Inchul Lee

Immunohistochemical evaluation of neuroendocrine cells and neoplasms of the lung.

The dispersed neuroendocrine (NE) system is represented in the bronchopulmonary tract by submucosal nerves and ganglion cells and, in the mucosal lining by solitary NE cells and neuroepithalial bodies (NEBs). The latter two components variably express pan-NE markers including NSE, chromogranin (s) and, notably, synaptophysin. The expression of serotonin, bombesin, calcitonin and leu-enkephalin has been well established; additional eutopic materials include somatostatin and calcitonin gene-related peptide. Solitary NE cells and NEBs are epithelial structures as defined by their consistent cytokeratin expression. Hyperplasia and dysplasia of NE cells may be found in association with various forms of chronic injury; they have been noted in chronic bronchiectasis and in the vicinity of neoplasms of various types. Hyperplastic and dysplastic pulmonary NE cells frequently express ectopic materials particularly ACTH. NE neoplasms of the bronchopulmonary tract comprice a spectrum that includes a) carcinoids, b) well differentiated NE carcinomas, c) intermediate cell NE carcinomas and d) small cell NE carcinomas. The precise pathologic criteria defining these entities are discussed in detail as are their clinical implications. The entire spectrum of lung NE neoplasms express NE markers demonstrable by immunocytochemistry; these include pan-NE markers, serotonin and numerous neuropeptides. The expression of multiple hormonal materials is frequent. Within any given tumor, some variation in expression may be noted in different sites and in different periods of the normal or therapeutically modified lifespan of the tumor. The entire spectrum of lung NE neoplasms is epithelial for they express cytokeratin polypeptides and desmoplakin; subsets of the tumors coexpress cytokeratins and neurofilament proteins. Also, subsets of these NE neoplasms may be immunostained with monoclonal antibodies to antigens related to exocrine phenotype suggesting focal amphicrine features.

Immunohistochemical Identification of Exocrine and Neuroendocrine Subsets of Large Cell Lung Carcinomas

Formalin fixed, paraffin embedded sections of 52 cases of pulmonary large cell undifferentiated carcinoma (LCUC) as defined in the current WHO classification were studied immunohistochemically to assess features of exocrine and neuroendocrine (NE) differentiation. Monoclonal antibody 44-3A6 was applied to detect a membrane association protein related to exocrine differentiation. A panel of ten neuroendocrine markers including antibodies to synaptophysin, chromogranin A, serotonin, and seven neuropeptides was used to assess NE differentiation. The broad spectrum anticytokeratin antibody PKK1 was used to confirm the epithelial differentiation of these tumors. Exocrine differentiation was detected in 40/52 (77%) of surgically resected LCUC, despite the absence of recognizable glands by light microscopy. Eighteen of 52 (35%) LCUC exhibited NE differentiation; synaptophysin was the most frequently detected NE marker. Cytokeratin immunostaining with PKK1 was demonstrated in 41/52 (79%) cases. Subsets of LCUC were defined based on their expression of exocrine or NE phenotypic markers. Accordingly, 28/52 (54%) LCUC displayed an exocrine phenotype, 6/52 (12%) a NE phenotype, 12/52 (23%) had combined exocrine and NE phenotypes, and 6/52 (12%) exhibited neither phenotype. In this surgical series, there were no significant differences in stage at presentation for the four subsets. Interestingly, two year survival appeared decreased in patients with tumors displaying the pure NE phenotype.

Paragangliomas of the Head and Neck: Ultrastructural and Immunohistochemical Analysis

Eighteen head and neck paragangliomas were studied by light microscopy and light microscopic immunohistochemistry by the peroxidase technique for the presence of NSE (neuron-specific enolase), serotonin, and a battery of neuropeptides. Seven of these tumors were also studied by electron microscopy. All 18 cases demonstrated immunostaining for NSE; 10 of the 11 carotid body tumors had immunostaining for multiple hormones. Considering all 18 cases, the most frequently demonstrated hormonal substances were in order: serotonin, leu-enkephalin, gastrin, substance P, vasoactive intestinal polypeptide (VIP), somatostatin, bombesin, calcitonin, and alpha MSH. In several tumors, adjacent-step sections stained for different hormonal substances strongly suggested reactivity for more than one hormone in given tumor cells. By electron microscopy, all 7 cases studied displayed considerable heterogeneity of the neurosecretory granules with respect to size, shape, and electron density. This demonstrated that branchiomeric paragangliomas are capable of producing a spectrum of neuropeptides in addition to their known amine content. The presence of immunoreactive serotonin in most of these neoplasms was confirmed. In addition to these findings, neurofibrils within the substance of carotid body paragangliomas demonstrated immunoreactivity for somatostatin and a gastrinlike neuropeptide. The significance of the neuropeptides in these neoplasms and their possible presence and role in normal and hyperplastic paraganglia remain to be defined.

