RESUMEN
1. The alterations of relative telomere length and expression of shelterin genes (TRF1, TRF2, RAP1, POT1, and TPP1) were evaluated from the chickens' right heart ventricle in the early and last stages of cold-induced pulmonary hypertension (PHS) at 21 and 42 d of age.2. The relative telomere length in the right ventricular tissues was significantly shorter in the PHS group of broilers than in the control group at 42 d, but did not statistically change at 21 d of age. There was a significant negative correlation between relative telomere length and RV:TV ratio in the broilers at 42 d of age.3. The relative expression of POT1, RAP1 and TPP1 genes in the right ventricular tissues was significantly lower in the PHS group than in the control group at 21 d. The relative expression of the TRF2 gene was only higher in the PHS group of broilers than control at 42 d. The mRNA level of the TRF2 gene exhibited a significant positive correlation with RV:TV ratio at 42 d.4. It was concluded that most shelterin genes are dysregulated in the early stage of PHS (right ventricular hypertrophy) while telomere attrition occurs only at the last stage (heart dilation/failure).
Asunto(s)
Pollos , Proteínas de Unión a Telómeros , Animales , Pollos/genética , Pollos/metabolismo , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo , Complejo Shelterina , Telómero/metabolismo , CorazónRESUMEN
Cold stress is an environmental cause of pulmonary hypertension syndrome (PHS) in broiler chickens. This factor could increase the rate of metabolic activity via thyroid hormones (T3 and T4). To evaluate the effect of these hormones on the heart, the plasma concentration of T3, T4, and the gene expression of their receptors (THRα and THRß) and many contractile proteins (ACTC1, MHCα, MHCß, RYR2, SERCA2, THRα, THRß, and troponin I) were measured in the right ventricle in 2 periods of age (21 and 35 d). Plasma T3 concentration was significantly higher in the PHS group of chickens than in the control one at 21 and 35 d while plasma T4 did not change. The relative expression of MHCα, RYR2, SERCA2, and THRα genes in the right ventricle tissues was only higher in PHS group of broilers than control group at 21 d (P < 0.05) whereas the expression of ACTC1, MHCß, and troponin I did not differ at 2 periods of age. The positive correlations between MHCα, RYR2, SERCA2, and T3, THRα were confirmed. The expression of THRß gene was only higher in PHS group of broilers than control at 35 d (P < 0.05). The data determined that cold stress could increase thyroid hormones and the gene expression of their receptor (THRα) in the pick of chicken growth (21 d) that they themselves elevates the expression of many genes related to contractile elements (MHCα, RYR2, and SERCA2), leading to adaptive right ventricle hypertrophy.
Asunto(s)
Hipertensión Pulmonar , Enfermedades de las Aves de Corral , Animales , Pollos/genética , Ventrículos Cardíacos , Hipertensión Pulmonar/genética , Hipertensión Pulmonar/veterinaria , Hormonas TiroideasRESUMEN
1. The relative expression of angiotensinogen (AGT), renin, angiotensin-converting enzyme (ACE) and angiotensin II type 1 receptor (AT1R) was determined using quantitative real-time PCR on tissue from the brain (forebrain, midbrain and hindbrain) to investigate the effect of cold-induced pulmonary hypertension syndrome (PHS) in broilers aged 42 days. Brain angiotensin II (Ang II) and AT1R levels were measured using enzyme immunoassay. 2. The right ventricle/total ventricles (RV/TV) ratio of the heart was increased in broilers exposed to cold stress (PHS group) at the end of the experiment. 3. ACE and renin transcripts in three parts of the brain were significantly increased in the PHS group at 42 d of age compared to controls while AGT transcript was significantly increased only in the hindbrain of PHS birds. The amount of AT1R transcript did not differ between control and PHS groups. 4. The amount of Ang II significantly decreased only in the midbrain of PHS birds compared with controls while the amounts of AT1R were not different between treatments in the three segments of the brain. 5. It was concluded that brain gene expression of AGT (in the hindbrain), renin, and ACE was upregulated in broilers with PHS whereas Ang II and AT1R levels were not changed. These results provided evidence of diminished involvement of the renin-angiotensin system in the pathogenesis of chicken pulmonary hypertension.
