Characterization of Leptospira strains HY-1, HY-2, and HY-10 isolated in Korea by means of monoclonal antibodies and restriction endonuclease DNA analysis.

To analyze the characteristics of Leptospira strains HY-1, HY-2, and HY-10, which were isolated from patients with leptospirosis in Korea in 1985, 12 monoclonal antibodies (MAbs) against strain HY-1 and six MAbs against serovar lai strain 017 were produced, and their properties were determined by the microscopic agglutination test. Genetic relationships among the leptospires were determined by restriction endonuclease DNA analysis. Three MAbs reacted with all strains of serogroup Icterohaemorrhagiae, but did not react with any strains of the other 10 serogroups. All MAbs reacted with strains 017, HY-1, HY-2, and HY-10 at nearly identical titers. Two MAbs reacted only with these four strains. These four strains also had the same restriction endonuclease cleavage patterns. Based on these results, strains HY-1, HY-2, and HY-10 were identified as serovar lai, which is one of the common serovars in China. It is suggested that serovar lai is one of the prevalent serovars in Korea, and that the Mabs produced in this study are useful for the accurate and rapid identification of this serovar.

Increased polysomy 3 and 17 detected by fluorescence in situ hybridization (FISH) in oral squamous cell carcinoma.

Numerical chromosome aberrations were studied by fluorescence in situ hybridization (FISH) using pericentromeric DNA probes specific for chromosomes 3 and 17 in 18 oral squamous cell carcinomas (SCCs) and 3 lymph nodes as control. Disomy 3 and 17 was detected in approximately 90% of control cells, and in 60.9 +/- 4.0% and 62.7 +/- 3.6% of SCCs, respectively. Polysomy 3 and 17 significantly increased in SCCs when compared to controls. The pattern of chromosome aberrations varied considerably between cases. There was no obvious relationship between the degree of polysomies and clinicopathological factors such as tumor-differentiation, stage of the disease and clinical outcome. Chromosome aberrations by FISH did not correlate with DNA aneuploidy by flow cytometry. Our results indicate that oral SCCs are more frequently associated with the increased copy number of chromosomes 3 and 17 than previously thought, and that a correlation between chromosome aberrations by FISH and DNA aneuploidy by flow cytometry is not obvious.

Immunohistochemical detection of p53 protein and proliferating cell nuclear antigen (pcna) in salivary-gland tumors.

The expression of p53 protein and PCNA was immunohistochemically examined in 161 cases of salivary gland tumors. The p53 protein was positive in 23.8% (24/101) of malignant salivary gland tumors, while only one case of 60 pleomorphic adenomas was positive. p53 protein was frequently detectable in salivary duct carcinoma (60%), basal cell adenocarcinoma (66.7%), acinic cell carcinoma (53.3%) and undifferentiated carcinoma (50%), but was rarely seen in adenoid cystic carcinoma (17.2%) and mucoepidermoid carcinoma (0%). p53 protein was more frequently detectable in PCNA positive cases than in negative cases (p=0.0074).

The estimation of proliferative activity by pcna and AgNORs in leukoplakia and squamous-cell carcinoma of the oral cavity.

The proliferating activity of leukoplakia and squamous cell carcinoma (SCC) was estimated using proliferating cell nuclear antigen (PCNA) immunostaining and silver-binding argyrophilic nucleolar organizer region (AgNOR) staining. Twenty-eight leukoplakias and 15 SCCs of the oral cavity were used in this study. The mean+/-S.D. of PCNA LI and AgNOR counts were 7.7+/-6.0% and 2.43+/-0.68/nucleus in leukoplakias, and 22.3+/-11.6% and 4.77+/-1.49/nucleus in SCCs. Both of PCNA LI and AgNOR counts were significantly higher in SCCs than in leukoplakias. There was a significant linear correlation between PCNA LI and AgNOR counts (p=0.0022) in leukoplakias, but not in SCCs. In the series of leukoplakias, PCNA LI apparently increased in leukoplakias with dysplasia and malignant transformed cases. Our data suggest that PCNA LI and AgNOR counts are useful markers of proliferating activity, and PCNA LI might be a prognostic factor of leukoplakias.

Immunohistochemical detection of human lung and gastric cancer antigen in human salivary gland tumors.

