人胎盘胎儿侧间充质干细胞无血清培养上清的抗氧化活性分析
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摘要
背景:目前关于间充质干细胞的研究大多集中于其免疫调节功能,而关于细胞和培养上清的其抗氧化能力的研究较为少见。目的:观察人胎盘胎儿侧间充质干细胞在无血清培养条件下,其上清的抗氧化能力。方法:采用无血清培养基培养胎儿侧胎盘间充质干细胞,分别于48 h收集P2-P6代细胞培养上清。在空白培养基中添加维生素C 100μmol/L作为阳性对照。检测P2-P6代人胎盘胎儿侧间充质干细胞培养上清的总抗氧化能力、清除活性氧自由基的能力和抗氧化酶活性。结果与结论:(1)抗氧化能力:各代次人胎盘胎儿侧间充质干细胞培养上清具有一定的抗氧化能力,且不同代次间的上清具有一定的差异,其中总抗氧化能力与40-80μmol/L的维生素C相当,各代次上清均具有一定的清除二苯基苦基苯肼自由基、羟自由基、超氧阴离子自由基等自由基清除能力;(2)抗氧化酶活性分析:各代次间的人胎盘胎儿侧间充质干细胞培养上清中均可以检测到一定水平的超氧化物歧化酶和谷胱甘肽过氧化物酶活性;(3)结果提示,在无血清条件下培养胎儿侧胎盘间充质干细胞的上清具有一定的抗氧化能力和抗氧化酶的活性。人胎盘胎儿侧间充质干细胞的抗氧化能力与其旁分泌机制有关,但其中的抗氧化成分及分子机制有待进一步的研究。
BACKGROUND: Current research on mesenchymal stem cells(MSCs) is mostly focused on its immune regulatory function, while little is reported on the antioxidant capacity of the cells and culture supernatant.OBJECTIVE: To investigate the anti-oxidative capacity of the supernatant harvested from human fetal placenta MSCs(f PMSCs) under a condition of serum free culture. METHODS: f PMSCs were cultured with serum free media, and the supernatants of cells at passages 2-6 were collected at 48 hours after culture. Vitamin C was added into the culture medium, as a positive control, and its concentration was 100 μmol/L. The total antioxidant capacity, scavenging capacity of free radicals and antioxidant enzymatic activities of supernatants were measured. RESULTS AND CONCLUSION: By comparing anti-oxidative activities of vitamin C and na?ve culture medium, supernatants collected from f PMSCs cultures exhibited obvious antioxidant capacities at different extents between passages of cell cultures. The total antioxidant capacity of the culture supernatant was comparable to 40-80 μmol/L vitamin C. In addition, all supernatants derived from cells with different passages displayed capacities to scavenge free radicals, including 2,2-diphenyl-1-picrylhydrazyl radical(DPPH·), hydroxyl radical(·OH), superoxide anion radical(O2-). Even more, activities of antioxidant enzymes, including superoxide dismutase and glutathione peroxidase, were also detected in supernatants collected from different passages of f PMSCs. Under the serum-free condition, the culture supernatants of f PMSCs have antioxidant capacities at certain extent. However, the antioxidant components and underlying mechanisms need to be further studied.
BACKGROUND: Current research on mesenchymal stem cells(MSCs) is mostly focused on its immune regulatory function, while little is reported on the antioxidant capacity of the cells and culture supernatant.OBJECTIVE: To investigate the anti-oxidative capacity of the supernatant harvested from human fetal placenta MSCs(f PMSCs) under a condition of serum free culture. METHODS: f PMSCs were cultured with serum free media, and the supernatants of cells at passages 2-6 were collected at 48 hours after culture. Vitamin C was added into the culture medium, as a positive control, and its concentration was 100 μmol/L. The total antioxidant capacity, scavenging capacity of free radicals and antioxidant enzymatic activities of supernatants were measured. RESULTS AND CONCLUSION: By comparing anti-oxidative activities of vitamin C and na?ve culture medium, supernatants collected from f PMSCs cultures exhibited obvious antioxidant capacities at different extents between passages of cell cultures. The total antioxidant capacity of the culture supernatant was comparable to 40-80 μmol/L vitamin C. In addition, all supernatants derived from cells with different passages displayed capacities to scavenge free radicals, including 2,2-diphenyl-1-picrylhydrazyl radical(DPPH·), hydroxyl radical(·OH), superoxide anion radical(O2-). Even more, activities of antioxidant enzymes, including superoxide dismutase and glutathione peroxidase, were also detected in supernatants collected from different passages of f PMSCs. Under the serum-free condition, the culture supernatants of f PMSCs have antioxidant capacities at certain extent. However, the antioxidant components and underlying mechanisms need to be further studied.
