Irritable bowel syndrome is a gastrointestinal disorder of unknown etiology characterized by widespread, chronic abdominal pain associated with altered bowel movements. Increasing amounts of evidence indicate that injury and inflammation during the neonatal period have long-term effects on tissue structure and function in the adult that may predispose to gastrointestinal diseases. In this study we aimed to investigate how the epigenetic regulation of DNA demethylation of the p2x7r locus guided by the transcription factor GATA binding protein 1 (GATA1) in spinal astrocytes affects chronic visceral pain in adult rats with neonatal colonic inflammation (NCI). The spinal GATA1 targeting to DNA demethylation of p2x7r locus in these rats was assessed by assessing GATA1 function with luciferase assay, chromatin immunoprecipitation, patch clamp, and interference in vitro and in vivo. In addition, a decoy oligodeoxynucleotide was designed and applied to determine the influence of GATA1 on the DNA methylation of a p2x7r CpG island. We showed that NCI caused the induction of GATA1, Ten-eleven translocation 3 (TET3), and purinergic receptors (P2X7Rs) in astrocytes of the spinal dorsal horn, and demonstrated that inhibiting these molecules markedly increased the pain threshold, inhibited the activation of astrocytes, and decreased the spinal sEPSC frequency. NCI also markedly demethylated the p2x7r locus in a manner dependent on the enhancement of both a GATA1-TET3 physical interaction and GATA1 binding at the p2x7r promoter. Importantly, we showed that demethylation of the p2x7r locus (and the attendant increase in P2X7R expression) was reversed upon knockdown of GATA1 or TET3 expression, and demonstrated that a decoy oligodeoxynucleotide that selectively blocked the GATA1 binding site increased the methylation of a CpG island in the p2x7r promoter. These results demonstrate that chronic visceral pain is mediated synergistically by GATA1 and TET3 via a DNA-demethylation mechanism that controls p2x7r transcription in spinal dorsal horn astrocytes, and provide a potential therapeutic strategy by targeting GATA1 and p2x7r locus binding.
Animals ; Astrocytes/metabolism* ; DNA Demethylation ; Epigenesis, Genetic ; GATA1 Transcription Factor/metabolism* ; Inflammation/metabolism* ; Oligodeoxyribonucleotides/metabolism* ; Rats ; Rats, Sprague-Dawley ; Receptors, Purinergic P2X7/metabolism* ; Visceral Pain/metabolism*

ObjectiveTo investigate the efficacy of Bushen Shengxue prescription and Yiqi Yangxue prescription in the treatment of chronic aplastic anemia and the effect on T cell subsets and the expression of T-box expressed in T cells （T-bet） and GATA binding protein 3 （GATA3）. MethodA total of 585 patients with chronic aplastic anemia who were treated in 19 hospitals in China from May 2018 to June 2021 were enrolled. With the prospective， double-blind and randomized control methods， the patients were randomized into three groups： kidney deficiency group， Qi and blood deficiency group， and control group. The three groups were respectively treated with Bushen Shengxue prescription granule， Yiqi Yangxue prescription granule， and Placebo （half the dose of Bushen Shengxue formula granules）. In addition， all of them were given oral cyclosporin and androgen. The treatment lasted 6 months， with 3 months as a course. The blood routine indexes， T cell subsets， and fusion genes T-bet and GATA3 before and after treatment were analyzed， and the safety indexes were monitored. ResultDuring the observation， a total of 75 cases dropped out and 18 were rejected. Finally， 161 cases in the kidney deficiency group， 164 in the Qi and blood deficiency group， and 167 in the control group were included. After 6 months of treatment， the total effective rate was 98.8% （159/161） in the kidney deficiency group， which was higher than the 79.9% （131/164） in the Qi and blood deficiency group （χ²=30.135， P<0.01） and the 61.7% （103/167） in the control group （χ²=70.126， P<0.01）. The total effective rate was higher in the Qi and blood deficiency group than in the control group （χ²=13.232， P<0.01）. After treatment， the hemoglobin （HGB） content increased significantly in three groups （P<0.05） as compared with that before treatment， particularly the kidney deficiency group （P<0.01）. After treatment， the white blood cell （WBC） count and platelet （PLT） count in the kidney deficiency group and the control group increased compared with those in the Qi and blood deficiency group （P<0.01）. There was no specific difference in neutrophils （ANC） after treatment among the three groups. At the same time point， the level of T helper type 1 （Th1） cells， Th1/Th2 ratio （P<0.05）， level of CD4⁺， and CD4⁺/CD8⁺ ratio （P<0.05） were significantly low in the kidney deficiency group among three groups. There was no significant difference in CD19^-， HLA/DR⁺， and CD25⁺ between the kidney deficiency group and the other two groups， but the T-bet of the kidney deficiency group and the control group was lower than that of the Qi and blood deficiency group （P<0.05）. ConclusionBushen Shengxue prescription exerts therapeutic effect on the aplastic anemia by improving the immunoregulatory mechanism， inhibiting the activity of immune system， modulating T cell subsets， suppressing Th1 and CD4⁺， and promoting bone marrow hematopoiesis. Moreover， it is safe with little side effects， which is worthy of further promotion.

