Kittrawee Kritmetapak1, Ravinder J Singh2, Theodore A Craig3, Jolaine M Hines4, Rajiv Kumar5. 1. Division of Nephrology and Hypertension, Department of Medicine, Mayo Clinic Rochester, 200 First Street Southwest, Rochester, MN, 55905, USA; Division of Nephrology, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand. 2. Department of Laboratory Medicine and Pathology, Mayo Clinic Rochester, 200 First Street Southwest, Rochester, MN, 55905, USA. 3. Division of Nephrology and Hypertension, Department of Medicine, Mayo Clinic Rochester, 200 First Street Southwest, Rochester, MN, 55905, USA. 4. Immunochemical Core Laboratory, Mayo Clinic Rochester, 200 First Street Southwest, Rochester, MN, 55905, USA. 5. Division of Nephrology and Hypertension, Department of Medicine, Mayo Clinic Rochester, 200 First Street Southwest, Rochester, MN, 55905, USA; Department of Biochemistry and Molecular Biology, Mayo Clinic Rochester, 200 First Street Southwest, Rochester, MN, 55905, USA. Electronic address: rkumar@mayo.edu.
Abstract
BACKGROUND: Novel human parathyroid hormone (hPTH) peptides of unknown biological activity have recently been identified in the serum of subjects with normal renal function, chronic renal failure, and end-stage renal disease through the application of liquid chromatography-high resolution mass spectrometry. PURPOSE: of experiments: To determine the bioactivity of these peptides, we synthesized hPTH28-84, hPTH38-84, and hPTH45-84 peptides by solid phase peptide synthesis and tested their bioactivity in MC3T3-E1 mouse osteoblasts, either individually or together with the native hormone, hPTH1-84, by assessing the accumulation of 3´,5´-cyclic adenosine monophosphate (cAMP) and the induction of alkaline phosphatase activity. RESULTS: Increasing doses of hPTH1-84 (1-100 nM) increased the accumulation of cAMP and alkaline phosphatase activity in osteoblasts. hPTH28-84, hPTH38-84, and hPTH45-84 in concentrations of 1-100 nM were biologically inert. Surprisingly, 100 nM hPTH38-84 and hPTH45-84 increased the accumulation of cAMP in osteoblasts treated with increasing amounts of hPTH1-84. Human PTH28-84 had no effects on cAMP activity alone or in combination with hPTH1-84. Conversely, 100 nM hPTH38-84, hPTH45-84, and hPTH28-84 blocked the activation of alkaline phosphatase activity by hPTH1-84. CONCLUSIONS: The data show that the short carboxyl-terminal hPTH peptides, hPTH38-84 and hPTH45-84, increase the amount of cellular cAMP generated in cultured osteoblasts in response to treatment with full-length hPTH1-84 when compared to full-length hPTH1-84 alone. Human PTH28-84 had no effect on cAMP activity alone or in combination with hPTH1-84. Human PTH28-84, hPTH38-84 and hPTH45-84 reduced the effects of hPTH1-84 in osteoblasts with respect to the induction of alkaline phosphatase activity compared to hPTH1-84 alone. Short carboxyl peptides of human PTH are biologically inert but when administered together with full-length hPTH1-84 modulate the bioactivity of hPTH1-84 in osteoblasts.
BACKGROUND: Novel human parathyroid hormone (hPTH) peptides of unknown biological activity have recently been identified in the serum of subjects with normal renal function, chronic renal failure, and end-stage renal disease through the application of liquid chromatography-high resolution mass spectrometry. PURPOSE: of experiments: To determine the bioactivity of these peptides, we synthesized hPTH28-84, hPTH38-84, and hPTH45-84 peptides by solid phase peptide synthesis and tested their bioactivity in MC3T3-E1 mouse osteoblasts, either individually or together with the native hormone, hPTH1-84, by assessing the accumulation of 3´,5´-cyclic adenosine monophosphate (cAMP) and the induction of alkaline phosphatase activity. RESULTS: Increasing doses of hPTH1-84 (1-100 nM) increased the accumulation of cAMP and alkaline phosphatase activity in osteoblasts. hPTH28-84, hPTH38-84, and hPTH45-84 in concentrations of 1-100 nM were biologically inert. Surprisingly, 100 nM hPTH38-84 and hPTH45-84 increased the accumulation of cAMP in osteoblasts treated with increasing amounts of hPTH1-84. Human PTH28-84 had no effects on cAMP activity alone or in combination with hPTH1-84. Conversely, 100 nM hPTH38-84, hPTH45-84, and hPTH28-84 blocked the activation of alkaline phosphatase activity by hPTH1-84. CONCLUSIONS: The data show that the short carboxyl-terminal hPTH peptides, hPTH38-84 and hPTH45-84, increase the amount of cellular cAMP generated in cultured osteoblasts in response to treatment with full-length hPTH1-84 when compared to full-length hPTH1-84 alone. Human PTH28-84 had no effect on cAMP activity alone or in combination with hPTH1-84. Human PTH28-84, hPTH38-84 and hPTH45-84 reduced the effects of hPTH1-84 in osteoblasts with respect to the induction of alkaline phosphatase activity compared to hPTH1-84 alone. Short carboxyl peptides of human PTH are biologically inert but when administered together with full-length hPTH1-84 modulate the bioactivity of hPTH1-84 in osteoblasts.
Authors: Kittrawee Kritmetapak; Louis A Losbanos; Jolaine M Hines; Katherine L O'Grady; Candice Z Ulmer; Hubert W Vesper; Felicity T Enders; Ravinder J Singh; Rajiv Kumar Journal: Clin Chem Date: 2021-06-01 Impact factor: 8.327