Tissue engineering and regenerative medicine最新文献

Background: Peripheral nerve injuries are a major clinical challenge because of their complex nature and limited regenerative capacity. This study aimed to improve peripheral nerve regeneration using Wharton's jelly mesenchymal stem cells (WJ-MSCs) engineered to express brain-derived neurotrophic factor (BDNF) via a baculovirus (BV) vector. The cells were evaluated for efficacy when seeded into acellular nerve grafts (ANGs) in a rat sciatic nerve defect model.

Methods: WJ-MSCs were transfected with recombinant BV to upregulate BDNF expression. Conditioned medium (CM) from these cells was utilized to treat Schwann cells (SCs), and the impact on myelination-related markers, including KROX20, myelin basic protein (MBP), glial fibrillary acidic protein (GFAP), and S100 calcium-binding protein β (S100β), and the activation of the mammalian target of rapamycin (mTOR)/ protein kinase B (AKT)/p38 signaling pathways were evaluated. In vivo, BDNF-expressing WJ-MSCs were seeded into ANGs and implanted into a rat sciatic nerve defect model. Functional recovery was evaluated via video gait analysis, isometric tetanic force measurement, muscle weight evaluation, ankle contracture angle measurement, and histological analysis using toluidine blue staining.

Results: BDNF expression was significantly upregulated in WJ-MSCs post-transfection. BDNF-MSC CM substantially promoted the expression of myelination markers in SCs and activated the mTOR/AKT/p38 signaling pathway. In the rat model, seeding of ANGs with BDNF-expressing WJ-MSCs resulted in improved functional outcomes, including enhanced toe-off angles, increased isometric tetanic force, greater muscle weight recovery, and a higher total number of myelinated axons compared with controls.

Conclusion: WJ-MSCs engineered to express BDNF significantly enhanced peripheral nerve regeneration when utilized in conjunction with ANGs. These findings indicate BDNF-expressing WJ-MSCs are a promising therapeutic approach for treating peripheral nerve injuries.

Background: Pancreatic islet transplantation holds great potential as a therapeutic approach for treating type 1 diabetes mellitus (T1D). However, large islets suffer from hypoxia due to the limited diffusion distance of oxygen, leading to cell loss. Therefore, smaller spheroids are needed for better transplantation outcomes. This study aims to develop a method for forming highly functional islet spheroids using glycol chitosan (GC) derivatives, such as N-acetylated glycol chitosan (AGC) and N-hexanoyl glycol chitosan (HGC).

Methods: Thermogelling polymers were produced by performing N-acylation of GC using the correspondingly carboxylic anhydrides. Islet spheroids were formed using a dual application with AGC-coated plates and HGC gelation. The AGC solution was applied to the plate for coating and evenly distributed using a 1 mL syringe. Then, the HGC encapsulated with islet single cells was cultured on top of it. Spheroid viability and functionality were evaluated using CCK-8 assay and glucose-stimulated insulin secretion assay.

Results: The aqueous solutions of AGC (4%, w/v) and HGC (36% hexanoylation) (2%, w/v) demonstrated a sol-gel transition temperature around 37 °C, suitable for the physiological environment. These polymers also showed no cytotoxicity to intact islets. Islet single cells were cultured on HGC gels with varying degrees of hexanoylation (DH) values, where higher DH values led to smaller and more uniform spheroids. The resulting spheroids formed on AGC-coated plates and HGC36 gelation were smaller and more uniform than those formed on untreated plates. These spheroids exhibited significantly improved glucose responsiveness, with superior insulin secretion.

Conclusion: The optimized method using AGC and HGC offers a more efficient way to produce smaller, uniform, and functional spheroids.

Background: Atherosclerosis often leads to ischemic heart disease and peripheral artery disease. Traditional revascularization technique such as bypass grafting using autologous vessels are commonly employed. However, limitations arise when patients lack suitable grafts due to underlying diseases or previous surgeries, prompting the need to substitute vessel grafts. Due to the high biocompatibility of decellularized products (grafts or scaffolds) prepared using supercritical carbon dioxide (ScCO₂), it has been widely applied in decellularization-related technologies in recent years. Therefore, this review article will comprehensively discuss the current developments in tissue vascular scaffolds applied to the treatment of cardiovascular diseases, with a particular focus on the application of supercritical carbon dioxide technology in this field and the challenges it faces.

