UBTF mediates activation of L3MBTL2 to suppress NISCH expression through histone H2AK119 monoubiquitination modification in breast cancer.

Lethal(3)malignant brain tumor-like protein 2 (L3MBTL2) has been related to transcriptional inhibition and chromatin compaction. Nevertheless, the biological functions and mechanisms of L3MBTL2 are undefined in breast cancer (BRCA). Here, we revealed that L3MBTL2 is responsible for the decline of Nischarin (NISCH), a well-known tumor suppressor, in BRCA, and explored the detailed mechanism. Knockdown of L3MBTL2 reduced monoubiquitination of histone H2A at lysine-119 (H2AK119ub), leading to reduced binding to the NISCH promoter and increased expression of NISCH. Meanwhile, the knockdown of L3MBTL2 decreased proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of BRCA cells, and increased apoptosis, which were abated by NISCH knockdown. Nucleolar transcription factor 1 (UBTF) induced the transcription of L3MBTL2 in BRCA, and the suppressing effects of UBTF silencing on EMT in BRCA cells were also reversed by NISCH knockdown. Knockdown of UBTF slowed tumor progression and attenuated lung tumor infiltration, whereas simultaneous knockdown of NISCH accelerated EMT and increased tumor lung metastasis. Taken together, our results show that L3MBTL2, transcriptionally activated by UBTF, exerts oncogenic functions in BRCA, by catalyzing H2AK119Ub and reducing expression of NISCH.

Baicalein promotes KDM4E to induce BICD1 and inhibit triple-negative breast cancer progression by blocking PAR1 signaling.

Baicalein has been implicated in the chemotherapy overcoming triple-negative breast cancer (TNBC). However, many unanswered questions remain regarding its role in treating TNBC. Here, we sought to demonstrate the molecular pathway mediated by baicalein in TNBC. Lysine-specific demethylase 4E (KDM4E), reduced in TNBC cells, was identified as a target protein of baicalein, and baicalein enhanced the protein expression and stability of KDM4E in TNBC cells. Knockdown of KDM4E attenuated the inhibitory effect of baicalein on TNBC cell activity, as demonstrated by intensified mobility, viability, and apoptosis resistance in TNBC cells. KDM4E activated protein bicaudal D homolog 1 (BICD1) expression by reducing the deposition of histone H3 lysine 9 trimethylation (H3K9me3) in its promoter, whereas BICD1 promoted protease-activated receptor-1 (PAR1) endocytosis and blocked PAR1 signaling through physical interaction with PAR1. Knockdown of KDM4E strengthened the PAR1-dependent activity of TNBC cells in response to thrombin activation, whereas TNBC progression activated by PAR1 signaling was blocked by combined overexpression of BICD1. Taken together, our data indicate that baicalein-promoted KDM4E enhanced the expression of BICD1 and activated the inhibitory effect of BICD1 on PAR1 signaling, thereby inhibiting TNBC progression.

Asiaticoside hampers epithelial-mesenchymal transition by promoting PPARG expression and suppressing P2RX7-mediated TGF-ß/Smad signaling in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) accounts for 10-20% of all human ductal adenocarcinomas and has a poor prognosis relative to other subtypes because of its high propensity to develop metastases. Here, the anticancer effects of asiaticoside (AC) against TNBC and the possible underlying mechanism were examined. We found that AC inhibited the TGF-ß1 expression and the SMAD2/3 phosphorylation in TNBC cells, thereby impairing the TGF-ß/SMAD signaling. AC inhibited the migration, invasion, and epithelial-mesenchymal transition (EMT) of TNBC cells by suppressing the TGF-ß/SMAD signaling. Meanwhile, AC inhibited the lung metastasis of TNBC cells in vivo and the expression of p-SMAD2/3 and vimentin, and increased the expression of E-cadherin and ZO-1 in the lung. Peroxisome proliferator activated receptor gamma (PPARG) was identified as a potential target of AC. AC increased PPARG expression, while PPARG knockdown attenuated the therapeutic effect of AC. AC-mediated PPARG overexpression suppressed the transcription of P2X purinoceptor 7 (P2RX7). The restoration of P2RX7 reversed the therapeutic effect of AC. These results suggested that AC blocked P2RX7-mediated TGF-ß/SMAD signaling by increasing PPARG expression, thereby suppressing EMT in TNBC.

miR-381 Reverses Multidrug Resistance by Negative Regulation of the CTNNB1/ABCB1 Pathway in HepG2/Dox Cells, and the Diagnostic and Prognostic Values of CTNNB1/ABCB1 Are Identified in Patients with LIHC.

