Repair effects of rat adipose-derived stem cells on DNA damage induced by ultraviolet in chondrocytes_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

ZHANG Jinli ¹ , LIU Zhihe ¹ , TANG Wenbin ² , XIONG Xifeng ¹ , ZHANG Zhi ² , CAO Wenjuan ¹ ,  LI Xiaojian ²

1. Guangzhou Institute of Traumatic Surgery, Guangzhou Red Cross Hospital, Medical College of Jinan University, Guangzhou Guangdong, 510220, P.R.China;
2. Department of Burn and Plastic Surgery, Guangzhou Red Cross Hospital, Medical College of Jinan University, Guangzhou Guangdong, 510220, P.R.China;

Corresponding author：

LI Xiaojian, Email: lixj64@163.com

Keywords：

Adipose-derived stem cells; chondrocyte; radiation; repair; rat

DOI：

10.7507/1002-1892.201610106

Video：

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Objective To explore the DNA repair effect of rat adipose-derived stem cells (ADSCs) on chond-rocytes exposed to ultraviolet (UV) radiation. Methods ADSCs were isolated and cultured from the inguinal adipose tissue of Sprague Dawley rat by digestion with collagenase type I. ADSCs cell phenotype was assayed with flow cytometry. Multiple differentiation capability of ADSCs at passage 3 was identified with osteogenic and adipogenic induction. The chondrocytes were obtained from rat articular cartilage by digestion with collagenase type II and were identified with toluidine blue staining. The chondrocytes at passage 3 were irradiated with 40 J/m² UV and cultured with normal medium (irradiated group), and medium containing the ADSCs supernatant (ADSCs supernatant group) or ADSCs was used for co-culture (ADSCs group) for 24 hours; no irradiation chondrocytes served as control group. The cell proliferation was estimated by MTS method. The expression of phosphorylated histone family 2A variant (γH2AX) was detected by immunofluorescence and Western blot. Results ADSCs presented CD29(+), CD44(+), CD106(-), and CD34(-); and results of the alizarin red staining and oil red O staining were positive after osteogenic and adipogenic induction. Cell proliferation assay demonstrated the absorbance (A) values were 2.20±0.10 (control group), 1.34±0.04 (irradiated group), and 1.57±0.06 (ADSCs supernatant group), showing significant difference between groups (P<0.05). Immunofluorescence and Western blot showed that the γH2AX protein expression was significantly increased in irradiated group, ADSCs supernatant group, and ADSCs group when compared with control group (P<0.05), and the expression was significantly decreased in ADSCs supernatant group and ADSCs group when compared with irradiated group (P<0.05), but no significant difference was found between ADSCs supernatant group and ADSCs group (P>0.05). Conclusion ADSCs can increase the cell proliferation and down-regulate the γH2AX protein expression of irradiated cells, indicating ADSCs contribute to the repair of irradiated chondrocyte.

Citation： ZHANG Jinli, LIU Zhihe, TANG Wenbin, XIONG Xifeng, ZHANG Zhi, CAO Wenjuan, LI Xiaojian. Repair effects of rat adipose-derived stem cells on DNA damage induced by ultraviolet in chondrocytes. Chinese Journal of Reparative and Reconstructive Surgery, 2017, 31(5): 600-606. doi: 10.7507/1002-1892.201610106 Copy

