Cloning of some heat shock proteins genes for further transcriptional study of Planktothrix agardhii exposed to abiotic stress.
Planktothrix agardhii is one of the freshwater cyanobacteria that can produce the hepatotoxin microcystins (MC)-a real threat to human and animal health. Knowledge of the biological role of MC in producing organisms is highly desired to understand the driving force of MC production. Recently, emerging evidences have suggested that MC may have protective role in cells facing environmental stress. If this is true, one should expect differences in the cellular protective mechanisms between MC-containing and MC-deficient mutant strains. To test this hypothesis, it would be essential to investigate the consequences of the loss of MC in Planktothrix in the transcriptional responses of its heat shock proteins (Hsps) to abiotic stresses-an important component of cellular stress response. However, a crucial first step is prerequisite for the isolation of hsp genes here, as the genome of Planktothrix has not been fully published. Therefore, we have successfully isolated four hsp genes including clpC (hsp100), htpG (hsp90), groEL (hsp60), and groES (hsp10) from Planktothrix agardhii PCC 7805 using ramped annealing PCR (RAN-PCR) with consensus-degenerate hybrid oligonucleotide primers (CODEHOP) and annealing control primer (ACP) system. In addition, some putative regulatory sequences found in the upstream region of groESL operon of Planktothrix agardhii were also discussed.


[Studies on kinetic viscoelasticity of slow muscle fibers--2. Dynamic stiffness changes studied by quick release method and frequency response method].
To evaluate the mechanical properties of slow fibers and fast fibers which make up the extraocular muscles in rabbits. I studied the contractile properties and viscoelastic properties of the superior rectus muscle(SR) and the retractor bulbi muscle (RB) of rabbits using the quick release method and the frequency response method. In the quick release method, isometric tension transients were slower in the SR than in the RB. In the frequency response method, the muscle length is perturbed sinusoidally to measure kinetic viscoelasticity. In resting muscle, dynamic stiffness was not changed with increasing frequency. In contracted muscle, dynamic stiffness was increased with increasing frequency, and the rate of increase was greater in the SR than in the RB. It was concluded that the viscoelasticity of the activated cross bridge is greater in slow fibers than in fast fibers.


Low-dose oral use of human interferon-alpha in cancer patients.
In a double-blind placebo-controlled trial, 57 adult subjects with disseminated malignancies were given orally low doses of recombinant human interferon-alpha (rHuIFN-alpha) at 0.05 IU, 0.5 IU, or 5.0 IU/kg body weight. The objective was to determine the efficacy of orally administered rHuIFN-alpha on appetite stimulation and/or weight loss prevention in anorectic cancer patients. Almost two-thirds (64%) of the subjects given 5.0 IU/kg reported an increase in appetite or body weight after 5 weeks in contrast to only 29% of the placebo-treated subjects. However, at the end of the 91-day trial, no significant differences in appetite enhancement or weight gain were noted between these two groups. Additionally, the 5.0 IU/kg treated group experienced half as many deaths as the control group by the conclusion of this 91-day trial.


[The gas chromatographic analysis of the fatty acids in the expired air in ischemic heart dis].
Lipids of exhaled air condensate (EAC) and sweat are analyzed by gas chromatography in coronary patients with myocardial infarction complicated by lung edema. Changes of lipid fatty acid spectrum in EAC and sweat were uniform: the levels of linoleic and arachidonic acids were increased, which can aggravate the disease and lead to complications.


Enhancement of rat hepatocellular-altered foci by the liver tumor promoter phenobarbital: evidence that foci are precursors of neoplasms and that the promoter acts on carcinogen-induced lesions.
Altered foci resistant to iron accumulation were induced by N-2-fluorenylacetamide in livers made siderotic by feeding 8-hydroxyquinoline and ferrous gluconate to inbred F344 rats. Following cessation of carcinogen feeding, most foci reverted to iron accumulation and could no longer be detected by 24 weeks, but some persisted for this interval. Futhermore, in 2 groups, 33 and 50% of rats developed liver tumors by 24 weeks after removal of the carcinogen. The addition of phenobarbital to the diet after cessation of carcinogen feeding enhanced persistence of altered foci and increased the incidence of liver tumors to 78--89%.


Cardiac rehabilitation services in low- and middle-income countries: a scoping review.
Despite the decreasing rate of cardiovascular disease-related mortality in developed nations, low- and middle-income countries (LMICs) are experiencing an increase. Cardiac rehabilitation (CR) successfully addresses this burden; however, the availability and nature of CR service delivery in LMICs are not well known. This scoping review examined the (1) presence and accessibility of CR services, (2) structure of CR services, and (3) effects of CR on patient outcomes in LMICs. Search criteria consisted of (1) nations considered to be low- or middle-income according to World Bank criteria, (2) CR, defined as programs including exercise and education, and (3) adults with cardiovascular diseases. Literature was identified through searching (a) the MEDLINE and EMBASE electronic databases, (b) proceedings from international cardiac conferences, (c) the grey literature and (d) through consulting experts in the field. Thirty peer-reviewed publications were identified. Grey literature, including Web sites for individual CR programs, revealed that CR is available in 32 (22.1%) LMICs. The most comprehensive data on accessibility stem from Latin America and the Caribbean, where 56% of institutions with cardiac catheterization facilities offered CR. Literature showed that some programs offered exercise, dietary advice, education, and psychological support, to assist patients to resume work and other activities of daily living. Fifteen peer-reviewed studies reported on CR outcomes, most of which were positive. Although patients similarly benefit from CR, few programs are available in LMICs. Policies need to be implemented to increase provision of tailored CR models at the global and national level, with evaluation.


Acute respiratory failure due to "pulmonary calciphylaxis" in a maintenance haemodialysis patient.
Calciphylaxis is a rapidly developing, fatal process of vascular calcium deposition with prominent cutaneous manifestation. We treated a long-term haemodialysis patient who developed an analogous disorder limited to the lungs. A 57-year-old man was admitted for initiation of peritoneal dialysis because limited cardiac reserve precluded further haemodialysis. He was treated successfully for pneumonia until hypoxia and progressive hypercalcaemia developed. (99m)Tc-methylene disphosphonate scintigraphy showed diffusely increased pulmonary uptake. Death supervened despite aggressive and successful treatment of hypercalcaemia. Autopsy studies included immunohistochemistry and morphometric studies of bone. Alveolar capillary walls showed diffuse calcium deposition. Both gross and microscopical findings differed from those of typical metastatic calcification in dialysis patients. Immunoreactivity for parathyroid hormone-related protein was present in the lesions. Bone histomorphometry indicated mild osteitis fibrosa. Pneumonia is believed to have caused local synthesis of parathyroid hormone-related protein that, along with high calcium x phosphorus product, contributed to calcium deposition. By analogy with the cutaneous process we termed the deposition "pulmonary calciphylaxis".


Comparative hardness and modulus of tooth-colored restoratives: a depth-sensing microindentation study.
The objective of this study was to compare the hardness and modulus of the continuum of direct tooth-colored restorative materials using a depth-sensing microindentation approach. The effects of thermal fatigue on mechanical properties were also evaluated. Six restorative materials representing the continuum were selected. They included an ormocer (Admira [AM], Voco), a giomer (Beautifil [BF], Shofu), a compomer (Dyract Extra [DE], Dentsply), a minifill composite (Esthet-X [EX], Dentsply), resin-modified (Fuji II LC [FL], GC) and highly viscous (Fuji IX [FN], GC) glass ionomer cements (GICs). Fourteen specimens (3 mm wide x 3 mm long x 2 mm deep) were made for each material. The specimens were randomly divided into two groups and treated as follows: Group A--stored in distilled water at 37 degrees C for 30 days; Group B--thermal cycled for 5000 cycles (35 degrees C [28s], 15 degrees C [2s], 35 degrees C [28s], 45 degrees C [2s]) and stored for 26.5 days. Hardness and modulus of the materials were determined using depth-sensing microindentation testing with the Instron MicroTester. Hardness was computed by dividing the peak load over the maximum projected contact area while modulus was calculated by analysis of the loading/unloading load-displacement (P-h) curves and the analytical model according to Oliver and Pharr (J. Mater. Res. 7 (1992) 1564). Results were analyzed using ANOVA/Scheffe's post hoc test and independent samples T-test (p<0.05). Hardness ranged 46.44-72.65 and 49.11-78.97 HV, while modulus ranged 7.86-12.78 and 8.12-13.13 GPa for Groups A and B, respectively. Although the ranking of mechanical properties were generally similar for both groups, disparities in statistical differences between materials were observed between Groups A and B for both hardness and modulus. For both groups, BF was significantly harder than DE, AM, FL and EX was significantly harder than FL. The modulus of FN was significantly greater than EX, DE, AM and FL was significantly stiffer than AM. With the exception of BF, no significant change in hardness and modulus was observed for all materials with thermocycling. The hardness and modulus of some glass ionomer-based/containing materials may be comparable or even superior to minifill and ormocer composites. Thermal fatigue should be considered when comparing mechanical properties between materials.


MicroRNAs relate to early worsening of renal function in patients with acute heart failure.
Deregulation of microRNAs (miRNAs) may be involved in the pathogenesis of heart failure (HF) and renal disease. Our aim is to describe miRNA levels related to early worsening renal function in acute HF patients. We studied the association between 12 circulating miRNAs and Worsening Renal Function (WRF; defined as an increase in the serum creatinine level of 0.3mg per deciliter or more from admission to day 3), absolute change in creatinine and Neutrophil Gelatinase Associated Lipocalin (NGAL) from admission to day 3 in 98 patients hospitalized for acute HF. At baseline, circulating levels of all miRNAs were lower in patients with WRF, with statistically significant decreased levels of miR-199a-3p, miR-423-3p, and miR-let-7i-5p (p-value<0.05). The increase in creatinine during the first 3 days of hospitalization was significantly associated with lower levels of miR-199a-3p, miR-27a-3p, miR-652-3p, miR-423-5p, and miR-let-7i-5p, while the increase in NGAL was significantly associated with lower levels of miR-18a-5p, miR-106a-5p, miR-223-3p, miR-199a-3p and miR-423-3p. MiR-199a-3p was the strongest predictor of WRF, with an Odds Ratio of 1.48 (1.061-2.065; p-value=0.021) and a C-index of 0.701. Our results show that the levels of circulating miRNAs at hospital admission for acute HF were consistently lower in patients who developed worsening of renal function. MiR-199a-3p was the best predictor of WRF in these patients.


