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Introduction. LAMA1, also known as laminin subunit α1, is a member of the laminin family, which is widely reported to be a key basement membrane molecule that affects various biological activities and is associated with many kinds of diseases. We aimed to investigate the association between LAMA1single-nucleotide polymorphisms and the occurrence and progression of esophageal squamous cell carcinoma in the Chinese population. Method. 2,186 participants were collected retrospectively between October 2008 and January 2017, including 1,043 ESCC patients and 1,143 noncancer patients. A 2 mL blood sample was obtained intravenously for the LDR for SNP analysis. The 6 SNP loci of LAMA1 were selected and examined. We analyzed the association of several genetic models of 6 LAMA1 SNP loci, sex, age, smoking and drinking status, and the occurrence of esophageal squamous cell carcinoma. Results. In the rs62081531 G > A locus, genotype GA was a protective factor for ESCC compared with GG (OR: 0.830, P = 0.046), especially among the younger and nondrinkers. At rs607230 T > C, genotype TC was linked with a lower risk of ESCC compared with TT. (OR: 0.613, P = 0.034). Haplotype Frequencies revealed that Ars62081531Grs621993Ars539713Trs566655Ars73938538Crs607230 (OR: 0.803, P = 0.028) and Grs62081531Grs621993Ars539713Trs566655Crs73938538Crs607230 (OR: 0.679, P = 0.010) were strongly associated with lower susceptibility of ESCC. Conclusion. The LAMA1 rs62081531, rs539713, rs566655, and rs607230 polymorphisms were demonstrated to be related to susceptibility to ESCC in the Chinese population. LAMA1 SNPs may have a significant impact on the occurrence of esophageal cancer and may serve as potential diagnostic biomarkers. [ABSTRACT FROM AUTHOR]

J Oral Pathol Med (2012) 41: 322-331 Background: To clarify the efficacy of grape seed procyanidin (GSP) on antiproliferative effects related to p53 functional status of oral squamous cell carcinoma (OSCC) for its chemoadjuvant potential. Methods: We used GSP to investigate SCC-25 cells with wild-type p53 gene and OEC-M1 cells with mutant p53 gene for the assessment of antiproliferative effects including cell viability, cell cycle, apoptosis, migration and invasion potential, and alterations of associated oncoproteins involved in cellular and molecular events. Results: The findings suggest that GSP on OEC-M1 cells leads to cell cycle arrest by increasing the expression of p21Cip1/p27Kip1 protein without functioning mitochondria-mediated apoptosis, whereas GSP on SCC-25 cells inhibits cell proliferation via both G1-phase arrest and mitochondria-mediated apoptosis in a dose-dependent manner as a result of alterations of Bcl-2. GSP also inhibits the migration and invasion of both cells, which are associated with the suppression of matrix metalloproteinases (MMPs), MMP-2 and MMP-9. Conclusion: Antiproliferative effectiveness of GSP is closely associated with the p53 status of OSCC cells. GSP displays chemoadjuvant potential via cell cycle blockage and apoptotic induction. Our findings clearly suggest that GSP may play a role as a novel chemopreventive or therapeutic agent for OSCC. [ABSTRACT FROM AUTHOR]

Background: Oral squamous cell carcinoma (OSCC) is one of the most common head and neck cancers. The prognosis of OSCC is usually poor because of extensive local invasion at initial diagnosis. In the literature, Fascin has been reported responsible for cell motility and over-expression of Fascin contributes to an unfavorable clinical course. Nevertheless, the roles of Fascin protein playing in aggressiveness of OSCC and their potential mechanisms need to be elucidated. Methods: Two cell lines of OSCC (OECM-1 and SCC-25) via the vector-based small interfering RNA (siRNA) to suppress the expression of the Fascin gene were used. Subsequent analyses and observation regarding the expression of Fascin protein and cyto-morphological alterations were detected by Western blot and immunofluorescent microscopy. Boyden chamber invasion assay, cell migration assay and adhesion assay were also applied to investigate the functional changes of OSCC. Results: There were statistically significant differences ( P < 0.05) of Fascin expression before and after silencing. Down-regulation of Fascin protein directly led to changes of cell surface protrusions under immunofluorescent microscopy and resulted in suppression of migration, invasion and increase of adhesion in both cell lines ( P < 0.05). Furthermore, down-regulation of Fascin expression also resulted in alterations of E-cadherin, β-catenin and TWIST at certain level, implicative of an association with epithelial-mesenchymal transition (EMT). Conclusions: Our results suggest that expression of Fascin protein may play an essential role in regulation of progression of OSCC and contributes to the event of EMT in the early aggressiveness of OSCC. [ABSTRACT FROM AUTHOR]

