层粘连蛋白和口腔癌关系的研究进展

肿瘤的侵袭 (invasion)和转移 (metastasis)是成功治疗肿瘤的最大障碍 ,上皮基底膜 (basement membrane,BM)的缺损是癌侵袭的标志。 BM是一复杂的细胞外基质结构 ,位于上皮 -基质界面 ,为维持实质细胞的结构提供了支持“骨架”(scaffold)。BM由层粘连蛋白 (laminin,L N) , 型胶原 ( collagen, -col)和硫酸肝素蛋白多糖 (haparansulfate proteoglycans,HS-PG)等胞外基质 (extracellular matrix,ECM)组成。电镜下 BM由致密板 (lamina densa)和透明板 (lamina lucida)构成 ,L N只分布于 BM的透明板中 ,由上皮细胞合成 ,并对上皮细胞…     

细胞角蛋白表达在诊断皮肤鳞状细胞癌和基底细胞癌中的应用

目的观察7种细胞角蛋白(Cytokeratin,CK)在皮肤鳞状细胞癌和基底细胞癌中的表达,并探讨其鉴别诊断意义。方法应用免疫组织化学S-P法对41例皮肤鳞状细胞癌,38例基底细胞癌和20例正常皮肤进行Cytokeratin7(K72.2)、Cytokeratin8(C-51)、Cytokeratin109(DE-K10)、Cytokeratin14(LL002)、Cytokeratin17(E3)、Cytokeratin18(D10)、Cytokeratin19(KS19.1)标记,观察不同细胞角蛋白的表达。结果41例皮肤鳞状细胞癌(包括高分化癌36例,中分化癌5例)和38例基底细胞癌中,7种细胞角蛋白的表达和分布有所不同,鳞状细胞癌显示CK10+/18+/19-;基底细胞癌为CK10-/18-/19+。结论有选择地检测一组CK的联合表达,有助于皮肤上皮性肿瘤的鉴别诊断。     

上皮样间质过度增生伴细胞角蛋白异常表达的恶性叶状肿瘤

目的分析伴上皮样间质过度增生及细胞角蛋白异常表达的恶性叶状肿瘤的诊断陷阱。方法对1例伴上皮样间质过度增生及细胞角蛋白异常表达的恶性叶状肿瘤的临床、形态、免疫组化特点,进行描述并结合文献进行分析。结果乳腺肿物穿刺组织表现为在黏液炎性的背景中见上皮样细胞弥漫分布,免疫组化:AE1/AE3、CK5/6、CK19、p63(+),CD34、bcl-2(-),组织形态及免疫组化表达均支持肉瘤样癌(化生性癌),切除标本广泛取材,可见到形态和免疫组化表达与穿刺标本相似的区域,同时也可见到典型的叶状肿瘤区域。结论伴上皮样间质过度增生的恶性叶状肿瘤在取材有限的情况下,即使加做免疫组化,依然容易误诊为肉瘤样癌。     

皮肤损害过程中肉芽组织形成与微血管结构的关系

目的:皮肤创伤处理不当可引发以上皮和肉芽组织过度增生为特征的假性上皮瘤样增生(PEH)病变。探讨微血管密度及其基底膜相关成分缺乏机制与PEH病变形成的关系。方法:将11例来自创(烧)伤后继发PEH病变(n=11)及其边缘正常皮肤(PEH-N,n=6)标本,采用免疫组织化学双染色方法观察微血管内皮细胞(micovascularordothelialcell,MEC)CD34、层粘连蛋白(LN)、Ⅳ型胶原、基质金属蛋白酶-2(MMP-2),MMP-3和MMP-9蛋白的表达水平和特征,并结合组织病理学和透射电镜观察MEC组织形态学改变。结果:与PEH-N相比,PEH组织中CD34,LN和Ⅳ型胶原阳性微血管分布密集,但浅层组织MEC的MMP-2,-3,-9蛋白水平升高,尤以MMP-9升高明显,且基底膜LN和Ⅳ型胶原信号显著减弱甚至消失,血管外炎症细胞密集浸润。超微结构显示,浅层肉芽组织中MEC分裂旺盛,缺乏完整的基底膜结构。结论:MEC过度表达MMPs是导致基底膜和间质相关成分缺失、炎症细胞浸润和上皮组织假性瘤样增生等连锁反应的重要环节。     

