삼백초가 CCl_4 투여로 유발된 rat의 지질대사에 미치는 효과를 알아보기 위해 NCT, CTA, SCT군으로 각각 10마리씩 나누어 NCT군과 CTA군은 0.9% saline을 SCT군은 삼백초 100mg/kg이 되게 2주 동안 매일 복강내에 투여하였다. 15일째에 CCl_4를 CTA군과 SCT군에 투여하였다. 희생시켜서 혈청과 간의지질대사를 관찰한 결과는 다음과 같다. 1. 혈청 중의 AST와 ALT의 활성도에서 SCT군은 CTA군에 비교해서 각각 43.18%, 96.05% 억제되었다. 2. Total cholesterol에서 삼백초를 투여한 SCT군은 NCT군 비교해서 0.95배 증가하였으며, 혈청중의 HDL-cholesterol 보다 0.73배 감소하였다. 3. 혈청중의 TG의 함량은 CTA군이 NCT군에 비해 1.53배 증가하였다. CTA군에 비해 SCT군은 0.93배 감소하였으며, 19.00%로 억제되었다. 4. 간조직 중의 MDA량은 SCT군이 CTA군에 비해 0.85배 감소하였고, 33.00%의 억제율을 보였다.

복제수정란 생산에 있어서 수핵란 내 체세포 주입 후 전기적인 융합은 필수과정인데, 이 과정을 거치는 동안 많은 수의 체세포 주입 난자가 융합에 실패하거나 lysis가 일어나게 된다. 본 실험에서는 한우 체세포를 이용하여 핵이식을 실시한 후 수핵세포질과 응합을 시도할 때 전기융합 방법에 따른 융합율과 배발달율을 검토하고자 실시하였다. 공여세포는 한우 귀 세포조직을 채취하여 0.05% trypsin과 EDTA가 첨가된 D-PBS로 세포를 분리한 후 DMEM

본 연구는 난자의 성숙시간, PHA-P 처리 또는 활성화 방법이 소 미수정란의 탈핵, 재구축란의 융합, 활성화 또는 체외발육에 미치는 영향을 검토하였다. 미수정란은 성숙 후 16∼24시간에 탈핵을 실시하고, PHA-P 처리 또는 무처리된 귀 피부세포를 이식 후 전기융합을 실시하였다. 후자의 경우는 융합 전에 PHA-P로 15분간 배양하였다. 융합란은 A23187과 CHXM 혹은 DMAP의 병용처리에 의해 활성화를 유기하고, 7∼9일간 체외배양하였다. 탈핵율은 성숙 후 16∼18시간에 실시한 경우(70.2∼92.3%)가 성숙 후 20∼24시간(44.3∼3.4%)에 비하여 유의적으로 높았다(P<0.05). M-II기 염색체의 위치는 성숙배양 시간이 길어짐에 따라 제 1 극체와의 간격이 멀어졌다. Donor 세포 혹은 재구축란에 PHA-P를 처리한 경우는 무처리구에 비하여 융합율이 향상되었다(P<0.05). 핵이식배의 분할율 및 배반포 발달율은 A23187+DMAP 처리구에서 78.6%와 32.9%로, A23187+CHXM 처리구에 비하여 유의적으로 높았다(P<0.05). 본 실험 결과는 성숙후 18시간에 탈핵을 실시하는 것이 효과적이며, donor세포 또는 융합 전 재구축란의 PHA-P 처리가 융합율 향상시킬 수 있고, 또한, 융합란을 A23187과 DMAP으로 병용처리 함으로써 난자의 활성화 및 배반포 발육율을 향상시켜, 결과적으로 핵이식기술의 효율성을 증진시킬 수 있을 것으로 사료된다.

