ALBERT

Description: Evolutionary adaptations to high altitude in Tibetans, Ethiopians, and Andean populations of South America have shown that Tibetans and Ethiopians have normal hemoglobin %, while most of Aymara and Quechua of the Andean highlands are polycythemic. Whole genome sequencing (WGS) in Quechua identified enriched SENP1 and ANP32D genes correlating with polycythemia (Zhou et al, Am J Hum Genet. 2013 Sep 5; 93(3): 452-462) but these genes were neither enriched nor segregated with polycythemia in Aymara. Instead, we identified that genes enriched in Aymara are related to regulation of cardiovascular development in high-altitude adapted Andeans, BRINP3, NOS2, and TBX5 (Crawford et al, Am J Hum Genet. 2017 Nov 2;101(5):752-767). To further search for Aymara propensity to polycythemia, we analyzed transcriptomes from Aymara and Europeans living in La Paz, Bolivia (3,639-4,150m) from limited amount of peripheral blood reticulocytes, platelets and granulocytes, but only granulocyte RNA was adequate for unbiased whole transcriptome analyses. In Aymaras, 2,585 genes were upregulated and 365 genes were downregulated (Adjp

Description: The transmembrane 4 superfamily (TM4SF) has come into prominence for its association with a wide range of cell surface molecules, especially integrins. We report that TM4SF molecules CD9, CD63, and CD81 are physically associated with c-kit receptor tyrosine kinase in the human factor–dependent myeloid cell line, MO7e. We characterized this complex using coimmunoprecipitation and colocalization methods. The c-kit coimmunoprecipitated with anti-TM4SF antibodies showed several distinct phenotypes compared to the total c-kit immunoprecipitated with anti–c-kit antibody. These included: (1) higher basal level of tyrosine phosphorylation without elevated kinase activity in the absence of Steel factor (SLF), (2) deficient enhancement of tyrosine phosphorylation and kinase activity in response to SLF, (3) elevated binding rate of SLF shown in chemical cross-linking studies, and (4) little internalization and degradation after SLF treatment. Cocapping studies in living cells showed that c-kit colocalized with TM4SF molecules after SLF stimulation, suggesting confirmation of the biochemical data obtained by the coimmunoprecipitation studies. Colocalization of c-kit with CD81 by SLF was also observed in cord blood CD34+ cells, suggesting the existence of functional units of c-kit in TM4SF complexes in primary hematopoietic cells. This suggests that some TM4SF members may negatively modulate function of c-kit receptor tyrosine kinase and thus regulate receptor sensitivity to SLF in hematopoietic progenitors.

Description: Stromal cell–derived factor 1 (SDF-1/CXCL12) is a multifunctional cytokine. We previously reported that myelopoiesis was enhanced in SDF-1α transgenic mice, probably due in part to SDF-1α enhancement of myeloid progenitor cell (MPC) survival. To understand signaling pathways involved in this activity, we studied the effects on factor-dependent cell line MO7e cells incubated with SDF-1α alone or in combination with other cytokines. SDF-1α induced transient activation of extracellular stress–regulated kinase (ERK1/2), ribosomal S6 kinase (p90RSK) and Akt, molecules implicated in cell survival. Moreover, ERK1/2, p90RSK, and Akt were synergistically activated by SDF-1α in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF), Steel factor (SLF), or thrombopoietin (TPO). Similar effects were seen after pretreatment of MO7e cells with SDF-1α followed by stimulation with the other cytokines, suggesting a priming effect of SDF-1α. Nuclear factor-κB (NF-κB) did not appear to be involved in SDF-1α actions, alone or in combination with other cytokines. These intracellular effects were consistent with enhanced myeloid progenitor cell survival by SDF-1α after delayed addition of growth factors. SDF-1α alone supported survival of highly purified human cord blood CD34+++ cells, less purified human cord blood, and MO7e cells; this effect was synergistically enhanced when SDF-1α was combined with low amounts of other survival-promoting cytokines (GM-CSF, SLF, TPO, and FL). SDF-1 may contribute to maintenance of MPCs in bone marrow by enhancing cell survival alone and in combination with other cytokines.

Description: Cell cycle checkpoints ensure orderly progression of events during cell division. A microtubule damage (MTD)-induced checkpoint has been described in G1 phase of the cell cycle (G1MTC) for which little is known. The present study shows that the G1MTC is intact in activated T lymphocytes from mice with the p21waf-1 gene deleted. However, p21waf-1 gene deletion does affect the ratio of cells that arrest at the G1MTC and the spindle checkpoint after MTD. The G1MTC arrests T lymphocytes in G1 prior to cdc2 up-regulation and prior to G1arrest by p21waf-1. Once cells have progressed past the G1MTC, they are committed to chromosome replication and metaphase progression, even with extreme MTD. The G1MTC is also present in a human myeloid cell line deficient in p21waf-1gene expression. The p21-independent G1MTC may be important in cellular responses to MTD such as those induced by drugs used to treat cancer.

Description: Stromal cell-derived factor-1 (SDF-1) plays a central role in the homing of hematopoietic stem cells and progenitor cells (HPCs) to bone marrow (BM). Tissue damage, most notably ischemia, induces the recruitment of bone marrow (BM)-derived stem/progenitor cells, which can lead to tissue repair. Moreover, damaged tissues and endothelium express and produce SDF-1. However, the effects of aberrant expression and production of SDF-1 in human endothelium on the biology of BM-derived stem/progenitor cells are not fully understood. To clarify this, we over-expressed the SDF-1 gene in human umbilical vein endothelial cells using an adenoviral vector (HUVEC/AdeSDF-1) and examined the transmigration, maintenance, and proliferation of HPCs supported by the endothelium. HUVEC/AdeSDF-1 monolayers induced the migration of MO7e cells and mobilized peripheral blood (mPB) CD34+ cells underneath the endothelium within a few hours, while HUVEC monolayers transfected with adenoviral vectors expressing LacZ (HUVEC/AdeLacZ) did not. A transmigration assay revealed that HUVEC/AdeSDF-1 monolayers supported the spontaneous transmigration of CD34+ cells to a greater degree than did HUVEC/LacZ monolayers (15 ± 4% vs. 5 ± 3%) and that transmigration was blocked with pretreatment of the input cells with pertussis toxin (PTX). The co-culturing of mPB CD34+ cells with HUVEC/AdeSDF-1 for up to 2 weeks led to a greater expansion of CD45+ cells and colony-forming cells, and reduced cell apoptosis. Furthermore, the co-culturing of mPB CD34+ cells with HUVEC/AdeSDF-1 led to the formation of numerous cobblestone areas, while mPB CD34+ cells plus HUVEC/AdeLacZ supported the formation of only a few cobblestone areas (253 ± 28 vs. 10 ± 3 per 2 x 104 mPB CD34+ cells at week-2), which were further enhanced by adding a combination of early-acting hematopoietic growth factors at low concentrations. To further elucidate whether SDF-1 is directly involved in the formation of cobblestone areas, mPB CD34+ cells were co-cultured with murine BM MS-5 stromal cells. Weekly PTX treatment, beginning 24 hours after start of co-culturing, markedly decreased the numbers of cobblestone areas by weeks 2 and 5, which indicates that SDF-1 is involved not only in the localization of primitive HPCs underneath the endothelium or BM stroma but also in the maintenance and proliferation of the cells. These results indicate that aberrant SDF-1 over-expression in any non-specialized endothelium induces BM-derived progenitor cell trafficking into extravasculr sites and promotes the maintenance and proliferation of the migrated cells.