Biomedical subjects

J E Groopman

Publications and source records attributed to J E Groopman.

At least 19 recordsLinked to original sources

PubMed1998

Stromal cell-derived factor-1 alpha and stem cell factor/kit ligand share signaling pathways in hemopoietic progenitors: a potential mechanism for cooperative induction of chemotaxis.

Stromal cell-derived factor (SDF-1alpha), the ligand for CXCR4, is a chemokine that acts as a potent chemoattractant for hemopoietic progenitor cells. Stem cell factor/kit ligand (SCF/KL), an early acting cytokine, has recently been reported to enhance the chemotaxis induced by SDF-1alpha. However, very little is known about downstream signaling events following these receptor-ligand interactions. To investigate these events, we utilized a model progenitor cell line, CTS, which expresses both the CXCR4 and c-kit receptors. We observed strong Ca2+ mobilization and enhancement of chemotaxis following treatment with SDF-1alpha or SCF/KL. A combination of these factors enhanced this chemotaxis in CTS cells as well as in CD34+ bone marrow cells. Prior treatment of CTS cells with pertussis toxin inhibited the SDF-1alpha-induced chemotaxis, suggesting that SDF-1alpha signaling involves a pertussis-sensitive Gi-coupled protein. SDF-1alpha treatment resulted in a rapid phosphorylation of the focal adhesion molecules RAFTK (related adhesion focal tyrosine kinase), paxillin, and p130cas, which then declined within minutes. SCF/KL alone or in combination with SDF-1alpha induced a rapid and sustained effect on phosphorylation of these substrates. SDF-1alpha treatment resulted in a rapid and robust activation of p44/42 mitogen-activated protein kinase compared with the relatively weak and delayed effect of SCF/KL treatment. Interestingly, a delayed but sustained activation of mitogen-activated protein kinase activation was observed when the factors were used in combination. Such cooperativity in downstream signaling pathways may explain the enhanced chemotaxis of progenitors observed with SDF-1alpha in combination with SCF/KL.

Antigens, CD34↗

PubMed1998

The alpha-chemokine, stromal cell-derived factor-1alpha, binds to the transmembrane G-protein-coupled CXCR-4 receptor and activates multiple signal transduction pathways.

The alpha-chemokine stromal cell-derived factor (SDF)-1alpha binds to the seven transmembrane G-protein-coupled CXCR-4 receptor and acts to modulate cell migration and proliferation. The signaling pathways that mediate the effects of SDF-1alpha are not well characterized. We studied events following SDF-1alpha binding to CXCR-4 in a model murine pre-B cell line transfected with human CXCR-4. There was enhanced tyrosine phosphorylation and association of components of focal adhesion complexes such as the related adhesion focal tyrosine kinase, paxillin, and Crk. We also observed activation of phosphatidylinositol 3-kinase. Wortmannin, a selective inhibitor of phosphatidylinositol 3-kinase, partially inhibited the SDF-1alpha-induced migration and tyrosine phosphorylation of paxillin. SDF-1alpha treatment selectively activated p44/42 mitogen-activated protein kinase (Erk 1 and Erk 2) and its upstream kinase mitogen-activated protein kinase kinase but not p38 mitogen-activated protein kinase, c-Jun amino-terminal kinase or mitogen activated protein kinase kinase. We also observed that SDF-1alpha treatment increased NF-kappaB activity in nuclear extracts from the CXCR-4 transfectants. Taken together, these studies revealed that SDF-1alpha activates distinct signaling pathways that may mediate cell growth, migration, and transcriptional activation.

Animals↗

PubMed1998

The alpha-chemokine receptor CXCR4 is expressed on the megakaryocytic lineage from progenitor to platelets and modulates migration and adhesion.

