Adult Stem Cells Research Articles

Comparison of intermittent intravenous tacrolimus to continuous tacrolimus in adult allogeneic stem cell transplant recipients.

Introduction: Initial continuous intravenous (CIV) tacrolimus (0.03 mg/kg/day based on ideal body weight [IBW]) has been favored for graft versus host disease (GVHD) prevention in allogeneic stem cell transplant patients due to the consistent, steady-state degree of immunosuppression; however, this method poses many logistical challenges. We implemented intermittent (IIV) tacrolimus at a starting dose of 0.015 mg/kg IBW twice daily over 4 h. To our knowledge this is the first retrospective comparison of CIV to IIV tacrolimus. Objectives: The primary objective was to evaluate the safety of IIV tacrolimus in comparison to CIV with respect to nephrotoxicity and neurotoxicity. The secondary objectives were to compare the incidence of grade II-IV acute GVHD (aGVHD) and chronic GVHD (cGVHD) at day +180, outcomes including relapse and overall survival, cell engraftment, and reactivation of cytomegalovirus and Epstein-Barr virus. Methods: This retrospective, single-center review evaluated adults who received an allogeneic stem cell transplant patients between January 1, 2020, and December 31, 2022. Results: Fifty-one unique patients were eligible for evaluation - 28 in the IIV cohort and 23 in the CIV group. The number of patients who developed nephrotoxicity and neurotoxicity were comparable between groups with no significant differences noted. No severe neurotoxicity was identified in either population. Secondary objectives revealed no significant difference in GVHD incidence or survival outcomes. Conclusion: IIV tacrolimus is comparable to CIV in terms of safety while also maintaining similar outcomes at day +180. IIV is a safe and feasible alternative to CIV in adult allogeneic stem cell transplant recipients.

Modulation of Human Bone Marrow Mesenchymal Stem Cells (BMMSC) by Nigella Sativa and Trigona Honey: An in vitro Study

Maintenance of bone marrow mesenchymal stem cells (BMMSC) numbers is crucial for tissue repair and regeneration since adult stem cells are often limited in number, expansion capacity and lifespan. It is believed that certain types of foods are beneficial in the preservation and stimulation of stem cells throughout life. Black seeds and stingless bee honey are among the widely investigated functional food with general health promoting benefit. We aim to explore the proliferative, anti-apoptosis and anti-senescence effect of Nigella sativa and Trigona honey on BMMSC. Cell proliferation was evaluated using MTT assay. We performed flow cytometric analysis to verify stem cell surface markers while quantitative PCR was done to measure the relative expression of target genes. Results showed favourable concentration dependent enhancement of cell proliferation by 1 to 10 ug/mL Nigella sativa and 0.097% to 0.195% Trigona honey. Observation on BMMSC’s morphology and surface markers expression revealed no alteration on BMMSC stemness properties. In addition, gene expression analysis supported that there was a significant (P<0.05) increase in proliferation related gene, β-Catenin, and lower expression of apoptotic- and senescence-related gene, BAX and p21, respectively. These preliminary findings suggest a promising effect of Nigella sativaand Trigona honey on stem cell proliferation.

Kinase-inactivated CDK6 preserves the long-term functionality of adult hematopoietic stem cells.

Hematopoietic stem cells (HSCs) are characterized by the ability to self-renew and to replenish the hematopoietic system. The cell-cycle kinase cyclin dependent-kinase 6 (CDK6) regulates transcription, whereby it has both kinase-dependent and kinase-independent functions. We here describe the complex role of CDK6, balancing quiescence, proliferation, self-renewal and differentiation in activated HSCs. Mouse HSCs expressing kinase-inactivated CDK6 show enhanced long-term repopulation and homing, whereas HSCs lacking CDK6 have impaired functionality. The transcriptomes of basal and serially transplanted HSCs expressing kinase-inactivated CDK6 exhibit an expression pattern dominated by HSC quiescence and self-renewal, proposing a concept where MAZ and NFY-A are critical CDK6 interactors. Pharmacologic kinase inhibition with a clinically used CDK4/6 inhibitor in murine and human HSCs validated our findings and resulted in increased repopulation capability and enhanced stemness. Our findings highlight a kinase-independent role of CDK6 in long-term HSC functionality. CDK6 kinase inhibition represents a possible strategy to improve HSC fitness.

