Journal: The FASEB Journal

Article Title: Human Dorsal Root Ganglia Neuronal Cell Line to Study Nociceptive Signaling: A New Pipeline for Pain Therapy

doi: 10.1096/fj.202503698R

Figure Lengend Snippet: Key nociceptive ion channel abundance, localization, and activity in HD10.6 cells. (A, B) D0 and D10 HD10.6 cells were fixed and stained for DAPI (blue), NF200 (green), and nociceptive ion channel TRPV1 (A) or NaV1.7 (B) in red. Representative images shown from three independent experiments. Scale bar 50 μm; filled arrows indicate examples of signal at soma whereas empty arrows indicate signal within axons. (C–F) Calcium influx in response to buffer only (BUF., blue) as well as during and following 10 μM TRPV1 agonist capsaicin (CAP., C, D, orange) or 1 μM NaV1.7 agonist OD1 (E, F, orange) was recorded from differentiated HD10.6 cells with Ca 2+ indicator Fluo‐4 AM (5 μM). Buffer was applied for 2 min and followed by the application of capsaicin or OD1 for 30 s. After up to 4 additional minutes of buffer, 50 mM KCl was applied for 5 s at the termination of the experiment to verify cell viability. Representative traces in response to respective stimuli shown (C,E), as well as the maximal normalized intensity (D,F). Replicates from three independent experiments. Statistical comparisons were made using Wilcoxon signed‐rank test due to non‐normality. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: NaV1.7 , Alomone Labs ASC‐008 , 1:100.

Techniques: Activity Assay, Staining

Journal: The FASEB Journal

Article Title: Human Dorsal Root Ganglia Neuronal Cell Line to Study Nociceptive Signaling: A New Pipeline for Pain Therapy

doi: 10.1096/fj.202503698R

Figure Lengend Snippet: Culture of HD10.6 cells into microfluidic devices to study peripheral administration. (A, B) D0 HD10.6 cells were seeded into the soma compartment of a microfluidic chamber, and media was changed to differentiation media 24 h post‐seeding. A 2X nerve growth factor (NGF) gradient was used to guide axons towards the axon‐only compartment. At D14, the cultures within the chamber were fixed and stained for neuronal marker β3 tubulin (magenta), marker for NaV1.7 PTX‐488 (green), and DAPI (blue). Empty arrows point to axons traveling through microgrooves. (A) captures a representative image of the soma compartment and axons within the proximal end of the microgrooves, whereas (B) shows the axon‐only compartment and distal microgrooves. Scale bar 50 μm. Respective compartments are highlighted in green within the right‐hand animated microfluidic chambers; images of microfluidic chambers created with BioRender.com . (C, D) D10 HD10.6 cells in 24‐well cell culture wells were infected with AAV9‐mCherry. 3 days following infection, cells were fixed and stained for β3 tubulin (green), mCherry (red), and DAPI (blue) The percentage of mCherry+ HD10.6 cells based on the total number of cells identified by DAPI and β3 tubulin‐positive staining is demonstrated in (C). A representative image of AAV9‐mCherry‐treated (1E9, top) and untreated (bottom) HD10.6 cells at this time‐point is shown in (D). (E) 3 days following AAV9‐mCherry infection in the peripheral compartment of HD10.6 cells cultured in microfluidic chambers, the cultures were fixed and stained for β3 tubulin (green), mCherry (red), and DAPI (blue). Representative image of soma (top) and peripheral terminal compartments (bottom) demonstrate mCherry signal within the soma and axons of HD10.6 cells. Replicates from three independent experiments; statistical comparisons were made using an unpaired non‐normal t ‐test. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: NaV1.7 , Alomone Labs ASC‐008 , 1:100.

Techniques: Staining, Marker, Cell Culture, Infection