應(yīng)趙建， 黃曉敏， 余埡妮， 林曉曉， 陳 晶， 李文君， 董 年， 陳成水

(溫州醫(yī)科大學(xué)附屬第一醫(yī)院呼吸與危重癥醫(yī)學(xué)科，浙江 溫州 325000)

PI3K/CTGF信號(hào)通路在TGF-β1誘導(dǎo)A549細(xì)胞表達(dá)collagen I過(guò)程中的作用*

應(yīng)趙建， 黃曉敏▲， 余埡妮， 林曉曉， 陳 晶， 李文君， 董 年△， 陳成水△

(溫州醫(yī)科大學(xué)附屬第一醫(yī)院呼吸與危重癥醫(yī)學(xué)科，浙江 溫州 325000)

目的： 研究磷脂酰肌醇3-激酶/結(jié)締組織生長(zhǎng)因子(PI3K/CTGF) 信號(hào)通路在轉(zhuǎn)化生長(zhǎng)因子β1(TGF-β1)誘導(dǎo)人肺腺癌A549細(xì)胞表達(dá)I型膠原蛋白(collagen I)過(guò)程中的分子機(jī)制。方法: 體外培養(yǎng)A549細(xì)胞，予TGF-β1刺激，觀察CTGF和collagen I的mRNA和蛋白表達(dá)及PI3K信號(hào)通路的活化；PI3K抑制劑LY294002預(yù)先處理A549細(xì)胞后，觀察TGF-β1刺激下CTGF和collagen I mRNA和蛋白表達(dá)的變化；CTGF特異性siRNA干擾A549細(xì)胞中CTGF的表達(dá)后，觀察TGF-β1刺激下collagen I mRNA和蛋白表達(dá)的變化和PI3K信號(hào)通路的活化。結(jié)果: TGF-β1可以誘導(dǎo)A549細(xì)胞中CTGF和collagen I的mRNA和蛋白表達(dá)以及PI3K信號(hào)通路的活化；PI3K特異性抑制劑LY294002可以部分逆轉(zhuǎn)TGF-β1誘導(dǎo)的A549細(xì)胞中CTGF和Collagen I mRNA和蛋白表達(dá)的升高。干擾CTGF可以降低TGF-β1誘導(dǎo)的A549細(xì)胞collagen I mRNA和蛋白表達(dá)，而不影響PI3K信號(hào)通路的活化。結(jié)論: CTGF是TGF-β1/PI3K信號(hào)通路調(diào)控的即刻早期反應(yīng)效應(yīng)蛋白，參與了TGF-β1誘導(dǎo)的A549細(xì)胞中collagen I表達(dá)。

轉(zhuǎn)化生長(zhǎng)因子β1； I型膠原蛋白； 結(jié)締組織生長(zhǎng)因子

細(xì)胞外基質(zhì)(extracellular matrix，ECM)代謝異常是腫瘤的重要特征之一，肺癌是高度纖維化的腫瘤，以I型膠原蛋白(collagen I)為代表的ECM過(guò)度沉積在肺癌的發(fā)生發(fā)展中扮演了重要角色[1-2]。既往認(rèn)為腫瘤間質(zhì)中成纖維細(xì)胞的激活及表型轉(zhuǎn)化是ECM沉積的中心環(huán)節(jié)，目前認(rèn)識(shí)到肺癌細(xì)胞在轉(zhuǎn)化生長(zhǎng)因子β1(transforming growth factor β1，TGF-β1)誘導(dǎo)下可以向成纖維細(xì)胞等間質(zhì)細(xì)胞分化從而參與ECM的異常沉積[3-4]。結(jié)締組織生長(zhǎng)因子(connective tissue growth factor，CTGF)是新近發(fā)現(xiàn)的早期即刻反應(yīng)蛋白，是TGF-β1調(diào)控的效應(yīng)蛋白，可以協(xié)同TGF-β1促進(jìn)細(xì)胞表型轉(zhuǎn)化和ECM沉積[5]，然而關(guān)于其在肺癌ECM代謝異常中的確切機(jī)制尚不清楚。結(jié)合我們之前的研究發(fā)現(xiàn)磷脂酰肌醇3-激酶(phosphatidyl-inositol 3-kinase，PI3K)信號(hào)通路的活化與上皮-間質(zhì)轉(zhuǎn)化和ECM的沉積密切相關(guān)[6-7]，本研究擬進(jìn)一步探討PI3K信號(hào)通路和效應(yīng)蛋白CTGF在TGF-β1誘導(dǎo)人肺腺癌A549細(xì)胞 collagen I表達(dá)中的潛在角色，闡明TGF-β1/PI3K/CTGF信號(hào)通路在A549細(xì)胞ECM合成的調(diào)控機(jī)制。

