Relationship of microRNA-133a and TGF-β1 protein in myocardial tissues of spontaneously hypertensive rats

AIM：To observe the changes of microRNA-133a and transforming growth factor β1 (TGF-β1) protein in the myocardium of spontaneously hypertensive rats (SHR). METHODS：Male SHR (18 weeks old, n=12) and male Wistar-Kyoto rats (WKY, 18 weeks old, n=12) served as SHR group and control group, respectively. Caudal arterial blood pressure was detected by a noninvasive blood pressure measurement and analysis system. Myocardial collagen volume fraction (CVF) and perivascular collagen area ratio (PVCA) were determined by Masson staining. The level of miR-133a in the heart was detected by real-time quantitative PCR. The protein level of TGF-β1 in the heart was also analyzed by the methods of immunohistochemisty and Western blotting. RESULTS：Compared with control group, systolic and diastolic blood pressure, CVF and PVCA significantly increased, the expression of TGF-β1 protein was significantly up-regulated, and the level of miR-133a was significantly reduced in SHR group. In SHR group, the expression of miR-133a was decreased to (23.9±4.6)% in control group. A negative correlation between the levels of miR-133a and TGF-β1 protein in SHR group was observed (r=-0.791, P<0.01). CONCLUSION：The level of miR-133a is down-regulated along with the up-regulation of TGF-β1 protein expression and collagen synthesis in the myocardial tissues of SHR. miR-133a and TGF-β1 may be involved in myocardial fibrosis in SHR.     

Ginsenoside Rh1 attenuates unilateral ureteral obstruction-induced renal interstitial fibrosis in rats

AIM:To observe the effects of ginsenoside Rh1 (G-Rh1) on unilateral ureteral obstruction (UUO)-induced renal interstitial fibrosis and to investigate the underlying mechanisms. METHODS:Male Sprague-Dawley (SD) rats (n=40) were divided into the following 4 groups:UUO-operated group (UUO group), sham-operated group (sham group), UUO-operated plus a low dose (50 mg·kg^-1·d^-1) of G-Rh1 treatment (low G-Rh1 group) and UUO-operated plus a high dose (100 mg·kg^-1·d^-1) of G-Rh1 treatment group (high G-Rh1 group). The G-Rh1 treatment was carried out by gastric gavage from the next day after the UUO operation once a day for 2 weeks (14 d). Immediately after the final dose of G-Rh1, 24 h urine was collected for the urine protein test, and then the rats were euthanized. The blood was collected for the blood urea nitrogen (BUN) and serum creatinine (SCr) assays, and the kidney was removed for pathological and biochemical evaluations. RESULTS:The levels of 24 h urine protein did not show any significant diffe-rence among the groups, while significantly increased levels of BUN and SCr in UUO group were observed (P<0.05), which was prevented by the treatment with G-Rh1 at both doses in a dose-dependent manner. Pathological evaluation showed the renal tissue damage was obvious in UUO group, which was improved by the treatment with G-Rh1 at both doses. Immunohistochemcial analysis exhibited that UUO increased renal interstitial transforming growth factor-β₁ (TGF-β₁) expression, which was also inhibited by the treatment with G-Rh1 at both doses(P<0.05). Significantly increased protein expression of renal interstitial collagen type I, α-smooth muscle actin (α-SMA) and connective tissue growth factor (CTGF) in UUO group was detected, which was suppressed by the treatment with G-Rh1 at both doses. CONCLUSION:G-Rh1 improves UUO-induced renal dysfunction and attenuates interstitial fibrosis, which is mediated via modulation of TGF-β₁-related pro-fibrogenic signaling pathway.     

Expression and modulation of connective tissue growth factor in renal interstitial fibrosis

CTGF, a member of the CCN family of immediate early genes, is a recently discovered profibrotic growth factor, which is involved in many pathophysiologic procedures. CTGF acts as a downstream effector of TGF-β acting on interstitial cells to enhance the progression of fibrotic renal diseases. It has been shown that CTGF gene expression can be induced or blocked by some kinds of cytokine and drugs. It is an interesting candidate target for future intervention strategies of renal interstitial fibrosis.     