What happened to Eric? The derailment of sexual development.
A variety of theoretical approaches, from the individual to ecological levels, was applied to the following clinical case in order to understand why a five-year-old boy demonstrated sexual behavior. Inappropriate sexual behavior in children is presented as symptomatic of problems in intrapersonal and interpersonal boundaries, relationship capabilities, and superego consolidation. The case material emphasizes that only when the child is helped to stop his sexual acting out can he access the sources of his emotional distress. The theoretical underpinnings that are relevant to this case will be shown as instrumental in the therapy of this child and his family. By integrating theory with practice, we hope to highlight this sensitive and prevalent issue as well as aid in the early identification and treatment of sexually problematic behavior.


Resting state networks in human cervical spinal cord observed with fMRI.
It has been reported that spontaneous fluctuations of blood oxygen level dependent (BOLD) signals can be detected in human brain and constitute resting state networks. It has not been reported whether resting state networks also exist in human spinal cord. In the present study, we investigate spontaneous BOLD signal changes in human cervical spinal cord during resting state. fMRI data were analyzed with independent component analysis and SPM software package. Acceptable reproducibility of spatial maps of BOLD signal changes was found across sessions, with the highest correlation values ranging from 0.18 to 0.44. The dominant frequency of signal changes from independent components with the highest correlation values was approximately the frequency range of the respiratory circle. Activities of spinal motor neurons innervating the scalenes were considered as a major factor in the production of BOLD signal fluctuations were observed in this study. Our findings suggest that BOLD fMRI can be applied to study the features of low-frequency rhythmic activities and corresponding mechanisms in the spinal cord during resting state.


Peutz-Jeghers syndrome-associated atypical mucinous proliferation of the uterine cervix: a case of minimal deviation adenocarcinoma ('adenoma malignum') in situ.
We describe a case of a non-invasive precursor of minimal deviation adenocarcinoma (MDA) of the uterine cervix, associated with Peutz-Jeghers syndrome (PJS). A 27-year-old woman, who had been followed for PJS, was referred to the gynecology clinic. Colposcopic examination demonstrated a small polypoid lesion in the transformation zone. Microscopic examination of the biopsy specimen demonstrated papillary proliferation of the mucinous epithelium with bland nuclear morphology. Conization revealed lobular endocervical glandular hyperplasia (LEGH) with distinct nuclear anaplasia, as well as papillary proliferation of the mucinous epithelium with mild to moderate nuclear abnormalities. This case suggests that the incipient phase of PJS-associated MDA is related to atypical LEGH ("MDA in situ"), and indicates the importance of early screening and surveillance by gynecologists in cases of PJS to detect cervical adenocarcinomas.


[Antibacterial activity of ceftriaxone].
In this study the author reports the main in vitro characteristics of ceftriaxone, a new aminothiazolyl methoxy-iminocephalosporin. The relationship between its structure and the antibacterial activity compared with cefotaxime reveals an expected longer antibacterial efficacy due to a long elimination half-life (8 h). It has been shown that ceftriaxone is highly stable against most either plasmid or chromosome-mediated beta-lactamases. Several studies demonstrated that the spectrum of ceftriaxone included Gram-positive cocci as well as most Gram-negative bacilli even beta-lactamase producers; a variable in vitro activity against Gram-negative aerobic bacteria (Pseudomonas sp., Acinetobacter sp.) has been proved, with a frequent synergistic bactericidal activity when combined with aminoglycosides, or with other beta-lactam antibiotics (carbenicillin, piperacillin). Multiple PBPs targets have been identified, including PBP1b, 2 and 3, leading to filamentation of E. coli, Ps. aeruginosa and Haemophilus cells. As a result, ceftriaxone displays a high intrinsic in vitro activity against troublesome strains and might be considered as a promising new cephalosporin for treatment of severe infections.


Educational interventions by pharmacists to patients with chronic pain: systematic review and meta-analysis.
We hypothesized that educational interventions delivered by pharmacists to patients with chronic pain might improve pain-related outcomes and sought to establish "proof of concept" for this hypothesis. We searched electronic databases and published literature for randomized studies that examined an educational intervention in relation to the management of chronic pain that was delivered by a pharmacist to an adult patient. Four studies were included that randomized 400 patients with chronic pain and which followed up patients between 1 and 16 weeks. Patients receiving these interventions experienced statistically significant benefits in the following outcomes compared with controls: a reduction in average pain intensity of 0.5 on a 0 to 10 rating scale, a reduction in adverse effects by more than 50%, and an improvement in satisfaction with treatment equivalent to approximately 1 point on a 0 to 10 rating scale. The interventions neither had effect on reducing interference from pain on daily life, nor on improving self-efficacy. Pharmacist-delivered educational interventions seem to reduce adverse events and improve satisfaction, but their clinical benefit on pain intensity is debatable. Our analysis suggests that the role of pharmacists may be important but a deeper understanding and evaluation of the active components of these interventions is needed within clinical trials before wider implementation into clinical practice can be recommended.


Computer analysis of manometric recordings. A study of overnight rectal activity in normal children.
To read and analyze manometric tracings from selected biological systems, a computer program was produced using low cost personal computer hardware. The program contains facilities for automatic scoring of contractions, intercontractile intervals (modality) and frequency analysis. This report concerns analysis of two overnight rectal motility recordings in 11 normal teenagers. After A/D conversion of data the developed software offers a set of options i.e. automatized scoring of contraction, detection of intervals between contractions and analytical tools such as histographic presentation of data, and presentation in the time domain. The A/D conversion time and computer analysis time for an 8 hour recording was 15 and 13 minutes, respectively. The analysis of frequency distribution overnight showed for the material as a whole 63% of no activity, 33% of 0-2 contractions x min-1 and 4% of more than 2 contractions x min-1. Modality analysis showed a peak between 15 and 25 seconds. Approximately 30% of the tracings showed accumulated intervals, which were multiples of the peak. No significant correlation was found between the first and second night of observation regarding the number of contractions and maximum frequency overnight. Significant correlation was found for the number of periods with high frequency activity between the two nights (p less than 0.05). Long-term pressure activity can be analyzed using the presented computer-aided method, providing a rapid and objective measurement of classical parameters and access to more in-depth analysis. This actual study of normal overnight rectal activity showed a large inter- and intra-individual variation.


Tumour necrosis factor-alpha, interleukin-1 beta and interleukin-6 in patients with renal cell carcinoma.
Patients with renal cell carcinoma (RCC) can exhibit fever, weight loss and increases in acute phase proteins. Interleukin (IL)-1, tumour necrosis factor (TNF) and IL-6 are considered major mediators of local and systemic inflammation. We measured plasma IL-1 beta, TNF-alpha (immunoradiometric assay) and IL-6 (ELISA) in 78 consecutive patients with untreated RCC and in 56 normal subjects. IL-6 plasma levels were higher in patients with RCC (mean 24.2 pg/ml, 11.1-37.3, 95% confidence interval) than in normal subjects (11.6 pg/ml, 10.1-13.1, n = 39, P < 0.01). The patients with fever or weight loss had higher blood levels of IL-6. IL-6 blood levels were also higher in patients with lymph node invasion and/or distant metastases (94.7 pg/ml, 39.0-150.4, n = 15) than in patients with undisseminated RCC (7.4, 4.1-10.7, n = 63, P < 0.0001). An abnormal IL-6 plasma value (> 40 pg/ml) had a positive predictive value of 91.0% for lymph node and/or metastatic spread of RCC. IL-6 was statistically correlated with C-reactive protein (nephelometric assay) blood values r' = 0.67, n = 78, P < 0.001). The TNF-alpha and IL-1 beta levels were not significantly different in patients with or without fever or weight loss. The plasma levels of the three cytokines were not correlated with the size of the primary tumour. An increased plasma value of IL-6 is a good marker for tumour dissemination in patients with untreated RCC.


Patient satisfaction in the long-term effects of Eustachian tube balloon dilation is encouraging.
To investigate the long-term effects of balloon Eustachian tuboplasty (BET) from patient's perspective and to discover which symptoms of Eustachian tube dysfunction (ETD) benefit the most from BET. We designed a retrospective postal questionnaire based on the seven-item ETD questionnaire (ETDQ-7). Our questionnaire covered the severity of present ETD symptoms in comparison with the preoperative situation, the severity of current overall ear symptoms, and possible surgical interventions after BET. Forty-six patients treated in our institution between 2011 and 2013 fulfilled the inclusion criteria and 74% (34 patients; total 52 ears treated with BET) returned the questionnaire with a mean follow-up time of 3.1 years (range 1.8-4.6 years). Pain in the ears, feeling of pressure in the ears, and feeling that ears are clogged had reduced in 75% of the ears that suffered from these symptoms preoperatively. Seventy-seven percent of all the responders felt that their overall ear symptoms were reduced. Altogether, 82% of all the patients stated that they would undergo BET again if their ear symptoms returned to the preoperative level. Patient satisfaction in the long-term effects of BET is encouraging. These results may help clinicians in preoperative patient selection and counselling.


Characterization of voltage-dependent calcium channel blocking peptides from the venom of the tarantula Grammostola rosea.
Voltage-dependent calcium channel blocking peptides were purified and sequenced from the venom of the tarantula, Grammostola rosea. cDNAs encoding the peptide sequences were cloned from the venom gland cDNA library. The electrophysiological effects of the peptides on several types of voltage-dependent calcium channels were evaluated using a Xenopus laevis oocyte expression system. A peptide contained in one of the HPLC peak fractions inhibited P/Q type voltage-dependent calcium channels (Ca(v)2.1). The amino acid sequence of this peptide is identical to that of ω-grammotoxin SIA. A peptide from another discrete peak, which is identical to GsAFII except for one tryptophan residue in the C-terminus, inhibited L-type voltage-dependent calcium channels (Ca(v)1.2). A novel peptide, named GTx1-15 (Accession number, AB201016), shows 76.5% sequence homology with the sodium channel blocker phrixotoxin 3, however, GTx1-15 preferentially inhibited T-type voltage-dependent calcium channels (Ca(v)3.1). In silico secondary and tertiary structure prediction revealed that GTx1-15 and sodium channel blockers such as hainantoxin-IV, phrixotoxin 3, and ceratotoxin 2 show very similar β-strand composition, distribution of Optimal Docking Areas (continuous surface patches likely to be involved in protein-protein interactions), and surface electrostatic potential. These findings suggest that these peptide toxins evolved from common ancestors by gene duplication to maintain surface atmospheres appropriate for interaction with low-voltage-dependent ion channels.