Evaluation of Skin Wound Healing with Biosheets Containing Somatic Stem Cells in a Dog Model: A Pilot Study

The administration of mesenchymal stem cells (MSCs) has a positive effect on wound healing; however, the lack of adequate MSC engraftment at the wound site is a major limiting factor in current MSC-based therapies. In this study, a biosheet prepared using in-body tissue architecture (iBTA) was used as a material to address these problems. This study aimed to assess and evaluate whether biosheets containing somatic stem cells would affect the wound healing process in dogs. Biosheets were prepared by subcutaneously embedding molds in beagles. These were then evaluated grossly and histologically, and the mRNA expression of inflammatory cytokines, interleukins, and Nanog was examined in some biosheets. Skin defects were created on the skin of the beagles to which the biosheets were applied. The wound healing processes of the biosheet and control (no biosheet application) groups were compared for 8 weeks. Nanog mRNA was expressed in the biosheets, and SSEA4/CD105 positive cells were observed histologically. Although the wound contraction rates differed significantly in the first week, the biosheet group tended to heal faster than the control group. This study revealed that biosheets containing somatic stem cells may have a positive effect on wound healing.

A novel method for culturing enteric neurons generates neurospheres containing functional myenteric neuronal subtypes

BackgroundThe enteric nervous system (ENS) is comprised of neurons, glia, and neural progenitor cells that regulate essential gastrointestinal functions. Advances in high-efficiency enteric neuron culture would facilitate discoveries surrounding ENS regulatory processes, pathophysiology, and therapeutics. New methodDevelopment of a simple, robust, one-step method to culture murine enteric neurospheres in a 3D matrix that supports neural growth and differentiation. ResultsMyenteric plexus cells isolated from the entire length of adult murine small intestine formed ≥3000 neurospheres within 7 days. Matrigel-embedded neurospheres exhibited abundant neural stem and progenitor cells expressing Sox2, Sox10 and Msi1 by day 4. By day 5, neural progenitor cell marker Nestin appeared in the periphery of neurospheres prior to differentiation. Neurospheres produced extensive neurons and neurites, confirmed by Tubulin beta III, PGP9.5, HuD/C, and NeuN immunofluorescence, including neural subtypes Calretinin, ChAT, and nNOS following 8 days of differentiation. Individual neurons within and external to neurospheres generated depolarization induced action potentials which were inhibited in the presence of sodium channel blocker, Tetrodotoxin. Differentiated neurospheres also contained a limited number of glia and endothelial cells. Comparison with existing methodsThis novel one-step neurosphere growth and differentiation culture system, in 3D format (in the presence of GDNF, EGF, and FGF2), allows for ∼2-fold increase in neurosphere count in the derivation of enteric neurons with measurable action potentials. ConclusionOur method describes a novel, robust 3D culture of electrophysiologically active enteric neurons from adult myenteric neural stem and progenitor cells.

Adult hair follicle stem cells differentiate into neuronal cells in explanted rat intestinal tissue.

Hair follicle stem cells (HFSCs) are adult stem cells located in the outer root sheath of the follicle bulge with high neural plasticity, which promise a potential for the stem cell therapy for neurological diseases. Hirschsprung's disease (HD) is characterized by the absence of ganglia in the distant bowel. In this study, we elucidated the capacity of HFSCs to differentiate into neuronal cells in the aganglionic colon from embryonic rat. HFSCs were isolated from adult Sprague-Dawley (SD) rats and formed spheres that could be passaged. The cultured HFSCs expressed neural crest stem cells (NCSCs) markers such as SOX10, CD34, and nestin, which indicated their neural crest lineage. Subsequent differentiation assays demonstrated that these cells could give rise to neural progeny that expressed neuronal or glial markers. The aganglionic colon from the embryonic intestine was applied as in vitro explant to test the capacity of proliferation and differentiation of HFSCs. The HFSCs expressing GFP or RFP integrated in intestinal explants and maintained proliferative capacity. Moreover, the HFSCs differentiated into Tuj1- or S100β-positive cells in the cultured intestinal explants. The results proposed that the HFSCs might be an alternative source of neural stem cells for the HD therapy.

