糖尿病大鼠创面感染的铜绿假单胞菌与自噬相互作用的实验研究
发布时间：2018-07-24 09:22
【摘要】：目的:1、探讨PA的T3SS结构对糖尿病溃疡创面组织中自噬相关蛋白表达的影响,并进一步探讨其对糖尿病溃疡创面愈合的影响。2、探讨PA的T3SS分泌的毒力蛋白ExoU、ExoS分别对糖尿病溃疡创面组织中自噬相关蛋白表达的影响,并进一步探讨其对糖尿病溃疡创面愈合的影响。3、探讨雷帕霉素及庆大霉素对PA感染的糖尿病溃疡创面组织中自噬相关蛋白表达的影响及对糖尿病溃疡创面愈合的影响。方法:一、第一部分:36只SD大鼠随机分为六组:糖尿病非感染组(DM+未感染),糖尿病感染PAK组(DM+PAK),糖尿病感染PAK△pcrV-(DM+PAK△pcrV-),非糖尿病非感染组(N+未感染),非糖尿病感染PAK组(N+PAK),非糖尿病感染PAK△pcrV-(N+PAK△pcrV-),每组各6只。分别在造模第0天、3天、7天、14天、21天观察创面愈合情况并进行创面内菌落计数;分别在造模第0、7、14天取创面组织,HE染色观察各组大鼠创面组织内细胞形态、结构变化;免疫组化及Western blotting检测各组大鼠创面组织内自噬相关蛋白LC3、Beclin-1、p62的表达;免疫荧光双染色检测各组大鼠创面组织内巨噬细胞中LC3的表达;二、第二部分:24只SD大鼠随机分为四组:糖尿病感染PA103组(DM+PA103),糖尿病感染PA01(DM+PA01),非糖尿病感染PA103组(N+PA103),非糖尿病感染PA01(N+PA01),每组各6只。方法同第一部分;三、第三部分:36只SD大鼠随机分为六组:糖尿病感染PA103组(DM+PA103),糖尿病感染PA103且雷帕霉素干预组(DM+PA103+雷帕霉素),非糖尿病感染PA103组(N+PA103),非糖尿病感染PA103且雷帕霉素干预组(N+PA103+雷帕霉素),糖尿病感染PA103且庆大霉素干预组(DM+PA103+庆大霉素),非糖尿病感染PA103且庆大霉素干预组(N+PA103+庆大霉素),每组各6只。方法同第一部分。结果:一、第一部分:1、创面愈合情况:与非糖尿病组大鼠相比,糖尿病组大鼠第3天有黑痂形成,第7天有较多脓性分泌物,创面未见缩小,第21天创面仍未愈合,伤愈合时间明显延迟。在糖尿病组中,与DM+未感染及DM+PAK△pcrV-相比,第3天DM+PAK组黑痂形成、坏死较严重、周围较红肿,第7天有脓性分泌物渗出且最多,第21天仍有较多分泌物,第21伤口直径较大。2、创面内菌落计数:与非糖尿病感染组相比,糖尿病感染组菌落增长速度较快且减少速度较慢,其中与DM+PAK△pcrV-相比,DM+PAK组第7天以后菌落数开始减少,但减少不明显。3、HE结果:糖尿病感染组在第7、14天炎性粒细胞浸润逐渐增加,与DM+PAK△pcrV-相比,DM+PAK分别在第7、14天有更多炎性细胞浸润。非糖尿病组感染组在第7天炎性细胞浸润达到高峰,第14天炎性细胞下降,与N+PAK△pcrV-组相比,N+PAK分别在第7、14天有更多炎性细胞浸润。4、免疫组化结果:各组LC3、Beclin-1在第0、7、14天表达递增,p62蛋白第7、14天表达均逐渐下降(P均0.05)。N+非感染组LC3、Beclin-1表达在第7、14天表达明显高于DM+非感染组,p62表达在第7、14天低于DM+非感染组(P均0.05)。DM+PAK△pcrV-组LC3、Beclin-1表达在第14天高于DM+PAK,p62表达在第14天低于DM+PAK(P均0.05);N+PAK△pcrV-组中LC3、Beclin-1表达在第7、14天高于N+PAK,p62表达在第7、14天低于N+PAK(P均0.05);与N+PAK组相比,DM+PAK组在第14天LC3、Beclin-1表达较低,p62表达较高(P均0.05);与N+PAK△pcrV-组相比,DM+PAK组LC3、Beclin-1表达在第7、14天较低,p62表达在第7、14天较高(P均0.05)。5、免疫印迹结果:同免疫组化结果相一致。6、免疫荧光结果:DM+PAK△pcrV-组巨噬细胞中LC3表达在第14天高于DM+PAK组;N+PAK△pcrV-组中巨噬细胞中LC3表达在第7、14天高于N+PAK。二、第二部分:1、创面愈合情况:与DM+PA01相比,DM+103组第3天有黑痂形成、坏死较严重、周围较红肿,伤口直径扩大,第14天伤口直径仍未缩小,坏死仍严重,第21天仍有较多分泌物,创面仍未愈合。2、创面内菌落计数:DM+PA103和DM+PA01在第0-7天持续增长,与DM+PA01相比,DM+PA103增加较快,第7天以后菌落数开始减少,减少较缓慢。3、HE结果:与DM+PA01组相比,DM+PA103分别在第7、14天有较多中性粒细胞、巨噬细胞浸润。4、免疫组化结果:各组LC3、Beclin-1在第0、7、14天表达递增,p62蛋白第7、14天表达均逐渐下降(P均0.05)。DM+PA103组LC3、Beclin-1表达在第14天低于DM+PA01,p62表达在第14天高于DM+PA01(P均0.05);N+PA103组LC3、Beclin-1表达在第7天低于N+PA01,p62表达在第7天高于N+PA01(P均0.05);与N+103组相比,DM+103组LC3、Beclin-1表达在第7、14天较低,p62表达在第7、14天较高(P均0.05)。5、免疫印迹结果:同免疫组化结果。6、免疫荧光结果:DM+103组巨噬细胞中LC3表达在第14天低于DM+PA01;N+PA01组巨噬细胞中LC3表达在第7天高于N+PA103。三、第三部分:1、创面愈合情况:与DM+PA103相比,DM+PA103+雷帕霉素在第7天脓性分泌物较前减少,第14天创面直径明显缩小。DM+PA103+庆大霉素组在第7天创面直径明显缩小,第21天伤口趋于愈合。与N+PA103相比,N+PA103+雷帕霉素在第3天组织破溃较轻,脓性分泌物渗出较少,第7天创面直径明显缩小,第21天创面基本愈合。N+PA103+庆大霉素组第3天结痂较轻,第21天创面基本愈合。2、创面内菌落计数:DM+PA103和DM+PA103+雷帕霉素在第0-7天持续增长,与DM+PA103组相比,DM+PA103+雷帕霉素增加较慢,第7天以后菌落数减少较快,DM+PA103+庆大霉素组第7天菌落数减少较快;与N+PA103组相比,N+PA103+雷帕霉素在第3天后菌落明显减少,N+PA103+庆大霉素第3天后菌落明显减少(P0.05)。3、HE结果:与DM+PA103组相比,DM+PA103+雷帕霉素与DM+PA103+庆大霉素组分别在第7、14天中性粒细胞、巨噬细胞浸润等炎症细胞数目较少。与N+PA103组相比,N+PA103+雷帕霉素与N+PA103+庆大霉素分别在第7、14天炎症细胞浸润减少。4、免疫组化结果:各组LC3、Beclin-1在第0、7、14天表达递增,p62蛋白第7、14天表达均逐渐下降(P均0.05)。DM+PA103+雷帕霉素组LC3表达在第7、14天高于DM+PA103,Beclin-1表达在第14天高于DM+PA103,p62表达在7、14低于N+PA103(P均0.05);N+PA103+雷帕霉素组LC3在第7、14天高于DM+PA103,Beclin-1表达在第7天高于N+PA103,p62表达在第7天低于N+PA103(P均0.05);与DM+PA103+雷帕霉素组相比,N+PA103+雷帕霉素组LC3、Beclin-1表达在第7、14天较高,p62表达在第7、14天较高(P均0.05)。DM+PA103+庆大霉素组和DM+PA103组在各个时间点Beclin-1、LC3、p62的表达量均无显著差异,N+PA103+庆大霉素组与N+PA103组在各个时间点Beclin-1、LC3、p62的表达量均无显著差异(P均0.05)。5、免疫印迹结果:同免疫组化。6、免疫荧光结果:DM+PA103+雷帕霉素组巨噬细胞中LC3表达在第7、14天高于DM+PA103;N+PA103+雷帕霉素组巨噬细胞中LC3在第7、14天高于N+PA103。结论:1、在创面愈合过程中,非糖尿病及糖尿病大鼠创面组织中细胞自噬蛋白水平均增强,但是糖尿病组的自噬水平明显低于非糖尿病组,且糖尿病创面炎症持续,创面愈合延迟。2、在糖尿病感染创面中,PA的T3SS均可抑制自噬蛋白水平的表达,自噬对细菌的清除能力减弱,PA的T3SS分泌的毒力蛋白ExoU抑制自噬蛋白水平的表达更明显,自噬对细菌的清除能力明显减弱,ExoU可导致糖尿病创面坏死更严重,感染持续且糖尿病创面愈合受损。3、在非糖尿病感染创面和糖尿病感染创面中,雷帕霉素可通过增强自噬促进对PA的清除可加速创面愈合,但其在糖尿病状态的效果弱于非糖尿病状态。4、庆大霉素在感染病创面可有效地清除PA以减少感染,进而促进创愈合,无论在糖尿病创面还是非糖尿创面,庆大霉素对创面内自噬蛋白水平无明显影响。
