Suppression of DNMT2/3 by proinflammatory cytokines inhibits CtBP1/2-dependent genes to promote the occurrence of atrophic nonunion.

Failure of bone healing after fracture often results in nonunion, but the underlying mechanism of nonunion pathogenesis is poorly understood. Herein, we provide evidence to clarify that the inflammatory microenvironment of atrophic nonunion (AN) mice suppresses the expression levels of DNA methyltransferases 2 (DNMT2) and 3A (DNMT3a), preventing the methylation of CpG islands on the promoters of C-terminal binding protein 1/2 (CtBP1/2) and resulting in their overexpression. Increased CtBP1/2 acts as transcriptional corepressors that, along with histone acetyltransferase p300 and Runt-related transcription factor 2 (Runx2), suppress the expression levels of six genes involved in bone healing: BGLAP (bone gamma-carboxyglutamate protein), ALPL (alkaline phosphatase), SPP1 (secreted phosphoprotein 1), COL1A1 (collagen 1a1), IBSP (integrin binding sialoprotein), and MMP13 (matrix metallopeptidase 13). We also observe a similar phenomenon in osteoblast cells treated with proinflammatory cytokines or treated with a DNMT inhibitor (5-azacytidine). Forced expression of DNMT2/3a or blockage of CtBP1/2 with their inhibitors can reverse the expression levels of BGLAP/ALPL/SPP1/COL1A1/IBSP/MMP13 in the presence of proinflammatory cytokines. Administration of CtBP1/2 inhibitors in fractured mice can prevent the incidence of AN. Thus, we demonstrate that the downregulation of bone healing genes dependent on proinflammatory cytokines/DNMT2/3a/CtBP1/2-p300-Runx2 axis signaling plays a critical role in the pathogenesis of AN. Disruption of this signaling may represent a new therapeutic strategy to prevent AN incidence after bone fracture.

A novel and superior Lasso-plate technique in treatment for coronoid process fracture in the terrible triad of elbow.

The treatment of ulna coronal process fractures in the terrible triad of elbow, especially type I and II Regan-Morrey coronoid fractures, still have been controversial. The purpose of this retrospective study was to evaluate the novel Lasso-plate technique to have a more reliable fixation and a well clinical outcomes for type I and II Regan-Morrey coronoid fractures in a terrible triad of the elbow (TTE). Patients with simple TTE, closed fracture, aged > 18 years, duration of injury < 2 weeks, type I and II Regan-Morrey coronoid process fracture fixed by the Lasso-plate technique or ORIF were enrolled in the study. Total 144 patients with type I and II Regan-Morrey coronoid fracture in TTE were included in the Lasso-plate group or ORIF (open reduction and internal fixation) group in the Xi'an Honghui Hospital from January 2017 to December 2020. Eighty-six patients in Lasso-plate group underwent surgery using a novel Lasso-plate technique. And other 58 patients in ORIF group underwent surgery using ORIF. The data of two groups, including the X-ray films, Computed tomography (CT), the range of elbow motion, Mayo Elbow Performance Score (MEPS) and the surgical complications, were extracted from the hospital's patient records. All patients in both groups were followed up at least 12 months. The mean operation time (88.2 ± 12.3 min) in Lasso-plate group is shorter than that of ORIF group (109.1 ± 13.0 min). There was one patient with injury of deep branch of radial nerve and one patient with superficial surgical incision infection in Lasso-plate group. There were two patients with surgical incision infection in ORIF group. There were three heterotopic ossifications in Lasso-plate group and eight heterotopic ossifications in ORIF group. There were 5 elbow joints stiffness in Lasso-plate group and 12 in ORIF group. At 12 months follow up, the mean range of flexion-extension motion in Lasso-plate group was 122.9° ± 13.4° versus 113.2° ± 18.1° in ORIF group (p < 0.01), the mean 89.7 ± 5.6 MEPS in Lasso-plate group versus mean 83.7 ± 6.1 MEPSin ORIF group. The fixation of coronoid process fracture in TTE by the Lasso-plate technique, especially type I and II Regan-Morrey coronoid fracture, could be easier to master and operate, could provide the sufficient stability of elbow joint to enable early functional exercise, along with a better clinical outcome, a lower surgical complication. For the treatment of TTE, we recommend the fixation of type I and II Regan-Morrey coronoid fracture with the Lasso-plate technique, which would result in a better clinical outcome.

The treatment of a femoral shaft fracture in patients with a previous post-traumatic femoral deformity using a Clamshell osteotomy.

