Journal: Scientific Reports

Article Title: FGF23, a novel muscle biomarker detected in the early stages of ALS

doi: 10.1038/s41598-021-91496-6

Figure Lengend Snippet: FGF23 mRNA levels are increased in ALS muscle tissue. FGF23 mRNA levels were assessed in human muscle samples by qPCR using GAPDH as an internal housekeeping control. Disease samples were expressed as a fold-change (mean ± SEM) compared to normal control tissue (set at 1). *** P = 0.0001. BI, biceps brachii; DL, deltoid; GC, gastrocnemius; Myo, myopathy disease control; neuro, neuropathy disease control, TA, tibialis anterior; VL, vastus lateralis.

Article Snippet: Human plasma FGF-23 was analyzed using U-PLEX Human FGF-23 Assay (K1516EK, MSD), and mouse plasma FGF23 was measure using mouse/rat FGF23 (intact) ELISA kit (60-6800, Quidel) according to the manufacturer’s instruction.

Techniques: Control

Journal: Scientific Reports

Article Title: FGF23, a novel muscle biomarker detected in the early stages of ALS

doi: 10.1038/s41598-021-91496-6

Figure Lengend Snippet: FGF23 protein is increased in ALS muscle tissue. Sections from 6 ALS and 4 normal muscle biopsy samples were immunostained with an anti-FGF23 antibody and counterstained with Hoechst and wheat germ agglutin (WGA). All 6 ALS patient samples but no control samples showed positive staining. Three of the ALS and two of the normal control sections are shown here. ALSp1 (deltoid), ALSp2 (vastus lateralis), ALSp3 (vastus lateralis), Ctrl1 (deltoid), Ctrl 2 (vastus lateralis). Asterisks highlight areas of grouped atrophy and arrowheads highlight several of the loci where FGF23 and WGA immunostaining colocalizes. Scale bars, 100 μm.

Article Snippet: Human plasma FGF-23 was analyzed using U-PLEX Human FGF-23 Assay (K1516EK, MSD), and mouse plasma FGF23 was measure using mouse/rat FGF23 (intact) ELISA kit (60-6800, Quidel) according to the manufacturer’s instruction.

Techniques: Control, Staining, Immunostaining

Journal: Scientific Reports

Article Title: FGF23, a novel muscle biomarker detected in the early stages of ALS

doi: 10.1038/s41598-021-91496-6

Figure Lengend Snippet: FGF23 immunoreactivity is higher in areas of grouped atrophy in human ALS muscle tissue. Using ImageJ Fluorescence Intensity (FI) Analysis function, we compared the FGF23 FI (per μM 2 ) of 37 atrophic fibers (< 25 μM minimal feret’s diameter) sampled from areas of grouped atrophy seen in 5 human ALS muscle samples with 37 non-atrophic fibers (> 25 μM minimal feret’s diameter) from the same sections. Representative photomicrographs of patient ALSp1 are shown with regions of interest highlighted for 6 grouped atrophic fibers (AF) and 6 non-atrophic fibers (NAF). An FGF23 intensity ratio was calculated by dividing the FI in atrophic fibers by the FI in non-atrophic factors for each patient. A ratio was also calculated between a similar number of non-atrophic fibers in the same section to the NAF region of interest as a control. The FI ratio was nearly sixfold higher in areas of grouped atrophy versus non-atrophic fibers. ** P = 0.006. Scale bar, 100 μm.

Article Snippet: Human plasma FGF-23 was analyzed using U-PLEX Human FGF-23 Assay (K1516EK, MSD), and mouse plasma FGF23 was measure using mouse/rat FGF23 (intact) ELISA kit (60-6800, Quidel) according to the manufacturer’s instruction.

