Cardiac telomere attrition following changes in the expression of shelterin genes in pulmonary hypertensive chickens.

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).

Differential expression of genes implicated in liver lipid metabolism in broiler chickens differing in weight.

1. Lipid parameters and expression of ACACA, APOA1, CPT1A, FASN, FOXO1, LIPG, PPARα and SIRT1 genes involved in lipid metabolism were investigated in two groups of high (HW) and low (LW) weight broilers from the same strain.2. Blood cholesterol and liver triglyceride levels were significantly increased in HW chickens compared to LW broilers, while other parameters, i.e. blood triglyceride, blood HDL/LDL, liver cholesterol and total liver fat showed no significant changes in either group.3. The relative expression of ACACA, APOA1 and CPT1A genes was significantly lower in the liver tissues of HW broilers than in the LW group. The mRNA levels of these three genes showed a significant negative correlation with abdominal fat deposition and live weight of broilers. However, relative expression of FASN, FOXO1, LIPG, PPARα and SIRT1 hepatic genes did not differ among broilers.4. It was concluded that, of eight hepatic genes implicated in lipid metabolism, only the expression of three (ACACA, APOA1 and CPT1A) were significant for fat and leanness within the same strain of chicken. Since reducing body fat is a major goal in the broiler industry, these data can provide fresh insight into the molecular processes underlying the regulation of fat deposition in broilers.

Brain renin-angiotensin system in broiler chickens with cold-induced pulmonary hypertension.

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.

Cardiac renin-angiotensin system (gene expression) and plasma angiotensin II in chickens with T3-induced pulmonary hypertension.

The relative expression of renin, angiotensin converting enzyme (ACE) and angiotensin II type 1 receptor (AT1R) 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 syndrome (PHS) in broiler chickens at 12 and 42 d of age. Plasma angiotensin II was also measured using enzyme immunoassay at 12, 21, 28, 32 and 42 d. Renin transcript in the right ventricle of the treated group was significantly increased at 12 d and decreased at 42 d of age compared to controls. ACE transcript in the left ventricle of the treated group was significantly increased at 42 d of age compared to controls. In the right ventricle, the expression of this gene was significantly increased at 12 d while decreased at 42 d of age in the treated group compared to control. AT1R transcript in the right ventricle of the treated group was significantly increased at 12 and 42 d of age compared to control. In the left ventricle of the treated group, the transcript of this gene was only higher at 12 d of age than control. Plasma angiotensin II was significantly increased in pulmonary hypertensive chickens at 28 d of age compared to control. It is concluded that gene expression of renin, ACE and AT1R was relatively upregulated in the heart of chickens developing pulmonary hypertension. The right ventricle of hearts from pulmonary hypertensive chickens showed considerable reductions of renin/ACE and elevation of AT1R which may provide evidence of diminished responsiveness of the renin-angiotensin system.

Variation of heat shock protein gene expression in the brain of cold-induced pulmonary hypertensive chickens.

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.

Antioxidant supplementation of low-protein diets reduced susceptibility to pulmonary hypertension in broiler chickens raised at high altitude.

A reduced-protein diet (designated as RPD) was prepared and its effects on growth performance and the development of pulmonary hypertension syndrome (PHS) were evaluated in broiler chickens compared to a normal-protein diet (designated as NPD) or to the RPD supplemented with CoQ10 alone (30 mg/kg) or in combination with vitamin E (30 mg/kg CoQ10 + 100 mg/kg vitamin E). The RPD had 30 g/kg less crude protein compared to the NPD. A total of 208 1-day-old male broilers (Ross 308 strain) were used in a 42-day trial. Serum concentrations of uric acid (UA) and nitric oxide (NO) significantly (p < 0.05) declined when chickens fed on the RPD. However, supplementing RPD with the antioxidants significantly (p < 0.05) increased the serum NO concentration. Although serum malondialdehyde (MDA) concentration was significantly (p < 0.05) higher in the RPD than the NDP, supplementing RPD with CoQ10 and CoQ10 + VE decreased serum MDA concentration to similar levels found in the NPD. Significant overexpression in GPX1 gene observed in the heart and lungs of broilers fed on the RPD, which was effectively restored by supplementation of CoQ10 . The right to total ventricular weight ratio (RV:TV) was significantly (p < 0.05) increased in birds fed the RPD, which concurred with an increase in mortality from pulmonary hypertension syndrome (PHS). However, a significant decline in mortality from PHS was observed when birds on RPD received CoQ10 or CoQ10 + VE. In conclusion, antioxidant supplementation effectively improves pulmonary hypertensive response in broiler chicken fed of reduced-protein diets.

