많은 사람들이 불멸에 관심이 있지만, 죽은 이의 불멸은 사회과학에서 지금까지 거의 다루어지지 않은 주제다. 본 연구에서는 죽은 이들의 도덕성 판단과 마음지각에 따라 불멸지각이 달라지는지를 검증하였다. 연구 1에서는 도 덕적으로 악한 사람, 선한 사람 및 도덕성과 연관되지 않은 인물에 대한 글을 제시하고, 도덕성 판단, 마음지각 및 불멸지각의 관계를 분석하였다. 그 결과 도덕적으로 선한 인물에 대한 불멸의 지각정도가 악한 인물 또는 중립적인 인물에 대한 불멸의 지각정도보다 더 높게 나타났다. 또한 인물의 도덕성 판단과 마음지각은 불멸을 지각하는 정도 와 유의한 상관관계가 나타났다. 연구 2에서는 도덕적으로 선한 로봇, 악한 로봇 및 도덕성과 연관되지 않은, 기능적 로봇에 대한 글을 제시하고, 로봇에 대한 마음지각 및 불멸지각의 관계를 분석하였다. 그 결과 로봇의 도덕성판단과 마음지각이 높을수록 로봇의 불멸을 지각하는 정도가 증가하였다. 이 결과는 도덕성 판단과 마음지각이 인간-로봇의 상호작용에서 어려움을 극복하는 변인으로 기여할 가능성을 보여준다.

Immortalization is an essential process of the transformation of cells to a neoplastic growth. High risk human papillomavirus (hrHPV) infection has been the major cause of head and neck squamous cell carcinoma (HNSCC). The aim of this study was to search for a novel pathway causing immortalization in HPV16 E6/E7 transfected immortalized oral keratinocytes (IHOK). hrHPV integration sites were identified through DNA sequencing. HPV16 E6/E7 genes were integrated into 1q32.2, 12q21.2, 15q15.2, and 19q13.43 in IHOKs. Array-CGH was conducted to examine the deranged sites of the genes of IHOK. Of the 587 amplification genes, 70 genes were resided on chromosome 20. We selected PLAGL2 and MAPRE1 as the most amplified genes. PLAGL2 and MAPRE1 mRNA showed higher expression in IHOK than in normal keratinocytes. Knockdown of MAPRE1 significantly reduced telomerase activity. The analysis using a public database substantiated our data, showing the amplification of chromosome 20 and MAPRE1. In conclusion, our results suggest that MAPRE1 could play a crucial role in activating telomerase activity in hrHPV-infected cells. This finding may provide basic data to develop a novel target therapy for hrHPV-related HNSCC.

본 논문은 『히브리 선율』 집에 들어있는 바이런의 초기시가 영혼과 하느님 의 불멸성에 대해 어떠한 종교적인 고찰을 보여주고 있는가를 다룬다. 종교시는 1810년대에 출간되어 시인의 이전 작품 『한가한 시간들』에서 보이는 자연의 항구성을 전개시켜 인간의 영혼과 전능하신 신의 영역까지 다루고 있다. 바이런 은 『히브리 선율』 중 다섯 편의 시를 통해서 인간 영혼의 불가침성을 암시해주 고 있으며 이는 그의 후기시와 여러 수상록에서도 이 부분에 대한 시인의 확고한 믿음을 보여주고 있다. 이러한 바이런의 초기에 나타난 종교적 명상의 씨앗 들은 시인을 1813-1814년에 지은 작품과 1823-1824년에 사이에 지은 『천지』라 는 작품을 토대로 성상파괴자라고 혹평했던 당대 비평가들의 태도가 왜곡되었 음을 여실히 보여주고 있다. 본 논문은 『히브리 선율』이라는 작품을 통하여 이 러한 당대의 비평시류에 반대되는 다양한 예증들을 통하여 바이런이 이들과 상 반되는 종교적 시각을 드러냄을 보여주고자 한다.

