復(fù)方天然植物制劑對(duì)奶?；厝樾Ч罢{(diào)控激素的影響

沈留紅，肖勁邦，巫曉峰，姜思汛，江濤，鄧俊良，左之才，余樹民，曹隨忠

（四川農(nóng)業(yè)大學(xué)動(dòng)物醫(yī)學(xué)院，動(dòng)物疫病與人類健康四川省重點(diǎn)實(shí)驗(yàn)室/奶牛疾病研究中心，成都 611130）

復(fù)方天然植物制劑對(duì)奶?；厝樾Ч罢{(diào)控激素的影響

沈留紅，肖勁邦，巫曉峰，姜思汛，江濤，鄧俊良，左之才，余樹民，曹隨忠

（四川農(nóng)業(yè)大學(xué)動(dòng)物醫(yī)學(xué)院，動(dòng)物疫病與人類健康四川省重點(diǎn)實(shí)驗(yàn)室/奶牛疾病研究中心，成都 611130）

【目的】探究復(fù)方天然植物制劑對(duì)奶?；厝樾Ч盎厝槠谀膛Ｑ逡葝u素（INS）、氫化可的松（HC）、生長(zhǎng)激素（GH）和胰島素生長(zhǎng)因子1（IGF-1）含量的影響?！痉椒ā吭囼?yàn)于2016年1—6月進(jìn)行，選擇四川省某規(guī)?；膛?chǎng)半封閉統(tǒng)一舍飼，體重（582±41）kg、第2—4胎中國(guó)荷斯坦奶牛150頭。從中選取體況良好，健康，乳房、乳汁均正常，即將進(jìn)入回乳期，產(chǎn)奶量為（15.42±0.71）kg的妊娠后期即將干乳的健康奶牛80頭。隨機(jī)分為A、B、C、D組，每組20頭，均采用逐漸干奶法回乳，回乳開始當(dāng)天記為第0天，A組為對(duì)照組，不飼喂復(fù)方天然植物制劑。B、C、D組均從回乳開始第1天，每日上午8∶00分別飼喂復(fù)方天然植物制劑400、500、600g，回乳完成后停止飼喂復(fù)方天然植物制劑。復(fù)方天然植物制劑由麥芽，樸硝，升麻，柴胡，香附，薏仁，蠶蛻，白術(shù)，黃芩，知母，蘇梗，芡實(shí)，五味子，蒲公英，甘草等按一定比例組成。分別采集奶牛回乳期第0、1、3、5、7、9和11天尾靜脈血，置于未加抗凝劑的離心管中，室溫下靜置1h，置離心機(jī)352×g離心10min，轉(zhuǎn)移上層血清于EP管中，-20℃凍存。采用雙抗體夾心酶聯(lián)免疫吸附技術(shù)（ELISA）法檢測(cè)血清INS、HC、GH和IGF-1含量，并記錄回乳期奶牛單日產(chǎn)奶量?！窘Y(jié)果】A、B、C、D 4組奶牛回乳時(shí)間分別為 11、7、5、5天。各組回乳期奶牛血清 IGF-1和 HC含量均呈下降趨勢(shì)，且對(duì)照組下降趨勢(shì)較各復(fù)方天然植物制劑組慢，復(fù)方天然植物制劑組血清IGF-1、HC和GH含量在回乳期后回升，對(duì)照組血清INS含量上升趨勢(shì)較各復(fù)方天然植物制劑組慢，第0至1天，各組奶牛血清HC、IGF-1和GH含量差異均不顯著（P＞0.05）；第3—7天，A組奶牛血清HC、GH和IGF-1含量極顯著高于B、C、D組（P＜0.01），而血清INS含量極顯著低于B、C、D組（P＜0.01）；第9天，A、D組奶牛血清HC含量均顯著高于C組（P＜0.05），A、B組奶牛血清GH和IGF-1含量均極顯著低于C、D組（P＜0.01），A組奶牛血清INS含量極顯著低于B、C、D組（P＜0.01）；第11天，A組奶牛血清HC、GH和IGF-1含量極顯著低于B、C、D組（P＜0.01），A組奶牛血清INS顯著低于D組（P＜0.05），但極顯著低于B、C組（P＜0.01）； C、D組奶牛血清IGF-1、GH、HC和INS含量在整個(gè)回乳期均差異性不顯著（P＞0.05）。在奶?；厝槠?，產(chǎn)奶量、HC、GH和IGF-1兩兩間均呈極顯著正相關(guān)（P＜0.01），INS與產(chǎn)奶量、HC、GH和IGF-1均呈極顯著負(fù)相關(guān)（P＜0.01）?！窘Y(jié)論】復(fù)方天然植物制劑能顯著促進(jìn)回乳期奶?；厝?，飼喂500g/日復(fù)方天然植物制劑，奶?；厝闀r(shí)間為5d，回乳效果即可達(dá)到最佳。回乳期奶牛血清INS、HC、IGF-1含量和產(chǎn)奶量變化兩兩間均呈極顯著相關(guān)，IGF-1、GH和HC對(duì)奶牛回乳起負(fù)向調(diào)控功能，而INS對(duì)奶牛回乳起正調(diào)控作用。