材 料 和 方 法

1 細(xì)胞

人肺腺癌細(xì)胞A549細(xì)胞購(gòu)于上海中科院細(xì)胞庫(kù)。

2 主要試劑

RPMI-1640培養(yǎng)基和胎牛血清購(gòu)自Gibco；人重組TGF-β1購(gòu)自PeproTech；PI3K抑制劑LY294002購(gòu)自Biovision；BCA蛋白濃度測(cè)定試劑盒、預(yù)染蛋白Marker和ECL發(fā)光液購(gòu)自Thermo；兔抗人p-Akt和Akt單抗購(gòu)自CST；兔抗人CTGF和collagen I單抗購(gòu)自Abcam；TRIzol購(gòu)自Invitrogen；cDNA逆轉(zhuǎn)錄試劑盒購(gòu)自TaKaRa；其它生化試劑購(gòu)自上海生工生物工程公司。Real-time PCR引物由上海生工生物工程公司設(shè)計(jì)合成，見(jiàn)表1；siRNA-CTGF由上海吉?jiǎng)P基因技術(shù)有限公司設(shè)計(jì)合成，見(jiàn)表2。

表1 Real-time PCR引物序列

表2 CTGF干擾RNA序列

3 方法

3.1 細(xì)胞培養(yǎng) A549細(xì)胞株使用包含10%胎牛血清和1%雙抗的RPMI-1640培養(yǎng)基于37 ℃、5% CO2恒溫培養(yǎng)箱中培養(yǎng)。

3.2 細(xì)胞干預(yù) 取生長(zhǎng)狀態(tài)良好處于對(duì)數(shù)生長(zhǎng)期的細(xì)胞，接種于12孔板(抽提RNA)或6孔板(抽提蛋白)，待細(xì)胞長(zhǎng)至孔板面積80%時(shí)，進(jìn)行分組干預(yù)，每個(gè)分組設(shè)立3個(gè)復(fù)孔，每個(gè)實(shí)驗(yàn)重復(fù)3次。

3.3 Real-time PCR實(shí)驗(yàn) 按照TRIzol說(shuō)明書提取細(xì)胞總RNA，分光光度法測(cè)定計(jì)算提取的總RNA含量及濃度。取總RNA 2 μg反轉(zhuǎn)錄為cDNA，再以適量cDNA為模板進(jìn)行real-time PCR實(shí)驗(yàn)。PCR反應(yīng)條件為95 ℃10 min； 95 ℃ 30 s, 60 ℃ 30 s, 72 ℃ 30 s，共40個(gè)循環(huán)。結(jié)果以GAPDH為內(nèi)參照，對(duì)目的基因進(jìn)行相對(duì)定量。

3.4 Western blot實(shí)驗(yàn) 收集各組細(xì)胞，提取總蛋白，BCA法測(cè)定蛋白濃度。每組取30 μg蛋白進(jìn)行凝膠電泳，濕轉(zhuǎn)至PVDF膜，5%脫脂牛奶室溫下封閉2 h， Ⅰ抗(1∶1 000)4 ℃孵育過(guò)夜，TBST緩沖液洗膜10 min、3次，Ⅱ抗(1∶5 000)室溫孵育1.5 h，再用TBST緩沖液洗膜10 min×3次，ECL化學(xué)發(fā)光法顯影。以β-actin為內(nèi)參照，結(jié)果以各蛋白與β-actin灰度值比值表示。

3.5 siRNA轉(zhuǎn)染 siRNA轉(zhuǎn)染方法按照Lipofectamine 2000產(chǎn)品說(shuō)明書進(jìn)行，提前1 d將1×105個(gè)細(xì)胞接種于24孔板，次日按照24孔板每孔取2 μL CTGF-siRNA、1 μL Lipofectamine 2000和50 μL Opti-MEM I的比例配制CTGF-siRNA-轉(zhuǎn)染試劑混合液，將siRNA-轉(zhuǎn)染試劑混合液添加入細(xì)胞培養(yǎng)板中，置于37 ℃、5% CO2的培養(yǎng)箱中培養(yǎng)，6 h后更換為含10%胎牛血清的RPMI-1640培養(yǎng)基。

