EGCG通过抑制TGF-β1/Smads信号通路改善高脂高糖饮食诱导的肥胖大鼠心肌纤维化

目的研究表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)对高脂高糖饮食诱导的肥胖大鼠心肌纤维化影响,探讨EGCG能否通过TGF-β1/Smads信号通路减轻心肌纤维化的程度。方法采用长期高脂高糖饮食形成肥胖大鼠模型,给予EGCG每天灌胃,治疗4周后,称取各组大鼠体重,Masson染色检测心肌纤维化的程度。生化检测血清中空腹血糖(FBG)、甘油三酯(TG)、总胆固醇(TC)、丙二醛(MDA)和谷胱甘肽(GSH)的含量。Western blotting检测各组大鼠心脏组织中相关蛋白(CollagenⅠ、CollagenⅢ、TGF-β1、Smad2/3、p-Smad2/3、Smad7)的表达水平。结果与正常组比较,模型组大鼠体重增加,给予EGCG治疗后减轻。与正常组比较,模型组大鼠心肌纤维化程度严重,给予EGCG治疗后减轻。与正常组比较,模型组大鼠FBG、TC、TG、MDA高于正常组,给予EGCG治疗后下降。与正常组比较,模型组大鼠GSH活性低于正常组,给予EGCG治疗后升高。与正常组比较,模型组大鼠CollagenⅠ、CollagenⅢ、TGF-β1、Smad2/3、pSmad2/3蛋白表达升高,给予EGCG治疗后下降。与正常组比较,模型组大鼠Smad7蛋白表达低于正常组,给予EGCG治疗后升高。结论EGCG能够改善肥胖大鼠血糖和脂质代谢紊乱,减轻心肌纤维化程度,其机制可能与氧化应激介导的TGF-β1/Smads信号通路有关。

Effects of Fertilization and Transplantation in the Neighbouring Area on Quality of Artemisia argyi Leaf

[Objectives]To make a preliminary study of the effects of the fertilizer application and transplantation from Qichun County,Hubei Province,China,to neighbouring area on the chemical substance contents and moxa yield of Artemisia argyi leaf in Qichun(AALQ).[Methods]The content of the total flavonoids in AALQ was determined by UV-Vis spectrophotometry with apigenin as reference substance.The contents of volatile oil and tannins were determined by the methods in Chinese Pharmacopoeia(2015 edition).The moxa yield was detected by the self-developed method of grinding and sifting with high-speed pulverizer.[Results]Whether applying cake fertilizer,or compound fertilizer after applying barnyard manure,the contents of total flavonoids and tannins in AALQ were reduced in varying degrees,and the content of volatile oil was changed,which had adversely affect the accumulation of effective substances in the leaves of medicinal A.argyi as a whole.The contents of related chemical substances and moxa yield of A.argyi planted in the neighbouring Xishui County from Qichun,Hubei Province were relatively high.[Conclusions]The contents of effective substances and moxa yield of AALQ were significantly reduced as a whole when applied fertilizer in planting,especially after the application of barnyard manure,compound fertilizer was added.The quality of A.argyi leaf would not be adversely affected if it was transplanted from Qichun County,Hubei Province,which is the genuine producing area,to nearby areas with similar ecological environment.

Upconversion nanoscopy revealing surface heterogeneity of tumor-secreted extracellular vesicles

Super-resolution microscopic imaging employing upconversion nanoparticles is applied to reveal the surface heterogeneity of tumor cell-derived small extracellular vesicles,i.e.,exosome.The number of surface antigens of every extracellular vesicles can be quantified by both the high imaging resolution and stable brightness of upconversion nanoparticles.This method proves its great potential in nanoscale biological studies.

