npj Regenerative Medicine最新文献

Chronic diabetic wounds represent a major clinical challenge, compounded by persistent inflammation, microbial invasion, and deficient angiogenesis. To address these intertwined pathophysiological features, we developed a copper-ion coordinated andrographolide-loaded hydrogel (ASFH), significantly enhancing andrographolide solubility and promoting wound healing dynamics. In vitro assessments demonstrated superior antimicrobial activity, optimal mechanical strength, self-healing ability, and cytocompatibility. In diabetic mice, ASFH notably accelerated wound closure, stimulated collagen maturation and re-epithelialization, dynamically shifted macrophages toward an anti-inflammatory phenotype, and markedly enhanced angiogenesis. Mechanistic studies integrating network pharmacology, molecular docking, dynamics simulations, and SPR validation pinpointed the Rac1/JNK1/Jun/Fos signaling cascade as a primary mediator of these regenerative effects. This work presents ASFH as a translationally relevant dressing system, simultaneously addressing critical limitations in diabetic wound management through targeted molecular therapeutic intervention.

Peripheral artery disease (PAD) causes progressive arterial narrowing in the lower limbs and can advance to critical limb ischemia (CLI). Limited revascularization options highlight the need for safer, more effective therapies. Vascular multipotent stem cells (VMSCs) and adipose-derived stem cells (ADSCs) were isolated from adipose tissue, characterized phenotypically, and tested for angiogenic activity in vitro. Their therapeutic efficacy was then examined in a murine critical limb ischemia model through intramuscular transplantation, assessing limb preservation, neovascularization, and cell integration. VMSCs shared mesenchymal stem cell-like features with ADSCs and exhibited robust proliferative capacity, enabling rapid expansion to clinically relevant numbers. VMSCs also demonstrated endothelial-like properties, including CD31, VE-cadherin, and CD141 expression, and formed capillary-like structures in vitro. In contrast, ADSCs displayed perivascular characteristics with α-SMA and Transgelin expression. Co-culture of VMSCs and ADSCs promoted the development of mature tubular networks in vitro. Combined cell transplantation markedly decreased limb loss and promoted both angiogenesis and arteriogenesis in ischemic tissue, with transplanted cells partially integrating into the host vasculature to form hybrid vascular structures. VMSCs and ADSCs show complementary regenerative functions, sustained engraftment, and support for large-vessel formation, underscoring their potential for stem cell-based vascular therapies.

Childhood interstitial lung disease (chILD) secondary to pulmonary surfactant deficiency is a devastating chronic lung disease in children. Clinical presentation includes mild to severe respiratory failure and fibrosis. There is no specific treatment, except lung transplantation, which is hampered by a severe shortage of donor organs, especially for young patients. Repair of lungs with chILD represents a longstanding therapeutic challenge but cell therapy is a promising strategy. As surfactant is produced by alveolar epithelial type II (ATII) cells, engraftment with normal or gene-corrected ATII cells might provide an avenue to cure. Here, we used a chILD disease-like model, Sftpc^-/- mice, to provide proof-of-principle for this approach. Sftpc^-/- mice developed chronic interstitial lung disease with age and were hypersensitive to bleomycin. We could engraft wild-type ATII cells after low dose bleomycin conditioning. Transplanted ATII cells produced mature SPC and attenuated bleomycin-induced lung injury up to two months post-transplant. This study demonstrates that partial replacement of mutant ATII cells can promote lung repair in a mouse model of chILD-like disease.