奶牛；回乳；復(fù)方天然植物制劑；泌乳激素

Abstract:【Objective】the objective of this study is to explore the impact of compound natural plant preparation on the milk withdrawal and serum insulin(INS), hydrocortisone(HC), growth hormone(GH) and insulin growth factor 1(TGF-1)levels in dairy cows.【Method】A total of 150 Chinese Holstein cows, weighing (582±41) kg, pregnant with 2-4 fetuses, and fed in a large-scale dairy farm in Sichuan Province from January 2016 to June 2016, were used in the present study. Eighty healthy cows with normal milk and udder that in late pregnancy and gave (15.42±0.71) kg milk per day were chosen. The cows were randomly divided into four groups: groups A, B, C and D, with 20 of them in each group. Stop dairy cows’ milk by gradually dry milk method and the day when cows gave less milk was recorded as 0 d. Group A was a control group, did not feed compound natural plant preparation and the groups B, C and D were fed 400g, 500g, 600g compound natural plant preparation,respectively, at 8.00 am from 1 d to the day of dry milk completely. Compound natural plant preparation comprises fructus hordei germinatus, mirabilite, rhizoma cimicifugae, bupleurum, rhizoma cyperi, coix seed, silkworm slough, rhizome atratylodis, scutellaria baicalensis, rhizome anemarrhenae, caulis perllae, semen euryales, schisandra chinensis, dandelion,licorice and so on. The cows’ venous blood was collected from tail on the day 0, 1, 3, 5, 7, 9 and 11, the samples were placed in the centrifuge tubes without anticoagulant at room temperature for 1h. The suspension was centrifuged at 352 × g for 10 min and then put into the refrigerator at -20℃. ELISA was used to evaluate the expression levels of serum INS, HC and IGF-I and the daily milk production was recorded.【Result】Groups A, B, C and D returned milk on the 11 d, 7 d, 5 d and 5 d respectively.The expression levels of serum IGF-1 and HC were declining in all 4 groups and the decline trend of the control group was slower than compound natural plant preparation groups. The expression levels of serum IGF-1, HC and GH were raised after the period of milk withdrawal, the upward trend of the expression levels of serum INS in the control group was slower than compound natural plant preparation groups. From the day 0 to the 1, the expression levels of serum HC, IGF-1 and GH had no significant difference(P＞0.05). From the day 3 to the day7, the expression levels of serum HC, GH and IGF-1 in group A were highly and significantly higher than those of groups B, C and D (P＜0.01), while the expression level of INS in group A was highly significant loser than those of groups B, C and D(P＜0.01). On the day 9, the expression levels of serum HC in groups A and D were significantly higher than that of group C(P＜0.05), the expression levels of serum GH and IGF-1 in groups A and B were highly and significantly lower than those of groups C and D(P＜0.01), the expression levels of serum INS in group A was highly and significantly lower than that of groups B, C and D(P＜0.01). On the day 11, the expression levels of serum HC, GH and IGF-1 in group A were highly significantly lower than those of groups B, C and D(P＜0.01), the expression levels of serum INS in group A was highly significant lower than that of groups B and C(P＜0.01) and significantly lower than that of group D(P＜0.05). The expression levels of serum IGF-1, GH, HC and INS had no significant difference between group C and D during the period of milk withdrawal (P＞0.05). During the period of milk withdrawal, there were significant positive correlations among milk production, HC, GH and IGF-1(P＜0.01) and there was a significant negative correlation between INS and milk production, HC, GH, IGF-1(P＜0.01).【Conclusion】 Compound natural plant preparation can help cows milk withdrawal significantly, feeding 500g·d-1can shorten the period of milk withdrawal to 5 days can make the best effect on milk withdrawal. The expression levels of serum INS, HC, IGF-1 and milk production had a highly significant correlation. IGF-1 GH and HC are negative regulation factors of milk withdrawal, while INS is a positive regulation factor.

Key words:dairy cow; milk withdrawal; compound natural plant preparation; galactin

