title: Macrophage migration inhibitory factor confers resistance to senescence through CD74-dependent AMPK-FOXO3a signaling in mesenchymal stem cells.
abstract: BACKGROUND
  Mesenchymal stem cells (MSCs)-based therapies have had positive outcomes in animal models of cardiovascular diseases. However, the number and function of MSCs decline with age, reducing their ability to contribute to endogenous injury repair. The potential of stem cells to restore damaged tissue in older individuals can be improved by specific pretreatment aimed at delaying senescence and improving their regenerative properties. Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that modulates age-related signaling pathways, and hence is a good candidate for rejuvenative function.
  
  
  METHODS
  Bone marrow-derived mesenchymal stem cells (BM-MSCs) were isolated from young (6-month-old) or aged (24-month-old) male donor rats. Cell proliferation was measured using the CCK8 cell proliferation assay; secretion of VEGF, bFGF, HGF, and IGF was assessed by RT-qPCR and ELISA. Apoptosis was induced by hypoxia and serum deprivation (hypoxia/SD) for up to 6 hr, and examined by flow cytometry. Expression levels of AMP-activated protein kinase (AMPK) and forkhead box class O 3a (FOXO3a) were detected by Western blotting. CD74 expression was assayed using RT-qPCR, Western blotting, and immunofluorescence.
  
  
  RESULTS
  In this study, we found that MSCs isolated from the bone marrow of aged rats displayed reduced proliferative capacity, impaired ability to mediate paracrine signaling, and lower resistance to hypoxia/serum deprivation-induced apoptosis, when compared to younger MSCs. Interestingly, pretreatment of aged MSCs with MIF enhanced their growth, paracrine function and survival. We detected enhanced secretion of VEGF, bFGF, HGF, and IGF from MIF-treated MSCs using ELISA. Finally, we show that hypoxia/serum deprivation-induced apoptosis is inhibited in aged MSCs following MIF exposure. Next, we found that the mechanism underlying the rejuvenating function of MIF involves increased CD74-dependent phosphorylation of AMPK and FOXO3a. Furthermore, this effect was abolished when CD74, AMPK, or FOXO3a expression was silenced using small-interfering RNAs(siRNA).
  
  
  CONCLUSIONS
  MIF can rejuvenate MSCs from a state of age-induced senescence by interacting with CD74 and subsequently activating AMPK-FOXO3a signaling pathways. Pretreatment of MSCs with MIF may have important therapeutic implications in restoration or rejuvenation of endogenous bone marrow-MSCs in aged individuals.
authors.affiliation: Department of Neurosurgery, First Affiliated Hospital, Wenzhou Medical University, Wenzhou, 325000, PR China. wisdomzheng@126.com.
authors.forename: Wenzheng
authors.initials: W
authors.lastname: Xia
authors.affiliation: Key Laboratories of Education Ministry for Myocardial Ischemia Mechanism and Treatment, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, 150086, PR China. 805137949@qq.com.|Department of Cardiology, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, 150086, PR China. 805137949@qq.com.
authors.forename: Fengyun
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Department of Radiation Oncology, First Affiliated Hospital, Wenzhou Medical University, Wenzhou, 325000, PR China. xiecongying@163.com.
authors.forename: Congying
authors.initials: C
authors.lastname: Xie
authors.affiliation: Key Laboratories of Education Ministry for Myocardial Ischemia Mechanism and Treatment, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, 150086, PR China. lium_65@163.com.|Department of Cardiology, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, 150086, PR China. lium_65@163.com.
authors.forename: Miaomiao
authors.initials: M
authors.lastname: Jiang
authors.affiliation: Department of Radiation Oncology, First Affiliated Hospital, Wenzhou Medical University, Wenzhou, 325000, PR China. 244517813@qq.com.
authors.forename: Meng
authors.initials: M
authors.lastname: Hou
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000375:Aging; D000818:Animals; D000944:Antigens, Differentiation, B-Lymphocyte / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017209:Apoptosis; D001854:Bone Marrow Cells / Q000166:cytology; D015687:Cell Hypoxia; D049109:Cell Proliferation; D002478:Cells, Cultured; D015536:Down-Regulation; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000949:Histocompatibility Antigens Class II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036341:Intercellular Signaling Peptides and Proteins / Q000032:analysis / Q000235:genetics; D019746:Intramolecular Oxidoreductases / Q000378:metabolism*; D008263:Macrophage Migration-Inhibitory Factors / Q000378:metabolism*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism; D019899:Paracrine Communication; D010766:Phosphorylation; D034622:RNA Interference; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction
chemical_list: D000944:Antigens, Differentiation, B-Lymphocyte; C499515:FOXO3 protein, rat; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D000949:Histocompatibility Antigens Class II; D036341:Intercellular Signaling Peptides and Proteins; D008263:Macrophage Migration-Inhibitory Factors; C045084:invariant chain; D055372:AMP-Activated Protein Kinases; D019746:Intramolecular Oxidoreductases; C515599:Mif protein, rat