• Bulge progenitors’ cells (BPC) are multipotent cells that fuel hair follicle cycling, sebaceous turnover, and epidermal repair. • BPC activation from quiescence is governed by niche-derived Wnt/β-catenin (anagen entry), BMP antagonism (licensing transit), and TGF-β/Smad (catagen enforcement). • Preclinical models demonstrate de novo folliculogenesis using BPC- dermal papilla cell recombinants in 3D scaffolds, with > 70
B-cell maturation antigen (BCMA)-directed chimeric antigen receptor (CAR) T-cell therapy can induce deep responses in relapsed or refractory multiple myeloma, yet relapse remains frequent. Neither post-infusion maintenance nor salvage after CAR T-cell failure has an established standard. This review synthesizes pivotal trials, real-world cohorts, and translational studies into a risk-adapted framework for longitudinal care. Relapse biology, target modulation, CAR T-cell persistence, immune reconstitution, measurable residual disease (MRD), and imaging are integrated to guide structured observation, time-limited maintenance, pre-emptive intervention, and salvage. Preventive treatment is defensible only when the estimated risk posed by residual disease exceeds the competing risks of infection, cytopenia, and delayed immune recovery. At relapse, treatment selection should begin with clinical tempo, disease compartment, contemporary expression of BCMA, G protein-coupled receptor class C group 5 member D (GPRC5D), and Fc receptor-homolog 5 (FcRH5), prior target pressure, and host immune fitness. Structured observation is appropriate for many patients with sustained MRD negativity, no extramedullary disease, and recovering immune function. Persistent or rising MRD, residual lesions, aggressive disease biology, or unfavorable CAR T-cell kinetics should prompt early trial referral rather than automatic chronic therapy. Salvage may involve BCMA retargeting, a switch to GPRC5D or FcRH5, conventional cytoreduction, radiotherapy, antibody-drug conjugates, bispecific antibodies, or a second cellular platform. Future trials should assess progression-free survival together with infection-free survival, treatment-free interval, immune recovery, quality of life, and responsiveness to subsequent salvage.
Precise genome editing of induced pluripotent stem cells (iPSCs) using clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9 (CRISPR-Cas9) has opened unprecedented avenues for advancements in regenerative medicine and disease modelling. However, the establishment of isogenic single-cell-derived iPSC populations, particularly upon CRISPR-Cas9 gene editing modifications, is one of the major challenges still associated with these advanced methodologies relying on low-rate editing events and requiring defined clonogenicity. In response, we have developed a systematic, comprehensive and efficient workflow combining generation, genotyping and expansion of high-quality monoclonal iPSC lines following CRISPR-Cas9 genome editing. In particular, the protocol incorporates optimized single-cell cloning procedures for two commercially available dispensing platforms, one based on microfluidic imaging and the other on impedance technology, together with rapid droplet digital PCR (ddPCR)-based screening of non-homologous end joining (NHEJ) and homology-directed repair (HDR) outcomes. By combining gentle single-cell handling with advanced genotyping methodologies, the protocol enables efficient early assessment of editing outcomes before commitment to labour-intensive clonal derivation, thereby accelerating project timelines, minimising cell stress and loss, preserving genetic fidelity and supporting scalability. Coupled with precisely defined culture conditions tailored for post-seeding recovery, these approaches aim to improve iPSC viability and clonal outgrowth, achieving at least 60
m5C (5-methylcytosine) modification is a significant epigenetic modification in RNA that has garnered substantial attention in recent years in stem cell biology and disease research. The dynamic regulation of m5C involves methyltransferases, demethylases, and specific binding proteins, which collectively influence RNA stability, translation efficiency, and cellular function. Especially in stem cell differentiation, m5C modification not only affects stem cell self-renewal and differentiation fate but is also closely associated with the onset and progression of various diseases. This review explores the mechanisms of m5C modification in stem cell differentiation and related diseases, highlighting its emerging roles in gene expression regulation and disease pathogenesis. Not applicable.
