CD14基因多态性对严重烧伤后CD14表达的影响及其临床意义

目的探讨内毒素受体CD14C-159T基因多态性对烧伤后CD14合成与释放的影响及其与脓毒症易感性、患者预后的关系。方法26例烧伤面积大于30％的患者,采用聚合酶链反应及限制性内切酶HaeuI对PCR产物的消化作用检测CD14基因多态性。同时,对患者白细胞CD14、肿瘤坏死因子-α（TNF-α）mRNA表达,血清sCD14浓度与CD14-159位点基因型以及烧伤患者预后的关系进行了分析。结果Tr、TC、CC三种基因型患者白细胞CD14mRNA、TNF—amRNA表达,血清sCD14浓度存在明显差异。其中Tr、TC基因型CDl4mRNA表达均明显高于CC纯合子（P＜0．05或P＜0．01）,伤后第7天CC基因型血浆sCDl4水平显著低于TC基因型（P＜0．05）;同时,TT纯合子的TNF-αmRNA表达强度明显高于CC纯合子（P＜0．05）。此外,伤后第7、21天死亡组CDl4mRNA表达量显著高于存活组（P＜0．05）。结论CD14C-159T基因多态性可显著影响严重烧伤后CDl4的合成与释放,并与烧伤患者脓毒症易感性有关,T等位基因可能是患者预后不良的高危基因标志物。     

CD14基因启动子区-159位点多态性在重度烧伤患者中的分布及意义

目的 了解CD14基因启动子-159C/T基因多态性与严重烧伤患者CD14 mRNA表达及血浆可溶性CD14(sCD14)水平的关系.方法 2004年6月-2006年6月,选择2个笔者单位收治的烧伤总面积大于或等于30%TBSA的患者77例,分别于伤后1、3、5、7、14、21、28 d抽取患者静脉血,采用PCR-限制性片段长度多态性法检测血浆CD14-159C/T基因多态性,酶联免疫吸附测定法、RT-PCR法检测患者血浆sCD14的含量及白细胞CD14 mRNA表达.结果 77例患者的CD14基因C-159T基因型中,CC纯合子型7例占9.1%、TC杂合子型49例占63.6%、TT等位基因纯合子型21例占27.3%,T等位基因和C等位基因分布频率分别为59.1%和40.9%.经检验表明,此研究群体达到了Hard-Weinberg平衡.7例CC纯合子型患者中并发脓毒症3例占42.9%,49例TC杂合子型并发该症38例占77.6%,21例TT等位基因纯合子型并发该症15例占71.4%.3例CC纯合子型脓毒症患者中,1例出现MODS;38例TC杂合子型脓毒症患者19例出现MODS占50.0%;15例TT等位基因纯合子型脓毒症患者10例出现MODS占66.7%.伤后7～21 d TC杂合子型、TT等位基因纯合子型患者外周血CD14 mRNA表达明显高于CC纯合子型患者(P＜0.05或P＜0.01).伤后7d TC杂合子型、TT等位基因纯合子型患者CD14 mRNA表达达高峰,分别为1.18±0.25、1.15±0.35.烧伤后TC杂合子型、TT等位基因纯合子型患者血浆中sCD14含量较高,伤后5 d CC纯合子型患者血浆sCD14含量(85±46)μg/L显著低于TC杂合子型患者[(134±43)μg/L,P＜0.01];伤后21、28 d TC杂合子型、TT等位基因纯合子型患者sCD14含量明显高于CC纯合子型患者(P＜0.01).结论 大面积烧伤后CD14基因启动子-159位点多态性TT基因型可能是烧伤感染患者发生MODS的主要基因标志物之一.携带TT基因型的烧伤脓毒症患者并发MODS概率高于其他基因型.     