hFSH는 α와 β subunit으로 구성된 heterodimer로서 두 subunit의 조합은 활성을 지닌 호르몬의 생산에 있어서 매우 중요한 단계이다. 이 조합과정의 효율을 증대하기 위하여 본 연구에서는 hFSH를 단일사슬의 단백질로 구축하고자 하였으며, 이의 일환으로 각 subunit 대한 cDNA단편을 연결하는 서열로 CTP linker를 도입하였다. 재조합한 hFSH-CTP 유전자는 pseudotype의 retrovirus vector system을 이용하여 CHO 세포와 닭의 배로 각각 전이되었다. CHO 세포에서의 FSH의 생산은 α와 β를 각각 전이한 경우에 비해 hFSH-CTP를 전이한 경우에서 17배 이상 높은 것으로 나타났다. 닭에서는 유전자 전이를 시도한 62개체 중에서 11마리가 부화하였으며 그 중 10마리가 형질전환된 닭인 것으로 RT-PCR을 통하여 확인되었다. 그러나 개체의 혈중 FSH의 생산은 확인하지 못하였다. 이상의 실험 결과를 바탕으로 하여 재조합된 hFSH-CTP는 FSH의 발현에 매우 효율적인 구조로 생각되며, 또한 retrovirus를 이용한 유전자 전이 방법은 형질전환 가금의 생산에 있어서 매우 적절한 방법으로 사료된다.

We used three-dimensional Matrigel culture system to examine the morphognesis of normal and malignant salivary glands cell in vitro including acinar cells(AC), myoepithelial cell(MC), salivary gland adenocarcinoma cells(SGT), mucoepidermoid carcinoma cells(MEC), and immortalized human salivary gland cells(HSG). For this purpose, normal and salivary gland tumor cells cultured in 3-D Matrigel, and characterized histologically and immunohistochemically, compared with same cells grown on monolayer culture and patient tissue from biopsy. 1. In three-dimensional Matrigel culture, HSG cells form acinar structure, SGT cells shows duct like structure, and other AC, MC and MEC cells dont' form any structure , and their morphology was different from that of monolayer cells. 2. Matrigel involved cell proliferation at a similar pattern to cells on plastic monolayer cell cultures, and monolayer cell revealed higher cell viability than that of Matrigel cultured cells. 3. All salivary glands cells on Matrigel or monolayer showed strong PCNA expression, and there is no expression difference in these cells. But some cells including myoepithelial cells in normal and salivary gland tumor tissue showing PCNA lavelling, so there is PCNA expression difference among normal and tumor tissue cells. 4. Actin expression was noted in AC cells on Matrigel, were rare expressed in the other cells except in MEC cells, and was present in myoepithelial cell and ductal cells of normal gland tissue. There is actin expression difference between tissue and cultured cells . 5. S-100 immunoreaction was moderateively positive in MC cells of monolayer culture, myoepithelial cells of normal tissue and pleomorphic adenoma, all cancer cells of mucoepidermoid carcinoma tissue, but significantly decreased in all salivary cells on Matrigel. 6. TGase 2 expression was prominent in MC cells of monolayer and Matrigel cultured, in myoepithelial cells of normal gland and pleomorphic adenoma, epidermoid cells of mucoepidermopid carcioma, and strong reaction in MEC and AC cells of monolayer and Matrigel cultured. 7. Expression of CK in monolayer culture showed strong reaction to CK6 in all sailvary gland cells, and mild reaction to CK10 and CK16 for all salivary cells, CK16 and CK19 expression in monolayer culture was similar to that of Matrigel culture. 8. CK6 and CK10 expression was strongest in AC and MC cells on Matrigel, and CK 4 was negative reaction in AC, SGT, MEC cells, strong reaction in MC cells but mild in SGT cells on Matrigel. Expression of CK was rare in HSG cells compared with other salivary gland cells, CK16 was prominent in SGT cells, CK10 and CK16 showed strongest expression in MEC cells of Matrigel. 9. Monolayer culture of HSG cell shwoing strong reaction to CK6, moderate to CK19 and mild to the others CK, but 3D cultured HSG cells reveal mild expression to CK16, and rare to others CK, intercallated duct in normal gland tissue showing strong to CK19, and mild to the others Ck, so there are CK expression difference in tissue, monolayer and 3-D cultured cells. 10. Monolayer culture of MEC cells represent strong reaction to CK6, mild to other CK, 3-D cells showing increased CK expression including CK6, epidermoid cells and intermediate cells in mucoepidermoid carcinoma tissue reveal positive to CK6 and CK16, mucous cell positive to CK10 and CK19, so Matrigel showed similar CK pattern compared to mucoepidermoid carcinoma tissue rather than monolThese data indicate that the interaction of salivary gland cells with basement membrane is an important factor in salivary gland development and cytodifferentiation, so this model system will be useful to study acinar or ductal differentiation in vitro.ayer cultred.