Multidrug resistance (MDR) is the biggest challenge in cancer therapy. In this study, we explored the molecular mechanism of MDR in human liver cancer and explored the related diagnostic and prognostic values of the targeted genes in patients with hepatocellular carcinoma. We constructed a multidrug-resistant liver cancer cell line, HepG2/Dox, using the parental subline HepG2. The (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) (MTT) assay was used to test the viability of the liver cancer cells. Western blotting was performed to test the expression of ABCB1, ß-catenin, and ß-actin. Luciferase assays were performed to confirm the relationship between miR-381 and its target genes. The diagnostic and prognostic values of target genes were analyzed using publicly available data from The Cancer Genome Atlas. The Mann-Whitney U test and logistic regression were performed to evaluate the association between ABCB1 or CTNNB1 expression and clinical features in patients with liver hepatocellular carcinoma (LIHC). Finally, Kaplan-Meier and Cox regression analyses were performed to test the effect of ABCB1 or CTNNB1 expression on the overall survival of patients with LIHC. ABCB1 expression was upregulated in HepG2/Dox cells. ABCB1 was found to be a direct target of hsa-miR-381 and was negatively regulated by has-miR-381. Moreover, hsa-miR-381 directly targeted the CTNNB1 3' UTR and decreased the luciferase activity of CTNNB1. Transfection with miR-183 partially reversed chemotherapeutic drug resistance by downregulating the expression of ABCB1 and CTNNB1 in HepG2/Dox cells. Spearman's analysis results showed that CTNNB1 and ABCB1 were positively correlated in patients with liver cancer, and increased CTNNB1 and ABCB1 expression occurred in patients with liver cancer. High expression of ABCB1 and CTNNB1 indicated poor prognosis in patients with liver cancer; however, neither ABCB1 nor CTNNB1 expression was an independent diagnostic factor in patients with LIHC. Overexpression of hsa-miR-381 partially reversed the MDR of HepG2 cells by directly targeting and negatively regulating the expression of CTTNB1 and ABCB1. Moreover, high expression of ABCB1 or CTNNB1 indicated poor prognosis in patients with liver cancer.

SNHG1 represses the anti-cancer roles of baicalein in cervical cancer through regulating miR-3127-5p/FZD4/Wnt/ß-catenin signaling.

IMPACT STATEMENT: Baicalein exhibits anti-cancer roles in several cancers. However, the factors influencing the antitumorigenic efficiencies of baicalein in CC remain largely unclear. Here, we provide convincing evidences that lncRNA SNHG1 attenuates the tumor-suppressive roles of baicalein in CC cell viability, apoptosis, migration, and CC tumor growth. This study further demonstrates that the influences of SNHG1 in the antitumorigenic process of baicalein are achieved through modulating the miR-3127-5p/FZD4Wnt/ß-catenin axis. SNHG1 attenuates the repressive role of baicalein on Wnt/ß-catenin. Therefore, SNHG1 is a novel modulator of the tumor-suppressive roles of baicalein and SNHG1 represents a therapeutic intervention target to reinforce the tumor-suppressive roles of baicalein in CC.

Improved method of structure-based virtual screening based on ensemble learning.

Virtual screening has become a successful alternative and complementary technique to experimental high-throughput screening technologies for drug design. Since the scoring function of docking software cannot predict binding affinity accurately, how to improve the hit rate remains a common issue in structure-based virtual screening. This paper proposed a target-specific virtual screening method based on ensemble learning named ENS-VS. In this method, protein-ligand interaction energy terms and structure vectors of the ligands were used as a combination descriptor. Support vector machine, decision tree and Fisher linear discriminant classifiers were integrated into ENS-VS for predicting the activity of the compounds. The results showed that the enrichment factor (EF) 1% of ENS-VS was 6 times higher than that of Autodock vina. Compared with the newest virtual screening method SIEVE-Score, the mean EF 1% and AUC of ENS-VS (mean EF 1% = 52.77, AUC = 0.982) were statistically significantly higher than those of SIEVE-Score (mean EF 1% = 42.64, AUC = 0.912) on DUD-E datasets; and the mean EF 1% and AUC of ENS-VS (mean EF 1% = 29.73, AUC = 0.793) were also higher than those of SIEVE-Score (mean EF 1% = 25.56, AUC = 0.765) on eight DEKOIS datasets. ENS-VS also showed significant improvements compared with other similar research. The source code is available at https://github.com/eddyblue/ENS-VS.

