| 1. |
- Li, Yuejuan, et al.
(författare)
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Silencing of hyaluronan synthase 2 suppresses the malignant phenotype of invasive breast cancer cells
- 2007
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Ingår i: International Journal of Cancer. - : Wiley. - 0020-7136 .- 1097-0215. ; 120:12, s. 2557-2567
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Tidskriftsartikel (refereegranskat)abstract
- Accumulation of hyaluronan has been demonstrated in the peritumoral breast cancer stroma and nests of tumor cells. In this study, we have quantified the production of hyaluronan and the expression of mRNAs encoding hyaluronan synthesizing (HAS) and hyaluronan degrading (HYAL) enzymes in a panel of breast cancer cell lines. The analysis revealed that highly invasive breast cancer cells produce high amounts of hyaluronan and express preferentially HAS2 mRNA, whereas less invasive breast cancer cells produce low amount of hyaluronan and express HAS1 and HYAL1 mRNAs. We explored the importance of HAS2 expression for breast cancer tumorigenicity, by specifically silencing the HAS2 gene using RNA interference (RNAi)-mediated suppression in the invasive breast cancer cell line Hs578T. This led to a less aggressive phenotype of the breast tumor cells, as assessed by cell growth, both in anchorage-dependent and anchorage-independent cultures. siRNA-mediated knock down of HAS2 in Hs578T breast tumor cells led to an up-regulation of HAS1, HAS3 and HYAL1 mRNAs, resulting in only a 50% decrease in the net hyaluronan production; however, the synthesized hyaluronan was of lower size and more polydisparse compared to control siRNA-treated cells. Interestingly, Hs578T cells deprived of HAS2 migrated only half as efficiently as HAS2 expressing cells through cell-free areas in a culture wounding assay and through Transwell polycarbonate membrane as well as invaded a Matrigel layer. These results imply that alterations in HAS2 expression and endogenously synthesized hyaluronan affect the malignant phenotype of Hs578T breast cancer cells.
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| 2. |
- Ejeskär, Katarina, 1969, et al.
(författare)
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Introduction of in vitro transcribed ENO1 mRNA into neuroblastoma cells induces cell death.
- 2005
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Ingår i: BMC cancer. - : Springer Science and Business Media LLC. - 1471-2407. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Neuroblastoma is a solid tumour of childhood often with an unfavourable outcome. One common genetic feature in aggressive tumours is 1p-deletion.The alpha-enolase (ENO1) gene is located in chromosome region 1p36.2, within the common region of deletion in neuroblastoma. One alternative translated product of the ENO1 gene, known as MBP-1, acts as a negative regulator of the c-myc oncogene, making the ENO1 gene a candidate as a tumour suppressor gene. METHODS: Methods used in this study are transfection of cDNA-vectors and in vitro transcribed mRNA, cell growth assay, TUNEL-assay, real-time RT-PCR (TaqMan) for expression studies, genomic sequencing and DHPLC for mutation detection. RESULTS: Here we demonstrate that transfection of ENO1 cDNA into 1p-deleted neuroblastoma cell lines causes' reduced number of viable cells over time compared to a negative control and that it induces apoptosis. Interestingly, a similar but much stronger dose-dependent reduction of cell growth was observed by transfection of in vitro transcribed ENO1 mRNA into neuroblastoma cells. These effects could also be shown in non-neuroblastoma cells (293-cells), indicating ENO1 to have general tumour suppressor activity.Expression of ENO1 is detectable in primary neuroblastomas of all different stages and no difference in the level of expression can be detected between 1p-deleted and 1p-intact tumour samples. Although small numbers (11 primary neuroblastomas), there is some evidence that Stage 4 tumours has a lower level of ENO1-mRNA than Stage 2 tumours (p = 0.01). However, mutation screening of 44 primary neuroblastomas of all different stages, failed to detect any mutations. CONCLUSION: Our studies indicate that ENO1 has tumour suppressor activity and that high level of ENO1 expression has growth inhibitory effects.
