p27Kip1 (p27) is a member of the cyclin dependent kinase inhibitor (CDKl) family of cell cycle proteins. We analyzed p27 ex-pression in normal endocrine cells, endocrine cell hyperplasia, adenomas and carcinomas. The levels of p27 protein expression decreased during tumor development and progression. This decrease occurs mainly at the post-translational level with protein degradation by the ubiquitin-proteasome pathway. These results indicate that p27 may be a useful immunohistochemical marker to help separate normal from hyper-plastic endocrine cells and adenomas from carcinomas.
Recent technological development has disclosed various transcription factors in the pituitary glands. The cloning of Pit-1, homeodomain containing nuclear bindi-ing transcription factors stimulated the subsequent discoveries of many factors participating in the transciption for the specific pituitary hormones. These include Pituitary homeobox1 (Ptx1), Prophet of Pit-1 (Prop-1), NeuroD1, steroidogenic factor-1 (SF-1) and DAX-1. These factors have been shown not only to interact but also to func-tion with various receptors synergistically to promote specific hormones, such as GH-PRL-TSH group, POMC and gonadotropin (FSH/LH).
Thyroid gland function is thought to be tightly regulated by thyrotropin (TSH). However, the function of each thyroid follicle is heterogeneous despite having the same blood supply of TSH and despite homogeneous thyrocyte expression of the TSH receptor (TSHR). The nature of this heterogeneity is not fully understood. Recent studies showed that thyroglobulin (TG) protein, stored in the follicular lumen, is a potent negative feedback regulator of follicular function. Thus, physiological concentrations of TG significantly suppress thyroidspecific gene expression in cultured thyrocytes and antagonizes the maximal TSH stimulation of thyroid-specific genes: thyroglobulin, thyroid peroxidase, sodium iodide symporter and TSH receptor. In vivo studies are consistent with these results. This regulation is mediated by TG suppression of thyroid-specific transcription factors: thyroid-transcription factors 1 and 2 as well as Pax-8. We propose a model of follicular activity wherein each follicle has its own cycle of thyroid hormone synthesis and secretion and wherein follicular heterogeneity reflects the asynchronous function of individual follicles. We provide evidence that this mechanism may be related to the phenotype of some thyroid diseases.
Adult rat chromaffin cells proliferate at a low rate throughout life and show robust mitogenic responses to a variety of agents in vivo and in vitro. Prolonged treatment of rats with some of these agents leads to development of pheochromocytomas, probably by exacerbating an age-dependent proclivity. The rat adrenal medulla is thus a remarkable model for studies of mitogenic signaling and neoplastic progression. Immunohistochemical approaches to the study of this model have helped to overcome obstacles presented by heterogeneity of the chromaffin cell population and complex relationships between medulla, cortex and nerve supply. Current evidence suggests that these relationships are important both in normal chromaffin cell turnover and in development of pheochromocytomas. The prototypical mitogen for adult rat chromaffin cells in vitro is nerve growth factor (NGF), which, parodoxically, arrests proliferation and causes neuronal differentiation of rat pheochromocytoma cells. Cell culture studies suggest that activation of phosphatidyl inositol 3-kinase by NGF is a critical determinant of mitogenic specificity in normal chromaffin cells. Numerous unanswered questions concern differences between normal chromaffin cells and their neoplastic counterparts, differences between chromaffin cells of rats and other species, and influences of the in vitro environment on responses to trophic signals.
Ovarian cell death is an essential process for the homeostasis of ovarian function in humans and in other mammalian species. It ensures the selection of the dominant follicle and the demise of an excessive number of follicles. Degeneration of the old corpora lutea also has a characteristic of programmed cell death. Cross-talk between endocrine, paracrine and autocrine factors as well as between protooncogenes, tumor-suppressor genes, survival genes and death genes can determine the fate of the ovarian somatic and germ cells. Establishment of highly transformed human and rat granulosa cells enables detailed studies on the role of the p53 tumor suppressor gene, cyclic AMP (cAMP), growth factors and extracellular matrices in controlling growth and apoptosis in experimental ovarian cells. It is hoped that stimulation of apoptosis in ovarian cancer cells will become one of the strategies in the treatment of ovarian cancer.
