| BackgroundPulmonary hypertension (PH) is a common pathophysiologic syndrome caused by a variety of diseases and characterized by a progressive increase of pulmonary vascular resistance and pulmonary artery pressure that finally causes right ventricular failure and premature death. Pulmonary arterial hypertension (PAH) is a disease of the small pulmonary arteries that is characterized by vascular remodeling. Characteristic features of this vascular remodeling include vessel wall thickening as a result of resident vessel wall cell proliferation, migration, and excessive deposition of extracellular matrix. In spite of recent advancements in the treatment of PAH, successful control has yet to be accomplished. The rennin angiotensin system (RAS) has been implicated in the pathogenesis of pulmonary vascular remodeling and PAH in a number of studies. Telmisartan is an angiotensin II type 1 receptor (AT1R) blocker, currently was used to treat patients with hypertension. Moreover, it has been associated with beneficial effects on anti-inflammation, vascular function, cardiac remodeling and renal function. However, little is known about the effects of telmisartan on PAH. To determine the effect of telmisartan on monocrotaline-induced PAH and its possible mechanism, the following study was carried out.Objective1. To investigate the interventional effects of telmisartan on monocrotaline-induced PAH in rats.2. To further explore the possible mechanisms of the protective effects of telmisartan on monocrotaline-induced PAH in rats.Methods1.32 male Sprague-Dawley rats were randomly divided into four groups:Group C (n=8) rats were injected subcutaneously with normal saline; Group P (n8) rats were injected subcutaneously with monocrotaline (60mg/kg) and received with normal saline by daily oral gavage for 3 weeks; Group PTh (n=8) rats were injected subcutaneously with monocrotaline (60mg/kg) and received telmisartan (10mg/kg) by daily oral gavage for 3 weeks; Group PTl (n=8) rats were injected subcutaneously with monocrotaline (60mg/kg) and received telmisartan (5mg/kg) by daily oral gavage for 3 weeks. 2. Right ventricular systolic pressure (RVSP) and mean pulmonary arterial pressure (mPAP) were measured by right heart catheter after 3 weeks. All rats were killed by exanguination and their hearts and lungs were harvested. The right ventriclular hypertrophy index (RVHI), percentage of small pulmonary arteries media thickness (WT%), muscularization degree of pulmonary vessels and score of pulmonary vascular inflammation were evaluated.3. ELISA kits were used to evaluate the proinflammatory cytokines (IL-6, TNF-a and MCP-1) in pulmonary homogenate at protein level.4. The protein expression of MMP-2 and MMP-9 at pulmonary vascular was examined by immunohistochemistry. The mRNA of MMP-2, MMP-9 and TIMP-1 in the lung were measured by real-time PCR. The enzymic activity of MMP-2 and MMP-9 were detected in gelatin zymography.5. The protein expression of ACE2 on pulmonary vascular was examined by immunohistochemistry. The mRNA of ACE2 of the lung was measured by real-time PCR.Results1. Effects of telmisartan on hemodynamic parameter, right ventricular hypertrophy, pulmonary vascular remodeling and perivascular inflammation:The mPAP, RVSP and RVHI of group C were 17.13±3.30mmHg, 40.33±5.77mmHg and 0.25±0.02. These indexes of group P were increased significantly compared with group C (37.00±4.98mmHg,67.04±4.87mmHg and 0.40±0.02, P<0.01), and attenuated significantly with telmisartan administration (24.75±4.69mmHg, 52.30±8.37mmHg and 0.29±0.01 of group PTh; 26.27±5.46mmHg,53.25±8.65mmHg and 0.34±0.02 of group PTl; P<0.01). No significant differences in mPAP and RVSP were observed in two telmisartan dosage groups (P>0.05), but RVHI of group PTh was lower than group PT1 with statistical significance (0.29±0.01 vs 0.34±0.02, P<0.01).Prominent medial wall hypertrophy, smooth muscle proliferation in muscular pulmonary arteries and muscularized arterioles were evident from rats treated with MCT (group P). Compared with group C, percentage of small pulmonary arteries media thickness, muscularization of acinus pulmonary arterioles and a-smooth muscle actin expression (IOD/Area) of pulmonary arterioles of group P increased significantly (45.52±5.48% vs 8.91±2.30%,67.55±2.95% vs 19.92±2.27%,0.49±0.03 vs 0.22±0.04; P<0.01). These increases were also prevented notably with telmisartan intervention (13.83±1.83%,50.83±3.12% and 