| Pulmonary hypertension syndrome (PHS) of broilers is also called ascites syndrome in broiler or heart failure syndrome (HFS) in broiler. The term "ascites" actually refers to the fluid accumulated in the abdominal cavity as a consequence of heart failure. A frequent occurrence of ascites has been a difficult problem detrimental to the poultry industry around the world. Approximately 4% of all broilers die from pulmonary hypertension syndrome, causing a loss estimated at $1 billion annually. General agreement exists regarding the reasonable etiology of ascites irresoluble. Now most of the investigators focused on pulmonary hypertension because they thought it was the center link of this disease. Endothelin-1 (ET-1) is a potent vasoconstrictive peptide that was isolated initially from the conditioned medium of cultured endothelial cells. It is a pathogenic mediator with a number of deleterious effects, including vasoconstriction, fibrosis, vascular hypertrophy, and inflammation. Atrial natriuretic peptide, angiotensin â…¡ and nitric oxide also takes an important role in cardiovascular system. In order to illuminate the etiopathogenesis of pulmonary hypertension syndrome in broiler from the point of endothelin-1, natural pulmonary hypertension syndrome broiler cases were got through using hyperalimentation dietary to feed a great deal of commercial broiler chickens. Radioimmunoassay was used to detect endothelin-1(ET-1), ANP and Aâ…¡ in plasma, heart, liver, lung and kidney of different degree ascites broilers and normal broilers at different weeks of age. The AHI and mPAP of 3 to 6 weeks were measured as well. At the same time, nitrate reductase method was used to detect the level of nitric oxide (NO) and nitric oxide synthase (NOS) in plasma of different degree ascites broilers at the age of 3,4,5 and 6 weeks. Imunohistochemistry and reverse transcriptase polymerase chain reaction (RT-PCR) method were used to illustrate the distribution and changes of ETA receptor in different degree of ascites in broiler. In order to study the effect of ET-1 and BQ123 on the mean pulmonary arterial pressure (mPAP), morbidity of PHS, pathological change of the main tissues, physiological index and biochemical indicator, different dose of ET-1 and BQ123 were i.v. injected into the broiler. Myocardial cells of broiler were also cultured to illustrate the effection of different doses of ET-1, BQ123, L-Arg and L-NAME on the cultured myocardial cells.(1) Natural pulmonary hypertension syndrome broiler cases were appear from the age of 10 days after hyperalimentation dietary were used to feed a great deal of commercial Arbor Acre broiler chickens, the morbility is about 5.3% or 6.7%. Right heart catheterwas used to determine the mean pulmonary arterial pressure, it was found that, compared with the normal group, the AHI and mPAP of PH group were higher or significantly higher, the tendency of increment with the days delaying was observed. From 3 weeks of age, the level of plasma and tissue ET-1 of the PH group was higher or significantly higher than that of the normal group. At 6 weeks of age, the level of ET-1 in plasma, lung, kidney of PH group decreased slightly, though it was still higher than that of the normal group. The level of ET-l of tissues was higher than that of plasma, and the level of lung ET-1 was higher than that of other organs. The level of NO and NOS of the two groups increased with the days delayed, and the PH group was significantly higher than the normal group. Besides, there was a significant positive correlation between mPAP and ET-1, but no significant correlation was found between mPAP and plasma NO of the two groups. It is concluded that ET-1 and NO took part in the formation and development of pulmonary hypertension syndrome in broilers and played an important role in it.(2) Immunohistochemical method and reverse transcriptase polymerase chain reaction (RT-PCR) method were used to study the distribution and expression of ETA receptor (ETAR) in different degree of ascites broilers. It was found that the ETAR expressed extensively in the main tissues of the broilers. In heart, it mainly expressed in cardiac muscle cell envelope, cardiac pericardium and vascular smooth muscle. In liver, it mainly distributed in hepatic cell envelope and Kupffer's cells. In lung, it mainly expressed in blood vessel, bronchus secundarius, parabronchus and pulmonary alveoli. In distal convoluted tubule, renal glomeruli, macula densa, medullary loop and vessel of kidney of kidney, it was also has different degree of expression. Compared with the normal group, the expression of ETAR in lung blood vessel and pulmonary alveoli of PH group, light ascites group and heavy ascites group is higher or significant higher. But with the development of the degree of ascites, the expression of ETAR was decreased, and the expression of ETAR in the PH group is the highest. With the development of the degree of ascetes, the expression of ETAR of the heart was decreased or significant decreased, but it was still higher than the normal group. The expression of ETAR of myocardium of right ventricle was significant higher than of myocardium of left ventricle. In liver, the expression of ETAR in liver cells was higher than that of the normal group and the heavy ascites group. Compared with the normal group, the expression of ETAR in hepatic blood vessel was obviously decreased. In kidney, the expression of ETAR in nephric tubule, glomeruli of kidney and macula densa also showed complicated changes with the development of the degree of ascites. The primers of broiler endothelin type A receptorgene was used to RT-PCR. The heart cDNA was obtained from the normal group broiler of 16, 30 days old and heavy ascites broiler of 30 days old. The results showed that the expression of EDNRA mRNA of the normal 16days old broilers was obviously lower than that of the 30days old broilers. The expression of EDNRA mRNA of the heavy ascites broiler was obviously lower than that of the normal group of broilers.