Osteoblasts are considered as a major factor contributing to bone development and mineralization,however,few studies have been done to establish and evaluate the primary cultured tibial osteoblast model of broiler chi...Osteoblasts are considered as a major factor contributing to bone development and mineralization,however,few studies have been done to establish and evaluate the primary cultured tibial osteoblast model of broiler chicks.Therefore,in the present study,two experiments were conducted to establish and evaluate the primary cultured tibial osteoblast model of broiler chicks.In experiment 1,osteoblasts were isolated from the tibia of one-day-old Arbor Acre male broiler chicks using the explant method and identified through the cell morphology,alkaline phosphatase(ALP)and alizarin red staining.Experiment 2 was carried out to evaluate the vitality and mineralization of primary cultured tibial osteoblasts of broilers on days 4,8,12,16,20,24,28 and 32 after incubation,respectively.The results from experiment 1 demonstrated that primary cultured tibial osteoblasts of broilers showed a spindle-shaped,triangular or polygonal morphology.More than 95%of the cells were stained blue-black after ALP staining,and mineralized nodules were formed after 4 days of continuous incubation.In experiment 2,lactate dehydrogenase(LDH)activity stayed at a relatively stabilized level although incubation time affected(P=0.0012)it during the whole culture period.Additionally,incubation time affected(P≤0.0001)the number and proportion of the area of mineralized nodules.They increased linearly and quadratically(P<0.04)with the increase of incubation time,and remained at a stabilized level from 24 to 32 days of incubation.The estimates of the optimal incubation time were 17 and 26 days based on the best fitted broken-line or quadratic models(P<0.0001)of the number and proportion of the area of mineralized nodules,respectively.These results indicate that the primary cultured tibial osteoblast model of broilers has been established successfully by the explant method,and it showed typical osteoblast morphology and characteristics of ALP activity and mineralization,and could maintain a relatively stabilized vitality from 4 to 32 days of incubation;and the optimal incubation time of primary tibial osteoblasts was 17 to 26 days.Therefore,it could be used to further study the underlying mechanisms of bone development and mineralization of broiler chicks.展开更多
Cinnamon essential oil(CEO)and its combination with bamboo leaf flavonoid(BLF)have been shown to exhibit an additive antibacterial effect in vitro,but their functions in broilers were not clear.An experiment was condu...Cinnamon essential oil(CEO)and its combination with bamboo leaf flavonoid(BLF)have been shown to exhibit an additive antibacterial effect in vitro,but their functions in broilers were not clear.An experiment was conducted to investigate the effects of dietary graded levels of CEO and its combination with BLF on the growth performance,immune responses,antioxidative ability,and intestinal morphology and microbiota of broilers fed a corn-soybean meal basal diet.A total of 576 1-d-old Arbor Acres commercial male broilers were randomly allotted to 9 treatments with 8 replicates of 8 birds each in a completely randomized design.Birds were fed on a basal corn-soybean meal diet(control,without plant extracts and antibiotics),or the basal diet supplemented with 50 mg of aureomycin kg^-1,50,100,200,400,or 800 mg of CEO kg^-1,a combination of 100 mg of CEO and 16.7 mg of BLF kg^-1,or a combination of 200 mg of CEO kg^-1 and 33.3 mg of BLF kg^-1 for 42 d.Dietary treatment affected(P<0.05)the serum immune globulin M(IgM)contents on d 42,liver malondialdehyde(MDA)contents on d 21,duodenal crypt depth on d 42,relative abundance of Lactobacillus in the cecal contents on d 21,and relative abundances of Escherichia coli and Bifidobacterium in the cecal contents on d 42,but had no effect(P>0.16)on all other measured indices.The addition of 400 mg of CEO kg^-1 or a combination of 200 mg CEO kg^-1 and 33.3 mg BLF kg^-1 increased(P<0.02)serum IgM contents on d 42.Dietary supplementation with 100 or 200 mg CEO kg^-1,or 50 mg aureomycin kg^-1 decreased(P<0.003)liver MDA contents on d 21.In addition,the supplement of 100 mg CEO kg^-1 increased(P<0.002)the Lactobacillus relative abundance in caecum on d 21 and Bifidobacterium relative abundance in caecum on d 42,and decreased(P<0.0001)E.coli relative abundance in caecum on d 42.The results indicated that dietary supplementation with CEO,an alternative to aureomycin,improved the immune status,antioxidantative ability and cecal microbiota of broilers,and dietary supplementation with the combinations of CEO and BLF did not exhibit further effects.Dietary supplementation with 100 mg CEO kg^-1 is beneficial for broilers fed a corn-soybean meal basal diet.展开更多