皮肤损害过程中肉芽组织形成与微血管结构的关系

目的：皮肤创伤处理不当可引发以上皮和肉芽组织过度增生为特征的假性上皮瘤样增生(PEH)病变。探讨微血管密度及其基底膜相关成分缺乏机制与PEH病变形成的关系。方法：将11例来自创(烧)伤后继发PEH病变(n=11)及其边缘正常皮肤(PEH-N，n=6)标本，采用免疫组织化学双染色方法观察微血管内皮细胞(micovascular ordothelial cell，MEC)CD34、层粘连蛋白(LN)、Ⅳ型胶原、基质金属蛋白酶-2(MMP-2)，MMP-3和MMP-9蛋白的表达水平和特征，并结合组织病理学和透射电镜观察MEC组织形态学改变。结果：与PEH-N相比，PEH组织中CD34，LN和Ⅳ型胶原阳性微血管分布密集，但浅层组织MEC的MMP-2，-3，-9蛋白水平升高，尤以MMP-9升高明显，且基底膜LN和Ⅳ型胶原信号显著减弱甚至消失，血管外炎症细胞密集浸润。超微结构显示，浅层肉芽组织中MEC分裂旺盛，缺乏完整的基底膜结构。结论：MEC过度表达MMPs是导致基底膜和间质相关成分缺失、炎症细胞浸润和上皮组织假性瘤样增生等连锁反应的重要环节。     

甲状腺微小乳头状癌19例临床病理分析

目的探讨甲状腺微小乳头状癌的临床病理特点。方法常规病理检查400例经手术切除的甲状腺良性疾病标本，筛选出微小癌病例，应用光镜观察其形态改变，并用SP法检测甲状腺球蛋白(Tg)、细胞角蛋白(CK／34βE12)、上皮膜抗原(EMA)的表达特点。结果400例中共检出微小乳头状癌19例，检出率为4．80%。具有诊断意义的主要组织学特征为毛玻璃状(透明)细胞核；核内假包涵体；核沟；纤维性间质反应或砂砾体。免疫组化标记17例高分子质量细胞角蛋白、16例上皮膜抗原、19例甲状腺球蛋白均呈( )。结论甲状腺微小乳头状癌为乳头状癌变型的特殊表现。高分子质量角蛋白和上皮膜抗原表达强阳性对于诊断具有较高的特异性。     

明胶酶B及其抑制物与大鼠肾脏衰老关系的研究

目的　探讨明胶酶B(MMP 9)及其抑制物 (TIMP 1)在大鼠肾脏衰老过程中的作用。方法　选用 3月、12月、2 4月龄大鼠 ,采用免疫组化技术分别检测MMP 9、TIMP 1、Ⅲ型胶原 (ColⅢ )、纤连蛋白 (FN)、增殖细胞核抗原 (PCNA)及α 平滑肌肌动蛋白 (α SMA)在不同鼠龄大鼠肾组织中的表达。结果　TIMP 1主要表达在肾小球、小管间质及血管 ,并随增龄表达增强 (P <0 0 1) ;MMP 9、主要表达在肾小管上皮细胞 ,随增龄表达无变化 ;ColⅢ主要表达在肾小球及小管间质 ,随增龄表达增强 (P <0 0 1) ;FN主要表达在肾小球及小管间质 ,随增龄表达增强 (P <0 0 1) ;α SMA除正常在血管平滑肌表达以外 ,在 2 4月鼠肾间质纤维化部位、肾小球系膜细胞及增厚的包曼氏囊有阳性表达 ;PCNA在 2 4月鼠肾小球、肾小管的细胞核及肾间质纤维化部位的成纤维细胞核有表达。TIMP 1与肾小球硬化有相关性 (P <0 0 5 )。TIMP 1与小管间质纤维化面积有相关性 (P <0 0 5 )。结论　MMP 9/TIMP 1表达失衡在大鼠肾脏衰老过程中可能起重要作用。     

