STAT3在溃疡性结肠炎患者肠黏膜的表达及活化增强

目的 检测溃疡性结肠炎（UC）患者结肠黏膜STAT3、pSTAT3蛋白及STAT3 mRNA的表达及与UC疾病活动度的关系。方法 47例UC患者根据内镜及临床活动度进行分级，收集正常对照结肠黏膜标本13例；用免疫组织化学法检测STAT3、pSTAT3在结肠黏膜的定位；用Western blot及RT-PCR法检测STAT3、pSTAT3蛋白及STAT3 mRNA在结肠黏膜的表达。结果 ⑴ STAT3为胞质着色；pSTAT3主要为胞核着色，胞质中也有表达，二者均主要表达于大量浸润的单个核细胞，且二者在UC患者黏膜的阳性细胞数及染色强度均有随病变的加重逐渐增加的趋势；⑵UC患者肠黏膜STAT3、pSTAT3蛋白的表达量高于对照者，且随着病变分级逐渐增加。⑶UC患者肠黏膜STAT3 mRNA的表达与蛋白表达一致。结论 UC患者结肠黏膜中STAT及pSTAT3的表达随病变程度逐渐增加，在一定程度上可以反映疾病的活动度。     

大鼠IgA肾病模型肾脏变化分析

目的探讨IgA肾病(IgA nephropathy,IgAN)大鼠肾小体、肾小囊、肾小球和近端肾小管病理变化。方法将20只SD雌性大鼠随机分成2组,即对照组和IgAN组(n=10)。用免疫荧光、HE染色和体视学方法,测出2组动物肾小体、肾小囊和肾小球体密度、球囊体积比、肾小体数密度、肾小体和肾小球长径、近端肾小管管腔和管壁面积及其比值,比较其差异。结果IgA免疫荧光染色,IgAN组皮质部见较强绿色荧光。与对照组比,IgAN组肾小体体密度增大,肾小囊体密度增大,肾小球体密度减小,球囊比减小,肾小体数密度变化差异无显著性,肾小体长径增大,肾小球长径减小,近端肾小管管腔面积减小,管壁面积增大,腔壁比减小。结论 IgAN大鼠肾小体和肾小囊增大,肾小球减小,近曲小管细胞体积增大,管腔变小。     

威灵仙总皂苷抑制佐剂性关节炎大鼠JAK2/STAT3信号通路

目的研究威灵仙总皂苷(TSC)对佐剂性关节炎(AA)大鼠的JAK2/STAT3信号通路的作用机制。方法雄性SD大鼠60只,随机分为6组,每组10只。除正常组外,采用Freund完全佐剂诱导AA大鼠模型。造模第12天,灌胃给予相应药物,1次/d,连续16 d,观察药物对AA大鼠体质量及足肿胀度的影响;取固定部位踝关节组织,HE染色观察病理学改变;实时荧光定量PCR法测定滑膜细胞中Janus激酶2(JAK2)、信号转导子和转录激活子3(STAT3)mRNA表达;Western blot法检测磷酸化JAK2及非磷酸化JAK2(p-JAK2、JAK2)、磷酸化STAT3及非磷酸化STAT3(p-STAT3、STAT3)的蛋白表达。结果 TSC可有效缓解AA大鼠体质量减轻及明显抑制AA大鼠足肿胀,明显改变炎细胞浸润,减少血管翳生成,减轻组织增生,有效抑制JAK2、STAT3 mRNA表达及降低p-JAK2/JAK2、p-STAT3/STAT3的相对表达。结论 TSC降低AA大鼠JAK2、STAT3 mRNA表达并抑制p-JAK2/JAK2、p-STAT3/STAT3相对表达,抑制JAK2/STAT3信号通路。     

