La、hVAP-33、eIF2B γ和HCV IRES特异性小干扰RNA抑制HCV的复制和表达

目的 证实La自身抗原、33kD人类囊相关膜蛋白(hVAP-33)和真核细胞翻译起始因子第3亚单位(eIF2B γ)是HCV在细胞内的协同感染因子,通过抑制Huh7细胞内这些因子的表达可抑制HCV复制和表达. 方法 分别设计合成3条HCV内部核糖体进入位点(IRES)的小干扰RNA(siRNAs),转染Huh7-HCV细胞后筛选出其中沉默效率最高的1条.以HCV假病毒感染Huh7细胞后48 h,分别以上述HCV IRES siRNA和之前试验中已经筛选出的La、hVAP-33和eIF2B γ特异性siRNAs单独或者不同组合转染Huh7-HCV细胞,利用荧光定量PCR方法检测HCV核心基因并计算△△CT值(相对定量法),比较不同siRNAs及组合对目的基因沉默的效率;同时以Western blot观察HCV核心蛋白表达量的差异.结果 La自身抗原与IRES特异性siRNA共转染对HCV表达的抑制效率最高,使HCV核心蛋白基因相对表达量下降了约41％;4种基因特异性siRNAs 对HCV在Huh7细胞中的复制和表达均有不同程度的抑制作用,La、hVAP-33和eIF2Bγ特异性siRNAs分别与IRES siRNA联合均较其单独转染对目的基因的抑制效率高.结论 可以认为La自身抗原、hVAP-33和eIF2Bγ是HCV在宿主细胞内的协同感染因子,通过对Huh7细胞内协同感染因子及HCV IRES基因沉默后可以显著减少HCV的表达.     

ADAM33基因单核苷酸多态性及单体型与慢性阻塞性肺疾病的关联性研究

目的 探讨中国华北地区汉族人群ADAM33基因S1、S2位点单核苷酸多态性及单体型与慢性阻塞性肺疾病(COPD)及肺功能的关联性.方法 应用DNA直接测序的方法,对90例COPD患者和90名健康对照者的ADAM33基因S1、S2位点基因型进行检测;应用SHEsis在线软件构建单体型并进行单体型关联分析.结果 ①病例组和对照组中S1位点基因型及等位基因频率分布比较差异无统计学意义(P＞0.05),S2位点基因型及等位基因频率分布比较差异有统计学意义(P ＜0.05).②Logistic回归分析表明:ADAM33基因S1位点不同基因型COPD发生的相对危险度比较差异无统计学意义(P＞0.05);S2位点不同基因型COPD发生的相对危险度比较,差异有统计学意义(P＜0.05),其中G/G+C/G基因型的OR值为2.364(95％CI 1.251～4.466).③COPD病例组S2位点基因型与肺功能相关临床指标的关系显示:3种基因型FEV1％预计值比较差异无统计学意义而FEV1/FVC比较差异有统计学意义,其中G/G基因型与C/C、C/G基因型相比FEV1/FVC下降更明显.④SHEsis在线软件对S1、S2位点进行单体型分析结果显示,单体型CG在COPD组和对照组中比较差异有统计学意义(P＜0.05).结论 在中国华北地区汉族人群中,ADAM33基因与COPD的易感性有关,但与疾病的严重程度无关.     

The possible role of interleukin-33 as a new player in the pathogenesis of contrast-induced nephropathy in diabetic rats

Introduction: Patients with diabetic kidney disease (DKD) are more prone to contrast-induced nephropathy (CN). Apoptosis and autophagy were found to be essential in the pathogenesis of DKD. Interleukin-33 (IL-33) is a cytokine, but its role in DKD and CN is unknown. As IL-33 is modulated by apoptosis, we aimed to determine the relationship between IL-33 apoptosis and autophagy in DKD with CN. Materials and methods: Thirty male Sprague–Dawley rats were enrolled and randomly allocated into three groups. The first group was comprised of healthy rats (HRs), whereas the other two groups were made up of diabetic rats (DRs) and diabetic rats with CN (DRs?+?CN). All groups except the HRs received 50?mg/kg/day of streptozotocin (STZ). The DRs?+?CN group was induced by administering 1.5?mg/kg of intravenous radiocontrast dye on the 35th day. Results: We observed increased IL-33 in the kidney tissue following induction of CN in the DRs. The DRs showed moderate immunopositivity, and the DRs?+?CN showed severe immunopositivity for caspase-3, cleaved caspase-3, caspase-8, caspase-9, LC3B, and Beclin-1 in tubular cells and glomeruli. The DRs also showed moderate immunopositivity in tubular cells, and the DRs?+?CN group showed severe immunopositivity for IL-33 in tubular cells. Increased caspase-3 was found in both glomeruli and tubuli; however, we could not demonstrate IL-33 in glomeruli. This could be secondary to inactivation of IL-33 via increased caspase-3 activity. Conclusion: The release of IL-33 from necrotic cells might induce autophagy, which can further balance the effects of increased apoptosis secondary to CN in DKD.     