CXCR4 is the receptor for the alpha-chemokine stromal cell-derived factor 1 (SDF-1) and has been shown to be expressed on a diversity of leukocytes. In this report, the expression of the CXCR4 receptor in cells of megakaryocytic lineage and the role of SDF-1 in megakaryocytopoiesis were investigated. Using flow cytometry in combination with reverse transcriptase-polymerase chain reaction (RT-PCR), we observed that bone marrow CD34(+), CD61(+) cells, blood platelets, and megakaryocytic leukemia cell lines all expressed the CXCR4 receptor. To examine the expression of the CXCR4 receptor on megakaryocyte progenitors (colony-forming units-megakaryocyte [CFU-Meg]), CXCR4-positive and -negative CD34(+) populations were separated from bone marrow and cultured in a plasma clot culture system. A subpopulation of the CFU-Meg was found in the CXCR4-positive fraction. The functional significance of CXCR4 expression on cells of the megakaryocytic lineage was examined by studying the effects of SDF-1alpha on migration and proliferation of megakaryocyte progenitor cells in vitro. We found that SDF-1alpha potently induced megakaryocyte progenitor migration and significantly enhanced adhesion of mature marrow megakaryocytes to endothelium. No marked effects of SDF-1alpha alone or in combination with thrombopoietin and stem cell factor/kit ligand on megakaryocyte production in vitro were noted. These results demonstrate for the first time that the CXCR4 alpha-chemokine receptor is expressed on cells of the megakaryocytic lineage from progenitors to platelets and that its ligand SDF-1alpha may modulate several aspects of megakaryocytopoiesis.

Antigens, CD34↗

PubMed1998

The related adhesion focal tyrosine kinase (RAFTK) is tyrosine phosphorylated and participates in colony-stimulating factor-1/macrophage colony-stimulating factor signaling in monocyte-macrophages.

RAFTK, a novel nonreceptor protein kinase, has been shown to be involved in focal adhesion signal transduction pathways in neuronal PC12 cells, megakaryocytes, platelets, and T cells. Because focal adhesions may modulate cytoskeletal functions and thereby alter phagocytosis, cell migration, and adhesion in monocyte-macrophages, we investigated the role of RAFTK signaling in these cells. RAFTK was abundantly expressed in THP1 monocytic cells as well as in primary alveolar and peripheral blood-derived macrophages. Colony-stimulating factor-1 (CSF-1)/macrophage colony-stimulating factor (M-CSF) stimulation of THP1 cells increased the tyrosine phosphorylation of RAFTK; similar increases in phosphorylation were also detected after lipopolysaccharide stimulation. RAFTK was phosphorylated with similar kinetics in THP1 cells and peripheral blood-derived macrophages. Immunoprecipitation analysis showed associations between RAFTK and the signaling molecule phosphatidylinositol-3 (PI-3) kinase. PI-3 kinase enzyme activity also coprecipitated with the RAFTK antibody, further confirming this association. The CSF-1/M-CSF receptor c-fms and RAFTK appeared to associate in response to CSF-1/M-CSF treatment of THP1 cells. Inhibition of RAFTK by a dominant-negative kinase mutant reduced CSF-1/M-CSF-induced MAPK activity. These data indicate that RAFTK participates in signal transduction pathways mediated by CSF-1/M-CSF, a cytokine that regulates monocyte-macrophage growth and function.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed1998

Beta-chemokine receptor CCR5 signals via the novel tyrosine kinase RAFTK.

Chemokine receptors are coupled to G-proteins and their activation results in prominent changes in cell migration and growth. The downstream signaling pathways that mediate these effects of chemokines are largely uncharacterized. Macrophage inflammatory protein 1 beta (MIP 1 beta) binding to its cognate receptor CCR5 resulted in activation of the related adhesion focal tyrosine kinase (RAFTK), with subsequent activation of the cytoskeletal protein paxillin and the down-stream transcriptional activators, c-Jun N-terminal kinase (JNK)/stress-activated protein kinase (SAPK) and p38 mitogen-activated protein (MAP) kinase. Inhibition of RAFTK by a dominant-negative kinase mutant markedly attenuated JNK/ SAPK activity. Thus, RAFTK appears to provide a functional "bridge" for the transmission of CCR5 receptor signaling to the cytoskeleton and nucleus, primary sites of chemotaxis and growth regulation.

Animals↗

PubMed1998

Human immunodeficiency virus tat modulates the Flk-1/KDR receptor, mitogen-activated protein kinases, and components of focal adhesion in Kaposi's sarcoma cells.

