1,25-二羟维生素D3处理的树突细胞在变应性气道炎症中的免疫调节作用

目的 探讨1,25-二羟维生素D,[1,25(OH)2D3]处理的树突细胞(DC)在变应性气道炎症中的免疫调节作用及其机制.方法 小鼠骨髓来源DC分2组,分别用1,25(OH):D,和磷酸盐缓冲液(PBS)处理,RT-PCR和蛋白质印迹法检测2组Dc Notch配体Mrna和蛋白表达.将2组DC和Notch配体中和抗体封闭的1,25(OH)2D3处理DC分别与小鼠脾脏来源CIM+T细胞共培养,流式细胞仪检测CD4+T细胞中CD4+CD25+Foxp3+T细胞百分比.将卵清蛋白(OVA)致敏小鼠分2组,每组5只,分别过继转移1,25(OH)2D3组DC[1,25(OH)2D3-DC组]和PBS组DC(PBS-DC组),以OVA激发气道炎症后行肺脏病理、支气管肺泡灌洗液(BALF)中自细胞介素4(IL-4)、IL-5、IL-13和干扰素γ(IFN-γ)水平以及脾脏CD4+T细胞中CD4+CD25+Foxp3+T细胞百分比检查.结果 1,25(OH)2D3组DC Notch配体Jaggedl和Jagged2 Mrna表达(0.376±0.029、0.564±0.018)和蛋白表达(0.786±0.034、0.632±0.026)均明显高于PBS组(分别为0.146±0.032、0.267±0.012和0.124±0.025、0.098±0.012,均P＜0.01).与CIM+T细胞共培养后,1,25(OH)2D3组CD4+T细胞中CD4+C1)25+Foxp3+T细胞百分比(22.49%±0.56%)明显高于PBS组(6.67%±0.60%,P＜0.01);经Jagged2中和抗体封闭组CIM4CD25+Foxp3+T细胞百分比(6.56%±1.89%)明显低于未封闭组(20.37%±1.64%,P＜0.01),1,25(OH)2D3-DC组小鼠气道炎症明显轻于PBS-DC组;BALF中IL-4、IL-5、IL-13和IFN-γ水平(pg/ml)分别为33±5、134±23、91±11和未检测到(＜12.5),均明显低于PBS-DC组(分别为55±7、332±49、152±19、23±6,均P＜0.01);脾脏CD4+T细胞中CIM+CD25+Foxp3+T细胞百分比(14.69%±1.14%)明显高于PBS-DC组(2.38%±0.14%,P＜0.01).结论 1,25(OH)2D3处理DC对变应性气道炎症有抑制作用,这种作用可能与1,25(OH)2D3处理DC通过Jaggect2介导的Notch信号途径诱导CD4+CD25+Foxp3+T细胞生成有关.     

Inactivation of mammalian target of rapamycin (mTOR) by rapamycin in a murine model of lipopolysaccharide-induced acute lung injury

Background  The mammalian target of rapamycin (mTOR) pathway, a key cellular signaling pathway associated with various cellular functions, has distinct roles in the inflammatory process. In this study, the mTOR inhibitor rapamycin (Rapa) was used to test whether inhibition of mTOR activation attenuates lipopolysaccharide (LPS)-induced acute lung injury (ALI) in a murine model.

Methods  Mice pretreated with Rapa or vehicle were given LPS intratracheally. Local cell numbers and inflammatory cytokines present in the bronchoalveolar lavage fluid (BAL), wet-to-dry weight ratio, histopathology of the lungs, and survival were evaluated.

Results  The phosphorylation of S6, a major downstream target of mTOR, had a 3-fold increase in lung tissue after LPS stimulation, but the increase was blocked by Rapa. Rapa reduced the levels of TNF-α (LPS vs. LPS + Rapa, (1672.74±193.73) vs. (539.17±140.48) pg/ml, respectively; P <0.01) and IL-6 (LPS vs. LPS + Rapa: (7790.88±1170.54) vs. (1968.57±474.62) pg/ml, respectively; P <0.01) in the BAL fluid. However, Rapa had limited effects on the overall severity of ALI, as determined by the wet-to-dry weight ratio of the lungs, number of neutrophils in the BAL fluid, and changes in histopathology. In addition, Rapa failed to reduce mortality in the LPS-induced ALI model.

Conclusions  We confirmed that mTOR was activated during LPS-induced ALI and strongly inhibited by Rapa. Although Rapa reduced the levels of the mediators of inflammation, the overall severity and survival of the ALI murine model were unchanged.

     