Association of miRNA-122-binding site polymorphism at the interleukin-1 a gene and its interaction with hepatitis B virus mutations with hepatocellular carcinoma risk

This study was designed to investigate the contribution of miRNA-122-binding site polymorphism at the IL-1A gene and its multiplicative interactions with hepatitis B virus （HBV） mutations in the risk of hepatocellular carcinoma （HCC）. A total of 1021 healthy controls, 302 HBV surface antigen （HBsAg） seroclearance subjects, and 2011 HBsAg-positive subjects （including 1021 HCC patients） were enrolled in this study. Quantitative PCR was used to genotype rs3783553. HBV mutations were determined by direct sequencing. Multivariate logistic regression analyses were performed to test the associations of rs3783553, mutations, and their interactions with the risk of HCC. No significant association was found between rs3783553 and the risk of HCC among healthy controls, HBsAg seroclearance subjects, HBsAg-positive subjects without HCC, and all controls. Additionally, rs3783553 was not significantly associated with chronic HBV infection, liver cirrhosis, HBV e antigen seroconversion, abnormal alanine aminotransferase, and high viral load （ 〉 10^4 copies/ml）. However, the TTCA insertion allele of rs3783553 was significantly associated with an increased frequency of HBV C7A mutation compared with homozygous TTCA deletion carriers [（del/ins ＋ ins/ins） vs. del/del, adjusted odds ratio （OR）= 1.48, 95% confidence interval （CI）= 1.09-2.02, P = 0.013]. Multiplicative interaction of rs3783553 with HBV preS deletion significantly reduced the risk of HCC in males, with an adjusted OR of 0.64 （95% CI = 0.42-0.98; P = 0.041） after age and HBV genotype were adjusted. Although rs3783553 did not significantly affect genetic susceptibility to HBV-related HCC, its variant allele may predispose the host to selecting HBV C7A mutation during evolution and significantly reduce the risk of HCC caused by HBV preS deletion. This study provides an insight into the complex host-virus interaction in HBV-induced hepatocarcinogenesis and is helpful in determining HBsAg-positive subjects who are likely to develop HCC.

Phosphatase and Tensin Homologue Genetic Polymorphisms and their Interactions with Viral Mutations on the Risk of Hepatocellular Carcinoma

Background： Chronic hepatitis B virus （HBV） infection is the major cause of hepatocellular carcinoma （HCC）. Some HBV mutants and dysregulation of phosphatase and tensin homolog （PTEN） may promote the development of HCC synergistically. We aimed to test the effects of PTEN genetic polymorphisms and their interactions with important HBV mutations on the development of HCC in HBV-infected subjects. Methods： Quantitative po[ymerase chain reaction was applied to genotype PTEN polymorphisms （rs1234220, rs2299939, rsl234213） in 1012 healthy controls, 302 natural clearance subjects, and 2011 chronic H BV-infected subjects including 1021 HCC patients. HBV mutations were determined by sequencing. The associations of PTEN polymorphisms and their interactions with HBV mutations with HCC risk were assessed using multivariate logistic regression analysis. Results： Rs1234220 C allele was significantly associated with HCC risk compared to healthy controls （adjusted odds ratio [AOR] = 1.35, 95% confidence interval [CI] = 1,07-1.69） and HCC-free HBV-infected subjects （AOR = 1.27, 95% CI = 1.01-1 .57）. rsl234220 C allele was significantly associated with increased frequencies of HCC-risk A 1652G, C 1673T, and C 1730G mutations in genotype B H BV-in fected subjects. Rs2299939 GT genotype was inversely associated with HCC risk in HBV-infected patients （AOR 0.75, 95% CI 0.62-0.92）. The interaction of rs2299939 variant genotypes （GT＋TT） with A3054T mutation significantly increased HCC risk （AOR = 2.41, 95% CI = 1.08-5.35）; whereas its interaction with C3116T mutation significantly reduced HCC risk （AOR = 0.34, 95% CI 0.18-0.66）. These significant effects were only evident in males alter stratification. Conclusions： PTEN polymorphisms and their interactions with HBV mutations may contribute to hepatocarcinogenesis in males. The host-virus interactions are important in identifying HBV-infected subjects who are more likely to develop HCC.

