SUMMARY Fanconi anemia (FA) is an inherited DNA repair deficiency syndrome. FA patients undergo progressive bone marrow failure (BMF) during childhood, which frequently requires allogeneic hematopoietic stem cell transplantation. The pathogenesis of this BMF has been elusive to date. Here we found that FA patients exhibit a profound defect in hematopoietic stem and progenitor cells (HSPCs) that is present before the onset of clinical BMF. In response to replicative stress and unresolved DNA damage, p53 is hyperactivated in FA cells and triggers a late p21Cdkn1a-dependent G0/G1 cell-cycle arrest. Knockdown of p53 rescued the HSPC defects observed in several in vitro and in vivo models, including human FA or FA-like cells. Taken together, our results identify an exacerbated p53/p21 “physiological” response to cellular stress and DNA damage accumulation as a central mechanism for progressive HSPC elimination in FA patients, and have implications for clinical care.

Summary Severe sepsis remains a poorly understood systemic inflammatory condition with high mortality rates and limited therapeutic options in addition to organ support measures. Here we show that the clinically approved group of anthracyclines acts therapeutically at a low dose regimen to confer robust protection against severe sepsis in mice. This salutary effect is strictly dependent on the activation of DNA damage response and autophagy pathways in the lung, as demonstrated by deletion of the ataxia telangiectasia mutated (Atm) or the autophagy-related protein 7 (Atg7) specifically in this organ. The protective effect of anthracyclines occurs irrespectively of pathogen burden, conferring disease tolerance to severe sepsis. These findings demonstrate that DNA damage responses, including the ATM and Fancony Anemia pathways, are important modulators of immune responses and might be exploited to confer protection to inflammation-driven conditions, including severe sepsis.

Telomerase is a ribonucleoprotein enzyme complex that reverse-transcribes an integral RNA template to add short DNA repeats to the 39-ends of telomeres. G-quadruplex structure in a DNA substrate can block its extension by telomerase. We have found that hnRNP A1-which was previously implicated in telomere length regulation-binds to both single-stranded and structured human telomeric repeats, and in the latter case, it disrupts their higher-order structure. Using an in vitro telomerase assay, we observed that depletion of hnRNP A/B proteins from 293 human embryonic kidney cell extracts dramatically reduced telomerase activity, which was fully recovered upon addition of purified recombinant hnRNP A1. This finding suggests that hnRNP A1 functions as an auxiliary, if not essential, factor of telomerase holoenzyme. We further show, using chromatin immunoprecipitation, that hnRNP A1 associates with human telomeres in vivo. We propose that hnRNP A1 stimulates telomere elongation through unwinding of a G-quadruplex or G-G hairpin structure formed at each translocation step.

Progressive bone marrow failure is a major cause of morbidity and mortality in human Fanconi Anemia patients. In an effort to develop a Fanconi Anemia murine model to study bone marrow failure, we found that Fancd2 ؊/؊ mice have readily measurable hematopoietic defects. Fancd2 deficiency was associated with a significant decline in the size of the c-Kit ؉ Sca-1 ؉ Lineage ؊ (KSL) pool and reduced stem cell repopulation and spleen colony-forming capacity. Fancd2 ؊/؊ KSL cells showed an abnormal cell cycle status and loss of quiescence. In addition, the supportive function of the marrow microenvironment was compromised in Fancd2 ؊/؊ mice. Treatment with Sirt1-mimetic and the antioxidant drug, resveratrol, maintained Fancd2 ؊/؊ KSL cells in quiescence, improved the marrow microenvironment, partially corrected the abnormal cell cycle status, and significantly improved the spleen colony-forming capacity of Fancd2 ؊/؊ bone marrow cells. We conclude that Fancd2 ؊/؊ mice have readily quantifiable hematopoietic defects, and that this model is well suited for pharmacologic screening studies. IntroductionFanconi anemia (FA) is a rare, autosomal, recessive genetic disorder associated with severe birth defects, cancer predisposition, and bone marrow failure. Thirteen causative genes (FANCA, FANCB, FANCC, FANCD1/BRCA2, FANCD2, FANCE, FANCF, FANCG/XRCC9, FANCI, FANCL/PHF9/Pog, FANCJ/BRIP1/BACH1, FANCM/Hef, and FANCN/ PALB2) have been identified and cloned to date, and the encoded proteins are believed to work together in a common DNA damageresponse pathway to maintain genomic integrity and protect the genome from DNA damage induced by cross-linking agents. 1,2 Although deficiency in DNA cross-link repair renders all FA cells susceptible to cross-linking agents, bone marrow is the most affected organ system. Mutations in any of the different FA genes almost universally lead to bone marrow failure, which is the primary cause of mortality in FA. 3 The pathogenesis of bone marrow failure in FA remains elusive. Mutations in several genes involved in DNA damage repair, including Atr, XPD, and Ercc1, caused either hematopoietic stem cell (HSC) loss or impaired HSC function under conditions of stress. [4][5][6] These studies suggest that the maintenance of genome integrity is critical for HSC survival and function. However, the extent to which genotoxicity, resulting from impaired DNA damage repair, contributes to bone marrow failure in FA is unclear. 7 Other pathways associated with hematopoietic failure, such as altered cytokine signaling, may also contribute to FA pathogenesis. 8,9 For example, levels of proapoptotic cytokines tumor necrosis factor-␣ (TNF-␣) and interferon-␥ (IFN-␥) are elevated in FA lymphocytes, bone marrow cells, and FA patient serum samples. 10-12 FA bone marrow cells (at least of the C complementation group) are also hypersensitive to these cytokines and undergo apoptosis when exposed to even low levels of them. [13][14][15] To better understand FA, multiple murine knockout models, includingand Fancl Ϫ/Ϫ m...

BackgroundRetinitis pigmentosa (RP) is characterized by progressive night blindness, visual field loss, altered vascular permeability and loss of central vision. Currently there is no effective treatment available except gene replacement therapy has shown promise in a few patients with specific gene defects. There is an urgent need to develop therapies that offer generic neuro-and vascular-protective effects with non-invasive intervention. Here we explored the potential of systemic administration of pluripotent bone marrow-derived mesenchymal stem cells (MSCs) to rescue vision and associated vascular pathology in the Royal College Surgeons (RCS) rat, a well-established animal model for RP.Methodology/Principal FindingsAnimals received syngeneic MSCs (1×106 cells) by tail vein at an age before major photoreceptor loss. Principal results: both rod and cone photoreceptors were preserved (5–6 cells thick) at the time when control animal has a single layer of photoreceptors remained; Visual function was significantly preserved compared with controls as determined by visual acuity and luminance threshold recording from the superior colliculus; The number of pathological vascular complexes (abnormal vessels associated with migrating pigment epithelium cells) and area of vascular leakage that would ordinarily develop were dramatically reduced; Semi-quantitative RT-PCR analysis indicated there was upregulation of growth factors and immunohistochemistry revealed that there was an increase in neurotrophic factors within eyes of animals that received MSCs.Conclusions/SignificanceThese results underscore the potential application of MSCs in treating retinal degeneration. The advantages of this non-invasive cell-based therapy are: cells are easily isolated and can be expanded in large quantity for autologous graft; hypoimmunogenic nature as allogeneic donors; less controversial in nature than other stem cells; can be readministered with minor discomfort. Therefore, MSCs may prove to be the ideal cell source for auto-cell therapy for retinal degeneration and other ocular vascular diseases.