Immunohistochemical identification of human lung and gastric carcinoma antigen detected with monoclonal antibodies (MoAbs) KM-93 and KM-231 respectively, was described in 83 salivary gland tumors, including 67 pleomorphic adenomas, 5 adenolymphomas, 3 mucoepidermoid tumors, 6 sialadenocarcinomas, and 2 adenoid cystic carcinomas as well as in normal salivary glands. The binding patterns of these two MoAbs was compared with that of MoAb recognizing epithelial membrane antigen (EMA). Serous cells of normal salivary glands showed positive KM-93 staining, whereas ductal cells were positive with KM-231, with ductal basal cells being characteristically so. EMA staining was confined to luminal and lateral borders of serous acini and ducts. Pleomorphic adenomas indicated positive depositions for both KM-93 and KM-231 in luminal tumor cells or luminal borders of tubuloductal structures. Adenolymphomas showed positive KM-231 staining in basal tumor cells and a positive KM-93 reaction in luminal tumor cells. Mucoepidermoid tumors revealed positive KM-231 staining in mucous-secreting cells, whereas weak KM-93 staining was found in all tumor cells. Sialoadenocarcinomas exhibited varying degrees of positive staining with KM-93 and KM-231 in their neoplastic cells. Adenoid cystic carcinomas showed luminal staining with KM-93 and KM-231 to their neoplastic cells. Adenoid cystic carcinomas showed luminal staining with KM-93 and MoAb EMA. The histogenesis of these salivary gland tumors is discussed in terms of the immunohistochemical features of staining patterns obtained with MoAbs KM-93 and KM-231.

Chromosome 17 abnormalities in squamous cell carcinoma of the oral cavity, and its relationship with p53 and Bcl-2 expression.

Numerical aberrations of chromosome 17 were studied by fluorescence in situ hybridization (FISH) using pericentromere specific DNA probe in 27 oral squamous cell carcinomas (SCCs), and its relationship with p53 and Bcl-2 protein expression was investigated. Since cells with polysomy 17 are significantly increased in SCC (p = 0.0005), chromosome 17 abnormality seems to be correlated with carcinogenesis of oral SCC. Chromosome 17 abnormality varied from case to case, and the degree of the abnormality did not correlate with the stage of the disease, the histological differentiation of SCC, or relapse of the disease and lymph node metastasis. However, there was a correlation between polysomy 17 and p53 immunoreactivity (p = 0.0228). p53 immunoreactivity showed a correlation with relapse of the disease (p = 0.0441). An inverse relationship was found between p53 and Bcl-2 immunoreactivity (p = 0.0421). In conclusion, we suggest that polysomy 17 is closely related carcinogenesis of oral SCC, and with p53 overexpression. It is assumed that polysomy 17 correlates with the mutation of p53 which results in an accumulation of aberrant p53 protein, and that its activation causes an increase of p53 protein.

Detection and analysis of human papillomavirus 16 and 18 homologous DNA sequences in oral lesions.

The prevalence of human papillomavirus (HPV) 16 and 18 was investigated in oral lesions of the population of northeast China including squamous cell carcinomas (SCCs), candida leukoplakias, lichen planuses and papillomas, by southern blot hybridization with polymerase chain reaction (PCR). Amplified HPV16 and 18 E6 DNA was analyzed by cycle sequence. HPV DNA was detected in 14 of 45 SCCs (31.1%). HPV18 E6 DNA and HPV16 E6. DNA were detected in 24.4% and 20.0% of SCCs. respectively. Dual infection of both HPV 16 and HPV 18 was detected in 6 of 45 SCCs (13.3%), but not in other oral lesions. HPV 18 E6 DNA was also detected in 2 of 3 oral candida leukoplakias, but in none of the 5 papillomas. Our study indicated that HPV 18 infection might be more frequent than HPV 16 infection in oral SCCs in northeast Chinese, dual infection of high risk HPV types was restricted in oral SCCs, and that HPV infection might be involved in the pathogenesis of oral candida leukoplakia.

Numerical aberrations of chromosome 17 detected by FISH with DNA-specific probe in oral tumors.

Numerical aberrations of chromosomes were investigated in oral tumors by using fluorescence in situ hybridization (F1SH) with 4 chromosome centromere-specific DNA probes. The hybridization signals of the centromere sequence were easily detected and counted within interphase nuclei as well as metaphase chromosomes. The number of hybridized signals for chromosome 17 significantly increased in oral malignant tumors compared to benign tumors, which had two spots as well as normal lymphocytes. Cell lines derived from oral squamous cell carcinoma also showed numerical aberrations of chromosomes 1, 4 and 11, in addition to chromosome 17. These findings suggested that numerical aberrations of chromosomes occur during the development of malignant tumors of the oral cavity.

