Zibo Yang

miRNA expression profile during osteogenic differentiation of human adipose-derived stem cells.

Human adipose‐derived stem cells (hADSC) are capable of differentiating into an osteogenic lineage. It is believed that microRNAs (miRNAs) play important roles in regulating this osteogenic differentiation of human adipose‐derived cells, although its molecular mechanism remains unclear. We investigated the miRNA expression profile during osteogenic differentiation of hADSCs, and assessed the roles of involved miRNAs during the osteogenic differentiation. We obtained and cultured human adipose‐derived stems cells from donors who underwent elective liposuction or other abdominal surgery at our institution. miRNA expression profiles pre‐ and post‐osteogenic induction were obtained using microarray essay, and differently expressed miRNAs were verified using quantitative real‐time polymerase chain reaction (qRT‐PCR). The expression of osteogenic proteins was detected using an enzyme‐linked immunosorbent assay. Putative targets of the miRNAs were predicted using online software MiRanda, TargetScan, and miRBase. Eight miRNAs were found differently expressed pre‐ and post‐osteogenic induction, among which four miRNAs (miR‐17, miR‐20a, miR‐20b, and miR‐106a) were up‐regulated and four miRNAs (miR‐31, miR‐125a‐5p, miR‐125b, and miR‐193a) were down‐regulated. qRT‐PCR analysis further confirmed the results. Predicted target genes of the differentially expressed miRNAs based on the overlap from three public prediction algorithms: MiRanda, TargetScan, and miRBase Target have the known functions of regulating stem cell osteogenic differentiation, self‐renewal, signal transduction, and cell cycle control. We identified a group of miRNAs that may play important roles in regulating hADSC cell differentiation toward an osteoblast lineage. Further study of these miRNAs may elucidate the mechanism of hADSC differentiation into adipose tissue, and thus provide basis for tissue engineering. J. Cell. Biochem. 113: 888–898, 2012.

MiR‐193b regulates early chondrogenesis by inhibiting the TGF‐beta2 signaling pathway

Cartilage generation and degradation are regulated by miRNAs. Our previous study has shown altered expression of miR‐193b in chondrogenic human adipose‐derived mesenchymal stem cells (hADSCs). In the current study, we investigated the role of miR‐193b in chondrogenesis and cartilage degradation. Luciferase reporter assays showed that miR‐193b targeted seed sequences of the TGFB2 and TGFBR3 3′‐UTRs. MiR‐193b suppressed the expression of early chondrogenic markers in chondrogenic ATDC5 cells, and TNF‐alpha expression in IL‐1b‐induced PMCs. In conclusion, MiR‐193b may inhibit early chondrogenesis by targeting TGFB2 and TGFBR3, and may regulate inflammation by repressing TNF‐alpha expression in inflamed chondrocytes.

Chondrogenic differentiation of ATDC5 and hMSCs could be induced by a novel scaffold-tricalcium phosphate-collagen-hyaluronan without any exogenous growth factors in vitro.

Application of chondrogenic growth factors is a routine strategy to induce chondrogenesis of hMSCs, but they have economic and safety problems in the long term. It is expected that scaffold material itself could play an important role in chondrogenesis of hMSCs. In this study we tested whether a novel tricalcium phosphate-collagen-hyaluronan scaffold (TCP-COL-HA) had inherent chondro-inductive capacity for chondrogenesis of both ATDC5 and hMSCs without any exogenous growth factors in vitro. hMSCs and ATDC5 were seeded onto TCP-COL-HA scaffolds and cultured in basal medium for 3 weeks to investigate whether the TCP-COL-HA scaffold itself had differentiation-inductive capacity in basal culture. With hMSCs-seeded scaffold in chondrogenic medium (including TGF-β1) as positive control, we then compared the chondrogenic induction of TCP-COL-HA in basal culture and in chondrogenic culture. The chondrogenic differentiation was evaluated by sulfated glycosaminoglycans (GAGs) quantification, type II collagen immunohistochemistry, and RT-PCR. Mechanical strength was evaluated by compression test and the cell death rate of hMSCs was assessed with TUNEL assay. The results showed TCP-COL-HA scaffold itself could efficiently induce chondrogenic differentiation of both ATDC5 and hMSCs after 3 weeks in basal culture. The accumulation of GAGs and the expression of chondrocyte marker genes were all significantly increased. In addition, hMSCs-seeded scaffold showed a significantly higher mechanical strength after 3 weeks in basal culture. The chondrogenic induction of TCP-COL-HA scaffolds in basal medium were almost similar to that in chondrogenic medium on hMSCs. The chondrogenesis-inducing capacity of TCP-COL-HA scaffold might help to improve cartilage tissue engineering with economic and safe benefits.

