中华传染病杂志
中華傳染病雜誌
중화전염병잡지
CHINESE JOURNAL OF INFECTIOUS DISEASES
2011年
9期
529-533
,共5页
陈永平%程瑗%金晓芝%阳韬%王春莹%刘俊%郑明华
陳永平%程瑗%金曉芝%暘韜%王春瑩%劉俊%鄭明華
진영평%정원%금효지%양도%왕춘형%류준%정명화
CD4阳性T淋巴细胞%受体,白细胞介素7%白细胞介素2受体α亚单位%肝硬化%星形细胞%细胞,培养的%转化生长因子β1
CD4暘性T淋巴細胞%受體,白細胞介素7%白細胞介素2受體α亞單位%肝硬化%星形細胞%細胞,培養的%轉化生長因子β1
CD4양성T림파세포%수체,백세포개소7%백세포개소2수체α아단위%간경화%성형세포%세포,배양적%전화생장인자β1
CD4-positive T-lymphocytes%Receptors,interleukin-7%Interleukin-2 receptor alpha subunit%Liver cirrhosis%Astrocytes%Cells,cultured%Transforming growth factor beta 1
目的 了解CD4+ CD25+ CD127dim/-调节性T淋巴细胞在体外对肝星状细胞(HSC)增殖以及功能的影响，初步探讨调节性T淋巴细胞促肝纤维化的机制。方法传代培养HSC LX-2，将免疫磁珠细胞分选(MASC)法分离所得慢性乙型肝炎患者调节性T淋巴细胞与HSC LX-2按不同方式共培养5d，以单独培养的HSC作为对照，细胞计数试剂盒-8（CCK-8）法检测共培养HSC增殖情况，ELISA法检测上清液中转化生长因子(TGF)β1含量，放射免疫法检测HSC分泌HA、PCⅢ水平。统计学处理采用LSD-t检验。结果 5例调节性T淋巴细胞与HSC比例为1.5∶1时HSC增殖最为明显，10例直接接触共培养与使用Transwell系统共培养调节性T淋巴细胞与HSC吸光度值分别为(0.713±0.032)、(0.735±0.028) cpm，均较对照组的（0.677±0.029）cpm增殖明显(t=5.4003，8.7878；均P＜0.01)。10例直接接触共培养与Transwell组细胞上清液中TGFβ1浓度分别为(781.59±76.45)、（813.53±60.62）pg/mL，显著高于对照组的(722.51±59.66) pg/mL（t=4.0014，6.1653;均P＜0.01）；HA浓度分别为（433.57±27.90）、（445.40±23.73）ng/mL，显著高于对照组的（415.83±19.44）ng/mL（t=3.3124，5.5231;均P＜0.01）；PCⅢ浓度分别为(21.93±1.71)、(23.12±1.87) ng/mL，显著高于对照组的（20.10±1.49）ng/mL(f=4.8082,7.9436;均P＜0.01)。且Transwell组各项结果均显著高于直接接触组(t=3.3875，2.1639，2.2107，3.1354;均P＜0.05)。结论CD4+ CD25+ CD127dim调节性T淋巴细胞可促进共培养的HSC增殖及其HA、PCⅢ的分泌。体外实验证明，CD4+ CD25+ CD127dim/-调节性T淋巴细胞具有促进肝纤维化的重要作用。
目的 瞭解CD4+ CD25+ CD127dim/-調節性T淋巴細胞在體外對肝星狀細胞(HSC)增殖以及功能的影響，初步探討調節性T淋巴細胞促肝纖維化的機製。方法傳代培養HSC LX-2，將免疫磁珠細胞分選(MASC)法分離所得慢性乙型肝炎患者調節性T淋巴細胞與HSC LX-2按不同方式共培養5d，以單獨培養的HSC作為對照，細胞計數試劑盒-8（CCK-8）法檢測共培養HSC增殖情況，ELISA法檢測上清液中轉化生長因子(TGF)β1含量，放射免疫法檢測HSC分泌HA、PCⅢ水平。統計學處理採用LSD-t檢驗。結果 5例調節性T淋巴細胞與HSC比例為1.5∶1時HSC增殖最為明顯，10例直接接觸共培養與使用Transwell繫統共培養調節性T淋巴細胞與HSC吸光度值分彆為(0.713±0.032)、(0.735±0.028) cpm，均較對照組的（0.677±0.029）cpm增殖明顯(t=5.4003，8.7878；均P＜0.01)。10例直接接觸共培養與Transwell組細胞上清液中TGFβ1濃度分彆為(781.59±76.45)、（813.53±60.62）pg/mL，顯著高于對照組的(722.51±59.66) pg/mL（t=4.0014，6.1653;均P＜0.01）；HA濃度分彆為（433.57±27.90）、（445.40±23.73）ng/mL，顯著高于對照組的（415.83±19.44）ng/mL（t=3.3124，5.5231;均P＜0.01）；PCⅢ濃度分彆為(21.93±1.71)、(23.12±1.87) ng/mL，顯著高于對照組的（20.10±1.49）ng/mL(f=4.8082,7.9436;均P＜0.01)。且Transwell組各項結果均顯著高于直接接觸組(t=3.3875，2.1639，2.2107，3.1354;均P＜0.05)。結論CD4+ CD25+ CD127dim調節性T淋巴細胞可促進共培養的HSC增殖及其HA、PCⅢ的分泌。體外實驗證明，CD4+ CD25+ CD127dim/-調節性T淋巴細胞具有促進肝纖維化的重要作用。
목적 료해CD4+ CD25+ CD127dim/-조절성T림파세포재체외대간성상세포(HSC)증식이급공능적영향，초보탐토조절성T림파세포촉간섬유화적궤제。방법전대배양HSC LX-2，장면역자주세포분선(MASC)법분리소득만성을형간염환자조절성T림파세포여HSC LX-2안불동방식공배양5d，이단독배양적HSC작위대조，세포계수시제합-8（CCK-8）법검측공배양HSC증식정황，ELISA법검측상청액중전화생장인자(TGF)β1함량，방사면역법검측HSC분비HA、PCⅢ수평。통계학처리채용LSD-t검험。결과 5례조절성T림파세포여HSC비례위1.5∶1시HSC증식최위명현，10례직접접촉공배양여사용Transwell계통공배양조절성T림파세포여HSC흡광도치분별위(0.713±0.032)、(0.735±0.028) cpm，균교대조조적（0.677±0.029）cpm증식명현(t=5.4003，8.7878；균P＜0.01)。10례직접접촉공배양여Transwell조세포상청액중TGFβ1농도분별위(781.59±76.45)、（813.53±60.62）pg/mL，현저고우대조조적(722.51±59.66) pg/mL（t=4.0014，6.1653;균P＜0.01）；HA농도분별위（433.57±27.90）、（445.40±23.73）ng/mL，현저고우대조조적（415.83±19.44）ng/mL（t=3.3124，5.5231;균P＜0.01）；PCⅢ농도분별위(21.93±1.71)、(23.12±1.87) ng/mL，현저고우대조조적（20.10±1.49）ng/mL(f=4.8082,7.9436;균P＜0.01)。차Transwell조각항결과균현저고우직접접촉조(t=3.3875，2.1639，2.2107，3.1354;균P＜0.05)。결론CD4+ CD25+ CD127dim조절성T림파세포가촉진공배양적HSC증식급기HA、PCⅢ적분비。체외실험증명，CD4+ CD25+ CD127dim/-조절성T림파세포구유촉진간섬유화적중요작용。
