Experimental Study On The Regulation Of LncRNA In Keratinocytes By Periplaneta Americana Extract

Objective: To explore the mechanism of Periplaneta americana extract to regulate lncRNA H19 in keratinocytes,to clarify the function of lncRNA H19,and to analyze the healing mechanism of Periplaneta americana extract.Methods: 1.Preparation of medicated serum with extract of Periplaneta americana.2.Construct lncRNA H19 overexpression and knockout plasmids and transfect them into keratinocytes(HaCaT)respectively.3.Drug intervention cells: Take the cells that grow in the logarithmic phase,divide them into a blank group and a medicated serum with extract of Periplaneta americana group,and add corresponding medicated serum for intervention.4.After the intervention of drug serum,use CCK-8 method to detect HaCaT cell proliferation,flow cytometry to detect apoptosis,scratch test to detect migration,ELISA method to detect TNF-?,IL-6 expression.Results: The CCK-8 experiment results showed that the medicated serum and blank serum of the American cockroach extract were used to intervene the cells of each group respectively.At 0h,there was no significant difference in the cell proliferation level between the American cockroach extract and the blank serum control group(p>0.05)After 24 h and 48 h intervention,the cell proliferation level of the LncRNA H19 group was significantly increased(p<0.001),and the cell proliferation of the LncRNA H19 knockout group was significantly slower(p<0.001);compared with the blank serum control group,The cell proliferation level of Periplaneta americana extract drug serum + idling group was significantly higher than that of the control group(p<0.001);compared with the LncRNA H19 knockout group alone,the periplaneta americana extract serum group+ LncRNA H19 knockout group followed the drug The intervention time was prolonged,and the cell proliferation level gradually increased;the cell proliferation level of the American cockroach extract drug serum + overexpression LncRNA H19 group was the most significant in each treatment group.Flow cytometry(Annexin,V-FITC/PI double staining method)experiments showed that after 24 hours of drug treatment,the apoptotic rate of the LncRNA H19 knockout group was 15.32%,and the apoptotic rate was significantly higher than that of the lncRNA H19 overexpression group.The apoptotic rate of the LV-OE-NC group under the intervention of the drug serum of Periplaneta americana extract was 17.77%,the apoptotic rate of the LV-sg-NC group was 17.98%,and the apoptotic rate of the two groups was the same as that of the blank serum +knockout LncRNA H19 group There is no significant difference,but compared with the corresponding blank serum idling group,the apoptotic rate was significantly increased;the apoptotic rate of the LV-OE-LncRNA H19 group was 10.65%,which was slightly lower than that of the idling group;LV-sg-LncRNA The apoptotic rate of H19 group was 33.27%,and apoptosis was the most significant in each treatment group.Scratch experiments showed that both the overexpression of LncRNA H19 and the drug serum of Periplaneta americana extract could promote the migration of HaCaT cells,and the cell migration ability of the group of Periplaneta americana extract drug serum + overexpression of LncRNA H19 was the strongest(p<0.001).ELISA experiments showed that both the drug serum of the American cockroach extract and the overexpression of LncRNA H19 can attenuate the expression of inflammatory factors TNF-a/IL-6;the expression level of the drug serum of the American cockroach extract + the overexpression of LncRNA H19 group decreased the most significantly(p<0.001);knock-out LncRNA H19 inflammatory factor expression level is up-regulated(p<0.001).Conclusion: Periplaneta americana extract may increase the expression of lncRNA H19 to promote the proliferation and migration of HaCaT cells,and promote the re-epithelialization of the wound;promote cell apoptosis in the early stage,accelerate the shedding of damaged tissues,and increase lncRNA H19 in the later stage to slow down cell apoptosis,promote tissue repair,and avoid Scar formation;up-regulate the expression of LncRNA H19,inhibit the expression of inflammatory factors TNF-a/IL-6,slow down the stimulation of HaCaT,and promote wound healing.