1.	Nie C, Yang D, Xu J, et al. Locallty administered adipos-derived stem cells accelerate wound healing through differentiation and vasculogenesis. Cell Transplant, 2011, 20(2): 205-216.
2.	Zhou P. DNA damage, signaling and repair: protecting genomic integrity and reducing the risk of human disease. Chinese Sci Bull, 2011, 56(30): 3119-3121.
3.	Fenech M, Kirsch-Volders M, Natarajan AT, et al. Molecular mechanisms of micronucleus, nucleoplasmic bridge and nuclear bud formation in mammalianand human cells. Mutagenesis, 2011, 26(1): 125-132.
4.	王宗烨, 徐维邦, 宋淑军. 电离辐射对骨与软骨的损伤. 中华放射医学与防护杂志, 1996, 16(3): 208-210.
5.	Kraus A, Woon C, Raghavan S, et al. Co-culture of human adipose-derived stem cells with tenocytes increases proliferation and induces differentiation into a tenogenic lineage. Plast Reconstr Surg, 2013, 132(5): 754e-766e.
6.	Granéli C, Thorfve A, Rustschi U, et al. Novel markers of osteogenic and adipogenic differentiation of human bone marrow stromal cells identified using aquantitative proteomics approach. Stem Cell Res, 2014, 12(1): 153-165.
7.	Coleman CM, Vaughan EE, Browe DC, et al. Growth differentiation factor-5 enhancesin vitro mesenchymal stromal cell chondrogenesis and hypertrophy. Stem Cells Dev, 2013, 22(13): 1968-1976.
8.	Sándor GK, Tuovinen VJ, Wolff J, et al. Adipose stem cell tissue-engineered construct used to treat large anterior mandibular defect: a case report and review of the clinical application of good manufacturing practice-level adipose stem cells for bone regeneration. J Oral Maxillofac Surg, 2013, 71(5): 938-950.
9.	Prins HJ, Braat AK, Gawlitt AD, et al.In vitro induction of alkaline phosphatase levels predictsin vivo bone forming capacity of human bone marrow stromal cells. Stem Cells Res, 2013, 12(2): 428-440.
10.	Simões IN, Boura JS, dos Santos F, et al. Human mesenchymal stem cells from the umbilical cord matrix: succsessful isolation andex vivo expansion using serum-/xeno-free culture media. Biotechnol J, 2013, 8(4): 448-458.
11.	Fumimoto Y, Matsuyama A, Komoda H, et al. Creation of a rich subcutaneous vascular network withimplanted adipose tissue-derived stromal cells and adipose tissue enhances subcutaneous grafting of islets indiabetic mice. Tissue Eng Part C Methods, 2009, 15(3): 437-444.
12.	Matsumoto D, Sato K, Gonda K, et al. Cell-assisted lipotransfer: supportive use of human adipose-derived cells for soft tissue augmentation with lipoinjection. Tissue Eng, 2006, 12(12): 3375-3382.
13.	Yang JA, Chung HM, Won CH, et al. Potential application of adipose-derived stem cells and their secretory factors to skin: discussion fromboth clinical and industrial viewpoints. Expert Opin Biol Ther, 2010, 10(4): 495-503.
14.	Cai L, Johnstone BH, Cook TG, et al. Suppression of hepatocyte growth factor production impairs the ability ofadipose-derived stem cells to promote ischemic tissue revascularization. Stem Cells, 2007, 25(12): 3234-3243.
15.	Rehman J, Traktuev D, Li J, et al. Secretion of angiogenic and antiapoptotic factors by human adipose stromal cells. Circulation, 2004, 109(10): 1292-1298.
16.	Zuk PA. The adipose-derived stem cell: looking back and looking ahead. Mol Biol Cell, 2010, 21(11): 1783-1787.
17.	Lee EY, Xia Y, Kim WS, et al. Hypoxia-enhanced wound-healing function of adipose-derived stem cells: increase in stem cell proliferation and up-regulation of VEGF and bFGF. Wound Repair Regen, 2009, 17(4): 540-547.
18.	Fraser JK, Wulur I, Alfonso Z, et al. Fat tissue: an underappreciated source of stem cells forbiotechnology. Trends Biotechnol, 2006, 24(4): 150-154.
19.	Tu WZ, Li B, Huang B, et al. γH2AX foci formation in the absence of DNA damage: mitotic H2AX phosphorylation is mediated by the DNA-PKcs/CHK2 pathway. FEBS Lett, 2013, 587(21): 3437-3443.
20.	Benson LJ, Gu Y, Yakovleva T, et al. Modifications of H3 and H4 during chromatin replication, nucleosome assembly, and histone exchange. J Biol Chem, 2006, 281(14): 9287-9296.
21.	Gavrilov B, Vezhenkova I, Firsanov D, et al. Slow elimination of phosphorylated histone gamma-H2AX from DNA of terminally diferentiated mouse heart cellsin situ. Biochem Biophys ResCommun, 2006, 347(4): 1048-1052.