The biomechanical differences of rotational acetabular osteotomy, Chiari osteotomy and shelf procedure in developmental dysplasia of hip

BackgroundRotational acetabular osteotomy (RAO), Chiari osteotomy and shelf procedure are important treatments to delay the progression of osteoarthritis in developmental dysplasia of hip (DDH) patients, but their biomechanical differences are still unknown. This study was to evaluate the different biomechanical changes of hip joint after these three surgeries.MethodsSixteen DDH models of 8 human cadaver specimens were reconstructed, and treated by different surgeries, and then strain around femoral head was evaluated by strain gauges.ResultsHip strain value of DDH model was decreased after treated by shelf procedure (Pleft = 0.016 and Pright = 0.021) and rotational acetabular osteotomy (P = 0.004), but not in Chiari osteotomy (P = 0.856). Moreover, the improved ratio of RAO treatment was better than shelf procedure (P = 0.015) and Chiari osteotomy (P = 0.0007), and the descendent range of shelf procedure was greater than Chiari osteotomy (P = 0.018).ConclusionsFrom biomechanics points, RAO was more effective in relieving hip joint stress compared with shelf procedure and Chiari osteotomy.

CCL3 serves as a potential plasma biomarker in knee degeneration (osteoarthritis)

OBJECTIVE To explore the ability of chemokines in plasma to detect the presence of pre-X-rays defined knee degeneration and the extent (burden). METHODS A total of 181 subjects (75 control subjects, 47 pre-X-KD patients and 50 X-KOA patients) were included and subdivided into three subgroups. Articular cartilage loss in pre-X-KD patients were scored on the basis of the ICRS classification during the arthroscopy or documented on MRI with chondral WORMS. The severity of X-KOA was graded using the Kellgren-Lawrence classification through the posterior-anterior knee X-rays. The concentrations of the inflammatory cytokines and chemokines in plasma were quantified using Luminex microbead-based suspension array (SA) and were cross-validated by enzyme-linked immunosorbent assay (ELISA). RESULTS CCL3 in plasma showed the highest ability to discriminate pre-X-KD patients from the controls with an AUC of 0.799. At a cutoff value of 0.168 pg/ml, the sensitivity was 70.21%, the specificity was 96.00%, the positive predictive value was 91.67% and the negative predictive value was 83.72%. As to define disease burden, the plasma levels of resistin, IL6, IL8, CCL3 and CCL4 showed significant association with the severity of X-rays defined knee OA, with regard to the KL classification. Moreover, significant elevation of IL6, IL8, CCL3 and CCL4 levels in plasma were observed in severe knee OA patients (KL grade IV) compared with those with pre-X-KD (KL grade 0-I). CONCLUSION We firstly showed that the plasma CCL3 could be potential serum biomarker for knee OA with the capacity to detect pre-X-rays defined changes and stage the severity of damage in knee.

Roles of Sagittal Anatomical Parameters of the Pelvis in Primary Total Hip Replacement for Patients with Ankylosing Spondylitis

We examined the correlation between acetabular prostheses and sagittal anatomical parameters of the pelvis for the preoperative evaluation of total hip arthroplasty in 29 patients with ankylosing spondylitis between April 2004 and November 2011. No implant dislocation or subsidence was observed at 4.18 years. The relationship between sagittal parameters conformed to the equation Pelvic incidence (PI)=Pelvic tilt (PT)+Sacral slope (SS). Better outcomes were achieved in the SS>PT group, postoperative function was positively correlated with SS/PI. Functional abduction and anteversion were positively correlated with PT but negatively correlated with SS. Due to the compensatory changes in the pelvis and spine of patients with AS, the preoperative assessment of sagittal parameters plays pivotal roles in placing acetabular prostheses in optimal positions and preventing postoperative impingement and dislocation.

