Background: Nerve injuries are debilitating, leading to long-term motor deficits. Remyelination and axonal growth are supported and enhanced by growth factor and cytokines. Combination of nerve guidance conduits (NGCs) with adipose-tissue-derived multipotent mesenchymal stromal cells (AdMSCs) has been performing promising strategy for nerve regeneration. Methods: 3D-printed polycaprolactone (PCL)-NGCs were fabricated. Wistar rats subjected to critical sciatic nerve damage (12-mm gap) were divided into sham, autograft, PCL (empty NGC), and PCL + MSCs (NGC multi-functionalized with 10⁶ canine AdMSCs embedded in heterologous fibrin biopolymer) groups. In vitro, the cells were characterized and directly stimulated with interferon-gamma to evaluate their neuroregeneration potential. In vivo, the sciatic and tibial functional indices were evaluated for 12 weeks. Gait analysis and nerve conduction velocity were analyzed after 8 and 12 weeks. Morphometric analysis was performed after 8 and 12 weeks following lesion development. Real-time PCR was performed to evaluate the neurotrophic factors BDNF, GDNF, and HGF, and the cytokine and IL-10. Immunohistochemical analysis for the p75^NTR neurotrophic receptor, S100, and neurofilament was performed with the sciatic nerve. Results: The inflammatory environment in vitro have increased the expression of neurotrophins BDNF, GDNF, HGF, and IL-10 in canine AdMSCs. Nerve guidance conduits multi-functionalized with canine AdMSCs embedded in HFB improved functional motor and electrophysiological recovery compared with PCL group after 12 weeks. However, the results were not significantly different than those obtained using autografts. These findings were associated with a shift in the regeneration process towards the formation of myelinated fibers. Increased immunostaining of BDNF, GDNF, and growth factor receptor p75^NTR was associated with the upregulation of BDNF, GDNF, and HGF in the spinal cord of the PCL + MSCs group. A trend demonstrating higher reactivity of Schwann cells and axonal branching in the sciatic nerve was observed, and canine AdMSCs were engrafted at 30 days following repair. Conclusions: 3D-printed NGCs multi-functionalized with canine AdMSCs embedded in heterologous fibrin biopolymer as cell scaffold exerted neuroregenerative effects. Our multimodal approach supports the trophic microenvironment, resulting in a pro-regenerative state after critical sciatic nerve injury in rats.

Crush injuries in peripheral nerves are frequent and induce long-term disability with motor and sensory deficits. Due to axonal and myelin sheath disruptions, strategies for optimized axonal regeneration are needed. Multipotent mesenchymal stromal cells (MSC) are promising because of their anti-inflammatory properties and secretion of neurotrophins. The present study investigated the effect of canine adipose tissue MSC (Ad-MSC) transplantation in an experimental sciatic nerve crush injury. Wistar rats were divided into three groups: sham (n = 8); Crush+PBS (n = 8); Crush+MSC (n = 8). Measurements of sciatic nerve functional index (SFI), muscle mass, and electromyography (EMG) were performed. Canine Ad-MSC showed mesodermal characteristics (CD34-, CD45-, CD44+, CD90+ and CD105+) and multipotentiality due to chondrogenic, adipogenic, and osteogenic differentiation. SFI during weeks 3 and 4 was significantly higher in the Crush+MSC group (p < 0.001). During week 4, the EMG latency in the Crush+MSC groups had better near normality (p < 0.05). The EMG amplitude showed results close to normality during week 4 in the Crush+MSC group (p < 0.04). There were no statistical differences in muscle weight between the groups (p > 0.05), but there was a tendency toward weight gain in the Crush+MSC groups. Better motor functional recovery after crush and perineural canine Ad-MSC transplantation was observed during week 2. This was maintained till week 4. In conclusion, the canine Ad-MSC transplantation showed early pro-regenerative effects between 2–4 weeks in the rat model of sciatic nerve crush injury.