Protective Effect of Se-Methylselenocysteine on Elaidic Acid-Induced Inflammation in Human Arterial Endothelial Cells.

This study was designed to investigate the anti-inflammatory effect of Se-methylselenocysteine (MSC) on elaidic acid (9t18:1, EA) induced human arterial endothelial cells (HAECs). MTT and flow cytometry were used to determine cell viability and cell apoptosis respectively. Western blotting was used to assess protein expression of intercellular adhesion molecular 1 (ICAM-1), E-selectin, interleukin-8 (IL-8), endothelial nitric oxide synthase (e-NOS) and phospholipases A2 (PLA2), while enzyme-linked immunosorbent assay (ELISA) was performed to examine the secretion level of nitric oxide (NO). In the cell viability assay, EA significantly decreased cell viability when compared with negative control (NC) group, and MSC effectively reversed this adverse effect, especially at the concentration of 200 µmol/L with 24 h incubation. Also, the same concentration of MSC prevented HAECs cell apoptosis induced by EA. In addition, we found that the expression of ICAM-1, E-selectin, IL-8 and PLA2 were significantly increased and e-NOS decreased in EA group compared with NC group. Inhibition of PLA2 promoted ICAM-1, E-slectin and IL-8 expression in HAECs induced by EA. And MSC down-regulated the secretion of NO level in EA-induced HAECs. Based on these results, we concluded that MSC activated PLA2 which regulated the expression of ICAM-1, E-selectin and IL-8 to protect inflammation induced by EA in HEACs.

Baicalein blocked cervical carcinoma cell proliferation by targeting CCND1 via Wnt/ß-catenin signaling pathway.

The purpose of this study was to investigate the inhibitory effect of baicalein on the proliferation of cervical carcinoma cells and stimulate cervical carcinoma cells with baicalein. MTT method was used to observe cell proliferation. Flow cytometry was used to observe cell cycle, and gene technology was used to observe the expression of corresponding genes at the level of gene and protein. ß-catenin activity was assessed using Western blot and ChIP. Baicalein suppressed cervical carcinoma cell HeLa proliferation by enhancing the activity of caspase-3. Baicalein blocked cell cycle at G0/G1 stage by inhibiting the expression of some genes. At the same time, it can prevent the nuclear translocation of ß-catenin and inhibit the activity of Wnt. When the Wnt signaling pathway is increased, the proliferation of HeLa cells is inhibited, and apoptosis is promoted in this way. In conclusion, it indicated that baicalein inhibits cervical carcinoma progression by targeting CCND1 via Wnt/ß-catenin signaling pathway.

Effect of low androgen levels on the sulphur dioxide signalling pathway in rat penile corpus cavernosum.

The aim of this study was to investigate the relationship between sulphur dioxide (SO2 ) signalling pathway and the changes in erectile function under low androgen levels. Thirty-six healthy male Sprague Dawley (SD) rats aged eight weeks were randomly divided into androgen replacement group, castration group and sham group. Rats in the androgen replacement group were subcutaneously injected with testosterone propionate at 3 mg/kg every other day postcastration. The maximum intracavernous pressure/mean arterial pressure (ICPmax /MAP) and the relative content of SO2 in the penile corpus cavernosum were measured. The mRNA and protein expressions of aspartate aminotransferase (AAT1 and AAT2), cysteine oxidase (CDO), endothelial nitric oxide synthase (eNOS) and phosphorylation of endothelial nitric oxide synthase (P-eNOS) were detected. ICPmax /MAP, P-eNOS/eNOS and the level of SO2 decreased significantly in the castration group compared to the other groups (p < 0.05). The expressions of mRNA and protein decreased significantly in the castration group compared to the androgen replacement group and the sham group (p < 0.05), while there was no significant difference between the androgen replacement group and sham group. Low androgen levels can inhibit erectile function by downregulating the SO2 signalling pathway.