The ATF4-RPS19BP1 axis modulates ribosome biogenesis to promote erythropoiesis

Hematopoietic differentiation is controlled by intrinsic regulators and the extrinsic hematopoietic niche. Activating transcription factor 4 (ATF4) plays a crucial role in the function of fetal and adult hematopoietic stem cell maintenance; however, the precise function of ATF4 in the bone marrow niche and the mechanism by which ATF4 regulates adult hematopoiesis remain largely unknown. Here, we employ four cell-type-specific mouse Cre lines to achieve conditional knockout of Atf4 in Cdh5+ endothelial cells, Prx1+ bone marrow stromal cells, Osx+ osteo-progenitor cells, and Mx1+ hematopoietic cells, and uncover the role of Atf4 in niche cells and hematopoiesis. Intriguingly, depletion of Atf4 in niche cells does not affect hematopoiesis; however, Atf4-deficient hematopoietic cells exhibit erythroid differentiation defects, leading to hypoplastic anemia. Mechanistically, ATF4 mediates direct regulation of Rps19bp1 transcription, which is, in turn, involved in 40S ribosomal subunit assembly to coordinate ribosome biogenesis and promote erythropoiesis. Finally, we demonstrate that under conditions of 5-fluorouracil-induced stress, Atf4 depletion impedes the recovery of hematopoietic lineages, which requires efficient ribosome biogenesis. Taken together, our findings highlight the indispensable role of the ATF4-RPS19BP1 axis in the regulation of erythropoiesis.

Quiescent Adult Neural Stem Cells: Developmental Origin and Regulatory Mechanisms.

The existence of neural stem cells (NSCs) in the adult mammalian nervous system, although small in number and restricted to the sub-ventricular zone of the lateral ventricles, the dentate gyrus of the hippocampus, and the olfactory epithelium, is a gift of evolution for the adaptive brain function which requires persistent plastic changes of these regions. It is known that most adult NSCs are latent, showing long cell cycles. In the past decade, the concept of quiescent NSCs (qNSCs) has been widely accepted by researchers in the field, and great progress has been made in the biology of qNSCs. Although the spontaneous neuronal regeneration derived from adult NSCs is not significant, understanding how the behaviors of qNSCs are regulated sheds light on stimulating endogenous NSC-based neuronal regeneration. In this review, we mainly focus on the recent progress of the developmental origin and regulatory mechanisms that maintain qNSCs under normal conditions, and that mobilize qNSCs under pathological conditions, hoping to give some insights for future study.

Porphyrin overdrive rewires cancer cell metabolism.

All cancer cells reprogram metabolism to support aberrant growth. Here, we report that cancer cells employ and depend on imbalanced and dynamic heme metabolic pathways, to accumulate heme intermediates, that is, porphyrins. We coined this essential metabolic rewiring "porphyrin overdrive" and determined that it is cancer-essential and cancer-specific. Among the major drivers are genes encoding mid-step enzymes governing the production of heme intermediates. CRISPR/Cas9 editing to engineer leukemia cell lines with impaired heme biosynthetic steps confirmed our whole-genome data analyses that porphyrin overdrive is linked to oncogenic states and cellular differentiation. Although porphyrin overdrive is absent in differentiated cells or somatic stem cells, it is present in patient-derived tumor progenitor cells, demonstrated by single-cell RNAseq, and in early embryogenesis. In conclusion, we identified a dependence of cancer cells on non-homeostatic heme metabolism, and we targeted this cancer metabolic vulnerability with a novel "bait-and-kill" strategy to eradicate malignant cells.

Cell death as an architect of adult skin stem cell niches.

Our skin provides a physical and immunological barrier against dehydration and environmental insults ranging from microbial attacks, toxins and UV irradiation to wounding. Proper functioning of the skin barrier largely depends on the interplay between keratinocytes- the epithelial cells of the skin- and immune cells. Two spatially distinct populations of keratinocyte stem cells (SCs) maintain the epidermal barrier function and the hair follicle. These SCs are inherently long-lived, but cell death can occur within their niches and impacts their functionality. The default cell death programme in skin is apoptosis, an orderly and non-inflammatory suicide programme. However, recent findings are shedding light on the significance of various modes of regulated necrotic cell death, which are lytic and can provoke inflammation within the local skin environment. While the presence of dying cells was generally regarded as a mere consequence of inflammation, findings in various human dermatological conditions and experimental mouse models of aberrant cell death control demonstrated that cell death programmes in keratinocytes (KCs) can drive skin inflammation and even tumour initiation. When cells die, they need to be removed by phagocytosis and KCs can function as non-professional phagocytes of apoptotic cells with important implications for their SC capacities. It is becoming apparent that in conditions of heightened SC activity, distinct cell death modalities differentially impact the different skin SC populations in their local niches. Here, we describe how regulated cell death modalities functionally affect epidermal SC niches along with their relevance to injury repair, inflammatory skin disorders and cancer.

