Thrombospondin Receptor Research Articles

CD47 Receptor Globally Regulates Metabolic Pathways That Control Resistance to Ionizing Radiation

Modulating tissue responses to stress is an important therapeutic objective. Oxidative and genotoxic stresses caused by ionizing radiation are detrimental to healthy tissues but beneficial for treatment of cancer. CD47 is a signaling receptor for thrombospondin-1 and an attractive therapeutic target because blocking CD47 signaling protects normal tissues while sensitizing tumors to ionizing radiation. Here we utilized a metabolomic approach to define molecular mechanisms underlying this radioprotective activity. CD47-deficient cells and cd47-null mice exhibited global advantages in preserving metabolite levels after irradiation. Metabolic pathways required for controlling oxidative stress and mediating DNA repair were enhanced. Some cellular energetics pathways differed basally in CD47-deficient cells, and the global declines in the glycolytic and tricarboxylic acid cycle metabolites characteristic of normal cell and tissue responses to irradiation were prevented in the absence of CD47. Thus, CD47 mediates signaling from the extracellular matrix that coordinately regulates basal metabolism and cytoprotective responses to radiation injury.

Erratum to: Pick of the Year 2014

Erratum to: J. Cell Commun. Signal. DOI 10.1007/s12079-015-0274-x This manuscript was erroneously omitted from issue 1 of 2015. The publisher apologizes for this oversight and for any inconvenience that was caused. The introduction refers to the instructions that the invited authors received in the preparation of their contributions to the following Bits & Bytes manuscripts published in issue 1 of the Journal of Cell Communication and Signaling: Dopamine: an old target in a new therapy by Sushanta K. Banerjee (2015) How IGF-1 activates its receptor by Robert Baxter (2015) Heparan sulfate provides a mechanism to respond to FGFR2b and control regenerative expansion by Enrique Brandan (2015) Cancer resistance, high molecular weight hyaluronic acid, and longevity by Gary J. Fisher (2015) Stem cell quest by Alexandra Irvine (2015) Lovastatin rescues human and mice cartilage disorders by Satoshi Kubota (2015) Yin and Yang revisited: CCN3 as an anti-fibrotic therapeutic? By Andrew Leask (2015) Another dimension to the importance of the extracellular matrix in fibrosis by Laure Rittie (2015) Therapeutic targeting of the thrombospondin-1 receptor CD47 to treat liver cancer by David D. Roberts, Sukhbir Kaur and David R. Soto-Pantoja (2015)

Therapeutic targeting of the thrombospondin-1 receptor CD47 to treat liver cancer.

CD47 is a signaling receptor for the matricellular protein thrombospondin-1 and a counter-receptor for signal regulatory protein-α (SIRPα) on macrophages. Following its initial discovery in 1992 as a cell surface protein that is over-expressed by ovarian carcinoma, elevated CD47 expression has emerged as a negative prognostic factor for a variety of cancers. CD47 is also a potential therapeutic target based on the ability of CD47 blockade to cause regression of tumors in mice, and a humanized CD47 antibody has recently entered phase I clinical trials. CD47 blockade may control tumor growth by inhibiting thrombospondin-1 signaling or by preventing inhibitory SIRPα signaling in tumor-associated macrophages. A recent publication by Lee et al. (Hepatology 60:179-191, 2014) provides evidence that blocking CD47 signaling specifically depletes tumor-initiating stem cells in hepatocellular carcinoma and implicates cathepsin-S/protease-activated receptor-2 signaling in mediating this therapeutic response.

Protective effects of aerobic swimming training on high-fat diet induced nonalcoholic fatty liver disease: Regulation of lipid metabolism via PANDER-AKT pathway

This study aimed to investigate the mechanism by which aerobic swimming training prevents high-fat-diet-induced nonalcoholic fatty liver disease (NAFLD). Forty-two male C57BL/6 mice were randomized into normal-diet sedentary (ND; n = 8), ND exercised (n = 8), high-fat diet sedentary (HFD; n = 13), and HFD exercised groups (n = 13). After 2 weeks of training adaptation, the mice were subjected to an aerobic swimming protocol (60 min/day) 5 days/week for 10 weeks. The HFD group exhibited significantly higher mRNA levels of fatty acid transport-, lipogenesis-, and β-oxidation-associated gene expressions than the ND group. PANDER and FOXO1 expressions increased, whereas AKT expression decreased in the HFD group. The aerobic swimming program with the HFD reversed the effects of the HFD on the expressions of thrombospondin-1 receptor, liver fatty acid-binding protein, long-chain fatty-acid elongase-6, Fas cell surface death receptor, and stearoyl-coenzyme A desaturase-1, as well as PANDER, FOXO1, and AKT. In the HFD exercised group, PPARα and AOX expressions were much higher. Our findings suggest that aerobic swimming training can prevent NAFLD via the regulation of fatty acid transport-, lipogenesis-, and β-oxidation-associated genes. In addition, the benefits from aerobic swimming training were achieved partly through the PANDER-AKT-FOXO1 pathway.

