人脑挫伤后HSP70、bFGF和COX2免疫组织化学研究
发布时间：2019-05-23 07:26
【摘要】： 前言 随着医学界对中枢神经系统创伤后分子病理学认识的提高以及免疫组织化学技术的进步，人们从免疫组织化学角度，对脑挫伤与时间的关系做了大量的研究。由于受到检材要求所限，在此方面的研究多为模拟脑挫伤的动物实验，从而造成了理论研究与实际鉴定工作相“脱钩”的现象。 目前，已有人分别应用免疫组织化学技术对热休克蛋白70(HSP70)、碱性成纤维细胞生长因子(bFGF)和环氧合酶-2(COX2)在大鼠脑挫伤后的表达进行了研究，结果表明这三项指标的表达在伤后均呈现出一定的变化规律，但未见有人将HSP70、bFGF和COX2应用于人脑挫伤时间推断的研究。故本实验采用实际工作中的案例标本，应用免疫组织化学染色，，旨在了解人脑挫伤后HSP70、bFGF和COX2的表达，揭示其表达与人脑挫伤时间变化关系，同时观察尸体解剖标本与动物实验标本免疫组织化学的差别，为脑挫伤时间判定实际鉴定工作提供一条可行的理论依据。 实验材料与方法 1．实验分组。选择1995-2001年因闭合性颅脑损伤死亡并伴有明显大脑皮质挫伤灶的35个案例，分成7个实验组，对照组选取非颅脑损伤且急性损伤致死5个案例，各组男女不限，年龄在19-58岁之间，平均年龄为38.09岁。 2．免疫组织化学染色方法。石蜡切片常规脱蜡至水；3％H_2O_2灭活内源性过氧化物酶；PBS冲洗；抗原修复；PBS冲洗；正常血清封闭；一抗，37℃1h；PBS冲洗；二抗，37℃30min；PBS冲洗；滴加 SP（或 SABC），37℃30min；PBS冲洗；DAB显色后水洗；苏木素复 染，脱水、透明、封片。 3．图像分析与统计学处理。应用MOTICAM 1300图像分析 仪，对免疫组织化学染色呈阳性的细胞定量检测其灰度值，然后在 SPSS 10．0 for Window软件下进行统计学处理。 实验结果 ＊结构型HSP70免疫组织化学染色。即刻死亡组HSpoo阳 性神经细胞灰度值最小，与其它各实验组相比有显著差异（P＜ 0．01人随挫伤时间的延长，神经细胞的 HSP70灰度值逐渐增大， 至20－27h组，HSpoo灰度值达最大，与其它各实验组（除10．5－ 11．sd组）相比有显著差异（P＜O．01人 以后各实验组mn0 灰度值有所下降，10．5刁1．sd组HSP70灰度值又再次升高。 2．bFGF免疫组织化学染色。即刻死亡组hFGF阳性神经细 胞灰度值最小，与其它各实验组相比有显著差异（P＜O．01人 10－14h组bFGF灰度值急剧增力（P＜0．0），以后bFGF灰度 值缓慢升高，至 10．5叫．sd组，灰度值达最大。 3．COXZ免疫组织化学染色。10－1均组 COXZ阳性神经细 胞的灰度值开始显著不久p＜｛一n，至排一月h组COXZ灰度值 达最｛ 乡与其｛ 各实验主卜以一入了显名差异（ ＜O．OU哆＊ 一＊卜组 和 2．5－3．sd组阳性全月再度值开步 升高，6。5－7．sd组阳性细 胞灰度值达最大，与其他各实验组相比有显著差异冲＜O．01厂 10．5－11．sd组未见 COXZ 的表达。 讨 论 1．HSpoo免疫组织化学染色结果分析。Seidbers等应用 Western印迹发现8例脑外伤手术患者的受损脑组织的结构型 HSP70与对照组相比没有差异。本实验检测人脑挫伤后结构型 ·2· HSpoo的表达情况，结果显示，各实验组中，即刻死亡组神经细胞 HSP70表达最强，HSpoo表达在损伤后 id显著下降，以后随着脑 挫伤时间的延长而有所恢复，这与Seidbers的实验结果存在较大 差异，原因是本实验案例的颅脑损伤程度较Seidbers的实验对象 更为严重，同时所采用的实验方法也不相同，所以，实验结果存在 较大的差异。因此，可以用结构型HSP70免疫组织化学染色推断 人脑挫伤时间。 2．bFGF免疫组织化学染色结果分析。Finklestein等发现大 鼠脑损伤后五周，bFGF免疫活性显著增加，其阳性细胞大多数类 似于反应性星形胶质细胞。Frantschy等检测大鼠中枢神经系统 损伤后bFGF蛋白表达情况，发现在损伤灶周边区的早期反应kh 到7d）中增加的bFGF表达大多数存在于MAC阳性细胞（巨噬细 胞和／或胶质细胞）中。本实验即刻死亡组的阳性神经细胞bFGF 表达最强，伤后lid内随时间的增加其表达不断下降。本实验 bFGF表达与动物实验差异的形成原因可能有以下几个方面：①实 验对象的差异。②颅脑损伤程度不一致。③由于脑损伤后，挫伤 周边区反应性缺血、缺氧，细胞反应性减低，从而抑制蛋白质的合 成。因此，可以用bFGF免疫组织化学染色推断人脑挫伤时间。 3．COXZ免疫组织化学染色结果分析。Str3SSS等人在大鼠外 伤性脑损伤后COXZ表达研究中发现，神经细胞COXZ免疫活性 于伤后Zh在4h和3d时升高，其中在24h显著增加，于伤后7d返 回至对照组水平。本实验检测人脑挫伤后COXZ的表达情况，结 果发现，重度颅脑损伤后 COXZ表达开始升高，在伤后 id左右达 峰值，以后逐步下降，其表达变化规律同Str8SSS等人的实验结果 基本一致。但本实验发现2．5－3．sd组和 6．5－7．sd组 COXZ表 达迅速下降，显著低于对照组，伤后 11d，COXZ表达消失，造成此 种现象的
[Abstract]:foreword With the improvement of the molecular pathology and the progress of the immunohistochemical technique in the treatment of the central nervous system after the trauma of the central nervous system in the medical community, it is known from the angle of the immunohistochemistry, the brain contusion and the time. The relationship is a lot of research. Because of the limited requirements of the material, the research on this aspect is mostly the animal experiment to simulate the contusion of the brain, thus causing the theoretical research and the practical appraisal work. At present, the heat shock protein 70 (HSP70), the basic fibroblast growth factor (bFGF) and the cyclooxygenase-2 (C) The expression of OX2 in the rat brain contusion was studied. The results showed that the expression of these three indexes showed a certain change in the postburn, but the expression of HSP70 and bFGF was not found. And the expression of HSP70, bFGF and COX2 after the contusion of the human brain is known, the relationship between the