Osteogenesis associated with persistent inflammation or infection exists in a broad range of conditions including rheumatoid arthritis and traumatic bone fracture. The poor outcomes of these conditions will benefit from more effective treatments. Here we investigated the molecular mechanisms and tested NEMO-binding domain peptide as a new approach of circumventing TNF-alpha inhibition of osteoblast differentiation. Our results showed: TNF-alpha markedly decreased BMP-2-induced alkaline phosphatase activity in the multipotent myoblast C2C12 cells in a dose dependent manner; stepwise experiments demonstrated that BMP-2-induced Smad1 activity was abrogated by addition of exogenous TNF-alpha or overexpression of NF-kappaB, and it was significantly elevated by overexpression of IkappaBalpha, an inhibitor of NF-kappaB; Western blotting showed that TNF-alpha markedly decreased the amount of phospho-Smad1 in BMP-2-activated C2C12 cells, but it did not alter Smad1 mRNA abundance as measured by real-time PCR; addition of a functional cell-permeable NEMO-binding domain (NBD) peptide antagonized NF-kappaB activity and ameliorated TNF-alpha inhibition of osteoblast differentiation. Taken together, our study reveals for the first time that NF-kappaB activation inhibits osteoblast differentiation by attenuating Smad1 activity and application of NBD peptide ameliorates this inhibitory effect. This could lead to new therapeutic drugs that circumvent the inflammatory inhibition of osteogenesis for treatment of traumatic open fractures with infection, rheumatoid arthritis and other bone loss disorders.

Circadian patterns of cardiovascular vulnerability were well characterized, with a peak incidence of acute myocardial infarction and stroke secondary to atherosclerosis in the morning, which showed the circadian clock may take part in the pathological process of atherosclerosis induced by hyperlipidemia. Hence, the effect of hyperlipidemia on the expression of circadian genes was investigated in atherosclerotic mouse model.

Chronic kidney disease (CKD) is a major healthcare problem with increasing prevalence in the population. CKD leads to end stage renal disease and increases the risk of cardiovascular disease. As such, it is important to study the mechanisms underlying CKD progression. To this end, an animal model was developed to allow the testing of new treatment strategies or molecular targets for CKD prevention. Many underlying risk factors result in CKD but the disease itself has common features, including renal interstitial fibrosis, tubular epithelial cell loss through apoptosis, glomerular damage, and renal inflammation. Further, CKD shows differences in prevalence between the genders with premenopausal women being relatively resistant to CKD. We sought to develop and characterize an animal model with these common features of human CKD in the C57BL/6 mouse. Mice of this genetic background have been used to produce transgenic strains that are commercially available. Thus, a CKD model in this strain would allow the testing of the effects of numerous genes on the severity or progression of CKD with minimal cost. This paper describes such a mouse model of CKD utilizing angiotensin II and deoxycorticosterone acetate as inducers.

Colorectal carcinoma (CRC) is one of the most prevalent types of malignancy‑associated mortality worldwide. Previous studies have demonstrated that amplification and overexpression of the human circadian locomotor output cycles kaput gene (hClock) was closely associated with a high risk for CRC as well as poor prognosis in CRC patients. However, the underlying molecular mechanisms of CRC remain to be fully elucidated. In the present study, hClock was exogenously overexpressed in the CRC cell line SW480 via infection of a lentivirus vector expressing hClock; in addition, a lentivirus vector‑based RNA interference approach, using short hairpin RNA, was performed in order to knockdown hClock in SW620 cells. The results showed that upregulation of hClock promoted proliferation and inhibited apoptosis in SW480 cells in vitro and in vivo, while downregulation of hClock inhibited SW620 cell proliferation and accelerated apoptosis in vitro. Upregulation of hClock enhanced the activity of the anti‑apoptotic gene phosphorpylated (p‑)AKT and inhibited the expression of the pro‑apoptotic gene B cell lymphoma‑2 (Bcl‑2)‑associated X protein and Bcl‑2 homology 3 interacting domain death agonist. Furthermore, targeted inhibition of hClock activity reduced p‑AKT expression. In conclusion, the results of the present study suggested that the circadian gene hClock promoted CRC progression and inhibit tumor cell apoptosis in vitro and in vivo, while silencing hClock was able to reverse this effect.

The expression levels of miR-365 vary in different malignancies. Herein, we found that miR-365 was overexpressed in both cells and clinical specimens of cutaneous squamous cell carcinoma (SCC). We demonstrated that the HaCaT(pre-miR-365-2) cell line, which overexpressed miR-365, could induce subcutaneous tumors in vivo. Antagomir-365, an anti-miR-365 oligonucleotide, inhibited cutaneous tumor formation in vivo, along with G1 phase arrest and apoptosis of cancer cells. These findings suggest that miR-365 may act as an onco-miR in cutaneous SCC both in vitro and in vivo. The present study provides valuable insight into the role of miR-365 in cutaneous SCC formation, which can help develop new drug and miR-365 target-based therapies for cutaneous SCC.

