Metastatic spread to cervical lymph nodes(LNs) is a major determinant of outcome in oral squamous cell carcinoma (SCC). To provide an useful method for the detection of lymph node micrometastases, we fulfilled the histopathological examination and reverse transcriptase polymerase chain reaction(RT-PCR) using the paraffin-embedded LNs of oral SCC patients. In this study, 78 LNs from 12 patients with primary oral SCC were analyzed. Metastases in the regional LNs were evaluated by RT-PCR for squamous cell carcinoma antigen(SCCA) and cytokeratin 5(CK5). Detectability of metastatic LNs by RT-PCR was compared with histopathological examination. Of 78 LNs, CK5 and SCCA mRNA were detected in 32(41.0%) and 8(10.3%), respectively. Histopathologically, 10(12.8%) of 78 LNs were positive. CK5 mRNA was detected in all 10 histopathologically positive LNs. In contrast, SCCA mRNA was detected in 5 of 10 histopathologically positive LNs. These findings suggest that genetic diagnosis by RT-PCR based on CK5 mRNA expression may be sensitive and clinically useful technique to detect the presence of metastatic carcinoma cells in regional lymph nodes of oral SCC.

The purpose of this study was to evaluate the role of c-fos and c-jun expression in the oral squamous cell carcinoma and ameloblastomas. For this study, 12 subjects diagnosed as squamous cell carcinoma and 7 subjects of ameloblastomas referred to the Dept. of Oral Path. School of Dentistry, Kyung Hee University, 2 subjects of normal oral mucosa without any inflammatory changes were used as experimental, control groups respectively. All the tissues ; experimental and control group were fixed in neutral formalin solution and embedded in paraffin, serial tissue section were made 5㎛ in thickness and processed in the standard way for immunohistochemical method, using primary and secondary antibodies, for c-fos (Anti-c-fos protein, rabbit polyclonal kit at 1:100 dilution), c-jun( anti-c-jun protein, rabbit polyclonal at 1:100 dilution), all BioGenex U.S.A. made, followed by the Streptavidin - Horse Radish Peroxidase(InnoGenex, Human-Avidin kit) application, counter stained with Mayer's hematoxylin stain method, mounted. And examined under the biologic microscope, with the criteria-(no epithelial stain), +(weak or focal epithelial stain), ++(moderate or focal intensive epithelial stain), +++(intense generalized epithelial staining) for the epithelial, and connective tissue component in squamous cell carcinomas, ameloblastomas and normal mucosal epithelium on each. Attained results as follows ; 1. In oral mucosa, c-fos and c-jun intensely expressed on the all cell layers except on the basal layer. Intense reaction is noted in the c-jun than in the c-fos. and a few cells with positive cytoplasm, negative nuclear are scattered in all layer. 2. The response to c-fos in ameloblastomas, is various according to the histological type, but intense resposes are nodted in nuclear and cytoplasm on the tall columnar cells at the periphery of the follicles compare to that on the stellate cells. 3. The respone to c-fos in squamous cell carcinoma, intense reaction is noted in cytoplasm and nuclei of the tumor cells in well differentiated, poorly differentiated type. 4. The response to c-jun in ameloblastoma, it is noted that moderate respone in nuclear and cytoplasm, at the tall columnar cells at the periphery of follicular or plexiform type but intense respone was notes on the columnar cells, and stellate cell in cytoplasm and nuclear of acanthomatous type. 5. The respon to c-jun in squamous cell carcinoma, it is noted that intensive responses only in cytoplasm in well differentiated type, but intensive responses in nuclei and cytoplasm in the poorly differentiated type are revealed. Intensive responses on c-fos, c-jun were noted on the high atypical cells. This results suggest that c-fos and c-jun may be affected to the reactivation on growth and development of the squamous cell carcinoma.

