【摘要】：中獸醫(yī)病因病理及辯證診治的基礎理論認為,熱應激是由于高熱邪氣所致的陽盛陰虛病證,而中藥女貞子具有滋陰補腎之功效；本課題組前期研究結果顯示富硒女貞子能有效降低急性熱應激小鼠的死亡率,緩解熱應激造成小鼠的肝臟損傷程度,既提高熱應激小鼠血清總蛋白、球蛋白和白蛋白的含量及免疫功能。因此我們對富硒女貞子進行了抗山羊熱應激的研究,通過富硒女貞子安全性試驗和抗熱應激療效試驗,觀察山羊生產(chǎn)性能、血液學、血清生化指標及對山羊外周血淋巴細胞增殖活性、細胞因子分泌、抗氧化活性的影響,來探索其抗山羊熱應激的效果及作用機制。試驗一富硒女貞子對山羊的安全性試驗旨在研究富硒女貞子對山羊的用藥安全性。選取30頭成年產(chǎn)奶山羊隨機均分為5組,其中C組飼喂基礎日糧,CLL組基礎日糧中添加女貞于30 g/頭/d,LSLL組、MSLL組、HSLL基礎日糧中分別添加富硒女貞子10g/頭/d、30 g/頭/d、50 g/頭/d,給藥組于試驗開始的第5d給藥,連續(xù)5d。經(jīng)產(chǎn)奶量、血液中WBC、RBC、HGB含量、血清中TP、ALB、 ALT、AST、CK、LDH的水平測定。結果顯示：富硒女貞子和普通女貞子均能提高山羊的產(chǎn)奶量,在試驗1～7、22-38 d產(chǎn)奶量增長率由大到小的排序為：LSLL組MSLL組HSLL組CLL組C組；CLL組能顯著提高山羊RBC(P0.05),MSLL組、CLL組均能顯著提高山羊HGB含量(P0.05)；LSLL組、MSLL組、HSLL組和CLL組均能顯著提高山羊血清中TP含量(P0.05),LSLL組、MSLL組和CLL組均能顯著提高山羊血清中ALB含量(P0.05),LSLL組能顯著降低血清中AST活性(P0.05)；添加富硒女貞子對山羊WBC含量、ALT、CK、LDH活性均無顯著影響(P0.05)。說明日糧中添加富硒女貞子在提高山羊產(chǎn)奶量的同時,亦提高了血液RBC、HGB和血清TP、ALB的含量,降低了血清AST活性,而對山羊WBC、ALT、CK、LDH舌性影響不明顯,提示基礎日糧中按≤50 g/頭/d劑量添加富硒女貞子,連用5 d,對山羊是安全的。試驗二富硒女貞子對山羊熱應激的影響為觀察富硒女貞子對熱應激山羊生產(chǎn)性能、血液學和血清生化指標的影響。選取30頭泌乳期薩能山羊隨機均分為5組,其中HS組飼喂基礎日糧,CLL組基礎日糧中添加女貞子10g/頭/d,LSLL組、MSLL組、HSLL組基礎日糧中分別添加富硒女貞子5 g/頭/d、10 g/頭/d、20 g/頭/d。試驗期為2014年7月10日至8月27日,給藥組于每旬前3天混飼給藥,給藥4批次。經(jīng)產(chǎn)奶量、乳成分、血液中WBC、RBC、HGB含量、血清中TP、ALB、ALT、AST、LDH、IL-2、IFN-γ、IL-6、SOD、GSH-Px、ACTH、COR的水平測定。結果顯示：日糧中添加富硒女貞子能提高熱應激山羊的產(chǎn)奶量,產(chǎn)奶量下降率由小到大的排序為：MSLL組LSLL組HSLL組CLL組HS組,其中MSLL組較非熱應激期增加了2.89%,極顯著高于HS組的下降10.07%(P0.01)；顯著升高熱應激山羊乳脂肪含量(P0.05)、提高了乳蛋白含量；明顯提高了血清中TP、ALB含量；28 d時HSLL組的HSP70表達量、LSLL組的LDH活性顯著低于HS組,LSLL組的IL-2、IFN-γ含量、SOD活性顯著高于HS組(P0.05),14 d時MSLL組、HSLL組、CLL組的COR濃度均顯著低于HS組(P0.05),對乳糖、總乳固體、非脂乳固體、WBC、RBC、HGB、ALT、AST、 IL-6的影響不明顯(P0.05)。表明富硒女貞子對熱應激山羊具有通過調(diào)節(jié)內(nèi)分泌-免疫、改善機體代謝而提高產(chǎn)奶量的作用,尤以5～10g/頭/d富硒女貞子的作用效果為佳。試驗三富硒女貞子對山羊外周血淋巴細胞增殖活性、細胞因子分泌及抗氧化活性的影響分析富硒女貞子對山羊外周血淋巴細胞增殖活性、細胞因子分泌及抗氧化活性的影響。采用MTT法測定富硒女貞子多糖、蛋白質、齊墩果酸對山羊外周血淋巴細胞增殖活性的影響,并且對其培養(yǎng)上清細胞因子IL-2、IL-4、IL-6、IFN-γ水平和SOD活性進行檢測。結果顯示：富硒女貞子多糖、蛋白質和齊墩果酸體外均具有促進ConA刺激山羊外周血淋巴細胞分裂增殖的作用,且富硒女貞子蛋白質組分、齊墩果酸組分的作用效果明顯優(yōu)于普通女貞子組,各相應濃度比較差異顯著比例分別達到63.64%、40.00%1富硒和普通女貞子均極顯著提高外周血淋巴細胞中SOD的活性(P0.01),且富硒女貞子的作用效果明顯優(yōu)于普通女貞子；富硒女貞子可刺激淋巴細胞產(chǎn)生IL-2,但與對照組比較差異不顯著(P0.05)。結果表明富硒女貞子多糖、蛋白質和齊墩果酸均具有促進山羊外周血淋巴細胞增殖活性的作用,同時明顯提高其抗氧化能力。結論：富硒女貞子,按≤50g/頭/d,連用5 d,對山羊具有較好的安全性：能緩解山羊熱應激影響導致的產(chǎn)奶量下降(提高了2.89%),以5～10g/頭/d富硒女貞子的作用效果為佳,且作用效果優(yōu)于普通女貞子。其作用機制是通過HPA軸來調(diào)低熱應激性升高的ACTH、COR,調(diào)升熱應激降低的IL-2、IL-6、IFN-γ水平,使得機體維持良好的內(nèi)環(huán)境穩(wěn)態(tài),提高其免疫功能,同時提高血清TP、ALB含量、GSH-Px、SOD活性,降低LDH活性、HSP70表達量,從而使機體降低應激性代償,維持更接近正常期的產(chǎn)奶量。
