AIM: To further analyze the interaction of tupaia CD81 with hepatitis C virus (HCV) envelope protein E2. METHODS: A tupaia CD81 large extracellular loop (CD81 LEL), which binds to HCV E2 protein, was cloned and expres...AIM: To further analyze the interaction of tupaia CD81 with hepatitis C virus (HCV) envelope protein E2. METHODS: A tupaia CD81 large extracellular loop (CD81 LEL), which binds to HCV E2 protein, was cloned and expressed as a GST-fusion protein, and interaction of HCV E2 protein with a tupaia CD81 LEL was evaluated by enzyme-linked immunosorbent assay (EIA). RESULTS: Although tupaia and human CD81 LEL differed in 6 amino acid changes, tupaia CD81 LEL was strongly recognized by anti-CD81 antibodies against human CD81 LEL conformation-dependent epitopes. Investigating LEL CD81-E2 interactions by EIA, we demonstrated that binding of tupaia CD81 LEL GST fusion protein to recombinant HCV E2 protein was markedly reduced compared to binding of human CD81 LEL GST fusion protein to recombinant HCV E2 protein. CONCLUSION: These data suggest that the structural differences in-between the tupaia and human CD81 may alter the interaction of the large extracellular loop with HCV envelope glycoprotein E2. These findings may be important for the understanding of the mechanisms of binding and entry of HCV to PTHs.展开更多
BACKGROUND Special AT-rich sequence binding protein 2(SATB2)-associated syndrome(SAS;OMIM 612313)is an autosomal dominant disorder.Alterations in the SATB2 gene have been identified as causative.CASE SUMMARY We report...BACKGROUND Special AT-rich sequence binding protein 2(SATB2)-associated syndrome(SAS;OMIM 612313)is an autosomal dominant disorder.Alterations in the SATB2 gene have been identified as causative.CASE SUMMARY We report a case of a 13-year-old Chinese boy with lifelong global developmental delay,speech and language delay,and intellectual disabilities.He had short stature and irregular dentition,but no other abnormal clinical findings.A de novo heterozygous nonsense point mutation was detected by genetic analysis in exon 6 of SATB2,c.687C>A(p.Y229X)(NCBI reference sequence:NM_001172509.2),and neither of his parents had the mutation.This mutation is the first reported and was evaluated as pathogenic according to the guidelines from the American College of Medical Genetics and Genomics.SAS was diagnosed,and special education performed.Our report of a SAS case in China caused by a SATB2 mutation expanded the genotype options for the disease.The heterogeneous manifestations can be induced by complicated pathogenic involvements and functions of SATB2 from reviewed literatures:(1)SATB2 haploinsufficiency;(2)the interference of truncated SATB2 protein to wild-type SATB2;and(3)different numerous genes regulated by SATB2 in brain and skeletal development in different developmental stages.CONCLUSION Global developmental delays are usually the initial presentations,and the diagnosis was challenging before other presentations occurred.Regular follow-up and genetic analysis can help to diagnose SAS early.Verification for genes affected by SATB2 mutations for heterogeneous manifestations may help to clarify the possible pathogenesis of SAS in the future.展开更多
本研究以绵羊睾丸组织为材料,利用RT-PCR技术获得绵羊硫氧还蛋白类蛋白2(Thioredoxin like protein 2,Txl-2)基因的CDS区序列,并对其进行生物信息学分析。结果表明,克隆得到的绵羊Txl-2基因CDS区为795 bp,编码264个氨基酸。生物信息学...本研究以绵羊睾丸组织为材料,利用RT-PCR技术获得绵羊硫氧还蛋白类蛋白2(Thioredoxin like protein 2,Txl-2)基因的CDS区序列,并对其进行生物信息学分析。结果表明,克隆得到的绵羊Txl-2基因CDS区为795 bp,编码264个氨基酸。生物信息学分析结果表明,Txl-2编码的蛋白无跨膜区、信号肽和N-糖基化位点,存在多个磷酸化位点;预测出了Txl-2蛋白的二级结构和三级结构;Clustel W方法对比绵羊Txl-2与绵羊、山羊预测序列同源性最高。展开更多
Objective To express human B7. 2 extracellular domain with prokaryote expression system and to evaluate its biological activity in vitro. Methods PCR was used to amplify the extracellular region of human B7. 2 which c...Objective To express human B7. 2 extracellular domain with prokaryote expression system and to evaluate its biological activity in vitro. Methods PCR was used to amplify the extracellular region of human B7. 2 which contained both the IgV and IgC domains. The recombinant PGEX-4T-3/hB7. 2 (IgV+C) was obtained by cloning the PCR product into a prokaryote expression plasmid PGEX-4T-3 and was transformed into the host strain of DH5-a. Tke fusion protein consisted of GST and hB7. 2(IgV+C) was identified by SDS-PAGE and Western blotting. T cell activation was observed by exposing purified T lymphocytes to the fusion protein and [3H]-TdR incorporation with the presence of the first signal imitated hy anti-CD3 antibody. Results The fusion protein GST-hB7. 2 (IgV+ C) was produced and detected in inclusive body form from engineered bacterial cells. With the first signal existed,T lymphocytes proliferated when it was co-stimulated by the fusion protein. Conclusion These results indicated that the functional human B7. 2(IgV+C) fusion protein can be produced in bacterial cells and the fusion protein displays the co-stimulatory activity in T lymphocytes activation.展开更多