Rhein attenuates bleomycin-induced rats pulmonary fibrosis through TGF-β1/Smad pathway by inhibiting miR-21 expression

AIM: To investigate the effect of rhein on bleomycin-induced pulmonary fibrosis and the expression of microRNA-21 (miR-21) and transforming growth factor-β1 (TGF-β1)/Smad signaling molecules in rats. METHODS: A single dose of bleomycin was intratracheal injected into the SD rats to induce pulmonary fibrosis. After injection of bleomycin, the rats were randomly divided into low-, medium-and high-dose rhein treatment groups and model group. The rats that were instilled with normal saline intratracheally served as control group. After the treatment for 28 d, the pulmonary pathologic changes were observed under microscope with hematoxylin-eosin staining. The lung coefficient and hydroxyproline content were also measured. The expression of miR-21 and the mRNA levels of TGF-β1 and Smad7 in the lung tissues were detected by real-time PCR. The protein levels of TGF-β1 and Smad7 were determined by Western blot. RESULTS: Rhein significantly attenuated the experimental alveolitis, pulmonary fibrosis, lung coefficient and hydroxyproline contents in the rats. Rhein obviously decreased the expression of miR-21,and the mRNA and protein levels of TGF-β1, but significantly increased the mRNA and protein levels of Smad7 in the lung tissues. CONCLUSION: Rhein effectively prevents bleomycin-induced pulmonary fibrosis by inhibiting the expression of miR-21 and promoting the expression of Smad7, thus regulating the TGF/Smad signaling pathway to decrease extracellular matrix deposition.     

Effect of imatinib on myocardial fibrosis in DOCA-salt hypertensive rats

AIM: To investigate the effects of imatinib (IMA), one of platelet-derived growth factor receptor (PDGFR) inhibitors, on myocardial fibrosis in deoxycorticosterone acetate (DOCA)-salt hypertensive rats. METHODS: Sixty male uninephrectomized SD rats were treated with 1% NaCl and 0.2% KCl in the drinking water for 4 weeks and assigned to 3 groups: vehicle control group (control group), DOCA treatment group (DOCA group), DOCA and IMA treatment group (DOCA+IMA group). Systolic blood pressure (SBP) was measured using the tail-cuff method. Myocardial tissue inflammation was analyzed by hematoxylin-eosin staining. Collagen volume fraction (CVF) and perivascular collagen area (PVCA) were analyzed by Sirius red staining. Ectodermal dysplasia-1 (ED-1) was analyzed by immunohistochemistry. Expression levels of platelet-derived growth factors (PDGF-A and PDGF-C), PDGF receptor α (PDGFRα) and phosphorylated PDGFRα (p-PDGFRα) were detected by Western blotting. RESULTS: SBP in DOCA group and DOCA+IMA group were signficantly higher than that in control group. No significant difference of SBP between DOCA group and DOCA+IM group was observed. In DOCA group, severe myocardial fibrosis was found, and CVF and PVCA were higher than those in control group. The differences of the CVF and PVCA between DOCA+IMA group and control group were detected, but the CVF and PVCA in DOCA+IMA group were significantly lower than those in DOCA group. Compared with control group, different degrees of myocardial tissue inflammation and monocyte/macrophage infiltration were observed in DOCA group and DOCA+IMA group. The expression levels of PDGF-A, PDGF-C and PDGFRα in DOCA group and DOCA+IMA group were much higher than those in control group, but the expression of p-PDGFRα in DOCA+IMA group were signficantly lower than that in DOCA group. CONCLUSION: Mineralocorticoid-induced myocardial fibrosis is related to cardiac tissue inflammatory response, excessive monocyte/macrophage infiltration and expressions of PDGF-A,PDGF-C and PDGFRα. Imatinib has an inhibitory effect on the myocardial fibrosis. The mechanisms may be associated with the inhibition of PDGFRα activity on the surface of fibroblast, thus interrupting PDGFs signaling-induced fibroblast division and proliferation.     

