Thrombospondin-1 is an endogenous substrate of cereblon responsible for immunomodulatory drug-induced thromboembolism.

ABSTRACT: Immunomodulatory drugs (IMiDs) are key drugs for treating multiple myeloma and myelodysplastic syndrome with chromosome 5q deletion. IMiDs exert their pleiotropic effects through the interaction between cell-specific substrates and cereblon, a substrate receptor of the E3 ubiquitin ligase complex. Thus, identification of cell-specific substrates is important for understanding the effects of IMiDs. IMiDs increase the risk of thromboembolism, which sometimes results in fatal clinical outcomes. In this study, we sought to clarify the molecular mechanisms underlying IMiDs-induced thrombosis. We investigated cereblon substrates in human megakaryocytes using liquid chromatography-mass spectrometry and found that thrombospondin-1 (THBS-1), which is an inhibitor of a disintegrin-like and metalloproteinase with thrombospondin type 1 motifs 13, functions as an endogenous substrate in human megakaryocytes. IMiDs inhibited the proteasomal degradation of THBS-1 by impairing the recruitment of cereblon to THBS-1, leading to aberrant accumulation of THBS-1. We observed a significant increase in THBS-1 in peripheral blood mononuclear cells as well as larger von Willebrand factor multimers in the plasma of patients with myeloma, who were treated with IMiDs. These results collectively suggest that THBS-1 represents an endogenous substrate of cereblon. This pairing is disrupted by IMiDs, and the aberrant accumulation of THBS-1 plays an important role in the pathogenesis of IMiDs-induced thromboembolism.

Serum thrombospondin-2 level changes with liver stiffness improvement in patients with type 2 diabetes.

OBJECTIVE: Baseline circulating thrombospondin-2 (TSP2) level was identified as a potential novel hepatic fibrosis biomarker that associates with development and progression of hepatic fibrosis in patients with nonalcoholic fatty liver disease and type 2 diabetes. Here, we investigated whether circulating TSP2 levels changed with improvement in liver stiffness (LS), which reflects liver fibrosis on transient elastography. DESIGN: Serum TSP2 levels were measured in participants from a randomized, open-label intervention study, at baseline and after 24-weeks treatment of either dapagliflozin 10 mg (N = 30) or sitagliptin 100 mg daily (N = 30). Vibration-controlled transient elastography was performed to evaluate the severity of hepatic fibrosis and steatosis using LS and controlled attenuation parameter (CAP), respectively. PATIENTS AND MEASUREMENTS: Among all 60 participants with similar clinical characteristics at baseline (mean HbA1c 8.9%, CAP 289 dB/m and LS 5.8 kPa), despite similar HbA1c lowering, treatment with dapagliflozin, but not sitagliptin, led to significant improvements in body weight (BW) (p = .012), CAP (p = .015) and LS (p = .011) after 24 weeks. RESULTS: Serum TSP2 level decreased significantly from baseline in dapagliflozin-treated participants (p = .035), whereas no significant change was observed with sitagliptin. In correlation analysis, change in serum TSP2 levels only positively correlated with change in LS (r = .487, p = .006), but not with changes in BW, CAP or HbA1c after dapagliflozin treatment. CONCLUSIONS: Serum TSP2 level decreased with LS after dapagliflozin treatment, and was independent of improvements in BW, glycemic control and hepatic steatosis, further supporting the potential of serum TSP2 level as a novel hepatic fibrosis biomarker in type 2 diabetes.

Thrombospondin-1, CD47, and SIRPα display cell-specific molecular signatures in human islets and pancreata.

