Adult hematopoietic stem cells (HSCs) with serially transplantable activity comprise two subtypes. One shows a balanced output of mature lymphoid and myeloid cells; the other appears selectively lymphoid deficient. We now show that both of these HSC subtypes are present in the fetal liver (at a 1:10 ratio) with the rarer, lymphoid-deficient HSCs immediately gaining an increased representation in the fetal bone marrow, suggesting that the marrow niche plays a key role in regulating their ensuing preferential amplification. Clonal analysis of HSC expansion posttransplant showed that both subtypes display an extensive but variable self-renewal activity with occasional interconversion. Clonal analysis of their differentiation programs demonstrated functional and molecular as well as quantitative HSC subtype-specific differences in the lymphoid progenitors they generate but an indistinguishable production of multipotent and myeloid-restricted progenitors. These findings establish a level of heterogeneity in HSC differentiation and expansion control that may have relevance to stem cell populations in other hierarchically organized tissues.

IntroductionEach day, the hematopoietic system of the adult mouse produces billions of mature blood cells. The multistep process that underlies the production of these cells is controlled by complex mechanisms that enable changing physiologic demands to be met without overwhelming the system. It is now clear that a small population of cells known as hematopoietic stem cells (HSCs) are ultimately responsible for maintaining the lifelong output of new blood cells. 1 Nevertheless, a molecular understanding of what constitutes an HSC and how its key functions are maintained are still poorly understood.The existence of hematopoietic cells with the individual potential to produce large numbers of multiple blood cell types in vivo for prolonged periods was first suggested by retrospective clonal tracking experiments that used unique chromosomal, 2 and later, retroviral marking approaches. 3,4 The relatively short lifespan of many mature blood cell types and the contrasting longevity of some of the multilineage clones identified in these experiments implied an origin of the clones from undifferentiated cells with an extensive ability to divide and maintain a derivative population of similarly undifferentiated cells. Serial transplantation experiments demonstrated that such cell divisions did occur and thus provided the first definitive evidence of HSC self-renewal. 3,4 These findings prompted a search for functional end points that would allow HSCs to be specifically quantified independent of the presence of other cell types when assayed in limiting dilution transplantation strategies using suitably irradiated congenic hosts. 5,6 A sustained output of at least 1% of all the circulating white blood cells (WBCs) for at least 4 months is now widely assumed to be suitable for this purpose. 7 Interestingly, most analyses of individual pluripotent hematopoietic cells proliferating either in vivo or in vitro have generally found the self-renewal activity actually displayed to be highly variable. 3,4,8,9 How such a variable behavior is related to the molecular state of the initial cells is not well defined and remains a subject of intense interest and investigation. 10 Variations in external cues may be one contributing parameter, at least in vivo, because it is known that self-renewal responses can be directly and rapidly modulated in this way in vitro. 11,12 In addition, there is some evidence of predetermined heterogeneity in HSC self-renewal potential. This is exemplified by the differences in regenerative activity of HSCs from fetal and adult sources 13,14 and the finding of a consistent association of short-term and long-term multilineage WBC outputs with distinct phenotypes of adult bone marrow (BM) cells (eg, according to their expression of CD49b and CD34). 15,16 Taken together, these results suggest that some hematopoietic cells can remain pluripotent for several divisions even though they are already destined to undergo terminal differentiation within a finite period. This is the basis of the concept of durable versu...