引文
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[32]Quek C,Bellingham SA,Jung CH,et al.Defining the purity of exosomes required for diagnostic profiling of small RNAsuitable for biomarker discovery.RNA Biol.2016.
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[39]Tsukita S,Yamada T,Takahashi K,et al.Micro RNAs 106b and 222 Improve Hyperglycemia in a Mouse Model of Insulin-Deficient Diabetes via Pancreaticβ-Cell Proliferation.EBio Medicine.2016.
[40]Mirzaei H,Sahebkar A,Jaafari MR,et al.Diagnostic and Therapeutic Potential of Exosomes in Cancer:The Beginning of a New Tale?J Cell Physiol.2016.
[2]Poljsak B,Suput D,Milisav I.Achieving the balance between ROS and antioxidants:when to use the synthetic antioxidants.Oxid Med Cell Longev.2013;2013(1):1-11.
[3]Ni S,Wang D,Qiu X,et al.Bone marrow mesenchymal stem cells protect against bleomycin-induced pulmonary fibrosis in rat by activating Nrf2 signaling.Int J Clin Exp Pathol.2015;8(7):7752-7761.
[4]Lin KC,Yip HK,Shao PL,et al.Combination of adipose-derived mesenchymal stem cells(ADMSC)and ADMSC-derived exosomes for protecting kidney from acute ischemia-reperfusion injury.Int J Cardiol.2016;216:173-185.
[5]Dey R,Kemp K,Gray E,et al.Human mesenchymal stem cells increase anti-oxidant defences in cells derived from patients with Friedreich's ataxia.Cerebellum.2012;11(4):861-871.
[6]Liu X,Zhou L,Chen X,et al.Culturing on decellularized extracellular matrix enhances antioxidant properties of human umbilical cord-derived mesenchymal stem cells.Mater Sci Eng C Mater Biol Appl.2016;61:437-448.
[7]Park CM,Kim MJ,Kim SM,et al.Umbilical cord mesenchymal stem cell-conditioned media prevent muscle atrophy by suppressing muscle atrophy-related proteins and ROS generation.In vitro cell dev biol Anim.2016;52(1):68-76.
[8]Turkoglu A,Duru ME,Mercan N,et al.Antioxidant and antimicrobial activities of Laetiporus sulphureus(Bull.)Murrill.Food Chem.2007;101(1):267-273.
[9]Zhang Z,Liu X,Zhang X,et al.Comparative evaluation of the antioxidant effects of the natural vitamin C analog2-O-beta-D-glucopyranosyl-L-ascorbic acid isolated from Goji berry fruit.Arch Pharmacal Res.2011;34(5):801-810.
[10]Costa S,Reina-Couto M,Albino-Teixeira A,et al.Statins and oxidative stress in chronic heart failure.Rev Port Cardiol.2016;35(1):41-57.
[11]Mota S I,Costa R O,Ferreira I L,et al.Oxidative stress involving changes in Nrf2 and ER stress in early stages of Alzheimer's disease.Biochim Biophys Acta.2015;1852(7):1428-1441.
[12]Aouacheri O,Saka S,Krim M,et al.The investigation of the oxidative stress-related parameters in type 2 diabetes mellitus.Can J Diabetes.2015;39(1):44-49.
[13]Okon IS,Zou MH.Mitochondrial ROS and cancer drug resistance:Implications for therapy.Pharmacol Res.2015;100:170-174.
[14]刘冲,彭御冰,王忠.Keap1-Nrf2-ARE信号通路在多器官疾病中的研究进展[J].中国临床医学,2015,22(2):239-243.
[15]Cao W,Cao K,Cao J,et al.Mesenchymal stem cells and adaptive immune responses.Immunol Lett.2015;168(2):147-153.
[16]Baglio SR,Pegtel DM,Baldini N.Mesenchymal stem cell secreted vesicles provide novel opportunities in(stem)cell-free therapy.Front Physiol.2012;3(3):1-11.
[17]Merino-Gonzalez C,Zuniga FA,Escudero C,et al.Mesenchymal stem cell-derived extracellular vesicles promote angiogenesis:potencial clinical application.Front Physiol.2016;7:1-9.
[18]Shalaby SM,El-Shal AS,Abd-Allah SH,et al.Mesenchymal stromal cell injection protects against oxidative stress in Escherichia coli-induced acute lung injury in mice.Cytotherapy.2014;16(6):764-75.