Pulmonary hypertension and right ventricular failure in broiler chickens reared at high altitude is affected by dietary source of n-6 and n-3 fatty acids.

The present study evaluated the development of pulmonary hypertension and right ventricular failure in broiler chickens reared at high altitude (2100 m) as affected by dietary intake of n-3 and n-6 fatty acid sources. Flax oil and soy oil were used as sources of n-3 and n-6 fatty acids, respectively, either with or without α-tocopheryl acetate. A total of 192 day-old broiler chicks (Ross 308) were used in a completely randomized design using isoenergetic and isonitrogenous experimental diets. Results showed that dietary flax oil significantly (p < 0.05) improved feed conversion ratio during 21-42 days of age. However, body weight gain did not significantly differ among the experimental groups in entire trial. Birds received flax oil had significantly higher serum concentration of nitric oxide (NO) but they had lower serum concentration of malondialdehyde when compared with their counterparts fed with soy oil. Liver and abdominal fat weights were significantly (p < 0.05) reduced by substitution of soy oil for flax oil. The right-to-total ventricle weight ratio (RV/TV) and mortality from pulmonary arterial hypertension (PAH) were significantly (p < 0.05) decreased in birds that received flax oil. In conclusion, n-3 fatty acids could significantly reduce RV:TV and PAH mortality in birds by increasing circulatory level of NO and suppressing hepatic lipogenesis.

Differential gene expression of three nitric oxide synthases is consistent with increased nitric oxide in the hindbrain of broilers with cold-induced pulmonary hypertension.

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.

L-arginine supplementation of reduced-protein diets improves pulmonary hypertensive response in broiler chickens reared at high altitude.

This study was conducted at high altitude (2100 m above sea level). A total of 208 d-old male broilers (Ross 308) were randomised across 16 floor pens and reared up to 42 d. A normal-protein diet (NPD) was formulated according to the National Research Council. A reduced-protein diet (RPD) was prepared with dietary protein reduced by 30 g/kg relative to that of the NPD. Two additional diets were prepared by adding 2 and 4 g L-arginine (ARG)/kg to the RPD. At the end of trial (42 d), blood sampling was done and carcass characteristics were recorded. ARG supplementation of the RPD at 4 g improved feed:gain in the growing stage and throughout the trial compared with the RPD. The right-to-total ventricular weight ratio was significantly increased in birds fed on RPD compared with those fed on NPD or ARG-supplemented RPD. Feeding RPD caused a significant decrease in plasma concentrations of nitric oxide (NO) and uric acid. Plasma NO level, however, was restored by ARG supplementation of RPD. ARG supplementation of RPD, however, significantly reduced the rate of pulmonary hypertension syndrome mortality compared to the negative control fed on RPD.

Diminished gene expression of cardiac heat shock proteins in pulmonary hypertensive chickens.

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.

Feeding reduced-protein diets to broilers subjected to hypobaric hypoxia is associated with the development of pulmonary hypertension syndrome.