Cellular microenvironment is an essential issue for regulating epithelial characteristics through the alteration of intricate signaling pathways and intercellular communications in different cell types. Thus, microenvironment influences tumor initiation, progression, and metastasis. This study aimed to investigate the relationship between microenvironment and epithelial property in HPV16 E6/E7-immortalized human oral keratinocytes (IHOKs). To investigate characteristics of IHOK cultured in different media, two media were used, which included keratinocyte growth media (KGM), F-medium composed of 3:1 ratio of DMEM and F-12 (P media) supplemented with 10% FBS and 1% penicillin/streptomycin. Proliferative property and invasive and migratory activity were observed. As results, proliferating activities of IHOK in different culture condition were changed. Likewise, migratory and invasive activities were also different depending on media types. These results suggest that cellular microenvironment can affect modification of biological properties of epithelial cells.

Plasminogen activators(PA) such as urokinase(uPA) and tissue type plasminogen activators(tPA), physiologically catalyze the conversion of the plasminogen to the wide spectrum proteinase plasmin. Because uPA and tPA are involved in cell growth, differentiation and migration of oral cancer, oral epithelial carcinogenesis including transformation of precancerous lesion into oral squamous cell carcinoma with PA is very interesting. It is important to prevent precancerous condition from transfoming into oral squamous cell carcinoma by the inhibitory effect of various drug. It is well known that cyclosporine A(CsA) as immunosuppressive properties exerts anti-cancer effects. Recently it is widely accepted that cultured immortalized oral keratinocyte (IHOK) is considered as an intermediate stage of oral carcinogenesis and used as precancerous condition in vitro. Thus it was thought that it might be interesting to investigate CsA effect on PA expression of IHOK. IHOK was cultured under KBM bullet kit at 37℃ under 95% CO2 incubator. Subconfluent IHOK cells was treated at different CsA concentration. uPA and tPA protein expression from cultured IHOK cell line has been detected by ELISA analysis in the CsA-treated samples. uPA expression of IHOK was higher than that of NHOK, while tPA was similar to that of NHOK. After CsA treatment, CsA might not effect the expression of uPA of IHOK, while showed a little effect on tPA of IHOK. It suggested that CsA had no effect in uPA expression of IHOK although uPA could be used as a marker for precancerous lesion.

Oral squamous cell carcinoma (SCC) is one of the leading causes of cancer mortality worldwide. It is generally thought that adjuvant chemotherapy provides modest prolongation of survival in various carcinoma. Docetaxel (Taxotere, TXT) play a significant role in the treatment of various solid tumors of epithelial origin. CsA (immunosuppressive drug) was widely used as adjunct for the treatment of cancer. Thus, it is important to pursue the apoptosis of IHOK and oral SCC induced by TXT combined with CsA related to the pathogenesis of oral SCC. But TXT combined CsA effect on IHOK and oral SCC remains unclear. After cultured IHOK and HN 22 oral squamous cell carcinoma cell line treated by 10 nM TXT and 1 μM, and caspase inhobitor, respectively, apoptosis index, cytochrome c and caspase-3 -8, -9 mRNA expression by RT-PCR, and procaspase-3 protein amount by immunoslot blotting was prepared. The purpose of this study were to examine the TXT-induced apoptosis pathway via caspase activation by CsA enhancement, and to apply these results to an effective therapeutic treatment plan for oral SCC by TXT combined CsA . 10 nM TXT showed about 60%, 55% celluar apoptosis of IHOK and HN 22, cell line, respectively, while CsA alone did not induce apoptosis in IHOK and HN 22 cell line. 1 μM CsA combined with 10 nM TXT increased apoptosis in IHOK and HN 22 cell line through caspase-3 and cytochrome c mRNA expression, while could not effect on caspase-8 and -9. Caspase inhibitor suppressed apoptosis of IHOK and HN 22 cell line induced by a combination of 1 μM CsA and 10 nM TXT. Immnoslot blotting showed procaspase-3 activation by a combination 1 μM CsA and 10 nM TXT, while caspase inhibitor inhibited activation. It suggested that a combination of CsA and TXT might induce increased apoptosis of IHOK and HN 22 oral squamous cell carcinoma cell line through caspase-3 activation. This treatment with a combination of TXT and CsA may be an effective therapeutic strategy for oral squamous cell carcinoma