Simple Summary: Certain tumor aggressiveness-associated mediators from cancer-associated fibroblasts (CAFs) in tumor microenvironment have been reported. Using gene expression analysis, we identified that CAFs overexpress Chemokine ligand 11 (CCL11), which is associated with tumor migration and invasion, increased expression of cancer stem cell properties, and induction of the epithelial-to-mesenchymal transition. Neutralization of CAF-induced CCL11 reversed the aggressive phenotype of cancer cells. Based on the immunohistochemical staining of clinical samples, we found that increased co-expression of CCL11 and its receptor, C-C Motif Chemokine Receptor 3 (CCR3), was associated with poor overall survival. Our results suggest that targeting CCL11-CCR3 signaling is a potential therapeutic strategy for patients with aggressive head and neck cancer. The tumor microenvironment (TME) plays a crucial role in tumor progression. One of its key stromal components, cancer-associated fibroblasts (CAFs), may crosstalk with cancer cells by secreting certain cytokines or chemokines. However, which important mediator(s) are released by CAFs, and the underlying molecular mechanism, remain largely unknown. In the present study, we isolated patient-derived CAFs and normal fibroblasts (NFs). Using microarray analysis, we detected chemokine ligand 11 (CCL11) overexpression in CAFs compared to NFs. CCL11 administration promoted the migration and invasion of head and neck cancer (HNC) cells with enhanced cancer stem cell-like properties and induction of epithelial-to-mesenchymal transition. Furthermore, neutralization of CCL11 activity reversed the aggressive phenotype of CAF-induced cancer cells. Confocal microscopy showed colocalization of CCL11 and CC chemokine receptor 3 (CCR3) on HNC cells. Moreover, immunohistochemical analysis of clinical samples from 104 patients with HNC showed that expression of CCL11 and CCR3 were significantly correlated with poor overall survival (p = 0.003 and 0.044, respectively). Collectively, CCL11 expressed on CAFs promotes HNC invasiveness, and neutralization of CCL11 reverses this effect. We propose that the CCL11/CCR3 signaling circuit is a potential target for optimizing therapeutic strategies against HNC. [ABSTRACT FROM AUTHOR]

Background Transient receptor potential (TRP) family plays important roles in cardiovascular system. We investigated the relationship between transient receptor potential channel subfamily M6 (TRPM6) and atrial fibrosis in rheumatic heart disease patients with atrial fibrillation (AF). Methods The right atrial tissue samples were obtained from 64 patients with rheumatic heart diseases who underwent heart valve replacement surgery, and composed of 34 sinus rhythm (SR) patients and 30 AF patients. Hematoxylin and Eosin (HE) staining was used to observe cross-sectional area (CSA) of myocardial cell. Masson staining and measurement of connective tissue growth factor (CTGF), transforming growth factor beta 1 (TGF-β 1), and collagen type I/III (Collagen I/III) were performed to determine atrial fibrosis. The mRNA and protein levels of TRPM6 were detected by real-time quantitative polymerase chain reaction (RT-PCR) and western blotting, respectively. Results Marked increases were observed in CSA of myocardial cell and myocardial collagen volume fraction in AF group compared with the SR group (all P < 0.05). The mRNA levels of myocardial fibrosis markers (CTGF, TGF-beta 1, Collagen I/III) in AF group increased significantly compared to the SR group (all P < 0.05). TRPM6 mRNA and protein levels in AF group were elevated markedly in comparison with SR group ( P < 0.01). Conclusion These findings revealed that increased TRPM6 mRNA and protein levels may contribute to atrial fibrosis, and suggested that TRPM6 might be involved in AF development by promoting fibrogenesis. [ABSTRACT FROM AUTHOR]