A new class of blockers of the voltage-gated potassium channel Kv1.3 via modification of the 4- or 7-position of khellinone.
The voltage-gated potassium channel Kv1.3 constitutes an attractive target for the selective suppression of effector memory T cells in autoimmune diseases. We have previously reported the natural product khellinone, 1a, as a versatile lead molecule and identified two new classes of Kv1.3 blockers: (i) chalcone derivatives of khellinone, and (ii) khellinone dimers linked through the 6-position. Here we describe the multiple parallel synthesis of a new class of khellinone derivatives selectively alkylated at either the 4- or 7-position via the phenolic OH and show that several chloro, bromo, methoxy, and nitro substituted benzyl derivatives inhibit Kv1.3 with submicromolar potencies. Representative examples of the most potent compounds from each subclass, 11m (5-acetyl-4-(4'-chloro)benzyloxy-6-hydroxy-7-methoxybenzofuran) and 14m (5-acetyl-7-(4'-bromo)benzyloxy-6-hydroxy-4-methoxybenzofuran), block Kv1.3 with EC50 values of 480 and 400 nM, respectively. Both compounds exhibit moderate selectivity over other Kv1-family channels and HERG, are not cytotoxic, and suppress human T cell proliferation at low micromolar concentrations.


Examining spatially varying relationships between land use and water quality using geographically weighted regression I: model design and evaluation.
Traditional regression techniques such as ordinary least squares (OLS) can hide important local variations in the model parameters, and are not able to deal with spatial autocorrelations existing in the variables. A recently developed technique, geographically weighted regression (GWR), is used to examine the relationships between land use and water quality in eastern Massachusetts, USA. GWR models make great improvements of model performance over OLS models, which is proved by F-test and comparisons of model R2 and corrected Akaike Information Criterion (AICc) from both GWR and OLS. GWR models also improve the reliabilities of the relationships by reducing spatial autocorrelations. The application of GWR models finds that the relationships between land use and water quality are not constant over space but show great spatial non-stationarity. GWR models are able to reveal the information previously ignored by OLS models on the local causes of water pollution, and so improve the model ability to explain local situation of water quality. The results of this study suggest that GWR technique has the potential to serve as a useful tool for environmental research and management at watershed, regional, national and even global scales.


Local recurrence and progression of non-muscle-invasive bladder cancer in Sweden: a population-based follow-up study.
The aim of this study was to investigate recurrence and progression of non-muscle-invasive bladder cancer (NMIBC) in a large population-based setting. Patients with bladder cancer (stage Ta, T1 or carcinoma in situ) diagnosed in 2004-2007 (n = 5839) in Sweden were investigated 5 years after diagnosis using a questionnaire. Differences in time to recurrence and progression were analysed in relation to age, gender, tumour stage and grade, intravesical treatment, healthcare region, and hospital volume of NMIBC patients (stratified in three equally large groups). Local bladder recurrence and progression occurred in 50 and 9% of the patients, respectively. The rate of local recurrence was 56% in the southern healthcare region compared to 37% in the northern region. A multivariate Cox proportional hazards model, adjusting for age, gender, tumour stage and grade, intravesical treatment, healthcare region and hospital volume, showed that recurrence was associated with TaG2 and T1 disease, no intravesical treatment and treatment in the southern healthcare region, but indicated a lower risk of recurrence in the northern healthcare region. Adjusting for the same factors in a multivariate analysis suggested that increased relative risk of progression correlated with older age, higher tumour stage and grade, and diagnosis in the Uppsala/Örebro healthcare region, whereas such risk was decreased by intravesical treatment (relative risk 0.72, 95% confidence interval 0.55-0.93, p = 0.012). The incidence of NMIBC recurrence and progression was found to be high in Sweden, and important disparities in outcome related to care patterns appear to exist between different healthcare regions.


Species- and sex-specific renal cytotoxicity of ochratoxin A and B in vitro.
Four different cell models were chosen for comparison of OTA and OTB toxicity: primary porcine (PKC), rat (RPTC) and human renal proximal epithelial cells (HKC) from both sexes and a porcine renal cell line: LLC-PK1. Culture conditions were tested and optimized for each respective cell type (species/sex and origin). All cell types were characterized for epithelial origin and growth patterns and following optimization of dosing strategies and assay procedures, a strict study design was implemented to avoid systemic variations. Due to possible sensitivity differences, three simple endpoints were chosen to provide basic data for interspecies comparison: neutral red uptake, MTT reduction and cell number. Of the endpoints tested neutral red appeared the most sensitive, although all three parameters yielded comparable EC50's. Sex-differences were observed between male and female HKC cells following 96 h exposure to OTA, with HKC(m) being more sensitive than HKC(f). No sex-difference was observed in PKC cells, however, the PKC were approximately 3 and 10 times more sensitive than HKC(m) and HKC(f), respectively, to OTA and OTB. Interestingly, the CI95 of the EC50 values obtained for OTA (15.5-16.5 microM) and OTB (17.0-2 1.0 microM) were comparable in the PKC cells. In contrast, OTB had lower cytotoxicity than OTA in HKC and LLC-PK1 (approx. 2-fold) and no effects in RPTC. Overall, HKC(m) were nearly as sensitive as PKC towards OTA, followed by RPTC, LLC-PK1 and HKC(f), thus suggesting a sex specific sensitivity in humans towards OTA induced cytotoxicity.


All-trans retinoic acid ameliorates inflammatory response mediated by TLR4/NF-κB during initiation of diabetic nephropathy.
Diabetic nephropathy (DN) is the leading cause of renal failure worldwide and its complications have become a public health problem. Inflammation, oxidative stress and fibrosis play central roles in the progression of DN that lead to renal failure. Potential deleterious effect of inflammation in early evolution of DN is not fully disclosed. Therefore, it is relevant to explore therapies that might modulate this process in order to reduce DN progression. We explored the beneficial effect of all-trans retinoic acid (ATRA) in early inflammation in glomeruli, proximal and distal tubules in streptozotocin (STZ)-induced diabetes. ATRA was administered (1 mg/kg daily by gavage) on days 3 to 21 after STZ administration. It was found that 21 days after STZ injection, diabetic rats exhibited proteinuria, increased natriuresis and loss of body weight. Besides, diabetes induced an increase in interleukins [IL-1β, IL-1α, IL-16, IL-13, IL-2; tumor necrosis factor alpha (TNF-α)] and transforming growth factor-beta 1 (TGF-β1), chemokines (CCL2, CCL20, CXCL5 and CXCL7), adhesion molecules (ICAM-1 and L-selectin) and growth factors (GM-CSF, VEGF, PDGF) in glomeruli and proximal tubules, whereas ATRA treatment remarkably ameliorated these alterations. To further explore the mechanisms through which ATRA decreased inflammatory response, the NF-κB/p65 signaling mediated by TLR4 was studied. We found that ATRA administration attenuates the TLR4/NF-κB inflammatory signaling and prevents NF-κB nuclear translocation in glomeruli and proximal tubules.


Neurologic defects and selective disruption of basement membranes in mice lacking entactin-1/nidogen-1.
Entactin-1 (nidogen-1) is an ubiquitous component of basement membranes. From in vitro experiments, entactin-1 was assigned a role in maintaining the structural integrity of the basement membrane because of its binding affinity to other components, such as type IV collagen and laminin. Entactin-1 also interacts with integrin receptors on the cell surface to mediate cell adhesion, spreading, and motility. Targeted disruption of the entactin-1 gene in the mouse presented in this study revealed a duplication of the entacin-1 locus. Homozygous mutants for the functional locus lacked entactin-1 mRNA and protein and often displayed seizure-like symptoms and loss of muscle control in the hind legs. The behavior patterns suggested the presence of neurologic deficits in the central nervous system, thus providing genetic evidence linking entactin-1 to proper functions of the neuromuscular system. In homozygous mutants, structural alterations in the basement membranes were found only in selected locations including brain capillaries and the lens capsule. The morphology of the basement membranes in other tissues examined superficially appeared to be normal. These observations suggest that the lost functions of entactin-1 result in pathologic changes that are highly tissue specific.


Experimental platform to study heavy metal ion-enzyme interactions and amperometric inhibitive assay of Ag+ based on solution state and immobilized glucose oxidase.
The heavy metal (HM) ion-enzyme interaction is an important research topic in many areas. Using glucose oxidase (GOx) as an example, a comprehensive experimental platform based on quartz crystal microbalance and electroanalysis techniques is developed here to quantitatively study the HM ion-enzyme interactions and amperometric inhibitive assays of HM ions. The effects of some common HM ions on the bioactivities of solution-state GOx (GOx(s)), electrode surface-adsorbed GOx (GOx(ads)), and polymer-entrapped GOx (GOx(e)) are comparatively examined on the basis of anodic amperometric detection of enzymatically generated H(2)O(2). Ag(+) shows the strongest inhibition effect among the HM ions examined, and the inhibitive assays of Ag(+) based on GOx(s), GOx(ads), and GOx(e) entrapped in poly(l-noradrenalin) (PNA) give limits of detection (LOD) of 2.0, 8.0, and 5.0 nM (S/N = 3), respectively. Inhibition effects of Hg(2+), Cu(2+), and Co(2+) are detectable only at 15 μM or higher concentrations, and the other HM ions show undetectable inhibition even at 1.0 mM. The developed experimental platform allows one to quantify the number of the bound HM ions per GOx(ads) molecule at various inhibition percentages. In addition, the electrosynthesized PNA matrix to entrap GOx for an inhibitive assay of Ag(+) shows the lowest competitive affinity to HM ions and gives the highest sensitivity, as compared with several other polymer matrixes commonly used for the inhibitive assay. The suggested experimental platform is recommended for wide applications in enzymatic inhibitive assays and quantitative studies of the inhibition effects of HM ions on many other redox-event-relevant enzymes.


Determination of heme and non-heme iron content of mouse erythropoietic organs.
A procedure is described for determining the content of heme and non-heme iron in organs and tissues of the mouse. Heme iron after homogenization and hemolysis of the samples is extracted as hemin by means of a mixture of ethyl acetate and glacial acetic acid, the extraction being followed by washing with hydrochloric acid. Total iron content is determined with commercial analytical sets exploiting color reaction with bathophenanthroline after mineralization of the samples. The presence of bone or other tissues does not distort the results. The procedure is relatively simple and suitable for serial analyses. The results obtained for the mouse by this method are in good agreement with recognized principles of ferrokinetics in mammals.