Thrombospondin-1 (TSP1) is a secreted protein minimally expressed in health but increased in disease and age. TSP1 binds to the cell membrane receptor CD47, which itself engages signal regulatory protein α (SIRPα), and the latter creates a checkpoint for immune activation. Individuals with cancer administered checkpoint-blocking molecules developed insulin-dependent diabetes. Relevant to this, CD47 blocking antibodies and SIRPα fusion proteins are in clinical trials. We characterized the molecular signature of TSP1, CD47, and SIRPα in human islets and pancreata. Fresh islets and pancreatic tissue from nondiabetic individuals were obtained. The expression of THBS1, CD47, and SIRPA was determined using single-cell mRNA sequencing, immunofluorescence microscopy, Western blot, and flow cytometry. Islets were exposed to diabetes-affiliated inflammatory cytokines and changes in protein expression were determined. CD47 mRNA was expressed in all islet cell types. THBS1 mRNA was restricted primarily to endothelial and mesenchymal cells, whereas SIRPA mRNA was found mostly in macrophages. Immunofluorescence staining showed CD47 protein expressed by ß cells and present in the exocrine pancreas. TSP1 and SIRPα proteins were not seen in islets or the exocrine pancreas. Western blot and flow cytometry confirmed immunofluorescent expression patterns. Importantly, human islets produced substantial quantities of secreted TSP1. Human pancreatic exocrine and endocrine tissue expressed CD47, whereas fresh islets displayed cell surface CD47 and secreted TSP1 at baseline and in inflammation. These findings suggest unexpected effects on islets from agents that intersect TSP1-CD47-SIRPα.NEW & NOTEWORTHY CD47 is a cell surface receptor with two primary ligands, soluble thrombospondin-1 (TSP1) and cell surface signal regulatory protein alpha (SIRPα). Both interactions provide checkpoints for immune cell activity. We determined that fresh human islets display CD47 and secrete TSP1. However, human islet endocrine cells lack SIRPα. These gene signatures are likely important given the increasing use of CD47 and SIRPα blocking molecules in individuals with cancer.

Serum thrombospondin-1 serves as a novel biomarker and agonist of gemcitabine-based chemotherapy in intrahepatic cholangiocarcinoma.

BACKGROUND: At present, the first-line treatment for advanced intrahepatic cholangiocarcinoma (ICC) is gemcitabine combined with cisplatin, but a considerable portion of ICC patients exhibit resistance to gemcitabine. Therefore, finding sensitisers for gemcitabine chemotherapy in ICC patients and predicting molecular markers for chemotherapy efficacy have become urgent needs. METHODS: In this study, PDX models were established to conduct gemcitabine susceptibility tests. The selected PDX tissues of the chemotherapy-sensitive group and drug-resistant group were subjected to transcriptome sequencing and protein chip technology to identify the key genes. Sixty-one ICC patients treated with gemcitabine chemotherapy were recruited for clinical follow-up validation. RESULTS: We found that thrombospondin-1 (TSP1) can predict gemcitabine chemosensitivity in ICC patients. The expression level of TSP1 could reflect the sensitivity of ICC patients to gemcitabine chemotherapy. Functional experiments further confirmed that TSP1 can increase the efficacy of gemcitabine chemotherapy for ICC. A mechanism study showed that TSP1 may affect the intake of oleic acid by binding to the CD36 receptor. CONCLUSIONS: In summary, we found a key molecule-TSP1-that can predict and improve the sensitivity of ICC patients to gemcitabine chemotherapy, which is of great significance for the treatment of advanced cholangiocarcinoma.

Thrombospondin 2 is a Functional Predictive and Prognostic Biomarker for Triple-Negative Breast Cancer Patients With Neoadjuvant Chemotherapy.