Immunohistochemical determination of growth fraction in human tumors.

The growth fraction in 93 cases of human tumors was estimated by an immunohistochemical staining using Ki-67. The tumors consisted of the following: 14 oral cancers, 14 breast cancers, 9 gastric cancers, 9 uterine cancers, 8 ovarian cancers, 6 colo-rectal cancers, 6 thyroid cancers, 5 esophagus cancers and 22 miscellaneous tumors. Regional labeled cells were predominantly found in the periphery of the tumor nests in squamous cell carcinomas. However, in adenocarcinomas the labeled cells were randomly distributed in tumor cell nests. The Ki-67 labeling index varied greatly from case to case (almost 0 to 50.9% with an average of 17.3%), even within the same organ group. The growth fraction was independent of the histological pattern, although thyroid cancers showed a lower labeling index than other malignant tumors. The labeling indices in benign tumors were lower than those in malignant tumors. The usefulness of this method for the estimation of biological behavior of human tumors is suggested.

Branched-chain amino acid aminotransferase in submandibular gland of the rat.

Crude extracts from submandibular glands of the rat, rabbit, guinea pig and macaque monkey contained high activities of branched-chain amino acid aminotransferase comparable to those from the heart and kidney. In each species, the isoelectric point and relative elution volume on Sephacryl of the aminotransferase from the submandibular gland had the same values as the heart enzyme. The enzyme of rat submandibular gland was partially purified; the properties of the purified enzyme suggest that it is the same type as that from the heart.

Analysis of major histocompatibility complex (MHC) class I antigen in the rat salivary gland.

This antigen was examined in rats of different ages (new-born, 3, 5, 7, 10, 12 and 14 days after birth and adult) by immunofluorescence and immunoelectron microscopy. Changes in each kind of salivary gland when graft versus host disease was induced in recipient rats were also investigated. Monoclonal antibodies (HAM 2 or OX 18) specific to rat MHC class I antigen were used and these were detected by FITC-conjugated anti-mouse immunoglobulin. With HAM 2, MHC class I antigen in the submandibular gland was mostly located in the secretory duct cells; this expression was first found 10 days after birth. The antigen was found on the cell surfaces of the secretory duct cells by immunoelectron microscopy. With OX 18, MHC class I antigen was mainly found in the secretory duct cells, but weak expression was also found in the acinar cells. Localization of the antigen, by HAM 2 and OX 18 was less evident in the secretory duct cells of parotid and sublingual glands. When graft versus host disease was induced, MHC class I antigen (HAM 2) was observed in both acinar and secretory duct cells of the submandibular gland.

An expression of squamous cell carcinoma-related antigen in oral cancers.

The relationship between serum level and stainability of squamous cell carcinoma-related antigen (SCC-Ag) in carcinoma of the oral cavity was examined. The amount of serum SCC-Ag was measured in 60 of 97 patients by a radioimmunoassay system. 63.3% showed elevated serum SCC-Ag levels above 2.0 ng/ml. The specimens taken from 97 patients with oral squamous cell carcinoma (SCC) were stained with a monoclonal antibody against the SCC-Ag by using an immunoperoxidase method. Tumor cells with keratinization in well-differentiated SCC stained well with the antibody, whereas cells without keratinization in poorly-differentiated SCC stained weakly, or not at all. Serum levels of the antigen were significantly higher in cases of invasive cancers, but serum levels of antigen correlated with neither the degree of morphological differentiation of the tumor, nor with the intracellular content of the antigen. These observations suggest that the parenchymalstromal relationship is one of the decisive factors in determining the serum SCC-Ag level.

The nucleolar organizer regions associated protein (Ag-NORs) in salivary gland tumors.