IgA肾病肾小球毛细血管内血浆蛋白淤积的临床病理意义

观察ＩａＡ肾病肾小球毛细血管内血浆蛋白平均吸光度变化与微血栓形成率、ＩｇＡ沉积量、临床表现及病理类型之间的关系。方法用图像分析仪定量测定５５例ＩｇＡＮ肾穿标本ＭＡＰＧ和ＩｇＡ沉积物,比较高ＭＡＰＧ组与低ＭＡＰＧ组之间微血栓形成率、ＩｇＡ沉积量、临床表现及病理类型的差异。     

雷帕霉素通过PI3K/AKT通路改善碘克沙醇致糖尿病大鼠肾损伤

目的观察雷帕霉素对碘克沙醇所致糖尿病大鼠肾损伤的保护作用,探讨其作用机制,为临床药物防治造影剂肾损伤提供理论依据。方法糖尿病大鼠45只,随机分为造影剂组(CM组,n=15)、雷帕霉素组(Rapa组,n=15)与空白对照组(NS组,n=15),SD大鼠被腹腔注射链脲佐菌素(STZ)建立糖尿病模型,随后给予碘克沙醇制备CIAKI模型,Rapa组给予雷帕霉素进行治疗,分别观察各组大鼠血中肌酐(Scr)、血尿素氮(BUN)的浓度变化;HE染色观察肾脏病理学变化;TUNEL检测肾上皮细胞凋亡情况、Western blot检测Bcl-2、Bax、PI3K、AKT、p-AKT表达情况。结果 CM组大鼠肾功能损伤严重,小管上皮细胞破坏,肾小管空泡变性,细胞脱落至小管腔,肾小管上皮细胞凋亡。雷帕霉素干预后,Rapa组肾功能明显改善,肾小管上皮细胞损伤较轻,肾组织中Bax蛋白的表达较CM组明显降低,Bcl-2、PI3K、AKT、p-AKT表达明显升高,细胞凋亡数减少。结论雷帕霉素改善碘克沙醇导致的糖尿病大鼠肾功能损伤,与下调Bax表达、上调Bcl-2、PI3K、AKT、p-AKT表达相关。     

雷帕霉素介导丝裂原活化蛋白激酶信号通路对肝癌细胞的影响

目的： 探讨雷帕霉素介导丝裂原活化蛋白激酶（MAPK）信号通路对肝癌细胞Bcl-2、Bcl-xl 及Bax 蛋白表 达的影响。方法：将人肝癌细胞BEL-7402 细胞分为对照组和雷帕霉素处理组（20、50、100 ng/mL）,观察各组 BEL-7402 细胞的形态变化、细胞增殖抑制率、凋亡及蛋白（MAPK、Bcl-2、Bcl-xl 及Bax）表达。结果：随着 雷帕霉素浓度的升高,BEL-7402 细胞呈现崩解和坏死;对照组BEL-7402 细胞增殖抑制率最低,雷帕霉素干预组 BEL-7402 细胞增殖抑制率高于对照组,随着时间及雷帕霉素浓度的升高,细胞生长抑制率逐渐升高;雷帕霉素 处理组与对照组相比差异有统计学意义;随着浓度的升高,BEL-7402 细胞凋亡率升高;对照组BEL-7402 细胞中 MAPK阳性表达率高于雷帕霉素处理组,雷帕霉素处理组随着浓度升高,MAPK阳性表达率不断降低;对照组 BEL-7402 细胞中Bcl-2、Bcl-xl 蛋白升高,与雷帕霉素处理组比较存在显著差异,雷帕霉素处理组Bcl-2、Bcl-xl 蛋白水平随着雷帕霉素的浓度升高而降低,Bax 表达水平随着雷帕霉素的浓度升高而增加。结论：雷帕霉素能 够抑制肝癌细胞增殖,加快坏死及破裂,随着雷帕霉素的浓度升高,凋亡程度增大,其作用机制可能与抑制 MAPK、Bcl-2、Bcl-xl 表达,促进Bax 水平升高有关。     