The aim of this study was to assess the safety and clinical outcome of patients with a femoral shaft fracture and a previous complex post-traumatic femoral malunion who were treated with a clamshell osteotomy and fixation with an intramedullary nail (IMN). The study involved a retrospective analysis of 23 patients. All had a previous, operatively managed, femoral shaft fracture with malunion due to hardware failure. They were treated with a clamshell osteotomy between May 2015 and March 2020. The mean age was 42.6 years (26 to 62) and 15 (65.2%) were male. The mean follow-up was 2.3 years (1 to 5). Details from their medical records were analyzed. Clinical outcomes were assessed using the quality of correction of the deformity, functional recovery, the healing time of the fracture, and complications. The mean length of time between the initial injury and surgery was 4.5 years (3 to 10). The mean operating time was 2.8 hours (2.05 to 4.4)), and the mean blood loss was 850 ml (650 to 1,020). Complications occurred in five patients (21.7%): two with wound necrosis, and three with deep vein thrombosis. The mean coronal deformity was significantly corrected from 17.78° (SD 4.62°) preoperatively to 1.35° (SD 1.72°) postoperatively (p < 0.001), and the mean sagittal deformity was significantly corrected from 20.65° (SD 5.88°) preoperatively to 1.61° (SD 1.95°; p < 0.001) postoperatively. The mean leg length discrepancy was significantly corrected from 3.57 cm (SD 1.27) preoperatively to 1.13 cm (SD 0.76) postoperatively (p < 0.001). All fractures healed at a mean of seven months (4 to 12) postoperatively. The mean Lower Extremity Functional Scale score improved significantly from 45.4 (SD 9.1) preoperatively to 66.2 (SD 5.5) postoperatively (p < 0.001). Partial cortical nonunion in the deformed segment occurred in eight patients (34.8%) and healed at a mean of 2.4 years (2 to 3) postoperatively. A clamshell osteotomy combined with IMN fixation in the treatment of patients with a femoral shaft fracture and a previous post-traumatic femoral malunion achieved excellent outcomes. Partial cortical nonunions in the deformed segment also healed satisfactorily.

Kirschner Wire as a Reference Marker for the Positioning of a Syndesmotic Screw: A Radiological Study and Clinical Evaluation.

OBJECTIVE: No consensus has been reached regarding optimal implantation for a syndesmotic screw. Thus, we aimed to explore the feasibility of a reliable and static fibular incisura plane reference for ideal syndesmotic screw placement. METHODS: A retrospective review of computed tomography (CT) scans of 42 uninjured adult ankles with foot fractures were analyzed to measure the tibiofibular vertical distance (TFVD) at 2.5 cm proximal to the plafond from August 2016 to June 2017 in our hospital. The patients (20 females, 22 males) were divided into four groups according to their TFVD: 0-1, 1-2, 2-3, and 3-4 mm, and patients in each group were counted. We retrospectively assessed 41 patients (15 females, 26 males) who underwent syndesmotic screw fixation for ankle fractures from December 2015 to June 2020. We performed t-testing of two independent samples to determine the differences in the angle between the anatomic axis of the syndesmosis and screw axis (AAS) and ankle function using the American Orthopaedic Foot and Ankle Society (AOFAS) score at 3 and 6 months postoperatively between the conventional (20 patients) and K-wire marker (21 patients) groups. The correlation between the AAS and AOFAS score was analyzed. RESULTS: The TFVD measured 2.23 ± 1.01 mm at 2.5 cm proximal to the plafond, and occurred at 25% of the distance from 2 to 3 mm in 47.6% of the patients. This new technique decreased AAS deformation by 62%, from 13.01° ± 2.84° to 4.89° ± 2.43°, in the conventional group (p < 0.001). At 3 months postoperatively, the AOFAS scores of ankle function were similar in both groups, but it was significantly better in the new group than that of conventional group at the 6-month follow-up (p = 0.024). There was a moderate negative correlation between AAS and AOFAS score at 6 months postoperatively (R = -0.684). No obvious complications affecting ankle function were observed in either group postoperatively. CONCLUSIONS: Surgeons can accurately place a screw trajectory using the fibular incisura plane as a reliable intraoperative reference. A 1.6-mm K-wire placed in the syndesmosis at 2.5 cm proximal to the tibial plafond could act as a static marker of the syndesmotic plane.

Flow Cytometric Analysis of Bone Marrow Particle Cells for Measuring Minimal Residual Disease in Multiple Myeloma.