Method: This review was compiled by searching relevant references on PubMed database (before June 2024) based on selected key words and specific terms.

Results: ScCO₂ is an effective and eco-friendly extraction agent widely used in industries like food, pharmaceuticals, and cosmetics. It has been applied in decellularization processes to obtain extracellular matrices (ECMs) from tissues. ScCO₂ technology has emerged as a promising method in cardiovascular disease treatment, particularly for developing tissue vascular scaffolds. ScCO₂ effectively removes cellular components while preserving the ECM, ensuring high biocompatibility and reduced immune response. It has been applied to decellularize tissues like heart valves and arteries, creating scaffolds that mimic natural ECM to support cell proliferation and tissue regeneration. Despite challenges such as solubility limitations and cost, ScCO₂ offers advantages like low toxicity and ease of use, making it a valuable tool in advancing regenerative medicine for cardiovascular applications.

Conclusion: ScCO₂ has the advantages of low cellular toxicity, cost-effectiveness, and ease of manipulation. These characteristics have the potential to lead to significant progress in cardiovascular research on tissue regeneration.

Background: Cell replacement therapy is the only treatment that restores or repairs the function of impaired tissues in neurodegenerative diseases, including vascular dementia (VaD); however, current VaD treatments focus on slowing or mitigating the underlying small vessel disease progression. We aimed to verify the improvement in neurocognition after administering human-induced pluripotent stem cell (hiPSC)-derived neural progenitor cells (NPCs) from in a VaD animal model.

Methods: After anesthesia, 10-12-week-old male C5BL/6 mice underwent sham or bilateral carotid artery stenosis (BCAS) surgeries. For BCAS, 0.18-mm micro-coils were wound around the bilateral common carotid arteries to induce chronic vascular insufficiency in the global brain. One day after surgery, the mice were administered phosphate buffer solution or NPC from hiPSCs via the tail vein for 15 d, and divided into sham (n = 6), VEH (n = 6), and NPC (n = 7) groups. Three months after the surgery, neurobehavioral tests including the Y-maze test (YMT), passive avoidance test (PAT), and novel object recognition test (NORT) were performed. Finally, mice brains were sectioned for evaluating microglia (Iba-1), astrocyte (GFAP) activation, and myelin (MBP) degeneration through immunohistochemistry (IHC).

Results: PAT latency (p = 0.01) and discrimination index in the NORT (p = 0.043) increased considerably in the NPC group than in the VEH group. However, alterations in YMT were not considerably higher in the NPC group than in the VEH group (p = 0.65). IHC tests revealed that the GFAP- and IBA-1-positive cell number was remarkably lower in the NPC group than in the VEH group (p < 0.05). Moreover, MBP density was higher in the NPC group.

Conclusion: hiPSC-derived NPCs have therapeutic potential in cerebral hypoperfusion VaD mice; it improves the working memory of VaD animals by diminishing inflammatory reactions and protecting them from demyelination.

Background: Mild head trauma often leads to long-term cognitive and neurological deficits. PLX3397, an inhibitor of colony-stimulating factor 1 receptor (CSF1R), offers promise as a therapeutic agent for traumatic brain injury (TBI) by targeting neuro-inflammation. Stem cell-based approaches are widely studied for neurological disorders. The objective of this study was to investigate therapeutic effect of intranasal administration of human neural crest-derived nasal turbinate stem cells (hNTSCs) on mild TBI in comparison with that of PLX3397.

Methods: We developed a model of mice with repetitive and mild TBI following a weight-drop once a day for 5 days. PLX3397 (50 mg/kg, p. o.) was administered for 21 days. Intranasal administration of hNTSCs (1 × 10⁶) was performed once.

Results: Iba1 + and GFAP + cells were increased in the cortex and hippocampus of TBI models. Iba1 + cells and GFAP + cells were remarkably decreased in PLX3397 or hNTSC-treated TBI models. Administration of PLX3397 attenuated the decrease in neurobehavioral activity. Similar effects were observed in a TBI model with a single dose of hNTSC.

Conclusion: Intranasal administration of hNTSCs had a microglia-depleting effect. Administered hNTSCs were found around the cortex and hippocampus of TBI brains. This investigation may provide a promising path for therapeutic initiatives for repetitive and mild TBI.

