Over the past three decades, cell sheet technology has evolved from its original scaffold-free approach developed for epithelial cells into a versatile platform applicable to mesenchymal stromal cells (MSC). Despite numerous experimental achievements and convincing preclinical outcomes, MSC-based cell sheets have yet to reach the stage of a clinically reproducible product. This gap reflects not so much the technological limitations of the method as an incomplete understanding of its biological nature. MSC-based cell sheets represent more than a vehicle for cell delivery; they are self-organizing systems governed by intrinsic biophysical and morphogenetic principles. Their structural maturation involves (1) cellular condensation, (2) extracellular matrix deposition, and (3) contractile remodeling-processes that mirror the early phases of granulation tissue formation. Viewing the cell sheet as an in vitro model of connective tissue regeneration opens new research avenues extending beyond its therapeutic applications. This review summarizes the key milestones in the development of MSC-derived cell sheet technology, identifies major challenges and conceptual inconsistencies, and discusses the potential of studying these constructs as autonomous biological systems. The integration of mechanobiology, spatial omics technologies, and tissue engineering approaches may help reconceptualize MSC-based cell sheets both as tools for translational therapy and as a fundamental model for studying self-organizing regenerative processes.

Cucurbitacin B is a naturally occurring tetracyclic triterpenoid extracted from plants in the Cucumis melo L. It demonstrates various pharmacological activities, such as hepatoprotective, anti-inflammatory, and anti-tumor effects. However, its therapeutic effect on inflammation-associated colorectal cancer (CRC) and its mechanism of action have not been elucidated. This study is aimed to investigate the effects of cucurbitacin B on inflammation-associated CRC and the mechanism of action. In vitro, the effect of cucurbitacin B on the inflammatory response and the related ZNF70/NLRP3 pathway was examined using western blotting, CCK-8, ELISA, and immunofluorescence assays. In the HCT116/THP-1 supernatant co-culture system, EdU, colony formation, and wound healing assays were performed to evaluate the effects of cucurbitacin B on the proliferation, migration, and epithelial-mesenchymal transition (EMT) progression of HCT116 cells in response to inflammation. In vivo, a mouse model of inflammatory CRC was constructed by administering azoxymethane (AOM) and dextran sodium sulfate (DSS). To study the role of ZNF70, an adenoviral vector AAV was used to knock out ZNF70 in mice. High-performance liquid chromatography (HPLC) was utilized to assess the toxic impact of cucurbitacin B on mice. Our results indicate that cucurbitacin B reduces the expression of NLRP3 inflammasome-associated proteins by downregulating the production of ZNF70. The co-culture experiment showed that cucurbitacin B inhibited the proliferation, migration, and EMT of HCT116 cells in the inflammatory microenvironment. In vivo studies demonstrated that the knockdown of ZNF70 restored body weight, improved colon length, reduced tumor burden, and increased survival rate in AOM/DSS model mice. This effect was further enhanced by the addition of cucurbitacin B. The HPLC results showed that cucurbitacin B was non-toxic to the heart, liver, spleen, lungs, and kidneys of mice. In conclusion, the downregulation of ZNF70 by cucurbitacin B inhibits the activation of NLRP3 inflammasomes and reduces the promoting effect of the inflammatory microenvironment on CRC proliferation, migration, and the EMT process. Our findings may provide new insights for the development of treatments for inflammation-associated CRC.

Major lower-limb amputation often leads to wound complications that delay recovery and prosthetic use. Negative Pressure Wound Therapy (NPWT) has been suggested to improve outcomes, but evidence is inconsistent. The objective of this study is to compare NPWT with standard wound therapy in fresh amputation stumps. A systematic review and meta-analysis following PRISMA and Cochrane guidelines included randomised and controlled observational trials. Primary outcomes were wound complications, including surgical-site infections (SSI) and wound dehiscence. Secondary outcomes included re-amputation, healing, and resource use. Seventeen papers including over 6745 patients were analysed. NPWT significantly reduced total wound complications (RR 0.49, 95% CI 0.37-0.66) and SSIs (RR 0.50, 95% CI 0.27-0.90), including both superficial (RR 0.30, 95% CI 0.12-0.73) and deep infections (RR 0.26, 95% CI 0.11-0.63). The secondary amputation risk was lowered by one-third (RR 0.67, 95% CI 0.50-0.89). Wound dehiscence risk decreased (RR 0.53, 95% CI 0.30-0.94). Mortality, readmission, and hospital stay showed no significant differences between groups. Healing outcomes strongly favoured NPWT: mean time to ≥ 76% granulation was shortened by 25 days, while complete closure occurred nearly two weeks earlier. Kaplan-Meier reconstruction confirmed that NPWT accelerated wound maturation by 77% (HR 0.23, 95% CI 0.12-0.44). Economic evaluations, though limited, indicated fewer dressing changes, reduced outpatient visits, and overall cost savings. NPWT after major lower-limb amputation halves wound complications and infections, lowers re-amputation, and accelerates healing. Despite unchanged systemic outcomes, the local benefits are clinically significant, supporting its use in standard post-amputation care. Trial Registration: CRD420251088214.

