中华实验眼科杂志
中華實驗眼科雜誌
중화실험안과잡지
CHINESE JOURNAL OF EXPERIMENTAL OPHTHALMOLOGY
2012年
6期
520-524
,共5页
三氧化二砷%表皮生长因子%视网膜色素上皮细胞%细胞增生%细胞迁移
三氧化二砷%錶皮生長因子%視網膜色素上皮細胞%細胞增生%細胞遷移
삼양화이신%표피생장인자%시망막색소상피세포%세포증생%세포천이
Arsenic trioxide%Epidermal growth factor%Retinal pigment epithelial cell%Proliferation%Migration
背景 细胞因子失衡所导致的视网膜色素上皮(RPE)细胞的异常增生和迁移是增生性玻璃体视网膜病变( PVR)的主要病理变化之一.三氧化二砷(As2O3)是中国传统中药中的有效成分,可有效抑制肿瘤细胞的增生和迁移.但As2O3对细胞生长因子引起的RPE细胞增生和迁移的影响尚未明确. 目的 探讨As2O3对表皮生长因子(EGF)诱导的ARPE-19细胞增生和迁移的影响.方法 用无血清培养基对RPE细胞系ARPE-19细胞进行培养,将终浓度为0、0.5、1.0、2.0、5.0、10.0和20.0 μmol/L的As2O3分别加入到无血清培养基和含10 mg/L EGF的ARPE-19细胞培养液中作用24 h和48 h,通过噻唑蓝(MTT)比色法检测各培养组ARPE-19细胞活性的吸光度(A)值,以探讨As2O3对细胞的药物毒性作用,并筛选安全、有效的As2O3作用浓度.用10 mg/L EGF加入培养基诱导ARPE-19细胞迁移,分别在培养板中加入0、0.5、1.0、2.0μmol/L As2O3 作用24 h和48 h,并通过划痕试验和Transwell试验检测As2O3对EGF诱导的ARPE-19细胞迁移的影响.结果 MTT法检测发现不同浓度As2O3组作用24 h和48 h后,无血清培养组细胞A值随As2O3浓度的升高而逐渐下降,总体差异有统计学意义(F浓度=38.269,P=0.000；F时间=0.874,P=0.358).与空白对照组(0μmol/LAs2O3组)比较,0.5～ 5.0 μmol/L As2O3组ARPE-19细胞A值的差异均无统计学意义(P＞0.05).对含10 mg/LEGF组的ARPE-19细胞,药物对细胞A值的影响呈现浓度和时间依赖性(F浓度=152.155,P=0.000;F时间=51.649,P=0.000).与对照组比较,0.5～2.0μmol/L As2O3加入24 h和48 h后,A值的变化差异均无统计学意义(P＞0.05),而0.5、1.0、2.0μmol/L As2O3加入10 mg/L EGF诱导的ARPE-19细胞中作用24h和48 h后,A值的变化差异均无统计学意义(F浓度=2.215,P=0.126;F时间 =2.230,P=0.155).5.0 ～20.0 μmol/L As2O3作用于EGF诱导的ARPE-19细胞中作用后,细胞A值明显下降,与空白对照组比较差异均有统计学意义(P＜0.05),5.0～20.0 μmol/L As2O3作用24 h后,对EGF诱导的ARPE-19细胞增生抑制率分别为12％、32％、37％；作用48 h后细胞抑制率分别为39％、44％和53％.划痕试验结果显示,0.5～2.0 μmol/L As2O3对EGF诱导的ARPE-19细胞的横向迁移具有抑制作用.Transwell试验结果表明,0.5～2.0 μmol/L As2O3对10 mg/L EGF诱导的ARPE-19细胞纵向迁移有明显的抑制作用,0.5、1.0、2.0μmol/L As2O3作用12h对ARPE-19细胞的抑制率分别为22％、33％和46％. 结论 As2O3在一定浓度范围内对ARPE-19细胞无毒性作用,2.0 μmol/L以下浓度的As2O3对EGF诱导的ARPE-19细胞增生无明显影响,但可影响细胞的迁移能力,5.0 μmol/L以上浓度的As2O3可明显抑制ARPE-19细胞的增生.