Baicalein and Ly294002 induces liver cancer cells apoptosis via regulating phosphatidyl inositol 3-kinase/Akt signaling pathway.

AIM: The aim of this study is to investigate the mechanism of baicalein in inducing human liver cell line SMMC-7721 apoptosis. MATERIALS AND METHODS: Twenty micromoles baicalein or 10 µM LY294002 was adopted to treat SMMC-7721 cells. Cell proliferation was tested by cell counting kit-8 assay. Cell cycle was determined by flow cytometry and cyclin D1 expression. Cell apoptosis was detected by annexin V/propidium iodide double staining. Phosphatidyl inositol 3-kinase (PI3K)/Akt signaling pathway was assessed by real-time polymerase chain reaction and Western blot. RESULTS: Baicalein suppressed liver cancer cell SMMC-7721 proliferation and induced cell apoptosis together with LY294002. Baicalein blocked cell cycle in G0-G1 phase and downregulated cyclin D1 level. Baicalein and LY294002 significantly suppressed PI3K/Akt signaling pathway-related molecule activity at both mRNA and protein levels (P < 0.05). CONCLUSION: Baicalein can inhibit liver cancer cell proliferation and promote cell apoptosis by affecting PI3K/Akt signaling pathway together with LY294002.

Icariin improves SHR erectile function via inhibiting eNOS uncoupling.

The aim of the study was to investigate whether or not the effect of icariin on erectile function of SHR is associated with inhibition of eNOS uncoupling. Ten 12-week-old male WKY rats and 10 age-matched male SHR were evenly randomised into SHR control group, SHR+ icariin treatment group, WKY control group and WKY+ icariin group. After being treated for 4 weeks, ICPmax/MAP, the expression of NT, monomer and dimer of eNOS and the level of BH4, BH2, DHFR, NADPH oxidase and GTPCH1 in the corpus cavernosum were determined. The ICPmax/MAP and the value of BH4, DHFR and GTPCH1 in the SHR icariin treatment group were significantly higher than that of the SHR group and less than that of the WYK group and icariin-treated WKY group (p < 0.05). The value of BH2, NADPH oxidase, the ratio of eNOS monomers/dimmers and NT in the SHR icariin treatment group was significantly less than that of in the SHR group and higher than that of the WYK control group and icariin-treated WKY group (p < 0.05). Hypertension increases eNOS uncoupling in the corpus cavernosum of SHR. Inhibiting uncoupling of eNOS may be an important mechanism of icariin to improve SHR erectile function.

Effect of low androgen levels on IKca and SKca3 channels in rat penile corpus cavernosum.

We investigated whether low androgen levels affected erectile function by regulating the expressions of intermediate-conductance Ca2+ -activated K+ channel (IKca) and small-conductance Ca2+ -activated K+ channel 3 (SKca3) in corpus cavernous of rats. Thirty-six healthy male SD rats were randomly divided into the 4-week control group, 4-week castration group, 4-week androgen replacement after castration group, 8-week control group, 8-week castration group and 8-week androgen replacement after castration group, respectively. The rats in the androgen replacement groups were subcutaneously injected with testosterone (3 mg/kg) every other day after castration. After 4 and 8 weeks, maximum intracavernous pressure/mean arterial pressure (ICPmax /MAP) was measured. Expressions of IKca, SKca3, endothelial nitric oxide synthase (eNOS) and P-eNOS in penile corpus cavernosum were detected. ICPmax /MAP decreased significantly in the castration groups as compared to the control groups and the androgen replacement groups (p < 0.01). mRNA expressions of IKca and SKca3 decreased significantly in the castration groups as compared to the control groups and androgen replacement groups (p < 0.01). Protein expressions of eNOS, P-eNOS, IKca and SKca3 in the castration groups were significantly reduced as compared to the control groups and androgen replacement groups (p < 0.01). Under low androgen levels, ICPmax /MAP can be reduced by down-regulating the expressions of SKca3 and IKca, inhibiting P-eNOS/eNOS and reducing eNOS bioactivity.

