• 대한한의학회지
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• 제20권1호
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• pp.91-105
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• 1999

The effects of Yinjin and Yinjinsaryongsangagambang on a DNA damaging agent, etoposide-induced apoptosis, cell viability, cell cycle progression, and mRNA expression of apoptosis-related genes of human hepatocyte cell line HepG2 were investigated using tryphan blue exclusion assay, MTT assay, flow cytometry, immunocytometric analysis of PCNA, and quantitative RT-PCR analysis. MTT assay showed that Yinjin and Yinjinsaryongsangagambang increases cellular viability of HepG2 cells in a dosage-dependent manner. Stimulation of cell cycle progression by Yinjin or Yinjinsaryongsangagambang was detected by flow cytometric analysis of the DNA content and immunocytometric analysis of PCNA expression. A significant reduction of a DNA-damaging agent, etoposide-induced apoptosis were found in both Yinjin and Yinjinsaryongsangagambang-treated cells in dosage-dependent manner. In overall, 3-fold reduction of apoptosis was recognized in $10.0\;{\mu}g/ml$ of Yinjin or Yinjinsaryongsangagambang-treated cells compared to untreated cells. Although the difference is not significant, Yinjinsaryongsangagambang showed slightly higher effect on the inhibition of apoptosis than Yinjin. From flow cytometric analysis of apoptosis, while 39.9% of untreated cells showed etoposide-induced apoptotic cell death, only 19.6% or 17.4% of Yinjin or Yinjinsaryongsangagambang-treated cells were fond at apoptotic sub G1 phase, respectively. Interestingly, strong induction of Gadd45-mRNA was observed from Yinjin or Yinjinsaryongsangagambang-treated cells. However, no changes in expression levels of p53 and Waf1 were detected, demonstrating that induction of Gadd45 mRNA expression by Yinjin or Yinjinsaryongsangagambang occurs by p53-independent mechanism. Marked mRNA inductions of two apoptosis-inhibiting genes, Bcl-2 and Bcl- XL, were found in both Yinjin or Yinjinsaryongsangagambang-treated HepG2 cells while no changes was detected in expression levels of an apoptosis-promoting gene, Bax.

• 한국식품영양과학회지
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• 제45권1호
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• pp.137-142
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• 2016

본 연구에서는 국내산 천일염 두 종류와 정제염이 HepG2 인체 간암 세포에서의 세포 성장 억제 효과, apoptosis 관련 유전자 Bcl-2, Bax, p53 및 p21의 mRNA 발현과 단백질 발현을 확인하였다. 소금 시료들은 HepG2 인체 간암세포에 0.5% 및 1% 농도로 처리하였을 때 세포 성장을 억제시켰으며 T염전의 천일염(SS-T)과 Y염전의 천일염(SS-Y)은 정제염(PS)에 비해 암세포 성장을 유의적으로 억제하였다(P<0.05). 또한 HepG2 세포에 1% 농도로 소금 시료를 처리하였을 때 대조군에 비해 apoptosis를 억제하는 유전자들을 mRNA 및 단백질 수준에서 조절하여 Bcl-2는 낮게 나타났고, Bax, p53, p21은 높게 발현되었다. SS-T와 SS-Y는 PS에 비하여 Ca, Mg, S, K 함량이 많았으며, 천일염 중에서도 SS-T가 SS-Y보다 많이 함유되어 있었다. 전반적으로 무기질이 많이 함유된 천일염이 정제염보다 항암 효과가 높았으며 이는 소금의 Na 및 그 외 다른 무기질들이 항암 관련 유전자들을 조절한 것으로 보인다. 이상의 결과를 통해 국내산 천일염과 정제염은 HepG2 세포에서 apoptosis와 cell cycle 관련 유전자 조절을 통해 항암 효과를 나타냄을 확인하였으며 그중에서도 천일염의 효과가 더 높게 나타나 그 원인 물질 및 항암 기작에 대한 후속 연구가 필요하다.

• Tuberculosis and Respiratory Diseases
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• 제49권4호
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• pp.453-465
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• 2000