利巴韦林对呼吸道合胞病毒感染哮喘加重小鼠胸腺基质淋巴细胞生成素分泌的影响

目的 明确利巴韦林(RIB)在呼吸道合胞病毒(RSV)感染哮喘加重治疗中的作用.方法 (1)细胞试验:人支气管上皮细胞16HBE随机分为RSV组、RSV/RIB组、RIB组和对照组,每组8瓶.感染细胞用RSV病毒液的感染复数(MOI)为2;RIB浓度为50 μg/ml.蛋白质印迹法检测各组细胞胸腺基质淋巴细胞生成素(TSLP)蛋白表达.(2)动物实验:雌性BALB/c小鼠随机分为卵清蛋白(OVA)组、OVA/RSV组、OVA/RSV/RIB组和对照组,每组8只.应用OVA腹腔注射致敏、OVA雾化吸入结合RSV病毒液滴鼻激发哮喘,RIB 10 mg/kg肌内注射.动物肺功能分析系统检测各组小鼠气道反应性;酶联免疫吸附试验检测小鼠血清白细胞介素4(IL-4)、IL-5、IL-13、干扰素γ(IFN-γ)和支气管肺泡灌洗液(BALF)TSLP浓度;取肺组织观察炎症反应和气道上皮细胞TSLP表达水平.结果 细胞试验显示RSV、RSV/RIB、RIB和对照组TSLP蛋白表达量分别为1.97±0.22、1.16±0.19、0.99±0.17和0.89±0.08,RSV/RIB组明显低于RSV组(P＜0.01).动物实验显示OVA/RSV/RIB组小鼠气道反应性明显低于OVA/RSV组(P＜0.01);血清IL-4、IL-5、IFN-γ和BALF中TSLP浓度分别为(109.7±41.9)、(220.8±30.9)、(13.0±3.4)和(1945±82)pg/ml,均明显低于OVA/RSV组[分别为(274.2±103.7)、(293.3±46.1)、(30.1±5.7)、(2127±46)pg/ml,均P＜0.01];气道炎症细胞浸润和气道上皮细胞TSLP表达均明显少于OVA/RSV组.结论 RIB可以抑制RSV感染引起的TSLP分泌增加和哮喘加重.     

Levels and dynamic changes of serum fibroblast growth factor 23 in hypophosphatemic rickets/osteomalacia

Background Hypophosphatemic rickets/osteomalacia is a group of diseases characterised by defective mineralization of bone due to hypophosphatemia and low 1,25-dihydroxy vitamin D. To explore the role of fibroblast growth factor 23 （FGF-23） in the regulation of phosphate homeostasis, we measured the circulating concentrations of this growth factor in healthy individuals and in patients with hypophosphatemic rickets/osteomalacia. Methods Nineteen patients with hypophosphatemic rickets/osteomalacia were included in hypophosphatemic group （HP, 12 female and 7 male, mean age was 30 years）, and 19 healthy age-matched individuals served as the control group. Full length FGF-23 fragments were measured by two-site enzyme-linked immunosorbent assay.Results Mean FGF-23 concentrations were significantly higher in the HP group （（87.4±43.6） pg/ml） compared with the control group （（19.2±6.16） pg/ml; P 〈0.001）. In 1 patient with tumour-induced osteomalacia, serum FGF-23 concentrations were 84.1 pg/ml; these concentrations were normalized 2 hours after a hemangiopericytoma resection （7.8 pg/ml）. Subsequently, serum 1,25（OH）2 vitamin D3 concentrations significantly increased from 21.3 pg/ml to 89.3 pg/ml, and serum phosphorus levels were normalized. Conclusions Serum FGF-23 concentrations were markedly elevated in patients with hypophosphatemic rickets. FGF-23 plays an important role in the pathogenesis of hypophosphatemic rickets/osteomalacia.     

Relationship between endothelial cell protein C receptor gene 6936A/G polymorphisms and deep venous thrombosis

Background  Deep venous thrombosis (DVT) can result in pulmonary embolism, a fatal complication that is due to the dislodgement and movement of a blood clot (thrombus) from a limb into the lungs. Genetic risk factors related to DVT development include mutations in coagulation proteins, especially the endothelial protein C receptor (EPCR), a component of the anticoagulation protein C (PC) pathway. The objective of the present study was to analyze the relationship between the 6936A/G polymorphism in the EPCR gene and the occurrence of DVT. 

Methods  This study involved 65 patients with DVT and 71 age- and gender-matched healthy controls. Peripheral blood samples were collected from all subjects. Plasma levels of soluble EPCR (sEPCR) were measured by enzyme-linked immunosorbent assay. Genomic DNA was extracted and EPCR gene product was amplified by a standard PCR reaction. Gene product bands were sequenced to identify EPCR gene polymorphisms.

Results  In the control group, the level of sEPCR in subjects with 6936AG genotype was significantly higher than that in subjects with 6936AA genotype ((0.97±0.32) pg/ml vs. (0.61±0.24) pg/ml, P <0.01). Similarly in the DVT group, the level of sEPCR in subjects with the 6936AG were greater than that in subjects with the 6936AA genotype ((0.87±0.21) pg/ml vs. (0.50±0.18) pg/ml, P <0.01). The sEPCR level in DVT patients was significantly higher than that in healthy controls ((0.68±0.32) pg/ml vs. (0.54±0.22) pg/ml, P <0.05). The 6936AG genotype frequency in DVT patients was significantly higher than that in healthy controls (P <0.05). In contrast, the 6936AA genotype frequency in DVT patients was lower than that in healthy controls (P <0.05). Subjects carrying 6936AG had an increased risk of thrombosis (OR=2.75, 95% CI: 1.04–7.30, P <0.05).

Conclusions  EPCR gene 6936A/G polymorphism is associated with increased plasma levels of sEPCR. Subjects carrying 6936AG likely have an increased risk of thrombosis.