1. Nie C, Yang D, Xu J, et al. Locallty administered adipos-derived stem cells accelerate wound healing through differentiation and vasculogenesis. Cell Transplant, 2011, 20(2): 205-216.
2. Zhou P. DNA damage, signaling and repair: protecting genomic integrity and reducing the risk of human disease. Chinese Sci Bull, 2011, 56(30): 3119-3121.
3. Fenech M, Kirsch-Volders M, Natarajan AT, et al. Molecular mechanisms of micronucleus, nucleoplasmic bridge and nuclear bud formation in mammalianand human cells. Mutagenesis, 2011, 26(1): 125-132.
4. 王宗烨, 徐维邦, 宋淑军. 电离辐射对骨与软骨的损伤. 中华放射医学与防护杂志, 1996, 16(3): 208-210.
5. Kraus A, Woon C, Raghavan S, et al. Co-culture of human adipose-derived stem cells with tenocytes increases proliferation and induces differentiation into a tenogenic lineage. Plast Reconstr Surg, 2013, 132(5): 754e-766e.
6. Granéli C, Thorfve A, Rustschi U, et al. Novel markers of osteogenic and adipogenic differentiation of human bone marrow stromal cells identified using aquantitative proteomics approach. Stem Cell Res, 2014, 12(1): 153-165.
7. Coleman CM, Vaughan EE, Browe DC, et al. Growth differentiation factor-5 enhancesin vitro mesenchymal stromal cell chondrogenesis and hypertrophy. Stem Cells Dev, 2013, 22(13): 1968-1976.
8. Sándor GK, Tuovinen VJ, Wolff J, et al. Adipose stem cell tissue-engineered construct used to treat large anterior mandibular defect: a case report and review of the clinical application of good manufacturing practice-level adipose stem cells for bone regeneration. J Oral Maxillofac Surg, 2013, 71(5): 938-950.
9. Prins HJ, Braat AK, Gawlitt AD, et al.In vitro induction of alkaline phosphatase levels predictsin vivo bone forming capacity of human bone marrow stromal cells. Stem Cells Res, 2013, 12(2): 428-440.
10. Simões IN, Boura JS, dos Santos F, et al. Human mesenchymal stem cells from the umbilical cord matrix: succsessful isolation andex vivo expansion using serum-/xeno-free culture media. Biotechnol J, 2013, 8(4): 448-458.
11. Fumimoto Y, Matsuyama A, Komoda H, et al. Creation of a rich subcutaneous vascular network withimplanted adipose tissue-derived stromal cells and adipose tissue enhances subcutaneous grafting of islets indiabetic mice. Tissue Eng Part C Methods, 2009, 15(3): 437-444.
12. Matsumoto D, Sato K, Gonda K, et al. Cell-assisted lipotransfer: supportive use of human adipose-derived cells for soft tissue augmentation with lipoinjection. Tissue Eng, 2006, 12(12): 3375-3382.
13. Yang JA, Chung HM, Won CH, et al. Potential application of adipose-derived stem cells and their secretory factors to skin: discussion fromboth clinical and industrial viewpoints. Expert Opin Biol Ther, 2010, 10(4): 495-503.
14. Cai L, Johnstone BH, Cook TG, et al. Suppression of hepatocyte growth factor production impairs the ability ofadipose-derived stem cells to promote ischemic tissue revascularization. Stem Cells, 2007, 25(12): 3234-3243.
15. Rehman J, Traktuev D, Li J, et al. Secretion of angiogenic and antiapoptotic factors by human adipose stromal cells. Circulation, 2004, 109(10): 1292-1298.
16. Zuk PA. The adipose-derived stem cell: looking back and looking ahead. Mol Biol Cell, 2010, 21(11): 1783-1787.
17. Lee EY, Xia Y, Kim WS, et al. Hypoxia-enhanced wound-healing function of adipose-derived stem cells: increase in stem cell proliferation and up-regulation of VEGF and bFGF. Wound Repair Regen, 2009, 17(4): 540-547.
18. Fraser JK, Wulur I, Alfonso Z, et al. Fat tissue: an underappreciated source of stem cells forbiotechnology. Trends Biotechnol, 2006, 24(4): 150-154.
19. Tu WZ, Li B, Huang B, et al. γH2AX foci formation in the absence of DNA damage: mitotic H2AX phosphorylation is mediated by the DNA-PKcs/CHK2 pathway. FEBS Lett, 2013, 587(21): 3437-3443.
20. Benson LJ, Gu Y, Yakovleva T, et al. Modifications of H3 and H4 during chromatin replication, nucleosome assembly, and histone exchange. J Biol Chem, 2006, 281(14): 9287-9296.
21. Gavrilov B, Vezhenkova I, Firsanov D, et al. Slow elimination of phosphorylated histone gamma-H2AX from DNA of terminally diferentiated mouse heart cellsin situ. Biochem Biophys ResCommun, 2006, 347(4): 1048-1052.

Chinese Journal of Reparative and Reconstructive Surgery

Repair effects of rat adipose-derived stem cells on DNA damage induced by ultraviolet in chondrocytes

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