Plasma heme oxygenase-1 concentration in relation to impaired glucose regulation in a non-diabetic Chinese population

Background

Our previous study has recently shown that plasma heme oxygenase-1 (HO-1), a stress-responsive protein, is elevated in individuals with type 2 diabetes. The current study aimed to examine the association between plasma HO-1 concentration and impaired glucose regulation (IGR) in non-diabetic individuals.

Methods

We conducted a case-control study including a total of 865 subjects (262 IGR individuals and 603 healthy controls) in a Chinese population. Basic characteristics were collected by questionnaire and standardized anthropometric measurements. Plasma HO-1 concentration was determined by ELISA.

Results

Plasma HO-1 concentration was significantly increased in IGR individuals compared with healthy controls (1.34 (0.81–2.29) ng/ml vs 0.98 (0.56–1.55) ng/ml, P<0.001). After adjustment for age, sex, and BMI, the ORs for IGR in the highest quartile of plasma HO-1 concentrations, compared with the lowest, was 3.42 (95% CI 2.11–5.54; P for trend <0.001). The trend remained significant even after additional adjustment for smoking, alcohol drinking, hypertension, family history of diabetes, lipid profiles and C-reactive protein. In the receiver-operating characteristic curve analysis, addition of plasma HO-1 concentration to a model with known risk factors yielded significantly improved discriminative value for IGR (area under the curves 0.75 (95% CI 0.71–0.78) vs. 0.72 (95% CI 0.69–0.76); P for difference = 0.026).

Conclusions

Elevated plasma HO-1 concentration is significantly associated with increased ORs for IGR. However, its clinical utility should be validated in further studies, especially in prospective cohort studies.     

Natural Killer Cells Are Characterized by the Concomitantly Increased Interferon-γ and Cytotoxicity in Acute Resolved Hepatitis B Patients

Natural killer (NK) cells are abundant in the liver and have been implicated in inducing hepatocellular damage in patients with chronic hepatitis B virus (HBV) infection. However, the role of NK cells in acute HBV infection remains to be elucidated. We comprehensively characterized NK cells and investigated their roles in HBV clearance and liver pathology in 19 chronic hepatitis B (CHB) patients and 21 acute hepatitis B (AHB) patients as well as 16 healthy subjects. It was found that NKp46⁺ NK cells were enriched in the livers of AHB and CHB patients. We further found that peripheral NK cells from AHB patients expressed higher levels of activation receptors and lower levels of inhibitory receptors than those from CHB patients and HC subjects, thus displaying the increased cytolytic activity and interferon-γ production. NK cell activation levels were also correlated positively with serum alanine aminotransferase levels and negatively with plasma HBV DNA levels in AHB patients, which is further confirmed by the longitudinal follow-up of AHB patients. Serum pro-inflammatory cytokine and chemokine levels were also increased in AHB patients as compared with CHB and HC subjects. Thus, the concomitantly increased interferon-γ and cytotoxicity of NK cells were associated with liver injury and viral clearance in AHB patients.     

The effect of chronic antipsychotic drug on hypothalamic expression of neural nitric oxide synthase and dopamine D2 receptor in the male rat

Antipsychotic-induced sexual dysfunction is a common and serious clinical side effect. It has been demonstrated that both neuronal nitric oxide (nNOS) and dopamine D2 receptor (DRD2) in the medial preoptic area (MPOA) and the paraventricular nucleus (PVN) of the hypothalamus have important roles in the regulation of sexual behaviour. We investigated the influences of 21 days' antipsychotic drug administration on expression of nNOS and DRD2 in the rat hypothalamus. Haloperidol (0.5 mg/kg/day i.p.) significantly decreased nNOS integrated optical density in a sub-nucleus of the MPOA, medial preoptic nucleus (MPN), and decreased the nNOS integrated optical density and cell density in another sub-nucleus of the MPOA, anterodorsal preoptic nucleus (ADP). Risperidone (0.25 mg/kg) inhibited the nNOS integrated optical density in the ADP. nNOS mRNA and protein in the MPOA but not the PVN was also significantly decreased by haloperidol. Haloperidol and risperidone increased DRD2 mRNA and protein expression in both the MPOA and the PVN. Quetiapine (20 mg/kg/day i.p.) did not influence the expression of nNOS and DRD2 in either the MPOA or the PVN. These findings indicate that hypothalamic nNOS and DRD2 are affected to different extents by chronic administration of risperidone and haloperidol, but are unaffected by quetiapine. These central effects might play a role in sexual dysfunction induced by certain antipsychotic drugs.     