Annually, 10% of 15 million bone fractures in the US fail to heal, and fractures with compromised blood flow, i.e., ischemia, are five times more likely to become nonunions. While ischemia is known to impair healing, the cellular and molecular mechanisms underlying this deficiency are unclear. Wild-type mice with surgically-induced ischemia underwent tibia fractures, and single-cell RNA-sequencing was performed on calluses at days 4 and 7 post-fracture. We observed delayed chondrogenic differentiation and upregulation of Cyclin-Dependent Kinase 8 (Cdk8) by stromal progenitors and fibroblasts in the ischemic callus. Hypoxia induced CDK8 gene expression in human mesenchymal stromal cells (hMSC), and pharmacological CDK8 inhibition promoted hMSC chondrogenic and osteogenic potential. In vivo oral delivery of a CDK8 inhibitor enhanced callus chondrogenesis and mineralization, potentially improving ischemic fracture healing. Our results suggest that CDK8 impedes stromal cell differentiation and that its inhibition may be a clinically translatable approach to enhance ischemic fracture healing.

Neurogenic bladder (NB) is a disabling condition lacking effective therapies. This study investigated whether CD73-expressing adipose-derived stem cells (ADSCs) promote bladder repair in a rat model of NB and explored the underlying mechanisms. ADSCs were sorted into CD73⁺ and CD73⁻ populations, and CD73⁺ cells were further modified to generate CD73⁺^/ev ADSCs and CD73-overexpressing CD73⁺^/⁺ADSCs, while CD73 inhibition was achieved using APCP. Conditioned media were applied to rat bladder smooth muscle cells in vitro, and ADSCs were injected into the bladder wall of rats subjected to bilateral pelvic nerve crush. Four weeks after treatment, bladder function, histology, and molecular markers were evaluated. CD73 overexpression enhanced VEGF and SDF-1 expression, promoted cell proliferation, and reduced inflammatory cytokines, whereas APCP suppressed VEGF. In vivo, CD73⁺^/⁺ADSCs improved cystometric parameters, regenerated bladder tissue, reduced pyroptosis, and activated the PI3K/AKT/mTOR pathway, while suppressing NF-κB/NLRP3/caspase-1 signaling. CD73 expression and VEGF progressively declined in untreated NB rats but were restored by CD73⁺^/⁺ADSCs. These findings indicate that CD73 enhances ADSC-mediated bladder repair through dual pro-regenerative and anti-inflammatory actions, suggesting a promising therapeutic strategy for NB.

The repair of corneal injuries remains a major challenge in clinical practice. Impaired corneal wound healing is closely associated with aberrantly activated stromal keratocytes and disorganized extracellular matrix. Here, we identify aberrant Hedgehog signaling in corneal keratocytes as a key driver of defective wound repair. In adult mice, Hedgehog signaling is suppressed in quiescent keratocytes but is pathologically reactivated following chemical injury, correlating with impaired repair. Keratocyte-specific Hedgehog activation via Ptch1 ablation disrupted corneal wound healing after epithelial scraping-a process that would normally resolve seamlessly under physiological conditions. Mechanistically, Hedgehog activation induced stromal thinning and stiffening through disorganized collagen fibrils. Transcriptomics analysis revealed keratocyte transdifferentiation into fibroblast-like phenotypes, accompanied by downregulation of extracellular matrix genes. Hedgehog-mediated stromal stiffening suppressed YAP activity in the overlying epithelium via Hippo pathway activation, blocking epithelial differentiation-a defect that was reversed by Hippo inhibition (XMU-MP-1). In chemical injury models, genetic Smo deletion or pharmacological Gli1/2 inhibition (GANT61) restored stromal architecture, normalized collagen organization, and rescued epithelial differentiation defects. These findings establish Hedgehog signaling in keratocytes as a critical regulator of stromal-epithelial crosstalk and highlight its targeted inhibition as a potential therapeutic strategy to restore corneal transparency and repair fidelity after injury.

Children with congenital heart defects increasingly survive to adulthood, but the non-physiological Fontan circulation imposed by current surgical palliation leads to significant sequelae and reduced lifespan. Restoring subpulmonic pumping function remains a long-standing goal, and there have been several attempts using regenerative medicine approaches. These efforts have lacked biomechanical rigor, however, and have not achieved the requisite functionality. Here, we introduce an analytically based framework that grounds pulsatile conduit design in biomechanical principles, coupling the architecture and properties of a passive matrix with embedded myofibers to optimize performance within pediatric anatomical constraints. Parametric exploration of matrix properties and myofiber orientations yields biomechanically feasible designs. Sensitivity analyses demonstrate design robustness and highlight parameters critical for reproducible biomanufacturing and surgical implementation. To illustrate clinical potential, a patient-specific lumped-parameter hemodynamic model shows that an optimized pulsatile conduit can generate physiologically meaningful pressures and flows and outperform passive grafts.