Low-level laser therapy (LLLT) is being increasingly applied in regenerative medicine due to its ability to modulate cellular metabolism and support repair processes. However, the effect of various exposure parameters on fibroblast activity still does not remain fully understood. The study aim was to evaluate the effect of selected LLLT parameters (808 nm, 100 and 200 mW, 2 and 10 J/cm², pulsed beam) on the biological activity of fibroblasts in vitro. Fibroblasts were exposed to the established exposure parameters, and then the number of adherent and metabolically active cells, their migration, adenylate kinase (AK) as well as nitric oxide (NO) levels, cytokine secretion, metalloproteinase (MMP-2, MMP-9) activity, and oxidative-antioxidant potential were assessed. The most favourable effects were achieved with 200 mW; 2 J/cm² and 100 m; 10 J/cm² exposure parameters. In these conditions, increased number of adherent and metabolically active cells, and elevated migration were observed. Moreover, in this condition decreased AK release and NO secretion were found. Furthermore, LLLT reduced in fibroblasts interleukin 6 (IL-6) and interleukin 8 (IL-8) secretion as well as affected the activity of metalloproteinases, what corresponded with subsequent wound healing process. Additionally, a beneficial effect on maintaining the oxidative-antioxidant balance in fibroblasts was noted. In summary, the effects of LLLT on fibroblasts depend on the applied irradiation parameters, and appropriately selected LLLT treatment (200 mW; 2 J/cm² and 100 mW; 10 J/cm²,) stabilise fibroblast metabolism, limits their proinflammatory response and promotes extracellular matrix remodelling (ECM). These results confirm the potential of LLLT as an effective method supporting regenerative processes in the treatment of tissue damage.

Human amniotic membrane (hAM) is a widely used biomaterial with longstanding utility in ophthalmology and emerging therapeutic promise across orthopaedics, obstetrics and gynaecology, and dermatology. Clinically available biological tissue repair materials primarily encompass autologous, allogeneic, and xenogeneic tissues. However, autologous materials are limited by availability, while allogeneic and xenogeneic tissues often present challenges related to immunocompatibility. As a commonly used allogeneic biomaterial, the hAM is regarded as a highly promising tissue repair material owing to its favourable immunological profile and exceptional tissue-regenerative properties. HAM is the innermost natural barrier of the placenta and possesses unique structural and biological characteristics that facilitate tissue repair and regeneration. This review summarizes recent advances and clinical applications of hAM in orthopaedics, obstetrics and gynaecology, and dermatology, specifically focusing on its roles in promoting tendon repair, alleviating osteoarthritis, repairing endometrial injury, treating diabetic foot ulcer, and enhancing burn wound healing. With the continued development of regenerative medicine, hAM is expected to play an increasingly important role in diverse tissue repair and regenerative medicine applications.

Tendon injuries account for considerable and escalating clinical burden. After injury, tendons heal poorly with persistent fibrovascular scar. Unlike healthy tendon which is type I collagen (COL1)-rich and highly-aligned, fibrovascular scar is type III collagen (COL3)-rich and disorganized; this compositional and organizational change has been implicated in the functional deficits resulting from tendon injury. Accordingly, COL3 is historically considered a primary driver of poor tendon healing. Given compelling evidence of COL3's role in regulating matrix structure and cell behavior, we sought to define the role of COL3 in driving physiologic/pathologic tendon healing through regulation of all phases of healing. Leveraging a mouse model of global, inducible Col3a1 knockdown, we reduced Col3a1 expression at the time of patellar tendon injury and investigated extracellular matrix, cellular, and mechanical changes at 1-, 3-, and 6-weeks post-injury. Based on data from other tissues, we hypothesized that COL3 loss would exacerbate poor tendon healing, with temporally-distinct contributions to tendon healing outcomes resulting from COL3's critical regulation of matrix organization, cell phenotype and activities, and tissue mechanics. Unexpectedly, reduction of COL3 in young adult mice did not change collagen architecture and had few, nuanced impacts on cell behavior and tissue mechanics throughout healing. These findings challenge the conventional paradigm that COL3 drives poor tendon healing outcomes and emphasize the need to identify matrix and cell mechanisms contributing to poor tendon healing. Understanding COL3's role in other injury models and lifespan contexts, including tendon development and aging, will provide more insight into its regulatory and therapeutic potential.

