Tianyong Hou

Umbilical cord Wharton's Jelly: a new potential cell source of mesenchymal stromal cells for bone tissue engineering.

Although bone marrow-derived mesenchymal stromal cells (BMSCs) are a main cell source for tissue-engineered bone (TEB), the clinical use of BMSCs is restricted due to the invasive bone marrow aspiration procedure and the decline in available number of mesenchymal stromal cells (MSCs) and differentiation potential with increasing age. Umbilical cord-derived MSCs (UCMSCs) are likely to be a promising alternative cell source for TEB due to their higher availability and potential to proliferate and differentiate. To assess this possibility, we studied bone morphogenetic protein 2 (BMP2)-induced osteogenic differentiation and activation of signaling pathways in UCMSCs and BMSCs. UCMSCs showed a phenotype and differentiation potential similar to that of BMSCs. After 14 days of BMP2 treatment, the overall expression of several osteogenic-specific phenotypes (type I collagen, osteopontin, and osteocalcin) was similar for UCMSCs and BMSCs. The signaling pathway by which BMP2 induced differentiation of both cell types involved the membrane receptor-initiated signals including SMADs, P38, and extracellular regulated kinase. The similar characteristics of BMP2-induced osteogenic differentiation of UCMSCs and BMSCs in vitro would support the use of UCMSCs in TEB.

The effect of mechanical stimulation on the maturation of TDSCs-poly(L-lactide-co-e-caprolactone)/collagen scaffold constructs for tendon tissue engineering

Mechanical stimulation plays an important role in the development and remodeling of tendons. Tendon-derived stem cells (TDSCs) are an attractive cell source for tendon injury and tendon tissue engineering. However, these cells have not yet been fully explored for tendon tissue engineering application, and there is also lack of understanding to the effect of mechanical stimulation on the maturation of TDSCs-scaffold construct for tendon tissue engineering. In this study, we assessed the efficacy of TDSCs in a poly(L-lactide-co-ε-caprolactone)/collagen (P(LLA-CL)/Col) scaffold under mechanical stimulation for tendon tissue engineering both in vitro and in vivo, and evaluated the utility of the transplanted TDSCs-scaffold construct to promote rabbit patellar tendon defect regeneration. TDSCs displayed good proliferation and positive expressed tendon-related extracellular matrix (ECM) genes and proteins under mechanical stimulation in vitro. After implanting into the nude mice, the fluorescence imaging indicated that TDSCs had long-term survival, and the macroscopic evaluation, histology and immunohistochemistry examinations showed high-quality neo-tendon formation under mechanical stimulation in vivo. Furthermore, the histology, immunohistochemistry, collagen content assay and biomechanical testing data indicated that dynamically cultured TDSCs-scaffold construct could significantly contributed to tendon regeneration in a rabbit patellar tendon window defect model. TDSCs have significant potential to be used as seeded cells in the development of tissue-engineered tendons, which can be successfully fabricated through seeding of TDSCs in a P(LLA-CL)/Col scaffold followed by mechanical stimulation.

MicroRNA-24 inhibits osteosarcoma cell proliferation both in vitro and in vivo by targeting LPAATβ.

Lysophosphatidic Acid Acyltransferase β (LPAATβ) may be critically involved in osteosarcoma cell proliferation. However, the comprehensive mechanisms responsible for regulation of LPAATβ in osteosarcoma cells remain unclear. This study found that enhanced LPAATβ expression was correlated with osteosarcoma cell proliferation. MiR-24, targeted to LPAATβ, was down-regulated in osteosarcoma cells. Overexpression of miR-24 down-regulated LPAATβ expression in osteosarcoma cells. Specifically, overexpression of miR-24 inhibited osteosarcoma cell proliferation, however, such effect was blocked when LPAATβ activity was inhibited. In conclusion, our study indicates that miR-24 is reduced in osteosarcoma cells, contributing to up-regulation of LPAATβ and resultant osteosarcoma cell proliferation.

Changing expression profiles of lncRNAs, mRNAs, circRNAs and miRNAs during osteoclastogenesis

Bone is a dynamic organ continuously undergoing shaping, repairing and remodeling. The homeostasis of bone is maintained by the balance between osteoblastic bone formation and osteoclastic bone resorption. Osteoclasts (OCs) are specialized multinucleated cells derived from hematopoietic stem cells (HSCs) or monocytes/macrophage progenitor cells. There are different stages during osteoclastogenesis, and one of the most important steps to form functional osteoclasts is realized by cell-cell fusion. In our study, microarray was performed to detect the expression profiles of lncRNA, mRNA, circRNA and miRNA at different stages during osteoclastogenesis of RAW264.7 cells. Often changed RNAs were selected and clustered among the four groups with Venn analysis. The results revealed that expressions of 518 lncRNAs, 207 mRNAs, 24 circRNAs and 37 miRNAs were often altered at each stage during OC differentiation. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) biological pathway analysis were performed to predict the functions of differentially expressed lncRNAs and co-expressed potential targeting genes. Co-expression networks of lncRNA-mRNA and circRNA-miRNA were constructed based on the correlation analysis between the differentially expressed RNAs. The present study provided a systematic perspective on the potential function of non-coding RNAs (ncRNAs) during osteoclastogenesis.