     

色氨酸代谢物调控Th17/Treg分化在小鼠哮喘变应原特异性免疫治疗中的作用及机制研究

目的 对色氨酸代谢物调控Th17/Treg分化在小鼠哮喘变应原特异性免疫治疗中的作用及机制进行研究.方法 将30只BALB/c鼠随机法分为5组:对照组、哮喘组、OVA-SIT组(OVA:鸡卵白蛋白;SIT:变应原特异性免疫治疗)、OVA-SIT+ 1-MT组(1-MT:IDO抑制剂,1-甲基色氨酸),OVA-SIT+1-MT+ KYN组(KYN:色氨酸代谢产物,犬尿氨酸).哮喘组:第0、7d予OVA致敏,第6周每天予1％OVA雾化激发,50 d予10％ OVA加强激发;OVA-SIT组:第4周每天予大剂量OVA皮下注射,余同哮喘组;OVA-SIT+ 1-MT组:第4周每天在腹腔内注入1-MT,1h后予大剂量OVA皮下注射h,余同哮喘组;OVA-SIT+ 1-MT+ KYN组:第3周每天加入1-MT,第4周每天加入KYN,末次加入KYN后1h予大剂量OVA行免疫治疗,余同哮喘组.末次激发6h内检测气道高反应性;对支气管肺泡灌洗液(BALF)进行细胞计数分析;ELISA检测血清IgE及BALF中IL-5、IL-10、IL-17;流式细胞技术检测脾脏CD4+RORγt+T及CD4+Foxp3+T细胞分化情况.结果 OVA-SIT+1-MT+KYN组较OVA-SIT+1-MT组酸性粒细胞浸润减少,炎症反应明显减轻.前者BALF中Il-5为74.8 ～ 86.8(83.48±6.02) pg/mL,IL-17为33.8～46.5(38.72 ±4.61) pg/mL,CD4+ RORγt+T细胞为2.45 ～2.82(2.60±0.14)％,CD4+ Foxp3+T细胞为7.83 ～9.09(8.36 ±0.53)％;后者BALF中Il-5为240.3～285.1 (259.65±16.27) pg/mL,IL-17为55.2 ～ 65.8(59.97±3.76) pg/mL,CD4+ RORγt+T细胞为4.31 ～5.34(4.94±0.38)％,CD4+Foxp3+T细胞为5.93～6.59(6.33 ±0.28)％,因此OVA-SIT+ 1-MT+ KYN组中Il-5、IL-17细胞因子以及脾脏中CD4+ RORγt+T细胞均明显低于OVA-SIT+ 1-MT组,差异均有统计学意义(P＜0.01),而前组脾脏中CD4+ Foxp3+T细胞水平明显高于后者,差异有统计学意义(P＜0.01).结论 色氨酸代谢产物有助于特异性免疫治疗减轻气道炎症作用,其机制与通过调控Th17及Treg分化有关.     

Interleukin-18 and -12 synergistically enhance cytotoxic functions of tumor-infiltrating lymphocytes

Background  The role of tumor-infiltrating lymphocytes (TILs) in the immunopathogenesis of individual cancer is not clear and is a challenge for anti-tumor immunotherapy. This study aimed to investigate the effects of interleukin (IL)-18 and -12 on cytotoxic functions of TILs.

Methods  TILs from postoperative gastric cancer patients were costimulated with IL-18 and IL-12. SGC-7901 tumor cells were pre-incubated with TILs and subcutaneously injected into BALB/C SCID mice. The function of TILs was evaluated by measuring tumor sizes in tumor-bearing mice, T helper (Th)1 (tumor necrosis factor (TNF)-α, interferon (IFN)-γ) and Th2 cytokine levels (IL-10 and IL-4) in serum and cytotoxicity of mouse natural killer (NK) and CD8⁺ T cells.

Results  IL-18 and IL-12 synergistically inhibited the growth of SGC-7901 cells in vivo and significantly extended the survival rate of SGC-7901-bearing mice (66.7% vs. 13.7%, P <0.01). Moreover, TILs could promote the secretion of TNF-α and IFN-γ ((130.34±7.65) vs. (210.63±12.31) pg/ml, P <0.01; (14.23±1.97) vs. (30.52±2.12) pg/ml, P <0.01), and downregulate IL-10 and IL-4 secretion ((103.72±11.21) vs. (61.36±5.41) pg/ml, P=0.021; (49.36±4.67) vs. (28.48±3.86) pg/ml, P=0.024).

Conclusion  IL-18 and IL-12 can synergistically enhance cytotoxic functions of TILs from human gastric cancer.

     

重症肺炎患者血清和肺泡灌洗液中IL-6和IL-10水平的变化及其与预后的相关性

目的：观察重症肺炎患者血清和肺泡灌洗液中白介素-6(IL-6)和白介素-10(IL-10)水平的变化,并探讨其与临床预后的关系,为临床治疗提供相关依据。方法选取我院重症医学科2014年10月至2015年12月收治的99例重症肺炎患者为研究对象,依据患者病程第10天的临床转归,将其分为好转组59例和恶化组40例。分别在患者病程第1、5、10天,检测其血清和肺泡灌洗液中IL-6、IL-10的水平,同时统计临床肺部感染评分(CPIS),并分析CPIS评分与IL-6、IL-10的水平的相关性。结果第5天和第10天时,好转组血清中IL-6水平分别为(43.18±16.79) pg/mL和(26.88±7.65) pg/mL,明显低于恶化组的(49.77±14.26) pg/mL和(39.73±8.12) pg/mL;好转组肺泡灌洗液中IL-6水平分别为(70.43±28.65) pg/mL和(35.17±9.81) pg/mL,明显低于恶化组的(81.52±20.14) pg/mL和(72.17±19.81) pg/mL,差异均具有统计学意义(P<0.05);组内比较,好转组和恶化组患者血清和肺泡灌洗液中IL-6水平均随着时间推移出现显著下降,差异均具有统计学意义(P<0.05)。第5天和第10天好转组患者血清中IL-10水平分别为(5.21±1.77) pg/mL和(3.63±1.54) pg/mL,明显低于恶化组的(9.76±3.11) pg/mL和(13.16±3.82) pg/mL,好转组肺泡灌洗液中IL-10水平分别为(7.75±1.8) pg/mL和(4.13±1.21) pg/mL,明显低于恶化组的(12.17±3.26) pg/mL和(15.82±4.88) pg/mL,差异均具有统计学意义(P<0.05);好转组患者血清IL-10随着时间推移逐渐降低,差异均具有统计学意义(P<0.05),而恶化组患者血清和肺泡灌洗液中IL-10的水平则持续性升高,差异均具有统计学意义(P<0.05)。相关性分析结果显示,CPIS评分与血清和肺泡灌洗液中IL-6、IL-10的水平均呈明显的正相关(95%CI=0.8118~0.9992,P=0.0018;95%CI=0.9539~0.9998,P=0.0002)。结论动态监测重症肺炎患者血清和肺泡灌洗液中IL-6、IL-l0的水平及变化趋势,能够反映患者预后情况,值得在临床中借鉴和推广。     