Quantitative information on gene activity at single cell-type resolution is essential for the understanding of how cells work and interact. Root hairs, or trichoblasts, tubular-shaped outgrowths of specialized cells in the epidermis, represent an ideal model for cell fate acquisition and differentiation in plants.

Social caste determination in the honey bee is assumed to be determined by the dietary status of the young larvae and translated into physiological and epigenetic changes through nutrient-sensing pathways. We have employed Illumina/Solexa sequencing to examine the small RNA content in the bee larval food, and show that worker jelly is enriched in miRNA complexity and abundance relative to royal jelly. The miRNA levels in worker jelly were 7-215 fold higher than in royal jelly, and both jellies showed dynamic changes in miRNA content during the 4(th) to 6(th) day of larval development. Adding specific miRNAs to royal jelly elicited significant changes in queen larval mRNA expression and morphological characters of the emerging adult queen bee. We propose that miRNAs in the nurse bee secretions constitute an additional element in the regulatory control of caste determination in the honey bee.

Epithelial-to-mesenchymal transition (EMT) is an important event during tumorigenesis. The human high-mobility group A2 (HMGA2) is a chromatin-binding protein, which contains three AT-hook domains that enable its binding to the minor groove of DNA. HMGA2 organizes protein complexes on enhancers of various genes to regulate gene expression and cell differentiation. The HMGA2 protein has been reported to be overexpressed in many types of cancer. It is not known, however, whether HMGA2 regulates EMT in human hepatocellular carcinoma (HCC) cell lines, and the mechanism(s) have not been fully elucidated. In this study, the expression of HMGA2 in five HCC cell lines was examined. The levels of HMGA2 expression among the five HCC cell lines coincided with their invasiveness. The variation in HMGA2 expression significantly correlated with the expression of several putative EMT markers. In addition, assessment of the invasive potential, following transfection with HMGA2-siRNA, demonstrated that the rate of cell migration was significantly reduced, suggesting that HMGA2 may be an important contributor to the invasion of tumor cells and that expression levels of HMGA2 influence the metastatic behavior of HCC cells. To further confirm the conclusion and explore the molecular mechanism through which HMGA2 induces EMT, we found that HMGA2 upregulates the expression of Twist and Snail in HCC cell lines. In conclusion, this present study is the first to show that HMGA2 effectively regulates EMT to induce invasion and metastasis in HCC cells. The function of HMGA2 as an oncoprotein may be associated with several important molecules involved in invasion and metastasis of cancer cells.

High-frequency point mutations of genes encoding histones have been identified recently as novel drivers in a number of tumors. Specifically, the H3K36M/I mutations were shown to be oncogenic in chondroblastomas and undifferentiated sarcomas by inhibiting H3K36 methyltransferases, including SETD2. Here we report the crystal structures of the SETD2 catalytic domain bound to H3K36M or H3K36I peptides with SAH (S-adenosylhomocysteine). In the complex structure, the catalytic domain adopts an open conformation, with the K36M/I peptide snuggly positioned in a newly formed substrate channel. Our structural and biochemical data reveal the molecular basis underying oncohistone recognition by and inhibition of SETD2.

The identification of rare inherited and de novo copy number variations (CNVs) in human subjects has proven a productive approach to highlight risk genes for autism spectrum disorder (ASD). A variety of microarrays are available to detect CNVs, including single-nucleotide polymorphism (SNP) arrays and comparative genomic hybridization (CGH) arrays. Here, we examine a cohort of 696 unrelated ASD cases using a high-resolution one-million feature CGH microarray, the majority of which were previously genotyped with SNP arrays. Our objective was to discover new CNVs in ASD cases that were not detected by SNP microarray analysis and to delineate novel ASD risk loci via combined analysis of CGH and SNP array data sets on the ASD cohort and CGH data on an additional 1000 control samples. Of the 615 ASD cases analyzed on both SNP and CGH arrays, we found that 13,572 of 21,346 (64%) of the CNVs were exclusively detected by the CGH array. Several of the CGH-specific CNVs are rare in population frequency and impact previously reported ASD genes (e.g., NRXN1, GRM8, DPYD), as well as novel ASD candidate genes (e.g., CIB2, DAPP1, SAE1), and all were inherited except for a de novo CNV in the GPHN gene. A functional enrichment test of gene-sets in ASD cases over controls revealed nucleotide metabolism as a potential novel pathway involved in ASD, which includes several candidate genes for follow-up (e.g., DPYD, UPB1, UPP1, TYMP). Finally, this extensively phenotyped and genotyped ASD clinical cohort serves as an invaluable resource for the next step of genome sequencing for complete genetic variation detection.

