Marrow hematopoietic cells comprise a semi-solid tissue that differentiates into various mature cell types that then populate the peripheral blood, a fluid tissue. The transition from marrow to blood requires that the mature blood cells acquire viscoelastic properties that allow them to pass through apertures as small as three microns as they traverse capillaries and narrow fenestrations in the marrow vascular endothelium, in the splenic red pulp, and in lymph nodes. Now, in an elegant series of experiments, Dr. Jae-Won Shin and colleagues in Dr. Dennis Discher’s laboratory at the University of Pennsylvania demonstrate that, depending upon hematopoietic lineage, expression of nuclear lamins play differing roles in determining both the fate of hematopoietic progenitor cells and the viscoelastic properties of mature nucleated blood cells.
The two most common A-type lamins, lamin-A and lamin-C, are alternatively spliced products of one gene, while the two B-type lamins, B1 and B2, are products of separate genes. The lamin proteins form a mesh-like network, the nuclear lamina, which is located between and interacting with the inner membrane of the nuclear envelope and the peripheral heterochromatin of the nucleus.1 Nuclear lamins are also associated with nuclear scaffold proteins such as actin, with chromatin modifying proteins, and with transcription factors.1 These interactions can influence gene expression, nuclear shape, and nuclear deformability. Mutant A-type lamins are associated with myopathies, neuropathies, and progeria. In murine models, germ-line mutations in B-type lamins result in death early in embryogenesis, while mutant membrane receptors for B-type lamins are associated with progressive lymphopenia.2 In humans, mutations in B-type lamin receptors cause the hyposegmentation of neutrophils that characterizes the Pelger–Huet anomaly.3
Lamin-A Relative to B-Type Lamins, Transformed to a Measurable A:B Ratio Versus Calibrated Sum Intensity A+B. Log-log plot of intensity of nuclear lamins A + B versus ratio of lamin-A:lamin-B in human bone marrow and peripheral blood cells based on mean fluorescence intensity of A-type lamins and B-type lamins as quantified by mass spectrometrycalibrated intracellular flow cytometry. The dashed line demarcates cells residing in bone marrow (BM) in upper gray area and cells circulating in the peripheral blood (PB) in lower pink area. Abbreviations: CD34+,CD38- , cell population enriched in hematopoietic stem cells; CD34+,CD38+, cell population enriched in multipotent hematopoietic progenitors with limited proliferative capacity; MSC, mesenchymal stromal cells; MkP, megakaryocyte progenitors; MK, megakaryocytes; ProEry, proerythroblasts; lateEry, late-stage erythroblasts; BM-G and BM-M, bone marrow granulocytic and monocytic cells; PB GM, peripheral blood granulocytes and monocytes; T, T-lymphocytes; B, B-lymphocytes; RBC, erythrocytes; Plt, platelets.Shin J et al. Lamins regulate cell trafficking and lineage maturation of adult human hematopoietic cells. PNAS. 2013;110:18893.
Lamin-A Relative to B-Type Lamins, Transformed to a Measurable A:B Ratio Versus Calibrated Sum Intensity A+B. Log-log plot of intensity of nuclear lamins A + B versus ratio of lamin-A:lamin-B in human bone marrow and peripheral blood cells based on mean fluorescence intensity of A-type lamins and B-type lamins as quantified by mass spectrometrycalibrated intracellular flow cytometry. The dashed line demarcates cells residing in bone marrow (BM) in upper gray area and cells circulating in the peripheral blood (PB) in lower pink area. Abbreviations: CD34+,CD38- , cell population enriched in hematopoietic stem cells; CD34+,CD38+, cell population enriched in multipotent hematopoietic progenitors with limited proliferative capacity; MSC, mesenchymal stromal cells; MkP, megakaryocyte progenitors; MK, megakaryocytes; ProEry, proerythroblasts; lateEry, late-stage erythroblasts; BM-G and BM-M, bone marrow granulocytic and monocytic cells; PB GM, peripheral blood granulocytes and monocytes; T, T-lymphocytes; B, B-lymphocytes; RBC, erythrocytes; Plt, platelets.Shin J et al. Lamins regulate cell trafficking and lineage maturation of adult human hematopoietic cells. PNAS. 2013;110:18893.
Dr. Shin and colleagues used mass spectrometry-calibrated intracellular flow cytometry to determine amounts and ratios of A-type and B-type lamins in normal human marrow and in nucleated blood cells. Using data from such analyses, the investigators developed a method for distinguishing early-stage hematopoietic cells from their mature progeny by plotting the content of A-type and B-type lamins versus the ratio of A-type:B-type lamins (Figure). In other experiments, they perturbed expression of lamins in hematopoietic progenitors using specific interfering RNAs or retinoic acid, which inhibits type-A lamin expression. They also tested nuclear deformability and chemotactic factorinduced cellular migration through three micron pores to simulate transit across capillary endothelium.
Megakaryocytes contained the most total lamins, which increased with increasing ploidy, whereas the erythroid cells had the highest A-type:B-type lamin ratio, which increased with differentiation. The high lamin content of megakaryocytes and the nuclear rigidity associated with the high lamin A:lamin B ratio in erythroblasts make these marrow cell types least likely to migrate through the marrow vascular endothelium as they are the lineages that produce anucleate blood cells through proplatelet formation in the case of megakaryocytes and undergo nuclear extrusion prior to entering the circulation in the case of erythroblasts. Altered expression of A-type and B-type lamins also affected the differentiation of myeloid progenitor cells such that increased lamin-A expression and/or decreased lamin-B expression enhanced erythroid progenitor development but decreased granulocyte-monocyte progenitor growth. Lamin-A overexpression was also associated with increased megakaryocytic differentiation, while decreased lamin-A expression inhibited erythroid progenitor differentiation and enhanced granulocyte-monocyte progenitor development. With decreased lamin-A:lamin-B ratios compared with erythroid nuclei and decreased lamin amounts compared with megakaryocytic nuclei, lymphocytic and myeloid nuclei had greater nuclear deformability, consistent with circulation of the mature nucleated cells in blood. During differentiation, the lamin-A:lamin-B ratio increased in granulocytic-monocytic cells, but their total lamin content decreased. Mature granulocytes-monocytes as well as lymphocytes readily migrated through three micron pores and softening their nuclei by decreasing the lamin-A to lamin-B ratio increased the migration rate.
In Brief
Dr. Shin and colleagues demonstrated a role for nuclear lamins both in differentiation of marrow hematopoietic cells and in the nuclear flexibility required by mature nucleated blood cells to enter and remain in circulation. These results suggest that increased nuclear rigidity contributes both to the loss of circulating lymphocytes in lamin-B receptor-deficient mice2 and to the impaired chemotactic migration associated with the Pelger–Huet anomaly that is a consequence of mutant lamin B receptors in humans.4 The findings also suggest mechanisms that could account for the leukostasis observed in acute leukemias with high numbers of circulating blasts. The hypothesis in this case is that nuclei of the leukemic blast are more rigid than nuclei of normal mature granulocytic-monocyctic cells. Further, the high nuclear rigidity of circulating erythroblasts may contribute to the development of microvascular occlusions that complicate sickle cell anemia.
