Identification of a novel bone/calcium metabolism-regulating factor in porcine pancreas.
We purified from porcine pancreas a hypocalcemic peptide clearly distinguishable from other pancreatic osteotropic factors such as amylin, calcitonin, and glucagon. Porcine pancreas was processed by acetone extraction, anion exchange chromatography, isoelectric focusing, and reverse-phase high performance liquid chromatography. Fractions were assayed for their inhibitory effects on bone resorption in vitro. Amino acid sequence of a homogeneous 28-kDa protein revealed 92% homology to a human elastase IIIB in the N terminus. Recombinant human elastase IIIB (rhEIIIB) inhibited bone resorption in organ culture stimulated by 1,25-dihydroxyvitamin D3 at concentrations as low as 75 ng/ml. Antibodies to rhEIIIB recognized purified pancreatic factor in Western blots and blocked its inhibitory effect on bone resorption. This antiresorptive activity was abolished by phenylmethylsulfonyl fluoride, suggesting the importance of elastase proteolytic activity for inhibition of bone resorption. In vivo, rhEIIIB and purified pancreatic factor significantly decreased recombinant human interleukin-1alpha-induced hypercalcemia. In conclusion, a novel naturally occurring inhibitor of bone resorption and calcium-lowering peptide has been identified in porcine pancreas. Because this pancreatic peptide has systemic effects on bone resorption and blood ionized calcium at low concentrations, it may represent a physiological regulator of normal bone remodeling and calcium homeostasis.
Izbicka
E
E
Department of Medicine, University of Texas Health Science Center at San Antonio, 78284-7877, USA.
Yoneda
T
T
Takaoka
Y
Y
Horn
D
D
Williams
P
P
Mundy
G R
GR
D000595:Amino Acid Sequence; D000818:Animals; D001862:Bone Resorption / Q000378:metabolism; D001842:Bone and Bones / Q000378:metabolism*; D002117:Calcitriol / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D006801:Humans; D006996:Hypocalcemia / Q000378:metabolism; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D008969:Molecular Sequence Data; D010010:Osteoclasts / Q000187:drug effects; D010179:Pancreas / Q000737:chemistry*; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D013552:Swine
D007527:Isoenzymes; D011994:Recombinant Proteins; D010664:Phenylmethylsulfonyl Fluoride; D010196:Pancreatic Elastase; D002117:Calcitriol; D002118:Calcium