代乳粉添加甘露寡糖對(duì)7—28日齡湖羊羔羊胃腸道發(fā)育的影響

鄭琛，李發(fā)弟，李飛，周巨旺，段鵬偉，劉繪匯，樊海苗，朱威力，劉婷

代乳粉添加甘露寡糖對(duì)7—28日齡湖羊羔羊胃腸道發(fā)育的影響

鄭琛1，李發(fā)弟2,3，李飛2，周巨旺1，段鵬偉1，劉繪匯1，樊海苗1，朱威力1，劉婷1

（1甘肅農(nóng)業(yè)大學(xué)動(dòng)物科學(xué)技術(shù)學(xué)院，蘭州 730070；2蘭州大學(xué)草地農(nóng)業(yè)科技學(xué)院，草地農(nóng)業(yè)生態(tài)系統(tǒng)國家重點(diǎn)實(shí)驗(yàn)室/ 農(nóng)業(yè)農(nóng)村部草牧業(yè)創(chuàng)新重點(diǎn)實(shí)驗(yàn)室，蘭州 730020；3甘肅省肉羊繁育生物技術(shù)工程實(shí)驗(yàn)室，甘肅民勤 733300）

【】探討代乳粉中添加甘露寡糖（mannan oligosaccharides，MOS）對(duì)7—28日齡湖羊羔羊胃腸道生長發(fā)育的影響。選擇同質(zhì)性良好的7日齡湖羊公羔（雙羔）30只，隨機(jī)分為2組，每組15只，每只為1個(gè)重復(fù)，對(duì)照組羔羊飼喂不含MOS的代乳粉，試驗(yàn)組羔羊飼喂含0.2 % MOS的代乳粉，試驗(yàn)期21d。羔羊28日齡時(shí)，兩個(gè)試驗(yàn)組各隨機(jī)選擇8只羔羊屠宰，取出消化道，稱量各胃室和腸段包含內(nèi)容物的質(zhì)量和凈質(zhì)量，量取各腸段長度，用以計(jì)算各部位的相對(duì)質(zhì)量和內(nèi)容物分布，以及各腸段的相對(duì)長度。多聚甲醛固定皺胃胃底腺區(qū)及十二指腸、空腸和回腸中段的組織樣品，測定組織形態(tài)和小腸上皮細(xì)胞凋亡率。采集十二指腸、空腸和回腸的黏膜樣品，測定緊密連接蛋白1 (claudin 1)、閉鎖小帶1（zonula occludens-1，ZO-1）和閉鎖蛋白（occludin）的mRNA表達(dá)量。除空腸相對(duì)長度外（%全腸長度，=0.040），MOS對(duì)羔羊胃腸指數(shù)（%活體質(zhì)量）、胃腸相對(duì)質(zhì)量（%全胃質(zhì)量、%全腸質(zhì)量和%全胃腸質(zhì)量）、腸道相對(duì)長度（%全腸長度）、內(nèi)容物相對(duì)活體質(zhì)量（%活體質(zhì)量）、胃腸內(nèi)容物相對(duì)總胃/腸內(nèi)容物及總胃腸內(nèi)容物相對(duì)質(zhì)量（%總胃內(nèi)容物、總腸內(nèi)容物、總胃腸內(nèi)容物）、小腸上皮細(xì)胞凋亡率和小腸黏膜claudin 1蛋白mRNA的表達(dá)量均沒有產(chǎn)生顯著影響（＞0.05），但MOS顯著提高羔羊十二指腸絨毛高度和肌層厚度并顯著降低絨毛寬度（=0.033，=0.047，=0.015），顯著上調(diào)空腸ZO-1蛋白mRNA表達(dá)量（=0.028），此外，MOS有提高羔羊回腸絨毛高度、絨毛寬度和隱窩深度、皺胃肌層厚度及回腸occludin蛋白mRNA表達(dá)量的趨勢（=0.075，=0.078，=0.085，=0.084，=0.052）。MOS對(duì)7—28日齡湖羊羔羊胃腸道相對(duì)質(zhì)量、長度和內(nèi)容物分布基本無顯著影響，但可改善十二指腸和回腸絨毛及肌層的組織形態(tài)，維持小腸屏障功能，有利于提高養(yǎng)分消化率。

