Functional dissection of the spike glycoprotein S1 subunit and identification of cellular cofactors for regulation of swine acute diarrhea syndrome coronavirus entry.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a novel porcine enteric coronavirus, and the broad interspecies infection of SADS-CoV poses a potential threat to human health. This study provides experimental evidence to dissect the roles of distinct domains within the SADS-CoV spike S1 subunit in cellular entry. Specifically, we expressed the S1 and its subdomains, S1A and S1B. Cell binding and invasion inhibition assays revealed a preference for the S1B subdomain in binding to the receptors on the cell surface, and this unknown receptor is not utilized by the porcine epidemic diarrhea virus. Nanoparticle display demonstrated hemagglutination of erythrocytes from pigs, humans, and mice, linking the S1A subdomain to the binding of sialic acid (Sia) involved in virus attachment. We successfully rescued GFP-labeled SADS-CoV (rSADS-GFP) from a recombinant cDNA clone to track viral infection. Antisera raised against S1, S1A, or S1B contained highly potent neutralizing antibodies, with anti-S1B showing better efficiency in neutralizing rSADS-GFP infection compared to anti-S1A. Furthermore, depletion of heparan sulfate (HS) by heparinase treatment or pre-incubation of rSADS-GFP with HS or constituent monosaccharides could inhibit SADS-CoV entry. Finally, we demonstrated that active furin cleavage of S glycoprotein and the presence of type II transmembrane serine protease (TMPRSS2) are essential for SADS-CoV infection. These combined observations suggest that the wide cell tropism of SADS-CoV may be related to the distribution of Sia or HS on the cell surface, whereas the S1B contains the main protein receptor binding site. Specific host proteases also play important roles in facilitating SADS-CoV entry.IMPORTANCESwine acute diarrhea syndrome coronavirus (SADS-CoV) is a novel pathogen infecting piglet, and its unique genetic evolution characteristics and broad species tropism suggest the potential for cross-species transmission. The virus enters cells through its spike (S) glycoprotein. In this study, we identify the receptor binding domain on the C-terminal part of the S1 subunit (S1B) of SADS-CoV, whereas the sugar-binding domain located at the S1 N-terminal part of S1 (S1A). Sialic acid, heparan sulfate, and specific host proteases play essential roles in viral attachment and entry. The dissection of SADS-CoV S1 subunit's functional domains and identification of cellular entry cofactors will help to explore the receptors used by SADS-CoV, which may contribute to exploring the mechanisms behind cross-species transmission and host tropism.

Analysis of X-ray diffraction fingerprint of Calamina and its processed products / 中草药

Objective: To establish X-ray diffraction (XRD) fingerprint of Calamina and its processed products, compare the effects of different processing Methods on the main components of medicinal materials and determine the content of ZnO in the processed products. Methods: XRD was used to analyze 10 batches of Calamina and its processed products, and fingerprints of Calamina and its processed products were established respectively. Six different processing methods were compared, and the content of ZnO in all processed products was determined by K value method. Results: Fingerprints of Calamina and its processed products were preliminarily established. There were 23 common peaks in the fingerprints of Calamina, and there were 10 common peaks in the fingerprints of its processed products. After calcination, the ZnCO3 characteristic peak of the raw material was transformed into the characteristic peak of ZnO; The content of ZnO in the calcined product exceeded 56%. Conclusion: XRD fingerprints could be used for the identification and analysis of Calamina and its processed products. The new and reliable method was provided for quality evaluation of Calamina and its processed products.

[Mechanisms of tanshinone â ¡_A in reducing 4-HNE-induced hepatocyte damage by activating PPARα].

Tanshinone Ⅱ_A( Tan Ⅱ_A),the liposoluble constituents of Salvia miltiorrhiza,can not only ameliorate the lipidic metabolism and decrease the concentration of lipid peroxidation,but also resist oxidation damage,scavenge free radicals and control inflammation,with a protective effect on prognosis after liver function impairment. Therefore,the studies on the exact mechanism of Tan Ⅱ_A in protecting the liver can provide important theoretical and experimental basis for the prevention and treatment effect of Tan Ⅱ_A for liver injury. In the present study,the protective effects and mechanism of Tan Ⅱ_A on 4-hydroxynonenal( 4-HNE)-induced liver injury were investigated in vitro. Normal liver tissues NCTC 1469 cells were used to induce hepatocytes oxidative damages by 4-HNE treatment. The protective effect of Tan Ⅱ_A on hepatocytes oxidative damages was detected by release amount of lactate dehydrogenase( LDH) analysis and hoechst staining. The protein expression changes of peroxisome proliferator-activated receptor α( PPARα) and peroxisome proliferator response element( PPRE) were analyzed by Western blot analysis in NCTC 1469 cells before and after Tan Ⅱ_A treatment. The gene expression changes of fatty aldehyde dehydrogenase( FALDH) were analyzed by Real-time polymerase chain reaction( PCR) analysis. The results showed that 4-HNE increased the release amount of LDH,lowered the cell viability of NCTC 1469 cells,and Tan Ⅱ_A reversed 4-HNE-induced hepatocyte damage. Western blot analysis and RT-PCR analysis results showed that 4-HNE decreased the expression of PPARα and FALDH and increased the expression of 4-HNE. However,the expression of PPARα and FALDH were increased significantly and the expression of 4-HNE was decreased obviously after Tan Ⅱ_A treatment. This study confirmed that the curative effect of Tan Ⅱ_A was obvious on hepatocytes damage,and the mechanism may be associated with activating PPARα and FALDH expression as well as scavenging 4-HNE.