绝经后妇女骨密度与血清1，25-（OH）2D3，25-OHD3浓度关系的研究

目的：研究绝经后妇女无优势手臂骨密度与血清1,25-（OH）2D3,25-OHD3含量的关系。方法：采用定量超声骨量分析系统（QUS）测量82例绝经后妇女无优势前臂远端骨密度（BMD）以诊断骨质疏松,采用放射免疫法（RIA）和ELISA检测骨质疏松组和骨量正常组妇女血清中1,25-（OH）2D3,25-OHD3水平。结果：绝经后骨质疏松组妇女血清1,25-（OH）2D3,25-OHD3含量分别为（26.97±6.78）pg/ml和（34.75±12.62）nmol/L,骨量正常组分别为（34.75±12.62）pg/ml和（42.03±12.63）nmol/L,骨质疏松妇女血清1,25-（OH）2D3、25-OHD3含量明显低于骨量正常组（P〈0.01）。绝经后妇女无优势前臂骨密度与血清1,25-（OH）2D3含量存在明显相关性（r=0.387,P=0.001）,25-OHD3含量与前臂骨密度也存在相关性。结论：血清1,25-（OH）2D3水平与无优势前臂骨密度明显相关,是绝经后妇女骨量降低的重要原因之一。     

IL-13在盆腔子宫内膜异位症患者血清和腹腔液中的表达及意义

目的 通过检测及探讨子宫内膜异位症（EMs）患者血清及腹腔液中IL-13水平的变化,初步推测其在EMS发病中的作用.方法 采用酶联免疫吸附试验（ELISA）,检测EMs组32例患者和对照组20例非EMs患者腹腔液及血清中IL-13的水平,比较两组之间的差异;并将EMs组血清及腹水IL-13水平分别与EMs R-AFS评分进行相关性分析.结果 EMs组血清IL-13水平[（6.95±2.74）pg/ml]明显低于对照组[（12.87±9.70） pg/ml]（P＜0.01）;EMs组腹腔液IL-13水平[（3.26±1.81）pg/ml]明显低于对照组[（8.49±6.65）pg/ml]（P＜0.01）.根据期别将EMs组分为轻度亚组（Ⅰ、Ⅱ期）和重度亚组（Ⅲ、Ⅳ期）,血清IL-13水平轻度亚组[（7.85±2.09）pg/ml]与对照组[（12.87±9.70） pg/ml]及轻度亚组[（7.85±2.09 ）pg/ml]与重度亚组[（6.34±3.00） pg/ml]均无统计学差异,重度亚组[（6.34±3.00）pg/ml]明显低于对照组[（12.87±9.70）pg/ml]（P＜0.05）;腹腔液IL-13水平重度亚组[（2.43±1.43）pg/ml]明显低于轻度组[（4.48±1.63）pg/ml]（P＜0.01）,后者又显著低于对照组[（8.49±6.65）pg/ml]（P＜0.05）.EMs组腹腔液IL-13表达水平与其R-AFS评分呈负相关（r=-0.52,P＜0.05）.结论 EMs患者腹腔液中IL-13的水平显著降低,且与疾病的严重程度成负相关,推测IL-13在EMs的发病中起重要的作用.     

初诊2型糖尿病患者血清活性维生素D3水平的变化及其临床意义

目的：观察初诊2型糖尿病患者血清活性维生素D3水平的变化情况,为临床治疗和干预提供理论依据。方法选取2014年1月至2015年6月期间就诊于我院干部病科和内分泌科的200例2型糖尿病患者设为观察组,选取同期就诊于我院的100健康查体者作为对照组。比较两组受检者的年龄、病程、体质指数(BMI)、甘油三酯(TG)、高密度脂蛋白(HDL-C)、低密度脂蛋白(LDL-C)及胰岛素分泌指数(HOMR-β)、胰岛素抵抗指数(HOMR-IR)、血清活性维生素D3水平,以及谷丙转氨酶(ALT)、谷草转氨酶(AST)、肌酐(CRE)、血钙、血磷水平。结果(1)两组受检者的血清TG、TC、ALT、AST、CRE、血钙、血磷水平比较差异均无统计学意义(P>0.05);(2)与对照组比较,观察组患者的FPG [(9.85±2.50) mmol/L vs (4.84±0.54) mmol/L]、2 hPG [(10.28±2.28) mmol/L vs (5.21±0.56) mmol/L]、HbA1c [(7.66±2.36)% vs (4.62±0.47)%]明显升高,FINS [(7.40±5.06)μU/ml vs (11.87±6.81)μU/ml ]、HOMR-β[(25.81±20.13) vs (254.33±102.14)]、1,25-(OH)D3[(15.57±3.58) ng/ml vs (51.72±8.96) ng/ml]水平明显降低,差异具有统计学意义(P<0.05),而HOMR-IR[(3.40±2.83) vs (2.32±0.63)]与对照组比较差异无统计学意义(P>0.05);(3)1,25-(OH)D3与HOMR-β呈正相关(r=0.72,P<0.05),与HOMR-IR无明显相关性(r=-0.11,P>0.05)。结论初诊2型糖尿病患者血清1,25-(OH)D3降低,低1,25-(OH)D3水平影响患者的胰岛素β细胞的分泌功能。     

