摘要
目的从健康志愿者新鲜粪便中筛选人源产甲酸草酸杆菌并研究其防治草酸钙结石的作用。方法从健康志愿者新鲜粪便中筛选并分离培养人源产甲酸草酸杆菌。采用0.8%乙二醇灌胃建立草酸钙结石大鼠模型。大鼠按随机数字表法分为对照组及4组乙二醇诱导的草酸钙结石模型组,其中3组草酸钙结石大鼠分别以10^6、10^7、10^8菌落形成单位(CFU)活菌干预4周。每周收集大鼠血及24 h尿液检测血钙、血镁、血磷、BUN、Scr、尿草酸、尿钙、尿镁、尿磷的变化。于第4周末处死大鼠,肾组织行HE、Yasue染色,偏折光镜下观察肾脏草酸钙结晶沉积部位及含量。结果分离出的菌株与已知美国模式菌种收集中心(ATCC)35274相似度为100%,由此判定已成功筛选出产甲酸草酸杆菌。第4周末5组大鼠间体重、Scr、BUN、血钙、血镁、血磷、尿镁、尿磷的差异均无统计学意义。第4周末造模组24 h尿钙排泄量显著低于对照组(P<0.05),经产甲酸草酸杆菌菌液干预后,10^8 CFU组24 h尿钙排泄量升高,显著高于造模组(P<0.05),其余干预组24 h尿钙排泄量与造模组差异无统计学意义。10^6 CFU组、10^7 CFU组尿草酸排泄量较造模组有所下降,但差异未达到统计学意义;10^8 CFU干预组在第1周末24 h尿草酸排泄量即显著低于造模组(P<0.05),且持续至第4周末。干预4周后,偏折光镜下对照组无结晶形成,造模组可见肾皮质和肾髓质内大量草酸钙结晶形成,结晶成堆分布,相互连接;Yasue染色与肾脏同一部位草酸钙结晶重合;与造模组相比,10^6 CFU组、10^7 CFU组草酸钙结晶评分无明显改变,而10^8 CFU组肾脏草酸钙结晶评分显著降低(P<0.05)。结论本研究已成功筛选出人源产甲酸草酸杆菌。每日给予10^8 CFU产甲酸草酸杆菌可安全、有效降低大鼠尿草酸排泄量,预防肾脏草酸钙结晶形成从而抑制结石形成。
Objective To screen Oxalobacter formigenes (OxF) from fresh feces of healthy adults, and study its effect on the the prevention of calcium oxalate kidney stones. Methods OxF was screened and cultured from fresh feces of healthy adults. The rat model of calcium oxalate stone was established by esophageal gavage of 0.8% of ethylene glycol. Rats were divided into a control group and four groups of rats with ethylene glycol-induced calcium oxalate kidney stones according to random number table. Three groups were treated with 10^6 CFU, 10^7 CFU, 10^8 CFU viable OxF every day, respectively, for 4 weeks. The blood and 24-hour urine samples were collected to detect the serum creatinine, urea nitrogen, serum and urine calcium, phosphorus, magnesium and urine oxalate every week. At the end of the 4th week, the rats were sacrificed and the kidney tissues were stained with HE and Yasue. The deposition and content of calcium oxalate crystals were observed under a light microscope. Results The bacteria strain isolated from fresh feces of healthy adults was 100% as same as the known ATCC35274 bacteria strain, which means the strain screened is OxF. Among the 5 groups, there were no significant differences in body weight, Scr, BUN, serum calcium, blood magnesium, blood phosphorus, urinary magnesium and urinary phosphorus. The 24-hour urinary calcium excretion in the model group was significantly lower than that of the control group (P<0.05). After intervention with OxF solution, the 24-hour urinary calcium excretion in the 10^8 CFU OxF group was significantly higher than that in the model group (P<0.05), while there was no significant difference between the other intervention groups and the model. The oxalic acid excretion of 10^6 CFU OxF group and 10^7 CFU OxF group was lower than that of the model, but the difference did not reach statistical significance (P>0.05). The 24 h oxalic acid excretion in the 10^8 CFU OxF group was significantly lower than that of the model at the end of first week (P<0.05), and continued to decrease for the next 3 weeks. After 4 weeks of intervention, no crystal formation was observed in the control group under the deflection microscope, but a large amount of calcium oxalate crystals were formed in the renal cortex and renal medulla. The crystals were piled up and connected to each other. Yasue staining coincided with the calcium oxalate crystal in the same part of the kidneys. Compared with the model, there was no significant change in the score of calcium oxalate crystal in the kidneys of 10^6 CFU OxF group and 10^7 CFU OxF group, while the score of calcium oxalate crystal in the kidneys of 10^8 CFU OxF group was significantly lower (P<0.05). Conclusions OxF are successively screened from healthy adults. Daily administration of 10^8 CFU OxF can safely and effectively reduce the urinary oxalic acid excretion, prevent the formation of calcium oxalate crystals and inhibit the formation of stones in kidneys of rats.
作者
张帝
汤晓静
高远
于鸿晶
文彬
付莉莉
宋书伟
梅长林
Zhang Di;Tang Xiaojing;Gao Yuan;Yu Hongjing;Wen Bin;Fu Lili;Song Shuwei;Mei Changlin(Kidney Institute, Department of Nephrology, Shanghai Changzheng Hospital, Second Military University,Shanghai 200003, China;SPH Sine Pharmaceutical Laboratories Co., Ltd,., Shanghai EngineeringResearch Center of Innovative Probiotic Drugs, Shanghai 201206, China)
出处
《中华肾脏病杂志》
CAS
CSCD
北大核心
2019年第4期288-294,共7页
Chinese Journal of Nephrology
基金
国家自然科学基金(81500533、81670612)
国家重点研发计划(2016YFC0901502)
上海市重中之重临床重点学科(2017ZZ02009)
上海市卫生和计划生育委员会科研课题(20164Y0159).
关键词
食草酸杆菌
肾结石
草酸钙
Oxalobacter formigenes
Kidney calculi
Calcium oxalate
