Objective To investigate the effects and underlying mechanisms of total flavonoids of sarcandra glabra(TFFSG)on bone marrow mesenchymal stem cells(BMSCs)and their derived exosomes in immune thrombo-cytopenia(ITP),with a focus on promoting megakaryocyte differentiation and maturation.Methods BMSCs induced by rabbit anti-rat platelet serum(APS)were divided into five groups:a blank control group,an ITP-BMSCs model group,and three TFFSG intervention groups with low(1.95 μg/mL),medium(3.90 μg/mL),and high doses(7.80 μg/mL).The apoptosis rates and the expression levels of apoptosis-related proteins-B-cell lymphoma 2(Bcl-2),Bcl-2-associated X protein(BAX),and Cysteinyl aspartate-specific proteinase-3(Caspase-3)-were assessed.Exosomes were isolated from the blank control group(NC-BMSCs-Exos),the ITP-BMSCs model group(ITP-BMSCs-Exos),and the medium-dose TFFSG group(TFFSG-BMSCs-Exos).Each group's exosomes(5 μg/mL)were co-cultured with megakaryocytic lineage Dami cells for 96 hours.Flow cytometry was employed to evaluate the expression of megakaryocytic differentiation markers(CD41a,CD42b,CD61)and the proportion of polyploid cells(≥4 N)in each group.Western Blot analysis was conducted to examine the expression of p-MEK1/2,MEK1/2,p-ERK1/2,and ERK1/2 across all groups.Results Compared with the ITP-BMSCs model group,the apoptosis rates in all TFFSG intervention groups were significantly reduced(P<0.01).In the medium-and high-dose TFFSG groups,BAX and Caspase-3 expression levels were markedly downregulated,whereas Bcl-2 expression was upregulated(P<0.05,P<0.01).Compared with the ITP-BMSCs-Exos group,the TFFSG-BMSCs-Exos group demonstrated increased expression of CD41a+,CD42b+,and CD61+,a higher proportion of polyploid cells(≥4 N)(P<0.05),as well as elevated ratios of p-MEK1/2 to MEK1/2 and p-ERK1/2 to ERK1/2(P<0.05).Conclusion TFFSG inhibits apopto-sis of ITP-state BMSCs in vitro and promotes megakaryocyte differentiation and polyploidization maturation through BMSC-derived exosomes by activating the MEK1/2-ERK1/2 signaling pathway.

Objective To investigate the effects and underlying mechanisms of total flavonoids of sarcandra glabra(TFFSG)on bone marrow mesenchymal stem cells(BMSCs)and their derived exosomes in immune thrombo-cytopenia(ITP),with a focus on promoting megakaryocyte differentiation and maturation.Methods BMSCs induced by rabbit anti-rat platelet serum(APS)were divided into five groups:a blank control group,an ITP-BMSCs model group,and three TFFSG intervention groups with low(1.95 μg/mL),medium(3.90 μg/mL),and high doses(7.80 μg/mL).The apoptosis rates and the expression levels of apoptosis-related proteins-B-cell lymphoma 2(Bcl-2),Bcl-2-associated X protein(BAX),and Cysteinyl aspartate-specific proteinase-3(Caspase-3)-were assessed.Exosomes were isolated from the blank control group(NC-BMSCs-Exos),the ITP-BMSCs model group(ITP-BMSCs-Exos),and the medium-dose TFFSG group(TFFSG-BMSCs-Exos).Each group's exosomes(5 μg/mL)were co-cultured with megakaryocytic lineage Dami cells for 96 hours.Flow cytometry was employed to evaluate the expression of megakaryocytic differentiation markers(CD41a,CD42b,CD61)and the proportion of polyploid cells(≥4 N)in each group.Western Blot analysis was conducted to examine the expression of p-MEK1/2,MEK1/2,p-ERK1/2,and ERK1/2 across all groups.Results Compared with the ITP-BMSCs model group,the apoptosis rates in all TFFSG intervention groups were significantly reduced(P<0.01).In the medium-and high-dose TFFSG groups,BAX and Caspase-3 expression levels were markedly downregulated,whereas Bcl-2 expression was upregulated(P<0.05,P<0.01).Compared with the ITP-BMSCs-Exos group,the TFFSG-BMSCs-Exos group demonstrated increased expression of CD41a+,CD42b+,and CD61+,a higher proportion of polyploid cells(≥4 N)(P<0.05),as well as elevated ratios of p-MEK1/2 to MEK1/2 and p-ERK1/2 to ERK1/2(P<0.05).Conclusion TFFSG inhibits apopto-sis of ITP-state BMSCs in vitro and promotes megakaryocyte differentiation and polyploidization maturation through BMSC-derived exosomes by activating the MEK1/2-ERK1/2 signaling pathway.