严重烧伤患者CD14基因多态性对高迁移率族蛋白B1表达的影响

目的 了解LPS受体CD14C-159T基因多态性对严重烧伤患者伤后高迁移率族蛋白B1(HMGB1)合成、释放的影响以及与脓毒症的关系.方法 采集35例烧伤总面积大于或等于30%TBSA患者伤后1、3、5、7、14、21、28 d静脉血.另设11名志愿者作为健康对照组.采用PCR-限制性片段长度多态性方法检测CD14-159C/T基因多态性,ELISA法检测血浆HMGB1水平,RT-PCR法检测HMGB1 tuRNA表达.对数据行χ~2检验、方差分析和t检验.结果 35例患者的CD14基因C-159T基因型中,CC纯合子型7例占20.0%、TC杂合子型16例占45.7%、TT等位基因纯合子型12例占34.3%.T等位基因和C等位基因分布的频率为57.2%和42.8%.验证表明,此研究群体达到了Hard-Weinberg平衡.在CD14C-159T基因型中,CC纯合子型患者发生脓毒症的概率较TC杂合子型、TT等位基因纯合子型低.3例CC纯合子型脓毒症患者中,仅1例死亡;9例TC杂合子型脓毒症患者中4例死亡;7例TT等位基因纯合子型脓毒症患者中4例死亡.与健康对照组比较伤后1 d患者血浆HMGB1水平即迅速升高,伤后14、21、28 d TC杂合子型、TT等位基因纯合子型患者血浆HMGB1水平均显著高于CC纯合子型(F值为3.5671、4.2035、3.8529,P<0.05或P<0.01).伤后14 d脓毒症组患者外周血白细胞HMGB1 mRNA表达量为1.5±0.5,显著高于非脓毒症组患者(1.2±0.4,t=-2.205,P<0.05).伤后7、21 d脓毒症组患者血浆HMGB1水平分别为(44±29)、(25±15)ng/mL,均高于非脓毒症组患者的(26±12)、(10±6)ng/mL(t值分别为-2.355、-3.872,P<0.05或P<0.01).结论 CD14C-159T基因多态性可显著影响严重烧伤后HMGB1的合成与释放,并与烧伤患者脓毒症易感性有关.     

CD14基因多态性与严重烧伤患者T淋巴细胞免疫功能的相关性研究

目的 探讨严重烧伤患者CD14基因多态性与T淋巴细胞免疫功能的关系.方法 采集77例烧伤体表总面积大于30%患者血标本,通过聚合酶链反应.限制性片段长度多态性方法检测CD14-159C/T基因多态性,观察其外周血T淋巴细胞增殖反应和白细胞介素-2(IL-2)的产生能力,并通过流式细胞仪检测T淋巴细胞CD4+/CD8+的比值、CD4+细胞的凋亡率.结果 严重烧伤后患者T淋巴细胞增殖能力明显下降,与CC纯合子患者比较,TT、TC基因型患者伤后第5、21、28天T淋巴细胞对丝裂原刺激增殖反应显著受抑(P<0.05或P<0.01).烧伤后携带TC、TT型患者IL-2产生一直处于较低水平,而CC型在伤后14 d分泌IL-2逐渐上升.与CC型患者比较,携带TT、TC型患者T淋巴细胞比值均降低,尤其在伤后第1、3、14、21、28天差异明显(P<0.05或P<0.01).三型CD3+CD4+T淋巴细胞凋亡率比较,TT型患者伤后第5、7、21天凋亡率显著高于CC型患者(P<0.05),TC型患者伤后7、14 d其凋亡率高于CC型患者(P<0.05),而,TT、TC型之间上述免疫功能指标比较均无显著差异(P>0.05).结论 CD14-159C/T多态性可影响大面积烧伤患者T淋巴细胞免疫功能状态,进而参与了严重感染并发症的发生与发展过程.     

肠内免疫营养对烫伤大鼠内毒素和CD14/肿瘤坏死因子-α mRNA表达的影响

目的研究肠内免疫营养物对烫伤大鼠内毒素和CD14 mRNA、肿瘤坏死因子 (TNF)-α mRNA表达的影响及机制。方法致64只SD大鼠总体表面积(TBSA)30％Ⅲ度烫伤, 随机分为肠内免疫营养组(EIN组,32只)和标准肠内营养组(EN组,32只),另取8只大鼠作为伤前正常对照组(N组)。EIN组和EN组均给予等热量(125 Kcal／dl)肠内营养液。分别于伤前、伤后 1、4、7、10 d抽取静脉血和肝组织,检测血清内毒素和TNF-α浓度,RT-PCR检测肝组织CD14 mR- NA、TNF-α mRNA。结果烫伤后EN、EIN组血清内毒素、TNF-α浓度比伤前血清内毒素 (0．125±0．050)和TNF-α浓度(0．85±0．27)显著升高(P<0．01),且肝组织CD14 mRNA、TNF-α mRNA的表达明显增多;在伤后4、7、10 d,EIN组血清内毒素和TNF-α浓度以及肝组织CD14 mR- NA、TNF-α mRNA的表达均比EN组显著降低(P<0．05或P<0．01)。结论肠内免疫营养与标准肠内营养相比,可明显降低烫伤后血清内毒素水平,减少肝组织CD14 mRNA、TNF-α mRNA的表达,从而降低血清TNF-α浓度,改善烫伤后机体炎症反应。     