Human papillomavirus (HPV) has been classified as one of the causing factors of head and neck squamous cell carcinoma (HNSCC). However, little is known about HPV-related carcinogenesis in HNSCC. The purpose o f this s tudy i s to characterize immortalized human oral keratinocyte (IHOK) transfected by HPV16 E6/E7, IHOK/hcdk4 (IHOK transfected by pLXRN-hcdk4) and IHOK/hcdk4/hTERT (IHOK transfected by pLPC-hTERT-hcdk4) to reconstitute HNSCC in vitro. Conclusively, we established a new immortalized cell lines, IHOK/hcdk4 and IHOK/hcdk4/hTERT, to understand multistep carcinogenic process of oncogenic HPV16 E6/E7 in HNSCC.

Intracellular reactive oxygen species(ROS) produced in a various pathologic state was known to intermediate many cellular response such as inflammation. Recently, low level light irradiation by HeNe laser used in many clinical field could improve inflammatory state by scavenging intracellular ROS through photo-detachment/dissociation process. The purpose of this study is to investigate the differential effects of blue and red light irradiation on ROS scavenging effects. Immortalized human oral keratinocyte HaCat cells were used. Phorbol 12-myristate 13-acetate(PMA) was treated for inflammation. Red(635nm) and blue(470nm) light irradiation was carried out. To asses the intracellular ROS by light irradiation, confocal microscopic and flow cytometric assay with DCF fluorescence for total ROS and ESR spectrometry of DMPO-O2 - for superoxide anion were caried out. And microarray was performed for mRNA expression level. Released intracellular total ROS in PMA treated HaCat cell lines was dissociated efficiently by red light irradiation, while blue light irradiation did not. Rather, blue light irradiation increased ROS formation. For superoxide anion generated the first synthetic form of ROS, red light irradiation reduced its amount but blue light irradiation did not. In the mRNA expression in line with cyclooxygenase(COX) pathway, prostagrandin endoperoxide synthase 1(PTGS 1), prostagrandin endoperoxide synthase 2(PTGS 2) and phospholipase A2(PLA2) were increased by both light irradiation and they were decreased as time flows. And genes associated with ROS releasing, mRNA expressions of tumor necrosis factor receptor (TNFR) and interleukin 1beta(IL1B) were increased by 1 hour red light irradiation but did not by blue light irradiation. As a result, red and blue light irradiation showed different response in affecting the level of ROS. These findings indicate that red light rather than blue light is more useful for anti-inflammation in clinical field

Considering the great potential of iron chelators at inhibiting the proliferation of tumor cells, in order to determine the molecular and biological basis for the effects of iron chelator in oral cancer, we investigated the effects of iron chelator, desferrioxamine (DFO), on the gene profiling analysis of immortalized human oral keratinocytes (IHOK), and oral cancer cells (HN12), using the cDNA microarray. We identified 46 clones cDNA exhibiting more than 2 fold overexpression in DFO treated IHOK and HN12 cells, and 94 cDNA reveal more than 2 fold down-regulated expression. Examination of gene expression that differs between DFO treated vs. control IHOK and HN12 cells apprear to be related to : cell cycle regulator, cell growth and apoptosis, signal transduction and stress. p21 for cell cell cycle factor was upregualted, and cyclin-cdk gene was decreased expression, so we observed cell cycle arrest in DFO treated IHOK and HN12 cells. In tumor growth, we have identified downregulation of hemidesmosomal protein (bullous pemphigoid antigen 1) and epiregulin expression in DFO treated IHOK and oral cancer cells. Signal transducers including mitogen-activated protein kinase-activated protein kinase 5, serine/thereonine kinase 6 were downregulated with DFO treated cells, suggesting the DFO regulates the p38 MAP kianse pathway in immortalized and maignant oral keratincytes. In conclusion, we have demonstrated the high-throughput utility of cDNA array hybridization in parallel to the gene expression analysis to identify genes that are expressed differentially in DFO treated with immortalized and malignant oral keratinocytes. The differentially expressed genes identified here should be informative in DFO-induced anti-cancer effects.