[Abstract]:Objective: 1, to investigate the effect of T3SS structure of PA on the expression of autophagy related protein in the tissue of diabetic ulcer wound, and further explore the effect of.2 on the healing of diabetic ulcer wound, and explore the effect of ExoS on the expression of autophagy related protein in the tissue of diabetic ulcer wound, and further explore the effect of ExoS on the expression of autophagic protein in the tissue of diabetic ulcer wound. Effect of.3, the effect of rapamycin and gentamicin on the expression of autophagy related protein in the tissue of diabetic ulcer wound with PA infection and the effect on the healing of diabetic ulcer wound. Methods: 1. Part one: 36 SD rats were randomly divided into six groups: diabetes non infected group (DM+ uninfected), diabetes sense PAK group (DM+PAK), diabetes infection PAK Delta pcrV- (DM+PAK Delta pcrV-), non diabetic non infection group (N+ uninfected), non diabetic PAK group (N+PAK), non diabetic PAK Delta pcrV- (N+PAK delta), each group of 6 rats, respectively in the model zeroth days, 3 days, 7 days, 14 days, 21 days to observe the wound healing and to carry on the count of bacterial colonies in the wound; respectively The wound tissue was taken on day 0,7,14, and the morphological and structural changes in the wound tissue were observed by HE staining. Immunohistochemistry and Western blotting were used to detect the expression of autophagy related protein LC3, Beclin-1, p62 in the wound tissue of each group, and the expression of LC3 in the macrophages in the wound tissue of the rats was detected by immunofluorescence; two The two part: 24 SD rats were randomly divided into four groups: diabetic PA103 group (DM+PA103), diabetes infected with PA01 (DM+PA01), non diabetic infection PA103 group (N+PA103), non diabetic infection PA01 (N+PA01), each group with the first part; three, third parts: 36 SD rats were randomly divided into six groups: diabetes PA103 group (DM+PA103), diabetes mellitus Disease infection PA103, rapamycin intervention group (DM+PA103+ rapamycin), non diabetic PA103 group (N+PA103), non diabetic PA103 and rapamycin intervention group (N+PA103+ rapamycin), diabetes infection PA103 and gentamicin intervention group (gentamicin), non diabetic infection PA103 and gentamicin intervention group (N+PA103+ celebration group) 6 in each group. Methods and the first part. Results: first, part one: 1, wound healing: compared with the non diabetic rats, the rats in the diabetic group had black eschar formation on third days, there were more purulent secretions on seventh days, the wound was not reduced, the wounds were still not healed on the twenty-first day, and the healing time was delayed. In the diabetic group, the DM+ was not felt with the diabetic group. Compared with DM+PAK Delta pcrV-, the third day group of DM+PAK group black scab formed, the necrosis was more serious, the surrounding was more red and swelling, the purulent secretions were exudative and most in the seventh day, there were more secretions in twenty-first days, the twenty-first wound diameter was larger than that