揹景 細胞因子失衡所導緻的視網膜色素上皮(RPE)細胞的異常增生和遷移是增生性玻璃體視網膜病變( PVR)的主要病理變化之一.三氧化二砷(As2O3)是中國傳統中藥中的有效成分,可有效抑製腫瘤細胞的增生和遷移.但As2O3對細胞生長因子引起的RPE細胞增生和遷移的影響尚未明確. 目的 探討As2O3對錶皮生長因子(EGF)誘導的ARPE-19細胞增生和遷移的影響.方法 用無血清培養基對RPE細胞繫ARPE-19細胞進行培養,將終濃度為0、0.5、1.0、2.0、5.0、10.0和20.0 μmol/L的As2O3分彆加入到無血清培養基和含10 mg/L EGF的ARPE-19細胞培養液中作用24 h和48 h,通過噻唑藍(MTT)比色法檢測各培養組ARPE-19細胞活性的吸光度(A)值,以探討As2O3對細胞的藥物毒性作用,併篩選安全、有效的As2O3作用濃度.用10 mg/L EGF加入培養基誘導ARPE-19細胞遷移,分彆在培養闆中加入0、0.5、1.0、2.0μmol/L As2O3 作用24 h和48 h,併通過劃痕試驗和Transwell試驗檢測As2O3對EGF誘導的ARPE-19細胞遷移的影響.結果 MTT法檢測髮現不同濃度As2O3組作用24 h和48 h後,無血清培養組細胞A值隨As2O3濃度的升高而逐漸下降,總體差異有統計學意義(F濃度=38.269,P=0.000；F時間=0.874,P=0.358).與空白對照組(0μmol/LAs2O3組)比較,0.5～ 5.0 μmol/L As2O3組ARPE-19細胞A值的差異均無統計學意義(P＞0.05).對含10 mg/LEGF組的ARPE-19細胞,藥物對細胞A值的影響呈現濃度和時間依賴性(F濃度=152.155,P=0.000;F時間=51.649,P=0.000).與對照組比較,0.5～2.0μmol/L As2O3加入24 h和48 h後,A值的變化差異均無統計學意義(P＞0.05),而0.5、1.0、2.0μmol/L As2O3加入10 mg/L EGF誘導的ARPE-19細胞中作用24h和48 h後,A值的變化差異均無統計學意義(F濃度=2.215,P=0.126;F時間 =2.230,P=0.155).5.0 ～20.0 μmol/L As2O3作用于EGF誘導的ARPE-19細胞中作用後,細胞A值明顯下降,與空白對照組比較差異均有統計學意義(P＜0.05),5.0～20.0 μmol/L As2O3作用24 h後,對EGF誘導的ARPE-19細胞增生抑製率分彆為12％、32％、37％；作用48 h後細胞抑製率分彆為39％、44％和53％.劃痕試驗結果顯示,0.5～2.0 μmol/L As2O3對EGF誘導的ARPE-19細胞的橫嚮遷移具有抑製作用.Transwell試驗結果錶明,0.5～2.0 μmol/L As2O3對10 mg/L EGF誘導的ARPE-19細胞縱嚮遷移有明顯的抑製作用,0.5、1.0、2.0μmol/L As2O3作用12h對ARPE-19細胞的抑製率分彆為22％、33％和46％. 結論 As2O3在一定濃度範圍內對ARPE-19細胞無毒性作用,2.0 μmol/L以下濃度的As2O3對EGF誘導的ARPE-19細胞增生無明顯影響,但可影響細胞的遷移能力,5.0 μmol/L以上濃度的As2O3可明顯抑製ARPE-19細胞的增生.
배경 세포인자실형소도치적시망막색소상피(RPE)세포적이상증생화천이시증생성파리체시망막병변( PVR)적주요병리변화지일.삼양화이신(As2O3)시중국전통중약중적유효성분,가유효억제종류세포적증생화천이.단As2O3대세포생장인자인기적RPE세포증생화천이적영향상미명학. 목적 탐토As2O3대표피생장인자(EGF)유도적ARPE-19세포증생화천이적영향.방법 용무혈청배양기대RPE세포계ARPE-19세포진행배양,장종농도위0、0.5、1.0、2.0、5.0、10.0화20.0 μmol/L적As2O3분별가입도무혈청배양기화함10 mg/L EGF적ARPE-19세포배양액중작용24 h화48 h,통과새서람(MTT)비색법검측각배양조ARPE-19세포활성적흡광도(A)치,이탐토As2O3대세포적약물독성작용,병사선안전、유효적As2O3작용농도.용10 mg/L EGF가입배양기유도ARPE-19세포천이,분별재배양판중가입0、0.5、1.0、2.0μmol/L As2O3 작용24 h화48 h,병통과화흔시험화Transwell시험검측As2O3대EGF유도적ARPE-19세포천이적영향.결과 MTT법검측발현불동농도As2O3조작용24 h화48 h후,무혈청배양조세포A치수As2O3농도적승고이축점하강,총체차이유통계학의의(F농도=38.269,P=0.000；F시간=0.874,P=0.358).여공백대조조(0μmol/LAs2O3조)비교,0.5～ 5.0 μmol/L As2O3조ARPE-19세포A치적차이균무통계학의의(P＞0.05).대함10 mg/LEGF조적ARPE-19세포,약물대세포A치적영향정현농도화시간의뢰성(F농도=152.155,P=0.000;F시간=51.649,P=0.000).여대조조비교,0.5～2.0μmol/L As2O3가입24 h화48 h후,A치적변화차이균무통계학의의(P＞0.05),이0.5、1.0、2.0μmol/L As2O3가입10 mg/L EGF유도적ARPE-19세포중작용24h화48 h후,A치적변화차이균무통계학의의(F농도=2.215,P=0.126;F시간 =2.230,P=0.155).5.0 ～20.0 μmol/L As2O3작용우EGF유도적ARPE-19세포중작용후,세포A치명현하강,여공백대조조비교차이균유통계학의의(P＜0.05),5.0～20.0 μmol/L As2O3작용24 h후,대EGF유도적ARPE-19세포증생억제솔분별위12％、32％、37％；작용48 h후세포억제솔분별위39％、44％화53％.화흔시험결과현시,0.5～2.0 μmol/L As2O3대EGF유도적ARPE-19세포적횡향천이구유억제작용.Transwell시험결과표명,0.5～2.0 μmol/L As2O3대10 mg/L EGF유도적ARPE-19세포종향천이유명현적억제작용,0.5、1.0、2.0μmol/L As2O3작용12h대ARPE-19세포적억제솔분별위22％、33％화46％. 결론 As2O3재일정농도범위내대ARPE-19세포무독성작용,2.0 μmol/L이하농도적As2O3대EGF유도적ARPE-19세포증생무명현영향,단가영향세포적천이능력,5.0 μmol/L이상농도적As2O3가명현억제ARPE-19세포적증생.
