Expression Of PPAR γ In Mesenchymal Stem Cells From Patients With Aplastic Anemia And Its Role On Replacement Of Hematopoietically Active Marrow With Fat Cells

ObjectiObjective Aplastic anemia (AA) is an acquired bone marrow failure disease. Thepathogenesis of AA remains unclear. Most efforts by investigators have concentrated on theelucidation of cellular immune-mediated mechanisms of hematopoietic cell destruction.Hematogenesis negative regulators secreted by T lymphocyte induced the apoptosis of CD34+cells in bone marrow from patients with AA increase by Fas/FasL way, and resulted in thequantity of hemopoietic stem cells decrease. Immunosuppressive therapy (IST) has been used inthe treatment of AA, but there exist some nonresponders accounted for 30% of AA patients afterIST. So, are there other ways other than Fas way make excessive apoptosis of hematopoieticcells? That will be a new breakthrough in AA pathogenesis. Bone marrow biopsy important fordiagnosis of AA is characterized by a reduction of hematopoietic cells replaced by adipocytes,but the mechanisms of fatty marrow replacement remain unclear. The peroxisomeproliferator-activated receptors gamma(PPARγ), ligand-dependent transcription factors, are afamily of transcription factors belonging to the nuclear receptor superfamily. Although manytranscription factors play an important role in adipocyte differentiation and gene expression,PPARγis essential for adipocyte differentiation. This study was aimed to quantitatively detectthe expression of PPARγin bone mesenchymal stem cells (BMSCs) from patients with aplasticanemia, with PPARγligand(pioglitazone) and inhibitor(GW9662), influencing on MSCsdifferentiation into adipocytes, so as to clarify the mechanisms of fatty marrow replacement inAA patients and provide theoretical basis for the mechanisms of hematopoietic cells destructionand treatment in aplastic anemia.Methods 1. The expression of PPARγin BMSCs from AA patients:①Real time fluorescentquantitative PCR was used to detect the expression of PPARγmRNA in MSCs from patients withaplastic anemia;②Western blot was used to detect the expression of PPARγprotein in MSCsfrom patients with aplastic anemia. The role of PPARγon fatty marrow replacement: MSCs fromcontrol group were divided into three groups, namely pure induction group (MSCs in fatinductive medium), pioglitazone group (MSCs in fat inductive medium+pioglitazone) andGW9662 group (MSCs in fat inductive medium + pioglitazone+GW9662).Induction of the threegroups of MSCs into adipocytes, we counted differentiation rate of adipocytes under amicroscope and detected TNF-αexpression in differentiated cell culture supernatant. ResultResults①The level of PPARγmRNA expression in BMSCs from AA patients was higherthan that in those from controls(3.34±0.033vs1.03±0.290;p＜0.05);②The level of PPARγprotein expression in BMSCs from AA patients was higher than that in those fromcontrols(1.46±0.099vs0.86±0.064;p＜0.05);③BMSCs from control group were divided intothree groups, inducing into adipocytes. Differentiation rate of adipocytes from the pioglitazonegroup(87.42±0.67)% was higher than that of the pure induction group(44.69±2.61)% andGW9662 group(39.29±1.59)%(p＜0.05); There was no difference in statistics between the pureinduction group and GW9662 group (p＞0.05).④TNF-αexpression in cell culture supernatantfrom the pioglitazone group(95.04±3.413)pg/ml was higher than that of the pure inductiongroup(30.84±3.478)pg/ml and GW9662 group(31.43±3.508)pg/ml; There was no differencein statistics between the pure induction group and GW9662 group (p＞0.05).Conclusion The PPARγmRNA and protein highly express in AA patients, and GW9662inhibits the process of transformation from BMSCs to adipocytes, which suggests PPARγmayplay an important role in replacement of hematopoietically active marrow with fat cells and thepathogenesis in aplastic anemia.