Hematopoietic stem cells (HSCs) regenerated in vivo display sustained differences in their self-renewal and differentiation activities. Variations in Steel factor (SF) signaling are known to affect these functions in vitro, but the cellular and molecular mechanisms involved are not understood. To address these issues, we evaluated highly purified HSCs maintained in single-cell serum-free cultures containing 20 ng/mL IL-11 plus 1, 10, or 300 ng/mL SF. Under all conditions, more than 99% of the cells traversed a first cell cycle with similar kinetics. After 8 hours in the 10 or 300 ng/mL SF conditions, the frequency of HSCs remained unchanged. However, in the next 8 hours (ie, 6 hours before any cell divided), HSC integrity was sustained only in the 300 ng/mL SF cultures. The cells in these cultures also contained significantly higher levels of Bmi1, Lnk, and Ezh2 transcripts but not of several other regulators. Assessment of 21 first division progeny pairs further showed that only those generated in 300 ng/mL SF cultures contained HSCs and pairs of progeny with similar differentiation programs were not observed. Thus, SF signaling intensity can directly and coordinately alter the transcription factor profile and long-term repopulating ability of quiescent HSCs before their first division. (Blood. 2008;112:560-567) IntroductionThe hematopoietic system of the adult mouse is responsible for the daily production of billions of differentiated blood cells of various types. Because most of these cells have a limited lifespan and proliferative ability, they must be continuously generated from a population of more primitive cells that collectively have life-long self-sustaining ability. This function is restricted to a tiny subset of multipotent cells generally referred to as hematopoietic stem cells (HSCs). Historically, HSCs have been both defined and quantified retrospectively by their ability to generate clones containing both lymphoid and myeloid blood cells for at least 4 months when transplanted into irradiated recipients at limiting dilutions. 1 Analyses of such mice have also allowed the long-term differentiation activity of individual HSCs to be characterized. 2,3 Methods have now been developed for isolating purified populations in which 20% to 60% of the cells display this durability of reconstituting activity, indicating that intravenously injected HSCs can have very high seeding efficiencies in irradiated mice. [4][5][6][7] The ability to isolate such highly purified HSC populations has also permitted a more direct and complete analysis of their in vivo differentiation activity in single-cell transplant experiments. 6,[8][9][10][11] More recently, serial transplants of such clonally repopulated mice have been performed. Together, these experiments have revealed that HSCs possess one of 4 distinct differentiation programs that are propagated over many generations in vivo. 10,12 In addition, these studies have shown that extensive self-renewal ability in vivo is strongly associated with the display of either a...

Mechanistically driven therapies for atrial fibrillation (AF), the most common cardiac arrhythmia, are urgently needed, the development of which requires improved understanding of the cellular signaling pathways that facilitate the structural and electrophysiological remodeling that occurs in the atria. Similar to humans, increased persistent Na + current leads to the development of an atrial myopathy and spontaneous and long-lasting episodes of AF in mice. How increased persistent Na + current causes both structural and electrophysiological remodeling in the atria is unknown. We crossbred mice expressing human F1759A-Na V 1.5 channels with mice expressing human mitochondrial catalase (mCAT). Increased expression of mCAT attenuated mitochondrial and cellular reactive oxygen species (ROS) and the structural remodeling that was induced by persistent F1759A-Na + current. Despite the heterogeneously prolonged atrial action potential, which was unaffected by the reduction in ROS, the incidences of spontaneous AF, pacing-induced after-depolarizations, and AF were substantially reduced. Expression of mCAT markedly reduced persistent Na + current–induced ryanodine receptor oxidation and dysfunction. In summary, increased persistent Na + current in atrial cardiomyocytes, which is observed in patients with AF, induced atrial enlargement, fibrosis, mitochondrial dysmorphology, early after-depolarizations, and AF, all of which can be attenuated by resolving mitochondrial oxidative stress.

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Hidradenitis suppurativa (HS) is a painful, inflammatory skin disease that has historically been understudied in the pediatric population. Procedural interventions, such as surgical excisions, skin grafts, and lasers, are important for comprehensive HS disease management. However, there is a lack of data on procedural treatments for HS in pediatric patients. The purpose of this study was to conduct a systematic review of the literature on the efficacy and safety of procedural treatments for HS in pediatric patients. In April 2022, MEDLINE and EMBASE databases were searched for articles on the efficacy of procedural treatments for HS in patients <18 years of age. Two independent reviewers extracted data from relevant studies. From 1974 to 2021, 23 articles with 81 patients were identified. Patients' Hurley stages included stage I (9.1%, 1/11), II (36.4%, 4/11), and III (54.5%, 6/11). The most extensively studied procedural interventions include negative pressure wound therapy (n = 30), surgical excision with skin graft/flap (n = 19),and endoscopic electrode or laser treatment (n = 11). In all, promising response rates for procedural management strategies were observed in the literature but the findings were largely based on case reports/series. Randomized controlled trials (RCTs), especially those geared toward minimally invasive procedural treatments, are needed to help guide clinicians on the most efficacious treatment modalities for pediatric patients with HS.

