Relationship between plasma fibroblast growth factor-23 concentration and survival time in cats with chronic kidney disease

BACKGROUND: Fibroblast growth factor‐23 (FGF‐23) and parathyroid hormone (PTH) are commonly increased in cats with azotemic chronic kidney disease (CKD). Both are predictors of survival time in human patients, but these relationships have not previously been examined in the cat. OBJECTIVES: To investigate the relationship between plasma FGF‐23 and PTH concentrations at diagnosis of CKD in cats with survival time and with disease progression over 12 months. ANIMALS: 214 azotemic, client‐owned cats (≥9 years). METHODS: Retrospective study: Biochemical and urinary variables at diagnosis of azotemic CKD, including plasma FGF‐23 and PTH concentrations were assessed as predictors of survival time (all‐cause mortality) using Cox regression, and as predictors of CKD progression over 12 months using logistic regression. RESULTS: In the final multivariable Cox regression model, survival was negatively associated with plasma creatinine (P = .002) and FGF‐23 concentrations (P = .014), urine protein‐to‐creatinine ratio (P < .001) and age (P < .001). Survival was positively associated with PCV (P = .004). In the final multivariable logistic regression model, independent predictors of CKD progression included logFGF‐23 and age. Neither plasma phosphate nor PTH was found to be an independent predictor of survival time or of CKD progression. CONCLUSIONS AND CLINICAL IMPORTANCE: Plasma FGF‐23 concentration is a novel prognostic indicator in cats with CKD, independent of other factors including plasma creatinine and phosphate concentrations. Further work is required to assess if FGF‐23 contributes directly to CKD progression, but regardless these findings may make FGF‐23 a useful biomarker for predicting poorer outcomes in cats with CKD

Evaluation of Methodologies for Microrna Biomarker Detection by Next Generation Sequencing

In recent years, microRNAs (miRNAs) in tissues and biofluids have emerged as a new class of promising biomarkers for numerous diseases. Blood-based biomarkers are particularly desirable since serum or plasma is easily accessible and can be sampled repeatedly. To comprehensively explore the biomarker potential of miRNAs, sensitive, accurate and cost-efficient miRNA profiling techniques are required. Next generation sequencing (NGS) is emerging as the preferred method for miRNA profiling; offering high sensitivity, single-nucleotide resolution and the possibility to profile a considerable number of samples in parallel. Despite the excitement about miRNA biomarkers, challenges associated with insufficient characterization of the sequencing library preparation efficacy, precision and method-related quantification bias have not been addressed in detail and are generally underappreciated in the wider research community. Here, we have tested in parallel four commercially available small RNA sequencing kits against a cohort of samples comprised of human plasma, human serum, murine brain tissue and a reference library containing ~ 950 synthetic miRNAs. We discuss the advantages and limits of these methodologies for massive parallel microRNAs profiling. This work can serve as guideline for choosing an adequate library preparation method, based on sensitivity, specificity and accuracy of miRNA quantification, workflow convenience and potential for automation

Deletion of PTH Rescues Skeletal Abnormalities and High Osteopontin Levels in Klotho−/− Mice

Maintenance of normal mineral ion homeostasis is crucial for many biological activities, including proper mineralization of the skeleton. Parathyroid hormone (PTH), Klotho, and FGF23 have been shown to act as key regulators of serum calcium and phosphate homeostasis through a complex feedback mechanism. The phenotypes of Fgf23−/− and Klotho−/− (Kl−/−) mice are very similar and include hypercalcemia, hyperphosphatemia, hypervitaminosis D, suppressed PTH levels, and severe osteomalacia/osteoidosis. We recently reported that complete ablation of PTH from Fgf23−/− mice ameliorated the phenotype in Fgf23−/−/PTH−/− mice by suppressing serum vitamin D and calcium levels. The severe osteomalacia in Fgf23−/− mice, however, persisted, suggesting that a different mechanism is responsible for this mineralization defect. In the current study, we demonstrate that deletion of PTH from Kl−/− (Kl−/−/PTH−/− or DKO) mice corrects the abnormal skeletal phenotype. Bone turnover markers are restored to wild-type levels; and, more importantly, the skeletal mineralization defect is completely rescued in Kl−/−/PTH−/− mice. Interestingly, the correction of the osteomalacia is accompanied by a reduction in the high levels of osteopontin (Opn) in bone and serum. Such a reduction in Opn levels could not be observed in Fgf23−/−/PTH−/− mice, and these mice showed sustained osteomalacia. This significant in vivo finding is corroborated by in vitro studies using calvarial osteoblast cultures that show normalized Opn expression and rescued mineralization in Kl−/−/PTH−/− mice. Moreover, continuous PTH infusion of Kl−/− mice significantly increased Opn levels and osteoid volume, and decreased trabecular bone volume. In summary, our results demonstrate for the first time that PTH directly impacts the mineralization disorders and skeletal deformities of Kl−/−, but not of Fgf23−/− mice, possibly by regulating Opn expression. These are significant new perceptions into the role of PTH in skeletal and disease processes and suggest FGF23-independent interactions of PTH with Klotho

FGF-23 in bone biology

Recent studies have demonstrated that levels of fibroblast growth factor 23 (FGF-23), a key regulator of phosphorus and vitamin D metabolism, rise dramatically as renal function declines and may play a key initiating role in disordered mineral and bone metabolism in patients with chronic kidney disease (CKD). The physiologic importance of FGF-23 in mineral metabolism was first identified in human genetic and acquired rachitic diseases and further characterized in animal models. FGF-23 and its regulators, including phosphate regulating endopeptidase homolog, dentin matrix 1 (DMP1), and matrix extracellular phosphoglycoprotein, are made primarily in bone, specifically in osteocytes. Dysregulation of these proteins results in osteomalacia, implicating the osteocyte in the regulation of skeletal mineralization. Studies in pediatric patients with CKD, the majority of whom have altered skeletal mineralization in early stages of CKD, have demonstrated that skeletal expression of both FGF-23 and its regulator, DMP1, are increased in early stages of CKD and that expression of these proteins is associated with alterations in skeletal mineralization. Thus, dysregulation of osteocytic proteins occur very early in the course of CKD and appear to be central to altered bone and mineral metabolism in this patient population