• Biomolecules & Therapeutics
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• v.25 no.6
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• pp.625-633
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• 2017

Sphingosylphosphorylcholine (SPC) is one of the bioactive phospholipids that has many cellular functions such as cell migration, adhesion, proliferation, angiogenesis, and $Ca^{2+}$ signaling. Recent studies have reported that SPC induces invasion of breast cancer cells via matrix metalloproteinase-3 (MMP-3) secretion leading to WNT activation. Thrombospondin-1 (TSP-1) is a matricellular and calcium-binding protein that binds to a wide variety of integrin and non-integrin cell surface receptors. It regulates cell proliferation, migration, and apoptosis in inflammation, angiogenesis and neoplasia. TSP-1 promotes aggressive phenotype via epithelial mesenchymal transition (EMT). The relationship between SPC and TSP-1 is unclear. We found SPC induced EMT leading to mesenchymal morphology, decrease of E-cadherin expression and increases of N-cadherin and vimentin. SPC induced secretion of thrombospondin-1 (TSP-1) during SPC-induced EMT of various breast cancer cells. Gene silencing of TSP-1 suppressed SPC-induced EMT as well as migration and invasion of MCF10A cells. An extracellular signal-regulated kinase inhibitor, PD98059, significantly suppressed the secretion of TSP-1, expressions of N-cadherin and vimentin, and decrease of E-cadherin in MCF10A cells. ERK2 siRNA suppressed TSP-1 secretion and EMT. From online PROGgene V2, relapse free survival is low in patients having high TSP-1 expressed breast cancer. Taken together, we found that SPC induced EMT and TSP-1 secretion via ERK2 signaling pathway. These results suggests that SPC-induced TSP-1 might be a new target for suppression of metastasis of breast cancer cells.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.3
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• pp.253-259
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• 2005

Objective: We investigated the expression of TSP-1 and -2 in eutopic endometrium of advanced endometriosis and control patients. Methods: The 33 endometriosis patients and 32 controls were enrolled. Endometrial samples were obtained from 65 premenopausal women aged 29-44 years, undergoing laparoscopic surgery or hysterectomy for non-malignant lesions. Sufficient samples were collected from 33 patients with endometriosis stage III and IV and 32 controls without endometriosis confirmed by laparoscopic surgery. The mRNA expression from eutopic endometrium for TSP-1 and -2 were analyzed by RT-QC PCR. Results: The mRNAs of TSP-1 and -2 were expressed in eutopic endometrium from endometriosis and normal controls throughout the menstrual cycle. There were no significant differences in expression of TSP-1 and TSP-2 in eutopic endometrium between controls and endometriosis patients. Conclusion: Our results indicated that TSP-1 and -2 had no crucial role compared to other molecules in the regulation of angiogenesis. These findings also suggest that dysregulation of other angiogenic regulators would be concerned in pathophysiologic role in endometriosis development.

• Journal of Life Science
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• v.14 no.1
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• pp.154-162
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• 2004

Thrombospondin-1 (TSP-1), a negative regulator in tumor growth and angiogenesis, is cell-type specifically regulated and at transcriptional level by external stimuli. Previously, we found that phorbol 12-myristate 13-acetate (PMA) suppressed TSP-1 expression in porcine aortic endothelial (PAE) cell, but enhanced in hepatoma cell line, Hep 3B cell. A region between -767 and -723 on the tsp-1 promoter was defined as a responsive site to the suppression in PAE cell. eased on the previous results, the molecular mechanism of TSP-1 expression was determined by characterizing interactions between cis-elements and trans-factors using three overlapped oligonucleotide probes, oligo a-1 (from -767 to -738), a-2 (-759 to -730) and a-3 (-752 to -723). The results from electromobility shift assay showed that PMA-induced suppression of TSP-1 transcription in PAE cell might be caused via a negative regulator binding to the region from -752 to -730 and additionally generated by lacking two positive regulators binding to the sites from -767 to -760 and from -752 to -730. Especially, PMA enhanced the binding ability of the negative regulator to the site from -752 to -730 in PAE cell, but anti-c-Jun did not affected its binding ability.

