A replicon-based bioassay for the measurement of interferons in patients with chronic hepatitis C

Overall treatment results of chronic hepatitis C have improved markedly with the introduction of pegylated interferon-alpha (PEG–IFN-) and ribavirin combination therapy. However, cure rates in the most common genotype 1 infection are still unsatisfactory. IFN- dose–response studies on viral kinetics suggest that inadequate dosing might be a key factor but drug levels have hardly been tested, which is in part due to difficulties in measuring this cytokine in patient samples. We have shown recently that hepatitis C virus (HCV) replicons are highly sensitive to IFN-. In this report we tested whether the replicon system could be used as a sensitive bioassay to determine the amount of biologically active IFN- in serum or heparinized plasma of patients under therapy. To facilitate the measurements, a stably replicating subgenomic HCV RNA was developed that carries the gene encoding the firefly luciferase. Dose response studies with IFN- demonstrate that the amount of expressed luciferase directly correlates with the level of HCV replication. By using this cell-based assay, serum samples of HCV patients treated with different types and doses of IFN- were analyzed in parallel to IFN- standards made by serial dilutions of the same type of IFN- the patient was treated with. Based on nonlinear logistic models serum concentrations corresponding to 1.3–19 U/ml were determined in patients under standard or high dose IFN- therapy, and from 3.8 to 4.1 ng/ml in patients treated with PEG IFN-. In conclusion, the HCV-replicon based bioassay allows determining the levels of biologically active IFN- in serum and heparinized plasma of patients under treatment.     

Comparison of bone mineral density and its variables between premenopausal and postmenopausal women

Objectives

To compare the bone mineral density (BMD) and its variables in premenopausal and postmenopausal women.

Methods

In this cross sectional study, 62 premenopausal and 62 postmenopausal apparently healthy women were evaluated by a questionnaire. The dietary intake of calcium was evaluated by 24 hours recall method and using table for proximate principle of common Indian food by Indian Council of Medical Research (ICMR). BMD at lumbar spine, femoral neck and Ward’s triangle were measured by dual energy X-ray absorptiometry (DXA). A correlation between BMD and various variables were calculated for each of the two groups.

Results

The mean age of premenopausal and postmenopausal women was 32.46±7.8 and 51.74±7.1 years respectively. The body mass index (BMI), height and weight were comparable in both the groups. The daily intake of calcium was significantly higher in premenopausal women (p<0.01). Approximately, 17% of the postmenopausal women and 9.6% of the premenopausal women were having osteoporosis; 28.56% of the postmenopausal women and 43.54% of the premenopausal women were having osteopenia at the lumbar spine. The BMD at lumber spine was found to be statistically significantly higher in premenopausal women than that in postmenopausal women (p=0.03). BMD at lumbar spine, femoral neck and Ward’s triangle were positively correlated with height, weight, BMI in premenopausal as well in postmenopausal women.

Conclusion

A significant number of women had osteopenia during premenopausal period and osteoporosis in postmenopausal phase. By increasing awareness towards bone health in second and third decade, morbidity of osteoporosis can be reduced.     

Global molecular epidemiology of the O15:K52:H1 extraintestinal pathogenic Escherichia coli clonal group: evidence of distribution beyond Europe

Escherichia coli O15:K52:H1 is a significant extraintestinal pathogen in Europe (G. Prats et al., J. Clin. Microbiol. 38:201-209, 2000). To search for evidence of this clonal group outside of Europe, 75 non-European E. coli isolates of serogroup O15 were compared with five members of the O15:K52:H1 clonal group from Barcelona, Spain, according to genomic background, virulence genotypes, and antimicrobial resistance profiles. Amplification phylotyping showed that 16 (21%) of the 75 non-European O15 isolates corresponded with the O15:K52:H1 clonal group. The 16 non-European O15:K52:H1 clonal group members represented diverse geographic locales. They were isolated almost exclusively from humans with extraintestinal infections and accounted for 50% of all O15 isolates from five human clinical collections studied. Most non-European clonal group members exhibited a consensus virulence factor profile that included the F16 or F7-2 papA alleles (P fimbrial structural subunit), papG allele II (P fimbrial adhesin), iha (putative adhesin siderophore), and iutA (aerobactin receptor). This resembles the virulence profiles of (i) European representatives of the O15:K52:H1 clonal group and (ii) phylogenetically related "clonal group A," a recently recognized significant contributor to trimethoprim-sulfamethoxazole resistance in the United States (A. R. Manges et al., N. Engl. J. Med. 345:1007-1013, 2001). Antimicrobial resistance profiles were variable, and resistance was inconsistently transferred by conjugation. These findings indicate that the O15:K52:H1 clonal group is broadly distributed beyond Europe, exhibits previously unrecognized phenotypic and genotypic diversity, and contributes significantly to extraintestinal infections in humans.     

