硼對非洲雛鴕鳥肺纖維化的影響
發(fā)布時(shí)間：2018-08-29 19:18
【摘要】：硼是動物機(jī)體生長發(fā)育的必須微量元素,在機(jī)體中參與了多項(xiàng)生理活動。硼的主要吸收形式是無機(jī)鹽,無機(jī)硼酸鹽主要通過胃腸道、呼吸道和破損皮膚吸收。呼吸系統(tǒng)與外界相通因而很容易受到外來刺激物的損傷。雛鴕鳥的肺組織尚未發(fā)育完善,易受到外界病原攻擊,是呼吸系統(tǒng)疾病的易感動物,雛鴕鳥有著高的患病率和死亡率。目前關(guān)于硼對機(jī)體諸多生理功能的研究已相繼報(bào)道,尤其是硼對骨、腦和免疫器官等的作用,含硼化合物對癌癥的治療等,而有關(guān)硼對呼吸系統(tǒng)的作用尚未報(bào)道。為了探明雛鴕鳥呼吸系統(tǒng)對硼的耐受程度,本研究以肺為研究對象,通過飲水添加不同濃度的硼酸,探明硼對非洲雛鴕鳥肺纖維化的影響。剛出殼的1 d鴕鳥,隨機(jī)分為6組,分別在飲水中添加濃度為0 mg/L(A組)、40 mg/L(B組)、80 mg/L(C組)、160 mg/L(D組)、320 mg/L(E組)、640 mg/L(F組)的硼酸,飼養(yǎng)至90 d取肺組織進(jìn)行石蠟切片,HE和MASSON染色,SOD、MDA、GSH測定,TUNEL技術(shù)和實(shí)時(shí)熒光定量PCR等方法,探討不同濃度飲水硼對非洲雛鴕鳥肺纖維化的影響。具體研究內(nèi)容如下:1硼對雛鴕鳥肺組織形態(tài)結(jié)構(gòu)的影響通過HE染色對雛鴕鳥肺組織形態(tài)結(jié)構(gòu)變化作初步評估,并對肺組織病理變化進(jìn)行標(biāo)準(zhǔn)量化分析。結(jié)果顯示,與對照組相較,硼劑量低于160 mg/L時(shí),肺組織結(jié)構(gòu)清晰,發(fā)育良好,硼劑量高于320 mg/L時(shí),肺組織形態(tài)結(jié)構(gòu)出現(xiàn)病變,異嗜性粒細(xì)胞顯著增多,血管內(nèi)有淋巴細(xì)胞,出現(xiàn)炎癥反應(yīng),細(xì)胞間質(zhì)增厚,疑似肺纖維化,細(xì)胞核有碎裂現(xiàn)象,疑似細(xì)胞凋亡。2硼對雛鴕鳥血清抗氧化能力的影響使用SOD、MDA和GSH試劑盒測定雛鴕鳥血清的SOD活性,GSH和MDA含量。結(jié)果顯示:與對照組相比,血清SOD活性在硼劑量低于160 mg/L時(shí)升高,且在硼劑量為80 mg/L時(shí)有顯著差異性(P0.05),在硼劑量高于320 mg/L時(shí)降低,且在硼劑量為640 mg/L時(shí)有顯著差異性(P0.05);血清GSH含量在硼劑量低于160mg/L時(shí)升高,且在硼劑量為80 mg/L時(shí)差異顯著(P0.05),在硼劑量高于320 mg/L時(shí)顯著降低(P0.05);血清MDA含量在硼劑量低于160 mg/L時(shí)降低,但差異不顯著(P0.05),在硼劑量高于320 mg/L時(shí)升高,且在硼劑量為640 mg/L時(shí)有顯著差異(P0.05)。綜上所述,硼劑量低于160 mg/L時(shí),可提高雛鴕鳥血清SOD活性和GSH含量,降低血清MDA含量,提高機(jī)體抗氧化能力;硼劑量高于320 mg/L時(shí),可降低鴕鳥血清SOD活性和GSH含量,提高血清MDA含量,降低機(jī)體抗氧化能力。3硼對雛鴕鳥肺組織炎癥反應(yīng)的影響通過HE染色結(jié)果可知,硼劑量高于320 mg/L時(shí)雛鴕鳥肺組織異嗜性粒細(xì)胞增多,有炎癥反應(yīng)。為驗(yàn)證該結(jié)果,本試驗(yàn)通過熒光定量PCR技術(shù)對肺組織IL-1β、IL-6、IL-10 m RNA表達(dá)水平進(jìn)行相對定量。與對照組相比,IL-1βm RNA表達(dá)水平在硼劑量為40-80 mg/L時(shí)降低,但差異不顯著,在160 mg/L時(shí)顯著升高(P0.05),在320-640 mg/L時(shí)升高極顯著(P0.01);IL-6 m RNA表達(dá)水平在硼劑量為40-80mg/L時(shí)降低,但是降低不顯著,在160 mg/L時(shí)升高,且升高不顯著,在320-640 mg/L時(shí)升高極顯著(P0.01);IL-10 m RNA表達(dá)水平一直在升高,在40 mg/L時(shí)變化不顯著,在80-160 mg/L時(shí)顯著升高(P0.05),在320-640 mg/L時(shí)升高極顯著(P0.01)。由此可見硼劑量低于160 mg/L時(shí)可抑制細(xì)胞炎癥反應(yīng);當(dāng)硼劑量高于320 mg/L時(shí),IL-1β、IL-6、IL-10 m RNA表達(dá)水平均顯著升高(P0.01),在促炎因子IL-1β、IL-6引起炎癥反應(yīng)時(shí),抗炎因子IL-10也隨著增多以抑制過度炎癥反應(yīng),這是高劑量硼酸引起炎癥反應(yīng)的結(jié)果。由此得出硼劑量為40-160 mg/L時(shí)可抑制炎癥反應(yīng),硼劑量為320-640 mg/L時(shí)可促進(jìn)炎癥反應(yīng)。4硼對雛鴕鳥肺組織細(xì)胞凋亡的影響已知HE染色在高倍顯微鏡(1000×)下觀察到,硼劑量高于320 mg/L時(shí),肺組織細(xì)胞核有碎裂現(xiàn)象,疑似細(xì)胞凋亡。為應(yīng)證該結(jié)論,采用TUNEL技術(shù),熒光顯微鏡成像發(fā)現(xiàn),硼劑量低于160 mg/L時(shí)顯著抑制細(xì)胞凋亡,硼劑量高于320 mg/L時(shí)促進(jìn)細(xì)胞凋亡。運(yùn)用熒光定量PCR技術(shù)對caspase-3 m RNA表達(dá)水平進(jìn)行相對定量測定,與對照組相比,硼劑量低于160 mg/L時(shí)caspase-3 m RNA表達(dá)水平顯著降低(P0.05),硼劑量高于320 mg/L時(shí)caspase-3 m RNA表達(dá)水平顯著增加(P0.05)。5硼對雛鴕鳥肺纖維化的影響利用MASSON染色標(biāo)記膠原纖維,并進(jìn)行IOD值統(tǒng)計(jì)分析。結(jié)果發(fā)現(xiàn),硼劑量高于320 mg/L時(shí)肺組織膠原纖維明顯增加。本試驗(yàn)以細(xì)胞外基質(zhì)和上皮間質(zhì)轉(zhuǎn)化為肺纖維化的直接評定指標(biāo),為探明該現(xiàn)象,本試驗(yàn)通過熒光定量PCR技術(shù)對ECM、E-Cadherin、a-SMA和Vimentin m RNA表達(dá)水平進(jìn)行相對定量。隨著硼劑量的增加,ECM、a-SMA和Vimentin m RNA表達(dá)水平呈上升趨勢,E-Cadherin m RNA表達(dá)水平呈下降趨勢。通過以上指標(biāo)驗(yàn)證,得出硼劑量低于160 mg/L時(shí)對肺纖維化的作用不明顯(P0.05),硼劑量高于320 mg/L時(shí)能顯著促進(jìn)肺纖維化(P0.05)的結(jié)論。綜合以上試驗(yàn)結(jié)果,得出硼劑量低于160 mg/L時(shí)可提高血清抗氧化能力,抑制肺組織發(fā)生炎癥反應(yīng)和細(xì)胞凋亡,進(jìn)而降低肺纖維化的發(fā)生;硼劑量高于320 mg/L時(shí)可損傷血清抗氧化能力,促進(jìn)肺組織炎性損傷和細(xì)胞凋亡,并增加肺纖維化的發(fā)生發(fā)展。