SCD14、TNF-α、E-SLT及IL-10在感染发病中的作用及临床意义

目的探讨SCD14、TNF-α、E-SLT及IL-10在感染发病机制中的作用及其意义.方法采用酶联免疫吸附法测定37例腹部外伤合并感染患者及内毒素血症兔模型中可溶性CD14(SCD14)、肿瘤坏死因子α(TNF-α)、E选择素(E-SLT)及白细胞介素10(IL-10)血清浓度变化情况.结果与对照组相比,SCD14与TNF-α在术后第1天开始升高,E-SLT与IL-10在术后合并感染组中于第3天出现增高[(1.61±0.47)μg/ml与(28.63±8.29)pg/ml,(153.6±48.9)ng/ml 与(38.21±10.87)pg/ml,P均＜0.05];在兔内毒素血症中则分别于第30、120min开始增高[(0.50±0.02)μg/ml与(1.00±0.02)pg/ml,(20.9±0.5)ng/ml 与(49.7±0.5)pg/ml,P均＜0.05].结论血清SCD14、TNF-α、E-SLT及IL-10浓度的变化在反映感染发生发展及预后方面具有一定的价值.     

脂多糖结合蛋白(LBP)基因多态性对LBP及细胞因子诱生的影响

目的探讨脂多糖结合蛋白（LBP）C1306→T单核苷酸多态性（SNP）对全血培养LBP表达及细胞因子生成的影响。方法采集118例健康献血员静脉血，运用聚合酶链反应（PCR）及限制性内切酶Stu Ⅰ对PCR产物的消化作用检测C1306→T基因多态性，并采用全血细胞培养模型检测内毒素刺激前后LBP、肿瘤坏死因子（TNF）-α、白细胞介素（IL）-6及IL-10蛋白水平。结果118例健康献血员中，14例为T／C杂合子，104例为T／T纯合子。对照组及内毒索刺激组中T／C基因型上清液LBP浓度均明显高于T／T纯合子，而TNF-α、IL-6及IL-10对内毒索的反应性在两种基因型之间差异不明显（P〉0．05）。结论内毒素受体LBP C1306→T单核苷酸多态性可能间接影响LBP的蛋白水平，但与体外炎症介质的生成关系不明显。     

荆门地区CD14-159C/T多态性和幽门螺杆菌在胃癌中交联作用研究

目的探讨荆门地区人群CD14-159C/T多态性与胃癌关联性及其与幽门螺杆菌交互作用。 方法采用多聚酶链反应-限制性片段长度多态性（PCR-RFLP）方法,分析127例胃癌患者和127名健康者的CD14-159C/T基因型。非条件Logistic分析各基因型与发病中易感性关系以及与幽门螺杆菌的交互作用。 结果携带C（CC/CT）基因个体患病风险较非C基因携带者（TT）风险明显增加（OR=1.35,95% CI=1.22~2.56,P=0.000;校正OR=1.61,95% CI=1.21~3.01,P=0.000）。条件Logistic分析表明,携带CC/CT基因型幽门螺杆菌个体胃癌罹患风险是携带TT基因非幽门螺杆菌个体的3.39倍（OR=3.39,95% CI=2.66~5.36,P=0.000）（RERI=1.94,95% CI=1.41~2.77;API=0.59,95% CI=0.33~0.84;S=1.46,95% CI=1.37~2.66）。 结论CD14-159C/T多态性增加荆门地区个体胃癌的罹患风险,且与幽门螺杆菌在胃癌发病中存在协同效应。     

烫伤大鼠肝脏脂多糖结合蛋白和脂多糖受体CD14 mRNA表达的意义

目的　观察烫伤后肠源性内毒素移位与肝组织内毒素增敏系统———脂多糖结合蛋白/脂多糖受体CD14(LBP/CD14)的相互关系及其意义。　方法　采用大鼠 35 %TBSAⅢ度烫伤模型 ,动物随机分为正常对照组、烫伤和重组杀菌 /通透性增加蛋白治疗组。烫伤组和rBPI2 1　治疗组动物于伤后 12h活杀。留取肝组织和血标本分别检测组织LBP、CD14、肿瘤坏死因子α(TNFα)mR NA表达及肝脏功能指标。　结果　烫伤后肝组织内毒素含量显著降增高 (P <0 .0 1) ,且肝组织LBP/CD14、TNFαmRNA表达亦明显增多。给予rBPI2 1　治疗可显著降低肝组织内毒素水平 ,并不同程度抑制组织LBP/CD14、TNFα基因表达。同时 ,rBPI2 1　治疗组动物血清谷丙转氨酶水平显著降低 (P <0 .0 1)。　结论　烫伤后移位内毒素聚积于肝脏 ,可明显刺激机体局部组织LBP/CD14mRNA的表达 ,而组织LBP/CD14表达上调可能是严重烫伤增敏内源性内毒素作用的主要分子生物基础     