Accelerating the healing of infected wounds remains a significant clinical challenge due to the complex interplay between persistent infection and impaired cellular signaling. Herein, we developed a bio-inspired, multifunctional hydrogel (KGMBSP@ZnO) by synergistically crosslinking konjac glucomannan (KGM) and Bletilla striata polysaccharide (BSP) to create a pro-regenerative microenvironment. Beyond its robust mechanical toughness and self-healing properties, this study uncovers the hydrogel's dual-functional role in infection eradication and bio-instructed tissue remodeling. Mechanistically, KGMBSP@ZnO does not merely act as an antibacterial barrier; it actively directs wound repair by orchestrating the macrophage-to-myofibroblast transition (MMT)-a critical yet under-explored pathway in material-mediated regeneration. In vitro analysis reveals that the hydrogel's bioactive matrix triggers endothelial proliferation and migration through the integrated activation of PI3K/Akt, MAPK/ERK, and Rho GTPase pathways. In a murine infected wound model, the hydrogel effectively suppressed chronic inflammation by modulating the TNF-α/NF-κB and IL-10 signaling axes while significantly promoting functional angiogenesis and organized collagen deposition. Our findings demonstrate that the KGMBSP@ZnO hydrogel provides a promising "immunomodulatory-to-regenerative" strategy, shifting the paradigm from passive wound dressing to an active therapeutic platform that modulates macrophage plasticity to achieve rapid, high-quality tissue regeneration.

Wounds, particularly chronic ones, pose significant clinical challenges due to their persistent nature and high risk of complications, which severely compromise patient quality of life and impose substantial socioeconomic burdens. This study presents a novel and sustainable strategy for advanced wound management by developing multifunctional dressing platforms based on β-chitosan (β-CS) derived from squid pen waste. An optimized biochemical extraction process was developed to obtain high-purity β-CS with suitable crystallinity and an elevated deacetylation degree. To harness these properties, β-CS was formulated with thermoresponsive Poloxamer 407 (P407), yielding two innovative wound healing systems: an in-situ forming film (CSPF) and a fast-powder (CSPP). To fully exploit the inherent bioactivity of β-CS, two novel wound healing platforms, namely an in-situ formed composite film (CSPF) and fast-deployable composite powder (CSPP), were fabricated by integrating β-CS with thermosensitive Poloxamer 407 (P407). In vitro characterization revealed that both formulations exhibited potent hemostatic capacity, broad-spectrum antibacterial activity, and outstanding hemocompatibility. In a murine full-thickness wound model, CSPF and CSPP significantly accelerated wound closure, enhanced collagen deposition, and promoted neovascularization. These dressings attenuated early inflammation through downregulation of key pro-inflammatory cytokines (IL-1β, IL-6, TNF-α), while activating the TGF-β/β-catenin pathway to facilitate fibroblast-to-myofibroblast transition and extracellular matrix remodeling. Furthermore, CSPF/CSPP enhanced VEGF-mediated angiogenesis, as evidenced by increased CD31+ microvessel density. By converting marine waste into bioactive therapeutics, this work advances sustainable biomaterial development and shifts wound care from passive coverage to active biological intervention.

The persistent non-healing of cutaneous wounds in diabetic patients represents a critical clinical challenge requiring urgent resolution. Excessive inflammation, impaired angiogenesis, and aberrant collagen remodeling profoundly hinder wound recovery. This study explores the potential therapeutic value of miR-221-3p in diabetic cutaneous wound healing using murine models. Subcutaneous administration of miR-221-3p was associated with accelerated wound closure; histological staining and Western blot analyses showed decreased expression of inflammatory markers (IL-1β, IL-6, MPO, CD68), increased levels of angiogenic markers (CD31, VEGFA), and enhanced collagen I/III deposition at wound margins. Complementary experiments using Mir221 knockout mice showed delayed healing, which was accompanied by upregulated MPO/CD68 expression, reduced CD31 levels, and decreased collagen fiber formation. These bidirectional observations suggest that miR-221-3p may contribute to diabetic wound healing, potentially through effects on inflammatory responses, neovascularization, and collagen synthesis. These findings suggest that miR-221-3p could serve as a potential therapeutic target for refractory wounds in diabetic patients, including diabetic foot ulcers and other chronic cutaneous lesions.