Study On The Function Of Glucose Metabolism In Macrophage-Mediated Inflammatory Response Post Hematopoietic Stem Cell Transplantation In Mice

Hematopoietic stem cell transplantation(HSCT)is an important means to treat hematological diseases.Clinically,graft versus host disease(GVHD)and infection are common complications after HSCT.GVHD is an inflammatory disease.After the occurrence of GVHD,the probability of infection will increase,resulting in the extremely complex inflammatory response of the body.Our laboratory previously found that in a mouse hematopoietic stem cell transplantation model,the reconstructed innate immune cells(such as macrophages)in the GVHD group were much more sensitive to LPS stimulation than those in the control group.Therefore,it is speculated that the reconstructed innate immune cells are in a state of high inflammatory response.At present,the relationship between inflammatory response and glucose metabolism has been widely concerned.Based on the mouse haploidentical hematopoietic stem cell transplantation model,this study explored the correlation between the high inflammatory response state and glucose metabolism pathway in GVHD mice.Firstly,we confirmed the inflammatory response state of innate immune cells after HSCT:ELISA analysis showed:Under the stimulation of LPS in vitro,innate immune cells(macrophages and monocytes)in GVHD group had higher levels of proinflammatory cytokines TNF-α,IL-6;In the mouse model of E.coli infection established after transplantation,the mortality rate was 0 in the WT group and 100%in the GVHD group when the E.coli dose was 6×10~8 CFU.Compared with WT group,GVHD group produced higher levels of pro-inflammatory cytokines TNF-αand IL-6;In the E.coli infection model of macrophage removal in GVHD group after transplantation,it shows that macrophages play an important role in bacterial clearance and the production of inflammatory cytokines.The above results show that GVHD group is more sensitive to the stimulation of inflammatory mediators,shows a high inflammatory immune response,and macrophages play an important role in the production of proinflammatory cytokines.In order to confirm the correlation between inflammatory response and glucose metabolism after HSCT,Firstly,the blood glucose level of the body after transplantation was detected.It was found that the blood glucose level in GVHD group was significantly reduced,suggesting that there is a difference in blood glucose level between GVHD group and control group.In the early stage of the laboratory,it was found that macrophages in GVHD group polarized to M1 and blocked to M2.It has also been reported that M1 macrophages are inflammatory macrophages,which tend to glycolysis.It is speculated that GVHD group in M1-polarized state may be more inclined to glycolytic pathway;Secondly,ELISA analysis showed that WT group,Non-GVHD group and GVHD group had higher levels of pro-inflammatory cytokines under IFN-γcombined with LPS stimulation compared with LPS only stimulation group.In addition,compared with the WT and Non-GVHD groups,the GVHD group had a higher level of inflammation after co-stimulation.ELISA analysis showed that glucose significantly promoted the secretion of pro-inflammatory cytokines(TNF-α,IL-6)in murine-derived peritoneal macrophages under LPS or IFN-γstimulation in a dose-dependent manner.These results suggest that the GVHD group exhibited a higher level of inflammation under LPS or IFN-γstimulation,which may be related to glucose.Finally,the changes of glucose metabolism pathway after HSCT were further explored:Firstly,before and after LPS stimulation,the expression of glycolysis related genes,glucose consumption,lactate production and ATP content of macrophages in GVHD group were detected,which indicating that glycolysis is the main pathway in peritoneal macrophages of the GVHD group;In addition,these glycolysis inhibitors could significantly inhibit the level of pro-inflammatory cytokines in each group and improve the survival rate of GVHD group;Finally,after LPS stimulation,the mitochondrial membrane potential of peritoneal macrophages in GVHD group was decreased significantly,and the level of cell death was increased significantly,suggesting that the mitochondrial function of GVHD group is impaired after LPS stimulation.The above shows that the high inflammatory response of macrophages in GVHD group is related to the enhancement of glycolysis pathway,and there may be obstacles in oxidative phosphorylation pathway,but it needs to be further verified.In conclusion,this study preliminarily revealed the effect of changes in glucose metabolic characteristics on inflammatory response after hematopoietic stem cell transplantation.The serology and cytology of the donor-T-cell-mediated GVHD group showed a significantly high inflammatory response,which was associated with cell metabolism.Therefore, we believe that the glucose metabolic pathway can regulate the inflammatory immune response in macrophages,and the GVHD group shows a preference for using the glycolytic pathway.When the glycolytic pathway is inhibited,the inflammatory level in the GVHD group is significantly reduced,and the survival rate of the GVHD group can be improved.This study provides a new target and experimental basis for the treatment of GVHD complications.