of the non diabetic infection group, and the growth rate of the diabetic infection group was faster and less speed than that of the non diabetic infection group. As compared with DM+PAK Delta pcrV-, the colony number of the DM+PAK group began to decrease after seventh days, but the decrease was not obvious.3. The HE result: the inflammatory granulocyte infiltration in the diabetic infection group gradually increased on the 7,14 day, and the DM+PAK was more inflammatory cell infiltration in the 7,14 day, compared with DM+PAK Delta pcrV-. Up to the peak, the fourteenth days of inflammatory cells decreased. Compared with the N+PAK Delta pcrV- group, N+PAK had more inflammatory cells infiltrating.4 on day 7,14. The results of immunohistochemistry: LC3, Beclin-1 in 0,7,14 days were increased, and the expression of p62 protein in 7,14 day decreased gradually (P was 0.05). In the non infected DM+ group, the expression of p62 was lower than the DM+ non infection group (P 0.05).DM+PAK Delta pcrV- group LC3, the Beclin-1 expression was higher than DM+PAK in the fourteenth day, and the p62 expression was lower than DM+PAK (all 0.05) at fourteenth days. In group DM+PAK, the expression of Beclin-1 was lower at fourteenth days and the expression of p62 was higher (P 0.05). Compared with N+PAK Delta pcrV- group, DM+PAK group LC3, Beclin-1 expression was lower in 7,14 day, p62 expression was higher (0.05). Immunoblotting results: immunoblotting results: immunofluorescence results: expression of immunofluorescence On the fourteenth day higher than the DM+PAK group, the expression of LC3 in the macrophages in the N+PAK Delta pcrV- group was higher than N+PAK. two, the second part: 1, the wound healing. Compared with DM+PA01, the DM+103 group had black eschar formation on third days, the necrosis was more severe, the diameter of the wound was more red, the diameter of the wound was enlarged, the diameter of the wound was still not narrowed, the necrosis was still serious and twenty-first days was still more serious. .2 was still not healed in the wound, and the colony count in the wound was not healed. DM+PA103 and DM+PA01 continued to increase on day 0-7. Compared with DM+PA01, DM+PA103 increased rapidly. After seventh days the colony number began to decrease, and the slow.3, HE result: DM+PA103 was more neutrophils in 7,14 day than DM+PA01 group, and macrophage infiltration was.4, immunization. The expression of LC3 and Beclin-1 increased gradually on day 0,7,14. The expression of p62 protein on day 7,14 decreased gradually (P 0.05).DM+PA103 group LC3, Beclin-1 expression was lower than DM+PA01 in the fourteenth day, p62 expression was higher in Fourteenth days than DM+PA01 (0.05). Compared with the N+103 group, the expression of LC3 and Beclin-1 in group DM+103 was lower on day 7,14, p62 expression was higher on day 7,14 (P 0.05).5, and immunoblotting results: the result of immunoblotting.6, immunofluorescence results: LC3 expression in the DM+103 group macrophages was lower than three, third part of macrophage in seventh days. 1, wound healing: compared with DM+PA103, the seventh days of rapamycin was reduced in seventh days, the diameter of the wound was significantly reduced in Fourteenth days and the diameter of.DM+PA103+ gentamicin group narrowed at seventh days, and the wounds tended to heal in twenty-first days. Compared with N+PA103, N+ PA103+ rapamycin had a lighter tissue break and a purulent secretion in the