Microinflammation is involved in the dysfunction of arteriovenous fistula in patients with maintenance hemodialysis

Background Vascular access （VA） dysfunction is a major clinical complication in the hemodialysis population and has a direct effect on dialysis outcome. This study was conducted to explore the role of microinflammation in the VA dysfunction in maintenance hemodialysis patients.
Methods Forty-seven patients （male 35 and female 12） receiving maintenance hemodialysis were included for this study. They were divided into three groups： group 1 （n=15）, patients with initial hemodialysis and new arteriovenous fistula （AVF）; group 2 （n=18）, patients treated with hemodialysis for long term with well-functional VA; group 3 （n=14）, maintenance hemodialysis patients with VA dysfunction. Biochemical parameters and serum tumor necrosis factor-α （TNF-α）, interleukin 6 （IL-6） and monocyte chemoattractant protein-1 （MCP-1） were determined. High-sensitivity C-reactive protein （hs-CRP） was determined by latex-enhanced immuno-nephelometric method. Tissues of radial artery were taken from group 1 and group 3 for the histological study. Expression of CD68 and MCP-1 in the radial artery was determined by immunohistochemistry.
Results Serum hs-CRP in group 3 was significantly higher than those in group 1 and group 2 （（7.40±2.42） mg/L vs （4.21±1.62） mg/L and （5.04±3.65） mg/L, P 〈0.01 and P 〈0.05, respectively）. Serum TNF-α in group 3 was significantly higher than those in group 1 and group 2 （（64.03±9.29） pg/ml vs （54.69±12.39） pg/ml and （54.05±7.68） pg/ml, P 〈0.05 and P 〈0.01, respectively）. Serum IL-6 in group 3 was also significantly higher than those in group 1 and group 2 （（70.09±14.53） pg/ml vs （56.43±10.11） pg/ml and （60.77±9.70） pg/ml, P 〈0.01 and P 〈0.05, respectively）. Patients in group 3 had a thicker internal layer of vessels than in group 1 （（0.356±0.056） mm vs （0.111±0.021） mm, P 〈0.01）. Expression of CD68 and MCP-1 in the fistula vessel walls in group 3 were much higher than those in group 1     

淋巴引流及ω-3多不饱和脂肪酸干预对大鼠肠道缺血再灌注的影响

目的 观察大鼠肠道缺血再灌注(Ⅰ/R)损伤时肠淋巴液引流对高迁移率族蛋白1(HMGB1)、炎症因子和内毒素的影响以及ω-3多不饱和脂肪酸(ω-3 PUFA)干预的效果.方法 72只SD大鼠随机区组法随机分为单纯引流组、Ⅰ/R组、Ⅰ/R+引流组(每组8只)和胃造口组[正常饮食(N)组、普通肠内营养(EN)组、普通肠内营养加ω-3 PUFA(PUFA)3大组,每大组再根据是否行Ⅰ/R 和引流分为2组,每组8只].单纯引流组只引流180 mⅠn淋巴液不行Ⅰ/R损伤;Ⅰ/R、VR+引流组行肠系膜上动脉夹闭60 mⅠn再灌注120 mⅠn;Ⅰ/R+引流组同时行肠淋巴液引流180 mⅠn.胃造口组大鼠均先行胃造口手术,分别给予不同营养5 d后造模,各引流组同前进行肠淋巴液引流180 mⅠn.手术完毕后分别取血清和淋巴液,定量检测内毒素,酶联免疫吸附试验(ELⅠSA)定量检测炎症因子以及HMGB1.结果 Ⅰ/R+引流组淋巴液中内毒素、炎症因子以及HMGBl均高于单纯引流组[均P＜0.05,白细胞介素(ⅠL)-6(30±8)pg/ml比(20±6)pg/ml,内毒素(0 029±0.011)U/ml比(0 008±0 005)U/ml];Ⅰ/R+引流组血清中内毒素、炎症因子均低于Ⅰ/R组(均P＜0 05).在胃造口组中,N 组和EN组的淋巴液中肿瘤坏死因子(TNF)-α与HMGBl均高于PUFA组[(46±17)pg/ml、(54±16)pg/ml比(28±9)pg/ml,(4.8±1.6)ng/ml、(5.3±1.8)ns/ml比(3.0±1.0)ng/ml,均P＜0.05].PUFA(Ⅰ/R)组血清中内毒素、炎症因子以及HMGBl均低于N(Ⅰ/R)组(均P＜0.05),PUFA(Ⅰ/R+引流)组血清中TNF-α与HMGBl均低于N(Ⅰ/R+引流)组(均P＜0 05).结论 引流肠淋巴液能够降低肠道Ⅰ/R损伤时内毒素、炎症因子和HMGB1的水平,减轻大鼠肠道Ⅰ/R引起的损伤.ω-3PUFA的干预对于肠道Ⅰ/R引起的损伤有一定的保护作用,对于减轻炎症反应有积极作用.

