There is a close relationship between low expression of BIM and resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI). Vorinostat is a pan-histone deacetylase inhibitor (HDACi) that augments BIM expression in various types of tumor cells, however, this effect is attenuated by the high expression of anti-apoptotic proteins in EGFR-TKI resistant non-small cell lung cancer (NSCLC) cells. Vorinostat in combination with metformin - a compound that can inhibit anti-apoptotic proteins expression, might cooperate to activate apoptotic signaling and overcome EGFR-TKI resistance. This study aimed to investigate the cooperative effect and evaluate possible molecular mechanisms. The results showed that vorinostat combined with gefitinib augmented BIM expression and increased the sensitivity of EGFR-TKI resistant NSCLC cells to gefitinib, adding metformin simultaneously could obviously inhibit the expression of anti-apoptotic proteins, and further increased expression levels of BIM and BAX, and as a result, further improved the sensitivity of gefitinib both on the NSCLC cells with intrinsic and acquired resistance to EGFR-TKI. In addition, autophagy induced by gefitinib and vorinostat could be significantly suppressed by metformin, which might also contribute to enhance apoptosis and improve sensitivity of gefitinib. These results suggested that the combination of vorinostat and metformin might represent a novel strategy to overcome EGFR-TKI resistance associated with BIM-dependent apoptosis in larger heterogeneous populations.

Gliomas are the most common malignant tumors of the brain. Long non-coding RNAs (lncRNAs) play key regulatory roles in various tumors. In this study, we aimed to determine the expression and biological roles of lncRNA RMRP in glioma.

Constitutive activation of TGF‑β signaling pathway is a well-documented mechanism responsible for the bone metastasis of prostate cancer (PCa). MicroRNAs (miRNAs) have been reported to be crucial for the activation of TGF‑β signaling via targeting downstream components of TGF‑β signaling pathway. Here, we report that miR‑19a‑3p is downregulated in bone metastatic PCa tissues and cells. Upregulation of miR‑19a‑3p suppresses invasion, migration in vitro and inhibits bone metastasis in vivo in PCa cells. Conversely, silencing miR‑19a‑3p yields the opposite effect. Our results further demonstrate that miR‑19a‑3p inhibits invasion and migration abilities of PCa cells via targeting downstream effectors of TGF‑β signaling, SMAD2 and SMAD4, resulting in the inactivation of TGF‑β signaling. Therefore, our results uncover a novel mechanistic understanding of miR‑19a‑3p-induced suppressive role in bone metastasis of PCa, which will facilitate the development of effective cancer therapy methods against PCa.

Hippo-YAP1 is a tumor-suppressor signaling pathway that inhibits cell proliferation and accelerates apoptosis. However, the role of YAP1 in gastric cancer (GC) is still in dispute. Ribosomal biogenesis is closely correlated with human malignancies. HBV pre-S2 trans-regulated protein 3 (HSPC111) is a portion of an RNA-dependent complex and plays a crucial role in ribosome biosynthesis. Nevertheless, little is known about the expression and function of this factor in GC. In the present study, we evaluated the significance of YAP1 together with HSPC111 in gastric cancer. According to The Cancer Genome Atlas database, high YAP1 mRNA expression was significantly associated with poor prognosis of GC patients, and dramatically increased mRNA levels of HSPC111 are observed in GC tissues. Consistent with these findings, we detected increased expression of both YAP1 and HSPC111 in GC cell lines and clinical samples. Notably, nuclear expression of YAP1 was positively correlated with clinical stage (P = 0.041), tumor size (P = 0.023), and lymph node metastasis (P = 0.007), while HSPC111 expression was correlated with lymph node metastasis (P = 0.014). Our analyses also detected a correlation between HSPC111 expression and nuclear and cytoplasmic YAP1 in clinical samples (nuclear: r = 0.2615, P = 0.004; cytoplasm: r = -0.3721, P < 0.001) and cell lines. Finally, we showed that patients who were HSPC111- and nuclear YAP1-positive were associated with the worst prognosis (34.5 ± 4.8 months, p = 0.001), and that nuclear expression of YAP1 might act as an independent prognostic factor for GC patients.