Asunto(s)
Proteínas Aviares/genética , Encéfalo/metabolismo , Pollos/genética , Hipertensión Pulmonar/veterinaria , Sistema Renina-Angiotensina/genética , Animales , Proteínas Aviares/metabolismo , Pollos/metabolismo , Frío/efectos adversos , Ensayo de Inmunoadsorción Enzimática/veterinaria , Hipertensión Pulmonar/etiología , Hipertensión Pulmonar/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinariaRESUMEN
Accurate normalization in real-time quantitative PCR is an important step in quantification of gene transcription pattern, in which proper application of stable reference gene(s) is crucial. To identify the most stable reference gene (s) in pulmonary hypertensive chickens, from a panel of 9 typical candidate genes, the expression of ACTB, HMBS, HPRT1, RPL13, RPL32, 18SrRNA, TBP, TFRC, and YWHAZ was determined in the lung and heart (right ventricle) of both healthy and cold-induced pulmonary hypertensive chickens at 42 d of age. The BestKeeper, geNorm, and NormFinder software programs were used to analyze this set of genes. Also, the ratio of right ventricle to the total ventricle was used as an index of induced pulmonary hypertension, which increased in the cold-treated chickens compared to the control at 42 d of age. Candidate reference genes ranking in the lung of pulmonary hypertensive chickens vs. healthy individuals included RPL13, YWHAZ, HMBS, ACTB, HPRT1, TFRC, RPL32, 18SrRNA, and TBP; those in the heart were YWHAZ, RPL13, HMBS, ACTB, HPRT1, TBP, 18SrRNA, TFRC, and RPL32; and those in the heart-lung combination included RPL13, YWHAZ, HMBS, HBRT1, TFRC, ACTB, 18SrRNA, RPL32, and TBP. The overall results showed that the most stable genes are YWHAZ, RPL13, HMBS, ACTB, HBRT1, TFRC, TBP, RPL32, and 18SrRNA, respectively. In addition, the combination of YWHAZ, RPL13, and HMBS is recommended as the reference gene panel for more accurate quantitative data normalization of heart or lung in the chicken pulmonary hypertension.
Asunto(s)
Pollos/genética , Perfilación de la Expresión Génica/veterinaria , Hipertensión Pulmonar/veterinaria , Enfermedades de las Aves de Corral/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Animales , Perfilación de la Expresión Génica/métodos , Hipertensión Pulmonar/genética , Pulmón/metabolismo , Miocardio/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa/métodosRESUMEN
Quantitative real-time PCR was carried out to evaluate gene expression of heat shock proteins (HSP) (HSP27, HSP56, HSP60, HSP70, HSP90 and ubiquitin) in the brain (hindbrain, midbrain, forebrain) of chickens with cold-induced pulmonary hypertension. The ratio of the right ventricle to the total ventricle (index of pulmonary hypertension in chickens) was increased in the cold-induced pulmonary hypertensive chickens at 42 d of age compared with control. The HSP genes were expressed in the three parts of the brain in the two experimental groups. In the hindbrain of cold-induced pulmonary hypertensive chickens, the relative gene expression of HSP27, HSP60, HSP70 and HSP90 was decreased while gene expression of HSP56 and ubiquitin was increased compared with controls. In the midbrain of cold induced-pulmonary hypertensive chickens, the expression of HSP56, HSP60, HSP70 and ubiquitin genes was increased compared with controls while HSP27 and HSP90 were decreased. In the forebrain of cold induced-pulmonary hypertensive chickens, the expression of HSP56, HSP60, HSP70 and ubiquitin genes was increased while the expression of the HSP27 gene was decreased compared with controls. It is concluded that overexpression of HSPs in the forebrain and midbrain probably delays the pathological process of cold stress whereas diminished expression of HSP genes in the hindbrain may affect the normal function of brain centres in this area to exacerbate pulmonary hypertension.
