HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) All Versions of this Article: 78/5/903    most recent biolreprod.107.064832v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Nagyova, E. Articles by Salustri, A. PubMed PubMed Citation Articles by Nagyova, E. Articles by Salustri, A. Agricola Articles by Nagyova, E. Articles by Salustri, A.

Synthesis of Tumor Necrosis Factor Alpha-Induced Protein 6 in Porcine Preovulatory Follicles: A Study with A38 Antibody¹

Academy of Sciences of the Czech Republic,³ Institute of Animal Physiology and Genetics, 277 21 Libechov, Czech Republic Department of Public Health and Cell Biology,⁴ University of Rome Tor Vergata, 00133 Rome, Italy Wellcome Trust Centre for Cell-Matrix Research,⁵ Faculty of Life Sciences, University of Manchester, M13 9PT Manchester, United Kingdom

ABSTRACT

We have previously shown that the heavy chains (HCs) of inter-alpha-trypsin inhibitor (IalphaI) become covalently linked to hyaluronan (HA) during in vivo and in vitro expansion of porcine oocyte-cumulus cell complexes (OCCs). We have now studied by immunoblotting the synthesis of tumor necrosis factor alpha-induced protein 6 (TNFAIP6), which is essential for catalyzing this reaction in expanding mouse OCCs. Expanding OCCs were collected from preovulatory follicles of naturally cycling pigs and also after in vitro culture (24 or 42 h) in medium supplemented with FSH and pig serum. After isolation, OCCs were treated with Streptomyces hyaluronidase or Chondroitinase ABC. Matrix, cell pellet, and total extracts were analyzed by Western blotting. A band of about 35 kDa and a doublet of about 120 kDa, corresponding to the molecular weight of the native and HC-linked forms of TNFAIP6, respectively, were detected by a rabbit anti-human TNFAIP6 polyclonal antibody in matrix extracts of expanded cumuli. Moreover, we found by using a cell-free assay that porcine follicular fluid collected from follicles at 24 h after hCG stimulation contains HC-HA coupling activity. This activity was abolished by the rat anti-human monoclonal antibody A38, which has an epitope within the Link module domain of TNFAIP6. These experiments suggest that free TNFAIP6 protein was present in follicular fluid aspirated from porcine follicles 24 h after hCG stimulation. In contrast to mouse, we show that the A38 monoclonal antibody does not affect in vitro cumulus expansion of porcine OCCs.

cumulus cells, extracellular matrix, hyaluronan, oocyte-cumulus complex, ovary, pig, TNFAIP6

¹Supported by grant 305/050960 from Grant Agency of the Czech Republic; by grant 52/ZV2 from Ministry of School, Youth, and Education, Program of Scientific and Technological Co-operation between Czech Republic and Italy; and by grant PRIN 2005 from the Italian Ministry of Education.

Correspondence: ²Eva Nagyova, Academy of Sciences of the Czech Republic, Institute of Animal Physiology and Genetics, Rumburska 89, 277 21 Libechov, Czech Republic. FAX: 420 315 639 510; e-mail: nagyova{at}iapg.cas.cz