Kouji Hashimura

Immunoreactivity of proliferating cell nuclear antigen in salivary gland tumours: An assessment of growth potential

Immunoreactivity of proliferating cell nuclear antigen (PCNA) was assessed to evaluate growth potential in surgically resected tissue specimens from 70 cases of benign and malignant salivary gland tumours. Three stage streptavidin-biotin immunoperoxidase immunostaining using monoclonal antibody to PCNA showed a heterogeneity of PCNA index and distribution. In normal salivary gland specimens, PCNA was demonstrated in the nuclei of few ductal and acinar cells. In pleomorphic adenoma a multiple nodular growth pattern was observed with positive immunoreactivity restricted to the nuclei of tubulo-ductal structures. Warthins tumour had positive nuclei in the outer cuboidal cells of epithelial component and germinal centres of lymphoid tissue. Myoepithelioma and acinic cell carcinoma showed slightly differing values and a statistically significant difference in the value of the index was observed in tumour cell aggregates of the cribiform type of adenoid cystic carcinoma and the solid undifferentiated type and between low/intermediate and high-grade mucoepidermoid tumours. PCNA is a useful marker of tumour cell proliferation; the index correlates with the grade of malignancy in salivary gland tumours.

Prenatal development of human major salivary glands and immunohistochemical detection of keratins using monoclonal antibodies

The major salivary glands were examined from 69 human fetuses ranging from 10 to 40 weeks of gestation. Prenatal growth curves of developing salivary glands could be established by histological scoring, and development was divided into the early developmental stage (EDS) from 10 to 18 weeks, early intermediate developmental stage (EIDS) from 19 to 24 weeks, late intermediate developmental stage (LIDS) from 15 to 32 weeks, late developmental stage (LDS) from 33 to 40 weeks. Characteristic morphogenesis and cytodifferentiation occurred in glandular duct cells during the period of EIDS and LIDS. In the LDS, acini and ducts of the salivary glands histologically developed into a mature state similar to adult glands. Immunohistochemical staining with monoclonal antibodies (MoAbs) PKK1, KL1, K8.12, K8.13, K4.62, RPN 1160, 1162, 1163, 1164, and 1165 was performed. During the fetal period, keratin expression as revealed by MoAbs PKK1, KL1, K8.12 was well established, and the staining pattern for each of these antibodies was comparable. Other antibodies showed rare or negative staining except K8.13 which had a diffuse, non-specific staining pattern. Accordingly, the proliferation and cytodifferentiation of fetal stage keratin staining in ductal cells as revealed by MoAbs PKK1, KL1, and K8.12 showed a heterogenic distribution in both luminal and basal cells. It is a characteristic finding that the cytodifferentiation of ductal luminal cells precedes ductal basal cells. Ductal basal cells stained with MoAb K8.12 and show heterogeneity of keratin distribution continuously until the full term of gestation. The keratin staining of oral epithelium was also examined to compare with distribution of salivary gland ductal cells and oral epithelial cells. In the present study, the developmental sequence of salivary gland cells and the immunohistochemical properties of keratin proteins in these cells were described in relation to the histogenesis of salivary gland tumours.

Immunohistochemical Detection of S-100, S-1OOα, S-100β Proteins, Glial Fibrillary Acidic Protein, and Neuron Specific Enolase in the Prenatal and Adult Human Salivary Glands

Developing human fetal salivary glands of gestational age from 10 to 40 weeks (n = 100) and normal adult glands (n = 10) were examined for immunoreactivity to S-100 protein and its subunits S-100 alpha, S-100 beta, glial fibrillary acidic protein (GFAP) and neuron specific enolase (NSE). In the early intermediate developmental stage (19-32 weeks) some acinar basal cells showed immunoreactivity to S-100 protein which rapidly disappeared in the late developmental stage (33-40 weeks). Adult salivary glands were negative for S-100 protein. The S-100 alpha subunit was strongly positive in the glandular ducts and acini of both fetal and adult glands. The S-100 beta, although present in some acini and ductal cells during the late intermediate developmental stage, was rarely seen in the adult glands. GFAP and NSE was positive at the developing salivary epithelium in the early developmental stage (15-18 weeks). The above findings indicated that the developing salivary epithelia showed transient appearance of the neuronal phenotype during active cytodifferentiation stage of glandular acini and ducts. Therefore, after evaluation of normal developmental and neoplastic transformation of the salivary glands a suggestion that neuronal differentiation of ductal reserve cells is responsible for the production of modified myoepithelial cells in both normal developmental salivary gland and neoplastic transformation is made.