A comparison of the relaxant effects of olprinone and aminophylline on methacholine-induced bronchoconstriction in dogs.
IV aminophylline, a nonselective phosphodiesterase (PDE) inhibitor, is often used to treat an asthma attack during anesthesia. However, in some instances, aminophylline-resistant attacks are observed. Selective PDE3 inhibitors are now clinically available and have been reported to produce bronchodilation. Thus, we compared the relaxant effects of olprinone, a novel PDE3 inhibitor, and aminophylline on methacholine-induced bronchoconstriction. Dogs were anesthetized with pentobarbital. Bronchoconstriction was elicited with methacholine (0.5 microg/kg + 5.0 microg. kg(-1). min(-1)) and assessed as percentage of changes in the bronchial cross-sectional area (BCA; basal = 100%) monitored by bronchoscope. Initially, the relaxant effects of olprinone (n = 8; 0-1000 microg/kg) and aminophylline (n = 8; 0-50 mg/kg) were compared. The bronchial cross-sectional areas were assessed before and 30 min after methacholine infusion began and 5 min after each dose of olprinone or aminophylline. We then determined whether propranolol (0.4 mg/kg) reversed the relaxation induced by olprinone (1000 microg/kg) and aminophylline (50 mg/kg). Olprinone and aminophylline dose-dependently antagonized bronchoconstriction by 56.2% +/- 21.3% (SD) and 68.0% +/- 30.3% with -log 50% effective dose (mean) of 4.80 +/- 0.38 (15.8) microg/kg and 1.96 +/- 0.42 (10.9) mg/kg, respectively. Aminophylline 50 mg/kg significantly increased plasma epinephrine, whereas olprinone did not. In addition, propranolol significantly reduced aminophylline-induced relaxation, but not olprinone-induced relaxation. Therefore, the relaxant effects of olprinone are independent of plasma epinephrine, whereas aminophylline effects may partially result from increased circulating concentrations of epinephrine. We compared the relaxant effects of olprinone and aminophylline on methacholine-induced bronchoconstriction in dogs. The relaxant effects of olprinone are independent of plasma epinephrine, whereas the aminophylline effects may be partly caused by an increase in plasma epinephrine.


Neuroectodermal tumors expressing c-, L-, and N-myc in transgenic mice that carry the E1A/E1B gene of human adenovirus type 12.
Adenovirus early 1 (E1) region gene products, including E1A and E1B, are required for transformation of primary cultured rodent cells. In order to investigate in vivo action of the E1 region, we established a line of transgenic mice carrying the oncogenic E1A and E1B genes of human adenovirus type 12 under control of the human angiotensinogen promoter. Histopathological analyses indicated that transgenic mice heritably develop neuroectodermal tumors arising from the pelvic region with varying degrees of incidence. The transgene was expressed in the neuroectodermal tumors as well as in TNT-1 cells, a cell line established from the tumors, where the human angiotensinogen promoter was constitutively active. The high level expression of c-, L-, and N-myc without gene amplification was notable in the original tumors and TNT-1 cells, but not in another tissue examined. The co-expression of the three sets of myc family genes in both the original tumors and the established cell line provided the possibility that the target cells for transformation may belong to a specific cell type that expresses all these oncogenes during development.


Glycogen synthase kinase-3β opens mitochondrial permeability transition pore through mitochondrial hexokinase II dissociation.
Accumulating evidence has revealed pivotal roles of glycogen synthase kinase-3β (GSK3β) inactivation on cardiac protection. Because the precise mechanisms of cardiac protection against ischemia/reperfusion (I/R) injury by GSK3β-inactivation remain elusive, we investigated the relationship between GSK3β-mediated mitochondrial hexokinase II (mitoHK-II; a downstream target of GSK3β) dissociation and mitochondrial permeability transition pore (mPTP) opening. In Langendorff-perfused hearts, GSK3β inactivation by SB216763 improved the left ventricular-developed pressure and retained mitoHK-II binding after I/R. In permeabilized myocytes, GSK3β depolarized mitochondrial membrane potential with accelerated mitochondrial calcein release (suggesting GSK3β-mediated mPTP opening) and decreased mitoHK-II bindings. GSK3β-mediated mPTP opening depended on mitoHK-II binding, i.e., it was accelerated by dissociation of mitoHK-II (dicyclohexylcarbodiimide) and attenuated by enhancement of mitoHK-II binding (dextran). However, inactivation of mitoHK-II by glucose-depletion or glucose-6-phosphate inhibited the GSK3β-mediated mPTP opening. We conclude that GSK3β-mediated mPTP opening may be involved in I/R injury and regulated by mitoHK-II binding and activity.


Evaluating countermeasures in spaceflight analogs.
Countermeasures are defined as solutions to prevent the undesirable physiologic outcomes associated with spaceflight. Spaceflight analogs provide a valuable opportunity for the evaluation of countermeasures because they allow for the evaluation of more subjects, more experimental control, and are considerably less expensive than actual spaceflight. The various human analogs have differing strengths and weaknesses with respect to the development and evaluation of countermeasures. The human analogs are briefly reviewed with a focus on their suitability for countermeasure evaluation. Bed rest is the most commonly used analog for evaluating countermeasures. While countermeasures are typically developed to target one or maybe two particular physiologic issues, it is increasingly important to evaluate all of the organ systems to discern whether they might be unintended consequences on nontargeted tissues. In preparation for Mars exploration it will be necessary to fully integrate countermeasures to protect all organ systems. The synergistic and antagonistic effects of multiple countermeasures needs to be the focus of future work.


[Relative non curative resection of gastric cancer].
Relative non curative resection of gastric cancer was studied according to The General Rules of Japanese Research Society for gastric cancer. The five year survival rate was 31.2%, while 39.6% for relative curative resection and 4.9% for absolute non curative resection. The cases whose relative non curative factor was due to lymph nodes (n greater than R) resulted in a fair prognosis, but the prognosis of those concerned with peritoneal dissemination (P1) or liver metastasis (H1) was very poor. Surgical achievement to improve postoperative results may be obtained when a systemic (13), (14v) (16) lymph node dissection is performed whenever only one of them was involved in metastasis.


Crystal structure, physical properties and HRTEM investigation of the new oxonitridosilicate EuSi2O2N2.
The new layered oxonitridosilicate EuSi(2)O(2)N(2) has been synthesized in a radio-frequency furnace at temperatures of about 1400 degrees C starting from europium(III) oxide (Eu(2)O(3)) and silicon diimide (Si(NH)(2)). The structure of the yellow material has been determined by single-crystal X-ray diffraction analysis (space group P1 (no. 1), a=709.5(1), b=724.6(1), c=725.6(1) pm, alpha=88.69(2), beta=84.77(2), gamma=75.84(2) degrees ,V=360.19(9)x10(6) pm(3), Z=4, R1=0.0631, 4551 independent reflections, 175 parameters). Its anionic Si(2)O(2)N(2) (2-) layers consist of corner-sharing SiON(3) tetrahedra with threefold connecting nitrogen and terminal oxygen atoms. High-resolution transmission electron micrographs indicate both ordered and disordered crystallites as well as twinning. Magnetic susceptibility measurements of EuSi(2)O(2)N(2) exhibit Curie-Weiss behavior above 20 K with an effective magnetic moment of 7.80(5) mu(B) Eu(-1), indicating divalent europium. Antiferromagnetic ordering is detected at 4.5(2) K. EuSi(2)O(2)N(2) shows a field-induced transition with a critical field of 0.50(5) T. The four crystallographically different europium sites cannot be distinguished by (151)Eu Mössbauer spectroscopy. The room-temperature spectrum is fitted by one signal at an isomer shift of delta=-12.3(1) mm s(-1) subject to quadrupole splitting of DeltaE(Q)=-2.3(1) mm s(-1) and an asymmetry parameter of 0.46(3). Luminescence measurements show a narrow emission band with regard to the four crystallographic europium sites with an emission maximum at lambda=575 nm.


Posterior glenoid bone grafting in total shoulder arthroplasty for osteoarthritis with severe posterior glenoid wear.
Total shoulder arthroplasty (TSA) in cases with posterior wear can be addressed by eccentric reaming of the anterior glenoid or by augmenting the posterior glenoid with bone grafting or augmented glenoid implants. We report the results of TSA with posterior glenoid bone grafting (PGBG) with humeral head autograft in patients with shoulder osteoarthritis and severe posterior glenoid wear. A retrospective review of cases from 2004 to 2014 revealed 34 patients. Preoperative and postoperative radiographs were evaluated for glenoid version and humeral head subluxation as well as component loosening. Patient-reported outcomes were compared preoperatively and postoperatively. Complications and reoperations were also evaluated. Of the 34 patients, 28 (82.4%) were available at a minimum of 2 years' follow-up. PGBG corrected glenoid retroversion from -28° ± 4° preoperatively to -4° ± 2° (P < .001). Humeral head subluxation also improved after PGBG with respect to the scapular axis and to the midglenoid face (P < .001). Radiographic analysis revealed all PGBGs had incorporated. Radiographically, 3 patients (10.7%) had a total of 5 broken or displaced screws. In addition, 3 patients (10.7%) had a broken metal marker in the center peg of the glenoid component. No patients required component revision surgery by final follow-up. Only 1 reoperation occurred for capsular release. Patients showed significant improvements in all patient-reported outcomes. Patients undergoing primary TSA with humeral head autograft PGBG showed significant improvements in glenoid version, humeral head subluxation, patient-reported outcomes, and range of motion at an average of 4 years' follow-up. There was a low revision rate and a high rate of graft incorporation.


Nongenomic actions of estrogens: exciting opportunities for pharmacology.
Estrogens, like other steroids, elicit a variety of rapid effects in many tissues in addition to their delayed action on gene expression in the cell nucleus. The rapid responses occur without participation of the genome, and are therefore termed nongenomic. Some of the estrogen induced effects acutely modulate vascular function and may contribute to the gender difference in cardiovascular susceptibility. While some actions may be mediated by novel, nonclassic receptors, the classic estrogen receptor has been shown to also act on signalling cascades. There are sparse examples for compounds structurally related to the endogenous hormone estradiol that bind to the estrogen receptor but may selectively elicit nongenomic responses. The further development of such selectively acting drugs holds much promise for better therapies with fewer side effects, e.g. for vascular malfunction, but also for estrogen-dependent cancer.


