Persistent Link:
http://hdl.handle.net/10150/183848
Title:
RENAL METABOLISM AND ACTION OF VITAMIN-D3.
Author:
CHANDLER, JOHN STEWART.
Issue Date:
1982
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
The kidney represents a unique endocrine organ for the metabolism and action of vitamin D₃. Renal biosynthesis of 1,25-hydroxyvitamin D₃ (1,25(OH)₂D₃), and other vitamin D₃ metabolites, is regulated by ionic and hormonal factors. 1,25(OH)₂D₃ elicits a dramatic increase in 25-hydroxyvitamin D₃-24-hydroxylase (24-OHase) activity while concomitantly suppressing 25-hydroxyvitamin D₃-1α-hydroxylase (1-OHase) activity. To study this mechanism(s), biosynthesis and purification of the radiolabeled metabolites in question was necessary. Protocols for the selective induction of either 1-OHase or 24-OHase activity in chick kidney (in vivo) and the use of these kidneys (homogenates) for the biosynthesis of very high specific radioactivity metabolites, in vitro, are presented. A number of chromatographic systems including HPLC were developed and employed for the detection and purification of these biosynthetic metabolites (1,25(OH)₂[³H]D₃; 1,24,25(OH)₃[³H]D₃; 24,25(OH)₂[³H]D₃). Characterization of specific receptors for 1,25(OH)₂D₃ in the rat and chick kidney is presented and by comparison, these receptors appear similar to the chick intestinal component. 1,25(OH)₂D₃ receptors were also identified in a number of cultured mammalian cells. These receptors, irrespective of origin (tissue or species), selectively bind to immobilized DNA, a property which may indicate a physiologic function. Extensive metabolite binding studies indicate that 1,24,25(OH)₃D₃ is the second most active metabolite at the receptor level. To evaluate the role of the 1,25(OH)₂D₃ receptor in modulating the 1,25(OH)₂D₃-responsive activity of the renal 24-OHase enzyme, two renal cell lines (LLC-PK₁ and LLC-MK₂) were studied. A sensitive microassay for 24-OHase activity in intact cells was developed to monitor the enzyme responses. Although both cell lines contain 1,25(OH)₂D₃-responsive 24-OHase activity, only the LLC-PK₁ cells contain receptors for 1,25(OH)₂D₃. It is concluded that 1,25(OH)₂D₃ can rapidly and specifically regulate (induce) the activity of the 24-OHase enzyme in the absence of its specific receptor protein and this induction is inhibited by actinomycin D. Hence, although 1,25(OH)₂D₃ may act in renal cells to induce enzymes for the metabolism of vitamin D via its receptor component, this mechanism is not obligatory.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Cholecalciferol.; Kidneys.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Biochemistry; Graduate College
Degree Grantor:
University of Arizona

Full metadata record

DC FieldValue Language
dc.language.isoenen_US
dc.titleRENAL METABOLISM AND ACTION OF VITAMIN-D3.en_US
dc.creatorCHANDLER, JOHN STEWART.en_US
dc.contributor.authorCHANDLER, JOHN STEWART.en_US
dc.date.issued1982en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractThe kidney represents a unique endocrine organ for the metabolism and action of vitamin D₃. Renal biosynthesis of 1,25-hydroxyvitamin D₃ (1,25(OH)₂D₃), and other vitamin D₃ metabolites, is regulated by ionic and hormonal factors. 1,25(OH)₂D₃ elicits a dramatic increase in 25-hydroxyvitamin D₃-24-hydroxylase (24-OHase) activity while concomitantly suppressing 25-hydroxyvitamin D₃-1α-hydroxylase (1-OHase) activity. To study this mechanism(s), biosynthesis and purification of the radiolabeled metabolites in question was necessary. Protocols for the selective induction of either 1-OHase or 24-OHase activity in chick kidney (in vivo) and the use of these kidneys (homogenates) for the biosynthesis of very high specific radioactivity metabolites, in vitro, are presented. A number of chromatographic systems including HPLC were developed and employed for the detection and purification of these biosynthetic metabolites (1,25(OH)₂[³H]D₃; 1,24,25(OH)₃[³H]D₃; 24,25(OH)₂[³H]D₃). Characterization of specific receptors for 1,25(OH)₂D₃ in the rat and chick kidney is presented and by comparison, these receptors appear similar to the chick intestinal component. 1,25(OH)₂D₃ receptors were also identified in a number of cultured mammalian cells. These receptors, irrespective of origin (tissue or species), selectively bind to immobilized DNA, a property which may indicate a physiologic function. Extensive metabolite binding studies indicate that 1,24,25(OH)₃D₃ is the second most active metabolite at the receptor level. To evaluate the role of the 1,25(OH)₂D₃ receptor in modulating the 1,25(OH)₂D₃-responsive activity of the renal 24-OHase enzyme, two renal cell lines (LLC-PK₁ and LLC-MK₂) were studied. A sensitive microassay for 24-OHase activity in intact cells was developed to monitor the enzyme responses. Although both cell lines contain 1,25(OH)₂D₃-responsive 24-OHase activity, only the LLC-PK₁ cells contain receptors for 1,25(OH)₂D₃. It is concluded that 1,25(OH)₂D₃ can rapidly and specifically regulate (induce) the activity of the 24-OHase enzyme in the absence of its specific receptor protein and this induction is inhibited by actinomycin D. Hence, although 1,25(OH)₂D₃ may act in renal cells to induce enzymes for the metabolism of vitamin D via its receptor component, this mechanism is not obligatory.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectCholecalciferol.en_US
dc.subjectKidneys.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineBiochemistryen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.identifier.proquest8217494en_US
dc.identifier.oclc681974363en_US
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