A structurally derived model of subunit‐dependent NMDA receptor function

NMDA receptors (NMDARs) are tetrameric complexes comprising two glycine‐binding GluN1 and two glutamate‐binding GluN2 subunits. Four GluN2 subunits encoded by different genes can produce up to 10 different di‐ and triheteromeric receptors. In addition, some neurological patients contain a de novo mutation or inherited rare variant in only one subunit. There is currently no mechanistic framework to describe tetrameric receptor function that can be extended to receptors with two different GluN1 or GluN2 subunits. Here we use the structural features of glutamate receptors to develop a mechanism describing both single channel and macroscopic NMDAR currents. We propose that each agonist‐bound subunit undergoes some rate‐limiting conformational change after agonist binding, prior to channel opening. We hypothesize that this conformational change occurs within a triad of interactions between a short helix preceding the M1 transmembrane helix, the highly conserved M3 motif encoded by the residues SYTANLAAF, and the linker preceding the M4 transmembrane helix of the adjacent subunit. Molecular dynamics simulations suggest that pre‐M1 helix motion is uncorrelated between subunits, which we interpret to suggest independent subunit‐specific conformational changes may influence these pre‐gating steps. According to this interpretation, these conformational changes are the main determinants of the key kinetic properties of NMDA receptor activation following agonist binding, and so these steps sculpt their physiological role. We show that this structurally derived tetrameric model describes both single channel and macroscopic data, giving a new approach to interpreting functional properties of synaptic NMDARs that provides a logical framework to understanding receptors with non‐identical subunits

Molecular biology of breast cancer metastasis: Clinical implications of experimental studies on metastatic inefficiency

Recent technological advances have led to an increasing ability to detect isolated tumour cells and groups of tumour cells in patients' blood, lymph nodes or bone marrow. However, the clinical significance of these cells is unclear. Should they be considered as evidence of metastasis, necessitating aggressive treatment, or are they in some cases unrelated to clinical outcome? Quantitative experimental studies on the basic biology of metastatic inefficiency are providing clues that may help in understanding the significance of these cells. This understanding will be of use in guiding clinical studies to assess the significance of isolated tumour cells and micrometastases in cancer patients

Reverse-Phase Phosphoproteome Analysis of Signaling Pathways Induced by Rift Valley Fever Virus in Human Small Airway Epithelial Cells

Rift valley fever virus (RVFV) infection is an emerging zoonotic disease endemic in many countries of sub-Saharan Africa and in Egypt. In this study we show that human small airway epithelial cells are highly susceptible to RVFV virulent strain ZH-501 and the attenuated strain MP-12. We used the reverse-phase protein arrays technology to identify phosphoprotein signaling pathways modulated during infection of cultured airway epithelium. ZH-501 infection induced activation of MAP kinases (p38, JNK and ERK) and downstream transcriptional factors [STAT1 (Y701), ATF2 (T69/71), MSK1 (S360) and CREB (S133)]. NF-κB phosphorylation was also increased. Activation of p53 (S15, S46) correlated with the increased levels of cleaved effector caspase-3, -6 and -7, indicating activation of the extrinsic apoptotic pathway. RVFV infection downregulated phosphorylation of a major anti-apoptotic regulator of survival pathways, AKT (S473), along with phosphorylation of FOX 01/03 (T24/31) which controls cell cycle arrest downstream from AKT. Consistent with this, the level of apoptosis inhibitor XIAP was decreased. However, the intrinsic apoptotic pathway marker, caspase-9, demonstrated only a marginal activation accompanied by an increased level of the inhibitor of apoptosome formation, HSP27. Concentration of the autophagy marker, LC3B, which often accompanies the pro-survival signaling, was decreased. Cumulatively, our analysis of RVFV infection in lung epithelium indicated a viral strategy directed toward the control of cell apoptosis through a number of transcriptional factors. Analyses of MP-12 titers in challenged cells in the presence of MAPK inhibitors indicated that activation of p38 represents a protective cell response while ERK activation controls viral replication

TIMP-1 Induces an EMT-Like Phenotypic Conversion in MDCK Cells Independent of Its MMP-Inhibitory Domain

Matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) regulate epithelial-mesenchymal transition (EMT) critical for the development of epithelial organs as well as cancer cell invasion. TIMP-1 is frequently overexpressed in several types of human cancers and serves as a prognostic marker. The present study investigates the roles of TIMP-1 on the EMT process and formation of the lumen-like structure in a 3D Matrigel culture of MDCK cells. We show that TIMP-1 overexpression effectively prevents cell polarization and acinar-like structure formation. TIMP-1 induces expression of the developmental EMT transcription factors such as SLUG, TWIST, ZEB1 and ZEB2, leading to downregulation of epithelial marker and upregulation of mesenchymal markers. Importantly, TIMP-1′s ability to induce the EMT-like process is independent of its MMP-inhibitory domain. To our surprise, TIMP-1 induces migratory and invasive properties in MDCK cells. Here, we present a novel finding that TIMP-1 signaling upregulates MT1-MMP and MMP-2 expression, and potentiates MT1-MMP activation of pro-MMP-2, contributing to tumor cell invasion. In spite of the fact that TIMP-1, as opposed to TIMP-2, does not interact with and inhibit MT1-MMP, TIMP-1 may act as a key regulator of MT1-MMP/MMP-2 axis. Collectively, our findings suggest a model in which TIMP-1 functions as a signaling molecule and also as an endogenous inhibitor of MMPs. This concept represents a paradigm shift in the current view of TIMP-1/MT1-MMP interactions and functions during cancer development/progression

