Calcific aortic valve disease (CAVD) is a progressive and life-threatening condition characterized by fibrocalcific remodeling of the valve leaflets. Valvular interstitial cells (VICs) are central mediators of calcific aortic valve disease (CAVD), as their osteogenic trans-differentiation drives pathological matrix remodeling and calcium deposition within the valve leaflets. Human-induced pluripotent stem cell-derived valvular interstitial cells (hiVICs) represent a promising patient-specific platform for disease modeling. While primary VICs (pVICs) readily undergo mineralization under osteogenic stimulation, hiVICs fail to calcify in conventional two-dimensional (2D) cultures. Our data suggests that the inability of hiVICs to calcify in 2D culture is related to FOXO1 (Forkhead box protein O1) activity, which suppresses the osteogenic transcriptional program by inhibiting RUNX2 (Runt-related transcription factor 2). To address this limitation, we then developed a three-dimensional tissue ring construct using hiVICs. When cultured in osteogenic medium, these constructs exhibited robust calcification, as confirmed by Alizarin Red and Von Kossa staining. FOXO1 was also identified as a mediator of calcification in the tissue ring constructs. Metformin treatment restored FOXO1 expression and inhibited calcification, while AS1842856, a selective FOXO1 inhibitor, exacerbated tissue construct mineralization and led to a near-complete tissue collapse. In summary, we establish a functional 3D hiVIC-based model of CAVD enabling mechanistic investigation and pharmacological screening and identify FOXO1 as a critical regulator of osteogenic transition.

It is widely believed that epithelial cells in solid tissues undergo epithelial-mesenchymal transition (EMT) during carcinogenesis. EMT transforms polar and adherent epithelial cells in solid tumors into mesenchymal cells that get mobilized as circulating tumor cells (CTCs) and trigger metastasis. Isolating normal and neoplastic epithelial stem cells and their characterization remains challenging and vague even today. Most deaths in cancer patients are due to metastasis and hence a huge interest exists in understanding and developing tools to prevent and overcome metastasis. EMT during cancer remains clouded by controversies and questions persist as to its precise role. Besides a lack of histological evidence, lineage tracing studies have also failed to provide definitive proof supporting role of EMT in metastasis. Pluripotent, very small embryonic-like stem cells (VSELs) express sex hormone receptors and exist in a quiescent state in all tissues. They are responsible for regular turnover of epithelial cells, maintain lifelong homeostasis and their dysfunctions result in various pathologies including cancer. Developmental exposure to endocrine disrupting chemicals directly impacts VSELs, results in epigenetic changes that transform VSELs into cancer stem cells (CSCs). CSCs enter cell cycle, undergo excessive self-renewal and initiate cancer. CSCs (epigenetically altered and dysfunctional VSELs) are mobilized into circulation and are studied by our group for early prediction of cancer unlike CTCs, in a liquid biopsy, that fail to detect cancer in early stages. In this article, we discuss that besides initiation, CSCs also play a key role in cancer spread. Open questions surrounding EMT are reviewed and discussed in the context of VSELs biology. Existing hallmarks of metastasis-initiating cells produced by EMT are critically examined considering CSCs with a crucial role in cancer initiation, progression, metastasis and recurrence, challenging the existing focus on EMT and CTCs.

Hematopoiesis is a dynamic, adaptive process that governs blood and immune cell production through coordinated self-renewal, proliferation and differentiation. Shaped by intrinsic cell heterogeneity, environmental cues and physiological demands, it ensures effective blood cell production under both steady-state and stress conditions. Advances in single-cell and lineage-tracing technologies have shifted the traditional hierarchical view of hematopoiesis toward a flexible, interconnected network. B cell development exemplifies this plasticity, involving coordinated genetic and environmental signals to generate diverse subsets. Beyond the classical common lymphoid progenitors (CLP)-dependent model, B cells can also arise through alternative pathways, including direct differentiation from HSCs or at extramedullary sites like the spleen. Environmental signals and niche-specific factors support this diversity. Bipotent progenitors linking B lymphoid and myeloid (macrophage/osteoclast) fates have been identified in both fetal and adult hematopoiesis, revealing overlapping lineage potential and developmental flexibility. Moreover, mature B cells exhibit functional adaptability. B2 cells can convert into B1 cells under certain conditions, while CD11b⁺ myeloid-like B cells (M-B cells) emerge during emergency myelopoiesis, highlighting functional plasticity beyond antibody production. This evolving understanding redefines B cells as versatile immunoregulatory players, especially during inflammation and immune stress and opens new avenues for therapeutic interventions in immunity and hematologic disorders.

