雷帕霉素纳米粒滴眼液联合环孢素A缓释膜治疗兔高危角膜移植术后免疫排斥反应的研究

Objective To evaluate the combined effect of topical rapamycin (RAPA) eye drop in nanometer vector and poly (lactic acid) (PLA) wafers of cyclospoirne A(CsA) in the prevention of acute allograft rejection after rabbit corneal transplantation. Methods It was an experimental study. RAPA was incorporated into the nanometer particles and CsA was incorporated into PLA wafers. A was syngeneic control whose both donor and recipient are New Zealand rabbit. Gray donor corneas were implanted into the 102 recipients of New Zealand albino rabbits with corneal neovascularization who were randomly divided into B, C, D, E, F, G 6 groups to receive the different types of therapy: B was no therapy control; C was eye drop of nanometer vector but no RAPA twice a day,28 days; D was PLA wafers in the anterior chamber of rabbit eyes but no drugs; E was 0.5% RAPA eye drop of nanometer vector twice a day,28 days; F was PLA wafers of CsA in the anterior chamber of rabbit eyes; G was PLA wafers of CsA in the anterior chamber of rabbit eyes and 0.5% RAPA eye drop of nanometer vector eye drop twice a day for 28 days together. Postoperative evaluation included slit-lamp biomicroscopy, histopathology and immunohistoiogy, Cytokines related with neovascularization and immunosuppression in the corneal tissue by RT-PCR. The graft survival was assessed by One-Way ANOVA and q test. Results Corneal allograft survival time: A (100.00±0.00), B ( 8.44±1.24), C (8.89±2.57), D (8.56±2.30), E (43.11±5.58), F (43.67±9.54), G (72.00±15.34)d. Group G led to a statistically significant prolongation of transplant survival and was superior than group E and F which was a statistical prolongation compared with group B, C and D (qGE=11.42, qGF=11.24,qEB=13.64, qEC=13.38, q<=13.46, qFB=13. 82, qFC=13.56, qFD=13.64; P<0.01). Immunohistopathologically, the grafts were subjected to an immune response contained a dense infiltrate of neutorphils,CD4+ and CD8+ T lymphocytes in the group B ,C and D. This cellular infiltrate was a significant reduction in group E,F,G. RT-PCR showed that the gene expression of IL-2 was inhibited earlier (3 days) in group F, G and VEGF gene expression being suppressed later (14 days) in group E, G. Conclusions Combined therapy with topical application of RAPA eye drop of nanometer vector and CsA PLA wafers can significantly prolong the survival of allograft at high-risk. Moreover, topical combined treatment of them is more effective, lower dosage, less side-effects and cheaper than the treatment with topical individual immunosuppressive drug.     