1. There are few studies on the effect of dietary protein content on pulmonary hypertension syndrome (PHS) in broiler chickens. Conflicting results from these studies prevent a clear conclusion on the effects of reduced-protein diets on development of the syndrome. 2. To obtain an understanding of the mechanisms involved, the current study, conducted at a high altitude (2100 m above sea level), evaluated the effect of three treatments that varied only according to dietary protein (CP) levels. One treatment with dietary CP advocated by National Research Council (1994) acted as a control. Two reduced-protein diets were also prepared with CP reduced 20 and 40 g/kg relative to the CP of the control, which were designated as LCPD2 and LCPD4, respectively. A total of 180 d-old male broilers (Ross 308) were randomised across 15 floor pens measuring 1.5 m(2) (12 birds per pen). Five such pens (replicates) were allotted to each dietary treatment. The protein treatments were applied from 1 to 42 d of age in which growth performance was measured and the mortality from PHS was monitored. At the end of trial (42 d), blood sampling was done and carcase characteristics were recorded. 3. Birds receiving LCPD4 gained more weight throughout the trial and had increased right ventricular weight ratio (RV:TV), relative liver weight, haematocrit, and heterophil: lymphocyte ratio at the end of the trial compared to the control. Plasma concentrations of nitric oxide (NO) and uric acid, however, were significantly lower in birds fed on LCPD4 than in those receiving the control diet. 4. Mortality from PHS was increased in birds fed the reduced-protein diets, which may have been associated with reduced concentration of plasma NO and increased haematocrit and RV:TV. In conclusion, feeding reduced-protein diets to broilers subjected to hypobaric hypoxia was associated with the development of PHS.

Event prediction in social network through Twitter messages analysis.

Event detection using social media analysis has attracted researchers' attention. Prediction of events especially in the management of social crises can be of particular significance. In this study, events are predicted through analyzing Twitter messages and examining the changes in the rate of Tweets in a specified subject. In the proposed method, the Tweets are initially preprocessed in consecutive fixed-length time windows. Tweets are then categorized using the non-negative matrix factorization analysis and the distance dependent Chinese restaurant process incremental clustering. The categorization results show that a high rate of Tweets entering a cluster represents the occurrence of a new event in near future. Finally, a description of the event is presented in the form of some frequent words in each cluster. In this paper, investigations on a Tweet dataset during a 6-month period indicate that the rate of sending Tweets about predictable events considerably changes before their occurrence. The use of this feature can make it possible to predict events with high degrees of precision.

Effects of supplementation of canola meal-based diets with arginine on performance, plasma nitric oxide, and carcass characteristics of broiler chickens grown at high altitude.

A total of 300 male broilers (Ross 308) were exposed to cool conditions at high altitudes to study the effects of dietary Arg supplementation on performance and physiological and zootechnical variables. A corn-soybean meal (SBM) and a corn-canola meal (CM) diet were formulated for the starting (1 to 3 wk of age) and growing (3 to 6 wk of age) stages according to NRC recommendations. Two additional diets were prepared by supplementing 0.2 and 0.4% l-Arg to the corn-CM diet. Substitution of CM for SBM caused a significant (P < 0.05) reduction in weight gain and feed intake and resulted in impaired feed:gain. Supplementing Arg in the CM diet restored the feed and weight losses to a significant extent so that a significant difference was found between CM diet and CM + 0.4% Arg in terms of weight gain for the growing (3 to 6 wk) stage and the entire study (1 to 6 wk; P < 0.05). Total plasma nitric oxide (NO) concentration analyzed by nitrate plus nitrite assay was measured in the treatment groups. A significant (P < 0.05) decrease in plasma NO level was observed by substituting CM for SBM in the diet. Supplementing the CM diet with Arg increased the plasma NO level above that of SBM group. Carcass and breast yields were significantly decreased (P < 0.05) as a result of substituting CM for SBM. The substitution of CM for SBM, however, significantly (P < 0.05) increased the proportions of thighs and heart. The right ventricular weight:total ventricular weight ratio and ascites mortality showed a significant (P < 0.05) increase when SBM was replaced by CM in the diet. Fortification of the CM diet with Arg eliminated the significant difference in the right-to-total ventricular weight ratios when compared with the SBM diet. In conclusion, feeding CM to broiler chickens raised at high altitude caused reduced growth performance and predisposed the birds to pulmonary hypertension and ascites, which were partly restored by Arg supplementation.