Functional and molecular characterisation of EO771.LMB tumours, a new C57BL/6-mouse-derived model of spontaneously metastatic mammary cancer.

The translation of basic research into improved therapies for breast cancer patients requires relevant preclinical models that incorporate spontaneous metastasis. We have completed a functional and molecular characterisation of a new isogenic C57BL/6 mouse model of breast cancer metastasis, comparing and contrasting it with the established BALB/c 4T1 model. Metastatic EO771.LMB tumours were derived from poorly metastatic parental EO771 mammary tumours. Functional differences were evaluated using both in vitro assays and spontaneous metastasis assays in mice. Results were compared to non-metastatic 67NR and metastatic 4T1.2 tumours of the 4T1 model. Protein and transcript levels of markers of human breast cancer molecular subtypes were measured in the four tumour lines, as well as p53 (Tp53) tumour-suppressor gene status and responses to tamoxifen in vivo and in vitro. Array-based expression profiling of whole tumours identified genes and pathways that were deregulated in metastatic tumours. EO771.LMB cells metastasised spontaneously to lung in C57BL/6 mice and displayed increased invasive capacity compared with parental EO771. By immunohistochemical assessment, EO771 and EO771.LMB were basal-like, as was the 4T1.2 tumour, whereas 67NR had a luminal phenotype. Primary tumours from all lines were negative for progesterone receptor, Erb-b2/Neu and cytokeratin 5/6, but positive for epidermal growth factor receptor (EGFR). Only 67NR displayed nuclear estrogen receptor alpha (ERα) positivity. EO771 and EO771.LMB expressed mutant p53, whereas 67NR and 4T1.2 were p53-null. Integrated molecular analysis of both the EO771/EO771.LMB and 67NR/4T1.2 pairs indicated that upregulation of matrix metalloproteinase-3 (MMP-3), parathyroid hormone-like hormone (Pthlh) and S100 calcium binding protein A8 (S100a8) and downregulation of the thrombospondin receptor (Cd36) might be causally involved in metastatic dissemination of breast cancer.

Identification of Actionable Genomic Alterations Across Different Lymphoma Histologies Using a Comprehensive Next Generation Genomic Sequencing Clinical Assay