Background: Buccal mucosa urethroplasty represents the primary strategy for reconstruction of long urethral strictures (US). However, significant complications including stricture recurrence and donor site morbidity currently hamper this approach. The goal of this study was to determine the efficacy of acellular, bi-layer silk fibroin (BLSF) biomaterials to serve as superior alternatives to buccal mucosal (BM) grafts for repair of 4 cm long US in female swine.

Methods: Urethral mucosal damage was induced over 4-5 cm long segments via electrocoagulation in adult female swine (N = 10) to promote US over the course of 2-4 weeks. Onlay urethroplasty with BLSF scaffolds or autologous BM grafts (N = 5 per group, ~ 4 cm²) was subsequently performed and animals were maintained for 3 months. Outcome analyses included urethroscopy, retrograde urethrography (RUG), and histological and immunohistochemical (IHC) analyses. Non operated urethral segments served as internal controls (N = 10).

Results: All swine survived the study with no severe complications and exhibited US formation following electrocoagulation with a 43-57% reduction in baseline calibers. At 3 months post-op, imaging modalities revealed both graft cohorts promoted > 80% restoration of native urethral calibers. Histological and IHC evaluations showed BLSF grafts supported the formation of innervated, vascularized urethral-like neotissues with α-smooth muscle actin + and SM22α + smooth muscle bundles as well as pan-cytokeratin + epithelia reminiscent of controls. In contrast, BM grafts primarily retained native oral tissue morphology after urethral transposition exhibiting cytokeratin 1 + stratified, squamous epithelia and scant muscle formation.

Conclusions: BLSF matrices can promote functional restoration of long US via regeneration of native urethral tissues.

Background: Systemic inflammation, often induced by elevated circulating lipopolysaccharide (LPS) levels, is a common consequence of intestinal epithelial barrier damage and microbial translocation. This condition is particularly prevalent in menopausal women, who are at increased risk for chronic inflammation and metabolic syndrome due to physiological changes during menopause. Myo-inositol has been shown to improve metabolic profiles in menopausal women with metabolic syndrome. In this study, we investigated whether small extracellular vesicles (sEVs) from human palatine tonsil-derived mesenchymal stem cells (T-MSCs) can restore circulating myo-inositol levels and promote myo-inositol synthesis in skeletal muscle under repeated LPS exposure, mimicking the intestinal leakage seen in menopausal women.

Methods: Over 2 weeks period, LPS was repeatedly administered to mice, along with a group that also received T-MSC-derived sEVs. After 15 days, the expression of proteins involved in inositol synthesis in skeletal muscle, and serum inositol levels were measured. Additionally, intracellular inositol expression was compared in LPS-treated skeletal muscle cells with and without T-MSC sEVs treatment in vitro. Lastly, the protein and microRNA composition of T-MSC sEVs was analyzed.

Results: Our results demonstrated that T-MSC-derived sEVs significantly increased serum myo-inositol levels and enhanced the expression of myo-inositol synthesis proteins in mice exposed to LPS. Similarly, LPS-treated myotubes supplemented with T-MSC sEVs exhibited restored myo-inositol expression. Moreover, T-MSC sEVs were found to contain high levels of muscle-related proteins.

Conclusion: These findings suggest that T-MSC sEVs may serve as a promising therapeutic strategy for mitigating the effects of intestinal leakage and chronic inflammation in menopausal women. By improving skeletal muscle mass and maintaining myo-inositol levels, T-MSC sEVs offer potential for addressing metabolic disturbances associated with menopause.

Background: The bone lacunar-canalicular system (LCS) is an important microscopic infrastructure for signaling and solute transport in bone tissue, which guarantees the normal physiological processes of bone tissue, and there is a direct relationship between osteoporosis and intrabody mass transfer; however, the mass transfer pattern of the LCS has not yet been clarified under different intensities of in extracorporeal shock waves. The present study aims to assess the effect of extracorporeal shock waves on mass transfer in LCS.

