Activation Of Signaling Proteins Research Articles

Stoichiometry controls activity of phase-separated clusters of actin signaling proteins.

Biomolecular condensates concentrate macromolecules into foci without a surrounding membrane. Many condensates appear to form through multivalent interactions that drive liquid-liquid phase separation (LLPS). LLPS increases the specific activity of actin regulatory proteins toward actin assembly by the Arp2/3 complex. We show that this increase occurs because LLPS of the Nephrin-Nck-N-WASP signaling pathway on lipid bilayers increases membrane dwell time of N-WASP and Arp2/3 complex, consequently increasing actin assembly. Dwell time varies with relative stoichiometry of the signaling proteins in the phase-separated clusters, rendering N-WASP and Arp2/3 activity stoichiometry dependent. This mechanism of controlling protein activity is enabled by the stoichiometrically undefined nature of biomolecular condensates. Such regulation should be a general feature of signaling systems that assemble through multivalent interactions and drive nonequilibrium outputs.

Tiliroside is a new potential therapeutic drug for osteoporosis in mice.

Osteoporosis is a class of metabolic bone disease caused by complexed ramifications. Overactivation of osteoclasts due to a sudden decreased estrogen level plays a pivotal role for postmenopausal women suffering from osteoporosis. Therefore, inhibiting osteoclast formation and function has become a major direction for the treatment of osteoporosis. Tiliroside (Tle) is a salutary dietary glycosidic flavonoid extracted from Oriental Paperbush flower, which has been reported to have an anti-inflammation effect. However, whether Tle affects the osteoclastogenesis and bone resorption remains unknown. Herein, we demonstrate that Tle prevents bone loss in ovariectomy in mice and inhibits osteoclast differentiation and bone resorption stimulated by receptor activator of nuclear factor-κB ligand (RANKL) in vitro. Molecular mechanism studies reveal that Tle reduces RANKL-induced activation of mitogen-activated protein kinase and T-cell nuclear factor 1 pathways, and osteoclastogenesis-related marker gene expression, including cathepsin K (Ctsk), matrix metalloproteinase 9, tartrate-resistant acid phosphatase (Acp5), and Atp6v0d2. Our research indicates that Tle suppresses osteoclastogenesis and bone loss by downregulating the RANKL-mediated signaling protein activation and expression. In addition, Tle inhibits intracellular reactive oxygen species generation which is related to the formation of osteoclasts. Therefore, Tle might serve as a potential drug for osteolytic disease such as osteoporosis.

Detection and functional analysis of BMP2 gene mutation in patients with tooth agenesis

To screen for BMP2 mutation with functional impact in patients with congenital tooth agenesis and to make oral and skeletal phenotype record and functional analysis with in vitro experiments. We enrolled eighteen patients with congenital tooth agenesis. The medical and dental history was collected,and clinical and dental examinations including the X-ray examination of oral-facial and skeletal bone were performed for the phenotypic analysis. Blood samples were collected to extract DNA and whole exome sequencing was conducted. The genes involved in oral-facial development and congenital skeletal diseases were investigated for mutation screening. The mutations with functional impact were then investigated. In one patient, the BMP2 mutation with putative functional impact was selected for functional analysis. Wild type and mutant BMP2 plasmids with green fluorescent protein (GFP) tag were constructed and transfected into HEK293T cells. Subcellular protein distribution was observed under laser scanning confocal microscope. The activation of downstream SMAD1/5/9 phosphorylation by BMP2 was detected by Western blotting to investigate the functional impact and genetic pathogenicity. BMP2 mutation NM_001200.3:c.393A>T (p.Arg131Ser), rs140417301 was detected in one patient with congenital tooth agenesis, while for other genes involved in oral-facial development and congenital skeletal diseases, no functionally significant mutation was found. The proband's parents didn't carry this mutation. The father had normal dentition, while the mother lacked one premolar, and both the parents showed normal palate and maxilla. The patient also had maxillary hypoplasia in both sagittal and coronal planes, palatal dysmorphology, and malocclusion, and was diagonsed with osteopenia after the X-ray examnination of his skeletal bone. Functional analysis showed this mutation had normal subcelluar localization but reduced phosphorylation of SMAD1/5/9 (reduction by 32%, 22%, and 27% in three independent replicates). Taken together with family co-segregation, this mutaion was considered as "likely pathogenic". BMP2 mutation c.393A>T (p. Arg131Ser) affects bone morphogenetic protein signaling activity, and may affect the number of teeth, growth of maxilla and palate, and bone mineral density.