Many studies have established that microRNAs (miRNAs) regulate gene expression in various biological processes in mammals and insects including honey bees. Dancing behavior is a form of communication unique to honey bees. However, it remains unclear which miRNAs regulate the dancing behavior in honey bees, and how. In the present study, total small RNAs (sRNAs) in Apis mellifera foragers and dancers were extracted and analyzed by a Solexa Sequencer to determine differentially expressed miRNAs. A small percentage (12.62%) of the unique sRNAs (the number of sequence types) were shared between foragers and dancers, but their expression accounted for 92.92% of the total sRNAs (the number of all sequence reads), and the length of them centered around 22nt. Out of 58 previously identified miRNAs, 54 were present in both foragers and dancers and most of them were down-regulated in dancers. The fold-changes of ame-miR-34, ame-miR-210, ame-miR-278 and ame-miR-282 were higher than 2. 86 and 104 novel miRNAs were detected in foragers and dancers, respectively. Furthermore, two known miRNAs (ame-miR-278 and ame-miR-282) were confirmed, by qPCR, to have lower expressions in dancers. The target genes of ame-miR-278 and ame-miR-282 were associated with kinase, neural function, synaptotagmin and energy. These results indicate that miRNAs are substantially different between the foraging and dancing stages, and suggest that miRNAs might play important roles in regulating dancing behaviors in honey bees.

In the detection of seizure onset zones, arterial spin labeling (ASL) can overcome the limitations of positron emission tomography (PET) with 18F-fluorodeoxyglucose (18F-FDG), which is invasive, expensive, and radioactive. PET/magnetic resonance (MR) systems have been introduced that allow simultaneous performance of ASL and PET, but comparisons of these techniques with stereoelectroencephalography (SEEG) and comparisons among the treatment outcomes of these techniques are still lacking. Here, we investigate the effectiveness of ASL compared with that of SEEG and their outcomes in localizing mesial temporal lobe epilepsy (MTLE) and assess the correlation between simultaneously acquired PET and ASL.

The aim of this study was to perform a systemic review of the studies addressing the use of moderately and severely steatotic donor livers for liver transplantation.

Here we present a visual reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detecting the gene encoding the σB major outer-capsid protein of novel duck reovirus (NDRV). A set of primers, composed of two outer primers, two inner primers and two loop primers, was designed based on the gene of interest. The LAMP reaction was conducted in a traditional laboratory water bath at 65 °C for 50 min. We compared the performance of calcein/Mn2+ and SYBR Green I dyes, as well as electrophoresis on agarose gel stained with GoldView nucleic acid dye to detect the RT-LAMP-amplified products and all assays could be employed to discriminate between positive and negative specimens in visible or UV light. Our data showed that there is no cross-reaction with other viruses and the RT-LAMP technique displayed high sensitivity for detecting NDRV with a minimal detection limit of 200 fg RNA input. This assay was more sensitive than conventional PCR in detecting NDRV both in natural and experimental infection. In conclusion, the RT-LAMP technique was remarkably sensitive, specific, rapid, simple and profitable for the identification of NDRV.

Successful RNAi applications depend on strategies allowing stable and persistent expression of minimal gene silencing triggers without perturbing endogenous gene expression. In this study, we proposed an endogenous microRNA (miRNA) cluster as a novel integration site for small hairpin RNAs (shRNAs). We successfully integrated exogenous shRNAs at the porcine miRNA-17-92 (pmiR-17-92) cluster via a CRISPR/Cas9-mediated knock-in strategy. The anti-EGFP or anti-CSFV shRNAs could be stably and effectively expressed at the control of the endogenous promoter of the pmiR-17-92 cluster. Importantly, we confirmed that hitchhike expression of anti- classical swine fever (CSFV) shRNA had no effect on cell growth, blastocyst development and endogenous pmiR-17-92 expression in selected transgene (TG) porcine fetal fibroblasts (PFFs) clones. Moreover, these TG PFFs could inhibit the replication of CSFV by half and could be further used for generation of transgenic pigs. Taken together, these results show that our RNA interference (RNAi) expression strategy benefits numerous applications, from miRNA, genome and transgenic research, to gene therapy.