Background: Triple-negative breast cancer (TNBC) is characterized by a more aggressive biological behavior and unfavorable outcome. Circulating and histological expression of THBS2 has been demonstrated to be a novel diagnostic and prognostic biomarker in patients with various types of tumors. However, few studies have evaluated the predictive and prognostic value of THBS2 in TNBC specifically. Methods: In total, 185 triple-negative breast cancer patients (TNBC) with preoperative neoadjuvant chemotherapy were enrolled in this study. Serum THBS2 (sTHBS2) level was measured both prior to the start of NAC and at surgery by enzyme-linked immunosorbent assay (ELISA). Histological THBS2 (hTHBS2) expression in patients with residual tumors was evaluated by immunohistochemistry (IHC) staining method. Correlations between variables and treatment response were studied. Kaplan-Meier plots and Cox proportional hazard regression model were applied for survival analysis. Functional activities of THBS2 in TNBC cells were determined by CCK-8 assay, colony formation, wound healing, and transwell assay. Results: Of the 185 patients, 48 (25.9%) achieved pathological complete response (pCR) after completion of NAC. Elevated pCR rates were observed in patients with a lower level of sTHBS2 at surgery and higher level of sTHBS2 change (OR = 0.88, 95%CI: 0.79-0.98, p = 0.020 and OR = 1.12, 95%CI: 1.02-1.23, p = 0.015, respectively). In survival analysis, hTHBS2 expression in residual tumor was of independent prognostic value for both disease-free survival (HR = 2.21, 95%CI = 1.24-3.94, p = 0.007) and overall survival (HR = 2.07, 95%CI = 1.09-3.92, p = 0.026). For functional studies, THBS2 was indicated to inhibit proliferation, migration, and invasion abilities of TNBC cells in vitro. Conclusion: Our findings confirmed the value of serum THBS2 level to predict pCR for TNBC patients and the prognostic performance of histological THBS2 expression in non-pCR responders after NAC. THBS2 might serve as a promising functional biomarker for patients with triple-negative breast cancer.

Melatonin Prevents Chondrocyte Matrix Degradation in Rats with Experimentally Induced Osteoarthritis by Inhibiting Nuclear Factor-κB via SIRT1.

Osteoarthritis (OA) is a common degenerative joint disease characterized by an imbalance of cartilage extracellular matrix (ECM) breakdown and anabolism. Melatonin (MT) is one of the hormones secreted by the pineal gland of the brain and has anti-inflammatory, antioxidant, and anti-aging functions. To explore the role of MT in rats, we established an OA model in rats by anterior cruciate ligament transection (ACLT). Safranin O-fast green staining showed that intraperitoneal injection of MT (30 mg/kg) could alleviate the degeneration of articular cartilage in ACLT rats. Immunohistochemical (IHC) analysis found that MT could up-regulate the expression levels of collagen type II and Aggrecan and inhibit the expression levels of matrix metalloproteinase-3 (MMP-3), matrix metalloproteinase-13 (MMP-13), and ADAM metallopeptidase with thrombospondin type 1 motif 4 (ADAMTS-4) in ACLT rats. To elucidate the mechanism of MT in protecting the ECM in inflammatory factor-induced rat chondrocytes, we conducted in vitro experiments by co-culturing MT with a culture medium. Western blot (WB) showed that MT could promote the expression levels of transforming growth factor-beta 1 (TGF-ß1)/SMAD family member 2 (Smad2) and sirtuin 2-related enzyme 1 (SIRT1) and inhibit the expression of levels of phosphorylated nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibi-tor (p-p65) and phosphorylated IκB kinase-α (p-IκBα). In addition, WB and real-time PCR (qRT-PCR) results showed that MT could inhibit the expression levels of MMP-3, MMP-13, ADAMTS-4, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) in chondrocytes induced by interleukin-1ß (IL-1ß), and up-regulate the expression of chondroprotective protein type II collagen. We found that in vivo, MT treatment protected articular cartilage in the rat ACLT model. In IL-1ß-induced rat chondrocytes, MT could reduce chondrocyte matrix degradation by up-regulating nuclear factor-kB (NF-κB) signaling pathway-dependent expression of SIRT1 and protecting chondrocyte by activating the TGF-ß1/Smad2 pathway.

Salinomycin alleviates osteoarthritis progression via inhibiting Wnt/ß-catenin signaling.