C-Src regulates Nrf2 activity through PKCδ after oxidant stimulus

Nrf2 is the main transcription factor involved in expression of cell defense enzymes, which is altered in several oxidant-related disorders. Cytosolic Nrf2 activation is modulated through phosphorylation by PKCδ, an enzyme controlled by Src tyrosine kinases. Of relevance, Src family members are involved in numerous cellular processes and regulated by hydrogen peroxide (H2O2). In this study we analysed the activation of cell survival-related signaling proteins, c-Src and Nrf2, and the influence of c-Src kinase on Nrf2 regulation after exposure to H2O2. Acute exposure of HT22 mouse hippocampal neural cells to H2O2 increased c-Src and Nrf2 phosphorylation/activation at Tyr416 and Ser40, respectively. Nrf2 phosphorylation at Ser40, its nuclear accumulation and transcriptional activity involving heme oxygenase-1 (HO-1) expression were dependent on c-Src kinase activation. Moreover, modulation of Nrf2 activity by c-Src occurred through PKCδ phosphorylation at Tyr311. We demonstrate, for the first time, c-Src-mediated regulation of Nrf2 transcriptional activity, via PKCδ activation, following an acute H2O2 stimulus. This work supports that the c-Src/PKCδ/Nrf2 pathway may constitute a novel signaling pathway stimulated by H2O2 and a potential target for the treatment of diseases involving redox deregulation.

Circuit Dysfunction in SOD1-ALS Model First Detected in Sensory Feedback Prior to Motor Neuron Degeneration Is Alleviated by BMP Signaling.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease for which the origin and underlying cellular defects are not fully understood. Although motor neuron degeneration is the signature feature of ALS, it is not clear whether motor neurons or other cells of the motor circuit are the site of disease initiation. To better understand the contribution of multiple cell types in ALS, we made use of a Drosophila Sod1G85R knock-in model, in which all cells harbor the disease allele. End-stage dSod1G85R animals of both sexes exhibit severe motor deficits with clear degeneration of motor neurons. Interestingly, earlier in dSod1G85R larvae, motor function is also compromised, but their motor neurons exhibit only subtle morphological and electrophysiological changes that are unlikely to cause the observed decrease in locomotion. We analyzed the intact motor circuit and identified a defect in sensory feedback that likely accounts for the altered motor activity of dSod1G85R We found cell-autonomous activation of bone morphogenetic protein signaling in proprioceptor sensory neurons which are critical for the relay of the contractile status of muscles back to the central nerve cord, completely rescues early-stage motor defects and partially rescue late-stage motor function to extend lifespan. Identification of a defect in sensory feedback as a potential initiating event in ALS motor dysfunction, coupled with the ability of modified proprioceptors to alleviate such motor deficits, underscores the critical role that nonmotor neurons play in disease progression and highlights their potential as a site to identify early-stage ALS biomarkers and for therapeutic intervention.SIGNIFICANCE STATEMENT At diagnosis, many cellular processes are already disrupted in the amyotrophic lateral sclerosis (ALS) patient. Identifying the initiating cellular events is critical for achieving an earlier diagnosis to slow or prevent disease progression. Our findings indicate that neurons relaying sensory information underlie early stage motor deficits in a Drosophila knock-in model of ALS that best replicates gene dosage in familial ALS (fALS). Importantly, studies on intact motor circuits revealed defects in sensory feedback before evidence of motor neuron degeneration. These findings strengthen our understanding of how neural circuit dysfunctions lead to neurodegeneration and, coupled with our demonstration that the activation of bone morphogenetic protein signaling in proprioceptors alleviates both early and late motor dysfunction, underscores the importance of considering nonmotor neurons as therapeutic targets.