This experiment was conducted to investigate the effect of dietary calcium(Ca)or phosphorus(P)deficiency on bone development and related Ca or P metabolic utilization parameters of broilers from 22 to 42 days of age b...This experiment was conducted to investigate the effect of dietary calcium(Ca)or phosphorus(P)deficiency on bone development and related Ca or P metabolic utilization parameters of broilers from 22 to 42 days of age based on our previous study,which indicated that dietary Ca or P deficiency impaired the bone development by regulating related Ca or P metabolic utilization parameters of broilers from 1 to 21 days of age.A total of 504 one-day-old Arbor Acres male broilers were randomly assigned to 1 of 4 treatments with 7 replicates in a completely randomized design,and fed the normal control and Ca-or P-deficient diets from 1 to 21 days of age.At 22 days of age,the broilers were further fed the normal control diet(0.90%Ca+0.35%non-phytate P(NPP)),the P-deficient diet(0.90%Ca+0.18%NPP),the Ca-deficient diet(0.30%Ca+0.35%NPP)or the Ca and P-deficient diet(0.30%Ca+0.18%NPP),respectively.The results showed that dietary Ca or P deficiency decreased(P<0.05)tibia bone mineral density(BMD),bone breaking strength(BBS),ash content,tibia ash Ca content and serum P content on days 28 and 42,but increased(P<0.05)tibia alkaline phosphatase(ALP)activity of broilers on day 42 compared with the control group.Furthermore,the broilers fed the P-deficient diet had the lowest(P<0.05)tibia BMD,BBS,ash content,serum P content and the highest(P<0.05)serum Ca content on day 28 compared with those fed the Ca-deficient or Ca and P-deficient diets.The results from the present study indicated that the bone development and related Ca or P metabolic utilization parameters of broilers were the most sensitive to dietary P deficiency,followed by dietary Ca deficiency or Ca and P-deficiency;dietary Ca or P deficiency impaired the bone development possibly by regulating serum Ca and P contents as well as tibia Ca content and ALP activity of broilers from 22 to 42 days of age.展开更多
The bone morphogenetic protein(BMP)and mitogen-activated protein kinase(MAPK)signaling pathways play an important role in regulation of bone formation and development,however,it remains unclear that the effect of diet...The bone morphogenetic protein(BMP)and mitogen-activated protein kinase(MAPK)signaling pathways play an important role in regulation of bone formation and development,however,it remains unclear that the effect of dietary different levels of non-phytate phosphorus(NPP)on these signaling pathways and their correlations with bone phosphorus(P)retention and bone development in broilers.Therefore,this experiment was conducted to investigate the effect of dietary P supplementation on BMP and MAPK signaling pathways and their correlations with bone P retention and bone development in broilers.A total of 800 one-day-old Arbor Acres male broilers were randomly allotted to 1 of 5 treatments with 8 replicates in a completely randomized design.The 5 treatments of dietary NPP levels were 0.15,0.25,0.35,0.45 and 0.55%or 0.15,0.22,0.29,0.36 and 0.43%for broilers from 1 to 21 days of age or 22 to 42 days of age,respectively.The results showed that extracellular signal-regulated kinase 1(ERK1)mRNA expression in the tibia of broilers on days 14 and 28,phosphorylated-ERK1(p-ERK1)on day 14,and BMP2 protein expression on days 28 and42 decreased linearly(P<0.04),while c-Jun N-terminal kinase 1(JNK1)mRNA expression on day 42 increased linearly(P<0.02)with the increase of dietary NPP level.At 14 days of age,total P accumulation in tibia ash(TP),bone mineral concentration(BMC),bone mineral density(BMD),bone breaking strength(BBS)and tibia ash were negatively correlated(r=-0.726 to-0.359,P<0.05)with ERK1 and JNK1 mRNA as well as p-ERK1;tibia alkaline phosphatase(ALP)and bone gal protein(BGP)were positively correlated(r=0.405 to 0.665,P<0.01)with ERK1 mRNA and p-ERK1.At 28 days of age,TP,BMC,BMD,BBS and tibia ash were negatively correlated(r=-0.518 to-0.370,P<0.05)with ERK1 mRNA and BMP2 protein,while tibia ALP was positively correlated(r=0.382 to 0.648,P<0.05)with them.The results indicated that TP,BMC,BMD,BBS or tibia ash had negative correlations,while tibia ALP and BGP had positive correlations with ERK1 and JNK1 mRNAs,BMP2 protein and p-ERK1,suggesting that bone P retention and bone development might be regulated by BMP and MAPK signaling pathways in broiler chickens.展开更多