Inhibitory effect of metformin on alveolar epithelial-mesenchymal transition in lung tissue of rats with pulmonary fibrosis

AIM: To study the inhibitory effect of metformin on alveolar epithelial-mesenchymal transition (EMT) in rats with pulmonary fibrosis and the possible mechanism. METHODS: SD rats (n=48) were used, 12 of which were set up as normal control group, and 36 of which were induced by bleomycin (5 mg/kg) by tracheal instillation to establish pulmonary fibrosis. The pulmonary fibrosis rats were randomly divided into bleomycin group, low dose (100 mg/kg) of metformin group, and high dose (300 mg/kg) of metformin group. The rats in metformin groups were given the corresponding dose of metformin daily for 4 weeks. HE staining and Masson staining were used to observe the changes of lung histopathology and collagen deposition. Real-time PCR, Western blot and innunohistochemical staining were used to detect the mRNA and protein expression of α-smooth muscle actin (α-SMA), E-cadherin, vimentin, zonula occludens-1 (ZO-1), collagen I, collagen III and transforming growth factor-β₁ (TGF-β₁), and the protein phosphorylation levels of Smad2/3 and extracellular signal-regulated kinase 1/2 (ERK1/2) were also determined. RESULTS: Metformin up-regulated the expression of E-cadherin and ZO-1, down-regulated the expression of α-SMA, vimentin, collagen I and collagen III, and the protein phosphorylation levels of Smad2/3 and ERK1/2 were also decreased (P<0.05). CONCLUSION: Metformin inhibits alveolar EMT in the rats with pulmonary fibrosis, and its mechanism may be related to the inhibition of TGF-β₁ signal transduction pathway.     

Antagonistic effect of thalidomide on TGF-β1-induced change of CTGF gene promoter-binding proteins in HELF cells

AIM:To investigate the antagonistic effect of thalidomide (THD) on the activation of connective tissue growth factor (CTGF) gene promoter induced by transforming growth factor-β₁ (TGF-β₁) in human embryonic lung fibroblasts (HELF). METHODS:Luciferase reporter vector driven by CTGF gene promoter was used to detect the effects of TGF-β₁ and THD on the activity of CTGF gene promoter, and DNA pull-down assay with CTGF gene promoter as a probe was used to analyze the changes of CTGF promoter-binding proteins under different conditions. RESULTS:TGF-β₁ increased the activity of reporter driven by CTGF gene promoter (P<0.01), while THD significantly inhibited TGF-β₁-induced increase in the reporter activity dose-dependently (P<0.01). At the same time, THD had inhibitory effect on the TGF-β₁-induced change of CTGF gene promoter-binding proteins (P<0.05). CONCLUSION:Regulation of CTGF gene promoter-binding proteins takes part in TGF-β₁-induced activation of CTGF gene promoter, while THD has antagonistic effect on this process.     

Effect of leonurine on expression of miR-1 in rats with myocardial fibrosis induced by isoproterenol

AIM: To investigate effect of leonurine on the expression of microRNA-1 (miR-1) in rats with myocardial fibrosis induced by isoproterenol (ISO). METHODS: SD rats (n=10) were used as normal control group, and 80 rats were given ISO by intraperitoneal injection daily for 2 weeks to establish the model of myocardial fibrosis. The model rats were randomly divided into 5 groups:model group, low-dose (7.5 mg·kg^-1·d^-1) leonurine group, middle-dose (15 mg·kg^-1·d^-1) leonurine group, high-dose (30 mg·kg^-1·d^-1) leonurine group and p38 mitogen-activated protein kinase (p38 MAPK) inhibitor (0.3 mg·kg^-1·d^-1) group. After the treatment for 2 weeks, the ultrastructure of left ventricular myocardial tissues was observed under electron microscope. Masson staining was used to detect collagen fibrosis, and the expression of collagen I and collagen Ⅲ was determined by the method of immunohistochemistry. The contents of endothelin-1 (ET-1) and angiotensin Ⅱ (Ang Ⅱ) were measured by ELISA. The expression of miR-1 and ET-1 mRNA was detected by real-time PCR, and the protein expression of p38 MAPK, β-myosin heavy chain (MHC) and α-MHC was determined by Western blot. RESULTS: Compared with model group, the ultrastructure of left ventricular myocardial tissues in high-dose leonurine group was attenuated, and the expression of miR-1 and the protein expression of α-MHC in left ventricular myocardial tissues of high-dose leonurine group were increased (P<0.05). Collagen volume fraction, collagen I, collagen Ⅲ, the ratio of collagen Ⅰ/collagen Ⅲ, the contents of ET-1 and Ang Ⅱ, the mRNA expression of ET-1, and the protein expression of p38 MAPK and β-MHC in high-dose leonurine group were lower than those in model group (P<0.05). CONCLUSION: Leonurine attenuates myocardial fibrosis in the rats induced by ISO, and it is potentially associated with affecting the expression of miR-1, and inhibiting ET-1/p38 MAPK signaling pathway.     

