Effects Of Dietary Calcium And Vitamin On The Growth And Mineralization Of Both Sternum And Tibia Bone In Meat Ducks And Underlying Mechanism

In this study,five studies were carried out to study on the morphology and mineralization of the tibia and sternum in meat ducks,and further investigate the effects of dietary nutrient density,calcium(Ca)and vitamin levels,especially 25-hydroxycholecalciferol(25-OH-D₃),on bone mass of tibia or/and sternum and its mechanism for meat-type poultry.Expt.1 Study on the morphology and mineralization of the tibia and sternum in meat ducksThis study aimed to understand how gait has been altered by BW and to characterize the basic physiological data on morphological and mineralized properties of the tibia and sternum of meat ducks.In experiment(Expt.)1,144 1-day-old ducks were allocated into eight pens(18 ducks/pen)and fed with the same diets until the age of 56 days.Eight birds(1 duck/pen)were weighted and selected according to average BW weekly for tibia characteristics.The body weight(BW)of ducks was increased with age throughout the whole study.Tibial length and width rapidly grew from 1 to 35 days,when it leveled off.The fat-free weight,ash,Ca,and phosphorus(P)content of tibia increased rapidly from 1 to 42 days and reached its plateau from42 days onwards.Tibial density and tibia-breaking strength increased in a regular manner with age until 42 days.When compared with BW,tibial fat-free weight and tibia-breaking strength increased isometrically,tibia length displayed a negative allometry,and tibial width showed a positive allometric growth,indicating that intensive selection for growth rate in ducks has resulted in tibial morphology changes,which represents an interesting compromise of adaptation for increasing BW.In Expt.2,a total of 72 1-day-old male and 72 female ducks were grouped in 8-unit cages(9 females and 9 males)in terms of similar BW,and all birds fed with the same diets.Eight birds(1 duck/pen)were selected weekly for measuring BW and sternal characteristics during 35 to 63 days.Results show that the sternal wide rapid grew from 35 to 42 days and the value exhibited little change after 42 days,while the keel length and the sternal depth had not significantly changed until 49 days.The male ducks possessed a greater value in coracoid distance,sternum central distance,and length than female birds.The sternal fat-free weight and density increased with ducks'age following the“S-shape”and to plateau around 56 days.The sternal ash,Ca and P content increased with age and reached a plateau in 49 days.The sex had not notably effect on the mineral content of sternum.Collectively,the tibia displayed rapid bone growth from1 to 35 days and mineralization during the period of 1 to 42 days,whereas the rapid growth and mineralization of sternum occurred for 35-49 days and 42-49days,respectively,and subsequently both achieved the plateau phase after 49 days.Expt.2 Effect of graded Ca supplementation in low nutrient density feed on sternum and tibia composition and bone turnover in meat ducksGenetic progress and increasing nutrient density for greater body mass and meat yield in poultry has inadvertently led to an imbalance between muscle mass and bone development which may or may not be detrimental to productivity and welfare.Slowing weight gain while promoting bone mineralization could positively influence skeletal health.Thus,this study aimed to evaluate the effect of graded Ca supplementation in low nutrient density(LND)diets on the bone mass and bone turnover of both sternum and tibia in meat ducks.Male meat ducks(720,15-day-old)were randomly assigned and fed a standard nutrient density positive control(PC)diet,and 4 LND diets with 0.5,0.7,0.9,and 1.1%Ca.Apparent metabolic energy(AME)was reduced to 11.1 and10.27 MJ/kg in the LND diet at 15-35 days and 36-56 days,respectively,as compared to PC diet,while maintaining proportionate essential nutrient relative to energy as in the PC diet.The results showed that the 0.9%Ca LND diet significantly decreased duck's BW and feed efficiency as compared with PC diet.At 35-day-old,ducks fed PC diets and 0.5%Ca LND diets had higher incidence of tibial dyschondroplasia(TD)than other groups.When compared with 0.5%Ca LND diet,LND diets with 0.7%or more Ca significantly improved tibia composition,microarchitecture,and mechanical properties,and consequently decreased the incidence of TD.Furthermore,LND diets with 0.7%or more Ca increased osteocyte-specific gene m RNA expression,and it blocked the expression of osteoblast differentiation marker genes including osteocalcin and collagen type 1(Coll-I),they also deceased the expression of osteoclast