羔羊；甘露寡糖；代乳粉；胃腸道；發(fā)育

0 引言

【研究意義】胃腸道的結(jié)構(gòu)是保障消化功能的前提，當(dāng)組織形態(tài)發(fā)育正常及功能完善時(shí)，胃腸道中的營養(yǎng)物質(zhì)才會(huì)被充分消化吸收[1]。反芻動(dòng)物胃腸道發(fā)育受多種因素影響和調(diào)節(jié)，如年齡[2]、斷奶[3-4]、飼糧類型[5-7]、腸營養(yǎng)素、激素和生長因子[8]等。飼喂幼畜時(shí)，除了需要提供高濃度的能量和養(yǎng)分以滿足幼畜快速生長發(fā)育和器官發(fā)育的需要[9-11]，還需要強(qiáng)化一些微量營養(yǎng)性和非營養(yǎng)性飼料添加劑以提高幼畜成活率和機(jī)體免疫力，益生菌和化學(xué)益生素就是目前幼齡動(dòng)物養(yǎng)殖中常用的免疫增強(qiáng)劑[12]。化學(xué)益生素被定義為營養(yǎng)活性物質(zhì)（Nutricine），是一種非藥品類功能食品，雖然不具備直接營養(yǎng)功能，但可以維持腸道消化吸收功能，因而增強(qiáng)動(dòng)物健康和生長發(fā)育[13]。【前人研究進(jìn)展】甘露寡糖（mannan oligosaccharides, MOS）是化學(xué)益生素的一種，來自于酵母（）細(xì)胞壁，富含甘露蛋白和β-葡聚糖等復(fù)雜碳水化合物等[14]，廣泛應(yīng)用于養(yǎng)殖業(yè)以提高動(dòng)物機(jī)體免疫機(jī)能并消除腸道病原菌[15-16]。在單胃動(dòng)物和水產(chǎn)動(dòng)物養(yǎng)殖中，添加MOS具有提高生產(chǎn)性能和促進(jìn)動(dòng)物健康的作用[17-20]，也促進(jìn)腸道發(fā)育，如前期和后期飼糧中分別添加0.2 %和0.1 % MOS可提高肉仔雞小腸絨毛高度并降低隱窩深度[21]，添加0.1 % MOS可顯著提高仔豬小腸黏膜絨毛高度/隱窩深度值（V/C）[22]，添加0.1 %、0.15 %和0.2 % MOS均可顯著提高兔小腸絨毛高度[23]?！颈狙芯壳腥朦c(diǎn)】MOS在反芻動(dòng)物上的應(yīng)用研究較少，主要是因?yàn)楹芏鄬W(xué)者認(rèn)為瘤胃微生物能夠降解MOS，從而消除其保健功能。然而，有限的資料仍然顯示MOS在反芻動(dòng)物養(yǎng)殖中起到了一定的有益作用，如改善綿羊瘤胃健康[24]和提高抗氧化能力[25]，提高羔羊血液免疫球蛋白水平[26]等，但MOS對(duì)幼齡反芻動(dòng)物消化道生長發(fā)育的影響鮮見報(bào)道。鑒于MOS在單胃動(dòng)物胃腸道發(fā)育中所表現(xiàn)出的積極作用，本研究提出假設(shè)，添加MOS對(duì)羔羊胃腸道發(fā)育也具有一定的影響，可為幼齡反芻動(dòng)物的健康養(yǎng)殖提供幫助?！緮M解決的關(guān)鍵問題】本試驗(yàn)以7日齡湖羊公雙羔作為試驗(yàn)對(duì)象，研究代乳粉中添加MOS對(duì)羔羊消化道生長發(fā)育的影響，為幼齡反芻動(dòng)物養(yǎng)殖中化學(xué)益生素的使用提供基礎(chǔ)數(shù)據(jù)。

1 材料與方法

1.1 試驗(yàn)設(shè)計(jì)及動(dòng)物

試驗(yàn)選用同質(zhì)性良好的30只7日齡湖羊公雙羔（選自甘肅省金昌中天羊業(yè)有限公司）作為試驗(yàn)動(dòng)物，采用對(duì)照試驗(yàn)設(shè)計(jì)，將羔羊隨機(jī)分為2個(gè)處理組，每組15只，每只為1個(gè)重復(fù)。羔羊分別飼喂對(duì)照代乳粉（北京精準(zhǔn)動(dòng)物營養(yǎng)研究中心，營養(yǎng)物質(zhì)濃度見表1）或添加0.2 % MOS（SCIPHAR?，陜西森弗天然制品有限公司，純度＞90 %）的代乳粉。飼養(yǎng)試驗(yàn)持續(xù)21 d。