4 統(tǒng)計(jì)學(xué)處理

使用SPSS 20.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)分析。數(shù)據(jù)均以均數(shù)±標(biāo)準(zhǔn)差(mean±SD)，多組間比較采用單因素方差分析(one-way ANOVA)，組間兩兩比較采用Bonferroni校正的t檢驗(yàn)，以P<0.05為差異有統(tǒng)計(jì)學(xué)意義。

結(jié) 果

1 TGF-β1誘導(dǎo)collagen I表達(dá)升高

選取不同濃度的TGF-β1刺激A549細(xì)胞不同時(shí)間， real-time qPCR和Western blot法檢測(cè)collagen I的mRNA和蛋白的表達(dá)。結(jié)果顯示，TGF-β1可以誘導(dǎo)A549細(xì)胞中collagen I的表達(dá)，呈現(xiàn)濃度和時(shí)間依賴性升高，其中5 μg/L TGF-β1是最適刺激濃度，TGF-β1刺激48 h collagen I升高最為顯著，差異有統(tǒng)計(jì)學(xué)意義(P<0.05)，見(jiàn)圖1。

Figure 1.The expression of collagen I induced by TGF-β1. A: the mRNA expression of collagen I induced by TGF-β1 at different doses for the indicated time; B: the protein level of collagen I changed by TGF-β1 at different doses for 48 h; C: the protein level of collagen I changed by TGF-β1 at 5 μg/L for the indicated time. Mean±SD. n=3. *P<0.05, **P<0.01 vs 0 h; #P<0.05, ##P<0.01 vs 0 μg/L.

2 TGF-β1誘導(dǎo)CTGF的即刻早期表達(dá)

選取5 μg/L TGF-β1刺激A549細(xì)胞不同時(shí)間，結(jié)果顯示TGF-β1可以誘導(dǎo)A549細(xì)胞中CTGF即刻早期表達(dá)，mRNA和蛋白在TGF-β1刺激3 h時(shí)即發(fā)生升高，TGF-β1刺激6 h時(shí)CTGF蛋白表達(dá)最高，差異有統(tǒng)計(jì)學(xué)意義(P<0.05)，見(jiàn)圖2。

Figure 2.The expression of CTGF induced by TGF-β1. A: the mRNA expression of CTGF induced by TGF-β1 at different doses for the indicated time; B: the protein level of CTGF changed by TGF-β1 at different doses for 48 h; C: the protein level of CTGF changed by TGF-β1 at 5 μg/L for the indicated time. Mean±SD. n=3. *P<0.05, **P<0.01 vs 0 h group; #P<0.05, ##P<0.01 vs 0 μg/L.

3 TGF-β1活化PI3K信號(hào)通路,PI3K抑制劑逆轉(zhuǎn)TGF-β1誘導(dǎo)的CTGF和collagen I表達(dá)升高

選取不同濃度的TGF-β1刺激A549細(xì)胞不同時(shí)間，用Western blot法檢測(cè)，可見(jiàn)TGF-β1可以激活PI3K信號(hào)通路，呈現(xiàn)濃度和時(shí)間依賴性。使用不同濃度的PI3K抑制劑LY294002預(yù)先處理A549細(xì)胞，再使用5 μg/L TGF-β1刺激48 h，結(jié)果可見(jiàn)PI3K抑制劑 LY294002可以逆轉(zhuǎn)TGF-β1誘導(dǎo)的A549細(xì)胞中CTGF和collagen I表達(dá)，呈現(xiàn)劑量依賴性，差異有統(tǒng)計(jì)學(xué)意義(P<0.05)，見(jiàn)圖3。

Figure 3.TGF-β1 induced PI3K/Akt signaling pathway activation, and PI3K inhibitor reversed the expression of CTGF and collagen I induced by TGF-β. A: the protein levels of Akt and p-Akt changed by with the vehicle or TGF-β1 at different doses for 10 min; B: the protein levels of Akt and p-Akt changed by TGF-β1 at dose of 5 μg/L for different time; C: the mRNA and protein levels of CTGF were measured after incubation with PI3K inhibitor LY294002 at different doses and TGF-β1 (5 μg/L) for 6 h; D: the mRNA and protein levels of collagen I were measured after incubation with PI3K inhibitor LY294002 at different doses and TGF-β1 (5 μg/L) for 48 h. Mean±SD. n=3. *P<0.05, **P<0.01 vs Con (control) group; △△P<0.01 vs 0 min group; #P<0.05, ##P<0.01 vs TGF-β1 group.

