Fibrosis is a complex and multifactorial process, affecting the structure and compromising the function of several organs. Among those, renal fibrosis is an important pathological change, eventually leading to renal failure. Proteomic analysis of the renal parenchyma in the well-established rat model of unilateral ureteral obstruction (UUO model) suggested that transgelin was up-regulated during the development of fibrosis. Transgelin up-regulation was confirmed both at the protein and at the mRNA level. It was observed that at early stages of fibrosis transgelin was mainly expressed in the interstitial compartment and, more specifically, in cells surrounding the glomeruli. Subsequently, it was confirmed that transgelin expressing cells were activated fibroblasts, based on their extensive co-expression of α-SMA and their complete lack of co-distribution with markers of other cell types (endothelial, epithelial and cells of the immune system). These periglomerular fibroblasts exhibited staining for transgelin mainly cytoplasmic but occasionally nuclear as well. In addition, transgelin expression in periglomerular fibroblasts was absent in renal fibrosis developed in a hypertensive model, compared to the UUO model. Promoter analysis indicated that there are several conserved motifs for transcription factor binding. Among those, Kruppel-like factor 6 was found to be up-regulated in transgelin positive periglomerular activated fibroblasts, suggesting a possible involvement in the mechanism of transgelin up-regulation. These data strongly suggest that transgelin is up-regulated in the obstructive nephropathy and could be used as a novel marker for renal fibrosis in the future.

In this study, expression of the SPC25 gene was characterized in breast cancer (BC), and its effects on BC development and progression, functions in BC cells, and potential underlying mechanisms were examined. Data from TCGAportal and FIREBROWSE indicated that SPC25 was upregulated in BC tissues compared to normal tissues, and CANCERTOOL indicated that higher SPC25 mRNA levels were associated with increased probability of recurrence and poorer survival in BC patients. BC patients with higher SPC25 expression displayed shorter distant metastasis-free survival, relapse-free survival, and overall survival. Colony formation and CCK-8 experiments confirmed that SPC25 promoted proliferation of BC cells. Single-cell analysis indicated that SPC25 is associated with cell cycle regulation, DNA damage and repair, and BC cell proliferation. SPC25 knockdown suppressed proliferation of BC cells. MiRNAs, circRNAs, RNA-binding proteins, transcription factors, and immune factors that might interact with SPC25 mRNA to promote BC were also identified. These findings suggest that SPC25 levels are higher in more malignant BC subtypes and are associated with poor prognosis in BC patients. In addition, DNA methyltransferase inhibitor and transcription factors inhibitor treatments targeting SPC25 might improve survival in BC patients.

Neuronal nicotinic acetylcholine receptors (nAChR) composed of alpha4 + beta2 subunits, the high affinity nicotine-binding site in the mammalian brain, up-regulate in response to chronic nicotine exposure. The identities of endogenous mediators of this process are unknown. We find that choline also up-regulates alpha4 + beta2 nAChRs stably expressed by HEK293 cells as measured by increased [(3)H]epibatidine density. Choline-mediated up-regulation is dose-dependent and corresponds with an increase in beta2 subunit protein expression. The choline kinase inhibitor hemicholinium-3 inhibits approximately 60% of choline-mediated up-regulation revealing both an HC3-dependent and -independent pathway. Furthermore, choline-mediated up-regulation is not additive with up-regulation agents such as nicotine, but it is additive with weaker promoters of the up-regulation process. When co-applied with the pro-inflammatory cytokine tumor necrosis factor alpha, choline-mediated up-regulation is increased further through a mechanism that includes an increase in both alpha4 and beta2 protein expression, and this is inhibited by the p38 MAPK inhibitor SB202190. These findings extend the view that up-regulation of alpha4 + beta2 nAChRs is a normal physiological response to altered metabolic and inflammatory conditions.