Kaposi's sarcoma (KS) spindle cell growth and spread have been reported to be modulated by various cytokines as well as the human immunodeficiency virus (HIV) gene product Tat. Recently, HIV-1 Tat has been shown to act like a cytokine and bind to the Flk-1/KDR receptor for the vascular endothelial growth factor A (VEGF-A), which is expressed by KS cells. We have characterized signal transduction pathways stimulated by HIV-1 Tat upon its binding to surface receptors on KS cells. We observed that stimulation in KS 38 spindle cells resulted in tyrosine phosphorylation and activation of the Flk-1/KDR receptor. We also report that HIV-1 Tat treatment enhanced the phosphorylation and association of proteins found in focal adhesions, such as the related adhesion focal tyrosine kinase RAFTK, paxillin, and p130(cas). Further characterization revealed the activation of mitogen-activated protein kinase, c-Jun amino-terminal kinase (JNK), and Src kinase. HIV-1 Tat contains a basic domain which can interact with growth factor tyrosine kinase receptors and a classical RGD sequence which may bind to and activate the surface integrin receptors for fibronectin and vitronectin. We observed that stimulation of KS cells with basic as well as RGD sequence-containing Tat peptides resulted in enhanced phosphorylation of RAFTK and activation of MAP kinase. These studies reveal that Tat stimulation activates a number of signal transduction pathways that are associated with cell growth and migration.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed1998

Fatigue in cancer and HIV/AIDS.

Fatigue is a common and troubling symptom in patients with cancer or HIV/AIDS, resulting in significant disability and adverse effects on quality of life. Its etiology remains complex and is most likely multifactorial. Despite its impact and prevalence, fatigue is often overlooked and undertreated in these patient populations. The general perceptions of fatigue are that its etiology cannot be determined, it is an inevitable manifestation that must be endured, and few interventions are available. Efforts are ongoing to better understand the etiology, characteristics, and consequences of fatigue in patients with cancer or HIV/AIDS. New practical methods of assessing it in cancer patients are now available. In order to improve the quality of life in these patients, physicians need to reassess their perceptions of fatigue and their approach to its diagnosis and management. There are recognizable causes and correlates for which interventions can be beneficial. These include anemia, pain, infection/fever, hormonal or nutritional deficiencies, depression/anxiety, sleep disturbances, and excessive inactivity or rest. Physicians should fully evaluate patients to identify the factors amenable to management. Fatigue is also seldom discussed by patients and their physicians. Improved communication with and counseling of patients and their caregivers can play an important role in the effective assessment and management of fatigue in patients with cancer or HIV/AIDS. Many patients may benefit from wider implementation of recent advances in the understanding and treatment of fatigue in these oncologic and infectious conditions.

Fatigue↗

PubMed1997

Signal transduction in human hematopoietic cells by vascular endothelial growth factor related protein, a novel ligand for the FLT4 receptor.

We have recently identified a novel ligand of the vascular endothelial growth factor (VEGF) family termed VEGF-related protein (VRP), which specifically binds to the FLT4 receptor. To characterize the signaling events after VRP engagement of its cognate receptor in hematopoietic cells, a population of human erythroleukemia (HEL) cells, termed HEL-JW, expressing high levels of FLT4 receptor was isolated. Stimulation of HEL-JW cells with VRP alone and in combination with the c-kit ligand/stem cell factor increased cell growth. VRP induced tyrosine phosphorylation of various proteins, including the FLT4 receptor. Further characterization of these tyrosine phosphorylated molecules revealed that Shc, Grb2, and SOS form a complex with the activated FLT4 receptor. HEL-JW cells also expressed RAFTK, a recently identified member of the focal adhesion kinase family. RAFTK was phosphorylated and activated upon VRP treatment, and there was an enhanced association of this kinase with the adaptor protein Grb2. Furthermore, the c-Jun NH2-terminal kinase (JNK), involved in growth activation and shown to mediate RAFTK signaling in other cell types, was activated by VRP stimulation. We also observed that VRP treatment of HEL-JW cells resulted in the phosphorylation of the cytoskeletal protein paxillin. This treatment resulted in an increased association of paxillin with RAFTK, which was mediated by the C-terminal region of RAFTK. These studies indicate that VRP stimulation induced the formation of a signaling complex at its activated receptor as well as activation of RAFTK. VRP-mediated activation of RAFTK may facilitate signal transduction to the cytoskeleton and downstream to the JNK pathway in FLT4-expressing blood cells.

