添加丙酸鈣對奶公犢生長性能的影響和調(diào)控瘤胃發(fā)育機(jī)理的研究
發(fā)布時(shí)間：2018-08-14 14:54
【摘要】：本課題研究了丙酸鈣添加水平對奶公犢生長性能的影響和調(diào)控瘤胃發(fā)育的機(jī)理。通過不同時(shí)間的飼養(yǎng)屠宰試驗(yàn),研究了丙酸鈣添加水平對犢牛生長性能、內(nèi)臟器官以及胃腸道發(fā)育的影響；通過對丙酸鈣影響瘤胃發(fā)酵、血液生化指標(biāo)和瘤胃上皮相關(guān)mRNA基因表達(dá)量的研究,闡明了丙酸通過內(nèi)分泌途徑和G蛋白偶聯(lián)受體途徑促進(jìn)瘤胃上皮發(fā)育的機(jī)理；應(yīng)用基于轉(zhuǎn)錄組學(xué)技術(shù)篩選了丙酸對瘤胃上皮差異表達(dá)的基因,為調(diào)控犢牛瘤胃發(fā)育提供了更為全面的理論框架和研究基礎(chǔ)。試驗(yàn)1：研究了丙酸鈣添加水平對不同飼喂階段奶公犢生長性能、內(nèi)臟器官以及胃腸道發(fā)育的影響。選用娟姍公犢牛54頭(日齡7±1d,體重23.1±1.2kg),隨機(jī)分成3個(gè)處理：1)對照組不添加丙酸鈣(CON)；2)添加5%丙酸鈣(5%CaP,DM基礎(chǔ))；3)添加10%丙酸鈣(10%CaP,DM基礎(chǔ))。試驗(yàn)期160 d,分為3階段：第1階段(飼喂0d-30d),第2階段(飼喂30d-90d),第三階段(飼喂90d～160d)。在每個(gè)階段結(jié)束時(shí),隨機(jī)從每個(gè)處理組中選擇6頭犢牛屠宰并取樣。添加丙酸鈣能夠提高犢牛飼喂90d(P=0.013)、120d(P=0.018)以及160d(P=0.006)的體重。飼喂90d的犢牛體重隨著丙酸鈣添加水平的提高而提高,飼喂120d和160d的犢牛體重在丙酸鈣的添加水平之間沒有顯著性差異。10%CaP組顯著提高了30d～60d(P=0.013)、60d～90d(P=0.013)和120d～160d(P=0.080)的平均日增重,而5%CaP組只提高了120d～160d的犢牛平均日增重(P=0.046)。10%CaP組脾臟重量在飼喂160天時(shí)顯著高于CON組和5%CaP組(P=0.010)。5%CaP和10%CaP組肝臟重量在飼喂90d時(shí)都顯著高于CON組(P=0.003),而在160d時(shí),只有10%CaP組肝臟重量顯著高于CON組(P=0.037)。添加丙酸鈣可以提高犢牛飼喂90d和160d的瘤胃重量(P=0.006,0.010),提高十二指腸絨毛高度(P=0.007)和隱窩深度(P=0.0002),提高空腸絨毛高度(P=0.0037)、隱窩深度(P=0.0065)以及絨毛高度與隱窩深度的比例(P=0.0080),并提高回腸的絨毛高度(P0.001)和隱窩深度(P0.001)。小結(jié)：添加丙酸鈣可以提高犢牛的生長性能、促進(jìn)肝臟和脾臟等內(nèi)臟器官以及胃腸道的發(fā)育。丙酸鈣添加量在飼喂90d以前添加10%效果較好,而在飼喂90d～160d期間,添加5%的丙酸鈣效果與10%效果相當(dāng)。試驗(yàn)2：研究了丙酸鈣添加水平對不同飼喂階段犢牛瘤胃發(fā)酵以及瘤胃發(fā)育的影響。試驗(yàn)動(dòng)物和處理同試驗(yàn)1。添加丙酸鈣可以顯著提高犢牛飼喂30d瘤胃內(nèi)總揮發(fā)酸含量(P=0.0370),并有提高乙酸(P=0.0632)、丙酸(P=0.0903)和異丁酸(P=0.0591)濃度的趨勢。10%CaP組降低了犢牛飼喂90d瘤胃內(nèi)氨態(tài)氮的含量(P=0.0013)。添加5%丙酸鈣可以提高飼喂160d犢牛瘤胃內(nèi)丙酸(P=0.0029)和戊酸的含量(P=0.0133)。添加丙酸鈣提高了犢牛飼喂30d(P=0.0085)和90d(P=0.0126)的瘤胃壁厚度,提高了飼喂30d犢牛瘤胃乳頭的長度(P=0.0012),且隨著丙酸鈣添加量的增加,瘤胃乳頭長度加大,但對乳頭寬度沒有影響(P=0.2754)。提高丙酸鈣添加水平可以增加飼喂90d犢牛瘤胃上皮乳頭的長度(P=0.0016)和上皮乳頭的寬度(P=0.0293),其中添加5%丙酸鈣可以提高飼喂160d犢牛瘤胃上皮乳頭的長度(P0.001),而添加10%的丙酸鈣可以提高瘤胃上皮乳頭的寬度(P=0.0025)。小結(jié)：添加丙酸鈣能夠促進(jìn)瘤胃發(fā)酵,促進(jìn)瘤胃上皮乳頭的發(fā)育。瘤胃上皮乳頭的長度與瘤胃內(nèi)的丙酸含量呈正相關(guān)關(guān)系。在飼喂90d以前添加10%丙酸鈣對犢牛瘤胃發(fā)酵和瘤胃發(fā)育的效果較好,飼喂90d到160天期間添加5%的丙酸鈣效果更好。試驗(yàn)3：研究了丙酸鈣添加水平對犢牛血液生化指標(biāo)和瘤胃上皮基因表達(dá)量的影響。試驗(yàn)動(dòng)物和處理同試驗(yàn)1。在每個(gè)階段結(jié)束時(shí)前一天,隨機(jī)從每個(gè)處理組中選擇6頭犢牛頸靜脈采血,并且從飼喂160d的CON和5%CaP組隨機(jī)選擇3頭犢牛的瘤胃背囊部樣品,用于熒光定量PCR測定。添加丙酸鈣可以提高不同飼喂階段犢牛血糖含量,血糖含量隨著丙酸鈣添加水平的提高而增加。10%CaP組顯著提高了飼喂30d犢牛血糖含量(P0.05)。添加丙酸鈣對犢牛血清胰島素含量的影響和血糖一致。添加丙酸鈣可以在數(shù)值上提高犢牛血清中1GF-1的濃度,其中在飼喂160d時(shí),5%CaP組濃度最高。添加5%丙酸鈣可以提高瘤胃上皮CCND1、CDK4、GPCR43、IGF-1R和INSR的基因表達(dá)量,其中CCND1和GPCR43都達(dá)到了顯著性水平。小結(jié)：添加丙酸鈣一方面可以提高血液中的葡萄糖、胰島素和1GF-1的含量,進(jìn)而提高瘤胃上皮INSR和IGF-1R基因表達(dá)量：另一方面丙酸可以直接作為信號分子通過G蛋白偶聯(lián)受體途徑提高GPCR43的基因表達(dá)量,來直接促進(jìn)瘤胃上皮的發(fā)育。試驗(yàn)4：通過高通量轉(zhuǎn)錄組測序(RNA-Seq)方法篩選添加丙酸鈣對犢牛瘤胃上皮差異表達(dá)的基因,探討丙酸對瘤胃上皮發(fā)育影響的潛在分子機(jī)制。從試驗(yàn)一中CON和5%CaP處理組中隨機(jī)選擇3頭飼喂160d的犢牛屠宰后取得的瘤胃上皮作為研究對象,經(jīng)過樣品總RNA提取、轉(zhuǎn)錄組文庫的制備,然后在HiSeq2500測序平臺上測序。樣本匹配到�；蚪M的基因共有18084個(gè),其中只在5%CaP組表達(dá)的基因有718個(gè),只在CON組表達(dá)的基因有696個(gè)。在兩個(gè)處理組中均有表達(dá)的基因有16670個(gè),顯著差異表達(dá)的基因數(shù)量為369個(gè),其中5%CaP組比CON組高表達(dá)的基因有216個(gè),低表達(dá)的基因有153個(gè)。通過GO富集分析發(fā)現(xiàn),添加丙酸鈣可以上調(diào)與上皮細(xì)胞發(fā)育相關(guān)的TGM3、TGM1、OVOL1、DSP、IVL、SPINK5、CLDN4、CNFN、KRT36基因的表達(dá)量促進(jìn)瘤胃上皮的發(fā)育。KEGG結(jié)果分析表明,差異基因主要通過代謝、信號轉(zhuǎn)導(dǎo)以及內(nèi)分泌系統(tǒng)影響瘤胃上皮的發(fā)育。小結(jié)：本試驗(yàn)成功篩選了丙酸鈣調(diào)控犢牛瘤胃上皮發(fā)育的候選基因,為調(diào)控犢牛瘤胃上皮發(fā)育提供了理論基礎(chǔ)以及更多的研究靶點(diǎn)和思路。