4 siRNA-CTGF-2是最佳干擾序列并影響collagen I的基礎(chǔ)表達(dá)

4條不同的干擾序列siRNA-CTGF轉(zhuǎn)染A549細(xì)胞24 h后，用real-time qPCR和Western blot分別檢測(cè)CTGF的mRNA和蛋白表達(dá)，結(jié)果可見(jiàn)siRNA-CTGF-2干擾A549細(xì)胞中CTGF表達(dá)的效果最佳，見(jiàn)圖4A。與對(duì)照組及siRNA陰性對(duì)照組相比，siRNA-CTGF-2組的collagen I基礎(chǔ)表達(dá)降低，差異具有統(tǒng)計(jì)學(xué)意義(P<0.05)，見(jiàn)圖4B。

Figure 4.siRNA-CTGF-2 was the best interference sequence and impaired the basal collagen I synthesis. A: the mRNA and protein expression of CTGF modified by different siRNAs; B: the mRNA and protein expression of collagen I were measured after CTGF interference by siRNA-CTGF2. Mean±SD. n=3. *P<0.05,**P<0.01 vs Con (control) group; #P<0.05, ##P<0.01 vs siRNA-Con group.

5 干擾CTGF表達(dá)影響TGF-β1誘導(dǎo)的collagen I表達(dá)升高而不影響PI3K信號(hào)通路的活化

siRNA-CTGF-2轉(zhuǎn)染24 h干擾A549細(xì)胞中CTGF的表達(dá)，再用5 μg/L TGF-β1刺激10 min，結(jié)果顯示siRNA-CTGF-2轉(zhuǎn)染并不影響TGF-β1活化PI3K信號(hào)通路，差異無(wú)統(tǒng)計(jì)學(xué)顯著性，見(jiàn)圖5A。然而，siRNA-CTGF-2轉(zhuǎn)染明顯降低TGF-β1誘導(dǎo)的collagen I表達(dá)升高，差異有統(tǒng)計(jì)學(xué)意義(P<0.05)，見(jiàn)圖5B。

Figure 5.Interference of CTGF expression impaired TGF-β1-induced collagen I synthesis but not PI3K/AKT signaling pathway activation. A: the protein levels of Akt and p-Akt changed by the vehicle or TGF-β1 at dose of 5 μg/L for 10 min after CTGF interference; B: TGF-β1-induced mRNA and protein expression of collagen I was measured after CTGF interference. Mean±SD. n=3. *P<0.05, **P<0.01 vs Con (control) or siRNA-Con group; ##P<0.01 vs siRNA-CTGF-2 group; △P<0.05, △△P<0.01 vs TGF-β1+siRNA-Con group.

討 論

既往認(rèn)為腫瘤間質(zhì)中成纖維細(xì)胞的激活及表型轉(zhuǎn)化是腫瘤胞外基質(zhì)代謝異常的關(guān)鍵[8]，目前逐漸認(rèn)識(shí)到腫瘤細(xì)胞可以發(fā)生間質(zhì)轉(zhuǎn)化不僅扮演了ECM沉積啟動(dòng)細(xì)胞，釋放的TGF-β1等轉(zhuǎn)化生長(zhǎng)因子參與腫瘤間質(zhì)中成纖維細(xì)胞的活化，更扮演了繼發(fā)受體的角色，其分泌的炎癥因子可以以自分泌或旁分泌的方式作用于自身[9]。TGF-β1是關(guān)鍵的促纖維化因子，主要激活經(jīng)典的Smad通路和非經(jīng)典Smad通路共同介導(dǎo)ECM中膠原蛋白(collagen)和纖連蛋白(fibronectin)等的沉積[10-11]。近來(lái)的研究揭示非經(jīng)典Smad通路PI3K的過(guò)度活化參與了肺癌的發(fā)生發(fā)展，我們之前的實(shí)驗(yàn)發(fā)現(xiàn)PI3K抑制劑可以逆轉(zhuǎn)慢性氣道重塑中膠原的沉積，然而關(guān)于PI3K如何參與肺癌胞外機(jī)制代謝異常研究甚少。CTGF屬于早期即刻反應(yīng)蛋白，影響成纖維細(xì)胞的活性，在纖維化的發(fā)生發(fā)展中發(fā)揮重要作用[12-13]。作為TGF-β1的調(diào)控蛋白，研究報(bào)道TGF-β1主要是經(jīng)Smad通路調(diào)控CTGF，然而有報(bào)道稱哺乳動(dòng)物雷帕霉素靶蛋白抑制劑可以反向激活PI3K通路促進(jìn)肺癌細(xì)胞中CTGF的過(guò)度表達(dá)，協(xié)同TGF-β1參與了ECM沉積[14-15]。本研究旨在探究PI3K信號(hào)通路和效應(yīng)蛋白CTGF在TGF-β1誘導(dǎo)A549細(xì)胞的collagen I表達(dá)中的的潛在角色，為未來(lái)針對(duì)肺癌ECM代謝異常揭示潛在的干預(yù)靶點(diǎn)。