In the gastrointestinal tract, Class III mucinsecreting cells include the cardiac mucous cells, mucous neck cells, pyloric gland cells and Brunner's gland cells. It has been found that these cells are stained by a recently developed monoclonal antibody, HIK 1083, at the light microscopic level. In the present study, we compared HIK 1083 staining with paradoxical Concanavalin A type III staining (PCS) and GSA-II lectin staining, and found that all three methods revealed the same reactivity at both the light and electron microscopic levels. HIK 1083 staining exhibited highly selective reactivity with cardiac mucous cells, mucous neck cells, pyloric gland cells and Brunner's gland cells. These results indicate that mucus-secreting cells of foregut origin share a common lineage and contain the same cell phenotype. Class III mucin secreted by these cells was characterized by the presence of a peripheral α-GlcNAc residue on the carbohydrate moiety of its constituent glycoproteins. Because of the ease and specificity of immunostaining using monoclonal antibody HIK 1083, it could replace PCS staining for both light and electron microscopy.
Proteoglycans are one of the major constituents of the extracellular matrix and cell membranes. In the brain, there are two major proteoglycans, chondroitin sulfate proteoglycans and heparan sulfate proteoglycans. In the adult mammalian central nervous system, some species of chondroitin sulfate proteoglycan are localized in the ‘perineuronal net’ around a restricted number of neurons. The ‘perineuronal net’ is a reticular structure covering the cell bodies and proximal dendrites of certain neurons. Various glycoproteins, proteoglycans and hyaluronic acid are constituents of the perineuronal nets. Recently, an N-terminal proteolytic fragment of neurocan named neurocan-130 was found to be a member of the perineuronal net-constituting molecules. Neurocan is a nervous tissue-unique chondroitin sulfate proteoglycan whose expression and proteolytic cleavage are developmentally regulated. Most of the perineuronal netconstituting molecules are considered to extracellularly exist and some are considered to be localized at the surface of glial cell processes enwrapping the neural cell body. However, neurocan-130 was detected in the cytoplasm of the glial cell processes. Perineuronal nets could be involved in synapse stabilization or neuronal maturation because they appear in the vicinity of the synapses relatively late in neuronal development. The presence of perineuronal nets around a limited number of cells may reflect some functional heterogeneity of the neuron and/or glia.
Immuno-molecular techniques specifically covers to the recent development of immunohistochemistry (IHC) and in situ hybridization (ISH). Particularly, in this review article, IHC emphasizes heat induced epitope retrieval (HIER) and tyramide (catalyzed) signal amplification (TSA or CSA) and ISH; focuses on peptide nucleic acid (PNA). The TSA (or LSA) is the reaction with marked amplification and has been useful in detecting trivial amounts of antigens. It is also applicable to ISH. PNA technology has been reported to be a stable detection method for RNA such as EBER. These methods have contributed to diagnostic pathology in the aspects of achievement and control of quality.
Immunohistochemical analysis can help determine the primary site of origin of carcinomas presenting at metastatic sites. Different types of epithelium, and their corresponding carcinomas, express different subsets of cytokeratins. Thus, antibodies to unique cytokeratins, such as “high molecular weight” cytokeratins, cytokeratin 7, cytokeratin 8, and cytokeratin 20, can help distinguish among different types of carcinomas that may have similar histologic appearances. Also helpful in this analysis are antibodies to the CEA family of proteins, as carcinomas of different sites vary widely in the quantitative and qualitative expression of these proteins. Another approach is the use of antibodies to vimentin, the intermediate filament protein that is co-expressed along with cytokeratin in a limited subset of carcinomas. Additionally, determination of primary site of origin of carcinomas can be assisted by the use of organ-associated markers such as prostatic specific antigen (prostate), gross cystic disease fluid protein-15 (breast), mesothelin (ovary, mesothelium), and thyroid transcription factor-1 (lung, thyroid). Selection of the appropriate subset of these antibodies can be most helpful in clinical situations where there is a limited set of possible primary sites (e. g., breast vs. lung or bladder vs. prostate).
