static-adhesion-assay-for-study-integrin-activation-t

Static Adhesion Assay for the Study of Integrin Activation in T Lymphocytes

T lymphocyte adhesion is required for multiple T cell functions, including migration to sites of inflammation and formation of immunological synapses with ...

assay-adhesion-under-shear-stress-for-study-t-lymphocyte-adhesion

Assay of Adhesion Under Shear Stress for the Study of T Lymphocyte-Adhesion Molecule Interactions

Overall, T cell adhesion is a critical component of function, contributing to the distinct processes of cellular recruitment to sites of inflammation and ...

co-immunoprecipitation-assay-for-studying-functional-interactions

Co-immunoprecipitation Assay for Studying Functional Interactions Between Receptors and Enzymes

Receptor-associated enzymes are the major mediators of cellular activation. These enzymes are regulated, at least in part, by physical interactions with ...

Ischemia is a stimulus for production of angiogenic cytokines that activate local vascular cells and mobilize angiogenic cells to the circulation. These responses are impaired in elderly patients with peripheral arterial disease. Hypoxia-inducible factor (HIF)-1 mediates adaptive responses to ischemia, including production of angiogenic cytokines. In this study, we demonstrate that aging and HIF-1 loss-of-function impair the expression of multiple angiogenic cytokines, mobilization of angiogenic cells, maintenance of tissue viability, and recovery of limb perfusion following femoral artery ligation. We show that HIF-1 directly activates transcription of the gene encoding stem cell factor and that mice lacking the cognate receptor C-KIT have impaired recovery from ischemia. Administration of AdCA5, an adenovirus encoding a constitutively active form of HIF-1alpha, improved the recovery of perfusion in older mice to levels similar to those in young mice. Injection of AdCA5 into nonischemic limb was sufficient to increase the number of circulating angiogenic cells. These results indicate that HIF-1 activity is necessary and sufficient for the mobilization of angiogenic cells and that HIF-1alpha gene therapy can counteract the pathological effects of aging in a mouse model of limb ischemia.

Effects of aging and hypoxia-inducible factor-1 activity on angiogenic cell mobilization and recovery of perfusion after limb ischemia.

Human immunodeficiency virus type 1 (HIV-1) primarily infects CD4(+) T cells and cells of the monocyte-macrophage lineage, resulting in immunodeficiency in an infected patient. Along with this immune deficiency, HIV-1 has been linked to a number of neurological symptoms in the absence of opportunistic infections or other co-morbidities, suggesting that HIV-1 is able to cross the blood-brain barrier (BBB), enter the central nervous system (CNS), and cause neurocognitive impairment. HIV-1-infected monocyte-macrophages traverse the BBB and enter the CNS throughout the course of HIV-1 disease. Once in the brain, both free virus and virus-infected cells are able to infect neighboring resident microglia and astrocytes and possibly other cell types. HIV-1-infected cells in both the periphery and the CNS give rise to elevated levels of viral proteins, including gp120, Tat, and Nef, and of host inflammatory mediators such as cytokines and chemokines. It has been shown that the viral proteins may act alone or in concert with host cytokines and chemokines, affecting the integrity of the BBB. The pathological end point of these interactions may facilitate a positive feedback loop resulting in increased penetration of HIV into the CNS. It is proposed in this review that the dysregulation of the BBB during and after neuroinvasion is a critical component of the neuropathogenic process and that dysregulation of this protective barrier is caused by a combination of viral and host factors including secreted viral proteins, components of the inflammatory process, the aging process, therapeutics, and drug or alcohol abuse.

Breaking down the barrier: the effects of HIV-1 on the blood-brain barrier.

