Patterns of nestin and other intermediate filament expression distinguish between gastrointestinal stromal tumors,leiomyomas and schwannomas

The KIT-positive specific gastrointestinal stromal tumors (GISTs), leiomyomas, and schwannomas are the three most common types of primary mesenchymal tumors of the gastrointestinal (GI) tract. The intermediate filaments are abundant cytoskeletal proteins commonly used as cell differentiation markers in diagnostic immunohistochemistry. Their patterns have not been fully characterized in GI mesenchymal tumors, and could offer differential diagnostically useful parameters. Very recently, nestin, a class VI intermediate filament expressed in neuroectodermal stem cells and skeletal muscle progenitor cells, has been shown in GISTs and suggested as a marker for these tumors. In this study we immunohistochemically examined the expression of nestin and other intermediate filament proteins, including desmin, keratins (Ks), glial fibrillary acidic protein (GFAP), neurofilament, and vimentin in GISTs of different sites, esophageal leiomyomas and GI schwannomas. Nestin was nearly consistently present in GISTs of different locations whether spindle cell or epithelioid, and benign or malignant. It was also detected in 23 of 24 (96%) GI schwannomas, whereas leiomyomas were uniformly negative. Vimentin was present in both GISTs and schwannomas, whereas it was typically absent in leiomyomas (25% positive, usually focally). Desmin was present in all leiomyomas, whereas only 3% of GISTs (4 of 140) were positive, and all schwannomas were negative. K18 was detected in a minority of GISTs, leiomyomas, and schwannomas. Malignant GISTs were more commonly keratin positive than the benign ones; there was 18% K18 positivity in malignant gastric and small intestinal GISTs, but 9% K18 positivity in benign gastric and small intestinal GISTs. Moreover, K8, albeit to a lesser degree, was detected in a minority of GISTs, but K7, K14, K19 and K20 were not detected. GFAP was present in the majority of schwannomas, whereas all GISTs were negative; some leiomyomas had weak cytoplasmic positivity. These results document distinctive patterns of intermediate filament proteins in GI mesenchymal tumors. Nestin is confirmed to be consistently expressed in GISTs but it is also present in most GI schwannomas; GFAP is helpful when separating GISTs and schwannomas, since only the latter are positive. The potential presence of K8 and K18 in GISTs should not lead to the misdiagnosis of carcinoma on biopsy.     

胃肠道间质瘤中DOG1和nestin的表达及意义

目的:探讨DOG1和nestin在胃肠道间质瘤(gastrointestinal stromal tumor, GIST)中的表达及其诊断价值.方法:采用免疫组织化学EnVision法分别检测25例GIST、5例平滑肌瘤和5例神经鞘瘤中DOG1和nestin的表达.结果:DOG1和nestin在GIST中的阳性表达率分别为100%和88%,而CD117和CD34在GIST中的阳性表达率分别为84%和64%.GIST中DOG1和nestin的表达与肿瘤部位、肿瘤大小、危险度分级和组织学形态均无关(P>0.05).5例平滑肌瘤中DOG1和nestin均阴性表达,DOG1和nestin在神经鞘瘤中的阳性表达率分别为0和100%.DOG1,nestin,CD117和CD34联合检测阳性率为48%.结论:DOG1在诊断GIST中具有敏感性和特异性,而nestin则是GIST诊断中比较敏感的一种标志物,但特异性差.DOG1,nestin,CD117和CD34联合检测可提高GIST的正确诊断.     

Gastrointestinal pacemaker cell tumor (GIPACT): gastrointestinal stromal tumors show phenotypic characteristics of the interstitial cells of Cajal.