The aim of the study was to investigate whether phosphorus(P) transporters, type IIb sodium-dependent phosphate cotransporter(NaP-IIb) and inorganic phosphate transporter 2(PiT2), were directly involved in P absorptio...The aim of the study was to investigate whether phosphorus(P) transporters, type IIb sodium-dependent phosphate cotransporter(NaP-IIb) and inorganic phosphate transporter 2(PiT2), were directly involved in P absorption across primary cultured duodenal epithelial cell monolayers of chick embryos. The siRNAs against NaP-IIb or PiT2 were designed, synthesized and transfected into primary cultured duodenal epithelial cells of chick embryos. Then, the inhibitory efficiency of siRNAs against NaP-IIb or PiT2 was analyzed, and the most efficacious siRNAs were selected to be used for subsequent P absorption experiments. Briefly, primary cultured duodenal epithelial cells of chick embryos were transfected with either NaP-IIb or PiT2 siRNAs and grown in confluent monolayers on transwell plates. The untransfected or transfected cell monolayers were then incubated in an uptake medium containing 0 or 0.25 mmol L^(–1) of P as KH_(2) PO_(4) to measure the P absorption across duodenal epithelial cell monolayers. The results showed that among the siRNAs designed, si-1372 and si-890 were demonstrated to be the most effective in inhibiting the NaPIIb and PiT2 expressions, respectively. Supplemental P increased(P=0.065) the protein abundance of PiT2 and enhanced(P<0.0001) P absorption in primary cultured duodenal epithelial cell of chick embryos. Furthermore, NaPIIb silencing decreased(P=0.07) P absorption across duodenal epithelial cell monolayers, while PiT2 silencing had no effect(P=0.345). It is concluded that the NaP-IIb, but not PiT2, might be directly involved in the P absorption of chick duodenal epithelial cells.展开更多
基金supported by the Key Program of the National Natural Science Foundation of China(31630073)the Initiation Funds of Yangzhou University for Distinguished Scientists,Chinathe Agricultural Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences(ASTIP-IAS09)。
文摘Osteoblasts are considered as a major factor contributing to bone development and mineralization,however,few studies have been done to establish and evaluate the primary cultured tibial osteoblast model of broiler chicks.Therefore,in the present study,two experiments were conducted to establish and evaluate the primary cultured tibial osteoblast model of broiler chicks.In experiment 1,osteoblasts were isolated from the tibia of one-day-old Arbor Acre male broiler chicks using the explant method and identified through the cell morphology,alkaline phosphatase(ALP)and alizarin red staining.Experiment 2 was carried out to evaluate the vitality and mineralization of primary cultured tibial osteoblasts of broilers on days 4,8,12,16,20,24,28 and 32 after incubation,respectively.The results from experiment 1 demonstrated that primary cultured tibial osteoblasts of broilers showed a spindle-shaped,triangular or polygonal morphology.More than 95%of the cells were stained blue-black after ALP staining,and mineralized nodules were formed after 4 days of continuous incubation.In experiment 2,lactate dehydrogenase(LDH)activity stayed at a relatively stabilized level although incubation time affected(P=0.0012)it during the whole culture period.Additionally,incubation time affected(P≤0.0001)the number and proportion of the area of mineralized nodules.They increased linearly and quadratically(P<0.04)with the increase of incubation time,and remained at a stabilized level from 24 to 32 days of incubation.The estimates of the optimal incubation time were 17 and 26 days based on the best fitted broken-line or quadratic models(P<0.0001)of the number and proportion of the area of mineralized nodules,respectively.These results indicate that the primary cultured tibial osteoblast model of broilers has been established successfully by the explant method,and it showed typical osteoblast morphology and characteristics of ALP activity and mineralization,and could maintain a relatively stabilized vitality from 4 to 32 days of incubation;and the optimal incubation time of primary tibial osteoblasts was 17 to 26 days.Therefore,it could be used to further study the underlying mechanisms of bone development and mineralization of broiler chicks.