Cytokines play a vital role in the host immune response by regulating the development and function of im munocompetent ce11s One immunomodulatory agent that has received attention in oncology research recently is interleukin - lO(IL-lO). IL-IO inhibi ted tumor antigen presenta tion and induced energy in T lymphocytes that had been s timu lated by autologous MHC class II positive tumor ce11s Patients with head and neck cancer have been shown to exhibit profound irnmunosuppression. The mechani sm by which tumor ce11s alter immunological function in the host is poorly understood. Recently. production of biological active IL- IO was confirmed in ovar‘ian cancer, melanoma, skin cancel‘ & head and neck cancer, suggesting that IL- lO reduces the function of tumor infiltrating lymphocytes and contributes to the tumor growth. IL-IO expression has not been examined extensively in human oral cancer and has not yet been cla rified. The purpose of t his study were to investigate IL-IO mRNA and protein expression in NHOK, IHOK and oral squamous ce11 carcinoma(OSCC) ce11 line by RT-PCR and irnmunoslot blotting, and to apply its results to examine its thera peutic significance for oraJ cancers. Cultured NHOK showed a lower level of IL-IO mRNA and protein expression than cultured IHOK and HN 22 OSCC cell line under pre and postconfluency. HN 22 OSCC cell line under pre and postconfl u ency. showed the highest level of IL-I0 Cul tured IHOK showing a intermediate expression of IL- IO could be as a vaJ u a bJe marker for oral carci nogenesis ste p. During the terminal differentiation of a11 the ce11 lines, IL- IO ex pression was significantly unchangeabl e. IL- IO mRNA expression of a11 the ce11 lines was consistent with IL-10 protein expression. It suggested that IL- lO expression might play an important role in oral carcinogenesis and IHOK could be a valuable marker for oral carcinogenesis step. And aJso IL- 10 related gene may be future targets for gene discovery and possi bJy therapeutic intervention

We have examined the effect of NO donor, S-nitl‘ oso-N-acetyl-DL-penicillamine(SNAP) on heme oxygenase-1 (HQ-l) ex pression in human oral immortalized & malignant keratinocytes, and investigated in the control of keratinocyte proliferation evidence tha t HO-1 cou ld be involved in a low dose of NO, NO inhibitor, HOinducer, and HO inhibitor medi ated cytoprotect ion against cytotoxi city induced by a high dose of NO Oral keratinocyte growth inhibitory or anti-proliferative effects were exerted by with SNAP and hemin in a dose- and cul tivation time dependent manner The level of HQ-1 protein was increased in all cell types after exposure hernin dose, and the hemin induced HQ-1 protein achieved at higher maximum level by 12 hrs in all kind of cells , The pretreatment of cells with 0, 2 μ M SNAP reduced 1 mM SNAP-induced death in IHOK and HN4 cells , These cytoprotective effects on high dose of NO induced HQ-1 expresion and cell ular toxicity were blocked by low dose of SNAP, HCB, and ZnPP IX supporting the involvement of HQ-1 in high dose NO induced growth arrest or cell death, But these cytoprotection pattern is different from immortalized and malignant keratinocytes , These results indirectly demonstrate that HQ-1 could be involved in cytoprotection by NO priming against high dose NO induced cytotoxicity in immortalized and maigla nt oral keratinocytes, Thus, HQ-1 might be an important cellular target of NO donor, with clinical implications for the pre vention of inJlammatory di seases and anti-tumor immunity

Pro-inflammatory cytokines are important mediators of cutaneous cellular activities during many oral mucosal diseases. IHOK culture model transfected by E6/E7 genes provide further evidence for the role of HPV in tumorogenesis. It is interesting to investigate cytokine expression of immortalized human oral keratinocyte(IHOK). The purpose of this study were to analysis cytokine mRNA expression levels of NHOK and IHOK by RT-PCR. IHOK showed about 5 fold increases of IL-6 compared with NHOK, while TNF-α was the lowest. It suggested that immortalization of NHOK with E6/E7 could result in elevated expression of IL-6, and IHOK be in the intermediate stage of oral carcinogenesis.