High-risk human papillomavirus detection: a split-sample comparison of hybrid capture and chromogenic in situ hybridization

The American Society for Colposcopy and Cervical Pathology (ASCCP) has proposed high-risk human papillomavirus (HPV) testing as the "preferred" triage for women with atypical squamous cells of undetermined significance. We studied 401 atypical squamous cells of undetermined significance liquid-based cervicovaginal cytology split samples for HPV by chromogenic in situ hybridization (CISH) and by Hybrid Capture (HC) II (Digene, Gaithersburg, MD); 202 underwent HC II followed by CISH, and 199 underwent CISH followed by HC II. Of 401 vials, 101 (25.2%) were positive for HPV by 1 or more methods. HC II labeled 83 of 401 (20.7%) samples as positive, while 38 of 401 (9.5%) were positive by CISH. Positive attributes of CISH include the provision of a cytomorphologic link in assessing HPV positivity and comparative ease of use in laboratories without trained molecular diagnosticians. Greater efficacy and quantitative design are advantages of HC II. Comparing data by sequence of testing showed a lower likelihood of positive test results on the second ancillary test than on the first ancillary test, regardless of age or testing method (odds ratio, second/first = 0.58; P = .003). This finding suggests that liquid-based cervicovaginal cytology samples are not homogeneous throughout. Correlative studies with histology and polymerase chain reaction may clarify predictive values for both methods.     

Paclitaxel loaded PEGylated gleceryl monooleate based nanoparticulate carriers in chemotherapy

A PEGylated drug delivery system of paclitaxel (PTX), based on glyceryl monooleate (GMO) was prepared by optimizing various parameters to explore its potential in anticancer therapy. The prepared system was characterized through polarized light microscopy, TEM, AFM and SAXS to reveal its liquid crystalline nature. As GMO based LCNPs exhibit high hemolytic toxicity and faster release of entrapped drug (66.2?±?2.5% in 24?h), PEGylation strategy was utilized to increase the hemocompatibility (reduction in hemolysis from 60.3?±?10.2 to 4.4?±?1.3%) and control the release of PTX (43.6?±?3.2% released in 24?h). The cytotoxic potential and cellular uptake was assessed in MCF-7 cell lines. Further, biodistribution studies were carried out in EAT (Ehrlich Ascites tumor) bearing mice using (99m)Tc-(Technetium radionuclide) labeled formulations and an enhanced circulation time and tumor accumulation (14 and 8 times, respectively) were observed with PEGylated carriers over plain ones, at 24?h. Finally, tumor growth inhibition experiment was performed and after 15 days, control group exhibited 15 times enhancement in tumor volume, while plain and PEGylated systems exhibited only 8 and 4 times enhancement, respectively, as compared to initial tumor volume. The results suggest that PEGylation enhances the hemocompatibility and efficacy of GMO based system that may serve as an efficient i.v. delivery vehicle for paclitaxel.     

Status of bacterial colonization, Toll-like receptor expression and nuclear factor-kappa B activation in normal and diseased human livers

Epidemiological data on bacterial translocation (BT), colonization and inflammation in normal human livers is lacking. In this study we investigated the status of bacterial colonization and inflammation in the normal, cirrhotic primary biliary cirrhosis (PBC), and nonalcoholic steatohepatitis (NASH) human liver tissues. Comparatively normal livers showed increased bacterial colonization than PBC and NASH. We analyzed mRNA levels of Toll-like receptors (TLR) 2 and TLR4, and protein levels of TLR4. Phosphorylated IKKα (pIKKα) protein estimation served as a marker for nuclear factor-kappa B (NF-κB) activation. In spite of the increased bacterial colonization in normal liver tissues, lower levels of TLR2/4 mRNA and TLR4 and pIKKα proteins were found compared to PBC and NASH indicating the maintenance of suppressed inflammation and immune tolerance in normal livers. To our knowledge, this is the first clinical evidence showing suppressed inflammation despite bacterial colonization in normal human livers thus maintaining liver immune homeostasis.     

Multiple Broad-Spectrum Beta-Lactamase Targets for Comprehensive Surveillance

Real-time PCR testing for bla_KPC, bla_NDM, bla_VIM, bla_IMP, and bla_CTX-M was performed on rectal swabs obtained from residents of two long-term acute-care facilities. While bla_KPC was detected in 69/102 swabs (67.6%), testing for four other targets increased the positivity rate for a broad-spectrum β-lactamase to 73.5% (McNemar''s P = 0.03).     

Association of Physical Function and Survival in Older-Adult Renal Transplant Recipients

There is an increase in older-adult renal transplant recipients in United States. The objective of this study was to assess the association between physical function (PF) and patient survival in renal transplant recipients who are aged 65 years or older. Using United Network for Organ Sharing (UNOS) data from 2007 to 2016, renal transplant recipients aged 65 years or older were included. Multivariable Cox regression was used to assess associations between survival and functional status adjusted for age, sex, race, donor quality, diabetes, and dialysis vintage. The study identified 26,721 patients. Patient survival was significantly higher in recipients who needed no assistance and lowest in patients in need of total assistance (P < .0001). In deceased donor (DD) transplants, the relative risk for mortality was 2.06 (1.74-2.43) for total assistance and 1.17 (1.08-1.28) for moderate assistance compared to no assistance (P < .0001). In living donor (LD) transplants, the relative risk of mortality was 1.38 (0.78-2.42) for patients needing total assistance and 1.37 (1.14-1.65) for patients needing moderate assistance compared to patients who did not need assistance (0.003). PF is an independent predictor of post-transplant mortality. Assessment of older potential renal transplant recipients should include assessment and standardization of functional status to counsel about post-transplant survival.