The interstitial cells of Cajal (ICC) form a complex cell network within the gastrointestinal tract wall where they function as a pacemaker system. Expression of the kit proto-oncogene is essential for the development of this system. The aim of our study was to examine the hypothesis that gastrointestinal stromal tumors differentiate toward cells with an ICC phenotype. Ultrastructurally, 58 stromal tumors were characterized and found to share many features with ICC. Seventy-eight stromal tumors were immunophenotyped, particularly with regard to the kit receptor. All 78 tumors revealed strong, homogeneous immunoreactivity for the kit receptor as did ICC of adjacent and control gastrointestinal walls. Focal hyperplasia and hypertrophy of kit receptor positive cells were also observed in the gastrointestinal wall adjacent to the tumors. CD34 immunoreactivity observed in interstitial cells surrounding Auerbach's ganglia suggests that a subpopulation of ICC is CD34 positive and may explain why 56 of 78 stromal tumors were CD34 positive. Thirty control tumors, including gastrointestinal leiomyomas and leiomyosarcomas, were all negative for the kit receptor. We conclude that gastrointestinal stromal tumors show striking morphological and immunophenotypic similarities with ICC and that they may originate from stem cells that differentiate toward a pacemaker cell phenotype. We propose that the noncommittal name "gastrointestinal stromal tumor" be replaced by gastrointestinal pacemaker cell tumor.     

Structure and organization of interstitial cells of Cajal in the gastrointestinal tract

The morphological features of interstitial cells of Cajal (ICC) in the gastrointestinal (GI) tract are described based on observations of laboratory animals including mice, rats and guinea-pigs, using immunohistochemical staining for Kit and electron microscopy. ICC show a specific distribution, arrangement and cell shape depending on their location within various regions and tissue layers of the GI tract. Hence they are classified into several subtypes. The stomach shows distinct regional variations in the distribution of subtypes of ICC from the cardia to pylorus, whereas the small intestine and colon both seem to retain nearly the same distribution pattern of subtypes of ICC throughout each organ. All subtypes of ICC share common ultrastructural features, such as the presence of numerous mitochondria, abundant intermediate filaments, and formation of gap junctions with the same type of cells and with smooth muscle cells. In addition, depending on their species and anatomical location, some subtypes of ICC show some features typical of smooth muscle cells including a basal lamina, caveolae, subsurface cisterns and dense bodies. ICC are somewhat heterogeneous morphologically. A question is raised on a special relationship between their ultrastructural features and dependency on Kit/stem cell factor system. As the neuromediator function of ICC, reciprocal distribution of ICC and gap junctions in the muscle coat is demonstrated by the comparison of Kit immunoreactive cells and gap junction protein connexin 43 in both small intestine and colon.     

Clinicopathologic and immunohistochemical characteristics of upper gastrointestinal leiomyomas harboring interstitial cells of Cajal: A potential mimicker of gastrointestinal stromal tumor

ObjectiveTo analyze clinicopathologic characteristics of upper gastrointestinal leiomyomas and to determine the distribution and immunohistochemical features of interstitial cells of Cajal, in order to designate whether they can cause diagnostic challenges.Materials and methodsTwenty-four upper gastrointestinal leiomyomas (14 esophagus, 10 stomach) were retrieved. CD117, DOG-1 and muscle markers were performed. The staining was analyzed based on the distribution and percentage. Interstitial cells of Cajal were distinguished based on their positivity for both CD117 and DOG-1 immunohistochemistry, along with their morphological features.ResultsMean age of patients was 49 years, M/F ratio was 2.4. Patients with gastric leiomyomas were significantly younger than those with esophageal leiomyomas (41.5 vs. 54.3, p = 0.012). Histologically, leiomyomas were similar to their endometrial counterpart. Immunohistochemically, all tumors had strong/diffuse positivity for muscle markers. CD117 highlighted mast cells in all cases. Three cases had prominently increased mast cells. Both CD117 and DOG-1 also highlighted interstitial cells of Cajal in 24/24 (100%) of cases. Interstitial cells of Cajal were distributed in variable proportions, from focal to homogenous. In one case, they constituted 50% of tumor cells. In 16 cases, the distribution was homogenous. Superficial leiomyomas (n = 3) had only focal CD117 and DOG-1 positivity.ConclusionUpper gastrointestinal leiomyomas harbor expression of CD117 and DOG-1 in entrapped/colonized interstitial cells of Cajal, which can cause a potential pitfall in the differential diagnosis, especially in cases that show prominent immunohistochemical positivity. Evaluation of the immunohistochemistry can be exceptionally challenging in small biopsy/cytology specimens. Careful histologic evaluation of the tumor as well as the recognition of interstitial cells of Cajal will help the pathologist render the accurate diagnosis.     