0.34±0.02 of group PTh; 20.84±15.21%,53.22±1.97% and 0.36±0.02 of group PTl; P<0.01).No significant differences in WT% and muscularization were observed in two telmisartan dosage groups (P>0.05), but IOD/Area of group PTh was lower than group PTl with statistical significance (P<0.05).Macrophages significantly increased in alveoli and primarily mononuclear cells infiltrated around the arterioles in group P. The perivascular inflammation score of group P was remarkably higher than that of group C with statistical significance (3.24±0.41 vs 0.40±0.16, P<0.01), and was reduced significantly with telmisartan administration (1.00±0.19 of group PTh; 1.46±0.31 of group PTl; P<0.01). The score of group PTh was lower than group PTl with statistical significance (P<0.05).2. Effects of telmisartan on pulmonary homogenate IL-6, TNF-a and MCP-1 of rats injected with MCT:The IL-6, TNF-a and MCP-1 level of group P were significantly higher than that of group C with statistical significance (459.40±94.77pg/ml vs 64.97±17.65pg/ml, 436.46±62.20pg/ml vs 56.51±14.92pg/ml,6265.53±2014.83pg/ml vs 65.18±23.37pg/ml; P<0.01). Also, MCT-induced increases in proinflammatory cytokines were significantly attenuated by telmisartan intervention (118.87±25.80pg/ml,242.61±37.52pg/ml and 1094.39±577.75pg/ml of group PTh; 155.87±50.64pg/ml,278.69±42.35pg/ml and 1288.57±593.14pg/ml of group PTl; P＜0.01). No significant differences in these proinflammatory cytokines were observed in two telmisartan dosage groups (P>0.05).3. Effects of telmisartan on expression of MMP-2, MMP-9 and TIMP-1 mRNA in the lungs and MMP-2, MMP-9 protein and activity:Immunohistochemistry for MMP-2 and MMP-9 in pulmonary arterioles of group P revealed that positive staining was localized mainly in the tunica intima and adventitia, which showed a notable increase as compared to group C. Moreover, this expression was decreased after administration of telmisartan. The MMP-2, MMP-9 and TIMP-1 mRNA of group P were significantly higher than that of group C with statistical significance (3.30±0.39 vs 0.96±0.08,7.07±0.54 vs 0.96±0.06,3.52±0.44 vs 1.02±0.14;P<0.01). These increases were also reduced notably with telmisartan intervention (1.97±0.24, 3.65±0.70 and 2.22±0.35 of group PTh; 2.43±0.21,4.91±0.68 and 2.54±0.43 of group PTl; P<0.01). No significant difference in TIMP-1 mRNA was observed in two telmisartan dosage groups (P＞0.05), but MMP-2 and MMP-9 mRNA of group PTh was lower than that of group PTl with statistical significance (P＜0.05).MMP-2 and MMP-9 enzymatic activity in group P was significantly higher than that of group C (54893.89±8480.12 vs 40669.68±4978.39, P<0.05; 23864.69±5121.83 vs 9511.13±3429.93, P＜0.01). MMP-2 activity in group PTh and PTl (52401.41±6234.28 and 51651.18±5289.70, respectively) was observed no significant differences compared with that of group P (P<0.05). In contrast, MMP-9 activity was significantly lower in two telmisartan dosage groups compared with that of group P (14244.99±2601.82 of group PTh, P<0.01; 14896.96±3625.29 of group PTl, P<0.05). No significant differences in MMP-2 and MMP-9 activity were observed in two telmisartan dosage groups (P>0.05).4. Effects of telmisartan on expression of ACE2 mRNA or protein in the lung:In pulmonary arterioles ACE2 is preferentially localized in the endothelial layer. Immunohistochemistry results showed a notable decrease of ACE2 positive staining in group P as compared to that of control rats, and its expression is increased after administration of telmisartan. The ACE2 mRNA of group P were significantly lower than that of group C with statistical significance (0.56±0.09 vs 1.04±0.14, P＜0.01), and this decrease was reversed notably with telmisartan intervention (2.08±0.40 of group PTh, 1.65±0.44 of group PTl; P＜0.01), furthermore group PTh was higher than group PTl with statistical significance (P＜0.05).Conclusion1. Telmisartan can attenuate MCT-induced pulmonary vascular remodeling and PAH.2. The mechanisms of protective effects partly by inhibiting the pervascular inflammation and proinflammatory cytokines in lung.3. Modulating the expression and activity of MMPs leads to amelioration of extracellular matrix remodeling was probably related with the protective effects of telmisartan.4. Telmisartan may be therapeutically useful in MCT-induced pulmonary vascular remodeling and PAH and ACE2 may be involved as part of its mechanisms.
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