Oral squamous cell carcinoma is the 1st most common malignancy in oral and maxillofacial area. HPV 16 has been strongly linked to progression of cervical carcinoma. E6 and E7 as a small DNA virus encoding two major oncoproteins of HPV 16 can act together to produce efficient immortalization of primary human epithelial cells. Thus it is important to pursue the development of Immortalized human oral keratinocyte(IHOK) culture model which could be related to the pathogenesis of oral squamous cell carcinoma. If we establish IHOK transfected by E6/E7 genes, IHOK will be accepted as a model system for HPV-linked oral carcinogenesis. The purpose of this study were to culture primary normal human oral keratinocyte(NHOK), and to establish IHOK for studying oral carcinogenesis in the future. NHOK was primarily cultured under normal culture condition, and transformed into IHOK by transfection of E6/E7 genes. After 100 passages depend on Ca++ condition, cultured IHOK was confirmed by growth curve, cornified cell envelope measurement, TGase 1activity, mRNA detection, tumorogenecity and anchorage independence assay. After 100 passages, cultured IHOK showed most basal cell and monolayer of polyhedral cells under 0.15mM Ca++, and small area of stratification and flattened epithelial cells with irregular border under 1.2mM Ca++. The cultured IHOK showed relatively resistant growth under high calcium condition. The E6/E7 mRNA was detected in cultured IHOK by RT-PCR. During the terminal differentiation in cultured IHOK, increased insoluble cornified cell envelope formation was accompanied with induction of TGase 1 activity. But the cultured IHOK showed less CEM and TGase 1 activity than those of cultured NHOK. Cultured IHOK showed non-tumorogenecity, but slight anchorage independence. We had developed a technique to transform NHOK into IHOK by transfection of E6/E7 genes. Cultured IHOK was established as intermediate stage cell to study the pathogenesis of human oral squamous cell carcinoma.

Cultured normal human oral kera tinocyte(NHOK) & inunortalized human oral keratinocyte(IHOK) provide a valuable model in ce llular proliferation and differentiation after proper stimulation , And it is interesting to study these estab lished cell lines esca ping normal control on their growth and differentiation, SPRR1 is induced during t erminal differ entiation 0 1' human epiderma l kerat inocytes but is rarely in anaplastic cells of keratinocyte origin, But SPR1 expression has not yet been explained during differ entiation uf NHOK and Lransformed oral keraLinocyLes , The purpose of this study were to examine mHNA and protein expression of SPR1 in response to a known differentiation signal, calcium conc in NHOK, lHOK a nd oral SCC ce ll line(HN 4) , and to apply these results for investigating the molecular mecha nisms of tra nsformed cellular differentiation , Primary cultured NHOK, established IHOK and HN 4 cell line were cul tured in KBM bullet kit Preconfluency of NHOK as control group was used Under O, 15mM Ca++ conc(Precon, Postcon) , and 1, 2mM Ca++ conc(Pos tcon)‘ the insoluble final pellets were measured fo1' cornified cell envelope measurements, and RT- PCR for SPRR1 mRNA meas urement, and immunoblotting for SPRR1 protein measurements in tripli cate , resp ectively , The terminal different ia tion of cu ltured NHOK and IHOK was depend on calcium concentration, while HN4 cell line was not SPRR1 mRNA and protein expression of cultured NHOK showed the highest among cultured IHOK & HN 4 cell line in hi gher ca lcium condition , SPRR1 mRNA and protein expression of cultured IHOK showed higher‘ than tha t of HN 4 cell line in hjgher cacium condi tion , SPRH1 was expressed in differentiation of NHOK and IHOK t ransfected by E6/E7 genes but ra rely expressed in malignant oral keratinocytes , It suggested that SPRR1 ex pression as kera tinocyte terminal diff‘erent ia tion marker involved in cellular cornification would be differentially effected by immorta li zation and ca rcinogenic transforma tion