[Abstract]:The effects of calcium propionate on growth performance and rumen development of dairy male calves were studied. The effects of calcium propionate on growth performance, visceral organs and gastrointestinal development of dairy male calves were studied by feeding and slaughtering experiments at different times. The study on the expression of rumen epithelium-related mRNA genes clarified the mechanism of propionic acid promoting the development of rumen epithelium through endocrine pathway and G protein-coupled receptor pathway; the differentially expressed genes of rumen epithelium were screened by transcriptome-based technology, which provided a more comprehensive theoretical framework and research for the regulation of rumen development of calves. Experiment 1: The effects of calcium propionate on growth performance, visceral organs and gastrointestinal development of male calves at different feeding stages were studied. Fifty-four Juanshan male calves (7 6550 At the end of each stage, six calves were randomly selected from each treatment group for slaughter and sampling. Calcium propionate could increase the feeding time of calves by 90 days (P = 0.013) and 120 days (P = 0.018). The body weight of the calves fed 90 days increased with the increase of calcium propionate, and there was no significant difference in the body weight of the calves fed 120 days and 160 days. The body weight of the calves fed 10% CaP increased significantly by 30 days to 60 days (P = 0.013), 60 days to 90 days (P = 0.013) and 120 days to 160 days (P = 0.080). The average daily gain (P = 0.046) of calves increased only 120 days to 160 days. The spleen weight of 10% CaP group was significantly higher than that of CON group and 5% CaP group at 160 days (P = 0.010). The liver weight of 5% CaP and 10% CaP group was significantly higher than that of CON group at 90 days (P = 0.003) and only 10% CaP group was significantly higher than that of CON group at 160 days (P = 0.037). The rumen weight (P=0.006,0.010) and the height of duodenal villi (P=0.007) and the depth of crypt (P=0.0002) were increased, the height of jejunal villi (P=0.0037), the depth of crypt (P=0.0065) and the ratio of the height of villi to the depth of crypt (P=0.0080) were increased, and the height of ileal villi (P=0.001) and the depth of crypt (P=0.001) were also increased. Summary: Calcium propionate can improve the growth performance of calves, promote the development of liver, spleen and other visceral organs as well as gastrointestinal tract. Calcium propionate supplementation was better when 10% was added before 90 days, but the effect of adding 5% calcium propionate was similar to that of 10% during 90 days to 160 days. The effect of calcium propionate on rumen fermentation and rumen development of calves at the same feeding stage was the same as that of experimental animals and treatments. 