Effect Of FoxC2 From Cutaneous Squamous Cell Carcinoma Exosomes On Biological Behavior Of Lymphatic Endothelial Cells And Its Expression In Carcinoma Tissue

Objective: Forkhead box C2(FOXC2)is a member of forkhead box transcription factor family.It plays an important role in the development of lymphatic vessels,and participates in tumor proliferation,metastasis,angiogenesis and lymphangiogenesis by inducing epithelial to mesenchymal transition.Cutaneous squamous cell carcinoma(CSCC)is a malignant tumor originating from epidermal or appendage keratinocytes.Domestic research has found that in non melanoma skin tumors,CSCC incidence rate is second,next only to basal cell carcinoma.The purpose of this study was to investigate the effect of FOXC2 on the biological behavior of lymphatic endothelial cells(LECs)and the relationship between the expression of FOXC2 and clinical and pathological features in CSCC.Methods:(1)Lentivirus was used to knock down FOXC2 gene of CSCC cells to construct CSCC shfoxc2,and the proliferation ability of CSCC shfoxc2 in control group was compared with that in experimental group;(2)The exosomes of csccgfp cells and csccsshfoxc2 cells were extracted by exosome kit;The morphology of exosomes was observed by transmission electron microscope(TEM)and the size of exosomes was analyzed by nanoparticle tracer analyzer;Western blot was used to detect the expression of FOXC2,CD9 and CD81;(3)The exosomes of csccgfp cells and csccsshfoxc2 cells were co cμltured with LECs,and the biological behaviors of LECs such as proliferation,migration and tubμlation were detected;(4)Methods: 30 patients with primary CSCC who underwent surgical resection in our hospital from 2013 to 2020 were selected and divided into two groups according to their clinical and pathological characteristics;(5)The expression levels of FOXC2 and lymphatic endothelial hyaluronic acid receptor 1(LYVE-1)in CSCC were detected by immunohistochemistry,and the lymphatic vessel density(LVD)was calcμlated by LYVE-1 labeled lymphatic vessels The expression levels of LVD and FOXC2 were compared between pathological groups,and the differences of LVD between high expression group and low expression group of FOXC2 were compared.Resμlts:(1)After knockdown of FOXC2 gene,the proliferation of CSCC cells decreased significantly;(2)The resμlts of transmission electron microscopy and nanoparticle tracking showed that the exosomes of CSCC cells were spherical in shape,and the diameter of the particles ranged from 50 nm to 170 nm;Western blot resμlts showed that the exosomes of CSCC cells were highly expressed in CD9 and CD81;(3)CSCC exosomes can promote the proliferation,migration and tube formation of LECs;The ability of csccshfoxc2 exosomes to influence the biological behavior of LECs was significantly decreased;(4)Of the 30 CSCC patients,5 were younger than 65 years old,18 were 65-80 years old,and 7 were older than 80 years old;There were 19 males and 11 females;There were 9 cases of lymphatic metastasis and 21 cases of non metastasis;The tumor diameter was less than 3cm in 8 cases,3-5cm in 16 cases,and more than 5cm in 6 cases;There were 18 cases of high differentiation and 12 cases of low differentiation;(5)There was no significant difference in the expression of FOXC2 in CSCC patients with different age,gender and tumor size(P > 0.05);The expression level of FOXC2 in middle and low differentiation group and metastasis group was higher than that in high differentiation group and non metastasis group(P < 0.05).The LVD of FOXC2 high expression group,low differentiation group and metastasis group was higher than that of FOXC2 low expression group,high differentiation group and non metastasis group(P < 0.05);There was no significant difference in LVD among CSCC patients with different age,gender and tumor size(P > 0.05).Conclusions: FOXC2 is involved in maintaining the normal proliferation of CSCC cells;FOXC2 transported by CSCC exosomes promoted the proliferation,migration and tube formation of LECs;The expression level of FOXC2 is related to the lymphatic vessel growth,lymph node metastasis and tissue differentiation of CSCC,suggesting that FOXC2 may be an important marker of the malignant degree and lymphatic metastasis of CSCC.It can provide new theoretical basis and strategies for the pathogenesis,diagnosis and treatment of CSCC.