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  • 1. Bart, Genevieve
    et al.
    Vico, Nuria Ortega
    Hassinen, Antti
    Pujol, Francois M.
    Deen, Ashik Jawahar
    Ruusala, Aino
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Tammi, Raija H.
    Squire, Anthony
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kellokumpu, Sakari
    Tammi, Markku I.
    Fluorescence Resonance Energy Transfer (FRET) and Proximity Ligation Assays Reveal Functionally Relevant Homo-and Heteromeric Complexes among Hyaluronan Synthases HAS1, HAS2, and HAS32015In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 290, no 18, p. 11479-11490Article in journal (Refereed)
    Abstract [en]

    In vertebrates, hyaluronan is produced in the plasma membrane from cytosolic UDP-sugar substrates by hyaluronan synthase 1-3 (HAS1-3) isoenzymes that transfer N-acetylglucosamine (GlcNAc) and glucuronic acid (GlcUA) in alternative positions in the growing polysaccharide chain during its simultaneous extrusion into the extracellular space. It has been shown that HAS2 immunoprecipitates contain functional HAS2 homomers and also heteromers with HAS3 (Karousou, E., Kamiryo, M., Skandalis, S. S., Ruusala, A., Asteriou, T., Passi, A., Yamashita, H., Hellman, U., Heldin, C. H., and Heldin, P. (2010) The activity of hyaluronan synthase 2 is regulated by dimerization and ubiquitination. J. Biol. Chem. 285, 23647-23654). Here we have systematically screened in live cells, potential interactions among the HAS isoenzymes using fluorescence resonance energy transfer (FRET) and flow cytometric quantification. We show that all HAS isoenzymes form homomeric and also heteromeric complexes with each other. The same complexes were detected both in Golgi apparatus and plasma membrane by using FRET microscopy and the acceptor photobleaching method. Proximity ligation assays with HAS antibodies confirmed the presence of HAS1-HAS2, HAS2-HAS2, and HAS2-HAS3 complexes between endogenously expressed HASs. C-terminal deletions revealed that the enzymes interact mainly via uncharacterized N-terminal 86-amino acid domain(s), but additional binding site(s) probably exist in their C-terminal parts. Of all the homomeric complexes HAS1 had the lowest and HAS3 the highest synthetic activity. Interestingly, HAS1 transfection reduced the synthesis of hyaluronan obtained by HAS2 and HAS3, suggesting functional cooperation between the isoenzymes. These data indicate a general tendency of HAS isoenzymes to form both homomeric and heteromeric complexes with potentially important functional consequences on hyaluronan synthesis.

  • 2.
    Bergqvist, Ann-Sofi
    et al.
    Division of Comparative Reproduction, Obstetrics and Udder Health, Department of Clinical Sciences, Faculty of Veterinary Medicine andAnimal Science, Swedish University of Agricultural Sciences, SLU, Uppsala, Sweden.
    Yokoo, Masaki
    Laboratory of Animal Reproduction, Graduate School of Agricultural Science, Tohuku University, Sendai, Japan.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Frendin, Jan
    Division of Large Animal Surgery and Medicine, Section of Anaesthesiology and Intensive Care, Department of Clinical Sciences, Faculty of Veterinary Medicine andAnimal Science, Swedish University of Agricultural Sciences, SLU, Uppsala, Sweden.
    Sato, Eimei
    Laboratory of Animal Reproduction, Graduate School of Agricultural Science, Tohuku University, Sendai, Japan.
    Rodríguez-Martínez, Heriberto
    Division of Comparative Reproduction, Obstetrics and Udder Health, Department of Clinical Sciences, Faculty of Veterinary Medicine andAnimal Science, Swedish University of Agricultural Sciences, SLU, Uppsala, Sweden.
    Hyaluronan and its binding proteins in the epithelium and intraluminal fluid of the bovine oviduct2005In: Zygote (Cambridge. Print), ISSN 0967-1994, E-ISSN 1469-8730, Vol. 13, no 3, p. 207-218Article in journal (Refereed)
    Abstract [en]

    Hyaluronan (HA) is involved in several important steps of sperm storage and of fertilization. This study investigates the presence and concentration of HA in oviductal fluid (ODF), together with the localization of HA and the presence of hyaluronan-binding proteins (HABPs) in the oviductal epithelium of normally cycling dairy heifers and cows. The concentration and amount of HA in ODF, collected over the course of several oestrous cycles via catheters placed in the isthmic and ampullar tubal segments, were measured using an ELISA. The concentration and amount of HA in ODF did not vary significantly between these anatomical regions, nor between the stages of the oestrous cycle (p > 0.05), although the amount of HA seemed to peak during oestrous. The most HA per day (2.9 +/- 0.64 microg, least square mean +/- SEM) was produced on the day of ovulation, whereas the lowest amount (1.25 +/- 0.68 microg) was produced 4 days before ovulation. To investigate the localization of HA, tissue samples were retrieved at well-defined stages of the oestrous cycle and from corresponding regions of the oviduct. Sections and protein extracts from the tissue samples were studied histochemically using biotinylated HABP and immunoblotted with fluorescein isothiocyanate (FITC)-HA, respectively. Presence of HA labelling in the oviductal epithelium was restricted to the sperm reservoir, a localization that seemed to be cycle-independent. The immunoblotting of samples from the lining epithelium revealed seven bands of HABPs. We confirm that the bovine oviduct produces HA and its binding proteins, and that HA is mainly localized to the epithelium of the sperm reservoir.

  • 3. Gianoukakis, Andrew G.
    et al.
    Jennings, Timothy A.
    King, Chris S.
    Sheehan, Christine E.
    Hoa, Neil
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Smith, Terry J.
    Hyaluronan accumulation in thyroid tissue: evidence for contributions from epithelial cells and fibroblasts2007In: Endocrinology, ISSN 0013-7227, E-ISSN 1945-7170, Vol. 148, no 1, p. 54-62Article in journal (Refereed)
    Abstract [en]

    Graves' disease (GD) and Hashimoto's thyroiditis (HT) are autoimmune processes often associated with hyperthyroidism and hypothyroidism, respectively. Despite their diverging clinical presentations, immune activation drives both diseases and results in connective tissue accumulation of the nonsulfated glycosaminoglycan, hyaluronan. The hydrophilic property of hyaluronan contributes to the pathogenesis of thyroid-associated ophthalmopathy, dermopathy and hypothyroid myxedema. Whether hyaluronan accumulates in the thyroid and plays a role in goiter formation in GD and HT remains unknown. We report here that levels of hyaluronan are increased in thyroid tissue from individuals with both diseases compared with glands uninvolved with autoimmune disorders. The transcript encoding hyaluronan synthase (HAS)-3, one of three mammalian HAS isoforms, was detected in thyroid tissue. Isolated thyrocytes in primary culture express all three HAS isoforms when treated with IL-1beta. Thyrocytes and thyroid fibroblasts produce hyaluronan under basal culture conditions and IL-1beta enhances levels of this molecule in both cell types. On a per-cell basis, fibroblasts produce more hyaluronan than do thyrocytes under basal conditions and after cytokine treatment. Synthesis in thyrocytes can also be altered by increasing serum concentration in the medium and by modifying culture density. Our findings suggest that hyaluronan accumulation in thyroid tissue might derive from thyrocytes and fibroblasts. Moreover, this glycosaminoglycan becomes more abundant as a consequence of autoimmune disease. It may therefore contribute to increased thyroid volume in GD and HT. Coupled with the newly identified influence exerted by hyaluronan on immunocompetent cells, our findings represent potentially important insights into the pathogenesis of autoimmune thyroid diseases.