Celiac disease (CD) is an autoimmune enteropathy linked to alterations of metabolism. Currently, limited untargeted metabolomic studies evaluating differences in the plasma metabolome of CD subjects have been documented. We engage in a metabolomic study that analyzes plasma metabolome in 17 children with CD treated with a gluten-free diet and 17 healthy control siblings in order to recognize potential changes in metabolic networks. Our data demonstrates the persistence of metabolic defects in CD subjects in spite of the dietary treatment, affecting a minor but significant fraction (around 4%, 209 out of 4893 molecular features) of the analyzed plasma metabolome. The affected molecular species are mainly, but not exclusively, lipid species with a particular affectation of steroids and derivatives (indicating an adrenal gland affectation), glycerophospholipids (to highlight phosphatidic acid), glycerolipids (with a special affectation of diacylglycerols), and fatty acyls (eicosanoids). Our findings are suggestive of an activation of the diacylglycerol-phosphatidic acid signaling pathway in CD that may potentially have detrimental effects via activation of several targets including protein kinases such as mTOR, which could be the basis of the morbidity and mortality connected with untreated CD. However, more studies are necessary to validate this idea regarding CD.

The transposon theory of aging hypothesizes the activation of transposable elements (TEs) in somatic tissues with age, leading to a shortening of the lifespan. It is thought that TE activation in aging produces an increase in DNA double-strand breaks, contributing to genome instability and promoting the activation of inflammatory responses. To investigate how TE regulation changes in somatic tissues during aging, we analyzed the expression of some TEs, as well as a source of small RNAs that specifically silence the analyzed TEs; the Drosophila cluster named flamenco. We found significant variations in the expression levels of all the analyzed TEs during aging, with a trend toward reduction in middle-aged adults and reactivation in older individuals that suggests dynamic regulation during the lifespan.

Human ether-a-go-go related-gene K⁺ channels (hERG) participate in the regulation of tumor cell proliferation and apoptosis. HERG channel activity is up-regulated by growth factors. Kinases sensitive to growth factor signaling include the serine/threonine protein kinase B-RAF. The present study thus explored whether B-RAF influences hERG channel expression and activity. To this end, hERG channels were expressed in Xenopus oocytes with or without wild-type B-RAF, hERG channel activity was determined utilizing dual-electrode voltage clamp and hERG protein abundance in the cell membrane was analyzed utilizing confocal microscopy as well as chemiluminescence. Moreover, in rhabdomyosarcoma RD cells the effect of B-RAF inhibitor PLX-4720 on hERG-mediated current was quantified by whole-cell patch clamp and hERG cell surface protein abundance by utilizing biotinylation of cell surface proteins as well as flow cytometry. As a result, co-expression of wild-type B-RAF in hERG-expressing Xenopus oocytes significantly increased hERG channel activity and hERG channel protein abundance in the cell membrane. Treatment for 24 hours of B-RAF and hERG-expressing Xenopus oocytes with B-RAF inhibitor PLX-4720 (10 µM) significantly decreased hERG-mediated current and hERG cell surface expression. Similarly, in rhabdomyosarcoma RD cells, treatment for 24 hours with B-RAF inhibitor PLX-4720 significantly decreased hERG cell membrane protein abundance and hERG-mediated current. In conclusion, B-RAF is a powerful regulator of hERG channel activity and cell surface hERG protein abundance.

Essential hypertension (EH) is a high prevalence disease facing a public health challenge. People were little known about the genetics of diagnosing the cause of EH. Circular RNAs that have a continuous cycle of covalent closure, without affected by RNA exonuclease, and are more stable and hard to degrade may involve into the molecule regulation mechanism of EH as an important biomedical.

During microRNA (miRNA) biogenesis, one strand of a 21-23 nucleotide RNA duplex is preferentially selected for entry into an RNA-induced silencing complex (RISC). The other strand, known as the miRNA* species, is typically thought to be degraded. Previous studies have provided miRNA* selection models, but it remains unclear how the dominance of one arm arises during the biogenesis of miRNA. Using miRNA sponge-like methods, we cloned four tandem target sequences (artificial target) of miR-7b* and then measured miR-7b* expression levels after transfection of the artificial target. miR-7b* levels were found to significantly increase after transfection of the artificial target. We postulate that the abundance of target transcripts drives miRNA arm selection.

