Bone Marrow Connexin-43 Expression Is Critical for Hematopoietic Regeneration After Chemotherapy

Abstract

Contact between bone marrow (BM) hematopoietic stem cells (HSC) and osteoblast/stromal (OS) cells has been shown to be crucial in the regulation of hematopoiesis. However, very little is known about the regulatory mechanisms of direct cell-to-cell communication in the hematopoietic microenvironment. Gap junction channels (connexons) are formed by polypeptides (connexins) arranged in hexamers and represent the best described intercellular communication system. Connexin-43 (Cx43) is expressed by BM OS cells and has been associated with the cadherin/β -catenin signaling pathway, recently reported as relevant in the OS/HSC interaction at the stem cell niche. Here, we employed an inducible gene-targeted murine approach to study the role of Cx43 in HSC proliferation and differentiation in vivo. Mx-Cre/Cx43^+/+ and Mx-Cre/Cx43^flox/flox littermates have been analyzed after gene deletion induced in vivo by the interferon-inducer poly (I)-poly (C), generating control (Cx43⁺) and Cx43-deficient (Cx43^−/−) mice. After one week, Cx43⁺ and Cx43^−/− mice were treated with 5-fluorouracil (5-FU). Cx43 expression in Cx43^−/− BM was markedly reduced (>90%) as analyzed on day +14 post-5-FU treatment. Cx43 deficiency did not induce a significant change in peripheral blood counts before 5-FU treatment, but the hematopoiesis recovery after 5-FU treatment was severely impaired as demonstrated by absence of recovery of peripheral blood counts, including profound neutropenia, anemia with reticulocytopenia, thrombocytopenia and a 5- to 8-fold decrease of cellularity and hematopoietic progenitor content (granulomacrophagic colony-forming-units (CFU-GM-), erythroid burst forming units (BFU-E) and mixed colony forming units (CFU-mix-) in BM and spleen on day +14 post-5-FU treatment. However, the femoral content of Lin⁻/c-kit⁺/Sca1⁺ cells in Cx43^−/− BM was maintained when compared to Cx43⁺ BM. Short-term competitive repopulation ability of Cx43^−/− BM cells was diminished as compared to Cx43⁺ mice, specifically for myeloid and B lymphoid cells, but showed spared long-term competitive repopulation ability with roughly normal hematopoietic differentiation. These data suggest that hematopoietic regeneration after cycle-specific chemotherapy is blocked in Cx43-deficient mice at the long-term HSC repopulating level. Cx43 expression within the BM appears to be crucial in the development of an efficient response to hematopoietic stress.

Keywords:

• Chemotherapy
• Connexin-43
• stem cells
• stress hematopoisis