  • 4.
    Heldin, Paraskevi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Basu, Kaustuv
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Kozlova, Inna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Porsch, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    HAS2 and CD44 in Breast Tumorigenesis2014In: Hyaluronan Signaling and Turnover / [ed] Simpson, MA and Heldin, P, Elsevier, 2014, Vol. 123, p. 211-229Chapter in book (Refereed)
    Abstract [en]

    Metastatic spread of breast cancer cells, facilitated by the epithelial-mesenchymal transition (EMT) process, is responsible for the majority of breast cancer mortality. Increased levels of hyaluronan due to deregulation of hyaluronan-synthesizing enzymes, like HAS2, and expression of CD44, the key receptor for hyaluronan, are correlated to poor outcome of patients with basal-like breast cancer. TGFβ induces HAS2 and CD44, both of which are required in the course of efficient TGFβ-induced EMT processes by mammary epithelial cells. Elucidation of the molecular mechanisms underlying tumor-stroma interactions in breast cancer including the regulation of HAS2 and CD44 expression may contribute to the development of better strategies to treat breast cancer patients.

  • 5.
    Heldin, Paraskevi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Basu, Kaustuv
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Olofsson, Berit
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Porsch, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Kozlova, Inna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Kahata, Kaoru
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Deregulation of hyaluronan synthesis, degradation and binding promotes breast cancer2013In: Journal of Biochemistry (Tokyo), ISSN 0021-924X, E-ISSN 1756-2651, Vol. 154, no 5, p. 395-408Article in journal (Refereed)
    Abstract [en]

    Clinical and experimental data indicate that hyaluronan accumulates in breast cancer compared with normal breast epithelium, which correlates to poor prognosis. In this review, we discuss the expression of genes encoding enzymes that synthesize or degrade hyaluronan, i.e. hyaluronan synthases and hyaluronidases or bind hyaluronan, i.e. CD44 and receptor for hyaluronan-mediated motility (RHAMM, also designated as HMMR or CD168), in relation to breast cancer progression. Hyaluronan and hyaluronan receptors have multi-faceted roles in signalling events in breast cancer. A better understanding of the molecular mechanisms underlying these signalling pathways is highly warranted and may lead to improvement of cancer treatment.

  • 6.
    Heldin, Paraskevi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lin, Chun-Yu
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Kaohsiung Med Univ Hosp, Div Infect Dis, Dept Internal Med, Kaohsiung, Taiwan;Kaohsiung Med Univ, Coll Med, Sepsis Res Ctr, Sch Med,Grad Inst Med, Kaohsiung, Taiwan.
    Kolliopoulos, Constantinos
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Chen, Yen-Hsu
    Kaohsiung Med Univ Hosp, Div Infect Dis, Dept Internal Med, Kaohsiung, Taiwan;Kaohsiung Med Univ, Coll Med, Sepsis Res Ctr, Sch Med,Grad Inst Med, Kaohsiung, Taiwan;Natl Chiao Tung Univ, Coll Biol Sci & Technol, Dept Biol Sci & Technol, Hsinchu, Taiwan.
    Skandalis, Spyros S.
    Univ Patras, Dept Chem, Lab Biochem, Biochem Biochem Anal & Matrix Pathobiol Res Grp, GR-26110 Patras, Greece.
    Regulation of hyaluronan biosynthesis and clinical impact of excessive hyaluronan production2019In: Matrix Biology, ISSN 0945-053X, E-ISSN 1569-1802, Vol. 78-79, p. 100-117Article in journal (Refereed)
    Abstract [en]

    The tightly regulated biosynthesis and catabolism of the glycosaminoglycan hyaluronan, as well as its role in organizing tissues and cell signaling, is crucial for the homeostasis of tissues. Overexpression of hyaluronan plays pivotal roles in inflammation and cancer, and markedly high serum and tissue levels of hyaluronan are noted under such pathological conditions. This review focuses on the complexity of the regulation at transcriptional and posttranslational level of hyaluronan synthetic enzymes, and the outcome of their aberrant expression and accumulation of hyaluronan in clinical conditions, such as systemic B-cell cancers, aggressive breast carcinomas, metabolic diseases and virus infection.

  • 7.
    Jacobson, Annica
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Rahmanian, Mehdi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Rubin, Kristofer
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Ludwiginstitutet för Cancerforskning.
    Expression of hyaluronan synthase 2 or hyaluronidase 1 differentially affect the growth rate of transplantable colon carcinoma cell tumors.2002In: Int J Cancer, Vol. 102, p. 212-Article in journal (Refereed)
    Abstract [en]

    Advanced colorectal cancers are often associated with elevated amounts of hyaluronan. To investigate the importance of hyaluronan in colon carcinoma tumor progression, we have expressed by stable transfection hyaluronan synthase 2 (Has2) and hyaluronidase 1 (Hyal1) in the rat colon carcinoma cell line, PROb. We found that hyaluronan overproduction led to a higher growth rate of tumor cells in vitro, and to a faster development of transplantable tumors in syngeneic rats, compared to the mock-transfectants. Has2 transfected PROb cells gave rise to tumors that were significantly less vascularized, but had a significantly larger viable tumor fraction compared to tumors generated from mock-transfectants. In contrast, Hyal1 overexpression suppressed the growth rate of tumor cells both in vitro and in vivo. Moreover, tumors derived from Hyal1-transfected cells had a significantly larger necrotic area than tumors derived from mock- and Has2-transfectants. Our study demonstrates that Has2 overproduction promotes tumorigenicity, whereas Hyal1 overexpression suppresses tumorigenicity in an experimental model for colon carcinoma.

