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• W2547213275 abstract "Abstract Background: Our preliminary research found that STAT3, IL-17A, and IL-21 expressed in cGVHD patients. So, we provide the blocking STAT3 signal to the induction of Treg cells differentiation, and to provide experimental basis on new targets of cGVHD immunotherapy. Methods: 1. Mice spleen CD4+ CD62L+ naiveT cells were separated by immune magnetic bead and then activated for 72 h. After 96 h infection with STAT3-shRNA and negative control lentivirus, the Th subgroup proportion were measured by flow cytometry. Th related cytokines levels test by Luminex. Real time quantitative PCR was to detect STAT3 and Th subgroup related transcription factor mRNA levels. CD117+ mouse bone marrow stem progenitor cells were sorted by flow cytometry, and transfected by STAT3-shRNA. Inhibition of STAT3 gene in mRNA level was measured at 96 h. Cell proliferation activity was test with CCK8 kit, and cell apoptosis rate determined by flow cytometry. Differentiation of CD117+ cells was induced by 2.2% of methyl cellulose and different cytokines. 2. BALB/c female mice, after the linear accelerator 700cGy of whole body irradiation, accepted miHA mismatched male B10. D2 mice bone marrow cells and spleen cells (8 x 106, 1:1). Randomly assigned 6 mice of cGVHD clinical score of 0.6 or above to each group. After STAT3-shRNA or negative control lentivirus treatment, the observe end point was 58th day after transplantation. The clinical and pathologic scores compared. Th17 and Treg cells measured by flow cytometry. Th related cytokines measured by Luminex. Purpose genes in blood and protein expression levels in target organs were found by Q-PCR and western blot test, respectively. Results: 1. The Th17 / Treg ratio of shRNA group was significantly decreased than that in the NC group (P < 0.05). Except for the Foxp3 gene, other purpose genes, including T-bet, Gata3, RORγt, TGFβ, Notch1, and Jagged2 mRNA levelsin interference group were cut. GM-CSF, IFN-gamma, beta, IL- 3, IL-2, IL-4, IL-6, TNF alpha, IL-17, IL-22a, IL-27, and IL-9 factor expression levels were significant difference between shRNA and negative control group (P < 0.05). There was no significant difference of cell proliferation activity, early apoptosis rate, and differentiation ability in STAT3-shRNA treated CD117+ bone marrow, compared with negative control group and blank control group (P > 0.05). 2. After 50th day, shRNA treatment group appear hair recovery, energy recovery, weight gain, shortness of breath better, mean of cGVHD score decreased. At the 58th day, clinical scores of cGVHD between shRNA treatment group and the negative control group overall mean difference was statistically significant (P < 0.05). cGVHD pathological score of lungs in shRNA treat group reduced (P < 0.05). STAT3mRNA levels in peripheral blood, phosphorylated STAT3, and STAT3 expression level of lung declined than control groups. The proportion of Th17 / Treg cells of spleen was significant reduced in shRNA group, compared with negative control group (P < 0.05). Conclusion: 1. STAT3 knocking down in naïve CD4 + Th cells induced the increased Treg cells, and the decreased Th17 cells. IL-2 confirmed to promote the growth of Treg cells. It speculated that blocking STAT3 might bring Th9 cells differentiation. STAT3 blocking in CD117+ stem progenitor cells have no significant effect on the proliferation, apoptosis and differentiation, validation the safety of STAT3-shRNA. 2. STAT3-shRNA treatment cGVHD mice in vivo achieved curative effect. The main target organs was the lung, which might be closely related to the fall in the proportion of Th17 /Treg. STAT3 may be used as a new target for immunotherapy of cGVHD. Acknowledgment The project was sponsored by grants from National Natural Science Foundation of China (No. 30972790; No.81270648; No.81370665; No.81300446), Provincial Natural Science Foundation of Guangdong (No. S2012010009560), Provincial Science and Technology Planning Project of Guangdong (No.2013B021800186; No.2013B021800201), and Science and Technology Planning Project of Guangzhou (No. 201400000003-4, 201400000003-1). Disclosures No relevant conflicts of interest to declare." @default.
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• W2547213275 date "2015-12-03" @default.
• W2547213275 modified "2023-09-26" @default.
• W2547213275 title "Blocking Stat3 to Regulate the Balance of Th17/Treg in Cgvhd Therapy" @default.
• W2547213275 doi "https://doi.org/10.1182/blood.v126.23.5420.5420" @default.
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