OBJECTIVE:To investigate the effects of icariin on apoptosis and the expression of Fas, Fas ligand(Fas L), B cell lymphoma(Bcl-2), and Bcl-2-associated X protein(Bax) in CD4+ T lymphocytes from patients with ankylosin...OBJECTIVE:To investigate the effects of icariin on apoptosis and the expression of Fas, Fas ligand(Fas L), B cell lymphoma(Bcl-2), and Bcl-2-associated X protein(Bax) in CD4+ T lymphocytes from patients with ankylosing spondylitis.METHODS:Primary cultures of peripheral blood CD4+ T lymphocytes were established and treated with icariin at high, medium, and low doses(0.5,0.25, and 0.125 mg/mL).Sulfasalazine treated and helthy cells were used as controls.Apoptosis of treated cells was determined by flow cytometry.Reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assays were used to determine the effects of icariin on the expression of Fas, Fas L, Bcl-2, and Bax.The activity of caspase 8 and caspase 3 was determined by a colorimetric assay.RESULTS:The m RNA and protein expression of Fas,and activity of caspase 8 and caspase 3 in CD4+ T lymphocytes were increased by icariin(P < 0.05).Conversely, the m RNA and protein expression of Bcl-2 was decreased(P < 0.05).The expression of Fas L and Bax were not significantly different between groups.The proapoptotic effects of icariin were dose-dependent.CONCLUSION:Icariin induces the apoptosis of CD4 + T cells from patients with AS comparing to normal control.Therefore, the induction of apoptosis may be the likely mechanism of action of icariin's antirheumatics activities.展开更多
Nod-like receptor family pyrin domain-containing protein 12 (NLRP12) is one of the critical pattern recognition receptors which participates in the regulation of multiple inflammatory responses. Mutations in NLRP12 ca...Nod-like receptor family pyrin domain-containing protein 12 (NLRP12) is one of the critical pattern recognition receptors which participates in the regulation of multiple inflammatory responses. Mutations in NLRP12 cause exceptionally rare NLRP12-associated autoinflammatory disease (NLRP12-AID). So far, very few patients with NLRP12-AID have been identified worldwide;therefore, data on the clinical phenotype and genetic profile are limited. In this study, we reported 10 patients who presented mainly with periodic fever syndrome or arthritis. Next-generation sequencing (NGS) identified 6 heterozygous mutations of NLRP12, including 2 novel null mutations. Of the patients, some with same mutations showed different clinical features. Compared to healthy controls, the increased levels of cytokines were revealed in the patients' plasmas, as well as in the supernatants of patients’ cells stimulated with lipopolysaccharide (LPS) or tumor necrosis factor-α (TNF-α). The missense mutations did not change the protein expression;but decreased level of NLRP12 protein was shown in the null mutations. And in vitro expression assay demonstrated a truncating protein induced by the frameshift mutation. Further functional studies revealed the deleterious effect of mutations on nuclear factor-kappa B (NF-κB) signaling. Both the null and missense mutations impaired their inhibition of NF-κB activation induced by p65. Collectively, this study reported a relatively large NLRP12-AID case series. Our findings expand the clinical spectrum, and reinforce the diversity of genetic mutations and clinical phenotypes. The NLRP12-associated disorder should be considered when autoinflammatory diseases are encountered in the clinical practice, especially for patients presenting with periodic fever but no other genetic cause identified.展开更多
OBJECTIVE:To study on effects of Lichong decoction on expression of apoptosis-controlling genes,Bcl-2 and Bcl-2-associated X protein(Bax) mRNAs in hysteromyoma tissue of the hysteromyoma model rat.METHODS:Fifty Wistar...OBJECTIVE:To study on effects of Lichong decoction on expression of apoptosis-controlling genes,Bcl-2 and Bcl-2-associated X protein(Bax) mRNAs in hysteromyoma tissue of the hysteromyoma model rat.METHODS:Fifty Wistar female rats were randomly divided into a normal group,a model group,a Lichong decoction group,a Guizifuling capsule group and a Mifepristone group.The hysteromyoma rat model was established by intraperitoneal injection of exogenous estrin and progestogens.Pathological examination of uterine tissue,uterine coefficient and uterine transverse diameter were made under optic microscope and expressions of Bcl-2 and Bax mRNAs in uterine tissue in the groups were detected with real-time fluorescent quantitative polymerase chain reaction(PCR) technique.RESULTS:After treatment,under microscope it was found that in the Lichong decoction group myometrium thinned,muscle fiber slightly overgrowth or long and thin,regular arrangement,inserting phenomenon of inner circular muscle and external longitudinal muscle was occasionally or not seen in the Lichong decoction group.The uterine coefficient and the uterine transverse diameter significantly decreased(P<0.01),and Bcl-2 mRNA expression significantly decreased(P<0.01) and Bax mRNA expression significantly increased in hysteromyoma tissue(P<0.01) in the Lichong decoction group as compared with the model group.CONCLUSION:Therapeutic effects of Lichong decoction on hysteromyoma is related with decrease of Bcl-2 mRNA expression and increase of Bax mRNA expression.展开更多