• Journal of Life Science
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• v.15 no.6 s.73
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• pp.895-903
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• 2005

Thrombospondin-1 (TSP-1), a negative regulator in tumor growth and angiogenesis, is cell-type specifically regulated under pathological conditions or by extracellular stimuli, and the regulation of TSP-1 gene expression is important for developing new approaches in tumor therapy. Mistletoe is a parasitir plant that have been used for immunomodulation and antitumor therapy. Helixor A is an aqueous part of mistletoes extract. Here we showed that TSP-1 expression was significantly induced at both mRNA and protein levels in the Hepatocarcinorna cell line (Hep3B) and primary bovine endothelial cell line (BAE) exposed to Helixor A. Our promoter analysis confirmed that the expression of TSP-1 gene was regulated by Helixor A at the transcriptional level. In cell invasion assay, the conditioned media obtained from treatment of these cells significantly reduced the number of invasive cells and also inhibited capillary-like tube formation of BAE cells on Matrigel. Moreover, the inhibitory efforts of the conditioned media on cell invasion and tube formation were reversed by blocking with anti-TSP-1 neutralizing antibodies, suggesting that TSP-1 is involved in Helixor A-indured antiangiogenic effect. Taken together, our results suggest that Helixor A have an antiangiogenic effects through upregulation of TSP-1.

• YAKHAK HOEJI
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• v.53 no.3
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• pp.151-155
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• 2009

Meteorin in astrocytes has antiangiogenic activities via thrombospondin-1 (TSP-1), however, the regulatory mechanism has been unclear. Here we report that Meteorin upregulates TSP transcriptionally through luciferase reporter assays in astrocytes. Moreover, Meteorin activates $PKC\delta$ and ERK1/2 in astrocytes. Inhibition of $PKC\delta$ and ERK1/2 activities attenuated expression of TSP-1 by Meteorin in astrocytes. We, therefore, demonstrate that Meteorin activates $PKC\delta$ signaling and, in turn, increases TSP-1 expression in astrocytes to inhibit angiogenesis in the brain.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.8
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• pp.1366-1372
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• 2018

Objective: A disintegrin and metallopeptidase with thrombospondin motifs type 8 (ADAMTS8) is crucial for diverse physiological processes, such as inflammation, tissue morphogenesis, and tumorigenesis. The chicken ADAMTS8 (chADAMTS8) gene was differentially expressed in the kidney following exposure to different calcium concentrations, suggesting a pathological role of this protein in metabolic diseases. We aimed to examine the molecular characteristics of chADAMTS8 and analyze the gene-expression differences in response to toll-like receptor 3 (TLR3) stimulation. Methods: The ADAMTS8 mRNA and amino acid sequences of various species (chicken, duck, cow, mouse, rat, human, chimpanzee, pig, and horse) were retrieved from the Ensembl database and subjected to bioinformatics analyses. Reverse-transcription polymerase chain reaction (RT-PCR) and quantitative PCR (qPCR) experiments were performed with various chicken tissues and the chicken fibroblast DF-1 cell line, which was stimulated with polyinosinic-polycytidylic acid (poly[I:C]; a TLR3 ligand). Results: The chADAMTS8 gene was predicted to contain three thrombospondin type 1 (TSP1) domains, whose amino acid sequences shared homology among the different species, whereas sequences outside the TSP1 domains (especially the amino-terminal region) were very dif­ferent. Phylogenetic analysis revealed that chADAMTS8 is evolutionarily clustered in the same clade with that of the duck. chADAMTS8 mRNA was broadly expressed in chicken tissues, and the expression was significantly up-regulated in the DF-1 cells in response to poly(I:C) stimulation (p<0.05). These results showed that chADAMTS8 may be a target gene for TLR3 signaling. Conclusion: In this report, the genetic information of chADAMTS8 gene, its expression in chicken tissues, and chicken DF-1 cells under the stimulation of TLR3 were shown. The result suggests that chADAMTS8 expression may be induced by viral infection and correlated with TLR3-mediated signaling pathway. Further study of the function of chADAMTS8 during TLR3-dependent inflammation (which represents RNA viral infection) is needed and it will also be important to examine the molecular mechanisms during different regulation, depending on innate immune receptor activation.

• Journal of Life Science
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• v.20 no.12
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• pp.1784-1791
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• 2010

In this study, we investigated the effects of ethyl alcohol extracts of Hizikia fusiforme (EHF) on the correlation between tightening of tight junctions (TJs) and anti-invasive activity in human gastric adenocarcinoma AGS cells. Inhibitory effects of EHF on cell proliferation, motility, and invasiveness were found to be associated with increased tightness of the TJs, which was demonstrated by an increase in transepithelial electrical resistance. Activities of matrix metalloprotease (MMP)-2 and -9 in AGS cells were dose-dependently inhibited by treatment with EHF, and this was also correlated with a decrease in expression of their mRNA and proteins; however, tissue inhibitor of metalloproteinase (TIMP)-1 and -2 mRNA levels were increased. Additionally, immunoblotting results indicated that EHF repressed the levels of claudin proteins (claudin-1, -3, and -4), major components of TJs that play key roles in control and selectivity of paracellular transport. Furthermore, EHF decreased expression of insulin such as growth factor-1 receptor proteins, while concurrently increasing that of thrombospondin-1 and E-cadherin. In conclusion, these results suggest that EHF treatment may inhibit tumor cell motility and invasion, and therefore act as a dietary source to decrease the risk of cancer metastasis.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.22
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• pp.9805-9812
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• 2014