Effects of IL-1 Receptor-Associated Kinase-4 Gene Silencing on Human Osteoblast-Like Cells

The aim of this study is to identify the effects of interleukin-1 receptor-associated kinase-4 (IRAK-4) gene silencing on human osteoblast-like cells. The siRNA sequences of the target gene, IRAK-4, were constructed and transferred into MG63 cells (control group = MG63 cells; SC group = MG63 cells transfected with scrambled IRAK-4 siRNA; KD group = MG63 cells transfected with 75 nM IRAK-4 siRNA). The morphological changes, cell growth, cell-cycle progression, apoptosis, and the expression of various cytokines and proteins were compared. Compared with the control and SC groups, IRAK-4 gene silencing in MG63 cells caused morphological changes, inhibited growth, altered the cell-cycle distribution, increased apoptosis (p < 0.05), decreased bone alkaline phosphatase and osteocalcin levels (p < 0.05), and decreased protein expression of Bcl-2/Bax and Bcl-2, p-JNK1/2, p-ERK1/2, and p-p38MAPK (p < 0.05). The results indicated that IRAK-4 gene silencing in MG63 cells inhibited cell proliferation and function and increase apoptosis, which may be related to the decreased Bcl-2/Bax ratio and inhibition of the protein expression of various components of the mitogen-activated protein kinase pathways. The results of this study may help improve the understanding of the relationship between IRAK-4 and osteoblast-like cells and the interactions between various cytokines in the periprosthetic inflammatory environment.

MicroRNA-193b-3p regulates chondrogenesis and chondrocyte metabolism by targeting HDAC3

Histone deacetylase 3 (HDAC3) plays a pivotal role in the repression of cartilage-specific gene expression in human chondrocytes. The aim of this study was to determine whether microRNA-193b-3p (miR-193b-3p) regulates the expression of HDAC3 during chondrogenesis and chondrocyte metabolism. Methods: miR-193b-3p expression was assessed in a human mesenchymal stem cell (hMSC) model of chondrogenesis, in interleukin-1β (IL-1β)-treated primary human chondrocytes (PHCs), and in non-degraded and degraded cartilage. hMSCs and PHCs were transfected with miR-193b-3p or its antisense inhibitor. A direct interaction between miR-193b-3p and its putative binding site in the 3′-untranslated region (3′-UTR) of HDAC3 mRNA was confirmed by performing luciferase reporter assays. Chondrocytes were transfected with miR-193b-3p before performing a chromatin immunoprecipitation assay with an anti-acetylated histone H3 antibody. To investigate miR-193b-3p-transfected PHCs in vivo, they were seeded in tricalcium phosphate-collagen-hyaluronate (TCP-COL-HA) scaffolds, which were then implanted in nude mice. In addition, plasma exosomal miR-193b-3p in samples from normal controls and patients with osteoarthritis (OA) were measured. Results: miR-193b-3p expression was elevated in chondrogenic and hypertrophic hMSCs, while expression was significantly reduced in degraded cartilage compared to non-degraded cartilage. In addition, miR-193b-3p suppressed the activity of reporter constructs containing the 3′-UTR of HDAC3, inhibited HDAC3 expression, and promoted histone H3 acetylation in the COL2A1, AGGRECAN, COMP, and SOX9 promoters. Treatment with the HDAC inhibitor trichostatin A (TSA) increased cartilage-specific gene expression and enhanced hMSCs chondrogenesis. TSA also increased AGGRECAN expression and decreased MMP13 expression in IL-1β-treated PHCs. Further, 8 weeks after implanting PHC-seeded TCP-COL-HA scaffolds subcutaneously in nude mice, we found that miR-193b overexpression strongly enhanced in vivo cartilage formation compared to that found under control conditions. We also found that patients with OA had lower plasma exosomal miR-193b levels than control subjects. Conclusions: These findings indicate that miR-193b-3p directly targets HDAC3, promotes H3 acetylation, and regulates hMSC chondrogenesis and metabolism in PHCs.

