Radiography shows bone formation in DBM/AF constructs that were implanted into the dorsal skin pocket of athymic mice.

<p>Rabbit annulus tissue was dissected and fabricated into a DBM cylinder and the constructs were implanted into the dorsal skin pocket of mice. Specimens were harvested at different time points. X-ray images A) in <i>vitro</i>, and, B) <i>in vivo</i>. Scale bar = 1 mm.</p

Hypertrophic chondrocytes and osteoblasts in punctured intervertebral disc specimens with Safranin-O staining.

<p>The rat intervertebral discs including endplates, NP, and AF tissues were punctured with a needle and implanted into the dorsal skin pocket of athymic mice. The specimens were harvested at 1, 2, 4, and 6 months postoperatively. Tissue sections stained with Safranin-O for disc structure and constituent proteoglycan staining shown in red (A, 4 x magnification; B,10 x magnification). Arrow: hypertrophy chondrocytes. Five sections from four discs were used at each time point.</p

Primer Sequences for real-time PCR.

<p>Primer Sequences for real-time PCR.</p

Chondrogenesis of rabbit annulus fibrosus cells <i>in vitro</i>.

<p>Rabbit annulus fibrosus cells were cultured in a pellet culture system, for three weeks, with chondrogenic medium or control medium (DMEM+1%FBS+1% ITS). A) Gross morphology of cell-pellet. B) GAG in the cell-pellet was measured with the dimethymethylene blue colorimetric assay using chondroitin sulfate as a standard; values are normalized to cell DNA content. C) Expression of collagen II and aggrecan genes are significantly increased in the chondrogenic culture medium compared to control medium. D) Representative images of Safranin-O staining for proteoglycan and immunostaining for type II collagen in the cell-pellet. Scale bar = 500 µm. * p<0.05; ** p<0.01.</p

Immunohistochemitry detection of subcutaneously implanted DBM/AF (A) or AF alone constructs (B).

<p>Separate sections were treated with collagen I, collagen II, collagen X, and osteocalcin primary antibodies and corresponding secondary biotinylated IgG. The signals were developed with 3,3′-diaminobenzidine and photographed on an Axioskop 2 Zeiss microscope. Five sections from four discs were used at each time point. Arrows showing hypertrophy chondrocytes.</p

Hydroxylated Fullerene: A Stellar Nanomedicine to Treat Lumbar Radiculopathy via Antagonizing TNF-α-Induced Ion Channel Activation, Calcium Signaling, and Neuropeptide Production

Current nonsurgical treatments of discogenic lumbar radiculopathy are neither effective nor safe. Our prior studies have suggested that hydroxylated fullerene (fullerol) nanomaterial could attenuate proinflammatory cytokine tumor necrosis factor alpha (TNF-α)-induced neuroinflammation and oxidative stress in mouse dorsal root ganglia (DRG) and primary neurons. Here, we aim to investigate the analgesic effect of fullerol in a clinically relevant lumbar radiculopathy mouse model and to understand its underlying molecular mechanism in mouse DRGs and neurons. Surprisingly, single and local application of fullerol solution (1 μM, 10 μL) was sufficient to alleviate ipsilateral paw pain sensation in mice up to 2 weeks postsurgery. In addition, microCT data suggested fullerol potentially promoted disc height recovery following injury-induced disc herniation. Alcian blue/picrosirius red staining also suggested that fullerol promoted regeneration of extracellular matrix proteins visualized by the presence of abundant newly formed collagen and proteoglycan in herniated discs. For in vitro DRG culture, fullerol attenuated TNF-α-elicited expression of transient receptor potential cation channel subfamily V member 1 (TRPV-1) and neuropeptides release (substance P and calcitonin gene-related peptide). In addition, fullerol suppressed TNF-α-stimulated increase in intracellular Ca²⁺ concentrations in primary neurons. Moreover, Western blot analysis in DRG revealed that fullerol’s beneficial effects against TNF-α might be mediated through protein kinase B (AKT) and extracellular protein-regulated kinase (ERK) pathways. These TNF-α antagonizing and analgesic effects indicated therapeutic potential of fullerol in treating lumbar radiculopathy, providing solid preclinical evidence toward further translational studies