Introduction: Melatonin, an endogenous neurohormone, modulates the biological circadian rhythms of vertebrates. It functions have been reported in previous stomatological studies as anti-inflammation, antioxidant, osseointegration of dental implants and stimulation to dental pulp stem cells differentiation, but its role in ameloblastic differentiation and mineralization has been rarely studied. Objective: To reveal the effects of melatonin on the mineralization of ameloblast lineage cells (ALCs), and to identify the change in gene expression and the potential mechanism based on ribonucleic acid sequencing (RNA-seq) analysis. Method: ALCs were induced in melatonin-conditioned medium. After 7-days culture, Western blot, real-time PCR, alkaline phosphatase (ALP) activity test, RNA-seq were accordingly used to detect the change in molecular level. After 1-month odontogenic induction in melatonin medium, Alizarin Red-S (ARS) staining showed the changes of mineral nodules. Differentially expressed genes (DEGs), enrichment of functions and signaling pathways analysis based on Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) database were performed. The JNK3 antagonist (JNK3 inhibitor IX, SR3576) and β-arrestin1 (Arrb1) overexpression were applied to confirm the fluctuation of melatonin-medicated JNK3 and Arrb1 expression. Results: In this study, we found out melatonin contributed to the ameloblastic mineralization, from which we can observed the elevated expression of enamel matrix protein, and increased ALP activity and mineralized nodules formation. RNA-seq analysis showed the up-regulation of neural JNK3 and down-regulation of Arrb1 in ALCs. Meanwhile, phosphorylated JNK3 deficiency (phosphorylated JNK3 inhibitor---SR3576 added to culture medium) led to mineralization delay, and Arrb1 overexpression proved Arrb1 takes bridge between melatonin receptors (MTNR) and JNK3 in MAPK signaling pathway.

Osteoprotegerin (OPG)/osteoclastogenesis inhibitory factor regulates bone mass by inhibiting osteoclastic bone resorption. mTOR, which is the mammalian target of rapamycin, is a kinase and central regulator of cell growth, proliferation, and survival. By using Rapamycin, we studied whether mTOR pathway is associated with OPG protein production in the mouse bone marrow-derived stromal cell line ST2. Rapamycin markedly increased the level of soluble OPG in ST2 cells. This antibiotic treatment resulted in the suppression of phosphorylation of mTOR. Rapamycin had no effects on the proliferation, differentiation, or apoptosis of the cells. Treatment with bone morphogenetic protein-4, which can induce OPG protein in ST2 cells, also resulted in a decrease in the density of the phospho-mTOR-band, suggesting that the suppression of the phospho-mTOR pathway is necessary for OPG production in ST2 cells. Thus, suitable suppression of mTOR phosphorylation is a necessary requirement for OPG production in bone marrow stromal cells.

Epithelial-to-mesenchymal transition (EMT) facilitates tumor metastasis. Twist is a basic helix-loop-helix protein that modulates many target genes through E-box-responsive elements. There are two twist-like proteins, Twist-1 and Twist-2, sharing high structural homology in mammals. Twist-1 was found to be a key factor in the promotion of metastasis of cancer cells, and is known to induce EMT. Twist-1 participation in carcinoma progression and metastasis has been reported in a variety of tumors. However, controversy exists concerning the correlation between Twist-1 and prognostic value with respect to carcinoma. A systematic review and meta-analysis were performed to determine whether the expression of Twist-1 was associated with the prognosis of carcinoma patients. This analysis included 17 studies: four studies evaluated lung cancer, three evaluated head and neck cancer, two evaluated breast cancer, two evaluated esophageal cancer, two evaluated liver cancer and one each evaluated osteosarcoma, bladder, cervical and ovarian cancer. A total of 2006 patients were enrolled in these studies, and the median trial sample size was 118 patients. Twist-1 expression was associated with worse overall survival (OS) at both 3 years (hazard ratio "HR" for death = 2.13, 95% CI = 1.86 to 2.45, p < 0.001) and 5 years (HR for death = 2.01, 95% CI = 1.76 to 2.29, p < 0.001). Expression of Twist-1 is associated with worse survival in carcinoma.

CacyBP/SIP (calcyclin binding protein/Siah-1 interacting protein) was originally discovered in Ehrlich ascities tumor cells but was later found also in many different tumors.

Cytosolic nonspecific dipetidase (CN2) belongs to the family of M20 metallopeptidases. It was stated in previous articles that higher expression levels of CN2 were observed in renal cell carcinoma and breast cancer. Our study explored the correlation between CN2 and colon carcinogenesis.

