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B cells are a source of GABA in the bone marrow (A) Representative flow plots showing emergence of B lineage cells in OP9 co-culture. (B) Schematic of WT or Cd19-Cre CKO HSPC OP9 B cell differentiation co-culture. (C) Left: quantification of Cd19 + B cells percentage at multiple time points. Right: mass spectrometric quantification of GABA level in the media throughout B cell differentiation. One-way ANOVA with Tukey’s test was performed. (D) Breeding scheme to generate Cd19-Cre CKO and control mice. (E) Left: quantification of B cells produced during OP9 co-culture differentiation of HSPCs isolated from Cd19-Cre CKO . Welch’s test was performed. Right: GABA level in media from Cd19-Cre mice and WT mice at day 12 of OP9 co-culture. OP9 stromal cells alone do not produce GABA. One-way ANOVA with Tukey’s test was performed. (F) Mass spectrometric quantification of GABA in Cd19-Cre and control mouse femurs ( n = 5 mutants vs. n = 5 controls). Paired t test was performed. (G) Left: representative flow plots showing HSPC gates (LSK: Lin - /Sca-1 + /c-Kit + ). Right: quantification of HSPC abundance in Cd19-Cre CKO and control mice ( n = 9 mutants vs. n = 9 controls). Data represented as mean ± standard deviation (SD) ( ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001), Welch’s test was performed. Graphics created with BioRender.

Journal: Stem Cell Reports

Article Title: GABA produced by multiple bone marrow cell types regulates hematopoietic stem and progenitor cells

doi: 10.1016/j.stemcr.2025.102677

Figure Lengend Snippet: B cells are a source of GABA in the bone marrow (A) Representative flow plots showing emergence of B lineage cells in OP9 co-culture. (B) Schematic of WT or Cd19-Cre CKO HSPC OP9 B cell differentiation co-culture. (C) Left: quantification of Cd19 + B cells percentage at multiple time points. Right: mass spectrometric quantification of GABA level in the media throughout B cell differentiation. One-way ANOVA with Tukey’s test was performed. (D) Breeding scheme to generate Cd19-Cre CKO and control mice. (E) Left: quantification of B cells produced during OP9 co-culture differentiation of HSPCs isolated from Cd19-Cre CKO . Welch’s test was performed. Right: GABA level in media from Cd19-Cre mice and WT mice at day 12 of OP9 co-culture. OP9 stromal cells alone do not produce GABA. One-way ANOVA with Tukey’s test was performed. (F) Mass spectrometric quantification of GABA in Cd19-Cre and control mouse femurs ( n = 5 mutants vs. n = 5 controls). Paired t test was performed. (G) Left: representative flow plots showing HSPC gates (LSK: Lin - /Sca-1 + /c-Kit + ). Right: quantification of HSPC abundance in Cd19-Cre CKO and control mice ( n = 9 mutants vs. n = 9 controls). Data represented as mean ± standard deviation (SD) ( ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001), Welch’s test was performed. Graphics created with BioRender.

Article Snippet: For HSPC isolation, BM cells were pre-enriched for c-Kit + cells using c-Kit microBeads (Miltenyi Biotec, 130-091-224) and a Midi MACS separator (Miltenyi Biotec, 130-042-301).

Techniques: Co-Culture Assay, Cell Differentiation, Control, Produced, Isolation, Standard Deviation

BMECs are a source of GABA in the bone marrow (A) Breeding scheme to generate Cdh5-CreERT2 CKO and control mice. Graphics created with BioRender. (B) Mass spectrometric quantification of GABA in Cdh5-CreERT2 CKO and control mouse femurs ( n = 3 mutants vs. n = 3 controls). Paired t test was performed. (C) Left: representative flow plots showing the HSPC gates (LSK: Lin - /Sca-1 + /c-Kit + ). Right: quantification of LSK cell abundance in Cdh5-CreERT2 CKO and control mice ( n = 11 mutants vs. n = 12 control). Data represented as mean ± standard deviation (SD) ( ∗ p < 0.05 and ∗∗ p < 0.01), Welch’s test was performed.