Carrier Proteins↗

PubMed1997

Characterization of signal transduction pathways in human bone marrow endothelial cells.

Human bone marrow endothelial cells immortalized with the T antigen of SV40 (TrHBMEC) have previously been characterized by us with regard to their properties that are similar to primary marrow endothelial cells and their utility as a model system. We now report that TrHBMEC express a recently discovered signal transduction molecule termed RAFTK (related adhesion focal tyrosine kinase), also called Pyk2 or CAK-beta. RAFTK, the second member of the focal adhesion kinase (FAK) family, is known to be activated in response to calcium flux in neuronal cells and integrin stimulation in megakaryocytes and B cells. We have studied the effects of cytokines on RAFTK activation in TrHBMEC. Treatment of TrHBMEC with the vascular endothelial growth factor (VEGF), as well as the VEGF-related protein (VRP), the recently identified ligand for the FLT-4 receptor, resulted in enhanced tyrosine phosphorylation of RAFTK. Similar changes in RAFTK phosphorylation were observed upon stimulation of TrHBMEC with basic fibroblast growth factor (bFGF) or oncostatin M (OSM). Stimulation of these cells with growth factors also resulted in an increase in RAFTK activity and the c-Jun NH2-terminal kinase (JNK). RAFTK coimmunoprecipitated with the cytoskeletal protein paxillin through its C-terminal proline-rich domain in TrHBMEC. These results suggest that, in marrow endothelium, activation of RAFTK by VEGF, VRP, OSM, and bFGF represents a new element in the signal transduction pathways used by these growth factors and likely acts to coordinate signaling from their surface receptors to the cytoskeleton, thereby modulating cell growth and function.

Bone Marrow↗

PubMed1997

Differential effects of interleukin-13 on cytomegalovirus and human immunodeficiency virus infection in human alveolar macrophages.

Alveolar macrophages, which form a principal line of defense against a variety of pulmonary pathogens, may themselves be infected by viruses like human immunodeficiency virus-1 (HIV-1), which impair their defensive functions. Interleukin-13 (IL-13), a multifunctional cytokine, has been considered for therapeutic use based on its potent inhibition of HIV-1 in these cells. We have further examined the effects of IL-13 on alveolar macrophages under conditions that reflect those seen in acquired immune deficiency syndrome, where this cell type is often infected by the opportunistic pathogen human cytomegalovirus (HCMV). Alveolar macrophages exposed to both HCMV and HIV-1 consistently exhibited higher levels of HIV-1 replication than cells exposed to HIV-1 alone. HIV-1 production was strongly suppressed in alveolar macrophages treated with IL-13 regardless of whether or not the cultures were coinfected with HCMV. However, IL-13 treatment markedly enhanced the expression of HCMV in otherwise latently infected macrophages in a dose dependent manner. These unexpected differential effects of IL-13 on host-virus interactions are important considerations in guiding its potential therapeutic applications.

AIDS-Related Opportunistic Infections↗

PubMed1997

Cytokine signaling through the novel tyrosine kinase RAFTK in Kaposi's sarcoma cells.

A number of cytokines, including basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), oncostatin M (OSM), IL-6, and tumor necrosis factor alpha (TNF-alpha), have been postulated to have a role in the pathogenesis of Kaposi's sarcoma (KS). The proliferative effects of bFGF and OSM may be via their reported activation of the c-Jun NH2-terminal kinase (JNK) signaling pathway in KS cells. We now report that KS cells express a recently identified focal adhesion kinase termed RAFTK which appears in other cell systems to coordinate surface signals between cytokine and integrin receptors and the cytoskeleton as well as act downstream to modulate JNK activation. We also report that the tyrosine kinase receptor FLT-4, present on normal lymphatic endothelium, is robustly expressed in KS cells. Treatment of KS cells with VEGF-related protein (VRP), the ligand for the FLT-4 receptor, as well as with the cytokines bFGF, OSM, IL-6, VEGF, or TNF-alpha resulted in phosphorylation and activation of RAFTK. Following its activation, there was an enhanced association of RAFTK with the cytoskeletal protein paxillin. This association was mediated by the hydrophobic COOH-terminal domain of the kinase. Furthermore, JNK activity was increased in KS cells after VEGF or VRP stimulation. We postulate that in these tumor cells RAFTK may be activated by a diverse group of stimulatory cytokines and facilitate signal transduction to the cytoskeleton and downstream to the growth promoting JNK pathway.