A deficiency of the dysferlin protein results in limb girdle muscular dystrophy type 2B and Miyoshi myopathy, with resulting plasma membrane abnormalities in myofibers. Many patients show muscle inflammation, but the molecular mechanisms that initiate and perpetuate this inflammation are not well understood. We previously showed abnormal activation of macrophages and hypothesized that activation of the inflammasome pathway may play a role in disease progression. To test this, we studied the inflammasome molecular platform in dysferlin-deficient human and mouse muscle. Consistent with our model, components of the NACHT, LRR and PYD-containing proteins (NALP)-3 inflammasome pathway were specifically up-regulated and activated in dysferlin-deficient but not in dystrophin-deficient and normal muscle. We demonstrate for the first time that normal primary skeletal muscle cells are capable of secreting IL-1beta in response to combined treatment with lipopolysaccharide and the P2X7 receptor agonist, benzylated ATP, suggesting that not only immune cells but also muscle cells can actively participate in inflammasome formation. In addition, we show that dysferlin-deficient primary muscle cells express toll-like receptors (TLRs; TLR-2 and TLR-4) and can efficiently produce IL-1beta in response to lipopolysaccharide and benzylated ATP. These data indicate that skeletal muscle is an active contributor of IL-1beta and strategies that interfere with this pathway may be therapeutically useful for patients with limb girdle muscular dystrophy type 2B.

Cellular senescence is a driver of various aging-associated disorders, including osteoarthritis. Here, we identified a critical role for Yes-associated protein (YAP), a major effector of Hippo signaling, in maintaining a younger state of human mesenchymal stem cells (hMSCs) and ameliorating osteoarthritis in mice. Clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR associated protein 9 nuclease (Cas9)-mediated knockout (KO) of YAP in hMSCs resulted in premature cellular senescence. Mechanistically, YAP cooperated with TEA domain transcriptional factor (TEAD) to activate the expression of forkhead box D1 (FOXD1), a geroprotective protein. YAP deficiency led to the down-regulation of FOXD1. In turn, overexpression of YAP or FOXD1 rejuvenated aged hMSCs. Moreover, intra-articular administration of lentiviral vector encoding YAP or FOXD1 attenuated the development of osteoarthritis in mice. Collectively, our findings reveal YAP-FOXD1, a novel aging-associated regulatory axis, as a potential target for gene therapy to alleviate osteoarthritis.

Osteoporosis and bone disorders related to the metabolic syndrome are often associated with adipokines secreted by adipocytes in bone. Adiponectin, a type of adipokine, is a regulator of immune responses and metabolic processes, but its role in bone biology remains uncertain. We investigated the role of adiponectin in bone metabolism using adiponectin-deficient mice in vivo and in vitro. Adiponectin-deficient mice exhibited reduced bone mass and increased adiposity. Adiponectin-deficient calvarial cells were prone to differentiate into adipocytes rather than osteoblasts. Although bone marrow macrophages (BMMs) from adiponectin-deficient mice had low osteoclastogenic potential as osteoclast precursors with increasing interferon regulatory factor 5 expression, under co-culture conditions of calvarial cells and BMMs, the enhanced receptor activator of nuclear factor κB ligand/osteoprotegerin (RANKL/OPG) ratio of adiponectin-deficient mesenchymal progenitor cells facilitated osteoclast differentiation. In addition, increased RANKL/OPG ratio was observed in the bone marrow extracellular fluid of adiponectin-deficient mice compared to that of wild-type mice. Notably, recombinant adiponectin treatment enhanced RANKL-induced osteoclast differentiation from BMMs but up-regulated OPG production in recombinant adiponectin-exposed calvarial cells, which inhibited osteoclast differentiation. Taken together, these results suggest that adiponectin plays an inhibitory role in bone metabolism through cross talk between precursor cells of both osteoclasts and osteoblasts by regulating RANKL/OPG ratio in the bone marrow microenvironment.

Abnormal expression of Aurora-A and epidermal growth factor receptor (EGFR) is observed in different kinds of cancer and associated with poor prognosis in cancer patients. However, the relationship between Aurora-A and EGFR in tumour development was not clear. In previous reports, we found that EGFR translocates to nucleus to activate Aurora-A expression after EGF treatment in EGFR-overexpressed cells. However, we also observed that not all the EGFR-overexpressed cells have the nuclear EGFR pathway to mediate the Aurora-A expression. In this study, we demonstrated that EGF signalling increased the Aurora-A protein expression in EGFR-overexpressed colorectal cancer cell lines via increasing the translational efficiency. In addition, the overexpression of EGFR was also associated with higher expression of Aurora-A in clinical colorectal samples. Activation of the PI3K/Akt/mTOR and MEK/ERK pathways mediated the effect of EGF-induced translational up-regulation. Besides, only the splicing variants containing exon 2 of Aurora-A mRNA showed increased interaction with the translational complex to synthesize Aurora-A protein under EGF stimulus. Besides, the exon 2 containing splicing variants were the major Aurora-A splicing forms expressed in human colorectal cancers. Taken together, our results propose a novel regulatory mechanism for the abnormal expression of Aurora-A in EGFR-overexpressed cancers, and highlight the importance of alternative 5'-UTR splicing variants in regulating Aurora-A expression. Furthermore, the specific expression of exon 2 containing splicing variants in cancer tissues may serve as a potential target for cancer therapy in the future.