Increased circulating level of interleukin-6 and CD8+T cell exhaustion are associated with progression of COVID-19

Background: Coronavirus disease 2019(COVID-19)is pandemic.It is critical to identify COVID-19 patients who are most likely to develop a severe disease.This study was designed to determine the clinical and epidemiological features of COVID-19 patients associated with the development of pneumonia and factors associated with disease progression.Methods:: Seventy consecutive patients with etiologically confirmed COVID-19 admitted to PLA General Hospital in Beijing,China from December 27,2019 to March 12,2020 were enrolled in this study and followed-up to March 16,2020.Differences in clinical and laboratory findings between COVID-19 patients with pneumonia and those without were determined by theχ2 test or the Fisher exact test(categorical variables)and independent group t test or Mann–Whitney U test(continuous variables).The Cox proportional hazard model and Generalized Estimating Equations were applied to evaluate factors that predicted the progression of COVID-19.Results: The mean incubation was 8.67(95%confidence interval,6.78–10.56)days.Mean duration from the first test severe acute respiratory syndrome coronavirus 2-positive to conversion was 11.38(9.86–12.90)days.Compared to pneumonia-free patients,pneumonia patients were 16.5 years older and had higher frequencies of having hypertension,fever,and cough and higher circulating levels of neutrophil proportion,interleukin-6,low count(<190/µl)of CD8+T cells,and neutrophil/lymphocyte ratio.Thirteen patients deteriorated during hospitalization.Cox regression analysis indicated that older age and higher serum levels of interleukin-6,C-reactive protein,procalcitonin,and lactate at admission significantly predicted the progression of COVID-19.During hospitalization,circulating counts of T lymphocytes,CD4+T cells,and CD8+T cells were lower,whereas neutrophil proportion,neutrophil/lymphocyte ratio,and the circulating levels of interleukin-6,C-reactive protein,and procalcitonin were higher,in pneumonia patients than in pneumonia-free patients.CD8+lymphocyte count in pneumonia patients did not recover when discharged.Conclusions: Older age and higher levels of C-reactive protein,procalcitionin,interleukin-6,and lactate might predict COVID-19 progression.T lymphocyte,especially CD8+cell-mediated immunity is critical in recovery of COVID-19.This study may help in predicting disease progression and designing immunotherapy for COVID-19.

Characterization of SARS-CoV-2 worldwide transmission based on evolutionary dynamics and specific viral mutations in the spike protein

Background:The coronavirus disease 2019(COVID-19)caused by severe acute respiratory syndrome-related coronavirus-2(SARS-CoV-2)is pandemic.However,the origins and global transmission pattern of SARS-CoV-2 remain largely unknown.We aimed to characterize the origination and transmission of SARS-CoV-2 based on evolutionary dynamics.Methods:Using the full-length sequences of SARS-CoV-2 with intact geographic,demographic,and temporal information worldwide from the GISAID database during 26 December 2019 and 30 November 2020,we constructed the transmission tree to depict the evolutionary process by the R package"outbreaker".The affinity of the mutated receptor-binding region of the spike protein to angiotensin-converting enzyme 2(ACE2)was predicted using mCSM-PPI2 software.Viral infectivity and antigenicity were tested in ACE2-transfected HEK293T cells by pseudovirus transfection and neutralizing antibody test.Results:From 26 December 2019 to 8 March 2020,early stage of the COVID-19 pandemic,SARS-CoV-2 strains identified worldwide were mainly composed of three clusters:the Europe-based cluster including two USA-based subclusters;the Asia-based cluster including isolates in China,Japan,the USA,Singapore,Australia,Malaysia,and Italy;and the USA-based cluster.The SARS-CoV-2 strains identified in the USA formed four independent clades while those identified in China formed one clade.After 8 March 2020,the clusters of SARS-CoV-2 strains tended to be independent and became"pure"in each of the major countries.Twenty-two of 60 mutations in the receptor-binding domain of the spike protein were predicted to increase the binding affinity of SARS-CoV-2 to ACE2.Of all predicted mutants,the number of E484K was the largest one with 86585 sequences,followed by S477N with 55442 sequences worldwide.In more than ten countries,the frequencies of the isolates with E484K and S477N increased significantly.V367F and N354D mutations increased the infectivity of SARS-CoV-2 pseudoviruses(P<0.001).SARS-CoV-2 with V367F was more sensitive to the S1-targeting neutralizing antibody than the wild-type counterpart(P<0.001).Conclusions:SARS-CoV-2 strains might have originated in several countries simultaneously under certain evolutionary pressure.Travel restrictions might cause location-specific SARS-CoV-2 clustering.The SARS-CoV-2 evolution appears to facilitate its transmission via altering the affinity to ACE2 or immune evasion.