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| 3. |
- Li, Lingli, 1963-
(författare)
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Effect of Hyaluronan-activation of CD44 on Cell Signaling and Tumorigenesis
- 2006
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Hyaluronan (HA), a structural component in the extracellular matrix (ECM), has been recognized as a signaling molecule. It is important during various biological activities such as embryogenesis, angiogenesis, wound healing and tumor progression. Increased amount of hyaluronan during embryonic development is necessary for cell migration and differentiation, but the increased production of hyaluronan by tumor cells or tissue fibroblasts is correlated to poor prognosis for tumor progression and chronic inflammation, respectively. Therefore, understanding the mechanisms regulating HA-enriched matrices and the roles of HA in the biological functions is of fundamental biological importance.Four novel findings are described in this thesis: (1) HA fragments (HA12) and the known angiogenic factor FGF-2 promote endothelial cell differentiation by induction of common but also distinct sets of genes, particularly, upregulation of the chemokine CXCL1/GRO1 gene is necessary for HA12-induced angiogenesis and this effect is dependent on CD44 activation. (2) High concentrations of hyaluronan suppress PDGF-BB-induced fibroblasts migration and PDGFRβ tyrosine phosphorylation upon activation of hyaluronan receptor CD44, probably by recruiting a CD44-associated phosphatase to the PDGFRβ. (3) PDGF-BB stimulates HAS2 transcriptional activity and HA synthesis through upregulation of MAP kinase and PI3 kinase signaling pathways in human dermal fibroblasts. (4) Specific suppression of HAS2 gene in the invasive breast cancer cell line Hs578T by RNA interference (RNAi) leads to a less aggressive phenotype of breast tumor cells. This suppressive effect can be reversed by exogenously added hyaluronan.In conclusion, binding of hyaluronan to CD44 plays an important role in cell signaling, inflammation and tumor progression. Further studies are required to elucidate the molecular mechanisms through which hyaluronan levels are regulated under physiological or pathological conditions, and to explore compounds involved in hyaluronan accumulation and activity as targets for therapies of chronic inflammation and tumors.
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| 4. |
- Li, Lingli, et al.
(författare)
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Growth factor regulation of hyaluronan synthesis and degradation in human dermal fibroblasts : importance of hyaluronan for the mitogenic response of PDGF-BB
- 2007
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Ingår i: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 404, s. 327-336
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Tidskriftsartikel (refereegranskat)abstract
- The glycosaminoglycan hyaluronan is important in many tissue-repair processes. We have investigated the synthesis of hyaluronan in a panel of cell lines of fibroblastic and epithelial origin in response to PDGF (platelet-derived growth factor)-BB and other growth factors. Human dermal fibroblasts exhibited the highest hyaluronan-synthesizing activity in response to PDGF-BB. Analysis of HAS (hyaluronan synthase) and HYAL (hyaluronidase) mRNA expression showed that PDGF-BB treatment induced a 3-fold increase in the already high level of HAS2 mRNA, and increases in HAS1 and HYAL1 mRNA, whereas the levels of HAS3 and HYAL2 mRNA were not affected. Furthermore, PDGF-BB also increased the amount and activity of HAS2 protein, but not of HYAL1 and HYAL2 proteins. Using inhibitors for MEK 1/2 [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase 1/2] (U0126) and for PI3K (phosphoinositide 3-kinase) (LY294002), as well as the SN50 inhibitor, which prevents translocation of the active NF-kappa B (nuclear factor KB) to the nucleus, we observed a complete inhibition of both HAS2 transcriptional activity and hyaluronan synthesis, whereas inhibitors of other signalling pathways were without any significant effect. TGF-beta 1 (transforming growth factor-beta 1) did not increase the activity of hyaluronan synthesis in dermal fibroblasts, but increased the activity of HYALs. Imponantly, inhibition of hyaluronan binding to its receptor CD44 by the monoclonal antibody Hennes-1, inhibited PDGF-BB-stimulated [H-3]thymidine incorporation of dermal fibroblasts. We conclude that the ERK MAPK and PI3K signalling pathways are necessary for the regulation of hyaluronan synthesis by PDGF-BB, and that prevention of its binding to CD44 inhibits PDGF-BB-induced cell growth.
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| 5. |
- Li, Lingli, et al.
