Limits...
Quantitative proteomic analysis of mitochondria from human ovarian cancer cells and their paclitaxel-resistant sublines.

Chen M, Huang H, He H, Ying W, Liu X, Dai Z, Yin J, Mao N, Qian X, Pan L - Cancer Sci. (2015)

Bottom Line: Identification of the relevant proteins in mitochondria will help in clarifying the possible mechanisms and in selecting effective chemotherapy for patients with paclitaxel resistance.Among them, mimitin and 14-3-3 ζ/δ were selected for further research.Multivariate analyses demonstrated that lower expression levels of the mimitin and 14-3-3 ζ/δ proteins were positively associated with shorter progression-free survival (PFS) and overall survival (OS) in patients with primary ovarian cancer (mimitin: PFS: P = 0.041, OS: P = 0.003; 14-3-3 ζ/δ: PFS: P = 0.031, OS: P = 0.011).

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

Show MeSH

Related in: MedlinePlus

(a) Electron microscopy image of mitochondria morphology (10 000×). The purity of the mitochondria was confirmed using electron microscopy, including intactness and contamination. (b) Western blot analyses of total cell proteins (Cell) and mitochondrial preparations isolated from paclitaxel-sensitive SKOV3 cells (Smt), paclitaxel-resistant SKOV3 cells (TSmt), paclitaxel-sensitive A2780 cells (Amt), and paclitaxel-resistant A2780 cells (TAmt) for COX4 (mitochondrial marker), lamin-B (nuclear marker), flotillin-1 (cell membrane marker) and β-actin (cytoskeletal protein). There was little contamination in the mitochondria-enriched fractions.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4556398&req=5

fig02: (a) Electron microscopy image of mitochondria morphology (10 000×). The purity of the mitochondria was confirmed using electron microscopy, including intactness and contamination. (b) Western blot analyses of total cell proteins (Cell) and mitochondrial preparations isolated from paclitaxel-sensitive SKOV3 cells (Smt), paclitaxel-resistant SKOV3 cells (TSmt), paclitaxel-sensitive A2780 cells (Amt), and paclitaxel-resistant A2780 cells (TAmt) for COX4 (mitochondrial marker), lamin-B (nuclear marker), flotillin-1 (cell membrane marker) and β-actin (cytoskeletal protein). There was little contamination in the mitochondria-enriched fractions.

Mentions: The purity of the isolated mitochondria was determined using electron microscopy. The examination of multiple electron microscopic fields revealed that the preparations contained few, if any, nuclear fragments or other cytosolic organelles (Fig.2a). In addition, we used western blotting to validate the purity of the isolated mitochondria. The total cell protein extraction was used as a control. Western blot results showed that mitochondrial proteins were greatly enriched in the purified samples. A mitochondrial marker COX4 was determined in total cell proteins and mitochondrial preparations were isolated from SKOV3, A2780, SKOV3-TR and A2780-TR cell lines. In contrast, lamin-B, flotillin-1 and β-actin were only detected in the cellular proteins and were absent in the mitochondrial preparations (Fig.2b).


Quantitative proteomic analysis of mitochondria from human ovarian cancer cells and their paclitaxel-resistant sublines.

Chen M, Huang H, He H, Ying W, Liu X, Dai Z, Yin J, Mao N, Qian X, Pan L - Cancer Sci. (2015)

(a) Electron microscopy image of mitochondria morphology (10 000×). The purity of the mitochondria was confirmed using electron microscopy, including intactness and contamination. (b) Western blot analyses of total cell proteins (Cell) and mitochondrial preparations isolated from paclitaxel-sensitive SKOV3 cells (Smt), paclitaxel-resistant SKOV3 cells (TSmt), paclitaxel-sensitive A2780 cells (Amt), and paclitaxel-resistant A2780 cells (TAmt) for COX4 (mitochondrial marker), lamin-B (nuclear marker), flotillin-1 (cell membrane marker) and β-actin (cytoskeletal protein). There was little contamination in the mitochondria-enriched fractions.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556398&req=5

fig02: (a) Electron microscopy image of mitochondria morphology (10 000×). The purity of the mitochondria was confirmed using electron microscopy, including intactness and contamination. (b) Western blot analyses of total cell proteins (Cell) and mitochondrial preparations isolated from paclitaxel-sensitive SKOV3 cells (Smt), paclitaxel-resistant SKOV3 cells (TSmt), paclitaxel-sensitive A2780 cells (Amt), and paclitaxel-resistant A2780 cells (TAmt) for COX4 (mitochondrial marker), lamin-B (nuclear marker), flotillin-1 (cell membrane marker) and β-actin (cytoskeletal protein). There was little contamination in the mitochondria-enriched fractions.
Mentions: The purity of the isolated mitochondria was determined using electron microscopy. The examination of multiple electron microscopic fields revealed that the preparations contained few, if any, nuclear fragments or other cytosolic organelles (Fig.2a). In addition, we used western blotting to validate the purity of the isolated mitochondria. The total cell protein extraction was used as a control. Western blot results showed that mitochondrial proteins were greatly enriched in the purified samples. A mitochondrial marker COX4 was determined in total cell proteins and mitochondrial preparations were isolated from SKOV3, A2780, SKOV3-TR and A2780-TR cell lines. In contrast, lamin-B, flotillin-1 and β-actin were only detected in the cellular proteins and were absent in the mitochondrial preparations (Fig.2b).

Bottom Line: Identification of the relevant proteins in mitochondria will help in clarifying the possible mechanisms and in selecting effective chemotherapy for patients with paclitaxel resistance.Among them, mimitin and 14-3-3 ζ/δ were selected for further research.Multivariate analyses demonstrated that lower expression levels of the mimitin and 14-3-3 ζ/δ proteins were positively associated with shorter progression-free survival (PFS) and overall survival (OS) in patients with primary ovarian cancer (mimitin: PFS: P = 0.041, OS: P = 0.003; 14-3-3 ζ/δ: PFS: P = 0.031, OS: P = 0.011).

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

Show MeSH
Related in: MedlinePlus