In December, the outbreak of a novel coronavirus (2019-nCoV) in Wuhan, China, has attracted extensive global attention. On January 20, 2020,the Chinese health authorities upgraded the coronavirus to a Class B infectious disease in the Law of the People's Republic of China on the Prevention and Treatment of Infectious Diseases, and considered it as Class A infectious diseases in disease control and prevention. On January 22, 2020, the 2019-nCoV nucleic acid detection test was listed as the diagnostic criteria in the "guidelines for diagnosis and treatment of pneumonia due to 2019-nCoV (Trial Version 2)" . Therefore, standardizing the operation process of the 2019-nCoV nucleic acid detection in clinical laboratories has become a top priority. It is of paramount importance to establish standard protocols for detection of the 2019-nCoV nucleic acids in clinical laboratories to improve the reliability of the results and ensure the biosafety of laboratory personnel.

Objective: To investigate the diagnostic value of immunoglobulin M (IgM) and immunoglobulin G(IgG) antibodies to 2019 Novel Coronavirus (2019-nCoV) in 2019-nCoV infection. Method: This is a retrospective study. Serum samples were collected from 284 patients including outpatients and inpatients in the Renmin Hospital of Wuhan University from January 20, 2020 to February 17, 2020. Among them 205 cases were 2019-nCoV infected patients, including 186 cases confirmed with nucleic acid test and 19 cases diagnosed by clinical symptoms and CT characteristics according to "the New Coronavirus Pneumonia Control Protocol (5th edition)" . A total of 79 subjects with other diseases but negative to 2019-nCoV infection were recruited as control group. Serum IgM and IgG antibodies to 2019-nCoV were measured with fully automated immunoassay technology for all subjects. Statistical significance between 2019-nCoV antibodies test and 2019-nCoV nucleic acid test was determined using the χ² tests. Result: The sensitivity of serum IgM and IgG antibodies to 2019-nCoV were 70.24%(144/205) and 96.10%(197/205) respectively and the specificity were 96.20%(76/79) and 92.41%(73/79) respectively. The positive and negative predictive values of 2019-nCoV antibodies were 95.63%(197/206) and 91.03% (71/78) respectively, and the positive and negative predictive values of 2019-nCoV nucleic acid test were 100%(186/186) and 80.61%(79/98) respectively. The total coincidence rate of diagnosing 2019-nCoV infection between antibody tests and nucleic acid test for 2019-nCoV were 88.03%(250/284). Conclusion Joint detection of serum IgM and IgG antibodies to 2019-nCoV is an effective screening and diagnostic indicators for 2019-nCoV infection, and an effective complement to the false negative results to nucleic acid test.

