Local IL-23 Expression in Murine Vaginal Candidiasis and Its Relationship with Infection and Immune Status

IL-12 is a heterodi meric cytokine which con-sist of p40 and p35 and plays a key role in the dif-ferentiation of T helper1 ( Th1)cells[1].Its func-tions in murine vaginal candidiasis have beenrepor-ted[2 ,3].IL-23 is a new member inthe IL-12 familyof heterodi meric cytokines which is composed ofp40 and p19[4]. IL-23 shares some features withIL-12 but they also differ ,functionally and struc-turally . Many of its biological features ,such as itsroleinthe development of murine vaginal candid…     

Targeting of p38 Mitogen-activated Protein Kinases to Early Growth Response gene 1 （EGR-1） in the Human Paclitaxel-resistance Ovarian Carcinoma Cells

To investigate the relationship between the expression of early growth response gene 1 (EGR-1) and p38MAPK pathway in the paclitaxel resistance of ovarian carcinoma cells, the effect of p38MAPK inhibitor SB203580 on cell apoptosis was examined by using Hoechst 33258 staining. The intracellular Rh123 (Rhodamine 123) accumulation was detected by the flow cytometry (FCM). The 50% inhibition concentration (IC50) of paclitaxel for A2780/Taxol cells was determined by MTT method. Electrophoretic motility shift assay (EMSA) was employed to examine the EGR-1DNA binding activity. MDR1 and EGR-1 mRNA were assessed by RT-PCR. The expressed of p-gp, phos- phorylated p53 and p38 were detected by Western blotting. SB203580 could remarkably promote the apoptosis of A2780/Taxol cells, and the cell apoptosis was in a time-dependent manner. Cellular Rh123 accumulation was increased, and the IC50 of paclitaxel for A2780/Taxol cells was decreased significantly. A2780/Taxol cell line after SB203580 treatment was shown to have a significantly higher level of EGR-1 DNA binding activity. SB203580 down-regulated the activity of p38MAPK pathway, but up-regulated EGR-1 expression. SB203580 significantly increased the level of cellular phosphorylated p53 protein, but decreased the p-gp protein level and MDR1 mRNA level in A2780/Taxol cells. There existed a close relationship between p38MAPK pathway and the paclitaxel resistance of ovarian carcinoma cells. The expression of EGR-1 mediated by p38MAPK pathway plays a critical role in paclitaxel resistance of ovarian carcinoma cells.     

CLONING AND EXPRESSION OF THE GENE CODING FOR IL-2 （60）-PE40, A MOLECULAR TARGETED PROTEIN

It has recently been shown that chimeric toxin composed of IL2 fused tp PE40,a mutant form of Pseu-domonas Exotoxin A devoid of its native cell recognition and binding domain was cytotoxic to IL-2 receptor bearing cells.We here amplified the gene IL-2 (60),which codes for the N-terminal 1-60 amino acids of human IL-2 by PCR.After that,we fused it to PE40 and the new chimteric protein IL-2 (60)-PE40 was expressed in E.coli.SDS-PAGE revealed that IL-2(60)-PE40 chimeric protein accounts for more than 18% of total cell proteins.As the region IL-2 binds with its receptor was defined in the N-terminal residuse 8-54 of IL-2,such fusion proteins will have the same activity with IL-2-PE40.Following primany purification,IL-2(60)-PE40 was shown to be very toxic to IL-2 receptor-positive cells but non measurable effect on the cells lacking IL-2 receptors.Such a structure has not been reported by now.The fusion pro-tein is useful for suppressing the immune response in cases of rejection of allografts and organ transplants and as therapeutic agents for the treatment of IL-2 receptor related diseases such as autoimmune disease,and as therapeutic agents for the treatment of IL-2 receptor related diseases such as autoimmune disease,ATL(adult T-cell leukemia),et al.     

