NGAL在ADV早期肾损伤中的表达及临床应用价值的初步研究

发布时间：2018-05-20 23:09

本文选题：中性粒细胞明胶酶相关脂质运载蛋白 + 慢性乙型肝炎　；参考：《第二军医大学》2017年硕士论文

【摘要】：目的:长期口服阿德福韦酯(ADV)治疗慢性乙型病毒性肝炎最常见的不良反应是肾功能损伤,目前临床上常用SCr、BUN、eGFR、血磷等作为ADV肾损伤的诊断指标,但这些传统指标对诊断早期肾损伤敏感性和特异性较低。已有大量研究证明中性粒细胞明胶酶相关脂质运载蛋白(NGAL)对各种病因引起的急性或慢性肾损伤均有很好的诊断价值。本研究通过横断面研究检测ADV单药或联用其它核苷(酸)类似物(NA)的慢性乙型病毒性肝炎(CHB)患者的血清及尿NGAL浓度水平,与SCr、eGFR、CysC、血磷等比较在ADV早期肾损伤的临床诊断价值。同时通过动物实验,研究NGAL在长期灌服ADV等核苷(酸)类似物大鼠肾脏损伤模型中的表达水平及部位。方法:本研究共有两部分:第一部分,临床横断面研究。选取2016年2月到2016年11月在长海医院感染科门诊长期口服ADV单药或ADV联合LAM、ADV联合ETV的CHB患者124例。跟据患者的eGFR水平分为正常肾功能组(A组:eGFR≥90ml/min)91例,肾功能损伤组(B组:eGFR90ml/min)33例,选取同期在门诊随访的55例HBV稳定携带者,63例口服ETV患者作为对照组。整理患者门诊检查的eGFR、SCr、Cys C、血磷等肾脏传统指标。通过酶联免疫吸附试验获得各组CHB患者的血清和尿NGAL浓度。统计分析比较NGAL与其他传统肾损伤标记物对诊断ADV引起的早期肾损伤的敏感性,Pearson相关分析进一步了解血清和尿NGAL与年龄、性别、SCr、eGFR,CysC、β2-MG、尿微量白蛋白、血磷之间的相关性。年龄分层下比较ADV肾功能损伤组NGAL浓度差异。第二部分,研究NGAL在ADV、TDF、ETV诱导的大鼠肾脏中的表达及部位。通过长期给予SD大鼠ADV、TDF、ETV,建立相应药物诱导的肾损伤模型,分别于8周、12周、16周检测并分析各组大鼠血清和尿NGAL的水平,通过HE染色了解各组大鼠肾脏的病理变化,免疫组化法检测肾脏组织中NGAL表达程度及部位。采用SPSS 22.0软件处理数据,P0.05代表差异有统计学意义。结果:1.血清NGAL虽然在ADV肾损伤组高于其它各组,但无统计学意义(P0.05)。ADV肾损伤组尿NGAL明显高于ETV组和稳定携带组,差异有统计学意义,但尿NGAL升高并没有血清肌酐和胱抑素C明显。2.相关性分析发现,血清和尿NGAL成正相关,两者与年龄成正相关(P0.05),与性别、服药时间、TB、eGFR、肌酐、胱抑素C、血钙、血磷、β2微球蛋白无相关性,此外尿NGAL与尿微量白蛋白成正相关(r=0.435,P0.05)。根据年龄分层结果显示,年龄越高,血清和尿NGAL越高,且50-59岁组、大于59岁组与小于50岁组比较有统计学差异(P0.01)。3.动物试验中,血清和尿NGAL随着用药时间延长逐渐升高,在ADV组,16周血NGAL明显高于12周,差异有统计学意义(P0.05),16周尿NGAL高于8周和12周,差异均有统计学意义;在TDF组中,16周血和尿NGAL明显高于8周(P0.05);ETV组和对照组在8周、12周、16周均无统计学差异(P0.05)。各组在同一时间点分析发现,ADV组血清和尿NGAL水平在8周、16周明显高于空白对照组和ETV组;TDF组血清和尿NGAL仅在16周与对照组比较有差异;ETV组血清和尿NGAL在8周、12周、16周与对照组均无明显差异。HE染色发现,在第8周,ADV组肾小管上皮细胞出现肿胀,12周、16周肾小管上皮细胞肿胀加重,可见空泡变性,肾小管细胞刷状缘脱落,部分管腔内可见蛋白管型。TDF组在16周肾小管上皮细胞有轻度颗粒变性,未见管型;所有各组肾脏间质和肾小球无明显变化。免疫组化显示NGAL主要表达于肾小管上皮细胞,肾间质和肾小球有少量表达。ADV组在12周、16周NGAL表达高于对照组,TDF组在16周NGAL免疫组化结果与对照组有统计学差异,且表达强度与病理损伤时间有一定的相关性,ETV组和与对照组无明显差异(P0.05)。结论:1.ADV肾损伤组患者尿NGAL明显高于对照组,但尿NGAL诊断ADV早期肾损伤敏感性没有肌酐、胱抑素C高,血清NGAL对ADV早期肾损伤无诊断价值。2.血清NGAL与年龄成正相关,尿NGAL与年龄、尿微量白蛋白成正相关。NGAL对ADV引起的高龄CHB患者肾损伤有更好的诊断价值。3.在大鼠实验组,随着ADV、TDF用药时间的增加,血清和尿NGAL水平明显升高,尿NGAL比血清NGAL升高更明显。肾组织中NGAL主要表达于肾小管上皮细胞,在ADV组表达最强,且表达程度与病理损伤时间有一定的相关性。
[Abstract]:Objective: the most common adverse reaction of long-term oral adfovir (ADV) in the treatment of chronic viral hepatitis is renal impairment. Currently, SCr, BUN, eGFR, and blood phosphorus are commonly used as diagnostic indicators of ADV renal injury, but these traditional indexes are less sensitive and specific for the diagnosis of early renal injury. Cell gelatinase related lipid carrier protein (NGAL) has a good diagnostic value for acute or chronic renal injury caused by various causes. This study examined serum and urinary NGAL levels in patients with chronic hepatitis B (CHB) with ADV single or other nucleoside analogues (NA), and SCr, eGFR, CysC, and blood by cross-sectional study. The clinical diagnostic value of phosphorus and so on in the early renal injury of ADV. At the same time, the expression level and location of NGAL in rat kidney damage model of ADV and other nucleoside (acid) analogues were studied by animal experiments. Methods: This study had two parts: the first part, clinical cross-sectional study, selected from February 2016 to November 2016 in Changhai Hospital. In the outpatient department of infection, 124 cases of CHB with LAM and ADV combined with ETV were taken for a long time. According to the eGFR level of the patients, 91 cases were divided into normal renal function group (A group: eGFR > 90ml/min), 33 cases of renal function injury group (B group: eGFR90ml/min), 55 cases of stable carriers in the same period, and 63 cases of oral administration as the control group. EGFR, SCr, Cys C, blood phosphorus and other renal traditional indicators were examined by an outpatient clinic. The serum and urinary NGAL concentrations of CHB patients in each group were obtained by enzyme linked immunosorbent assay. The sensitivity of NGAL and other traditional renal damage markers to the diagnosis of early renal injury caused by ADV was compared and compared. The Pearson correlation analysis was used to further understand the serum and urinary NGAL. The correlation between age, sex, SCr, eGFR, CysC, beta 2-MG, urine microalbuminuria, and blood phosphorus. The difference of NGAL concentration in ADV renal function injury group was compared under age