Arthroscopic airbrush assisted cell implantation for cartilage repair in the knee: a controlled laboratory and human cadaveric study.

OBJECTIVE: The objective of this study was to investigate the feasibility of arthroscopic airbrush assisted cartilage repair. METHODS: An airbrush device (Baxter) was used to spray both human expanded osteoarthritic chondrocytes and choncrocytes with their pericellular matrix (chondrons) at 1 × 10(6) cells/ml fibrin glue (Tissucol, Baxter) in vitro. Depth-dependent cell viability was assessed for both methods with confocal microscopy. Constructs were cultured for 21 days to assess matrix production. A controlled human cadaveric study (n = 8) was performed to test the feasibility of the procedure in which defects were filled with either arthroscopic airbrushing or needle extrusion. All knees were subjected to 60 min of continuous passive motion and scored on outline attachment and defect filling. RESULTS: Spraying both chondrocytes and chondrons in fibrin glue resulted in a homogenous cell distribution throughout the scaffold. No difference in viability or matrix production between application methods was found nor between chondrons and chondrocytes. The cadaveric study revealed that airbrushing was highly feasible, and that defect filling through needle extrusion was more difficult to perform based on fibrin glue adhesion and gravity-induced seepage. Defect outline and coverage scores were consistently higher for extrusion, albeit not statistically significant. CONCLUSION: Both chondrons and chondrocytes can be evenly distributed in a sprayed fibrin glue scaffold without affecting viability while supporting matrix production. The airbrush technology is feasible, easier to perform than needle extrusion and allows for reproducible arthroscopic filling of cartilage defects.

Design and fabrication of a hybrid alginate hydrogel/poly(ε-caprolactone) mold for auricular cartilage reconstruction.

The aim of this study was to design and manufacture an easily assembled cartilage implant model for auricular reconstruction. First, the printing accuracy and mechanical properties of 3D-printed poly-ε-caprolactone (PCL) scaffolds with varying porosities were determined to assess overall material properties. Next, the applicability of alginate as cell carrier for the cartilage implant model was determined. Using the optimal outcomes of both experiments (in terms of (bio)mechanical properties, cell survival, neocartilage formation, and printing accuracy), a hybrid auricular implant model was developed. PCL scaffolds with 600 µm distances between strands exhibited the best mechanical properties and most optimal printing quality for further exploration. In alginate, chondrocytes displayed high cell survival (~83% after 21 days) and produced cartilage-like matrix in vitro. Alginate beads cultured in proliferation medium exhibited slightly higher compressive moduli (6 kPa) compared to beads cultured in chondrogenic medium (3.5 kPa, p > .05). The final auricular mold could be printed with 300 µm pores and high fidelity, and the injected chondrocytes survived the culture period of 21 days. The presented hybrid auricular mold appears to be an adequate model for cartilage tissue engineering and may provide a novel approach to auricular cartilage regeneration for facial reconstruction. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1711-1721, 2019.

Saliva-Derived Commensal and Pathogenic Biofilms in a Human Gingiva Model.

In vitro models that closely mimic human host-microbiome interactions can be a powerful screening tool for antimicrobials and will hold great potential for drug validation and discovery. The aim of this study was to develop an organotypic oral mucosa model that could be exposed to in vitro cultured commensal and pathogenic biofilms in a standardized and scalable manner. The oral mucosa model consisted of a tissue-engineered human gingiva equivalent containing a multilayered differentiated gingiva epithelium (keratinocytes) grown on a collagen hydrogel, containing gingiva fibroblasts, which represented the lamina propria. Keratinocyte and fibroblast telomerase reverse transcriptase-immortalized cell lines were used to overcome the limitations of isolating cells from small biopsies when scalable culture experiments were required. The oral biofilms were grown under defined conditions from human saliva to represent 3 distinct phenotypes: commensal, gingivitis, and cariogenic. The in vitro grown biofilms contained physiologic numbers of bacterial species, averaging >70 operational taxonomic units, including 20 differentiating operational taxonomic units. When the biofilms were applied topically to the gingiva equivalents for 24 h, the gingiva epithelium increased its expression of elafin, a protease inhibitor and antimicrobial protein. This increased elafin expression was observed as a response to all 3 biofilm types, commensal as well as pathogenic (gingivitis and cariogenic). Biofilm exposure also increased secretion of the antimicrobial cytokine CCL20 and inflammatory cytokines IL-6, CXCL8, and CCL2 from gingiva equivalents. This inflammatory response was far greater after commensal biofilm exposure than after pathogenic biofilm exposure. These results show that pathogenic oral biofilms have early immune evasion properties as compared with commensal oral biofilms. The novel host-microbiome model provides an ideal tool for future investigations of gingiva responses to commensal and pathogenic biofilms and for testing novel therapeutics.

Long-term vitamin D deficiency in older adult C57BL/6 mice does not affect bone structure, remodeling and mineralization.

Animal models show that vitamin D deficiency may have severe consequences for skeletal health. However, most studies have been performed in young rodents for a relatively short period, while in older adult rodents the effects of long-term vitamin D deficiency on skeletal health have not been extensively studied. Therefore, the first aim of this study was to determine the effects of long-term vitamin D deficiency on bone structure, remodeling and mineralization in bones from older adult mice. The second aim was to determine the effects of long-term vitamin D deficiency on mRNA levels of genes involved in vitamin D metabolism in bones from older adult mice. Ten months old male C57BL/6 mice were fed a diet containing 0.5% calcium, 0.2% phosphate and 0 (n=8) or 1 (n=9) IU vitamin D3/gram for 14 months. At an age of 24 months, mice were sacrificed for histomorphometric and micro-computed tomography (micro-CT) analysis of humeri as well as analysis of CYP27B1, CYP24 and VDR mRNA levels in tibiae and kidneys using RT-qPCR. Plasma samples, obtained at 17 and 24 months of age, were used for measurements of 25-hydroxyvitamin D (25(OH)D) (all samples), phosphate and parathyroid hormone (PTH) (terminal samples) concentrations. At the age of 17 and 24 months, mean plasma 25(OH)D concentrations were below the detection limit (<4nmol/L) in mice receiving vitamin D deficient diets. Plasma phosphate and PTH concentrations did not differ between both groups. Micro-CT and histomorphometric analysis of bone mineral density, structure and remodeling did not reveal differences between control and vitamin D deficient mice. Long-term vitamin D deficiency did also not affect CYP27B1 mRNA levels in tibiae, while CYP24 mRNA levels in tibiae were below the detection threshold in both groups. VDR mRNA levels in tibiae from vitamin D deficient mice were 0.7 fold lower than those in control mice. In conclusion, long-term vitamin D deficiency in older adult C57BL/6 mice, accompanied by normal plasma PTH and phosphate concentrations, does not affect bone structure, remodeling and mineralization. In bone, expression levels of CYP27B1 are also not affected by long-term vitamin D deficiency in older adult C57BL/6 mice. Our results suggest that mice at old age have a low or absent response to vitamin D deficiency probably due to factors such as a decreased bone formation rate or a reduced response of bone cells to 25(OH)D and 1,25(OH)2D. Older adult mice may therefore be less useful for the study of the effects of vitamin D deficiency on bone health in older people.