Animals↗

PubMed1997

RAFTK, a novel member of the focal adhesion kinase family, is phosphorylated and associates with signaling molecules upon activation of mature T lymphocytes.

The related adhesion focal tyrosine kinase (RAFTK), a recently discovered member of the focal adhesion kinase family, has previously been reported to participate in signal transduction in neuronal cells, megakaryocytes, and B lymphocytes. We have found that RAFTK is constitutively expressed in human T cells and is rapidly phosphorylated upon the activation of the T cell receptor (TCR). This activation also results in an increase in the autophosphorylation and kinase activity of RAFTK. After its stimulation, there was an increase in the association of the src cytoplasmic tyrosine kinase Fyn and the adapter protein Grb2. This association was mediated through the SH2 domains of Fyn and Grb2. RAFTK also co-immunoprecipitates with the SH2 domain of Lck and with the cytoskeletal protein paxillin through its COOH-terminal proline-rich domain. The tyrosine phosphorylation of RAFTK after T cell receptor-mediated stimulation was reduced by the pretreatment of cells with cytochalasin D, suggesting the role of the cytoskeleton in this process. These observations indicate that RAFTK participates in T cell receptor signaling and may act to link signals from the cell surface to the cytoskeleton and thereby affect the host immune response.

Adaptor Proteins, Signal Transducing↗

PubMed1997

Association of csk-homologous kinase (CHK) (formerly MATK) with HER-2/ErbB-2 in breast cancer cells.

Protein-tyrosine kinases, such as HER-2/ErbB-2, have been specifically linked to breast cancer. The Csk-homologous kinase (CHK), formerly MATK, is a tyrosine kinase that contains the Src homology 2 and 3 (SH2 and SH3) domains and demonstrates homology ( approximately 50%) to the Csk tyrosine kinase. Like Csk, CHK is able to phosphorylate and inactivate Src family kinases. In this report, we investigated whether CHK is expressed in breast cancer tissues and whether it participates in the ErbB-2 signaling pathway in T47D and MCF-7 breast cancer cell lines. Immunostaining of the CHK protein in breast tissues demonstrated that primary invasive ductal carcinomas, stage II (13 of 15 cases) and stage I (8 of 15 cases), expressed the CHK protein, while this protein was not detected in the adjacent normal tissues from the same patients. To study the role of CHK in the ErbB-2 signaling pathway, glutathione S-transferase fusion proteins containing the SH2 and SH3 domains of CHK were generated. CHK-SH2 and CHK-SH3-SH2, but not CHK-SH3 or CHK-NH2-SH3, precipitated the tyrosine-phosphorylated ErbB-2 upon stimulation with heregulin. EGF or interleukin-6 stimulation of T47D cells failed to induce CHK-SH2 association with ErbB-2, the EGF-receptor, or the interleukin-6 receptor. In vivo association of the tyrosine-phosphorylated ErbB-2 with CHK was observed in co-immunoprecipitation studies with anti-CHK antibodies. EGF-R, ErbB-3, and ErbB-4 were not detected in the CHK immunoprecipitates or in the precipitates of the GST-SH2 fusion proteins of CHK, suggesting that the association of CHK with ErbB-2 upon heregulin stimulation is receptor-specific (ErbB-2) and ligand-specific (heregulin). These results indicate that CHK might participate in signaling in breast cancer cells by associating, via its SH2 domain, with ErbB-2 following heregulin stimulation.

Glycoproteins↗

PubMed1997

Transfusion in HIV disease.