Osteoarthritis (OA) is the most prevalent degenerative whole-joint disease characterized by cartilage degeneration, synovial hyperplasia, osteophyte formation, and subchondral bone sclerosis. Currently there are no disease-modifying treatments available for OA because its etiology and pathogenesis are largely unknown. Here we report that a natural carboxylic polyether ionophore that is used as an anti-tumor drug, salinomycin (SAL), may be a promising therapeutic drug for OA in the future. We found that SAL showed no cytotoxicity on mouse chondrocytes and displayed a protective effect against interleukin-1ß (IL-1ß), in cultured mouse chondrocytes and cartilage explants. Treatment with low SAL concentrations directly upregulated the anabolism factors collagen II and aggrecan, while it inhibited the catabolic factors matrix metalloproteinase-13 (MMP13) and metalloproteinase with thrombospondin motifs-5 (ADAMTS5) to protect against extracellular matrix (ECM) degradation, and also suppressed inflammatory responses in mouse chondrocytes. Furthermore, SAL reduced the severity of OA-associated changes and delayed cartilage destruction, subchondral bone sclerosis, and osteophyte formation in a destabilized medial meniscus (DMM) surgery-induced mouse OA model. Mechanistically, a low SAL concentration induced anabolism and inhibited catabolism in chondrocytes via inhibiting Lrp6 phosphorylation and Wnt/ß-catenin signaling. Our results suggested that SAL may serve as a potential disease-modifying therapeutic against OA pathogenesis.

Polydatin inhibits IL-1ß-mediated chondrocyte inflammation and ameliorates cartilage degradation: Involvement of the NF-κB and Wnt/ß-catenin pathways.

Osteoarthritis (OA) is a highly prevalent chronic joint disease that involves extracellular matrix (ECM) degradation and articular cartilage inflammation. Polydatin (PD) can alleviate inflammatory reactions in numerous diseases. The present study aimed to investigate the chondroprotective and anti-inflammatory effects of PD on interleukin (IL)- 1ß-treated chondrocytes in vitro and anterior cruciate ligament transection-induced rat OA models in vivo. Primary chondrocytes were isolated from SD rats and cultured. Only second-passage cells were used for subsequent experiments. Counting kit-8, quantitative real-time polymerase chain reaction, western blotting, enzyme-linked immunosorbent assay, and immunofluorescence were used to detect relevant indices. Rat OA models were established to obtain in vivo data. PD treatment decreased the production of nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α), and IL-6 during IL-1ß-stimulated chondrocyte inflammation. Moreover, PD upregulated aggrecan and collagen II expression, whereas downregulated a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) and matrix metalloproteinase-13 (MMP-13) expression on IL-1ß-mediated chondrocytes. Additionally, PD reduced IL-1ß-stimulated NF-κB and Wnt/ß-catenin activation and nuclear translocation. The results of histological analysis and scoring revealed that OA in the rat models was effectively ameliorated by the intra-articular injection of PD. PD suppressed IL-1ß-stimulated iNOS, COX-2, NO, and PGE2 production, TNF-α, IL-6, collagen X, MMP-13, and ADAMTS-5 expression, collagen II and aggrecan degeneration by inhibiting NF-κB and Wnt/ß-catenin signaling in vitro. PD also mitigated OA progression in the rat models, thereby providing reliable data that PD could serve as a promising candidate for OA therapy.

Tissue-targeted R-spondin mimetics for liver regeneration.

R-spondin (RSPO) proteins amplify Wnt signaling and stimulate regeneration in a variety of tissues. To repair tissue in a tissue-specific manner, tissue-targeted RSPO mimetic molecules are desired. Here, we mutated RSPO (RSPO2 F105R/F109A) to eliminate LGR binding while preserving ZNRF3/RNF43 binding and targeted the mutated RSPO to a liver specific receptor, ASGR1. The resulting bi-specific molecule (αASGR1-RSPO2-RA) enhanced Wnt signaling effectively in vitro, and its activity was limited to ASGR1 expressing cells. Systemic administration of αASGR1-RSPO2-RA in mice specifically upregulated Wnt target genes and stimulated cell proliferation in liver but not intestine (which is more responsive to non-targeted RSPO2) in healthy mice, and improved liver function in diseased mice. These results not only suggest that a tissue-specific RSPO mimetic protein can stimulate regeneration in a cell-specific manner, but also provide a blueprint of how a tissue-specific molecule might be constructed for applications in a broader context.

Therapeutic Role for TSP-2 Antibody in a Murine Asthma Model.