Analysis of tissue turnover, the balance between cell proliferation and cell death, can provide important information about our understanding of the biological characteristics of various tissues and their functions. Especially it is very important to study apoptosis, one of the modes of cell death, in surgical pathology specimens. It is very important to note that the whole concept of cell death including apoptosis and oncosis is based on the morphological findings. Therefore, an analysis of morphological features of the cells undergoing the processes of various modes of cell death is very important. DNA fragmentation can be detected in situ by labeling 3′-OH ends with biotinylated deoxyuridine triphosphate (dUTP) through the action of terminal deoxynucleotidyl transferase (TdT). Since then, this method, subsequently termed 3′-OH nick end labeling, or TdT-mediated dUTP-biotin nick end labeling (TUNEL), has been widely used to detect cells with DNA fragmentation, especially in surgical pathology materials of human disorders. This TUNEL method is now being incorporated into many pathology laboratories throughout the world. However, increasing evidence suggest that TUNEL method is by no means specific for the detection of apoptosis and this method may also detect cells which are committed to, but not yet in the process of apoptosis. Therefore, it is very important to correlate the findings of TUNEL with morphological features of surgical pathology specimens. In addition, it is also important to note that over-fixation is associated with false negative results and insufficient fixation is likely to be associated with false positive results. Therefore, the processing of the specimens including fixation and others is critical in performing TUNEL and its interpretation.
Application of immunohistochemical methods has greatly contributed to diagnostic thyroid pathology, especially to the diagnosis of thyroid tumors. These techniques can demonstrate specific markers for follicular and C (parafollicular) cells and their tumors. Therefore, they facilitate defining the spectrum of thyroid tumors, and also are useful for the distinction between tumors of different histogenesis. In this paper, we reviewed thyroid specific proteins and compounds as immunohistochemical markers, i. e. thyroglobulin (TG), thyroxine (T4), triiodothyronine (T3), thyroperoxidase (TPO), TSH receptor (TSHR), Na/I symporter (NIS), thyroid transcription factor-1 (TTF-1), and calcitonin. In addition, we also reviewed intermediate filaments and other proteins having a considerable diagnostic value in thyroid tumor pathology.
Since immunohistochemical procedures started to be used in the field of tumor pathology, they have been actively applied to the study of soft tissue tumors as techniques which remove much of the difficulty involved in histological diagnosis. A number of reports are available to show that these procedures have successfully yielded objective findings which allow for diagnostic agreement in many cases. Another opinion is that what is most important in the diagnosis of soft tissue tumors is an accurate determination of the morphological characteristics, making it doubtful that immunostaining would be performed unnecessarily because the accurate determination of morphological characteristics allows for the diagnosis of almost all soft tissue tumors. Clearly, the point that it is important to determine morphological characteristics is quite right. It is also a fact that many soft tissue tumors can be diagnosed without immunostaining. However, there are also a large number of patients for whom definite diagnosis can be made only after immunostaining and in whom reconfirmation by immunostaining is needed before a final diagnosis is made.
γ-Aminobutyric acid (GABA) immunoreactivities were found by immunohistochemistry in the chemoreceptor cells of the cat carotid body at the light microscopic level. The immunohistochemical process used consisted of a strept-avidin-biotin-peroxidase method. The immunostaining was mostly weak and appeared in about one-third of the cells. The scarcity of GABA immunoreactivities provides an explanation for the previously reported meager interactive effects on respiration of the elaboration of the GABA-ergic and chemoreceptor systems found in the cat at the functional level. Therefore, GABA would be an unlikely essential modulator of the chemosensory function in the cat.
The objective of this study is to confirm the relationship between expression of cytokeratin no. 8 (CK8) and types of human papillomavirus (HPV) DNA in cervical squamous intraepithelial lesions (SIL). Expression of CK8, and types of HPV DNA as 6/11, 16/18 and 31/33/51 were investigated in normal and SIL tissues by immunohistochemistry and in situ hybridization techniques. CK8 positive staining was detected in 68 (51.9%) out of 131 tissues of HPV DNA positive (p<0.01). The located patterns of CK8 were A-type (only basal layer), B-type (basal to intermediate layers), and C-type (all layers). In 54 tissues of HPV 16/18 types, there were 20 (37%) tissues that showed C-type (p<0.001). In HPV 6/11 and 31/33/51 types, CK8 positive staining was mostly shown in A-type. The malignant transformed tissues were detected as HPV 16/18 types and CK8 C-type. HPV 16/18 types promote expression of CK8. Tissues expressed as C-type with HPV 16/18 types signify abnormal cytodifferenciation and are probably real pre-malignant lesions. The detection of CK8 and HPV DNA types are useful for prediction of malignant transformation.