Robustness and Information Transfer within IL-6-induced JAK/STAT Signalling

Cellular communication via intracellular signalling pathways is crucial. Expression and activation of signalling proteins is heterogenous between isogenic cells of the same cell-type. However, mechanisms evolved to enable sufficient communication and to ensure cellular functions. We use information theory to clarify mechanisms facilitating IL-6-induced JAK/STAT signalling despite cell-to-cell variability. We show that different mechanisms enabling robustness against variability complement each other. Early STAT3 activation is robust as long as cytokine concentrations are low. Robustness at high cytokine concentrations is ensured by high STAT3 expression or serine phosphorylation. Later the feedback-inhibitor SOCS3 increases robustness. Channel Capacity of JAK/STAT signalling is limited by cell-to-cell variability in STAT3 expression and is affected by the same mechanisms governing robustness. Increasing STAT3 amount increases Channel Capacity and robustness, whereas increasing STAT3 tyrosine phosphorylation reduces robustness but increases Channel Capacity. In summary, we elucidate mechanisms preventing dysregulated signalling by enabling reliable JAK/STAT signalling despite cell-to-cell heterogeneity.

In Vitro Analysis of Hedgehog Acyltransferase and Porcupine Fatty Acyltransferase Activities.

Hedgehog and Wnt proteins are modified by covalent attachment of the fatty acids palmitate and palmitoleate, respectively. These lipid modifications are essential for Hedgehog and Wnt protein signaling activities and are catalyzed by related, but distinct fatty acyltransferases: Hedgehog acyltransferase (Hedgehog) and Porcupine (Wnt). In this chapter, we provide detailed methods to directly monitor Hedgehog and Wnt protein fatty acylation in vitro. Palmitoylation of Sonic hedgehog (Shh), a representative Hedgehog family member, is assayed using purified Hedgehog acyltransferase (Hhat) or Hhat-enriched membranes, a recombinant 19kDa Shh protein or C-terminally biotinylated Shh 10-mer peptide, and 125I-iodopalmitoyl CoA as the donor fatty acyl CoA substrate. The radiolabeled reaction products are quantified by SDS-PAGE and phosphorimaging or by γ-counting. To assay Wnt acylation, the reaction consists of a biotinylated, double disulfide-bonded Wnt peptide containing the sequence surrounding the Wnt3a acylation site, [125I] iodo-cis-9-pentadecenoyl CoA, and Porcupine-enriched membranes. Radiolabeled, biotinylated Wnt3a peptide is captured on streptavidin coated beads and the reaction product is quantified by γ-counting.

Single-Cell Mass Cytometry Characterizes Phenotypic and Functional Heterogeneity in Acute Myeloid Leukemia at Diagnosis, in Remission and Relapse

Single-Cell Mass Cytometry Characterizes Phenotypic and Functional Heterogeneity in Acute Myeloid Leukemia at Diagnosis, in Remission and Relapse

Multiregion Quantification of Extracellular Signal-regulated Kinase Activity in Renal Cell Carcinoma

To personalize treatment for renal cell carcinoma (RCC), it would be ideal to confirm the activity of druggable protein pathways within individual tumors. We have developed a high-resolution nanoimmunoassay (NIA) to measure protein activity with high precision in scant specimens (eg, fine needle aspirates [FNAs]). Here, we used NIA to determine whether protein activation varied in different regions of RCC tumors. Since most RCC therapies target angiogenesis by inhibiting the vascular endothelial growth factor (VEGF) receptor, we quantified phosphorylation of extracellular signal-regulated kinase (ERK), a downstream effector of the VEGF signaling pathway. In 90 ex vivo FNA biopsies sampled from multiple regions of 38 primary clear cell RCC tumors, ERK phosphorylation differed among patients. In contrast, within individual patients, we found limited intratumoral heterogeneity of ERK phosphorylation. Our results suggest that measuring ERK in a single FNA may be representative of ERK activity in different regions of the same tumor. As diagnostic and therapeutic protein biomarkers are being sought, NIA measurements of protein signaling may increase the clinical utility of renal mass biopsy and allow for the application of precision oncology for patients with localized and advanced RCC. Patient summaryIn this report, we applied a new approach to measure the activity of extracellular signal-regulated kinase (ERK), a key cancer signaling protein, in different areas within kidney cancers. We found that ERK activity varied between patients, but that different regions within individual kidney tumors showed similar ERK activity. This suggests that a single biopsy of renal cell carcinoma may be sufficient to measure protein signaling activity to aid in precision oncology approaches.