About one third of acquired immunodeficiency syndrome cases in the USA have been attributed to the use of injected addictive drugs, frequently involving opioids like heroin and morphine, establishing them as significant predisposing risk factors for contracting human immunodeficiency virus type 1 (HIV-1). Accumulating evidence from in vitro and in vivo experimental systems indicates that opioids act in concert with HIV-1 proteins to exacerbate dysregulation of neural and immune cell function and survival through diverse molecular mechanisms. In contrast, the impact of opioid exposure and withdrawal on the viral life cycle and HIV-1 disease progression itself is unclear, with conflicting reports emerging from the simian immunodeficiency virus and simian-human immunodeficiency virus infection models. However, these studies suggest a potential role of opioids in elevated viral production. Because human microglia, astrocytes, CD4+ T lymphocytes, and monocyte-derived macrophages express opioid receptors, it is likely that intracellular signaling events triggered by morphine facilitate enhancement of HIV-1 infection in these target cell populations. This review highlights the biochemical changes that accompany prolonged exposure to and withdrawal from morphine that synergize with HIV-1 proteins to disrupt normal cellular physiological functions especially within the central nervous system. More importantly, it collates evidence from epidemiological studies, animal models, and heterologous cell systems to propose a mechanistic link between such physiological adaptations and direct modulation of HIV-1 production. Understanding the opioid-HIV-1 interface at the molecular level is vitally important in designing better treatment strategies for HIV-1-infected patients who abuse opioids.

Role of mu-opioids as cofactors in human immunodeficiency virus type 1 disease progression and neuropathogenesis.

Etwa ein Drittel der erworbene Immunschw&auml;che-Syndrom-F&auml;lle in den USA haben die Verwendung von injizierten s&uuml;chtig machenden Drogen, bei denen Opioide wie Heroin und Morphin, &uuml;ber ihre Errichtung als wesentliche pr&auml;disponierende Risikofaktoren f&uuml;r contracting humanen Immundefizienz-Virustyp 1 (HIV-1) zugeschrieben. Sammeln Beweise von in-vitro und in vivo experimentelle Systeme gibt, dass Opioide mit HIV-1 Proteine Zusammenwirken, Dysregulation der neuronalen und immun Zelle Funktion und &Uuml;berleben durch verschiedene Molekulare Mechanismen zu versch&auml;rfen. Im Gegensatz dazu ist die Auswirkungen der opioid-Exposition und der R&uuml;ckzug auf den viralen Lebenszyklus und Fortschreiten der HIV-1-Erkrankung selbst unklar, mit widerspr&uuml;chliche Berichte aus dem Simianes Immundefizienz-Virus und Simian-menschlichen Immunschw&auml;che-Virus-Infektion-Modelle. Jedoch deutet diese Untersuchungen eine m&ouml;gliche Rolle von Opioiden in der erh&ouml;hten virale Produktion. Weil menschliche Mikroglia, Astrozyten, CD4 + T-Lymphozyten und Makrophagen Monozyte abgeleitete Opioidrezeptoren Ausdr&uuml;cken, ist es wahrscheinlich, dass die intrazellul&auml;re signalisierende Ereignisse, die ausgel&ouml;st durch Morphin Verbesserung der HIV-1 Infektion in diese Zelle Zielgruppen erleichtern. Diesen Bericht unterstreicht die biochemischen &Auml;nderungen, die begleiten sehr intensiver und Entzug von Morphin synergize mit HIV-1 Proteine normale physiologische Zellfunktionen vor allem innerhalb des zentralen Nervensystems zu st&ouml;ren. Noch wichtiger ist, sammelt es Hinweise aus epidemiologischen Studien, Tiermodelle und heterologe Zellsysteme eine mechanistische Verbindung zwischen solchen physiologischen Anpassungen und direkten Modulation der HIV-1-Produktion vorschlagen. Grundlagen der Opioid-HIV-1-Schnittstelle auf molekularer Ebene ist unbedingt bei der Gestaltung von besserer Strategien zur Behandlung von HIV-1-infizierten Patienten, die Opioide zu missbrauchen.

Rolle des Mu-Opioiden als Kofaktoren in humanen Immundefizienz-Virus Typ 1 Krankheitsverlauf und Neuropathogenesis.