Effect of Ginkgo biloba extract on the expression of connective tissue growth factor in the initiating stage of fibrosis in lungs of rats

AIM: To study the effect of Ginkgo biloba extract (GbE) on the expression of connective tissue growth factor (CTGF) in the initiating stage of pulmonary fibrosis of rats after administration of bleomycin (BLM).METHODS: The expression of CTGF in lungs was detected by Western blotting. The content of hydroxyproline was assayed by the method of chloramines T. The content of malondialdehyde (MDA) in plasma was investigated by colorimetry.RESULTS: On day 14 after administration of BLM, the contents of CTGF in lungs and MDA in plasma in BLM+NS group were higher than those in NS group, respectively (P<0.05; P<0.01). On day 30 after BLM, the contents of hydroxyproline in lungs and MDA in plasma in BLM+NS group were higher than those in NS group, respectively (both P<0.01). Treatment with GbE ameliorated the above changes induced by BLM. CONCLUSION: GbE ameliorates the up-regulation of CTGF in the initial stage of fibrosis in lungs of rats after administration of BLM. GbE prevents the hyperoxidative injury in lungs of rats after BLM, which might be one of mechanisms underling the effect of GbE on CTGF.     

Effect of bFGF on promoting angiogenesis in infarct area and improving myocardial fibrosis in mouse myocardial infarction model

AIM: To investigate the protective effect of basic fibroblast growth factor (bFGF) on the heart of mice with myocardial infarction and its mechanism. METHODS: The model of myocardial infarction was established by the ligation of left anterior descending artery of C57/B6 mice (8~12 weeks old) after lateral thoracotomy. The mice were divided into sham operation group, myocardial infarction group and bFGF administration group. bFGF at 0.5 μg was intraperitoneally injected on alternate days after myocardial infarction for 7 d. Cardiac Doppler ultrasonography was used to detect cardiac function after myocardial infarction for 28 d, and left ventricular end-diastolic diameter, left ventricular end-systolic diameter, left ventricular ejection fraction and left ventricular shortening fraction (LVFS) were used to evaluate cardiac function. After myocardial infarction for 28 d, the mice were sacrificed and the hearts were collected for preparing pathological sections. The degrees of myocardial fibrosis and angiogenesis in the myocardial infarction area were observed. Western blot was used to detect the indicators of angiogenesis. RESULTS: The results of Masson staining showed that bFGF administration significantly reduced myocardial fibrosis at 28 d after myocardial infarction. Cardiac ultrasound data showed that cardiac functions in myocardial infarction group were poorer than those in sham group, and bFGF administration significantly improved cardiac functions. Immunofluorescence staining showed that neovascularization in myocardial infarction area of bFGF administration group was more than that in myocardial infarction group. The results of Western blot showed that bFGF activated AKT/HIF-1α/VEGF signaling pathway. CONCLUSION: Intraperitoneal injection of bFGF reduces myocardial fibrosis and improves cardiac function in myocardial infarction mice. bFGF may promote angiogenesis by activating AKT/HIF-1α/VEGF signaling pathway.     