[Abstract]:Boron is an essential trace element for the growth and development of animals and participates in many physiological activities in the body. The main absorption form of boron is inorganic salts. Inorganic borates are mainly absorbed through the gastrointestinal tract, respiratory tract and damaged skin. Respiratory system is connected with the outside world and therefore vulnerable to foreign stimuli. The lung tissue of ostrich chicks has not yet been found. Being well-developed and vulnerable to external pathogens, ostrich is a susceptible animal to respiratory diseases. Young ostrich has a high morbidity and mortality rate. At present, studies on the physiological functions of boron have been reported, especially the effects of boron bone, brain and immune organs, boron-containing compounds on cancer treatment, etc., while boron on the respiratory system. The effects of Boron on pulmonary fibrosis in African ostrich chicks were studied by adding boric acid to drinking water. The newly-hatched ostriches were randomly divided into 6 groups: 0 mg/L (group A) and 40 mg/L (group B). 80 mg/L (group C), 160 mg/L (group D), 320 mg/L (group E), 640 mg/L (group F) of boric acid were fed to 90 days for paraffin section, HE and MASSON staining, SOD, MDA, GSH determination, TUNEL and real-time fluorescence quantitative PCR to investigate the effects of different concentrations of Boron on pulmonary fibrosis in African ostrich chicks. The results showed that the lung tissue structure was clear and well developed when boron dosage was lower than 160 mg/L, and when boron dosage was higher than 320 mg/L, the lung tissue structure was out of shape. Now pathological changes, heterophilic granulocytes increased significantly, lymphocytes in the blood vessels, inflammation, thickening of the interstitial cells, suspected pulmonary fibrosis, nuclear fragmentation, suspected apoptosis. 2 boron on the serum antioxidant capacity of ostrich chicks using SOD, MDA and GSH kits to determine the serum SOD activity, GSH and MDA content. Compared with the control group, the serum SOD activity increased when the boron dose was lower than 160 mg/L, and there was a significant difference when the boron dose was 80 mg/L (P 0.05), decreased when the boron dose was higher than 320 mg/L, and there was a significant difference when the boron dose was 640 mg/L (P 0.05); the serum GSH content increased when the boron dose was lower than 160 mg/L, and the boron dose was 80 mg/L. There was a significant difference (P 0.05), when the boron dose was higher than 320 mg/L, it decreased significantly (P 0.05); the serum MDA content was lower when the boron dose was lower than 160 mg/L, but the difference was not significant (P 0.05), when the boron dose was higher than 320 mg/L, and when the boron dose was 640 mg/L, there was a significant difference (P 0.05). When boron dosage