皮肤增生性病变研究的最新进展：上皮问质过度增生的假上皮瘤样增生

假上皮瘤样增生(pseudoepitheliomatous hyperplasia，PEH)病变是临床上罕见的以上皮一间质过超长生为特征的良性病变，多继发于皮肤创(烧)伤、炎症刺激、慢性皮肤病和皮肤肿瘤等多种疾病。部分病例可见。calretinin等神经相关蛋白异常表达和表皮生长因子受体信号通路的活化。上皮一间质细胞之间激烈的相互作用引起是上皮细胞过度增殖和异常分化，可能是PEH病变发生的重要机制，对研究肿瘤发生机制具有指导意义。就PEH的原发病、临床和病理组织学特征、组织学起源和发病机制等内容进行概述。     

宫颈腺样基底细胞癌临床病理观察

目的进一步认识宫颈腺样基底细胞癌的病理形态及临床特点。方法应用细胞学、组织病理学、免疫组化等方法对1例宫颈腺样基底细胞癌进行分析,并结合相关文献讨论。结果液基涂片细胞学显示有异型的鳞状上皮细胞,符合高级别鳞状上皮内病变的形态学改变。光学显微镜下观察发现,宫颈鳞状上皮下方的间质内有圆形至卵圆形的小细胞巢,部分癌细胞巢的中央出现鳞状分化,周围呈栅栏状结构;肿瘤细胞巢与表面CIN3融合。免疫组化显示p63、p16和CK5/6(+),Ki-67灶状(+);宫颈间质内的部分血管平滑肌及纤维母细胞actin(+),而在癌巢周围呈(-)。结论宫颈腺样基底细胞癌罕见,具有独特的组织病理学特点。宫颈间质内的瘤组织易误认为鳞状上皮化生或被认为CIN累腺。诊断宫颈腺样基底细胞癌,须掌握严格的诊断标准,以作出正确诊断。     

子宫上皮样滋养细胞肿瘤

目的:探讨子宫上皮样滋养细胞肿瘤的临床病理学特征。方法:复习3例子宫上皮样滋养细胞肿瘤的临床资料,并进行组织病理学和免疫组织化学方法观察。结果:3例子宫上皮样滋养细胞肿瘤患者的年龄分别36、46和48岁。临床上主要表现为不规则的阴道流血和盆腔包块。组织学检查显示肿瘤主要由相对单一的单个核滋养细胞组成,排列成片状和条索状的紧密细胞巢,常见广泛坏死区周围绕以存活的瘤细胞岛,形成“地图样外观”。免疫组化标记检查显示瘤细胞表达人体胎盘催乳素(hPL)、β-人绒毛膜促性腺激素(β-hCG)、胎盘碱性磷酸酶(PLAP)、细胞角蛋白(CKpan)、CK18、上皮膜抗原(EMA)、表皮因子生长受体(EGFR)、上皮钙黏素、黑色素瘤细胞黏附分子(Mel-CAM)和抑制素-α。结论:子宫上皮样滋养细胞肿瘤是罕见的、起源于中间滋养细胞的肿瘤。确诊依赖于组织病理学观察和免疫组织化学标记。     