differentiation genes,such as vacuolar H⁺-ATPases(V-ATPases),cathepsin K and receptor activator of NF-?b(RANK).Meanwhile,serum bone markers such as ALP,osteocalcin,both osteoblast markers,and tartrate-resistant acid phosphatase(TRAP),an osteoclast marker,were significantly decreased in at least 0.7%Ca treated groups.Regarding the sternum,although the 0.9%Ca LND diet decreased sternal dimension,it increased trabecular bone volume/total volume(BV/TV)and Ca content of the sternum compared with PC diet.Feeding 0.7%or more Ca with the LND diet significantly increased the mineral content,bone density,BV/TV and trabecular number(Tb.N)of the sternum in 49-day-old ducks.Furthermore,the LND diet with0.7%or more Ca increased osteocyte-specific gene m RNA and osteoprotegerin(OPG)expression,and it blocked the expression of cathepsin K.TRAP staining also revealed that the addition 0.7%or more Ca to the LND diet significantly decreased the number of osteoclasts compared with the 0.5%Ca LND diet.Meanwhile serum TRAP activity was significantly decreased in 0.7%or more Ca treated groups.We concluded that LND diet with 0.7%or more Ca may maintain optimal bone mass of both sternum and tibia through suppressing bone resorption for meat ducks.Expt.3 Effects of commercial premix vitamin level on sternum growth and calcification in meat ducksThe present study was conducted to investigate the effects of dietary vitamin level on sternum growth and calcification in meat ducks.A total of 432 1-day-old mixed-sex Cherry Valley ducks(216 males and 216 females)was randomly allocated 3 treatments with 8 replicates(18 ducks/pen)and fed low-vitamin level diet(70%NRC vitamin regimen),high-vitamin level diet(DSM vitamin regimen)or medium-vitamin level diet(50%low-vitamin level diet and 50%high-vitamin level diet).Sternum and serum were harvested after 49 days of feeding.The results showed that dietary high-vitamin level increased BW at 49 days(P<0.05).Compared with the low-vitamin level group,the medium and high-vitamin level diets decreased the sternum length and depth(P<0.05).However,medium-and high-vitamin level increased sternal fat-free weight,calcified ratio,density,ash,and Ca concentration(P<0.05).Likewise,high-vitamin level improved serum Ca and P content(P<0.05)and declined serum ALP activity(P<0.05)compared with the low-vitamin level group.Our study indicates that diets with high-vitamin level have a positive effect on the growth performance and sternum calcification of 49-day-old meat ducks.Expt.4 Effect of dietary 25-OH-D₃ on the tibia and sternum mass of meat ducks under different vitamin regimensBone problems often result from the rapid weight gain and poor bone quality in modern ducks,leading to high risk of fractures and continuous pain.We hypothesized that increasing bone quality in combination with delaying weight gain via an LND diet probably reverses these skeletal abnormalities.Studies indicated that 25-OH-D₃,a vitamin D₃ metabolite,is effective in treating bone-related disorders.Therefore,the experiment was conducted to investigate the effects of 25-OH-D₃ on tibia and sternum mass of meat ducks.1024 male meat ducklings were divided into 8 groups(8replicates pens;16 ducks/pen)and fed two standard nutrient density diets(containing NRC vitamin regimen)with or without 0.069 mg/kg 25-OH-D₃,and six LND diets made up of a 2×3 factorial design including 3 kinds of vitamin premixes(NRC,NY/T and CVF),and with or without 0.069 mg 25-OH-D₃/kg for 56 days.PC and LND diets were designed according to Expt.2.In standard nutrient density diets,25-OH-D₃ addition increased the BW at 14 days,gain(1-14 days),feed intake(FI)and the ratio of feed intake and weight gain(F:G)(15-35 days and 15-56 days).In LND diets,birds fed NY/T and CVF vitamin diets displayed higher BW and gain(15-35days and 15-56 days),while 25-OH-D₃ supplementation did not notably change growth performance expect for decreasing the FI and F:G for 15-35 days and 15-56days.In standard nutrient density diets,25-OH-D₃ addition improved the mineral content,microarchitecture,and mechanical properties of tibias,and this was accompanied by decreased serum bone resorption marker and a concomitant decrement in expression of osteoclast-specific marker genes expression at 35 days.Analogously,25-OH-D₃ treatment also notably improved sternal mineral accumulation and microarchitecture,along with decreasing serum bone turnover markers levels and increasing serum Ca concentration in 49-day-old birds.In LND diets,birds fed NY/T vitamin diets exhibited higher tibia minerals deposition and lower bone turnover rate that those fed NRC vitamin diets.25-OH-D₃supplementation increased tibial mineral content,microarchitecture,and