Objective To investigate the effects of AIDS and tuberculosis(TB)co-infection on CD8+T lymphocytes.Methods Serum CD8+T cells and CD4+T cells were analyzed and compared between human immunodeficiency virus(HIV)alone patients(n=200),TB alone patients(n=200),and HIV/TB co-infection patients(n=200).Results CD8+T cells were significantly higher in HIV alone group compared to HIV/TB co-infection group and TB alone group(x2=128.779,P＜0.001).The level of CD8+T lymphocytes in HIV/TB co-infection group and HIV alone group was higher than normal level(ZHIV/TB=8.343,PHIV/TB＜0.001,ZHIV=7.988,PHIV＜0.001).The CD8+T cells in TB alone group decreased significantly compared with normal levels(ZTB=8.682,PTB＜0.001).The levels of CD4+T cells in the three groups of patients were all lower than normal(ZHIV=11.088,PHIV＜0.001,ZTB=5.562,PTB＜0.001,ZHIV/TB=12.077,PHIV/TB＜0.001).The stratified analysis of CD8+T cell levels by CD4+T cell counts,showed that when CD4+T cell count was≤100 cells/μL,the level of CD8+T cells in HIV alone group was higher than that in TB alone group.HIV alone group had higher CD8+T cells than HIV/TB co-infection group,and the level of CD8+T cells in HIV/TB co-infection group was significantly lower than normal(Z0-100=1.604,P0-100=0.109).When CD4+T cells ≥101 cells/μL,HIV/TB co-infection group had higher CD8+T cells than TB alone group.HIV alone group had higher CD8+T cells than TB alone group.When CD4+T cells＞500 cells/μL,CD8+T cell levels were within the normal range in all three groups of patients.Taking the HIV alone group as a reference,with the decrease of CD4+T cell count,the CD8+T cell count decreased more significantly in TB alone group and HIV/TB co-infection group than in HIV alone group(slope was 0.344 and 0.216,respectively,P＜0.001).Conclusions With the decline of CD4+T cells,both HIV/TB co-infection and TB alone are associated with decrease in CD8+T cells,and the decrease in the TB alone group is more prominent.

Objective:To investigate the effectiveness and feasibility of whole exome sequencing (WES), as a molecular diagnosis technique, for adult patients with genetic diseases.Methods:The present retrospective analysis included 445 adult patients (ages 18-80 years) with suspected genetic diseases who underwent whole exome sequencing (WES) from August 2021 to December 2022. The pathogenicity classification of each variant was assessed in accordance with the recommendations developed by the American Society of Medical Genetics and Genomics.Results:The overall positive rate of WES among adult patients with suspected genetic diseases was 28.08% (125/445). The highest positive rate was observed in the age group of 41-50 years (34.33%, 23/67). Among the diagnosed genetic diseases, those affecting the cardiovascular system (63.16%, 84/133), nervous system (18.05%, 24/133), and endocrine system (13.53%, 18/133) ranked as the top three. The most common genetic diseases identified through WES in adult patients were hypertrophic cardiomyopathy (18.80%, 25/133), dilated cardiomyopathy (16.54%, 22/133), Marfan syndrome (15.04%, 20/133), epilepsy (9.02%, 12/133), and familial hypercholesterolemia (4.51%, 6/133). The main causative genes identified included FBN1 (14.29%, 19/133), MYBPC3 (9.02%, 12/133), MYH7 (9.02%, 12/133), LDLR (3.76%, 5/133), TTN (3.76%, 5/133), and TNNI3 (3.01%, 4/133).Conclusion:Applying the WES technique in clinical practice can improve the diagnostic rate of adult genetic diseases, especially in adult patients with suspected genetic conditions involving the cardiovascular system, nervous system, and endocrine system.

