本文選題：亞急性瘤胃酸中毒 + 鈉離子耦合單羧酸轉(zhuǎn)運(yùn)蛋白1��； 參考：《吉林大學(xué)》2017年碩士論文

【摘要】：亞急性瘤胃酸中毒(SARA)是公認(rèn)的高產(chǎn)奶牛消化系統(tǒng)性疾病,因其危害奶牛健康和生產(chǎn)性能而受到重視。在規(guī)�；B(yǎng)殖條件下,為滿足高產(chǎn)奶牛對(duì)能量的需求,過(guò)度飼喂精飼料,使短鏈脂肪酸(SCFA)在瘤胃內(nèi)的產(chǎn)生速率超過(guò)吸收速率,從而引起SCFA在瘤胃內(nèi)大量蓄積,導(dǎo)致瘤胃p H下降,并增加SARA發(fā)生的風(fēng)險(xiǎn)。SCFA是反芻動(dòng)物重要的能量代謝基底物,其在機(jī)體內(nèi)代謝產(chǎn)生大量的能量,因此,研究SCFA在瘤胃內(nèi)的轉(zhuǎn)運(yùn)和吸收機(jī)制,對(duì)反芻動(dòng)物能量供應(yīng)具有至關(guān)重要的作用,然而,目前SCFA在瘤胃內(nèi)的轉(zhuǎn)運(yùn)和吸收機(jī)制尚不清楚。在人和鼠的腸上皮細(xì)胞,鈉離子耦合單羧酸轉(zhuǎn)運(yùn)蛋白1(SLC5A8)和氫離子耦合單羧酸轉(zhuǎn)運(yùn)蛋白1(MCT1)對(duì)SCFA的吸收和轉(zhuǎn)運(yùn)至關(guān)重要,然而對(duì)于奶牛SLC5A8和MCT1的研究卻鮮有報(bào)道。本研究通過(guò)體內(nèi)實(shí)驗(yàn)和體外實(shí)驗(yàn)相結(jié)合的方法對(duì)SLC5A8和MCT1進(jìn)行了研究。在體內(nèi)試驗(yàn)(1)通過(guò)Western blot和實(shí)時(shí)熒光定量PCR檢測(cè)SLC5A8和MCT1在犢牛和成年牛消化道內(nèi)的分布和表達(dá),結(jié)果表明,SLC5A8和MCT1在犢牛和成年牛的消化道內(nèi)均有表達(dá)和分布,其中瘤胃的表達(dá)水平顯著高于其它部位,且SLC5A8和MCT1在成年牛的大腸中的表達(dá)顯著升高;(2)為了確定SLC5A8和MCT1的表達(dá)變化與奶牛健康的關(guān)系,檢測(cè)了SLC5A8和MCT1在正常奶牛和SARA奶牛瘤胃內(nèi)的表達(dá)變化,結(jié)果顯示,SARA奶牛抑制了SLC5A8和MCT1的表達(dá);(3)通過(guò)免疫組織化學(xué)的方法檢測(cè)SLC5A8和MCT1在瘤胃組織中的定位,發(fā)現(xiàn)SLC5A8主要分布于近管腔側(cè)的瘤胃上皮組織中,MCT1主要位于瘤胃上皮的基底層。為了確定SLC5A8和MCT1的表達(dá)在SARA奶牛被抑制的原因,體外分離培養(yǎng)犢牛瘤胃上皮細(xì)胞,根據(jù)正常和SARA奶牛瘤胃內(nèi)SCFA的濃度,將SCFA分為SARA組(乙酸90 m M、丙酸40 m M和丁酸30 m M)和對(duì)照組(乙酸60 m M,丙酸30 m M和丁酸20 m M)處理瘤胃上皮細(xì)胞,發(fā)現(xiàn)低濃度的SCFA促進(jìn)SLC5A8和MCT1的表達(dá),高濃度的SCFA抑制SLC5A8和MCT1的表達(dá),這與瘤胃體內(nèi)的結(jié)果一致。為了進(jìn)一步確定影響SLC5A8和MCT1表達(dá)的具體因素,用不同濃度的乙酸(0,30,60和90 m M)、丙酸(0,20,30和40 m M)和丁酸(0,10,20和30 m M)處理瘤胃上皮細(xì)胞,結(jié)果顯示,低濃度的丙酸和丁酸促進(jìn)SLC5A8和MCT1的表達(dá),高濃度的丙酸和丁酸抑制SLC5A8和MCT1的表達(dá);乙酸對(duì)SLC5A8的表達(dá)具有促進(jìn)作用,并隨濃度的升高而逐漸上升,然而高濃度的乙酸抑制了MCT1的表達(dá)。另外,通過(guò)瘤胃上皮細(xì)胞免疫熒光實(shí)驗(yàn),發(fā)現(xiàn)SLC5A8和MCT1主要位于細(xì)胞膜的表面,其主要通過(guò)跨膜的方式進(jìn)行轉(zhuǎn)運(yùn)。綜上所述,這些結(jié)果表明高濃度的SCFA抑制SLC5A8在SARA奶牛的瘤胃中的表達(dá),降低SCFA的吸收,從而加重SARA。
[Abstract]:Subacute rumen acidosis (SARAA) is a widely recognized digestive system disease in dairy cows. In order to meet the energy demand of high-yield dairy cattle, the short chain fatty acid (SCFA) production rate in rumen exceeded the absorption rate in order to satisfy the energy demand of high-yield dairy cows, which led to the accumulation of SCFA in rumen. SARA is an important energy metabolic substrate in ruminants, which produces a large amount of energy in the body. Therefore, the mechanism of SCFA transport and absorption in rumen is studied. However, the mechanism of SCFA transport and absorption in rumen is still unclear. In human and mouse intestinal epithelial cells, sodium ion-coupled monocarboxylate transporter (1SLC5A8) and hydrogen ion-coupled monocarboxylic acid transporter (1MCT1) play an important role in the absorption and transport of SCFA. However, there are few studies on SLC5A8 and