Aldh1a1 expression increases with age in HSCs, but Aldh1a1 deficiency is not compensated by increased transcription of other ALDHS. (A) Gene expression profiles for ALDHs in Thy-1.1^lowSca-1⁺Lineage⁻c-kit⁺ HSCs, Thy-1.1^lowSca-1⁺Mac-1^lowCD4^lowB220⁻ non–self-renewing multipotent progenitors and CD45⁺ bone marrow cells from young adult mice. These data were extracted from genome-wide data published as supplementary material from an earlier study.²⁰  (B) Aldh1a1 transcript levels were compared by qPCR in CD150⁺CD48⁻CD41⁻Sca1⁺c-Kit⁺ HSCs or whole bone marrow cells independently isolated from three 2.5-month-old mice and three 22-month-old mice. cDNA content was normalized between samples based on ß-actin expression (data not shown). Aldh1a1 transcript levels are expressed in terms of fold change relative to old bone marrow (set to 1). Aldh1a1 was expressed at significantly (P < .01) higher levels in old HSCs compared with old bone marrow or young HSCs. (C) A schematic representation of Aldh1a1 showing the position of exons (numbered black boxes), 4 sets of qPCR primer binding sites (arrows), and the neomycin resistance cassette in the targeted allele. Bone marrow cells (D) or CD150⁺CD48⁻CD41⁻Sca1⁺c-Kit⁺ HSCs (E) from 3 littermate pairs aged 2.5 to 6 months were tested for expression of Aldh1a1 with the above primers, and all other Aldh family members by quantitative RT-PCR. cDNA content was normalized between samples based on ß-actin expression (data not shown) and data are expressed as fold change relative to the littermate control. #P < .1; *P < .05. Error bars represent SD. The mutant Aldh1a1 transcript was present at greatly reduced levels, and we did not detect any Aldh transcript with a compensatory increase in expression.