雷帕霉素抑制人多囊肾囊肿衬里上皮细胞增殖及VEGF表达

目的 探讨雷帕霉素对常染色体显性多囊肾病囊肿衬里上皮细胞增殖及血管内皮细胞生长因子（VEGF）表达的抑制作用及其机制。方法MTT法检测WT9-12细胞增殖;流式细胞术检测细胞周期及凋亡;Western Blot检测周期相关蛋白(cyclinD、p21)、凋亡相关蛋白(Bcl2/Bax)及VEGF表达。结果 雷帕霉素可抑制WT9-12细胞的增殖,使细胞周期停滞在G0/G1期并促进细胞凋亡。雷帕霉素可下调WT9-12细胞cyclinD 、上调p21表达,下调Bcl2、上调Bax表达。原代培养的多囊肾囊肿衬里上皮细胞及WT9-12细胞的VEGFmRNA表达明显高于正常肾小管上皮细胞（P<0.05）。雷帕霉素可抑制WT9-12细胞VEGF的表达（P<0.05）。结论 雷帕霉素可通过抑制多囊肾囊肿衬里上皮细胞增殖、促进凋亡及抑制VEGF表达抑制血管形成,从而抑制多囊肾病进展。     

褪黑素对心力衰竭大鼠JAK1/STAT3信号通路的调节作用

目的 探讨褪黑素对心力衰竭(HF)大鼠JAK1/STAT3信号通路的影响。方法 将SD雄性大鼠通过左前降支冠状动脉结扎术建立HF模型并随机分为模型组和褪黑素低剂量组(5 mg/kg)、中剂量组(10 mg/kg)、高剂量组(20 mg/kg),将正常大鼠作为对照组,20只/组。对照组和模型组灌胃给药生理盐水,所有组别连续灌胃4周。采用无创尾动脉血压测量分析系统检测各组大鼠给药后的血压及心率;采用彩色多普勒超声诊断仪检测大鼠左心室舒张末期容积(left ventricular end diastolic volume,LVEDV)、左心室收缩末期容积(left ventricular end systolic volume, LVESV)、心室舒张末期室间隔厚度(IvSd)、左心室舒张末期直径(left ventricular end diastolic diameter, LVIDd)、左心室收缩末期直径(left ventricular end systolic diameter, LVID)、舒张早期二尖瓣血流速度/舒张晚期二尖瓣血流速度比值(E/A)、射血分数(ejection ...     

阿托伐他汀抑制NLRP3炎性小体缓解实验性IgA肾病大鼠肾损伤

目的 观察阿托伐他汀对IgA肾病大鼠的肾保护作用,探讨其介导NLRP3炎性小体的作用机制.方法 Sd大鼠随机分为对照组、模型组、阿托伐他汀低、高剂量组,每组10只;利用口服牛γ-球蛋白(BGG)联合尾静脉注射BGG方法建立IgA肾病大鼠模型;于造模完成后灌胃给予低、高剂量阿托伐他汀治疗6周;取血液分离血清检测肌酐(Scr)、尿素氮(BUN)、总蛋白(TP)、白蛋白丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)含量;检测大鼠尿液中红细胞数目及24 h尿蛋白(24 h TUP)水平;HE染色检测肾病理组织学变化;免疫荧光染色检测肾组织中IgA沉积情况;Western blot检测肾组织中IL-1、Caspase-l及NLRP3的表达.结果 阿托伐他汀显著降低血清中Scr、BUN及尿液中红细胞数目、24hTUP含量;改善IgA的过度沉积和病理损伤,同时阿托伐他汀抑制了IL-1、Caspase-1及NLRP3蛋白的表达(P＜0.05).结论 阿托伐他汀改善IgA肾病大鼠肾功能,缓解肾损伤,与抑制NLRP3-(Caspase-l)/IL-l信号通路相关.     