OBJECTIVE:To evaluate the effects of baicalin in human gastric cancer cells, including apoptosis-inducing effects, and to investigate its underlying mechanisms of action.METHODS:Cell proliferation and apoptosis assays...OBJECTIVE:To evaluate the effects of baicalin in human gastric cancer cells, including apoptosis-inducing effects, and to investigate its underlying mechanisms of action.METHODS:Cell proliferation and apoptosis assays were performed to investigate the anti-proliferation effects of baicalin in human gastric cancer BGC-823 and MGC-803 cells.Real time-quantitative polymerase chain reaction and Western blotting analysis were performed to elucidate the molecular mechanisms underlying the anti-tumor properties of baicalin.RESULTS:In BGC-823 and MGC-803 gastric cancer cells treated with 80, 120, and 160 μmol/L baicalin for 48 h, a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay showed that baicalin significantly inhibited cell proliferation in a dose-dependent manner, while flow cytometric analysis demonstrated that baicalin could induce apoptosis, also in a dose-dependent manner.Moreover, baicalin up-regulated the expression of caspase-3, caspase-9, and B cell lymphoma(Bcl-2)-associated X protein and down-regulated the expression of Bcl-2 at both the m RNA and protein level.CONCLUSION:Baicalin has potential as a therapeutic agent for gastric cancer by inducing apoptosis in cancer cells.展开更多
In order to study the mechanism of the effect of heparin on apoptosis in carcinoma cells, the nasopharyngeal carcinoma cell line CNE2 was used to identify the effect of heparin on apoptosis associated with the express...In order to study the mechanism of the effect of heparin on apoptosis in carcinoma cells, the nasopharyngeal carcinoma cell line CNE2 was used to identify the effect of heparin on apoptosis associated with the expression of c-myc, bax, bcl-2 proteins by use of Hoechst 33258 staining, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL), agarose gel electrophoresis, and flow cytometry, as well as Western blot analysis. The results showed that heparin induced apoptosis of CNE2 cells including the morphologic changes such as reduction in the volume, and the nuclear chromatin condensation, as well as the 'ladder pattern' revealed by agarose gel electrophoresis of DNA in a concentration-dependent manner. The number of TUNEL-positive cells was dramatically increased to 33.6+/-1.2% from 2.8+/-0.3% by treatment with heparin in different concentrations (10 to approximately 40 kU/L). The apoptotic index was increased to 32.5% from 3.5% by detecting SubG1 peaks on flow cytometry. Western blot analysis showed that levels of bcl-2, bax and c-myc were significantly overexpressed by treatment with the increase of heparin concentrations. These results suggest that heparin induces apoptosis of CNE2 cells, which may be regulated by differential expression of apoptosis-related genes.展开更多
AIM: To investigate the role of the mitochondrial pathway in JTE-522-induced apoptosis and to investigate the relationship between cytochrome C release, caspase activity and loss of mitochondrial membrane potential (D...AIM: To investigate the role of the mitochondrial pathway in JTE-522-induced apoptosis and to investigate the relationship between cytochrome C release, caspase activity and loss of mitochondrial membrane potential (Deltapsim). METHODS: Cell culture, cell counting, ELISA assay, TUNEL, flow cytometry, Western blot and fluorometric assay were employed to investigate the effect of JTE-522 on cell proliferation and apoptosis in AGS cells and related molecular mechanism. RESULTS: JTE-522 inhibited the growth of AGS cells and induced the apoptosis. Caspases 8 and 9 were activated during apoptosis as judged by the appearance of cleavage products from procaspase and the caspase activities to cleave specific fluorogenic substrates. To elucidate whether the activation of caspases 8 and 9 was required for the apoptosis induction, we examined the effect of caspase-specific inhibitors on apoptosis. The results showed that caspase inhibitors significantly inhibited the apoptosis induced by JTE-522. In addition, the membrane translocation of Bax and cytosolic release of cytochrome C accompanying with the decrease of the uptake of Rhodamin 123, were detected at an early stage of apoptosis. Furthermore, Bax translocation, cytochrome C release, and caspase 9 activation were blocked by Z-VAD.fmk and Z-IETD-CHO. CONCLUSION: The present data indicate a crucial association between activation of caspases 8, 9, cytochrome C release, membrane translocation of Bax, loss of Deltapsim and JTE-522-induced apoptosis in AGS cells.展开更多