Allergen-Dependent Differences in ILC2s Frequencies in Patients With Allergic Rhinitis

PurposeGroup 2 innate lymphoid cells (ILC2s) are a novel population of lineage-negative cells that induce innate type 2 responses by producing the critical Th2-type cytokines IL-5 and IL-13 in response to IL-25 and IL-33 stimulation. ILC2s accumulation in the peripheral blood of patients with allergic rhinitis (AR) is controversial; the precise role of ILC2s in the immunopathogenesis of AR is still not clear. We investigated the role of ILC2s in phenotypic AR sensitized to distinct allergens.MethodsFlow cytometric analysis of the peripheral blood of 7 healthy controls (HCs), 9 patients monosensitized to house dust mite (HDM), and 8 patients monosensitized to mugwort was performed to quantify ILC2s frequency. Peripheral blood mononuclear cells (PBMCs) were isolated from HDM-AR and mugwort-AR patients, and Lineage^- and Lineage⁺ cells were separated using a fluorescence-activated cell sorter (FACS). IL-5 and IL-13 levels in the supernatants of PBMCs, and Lineage^- and Lineage⁺ cells stimulated with IL-25 and/or IL-33 combined with IL-2 in vitro were assessed using the Milliplex magnetic bead kit.ResultsThe percentage of ILC2s was significantly elevated in HDM-AR patients compared to mugwort-AR patients and HCs, while no significant difference was found between mugwort-AR patients and HCs. IL-33±IL-25 plus IL-2 induced a significantly greater release of IL-5 and IL-13 in the PBMCs of HDM-AR patients compared to PBMCs of mugwort-AR patients. IL-25 plus IL-2 also induced a significantly greater release of IL-13 in the PBMCs of HDM-AR patients compared to PBMCs of mugwort-AR patients. Stimulation with IL-33 and/or IL-25 combined with IL-2 also induced a significantly greater IL-5 and IL-13 release from Lineage^- cells compared to Lineage⁺ cells.ConclusionsAR patients sensitized to HDM or mugwort allergen have distinct phenotypic and functional profiles in ILC2s frequencies. ILC2s mediate major type 2 immunity in the development of HDM-AR and may be a potential therapeutic target.     

Interleukin-33 facilitates neutrophil recruitment and bacterial clearance in S. aureus-caused peritonitis

Interleukin (IL)-33, a newly recognized member of IL-1 family of cytokines, plays an important role in polarizing Th2-associated immunity. Recently growing evidence indicates that IL-33 also represents a crucial mediator of antimicrobial infection. In this study, we investigated the effect of IL-33 on antibacterial response using an acute Staphylococcus aureus peritoneal infection model. Our results showed that IL-33 administration induced a rapid bacterial clearance and markedly reduced the S. aureus infection-related mortality. IL-33-treated mice displayed increased neutrophil influx into the focus of infection and higher concentrations of chemokine CXCL2 in the peritoneum than untreated mice. The beneficial effect of IL-33 priming was related to reversal of the S. aureus-induced reduction of CXCR2 expression on the surface of neutrophils. Furthermore, conditioning of neutrophils by IL-33 led to the enhancement of complement receptor 3 expression induced by S. aureus, which in turn facilitates the phagocytosis of opsonized S. aureus. Finally, neutrophils primed by IL-33 upregulated the production of reactive oxygen species and the subsequent killing activity for S. aureus. All together, these findings suggest that IL-33, through regulating multiple steps of neutrophil-mediated bactericidal function, provides a profound effect in host antimicrobial defense response.     

