Summary
Calbindin-d 28K expression in insulin-producing tumoral cells of the RINm5F line was assessed by Western-blot and high pressure liquid chromatography. Western blot analysis demonstrated the presence in RINm5F cell homogenates of a protein recognized by a specific polyclonal antibody against chick calbindin. Proteins with apparent molecular weights (mol wt) of 44, 47, 56, and 85 kD were also recognized by the antiserum in RINm5F cell extract, but not in normal rat islet extract. HPLC heat-resistant protein extract from RINm5F cell homogenates revealed three calbindin positive peaks: a major peak with a retention time (20.5 min) identical to that found in a rat cerebellar extract and two minor peaks with shorter retention times. The calbindin content of RINm5F cells was apparently unaffected after 9 d culture in a medium supplemented with 10% calf serum pretreated with dextran-charcoal to remove 1,25-dihydroxyvitamin D3.
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Parmentier M, Lawson DE, Vassart G. Human calbindin complementary DNA sequence: evolutionary and functional implications. Eur. J. Biochem. 1987; 170: 207–215.
Wilson PW, Harding M, Lawson DEM. Putative amino acid sequence of chick calcium- binding protein deduced from a complementary DNA sequence. Nucl. Acids Res. 1985; 13: 8867–8881.
Roth J, Bonner-Weir S, Norman AW, Orci L. Immunocytochemistry of vitamin D- dependent calcium binding protein in chick pancreas: exclusive localization in B-cells. Endocrinology 1982; 110: 2216–2218.
Pochet R, Pipeleers DG, Malaisse WJ, Calbindin D-27kDa: preferential localization in non-B islet cells of the rat. pancreas. Biol. of the Cell 1987; 61: 155–161.
Norman AW, Frankel BJ, Heldt AM, Grodsky GM. Vitamin D deficiency inhibits pancreatic secretion of insulin. Science 1980; 209: 823–825.
Chertow BS, Silvitz WI, Baranetsky NG, Clark SA, Waite A, Deluca HF. Cellular mechanisms of insulin release: the effects of vitamin D deficiency and repletion on rat insulin secretion. Endocrinology 1983; 113: 1511–1518.
Kadowski S, Norman AW. Dietary vitamin D is essential for normal insulin secretion from the perfused rat pancreas. J. Clin. Invest. 1984; 73: 759–766.
Kadowski S, Norman AW. Demonstration that the vitamin D metabolite 1,25(OH)2 − vitamin D3 and not 24 R,25(OH)2-vitamin D3 is essential for normal insulin secretion in the perfused rat pancreas. Diabetes 1985; 34: 315–320.
Billaudel B, Labriji-Mestaghanmi H, Sutter BCJ, Malaisse WJ. Vitamin D and pancreatic islet function. II. Dynamics of insulin release and cationic fluxes. J. Endocrinol. Invest. 1988; 11: 585–593.
Tanaka Y, Seino Y, Ishida M, Yamaoka K, Satomura K, Yabuuchi H, Seino Y, Imura H. Effect of 1,25-dihydroxyvitamin D3 on insulin secretion: direct or mediated. Endocrinology 1986; 118: 1971–1976.
Labriji-Mestaghanmi H, Billaudel B, Garnier PE, Malaisse WJ, Sutter BCJ. Vitamin D and pancreatic islet function I. Time course for changes in insulin secretion and content during vitamin D deprivation and repletion. J. EndocrinoL Invest. 1988; 11: 577–584.
Gadzar AF, Chick WL, Oie HK, Sims HL, King DL, Weir GC, Lauris V. Continuous, clonal, insulin- and somatostatin-secreting cell lines established from a transplantable rat islet cell tumor. Proc. Natl. Acad, Sci. USA 1980; 77: 3519–3523.
Spencer R, Charman M, Wilson PW, Lawson DEM. The relationship between vitamin D-stimuIated Ca transport and intestinal CaBP in the chicken. Biochem. J. 1978; 170: 93–101.
Pasteels B, Miki N, Hatakenaka S, Pochet R. Immunohistochemical comigration between calbindin D-27kDa and visinin. Brain Res. 1987; 412: 107–113.
Malaisse-Lagae F, Malaisse WJ. Insulin release by pancreatic islets. Methods in Diabetes Research, vol. 1, Larner J, and Pohl, SF, Eds, Wiley, NY, 147–152, 1984.
Giroix M-H, Malaisse-Lagae F, Sener A, Malaisse WJ. Hexose metabolism in pancreatic iselts. Galactose transport, phosphorylation and oxidation. Mol. Cell. Biochem. 1985; 66: 61–64.
Bouillon R, De Moor P, Bagglioni EG, Uskovic MR. A radioimmunoassay for 1,25-dihydroxycholecalciferol. Clin. Chem. 1980; 26: 562–567.
Lawson DEM. Methods in Enzymology, vol. 67, McCornick DB, Wright LD, Eds, Academic, New York, 459–465, 1980.
Pochet R, Parmentier M, Lawson DEM, Pasteels, JL. Rat brain synthesizes two “vitamin D-dependent” calcium-binding proteins. Brain Res. 1985; 345: 251–256.
Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature 1970; 227: 680–685.
Johnson DA, Gautsch JW, Sportsman JR, Elder JH. Improved technique utilizing nonfat dry milk for analysis of proteins and nucleic acids transferred to nitrocellulose. Gene Anal. Techn. 1984; 1: 3–8.
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J. Biol. Chem. 1951; 193: 265–275.
Intrator S, Elion J, Thomasset M, Brehier A. Purification, immunological and biochemical characterisation of rat 28kDa cholecalcin (cholecalciferol-induced calcium-binding proteins). Biochem. J. 1985; 231: 89–95.
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Pochet, R., Blachier, F., Lawson, D.E.M. et al. Presence of calbindin-d 28k in endocrine pancreatic tumoral cells of the RINm5F Line. Int J Pancreatol 5, 295–304 (1989). https://doi.org/10.1007/BF02924474
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DOI: https://doi.org/10.1007/BF02924474