was higher than 320 mg/L, SOD activity and GSH content in serum of ostrich could be decreased, MDA content in serum could be increased and antioxidant capacity could be decreased. The effect of Boron on inflammatory reaction of lung tissue of ostrich was observed by HE staining. In order to verify the results, the expression levels of IL-1 beta, IL-6 and IL-10 m RNA in ostrich lung tissues were quantified by fluorescence quantitative PCR. Compared with the control group, the expression levels of IL-1 beta m RNA in ostrich lung tissues decreased at boron dosage of 40-80 mg/L, but the difference was not significant. The expression levels increased significantly at 160 mg/L. (P 0.05), at 320-640 mg/L increased significantly (P 0.01); IL-6 m RNA expression level decreased at boron dosage of 40-80 mg/L, but the decrease was not significant, at 160 mg/L increased, and the increase was not significant, at 320-640 mg/L increased extremely significantly (P 0.01); IL-10 m RNA expression level has been increasing, at 40 mg/L did not change significantly, at 80-160 mg/L significantly. The levels of IL-1 beta, IL-6 and IL-10 m RNA were significantly increased when the dose of boron was lower than 160 mg/L (P 0.05). The levels of IL-10, IL-1 beta and IL-6 were inhibited when the dose of boron was higher than 320 mg/L (P 0.01). Excessive inflammation is the result of inflammation induced by high dose of boric acid. It is concluded that boron dose is 40-160 mg/L and 320-640 mg/L can inhibit inflammation. The effect of Boron on apoptosis of lung tissue in ostrich chicks is known to be observed under high power microscopy (1000 x). Boron dose is higher than 320 mg/L. To confirm this conclusion, TUNEL and fluorescence microscopy showed that boron dosage below 160 mg/L significantly inhibited cell apoptosis, and boron dosage above 320 mg/L promoted cell apoptosis. The expression of Caspase-3 m RNA was quantitatively determined by fluorescence quantitative PCR. Compared with the control group, the expression level of Caspase-3 m RNA was significantly decreased (P 0.05) when boron dosage was lower than 160 mg/L, and significantly increased (P 0.05) when boron dosage was higher than 320 mg/L. The effect of boron pulmonary fibrosis in ostrich chicks was studied by MASSON staining and IOD value was statistically analyzed. The expression levels of ECM, E-Cadherin, a-SMA and Vimentin m RNA were quantified by fluorescence quantitative PCR. With the increase of boron dosage, the expression levels of ECM, a-SMA and Vimentin m RNA were measured. The expression of M RNA increased and the expression of E-Cadherin m RNA decreased. The results showed that the effect of boron dosage below 160 mg/L on pulmonary fibrosis was not obvious (P 0.05), and that boron dosage above 320 mg/L could significantly promote pulmonary fibrosis (P 0.05). When boron dosage is higher than 320 mg/L, the serum antioxidant capacity can be damaged, the inflammatory injury and cell apoptosis of lung tissue can be promoted, and the occurrence and development of pulmonary fibrosis can be increased.