Aim: The expression of the cytokines IL-2, IL-6, IL-10, IFN-γ and TNF-α and the adhesion proteins CD99 and CD106 was studied in the human testis at the protein level. Methods: The expression of the cytokines and the...Aim: The expression of the cytokines IL-2, IL-6, IL-10, IFN-γ and TNF-α and the adhesion proteins CD99 and CD106 was studied in the human testis at the protein level. Methods: The expression of the cytokines and the adhesion proteins was assessed using immunohistochemistry and immunoblotting. Results: None of the cytokines studied was present in the human testis, but CD99 and CD106 (VCAM-1) strongly were expressed in all the testes investigated. CD99 was present in the interstitial tissue of the human testis as well as in the Sertoli cells. The identity of the CD99+ interstitial cells is unclear. CD106 (VCAM-1) was present in Leydig cells as well as the basal parts of the Sertoli cells in the seminiferous tubules. In immunoblotting, CD99 was demonstrated at molecular ratios of 46-57 (kD). This is a novel isoform of the molecule. Conclusion: The human testis produces both CD99 and CD106 and as CD106 mediates cell binding to lymphocytes, it is possible that the human Leydig cells adhere to lymphocytes like the rodent Leydig cells. (Asian J Androl 2002 Dec; 4: 243-248)展开更多
CD4 + CD25 + CD127 dim/regulatory T cells(Tregs) have been implicated in suppressing T cell immune responses to hepatitis B virus(HBV),but the inhibition mechanism has not being clear yet.This study investigated the e...CD4 + CD25 + CD127 dim/regulatory T cells(Tregs) have been implicated in suppressing T cell immune responses to hepatitis B virus(HBV),but the inhibition mechanism has not being clear yet.This study investigated the effects of soluble FGL2(sFGL2) secreted by Tregs on immune suppression in chronic HBV-infected patients.We verified that sFGL2 protein and mRNA were highly expressed in Tregs.The separated Tregs by using magnetic beads from peripheral blood mononuclear cells(PBMCs) in 20 patients with chronic hepatitis B were co-cultured with PBMCs at a ratio of 1:3 with anti-CD3 stimulating antibody or FGL2 blocking antibody.The proliferation index of CD8 + T cells after blocking FGL2 was higher than that in blank group(3.58±0.18 vs.3.28±0.17,P=0.034) in 18 of 20 samples,and lower than that in CD3 stimulation group(3.82±0.19,P=0.026) in 16 of 20 samples.The IFN-γ secreted in the mixed culture in the absence of Tregs was higher than that in the culture in the presence of Tregs,but it could be abolished by FGL2 blocking antibody.These results suggest that sFGL2 protein secreted by Tregs suppresses the proliferation and function of CD8 + T cells in chronic hepatitis B.展开更多
Objective: To investigate the effects of synthetic retinoid CD437 and acitretin on cell proliferation, apoptosis, cycle arrest and Bax/ Bcl-2 protein expression of melanoma A375 cell, Methods:MTT assay was used to d...Objective: To investigate the effects of synthetic retinoid CD437 and acitretin on cell proliferation, apoptosis, cycle arrest and Bax/ Bcl-2 protein expression of melanoma A375 cell, Methods:MTT assay was used to determine the anti-proliferative effects of CD437 and acitretin on melanoma A375 cell, Flow cytometry was performed to investigate the influence of CD437 and acitretin on cell cycle and cell apoptosis. SABC immunocytochemistry was employed for detection of Bax/bcl-2 protein expressions. Results:10^-5 mol/L CD437 was more effective than acitretin in inhibiting proliferation and inducing apoptosis of A375 cell after 24 h treatment, growth inhibiting ratio and apoptosis ratio(58.6%vs43.25% and 28.03%vs17.13%, P 〈 0.05 respectively). CD437 promoted G0/G1 arrest in melanoma A375 cell, however acitretin could not. CD437 and acitretin could up-regulate the expression of Bax protein and downregulate the expression of bcl-2 protein(P 〈 0.05). Conclusion:CD437 is more effective than acitretin in inhibiting proliferation and inducing apoptosis and cycle arrest on A375 cell, CD437 may have more potentialities than acitretin for subsidiary treatment of melanoma. Mitochondrial apoptosis pathway is partially involved in two drugs inducing apoptosis on A375 cell.展开更多
Burkholderia mallei is the etiologic agent of glanders in solipeds and humans. Lipopolysaccharide (LPS) is a major component of cell envelop of this pathogen. O-antigen, the most external component of LPS, is a virule...Burkholderia mallei is the etiologic agent of glanders in solipeds and humans. Lipopolysaccharide (LPS) is a major component of cell envelop of this pathogen. O-antigen, the most external component of LPS, is a virulence factor and a protective antigen in many pathogenic bacteria. Two putative proteins named Wzm (integral membrane protein) and Wzt (hydrophilic ATP-binding protein) are believed to make up an ABC-2 transporter of B. mallei that facilitates transport of components of O-antigen from cytosol to outer-membrane. We studied the importance of wzt (encoding Wzt) to growth, LPS O-antigen profile, and pathogenicity of B. mallei. A wzt mutant strain was generated by deleting a portion of the wzt in B. mallei wild type strain ATCC 23344 by gene replacement. Compared to the wild type strain, the wzt mutant displayed slower growth in vitro and less lethality in CD1 mice when inoculated intraperitoneally. The 50% lethal doses (LD50) of the wild type and the wzt mutant strains were 5.9 × 105 and 9.1 × 105 cfu, respectively. CD1 mice inoculated with a non-lethal dose of the wzt mutant produced specific serum immunoglobulins IgG1 and IgG2a and were partially protected against challenge with 11.2 times LD50 of the wild type strain. These findings suggest that the wzt is required for optimal in vitro growth and pathogenesis of B. mallei, and a wzt mutant protects CD1 mice against glanders.展开更多