Abstract：

Objective To investigate the effects of lymphatic drainage and ω-3 polyunsaturated fatty acid (ω-3PUFA) on high mobility group box 1 (HMGB1) , inflammatory cytokines and endotoxin in rats with intestinal ischemia-reperfusion (Ⅰ/R) injury. Methods A total of 72 SD rats were randomly divided into drainage-alone group, Ⅰ/R group, ischemia-reperfusion plus drainage (Ⅰ/R+D) group (n=8 each)and 3 groups with 16 rats undergoing gastrostomy in each group: normal diet (N) group, enteral nutrition (EN) group and enteral nutrition & ω-3PUFA (PUFA) group. And they were further divided into 2 subgroups (n=8). The rats in Ⅰ/R and Ⅰ/R+D groups were subjected to a 60-min ischemia follow by 120-min reperfusion injury of superior mesenteric artery. When the rats suffered Ⅰ/R injury, intestinal lymph was drained for 180 min in the Ⅰ/R+D group. The rats in the drainage-alone group received 180-min lymph drainage without Ⅰ/R injury. After 5 days with different nutrition regimes, the models were established similarly. The rats in the Ⅰ/R+D sub-groups were treated with intestinal lymph drainage for 180 min. The serum and lymph samples were collected post-operatively. Endotoxin was detected by a Limulus kit. The inflammatory cytokines and high mobility group box 1 (HMGB1) were analyzed by enzyme-linked immunosorbent assay (ELISA).Results Endotoxin, inflammatory cytokines and lymphatic HMGB1 of lymphatic in the Ⅰ/R+D group were higher than those in the drainage-alone group [all P＜0.05 , IL-6 :(30±8) pg/ml vs (20±6) pg/ml, endotoxin: (0.029±0.011) U/ml vs (0.008+0.005) U/ml].The serum levels of endotoxin and inflammatory cytokines in the Ⅰ/R+ D group were lower than those in the Ⅰ/R group (P＜0.05).The lymphatic levels of TNF-a (tumor necrosis factor-alpha) and HMGB1 in the N and EN groups were higher than those in the PUFA group[TNF-α: (46±17)pg/ml, (54±16)pg/ml vs(28±9) pg/ml, HMGB1: (4.8±1.6) ng/ml, (5.3±1.8) ng/ml, (3.0±1.0) ng/ml, all P＜0.05) ].The serum levels of endotoxin, inflammatory cytokines and HMGB1 in the PUFA(l/R) group were lower than those in the N(Ⅰ/R) group (F＜0.05).The levels of TNF-a and HMGB1 were lower in the PUFA (Ⅰ/R+D) group than those in the N(Ⅰ/R+ D) group (both P＜0.05).Conclusion Lymphatic drainage may reduce the levels of endotoxin, inflammatory cytokines and HMGB1 so as to alleviate the intestinal Ⅰ/R injury. The intervention of ω-3PUFA has some protective effect through relieving inflammation.     

Effects of heme oxygenase-1 gene modulated mesenchymal stem cells on vasculogenesis in ischemic swine hearts

Background  Mesenchymal stem cells (MSCs) transplantation may partially restore heart function in the treatment of acute myocardial infarction (AMI). The aim of this study was to explore the beneficial effects of MSCs modified with heme xygenase-1 (HO-1) on post-infarct swine hearts to determine whether the induction of therapeutic angiogenesis is modified by the angiogenic cytokines released from the implanted cells.

Methods  In vitro, MSCs were divided into four groups: (1) non-transfected MSCs (MSCs group), (2) MSCs transfected with the pcDNA3.1-Lacz plasmid (Lacz-MSCs group), (3) MSCs transfected with pcDNA3.1-hHO-1 (HO-1-MSCs group), and (4) MSCs transfected with pcDNA3.1-hHO-1 and pretreatment with an HO inhibitor, tin protoporphyrin (SnPP) (HO-1-MSCs+SnPP group). Cells were cultured in an airtight incubation bottle for 24 hours, in which the oxygen concentration was maintained at <1%, followed by 12 hours of reoxygenation. After hypoxia/reoxygen treatment, ELISA was used to measure transforming growth factor (TGF-β) and fibroblast growth factor (FGF-2) in the supernatant. In vivo, 28 Chinese mini-pigs were randomly allocated to the following treatment groups: (1) control group (saline), (2) Lacz-MSCs group, (3) HO-1-MSCs group, and (4) HO-1-MSCs + SnPP group. About 1×10⁷ of autologous stem cells or an identical volume of saline was injected intracoronary into porcine hearts 1 hour after MI. Magnetic resonance imaging (MRI) assay and postmortem analysis were assessed four weeks after stem cell transplantation.

Results  Post hypoxia/reoxygenation in vitro, TGF-β in the supernatant was significantly increased in the HO-1-MSCs ((874.88±68.23) pg/ml) compared with Lacz-MSCs ((687.81±57.64) pg/ml, P <0.001). FGF-2 was also significantly increased in the HO-1-MSCs ((1106.48±107.06) pg/ml) compared with the Lacz-MSCs ((853.85±74.44) pg/ml, P <0.001). In vivo, at four weeks after transplantation, HO-1 gene transfer increased the capillary density in the peri-infarct area compared with the Lacz-MSCs group (14.24±1.66/HPFs vs. 11.51±1.34/HPFs, P <0.001). Arteriolar density was also significantly higher in HO-1-MSCs group than in the Lacz-MSCs group (7.86±2.00/HPFs vs. 6.45±1.74/HPFs, P=0.001). At the same time, the cardiac function was significantly improved in the HO-1-MSCs group compared with the Lacz-MSCs group ((53.17±3.55)% vs. (48.82±2.98)%, P <0.05). However, all these effects were significantly abrogated by SnPP.