연구배경 : 폐암의 발생과정은 다양한 유전자 이상과 여러 가지경로 이상을 포함한 다단계 과정이다. 암유전자의 활성화나 종양억제유전자의 불활성화, 그리고 결과적인 유전적 불안정성의 증가는 폐암의 발암과정에서 일어나는 주요한 사건이며 임상적으로 폐암이 진단되기까지 10내지 20여 가지의 유전적 변화가 축적되는 것으로 알려져 있다. 본 연구에서 저자들은 비소세포폐암에서 종양억제유전자인 p53과 FHIT의 돌연변이, FHIT 유전자의 전사체 이상 여부를 확인하고 종양억제유전자부근에 위치하는 극소위성의 유전적 변화를 관찰하였다. 대상 및 방법 : 비소세포폐암으로 진단된 후 외과적 적출술을 시행받은 환자 29명의 생검조직과 그에 대응하는 동일인의 정상조직을 대상으로 하였다. p53과 FHIT의 돌연변이 여부는 PCR-SSCP, DNA 염기분석으로 확인하였고 D3S1285, D9S171, TP53에서 극소위성 불안정성과 이형접합성 상실은 PCR로 확인하였다. FHIT 유전자의 전사체 이상 여부 확인을 위해서는 RT-PCR을 사용하였다. 결과 : 1) p53 유전자의 2예에서 관찰되었고 모두 exon 5에서 1개의 염기가 치환되는 점돌연변이였다. 2) 극소위성 불안정성은 D3S1285와 D9S171에서 각각 2예, 1예, 이형접합성 상실은 D3S1285, D9S171, TP53에서 각각 3예, 4예, 7예가 관찰되었다. 3) FHIT 유전자의 변이는 11예에서 관찰되었으며 이중 6예는 exon 8의 codon 98에서 염기서열이 CAT가 CAC로 바뀌는 잠재적 치환이었다. 4) FHIT 유전자의 전사체 이상은 $\beta$-actin이 제대로 발현되는 15예중 4예에서 관찰되었으며 exon 6-9의 결실로 확인되었다. 결론 : 이상으로 비소세포폐암 발생에 p53, FHIT 유전자의 변이, 극소위성 불안정성과 이형접합성 상실 등 다양한 분자유전학적 기전이 복합적으로 작용할 것으로 생각되며 이번 연구에서 조사된 유전적 이상의 빈도는 앞서 발표된 서양의 연구결과와 대체적으로 일치한다. 특히 극소위성의 분석은 편평세포암에서 종양표지자로서의 역할이 기대된다. 이런 발암과정에 대한 이해는 예방, 진단 및 치료적 접근을 발전시키는데 도움을 줄 수 있을 것이고 향후 이들에 관한 가능적 연구들이 수행되어야 할 것이다.

• Asian Pacific Journal of Cancer Prevention
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• 제15권22호
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• pp.9791-9795
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• 2014

To study the gene expression change and possible signal pathway during androgen-dependent prostate cancer (ADPC) becoming androgen-independent prostate cancer (AIPC), an LNCaP cell model of AIPC was established using flutamide in combination with androgen-free environment inducement, and differential expression genes were screened by microarray. Then the biological process, molecular function and KEGG pathway of differential expression genes are analyzed by Molecule Annotation System (MAS). By comparison of 12,207 expression genes, 347 expression genes were acquired, of which 156 were up-ragulated and 191 down-regulated. After analyzing the biological process and molecule function of differential expression genes, these genes are found to play crucial roles in cell proliferation, differntiation, cell cycle control, protein metabolism and modification and other biological process, serve as signal molecules, enzymes, peptide hormones, cytokines, cytoskeletal proteins and adhesion molecules. The analysis of KEGG show that the relevant genes of AIPC transformation participate in glutathione metabolism, cell cycle, P53 signal pathway, cytochrome P450 metabolism, Hedgehog signal pathway, MAPK signal pathway, adipocytokines signal pathway, PPAR signal pathway, TGF-${\beta}$ signal pathway and JAK-STAT signal pathway. In conclusion, during the process of ADPC becoming AIPC, it is not only one specific gene or pathway, but multiple genes and pathways that change. The findings above lay the foundation for study of AIPC mechanism and development of AIPC targeting drugs.

• IMMUNE NETWORK
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• 제1권3호
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• pp.221-229
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• 2001