Effects of Pulsed Electromagnetic Field Frequencies on the Osteogenic Differentiation of Human Mesenchymal Stem Cells

The purpose of this study was to evaluate the effect of different frequencies of pulsed electromagnetic fields on the osteogenic differentiation of human mesenchymal stem cells. Third-generation human mesenchymal stem cells were irradiated with different frequencies of pulsed electromagnetic fields, including 5, 25, 50, 75, 100, and 150 Hz, with a field intensity of 1.1 mT, for 30 minutes per day for 21 days. Changes in human mesenchymal stem cell morphology were observed using phase contrast microscopy. Alkaline phosphatase activity and osteocalcin expression were also determined to evaluate human mesenchymal stem cell osteogenic differentiation.Different effects were observed on human mesenchymal stem cell osteoblast induction following exposure to different pulsed electromagnetic field frequencies. Levels of human mesenchymal stem cell differentiation increased when the pulsed electromagnetic field frequency was increased from 5 hz to 50 hz, but the effect was weaker when the pulsed electromagnetic field frequency was increased from 50 Hz to 150 hz. The most significant effect on human mesenchymal stem cell differentiation was observed at of 50 hz.The results of the current study show that pulsed electromagnetic field frequency is an important factor with regard to the induction of human mesenchymal stem cell differentiation. Furthermore, a pulsed electromagnetic field frequency of 50 Hz was the most effective at inducing human mesenchymal stem cell osteoblast differentiation in vitro.

A composite demineralized bone matrix--self assembling peptide scaffold for enhancing cell and growth factor activity in bone marrow.

The need for suitable bone grafts is high; however, there are limitations to all current graft sources, such as limited availability, the invasive harvest procedure, insufficient osteoinductive properties, poor biocompatibility, ethical problems, and degradation properties. The lack of osteoinductive properties is a common problem. As an allogenic bone graft, demineralized bone matrix (DBM) can overcome issues such as limited sources and comorbidities caused by invasive harvest; however, DBM is not sufficiently osteoinductive. Bone marrow has been known to magnify osteoinductive components for bone reconstruction because it contains osteogenic cells and factors. Mesenchymal stem cells (MSCs) derived from bone marrow are the gold standard for cell seeding in tissue-engineered biomaterials for bone repair, and these cells have demonstrated beneficial effects. However, the associated high cost and the complicated procedures limit the use of tissue-engineered bone constructs. To easily enrich more osteogenic cells and factors to DBM by selective cell retention technology, DBM is modified by a nanoscale self-assembling peptide (SAP) to form a composite DBM/SAP scaffold. By decreasing the pore size and increasing the charge interaction, DBM/SAP scaffolds possess a much higher enriching yield for osteogenic cells and factors compared with DBM alone scaffolds. At the same time, SAP can build a cellular microenvironment for cell adhesion, proliferation, and differentiation that promotes bone reconstruction. As a result, a suitable bone graft fabricated by DBM/SAP scaffolds and bone marrow represents a new strategy and product for bone transplantation in the clinic.

Rapid and accurate detection of RMP- and INH- resistant Mycobacterium tuberculosis in spinal tuberculosis specimens by CapitalBio™ DNA microarray: A prospective validation study

BackgroundDNA microarrays can detect tuberculosis and its multi-drug resistant form in M. tuberculosis isolates and sputum specimens with high sensitivity and specificity. However, no performance data currently exists for its use in spinal tuberculosis specimens. This study was aimed to assess the performance of the CapitalBio™ DNA microarray in the detection of isoniazid (INH) and rifampicin (RMP) resistance in spinal tuberculosis compared with the BACT/MGIT 960 system.MethodsFrom March 2009 to December 2011, 153 consecutive patients from Southwest Hospital, Chongqing with clinically and pathologically diagnosed spinal tuberculosis were enrolled into this study. Specimens collected during surgery from the tuberculosis patients were subjected to M. tuberculosis species identification and drug-resistance detection by the CapitalBio™ DNA microarray, and results were compared with those obtained from the absolute concentration drug susceptibility testing.ResultsThe CapitalBio™ DNA microarray achieved 93.55% sensitivity for the correct M. tuberculosis species identification of the 93 specimens that tested positive for spinal tuberculosis through culture. In addition, twenty-seven additional patients (45.0%) were detected by the DNA microarray to be positive for M. tuberculosis among sixty spinal tuberculosis patients who were culture negative. Moreover, the DNA microarray had a sensitivity of 88.9% and a specificity of 90.7% for RMP resistance, and the microarray had a sensitivity of 80.0% and a specificity of 91.0% for INH resistance. The mean turn-around time of M. tuberculosis species identification and drug resistance detection using the DNA microarray was 5.8 (range, 4–9) hours.ConclusionsThe CapitalBio™ DNA microarray is a feasible and accurate tool for the species identification of M. tuberculosis and for directly detecting RMP and INH resistance from spinal tuberculosis specimens in fewer than 9 hours.