Peripheral nerve injuries (PNIs) by transection require reconstructive surgery, often with highly variable results and persistent sensory and motor deficits. Three-dimensional (3D) printing enables the biofabrication of nerve guidance conduits (NGCs) with the ability to release neurotrophic factors, showing therapeutic potential. We developed a 3D printing process of NGCs using polycaprolactone (PCL) and gelatin methacryloyl (GelMA) integrated with a thermostable fibroblast growth factor 2 (FGF-2). The synthesized GelMA at 10% (w/v) concentration showed superior rheological, mechanical, and ultrastructural characteristics, ensuring 3D printing fidelity. Incorporating FGF-2 into GelMA resulted in a controlled release pattern over 30 days along with biocompatibility and an increase of metabolism in rat S16 Schwann cells and human mesenchymal stem cells (MSCs). MSCs exhibited gene regulation linked to vascularization after FGF-2 stimulation. The PCL polymer facilitated the buildability of a spiral-patterned tubular structure, which was functionalized with a combination of GelMA and UV photocrosslinked. At 12 weeks, following a long-gap nerve injury in rats, NGC implantation enhanced sensory and motor recovery, improved electrophysiological function, and promoted morphological and ultrastructural nerve reorganization and regeneration. At 4 weeks, significant Schwann cell proliferation (S100), expression of the pan-neurotrophin receptor (P75NTR), myelination of newly grown axons, and organization of neurofilaments were observed. The bioactive NGCs represent a promising alternative to nerve autografts for the repair of long-gap injuries.

The route used in the transplantation of mesenchymal stem cells (MSCs) can directly affect the treatment success. The transplantation of MSCs via the intrathecal (IT) route can be an important therapeutic strategy for neurological disorders. The objective of this study was to evaluate the safety and feasibility of the IT transplantation of autologous (Auto-MSCs) and allogeneic (Allo-MSCs) bone marrow mesenchymal stem cells (BM-MSCs) in healthy dogs. Based on neurodisability score, cerebrospinal fluid (CSF) and magnetic resonance imaging (MRI), no significant differences from the control group were observed on day 1 or day 5 after IT Auto- or Allo-MSCs transplantation (P > 0.05). In addition, analysis of matrix metalloproteinase (MMP)-2 and MMP-9 expression in the CSF revealed no significant differences (P > 0.05) at 5 days after IT transplantation in the Auto- or Allo-MSCs group when compared to the control. Intrathecal transplantation of BM-MSCs in dogs provides a safe, easy and minimally invasive route for the use of cell-based therapeutics in central nervous system diseases.

Background: To evaluate tissue regeneration of the urinary bladder after the implantation of a decellularized vein sown with autologous adipose-derived mesenchymal stem cells (ASC) on luminal surfaces. Methods: New Zealand rabbits (n = 10) were distributed in two groups: Group Bioscaffold alone (G1)-decellularized vena cava (1 cm²) was implanted, and Group Bioscaffold plus ACSs (G2)-decellularized vena cava (1 cm²) containing ASCs were implanted. ASCs were expanded, characterized, and maintained for one week in culture with a decellularized vein scaffold. The implants were performed under general anesthesia using a continuous suture pattern. Afterward, 21 d (day) specimens were collected and analyzed by hematoxylin and eosin (HE) histology and scanning electron microscopy (SEM). Results: The integrity of the urinary bladder was maintained in both groups. A superior regenerative process was observed in the G2 group, compared to the G1 group. We observed a greater urothelial epithelialization and maturity of the mucosa and submucosa fibroblasts. Furthermore, SEM demonstrated a notable amount of urothelial villus in the G2 group. Conclusion: Decellularized vena cava scaffolds were able to maintain the integrity of the urinary bladder in the proposed model. In addition, ASCs accelerated the regenerative process development, observed primarily by the new urothelial epithelization and the maturity of mucosa and submucosa fibroblasts.

Peripheral arterial disease (PAD) is the main cause of mortality in the western population and requires surgical intervention with the use of vascular substitutes, such as autologous veins or Dacron or PTFE prostheses. When this is not possible, it progresses to limb amputation. For cases where there is no autologous vascular substitute, tissue engineering with the production of neovessels may be a promising option. Previous experimental studies have shown in vitro that rabbit vena cava can be decellularized and serve as a scaffold for receiving mesenchymal stem cells (MSC), with subsequent differentiation into endothelial cells. The current study aimed to evaluate the behavior of a 3D product structure based on decellularized rabbit inferior vena cava (IVC) scaffolds seeded with adipose-tissue-derived stem cells (ASCs) and implanted in rabbits dorsally subcutaneously. We evaluated the induction of the inflammatory response in the animal. We found that stem cells were positive in reducing the inflammatory response induced by the decellularized scaffolds.