【學(xué)位授予單位】：華中農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S858.39
本文編號：2212160
[Abstract]:Boron is an essential trace element for the growth and development of animals and participates in many physiological activities in the body. The main absorption form of boron is inorganic salts. Inorganic borates are mainly absorbed through the gastrointestinal tract, respiratory tract and damaged skin. Respiratory system is connected with the outside world and therefore vulnerable to foreign stimuli. The lung tissue of ostrich chicks has not yet been found. Being well-developed and vulnerable to external pathogens, ostrich is a susceptible animal to respiratory diseases. Young ostrich has a high morbidity and mortality rate. At present, studies on the physiological functions of boron have been reported, especially the effects of boron bone, brain and immune organs, boron-containing compounds on cancer treatment, etc., while boron on the respiratory system. The effects of Boron on pulmonary fibrosis in African ostrich chicks were studied by adding boric acid to drinking water. The newly-hatched ostriches were randomly divided into 6 groups: 0 mg/L (group A) and 40 mg/L (group B). 80 mg/L (group C), 160 mg/L (group D), 320 mg/L (group E), 640 mg/L (group F) of boric acid were fed to 90 days for paraffin section, HE and MASSON staining, SOD, MDA, GSH determination, TUNEL and real-time fluorescence quantitative PCR to investigate the effects of different concentrations of Boron on pulmonary fibrosis in African ostrich chicks. The results showed that the lung tissue structure was clear and well developed when boron dosage was lower than 160 mg/L, and when boron dosage was higher than 320 mg/L, the lung tissue structure was out of shape. Now pathological changes, heterophilic granulocytes increased significantly, lymphocytes in the blood vessels, inflammation, thickening of the interstitial cells, suspected pulmonary fibrosis, nuclear fragmentation, suspected apoptosis. 2 boron on the serum antioxidant capacity of ostrich chicks using SOD, MDA and GSH kits to determine the serum SOD activity, GSH and MDA content. Compared with the control group, the serum SOD activity increased when the boron dose was lower than 160 mg/L, and there was a significant difference when the boron dose was 80 mg/L (P 0.05), decreased when the boron dose was higher than 320 mg/L, and there was a significant difference when the boron dose was 640 mg/L (P 0.05); the serum GSH content increased when the boron dose was lower than 160 mg/L, and the boron dose was 80 mg/L. There was a significant difference (P 0.05), when the boron dose was higher than 320 mg/L, it decreased significantly (P 0.05); the serum MDA content was lower when the boron dose was lower than 160 mg/L, but the difference was not significant (P 0.05), when the boron dose was higher than 320 mg/L, and when the boron dose was 640 mg/L, there was a significant difference (P 0.05). When boron dosage was higher than 320 mg/L, SOD activity and GSH content in serum of ostrich could be decreased, MDA content in serum could be