Malignant stromal tumor of the gallbladder with interstitial cells of Cajal phenotype

Malignant mesenchymal tumors of the gallbladder are exceedingly rare. We report a malignant stromal tumor of the gallbladder with a phenotype of interstitial cells of Cajal. To our knowledge, only the benign counterpart of this tumor has been described previously. A 34-year-old woman presented with right upper quadrant abdominal pain. At the time of cholecystectomy, the gallbladder was noted to have a thickened wall and a polypoid mass arising in the neck of the gallbladder. Histologic sections showed a cellular proliferation of spindle neoplastic cells that were arranged in short fascicles. Numerous mitotic figures and foci of necrosis were noted. The neoplastic cells expressed CD117 (c-Kit protein) and vimentin. They were negative for smooth muscle actin, desmin, myoglobin, cytokeratin, S100 protein, and CD34. Our case demonstrates that a malignant stromal tumor that is histologically and immunohistochemically identical to gastrointestinal stromal tumor can occur in the gallbladder, and that the expression of CD117 may be of therapeutic importance.     

Expression of intermediate filament proteins in thyroid gland and thyroid tumors

The presence of intermediate filament proteins of cytokeratin/prekeratin type and vimentin type was evaluated in non-neoplastic thyroid glands and in different types of thyroid neoplasms. Follicular epithelium of both normal and goitrous thyroids showed a strong reaction with anticytokeratin antibodies that widely cross-react with various simple epithelia. On the other hand, in normal thyroid, there were only occasionally (in one of 12 cases) solitary cells reacting with antibodies to epidermal prekeratin. In nodular goiters, such cells were often seen (eight of 18), especially among the lining cells of cysts, and in chronic thyroiditis in all (12 of 12) cases. Only the stromal cells and intraluminal macrophages reacted with antibodies to vimentin. Neoplastic cells of papillary carcinomas showed a positive staining reaction both with antibodies to cytokeratins and to epidermal prekeratin. Follicular carcinoma cells, although positive for cytokeratins, could generally not be stained with antibodies to epidermal prekeratin. Medullary carcinoma cells also showed cytokeratin positivity and, only occasionally, positivity for epidermal prekeratin. Anaplastic carcinomas were also reactive with antibodies to cytokeratin but, for the most part, were negative for epidermal prekeratin. Interestingly, some neoplastic cells of all types of thyroid carcinomas also appeared to contain vimentin, as shown with both polyclonal and monoclonal antivimentin antibodies. In contrast to carcinomas, the intermediate filaments of thyroid sarcomas and lymphomas were only of vimentin type. Furthermore, it was found that the papillary structures in benign goiters were only reactive with cytokeratin antibodies and lacked, in contrast to papillary carcinomas, epidermal prekeratin-like immunoreactivity. Hence, the analysis of intermediate filament proteins of thyroid tumors can be utilized to differentiate between papillary and follicular carcinomas and between benign and malignant papillary lesions as well as between anaplastic thyroid carcinomas and sarcomas or lymphomas.     

长爪沙鼠胃肠道Cajal间质细胞的形态学

目的 探讨长爪沙鼠胃肠道Cajal间质细胞（ICCs）的形态和分布规律。 方法 采用10只成年长爪沙鼠,体重50~70g,取胃、小肠、结肠制作冷冻切片,结合全层铺片的c-Kit免疫荧光染色。结果 ICCs呈网络状分布于整个胃肠道,不同部位ICCs的分布及形态有所不同。在胃底部,仅见肌内ICCs(ICC-IM）,而在胃体和胃窦部除ICC-IM外,可见肌间ICCs(ICC-MY)分布在肌间神经丛周围;其细胞密度胃底ICC-IM最多,由胃底至胃窦逐渐减少,而ICC-MY由胃体至胃窦逐渐增多。在小肠可见ICC-IM, ICC-MY和深肌层ICCs(ICC-DMP)3个亚群,结肠管壁内也分布有ICC-IM、ICC-MY和黏膜下ICCs(ICC-SM)3个亚群。结论 沙鼠可用于有关ICCs正常形态、结构及功能的研究。     

