Ryosuke Iwai

Postoperative aneurysm sac enlargement is a significant clinical issue in endovascular aortic aneurysm repair that is potentially associated with impaired microcirculation in the aneurysmal wall. We developed centimeter-long, fiber-shaped aggregates of human bone-marrow-derived mesenchymal stromal cells (HMSC fiber) to function as a scaffold-free cellular construct applicable to endovascular treatment. HMSC fibers were prepared using a cell self-aggregation technique and optimized by controlling the cell number per unit length to preserve cellular viability and mechanical stability. The resulting fibers retained mesenchymal stromal cell characteristics and endogenous extracellular matrix, facilitating smooth handling and intraluminal delivery without structural collapse. After transcatheter administration into a swine aortic aneurysm model, HMSC fiber-induced fibroconnective tissue formation occurred with capillary-like structures within the aneurysm sac. These findings demonstrate the feasibility of HMSC fiber as a controllable and stable platform for localized endovascular cell delivery. Furthermore, this study established their potential utility as a regenerative adjunct to current endovascular treatment for aortic disease.

Background: Diabetic foot ulcers (DFUs) are a severe complication of diabetes, associated with high morbidity and amputation risk. In-body tissue architecture (iBTA)-induced autologous biococktails, generated using a newly designed device (BSM1), offer a novel in vivo regenerative approach using somatic pluripotent stem cells (PSCs).Methods: This exploratory, single-arm clinical study evaluated the feasibility, safety, and efficacy of BSM1-derived biococktails in ten patients with chronic DFUs. Biococktails were fabricated through subcutaneous embedding of the BSM1 device for three weeks and applied to DFUs. The primary endpoint was successful formation of PSC-containing biococktails. Secondary endpoints included wound closure, reduction in wound area and depth, granulation, and adverse events over 12 weeks. Histology and mRNA analyses were performed to characterize biococktail components.Findings: BSM1 embedding and extraction were successful in all participants (100%). All participants completed the study without major complications. Biococktails from 37 devices were applied at an average density of 0·35 g/cm² over a mean wound area of 10·24 cm2. During the embedding period, when only standard wound care was provided, wound area and depth showed minimal change. Following application, both parameters decreased, reaching statistically significant improvement at 2 and 3 months. At 3 months, wound area decreased by 84·15 ± 30·46% (median 98·48%), while wound depth halved within 1 week and decreased continuously thereafter. Histology confirmed the presence of markers associated with PSCs, mesenchymal stem cells, M2 macrophages, myofibroblasts, capillaries, and extracellular matrix components. Gene expression profiling identified mRNAs encoding growth factors (VEGFA, HGF, PDGF, TGF-β1) and anti-inflammatory cytokines (IL-10). No amputations or severe biococktail-related adverse events were observed.Interpretation: BSM1-derived autologous biococktails represent a safe, feasible, and innovative in vivo-assembled regenerative strategy that may induce sustained wound improvement beyond standard care in patients with chronic DFUs. Larger controlled studies are warranted.

Postoperative aneurysm sac enlargement is a significant clinical issue in endovascular aortic aneurysm repair that is potentially associated with impaired microcirculation in the aneurysmal wall. We developed centimeter-long, fiber-shaped aggregates of human bone marrow-derived mesenchymal stromal cells (HMSC fiber) to function as a scaffold-free cellular construct applicable to endovascular treatment. HMSC fibers were fabricated using a cell self-aggregation technique and optimized by controlling the cell number per unit length to preserve cellular viability and mechanical stability. The resulting fibers retained mesenchymal stromal cell characteristics and endogenous extracellular matrix, facilitating smooth handling and intraluminal delivery without structural collapse. After transcatheter administration into a swine aortic aneurysm model, HMSC fiber-induced fibroconnective tissue formation stimulated neovascularization within the aneurysm cavity. These findings demonstrate the feasibility of HMSC fiber as a controllable and stable platform for localized endovascular cell delivery. Furthermore, this study established their potential utility as a regenerative adjunct to current endovascular treatment for aortic disease.

Several studies have investigated the location of transplanted cells and tissue‐engineered cell constructs in the body by incorporating contrast nanoparticles into cells by endocytosis; however, these have yet to be applied clinically because of the complexity of assessing the safety of nanoparticles. In this study, we proposed that our developed adherent cell self‐aggregation technique (CAT) could be used to develop cell aggregates loaded with contrast particles of a size that would exclude the possibility of endocytosis, and aimed to prepare these aggregates followed by biological and computed tomography (CT) contrast evaluation under X‐rays. Once human bone marrow‐derived mesenchymal stem cells (HBMSCs) were seeded into culture dishes coated with CAT‐inducing polymer to form gapless cell monolayer sheets, tungsten carbide (WC) particles smaller than 1 μm or titanium (Ti) particles larger than 10 μm were added, and thus each particle deposited on the surface of the cell monolayer sheet. During the subsequent overnight incubation, spontaneous detachment and aggregation of the cell monolayer sheets with deposited WC and Ti particles occurred, forming single spherical cell aggregates (spheroids) and loading these particles. Histological analysis confirmed that Ti particles with a diameter of at least 10 μm were not endocytosed and remained attached to the outside of cells forming spheroids, while WC particles were endocytosed into the cells. The CT images of the Ti‐loaded spheroids were clearly visible along the spheroid shape under X‐ray irradiation. Then, we confirmed that there was no toxicity to the cells forming the spheroids by loading Ti particles, and the cells could sprout and proliferate by culturing the spheroids. We successfully prepared Ti particle‐loaded HBMSCs aggregates with long fiber shape (> 10 cm) by applying CAT to a culture dish with a ring‐fiber‐shaped culture groove and confirmed their clear visibility on CT images under X‐ray irradiation, as well as their containment and ejection into a catheter, demonstrating their applicability to catheter‐mediated regenerative therapy.