表1 羔羊代乳粉營養(yǎng)物質(zhì)濃度（風(fēng)干基礎(chǔ)）

1)營養(yǎng)水平均為實(shí)測值The nutrient levels are measured values

1.2 羔羊飼養(yǎng)管理

羔羊出生的1—3 d內(nèi)采食母乳，4日齡與母羊分離后奶瓶訓(xùn)飼代乳粉。7日齡清晨空腹稱重，按組間體重?zé)o差異（對(duì)照組4.09 ± 0.66 kg，MOS組4.07 ± 0.61 kg）的原則將羔羊隨機(jī)分為2組。8日齡時(shí)按試驗(yàn)設(shè)計(jì)飼喂羔羊，喂量為羔羊體重的2 %，每日飼喂4次，分別為6:00、12:00、18:00和24:00，代乳粉與水的比例為1:5。羔羊單籠飼養(yǎng)，自由飲水。

1.3 羔羊屠宰及胃腸道相關(guān)指標(biāo)測定

羔羊28日齡時(shí)屠宰，宰前不禁食禁水，稱量活重后立即頸靜脈放血致死。打開腹腔后按照馬仲華[27]的方法分離瘤胃、網(wǎng)胃、瓣胃、皺胃、十二指腸、空腸、回腸、盲腸、結(jié)腸和直腸。稱量胃腸道各部位凈質(zhì)量和含內(nèi)容物的質(zhì)量，計(jì)算各部位的相對(duì)質(zhì)量及其內(nèi)容物分布。測量腸道各段的長度，計(jì)算相對(duì)長度。

1.4 組織樣品采集及組織形態(tài)和小腸上皮細(xì)胞凋亡率測定

采集皺胃胃底腺區(qū)以及十二指腸、空腸和回腸中部約1 cm2的組織樣品，轉(zhuǎn)入多聚甲醛固定液中，在成都里來生物科技有限公司用H.E染色法和Tunel法觀察胃腸道組織形態(tài)和測定小腸上皮細(xì)胞凋亡率。

1.5 小腸黏膜緊密連接蛋白mRNA表達(dá)量測定

采集十二指腸、空腸和回腸中段黏膜樣品測定緊密連接蛋白1（claudin 1）、閉鎖小帶1（zonula occludens-1，ZO-1）和小腸黏膜閉鎖蛋白（occludin）的mRNA表達(dá)量。提取樣品總RNA，檢測RNA濃度和純度合格后將各樣品RNA反轉(zhuǎn)錄為cDNA。使用Oligo 7.0設(shè)計(jì)引物，其中claudin 1和ZO-1參考LIU等[28]方法設(shè)計(jì)，引物長度分別為216和163 bp，occludin參考GenBank設(shè)計(jì)（NC_040267），引物長度為93 bp，以β-Actin為內(nèi)參基因（NC_040362，長度97 bp）。RT-PCR使用20 μL擴(kuò)增體系：10 μL 2×Biogold qPCR SuperMix（2×Biogold qPCR Mixture，浙江博而金科技股份有限公司），0.4 μL上下游引物，1 μL cDNA，8.2 μL ddH2O。RT-PCR在Roche LightCycler?480II進(jìn)行，反應(yīng)條件為：95 ℃預(yù)變性3 min，95 ℃變性10 s，60 ℃退火20 s，72 ℃延伸10 s，40個(gè)循環(huán)，72 ℃延伸10 min。目的基因的相對(duì)表達(dá)量用2-ΔΔCt法計(jì)算。

1.6 數(shù)據(jù)統(tǒng)計(jì)分析

使用SPSS 22.0對(duì)試驗(yàn)數(shù)據(jù)進(jìn)行獨(dú)立樣本t檢驗(yàn)，以≤0.05表示為差異顯著，以0.05＜≤0.10表示差異具有顯著趨勢。

2 結(jié)果

2.1 MOS對(duì)羔羊胃腸道相對(duì)質(zhì)量的影響

2.1.1 羔羊胃腸指數(shù) 從表2可以看出，MOS對(duì)羔羊胃腸指數(shù)（%活體質(zhì)量）并未產(chǎn)生顯著影響（＞0.05），僅有降低羔羊十二指腸指數(shù)的趨勢（=0.066）。

