Effect Of Vitamin D On The Expression Of VDR,PPARγ And VEGFC In Diabetic Foot Ulcer Tissues And Macrophages Through VDR-PPARγ Pathway

Part oneEffect of vitamin D on the expression of Vitamin D receptor、Peroxisome proliferator-activated receptorγ、and Vascular Endothelial Growth Factor C in ulcer tissue of diabetic foot patientsObjective:To detect the level of 25 hydroxyvitamin D[25（OH）D)]in serum of patients with diabetic foot ulcer（DFU）and the levels of vitamin D receptor（VDR）、peroxisome proliferator activated receptorγ（PPARγ）、vascular endothelial growth factor C（VEGFC）in ulcer tissue expression of gene and protein,and through the intervention of vitamin D（VitD）deficiency and lack of patients,the changes of target genes and proteins before and after treatment were compared and analyzed,and the protective effect and mechanism of vitamin D on DFU were preliminarily explored.Methods:90 patients with diabetic foot ulcer in our department of endocrine department from August 2018 to January 2020 were selected and divided into three groups:30 patients with normal VitD（group A）;30 patients with insufficient VitD（group B）;30 patients with deficiency VitD（group C）.Group B and group C received intramuscular injection of VitD₂,600000 U/time,once a month,three times for 12weeks.At the same time,30 patients with normal foot skin tissue injury in burn and plastic surgery and orthopedic surgery were selected as control group（NC group）.25（OH）D was detected by chemiluminescence method,the expression of VDR,PPARγ,VEGFC gene and protein were detected by RT-PCR and Western blot.Results:（1）Compared with NC group,TG、FINS、HbA1c and HOMA-IR in group A were significantly higher,The level of QUICKI decreased（P<0.05）;Group B,TG、FINS、HbA1c and HOMA-IR level increased,The level of HDL-C、25（OH）D、QUICKI decreased（P<0.05）;Group C,TG、FINS、HbA1c and HOMA-IR level increased,The level of HDL-C、25（OH）D、QUICKI decreased（P<0.05）;（2）Compared with group A,TG、FINS and HOMA-IR were significantly higher in group B,The level of HDL-C、25（OH）D、QUICKI decreased（P<0.05）;Group C,TG、FINS、HbA1c and HOMA-IR level increased,The level of HDL-C、25（OH）D、QUICKI decreased（P<0.05）;（3）Compared with group B,TG、FINS、HbA1c and HOMA-IR were significantly higher in group C,HDL-C、25（OH）D level decreased（P<0.05）;（4）After 4 weeks of treatment,TG level decreased in group B and group C（P<0.05）;The expression of VDR,PPARγ,VEGFC gene and protein increased in group B and group C;There were no cured patients in group A,group B and group C,The difference of ulcer treatment effect in each group was statistically significant（P=0.0001）;（5）After12 weeks of treatment,the levels of TG、HbA1c and HOMA-IR in group B and group C decreased,the levels of 25（OH）D and QUICKI increased（P<0.05）;Most patients in each group recovered,There was no significant difference in the efficacy of ulcer treatment in each group（P=0.0927）;（6）Pearson correlation analysis showed that serum25（OH）D level was positively correlated with QUICKI level（r=0.247,P<0.05）,and negatively correlated with TG、LDL-C、FINS、HbA1c and HOMA-IR levels（r=-0.197,-0.168,-0.274,-0.215,-0.357,P<0.05）;（7）Logistic regression analysis showed that high levels of HbA1c,HOMA-IR,TG,LDL-C and low levels of 25（OH）D and QUICKI were risk factors for diabetic foot ulcer.Conclusion:VitD may affect the occurrence and development of DFU by affecting the expression of VDR、PPARγand VEGFC in ulcer tissue of DFU patients.Part two Effect of vitamin D on the expression of Vitamin D receptor、peroxisome proliferator-activated receptorγ、and Vascular Endothelial Growth Factor C in macrophages induced by high glucose through VDR-PPARγ pathwayObjective: To observe the effect of vitamin D on the expression of VDR、PPARγ、VEGFC gene and protein in macrophage under high glucose environment,and to explore the mechanism of vitamin D affecting DFU healing through VDR-PPARγpathway.Methods: Through concentration and time-dependent experiments,the mouse macrophage line（U937）was cultured in 25 mmol/L glucose for 24 hours.They were divided into four groups: control group（CON）,VitD group,PPARγ inhibitor（T0070907）group,VitD + T0070907 group.The levels of tumor necrosis factor（TNF-a）,interleukin-6（IL-6）and VEGFC were detected by ELISA.The expression levels of VDR、PPARγ、VEGFC gene and protein were detected by real time PCR and Western blot.Results:（1）In macrophages,VitD was expressed in a concentration and time-dependent manner,which increased the level of VEGFC and decreased the levels of TNF-a and IL-6,and the maximal effect was observed at the concentration of 0.1mmol / L for 24h;（2）Compared with CON group,the expression of VDR、PPARγ、VEGFC gene and protein in VitD group was increased（P<0.05）;there was no significant difference in the expression of VDR gene and protein in T0070907group（P>0.05）,but the expression of PPARγ and VEGFC protein was decreased（P<0.05）;the expression of VDR gene and protein in VitD + T0070907 group was increased,but the expression of PPARγ、VEGFC gene and protein was decreased（P<0.05）;（3）Compared with VitD group,the expression of VDR、PPARγ、VEGFC gene and protein in T0070907 group was lower（P<0.05）;the expression of VDR gene and protein in VitD+ T0070907 group had no significant difference（P>0.05）,but the expression of PPARγ、VEGFC gene and protein was lower（P<0.05）;（4）Compared with T0070907 group,the expression of VDR gene and protein in VitD+ T0070907 group was increased（P<0.05）,but the expression of PPARγ and VEGFC gene and protein had no significant difference（P>0.05）;（5）Spearman correlation analysis showed that vitamin D was negatively correlated with IL-6 and TNF-α（r=-0.186,-0.342,P<0.05）,and positively correlated with VDR 、 PPARγ and VEGFC（r=0.457,0.387,0.262,P<0.05）.Conclusion:VitD may affect the expression of VDR 、 PPARγ and VEGFC in macrophages in high glucose environment through VDR-PPARγ pathway,reduce inflammatory reaction.