碘/钠摄入量对小鼠血脂代谢影响的实验研究

Objective The present study has been designed to investigate the impact of dietary iodine/sodium intake on blood lipid metabolism in mice. Methods According to body weight and gender, two hundred and sixty Balb/c mice were randomly divided into 2 groups including normal sodium group(Na) and low sodium group(LNa), with 130 animals per group. Each group were then randomly further divided into 5 sub-groups according to the amount of iodine intake: ① severe iodine deficiency(SID); ② mild iodine deficiency(MID); (③normal iodine (NI); ④ 10-fold high iodine ( 10HI ); (⑤ 50-fold high iodine (50HI), 10 groups in total, 26 per group.Eight months later, the body weight and the levels of urinary iodine, thyroid hormones and total cholesterol (TC),Results In Na group, the levels of TG and TC in male mice of SID group[ (1.64 ± 0.35), (3.88 ± 0.35 )mmol/L]and MID group[ ( 1.67 ± 0.31 ), (3.41 ± 0.66)mmol/L] were significantly higher than that of NI group[ ( 1.49 ± 0.42), (3.25 ± 0.47)mmol/L] and the levels of TG in female mice of SID group[(1.52 ± 0.22)mmol/L] were significantly higher than that of NI group[ (1.23 ± 0.22)mmol/L]. In addition, the levels of TG in male mice of 10HI and 50HI groups [ ( 1.16 ± 0.23 ), ( 1.21 ± 0.27 ) mmol/L ] were significantly lower than that of NI group [ ( 1.49 ± 0.42)mmol/L, all P ＜ 0.05], the levels of TC in female mice of 10HI and 50HI groups[(2.37 ± 0.49), (2.48 ± 0.37)mmol/L] were significantly lower than that of NI group[ (2.84 ± 0.37) mmol/L, all P ＜ 0.05 ]. In LNa group,the levels of TG and TC in male mice of SID group[ (1.39 ± 0.40), (3.33 ± 0.46 )mmol/L] were significantly lower than that of NI group [(1.30 ± 0.28), (3.00 ± 0.53) mmol/L, all P ＜ 0.05], the levels of TG, TC and LDL in female mice of SID group[ (1.48 ± 0.26), (2.76 ± 0.43), (0.62 ± 0.22)mmol/L], the levels of LDL in female mice of MID group[ (0.60 ± 0.17 )mmol/L] were significantly lower than that of NI group[(l.22 ± 0.36), (2.51 ± 0.38),(0.48 ± 0.08), (0.48 ± 0.08)mmol/L, all P ＜ 0.05], the levels of TG in male mice of 10HI and 50HI group [ (1.12 ± 0.22), (0.90 ± 0.11 )mmol/L] were significantly lower than that of NI group (all P ＜ 0.05 ), the levels of TC in female mice of 10HI and 50HI groups[ (2.35 ± 0.34), (2.37 ± 0.37)mmol/L], the levels of LDL in female mice of 50HI group[(0.65 ± 0.18)mmol/L], were significantly lower than that of NI group(all P ＜ 0.05). In Na group, the levels of thyroid hormones were distinctively decreased in SID group[TT4(0.00 ± 0.00)nmol/L, FT4 (0.93 ± 0.42)pmol/L, TT3(0.49 ± 0.07)nmol/L, FT3(2.86 ± 0.37)pmol/L] and MID group [TT4 (17.15 ± 15.26)nmol/L, FT4( 18.46 ± 4.31 )pmol/L, TT3(0.67 ± 0. 10)nmol/L, FT3(3.18 ± 0.24)pmol/L] compared with that of the NI group [TT4 (37.15 ± 15.26)nmol/L, FT4(28.46 ± 4.31)pmol/L, TT3(0.85 ± 0.10)pmol/L, FT3(3.87 ± 0.24)pmol/L, all P ＜ 0.05 ]. In LNa group, the levels of thyroid hormones were distinctively decreased in SID group [TT4 (0.00 ± 0.00) nmol/L,FT4(1.03 ± 0.78)pmol/L, TT3(0.51 ± 0.05)nmol/L, FT3(3.01 ± 0.17)pmol/L] and MID group[TT4(19.76 ± 12.22)nmol/L, FT4(21.46 ± 5.37)pmol/L, TT3(0.71 ± 0.21)nmol/L, FT3(3.56 ± 0.23)pmol/L] compared with that of the NI group[TT4(36.23 ± 14.72)nmol/L, FT4(30.96 ± 6.33)pmol/L, TT3(0.89 ± 0.20)nmol/L, FT3(4.05 ± 0.24)pmol/L, all P ＜ 0.05]. Conclusions Dietary iodine intake plays an important role in the blood lipid metabolism. Iodine deficiency could increase while iodine excess could decrease the levels of serum TG, TC or LDL in mice. Monitoring the amount of iodine intake during sodium restriction should have an important role in effective prevention and treatment of cardiovascular disease.     

碘对诱发实验性自身免疫性甲状腺炎形成的影响

目的建立诱发型实验性自身免疫性甲状腺炎(EAT)动物模型,研究碘对诱发EAT形成的影响,进一步探讨其可能的作用机制。方法:采用雌性Lewis大鼠诱发EAT并饲以不同碘含量的饮食,观察各组大鼠甲状腺病理改变、血清中Tg自身抗体水平、血清中甲状腺激素水平、脾细胞分泌TNF水平和甲状腺FasmRNA表达水平。结果1.诱发EAT的HI组大鼠炎症范围较广,为较重度炎症。NI组显示为轻度炎症,且为恢复期。LI组显示为滤泡内少量淋巴细胞浸润。2.诱发EAT的HI、NI和LI组大鼠血清中均出现较高水平的Tg自身抗体,3组间无显著性差异。3.诱发EAT的HI组大鼠脾细胞分泌TNE水平高于NI和LI组(P<0.05)。4.诱发EAT的HI、NI和LI组大鼠甲状腺FasmRNA的表达水平均较各自未诱发EAT的大鼠明显升高,其中HI组高于同组的NI和LI组,P<0.01。结论1.高碘饮食可使已诱发EAT的Lewis大鼠甲状腺组织炎症加重且炎症反应持续时间较长,考虑与脾细胞分泌的TNF水平和甲状腺FasmRNA的表达水平较高有关。2.未发现甲状腺组织炎症的程度与自身抗体水平相关。     