基金supported by the Special Fund for Agro-scientific Research in the Public Interest, China (201403047)the Special Funds of CAAS for Distinguished Scientists, China+2 种基金the earmarked fund for China Agriculture Research System (CARS-41)the Agricultural Science and Technology Innovation Program, China (ASTIP-IAS08)the Key Agricultural Science and Technology Program of Datong, Shanxi, China (2016047)
文摘Cinnamon essential oil(CEO)and its combination with bamboo leaf flavonoid(BLF)have been shown to exhibit an additive antibacterial effect in vitro,but their functions in broilers were not clear.An experiment was conducted to investigate the effects of dietary graded levels of CEO and its combination with BLF on the growth performance,immune responses,antioxidative ability,and intestinal morphology and microbiota of broilers fed a corn-soybean meal basal diet.A total of 576 1-d-old Arbor Acres commercial male broilers were randomly allotted to 9 treatments with 8 replicates of 8 birds each in a completely randomized design.Birds were fed on a basal corn-soybean meal diet(control,without plant extracts and antibiotics),or the basal diet supplemented with 50 mg of aureomycin kg^-1,50,100,200,400,or 800 mg of CEO kg^-1,a combination of 100 mg of CEO and 16.7 mg of BLF kg^-1,or a combination of 200 mg of CEO kg^-1 and 33.3 mg of BLF kg^-1 for 42 d.Dietary treatment affected(P<0.05)the serum immune globulin M(IgM)contents on d 42,liver malondialdehyde(MDA)contents on d 21,duodenal crypt depth on d 42,relative abundance of Lactobacillus in the cecal contents on d 21,and relative abundances of Escherichia coli and Bifidobacterium in the cecal contents on d 42,but had no effect(P>0.16)on all other measured indices.The addition of 400 mg of CEO kg^-1 or a combination of 200 mg CEO kg^-1 and 33.3 mg BLF kg^-1 increased(P<0.02)serum IgM contents on d 42.Dietary supplementation with 100 or 200 mg CEO kg^-1,or 50 mg aureomycin kg^-1 decreased(P<0.003)liver MDA contents on d 21.In addition,the supplement of 100 mg CEO kg^-1 increased(P<0.002)the Lactobacillus relative abundance in caecum on d 21 and Bifidobacterium relative abundance in caecum on d 42,and decreased(P<0.0001)E.coli relative abundance in caecum on d 42.The results indicated that dietary supplementation with CEO,an alternative to aureomycin,improved the immune status,antioxidantative ability and cecal microbiota of broilers,and dietary supplementation with the combinations of CEO and BLF did not exhibit further effects.Dietary supplementation with 100 mg CEO kg^-1 is beneficial for broilers fed a corn-soybean meal basal diet.
基金The present study was financially supported by the National Key R&D Program of China(2017YF0502200)the Key Program of the National Natural Science Foundation of China(31630073)+2 种基金the earmarked fund for China Agriculture Research System(CARS-41)the Agricultural Science and Technology Innovation Program,China(ASTIP-IAS09)the earmarked fund for Hebei Chicken Innovation Team of Modern Agro-Industry Technology Research System,China(HBCT2018150203 and HBCT2018150206).
文摘This experiment was conducted to investigate the effect of dietary calcium(Ca)or phosphorus(P)deficiency on bone development and related Ca or P metabolic utilization parameters of broilers from 22 to 42 days of age based on our previous study,which indicated that dietary Ca or P deficiency impaired the bone development by regulating related Ca or P metabolic utilization parameters of broilers from 1 to 21 days of age.A total of 504 one-day-old Arbor Acres male broilers were randomly assigned to 1 of 4 treatments with 7 replicates in a completely randomized design,and fed the normal control and Ca-or P-deficient diets from 1 to 21 days of age.At 22 days of age,the broilers were further fed the normal control diet(0.90%Ca+0.35%non-phytate P(NPP)),the P-deficient diet(0.90%Ca+0.18%NPP),the Ca-deficient diet(0.30%Ca+0.35%NPP)or the Ca and P-deficient diet(0.30%Ca+0.18%NPP),respectively.The results showed that dietary Ca or P deficiency decreased(P<0.05)tibia bone mineral density(BMD),bone breaking strength(BBS),ash content,tibia ash Ca content and serum P content on days 28 and 42,but increased(P<0.05)tibia alkaline phosphatase(ALP)activity of broilers on day 42 compared with the control group.Furthermore,the broilers fed the P-deficient diet had the lowest(P<0.05)tibia BMD,BBS,ash content,serum P content and the highest(P<0.05)serum Ca content on day 28 compared with those fed the Ca-deficient or Ca and P-deficient diets.The results from the present study indicated that the bone development and related Ca or P metabolic utilization parameters of broilers were the most sensitive to dietary P deficiency,followed by dietary Ca deficiency or Ca and P-deficiency;dietary Ca or P deficiency impaired the bone development possibly by regulating serum Ca and P contents as well as tibia Ca content and ALP activity of broilers from 22 to 42 days of age.
