Oral Squamous Cell Carcinoma Biopsies Research Articles

Prognostic value of transforming growth factor beta 1 [TGF-β1] and matrix metalloproteinase 9 [MMP-9] in oral squamous cell carcinoma

Objectives: The prognostic value of MMP-9 and TGF-β1 in oral squamous cell carcinoma (OSCC) is not clear. This study aimed to assess this subject.Methods: After immunohistochemistry staining of 48 OSCC biopsies with MMP-9 and TGF-β1 markers, marker expression in the stroma was estimated, and the correlations with OSCC prognosis determinants were analyzed (α = 0.05).Results: Mode of invasion was associated with metastasis (rho = 0.449, p = 0.008). MMP-9 was positively associated with metastasis and mode of invasion. TGF-β1 was negatively correlated to tumor histologic grade but was positively associated with metastasis.Conclusions: Unlike TGF-β1, MMP-9 might be useful for prognosis determination.

Correlation between koilocytes and human papillomavirus detection by PCR in oral and oropharynx squamous cell carcinoma biopsies

The purpose of this study was to compare the histopathological analysis with polymerase chain reaction (PCR) methods to predict the presence of human papillomavirus (HPV) infection in oral squamous cell carcinoma biopsies. Eighty-three paraffin-embedded tissue specimens from patients with oropharynx and mouth floor squamous cell carcinoma were submitted to histopathological analysis under light microscopy, specifically for the determination of the presence of koilocytes. Subsequently, DNA was purified from the same paraffin-embedded specimens and submitted to PCR. Fisher's exact test showed no statistically significant correlation between the two methods. The results suggest that the presence of koilocytes is unreliable for the detection of HPV presence in oral and oropharynx squamous cell carcinoma.

Stress hormones increase cell proliferation and regulates interleukin-6 secretion in human oral squamous cell carcinoma cells

Patients with oral cancer can have high psychological distress levels, but the effects of stress-related hormones on oral cancer cells and possible mechanisms underlying these relationships are unknown. In this study, we have investigated the effects of stress-related hormones on interleukin-6 (IL-6) secretion and proliferation of oral squamous cell carcinoma (OSCC) cells. The effects of norepinephrine (NE), and cortisol were studied in SCC9, SCC15, and SCC25 cells and effects of isoproterenol in SCC9 and SCC25 cells. Real-time PCR studies revealed constitutive β1- and β2-adrenergic receptors (β-ARs) expression in the SCC9, SCC15, and SCC25 cells. The results showed that NE and isoproterenol significantly enhanced IL-6 mRNA expression and protein production in supernatants of SCC9 and SCC25 cells. Physiological stress levels of NE and isoproterenol (10μM) at 1h elicited the most robust IL-6 increase. Regarding IL-6 secretion, 10μM NE induced a 5-fold increase at 1h, 3.7-fold increase at 6h, and 3.2-fold at 24h in SCC9 cells. These effects were blocked by the β-adrenergic antagonist propranolol, supporting a role for β-ARs in IL-6 secretion. The effects of cortisol varied according to the hormone concentration. Pharmacological concentrations of cortisol (1000nM) inhibited IL-6 production by SCC9 and SCC25 cells. Cortisol dose that simulates stress conditions (10nM) tended to increase IL-6 expression in SCC9 cells. Hormonal doses that simulate stress conditions (10μM NE, at 6h in SCC9 and SCC15 cells and 10nM cortisol, at 48h in SCC15 cells) stimulated increased cell proliferation. Treatment of SCC9 cells with IL-6 neutralizing ab (10μg/mL) partially inhibited NE-induced proliferation. Finally, 20 OSCC biopsies were shown to express β1- and β2-ARs. These findings suggest that stress hormones can affect oral cancer cells behavior.