豚鼠输尿管Cajal间质细胞毒蕈碱受体的表达及功能研究

目的研究毒蕈碱受体在豚鼠输尿管Cajal间质细胞（ICC）上的表达,初步探讨ICC在胆碱能神经递质传递中的作用。方法将20只豚鼠随机分成对照组和实验组,进行输尿管肌条实验,观察在卡巴胆碱作用下输尿管肌条的收缩幅度和频率的变化;10只豚鼠,将输尿管进行固定、撕片和荧光染色,检测豚鼠输尿管毒蕈碱受体亚型（M1-M5）在ICC上的表达情况。结果卡巴胆碱作用后,实验组肌条的收缩幅度为0 g,频率为0次/分,对照组的幅度为（0.106±0.021）g,频率为（4.900±0.875）次/分,两组差异显著（P〈0.01）。免疫荧光双标显示,输尿管ICC仅表达M2、M3受体亚型。结论胆碱能神经可能通过ICC来调控输尿管平滑肌。     

Involvement of intramuscular interstitial cells of Cajal in neuroeffector transmission in the gastrointestinal tract

Specialized cells known as interstitial cells of Cajal (ICC) are distributed in specific locations within the tunica muscularis of the gastrointestinal (GI) tract. ICC serve as electrical pacemakers, provide pathways for the active propagation of slow waves, are mediators of enteric motor neurotransmission and play a role in afferent neural signalling. Morphological studies have provided evidence that motor neurotransmission in the GI tract does not occur through poorly defined structures between nerves and smooth muscle, but rather via specialized synapses that exist between enteric nerve terminals and intramuscular ICC or ICC-IM. ICC-IM are coupled to smooth muscle cells via gap junctions and post-junctional responses elicited in ICC-IM are conducted to neighbouring smooth muscle cells. Electrophysiological studies from the stomachs and sphincters of wild-type and mutant animals that lack ICC-IM have provided functional evidence for the importance of ICC in cholinergic excitatory and nitrergic inhibitory motor neurotransmission. Intraperitoneal injection of animals with Kit neutralizing antibody or organ culture of gastrointestinal tissues in the presence of neutralizing antibody, which blocks the development and maintenance of ICC, has provided further evidence for the role of ICC in enteric motor transmission. ICC-IM also generate an ongoing discharge of unitary potentials in the gastric fundus and antrum that contributes to the overall excitability of the stomach.     

胃肠道Cajal间质细胞起搏功能的研究进展

正正常的胃肠运动功能依赖于节律性的胃肠蠕动,这种规律性的蠕动目前主要认为与肠神经系统及众多的神经递质有关,但其具体的调控机制目前尚有争议,主要争论的焦点在于肠神经系统所释放的神经递质是否直接作用于平滑肌细胞~([1-2])。近年来,越来越多的研究发现,Cajal间质细胞(interstitial cells of Cajal,ICC)在胃肠动力调节方面扮演着至关重要的作用~([3])。因此,本文拟就从生理结构基础、起     

bcl-2和p16在胃肠道间质瘤中的表达及其临床意义

目的 探讨bcl-2和p16的表达与胃肠道间质瘤(gastrointestinal stromal tumor，GIST)的良恶性及发生发展的关系。方法 应用免疫组织化学技术EnVision微波二步法检测40例GIST(良性20例，恶性20例)中bcl-2和p16蛋白的表达。结果 40例GIST中有29例表达bcl-2，23例表达p16，阳性率分别为72．5％和57．5％。bcl-2阳性信号定位于细胞质，p16阳性信号定位于细胞质和细胞核。bcl-2、p16的表达与GIST的良恶性、组织学分型、部位、性别及年龄均无关。结论 bcl-2及p16可能在GIST的早期阶段即参与肿瘤的发生发展，但两者不能作为判断肿瘤良恶性的指标。     

Expression of microtubule-associated protein tau by gastrointestinal stromal tumors.

