Human cytochrome P450scc (CYP11A1) catalyzes epoxide formation with ergosterol

Cytochrome P450scc (P450scc) catalyzes the cleavage of the side chain of both cholesterol and the vitamin D(3) precursor, 7-dehydrocholesterol. The aim of this study was to test the ability of human P450scc to metabolize ergosterol, the vitamin D(2) precursor, and define the structure of the major products. P450scc incorporated into the bilayer of phospholipid vesicles converted ergosterol to two major and four minor products with a k(cat) of 53 mol · min(-1) · mol P450scc(-1) and a K(m) of 0.18 mol ergosterol/mol phospholipid, similar to the values observed for cholesterol metabolism. The reaction of ergosterol with P450scc was scaled up to make enough of the two major products for structural analysis. From mass spectrometry, NMR, and comparison of the NMR data to that for similar molecules, we determined the structures of the two major products as 20-hydroxy-22,23-epoxy-22,23-dihydroergosterol and 22-keto-23-hydroxy-22,23-dihydroergosterol. Molecular modeling and nuclear Overhauser effect (or enhancement) spectroscopy spectra analysis helped to establish the configurations at C20, C22, and C23 and determine the final structures of major products as 22R,23S-epoxyergosta-5,7-diene-3β,20α-diol and 3β,23S-dihydroxyergosta-5,7-dien-22-one. It is likely that the formation of the second product is through a 22,23-epoxy (oxirane) intermediate followed by C22 hydroxylation with the formation of strained 22-hydroxy-22,23-epoxide (oxiranol), which is immediately transformed to the more stable α-hydroxyketone. Molecular modeling of ergosterol into the P450scc crystal structure positioned the ergosterol side chain consistent with formation of the above products. Thus, we have shown that P450scc efficiently catalyzes epoxide formation with ergosterol giving rise to novel epoxy, hydroxy, and keto derivatives, without causing cleavage of the side chain.     

C8orf13–BLK is a genetic risk locus for systemic sclerosis and has additive effects with BANK1: Results from a large french cohort and meta‐analysis

Objective

Accumulating evidence suggests that B cells are involved in systemic sclerosis (SSc). BANK1 has been reproducibly reported to be associated with diffuse cutaneous SSc (dcSSc). BLK encodes another B cell signal transducer, and a functional variant at the C8orf13–BLK locus has been found to be associated with SSc in Caucasians. However, no independent replication has been reported, and there are discrepancies in the genotype–phenotype correlation between these studies in Caucasians and another study performed in the Japanese population. Therefore, in a large cohort of French Caucasians and using a meta‐analysis of the available data, this study was undertaken to determine whether the C8orf13–BLK locus is associated with SSc, and to assess the possibility of interaction between BLK and BANK1 in SSc.

Methods

The C8orf13–BLK rs13277113 genotype was determined in 1,031 patients with SSc and 1,014 control subjects for whom BANK1 genotypes were available. Meta‐analysis of the 3 available data sets (6,078 individuals) was also performed.

Results

Minor allele frequencies for rs13277113 revealed an association restricted to the dcSSc subtype (P = 0.012, odds ratio [OR] 1.29) in the French sample. Meta‐analysis of the combined Caucasian populations showed an association of this genotype with both SSc (P = 0.0013, OR 1.16, 95% confidence interval [95% CI] 1.06–1.26) and dcSSc (P = 0.0012, OR 1.23, 95% CI 1.08–1.39). Inclusion of the Japanese population confirmed the overall association with the disease, with the strongest association observed with dcSSc (P = 3.27 × 10⁻⁵, OR 1.27). Secondary analysis in the French sample revealed additive effects between C8orf13–BLK and BANK1, mainly in the dcSSc subset.

Conclusion

These results confirm C8orf13–BLK as an SSc risk locus. The strongest effects, and particularly additive effects, were observed in the interaction between C8orf13–BLK and BANK1 in the dcSSc subset.

     

Cardiovascular risk assessment in persons with HIV infection in the developing world: comparing three risk equations in a cohort of HIV-infected Thais

Objective

There is growing concern regarding cardiovascular disease in HIV‐infected individuals in developing countries such as Thailand. We evaluated the 10‐year risk of coronary heart disease (CHD) in a Thai HIV‐infected cohort using three cardiovascular risk equations, and assessed the level of agreement among their predictions.

Methods

We carried out a cross‐sectional analysis of data on 785 Thai subjects followed prospectively in the HIV Netherlands Australia Thailand Collaboration (HIV‐NAT) cohort study from 1996 to 2009. Cardiovascular risk factor history, along with relevant laboratory and clinical data, was collected at follow‐up clinic visits. Ten‐year risks of CHD were calculated using the Framingham, Ramathibodi–Electricity Generating Authority of Thailand (Rama‐EGAT) and Data Collection on Adverse Effects of Anti‐HIV Drugs (D:A:D) risk equations.

Results

The mean age of the patients was 41.0 years; 55% of the subjects were male. The mean duration of antiretroviral therapy was 7.7 years. The prevalence of cardiovascular risk factors was low, with the most common risk factor being low high‐density lipoprotein (HDL) (36.3%). The prevalence of high cardiovascular risk scores (defined as 10‐year risk of CHD≥10%) was also low: 9.9, 2.1 and 0.8%, by the Framingham, Rama‐EGAT and D:A:D scoring systems, respectively. Only eight subjects (1.0%) had a history of CHD. Bland–Altman plots showed that the Framingham equation predicted a higher risk of CVD compared with the Rama‐EGAT and D:A:D equations, which agreed relatively well.

Conclusion

The predicted cardiovascular risk in this HIV‐infected Thai cohort was relatively low. The agreement among the Rama‐EGAT and D:A:D risk scores suggests that both equations may be appropriate estimators of cardiovascular risk in this population.     