[Predictive value of CA 125 in detection of ovarian cancer in pre- and postmenopausal patients].
The aim of the study was to evaluate the usefulness of CA125 measurements in ovarian cancer diagnostics in pre- versus postmenopausal women. The study group consisted of 99 serum specimens of women diagnosed with ovarian cancer (28 pre- and 71 postmenopausal). The control group consisted of 86 specimens collected from women without any ovarian pathology (34 pre- and 52 postmenopausal). CA 125 measurements were performed with Zymed Laboratories CA 125 ELISA Kit. Descriptive statistics, including mean values, standard deviation (SD) and 95CI of CA 125 among pre- and postmenopausal women, were calculated. The usefulness of CA 125 measurements in ovarian cancer diagnostics was assessed by calculating specificity sensitivity positive and negative predictive rates among pre- versus postmenopausal women. Statistical calculation was preformed with the use of STATISTICA 6.0 StatSoft Inc. (2001) software (wwwstatsoft. com). Higher CA 125 concentrations were observed in controls among pre- versus postmenopausal women (10.01 +/- 15.41; 95% CI: 4.63-15.39 IU/ml vs. 8.88 +/- 13.74; 95% CI: 5.06-12.71/U/ml, respectively NS). In contrary among women with ovarian cancer CA 125 levels were higher in postmenopausal women when compared to premenopausal (352.11 +/- 432.07; 95% CI: 184.57-519.65 vs. 541.59 +/- 547.98 95% CI: 411.89-671.29, respectively p < 0.05). In premenopausal group the CA 125 sensitivity was calculated at 64% whereas specificity 94.12% (FPR 5.9%; OR: 10.9), positive predictive value 90% and negative predictive value 94.12%. Among postmenopausal women results were as follows: sensitivity 88.73%, specificity 98.07% (FPR 1.9%, OR: 46.7), positive predictive value 98.44% and negative predictive value 86.44%. CA 125 measurements were more useful in the postmenopausal group. Higher CA 125 levels were observed in women with ovarian cancer diagnosis when compared to controls, in which lower CA 125 were observed. Higher sensitivity specificity and positive predictive values were observed in the postmenopausal group.


A high-throughput colorimetric-based bioassay method for determination of fosmidomycin in plasma and urine and application for pharmacokinetic study.
Fosmidomycin is a phosphonic acid derivative originally isolated as a natural antibiotic from Streptomyces lavendulae. It was originally used as an antibiotic, and later on as an antimalarial drug. A simple, sensitive, selective and reproducible bioassay based on colorimetric method was developed for the determination of fosmidomycin in human plasma and urine. Enterobacter cloacae ATCC 23355 strain was used as a test organism. Inhibition of bacterial growth was assessed using MTT assay. Calibration curves were prepared from concentration response curves in plasma (0, 0.5, 1, 2.5, 5, 12.5, 25, 50 ng/microl) and urine (0, 0.5, 1, 2.5, 5, 12.5, 25, 50 and 100 ng/microl) and were all linear with correlation coefficients better than 0.990. The precision of the method based on within-day repeatability and reproducibility (day-to-day variation) was below 10% (% coefficient of variations: % C.V.) Good accuracy was observed for both the intra-day and inter-day assays, as indicated by the minimal deviation of mean values found between the measured samples and the theoretical values (below +/-10%). Limit of quantification (L.O.Q.) was accepted as 0.5 ng using 20 microl plasma or 10 microl urine sample. The mean recovery for fosmidomycin was greater than 99%. The method was free from interference from other commonly used antibiotics. The analytical method established in this study meets the criteria of high sensitivity, accuracy and reproducibility for routine use in pharmacokinetic studies. The method has advantages over the recently developed bioassay in its simplicity, accuracy, short analysis time, good sensitivity, and the requirement for smaller volumes of samples. The method appears to be robust and has been applied to a pharmacokinetic study to determine the plasma and urinary excretion of fosmidomycin in a total of 24 Thai patients with acute uncomplicated falciparum malaria following oral doses of 1800 fosmidomycin given every 10 h for 3 days.


Temporal trend studies on polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs) in ringed seals from east Greenland.
The concentrations of polybrominated diphenyl ethers (PBDEs) 17, 28, 47, 49, 66, 85, 99, 100, 153, 154, and 183 were determined in ringed seal blubber from central East Greenland collected in 1986, 1994, 1999 and during the period 2001 to 2004. The trend of PBDEs was compared with the trends of polychlorinated biphenyls (PCBs) 28, 31, 52, 101, 105, 118, 138, 153, 156, and 180 during the same period. The levels of sigmaPBDE in East Greenland ringed seals ranged from 21.8 ng g(-1) lipid weight (1w) in 1986 to 39.3 ng g(-1) lw in 2001 and are among the highest observed in ringed seal from the Arctic. The dominating congeners were BDE-47 (75.4%) and BDE-99 (9.7%). The concentrations of PBDEs and PCBs increased with the age of the seals, and therefore only young seals < or =4 years old) were included in the temporal trend analyses. No significant trend (p > 0.14) was observed in sigmaPBDE or the congeners BDE 28, 47 and, 99 during the period while sigmaPCB decreased significantly (p = 0.004) over the period from 1986 to 2004 with an estimated annual rate of 4.3%.


Complete inversion of the uterus during caesarean section: a case report.
A case of complete inversion of the uterus through the uterine incision at the time of Caesarean section is presented. The possible aetiological factors were a short cord and fundal insertion of the placenta. Disappearance of the lower segment incision and the raw appearance of the endometrial surface of a completely inverted uterus posed a diagnostic dilemma. Prompt understanding and immediate repositioning by a simple technique prevented any further complications. This rare complication of Caesarean section and its easy principles of management are described for the benefit of an unwary obstetrician.


Mitochondrial free radical overproduction due to respiratory chain impairment in the brain of a mouse model of Rett syndrome: protective effect of CNF1.
Rett syndrome (RTT) is a pervasive neurodevelopmental disorder mainly caused by mutations in the X-linked MECP2 gene associated with severe intellectual disability, movement disorders, and autistic-like behaviors. Its pathogenesis remains mostly not understood and no effective therapy is available. High circulating levels of oxidative stress markers in patients and the occurrence of oxidative brain damage in MeCP2-deficient mouse models suggest the involvement of oxidative stress in RTT pathogenesis. However, the molecular mechanism and the origin of the oxidative stress have not been elucidated. Here we demonstrate that a redox imbalance arises from aberrant mitochondrial functionality in the brain of MeCP2-308 heterozygous female mice, a condition that more closely recapitulates that of RTT patients. The marked increase in the rate of hydrogen peroxide generation in the brain of RTT mice seems mainly produced by the dysfunctional complex II of the mitochondrial respiratory chain. In addition, both membrane potential generation and mitochondrial ATP synthesis are decreased in RTT mouse brains when succinate, the complex II respiratory substrate, is used as an energy source. Respiratory chain impairment is brain area specific, owing to a decrease in either cAMP-dependent phosphorylation or protein levels of specific complex subunits. Further, we investigated whether the treatment of RTT mice with the bacterial protein CNF1, previously reported to ameliorate the neurobehavioral phenotype and brain bioenergetic markers in an RTT mouse model, exerts specific effects on brain mitochondrial function and consequently on hydrogen peroxide production. In RTT brains treated with CNF1, we observed the reactivation of respiratory chain complexes, the rescue of mitochondrial functionality, and the prevention of brain hydrogen peroxide overproduction. These results provide definitive evidence of mitochondrial reactive oxygen species overproduction in RTT mouse brain and highlight CNF1 efficacy in counteracting RTT-related mitochondrial defects.


Vitreous humour - routine or alternative material for analysis in forensic medicine.
Biological materials used in toxicological analyses in forensic medicine traditionally include blood, urine and vitreous humour. Forensic use of the vitreous body is mostly due to the need to assess the endogenous concentration of ethyl alcohol in the process of human body decomposition. The vitreous body is an underestimated biological material, even though its biochemical properties and anatomical location make it suitable for specific forensic toxicology tests as a reliable material for the preparation of forensic expert opinions. Based on the available literature the paper gathers information on the biochemical structure of the vitreous body, ways to secure the material after collection and its use in postmortem diagnostics. Specific applications of the vitreous humour for biochemical and toxicological tests are discussed, with a focus on its advantages and limitations in forensic medical assessment which are attributable to its biochemical properties, anatomical location and limited scientific studies on the distribution of xenobiotics in the vitreous body.


Review of PI3K/mTOR Inhibitors Entering Clinical Trials to Treat Triple Negative Breast Cancers.
Constitutive activation of the PI3K/mTOR signaling pathway is observed in most, if not all, breast cancers. Accordingly, many PI3K and/or mTOR inhibitors have entered clinical trials, and completed studies should soon reveal the efficacy of these new drug families in the treatment of cancer patients. We present the PI3K/Akt/mTOR signaling pathway and the structure and the anti-tumor efficiency of some mTOR inhibitors such as rapalogues and competitive inhibitors, which have entered clinical trials. We also discuss some of the clinical trial results associated with these molecules mainly focusing on studies performed on relapsing breast cancer patients - but not only. Most of the clinical trials with PI3K/mTOR inhibitors alone or in combination with chemotherapies were performed in heavily pre-treated patients and revealed non-negligible amounts of partial responses and long-term stable disease for these patients. Therefore, these compounds seem to prevent tumor growth and survival of cancer cells in Human, representing a new range of anti-tumor drugs that can be utilized not only as first-line treatments but as second- and third-line agents for patients who relapse. Drugs inhibiting the PI3K/mTOR signaling pathway may represent tailored anti-tumor agents, paving the way for their clinical application in different tumor types.


Responding to a murder/suicide at a rural junior high school.
After a murder/suicide at the Red Lion Junior High School in Red Lion, Pennsylvania, (the second critical incident in two years for this school district), our crisis response team provided critical incident stress management (CISM) services, response coordination and ongoing support to the students, teachers, parents and community of this small town. Our unique response team was a diverse blend of professionals from the school district, Pennsylvania State Education Association, school psychologists, and the local employee assistance program (EAP). Within eight days, the school population had moved beyond a horrifying critical incident to begin the process of recovery.


The most deviated codon position in AT-rich bacterial genomes: a function related analysis.
We have performed systematic study on more than 120 archaeal and bacterial genomes. Based on the index proposed in the current paper, clear patterns are observed showing the relation between the base compositional deviation at three codon positions and the genomic GC content. For AT-rich genomes, the Most Deviated Codon Position (MDCP) is the 1st codon position, while for GC-rich genomes, MDCP appears at the 2nd or 3rd codon position alternatively. According to MDCP, the CDSs of a genome can be classified into two types: typical and atypical. In AT-rich genomes the typical represent the majority and account for about 3/4 of all the CDSs. Based on the functional classification of COG database, the two types of CDSs are examined. An apparent bias of distribution is observed that the CDSs with the function of 'information processing' are more likely to present in typical type.


An in vitro release study of 5-fluoro-uracil (5-FU) from swellable poly-(2-hydroxyethyl methacrylate) (PHEMA) nanoparticles.
Nanomaterials are at the leading edge of the rapidly developing field of nanotechnology. The use of nanoparticles as drug delivery vehicles for anticancer therapeutics has great potential to revolutionize the future of cancer therapy. The present paper concerns both the optimizations of anticancer drug loading and its release from polymeric nanoparticles. The major aim of this study was to design poly (HEMA) nanoparticles as swelling controlled drug release system for anticancer drug. The prepared nanoparticles were characterized by Infra-Red (IR) Spectra, Particle size Analysis, and Scanning Electron Microscopy (SEM). The nanoparticles were loaded with widely used anticancer drug, 5-Fluorouracil, and controlled release of drug was investigated to observe the effects of various parameters such as percent loading of the drug, chemical architecture of the nanocarriers, pH, temperature, and nature of release media on the release profiles. The chemical stability of 5-Fluorouracil (5-FU) was also tested in phosphate buffer saline (PBS) (pH = 7.4) and release was studied in various simulated biological fluids. The prepared nanoparticles could provide a possible pathway for controlled and targeted delivery of anticancer drug, thus causing lower side effects and higher efficacy.