The purpose of this work was to study the expression in gastrointestinal stromal tumors (GISTs) of various antigens, including the protein tau associated with enteric neuronal differentiation; to compare their expression with that of c-kit, known to be associated with interstitial cell of Cajal differentiation; and to correlate their expression with the observation of ultrastructural features of gastrointestinal autonomic nerve tumors. Twenty-six GISTs of the stomach and 16 GISTs of the small bowel were included in the study group. Thirty-five tumors served as controls. Tissue sections were immunostained with vimentin, CD34, desmin, specific smooth muscle actin, S100 protein, neuron-specific enolase, PGP9.5, neurofilament, bcl-2 oncoprotein, synaptophysin, chromogranin A, c-kit, and tau. Twenty-one of these tumors were also analyzed ultrastructurally. Of the 42 GISTs, 28 were predominantly spindled, 7 were predominantly epithelioid, and 7 were a mixture of epithelioid and spindle cells. Ten primary GISTs were classified as benign, 9 as borderline, and 23 as malignant. Metastatic dissemination was present at primary surgery in 1 case and eventually developed in 6 patients. Six disease-related deaths were counted. In normal submucous and myenteric plexuses of stomach and small bowel, ganglion cell bodies and nerve fibers strongly expressed tau. Twenty (76.9%) GISTs of the stomach and 12 (75%) of the small bowel expressed tau. Tau often showed intense, diffuse staining patterns in both spindled and epithelioid tumors. Ten (100%) of the 10 benign GISTs, 7 (77.8%) of the borderline GISTs, and 15 (65.2%) of the 23 frankly malignant GISTs expressed tau. Thirty-six GISTs expressed at least 2 different neuronal markers. A coexpression of the neuronal markers and c-kit was observed in 90% of GISTs. The expression of tau was observed in 12 of the 15 GISTs with dense core granules, considered as the definitive finding for a diagnosis of gastrointestinal autonomic nerve tumors. Ten of these also expressed c-kit; 9 were malignant. Tau also immunostained other intra-abdominal tumors, including neuroendocrine carcinomas, paragangliomas and desmoplastic round cell tumors. This immunohistochemical study shows that GISTs are specific tumors of the digestive tract and are nearly always characterized by simultaneous neuronal and interstitial cell of Cajal differentiation. Although the loss of tau expression is observed only in borderline and malignant tumors, its prognostic value is not clear cut.     

Ultrastructural characterization of the interstitial cells of Cajal

Recent studies on the interstitial cells of Cajal (ICC) have determined ultrastructural criteria for the identification of these previously enigmatic cells. This review deals with the electron microscopic findings obtained by the author's research group in different tissue regions of the gut in mice, rats and guinea-pigs, comparing these with reports from other groups in different species and in humans. ICC are characterized by the following morphological criteria: numerous mitochondria, abundant intermediate filaments and large gap junctions which connect the cells with each other and with smooth muscle cells. Due to their location in the gut and the specific species, the ICC are markedly heterogeneous in appearance, ranging from cells closely resembling smooth muscle cells to those similar to fibroblasts (Table 1). Nevertheless, the above-mentioned morphological features are shared by all types of ICC and serve in identifying them. Recent discoveries on a significant role of c- kit in the maturation of the ICC and their specific immunoreactivity to anti-c-Kit antibody have confirmed the view that the ICC comprise an independent and specific entity of cells. This view is reinforced by the findings of the author's group that the ICC characteristically possess vimentin filaments and are stained with the zinc iodide-osmium tetroxide method which provides a staining affinity similar to methylene blue, the dye used in the original work by Cajal, (1911). Developmental studies indicate that the ICC are derived from a non-neuronal, mesenchymal origin. This paper further reviews advances in the physiological studies on the ICC, in support of the hypothesis by THUNEBERG (1982) that they function as a pacemaker in the digestive tract and a mediator transmitting impulses from the nerve terminals to the smooth muscle cells.     

