The encapsulation of bismuth as BiOCl/Bi2O3 within ultra-short (ca. 50 nm) single-walled carbon nanocapsules (US-tubes) has been achieved. The Bi@US-tubes have been characterized by high-resolution transmission electron microscopy (HR-TEM), energy-dispersive X-ray spectroscopy (EDS), thermogravimetric analysis (TGA), X-ray photoelectron spectroscopy (XPS), and Raman spectroscopy. Bi@US-tubes have been used for intracellular labeling of pig bone marrow-derived mesenchymal stem cells (MSCs) to show high X-ray contrast in computed tomography (CT) cellular imaging for the first time. The relatively high contrast is achieved with low bismuth loading (2.66% by weight) within the US-tubes and without compromising cell viability. X-ray CT imaging of Bi@US-tubes-labeled MSCs showed a nearly two-fold increase in contrast enhancement when compared to unlabeled MSCs in a 100 kV CT clinical scanner. The CT signal enhancement from the Bi@US-tubes is 500 times greater than polymer-coated Bi2S3 nanoparticles and several-fold that of any clinical iodinated contrast agent (CA) at the same concentration. Our findings suggest that the Bi@US-tubes can be used as a potential new class of X-ray CT agent for stem cell labeling and possibly in vivo tracking.

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Cartilage tissue engineering remains a significant challenge for both researchers and clinicians. Many strategic approaches, such as the delivery of growth factors to an in vitro cultured cartilage construct, continue to receive significant attention. However, the effects of delivering exogenous signaling molecules on endogenous signaling pathways within an engineered tissue are not well understood. In order to address this concern, we have investigated how the delivery of insulin-like growth factor-1 (IGF-1, delivered at concentrations of 0, 10, 50, and 100 ng/mL) affects the endogenous expression of IGF-1, its receptor (IGF-1R), and a well known IGF-1 binding protein (IGFBP-3) by articular chondrocytes embedded in alginate hydrogels over 8 days. To the best of our knowledge, this is the first report of delivery effects upon endogenous signal expression in a three-dimensional system relevant to tissue engineering objectives. Results showed significant differences in mRNA expression of IGF-1, IGF-1R, type II collagen, and type I collagen by day 8 between the induced versus noninduced IGF-1 groups. At day 8, the induced IGF-1 groups expressed IGF-1 mRNA four times lower than the 0 ng/mL IGF-1 group. Further, the IGF-1R mRNA expression was five times higher for the groups exposed to exogenous IGF-1 versus the 0 ng/mL IGF-1 case. Interestingly, the expression of IGFBP-3 decreased for all groups. Type II collagen expression was the highest and type I collagen was the lowest for the IGF-1 delivered samples. Finally, the different concentrations of IGF-1 investigated did not demonstrate significantly different trends in mRNA expression levels. Overall, results indicate that exogenous IGF-1 delivery does affect signaling molecule expression by chondrocytes embedded in alginate hydrogels, particularly downregulating the delivered signal while upregulating its receptor.

The development of an engineered tissue requires a clear understanding of the interactions between the individual components. In this study, we investigated how the addition of hyaluronic acid (HA) to a cartilage tissue engineered scaffold alters chondrocytic expression, and specifically the expression of insulin-like growth factor-1 (IGF-1) signaling molecules. Bovine chondrocytes were embedded (7 million cells/mL) in 2.0% w/v alginate hydrogels containing varying HA concentrations (0, 0.05, 0.50, and 5.00 mg/mL). In vitro constructs were cultured with exogenous IGF-1, and gene expression was monitored at days 1, 4, and 8 for IGF-1, IGF-1 receptor (IGF-1R), IGF binding protein 3 (IGFBP-3), type II collagen and type I collagen. In vivo constructs were precultured for 24 h with exogenous IGF-1 before being implanted subcutaneously in severe combined immunodeficient mice; samples were analyzed using histology at days 7, 14, and 21. Results indicate that, with the addition of high levels (5.00 mg/mL) of HA, IGF-1 can become entrapped within the matrix and therefore interfere with the delivery of IGF-1 to chondrocytes. In vitro and in vivo data showed that increasing the concentration of HA in an alginate hydrogel can decrease chondrocyte IGF-1 expression. IGF-1R expression did not change with HA concentration, and the addition of any HA did not significantly alter IGFBP-3 expression. Chondrocytes continuously expressed phenotypic type II collagen in vitro and in vivo throughout the study for all the groups. However, for all the HA concentrations investigated, chondrocytes showed more of a fibroblastic phenotype, as indicated by greater expression of type I collagen than with no HA, in vitro and in vivo. In conclusion, these results indicate that HA interferes with the delivery of IGF-1 to chondrocytes, affecting the endogenous expression of IGF-1 signaling molecules and the resulting chondrocyte phenotype, and therefore demonstrating the critical effect of biomaterial scaffolds on encapsulated cell function.

In this study, a two part bone tissue engineering scaffold was investigated consisting of a solid poly(propylene fumarate) (PPF) intramedullary rod for mechanical support surrounded by a porous PPF sleeve for osseointegration and delivery of poly(DL-lactic-co-glycolic acid) (PLGA) microspheres with adsorbed recombinant human bone morphogenetic protein-2 (rhBMP-2). Scaffolds were implanted into critical size rat segmental femoral defects with internal fixation for 12 weeks. Bone formation was assessed throughout the study via radiography, and following euthanasia via micro-CT and histology. Mechanical stabilization was evaluated further via torsional testing. Experimental implant groups included the PPF rod alone and the rod with a porous PPF sleeve containing PLGA microspheres with 0, 2, or 8 μg of rhBMP-2 adsorbed onto their surface. Results showed that presence of the scaffold increased mechanical stabilization of the defect, as evidenced by the increased torsional stiffness of the femurs by the presence of a rod compared to the empty defect. Although the presence of a rod decreased bone formation, the presence of a sleeve combined with a low or high dose of rhBMP-2 increased the torsional stiffness to 2.06 ± 0.63 N·mm and 1.68 ± 0.56 N·mm, respectively, from 0.56 ± 0.24 N·mm for the rod alone. The results indicate that, while scaffolds may provide structural support to regenerating tissues and increase their mechanical properties, the presence of scaffolds within defects may hinder overall bone formation if they interfere with cellular processes.