Cytopenias often accompany infection with HIV. The pathophysiology responsible for the anemia, neutropenia, and thrombocytopenia in AIDS is complex and incompletely understood. Although HIV-infected patients with advanced disease were once routinely transfused for anemia accompanying zidovudine therapy, this practice has decreased as a result of decreased doses of zidovudine now being used and the more frequent use of HIV protease inhibitors. A more thorough understanding of the adverse effects of transfusion and the potential of immune modulation and facilitation of viral replication has no doubt led physicians to consider more thoroughly other factors before transfusing their patient. The results of trials examining the effect of leukodepletion on transfusion-related immune modulation are eagerly awaited. The availability of hematopoietic growth factors has also led to more effective treatment of the cytopenias without significant complications.

AIDS-Related Opportunistic Infections↗

PubMed1997

Potent inhibition of human immunodeficiency virus type 1 in primary T cells and alveolar macrophages by a combination anti-Rev strategy delivered in an adeno-associated virus vector.

The rate of viral replication appears to play a pivotal role in human immunodeficiency virus type 1 (HIV-1) pathogenesis and disease progression as it outstrips the capacity of the immune system to respond. Important cellular sites for HIV-1 production include T lymphocytes and tissue macrophages. Antiviral strategies, including newer treatment modalities such as gene therapy of HIV-1-susceptible cell populations, must be capable of engendering durable inhibitory effects to HIV-1 replication in both of these primary cell types in order to be effective. Among the potential genetic targets for intervention in the HIV-1 life cycle, the Rev regulatory system, consisting of Rev and its binding site, the Rev-responsive element (RRE), stands out as particularly attractive. Rev is essential for maintaining the stability of the viral genomic RNA as well as viral mRNAs encoding key structural and regulatory proteins. Moreover, it exhibits favorable threshold kinetics, in that Rev concentrations must rise above a critical level to exert their effect. To disable Rev function, primary T cells or macrophages were transduced with anti-Rev single-chain immunoglobulin (SFv) or RRE decoy genes either singly or in combination by employing adeno-associated virus vectors and then challenged with HIV-1. By directing both a protein and a nucleic acid against the normal interaction between Rev and the RRE, this genetic antiviral strategy effectively inhibited infection by either clinical or laboratory virus isolates. These results provide a framework for novel interventions to reduce virus production in the infected host.

Acquired Immunodeficiency Syndrome↗

PubMed1997

Patient, caregiver, and oncologist perceptions of cancer-related fatigue: results of a tripart assessment survey. The Fatigue Coalition.

Although fatigue is the most common symptom reported by cancer patients and has serious adverse effects on quality of life, it remains poorly understood. A survey was designed to characterize the epidemiology of cancer-related fatigue from the perspectives of the patient, primary caregiver, and oncologist. A telephone survey included 419 cancer patients recruited from 100,000 randomly selected households nationwide. Patients provided access to 200 primary caregivers (usually family members) who were also interviewed by telephone. In a separate mail survey, 197 of 600 randomly sampled oncologists (unrelated to the patients) responded to a questionnaire that assessed perceptions and attitudes concerning fatigue in cancer patients who had received chemotherapy or radiotherapy and their caregivers. The median patient age was 65 years, and the principal cancer diagnoses were breast (females) and genitourinary (males). Fifty-nine percent of the patients had received chemotherapy, 63% radiation therapy, and 24% both; 20% of patients received their last treatment within 6 weeks, 31% within 7 weeks to 1 year, and 49% more than 1 year ago. More than three quarters of patients (78%) experienced fatigue (defined as a general feeling of debilitating tiredness or loss of energy) during the course of their disease and treatment. Thirty-two percent experienced fatigue daily, and 32% reported fatigue significantly affected their daily routines. Caregivers reported observing fatigue in 86% of the index patients, and oncologists perceived that 76% of their patients experienced fatigue. Although oncologists believed that pain adversely affected their patients to a greater degree than fatigue (61% v 37%), patients felt that fatigue adversely affected their daily lives more than pain (61% v 19%). Most oncologists (80%) believed fatigue is overlooked or undertreated, and most patients (74%) considered fatigue a symptom to be endured. Fifty percent of patients did not discuss treatment options with their oncologists, and only 27% reported that their oncologists recommended any treatment for fatigue. When used, treatments for fatigue were generally perceived by patients and caregivers to be successful. These data confirm the high prevalence and adverse impact of cancer-related fatigue, although it is seldom discussed and infrequently treated. For patients and oncologists, improving the quality of life of cancer patients requires a heightened awareness of fatigue, a better understanding of its impact, and improve communication and familiarity with interventions that can reduce its debilitating effects.