Background Growth factor-induced proliferation and migration of retinal pigment epithelium (RPE) cells are the major pathological changes of proliferative vitreoretinopathy (PVR).Arsenic trioxide ( As2O3 ) is an active ingredient of Chinese traditional medicines,which has an inhibition on proliferation and migration of tumor cells.However,it is not clear whether As2O3 could inhibit growth factor-induced proliferation and migration of RPE cells. Objective This study was to explore the effects of As2O3 on epidermal growth factor (EGF)-induced proliferation and migration of ARPE-19 cells. Methods RPE cell line (ARPE-19 cells) were cultured.Different concentrations of As2O3(0,0.5,1.0,2.0,5.0,10.0,20.0 μmol/L) were added in the culture plate to treat ARPE-19 cells with or without 10 mg/L EGF in serum-free group for 24 and 48 hours,respectively.The MTT colorimetric assay was used to check the cell viability and evaluate the drug toxicity.The effects of As2O3 on EGF-induced proliferation of ARPE-19 cells were analyzed to get an effective and avirulent concentrations of As2O3.The effects of As2O3 on EGF-induced migration of ARPE-19 cells were observed by scratch-wound assay and the Boyden chamber assay.Results MTT assay showed that the A values were gradually declined with the increase of As2O3 concentrations after As2O3 treatment without EGF for 24 hours and 48 hours ( Fgroup =38.269,P =0.000 ; Ftime =0.874,P =0.358 ).Compared with the control group,no significant differences were seen in the A values of ARPE-19 cells in 0.5-5.0 μmol/L groups (all P＞0.05).Meantime,As2O3 reduced the A values of ARPE-19 cell with 10 mg/L EGF in dose- and time-dependent manner ( Fgroup =152.155,P =0.000 ; Ftime =51.649,P =0.000 ).There were not significant differences in 10 mg/L EGF-induced cell growth after 0.5,1.0,2.0 μmol/L As2O3 was added for 24 and 48 hours ( Fgroup =2.215,P =0.126 ;Ftime =2.230,P =0.155).However,when 5.0-20.0 μmol/L As2O3 added,the A values of 10 mg/L EGF-induced ARPE-19 cells lowed,showing a significant difference in comparison with the control groups ( all P＜0.05),with the cellular inhibiting rate 12％,32％,37％ in 24 hours and 39％,44％ and 53％ in 48 hours.Scratch-wound assay showed that EGF-induced horizontal migration of ARPE-19 cells was slow after 0.5-2.0 μmol/L As2O3 treated,and the same results also appeared in cell lognitudinal migration by Boyden chamber assay,with the inhibitory rates 22％,33％ and 46％ respectively. Conclusions As2O3 is avirulent on ARPE-19 cells within definite concentration range.At ≤ 2.0 μmol/L concentrations,As2O3 dose not affect EGF-induced proliferation of ARPE-19 cells,but it suppresses EGF-induced cell migration.At ≥ 5.0 μmol/L concentrations,As2O3 plays an inhibitory role to EGF-induced proliferation of ARPE-19 cells.