Objective: To investigate the positive rate for 2019-nCoV tests and co-infections in Wuhan district. Methods: A total of 8 274 cases in Wuhan were enrolled in this cross-sectional study during January 20 to February 9, 2020, and were tested for 2019-nCoV using fluorescence quantitative PCR. Both respiratory tract samples (nasopharynx, oropharynx, sputum and alveolar lavage fluid) and non-respiratory tract samples (urine, feces, anal swabs, blood and conjunctival sac swabs) were collected. If both orf1ab and N genes are positive, they are classified as nucleic acid test positive group; if both orf1ab and N genes are negative, they are classified as negative group; if single gene target is positive, they are classified as suspicious group. Individuals were divided into male group and female group according to sex. At the same time, 316 patients were tested for 13 respiratory pathogens by multiplex PCR. Results: Among the 8 274 subjects, 2 745 (33.2%) were 2019-nCoV infected; 5 277 (63.8%) subjects showed negative results in the 2019-nCoV nucleic acid test; and 252 cases (3.05%) was not definitive (inconclusive result). The age of cases with COVID-19 patients and inconclusive cases was significantly higher than that of cases without 2019-nCoV infection (40 vs 56, t=27.569, P<0.001; 52 vs 56, t=6.774, P<0.001). The positive rate of 13 respiratory pathogens multiple tests was significantly lower in 104 subjects who were positive for 2019-nCoV compared with those in subjects who were negative for 2019-nCoV test (5.77% vs 18.39%, χ²=24.105, P=0.003). Four types of respiratory tract samples and five types of non-respiratory tract samples were found to be positive for 2019-nCoV nucleic acid test. Conclusion The 2019-nCoV nucleic acid positive rate in male is higher than in female. Co-infections should be pay close attention in COVID-19 patients. 2019-nCoV nucleic acid can be detected in non-respiratory tract samples.

ObjectiveTo investigate the influence of hepatitis C virus (HCV) on the expression of lipid metabolism indices in vitro and in vivo. MethodsA total of 114 samples of patients with HCV infection who were treated in Renmin Hospital of Wuhan University from September 2017 to September 2018 were collected as experimental group, and 96 samples of healthy individuals who underwent physical examination were collected as control group. An automatic biochemical analyzer was used to measure blood lipid parameters, including triglyceride (TG), total cholesterol (TCh), high-density lipoprotein (HDL), low-density lipoprotein (LDL), small and low-density lipoprotein (sdLDL), lipoprotein (a), apolipoprotein A1 (ApoA1), and apolipoprotein B (ApoB). RT-qPCR was used to measure the mRNA expression of fatty acid synthase (FASN), acetyl-CoA carboxylase (ACACA), hydroxymethylglutarate mono-acyl CoA reductase (HMGR), ApoA1, ApoB, and low-density lipoprotein receptor (LDLR) in Huh7.5.1 cells with or without HCV infection. The t-test was used for comparison of normally distributed continuous data between two groups, and the Mann-Whitney U test was used for comparison of non-normally distributed continuous data between two groups. ResultsCompared with the control group, the experimental group had significantly lower serum levels of TCh (2.98±0.51 mmol/L vs 4.24±0.43 mmol/L, t=4.96, P＜0.05), HDL (0.87±0.16 mmol/L vs 1.24±0.21 mmol/L, t=5.65, P＜0.05), LDL (1.75±0.24 mmol/L vs 2.64±0.37 mmol/L, t=3.88, P＜0.05), ApoA1 (0.94±0.18 mmol/L vs 1.47±0.26 mmol/L, t=3.71, P＜0.05), and ApoB (0.67±0.31 mmol/L vs 0.98±0.14 mmol/L, t=4.41, P＜0.05). Huh7.5.1 cells with HCV infection had significantly lower mRNA expression of ApoA1 than those without HCV infection (t=-3.43, P＜0.05), as well as significantly higher mRNA expression of FASN, ACACA, HMGR, ApoB, and LDLR (t=5.40, 4.93, 3.34, 6.88, and 3.84, all P＜0.05). ConclusionHCV infection can upregulate the mRNA levels of enzymes involved in the synthesis of fatty acids and cholesterol and thus affect lipid metabolism in vivo and in vitro.

Hepatitis C virus (HCV) infection is one of the leading causes of liver cirrhosis, end-stage liver disease, and even liver cancer. Patients with HCV infection tend to have insulin resistance and dyslipidemia, which may lead to a series of metabolic syndromes and greatly threaten human health. At present, there are still no effective vaccines to prevent HCV infection, and therefore, the mechanism of HCV infection remains a hot topic in clinical research. Many studies have shown that there is a complex relationship between HCV infection and lipids, but the role of lipids in HCV infection remains unclear. This article reviews the current research status of the role of lipid metabolism in HCV infection and life cycle, in order to understand the mechanism of abnormal fat metabolism in the liver and the pathogenesis of fatty liver disease.