Age-related reduction in the functional properties of adult stem cells located in the peripheral region of human retinal pigment epithelium.

Adult stem cells (SCs) with self-renewal and multilineage potential have been reported upon culturing human retinal pigment epithelial (RPE) cells. The current study aimed to identify the location of SCs in human RPE and to elucidate the age-related changes. Peripheral, equatorial, and central RPE cells from donors of three age groups were analyzed for their sphere-forming, clonal, and label-retaining cell properties. Furthermore, native human RPE flatmounts were immunostained for SC and proliferating cell markers. Cells with higher sphere-forming and clonal ability were identified only in young donors (<30 years) and were restricted to the periphery. Upon culturing, cells from peripheral and equatorial regions had the label-retaining cell (LRC) property. With aging, the LRCs were restricted to the periphery and were reduced. In young donors, Ki67+ proliferating cells were not observed in native RPE. However, such cells were observed in the peripheral RPE of older donors correlating with the need for regeneration. The native RPE cells were negative for SC marker expression. The above findings highlighted the presence of SCs with the ability to proliferate in the peripheral RPE and a reduction in these functional properties of SCs with aging.

A retrospective study of intravenous pentamidine for Pneumocystis jirovecii pneumonia prophylaxis in adult patients with hematologic malignancies—its utility during respiratory virus pandemics

ObjectivesIn hematology, prophylaxis for Pneumocystis jirovecii pneumonia (PCP) is recommended for patients undergoing hematopoietic stem cell transplantation and in selected categories of intensive chemotherapy for hematologic malignancies. Trimethoprim-sulfamethoxazole (TMP-SMX) is the recommended first-line agent; however, its use is not straightforward. Inhaled pentamidine is the recommended second-line agent; however, aerosolized medications were discouraged during respiratory virus outbreaks, especially during the COVID-19 pandemic, in view of potential contamination risks. Intravenous (IV) pentamidine is a potential alternative agent. We evaluated the effectiveness and tolerability of IV pentamidine use for PCP prophylaxis in adult allogeneic hematopoietic stem cell transplantation recipients and patients with hematologic malignancies during COVID-19. ResultsA total of 202 unique patients who received 239 courses of IV pentamidine, with a median of three doses received (1-29). The largest group of the patients (49.5%) who received IV pentamidine were undergoing or had received a hematopoietic stem cell transplant. The most common reason for not using TMP-SMX prophylaxis was cytopenia (34.7%). We have no patients who had breakthrough PCP infection while on IV pentamidine. None of the patients developed an infusion reaction or experienced adverse effects from IV pentamidine. ConclusionsPentamidine administered IV monthly is safe and effective.

Tracking of Nascent Matrix Deposition during Muscle Stem Cell Activation across Lifespan Using Engineered Hydrogels.

Adult stem cells occupy a niche that contributes to their function, but how stem cells rebuild their microenvironment after injury remains an open-ended question. Herein, biomaterial-based systems and metabolic labeling are utilized to evaluate how skeletal muscle stem cells deposit extracellular matrix. Muscle stem cells and committed myoblasts are observed to generate less nascent matrix than muscle resident fibro-adipogenic progenitors. When cultured on substrates that matched the stiffness of physiological uninjured and injured muscles, muscle stem cells increased nascent matrix deposition with activation kinetics. Reducing the ability to deposit nascent matrix by an inhibitor of vesicle trafficking (Exo-1) attenuated muscle stem cell function and mimicked impairments observed from muscle stem cells isolated from old muscles. Old muscle stem cells are observed to deposit less nascent matrix than young muscle stem cells, which is rescued with therapeutic supplementation of insulin-like growth factors. These results highlight the role of nascent matrix production with muscle stem cell activation.

Current Perspectives and Progress in Preoperative Portal Vein Embolization with Stem Cell Augmentation (PVESA).