0 引言

【研究意義】奶牛經(jīng)過(guò)漫長(zhǎng)的泌乳期，機(jī)體損耗大量營(yíng)養(yǎng)物質(zhì)，需經(jīng)回乳期，進(jìn)入干奶期，蓄積機(jī)體儲(chǔ)備，更新乳腺組織及優(yōu)化內(nèi)分泌系統(tǒng)。為分娩后高效泌乳做物質(zhì)準(zhǔn)備?；厝槠谀膛５幕厝樾Ч麑⒅苯佑绊懏a(chǎn)后泌乳量及乳汁品質(zhì)。當(dāng)下次泌乳期再次來(lái)臨，乳腺細(xì)胞必須具有較強(qiáng)活力才能夠支撐大量泌乳而引起的營(yíng)養(yǎng)損耗[1]。若乳腺上皮細(xì)胞持續(xù)分泌乳汁，乳腺細(xì)胞無(wú)法休整，不僅影響下一個(gè)泌乳期的泌乳量，且對(duì)之后胎次的泌乳均有不利影響[2]。因此，縮短回乳期，使奶牛快速進(jìn)入干乳期，對(duì)奶牛乳腺的保護(hù)具有重要意義。目前還未見專一縮短奶?；厝槠诘乃幬?，復(fù)方天然植物制劑因具有多種功效，無(wú)抗藥性和耐藥性，無(wú)藥物殘留和不良反應(yīng)少等獨(dú)特優(yōu)勢(shì)，自古以來(lái)一直被廣泛應(yīng)用于提高免疫、促進(jìn)生長(zhǎng)、催乳和回乳等[3]。探究復(fù)方天然植物制劑對(duì)回乳期奶?；厝樾Ч把迕谌榧に?、因子的影響，有助于奶牛乳房保健產(chǎn)品的開發(fā)，改進(jìn)奶?；厝榉椒?，降低回乳期奶牛乳房炎發(fā)生率?！厩叭搜芯窟M(jìn)展】影響奶牛乳腺合成、乳汁分泌的因素很多，如飼養(yǎng)管理、遺傳育種、營(yíng)養(yǎng)代謝、內(nèi)分泌因素等[4]。但在相同的飼養(yǎng)管理模式下，奶牛乳腺合成、乳汁分泌主要受到泌乳激素和因子的調(diào)控，其中直接調(diào)控乳腺發(fā)育和泌乳的激素主要有胰島素（insulin, INS）、氫化可的松（hydrocortisone,HC）、生長(zhǎng)激素（growth hormone，GH）、催乳素（prolactin,PRL）、雌激素（estrogen, E）和孕酮（progesterone，P4）等[5-6]，其通過(guò)與胰島素樣生長(zhǎng)因子1（Insulin-like growth factor 1，IGF-1）、轉(zhuǎn)化生長(zhǎng)因子 β1（transforming growth factor β1，TGF-β1）和信號(hào)轉(zhuǎn)導(dǎo)和轉(zhuǎn)錄激活因子5（signal transduction and transcriptional activation factor of 5，STAT5）等[7]細(xì)胞因子相互作用而形成乳腺發(fā)育和泌乳的調(diào)控網(wǎng)絡(luò)。運(yùn)用復(fù)方天然植物制劑可提高人、小鼠、大鼠、兔、肉牛等動(dòng)物機(jī)體INS、HC、GH、IGF-1含量[3,8-13]以及促進(jìn)人和小鼠等動(dòng)物回乳或泌乳已有研究[14-16]，并顯示均與泌乳激素及因子呈一定的相關(guān)性[17-18]。【本研究切入點(diǎn)】奶牛經(jīng)過(guò)泌乳期，機(jī)體消耗嚴(yán)重，需要干奶期恢復(fù)體質(zhì)，更新乳腺組織，為下一個(gè)泌乳期做充分準(zhǔn)備，而傳統(tǒng)回乳方法效果差，容易造成乳房炎、流產(chǎn)、抗生素殘留等副作用[19]，回乳復(fù)方天然植物制劑在奶?；厝槠诘倪\(yùn)用有巨大研究前景[20]。因此本文旨在研究復(fù)方天然植物制劑對(duì)回乳期奶?；厝樾Ч把迕谌榧に亍⒁蜃拥挠绊?。為進(jìn)一步研究復(fù)方天然植物制劑對(duì)回乳期奶牛泌乳相關(guān)激素的調(diào)控機(jī)理，以及為尋求更安全有效的回乳技術(shù)措施提供理論依據(jù)?！緮M解決的關(guān)鍵問(wèn)題】通過(guò)對(duì)回乳期奶牛飼喂復(fù)方天然植物制劑，探究血清INS、HC、GH和IGF-1含量變化，以期為復(fù)方天然植物制劑在回乳期奶牛的應(yīng)用提供理論依據(jù)，確保復(fù)方天然植物制劑在奶?；厝槠诘膽?yīng)用正確合理。

1 材料與方法

1.1 材料

1.1.1 試驗(yàn)動(dòng)物 2016年1—6月，選擇四川省某規(guī)?；膛?chǎng)半封閉統(tǒng)一舍飼，體重（582±41）kg、2—4胎中國(guó)荷斯坦奶牛150頭。從中選取體況良好，健康，乳房、乳汁均正常，即將進(jìn)入回乳期，產(chǎn)奶量為（15.42±0.71）kg的妊娠后期奶牛80頭。隨機(jī)分為A、B、C、D 4組，每組20頭。各組均采用逐漸干奶法進(jìn)行回乳，回乳當(dāng)天記為第0天，從回乳第1天，A組為對(duì)照組，不飼喂復(fù)方天然植物制劑，B、C、D組每日上午 8:00分別飼喂復(fù)方天然植物制劑 400、500、600g，直至停奶，停止飼喂。

1.1.2 試驗(yàn)藥物 復(fù)方天然植物制劑由四川農(nóng)業(yè)大學(xué)動(dòng)物醫(yī)學(xué)院奶牛疾病研究中心研發(fā)。由麥芽，樸硝，升麻，柴胡，香附，薏仁，蠶蛻，白術(shù)，黃芩，知母，蘇梗，芡實(shí)，五味子，蒲公英和甘草等組成。均購(gòu)自成都荷花池中藥材專業(yè)市場(chǎng)（一等品）。按比例配方，粉碎過(guò)60目篩，塑料袋500 g/袋分裝備用。