Objective To investigate the effect of plasma microRNA(miR)-93-5p on ovarian granulosa cell proliferation in patients with polycystic ovary syndrome(PCOS)and its possible mechanism.Methods A total of 120 PCOS patients of childbearing period who were treated at Jiaozuo People's Hospital from January 2022 to January 2023 were selected as the PCOS group,and 18 healthy women of childbearing period who were physically examined at the same hospital during the same period were selected as the control group.The age,body weight and height of the subjects in the two groups were recorded,and the body mass index(BMI)was calculated.Fasting venous blood was collected from the subjects during the follicular phase of the natural menstrual cycle or the progesterone-induced withdrawal bleeding phase,and serum levels of luteinizing hormone(LH),follicle-stimulating hormone(FSH),total testosterone(TT),anti-Mullerian hormone(AMH),aspartate aminotransferase(AST),alanine aminotransferase(ALT),fasting insulin(FINS),and fasting blood glucose(FBG)were measured by using the electrochemical method;and the homeostasis model assessment of insulin resistance(HOMA-IR)was calculated.Quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect the expression level of miR-93-5p in plasma of patients in the two groups.Human ovarian granulosa cells(KGN cells)in logarithmic growth phase were cultured in 6-well plates with 3 × 105 cells per well,and they were randomly divided into the miR-93-5p mimics group,LY294002+miR-93-5p group,AZD5363+miR-93-5p group,negative control(NC)group,and blank control group.The KGN cells in the miR-93-5p group were transfected with miR-93-5p mimics;the KGN cells in the LY294002+miR-93-5p group were transfected with miR-93-5p mimics and treated with 50 mmol·L-1 LY294002 one hour before transfection;the KGN cells in the AZD5363+miR-93-5p group were transfected with miR-93-5p mimics and treated with 50 μmol·L-1 AZD5363 one hour before transfection;the KGN cells in the NC group were transfected with the negative control plasmids;and the KGN cells in the blank control group were not treated at all.The expression of miR-93-5p in the KGN cells in each group was detected by qRT-PCR,and the proliferation of the KGN cells in each group was detected by cell counting kit-8.Results There was no significant differences in age,FSH,ALT,and AST levels of patients between the PCOS group and the blank control group(P＞0.05).The BMI,TT,AMH,LH,FINS,FBG,and HOMA-IR of patients in the PCOS group were significantly higher than those in the blank control group(P＜0.05).The relative expression of miR-93-5p in plasma of patients in the PCOS group was significantly higher than that in the blank control group(t=-5.549,P＜0.001).miR-93-5p was moderately positively correlated with TT,FINS and HOMA-IR(r=0.434,0.622,0.586;P＜0.001)and was mildly positively correlated with FBG and LH(r=0.398,0.398;P＜0.001).The receiver operating characteristic curve showed that the optimal cut-off value for plasma miR-93-5p in diagnosing PCOS was 1.380,the area under the curve was 0.906(95％confidence interval:0.839-0.973,P＜0.001),the sensitivity was 0.858,the specificity was 0.833,and the Youden index was 0.691.The relative expression of miR-93-5p in the KGN cells in the miR-93-5p mimics group,LY294002+miR-93-5p group and AZD5363+miR-93-5p group was significantly higher than that in the blank control group and NC group(P＜0.05).After 24,48 and 72 hours of culture,the proliferation of the KGN cells in the miR-93-5p mimics group,LY294002+miR-93-5p group and AZD5363+miR-93-5p group was significantly higher than that in the blank control group and the NC group(P＜0.05);the proliferation of the KGN cells in the LY294002+miR-93-5p group and AZD5363+miR-93-5p group was significantly lower than that in the miR-93-5p mimics group(P＜0.05).Conclusion miR-93-5p in plasma is overexpressed in PCOS patients,and it may be involved in the occurrence and development of PCOS by mediating the proliferation of ovarian granulosa cells through the phosphoinositide 3 kinase/protein kinase B signaling pathway.The miR-93-5p level in plasma has a certain diagnostic value for PCOS.