Techniques: Fluorescence, Control

Journal: Scientific Reports

Article Title: FGF23, a novel muscle biomarker detected in the early stages of ALS

doi: 10.1038/s41598-021-91496-6

Figure Lengend Snippet: FGF23 is increased in SOD1 G93A muscle. ( A ) The clinical timeline of disease progression in the SOD1 G93A mouse is shown above . Below is a qPCR analysis of gastrocnemius muscle samples from littermate controls (WT) and SOD1 G93A mice at different ages as indicated. Data points are the mean ± SEM of 6–8 mice. * P < 0.05, *** P < 0.0005. ( B ) Photomicrographs of gastrocnemius muscle sections from a WT and SOD1 G93A mouse (60 d) immunostained with an anti-FGF23 antibody and counterstained with Hoechst and WGA. Scale bar, 100 μm. Arrowheads highlight several areas of merged FGF23 and WGA staining. ( C ) ELISA analysis of FGF23 in plasma samples obtained at the ages indicated. Data points are the mean ± SEM of 3 mice per group. ** P < 0.01.

Article Snippet: Human plasma FGF-23 was analyzed using U-PLEX Human FGF-23 Assay (K1516EK, MSD), and mouse plasma FGF23 was measure using mouse/rat FGF23 (intact) ELISA kit (60-6800, Quidel) according to the manufacturer’s instruction.

Techniques: Biomarker Discovery, Staining, Enzyme-linked Immunosorbent Assay, Clinical Proteomics

Journal: Scientific Reports

Article Title: FGF23, a novel muscle biomarker detected in the early stages of ALS

doi: 10.1038/s41598-021-91496-6

Figure Lengend Snippet: Plasma FGF23 concentration.

Article Snippet: Human plasma FGF-23 was analyzed using U-PLEX Human FGF-23 Assay (K1516EK, MSD), and mouse plasma FGF23 was measure using mouse/rat FGF23 (intact) ELISA kit (60-6800, Quidel) according to the manufacturer’s instruction.

Techniques: Clinical Proteomics, Concentration Assay, Control

Journal: Scientific Reports

Article Title: FGF23, a novel muscle biomarker detected in the early stages of ALS

doi: 10.1038/s41598-021-91496-6

Figure Lengend Snippet: FGF23 in human plasma samples from ALS patients and healthy controls. ( A ) Baseline levels of log-transformed plasma FGF23 concentration (pg/ml) among controls, and faster and slower progressing ALS patients. Boxes show median (middle line), and 25th and 75th percentiles (lower and upper border, respectively); whiskers extend to a maximum of 1.5 × interquartile range (IQR), or to the most extreme value if it is less than 1.5 × IQR from the 25th or 75th percentile. ( B ) Longitudinal changes in log-transformed plasma FGF23 among controls. ( C ) Longitudinal changes in log-transformed plasma FGF23 among ALS slower progressors (ALSFRS-R decline < 0.8 point/month). ( D ) Longitudinal changes in log-transformed plasma FGF23 among ALS faster progressors (ALSFRS-R decline > 1.2 points/month).

Article Snippet: Human plasma FGF-23 was analyzed using U-PLEX Human FGF-23 Assay (K1516EK, MSD), and mouse plasma FGF23 was measure using mouse/rat FGF23 (intact) ELISA kit (60-6800, Quidel) according to the manufacturer’s instruction.

Techniques: Clinical Proteomics, Transformation Assay, Concentration Assay

Journal: The FASEB Journal

Article Title: High phosphate‐induced progressive proximal tubular injury is associated with the activation of Stat3/Kim‐1 signaling pathway and macrophage recruitment

doi: 10.1096/fj.202200167rr

Figure Lengend Snippet: FIGURE 1 Chronic high phosphate intake causes hyperphosphatemia and increases Fgf23 and PTH levels. (A) Schematic representation of experimental setup. Eight-week-old male C57BL/6N mice were fed with a 2% high phosphate diet (HPD) or 0.8% phosphate control diet (Ctrl) for one up to six months. (B) Measurement of serum phosphate and (C) quantification of urinary phosphate to creatinine ratio were performed to analyze phosphate homeostasis. Measurement of (D) intact fibroblast growth factor 23 (iFgf23) and (E) parathyroid hormone (PTH) in plasma by ELISA was done to quantify the levels of phosphaturic hormones. (F) Measurement of serum calcium and (G) quantification of urinary calcium to creatinine ratio were performed to analyze calcium homeostasis.

