CAJ | 학술논문

目的探讨银屑病患者皮损间充质干细胞(MSC)对T淋巴细胞增殖的抑制能力.方法寻常性银屑病患者15例,进行期7例,静止期8例.健康对照组15例,取自我院泌尿外科和整形外科手术切除的正常皮肤.分离、培养患者皮损与健康人皮肤MSC,流式细胞仪及多向分化法进行细胞鉴定；将第3代皮肤MSC与健康人外周血T细胞共培养,用细胞计数法及MTT法观察其对T细胞增殖的影响；ELISA法检测皮肤MSC培养上清液中白介素6(IL-6)、白介素11(IL-11)、肝细胞生长因子(HGF)和转化生长因子β1(TGF-β1)的浓度.患者组和对照组T淋巴细胞增殖比较采用单因素方差分析及SNK检验,细胞因子水平比较采用两独立样本均数比较的t检验.结果倒置显微镜下,银屑病组与健康对照组皮肤MSC的细胞形态和多向分化能力相似,细胞表面均表达CD29、CD44、CD73、CD90、CD105,而CD34、CD45及HLA-DR表达阴性.健康人外周血T细胞与银屑病患者及健康对照皮肤MSC共培养,T淋巴细胞计数健康对照组为(1.04±0.29)×105/ml,银屑病患者组为(1.67±0.34)×105/ml,两组比较,P＜ 0.01；MTT检测细胞增殖结果(A值)分别为0.275±0.007和0.317±0.021,两组比较,P＜ 0.01.培养上清液中,患者组MSC分泌IL-11水平(181.37±31.74 ng/L高于健康对照组(130.07±29.20 ng/L),两组比较,t=5.32,P＜ 0.01;患者组MSC分泌IL-6(61.67±17.53 ng/L)和HGF(319.24±41.03 ng/L)水平低于健康对照组(分别为76.74±18.96 ng/L和352.35±51.47 ng/L),两组比较,t值分别为2.61和2.25,均P＜ 0.05)；TGF-β1两组差异无统计学意义.结论银屑病患者皮损MSC对T细胞增殖的抑制能力减弱,可能是银屑病的发病机制之一.
목적 탐토은설병환자피손간충질간세포(MSC)대T림파세포증식적억제능력.방법 심상성은설병환자15례,진행기7례,정지기8례.건강대조조15례,취자아원비뇨외과화정형외과수술절제적정상피부.분리、배양환자피손여건강인피부MSC,류식세포의급다향분화법진행세포감정；장제3대피부MSC여건강인외주혈T세포공배양,용세포계수법급MTT법관찰기대T세포증식적영향；ELISA법검측피부MSC배양상청액중백개소6(IL-6)、백개소11(IL-11)、간세포생장인자(HGF)화전화생장인자β1(TGF-β1)적농도.환자조화대조조T림파세포증식비교채용단인소방차분석급SNK검험,세포인자수평비교채용량독립양본균수비교적t검험.결과 도치현미경하,은설병조여건강대조조피부MSC적세포형태화다향분화능력상사,세포표면균표체CD29、CD44、CD73、CD90、CD105,이CD34、CD45급HLA-DR표체음성.건강인외주혈T세포여은설병환자급건강대조피부MSC공배양,T림파세포계수건강대조조위(1.04±0.29)×105/ml,은설병환자조위(1.67±0.34)×105/ml,량조비교,P＜ 0.01；MTT검측세포증식결과(A치)분별위0.275±0.007화0.317±0.021,량조비교,P＜ 0.01.배양상청액중,환자조MSC분비IL-11수평(181.37±31.74 ng/L고우건강대조조(130.07±29.20 ng/L),량조비교,t=5.32,P＜ 0.01;환자조MSC분비IL-6(61.67±17.53 ng/L)화HGF(319.24±41.03 ng/L)수평저우건강대조조(분별위76.74±18.96 ng/L화352.35±51.47 ng/L),량조비교,t치분별위2.61화2.25,균P＜ 0.05)；TGF-β1량조차이무통계학의의.결론 은설병환자피손MSC대T세포증식적억제능력감약,가능시은설병적발병궤제지일.
Objective To evaluate the inhibitory effect of mesenchymal stem cells (MSCs) in lesions of patients with psoriasis on T lymphocyte proliferation.Methods Tissue specimens were obtained from the lesions of 15 patients with psoriasis vulgaris (7 at progressive stage and 8 at resting stage) and normal skin of 15 human controls from the Department of Urology and Plastic Surgery,Taiyuan City Centre Hospital.MSCs were isolated from these skin specimens,cultured,and identified using flow cytometry and in vitro differentiation assay.Enzyme-linked immunosorbent assay (ELISA) was performed to detect the concentration of interleukin (IL)-6,IL-1 1,hepatocyte growth factor (HGF) and transforming growth factor (TGF)-β1 in the culture supernatant of third-passage MSCs.Peripheral blood T cells were obtained from a healthy adult and cocultured with the third-passage MSCs for four days.Then,cells were counted and methyl thiazolyl tetrazolium (MTT) assay was conducted to evaluate the proliferation of T cells.One-way analysis of variance (ANOVA) and Student-Newman-Keuls (SNK) test were carried out to compare the proliferation of T lymphocytes,and two independent samples t test to compare the concentrations of cytokines.Results Inverted microscopy revealed that the patient-and control-derived MSCs shared similar morphological properties and multi-directional differentiation capacity,along with the expression of CD29,CD44,CD73,CD90 and CD105,but absence of CD34,CD45 and HLA-DR on cell surface.After coculture with MSCs from the patients and controls for four days,the count of T lymphocytes per milliliter was (1.67 ± 0.34) × 105 and (1.04 ± 0.29) × 105 respectively (P＜ 0.01),and the proliferative activity (expressed as absorbence at 492 nm)was 0.317 ± 0.021 and 0.275 ± 0.007 respectively (P ＜ 0.01).Compared with the control-derived MSCs,the patient-derived MSCs showed a significantly higher level of IL-1 1 ((181.37 ± 31.74) vs.(130.07 ± 29.20) ng/L,t =5.32,P ＜ 0.01),but a lower level of lL-6 ((61.67±17.53) vs.(76.74±18.96) ng/L,t=2.61,P＜0.05)and HGF ((319.24 ± 41.03) vs.(352.35 ± 51.47) ng/L,t =2.25,P＜ 0.05),as well as a similar level of TFG-β1,in the culture supernatant.Conclusions The inhibitory effect of MSCs in psoriatic lesions on T lymphocyte proliferation is diminished,which may contribute to the pathogenesis of psoriasis.