Objective To explore the expression level of microRNA(miR)-320a-3p in peripheral blood in patients with polycystic ovary syndrome(PCOS)and its possible mechanism for regulating PCOS.Methods A total of 149 PCOS patients admitted to the Jiaozuo People's Hospital from July 2021 to July 2022 were selected as the research subjects(PCOS group),and 18 healthy volunteers with a regular menstrual cycle(28-35 d),no clinical or biochemical manifestations of hyperandrogenism,and no polycystic ovary changes detected by ultrasonography were selected as the control group.Clinical data of the subjects in the two groups were collected and compared.The expression level of miR-320a-3p in plasma of subjects in the two groups was detected by using the real-time quantitative polymerase chain reaction.The correlation between plasma miR-320a-3p expression and clinical indexes was evaluated by using the Pearson partial correlation analysis.The predictive value of miR-320a-3p for PCOS was analyzed by using the receiver operating characteristic curve.The relationship between miR-320a-3p and androgen receptor was evaluated by using the dual luciferase reporter gene assay.Results There was no significant difference in age,hip circumference,follicle stimulating hormone(FSH),prolactin(PRL),and high-density lipoprotein cholesterol(HDL-C)between the two groups(P＞0.05).The body mass index,waist circumference,total testosterone(TT),anti-Miillerian hormone(AMH),luteinizing hormone(LH),fasting insulin(FINS),fasting blood glucose(FBG),2-hour postprandial blood glucose(2 h BG),homeostasis model assessment of insulin resistance(HOMA-IR),total cholesterol,triglycerides(TG),and low-density lipoprotein cholesterol(LDL-C)of patients in the PCOS group were significantly higher than those in the control group,while the level of estradiol(E2)was significantly lower than that in the control group(P＜0.05).The relative expression level of miR-320a-3p in plasma of patients in the PCOS group was significantly lower than that in the control group(P＜0.05).The expression level of miR-320a-3p was moderately negatively correlated with TT(r=-0.594,P＜0.05)and slightly negatively correlated with waist circumference,FINS,2 h BG,HOMA-IR,and LDL-C(r=-0.293,-0.208,-0.227,-0.208,-0.208;P＜0.05),and showed no significant correlation with hip circumference,AMH,FSH,E2,PRL,FBG,TG,and HDL-C(r=-0.079,0.020,-0.042,0.089,0.005,-0.141,-0.116,0.059;P＞0.05).The area under the curve of miR-320a-3p for predicting PCOS was 0.968(95％confidence interval:0.922-1.000,P＜0.05),with a cut-off value of 0.515,a sensitivity of 0.917,a specificity of 0.789,and a Jordon index of 0.706.miR-320a-3p negatively regulated androgen receptors.Conclusion miR-320a-3p is abnormally low expressed in peripheral blood of PCOS patients and may participate in the occurrence and development of PCOS by negatively regulating androgen receptors to increase TT levels.It has high predictive value for diagnosing PCOS.

Objective To investigate the mechanism of Yes-associated protein 1(YAP1)ameliorating aristolochic acid 1(AAI)-induced liver injury in mice based on untargeted metabolomics techniques.Methods There were 83-week-old male hepatocyte-specific Yap1 gene knockout mice(genotyped as Yap1Flox/Flox,Albumin-Cre,aka.Yap1LKO)were randomly selected as the Yap1LKO+AAI group,and 8 Yap1Flox control mice as the Yap1Flox+AAI group.Both groups were injected intraperitoneally with AAI at a dose of 2.5 mg·kg-1·d-1 for 14 consecutive days.Genotypes were identified by tail PCR;serum alanine transaminase(ALT)and aspartate transaminase(AST)activities were determined by microplate assay;histopathological changes of liver tissue were observed by HE staining;and the protein expression of YAP1 in liver tissue was determined by immunohistochemistry.The untargeted metabolomics approach was used to analyze the liver tissue differential metabolites,and the samples were analyzed by ultra performance liquid chromatography-quadrupole-electrostatic field orbit trap high-resolution mass spectrometry,and the differential metabolites were screened by principal component analysis(PCA),Partial least square-discriminant analysis(PLS-DA),and orthogonal partial least squares-discriminant analysis(OPLS-DA);using HMDB database and METLIN database to identify metabolites,and the pathway enrichment of differential metabolites was analyzed by KEGG database.Results(1)After 14 days of AAI induction,the increase of body mass in Yap1LKO mice was lower than that in Yap1Flox mice,but there was no statistical significance(P＞0.05).On day 14,compared with the Yap1Flox+AAI group,the serum ALT and AST enzyme activities in the Yap1LKO+AAI group of mice were significantly increased(P＜0.05),and the histopathological damage of the liver was significantly aggravated.The livers of the Yap1Flox mice had a positive protein expression of YAP1,whereas the Yap1LKO mice did not have a positive protein expression of YAP1.(2)A total of 139 differential metabolites with significant changes(VIP＞1 and P＜0.05)were screened by metabonomic analysis;compared with Yap1LKO+ AAI group,62 liver metabolites in Yap1Flox+AAI group were up-regulated,including choline,taurine,hypotaurine,α-linolenic acid,eleostearic acid,chenodeoxycholic acid and so on.Seventy-seven metabolites were down-regulated including glycerophosphocholine,L-phosphatidylcholine,L-glutamine,L-serine,L-glutathione,5-methionine,phenylalanine,glucose 6-phosphate,lactic acid,uric acid glycosides,etc..KEGG-enriched pathways were mainly choline metabolism,glycerophospholipid metabolism,insulin resistance,glutathione metabolism,etc..Conclusion Hepatocyte-specific Yap1 gene knockout exacerbated AAI-induced liver injury in mice,and YAP1 was involved in the regulation of choline metabolism and glycerophospholipid metabolism through the up-regulation of unsaturated fatty acids,such as choline and taurine,which ameliorated AAI-induced liver injury in mice.