β-carotene isolated from the marine red alga, Gracillaria sp. potently attenuates the growth of human hepatocellular carcinoma (HepG2) cells by modulating multiple molecular pathways

There remains a remarkable controversy on the anti-cancer stand of β-carotene specifically when high concentrations are used. This study aimed to examine the growth inhibitory effect of the lower concentrations of β-carotene in HepG2 cells. Beta-carotene purified from Gracillaria sp. inhibited viability of HepG2 cells in a concentration-dependent manner. This growth inhibitory effect was well correlated with increased apoptotic cell death. Apoptosis induction by β-carotene was associated with increased caspase-3 activity and reduced mitochondrial membrane potential. Western blot analysis showed a significant decrease in the expression of Bcl-2, PARP, and NF-kB protein, and an increase in the protein expression of Bax. Beta-carotene also blocked the activation of intracellular growth signaling proteins, Akt and ERK1/2. Further, it down-regulated the expression of endogenous antioxidant enzymes, SOD-2 and HO-1, and its transactivation factor, Nrf-2. The current study emphasizes the role of β-carotene as an effective inhibitor of the growth of hepatocarcinoma cell.

Canonical BMP signaling in tubular cells mediates recovery after acute kidney injury

Bone morphogenetic protein (BMP) signaling has been shown to modulate the development of renal fibrosis in animal models of kidney injury, but the downstream mediators are incompletely understood. In wild-type mice, canonical BMP signaling mediated by SMAD1/5/8 transcription factors was constitutively active in healthy renal tubules, transiently down-regulated after ischemia reperfusion injury (IRI), and reactivated during successful tubular regeneration. We then induced IRI in mice with a tubular-specific BMP receptor 1A (BMPR1A) deletion. These mice failed to reactivate SMAD1/5/8 signaling in the post-ischemic phase and developed renal fibrosis after injury. Using unbiased genomic analyses, we identified three genes encoding inhibitor of DNA-binding (ID) proteins (Id1, Id2, and Id4) as key targets of BMPR1A-SMAD1/5/8 signaling. BMPR1A-deficient mice failed to re-induce these targets following IRI. Instead, BMPR1A-deficiency resulted in activation of pro-fibrotic signaling proteins that are normally repressed by ID proteins, namely, p38 mitogen-activated protein kinase and cell cycle inhibitor p27. These data indicate that the post-ischemic activation of canonical BMP signaling acts endogenously to repress pro-fibrotic signaling in tubular cells and may help to prevent the progression of acute kidney injury to chronic kidney disease.

Structural Insight into H-NOX Gas Sensing and Cognate Signaling Protein Regulation.

Heme-nitric oxide/oxygen binding (H-NOX) proteins are a family of gas-binding hemoproteins that bind diatomic gas ligands such as nitric oxide (NO) and oxygen (O2 ). In bacteria, H-NOXs are often associated with signaling partners, including histidine kinases (HKs), diguanylate cyclases (DGCs) or methyl-accepting chemotaxis proteins (MCPs), either as a stand-alone protein or as a domain of a larger polypeptide. H-NOXs regulate the activity of cognate signaling proteins through ligand-induced conformational changes in the H-NOX domain and protein/protein interactions between the H-NOX and the cognate signaling partner. This review summarizes recent progress toward deciphering the molecular mechanism of bacterial H-NOX activation and the subsequent regulation of H-NOX-associated cognate sensor proteins from a structural and biochemical point of view.

An Anti-Cancer Drug Candidate CYC116 Suppresses Type I Hypersensitive Immune Responses through the Inhibition of Fyn Kinase in Mast Cells.

Mast cells are the most prominent effector cells of Type 1 hypersensitivity immune responses. CYC116 [4-(2-amino-4-methyl-1,3-thiazol-5-yl)-N-[4-(morpholin-4-yl)phenyl] pyrimidin-2-amine] is under development to be used as an anti-cancer drug, but the inhibitory effects of CYC116 on the activation of mast cells and related allergy diseases have not reported as of yet. In this study, we demonstrated, for the first time, that CYC116 inhibited the degranulation of mast cells by antigen stimulation (IC50, ∼1.42 µM). CYC116 also inhibited the secretion of pro-inflammatory cytokines including TNF-α (IC50, ∼1.10 µM), and IL-6 (IC50, ∼1.24 µM). CYC116 inhibited the mast cell-mediated allergic responses, passive cutaneous anaphylaxis (ED50, ∼22.5 mg/kg), and passive systemic anaphylaxis in a dose-dependent manner in laboratory experiments performed on mice. Specifically, CYC116 inhibited the activity of Fyn in mast cells and inhibited the activation of Syk and Syk-dependent signaling proteins including LAT, PLCγ, Akt, and MAP kinases. Our results suggest that CYC116 could be used as an alternative therapeutic medication for mast cell-mediated allergic disorders, such as atopic dermatitis and allergic rhinitis.