The p16 gene encodes an inhibitor of the cyclin-dependent kinase, which inactivates cyclin-dependent kinase and contro1s the cell cycle progression, The 10ss of p16 expression or overexpression has been reported in many kinds of tumors, Both p16 and PCNA regu1ates cell cycle progression at the Gl/8 checkpoint, Although many researches about the p16 expression in ora1 cancer have been carried out, there are few studies about the corre1ation between p16 ex pression and pro1iferation of ora1 cancer cells The object of this study was to eva1uate the avai1ability of p16 as ear1y diagnostic factor and prognostic factor through corre1atión ana1ysis of p16 expression in ora1 squamous cell carcinoma and its re1ation to PCNA index and clinicopatho1ogic factors 80 we investigated p16 immunohistochemica1 expression of 83 ora1 squmaous cell carcinomas, and obtained the resu1ts as followed, 18 out of the 83 cases(21, 69%) showed p16 positive and 65 samp1es(78,31%) showed p16 negative, Whi1e the mean va1ue of PCNA indices of p16 positive cases was 65,94 ::t 18,32, that of PCNA indices at p16 negati ve ones 54,79 ::t 18, 39, This difference between them showed statistica1 sígnificance, (P=O, 030) p16 positive group was 12/60(20, 0%) of well differentiated tumors and p16 negative group was 6/23(16, 1%) of moderate1y or poor1y differentiated tumors, This difference did not show statistica1 significance. (P=O. 372) From the resu1ts above, it was suggested p16 expression is re1ated to PCNA index in ora1 squamous cell carcinomas.

Amino acid transporters play an important role in supplying organic nutrient to cells. The expression of L-type arnino acid transporter 1 (LATl) and its subunit 4F2 heavy chain (4F2hc) was evaluated to deterrnine the alterations to these transporters in oral norrnal mucosa (ONM) , oral precancerous lesion (OPL) and oral squamous cell carcinoma (OSCC). Sections from formalin-ftxed, paraffm-embedded S따nples of ONM, OPL or OSCC were exarnined using immunohistochernical staining to detect LATl and 4F2hc proteins. 까le LATl and 4F강lC expression increased progressively from ONM to hypeφ，Iastic and to dysplastic lesions and OSCC. In partiαlar， LATl rnay be a more S야dftc indicator of tumor prog~않sion than 4F2hc. 까le gradually increasing LA Tl and 4F2hc expression detected during the multistep progressive change shows that the protein rnay have an important role in the early stages of multistep oral carcinogenesis. In addition, the specific inhibition of LA Tl and 4F2hc rnight be a new rationale to suppress oral cancer progression.

Squamous cell carcinoma comprises about 95% of oral cancers. 까le gene디C 없mage in carcinogen-exposed fields is accumulated to transforrn norrnal mucosa in dysplas디c tissue and fmally invasive carcinoma through multistep process. This carcinogenic process has been a cause of the development of secondary tumors after the removal of primary carcmoma. πle improvement of therapeutic modalities of oral cancer has driven into the increase of multiple cancer occurrence in head and neck region. We experienced 3 pa디ents who had mul디ple squamous cell carcinomas in oral cavlty. π1ÎS study aimed to report multiple pr따laπ squamous cell carcinoma by clinical and pathologic examination and to discuss their molecular mechanism

To cletermine the role of mismatch repair in the clevelopment of oral squamolls cell carcinomas (OSCCs) , the prevalence of microsatellite instability (MSI), expression of I뼈LH1 ancll띠SH2 ， ancl hypennethylation of I뻐LH1 ancl hMSH2 were explorecl. Bya panel of five markers (BAT25, BAT26 , 02S123, 05S346, ancl 017~‘250 ， the so-callecl Bethescla markers) for screening of MSI, MSI was observecl in 5 of the 15 sqllamolls cell carcinomas (33.3%). As MSI is callsecl by the clysfllnction of MMR genes, this stucly examinecl the methylation status of CpG sites in the hMLH 1 ancl hMSH2 promoters ancl the expression hMLH1 ancl hMSH2. Becallse of inappr이)riate efficiency of fonnalin-frxecl ancl paraffin-embeclclecl samples for methylation-specific PCR (MS-PCR) of hMLH1 ancl hMSH2, the role of promoter hypelmethylation 띠 the clevelopment of MSI ancl expression of hMLH1 ancl hMSH2. meùlylation was failecl to clefìne. However, loss of nllclear staining of 1ψIILH 1 ancl hMSH2 were seen in nine (69.2%) ancl four (30.7%) of 13 OSCCs, while four ancl fìve all corresponcling normal epiùlelial tisslles showecl positive nllclear stι ining ofl마ILH1 and hMSH2. These data suggest that MSI anclloss of expression of hMLH1 ancl hMSH2 play a role in the carcinc핑enesis of oral sqllamolls cell carcinomas thollgh the role of promoter hypermethylation of hMLH1 ancl hMSH2 in MSI and their expresslon IS lIncertam