Portal vein embolization with stem cell augmentation (PVESA) is an emerging approach for enhancing the growth of the liver segment that will remain after surgery (i.e., future liver remnant, FLR) in patients with liver cancer. Conventional portal vein embolization (PVE) aims to induce preoperative FLR growth, but it has a risk of failure in patients with underlying liver dysfunction and comorbid illnesses. PVESA combines PVE with stem cell therapy to potentially improve FLR size and function more effectively and efficiently. Various types of stem cells can help improve liver growth by secreting paracrine signals for hepatocyte growth or by transforming into hepatocytes. Mesenchymal stem cells (MSCs), unrestricted somatic stem cells, and small hepatocyte-like progenitor cells have been used to augment liver growth in preclinical animal models, while clinical studies have demonstrated the benefit of CD133 + bone marrow-derived MSCs and hematopoietic stem cells. These investigations have shown that PVESA is generally safe and enhances liver growth after PVE. However, optimizing the selection, collection, and application of stem cells remains crucial to maximize benefits and minimize risks. Additionally, advanced stem cell technologies, such as priming, genetic modification, and extracellular vesicle-based therapy, that could further enhance efficacy outcomes should be evaluated. Despite its potential, PVESA requires more investigations, particularly mechanistic studies that involve orthotopic animal models of liver cancer with concomitant liver injury as well as larger human trials.

Annelid adult cell type diversity and their pluripotent cellular origins

Many annelids can regenerate missing body parts or reproduce asexually, generating all cell types in adult stages. However, the putative adult stem cell populations involved in these processes, and the diversity of cell types generated by them, are still unknown. To address this, we recover 75,218 single cell transcriptomes of the highly regenerative and asexually-reproducing annelid Pristina leidyi. Our results uncover a rich cell type diversity including annelid specific types as well as novel types. Moreover, we characterise transcription factors and gene networks that are expressed specifically in these populations. Finally, we uncover a broadly abundant cluster of putative stem cells with a pluripotent signature. This population expresses well-known stem cell markers such as vasa, piwi and nanos homologues, but also shows heterogeneous expression of differentiated cell markers and their transcription factors. We find conserved expression of pluripotency regulators, including multiple chromatin remodelling and epigenetic factors, in piwi+ cells. Finally, lineage reconstruction analyses reveal computational differentiation trajectories from piwi+ cells to diverse adult types. Our data reveal the cell type diversity of adult annelids by single cell transcriptomics and suggest that a piwi+ cell population with a pluripotent stem cell signature is associated with adult cell type differentiation.

Rejuvenating aged stem cells: therapeutic strategies to extend health and lifespan.

Aging is associated with a global decline in stem cell function. To date, several strategies have been proposed to rejuvenate aged stem cells: most of these result in functional improvement of the tissue where the stem cells reside, but the impact on the lifespan of the whole organism has been less clearly established. Here, we review some of the most recent work dealing with interventions that improve the regenerative capacity of aged somatic stem cells in mammals and that might have important translational possibilities. Overall, we underscore that somatic stem cell rejuvenation represents a strategy to improve tissue homeostasis upon aging and present some recent approaches with the potential to affect health span and lifespan of the whole organism.

Islr regulates satellite cells asymmetric division through the SPARC/p-ERK1/2 signaling pathway.

Satellite cells (SCs) are adult muscle stem cells responsible for muscle regeneration after acute and chronic muscle injuries. The balance between stem cell self-renewal and differentiation determines the kinetics and efficiency of skeletal muscle regeneration. This study assessed the function of Islr in SC asymmetric division. The deletion of Islr reduced muscle regeneration in adult mice by decreasing the SC pool. Islr is pivotal for SC proliferation, and its deletion promoted the asymmetric division of SCs. A mechanistic search revealed that Islr bound to and degraded secreted protein acidic and rich in cysteine (SPARC), which activated p-ERK1/2 signaling required for asymmetric division. These findings demonstrate that Islr is a key regulator of SC division through the SPARC/p-ERK1/2 signaling pathway. These data provide a basis for treating myopathy.