third day. The exudation was less, the diameter of the wound was obviously reduced in seventh days, the basic healing of the wound was twenty-first days in the group of.N+PA103+ gentamicin group, and the wound healing was lighter in third days. The wound healing was basically.2 in the twenty-first day. The number of DM+PA103 and DM+PA103+ rapamycin continued to increase on the 0-7 day, compared with the DM+PA103 group, the increase of DM+PA103+ rapamycin was slower, and seventh days later, the bacteria were increased. The colony number of DM+PA103+ gentamicin group decreased faster in the seventh day of gentamicin group. Compared with group N+PA103, the colony of N+PA103+ rapamycin decreased obviously after third days, and the colony of N+PA103+ gentamicin decreased significantly (P0.05).3, HE result: DM+ PA103+ rapamycin and DM+PA103+ gentamicin group were in the first 7,14 days, respectively. The number of inflammatory cells in neutrophils and macrophage infiltration was less. Compared with the N+PA103 group, the infiltration of N+PA103+ rapamycin and N+PA103+ gentamicin decreased.4 respectively on day 7,14, and the immunohistochemical results showed that the expression of LC3, Beclin-1 in 0,7,14 days increased, and the expression of p62 egg white in 7,14 day decreased gradually (P all 0.05).DM+PA103+ Lei The expression of LC3 in the paramiomycin group was higher in the day 7,14 than in DM+PA103, and the expression of Beclin-1 was higher in Fourteenth days than in DM+PA103, and the expression of p62 in 7,14 was lower than N+PA103 (P 0.05); N+PA103+ rapamycin group was higher in 7,14 day than in seventh days. The expression of LC3 and Beclin-1 in +PA103+ rapamycin group was higher on 7,14 day, p62 expression was higher on 7,14 day (P all 0.05).DM+PA103+ gentamicin group and DM+PA103 group at all time points Beclin-1, LC3, p62 expression was not significant difference. Difference (P 0.05).5, immunoblotting results: immunofluorescence.6, immunofluorescence results: the expression of LC3 in macrophages in DM+PA103+ rapamycin group was higher in 7,14 day than DM+PA103; LC3 in N+PA103+ of rapamycin group was higher in N+PA103. conclusion on 7,14 day: 1, in wound healing process, the wound tissue of non diabetic and diabetic rats was fine. The level of autophagic protein was enhanced, but the level of autophagy in the diabetic group was significantly lower than that in the non diabetic group, and the inflammation of the diabetic wound continued and the wound healing delayed.2. In the diabetic infection wound, PA T3SS could inhibit the expression of autophagic protein, the ability to remove autophagy to bacteria and the inhibitory protein ExoU secreted by PA T3SS. The expression of autophagic protein is more obvious, the ability of autophagy to scavenge bacteria is significantly weakened, ExoU can lead to more severe diabetic wound necrosis, continuous infection and impaired healing of diabetic wound.3. In non diabetic wound and diabetic infection wounds, rapamycin can accelerate wound healing by enhancing autophagy to promote the clearance of PA. However, its effect on diabetes is weaker than non diabetic.4. Gentamicin can effectively remove PA to reduce infection and thus promote healing. Gentamicin has no significant effect on the level of autophagic protein in wound and non diabetic wound.
【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R587.2
本文编号：2140925
