Osteochondral allograft transplantation has a good clinical outcome, however, there is still debate on optimization of allograft storage protocol. Storage temperature and nutrient medium composition are the most critical factors for sustained biological activity of grafts before implantation. In this study, we performed a time-dependent in vitro experiment to investigate the effect of various storage conditions on electromechanical, histological and histochemical properties of articular cartilage. Osteochondral grafts derived from goat femoral condyles were frozen at −70 °C or stored at 4 °C and 37 °C in the medium supplemented with or without insulin-like growth factor-1 (IGF-1). After 14 and 28 days the cartilage samples were quantitatively analysed for electromechanical properties, glycosaminoglycan distribution, histological structure, chondrocyte viability and apoptosis. The results were compared between the experimental groups and correlations among different evaluation methods were determined. Storage at −70 °C and 37 °C significantly deteriorated cartilage electromechanical, histological and histochemical properties. Storage at 4 °C maintained the electromechanical quantitative parameter (QP) and glycosaminoglycan expression near the normal levels for 14 days. Although hypothermic storage revealed reduced chondrocyte viability and increased apoptosis, these parameters were superior compared with the storage at −70 °C and 37 °C. IGF-1 supplementation improved the electromechanical QP, chondrocyte viability and histological properties at 37 °C, but the effect lasted only 14 days. Electromechanical properties correlated with the histological grading score (r = 0.673, p < 0.001), chondrocyte viability (r = −0.654, p < 0.001) and apoptosis (r = 0.416, p < 0.02). In addition, apoptosis correlated with glycosaminoglycan distribution (r = −0.644, p < 0.001) and the histological grading score (r = 0.493, p = 0.006). Our results indicate that quality of allografts is better preserved at currently established 4 °C storage temperature. Storage at −70 °C or at 37 °C is unable to maintain cartilage function and metabolic activity. IGF-1 supplementation at 37 °C can enhance chondrocyte viability and improve electromechanical and histological properties of the cartilage, but the impact persists only 14 days. The correlations between cartilage electromechanical quantitative parameter (QP) and metabolic activity were detected. Our findings indicate that non-destructive assessment of cartilage by Arthro-BST is a simple and reliable method to evaluate allograft quality, and could be routinely used before implantation.

Ozonation has been proved as a viable surface modification technique providing certain properties to the scaffolds that are essential in tissue engineering. However, the ozone (O₃) treatment of PCL scaffolds in aqueous environments has not yet been presented. O₃ treatment performed in aqueous environments is more effective compared with traditional, executed in ambient air treatment due to more abundant production of hydroxyl radicals (•OH) within the O₃ reaction with water molecules. During interaction with •OH, the scaffold acquires functional groups which improve wettability properties and encapsulate growth factors. In this study, a poly(ε)caprolactone (PCL) scaffold was fabricated using solution electrospinning and was subsequently ozonated in a water reactor. The O₃ treatment resulted in the expected occurrence of oxygen-containing functional groups, which improved scaffold wettability by almost 27% and enhanced cell proliferation for up to 14 days. The PCL scaffold was able to withhold 120 min of O₃ treatment, maintaining fibrous morphology and mechanical properties.

Skeletal muscle‐derived stem/progenitor cells (MDSPCs) have been thoroughly investigated and already used in preclinical studies. However, therapeutic potential of MDSPCs isolated using preplate isolation technique for acute kidney injury (AKI) has not been evaluated. We aimed to characterize rat MDSPCs, compare them with bone marrow mesenchymal stem cells (BM‐MSCs), and evaluate the feasibility of MDSPCs therapy for gentamicin‐induced AKI in rats. We have isolated and characterized rat MDSPCs and BM‐MSCs. Characteristics of rat BM‐MSCs and MDSPCs were assessed by population doubling time, flow cytometry, immunofluorescence staining, RT‐PCR, and multipotent differentiation capacity. Gentamicin‐induced AKI model in rat was used to examine MDSPCs therapeutic effect. Physiological and histological kidney parameters were determined. MDSPCs exhibited similar immunophenotype, stem cell gene expression, and multilineage differentiation capacities as BM‐MSCs, but they demonstrated higher proliferation rate. Single intravenous MDSPCs injection accelerated functional and morphological kidney recovery, as reflected by significantly lower serum creatinine levels, renal injury score, higher urinary creatinine, and GFR levels. PKH‐26‐labeled MDSPCs were identified within renal cortex 1 and 2 weeks after cell administration, indicating MDSPCs capacity to migrate and populate renal tissue. In conclusion, MDSPCs are capable of mediating functional and histological kidney recovery and can be considered as potential strategy for AKI treatment.

Polycaprolactone (PCL) is a non-cytotoxic, completely biodegradable biomaterial, ideal for cartilage tissue engineering. Despite drawbacks such as low hydrophilicity and lack of functional groups necessary for incorporating growth factors, it provides a proper environment for different cells, including stem cells. In our study, we aimed to improve properties of scaffolds for better cell adherence and cartilage regeneration. Thus, electrospun PCL–scaffolds were functionalized with ozone and loaded with TGF-β3. Together, human-muscle-derived stem cells (hMDSCs) were isolated and assessed for their phenotype and potential to differentiate into specific lineages. Then, hMDSCs were seeded on ozonated (O) and non-ozonated (“naïve” (NO)) scaffolds with or without protein and submitted for in vitro and in vivo experiments. In vitro studies showed that hMDSC and control cells (human chondrocyte) could be tracked for at least 14 days. We observed better proliferation of hMDSCs in O scaffolds compared to NO scaffolds from day 7 to day 28. Protein analysis revealed slightly higher expression of type II collagen (Coll2) on O scaffolds compared to NO on days 21 and 28. We detected more pronounced formation of glycosaminoglycans in the O scaffolds containing TGF-β3 and hMDSC compared to NO and scaffolds without TGF-β3 in in vivo animal experiments. Coll2-positive extracellular matrix was observed within O and NO scaffolds containing TGF-β3 and hMDSC for up to 8 weeks after implantation. These findings suggest that ozone-treated, TGF-β3-loaded scaffold with hMDSC is a promising tool in neocartilage formation.

To determine the applicability of a minimally invasive diagnostic device to evaluate the quality of articular cartilage following autologous (OAT) and allogeneic (OCA) osteochondral graft transplantation in goat model. OAT grafts were harvested from lateral femoral condyles (LFCs) and transplanted into osteochondral defects created in medial femoral condyles (MFCs) of contralateral knees. OCA grafts were transplanted into MFC condyles after in vitro storage. Autologous platelet-rich plasma (PRP) was administered intraarticularly after the surgery and at 1 and 2 months postoperatively. OAT and OCA grafts were evaluated macroscopically (Oswestry arthroscopy score [OAS]), electromechanically (quantitative parameter, QP), and histologically (O’Driscoll score, safranin O staining intensity) at 3 and 6 months after transplantation. Results were compared with preoperative graft evaluation. Transplanted cartilage deteriorated within 6 months in all groups. Cartilage quality was better retained in OAT group compared with a decline in OCA group. QP and OAS scores were comparable in OAT and OCA groups at 3 months, but superior in OAT group at 6 months, according to all the methods applied. PRP injections significantly improved QP and OAS score at 6 months compared with 3 months in OAT group. QP moderately correlated with OAS, O’Driscoll score, and safranin O staining intensity. Grafts did not retain preoperative quality parameters at 6 months follow-up; however, OAT were superior to OCA grafts. PRP may have a beneficial effect on macroscopic and electromechanical properties of cartilage; however, histological improvement is yet to be proved. Electromechanical diagnostic device enables reliable assessment of transplanted cartilage.