[D]-H6L9, as a pH-responsive anti-microbial peptide (AMP), has been evidenced by us to be an excellent choice in tumor microenvironment-responsive delivery as it could render liposomes responsive to the acidified tumor microenvironment. However, [D]-H6L9-modified liposomes could not actively target to tumor area. Therefore, integrin αvβ3-targeted peptide RGD was co-modified with [D]-H6L9 onto liposomes [(R + D)-Lip] for improved tumor delivery efficiency. Under pH 6.3, (R + D)-Lip could be taken up by C26 cells and C26 tumor spheroids (integrin αvβ3-positive) with significantly improved efficiency compared with other groups, which was contributed by both RGD and [D]-H6L9, while RGD did not increase the cellular uptake performance on MCF-7 cells (integrin αvβ3-negative). Results showed that RGD could decrease cellular uptake of (R + D)-Lip while [D]-H6L9 could increase it, implying the role of both RGD and [D]-H6L9 in cellular internalization of (R + D)-Lip. On the other hand, (R + D)-Lip could escape the entrapment of lysosomes. PTX-loaded (R + D)-Lip could further increase the cellular toxicity against C26 cells compared with liposomes modified only with RGD and [D]-H6L9 respectively, and achieve remarkable tumor inhibition effect on C26 tumor models.

Prenatal ethanol exposure (PEE) is an established risk factor for intrauterine growth retardation. The present study was designed to determine whether PEE can increase the susceptibility of high-fat diet (HFD)-induced metabolic syndrome (MS) in adult offspring in a sex-specific manner, based on a generalized linear model analysis. Pregnant Wistar rats were administered ethanol (4 g/kg.d) from gestational day 11 until term delivery. All offspring were fed either a normal diet or a HFD after weaning and were sacrificed at postnatal week 20, and blood samples were collected. Results showed that PEE reduced serum adrenocorticotropic hormone (ACTH) and corticosterone levels but enhanced serum glucose, insulin, insulin resistant index (IRI), triglyceride and total cholesterol (TC) concentrations. Moreover, the analysis showed interactions among PEE, HFD and sex. In the PEE offspring, HFD aggravated the decrease in ACTH and corticosterone levels and further increased serum glucose, insulin, triglyceride and TC levels. The changes of serum ACTH, glucose and IRI levels in the female HFD rats were greater than those in the male HFD rats. Our findings suggest that PEE enhances the susceptibility to MS induced by HFD in a sex-specific manner, which might be primarily associated with the neuroendocrine metabolic programming by PEE.

Inhibition of the Wnt antagonist sclerostin increases bone mass in patients with osteoporosis and in preclinical animal models. Here we show increased levels of the Wnt antagonist Dickkopf-1 (DKK-1) in animals treated with sclerostin antibody, suggesting a negative feedback mechanism that limits Wnt-driven bone formation. To test our hypothesis that co-inhibition of both factors further increases bone mass, we engineer a first-in-class bispecific antibody with single residue pair mutations in the Fab region to promote efficient and stable cognate light-heavy chain pairing. We demonstrate that dual inhibition of sclerostin and DKK-1 leads to synergistic bone formation in rodents and non-human primates. Furthermore, by targeting distinct facets of fracture healing, the bispecific antibody shows superior bone repair activity compared with monotherapies. This work supports the potential of this agent both for treatment and prevention of fractures and offers a promising therapeutic approach to reduce the burden of low bone mass disorders.

Cell penetrating peptides (CPPs) were widely used for drug delivery to tumor. However, the nonselective in vivo penetration greatly limited the application of CPPs-mediated drug delivery systems. And the treatment of malignant tumors is usually followed by poor prognosis and relapse due to the existence of extravascular core regions of tumor. Thus it is important to endue selective targeting and stronger intratumoral diffusion abilities to CPPs. In this study, an RGD reverse sequence dGR was conjugated to a CPP octa-arginine to form a CendR (R/KXXR/K) motif contained tandem peptide R8-dGR (RRRRRRRRdGR) which could bind to both integrin αvβ3 and neuropilin-1 receptors. The dual receptor recognizing peptide R8-dGR displayed increased cellular uptake and efficient penetration ability into glioma spheroids in vitro. The following in vivo studies indicated the active targeting and intratumoral diffusion capabilities of R8-dGR modified liposomes. When paclitaxel was loaded in the liposomes, PTX-R8-dGR-Lip induced the strongest anti-proliferation effect on both tumor cells and cancer stem cells, and inhibited the formation of vasculogenic mimicry channels in vitro. Finally, the R8-dGR liposomal drug delivery system prolonged the medium survival time of intracranial C6 bearing mice by 2.1-fold compared to the untreated group, and achieved an exhaustive anti-glioma therapy including anti-tumor cells, anti-vasculogenic mimicry and anti-brain cancer stem cells. To sum up, all the results demonstrated that R8-dGR was an ideal dual receptor recognizing CPP with selective glioma targeting and efficient intratumoral diffusion, which could be further used to equip drug delivery system for effective glioma therapy.