Immunohistochemical evaluation of ras oncogene expression in pulmonary and pleural neoplasms.

SummaryWe undertook an immunohistochemical analysis of human bronchopulmonary epithelial neoplasms and pleural mesotheliomas using a monoclonal antibody which recognizes ras oncogene products (p21ras). The monoclonal antibody, RAP-5, recognizes both unaltered and certain mutated p21ras. Formalin fixed and paraffin embedded tissue samples of 187 lung epithelial tumors and 27 pleural mesotheliomas were investigated; normal and bronchiectatic lungs were similarly studied. Normal lung and pleural tissue did not immunostain except for occasional type II pneumocytes. Reactive type II pneumocytes adjacent to carcinomas and bronchiectasis immunostained consistently. Twenty four/34 (71%) squamous carcinomas immunostained. Only 8/50 (16%) adenocarcinomas immunostained focally and weakly whereas 19/24 (79%) bronchioloalveolar carcinomas immunostained. Eleven/18 (61%) large cell carcinomas immunostained with variable intensity. Eleven/13 (85%) carcinoids, 6/7 (85%) well differentiated neuroendocrine carcinomas, and 18/21 (86%) intermediate cell neuroendocrine carcinomas immunostained while none of 20 small cell neuroendocrine carcinomas immunostained. Only a few mesotheliomas were immunostained focally. Two/14 (14%) epithelial type and 1/9 (11%) biphasic type mesotheliomas immunostained weakly; none of 4 spindle cell mesotheliomas immuno stained.We conclude that while at least occasional cases of most types of pulmonary epithelial neoplasms express p21ras, the frequency and intensity of the expression are distinctly greater in certain tumor types such as squamous, bronchioloalveolar, and neuroendocrine neoplasm except for the small cell type. Contrary to these lung epithelial neoplasms, most mesotheliomas did not immunostain for p21ras. Whether the enhanced p21ras expression may point to a different mechanism of transformation or may merely reflect differentiation features remains undetermined.

Neuroendocrine skin carcinoma coexpressing cytokeratin and neurofilament proteins.

Neuroendocrine (Merkel cell) carcinomas of the skin have been recognized as such for several years. Given the reported wide variability in the morphology and clinical evolution of these tumors, the notion that may they comprise several variants rather than a single type has been advocated. Electron microscopy has played a key role in the early recognition of these tumors while immunohistochemical studies for various neuroendocrine markers have facilitated their subsequent diagnosis and improved our understanding as to their complexity by the demonstration of immunoreactivity for NSE (neuron specific enolase) and a number of neuropeptides. There has also been considerable interest in the study of the cytoskeletal intermediate filament complement of neuroendocrine neoplasms in general and of those of the skin in particular. Early reports indicated that neuroendocrine skin carcinomas had neurofilaments while subsequent investigations determined that they had cytokeratin. However, more recent studies have indicated that at least some neuroendocrine skin carcinomas could in fact coexpress both aforementioned classes of intermediate filament proteins. This brief report is presented to confirm the latter investigations.

Bombesin in human neuroendocrine (NE) neoplasms

Bombesin is a 14 amino acid peptide isolated from amphibian skin which was found to have stimulatory effects upon gastric and pancreatic secretions, release of gastrointestinal hormones, gallbladder contraction and bronchoconstriction. It is present in amphibian gastric endocrine cells, avian proventriculus endocrine cells and avian brain. In mammals it is present mainly in nerve cells and fibers. The only mammalian endocrine cell shown to date to have bombesin is the P-cell in fetal lung. Bombesin is also found in mammalian brain, with its highest concentration in the hypothalamus. We examined several groups of human neuroendocrine neoplasms for the presence of bombesin by immunohistochemistry. Our findings indicate that bombesin is present 68% of bronchial carcinoids, 65% of pulmonary neuroendocrine carcinomas, 62% of neuroendocrine carcinomas of the skin, 5-10% of pheochromocytomas and extraadrenal paragangliomas and 35% of gastrointestinal carcinoids and neuroendocrine carcinomas. Parallel studies in a wide variety of non neuroendocrine neoplasms failed to reveal the presence of bombesin. We conclude that bombesin is a highly specific marker of neuroendocrine differentiation and thus a valuable tumor marker. Furthermore, its specificity compares favorably with another neuroendocrine marker, neuron specific enolase, an enzyme thought to be present only in neural tissues and neuroendocrine cells but recently found in non neural human tissues and non neuroendocrine neoplasms.

