Radiation-associated bone marrow (BM) injury is one of the most serious limiting factors of radiotherapy. Radiation-induced hematopoietic injury, no matter how transient or long lasting, can ultimately impair HSC function and decrease the HSC reserve, leading to increased risk for the development of BM failure or cancer. However, molecular mechanisms underlying radiation-induced HSC functional decline are largely unknown.

We previously identified a stem cell regulatory gene, latexin (Lxn), as a novel negative regulator of HSCs in mice. HSCs in Lxn knockout mice (Lxn-/-) had increased self-renewal and survival. In our new findings, we surprisingly found that Lxn-/- mice had the significant survival advantages under lethal dose of total body irradiation (TBI). We further found that HSCs and hematopoietic progenitor cells (HPCs), measured by immunophenotypes and colony assay, recovered much faster in Lxn-/- mice than wild-type mice (WT) within one month after sub-lethal dose of TBI. The better preserved HSC/HPC pool was due to the decreased apoptosis in which the percentage of Annexin V + PI- apoptotic HSCs/HPCs cells was significantly lower in Lxn-/- mice than WT mice. These data suggest that Lxn inactivation protects HSCs and HPCs from radiation-induced cell death, thus mitigating acute hematopoietic suppression and conferring a survival advantage.

To determine the long-term effect of TBI on Lxn-/- HSCs, we performed limiting dilution competitive repopulation unit assay (CRU), and found that Lxn-/- CRU was significantly higher than WT CRU. Moreover, we performed serial transplantation experiment, and found that Lxn-/- HSC continuously regenerated blood and bone marrow cells even at the 4th round of transplantation whereas WT HSCs were exhausted. These data provide robust evidence that Lxn inactivation protects functional long-term HSCs from radiation-induced injury. Radiation can increase the risk of hematological malignancy later in the life. We thus maintained a group of mice that were subject to either a single dose of 6.5Gy TBI or split low doses of TBI (2 Gy daily for 6 days), and monitored their gross condition and blood cell counts for 20 months. At 20 month post-radiation, we performed bone marrow analysis and histopathology analysis. We found that Lxn-/- mice did not spontaneously develop hematopoietic malignancies, their bone marrow HSCs/HPCs had normal population size, and bone marrow had normal histopathology. These data suggest that Lxn inactivation mitigates radiation-induced short-term myelosuppression and long-term HSC functional impairment without induction of hematologic malignancy.

At the molecular level, we previously reported that Lxn sensitized leukemogenic cells to gamma-irradiation-induced cell-cycle arrest and cell death through Rps3 pathway, and Rps3 was a binding protein of Lxn. Rps3 has been shown to be involved in the NFkB pathway. We found that Rps3 bound Lxn in primary hematopoietic stem and progenitor cells (HSPCs) using Co-IP assay. Lxn-/- HSPCs had the increased expression of Rps3 and NFkB p65 before or post-irradiation. Knockdown of Rps3 in Lxn-/- HSPCs decreased NFkB p65 and increased radiation-induced apoptosis. Moreover, when Lxn-/- HSPCs were treated with NFkB p65 specific inhibitor, the similar phenotypes were also shown, suggesting that Lxn functions through Rps3-NFkB-mediated pro-survival pathway in primary HSPCs. We are currently proving this molecular pathway using the in vivo model by crossing p65 knockout mice with Lxn-/- mice.

In conclusion, latexin inhibition mitigates irradiation induced hematopoietic injury via Rps3-NFkB-mediated pro-survival pathway.


No relevant conflicts of interest to declare.

Author notes


Asterisk with author names denotes non-ASH members.