Discovery of low-affinity preproinsulin epitopes and detection of autoreactive CD8 T-cells using combinatorial MHC multimers

Autoreactive cytotoxic CD8 T-cells (CTLs) play a key pathogenic role in the destruction of insulin-producing beta-cells resulting in type 1 diabetes. However, knowledge regarding their targets is limited, restricting the ability to monitor the course of the disease and immune interventions. In a multi-step discovery process to identify novel CTL epitopes in human preproinsulin (PPI), PPI was digested with purified human proteasomes, and resulting COOH-fragments aligned with algorithm-predicted HLA-binding peptides to yield nine potential HLA-A1, -A2, -A3 or -B7-restricted candidates. An UV-exchange method allowed the generation of a repertoire of multimers including low-affinity HLA-binding peptides. These were labeled with quantum dot-fluorochromes and encoded in a combinatorial fashion, allowing parallel and sensitive detection of specific, low-avidity T-cells. Significantly increased frequencies of T-cells against four novel PPI epitopes (PPI_4-13/B7, PPI_29-38/A2, PPI_76-84/A3 and PPI_79-88/A3) were detected in stored blood of patients with recent onset diabetes but not in controls. Changes in frequencies of circulating CD8 T-cells against these novel epitopes were detected in blood of islet graft recipients at different time points after transplantation, which correlated with clinical outcome. In conclusion, our novel strategy involving a sensitive multiplex detection technology and requiring minimal volumes of stored blood represents a major improvement in the direct ex-vivo characterization and enumeration of immune cells in the pathogenesis of type 1 diabetes.     

Households that harbour zero-dose children after door-to-door national immunization campaigns

Data from two consecutive rounds of national immunization days were analysed to identify households that harboured zero-dose children after door-to-door immunization campaigns, and to highlight settings and features associated with these households in Chad. The proportion of households harbouring zero-dose children were higher among rural than among urban dwellers and were prevalent among parents who did not know the benefit of polio immunization. These results show the need for more attention to be given to rural dwellers and parent awareness of the benefit of polio immunization.     

Abnormal white matter microstructure in schizophrenia: a voxelwise analysis of axial and radial diffusivity

Diffusion Tensor Imaging (DTI) investigations in schizophrenia have provided evidence of impairment in white matter as indicated by reduced fractional anisotropy (FA). However, the neuropathological implications of these findings remain unclear. In the current study, we conducted a voxelwise analysis of the constituent parameters of FA, Axial (λ_||) and Radial Diffusivity (λ_┴), in 14 male participants with schizophrenia and 14 age, gender, education, and premorbid intelligence matched healthy controls. Significantly reduced FA and higher Radial Diffusivity were concurrently observed in several major white matter tracts in the schizophrenia group. This finding suggests that the loss of white matter integrity in schizophrenia is the result of demyelination and/or changes to the axonal cytoskeleton rather than gross axonal damage.     

Pioglitazone inhibits androgen production in NCI-H295R cells by regulating gene expression of CYP17 and HSD3B2

Thiazolidinediones (TZDs) such as pioglitazone and rosiglitazone are widely used as insulin sensitizers in the treatment of type 2 diabetes. In diabetic women with polycystic ovary syndrome, treatment with pioglitazone or rosiglitazone improves insulin resistance and hyperandrogenism, but the mechanism by which TZDs down-regulate androgen production is unknown. Androgens are synthesized in the human gonads as well as the adrenals. We studied the regulation of androgen production by analyzing the effect of pioglitazone and rosiglitazone on steroidogenesis in human adrenal NCI-H295R cells, an established in vitro model of steroidogenesis of the human adrenal cortex. Both TZDs changed the steroid profile of the NCI-H295R cells and inhibited the activities of P450c17 and 3betaHSDII, key enzymes of androgen biosynthesis. Pioglitazone but not rosiglitazone inhibited the expression of the CYP17 and HSD3B2 genes. Likewise, pioglitazone repressed basal and 8-bromo-cAMP-stimulated activities of CYP17 and HSD3B2 promoter reporters in NCI-H295R cells. However, pioglitazone did not change the activity of a cAMP-responsive luciferase reporter, indicating that it does not influence cAMP/protein kinase A/cAMP response element-binding protein pathway signaling. Although peroxisome proliferator-activated receptor gamma (PPARgamma) is the nuclear receptor for TZDs, suppression of PPARgamma by small interfering RNA technique did not alter the inhibitory effect of pioglitazone on CYP17 and HSD3B2 expression, suggesting that the action of pioglitazone is independent of PPARgamma. On the other hand, treatment of NCI-H295R cells with mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) inhibitor 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one (PD98059) enhanced promoter activity and expression of CYP17. This effect was reversed by pioglitazone treatment, indicating that the MEK/ERK signaling pathway plays a role in regulating androgen biosynthesis by pioglitazone.     