Cardiovascular disease (CVD) is a significant cause of cardiac and vascular-related deaths worldwide. While traditional drug and surgical treatments can alleviate symptoms and slow progression, they cannot regenerate heart tissue or reverse function. Heart transplantation, although a radical cure, is limited by donor availability, risks, and costs. Stem cell therapy has gained attention as a potential treatment option, but is hindered by low retention rates post-transplantation. Extracellular vesicles (EVs) are nanoscale membrane vesicles found in various cells and play a key role in the paracrine effects of stem cells. Despite being a promising treatment for cardiovascular diseases, the short plasma half-life and non-specific uptake by the liver and spleen significantly impact its therapeutic efficacy in the heart. This review examines the current understanding of extracellular vesicles and recent advancements in strategies to reduce EV loss and enhance targeted delivery for cardiovascular disease treatment. Approaches such as hydrogel incorporation, vesicular membrane modifications, fusion techniques, and inhibition of monocyte-macrophage system (MPS) clearance are discussed. The paper concludes by addressing the current status of extracellular vesicle therapy and provides insights into its future development.

Bone organoids mimic the structure and function of actual bone tissue and serve as novel tools for disease modeling, drug testing, and bone repair. However, their development is severely impeded by the limited availability of cell sources. Fortunately, human pluripotent stem cells (hPSCs) can differentiate into organoid constituent cells, including osteoblasts, osteoclasts, and endothelial cells. However, their differentiation efficiencies are relatively low and do not meet the requirements of clinical applications because of the use of undefined culture components such as fetal bovine serum. More importantly, nearly all the existing scaffolds cannot support the culture of hPSCs. Thus, much effort should be made to construct bone organoids using cells induced from hPSCs. This review starts with the in vivo development of bone tissue. We summarize the mechanisms, methods, and purification processes for differentiating hPSCs into the above cell types in bone organoids. On this basis, we described strategies related to hPSC-based bone organoids and the growth factors and bioactive materials needed to accelerate this process. Finally, we extensively discuss the existing challenges and prospects. This review is valuable for the future development and clinical application of hPSC-derived bone organoids.

Bone regeneration is a dynamic process regulated by the interplay between the immune and skeletal systems. Regulatory T cells (Treg), a specialized subset of CD4 T cells, play a crucial role in immunomodulation and bone regeneration by regulating the immune response and interacting with progenitor cells. However, the specific mechanisms through which Treg influence the osteogenic differentiation of bone marrow stromal cells (BMSC) remain unexplored. Treg were isolated from six healthy donors, expanded for 13 days, and starved for 24 h to collect Treg-conditioned media (Treg-CM). BMSC obtained from three different healthy donors were treated with Treg-CM at an optimized concentration (50 µg/mL) to assess its impact on BMSC metabolic activity, migration, and osteogenic differentiation. Label free proteomics and cytokine profiling were conducted to identify unique proteins and immunomodulatory factors in Treg-CM. The secretory cytokines of BMSC treated with Treg-CM were also analyzed. Treg-CM enhances BMSC osteogenic differentiation by upregulating the expression of key osteoblast-specific genes, increasing ALP activity, and facilitating calcium deposition. Proteomics identified unique proteins in Treg-CM that regulate cytoskeletal dynamics, metabolic processes and mRNA regulation, highlighting a complex mechanism underlying Treg-CM effects. Cytokine profiling provided key immune modulators in Treg-CM that regulate osteogenesis. Furthermore, elevated levels of MIP-1α and G-CSF were secreted by BMSC treated with Treg-CM further supporting its role in immune-mediated osteogenesis. Our findings reveal that Treg-CM enhances not only osteogenesis in vitro but also fosters a pro-regenerative microenvironment. This highlights its potential as a cell-free strategy for enhancing stem-cell based osteogenesis.