Radixin, a member of the ERM (ezrin-radixin-moesin) family, plays important roles in cell motility, invasion and tumor progression. It is expressed in a variety of normal and neoplastic cells, including many types of epithelial and lymphoid examples. However, its function in glioblastomas remains elusive. Thus, in this study, radixin gene expression was first examined in the glioblastoma cells, then suppressed with a lentivirus-mediated short-hairpin RNA (shRNA) method.We found that there were high levels of radixin expression in glioblastoma U251cells. Radixin shRNA caused down-regulation of radixin gene expression and when radixin-silenced cells were implanted into nude mice, tumor growth was significantly inhibited as compared to blank control cells or nonsense shRNA cells. In addition, microvessel density in the tumors was significantly reduced. Thrombospondin-1 (TSP-1) and E-cadherin were up-regulated in radixin- suppressed glioblastoma U251 cells. In contrast, MMP9 was down-regulated. Taken together, our findings suggest that radixin is involved in GBM cell migration and invasion, and implicate TSP-1, E-cadherin and MMP9 as metastasis-inducing factors.

• The Korean Journal of Pain
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• v.35 no.1
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• pp.66-77
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• 2022

Background: Thrombospondin-4 (TSP4) upregulates in the spinal cord following peripheral nerve injury and contributes to the development of neuropathic pain (NP). We investigated the effects of cyanocobalamin alone or in combination with morphine on pain and the relationship between these effects and spinal TSP4 expression in neuropathic rats. Methods: NP was induced by chronic constriction injury (CCI) of the sciatic nerve. Cyanocobalamin (5 and 10 mg/kg/day) was administered 15 days before CCI and then for 4 and 14 postoperative days. Morphine (2.5 and 5 mg/kg/day) was administered only post-CCI. Combination treatment included cyanocobalamin and morphine, 10 and 5 mg/kg/day, respectively. All drugs were administered intraperitoneally. Nociceptive thresholds were detected by esthesiometer, analgesia meter, and plantar test, and TSP4 expression was assessed by western blotting and fluorescence immunohistochemistry. Results: CCI decreased nociceptive thresholds in all tests and induced TSP4 expression on the 4th postoperative day. The decrease in nociceptive thresholds persisted except for the plantar test, and the increased TSP4 expression reversed on the 14th postoperative day. Cyanocobalamin and low-dose morphine alone did not produce any antinociceptive effects. High-dose morphine improved the decreased nociceptive thresholds in the esthesiometer when administered alone but combined with cyanocobalamin in all tests. Cyanocobalamin and morphine significantly induced TSP4 expression when administered alone in both doses for 4 or 14 days. However, this increase was less when the two drugs are combined. Conclusions: The combination of cyanocobalamin and morphine is more effective in antinociception and partially decreased the induced TSP4 expression compared to the use of either drug alone.

• Reproductive and Developmental Biology
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• v.29 no.3
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• pp.187-191
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• 2005

This study was designed to investigate the changes of quantity and quality of proteins in medium and cytoplasm during in vitro maturation of bovine oocytes. The total quantity of proteins in medium decreased from 0 to 4.5 hr, but increased from 13.5 to 18 hr after the onset of in vitro maturation. The total quantity of protein in cytoplasm increased from 0 to 4.5 hr, decreased from 4.5 to 9 hr, and increased after 18 hr after the onset of in vitro maturation. A total of 298 protein spots was detected on a gel of 2D SDS-PAGE form maturation medium. Among 28 protein spots expressed significant differences in their quantity, 8 proteins were identified by peptide mass fingerprinting (aldose reductase, alpha enolase, apolipoprotein A-1 precursor, 43kDa collectin precursor, heat shock 27kDa protein, plasminogen activator inhibitor-1 precursor, thrombospondin 1, transitional endoplasmic reticulum ATPase). Among total of 35 protein spots detected on gel of 2D SDS-PACE from oorytes cytoplasm, $\beta$-tubulin was identified by peptide mass fingerprinting.