[Abstract]:According to the basic theory of etiology, pathology and dialectical diagnosis and treatment of Chinese veterinary medicine, heat stress is due to yang-Sheng-yin deficiency caused by high-heat pathogenic qi, while Ligustrum lucidum has the effect of nourishing yin and tonifying kidney. The degree of visceral injury can not only increase the content of serum total protein, globulin and albumin, but also improve the immune function of heat-stressed mice.Therefore, we studied the anti-heat stress effect of Se-enriched Ligustrum lucidum on goats. To explore the effect and mechanism of anti-heat stress on peripheral blood lymphocyte proliferation, cytokine secretion and antioxidant activity. Experiment 1 The safety test of Se-enriched Ligustrum lucidum on goats was designed to study the medication safety of Se-enriched Ligustrum lucidum on goats. The basic diet of CLL group was fed with 30 g/head/d, LSLL group, MSLL group, HSLL base diet was fed with 10 g/head/d, 30 g/head/d, 50 g/head/d of Ligustrum lucidum enriched with selenium, and the treatment group was fed on the 5th day of the experiment. The milk yield, WBC, RBC, HGB content in blood, TP, ALB, ALT, AST, CK, LDH levels in serum were measured. The results showed that both Se-enriched Ligustrum lucidum and common Ligustrum lucidum could increase the milk yield of goats. The order of increase rate of milk yield from large to small was as follows: LSLL group MSLL group CLL group C group; CLL group RBC (P 0.05), MSLL group, CLL group could significantly increase the HGB content of goats (P 0.05); LL group can significantly increase the content of TP in goat serum (P 0.05), LSLL group, MSLL group and CLL group can significantly increase the content of ALB in goat serum (P 0.05), LSLL group can significantly reduce the activity of AST in serum (P 0.05); adding Se-enriched Ligustrum lucidum has no significant effect on WBC content, ALT, CK, LDH activity in goats (P 0.05). The content of RBC, HGB, TP and ALB in blood and the activity of AST in serum were increased, but the tongue of WBC, ALT, CK and LDH in goats were not affected obviously. It was suggested that adding Se-enriched Ligustrum lucidum to basal diet for 5 days was safe for goats. Thirty lactating Saanen goats were randomly divided into 5 groups, including HS group fed basal diet, CLL group fed basal diet supplemented with 10g Ligustrum lucidum per day, LSLL group, MSLL group, HSLL group fed basal diet supplemented with 5g Ligustrum lucidum enriched with selenium respectively. D, 10 g / head / d, 20 g / head / d. The trial period was from July 10 to August 27, 2014. The administration group was administered in four batches three days before every ten days. The milk yield, milk composition, WBC, RBC, HGB content in blood, serum TP, ALB, ALT, AST, LDH, IL-2, IFN-gamma, IL-6, SOD, GSH-Px, ACTH, COR levels were determined. The order of decreasing rate of milk yield and milk yield of high heat stress goats from small to large was: HS group of LSLL group of MSLL group of HSLL group, HS group of CLL group, MSLL group increased by 2.89% compared with non-heat stress group, significantly higher than HS group decreased by 10.07% (P 0.01); significantly increased milk fat content of heat stress goats (P 