2.1.2 羔羊胃腸相對(duì)質(zhì)量 從表3可以看出，MOS對(duì)羔羊胃腸相對(duì)質(zhì)量（%全胃質(zhì)量、%全腸質(zhì)量和%全胃腸質(zhì)量）均未產(chǎn)生顯著影響（＞0.05），但飼喂含MOS代乳粉羔羊的大腸相對(duì)質(zhì)量略高于對(duì)照組羔羊（＞0.05）。

2.1.3 羔羊腸道相對(duì)長度 從表4可以看出，MOS顯著降低了羔羊空腸相對(duì)長度（%全腸長度，=0.040），此外，MOS有增加盲腸相對(duì)長度的趨勢（=0.094）。

2.2 MOS對(duì)羔羊胃腸道內(nèi)容物分布的影響

2.2.1 羔羊胃腸內(nèi)容物相對(duì)活體質(zhì)量（%活體質(zhì)量） 從表5可以看出，MOS對(duì)羔羊胃腸內(nèi)容物相對(duì)活體質(zhì)量（%活體質(zhì)量）沒有產(chǎn)生顯著影響（＞0.05）。

2.2.2 羔羊胃腸內(nèi)容物相對(duì)總胃/腸內(nèi)容物及總胃腸內(nèi)容物相對(duì)質(zhì)量（%總胃內(nèi)容物、總腸內(nèi)容物、總胃腸內(nèi)容物） 從表6可以看出，MOS對(duì)羔羊胃腸內(nèi)容物相對(duì)總胃/腸內(nèi)容物及總胃腸內(nèi)容物相對(duì)質(zhì)量（%總胃內(nèi)容物、%總腸內(nèi)容物和%總胃腸內(nèi)容物）均沒有產(chǎn)生顯著影響（＞0.05），但采食含MOS代乳粉羔羊瘤胃內(nèi)容物相對(duì)質(zhì)量略高于對(duì)照組羔羊，而皺胃、盲腸和結(jié)腸內(nèi)容物相對(duì)質(zhì)量略低于對(duì)照組羔羊（＞0.05）。

表2 MOS對(duì)羔羊胃腸指數(shù)的影響

表3 MOS對(duì)羔羊胃腸相對(duì)質(zhì)量的影響

表4 MOS對(duì)羔羊腸道相對(duì)長度的影響

同行數(shù)據(jù)后所標(biāo)字母相異表示差異顯著（＜0.05），所標(biāo)字母相同表示差異不顯著（＞0.05）。下同

Different letters in the same row means significant difference between the treatments (＜0.05), same letter in the same row means not significant difference between treatments (＞0.05). The same as below

表5 MOS對(duì)羔羊胃腸內(nèi)容物相對(duì)活體質(zhì)量的影響

表6 MOS對(duì)羔羊胃腸內(nèi)容物相對(duì)總胃/腸內(nèi)容物及總胃腸內(nèi)容物相對(duì)質(zhì)量的影響

2.3 MOS對(duì)羔羊皺胃和小腸組織形態(tài)的影響

從表7可以看出，MOS顯著提高羔羊十二指腸絨毛高度和肌層厚度但顯著降低絨毛寬度（=0.033，=0.047，=0.015），此外，MOS有提高羔羊回腸絨毛高度、絨毛寬度和隱窩深度及皺胃肌層厚度的趨勢（=0.075，=0.078，=0.085，=0.084）。MOS對(duì)羔羊皺胃和小腸其他形態(tài)學(xué)指標(biāo)未產(chǎn)生顯著影響（＞0.05）。

從表8可以看出，MOS顯著上調(diào)了羔羊空腸ZO-1蛋白mRNA的表達(dá)量（=0.028），且有上調(diào)回腸occludin蛋白mRNA表達(dá)量的趨勢（=0.052）。MOS對(duì)羔羊十二指腸3種緊密連接蛋白、空腸claudin 1和occludin蛋白、回腸claudin 1和ZO-1蛋白mRNA的表達(dá)量以及各腸段上皮細(xì)胞凋亡率均沒有產(chǎn)生顯著影響（＞0.05）。