THE HISTOGENESIS OF BASEMENT MEMBRANES

A parietal yolk sac carcinoma of the mouse that secretes large quantities of basement membrane-like material has been used to study the formation of basement membranes. Suitably characterized fluorescein-labeled antibodies against this material stained basement membranes of epithelial structures and vessels, as well as reticulin. When absorbed with reticulin and vascular basement membranes of the spleen until these structures no longer fluoresced, the antibody still stained the basement membrane-like material of the tumor, its normal embryonic counterpart (Reichert's membrane), and the basement membranes at the bases of epithelial cells. The observation made previously that parietal yolk sac cells secreted, in the absence of connective tissue and reticulin, the basement membrane (Reichert's membrane) upon which they rested has been confirmed through the localization of ferritin-labeled antibody to the endoplasmic reticulin of the secreting cells. Since a basement membrane proven to be an epithelial secretion is antigenically similar to basement membranes at the bases of all epithelial cells studied but antigenically different from connective tissue elements, it is postulated that the basement membranes at the bases of epithelial cells in general are an epithelial secretion, and are not a condensation of ground substance as is commonly believed.     

脱细胞猪角膜表面基底膜成分的检测

背景:前期实验显示脱细胞猪角膜具有良好的组织相容性,可以支持角膜细胞和皮肤上皮细胞的生长。目的:检测脱细胞猪角膜是否保存了利于角膜上皮细胞生长的重要组织结构—基底膜。方法:利用荧光抗体对脱细胞猪角膜表面的基底膜成分(层粘蛋白和Ⅳ型胶原)进行免疫组织化学检测,荧光显微镜下观察脱细胞猪角膜表面是否保存了基底膜成分。结果与结论:免疫荧光染色显示脱细胞猪角膜前基质表面层粘蛋白和Ⅳ型胶原呈阳性表达,与新鲜猪角膜表面基底膜的荧光表达相同,表明脱细胞猪角膜保存了利于角膜上皮细胞生长的基底膜。     

MMP-3在人大肠癌组织中的表达及意义

目的:检测基质金属蛋白酶-3(MMP-3)在人大肠癌组织中的表达,以探讨其与人结肠癌进程的相关性。方法:应用光镜、电镜对结肠癌组织进行形态学观察,用免疫组化方法检测MMP-3的表达。结果:结肠癌晚期组MMP-3表达水平高于早期组,差异有非常显著性(P<0.01)。电镜观察可见在结肠癌早期,淋巴细胞和树突状细胞浸润较多,癌细胞穿基膜不明显;晚期,淋巴细胞和树突状细胞浸润较少,癌细胞穿基膜明显。结论:MMP-3在大肠癌晚期组织中的表达高于早期。提示MMP-3与结肠癌进程有直接关系。MMP-3的表达程度可作为判定结肠癌侵袭、转移和预后的指标。     

AN ULTRASTRUCTURAL STUDY OF GLOMERULAR PERMEABILITY IN AMINONUCLEOSIDE NEPHROSIS USING CATALASE AS A TRACER PROTEIN

Beef liver catalase (mol wt 240,000) was injected intravenously into normal rats and rats made nephrotic with aminonucleoside of puromycin. The localization of the tracer in the kidneys was then studied by ultrastructural cytochemistry, 3 min–12 hr after injection. Passage of catalase into the urinary space in normal rats was restricted by the basement membrane and by the epithelial slit pore. Nephrotic glomeruli showed extensive fusion of foot processes and formation of pockets and vacuoles in the fused epithelium; within 3 min after injection, catalase appeared in basal pockets, epithelial vacuoles, and the urinary space. Residual slit pores and close junctions in fused epithelium were impermeable to catalase. These studies indicate that alteration of the epithelial cells and basement membrane is responsible for protein leakage in aminonucleoside nephrosis.     

Thrombin suppresses matrix metalloproteinase 2 activity and increases tissue inhibitor of metalloproteinase 1 synthesis in cultured human peritoneal mesothelial cells.