Diabetic retinopathy (DR) is a leading cause of vision loss and is now increasingly understood as a neurovascular disease rather than a purely microvascular complication of diabetes. The retinal neurovascular unit (NVU), composed of neurons, Müller cells, microglia, astrocytes, endothelial cells, pericytes, and extracellular matrix components, integrates neural activity, vascular perfusion, barrier integrity, and inflammatory control. In diabetes, NVU injury develops through a hierarchical and self-amplifying process. Upstream metabolic and systemic triggers, including hyperglycemia, advanced glycation end products (AGEs), homocysteine and lipid imbalance, and NADPH oxidase-derived reactive oxygen species, initiate cellular stress. These triggers converge on mitochondrial dysfunction, impaired autophagy and mitophagy, endoplasmic reticulum stress, and mitochondrial DNA release, which subsequently activate innate immune pathways. Neuroinflammatory amplification driven by microglial activation, Müller cell gliosis, NLRP3 inflammasome signaling, pyroptosis, and cytokine feedback further links neuronal and vascular injury. The final common pathway is integrated NVU breakdown, characterized by retinal neurodegeneration, pericyte loss, endothelial dysfunction, blood-retinal barrier disruption, vascular leakage, and VEGF-driven neovascularization. Regulatory RNA networks, including lncRNAs, miRNAs, circRNAs, piRNAs, and tRNA-derived fragments, act as cross-cutting epigenetic regulators of these processes. This review integrates current evidence within an NVU-centered systems biology framework and aligns therapeutic strategies with the disease hierarchy, distinguishing established clinical treatments from preclinical and emerging interventions.
Bladder dysfunction comprises diverse lower urinary tract disorders lacking treatments that restore tissue integrity and neural control. While mesenchymal stem cells (MSCs) show promise in regenerative urology, their benefits primarily stem from paracrine mechanisms. Consequently, MSC-derived extracellular vesicles (MSC-EVs) have emerged as superior cell-free alternatives, recapitulating the parent cells’ regenerative potential with enhanced biosafety, stability, and controllability.This review summarizes advances in MSC- and MSC-EV-based therapies for bladder conditions, including neurogenic, diabetic, overactive/underactive bladder, outlet obstruction, and interstitial cystitis. Mechanistically, these therapies exert multi-target effects: modulating immunity, suppressing fibrosis (via TGF-β inhibition), attenuating inflammation (via NF-κB/NLRP3), promoting angiogenesis, and enhancing neuroregeneration. MSC-EVs drive these processes by precisely transferring bioactive microRNAs and proteins.We propose a paradigm shift from conventional cell therapies to MSC-EV-centric, cell-free strategies, which overcome traditional transplantation limitations like low engraftment and immunogenicity. While early data confirm feasibility and safety, challenges in standardizing EV isolation, dosing, and long-term efficacy remain. Ultimately, this review establishes MSC-EVs as a next-generation therapeutic framework, guiding future clinical translation in regenerative urology.
Osteoporotic fracture is the most severe complication of osteoporosis, and impaired bone regeneration and delayed healing after fracture remain major clinical challenges. Extracellular vesicles (EVs) derived from bone marrow mesenchymal stem cells (BMSCs) may play a critical role in promoting bone regeneration after osteoporotic fracture. Enhancing their osteogenic potential and elucidating the underlying mechanisms may contribute to accelerated fracture healing. In this study, resveratrol-stimulated BMSC-derived EVs (Res-EVs) were isolated using differential centrifugation combined with ultracentrifugation. We found that compared with EVs secreted by BMSCs under normal culture conditions (Nor-EVs), Res-EVs significantly enhanced BMSCs osteogenic differentiation capacity. Analysis of miRNA sequencing data identified miR-19a-3p as significantly downregulated and PC-3p-83,608 as significantly upregulated. The reliability of the sequencing data was validated by qPCR results. By transfecting BMSCs with inhibitor and mimic of miR-19a-3p, we found that miR-19a-3p was a negative regulator of osteogenic differentiation. Bioinformatics prediction and dual-luciferase reporter assays confirmed that miR-19a-3p directly targets Raf1. Western blot results indicated that miR-19a-3p regulates osteogenic differentiation of BMSCs by targeting Raf1. In vivo experiments further demonstrated that Res-EVs treatment enhanced repair at osteoporotic fracture sites and increased the expression of RUNX2 and RAF1. Collectively, these findings indicate that Res-EVs promote bone regeneration at osteoporotic bone defect sites in rats, with the miR-19a-3p/Raf1 axis representing a potential underlying mechanism.
