The paper investigates, by electron microscopy, the mechanism of regeneration of Clara cells killed selectively and completely by bromobenzene (BB) in mouse terminal bronchioles. Male specific pathogen free mice received an intraperitoneal injection of 0.76g/kg BB. Within 24hr, Clara cells showed vacuolization of the massive smooth endoplasmic reticulum (SER) followed by severe cell damage, and all Clara cells then exfoliated into the airway. Only ciliated cells remained intact; 99.9% of the lining cells were ciliated cells, which flattened and covered all of the terminal bronchioles. Many ciliated cells then lost the cilia and basal bodies; concurrently, nonciliated cells occurred although cell proliferation was negligible. Between 3 and 5 days, these nonciliated cells divided and increased in number, but some of them still maintained a few basal bodies, strongly suggesting that they are “post-ciliated” cells. At the same time, some cells showed ciliogenesis to replenish the decreased ciliated cell population, and at day 5 some formed a cluster of SER. After eight days, granules were formed in the SER-containing cells, and resulted in Clara cell development. Cycloheximide did not affect the reservation of ciliated cells but spoiled the reappearance of Clara cells during the bronchiolar regeneration after BB injection. At day 2 when Clara cells were thoroughly destroyed, [³H]-thymidine was incorporated into some ciliated cells, which transformed into nonciliated cells at days 3 and 4. Therefore, the present results may prove that a bronchiolar regeneration mechanism involves dedifferentiation of ciliated cells to form nonciliated cells, from which columnar epithelium regenerates.

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An immunohistochemical study was made of the expression of c-myc protein, p53, c-erbB-2 protein and epidermal growth factor receptor (EGF-R) in human normal thyroid and thyroid tumors including follicular adenoma and follicular, papillary, squamous cell, anaplastic and medullary carcinoma. Immunoreaction product for c-myc protein was located in the cytoplasm as well as nucleus of follicular cells in normal thyroid and neoplastic cells of all tumors. Semiquantitative analysis using serial dilutions of the antibody showed significant differences in immunoreactive cytoplasmic c-myc protein between normal thyroid and thyroid tumors. Immunoreaction product for p 53 was noted in the cytoplasm of follicular cells in normal thyroid and neoplastic cells of all tumors, however, reaction product was absent in the nucleus for all cases of papillary and anaplastic carcinoma and 8 out of 10 cases of squamous cell carcinoma. On the other hand, normal thyroid and other types of thyroid tumor exhibited positive nuclear reaction for the antibody in about 50% of the cases. Immunoreaction product for c-erbB-2 protein and EGF-R was seen as either diffuse or granular deposits in the cytoplasm of the follicular cells in normal thyroid and neoplastic cells in all tumors, except for negative results in the immunostaining for EGF-R in normal thyroid. In immunostaining for these two antigens, thyroid tumor showed a significantly higher positivity than normal thyroid, and thyroid carcinoma showed a significantly higher incidence of positive cases than that of follicular adenoma, especially in immunostaining for EGF-R. These results show that cytoplamic c-myc protein, c-erbB-2 protein and EGF-R are enhanced in many cases of thyroid tumor, and EGF-R tends to increase considerably in thyroid carcinoma. It is also suggested that nuclear p53 might bear some relation to the differentiation of thyroid tumors.

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Neurons containing the enzyme aromatic L-amino acid decarboxylase (AADC) but lacking tyrosine hydroxylase, dopamine and serotonin were found for the first time in the cortex and caudoputamen (CP) of 1 month-old mice by light and electron microscopic immunocytochemistry. These AADC-like immunoreactive cells appeared from postnatal (P) 12 days, reached maximum in number at P30 days and reduced cell numbers till the adult stage. The majority of these cells in the cortex localized in layers III∼IV. On the other hand, no particular topography was found in the distribution of these cells which were clustered throughout the CP. Ultrastructurally AADC-like immunoreactivity was found in the thin neuronal cytoplasm but not in the nucleus, Golgi apparatus, mitochondria, nor glia.

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A total of 3174 lymph nodes obtained from 156 cases of neck dissection and 31 cases of parotidectomy were evaluated for presence of heterotopic salivary gland tissue. The intranodal salivary tissue was found in 19 cases of neck dissection and 21 cases of parotidectomy; and they were more frequent in the intraparotid (67.7% cases) than in the extraparotid cervical nodes (12.2% cases). Salivary gland heterotopia in the lymph nodes histologically showed mature acini, small ducts, striated ducts, immature ducts and acini similar in structure to those seen in fetal salivary glands with proliferating ductal structures (22.7%), oncocytic metaplasia (10.6%), hyperplasia of oncocytes with cyst formation (10.6%), cyst formation in ductal epithelia (16.7%) and sebaceous metaplasia (0.5%). Monoclonal antibody to cytokeratins PKK1 and KL1 staining was positive in luminal ductal cells. Cytokeratin K8.12 reactivity was positive in both the luminal and basal ductal cells in contrast to negative luminal ductal cells in extra nodal salivary ducts. Immunoreactivity of epithelial membrane antigen (EMA), lysozyme (LY), lactoferrin (LF), alpha₁-an-tichymotrypsin and alpha₁-antitrypsin was irregular. On the basis of histological and immunohistochemical findings, the authors conclude that salivary tissues in lymph nodes are immature gland type, have potential for cytodifferentiation and various lymphoepithelial lesions may arise from these heterotopic salivary gland tissues.