Background:Several retrospective series have characterized the genomic landscape in many lymphoma subtypes. However, as these studies used different sequencing methods, depth of coverage, and specimen source, the incidence of alterations in many lymphoma subtypes remain unknown. Even when common genomic alterations were identified within a lymphoma subtype, the incidence and spectrum of the alterations varied. In this study, we prospectively examined the incidence of genomic alterations across different lymphoma subtypes using the Foundation One Heme (FOH) assay.Methods:FOH testing was offered to patients in routine clinical practice. Formalin-fixed paraffin-embedded (FFPE) tissue, bone marrow aspirate or peripheral blood were examined using the FOH targeted sequencing assay. Hybridization capture from 405 cancer related genes and 31 genes commonly rearranged in cancer was applied to ≥ 50ng of DNA extracted from 49 tumor specimens and sequenced to high, uniform coverage. Genomic alterations, which include: base substitutions, small insertions and deletions (indels), rearrangements, and copy number alterations, were determined.Results:Specimens from 48 non-consecutive patients with lymphoma (5 new diagnoses, 42 relapsed) were prospectively examined. Represented subtypes include DLBCL (44%), FL (17%), CLL (10%), MCL (10%), MZL (8%), T cell lymphoma (4%), and Hodgkin lymphoma (2%). Median age of diagnosis was 48.1 years (range 29.5-80.6) with a male predominance (69%). The majority (75%) had advanced stage disease at diagnosis, with a median of 3 lines of therapy (range 1-12, n=44). Forty-nine specimens, (1 patient with 2 specimens) were collected from FFPE (70%), peripheral blood (22%) or bone marrow aspirate (8%). Reports resulted after a median 28 days (range 10-48 days) from the time specimens were shipped to Foundation Medicine.Across 48 patients, 90 distinct alterations were detected with a median of 5 alterations/patient (range 0-11). Genomic alterations were more commonly observed in relapsed aggressive lymphoma compared to relapsed indolent lymphoma (median 6.4 vs 4 alterations/patient, p<0.05) or in newly diagnosed patients. The most common alteration was various IgH translocations (46%) and alterations in cyclin dependent kinases (CDK) genes (CDKN2A, CDKN2B and CDKN2C) (31%). Alterations of p53 (23%) and MLL2 (21%) were also frequently observed. Several rare alterations not previously described in primary lymphomas were identified, including alterations in ATRX, CDH1, MSH6, pyruvate carboxylase, and thrombospondin receptor (CD36). Alterations associated with approved agents or clinical trials were found in 89% of patients and were more frequent in relapsed aggressive B-cell NHL (median 2/patient) versus relapsed indolent B-cell NHL (median 1/patient, p<0.02). Commonly identified actionable alterations included CDK alterations in DLBCL, MCL and CLL, and alterations in the PI3K oncogenic pathway in DLBCL, FL and MZL.Conclusion:Application of a comprehensive next generation genomic sequencing assay provides an opportunity to both describe the spectrum and compare the incidence of genetic alterations across different lymphoma subtypes. Preliminary data suggest the vast majority of patients have one or more genomic alterations linked to approved agents or clinical trials. Data collection is ongoing. While the true incidence of each genomic alteration is not defined, the dataset provides the frequency of genomic alterations in a clinically relevant population. Moreover, these data will facilitate design of clinical trials by providing the opportunity to select patients based on shared genomic alterations rather than lymphoma subtype. Table 1Clinical characteristics, N=48Age at diagnosis57.1 (29.5-80.6)N (% of patients)Sex Male Female33 (68.7%)16 (33.3%)Subtype DLBCL21 (43.8%)FL8 (16.7%)CLL5 (10.4%)MCL5 (10.4%)MZL4 (8.3%)T cell lymphoma2 (4.2%)Hodgkin lymphoma1 (2.1%)Other3 (6.3%)Stage at Diagnosis I/II III/IV Not recorded10 (20.8%)36 (75.0%)2 (4.2%)Biopsy sent At diagnosis At relapse6 (12.5%)42 (87.5%)Response to First line Therapy Near CR or CR PR SD PD Unknown22 (45.8%)9 (18.8%)5 (10.4%)3 (8.3%)8 (16.7%)Genomic Alterations with clinical value Potentially Prognostic Potentially Actionable Alterations previously not well described37 (77.0%)43 (89.6%)7 (14.6%) [Display omitted] DisclosuresMoskowitz:Seattle Genetics, Inc.: Consultancy, Research Funding; Genentech: Research Funding; Merck: Research Funding. Horwitz:Celgene: Consultancy, Research Funding; Millennium: Consultancy, Research Funding; Infinity: Research Funding; Kiowa]Kirin: Research Funding; Seattle Genetics: Consultancy, Research Funding; Spectrum: Research Funding; Amgen: Consultancy; Bristol]Myers Squibb,: Consultancy; Jannsen: Consultancy. Hamlin:Gilead, Spectrum, Seattle Genetics, Genentech: Consultancy; Spectrum, GSK, Jansen and Jansen/Pharmacyclics, Portola, Seattle Genetics: Research Funding. Matasar:Genentech: Consultancy; Spectrum: Consultancy. Miller:Foundation Medicine: Employment. Stephens:Foundation Medicine: Employment, Equity Ownership. He:Foundation Medicine: Employment. Younes:Novartis: Research Funding; J & J: Research Funding; Curis: Research Funding; Bayer; Bristol Meyer Squibb; Celgene; Incyte; Janssen R & D; Sanofi; Seattle Genetics; Takeda Millenium: Honoraria.

Intermittent whole-body vibration attenuates a reduction in the number of the capillaries in unloaded rat skeletal muscle.