Imatinib attenuates DOCA/salt-induced rat myocardial fibrosis via PDGFs/PDGFRs signaling pathway

AIM：To investigate the role of imatinib (IMA) in reducing myocardial fibrosis and its relationship with platelet-derived growth factors (PDGFs)/PDGF receptors (PDGFRs) signaling pathway in deoxycorticosterone acetate (DOCA)/salt-induced hypertension in rats. METHODS:Sixty male uninephrectomized SD rats were treated with 1% NaCl and 0.2% KCl in the drinking water for 4 weeks and randomly divided into vehicle control (CON) group, DOCA treatment (DOCA) group and DOCA plus IMA treatment (DOCA+IMA) group. Systolic blood pressure (SBP) was mea-sured weekly using the tail-cuff method. The extent of myocardial interstitial fibrosis and the ratio of perivascular collagen area/vascular area (PVCA/VA) were analyzed by picro-Sirius red staining. The mRNA expression of fibroblast-specific protein 1 (FSP-1), α-smooth muscle actin (α-SMA), procollagenⅠ, procollagenⅢ, PDGFRα and PDGFRβ in the myocardium was measured by real-time PCR. The protein levels of PDGFRα, PDGFRβ, p-PDGFRβ, vimentin and α-SMA were analyzed by immunohistochemical staining. The cell types of PDGFRα and PDGFRβ localizations were examined by immunofluorescence method. RESULTS：(1) SBP in DOCA and DOCA+IMA groups was higher than that in CON group on day 14 and day 28, and no difference of SBP between DOCA group and DOCA+IMA group was found. The extent of myocardial interstitial fibrosis and the ratio of PVCA/VA in DCOA group were higher than those in CON group, and those in DOCA+IMA group were significantly lower than those in DOCA group on day 28. (2) Compared with CON group, the expression of PDGFRα and PDGFRβ in DOCA group was increased on day 14. Compared with DOCA group, the expression of PDGFRα and PDGFRβ in DOCA+IMA group was attenuated. Compared with CON group, the expression of PDGFRβ, p-PDGFRβ, FSP-1, α-SMA, procollagenⅠand procollagen Ⅲ in DOCA group was increased significantly on day 28, but the expression of PDGFRα was not increased significantly, and the expression of PDGFRβ was higher than PDGFRα. Compared with DOCA group, the expression of PDGFRβ, p-PDGFRβ, FSP-1, α-SMA, procollagenⅠand procollagen Ⅲ in DOCA+IMA group was attenuated on day 28. (3) The fibroblasts in the myocardial interstitium of CON group were mainly vimentin positive, but not α-SMA positive on day 28,while the number of α-SMA-positive spindle-shaped cells (myofibroblasts) increased significantly in DOCA group. Compared with DOCA group, the number of myofibroblasts in DOCA+IMA group was attenuated on day 28. (4) PDGFRα was located in fibroblasts and myofibroblasts but not in vascular smooth muscle cells (VSMCs). PDGFRβ was not only located in myofibroblasts but also in VSMCs. CONCLUSION:PDGFRα takes effect in the early phase of myocardial fibrosis in the DOCA/salt hypertensive rats, while PDGFRβ takes effect in the entire process. PDGFRα and PDGFRβ are expressed in cardiac fibroblasts and myofibroblasts. Imatinib reduces the proliferation and transformation of fibroblasts, thereby attenuating myocardial fibrosis by inhibiting PDGFs/PDGFRs signaling pathway.     