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A specific antibody against purified human liver guanase was raised in rabbits. The antiserum formed a single precipitin line with human liver extracts, and completely inhibited the activity of the enzyme derived from human liver and kidney. Immunoblotting showed that the antibody bound only to the one protein band of liver and kidney guanase activity. In immunohistochemical studies, the DAB (3, 3′-diaminobenzidine tetrahydrochloride)-reaction with this antibody was positive in the same locations as the histochemical guanase reaction. Moreover, biochemical data of enzymic activity in nephron segments of the rat indicated that guanase activity was located in the proximal tubules. Thus, the DAB reaction appeared to show the location of guanase itself. This immunohistochemical procedure for detection of guanase should useful in clinical studies on this enzyme.

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The purpose of this study was to show the immunohistochemical activity of the commercialized antibodies of oncogene products and related enzymes using extremely small lesions of tubular adenocarcinomas in human stomachs. Seven lesions among 11 carcinoma lesions (63.6%) measuring 1mm in diameter and 8 among 8 carcinoma lesions (100%) measuring 1∼5mm in diameter showed positive reaction in their cytoplasms for c-myc oncogene products (6E10). In immunohistochemical staining for c-erbB-2 gene product no carcinoma lesions (0%) measuring 1mm in diameter were positive in their cytoplasms and all carcinoma lesions (100%) measuring 1∼5mm in diameter were positive in their cytoplasms. One lesion (1/11 lesions, 9.1%) measuring 1mm in diameter and 6/8 lesions (75%) showed positive reactions in their cytoplasms for protein kinase C (Type III) immunohistochemically (x² analysis, p<0.05). All gastric minute tubular adenocarcinoma lesions in this study were positive in their cytoplasms for lysozyme immunohistochemically.

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Lectin histochemical studies were performed on frozen sections of biopsied skeletal muscle from two unrelated patients with lysosomal glycogen storage disease without acid maltase deficiency. The clinical manifestations of these patients, myopathological features and biochemical characteristics are similar to those first reported by Danon et al. in 1981. Routine myopathological findings showed vacuolar myopathy and excess of glycogen with or without glycogenosomes resembling acid maltase deficiency (AMD), but the biochemical activity of acid α-glucosidase was normal.
Ten lectins with varying sugar specificities were used as probes. The results were compared with normal muscle and with one patient who had the typical manifestations of adult onset AMD. Though myofibers themselves were never stained in normal muscle, positively stained myofibers with or without vacuolations were found in diseased muscle. Vacuolar membranes and their contents were strongly stained by almost all the lectins applied. Ulex europaeus agglutinin-I (UEA-I) specific to L-fucose stained the perimeter of the disturbed muscle in addition to the vacuoles.
The staining pattern of the vacuolations and perimeter of disturbed myofibers was different from that found in AMD. In AMD, Triticum vulgaris agglutinin (WGA) and Limax flavus aggutinin (LFA) faintly stained the vacuolar membranes and contents. These preliminary investigations of abnormal lectin staining patterns may provide a key to understanding the pathomechanism of this unique lysosomal glycogen storage disease.

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During pre-and postnatal development, the atrial natriuretic peptide (ANP)-granules in the right and left auricular cardiocytes of Syrian hamsters were studied immunohistochemically and by transmission electron microscopy. Moreover, the number and diameter of ANP-granules in the cardiocytes were analyzed by ultrastructural morphometry. Immunohistochemically, the ANP immunoreactivity was located primarily in the perinuclear region in auricular cardiocytes of the neonatal group, but was located diffusely in the cells of the fetal group. At the 15th day of gestation, the immunoreactivity was stronger than at the 13th day of gestation. The ANP immunoreactivity observed in the cardiocytes of the newborn was weaker than in the fetal group. During the postnatal growth period, the immunoreactivity became gradually stronger. Ultrastructurally, ANP-granules in the fetal cardiocytes were scattered in the cytoplasm with immature organelles. During the postnatal growth period, the granules were predominantly located in the perinuclear region associated with the Golgi apparatus. By ultrastructural morphometry, the number of granules in the right auricular cardiocytes was significantly greater than in the left auricular cardiocytes in all groups examined. In the newborn, the number of granules was significantly less than that in the fetal group. During the postnatal growth period, the granule number of cardiocytes significantly increased. There was no significant difference between the granule number of 10-day-old neonates and of 90-day-old neonates. During the postnatal period, the granule diameter in both auricles was significantly greater than in the fetal group. The granule size was not significantly different between the right auricular cardiocytes and the left ones in all groups examined. It is suggested that a large amount of ANP-granules is secreted at the perinatal period, resulting in granule depletion in both auricular cardiocytes.

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Localization of E-cadherin in the intact and wounded mouse corneal epithelium was examined by immunoelectron microscopy of ultrathin frozen sections. In the intact epithelium, immunogold particles for E-cadherin were found predominantly adjacent to desmosomes and gap junctions and hardly in the non-junctional plasma membrane. Six hr after artificial wounding, a majority of the labeling became independent of the junctions; immunogolds were seen on both sides of the widened intercellular space or bridging between the facing plasma membrane. The present results indicate that E-cadherin mediates cell-cell contact in conjunction with desmosomes and gap junctions in the normal epithelium; dissociation of E-cadherin from the junctions may be necessary for their loosening and cell migration in the injured epithelium.

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