구강암의 발생원인 중 하나인 인간유두종바이 러스(HPV)로 불멸화시 킨 구깅 각화세 포(1 HOK) 외 정상 인긴 구강각화세 포 (NHOK) 의 표지자를 비 교 연구하는 것은 정상과 그 전암냉소의 생화학적 및 세 포회 학적 띤호l을 평가히는 적젤힌 모탤이지띤 떤 구된바 많지 않았다 본 연구는 구강 각화 세 포의 형 질전환을 조절히는 분자적 상태를 연구하기 위 해 익 6000711 의 유전자가 pri nt된 cONA mì croarray를 이용하여 인간 정상 구강각화세 포(NHOK) 와 HPV16으로 불띨회한 각화세 포(J HO K) 간에 유전지 발현을 비교하였다， NHOK외 IHOK세포는 96%의 유전자가 유사한 발현을 보였으며 2배이 상의 발현을 보이 는 경우-는 NHOK가 IHOK에 비해 85개 유전지가‘ IHOK가 쩌OK에 비해 1477H 의 유전자 발현이 u p-regul at i on되 어 총 4%미 만이 발현차이를 보 였고 반복된 hybridì zation 으로부터 얻은 선택된 spot의 Pearson 싱관계수는 074 에서 091로 냐티 났다 NHOK외 IHOK간의 유전자 발현을 기능별로 분석한 결과 몇 가지 주요 발현 유전자 그룹을 확인하였는데 세포부착 및 인식 세포주기 초칠인지 세 포자떨사， 전사인지- 성장인자 및 수용기 ， 세포골격 및 세 포외기질 단백 . 신호진달 조절자 및 기 타 그룹으로 분류할 수 있었다 IHOK에서 는 세 포인식 인자 중 endothelin 1, collagen IV, fibronectin , SPR1 이 발현증가 되었고 CK7, POG1"2, F'G1"1"R2가 세 포골격 및 성장인지중에서 upregulation 되었지만 신호전달 인지중 발현증가된 것은 없었고 동일힌 유전자를 나타내 는 cJ ustered gene map을 그릴 수 있었다 따라서 이러힌 Illicroan‘ay를 이 용한 분자학적 표지자 얀구가 구강 임 빌임과정 에서 유전지발현 을 확인히는데 큰 도웅을 줄 수 있을 뿐 아니라 유전자 조절에 의힌 진단 에 후. 치 료의 정획성을 개선시 킬 수 있으리리 여겨진다

Studies to evaluate distribution of markers in normal keratinocyte and their immortalized keratinocyte are appropriate to evaluate the normal and preneoplastic lesion of oral cancers as biochemical and cytochemical changes associate with tumorigenesis being not completely understood. Complementary DNA microarray containing 6000 sequence -verified cDNA elements was used to systematically characterize the variation in gene expression patterns of NHOK cells vs. immortalized keratinocyte by HPV16 E6-E7(IHOK). Examination of gene expression that is 85 clones cDNAs exhibits greater than 2 fold overexpression in NHOK probes relative to IHOK probe, 147 cDNAs reveal greater 2 fold overexpression in IHOK relative to NHOK probe.The high similarity in gene expression (96.5%) between IHOK and NHOK cells suggests that only an additional 232/6720 (3.5%) of the genome is differentially gene activated during HPV16 immoratlized keratinocyte growth and differentiation. Examination of gene expression that differs between NHOK and IHOK cellsapprear to be related to : cell adhesion & recognition, cell cycle regulator, apoptosis, transciption factors, growth factors and therir receptors, cytoskeletal and extracellular matrix proteins, signal transduction modulators and effectors, and miscellaneous. The gene expression of cell recognition factor such as endothelin 1, collagen IV, fibronectin, and SPR1 in IHOK were upregulated. Distinct or duplicated cDNA clones representing the same gene were typically clustered in adjacent rows in the clustered gene map. Therefore the differentially expressed and identified genes should be informative in studying oral epithelial cell carcinogenesis and such studies should foster the research of molecular markers allowing to assess the phenotypeof malignant epithelial tumor.

