| Diabetes mellitus(DM),one of the most prevalent endocrine diseases worldwide today,is a metabolic disease mainly characterized by hyperglycemia,usually resulting from defects in insulin secretion or impairment of its biological actions.Long standing hyperglycemia can lead to chronic damage and dysfunction in various tissues and organs,especially the eye,kidney,heart,blood vessels,nerves,meantime epidemiological and laboratory studies have provided evidence proved that hyperglycemia plays an important role in the development and progression of tumors as an independent risk factor.At present,there has been great progress in the study of the effects of hyperglycemia on the occurrence and development of tumors.Prolonged hyperglycemic state could promote the proliferation,metastasis,and anti apoptotic effects of tumor cells and even promote the expression of oncogenes to a certain extent.The hyperglycemic state can also affect immune system to some extent.Some studies have shown that the proinflammatory effect produced by the increased secretion of some cytokines,such as TNF-αand IL-6,in the hyperglycemic state is able to stimulate oncogene expression and tumor proliferation,while the hyperglycemic state also has some degree of inhibitory effect on macrophages,neutrophils,which is one of the reasons for tumor susceptibility in diabetic patients.However,for some immune cells that play a role in tumor immunity,such as dendritic cells(DCs),T cells and NK cells,there is still a certain controversy about the specific effect of hyperglycemia on their function.Therefore,this paper mainly explores the effect of long-term hyperglycemic state on the function of DCs and T cells.In order to verify the effect of hyperglycemia on immune cells and the impact on tumorigenesis and progression under diabetic conditions in vivo,we established a type 1diabetes mellitus(T1DM)mouse model by intraperitoneal injection of streptozotocin(STZ).STZ,which can be used as a broad-spectrum antibiotic,also has some toxicity and is able to destroy pancreaticβcells to artificially create symptoms of T1DM such as insulin deficiency,hyperglycemia,polydipsia and polyuria.We subcutaneously inoculated a certain amount of B16-OVA melanoma cells into T1DM mice and C57 mice of the same age to construct the tumor model respectively.The results show that the tumor growth of T1DM mice was significantly faster than that of WT mice,and the mortality rate was higher.This phenomenon proved hyperglycemia have a promoting effect on the occurrence of tumor development,but whether it plays an important role in the process of cancer development of DC and T cells function is still uncertain.In order to find out,we performed a long-term dynamic monitoring of the functional changes of immune cells in vivo in T1DM mice.To determine whether hyperglycemia has any effect on DC function,we used magnetic beads sorting to obtain CD11c~+DCs from the spleens of T1DM and WT mice.By examined the expression of cell surface molecules,cytokine secretion levels after activation with LPS,we found that the expression of CD40,CD86,MHC class II molecules on the surface of DCs,as well as the secretion levels of cytokines such as TNF-αand IL-6,were not significantly different after activation of splenocytes from T1DM mice at the early stage of diabetes(2 weeks after building a successful model)compared with WT C57 mice.However as time extented,TNF-αand IL-6 secretion level by splenic CD11c~+DCs from T1DM mice was significantly reduced.This indicates that prolonged hyperglycemia is able to exert an inhibitory effect on DCs in vivo.To investigate the effects of high glucose on DC development and function,we stimulated bone marrow-derived dendritic cells(BMDCs)obtained from wild-type mice in vitro with various concentrations of glucose for a short time to observe changes in cell phenotype,inflammatory factor secretion,and its antigen-presenting capacity.Preliminary results showed that when stimulated with 25 mmol/L glucose,BMDCs displayed upregulation of the expression levels of CD40 and CD86 as well as MHC class II molecules,increased secretion of cytokines TNF-αand IL-6.Examination of DC antigen-presenting function found that DCs stimulated with 25 mmol/L glucose for a short time could enhance its function of proliferating T cells and promoted T cell surface molecule expression and cytokine secretion.To investigate whether prolonged culture with high glucose could affect DC development and its function,we cultured bone BMDCs from wild-type mice with medium containing various concentrations of glucose,and found that high glucose had no significant effect on the induced development of BMDCs,25 mmol/L glucose culture could promote DCs’CD40,CD86,as well as MHC-II expression and secretion of cytokines such as TNF-αand IL-6and its antigen-presenting function.This demonstrates that glucose can have a promoting effect on BMDC function to some extent.However,the detection of the ability of DCs to induce Th cells revealed a concomitant decrease in the level of Th cells induced by DC induction with increasing concentrations of glucose in the culture medium.At different time points after the model was successful constructed,we uses magnetic beads separated CD8~+T cells and CD4~+T cells from spleens,and measure cell surface molecule expression and cytokine secretion levels after activation.We found that in early stage oof diabetes(2 weeks after the model was build),There was also no clear change trend in the secretion levels of other cytokines,cell surface molecules,nor in the proportion of IFN-γ~+CD8~+T cells.This result may suggest that at the early stage of diabetes,short period of high blood glucose is difficult to affect the function of immune cells in vivo.However,examination of splenocytes from mice 4 weeks and 8 weeks after successful modeling revealed that the surface CD107a expression level on CD8~+T cells decreased in the spleen of T1DM mice,the proportion of IFN-γ~+CD8~+T cells was significantly reduced,and granzyme B secretion in the supernatant was reduced.This result shows that in a long-term hyperglycemic state,there is a certain degree of decline in immune cell function,and the body’s immune system begins to show a state of suppression.After the analysis of DC and T cells in the tumor tissues of T1DM mice and WT mice,it was found that there was no significant difference in the proportion of relevant immune cells in the tumor tissues of mice,but the proportion of IFN-γ~+CD8~+T cells in the tumor infiltration of T1DM mice was reduced.These results provide further evidence that chronic hyperglycemia has an inhibitory effect on immune cell function.Furthermore,to compare whether there are differences in the killing function of murine CD8~+T cells in the hyperglycemic state,we generated a T1DM model using OT-I mice,in which both T1DM CD8~+T cells and normal CD8~+T cells from OT-I mice maintained in hyperglycemia for 3 weeks were injected via the tail vein into a WT B16-OVA tumor model and tumor size was continuously monitored.The tumor size of the T1DM CD8~+T cell reinfusion group was still larger than that of the control group.This further verified that long-term high glucose status would exert inhibitory effects on T cell function and promote tumorigenesis and development.In summary,this paper through in vitro and in vivo tests shows that a short time of a certain degree of high glucose stimulation is able to promote the antigen-presenting function of DCs,but a long time of high glucose culture is able to inhibit DC maturation.At the same time,long time hyperglycemic state could promote the occurrence and development of tumors by inhibiting the function of DCS and T cells,and further specific mechanisms are still under investigation.In this paper,we hope to provide new ideas for the development and treatment of cancer in diabetic patients through the study of immune cell function under high glucose condition. |
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