1.1.3 試驗(yàn)試劑 牛胰島素、氫化可的松、生長(zhǎng)激素和胰島素樣生長(zhǎng)因子1雙抗體夾心酶聯(lián)免疫吸附檢測(cè)（ELISA）試劑盒，均由美國(guó)RD公司提供。

1.2 試驗(yàn)方法

1.2.1 回乳方式 采用逐漸干奶法[10-11]，方法為：停喂多汁飼料，減少精料喂量，以青干草為主，控制飲水，適當(dāng)加強(qiáng)運(yùn)動(dòng)。在回乳第1天，擠奶次數(shù)由3次改為2次，第2天改為1次，逐漸減少擠奶次數(shù)，當(dāng)日產(chǎn)奶量為3—4 kg時(shí)，停止擠奶。

1.2.2 血清收集 試驗(yàn)?zāi)膛；厝殚_始當(dāng)天記為第 0天，依次采集第0、1、3、5、7、9和11天尾靜脈血10mL，置于未加抗凝劑的離心管中，室溫下靜置1h，離心352×g離心10min，轉(zhuǎn)移上層血清于EP管中，-20℃凍存，待檢。

1.2.3 ELISA檢測(cè) 采用雙抗體夾心酶聯(lián)免疫吸附技術(shù)（ELISA）測(cè)定牛胰島素、氫化可的松、生長(zhǎng)激素和胰島素樣生長(zhǎng)因子1的水平，步驟嚴(yán)格按照說(shuō)明書進(jìn)行。血清樣品的檢測(cè)于2016 年7—8月在四川農(nóng)業(yè)大學(xué)動(dòng)物醫(yī)學(xué)院動(dòng)物疫病與人類健康四川省重點(diǎn)實(shí)驗(yàn)室和奶牛疾病研究中心進(jìn)行。

1.3 統(tǒng)計(jì)分析

SPSSl9.0軟件進(jìn)行統(tǒng)計(jì)學(xué)分析，K-S檢驗(yàn)計(jì)量資料是否服從正態(tài)分布，以平均數(shù)±標(biāo)準(zhǔn)差（ˉX±S）表示，兩組間采用獨(dú)立樣本t檢驗(yàn)，多組間比較采用單因素方差分析，相關(guān)性分析采用雙變量 Pearson相關(guān)分析，P＜0.05為差異有統(tǒng)計(jì)學(xué)意義，P＜0.01為差異性極顯著。

2 結(jié)果

2.1 復(fù)方天然植物制劑對(duì)回乳期奶牛產(chǎn)奶量的影響

如表1所示，A、B、C、D 4組奶?；厝槠谛枰獣r(shí)間分別是11、7、5和5d，回乳第0天，各組間奶產(chǎn)量差異不顯著（P＞0.05）；第1天，B組產(chǎn)奶量極顯著高于D組（P＜0.01）；第3—5天，A、B組產(chǎn)奶量均極顯著高于C、D組（P＜0.01），且A組產(chǎn)奶量極顯著高于B組（P＜0.01）；第7天，A組產(chǎn)奶量極顯著高于B組；C、D組產(chǎn)奶量在整個(gè)回乳期差異均不顯著（P＞0.05）。

表1 復(fù)方天然植物制劑對(duì)回乳期奶牛奶產(chǎn)量的影響（kg·d-1,ˉX±S）Table 1 The effects of compound natural plant preparation on the milk production of dairy cows during the period of milk withdrawal

2.2 復(fù)方天然植物制劑對(duì)回乳期奶牛血清 INS含量的影響

如表2所示，在奶?；厝槠?，各組INS含量均隨時(shí)間推移呈上升趨勢(shì)，回乳第0—1天，各組INS含量差異均不顯著（P＞0.05）；第3—5天，A組INS含量極顯著低于B、C、D組（P＜0.01），B組顯著低于C組（P＜0.05）；第7天，A、B組INS含量極顯著低于C、D組（P＜0.01），A組極顯著低于B組（P＜0.01）；第9天，A組INS含量極顯著低于B、C、D 組（P＜0.01），B 組顯著低于 C、D 組（P＜0.05）；第 11天，A組極顯著低于 B、C組（P＜0.01），A組顯著低于D組（P＜0.05）；C、D組INS含量在整個(gè)回乳期均差異性不顯著（P＞0.05）。

表2 復(fù)方天然植物制劑對(duì)回乳期奶牛血清INS含量的影響（nIU·mL-1,ˉX±S）Table 2 The effects of compound natural plant preparation on the insulin of dairy cows during period of milk withdrawal