阻断CCR2信号对IgA肾病模型大鼠肾病理损伤及炎性因子表达的影响

目的 探讨阻断CCR2信号对IgA肾病（IgAnephropathy,IgAN）模型大鼠肾组织病理变化及炎性因子表达的影响。方法将40只Wistar大鼠随机分为正常对照组、IgAN模型组、CCR2拮抗剂组和奥美沙坦组。联合应用LPS、BSA和CCl4建立IgAN大鼠模型、CCR2拮抗剂组和奥美沙坦组,大鼠成模后分别予以10mg/kg的CCR2拮抗剂或Olmesartan灌胃2周,正常对照组和IgAN模型组予以等量的生理盐水。测定24h尿蛋白（NPr）、血尿素氮（BUN）、血肌酐（SCre）含量;采用免疫荧光、免疫组化、病理染色等方法检测IgAN模型组大鼠肾组织病理改变及炎性因子的表达。结果 与正常对照组相比,IgAN模型组大鼠肾小球系膜区IgA沉积和系膜细胞增生明显,24hNPr、SCre和BUN水平明显升高（P0.01）,CCR2拮抗剂能减少系膜区IgA沉积和系膜细胞增生,并能明显降低大鼠24hNPr、SCre和BUN水平（P0.01）。与正常对照组相比,IgAN模型组大鼠肾组织炎症相关因子MCP 1、IL 6、IL 17和TNF α表达明显增强,CCR2拮抗剂能明显抑制上述炎症因子的表达（P0.05）。结论 阻断CCR2信号可缓解IgAN模型大鼠的肾组织病理损伤,并降低肾组织炎症因子的表达,提示CCR2信号通路可能参与IgAN的发生、发展。     

成人IgA肾病中CD20的表达及意义

目的探讨CD20在不同肾功能状态的成人IgA肾病(IgA nephropathy,IgAN)患者肾组织中的表达及其与肾脏临床病理特征、预后的关系。方法将47例IgAN依据改良MDRD公式评估肾小球滤过率(estimated glomerular filtration rate,eGFR),将入选患者分为三组:A组(肾功能正常或轻度受损):eGFR60 ml/min;B组(肾功能中度受损):eGFR 30~60 ml/min;C组(肾功能重度受损,不含尿毒症):eGFR 10~30 ml/min。按Katafuchi评分标准进行肾活检病理评分,应用免疫组化法检测三组肾穿刺标本中CD20的表达,并对所有患者进行随访,复查相关临床指标。结果 CD20主要表达于IgAN患者肾组织皮质区肾小管管周间质B淋巴细胞胞膜。与eGFR60 ml/min组相比,eGFR 30~60 ml/min组、eGFR 10~30 ml/min组CD20表达量明显升高,差异有统计学意义(P值分别为0.002,0.001);肾组织中CD20表达量与eGFR呈负相关(r=-0.697,P0.001),与24 h尿蛋白定量、血清肌酐、肾小管间质积分呈正相关(r值分别为0.524、0.647、0.769;P均0.001)。结论肾组织中CD20的表达量与IgAN患者肾脏病理损伤指数、24 h尿蛋白定量、肾功能状态均密切相关,提示检测IgAN患者肾组织中CD20表达对了解肾脏病变程度、评估患者预后有一定临床价值。     

Anti-inflammatory effects of triptolide on IgA nephropathy in rats

Abstract

IgA nephropathy (IgAN) is the finding of immune deposits predominantly containing polymeric IgA in the glomerular mesangium on renal biopsy. Recently studies show that inflammation may involve in the progression of renal glomerulosclerosis and tubulointerstitial scarring in IgAN. This study was designed to evaluate the renoprotective effect of triptolide on IgAN rat model. IgAN was induced in Sprague–Dawley rats by oral and intravenous immunization with BSA for 12 weeks. Rats were treated with triptolide (200?μg/kg/d intragastrically) from 12 to 28 weeks. At Week 28, the rats was sacrificed, kidneys and blood samples were collected for further analysis. Our data shown that IgAN rat model showed marked deterioration of proteinuria together with higher levels of the urine protein:creatinine ratio compared to the normal control. Animals that underwent intermittent exposure to triptolide treatment exhibited significant improvements in the functional parameters without severe side effects. Rats developing IgAN had profound mesangial proliferation and mesangial expansion, intense and diffuse glomerular IgA deposition, while triptolide treatment significantly attenuated it. We also observed that treatment with triptolide significantly decreases serum levels of IL-1β and IL-18, and may exerted anti-inflammatory effects by down-regulating NLRP3 and TLR4 expression. Our study clearly demonstrated that triptolide prevents IgAN progression via an amelioration of inflammasome-mediated proinflammatory cytokine production, thus brought a light of hope for treatment of IgAN.     