BackgroundWhole-body vibration has been suggested for the prevention of muscle mass loss and muscle wasting as an attractive measure for disuse atrophy. This study examined the effects of daily intermittent whole-body vibration and weight bearing during hindlimb suspension on capillary number and muscle atrophy in rat skeletal muscles.MethodsSixty male Wistar rats were randomly divided into four groups: control (CONT), hindlimb suspension (HS), HS + weight bearing (WB), and HS + whole-body vibration (VIB) (n = 15 each). Hindlimb suspension was applied for 2 weeks in HS, HS + WB, and HS + VIB groups. During suspension, rats in HS + VIB group were placed daily on a vibrating whole-body vibration platform for 20 min. In HS + WB group, suspension was interrupted for 20 min/day, allowing weight bearing. Untreated rats were used as controls.ResultsSoleus muscle wet weights and muscle fiber cross-sectional areas (CSA) significantly decreased in HS, HS + WB, and HS + VIB groups compared with CONT group. Both muscle weights and CSA were significantly greater in HS + WB and HS + VIB groups compared with HS group. Capillary numbers (represented by capillary-to-muscle fiber ratio) were significantly smaller in all hindlimb suspension-treated groups compared with CONT group. However, a reduction in capillary number by unloading hindlimbs was partially prevented by whole-body vibration. These findings were supported by examining mRNA for angiogenic-related factors. Expression levels of a pro-angiogenic factor, vascular endothelial growth factor-A mRNA, were significantly lower in all hindlimb suspension-treated groups compared with CONT group. There were no differences among hindlimb suspension-treated groups. Expression levels of an anti-angiogenic factor, CD36 (receptor for thrombospondin-1) mRNA, were significantly higher in all hindlimb suspension-treated groups compared with CONT group. Among the hindlimb suspension-treated groups, expression of CD36 mRNA in HS + VIB group tended to be suppressed (less than half the HS group).ConclusionsOur results suggest that weight bearing with or without vibration is effective for disuse-derived disturbance by preventing muscle atrophy, and whole-body vibration exercise has an additional benefit of maintaining microcirculation of skeletal muscle.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2474-15-315) contains supplementary material, which is available to authorized users.

Thrombospondin-1 regulates bone homeostasis through effects on bone matrix integrity and nitric oxide signaling in osteoclasts.

Thrombospondin-1 (TSP1), an endogenous antiangiogenic, is a widely expressed secreted ligand with roles in migration, adhesion, and proliferation and is a target for new therapeutics. While TSP1 is present in the bone matrix and several TSP1 receptors play roles in bone biology, the role of TSP1 in bone remodeling has not been fully elucidated. Bone turnover is characterized by coordinated activity of bone-forming osteoblasts (OB) and bone-resorbing osteoclasts (OC). TSP1-/- mice had increased bone mass and increased cortical bone size and thickness compared to wild type (WT). However, despite increased size, TSP1-/- femurs showed less resistance to bending than expected, indicative of diminished bone quality and a bone material defect. Additionally, we found that TSP1 deficiency resulted in decreased OC activity in vivo and reduced OC differentiation. TSP1 was critical during early osteoclastogenesis, and TSP1 deficiency resulted in a substantial overexpression of inducible nitric oxide synthase (iNOS). Importantly, administration of a NOS inhibitor rescued the OC function defects of TSP1-/- mice in vivo. To investigate the role of bone-derived TSP1 in osteoclastogenesis, we found that WT pre-OCs had defective iNOS expression when cultured on TSP1-/- bone compared to WT bone, suggesting that TSP1 in bone plays a critical role in iNOS signaling during OC development. These data implicate a new role for TSP1 in bone homeostasis with roles in maintaining bone matrix integrity and regulating OC formation. It will be critical to monitor bone health of patients administered TSP1-pathway directed therapeutics in clinical use and under development.

CD47-dependent immunomodulatory and angiogenic activities of extracellular vesicles produced by T cells

Intercellular communication is critical for integrating complex signals in multicellular eukaryotes. Vascular endothelial cells and T lymphocytes closely interact during the recirculation and trans-endothelial migration of T cells. In addition to direct cell–cell contact, we show that T cell derived extracellular vesicles can interact with endothelial cells and modulate their cellular functions. Thrombospondin-1 and its receptor CD47 are expressed on exosomes/ectosomes derived from T cells, and these extracellular vesicles are internalized and modulate signaling in both T cells and endothelial cells. Extracellular vesicles released from cells expressing or lacking CD47 differentially regulate activation of T cells induced by engaging the T cell receptor. Similarly, T cell-derived extracellular vesicles modulate endothelial cell responses to vascular endothelial growth factor and tube formation in a CD47-dependent manner. Uptake of T cell derived extracellular vesicles by recipient endothelial cells globally alters gene expression in a CD47-dependent manner. CD47 also regulates the mRNA content of extracellular vesicles in a manner consistent with some of the resulting alterations in target endothelial cell gene expression. Therefore, the thrombospondin-1 receptor CD47 directly or indirectly regulates intercellular communication mediated by the transfer of extracellular vesicles between vascular cells.