IL-1α and ATP mRNA expression after ultraviolet irradiation in human keratinocyte original-SCC 12F cells

Background Generally speaking, undifferentiated keratinocytes may synthesize a larger amount of IL-α and its production is decreased as cells complete differentiation gradually. Ultraviolet B (UVB)light can signigicantly stimulate the production and release of some cytokines. In this study we investigated the influence of UVB irradiation on IL-1α and adenosine triphosphoric (ATP) mRNA expressions in the human keratinocyte (KC) of original squamous cell carcinoma line (SCC 12F cells).Methods The cultured SCC 12F cells were irradiated with 30 mJ/cm^2 of UVB. Northern blot was employed to analyze the expression of IL-1α and ATP mRNA.Results There was a constitutive expression of IL-1α mRNA in SCC 12F cells. The expression increased in culturing time in regular KC medium and reached the highest expression at 120 hours.The expression level of IL-1α was up-regulated with two peaks at 6 hours and 72 hours, respectively after UVB irradiation. In comparison with IL-1α mRNA expression, ATP mRNA was down-regulated,with similar biphasic peaks, compared with the sham irradiated group.Conclusions SCC12F cells may express IL-1α mRNA constitutively. After UVB irradiation, the mRNA expression of IL-1α and ATP will show opposite effect because of inflammation/immunity and energy consumption mechanisms.     

EFFECT OF INTERLEUKIN-1β ON GROWTH HORMONE GENE EXPRESSION AND ITS POSSIBLE MOLECULAR MECHANISM IN RAT MtT/S SOMATOTROPH CELLS

Objective To elucidate the effect of interleukin-1β (IL-1β) on human growth hormone (hGH) gene expression in a rat somatotropic pituitary cell line MtT/S. Methods Stably transfected MtT/S cells were firstly established by transfecting 484-Luc1 plasmid which contained hGH gene promoter -484 to 30 bp and luciferase reporter gene. The effect of IL-1β on hGH gene expression was determined by assaying the luciferase activities. RT-PCR method was also used to determine whether IL-1 recepor mRNA was expressed in MtT/S cells. Results The 103 U/mL IL-1β stimulated secretion and synthesis of GH, and promoted the 5’-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.38 times above the control. Among inhibitors of signaling transduction pathways, mitogen-activated protein kinase kinase (MAPKK/MEK) inhibitor PD98059 (40 μmol/L) and p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 (5 μmol/L) completely blocked the stimulatory effect of IL-1β, and phosphatidylinositol-3-kinase (PI3-K) inhibitor LY294002 partly abolished the effect of IL-1β. Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells. Neither over-expression of Pit-1 nor inhibition of Pit-1 expression affected induction of hGH promoter activity by IL-1β. A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-1β, and results showed that the stimulatory effect of IL-1β was abolished following deletion of the -196 to -132 bp fragment. Conclusions IL-1β promotes GH secretion and synthesis in rat MtT/S somatotroph cells. The stimulatory effect of IL-1β on hGH gene promoter appears to require the activation of MEK, p38 MAPK, PI3-K, and a fragment of promoter sequence that spans the –196 to –132 bp of the gene, but it may be unlinked with Pit-1 protein.     

C-reactive protein decreases interleukin-8 production in human endothelial progenitor cells by inhibition of p38 MAPK pathway

Background C-reactive protein （CRP） has been reported to damage the vascular wall by inducing endothelial dysfunction and inflammation, and it is also speculated to have a role in attenuating angiogenic functions of human endothelial progenitor cells （EPCs）. Interleukin-8 （IL-8） is an important mediator of the paracrine mitogenic effect of EPCs which has direct angiogenic effects on mature endothelial cells. We, herein, investigated the direct effect of CRP on IL-8 production and gene expression in cultured human EPCs. Methods EPCs were isolated from the peripheral venous blood of healthy male volunteers. Cells were cultured in EndoCultTM liquid medium in the absence and presence of CRP at clinically relevant concentrations （5 to 25 μg/ml） for different durations （3 to 48 hours）. IL-8 protein and mRNA of cultured EPCs were evaluated using ELISA and real-time PCR. Results The results showed that CRP at a concentration of 10 μg/ml significantly reduced IL-8 secretion of cultured EPCs with a peak at 25 μg/ml, and also decreased mRNA expression in EPCs with a peak at 12 hours. In addition, preincubation of EPCs with SB203580, an inhibitor of p38 mitogen-activated protein kinase （MAPK） decreased CRP inhibition of IL-8 mRNA expression at 12 hours in EPCs. Conclusions Our study, for the first time, demonstrates that CRP directly inhibits EPCs IL-8 secretion, a key cytokine player of angiogenesis induced by EPCs. Inhibition occurred in part via an effect of CRP to active the p38 MAPK signal transduction pathway in EPC. The ability of CRP to inhibit EPCs IL-8 secretion may represent an important mechanism that further links inflammation to cardiovascular disease.     