Oligoclonally expanding gammadelta T lymphocytes induce IgA switching in IgA nephropathy

The aetiology of IgA nephropathy (IgAN) is closely related with abnormality of mucosal immunity. We investigated the roles of γδ T cells in the regulation of IgA production by B cells in IgAN patients. The proportion of γδ T cells in peripheral blood mononuclear cells (PBMNC) was higher in IgAN patients than in the controls and was found to be correlated with the proportion of surface IgA‐positive (sIgA+) B cells, which are precursors of IgA‐secreting plasma cells. After in vitro PWM stimulation, sIgA expression on B cells and IgA production were significantly enhanced in PBMNC obtained from IgAN patients, whereas the enhancements were abolished by removal of γδ T cells from the PBMNC. Purified γδ T cells from IgAN patients induced surface IgA expression on naïve sIgD+ B cells more effectively than did αβ T cells. Moreover, stimulated γδ T cells from IgAN patients produced a larger amount of TGF‐β1, which is one of the main cytokines that induces IgA class switching on B cells, as compared with αβ T cells and control γδ T cells. The expanded γδ T cells from IgAN patients exclusively expressed Vγ9, and the nucleotide sequences of junctional regions of Vγ9 showed very limited TCR diversities. It was therefore concluded that γδ T cells, which are expanded in response to specific antigens, enhance IgA class switching on B cells in IgAN patients.     

Urinary IgA in IgA nephropathy and Henoch-Schoenlein purpura

To determine the concentrations and molecular forms of urinary IgA in IgA nephropathy and Henoch-Schoenlein purpura, we studied 29 patients with these IgA-associated renal diseases (IgAN). Control groups comprised 10 patients with other diverse renal diseases and 11 healthy volunteers. Urinary IgA and IgG concentrations were higher in IgAN than in either control group and correlated positively with the serum creatinine concentration as well as the urinary protein excretion (P<0.01). However, IgA/IgG ratios did not differ among the three groups. Polymeric IgA (p-IgA) in the urine predominated only in normals; in IgAN and patients with other renal diseases, monomeric IgA (m-IgA) occurred almost exclusively. Serum IgA concentrations were generally normal in IgAN; four patients had concentrations greater than 500 mg/dl. Although the fraction of p-IgA in serum (median, 18%) was increased above normal (5–10%) in 13 of 16 (81%) subjects, neither the concentration of IgA or IgG nor the amount of p-IgA correlated with the serum creatinine concentration. These data suggest that the molecular form and concentration of urinary IgA are not discriminating for IgAN and are independent of these characteristics of serum IgA.     

Expression of prorenin receptor in renal biopsies from patients with IgA nephropathy

Prorenin receptor (PRR) has been implicated in the onset and progression of various renal diseases, though its possible association with immunoglobulin A (IgA) nephropathy remains unclear. In the present study, we tried to clarify expression and pathophysiological significance of PRR in IgA nephropathy. We immunohistochemically assessed PRR levels in renal biopsy specimens from 48 patients with IgA nephropathy and evaluated its relevance to the clinical and pathological features of the disease. PRR was detected mainly in renal tubular cells, which was confirmed at the subcellular level using immunoelectron microscopy. The PRR-positive area (%PRR area) correlated with daily urinary protein, which is known to reflect disease severity (r=0.286, P=0.049). PRR levels were weaker in tubular cells bordering areas of severe interstitial fibrosis, where α-smooth muscle actin-positive myofibroblasts were present. We also used immunohistochemical detection of microtubule-associated protein-1 light chain 3 (LC3) and electron microscopy to assess autophagy, a cytoprotective mechanism downstream of PRR. We noted an apparent coincidence between autophagy activation in tubular cells and PRR expression in the same cells. Taken together, our findings suggest that renal expression of PRR in IgA nephropathy may be a compensatory response slowing disease progression by preventing tubular cell death and subsequent fibrosis through activation of cytoprotective autophagic machinery. Further studies using different type of kidney diseases could draw conclusion if the present finding is a generalized observation beyond IgA nephropathy.     