低碘后补充不同碘水平饮食对大鼠肿瘤坏死因子和干扰素mRNA表达的影响

目的大鼠低碘后,补充不同碘水平饮食,观察其甲状腺组织形态、甲状腺激素水平和脾脏中肿瘤坏死因子(TNF-α)和干扰素(IFN-γ)的mRNA表达,探讨低碘后补碘对大鼠的影响。方法选用Wistar大鼠,低碘饲料加去离子水饲养3个月为低碘组(LI组),低碘后补充适碘饮食为LI-1×KI组,补充高碘饮食为LI-50×KI组,均补碘3个月,另设适碘对照组(NI组)。检测4组大鼠血清中甲状腺激素水平、甲状腺组织形态、脾脏中TNF-α和IFN-γmRNA的表达。结果LI组大鼠血清中T3和T4水平下降,甲状腺滤泡内胶质稀少,甲状腺上皮细胞呈增生、肥大等典型的低碘表现,脾脏中TNF-α和IFN-γ的mRNA表达也明显低于NI组。低碘后补充适碘饮食的LI-1×KI组,不仅使血清中T3和T4水平和甲状腺上皮细胞的形态逐渐恢复正常,TNF-α和IFN-γ的mRNA表达也明显提高。而低碘后补充高碘饮食的LI-50×KI组,甲状腺激素水平和TNF-α和IFN-γ的mRNA表达也均明显上升,但甲状腺组织形态虽有所好转,与LI-1×KI组比较尚有明显差异,且甲状腺滤泡有胶质潴留。结论低碘后补充适当水平的碘,其甲状腺激素水平、TNF-α和IFN-γ的mRNA表达及甲状腺组织形态的改变可逐渐恢复正常,呈可逆性。而低碘后补充高碘饮食,其甲状腺组织形态学的改变尚难恢复为正常。     

雌激素与自身免疫性疾病

自身免疫性疾病的发病存在明显的性别差异，女性的发病率常高于男性，文章就雌激素在自身免疫性疾病发生机制中的作用，特别是近年来雌激素对免疫细胞信号，细胞分化，下丘脑－垂体－肾上腺（HPA）轴的激活，自身免疫性疾病的发生和缓解等方面的研究作一介绍，进一步探讨雌激素调节异常对自身免疫性疾病发生的影响。     

雷帕霉素纳米粒滴眼液联合环孢素A缓释膜治疗兔高危角膜移植术后免疫排斥反应的研究

目的 探讨局部联合应用雷帕霉素(RAPA)纳米粒滴眼液与环孢素A(CsA)缓释膜治疗兔高危角膜移植术后免疫排斥反应的效果和协同机制.方法 实验研究.(1)制备0.5%聚乳酸/胆固醇改性壳聚糖RAPA纳米粒滴眼液和聚乳酸/聚乙二醇CsA缓释膜.(2)设A组为同源对照组(17只兔),供受体均为新西兰白兔.建立102只(102只眼)新西兰白兔角膜新生血管模型,采用随机数字表法将其分为B、C、D、E、F、G6组,每组17只,供体为青紫兰兔,各组行穿透性角膜移植术.A组同源对照组;B组未治疗组;C组空白纳米粒滴眼液滴眼,每天2次共28 d;D组术中前房植入空白缓释膜;E组0.5%RAPA纳米粒滴眼液滴眼,每天2次共28 d;F组术中前房植入CsA缓释膜;G组术中前房植入CsA缓释膜并0.5%RAPA纳米粒滴眼液滴眼,每天2次共28 d.(3)术后观察100 d,隔天用显微镜观察角膜植片情况,记录免疫排斥反应发生时间和程度.(4)术后3、14、28及35 d用CD4和CD8单克隆抗体行免疫组织化学检查;用逆转录聚合酶链反应(RT-PCR)检测植片白细胞介素2(IL-2)和血管内皮生长因子(VEGF)表达.应用单因素方差分析和q检验,比较各组角膜植片平均存活时间.结果 各组兔眼角膜植片存活时间分别为A组(100.00±0.00)d、B组(8.44±1.24)d、C组(8.89±2.57)d、D组(8.56±2.30)d、E组(43.11±5.58)d、F组(43.67±9.54)d、G组(72.00±15.34)d.G组较E、F组,E、F组较B、C、D组,能显著延长角膜植片存活(qGE=11.42,qGF=11.24,qEB=13.64,qEC=13.38,qED=13.46,qFB=13.82,qFC=13.56,qFD=13.64;均P<0.01).免疫组织化学检查显示,B、C、D组术后14 d左右角膜植片中CD4+和CD8+T淋巴细胞大量聚集,呈进行性加重;E、F组术后35 d左右角膜植床与植片中CIM+和CD8+T淋巴细胞数开始出现;G组术后60 d以后,植片中CD4+和CD8+T淋巴细胞浸润.角膜植片RT-PCR检查显示,A组IL-2和VEGF始终未表达;F、G组IL-2表达在3 d开始被抑制;E、G组VEGF表达在14 d开始被抑制.结论 局部联合应用药物缓释剂治疗高危角膜移植术后免疫排斥反应较单独用药效果好,联合用药能减少单独用药的剂量和毒副作用.     