BACKGROUND: Specific immunotherapy, including agonists for Toll-like receptor 2 (TLR2), have been shown to protect from allergies and to have a high immunomodulatory capacity. METHODS: A new antibody, TSP-2, reactive against an epitope of the extracellular domain of TLR2, was identified. The effect of the antibody on dendritic cells was assessed by immunohistochemistry, Western blot, and flow cytometric analysis. The effect of TSP-2 in a murine asthma model induced with ovalbumin (OVA) was assessed. The model is a form of airway hyperresponsiveness (AHR) and was analyzed by whole-body plethysmography, the measurement of Th1/Th2 cytokines in bronchial alveolar lavage fluid (BALF) and serum by ELISA, and the CCK-8 assay for lymphocyte proliferation. The effect of TSP-2 on the maturation of bone marrow-derived dendritic cells (BMDCs) was assessed by flow cytometric analysis. RESULTS: TSP-2 promoted the maturation of dendritic cells and the proliferation of lymphocyte in vitro and in vivo. The effect of TSP-2 on T helper 1 (Th1)/Th2 cytokine secretion was slightly more powerful than that of Pam3CSK4. TSP-2 antibody reduced AHR and OVA-specific IgE levels in allergic asthma. TSP-2 antibody also reduced lung inflammation and decreased leukocyte numbers in an OVA-sensitized and challenged asthma model. TSP-2 antibody increased OVA-stimulated I-A, CD80, CD86, and MHC-II levels on BMDCs. CONCLUSIONS: This study identifies a novel therapeutic strategy for AHR, which uses antibodies reactive against TLR2. It also provides theoretical evidence for the control of allergic asthma by targeting TLR2.

R-Spondin1 expands Paneth cells and prevents dysbiosis induced by graft-versus-host disease.

The intestinal microbial ecosystem is actively regulated by Paneth cell-derived antimicrobial peptides such as α-defensins. Various disorders, including graft-versus-host disease (GVHD), disrupt Paneth cell functions, resulting in unfavorably altered intestinal microbiota (dysbiosis), which further accelerates the underlying diseases. Current strategies to restore the gut ecosystem are bacteriotherapy such as fecal microbiota transplantation and probiotics, and no physiological approach has been developed so far. In this study, we demonstrate a novel approach to restore gut microbial ecology by Wnt agonist R-Spondin1 (R-Spo1) or recombinant α-defensin in mice. R-Spo1 stimulates intestinal stem cells to differentiate to Paneth cells and enhances luminal secretion of α-defensins. Administration of R-Spo1 or recombinant α-defensin prevents GVHD-mediated dysbiosis, thus representing a novel and physiological approach at modifying the gut ecosystem to restore intestinal homeostasis and host-microbiota cross talk toward therapeutic benefits.

Management of Mucositis During Chemotherapy: From Pathophysiology to Pragmatic Therapeutics.

Chemotherapy-induced mucositis is a common condition caused by the breakdown of the mucosal barrier. Symptoms can include pain, vomiting and diarrhoea, which can often necessitate chemotherapy treatment breaks or dose reductions, thus compromising survival outcomes. Despite the significant impact of mucositis, there are currently limited clinically effective pharmacological therapies for the pathology. New emerging areas of research have been proposed to play key roles in the development of mucositis, providing rationale for potential new therapeutics for the prevention, treatment or management of chemotherapy-induced mucositis. This review aims to address these new areas of research and to comment on the therapeutics arising from them.

Thrombospondin promoted anti-tumor of adenovirus-mediated calreticulin in breast cancer: Relationship with anti-CD47.