The present study aims to investigate the expression and function of lysine-specific demethylase 4B (KDM4B) in yak cumulus cells (CCs) in order to reveal the mechanisms by which KDM4B regulates biological characteristics and function of CCs. The cellular location of KDM4B and the methylation pattern of H3K9 were detected using immunofluorescence (IF) staining in CCs. The mRNA expression levels of apoptosis-related genes (BCL-2 , HAX1 and BAX) and genes related to the estrogen pathway (ESR2 , CYP17 and 3B-HSD) were estimated by qRT-PCR after knockdown of KDM4B expression by siRNA in yak CCs. Then, a proliferation assay, Annexin V-FITC staining, and ELISA were utilized to explore the effects of KDM4B silencing on CCs proliferation, apoptosis, and estrogen (E 2) secretion, respectively. The results showed that KDM4B is located in the nuclei of yak CCs and is distributed in a dotted pattern. Knockdown KDM4B induced a decrease in cell proliferation, an increase in apoptotic rate and a reduction in the levels of E 2 secretion of CCs. Additionally, the methylation patterns of H3K9me2 and H3K9me3 were significantly increased in CCs transfected with KDM4B siRNA-1 (P < 0.05). The mRNA expression level of apoptosis promoting BAX genes was significantly upregulated, but 3B-HSD, ESR2 and anti-apoptotic HAX1 genes were significantly downregulated in transfected CCs (P < 0.05). Furthermore, the rate of embryos developing from the 2-cell stage to blastocysts was lower in the siRNA-1 transfection group than that of the control group (28.6 ± 2.9% vs 40.4 ± 2.4%, P < 0.05). In conclusion, our study indicates that KDM4B regulates the biological characteristics and physiological function of yak CCs mainly through changing the methylation patterns of H3K9 and related gene expression levels. • Cumulus granulosa cells of yak were successfully isolated and cultured. • Knockdown KDM4B could induce a decrease in the levels of E2 secretion and cell proliferation, and an increase in apoptotic rate of CCs. • The embryonic developmental competence was downregulated significantly After CCs transfection with siRNA-1. [ABSTRACT FROM AUTHOR]

Aims To explore risk factors for stroke independent of hypertension and the relationship between riboflavin kinase ( RFK) and stroke. Methods Gene expression profiling in the brains of spontaneously hypertensive rats ( SHR) and stroke-prone spontaneously hypertensive rats ( SHRSP) was comparatively analyzed by gene chips. The differentially expressed gene RFK was further verified by q- PCR and Western blot. The protective role of RFK-regulated flavins (including riboflavin, flavin mononucleotide, and flavin adenine dinucleotide) in stroke was observed in middle cerebral artery occlusion ( MCAO) mice. Influence of flavins on apoptosis and death in oxygen and glucose deprivation ( OGD)-treated neurons was examined by flow cytometry. Bax and Bcl-2 proteins were detected by Western blot. Results Of the 76 differentially expressed genes, 41 genes were upregulated, and 35 genes were downregulated in SHRSP as compared with SHR. RFK was significantly downregulated in SHRSP. Flavins markedly decreased infarct area in MCAO mice, inhibited apoptosis and death in OGD-treated neurons, and decreased Bax protein expression. Conclusions Physiological downregulation of RFK may be a new potential risk factor for stroke, which probably affects the absorbance and utility of riboflavin and further destroys the protective effect of flavins on stroke. RFK might act as a therapeutic target for stroke. [ABSTRACT FROM AUTHOR]

Transposons are widely used for genetic engineering in various model organisms. Recently, piggyBac (PB) has been developed as a transposable and efficient gene transfer tool in mammalian cells. In the present study, we developed three types of PB transposon systems containing a dual plasmid system (DPS), a single plasmid system (SPS), and a DNA-mRNA combined system (DRPS) and characterized their basic properties in HEK293 cells. The basic elements of the donor plasmid included a selectable-reporter gene expression cassette, two loxP sites in the same orientation, a multiple cloning site, and two chicken β-globin insulator core elements. We further identified the function of the selectable-reporter and examined PB integration sites in the human genome. Moreover, we compared the transposition efficacy and found that SPS transposed more efficiently, as compared to DPS; integration into the host genome was determined by measuring PBase activity. Results discovered the loss of PBase activity in the DRPS, indicating that this system is much more biologically safe, as compared to DPS and SPS. Finally, we employed the DRPS to successfully perform a gene delivery into bovine mammary epithelial cells (BMECs). Taken together, the information from this study will improve the flexibility of PB transposon systems and reduce the genotoxicity of PBase in genetic engineering. [ABSTRACT FROM AUTHOR]

