ABSTRACT
Objective:To explore the effect of rapamycin on 1-methyl-4-phenylpyridinium iodide (MPP+ )-induced activation of Nod-like receptor protein 3 (NLRP3) inflammasome in microglia.Methods:The BV2 microglia cells were divided into control group, model group and rapamycin group.The model group and rapamycin group were treated by MPP+ to activate NLRP3 inflammasome, and rapamycin group was pretreated with rapamycin.Quantitative real-time PCR (RT-qPCR) was used to detect the mRNA levels of NLRP3, apoptosis-associated speck-like protein (ASC) and caspase-1.Immunofluorescence was used to detect the protein expression of NLRP3 and interleukin-1β (IL-1β). Western blot was carried out to assess the protein expression of NLRP3, ASC, caspase-1, beclin1 and microtubule-associated protein 1 light chain 3 (LC3).Results:The mRNA levels of NLRP3, ASC and caspase-1 in model group were higher than those in control group ( t=4.825, 3.015, 5.853, all P<0.05). The mRNA levels of NLRP3 and caspase-1 in rapamycin group were lower than those in model group ( t=2.75, 2.89, both P<0.05). In model group, the protein expressions of NLRP3 (1.54±0.22), ASC (1.02±0.13) and caspase-1 (1.42±0.30) were higher than NLRP3 (0.66±0.15), ASC (0.41±0.14) and caspase-1 (0.70±0.10) in control group ( t=5.653, 5.602, 3.964, all P<0.01), while the protein expression of beclin1 (0.28±0.09) and LC3II/LC3I ratio(0.69±0.14) were lower than beclin1 (0.60±0.11) and LC3II/LC3I (1.29±0.23) in control group ( t=4.010, 3.982, both P<0.01). The protein expressions of NLRP3 (0.80±0.18) and ASC (0.68±0.14) in rapamycin group were lower than those in model group ( t=4.413, 3.077, both P<0.05), while the protein expression of beclin1 (0.65±0.20) and LC3II/LC3I ratio(1.42±0.36) were higher than those in model group ( t=2.965, 3.278, both P<0.05). Conclusion:MPP+ activates NLRP3 inflammasome and impairs autophagic function in microglia.Rapamycin inhibits MPP+ -induced activation of NLRP3 inflammasome by restoring autophagic impairment in microglia.
ABSTRACT
OBJECTIVE@#To investigate the profile and potential significance of PTEN and NBS1 mutations among patients with familial or at early onset breast cancer in Hunan province. @*METHODS@#A total of 131 breast cancer patients with familial history or suffered from breast cancer at the age of less than 35 years old were included in this study. A comprehensive phosphatase and tensin homolog (PTEN) and nibrin (NBS1) mutation analysis was performed through denaturing high performance liquid chromatography (DHPLC) and subsequent DNA direct sequencing. @*RESULTS@#Among 131 patients, a reported mutation IVS4+109insTCTTA in PTEN gene were identified in two patients. The mutation frequency of IVS4+109insTCTTA was 1.15%. Two mutations in PTEN gene, 225 A>C (Thr 160 Pro) and IVS5+13T>C, was firstly discovered. Another reported missense mutation was rs121909229 G>A (Arg 130 Gln). Three mutations were detected in NBS1 gene, of which IVS6+43A>G and IVS6+127A>G were firstly discovered and another reported synonymous mutations was rs1805794 G>C (Glu 185 Gln). @*CONCLUSION@#The novel mutations in PTEN and NBS1 might be specific to the familial and early-onset breast cancer of Chinese Hunan population.