Effect of different sandblasting conditions on the metal-ceramic bonding strength of Co-Cr alloy fabricated by selective laser melting technology / 华西口腔医学杂志

OBJECTIVE@#To investigate the effect of different sandblasting conditions on the metal-ceramic bonding strength of Co-Cr alloy fabricated by selective laser melting (SLM) technology.@*METHODS@#A total of 63 specimens of Co-Cr alloy fabricated by SLM were prepared and randomly divided into nine groups (n=7). Each group was treated with different powder particles (A1=50 µm, A2=100 µm, and A3=150 µm) and pressures (B1=0.2 MPa, B2=0.4 MPa, and B3=0.6 MPa) in sandblasting. One sample was randomly selected from each group for microstructure observation by scanning electron microscope (SEM). Ceramic was fired at the center of the specimens. Metal-ceramic bonding strength was measured with universal testing machine. Results were statistically analyzed with SPSS 17.0 software.@*RESULTS@#The mean bond strengths were as follows: Group A1B1: 27.22 MPa±0.95 MPa, Group A1B2: 27.58 MPa±0.47 MPa, Group A1B3: 26.80 MPa±0.71 MPa, Group A2B1: 27.54 MPa±0.78 MPa, Group A2B2: 30.75 MPa±0.43 MPa, Group A2B3: 26.93 MPa±0.88 MPa, Group A3B1: 28.18 MPa±0.93 MPa, Group A3B2: 29.55 MPa±0.57 MPa, and Group A3B3: 28.11 MPa±0.91 MPa. The particle factor of Al₂O₃ and the pressure factor of blasting showed statistical significance (P<0.05). An interaction was observed between the factors of particle and pressure (P<0.05). Mixed fracture mode of all specimens was observed after the shear strength test.@*CONCLUSIONS@#In conclusion, metal-ceramic bonding strength reaches the maximum when specimens are sandblasted with 100 µm alumina oxide at 0.4 MPa pressure.

Genetic variants in CHEK1 gene are associated with the prognosis of thoracic esophageal squamous cell carcinoma patients treated with radical resection / 华中科技大学学报（医学）（英德文版）

CHEK1 gene is known to play an important role in tumor progression by cell cycle control. However, the association between CHEK1 and the prognosis of esophageal squamous cell carcinoma (ESCC) is unclear. In this study, we explored the association between genetic variants in CHEK1 gene and prognosis of ESCC patients treated with radical resection. A total of 131 thoracic ESCC patients who underwent radical resection were included in this retrospective study and genotyped using the MassArray method. According to the univariate Cox hazard analysis, the GT/TT genotype of CHEK1 rs555752 was shown to be strongly related to a decreased overall survival (OS) (HR=2.560, 95% CI: 1.415-4.631, P=0.002) and disease-free survival (DFS) (HR=2.160, 95% CI: 1.258-3.710, P=0.005). Furthermore, according to the multivariate Cox hazard analysis and multiple testing, patients with the GT/TT genotype of CHEK1 rs555752 had a notably decreased OS (HR=2.735, 95% CI: 1.468-5.096, P=0.002, Pc=0.006) and DFS (HR=2.282, 95% CI: 1.292-4.023, P=0.004, Pc=0.012). In conclusion, genetic variants of the CHEK1 gene are significantly related to OS and DFS of ESCC patients, and may therefore be predictors of the prognosis of thoracic ESCC after surgery.

Nonsynonymous single nucleotide polymorphisms in the complement component 3 gene are associated with risk of age-related macular degeneration: a meta-analysis.