Baicalein inhibits breast cancer growth via activating a novel isoform of the long noncoding RNA PAX8-AS1-N.

Baicalein, a natural flavonoid, has fascinating anti-cancer properties in breast cancer. Long noncoding RNAs (lncRNAs), a class of transcripts with no protein-coding potential, also exhibit critical roles in breast cancer. However, the molecular mechanisms mediating the anti-cancer properties of baicalein and whether lncRNAs are involved in the anti-cancer effects are still unclear. In this study, we identified a novel isoform of lncRNA PAX8-AS1 (PAX8-AS1-N), which is activated by baicalein in a dose- and time-dependent manner. Functional assays showed that PAX8-AS1-N reduced cell viability, inhibited cell-cycle progression, and induced apoptosis of breast cancer cells in vitro. Depletion of PAX8-AS1-N promoted breast xenograft tumor growth in vivo. Furthermore, depletion of PAX8-AS1-N attenuated the suppressive roles of baicalein on cell viability, the apoptosis induced by baicalein, and also the suppressive roles of baicalein on tumor growth in vivo. Mechanistically, PAX8-AS1-N bound to miR-17-5p, and up-regulated miR-17-5p targets, such as PTEN, CDKN1A, and ZBTB4. In addition, PAX8-AS1-N was down-regulated in breast cancer and reduced expression of PAX8-AS1-N indicated poor survival of breast cancer patients. In conclusion, our results demonstrated that PAX8-AS1-N activation mediated the anti-cancer effects of baicalein via regulating miR-17-5p, and suggested that baicalein and enhancing PAX8-AS1-N would be potential therapeutic strategies against breast cancer.

Effect of Low Androgen Status on the Expression of P2Y Receptors in the Corpus Cavernosum of Rats.

OBJECTIVE: To determine whether low androgen status impacts erectile function by regulating the expression of the G protein-coupled P2Y receptors (P2Y1, P2Y2, P2Y4, and P2Y6) in the corpus cavernosum penis of rats. MATERIALS AND METHODS: A total of 36 healthy, 8-week-old, male Sprague-Dawley rats were randomly allocated into 6 groups: 4 weeks of androgen replacement therapy post castration (group A); 4 weeks post castration (group B); 4 weeks post sham operation (group C); 8 weeks of androgen replacement therapy post castration (group D); 8 weeks post castration (group E); and 8 weeks post sham operation (group F). The ratio of maximum intracavernous pressure/mean arterial pressure was measured for every group and detected the expression of P2Y1, P2Y2, P2Y4, and P2Y6 in the rat corpus cavernosum penis of every group. RESULTS: The ratio of maximum intracavernous pressure/mean arterial pressure significantly declined in the castration group compared with the control group (P <.01). The expression of P2Y1, P2Y2, P2Y4, and P2Y6 and the ratio of the phosphorylated endothelial nitric oxide synthase (eNOS)/eNOS proteins were significantly lower in the castration group vs treatment or control (P <.01) whereas they were significantly lower in the group of 8 weeks vs 4 weeks post castration (P <.05). The rat serum testosterone levels in every group were positively correlated with the protein levels of P2Y1, P2Y2, P2Y4, and P2Y6. CONCLUSION: Downregulation of the expression of the P2Y1, P2Y2, P2Y4, and P2Y6 receptors that reduces the ratio of phosphorylated eNOS/eNOS and eNOS activity may be one of the important mechanisms of erectile dysfunction caused by low androgen status.

Long noncoding RNA NKILA enhances the anti-cancer effects of baicalein in hepatocellular carcinoma via the regulation of NF-κB signaling.