Cystic change in metastatic lymph nodes of papillary thyroid carcinoma (PTC) is a diagnostic challenge for fine needle aspiration (FNA) because of the scant cellularity. The aim of this study was to evaluate the measurement of thyroglobulin in fine needle aspirate (Tg-FNA) for detecting metastatic PTC in patients with cystic neck lesions and to validate the optimal cutoff value of Tg-FNA. A total of 75 FNA specimens of cystic lesions were identified, including 40 of metastatic PTC. Predetermined threshold levels of 0.04 (minimum detection level), 0.9, 10.0, and 77.0 ng/mL (maximum normal serum-Tg level) were used to evaluate the diagnostic accuracy of Tg-FNA for metastatic PTC detection. The areas under the receiver operating characteristic curve for diagnosing metastatic PTC of Tg-FNA values of 0.04, 0.9, 10.0, and 77.0 ng/mL were 0.5 (95% confidence interval [CI], 0.382-0.618), 0.645 (95% CI, 0.526-0.752), 0.945 (95% CI, 0.866-0.984), and 0.973 (95% CI, 0.907-0.996), respectively. With a cutoff value of 77.0 ng/mL, the combination of Tg-FNA and FNA cytology showed superior diagnostic power (97.5% sensitivity and 100% specificity) compared to FNA cytology alone (80% sensitivity and 100% specificity). We recommend a Tg-FNA cutoff of 77.0 ng/mL, the maximum normal serum-Tg level, for cystic neck lesions.

The carboxyl terminus of Hsp70-interacting protein (CHIP) is a U box-type ubiquitin ligase that plays crucial roles in various biological processes, including tumor progression. To date, the functional mechanism of CHIP in thyroid cancer remains unknown. Here, we obtained evidence of upregulation of CHIP in thyroid cancer tissues and cell lines. CHIP overexpression markedly enhanced thyroid cancer cell viability and colony formation in vitro and accelerated tumor growth in vivo. Conversely, CHIP knockdown impaired cell proliferation and tumor growth. Notably, CHIP promoted cell growth through activation of MAPK and AKT pathways, subsequently decreasing p27 and increasing cyclin D1 and p-FOXO3a expression. Our findings collectively indicate that CHIP functions as an oncogene in thyroid cancer, and is therefore a potential therapeutic target for this disease.

Our previous study showed that knockdown the endogenous expression of DCUN1D3 (also called SCCRO3 or DCNL3) blocked the S phase progression after UV irradiation. Here, we show that the silence of DCUN1D3 can increase the cyclin-dependent kinase inhibitor p27 protein levels after UV irradiation. Through Co-immunoprecipitation experiments, we found that DCUN1D3 bound to CAND1. And DCUN1D3 knockdown synergized with CAND1 over-expression in arresting the S phase. Given the CAND1's established role in Cullin-1 neddylation, we found Cullin-1 was less neddylated in DCUN1D3 deficient cells. So the silence of DCUN1D3 can inhibit the formation of SCFSKP2 complex by reducing Cullin-1 neddylation. Given that p27 is the primary target of SCFSKP2 complex, the cells lost DCUN1D3 showed a remarkable accumulation of p27 to cause S phase block.

The purpose of this study was to investigate the diagnostic value of 3.0-T (1)H magnetic resonance spectroscopy ((1)H MRS) in primary malignant hepatic tumors and to compare the effects of (1)H MRS on the diagnostic accuracy of liver-occupying lesions between junior and experienced radiologists.