Journal: Stem Cell Reports

Article Title: GABA produced by multiple bone marrow cell types regulates hematopoietic stem and progenitor cells

doi: 10.1016/j.stemcr.2025.102677

Figure Lengend Snippet: BMECs are a source of GABA in the bone marrow (A) Breeding scheme to generate Cdh5-CreERT2 CKO and control mice. Graphics created with BioRender. (B) Mass spectrometric quantification of GABA in Cdh5-CreERT2 CKO and control mouse femurs ( n = 3 mutants vs. n = 3 controls). Paired t test was performed. (C) Left: representative flow plots showing the HSPC gates (LSK: Lin - /Sca-1 + /c-Kit + ). Right: quantification of LSK cell abundance in Cdh5-CreERT2 CKO and control mice ( n = 11 mutants vs. n = 12 control). Data represented as mean ± standard deviation (SD) ( ∗ p < 0.05 and ∗∗ p < 0.01), Welch’s test was performed.

Article Snippet: For HSPC isolation, BM cells were pre-enriched for c-Kit + cells using c-Kit microBeads (Miltenyi Biotec, 130-091-224) and a Midi MACS separator (Miltenyi Biotec, 130-042-301).

Techniques: Control, Standard Deviation

BMEC and hematopoietic cell-derived GABA synergizes to regulate HSPC activity (A) Breeding scheme to generate Tie2-Cre CKO and control mice. Graphics created with BioRender. (B) Mass spectrometric quantification of GABA in Tie2-Cre CKO and control mouse femur ( n = 3 mutants vs. n = 3 controls). Paired t test was performed. (C) Left: representative flow plots showing the HSPC gates (LSK: Lin - /Sca-1 + /c-Kit + ). Right: quantification of LSK cell abundance in Tie2-Cre CKO mice and control mice ( n = 7 mutants vs. n = 7 controls). Welch’s test was performed. (D) Percentage of pre-B cells in Tie2-Cre CKO and control mice ( n = 8 mutants vs. n = 8 controls). Welch’s test was performed. (E) Percentage of B220 + cells in PB of Tie2-Cre CKO and control mice ( n = 9 mutants vs. n = 9 controls). Welch’s test was performed. Data are represented as mean ± standard deviation (SD) ( ∗ p < 0.05). (F) Summary of GABA levels and HSPC reduction in all CKO models.

Journal: Stem Cell Reports

Article Title: GABA produced by multiple bone marrow cell types regulates hematopoietic stem and progenitor cells

doi: 10.1016/j.stemcr.2025.102677

Figure Lengend Snippet: BMEC and hematopoietic cell-derived GABA synergizes to regulate HSPC activity (A) Breeding scheme to generate Tie2-Cre CKO and control mice. Graphics created with BioRender. (B) Mass spectrometric quantification of GABA in Tie2-Cre CKO and control mouse femur ( n = 3 mutants vs. n = 3 controls). Paired t test was performed. (C) Left: representative flow plots showing the HSPC gates (LSK: Lin - /Sca-1 + /c-Kit + ). Right: quantification of LSK cell abundance in Tie2-Cre CKO mice and control mice ( n = 7 mutants vs. n = 7 controls). Welch’s test was performed. (D) Percentage of pre-B cells in Tie2-Cre CKO and control mice ( n = 8 mutants vs. n = 8 controls). Welch’s test was performed. (E) Percentage of B220 + cells in PB of Tie2-Cre CKO and control mice ( n = 9 mutants vs. n = 9 controls). Welch’s test was performed. Data are represented as mean ± standard deviation (SD) ( ∗ p < 0.05). (F) Summary of GABA levels and HSPC reduction in all CKO models.

Article Snippet: For HSPC isolation, BM cells were pre-enriched for c-Kit + cells using c-Kit microBeads (Miltenyi Biotec, 130-091-224) and a Midi MACS separator (Miltenyi Biotec, 130-042-301).

Techniques: Derivative Assay, Activity Assay, Control, Standard Deviation