1. Calcium propionate significantly increased the content of total volatile acids (P = 0.0370) in the rumen of calves fed 30 days, and increased the concentration of acetic acid (P = 0.0632), propionic acid (P = 0.0903) and isobutyric acid (P = 0.0591) in the rumen of calves. The contents of propionic acid (P = 0.0029) and valeric acid (P = 0.0133) in rumen were increased by adding 5% calcium propionate (P = 0.0013). Calcium propionate increased the thickness of rumen wall and the length of rumen papilla (P = 0.0012) of calves fed with calcium propionate for 30 days (P = 0.0085) and 90 days (P = 0.0126). The length of rumen papilla and the width of epithelial papilla increased with the increase of calcium propionate (P = 0.0016) and 5% calcium propionate (P = 0.0293), but the length of rumen papilla increased with the increase of calcium propionate (P = 0.2754). 10% calcium propionate could increase the width of rumen papilla (P = 0.0025). Summary: Calcium propionate could promote rumen fermentation and the development of rumen papilla. The length of rumen papilla was positively correlated with the content of propionic acid in rumen. Experiment 3: The effects of calcium propionate supplementation on blood biochemical indexes and rumen epithelial gene expression in calves were studied. The animals and treatments were the same as experiment 1. Six calves were randomly selected from each treatment group one day before the end of each stage. Blood samples from the rumen backpack of three calves were randomly selected from the CON and 5% CaP groups fed for 160 days and used for quantitative fluorescence PCR determination. Calcium propionate could increase the concentration of 1GF-1 in calf serum, and the concentration of 5% CaP was the highest at 160 days. Adding 5% calcium propionate could increase the expression of CCND1, CDK4, GPCR43, IGF-1R and INSR genes in rumen epithelium, including CCND1 and GPCR43. Conclusion: On the one hand, calcium propionate can increase the levels of glucose, insulin and 1GF-1 in the blood, and then increase the expression of INSR and IGF-1R genes in the rumen epithelium. On the other hand, propionic acid can be directly used as a signal molecule to increase the expression of GPCR43 gene through G protein-coupled receptor pathway, thus directly promoting the tumor. Gastric epithelial development. Experiment 4: Genes differentially expressed in rumen epithelium of calves were screened by high-throughput transcriptome sequencing (RNA-Seq) to explore the potential molecular mechanism of the effects of calcium propionate on rumen epithelial development. Three calves were randomly selected from the CON and 5% CaP treatment groups in trial 1 after slaughtering for 160 days. The total RNA was extracted from the samples, the transcriptome library was prepared, and then sequenced on the HiSeq2500 sequencing platform. The number of differentially expressed genes was 369, of which 216 were overexpressed in 5% CaP group and 153 were underexpressed in CON group. The results of GO enrichment analysis showed that calcium propionate could up-regulate the expression of TGM3, TGM1, OVOL1, DSP, IVL, SPINK5, CLDN4, CNFN and KRT36 genes related to epithelial cell development. KEGG analysis showed that the differentially expressed genes affected the development of rumen epithelium mainly through metabolism, signal transduction and endocrine system.
【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：S823.5
