Biomedical subjects

Peter A Friedman

Publications and source records attributed to Peter A Friedman.

12 recordsLinked to original sources

PubMed2025

Network Interactions of Circulating FGF23, HRG-HMGB1, and Cardiac Disease in CKD.

KEY POINTS: Multitrait analysis of genome-wide association study boosts the statistical power to identify novel genetic traits for fibroblast growth factor 23. A functional genomics approach aided network discovery to identify histidine-rich glycoprotein (HRG) and high-mobility group protein box 1 (HMGB1) as key regulators of cardiac disease in CKD. Integration of clinical and genetic data enhances the discovery power and is crucial for understanding the genetic underpinnings of mineral bone disorder related to CKD. BACKGROUND: Genome-wide association studies (GWAS) have identified numerous genetic loci associated with mineral metabolism markers but have exclusively focused on single-trait analysis. In this study, we performed a multitrait analysis of GWAS (MTAG) of mineral metabolism, exploring overlapping genetic architecture between traits to identify novel genetic associations for fibroblast growth factor 23 (FGF23). METHODS: We applied MTAG to variants common to GWAS of five genetically correlated mineral metabolism markers in participants of European ancestry. We integrated UK Biobank GWAS for blood levels for phosphate, 25-hydroxyvitamin D, and calcium (n=366,484) and Cohorts for Heart and Aging Research in Genetic Epidemiology GWAS for parathyroid hormone (n=29,155) and FGF23 (n=13,716). We then used supervised and unsupervised deep machine learning to identify novel associations between genetic traits and FGF23. RESULTS: MTAG increased the effective sample size for mineral metabolism markers to n=50,325 for FGF23. After clumping, MTAG identified independent genome-wide significant single-nucleotide polymorphisms for all traits, including 62 loci for FGF23. Many of these loci have not been previously reported in single-trait analyses. Through a functional genomics approach, we identified histidine-rich glycoprotein (HRG) and high-mobility group box 1 (HMGB1) as master regulators of downstream canonical pathways associated with circulating FGF23, and both genes were highly enriched in hypertrophied cardiac tissue of deceased hemodialysis patients. In addition, we found that DNMT3A was associated with uremic toxin, 8-hydroxy-2-deoxyguanosine, a biomarker of DNA damage. In silico gene perturbation analysis revealed that DNMT3A is protective in patients with heart failure caused by hypertrophied or dilated cardiomyopathy. CONCLUSIONS: Our findings highlight the importance of MTAG analysis of mineral metabolism markers to boost the number of genome-wide significant loci for FGF23 to identify novel genetic traits. Functional genomics revealed novel networks that inform unique cellular functions and identified HRG and HMGB1 as key master regulators of FGF23 and cardiovascular disease in CKD.

bones, stones, and mineral metabolism↗

PubMed2006

Extracellular signal-regulated kinase activation by parathyroid hormone in distal tubule cells.

The PTH receptor (PTH1R) activates multiple signaling pathways, including extracellular signal-regulated kinases 1 and 2 (ERK1/2). The role of epidermal growth factor receptor (EGFR) transactivation in ERK1/2 activation by PTH in distal kidney cells, a primary site of PTH action, was characterized. ERK1/2 phosphorylation was stimulated by PTH and blocked by the EGFR inhibitor, AG1478. Upon PTH stimulation, metalloprotease cleavage of membrane-bound heparin-binding fragment (HB-EGF) induced EGFR transactivation of ERK. Conditioned media from PTH-treated distal kidney cells activated ERK in HEK-293 cells. AG1478 added to HEK-293 cells ablated transactivation by conditioned media. HB-EGF directly activated ERK1/2 in HEK-293 cells. Pretreatment of distal kidney cells with the metalloprotease inhibitor GM-6001 abolished transactivation of ERK1/2 by PTH. The role of the PTH1R COOH terminus in PTX-sensitive ERK1/2 activation was characterized in HEK-293 cells transfected with wild-type PTH1R, with a PTH1R mutated at its COOH terminus, or with PTH1R truncated at position 480. PTH stimulated ERK by wild-type, mutated and truncated PTH1Rs 21-, 27- and 57-fold, respectively. Thus, the PTH1R COOH terminus exerts an inhibitory effect on ERK activation. EBP50, a scaffolding protein that binds to the PDZ recognition domain of the PTH1R, impaired PTH but not isoproterenol or calcitonin-induced ERK activation. Pertussis toxin inhibited PTH-stimulated ERK1/2 by mutated and truncated PTH1Rs and abolished ERK1/2 activation by wild-type PTH1R. We conclude that ERK phosphorylation in distal kidney cells by PTH requires PTH1R activation of G(i), which leads to stimulation of metalloprotease-mediated cleavage of HB-EGF and transactivation of the EGFR and is regulated by EBP50.