Article Snippet: Plasma intact FGF23 (#60– 6800) and PTH 1– 84 (#60– 2305) were quantified by murine ELISA (both Quidel, San Diego, CA, USA).

Techniques: Control, Clinical Proteomics, Enzyme-linked Immunosorbent Assay

Journal: The FASEB Journal

Article Title: High phosphate‐induced progressive proximal tubular injury is associated with the activation of Stat3/Kim‐1 signaling pathway and macrophage recruitment

doi: 10.1096/fj.202200167rr

Figure Lengend Snippet: FIGURE 7 High phosphate activates Stat3/Kim-1 signaling in human proximal tubular epithelial cells. (A and B) Quantitative RT-PCR analysis of HAVCR1 (A) and CCL2 (B) in human proximal tubular epithelial (HK-2) cells stimulated with fibroblast growth factor 23 (Fgf23), parathyroid hormone (PTH) or increasing concentrations of phosphate (P) for 48 h. (C) Representative immunoblots of pStat3, Stat3 and Gapdh using protein lysates of human proximal tubular epithelial cells stimulated with fibroblast growth factor 23 (Fgf23), parathyroid hormone (PTH) or two different concentrations of phosphate for 10 min. (D) Representative immunofluorescence images of Kim-1 (orange) and pStat3 (green) co-stained HK-2 cells after stimulation with Fgf23, PTH or two different concentrations of phosphate for 48 h. Cell nuclei were counterstained with DAPI (blue) and actin filaments were stained with phalloidin (white). Scale bar: 20 μm.

Article Snippet: Plasma intact FGF23 (#60– 6800) and PTH 1– 84 (#60– 2305) were quantified by murine ELISA (both Quidel, San Diego, CA, USA).

Techniques: Quantitative RT-PCR, Western Blot, Immunofluorescence, Staining

Journal: The FASEB Journal

Article Title: High phosphate‐induced progressive proximal tubular injury is associated with the activation of Stat3/Kim‐1 signaling pathway and macrophage recruitment

doi: 10.1096/fj.202200167rr

Figure Lengend Snippet: FIGURE 8 Schematic representation of systemic and renal effects of high inorganic phosphate intake. (A) The enhanced consumption of food containing considerable amounts of inorganic phosphates induces fibroblast growth factor 23 (FGF23) and parathyroid hormone (PTH) levels and causes hyperphosphatemia as well as hyperphosphaturia. Moreover, it promotes progressive tubular injury and tubulointerstitial fibrosis with severe macrophage recruitment. (B) High dietary phosphate intake in health promotes proximal tubular epithelial cell damage with loss of cell polarity, flattened epithelia, disrupted brush border membranes, and vacuolization. The marked interstitial infiltration of macrophages located around proximal tubular lesions is accompanied by increased accumulation of collagen fibers. (C) High phosphate stimulates KIM-1 expression via phosphorylation and thereby activation of STAT3 in proximal tubule cells. Increased KIM-1 synthesis enhances MCP-1 expression in proximal tubule cells leading to enhanced macrophage recruitment. Concomitantly, activation of STAT3/KIM-1 signaling upregulates pro-fibrotic gene expressions that promote tubulointerstitial fibrosis. Altogether this leads to the progression of tubular injury. This figure is created with BioRender.com.

Article Snippet: Plasma intact FGF23 (#60– 6800) and PTH 1– 84 (#60– 2305) were quantified by murine ELISA (both Quidel, San Diego, CA, USA).

Techniques: Expressing, Phospho-proteomics, Activation Assay