生长抑素DNA免疫对小鼠体组织的影响及表达产物的分布规律
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摘要
本研究应用免疫组织化学(streptavidin-biotin-peroxidase-complex, SABC)、HE染色等技术,在无抗性筛选生长抑素DNA疫苗(pGS/2SS-M4GFP-asd)肌肉注射7周龄的雄性昆明小鼠后,检测DNA疫苗表达产物在小鼠各组织分布的细胞类型和消长规律,并且对其免疫效果进行评价。旨在探讨疫苗的免疫机制和安全性,为开发高效、安全的促进动物生长的DNA疫苗,并加快其临床应用奠定基础。
1生长抑素DNA免疫的表达产物在小鼠体组织中的分布
将生长抑素DNA疫苗肌肉注射免疫7周龄的雄性昆明小鼠,分别于免疫后24h、48h、5d、7d、9d、11d采集小鼠心脏、肝脏、脾脏、肺脏、肾脏、肌肉组织,经固定、浸蜡、包埋、组织切片后,免疫组织化学染色,旨在探讨生长抑素DNA疫苗的表达产物在小鼠各组织细胞中的分布。结果表明：DNA疫苗表达产物主要分布于肌肉组织的中性粒细胞、巨噬细胞、成纤维细胞,脾脏组织中的巨噬细胞,肝脏组织中的肝细胞,而心脏、肺脏、肾脏各组织细胞中尚未检测到SS阳性信号。
2生长抑素DNA免疫的表达产物在小鼠体组织中的消长规律
分析上述不同时期的切片结果,探讨生长抑素DNA疫苗的表达产物在小鼠各组织及各种细胞中的消长规律。结果表明：肌肉组织中成纤维细胞在免疫24h后,能摄取减毒沙门氏菌并在其胞浆中出现了疫苗表达产物,且在免疫48 h达到高峰,而在免疫第5d后成纤维细胞中检测不到SS阳性信号。从免疫后7d至11d,可在巨噬细胞中检测到疫苗表达产物的分布。
免疫48 h后,显微显微镜下可观察到疫苗表达产物分布于脾脏组织的巨噬细胞的胞浆中,一直持续到免疫后11 d。与此类似的是,在免疫48 h后可观察到疫苗表达产物在肝细胞的胞浆中的分布,但是仅持续到免疫后第7 d。
3生长抑素DNA免疫对小鼠体组织的影响
为了分析生长抑素DNA疫苗中所含有的猪霍乱减毒沙门氏活菌对小鼠的生长发育是否有毒害作用,本试验对小鼠进行了全程健康状况观察记录,并用HE染色方法分析了各种组织切片在不同时期的变化情况,结果在整个实验过程中未观察到小鼠的死亡,猪霍乱减毒沙门氏活菌对肝脏造成一定的病理损伤,但是小鼠在免疫第7d后肝脏能对损伤的组织进行自我修复,并能维持肝脏正常的机能,除此之外,未检测到其他组织有所病理性损伤。
This study discussed the immunologic mechanism of DNA vaccine and its safety. The somatostatin DNA vaccine without antibiotic resistance gene (pGS/2SS-M4GFP-asd) was injected to the male Kunming mice of 7 weeks old. The technologies of streptavidin-biotin-peroxidase-complex and HE staining techniques were used to detect the somatostatin which was expressed by DNA vaccine. The SS in Kunming mice tissue distribution, descents and ascent dynamics were studied. The results of this study can provide a theory reference to the DNA vaccine safety apply.
1 The expression products of somatostatin DNA vaccine in mice was distributed in body tissue
In order to analyze the tissues distribution of SS DNA vaccine expression products, the SS DNA vaccine was injected to the Kunming mice of 7 weeks old. The heart, liver, spleen, lung, kidney, muscle tissue were taken after 24 h、48 h、5 d、7 d、9 d、11d, respectively. The tissues were embedded in paraffin, and then the tissue sections were stained with immunohistochemistry technique. The results indicated that the SS DNA vaccine expressions were mainly distributed in muscle tissue neutrophils, macrophages, fibroblasts, in the spleen macrophages, in the liver cells. However, little was detected in heart, lung, kidney.
2 Descents and ascent dynamics of SS DNA vaccine expression products in mice tissues
We analyzed descents and ascent dynamics of SS DNA vaccine in mice tissues with the result of slice in different experimental time. The results show that the product of SS DNA vaccine was detected. The salmonella were attacked in fibroblasts cytoplasm after 24h of injecting. Immunologic reaction rose peak after 48h and SS positive signal was lost at the 5th day. From 7 d to 11 d in the immune, the expression products of vaccine can be detected in macrophages.
48h to 11 d after immunization, the expression pruducts of vaccine in the spleen of the cytoplasm of macrophages were observed in the microscope. Similarly, after the 48 h immunization, the expression pruducts of vaccine in the cytoplasm of liver cells were observed in the microscope, but only until 7 d.