我們研究發(fā)現(xiàn)TGF-β1可以誘導(dǎo)A549細(xì)胞的collagen I的表達(dá)升高，與之前的研究報(bào)道類似，除此之外，TGF-β1可以誘導(dǎo)CTGF的早期表達(dá)和PI3K信號(hào)通路的活化。使用PI3K抑制劑可以逆轉(zhuǎn)TGF-β1誘導(dǎo)的CTGF的早期表達(dá)和collagen I的表達(dá)升高，提示我們PI3K信號(hào)通路和經(jīng)典的Smad通路共同參與了TGF-β1調(diào)控的CTGF早期表達(dá)。我們進(jìn)一步經(jīng)siRNA-CTGF特異性降低A549細(xì)胞中CTGF的表達(dá)，發(fā)現(xiàn)A549細(xì)胞的collagen I基礎(chǔ)表達(dá)及TGF-β1誘導(dǎo)的collagen I表達(dá)同時(shí)發(fā)生了降低，而不影響TGF-β1活化PI3K信號(hào)通路。

總而言之，我們研究發(fā)現(xiàn)TGF-β1經(jīng)活化PI3K信號(hào)通路調(diào)控A549細(xì)胞中collagen I的表達(dá)升高，CTGF是TGF-β1/PI3K信號(hào)通路潛在的調(diào)控蛋白，協(xié)同TGF-β1誘導(dǎo)A549細(xì)胞中collagen I的表達(dá)升高，初步闡述了TGF-β1/PI3K/CTGF信號(hào)通路在調(diào)控A549細(xì)胞collagen I表達(dá)中的分子機(jī)制。CTGF和PI3K信號(hào)通路可能是未來(lái)針對(duì)肺癌ECM過(guò)度沉積的潛在干預(yù)靶點(diǎn)。

[1] Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation[J]. Cell, 2011, 144(5): 646-674.

[2] Shintani Y, Maeda M, Chaika N, et al. Collagen I promotes epithelial-to-mesenchymal transition in lung cancer cells via transforming growth factor-beta signaling[J]. Am J Respir Cell Mol Biol, 2008, 38(1): 95-104.

[3] 岳喜磊, 成 瑩, 許繼德, 等. TRPC1在 TGF-β1誘導(dǎo)支氣管上皮細(xì)胞間充質(zhì)轉(zhuǎn)化中的作用[J]. 中國(guó)病理生理雜志, 2015,31(3): 492-498.

[4] Wang X, Adler KB, Erjefalt J, et al. Airway epithelial dysfunction in the development of acute lung injury and acute respiratory distress syndrome[J]. Expert Rev Respir Med, 2007, 1(1): 149-155.

[5] 楊 敏, 黃海長(zhǎng), 王海燕. 結(jié)締組織生長(zhǎng)因子促纖維化作用及其表達(dá)調(diào)節(jié)的研究進(jìn)展[J]. 中國(guó)病理生理雜志, 2005, 21(1): 199-202,208.

[6] Chen C, Fang X, Wang Y, et al. Preventive and therapeutic effects of phosphoinositide 3-kinase inhibitors on acute lung injury[J]. Chest, 2011, 140(2): 391-400.

[7] Fang X, Li K, Tao X, et al. Effects of phosphoinositide 3-kinase on protease-induced acute and chronic lung inflammation, remodeling, and emphysema in rats[J]. Chest, 2013, 143(4): 1025-1035.

[8] Phan SH. Genesis of the myofibroblast in lung injury and fibrosis[J]. Proc Am Thorac Soc, 2012, 9(3): 148-152.

