| Purpose: To analysis the potentail role for apoptosis related factors in vascular remodeling, such as TNF-??bax?BCL-xl,and provide a new direction for the study of the patients with chronic thromboembolic pulmonary hypertension(CTEPH), we construct CTEPH of rats model in autologous blood clots injection method.Methods: After the autologous blood clots were made, we repeatedly embolized pulmonary artery with the clots through the jugular vein injection to establish CTEPH experimental animal model of rat. We also measured mean pulmonary artery pressure(m PAP), and observed pulmonary artery changes by HE stain. The plasma level of TNF-? were measured with ELISA. The expressions of m RNA of TNF-?, bax, BCL- xl in pulmonary artery were tested by RT-PCR; The expressions of protein of TNF-?, bax,BCL-xl in pulmonary artery were tested by western blot. The expressions of TNF-?, bax,BCL-xl in pulmonary artery were tested by immunohistochemistry.Results:(1) The m PAP in the 1-week, 2-week, and 4-week subgroups in the experimental group increased significantly compared with the sham operation group(20.33±2.56 mm Hg vs 15.21±1.54 mm Hg, 23.12±2.56 mm Hg vs 14.93±1.43 mm Hg,24.96±1.58 mm Hg vs 15.4±1.41 mm Hg, P<0.05, respectively). The WA/TA ratio in the1-week, 2-week, and 4-week subgroups in the experimental group increased significantly compared with the sham operation group(41.86±3.42 vs 30.16±2.98, 46.33±3.23 vs 30.16±2.42, 52.18±3.21 vs 29.71±2.18, P<0.05, respectively). In the experimental group, m PAP and WA/TA, compared the 1-week subgroup with 2-week subgroup, the 1-week subgroup with 4-week subgroup, altered significantly(P < 0.05).The m PAP had no significant difference between 4-week and 2-week subgroups (P>0.05). The WA/TA ratio had statistically significant difference between 4-week and2-week subgroups(P < 0.05).(2) The plasma TNF-? concentration in the 1-week, 2-week, and 4-week subgroups in the experimental group increased significantly compared with the sham operation group(126.58±21.44 pg/ml vs 46.91±1.44pg/ml, P<0.05;122.30±35.29 pg/ml vs 46.81 ± 1.43pg/ml, P <0.05;126.52 ± 14.89pg/ml vs 46.43 ± 1.86pg/ml,P <0.05, respectively).(3) The TNF-? m RNA expressed in pulmonary artery in the 1-week, 2-week, and4-week subgroups of experimental group increased significantly compared with the sham operation group(0.77±0.14 vs 0.31±0.07, P<0.05; 0.96±0.08 vs 0.33±0.06,P<0.05; 0.64±0.12 vs 0.35±0.06, P<0.05, respectively).The Bax m RNA expressed in pulmonary artery in the 1-week, 2-week, and 4-week subgroups in the experimental group increased significantly compared with the sham operation group(0.77±0.07 vs0.52±0.09, P<0.05; 0.93±0.06 vs 0.55±0.1, P<0.05; 0.92±0.08 vs 0.57±0.06, P<0.05,respectively).The Bcl-xl m RNA expressed in pulmonary artery in the 1-week,2-week,and 4-week subgroups in the experimental group increased significantly compared with the sham operation group(0.43±0.07 vs 0.84±0.12, P<0.05; 0.52±0.06 vs 0.84 ± 0.14, P < 0.05; 0.52 ± 0.07 vs 0.82 ± 0.11, P<0.05, respectively).(4) The TNF-? proteins expressed in pulmonary artery in the 1-week, 2-week, and4-week subgroups of experimental group increased significantly compared with the sham operation group(1.62±0.08 vs 0.85±0.12, P<0.05;1.85±0.08 vs 0.89±0.13,P<0.05;1.37±0.12 vs 0.91±0.15,P<0.05, respectively); The Bax proteins in the 1-week,2-week, and 4-week subgroups in the experimental group increased significantly compared with the sham operation group(1.52±0.06 vs 0.96±0.1, P<0.05; 1.80±0.08 vs0.98±0.13, P<0.05; 1.94 ±0.06 vs 0.97±0.16, P<0.05,respectively); The Bcl-xl proteins in the 1-week, 2-week, and 4-week subgroups of experimental group increased significantly compared with the sham operation group(1.00±0.09 vs 1.79±0.06, P<0.05;0.96±0.06 vs 1.75 ± 0.05, P<0.05; 0.76 ± 0.07 vs 1.76 ± 0.06, P<0.05, respectively).(5) Immunohistochemical results showed that TNF-?, Bax antigens were expressed in pulmonary artery endothelial cells. When comparing the experimental group with sham-operated group, dyeing was obviously higher and expression level was raised. The Bcl-xl antigens dyeing is abated compared the experimental group with sham-operated group. In the experimental group, among the 1-week group, 2-week group and 4-week group, TNF-?, Bax antigens gradually dyeing deepened in the pulmonary artery endothelial cells with the extension of time and higher expression. while the Bcl-xl dyeing is weakened and expression is lowered.(6) Correlation analysis: The expression of pulmonary artery TNF-? protein was positively related with m PAP(r=0.605,P<0.01), and with WA/TA(r=0.629,P<0.01);The expression of serum TNF-? was positively related with that of pulmonary artery Bax protein(r=0.754,P<0.01), while negatively correlated with the pulmonary artery Bcl–xl protein(r=-0.831, P<0.01).The expression of serum TNF-? was positively related with that of pulmonary artery TNF-? protein(r=0.721,P<0.01); MPAP and the expression of pulmonary artery Bax protein were positively correlated(r=0.840,P<0.01),while negatively correlated with the expression of pulmonary artery Bcl-xl protein(r=-0.841,P<0.01).WA/TA and the expression of pulmonary artery Bax protein were in positively correlation with each other(r=0.896, P<0.01),while negatively correlated with the pulmonary artery Bcl–xl protein(r=-0.897,P<0.01).Conclusion:(1) By embolizing autologous blood clots repeatedly and injecting anticoagulants tranexamic acid, a rat model of chronic thromboembolic pulmonary hypertension can be established and provided for CTEPH pathological and pharmacological experimental study.(2) In a CTEPH rat model, thrombosis induced higher expression of TNF-? in pulmonary artery endothelial cells. There are significant positive correlations between TNF-? and pulmonary arterial pressure, vascular remodeling, and apoptosis. Therefore,the TNF-? may play an important role in vascular remodeling of CTEPH, and lead to pulmonary hypertesion. (3)The expression of TNF-? in plasma is in accordance with the level of TNF-? in the pulmonary artery. So it may be a relatively simple method to examine CTEPH patient's condition.(4) We detected that the expression of Bax is increased and the Bcl-xl is reduced. The apoptosis of pulmonary artery endothelial cell enhanced obviously, and its expression obviously positive correlation with TNF-?, and apoptosis is positively correlated with pulmonary hypertension and vascular remodeling. Apoptosis plays an important role in the process of vascular remodeling and raised pulmonary artery pressure in CTEPH.The TNF-? may regulate the apoptosis of pulmonary artery endothelial cells, promote vascular remodeling, finally cause the pulmonary hypertesion. |
PDF Full Text Request