【作者】 周智；

【导师】 李荟元；

【作者基本信息】 中国人民解放军第四军医大学 ， 外科学， 2003， 博士

【摘要】 瘢痕是创伤修复的必然产物，是由肉芽组织逐渐发展和演变而来的，主要由胶原纤维组成的血管稀少的具有透明变性改变的纤维组织，瘢痕从形态和转归方面可以分为普通瘢痕和病理性瘢痕，病理性瘢痕又分为增生性瘢痕(Hypertrophic scar HS)和瘢痕疙瘩(Keloid K)两种。而瘢痕根据其发展时间和自身发展的特点，我们又可以分为生长活跃的瘢痕和相对静息期的瘢痕，相对静息期的瘢痕中，血管常常硬化闭锁，小血管、皮肤附件和成纤维细胞的结构大多消失，瘢痕主要由透明变性的胶原纤维组成。对于瘢痕的发生和演变过程和原因目前尚没有一个完整的解释，而瘢痕之间又存在着不同的生长转归特性，研究不同瘢痕之间的增殖和转化规律就成为研究瘢痕本身特质的方向。 以往的研究把目光大多投到了瘢痕的增生阶段，这个阶段瘢痕中细胞数量很多、细胞功能活跃、相关影响因素复杂，既为研究提供了机遇，但同时也增加了研究的难度和成本。在实践中我们注意 第四军医大学博士学位论文到，并不是所有的静息期疲痕都是稳定的和一成不变的，在一定的刺激因素下他们仍然会复发和重新拥有增殖和分泌的活性。对于静息期癫痕的研究国内目前尚没有报道，国外也罕见文献，我们研究相对静息期病理性癫痕，就是试图找出在癫痕相对稳定的时期本身所具有的增殖和凋亡潜质，这对于揭示病理性疲痕的性质是一个新的研究思路和提示。 由于成熟的癫痕的固有特征，在这个阶段，瘤痕相对稳定、细胞数目少、增殖等生理活性降低，基本上由胶原纤维组成。在研究中我们更多的使用了免疫组化检测抗原表达的方式，通过对静息期病理性瘫痕中影响增殖和凋亡的相关抗原表达的检测和分析，我们得到了一些有益的结果。 一、我们通过常规病理染色在镜下确定了研究的对象，将实验的标本定位在处于相对静息期的病理性瘟痕上，并确定了可能影响静息期瘤痕转归的因素，即疲痕中残存的血管及皮肤附件周围具有增殖活性的成纤维细胞。为继续的研究提供了良好的思路和方向。同时发现HS和K有着不同的镜下胶原纤维的形态特征，即：HS大多为松散的，旋涡状、结节状结构。而K 中，为相对平行的，均质样结构。两者均表现出明显的玻璃样变性的特点。 4 第四军医大学博士学位论文 二、检测了凋亡相关基因仇 1－2、Bax在相对静息期 HS、K中的表达及分布。通过实验发现：HS和 K的成纤维细胞中他们的表达强度及分布有所不同，在 HS中，BC－2主要分布在瘫痕上皮基底层，真皮浅层也有少量散在表达分布，但是表达较弱；BSX表达阳性率及强度较 k－2 高，多在上皮基底层及小血管的附近、皮肤附件周围的成纤维细胞中出现，镜下可见的少量的游离成纤维细胞上也有表达；而在K中，BCI－2的表达也主要在瘫痕上皮的棘细胞层和基底细胞层，在血管周围的成纤维细胞内也有少量表达；中央区罕见的成纤维细胞中也能看到Bcl－2着色。同时Bax的表达较k－2的弱，而且多发生在基底细胞层。观察中，有两个K标本在瘤痕深处发现了有凋亡抗原仇－2的染色（弱表达）。我们认为凋亡基因在癫痕表面上皮、微小血管周围、皮肤残留附件上的表达，提示着我们，它们在相对静息期疲痕未来的转归中可能扮演着重要的角色。 三、我们检测了相对静息期的HS、K中IV型胶原的分布及表达，这在国内尚没有报道，结合文献我们认为IV型胶原的分布及表达对于癫痕的转归应该有着重要的影响。实验中我们发现：在所有的病理性疲痕标本中均有IV型胶原的表达。在HS中血管壁中可看到深褐色的较为粗大的颗粒样物质存在，而同样在K 中的表达无论色彩深 5 第四军医大学博士学位论文度及浓密度都远不如 HS。在对皮肤残留附件、癫痕表皮组织中的表达的观察中我们也得到了与在血管壁上观察到的几乎相似的结论。结合文献，我们认为在静息期的疲痕中，IV型胶原也是诱发疲痕转归的重要的因素，但是它本身的转归和以及在活跃期痰痕中的作用还有待进一步研究。同时我们结合所观察到的结果并结合文献提出了以下的假设：①IV型胶原或许是在成熟期决定瘫痕走向及增生性瘫痕消退的重要因素；②IV型胶原对于成纤维细胞分泌的I、Ill型胶原的比例及转归可能有一定的影响作用；③w型胶原的高表达对于上皮细胞的反馈作用也应该对上皮细胞，并通过上皮细胞对疲痕本身的转归产生作用；④IV型胶原可能是未来诱发痰痕凋亡转归的诱发因素。 四、我们进行了相对静息期HS爪核仁形成区嗜银染色ug－NORS）及增殖细胞核抗原（PCNA）的检测，发现在相对静息期的HS 中AS－NORs染色阳性区主要集中在小血管和附件周围的细胞区域，在分布区域上K与HS相同，两者在表达强度上无显著差异。PCNA检测中HS和K的抗原表达在分布区域上没更多还原