Role of miR-219 and TGFBR2 in pathogenesis of renal fibrosis

AIM:To study the role of microRNA-219 (miR-219) in regulation of transforming growth factor-β receptor type 2 (TGFBR2) in renal fibrosis. METHODS:The renal fibrosis patients (n=70) were selected in this stu-dy, and 20 cases of healthy people were selected as control group. RT-qPCR was used to detect the expression of miR-219 in the serum of the patients with renal fibrosis and control group, and the expression of miR-219 in NRK49F cells after stimulation with angiotensin Ⅱ(AngⅡ) was detected. The protein expression of α-smooth muscle actin (α-SMA) in the NRK49F cells transfected with miR-219 mimics after stimulation with AngⅡ was determined by Western blot. The potential target gene TGFBR2 of miR-219 was screened and verified by the method of luciferase reporter gene. RT-qPCR and Western blot were used to detected the effect of miR-219 mimics on the expression of TGFBR2 at mRNA and protein levels, and the mRNA expression of α-SMA, connective tissue growth factor (CTGF), type I collagen α1 (COL1A1) and COL3A1 in the NRK49F cells was also detected, respectively. The unilateral ureteral occlusion (UUO) mouse model was established and the expression of miR-219 in the renal tissue was monitored. The morphological change of renal fibrosis was observed in the UUO mice after injection of miR-219, and the mRNA expression levels of COL1A1 and COL3A1 were detected. RESULTS:The expression level of miR-219 in the patients with renal fibrosis was significantly lower than that in control group, and the expression of miR-219 in the UUO mice was decreased significantly (P<0.01). The expression level of miR-219 was significantly decreased in the NRK49F cells after AngⅡ stimulation, and miR-219 mimics inhibited the protein expression of α-SMA(P<0.01). miR-219 mimics had a targeted regulatory effect on TGFBR2 gene, which inhibited the mRNA and protein expression of TGFBR2. miR-219 mimics inhibited the mRNA expression of α-SMA, CTGF, COL1A1 and COL3A1. miR-219 also down-regulated the mRNA expression of COL1A1 and COL3A1 in the UUO mice and inhibited the process of renal fibrosis. CONCLUSION:miR-219 inhibits the development of renal fibrosis by inhibiting the expression of TGFBR2, which may become a new target for the diagnosis and treatment of renal fibrosis.     

Effects of Danshensu on TGF-β1/Smads signaling pathways in rats with pulmonary fibrosis

AIM: To study the preventive and curative roles of Danshensu (DA) in bleomycin (BLM)-induced pulmonary fibrosis in rats. METHODS: Pulmonary fibrosis was induced in SD rats by intratracheal instillation of BLM. The rats were intraperitoneally injected with dexamethasone (1 mg·kg-1·d-1, DXM group), DA (15 mg·kg-1·d-1, DA group), or physiological saline (2 mL·d-1, BLM group). Normal controls (NC group) received physiological saline both intratracheally and intraperitoneally. At the 28th day after modeling, the histological changes of the lungs were evaluated by hematoxylin-eosin (HE) and Masson’s trichrome staining. The protein levels of α-smooth muscle actin (α-SMA) in the lung tissues were detected by the method of immunohistochemistry. The mRNA expression of transforming growth factor beta 1 (TGF-β1), Smad3 and Smad7 was assessed by real-time fluorescence quantitative PCR. RESULTS: Compared with BLM group, the degree of inflammation and fibrosis of the lung in DA group was obviously reduced, and so was the expression of α-SMA in the lung tissues. The mRNA expression of TGF-β1 and Smad3 in the lung tissues of the rats decreased and the mRNA expression of Smad7 increased. CONCLUSION: DA alleviates BLM-induced pulmonary fibrosis in rats in the early stage by inhibiting the expression of TGF-β1/Smad3 and stimulating the expression of Smad7 in the lung tissues.     

Over-expressed calreticulin interacts with integrin-α5 and calcineurin system to induce atrial remodeling in patients with atrial fibrillation and valvular disease