2.3 復(fù)方天然植物制劑對(duì)回乳期奶牛血清HC含量的影響

如表3所示，在奶?；厝槠?，各組HC含量均隨時(shí)間推移呈下降趨勢(shì)，回乳第0—1天，各組HC含量差異均不顯著（P＞0.05）；第3—7天，A組HC含量極顯著高于B、C、D組（P＜0.01）；第9天，A、D組HC含量均顯著高于C組（P＜0.05）；第11天，A組HC含量極顯著低于B、C、D組（P＜0.01），且B組顯著低于C、D組（P＜0.05）；C、D組HC含量在整個(gè)回乳期差異均不顯著（P＞0.05）。

2.4 復(fù)方天然植物制劑對(duì)回乳期奶牛血清GH含量的影響

如表4所示，在奶牛回乳期，各組奶牛血清GH含量均呈下降趨勢(shì)，并在回乳期后上升。第0—1 天，GH含量在四組間差異均不顯著（P＞0.05）；第3—7天，A組GH含量均極顯著高于B、C、D組（P＜0.01）；第9天，A、B組GH含量均極顯著低于C、D組（P＜0.01）；第11天，A組GH含量極顯著低于B、C、D組（P＜0.01），B組GH含量顯著低于C組（P＜0.05），B組GH含量極顯著低于D組（P＜0.01）；C、D組GH含量在整個(gè)回乳期差異均不顯著（P＞0.05）。

表3 復(fù)方天然植物制劑對(duì)回乳期奶牛血清HC含量的影響（ng·mL-1,ˉX±S）Table 3 The effects of compound natural plant preparation on the HC of dairy cows during the period of milk withdrawal

表4 復(fù)方天然植物制劑對(duì)回乳期奶牛血清GH含量的影響（ng·mL-1,ˉX±S）Table 4 The effects of compound natural plant preparation on GH of dairy cows during the period of milk withdrawal

2.5 復(fù)方天然植物制劑對(duì)回乳期奶牛血清 IGF-1含量的影響

如表5所示，在奶牛回乳期，各組IGF-1含量均隨時(shí)間推移呈下降趨勢(shì)，并在回乳期后回升，回乳第0天，B組IGF-1含量顯著高于C組（P＜0.05）；第1天，各組IGF-1含量差異均不顯著（P＞0.05）；第37天，A組IGF-1含量極顯著高于B、C、D組（P＜0.01）；第9天，A、B組IGF-1含量均極顯著低于C、D組（P＜0.01），A組極顯著高于B組（P＜0.01）；第11天，A組IGF-1含量極顯著低于B、C、D組（P＜0.01）；C、D組IGF-1含量在整個(gè)回乳期均差異性不顯著（P＞0.05）。

表5 復(fù)方天然植物制劑對(duì)回乳期奶牛血清IGF-1含量的影響（ng·mL-1,ˉX±S）Table 5 The effects of compound natural plant preparation on IGF-1 of dairy cows during the period of milk withdrawal

2.6 回乳期奶牛血清INS、HC、IGF-1含量和產(chǎn)奶量間的相關(guān)性

利用對(duì)照組進(jìn)行相關(guān)性分析，結(jié)果如表6所示，在奶牛回乳期，產(chǎn)奶量、HC、GH和IGF-1兩兩間均呈極顯著正相關(guān)（P＜0.01），INS與產(chǎn)奶量、HC、GH和IGF-1均呈極顯著負(fù)相關(guān)（P＜0.01）。

表6 回乳期奶牛血清INS、HC、IGF-1含量和產(chǎn)奶量的相關(guān)性Table 6 Correlation between the expression levels of INS, HC and IGF-1 milk yield during the period of withdrawal

3 討論

3.1 對(duì)回乳期奶牛產(chǎn)奶量的影響

目前，人醫(yī)運(yùn)用麥芽口服，芒硝外敷等回乳方式已不少見[14,16]。王雄[14]使用“回乳抑增一號(hào)”（麥芽、牡蠣、浙川貝等）使婦女溢乳改善，郝振華等[15]利用麥芽治療婦女產(chǎn)后溢乳有明顯療效?，F(xiàn)有回乳復(fù)方天然植物制劑中多有麥芽，且已經(jīng)人工合成麥角衍生物——卡麥角林能明顯抑制PRL生成，進(jìn)而控制泌乳[22]。然而，復(fù)方天然植物制劑運(yùn)用于奶牛回乳還鮮有報(bào)道，本研究結(jié)果顯示，在奶?；厝檫^(guò)程中，各組奶牛產(chǎn)奶量均隨時(shí)間推移呈降低趨勢(shì)，回乳第 1天，400g組產(chǎn)奶量極顯著高于 500g組（P＜0.01）；第3—5天，對(duì)照組和400g組產(chǎn)奶量極顯著高于500、600g組（P＜0.01），且對(duì)照組產(chǎn)奶量極顯著高于 400g組（P＜0.01），表明了復(fù)方天然植物制劑具有較好回乳效果，縮短了奶?；厝槠冢矣行p少了日產(chǎn)奶量，推測(cè)其機(jī)理可能是復(fù)方天然植物制劑以炒麥芽消食、回乳[14,16]；升麻、柴胡、香附、白術(shù)疏肝理氣；黃芩、知母、蘇梗安胎；五味子、蒲公英固精斂陰，清熱解毒，統(tǒng)籌全局，斷其生化之源，使肝氣調(diào)達(dá)四運(yùn)，有效抑制乳汁分泌而回乳，500、600g組產(chǎn)奶量在整個(gè)回乳期差異均不顯著（P＞0.05），表明飼喂復(fù)方天然植物制劑達(dá)到500g/日時(shí)，回乳效果即可達(dá)到最佳。