Nonsynonymous single nucleotide polymorphisms (SNPs) in complement component 3 (CC3) are associated with the risk of age-related macular degeneration (AMD), however, this association is not consistent among studies. To thoroughly address this issue, we performed an updated meta-analysis to evaluate the association between nine SNPs in the CC3 gene and AMD risk. A search was conducted of the PubMed database through 3rd Aug, 2014. Odds ratios (ORs) and 95% confidence intervals (CIs) were used to assess the strength of associations. Based on the search criteria for manuscripts reporting AMD susceptibility related to CC3 in nine SNPs, 57 case-control studies from 22 different articles were retrieved. Significantly positive associations were found for the rs2230199 C/G SNP and AMD in the Caucasian population, as well as for the rs1047286 C/T SNP. Moreover, a relationship between the rs11569536 G/A SNP and AMD was detected. By contrast, a negative association was observed between rs2250656 A/G SNP and AMD risk. The present meta-analysis suggests that these four SNPs in the CC3 gene are potentially associated with the risk of AMD development. Further studies using larger sample sizes and accounting for gene-environment interactions should be conducted to elucidate the role of CC3 gene polymorphisms in AMD risk.

Association between a 27-bp variable number of tandem repeat polymorphism in intron 4 of the eNOS gene and risk for diabetic retinopathy Type 2 diabetes mellitus: a meta-analysis.

PURPOSE: Imbalance in the production of endothelial nitric oxide synthase (eNOS), which plays an essential role in retinal vascular function, can lead to the development of diabetic retinopathy (DR). To thoroughly address this issue, we performed an updated meta-analysis to evaluate the association between the eNOS 27VNTR (4b/4a) polymorphism and DR in type 2 diabetes mellitus (T2DM). METHODS: A search was conducted of PubMed and Chinese language (WanFang) databases through 3 March 2013. Data were retrieved in a systematic manner and analyzed using Stata Statistical Software. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were used to assess the strength of associations. RESULTS: Based on the search criteria for DR susceptibility related to the 27VNTR (4b/4a) polymorphism of the eNOS gene, 16 case-control studies (15 articles), comprising 3227 T2DM patients with DR and 3437 T2DM patients without DR, were retrieved. Although no significant associations were uncovered in either the overall analysis or DR subtype groups, a decreased association was detected between the African- (allelic contrast: OR = 0.75, 95% CI = 0.65-0.88) or population-based (PB) studies (dominant genetic model: OR = 0.91, 95% CI = 0.83-0.98) and the eNOS 27VNTR (4b/4a) polymorphism. Stratification according to average duration of DM revealed that T2DM patients with histories of >10 years had an elevated susceptibility to DR compared with those with histories of shorter durations (homozygote comparison: OR = 1.67, 95% CI = 1.09-2.58). CONCLUSIONS: The present meta-analysis suggests that the eNOS 27VNTR (4b/4a) polymorphism potentially decreases the risk of developing DR in T2DM African individuals. The higher degree of susceptibility in patients with longer (>10 years) durations of DM is indicative of the involvement of a gene-environment interaction in determining the risk for DR. Further studies, based on larger sample sizes and additional gene-environment interactions, should be conducted to elucidate the role of eNOS gene polymorphisms, especially 27VNTR (4b/4a), in the risk for DR.

Difference analysis of proteome between diabetic cataract and age related cataract / 中华实验眼科杂志

Background With the changes of diet and living style,the diabetes has become the major diseases affecting human health.Diabetic cataract is a common complication of diabetes. Objective The present study was to investigate the difference of lens proteomics between diabetic cataract and age related cataract using two dimensional electrophoresis (2-DE) and mass spectrometry in order to postpone happening of diabetic cataract and offer the effective approach to the prevention and therapy of diabetic cataract. Methods The lenses were obtained from 8 diabetic patients and 12 age-related cataract patients during the surgery to extract the protein by lysis and centrifugation.The lens proteins were separated using immobilized pH gradients 2-DE.Image analysis was carried out using PDQuest Advanced-8.0.1 software package.Significant difference of the crystallines was identified by matrixassisted laser adsorption/ionization time of-flight-mass spectrometry (MALDI-TOF-MS) and peptide mass fingerprint combined with protein database. Results The maps of 2-DE showed that lens proteins of diabetic cataract and age related cataract were in the section of pH 5-9 with the relative molecular weight 14000-97000;while relative molecular weight of more abundant crystalline was localized at 20000-31000.About 3 differential protein spots were detected by image analysis software.Two crystallines,αB and βB1 crystallin,were identified using MALDI-TOF-MS.Conclusions Proteomic analysis of lens can be accomplished and the proteins can be well separated,moreover,differential proteins can be analyzed using 2-DE and mass spectrometry between diabetic cataract and age related cataract.These results indicate that αB and βB1 crystallin proteins accelerate the development of diabetic cataract.This technique offers a new avenue for clarity of lens proteins of diabetic cataract other than age related cataract.