Astrocytes and therapeutics for Parkinson’s disease

Astrocytes play direct, active, and critical roles in mediating neuronal survival and function in various neurodegenerative disorders. This role of astrocytes is well illustrated in amyotrophic lateral sclerosis (ALS), in which the removal of glutamate from the extracellular space by astrocytes confers neuroprotection, whereas astrocytic release of soluble toxic molecules promotes neurodegeneration. In recent years, this context-dependent dual role of astrocytes has also been documented in experimental models of Parkinson’s disease. The present review addresses these studies and some potential mechanisms by which astrocytes may influence the neurodegenerative processes in Parkinson’s disease, and in particular examines how astrocytes confer neuroprotection either through the removal of toxic molecules from the extracellular space or through the release of trophic factors and antioxidant molecules. In contrast, under pathological conditions, astrocytes release proinflammatory cytokines and other toxic molecules that are detrimental to dopaminergic neurons. These emerging roles of astrocytes in the pathogenesis of Parkinson’s disease constitute an exciting development with promising novel therapeutic targets.     

MiR-378 as a biomarker for response to anti-angiogenic treatment in ovarian cancer

Objective

To determine the role of miR-378 as a biomarker for anti-angiogenic therapy response in ovarian cancer.

Methods

Expression of miR-378 was analyzed in ovarian cancer cell lines and human tumors vs. normal ovarian epithelial cells by qRT-PCR. After miR-378 transfection in SKOV3 cells, dysregulated genes were identified using microarray. Data from The Cancer Genome Atlas (TCGA) was utilized to correlate miR-378 expression with progression-free survival (PFS) among patients treated with anti-angiogenic therapy by using Kaplan–Meier and Cox proportional hazards.

Results

MiR-378 was overexpressed in ovarian cancer cells and tumors vs. normal ovarian epithelial cells. Overexpressing miR-378 in ovarian cancer cells altered expression of genes associated with angiogenesis (ALCAM, EHD1, ELK3, TLN1), apoptosis (RPN2, HIPK3), and cell cycle regulation (SWAP-70, LSM14A, RDX). In the TCGA dataset, low vs. high miR-378 expression was associated with longer PFS in a subset of patients with recurrent ovarian cancer treated with bevacizumab (9.2 vs. 4.2 months; p = 0.04). On multivariate analysis, miR-378 expression was an independent predictor for PFS after anti-angiogenic treatment (HR = 2.04, 95% CI: 1.12–3.72; p = 0.02). Furthermore, expression levels of two miR-378 targets (ALCAM and EHD1) were associated with PFS in this subgroup of patients who received anti-angiogenic therapy (9.4 vs. 4.2 months, p = 0.04 for high vs. low ALCAM; 7.9 vs. 2.3 months, p < 0.01 for low vs. high EHD1).

Conclusions

Our data suggest that miR-378 is overexpressed in ovarian cancer cells and tumors vs. normal ovarian epithelial cells. MiR-378 and its downstream targets may serve as markers for response to anti-angiogenic therapy.     

Risk factors for repeat abortion and implications for addressing unintended pregnancy in Vietnam

Objective

To determine predictors of repeat abortion in 3 provinces in Vietnam.

Methods

In a cross-sectional study between August and December 2011, women who underwent abortion were interviewed after the procedure in 62 public health facilities in Hanoi, Khanh Hoa, and Ho Chi Minh City (HCMC). Information on sociodemographic factors, contraceptive and reproductive history and intentions, and opinions and experience of abortion services was collected. The primary outcome was repeat (≥ 2) abortions.

Results

Overall, 1224 women were interviewed: 534 from Hanoi, 163 from Khanh Hoa, and 527 from HCMC. The mean age and parity of the respondents were 29 years and 1.8, respectively, and 79.6% were married. Approximately half of the respondents were not using contraception before pregnancy. The prevalence of repeat abortion was 31.7%. In multivariate models, significant predictors of repeat abortion included living in Hanoi, higher parity, age 35 years or older, and having 2 or more daughters (versus 1) or no sons (versus 1) after controlling for parity (all P < 0.05).

Conclusion

Repeat abortion remains high in Vietnam, fueled partly by inadequate contraceptive use. Son preference seems to be an important predictor of repeat abortion. Strengthening post-abortion contraceptive counseling and promoting long-acting contraceptive methods are essential to reduce repeat abortion.     

Service users’ attributes associated with the uptake of medical versus surgical abortion at public health facilities in Vietnam

Objective

To investigate the attributes of service users associated with uptake of medical abortion (MA) versus manual vacuum aspiration (MVA) at public health facilities in Vietnam.

Methods

Structured exit interviews were conducted among women who underwent termination at 62 public health facilities in Hanoi, Khanh Hoa, and Ho Chi Minh City (HCMC) between August and December 2011. Data on sociodemographic, abortion-related, and service-related factors were compared between women who underwent MVA versus MA.

Results

Overall, 1233 women completed the study survey: 541 (43.9%) from Hanoi; 163 (13.2%) from Khanh Hoa; and 529 (42.9%) from HCMC. Almost one-quarter of women (23.1%) had chosen MA. After controlling for sociodemographic factors, women living in Khanh Hoa (odds ratio [OR], 13.4; 95% confidence interval [CI], 5.3–33.8) and HCMC (OR, 5.8; 95% CI, 2.1–15.9) were more likely to have undergone MA than women in Hanoi. Older women were less likely to have undergone MA (P < 0.05), and those who had previously heard of MA were twice as likely to have undergone MA (P = 0.020).

Conclusion

Uptake of MA was lower than that of MVA and varied by province. Women in Vietnam will make their own judgment about which method to choose if they have prior knowledge of both.