  • 8.
    Karalis, Theodoros T.
    et al.
    Univ Patras, Dept Chem, Lab Biochem, Biochem Biochem Anal & Matrix Pathobiol Res Grp, GR-26110 Patras, Greece.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Vynios, Demitrios H.
    Univ Patras, Dept Chem, Lab Biochem, Biochem Biochem Anal & Matrix Pathobiol Res Grp, GR-26110 Patras, Greece.
    Neill, Thomas
    Thomas Jefferson Univ, Kimmel Canc Ctr, Dept Pathol Anat & Cell Biol, Philadelphia, PA 19107 USA;Thomas Jefferson Univ, Kimmel Canc Ctr, Canc Cell Biol & Signaling Program, Philadelphia, PA 19107 USA.
    Buraschi, Simone
    Thomas Jefferson Univ, Kimmel Canc Ctr, Dept Pathol Anat & Cell Biol, Philadelphia, PA 19107 USA;Thomas Jefferson Univ, Kimmel Canc Ctr, Canc Cell Biol & Signaling Program, Philadelphia, PA 19107 USA.
    Iozzo, Renato V.
    Thomas Jefferson Univ, Kimmel Canc Ctr, Dept Pathol Anat & Cell Biol, Philadelphia, PA 19107 USA;Thomas Jefferson Univ, Kimmel Canc Ctr, Canc Cell Biol & Signaling Program, Philadelphia, PA 19107 USA.
    Karamanos, Nikos K.
    Univ Patras, Dept Chem, Lab Biochem, Biochem Biochem Anal & Matrix Pathobiol Res Grp, GR-26110 Patras, Greece.
    Skandalis, Spyros S.
    Univ Patras, Dept Chem, Lab Biochem, Biochem Biochem Anal & Matrix Pathobiol Res Grp, GR-26110 Patras, Greece.
    Tumor-suppressive functions of 4-MU on breast cancer cells of different ER status: Regulation of hyaluronan/HAS2/CD44 and specific matrix effectors2019In: Matrix Biology, ISSN 0945-053X, E-ISSN 1569-1802, Vol. 78-79, p. 118-138Article in journal (Refereed)
    Abstract [en]

    The malignant phenotype of various cancers is linked to enhanced expression of hyaluronan, a proangiogenic glycosaminoglycan whose expression is suppressed by 4-methylumbelliferone (4-MU), a non-toxic oral agent used as a dietary supplement to improve health and combat prostate cancer. In this study, we investigated the role of 4-MU in mammary carcinoma cells with distinct malignant phenotypes and estrogen receptor (ER) status, a major prognostic factor in the clinical management of breast cancers. We focused on two breast cancer cell lines, the low metastatic and ER alpha+ MCF-7 cells, and the highly-aggressive and ER alpha-MDA-MB-231 cells. Treatment with 4-MU caused a dose-dependent decrease of hyaluronan accumulation in the extracellular matrix as well as within the breast cancer cells, most prevalent in cells lacking ER alpha. This decrease in hyaluronan was accompanied by suppression of Hyaluronan Synthase 2 (HAS2), the major enzyme responsible for the synthesis of hyaluronan, and by induction of hyaluronidases (HYALs) -1 and -2. Moreover, 4-MU induced intense phenotypic changes and substantial loss of CD44, a major hyaluronan receptor, from cell protrusions. Importantly, 4-MU evoked differential effects depending on the absence or presence of ER alpha. Only the ER alpha+ cells showed signs of apoptosis, as determined by cleaved PARP-1, and anoikis as shown by concurrent loss of E-cadherin and beta-catenin. Interestingly, 4-MU significantly reduced migration, adhesion and invasion of ER alpha- breast cancer cells, and concurrently reduced the expression and activity of several matrix degrading enzymes and pro-inflammatory molecules with tumor-promoting functions. Collectively, our findings suggest that 4-MU could represent a novel therapeutic for specific breast cancer subtypes with regard to their ER status via suppression of hyaluronan synthesis and regulation of HAS2, CD44, matrix-degrading enzymes and inflammatory mediators. 

  • 9.
    Kolliopoulos, Constantinos
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Lin, Chun-Yu
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care. Uppsala University, Science for Life Laboratory, SciLifeLab. Division of Infectious Diseases, Department of Internal Medicine, Kaohsiung Medical University Hospital, Sepsis Research Center, Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Moustakas, Aristidis
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Has2 natural antisense RNA and Hmga2 promote Has2 expression during TGFβ-induced EMT in breast cancer2019In: Matrix Biology, ISSN 0945-053X, E-ISSN 1569-1802, Vol. 80, p. 29-45Article in journal (Refereed)
    Abstract [en]

    The glycosaminoglycan hyaluronan has a crucial role in tissue organization and cell signaling. Hyaluronan accumulates in conjunction with rapid tissue remodeling during embryogenesis, as well as in inflammatory conditions and cancer. We report a negative correlation between the expression of genes encoding hyaluronan synthase HAS2, its natural antisense transcript HAS2-AS, the chromatin modulating factor HMGA2 and transforming growth factor-β (TGFβ), and survival of patients with invasive breast carcinomas. In mouse mammary epithelial cells, TGFβ activates Smad and non-Smad signaling pathways, resulting in the transcriptional induction of Has2, Has2as (the mouse ortholog of HAS2-AS) and Hmga2, as well as epithelial-mesenchymal transition (EMT)-promoting transcription factors, such as Snail. Importantly, Has2as abrogation suppressed the TGFβ induction of EMT markers, including Snai1, Hmga2, Fn1, and suppressed the mesenchymal phenotype. TGFβ induction of Hmga2, Has2as and Has2, and synthesis of hyaluronan were accompanied with activation of Akt and Erk1/2 MAP-kinase signaling and were required for breast cancer cell motility. Importantly, the hyaluronan receptor Cd44, but not Hmmr, was required for TGFβ-mediated EMT phenotype. Interestingly, Has2as was found to contribute to the maintenance of stem cell factors and breast cancer stemness. Our findings show that Has2as has a key role in TGFβ- and HAS2-induced breast cancer EMT, migration and acquisition of stemness.