strength of birds under an NRC rather than NY/T vitamin regimen.At 49 days of age,regardless of 25-OH-D₃ supplementation,NY/T vitamin regimen inhibited bone turnover,and consequently increased sternal trabecular bone volume and mineral deposition compared with NRC vitamin premix.Likely,CVF vitamin diets also increased sternal bone volume at 49 days as compared to NRC vitamin diets.Supplementing 25-OH-D₃ to NRC but not NY/T vitamin regimen significantly improved sternal microarchitecture and mineral content,which was accompanied by decreased serum bone resorption markers concentration,as well as down-regulation the gene expressions of osteoclast differentiation and activity.CVF vitamin with 25-OH-D₃diets similarly displayed lower bone resorption and higher serum Ca level,and improved microarchitecture.To conclude,dietary administration of 25-OH-D₃increased tibia and sternum mass in NRC vitamin diet by suppressing bone resorption in 35 or 49 days of age meat ducks,respectively,and this positive impact was only observed in NRC but not NY/T vitamin regimens when birds were fed an LND diet.Expt.5 25-OH-D₃ reverses heat induced alterations in bone quality in finisher broilers associated with effects on intestinal integrity and inflammationAlterations in ambient temperature have been associated with multiple detrimental effects on broilers such as intestinal barrier disruption and dysbiosis resulting in systemic inflammation.Inflammation and 25-OH-D₃ have shown to play a negative and positive role,respectively,in the regulation of bone mass.Hence the potential of 25-OH-D₃ in alleviating heat induced bone alterations and its mechanisms was studied.At15 days of age,birds were allotted to either thermoneutral conditions;control(ad libitum feeding basal diets)and pair feeding(restrictive feeding according to FI of heat-stressed birds of basal diets),or room with heat stress,i.e.,daily 7 h at 34°C with relative humidity(RH)between 50 and 60%and rest of day 26°C with no RH control;group fed basal diets(ad libitum feeding of basal diets)and fed basal diets supplemented with0.069 mg/kg 25-OH-D₃(ad libitum).Performance was monitored,and various physiological parameters were measured at the end of the study.Heat stress directly induced a decrease in tibia material properties and bone mass,as demonstrated by lower mineral content,and heat stress caused a notable increase in intestinal permeability.Treatment with dietary 25-OH-D₃ reversed the heat stress-induced bone loss and barrier leak.Broilers suffering from heat stress exhibited dysbiosis and increased expression of inflammatory cytokines in ileum and bone marrow,as well as increased osteoclast number and activity.Changes that were prevented by dietary 25-OH-D₃ administration.Specifically,dietary 25-OH-D₃ addition decreased abundance of B-and T-cells in blood,and the expression of inflammatory cytokines,especially TNF-?,in both ileum and bone marrow,but did not alter the diversity and repopulation of major bacterial phyla.With regard to bone remodeling,dietary 25-OH-D₃ supplementation was linked to a decrease in serum C-terminal telopeptide of type I collagen(CTx)reflecting bone resorption and a concomitant decrement in osteoclast-specific marker genes expression(e.g.cathepsin K),whereas it did not apparently change serum bone formation markers during heat stress.These data underscore the damage of heat stress to intestinal integrity and bone health,as well as that dietary 25-OH-D₃ supplementation was identified as a potential therapy for preventing these adverse effects.Conclusion1.The tibia displayed rapid bone growth(1 to 35d)and mineralization(1 to 42d),whereas the age of rapid growth and mineralization occurred in 1-49 days and 42-29days,respectively,for sternum in meat ducks.2.LND diets with 0.7%or more Ca decreased bone turnover,which subsequently increased tibial quality for 35-day-old meat ducks,and LND diet with 0.7%or more Ca also may suppress bone resorption and maintain optimal sternal mass for 49-d-old meat ducks.3.A high vitamin level diet,such as DSM(2016)vitamin diets,can benefit BW and sternum calcification of 49-day-old meat ducks.4.No matter what nutrient density,25-OH-D₃ treatment increased tibial and sternal mass in NRC vitamin regimen by suppressing bone resorption and inducing Ca absorption for meat ducks,and NY/T vitamin regimen made a favorable effect on the sternal and tibial mass of duck in LND diet.5)Dietary supplementation with 25-OH-D₃ could also improve intestinal integrity and depressing bone resorption through decreasing inflammatory response,and/or through directly modulating osteoclast to increase tibia quality during heat stress.