Disease-associated mutations prevent GPR56-collagen III interaction

GPR56 is a member of the adhesion G protein-coupled receptor (GPCR) family. Mutations in GPR56 cause a devastating human brain malformation called bilateral frontoparietal polymicrogyria (BFPP). Using the N-terminal fragment of GPR56 (GPR56(N)) as a probe, we have recently demonstrated that collagen III is the ligand of GPR56 in the developing brain. In this report, we discover a new functional domain in GPR56(N), the ligand binding domain. This domain contains four disease-associated mutations and two N-glycosylation sites. Our study reveals that although glycosylation is not required for ligand binding, each of the four disease-associated mutations completely abolish the ligand binding ability of GPR56. Our data indicates that these four single missense mutations cause BFPP mostly by abolishing the ability of GPR56 to bind to its ligand, collagen III, in addition to affecting GPR56 protein surface expression as previously shown.     

Identification of in vivo protein phosphorylation sites in human pathogen Schistosoma japonicum by a phosphoproteomic approach

Schistosome is the causative agent of human schistosomiasis and related animal disease. Reversible protein phosphorylation plays a key role in signaling processing that are vital for a cell and organism. However, it remains to be undercharacterized in schistosomes. In the present study, we characterized in vivo protein phosphorylation events in different developmental stages (schistosomula and adult worms) of Schistosoma japonicum by using microvolume immobilized metal-ion affinity chromatography (IMAC) pipette tips coupled to nanoLC-ESI-MS/MS. In total, 127 distinct phosphorylation sites were identified in 92 proteins in S. japonicum. A comparison of the phosphopeptides identified between the schistosomula and the adult worms revealed 30 phosphoproteins co-detected in both of the two worms. These proteins included several signal molecules and enzymes such as 14-3-3 protein, cysteine string protein, heat shock protein 90, epidermal growth factor receptor pathway substrate 8, proliferation-associated protein 2G4, peptidyl-prolyl isomerase G, phosphofructokinase and thymidylate kinase. Additionally, the phosphorylation sites were examined for phosphorylation specific motif and evolutionarily conservation. The study represents the first attempt to determine in vivo protein phosphorylation in S. japonicum by using a phosphoproteomic approach. The results by providing an inventory of phosphorylated proteins may facilitate to further understand the mechanisms involved in schistosome development and growth, and then may result in the development of novel vaccine candidates and drug targets for schistosomiasis control.     

Microbial transformation of the triterpene nigranoic acid in Trichoderma sp.

Two new metabolites were obtained by microbial transformation of the triterpene nigranoic acid (3,4-secocyloarta-4 (28), 24 (Z)-diene-3,26-dioic acid), (1) in the culture of Trichoderma sp. JY-1, a fungus obtained from the branches of Kadsura angustifolia. Their structures were established as 15α, 16α-dihydroxy-3,4-secocyloarta-4 (28), 17 (20), 17 (E), 24 (E)-triene-3,26-dioic acid (2) and 16α, 20α-dihydroxy-18 (13 → 17β) abeo-3,4-secocyloarta-4 (28), 12 (13), 24 (Z)-triene-3,26-dioic acid (3) by analysis of NMR and MS data and by analogy with the data for the substrate nigranoic acid (1). Compound 2 was found to possess an unusual 17(20), 17 (E)-ene structure while compound 3 featured an unprecedented 18(13 → 17β)-abeo-secocyloarta skeleton. Additionally, compounds 1–3 showed weak anti-HIV activity with EC₅₀ values of 10.5, 8.8 and 7.6 μg/mL, therapeutic index values (CC₅₀/EC₅₀) of 8.48, 9.12 and 10.1, respectively.     