The present study was aimed to evaluate resting-state functional connectivity and topological properties of brain networks in narcolepsy patients compared with healthy controls. Resting-state fMRI was performed in 26 adult narcolepsy patients and 30 matched healthy controls. MRI data were first analyzed by group independent component analysis, then a graph theoretical method was applied to evaluate the topological properties in the whole brain. Small-world network parameters and nodal topological properties were measured. Altered topological properties in brain areas between groups were selected as region-of-interest seeds, then the functional connectivity among these seeds was compared between groups. Partial correlation analysis was performed to evaluate the relationship between the severity of sleepiness and functional connectivity or topological properties in the narcolepsy patients. Twenty-one independent components out of 48 were obtained. Compared with healthy controls, the narcolepsy patients exhibited significantly decreased functional connectivity within the executive and salience networks, along with increased functional connectivity in the bilateral frontal lobes within the executive network. There were no differences in small-world network properties between patients and controls. The altered brain areas in nodal topological properties between groups were mainly in the inferior frontal cortex, basal ganglia, anterior cingulate, sensory cortex, supplementary motor cortex, and visual cortex. In the partial correlation analysis, nodal topological properties in the putamen, anterior cingulate, and sensory cortex as well as functional connectivity between these regions were correlated with the severity of sleepiness (sleep latency, REM sleep latency, and Epworth sleepiness score) among narcolepsy patients. Altered connectivity within the executive and salience networks was found in narcolepsy patients. Functional connection changes between the left frontal cortex and left caudate nucleus may be one of the parameters describing the severity of narcolepsy. Changes in the nodal topological properties in the left putamen and left posterior cingulate, changes in functional connectivity between the left supplementary motor area and right occipital as well as in functional connectivity between the left anterior cingulate gyrus and bilateral postcentral gyrus can be considered as a specific indicator for evaluating the severity of narcolepsy.

Nanotechnology-based drug delivery systems have been widely used for oral and systemic dosage forms delivery depending on the mucoadhesive interaction, and keratin has been applied for biomedical applications and drug delivery. However, few reports have focused on the keratin-based mucoadhesive drug delivery system and their mechanisms of mucoadhesion. Thus, the mucoadhesion controlled kerateine (reduced keratin, KTN)/keratose (oxidized keratin, KOS) composite nanoparticles were prepared via adjusting the proportion of KTN and KOS to achieve controlled gastric mucoadhesion and drug release based on their different mucoadhesive abilities and pH-sensitive properties. Furthermore, the mechanisms of mucoadhesion for KTN and KOS were also investigated in the present study.

Breast cancer (BC) is a common malignancy among women worldwide. Fibroblast growth factor receptor 2 (FGFR2) rs2981582 is reported to play a vital role in BC development. However, the relationship between them remains unclear.

Carminic acid (CA) extracted from cochineal is widely used in food additives as a natural colorant, and its potential risk to human health makes its detection important. In this work, a layered doubled hydroxide (LDH)-luminol-H2O2 system-based chemiluminescence (CL) platform has been successfully applied for CA sensing. The principle detection consists of two steps: first, LDH adsorbs CA onto the surface via electrostatic attraction; second, CA quenches the CL of the LDH-luminol-H2O2 system via the synergistic effect of CL resonance energy transfer, reduction of reactive oxygen species, and occupation of positively charged centers of brucite-like layers. With this CL approach, 0.5 μM CA is detectable using a CL spectrometer, and the limit of detection is 0.03 μM. This CL system exhibited a linear response to CA in the concentration range from 0.5 to 10 μM. In addition, the practical application of the designed CL sensing system is evaluated with dried pork slice samples.

Rapid increase of antimicrobial resistance has become an urgent threat to global public health. In this research, since photothermal therapy is a potential antibacterial strategy, which is less likely to cause resistance, a metal-organic framework-based chemo-photothermal combinational system was constructed. Zeolitic imidazolate frameworks-8 (ZIF-8), a porous carrier with unique features such as high loading and pH-sensitive degradation, was synthesized, and then encapsulated photothermal agent indocyanine green (ICG). First, ICG with improved stability in ZIF-8 (ZIF-8-ICG) can effectively produce heat in response to NIR laser irradiation for precise, rapid, and efficient photothermal bacterial ablation. Meanwhile, Zn2+ ions released from ZIF-8 can inhibit bacterial growth by increasing the permeability of bacterial cell membrane and further strengthen photothermal therapy efficacy by reducing the heat resistance of bacteria. Study showed that bacteria suffered from significant changes in morphology after treatment with ZIF-8-ICG under laser irradiation. The combinational chemo-hyperthermia therapy of ZIF-8-ICG could thoroughly ablate murine subcutaneous abscess induced by methicillin-resistant Staphylococcus aureus (MRSA), exhibiting a nearly 100% bactericidal ratio. Both in vitro and in vivo safety evaluation confirmed that ZIF-8-ICG was low toxic. Overall, our researches demonstrated that ZIF-8-ICG has great potential to be served as an alternative to antibiotics in combating multidrug-resistant bacterial pathogens.