The Study Of Autophagy Induced By MicroRNA99 Derived From HIV

Objective:To explore whether the novel HIV-derived microRNA99 have ability to cause autophagy in macrophages and its possible influence on the function of autophagy,thus further study the mechanism of HIV chronic immunoactivation.Methods:1.The construction of macrophages model:With PMA stimulates the human monocytes leukemia cells differentiate into macrophages that observed by microscope.2.Macrophages be divided into experimental group(group miR99),rapamyc in group(RP)and control group.Disposal respectively in each group with miR99,rapamycin,do not add any reagents.Each group were set repeat group.3.After each treatments of 2,4,8,12,24 hours,Macrophages of each grou ops were crushed to extract protein,Macrophages in repeating group were scrap ed in 8 hours,delivering electron microscopy office for specimens.4.Autophagosomes were observed by electron microscopy and counted;int erested protein P62 and LC3 I and LC3 ? were detected by Western blot metho d and analysised by Image J software.5.Data collection,analysis and comparison between groups.Results:1.Most of THP-1 cells were transformed into macrophages.after 48 hours PMA stimulation.2.Autophagosome: compared to the normal control group,the RP group and miR99 group was obviously increased(P < 0.05),there is no significant difference between the RP group and miR99(P > 0.05).3.Interested protein expression:(1)the RP group: compared to normal control group,the expression of P62 and LC3 ? along time(2 h,4 h,8 h,12 h,24 h)decreasing(P62,P < 0.05),and increasing(LC3-?,P < 0.05)respectively.(2)miR99 group: compared to the normal control group,the expression of LC3 ? increasing over time(ditto)(P < 0.05),no significant change in P62 in each time period.(3)compare miR99 group with RP group he expression of LC3 ? increasing over time,while P62 of miR99 group is no time-varying.Conclusions:1.HIV-derived microRNA99 cause autophagy occurs in macrophages.2.HIV-derived microRNA99 could inhibit the degradation of autophagy in macrophages.3.Autophagy may be one of the repositories of HIV in macrophages4.The effects of autophagy may be involved in the mechanisms of HIV ch ronic immunoactivation.