expression of the expression and the time of the human brain contusion is disclosed, and the body solution is observed at the same time. The difference of the immunohistochemistry between the sample and the experimental specimen of the animal is the time of the brain contusion. Determine the actual qualification to provide a viable option The experimental materials and methods 1. The experimental materials and methods 1. The experimental group was selected. In 1995-2001,35 cases of closed brain injury and with significant cerebral cortex contusion were selected and divided into 7 experimental groups, and the control group was selected from the group of non-craniocerebral injury and acute cerebral cortex contusion. 5 cases of injury, male and female in each group The age ranged from 19 to 58 years with a mean age of 38.09 years.2. Immunohistochemistry staining. The paraffin sections were routinely dewaxed to water;3% H _ 2O _ 2 inactivates the endogenous peroxidase; PBS rinse; Antigens complex; PBS wash; normal serum closure;1-,37 & deg; C for 1 h; PBS wash;2-,3 7 鈩
本文编号：2483717
[Abstract]:foreword With the improvement of the molecular pathology and the progress of the immunohistochemical technique in the treatment of the central nervous system after the trauma of the central nervous system in the medical community, it is known from the angle of the immunohistochemistry, the brain contusion and the time. The relationship is a lot of research. Because of the limited requirements of the material, the research on this aspect is mostly the animal experiment to simulate the contusion of the brain, thus causing the theoretical research and the practical appraisal work. At present, the heat shock protein 70 (HSP70), the basic fibroblast growth factor (bFGF) and the cyclooxygenase-2 (C) The expression of OX2 in the rat brain contusion was studied. The results showed that the expression of these three indexes showed a certain change in the postburn, but the expression of HSP70 and bFGF was not found. And the expression of HSP70, bFGF and COX2 after the contusion of the human brain is known, the relationship between the expression of the expression and the time of the human brain contusion is disclosed, and the body solution is observed at the same time. The difference of the immunohistochemistry between the sample and the experimental specimen of the animal is the time of the brain contusion. Determine the actual qualification to provide a viable option The experimental materials and methods 1. The experimental materials and methods 1. The experimental group was selected. In 1995-2001,35 cases of closed brain injury and with significant cerebral cortex contusion were selected and divided into 7 experimental groups, and the control group was selected from the group of non-craniocerebral injury and acute cerebral cortex contusion. 5 cases of injury, male and female in each group The age ranged from 19 to 58 years with a mean age of 38.09 years.2. Immunohistochemistry staining. The paraffin sections were routinely dewaxed to water;3% H _ 2O _ 2 inactivates the endogenous peroxidase; PBS rinse; Antigens complex; PBS wash; normal serum closure;1-,37 & deg; C for 1 h; PBS wash;2-,3 7 鈩
本文编号：2483717
本文链接：https://www.wllwen.com/shekelunwen/gongan/2483717.html