[Abstract]:Objective: 1, to investigate the effect of T3SS structure of PA on the expression of autophagy related protein in the tissue of diabetic ulcer wound, and further explore the effect of.2 on the healing of diabetic ulcer wound, and explore the effect of ExoS on the expression of autophagy related protein in the tissue of diabetic ulcer wound, and further explore the effect of ExoS on the expression of autophagic protein in the tissue of diabetic ulcer wound. Effect of.3, the effect of rapamycin and gentamicin on the expression of autophagy related protein in the tissue of diabetic ulcer wound with PA infection and the effect on the healing of diabetic ulcer wound. Methods: 1. Part one: 36 SD rats were randomly divided into six groups: diabetes non infected group (DM+ uninfected), diabetes sense PAK group (DM+PAK), diabetes infection PAK Delta pcrV- (DM+PAK Delta pcrV-), non diabetic non infection group (N+ uninfected), non diabetic PAK group (N+PAK), non diabetic PAK Delta pcrV- (N+PAK delta), each group of 6 rats, respectively in the model zeroth days, 3 days, 7 days, 14 days, 21 days to observe the wound healing and to carry on the count of bacterial colonies in the wound; respectively The wound tissue was taken on day 0,7,14, and the morphological and structural changes in the wound tissue were observed by HE staining. Immunohistochemistry and Western blotting were used to detect the expression of autophagy related protein LC3, Beclin-1, p62 in the wound tissue of each group, and the expression of LC3 in the macrophages in the wound tissue of the rats was detected by immunofluorescence; two The two part: 24 SD rats were randomly divided into four groups: diabetic PA103 group (DM+PA103), diabetes infected with PA01 (DM+PA01), non diabetic infection PA103 group (N+PA103), non diabetic infection PA01 (N+PA01), each group with the first part; three, third parts: 36 SD rats were randomly divided into six groups: diabetes PA103 group (DM+PA103), diabetes mellitus Disease infection PA103, rapamycin intervention group (DM+PA103+ rapamycin), non diabetic PA103 group (N+PA103), non diabetic PA103 and rapamycin intervention group (N+PA103+ rapamycin), diabetes infection PA103 and gentamicin intervention group (gentamicin), non diabetic infection PA103 and gentamicin intervention group (N+PA103+ celebration group) 6 in each group. Methods and the first part. Results: first, part one: 1, wound healing: compared with the non diabetic rats, the rats in the diabetic group had black eschar formation on third days, there were more purulent secretions on seventh days, the wound was not reduced, the wounds were still not healed on the twenty-first day, and the healing time was delayed. In the diabetic group, the DM+ was not felt with the diabetic group. Compared with DM+PAK Delta pcrV-, the third day group of DM+PAK group black scab formed, the necrosis was more serious, the surrounding was more red and swelling, the purulent secretions were exudative and most in the seventh day, there were more secretions in twenty-first days, the twenty-first wound diameter was larger than that of the non diabetic infection group, and the growth rate of the diabetic infection group was faster and less speed than that of the non diabetic infection group. As compared with DM+PAK Delta pcrV-, the colony number of the DM+PAK group began to decrease after seventh days, but the decrease was not obvious.3. The HE result: the inflammatory granulocyte infiltration in the diabetic