Decellularized Annulus Fibrosus Matrix/Chitosan Hybrid Hydrogels with Basic Fibroblast Growth Factor for Annulus Fibrosus Tissue Engineering.

Low back pain caused by degenerative disc disease affects many people worldwide and brings huge economical burden. Thus, attentions have focused on annulus fibrosus (AF) tissue engineering for treatment of intervertebral disc degeneration. To engineer a functional replacement for the AF, it is important to fabricate scaffolds that mimic the structural and mechanical properties of native tissue. AF-derived stem cells are promising seed cells for AF tissue engineering due to their tissue specificity. In the present study, decellularized AF matrix (DAFM)/chitosan hybrid hydrogels were fabricated using genipin as a crosslinker. AF stem cells were cultured on hydrogel scaffolds with or without basic fibroblast growth factor (bFGF), and cell proliferation, morphology, gene expression, and AF tissue synthesis were examined. Overall, more collagen-I, collagen-II, and aggrecan were secreted by AF stem cells grown on hydrogels with bFGF compared to those without. These results support the application of DAFM/chitosan hybrid hydrogels as an appropriate candidate for AF tissue engineering. Furthermore, incorporation of bFGF into hydrogels promoted AF-related tissue synthesis. Impact Statement The investigation of annulus fibrosus (AF)-related tissue secretion and gene expression in extracellular matrix (ECM) of AF-derived stem cells (AFSCs) provided theoretical and practical basis for the choice of scaffold materials and growth factors for AF tissue engineering. The innovations of the present work are obvious. First, AFSCs were used because they are more easily differentiated into AF cells, thereby producing more AF-related ECM. Second, the decellularized AF matrix (DAFM) was derived from native AF tissue, but had reduced immunogenicity after decellularization. Furthermore, the DAFM structure mimicked the fibrous network of actual AF tissue, which was advantageous to AFSC adhesion and growth. Third, basic fibroblast growth factor was successfully incorporated into the DAFM, showed gradual sustained release, and effectively promoted production of AF tissue ECM factors collagen-I, collagen-II, aggrecan, and glycosaminoglycan.

Isolation and characterization of stem cells from differentially degenerated human lumbar zygapophyseal articular cartilage.

The present study aimed to verify the presence of stem cells with multilineage differentiation potential in human lumbar zygapophyseal articular cartilage (LZAC) and to compare the chondrogenic potential of cells obtained from differentially degenerated articular cartilage samples. Surgically obtained human lumbar zygapophyseal joint tissues were classified into the normal, mildly degenerated and severely degenerated groups, according to their pathological characteristics. Primary chondrocytes from these groups were cultured, and stem cells were selected using a monoclonal cell culture method. Differences in stem cell morphology between the three groups were observed using inverted microscopy and phalloidin staining. In addition, stem cell chondrogenic potential was determined through induced differentiation and cellular staining. Gene and protein expression levels of the chondrogenic­specific markers aggrecan, collagen type­II and SRY­related high­mobility­group box 9 were determined using reverse transcription­quantitative polymerase chain reaction and western blotting. The clonogenic ability of stem cells in the three groups was determined using a clonogenic assay. It was revealed that stem cells with multilineage differentiation potential were isolated from all three cartilage groups; however, the cells obtained from severely degenerated articular cartilage resulted in severe fibrosis, whilst those obtained from mildly degenerated articular cartilage possessed stronger chondrogenic and clonogenic abilities. Taken together, stem cells with multilineage differentiation potential and clonal properties were identified in human LZAC, and these characteristics were more prominent in mildly degenerated as compared with severely degenerated articular cartilage.