Association of human leukocyte antigen-DR-DQ-DP haplotypes with the risk of hepatitis B virus-related hepatocellular carcinoma

Aim:Genetic polymorphisms of human leukocyte antigen(HLA)class II molecules are associated with chronic hepatitis B virus(HBV)infection.We aimed to investigate the impacts of HLA-II haplotypes on viral evolution and the risks of HBV-caused liver diseases.Methods:HLA-DR-DQ-DP haplotypes were estimated in 1210 healthy controls,296 HBV clearance subjects,301 asymptomatic hepatitis B surface antigen carriers,770 chronic hepatitis B patients,443 HBV-related liver cirrhosis(LC)patients,and 1037 HBV-related hepatocellular carcinoma(HCC)patients.HBV mutations were determined by sequencing.The associations of HLA-DR-DQ-DP haplotypes with viral mutations and the risks of liver diseases were assessed by multivariate logistic regression.Results:Compared to HBV-free subjects,the haplotypes CCAACG,CCGACG,TCAATA,and TCGATA were associated with decreased HCC risk,with an odds ratio(OR)[95%confidence interval(CI)]of 0.62(0.40-0.95),0.60(0.39-0.92),0.73(0.54-0.98),and 0.58(0.42-0.78),respectively.CCAACG,CCGACG,and TCAATA were significantly associated with decreased frequencies of the HCC-risk HBV mutations:preS1 deletion,APOBECsignature HBV mutations in the core promoter and preS regions,A51C/T,G104C/T,and G146C/T.TCGATA and TTAACG were associated with increased LC risk,with an OR(95%CI)of 1.54(1.03-2.30)and 2.23(1.50-3.33),respectively.However,TCGATA and TTAACG were not consistently associated with the cirrhosis-risk HBV mutations.Conclusion:CCAACG,CCGACG,and TCAATA are inversely associated with HCC risk,possibly because they are involved in creating an immune microenvironment attenuating the generation of HCC-risk HBV mutations.TCGATA and TTAACG might predispose the polarity of immunity towards Th17 isotype related to LC.

Association of novel mutations and heplotypes in the preSregion of hepatitis B virus with hepatocellular carcinoma

The association of viral mutations and haplo-typic carriages with mutations in the preS region of hepatitis B virus(HBV)genotypes B and C with hepatocellular carcinoma(HCC)is of great significance for the prediction of this malignancy,but it remains obscure.We analyzed the preS sequences of HBV genotypes B and C from 1172 HBV-infected subjects including 231 patients with HCC.As compared with the HBV-infected subjects without HCC,C2875T,G2946C,A3054C,C3060A,T3066C,C3116T,A3120C,G3191A,A1C,C7A,C10A,A31C,C76T,G105C,and G147C in both genotypes were significantly associated with increased risks of HCC.C2875A,G2950A,G2951A,A3054T,C3060T,T3066A,T3069G,A3120T,and G3191C were significantly associated with increased risks of HCC in genotype C,whereas these mutations were inversely associated with HCC in genotype B.Multivariate regression analyses showed that C76A/T was a novel factor independently associated with an increased risk of HCC,as compared with those without HCC.The frequencies of haplotypes 2964A-3116T-preS2 start codon wild-type-7C,2964C-3116T-7A-76C,and 2964A-3116T-7C-76A/T were significantly higher in the patients with HCC(P<0.001),whereas a haplotypic carriage with a single mutation and another three wild-types were inversely associated with HCC.Conclusively,the association of HBV mutations in the preS region with HCC depends on HBV genotype and haplotypic carriage with two or more mutations that are each associated with an increased risk of HCC independently.