Hepatocellular carcinoma (HCC) is one of the most common cancer and leading cause of cancer-related death worldwide. Baicalein, a principle flavonoid, has shown attractive anti-cancer effects on HCC. However, the underlying molecular mechanisms and influencing factors contributing to the anti-cancer effects of baicalein on HCC are still largely unknown. Long noncoding RNAs (lncRNAs) have been revealed to be fascinating therapeutic targets for cancers. The roles of NF-κB Interacting LncRNA (NKILA) are recently explored in several cancers. However, the expressions, clinical significances, roles and action mechanisms of NKILA in the anti-cancer effects of baicalein on HCC are unknown. In this study, we found that NKILA is down-regulated in HCC and reduced expression of NKILA indicts poor survival of HCC patients. Functional assays showed that overexpression of NKILA enhances the roles of baicalein on HCC cell proliferation inhibition, apoptosis induction, and migration inhibition in vitro and tumor growth suppression in vivo. Conversely, knockdown of NKILA suppresses the effects of baicalein. Mechanistically, we found that NKILA inhibits IκBα phosphorylation, NF-κB nuclear translocation, and NF-κB activity. NKILA also enhances the inhibitory effects of baicalein on NF-κB signaling. Furthermore, the effects of NKILA on baicalein-induced NF-κB activity inhibition, cell growth inhibition, apoptosis induction, and migration inhibition are reversed by NF-κB nuclear translocation inhibitor JSH-23. Collectively, our data demonstrated that NKILA enhances the anti-cancer effects of baicalein on HCC in vitro and in vivo via the regulation of NF-κB signaling, and implied that the combination of NKILA and baicalein would be potential therapeutic strategies for HCC.

Baicalein induces cervical cancer apoptosis through the NF-κB signaling pathway.

To investigate the mechanism of baicalein in inducing human cervical cancer cell line C33A apoptosis. Baicalein (200 µM) was used to treat C33A cells. Cell proliferation was tested by the MTT assay. Cell apoptosis was detected by the TUNEL assay and caspase­3 activity measurement. Cell cycle was determined by flow cytometry and associated gene expression at mRNA and protein levels. Nuclear factor (NF)­κB activity was assessed by luciferase assay and western blotting. Baicalein suppressed cervical cancer cell C33A proliferation and induced cell apoptosis by activating caspase­3 activity. Baicalein blocked cell cycle in G0/G1 phase through regulating the expression of associated genes. Baicalein inhibited NF­κB activity by repressing nuclear translocation. Baicalein suppressed C33A proliferation and promoted cellular apoptosis by inhibiting NF­κB signaling pathway. In conclusion, the results indicate that baicalein can inhibit cervical cancer cell proliferation and promote cell apoptosis by affecting NF-κB activity.

Baicalein inhibits cervical cancer progression via downregulating long noncoding RNA BDLNR and its downstream PI3K/Akt pathway.

Baicalein, an active flavonoid extracted from the root of Scutellaria baicalensis Georgi, has fascinating anti-cancer effects on many cancers. Our previous study also found that baicalein inhibited cervical cancer cell proliferation and migration, and induced cervical cancer cell apoptosis and cell cycle arrest. However, the molecular mechanisms underlying the anti-cancer effects of baicalein are largely unknown. In this study, we identified a novel long noncoding RNA (lncRNA), which is downregulated by baicalein in a dose- and time-dependent manner in cervical cancer. We named this lncRNA as baicalein down-regulated long noncoding RNA (BDLNR). Gain-of- and loss-of-function assays showed that BDLNR was required for baicalein-induced cell proliferation inhibition, cell death induction, migration inhibition, and in vivo tumor growth inhibition of cervical cancer. Mechanistically, BDLNR physically bound to YBX1, recruited YBX1 to PIK3CA promoter, activated PIK3CA expression and PI3K/Akt pathway. Furthermore, BDLNR was upregulated in cervical cancer and associated with poor prognosis of cervical cancer patients. Collectively, our data demonstrated that BDLNR mediated the anti-cancer effects of baicalein in cervical cancer via activating PI3K/Akt pathway, and implied that BDLNR would be potential therapeutic target for enhancing the anti-cancer effects of baicalein in cervical cancer.

ATP2B1 gene Silencing Increases Insulin Sensitivity through Facilitating Akt Activation via the Ca2+/calmodulin Signaling Pathway and Ca2+-associated eNOS Activation in Endothelial Cells.