  • 10.
    Kolliopoulos, Constantinos
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Raja, Erna
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Razmara, Masoud
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Moustakas, Aristidis
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    van der Heide, Lars P
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Transforming growth factor β (TGFβ) induces NUAK kinase expression to fine-tune its signaling output2019In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 294, no 11, p. 4119-4136Article in journal (Refereed)
    Abstract [en]

    TGFβ signaling via SMAD proteins and protein kinase pathways up- or down-regulates the expression of many genes and thus affects physiological processes, such as differentiation, migration, cell cycle arrest, and apoptosis during developmental or adult tissue homeostasis. We here report that NUAK family kinase 1 (NUAK1) and NUAK2 are two TGFβ target genes. NUAK1/2 belong to the AMP-activated protein kinase (AMPK) family, whose members control central and protein metabolism, polarity and overall cellular homeostasis. We found that TGFβ-mediated transcriptional induction of NUAK1 and NUAK2 requires SMAD family members 2, 3 and 4 (SMAD2/3/4) and mitogen activated protein kinase (MAPK) activities, which provided immediate and early signals for the transient expression of these two kinases. Genomic mapping identified an enhancer element within the first intron of the NUAK2 gene that can recruit SMAD proteins, which, when cloned, could confer induction by TGFβ.  Furthermore, NUAK2 formed protein complexes with SMAD3 and the TGFβ type I receptor. Functionally, NUAK1 suppressed and NUAK2 induced TGFβ signaling. This was evident during TGFβ-induced epithelial cytostasis, mesenchymal differentiation and myofibroblast contractility, in which NUAK1 or NUAK2 silencing enhanced or inhibited these responses, respectively. In conclusion, we have identified a bifurcating loop during TGFβ signaling, whereby transcriptional induction of NUAK1 serves as a negative checkpoint and NUAK2 induction positively contributes to signaling and terminal differentiation responses to TGFβ activity.

  • 11.
    Kozlova, Inna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Ruusala, Aino
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Voytyuk, Oleksandr
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Skandalis, Spyros S
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    IQGAP1 regulates hyaluronan-mediated fibroblast motility and proliferation2012In: Cellular Signalling, ISSN 0898-6568, E-ISSN 1873-3913, Vol. 24, no 9, p. 1856-1862Article in journal (Refereed)
    Abstract [en]

    IQGAP1, an essential scaffolding protein, forms a complex with the hyaluronan receptor CD44. In this study, we have examined the importance of IQGAP1 for hyaluronan-mediated fibroblast migration and proliferation. Hyaluronan induced formation of F-actin fibers and focal adhesions, which was dependent on IQGAP1. IQGAP1 was required for hyaluronan- but not for platelet-derived growth factor (PDGF)-BB-induced cell migration, and was required for both hyaluronan- and PDGF-BB-mediated fibroblast proliferation, but not for proliferation induced by 10% fetal bovine serum. Depletion of IQGAP1 suppressed hyaluronan-induced activation of Rac1 and enhanced the activation of RhoA. Taken together, these findings indicate important roles for IQGAP1 in hyaluronan-stimulated migration and proliferation of fibroblasts.

  • 12.
    Li, Lingli
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Inhibition of Platelet-derived Growth Factor-BB-induced Receptor Activation and Fibroblast Migration by Hyaluronan Activation of CD442006In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 281, no 36, p. 26512-26519Article in journal (Refereed)
    Abstract [en]

    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.

  • 13. Li, Y
    et al.
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hyaluronan production increases the malignant properties of mesotheliomacells.2001In: Br J Cancer, Vol. 85, p. 600-Article in journal (Refereed)
    Abstract [en]

    Malignant pleural mesotheliomas is in most cases associated with elevated amounts of hyaluronan. To investigate the importance of hyaluronan for the malignant properties of mesotheliomas, we have expressed murine hyaluronan synthase 2 (HAS2) in the non-hyaluronan producing mesothelioma cell line, Mero-25. We found that upon hyaluronan overproduction the mesothelioma cells changed their epitheloid character to a fibroblastic phenotype and were surrounded by pericellular matrices, the size of which correlated to the amount of synthesized hyaluronan. HAS2-transfected cells with the ability to synthesize about 520 ng hyaluronan/5 x 10(4)cells/24 h exhibited about a 2-fold increase in the expression of the cell surface hyaluronan receptor CD44 and their locomotion increased compared to that of mock-transfected Mero-25 cells. Furthermore, the malignant properties of mesothelioma cell clones as determined by the ability to grow in a soft agar assay correlated to their hyaluronan production. These results provide evidence for an important role of hyaluronan in the aggressive spread of mesotheliomas in adjacent non-cancerous stromal tissues.

  • 14.
    Lin, Chun-Yu
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care. Kaohsiung Med Univ, Kaohsiung Med Univ Hosp, Infect Control Ctr, Div Infect Dis,Dept Internal Med, Kaohsiung 807, Taiwan;Kaohsiung Med Univ, Sepsis Res Ctr, Ctr Dengue Fever Control & Res, Sch Med,Grad Inst Med, Kaohsiung, Taiwan.
    Kolliopoulos, Constantinos
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Huang, Chung-Hao
    Kaohsiung Med Univ, Kaohsiung Med Univ Hosp, Infect Control Ctr, Div Infect Dis,Dept Internal Med, Kaohsiung 807, Taiwan;Kaohsiung Med Univ, Sepsis Res Ctr, Ctr Dengue Fever Control & Res, Sch Med,Grad Inst Med, Kaohsiung, Taiwan.
    Tenhunen, Jyrki
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care. Tampere Univ Hosp, Dept Intens Care, Crit Care Med Res Grp, Tampere, Finland.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Chen, Yen-Hsu
    Kaohsiung Med Univ, Sepsis Res Ctr, Ctr Dengue Fever Control & Res, Sch Med,Grad Inst Med, Kaohsiung, Taiwan;Kaohsiung Municipal Tatung Hosp, Dept Internal Med, 68 Jhonghua 3rd Rd, Kaohsiung 80145, Taiwan;Natl Chiao Tung Univ, Coll Biol Sci & Technol, Dept Biol Sci & Technol, Hsinchu, Taiwan.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    High levels of serum hyaluronan is an early predictor of dengue warning signs and perturbs vascular integrity2019In: EBioMedicine, E-ISSN 2352-3964, Vol. 48, p. 425-441Article in journal (Refereed)
    Abstract [en]

    Background: A main pathological feature of severe dengue virus infection is endothelial hyper-permeability. The dengue virus nonstructural protein 1 (NS1) has been implicated in the vascular leakage that characterizes severe dengue virus infection, however, the molecular mechanisms involved are not known.

    Methods: A cohort of 250 dengue patients has been followed from the onset of symptoms to the recovery phase. Set urn hyaluronan levels and several other clinical parameters were recorded. The effect of NS1 treatment of cultured fibroblasts and endothelial cells on the expressions of hyaluronan synthetic and catabolic enzymes and the hyaluronan receptor CD44, were determined, as have the effects on the formation of hyaluronan-rich matrices and endothelial permeability.