stratification. The second part was to study the expression and location of NGAL in ADV, TDF, ETV induced rat kidneys. The levels of serum and urine NGAL were detected and analyzed in 8 weeks, 12 weeks and 16 weeks respectively. The pathological changes of kidney in each group were detected by HE staining. The degree and location of NGAL expression in kidney tissues were detected by immunohistochemical method. The data of SPSS 22 were used to deal with the data, P0.05 was statistically significant. The result: 1. serum NGAL was in ADV kidney. The injury group was higher than the other groups, but there was no statistical significance (P0.05) in the.ADV kidney injury group, the urine NGAL was significantly higher than that of the ETV group and the stable carrying group. The difference was statistically significant, but the urine NGAL increased without serum creatinine and cystatin C.2. correlation analysis. The serum and urine NGAL were positively correlated, both were positively correlated with age (P0.05), and sex, and uniform. TB, eGFR, creatinine, creatinine, Cystatin C, blood calcium, blood phosphorus, and beta 2 microglobulin were not related, and urine NGAL was positively correlated with urine microalbumin (r=0.435, P0.05). According to age stratification, the higher the age, the higher the serum and urinary NGAL, and the 50-59 year old group, larger than the 59 year old group and the group less than 50 years of age (P0.01).3. animal test In the group ADV, the serum and urinary NGAL increased gradually with the time of drug use, and the blood NGAL in the 16 week was significantly higher than that in the 12 week. The difference was statistically significant (P0.05). The 16 week urine NGAL was higher than 8 weeks and 12 weeks. In group TDF, the 16 weeks of blood and urine NGAL were significantly higher than 8 weeks (P0.05), and there was no statistical difference between the ETV group and the control group at 8, 12, and 16 weeks. Difference (P0.05). At the same time point analysis, the serum and urinary NGAL levels in group ADV were 8 weeks, 16 Zhou Mingxian was higher than that in the blank control group and ETV group. The serum and urine NGAL in TDF group were only compared with the control group. The serum and urinary NGAL in the ETV group were found to have no significant difference between the control group and the control group at 8 weeks, 12 weeks and the 16 weeks, and in eighth weeks, the ADV group has small kidneys. The tubular epithelial cells were swelling, 12 weeks, 16 weeks of renal tubular epithelial cell swelling increased, visible vacuolation, renal tubular cells brush edge shedding, part of the tubule.TDF group in the 16 weeks in the renal tubular epithelial cells in 16 weeks of mild granular degeneration, no tube type, all groups of renal interstitium and glomeruli have no obvious changes. Immunohistochemistry showed NG AL mainly expressed in renal tubular epithelial cells, renal interstitium and glomeruli had a small amount of expression of.ADV group in 12 weeks, 16 weeks NGAL expression higher than the control group, TDF group in 16 weeks of NGAL immunization results and the control group had statistical difference, and the expression intensity and pathological injury time has certain correlation, ETV group and the control group have no significant difference (P0.05). Conclusion: 1 The urine NGAL of.ADV renal injury group was significantly higher than that of the control group, but the urine NGAL diagnosis of early renal injury was not creatinine, Cystatin C was high, serum NGAL had no diagnostic value for early renal injury,.2. serum NGAL was positively correlated with age, and urine NGAL was positively related to age and urine microalbuminuria. The good diagnostic value.3. was in the experimental group of rats. With the increase of ADV and TDF, the level of serum and urine NGAL increased obviously, and the urine NGAL was more obvious than the serum NGAL. The expression of NGAL in renal tissue was mainly expressed in the renal tubular epithelial cells, and the expression was strongest in the ADV group, and the expression level was related to the time of the disease injury.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R512.62;R692

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