This study investigated the long-term patency of regenerative Biotube grafts and discusses their feasibility as an alternative to autologous vein grafts for peripheral artery disease. Six Biotubes with a diameter of 4 mm were autologously fabricated in recipients using in vivo tissue engineering (in-body tissue architecture) technology and implanted as carotid artery bypass grafts in a goat model. All six grafts remained patent at 6 months despite exceeding 10 cm in length, demonstrating their biocompatibility and durability. Histological analysis revealed neointima formation, endothelialization, and minimal inflammation. However, in one goat, a graft developed stenosis, while another showed dilatation. These findings demonstrate the use of Biotubes as a viable option for peripheral vascular reconstruction as tissue-engineered vascular grafts. However, further optimization is needed to address emerging issues with their use, such as stenosis and aneurysm formation, to improve long-term patency.

The limited capacity of articular cartilage for self-repair is a critical challenge in orthopedic medicine. Here, we aimed to develop a simplified method of generating chondrocyte particles from human-induced pluripotent stem cell-derived expandable limb-bud mesenchymal cells (ExpLBM) using a cell self-aggregation technique (CAT). ExpLBM cells were induced to form chondrocyte particles through a stepwise differentiation protocol performed on a CAT plate (prevelex-CAT^®), which enables efficient and consistent production of an arbitrary number of uniformly sized particles. Histological and immunohistochemical analyses confirmed that the generated chondrocyte particles expressed key cartilage markers, such as type II collagen and aggrecan, but not hypertrophic markers, such as type X collagen. Additionally, when these particles were transplanted into osteochondral defects in rats with X-linked severe combined immunodeficiency, they demonstrated successful engraftment and extracellular matrix production, as evidenced by Safranin O and Toluidine Blue staining. These data suggest that the plate-based CAT system offers a robust and scalable approach to produce a large number of chondrocyte particles in a simplified and efficient manner, with potential application to cartilage regeneration. Future studies will focus on refining the system and exploring its clinical applications to the treatment of cartilage defects.

Securing high-quality cell sources is important in regenerative medicine. In this study, we developed a device that can accumulate autologous stem cells in the body. When small wire-assembled molds were embedded in the dorsal subcutaneous pouches of beagles for several weeks, collagen-based tissues with minimal inflammation formed inside the molds. At 3 weeks of embedding, the outer areas of the tissues were composed of immature type III collagen with large amounts of cells expressing SSEA3 or SSEA4 markers, in addition to growth factors such as HGF or VEGF. When separated from the tissues by collagenase treatment, approximately four million cells with a proportion of 70% CD90-positive and 20% SSEA3- or SSEA4-positive cells were recovered from the single mold. The cells could differentiate into bone or cartilage cells. The obtained cell-containing tissues are expected to have potential as therapeutic materials or cell sources in regenerative medicine.

The administration of mesenchymal stem cells (MSCs) has a positive effect on wound healing; however, the lack of adequate MSC engraftment at the wound site is a major limiting factor in current MSC-based therapies. In this study, a biosheet prepared using in-body tissue architecture (iBTA) was used as a material to address these problems. This study aimed to assess and evaluate whether biosheets containing somatic stem cells would affect the wound healing process in dogs. Biosheets were prepared by subcutaneously embedding molds in beagles. These were then evaluated grossly and histologically, and the mRNA expression of inflammatory cytokines, interleukins, and Nanog was examined in some biosheets. Skin defects were created on the skin of the beagles to which the biosheets were applied. The wound healing processes of the biosheet and control (no biosheet application) groups were compared for 8 weeks. Nanog mRNA was expressed in the biosheets, and SSEA4/CD105 positive cells were observed histologically. Although the wound contraction rates differed significantly in the first week, the biosheet group tended to heal faster than the control group. This study revealed that biosheets containing somatic stem cells may have a positive effect on wound healing.

This study investigated six-month outcomes of first models of ascending aortic replacement. The molds used to produce the Biotube were implanted subcutaneously in goats. After 2–3 months, the molds were explanted to obtain the Biotubes (inner diameter, 12 mm; wall thickness, 1.5 mm). Next, we performed ascending aortic replacement using the Biotube in five allogenic goats. At 6 months, the animals underwent computed tomography (CT) and histologic evaluation. As a comparison, we performed similar surgeries using glutaraldehyde-fixed autologous pericardial rolls or pig-derived heterogenous Biotubes. At 6 months, CT revealed no aneurysmalization of the Biotube or pseudoaneurysm formation. The histologic evaluation showed development of endothelial cells, smooth muscle cells, and elastic fibers along the Biotube. In the autologous pericardium group, there was no evidence of new cell development, but there was calcification. The histologic changes observed in the heterologous Biotube group were similar to those in the allogenic Biotube group. However, there was inflammatory cell infiltration in some heterologous Biotubes. Based on the above, we could successfully create the world’s first Biotube-based ascending aortic replacement models. The present results indicate that the Biotube may serve as a scaffold for aortic tissue regeneration.