大鼠肝移植缺血再灌注后Kupffer细胞CD14基因及蛋白的表达

目的研究大鼠肝移植缺血再灌注后Kupffer细胞CD14基因及蛋白的表达,探讨其在再灌注损伤中的作用.方法分离培养大鼠肝移植缺血再灌注后0（对照组）、2、6、12 h（IR组）的Kupffer细胞,用逆转录聚合酶联反应（RT-PCR）检测Kupffer细胞CD14 mRNA的表达,用免疫印迹检测CD14蛋白合成,用酶联免疫吸附试验（ELISA）法测定培养上清TNF-α的分泌量.然后在上述时间点的细胞培养液中加入抗CD14抗体（anti-CD14组）,观察CD14抗体对TNF-α分泌的影响.结果再灌注后Kupffer细胞CD14 mRNA、蛋白以及TNF-α随观察时间点呈逐步上升趋势（与对照组相比,P＜0.01）.应用抗CD14抗体后,TNF-α表达较IR组明显降低（P＜0.01）.结论再灌注后Kupffer细胞CD14基因及蛋白的表达明显升高,TNF-α的合成和分泌也明显增强;抗CD14单抗能明显抑制TNF-α的产生;CD14在介导Kupffer细胞激活和肝移植缺血再灌注损伤中可能起重要作用.     

Influence of CD14 polymorphism on CD14 expression in patients with extensive burns

We sought to investigate the association of CD14 genotype with the risk of mortality after burn, and we also attempted to evaluate whether CD14-159 C/T polymorphism affects the kinetics and extent of CD14 expression as well as its release, and TNF-alpha expression in burned patients. The study involved 64 patients in Chinese Han population incurring burns covering more than 30% of the total body surface area. CD14 polymorphism was determined by polymerase chain reaction (PCR) and subsequent restriction fragment length polymorphism (RFLP) analysis. Meanwhile, leukocyte CD14 mRNA expression and soluble CD14 (sCD14) levels were measured during a 28-day observation period. TNF-alpha mRNA and protein levels were also determined in patients with different genotypes of CD14. On day 21 after burn, CD14 mRNA expression and sCD14 levels were significantly higher in TT homozygotes than in CC genotypes (1.33+/-0.36 microg/ml vs. 0.75+/-0.28 microg/ml and 16.1+/-4.6 microg/ml vs. 9.7+/-3.4 microg/ml, P<0.05), and these values were also higher in non-survivors than in survivors (1.32+/-0.40 microg/ml vs. 0.87+/-0.32 microg/ml and 14.8+/-4.5 microg/ml vs. 11.1+/-4.8 microg/ml, P<0.05). In addition, TNF-alpha mRNA and protein levels were significantly lower in both CC homozygotes and survivors than in TT genotypes or non-survivors during the 28-day observation period (P<0.05). However, TT genotype did not impart an increased risk for burn mortality in this small study. In conclusion, CD14-159 C/T polymorphism might be associated with the kinetics and extent of CD14 expression as well as its release, and it was also related to TNF-alpha expression. However, this study did not confirm CD14-159 C/T polymorphism was associated with the outcome of extensive burns.     

Association between decreased kidney function and endotoxin receptor CD14 C-159T polymorphism among Japanese health check-up examinees

BACKGROUND: A recently identified promoter polymorphism of the endotoxin receptor (CD14 C-159T) was shown to be associated with atherosclerotic diseases such as myocardial infarction. This study was conducted to determine whether this polymorphism is associated with decreased kidney function. METHODS: A total of 281 male and 522 female health check-up examinees, aged 39-88 years, were genotyped for CD14 C-159T. The glomerular filtration rate (GFR) was estimated by the Modification of Diet in Renal Disease (MDRD) Study equation. Estimated GFR (eGFR) and the proportion of subjects with mildly decreased eGFR (eGFR under 90 mL/min/1.73 m(2)) were compared among the genotypes. RESULTS: Subjects carrying the T allele showed decreased age- and sex-adjusted eGFR compared with those with CC genotype (101+/-22 vs. 105+/-23 mL/min/1.73 m(2); mean+/-SD, p = 0.012). The proportion of subjects with mildly decreased eGFR was higher in T allele carriers (34.2% for TT+CT and 26.3% for CC genotype, p = 0.041), but not statistically significant when adjusted for age and sex (odds ratio [OR] 1.41, 95% CI 0.97-2.05, p = 0.076). In subjects under 65 years, T allele carriers had a significantly increased risk for mildly decreased eGFR (27.1% for TT+CT and 18.0% for CC; age- and sex-adjusted OR 1.82, 95% CI 1.06-3.12, p = 0.030). CONCLUSION: CD14-159T allele was associated with decreased eGFR compared with CC genotype, and with a higher prevalence of mildly decreased eGFR in younger subjects under 65.     