雷帕霉素纳米粒滴眼液联合环孢素A缓释膜治疗兔高危角膜移植术后免疫排斥反应的研究

Objective To evaluate the combined effect of topical rapamycin (RAPA) eye drop in nanometer vector and poly (lactic acid) (PLA) wafers of cyclospoirne A(CsA) in the prevention of acute allograft rejection after rabbit corneal transplantation. Methods It was an experimental study. RAPA was incorporated into the nanometer particles and CsA was incorporated into PLA wafers. A was syngeneic control whose both donor and recipient are New Zealand rabbit. Gray donor corneas were implanted into the 102 recipients of New Zealand albino rabbits with corneal neovascularization who were randomly divided into B, C, D, E, F, G 6 groups to receive the different types of therapy: B was no therapy control; C was eye drop of nanometer vector but no RAPA twice a day,28 days; D was PLA wafers in the anterior chamber of rabbit eyes but no drugs; E was 0.5% RAPA eye drop of nanometer vector twice a day,28 days; F was PLA wafers of CsA in the anterior chamber of rabbit eyes; G was PLA wafers of CsA in the anterior chamber of rabbit eyes and 0.5% RAPA eye drop of nanometer vector eye drop twice a day for 28 days together. Postoperative evaluation included slit-lamp biomicroscopy, histopathology and immunohistoiogy, Cytokines related with neovascularization and immunosuppression in the corneal tissue by RT-PCR. The graft survival was assessed by One-Way ANOVA and q test. Results Corneal allograft survival time: A (100.00±0.00), B ( 8.44±1.24), C (8.89±2.57), D (8.56±2.30), E (43.11±5.58), F (43.67±9.54), G (72.00±15.34)d. Group G led to a statistically significant prolongation of transplant survival and was superior than group E and F which was a statistical prolongation compared with group B, C and D (qGE=11.42, qGF=11.24,qEB=13.64, qEC=13.38, q<=13.46, qFB=13. 82, qFC=13.56, qFD=13.64; P<0.01). Immunohistopathologically, the grafts were subjected to an immune response contained a dense infiltrate of neutorphils,CD4+ and CD8+ T lymphocytes in the group B ,C and D. This cellular infiltrate was a significant reduction in group E,F,G. RT-PCR showed that the gene expression of IL-2 was inhibited earlier (3 days) in group F, G and VEGF gene expression being suppressed later (14 days) in group E, G. Conclusions Combined therapy with topical application of RAPA eye drop of nanometer vector and CsA PLA wafers can significantly prolong the survival of allograft at high-risk. Moreover, topical combined treatment of them is more effective, lower dosage, less side-effects and cheaper than the treatment with topical individual immunosuppressive drug.     

抗甲状腺刺激性免疫球蛋白单链抗体基因的构建及序列测定

 目的构建抗甲状腺刺激性免疫球蛋白(Thyroid stimulating immunoglobulin,TSI)单链抗体基因,获得基因序列.方法利用噬菌体表面呈现技术,结合间接ELISA方法和竞争抑制实验,筛选出抗TSI单链噬菌体抗体,进一步获得抗TSI单链抗体,对抗TSI单链抗体基因进行测序.结果抗TSI单链抗体基因长759bp,其中VL长342bp,VH长372bp,具有开放性阅读框架,为功能性基因.结论抗TSI单链抗体基因构建成功,噬菌体表面呈现技术为获得功能性单链抗体基因的可靠方法.     

产后甲状腺炎发病机制的研究进展

产后甲状腺炎是发生于产后一年内的一种暂时性或永久性甲状腺功能紊乱的综合征,属于自身免疫性疾病.甲状腺超氧化物酶抗体与产后甲状腺炎的发生密切相关,目前认为,妊娠相关的免疫抑制在产后消失,出现暂时性免疫反弹是引起产后甲状腺炎的主要因素.此外,神经-内分泌-免疫网络调节、环境因素及遗传因素等的相互作用,决定了产后甲状腺炎的发生、发展.     

持久稳定分泌抗人甲状腺球蛋白单克隆抗体的杂交瘤细胞株的建立

杂交瘤技术创立以来,在生物、医学各领域已广泛应用,各地陆续报导建立了分泌抗白细胞抗原、肿瘤抗原和微生物抗原的单克隆抗体(McAb)杂交瘤细胞株。但在内分泌免疫病方面,有关报导还不多。内分泌免疫病是免疫介导性疾病中尤为复杂的一组疾病,单克隆抗体技术为诊断和深入细致研究其发病机制提供了有力工具。我们选择了