基金Supported by Grants from Health Department of Hubei Province (JX2B09)NSFC (30771911)National 973 Key Program (2009CB522502), and Tongji Hospital
文摘AIM: To further analyze the interaction of tupaia CD81 with hepatitis C virus (HCV) envelope protein E2. METHODS: A tupaia CD81 large extracellular loop (CD81 LEL), which binds to HCV E2 protein, was cloned and expressed as a GST-fusion protein, and interaction of HCV E2 protein with a tupaia CD81 LEL was evaluated by enzyme-linked immunosorbent assay (EIA). RESULTS: Although tupaia and human CD81 LEL differed in 6 amino acid changes, tupaia CD81 LEL was strongly recognized by anti-CD81 antibodies against human CD81 LEL conformation-dependent epitopes. Investigating LEL CD81-E2 interactions by EIA, we demonstrated that binding of tupaia CD81 LEL GST fusion protein to recombinant HCV E2 protein was markedly reduced compared to binding of human CD81 LEL GST fusion protein to recombinant HCV E2 protein. CONCLUSION: These data suggest that the structural differences in-between the tupaia and human CD81 may alter the interaction of the large extracellular loop with HCV envelope glycoprotein E2. These findings may be important for the understanding of the mechanisms of binding and entry of HCV to PTHs.
文摘BACKGROUND Special AT-rich sequence binding protein 2(SATB2)-associated syndrome(SAS;OMIM 612313)is an autosomal dominant disorder.Alterations in the SATB2 gene have been identified as causative.CASE SUMMARY We report a case of a 13-year-old Chinese boy with lifelong global developmental delay,speech and language delay,and intellectual disabilities.He had short stature and irregular dentition,but no other abnormal clinical findings.A de novo heterozygous nonsense point mutation was detected by genetic analysis in exon 6 of SATB2,c.687C>A(p.Y229X)(NCBI reference sequence:NM_001172509.2),and neither of his parents had the mutation.This mutation is the first reported and was evaluated as pathogenic according to the guidelines from the American College of Medical Genetics and Genomics.SAS was diagnosed,and special education performed.Our report of a SAS case in China caused by a SATB2 mutation expanded the genotype options for the disease.The heterogeneous manifestations can be induced by complicated pathogenic involvements and functions of SATB2 from reviewed literatures:(1)SATB2 haploinsufficiency;(2)the interference of truncated SATB2 protein to wild-type SATB2;and(3)different numerous genes regulated by SATB2 in brain and skeletal development in different developmental stages.CONCLUSION Global developmental delays are usually the initial presentations,and the diagnosis was challenging before other presentations occurred.Regular follow-up and genetic analysis can help to diagnose SAS early.Verification for genes affected by SATB2 mutations for heterogeneous manifestations may help to clarify the possible pathogenesis of SAS in the future.
文摘本研究以绵羊睾丸组织为材料,利用RT-PCR技术获得绵羊硫氧还蛋白类蛋白2(Thioredoxin like protein 2,Txl-2)基因的CDS区序列,并对其进行生物信息学分析。结果表明,克隆得到的绵羊Txl-2基因CDS区为795 bp,编码264个氨基酸。生物信息学分析结果表明,Txl-2编码的蛋白无跨膜区、信号肽和N-糖基化位点,存在多个磷酸化位点;预测出了Txl-2蛋白的二级结构和三级结构;Clustel W方法对比绵羊Txl-2与绵羊、山羊预测序列同源性最高。
基金This work was supported by the National Natural Science Foundation of China(No. 39470293).
文摘Objective To express human B7. 2 extracellular domain with prokaryote expression system and to evaluate its biological activity in vitro. Methods PCR was used to amplify the extracellular region of human B7. 2 which contained both the IgV and IgC domains. The recombinant PGEX-4T-3/hB7. 2 (IgV+C) was obtained by cloning the PCR product into a prokaryote expression plasmid PGEX-4T-3 and was transformed into the host strain of DH5-a. Tke fusion protein consisted of GST and hB7. 2(IgV+C) was identified by SDS-PAGE and Western blotting. T cell activation was observed by exposing purified T lymphocytes to the fusion protein and [3H]-TdR incorporation with the presence of the first signal imitated hy anti-CD3 antibody. Results The fusion protein GST-hB7. 2 (IgV+ C) was produced and detected in inclusive body form from engineered bacterial cells. With the first signal existed,T lymphocytes proliferated when it was co-stimulated by the fusion protein. Conclusion These results indicated that the functional human B7. 2(IgV+C) fusion protein can be produced in bacterial cells and the fusion protein displays the co-stimulatory activity in T lymphocytes activation.