Commercial hospital discharge packs for breastfeeding women.
Exclusive breastfeeding until around six months of age, followed by the introduction of solids with continued breastfeeding, is considered to be the optimal nutritional start for newborn infants. To determine whether the exclusivity and duration of breastfeeding is affected by giving mothers commercial discharge packs in hospital which contain artificial formula or promotional material for artificial formula. These packs are those which are commonly given to mothers on leaving hospital after giving birth (thus discharge packs). Comprehensive electronic search of the register of clinical trials maintained and updated by the Cochrane Pregnancy and Childbirth Group and CINAHL and MEDLINE. All randomised controlled trials with or without blinding to examine the effects of commercial discharge packs on breastfeeding. Consenting postpartum women who initiate breastfeeding while in hospital or immediately upon discharge. Commercial discharge packs which contain free samples of infant formula or promotional material versus non commercial discharge packs (specifically those from which free samples of infant formula have been removed or have been replaced with e.g. breast pads) or no pack. The proportion of women breastfeeding at six weeks and 3 months (13 weeks) postpartum. Other outcomes: Rates of breastfeeding at other fixed time points between 0 and 6 months postpartum. Data were extracted by one reviewer and checked by a second reviewer. Nine randomised controlled trials involving a total of 3730 women were analysed. The studies only included women from North America. The meta-analysis showed that when comparing commercial discharge packs with any of the controls (no intervention, non-commercial pack and combinations of these), exclusive breastfeeding was reduced at all time points in the presence of commercial hospital discharge packs. There was no evidence to support the conjecture that use of hospital discharge packs causes the early termination of non-exclusive breastfeeding. Where the introduction of solid food was measured, giving a commercial pack (with or without formula) reduced the time before solid food was introduced. The giving of commercial hospital discharge packs (with or without formula) appears to reduce the number of women exclusively breastfeeding at all times but has no significant effect upon the earlier termination of non-exclusive breastfeeding.


Plaque-based competitive hybridization.
The authors have developed a simple, cost-saving experimental design, plaque-based competitive hybridization (PBCH), for genome-wide identification of genes differentially expressed in different tissues. PBCH offers advantages in comparison with other methods used in comparative genomics by combining the principles of differential hybridization with the subtractive hybridization. PBCH is particularly advantageous when libraries with few differences are to be analyzed. The authors demonstrate the use of PBCH by identifying 3 genes, up-regulated in the developing velvet antler of red deer (Cervus elaphus): ApoD, C011A2, and S100a1. The fidelity and sensitivity of PBCH is also shown: 1 specific clone among a library sample of 15,000 can be recognized. Possibilities for further utilizations are discussed.


Acute glycaemic load breakfast manipulations do not attenuate cognitive impairments in adults with type 2 diabetes.
Research on young healthy samples suggests that low glycaemic load foods can confer benefits for cognitive performance. The aim was to examine the effects of type 2 diabetes on cognitive function, and to investigate whether consumption of low glycaemic load breakfasts affects cognitive function in adults with type 2 diabetes. Memory, psychomotor skill and executive function were examined at two morning test sessions in 24 adults with type 2 diabetes and 10 adults with normal glucose tolerance (NGT) aged 45-77 years without dementia after water, low, and high glycaemic load breakfasts were consumed in accordance with a crossover, counterbalanced design. The type 2 diabetes and NGT groups were matched for education, depression, and IQ. Type 2 diabetes was associated with impairments in verbal memory, spatial memory, psychomotor skill, and executive function compared to adults with NGT. Consumption of the three breakfast conditions did not impact on cognitive performance in the type 2 diabetes or NGT participants. Abnormalities in glucose tolerance such as type 2 diabetes can have demonstrable negative effects on a range of cognitive functions. However, there was no evidence that low GL breakfasts administered acutely could confer benefits for cognitive function (ClincalTrials.gov identifier, NCT01047813).


The interleukin-1 system and female reproduction.
Interleukins (ILs) are known best for their involvement in the immune system and their role during inflammation. In the ovary, a growing body of evidence suggests that the ovarian follicle is a site of inflammatory reactions. Thus ovarian cells could represent sources and targets of ILs. Since then, the IL-1 system components (IL-1alpha, IL-1beta, IL-1 receptor antagonist, IL-1 receptors) have been demonstrated to have several sites of synthesis in the ovary. These factors have been localized in the various ovarian cell types, such as the oocyte, granulosa and theca cells, in several mammalian species. IL-1-like bioactivity has been reported in human and porcine follicular fluid at the time of ovulation. The role of IL-1 in local processes is still poorly known, although there is evidence for involvement in the ovulation process, and in oocyte maturation. More precisely, IL-1 may be involved in several ovulation-associated events such as the synthesis of proteases, regulation of plasminogen activator activity, prostaglandin and nitric oxide production. IL-1 also regulates ovarian steroidogenesis. These different aspects of the involvement of the IL-1 system in important aspects of female reproduction are discussed.


Altered Nocturnal Cardiovascular Control in Children With Sleep-Disordered Breathing.
To assess cardiovascular control during sleep in children with sleep-disordered breathing (SDB) and the effect of adenotonsillectomy in comparison to healthy nonsnoring children. Cardiorespiratory signals obtained from overnight polysomnographic recordings of 28 children with SDB and 34 healthy nonsnoring children were analyzed. We employed an autoregressive closed-loop model with heart period (RR) and pulse transit time (PTT) as outputs and respiration as an external input to obtain estimates of respiratory gain and baroreflex gain. Mean and variability of PTT were increased in children with SDB across all stages of sleep. Low frequency power of RR and PTT were attenuated during non-rapid eye movement (REM) sleep. Baroreflex sensitivity was reduced in children with SDB in stage 2 sleep, while respiratory gain was increased in slow wave sleep. After adenotonsillectomy, these indices normalized in the SDB group attaining values comparable to those of healthy children. In children with mild-to-moderate SDB, vasomotor activity is increased and baroreflex sensitivity decreased during quiet, event-free non-REM sleep. Adenotonsillectomy appears to reverse this effect.


Quantitative trait loci affecting milk production traits on bovine chromosome 6 in zebuine Gyr breed.
Fourteen Brazilian dairy Gyr sire families with 657 daughters were analyzed for quantitative trait loci (QTL) on chromosome 6 by using a daughter design for 5 economic traits: milk, fat, and protein production, fat and protein percentage. The cows and sires were genotyped for 27 microsatellites with average spacing between markers of 4.9 cM. In the analyses across 14 families, for the largest significant families, and within family, a QTL was located for milk yield and fat yield close to marker BMS2508 at the 5% chromosome-wide significance level across families and 1% chromosome-wide within families. For fat percentage, a QTL near DIK4482 was identified at the 5% chromosome-wide significance level when all families were analyzed together and at the 1% chromosome-wide significance level within the largest significant families. The different analyses yielded results that were generally consistent for milk yield, fat yield, and fat percentage. The order of the markers in the derived map was consistent with that in the consensus map. Some QTL and candidate genes in dairy cattle for milk production traits are probably preserved in Bos taurus and Bos indicus.


Possibilities and problems with identification and determination of "new" hypnotics.
Authors discuss problems with identification and determination of flunitrazepam and zolpidem in biological material (BM). Over the recent years, these two structurally different substances have become the most frequently used as well as abused hypnotic drugs. This study presents applicability of immunochemical methods in the screening of flunitrazepam, one of the most commonly prescribed drugs among the benzodiazepines. Herein described techniques, a liquid-liquid (L-L) extraction, solid phase extraction (SPE) and the so-called "freeze out" method are used for isolation of the above mentioned compounds from BM. Besides the thin layer chromatography (TLC) and gas chromatography - mass spectrometry (GC-MS) applied in qualitative analysis, the study also describes a gas chromatography with electron capture detector (GC-ECD) and gas chromatography with nitrogen phosphorus detector (GC-NPD) optimized for the determination of flunitrazepam and zolpidem in blood (serum). Successful analyses of these two substances are of major importance, especially in interpreting the results of forensic toxicological examinations.


[Analysis of factors causing peripheral visual field defects after vitreomacular surgery].
As a result of the study analysis of potential causes of visual field defects (VFD) after vitreomacular surgery is performed. 110 cases are enrolled in the study: 81 patient with macular hole and 29 patients with epiretinal fibrosis. Correspondence of VFD to position of irrigation port including unusual inferior-nasal position was revealed as well as VFD absence in epiretinal fibrosis operated without air pump. VFD did not appear after lowering of air pump pressure by 15-20 mmHg. The cause of VFD after vitreomacular surgery is a mechanical damage of internal retinal surface by directed air/gas jet after total fluid exchange with too high pressure in an air pump. This complication is an easily preventable iatrogenic effect and may serve as one of quality criterion for this treatment procedure.


A review on the use of bulk specimen X-ray microanalysis in cancer research.
The freeze-fracture, freeze-drying (FFFD) method of biological bulk specimen preparation combined with quantitative X-ray microanalysis is suitable for the measurement of intracellular concentrations of biologically relevant elements in human biopsy or experimental animal materials. Especially useful information can be obtained regarding the intracellular Na+/K+ ratios being independent of the actual (and unknown) water content of the cytoplasm. The sustained increase of this ratio indicates a sustained depolarization of the cell membrane. These data are of importance from the point of view of the membrane hypothesis of mitogenesis (MHM). It has been revealed that the distribution histograms of the intracellular Na+/K+ ratio display a very significant broadening and an increase of the average values in human urogenital, thyroid and laryngeal tumors, as well as in experimentally induced cell proliferation models. Although MHM has been claimed to be invalid on the basis of some atomic absorption measurements of the intracellular monovalent ion concentrations as well as of some in vitro results obtained with amiloride, this review paper demonstrates that MHM may still be a valid hypothesis for the explanation of mitotic regulation.