MCT1 in dairy cattle. In this study, SLC5A8 and MCT1 were studied by in vivo and in vitro experiments. The distribution and expression of SLC5A8 and MCT1 in the digestive tract of calves and adult cattle were detected by Western blot and real-time fluorescence quantitative PCR. The results showed that SLC5A8 and MCT1 were expressed and distributed in the digestive tract of calves and adult cattle. In order to determine the relationship between the expression of SLC5A8 and MCT1 and the health of cows, the expression of SLC5A8 and MCT1 in the large intestine of adult cattle was significantly higher than that in other parts. The expression of SLC5A8 and MCT1 in the rumen of normal cows and SARA cows were detected. The results showed that the expression of SLC5A8 and MCT1 was inhibited in Sara cows. The localization of SLC5A8 and MCT1 in rumen tissues was detected by immunohistochemical method. It was found that SLC5A8 was mainly located in the basal layer of rumen epithelium in the proximal lumen of rumen epithelium. In order to determine the reason for the inhibition of SLC5A8 and MCT1 expression in SARA cows, calf rumen epithelial cells were isolated and cultured in vitro, and the concentrations of SCFA in the rumen of normal and SARA cows were determined. SCFA was divided into SARA group (acetic acid 90 mm, propionic acid 40 mm and butyric acid 30 mm) and control group (acetic acid 60 mm, propionic acid 30 mm and butyric acid 20 mm). It was found that low concentration of SCFA promoted the expression of SLC5A8 and MCT1. High concentration of SCFA inhibits the expression of SLC5A8 and MCT1, which is consistent with the results in rumen. In order to further determine the specific factors that affect the expression of SLC5A8 and MCT1, the rumen epithelial cells were treated with different concentrations of 30 and 90 mg / L acetate, 2030 and 40 mm propionate, and 1020 and 30 mm butyrate, respectively. Low concentration of propionic acid and butyric acid promoted the expression of SLC5A8 and MCT1, high concentration of propionic acid and butyric acid inhibited the expression of SLC5A8 and MCT1, acetic acid promoted the expression of SLC5A8 and increased gradually with the increase of concentration. However, high concentration of acetic acid inhibited the expression of MCT1. In addition, it was found that SLC5A8 and MCT1 were mainly located on the surface of the cell membrane and transported mainly through the membrane through the immunofluorescence assay of rumen epithelial cells. In conclusion, these results suggest that high concentration of SCFA inhibits the expression of SLC5A8 in the rumen of SARA cows and decreases the absorption of SCFA, thus exacerbating SARAA.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S858.23

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