fgfr3  (R&D Systems)

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Nine to fourteen somite mouse embryos, cultured for 24 h. (A) Control embryo, exposed to the solvent dimethyl sulfoxide (DMSO) only, reveals age-appropriate telencephalic hemispheres (green asterisk) and upper limb buds (green arrowheads); for comparison (green arrow), see corresponding hematoxylin stained histological sections (green colored areas). (B) Treatment with 2.5 μM of the pan-FGFR inhibitor PD173074 considerably impairs the development of telencephalic hemispheres (red asterisk) and upper limb buds (red arrowheads); for comparison (red arrow), see corresponding hematoxylin stained histological sections (red colored areas). (C) Application of 0.5 μM PD173074 did not cause any obvious abnormalities. (D) Embryos incubated with 40 μM of the pan-FGFR inhibitor SU5402 exhibit general growth disturbances (also see ). (E,F) Treatment with 40 μg/ml of anti-FGFR3 neutralizing antibodies or 10 μM of the pan-BMPR inhibitor LDN193189 were well tolerated. Scale bars: 500 μm for stereomicrographs (A–F) , 200 μm for light micrographs (A,B) .

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Cell Culture, Control, Solvent, Comparison, Staining, Incubation

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Impact of FGFR and BMPR inhibition on epibranchial placode morphogenesis and neurogenesis in 9–14 somite mouse embryos, cultured for 24 h. Anti-Neurogenin-2 (Ngn2) immunohistochemistry (brown precipitate); shown are maximum numbers of Ngn2 + neuroblasts found per section. (A–C) Control placodes reveal high-grade thickened pseudostratified epithelium (2–4 cell nuclei) with 12–16 Ngn2 + neuroblasts. (D–F) Pan-FGFR inhibitor SU5402 (40 μM) causes moderate reductions in placode thickness and statistically significant decreases in neuroblast numbers in the epibranchial placodes 1 and 2, but not 3 (also see ). (G–I) Pan-FGFR inhibitor PD173074 (0.5 μM) elicits strong decreases in the thickness of the epibranchial placodes 1 and 2 as well as statistically significant decreases in neuroblast numbers in all three epibranchial placodes (also see ). (J–L) High doses of PD173074 (2.5 μM) result in the complete absence of high-grade thickened epibranchial placodes. Only single Ngn2 + neuroblasts can be detected, if at all (arrowhead in panel L ). (M–O) Anti-FGFR3 neutralizing antibodies (40 μg/ml) reduce Ngn2 + neuroblast numbers, but do not have negative effects on placode thickness. (P–R) Following pan-BMPR inhibition (10 μM LDN193189), epibranchial placode 1 reveals reduced numbers of Ngn2 + neuroblasts, but normal morphology (P) ; epibranchial placode 2 lacks any obvious impairment (Q) ; epibranchial placode 3 simultaneously presents slight reductions in Ngn2 + neuroblast numbers and placode thickness (R) . p1, p2, pharyngeal pouches 1 and 2. Scale bars: 20 μm.

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Inhibition, Cell Culture, Immunohistochemistry, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Impact of FGFR inhibition on the neurogenesis of epibranchial placodes (EP) in 9–14 somite mouse embryos, cultured for 24 h. Box plots indicate the numbers of Neurogenin (Ngn)2 + (A–C) or Ngn1 + (D–F) neuroblasts in EP1 (A,D) , EP2 (B,E) , or EP3 (C,F) following exposure to either dimethyl sulfoxide (DMSO) only (control; gray), or increasing doses of pan-FGFR inhibitor SU5402 (light blue), or 0.5 μM of pan-FGFR inhibitor PD173074 (dark blue), or increasing concentrations of anti-FGFR3 neutralizing antibodies (yellow). Significant differences between each treatment group and the controls are indicated by gray asterisks (* P < 0.05, ** P < 0.001; Mann–Whitney test). Additionally, PD173074-treated embryos were compared to groups exposed to either highest levels of SU5402 or anti-FGFR3 antibodies (significant differences: # P < 0.05, ## P < 0.001; or specific P values, respectively). Whiskers, lower and upper extremes; box limits, 25th and 75th percentiles; red center lines, medians; black dots, data points; open circles, outliers.

