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. Author manuscript; available in PMC: 2022 Sep 1.
Published in final edited form as: Dev Dyn. 2020 Oct 26;250(9):1330–1339. doi: 10.1002/dvdy.261

Figure 3. Systemic and targeted elevation of vitamin D signaling promotes fin regeneration.

Figure 3.

(A) Three daily injections of 0.8 μg alfacalcidol increases cyp24a1-directed GFP fluorescence in uninjured caudal fins. (B) The lengths of 4 dpa regenerates increased by 5% with daily injections of 0.8 μg alfacalcidol injection, compared to vehicle (10% EtOH) injection. Values were normalized to the 4 dpa average lengths of vehicle-injected fish to correct for batch differences and plotted as relative lengths, as pooled data from three experimental groups were used. n = 41 for each group. (C) Daily injections of 0.8 μg alfacalcidol to osx:H2A-mCherry; osx:Venus-hGeminin transgenic fish following fin amputation increased osteoblast proliferation compared to 10% EtOH treated ones in regenerating fins at 4 dpa. Red, osteoblast nuclei. Green, osteoblast nuclei in S/G2/M. n = 9 for each group. (D) Microinjection of 25 ng/μL vdra RNA into one-cell stage embryos sensitized the response to 24 hours treatment with 0.1 μM alfacalcidol. Green represents cyp24a1-directed fluorescence, a readout for VDR activity, which is higher in the RNA-injected group. Vehicle for RNA injection is water, and vehicle for alfacalcidol treatment is 10% DMSO. Representative embryos shown, n = 20 for each group. (E, F) A low dose (0.4 μg) daily injection of alfacalcidol is sufficient to improve fin regeneration in lepb:vdra and lepb:vdrb transgenic lines, which induce wild-type VDR expression in regenerating fins. Wild-type animals show no difference between vehicle- (10% EtOH) and alfacalcidol-injected groups. n = 10 for all groups. Dashed lines indicate approximate amputation planes.