[9] Yang J, Velikoff M, Canalis E, et al. Activated alveolar epithelial cells initiate fibrosis through autocrine and paracrine secretion of connective tissue growth factor[J]. Am J Physiol Lung Cell Mol Physiol, 2014, 306(8): L786-L796.

[10]王來(lái)亮, 羅 群. 腎小管上皮間充質(zhì)轉(zhuǎn)化與腎臟纖維化[J]. 中國(guó)病理生理雜志, 2014, 30(10): 1910-1914,1920.

[11]馬文東, 袁 媛, 楊 奕, 等. TGF-β1介導(dǎo)的RhoA/ROCK通路在大鼠肺肌成纖維細(xì)胞分化中的調(diào)節(jié)作用[J]. 中國(guó)病理生理雜志, 2013, 29(10): 1758-1763.

[12]王達(dá)安, 林逸心, 王 崢, 等. 沙利度胺抑制 TGF-β1誘導(dǎo)的 HELF 細(xì)胞中結(jié)締組織生長(zhǎng)因子基因啟動(dòng)子的激活[J]. 中國(guó)病理生理雜志, 2014,30(4): 693-697.

[13]Mason RM. Connective tissue growth factor(CCN2), a pathogenic factor in diabetic nephropathy. What does it do? How does it do it? [J].J Cell Commun Signal, 2009, 3(2): 95-104.

[14]Xu X, Wan X, Geng J, et al. Rapamycin regulates connective tissue growth factor expression of lung epithelial cells via phosphoinositide 3-kinase[J]. Exp Biol Med (Maywood), 2013, 238(9): 1082-1094.

[15]Xu X, Dai H, Geng J, et al. Rapamycin increases CCN2 expression of lung fibroblasts via phosphoinositide 3-kinase[J]. Lab Invest, 2015, 95(8): 846-859.

(責(zé)任編輯： 林白霜， 羅 森)

Effects of PI3K/CTGF signaling pathway on TGF-β1-induced collagen I expression in A549 cells

YING Zhao-jian, HUANG Xiao-min, YU Ya-ni, LIN Xiao-xiao, CHEN Jing, LI Wen-jun, DONG Nian, CHEN Cheng-shui

(DepartmentofPulmonaryandCriticalCareMedicine,TheFirstAffiliatedHospital,WenzhouMedicalUniversity,Wenzhou325000,China.E-mail:wzmudongnian@foxmail.com;wzchencs@163.com)

AIM: To investigate the molecular mechanism of transforming growth factor β1/phosphatylinositol 3-kinase/connective tissue growth factor (TGF-β1/PI3K/CTGF) signaling pathway in the induction of collagen I expression in human lung adenocarcinoma A549 cells. METHODS: The A549 cells were culturedinvitroand stimulated by TGF-β1. The expression of CTGF and collagen I at mRNA and protein levels as well as the activation of PI3K was measured. The A549 cells were pre-treated with PI3K inhibitor LY294002, the expression of CTGF and collagen I at mRNA and protein levels was measured upon TGF-β1 stimulation. Under the condition ofCTGFinterference by siRNA-CTGF, the expression of collagen I at mRNA and protein levels as well as the activation of PI3K by TGF-β1 stimulation was measured. RESULTS: TGF-β1 stimulated the expression of CTGF and collagen I at mRNA and protein levels as well as the activation of PI3K at a dose- and time-dependant manner. PI3K inhibitor LY294002 partly reversed TGF-β1-induced CTGF and collagen I expression. Interference ofCTGFdown-regulated TGF-β1-induced collagen I expression without the effect on the activation of PI3K. CONCLUSION: PI3K signaling pathway plays a pivotal role in TGF-β1-induced collagen I expression in the A549 cells. CTGF is an immediate early response effector protein of TGF-β1/PI3K signaling pathway and synergizes with TGF-β1 in the induction of collagen I expression in the A549 cells.

Transforming growth factor β1; Collagen I; Connective tissue growth factor

1000- 4718(2017)03- 0489- 06

2016- 09- 21

2016- 12- 01

國(guó)家自然科學(xué)基金資助項(xiàng)目(No. 81270131； No. 81570075)

△通訊作者 董 年 Tel: 0577-86352962; E-mail: wzmudongnian@foxmail.com; 陳成水 Tel: 0577-55578186; E-mail: wzchencs@163.com

▲并列第1作者

R730.23

A

10.3969/j.issn.1000- 4718.2017.03.017