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Inhibition, Cell Culture, Control, MANN-WHITNEY

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Developmental profiles of the epibranchial placodes of C57BL/6N mice (embryonic days (E) 8.5 to E11.5) as reflected by earlier reconstructions of Ngn2 + intraplacodal neuroblasts recounted for present purposes ( n = 11 mouse embryos with 22 body sides). (A–C) Using CurveExpert Professional (Hyams Development, Chattanooga, TN, United States), regression curves were generated that represent the developmental profiles of the epibranchial placodes 1, 2, and 3 (EP1, red; EP2, blue; EP3, green). Confidence bands (medium red, blue, or green) most likely contain 95% of all values (black dots). Prediction bands (faint red, blue, or green) indicate the range in which 95% of all future values will fall. Here, embryos possessing 9–14 pairs of somites were included in our whole embryo cultures (WECs). This range is indicated by the left of the two gray columns and is correlated with the embryonic age plotted on the x -axis. The right gray column marks the period within which embryos were removed from the culture after 24 h. Asterisks indicate the respective medians of Ngn2 + neuroblasts for EP1, EP2, and EP3 of DMSO control embryos . (D–I) Box plots display the relative changes in the numbers of Ngn2 + (E–G,I) or Ngn1 + (D,H) neuroblasts following exposure to either the pan-FGFR inhibitor PD173074 (D,I) , to anti-FGFR3 neutralizing antibodies (F–H) , or to the pan-BMPR inhibitor LDN193189 (E) , respectively (see , also for n values). Significant differences between EP1 (red), EP2 (blue), and/or EP3 (green) are indicated by asterisks (* P < 0.05, ** P < 0.001; Mann–Whitney test). Whiskers, lower and upper extremes; box limits, 25th and 75th percentiles; red, blue, or green center lines, medians; open circles, outliers. (D,E) Plausibility checks demonstrate that EP2 and EP3, which almost synchronously produce neuroblasts, by no means always react in the same way to certain inhibitors. (D,F–I) Conversely, EP1, which slightly precedes EP2 and EP3 in terms of neuroblast production, does not always deviate in its behavior toward certain inhibitors from EP2 and/or EP3 (for details, see text).

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Generated, Control, MANN-WHITNEY

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Nine to fourteen somite mouse embryos, cultured for 24 h. (A) Control embryo, exposed to the solvent dimethyl sulfoxide (DMSO) only, reveals age-appropriate telencephalic hemispheres (green asterisk) and upper limb buds (green arrowheads); for comparison (green arrow), see corresponding hematoxylin stained histological sections (green colored areas). (B) Treatment with 2.5 μM of the pan-FGFR inhibitor PD173074 considerably impairs the development of telencephalic hemispheres (red asterisk) and upper limb buds (red arrowheads); for comparison (red arrow), see corresponding hematoxylin stained histological sections (red colored areas). (C) Application of 0.5 μM PD173074 did not cause any obvious abnormalities. (D) Embryos incubated with 40 μM of the pan-FGFR inhibitor SU5402 exhibit general growth disturbances (also see ). (E,F) Treatment with 40 μg/ml of anti-FGFR3 neutralizing antibodies or 10 μM of the pan-BMPR inhibitor LDN193189 were well tolerated. Scale bars: 500 μm for stereomicrographs (A–F) , 200 μm for light micrographs (A,B) .