ObjectiveTo evaluate the reliability and validity of the Dampness Syndrome Scale of Chinese Medicine （DSSCM） among patients with persistent asthma， and to explore the correlation between dampness syndrome and clinical characteristics of persistent asthma. MethodsA cross-sectional survey was conducted. Basic information， examination results， DSSCM， Asthma Control Test （ACT）， Generalized Anxiety Disorder-7 （GAD-7）， and Patient Health Questionnaire-9 （PHQ-9） scores were collected from 206 patients with persistent asthma to evaluate the reliability and validity of DSSCM and to explore the correlation between dampness syndrome and clinical characteristics. ResultsThe mean score of DSSCM among 206 patients was 14.59 ± 10.53. The overall Cronbach α coefficient and Spearman-Brown split-half reliability coefficient of the scale were both greater than 0.8， and the success rate of scale convergent and discriminant validity calibration were greater than 80%. The confirmatory factor analysis showed that the χ²/df was 2.309， and the root mean square error of approximation （RMSEA） was 0.08； the root mean square residual （RMR） was 0.049， whereas the comparative fit index （CFI）， the goodness of fit index （GFI）， the adjusted goodness of fit index （AGFI）， the normed fit index （NFI） and the incremental fit index （IFI） were less than 0.9. Correlation analysis showed that DSSCM scores were positively correlated with disease duration， GAD-7 scores， and PHQ-9 scores （P＜0.05）， and negatively correlated with ACT scores （P＜0.01）. The DSSCM scores were significantly different between patients with different disease severity （H = 10.92， P = 0.01）， and the DSSCM scores of allergic patients were higher than those of non-allergic patients （Z = -4.19， P＜0.001）. ConclusionDSSCM has acceptable reliability and validity for patients with persistent asthma. The scores of DSSCM correlated with the disease duration， ACT score， GAD-7 score， PHQ-9 score， disease severity and allergic status of persistent asthmatics.

Objective To study the relationship between economic level and cognitive level of the elderly, and to test the mediating effect of social participation between the two. Methods SPSS 26.0 was used for data processing. Ordinary least squares (OLS) linear regression was used to analyze the relationship between economic level, social participation and cognitive level. The mediation effect test was used to test the mediating effect of social participation between economic level and cognitive level. Results In the OLS regression model, model 2 showed that the estimation of economic level was 0.000 003 52, model 3 showed that the estimation of social participation was -0.316 907 5, and model 4 showed that the economic level was -0.000 003 39, all of which were significant at the level of 1%. The mediating effect size of social participation was 0.298 295 45, and the robustness test showed that the correlation coefficient between economic level and cognitive level orientation was 0.000 002 15, which was highly significant at the 1% level. Conclusion There is a positive correlation between the economic level and cognitive level of the elderly, and there is a positive correlation between the social participation and cognitive level of the elderly. The economic level of the elderly improves their cognitive level through the mediating effect of social participation, and the impact effect of social participation is 29.83%.

The Ehlers-Danlos syndrome(EDS)is a rare inherent connective tissue disorder.The prev-alence of EDS in the population is estimated at one out of ten thousand to one out of a hundred thousand.The vascular EDS(vEDS)are rare among the subtypes but are the worst in prognosis.The article reports a case of vEDS admitted to the hospital.The patient was a young man complaining of a sudden onset of aphasia in right hemiparalysis and severe left abdominal pain for unknown reasons.The diagnosis was made after the genetic testing.The patient suffered from vEDS.Then,the multi-disciplinary team(MDT)made a treatment plan tailored to this young patient.The complexity in classification and delusive presentations of the EDS make the correct diagnosis very challenging.This article hopes to report this case and to share the experiences to the bet-ter understanding of this disease.

