[Icaritin promotes chondrogenic differentiation of BMSCs by Wnt/ß-catenin signaling pathway].

To investigate the effect of icaritin (ICT) combined with GDF-5 on chondrogenic differentiation of bone marrow stromal cells (BMSCs), and discuss the action of Wnt signaling pathway, full bone marrow adherent method was used to isolate and culture SD rats BMSCs, and the cells at P3 generation were taken and divided into 6 groups： BMSCs group, ICT group, GDF-5 group, GDF-5+ICT group, GDF-5+ICT+SB216763 group, and GDF-5+ICT+ XAV-939 group. The cells were induced and cultured for 14 days. The morphology change was observed by inverted microscope. Alcian blue staining method was used to detect the changes of proteoglycans. RT-PCR was used to detect the mRNA expressions of aggrecan, Col2, Sox9, Dvl1, Gsk3ß, and ß-catenin. The protein expressions of collagen 2 (COL2) and ß-catenin were detected by Western blot. The results indicated that, compared with the BMSCs group, gradual increase was present in proteoglycan Alcian blue staining; mRNA expressions of cartilage differentiation marker genes aggrecan, COL2, Sox9 and the protein expression of COL2, as well as mRNA and protein expressions of Wnt signaling pathway-related gene ß-catenin, but with gradual decrease in Gsk3ß mRNA expressions in GDF-5 group, GDF-5+ICT group and GDF-5+ICT+SB216763 group. On the contrary, compared with GDF-5+ICT group, there was a decrease in expressions of Dvl1, and ß-catenin related to chondrogenic differentiation and Wnt signaling pathway, a increase in Gsk3ß mRNA expression, and also a decrease in protein expressions of COL2 and ß-catenin in GDF-5+ICT+XAV-939 group, with statistically significant difference between two groups. GDF-5 in combination with icaritin can induce chondrogenic differentiation of BMSCs in rats, and icaritin (ICT) can promote the chondrogenic differentiation. ICT can promote the chondrogenic differentiation of BMSCs in vitro probably by activating the Wnt/ß-catenin signaling pathway.

The PGC-1α/ERRα/ULK1 pathway contributes to Perioperative neurocognitive disorders by inducing mitochondrial dysfunction and activating NLRP3 inflammasome in aged mice.

Perioperative neurocognitive disorders (PND) are intractable, indistinct, and considerably diminish the postoperative quality of life of patients. It has been proved that Peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) was involved in neurodegenerative diseases by regulating mitochondrial biogenesis. The underlying mechanisms of PGC-1α and Nod-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome in PND are not well understood. In this study, we constructed a model of laparotomy in aged mice, and then examined the cognition changes with novel object recognition tests and fear condition tests. The protein levels of PGC-1α and NLRP3 in the hippocampus were detect after surgery. Our results showed that NLRP3 and downstream PI3K/AKT pathway expressions were augmented in the hippocampus after surgery, whereas, the expressions of PGC-1α/estrogen-related receptor α (ERRα)/Unc-51-like autophagy activating kinase 1 (ULK1) pathway were diminished after surgery. In addition, we found that NLRP3 was mainly co-localized with neurons in the hippocampus, and synaptic-related proteins were reduced after surgery. At the same time, transmission electron microscopy (TEM) showed that mitochondria were impaired after surgery. Pharmacological treatment of MCC950, a selective NLRP3 inhibitor, effectively alleviated PND. Activation of PGC-1α with ZLN005 significantly ameliorated PND by enhancing the PGC-1α/ERRα/ULK1 signaling pathway, and further suppressing NLRP3 activation. As a result, we conclude that suppression of the PGC-1α/ERRα/ULK1 signaling pathway is the primary mechanism of PND which caused mitochondrial dysfunction, and activated NLRP3 inflammasome and downstream PI3K/AKT pathway, eventually improved cognitive dysfunction.

[Inhibition of Ferulic Acid on U937 Cell Growth and Its Related Mechanism].