Tribbles expression in cumulus cells is related to oocyte maturation and fatty acid metabolism.

BackgroundIn mammals, the Tribbles family includes widely expressed serine-threonine kinase-like proteins (TRIB1, TRIB2 and TRIB3) that are involved in multiple biological processes including cell proliferation and fatty acid (FA) metabolism. Our recent studies highlighted the importance of FA metabolism in cumulus cells (CC) during oocyte maturation in vertebrates and reported a higher TRIB1 expression in CC surrounding in vivo mature oocytes as compared to immature ooocytes in mice and cows. The objective was to investigate Tribbles expression patterns in bovine CC during in vitro maturation (IVM) and to examine their roles in the cumulus-oocyte complex.MethodsTribbles gene expression was analyzed in bovine and murine CC using quantitative RT-PCR. Proteins were detected using Western blot and intracellular localization was assessed by immunofluorescence. Bovine COCs were treated with etomoxir, an inhibitor of FA oxidation (FAO) which blocks CPT1 activity, during 6 h and 18 h IVM. Oocyte meiotic stage was assessed and expression of Tribbles and lipid metabolism genes was quantified in CC.Results and discussionTRIB1 and TRIB3 were more strongly expressed whereas TRIB2 was less expressed in CC surrounding the oocytes from preovulatory follicles than in CC of immature ones. In CC, Tribbles were located in the cytoplasm and nucleus; in mitotic cells TRIB2 and TRIB3 were detected in the spindle. In the oocyte, Tribbles proteins were detected in the ooplasm; also TRIB2 and TRIB3 were more accumulated in the germinal vesicle. In bovine CC, expression of TRIB1 and TRIB3 was transiently increased at a time preceding oocyte meiosis resumption in vitro. Treatment with etomoxir (150 μM) during IVM resulted in a significant reduction of oocyte maturation rate and decreased MAPK3/1 phosphorylation in the oocytes. In CC, 18 h IVM of etomoxir treatment significantly increased expression of TRIB1, TRIB3, CPTA1 (enzyme regulating FA entry in mitochondria for FAO) and CD36 (thrombospondin receptor involved in FA transport). Under the same conditions, expression of TRIB2 and ACACA (Acetyl coenzyme A carboxylase involved in FA synthesis) decreased in CC.All considered, Tribbles family members may be involved in cell proliferation and in FAO signaling in CC and participate in oocyte meiotic resumption regulation.

Alpha2delta‐1 Identified as a novel effector underlying the action of brain‐derived neurotrophic factor in hypothalamic control of feeding in mice (641.21)

Brain‐derived neurotrophic factor (BDNF), a protein that supports neuronal survival and synaptic plasticity, is essential for energy balance regulation. Genetic polymorphisms that impede BDNF function pose risk factors for obesity in humans. Similarly, blunted BDNF signaling in the rodent brain is associated with overeating (OV), body weight (BW) gain and metabolic dysfunction. Here we identified alpha2delta‐1 (α2δ‐1), a high voltage‐gated calcium channel subunit and thrombospondin receptor, as a novel appetite regulator underlying BDNF function in the ventromedial hypothalamus (VMH). α2δ‐1 is highly expressed in the VMH of wild‐type (WT) mice. Moreover, mice lacking central Bdnf (CM) have reduced cell surface α2δ‐1 expression in VMH cells. Selective inhibition of VMH α2δ‐1 in WT mice through chronic delivery of gabapentin resulted in a significant increase in food intake and BW gain after 7 days. Rescuing the α2δ‐1 deficit in VMH cells of CM, through targeted delivery of an adeno‐associated virus encoding α2δ‐1, mitigated the OV and BW gain and normalized glucose homeostasis. These data inform mechanisms underlying effects of BDNF in the VMH and identify a critical role for α2δ‐1 in feeding control.

The thrombospondin-1 receptor CD36 is an important mediator of ovarian angiogenesis and folliculogenesis.