Profiling signaling proteins in human spermatozoa: Biomarker identification for sperm quality evaluation

In this study we unraveled human spermatozoa signaling pathways and correlated the activity of sperm signaling proteins with clinical data. Thirty seven semen samples were used and basic semen parameters, sperm DNA fragmentation and the expression levels of 75 protein kinases and the phosphorylation/cleavage patterns of 18 signaling proteins were evaluated to determine the correlation between semen basic parameters and the expression and activity of signaling proteins. We have contributed towards establishing a biomarker “fingerprint” to assess sperm quality based on molecular parameters.

FSP1-positive fibroblasts are adipogenic niche and regulate adipose homeostasis.

Adipocyte progenitors reside in the stromal vascular fraction (SVF) of adipose tissues that are composed of fibroblasts, immune cells, and endothelial cells. It remains to be elucidated how the SVF regulates adipocyte progenitor fate determination and adipose homeostasis. Here, we report that fibroblast-specific protein-1 (FSP1)+ fibroblasts in the SVF are essential to adipose homeostasis. FSP1+ fibroblasts, devoid of adipogenic potential, are adjacent to the preadipocytes in the SVF. Ablation of FSP1+ fibroblasts in mice severely diminishes fat content of adipose depots. Activation of canonical Wnt signaling in the FSP1+ fibroblasts results in gradual loss of adipose tissues and resistance to diet-induced obesity. Alterations in the FSP1+ fibroblasts reduce platelet-derived growth factor (PDGF)-BB signaling and result in the loss of preadipocytes. Reduced PDGF-BB signaling, meanwhile, impairs the adipogenic differentiation capability of preadipocytes by regulating matrix metalloproteinase (MMP) expression, extracellular matrix remodeling, and the activation of Yes-associated protein (YAP) signaling. Thus, FSP1+ fibroblasts are an important niche essential to the maintenance of the preadipocyte pool and its adipogenic potential in adipose homeostasis.

Constitutively Active MDA5 Proteins Are Inhibited by Paramyxovirus V Proteins

Excessive interferon (IFN) production and signaling can lead to immunological and developmental defects giving rise to autoimmune diseases referred to collectively as "type I interferonopathies." A subset of these diseases is caused by monogenic mutations affecting proteins involved in nucleic acid sensing, homeostasis, and metabolism. Interferonopathic mutations in the cytosolic antiviral sensor MDA5 render it constitutively hyperactive, resulting in chronic IFN production and IFN-stimulated gene expression. Few therapeutic options are available for patients with interferonopathic diseases, but a large number of IFN evasion and antagonism strategies have evolved in viral pathogens that can counteract IFN production and signaling to enhance virus replication. To test the hypothesis that these natural IFN suppressors could be used to subdue the activity of interferonopathic signaling proteins, hyperactive MDA5 variants were assessed for susceptibility to a family of viral MDA5 inhibitors. In this study, Paramyxovirus V proteins were tested for their ability to counteract constitutively active MDA5 proteins. Results indicate that the V proteins are able to bind to and disrupt the signaling activity of these MDA5 proteins, irrespective of their specific mutations, reducing IFN production and IFN-stimulated gene expression to effectively suppress the hyperactive antiviral response.