A silver colloid technique used to identify nucleolar organizer regions associated protein (Ag-NORs) has been applied to 20 salivary gland tumors. The method was readily applicable to the preparations of paraffin-embedded sections and the Ag-NORs were enumerated with ease. A significant difference was found between the numbers of Ag-NORs in the nuclei of malignant salivary gland tumors, such as adenoid cystic carcinoma, mucoepidermoid tumor and adenocarcinoma (with a mean of from 2.05 to 2.78 per nucleus) and those of benign salivary gland, such as pleomorphic adenoma, adenolymphoma (Wartin tumor) and clear cell adenoma (with a mean of from 1.47 to 1.72 per nucleus). It is proposed that the Ag-NORs technique, which is rapid, simple, and inexpensive, may be useful in the differential diagnosis of malignant and benign salivary gland tumors.

Measurement of proliferating cell nuclear antigen (PCNA) and its clinical application in oral cancers.

The PCNA score was measured in oral squamous cell carcinoma (SCC), and its relationship to other cell proliferation markers, Ki-67 score, S-phase fraction (SPF), and AgNORs counts was investigated. The PCNA score ranged from 0.4% to 43.5% with an average value of 22.8%, the Ki-67 score ranged from 4.9% to 40% with an average of 24.1%, and the SPF ranged from 0.4% to 32.5% with an average of 12.4%, while AgNORs counts ranged from 2.53/nucleus to 7.03/nucleus with an average of 4.74/nucleus. These four parameters were closely interrelated. There was a significant difference in PCNA score between malignant and nonmalignant lesions, suggesting a difference in growth activity. The mean PCNA score decreased significantly from 20.0% to 8.0% after cancer chemotherapy. The response of cancer cells to anticancer agents may be estimated by consecutive measurement of PCNA, since the PCNA score dropped after treatment in cases showing a favorable prognosis.

The significance of PCNA and p53 protein in some oral tumors.

The expression of PCNA and p53 protein was evaluated in a total of 75 cases of benign and malignant lesions of the oral cavity, comprising 50 squamous cell carcinomas (SCCs), 14 leukoplakias, and 11 pleomorphic adenomas. The DNA histogram of 20 SCCs was measured by flow cytometry. p53-positive cells were frequently seen in SCCs, but were rare in leukoplakias and pleomorphic adenomas. The PCNA labeling index (LI) was higher in SCCs than in other benign lesions. The expression rate of p53 protein was markedly elevated in SCCs obtained from smoking patients, when compared to nonsmoking patients. DNA ploidy did not show a close relationship with PCNA and p53 expression. The mean value of PCNA LI for 22 cases carrying positive p53 protein was 52.3%, which was higher than that of p53 protein negative cases (35.7%). The Kaplan-Meier survival curve of the patients who were negative for p53 was significantly more favorable than for patients who were positive for p53 (P < 0.01, Cox-Mantel test). These results suggest that PCNA and p53 LI are markers for the malignant potential of the oral mucosa, and are a useful indicator suggesting a poor prognosis.

[Strongyloidiasis associated with multiple myeloma in Ehime prefecture].

A 76-year-old male with multiple myeloma, who was born and had lived in Ehime Prefecture was admitted to our hospital because of high fever. The chest X-p film showed right middle lobal pneumonia. Eosinophilia was detected and a stool smear examination revealed rhabditis form larvae of the nematode. The filaria form larvae of Strongyloides stercoralis were detected by stool culture. Antibiotics and pyruvinium pamoate were administered to bacterial pneumonia and hyperinfection of the strongyloides, respectively. Consequently, pneumonia and strongyloidiasis were promptly improved. It was considered that he was infected with strongyloides in south-east Asia during World-War II and a little autoinfection of this nematode had continued about 50 years. In addition, it was suggested that hyperinfection of strongyloides resulted from the immunosuppressive state induced by the chemotherapy for multiple myeloma.

[Hereditary gingival fibromatosis].

Fibromatosis may be defined as diffuse poorly circumscribed overgrowth of the fibrous tissue that infiltrates adjacent normal tissues. They are difficult to eradicate surgically, and recur but not metastasize. Gingival fibromatosis is generally regarded as a disease that leads to an extensively diffuse and remarkable hyperplasia of the maxillo-mandibular gingiva. Occasionally, this lesion covers all teeth. The histogenesis of the fibromatosis remains unexplained. Trauma, endocrine, idiopathic factors and genetic factors have been implicated, but it is uncertain whether any of then play a major role in the development of the disease. Occasional cases with familial history have been reported. The treatment of choice would appear to been block resection of the tumor and surrounding normal structures. Although, this lesion has a high recurrent rate. For this reason, in many of the case reports, that it has been recommended that the follow up period is considerably less than three years.