[논문]위암 조직과 세포주에서 mDNA와 OXPHOS 단백질 분석

아드리안 시레가; 하영술; 문동규; 우동균

doi:10.5352/jls.2021.31.12.1057

[국내논문] 위암 조직과 세포주에서 mDNA와 OXPHOS 단백질 분석
Alterations in Mitochondrial DNA Copy Numbers and Mitochondrial Oxidative Phosphorylation (OXPHOS) Protein Levels in Gastric Cancer Tissues and Cell Lines 원문보기

생명과학회지 = Journal of life science, v.31 no.12, 2021년, pp.1057 - 1065

아드리안 시레가 (경상국립대학교 약학과) , 하영술 (경상국립학교병원 의생명연구원) , 문동규 (경상국립대학교병원 정형외과) , 우동균 (경상국립대학교 약학과)

초록
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위암 환자에서 미토콘드리아 DNA (mtDNA)의 양적 변화가 보고 되고 있으며 이러한 변화가 위암의 발암이나 진행에 관여되는 것으로 추정되고 있다. 그리나 위암에서 미토콘드리아 단백질이나 mtDNA에 의해 암호화된 산화적 인산화(OXPHOS) 단백질의 양적 변화에 관한 연구는 아직까지 미비한 실정이다. 본 연구에서는 위암환자 조직 및 세포주를 이용하여 mtDNA 양 그리고 미토콘드리아 단백질 및 OXPHOS 단백질의 양을 분석하였다. 또한, mtDNA 양적 변화와 위암 환자의 임상병리학적 특징을 연관 분석하였다. MtDNA 양을 분석하기 위하여 qPCR 기법을 그리고 단백질 분석에는 Western blot 기법을 각각 활용하였다. 총 27개의 위암 환자 샘플에서 약 80%에 해당하는 22개의 환자 위암조직에서 정상조직에 비해 mtDNA 양이 감소하였으며, 나머지 환자에서는 mtDNA 양이 증가하였다. 이러한 mtDNA 양이 감소한 위암 조직 샘플에서는 미토콘드리아 단백질 및 OXPHOS 단백질의 양도 같이 감소하였다. 한편, 본 연구에 사용된 총 5개의 위암 세포주 모두에서 mtDNA 양이 감소하였다 그러나 위암 세포주에서는 mtDNA 양적 감소와 미토콘드리아 단백질 및 OXPHOS 단백질의 양적 감소가 항상 일치하지는 않았다. 이러한 연구결과는 위암 조직 및 세포주에서 mtDNA 양의 감소가 흔하며 이는 mtDNA 양적 변화가 위암의 생성에 관여함을 제시한다.

Abstract ▼ AI-Helper

Alterations in mitochondrial DNA (mtDNA) copy numbers have been reported in patients with stomach cancer and suggested to play a role in gastric carcinogenesis or gastric cancer progression. However, changes in the levels of mitochondrial proteins or mtDNA-encoded oxidative phosphorylation (OXPHOS) proteins in gastric cancer remain unclear. In this study, we investigated mtDNA contents, mitochondrial protein levels, and mtDNA-encoded OXPHOS protein levels in gastric cancer tissues and cell lines. We correlated mtDNA copy numbers with clinicopathologic features of the gastric cancer samples used in this study and used quantitative PCR to analyze the mtDNA copy numbers of the gastric cancer tissues and cell lines. Western blot analysis was used for assessing the amounts of mitochondrial proteins and mtDNA-encoded OXPHOS proteins. Among the 27 gastric cancer samples, 22 showed a reduction in mtDNA copy numbers. The mtDNA content was increased in the other five samples relative to that in normal matched gastric tissues. Mitochondrial protein and OXPHOS protein levels were reduced in some gastric cancer tissues. However, mitochondrial protein and OXPHOS protein levels in gastric cancer cell lines were not always in line with their mtDNA contents. The mtDNA copy numbers were reduced in five gastric cancer cell lines tested in this study. In summary, this study reports a common reduction in mtDNA contents in gastric carcinoma tissues and cell lines, pointing to the possible involvement of mtDNA content alterations in tumorigenesis of the stomach.

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표/그림 (4)

표 Table 1. Correlation between mtDNA copy numbers and clinicopathologic features in 27 gastric cancer patients
그림 Fig. 1. mtDNA copy numbers in gastric cancer tissues by quantitative PCR. (A) Alterations of mtDNA copy numbers in 27 gastric cancer tissues relative to their matching normal tissues. (B) The overall mtDNA copy number ratio of 27 gastric cancer tissues to their normal tissues. *indicates p<0.05, **indicates p<0.01, and ***indicates p<0.001.
그림 Fig. 2. VDAC and mtDNA-encoded OXPHOS protein levels in gastric cancer tissues by Western blot analysis. (A) VDAC protein levels in six gastric cancer tissues relative to their matching normal tissues. (B) mtDNA-encoded OXPHOS protein (UQCRC2, COX2, and ATPB) levels in three gastric cancer tissues relative to their matching normal tissues. N indicates normal tissue and T indicates gastric cancer tissue. Numbers under the blot images indicate quantitated amounts of corresponding target protein relative to Actin.
그림 Fig. 3. mtDNA copy numbers, VDAC protein levels, and mtDNA-encoded OXPHOS protein levels in five gastric cancer cell lines (SNU1, SNU484, SNU638, SNU719 and MKN28). (A) mtDNA copy numbers in gastric cancer cell lines were assessed by qPCR. (B) VDAC protein levels in gastric cancer cell lines were assessed by Western blot analysis. (C) mtDNA-encoded OXPHOS protein (UQCRC2, COX2, and ATPB) levels in gastric cancer cell lines were assessed by Western blot analysis. HEK293T cell line was used as a control. Numbers under the blot images indicate quantitated amounts of corresponding target protein relative to Actin. ***indicates p<0.001.

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The Student’s two tailed t-test was used for the determination of statistical relevance between groups

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