Objective To correctly identify the blood group of ABO and study its molecular biological characteristics.Methods Blood samples from blood donors with discrepancies in forward and reverse typing using the microplate method were subjected to both saline tube agglutination test for serological identification and polymerase chain reaction-sequence specific primers(PCR-SSP)for genotyping.Additionally,direct sequencing of exons 1 to 7 of the ABO gene was performed on some donor samples to analyze their blood phenotype,genotype and gene sequence.Results For 256 samples showing discrepancy between forward and reverse typing in microwell method,119 were identified as normal ABO blood types,90 were weakened ABO antibodies and 47 were ABO subtypes by serology tube test.According to the PCR-SSP genotyping test,233 of 256(91.02%)were consistent with serological phenotype and genotype,17 of 256(6.64%)were inconsistent,and 6 samples can′t read genotype based on the kit result typing table.A total of 17 genotypes were identified in 250 samples as AO1 in 56,AO2 in 58,AA in 50,BO1 in 31,BO2 in 17,BB in 8,O1O1 in 2,O1O2 in 7,AB in 13,AO4 in 1,A205O2 in 1,A205A in 1,A201A in 1,O1O4 in 1,O2O2 in 1,A201B in 1 and A205B in 1.Sequencing of exons 1 to 7 of the ABO gene was carried out in 78 samples,and 29 ABO alleles were detected,seven of which were common alleles(?A101,?A102,?A104,?B101,?O01,?O02,?O04),22 of which were rare alleles(?A201,?A205,?Ax01,?Ax03,?Ax13,?Ax19,?Ax22,?Ael10,?B305,?Bel03,?Bel06/?Bx02,?Bw07,?Bw12,?Bw17,?Bw37,?O05,?O26,?O61,?B(A)04,?B(A)07,?cisAB01 and ?cisAB01var).In addition,six rare allele mutation sites were identified(c.101A＞G;c.103_106delG;c.146_147insGC;c.259G＞T;c.322C＞T;c.932T＞C).Conclusion The identification of ambiguous ABO blood group requires the combination of serological testing and molecular biological examination to correctly identify the blood type and ensure the safety of clinical blood transfusion.

Objective To express recombinant Burkholderia pseudomallei(B.pseudomallei)type Ⅲ secretion system BipD protein,prepare its polyclonal antibodies and verify their immunological traits.Methods The recombinant pET-28a-BipD plasmid was generated,and the pET-28a-BipD-carried E.coli BL21(DE3)bacteria were induced with isopropyl-β-d-thiogalactoside(IPTG)to express recombinant BipD(rBipD)protein.The rBipD was obtained by affinity chromatography using His Trap column,then mixed with Fredrick's adjuvant to immunize BALB/c mice by intraperitoneal injection in order to obtain anti-rBipD polyclonal antibodies.The immunoreactivity of rBipD was detected by Western blot assay using rabbit anti-melioidosis serum and the serum from melioidosis patients.The immunogenicity of rBipD was evaluated using Western blotting and immunofluorescence staining.Finally,rBipD was used to establish an indirect ELISA to detect serum antibodies of clinical melioidosis patients.Results The recombinant plasmid pET-28a-BipD was successfully constructed and transformed into E.coli BL21(DE3)to induce rBipD expression with IPTG treatment.The obtained rBipD had a relative molecular weight of 36×103 and a purity of 95.4％,and had good immunogenicity and immunoreactivity.It could induce the production of specific antibodies after immunizing mice,and mouse polyclonal antibodies against rBipD were prepared with the titer of 1∶512 000.rBipD of 5.0 μg/mL produced specific immune response with the serum of melioidosis patients,but had no specific reaction with the serum of tuberculosis patients,with statistical difference(P＜0.01).Conclusion rBipD with immunological activity is successfully prepared and purified,and its polyclonal antibodies are also developed,which provide a good tool for clinical immunological diagnosis and study of immune mechanism of B.pseudomallei infection.