Cardiac abnormalities as a new manifestation of nonalcoholic fatty liver disease: echocardiographic and tissue Doppler imaging assessment

Nonalcoholic fatty liver disease (NAFLD) is linked to the metabolic syndrome. The aim of the present study is to determine the effect of the metabolic syndrome on left ventricular (LV) geometry and function using as a model patients with NAFLD. Thirty-eight patients with NAFLD, less than 55 years of age and with a normal exercise test, were compared with an age and sex-matched control group. Patients with diabetes mellitus, hypertension, and body mass index>40 were excluded. A complete echocardiographic study including tissue Doppler imaging (TDI) was performed. The following parameters were assessed by echo Doppler: peak velocities of early (E) and late (A) diastolic filling, E/A ratio, flow propagation velocity (Vp). Using TDI early diastolic velocity (E'), and systolic velocity (S') of mitral annulus were obtained. The patients with NAFLD had a significantly higher body mass index (31.4+/-5 vs. 26.4+/-4 kg/m, P=0.01), higher glucose (100.6+/-13 vs. 83.0+/-10 mg/dL, P=0.01), and triglyceride levels (126.5+/-44 vs. 206.5+/-67 mg/dL, P<0.001). Increased thickness of the intraventricular septum, posterior wall (11.03+/-2.2 vs. 8.9+/-2.9 mm, P=0.001; 8.5+/-1.7 vs. 9.7+/-2.3 mm, P=0.04), and larger LV mass and LV mass/height (160.7+/-58.7 vs.115.3+/-35.4 g, P=0.001 and 92.6+/-29.5 vs. 69.2+/-19.8 g/m, P=0.001, respectively) were found in NAFLD group. LV systolic function was similar in both groups. Patients with NAFLD had a lower E (73.6+/-11.0 vs. 86.4+/-20.0 cm/s, P<0.006) and E/A ratio (1.0+/-0.3 vs. 1.76+/-0.8 P<0.0001). Moreover, the Vp and the E' on TDI were significantly lower compared with the control group (49.0+/-9.7 vs. 74.7+/-18.4 cm/s, P<0.0001 and 10.3+/-2.0 vs. 13.8+/-1.7 cm/s, P<0.0001, respectively). On multivariate analysis the E' on TDI was the only independent parameter associated with NAFLD. In conclusion, patients with NAFLD in the absence of morbid obesity, hypertension, and diabetes have mildly altered LV geometry and early features of left ventricular diastolic dysfunction. Early diastolic velocity on TDI was found to be the only index that could identify the patients with NAFLD and metabolic syndrome.     

Long-term benefits and harms of early colorectal cancer screening in German individuals with familial cancer risk

Individuals with a family history of colorectal cancer (CRC) may benefit from early screening with colonoscopy or immunologic fecal occult blood testing (iFOBT). We systematically evaluated the benefit-harm trade-offs of various screening strategies differing by screening test (colonoscopy or iFOBT), interval (iFOBT: annual/biennial; colonoscopy: 10-yearly) and age at start (30, 35, 40, 45, 50 and 55 years) and end of screening (65, 70 and 75 years) offered to individuals identified with familial CRC risk in Germany. A Markov-state-transition model was developed and used to estimate health benefits (CRC-related deaths avoided, life-years gained [LYG]), potential harms (eg, associated with additional colonoscopies) and incremental harm-benefit ratios (IHBR) for each strategy. Both benefits and harms increased with earlier start and shorter intervals of screening. When screening started before age 50, 32-36 CRC-related deaths per 1000 persons were avoided with colonoscopy and 29-34 with iFOBT screening, compared to 29-31 (colonoscopy) and 28-30 (iFOBT) CRC-related deaths per 1000 persons when starting age 50 or older, respectively. For iFOBT screening, the IHBRs expressed as additional colonoscopies per LYG were one (biennial, age 45-65 vs no screening), four (biennial, age 35-65), six (biennial, age 30-70) and 34 (annual, age 30-54; biennial, age 55-75). Corresponding IHBRs for 10-yearly colonoscopy were four (age 55-65), 10 (age 45-65), 15 (age 35-65) and 29 (age 30-70). Offering screening with colonoscopy or iFOBT to individuals with familial CRC risk before age 50 is expected to be beneficial. Depending on the accepted IHBR threshold, 10-yearly colonoscopy or alternatively biennial iFOBT from age 30 to 70 should be recommended for this target group.