3.2 對(duì)回乳期奶牛血清INS含量的影響

INS具有加強(qiáng)糖原合成、維持血糖恒定并調(diào)控泌乳等作用[23]。其可激活胰島素亞基受體I（IRS-I），通過(guò)配體約束力和自動(dòng)磷酸化的誘導(dǎo)，促使生成INS受體蛋白位點(diǎn)，從而調(diào)控乳腺上皮細(xì)胞乳糖、乳脂的生物合成[24]。但I(xiàn)NS對(duì)反芻動(dòng)物乳腺發(fā)育的作用仍存在爭(zhēng)議，佟慧麗等[25]利用INS處理奶山羊乳腺上皮細(xì)胞后發(fā)現(xiàn)，細(xì)胞活力無(wú)明顯變化，而陳建暉等[26]對(duì)奶牛乳腺上皮細(xì)胞進(jìn)行類似處理后細(xì)胞活力下降，田青等[27]則表示INS對(duì)奶牛泌乳細(xì)胞生長(zhǎng)及分化均有促進(jìn)作用。本研究結(jié)果顯示，在奶牛回乳過(guò)程中，血清INS含量在第0—1天變化不顯著（P＞0.05），在第3—11天依次升高且差異均極顯著（P＜0.01），可能由于回乳期間GH含量降低，其對(duì)INS的抑制作用減少，并且由于懷孕后期，胎犢宮內(nèi)發(fā)育需要大量糖脂沉積，共同促進(jìn)機(jī)體釋放INS。

PANG等[28]利用黃連治療糖尿病患者取得良好效果，王璟[8]發(fā)現(xiàn)生地、黃芪、當(dāng)歸等組成的復(fù)方天然植物制劑提高了糖尿病患者INS含量，張秋華等[9]指出，黃芩、知母、白術(shù)等復(fù)方天然植物制劑可提高小鼠體內(nèi)INS表達(dá)量，另有研究表明，黃連、黃芩、知母等復(fù)方天然植物制劑能夠顯著提高INS對(duì)糖代謝作用率[29]，本研究顯示，在奶牛回乳過(guò)程中，復(fù)方天然植物制劑能夠極顯著提高回乳期奶牛血清 INS含量（P＜0.01），400g組INS含量升高趨勢(shì)較500、600g組慢，但500、600g組INS含量在整個(gè)回乳期差異均不顯著（P＞0.05），表明飼喂500和600g復(fù)方天然植物制劑對(duì)奶牛回乳期血清INS含量的影響不顯著?？赡苁菑?fù)方天然植物制劑中黃芩和知母等可提升奶牛血清INS含量，促進(jìn)奶牛機(jī)體糖脂沉積，為滿足奶牛自身及胎兒生長(zhǎng)發(fā)育的營(yíng)養(yǎng)和能量需要，從而減少了產(chǎn)奶量。

3.3 對(duì)回乳期奶牛血清HC含量的影響

HC是哺乳動(dòng)物腎上腺皮質(zhì)分泌的主要糖皮質(zhì)激素之一，近年來(lái)，關(guān)于HC對(duì)提高奶牛泌乳量及改善乳品質(zhì)逐漸成為研究熱點(diǎn)。HC與乳腺細(xì)胞胞內(nèi)核受體結(jié)合，啟動(dòng)并調(diào)控泌乳相關(guān)基因表達(dá)和乳蛋白合成，通過(guò)基因組機(jī)制實(shí)現(xiàn)其生理和藥理功能[30-31]。KABOTANSKI等[32]研究發(fā)現(xiàn)HC能夠增強(qiáng)PRL對(duì)酪蛋白mRNAs累積，進(jìn)而對(duì)奶牛乳腺上皮細(xì)胞增殖起重要調(diào)控。本研究結(jié)果顯示，在奶?；厝檫^(guò)程中，血清HC含量在第0—1天變化不顯著（P＞0.05），第3天極顯著高于其余各天（P＜0.01），可能是由于奶牛飼糧成分改變和飼舍轉(zhuǎn)換等造成的應(yīng)激使HC含量升高，而第 3—11天依次降低且差異均極顯著（P＜0.01），提示奶?；厝槠谘錚RL降低，促使乳腺減少其泌乳細(xì)胞胞內(nèi)核受體表達(dá)量，反饋抑制腎上腺皮質(zhì)分泌 HC[33]，降低泌乳量，也從正面表明了 HC在奶?；厝橹械闹匾饔?。