Effect of trifluoperazine on renal epithelioid Madin-Darby canine kidney cells.
Following exposure to a number of hormones, the cell membrane in Madin-Darby Canine Kidney (MDCK) cells is hyperpolarized by increase of intracellular calcium activity. The present study has been performed to elucidate the possible role of calmodulin in the regulation of intracellular calcium activity and cell membrane potential. To this end trifluoperazine has been added during continuous recording of cell membrane potential or intracellular calcium. Trifluoperazine leads to a transient increase of intracellular calcium as well as a sustained hyperpolarization of the cell membrane by activation of calcium sensitive K+ channels. Half-maximal effects are observed between 1 and 10 mumol/L trifluoperazine. A further calmodulin antagonist, chlorpromazine, (50 mumol/L), similarly hyperpolarizes the cell membrane. The effects of trifluoperazine are virtually abolished in the absence of extracellular calcium. Pretreatment of the cells with either pertussis toxin or phorbol-ester TPA does not interfere with the hyperpolarizing effect of trifluoperazine. In conclusion, calmodulin is apparently involved in the regulation of calcium transfer across the cell membrane but not in the stimulation of K+ channels by intracellular calcium.


[The temporal correlates of evoked potentials from the visual and motor cortices in affective states].
Peak latencies (PLs) of seven isopolar components of the evoked potentials (EPs) of the occipital and central areas in patients with different extent of depressions with the predominance of alarm or anguish affects were compared with healthy ones. In alarm affect the differences in PLs of EP components dominant in the occipital areas while in anguish affect they prevail in the central cortical zones.


Comprehensive circular RNA profiling reveals circ_0002060 as a potential diagnostic biomarkers for osteoporosis.
Osteoporosis (OP) is a multifactorial and common bone disease worldwide. The clinical diagnosis and treatment of osteoporosis has limitations since changes in bone density occur slightly. Thus, it is critically needed to look for new biomarkers to identify and cure osteoporosis. CircRNAs are important molecular regulators that participate in various biological processes and present tremendous therapeutic potential. The role of circRNAs in osteoporosis remains largely unknown. Here, we explore the expression of circRNAs in osteoporosis using microarray analysis, and identify potential circRNAs associated with osteoporosis by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) validation and statistical analysis in clinical samples. As a result, circ_0006873 and circ_0002060 were found to be significantly upregulated between nonosteoporotic controls and osteoporosis patients (n = 40, respectively). Pearson correlation analysis showed that the level of circ_0006873 and circ_0002060 were associated with the low-bone mineral density in patients with osteoporosis. Furthermore, ROC analysis indicated that circ_0002060 showed potential diagnostic value for osteoporosis. Taken together results using bioinformatics, experimental analysis, and disease models, we revealed that circ_0002060 could be used as potential diagnostic biomarker and therapeutic target in osteoporosis.


Identification by mutagenesis of conserved arginine and glutamate residues in the C-terminal domain of rat liver carnitine palmitoyltransferase I that are important for catalytic activity and malonyl-CoA sensitivity.
Carnitine palmitoyltransferase I (CPTI) catalyzes the conversion of long chain fatty acyl-CoAs to acylcarnitines in the presence of l-carnitine. To determine the role of the conserved glutamate residue, Glu-603, on catalysis and malonyl-CoA sensitivity, we separately changed the residue to alanine, histidine, glutamine, and aspartate. Substitution of Glu-603 with alanine or histidine resulted in complete loss of L-CPTI activity. A change of Glu-603 to glutamine caused a significant decrease in catalytic activity and malonyl-CoA sensitivity. Substitution of Glu-603 with aspartate, a negatively charged amino acid with only one methyl group less than the glutamate residue in the wild type enzyme, resulted in partial loss in CPTI activity and a 15-fold decrease in malonyl-CoA sensitivity. The mutant L-CPTI with a replacement of the conserved Arg-601 or Arg-606 with alanine also showed over 40-fold decrease in malonyl-CoA sensitivity, suggesting that these two conserved residues may be important for substrate and inhibitor binding. Since a conservative substitution of Glu-603 to aspartate or glutamine resulted in partial loss of activity and malonyl-CoA sensitivity, it further suggests that the negative charge and the longer side chain of glutamate are essential for catalysis and malonyl-CoA sensitivity. We predict that this region of L-CPTI spanning these conserved C-terminal residues may be the region of the protein involved in binding the CoA moiety of palmitoyl-CoA and malonyl-CoA and/or the putative low affinity acyl-CoA/malonyl-CoA binding site.


Dietary restriction, polyamines and monocrotaline-induced pulmonary hypertension.
Dietary restriction (DR), i.e. reduction of total caloric intake, has been shown to result in protection against monocrotaline (MCT)-induced pulmonary hypertension (PH). Restriction of the diet to 8 g/rat/day instead of the usual intake (18 g/rat/day), inhibits the progression of cardiopulmonary changes and prolongs survival after a single dose of MCT. We have shown previously that the development of MCT-induced pulmonary hypertension is associated with inhibition of polyamine biosynthesis in the lungs of MCT-treated rats. In the present study, we tested the hypothesis that DR provides protection against the development of chronic PH in the rat by limiting increases in polyamine and DNA synthesis. We randomly divided animals into four groups each (MCT, MCT + DR, control, and control + DR). We injected rats with a single dose of MCT (60 mg/kg, s.c.) and a corresponding number of control rats with vehicle. Animals in all groups were given free access to food and water prior to administration of MCT. Immediately following injection of MCT both the MCT and control groups were given free access to food and water, while the other groups (MCT + DR and control + DR) we given the restricted diet (8 g/rat/day). Daily measurements were made of body weight and of water and food intake. Animals were killed in each group at 1, 4, 7, 14, and 21 days post MCT to determine right ventricular hypertrophy (RVH), lung wet weight, ornithine decarboxylase (ODC) activity, and polyamine and DNA contents. We measured DNA synthesis 7 days after MCT by determining [3H]thymidine incorporation into the whole lung DNA. We found that 7 days after MCT treatment DNA synthesis increased compared to control. However, DR (MCT + DR) treatmen prevented the increase in DNA synthesis following MCT. Right ventricular hypertrophy, lung wet weight, ODC activity and lung polyamine levels were increased following MCT. Treatment with DR (MCT + DR) prevented increases in RVH, lung wet weight, ODC activity and lung polyamine levels. We conclude that DR to 8 g/day/rat protects against MCT-induced PH and is associated with an inhibition of increased lung polyamine and DNA synthesis that occur in the lung during the development of MCT-induced PH. These results are consistent with a recent report which suggests that increased lung polyamine biosynthesis is required for the development of MCT-induced PH. The data are also consistent with the hypothesis that inhibition of polyamine biosynthesis influences the development of MCT-induced PH in part by regulating DNA synthesis in key lung cells.


Acupuncture treatment of urticaria.
Acupuncture has long been used to treat urticaria in the Asian world. Acute urticaria can be easily and effectively treated with acupuncture. L111 (Quchi), Sp10 (Xuehai), Sp6 (Sanyinjiao), and S36 (Zusanli) are the 4 acupuncture points most commonly prescribed. Chronic urticaria is a challenge for medical therapy. There are at least 6 kinds of acupuncture methods developed to overcome this challenge. The combination of ordinary acupuncture and auricular acupuncture has been observed to be a highly effective cure of chronic urticaria. Acupuncture point injection with thiamine hydrochloride (vitamin B1) is also an effective treatment. However, results of these clinical observations have not been systematically confirmed for lack of a control group and the need for standard classification of urticaria. Although these observational results have clinical limitations, they do offer insight into an alternative to conventional treatment of urticaria. In the future, acupuncture or acupuncturelike techniques may provide an effective alternative for treatment of patients with urticaria, particularly those refractory to medication therapy.


Comparison of the discriminative stimulus effects of salvinorin A and its derivatives to U69,593 and U50,488 in rats.
Research interests regarding the psychopharmacology of salvinorin A have been motivated by the recreational use and widespread media focus on the hallucinogenic plant, Salvia divinorum. Additionally, kappa opioid (KOP) receptor ligands may have therapeutic potential in the treatment of some neuropsychiatric conditions, including drug dependence and mood disorders. Salvinorin A is a selective KOP agonist, but only a few studies have explored the discriminative stimulus effects of this compound. This study compared the discriminative stimulus effects of salvinorin A and two synthetic derivatives of salvinorin B to the KOP agonists, U69,593 and U50,488. Sixteen male Sprague-Dawley rats trained to discriminate U69,593 (0.13 mg/kg, s.c., N = 8) or U50,488 (3.0 mg/kg, i.p., N = 8) under a fixed-ratio 20 schedule of food reinforcement were administered substitution tests with salvinorin A (0.125-3.0 mg/kg, i.p.). The animals trained to discriminate U69,593 were also administered substitution tests with salvinorin B ethoxymethyl ether (0.005-0.10 mg/kg, i.p.) and salvinorin B methoxymethyl ether (0.03-0.10 mg/kg, i.p.). Another eight rats were trained to discriminate 2.0 mg/kg salvinorin A and tested with U69,593 (0.04-0.32 mg/kg) and U50,488 (0.4-3.2 mg/kg). Salvinorin A and both synthetic derivatives of salvinorin B substituted completely for U69,593. Additionally, cross-generalization was observed between salvinorin A and both KOP agonists. These findings support previous reports indicating that the discriminative stimulus effects of salvinorin A are mediated by kappa receptors. Future studies may assist in the development and screening of salvinorin A analogs for potential pharmacotherapy.


Abnormal lymphocyte subsets in Kuwaiti patients with type-1 insulin-dependent diabetes mellitus and their first-degree relatives.
Circulating lymphocyte subset imbalance is associated with type-1 insulin-dependent diabetes mellitus (IDDM). To examine the imbalance in these immunoregulatory cells in Kuwaitis with type-1 diabetes and their first-degree relatives we analysed T-lymphocyte subsets and HLA-DR expression (activation) in 18 IDDM patients with a family history of IDDM and 18 non-diabetic first-degree relatives of the IDDM patients. Both IDDM patients and their first-degree relatives showed a mild lymphopenia. Total T lymphocytes, CD3+ cells, in IDDM patients and their first-degree relatives were reduced compared to control subjects (P < 0.001). Total B lymphocytes, CD19+ cells, was increased in IDDM patients (P = 0.001), but was comparable to controls in IDDM patients' first-degree relatives. No quantitative abnormality was demonstrated in CD4+ cells in IDDM patients; however, these cells were higher in their first-degree relatives (P = 0.0089). Suppressor T lymphocytes, CD8+ cells, in first-degree relatives and controls were not significantly different; however, these cells were significantly reduced in IDDM patients (P = 0.001). The ratio of CD4+/CD8+ cells was higher in IDDM patients and their first-degree relatives compared to controls (P = 0.0007 and 0.0103, respectively). Activated T lymphocytes, HLA-DR+ CD3+ cells, were significantly increased in IDDM patients and their first-degree relatives. HLA-DR3 was the most common antigen found in IDDM patients (77% vs. 20% in controls, P = 0.00021). The second most common antigen was HLA-DR4 (55% vs. 24% in controls, P = 0.0566). However, no relationship was found in the levels of CD3+, CD4+ or CD8+ cells in patients possessing either DR3 or DR4. These results suggest that T-lymphocyte subset imbalance not only characterizes the cellular autoimmunity in the pathogenesis of IDDM but may also be significant in early pre-diabetic stages in those with a family history of IDDM.