Atherosclerotic plaque formation is characterized by the persistence of lipid-laden macrophages on the inner walls of arteries. Chronic inflammation and imbalanced macrophage function are likely to play a critical role. Herein, we investigated whether bromodomain-containing protein 7 (Brd7), a member of the bromodomain-containing protein family, regulates atherosclerosis, and if so, which mechanisms are responsible for the process. We found that Brd7 is expressed in mouse atherosclerotic plaques, and mostly in macrophages. Inhibition of Brd7 accelerates atherosclerotic lesion formation in ApoE-/- mice by promoting NF-κB-mediated inflammation. Furthermore, Brd7 inhibition alters the phenotype of macrophages and promotes plaque instability, at least partly via STAT6 signaling. Our data define a previously undescribed role of Brd7 in the development of atherosclerosis.

A novel endo-type β-agarase was cloned from an agar-degrading bacterium, Microbulbifer sp. Q7 (CGMCC No. 14061), that was isolated from sea cucumber gut. The agarase-encoding gene, ID2563, consisted of 1800 bp that encoded a 599-residue protein with a signal peptide of 19 amino acids. Sequence analysis suggested that the agarase belongs to the GH16 family. The agarase was expressed in Escherichia coli with a total activity of 4.99 U/mL in fermentation medium. The extracellular enzyme activity accounted for 65.73% of the total activity, which indicated that the agarase can be extracellularly secreted using the wild-type signal peptide from Microbulbifer sp. Q7. The agarase exhibited maximal activity at approximately 40 °C and pH 6.0. It was stable between pH 6.0 and pH 9.0, which was a much wider range than most of the reported agarases. The agarase was sensitive to some metal ions (Cu2+, Zn2+ and Fe3+), but was resistant to urea and SDS. The agarase hydrolyzed β-1,4-glycosidic linkages of agarose, primarily yielding neoagarotetraose and neoagarohexaose as the final products. These indicate that this recombinant agarase can be an effective tool for the preparing functional neoagaro-oligosaccharides.

Vascular smooth muscle cell (VSMC) phenotypic modulation is characterized by the downregulation of SMC actin cytoskeleton proteins. Our published study shows that depletion of SM22α (aka SM22, Transgelin, an actin cytoskeleton binding protein) promotes inflammation in SMCs by activating NF-κB signal pathways both in cultured VSMCs and in response to vascular injury. The goal of this study is to investigate the underlying molecular mechanisms whereby SM22 suppresses NF-κB signaling pathways under inflammatory condition. NF-κB inducing kinase (Nik, aka MAP3K14, activated by the LTβR) is a key upstream regulator of NF-κB signal pathways. Here, we show that SM22 overexpression suppresses the expression of NIK and its downstream NF-κB canonical and noncanonical signal pathways in a VSMC line treated with a LTβR agonist. SM22 regulates NIK expression at both transcriptional and the proteasome-mediated post-translational levels in VSMCs depending on the culture condition. By qPCR, chromatin immunoprecipitation and luciferase assays, we found that Nik is a transcription target of serum response factor (SRF). Although SM22 is known to be expressed in the cytoplasm, we found that SM22 is also expressed in the nucleus where SM22 interacts with SRF to inhibit the transcription of Nik and prototypical SRF regulated genes including c-fos and Egr3. Moreover, carotid injury increases NIK expression in Sm22-/- mice, which is partially relieved by adenovirally transduced SM22. These findings reveal for the first time that SM22 is expressed in the nucleus in addition to the cytoplasm of VSMCs to regulate the transcription of Nik and its downstream proinflammatory NF-kB signal pathways as a modulator of SRF during vascular inflammation.