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Cell Culture, Control, Solvent, Comparison, Staining, Incubation

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Impact of FGFR and BMPR inhibition on epibranchial placode morphogenesis and neurogenesis in 9–14 somite mouse embryos, cultured for 24 h. Anti-Neurogenin-2 (Ngn2) immunohistochemistry (brown precipitate); shown are maximum numbers of Ngn2 + neuroblasts found per section. (A–C) Control placodes reveal high-grade thickened pseudostratified epithelium (2–4 cell nuclei) with 12–16 Ngn2 + neuroblasts. (D–F) Pan-FGFR inhibitor SU5402 (40 μM) causes moderate reductions in placode thickness and statistically significant decreases in neuroblast numbers in the epibranchial placodes 1 and 2, but not 3 (also see ). (G–I) Pan-FGFR inhibitor PD173074 (0.5 μM) elicits strong decreases in the thickness of the epibranchial placodes 1 and 2 as well as statistically significant decreases in neuroblast numbers in all three epibranchial placodes (also see ). (J–L) High doses of PD173074 (2.5 μM) result in the complete absence of high-grade thickened epibranchial placodes. Only single Ngn2 + neuroblasts can be detected, if at all (arrowhead in panel L ). (M–O) Anti-FGFR3 neutralizing antibodies (40 μg/ml) reduce Ngn2 + neuroblast numbers, but do not have negative effects on placode thickness. (P–R) Following pan-BMPR inhibition (10 μM LDN193189), epibranchial placode 1 reveals reduced numbers of Ngn2 + neuroblasts, but normal morphology (P) ; epibranchial placode 2 lacks any obvious impairment (Q) ; epibranchial placode 3 simultaneously presents slight reductions in Ngn2 + neuroblast numbers and placode thickness (R) . p1, p2, pharyngeal pouches 1 and 2. Scale bars: 20 μm.

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Inhibition, Cell Culture, Immunohistochemistry, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Impact of FGFR inhibition on the neurogenesis of epibranchial placodes (EP) in 9–14 somite mouse embryos, cultured for 24 h. Box plots indicate the numbers of Neurogenin (Ngn)2 + (A–C) or Ngn1 + (D–F) neuroblasts in EP1 (A,D) , EP2 (B,E) , or EP3 (C,F) following exposure to either dimethyl sulfoxide (DMSO) only (control; gray), or increasing doses of pan-FGFR inhibitor SU5402 (light blue), or 0.5 μM of pan-FGFR inhibitor PD173074 (dark blue), or increasing concentrations of anti-FGFR3 neutralizing antibodies (yellow). Significant differences between each treatment group and the controls are indicated by gray asterisks (* P < 0.05, ** P < 0.001; Mann–Whitney test). Additionally, PD173074-treated embryos were compared to groups exposed to either highest levels of SU5402 or anti-FGFR3 antibodies (significant differences: # P < 0.05, ## P < 0.001; or specific P values, respectively). Whiskers, lower and upper extremes; box limits, 25th and 75th percentiles; red center lines, medians; black dots, data points; open circles, outliers.

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Inhibition, Cell Culture, Control, MANN-WHITNEY

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Developmental profiles of the epibranchial placodes of C57BL/6N mice (embryonic days (E) 8.5 to E11.5) as reflected by earlier reconstructions of Ngn2 + intraplacodal neuroblasts recounted for present purposes ( n = 11 mouse embryos with 22 body sides). (A–C) Using CurveExpert Professional (Hyams Development, Chattanooga, TN, United States), regression curves were generated that represent the developmental profiles of the epibranchial placodes 1, 2, and 3 (EP1, red; EP2, blue; EP3, green). Confidence bands (medium red, blue, or green) most likely contain 95% of all values (black dots). Prediction bands (faint red, blue, or green) indicate the range in which 95% of all future values will fall. Here, embryos possessing 9–14 pairs of somites were included in our whole embryo cultures (WECs). This range is indicated by the left of the two gray columns and is correlated with the embryonic age plotted on the x -axis. The right gray column marks the period within which embryos were removed from the culture after 24 h. Asterisks indicate the respective medians of Ngn2 + neuroblasts for EP1, EP2, and EP3 of DMSO control embryos . (D–I) Box plots display the relative changes in the numbers of Ngn2 + (E–G,I) or Ngn1 + (D,H) neuroblasts following exposure to either the pan-FGFR inhibitor PD173074 (D,I) , to anti-FGFR3 neutralizing antibodies (F–H) , or to the pan-BMPR inhibitor LDN193189 (E) , respectively (see , also for n values). Significant differences between EP1 (red), EP2 (blue), and/or EP3 (green) are indicated by asterisks (* P < 0.05, ** P < 0.001; Mann–Whitney test). Whiskers, lower and upper extremes; box limits, 25th and 75th percentiles; red, blue, or green center lines, medians; open circles, outliers. (D,E) Plausibility checks demonstrate that EP2 and EP3, which almost synchronously produce neuroblasts, by no means always react in the same way to certain inhibitors. (D,F–I) Conversely, EP1, which slightly precedes EP2 and EP3 in terms of neuroblast production, does not always deviate in its behavior toward certain inhibitors from EP2 and/or EP3 (for details, see text).

