Tachykinins in Monocrotaline-induced Pulmonary Hypertension in Wistar Rats

• Main Authors: Mei-Jung Chen, 陳玫蓉
• Other Authors: Yih-Loong Lai
• Format: Others
• Language: zh-TW
• Published: 2003
• Online Access: http://ndltd.ncl.edu.tw/handle/23314890496177420968

博士 === 國立臺灣大學 === 生理學研究所 === 91 === The purpose of this study was to explore the effect of substance P on monocrotaline (MCT)-induced pulmonary hypertension (PH). The pulmonary arterial blood pressure (Ppa) and right ventricular hypertrophy are two main characteristics for the MCT-induced PH. The metabolite of MCT causes injury of arterial wall, which in turn induces remodeling of vessel, increase in pulmonary vascular resistance, and right ventricular hypertrophy. It is well known that tachykinins, the neruotransmitters of the nonadrenergic-noncholinergic nervous system, have proliferative and pro-inflammative effects. It was proposed that tachykinins (mainly substance P and neurokinin A in mammals) promote the development of MCT-induced PH. Because substance P accounts for over 95% of tachykinins in mammals, it was focused mainly in the present thesis. The first study was designed to evaluate whether antioxidants and depletion of substance P attenuate the PH. The study was carreid out in three parts. In the first part, using 1,3-dimethyl-2-thiourea (DMTU) and hexa(sulfobutyl)fullerenes (FC4S) as antioxidants. Wistar rats were divided into six groups: control; MCT; capsaicin+MCT; MCT+DMTU-1; MCT+DMTU-2; and MCT+FC4S. Rats in the control group were kept in normoxia condition and received saline injection; MCT rats received a subcutaneous injection of MCT (60 mg/kg). Animals in the capsaicin+MCT group received capsaicin pretreatment to deplete substance P, then followed by MCT treatment. The antioxidant DMTU was given in the early (DMTU-1) and the late (DMTU-2) periods post MCT. The rats in the last group received another antioxidant FC4S treatment. Functional study was carried out three weeks after MCT treatment. Compared to the control group, MCT treatment caused increases in pulmonary arterial blood pressure and blood vessel wall thickness, and right ventricular hypertrophy. Depletion of substance P or antioxidants attenuated the MCT-induced alterations. The second part was to test if a change in pulmonary vascular response to substance P occurs following MCT treatment. Animals were divided into two groups: control and MCT-2wk. Two weeks after MCT treatment, the increase in pulmonary arterial blood pressure caused by substance P was exaggerated in the MCT-2wk group compared to that in the control group. For the third part, reactive oxygen species were measured by luminol-enhanced chemiluminescence counts. An increase trend in reactive oxygen species was detected during the early period of the MCT pneumotoxicity. For the second study, it was designed to evaluate whether substance P and reactive oxygen species play a role in perinatal hypoxia-exaggerated, monocrotaline-induced pulmonary hypertension. The first part was carried out to distinguish if a difference in MCT-induced PH occurs between normoxia and prenatal hypoxia rats. To breed prenatal hypoxia rats, pregnant Wistar rats were exposed to intermittent hypoxia (from 5 PM to 8 AM) by placing in a decompression chamber with a barometric pressure of 380 Torr for 17 days, from the 5th day to the 21st day of the pregnancy. The infants were born from these pregnant rats and grew up in normoxia. Three weeks after MCT treatment, increases in pulmonary arterial blood pressure and lung tissue substance P level were found in prenatal hypoxia rats compared to those in normoxia rats. However, a right ventricular hypertrophy was failed to detect in prenatal hypoxia rats. The second part was performed to evaluate the effects of depletion of substance P and antioxidants on the prenatal hypoxia-exaggerated, MCT-induced PH. Prenatal hypoxia rats were divided into six groups: control, MCT, capsaicin+MCT, MCT+DMTU-1, MCT+DMTU-2, and MCT+FC4S. The animal preparation in each group was the same as that described above. MCT-induced increase in pulmonary arterial blood pressure was attenuated by either capsaicin pretreatment or antioxidants. These results suggest that both substance P and reactive oxygen species are involved in the prenatal hypoxia-augmented, MCT-induced PH. In the third study, the hypothesis that the up-regulation of preprotachykinin (PPT) gene might occur during the development of MCT-induced PH, accompanying with an increase in the lung tissue substance P level, was tested. Thus interference with substance P synthesis or blockage of substance P receptors [neurokinin1 (NK1) and NK2] may attenuate the development of PH. To interfere the PPT gene expression, double-stranded PPT RNA was used [RNA interference (RNAi)] in this study. Animals were divided into four groups in part 1: control; MCT; RNAi+MCT; and solvent+MCT. One day prior to MCT, bilateral nodose ganglia were microinjected with ds PPT RNA in rats of the RNAi+MCT group, whereas solvent in the solvent+MCT group. Functional study was carried out two weeks after MCT. MCT induced right ventricular hypertrophy, as well as increases in pulmonary arterial pressure, PPT mRNA (nodose ganglia and lung tissue), and lung tissue substance P level. All the MCT-induced alterations were attenuated by RNAi, indicating that the up-regulation of PPT mRNA leads to the increase in lung tissue substance P. The second part was designed to evaluate the effects of NK1 receptor antagonist CP-96345 or NK2 receptor antagonist SR-48968 on the MCT-induced PH. There were seven groups of animals: control; MCT; MCT+CP-96345-3.4; MCT+CP-96345-10; MCT+CP-96344-10; MCT+SR-48968; and MCT+SR-48965. Started from one day post MCT, MCT-treated rats received daily injection of CP-96345 (3.4 or 10 mg/kg, ip) or its inactive enantiomer CP-96344 (10 mg/kg, ip). SR-48968 (3 mg/kg, ip) or its inactive enantiomer SR-48965 (3 mg/kg, ip) was injected to MCT-treated rats every other day started one day post MCT. Functional study was carried out three weeks after MCT. MCT-induced increase in pulmonary arterial pressure and right ventricular hypertrophy were attenuated by NK receptor antagonists, indicating the useful treatment of NK receptor antagonists on the PH. It was concluded that an increase in tachykinins was demonstrated in MCT-induced PH, and up-regulation of PPT gene and increase in reactive oxygen species were the underlying mechanism. Depletion of substance P, antioxidant treatment, and NK receptor antagonists attenuated PH.