Methamphetamine (METH) is a widespread illicit drug. If it is taken by pregnant women, it passes through the placenta and just as it affects the mother, it can impair the development of the offspring. The aim of our study was to identify candidates to investigate for changes in the gene expression in the specific regions of the brain associated with addiction to METH in rats. We examined the various areas of the central nervous system (striatum, hippocampus, prefrontal cortex) for signs of impairment in postnatal day 80 in experimental rats, whose mothers had been administered METH (5 mg/kg/day) during the entire gestation period. Changes in the gene expression at the mRNA level were determined by two techniques, microarray and real-time PCR. Results of two microarray trials were evaluated by LIMMA analysis. The first microarray trial detected either up-regulated or down-regulated expression of 2189 genes in the striatum; the second microarray trial detected either up-regulated or down-regulated expression of 1344 genes in the hippocampus of prenatally METH-exposed rats. We examined the expression of 10 genes using the real-time PCR technique. Differences in the gene expression were counted by the Mann-Whitney U-test. Significant changes were observed in the cocaine- and amphetamine-regulated transcript prepropeptide, tachykinin receptor 3, dopamine receptor D3 gene expression in the striatum regions, in the glucocorticoid nuclear receptor Nr3c1 gene expression in the prefrontal cortex and in the carboxylesterase 2 gene expression in the hippocampus of prenatally METH-exposed rats. The microarray technique also detected up-regulated expression of trace amine-associated receptor 7 h gene in the hippocampus of prenatally METH-exposed rats. We have identified susceptible genes; candidates for the study of an impairment related to methamphetamine addiction in the specific regions of the brain.

Hepatocellular carcinoma (HCC) is a common cause of cancer mortality worldwide. Recent studies have shown that the polytopic enzyme membrane associated ring-CH-type finger 6 (MARCH6) participates in tumorigenesis, but its function in HCC development needs to be investigated. This study aimed to explore the role of MARCH6 in HCC.

Although huge progress has been made in therapeutics against hepatocellular carcinoma (HCC) over the decades, the prognosis of this lethal disease remains poor. To find out risk factors for HCC-related outcome and better predict the prognosis, there is an unmet need to identify novel biomarkers of HCC. Accumulating evidence suggests that circRNAs play pivotal roles in carcinogenesis of several malignancies. In this study, we analyzed two datasets (GSE 94508 and GSE 97332) to examine differentially expressed circRNAs markedly related to HCC pathogenesis. Using Limma package in R and WGCNA analysis, hsa_circ_0000517 was significantly up-regulated in HCC (adjusted P < 0.01). Thereafter, a hsa_circ_0000517-related regulatory network was built based on application of databases including CSCD, TargetScan, miRDB, and miRTarBase. We uncovered the potential function of hsa_circ_0000517 through bioinformatics approaches, such as PPI network, GO, and KEGG pathway analyses. Specifically, functional analysis unveiled that hsa_circ_0000517 was likely to regulate the MAPK and Ras pathway through sponging several miRNAs and having an impact on the expression of TP53, MYC, and AKT1. To verify our initial finding, the expression of hsa_circ_0000517 in 60 HCC patients was detected by qRT-PCR and the expression in cancer tissues was higher compared with the paracarcinoma tissues. Survival analysis suggests high hsa_circ_0000517 expression was associated with adverse prognosis in HCC patients. Furthermore, this circRNA was significantly up-regulated in worse TNM stage, consistent with the progressive-stage-specific characteristic of circRNAs. A prognostic nomogram built on AFP and has_circ_0000517 showed significant diagnostic value. In all, we concluded that hsa_circ_0000517, a promising molecular in underlying mechanism of HCC, is a potent valuable biomarker for prognosis prediction.