OBJECTIVE: Recently, high levels of intraperitoneally generated thrombin were found in the effluent of patients treated with continuous ambulatory peritoneal dialysis (CAPD). The aim of the present study was to investigate in human peritoneal mesothelial cells (HMCs) the effect of thrombin on the activity and synthesis of matrix metalloproteinases (MMPs), which regulate the degradation of basement membrane collagen. METHODS: Cultured HMCs were isolated from omental tissue and used at confluence for the experiments. Conditioned media were obtained by incubating cells with serum-free M199 containing the relevant doses of thrombin. Activity of MMP-2 and MMP-9 were determined by an activity assay system. The antigen levels of MMPs and of the specific tissue inhibitor of metalloproteinases 1 (TIMP-1) were measured by ELISA. Northern blot analysis was applied to analyze mRNA expression of MMP-2 and TIMP-1. RESULTS: Incubation of HMCs with increasing doses of thrombin resulted in a concentration- and time-dependent suppression of MMP-2 activity. No changes in MMP-9 activity were seen. After a 48-hour stimulation period with thrombin (5 U/mL), MMP-2 activity decreased to 53% of that seen in control conditions. Antigen measurements revealed that this decrease was paralleled by a slight reduction in MMP-2 levels, which became significant at a thrombin dose of 5 U/mL [50.65 +/- 7.5 ng/10(5) cells (48 hours, 5 U/mL) vs 64.6 +/- 10.1 ng/10(5) cells (control)]. Under the same conditions, TIMP-1 levels were considerably increased [3.9 +/- 0.46 microg/10(5) cells (48 hours, 5 U/mL) vs 1.2 +/- 0.14 microg/10(5) cells (control)]. Hirudin (10 U/mL) completely inhibited the thrombin-induced effects on MMP-2 and TIMP-1 synthesis. These results were also reflected by Northern blot hybridization, where a slight decrease in MMP-2 and an increase in TIMP-1 mRNA expression were observed in response to thrombin. CONCLUSIONS: Our results suggest that high thrombin levels suppress MMP-2 activity through decreased MMP-2 and increased TIMP-1 synthesis. Thus, thrombin may promote the accumulation of basement membrane collagen. In addition to fibrin formation, this mechanism may represent a further contribution by thrombin to peritoneal thickening during CAPD.     

A bioartificial environment for kidney epithelial cells based on a supramolecular polymer basement membrane mimic and an organotypical culture system

Renal applications in healthcare, such as renal replacement therapies and nephrotoxicity tests, could potentially benefit from bioartificial kidney membranes with fully differentiated and functional human tubular epithelial cells. A replacement of the natural environment of these cells is required to maintain and study cell functionality cell differentiation in vitro. Our approach was based on synthetic supramolecular biomaterials to mimic the natural basement membrane (BM) on which these cells grow and a bioreactor to provide the desired organotypical culture parameters. The BM mimics were constructed from ureidopyrimidinone (UPy)‐functionalized polymer and bioactive peptides by electrospinning. The resultant membranes were shown to have a hierarchical fibrous BM‐like structure consisting of self‐assembled nanofibres within the electrospun microfibres. Human kidney‐2 (HK‐2) epithelial cells were cultured on the BM mimics under organotypical conditions in a custom‐built bioreactor. The bioreactor facilitated in situ monitoring and functionality testing of the cultures. Cell viability and the integrity of the epithelial cell barrier were demonstrated inside the bioreactor by microscopy and transmembrane leakage of fluorescently labelled inulin, respectively. Furthermore, HK‐2 cells maintained a polarized cell layer and showed modulation of both gene expression of membrane transporter proteins and metabolic activity of brush border enzymes when subjected to a continuous flow of culture medium inside the new bioreactor for 21 days. These results demonstrated that both the culture and study of renal epithelial cells was facilitated by the bioartificial in vitro environment that is formed by synthetic supramolecular BM mimics in our custom‐built bioreactor. Copyright © 2015 John Wiley & Sons, Ltd.     

