• TSP-1
• angiogenesis
• cancer
• metalloproteases
• serine proteases
• tumor microenvironment

•

Comprehensive overview of different classes of ECM molecules in the HSC niche.

Hematopoietic stem cells (HSCs) are the life-long source of all types of blood cells. Their function is controlled by their direct microenvironment, the HSC niche in the bone marrow. Although the importance of the extracellular matrix (ECM) in the niche by orchestrating niche architecture and cellular function is widely acknowledged, it is still underexplored. In this review, we provide a comprehensive overview of the ECM in HSC niches. For this purpose, we first briefly outline HSC niche biology and then review the role of the different classes of ECM molecules in the niche one by one and how they are perceived by cells. Matrix remodeling and the emerging importance of biophysics in HSC niche function are discussed. Finally, the application of the current knowledge of ECM in the niche in form of artificial HSC niches for HSC expansion or targeted differentiation as well as drug testing is reviewed.

• Bone marrow
• Extracellular matrix
• Hematopoietic stem cell niche

Thrombospondin-4 (TSP-4) attracted renewed attention recently as a result of assignment of new functions to this matricellular protein in cardiovascular, muscular, and nervous systems. We have previously reported that TSP-4 promotes local vascular inflammation in a mouse atherosclerosis model. A common variant of TSP-4, P387-TSP-4, was associated with increased cardiovascular disease risk in human population studies. In a mouse atherosclerosis model, TSP-4 had profound effect on accumulation of macrophages in lesions, which prompted us to examine its effects on macrophages in more detail. We examined the effects of A387-TSP-4 and P387-TSP-4 on mouse macrophages in cell culture and in vivo in the model of LPS-induced peritonitis. In tissues and in cell culture, TSP-4 expression was associated with inflammation: TSP-4 expression was upregulated in peritoneal tissues in LPS-induced peritonitis, and pro-inflammatory signals, INFγ, GM-CSF, and LPS, induced TSP-4 expression in macrophages in vivo and in cell culture. Deficiency in TSP-4 in macrophages from Thbs4^−/− mice reduced the expression of pro-inflammatory macrophage markers, suggesting that TSP-4 facilitates macrophage differentiation into a pro-inflammatory phenotype. Expression of TSP-4, especially more active P387-TSP-4, was associated with higher cellular apoptosis. Cultured macrophages displayed increased adhesion to TSP-4 and reduced migration in presence of TSP-4, and these responses were further increased with P387 variant. We concluded that TSP-4 expression in macrophages increases their accumulation in tissues during the acute inflammatory process and supports macrophage differentiation into a pro-inflammatory phenotype. In a model of acute inflammation, TSP-4 supports pro-inflammatory macrophage apoptosis, a response that is closely related to their pro-inflammatory activity and release of pro-inflammatory signals. P387-TSP-4 was found to be the more active form of TSP-4 in all examined functions.

TGF-β is a multifunctional cytokine affecting many cell types and implicated in tissue remodeling processes. Due to its many functions and cell-specific effects, the consequences of TGF-β signaling are process-and stage-dependent, and it is not uncommon that TGF-β exerts distinct and sometimes opposing effects on a disease progression depending on the stage and on the pathological changes associated with the stage. The mechanisms underlying cell- and process-specific effects of TGF-β are poorly understood.

We are describing a novel pathway that mediates induction of angiogenesis in response to TGF-β1. We found that in endothelial cells (EC) TSP-4, a secreted extracellular matrix (ECM) protein is upregulated in response to TGF-β1 and mediates the effects of TGF-β1 on angiogenesis.

Upregulation of TSP-4 does not require the synthesis of new protein, is not caused by decreased secretion of TSP-4, and is mediated by activation of SMAD3. Using Thbs4^−/− mice and TSP-4 shRNA, we found that TSP-4 mediated pro-angiogenic functions on cultured EC and angiogenesis in vivo in response to TGF-β1. We observed ~ 3-fold increases in tumor mass and levels of angiogenesis markers in animals injected with TGF-β1, and these effects did not occur in Thbs4^−/− animals. Injections of an inhibitor of TGF-β1 signaling SB431542 also decreased the weights of tumors and cancer angiogenesis.