Surgical strategy and management outcomes for adjacent multisegmental spinal tuberculosis: a retrospective study of forty-eight patients.

Study Design. Retrospective study. Objective. To evaluate the clinical outcomes of 4 different procedures for the treatment of adjacent multisegmental spinal tuberculosis (AMSST) and to investigate the selection strategy of the optimal procedure with respect to focal characteristics. Summary of Background Data. Because of the lack of support of the anterior columns of multiple segments, AMSST is thought to confer high risks for the development of kyphosis and paraplegia. However, there are few reports regarding the clinical outcomes of the surgical treatment for AMSST have been investigated. Methods. From August 1999 to June 2010, 48 patients with AMSST were enrolled in this study. Seven patients (A group) underwent a single-stage anterior operation. Eighteen patients (AP group) underwent a single-stage posterior and anterior combined operation. Eighteen patients (P group) underwent a single-stage posterior operation. Five patients (DP group) underwent computed tomography-guided drainage and local chemotherapy combined with a 2-stage posterior operation. The patients were followed up clinically and radiologically for an average of 29.6 months. Results. The cohort consisted of 29 males and 19 females, aged 4 to 54 years. The patients exhibited significant improvements in deformity and neurological deficits. Graft union was achieved in all patients 5 to 12 months postoperatively. Surgery-related complications were noted in 1 and 2 patients in the AP and P groups, respectively. Postoperative recurrence occurred in 1 and 2 patients in the AP and P groups, respectively. All 48 patients had been cured at the final follow-up. Conclusion. This study demonstrated that the 4 procedures can safely and effectively achieve nerve decompression, graft fusion, and kyphosis correction. Individualized procedures should be chosen according to the patients general condition, focal characteristic, type of complication, and surgeons experience. Level of Evidence: 3

Prevascularisation with endothelial progenitor cells improved restoration of the architectural and functional properties of newly formed bone for bone reconstruction

PurposeThe aim of this study was to examine whether the addition of endothelial progenitor cells (EPCs) contributes to restoring the architectural and functional properties of newly formed bone for reconstruction of bone defects.MethodsBone marrow-derived EPCs and mesenchymal stem cells (MSCs) were co-seeded onto demineralized bone matrix (DBM) as a prevascularized tissue-engineered bone (TEB) for the repair of segmental bone defects to evaluate the effects of prevascularization of TEB on ameliorating morphological, haemodynamic and mechanical characteristics.ResultsThe restoration of the intraosseous vasculature and medullary cavity was improved markedly compared to the non-prevascularized groups. The blood supply, biomechanical strength, and bone mineral density of the prevascularized group were significantly higher than those of the non-prevascularized groups during bone reconstruction.ConclusionsThe present study indicates that EPC-dependent prevascularization contributes to bone healing with structural reconstruction and functional recovery and may improve the understanding of correlation between angiogenesis and osteogenesis.

Effects of Initial Cell Density and Hydrodynamic Culture on Osteogenic Activity of Tissue-Engineered Bone Grafts

This study aimed to study the effects of initial cell density and in vitro culture method on the construction of tissue-engineered bone grafts and osteogenic activities. Human mesenchymal stem cells (hMSCs) were seeded onto cubic scaffolds prepared from demineralized bone matrix (DBM) by three methods - static, hydrodynamic, or fibrin hydrogel-assisted seeding. The resulting cell-scaffold constructs were cultured in vitro by static flask culture or hydrodynamic culture. The initial cell density and the subsequent in vitro proliferation and alkaline phosphate activities of the constructs were analyzed. The constructs were also subcutaneously implanted in nude mice to examine their in vivo osteogenic activities. Hydrogel-assisted seeding gave the highest seeding efficiency, followed by hydrodynamic and conventional static seeding. During in vitro culture, hydrodynamic culture produced higher plateau cell densities, alkaline phosphatase (ALP) activities, and extracellular matrix production than static culture. After subcutaneous implantation in nude mice, the implants prepared by the combination of hydrogel-assisted seeding and hydrodynamic culture produced higher wet weight and bone mineral density than implants prepared by other methods. The results suggest that the hydrogel-assisted seeding can substantially increase the initial seed cell density in scaffolds. Subsequent hydrodynamic culture can promote the proliferation and osteoblastic differentiation of the seeded cells. Correspondingly, bone grafts produced by the combination of these two methods achieved the highest osteogenic activity among the three methods employed.