BackgroundOvarian angiogenesis is a complex process that is regulated by a balance between pro- and anti-angiogenic factors. Physiological processes within the ovary, such as folliculogenesis, ovulation, and luteal formation are dependent upon adequate vascularization and anything that disrupts normal angiogenic processes may result in ovarian dysfunction, and possibly infertility. The objective of this study was to evaluate the role of the thrombospondin-1 (TSP-1) receptor CD36 in mediating ovarian angiogenesis and regulating ovarian function.MethodsThe role of CD36 was evaluated in granulosa cells in vitro and ovarian morphology and protein expression were determined in wild type and CD36 null mice.ResultsIn vitro, CD36 inhibition increased granulosa cell proliferation and decreased apoptosis. Granulosa cells in which CD36 was knocked down also exhibited an increase in expression of survival and angiogenic proteins. Ovaries from CD36 null mice were hypervascularized, with increased expression of pro-angiogenic vascular endothelial growth factor (VEGF) and its receptor VEGFR-2. Ovaries from CD36 null mice contained an increase in the numbers of pre-ovulatory follicles and decreased numbers of corpora lutea. CD36 null mice also had fewer number of offspring compared to wild type controls.ConclusionsThe results from this study demonstrate that CD36 is integral to the regulation of ovarian angiogenesis by TSP-1 and the expression of these family members may be useful in the control of ovarian vascular disorders.

Hypothalamic dysfunction of the thrombospondin receptor α2δ-1 underlies the overeating and obesity triggered by brain-derived neurotrophic factor deficiency.

Brain-derived neurotrophic factor (BDNF) and its receptor, TrkB, are critical components of the neural circuitry controlling appetite and body weight. Diminished BDNF signaling in mice results in severe hyperphagia and obesity. In humans, BDNF haploinsufficiency and the functional Bdnf Val66Met polymorphism have been linked to elevated food intake and body weight. The mechanisms underlying this dysfunction are poorly defined. We demonstrate a chief role of α2δ-1, a calcium channel subunit and thrombospondin receptor, in triggering overeating in mice with central BDNF depletion. We show reduced α2δ-1 cell-surface expression in the BDNF mutant ventromedial hypothalamus (VMH), an energy balance-regulating center. This deficit contributes to the hyperphagia exhibited by BDNF mutant mice because selective inhibition of α2δ-1 by gabapentin infusion into wild-type VMH significantly increases feeding and body weight gain. Importantly, viral-mediated α2δ-1 rescue in BDNF mutant VMH significantly mitigates their hyperphagia, obesity, and liver steatosis and normalizes deficits in glucose homeostasis. Whole-cell recordings in BDNF mutant VMH neurons revealed normal calcium currents but reduced frequency of EPSCs. These results suggest calcium channel-independent effects of α2δ-1 on feeding and implicate α2δ-1-thrombospondin interactions known to facilitate excitatory synapse assembly. Our findings identify a central mechanism mediating the inhibitory effects of BDNF on feeding. They also demonstrate a novel and critical role for α2δ-1 in appetite control and suggest a mechanism underlying weight gain in humans treated with gabapentinoid drugs.

Gabapentin Administration Reduces Reactive Gliosis and Neurodegeneration after Pilocarpine-Induced Status Epilepticus

The lithium-pilocarpine model of epilepsy reproduces in rodents several features of human temporal lobe epilepsy, by inducing an acute status epilepticus (SE) followed by a latency period. It has been proposed that the neuronal network reorganization that occurs during latency determines the subsequent appearance of spontaneous recurrent seizures. The aim of this study was to evaluate neuronal and glial responses during the latency period that follows SE. Given the potential role of astrocytes in the post-SE network reorganization, through the secretion of synaptogenic molecules such as thrombospondins, we also studied the effect of treatment with the α2δ1 thrombospondin receptor antagonist gabapentin. Adult male Wistar rats received 3 mEq/kg LiCl, and 20 h later 30 mg/kg pilocarpine. Once SE was achieved, seizures were stopped with 20 mg/kg diazepam. Animals then received 400 mg/kg/day gabapentin or saline for either 4 or 14 days. In vitro experiments were performed in dissociated mixed hippocampal cell culture exposed to glutamate, and subsequently treated with gabapentin or vehicle. During the latency period, the hippocampus and pyriform cortex of SE-animals presented a profuse reactive astrogliosis, with increased GFAP and nestin expression. Gliosis intensity was dependent on the Racine stage attained by the animals and peaked 15 days after SE. Microglia was also reactive after SE, and followed the same pattern. Neuronal degeneration was present in SE-animals, and also depended on the Racine stage and the SE duration. Polysialic-acid NCAM (PSA-NCAM) expression was increased in hippocampal CA-1 and dentate gyrus of SE-animals. Gabapentin treatment was able to reduce reactive gliosis, decrease neuronal loss and normalize PSA-NCAM staining in hippocampal CA-1. In vitro, gabapentin treatment partially prevented the dendritic loss and reactive gliosis caused by glutamate excitotoxicity. Our results show that gabapentin treatment during the latency period after SE protects neurons and normalizes PSA-NCAM probably by direct interaction with neurons and glia.