The integration of CRISPR-based functional genomics with pluripotent stem cell (PSC) technologies has been recognized as a transformative approach for investigating gene function, modeling human disease, and advancing regenerative medicine. The aim of this review is to provide a comprehensive evaluation of recent developments in CRISPR-Cas platforms, including gene knockouts, base and prime editing, and CRISPR activation or interference (CRISPRa/i), as applied to PSC systems. Studies employing human PSCs, including embryonic stem cells and induced pluripotent stem cells, have been examined to summarize methodologies for genome-wide screening, lineage tracing, and therapeutic engineering. Advances in editing efficiency, delivery strategies, and genomic safety have been reported, while limitations persist in the form of off-target modifications, epigenetic variability, and cell-type-specific responses. Notable applications include the generation of immune-evasive PSC lines, the development of organoid models for physiological and pathological studies, and the implementation of phenotypic screening for disease-relevant traits. Collectively, these technological and methodological advancements have established functional genomics of PSC-CRISPRSPR as a versatile and powerful framework for elucidating fundamental aspects of human biology, dissecting complex traits, and accelerating the translation of discoveries from experimental research to clinical implementation.

Allogeneic hematopoietic stem cell transplantation (HSCT) for pediatric acute leukemia is limited by non-relapse mortality (NRM) and relapse. This study evaluated whether the endothelial activation and stress index (EASIX) score-calculated with the formula [lactate dehydrogenase (LDH; U/L) × serum creatinine (mg/dL)]/platelets (10/L)]-could be associated with NRM and overall survival (OS). We analyzed 195 patients (< 25 years) with acute lymphoblastic leukemia (ALL) or acute myeloid leukemia (AML) who underwent first-time peripheral blood HSCT at a single center between 2014 and 2022. The EASIX score was assessed pre-transplant (EASIX.PRE) and on day + 7 post-transplant (EASIX.POST). Cutoff values for EASIX.PRE (0.836) and EASIX.POST (1.632) were established using receiver operating characteristic (ROC) curves. Patients with EASIX.PRE scores below the cutoff exhibited significantly improved 12-month OS (84.67% vs. 66.12%, P = 0.028). Multivariable analysis confirmed that an EASIX.PRE score above the cutoff was an independent prognostic factor for inferior OS (HR = 1.83, P = 0.039). Similarly, patients with an EASIX.POST score below the cutoff showed a higher 12-month OS rate (87.78% vs. 72.65%, P = 0.046). However, in multivariable analysis, an EASIX.POST score above the cutoff did not demonstrate a significant relationship with reduced OS. Neither EASIX.PRE nor EASIX.POST score was independently associated with NRM, relapse, graft-versus-host disease (GvHD), leukemia-free survival (LFS), or GvHD-free, relapse-free survival (GRFS). This study highlights the prognostic utility of EASIX.PRE for OS in pediatric HSCT recipients but underscores its limited role in predicting NRM or GvHD. Further studies with larger cohorts and dynamic EASIX assessments are required to confirm these findings and refine risk stratification in pediatric HSCT.