IL-12p40、IL-12p70在经典型霍奇金淋巴瘤中的表达及意义

目的 探讨白细胞介素-12p40(IL-12p40)、白细胞介素-12p70(IL-12p70)在经典型霍奇金淋巴瘤(CHL)组织中的表达及意义. 方法 应用免疫组织化学法检测IL-12p40、IL-12p70在41 例CHL组织、92 例弥漫性大B细胞淋巴瘤(DLBCL)组织及20 例反应性淋巴组织增生(RLH)中的表达. 结果 IL-12p40、IL-12p70在41例CHL及92例DLBCL中的阳性表达率分别是 51. 22%、70. 73% 及 0. 00%、4. 35%,20 例 RLH 中未见 IL-12p40 及 IL-12p70 表达. IL-12p40、IL-12p70在CHL 中及各亚型中的表达均明显高于DLBCL及RLH(P<0. 05). IL-12p40及IL-12p70的表达与患者性别、年龄、肿块大小、B症状、临床分期、血清乳酸脱氢酶水平、国际预后评分等临床病理因素均无关. IL-12p40与IL-12p70在CHL中的表达有相关性(P<0. 05). 结论 IL-12p40、IL-12p70在CHL发病中有重要作用.     

雷公藤多苷对结肠炎小鼠结肠组织IL-23、IL-17和IL-12表达的影响

目的:观察雷公藤多苷( GTT)、美沙拉嗪对三硝基苯磺酸(TNBS)诱导的急性结肠炎小鼠结肠组织中IL-23、IL-17、IL-12表达的影响,探讨其可能的作用机制.方法:C57BL/6小鼠随机分为4组:正常对照组、模型组、美沙拉嗪组、雷公藤多苷组,后3组应用TNBS建立小鼠急性结肠炎模型,模型组不作其他处理,雷公藤多苷组和美沙拉嗪组于造模前4 d开始每天给予雷公藤多苷灌胃或美沙拉嗪灌肠液灌肠直到实验结束,正常对照组不作任何特殊处理.各组小鼠均于TNBS灌肠后48 h处死,检测各组小鼠结肠组织大体、组织学损伤评分及髓过氧化物酶(MPO)活性;ELISA法检测结肠组织IL-23 p19、IL-17蛋白含量,实时荧光定量RT-PCR检测结肠组织IL-23 p19、IL-17、IL-12 p35的mRNA表达.结果:美沙拉嗪组和雷公藤多苷组小鼠结肠大体及组织学损伤记分、MPO活性明显低于模型组(P<0.05);模型组结肠组织IL-23 p19、IL-17、IL-12 p35 mRNA表达水平明显高于正常对照组、美沙拉嗪组、雷公藤多苷组(P<0.05);模型组结肠组织IL-23 p19、IL-17蛋白水平明显高于正常对照组、美沙拉嗪组、雷公藤多苷组(P<0.05),后三者间无统计学差异.结论:雷公藤多苷可能通过非选择性抑制IL-23 p19、IL-17、IL-12 p35的表达来抑制结肠炎小鼠的炎症反应,效果与美沙拉嗪类似.     