Stem Cell and its Applications in Medical Biotechnology

Stem cells are undifferentiated cells in multicellular organisms that can be specialized into many types of cells and multiply into different types of cells. Stem cells can be produced from many sources in the body such as early embryos, embryonic tissue, foetal membranes i e; chorion and amnion, umbilical cord, and amniotic fluid. Stem cells are also present in organs of adult organisms like blood, skin, skeletal muscle, bone marrow, etc., Stem cells can be totipotent, pluripotent, multipotent, and unipotent. Stem cells are used to regenerate injured cells in the body, and repair the cells due to any kind of disease. From all types of cells, totipotent stem cells can generate any kind of cells in the body so they are the most important type of cells. Along with this, pluripotent stem cells also produce all types of cells, except the placenta or embryonic cells. These cells can be isolated from the body from the late embryonic stage after 1 week of the birth of a child, from the miscarriage of 12 weeks, and also from the umbilical cord of the child. These can be stored for another 2 decades. Another type is multipotent stem cells, which can generate a limited number of cells in a specific type; the last fourth type is unipotent stem cells, which can only produce cells of their own type. Diagnosis with stem cells is a useful technique to treat patients with many blood-related diseases for example blood cancer, thalassemia, etc., It also cures diseases like diabetes, liver disease, bone and cartilage, and organ transplantation.

Manipulating Stem Cell Fate with Disordered Bioactive Cues on Surfaces: The Role of Bioactive Ligand Selection.

The development of 2D or 3D bioactive platforms for rapidly isolating pure populations of cells from adult stem cells holds promise for advancing the understanding of cellular mechanisms, drug testing, and tissue engineering. Over the years, methods have emerged to synthesize bioactive micro- and nanostructured 2D materials capable of directing stem cell fate. We introduce a novel method for randomly micro- or nanopatterning any protein/peptide onto both 2D and 3D scaffolds via spray technology. Our goal is to investigate the impact of arranging bioactive micropatterns (ordered vs disordered) on surfaces to guide human mesenchymal stem cell (hMSC) differentiation. The spray technology efficiently coats materials with controlled, cost-effective bioactive micropatterns in various sizes and shapes. BMP-2 mimetic peptides were covalently grafted, individually or in combination with RGD peptides, onto activated polyethylene terephthalate (PET) surfaces through a spraying process, incorporating nano/microscale parameters like size, shape, and composition. The study explores different peptide distributions on surfaces and various peptide combinations. Four surfaces were homogeneously functionalized with these peptides (M1 to M4 with various densities of peptides), and six surfaces with disordered micro- and nanopatterns of peptides (S0 to S5 with different sizes of peptide patterns) were synthesized. Fluorescence microscopy assessed peptide distribution, followed by hMSC culture for 2 weeks, and evaluated osteogenic differentiation via immunocytochemistry and RT-qPCR for osteoblast and osteocyte markers. Cells on uniformly peptide-functionalized surfaces exhibited cuboidal forms, while those on surfaces with disordered patterns tended toward columnar or cuboidal shapes. Surfaces S4 and S5 showed dendrite-like formations resembling an osteocyte morphology. S5 showed significant overexpression of osteoblast (OPN) and osteocyte markers (E11, DMP1, and SOST) compared to control surfaces and other micropatterned surfaces. Notably, despite sharing an equivalent quantity of peptides with a homogeneous functionalized surface, S5 displayed a distinct distribution of peptides, resulting in enhanced osteogenic differentiation of hMSCs.

SOXC are critical regulators of adult bone mass

Pivotal in many ways for human health, the control of adult bone mass is governed by complex, incompletely understood crosstalk namely between mesenchymal stem cells, osteoblasts and osteoclasts. The SOX4, SOX11 and SOX12 (SOXC) transcription factors were previously shown to control many developmental processes, including skeletogenesis, and SOX4 was linked to osteoporosis, but how SOXC control adult bone mass remains unknown. Using SOXC loss- and gain-of-function mouse models, we show here that SOXC redundantly promote prepubertal cortical bone mass strengthening whereas only SOX4 mitigates adult trabecular bone mass accrual in early adulthood and subsequent maintenance. SOX4 favors bone resorption over formation by lowering osteoblastogenesis and increasing osteoclastogenesis. Single-cell transcriptomics reveals its prevalent expression in Lepr+ mesenchymal cells and ability to upregulate genes for prominent anti-osteoblastogenic and pro-osteoclastogenic factors, including interferon signaling-related chemokines, contributing to these adult stem cells’ secretome. SOXC, with SOX4 predominantly, are thus key regulators of adult bone mass.