基金Supported by the Fundamental Research Funds for the Central Public Welfare Research Institutes:Pharmacogenomics of Tripterygium wilfordii Hook F response and liver toxic reaction in Rheumatoid Arthritis(No.ZZ0708079)the Fundamental Research Funds for the Central Public Welfare Research Institutes Pharmacogenomics of Tripterygium wilfordii Hook F response reaction in Rheumatoid Arthritis(No.31470962)
文摘OBJECTIVE:To investigate the effects of icariin on apoptosis and the expression of Fas, Fas ligand(Fas L), B cell lymphoma(Bcl-2), and Bcl-2-associated X protein(Bax) in CD4+ T lymphocytes from patients with ankylosing spondylitis.METHODS:Primary cultures of peripheral blood CD4+ T lymphocytes were established and treated with icariin at high, medium, and low doses(0.5,0.25, and 0.125 mg/mL).Sulfasalazine treated and helthy cells were used as controls.Apoptosis of treated cells was determined by flow cytometry.Reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assays were used to determine the effects of icariin on the expression of Fas, Fas L, Bcl-2, and Bax.The activity of caspase 8 and caspase 3 was determined by a colorimetric assay.RESULTS:The m RNA and protein expression of Fas,and activity of caspase 8 and caspase 3 in CD4+ T lymphocytes were increased by icariin(P < 0.05).Conversely, the m RNA and protein expression of Bcl-2 was decreased(P < 0.05).The expression of Fas L and Bax were not significantly different between groups.The proapoptotic effects of icariin were dose-dependent.CONCLUSION:Icariin induces the apoptosis of CD4 + T cells from patients with AS comparing to normal control.Therefore, the induction of apoptosis may be the likely mechanism of action of icariin's antirheumatics activities.
基金supported in part by the National Key Research and Development Program of China(No.2021YFC2702005)National Natural Science Foundation of China(No.81971547)+1 种基金the Research Fund for Outstanding Youth Scholar of Chongqing Talents(No.CQYC201905003)the High-level Medical Reserved Personnel Training Project of Chongqing(No.2019181).
文摘Nod-like receptor family pyrin domain-containing protein 12 (NLRP12) is one of the critical pattern recognition receptors which participates in the regulation of multiple inflammatory responses. Mutations in NLRP12 cause exceptionally rare NLRP12-associated autoinflammatory disease (NLRP12-AID). So far, very few patients with NLRP12-AID have been identified worldwide;therefore, data on the clinical phenotype and genetic profile are limited. In this study, we reported 10 patients who presented mainly with periodic fever syndrome or arthritis. Next-generation sequencing (NGS) identified 6 heterozygous mutations of NLRP12, including 2 novel null mutations. Of the patients, some with same mutations showed different clinical features. Compared to healthy controls, the increased levels of cytokines were revealed in the patients' plasmas, as well as in the supernatants of patients’ cells stimulated with lipopolysaccharide (LPS) or tumor necrosis factor-α (TNF-α). The missense mutations did not change the protein expression;but decreased level of NLRP12 protein was shown in the null mutations. And in vitro expression assay demonstrated a truncating protein induced by the frameshift mutation. Further functional studies revealed the deleterious effect of mutations on nuclear factor-kappa B (NF-κB) signaling. Both the null and missense mutations impaired their inhibition of NF-κB activation induced by p65. Collectively, this study reported a relatively large NLRP12-AID case series. Our findings expand the clinical spectrum, and reinforce the diversity of genetic mutations and clinical phenotypes. The NLRP12-associated disorder should be considered when autoinflammatory diseases are encountered in the clinical practice, especially for patients presenting with periodic fever but no other genetic cause identified.
基金Supported by Beijing City Natural Science Fund (No.7082015)National Natural Science Fund (No. 81073096)"Middle-young Aged Core Talent Cultural Plant" of Beijing City University Talent Strengthening Education Plant (No.PHR201008403)
文摘OBJECTIVE:To study on effects of Lichong decoction on expression of apoptosis-controlling genes,Bcl-2 and Bcl-2-associated X protein(Bax) mRNAs in hysteromyoma tissue of the hysteromyoma model rat.METHODS:Fifty Wistar female rats were randomly divided into a normal group,a model group,a Lichong decoction group,a Guizifuling capsule group and a Mifepristone group.The hysteromyoma rat model was established by intraperitoneal injection of exogenous estrin and progestogens.Pathological examination of uterine tissue,uterine coefficient and uterine transverse diameter were made under optic microscope and expressions of Bcl-2 and Bax mRNAs in uterine tissue in the groups were detected with real-time fluorescent quantitative polymerase chain reaction(PCR) technique.RESULTS:After treatment,under microscope it was found that in the Lichong decoction group myometrium thinned,muscle fiber slightly overgrowth or long and thin,regular arrangement,inserting phenomenon of inner circular muscle and external longitudinal muscle was occasionally or not seen in the Lichong decoction group.The uterine coefficient and the uterine transverse diameter significantly decreased(P<0.01),and Bcl-2 mRNA expression significantly decreased(P<0.01) and Bax mRNA expression significantly increased in hysteromyoma tissue(P<0.01) in the Lichong decoction group as compared with the model group.CONCLUSION:Therapeutic effects of Lichong decoction on hysteromyoma is related with decrease of Bcl-2 mRNA expression and increase of Bax mRNA expression.
