PTPN22在大鼠脑出血后炎症损伤中的作用研究

Interference of protein tyrosine phosphatase non-receptor type 22 inhibits NLRP3 inflammasome activation to alleviate inflammation after cerebral hemorrhage in rats

目的探讨蛋白酪氨酸磷酸酶,非受体型22(protein tyrosine phosphatase non-receptor type 22,PTPN22)在大鼠脑出血后炎症反应中的作用及其机制。方法采用自体血注入脑内建立大鼠脑出血(intracerebral hemorrhage,ICH)模型,36只SD大鼠随机分为假手术组,ICH3、6、12、24、48 h组(n=6),Western blot筛选大鼠脑出血后PTPN22表达的时间窗。72只大鼠随机分为假手术组、ICH组、ICH+NC组、ICH+PTPN22干扰组(n=18)。ICH24 h后,对各组大鼠进行神经功能评分(mNSS),脑含水量测定,Western blot检测大鼠脑出血后血肿周围NLRP3,成熟的白介素-1β(cleaved-IL-1β),成熟的白介素-18(cleaved-IL-18)以及激活的半胱天冬酶-1(cleaved-caspase-1)的表达,HE和Nissl染色观察大鼠脑组织形态学改变,免疫荧光染色检测血肿周围髓过氧化物酶(MPO)的表达。结果大鼠脑出血后12hPTPN22的表达开始升高,24 h达到峰值,48 h开始降低。差异具有统计学意义(P<0.05);与ICH+NC组相比,ICH+PTPN22干扰组神经功能评分降低,脑含水量减少,差异具有统计学意义(P<0.05);HE和Nissl染色结果可见,ICH+PTPN22干扰组脑组织损伤程度与ICH+NC组相比明显减轻;ICH+PTPN22干扰组大鼠脑出血后血肿周围NLRP3、cleaved-IL-1β、cleaved-IL-18以及cleaved-caspase-1的表达与ICH+NC组相比明显降低(P<0.05);ICH+PTPN22干扰组MPO细胞与ICH+NC组相比明显减少。结论干扰PTPN22可通过抑制NLRP3炎性体的激活从而减轻大鼠脑出血后的炎症损伤。

Objective To investigate the role of protein tyrosine phosphatase non-receptor type 22(PTPN22)in inflammatory response following intracerebral hemorrhage(ICH)in rats and explore the mechanism that mediates its effect.Methods Rat models of ICH were established by injecting autologous blood into the brain of 30 SD rats,and Western blotting was performed at 3,6,12,24,and 48 h after ICH(6 rats at each time point)and in 6 sham-operated rats to determine the time window of PTPN22 expression after ICH.Another 72 rats were randomized equally into sham operation group,ICH group,ICH+negative control(ICH+NC)group,and ICH+PTPN22 interference group,and the neurological function score(mNSS)and brain water content were assessed at 24 h after ICH or the sham operation.Western blotting was used to detect the expression of cleaved interleukin-18(cleaved IL-18),cleaved-IL-1βand cleaved caspase-1 around the hematoma in the rats.Histomorphological changes of the brain tissues of the rats were observed using HE and Nissl staining,and the expression of myeloperoxidase(MPO)around the hematoma was detected using immunofluorescence staining.Results The expression of PTPN22 began to increase significantly at 12 h after ICH in the rats,peaked at 24 h and began to decrease at 48 h(P<0.05).Compared with those in ICH+NC group,the rats in ICH+PTPN22 interference group had significantly lowered neurological scores and reduced brain water content(P<0.05)with obviously alleviated brain tissue damage as shown by HE and Nissl staining.The expression of NLRP3,cleaved IL-1β,cleaved IL-18 and cleaved caspase-1 and the number of MPO-positive cells around the hematoma were all significantly lowered in PTPN22 interference group compared with ICH+NC group(P<0.05).Conclusion Small interfering RNA-mediated interference of PTPN22 attenuates inflammatory response after ICH in rats by inhibiting the activation of NLRP3.