王喜軍等[10]利用六味地黃丸降低了HC致大鼠腎虛動(dòng)物模型HC表達(dá)量，蘇成虎等[11]利用水芹提取物減少了HC致腎陽(yáng)虛小鼠體內(nèi)HC含量。本研究發(fā)現(xiàn)，在奶牛回乳過(guò)程中，飼喂復(fù)方天然植物制劑各組均極顯著降低血清HC含量（P＜0.01），說(shuō)明復(fù)方天然植物制劑降HC含量效果明顯，在回乳期后，中藥組血清 HC含量明顯回升，且各中藥組間差異不顯著（P＞0.05），可能是由于奶牛及胎犢對(duì)糖、脂肪和蛋白質(zhì)等代謝調(diào)節(jié)需求 HC[34]，而對(duì)照組繼續(xù)下降，可能是由于對(duì)照組回乳持續(xù)時(shí)間較長(zhǎng)，致使HC還未達(dá)到回升期，綜上所述，奶牛在回乳期HC明顯降低，并在回乳期后回升，表明各組復(fù)方天然植物制劑降奶牛血清HC含量均有良好效果。

3.4 對(duì)回乳期奶牛血清GH含量的影響

GH是由腦垂體分泌的一種單一肽鏈的蛋白質(zhì)激素，有促進(jìn)生長(zhǎng)發(fā)育，調(diào)控泌乳等作用[35-36]。運(yùn)用牛生長(zhǎng)素可調(diào)節(jié)奶牛機(jī)體物質(zhì)代謝，提高飼料轉(zhuǎn)化率，增加產(chǎn)奶量[37-38]， 對(duì)早期泌乳奶牛注射GH可增加產(chǎn)奶量[17]，其主要通過(guò)JAK2/STAT5信號(hào)通路、IGF-I和GH受體等增加乳腺血流量，使?fàn)I養(yǎng)成分向乳腺聚集，促進(jìn)乳腺發(fā)育[39]。本試驗(yàn)結(jié)果表明，在奶?；厝檫^(guò)程中，GH含量呈下降趨勢(shì)，并在回乳期后回升，可能是減少了擠奶刺激，乳房GH受體表達(dá)受到抑制，進(jìn)而反饋抑制垂體分泌 GH，也可能是由于懷孕末期，為滿足胎犢宮內(nèi)快速生長(zhǎng)發(fā)育需求導(dǎo)致。

李嬪等[3]利用生地、黃柏和知母等制成濃縮合劑飼喂大鼠，能夠調(diào)控GH蛋白表達(dá)。LO等[31]對(duì)大鼠灌服大黃衍生物使GH表達(dá)增加。王金合等[12]對(duì)肉牛拌喂柴胡、當(dāng)歸等提高肉牛血清GH含量。本研究表明，在奶?；厝檫^(guò)程中，對(duì)照組血清GH含量在回乳第1—11天均呈下降趨勢(shì)，且在第3—7天，對(duì)照組血清 GH含量極顯著高于中藥組（P＜0.01），400g組在第3—5天極顯著高于500、600g組（P＜0.01），隨后，復(fù)方天然植物制劑組GH含量均上升，且500與600g組在整個(gè)回乳期差異性不顯著（P＞0.05）。說(shuō)明復(fù)方天然植物制劑對(duì)回乳期奶牛降 GH效果明顯，飼喂500g/日即可達(dá)到最佳效果，且在回乳期后，復(fù)方天然植物制劑組GH含量回升，推測(cè)復(fù)方天然植物制劑對(duì)胎兒生長(zhǎng)發(fā)育有促進(jìn)作用，而對(duì)照組GH在回乳期呈現(xiàn)逐漸下降趨勢(shì)，可能是由于對(duì)照組奶牛回乳速度較慢，GH還未達(dá)到回升期。表明復(fù)方天然植物制劑能夠使奶?？焖倩厝?，使血清GH含量快速回升，進(jìn)而間接促進(jìn)胎犢生長(zhǎng)發(fā)育。

3.5 對(duì)回乳期奶牛血清IGF-1含量的影響

IGF-1是一類功能復(fù)雜的多肽因子，可促進(jìn)乳腺泌乳，特別是介導(dǎo) GH的催乳作用[7]。研究表明，乳腺組織中存在IGF-1及其受體，對(duì)早期泌乳奶牛注射GH，增加了IGF-1及其受體表達(dá)量[17]，另外，IGF-1可直接通過(guò)受體作用于乳腺，對(duì)乳腺最終分化完全、發(fā)育成熟與乳汁生成以及新生兒的生長(zhǎng)發(fā)育具有調(diào)控作用[41]，MURNEY 等[42]和 HERNANDEZ 等[43]也表示IGF-1與奶牛產(chǎn)奶量相關(guān)性顯著。本研究結(jié)果表示，在奶?；厝檫^(guò)程中，血清IGF-1含量依次呈階梯式下降，而第0和1天、第3和5天、第7和9天、第9和11天之間差異均不顯著（P＞0.05），與上述結(jié)果趨勢(shì)一致，可能是由于GH降低，其與IGF-1結(jié)合減少，反饋抑制乳腺泌乳細(xì)胞分泌。表明奶牛IGF-1不僅在奶牛青春期乳腺生長(zhǎng)和泌乳期起正向調(diào)控，且在奶?；厝槠诳山档虸GF-1含量，促進(jìn)回乳進(jìn)行。