Our results from in vivo angiogenesis models and cultured EC document that TSP-4 mediates upregulation of angiogenesis by TGF-β1. Upregulation of pro-angiogenic TSP-4 and selective effects of TSP-4 on EC may contribute to stimulation of tumor growth by TGF-β despite the inhibition of cancer cell proliferation.

• Amino Acid Sequence
• Animals
• Colon/cytology
• Colon/metabolism
• Flow Cytometry
• Interstitial Cells of Cajal/metabolism
• Ion Channels/chemistry
• Ion Channels/metabolism
• Jejunum/cytology
• Jejunum/metabolism
• Mice
• Sequence Homology, Amino Acid
• Transcriptome

• R01 DK094886 NIDDK NIH HHS
• R01 DK103055 NIDDK NIH HHS
• National Institute of Diabetes and Digestive and Kidney Diseases

• angiogenesis
• extracellular matrix
• thrombospondin-4

• Cell biology
• Microgravity
• Profilin-1
• Random Positioning Machine
• Spaceflight
• Thyroid cancer cells

• Astrocytes
• Extra-cellular matrix
• Neurogenesis
• Rostral-migratory stream
• Thrombospondin 1

AIM: To identify genes associated with gastric precancerous lesions in Helicobacter pylori (H. pylori)-susceptible ethnic Malays.

METHODS: Twenty-three Malay subjects with H. pylori infection and gastric precancerous lesions identified during endoscopy were included as “cases”. Thirty-seven Malay subjects who were H. pylori negative and had no precancerous lesions were included as “controls”. Venous blood was collected for genotyping with Affymetrix 50K Xba1 kit. Genotypes with call rates < 90% for autosomal single nucleotide polymorphisms (SNPs) were excluded. For each precancerous lesion, associated SNPs were identified from Manhattan plots, and only SNPs with a χ²P value < 0.05 and Hardy Weinberg Equilibrium P value > 0.5 was considered as significant markers.

RESULTS: Of the 23 H. pylori-positive subjects recruited, one sample was excluded from further analysis due to a low genotyping call rate. Of the 22 H. pylori-positive samples, atrophic gastritis only was present in 50.0%, complete intestinal metaplasia was present in 18.25%, both incomplete intestinal metaplasia and dysplasia was present in 22.7%, and dysplasia only was present in 9.1%. SNPs rs9315542 (UFM1 gene), rs6878265 (THBS4 gene), rs1042194 (CYP2C19 gene) and rs10505799 (MGST1 gene) were significantly associated with atrophic gastritis, complete intestinal metaplasia, incomplete metaplasia with foci of dysplasia and dysplasia, respectively. Allele frequencies in “cases”vs“controls” for rs9315542, rs6878265, rs1042194 and rs10505799 were 0.4 vs 0.06, 0.6 vs 0.01, 0.6 vs 0.01 and 0.5 vs 0.02, respectively.

CONCLUSION: Genetic variants possibly related to gastric precancerous lesions in ethnic Malays susceptible to H. pylori infection were identified for testing in subsequent trials.

• Gastric precancerous lesions
• Gene polymorphisms
• Genome-wide association
• Helicobacter pylori
• Malays

• Adult
• Aged
• Asian People/genetics
• Case-Control Studies
• Chi-Square Distribution
• Female
• Gastritis, Atrophic/diagnosis
• Gastritis, Atrophic/ethnology
• Gastritis, Atrophic/genetics
• Gastritis, Atrophic/microbiology
• Gastroscopy
• Gene Frequency
• Genetic Predisposition to Disease
• Helicobacter Infections/complications
• Helicobacter Infections/diagnosis
• Helicobacter Infections/ethnology
• Helicobacter Infections/microbiology
• Helicobacter pylori/pathogenicity
• Humans
• Malaysia/epidemiology
• Male
• Metaplasia
• Middle Aged
• Phenotype
• Polymorphism, Single Nucleotide
• Precancerous Conditions/diagnosis
• Precancerous Conditions/ethnology
• Precancerous Conditions/genetics
• Precancerous Conditions/microbiology
• Predictive Value of Tests
• Risk Factors
• Stomach Neoplasms/diagnosis
• Stomach Neoplasms/ethnology
• Stomach Neoplasms/genetics
• Stomach Neoplasms/microbiology