Minimal residual disease (MRD) is one of the most relevant prognostic factors in patients with multiple myeloma (MM). However, the hemodilution of bone marrow (BM) aspirates, the most common preanalytical problem, is known to affect MRD detection. In the present study, we analyzed a preanalytical method for routine BM aspirates and a bone marrow particle cell (BMPL) enrichment assay and validated it as a reliable preanalytical method for flow cytometric MRD determination. A total of 120 BM samples were taken from 103 MM patients consecutively recruited; 77 BM samples had BMPL enrichment analysis and 99 BM samples were routinely analyzed. Then, the two different samples from patients with MM were sent for MRD detection using an eight-color flow cytometry. Our data showed that assessment of the BMPL enrichment samples attenuated the overestimation of MRD-negative assessed in the routine BM samples, which was mainly caused by hemodilution. In conclusion, the BMPL enrichment assay is a functional and practical preanalytical method for flow cytometric MRD analysis.

Clinical and radiographic outcomes of revision with autogenous "structured" bone grafting combined with superior plate for recalcitrant atrophic nonunion of clavicular midshaft: a retrospective study.

PURPOSE: To assess the efficacy of autogenous "structured" bone grafting (ASBG), it was combined with superior plate (SP) revision operations for recalcitrant clavicular midshaft aseptic nonunion (CMAN). METHODS: This retrospective study included 12 patients who suffered from failure of autologous cancellous bone grafting (ACBG) and SP fixation because of CMAN. Visual analogue scale (VAS) data for pain and disabilities of arm, shoulder, and hand (DASH) scores of patients who underwent these procedures from January 2019 to December 2020, obtained before surgery and at the final follow-up time, were analysed. RESULTS: The average time between primitive fracture and this operative treatment was 29 months (15-38 months). The average duration of surgery was 153 minutes (range, 115-230 min), and the average blood loss was 560 ml (range, 350-860 ml). Complications occurred in two cases (16.67%): one was persistent pain at the donor site, and the other was a calf muscle vein thrombosis. No tissue infection was observed during follow-up. The mean follow-up time was 18 months (range, 12-30 months). All fractures progressed to osseous healing at a mean time of 14 weeks (range, 12-16 weeks). The mean pain VAS score significantly improved, from 4.8 ± 1.7 pre-operatively to 1.9 ± 1.1 at the final follow-up (P = 0.01). The mean DASH score improved significantly from 30.1 ± 11.2 pre-operatively to 7.8 ± 4. 2 at the final follow-up (P < 0.01). CONCLUSIONS: ASBG combined with SP revision surgery achieved excellent clinical outcomes in patients with recalcitrant CMAN.

Clinical Application Effects of Different Preoperative Blood Management Schemes in Older Patients with Delayed Intertrochanteric Fracture Surgery.

Introduction: Research on preoperative blood management in older patients with delayed surgery for intertrochanteric fracture is scarce, especially regarding hematopoiesis and hemostasis. We assessed the effectiveness of optimized blood management programs in older patients undergoing delayed surgery for intertrochanteric fractures. Methods: This retrospective study included 456 patients who underwent delayed surgery for intertrochanteric fractures. According to the optimized blood management plan, the patients were divided into four groups: group A was the control group; group B received 1 g of tranexamic acid (TXA) intravenously at admission; group C underwent sequential TXA treatment after admission until 1 day before surgery (1 g/day); and group D received iron supplements (200 mg/day) in addition to the treatment administered to group C, with or without recombinant human erythropoietin (rHuEPO; 40,000 IU). The primary outcomes were preoperative hidden blood loss (HBL), preoperative allogeneic blood transfusion (ABT) rate, hemoglobin (Hb) change, and actual Hb drop. Results: The Hb reduction, calculated HBL, and hospitalization duration in groups C and D were significantly lower than those in groups A and B. The preoperative ABT rates in groups C and D were significantly lower than those in groups A and B, with no significant difference between groups C and D. Discussion: The results of this study suggested that iron supplementation (with or without rHuEPO) combined with the sequential IV TXA scheme did not show a better clinical effect than the sequential IV TXA scheme in the management of patients undergoing delayed surgery for intertrochanteric fractures. Therefore, further evaluation is needed before recommending iron supplements and rHuEPO in older patients.

Self-assembling peptides with hBMP7 biological activity promote the differentiation of ADSCs into nucleus pulposus-like cells.