Simple Summary: The tumor microenvironment (TME) plays an important role of cancer recurrence and treatment resistance. The cytokines in the TME may involve the tumor pathogenesis. Therefore, it is critical to discover the novel TME-associated signal transduction pathways contributing for the tumor progression. In this study, we established stable clones of interleukin (IL)-33-overexpressing head and neck squamous cell carcinoma (HNSCC) cells to simulate IL-33-induced autocrine signaling and identify the role of IL-33 expression in more aggressive phenotypes with increased mobility and properties of cancer stemness to provide a potential therapeutic strategy for HNSCC patients, on the basis of targeting the IL-33-enhanced-CXCR4 regulatory circuit. Despite recent advances, treatment for head and neck squamous cell carcinoma (HNSCC) has limited efficacy in preventing tumor progression. We confirmed previously that carcinoma-associated fibroblasts (CAF)-induced interleukin-33 (IL-33) contributed to cancer progression. However, the molecular mechanisms underlying the complex communication network of the tumor microenvironment merited further evaluation. To simulate the IL-33-induced autocrine signaling, stable clones of IL-33-overexpressing HNSCC cells were established. Besides well-established IL-33/ST2 and SDF1/CXCR4 (stromal-derived factor 1/C-X-C motif chemokine receptor 4) signaling, the CAF-induced IL-33 upregulated CXCR4 via cancer cell induction of IL-33 self-production. The IL-33-enhanced-CXCR4 regulatory circuit involves SDF1/CXCR4 signaling activation and modulates tumor behavior. An in vivo study confirmed the functional role of IL-33/CXCR4 in tumor initiation and metastasis. The CXCR4 and/or IL-33 blockade reduced HNSCC cell aggressiveness, with attenuated invasions and metastases. Immunohistochemistry confirmed that IL-33 and CXCR4 expression correlated significantly with disease-free survival and IL-33-CXCR4 co-expression predicted a poor outcome. Besides paracrine signaling, the CAF-induced IL-33 reciprocally enhanced the autocrine cancer-cell self-production of IL-33 and the corresponding CXCR4 upregulation, leading to the activation of SDF1/CXCR4 signaling subsequent to cancer progression. Thus, targeting the IL-33-enhanced-CXCR4 regulatory circuit attenuates tumor aggressiveness and provides a potential therapeutic option for improving the prognosis in HNSCC patients. [ABSTRACT FROM AUTHOR]

To investigate the effect of mitochondrial heteroplasmy on embryo development, cloned embryos produced using bovine oocytes as the recipient cytoplasm and ovine granulosa cells as the donor nuclei were complemented with 2 pL mitochondrial suspension isolated from ovine (BOOMT embryos) or bovine (BOBMT embryos) granulosa cells; cloned embryos without mitochondrial injection served as the control group (BO embryos). Reverse transcription–quantitative polymerase chain reaction (RT-qPCR) and sodium bisulfite genomic sequencing were used to analyse mRNA and methylation levels of pluripotency genes (OCT4, SOX2) and mitochondrial genes (TFAM, POLRMT) in the early developmental stages of cloned embryos. The number of mitochondrial DNA copies in 2 pL ovine-derived and bovine-derived mitochondrial suspensions was 960 ± 110 and 1000 ± 120, respectively. The blastocyst formation rates were similar in BOBMT and BO embryos (P > 0.05), but significantly higher than in BOOMT embryos (P < 0.01). Expression of OCT4 and SOX2, as detected by RT-qPCR, decreased significantly in BOOMT embryos (P < 0.05), whereas the expression of TFAM and POLRMT increased significantly, compared with expression in BOOMT and BO embryos (P < 0.05). In addition, methylation levels of OCT4 and SOX2 were significantly greater (P < 0.05), whereas those of TFAM and POLRMT were significantly lower (P < 0.01), in BOOMT embryos compared with BOBMT and BO embryos. Together, the results of the present study suggest that the degree of mitochondrial heteroplasmy may affect embryonic development. The mitochondria play crucial roles throughout embryonic development. To investigate whether mitochondrial heteroplasmy affects embryo development, cloned embryos with different levels of mitochondrial heteroplasmy were constructed. Blastocyst formation rates, the expression of pluripotency and mitochondrial regulator genes, and the methylation levels of these genes differed significantly with mitochondrial heteroplasmy. Therefore, mitochondrial heteroplasmy may affect embryonic development. [ABSTRACT FROM AUTHOR]