Matrix metalloproteinases(MMPs) are involved in the degradation of extracellular matrix, which is re lated to infiltrative growth and metastasis of tumor and are regulated by tisslle inhibitor of metalloproteinases(TIMPs) or cell adhesion molecllles such as E-cadherin and epidermal growth factor receptor (EGFR). The aim of this study is to evaluate the relationship between MMP-2, MMP-9 expressions and clinico-pathologic factors, 끼MP-1 ， TIMP-2, EGFR and E-cadherin expressions. lmmunohistochemical stains were perfom1ed on 55 cases of squamous cell carcinoma of the ordl cavity and the results were as follows. MMP-2 and MMP-9 expressions were noted in 30(54.5%) and 22(40.0%) of 55 cases, TIMP-1 and πMP-2 in 21(38.2%) and 33(60.0%), and E-cadherin and EGFR expressions in 35(63.6%) and 26(47.3%) of 55 cases, respectively. MMP-2 expression rdte was slightly higher 띠 cases without recurrence, and 끼MP- 2 expression rate was slightly higher in cases showing more inftltrative growth pattem. 까1e expression rate of EGFR was higher in cases with well differentiation(p=0.OO47), but no posi디ve relationship between the expression rate of Ecadherin and histologic grade was found. Cases with positive reaction for MMP-9 showed an increasing tendency of nega디ve reaction for TIMP-1. π1e expression rate of MMP-2 was higher in cases with positive reaction for E-cadherin and EGFR with no statistical significance. 까1e expression rate of MMP-9 was significantly higher in cases with positive reaction for E-cadherin(p=0.022l). These results suggest that MMP-2, MMP-9, TIMP-1 and TIMP-2 expressions are involved in the development of oral squamous cell carcinomas, but MMP-2, MMP-9, 끼MP-1 ， and 끼MP-2 expressions might not seem to be a useful prognostic factors because there were no significant relationship between clinicopathologic parameters. EGFR expression showed positive correlation with low histologic grade, so EGFR expression could be regarded as a good prognostic factor. In the progression of sqllamous cell

까le purpose of this study was to evaluate the role of EGF(Epidermal Growth Factor) , EGFR(Epidemlal Growth Factor Receptor), aFGF(acidic Fibroblast Growth Factor, FGF-1), bFGF(basic Fibroblast Growth Factor, FGF-2), FGFR(Fibroblast Growth Factor Receptor), and VEGF(Vascular Endothelial Growth Factor) 띠 the development of the oral squamous cell carcinoma. For this study 6 subjects, diagnosed as squamous cell carcinoma refelTed to the Dept. of Oral Path. College of Dentistry, Kyung Hee University, 2 subjects of normal 이띠 mucosa with any inflammatolY changes were used as expelimental, control groups respectively. AlI the 디ssu es ; expe디me nta l and control group were fixed in 100;ú neutral fOlmalin solution and embeclded in paraffìn , seIial tissue section were made 511m in thickness ancl processecl in the standard way for immunohistochemical methocl, using primary ancl seconclalY antibodies, for EGF(Antirabbit Ig G, rabbit kit at 1:100 dilution), EGFR(Antimouse Ig G, mouse kit at 1:100 dilution), aFGF(Antirabbit Ig G, rabbit kit at 1:100 dilution), bFGF(Antirabbit Ig G, rabbit 써t at 1:100 dilution), FGFR(Antimouse Ig G, mouse kit at 1:100 dilution), ancl VEGF(Antirabbit Ig G, rabbit kit at 1:100 clilution), all BioGenex U.S.A. macle, followed by the Stre ptavidin - Horse Radish Peroxidase(InnoGenex, Human-avidin kit) application, counter stained with Mayer’s hematoxylin stain method. And examined under the biologic microscope, -(no epithelial stain), +(weak or focal epithelial stain), ++(mode rate or focal intensive epithelial stain), +++(intense generalized epithelial staining) for the epithelial, and connective tissue component in squamous cell carcinoma and in nomlal mucosal epithelium on each. Attained results as follows ; 1. It is noted that more intensed reactio n EGF, EGFR, aFGF, bFGF, FGFR, and VEGF on experimental group compare to that on the control group. 2. Increased reaction is noted on the tumor components compare to that in the stromal tissues. 3. Intensed reaction is noted on the basement membrane adjacent to cancer nest to EGF, EGFR, aFGF, bFGF, FGFR, and VEGF 4. It is noted that intensed positive reaction on cancer pearls, cancer components with hyperactivities, in cancer nest. And at the peIiphelY of cancer nest, diffuse moclerate reaction to EGF, EGFR, aFGF, bFGF, FGFR, and VEGF is notecl This results suggest that EGF, EGFR, aFGF, bFGF, FGFR, and VEGF mJy be effectecl to the growth ancl clevelopment of the squamous cell carcinoma.