3 Somatostatin DNA vaccine impact mice tissue
We observed and recoreded the whole health of the test mice, and analyzed the results of slice in the different time with hematoxylin-eosin staining in order to analyze live attenuated Salmonella choleraesuis in SS DNA vaccine have toxic effects in mice viable growth and development. The result showed that there was no one dead in the experiment and live attenuated Salmonella choleraesuis caused certain pathological damage on liver, but the first 7 days after immunization in mice liver can damage tissue to repair it and to maintain normal liver function, In this regard, the vaccine is safe. In addition, the vaccine did not have other pathological tissue damage detection.
1生长抑素DNA免疫的表达产物在小鼠体组织中的分布
将生长抑素DNA疫苗肌肉注射免疫7周龄的雄性昆明小鼠,分别于免疫后24h、48h、5d、7d、9d、11d采集小鼠心脏、肝脏、脾脏、肺脏、肾脏、肌肉组织,经固定、浸蜡、包埋、组织切片后,免疫组织化学染色,旨在探讨生长抑素DNA疫苗的表达产物在小鼠各组织细胞中的分布。结果表明：DNA疫苗表达产物主要分布于肌肉组织的中性粒细胞、巨噬细胞、成纤维细胞,脾脏组织中的巨噬细胞,肝脏组织中的肝细胞,而心脏、肺脏、肾脏各组织细胞中尚未检测到SS阳性信号。
2生长抑素DNA免疫的表达产物在小鼠体组织中的消长规律
分析上述不同时期的切片结果,探讨生长抑素DNA疫苗的表达产物在小鼠各组织及各种细胞中的消长规律。结果表明：肌肉组织中成纤维细胞在免疫24h后,能摄取减毒沙门氏菌并在其胞浆中出现了疫苗表达产物,且在免疫48 h达到高峰,而在免疫第5d后成纤维细胞中检测不到SS阳性信号。从免疫后7d至11d,可在巨噬细胞中检测到疫苗表达产物的分布。
免疫48 h后,显微显微镜下可观察到疫苗表达产物分布于脾脏组织的巨噬细胞的胞浆中,一直持续到免疫后11 d。与此类似的是,在免疫48 h后可观察到疫苗表达产物在肝细胞的胞浆中的分布,但是仅持续到免疫后第7 d。
3生长抑素DNA免疫对小鼠体组织的影响
为了分析生长抑素DNA疫苗中所含有的猪霍乱减毒沙门氏活菌对小鼠的生长发育是否有毒害作用,本试验对小鼠进行了全程健康状况观察记录,并用HE染色方法分析了各种组织切片在不同时期的变化情况,结果在整个实验过程中未观察到小鼠的死亡,猪霍乱减毒沙门氏活菌对肝脏造成一定的病理损伤,但是小鼠在免疫第7d后肝脏能对损伤的组织进行自我修复,并能维持肝脏正常的机能,除此之外,未检测到其他组织有所病理性损伤。
This study discussed the immunologic mechanism of DNA vaccine and its safety. The somatostatin DNA vaccine without antibiotic resistance gene (pGS/2SS-M4GFP-asd) was injected to the male Kunming mice of 7 weeks old. The technologies of streptavidin-biotin-peroxidase-complex and HE staining techniques were used to detect the somatostatin which was expressed by DNA vaccine. The SS in Kunming mice tissue distribution, descents and ascent dynamics were studied. The results of this study can provide a theory reference to the DNA vaccine safety apply.
1 The expression products of somatostatin DNA vaccine in mice was distributed in body tissue
In order to analyze the tissues distribution of SS DNA vaccine expression products, the SS DNA vaccine was injected to the Kunming mice of 7 weeks old. The heart, liver, spleen, lung, kidney, muscle tissue were taken after 24 h、48 h、5 d、7 d、9 d、11d, respectively. The tissues were embedded in paraffin, and then the tissue sections were stained with immunohistochemistry technique. The results indicated that the SS DNA vaccine expressions were mainly distributed in muscle tissue neutrophils, macrophages, fibroblasts, in the spleen macrophages, in the liver cells. However, little was detected in heart, lung, kidney.
2 Descents and ascent dynamics of SS DNA vaccine expression products in mice tissues
We analyzed descents and ascent dynamics of SS DNA vaccine in mice tissues with the result of slice in different experimental time. The results show that the product of SS DNA vaccine was detected. The salmonella were attacked in fibroblasts cytoplasm after 24h of injecting. Immunologic reaction rose peak after 48h and SS positive signal was lost at the 5th day. From 7 d to 11 d in the immune, the expression products of vaccine can be detected in macrophages.
48h to 11 d after immunization, the expression pruducts of vaccine in the spleen of the cytoplasm of macrophages were observed in the microscope. Similarly, after the 48 h immunization, the expression pruducts of vaccine in the cytoplasm of liver cells were observed in the microscope, but only until 7 d.
3 Somatostatin DNA vaccine impact mice tissue
We observed and recoreded the whole health of the test mice, and analyzed the results of slice in the different time with hematoxylin-eosin staining in order to analyze live attenuated Salmonella choleraesuis in SS DNA vaccine have toxic effects in mice viable growth and development. The result showed that there was no one dead in the experiment and live attenuated Salmonella choleraesuis caused certain pathological damage on liver, but the first 7 days after immunization in mice liver can damage tissue to repair it and to maintain normal liver function, In this regard, the vaccine is safe. In addition, the vaccine did not have other pathological tissue damage detection.
引文
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