    Findings: Elevated serum hyaluronan levels (70 ng/ml) during early infection was found to be an independent predictor for occurrence of warning signs, and thus severe dengue fever. High circulating levels of the viral protein NS1, indicative of disease severity, correlated with high concentrations of serum hyaluronan. NS1 exposure decreased the expression of CD44 in differentiating endothelial cells impairing the integrity of vessel-like structures, and promoted the synthesis of hyaluronan in dermal fibroblasts and endothelial cells in synergy with dengue-induced pro-inflammatory mediators. Deposited hyaluronan-rich matrices around cells cultured in vitro recruited CD44-expressing macrophage-like cells, suggesting a mechanism for enhancement of inflammation. In cultured endothelial cells, perturbed hyaluronan-CD44 interactions enhanced endothelial permeability through modulation of VE-cadherin and cytoskeleton re-organization, and exacerbated the NS1-induced disruption of endothelial integrity.

    Interpretation: Pharmacological targeting of hyaluronan biosynthesis and/or its CD44-mediated signaling may limit the life-threatening vascular leakiness during moderate-to-severe dengue virus infection. 

  • 15.
    Mehic, Merima, Sr.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    de Sa, Vanessa Karen, Sr.
    AC Camargo Canc Ctr, Sao Paulo, SP.
    Hebestreit, Sandra
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik, Sr.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Ludwig Inst Canc Res, Uppsala.
    The role of deubiquitinating enzyme USP17, hyaluronan synthase 2, and hyaluronan in non-small-cell lung cancer oncogenic transformation2018In: Clinical Cancer Research, ISSN 1078-0432, E-ISSN 1557-3265, Vol. 24, no 1, p. 96-96Article in journal (Other academic)
    Abstract [en]

    Introduction: Lung cancer is the result of a multistep accumulation of genetic and/or epigenetic alterations; therefore, a better understanding of the molecular mechanism by which these alterations affect lung cancer pathogenesis would provide new diagnostic procedures and prognostic factors for early detection of recurrence. The remarkable qualitative and quantitative modifications of extracellular matrix components as the deubiquitinating enzyme (USP17), hyaluronan (HA), and hyaluronan synthases 2 (HAS 2) may favor invasion, cellular motility, and proliferation in several cancers including lung.

    Results: The silencing of USP17 led to decreased hyaluronan production, whereas the suppression of USP4 increased hyaluronan synthesis. Importantly, high levels of USP17 and HAS2 were detected in a panel of cancer cell lines compared to normal cells, and immunohistochemical stainings revealed higher expression of USP17 and HAS2 in tissues of lung cancer patients compared to normal tissue. Numerous epithelial cells expressed USP17 and HAS2 in dysplasia compared to squamous cell carcinoma (SqCC) (p=0.001). USP17 and HAS2 were prominently expressed in adenocarcinoma (ADC) (p≤0.005). HA immunostaining indexes were increased in ADC and SqCC compared to normal and dysplasia cells (p=0.05). Consistent with the immunohistochemical analyses, low amounts of hyaluronan and USP17 were observed in SqCC by confocal analysis, coincident with less colocalization as determined by confocal microscopy. In contrast, a high expression of hyaluronan (48% of positive index) and high USP17 expression (78% of positive index) in ADC was consistent with a higher degree of colocalization.

    Conclusions: HAS2, hyaluronan and USP17 were expressed at higher levels in particular in preneoplastic lesions and ADC, suggesting a role in NSCLC oncogenic transformation, possibly by promoting cellular division by USP17-mediated. Elucidation of the mechanism of how USP17 and HAS2 cooperate in the regulation of the cell cycle might be of therapeutic importance.

  • 16.
    Mehić, Merima
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    de Sa, Vanessa Karen
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab. AC Camargo Canc Ctr, Int Res Ctr, Dept Genom & Mol Biol, Sao Paulo, Brazil.; Univ Sao Paulo, Fac Med, Dept Pathol, Sao Paulo, Brazil.
    Hebestreit, Sandra
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    The deubiquitinating enzymes USP4 and USP17 target hyaluronan synthase 2 and differentially affect its function2017In: Oncogenesis, E-ISSN 2157-9024, Vol. 6, article id e348Article in journal (Refereed)
    Abstract [en]

    The levels of hyaluronan, a ubiquitous glycosaminoglycan prominent in the extracellular matrix, is balanced through the actions of hyaluronan-synthesizing enzymes (HAS1, 2 and 3) and degrading hyaluronidases (Hyal 1, 2, 3 and PH20). Hyaluronan accumulates in rapidly remodeling tissues, such as breast cancer, due to deregulated expression of the HAS2 gene and/or alterations of HAS2 activity. The activity of HAS2 is regulated by post-translational modifications, including ubiquitination. In order to identify deubiquitinating enzymes (DUBs) that are involved in de-ubiquitination of HAS2, a complementary (cDNA) library of 69 Flag-HA-tagged human DUBs cloned into retroviral vectors was screened in human embryonic kidney (HEK) 293T cells for their ability to de-ubiquitinate myc-tagged HAS2. Several DUBs were found to decrease the ubiquitination of 6myc-HAS2, among which, the most effective were USP17 and USP4. USP17 efficiently removed polyubiquitination, whereas USP4 preferentially removed monoubiquitination of 6myc-HAS2. Co-immunoprecipitation studies revealed interactions between HAS2 and USP17, as well as between HAS2 and USP4, in membrane preparations of HEK293T cells. USP17 significantly stabilized 6myc-HAS2 protein levels, whereas USP4 did not. The silencing of USP17 led to decreased hyaluronan production, whereas the suppression of USP4 increased hyaluronan synthesis. Importantly, high levels of USP17 and HAS2 were detected in a panel of cancer cell lines compared to normal cells, and immunohistochemical stainings revealed higher expression of USP17 and HAS2 in tissues of lung cancer patients compared to normal tissue. In conclusion, USP17 and USP4 differently affect HAS2 ubiquitination, and the stability and function of HAS2.

