Haematopoietic Stem Cells

1. Purification & the Weissman Lineage

Irving Weissman’s lab at Stanford in 1988 purified HSCs prospectively using FACS: cells that were Lineage-negative (no mature-cell markers), Sca-1⁺, c-Kit⁺, Thy-1^lo (LSK Thy1^loin mouse). These cells rescued lethally irradiated mice at single-cell level; a single cell could reconstitute all blood lineages indefinitely. The modern gold standard uses additional markers (CD150⁺, CD48⁻, CD34⁻) that enrich for long-term HSCs by another order of magnitude.

Human HSCs are defined by Lin⁻, CD34⁺, CD38⁻, CD90⁺, CD49f⁺. CD34 is clinically central — CD34-enriched populations are what gets transplanted in bone-marrow transplantation.

2. Hierarchy

The classical hierarchy (still the organising picture, with known revisions):

• LT-HSC (long-term HSC): true self-renewing, multipotent, largely quiescent (~1 division per year in human marrow). Reconstitutes a transplant recipient for life.
• ST-HSC: multipotent, limited self-renewal (weeks to months); transient engraftment.
• MPP (multipotent progenitor): multipotent, no self-renewal.
• CMP / CLP: common myeloid / common lymphoid progenitors. First lineage commitment.
• Lineage-committed progenitors: MEP (megakaryocyte/erythroid), GMP (granulocyte/monocyte), CD8/CD4 T cells, B cells, NK.
• Mature cells: RBCs (3–4 month lifespan), neutrophils (days), platelets (~10 days), lymphocytes (variable).

Single-cell RNA-seq (Velten 2017, Nestorowa 2016) has shown that this tree is simplified — lineage commitment is more continuous than the boxes suggest — but the gross topology is confirmed.

Simulation: HSC Amplification Hierarchy

3. The Bone-Marrow Niche

HSCs reside primarily in perivascular niches in the bone marrow. Two niche types:

• Endosteal niche: osteoblasts and endosteal surface; classical resting-HSC location. Supports quiescence via Tie2/Ang1, Jagged/Notch, and N-cadherin.
• Perivascular niche: sinusoids, LepR⁺ stromal cells, CAR (CXCL12-abundant reticular) cells, and sympathetic nerve fibres. Supports both quiescent and activated HSCs. CXCL12/CXCR4 signalling is critical for HSC retention (plerixafor mobilises HSCs by antagonising CXCR4; used clinically for stem-cell harvest).

The bone marrow is hypoxic (O₂ ~1–3%). HSCs preferentially use glycolysis; mitochondrial activity is low, with low ROS a key feature of quiescent LT-HSCs. Activation triggers a metabolic switch to OxPhos; if this switch fails, HSCs accumulate ROS damage and exhaust (Tothova 2007).

4. Bone-Marrow Transplantation

E. Donnall Thomas at Fred Hutchinson (Nobel 1990) developed allogeneic BMT in the 1960s as treatment for leukaemia. The procedure:

1. Myeloablation: total-body irradiation + chemotherapy eradicates the patient’s haematopoietic system (including the leukaemia).
2. Transplant: donor HSCs infused intravenously; they home to the bone marrow niche via CXCR4/CXCL12.
3. Engraftment: 14–28 days; neutrophil count recovery.
4. Graft-vs-host / graft-vs-leukaemia: donor T cells (if present) attack recipient tissues but also residual leukaemia cells. A balancing act.

BMT is now used for leukaemias (AML, ALL, CML), lymphomas (NHL, HL), multiple myeloma, aplastic anaemia, severe combined immunodeficiency (SCID), sickle-cell disease, thalassaemia. Tens of thousands of procedures annually worldwide. Autologous BMT (the patient’s own collected HSCs, used to rescue the bone marrow after high-dose chemotherapy) is simpler; allogeneic BMT provides GvL but carries GvHD risk.

5. Gene Therapy via HSCs

HSCs’ permanent engraftment makes them the perfect vector for durable gene therapy. Autologous HSCs are collected, genetically modified ex vivo, and re-infused into the myeloablated patient. The corrected cells self-renew for life.

Approved HSC gene therapies:

• Casgevy (exa-cel, Vertex/CRISPR Therapeutics, 2023): CRISPR-Cas9 editing of BCL11A erythroid enhancer to reactivate fetal haemoglobin, for sickle-cell disease and transfusion-dependent β-thalassaemia. First approved CRISPR medicine.
• Zynteglo (Bluebird, 2022): lentiviral beta-globin gene addition for transfusion-dependent beta-thalassaemia. ~$2.8M per dose.
• Strimvelis (GSK, 2016): ADA-SCID gene therapy. First ex vivo gene therapy approved in Europe.
• Lyfgenia (Bluebird, 2023): lentiviral anti-sickling HbA-T87Q for sickle-cell disease.

Clinical trials expanding rapidly for MLD, X-CGD, Fanconi anaemia, adrenoleukodystrophy. Cost and bone-marrow-transplant burden are the remaining obstacles; in vivo HSC editing (without the BMT step) is the current frontier.

6. Clonal Hematopoiesis & HSC Aging

With age, somatic mutations accumulate in HSCs; some mutations (in DNMT3A, TET2, ASXL1, JAK2, TP53) give a small fitness advantage, and the mutant clone slowly expands. By age 70, ~10–20% of individuals have a detectable clone occupying > 2% of their mature myeloid cells (clonal haematopoiesis of indeterminate potential, CHIP; Jaiswal 2014). CHIP confers increased risk of haematological malignancy but — unexpectedly — also doubles cardiovascular disease risk, because the mutant myeloid cells drive inflammation in atherosclerotic plaques. The HSC compartment is the first place in which we routinely observe age-related clonal evolution in humans, and it is reshaping our understanding of aging biology.