Endothelial cell insulin resistance may be partially responsible for the higher risk of atherosclerosis and cardiovascular disease in populations with insulin resistance and type 2 diabetes mellitus (T2DM). A genome-wide association study revealed a significant association between the ATPase plasma membrane Ca2+ transporting 1 (ATP2B1) gene and T2DM in two community-based cohorts from the Korea Association Resource Project. However, little is known about the implication of the ATP2B1 gene on T2DM. In the present study, we investigated the role of the ATP2B1 gene in endothelial cell insulin sensitivity. ATP2B1 gene silencing resulted in enhanced intracellular calcium concentrations and increased insulin-induced Akt activation compared to that in the negative siRNA-transfected HUVECs (Human Umbilical Vein Endothelial Cells). The elevated insulin sensitivity mediated by ATP2B1 gene silencing was Ca2+/calmodulin-dependent, as verified by administration of the calcium chelator BAPTA-AM or the calmodulin-specific antagonist W7. Moreover, higher levels of phosphorylation of eNOS (Ser1177) were observed in ATP2B1-silenced HUVECs. In addition to BAPTA-AM and W7, L-NAME, an eNOS antagonist, abolished insulin-induced Akt phosphorylation at Ser473 in both si-Neg and si-ATP2B1-transfected endothelial cells. These results indicate that the enhanced insulin sensitivity in ATP2B1-silenced endothelial cells is alternatively dependent on an increase in intracellular Ca2+ and the subsequent activation of the Ca2+/calmodulin/eNOS/Akt signaling pathway. In summary, ATP2B1 gene silencing increased insulin sensitivity in endothelial cells by directly modulating the Ca2+/calmodulin signaling pathway and via the Ca2+/calmodulin/eNOS/Akt signaling pathway alternatively.

The restraining effect of baicalein and U0126 on human cervical cancer cell line HeLa.

To explore the restraining effect of baicalein and the mitogen-activation protein kinase kinase inhibitor, U0126, on human cervical cell line HeLa proliferation, apoptosis and migration. HeLa cells were treated by different concentrations of baicalein or U0126. A Cell Counting Kit (CCK)­8 assay was applied to examine cell viability. Flow cytometry was used to determine cell cycle and apoptosis. A wound healing assay was performed to detect cell migration. A terminal deoxynucleotidyl transferase dUTP nick end labeling assay was adopted to test cell apoptosis. Reverse transcription­quantitative polymerase chain reaction and western blot analysis was used to detect apoptosis gene and protein expression. CCK­8 assay demonstrated that baicalein and U0126 suppressed HeLa cell viability by dose dependence. TUNEL, Annexin V­fluorescein isothiocyanate/propidium iodide, and ratio of Bcl­2­associated X protein and B cell lymphoma 2 indicated that baicalein and U0126 induced HeLa cell apoptosis. Flow cytometry revealed that baicalein blocked the cell cycle of HeLa in G0/G1 phase. A wound healing assay demonstrated that baicalein significantly inhibited HeLa cell migration compared with control. Baicalein and U0126 markedly downregulated extracellular signal­regulated kinase 1/2, matrix metalloproteinase (MMP) 2 and MMP9 levels both in mRNA and protein. In the present study, the authors demonstrated that baicalein and U0126 may be used in cervical cancer treatment by inhibiting cell migration and inducing cell apoptosis.

Magnetic nanoparticle-loaded electrospun polymeric nanofibers for tissue engineering.

Magnetic nanoparticles have been one of the most attractive nanomaterials for various biomedical applications including magnetic resonance imaging (MRI), diagnostic contrast enhancement, magnetic cell separation, and targeted drug delivery. Three-dimensional (3-D) fibrous scaffolds have broad application prospects in the biomedical field, such as drug delivery and tissue engineering. In this work, a novel three-dimensional composite membrane composed of the tri-block copolymer poly(ε-caprolactone)-poly(ethylene glycol)-poly(ε-caprolactone) (PCL-PEG-PCL, PCEC) and magnetic iron oxide nanoparticles (Fe3O4 NPs) were fabricated using electrospinning technology. The physico-chemical properties of the PCEC/Fe3O4 membranes were investigated by Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD) and differential scanning calorimetry (DSC). Morphological observation using scanning electron microscopy (SEM) showed that the composite fibers containing 5% Fe3O4 nanoparticles had a diameter of 250nm. In vitro cell culture of NIH 3T3 cells on the PCEC/Fe3O4 membranes showed that the PCEC/Fe3O4 fibers might be a suitable scaffold for cell adhesion. Moreover, MTT analysis also demonstrated that the membranes possessed lower cytotoxicity. Therefore, this study revealed that the magnetic PCEC/Fe3O4 fibers might have great potential for using in skin tissue engineering.