肥大细胞及IL-33/ST2在复发性自然流产小鼠模型中的作用初探

目的探讨复发性流产小鼠子宫和胎盘组织中肥大细胞的数量及白细胞介素-33(IL-33)/ST2的表达变化。方法 20只雌性CBA/J小鼠随机分为两组,每组10只,分别与雄性DBA/2和Balb/c小鼠按雌雄比例2:1合笼交配,建立正常妊娠组(CBA/J♀×Balb/c/2♂)和自然流产组(CBA/J♀×DBA/2♂)模型。妊娠第13.5天处死各组雌性小鼠,计数存活胚胎数和丢失胚胎数,计算胚胎丢失率。甲苯胺蓝染色法检测小鼠子宫组织中肥大细胞的数量。ELISA检测血清中干扰素-γ(IFN-γ)、IL-4的浓度,qRT-PCR和Western-blotting分别检测胎盘组织中IL-4、IFN-γ、IL-33、ST2的mRNA和蛋白表达水平。结果成功构建了复发性自然流产(RSA)的小鼠模型,RSA模型组胚胎丢失率显著高于正常妊娠组(16.2%vs.4.92%,P0.05);RSA组子宫组织中肥大细胞数显著低于正常妊娠组[(1.50±0.83)vs.(3.35±1.63)个](P0.05);ELISA结果显示,与正常妊娠组相比,RSA组小鼠血清中IFN-γ水平显著升高[(346.79±4.34)vs.(168.84±2.35)ng/L],IL-4水平显著降低[(98.46±5.81)vs.(157.56±9.35)ng/L](P均0.05);qRT-PCR和Western-blotting结果显示RSA组小鼠胎盘组织中IFN-γ表达水平显著升高,IL-4、IL-33、ST2表达水平显著降低(P0.05)。结论肥大细胞和IL-33/ST2可能参与了RSA小鼠体内Th1/Th2的调节,有助于妊娠的维持。     

白细胞介素-33在不同严重程度脓毒症中的变化

目的  分析血浆白细胞介素-33（IL-33）在不同严重程度脓毒症中的变化。方法  选取2012年1月- 2015年4月该院重症加强治疗病房（ICU）收治的120例脓毒症患者为研究对象。根据器官功能、感染严重程度将患者分为一般脓毒症组（36例）、严重脓毒症组（36例）和感染性休克组（48例）,并选取同期在该院行健康检查的120例志愿者作为对照组。比较患者ICU各时间的APACHE Ⅱ评分、降钙素原（PCT）、白细胞介素-33（IL-33）、白细胞介素-6（IL-6）、白细胞介素-1β（IL-1β）、肿瘤坏死因子-α（TNF-α）和可溶性人基质裂解素2（sST2）的变化;比较患者ICU住院时间和机械通气时间;比较不同预后脓毒症患者ICU各时间的IL-33水平变化。结果  脓毒症患者的APACHE Ⅱ评分、PCT和IL-33水平高于对照组,且脓毒症患者的APACHE Ⅱ评分、PCT和IL-33水平随患者病情加重而升高,差异有统计学意义（P <0.05）。脓毒症患者的ICU住院和机械通气时间随患者病情加重而延长,且感染性休克组的死亡率显著高于其余两组,差异有统计学意义（P <0.05）。脓毒症患者的APACHE Ⅱ评分,PCT、IL-33、IL-6、IL-1β、TNF-α、sST2水平与对照组比较,差异有统计学意义（P <0.05）。脓毒症患者的APACHE Ⅱ评分,PCT、IL-33、IL-6水平随ICU住院时间延长而降低,差异有统计学意义（P <0.05）。脓毒症患者的IL-1β、TNF-α、sST2水平随ICU住院时间延长,变化不明显（P >0.05）。入ICU 30 d后有9例患者死亡为死亡组,111例患者存活为存活组。死亡组患者的IL-33水平随ICU住院时间延长而上升,但差异无统计学意义（P >0.05）。存活组患者的IL-33水平随ICU住院时间延长而下降,死亡组患者入ICU 3 h内IL-33水平较存活组低,入ICU 24 h和5 d时IL-33水平高于存活组,差异有统计学意义（P < 0.05）。结论  脓毒症患者入ICU时血浆IL-33水平升高,而后随ICU住院时间延长而降低,且随患者病情加重而升高,因此血浆IL-33水平能提示患者的预后。

     

Frequent but nonrandom expression of myeloid markers on de novo childhood acute lymphoblastic leukemia

The expression of the myeloid markers CD13, CD33, and CD15 in two hundred and eighty-three cases of de novo childhood acute lymphoblastic leukemia (ALL) is examined. The expression of at least one marker is a frequent event which is noted in 64% and 74% of B- and T-lineage ALL cases, respectively. Certain patterns of myeloid antigen expression can be recognized including: no expression of CD13, CD33, and CD15 in mature B-ALL, significantly higher levels of CD13 and CD33 and significantly lower levels of CD15 in TEL-AML1-positive B cell precursor ALL, no expression of CD13 and CD33 in E2A-PBX1-positive B cell precursor ALL cases and common T-ALL (double positive for CD4 and CD8), and no expression of CD13 in MLL-AF4-positive B cell precursor ALL cases. Although the numbers in some ALL subtypes are small, these patterns are consistent with nonrandom expression of myeloid markers in de novo childhood ALL.