Background: Inactivation of tumor suppressor genes is believed to be important in the development of many human malignancies. Recently, several lines of evidence have indicated that the wild type p53 gene located at 17p13.3, may function as a tumor suppressor gene and that a mutant p53 gene could promote transformation by inactivating normal p53 function in a dominant negative fashion. These broad spectrum of p53 mutation in human cancers provide that mutant p53 and their protein may be potential targets of tumor diagnostic and therapeutic interventions. Method: Colony formation was performed to investigate growth suppressional ability. p53 expression pattern was examined by western blot and p53-mediated transactivation ability was assessed by CAT activity. SNU C2A cells were observed in apoptotic aspects induced by etoposide and $H_2O_2$ treatment, detecting sensitivity on agent, DNA fragmentation through agarose gel, chromatin condensation by fluorescence microscope, and cell cycle distribution by FACS. Result: 1) p53 mutant his179arg ($histidine{\rightarrow}arginine$) detected in SNU C2A cells lost transcriptional activity and growth suppression ability, showing dominant negative effect on its wild type p53. 2) Etoposide-treated SNU C2A cells induced apoptosis, exhibiting dramatic reduction of cell growth, DNA fragmentation, nuclear condensation formation of apoptotic body and increment of sub-G1 cell fraction. 3) Etoposide and $H_2O_2$-treated SNU C2A cells have no high increase of p53 expression and overexpressed p53 protein changed localization, from cytoplasm to nucleus. Also, p53-mediated transcriptional activity was increased by agents-treatment. Conclusion: SNU C2A cells coexpress wild-type and mutant p53 protein induced apoptosis in the condition on DNA damage, through localizational shift from cytoplasm to nucleus of p53 protein rather than the induction of p53 protein. SNU C2A cells derived mutant p53 his179arg abrogated both the growth supression ability and transactivational activity, showing inhibition effect on transcriptional activity of wild type p53, but did not repress the activity of wild type p53 in SNU C2A cells owing to dominant activity of wild type. These cell condition may provide new gene therapeutic implications leading effective antiproliferation of cell when mutant and wild-type p53 protein were co-expressed in cell.

• Asian Pacific Journal of Cancer Prevention
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• 제14권12호
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• pp.7651-7656
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• 2013

Background: Breast cancer is the most common malignancy among women in both developed and developing countries. The burden is increasing in low-income and middle-income countries (LMCs) and threatens the public health of such societies. Introduction of expensive monoclonal antibodies to cancer treatment regimens poses a real challenge in the health systems of LMCs. Despite controversy of cost-effectiveness of bevacizumab in breast cancer, some studies indicate gain of patients from this drug. The present study aimed to propose a priority setting model for administration of anti-angiogenic agents in breast cancer via assessment of tumor angiogenesis by the microvessel density (MVD) method and associations with clinicopathological characteristics (including simultaneous mutations of TP53 and HER-2 genes). Materials and Methods: Age, axillary lymph nodes status, tumor size, stage and grade, estrogen and progesterone receptors status, HER-2/neu status (by immunohistochemistry and FISH test), TP53 mutation, Ki-67 (for proliferation assay) and CD34 (for angiogenesis assay) were assessed in 111 breast cancer patients. The molecular subtype of each tumor was also determined and correlations of simultaneous mutations of HER-2 and p53 genes with angiogenesis and other clinicopathological characteristics were evaluated. Results: There were significant associations between simultaneous mutations of HER-2 and p53 genes and all other parameters except tumor size. The degree of angiogenesis in the ERBB2 subtype was greater than the others. Younger patients showed a higher angiogenesis rate rather those older than 50 years. Conclusions: Our results demonstrated that patients with simultaneous mutations of HER-2 and p53 genes, those with ERBB2 molecular subtype and also younger women (often triple negative) seem more eligible for obtaining anti-angiogenic agents. These results suggest a model for priority setting of patients with breast cancer for treatment with anti-angiogenic drugs in LMCs.

• 대한임상검사과학회지
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• 제45권1호
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• pp.32-36
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• 2013

The thyroid is the organ that has the greatest risk of malignant tumors among the endocrine tumors. The papillary carcinoma occupies 80% of the entire thyroid tumors. Immunohistochemical staining of galectin-3 has usually been used in differentiating papillary carcinoma and follicular carcinoma. The p53 gene of the cell cycle is a tumor suppressor gene acting in on the control points. The cyclin D1 genes in the cell cycle, involved in the implementation of G1 and S phase, plays an important role in the progression of thyroid tumors. This research compares and analyzes correlation between papillary carcinoma, follicular carcinoma, p53, cyclin D1 and galectin-3 gene expression patterns. In a total of 30 cases from papillary carcinoma, 21 cases from p53 (70%), 27 cases in galectin-3 (90%), and 26 cases in cyclin D1 (86.7%) showed positive rate. The galectin-3 staining investigated, showed a significant difference between a papillary carcinoma and a follicular carcinoma. Follicular carcinoma from 15 cases, p53 in 13 cases (86.7%), galectin-3 in 5 cases (33.3%) and cyclin D1 in 12 cases (80%) showed a positive rate. The cyclin D1 in follicular carcinoma and staining between the p53 that had correlation was also investigated. In this study, as the examples of the expression of the 27 cases of galectin-3 (90%) in papillary carcinoma and 5 cases in follicular carcinoma (33.3%) indicate, it was concluded that there is a difference in the expression on both carcinoma. In addition, cyclin D1 and p53 has a positive rate in follicular carcinoma, when cyclin D1 in 12 cases (80%), there was a significant correlation that was investigated. Distinguishing between papillary carcinoma and follicular carcinoma can be identified by the expression of galectin-3. It is considered to get results that are more accurate in follicular carcinoma diagnosis depending on whether the cyclin D1 and p53 is expressed or not.