【Abstract】 Scar is the inevitable production of tissue repair, and it is gradually developed from granulation tissue. Scar is mainly made up of fiber tissue with a few blood vessels collagenous fibers undergo hyaline degeneration. According to its morphologic change, scar can be divided into two groups --- common scar and pathologic scar. And pathologic scar mainly includes hypertrophic scar and keloid. According to its development, scar can be defined as actively growing scar and relatively resting scar . The major features of actively growing scar are: the high multiplication rate of fibroblast and the high apoptosis rate, and strong collagen secretion. And there are many blood vessels vertical to epithelium in the actively growing scar. While in the relatively resting scar, blood vessels are generally sclerosis and blocked and small blood vessels, appendages of skin and structure of fibroblast mainly disappear. And the relatively resting scar is mainly made up of the collagenous fibers undergo hyaline degeneration. By far, there is any perfect explanation of the development of scar, and since among scars there are different development and transformation, the study on the multiplication and transformation among scars becomes a way to the study on the features of scars.In the past, the study on scars focused on the following aspects: 1. growing, multiplying and apoptosis characters of fibroblast in or fromscar and relation with ECM. 2. the development and transformation of ECM and relation with other related factors. 3. signal transfer passage of related cells and effects on induction, occurrence and development of different scars on genetic level.Previous study emphasizes the multiplying period of scar, in which the cellular number in scar is large, cells are active and the related factors are complex. All these not only provide opportunity but increase difficulty and cost of research as well. In practice, we notice that not all relatively resting scars are stable and unchangeable. By certain stimulating factors, they will still regain the activity of multiplication and secretion. So far there is no report on the research of the relatively resting scar in China, and rare reports abroad. Through the study on the relatively resting pathologic scars, we attempt to find their potentials to multiply and apoptosis in the relatively resting period and this is a new way to show the features of pathological scar.Owing to the features of relatively resting scars, in this period scars mainly made up of collagenous fibers are relatively stable, the number of cells is small, the biological activity such as multiplication decreases. As to their research, we conducted immunohistochemical determination of antigen to express antigen. We gained some results by the different expression of antigens influencing multiplication and apoptosis of relatively resting pathologic scars.1. We determined the research object under the microscope by regular pathological staining, and chose the pathologically relatively resting scars as the experimental samples and determined the possible factors affecting the transformation of relatively resting scars,namely the small blood vessels remaining in the skin and fibroblasts with multiplying activity around appendages of skin. At the same time, we found hyperplastic scar and keloid had the different morphologic features of collagenous fibers under microscope, that is hyperplastic scar always has loose, whirl-like and nodular structure, while keloid has relatively parallel and homogeneous structure. Hyaline degeneration significantly characterizes hyperplastic scar and keloid.2. We determined the expression distribution of genes Bcl-2 and Bax related to apoptosis in HS in relatively resting period and K. Through the analysis of the expression of different apoptosis antigens, we hoped to find its cellular elements and its potentials. We found the expression intensity and distribution of fibroblast of hyperplastic scar and keloid were different. In hyperplastic scar, expression of Bcl更多还原