Chronic constriction injury (CCI) of the sciatic nerve may induce dorsal root ganglion (DRG) neuronal hyperexcitability and behaviorally expressed hyperalgesia. CCI is a model of neuropathic pain. To investigate the association between the expression of protease activated receptor 2 (PAR2), TMEM16A and neuropathic pain, the expression of PAR2 and TMEM16A proteins in the DRG neurons of rats following CCI of the sciatic nerve was investigated. Following the creation of the CCI model, the thermal withdrawal latency (TWL) was examined by a hot plate test. An immunofluorescence assay and western blot assay were performed to determine the expression of PAR2 and TMEM16A proteins in the ipsilateral L4‑6 DRG neurons. The concentration of inositol 1,4,5‑triphosphate (IP3) in the L4‑6 DRG was determined by ELISA. In the CCI‑D7 (7 days after CCI) and CCI‑D14 (14 days after CCI) treatment groups, the TWL of rats was significantly shorter than that in the sham operated group (P<0.01; n=12). The expression of PAR2 and TMEM16A proteins in the CCI‑D7 and CCI‑D14 groups were significantly upregulated compared with the sham operated group (P<0.05; n=12). Additionally, it was revealed that PAR2 and TMEM16A were co‑expressed in DRG neurons. It was also observed that IP3 significantly increased in the CCI‑D7 and CCI‑D14 groups compared with the sham operation group (P<0.05; n=6) as PAR2 and TMEM16A also increased. These findings suggest that the upregulation of PAR2 and TMEM16A in DRG neurons, the co‑expression of the two proteins and increasing IP3 are critical to the development of neuropathic pain.

To study basal epithelial cell (BEC), sub-basal nerve plexus (SBN) and Langerhans cell (LC) density in patients with type 2 diabetes mellitus (T2DM) with corneal punctate epitheliopathy (CPE) and to assess their association with time to healing of CPE.

Leptin acts as a mediator of inflammation and energy homeostasis by activating leptin receptor (LEPR). We conducted this study to explore the association of polymorphisms in LEPR with type 2 diabetes mellitus (T2DM) and its related metabolic traits.

Mesenchymal stem cells (MSCs) are multipotent non-hematopoietic progenitor cells that can undergo self-renewal and differentiate into multi-lineages. Bone marrow stromal stem cells (BMSCs) represent one of the most commonly-used MSCs. In order to overcome the technical challenge of maintaining primary BMSCs in long-term culture, here we seek to establish reversibly immortalized mouse BMSCs (imBMSCs). By exploiting CRISPR/Cas9-based homology-directed-repair (HDR) mechanism, we target SV40T to mouse Rosa26 locus and efficiently immortalize mouse BMSCs (i.e., imBMSCs). We also immortalize BMSCs with retroviral vector SSR #41 and establish imBMSC41 as a control line. Both imBMSCs and imBMSC41 exhibit long-term proliferative capability although imBMSC41 cells have a higher proliferation rate. SV40T mRNA expression is 130% higher in imBMSC41 than that in imBMSCs. However, FLP expression leads to 86% reduction of SV40T expression in imBMSCs, compared with 63% in imBMSC41 cells. Quantitative genomic PCR analysis indicates that the average copy number of SV40T and hygromycin is 1.05 for imBMSCs and 2.07 for imBMSC41, respectively. Moreover, FLP expression removes 92% of SV40T in imBMSCs at the genome DNA level, compared with 58% of that in imBMSC41 cells, indicating CRISPR/Cas9 HDR-mediated immortalization of BMSCs can be more effectively reversed than that of retrovirus-mediated random integrations. Nonetheless, both imBMSCs and imBMSC41 lines express MSC markers and are highly responsive to BMP9-induced osteogenic, chondrogenic and adipogenic differentiation in vitro and in vivo. Thus, the engineered imBMSCs can be used as a promising alternative source of primary MSCs for basic and translational research in the fields of MSC biology and regenerative medicine.

This study aimed to explore the effects of overexpression of sirtuin 6 (SIRT6) on the tumorigenicity of hepatocellular carcinoma (HCC) cells.

