Figure 2.
Blockade of dopamine synthesis impairs HSC maintenance. Representative confocal images showing expression pattern of endogenous Th protein in combination with Tuj1 (A) and TrkA (B) in WT BM. (C) Immunostaining showing distribution of dopamine and its colocalization with Th in BM. (D) Tile-scan image showing Th-Cre–driven GFP expression in Th-mTmG recipient BM. Colocalization of Th-Cre–driven GFP with neuronal presynaptic marker synaptophysin (E) and Th protein (F) in Th-mTmG recipients. (G) Diagram depicting genetic strategy for Th+ cell ablation after DTX administration. (H) Quantification of the percentages and numbers of LSK cells and HSCs in BM of DTRiΔTh (Cre+/T flox+/f; n = 6) and control (Cre+/+ flox+/f; n = 8) mice. The P value was calculated using the Student t test. (I) Quantification of donor-derived (CD45.2) B lymphocytes, T lymphocytes, and myeloid cells in competitive repopulating assay. A total of 5 × 105 CD45.2 (Ctrl or DTRiΔTh) BM cells mixed with 5 × 105 CD45.1 BM cells were transplanted into lethally irradiated CD45.1 recipients, which were analyzed 16 weeks later. Ctrl = 7; DTRiΔTh = 7. The P value was calculated using the Student t test. (J) Schematic diagram depicting WT BM transplantation into DTRiΔTh and control mice. DTX injection and HSC analysis were done 2 months after transplantation. Th-mTmG reporter mice were analyzed without toxin injection. (K) Representative FACS plots of LSK cells and HSCs in control and DTRiΔTh(SC) recipients after DTX administration. Numbers represent the percentages of the indicated cell populations in BM. (L) Quantification of the percentages (left panels) and numbers (right panels) of LSK cells and HSCs in control (n = 11) and DTRiΔTh(SC) (n = 8) recipient BM. The P value was calculated using the Student t test. (M) Quantification of the numbers and percentages of LSK cells and HSCs in BM from vehicle-injected (n = 6) or carbidopa-injected (n = 6) WT mice. The P value was calculated using the Student t test. APC, allophycocyanin; Ctrl, control; FITC, fluorescein isothiocyanate; PE, phycoerythrin.

Blockade of dopamine synthesis impairs HSC maintenance. Representative confocal images showing expression pattern of endogenous Th protein in combination with Tuj1 (A) and TrkA (B) in WT BM. (C) Immunostaining showing distribution of dopamine and its colocalization with Th in BM. (D) Tile-scan image showing Th-Cre–driven GFP expression in Th-mTmG recipient BM. Colocalization of Th-Cre–driven GFP with neuronal presynaptic marker synaptophysin (E) and Th protein (F) in Th-mTmG recipients. (G) Diagram depicting genetic strategy for Th+ cell ablation after DTX administration. (H) Quantification of the percentages and numbers of LSK cells and HSCs in BM of DTRiΔTh (Cre+/T flox+/f; n = 6) and control (Cre+/+ flox+/f; n = 8) mice. The P value was calculated using the Student t test. (I) Quantification of donor-derived (CD45.2) B lymphocytes, T lymphocytes, and myeloid cells in competitive repopulating assay. A total of 5 × 105 CD45.2 (Ctrl or DTRiΔTh) BM cells mixed with 5 × 105 CD45.1 BM cells were transplanted into lethally irradiated CD45.1 recipients, which were analyzed 16 weeks later. Ctrl = 7; DTRiΔTh = 7. The P value was calculated using the Student t test. (J) Schematic diagram depicting WT BM transplantation into DTRiΔTh and control mice. DTX injection and HSC analysis were done 2 months after transplantation. Th-mTmG reporter mice were analyzed without toxin injection. (K) Representative FACS plots of LSK cells and HSCs in control and DTRiΔTh(SC) recipients after DTX administration. Numbers represent the percentages of the indicated cell populations in BM. (L) Quantification of the percentages (left panels) and numbers (right panels) of LSK cells and HSCs in control (n = 11) and DTRiΔTh(SC) (n = 8) recipient BM. The P value was calculated using the Student t test. (M) Quantification of the numbers and percentages of LSK cells and HSCs in BM from vehicle-injected (n = 6) or carbidopa-injected (n = 6) WT mice. The P value was calculated using the Student t test. APC, allophycocyanin; Ctrl, control; FITC, fluorescein isothiocyanate; PE, phycoerythrin.

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