aav2 dj dio ef1α mkate2 bipac  (Addgene inc)

Journal: Journal of cell science

Article Title: Planar polarized organization of mouse hair cells is established and maintained by STK32A, GPR156 and EMX2

doi: 10.1242/jcs.264340

Figure Lengend Snippet: ( A ) qRT-PCR measures of Stk32a transcript relative to the housekeeping gene GAPDH and the IHC-specific marker gene Fgf8 in organ of Corti tissue dissected from Emx2 KO and control cochlea at E18.5. Relative levels of Fgf8 mRNA remain comparable across genotypes consistent with the preservation of IHCs in Emx2 KOs. Error bars are standard deviation and significance was determined using Student’s t-test (* p<0.05, ** p<0.001). ( B ) Fluorescent ISH using the Hybridization Chain Reaction technique shows the distribution of Stk32a mRNA in Emx2 KO and extensive overlap with IHCs in the E18.5 cochlea. Brackets indicate the location of the single row of IHCs in controls and expanded rows of IHCs in the Emx2 KO. OHCs are not labeled in controls and are absent from the Emx2 KO. Scale bars are 10μm.

Article Snippet: HCR probes used in this study were: Fgf8 -B3 (Molecular Instruments PRC597), Gfi1 -B2 (Molecular Instruments PRG075), and Stk32a -B1 (Molecular Instruments PRF996).

Techniques: Quantitative RT-PCR, Marker, Control, Preserving, Standard Deviation, Hybridization, Labeling

Journal: The FASEB Journal

Article Title: Loss of the RNA Binding Protein HuR in Early Murine Limb Mesenchyme Does Not Affect Development but Leads to Impaired Bone Homeostasis in Adulthood

doi: 10.1096/fj.202500780RR

Figure Lengend Snippet: Effect of germline Cre recombination in HuR fl/fl mice (Elavl1 KO) through maternal inheritance of Prx1‐Cre. (A) Whole‐mount embryos at E13.5 stained with alcian blue. (B) Toluidine blue‐stained sections of E13.5 embryo forelimbs and hindlimbs. (C) Immunofluorescence detection of HuR in limb sections from E13.5 embryos. (D) Brightfield micrographs demonstrating morphology changes in Control and Elavl1 KO embryos isolated from E9.5 to E11.5. Scale bar = 500 μm. (E) Whole‐mount in situ hybridization chain reaction detection of limb bud markers mRNA in E10.5 and E11.5 embryos. Asterisk: Limb bud, Triangle: Apical ectodermal ridge, Arrowhead: The zone of polarizing activity. Scale bar = 150 μm. (F) Whole‐mount immunofluorescence detection of Fgf8 protein in E10.5 embryos. Asterisk: Limb bud, Triangle: Apical ectodermal ridge. Scale bar = 150 μm.

Article Snippet: Embryos were fixed in 4% formaldehyde for 1 h at 4°C, washed, and stored in 0.0025% Triton X‐100 in PBS (PBS‐Tr) at 4° C. Prior to immunofluorescence staining, embryos were permeabilized with 0.25% Triton X‐100 in PBS for 30 min and then blocked with 2% bovine serum albumin in PBS solution for 1 h before incubation with 1:100 Fgf8 antibody (MAB323, R&D Systems, UK) overnight at 4°C.

Techniques: Staining, Immunofluorescence, Control, Isolation, In Situ Hybridization, Activity Assay

Journal: Neuroscience Bulletin

Article Title: Fibroblast Growth Factor 8 Suppresses Neurotoxic Astrocytes and Alleviates Neuropathic Pain via Spinal FGFR3 Signaling