3 討論

3.1 MOS對(duì)羔羊胃腸道相對(duì)質(zhì)量的影響

幼齡反芻動(dòng)物胃腸道發(fā)育中，胃腸道的相對(duì)質(zhì)量（%活體質(zhì)量、%全胃質(zhì)量、%全腸質(zhì)量和%全胃腸道質(zhì)量）及相對(duì)長度（%全腸長度）等是反映機(jī)體消化道生長發(fā)育的重要指標(biāo)[29]。幼齡反芻動(dòng)物在生長發(fā)育中，體內(nèi)組織器官會(huì)因機(jī)體不同的功能需要而表現(xiàn)出不同的生長發(fā)育速度[29]。本次試驗(yàn)中，對(duì)照組和MOS處理組羔羊胃腸道相對(duì)質(zhì)量未出現(xiàn)顯著差異，這是因?yàn)?，整個(gè)試驗(yàn)期羔羊均飼喂液體代乳粉，盡管代乳粉營養(yǎng)均衡，但前胃得不到正常發(fā)育，單純吃奶的動(dòng)物，瘤胃缺乏粗糙物質(zhì)的刺激[30]，而固體飼料的攝入會(huì)為胃腸道發(fā)育帶來更好的物理刺激[31]?？苷加⒌萚32]也報(bào)道采食粗飼料可以刺激瘤胃發(fā)育，而幼齡反芻動(dòng)物僅喂乳汁或代乳品，會(huì)延滯前胃發(fā)育。但采食含MOS代乳粉的羔羊空腸相對(duì)長度顯著低于對(duì)照組羔羊，盲腸相對(duì)長度有高于對(duì)照組羔羊的趨勢且大腸各段的相對(duì)長度均高于對(duì)照組羔羊，主要是因?yàn)镸OS作為低聚糖，在羔羊小腸不能被消化吸收，而在大腸段可作為有益菌的發(fā)酵底物并促進(jìn)有害菌排出體外，促進(jìn)大腸發(fā)育[33]，使大腸相對(duì)長度增加而小腸相對(duì)長度有所降低，但由于其作為添加量很低的飼料添加劑，不能顯著改變羔羊胃腸道的生長發(fā)育，僅有微弱的作用。周懌[34]在犢牛代乳粉中添加75 mg·kg-1酵母β-葡聚糖后發(fā)現(xiàn)對(duì)胃腸道相對(duì)質(zhì)量無顯著影響，閆曉剛[35]在犢牛飼糧中添加20 g·d-1酵母培養(yǎng)物，對(duì)犢牛前胃相對(duì)質(zhì)量無顯著影響，也與本試驗(yàn)結(jié)果相同。

表7 MOS對(duì)羔羊皺胃和小腸組織形態(tài)的影響

表8 MOS對(duì)羔羊小腸黏膜緊密連接蛋白mRNA表達(dá)量及上皮細(xì)胞凋亡的影響

3.2 MOS對(duì)羔羊胃腸道內(nèi)容物分布的影響

胃腸道內(nèi)容物滯留時(shí)間決定養(yǎng)分的消化吸收率，而內(nèi)容物的滯留時(shí)間由食糜類型和胃腸道運(yùn)動(dòng)所決定。本次試驗(yàn)中，所有羔羊胃腸道內(nèi)容物含量均處于較低水平，主要是因?yàn)楦嵫蛑伙曃挂后w代乳粉，因此，會(huì)縮短食糜在胃腸道中的滯留時(shí)間，且食糜流通量顯著低于采食固體飼料的動(dòng)物[30]。胃腸道的節(jié)律性運(yùn)動(dòng)以及食糜的推送和分布，由食糜壓力差、體液因素、交感神經(jīng)、迷走神經(jīng)、平滑肌細(xì)胞興奮性、激素以及食糜的物理化學(xué)性質(zhì)等多種因素調(diào)控[36]。本次試驗(yàn)中，MOS對(duì)羔羊胃腸道內(nèi)容物分布并未產(chǎn)生顯著影響，也說明食物的物理形態(tài)是決定胃腸道內(nèi)容物分布的主要因素，MOS作為外源添加的益生素，不能對(duì)胃腸道運(yùn)動(dòng)和內(nèi)容物分布產(chǎn)生顯著影響，僅由于對(duì)胃腸道益生菌有促進(jìn)作用而改變食糜在胃腸道不同部位的滯留時(shí)間，導(dǎo)致采食含MOS代乳粉羔羊瘤胃內(nèi)容物相對(duì)質(zhì)量較高，而皺胃、盲腸和結(jié)腸內(nèi)容物相對(duì)質(zhì)量較低，也與前人在成年羊上的研究結(jié)果類似[33]。

