Volumetric muscle loss (VML), a condition affecting millions due to trauma, represents a critical unmet need in regenerative medicine, with no established standard of care. This study introduces a de novo therapeutic strategy using tissue-specific skeletal muscle acellular extracellular matrix (aECM) fibers fabricated using scaffold-free Anchored Cell Sheet Engineering technique. These engineered fibers replicate the native ECM composition and microarchitecture of skeletal muscle, incorporating essential structural and basement membrane proteins. In a rat VML model, aECM demonstrated promising regenerative capacity compared to commercial porcine-derived small intestine submucosa (SIS) ECM. Over an 8-week period, compared to contralateral muscle, aECM preserved muscle volume and weight, regulated inflammatory and fibrotic responses, and promoted vascularization. In contrast, SIS was rapidly degraded by week 4 and associated with fibrosis. Force recovery in aECM was lower at the 8-week time point (77 % compared to 91 % in control), but histological and immunohistochemical analyses revealed newly formed, dispersed muscle fibers exclusively in aECM treatment. Importantly, muscle weight was preserved only when aECM was used, resulting in similar normalized force-to-weight across all groups (87 % in aECM vs. 88 % in SIS). The histological analyses further demonstrated ongoing tissue remodeling, indicative of sustained regeneration, in contrast to the premature fibrotic healing observed in the other groups. An innovative quantitative image analysis workflow enabled assessment of spatial tissue heterogeneity through histology and immunohistochemistry images, setting a new standard for regeneration analysis. These findings establish engineered tissue-specific aECM as a transformative approach for VML treatment, laying the groundwork for translation to clinical applications. STATEMENT OF SIGNIFICANCE: The current study introduces a transformative approach to treating volumetric muscle loss (VML) through the development of tissue-specific acellular extracellular matrix (aECM) fibers engineered using a scaffold-free biofabrication platform uniquely suited for recreation of such aECM components. The engineered fibers represent a significant advancement over current commercial options by recreating native ECM composition and microarchitecture while eliminating complications associated with xenogenic materials. Through comprehensive in vivo evaluation in a rat model, it is demonstrated that these engineered fibers maintain muscle mass and promote controlled tissue regeneration, addressing key limitations of existing treatments. The scaffold-free biofabrication of tissue-specific aECM provides a new paradigm for biomaterial design in regenerative medicine.

The advent of intestinal organoids, three-dimensional structures derived from stem cells, has significantly advanced the field of biology by providing robust in vitro models that closely mimic the architecture and functionality of the human intestine. These organoids, generated from induced pluripotent stem cells (iPSCs), embryonic stem cells (ESCs), or adult stem cells, possess remarkable capabilities for self-renewal, differentiation into diverse intestinal cell types, and functional recapitulation of physiological processes, including nutrient absorption, epithelial barrier integrity, and host-microbe interactions. The utility of intestinal organoids has been extensively demonstrated in disease modeling, drug screening, and personalized medicine. Notable examples include iPSC-derived organoids, which have been effectively employed to model enteric infections, and ESC-derived organoids, which have provided critical insights into fetal intestinal development. Patient-derived organoids have emerged as powerful tools for investigating personalized therapeutics and regenerative interventions for conditions such as inflammatory bowel disease (IBD), cystic fibrosis, and colorectal cancer. Preclinical studies involving transplantation of human intestinal organoids into murine models have shown promising outcomes, including functional integration, epithelial restoration, and immune system interactions. Despite these advancements, several challenges persist, particularly in achieving reproducibility, scalability, and maturation of organoids, which hinder their widespread clinical translation. Addressing these limitations requires the establishment of standardized protocols for organoid generation, culture, storage, and analysis to ensure reproducibility and comparability of findings across studies. Nevertheless, intestinal organoids hold immense promise for transforming our understanding of gastrointestinal pathophysiology, enhancing drug development pipelines, and advancing personalized medicine. By bridging the gap between preclinical research and clinical applications, these organoids represent a paradigm shift in the exploration of novel therapeutic strategies and the investigation of gut-associated diseases.

Tracheal reconstruction presents a significant global clinical challenge due to the unique structure and function of the trachea, which complicates its repair. Key challenges include restoring tracheal function post-transplantation, managing surgical complications, and ensuring the long-term survival of transplanted tissue. Furthermore, adequate vascularization of the transplanted trachea, along with the repair of cartilage and epithelial cells, is critical for facilitating functional recovery and successful reconstruction. In this study, a decellularized tracheal matrix was extracted to preserve its natural bioactivity. Nanoclay and GelMA were then employed as carrier materials to integrate with the decellularized tracheal matrix, forming a hydrogel scaffold. Both nanoclay and GelMA offer tunable porous structures and surface properties that promote cell adhesion and proliferation, while providing sufficient mechanical support. By leveraging the biological functions of endothelial progenitor cell-derived exosomes, we successfully loaded exosomes onto the decellularized extracellular matrix, creating a novel cell-free tracheal scaffold for investigating tracheal defect repair. The scaffold exhibited excellent biocompatibility, promoting graft vascularization and facilitating the repair of tracheal cartilage and epithelium. These findings hold significant promise for advancing tracheal reconstruction surgery and offer valuable insights for future research in this area.