GLOMERULAR PERMEABILITY : ULTRASTRUCTURAL STUDIES IN EXPERIMENTAL NEPHROSIS USING HORSERADISH PEROXIDASE AS A TRACER

Wistar/Furth rats were made nephrotic by daily administration of amino-nucleoside of puromycin, and the ultrastructural localization of horseradish peroxidase (mol wt 40,000) in the renal glomerulus was studied from 1 min to 20 hr after intravenous injection of the tracer. In control rats, peroxidase permeated the endothelial fenestrae, the basement membrane, and the epithelial slits, and was present in tubular lumina. Nephrotic glomeruli showed relatively normal basement membranes, extensive fusion of foot processes with formation of "close" intercellular junctions, and large vacuoles and pockets in epithelial cells. On serial sections some of the epithelial vacuoles communicated on one side with the extracellular space overlying basement membrane, and on the other side with the urinary space. In nephrotic animals, peroxidase permeated the basement membrane and the close junctions, and was present in many of the vacuoles and pockets as early as 1 min after injection. Only small numbers of peroxidase-positive vacuoles remained in. epithelial cells 1 hr or more after injection of the tracer. It is suggested that the epithelial pockets and vacuoles form pathways across which leaking proteins can be transferred across the epithelium into the urinary space. Epithelial vacuoles may also be absorption droplets designed to "conserve" leaking proteins, but this function was not prominent in our experiments with peroxidase.     

体外培养人角膜缘上皮细胞抑制激活态角膜基质细胞的生长

背景：角膜受到损伤后,角膜基质细胞激活转变为成纤维细胞,引起角膜基质瘢痕化,导致视力下降甚至丧失。目的：观察角膜不同部位上皮细胞与角膜基质细胞的相互作用,探索角膜缘上皮细胞群能否抑制激活态角膜基质细胞的生长。方法：采用酶消化及机械外力相结合的方法获取人角膜中央、角膜旁中央及角膜缘处角膜上皮细胞与浅层角膜基质细胞,进行体外培养。相差显微镜下观察细胞形态及生长变化。待培养角膜上皮细胞与基质细胞发生接触抑制时,记作＂0周＂,采用免疫荧光染色技术检测培养细胞中PCNA及p63蛋白的表达。结果与结论：培养的角膜上皮细胞与成纤维细胞发生接触抑制时,两种细胞间有明显分界线。角膜缘组上皮细胞中PCNA及p63蛋白均有较高的表达;角膜旁中央组PCNA有较高的表达,p63蛋白阴性表达;角膜中央组PCNA表达较低,p63蛋白阴性表达;从鉴定结果中可以得出只有角膜缘组中存在一定比例的角膜缘上皮干细胞。角膜缘组上皮细胞逐渐包围并化解成纤维细胞,在相互作用4周后,成纤维细胞聚集成死细胞团,缺乏角膜缘干细胞的中央组及旁中央组中成纤维细胞生长面积增加,上皮细胞生长受到抑制甚至死亡。说明体外培养的角膜缘上皮细胞群可以抑制激活态角膜基质细胞的生长。     

Cellular pathology of Heyman nephritis: monoclonal antibodies against the pathogenic antigen Gp 330

Heymann nephritis is an experimental auto-immune glomerulonephritis, closely resembling human epimembranous nephritis, which is induced by identified antigens in the brush border membranes of kidney proximal tubules. The hallmark of the disease is the accumulation of immune deposits in a granular pattern in the lamina rara externa of the glomerular basement membrane. We have established that a large membrane glycoprotein with an apparent molecular weight of 330 kDal (pg 330) is the pathogenic antigen. By means of immunohistochemistry, using mono- and polyclonal anti-pg 330 IgG we found that gp 330 is present in the cell membranes of glomerular epithelial cells and that it is concentrated there in coated pits (specialized areas off the cell membrane which play a key role in receptor mediated endocytosis for ligands such as insulin and others). On the basis of these findings we propose the following mechanism of formation of an immune deposit: (1) "In-situ" formation of immune complexes of gp 330 and anti-gp 330 IgG; (2) shedding of the immune complexes into the basement membrane and (3) crosslinking of the immune complexes into the basement membrane. This scheme could be a general mechanism by which immune deposits are formed also in various other auto-immune diseases.