AIM: To determine whether calreticulin over-expression contributes to atrial fibrosis in the patients with atrial fibrillation (AF) and valvular heart disease (VHD).METHODS: Right and left atrial specimens were obtained from 78 patients undergoing valve replacement surgery. The patients were divided into sinus rhythm (SR) group, paroxysmal AF (PaAF) group and persistent AF (PeAF, AF lasting >6 months) group. The protein expression of calreticulin, integrin-α5, and transforming growth factor-β1 (TGF-β1) was measured. Immunoprecipitation was also performed to determine whether calreticulin interacted with either calcineurin B or integrin-α5. RESULTS: The protein expression of calreticulin, integrin-α5 and TGF-β1 was increased in AF groups, especially in the left atrium of the patients with mitral valve disease as compared with SR group. Calreticulin interacted with both calcineurin B and integrin-α5. The expression level of integrin-α5 was significantly correlated with the expression level of TGF-β1, while the expression level of calreticulin was correlated with that of integrin-α5 and TGF-β1. Under similar classification of the cardiac function, the expression level of calreticulin in PeAF group was higher than that in SR group. CONCLUSION: The expression of calreticulin, integrin-α5, and TGF-β1 is increased in the atrial tissues of the AF patients and is related to the AF type, suggesting that calreticulin is involved in atrial remodeling in AF and VHD patients.     

Intervention effects of apyrase on silica-induced pulmonary fibrosis in mice

AIM: To investigate the effect of apyrase on the experimental silicosis. METHODS: C57BL/6 male mice were randomly divided into control group, silica treatment group, silica+apyrase group and silica+NS group. A mouse model of lung fibrosis was induced by crystalline silica particles (50 mg/kg, via oropharyngeal instillation), and were sacrificed at 3 h, 7 d, 14 d and 28 d. Apyrase was delivered by oropharyngeal aspiration at the same time and 4 h after silica challenge. The lung indexes were calculated and the concentration of ATP was detected by bioluminescent assay. The mRNA expression levels of collagen type Ⅰ(Col Ⅰ), collagen type Ⅲ (Col Ⅲ) and transforming growth factor β1 (TGF-β1) were examined by real-time PCR. The protein levels of TGF-β1 in bronchoalveolar lavage fluid were measured by ELISA. RESULTS: The elevated lung index and collagen levels showed that silicosis model was established successfully. Compared with silica group, apyrase treatment significantly alleviated silica-induced inflammation, reduced inflammation score on day 7, and decreased the lung index, collagen volume fraction and the mRNA expression of Col Ⅰand Col Ⅲ on day 28. Treatment with apyrase effectively down-regulated the mRNA levels of TGF-β1 in the lung tissues and TGF-β1 protein levels in bronchoalveolar lavage fluid on day 7.CONCLUSION: Apyrase attenuates the pulmonary inflammation and fibrosis of silicosis, which may be related with down-regulation of ATP and TGF-β1 in the lung tissues.     

Effects of long-term cigarette smoke exposure on pulmonary vascular remodeling and TGF-β1 expression in rat pulmonary vessels

AIM: To observe the effects of long-term cigarette smoke exposure on pulmonary vascular remodeling and the protein expression of transforming growth factor-β1(TGF-β1) in the rats, and to explore the mechanism.METHODS: SD rats(n=36) were randomly divided into control group, 2-week smoke exposure(S-2W) group and 12-week smoke exposure(S-12W) groups. HE staining and α-smooth muscle actin staining were performed to observe the pulmonary vascular remodeling.The protein expression of proliferating cell nuclear antigen(PCNA) and TGF-β1 in the pulmonary arteries was determined by the method of immunohistochemistry. The mRNA expression of TGF-β1 in the pulmonary arteries was evaluated by RT-qPCR.RESULTS: Compared with control group, ratio of pulmonary vessel wall thickness to vessel diameter(WT%) and percentage of muscularized vessels were significantly increased in S-2W group and S-12W group(P<0.01). Significant increases in the protein expression of PCNA and TGF-β1 in smoke exposure groups were observed compared with control group. There was significant difference between 2 model groups(P<0.01). Compared with control group, the mRNA expression of TGF-β1 in pulmonary artery walls obviously increased in smoke exposure groups. There was significantly difference between S-2W and S-12W groups(P<0.05). The mRNA expression of TGF-β1 was positively correlated with pulmonary vascular muscularization, WT% and the protein expression of PCNA. CONCLUSION: Long-term cigarette smoke exposure results in pulmonary artery remodeling in rats. The possible mechanism is that cigarette smoking exposure induces the over-expression of TGF-β1 at mRNA level in pulmonary vessels and promotes the proliferation of pulmonary vascular smooth muscle cells in rats.