• Amino Acid Sequence
• Animals
• Cell Adhesion Molecules/metabolism
• Cell Membrane/metabolism
• Cell Nucleus/metabolism
• Chromatography, Affinity/methods
• Humans
• Hyaluronan Receptors/metabolism
• Models, Molecular
• Molecular Sequence Data
• Multiprotein Complexes
• Peptides/chemistry
• Peptides/genetics
• Peptides/metabolism
• Protein Structure, Tertiary
• Signal Transduction/physiology
• Surface-Active Agents/metabolism
• Thrombospondins/chemistry
• Thrombospondins/genetics
• Thrombospondins/metabolism

• Canadian Institutes of Health Research

C21, the C-terminal peptide of thrombospondin-4, has growth promoting activity and was discovered as one of several erythropoietin-dependent endothelial proteins. C21 stimulates red cell formation in anemic mice and is a growth factor for CD34+ and CD36+ hematopoietic cells, skin fibroblasts and kidney epithelial cells. ROD1 has been identified as an intracellular mediator. Nothing is known about the existence of putative C21 receptors on plasma membranes of target cells.

We analyzed the nature of C21-binding proteins in cell lysates of skin fibroblasts using C21 affinity columns. The membrane receptor CD44 was identified as C21-binding protein by mass spectrometry. We were unable to demonstrate any direct involvement of CD44 on cell growth or the effect of C21 on cell proliferation. A soluble form of CD44 was synthesized in insect cells and purified from culture supernatants with a combination of PVDF filtration in the presence of ammonium sulphate and HPLC. Both osteopontin and hyaluronic acid competitively displaced Biotin-C21 binding to CD44. In a colony-forming assay using primitive CD133+ hematopoietic stem cells from cord blood, osteopontin and C21 had opposite effects and C21 reduced the inhibitory action of osteopontin.

CD44 is a C21-binding membrane protein. We could not demonstrate an involvement of CD44 in the proliferative action of C21. Nevertheless, based on the antagonism of C21 and osteopontin in hematopoietic precursors, we speculate that C21 could indirectly have a major impact on hematopoietic stem cell proliferation, by hindering osteopontin membrane binding at the level of the bone marrow niche.

• Amino Acid Sequence
• Animals
• Cloning, Molecular
• DNA, Complementary/genetics
• Gene Expression
• Gene Expression Profiling
• Molecular Sequence Data
• Pancreas/cytology
• Penaeidae/classification
• Penaeidae/genetics
• Phylogeny
• Protein Structure, Tertiary
• RNA, Messenger/genetics
• RNA, Messenger/metabolism
• Sequence Alignment
• Stomach/cytology
• Thrombospondins/chemistry
• Thrombospondins/genetics

• Animals
• Cattle
• Cells, Cultured
• Endothelial Cells/drug effects
• Endothelial Cells/metabolism
• Erythroid Cells/drug effects
• Erythroid Cells/metabolism
• Erythropoietin/pharmacology
• Humans
• Insulin-Like Growth Factor Binding Protein 3/antagonists & inhibitors
• Insulin-Like Growth Factor Binding Protein 3/metabolism
• Interleukin-3/antagonists & inhibitors
• Interleukin-3/metabolism
• Peptide Fragments/pharmacology
• Thrombospondin 1/metabolism
• Thrombospondin 1/pharmacology
• Thymidine/metabolism
• Umbilical Cord/drug effects
• Umbilical Cord/metabolism