Article Snippet: For selective inhibition of the FGFR3 pathway, rat anti-FGFR3 neutralizing antibodies were deployed (MAB710, R&D Systems, Minneapolis, MN, United States, lot FTD0216021, RRID: AB_2103386 ).

Techniques: Generated, Control, MANN-WHITNEY

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Nine to fourteen somite mouse embryos, cultured for 24 h. (A) Control embryo, exposed to the solvent dimethyl sulfoxide (DMSO) only, reveals age-appropriate telencephalic hemispheres (green asterisk) and upper limb buds (green arrowheads); for comparison (green arrow), see corresponding hematoxylin stained histological sections (green colored areas). (B) Treatment with 2.5 μM of the pan-FGFR inhibitor PD173074 considerably impairs the development of telencephalic hemispheres (red asterisk) and upper limb buds (red arrowheads); for comparison (red arrow), see corresponding hematoxylin stained histological sections (red colored areas). (C) Application of 0.5 μM PD173074 did not cause any obvious abnormalities. (D) Embryos incubated with 40 μM of the pan-FGFR inhibitor SU5402 exhibit general growth disturbances (also see ). (E,F) Treatment with 40 μg/ml of anti-FGFR3 neutralizing antibodies or 10 μM of the pan-BMPR inhibitor LDN193189 were well tolerated. Scale bars: 500 μm for stereomicrographs (A–F) , 200 μm for light micrographs (A,B) .

Article Snippet: This is exactly the requirement that anti-FGFR3 neutralizing antibodies obtained from R&D Systems fulfill (MAB710).

Techniques: Cell Culture, Control, Solvent, Comparison, Staining, Incubation

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Impact of FGFR and BMPR inhibition on epibranchial placode morphogenesis and neurogenesis in 9–14 somite mouse embryos, cultured for 24 h. Anti-Neurogenin-2 (Ngn2) immunohistochemistry (brown precipitate); shown are maximum numbers of Ngn2 + neuroblasts found per section. (A–C) Control placodes reveal high-grade thickened pseudostratified epithelium (2–4 cell nuclei) with 12–16 Ngn2 + neuroblasts. (D–F) Pan-FGFR inhibitor SU5402 (40 μM) causes moderate reductions in placode thickness and statistically significant decreases in neuroblast numbers in the epibranchial placodes 1 and 2, but not 3 (also see ). (G–I) Pan-FGFR inhibitor PD173074 (0.5 μM) elicits strong decreases in the thickness of the epibranchial placodes 1 and 2 as well as statistically significant decreases in neuroblast numbers in all three epibranchial placodes (also see ). (J–L) High doses of PD173074 (2.5 μM) result in the complete absence of high-grade thickened epibranchial placodes. Only single Ngn2 + neuroblasts can be detected, if at all (arrowhead in panel L ). (M–O) Anti-FGFR3 neutralizing antibodies (40 μg/ml) reduce Ngn2 + neuroblast numbers, but do not have negative effects on placode thickness. (P–R) Following pan-BMPR inhibition (10 μM LDN193189), epibranchial placode 1 reveals reduced numbers of Ngn2 + neuroblasts, but normal morphology (P) ; epibranchial placode 2 lacks any obvious impairment (Q) ; epibranchial placode 3 simultaneously presents slight reductions in Ngn2 + neuroblast numbers and placode thickness (R) . p1, p2, pharyngeal pouches 1 and 2. Scale bars: 20 μm.

Article Snippet: This is exactly the requirement that anti-FGFR3 neutralizing antibodies obtained from R&D Systems fulfill (MAB710).