Programmed death-1 (PD-1) is a co-receptor that is expressed predominantly by T cells. The binding of PD-1 to its ligands, PD-L1 or PD-L2, is vital for the physiologic regulation of the immune system. A major functional role of the PD-1 signaling pathway is the inhibition of self-reactive T cells, which serve to protect against autoimmune diseases. Elimination of the PD-1 pathway can therefore result in the breakdown of immune tolerance that can ultimately lead to the development of pathogenic autoimmunity. Conversely, tumor cells can at times co-opt the PD-1 pathway to escape from immunosurveillance mechanisms. Therefore, blockade of the PD-1 pathway has become an attractive target in cancer therapy. Recent clinical trials have shown that anti-PD-1 agents have profound effects on solid tumor regression. Current approaches include six agents that are either PD-1 and PD-L1 targeted neutralizing antibodies or fusion proteins. More than forty clinical trials are underway to better define the role of PD-1 blockade in variety of tumor types. In this review we will highlight the basic biology of the PD-1 system and discuss its potential roles in both autoimmunity and cancer. We propose that future research on PD-1 may lead to the translation of fundamental regulatory pathways into the development of practical new approaches for the treatment of autoimmune diseases and cancer.

Programmed death-1 pathway in cancer and autoimmunity.

The coreceptor programmed death 1 inhibits T-cell adhesion by regulating Rap1.

Approximately one-third of the AIDS cases in the United States have been attributed to the use of injected drugs, frequently involving the abuse of opioids. Consequently, it is critical to address whether opioid use directly contributes to altered susceptibility to HIV-1 beyond the increased risk of exposure. Previous in vitro and in vivo studies addressing the role of μ-opioid agonists in altering levels of the co-receptor CXCR4 and subsequent HIV-1 replication have yielded contrasting results. The bone marrow is believed to be a potential anatomical sanctuary for HIV-1.

Effect of μ-opioid agonist DAMGO on surface CXCR4 and HIV-1 replication in TF-1 human bone marrow progenitor cells.

Ras and Rap proteins are closely related small guanosine triphosphatase (GTPases) that share similar effector-binding domains but operate in a very different signaling networks; Ras has a dominant role in cell proliferation, while Rap mediates cell adhesion. Ras and Rap proteins are regulated by several shared processes such as post-translational modification, phosphorylation, activation by guanine exchange factors and inhibition by GTPase-activating proteins. Sub-cellular localization and trafficking of these proteins to and from the plasma membrane are additional important regulatory features that impact small GTPases function. Despite its importance, the trafficking mechanisms of Ras and Rap proteins are not completely understood. Chaperone proteins play a critical role in trafficking of GTPases and will be the focus of the discussion in this work. We will review several aspects of chaperone biology focusing on specificity toward particular members of the small GTPase family. Understanding this specificity should provide key insights into drug development targeting individual small GTPases.

Chaperone-mediated specificity in Ras and Rap signaling.

CD28 is a coreceptor expressed on T lymphocytes. Signaling downstream of CD28 promotes multiple T cell functions such as proliferation, survival, and cytokine secretion. Adhesion to APCs is another function of T cells; however, little is known with regard to the role of CD28 in this process. Our previous studies have shown that CD28 inhibits T cell adhesion, but the underlying mechanism that mediates this process remains unknown. In the present study we discovered that signaling downstream of CD28 resulted in inhibition of Rap1 activity and decreased LFA-1-mediated adhesion. We showed that this was regulated by the recruitment of calcium-promoted Ras inactivator (CAPRI), a GTPase-activating protein, to the plasma membrane downstream of CD28 signaling. CAPRI trafficking to the plasma membrane was secondary to calcium influx and was mediated by its C2A and C2B domains. We conclude that CD28 inhibits Rap1-mediated adhesion by recruiting the protein CAPRI to the plasma membrane.

CD28 inhibits T cell adhesion by recruiting CAPRI to the plasma membrane.