Bone-Invasive Oral Squamous Cell Carcinoma in Cats

Feline oral squamous cell carcinoma (OSCC) is the most common oral tumor in cats. There is no effective treatment, and the average duration of survival after diagnosis is only 2 months. Feline OSCC is frequently associated with osteolysis; however, the mechanisms responsible are unknown. The objective of this study was to characterize the epidemiology and pathology of bone-invasive OSCC in cats and to determine the expression of select bone resorption agonists. In sum, 451 cases of feline OSCC were evaluated. There was no sex or breed predisposition, although there were more intact cats in the OSCC group compared to the control group. Gingiva was the most common site, followed by the sublingual region and tongue. Cats with lingual OSCC were younger (mean, 11.9 years) compared to cats with gingival OSCC (mean, 13.6 years). In addition to osteolysis, there was periosteal new bone formation, osseous metaplasia of tumor stroma, and direct apposition of OSCC to fragments of bone, suggestive of bone-binding behavior. Eighty-two cases were selected for immunohistochemical detection of parathyroid hormone-related protein (PTHrP). Specimens with osteolysis had increased PTHrP expression and nuclear localization, compared to OSCC without osteolysis. Thirty-eight biopsies of OSCC with osteolysis were evaluated for tumor necrosis factor α expression, and only 4 biopsies had such expression in a small proportion of tumor cells. Increased tumor expression of PTHrP and increased localization of PTHrP to the nucleus were associated with osteolysis and may play an important role in bone resorption and tumor invasion in cats with OSCC.

BUBR1 expression in benign oral lesions and squamous cell carcinomas: Correlation with human papillomavirus

Oral squamous cell carcinoma (OSCC) is the most common head and neck cancer. Only in Brazil, the estimate is for 14,160 new cases in 2009. HPV is associated with increasing risk of oral cancer, but its role in carcinogenesis is still controversial. BUBR1, an important protein in the mitotic spindle assembly checkpoint (SAC), has been associated with some virus-encoded proteins and cancer. The aim of the present study was to evaluate the expression of BUBR1 in non-malignant oral lesions and OSCC with and without metastasis associated with HPV infection. We performed immunohistochemistry for BUBR1 in 70 OSCC biopsies divided into three groups (in situ tumors, invasive tumors without metastasis and invasive tumors with metastasis) with their respective lymph nodes from samples with metastasis and in 16 non-malignant oral lesions. PCR was performed in order to detect HPV DNA. Significantly higher BUBR1 expression associated with shorter survival (p=0.0479) was observed in malignant lesions. There was also a significant correlation (r=1.000) with BUBR1 expression in lesions with metastasis and their lymph nodes. Ninety percent of OSCC and 100% of benign lesions were HPV positive. HPV16 and HVP18 were present in 13 and 24% of HPV-positive OSCC samples, respectively. HPV was more prevalent (76%) in samples with a high BUBR1 expression and the absence of viral DNA had no influence on BUBR1 expression. These findings suggest that HPV could be associated with overexpression of BUBR1 in OSCC, but not in benign oral lesions.

Upregulation of Notch pathway molecules in oral squamous cell carcinoma

The constitutive activation of the Notch pathway has been demonstrated in various types of malignancies. However, it remains unclear how the Notch pathway is involved in the pathogenesis of oral squamous cell carcinoma (OSCC). We investigated the expression of Notch pathway molecules in OSCC cell lines and biopsy specimens and examined the effect of Notch pathway inhibition. Reverse transcription-polymerase chain reaction revealed upregulation of Notch1, Notch2, Jagged1, HES1 and HEY1 in both OSCC cell lines and biopsy specimens. Immunohistochemical examination showed that the Notch intracellular domain accumulates in the nucleus of cells in OSCC cell lines and biopsy specimens. In addition, Jagged1 is expressed in the cytoplasm of cells in OSCC cell lines and biopsy specimens. Furthermore, Notch pathway inhibition using a gamma-secretase inhibitor prevented the growth of OSCC in vitro. These findings suggest that inhibition of the Notch pathway suppresses OSCC growth and may be a useful approach for the treatment of patients with OSCC.

Her-2 Immunohistochemical Expression in Oral Squamous Cell Carcinomas is Associated with Polysomy of Chromosome 17, Not Her-2 Amplification

Based on the prognostic role of Her-2 amplification and protein overexpression in breast cancer, various studies have been performed in oral squamous cell carcinomas (OSCC) with inconsistent results. As in invasive breast carcinomas Her-2 overexpression has been related to an increased number of chromosome 17 copies, a common chromosomal alteration in OSCC, we evaluated the association between polysomy 17 and Her-2 protein expression in a series of primary OSCC. Forty-one incisional biopsies of primary OSCC were included in the study. Protein expression was evaluated immunochistochemically with CB11 mouse monoclonal anti-human antibody. The reaction was arbitrarily characterized as absent, faint, moderate, and strong, and staining pattern as cytoplasmic and membranous. Positive cases were analyzed by chromogenic in situ hybridisation (CISH) to access Her-2 status. The association between polysomy 17 and Her-2 expression was checked by Fisher's exact test. Four cases were negative and 37 cases were positive for Her-2. Staining was faint in 15 cases and moderate in 22 cases. CISH showed that all cases with faint staining were diploid, while from the cases with moderate staining 10 were diploid and 12 polysomic for chromosome 17. Thirteen cases showed purely cytoplasmic staining, while in 24 there were areas of both cytoplasmic and membranous staining. There was a statistically significant correlation between intensity of the reaction and polysomy 17 (P = 0.0036), in particular for cases with both cytoplasmic and membranous staining (P = 0.0128). In some OSCC Her-2 immunohistochemical expression may be associated with chromosome 17 polysomy and not Her-2 amplification.