Aged↗

PubMed1997

Impact of transfusion on viral load in human immunodeficiency virus infection.

Recent research has shown that anemia and transfusion are associated with accelerated mortality in patients with human immunodeficiency virus type 1 (HIV-1) infection. It appears that blood transfusions may directly accelerate HIV-1 disease progression through activation of HIV-1 expression and/or transfusion related immunosuppression. In vitro and in vivo evidence suggests that immunization with common recall antigens (eg, tetanus toxoid) can significantly increase plasma HIV viremia, as well as the in vitro susceptibility of peripheral blood mononuclear cells to acute infection. Allogeneic leukocytes also present an antigenic challenge to HIV-infected mononuclear cells, which may cause them to proliferate and increase viral production. In vitro incubation of HIV-infected mononuclear cells with allogeneic blood components (mainly leukocytes) results in increased viral replication. Whether the effects of transfusion on HIV viral load are due to a reduced immune response and/or activation of the virus itself remains to be determined. An ongoing study is evaluating the effects of depleting leukocytes from blood prior to transfusion in HIV-infected patients. Patients are randomized to receive standard of leukocyte-depleted red blood cell (RBC) transfusions and the effect of transfusion on HIV replication is measured. Preliminary data suggest that leukocyte depletion prior to RBC transfusion does not alter viral replication compared with standard packed RBC transfusions.

Adult↗

PubMed1995

Regulation of platelet activation in vitro by the c-Mpl ligand, thrombopoietin.

Thrombopoietin (TPO) is a recently identified growth factor that regulates megakaryocytopoiesis. Its receptor, c-Mpl, is expressed in megakaryocyte progenitors, mature megakaryocytes, and human blood platelets. We have observed that TPO treatment of human platelets resulted in tyrosine phosphorylation of several cellular proteins, including the c-Mpl receptor and the 85-kD subunit of phosphatidylinositol 3-kinase (PI3-K). TPO stimulated this tyrosine phosphorylation in a time-dependent manner, reaching a maximum in 5 minutes. The tyrosine phosphorylation of PI 3-K was dependent on the concentration of TPO and reached a maximum at concentrations between 50 and 100 ng/mL. This phosphorylation was independent of extracellular fibrinogen and ligation of the alpha IIb beta 3 integrin. In contrast, TPO, in the presence of exogenous fibrinogen, induced concentration-dependent platelet aggregation, which was blocked by the soluble c-Mpl receptor. Increasing TPO concentrations modulated the degree of the primary wave of aggregation and the lag phase, but not the slope or maximum of the secondary wave of aggregation. This secondary aggregation was controlled by the addition of apyrase, suggesting an adenosine diphosphate (ADP)-dependent mechanism. Treatment of platelets with TPO resulted in augmented binding of 125I-fibrinogen to intact platelets, with a 50% effect (EC50) occurring between 5 and 10 ng/mL. TPO-induced binding of fibrinogen to platelets was comparable in degree with that observed by stimulation with 10 mumol/L ADP. In an immobilized collagen-platelet adhesion assay, a significant increase in the attachment of TPO-stimulated platelets was observed. This effect was dependent on the concentration of TPO. At 50 ng/mL of TPO, platelet attachment to collagen increased threefold compared with the buffer control. Furthermore, the presence of fibrinogen did not significantly alter TPO augmentation of the platelet-collagen interaction. This interaction was mediated by the Arg-Gly-Asp (RGD) adhesion recognition sequence, as it was completely abolished by 100 mumol/L of the RGDS peptide. A fraction of the TPO-dependent platelet attachment to a collagen-coated surface was insensitive to treatment with prostaglandin E1. Furthermore, antibody to alpha IIb integrin partially inhibited platelet attachment to collagen, suggesting that the integrin alpha IIb beta 3 participates in this association. These data indicate that TPO might function not only as a cytokine in megakaryocyte growth and differentiation, but may also participate in direct platelet activation and modulate platelet-extracellular matrix interactions.

Adenosine Diphosphate↗