Asunto(s)
Pollos , Expresión Génica , Hipertensión Pulmonar/veterinaria , Enfermedades de las Aves de Corral/genética , Animales , Encéfalo/metabolismo , Frío/efectos adversos , Proteínas de Choque Térmico/genética , Hipertensión Pulmonar/genética , Hipertensión Pulmonar/metabolismo , Enfermedades de las Aves de Corral/etiología , Enfermedades de las Aves de Corral/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinariaRESUMEN
Quantitative real-time PCR and Griess reaction were conducted to evaluate gene expression of nitric oxide synthases (eNOS, nNOS and iNOS) and nitric oxide (NO) production in the hindbrain, midbrain, forebrain of chickens with cold-induced pulmonary hypertension. The ratio of the right to total ventricular weight of heart as an indication of pulmonary hypertension was increased in the cold stress groups of chickens at 42 d of age. In the pulmonary hypertensive chickens, production of NO was increased in the hindbrain but was unchanged in the forebrain and midbrain. Relative gene expression of eNOS and nNOS was upregulated in the three segments of brain, whereas the iNOS transcript was downregulated in the forebrain and midbrain of the cold-induced pulmonary hypertensive chickens. It is concluded that in the chickens with cold-induced pulmonary hypertension, variations of nNOS, eNOS and iNOS gene expression would lead to overproduction of NO in the hindbrain, whereas the variations in the expression of these genes did not result in an elevation of NO in the forebrain and midbrain. It is suggested that high levels of NO in the hindbrain excites neural mechanisms involved in pulmonary hypertension.
Asunto(s)
Proteínas Aviares/genética , Pollos , Regulación de la Expresión Génica , Hipertensión Pulmonar/veterinaria , Óxido Nítrico Sintasa/genética , Enfermedades de las Aves de Corral/genética , Animales , Proteínas Aviares/metabolismo , Frío , Hipertensión Pulmonar/etiología , Hipertensión Pulmonar/genética , Mesencéfalo/metabolismo , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa de Tipo I/genética , Óxido Nítrico Sintasa de Tipo I/metabolismo , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismo , Óxido Nítrico Sintasa de Tipo III/genética , Óxido Nítrico Sintasa de Tipo III/metabolismo , Enfermedades de las Aves de Corral/etiología , Prosencéfalo/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Rombencéfalo/metabolismo , Estrés FisiológicoRESUMEN
1. The relative expression of heat shock protein (HSP) genes (HSP60, HSP70 and HSP90) was performed using quantitative real-time PCR on tissue from the heart ventricles to investigate the effect of 3,5,3'-l-triiodothyronine (T3)-induced pulmonary hypertension in broiler chickens. 2. The ratio of the right ventricle to total ventricle (index of pulmonary hypertension) was increased in the treated groups at 12 and 42 d of age compared to controls but was significant only at 42 d. 3. The HSP genes were expressed in the right and left ventricles of control but T3-treated broilers at 12 and 42 d of age. The relative amounts of HSP60 and HSP90 gene expression in the right ventricle of treated groups were significantly increased at 12 d and decreased at 42 d of age compared to controls. 4. Variations of HSP60 and HSP90 mRNAs in the left ventricle were not significant. The relative amount of HSP70 mRNA expression in the right and left ventricles of treated groups was significantly decreased at 42 d of age compared to controls. HSP70 mRNA expression did not differ between the right and left ventricles at 12 d of age. 5. It is concluded that gene expression of HSPs (i.e. HSP60 and HSP90) was upregulated in the heart of chickens developing pulmonary hypertension syndrome, probably to delay the pathological process of disease. The right ventricle from hearts of pulmonary hypertensive chickens showed considerable reductions of HSP60, HSP70 and HSP90, which is evidence of a loss of compensatory responsiveness in dilated heart.