VEGF-C and VEGF-D expression is correlated with lymphatic vessel density and lymph node metastasis in oral squamous cell carcinoma: Implications for use as a prognostic marker

The prognosis of patients with oral squamous cell carcinoma (SCC) is influenced by the presence of lymph node metastasis. In this study, we analyzed the relationship between lymphangiogenesis and the expression of VEGF-C and VEGF-D in association with lymph node metastasis in oral SCC. Oral SCC biopsy specimens (160 cases) were examined for lymphatic vessel density (LVD) and the expression of VEGF-C and VEGF-D immunohistochemically. The levels of VEGF-C and VEGF-D expression and LVD were significantly associated with lymph node metastasis (p<0.001). The expression of VEGF-C and VEGF-D increased the LVD significantly (p<0.001). Multivariate analysis showed that VEGF-C expression and LVD were significantly associated with lymph node metastasis (p<0.001). This study presents clinical evidence for the important roles of VEGF-C and VEGF-D in lymphangiogenesis and lymphatic metastasis of oral SCC, and suggests that VEGF-C or LVD can effectively predict lymphatic metastasis of oral SCC.

Detection of EBV Infection and Gene Expression in Oral Cancer from Patients in Taiwan by Microarray Analysis

Epstein-Barr virus is known to cause nasopharyngeal carcinoma. Although oral cavity is located close to the nasal pharynx, the pathogenetic role of Epstein-Barr virus (EBV) in oral cancers is unclear. This molecular epidemiology study uses EBV genomic microarray (EBV-chip) to simultaneously detect the prevalent rate and viral gene expression patterns in 57 oral squamous cell carcinoma biopsies (OSCC) collected from patients in Taiwan. The majority of the specimens (82.5%) were EBV-positive that probably expressed coincidently the genes for EBNAs, LMP2A and 2B, and certain structural proteins. Importantly, the genes fabricated at the spots 61 (BBRF1, BBRF2, and BBRF3) and 68 (BDLF4 and BDRF1) on EBV-chip were actively expressed in a significantly greater number of OSCC exhibiting exophytic morphology or ulceration than those tissues with deep invasive lesions (P = .0265 and .0141, resp.). The results may thus provide the lead information for understanding the role of EBV in oral cancer pathogenesis.

Imaging mass spectrometry using chemical inkjet printing reveals differential protein expression in human oral squamous cell carcinoma

Matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) is a recently developed technique that generates molecular profiles usually of peptide and protein signals directly from the surface of thin tissue sections and can be coupled with automation to generate two-dimensional ion density maps. This allows specific information to be obtained on the relative abundance and spatial distribution of the analytes of interest. The technique has potential for application in many diseases including cancer with respect to elucidating the molecular pathology and identifying potential biomarkers. In this proof-of-principle study we have evaluated inkjet printing of the sinapinic acid matrix used for MALDI-IMS directly onto the surface of human oral squamous cell carcinoma biopsy specimens. This MS profiling technique produced reproducible informative chemical images for clinical pathology. Analysis of the resulting protein profiles of highly expressed protein in squamous cell carcinoma of the tongue reveals spectral features at approximately 4500 and approximately 8360 Da.