Immunohistochemical analysis of normal and mutated ras oncogene p21 expression in human pulmonary and pleural neoplasms

SummaryIn this study we examined 214 cases of primary human pulmonary neoplasms for the expression of a mutated form of the ras oncogene p21 product, recognized by the monoclonal antibody (MCA) DWP. Adjacent serial sections from these same cases had previously been used to demonstrate the frequency of ras p21 expression using the broadly reactive anti-ras p21 MCA RAP-5. Confirmation of the increased expression of p21 was accomplished using MCA Yl3-259. The use of adjacent tissue sections from these cases allows the direct comparison of the expression of the mutated and non-mutated forms of ras p21. If reactivity with DWP would prove to be significantly more restrictive than that of the “pan” ras MCAs, RAP-5 and Yl3-259, it would lend support to the possibility that DWP (and similar MCAs which detect other specific mutations) could be used to define subsets of these neoplasms based on their specific ras p21 phenotype. Since one would anticipate that the valine/cysteine substitution at position 12 of the ras p21 would occur at only low frequencies in human tumors, our results with DWP are consistent with this hypothesis. As previously reported, RAP-5 reacted with a high proportion of lung tumors (100/214 or 47%). In this report, we demonstrate the selective expression of the mutation recognized by the MCA DWP in only 5% of these same tumors (13/214), and that the expression of this mutated form is not restricted to any of the conventional histological subclasses of pulmonary neoplasms.

Expression of ras oncogene p21 during human fetal development as determined by monoclonal antibodies RAP-5, Y13-259, and DWP

SummaryIn this report we describe the expression of the ras proto-oncogene p21 protein in various tissues during normal fetal development. Conventional, formalin fixed and paraffin-embedded sections of normal organs were examined from fetuses ranging 9 to 42 weeks of gestation. Immunohistochemical localization of ras p21 was accomplished using the broadly reactive, mouse monoclonal antibodies RAP-5 and Y13-259. The monoclonal antibody DWP, which is specific for a mutated form of ras p21 having a valine/cysteine at amino acid position 12, was also used. Detectable expression of the p21 protein was seen at different time periods during fetal development depending on the tissue. The expression of ras p21 (as detected by RAP-5 and Yl3-259) was noted in a wide range of cell types and tissues; intense immunostaining was noted in epithelial cells of the gastrointestinal tract, exocrine and endocrine pancreas, renal tubules and transitional urotheliem, as well as in other tissues. This immunostaining generally, but not invariably, corresponded with patterns previously reported in benign and/or malignant neoplasms of adult tissues. In most instances ras p21 expression, when present, occured during periods of rapid growth in given organ systems. However, some actively proliferating fetal tissues such as thymus and spleen, failed to express detectable ras p21 suggesting that factors other than cell cycle may influence its expression. No reactivity with DWP was noted in any of the tissues, suggesting that the mutated forms detected by this monoclonal antibody are not expressed during normal human embryogenesis. These data show that there is regulated expression, and broad distribution of this gene product in normal developing human fetal tissue.

Immunohistochemical demonstration of lacto-N-fucopentose III in lung carcinomas with monoclonal antibody 624a12

Bronchopulmonary carcinomas were analyzed immunohistochemically using monoclonal antibody 624A12. The antibody was raised against a human small cell carcinoma cell line NCI-H69. It recognizes a particular sugar sequence in lacto-N-fucopentose III, which is preserved in formalin fixed and paraffin embedded tissue. Various bronchopulmonary carcinomas revealed characteristic patterns of immunoreactivity. Forty nine/50 adenocarcinomas were immunoreactive either diffusely or focally. The immunostaining was usually limited to the cell membranes with occasional intracytoplasmic immunostaining in large cells. The only negative case had been irradiated before surgical resection. Twenty seven/38 squamous cells carcinomas did not immunostain while the remaining 11 displayed focal immunoreactivity in areas of loose cellular apposition associated with necrosis and, rarely, in squamous pearls. All of six adenosquamous carcinomas showed immunoreactivity focally. Eleven/30 large cell carcinomas and 10/11 bronchiolo-alveolar carcinomas were either diffusely or focally immunoreactive. Seven/26 intermediate cell neuroendocrine carcinomas were focally immunoreactive while none of 33 typical small cell neuroendocrine carcinomas, 21 carcinoids, and 10 well differentiated neuroendocrine carcinomas was immunoreactive. An adenoid cystic carcinoma was diffusely immunoreactive, and a mucoepidermoid carcinoma was focally immunoreactive. We conclude that various bronchopulmonary neoplasms have characteristic patterns of distribution of this antigen, and that monoclonal antibody 624A12 may be useful for the differential diagnose among bronchopulmonary carcinomas, and their differential diagnosis from pleural mesotheliomas.