Abstract 4808: Combinations of imatinib mesylate with AKT inhibitor (miransertib, ARQ 751) or FGFR inhibitor (derazantinib) show synergy in GIST cell lines and preclinical models

Abstract The majority of gastrointestinal stromal tumors (GIST) harbor oncogenic mutations in the receptor tyrosine kinase KIT or platelet-derived growth factor receptor alpha (PDGFRA). Small molecule kinase inhibitors such as imatinib mesylate (IM) have significantly improved the clinical management of GIST by targeting these mutant receptors. Despite strong overall response rates to IM, disease progression generally occurs with time. Inhibiting targets other than, or in addition to, KIT/PDGFRA may provide additional therapeutic benefit in GIST patients. Both AKT and FGFR signaling have recently been reported to be resistance mechanisms associated with survival of IM-resistant GIST cell lines and tumors. In this study, we performed in vitro and in vivo experiments to assess the potential benefit of combining IM with the ArQule AKT inhibitors Miransertib (ARQ 092) and ARQ 751 and the FGFR inhibitor, Derazantinib (ARQ 087). To evaluate in vitro drug sensitivity, a panel of IM-sensitive (GIST-T1, GIST882) and resistant GIST cell lines (GIST-T1/829, GIST430) were subjected to drug treatment for 72 hours before measuring viability with the Cell Titer Blue Viability Assay. Synergy between IM and Miransertib, ARQ 751 and Derazantinib was quantified using the Chou-Talalay algorithm to calculate CI values. CI values <1 are considered synergistic. The 3:1 molar ratio of the three tested combinations demonstrated synergistic CI values in all four GIST lines. Immunoblot assays confirmed that drugs inhibited their intended targets in each cell line following six-hour drug treatment. Interestingly, a significant decrease in the activation of a downstream signaling protein, p-S6, was observed in the AKTi+ IM -treated cells compared to cells treated with single agents. In vivo studies evaluating Miransertib, ARQ 751 and Derazantinib as monotherapies and in combination with IM were performed using GIST-T1 and GIST430 xenograft models, as well, as an IM-resistant, KIT exon 9-mutated GIST PDX model. In both IM-resistant GIST models, all three monotherapies (Miransertib, ARQ 751, Derazantinib) significantly inhibited tumor growth as compared to IM alone. In addition, combination therapy, with both AKTi and FGFRi, provided significantly greater efficacy in both IM-sensitive and resistant xenograft models of GIST. Together, these studies demonstrate that IM in combination with the novel ArQule AKT inhibitors (Miransertib and ARQ 751) and FGFR inhibitor (Derazantinib) provide significantly improved efficacy compared to monotherapy in the tested models. These results provide justification for development of translational trials evaluating these combinations in GIST patients. Citation Format: Marya Kozinova, Shalina Joshi, Karthik Devarajan, Phillip Zook, Jimson W. D'Souza, Jeffrey M. Farma, Nestor Esnaola, Reza Foroughi, Yi Yu, Brian Schwartz, Terence Hall, Margaret von Mehren, Lori A. Rink. Combinations of imatinib mesylate with AKT inhibitor (miransertib, ARQ 751) or FGFR inhibitor (derazantinib) show synergy in GIST cell lines and preclinical models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4808.

Cortical and hippocampal expression of inflammatory and intracellular signaling proteins in aged rats submitted to aerobic and resistance physical training.

Aging is often accompanied by an increase in pro-inflammatory markers. This inflammatory process is directly related to cellular dysfunctions that induce events such as the exacerbated activation of cell death signaling pathways. In the aged brain, dysregulation of the normal activities of neuronal cells compromises brain functions, thereby favoring the onset of neurodegenerative diseases and cognitive deficits. Interactions between various stimuli, such as stress, are responsible for the modulation of cellular processes and activities. Physical exercise is a controllable model of stress, largely used as a strategy for studying the physiological mechanisms of inflammatory responses and their consequences. However, different types of physical exercise promote different responses in the organism. The present study was designed to investigate the expression of inflammatory cytokines and chemokines, and expression and activation of intracellular signaling proteins (CREB, ERK, Akt, p70S6k, STAT5, JNK, NFkB e p38) in the cerebral cortex and hippocampal formation of aged rats submitted to aerobic and resistance exercise. Inflammatory analysis showed that aged rats that underwent resistance training had decreased cortical levels of RANTES and a reduction in the hippocampal levels of MIP-2 when compared with control animals (sedentary). No significant difference was detected in the cortical and hippocampal inflammatory response between aerobic and sedentary groups. However, when comparing the two training models (aerobic vs resistance), it was observed that aerobic training increased the cortical levels of IL-13, IL-6, IL-17α compared with resistance training. Regarding the signaling proteins, a significant increase in cortical expression of the proteins JNK, ERK and p70S6k was found in the aerobic group in relation to the sedentary group. No significant change in the cortical and hippocampal expression of signaling proteins was detected between resistance training and sedentary groups. Nevertheless, when training models were compared, it was observed that aerobic training increased cortical expression of the total proteins p38, ERK, Akt and p70S6k in relation to resistance training. Taken together, these results show that changes in the brain expression of inflammatory and cell survival proteins in aged rats depend on the type of physical training.