Electrocardiogram analysis of the normal unanesthetized green peafowl (Pavo muticus).

The aim of this study was to describe normal electrocardiogram (ECG) patterns and values in unanesthetized green peafowl (Pavo muticus). The standard bipolar and augmented unipolar limb leads ECGs were recorded in the birds. The waveforms were analyzed in all leads at 50 mm/sec and at 10 mm = 1 mV to determine P, PR (segment and interval), QRS, ST, QT durations, P, net QRS complex and T amplitudes. The polarity of each waveform was tabulated in all leads. The mean electrical axis (MEA) for the frontal plane was calculated using leads II and III. The mean heart rates were calculated from lead II that was 257.5 ± 5.33 beats/min. The P wave was dominantly positive in all leads. The dominant pattern of waveforms of the QRS complexes were QS in leads II, III, and aVF. In leads I, aVR and aVL, the R pattern was dominant. The T wave was almost positive in leads I, II, and aVF. The average value of the heart MEA was -96.75 ± 3.31°. Description of normal ECG parameters will facilitate a better realizing of ECG changes of unhealthy green peafowls.

Changes in the expression of cardiac genes responsive to thyroid hormones in the chickens with cold-induced pulmonary hypertension.

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.

Evaluation of endothelial and inducible nitric oxide synthase genes expression in the heart of broiler chickens with experimental pulmonary hypertension.

1. To clarify the effect of T(3)-induced pulmonary hypertension on endothelial and inducible nitric oxide synthase (eNOS and iNOS) mRNA expression in the ventricles of the heart, semi quantitative reverse transcription-PCR was performed on total RNAs isolated from broiler chicken hearts after feeding supplementary T(3) (15 mg T3/kg) for 6 weeks. NO metabolites (nitrite/nitrate) of serum were measured. 2. The eNOS and iNOS genes were expressed in the right and left ventricles of control and T(3)-treated broilers at 12, 28 and 49 d of age. The relative amount of eNOS mRNA expression in the right and left ventricles did not significantly differ between control and T(3)-treated broilers at any age. 3. The relative amount of iNOS mRNA expression in the right and left ventricles was lower in T(3)-treated broilers than in control broilers at 49 d of age, but not at 12 or 28 d. 4. The amount of NO metabolites was reduced in the serum of T(3)-treated chickens at 49 d of age when compared with the control. 5. It is concluded that eNOS and iNOS genes are normally expressed in the heart of broilers. It is probable that impaired NO synthesis and reduction of iNOS gene expression in the heart ventricles are involved in the pathophysiology of cardiac function in broilers with pulmonary hypertension.

Oral supplementation with vitamin C improves intestinal mucosa morphology in the pulmonary hypertensive broiler chicken.

1. The aim of this study was to determine the effects of added dietary vitamin C on the intestinal mucosa morphology in pulmonary hypertensive broiler chickens. 2. One-day-old chickens were reared at high altitude and treated with 400, 800 or 1200 mg vitamin C/l of drinking water for 48 d. Segments from the duodenum, jejunum and ileum were collected for morphometric analysis. 3. The duodenum and jejunum villus height, width, surface area and lamina propria thickness were greater in the highest vitamin C-treated groups compared to controls. 4. There were significant variations only in villus types of the jejunum and ileum: increased proportions of leaf + tongue-like villi and decreased proportions of convoluted + ridge-like villi at day 36 at the groups with 400 and 800 mg vitamin C/l. 5. It was concluded that vitamin C improves the gut morphology of the pulmonary hypertensive broiler chickens.

Role of nitric oxide produced by constitutive and inducible nitric oxide synthases in the mouse gastric fundus.