Androgen metabolism and JAK/STAT pathway genes and prostate cancer risk

Background: Prostate cancer (PC) is the most frequently diagnosed solid tumor in U.S. men. Genome-wide association studies (GWAS) have identified over 40 risk-associated single nucleotide polymorphisms (SNPs), including variants in androgen pathway genes (e.g., KLK3 and AR). Androgens are important in PC and genes involved in this pathway are therefore candidates for conferring susceptibility to PC. Methods: In this hypothesis-testing study, we evaluated PC risk in association with SNPs in 22 candidate genes involved in androgen metabolism or interactions with the androgen receptor (AR). A total of 187 SNPs were genotyped in 1458 cases and 1351 age-matched controls from a population-based study. PC risk was estimated using adjusted unconditional logistic regression and multinomial regression models. Results: Single SNP analyses showed evidence (p < 0.05) for associations with 14 SNPs in 9 genes: NKX3.1, HSD17B3, AKR1C3, SULT2A1, CYP17A1, KLK3, JAK2, NCOA4 and STAT3. The most significant result was observed for rs2253502 in HSD17B3 (odds ratio, OR = 0.57, 95% CI: 0.39–0.84). In addition, five SNPs in four genes (CYP17A1, HSD17B4, NCOA4, and SULT2A1) were associated with more aggressive disease (p < 0.01). Conclusions: Our results replicate previously reported associations for SNPs in CYP17A1, HSD17B3, ARK1C3, NKX3.1, NCOA4 and KLK3. In addition, novel associations were observed for SNPs in JAK2, HSD17B4, and SULT2A1. These results will require replication in larger studies.     

Discovery of multiple IGS haplotypes within genotypes of Puccinia striiformis

In a search for specific molecular markers for population analysis of Puccinia striiformis f. sp. tritici, the ribosomal DNA (rDNA) intergenic spacer (IGS) 1 region (rDNA-IGS1, between the 28S and the 5S rDNA genes) was amplified, cloned, and sequenced. It was found to exhibit multiple bands and length polymorphism. Surprisingly, single isolates were found to possess between three to five different IGS1 haplotypes. Bands were cloned and sequenced, and two highly variable regions (α and β) were found between conserved regions, with repeat units interspersed in both types of regions. There were 14 different repeat units, and these were sometimes grouped further into four combinations of repeat units, with a few individual nucleotides (A or C) inserted between the repeats. Among three geographically dispersed isolates, the variable region α was divided into eight types, and the variable region β was divided into two types based on repeat units. Most of the 14 repeat units were shared by the variable and the conserved regions. Among the three isolates, there were a total of 12 IGS1 haplotypes, but some of these were shared between isolates such that there were only eight unique haplotypes. The occurrence of multiple haplotypes within single isolates may be useful for analyzing the population structure, tracking the origin of annual epidemics and providing insights into evolutionary biology of this pathogen.     

Enhanced thermostability of a Rhizopus chinensis lipase by in vivo recombination in Pichia pastoris

ABSTRACT: BACKGROUND: Lipase from Rhizopus chinensis is a versatile biocatalyst for various bioconversions and has been expressed at high-level in Pichia pastoris. However, the use of R. chinensis lipase in industrial applications is restricted by its low thermostability. Directed evolution has been proven to be a powerful and efficient protein engineering tool for improvement of biocatalysts. The present work describes improvement of the thermostability of R. chinensis lipase by directed evolution using P. pastoris as the host. RESULTS: An efficient, fast and highly simplified method was developed to create a mutant gene library in P. pastoris based on in vivo recombination, whose recombination efficiency could reach 2.3 x 105 /mug DNA. The thermostability of r27RCL was improved significantly by two rounds of error-prone PCR and two rounds of DNA shuffling in P. pastoris. The S4-3 variant was found to be the most thermostable lipase, under the conditions tested. Compared with the parent, the optimum temperature of S4-3 was two degrees higher, Tm was 22 degrees higher and half-lives at 60degreesC and 65degreesC were 46- and 23- times longer. Moreover, the catalytic efficiency kcat/Km of S4-3 was comparable to the parent. Stabilizing mutations probably increased thermostability by increasing the hydrophilicity and polarity of the protein surface and creating hydrophobic contacts inside the protein. CONCLUSIONS: P. pastoris was shown to be a valuable cell factory to improve thermostability of enzymes by directed evolution and it also could be used for improving other properties of enzymes. In this study, by using P. pastoris as a host to build mutant pool, we succeeded in obtaining a thermostable variant S4-3 without compromising enzyme activity and making it a highly promising candidate for future applications at high temperatures.