Peroxisome proliferator-activated receptor α (PPARα) is a key regulator to ameliorate liver injury in cases of acute liver failure (ALF). However, its regulatory mechanisms remain largely undetermined. Endoplasmic reticulum stress (ER stress) plays an important role in a number of liver diseases. This study aimed to investigate whether PPARα activation inhibits ER stress-induced hepatocyte apoptosis, thereby protecting against ALF. In a murine model of D-galactosamine (D-GalN)- and lipopolysaccharide (LPS)-induced ALF, Wy-14643 was administered to activate PPARα, and 4-phenylbutyric acid (4-PBA) was administered to attenuate ER stress. PPARα activation ameliorated liver injury, because pre-administration of its specific inducer, Wy-14643, reduced the serum aminotransferase levels and preserved liver architecture compared with that of controls. The protective effect of PPARα activation resulted from the suppression of ER stress-induced hepatocyte apoptosis. Indeed, (1) PPARα activation decreased the expression of glucose-regulated protein 78 (Grp78), Grp94 and C/EBP-homologous protein (CHOP) in vivo; (2) the liver protection by 4-PBA resulted from the induction of PPARα expression, as 4-PBA pre-treatment promoted upregulation of PPARα, and inhibition of PPARα by small interfering RNA (siRNA) treatment reversed liver protection and increased hepatocyte apoptosis; (3) in vitro PPARα activation by Wy-14643 decreased hepatocyte apoptosis induced by severe ER stress, and PPARα inhibition by siRNA treatment decreased the hepatocyte survival induced by mild ER stress. Here, we demonstrate that PPARα activation contributes to liver protection and decreases hepatocyte apoptosis in ALF, particularly through regulating ER stress. Therefore, targeting PPARα could be a potential therapeutic strategy to ameliorate ALF.

RBM15, an RNA binding protein, determines cell-fate specification of many tissues including blood. We demonstrate that RBM15 is methylated by protein arginine methyltransferase 1 (PRMT1) at residue R578, leading to its degradation via ubiquitylation by an E3 ligase (CNOT4). Overexpression of PRMT1 in acute megakaryocytic leukemia cell lines blocks megakaryocyte terminal differentiation by downregulation of RBM15 protein level. Restoring RBM15 protein level rescues megakaryocyte terminal differentiation blocked by PRMT1 overexpression. At the molecular level, RBM15 binds to pre-messenger RNA intronic regions of genes important for megakaryopoiesis such as GATA1, RUNX1, TAL1 and c-MPL. Furthermore, preferential binding of RBM15 to specific intronic regions recruits the splicing factor SF3B1 to the same sites for alternative splicing. Therefore, PRMT1 regulates alternative RNA splicing via reducing RBM15 protein concentration. Targeting PRMT1 may be a curative therapy to restore megakaryocyte differentiation for acute megakaryocytic leukemia.

Hyperactivation of the transcriptional factor E2F1 occurs frequently in human cancers and contributes to malignant progression. E2F1 activity is regulated by proteolysis mediated by the ubiquitin-proteasome system. However, the deubiquitylase that controls E2F1 ubiquitylation and stability remains undefined. Here we demonstrate that the deubiquitylase POH1 stabilizes E2F1 protein through binding to and deubiquitylating E2F1. Conditional knockout of Poh1 alleles results in reduced E2F1 expression in primary mouse liver cells. The POH1-mediated regulation of E2F1 expression strengthens E2F1-downstream prosurvival signals, including upregulation of Survivin and FOXM1 protein levels, and efficiently facilitates tumour growth of liver cancer cells in nude mice. Importantly, human hepatocellular carcinomas (HCCs) recapitulate POH1 regulation of E2F1 expression, as nuclear abundance of POH1 is increased in HCCs and correlates with E2F1 overexpression and tumour growth. Thus, our study suggests that the hyperactivated POH1-E2F1 regulation may contribute to the development of liver cancer.

Abscisic acid (ABA) regulates various developmental processes and stress responses in plants. Protein phosphorylation/dephosphorylation is a central post-translational modification (PTM) in ABA signaling. However, the phosphoproteins regulated by ABA under osmotic stress remain unknown in maize. In this study, maize mutant vp5 (deficient in ABA biosynthesis) and wild-type Vp5 were used to identify leaf phosphoproteins regulated by ABA under osmotic stress. Up to 4052 phosphopeptides, corresponding to 3017 phosphoproteins, were identified by Multiplex run iTRAQ-based quantitative proteomic and LC-MS/MS methods. The 4052 phosphopeptides contained 5723 non-redundant phosphosites; 512 phosphopeptides (379 in Vp5, 133 in vp5) displayed at least a 1.5-fold change of phosphorylation level under osmotic stress, of which 40 shared common in both genotypes and were differentially regulated by ABA. Comparing the signaling pathways involved in vp5 response to osmotic stress and those that in Vp5, indicated that ABA played a vital role in regulating these pathways related to mRNA synthesis, protein synthesis and photosynthesis. Our results provide a comprehensive dataset of phosphopeptides and phosphorylation sites regulated by ABA in maize adaptation to osmotic stress. This will be helpful to elucidate the ABA-mediate mechanism of maize endurance to drought by triggering phosphorylation or dephosphorylation cascades.