本文編號：2183214
[Abstract]:The effects of calcium propionate on growth performance and rumen development of dairy male calves were studied. The effects of calcium propionate on growth performance, visceral organs and gastrointestinal development of dairy male calves were studied by feeding and slaughtering experiments at different times. The study on the expression of rumen epithelium-related mRNA genes clarified the mechanism of propionic acid promoting the development of rumen epithelium through endocrine pathway and G protein-coupled receptor pathway; the differentially expressed genes of rumen epithelium were screened by transcriptome-based technology, which provided a more comprehensive theoretical framework and research for the regulation of rumen development of calves. Experiment 1: The effects of calcium propionate on growth performance, visceral organs and gastrointestinal development of male calves at different feeding stages were studied. Fifty-four Juanshan male calves (7 6550 At the end of each stage, six calves were randomly selected from each treatment group for slaughter and sampling. Calcium propionate could increase the feeding time of calves by 90 days (P = 0.013) and 120 days (P = 0.018). The body weight of the calves fed 90 days increased with the increase of calcium propionate, and there was no significant difference in the body weight of the calves fed 120 days and 160 days. The body weight of the calves fed 10% CaP increased significantly by 30 days to 60 days (P = 0.013), 60 days to 90 days (P = 0.013) and 120 days to 160 days (P = 0.080). The average daily gain (P = 0.046) of calves increased only 120 days to 160 days. The spleen weight of 10% CaP group was significantly higher than that of CON group and 5% CaP group at 160 days (P = 0.010). The liver weight of 5% CaP and 10% CaP group was significantly higher than that of CON group at 90 days (P = 0.003) and only 10% CaP group was significantly higher than that of CON group at 160 days (P = 0.037). The rumen weight (P=0.006,0.010) and the height of duodenal villi (P=0.007) and the depth of crypt (P=0.0002) were increased, the height of jejunal villi (P=0.0037), the depth of crypt (P=0.0065) and the ratio of the height of villi to the depth of crypt (P=0.0080) were increased, and the height of ileal villi (P=0.001) and the depth of crypt (P=0.001) were also increased. Summary: Calcium propionate can improve the growth performance of calves, promote the development of liver, spleen and other visceral organs as well as gastrointestinal tract. Calcium propionate supplementation was better when 10% was added before 90 days, but the effect of adding 5% calcium propionate was similar to that of 10% during 90 days to 160 days. The effect of calcium propionate on rumen fermentation and rumen development of calves at the same feeding stage was the same as that of experimental animals and treatments. 