Animals↗

PubMed2006

The association of NHERF adaptor proteins with g protein-coupled receptors and receptor tyrosine kinases.

The sodium-hydrogen exchanger regulatory factors (NHERF-1 and NHERF-2) are a family of adaptor proteins characterized by the presence of two tandem PDZ protein interaction domains and a C-terminal domain that binds the cytoskeleton proteins ezrin, radixin, moesin, and merlin. The NHERF proteins are highly expressed in the kidney, small intestine, and other organs, where they associate with a number of transporters and ion channels, signaling proteins, and transcription factors. Recent evidence has revealed important associations between the NHERF proteins and several G protein-coupled receptors such as the beta2-adrenergic receptor, the kappa-opioid receptor, and the parathyroid hormone receptor, as well as growth factor tyrosine kinase receptors such as the platelet-derived growth factor receptor and the epidermal growth factor receptor. This review summarizes the emerging data on the biochemical mechanisms, physiologic outcomes, and potential clinical implications of the assembly and disassembly of receptor/NHERF complexes.

Animals↗

PubMed2006

PTH(1-84)/PTH(7-84): a balance of power.

This review considers many new basic and clinical aspects of parathyroid hormone (PTH). We focus especially on the identification of PTH fragments and how they may relate to renal failure, diagnosis, and treatment of secondary hyperparathyroidism and renal osteodystrophy. The biosynthesis and metabolism of PTH, measurement of circulating forms of PTH, the effects of PTH on receptor activation and turnover, the relationship between PTH levels and bone turnover in renal failure in humans, and the involvement of PTH in experimental models of renal failure are discussed. Despite these developments in understanding the etiology of renal failure and the availability of new assays for bioactive PTH, no adequate surrogate for bone biopsy and quantitative bone histomorphometry has been developed.

Amino Acid Sequence↗

PubMed2004

Parathyroid hormone receptor trafficking contributes to the activation of extracellular signal-regulated kinases but is not required for regulation of cAMP signaling.

Agonist-mediated activation of the type 1 parathyroid hormone receptor (PTH1R) results in several signaling events and receptor endocytosis. It is well documented that arrestins contribute to desensitization of both G(s)- and G(q)-mediated signaling and mediate PTH1R internalization. However, whether PTH1R trafficking directly contributes to signaling remains unclear. To address this question, we investigated the role of PTH1R trafficking in cAMP signaling and activation of extracellular signal-regulated kinases ERK1/2 in HEK-293 cells. Dominant negative forms of dynamin (K44A-dynamin) and beta-arrestin1 (beta-arrestin1-(319-418)) abrogated PTH1R internalization but had no effect on cAMP signaling; neither acute cAMP production by PTH nor desensitization and resensitization of cAMP signaling were affected. Therefore, PTH1R trafficking is not necessary for regulation of cAMP signaling. PTH-(1-34) induced rapid and robust activation of ERK1/2. A PTHrP-based analog ([p-benzoylphenylalanine1, Ile5,Arg(11,13),Tyr36]PTHrP-(1-36)NH2), which selectively activates the G(s)/cAMP pathway without inducing PTH1R endocytosis, failed to stimulate ERK1/2 activity. Inhibition of PTH1R endocytosis by K44A-dynamin dampened ERK1/2 activation in response to PTH-(1-34) by 69%. Incubation with the epidermal growth factor receptor inhibitor AG1478 reduced ERK1/2 phosphorylation further. In addition, ERK1/2 phosphorylation occurred following internalization of a PTH1R mutant induced by PTH-(7-34) in the absence of G protein signaling. Collectively, these data indicate that PTH1R trafficking and G(q) (but not G(s)) signaling independently contribute to ERK1/2 activation, predominantly via transactivation of the epidermal growth factor receptor.