Association between Decreased Kidney Function and Endotoxin Receptor CD14 C-159T Polymorphism among Japanese Health Check-up Examinees

Background. A recently identified promoter polymorphism of the endotoxin receptor (CD14 C-159T) was shown to be associated with atherosclerotic diseases such as myocardial infarction. This study was conducted to determine whether this polymorphism is associated with decreased kidney function. Methods. A total of 281 male and 522 female health check-up examinees, aged 39–88 years, were genotyped for CD14 C-159T. The glomerular filtration rate (GFR) was estimated by the Modification of Diet in Renal Disease (MDRD) Study equation. Estimated GFR (eGFR) and the proportion of subjects with mildly decreased eGFR (eGFR under 90 mL/min/1.73 m²) were compared among the genotypes. Results. Subjects carrying the T allele showed decreased age- and sex-adjusted eGFR compared with those with CC genotype (101±22 vs. 105±23 mL/min/1.73 m²; mean±SD, p?=?0.012). The proportion of subjects with mildly decreased eGFR was higher in T allele carriers (34.2% for TT+CT and 26.3% for CC genotype, p?=?0.041), but not statistically significant when adjusted for age and sex (odds ratio [OR] 1.41, 95% CI 0.97–2.05, p?=?0.076). In subjects under 65 years, T allele carriers had a significantly increased risk for mildly decreased eGFR (27.1% for TT+CT and 18.0% for CC; age- and sex-adjusted OR 1.82, 95% CI 1.06–3.12, p?=?0.030). Conclusion. CD14-159T allele was associated with decreased eGFR compared with CC genotype, and with a higher prevalence of mildly decreased eGFR in younger subjects under 65.     

Polymorphisms in the B-type natriuretic peptide (BNP) gene are associated with NT-proBNP levels but not with diabetic nephropathy or mortality in type 1 diabetic patients.

BACKGROUND: Circulating N-terminal pro-brain natriuretic peptide (NT-proBNP) levels are elevated in patients with diabetic nephropathy and independently predict excess cardiovascular morbidity and mortality. Therefore, we investigated the association between two polymorphisms -381T/C and 1551G/A of the BNP gene, plasma NT-proBNP levels and mortality prognosis in 380 type 1 diabetic patients with and without diabetic nephropathy. METHODS: In a prospective observational follow-up study, 197 type 1 diabetic patients with diabetic nephropathy {121 men, age [mean (SD)] 41 +/- 9.5 years, duration of diabetes 28 +/- 8.0 years, glomerular filtration rate 67 +/- 28 ml/min/1.73 m2}, and a matched control group of 183 patients with longstanding type 1 diabetes and persistent normoalbuminuria (111 men, age 43 +/- 10.0 years, duration of diabetes 27 +/- 8.3 years) were followed for 12.6 (0.0-12.9) years. Plasma NT-proBNP concentration was determined by immunoassay at baseline. The BNP genotypes were determined by TaqMan chemistry based assays. RESULTS: The two polymorphisms were in almost complete linkage disequilibrium (r2 = 0.883) and thus only the results of the -381T/C promoter polymorphism are shown. There was no significant difference between cases and controls in either genotype distributions (cases TT 32%, TC 53%, CC 15%; controls TT 28%, TC 52%, CC 20%) or allele frequencies (cases T/C 0.58/0.42; controls T/C 0.54/0.46) for the -381T/C polymorphism. Among the 164 normoalbuminuric patients without antihypertensive treatment and previous major cardiovascular disease (CVD), the -381T/C polymorphism was associated with circulating levels of NT-proBNP [median (interquartile range) 21 (5-32), 34 (12-67) and 32 (12-58) ng/l for TT, TC and CC, respectively (P = 0.041)] persisting after adjustment for covariates (P = 0.018). During follow-up, the -381T/C polymorphism did not predict all-cause or cardiovascular mortality among type 1 diabetic patients with or without diabetic nephropathy. CONCLUSIONS: The BNP -381T/C and 1551G/A polymorphisms are associated with circulating levels of NT-proBNP but not with prevalent overt diabetic nephropathy. These polymorphisms do not predict all-cause or cardiovascular mortality in Caucasian type 1 diabetic patients with or without diabetic nephropathy.