[19]杜莉莉,吕润潇,杨晓漪,等.胎盘间充质干细胞低氧培养液对肠黏膜上皮细胞氧化应激损伤的保护作用[J].中国医科大学学报,2016,45(2):131-140.
[20]Djouad F,Plence P,Bony C,et al.Immunosuppressive effect of mesenchymal stem cells favors tumor growth in allogeneic animals.Blood.2003;102(10):3837-3844.
[21]Martinet L,Fleury-Cappellesso S,Gadelorge M,et al.Aregulatory cross-talk between Vgamma9Vdelta2 Tlymphocytes and mesenchymal stem cells.Eur J Immunol.2009;39(3):752-762.
[22]Tasso R,Augello A,Carida M,et al.Development of sarcomas in mice implanted with mesenchymal stem cells seeded onto bioscaffolds.Carcinogenesis.2009;30(1):150-157.
[23]Sabin K,Kikyo N.Microvesicles as mediators of tissue regeneration.Transl Res.2014;163(4):286-295.
[24]Sdrimas K,Kourembanas S.MSC microvesicles for the treatment of lung disease:a new paradigm for cell-free therapy.Antioxid Redox Signal.2014;21(13):1905-1915.
[25]Chen W,Huang Y,Han J,et al.Immunomodulatory effects of mesenchymal stromal cells-derived exosome.Immunol Res.2016;64(4):831-840.
[26]Ahmadi M,Rahbarghazi R,Aslani M R,et al.Bone marrow mesenchymal stem cells and their conditioned media could potentially ameliorate ovalbumin-induced asthmatic changes.Biomed Pharmacother.2016;85:28-40.
[27]Chen KH,Chen CH,Wallace CG,et al.Intravenous administration of xenogenic adipose-derived mesenchymal stem cells(ADMSC)and ADMSC-derived exosomes markedly reduced brain infarct volume and preserved neurological function in rat after acute is.Oncotarget.2016;7(46):74537-74556.
[28]Toh WS,Lai RC,Hui JH,et al.MSC exosome as a cell-free MSC therapy for cartilage regeneration:implications for osteoarthritis treatment.Semin Cell Dev Biol.2016.
[29]Reza AM,Choi YJ,Yasuda H,et al.Human adipose mesenchymal stem cell-derived exosomal-mi RNAs are critical factors for inducing anti-proliferation signalling to A2780 and SKOV-3 ovarian cancer cells.Sci Rep.2016;6:38498.
[30]Ti D,Hao H,Fu X,et al.Mesenchymal stem cells-derived exosomal micro RNAs contribute to wound inflammation.Sci China Life Sci.2016.
[31]Gopal SK,Greening DW,Rai A,et al.Extracellular vesicles:their role in cancer biology and epithelial-mesenchymal transition.Biochem J.2017;474(1):21-45.
[32]Quek C,Bellingham SA,Jung CH,et al.Defining the purity of exosomes required for diagnostic profiling of small RNAsuitable for biomarker discovery.RNA Biol.2016.
[33]Nuzhat Z,Kinhal V,Sharma S,et al.Tumour-derived exosomes as a signature of pancreatic cancer-liquid biopsies as indicators of tumour progression.Oncotarget.2016.
[34]Foj L,Ferrer F,Serra M,et al.Exosomal and Non-Exosomal Urinary mi RNAs in Prostate Cancer Detection and Prognosis.Prostate.2016.
[35]Wu J,Wang Y,Li L.Functional significance of exosomes applied in sepsis:a novel approach to therapy.Biochim Biophys Acta.2016.
[36]Nedaeinia R,Manian M,Jazayeri MH,et al.Circulating exosomes and exosomal micro RNAs as biomarkers in gastrointestinal cancer.Cancer Gene Ther.2016.
[37]Thulin P,Hornby RJ,Auli M,et al.A longitudinal assessment of mi R-122 and GLDH as biomarkers of drug-induced liver injury in the rat.Biomarkers.2016:1-9.
[38]Ming Z,Zhou R,Chen XM.Regulation of Host Epithelial Responses to Cryptosporidium Infection by Micro RNAs.Parasite Immunol.2016.
[39]Tsukita S,Yamada T,Takahashi K,et al.Micro RNAs 106b and 222 Improve Hyperglycemia in a Mouse Model of Insulin-Deficient Diabetes via Pancreaticβ-Cell Proliferation.EBio Medicine.2016.
[40]Mirzaei H,Sahebkar A,Jaafari MR,et al.Diagnostic and Therapeutic Potential of Exosomes in Cancer:The Beginning of a New Tale?J Cell Physiol.2016.