Arrùno acid transpoπers play an important role in supplying nutrition to cells and for cell proliferation. System L is a major nutrient transport system responsible for the Na+-independent transport of large neutral amino acids including several essential amino acids. In malignant tumors, a system L transporter L-type amino acid transporter 1 (LATl) is upregulated to support tumor cell growth. In the present study, we have examined the expression and function of system L amino acid transporter in FaDu human pharyngeal squamous carcinoma cells. RT-PCR, real-time quantitative RT-PCR and westem blot analysis have revealed that the FaDu cells express LATl together with its associaω19 protem 4F2hc, whereas the FaDu cells do not express the other system L isoform L-type amino acid transporter 2 (LAT2). 까le uptake of L-(14Clleucine by FaDu cells is Na+-independent and almost completely inhibited by system L selective inhibitor 2-aminobicyclo-(2,2,1)-heptane-2- carboxylic acid (BCH). The profiles of the inhibition of L-[I4Cllellcine uptake by variolls amino acids in the FaDu cells are comparable with those for the LA T1 expressed in Xenopus 。()(찌es. π1e majority of L-[I4Clleucine uptake is, therefore, mediated by LAT1 in the FaDu cells. These results suggest that the transport of neutral amino acids including several essential amino acids in the FaDu human pharyngeal squamous carcinoma cells mediated by LAT1. In addition, specific inhibition of LAT1 by such agents as BCH in pharyngeal squamous cell carcinomas will be a new rationale for anti-cancer therapy.

p16INK4A and p15INK4B tumor suppressor genes are frequently altered in various human tumors. Hypermethylation of the promoter region of p16INK4A and p15INK4B seem to be the major mechanism of inactivation. To determine whether the change in p16INK4A and p15INK4B methylation status occur in oral squamous cell carcinomas (OSCCs) and benign oral epithelial hyperplasias, we analyzed 46 OSCCs and 20 benign oral epithelial hyperplasias by methylation-specific PCR. We also analyzed a subset of the samples for p16INK4A and p15INK4B protein expression by immunohistochemistry. The promoter region of p15INK4B was hypermethylated in 13 specimens of the 15 finally analyzed OSCCs and three specimens of the five analyzed benign oral epithelial hyperplasias. By immunohistochemical analysis, we confirmed the loss of p15INK4B expression of all hypermethylated specimens. The promoter region of p16INK4A was amplified by both an unmethylated- and a methylated-specific primers in just one OSCCs. The remaining specimens including 11 OSCCs and four benign oral epithelial hyperplasias were normally methylated. By immunohistochemistry, we analyzed the loss of p16INK4A expression in seven specimens of the 12 OSCCs and two specimens of the four benign oral epithelial hyperplasias. Except for one OSCC, however, all specimens showing loss of expression were normally methylated. These results suggest that loss of p16INK4A and p15INK4B protein expression play an important role in the development of both OSCCs and benign oral epithelial hyperplasias.

E6 and E7 as two major oncoproteins of HPV can act together to produce several tumors, providing an evidence for the role HPV in oral squamous cell tumorogenesis. It is worthy to detect E6/E7 mRNA expression of HPV in oral carcinoma. The purpose of this study were to detect HPV mRNA in HNSCC cell lines, and to use these results to confirm oral squamous cell carcinoma. Semi-quantitative RT-PCR method for E7 mRNA expression in HNSCC cell lines should play an important role in detecting the early stage of oral squamous cell carcinoma.