Muscle function and regeneration are impaired in type 1 diabetes, but whether this arises directly from muscle stem cell (MuSC) dysfunction has not been addressed. Here, we utilized three-dimensional MuSC cultures (micromuscles) to demonstrate that hyperglycemia drives deficits in muscle stem cell function, leading to impaired force production in differentiated myotubes. The functional capacity of skeletal muscle was shown to decline after repeated bouts of injury in mouse models of type 1 diabetes, and this was replicated in micromuscles derived from MuSCs isolated from diabetic mice, indicating MuSC dysfunction was linked to poor muscle regeneration and function. The loss of force producing capacity was associated with impaired myotube hypertrophy in vitro and in vivo after injury. Furthermore, poor muscle regeneration was exacerbated by a loss of MuSC number due to aberrant activation, even in the absence of injury. Deficits in MuSC function and number could be rescued by early treatment with the glucose-lowering drug dapagliflozin, indicating that MuSC defects were driven by exposure to a hyperglycemic environment. The findings reveal that MuSC dysfunction contributes to muscle functional deficits in models of type 1 diabetes.

Trans-sutural distraction osteogenesis (TSDO) is an effective treatment of midfacial hypoplasia, a craniofacial deformity frequently associated with cleft lip and palate. Though extracellular matrix (ECM) remodeling plays a pivotal role in craniofacial correction, the characteristics and mechanisms underlying collagen reorganization and cellular morphological adaptations during TSDO remain poorly understood. This study quantitatively delineates the spatiotemporal changes of sutural cells and ECM morphology, revealing a polarized alignment parallel to the direction of mechanical force. Multi-omics analysis demonstrates that macrophages regulate collagen remodeling in suture mesenchymal stem cells (SuSCs) via the PDGF signaling pathway. Subsequent in vitro stretch loading models confirmed PDGF pathway activation enhances SuSCs migration, collagen synthesis, and cellular morphological reorganization. Validation in macrophage-elimination murine models further corroborated this regulatory axis. Collectively, our work maps the dynamic microenvironmental changes during TSDO and elucidates cell-cell interaction-driven ECM collagen remodeling. These insights advance the understanding of TSDO-mediated osteogenesis and provide a foundation for developing optimized therapeutic strategies.

Leading gene therapy approaches for Duchenne muscular dystrophy (DMD) using AAV-mediated delivery of microdystrophin (µDys) have shown partial efficacy in patients, contrasting with the favorable outcomes observed in animal models. The identification of effective therapeutic strategies could be accelerated by using human high-throughput DMD models that replicate the molecular complexity driving pathology for accurate screening. To face this challenge, we develop MYOrganoids, an engineered muscle platform derived from patient-induced pluripotent stem cells (iPSC), recapitulating critical hallmarks of DMD, such as fibrosis and muscle dysfunction. We show that co-culture of fibroblasts with iPSC-derived muscle cells during organoid generation is pivotal for functional maturation and muscle force evaluation upon eccentric contractions. Notably, incorporation of DMD fibroblasts induced phenotypic exacerbation in DMD MYOrganoids by unraveling of fibrotic signature and fatiguability through cell-contact and paracrine mechanisms. We then exploited our system to interrogate gene therapy efficacy in this severe context. Although µDys gene transfer improves muscle resistance and partially restores membrane stability, it fails to reduce profibrotic signaling. These findings highlight the persistence of fibrotic activity post-gene therapy in our system, a limitedly explored aspect in DMD models, and provide the opportunity to study mechanisms of dysregulated cellular communication and empower gene therapy efficacy.