be involved in the development of oral SCC. If we compare the morphologic features of NHOK to IHOK according to calcium concentration by TEM, IHOK have been gained wide acceptance as a model system for HPV-linked oral carcinogenesis. We already have established immortalized oral keratinocytes(IHOK) transfected by E6 and E7 gene. The purpose of this study were to examined the ultrastructural features of cultured NHOK, IHOK, and HN4 oral squamous cell carcinoma cell line, and to apply these results to oral carcinogenesis in the future. NHOK from healthy retromolar pad was primarily cultured at 37oC and 5% CO2. IHOK, and HN 4 cell line which were cultured under 0.15 and 1.2mM Ca++ of KBM bullet kit. For transmission electronmicroscopy(TEM), under preconfluency, and after 3 days of postconfluency under 1.2mM Ca++, cultured NHOK, IHOK, and HN4 cell line were immediately fixed in 2.0% glutaraldehyde in 0.1M cacodylate buffer(pH 7.4) at 4OC for 1h. The ultrathin sections were stained with uranyl acetate and lead citrate, and examined by TEM. The obtained results were as follows. 1. TEM of cultured NHOK under 1.2mM Ca++ showed increased tonofilaments, and vaculated ovoid cells with cornified envelope, while cultured IHOK showed prominent microvilli, unilateral desmosome in microvillus, and tonofilaments. 2. TEM of HN 4 cell line sowed numerous microvilli, increased N/C ratio, and lateral desmosome in microvilli under 0.15mm, while under 1.2mM well forming desmosomes. From the aboving results, under high calcium cultured IHOK showed less tonofilaments than that of cultured NHOK, while cultured IHOK, and HN 4 cell lines showed more increased desmosomes under high calcium. It was suggested that the ultrastructural changes of cultured IHOK would be accepted as intermediate stage cells for studying oral carcinogenesis.

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.

한우의 수정란을 보존하기 위하여 고급육계통과 다유계통의 공란우를 선발하여 과배란처리에 따른 황체의 반응, 황체수에 대한 채란된 수정란의 수 그리고 채란 수정란수에 대한 이식가능한 수정란 생산에 미치는 영향을 조사하였다. 다유계통의 공란우는 이유시 체중에 대한 육종가 상위 20%이내, 고급육 계통의 공란우는 육질에 대한 육종가 상위 20%이내의 종빈우를 선발하였다. 과배란처리 방법은 발정후 11일차에 PEG 30% FSH를 견갑부에 피하주사로 1회 투여하

한우의 유전자원을 보존하기 위하여 정액동결 시험을 수행하였으며 고급육계통은 육질에 대한 육종가 상위 10% 이내의 자손에서 선발하였고 다 유계통은 어미의 이유시 체중에 대한 육종가 상위 10%이내의 자손에서 선발하였으며 2개 계통에서 종모우 총 13두를 선발하여 공시하였다. 정액채취는 인공질법으로 실시하였고 의빈대에 수소를 계류하고 채정대상우를 승가시켰으며, 3회 가승가후에 채정하였다. 채정후 10분 이내에 실험실로 옮겨와 검사항목을 조사하고 37에서

본 연구는 착상전 이배체 단위발생 돼지난자를 체외 배양시 우태아혈청 (FBS), 우혈청 알부민 (BSA) 및 상피세포성장인자 (EGF)를 배양액에 첨가하였을 때 배반포, 총 세포수, 세포사멸 및 세포사멸에 관여하는 유전자의 발현을 조사하고자 수행하였다. 0.4% BSA를 배양액에 첨가하였을 때 2세포기 단위발생 난자의 배반포까지의 발달율이 증가되었다(P<0.01). FBS는 배반포의 총세포수를 감소시 켰고 세포사멸을 증가하였다(P<0.01). 그리고 EGF는 BSA가 존재하는 조건하에서 배반포의 총세포수를 증가하였는데 EGF와 BSA가 각각 단독으로 존재할 때는 이런 작용이 없었다. 세포사멸도 이와 이슷한 경향을 보였는데 EGF와 BSA가 각각 존재할 때에는 비처리군과 차이가 없었지만 함께 존재할 때에는 세포사멸을 감소시켰다. RT-PCR의 결과에 의하면 EGF는 BSA가 존재하는 배양액에서 Bcl-xL 유전자의 상대적 발현량을 증가시키고 Bak 유전자의 상대적 발현량에는 영향을 주지 않는 과정을 통하여 세포사멸을 감소시키는것 같다. 반면에 FBS는 Bcl-xL의 발현량을 감소시키고 Bak 유전자의 상대적 발현량을 증가시킨다. 이러한 결과는 세포사멸에 관여하는 유전자의 발현은 배양액의 첨가물에 따라 유의적으로 영향을 받으며, 체외배양시 배아의 초기발달에 관여함을 시사한다.