Diabetes mellitus (DM) is characterized by hyperglycemia, leading to various systemic complications. Stem cell based regenerative applications hold revolutionary potential for treating various chronic disorders, including diabetes and its associated co-morbidities. This review highlights the regenerative potential of mesenchymal stem cells (MSCs) for diabetes induced systemic manifestations i.e., damage to pancreatic beta-cells, skin, neural, retinal, and renal tissues. Persistent hyperglycemic condition in DM causes mitochondria to produce reactive oxygen species (ROS) which further activates inflammatory processes. Pro-inflammatory mediators (TNF-α, IL-1, IL-6, and C-reactive protein) lead to metabolic inflammation and damage pancreatic β-cells, blood brain barrier (BBB), synaptic integrity contributing to neurodegenerative effects, impaired glomerulus filtration rate (GFR), and blood renal barrier (BRB). MSCs evidently dictate their potential to reduce inflammation, differentiation into specific cell types, and augment tissue repair and regeneration. A number of mechanisms have been proposed by which MSCs exert their effect to improve these complications. MSCs augment β-cell function by mitigating endoplasmic reticulum stress and even translocating healthy mitochondria to injured cells. MSCs improve oxidative stress and mitochondrial dysfcunction, key processes of retinal and nerve damage. MSCs also reduce fibrosis, revive glomerular function, enhance vascular stability, promote angiogenesis and wound healing. MSC secretome, rich in bioactive metabolites, also provides retinal- and neuronal protection. MSC-based therapies have emerged as a promising hope for affected individuals. Regardless of their advantages, challenges still endure which include selection of MSC source, scalability, systemic and long-term safety, therefore, extended preclinical and clinical research is needed to standardize the treatment.

Mesenchymal stromal cells (MSCs) have demonstrated therapeutic potential in hematologic diseases by modulating immune responses, supporting hematopoiesis, and remodeling the bone marrow microenvironment. Clinically, MSCs have been explored for graft-versus-host disease and hematopoietic stem cell transplantation support, while their applications in hematologic malignancies, including acute myeloid leukemia, multiple myeloma, and myelodysplastic syndromes, remain under investigation. However, therapeutic heterogeneity, safety concerns, and standardization challenges limit their clinical translation. Recent advances in MSC-derived extracellular vesicles, gene modification technologies, and integrative combination strategies have expanded the therapeutic landscape, enabling more precise and targeted modulation of immune responses and tumor microenvironments. Moreover, disease-specific evidence highlights the dual roles of MSCs-acting either as therapeutic agents or as contributors to disease progression-depending on stromal plasticity and niche conditioning. This review provides a comprehensive and mechanistic synthesis of MSC functions across both malignant and non-malignant hematologic disorders, integrating preclinical and clinical findings in immunoregulation, hematopoietic recovery, anti-fibrosis, and microenvironmental reprogramming. In addition, we critically evaluate emerging strategies to overcome translational bottlenecks, including inter-donor variability, lack of predictive potency markers, and the need for scalable, standardized manufacturing protocols. By bridging foundational mechanisms with translational potential, this review offers forward-looking perspectives to guide future optimization and clinical integration of MSC-based therapies in hematology.

Tissue engineering seeks to develop biomimetic substitutes for damaged tissues using natural or synthetic materials functionalized with cells or biologically active compounds. Understanding how cell-scaffold interactions influence cellular behavior is critical for optimizing tissue engineering strategies. Polylactic acid (PLA), a biodegradable and biocompatible polymer approved by the U.S. FDA, is commonly used for scaffolds fabrication. Mesenchymal stromal cells (MSCs) are widely utilized in regenerative medicine due to their capacity to differentiate into mesodermal lineages and their secretomes, which exhibit robust paracrine activity. To date, few studies have investigated how scaffold characteristics, such as materials and/or architecture, modulate secretome composition using integrated multi-omics approaches. An untargeted metabolomics workflow combined with label-free proteomics was employed to analyze ASC-derived secretomes, obtained from adipose tissue-derived MSCs (ASCs) cultured in PLA-discs and PLA-scaffolds, as well as on conventional 2D polystyrene (PS) culture surfaces. A similar proteomic distribution was observed when secretomes from PLA-discs and PLA-scaffolds were compared. On the other hand, significant changes in the proteomic profile were observed between PLA-discs and 2D-PS secretomes. Proteins belonging to carbohydrate metabolism, cell motility, vasculature development, and oxidative stress response, among others, were increased in PLA-discs. The metabolomic profile shows significant differences, with metabolites related to glucose metabolism, such as pyruvate and lactic acid, increased in PLA secretomes. Our multi-omic approach demonstrates that the PLA constructs introduced here can modulate the secretome of ASCs, inducing significant changes in its composition, highlighting the influence of culture format on the secretory capacity of ASCs.

