摘要
目的探讨谷氨酰胺预处理对缺氧/复氧损伤后HK-2细胞凋亡的影响。方法取离体培养的HK-2细胞,随机分成4组:对照组(Con组)。CoCl2组:加入300μM CoCl2处理4 h,然后更换正常的培养基培养24 h,之后更换无血清的培养基培养。谷氨酰胺组(GLN组):培养孔中加入10 mmol/L谷氨酰胺预处理24 h后同CoCl2组。ZnPP组:培养孔中加入3μM ZnPP(锌原卟啉,HO-1抑制剂)和10 mmol/L谷氨酰胺处理24 h后同CoCl2组。MTT法测定细胞增殖,流式细胞技术测定细胞的凋亡。RT-PCR技术检测血红素加氧酶-1(Heme oxygenase,HO-1)和凋亡相关基因的表达情况。结果 10 mmol/L谷氨酰胺预处理可以明显增加HK-2细胞的增殖能力,减少凋亡(P<0.01)。预处理上调HO-1的表达,同时上调凋亡抑制基因Bcl-2的表达,下调促凋亡基因caspase-3和NF-κB的表达。而这些调节作用可被ZnPP抑制(P<0.05或P<0.01)。结论 10 mmol/L谷氨酰胺预处理对缺氧/复氧后HK-2细胞有保护作用,HO-1和凋亡相关基因在预处理中起到重要的作用。
Objective To determine whether glutamine (GLN) pretreatment may protect human proximal re- nal tubular epithelial ( HK-2 ) cells against anoxia-reoxygenation injury and the role of Heme oxygenasel ( HO-1 ). Methods Cells were randomly assigned to four groups : control group ( Con ) : in which cells were untreated ; CoC12 group( CoC12 ) :in which cells were treated with 300 p.M CoC12 (chemical anoxia analogue) for 1 hour, cultured with normal medium for 24 hours, and then stimulated with no serum media ; GLN group : in which cells were pretreated with 10 mmol/L GLN for 24 hours, then treated by the same method as group CoC12 ;ZnPP group (inhibitor of HO-1 ) :cells were treated with 3 p^M ZnPP and 10 mmol/L GLN for 24 hours, then treated by the same method as group CoC12. U- sing MTT method and flow cytometry to detect the proliferation and apoptosis of HK-2 cells, RT-PCR method was used to investigate the regulation of HO-1, NF-KB, Bcl-2 and caspase-3 mRNA. Results After pretreated with 10 mmol/L GLN, the HK-2 cells activity was increased and the apoptosis was decreased (P 〈 0. 01 ) , the expression of HO-1 and Bcl-2 gene were up-regulated, NF-KB and caspase-3 gene were down-regulated ( P 〈 0.05 or 〈 0. 01 ). Condnsion Pretreatment with 10 mmol/L GLN effectively prevents the HK-2 cells against anoxia-reoxygenation injury by regula- ting the expression of apoptosis related genes ,HO-1 induced by GLN plays a key role in this cytoprotection.
出处
《实用药物与临床》
CAS
2013年第11期993-997,共5页
Practical Pharmacy and Clinical Remedies
基金
辽宁省科技厅科学技术计划项目(2011225020)