Cervical carcinoma is the 1st most common malignancy in korean females. HPV have been strongly linked to progression of cervical carcinoma. E6 and E7 as a small DNA virus encoding two major oncoproteins of HPV can act together to produce efficient immortalization of primary human epithelial cells, providing further evidence for the role of HPV in tumorogenesis. It is important to pursue the development of Immortalized human epithelial keratinocyte(IHEK) culture model which could be related to the pathogenesis between cervical and oral carcinoma. If we establish IHEK transfected by E6E7 gene, IHEK will be accepted as a model system for HPV-linked cervical carcinogenesis. The purpose of this study were to culture primarily normal human epithelial keratinocyte(NHEK), and to establish IHEK for applying these results to cervical and oral carcinogenesis in the future. The obtained results were as follows. 1. After 7-9 passages, cultured NHEK was almost senesce and disappeared, but cultured IHEK showed most basal cell and monolayer of polyhedral cells under 0.05mM Ca++, while small area of stratification and flattened epithelial cells with irregular border under 1.2mM Ca++. 2. The cultured IHEK showed relatively resistant growth to high calcium condition. 3. The mRNA E6E7 in cultured IHEK by RT-PCR was detected. 4. During the terminal defferentiation in cultured NHEK and IHEK, increase of insoluble cornified cell envelope formation was accompanied with induction of TGase 1 activity. But the cultured IHEK showed less CEM and TGase 1 activity than those of cultured NHEK. 5. Cultured IHEK showed non-tumorogenecity, but week anchorage independence. From the aboving results, we have developed technique to transform NHEK into IHEK by transfecting cells with E6E7 gene. Cultured IHEK was established as intermediate stage cell for studying the pathogenesis of human cervical carcinoma.

The study of cornified cell envelope has been used to investigate the differentiation factors and to advance oral carcinogenesis, CE of human oral keratinocytes are in wet condition as saliva containing many proteases, growth factors, and many kinds of bacteria, The analysis of CE in Immortalized human oral keratinocyte(IHOK) derived from normal human oral keratinocyte(NHOK) will be used to study the pathogenesis of oral squamous cell carcinoma, The purpose of this study was to analyze the amino acid component derived from CE of cultured NHOK and IHOK, It will be helpful to study the role of transfected E6/E7 gene in forming CE, and to examine the pathogenesis of oral squamous cell carcinoma, After primry culture of NHOK, IHOK were cultured in KBM bullet kit at 370C under 95% C02 incubator, Growth curve according to calcium concentration, cornified cell envelope measurement(CEM), and protein chemistry for amino acid component of CE were done(Mena :f::SD) , respectively. The obtained results were as follows, lHOK showed small areas of stratification, more compact, with irregular border and tightly apposed cells in 1,2 mM Ca++, Cornified cell envelope exhibited an aggregated group of empty space surrounded by the remained cell membrane, During the terminal differentiation in cultured NHOK and IHOK, insoluble cornified cell envelope formation was increased, CEM of NHOK was about 4 folds than that of lHOK under high calcium, Amino acid component of both groups showed Pro/Glu(SPR) , Gln/Glu(lnvolucrin) , and Gly(Loricrin) in descending order, From the aboving results, ít was suggested that when the terminal dífferentiation in cultured NHOK and IHOK, major amino acid component of CE in cultured lHOK was the same to that of cultured NHOK, It was thought that E6 and E7 gene should be involved in preventing the differentiation and proliferation of IHOK from making CE,