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Mitochondrial reactive oxygen species regulate adipocyte differentiation of mesenchymal stem cells in hematopoietic stress induced by arabinosylcytosine.

Wang W, Zhang Y, Lu W, Liu K - PLoS ONE (2015)

Bottom Line: ROS levels were detected using the CM-H2DCFDA probe and Mito-SOX dye.The elevated ROS levels induced by Ara-C were caused by both over-generation of mitochondrial ROS and reduction of antioxidant enzymes (Cu/Zn Superoxide dismutase and catalase).Our findings suggest that a mitochondrial-targeted antioxidant could diminish adipocyte differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Peking University People's Hospital, Beijing, China; Institute of Hematology, Peking University, Beijing, China.

ABSTRACT

Objective: The increase in adipocytes induced by chemotherapeutic drugs may play a negative role in hematopoietic recovery. However, the mechanism underlying adipocyte differentiation of mesenchymal stem cells (MSCs) in hematopoietic stress is still unknown. Hence, the involvement of reactive oxygen species (ROS) in adipocyte differentiation under hematopoietic stress was investigated in vitro and in vivo.

Methods: The roles of cellular ROS in adipogenesis were investigated in vivo through an adipocyte hyperplasia marrow model under hematopoietic stress induced by arabinosylcytosine (Ara-C) and in vitro via adipocyte differentiation of human MSCs. ROS levels were detected using the CM-H2DCFDA probe and Mito-SOX dye. Adipogenesis was evaluated by histopathology and oil red O staining, whereas detection of mRNA levels of antioxidant enzymes and adipogenesis markers was performed using quantitative real-time polymerase chain reaction analysis.

Results: ROS were found to play an important role in regulating adipocyte differentiation of MSCs by activating peroxisome proliferator-activated receptor gamma (PPARγ,) while the antioxidant N-acetyl-L-cysteine acts through ROS to inhibit adipocyte differentiation. The elevated ROS levels induced by Ara-C were caused by both over-generation of mitochondrial ROS and reduction of antioxidant enzymes (Cu/Zn Superoxide dismutase and catalase). Our findings suggest that a mitochondrial-targeted antioxidant could diminish adipocyte differentiation.

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Related in: MedlinePlus

Mito-Tempo diminished adipocyte differentiation.(A) ROS levels in human MSCs on Day 0 and Day 2 of differentiation. Cells were treated with 500 nM of TPP or Mito-Tempo for 4 h prior to measurement on Day 2. (B) Mito-Tempo diminished lipid accumulation. Human MSCs were treated with 500 nM of Mito-Tempo starting on Day 2 of differentiation. Cells were fixed and stained with oil red O on Day 14. The oil red-O was extracted with isopropanol and absorbance was measured at 518 nm. (C, D) Gene expression of PPARγ and adiponectin was decreased in the presence of Mito-Tempo on Day 7 and Day 14 of differentiation.*P < 0.05.
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pone.0120629.g004: Mito-Tempo diminished adipocyte differentiation.(A) ROS levels in human MSCs on Day 0 and Day 2 of differentiation. Cells were treated with 500 nM of TPP or Mito-Tempo for 4 h prior to measurement on Day 2. (B) Mito-Tempo diminished lipid accumulation. Human MSCs were treated with 500 nM of Mito-Tempo starting on Day 2 of differentiation. Cells were fixed and stained with oil red O on Day 14. The oil red-O was extracted with isopropanol and absorbance was measured at 518 nm. (C, D) Gene expression of PPARγ and adiponectin was decreased in the presence of Mito-Tempo on Day 7 and Day 14 of differentiation.*P < 0.05.

Mentions: To further specify the role of ROS produced by mitochondria in adipocyte differentiation, the effect of Mito-Tempo on adipocyte differentiation of MSCs was investigated in vitro. The Mito-Tempo antioxidant is a combination of piperidine nitroxide and the lipophilic cation triphenylphosphonium (TPP), giving Mito-Tempo the ability to pass through lipid bilayers with ease and accumulate several hundred-fold inside the mitochondria. Human MSCs were treated with control TPP and Mito-Tempo starting on Day 2 of differentiation. As shown in Fig. 4A, the increase in ROS in the early stage of differentiation was attenuated in the presence of Mito-Tempo. Furthermore, Mito-Tempo significantly reduced lipid accumulation (Fig. 4B) as well as the expression of PPARγ and adiponectin (Fig. 4C-D). These results demonstrate that mitochondrial ROS are required for adipocyte differentiation of MSCs.