doi: 10.1007/s12264-025-01533-x

Figure Lengend Snippet: FGF8 alleviates SNI-induced mechanical allodynia through FGFR3. A , B Paw withdrawal threshold ( A ) and von Frey–evoked response frequency ( B ) in SNI mice following a single intrathecal injection of FGF8 (1 μg in 5 μL) on day 14 post-SNI. C Time course of mechanical allodynia response over five consecutive days following FGF8 administration on day 14 post-SNI. D , E qRT-PCR analysis showed changes of Fgfr2 ( D ) and Fgfr3 ( E ) in spinal cord tissue on day 14 post-SNI. F , G Representative images showing co-localization of FGFR3 with GFAP in the SDH of sham and SNI mice ( F ) and quantification ( G ) on day 14 post-SNI. H , I Representative images and quantification of FGFR3 knockdown efficiency in spinal astrocytes using FGFR3-targeting shRNA. Arrowheads indicate the typical cell. J , K Selective knockdown of FGFR3 attenuated the analgesic effects of intrathecal FGF8 injection on day 14 post-SNI. n = 9 mice ( A , C) ; n = 4–6 mice ( B–E , J , K ); n = 10-12 slices ( G , I ) from 3 mice. Scale bar: 25 μm ( F , H ). Statistics by two-way ANOVA followed by post hoc Holm-Sidak test ( A–C , J , K ) and two-tailed Student’s t -test ( D–I ). * P < 0.05, ** P < 0.01. Data are presented as mean ± SEM. I.t, intrathecal injection.

Article Snippet: Recombinant FGF8 protein (0.5 μg or 1 μg in 5 μL PBS; R&D Systems, 423-F8-025/CF) was administered via intrathecal (i.t.) lumbar puncture between L5–L6 vertebrae under isoflurane anesthesia using a 31G needle-equipped Hamilton syringe (25 μL volume).

Techniques: Injection, Quantitative RT-PCR, Knockdown, shRNA, Two Tailed Test

Journal: Neuroscience Bulletin

Article Title: Fibroblast Growth Factor 8 Suppresses Neurotoxic Astrocytes and Alleviates Neuropathic Pain via Spinal FGFR3 Signaling

doi: 10.1007/s12264-025-01533-x

Figure Lengend Snippet: FGF8 alleviates inflammatory induction of neurotoxic astrocyte states in vitro. A Relative mRNA expression of C3 in isolated astrocytes treated with normal culture medium, LPS, or MCM, as measured by qRT-PCR. B – I Expression of reactive astrocyte markers in astrocytes treated with IL-1α, TNF-α, and C1q, with or without FGF8 co-treatment. A1-specific transcripts: C3 ( B ), H2-T23 ( D ), and Serping1 ( E ); A2-specific transcripts: S100a10 ( C ), Ptx3 ( F ), and Ptgs ( G ); pan-reactive transcripts: Lcn2 ( H ) and Cxcl10 ( I ). J–L Relative mRNA expression of synaptogenic genes Gpc4 ( J ), Gpc6 ( K ), Tsp1, and Tsp2 ( L ) in astrocytes under the same treatment conditions. M , N Relative mRNA expression of Fgfr2 ( M ) and Fgfr3 ( N ) in isolated astrocytes treated with normal culture medium, LPS, or MCM, as measured by qRT-PCR. n = 6–12 petri dishes ( A–C ) and n = 3–7 petri dishes ( M , N ) derived from pooled spinal cords of 6 mice per preparation; n = 3–6 petri dishes ( D–I ) and n = 4–6 petri dishes ( J–L ) from pooled spinal cords of 12 mice per preparation. Statistics by one-way ANOVA by post hoc Holm-Sidak test ( A–N ). * P < 0.05, ** P < 0.01. # P < 0.05, ## P < 0.01. Data are presented as mean ± SEM. LPS, lipopolysaccharide. MCM, microglia-conditioned media.

Article Snippet: Recombinant FGF8 protein (0.5 μg or 1 μg in 5 μL PBS; R&D Systems, 423-F8-025/CF) was administered via intrathecal (i.t.) lumbar puncture between L5–L6 vertebrae under isoflurane anesthesia using a 31G needle-equipped Hamilton syringe (25 μL volume).