infection group gradually increased on the 7,14 day, and the DM+PAK was more inflammatory cell infiltration in the 7,14 day, compared with DM+PAK Delta pcrV-. Up to the peak, the fourteenth days of inflammatory cells decreased. Compared with the N+PAK Delta pcrV- group, N+PAK had more inflammatory cells infiltrating.4 on day 7,14. The results of immunohistochemistry: LC3, Beclin-1 in 0,7,14 days were increased, and the expression of p62 protein in 7,14 day decreased gradually (P was 0.05). In the non infected DM+ group, the expression of p62 was lower than the DM+ non infection group (P 0.05).DM+PAK Delta pcrV- group LC3, the Beclin-1 expression was higher than DM+PAK in the fourteenth day, and the p62 expression was lower than DM+PAK (all 0.05) at fourteenth days. In group DM+PAK, the expression of Beclin-1 was lower at fourteenth days and the expression of p62 was higher (P 0.05). Compared with N+PAK Delta pcrV- group, DM+PAK group LC3, Beclin-1 expression was lower in 7,14 day, p62 expression was higher (0.05). Immunoblotting results: immunoblotting results: immunofluorescence results: expression of immunofluorescence On the fourteenth day higher than the DM+PAK group, the expression of LC3 in the macrophages in the N+PAK Delta pcrV- group was higher than N+PAK. two, the second part: 1, the wound healing. Compared with DM+PA01, the DM+103 group had black eschar formation on third days, the necrosis was more severe, the diameter of the wound was more red, the diameter of the wound was enlarged, the diameter of the wound was still not narrowed, the necrosis was still serious and twenty-first days was still more serious. .2 was still not healed in the wound, and the colony count in the wound was not healed. DM+PA103 and DM+PA01 continued to increase on day 0-7. Compared with DM+PA01, DM+PA103 increased rapidly. After seventh days the colony number began to decrease, and the slow.3, HE result: DM+PA103 was more neutrophils in 7,14 day than DM+PA01 group, and macrophage infiltration was.4, immunization. The expression of LC3 and Beclin-1 increased gradually on day 0,7,14. The expression of p62 protein on day 7,14 decreased gradually (P 0.05).DM+PA103 group LC3, Beclin-1 expression was lower than DM+PA01 in the fourteenth day, p62 expression was higher in Fourteenth days than DM+PA01 (0.05). Compared with the N+103 group, the expression of LC3 and Beclin-1 in group DM+103 was lower on day 7,14, p62 expression was higher on day 7,14 (P 0.05).5, and immunoblotting results: the result of immunoblotting.6, immunofluorescence results: LC3 expression in the DM+103 group macrophages was lower than three, third part of macrophage in seventh days. 1, wound healing: compared with DM+PA103, the seventh days of rapamycin was reduced in seventh days, the diameter of the wound was significantly reduced in Fourteenth days and the diameter of.DM+PA103+ gentamicin group narrowed at seventh days, and the wounds tended to heal in twenty-first days. Compared with N+PA103, N+ PA103+ rapamycin had a lighter tissue break and a purulent secretion in the third day. The exudation was less, the diameter of the wound was obviously reduced in seventh days, the basic healing of the wound was twenty-first days in the group of.N+PA103+ gentamicin group, and the wound healing was lighter in third days. The wound healing was basically.2 in the twenty-first day. The number of DM+PA103 and DM+PA103+ rapamycin continued to increase on the 