OBJECTIVE: Calreticulin (CRT) protein has multifaceted role in carcinogenesis, however its role in breast cancer remains unidentified. In this study, we attempted to evaluate the effect of overexpressed CRT on breast cancer cells viability and proliferation. METHODS: Levels of mRNA and protein expression for CRT and CD47 in cells were determined by Quantitative RT-PCR analysis and Western blot, respectively. Cells apoptosis was evaluated using Annexin V-FITC assay with flow cytometry. Cell viability was assessed using MTT assay. Cell migration and autophagy were also evaluated. RESULTS: In breast cancer cells of MCF-7 and MDA-MB-231, both CRT and CD47 expression were enhanced, compared with that in normal breast cells of MCF-10A. Overexpression of CRT by MCF-7 and MDA-MB-231 cells transfected with significantly suppressed cell migration, viability as well as promote cell apoptosis while exerted no effected on cell autophagy. Interestingly, combining of thrombospondin (TSP) and overexpression of CRT significantly induced cell autophagy and inhibited tumor growth in MCF-7 cells xenograft. In result of chip assay, we observed that TSP treatment promoted interaction of TSP with CRT and CD47. CONCLUSION: TSP promoted anti-tumor of adenovirus-mediated CRT via forming complexes with CRT and CD47 in breast cancer.

Thrombospondin-2 secreted by human umbilical cord blood-derived mesenchymal stem cells promotes chondrogenic differentiation.

Increasing evidence indicates that the secretome of mesenchymal stem cells (MSCs) has therapeutic potential for the treatment of various diseases, including cartilage disorders. However, the paracrine mechanisms underlying cartilage repair by MSCs are poorly understood. Here, we show that human umbilical cord blood-derived MSCs (hUCB-MSCs) promoted differentiation of chondroprogenitor cells by paracrine action. This paracrine effect of hUCB-MSCs on chondroprogenitor cells was increased by treatment with synovial fluid (SF) obtained from osteoarthritis (OA) patients but was decreased by SF of fracture patients, compared to that of an untreated group. To identify paracrine factors underlying the chondrogenic effect of hUCB-MSCs, the secretomes of hUCB-MSCs stimulated by OA SF or fracture SF were analyzed using a biotin label-based antibody array. Among the proteins increased in response to these two kinds of SF, thrombospondin-2 (TSP-2) was specifically increased in only OA SF-treated hUCB-MSCs. In order to determine the role of TSP-2, exogenous TSP-2 was added to a micromass culture of chondroprogenitor cells. We found that TSP-2 had chondrogenic effects on chondroprogenitor cells via PKCα, ERK, p38/MAPK, and Notch signaling pathways. Knockdown of TSP-2 expression on hUCB-MSCs using small interfering RNA abolished the chondrogenic effects of hUCB-MSCs on chondroprogenitor cells. In parallel with in vitro analysis, the cartilage regenerating effect of hUCB-MSCs and TSP-2 was also demonstrated using a rabbit full-thickness osteochondral-defect model. Our findings suggested that hUCB-MSCs can stimulate the differentiation of locally presented endogenous chondroprogenitor cells by TSP-2, which finally leads to cartilage regeneration.

Improved therapeutic responses for liposomal doxorubicin targeted via thrombospondin peptidomimetics versus untargeted doxorubicin.

New therapies in cancer treatment are focusing on multifaceted approaches to starve and kill tumors utilizing both antiangiogenic and chemotherapeutic compounds. In this work, we searched for a peptide vector that would home liposomes both to endothelial and tumor cells. [Abu6]TSPB and [Abu6]TSPA, aspartimide analogs of natural sequences of TSP-1 and TSP-2, respectively, were tested for adhesion of tumor and endothelial cells, in vivo and in vitro antiangiogenic effects, and in vivo antitumor action. Both peptides support the adhesion of both types of cells, but only [Abu6]TSPA inhibits the angiogenesis in vivo, and [Abu6]TSPA-targeted L-DOX decreases by 58% (P < 0.008) the HT29 tumor growth in nude mice. The improvement in the doxorubicin antitumor effect should be attributed to the antiangiogenic effect of [Abu6]TSPA, since [Abu6]TSPB, despite being a good ligand for both cell types, had no effect on tumor growth.

Thrombospondin and apoptosis: molecular mechanisms and use for design of complementation treatments.