  • 17.
    Melero-Fernandez de Mera, R. M.
    et al.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland.
    Arasu, U. T.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland.
    Karna, R.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland.
    Oikari, S.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland.
    Rilla, K.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland.
    Vigetti, D.
    Univ Insubria, Dept Med & Surg, Varese, Italy.
    Passi, A.
    Univ Insubria, Dept Med & Surg, Varese, Italy.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Tammi, M. I.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland.
    Deen, A. J.
    Univ Eastern Finland, Inst Biomed, Kuopio, Finland;Univ Eastern Finland, AI Virtanen Inst Mol Sci, Kuopio, Finland.
    Effects of mutations in the post-translational modification sites on the trafficking of hyaluronan synthase 2 (HAS2)2019In: Matrix Biology, ISSN 0945-053X, E-ISSN 1569-1802, Vol. 80, p. 85-103Article in journal (Refereed)
    Abstract [en]

    Vesicular trafficking of hyaluronan synthases (HAS1-3) from endoplasmic reticulum (ER) through Golgi to plasma membrane (PM), and either back to endosomes and lysosomes, or out into extracellular vesicles, is important for their activities. We studied how post-translational modifications affect the trafficking of HAS2 by mutagenesis of the sites of ubiquitination (K190R), phosphorylation (T110A) and 0-GIcNAcylation (S221A), using Dendra2- and EGFP-HAS2 transfected into COS1 cells. Confocal microscopy showed HAS2 wild type (wt) and its K19OR and S221A mutants in ER, Golgi and extracellular vesicles, while the T110A mutant remained mostly in the ER. HA synthesis was reduced by S221A, while completely blocked by K19OR and T110A. Cell-surface biotinylation indicated that T110A was absent from PM, while S221A was close to the level of wt, and K190R was increased in PM. TIRF microscopy analysis gave similar results. Rabl 0 silencing increased HA secretion by HAS2, likely by inhibiting endocytosis of the enzyme from PM, as reported before for HAS3. Green-to-red photo-conversion of Dendra2-HAS2 constructs suggested slower decay of K190R and S221A than HAS2 wt, while T110A was barely degraded at all. S221D and S221E, the phosphomimetic mutants of this site, decayed faster and blocked hyaluronan synthesis, suggesting alternative 0-GIcNAci-PO4 substitution to regulate the stability of the enzyme. Probing the role of dynamic 0-GIcNAcylation at S221 by adding glucosamine increased the half-life of only HAS2 wt. The Dendra2 " HAS2 disappearance from Golgi was slower for K190R. Of the two inactive constructs, K190R co-transfected with HAS2 wt suppressed, whereas T110A had no effect on HA synthesis. Interestingly, the HAS2stimulated shedding of extracellular vesicles was dependent on HAS residence in PM but independent of HA synthesis. The results indicate that post-translational modifications control the trafficking of HAS2, and that trafficking is an integral part of the post-translational regulation of HAS2 activity.

  • 18. Nishitsuka, Koichi
    et al.
    Kashiwagi, Yoshiko
    Tojo, Naoki
    Kanno, Chikako
    Takahashi, Yoshinori
    Yamamoto, Teiko
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Yamashita, Hidetoshi
    Hyaluronan production regulation from porcine hyalocyte cell line by cytokines2007In: Experimental Eye Research, ISSN 0014-4835, E-ISSN 1096-0007, Vol. 85, no 4, p. 539-545Article in journal (Refereed)
    Abstract [en]

    The objective of this study were to establish a cell line derived from porcine hyalocytes and to investigate the regulation of hyaluronan (HA) synthesis in response to cytokines. After 50 passages of the cells derived from porcine vitreous tissue, a cell line was generated. The immortalized cells showed fibroblastic morphology. The cell doubling time was 56.9h. In the mRNA level, the cells expressed plate-derived growth factor (PDGF) alpha receptor, PDGF beta receptor, transforming growth factor-beta (TGF-beta) type I receptor, TGF-beta type II receptor, CD44, collagen type I, collagen type II, glial fibrillary acidic protein (GFAP), hyaluronan synthase (HAS) 2, HAS 3 and beta-actin. In the protein level, GFAP was expressed in this cell line. S-100 protein and cytokeratin were not detected. Stimulation with TGF-beta1 and/or PDGF-BB induced a marked increase in the expression level of HAS2 mRNA, and induced HA production. TGF-beta1 stimulated HAS2 expression through the signal transduction pathway including Smad 2,3,4. In summary, this report constitutes the first successful immortalization of porcine hyalocyte cells. The production of HA was induced from the generated porcine hyalocyte cell line under the stimulation of TGF-beta1 and/or PDGF-BB, which may be related to the pathogenesis of proliferative membrane formation in proliferative vitreo-retinal diseases.

  • 19. Papanikolaou, Vassilios
    et al.
    Vrochides, Dionisios
    Gakis, Dimitrios
    Patsiaoura, Kalliopi
    Margari, Paraskevi
    Dogramatzi, Fani
    Antoniades, Antonios
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Ludwiginstitutet för Cancerforskning.
    Improvement of rat liver graft function after storage in University ofWisconsin solution containing testicular hyaluronidase.2002In: Liver Transpl, Vol. 8, p. 1028-Article in journal (Refereed)
    Abstract [en]

    Hyaluronan accumulates at sites of inflammation, which affects the organization of matrix and thereby the proliferation, migration, and adherence of cells. In this study we investigated possible beneficial effects of the hyaluronan-degrading enzyme hyaluronidase on rat liver graft viability. Orthotopic rat liver transplantation was performed using a cuff technique in Wistar AL Bacharach Glaxo (WAG) rats grafted with WAG livers, which had been stored in the University of Wisconsin (UW) solution or in UW solution enriched with testicular hyaluronidase. Liver tissue architecture, as well as tissue and serum hyaluronan levels, were determined using immunohistochemistry and biochemical assays. Addition of testicular hyaluronidase (0.4 mg/mL) to livers preserved for 24 hours in cold UW solution followed by brief exposure to Ringer's lactate both prolonged the function of the grafted livers and improved their viability (4 of 10 grafts survived, compared with 0 of 10 in the control group). Hyaluronidase treatment did not damage the liver tissue architecture, and a reduced edema was observed in the survivors. Furthermore, 10 minutes after restoration of circulation, higher serum hyaluronan levels were observed in nonsuccessful compared with successful transplantations, whereas no differences in the levels of other serum viability markers were detected. We conclude that addition of testicular hyaluronidase to storage UW solution limits liver cell damage and considerably improves graft function. Furthermore, our data suggest that serum hyaluronan level is a better marker than other serum markers for early evaluation of postoperative graft function.

  • 20.
    Porsch, Helena
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Bernert, Berit
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Mehić, Merima
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Theocharis, A D
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Heldin, Carl-Henrik
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Paraskevi
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Efficient TGF beta-induced epithelial-mesenchymal transition depends on hyaluronan synthase HAS22013In: Oncogene, ISSN 0950-9232, E-ISSN 1476-5594, Vol. 32, no 37, p. 4355-4365Article in journal (Refereed)
    Abstract [en]

    Epithelial-mesenchymal transition (EMT) is a developmental program, which can be adopted by cancer cells to increase their migration and ability to form metastases. Transforming growth factor β (TGFβ) is a well-studied inducer of EMT. We demonstrate that TGFβ potently stimulates hyaluronan synthesis via upregulation of hyaluronan synthase 2 (HAS2) in NMuMG mammary epithelial cells. This stimulatory effect requires the kinase active type I TGFβ receptor and is dependent on Smad signaling and activation of the p38 mitogen-activated protein kinase. Knockdown of HAS2 inhibited the TGFβ-induced EMT by about 50%, as determined by the phase contrast microscopy and immunostaining using the EMT marker ZO-1. Furthermore, real-time PCR analysis of the EMT markers fibronectin, Snail1 and Zeb1 revealed decreased expressions upon HAS2 suppression, using specific small interfering RNA (siRNA) for HAS2. Removal of the extracellular hyaluronan by Streptomyces hyaluronidase or inhibiting the binding to its cell surface receptor CD44 by blocking antibodies, did not inhibit TGFβ-induced EMT. Interestingly, HAS2 suppression completely abolished the TGFβ-induced cell migration, whereas CD44 knockdown did not. These observations suggest that TGFβ-dependent HAS2 expression, but not extracellular hyaluronan, has an important regulatory role in TGFβ-induced EMT.