Growth pattern of experimental squamous cell carcinoma in rat submandibular glands—An immunohistochemical evaluation

Histopathological and immunohistochemical studies during carcinogenesis in rat submandibular glands (SMGs) using a carcinogen (9,10-dimethyl-1,2-benzanthracene: DMBA) were evaluated. For carcinogenesis, the carcinogen-containing sponge was surgically inserted into the gland. Histopathological features during carcinogenesis were as follows; dilatation of ductal segments, the presence of duct-like structures and cystic lesion around the sponge were observed within 3 weeks of the experiment, squamous metaplasia in duct-like structures and lining epithelium of the cystic structures around the sponge were observed at 4-6 weeks of the experiment, and finally well differentiated squamous cell carcinomas (SCCs) were observed after 8 weeks of the experiment. The immunoreactivity of K8.12 keration (K8.12), S-100 protein (S-100), epidermal growth factor (EGF), laminin, and proliferating cell nuclear antigen (PCNA) were evaluated. In the normal SMG, EGF was confined to the granular cells and S-100 to the pillar cells of granular convoluted tubules (GCTs). K8.12 was found in striated (SD) and excretory duct (ED) cells and laminin showed linear staining of the basement membrane around the ducts, acini and blood vessels. PCNA-positive nuclei were rarely observed in the normal glandular parenchyma. During carcinogenesis, during the first stage, EGF in granular cells and S-100 in pillar cells of GCT segments disappeared, and cytokeration K8.12 was observed in duct-like structures and cystic epithelium around the DMBA sponge. PCNA-positive nuclei in the first stage were mainly confined to basal cells of morphologically altered ducts. During the second stage, squamous metaplastic cells showed an intense K8.12 reaction. During the third stage, the well differentiated SCC showed strong reaction for K8.12, and the linear staining for laminin staining had disappeared at the invading fronts. The PCNA index was nearly 40% in the tumour cell component. The stem cells or the progenitor cells during experimental carcinoma were most likely to be the ductal basal cells, and carcinogenesis was initiated with an increase of proliferating activity in small cell clusters surrounding a necrotic area, basal cells of dilated excretory ducts and duct-like structures. Thus, all ductal segments undergoing squamous metaplasia may participate in the genesis of neoplasia during experimental carcinogenesis.

Comparison of proliferating cell nuclear antigen index in benign and malignant salivary pleomorphic adenoma

The expression of proliferating cell nuclear antigen (PCNA) was studied in benign and malignant pleomorphic adenomas by using monoclonal antibody to PCNA. Carcinoma in pleomorphic adenoma (n = 8), cell-rich variant (n = 6) and typical pleomorphic adenoma (n = 6) were selected in this study. The PCNA index in carcinoma in pleomorphic adenoma showed a higher index of nuclear staining (mean 22.9%, S.D. 6.2) than in typical pleomorphic adenoma (mean 6.9%, S.D. 3.4) or a cell-rich variant of pleomorphic adenoma (mean 8.8%, S.D. 3.3). A significant difference in PCNA index was found between benign and malignant pleomorphic adenoma (P < 0.05). The present study suggests that PCNA index significantly differs between pleomorphic adenoma and carcinoma in pleomorphic adenoma, but in the prediction of malignant transformation potential it should be combined with routine histopathological examination.

Immunohistochemical localization of lysozyme, lactoferrin, al-antichymotrypsin, and α1-antichymotrypsin, and α1-antitrypsin in salivary gland of human fetuses

Summary 26 human fetuses were examined to elucidate the immunohistochemical distributions of lysozyme, lactoferrin, α 1 -antichymotrypsin, and α 1 -antitrypsin in prenatal salivary glands. Development of fetal salivary glands was divided into 4 stages: The early developmental stage (EDS) , the early intermediate developmental stage (EIDS) , the late intermediate developmental stage (LIDS) , and the late developmental stage (LDS) and were used to compare antigen localization during salivary gland development. Lysozyme (LY) staining was prominent in serous or demilune cells of the mucous acinar compartment. Lactoferrin (LF) was rarely seen in the fetal glands; only trace amounts were seen in serous cells. α 1 -antichymotrypsin (α 1 - ACT ) was diffusely positive particularly in glandular ducts. α 1 -antitrypsin (α 1 - AT ) was also diffusely distributed in all salivary gland elements and was more abundant in ductal cells than acinar cells. During the EDS, immunohistochemical staining of LY, LF, α 1 -ACT, and α 1 -AT could be observed with glandular intensity increases corresponding to the advance of cytodifferentiation of granular epithelium occuring in the subsequent EIDS and LIDS. Staining intensities were continuous during the LDS even though the amount of those materials in the fetal salivary glands was not of the extent seen in the adult salivary gland. These results suggest that production of LY, LF, α 1 -ACT, and α 1 -AT was positive during prenatal development of human salivary glands. The present study discusses the protective roles and defense mechanisms of LY, LF, α 1 -ACT, and α 1 -AT in developing human salivary glands.

Vimentin expression in sweat gland tumours

Immunohistochemical localization of vimentin was studied in 93 cases of sweat gland tumours using a monoclonal anti-vimentin antibody. A strong immunoreactivity of vimentin was observed in modified myoepithelial or neoplastic myoepithelial cells of mixed tumour of the skin, syringoma, and sweat gland adenoma. Tumour cells in outer layers of tubular, ductal, and duct-like structures usually showed positive staining for vimentin, which coincided with modified myoepithelial cells. All tumour cells of clear cell hydroadenoma showed positive vimentin staining. Tumour cells of the luminal border of tubulo-ductal structures of mixed tumours were rarely immunoreactive for vimentin. Positive vimentin staining of tumour cells in the outer zone of tubulo-ductal structures in sweat gland tumours may be related to reactive proliferation of modified myoepithelial cells and simultaneous growth of luminal tumour cells.