Techniques: In Vitro, Expressing, Isolation, Quantitative RT-PCR, Derivative Assay

Journal: Neuroscience Bulletin

Article Title: Fibroblast Growth Factor 8 Suppresses Neurotoxic Astrocytes and Alleviates Neuropathic Pain via Spinal FGFR3 Signaling

doi: 10.1007/s12264-025-01533-x

Figure Lengend Snippet: Central action of FGF8 enhances inhibitory synaptic transmission in the spinal cord. A – D Representative firing traces ( A ) and quantitative statistics showing no significant changes in firing frequencies ( B ), RMP ( C ), or rheobase current ( D ) in lamina II neurons following perfusion with FGF8 (500 ng/mL). E – G Representative traces of sEPSCs of lamina II neurons ( E ) and quantitative statistics of sEPSC frequency ( F ) and amplitude ( G ). H – J Representative traces of sIPSCs of lamina II neurons ( H ) and quantitative statistics of sIPSC frequency ( I ) and amplitude ( J ). n = 18–19 neurons ( B – D ) and n = 15–19 neurons ( F , G , I , J ) from 3 mice. Statistics by two-way ANOVA followed by post hoc Holm-Sidak test ( B ), two-tailed Student’s t -test ( C , D ), and Mann-Whitney U test ( F , G , I , J ).** P < 0.01. Data are presented as mean ± SEM. AP, action potential. RMP, resting membrane potential. sEPSC, spontaneous excitatory postsynaptic current. sIPSC, spontaneous inhibitory postsynaptic current.

Article Snippet: Recombinant FGF8 protein (0.5 μg or 1 μg in 5 μL PBS; R&D Systems, 423-F8-025/CF) was administered via intrathecal (i.t.) lumbar puncture between L5–L6 vertebrae under isoflurane anesthesia using a 31G needle-equipped Hamilton syringe (25 μL volume).

Techniques: Transmission Assay, Two Tailed Test, MANN-WHITNEY, Membrane

Journal: Neuroscience Bulletin

Article Title: Fibroblast Growth Factor 8 Suppresses Neurotoxic Astrocytes and Alleviates Neuropathic Pain via Spinal FGFR3 Signaling

doi: 10.1007/s12264-025-01533-x

Figure Lengend Snippet: SNI induces persistent microglial activation in the SDH. A , B Representative immunofluorescence images ( A ) and quantification ( B ) of IBA-1 staining in the ipsilateral SDH from day 1 to day 14 post-SNI. C , D Representative Western blot images ( C ) and quantification ( D ) of IBA-1 protein levels in the L2–L5 spinal cord segments from day 4 to day 14 post-SNI. E Representative skeletonized images of individual IBA-1⁺ microglia from sham and SNI mice. F , G Quantification of microglial morphology on days 4, 7, and 14 post-SNI, showing a significant increase in soma area ( F ) and a decrease in process length ( G ) compared to sham controls. H – J Sholl analysis demonstrates reduced microglial process complexity in SNI mice compared to sham controls. n = 9 slices. K qRT-PCR analysis of Tnf-α , Il-1α , and C1q in the ipsilateral SDH from sham and SNI mice on day 14 ( n = 5–6 mice). L qRT-PCR analysis of Tnf-α , Il-1α , and C1q in spinal cord tissue from SNI mice treated with FGF8 or vehicle on day 14 ( n = 6 per group). M Representative images of C3 and GFAP co-localization and quantification of C3 and GFAP in the SDH of control and FGFR3 shRNA mice following i.t. LPS administration. n = 9 slices ( B ), n = 8–11 cells ( F – J ) from 3 mice; n = 4–6 mice ( D , K , L ); n = 12-13 slices ( M ) from 3 mice. Statistics by two-way ANOVA followed by post hoc Holm-Sidak test ( B , D , F , G ). Scale bar: 25 μm ( A , E , M ). Statistics by two-way ANOVA ( H-I ), two-tailed Student’s t -test ( K , L ), and Mann-Whitney U test ( M ). * P < 0.05, ** P < 0.01. Data are presented as mean ± SEM.

Article Snippet: Recombinant FGF8 protein (0.5 μg or 1 μg in 5 μL PBS; R&D Systems, 423-F8-025/CF) was administered via intrathecal (i.t.) lumbar puncture between L5–L6 vertebrae under isoflurane anesthesia using a 31G needle-equipped Hamilton syringe (25 μL volume).

Techniques: Activation Assay, Immunofluorescence, Staining, Western Blot, Quantitative RT-PCR, Control, shRNA, Two Tailed Test, MANN-WHITNEY