Methods: Sodium fluorescein tracer was taken as the transport substance, and the fluorescence intensities of osteocytes at lacuna in bovine cortical bone were used to indicate the mass transfer effect. The free diffusion and different extracorporeal shock waves were performed in LCS experiments and the fluorescence intensities of the superficial, shallow, middle, and deep layers of osteocytes, which were arranged in a proximity-to-distant order away from the Haversian canal, were detected by laser scanning confocal microscopy.

Results: The results showed that, under different shock waves, the fluorescence intensities of superficial lacunae were the highest in an osteon, followed by shallow and middle layers, and the fluorescence intensities of deep lacunae furthest from the Haversian canal were the lowest, with a decreasing trend and a decreased range of 44.75-97.11%. Relative to free diffusion, the fluorescence intensities of the lacunae in each layer increased by 33.16%, 20.56%, 16.11%, and 26.64% in the superficial, shallow, middle, and deep layers of osteocytes, respectively, under the effect of the extracorporeal shock waves at 1 bar; the fluorescence intensities of the middle layer increased by 100.03% when the intensity was 5 bar, and average fluorescence intensities increased the most with an incremental value of 81.34% in all different shock waves; the fluorescence intensities of the lacunae of each layer was enhanced with a range of 110.93-161.03% by 8 bar.

Conclusion: Extracorporeal shock waves promoted tracer mass transfer within the LCS, and the higher the shock wave magnitudes, the larger the mass transfer in LCS. The transport of solute molecules, nutrients, and signaling molecules within the LCS was facilitated by the extracorporeal shock waves, which may help to address bone diseases such as osteoporosis from the direction of mass transfer in LCS.

Background: Recently, magnetic composite biomaterials have raised attention in bone tissue engineering as the application of dynamic magnetic fields proved to modulate the proliferation and differentiation of several cell types.

Methods: This study presents a novel method to fabricate biofunctional magnetic scaffolds by the deposition of superparamagnetic iron oxide nanoparticles (SPIONs) through thermal Drop-On-Demand inkjet printing on three-dimensional (3D) printed scaffolds. Firstly, 3D scaffolds based on thermoplastic polymeric composed by poly-L-lactic acid/poly-caprolactone/poly(3-hydroxybutyrate-co-3-hydroxyvalerate) were fabricated by Fused Deposition Modelling. Then, in a second step, SPIONs were incorporated onto the surface of the scaffolds by inkjet printing following a designed 2D pattern.

Results: A complete characterization of the resulting magnetic scaffolds was carried out attending to the surface SPIONs deposits, demonstrating the accuracy and versatility of the production technique, as well as the stability under physiological conditions and the magnetic properties. Biological evaluation with human bone marrow mesenchymal stems cells demonstrated biocompatibility of the scaffolds and increased osteogenic capability under the application of a magnetic field, due to the activation of mechanotransduction processes.

Conclusion: These results show that the developed 3D magnetic biofunctional scaffolds can be a very promising tool for advanced and personalised bone regeneration treatments.

Background: Fillers have become a viable treatment option for addressing volume deficits, whether for aesthetic purposes or due to trauma or congenital deformities. While most fillers effectively maintain volume, promoting adipogenesis remains a significant challenge. This study investigated a biomaterial designed to maintain volume both in the short and long term while promoting adipose tissue formation, focusing on the biological properties of a human acellular adipose matrix (AAM) combined with crosslinked hyaluronic acid (HA) and carboxymethyl cellulose (CMC) gels.

Methods: The AAM was prepared through delipidation and decellularization and evaluated for residual fat and cells. To assess its performance, the AAM was compared with conventional collagen scaffolds for the proliferation and adipogenic differentiation of human adipose-derived stem cells(hADSCs) in vitro. An injectable AAM filler was developed by combining AAM with crosslinked HA and CMC gels for the desired rheological properties. Over 12 weeks, the AAM filler, conventional HA filler, and adipose tissue were compared in a nude mice model, assessing volume retention, cell incorporation, and adipogenesis.

Results: The AAM showed effective fat and cell removal and promoted the viability and adipogenic differentiation of hADSCs in vitro. The AAM filler exhibited six times higher viscosity than HA filler. It also outperformed both HA filler and adipose tissue in volume retention and cell incorporation, and new adipose tissue formation.

Conclusions: These results suggest that AAM filler is a promising biomaterial for soft tissue augmentation, particularly in applications requiring volume retention and adipogenesis.