Techniques: Inhibition, Cell Culture, Immunohistochemistry, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Impact of FGFR inhibition on the neurogenesis of epibranchial placodes (EP) in 9–14 somite mouse embryos, cultured for 24 h. Box plots indicate the numbers of Neurogenin (Ngn)2 + (A–C) or Ngn1 + (D–F) neuroblasts in EP1 (A,D) , EP2 (B,E) , or EP3 (C,F) following exposure to either dimethyl sulfoxide (DMSO) only (control; gray), or increasing doses of pan-FGFR inhibitor SU5402 (light blue), or 0.5 μM of pan-FGFR inhibitor PD173074 (dark blue), or increasing concentrations of anti-FGFR3 neutralizing antibodies (yellow). Significant differences between each treatment group and the controls are indicated by gray asterisks (* P < 0.05, ** P < 0.001; Mann–Whitney test). Additionally, PD173074-treated embryos were compared to groups exposed to either highest levels of SU5402 or anti-FGFR3 antibodies (significant differences: # P < 0.05, ## P < 0.001; or specific P values, respectively). Whiskers, lower and upper extremes; box limits, 25th and 75th percentiles; red center lines, medians; black dots, data points; open circles, outliers.

Article Snippet: This is exactly the requirement that anti-FGFR3 neutralizing antibodies obtained from R&D Systems fulfill (MAB710).

Techniques: Inhibition, Cell Culture, Control, MANN-WHITNEY

Journal: Frontiers in Cell and Developmental Biology

Article Title: Responses of Epibranchial Placodes to Disruptions of the FGF and BMP Signaling Pathways in Embryonic Mice

doi: 10.3389/fcell.2021.712522

Figure Lengend Snippet: Developmental profiles of the epibranchial placodes of C57BL/6N mice (embryonic days (E) 8.5 to E11.5) as reflected by earlier reconstructions of Ngn2 + intraplacodal neuroblasts recounted for present purposes ( n = 11 mouse embryos with 22 body sides). (A–C) Using CurveExpert Professional (Hyams Development, Chattanooga, TN, United States), regression curves were generated that represent the developmental profiles of the epibranchial placodes 1, 2, and 3 (EP1, red; EP2, blue; EP3, green). Confidence bands (medium red, blue, or green) most likely contain 95% of all values (black dots). Prediction bands (faint red, blue, or green) indicate the range in which 95% of all future values will fall. Here, embryos possessing 9–14 pairs of somites were included in our whole embryo cultures (WECs). This range is indicated by the left of the two gray columns and is correlated with the embryonic age plotted on the x -axis. The right gray column marks the period within which embryos were removed from the culture after 24 h. Asterisks indicate the respective medians of Ngn2 + neuroblasts for EP1, EP2, and EP3 of DMSO control embryos . (D–I) Box plots display the relative changes in the numbers of Ngn2 + (E–G,I) or Ngn1 + (D,H) neuroblasts following exposure to either the pan-FGFR inhibitor PD173074 (D,I) , to anti-FGFR3 neutralizing antibodies (F–H) , or to the pan-BMPR inhibitor LDN193189 (E) , respectively (see , also for n values). Significant differences between EP1 (red), EP2 (blue), and/or EP3 (green) are indicated by asterisks (* P < 0.05, ** P < 0.001; Mann–Whitney test). Whiskers, lower and upper extremes; box limits, 25th and 75th percentiles; red, blue, or green center lines, medians; open circles, outliers. (D,E) Plausibility checks demonstrate that EP2 and EP3, which almost synchronously produce neuroblasts, by no means always react in the same way to certain inhibitors. (D,F–I) Conversely, EP1, which slightly precedes EP2 and EP3 in terms of neuroblast production, does not always deviate in its behavior toward certain inhibitors from EP2 and/or EP3 (for details, see text).

Article Snippet: This is exactly the requirement that anti-FGFR3 neutralizing antibodies obtained from R&D Systems fulfill (MAB710).

Techniques: Generated, Control, MANN-WHITNEY