| Vitamin D is used as an essential nutrient to regulate the intestinal calcium(Ca)absorption of poultry.Vitamin D3 and 25-hydroxycholecalciferol(25-OH-D3)have been authorized as feed additives in poultry.The final active product of vitamin D is 1,25-dihydroxycholecalciferol(1,25-(OH)2-D3).1,25-(OH)2-D3 binds to vitamin D receptor(VDR)to regulate the Ca transporter gene expression and promotes intestinal calcium absorption.TRPV6,CaBP-D28k,PMCAlb and NCX1 are key transporters for Ca absorption in poultry small intestine.Seldom reports about Ca transporter gene expression in broilers were found.Therefore,this study was conducted to investigate the effects of age,dietary Ca levels,and 1,25-(OH)2-D3 on the Ca transporter gene expressions in the small intestine of broiler chickens.Experiment 1 Effects of age on growth performance,bone development,and calcium transporter gene expression levels in the small intestine of broiler chickensThis study was conducted to evaluate the effects of age on body weight,bone growth,and mRNA expression levels of calbindin-D28k(CaBP-D28k),plasma membrane calcium pump(PMCAlb),nuclear vitamin D receptor(nVDR),and membrane vitamin D receptor(mVDR)in the duodenum of broiler chickens fiom 14 to 42 days of age.A total of 96 males of 1-day-old Ross 308 broilers were randomly allotted to 8 replicates of 12 chickens per replicate.Broilers were fed diets with adequate nutrients.Eight chickens were slaughtered at 14,21,28,35,and 42 days of age,respectively.The mRNA expression levels of Ca transporter genes in the duodenum were analyzed using RT-PCR.Results showed that body weight of broiler chickens linearly increased from 14 to 42 days of age.The weight,length,diameter,and ash weight of the femur linearly increased from 14 to 42 days of age.By contrast,the percentage contents of ash,calcium,and phosphorus in the femur linearly decreased with age.Similar results were found in the tibia.The mRNA expression levels of CaBP-D28k,PMCA1b,nVDR,and mVDR in the duodenum of broiler chickens quadratically increased with age.The highest mRNA levels of the four genes were observed at 28 days of age.No significant differences in mRNA expression levels of CaBP-D28k were observed among ages.These data revealed that body weight and the weight and ash weight of leg bone linearly increased with age;bone ash contents linearly decreased with age;and gene expression levels of CaBP-D28k,PMCAlb,nVDR,and mVDR quadratically increased with age.These results indicated that the Ca absorption ability in the small intestine of broilers changes with age.Experiment 2 Effect of dietary calcium levels on growth performance,bone development,and Ca transporter gene expression levels in the small intestine of broiler chickensThis study was conducted to investigate the effect of dietary calcium(Ca)levels on the growth performance,bone development and Ca transporter gene expression levels in the small intestine of broiler chickens from 1 to 18 days of age.On the day of hatch,350 males of Ross 308 broilers were randomly allotted to 5 treatments with 5 replicates(14 birds per replicate).Dietary Ca levels were 0.50%,0.70%,0.90%,1.10%,and 1.30%,in which 0.90%was the control.All diets contained 0.45%non-phytate phosphorus(NPP).The increase in dietary Ca levels from 0.50%to 1.30%did not affect the average daily feed intake,daily weight gain,or feed convertion ratio in 1-to 18-day-old broilers fed with 0.45%NPP(P>0.05).Dietary Ca levels did not affect the tibia weight in broilers at 18 days of age(P>0.05).Increasing the Ca levels from 0.50%to 1.30%linearly increased the ash weight and the percentage contents of ash,Ca and phosphorus(P)in the tibia(P<0.05).The percentage contents of minerals in broilers fed with 0.90%Ca were higher than those in birds fed with 0.50%Ca(P<0.05).No difference in tibia qualities was observed in broilers fed with 0.90%-1.30%-Ca(P>0.05).Increasing the Ca levels from 0.50%to 1.30%linearly decreased the mRNA expression levels of Ca-binding protein 