Programmed death-1 (PD-1) is an inhibitory co-receptor that is highly expressed in T lymphocytes that has been shown to downregulate inflammatory responses in several inflammatory diseases including systemic lupus erythematosus and rheumatoid arthritis. Yet, the role of PD-1 in psoriatic arthritis (PsA) has not been studied. In order to fill this gap, we measured the expression levels of PD-1 in peripheral T cells from patients with active disease. Twenty patients and fifteen age-matched healthy control subjects were recruited. The percentage of CD3(+)PD-1(+) T cells was measured by flow cytometry. Despite normal concentration of peripheral T cells, the expression levels of PD-1 were significantly higher in patients compared to healthy controls. Interestingly, among the patients, the expression levels inversely correlated with disease activity measured by disease activity scores (DAS28). PD-1 expression levels strongly correlated with the number of tender and swollen joints, but not with C-reactive protein (CRP) levels or psoriasis area and severity index (PASI). Functionally, in vitro ligation of PD-1 receptor in PsA T cells inhibited interleukin-2 (IL-2) secretion, Akt phosphorylation, and Rap1 activation. These findings suggest that PD-1 might serve as a biomarker for disease activity in PsA and highlight the need for additional studies in order to establish the role of PD-1 in PsA pathogenesis.

Analysis of Programmed Death-1 in Patients with Psoriatic Arthritis.

Autoimmune diseases arise from aberrant activation of immune cells directed against endogenous autoantigens expressed throughout the human body. While the initiating triggers remain poorly understood, the self-perpetuating phase of these diseases is directly linked to the ongoing recruitment of inflammatory cells that traffic to the affected anatomical sites. T lymphocytes are prominent drivers of many autoimmune diseases and the targeted trafficking of these cells to infiltrate the affected organs is often a common denominator. The regulation of T cell trafficking involves the coordinated expression of specific patterns of chemokines and the reciprocal expression of cognate chemokine receptors on T cell membranes. Thereby, chemokines direct the specific trafficking of a wide array of responsive activated immune cells. Specific patterns of chemokine receptor expression can correlate with disease activity in an autoimmune disease, confirming the importance of further characterizing the T cells that infiltrate specific sites of autoimmunity. Herein, we will review our current understanding of the roles of chemokines in two common autoimmune diseases: rheumatoid arthritis and multiple sclerosis. We also discuss the implications for chemokine receptor signatures in autoimmune pathogenesis, and how these may provide novel targets for therapeutic intervention.

T cell chemokine receptor patterns as pathogenic signatures in autoimmunity.

Erratum to: Analysis of Programmed Death-1 in Patients with Psoriatic Arthritis.

Regulation of integrins is critical for lymphocyte adhesion to endothelium and trafficking through secondary lymphoid organs. Inside-out signaling to integrins is mediated by the small GTPase Rap1. Two effectors of Rap1 regulate integrins, RapL and Rap1 interacting adaptor molecule (RIAM). Using mice conditionally deficient in both Rap1a and Rap1b and mice null for RIAM, we show that the Rap1/RIAM module is not required for T- or B-cell development but is essential for efficient adhesion to intercellular adhesion molecule (ICAM) 1 and vascular cell adhesion molecule (VCAM) 1 and for proper trafficking of lymphocytes to secondary lymphoid organs. Interestingly, in RIAM-deficient mice, whereas peripheral lymph nodes (pLNs) were depleted of both B and T cells and recirculating B cells were diminished in the bone barrow (BM), the spleen was hypercellular, albeit with a relative deficiency of marginal zone B cells. The abnormality in lymphocyte trafficking was accompanied by defective humoral immunity to T-cell-dependent antigens. Platelet function was intact in RIAM-deficient animals. These in vivo results confirm a role for RIAM in the regulation of some, but not all, leukocyte integrins and suggest that RIAM-regulated integrin activation is required for trafficking of lymphocytes from blood into pLNs and BM, where relatively high shear forces exist in high endothelial venules and sinusoids, respectively.