0.05), increased milk protein content; The expression of HSP70 in HSLL group and LDH activity in LSLL group were significantly lower than those in HS group at 28 d, while the content of IL-2, IFN-gamma and SOD activity in LSLL group were significantly higher than those in HS group (P 0.05). The COR concentration in MSLL group, HSLL group and CLL group was significantly lower than that in HS group at 14 d (P 0.05), and the effects on lactose, total milk solids, non-fat milk solids, WBC, RBC, HGB, ALT, AST, IL-6 were not clear. The results showed that the Se-enriched Ligustrum lucidum could increase milk yield by regulating Endocrine-Immunity and improving metabolism of the body. The effect of Se-enriched Ligustrum lucidum 5-10g/head/d was the best. The effects of Se-enriched Ligustrum lucidum on the proliferation, cytokine secretion and antioxidant activity of peripheral blood lymphocytes of goats were analyzed.The effects of Se-enriched Ligustrum lucidum polysaccharide, protein and oleanolic acid on the proliferation activity of peripheral blood lymphocytes of goats were determined by MTT method. The results showed that Se-enriched Ligustrum lucidum polysaccharides, proteins and oleanolic acid could promote the proliferation of peripheral blood lymphocytes stimulated by ConA in vitro, and the effects of Se-enriched Ligustrum lucidum protein components and oleanolic acid components were significantly better than those of ordinary Ligustrum lucidum group, and the corresponding concentrations were significantly different in proportion. Selenium-enriched Ligustrum lucidum L. and common Ligustrum lucidum L. both significantly increased SOD activity in peripheral blood lymphocytes (P 0.01), and the effect of Se-enriched Ligustrum lucidum L. was better than that of common Ligustrum lucidum L. Se-enriched Ligustrum L. could stimulate lymphocytes to produce IL-2, but there was no significant difference between the two groups (P 0.05). Conclusion: Selenium-enriched Ligustrum lucidum has better safety for goats when it is used for 5 days at a dose of <50g/head/d. It can alleviate the decrease of milk yield caused by heat stress in goats (2.89%) and enrich them at a dose of 5-10g/head/d. Ligustrum selenium has a better effect than Ligustrum lucidum. Its mechanism is to lower the levels of ACTH, COR, IL-2, IL-6 and IFN-gamma in the elevated heat stress through HPA axis, so that the body can maintain a good homeostasis of the internal environment, improve its immune function, and increase the content of TP, ALB, GSH-Px, SOD activity in serum. Reduce the activity of LDH and the expression of HSP70, so that the body can reduce stress compensation and maintain milk production closer to normal period.
【學位授予單位】：揚州大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：S853.7

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