A study on the expression of interleukin(IL)-10 and IL-12 P35, P40 mRNA in the psoriatic lesions

Psoriasisisa common chronic,relapsing and in-flammatory disease and its pathogenesis is still un-known.It is considered thatit is one kind of geneticand immunological diseases controlled by multiplegenes.Even though theexactpathogenesisisstillun-known,many studies showed that T cells mediatedimmune response played a very important role in thepathogenesis of psoriasis.In this study,the expres-sion of IL- 1 0 ,IL- 1 2 P35 and P40 m RNA in the pso-riasis lesions were detected by using RT- P…     

胸腺五肽对复发性外阴阴道念珠菌病患者IL-12p70和IL-23p19水平的影响及疗效分析

目的研究观察胸腺五肽对复发性外阴阴道念珠菌病患者阴道分泌物IL-12p70和IL-23p19水平的影响及疗效。方法采用酶联免疫吸附实验（ELISA）双抗体夹心法检测复发性外阴阴道念珠菌病患者治疗组、对照组治疗前后及正常人阴道分泌物IL-12p70和IL-23p19水平。结果两组治疗前复发性外阴阴道念珠菌病患者IL-12p70和IL-23p19水平均低于正常人（P〈0.01）,两组患者间IL-12p70和IL-23p19水平相近（P〉0.05）。治疗后胸腺五肽组IL-12p70和IL-23p19水平升高,对照组治疗前后水平差异无统计学意义（P〉0.05）。治疗组的疗效（总有效率80.00%）明显高于对照组的43.33%（P〈0.05）。结论复发性外阴阴道念珠菌病患者存在着Th1/Th2型细胞因子失衡,胸腺五肽可通过纠正两型细胞因子失衡而改善患者免疫功能,提高治疗效果。     

凋亡细胞调控巨噬细胞表达IL-12家族细胞因子的水平

［摘要］ 目的观察凋亡细胞对巨噬细胞中白细胞介素12 （interleukin-12,IL-12）家族细胞因子表达的影响。 方法将小鼠巨噬细胞系RAW264.7（RAW 细胞）和小鼠腹腔巨噬细胞分别分为空白对照组、脂多糖（lipopolysaccharides,LPS）刺激组（LPS组）、凋亡细胞刺激组（apo组）、脂多糖+凋亡细胞刺激组（LPS+apo组）,实时定量PCR检测各组细胞刺激前后mIL-12p35、mIL-12p40、mIL-23p19水平。采用酶联免疫吸附测定方法检测各组激活的RAW细胞培养液中IL-10、前列腺素E2（prostaglandin E2,PGE2）和转化生长因子β（transforming growth factor-β,TGF-β）的含量。 结果RAW细胞和小鼠腹腔巨噬细胞LPS组IL-12p35、IL-12p40、IL-23p19 mRNA表达量高于空白对照组和apo组,LPS+apo组IL-12p35、IL-12p40、IL-23p19 mRNA表达量高于空白对照组和apo组,低于LPS组（P＜0.05）。LPS组和LPS+apo组IL-10、PGE2、TGF-β浓度均高于对照组,LPS+apo组TGF-β浓度高于对照组和LPS组（P＜0.05）;LPS组和LPS+apo组IL-10、PGE2浓度差异无统计学意义（P＞0.05）。 结论凋亡细胞抑制活化巨噬细胞IL-12p35、IL-12p40、IL-23p19 mRNA的表达。凋亡细胞对IL-10和PGE2分泌无影响,但促进TGF-β的分泌。     

2型糖尿病患者IL-23p19 mRNA及血浆IL-23水平的变化和意义

目的探讨2型糖尿病(T2DM)治疗前后外周血单个核细胞中IL-23p19 mRNA水平以及血浆中IL-23水平的变化。方法检测空腹血糖水平选定分组对象,采用逆转录-聚合酶链式反应(RT-PCR)法检测58例2型糖尿病患者(初诊糖尿病患者28例,经治疗患者30例)和26例健康体检者(对照组)全血中IL-23p19 mRNA的水平,同时采用酶联免疫吸附试验(ELISA)检测各组血浆中IL-23的水平。结果初诊糖尿病组IL-23 mRNA表达水平高于经治疗组和对照组,差异具有统计学意义(P<0.05),IL-23血浆水平比较初诊糖尿病组与经治疗组及对照组差异有统计学意义(P<0.05)。结论 2型糖尿病治疗前IL-23及p19 mRNA水平明显高于经治疗组和对照组,IL-23可能与2型糖尿病的发病和病程进展有一定的关系。     