王昶[44]和唐中生等[45]發(fā)現(xiàn)丹參可使大鼠血清和肝組織IGF-1表達(dá)量上升，張弛等[46]利用丹參、黃芩等組成的復(fù)方天然植物制劑在兔上的研究結(jié)果類似，石寶明等[13]利用益母草、王不留行、黃芩等組成的復(fù)方天然植物制劑顯著增加泌乳大鼠產(chǎn)奶量及IGF-1含量。本試驗(yàn)結(jié)果顯示，在奶?；厝檫^(guò)程中，對(duì)照組血清IGF-1含量較復(fù)方天然植物制劑組下降慢，復(fù)方天然植物制劑500和600g組血清IGF-1含量下降較400g組快，而復(fù)方天然植物制劑組500和600g差異不顯著（P＞0.05）。表明復(fù)方天然植物制劑降IGF-1效果明顯，其中500、600g組效果較好，在回乳期后復(fù)方天然植物制劑IGF-1含量均上升，表明復(fù)方天然植物制劑可使奶?？焖倩厝椋瑴p少IGF-1低含量持續(xù)時(shí)間，間接促進(jìn)IGF-1對(duì)GH的介導(dǎo)作用，加快胎犢生長(zhǎng)發(fā)育。

3.6 血清INS、HC、GH、IGF-1含量和產(chǎn)奶量間的相關(guān)性

神經(jīng)內(nèi)分泌的多種激素與乳腺外組織及乳腺分泌的多種生長(zhǎng)因子相互協(xié)同，以內(nèi)分泌、旁分泌和自分泌等方式共同調(diào)節(jié)乳腺的生長(zhǎng)發(fā)育和泌乳[5]。有研究表明，IGF-1可介導(dǎo)GH的催乳作用，提高奶牛產(chǎn)奶量[47]，而INS與GH在泌乳過(guò)程中存在拮抗，當(dāng)血清中存在高濃度的 INS，其調(diào)控葡萄糖轉(zhuǎn)運(yùn)至非乳腺組織，使乳腺對(duì)葡萄糖的攝取不敏感，而IGF-1介導(dǎo)的GH在低水平INS下可調(diào)節(jié)乳腺和乳腺外營(yíng)養(yǎng)競(jìng)爭(zhēng)，使?fàn)I養(yǎng)向乳腺轉(zhuǎn)移，促進(jìn)乳腺發(fā)育和乳的合成[48]。本研究顯示，奶?；厝槠冢珿H含量與HC含量、IGF-1含量和產(chǎn)奶量呈極顯著正相關(guān)（P＜0.01），INS含量與IGF-1、HC含量、GH含量和奶產(chǎn)量呈極顯著負(fù)相關(guān)（P＜0.01），另有研究資料表明，HC可促進(jìn)乳腺對(duì)葡萄糖的攝取，提升血清IGF-1含量，增加泌乳量[47]，本研究顯示，奶牛回乳期，IGF-1含量、HC含量和產(chǎn)奶量?jī)蓛砷g呈極顯著正相關(guān)（P＜0.01），表明在奶牛回乳期，IGF-1、GH和HC對(duì)奶牛回乳起負(fù)向調(diào)控功能，而INS對(duì)奶?；厝槠鹫{(diào)控作用。

4 結(jié)論

復(fù)方天然植物制劑能顯著促進(jìn)回乳期奶?；厝?，飼喂500g/日復(fù)方天然植物制劑，奶?；厝闀r(shí)間為5d，回乳效果即可達(dá)到最佳。回乳期奶牛血清胰島素、氫化可的松、胰島素樣生長(zhǎng)因子1含量和產(chǎn)奶量變化兩兩間均呈極顯著相關(guān)，生長(zhǎng)激素、胰島素樣生長(zhǎng)因子1和氫化可的松對(duì)奶?；厝槠鹭?fù)向調(diào)控作用，而胰島素對(duì)奶?；厝槠鹫{(diào)控作用。

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（責(zé)任編輯 林鑒非）

Effects of Compound Natural Plant Preparation on Milk Withdrawal and Galactin in Dairy Cows

SHEN LiuHong, XIAO JinBang, WU XiaoFeng, JIANG SiXun, JIANG Tao, DENG JunLiang,ZUO ZhiCai, YU ShuMin, CAO SuiZhong
(The Key Laboratory of Animal Disease and Human Health of Sichuan Province/The Medical Research Center for Cow Disease,College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130)

2016-11-30；接受日期：2017-07-20

四川省教育廳重點(diǎn)項(xiàng)目（15ZA0024）、四川農(nóng)業(yè)大學(xué)雙支計(jì)劃（03571303）、四川農(nóng)業(yè)大學(xué)本科科研興趣培養(yǎng)計(jì)劃（KY2016062）

聯(lián)系方式：沈留紅，E-mail：shenlh@sicau.edu.cn。肖勁邦，E-mail：811031312@qq.com。沈留紅和肖勁邦為同等貢獻(xiàn)作者。通信作者曹隨忠，E-mail：suizhongcao@126.com