DETECTION OF p53 MUTATIONS ON ORAL SQUAMOUS CELL CARCINOMA

Oral squamous cell carcinoma (OSCC) is the most common malignant tumor of the oral cavity and it account for 80-90% of all malignancies in the oral cavity. The aim of this study was to determine the presence of p53 mutations and to associate these mutations with the histopathological type of OSCC such as well differentiated and poorly differentiated. Analytical observational comparative study by cross sectional design was used. Forty untreated well and poorly differentiated OSCC biopsy sample and normal tissue biopsy material taken from 16 normal patients were analyzed for the presence of mutation in the conserved region of the p53 gene especially in exon 5 by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). The results of this study showed that p53 gene mutations were detected in exon 5; 11/40 (27.55%) with heterozygous mutation 9/11 (81.8%). The incidence in exon 5 of p53 gene mutation was significantly associated with well differentiated 2/20 (10%) and poorly differentiated 9/20 (45%) OSCPP (P=0,013). This study concludes that 1) mutation in exon 5 of p53 gene occurred frequently in OSCC; 2) exon 5 of the p53 gene could be one of the specific targets for histopathological grade of OSCC; 3) mutation in the exon 5 of p53 gene could be important prognostic factor in OSCC.

Increased number of CD25+ FoxP3+ regulatory T cells in oral squamous cell carcinomas detected by chromogenic immunohistochemical double staining

The role of tumor-infiltrating regulatory T cells (Treg) compromising antitumor effects of immune cells in oral squamous cell carcinoma (OSCC) is largely unknown. The presence of CD25(+) FoxP3(+) Treg as well as of CD3(+) FoxP3(+) and of CD8(+) FoxP3(+) tumor-infiltrating lymphocytes (TIL) was verified in OSCC and compared with non-cancerous lymphoepithelial tissue. Three double stainings (CD3/FoxP3, CD8/FoxP3 and CD25/FoxP3) were performed on tissue sections of 15 OSCC and compared with 15 human tonsils. OSCC biopsy samples provide evidence for a strong infiltration of TIL, in particular, naturally occurring CD25(+) FoxP3(+) Treg. Whereas a comparison of OSCC and control tissue did not show significant changes in the number of CD3(+) FoxP3(+) TIL and of CD8(+) FoxP3(+) TIL, a significantly higher frequency of CD25(+) FoxP3(+) TIL (Treg) could be observed in OSCC (P < 0.001, two-sided t-test). Given the small number of specimens, a significant correlation with tumor stage could not be verified. Chromogenic double staining of CD4/FoxP3 is a promising tool for the detection of Treg in paraffin-embedded tissue of OSCC.

Decreased gene expression of human β-defensin-1 in the development of squamous cell carcinoma of the oral cavity

The aim of this study was to investigate the gene expression of human β-defensin-1, -2, -3 (hBD-1, -2, -3), interleukin-1β, tumour necrosis factor-α and cyclooxygenase-2 in oral squamous cell carcinoma (OSCC) compared to benign and premalignant lesions as well as healthy controls. Biopsies of healthy gingiva (n = 5), irritation fibroma (n = 5), leukoplakia (n = 5) and OSCC (n = 5) were obtained during routine surgical procedures. RNA was extracted according to standard protocols and transcripts of hBD-1, -2, -3, interleukin-1β, tumour necrosis factor-α and cyclooxygenase-2 were analysed by real-time polymerase chain reaction. The expression of hBD-1 was reduced in all lesions (5-fold in irritation fibroma and 2.5-fold in leukoplakia), but most significantly (50-fold) in OSCC. hBD-1 appears to play a role in the development of OSCC. The loss of its function might contribute to the malignant progression of these tumours.

The pattern of p53 gene mutations on oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is the result of accumulation genetic lesion and caused by specific mutation in specific key regulation genes. p53 gene is key target specific regulatory genes which function as negative regulators in cell cycle control. The highest mutation rates found in human cancers and the etiology in high risk populations and the pattern of molecular pathogenesis mechanism involved in the OSCC remain unclear. The purpose of this study was to determine the presence of alteration or mutation of p53 gene and to associate these mutations histopathological status of the patients such as well differentiated and poorly differentiated in OSCC in order to elucidate the molecular pathogenesis mechanism of OSCC based on the pattern of p53 gene. Using 40 untreated well and poorly differentiated OSCC biopsy samples and 16 normal patients were analyzed for the presence of mutation in the conserved region of the p53 gene exons 5 and or 7 by PCR-SSCP mutational analysis for p53 gene showed 70% of total samples : exon 5: 27.5% with heterozygous mutation 81.8%, exon 7: 55% with heterozygous mutation 100%. The incidence of p53 mutation was not significantly associated with well and poorly differentiated OSCC with the exception in exon 5 of p53 gene (p = 0.013) using contingency coefficient. This study concludes that mutation of p53 gene especially in exon 7 may not indirectly play in the progressivity of OSCC with the exception of mutation in exon 5 of p53 which indicates the essential role in the progressivity of OSCC.