• Biomolecules & Therapeutics
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• 제32권1호
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• pp.94-103
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• 2024

Non-alcoholic fatty liver disease (NAFLD) is characterized by excessive accumulation of fat in the liver, and there is a global increase in its incidence owing to changes in lifestyle and diet. Recent findings suggest that p53 is involved in the development of non-alcoholic fatty liver disease; however, the association between p53 expression and the disease remains unclear. Doxorubicin, an anticancer agent, increases the expression of p53. Therefore, this study aimed to investigate the role of doxorubicin-induced p53 upregulation in free fatty acid (FFA)-induced intracellular lipid accumulation. HepG2 cells were pretreated with 0.5 ㎍/mL of doxorubicin for 12 h, followed by treatment with FFA (0.5 mM) for 24 h to induce steatosis. Doxorubicin pretreatment upregulated p53 expression and downregulated the expression of endoplasmic reticulum stress- and lipid synthesis-associated genes in the FFA -treated HepG2 cells. Additionally, doxorubicin treatment upregulated the expression of AMP-activated protein kinase, a key modulator of lipid metabolism. Notably, siRNA-targeted p53 knockdown reversed the effects of doxorubicin in HepG2 cells. Moreover, doxorubicin treatment suppressed FFA -induced lipid accumulation in HepG2 spheroids. Conclusively, these results suggest that doxorubicin possesses potential application for the regulation of lipid metabolism by enhance the expression of p53 an in vitro NAFLD model.

• Journal of Animal Science and Technology
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• 제53권3호
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• pp.185-193
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• 2011

To investigate the influences of candidate genes on the birth weight and the early stages of life, genotyping of the prolactin receptor 3 (PRLR3) and retinol-binding protein 4 (RBP4) genes was performed in 156 and 141 Berkshire pigs, respectively. The frequency of both PRLR3 alleles A and a was 0.50. The frequencies of the RBP4 alleles B and b were 0.42 and 0.58, respectively. Neither locus was in Hardy-Weinberg equilibrium. No significant associations of the PRLR3 alleles with birth or weaning weights and of the RBP4 alleles with birth weight were observed. The proportions of the phenotype variances due to the genotypes of PRLR3 in the feeder weights was 4.0% and those of RBP4 in the weaning and feeder weights were 11.9 and 3.3%, respectively (P < 0.05). The dominance effect of PRLR3 and RBP4 on feeder weights was 2.40 and -1.86 kg, respectively (P < 0.01). The additive and dominance effects of RBP4 on weaning weights were 0.332 and -0.682 kg, respectively (P < 0.01). Even if no significant epistasis of PRLR3 and RBP4 was detected, a considerable trend of consistent positive epistasis estimates of AA/BB and Aa/Bb was observed for all traits. The results of this study may have a considerable impact on early-stage growth by both loci, and a selection strategy should be designed separately for each marker in Berkshire pigs.

• 대한본초학회지
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• 제28권6호
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• pp.53-58
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• 2013

Objectives : Pinelliae Tuber has been used as a typical unauthentic herbal medicines. Due to the morphological similarity between Pinelliae Tuber and adulterants, the correct authentication is very difficult. Therefore, we introduced DNA barcode to establish a powerful tool for the authentication of Pinelliae Tuner from adulterants. Methods : To obtain DNA barcode regions, genomic DNA was extracted from nineteen specimens of Pinellia ternata, Pinellia pedatisecta, Pinellia tripartita, and Typhonium flagelliforme, and matK and rbcL genes were amplified. For identification of species specific sequences and analysis phylogenetic relationship, a comparative analysis were performed by the ClastalW and UPGMA based on entire sequences of matK and rbcL genes, respectively. Results : In comparison of two DNA barcode sequences, we elucidated the phylogenetic relationship showing distinct four groups depending on species and identified 40 and 20 species specific nucleotides enough to distinguish each species from matK and rbcL gene, respectively. The sequence differences at the corresponding positions were avaliable genetic marker nulceotides to discriminate the correct species among analyzed four species. These results indicated that phylogentic and comparative analysis of matK and rbcL genes are useful genetic markers to authenticate Pinelliae Tubers. Conclusions : The marker nucleotides enough to distinguish P. ternata, P. tripatrita, P. peditisecta, and T. flagelliform, were observed at 40 positions in matK gene and 20 positions in rbcL gene sequence, respectively. These differences can be used to authenticate Pinelliae Tuber from adulterants as well as discriminate each four species.