Arrestins↗

PubMed2004

Ligand-selective dissociation of activation and internalization of the parathyroid hormone (PTH) receptor: conditional efficacy of PTH peptide fragments.

G protein-coupled receptors (GPCRs) mediate the action of many hormones, cytokines, and sensory and chemical signals. It is generally thought that receptor desensitization and internalization require occupancy and activation of the GPCR. PTH and PTHrP receptor (PTH1R) belongs to GPCR class B and is the major regulator of extracellular calcium homeostasis. Using kidney distal convoluted tubule cells transfected with a human PTH1R/enhanced green fluorescent protein fusion protein, quantitative, real-time fluorescence microscopy was used to analyze receptor internalization. In these cells, which are the target of the calcium-sparing action of PTH, PTH(1-34) activated adenylyl cyclase (AC) and phospholipase C (PLC) and PTH1R endocytosis. PTH(1-31), however, stimulated AC and PLC but not PTH1R endocytosis. Conversely, PTH(7-34) rapidly stimulated PTH1R internalization without activating AC or PLC. PTH(2-34) and (3-34) caused PTH1R internalization intermediate between PTH(1-34) and (7-34). PTH1R sequestration occurred in a dynamin- and clathrin-dependent manner. Directly activating AC inhibited PTH1R internalization in response to PTH(7-34). PTH1R endocytosis was sensitive to protein kinase C inhibition. PTH(1-34), (7-34), and (1-31) evoked PTH1R phosphorylation. Removal of most of the C terminus of the PTH1R eliminated receptor phosphorylation and the cAMP/protein kinase C sensitivity of internalization. PTH(1-34) and (7-34) internalized the truncated PTH1R with identical kinetics, and the response was unaffected by forskolin. Thus, the PTH1R C terminus contains regulatory sequences that are involved in, but not required for, PTH1R internalization. The results demonstrate that receptor activation and internalization can be selectively dissociated.

Animals↗

PubMed2004

Calcium-sensing receptor regulation of renal mineral ion transport.

Extracellular calcium has long been known to affect the rate and magnitude of renal calcium and phosphate recovery. In this review, we consider some of these findings in light of our present understanding of the tubular localization of the calcium-sensing receptor (CaSR). Experiments directly implicating the CaSR in regulating calcium and phosphate transport are described. These results point to an important role of the CaSR in regulating PTH-dependent calcium absorption by cortical thick ascending limbs and on PTH-sensitive proximal tubule phosphate transport. Possible avenues for further investigation are suggested.

Animals↗

PubMed2004

PTH revisited.

Recent investigations of parathyroid hormone (PTH) have advanced our understanding of its circulating forms as well as its action. It is now clear that first-generation immunoradiometric assays of so-called intact "PTH" not only measured full-length PTH(1-84) but also recognized large PTH fragments lacking the amino-terminus. New, second generation assays detect only full-length PTH. Under diverse pathological settings, second generation assays display lower levels of PTH (1-84). By measuring full-length PTH (bioactive PTH) and the combined full-length plus amino-terminal PTH fragments, the amount of non-PTH(1-84) in circulation can be estimated. The primary amino-terminal fragment is likely to be PTH(7-84). A considerable controversy surrounds the pathological significance of PTH(7-84) and its relation to adynamic bone disease. While these findings were emerging, other work uncovered the apparent basis by which PTH receptors signal through cAMP in some instances but through Ca/inositol phosphate in others. This signaling switch is dictated by the cytoplasmic adapter protein NHERF1 (EBP50), which is expressed in a cell-selective fashion. Other provocative findings may provide a means of unifying determinations of PTH(7-84) with the effects of NHERF1 on PTH receptor signaling. These latter studies reveal that in cells expressing NHERF1, PTH(7-84) has no effect on PTH receptor signaling or internalization. However, in cells lacking or expressing low levels of NHERF1, PTH(7-84) internalizes the PTH receptor without accompanying activation. Together, these findings suggest that the accumulation of PTH(7-84) in renal failure may lead to PTH resistance by internalizing and down-regulating PTH receptors.