Conclusion  MSCs provided a beneficial effect on cardiac function after ischemia/reperfusion by the induction of therapeutic angiogenesis, and this effect was amplified by HO-1 overexpression.

     

urantide对小鼠急性肝细胞凋亡的影响及其机制

目的 探讨urotensinⅡ(UⅡ)特异性受体(UT)特异性拮抗剂urantide对急性肝细胞凋亡的影响及其机制.方法 雄性BALB/c小鼠按随机排列表法随机分为4组(每组6只):健康对照组、预处理对照组、模型组和预处理模型组.预处理对照组和预处理模型组给予尾静脉注射0.6 mg/kg urantide预处理,健康对照组和模型组则给予相同体积生理盐水.30min后模型组及预处理模型组立即以脂多糖联合D-半乳糖胺( D-GalN)腹腔注射诱导急性肝细胞凋亡小鼠模型,健康对照组和预处理对照组则给予相应体积的生理盐水.用原位末端转移酶标记(TUNEL)技术分析和半胱氨酸蛋白酶(caspase)-3活性测定检测肝细胞凋亡程度;用反转录(RT)-PCR及酶联免疫吸附试验( ELISA)方法检测肝组织及血浆前炎细胞因子肿瘤坏死因子(TNF)-α、干扰素(IFN)-γ和白细胞介素(IL)-1β的表达与分泌水平.结果 模型组可见大量肝细胞凋亡,而预处理模型组肝细胞凋亡指数和caspase-3活性均明显低于模型组[(26±11)％比(77±20)％,(2.50±0.83) pmol·min-1·mg-1比(3.76±0.42) pmol·min -·mg-1,均P＜0.01];肝组织前炎细胞因子TNF-α、IL-1β和IFN-γ的mRNA相对表达水平及蛋白分泌水平也均明显低于模型组[1.69±0.47比3.57±0.79、0.31±0.02比0.46±0.06、2.81±0.72比3.35±0.84,(233±36) pg/ml比(441±157) pg/ml、(228 ±21) pg/ml比(364±20) pg/ml、(93.8±5.2) pg/ml比(180.3±4.3) pg/ml,均P＜0.01].结论 urantide可通过抑制前炎细胞因子的表达与分泌抑制脂多糖/D-GalN诱导的急性肝细胞凋亡.这表明UⅡ/UT受体系统在急性肝衰竭免疫炎性损伤中起关键性的作用,并有可能成为ALF药物治疗的新靶点.     

Intermittent hypercalcaemia and vitamin D sensitivity in Hodgkin's disease

A patient with Hodgkin's disease spontaneously developed steroid-responsive hypercalcaemia during two consecutive summers. Administration of 3000 U/day of vitamin D, while he was normocalcaemic, caused a sharp increase in serum 1,25(OH)2D3 (from 59 pg/ml to 142 pg/ml) and subsequently hypercalcaemia while serum 25(OH)D3 rose moderately within the normal range (from 2.8 ng/ml to 10 ng/ml). During a spontaneous episode of hypercalcaemia which was accompanied by increased circulating 1,25(OH)2D3 concentrations, administration of hydrocortisone decreased serum 1,25(OH)2D3 rapidly (from 115 pg/ml to 62 pg/ml) and eventually led to normocalcaemia while serum 25(OH)D3 remained unchanged. Thus the disturbances of mineral metabolism found in this patient with Hodgkin's disease are very similar to those previously described in sarcoidosis.     

胶质瘤环氧化酶-2抑制剂塞来昔布对肿瘤微环境及生物学行为的影响

目的 探讨选择性环氧化酶-2（cyclooxygenase-2,COX-2）抑制剂塞来昔布（celecoxib）对大鼠脑胶质瘤免疫微环境及生物学行为的影响.方法 建立大鼠脑胶质瘤模型,观察celecoxib治疗后肿瘤大小及大鼠生存期;免疫组化法检测肿瘤组织COX-2、转化生长因子-β（transforming growth factor-beta,TGF-β）、血管内皮生长因子（vascular endothelial growth factor,VEGF）蛋白表达;流式细胞术检测肿瘤微环境CD4＋CD25＋T细胞亚群数量;酶联免疫法（ELISA）检测大鼠血清中前列环素E2（Prostaglandin E2,PGE2）、白介素-l0（IL-l0）、白介素-12（IL-12）分泌水平.结果 Celecoxib治疗组肿瘤体积明显小于对照组[（103.67±5.54） mm3,（151.60±8.34） mm3,P＜0.01],并可延长大鼠生存期[（42.87± 1.72）d,（26.13± 1.53）d,P＜0.01],肿瘤组织中COX-2、VEGF及TGF-β蛋白的表达均出现下调,其肿瘤微环境中CD4＋CD25＋T细胞亚群比例明显降低（5.32％,7.84％,P＜0.05）.血清中PGE2与IL-10水平明显下降[（223.66±33.79）pg/ml,（344.15±41.09） pg/ml,（98.69± 10.99） pg/ml,（133.37± 13.15） pg/ml,P＜0.01],而IL-12分泌明显增多[（237.20±37.31） pg/ml,（117.90±19.20） pg/ml,P＜0.01].结论 Celecoxib可通过多种途径改善肿瘤局部免疫微环境,从而改变肿瘤的生物学行为,延缓肿瘤发展,并为肿瘤治疗提供良好的基础环境.     