Thrombospondin-1/CD36 pathway contributes to bone marrow-derived angiogenic cell dysfunction in type 1 diabetes via Sonic hedgehog pathway suppression

Refractory wounds in diabetic patients present a significant clinical problem. Sonic hedgehog (SHH), a morphogenic protein central to wound repair, is deficient in diabetes. Regulation of SHH in wound healing is poorly understood. We hypothesize that thrombospondin-1 (TSP-1), through its receptor CD36, contributes to the SHH signaling defect in bone marrow-derived angiogenic cells (BMACs) in type 1 diabetic mice. Isolated BMACs from TSP-1-knockout mice demonstrated improved tube formation, migration, and adhesion in parallel with active SHH signaling. BMACs from STZ-induced type 1 diabetic mice showed significantly impaired Matrigel tube formation (n = 5; P < 0.05 vs. control), which was rescued by TSP-1 depletion (n = 5; P < 0.05 STZ-TSP-1(-/-) vs. STZ-WT) or exogenous SHH (20 mg/l, 24 h, n = 4; P < 0.05 vs. STZ-control). The expression of CD36 was elevated in BMACs from STZ mice (n = 4; P < 0.05). SHH signaling was significantly higher in BMACs from TSP-1(-/-) mice and TSP-1 receptor CD36-knockout mice (n = 6; P < 0.05 vs. WT) but not CD47-knockout mice (n = 3; P > 0.05 vs. WT). The impairment of recombinant human TSP-1 (2.2 nM, 24 h) on BMAC Matrigel tube formation was delayed significantly by CD36 deletion (n = 5; P < 0.05). CD36(-/-) BMACs demonstrated better tube formation under both normal and diabetic conditions with active SHH signaling (n = 4; P < 0.05 vs. WT BMACs). In conclusion, The TSP-1/CD36 pathway contributes to the SHH signaling defect, resulting in BMAC dysfunction in type 1 diabetic mice.

Electroconvulsive seizure induces thrombospondin-1 in the adult rat hippocampus

Synaptic dysfunction has recently gained attention for its involvement in mood disorders. Electroconvulsive therapy (ECT) possibly plays a role in synaptic repair. However, the underlying mechanisms remain uncertain. Thrombospondin-1 (TSP-1), a member of the TSP family, is reported to be secreted by astrocytes and to regulate synaptogenesis. We investigated the effects of electroconvulsive seizure (ECS) on the expression of TSPs in the adult rat hippocampus. Single and repeated ECS significantly increased TSP-1 mRNA expression after 2h and returned to sham levels at 24h. Conversely, the TSP-2 and -4 mRNA levels did not change. Only repeated ECS induced TSP-1 proteins. ECS also induced glial fibrillary acidic protein (GFAP) expression. The GFAP expression occurred later than the TSP-1 mRNA expression following single ECS; however, it occurred earlier and was more persistent following repeated ECS. ECS had no effect on the α2δ-1 or neuroligin-1 expressions, both of which are TSP-1 receptors. Furthermore, chronic treatment with antidepressants did not induce the expression of TSP-1 or GFAP. These findings suggest that repeated ECS, but not chronic treatment with antidepressants, induces TSP-1 expression partially via the activation of astrocytes. Therefore, TSP-1 is possibly involved in the synaptogenic effects of ECS.

Thrombospondin-1 Signaling through CD47 Inhibits Self-renewal by Regulating c-Myc and Other Stem Cell Transcription Factors

Signaling through the thrombospondin-1 receptor CD47 broadly limits cell and tissue survival of stress, but the molecular mechanisms are incompletely understood. We now show that loss of CD47 permits sustained proliferation of primary murine endothelial cells, increases asymmetric division, and enables these cells to spontaneously reprogram to form multipotent embryoid body-like clusters. c-Myc, Klf4, Oct4, and Sox2 expression is elevated in CD47-null endothelial cells, in several tissues of CD47- and thrombospondin-1-null mice, and in a human T cell line lacking CD47. CD47 knockdown acutely increases mRNA levels of c-Myc and other stem cell transcription factors in cells and in vivo, whereas CD47 ligation by thrombospondin-1 suppresses c-Myc expression. The inhibitory effects of increasing CD47 levels can be overcome by maintaining c-Myc expression and are absent in cells with dysregulated c-Myc. Thus, CD47 antagonists enable cell self-renewal and reprogramming by overcoming negative regulation of c-Myc and other stem cell transcription factors.