increased and antioxidant capacity could be decreased. The effect of Boron on inflammatory reaction of lung tissue of ostrich was observed by HE staining. In order to verify the results, the expression levels of IL-1 beta, IL-6 and IL-10 m RNA in ostrich lung tissues were quantified by fluorescence quantitative PCR. Compared with the control group, the expression levels of IL-1 beta m RNA in ostrich lung tissues decreased at boron dosage of 40-80 mg/L, but the difference was not significant. The expression levels increased significantly at 160 mg/L. (P 0.05), at 320-640 mg/L increased significantly (P 0.01); IL-6 m RNA expression level decreased at boron dosage of 40-80 mg/L, but the decrease was not significant, at 160 mg/L increased, and the increase was not significant, at 320-640 mg/L increased extremely significantly (P 0.01); IL-10 m RNA expression level has been increasing, at 40 mg/L did not change significantly, at 80-160 mg/L significantly. The levels of IL-1 beta, IL-6 and IL-10 m RNA were significantly increased when the dose of boron was lower than 160 mg/L (P 0.05). The levels of IL-10, IL-1 beta and IL-6 were inhibited when the dose of boron was higher than 320 mg/L (P 0.01). Excessive inflammation is the result of inflammation induced by high dose of boric acid. It is concluded that boron dose is 40-160 mg/L and 320-640 mg/L can inhibit inflammation. The effect of Boron on apoptosis of lung tissue in ostrich chicks is known to be observed under high power microscopy (1000 x). Boron dose is higher than 320 mg/L. To confirm this conclusion, TUNEL and fluorescence microscopy showed that boron dosage below 160 mg/L significantly inhibited cell apoptosis, and boron dosage above 320 mg/L promoted cell apoptosis. The expression of Caspase-3 m RNA was quantitatively determined by fluorescence quantitative PCR. Compared with the control group, the expression level of Caspase-3 m RNA was significantly decreased (P 0.05) when boron dosage was lower than 160 mg/L, and significantly increased (P 0.05) when boron dosage was higher than 320 mg/L. The effect of boron pulmonary fibrosis in ostrich chicks was studied by MASSON staining and IOD value was statistically analyzed. The expression levels of ECM, E-Cadherin, a-SMA and Vimentin m RNA were quantified by fluorescence quantitative PCR. With the increase of boron dosage, the expression levels of ECM, a-SMA and Vimentin m RNA were measured. The expression of M RNA increased and the expression of E-Cadherin m RNA decreased. The results showed that the effect of boron dosage below 160 mg/L on pulmonary fibrosis was not obvious (P 0.05), and that boron dosage above 320 mg/L could significantly promote pulmonary fibrosis (P 0.05). When boron dosage is higher than 320 mg/L, the serum antioxidant capacity can be damaged, the inflammatory injury and cell apoptosis of lung tissue can be promoted, and the occurrence and development of pulmonary fibrosis can be increased.
【學(xué)位授予單位】：華中農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S858.39
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