Tissue engineering, a multidisciplinary research field aiming to revolutionize regenerative medicine, relies on scaffolds for optimal cell cultures and organ development. Decellularized tissue extracellular matrices (dECM) scaffolds, particularly from human amniotic membrane (hAM), show promise in clinical applications. This review discusses the significance of scaffolds, emphasizing dECM-based hAM scaffolds, delving into ECM complexities, decellularization processes, and evaluation methods. Raman spectroscopy emerges as a non-destructive tool for evaluating ECM preservation, presenting potential for quantifying ECM components in hAM before and after decellularization. The review explores the role of hAM as a biomaterial, detailing its composition and characteristics and emphasizes the importance of evaluating ultrastructural components and suggests Raman spectroscopy as a valuable technique for this purpose.

This study presents a novel method to enhance the biocompatibility of decellularized porcine aortic segments while preserving their mechanical properties and histological structure. Detergent-decellularized aortic segments were treated with modified globular chitosan (Novochizol™) at varying concentrations (0.5%, 1%, 2%, and 3%) by sonication and subsequently subjected to mechanical testing. To further improve cell infiltration, blind-ended laser channels were created within the decellularized segments. The modified grafts were then seeded with porcine vascular interstitial cells in vitro for 7 days or implanted into the thoracic aorta of minipigs for 30 days. Histological analysis was performed at each stage of the study. Impregnation with Novochizol™ significantly increased the specific strength (from 0.97 ± 0.19 MPa to 4.99 ± 2.43 MPa) and Young's modulus (from 0.73 ± 0.06 MPa to 14.66 ± 7.14 MPa) of the decellularized aortic segments. Histological examination confirmed the preservation of the connective tissue matrix's morphological structure. Optimal modification conditions were identified as a 30 min sonication in a 1% Novochizol™ solution at 25 °C. A 35 ms continuous laser treatment was sufficient to create a 1 mm deep blind-ended channel, thereby promoting the seeding of vascular interstitial cells within the acellular graft, as confirmed by implantation in minipigs.

The extracellular matrix (ECM) is a non-cellular three-dimensional structure present in all tissues that is essential for the intestinal maintenance, function and structure, as well as for providing physical support for tissue integrity and elasticity. ECM enables the regulation of various processes involved in tissue homeostasis, being vital for healing, growth, migration and cell differentiation. Structurally, ECM is composed of water, polysaccharides and proteins, such as collagen fibers and proteoglycans, which are specifically arranged for each tissue. In pathological scenarios, such as inflammatory bowel disease (IBD), the deposition and remodeling of the ECM can be altered in relation to the homeostatic composition. IBD, such as Ulcerative colitis and Crohn's disease, can be differentiated according to ECM alterations, such as circulating levels of collagen, laminin and vimentin neoepitopes. In this context, ECM presents particularities in both physiological and pathological processes, however, exploring methods of tissue decellularization is emerging as a promising frontier for new therapeutic interventions and clinical protocols, promoting the development of new approaches to intestinal diseases.

In this study, the optimal engraving parameters were determined through the analysis of scanning electron microscopy (SEM) data, as follows: a laser power density of 5.5 × 105 W/cm2, an irradiation rate of 0.1 mm/s, a well radius of 60 μm, a distance between well centers of 200 μm, and a number of passes for each well of 20. After 1 week of in vitro cultivation, chondrocytes were located on the surface of the scaffolds, in the sockets and lacunae of decellularized cartilage. When implanted into animals, both cellular and acellular scaffolds were able to support cartilage in-growth and complete regeneration of the defect without clear boundaries with normal tissue. Nevertheless, the scaffolds populated with cells exhibited superior biocompatibility and were not subject to rejection, in contrast to cell-free constructs.

Achilles tendon is vital in maintaining the stability and function of ankle joint. It is quite difficult to achieve the structural and functional repair of Achilles tendon in tissue engineering.

A tissue-engineered tendon micro-tissue was prepared using rat tail tendon extracellular matrix (TECM) combined with rat adipose stem cells (ADSCs) to repair Achilles tendon injuries. The TECM was prepared by repeated freezing and thawing. The in vitro characteristics of TECM and its effect on ADSCs proliferation were detected. This tissue-engineered tendon micro-tissue for Achilles tendon repair in vivo was evaluated based on general characteristics, gait analysis, ultrasound findings, histological analysis, and biomechanical testing.