(författare)
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Inhibition of Platelet-derived Growth Factor-BB-induced Receptor Activation and Fibroblast Migration by Hyaluronan Activation of CD44
- 2006
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 281:36, s. 26512-26519
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Tidskriftsartikel (refereegranskat)abstract
- The extracellular matrix molecule hyaluronan was found to suppress platelet-derived growth factor (PDGF) beta-receptor activation and PDGF-BB-induced migration of primary human dermal fibroblasts. The suppressive effect of hyaluronan was neutralized by a monoclonal antibody that specifically inhibits hyaluronan binding to its receptor CD44. Moreover, co-immunoprecipitation experiments showed that the PDGF beta-receptor and CD44 can form a complex. Interestingly, the inhibitory effect of hyaluronan on PDGF beta-receptor activation was not seen in the presence of the tyrosine phosphatase inhibitor pervanadate. Our observations suggest that hyaluronan suppresses PDGF beta-receptor activation by recruiting a CD44-associated tyrosine phosphatase to the receptor.
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| 6. |
- Rosenthal, Mark A, et al.
(författare)
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Phase I and Pharmacokinetic Evaluation of Intravenous Hyaluronic Acid in Combination with Doxorubicin or 5-Fluorouracil
- 2005
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Ingår i: Chemotherapy. - : S. Karger AG. - 0009-3157 .- 1421-9794. ; 51:2-3, s. 132-141
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Pre-clinically, hyaluronan (HA) has been demonstrated to systemically target chemotherapeutic drugs to tumours while ameliorating treatment toxicities. This study is a preliminary clinical investigation to determine if HA could be safely used in combination with 5-fluorouracil (5-FU) and doxorubicin (DOX). METHODS: Thirty patients with metastatic cancer were intravenously administered 500 mg/m2 HA in combination with escalating doses of DOX (30-60 mg/m2) or 5-FU (cumulative dose of 1,350-2,250 mg/m2 per cycle). The effect of pre-administration of 20 mg/m2 of folinic acid on HA/5-FU chemotherapy was also investigated. Patients were randomized to receive either HA/chemotherapy or chemotherapy alone in their first treatment cycle and vice versa for the second cycle. Patients received HA and chemotherapy in all subsequent cycles. RESULTS: Treatment was well tolerated, tumour responses were observed and the co-administration of HA did not alter the pharmacokinetics of clinically relevant doses of 5-FU or DOX. CONCLUSION: High doses of intravenous high-molecular-weight HA can be safely co-administered with clinical doses of chemotherapy without significantly altering the toxicity or pharmacokinetics of the drugs or HA.
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| 7. |
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| 8. |
- Takahashi, Yoshinori, et al.
(författare)
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Hyaluronan fragments induce endothelial cell differentiation in a CD44- and CXCL1/GRO1-dependent manner.
- 2005
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Ingår i: J Biol Chem. - 0021-9258.
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Tidskriftsartikel (refereegranskat)abstract
- Hyaluronan is a glycosaminoglycan of the extracellular matrix. In tumors and during chronic inflammatory diseases hyaluronan is degraded to smaller fragments which are known to stimulate endothelial cell differentiation. In this study we have compared the molecular mechanisms through which hyaluronan dodecasaccharides (HA12), and the known angiogenic factor, fibroblast growth factor 2 (FGF-2), induce capillary endothelial cell sprouting in a 3D collagen gel. The gene expression profiles of unstimulated and HA12 or FGF-2 stimulated endothelial cells were compared using a microarray analysis approach. The data revealed that both FGF-2 and HA12 promoted endothelial cell morphogenesis in a process depending on the expression of ornithine decarboxylase (Odc) and ornithine decarboxylase antizyme inhibitor (Oazi) genes. Among the genes selectively upregulated in response to HA12 was the chemokine CXCL1/GRO1 gene. The notion that the induction of CXCL1/GRO1 is of importance for HA12-induced endothelial cell sprouting was supported by the fact that morphogenesis was inhibited by antibodies specifically neutralizing the CXCL1/GRO1 protein product. HA12-stimulated endothelial cell differentiation was exerted via binding to CD44, since it was inhibited by antibodies blocking CD44 function. Our data show that hyaluronan fragments and FGF-2 affect endothelial cell morphogenesis by induction of overlapping, but also distinct sets of genes.
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