Prognostic molecular markers in early breast cancer

A multitude of molecules involved in breast cancer biology have been studied as potential prognostic markers. In the present review we discuss the role of established molecular markers, as well as potential applications of emerging new technologies. Those molecules used routinely to make treatment decisions in patients with early-stage breast cancer include markers of proliferation (e.g. Ki-67), hormone receptors, and the human epidermal growth factor receptor 2. Tumor markers shown to have prognostic value but not used routinely include cyclin D(1 )and cyclin E, urokinase-like plasminogen activator/plasminogen activator inhibitor, and cathepsin D. The level of evidence for other molecular markers is lower, in part because most studies were retrospective and not adequately powered, making their findings unsuitable for choosing treatments for individual patients. Gene microarrays have been successfuly used to classify breast cancers into subtypes with specific gene expression profiles and to evaluate prognosis. RT-PCR has also been used to evaluate expression of multiple genes in archival tissue. Proteomics technologies are in development

Antibody Responses against Xenotropic Murine Leukemia Virus-Related Virus Envelope in a Murine Model

Xenotropic murine leukemia virus-related virus (XMRV) was recently discovered to be the first human gammaretrovirus that is associated with chronic fatigue syndrome and prostate cancer (PC). Although a mechanism for XMRV carcinogenesis is yet to be established, this virus belongs to the family of gammaretroviruses well known for their ability to induce cancer in the infected hosts. Since its original identification XMRV has been detected in several independent investigations; however, at this time significant controversy remains regarding reports of XMRV detection/prevalence in other cohorts and cell type/tissue distribution. The potential risk of human infection, coupled with the lack of knowledge about the basic biology of XMRV, warrants further research, including investigation of adaptive immune responses. To study immunogenicity in vivo, we vaccinated mice with a combination of recombinant vectors expressing codon-optimized sequences of XMRV gag and env genes and virus-like particles (VLP) that had the size and morphology of live infectious XMRV.Immunization elicited Env-specific binding and neutralizing antibodies (NAb) against XMRV in mice. The peak titers for ELISA-binding antibodies and NAb were 1:1024 and 1:464, respectively; however, high ELISA-binding and NAb titers were not sustained and persisted for less than three weeks after immunizations.Vaccine-induced XMRV Env antibody titers were transiently high, but their duration was short. The relatively rapid diminution in antibody levels may in part explain the differing prevalences reported for XMRV in various prostate cancer and chronic fatigue syndrome cohorts. The low level of immunogenicity observed in the present study may be characteristic of a natural XMRV infection in humans

Are mesenchymal stromal cells immune cells?

Mesenchymal stromal cells (MSCs) are considered to be promising agents for the treatment of immunological disease. Although originally identified as precursor cells for mesenchymal lineages, in vitro studies have demonstrated that MSCs possess diverse immune regulatory capacities. Pre-clinical models have shown beneficial effects of MSCs in multiple immunological diseases and a number of phase 1/2 clinical trials carried out so far have reported signs of immune modulation after MSC infusion. These data indicate that MSCs play a central role in the immune response. This raises the academic question whether MSCs are immune cells or whether they are tissue precursor cells with immunoregulatory capacity. Correct understanding of the immunological properties and origin of MSCs will aid in the appropriate and safe use of the cells for clinical therapy. In this review the whole spectrum of immunological properties of MSCs is discussed with the aim of determining the position of MSCs in the immune system

Fascin overexpression promotes neoplastic progression in oral squamous cell carcinoma

<p>Abstract</p> <p>Background</p> <p>Fascin is a globular actin cross-linking protein, which plays a major role in forming parallel actin bundles in cell protrusions and is found to be associated with tumor cell invasion and metastasis in various type of cancers including oral squamous cell carcinoma (OSCC). Previously, we have demonstrated that fascin regulates actin polymerization and thereby promotes cell motility in K8-depleted OSCC cells. In the present study we have investigated the role of fascin in tumor progression of OSCC.</p> <p>Methods</p> <p>To understand the role of fascin in OSCC development and/or progression, fascin was overexpressed along with vector control in OSCC derived cells AW13516. The phenotype was studied using wound healing, Boyden chamber, cell adhesion, Hanging drop, soft agar and tumorigenicity assays. Further, fascin expression was examined in human OSCC samples (N = 131) using immunohistochemistry and level of its expression was correlated with clinico-pathological parameters of the patients.</p> <p>Results</p> <p>Fascin overexpression in OSCC derived cells led to significant increase in cell migration, cell invasion and MMP-2 activity. In addition these cells demonstrated increased levels of phosphorylated AKT, ERK1/2 and JNK1/2. Our in vitro results were consistent with correlative studies of fascin expression with the clinico-pathological parameters of the OSCC patients. Fascin expression in OSCC showed statistically significant correlation with increased tumor stage (<it>P </it>= 0.041), increased lymph node metastasis (<it>P </it>= 0.001), less differentiation (<it>P </it>= 0.005), increased recurrence (<it>P </it>= 0.038) and shorter survival (<it>P </it>= 0.004) of the patients.</p> <p>Conclusion</p> <p>In conclusion, our results indicate that fascin promotes tumor progression and activates AKT and MAPK pathways in OSCC-derived cells. Further, our correlative studies of fascin expression in OSCC with clinico-pathological parameters of the patients indicate that fascin may prove to be useful in prognostication and treatment of OSCC.</p

The Tumor Microenvironment: The Making of a Paradigm

What has been will be again, what has been done will be done again; there is nothing new under the su