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Significant advances have been made in the development of methods for purifying murine hematopoietic cells with longterm (&gt;4 months) in vivo reconstituting ability although these longterm repopulating cells (LTRCs) remain heterogeneous with regard to the self-renewal (SR) activity they display when transplanted into irradiated hosts. Furthermore our group has also identified cell culture conditions that differentially alter LTRC activity without immediate effects on their proliferation or survival. Here, we show that highly purified LTRCs with high and low SR properties can be prospectively isolated from normal adult mouse bone marrow (ABM) as 2 separate populations according to their expression of CD150 within the EPCR++CD48−CD45mid fraction of cells: 56% total LTRCs and 43% of the high SR type in the CD150+ subset vs. 39% total LTRCs and 32% of the low SR type in the CD150− subset (as determined from 62 and 28 single cell transplants, respectively). As a first test of whether these populations would likely be useful to search for new molecular differences associated with their different SR properties, we compared the level of expression in these 2 populations of a small set of genes previously reported to regulate LTRC SR activity: c-Kit, Bmi1, Gata3, Rae28, Ezh2 and Lnk by quantitative real-time PCR (Q-RT-PCR). This exercise revealed transcript levels of the first 4 of these genes to be significantly higher in the CD150+ subset that is selectively enriched in high SR LTRCs, thus validating the concept that they have a distinct molecular signature. Previous evidence shows that high SR LTRCs are present in both FL LTRCs and ABM LTRCs but they differ in some properties (i.e.: cell cycle status, regeneration kinetics). We therefore began a search for ontogeny-independent components of the SR machinery by comparing tags present in 2 LongSAGE libraries produced from CD45midlin−Rho−SP ABM cells and from lin−Sca1+CD43+Mac1+ embryonic day 14.5 fetal liver (FL) cells (each 20–30% total LTRCs and 12–20% of the high SR type, as determined by 132 (FL) and 352 (ABM) single cell transplants, respectively). From these comparisons and additional data in other publicly available datasets for primitive murine hematopoietic cells, we identified 28 genes not previously shown to have a functional role in LTRC SR control. We then compared the level of expression of these 28 genes between the CD150+ subsets of EPCR++CD48−CD45mid ABM cells and FL cells (24% total LTRCs and 12% high SR LTRCs in the FL subset) and their respective downstream lin− progeny. This comparison revealed 10 of these genes to be down-regulated in the lin− populations of both ABM and FL. Further comparison of the expression of these 10 genes between the high vs. low SR LTRCs (found in the CD150+ and CD150− subsets of EPCR++CD48−CD45mid) ABM cells showed the expression of 5 (Vwf, Rhob, Pld3, Prnp and Smarcc2) to be downregulated in the CD150− (low SR LTRC) subset. Interestingly, the first 4 of these genes, as well as 2 of the preliminary set of SR regulators (Bmi1 and Gata3), were also selectively down-regulated in EPCR++CD150+CD48−CD45mid ABM cells that had been incubated for 16 hours in 1 or 10 ng/ml Steel factor + 20 ng/ml IL-11 (conditions that decrease LTRC activity in vivo 4–5-fold before any of these divide or die). Taken together, these results point to the existence of more, although a rather small number of additional genes, including Vwf, Rhob, Pld3, and Prnp, whose products may be involved in controlling the SR potential of normal mouse LTRCs.