3.3 MOS對(duì)羔羊皺胃和小腸組織形態(tài)的影響

對(duì)前胃功能發(fā)育不完善的幼齡反芻動(dòng)物來說，皺胃和小腸是養(yǎng)分最主要的消化吸收部位，而養(yǎng)分的吸收取決于皺胃和小腸的組織形態(tài)。皺胃的黏膜和肌層厚度、小腸的絨毛高度、隱窩深度、黏膜厚度及V/C值等，是評(píng)價(jià)動(dòng)物消化道對(duì)養(yǎng)分消化吸收的重要指標(biāo)[37-39]，如絨毛高度與腸道上皮細(xì)胞發(fā)育呈正相關(guān)，高度越高養(yǎng)分吸收能力越強(qiáng)[40]，而隱窩深度與腸道上皮細(xì)胞成熟率呈負(fù)相關(guān)，隱窩越淺表明細(xì)胞成熟率高且分泌功能越強(qiáng)[41]，此外，V/C值與腸道上皮細(xì)胞更新程度有關(guān)，也與腸道養(yǎng)分吸收能力呈正相關(guān)[42-43]。本次試驗(yàn)中，羔羊腸道絨毛均保持在較高水平，這是因?yàn)?，試?yàn)羔羊全期只飼喂液體代乳粉，而固體飼料會(huì)加大對(duì)腸絨毛的刺激而導(dǎo)致腸絨毛脫落速度加快[44]，液體飼料可維持腸絨毛高度[45]。本試驗(yàn)中，MOS顯著提高了羔羊十二指腸絨毛高度和肌層厚度，并有提高回腸絨毛高度、絨毛寬度和皺胃肌層厚度的趨勢，表明MOS能提高羔羊皺胃和小腸段的養(yǎng)分消化吸收能力。周懌[34]報(bào)道飼糧添加75 mg·kg-1酵母β-葡聚糖可提高犢牛小腸絨毛高度和V/C值。在仔豬和肉仔雞的試驗(yàn)中，飼糧添加0.1 % MOS可顯著提高小腸V/C值[21-22]。本試驗(yàn)中采食含MOS代乳粉羔羊空腸ZO-1蛋白mRNA表達(dá)量顯著上調(diào)，回腸occludin蛋白mRNA表達(dá)量也有上調(diào)的趨勢，且小腸上皮細(xì)胞凋亡率均低于對(duì)照組羔羊，也說明MOS有利于維持小腸正常屏障功能，并使腸絨毛維持在較高水平。occludin蛋白、claudin 1蛋白和ZO-1蛋白是腸上皮細(xì)胞間的緊密連接蛋白，構(gòu)建腸道屏障，機(jī)械性阻止微生物入侵[46]。而外界刺激、生理和病理等會(huì)使腸道屏障發(fā)生改變，增加腸上皮細(xì)胞間隙通透性，導(dǎo)致病原菌侵入細(xì)胞引發(fā)感染性疾病。Puthenedam等[47]報(bào)道，腸道中的乳酸菌和雙歧桿菌可上調(diào)ZO-1蛋白及occludin蛋白的表達(dá)，修復(fù)腸道損傷。楊俊等[48]也報(bào)道，腸上皮細(xì)胞被大腸桿菌（，EIEC）感染后，用乳酸菌處理時(shí)，緊密連接相關(guān)蛋白（claudin，occludin，junction adherensive molecular-1（JAM-1），ZO-1）表達(dá)量上調(diào)，腸道通透性得到改善。本試驗(yàn)中，MOS作為化學(xué)益生素可以促進(jìn)腸道中乳酸桿菌和雙歧桿菌等益生菌的增殖[16]，因此發(fā)揮出維護(hù)腸道屏障健康的作用。腸道黏膜的更新和上皮細(xì)胞的轉(zhuǎn)型由上皮細(xì)胞凋亡和有絲分裂共同維持，凋亡一方面可以促進(jìn)腸黏膜的適度發(fā)育和成熟，但另一方面，如細(xì)胞過度凋亡，將引起腸道功能紊亂[49]。尚沁沁[50]指出，益生菌可通過抑制病原菌在腸道的定植，調(diào)控細(xì)胞凋亡通路，降低腸上皮細(xì)胞凋亡率。Yan等[51]研究發(fā)現(xiàn)，乳酸菌可以抑制由腫瘤壞死因子α（tumor necrosis factor, TNF-α）誘導(dǎo)的腸道上皮細(xì)胞凋亡。本次試驗(yàn)中采食含MOS代乳粉羔羊小腸上皮細(xì)胞凋亡率低于對(duì)照組羔羊，也是由于MOS促進(jìn)腸道有益菌增殖而引起的[33]。

