中国实验用小型猪肾小管的发育及各节段标志物的表达

Renal Tubular Development and Different Segments Renal Tubular Markers of Chinese Experimental Miniature pig

目的:明确解剖和生理上与人非常接近的中国实验用小型猪肾脏发育过程中肾小管的形态学变化和肾小管各节段的特异性标志物。方法:采用高碘酸-希夫(PAS)染色和免疫荧光染色技术,系统观察中国实验用小型猪妊娠28～112d(E28d～E112d)和出生后1d、7d、14d、21d(P1d～21d),共17个不同时间点猪肾小管的发育及肾小管特异性标志物雪莲花凝集素(LTL)、水通道蛋白1(AQP1)、钙结合蛋白(calbindin)-D28k在肾小管不同节段的表达。结果:(1)中国实验用小型猪E28d可见后肾间充质和输尿管芽,即后肾已经开始发育;但这时还没有肾小管。E35d可见不同节段的肾小管,即肾小管已开始发育。从E35d～P14d(E112d仔猪出生),肾皮质均有生肾区存在,即不断有新的肾单位发生;P21d生肾区消失,即不再有新的肾单位产生。(2)①LTL在E28d表达在输尿管芽,E35d开始在近端小管表达,以刷状缘表达最为明显;表达由弱到强,由点状到线状。②AQP1在E28d未见表达,E35d开始表达;AQP1表达在近端小管和髓袢的降支细段,主要表达在细胞膜,尤其在管腔侧的表达更为明显。③Calbindin-D28k在E28d表达在输尿管芽,E35d开始表达在远端小管和集合管;Calbindin-D28k主要表达在细胞质,随着肾小管发育,表达逐渐增强。(3)发现集合管来源于输尿管芽,发源于输尿管芽的集合管从被膜下的生肾区一直延伸到肾髓质。结论:中国实验用小型猪妊娠35d可以见到不同节段肾小管。LTL、AQP1、Calbindin-D28k可以分别作为猪近端小管、髓袢、远端小管和集合管的标志物。

Objective：To clear the morphological changes of the renal tubular and its markers during the kidney development of the Chinese experimental miniature pig,whose kidney is very close to human＇s in anatomy and physiology.Methods：The PAS staining and immunofluorescence staining were used to systematically study the development of renal tubular as well as the expression of the LTL,AQP1 and Calbindin-D28k in different segments of renal tubules at 17 different time points that is embryonic 28 days to 112 days（E28 d^E112 d） and postnatal 1 day,7 days,14 days,21 days（P1 d^E21 d） in the Chinese experimental miniature pig.Results：（1）In E28 d,the metanephric mesenchyme and ureteric bud were found that means the start of permanent kidney development while the renal tubular wasn＇t observed.In E35 d,the different segments of renal tubules could be found that means the start of the development of renal tubules.From E35 d to P14 d,the nephrogenic zone could be found that was contributing to the information of nephron,while it disappeared in P21d（Piglets born in E112d）.（2）① LTL was expressed in ureteric bud in E28 d and extended to proximal tubules in E35 d.LTL was mainly expressed in brush border,and displayed from weak to strong,from point to line.② AQP1 was expressed in the proximal tubule and the descending thin limb of Henle＇s loop,and was mainly in cell membrane especially the luminal side from E35 d.③ Calbindin-D28k was expressed in ureteric bud in E35 d,and extended to the distal tubule and collecting duct in E35 d.Calbindin-D28k was mainly expressed in the cytoplasm,and gradually enhanced with the development of renal tubular.（3）In this study,we found that collecting duct originated from the ureteric bud and extended from subcapsular nephrogenic zone to the renal medulla.Conclusion：The different segments renal tubular of Chinese experimental miniature pig could be observed as early as E35 d.LTL,AQP1 and Calbindin-D28k could be used as markers of proximal tubular,medullary loop,distal tubule and collecting duct respectively in this miniature pig.The collecting duct originated from the ureteric bud.