The results showed that the TECM scaffold had good biocompatibility for ADSCs. At 2 weeks post-surgery, collagen types I and III and tenomodulin expression were higher, and vascular endothelial growth factor expression was lower in the micro-tissue group than other groups. At 4 and 8 weeks post-surgery, the results of histological analysis and ultrasound findings showed that the repaired tendon tissue was smooth and lustrous, and was arranged regularly and evenly in the micro-tissue group. Gait analysis confirmed that better motor function recovery was noted in micro-tissue group than other groups. In addition, the mechanical properties of the repaired tendon tissue in micro-tissue group were better than other groups.

Tissue-engineered tendon micro-tissue fabricated by TECM and ADSCs has good biocompatibility and can promote structural and functional repair of tendon in vivo. This composite biomaterial has broad application prospects in tissue engineering.

The extracellular matrix (ECM) is a multifunctional network of macromolecules that regulate various cellular functions and physically support the tissues. Besides physiological conditions, the ECM also changes during pathological conditions such as cancer. As tumor cells proliferate, notable changes occur in the quantity and makeup of the surrounding ECM. Therefore, the role of this noncellular component of tissues in studies of tumor microenvironments should be considered. So far, many attempts have been made to create 2-dimensional (2D) or 3-dimensional (3D) models that can replicate the intricate connections within the tumor microenvironment. Decellularized tissues are proper scaffolds that imitate the complex nature of native ECM. This review aims to summarize 3D models of digestive system cancers based on decellularized ECMs. These ECM-based scaffolds will enable us to study the interactive communication between cells and their surrounding environment which brings new potential for a better understanding of the pathophysiology of cancer.

Articular cartilage is a highly specialized tissue able to tolerate physical stress. However, its capacity for restoration is restricted, and injuries to the cartilage do not recover spontaneously. Interest in mesenchymal stem cells derived from human adipose tissue (hASCs) is growing due to their potential to improve tissue healing and recovery. Decellularized extracellular matrix (dECM)-based hydrogels combined with hASCs could serve as an interface for studying behavior and differentiation properties in a cartilage microenvironment. In the present study, we described the behavior of hASCs cultured in a commercial dECM MatriXpec™. The structural microtopography of MatriXpec™ was analyzed by scanning electron micrography, and its protein composition was accessed by mass spectrometry. The protein composition of MatriXpec™ is mainly represented by collagen proteins, building its fibrous ultrastructure. hASCs were cultured three-dimensionally (3D) on MatriXpec™ to perform cell viability, growth, and cartilage differentiation analysis. We showed that MatriXpec™ could be loaded with hASCs and that it supports cell maintenance for several days. We observed that the three-dimensional ultrastructure of the biomaterial is composed of nanofibers, and its protein composition reflects the tissue from which it was harvested. Finally, we showed that the molecular cues from the hydrogel are biologically active as these influence cell behavior and differentiation phenotype, increasing the expression of fibrocartilage-related genes such as SOX9, COL1, COL10, and MMP13. MatriXpec™ hydrogel can be used as an interface for 3D hASCs culture studies as it maintains cell viability and supports its differentiation process.

Decellularized matrices derived from diseased tissues/organs have evolved in the most recent years, providing novel research perspectives for understanding disease occurrence and progression and providing accurate pseudo models for developing new disease treatments. Although decellularized matrix maintaining the native composition, ultrastructure, and biomechanical characteristics of extracellular matrix (ECM), alongside intact and perfusable vascular compartments, facilitates the construction of bioengineered organ explants in vitro and promotes angiogenesis and tissue/organ regeneration in vivo, the availability of healthy tissues and organs for the preparation of decellularized ECM materials is limited. In this paper, we review the research advancements in decellularized diseased matrices. Considering that current research focuses on the matrices derived from cancers and fibrotic organs (mainly fibrotic kidney, lungs, and liver), the pathological characterizations and the applications of these diseased matrices are mainly discussed. Additionally, a contrastive analysis between the decellularized diseased matrices and decellularized healthy matrices, along with the development in vitro 3D models, is discussed in this paper. And last, we have provided the challenges and future directions in this review. Deep and comprehensive research on decellularized diseased tissues and organs will promote in-depth exploration of source materials in tissue engineering field, thus providing new ideas for clinical transformation.

In recent years, significant biotechnological advancements have been made in engineering human cardiac tissues and organ-like models. This field of research is crucial for both basic and translational research due to cardiovascular disease being the leading cause of death in the developed world. Additionally, drug-associated cardiotoxicity poses a major challenge for drug development in the pharmaceutical and biotechnological industries. Progress in three-dimensional cell culture and microfluidic devices has enabled the generation of human cardiac models that faithfully recapitulate key aspects of human physiology. In this review, we will discuss 3D pluripotent stem cell (PSC)-models of the human heart, such as engineered heart tissues and organoids, and their applications in disease modeling and drug screening.