  • 21.
    Porsch, Helena
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Mehic, Merima
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Olofsson, Berit
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Platelet-derived Growth Factor beta-Receptor, Transforming Growth Factor beta Type I Receptor, and CD44 Protein Modulate Each Other's Signaling and Stability2014In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 289, no 28, p. 19747-19757Article in journal (Refereed)
    Abstract [en]

    Growth factors, such as platelet-derived growth factor BB (PDGF-BB) and transforming growth factor beta(TGF beta), are key regulators of cellular functions, including proliferation, migration, and differentiation. Growth factor signaling is modulated by context-dependent cross-talk between different signaling pathways. We demonstrate in this study that PDGF-BB induces phosphorylation of Smad2, a downstream mediator of the canonical TGF beta pathway, in primary dermal fibroblasts. The PDGF-BB-mediated Smad2 phosphorylation was dependent on the kinase activities of both TGF beta type I receptor (T beta RI) and PDGF beta-receptor (PDGFR beta), and it was prevented by inhibitory antibodies against TGF beta. Inhibition of the activity of the T beta RI kinase greatly reduced the PDGF-BB-dependent migration in dermal fibroblasts. Moreover, we demonstrate that the receptors for PDGF-BB and TGF beta interact physically in primary dermal fibroblasts and that stimulation with PDGF-BB induces internalization not only of PDGFR beta but also of T beta RI. In addition, silencing of PDGFR beta by siRNA decreased the stability of T beta RI and delayed TGF beta-induced signaling. We further show that the hyaluronan receptor CD44 interacts with both PDGFR beta and T beta RI. Depletion of CD44 by siRNA increased signaling via PDGFR beta and T beta RI by stabilizing the receptor proteins. Our data suggest that cross-talk between PDGFR beta and T beta RI occurs in dermal fibroblasts and that CD44 negatively modulates signaling via these receptors.

  • 22.
    Rahmanian, Mehdi
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Ludwiginstitutet för Cancerforskning.
    Testicular hyaluronidase induces tubular structures of endothelial cells grown in three-dimensional collagen gel through a CD44-mediated mechanism.2002In: Int J Cancer, Vol. 97, p. 601-Article in journal (Refereed)
    Abstract [en]

    Cultured brain capillary endothelial cells grown in a 3-dimensional collagen gel can form tubular structures after stimulation by angiogenic factors. We found that treatment of such cultures with testicular hyaluronidase led to formation of tubular structures and cell survival. Anion-exchange chromatography of the enzyme preparation on a MonoQ column revealed the presence of the angiogenic factor basic fibroblast growth factor (bFGF) in the flow-through fraction, as determined by immunoblotting; part of the effect on endothelial cell morphogenesis could thus be ascribed to bFGF. However, adsorbed fractions eluted with increasing concentrations of NaCl, which exhibited hyaluronan-degrading activity at neutral pH, did not contain bFGF but were still able to induce tube-like structures of the endothelial cells. Streptomyces hyaluronidase failed to evoke the same effect. Interestingly, blocking of hyaluronan binding to CD44 receptors by the monoclonal antibody KM114 inhibited the effect of hyaluronidase, but not of bFGF, on endothelial cell tube formation. Our data suggest a CD44-mediated specific role for certain populations of testicular hyaluronidase in the induction of angiogenesis.

  • 23. Rosenthal, Mark A
    et al.
    Gibbs, Peter
    Brown, Tracey J
    Wong, Shirley
    Uren, Shannon
    Ellis, Andrew
    Li, Lingli
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Fox, Richard M
    Phase I and Pharmacokinetic Evaluation of Intravenous Hyaluronic Acid in Combination with Doxorubicin or 5-Fluorouracil2005In: Chemotherapy, ISSN 0009-3157, E-ISSN 1421-9794, Vol. 51, no 2-3, p. 132-141Article in journal (Refereed)
    Abstract [en]

    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.

  • 24. Simpson, Melanie A.
    et al.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Advances in Cancer Research: Hyaluronan Signaling And Turnover, Volume One Hundred And Twenty Three, Preface2014In: Hyaluronan Signaling and Turnover / [ed] Simpson, MA andHeldin, P, Elsevier, 2014, p. 15-16Chapter in book (Other academic)
  • 25. Simpson, Melanie A
    et al.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Preface2014In: Advances in Cancer Research, ISSN 0065-230X, E-ISSN 2162-5557, Vol. 123, p. xv-xviArticle in journal (Refereed)
  • 26.
    Takahashi, Yoshinori
    et al.
    Ludwiginstitutet för Cancerforskning.
    Li, Lingli
    Ludwiginstitutet för Cancerforskning.
    Kamiryo, Masaru
    Ludwiginstitutet för Cancerforskning.
    Asteriou, Trias
    Ludwiginstitutet för Cancerforskning.
    Moustakas, Aristidis
    Ludwiginstitutet för Cancerforskning.
    Yamashita, Hidetoshi
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Ludwiginstitutet för Cancerforskning.
    Hyaluronan fragments induce endothelial cell differentiation in a CD44- and CXCL1/GRO1-dependent manner.2005In: J Biol Chem, ISSN 0021-9258Article in journal (Refereed)
    Abstract [en]

    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.