猪白细胞介素12 p35、p40亚基基因的克隆和序列分析

目的:克隆猪白细胞介素(IL)-12p35、p40亚基基因,为制备猪囊尾蚴的复合基因疫苗奠定基础。方法:分别分离成年猪的外周血单个核细胞和脾细胞,培养10h后用IFN-γ(100ng/ml)刺激增殖12h,随后加入细菌脂多糖(1μg/ml)刺激培养3h,分别收集细胞并提取细胞总RNA,用RT-PCR方法扩增猪IL-12p35、p40cDNA编码基因,克隆至pMD18-T载体,经PCR和双酶切鉴定后进行序列测定。结果:猪IL-12p35、p40cDNAPCR产物电泳结果证明所克隆的基因分别为616bp和1011bp,基因测序结果与GenBank报道的基因序列各有一个碱基差异,但编码氨基酸无差异,证实分别为猪IL-12p35、p40cDNA编码基因。结论:成功克隆了猪IL-12p35、p40cDNA编码基因。     

IL-23/IL-17轴因子在初诊Graves病患者治疗前后的变化

目的 检测IL-23/IL-17轴在Graves病(GD)患者外周血中的表达水平,以及1311I或抗甲状腺药物(ATD)治疗后的变化趋势.方法 收集初诊GD患者40例作为GD组,其中131I治疗组(20例)和ATD治疗组(20例);40例性别、年龄匹配的健康者设为对照组.使用ELISA法检测GD组治疗前后及对照组IL-17、IL-23水平.结果 ①两组比较,治疗前GD组血清IL-23、IL-17表达水平明显高于对照组(P＜0.05);②131I治疗后第6个月IL-23、IL-17的外周血水平较治疗前有明显降低(P＜0.05),仍明显高于对照组(P＜0.05).ATD组治疗前后差异无统计学意义.结论 IL-23、IL-17在初诊GD患者外周血中高表达,提示IL-23/IL-17轴可能参与Graves发病,同时放射性131 I治疗甲亢改善病情可能与IL-23/IL-17轴因子变化部分相关.     

寻常型银屑病患者血清中IL-17的表达及紫草素对IL-17刺激HaCaT细胞分泌IL-6和IL-23的影响

目的：探讨寻常型银屑病患者血清中白细胞介素17（IL-17）的表达水平及IL-17刺激后HaCaT细胞分泌白细胞介素6（IL-6）和白细胞介素23（IL-23）的变化,阐明其临床意义及紫草素的干预作用。方法：以25名正常对照者、29例寻常型银屑病患者和不同组别HaCaT细胞（空白对照组、IL-17刺激24h组、IL-17刺激36h组、IL-17刺激48h组、紫草素+IL-17组、环孢素A+IL-17组和IL-17组）为研究对象,采用双抗体夹心酶联免疫吸附法（ELISA）检测寻常型银屑病患者血清IL-17水平和HaCaT细胞各组上清液中IL-6、IL-23水平;实时荧光免疫定量-聚合酶链式反应法（RT-PCR）检测各组HaCaT细胞中IL-6和IL-23p19 mRNA表达水平;采用细胞计数盒-8（CCK-8）法检测各组HaCaT细胞活力。结果：与正常对照组比较,银屑病组患者血清IL-17水平明显升高,以重度皮损银屑病组升高最为明显（P < 0.05）;IL-17刺激24、36和48h组HaCaT细胞及其培养上清中IL-6和IL-23水平及其mRNA表达水平均高于空白对照组（P < 0.01）;紫草素+IL-17组和环孢素A+IL-17组HaCaT细胞及其培养上清中IL-6和IL-23水平及其mRAN表达水平均低于IL-17组（P < 0.05）。与空白对照组比较,其他各组细胞活力差异无统计学意义（P > 0.05）。结论：银屑病患者血清IL-17表达水平升高,尤其是重度皮损银屑病患者血清IL-17表达水平升高明显,IL-17可促进HaCaT细胞分泌IL-6和IL-23,呈时间依赖性,紫草素可抑制IL-17的促炎症作用。