Expression of caveolin-1 and its correlation with cisplatin sensitivity in oral squamous cell carcinoma

Cisplatin (CDDP) is widely used for chemotherapy of oral squamous cell carcinoma (OSCC). However, the mechanism of resistance to CDDP is unclear. Recently, caveolin-1 was identified as being associated with both metastasis and multidrug resistance. In the present study, we showed that caveolin-1 expression is significantly related to chemosensitivity in OSCC. We established a CDDP-resistant cell line, H-1R, from the parental OSCC cell line, H-1. Caveolin-1 expression was determined by reverse transcriptase-polymerase chain reaction (RT-PCR) in both cell lines. We analyzed expression of caveolin-1 in 30 OSCC biopsy specimens and investigated the relationship between expression of caveolin-1 and patients' clinicopathological parameters and chemotherapeutic responses. The 3-(3,4-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay indicated that H-1R has a ten-times greater resistance to CDDP than H-1 has. The level of caveolin-1 expression in H-1R was significantly decreased in comparison with that in H-1 by real-time RT-PCR analysis. Positive caveolin-1 immunostaining correlated positively with a complete response (16/20, 80.0%). However, negative immunostaining was found in 6/7 (85.7%) cases with no response. Positive immunohistochemical staining of caveolin-1 correlated positively with chemosensitivity to CDDP-based combination chemotherapy (P=0.02). These results suggest that overexpression of the caveolin-1 gene may provide novel diagnostic markers associated with CDDP sensitivity in OSCC.

Human Papillomavirus in Oral Cancer from Sri Lanka: Prevalence and relationship with clinico-pathological parameters.

The presence of human papillomavirus (HPV) in two groups of oral squamous cell carcinoma (OSCC) from Sri Lanka were analyzed by PCR and direct cycle sequencing using consensus GP5+/6+ primers. The two groups of OSCC specimens analyzed were 68 formalin-fixed paraffinembedded surgical specimens and 34 formalin-fixed unembedded biopsies. Twenty-five patients from the latter group were habitual betel quid chewers. The HPV prevalence of the overall sample was 38/102 (37.2%). However, HPV was detected in significantly higher rate from formalin-fixed unembedded biopsies (61.7%) compared to paraffin embedded surgical specimens (25%) (p<0.0003). Upon sequencing, high-risk HPV types, 16,18, 45 and 66 were detected in 65% of the HPV infected tumors. Moreover, HPV was detected at significantly higher rates from small primary tumors (T1&T2) (75%) compared to large primary tumors (T3&T4) (30%) (p<0.025). However, no correlation existed between other clinico-pathological variables such as age, gender, sites, habits or histological differentiation and HPV detection when analyzed statistically using Pearson's chi-square test (at 5% level of significance). In conclusion, high prevalence of HPV in formalin fixed-unembedded OSCC biopsies from Sri Lanka indicate that, in addition to betel quid, viral infection may play a role in oral carcinogenesis and support the multi-factor model for oral cancer causation.

Antioxidant enzyme levels in oral squamous cell carcinoma and normal human oral epithelium.

The antioxidant enzymes (manganese- and copper-zinc-containing superoxide dismutases, catalase and glutathione peroxidase) limit cell injury induced by reactive oxygen species. The purpose of the study was to determine whether human oral squamous cell carcinomas have altered antioxidant enzyme levels. This study is the first to undertake this task in human oral mucosa and squamous cell carcinoma. Semiquantitative immunohistochemistry was used to examine 26 archived oral squamous cell carcinoma biopsies. Fourteen well-differentiated and 12 poorly differentiated tumors were examined, as were 12 specimens of oral mucosa. All sections were reviewed by two oral and maxillofacial pathologists, and image analysis of the immunostained sections was performed using NIH Image. Antioxidant enzyme staining intensities were compared in the different groups by Duncan's multiple range test. In general, mucosal basal cells displayed lower antioxidant enzyme levels than spinous cells, and primary tumor cells displayed lower antioxidant enzyme staining intensities than did their normal cell counterparts. Moreover, poorly differentiated tumor cells showed lower antioxidant enzyme staining intensities than well-differentiated tumor cells. Manganese-containing superoxide dismutase staining intensities were, however, higher in well-differentiated oral squamous cell carcinomas than their normal cells of origin. Detection of antioxidant enzymes may be a useful future marker in the molecular diagnosis of the oral cancer. Moreover, it may be possible to not only monitor the effectiveness of chemopreventive and therapeutic strategies in oral cancer using these enzymes, but to monitor tumor recurrence.