Thrombospondin-1 is the first and most studied naturally occurring protein inhibitor of angiogenesis. Its characteristic multi-domain structure determines thrombospondin-1 divergent functions, which include but are not limited to the regulation of angiogenesis. Below we overview the structural determinants and receptors expressed on the endothelial and other cell types, that are at the root of thrombospondin-1 striking ability to block neovascularization. We specifically emphasize thrombospondin-1 direct apoptotic action on the remodeling vascular endothelium and summarize current knowledge of its pro-apoptotic signaling and transcriptional networks. Further, we provide comprehensive survey of the thrombospondin-based anti-angiogenic strategies with special focus on the combination treatments. We convincingly illustrate how precise knowledge of the pro-apoptotic events and intermediates elicited by thrombospondin in the vascular endothelial cells facilitates the design of the most effective treatment combinations, where the efficacy of thrombospondin-derived compounds is maximized by the partner drug(s) ("complementation" strategies) and provide examples of such fine-tuning of the thrombospondin-based anti-angiogenic treatments.

Thrombospondin-based antiangiogenic therapy.

Thrombospondins (TSPs) are a family of extracellular matrix proteins that regulate tissue genesis and remodeling. TSP-1 plays a pivotal role in the regulation of both physiological and pathological angiogenesis. The inhibitory effects of TSP-1 on angiogenesis have been established in numerous experimental models. Among other TSP members, TSP-2 has equivalent domain structure as TSP-1 and shares most functions of TSP-1. The mechanisms by which TSP-1 and -2 inhibit angiogenesis can be broadly characterized as direct effects on vascular endothelial cells and indirect effects on the various angiogenic regulators. The fact that TSP-1 and -2 are potent endogenous angiogenic inhibitors has prompted studies to explore their therapeutic applications, and detailed understanding of the mechanisms of action of TSP-1 and -2 has facilitated the design of therapeutic strategies to optimize these activities. The therapeutic effects can be achieved by up-regulation of endogenous TSPs, or by the delivery of recombinant proteins or synthetic peptides that contain sequences from the angiogenic domain of TSP-1. In this article, we review the progress in thrombospondin-based antiangiogenic therapy and discuss the perspectives on the significant challenges that remain.

R-spondin1, a novel intestinotrophic mitogen, ameliorates experimental colitis in mice.

BACKGROUND & AIMS: R-spondin 1 (Rspo1) is a novel epithelial mitogen that stimulates the growth of mucosa in both the small and large intestine. METHODS: We investigated the therapeutic potential of Rspo1 in ameliorating experimental colitis induced by dextran sulfate sodium (DSS) or trinitrobenzene sulfonic acid (TNBS) as well as nonsteroidal anti-inflammatory drug-induced colitis in interleukin (IL)-10-deficient mice. RESULTS: Therapeutic administration of recombinant Rspo1 protein reduced the loss of body weight, diarrhea, and rectal bleeding in a mouse model of acute or chronic DSS-induced colitis. Histologic evaluation revealed that Rspo1 improved mucosal integrity in both villus and/or crypt compartments in the small intestine and colon by stimulating crypt cell growth and mucosal regeneration in DSS-treated mice. Moreover, Rspo1 significantly reduced DSS-induced myeloperoxidase activity and inhibited the overproduction of proinflammatory cytokines, including tumor necrosis factor-alpha, IL-1alpha, IL-6, interferon-gamma, and granulocyte-macrophage colony-stimulating factor, in mouse intestinal tissue, indicating that Rspo1 may reduce DSS-induced inflammation by preserving the mucosal barrier function. Likewise, Rspo1 therapy also alleviated TNBS-induced interstitial inflammation and mucosal erosion in the mouse colon. Furthermore, Rspo1 substantially decreased the histopathologic severity of chronic enterocolitis by repairing crypt epithelium and simultaneously suppressing inflammatory infiltration in piroxicam-exposed IL-10(-/-) mice. Endogenous Rspo1 protein was localized to villus epithelium and crypt Paneth cells in mouse small intestine. CONCLUSIONS: Our results show that Rspo1 may be clinically useful in the therapeutic treatment of inflammatory bowel disease by stimulating crypt cell growth, accelerating mucosal regeneration, and restoring intestinal architecture.