  • 27.
    Teder, P
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Nettelbladt, O
    Department of Medical Sciences.
    Heldin, P
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Characterization of the mechanism involved in bleomycin-induced increased hyaluronan production in rat lung1995In: Am. J. Respir. Cell Mol. Biol., Vol. 12, p. 181-189Article in journal (Refereed)
  • 28.
    Teder, P
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Department of Medical Sciences.
    Versnel, M A
    Heldin, P
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Stimulatory effects of pleural fluids from mesothelioma patients on CD44 expression, hyaluronan production and cell proliferation in primary cultures of normal mesothelial and transformed cells1996In: Int J Cancer, Vol. 67, p. 393-398Article in journal (Refereed)
  • 29.
    Tzanakakis, George
    et al.
    Lab of Anatomy-Histology-Embryology, Medical School, University of Crete, 71003 Heraklion, Greece.
    Kovalszky, Ilona
    1st Department of Pathology and Experimental Cancer Research, Semmelweis University, Budapest H-1085, Hungary.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Nikitovic, Dragana
    Lab of Anatomy-Histology-Embryology, Medical School, University of Crete, 71003 Heraklion, Greece.
    Proteoglycans/Glycosaminoglycans: from basic research to clinical practice2014In: BioMed Research International, ISSN 2314-6133, E-ISSN 2314-6141, Vol. 2014, article id 295254Article in journal (Refereed)
  • 30. Udabage, Lishanthi
    et al.
    Brownlee, Gary R.
    Waltham, Mark
    Blick, Tony
    Walker, Emma C.
    Heldin, Paraskevi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Nilsson, Susan K.
    Thompson, Erik W.
    Brown, Tracey J
    Antisense-mediated suppression of hyaluronan synthase 2 inhibits the tumorigenesis and progression of breast cancer2005In: Cancer Research, ISSN 0008-5472, E-ISSN 1538-7445, Vol. 65, no 14, p. 6139-6150Article in journal (Refereed)
    Abstract [en]

    The progression of several cancers is correlated with the increased synthesis of the glycosaminoglycan, hyaluronan. Hyaluronan is synthesized at the plasma membrane by various isoforms of hyaluronan synthases (HAS). The importance of HAS2 expression in highly invasive breast cancer was characterized by the antisense inhibition of HAS2 (ASHAS2). The effect of HAS2 inhibition on cell proliferation, migration, hyaluronan metabolism, and receptor status was characterized in vitro, whereas the effect on tumorigenicity and metastasis was established in vivo. HAS2 inhibition resulted in a 24-hour lag in proliferation that was concomitant to transient arrest of 79% of the cell population in G0-G1. Inhibition of HAS2 did not alter the expression of the other HAS isoforms, whereas hyaluronidase (HYAL2) and the hyaluronan receptor, CD44, were significantly down-regulated. ASHAS2 cells accumulated greater amounts of high molecular weight hyaluronan (>10,000 kDa) in the culture medium, whereas mock and parental cells liberated less hyaluronan of three distinct molecular weights (100, 400, and 3,000 kDa). The inhibition of HAS2 in the highly invasive MDA-MB-231 breast cancer cell line inhibited the initiation and progression of primary and secondary tumor formation following s.c. and intracardiac inoculation into nude mice, whereas controls readily established both primary and secondary tumors. The lack of primary and secondary tumor formation was manifested by increased survival times where ASHAS2 animals survived 172% longer than the control animals. Collectively, these unique results strongly implicate the central role of HAS2 in the initiation and progression of breast cancer, potentially highlighting the co-dependency between HAS2, CD44, and HYAL2 expression.

  • 31. Usui, T.
    et al.
    Nakajima, F.
    Ideta, R.
    Kaji, Y.
    Suzuki, Y.
    Araie, M.
    Miyauchi, S.
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Ludwiginstitutet för Cancerforskning.
    Yamashita, H.
    Hyaluronan synthase in trabecular meshwork cells.2003In: Br J Ophthalmol, ISSN 0007-1161, Vol. 87, no 3, p. 357-60Article in journal (Refereed)
    Abstract [en]

    BACKGROUND/AIMS: Hyaluronan is present in the trabecular meshwork where it is involved in the pathophysiology of aqueous outflow environment. In this study, the expression and regulation of hyaluronan synthase (HAS), which is the enzyme synthesising hyaluronan, in trabecular meshwork cells were investigated. METHODS: Cultured bovine trabecular meshwork cells (BTMCs) were used. HAS expression in BTMCs was examined by RT-PCR. The effects of transforming growth factor beta (TGF-beta) and platelet derived growth factor BB (PDGF-BB) on HAS expression in BTMCs were examined by quantitative RT-PCR. The HAS2 expression by TGF-beta and PDGF-BB at the protein level was also confirmed immunohistochemically. The production of hyaluronan from BTMCs was detected by high performance liquid chromatography (HPLC). RESULTS: Three HAS isoforms were expressed in BTMCs at the mRNA level. Among HAS isoforms, only the expression of HAS2 mRNA was increased by the administration of TGF-beta or PDGF-BB. HAS2 upregulation by these growth factors was also confirmed at the protein level. Further, hyaluronan production from BTMCs was stimulated by TGF-beta or PDGF-BB. CONCLUSION: Expression of HAS in trabecular meshwork may maintain the hyaluronan content in the aqueous outflow pathway. Its production is regulated by TGF-beta and PDGF-BB. The regulation of the expression of HAS in trabecular meshwork might be useful for modulating the aqueous outflow environment.

  • 32. Warn, Richard
    et al.
    Harvey, Pascale
    Warn, Alba
    Foley-Comer, Adam
    Heldin, Paraskevi
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Versnel, Marjan
    Arakaki, Naokatu
    Daikuhara, Yasushi
    Laurent, Geoffrey J.
    Herrick, Sarah E.
    Mutsaers, Steven E.
    Hgf/sf induces mesothelial cell migration and proliferation by autocrineand paracrine pathways.2001In: Exp Cell Res, Vol. 267, p. 258-Article in journal (Refereed)
    Abstract [en]

    Mesothelial repair differs from that of other epithelial-like surfaces as healing does not occur solely by centripetal in-growth of cells as a sheet from the wound margins. Mesothelial cells lose their cell-cell junctions, divide, and adopt a fibroblast-like morphology while scattering across and covering the wound surface. These features are consistent with a cellular response to hepatocyte growth factor/scatter factor (HGF/SF). In this study, we examined the ability of mesothelial cells to secrete HGF/SF and investigated its possible role as an autocrine regulator of mesothelial cell motility and proliferation. We found that human primary mesothelial cells expressed HGF/SF mRNA and secreted active HGF/SF into conditioned medium as determined by ELISA and in a scattering bioassay. These cells also expressed the HGF/SF receptor, Met, as shown by RT-PCR and by Western blot analysis and immunofluorescence. Incubation of mesothelial cells with neutralizing antibodies to HGF/SF decreased cell migration to 25% of controls, whereas addition of HGF/SF disrupted cell-cell junctions and induced scattering and enhanced mesothelial cell migration. Furthermore, HGF/SF showed a small but significant mitogenic effect on all mesothelial cell lines examined. In conclusion, HGF/SF is produced by mesothelial cells and induces both motility and proliferation of these cells. These data are consistent with HGF/SF playing an autocrine role in mesothelial healing.

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