Bone Diseases↗

PubMed2003

Calcium-sensing receptor regulation of PTH-inhibitable proximal tubule phosphate transport.

Inorganic phosphate (Pi) is absorbed by proximal tubules through a cellular pathway that is inhibited by parathyroid hormone (PTH). The calcium-sensing receptor (CaSR) is expressed on apical membranes of proximal tubules. In the present studies, we determined the effect of luminal and/or basolateral PTH on phosphate absorption and tested the hypothesis that CaSR activation blocks PTH-inhibitable phosphate absorption. Single proximal S3 tubules were dissected from the kidneys of mice and studied by the Burg technique. Tubules were bathed with DMEM culture media supplemented with 6% BSA and perfused with an ultrafiltrate prepared from the bathing solution. 33P and FITC-inulin were added to the luminal perfusate to measure phosphate absorption (JPi) and fluid absorption (Jv), respectively. JPi averaged 2.9 pmol.min-1.mm-1 under control conditions and decreased by 20% upon addition of serosal PTH. PTH had no effect on Jv. Inclusion of PTH in the luminal perfusate reduced JPi to 2.1 pmol. min-1. mm-1. Combined addition of PTH to perfusate and bathing solutions reduced JPi to 1.5 pmol. min-1. mm-1 without affecting Jv. Indirect immunofluorescence studies revealed abundant PTH receptor (PTH1R) expression on brush-border membranes, with lower amounts on basolateral membranes. CaSRs were localized primarily, but not exclusively, to brush-border membranes. CaSR activation with luminal Gd3+ abolished the inhibitory action of PTH on JPi. Addition of Gd3+ to the serosal bathing solution had no effect on PTH-sensitive JPi. Gd3+ i.e., PTH-independent JPi. Gd3+ did not affect basal, had no effect on Jv when added to lumen or bath. Dopamine-inhibitable JPi was not affected by Gd3+. Experiments with proximal-like opossum kidney cells showed that elevated extracellular Ca2+ or NPS R467, a type II calcimimetic, inhibited PTH action on Pi uptake. In conclusion, PTH1Rs are expressed on apical and basolateral membranes of mouse proximal tubules. Stimulating apical or basolateral PTH1R inhibits phosphate absorption. CaSR activation specifically regulates PTH-suppressible phosphate absorption.

Animals↗

PubMed2003

Activation-independent parathyroid hormone receptor internalization is regulated by NHERF1 (EBP50).

Parathyroid hormone (PTH) regulates extracellular calcium homeostasis through the type 1 PTH receptor (PTH1R) expressed in kidney and bone. The PTH1R undergoes beta-arrestin/dynamin-mediated endocytosis in response to the biologically active forms of PTH, PTH-(1-34), and PTH-(1-84). We now show that amino-truncated forms of PTH that do not activate the PTH1R nonetheless induce PTH1R internalization in a cell-specific pattern. Activation-independent PTH1R endocytosis proceeds through a distinct arrestin-independent mechanism that is operative in cells lacking the adaptor protein Na/H exchange regulatory factor 1 (NHERF1) (ezrin-binding protein 50). Using a combination of radioligand binding experiments and quantitative, live cell confocal microscopy of fluorescently tagged PTH1Rs, we show that in kidney distal tubule cells and rat osteosarcoma cells, which lack NHERF1, the synthetic antagonist PTH-(7-34) and naturally circulating PTH-(7-84) induce internalization of PTH1R in a beta-arrestin-independent but dynamin-dependent manner. Expression of NHERF1 in these cells inhibited antagonist-induced endocytosis. Conversely, expression of dominant-negative forms of NHERF1 conferred internalization sensitivity to PTH-(7-34) in cells expressing NHERF1. Mutation of the PTH1R PDZ-binding motif abrogated interaction of the receptor with NHERF1. These mutated receptors were fully functional but were now internalized in response to PTH-(7-34) even in NHERF1-expressing cells. Removing the NHERF1 ERM domain or inhibiting actin polymerization allowed otherwise inactive ligands to internalize the PTH1R. These results demonstrate that NHERF1 acts as a molecular switch that legislates the conditional efficacy of PTH fragments. Distinct endocytic pathways are determined by NHERF1 that are operative for the PTH1R in kidney and bone cells.