Chronic Delivery of a Thrombospondin-1 Mimetic Decreases Skeletal Muscle Capillarity in Mice

Angiogenesis is an essential process for normal skeletal muscle function. There is a growing body of evidence suggesting that thrombospondin-1 (TSP-1), a potent antiangiogenic protein in tumorigenesis, is an important regulator of both physiological and pathological skeletal muscle angiogenesis. We tested the hypothesis that chronic exposure to a TSP-1 mimetic (ABT-510), which targets the CD36 TSP-1 receptor, would decrease skeletal muscle capillarity as well as alter the balance between positive and negative angiogenic proteins under basal conditions. Osmotic minipumps with either ABT-510 or vehicle (5% dextrose) were implanted subcutaneously in the subscapular region of C57/BL6 mice for 14 days. When compared to the vehicle treated mice, the ABT-510 group had a 20% decrease in capillarity in the superficial region of the gastrocnemius (GA), 11% decrease in the plantaris (PLT), and a 35% decrease in the soleus (SOL). ABT-510 also decreased muscle protein expression of vascular endothelial growth factor (VEGF) in both the GA (−140%) and SOL (−62%); however there was no change in VEGF in the PLT. Serum VEGF was not altered in ABT-510 treated animals. Endogenous TSP-1 protein expression in all muscles remained unaltered. Tunnel staining revealed no difference in muscle apoptosis between ABT-510 and vehicle treated groups. These data provide evidence that the anti-angiogenic effects of TSP-1 are mediated, at least in part, via the CD36 receptor. It also suggests that under physiologic conditions the TSP-1/CD36 axis plays a role in regulating basal skeletal muscle microvessel density.

Thrombospondin and VEGF-R: Is There a Correlation in Inflammatory Bowel Disease?

Up to date several authors discussed interactions between cells forming inflammatory infiltrates in the course of inflammatory bowel disease (IBD), mainly dealing with endoscopic biopsy specimens. These usually contain only mucosa. We have evaluated full bowel wall sections, which seems to be especially important in patients with Crohn's disease (CD). The purpose of our study was to evaluate the relationship between vascular density and expression of thrombospondin-1 (TSP-1) and vascular endothelial growth factor receptor 1 (VEGFR-1) in full-thickness tissue fragments of intestinal wall taken from patients after colectomy, comparing those with IBD to non-IBD control group. Histological sections were immunostained with antibodies against CD-31, TSP-1, and VEGFR-1 and analyzed by pathologists with the use of computer-assisted morphometrics. Our research showed significantly higher vascular density and vascular area percentage in all layers of bowel wall in patients with CD when compared to control. We have also demonstrated differences in vascular density distribution between ulcerative colitis (CU) and CD and between CU and control. However we have not found statistically significant correlation between those findings and VEGFR-1 or TSP-1 expression. Our results might suggest existence of different, TSP-1 independent pathways of antiangiogenesis in IBD.

Clinical relevance of thrombospondin receptor (CD36) expression in Egyptian de novo adult acute myeloid leukemia

Background Acute myeloid leukemias (AMLs) are a heterogeneous group of disorders that often present with different morphological, immunophenotypic, and cytogenetic patterns. Identification of these characteristics may be useful for a better prognostic evaluation and for a more appropriate therapeutic approach. CD36 is a transmembrane, highly glycosylated, glycoprotein commonly expressed on blasts in acute monocytic leukemia, megakaryoblastic leukemia, and erythroleukemia. Patients and methods We evaluated CD36 surface expression in 97 newly diagnosed AML patients, and the results were correlated with the morphology, immunophenotype, cytogenetic pattern, and clinical outcome. Results CD36 antigen was recorded in 48 of 97 patients (49.5%) and particularly in those with M5 and M6 FAB subtypes. Moreover, CD36 expression was significantly associated with the expression of CD11b ( P =0.001) and CD14 ( P =0.0001), unfavorable cytogenetic abnormalities ( P =0.001), shorter overall survival ( P >0.0001), and leukemia-free survival ( P =0.03). Conclusion On the basis of the results of the study, it can be concluded that CD36 expression in AML patients may identify a subgroup with a poor prognosis, and may thus be a valuable adjunct to be added to the current prognostic factors.