1. Calcium propionate significantly increased the content of total volatile acids (P = 0.0370) in the rumen of calves fed 30 days, and increased the concentration of acetic acid (P = 0.0632), propionic acid (P = 0.0903) and isobutyric acid (P = 0.0591) in the rumen of calves. The contents of propionic acid (P = 0.0029) and valeric acid (P = 0.0133) in rumen were increased by adding 5% calcium propionate (P = 0.0013). Calcium propionate increased the thickness of rumen wall and the length of rumen papilla (P = 0.0012) of calves fed with calcium propionate for 30 days (P = 0.0085) and 90 days (P = 0.0126). The length of rumen papilla and the width of epithelial papilla increased with the increase of calcium propionate (P = 0.0016) and 5% calcium propionate (P = 0.0293), but the length of rumen papilla increased with the increase of calcium propionate (P = 0.2754). 10% calcium propionate could increase the width of rumen papilla (P = 0.0025). Summary: Calcium propionate could promote rumen fermentation and the development of rumen papilla. The length of rumen papilla was positively correlated with the content of propionic acid in rumen. Experiment 3: The effects of calcium propionate supplementation on blood biochemical indexes and rumen epithelial gene expression in calves were studied. The animals and treatments were the same as experiment 1. Six calves were randomly selected from each treatment group one day before the end of each stage. Blood samples from the rumen backpack of three calves were randomly selected from the CON and 5% CaP groups fed for 160 days and used for quantitative fluorescence PCR determination. Calcium propionate could increase the concentration of 1GF-1 in calf serum, and the concentration of 5% CaP was the highest at 160 days. Adding 5% calcium propionate could increase the expression of CCND1, CDK4, GPCR43, IGF-1R and INSR genes in rumen epithelium, including CCND1 and GPCR43. Conclusion: On the one hand, calcium propionate can increase the levels of glucose, insulin and 1GF-1 in the blood, and then increase the expression of INSR and IGF-1R genes in the rumen epithelium. On the other hand, propionic acid can be directly used as a signal molecule to increase the expression of GPCR43 gene through G protein-coupled receptor pathway, thus directly promoting the tumor. Gastric epithelial development. Experiment 4: Genes differentially expressed in rumen epithelium of calves were screened by high-throughput transcriptome sequencing (RNA-Seq) to explore the potential molecular mechanism of the effects of calcium propionate on rumen epithelial development. Three calves were randomly selected from the CON and 5% CaP treatment groups in trial 1 after slaughtering for 160 days. The total RNA was extracted from the samples, the transcriptome library was prepared, and then sequenced on the HiSeq2500 sequencing platform. The number of differentially expressed genes was 369, of which 216 were overexpressed in 5% CaP group and 153 were underexpressed in CON group. The results of GO enrichment analysis showed that calcium propionate could up-regulate the expression of TGM3, TGM1, OVOL1, DSP, IVL, SPINK5, CLDN4, CNFN and KRT36 genes related to epithelial cell development. KEGG analysis showed that the differentially expressed genes affected the development of rumen epithelium mainly through metabolism, signal transduction and endocrine system.
【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：S823.5
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