Soluble fibronectin promotes migration of oral squamous-cell carcinoma cells.

We have shown that a fibronectin (FN) matrix is required for the organization of tenascin-C (TN-C) matrices by peritumor fibroblasts (PTF) cultured from tissue surrounding oral squamous-cell carcinoma (SCC). In the present study, we detected alternatively spliced FN containing both the EDA and EDB domains decorating the reactive stroma adjacent to the invading tumor nests in oral SCC biopsies. In vitro, PTF cells organized an extensive FN matrix rich in the EDA domain and containing a small amount of EDB. In contrast, normal human fibroblasts deposited a FN matrix which expressed only the EDA domain. PTF-conditioned medium (CM), shown to enhance migration of oral SCC cells on TN-C, was found to enhance their migration on FN and invasion of a reconstituted basement membrane. Addition of antibodies to FN to the PTF-CM inhibited SCC-cell migration on TN-C, and depletion of FN from the PTF-CM abolished its ability to induce migration or invasion by oral SCC cells, suggesting that FN promotes the migration and invasion of oral SCC cells. Western blots of the PTF-CM identified FN containing the EDA but not the EDB domain. When soluble FN was added to the control medium in the lower chamber of the Transwell system, SCC-cell migration increased significantly. These results demonstrate that both the EDA and the EDB domains of FN are expressed in the extracellular matrix of oral SCC in vivo and PTF in vitro and indicate that FN is the probable chemotactic factor in the PTF-conditioned medium.

P53 Expression and p53 gene mutation in oral cancer and dysplasia

BACKGROUND: The aim of the current study was to examine the possible association of the p53 tumor suppressor gene with the development of oral cancer. We examined biopsy material from patients with oral squamous cell carcinoma for p53 protein expression and p53 mutations. METHODS: Eighteen samples were analyzed. These comprised oral squamous cell carcinoma biopsy samples and oral dysplastic biopsy material taken from nine patients. An immunohistochemical technique was used to determine p53 protein expression. A direct sequencing technique was used to detect mutations in the p53 gene itself. RESULTS: Overall seven of nine (77%) oral squamous cell carcinomas and seven of nine (77%) oral dysplastic lesions analyzed showed evidence of p53 protein over-expression. Mutations in exons 5 and 6 of the p53 gene were detected in five of nine (55%) oral squamous cell carcinomas and four of nine (44%) dysplastic lesions analyzed. CONCLUSIONS: Both p53 protein over-expression and p53 mutations were noted in dysplastic oral lesions. Over-expression of the p53 protein and p53 gene mutations were not always coexistent, however, suggesting that gene mutation may be only one of the mechanisms responsible for stabilization of the p53 protein. (Otolaryngol Head Neck Surg 1998;118:115-23.)

Mode of tumor invasion in oral squamous cell carcinoma: improved grading based on immunohistochemical examination of extracellular matrices.

To predict the nodal involvement of oral squamous cell carcinoma (OSCC) many investigators have studied the histologic features of primary tumors. However, conventional histologic grading still is not sufficient to provide an objective and practical evaluation. In this study we tried to modify one standard type of histologic grading, namely, mode of invasion (MI), on the basis of the extracellular matrix (ECM) staining pattern. One hundred seventeen initial biopsies of primary OSCC were histologically examined, and the MI was graded. The expression of a series of ECMs-laminin, type IV collagen, heparin-sulphate proteoglycan, fibronectin, tenascin, decorin, and vitronectin-was immunohistochemically examined. The ECM staining pattern showed a close association with the MI, and was classified into types I and II. The type I staining pattern was observed in the noninvasive cases (MI grades 1 and 2), while type II was in the highly invasive cases (MI grades 4c and 4d). Because the ECM staining pattern in the moderately invasive cases (MI grade 3) included both types I and II, these cases were subdivided and compared. As a result, the type II cases showed a significantly higher incidence of nodal involvement than did the type I cases. An examination of a series of the ECMs, in addition to the conventional histologic examination, is thus considered to provide more objective and practical data to evaluate the invasive and metastatic potential of OSCC.