Mitochondrial reactive oxygen species regulate adipocyte differentiation of mesenchymal stem cells in hematopoietic stress induced by arabinosylcytosine.

Wang W, Zhang Y, Lu W, Liu K - PLoS ONE (2015)

Mito-Tempo diminished adipocyte differentiation.(A) ROS levels in human MSCs on Day 0 and Day 2 of differentiation. Cells were treated with 500 nM of TPP or Mito-Tempo for 4 h prior to measurement on Day 2. (B) Mito-Tempo diminished lipid accumulation. Human MSCs were treated with 500 nM of Mito-Tempo starting on Day 2 of differentiation. Cells were fixed and stained with oil red O on Day 14. The oil red-O was extracted with isopropanol and absorbance was measured at 518 nm. (C, D) Gene expression of PPARγ and adiponectin was decreased in the presence of Mito-Tempo on Day 7 and Day 14 of differentiation.*P < 0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4359087&req=5

pone.0120629.g004: Mito-Tempo diminished adipocyte differentiation.(A) ROS levels in human MSCs on Day 0 and Day 2 of differentiation. Cells were treated with 500 nM of TPP or Mito-Tempo for 4 h prior to measurement on Day 2. (B) Mito-Tempo diminished lipid accumulation. Human MSCs were treated with 500 nM of Mito-Tempo starting on Day 2 of differentiation. Cells were fixed and stained with oil red O on Day 14. The oil red-O was extracted with isopropanol and absorbance was measured at 518 nm. (C, D) Gene expression of PPARγ and adiponectin was decreased in the presence of Mito-Tempo on Day 7 and Day 14 of differentiation.*P < 0.05.
Mentions: To further specify the role of ROS produced by mitochondria in adipocyte differentiation, the effect of Mito-Tempo on adipocyte differentiation of MSCs was investigated in vitro. The Mito-Tempo antioxidant is a combination of piperidine nitroxide and the lipophilic cation triphenylphosphonium (TPP), giving Mito-Tempo the ability to pass through lipid bilayers with ease and accumulate several hundred-fold inside the mitochondria. Human MSCs were treated with control TPP and Mito-Tempo starting on Day 2 of differentiation. As shown in Fig. 4A, the increase in ROS in the early stage of differentiation was attenuated in the presence of Mito-Tempo. Furthermore, Mito-Tempo significantly reduced lipid accumulation (Fig. 4B) as well as the expression of PPARγ and adiponectin (Fig. 4C-D). These results demonstrate that mitochondrial ROS are required for adipocyte differentiation of MSCs.

Bottom Line: ROS levels were detected using the CM-H2DCFDA probe and Mito-SOX dye.The elevated ROS levels induced by Ara-C were caused by both over-generation of mitochondrial ROS and reduction of antioxidant enzymes (Cu/Zn Superoxide dismutase and catalase).Our findings suggest that a mitochondrial-targeted antioxidant could diminish adipocyte differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Peking University People's Hospital, Beijing, China; Institute of Hematology, Peking University, Beijing, China.

ABSTRACT

Objective: The increase in adipocytes induced by chemotherapeutic drugs may play a negative role in hematopoietic recovery. However, the mechanism underlying adipocyte differentiation of mesenchymal stem cells (MSCs) in hematopoietic stress is still unknown. Hence, the involvement of reactive oxygen species (ROS) in adipocyte differentiation under hematopoietic stress was investigated in vitro and in vivo.

Methods: The roles of cellular ROS in adipogenesis were investigated in vivo through an adipocyte hyperplasia marrow model under hematopoietic stress induced by arabinosylcytosine (Ara-C) and in vitro via adipocyte differentiation of human MSCs. ROS levels were detected using the CM-H2DCFDA probe and Mito-SOX dye. Adipogenesis was evaluated by histopathology and oil red O staining, whereas detection of mRNA levels of antioxidant enzymes and adipogenesis markers was performed using quantitative real-time polymerase chain reaction analysis.

Results: ROS were found to play an important role in regulating adipocyte differentiation of MSCs by activating peroxisome proliferator-activated receptor gamma (PPARγ,) while the antioxidant N-acetyl-L-cysteine acts through ROS to inhibit adipocyte differentiation. The elevated ROS levels induced by Ara-C were caused by both over-generation of mitochondrial ROS and reduction of antioxidant enzymes (Cu/Zn Superoxide dismutase and catalase). Our findings suggest that a mitochondrial-targeted antioxidant could diminish adipocyte differentiation.

Show MeSH
Related in: MedlinePlus