Deinagkistrodon acutus(also known as the five-paced viper and the traditional Chinese medicine ingredient is called Agkistrodon) venom is a viscous liquid from the venom glands of the Deinagkistrodon acutus. It contains a variety of protein and peptide components such as phospholipase A2, serine protease, metalloproteinase, C-type lectin, L-amino acid oxidase, and has a variety of biological activities, playing an important role in anti-tumour, anti-thrombotic, anti-inflammatory and anti-bacterial activities. In recent years, snake venom research has become increasingly widespread, but there is still a lack of comprehensive and systematic studies on snake venom from Deinagkistrodon acutus. In this paper, the source, identification, active ingredients, toxicity studies and quality researches of Deinagkistrodon acutus venom are summarized and analysed by searching the related research progress, in order to provide reference for further development and utilization of Deinagkistrodon acutus venom.

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.

Objective To investigate the clinicopathological characteristics and prognosis of patients with Borrmann type Ⅳ gastric cancer. Methods A cohort retrospective analysis of 2386 patients with gastric cancer who underwent radical surgery was used to screen out Borrmann type Ⅳ patients, and analyze their clinical features and prognostic factors. Results Among 2386 patients with gastric cancer, 363 cases (15.21%) were Borrmann type Ⅳ. Compared with non-Borrmann type Ⅳ gastric cancer patients, Borrmann type Ⅳ patients had higher rates of simultaneous liver metastasis, metachronous liver metastasis, lymph node metastasis and vascular infiltration. Moreover, the age of onset tended to be younger and the pathological type tended to be poorly differentiated-undifferentiated (all P < 0.05). The 5-year OS of the entire group was 49.32% and the 5-year DFS was 44.61%. There were significant differences in 5-year OS and DFS between Borrmann type Ⅳ and non-Borrmann type Ⅳpatients (all P < 0.001). The subgroup analyses showed that there were statistically significant differences in 5-year OS and DFS of gastric cancer patients between Borrmann type Ⅳ and non-Borrmann type Ⅳ in pT2-pT4a and pN0-pN3a stages (all P < 0.005). Multivariate analysis showed that the poorly differentiated-undifferentiated tumor, the T4a-pT4b stage of tumor invasion depth, lymph node metastasis, the ⅢA-ⅢC pTNM stage of the tumor, postoperative liver metastasis and peritoneal metastasis were independent risk factors affecting the prognosis of Borrmann type Ⅳ gastric cancer patients (all P < 0.05). Conclusion Borrmann type Ⅳ gastric cancer is prone to liver metastasis, lymph node metastasis, peritoneal metastasis and poor prognosis, and it's prognosis is affected by a variety of independent risk factors.

Objective: Apolipoprotein E (ApoE) is mainly synthesized in the liver. So far, it is unknown the relationship among APOE gene polymorphisms and WML, brain atrophy. Therefore, the aim of the study was to assess the associations of APOE gene polymorphisms in patients with WML and brain atrophy. Methods: A total of 58 patients with WML, 128 patients with brain atrophy, 112 patients with co-occurrence of WML and brain atrophy and 95 healthy elderly volunteers were recruited from Renmin Hospital of WuHan University. Results: Allele E3 was the most common allele. The alleles E2 had significantly higher levels of ApoB and lower age in WML group. The alleles E2 was associated with the lower level of ApoB, LDL-Ch, TCh, and sdLDL in co-occurrence group. The E3/E3 genotype has higher level of sdLDL, but lower age and female frequency in WML. The E3/E4 genotype had higher level of TG, but lower age in WML. Gender, Age, E2, Hyperhomocysteinemia and UA were also significantly associated with disease progression. Conclusion This study found that clinical data, lipids and metabolic complications were closely related to ApoE genotypes and alleles, and also disease progression and type.

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.