4 結(jié)論

7—28日齡湖羊羔羊代乳粉中添加MOS對(duì)羔羊胃腸道相對(duì)質(zhì)量和長度、內(nèi)容物分布和組織形態(tài)基本無顯著影響，但顯著提高羔羊十二指腸絨毛高度和肌層厚度并顯著降低絨毛寬度且上調(diào)空腸ZO-1蛋白mRNA表達(dá)量，還有提高羔羊回腸絨毛高度、絨毛寬度和隱窩深度、皺胃肌層厚度及回腸occludin蛋白mRNA表達(dá)量的趨勢。表明代乳粉中添加0.2 % MOS對(duì)湖羊羔羊胃腸道發(fā)育的影響較為微弱，主要對(duì)小腸絨毛形態(tài)和屏障功能有一定促進(jìn)作用。

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Effects of adding mannan oligosaccharides to milk replacer on the development of gastrointestinal tract of 7-28 days oldlambs

ZHENG Chen1, LI Fadi2,3, LI Fei2, ZHOU Juwang1, DUAN Pengwei1, LIU Huihui1, FAN Haimiao1, ZHU Weili1, LIU Ting1

(1College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070;2State Key Laboratory of Grassland Agro-ecosystems, Key laboratory of Grassland Livestock Industry Innovation/Ministry of Agriculture and Rural Affairs, College of Pastoral Agriculture Science and Technology, Lanzhou University, Lanzhou 730020;3Engineering Laboratory of Mutton Sheep Breeding and Reproduction Biotechnology in Gansu Province, Minqin 733300, Gansu)

【】This study was conducted to investigate the effects of mannan oligosaccharides (MOS) supplementation to milk replacer on the development of gastrointestinal tract of 7-28 day-oldlambs. 【】Thirty 7 day-oldmale lambs were chosen and divided into 2 groups randomly, fifteen lambs in each group and each lamb as a repeat. Lambs were fed milk replacer with or without 0.2 % MOS, respectively. The test lasted for 21 days. Eight lambs were selected from each group randomly and slaughtered at 28 day-old. The weights of the compound stomach and the intestinal tract with and without content, and lengths of the intestinal tract were measured, and the relative quality and length were calculated. While the tissue samples from fundus gland region of the abomasum, the middle part of duodenum, jejunum and ileum were fixed in paraformaldehyde to analyse the histomorphology, and the apoptotic rate of intestinal epithelial cells as well. And the mRNA expression of claudin 1, zonula occludens-1 (ZO-1), and occludin protein of duodenum, jejunum and ileum mucosa were measured.【】The results showed that except relative length of jejunum (=0.040), the relative weights (% body weight, % stomach weight, % intestinal tract weight, and % gastrointestinal tract weight), relative lengths (% intestinal tract length), content of stomach and intestinal tract (% body weight, % stomach content weight, % intestinal tract content weight, and % gastrointestinal tract content weight), the apoptotic rate of intestinal epithelial cells and mRNA expression of claudin 1 protein in intestinal tract of lambs were not affected by MOS (＞0.05). However, MOS elongated the villus height and the muscular thickness, and decreased the villus width of lamb duodenum significantly (=0.033=0.047=0.015). MOS also up-regulated the mRNA expression of ZO-1 protein of lamb jejunum significantly (=0.028). And there was a tendency that MOS elongated villus height, width and crypt depth of ileum, muscular thickness of abomasum, and mRNA expression of occludin protein of ileum (=0.075=0.078=0.085=0.084=0.052).【】MOS almost did not affect the relative weights, lengths, and content distribution of gastrointestinal tract of 7-28 days oldlambs, but improved the histomorphology of duodenum and ileum, indicating it could maintain barrier function of intestinal tract and benefit to nutrients digestibility.

lamb; mannan oligosaccharides; milk replacer; gastrointestinal tract; development

10.3864/j.issn.0578-1752.2020.02.014

2019-06-24；

2019-08-14

國家自然科學(xué)基金(31560646，31860657)

鄭琛，E-mail：zhengc@gsau.edu.cn。通信作者劉婷，E-mail：liuting@gsau.edu.cn

（責(zé)任編輯 林鑒非）