OBJECTIVE: To investigate the effects of ferulic acid (FA) on proliferation, apoptosis and Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway of human acute myeloid leukemia (AML) U937 cells. METHODS: Different concentrations of FA (0、10、25、50、100、200 µmol/L) was used to treat human AML cell lines Kasumi-1, HL-60, and U937 cells respectively for 24 h. The cell survival rate was detected by cell counting kit-8 method, and the 50% inhibitory concentration (IC50) of each cell line was calculated. The U937 cells were divided into control group, FA group (50 µmol/L FA), FA+pcDNA group (50 µmol/L FA+transfected with empty plasmid), FA+pcDNA-TLR4 group (50 µmol/L FA+transfected with TLR4 overexpression plasmid). Flow cytometry was used to detect U937 cell cycle and cell apoptosis; plate clone formation test was used to detect U937 cell clone formation ability; fluorescence quantitative PCR was used to detect the TLR4, NF-κB p65 mRNA levels in U937 cells; Western blot was used to detect the expression levels of CyclinD1, CyclinE, Bcl-2 related X protein (Bax), B cell lymphoma-2 (Bcl-2), Caspase-3, TLR4, and NF-κB p65 protein in U937 cells. RESULTS: With the increase of FA concentration, the survival rates of Kasumi-1, HL-60 and U937 cells gradually decreased(r=-0.919, r=-0.909, r=-0.900), the IC50 of U937 cells was 50.25±2.23 µmol/L. Compared with the control group, after drug treatment of U937 cells, the ratio of G0/G1 phase cells, apoptosis rate, expression levels of Bax and Caspase-3 proteins in FA group were significantly increased (P<0.05), the number of cell clones, the ratios of S phase and G2/M phase cells, expression levels of CyclinD1, CyclinE and Bcl-2 proteins, and TLR4, NF-κB p65 mRNA and protein were significantly decreased (P<0.05); compared with FA group and FA+pcDNA group, the ratio of G0/G1 phase cells, apoptosis rate, expression levels of Bax and Caspase-3 proteins in FA+pcDNA-TLR4 group were significantly decreased (P<0.05), the number of cell clones, the ratios of S phase and G2/M phase cells, expression levels of expression levels of CyclinD1, CyclinE and Bcl-2 proteins, and TLR4, NF-κB p65 mRNA and proteins were significantly increased (P<0.05). CONCLUSION: FA inhibits U937 cell proliferation and promotes cell apoptosis by inhibiting the TLR4/NF-κB signaling pathway.

Circ_0084188 promotes colorectal cancer progression by sponging miR-654-3p and regulating kruppel-like factor 12.

To investigate the biological role and mechanism of circ_0084188 in colorectal cancer (CRC). Real-time quantitative polymerase chain reaction and western blot assay were used to detect RNA levels and protein levels in CRC cell lines (HCT116 and SW480), respectively. Cell proliferation was evaluated by Cell Counting Kit-8 assay, 5-ethynyl-2'-deoxyuridine assay, and colony formation assays. Cell apoptosis was determined using flow cytometry. Cell migration and invasion were measured by transwell assay. Sphere formation efficiency was determined by sphere formation assay. The interaction between microRNA-654-3p (miR-654-3p) and circ_0084188 or Kruppel-like factor 12 (KLF12) was confirmed by a dual-luciferase reporter, RNA immunoprecipitation and RNA pull-down assays. Xenograft in CRC mice model was utilized for exploring the role of circ_0084188 in vivo.Circ_0084188 was overexpressed in CRC tissues and cells. Circ_0084188 silencing suppressed cell proliferation, migration, invasion, and stemness and induced apoptosis in CRC cells. Circ_0084188 acted as a sponge for miR-654-3p, and circ_0084188 regulated CRC cell behaviors via sponging miR-654-3p. Moreover, KLF12 was a target of miR-654-3p, and miR-654-3p overexpression inhibited the malignant behaviors of CRC cells by downregulating KLF12. Mechanically, circ_0084188 sponged miR-654-3p to regulate KLF12 expression in CRC cells. In addition, circ_0084188 downregulation inhibited tumor growth in vivo.Circ_0084188 knockdown might repress CRC progression partially via regulating the miR-654-3p/KLF12 axis, providing a novel insight into the pathogenesis of CRC.

microRNA­145 modulates migration and invasion of bladder cancer cells by targeting N­cadherin.

MicroRNA (miRNA)­145 has been demonstrated to serve a role in several types of tumors, however, the potential molecular mechanism of action of miRNA­145 in bladder cancer metastasis remains to be elucidated. This study aimed to investigate the potential modulation of miRNA­145 in bladder carcinoma and elucidate the underlying molecular mechanism. The expression of miRNA­145 in bladder adenocarcinoma tissues and bladder cancer cells was measured by reverse transcription­quantitative polymerase chain reaction. miRNA­145 mimics and inhibitor were transfected into bladder cancer (BC) cells to determine the role of miRNA­145 on cell motility and invasion measured by wound healing and transwell assays. Luciferase assay was performed to confirm whether N­cadherin was the direct target of miRNA­145. Subsequently, expression of N­cadherin and matrix metalloproteinase­9 (MMP9) in BC cells were detected by western blot analysis. miRNA­145 was significantly downregulated cells and tissues from patients with BC, compared with healthy controls. miRNA­145 markedly inhibited the ability of BC cells to migrate and invade. Furthermore, N­cadherin was identified as a target of miRNA­145 in BC cells. MMP9, acting downstream of N­cadherin, was downregulated in BC cells by miRNA­145. In the present study, miRNA­145 suppressed the migration and invasion of BC cells by regulating N­cadherin. The results of the present study indicated that miRNA­145 may function as a tumor suppressor and may have a potential to be a diagnostic and predictive biomarker, and a therapeutic target for treatment of BC.