基金Supported by the Natural Science Foundation of Gansu Province(No.148RJZA073)the Health Industry Scientific Research Project of Gansu Province(No.GWGL 2013-40)the Openning Plan Project of Key Laboratory for Transfer of Dunhuang Medicine at the Provincial and Ministerial Level(No.DHYX1213-008)
文摘OBJECTIVE:To evaluate the effects of baicalin in human gastric cancer cells, including apoptosis-inducing effects, and to investigate its underlying mechanisms of action.METHODS:Cell proliferation and apoptosis assays were performed to investigate the anti-proliferation effects of baicalin in human gastric cancer BGC-823 and MGC-803 cells.Real time-quantitative polymerase chain reaction and Western blotting analysis were performed to elucidate the molecular mechanisms underlying the anti-tumor properties of baicalin.RESULTS:In BGC-823 and MGC-803 gastric cancer cells treated with 80, 120, and 160 μmol/L baicalin for 48 h, a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay showed that baicalin significantly inhibited cell proliferation in a dose-dependent manner, while flow cytometric analysis demonstrated that baicalin could induce apoptosis, also in a dose-dependent manner.Moreover, baicalin up-regulated the expression of caspase-3, caspase-9, and B cell lymphoma(Bcl-2)-associated X protein and down-regulated the expression of Bcl-2 at both the m RNA and protein level.CONCLUSION:Baicalin has potential as a therapeutic agent for gastric cancer by inducing apoptosis in cancer cells.
文摘In order to study the mechanism of the effect of heparin on apoptosis in carcinoma cells, the nasopharyngeal carcinoma cell line CNE2 was used to identify the effect of heparin on apoptosis associated with the expression of c-myc, bax, bcl-2 proteins by use of Hoechst 33258 staining, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL), agarose gel electrophoresis, and flow cytometry, as well as Western blot analysis. The results showed that heparin induced apoptosis of CNE2 cells including the morphologic changes such as reduction in the volume, and the nuclear chromatin condensation, as well as the 'ladder pattern' revealed by agarose gel electrophoresis of DNA in a concentration-dependent manner. The number of TUNEL-positive cells was dramatically increased to 33.6+/-1.2% from 2.8+/-0.3% by treatment with heparin in different concentrations (10 to approximately 40 kU/L). The apoptotic index was increased to 32.5% from 3.5% by detecting SubG1 peaks on flow cytometry. Western blot analysis showed that levels of bcl-2, bax and c-myc were significantly overexpressed by treatment with the increase of heparin concentrations. These results suggest that heparin induces apoptosis of CNE2 cells, which may be regulated by differential expression of apoptosis-related genes.
基金National Natural Science Foundation of China,No.39770300,30070873the Overseas Chinese Affairs Office of the State Council Foundation,No.98-33
文摘AIM: To investigate the role of the mitochondrial pathway in JTE-522-induced apoptosis and to investigate the relationship between cytochrome C release, caspase activity and loss of mitochondrial membrane potential (Deltapsim). METHODS: Cell culture, cell counting, ELISA assay, TUNEL, flow cytometry, Western blot and fluorometric assay were employed to investigate the effect of JTE-522 on cell proliferation and apoptosis in AGS cells and related molecular mechanism. RESULTS: JTE-522 inhibited the growth of AGS cells and induced the apoptosis. Caspases 8 and 9 were activated during apoptosis as judged by the appearance of cleavage products from procaspase and the caspase activities to cleave specific fluorogenic substrates. To elucidate whether the activation of caspases 8 and 9 was required for the apoptosis induction, we examined the effect of caspase-specific inhibitors on apoptosis. The results showed that caspase inhibitors significantly inhibited the apoptosis induced by JTE-522. In addition, the membrane translocation of Bax and cytosolic release of cytochrome C accompanying with the decrease of the uptake of Rhodamin 123, were detected at an early stage of apoptosis. Furthermore, Bax translocation, cytochrome C release, and caspase 9 activation were blocked by Z-VAD.fmk and Z-IETD-CHO. CONCLUSION: The present data indicate a crucial association between activation of caspases 8, 9, cytochrome C release, membrane translocation of Bax, loss of Deltapsim and JTE-522-induced apoptosis in AGS cells.
基金Correspondence to: Dr. Esko Verijnkorva, University of Turku, Institute of Biomedicine, Department of Anatomy, Kiinamyllynkatu 10, FIN-20520 Turku, Finland.
