M2型巨噬细胞对草酸钙晶体刺激肾小管上皮细胞损伤的影响  

The effects of M2-type macrophages on renal tubular epithelial cells injury exposed to calcium oxalate crystal

作　　者：刘权[1] 关晓峰[1] 陶芝伟[1] 王翔[1] 吴基华[1] 刘云龙[1] 何子奇[1] 邓耀良[1] 

机构地区：[1]广西医科大学第一附属医院泌尿外科,广西南宁530021

出　　处：《广东医学》2018年第3期331-335,共5页Guangdong Medical Journal

基　　金：国家自然科学基金资助项目(编号:81360113;81760127);广西卫计委自筹经费科研课题(编号:Z2016311)

摘　　要：目的探讨M2型巨噬细胞对草酸钙晶体(CaOx)刺激肾小管上皮细胞(HK-2)损伤的影响。方法将人单核细胞(THP-1)经佛波酯(PMA)及白细胞介素(IL)-4、IL-13序贯诱导为M2型巨噬细胞,使用实时定量PCR、Western Blot、ELISA法检测其表型标志物,以鉴定诱导分化情况。用浓度为0.5 mg/m L的CaOx刺激HK-2细胞,利用Transwell建立M2型巨噬细胞(上室)与HK-2细胞(下室)共培养模型,实验分组为HK-2细胞组、HK-2细胞+M2型巨噬细胞共培养组、HK-2细胞+CaOx组、HK-2细胞+CaOx+M2型巨噬细胞共培养组。使用CCK-8法检测各组HK-2细胞活力、微量酶标仪法测各组HK-2细胞上清液的乳酸脱氢酶(LDH)活性、DAPI进行凋亡染色和Western Blot检测各组HK-2细胞抗凋亡蛋白Bcl-2的表达情况。划痕实验观测M2型巨噬细胞对HK-2细胞的迁移、修复的影响。结果表型标志物鉴定结果显示,成功将THP-1细胞序贯诱导为M2型巨噬细胞。CCK-8法结果显示M2型巨噬细胞显著增加CaOx对HK-2细胞损伤后的活力,且M2型巨噬细胞对HK-2细胞增殖具有促进作用(P<0.05)。LDH活性检测结果显示M2型巨噬细胞显著性降低CaOx刺激HK-2细胞的LDH活性(P<0.05)。DAPI染色及抗凋亡蛋白Bcl-2的表达结果显示M2型巨噬细胞具有抑制CaOx刺激HK-2细胞凋亡的作用。划痕实验显示M2型巨噬细胞对HK-2细胞具有促进迁移、修复的作用。结论 M2型巨噬细胞对CaOx刺激HK-2细胞的损伤具有一定的抑制作用,且可促进HK-2细胞增殖、迁移。Objective To investigate the effects of M2 - type macrophages on renal tubular epithelial cells （ HK - 2） injury exposed to calcium oxalate （CaOx） crystal. Methods THP- 1 cells were induced into M2 -type macrophages by Phorbol - 12 - myristate - 13 - acetate （ PMA）, IL -4 and IL - 13. Their phenotypie markers were identified by RT - PCR, Western Blot and ELISA. HK- 2 cells were incubated with CaOx at a concentration of 0. 5 mg/mL. The co- cul- tured model of M2 -type maerophages （upper chamber） and HK- 2 cells （lower chamber） were constructed by Trans - well. The experiment was divided into HK - 2 cells group, HK - 2 cells ＋ M2 - type macrophages co - cultured group, HK- 2 cells ＋ CaOx group, and HK- 2 cells ＋ CaOx ＋ M2 -type macrophages co- cultured group. The viability of HK - 2 cells was detected by CCK - 8. The activity of lactate dehydrogenase （LDH） in the supernatant of HK - 2 cells was detected by microplate reader. DAPI was used for the apoptosis staining and Western blot was used for detecting the expression of Bcl - 2 in HK - 2 cells. The effects of M2 macrophages on the migration and repairmen of HK - 2 ceils were observed by the scarification test. Results THP - 1 cells were successfully induced into M2 - type macrophages. The CCK- 8 test showed that M2 -type macrophages significantly increased the viability of CaOx on HK -2 cells and M2 - type macrophages proliferated on HK - 2 cells （ P 〈 0.05 ）. The LDH test showed that M2 macrophages significantly re- duced the LDH of HK - 2 cells exposed to CaOx （ P 〈 0. 05 ）. The DAPI test and the Bcl - 2 expression showed that M2macrophages inhibited the HK- 2 cells from apoptosis exposed to CaOx. The scarification experiment showed that M2 - type macrophages had a role of promoting the migration and repairmen to HK - 2 cells. Conclusion M2 - type macropha- ges have a certain inhibitory effect on HK-2 cells injury exposed to CaOx and can also promote the proliferation, migra- tion and repairmen of HK -2 cells.

关 键 词：M2型巨噬细胞 草酸钙晶体 肾小管上皮细胞 损伤 

分 类 号：R692.4[医药卫生—泌尿科学]