MMPs are catalytic enzymes involved in the degradation of extracellular marix, and associated with invasive growth and metastasis of malignant tumors along with angiogenesis. This study was to evaluate the prognostic significance of VEGF expression and microvessel density(MVD) and the relationship between VEGF expression, MVD and MMPs expression in oral squamous cell carcinomas(OSCC). The materials were from 52 cases of OSCC during a period from 1991 to 2001. Clinicopathologic parameters such as clinical stage, recurrence, histologic grade and invasion pattern were evaluated. Immunohistochemical staining for MMP-2, MMP-9, VEGF and CD34 were performed and statistical analyses between clinicopathologic parameters and VEGF expression and MVD were done. MMP-2, MMP-9 and VEGF expressions were noted in 30(57.7%), 21(40.1%) and 38(73.1%) of 52 cases, respectively. MVD was measurable in 35 cases, and cases with increased MVD more than average were 16(45.7%) of 35 cases. There was no significant relationship between clinicopathologic parameters and VEGF expression or MVD in OSCC, which suggests that VEGF expression and MVD can not be regarded as reliable prognostic factors. Cases with less infiltrative growth pattern showed a tendency of increased MVD, which can explain the possibility that neovascularization might be an early event of tumor invasion. Lack of significant relationship between MMPs, VEGF expressions and MVD might be due to limited number of cases, and positive correlation between MMP-2 and VEGF expression suggests that both factors might be involved in the process of angiogenesis in OSCC.

The imbalance between epithelial cell growth and inhibitory factors may cause human epithelial cancer. The dysregulation of growth inhibitory effect of TGF-β1 has been recognized in a variety of carcinomas. This study aimed to investigate the expression of TGF-β1 type II receptors(TβR-II) in the carcinogenesis of oral squamous cell carcinoma(OSCC). Six cell lines established from OSCC in the department of Oral Pathology, Yonsei University College of Dentistry were used. DNA was extracted from harvested cells by phenol-chloroform method. Polymerase chain reaction (PCR) was done with each primer of exon 3, 4, 5, 6 of TβR-II gene. PCR products were inserted to cloning vector (pGEM-T easy vector) and then analyzed to automatic DNA sequencing analyzer. Reverse transcriptase-PCR (RT-PCR) was performed to confirm the mRNA expression of TβR-II gene. Western blotting was performed to detect the expression of the TβR-II protein. As results, a frameshift within a polyadenine region of exon 3 was found in YD-8 cell line. In YD-17 cell line, a missense mutation at codon 238 of exon 4 was found, suggesting the alteration of amino acid from asparagine to aspartic acid. TβR-II mRNA was detected in all cancer cell lines, but it was slightly decreased as compared to that of normal oral mucosal cells. In Western blotting, no TβR-Ⅱ protein was detected in all OSCC cell lines. These results suggested that the altered regulation of TGF-β1 function might play a role in the development of OSCC.

It has been said that amino acid transporters play an important role in supplying nutrition to normal and cancer cells and for cell proliferation. System L is a major nutrient transport system responsible for the Na+-independent transport of large neutral amino acids including several essential amino acids. In malignant tumors, a system L transporter L-type amino acid transporter 1(LAT1) is up-regulated to support tumor cell growth. In the present study, we have examined the function of LAT1 and its expression in the KB human oral epidermoid carcinoma cells. RT-PCR, western blot analysis and immunohistochemical analysis have revealed that KB cells express LAT1 in the plasma membrane together with its associating protein 4F2hc, whereas KB cells do not express the other system L isoform LAT2. The uptake of L-[14C]leucine by KB cells is Na+-independent and almost completely inhibited by system L selective inhibitor BCH. The profiles of the inhibition of L-[14C]leucine uptake by amino acids in the KB cells are comparable with those for the LAT1 expressed in Xenopus oocytes. The majority of L-[14C]leucine uptake is, therefore, mediated by LAT1 in the KB cells. These results suggest that the uptakes of neutral amino acids including several essential amino acids in the KB oral epidermoid carcinoma cells mediated by LAT1. In addition, specific inhibition of LAT1 by such agents as BCH in human oral squamous cell carcinomas will be a new rationale for anti-cancer therapy.