IgA polyspecific autoantibodies in IgA nephropathy.

The specificity of circulating and kidney-bound IgA during IgA nephropathy is still a matter of discussion. In the present study, high levels of IgA antibodies directed against a panel of self and non-self antigens were found in the serum from patients with IgA nephropathy and were eluted from four out of the seven kidney biopsies studied. After immunoadsorption of pooled selected serum samples on TNP and actin-coated columns, polyspecific IgA antibodies were eluted. This supports the hypothesis that IgA-bearing B cells clones most probably producing polyspecific antibodies are a major feature of human IgA nephropathy. These findings also suggest that it may be hazardous to draw conclusions from the finding of apparently monospecific IgA antibodies in this condition.     

Protective and susceptible HLA polymorphisms in IgA nephropathy patients with end-stage renal failure

Idiopathic immunoglobulin A (IgA) nephropathy is characterised by an extreme variability in clinical course, leading to end-stage renal failure in 15-20% of adults. This subgroup of patients with IgA nephropathy is usually included in the waiting lists of organ exchange organisations. The frequency of HLA-A,B,DR antigens of this subset of IgA nephropathy patients was calculated and compared to controls. The antigens HLA-B35 and DR5 were significantly increased in the patients with relative risk values of 1.385 and 1.487, respectively. The antigens HLA-B7, B8, DR2, and DR3 were found in a significantly lower frequency in the patients as compared to the controls. The relative risk (RR) values ranged between 0.695 and 0.727. Consequently, the haplotypes HLA-A1, B8, DR3, HLA-A3, B7, DR2, HLA-A2, B7, DR2 together with HLA-A1, B15, DR4, HLA-A9, B12, DR7, and HLA-A10, B18, DR2 were found to be protective with RR values ranging from 0.309 to 0.587. The only susceptible haplotype observed was HLA-A2-B5, DR5 (RR=2.990).     

白花丹素调控ROS抑制NLRP3炎症小体减轻IgA肾损伤机制的实验研究

目的 研究白花丹素对IgA肾病的作用和机制。 方法 按Ying等改良的BSA+LPS+CCl4方法复制实验IgA肾病模型；然后，给药组大鼠每组分别腹腔注射10 mg/kg、20 mg/kg和50 mg/kg白花丹素，1次/d，对照组和模型组腹腔注射等量的生理盐水1次/d；8周后，全自动化学分析仪检测24 h尿蛋白、血肌酐、血尿素，流式检测ROS含量，试剂盒检测SOD活性和MDA含量，HE染色观察病理损伤，Elisa检测TNF-α、IL-18、IL-1β，蛋白印迹法检测NLRP3、ASC、caspase-1 p20、P13K、AKT和NF-kB蛋白表达。 结果 与模型组相比，给药组大鼠的尿蛋白、血清肌酸酐和尿素氮含量显著减少，ROS水平显著降低，SOD含量显著增多，MDA含量显著减少，MDA、IL-1β、IL-18和TNF-α的含量显著减少，NLRP3、ASC、caspase-1 p20、P13K、AKT和NF-kB的蛋白表达显著下调，并且随着给药量的增加效果越显著。 结论 白花丹素通过降低尿蛋白、血肌酐和血尿素含量，减轻病理损伤，抑制氧化应激、炎症反应和NLRP3/P13K/AKT/NF-kB通路激活来减轻IgA肾病。