Dendritic spines undergo continuous remodeling during development of the nervous system. Their stability is essential for maintaining a functional neuronal circuit. Spine dynamics and stability of cortical excitatory pyramidal neurons have been explored extensively in mammalian animal models. However, little is known about spiny interneurons in non-mammalian vertebrate models. In the present study, neuronal morphology was visualized by single-cell electroporation. Spiny neurons were surveyed in the Xenopus tadpole brain and observed to be widely distributed in the olfactory bulb and telencephalon. DsRed- or PSD95-GFP-expressing spiny interneurons in the olfactory bulb were selected for in vivo time-lapse imaging. Dendritic protrusions were classified as filopodia, thin, stubby, or mushroom spines based on morphology. Dendritic spines on the interneurons were highly dynamic, especially the filopodia and thin spines. The stubby and mushroom spines were relatively more stable, although their stability significantly decreased with longer observation intervals. The 4 spine types exhibited diverse preferences during morphological transitions from one spine type to others. Sensory deprivation induced by severing the olfactory nerve to block the input of mitral/tufted cells had no significant effects on interneuron spine stability. Hence, a new model was established in Xenopus laevis tadpoles to explore dendritic spine dynamics in vivo.

Protein turnover and homeostasis are regulated by the proteasomal system, which is critical for cell function and viability. Pluripotency of stem cells also relies on normal proteasomal activity that mitigates senescent phenotypes induced by intensive cell replications, as previously demonstrated in human bone marrow stromal cells. In this study, we investigated the role of proteasomes in self-renewal of neural progenitor cells (NPCs). Through both in vivo and in vitro analyses, we found that the expression of proteasomes was progressively decreased during aging. Likewise, proliferation and self-renewal of NPCs were also impaired in aged mice, suggesting that the down-regulation of proteasomes might be responsible for this senescent phenotype. Lowering proteasomal activity by loss-of-function manipulations mimicked the senescence of NPCs both in vitro and in vivo; conversely, enhancing proteasomal activity restored and improved self-renewal in aged NPCs. These results collectively indicate that proteasomes work as a key regulator in promoting self-renewal of NPCs. This potentially provides a promising therapeutic target for age-dependent neurodegenerative diseases.

Opioid drug abusers have a greater susceptibility to gram positive (Gram (+)) bacterial infections. However, the mechanism underlying opioid modulation of Gram (+) versus Gram (-) bacterial clearance has not been investigated. In this study, we show that opioid treatment resulted in reduced phagocytosis of Gram (+), when compared to Gram (-) bacteria. We further established that LPS priming of chronic morphine treated macrophages leads to potentiated phagocytosis and killing of both Gram (+) and Gram (-) bacteria in a P-38 MAP kinase dependent signaling pathway. In contrast, LTA priming lead to inhibition of both phagocytosis and bacterial killing. This study demonstrates for the first time the differential effects of TLR4 and TLR2 agonists on morphine induced inhibition of phagocytosis. Our results suggest that the incidence and severity of secondary infections with Gram (+) bacteria would be higher in opioid abusers.

Cell-penetrating peptides (CPPs) have been widely used to enhance the membrane translocation of various carriers for many years, but the non-specificity of CPPs seriously limits their utility in vivo. In this study, cholesterol-anchored, reduction-sensitive PEG (first synthesized by our laboratory) was applied to develop a co-modified liposome with improved tumor targeting. Following optimization of the formulation, the in vitro and in vivo properties of the co-modified liposome were evaluated. The co-modified liposome had a much lower cellular uptake and tumor spheroid uptake, but a much higher tumor accumulation compared to CPP-modified liposome, indicating the non-specific penetration of CPPs could be attenuated by the outer PEG coating. With the addition of exogenous reducing agent, both the in vitro and in vivo cellular uptake was markedly increased, demonstrating that the reduction-sensitive PEG coating achieved a controllable detachment from the surface of liposomes and did not affect the penetrating abilities of CPPs. The present results demonstrate that the combination of cholestervsitive PEG and CPPs is an ideal alternative for the application of CPP-modified carriers in vivo.