A case of odontogenic ghost cell tumor (OGCT) with clear cell components occurred in the mandible of a 63-yearold man. The tumor revealed ameloblastomatous type epithelial components together with clusters of ghost cells and dentinoid juxtaposed to the odontogenic epithelium. In addition, some areas of the tumor tissue showed sheets and islands of clear, glycogen-rich epithelial cells, separated by a thin fibrous connective tissue stroma. Both ameloblastic and clear cells showed positive immunoreactivities for cytokeratin 19 and AE1/3. It is not known whether this tumor represents a clear cell differentiation of a preexisting OGCT or a separate and distinct neoplasm derived de novo from odontogenic epithelium. This tumor was preferred the term clear cell odontogenic ghost cell tumor, which captures the clear cell components, one of the tumors most prominent distinguishing features

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.

Established SGT cell line from human submandibular gland adenocarcinoma was used to study the TGase expression on a cellular level in vitro. Transglutaminase 2(TGase 2) is assoacitated with apoptosis, GTP binding protein, and cell marix interaction. The role of TGase 2 in salivary gland tumors is not clear yet. The pupose of this study were to examine the TGase expression of SGT cell line compared to other tumor cell lines, and to apply these results to the pathogenesis of salivary gland tumor. TGase enzyme assay of SGT, SCC-15, HN 4 and HeLa tumor cell line was 3 times repeated, and calculated. Immunoslot blot for semiquantitative protein analysis was done. The obtained results were as follows. 1. SGT cell line showed the highest TGase 2 enzyme activity(about 6-16 folds) irrespective of pre or postconfluency. 2. HN 4 cell line showed the highest TGase 1 enzyme activity(about 2-3 folds) irrespective of pre or postconfluency. 3. Under postconfluency TGase 1 induction was not induced, but slightly increased in all tumor cell lines. 4. TGase enzyme activity in all tumor cell lines was accompanied with TGase protein formation. From the aboving results, the higher TGase 2 expression of SGT cell line suggested that they would come from submandibular ductal cells and have a important role in the pathogensis of salivary gland tumors.

Cell adhesion is used as a parameter to evaluate the biocompatibility of dental implant and also affected by the surface form of dental implant. Most study have showed different cell reaction by the composition and the surface morphology of implant. Therefore it is thought that the osteoblastic activity would be affected by the surface roughness and composition of implants. This study was performed to evaluate the biological activity and morphological change of normal human osteoblastic cells(NHost) depending on the variations of implant surfaces. We used grade 2 titanium disks which were being air-blasted with TiO2 50 ㎛, 110 ㎛, 250 ㎛ powder by 3psi compressed air and non-blasted as control. We evaluated and compared morphologic change, adhesion assay, and Ca, P, ALP concentration of NHost in vitro. The obtained results were as follows. 1. In the growth curve, although the growth of experimental groups were lower than that of the group of NHost only, there was no significant difference between each groups. 2. Inverted microscopic findings showed NHosts in early stage of each group were adherant perpendicular to the titanium disk and the multilayered NHosts were attached with various directions after 4 weeks. 3. Scanning electron microscopic(SEM) features showed that NHosts in all groups seemed to be attached multilayered and connected with each process after 2 weeks. 4. NHosts' processes were found by the SEM after one day culture. The cell adhesion of experiment group was higher than that of control group. 110 ㎛(the 3rd group) showed prominant process of NHost on the titanium disk surface. 5. Although the concentrations of Ca, P and ALP were gradually reduced, ANOVA analysis of each groups were partially different, and ANOVA analysis of 4th group were significantly different with others. From the aboving results, NHosts cultured on the titanium disks showed similar morphological change and cell proliferation. There were partially differences in each group except the 4th group, and the 4th group were significantly different with other's in biological activity. We thought that biological activity and adhesion of NHost cell on titanium had been affected by the variation of the titanium surface roughness.

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.