Objective To investigate whether CD4+ CD25+ CD127dim/- regulatory T lymphocytes (Treg) can induce the proliferation of hepatic stellate cells (HSC) and expression of fibrosis-related factors on HSC in vitro and further to explore the mechanism of Treg inducing fibrogenesis. Methods HSC LX-2 cells were subcultured. CD4+ CD25+ CD127dim/- cells were purified using magnetic cell separation. The HSC were co-cultured with Treg by direct contact or by Transwell system in vitro. The HSC cultured alone was used as control. Cell proliferation was measured by CCK-8 assay.The expression of transforming growth factor (TGF)-β1 was detected by enzyme-linked inmunosorbent assay (ELISA), and the expressions of hyaluronic acid (HA) and precollagen Ⅲ (PC Ⅲ ) were detected by radio immunoassay (RIA). The data were analyzed by LSD-t test. Results HSC proliferation was strongest when Treg∶ HSC= 1.5∶ 1. The absorbance in direct contact co-culture group and Transwell system co-culture group was (0. 713±0. 032) cpm and (0. 735±0. 028) cpm, respectively, both of which were higher than that in control group [(0. 677 ± 0. 029) cpm](t = 5. 4003 and 8. 7878,respectively; both P＜0. 01). The concentrations of TGF-β1 in the supernatant were (781. 59 ±76.45) pg/mL and (813. 53±60. 62) pg/mL, respectively in direct contact co-culture group and Transwell system co-culture group, which were significantly higher than that in control group [(722.51±59. 66) pg/mL](t = 4.0014 and 6. 1653, respectively; both P＜0.01). The concentrations of HA were (433. 575±27.90) ng/mL and (445.40±23.73) ng/mL, respectively in direct contact co-culture group and Transwell system co-culture group, which were higher compared to that in control group [-(415. 83±19.44) ng/mL](t =3. 3124 and 5. 5231, respectively; both P＜0.01). Likewise, the concentrations of PC Ⅲ were (21. 93± 1.71) and (23. 125± 1.87) ng/mL in direct contact group and Transwell group, respectively compared to (20. 10± 1.49) ng/mL in control group (t = 4. 8082 and 7. 9436, respectively; both P ＜ 0.01). Furthermore, the absorbance,concentrations of TGF-β1, HA and PC Ⅲ in Transwell co-culture group were all higher compared to direct contact group (t = 3. 3875, 2.1639, 2. 2107 and 3.1354, respectively; all P＜0. 05).Conclusions The cell proliferation and the expressions of fibrosis-related factors in HSC increase greatly after co-cultured with CD4+ CD25+ CD127dim/- Treg. Therefore, Treg may play an important role in inducing liver fibrogenesis.