基金supported by the Key Program of the National Natural Science Foundation of China(31630073)the China Agriculture Research System of MOF and MARA(CARS-41)the Agricultural Science and Technology Innovation Program,China(ASTIPIAS09)。
文摘The bone morphogenetic protein(BMP)and mitogen-activated protein kinase(MAPK)signaling pathways play an important role in regulation of bone formation and development,however,it remains unclear that the effect of dietary different levels of non-phytate phosphorus(NPP)on these signaling pathways and their correlations with bone phosphorus(P)retention and bone development in broilers.Therefore,this experiment was conducted to investigate the effect of dietary P supplementation on BMP and MAPK signaling pathways and their correlations with bone P retention and bone development in broilers.A total of 800 one-day-old Arbor Acres male broilers were randomly allotted to 1 of 5 treatments with 8 replicates in a completely randomized design.The 5 treatments of dietary NPP levels were 0.15,0.25,0.35,0.45 and 0.55%or 0.15,0.22,0.29,0.36 and 0.43%for broilers from 1 to 21 days of age or 22 to 42 days of age,respectively.The results showed that extracellular signal-regulated kinase 1(ERK1)mRNA expression in the tibia of broilers on days 14 and 28,phosphorylated-ERK1(p-ERK1)on day 14,and BMP2 protein expression on days 28 and42 decreased linearly(P<0.04),while c-Jun N-terminal kinase 1(JNK1)mRNA expression on day 42 increased linearly(P<0.02)with the increase of dietary NPP level.At 14 days of age,total P accumulation in tibia ash(TP),bone mineral concentration(BMC),bone mineral density(BMD),bone breaking strength(BBS)and tibia ash were negatively correlated(r=-0.726 to-0.359,P<0.05)with ERK1 and JNK1 mRNA as well as p-ERK1;tibia alkaline phosphatase(ALP)and bone gal protein(BGP)were positively correlated(r=0.405 to 0.665,P<0.01)with ERK1 mRNA and p-ERK1.At 28 days of age,TP,BMC,BMD,BBS and tibia ash were negatively correlated(r=-0.518 to-0.370,P<0.05)with ERK1 mRNA and BMP2 protein,while tibia ALP was positively correlated(r=0.382 to 0.648,P<0.05)with them.The results indicated that TP,BMC,BMD,BBS or tibia ash had negative correlations,while tibia ALP and BGP had positive correlations with ERK1 and JNK1 mRNAs,BMP2 protein and p-ERK1,suggesting that bone P retention and bone development might be regulated by BMP and MAPK signaling pathways in broiler chickens.
基金supported by the Key Program of the National Natural Science Foundation of China(31630073)the National Natural Science Foundation of China(31472116)+2 种基金the National Key R&D Program of China(2017YFD0502200)the China Agriculture Research System of MOF and MARA(CARS-41)the Agricultural Science and Technology Innovation Program(ASTIP-IAS09)。
文摘The aim of the study was to investigate whether phosphorus(P) transporters, type IIb sodium-dependent phosphate cotransporter(NaP-IIb) and inorganic phosphate transporter 2(PiT2), were directly involved in P absorption across primary cultured duodenal epithelial cell monolayers of chick embryos. The siRNAs against NaP-IIb or PiT2 were designed, synthesized and transfected into primary cultured duodenal epithelial cells of chick embryos. Then, the inhibitory efficiency of siRNAs against NaP-IIb or PiT2 was analyzed, and the most efficacious siRNAs were selected to be used for subsequent P absorption experiments. Briefly, primary cultured duodenal epithelial cells of chick embryos were transfected with either NaP-IIb or PiT2 siRNAs and grown in confluent monolayers on transwell plates. The untransfected or transfected cell monolayers were then incubated in an uptake medium containing 0 or 0.25 mmol L^(–1) of P as KH_(2) PO_(4) to measure the P absorption across duodenal epithelial cell monolayers. The results showed that among the siRNAs designed, si-1372 and si-890 were demonstrated to be the most effective in inhibiting the NaPIIb and PiT2 expressions, respectively. Supplemental P increased(P=0.065) the protein abundance of PiT2 and enhanced(P<0.0001) P absorption in primary cultured duodenal epithelial cell of chick embryos. Furthermore, NaPIIb silencing decreased(P=0.07) P absorption across duodenal epithelial cell monolayers, while PiT2 silencing had no effect(P=0.345). It is concluded that the NaP-IIb, but not PiT2, might be directly involved in the P absorption of chick duodenal epithelial cells.