文摘Aim: The expression of the cytokines IL-2, IL-6, IL-10, IFN-γ and TNF-α and the adhesion proteins CD99 and CD106 was studied in the human testis at the protein level. Methods: The expression of the cytokines and the adhesion proteins was assessed using immunohistochemistry and immunoblotting. Results: None of the cytokines studied was present in the human testis, but CD99 and CD106 (VCAM-1) strongly were expressed in all the testes investigated. CD99 was present in the interstitial tissue of the human testis as well as in the Sertoli cells. The identity of the CD99+ interstitial cells is unclear. CD106 (VCAM-1) was present in Leydig cells as well as the basal parts of the Sertoli cells in the seminiferous tubules. In immunoblotting, CD99 was demonstrated at molecular ratios of 46-57 (kD). This is a novel isoform of the molecule. Conclusion: The human testis produces both CD99 and CD106 and as CD106 mediates cell binding to lymphocytes, it is possible that the human Leydig cells adhere to lymphocytes like the rodent Leydig cells. (Asian J Androl 2002 Dec; 4: 243-248)
基金supported by National 973 Project in Viral Hepatitis,China (MOST,No. 2007CB512900)
文摘CD4 + CD25 + CD127 dim/regulatory T cells(Tregs) have been implicated in suppressing T cell immune responses to hepatitis B virus(HBV),but the inhibition mechanism has not being clear yet.This study investigated the effects of soluble FGL2(sFGL2) secreted by Tregs on immune suppression in chronic HBV-infected patients.We verified that sFGL2 protein and mRNA were highly expressed in Tregs.The separated Tregs by using magnetic beads from peripheral blood mononuclear cells(PBMCs) in 20 patients with chronic hepatitis B were co-cultured with PBMCs at a ratio of 1:3 with anti-CD3 stimulating antibody or FGL2 blocking antibody.The proliferation index of CD8 + T cells after blocking FGL2 was higher than that in blank group(3.58±0.18 vs.3.28±0.17,P=0.034) in 18 of 20 samples,and lower than that in CD3 stimulation group(3.82±0.19,P=0.026) in 16 of 20 samples.The IFN-γ secreted in the mixed culture in the absence of Tregs was higher than that in the culture in the presence of Tregs,but it could be abolished by FGL2 blocking antibody.These results suggest that sFGL2 protein secreted by Tregs suppresses the proliferation and function of CD8 + T cells in chronic hepatitis B.
基金supported by the National Natural Science Foundation of China(No.30371295).
文摘Objective: To investigate the effects of synthetic retinoid CD437 and acitretin on cell proliferation, apoptosis, cycle arrest and Bax/ Bcl-2 protein expression of melanoma A375 cell, Methods:MTT assay was used to determine the anti-proliferative effects of CD437 and acitretin on melanoma A375 cell, Flow cytometry was performed to investigate the influence of CD437 and acitretin on cell cycle and cell apoptosis. SABC immunocytochemistry was employed for detection of Bax/bcl-2 protein expressions. Results:10^-5 mol/L CD437 was more effective than acitretin in inhibiting proliferation and inducing apoptosis of A375 cell after 24 h treatment, growth inhibiting ratio and apoptosis ratio(58.6%vs43.25% and 28.03%vs17.13%, P 〈 0.05 respectively). CD437 promoted G0/G1 arrest in melanoma A375 cell, however acitretin could not. CD437 and acitretin could up-regulate the expression of Bax protein and downregulate the expression of bcl-2 protein(P 〈 0.05). Conclusion:CD437 is more effective than acitretin in inhibiting proliferation and inducing apoptosis and cycle arrest on A375 cell, CD437 may have more potentialities than acitretin for subsidiary treatment of melanoma. Mitochondrial apoptosis pathway is partially involved in two drugs inducing apoptosis on A375 cell.
文摘Burkholderia mallei is the etiologic agent of glanders in solipeds and humans. Lipopolysaccharide (LPS) is a major component of cell envelop of this pathogen. O-antigen, the most external component of LPS, is a virulence factor and a protective antigen in many pathogenic bacteria. Two putative proteins named Wzm (integral membrane protein) and Wzt (hydrophilic ATP-binding protein) are believed to make up an ABC-2 transporter of B. mallei that facilitates transport of components of O-antigen from cytosol to outer-membrane. We studied the importance of wzt (encoding Wzt) to growth, LPS O-antigen profile, and pathogenicity of B. mallei. A wzt mutant strain was generated by deleting a portion of the wzt in B. mallei wild type strain ATCC 23344 by gene replacement. Compared to the wild type strain, the wzt mutant displayed slower growth in vitro and less lethality in CD1 mice when inoculated intraperitoneally. The 50% lethal doses (LD50) of the wild type and the wzt mutant strains were 5.9 × 105 and 9.1 × 105 cfu, respectively. CD1 mice inoculated with a non-lethal dose of the wzt mutant produced specific serum immunoglobulins IgG1 and IgG2a and were partially protected against challenge with 11.2 times LD50 of the wild type strain. These findings suggest that the wzt is required for optimal in vitro growth and pathogenesis of B. mallei, and a wzt mutant protects CD1 mice against glanders.