舍曲林与文拉法辛对首发抑郁症患者血清细胞因子水平影响的对照研究

目的 探讨首发抑郁症患者的细胞因子水平变化以及比较不同种类的抗抑郁剂舍曲林与文拉法辛对患者血清细胞因子水平的影响.方法 分别测定80例首发抑郁症患者的血清IL-2、IL-6、TNF-α、IL-10水平,并与40例健康对照者进行比较;80例首发抑郁症患者依抽签法分为人数相等的2组,分别经舍曲林、文拉法辛治疗6周后进行疗效评估,对舍曲林、文拉法辛治疗敏感的患者进行IL-2、IL-6、TNF-α、IL-10水平测定,并进行统计分析.结果 首发抑郁症患者的血清IL-2、IL-6、TNF-α细胞因子水平[(6.79±2.89)pg/ml,(10.12±4.52)pg/ml,(14.81±4.38)pg/ml]显著高于健康对照组[(4.06±2.05)pg/ml,(6.04±1.79)pg/ml,(10.69±2.54)pg/ml];舍曲林、文拉法辛治疗后较治疗前IL-6水平有显著降低(P＜0.05),IL-2、TNF-α、IL-10差异无显著性(P＞0.05),而舍曲林、文拉法辛两种抗抑郁剂对细胞因子水平的影响差异无显著性(P＞0.05).结论 首发抑郁症患者存在细胞因子水平的改变,并可以通过舍曲林、文拉法辛的治疗来矫正,而舍曲林、文拉法辛可能对细胞因子有相同的免疫作用机制.     

旋毛虫感染对过敏性哮喘小鼠IL-4及IL-5的影响

目的通过研究旋毛虫感染鼠在过敏性哮喘发病时血清及支气管肺泡灌洗液中IL-4和IL-5的水平变化．探讨旋毛虫感染对过敏性哮喘的作用。方法取雌性、8周龄BALB／C小鼠20只,随机分为4组．A组为空白对照组,B组为单纯过敏性哮喘组,C组为感染旋毛虫后哮喘组,D组为单纯感染旋毛虫组。C、D组小鼠经灌胃法感染旋毛虫囊包幼虫200—300条;4周后,以卵清白蛋白（OVA）分别对B、C组小鼠进行致敏激发,建立过敏性哮喘模型;8周后,取小鼠血清、肺泡灌洗液,用ELISA法检测IL-4和IL．5水平。结果血清中A、B、C、D组小鼠IL-4水平分别为（136．4948±31．1670）、（209．8004±21．5652）、（162．7892±17．6829）、（182．7092±12．0841）pg／L;IL-5水平分别为（7．3101±1．8600）、（15．9997±2．28208）、（9．2411±1．7280）、（11．4135±2．10776）ng／L;与B组比较,C组小鼠IL-4、IL-5水平均降低,差异有统计学意义（P〈0．05）。肺泡灌洗液中A、B、c、D组小鼠IL．4水平分别为（169．1636±9．9997）、（232．9076＋17．9501）、（172．3508＋-19．1895）、（193．0676＋-16．5708）pg／L;IL-5水平分别为（12．8618±1．7799）、（23．8460±5．507）、（15．7560±4．5419）、（20．3440±3．9713）ng／L;与B组比较,C组小鼠IL-4、IL-5水平降低,差异有统计学意义（P〈0．05）。结论旋毛虫感染对过敏性哮喘小鼠IL-4、IL-5的表达有抑制作用。     

白细胞介素1受体拮抗剂对大鼠过敏性哮喘的治疗作用及相关机制研究

目的 研究白细胞介素1受体拮抗剂(IL-1ra)对大鼠过敏性哮喘的治疗作用及相关机制.方法 雌性SD大鼠分为正常对照组、哮喘模型组、哮喘模型低剂量IL-1ra(6 mg/kg)治疗组(低剂量治疗组)和高剂量IL-1ra(30 mg/ks)治疗组(高剂量治疗组).每组10只.采用卵白蛋白腹腔及皮下注射致敏加雾化吸入激发的方法建立大鼠过敏性哮喘模型.激发前尾静脉注射不同剂量的IL-1ra.通过检测各组大鼠肺功能、肺泡灌洗液(BALF)炎性细胞构成、肺组织病理切片、血清总IgE含量等指标评价治疗效果,利用半定量RT-PCR检测信号转导子和转录激活子6(STAT6)mRNA以及核因子κB(NF-κB)mRNA表达情况.结果 哮喘模型组大鼠呼吸速率[(206±11)次/min]、呼气流量[(77±8)μl/s]、BALF嗜酸粒细胞比例(24.8%±1.3%)、血清总IgE含量[(72.5±8.1)ng/ml]、STAT6表达(0.294±0.048)和NF-κB表达(0.686±0.052)均明显高于低剂量治疗组[分别为(183±9)移c/min,(64±5)μl/,s,18.5%±3.1%,(63.4±4.8)ng/ml,0.229±0.038,0.613±0.062,均P＜0.05]和高剂量治疗组[分别为(181±11)次/min,(57±4)μl/s,14.7%±2.1%,(41.4±7.8)ng/ml,0.194±0.076,0.352±0.267,均P＜0.05].高剂量治疗组治疗效应优于低剂量治疗组(P＜0.05).肺组织病理检查与以上结果相近.结论 IL-1ra对大鼠过敏性哮喘具有明显的治疗效果,这种作用可能是通过同时调控STAT6 mRNA和NF-κB mRNA的表达实现的.