28-kDa(CaBP-D28k),plasma membrane Ca-transporting ATPase lb(PMCAlb),sodium(Na)/Ca exchanger 1(NCX1),nuclear vitamin D receptor(nVDR)and membrane vitamin D receptor(mVDR)in the duodenum of broilers at 18 days of age(P<0.05).The mRNA expression levels of CaBP-D28k and PMCAlb in the duodenum of broilers fed with 0.90%Ca were lower than those of birds fed with 0.50%Ca(P<0.05).No differences in the mRNA expression levels of CaBP-D28k and PMCAlb in the duodenum of broilers fed with 0.90%-1.30%Ca(P>0.05).The mRNA expression levels of nVDR and mVDR in broilers fed with 1.10%Ca were lower than those of birds fed with 0.50%Ca(P<0.05).Similar results were found in the jejunum and ileum.Broilers fed with 0.90%-1.30%Ca had lower mRNA expression levels of CaBP-D28k and PMCAlb in the small intestine than birds fed with 0.50%Ca(P<0.05).No difference was noted in the mRNA expression levels of CaBP-D28k and PMCAlb of birds fed with 0.90%-1.30%Ca(P>0.05).Compared with those fed with 0.50%Ca,birds fed with 1.30%Ca had lower mRNA expression levels of NCX1 in the duodenum and jejunum(P<0.05),and birds fed with 1.10%Ca obtained lower mRNA expression levels of nVDR and mVDR in the duodenum(P<0.05).These data indicated that low levels of dietary Ca stimulated Ca transporter gene transcription,but high levels of Ca inhibited Ca transporter gene expression levels and prevented excessive Ca absorption in the small intestine of broiler chickens.Experiment 3 Effects of 1,25-dihydroxycholecalciferol on growth performance,bone development,and calcium transporter gene expression levels in the small intestine of broiler chickens1,25-Dihydroxycholecalciferol(1,25-(OH)2-D3)is the final active product of vitamin D.This study aimed to investigate the effects of 1,25-(OH)2-D3 on the growth performance,bone development,and calcium(Ca)transporter gene expression levels in the small intestine of broiler chickens.On the day of hatch,210 females of Ross 308 broilers were randomly allotted to 3 treatments with 5 replicates(14 birds per replicate).Three levels of 1,25-(OH)2-D3(0,1.25,and 2.50 μg/kg)were added to the basal diet without vitamin D.Results showed that the addition of 1,25-(OH)2-D3 increased average daily feed intake and average daily gain and decreased feed convertion ratio in 1-to 19-day-old broiler chickens compared with the basal diet without vitamin D(P<0.05).1,25-(OH)2-D3 also enhanced the weight,ash weight,and the percentage contents of ash,Ca,and P in the tibia and femur in broilers(P<0.05).The mRNA expression levels of Ca-binding protein(CaBP-D28k)in the duodenum,jejunum,and ileum of 19-day-old broilers were increased by 88.1,109.1,and 2.7 times after adding 1.25 μg/kg 1,25-(OH)2-D3(P<0.05).No differences in the three gene expression levels were observed between birds fed with 1.25 and 2.50μg/kg 1,25-(OH)2-D3(P>0.05).The mRNA expression levels of PMCA1b in the duodenum and jejunum were also enhanced by 1,25-(OH)2-D3 addition(P<0.05).By contrast,the mRNA expression levels of NCX1 in the duodenum and that of vitamin D receptor(VDR)in the duodenum and jejunum were not affected by 1,25-(OH)2-D3(P>0.05).These data indicated that 1,25-(OH)2-D3 upregulated Ca transporter gene transcription and promoted Ca2+ absorption in the small intestine especially in the proximal intestine(i.e.duodedum and jejunum),thereby improved growth performance and bone mineralization of broiler chickens.In conclusion,the Ca transporter gene expression and Ca absorption ability in the small intestine of broilers changes with age.Low levels of dietary Ca stimulate Ca transporter gene transcription,but high levels of Ca inhibite Ca transporter gene expression and prevent excessive Ca absorption in the small intestine of broilers.The addition of 1,25-(OH)2-D3 upregulates Ca transporter gene transcription and promotes Ca2+ absorption in the small intestine especially in the proximal intestine(i.e.duodedum and jejunum),thereby improves growth performance and bone mineralization of broiler chickens. |