NEDD9 promotes oncogenic signaling, a stem/mesenchymal gene signature, and aggressive ovarian cancer growth in mice.

Neural precursor cell expressed, developmentally downregulated 9 (NEDD9) supports oncogenic signaling in a number of solid and hematologic tumors. Little is known about the role of NEDD9 in ovarian carcinoma (OC), but available data suggest elevated mRNA and protein expression in advanced stage high grade cancers. We used a transgenic MISIIR-TAg mouse OC model combined with genetic ablation of Nedd9 to investigate its action in the development and progression of OC. A Nedd9−/− genotype delayed tumor growth rate, reduced incidence of ascites, and reduced expression and activation of signaling proteins including SRC, STAT3, E-cadherin and AURKA. Cell lines established from MISIIR-TAg;Nedd9−/− and MISIIR-TAg;Nedd9+/+ mice exhibited altered migration and invasion. Growth of these cells in a syngeneic allograft model indicated that systemic Nedd9 loss in the microenvironment had little impact on tumor allograft growth, but in a Nedd9 wild type background Nedd9−/− allografts exhibited significantly reduced growth, dissemination and oncogenic signaling compared to Nedd9+/+ allografts. Gene expression analysis revealed that Nedd9+/+ tumors exhibited more mesenchymal ‘stem-like’ transcriptional program, including increased expression of Aldh1a1 and Aldh1a2. Conversely, loss of Nedd9 resulted in increased expression of differentiation genes, including fallopian tube markers Foxj1, Ovgp1 and Pax8. Collectively, these data suggest that tumor cell-intrinsic Nedd9 expression promotes OC development and progression by broad induction of oncogenic protein signaling and stem/mesenchymal gene expression.

Deficiency of FAM3D (Family With Sequence Similarity 3, Member D), A Novel Chemokine, Attenuates Neutrophil Recruitment and Ameliorates Abdominal Aortic Aneurysm Development.

Chemokine-mediated neutrophil recruitment contributes to the pathogenesis of abdominal aortic aneurysm (AAA) and may serve as a promising therapeutic target. FAM3D (family with sequence similarity 3, member D) is a recently identified novel chemokine. Here, we aimed to explore the role of FAM3D in neutrophil recruitment and AAA development. FAM3D was markedly upregulated in human AAA tissues, as well as both elastase- and CaPO4-induced mouse aneurysmal aortas. FAM3D deficiency significantly attenuated the development of AAA in both mouse models. Flow cytometry analysis indicated that FAM3D-/- mice exhibited decreased neutrophil infiltration in the aorta during the early stage of AAA formation compared with their wild-type littermates. Moreover, application of FAM3D-neutralizing antibody 6D7 through intraperitoneal injection markedly ameliorated elastase-induced AAA formation and neutrophil infiltration. Further, in vitro coculture experiments with FAM3D-neutralizing antibody 6D7 and in vivo intravital microscopic analysis indicated that endothelial cell-derived FAM3D induced neutrophil recruitment. Mechanistically, FAM3D upregulated and activated Mac-1 (macrophage-1 antigen) in neutrophils, whereas inhibition of FPR1 (formyl peptide receptor 1) or FPR2 significantly blocked FAM3D-induced Mac-1 activation, indicating that the effect of FAM3D was dependent on both FPRs. Moreover, specific inhibitors of FPR signaling related to Gi protein or β-arrestin inhibited FAM3D-activated Mac-1 in vitro, whereas FAM3D deficiency decreased the activation of both FPR-Gi protein and β-arrestin signaling in neutrophils in vivo. FAM3D, as a dual agonist of FPR1 and FPR2, induced Mac-1-mediated neutrophil recruitment and aggravated AAA development through FPR-related Gi protein and β-arrestin signaling.