Actins↗

PubMed2002

Plasma membrane Ca2+-ATPase and NCX1 Na+/Ca2+ exchanger expression in distal convoluted tubule cells.

The plasma membrane Ca2+-ATPase (PMCA) and the NCX1 Na+/Ca2+ exchanger regulate intracellular Ca2+ concentrations and mediate Ca2+ efflux in absorptive epithelial cells. We characterized the PMCA isoforms and subtypes expressed in mouse distal convoluted tubule (mDCT) cells and Na+/Ca2+ exchanger protein expression in mDCT cells. In lysates of mDCT cells, immunoprecipitation and Western blot analysis, performed with a monoclonal antibody to PMCA, revealed a 140-kDa protein consistent with PMCA. Laser-scanning confocal fluorescence microscopy indicated that PMCA and NCX1 expression is restricted to basolateral membranes only in confluent mDCT cells, because subconfluent cultures predominately express intracellular localizations. PMCA isoform-specific PCR primers generated appropriately sized products only for PMCA1 and PMCA4 from DCT cells but PMCA1-4 from whole mouse kidney. Assessment of splice site C within the calmodulin-binding domain demonstrated the presence of PMCA1b and PMCA4b mRNAs in mDCT cells. Northern blot analysis of mDCT cell RNA revealed transcripts of 7.5 and 5.5 kb for PMCA1 and 8.5 and 7.5 kb for PMCA4. We conclude that DCT cells express PMCA transcripts encoding PMCA1b and PMCA4b. Basolateral localization of the Na+/Ca2+ exchanger and MCAs support the idea that multiple PMCA isoforms, in concert with the Na+/Ca2+ exchanger, mediate basal or hormone-stimulated Ca2+ efflux by distal tubules.

Animals↗

PubMed2002

Calcium-sensing receptor regulation of PTH-dependent calcium absorption by mouse cortical ascending limbs.

Resting Ca(2+) absorption by cortical thick ascending limbs (CALs) is passive and proceeds through the paracellular pathway. In contrast, parathyroid hormone (PTH) stimulates active, transcellular Ca(2+) absorption (J(Ca)). The Ca(2+)-sensing receptor (CaSR) is expressed on serosal membranes of CALs. In the present study, we tested the hypothesis that activation of the CAL CaSR indirectly inhibits passive Ca(2+) transport and directly suppresses PTH-induced cellular J(Ca). To test this theory, we measured J(Ca) and Na absorption (J(Na)) by single perfused mouse CALs. Net absorption was measured microfluorimetrically in samples collected from tubules perfused and bathed in symmetrical HEPES-buffered solutions or those in which luminal Na(+) was reduced from 150 to 50 mM. We first confirmed that Gd(3+) activated the CaSR by measuring intracellular Ca(2+) concentration ([Ca(2+)](i)) in CALs loaded with fura 2. On stepwise addition of Gd(3+) to the bath, [Ca(2+)](i) increased, with a half-maximal rise at 30 microM Gd(3+). J(Ca) and transepithelial voltage (V(e),) were measured in symmetrical Na(+)-containing solutions. PTH increased J(Ca) by 100%, and 30 microM Gd(3+) inhibited this effect. V(e) was unchanged by either PTH or Gd(3+). Similarly, NPS R-467, an organic CaSR agonist, inhibited PTH-stimulated J(Ca) without altering V(e). Neither PTH nor Gd(3+) affected J(Na). Addition of bumetanide to the luminal perfusate abolished J(Na) and V(e). These results show that CaSR activation directly inhibited PTH-induced transcellular J(Ca) and that cellular Ca(2+) and Na(+) transport can be dissociated. To test the effect of CaSR activation on passive paracellular Ca(2+) transport, J(Ca) was measured under asymmetrical Na conditions, in which passive Ca(2+) transport dominates transepithelial absorption. PTH stimulated J(Ca) by 24% and was suppressed by Gd(3+). In this setting, Gd(3+) reduced V(e) by 32%, indicating that CaSR activation inhibited both transcellular and paracellular Ca(2+) transport. We conclude that the CaSR regulates both active transcellular and passive paracellular Ca(2+) reabsorption but has no effect on J(Na) by CALs.

Absorption↗