0-7 day, compared with the DM+PA103 group, the increase of DM+PA103+ rapamycin was slower, and seventh days later, the bacteria were increased. The colony number of DM+PA103+ gentamicin group decreased faster in the seventh day of gentamicin group. Compared with group N+PA103, the colony of N+PA103+ rapamycin decreased obviously after third days, and the colony of N+PA103+ gentamicin decreased significantly (P0.05).3, HE result: DM+ PA103+ rapamycin and DM+PA103+ gentamicin group were in the first 7,14 days, respectively. The number of inflammatory cells in neutrophils and macrophage infiltration was less. Compared with the N+PA103 group, the infiltration of N+PA103+ rapamycin and N+PA103+ gentamicin decreased.4 respectively on day 7,14, and the immunohistochemical results showed that the expression of LC3, Beclin-1 in 0,7,14 days increased, and the expression of p62 egg white in 7,14 day decreased gradually (P all 0.05).DM+PA103+ Lei The expression of LC3 in the paramiomycin group was higher in the day 7,14 than in DM+PA103, and the expression of Beclin-1 was higher in Fourteenth days than in DM+PA103, and the expression of p62 in 7,14 was lower than N+PA103 (P 0.05); N+PA103+ rapamycin group was higher in 7,14 day than in seventh days. The expression of LC3 and Beclin-1 in +PA103+ rapamycin group was higher on 7,14 day, p62 expression was higher on 7,14 day (P all 0.05).DM+PA103+ gentamicin group and DM+PA103 group at all time points Beclin-1, LC3, p62 expression was not significant difference. Difference (P 0.05).5, immunoblotting results: immunofluorescence.6, immunofluorescence results: the expression of LC3 in macrophages in DM+PA103+ rapamycin group was higher in 7,14 day than DM+PA103; LC3 in N+PA103+ of rapamycin group was higher in N+PA103. conclusion on 7,14 day: 1, in wound healing process, the wound tissue of non diabetic and diabetic rats was fine. The level of autophagic protein was enhanced, but the level of autophagy in the diabetic group was significantly lower than that in the non diabetic group, and the inflammation of the diabetic wound continued and the wound healing delayed.2. In the diabetic infection wound, PA T3SS could inhibit the expression of autophagic protein, the ability to remove autophagy to bacteria and the inhibitory protein ExoU secreted by PA T3SS. The expression of autophagic protein is more obvious, the ability of autophagy to scavenge bacteria is significantly weakened, ExoU can lead to more severe diabetic wound necrosis, continuous infection and impaired healing of diabetic wound.3. In non diabetic wound and diabetic infection wounds, rapamycin can accelerate wound healing by enhancing autophagy to promote the clearance of PA. However, its effect on diabetes is weaker than non diabetic.4. Gentamicin can effectively remove PA to reduce infection and thus promote healing. Gentamicin has no significant effect on the level of autophagic protein in wound and non diabetic wound.
【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R587.2
【参考文献】
相关期刊论文 前5条
1 王宁;左国营;谢俊杰;程子梦;;铜绿假单胞菌Ⅲ型分泌系统与致病毒力因子的研究进展[J];解放军药学学报;2015年05期
2 周晓娟;徐根隆;应华永;;烧伤科耐药铜绿假单胞菌耐药性与毒力基因研究[J];中华医院感染学杂志;2014年07期
3 吕雷;林康;高伟阳;何智灵;高自勉;李浙峰;李俊杰;;饥饿诱导增生性瘢痕成纤维细胞自噬发生[J];中国病理生理杂志;2013年02期
4 董晨晓;宋诗铎;王悦;门昆;;43株临床铜绿假单胞菌exoS、exoU基因的携带及其耐药性[J];中国感染控制杂志;2010年02期
5 朱欣;李闻文;;糖尿病足感染的病原菌特点及耐药性分析研究[J];实用预防医学;2007年04期
相关博士学位论文 前1条
1 宣敏;自噬参与衰老皮肤创面愈合的相关实验研究[D];南方医科大学;2014年
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