Pathology of gastrointestinal stromal tumors

Gastrointestinal stromal tumors (GIST) are the most common mesenchymal tumors in the gastrointestinal tract. It was found that most GIST expressed KIT, a receptor tyrosine kinase encoded by protooncogene c-kit. In normal gastrointestinal wall, KIT is expressed by interstitial cells of Cajal (ICC), which are a pacemaker for autonomous gastrointestinal movement. Because both GIST and ICC are double-positive for KIT and CD34, and because familial and multiple GIST appear to develop from diffuse hyperplasia of ICC, GIST are considered to originate from ICC or their precursor cells. It was also found that approximately 90% of the sporadic GIST have somatic gain-of-function mutations of the c-kit gene, and that the patients with familial and multiple GIST have germline gain-of-function mutations of the c-kit gene. These facts strongly suggest that the c-kit gene mutations are a cause of GIST. Approximately half of the sporadic GIST without c-kit gene mutations were demonstrated to have gain-of-function mutations in platelet-derived growth factor receptor-alpha (PDGFRA) gene that encodes another receptor tyrosine kinase. Because KIT is immunohistochemically negative in a minority of GIST, especially in PDGFRA gene mutation-harboring GIST, mutational analyses of c-kit and PDGFRA genes may be required to diagnose such GIST definitely. Imatinib mesylate was developed as a selective tyrosine kinase inhibitor. It inhibits constitutive activation of mutated KIT and PDGFRA, and is now being used for KIT-positive metastatic or unresectable GIST as a molecular target drug. Confirmation of KIT expression by immunohistochemistry is necessary for application of the drug. The effect of imatinib mesylate is different in various types of c-kit and PDGFRA gene mutations, and the secondary resistance against imatinib mesylate is often acquired by the second mutation of the identical genes. Mutational analyses of c-kit and PDGFRA genes are also significant for prediction of effectiveness of drugs including newly developed agents.     

Increased expression of the embryonic intermediate filament, nestin, in the brains of scrapie-infected mice

This study investigated the immunohistochemical localization of nestin and its expression level in the brains of mice infected with scrapie, a disease which is characterized by spongiform neurodegeneration, neuronal vacuolation, and astrogliosis. Western blot analysis showed that the protein level of nestin was significantly increased in scrapie-infected brains compared with those of control brains. Nestin immunoreactivity in vascular endothelial cells was more intense in the brains of scrapie-infected mice compared with those of controls. There was marked nestin immunostaining in reactive astrocytes, but not in the neurons of the scrapie-infected brains. Considering all the findings, this study suggests that the nestin-positive cells, including glial cells and vascular endothelial cells, may have the potential to differentiate into mature glial cells and other neuronal elements, which would replenish lost neurons.     

Cell markers in gastrointestinal stromal tumors

Stromal tumors of the gastrointestinal (GI) tract have generated considerable controversy about their direction and level of differentiation, particularly about whether the tumor cells are smooth muscle or Schwann cells. In an attempt to characterize these tumors, the immunohistochemical staining patterns of desmin, vimentin, actin, and S-100 protein were studied in 41 GI stromal tumors, using the avidin-biotin method, and compared with normal host smooth muscle and nerve and with esophageal and uterine leiomyomas. Twenty gastric and one rectal tumor stained diffusely with vimentin and actin, but not with desmin, and had scattered strongly S-100-positive cells that might either be trapped Schwann cells or tumor cells. Twenty small bowel tumors stained similarly to the gastric tumors with regard to vimentin, actin, and desmin, but most (17/20) had a unique, strongly positive geographic staining pattern with S-100. No differences in staining were noted between benign and malignant tumors in either gastric or small bowel sites, and most histologic patterns in tumors from similar locations stained similarly. These results suggest that GI stromal tumors are not truly "leiomyomas and leiomyosarcomas," but relatively undifferentiated tumors, with the expression of various antigens depending on their location in the gut.