In the present study, the role of nitric oxide (NO) produced by constitutive and inducible nitric oxide synthases (cNOS and iNOS, resepctively) on the contraction and relaxation of fundus in normal and lipopolysaccharide (LPS)-treated mice was examined. A whole fundic ring isolated from mice pretreated with reserpine was mounted in an organ bath containing Krebs' solution with 0.001 mmol/L atropine. Rings were contracted initially by 5-hydroxytryptamine (5-HT; 0.03 mmol/L) before relaxation was induced using ATP (0.03 mmol/L), ADP (0.03 mmol/L), pentoxifylline (0.002 mmol/L), electrical field stimulation (EFS; 50 V, 1 msec, 50 Hz, 3 min) and L-arginine (0.05 mmol/L). All drugs and EFS induced significant relaxation of isolated rings. The relaxations induced were significantly inhibited by N(G)-nitro-L-arginine methyl ester (L-NAME; 1.0 mmol/L). However, the iNOS inhibitors L-N(6)-(1-iminoethyl) lysine hydrochloride (L-NIL; 1.0 mmol/L) and amino guanidine (AMG; 1.0 mmol/L) had no significant effect on tissue relaxation. Then, the relaxant effects of 0.03 mmol/L ATP were tested on precontracted isolated fundic rings taken from 10 mg/kg LPS-treated animals. The non-selective NOS inhibitor L-NAME (10 mg/kg), the iNOS inhibitors L-NIL (3 mg/kg) and AMG (20 mg/kg) and betamethasone (0.1 mg/kg) were used to examine the role of NO produced by iNOS in the relaxation responses. It was found that the level of contraction induced by 0.03 mmol/L 5-HT in rings isolated from LPS-treated animals was significantly (P < 0.5) less than that in rings from untreated mice. However, precontracted tissues from LPS-treated mice were significantly relaxed by ATP and the relaxation response to ATP was significantly inhibited by L-NIL, ANG and betamethasone, but not by L-NAME. We suggest that, in LPS-treated mice, the production of NO from iNOS produces a reduction in the contractile response, as well as a decrease in NO formation by cNOS, resulting in changes to smooth muscle cell function.

Identification of reliable reference genes for quantitative real-time PCR in lung and heart of pulmonary hypertensive chickens.

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.

Role of Selenium from Different Sources in Prevention of Pulmonary Arterial Hypertension Syndrome in Broiler Chickens.

Pulmonary arterial hypertension (PAH) syndrome in broilers is associated with hypoxia, which prevails at high altitude. Oxidative stress is the pathogenic mechanism underlying PAH. Because selenium is key element in the structure of antioxidant enzymes, we evaluated pulmonary hypertensive responses in broiler chickens fed with diets supplemented with organic or nano-selenium. One hundred forty-four broilers (starting at 5 days old) were fed with (i) control group: birds received a standard diet; (ii) nano-selenium group: birds were fed with basal diet supplemented with nano-selenium at 0.3 mg/kg; and (iii) organic selenium group: birds received basal diet supplemented with organic selenium at 0.3 mg/kg. We assessed growth performance, carcass characteristics, antioxidant variables, blood parameters, and small intestine morphology. Although Se supplementation did not affect growth performance, carcass traits, and organ weight (P > 0.05), the right to total ventricular weight ratio (RV:TV), malondialdehyde concentration in the liver, and heterophil to lymphocyte ratio were significantly lower in the nano-selenium group relative to the control (P < 0.05). Chickens that received nano-selenium also elicited significantly higher antibody titers after 24 h of an injection of sheep red blood cells (P < 0.05). Nano-selenium supplementation also significantly increased villus height, absorptive surface area, and lamina propria thickness relative to the control (P < 0.05) in different segments of the small intestine. In contrast, organic selenium supplement improved intestinal morphometry only in the jejunum. We conclude that dietary supplementation of 0.30 mg/kg nano-selenium could prevent right ventricular hypertrophy as reflected by reduced RV:TV, reduced levels of lipid peroxidation in the liver, and improved gut function.