(3) When different degree of ascites group compared each other, the mean pulmonary arterial pressure, ascites heart index (AHI), right ventricle and body weight ratio (TV/BW), the level of plasma ET-1, plasma ANP and plasma NO were all significantly or every significantly increased with the development of ascites degree. Plasma and tissue endothelin-1 (ET-1), atrial natriuretic polypeptide (ANP), mPAP, AHI and NO of the normal, light and heavy ascites broiler groups were measure. It was found that AHI and mPAP of the light ascites group were higher or significantly higher than that of the normal group, and the heavy ascites group was higher or significantly higher than that of the light ascites group. The trend of increment with the days delaying was observed too. The change tendency of plasma and lung inducible nitric oxide synthase (iNOS), constitutive nitric oxide synthase (cNOS) level was complex. Compared with the normal and light ascites group, the level of plasma iNOS in heavy ascites group were higher or significantly higher. The trend of increment of iNOS with the day delaying was complex, it was decreased at first and then increased. The level of cNOS in heavy ascites was lower or significant lower than the normal and light ascites group. It showed complex change as broilers growing. Radioimmunoassay was used to detect tissue ET-1 and ANP of broilers in different degree ascites group of 5 weeks old broilers. It found that the content of ANP and ET-1 is significantly or very significantly increased with the development of the ascites. The level of the ET-1 and ANP in lung and heart is the highest than in other tissues, and the level of ANP in tissues was higher than that of plasma. Besides, there were significant positive correlation between mPAP and AHI, between plasma ANP, ET-1 and AHI, mPAP. There were significant positive correlation among plasma mPAP, AHI and cardiac ET-1 at 5 weeks of age, but no significant correlation was found between plasma NO and plasma ET-1, plasma ANP, cardiac ET-1.(4) Twenty eight days old broilers were randomly divided into several groups with 8 replicates each, infused intravenously with different dose of ET-1 and BQ123. The control group was injected 0.86% NaCl. The mean pulmonary arterial pressure (mPAP) was measured, and plasma A I , ANP, ET-1 at different time after the broilers were injected different dose of ET-1 and BQ123. The mPAP decreased from time 0 to 5 min and thenbegan to increase from time 5 to 30 min when the broilers were injected with ET-1 24, 120, 240ng/kg BW. The effective dose of ET-1 was 240ng/kg BW to make mPAP increase. BQ123 can decrease mPAP of the broilers with dose dependent and it also can counteract the increasing effect of blood pressure after injected ET-1, but it can't counteract the decreasing effect of blood pressure at the beginning of the injection. In other experiment, 170 broilers were divided into 3 groups: a control group (group C, 50 broilers), group A and B (60 broilers each) that intravenously injected with 240ng/kg BW and 360ng/kg BW ET-1, respectively. The PHS morbidity of group A was higher than that of group C but lower than group B. The right/total ventricular weight ratio (RV/TV) and mPAP were higher in group A and B than in group C. There was the same tendency of the values of packed cell volume (PCV) and plasma nitric oxide (NO) when compared group A, B with group C. The tissue slice of broilers showed that pathology changes appeared in the main tissues after ET-1 was injected into the broiler with long-term.(5) Enzyme digestion was used to cultivate myocardial cell of broilers. In this experiment, methyl thiazolyl tetrazolium (MTT) and 3H-TdR, 3H-Pro, 3H-Leu were used to study the effect of ET-1 and BQ123 on the broiler myocardial cell. In the present study, it indicated that when the concentration of ET-1 exceeded 10"8mol/L, ET-1 was obviously promoted the growth and hypertrophy of the myocardial cells. If the ET-1 concentration was higher, the growth promotion effect on the myocardial cells was better. When the concentration of BQ123 exceeded 10"8mol/L, it can completely counteract the promotion effect of ET-1 on myocardial cells of broilers, and the inhibitory action was obvious if the concentration of BQ123 was higher. Fluorescent labeling was used to study the effect of ET-1, BQ123, L-Arg and L-NAME on the change of [Ca2+]i in myocardial cells. ET-1 could promote [Ca2+]i of myocardial cells inflowing and BQ123 could counteract this effect soundly, but BQ123 could not counteract the transient increase effect of ET-1.
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