Latency to traverse a T-maze at 2 days of age and later adrenocortical responses to an acute stressor in domestic chicks.
Latencies to escape from a T-maze, and thereby reinstate visual contact with conspecifics, were measured in broiler chicks at 2 days of age. Chicks were assigned to high- (HP) or low- (LP) performance categories if their escape latencies fell below 25 s or above 75 s, respectively. These chicks were then housed socially in 10 same-category groups (5 HP, 5 LP), each comprising eight birds. At 15 days of age, one chick was taken from each of two randomly selected cages (1 HP, 1 LP) and immediately bled (undisturbed controls). At the same time, another chick was taken from each of these boxes and immersed up to its neck in warm water (partial water immersion, PWI) for 15 min before blood was collected. All chicks were sexed after bleeding. There were no differences between the plasma corticosterone (CS) levels of undisturbed (control) HP and LP chicks. Exposure to PWI significantly increased circulating CS levels, and this elevation was more pronounced in LP than in HP chicks. Male chicks also showed higher stress-induced adrenocortical responses than did females. The present findings suggest that the T-maze responses of young chicks might predict their later adrenocortical responses to a known stressor. This relationship is discussed in terms of individual differences in fearfulness, ability to cope with challenge, and/or stress susceptibility.


The effect of improvements in cytometer sensitivity on the detection of CD5-positive B cells with dim fluorescence.
When antigen density on the surface of a cell population is low and variable, the percentage of that population determined to express the antigen (i.e., to be positively stained) depends directly on the sensitivity of the flow cytometer for resolving particles which are dimly fluorescent from those which are unstained. In this study, the sensitivity of a commercial flow cytometer has been improved by changes in the photomultiplier tube, the fluorescence filter, and the amount of stray light entering the fluorescence channel. In a model system with human lymphocytes, modifications to these factors increased the percent of the B-lymphocyte population found to express the CD5 antigen.


Metformin Inhibits N-Methyl-N-Nitrosourea Induced Gastric Tumorigenesis in db/db Mice.
Type 2 diabetes can elevate risk of gastric cancer and metformin, an anti-diabetic agent, has an inhibitory effect against gastric cancer cell in vitro. However, the effect of metformin on type 2 diabetes-related gastric tumorigenesis in vivo is still not clear. In the present study, we aim to detect whether metformin can inhibit increased risk of gastric cancer in diabetic db/db mice and which the potential anti-cancer mechanisms of metformin are. 4-week-old mice were divided into 3 groups (2 db/db mice groups and one wild type mice group). All diabetic and non-diabetic mice were treated with N-Methyl-N-Nitrosourea (MNU) for 20 weeks to induce gastric tumorigenesis. At week 21, one db/db mice group were treated with metformin (5 mg/ml) for 10 weeks and the other 2 groups were treated with saline. Blood samples were collected for testing insulin and insulin-like growth factor (IGF)-1. Stomach tissues were collected for histopathological evaluation and mRNAs analysis. Metformin significantly decreased incidence of MNU-induced gastric dysplasia and cancer in diabetic db/db mice. Furthermore, metformin reduced serum insulin as well as IGF-1, and also suppressed expression of insulin receptor, IGF-1, IGF-1 receptor and several pro-inflammatory cytokines mRNAs in stomach of db/db mice, but did not significantly influence IGF-2 and IGF-2 receptor expressions. The results show that metformin can prevent the risk of gastric cancer in type 2 diabetes and the protective mechanisms may involve in an inhibitory effect of metformin on insulin as well as IGF-1 signals and cancer related pro-inflammatory cytokines.


Neurological features of congenital fibrosis of the extraocular muscles type 2 with mutations in PHOX2A.
Congenital fibrosis of the extraocular muscles type 2 (CFEOM2) is a complex strabismus syndrome that results from mutations in the homeodomain transcription factor PHOX2A. To define the clinical and neuroimaging features of patients with this autosomal recessive syndrome, we studied 15 patients with genetically defined CFEOM2. All patients underwent full neurological, neuro-ophthalmological and orthoptic assessments. Twelve patients had pupillary pharmacological testing and nine had 3.0 tesla MRI of the brain, brainstem and orbits. Patients were born with severe bilateral ptosis and exotropia with almost complete bilateral absence of adduction, elevation, depression and intorsion. Variable abduction was present prior to strabismus surgery in 14 patients, and central ocular motility reflexes (smooth pursuit, saccades, vestibulo-ocular reflex and optokinetic reflex) were intact except for convergence. Pupillary light and near reflexes were not present, but irises were anatomically normal and responded to pupillary pharmacology. Neuroimaging of brain and brainstem was remarkable for the anatomical absence of cranial nerve (CN) 3 and probably CN 4 bilaterally. Therefore, the CFEOM2 phenotype and neuroimaging are both consistent with the congenital absence of CNs 3 and 4. Additional features included presence of most central ocular motility reflexes, a central lack of pupillary responsiveness of uncertain aetiology and modest phenotypic variability that does not correlate with specific PHOX2A mutations. Clinical presentation, neuroimaging and Phox2a-/- animal models all support the concept that CFEOM2 is a primary neurogenic abnormality with secondary myopathic changes.


Identification of protein receptors for coronaviruses by mass spectrometry.
As obligate intracellular parasites, viruses need to cross the plasma membrane and deliver their genome inside the cell. This step is initiated by the recognition of receptors present on the host cell surface. Receptors can be major determinants of tropism, host range, and pathogenesis. Identifying virus receptors can give clues to these aspects and can lead to the design of intervention strategies. Interfering with receptor recognition is an attractive antiviral therapy, since it occurs before the viral genome has reached the relative safe haven within the cell. This chapter describes the use of an immunoprecipitation approach with Fc-tagged viral spike proteins followed by mass spectrometry to identify and characterize the receptor for the Middle East respiratory syndrome coronavirus. This technique can be adapted to identify other viral receptors.


Demonstration of regurgitation of gut content during blood meals of the tick Ornithodoros moubata. Possible role in the transmission of pathogenic agents.
Nymphs of Ornithodoros moubata were fed tritiated ecdysteroid. These ingested hormones are conjugated to fatty acyl esters that accumulate in the midgut (Connat et al. 1988). A few months later, the same ticks which had molted, were fed on physiological medium without radiolabel. At the issue of the blood meal, the nutritive medium contained an amount of radiolabel corresponding to 0.5% of the total labelling in the animal before the blood meal; this corresponded to 1.3% of the midgut content. These results demonstrate that in addition to transmission of parasites by saliva and coxal fluid (Burgdorfer 1951), transmission through regurgitation of the blood content in the gut could occur. An equivalent quantity of radiolabel was also emitted in the feces during and after the meal, but no conclusion about parasite transmission can be drawn from these "metabolic" results.


Vitamin D attenuates inflammation in CFTR knockdown intestinal epithelial cells but has no effect in cells with intact CFTR.
The cystic fibrosis (CF) intestine is characterized by chronic inflammation. CF patients are instructed to ingest supplemental vitamin D on a daily basis thereby exposing their intestinal tract to pharmacological amounts of this vitamin. It has been shown that vitamin D exerts intestinal anti-inflammatory properties. We therefore postulate that vitamin D may be beneficial in the management of CF intestinal inflammation by attenuating cellular inflammatory responses. In this study, we investigated the anti-inflammatory effects of the oral form of vitamin D3 (cholecalciferol) and its metabolites, 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3, on cytokine-induced inflammatory responses in intestinal epithelial Caco-2/15 cells with intact expression of CF transmembrane conductance regulator (CFTR) and knockdown for CFTR. We show that 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3 inhibited p38MAPK phosphorylation and that these effects were not mediated by changes in the expression of MAPK phosphatase-1 (MKP-1). However, 1,25-dihydroxyvitamin D3 exhibited superior anti-inflammatory effects as it furthermore reduced cytokine-induced NF-κB nuclear translocation and interleukin-8 mRNA stability and secretion. Intriguingly, the anti-inflammatory effects of vitamin D metabolites were only observed in CFTR knockdown cells, which may be explained by alterations in its catabolism associated with changes in CYP24A1 expression. These observations were supported in vivo whereby Cftr(-/-) mice fed large amounts of vitamin D3 for 2 mo led to a reduction in the number of eosinophils and apoptotic cells in the duodenal mucosa of females but not males. Altogether, these findings suggest that vitamin D exerts intestinal anti-inflammatory actions under specific circumstances and may thus prove beneficial in CF.


Immunogenic effect of bacteriophage in patients subjected to phage therapy.
Fifty seven cases of bacterial infections subjected to phage therapy were tested for a production of antibodies against the applied bacteriophages. Monoinfections confirmed in 40 patients were caused in majority of cases by pyogenic Staphylococci (29 cases) and rarely by Gram-negative bacteria: Klebsiella, Escherichia, Proteus and Pseudomonas (11 cases). Polyinfections caused by the above types of bacteria were recorded in 17 cases. The titer of neutralizing and hemagglutinating antibodies was determined before phage therapy, in the 10th day and in some cases in the 21st day of its course. The effect of natural and immune antibodies on the final result of therapy was analyzed.


Understanding and revisiting properties of EuTiO3 bulk material and films from first principles.
Ab initio computations are performed to investigate properties of bulk material and epitaxial films made of EuTiO3 (ETO). A whole family of nanoscale twinned phases, that present complex oxygen octahedra tilting (OOT) and unusual antiferroelectricity, is found to be degenerate in energy with simpler phases (all possessing typical OOT) in bulk ETO. Such degeneracy provides a successful explanation of recently observed anomalous phenomena. The calculations also lead to revisiting the (rich) phase diagram of ETO films.


Diagnosis, staging, and prognostic factors of small cell lung cancer.
The proportion of small cell-large cell or combined small-cell subtypes is very low, and the clinical outcome in patients with these subtypes appears not to be significantly different from that of the majority of patients with small-cell lung cancer. Anatomic staging of small-cell lung cancer remains necessary if the outcome influences treatment decisions and for treatment in many clinical trials. The influence of simple clinical and biochemical variables on prognosis has been confirmed by the analysis of very large numbers of patients. Patients with small-cell lung cancer surviving at 2 years after chemotherapy continue to be at risk for relapse for up to 5 more years independent of the initial stage of disease. Major future directions will include the investigation into possible correlations of biologic properties of tumors with prognosis and the use of simple clinical and biochemical parameters to preselect patients for treatment strategies according to risk groups.