The Effect Of Ginkgo Biloba Combined With Collagen Peptide On The Proliferation And Apoptosis Of Skin Squamous Cell Carcinoma A431 Cells

Cutaneous squamous cell carcinoma(CSCC)is a skin malignant tumor that originates in keratinocytes of the epidermis or appendages.It is similar to "flower sore" and "tumor" in Chinese medicine.In cutaneous non-melanoma,the incidence of CSCC is second only to basal cell carcinoma.It is also the leading cause of death for non-melanoma of the skin.The Mayo Clinic study showed that the incidence of CSCC increased by 263%between 1976-1984 and 2000-2010.As the population ages,this ratio is likely to increase.CSCC occurs most frequently in exposed skin areas such as the head and neck,the upper limb extension,and the back of the hand.Its clinical manifestations are mainly papules,nodules and ulcers.CSCC develops rapidly,with lymph node metastasis and visceral metastasis occurring in about 5%of patients.The treatment of CSCC includes surgery,radiation therapy,photodynamic therapy,chemotherapy,local topical medicine,CO2 laser,cryotherapy and sonodynamic therapy.Currently,the main treatment is surgical resection.However,larger tumors and extensive resection can seriously affect the patient's quality of life.Mohs microresection is time-consuming and expensive,requiring the operator to have a high degree of expertise.Traditional chemotherapy drugs reduce sensitivity and have side effects such as skin erythema,erosion,and ulcers.Clinical observations have shown that verrucous squamous cell carcinoma increases the risk of metastasis after radiotherapy.Laser and cryotherapy have a higher risk of recurrence and have been gradually replaced by other treatment methods.The efficacy of photodynamic therapy is difficult to compare with other methods.Therefore,based on the increased incidence of CSCC and the deficiencies of existing treatments,it is necessary to develop new drugs for the treatment of CSCC.Chinese medicine plays an important role in the adjuvant treatment of tumors.The etiology and pathogenesis of the tumor are mainly six kinky foreign attacks,emotional trauma,phlegm and coagulation,blood stasis block,and righteousness deficiency.The blood stasis block and the deficiency of healthy atmosphere throughout the disease,so the treatment cannot be separated from promoting blood circulation and stasis and supporting healthy atmosphere.The body's qi and blood flow more than once.If the qi is weak,the temptation is evil.Coupled with blood stasis.it gathers to form cancer.Blood stasis is the basic product of tumors.The principle of promoting blood circulation and removing blood stasis is the basic method of treating tumors.Ginkgo leaves are sweet,bitter,astringent and flat.It belongs to the heart channel and lung channel.It has the functions of promoting blood circulation and removing blood stasis,and tonifying the collaterals and relieving pain.Several experiments have reported that ginkgo biloba extract(EGb)has anti-tumor effects.Modern pharmacological studies have shown that the main components of EGb are flavonoids and terpenoids.Its anti-cancer properties are anti-oxidation,anti-angiogenesis and gene regulation.Flavonoid-containing beverages or foods can reduce the risk of various cancers including prostate cancer,lung cancer,stomach cancer,and breast cancer.In vitro experiments show that EGb761 can inhibit the proliferation and apoptosis of various tumor cells.Doctors use methods of supplementing deficiency to support righteousness.Supporting righteousness and eliminating evil complement each other.The medical doctors Zhang Yuan and Li Dongyuan of the Jin Dynasty put forward the treatment of "supporting positive accumulation and self-elimination".Professor Liu Jiaxiang proposed that "no deficiency is not a tumor." He strongly promoted the method of helping righteousness to treat tumors.Studies have shown that collagen peptides have antioxidant,immunomodulatory and anti-tumor effects.Xu Xiaobing et al found that the combination of EGb and colla corii asini hydrolysate can inhibit the proliferation of breast cancer cells MCF-7 and MDA-MB-231.In addition,several studies have shown that collagen peptides have a protective effect on HaCaT cells.Zhang Xu et al.Found that the skin peptide of Rana chensinensis can promote the proliferation of HaCaT cells.Under the stimulation of UVB,the peptide inhibited the apoptosis of HaCaT cells.Objective:We will combine promoting blood circulation and removing blood stasis and supporting healthy atmosphere to explore the effect of EGb combined with collagen peptide on the proliferation and apoptosis of A431 cells.Methods:Detection of EGb,EGb-collagen peptide on A431 cells,HaCaT cells by 3-(4,5-dimethyl2-thiazolyl)-2,5-diphenyl-2H-tetrabromide(MTT)method Effects of Proliferative Activity.The effects of EGb and EGb-collagen peptides on the apoptosis rate of cutaneous squamous cell carcinoma A431 cells were detected by flow cytometry.The effects of EGb and EGb-collagen peptides on the apoptotic morphology of A431 cells were observed by Hoechst fluorescence staining.The expression levels of Fas,Bcl-2 and BAX proteins were determined by Western blot.Results:After 48 hours of treatment with gradient concentration of EGb and combination(EGb-collagen peptides),MTT assay showed that EGb inhibited the proliferation of A431 cells and HaCaT cells;collagen peptides had a two-way effect on the proliferation of HaCaT cells;the combination could enhance EGb on A431 Cell proliferation inhibition,and can reduce the killing effect of EGb on HaCaT cells.Flow cytometry analysis showed that the combination can increase the apoptotic rate of A431 cells by EGb after 48 hours of treatment.The Hochest 33258 method was used to fluorescently stain A431 cells.Under the fluorescence microscope,it was observed that the A431 cells treated with the complex had more apoptotic morphological characteristics than EGb,including the decrease in the number of cells,and the strongly stained nuclei and debris were observed.Nuclei.Western Blot analysis showed that the combination increased Fas expression compared with EGb,and increased the relative proportion of BAX and Bcl-2.Conclusion and significance:The combination can enhance the inhibitory effect of EGb on the proliferation of A431 cells,and can reduce the inhibitory effect of EGb on the proliferation of HaCaT cells.The combination can also enhance the pro-apoptotic effect of EGb on A431 cells,and its mechanism may be related to endogenous and exogenous apoptotic pathways.This study has certain significance for the development of new drugs for skin squamous cell carcinoma in the future.