Objective To investigate the relationship between the dynamic changes of interleukin-1β (IL-1β) levels and severity and complications of patients with multiple trauma at the early stage.Methods Among 97 patients with multiple trauma in Emergency Department of Shanghai Jiao Tong University Affiliated Sixth People's Hospital between August 2015 and May 2016,12 patients were excluded as follows,(1) with burns or chemical injuries;(2) pregnancy or menopausal women;(3) had bacterial infection a week ago;(4) with chronic diseases.The other 85 patients with multiple traumas were classified into three categories according to the injury severity score (ISS).That is,the slight group (22 cases,9≤ ISS ＜ 15),moderate group (35 cases,15 ≤ ISS ＜ 25) and severe group (28 cases,ISS ≥ 25).Their venous blood samples were collected at 6,12,24,48 and 72 hours after trauma respectively,and the serum IL-1 β levels were measured using a specific immunoluminometric assays.The basal conditions including age,the hospitalization days and so on among these three groups were compared via ANOVA.The mean IL-1 β levels at above time intervals among three groups were compared.Finally,the relationship between the peak concentration of IL-1β and injury severity and complications was analyzed by multiple Logistic regression.Results (1) As the increasing severity of trauma,the patients with longer days of hospitalization and higher rate of multiple organ dysfunction syndrome (MODS) (P ＜ 0.05).(2) The levels of IL-1 β in the moderate and severe groups were remarkably higher than those in the slight group (P ＜ 0.02).(3) The IL-1β levels in each group peaked at 6 hours after trauma and began to decline.(4) Multivariate logistic analysis showed that peak concentration of IL-1 β was still an independent predictor for injury severity (moderate group:odds ratio,1.21;95％ confidence interval:1.05-1.39,P =0.007;severe group:odds ratio,1.20;95％ confidence interval:1.03-1.40,P =0.019) and sepsis (odds ratio,1.28;95％ confidence interval:1.10-1.50,P =0.001),but had no significant association with MODS and trauma mortality even after controlling other risk factors.Conclusions The serum IL-1β at 6 hours after injury could be used as an early effective indicator to evaluate the injury severity and infectionrelated complications in patients with multiple trauma.

Objective To construct the eukaryotic expression vector of cyclin-dependent kinase ( CDK) 7 labeled with Myc tag, obtain the expressed product , and identify its interaction with FLAG-P53 at the protein level .Methods Human CDK7 coding gene region amplified from the mammary cDNA library by PCR was inserted into the pXJ -40 vector.The recombinant plasmid Myc-CDK7 transfected into human 293T cell lines was investigated and examined by SDS-PAGE and Western blotting.In addition,assay was applied to determine the interaction between Myc-CDK7 and FLAG-p53.Results The coding region of CDK7 was successfully amplified by PCR and cloned into pXJ-40 vector, which was identified by double enzyme digestion and gene sequencing .Myc-CDK7 was successfully expressed in human 293T cell lines according to SDS-PAGE and Western blotting assay indicated that Myc-CDK7 could interact with P53 protein, which verified its known function .Conclusion The eukaryotic expression vector Myc-CDK7 is successfully obtained , which will contribute to further research on CDK 7-mediated cell cycle regulation .

Objective To study the biomechanics of simple locking proximal femoral plate (LPFP),LPFP combined with calcium phosphate cement (CPC) and proximal femoral nail antirotation (PFNA) in treatment of unstable intertrochanteric fractures.Methods Thirty-six femurs from 18 specimens with no pathological defects,fractures,malformations or neoplasms were enrolled in the study and were divided into three groups (Groups A,B and C),with 12 femurs per group.Intertrochanteric fractures were duplicated in accordance with Evans classification of type Ⅲ A.The fractures in Group A were treated by LPFP combined with CPC fixation,that in Group B treated with only LPFP fixation,while that in Group C fixed with PFNA.A comparative mechanical test of femoral specimens was performed in a special clamp.Results Axial compression test showed linear change of the load-displacement of three surgical fixations for intertrochanteric fractures under biological load.The measurement of torque-torsion angle of three fixations for intertrochateric fractures showed that the torque was enlarged with increase of torsion angle till the failure of internal fixation.Conclusion LPFP combined with CPC or PFNA can attain solid fixation of unstable intertrochanteric fractures.