Diabetic foot ulcer (DFU) is a clinically challenging complication characterized by poor healing outcomes, and conventional therapies provide limited benefit. Mesenchymal stem cell (MSC) transplantation offers a promising strategy for DFU repair. However, the low survival of transplanted MSCs in the hostile wound microenvironment, coupled with the lack of real-time, non-invasive methods to track these cells in vivo, severely hampers their therapeutic efficacy and clinical translation. We engineered MSCs to co-express a dual reporter system comprising near-infrared fluorescent protein (iRFP) and ferritin heavy chain (FTH1). These modified cells were then integrated with a fibrin glue (FG) scaffold to create a unified platform that supports both multimodal imaging and therapeutic function within skin wounds. First, FTH1 overexpression enhances the antioxidant capacity of MSCs, while the FG scaffold provides structural support; this combination enhances cell survival and retention. Second, the iRFP/FTH1 dual reporter enables near-infrared fluorescence imaging and MRI-based localization, establishing a multimodal platform for real-time cell tracking. In a full-thickness skin defect model in diabetic mice, multimodal imaging revealed that transplanted cells persisted in the wound area for approximately seven days. Treatment with iRFP/FTH1-MSCs/FG significantly accelerated wound closure and promoted hair follicle regeneration and angiogenesis. Additionally, local iron deposition resulting from FTH1 expression enhanced fibroblast migration and collagen synthesis, further facilitating extracellular matrix remodeling. Mechanistic studies demonstrated that this therapy drives macrophage polarization toward the anti-inflammatory M2 phenotype and activates the PI3K–AKT–VEGF signaling pathway. These complementary effects synergistically enhance tissue regeneration and systematically improve diabetic wound healing. Collectively, this multimodal stem cell–scaffold system effectively integrates dynamic cell tracking with stem cell therapy during skin wound repair. It addresses a critical technical gap in visualizing stem cells within the wound microenvironment and provides valuable methodological and theoretical foundations for optimizing regenerative strategies for diabetic skin wounds.
Hematopoietic stem cell (HSCs) aging is a complex biological process driven by both cell-intrinsic alterations and extrinsic cues from the bone marrow niche. Understanding these mechanisms is critical for developing therapies against aging-related hematopoietic disorders. This review synthesizes recent advances in the molecular mechanisms underlying HSCs aging, including microenvironmental aging, genomic instability, epigenetic dysregulation, mitochondrial dysfunction, and aberrant nuclear mechanotransduction. We summarize that the functional decline of HSCs during aging drives a compensatory expansion of the phenotypically defined stem cell pool, leading to an aberrant increase in cell number. We also highlight aging-associated HSCs heterogeneity, including CD150high and P-selectin-positive subsets that enrich for myeloid-biased or functionally compromised HSCs states while emphasizing that surface phenotype alone may not fully indicate functional rejuvenation. Finally, we discuss emerging rejuvenation strategies-including targeting myeloid-biased HSCs, modulating inflammatory pathways, and implementing epigenetic or metabolic interventions-supported by cutting-edge technologies such as single-cell multi-omics, gene editing, and computational modeling. These approaches hold promise for counteracting age-related hematopoietic decline and restoring immune competence.
Umbilical cord-derived mesenchymal stromal cells (UC-MSCs) offer distinct advantages for clinical translation, including accessibility, scalability, and broad immunomodulatory capacity. However, the efficacy of systemically delivered UC-MSCs is constrained by suboptimal in vivo trafficking. Intravascular administration is limited by pulmonary first-pass sequestration, inefficient endothelial recruitment, blood-mediated inflammatory injury, and poor retention. While UC-MSCs homing is often conceptualized through a leukocyte adhesion paradigm, this model incompletely describes culture-expanded UC-MSCs, which exhibit heterogeneous expression of chemokine receptors, adhesion molecules, and selectin ligands. Furthermore, biodistribution and safety are critically determined by biophysical and hemocompatibility parameters, including cell size, deformability, cryopreservation status, and tissue factor (TF/CD142)-dependent procoagulant activity. This review synthesizes current understanding of UC-MSCs trafficking at the translational interface of biology and manufacturing. We examine canonical migratory mechanisms-chemokine signaling, integrin-mediated adhesion, extracellular matrix remodeling, and intracellular motility pathways-alongside underappreciated determinants of therapeutic performance: instant blood-mediated inflammatory reaction (IBMIR), complement-coagulation crosstalk, post-thaw functional impairment, donor variability, and route-dependent biodistribution. We also address the paradox wherein therapeutic benefit occurs despite minimal durable engraftment, implicating paracrine signaling, extracellular vesicles, and apoptosis-associated immune reprogramming as primary effectors. Finally, we evaluate strategies to enhance delivery and efficacy, including preconditioning, glycoengineering, receptor overexpression, route optimization, biomaterial-assisted retention, and migration-relevant potency assays under Good Manufacturing Practice (GMP). Advancing UC-MSCs therapy toward reproducible, mechanism-guided clinical application requires rigorous integration of hemocompatibility assessment, product characterization, and clinically informative cell tracking.