Better Prognosis of Senile Patients with Intertrochanteric Femoral Fracture by Treatment with Open Reduction Internal Fixation than by Hip Arthroplasty

ABSTRACT Purpose: To compare the postoperative survival and mortality rates in intertrochanteric femoral fracture (IFF) patients who underwent either open reduction internal fixation (ORIF) or hip arthroplasty. Methods: Clinical data from senior patients who had IFF and underwent ORIF or hip arthroplasty were analyzed retrospectively. Survival curves were compared between groups with Kaplan–Meier method and log-rank test. Significant independent prognostic factors were identified by Cox multivariate regression analysis. Results: All patients recovered fully post-surgery. Although 31 patients died during the follow-up period (ORIF, mean 45.4 months; arthroplasty, mean 51.6 months), mortality rate did not differ significantly between the groups. The 1-yr and 2-yr survival rate estimates for the ORIF group were 92.2%, and 86%, respectively; they were 85% and 74% for the arthroplasty group. Average survival lengths for ORIF and arthroplasty groups were 88 and 67 months, respectively. The effect of surgical approaches on survival differed significantly (log-rank test c2 = 6.402, p = 0.011). Multivariate Cox regression model indicated that surgical choice (p = 0.036) was a significant independent risk factor for the prognosis of senile IFF, even with adjustment for age (p = 0.002). Conclusion: The overall postoperative prognosis was superior in senile IFF patients treated with ORIF.

Preoperative Measurement of Tibial Resection in Total Knee Arthroplasty Improves Accuracy of Postoperative Limb Alignment Restoration

Background:Accuracy of implant placement in total knee arthroplasty (TKA) is crucial. Traditional extramedullary alignment instruments are fairly effective for achieving the desired mean tibial component coronal alignment. We modified the traditional tibial plateau resection technique and evaluated its effect on alignment restoration. Methods:Two hundred and eighty-two primary TKAs in our hospital between January 2013 and December 2014 were enrolled in this retrospective study. Group A consisted of 128 primary TKAs performed by one senior surgeon. Preoperative measurement of the tibial resection was conducted on radiographs, and the measured thicknesses of the lateral and medial plateau resection were used to place the tibial alignment guide. Group B consisted of 154 primary TKAs performed by the other senior surgeon, using a traditional tibial plateau resection technique. In all patients, an extramedullary guide was used for tibial resection, and preoperative and postoperative full-leg standing radiographs were used to assess the hip-knee-ankle angle (HKA), femoral component alignment angle (FA), and tibial component alignment angle (TA). A deviation ≥3° was considered unsatisfactory. Data were analyzed by unpaired Students t-test. Results:The mean postoperative HKA and TA angles were significantly different between Groups A and B (178.2 ± 3.2° vs. 177.0 ± 3.0°, t = 2.54, P = 0.01; 89.3 ± 1.8° vs. 88.3 ± 2.0°, t = 3.75, P = 0.00, respectively). The mean postoperative FA was 88.9 ± 2.5° in Group A and 88.9 ± 2.6° in Group B, and no significant difference was detected (t = 0.10, P = 0.92). There were 90 (70.3%) limbs with restoration of the mechanical axis to within 3° of neutral alignment and 38 (29.7%) outliers (>3° deviation) in Group A, whereas there were 89 (57.8%) limbs with restoration of the mechanical axis to within 3° of neutral alignment and 65 (42.2%) outliers (>3° deviation) in Group B. The severity of the preoperative alignment deformity was a strong predictor for postoperative alignment. Conclusions:Using conventional surgical instruments, preoperative measurement of resection thickness of the tibial plateau on radiographs could improve the accuracy of conventional surgical techniques.