Down syndrome (DS) is mostly caused by a trisomy of the entire Chromosome 21 (Trisomy 21, T21). Here, we use SWATH mass spectrometry to quantify protein abundance and protein turnover in fibroblasts from a monozygotic twin pair discordant for T21, and to profile protein expression in 11 unrelated DS individuals and matched controls. The integration of the steady-state and turnover proteomic data indicates that protein-specific degradation of members of stoichiometric complexes is a major determinant of T21 gene dosage outcome, both within and between individuals. This effect is not apparent from genomic and transcriptomic data. The data also reveal that T21 results in extensive proteome remodeling, affecting proteins encoded by all chromosomes. Finally, we find broad, organelle-specific post-transcriptional effects such as significant downregulation of the mitochondrial proteome contributing to T21 hallmarks. Overall, we provide a valuable proteomic resource to understand the origin of DS phenotypic manifestations.

Iron fortification of foods is currently a strategy employed to fight iron deficiency in countries. Liposomes were assumed to be a potential carrier of iron supplements.

The objective of this study was to analyze the effect of the expression of WWOX and p53 on the growth of MG-63 osteosarcoma cells and to explore the correlation between osteosarcoma and the expression of WWOX and p53. WWOX and p53-overexpressing MG-63 osteosarcoma cell lines were established by transfection and named the MW and MP cell lines, respectively. Untransfected MG-63 cells (blank control) were used as control. Quantitative polymerase chain reaction (qPCR) and western blot analysis were used to detect the expression of WWOX and wild-type p53 mRNA and protein, respectively. The effects of WWOX and p53 (wild-type) on the activity of MG-63 cells were determined by MTT assay and flow cytometry. The expression of mutant p53 protein in 65 cases of osteosarcoma was detected by immunohistochemistry to analyze the correlation between p53 and the development of osteosarcoma. qPCR showed that WWOX and p53 mRNA was overexpressed in MW and MP cells, respectively. Western blot analysis showed that the levels of WWOX and p53 protein in MW and MP cells were higher than in the blank control group. MTT assay showed that the cell proliferation ability of MW and MP cells was significantly lower than in the blank control group. Flow cytometry showed that 78.49% of MW and 66.76% of MP cells were arrested in the G0/G1 phase. Immunohistochemistry showed that mutant p53 was highly expressed in osteosarcoma, with a positive expression rate of 47.7%. The expression rate was positively correlated with the pathological grade of cancer. In conclusion, WWOX can affect the cell cycle of MG-63 osteosarcoma cells to inhibit cell proliferation, which provides new insights into gene therapy for osteosarcoma. The two types of the p53 gene have different functions in the development of osteosarcoma. Wild-type p53 acts as a tumor suppressor, while mutant p53, which is overexpressed in malignant osteosarcoma, has a carcinogenic effect associated with the degree of osteosarcoma.

Cancer stem cells are involved in cancer establishment, progression, and resistance to current treatments. We demonstrated the in vitro and in vivo anti-breast cancer effect of bakuchiol in a previous study. However, the ability of bakuchiol to target breast cancer stem cells (BCSCs) and inhibit breast cancer metastasis remains unknown. In the current study, we used the cell surface markers CD44 and CD24 to distinguish BCSCs from MCF-7 cells. Bakuchiol inhibited mammosphere formation and aldehyde dehydrogenase activity in BCSCs. Moreover, bakuchiol induced apoptosis and suppressed the mitochondrial membrane potential of BCSCs. Bakuchiol upregulated the expression levels of pro-apoptotic genes, BNIP3 and DAPK2. Bakuchiol induced oxidative stress and altered lipogenesis in BCSCs. In zebrafish xenografts, bakuchiol inhibited breast cancer cell metastasis in vivo. In addition, bakuchiol altered the expression levels of metastasis-related genes through upregulating CK18 and downregulating Notch3, FASN, TGFBR1, and ACVR1B. Our study provides evidence for the anti-breast cancer potential of bakuchiol.