An increasing number of studies have highlighted the potential link between EXO1 polymorphisms and cancer risk, although no consensus has yet been obtained. Thus, we aimed to obtain a thorough and current assessment of EXO1 polymorphisms and cancer susceptibility by performing a meta-analysis. A comprehensive literature retrieval was performed on PubMed, EMbase, Web of Science and Wanfang databases. The odds ratio (OR) and 95% confidence interval (CI) were applied to assess the results. Finally, 39 case-control studies of the nine EXO1 polymorphisms that involved 21,651 cases and 21,348 controls met our inclusion criteria. The pooled analysis indicated that the rs1047840 polymorphism conferred a significantly increased susceptibility to cancer in an allelic model. Similarly, the rs3754093, rs1776177, rs9350, rs10802996, rs1635498, rs1776148 and rs851797 polymorphisms were also associated with an increased susceptibility to cancer in an allelic model, respectively, while no significant association was identified for rs1635517 polymorphism. For the rs1047840 polymorphism, in an ethnicity subgroup analysis, a significantly increased susceptibility to cancer for Asians was identified in all the genetic models, and for Caucasians in an allelic model. Our findings provide the evidence that the rs1047840, rs9350, rs10802996, rs1635498, rs1776148, rs1776177, rs3754093 and rs851797 polymorphisms may act as risk factors for cancer.

Podocyte apoptosis is a major mechanism that leads to proteinuria in many kidney diseases. However, the concert mechanisms that cause podocyte apoptosis in these kidney diseases are not fully understood. RhoA is one of Rho GTPases that has been well studied and plays a key role in regulating cytoskeletal architecture. Previous study showed that insufficient RhoA could result in rat aortic smooth muscle cell apoptosis. However, whether RhoA is involved in podocyte apoptosis remains unknown.

The role of immunity in the pathogenesis of acute hepatitis B virus (HBV) infection is poorly understood. The purpose of this research was to define the intrahepatic immune factors responsible for viral clearance during acute HBV infection. The model of acute HBV infection was established by hydrodynamically transfecting mice with pCDNA3.1-HBV1.3 plasmids which contained a supergenomic HBV1.3-length transgene. The frequency of CD4+ CXCR5+ T cells, CD19+ B cells and their surface molecules in livers, spleens and peripheral blood were detected using flow cytometry. The lymphomononuclear cells isolated from the livers of transfected mice were further stimulated by HBc-derived peptides and then the frequency and cytokine secretion of HBV-specific CD4+CXCR5+ T cells were detected. We found that the frequency of CXCR5+ in CD4+ T cells was specifically increased; the expression of PD-1 was decreased while the expression of ICOS was increased on intrahepatic CD4+CXCR5+ T cells. Although the frequency of CD19+ B cells was not affected, the expression of PDL-1, ICOSL and IL-21R on B cells was increased in the livers of mice. The frequency of HBV-specific CD4+CXCR5+ T cells and the production of IL-21 by intrahepatic CD4+CXCR5+ T cells of mice with acute HBV infection were increased after stimulation. Furthermore, the expression of function-related molecules of intrahepatic CD4+CXCR5+ T, including Bcl-6, CXCR5, IL-6, IL-6R, IL-21 and IL-4 in the liver was increased during acute HBV infection. In conclusion, the activation of intrahepatic CD4+CXCR5+ T cells and B cells was associated with the clearance of HBV during acute infection.

The prominent feature of human cytomegalovirus (HCMV) is cell tropism specificity for human fetal nervous system, which leads to severe fetal nervous system damage especially in first-trimester gestation. In this study, human astrocytes isolated from fetal brain were infected with HCMV AD169 and whole genome transcriptome profile was performed. The results showed that the gene expression of interferon stimulated genes (ISGs), chemokine and chemokine receptors were significantly up-regulated (P < 0.01). The antiviral replication effects of IFIT1 (Interferon-induced protein with tetratricopeptide repeats 1, Fc = 148.17) was investigated. Lentivirus with IFIT1 overexpression or knockdown was transduced into astrocytes, respectively. The viral mRNA, protein expression and HCMV titers were determined. The results showed that IE1, IE2, pp65, and viral titers were significantly decreased in IFIT1 overexpression group and enhanced in the knockdown group compared with control one (P < 0.01). Taken together, this study revealed IFIT1 played an important antiviral role in HCMV infected fetal astrocytes. The prominent feature of human cytomegalovirus (HCMV) is cellular tropism specificity for human fetal brain nervous system leading to severe fetal nervous damage especially in first-trimester gestation. In this study, human astrocytes isolated from first-trimester fetal brain were infected with HCMV AD169 and IFIT1 was studied for its antiviral replication effects. The results provided insights into the function of IFIT1 as a key factor in antiviral defense contributing to development of targeted therapeutics to fetal brain with HCMV infection. J. Med. Virol. 89:672-684, 2017. © 2016 Wiley Periodicals, Inc.