Rap1 and its effector RIAM are required for lymphocyte trafficking.

Numerous systems exist to model the blood-brain barrier (BBB) with the goal of understanding the regulation of passage into the central nervous system (CNS) and the potential impact of selected insults on BBB function. These models typically focus on the intrinsic cellular properties of the BBB, yet studies of peripheral cell migration are often excluded due to technical restraints.

Co-culture model consisting of human brain microvascular endothelial and peripheral blood mononuclear cells.

The blood-brain barrier (BBB) has been defined as a critically important protective barrier that is involved in providing essential biologic, physiologic, and immunologic separation between the central nervous system (CNS) and the periphery. Insults to the BBB can cause overall barrier damage or deregulation of the careful homeostasis maintained between the periphery and the CNS. These insults can, therefore, yield numerous phenotypes including increased overall permeability, interendothelial gap formation, alterations in cytokine and chemokine secretion, and accelerated cellular passage. The current studies expose the human brain microvascular endothelial cell line, hCMEC/D3, to prolonged morphine exposure and aim to uncover the mechanisms underlying alterations in barrier function in vitro. These studies show alterations in the mRNA and protein levels of the cellular adhesion molecules (CAMs) intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and activated leukocyte cell adhesion molecule that correlate with an increased firm adhesion of the CD3⁺ subpopulation of peripheral blood mononuclear cells (PBMCs). Overall, these studies suggest that prolonged morphine exposure may result in increased cell migration into the CNS, which may accelerate pathological processes in many diseases that involve the BBB.

ArticleTotal : 3	Year
Publication title Cited by 21	2014
Publication title Cited by 3	2016
Publication title Cited by 1	2018

Article	Year
Effects of aging and hypoxia-inducible factor-1 activity on angiogenic cell mobilization and recovery of perfusion after limb ischemia. Circulation research\| PubMed ID: 17932327	2007
Breaking down the barrier: the effects of HIV-1 on the blood-brain barrier. Brain research\| PubMed ID: 21641584	2011
Role of mu-opioids as cofactors in human immunodeficiency virus type 1 disease progression and neuropathogenesis. Journal of neurovirology\| PubMed ID: 21735315	2011
Programmed death-1 pathway in cancer and autoimmunity. Clinical immunology (Orlando, Fla.)\| PubMed ID: 24780173	2014
The coreceptor programmed death 1 inhibits T-cell adhesion by regulating Rap1. The Journal of allergy and clinical immunology\| PubMed ID: 25240786	2015
Effect of μ-opioid agonist DAMGO on surface CXCR4 and HIV-1 replication in TF-1 human bone marrow progenitor cells. BMC research notes\| PubMed ID: 25338959	2014
Chaperone-mediated specificity in Ras and Rap signaling. Critical reviews in biochemistry and molecular biology\| PubMed ID: 25488471	2015
CD28 inhibits T cell adhesion by recruiting CAPRI to the plasma membrane. Journal of immunology (Baltimore, Md. : 1950)\| PubMed ID: 25637021	2015
Analysis of Programmed Death-1 in Patients with Psoriatic Arthritis. Inflammation\| PubMed ID: 25663558	2015
T cell chemokine receptor patterns as pathogenic signatures in autoimmunity. Discovery medicine\| PubMed ID: 25725226	2015
Erratum to: Analysis of Programmed Death-1 in Patients with Psoriatic Arthritis. Inflammation\| PubMed ID: 25847627	2015
Rap1 and its effector RIAM are required for lymphocyte trafficking. Blood\| PubMed ID: 26324702	2015
Co-culture model consisting of human brain microvascular endothelial and peripheral blood mononuclear cells. Journal of neuroscience methods\| PubMed ID: 27216631	2016
Prolonged Morphine Exposure Induces Increased Firm Adhesion in an in Vitro Model of the Blood-Brain Barrier. International journal of molecular sciences\| PubMed ID: 27294916	2016

Marianne Strazza

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