The cranial suture complex is a heterogeneous tissue consisting of osteogenic progenitor cells and mesenchymal stem cells (MSCs) from bone marrow and suture mesenchyme. The fusion of cranial sutures is a highly coordinated and tightly regulated process during development. Craniosynostosis is a congenital malformation caused by premature fusion of cranial sutures. While the progenitor cells derived from the cranial suture complex should prove valuable for studying the molecular mechanisms underlying suture development and pathogenic premature suture fusion, primary human cranial suture progenitors (SuPs) have limited life span and gradually lose osteoblastic ability over passages. To overcome technical challenges in maintaining sufficient and long-term culture of SuPs for suture biology studies, we establish and characterize the reversibly immortalized human cranial suture progenitors (iSuPs). Using a reversible immortalization system expressing SV40 T flanked with FRT sites, we demonstrate that primary human suture progenitor cells derived from the patent sutures of craniosynostosis patients can be efficiently immortalized. The iSuPs maintain long-term proliferative activity, express most of the consensus MSC markers and can differentiate into osteogenic and adipogenic lineages upon BMP9 stimulation in vitro and in vivo. The removal of SV40 T antigen by FLP recombinase results in a decrease in cell proliferation and an increase in the endogenous osteogenic and adipogenic capability in the iSuPs. Therefore, the iSuPs should be a valuable resource to study suture development, intramembranous ossification and the pathogenesis of craniosynostosis, as well as to explore cranial bone tissue engineering.

Aberrant Notch signalling has been linked to many cancers including choroid plexus (CP) tumours, a group of rare and predominantly paediatric brain neoplasms. We developed animal models of CP tumours, by inducing sustained expression of Notch1, that recapitulate properties of human CP tumours with aberrant NOTCH signalling. Whole-transcriptome and functional analyses showed that tumour cell proliferation is associated with Sonic Hedgehog (Shh) in the tumour microenvironment. Unlike CP epithelial cells, which have multiple primary cilia, tumour cells possess a solitary primary cilium as a result of Notch-mediated suppression of multiciliate differentiation. A Shh-driven signalling cascade in the primary cilium occurs in tumour cells but not in epithelial cells. Lineage studies show that CP tumours arise from monociliated progenitors in the roof plate characterized by elevated Notch signalling. Abnormal SHH signalling and distinct ciliogenesis are detected in human CP tumours, suggesting the SHH pathway and cilia differentiation as potential therapeutic avenues.

COPD is an abnormal inflammatory response characterized by decreased expression of TLR2 in patients, which is suggested to induce invasive pulmonary aspergillosis (IPA). MicroRNAs (miRNAs) have been shown to play important roles in the pathogenesis of human respiratory system disorders. Therefore, the aim of this study was to identify the miRNAs involved in the regulation of TLR2 signaling in COPD.

The combined effects of matrine (Mat) and cisplatin on the survival and apoptosis of rhabdomyosarcoma (RMS) RD cells, as well as the possible mechanism of the synergistic effect of Mat and cisplatin were investigated in the present study. RMS RD cells were divided and treated as follows: control group, 5 mg/l cisplatin group, Mat groups (0.5, 1.0 and 1.5 g/l), and Mat (0.5, 1.0 and 1.5 g/l) combined with 5 mg/l cisplatin groups. An MTT assay and flow cytometry were applied to detect the survival and apoptotic rates, respectively, while RT-PCR was applied to detect the expression levels of X-linked inhibitor of apoptosis protein (XIAP) mRNA in the RD cells of each group. The survival rates of RD cells in each experimental group were lower than in the control group, and the apoptotic rates were higher than those in the control group (P<0.05). An increase in drug concentrations led to the cell proliferation inhibitory and apoptotic rates of the single Mat groups increasing as a function of dose (pairwise comparison among the groups, P<0.05), while the proliferation inhibitory and apoptotic rates of Mat combined with the cisplatin groups under different concentration were significantly higher than those of the single Mat and single cisplatin groups under the same concentration (P<0.01). The expression levels of XIAP mRNA in the RD cells of each experimental group were lower than those in the control group (P<0.05). Additionally, the expression levels of XIAP mRNA in the group treated with Mat and cisplatin were significantly lower than those of the single cisplatin and single Mat groups (P<0.01). In conclusion, Mat and cisplatin are capable of inhibiting the proliferation of RD cells and inducing apoptosis by suppressing the XIAP mRNA expression levels.