CAJ | 학술논문

背景：研究发现在肾结石模型中肾间质晶体周围存在大量单核/巨噬细胞浸润，显示巨噬细胞可能参与晶体在肾脏中的沉积过程，而巨噬细胞是人体重要的固有免疫细胞，在肾脏中沉积的大量的单核/巨噬细胞吞噬晶体，会产生一些炎症因子损伤和破坏肾小管上皮细胞，最终有利于结石的形成。目的：探讨一水草酸钙晶体刺激人巨噬细胞后高迁移率族蛋白B1的表达水平。方法：用100 mg/L的一水草酸钙刺激巨噬细胞，分别于刺激后0，6，12，24和36 h，用Western blot检测细胞总蛋白和细胞浆内高迁移率族蛋白B1的含量；用实时荧光定量PCR，检测细胞中高迁移率族蛋白B1 mRNA表达情况；分别于一水草酸钙刺激后0，1，2和4 h，用酶联免疫吸附实验测定细胞培养液上清中肿瘤坏死因子α和白细胞介素6的水平。结果与结论：一水草酸钙刺激后0-6 h细胞浆内高迁移率族蛋白B1的含量较低，刺激后12-36 h细胞浆内高迁移率族蛋白B1逐渐增加。一水草酸钙刺激后0-6 h，巨噬细胞总蛋白中高迁移率族蛋白B1含量不高，在刺激后12 h细胞总蛋白高迁移率族蛋白B1的含量开始增加，并且在刺激后24-36 h保持在较高水平。RT-PCR结果显示，一水草酸钙刺激后0-12 h，培养细胞中高迁移率族蛋白B1的mRNA表达量无明显变化，刺激后18-24 h培养细胞中高迁移率族蛋白B1的mRNA表达量明显增加。ELISA结果显示，一水草酸钙刺激后2 h，细胞培养液上清中肿瘤坏死因子α和白细胞介素6表达和释放增加，4 h达到明显高峰。结果表明，一水草酸钙可以诱导人巨噬细胞高迁移率族蛋白B1的表达及mRNA表达增加；诱导人巨噬细胞内肿瘤坏死因子α和白细胞介素6的表达增加，且高迁移率族蛋白B1表达的时间明显晚于肿瘤坏死因子a和白细胞介素6的释放时间。
배경：연구발현재신결석모형중신간질정체주위존재대량단핵/거서세포침윤，현시거서세포가능삼여정체재신장중적침적과정，이거서세포시인체중요적고유면역세포，재신장중침적적대량적단핵/거서세포탄서정체，회산생일사염증인자손상화파배신소관상피세포，최종유리우결석적형성。목적：탐토일수초산개정체자격인거서세포후고천이솔족단백B1적표체수평。방법：용100 mg/L적일수초산개자격거서세포，분별우자격후0，6，12，24화36 h，용Western blot검측세포총단백화세포장내고천이솔족단백B1적함량；용실시형광정량PCR，검측세포중고천이솔족단백B1 mRNA표체정황；분별우일수초산개자격후0，1，2화4 h，용매련면역흡부실험측정세포배양액상청중종류배사인자α화백세포개소6적수평。결과여결론：일수초산개자격후0-6 h세포장내고천이솔족단백B1적함량교저，자격후12-36 h세포장내고천이솔족단백B1축점증가。일수초산개자격후0-6 h，거서세포총단백중고천이솔족단백B1함량불고，재자격후12 h세포총단백고천이솔족단백B1적함량개시증가，병차재자격후24-36 h보지재교고수평。RT-PCR결과현시，일수초산개자격후0-12 h，배양세포중고천이솔족단백B1적mRNA표체량무명현변화，자격후18-24 h배양세포중고천이솔족단백B1적mRNA표체량명현증가。ELISA결과현시，일수초산개자격후2 h，세포배양액상청중종류배사인자α화백세포개소6표체화석방증가，4 h체도명현고봉。결과표명，일수초산개가이유도인거서세포고천이솔족단백B1적표체급mRNA표체증가；유도인거서세포내종류배사인자α화백세포개소6적표체증가，차고천이솔족단백B1표체적시간명현만우종류배사인자a화백세포개소6적석방시간。
BACKGROUND:There are a large amount of infiltrated monocytes and macrophages around the renal interstitial crystals in a kidney model, indicating that macrophages may be involved in the process of crystal deposition in the kidney. As macrophages are important human innate immune cels, a large number of infiltrated monocytes and macrophages in the kidney wil produce some inflammatory injuries and damage renal tubular epithelial cels, which ultimately lead to the formation of stones. OBJECTIVE: To study the expression of after high mobility group box-1 protein (HMGB1) in macrophages stimulated by calcium oxalate monohydrate crystals. METHODS:Western blot assay was used to detect total protein and HMGB1 levels in the cytoplasm of macrophages under 100 mg/L calcium oxalate monohydrate stimulation for 0, 6, 12, 24, 36 hours. Meanwhile, <br> real-time quantitative PCR was adopted to determine the mRNA expression of HMGB1. Tumor necrosis factor α and interleukin-6 levels in the cultured cel supernatant at 0, 1, 2 and 4 hours after calcium oxalate monohydrate stimulation were measured using ELISA. RESULTS AND CONCLUSION: Under the calcium oxalate monohydrate stimulation, the level of HMGB1 in the cytoplasm of macrophages was lower within 0-6 hours, but gradualy increased within 12-36 hours; the level of HMGB1 in the total protein of macrophages was not high within 0-6 hours, began to increase at 12 hours, and then kept at a higher level within 24-36 hours. RT-PCR results showed that under the calcium oxalate monohydrate stimulation, the mRNA level of HMGB1 in the cultured cels had no changes within 0-12 hours, but significantly increased within 18-24 hours. ELISA results showed that thelevels of tumor necrosis factor α and interleukin-6 in the cultured cel supernatant subject to calcium oxalate monohydrate were increased at 2 hours and peaked at 4 hours. These findings indicate that calcium oxalate monohydrate can increase the HMGB1 expression in macrophages at mRNA and protein levels, and also induce the increase in tumor necrosis factor α and interleukin-6 expression in macrophages. Moreover, the expression of HMGB1 is later than the release of tumor necrosis factor α and interleukin-6.