The persistence and drug resistance of leukemic stem cells (LSCs) are major challenges in the treatment of acute myeloid leukemia (AML). Ferroptosis, a novel form of programmed cell death, has emerged as a promising strategy for eradicating LSCs. This review provides a systematic analysis of LSC ferroptosis resistance and explores the interplay between iron metabolism, lipid peroxidation, and antioxidant defense mechanisms. We propose a novel predictive model based on single-cell multiomics data that integrates iron homeostasis regulators (TfR1, GPX4, and FTH1) to assess the susceptibility of LSCs to ferroptosis. A key innovation of this study was the in-depth exploration of LSC ferroptotic heterogeneity and its interaction with the tumor microenvironment, shedding light on new approaches for precision AML therapy. Based on these findings, we introduced an innovative treatment paradigm combining ferroptosis inducers (e.g., erastin, RSL3) with immunotherapies (such as PD-L1 inhibitors and CAR-T cell therapy) to enhance LSC clearance and minimize measurable residual disease (MRD). This review fills a critical knowledge gap in the study of ferroptosis in LSCs, providing a theoretical foundation and translational insights for future AML treatment strategies.

Hematopoietic stem cell transplantation has been conducted in clinical settings to treat patients with malignant or non-malignant blood diseases for decades. Cord blood (CB) has been recognized as an essential graft source with beneficial characteristics, such as a lower risk of relapse and a lower rate of chronic graft-versus-host disease. However, the limited number of cells in CB impedes its broader use and hinders the ability to harness its benefits. Various expansion strategies have emerged to address this barrier, based on a deeper understanding of fate decisions and the maintenance of stemness in hematopoietic stem cells. To achieve an efficient transition from the laboratory to clinical application, several strategies have successfully managed scale-up manufacturing to satisfy clinically relevant requirements for both quality and scale. These approaches have progressed to the clinical stage and have demonstrated promising results. Novel expanded CB-derived hematopoietic stem and progenitor cells (HSPCs) therapies, including OMISIRGE (Omidubicel onlv.), Zemcelpro (Dorocubicel), and upcoming products with International Nonproprietary Name designations, introduce innovative concepts and comprehensive considerations for improving CB transplantation. This progress enables novel therapeutic options and represents a breakthrough in traditional CB transplants. In this context, we summarize and explore representative techniques and products to provide insights that inspire future developments in CB-derived HSPC therapies.

The liver is a highly versatile and resilient organ that is crucial for metabolism, detoxification, digestion, and immune regulation. Its remarkable regenerative capacity is driven primarily by two key cellular processes: hepatocyte polyploidy and cellular senescence. This review explores the complex roles of polyploidy, in which hepatocytes possess multiple chromosome sets, and senescence, characterized by irreversible cell cycle arrest, in maintaining liver homeostasis and facilitating regeneration. Polyploid hepatocytes increase genetic and metabolic diversity, enabling the liver to withstand stress and recover from injury through mechanisms such as compensatory regeneration, depolyploidization, and the fusion of extrinsic stem cells. Concurrently, cellular senescence acts as a protective barrier against uncontrolled cell proliferation and genomic instability while also promoting tissue repair via the senescence-associated secretory phenotype (SASP). The interplay between polyploidy and senescence is regulated by critical molecular pathways, including the Hippo, PI3K/Akt, and p53 signaling pathways, which balance cell proliferation, differentiation, and apoptosis. Additionally, this review discusses the therapeutic potential of targeting these processes to increase liver regeneration, prevent fibrosis, and reduce the risk of hepatocellular carcinoma (HCC). Emerging strategies such as senolytic drugs, stem cell therapies, and cytokine modulation offer promising avenues for treating chronic liver diseases. However, challenges remain in fully understanding the functional distinctions between diploid and polyploid hepatocytes and managing the dual roles of senescence. Future research should focus on molecular insights and targeted interventions to optimize liver health and regenerative outcomes.