Ionizing radiation and chemotherapy deplete hematopoietic stem cells and damage the vascular niche wherein hematopoietic stem cells reside. Hematopoietic stem cell regeneration requires signaling from an intact bone marrow (BM) vascular niche, but the mechanisms that control BM vascular niche regeneration are poorly understood. We report that BM vascular endothelial cells secrete semaphorin 3 A (SEMA3A) in response to myeloablation and SEMA3A induces p53 – mediated apoptosis in BM endothelial cells via signaling through its receptor, Neuropilin 1 (NRP1), and activation of cyclin dependent kinase 5. Endothelial cell – specific deletion of Nrp1 or Sema3a or administration of anti-NRP1 antibody suppresses BM endothelial cell apoptosis, accelerates BM vascular regeneration and concordantly drives hematopoietic reconstitution in irradiated mice. In response to NRP1 inhibition, BM endothelial cells increase expression and secretion of the Wnt signal amplifying protein, R spondin 2. Systemic administration of anti - R spondin 2 blocks HSC regeneration and hematopoietic reconstitution which otherwise occurrs in response to NRP1 inhibition. SEMA3A – NRP1 signaling promotes BM vascular regression following myelosuppression and therapeutic blockade of SEMA3A – NRP1 signaling in BM endothelial cells accelerates vascular and hematopoietic regeneration in vivo.

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Recent events encompassing social injustices, healthcare disparities, and police brutality against Black citizens highlight the continued need to strive toward unbiased and inclusive practices in all realms of the world. Our voices as cell biologists are powerful tools that can be used to combat inequities in the scientific landscape. In this inaugural Voices essay, we discuss how exclusion and inclusion events have contributed to our scientific journeys and how scientists can work to create an inclusive environment for our trainees and colleagues. As underrepresented minority scientists in the early and late stages of our scientific training, we frame the trainee experience to provide insight from unique perspectives. This essay also provides actionable items that the cell biology community can implement to promote inclusivity. We anticipate that initiating an open dialogue focused on diversity and inclusion will promote growth in the field of cell biology and enable scientists to assess and assume their role in creating welcoming environments. We believe that scientists at all stages in their careers can make meaningful and habitual contributions to supporting inclusivity in cell biology, thereby creating a future where diversity, equity, and inclusion are expected, not requested.

Chemotherapy and irradiation cause DNA damage to hematopoietic stem cells (HSCs), leading to HSC depletion and dysfunction and risk for malignant transformation over time. Extrinsic regulation of HSC DNA repair is not well understood and therapies to augment HSC DNA repair following myelosuppression remain undeveloped. We report that epidermal growth factor receptor (EGFR) regulates DNA repair in HSCs following irradiation via activation of the DNA-dependent protein kinase - catalytic subunit (DNA-PKcs) and nonhomologous end joining (NHEJ). We show that hematopoietic regeneration in vivo following total body irradiation (TBI) is dependent upon EGF - mediated repair of DNA damage via activation of DNA-PKcs. Conditional deletion of EGFR in hematopoietic stem/progenitor cells (HSPCs) significantly decreased DNA-PKcs activity following irradiation, causing increased HSC DNA damage and depressed HSC recovery over time. Systemic administration of EGF promoted HSC DNA repair and rapid hematologic recovery in chemotherapy - treated mice and had no effect on AML growth in vivo. Further, EGF treatment drove the recovery of human HSCs capable of multilineage in vivo repopulation following radiation injury. Whole genome sequence analysis revealed no increase in coding region mutations in HSPCs from EGF - treated mice, but increased intergenic copy number variant mutations were detected. These studies demonstrate that EGF promotes HSC DNA repair and hematopoietic regeneration in vivo via augmentation of NHEJ. EGF has therapeutic potential to promote human hematopoietic regeneration and further studies are warranted to assess long-term hematopoietic effects.

The discovery of novel hematopoietic stem cell (HSC) surface markers can enhance understanding of HSC identity and function. We have discovered a population of primitive bone marrow (BM) HSCs distinguished by their expression of the heparan sulfate proteoglycan, Syndecan-2, which serves as both a marker and regulator of HSC function. Syndecan-2 expression was increased 10-fold in CD150+CD48-CD34-c-Kit+Sca-1+Lineage- cells (long-term - HSCs, LT-HSCs) compared to differentiated hematopoietic cells. Isolation of BM cells based solely on Syndecan-2 surface expression produced a 24-fold enrichment for LT-HSCs, 6-fold enrichment for alpha-catulin+c-kit+ HSCs, and yielded HSCs with superior in vivo repopulating capacity compared to CD150+ cells. Competitive repopulation assays revealed the HSC frequency to be 17-fold higher in Syndecan-2+CD34-KSL cells compared to Syndecan-2-CD34-KSL cells and indistinguishable from CD150+CD34-KSL cells. Syndecan-2 expression also identified nearly all repopulating HSCs within the CD150+CD34-KSL population. Mechanistically, Syndecan-2 regulates HSC repopulating capacity through control of expression of Cdkn1c (p57) and HSC quiescence. Loss of Syndecan-2 expression caused increased HSC cell cycle entry, downregulation of Cdkn1c and loss of HSC long-term - repopulating capacity. Syndecan-2 is a novel marker of HSCs which regulates HSC repopulating capacity via control of HSC quiescence.