Functionalized self-assembling peptides, which display functional growth-factor bioactivity, can be designed by connecting the C-terminus of a pure self-assembling peptide with a short functional motif. In this study, we designed a novel functionalized peptide (RADA16-SNVI) in which an SNVI motif with hBMP-7 activity was conjugated onto the C-terminus of the RADA16 peptide via solid-phase synthesis. A mix of RADA16-SNVI and RADA16 solutions was used to create a functionalized peptide nanofiber scaffold (SNVI-RADA16). The hydrogels were analyzed by atomic force microscopy, circular dichroism, and scanning electron microscopy. The results showed that the SNVI-RADA16 solution effectively formed hydrogel. Next, we seeded the SNVI-RADA16 scaffold with adipose-derived stem cells (ADSCs) and investigated whether it displayed biological properties of nucleus pulposus tissue. SNVI-RADA16 displayed good biocompatibility with the ADSCs and induced their expression. Cells in SNVI-RADA16 gel had a greater secretion of the extracellular matrix marker collagen type II and aggrecan compared to ADSCs grown in monolayer and control gel (p < 0.05). The ratio of the aggrecan to collagen in cells in SNVI-RADA16 gel is approximately 29:1 after culture for 21 days. ADSCs in SNVI-RADA16 gels expressed the hypoxia-inducible factor 1α(HIF-1α) mRNA by real-time PCR. However, HIF-1 mRNA is absence in control gel and monolayer. The results suggested that the functionalized self-assembled peptide promotes the differentiation of ADSCs into nucleus pulposus-like cells. Thus, the designed SNVI-RADA16 self-assembling peptide hydrogel scaffolds may be suitable for application in nucleus pulposus tissue regeneration.

An Approach to Intraoperatively Identify the Coronal Plane Deformities of the Distal Tibia When Treating Tibial Fractures with Intramedullary Nail Fixation: a Retrospective Study.

OBJECTIVE: To develop a new approach to intraoperatively identify the presence of coronal plane deformities (both valgus and varus) when treating tibial fractures with closed reduction and intramedullary nail fixation. METHODS: A retrospective analysis was conducted by enrolling 33 consecutive patients with tibial fractures who received closed reduction and intramedullary nail fixation from January 2018 to January 2019 at our trauma center. Out of the 33 patients, 23 were males and 10 were females and the average age was 41 years (ranging 22 to 69 years of age). Standard anteroposterior and lateral preoperative radiographs were routinely performed. After intraoperatively inserting the tibial intramedullary nail through the standard entry point, the parallel relationship between the distal horizontal interlocking screw and the tibiotalar joint surface on the anteroposterior fluoroscopy was used to determine the occurrence of valgus or varus deformities of the distal tibial fragment. Radiographic and clinical outcomes were analyzed using the average interval from injury to surgery, the lateral distal tibial angle (LDTA) of the unaffected and affected sides, complications and the Olerud-Molander ankle score. RESULTS: All 33 patients were postoperatively followed for 13 to 25 months (mean 18.7 months). The fractures achieved bone union at an average of 4.3 months (ranging from 3 to 6 months). The total complication rate was 60.6% (20 cases), including four cases that showed deep vein thrombosis, one case showing an infection and delayed union and 15 cases showing slight to moderate anterior knee pain. The postoperative LDTA of the unaffected side measured 87.3° to 89.6 ° (average 88.7° ± 0.8°), and the LDTA of the affected side was 87.5° to 90.4° (average 88.9° ± 1.1°). There was no significant difference between the unaffected and affected sides (t = -1.865, P = 0.068). The intraoperative measurement indicated six cases of valgus angulation and three cases of varus angulation deformities, and all deformities were corrected during surgery. According to the Olerud-Molander ankle score, clinical outcomes demonstrated 22 excellent cases, eight good cases, two fair cases, and one poor case 12 months after surgery. CONCLUSION: The parallel relationship between the distal horizontal interlocking screw and tibiotalar joint surface on intraoperative anteroposterior films were able to determine the onset of valgus or varus angulations of the distal tibial fragment in the fracture zone after the tibial intramedullary nail was inserted through the standard entry point.

Dose tranexamic acid reduce blood loss associated with simultaneous bilateral distal tibial tubercle-high tibial osteotomy?

BACKGROUND: Simultaneous bilateral distal tibial tubercle high tibial osteotomy (SBDTT-HTO) can result in increased blood loss. The aim of this study is to evaluate the actual hemostatic effect of different tranexamic acid (TXA) treatment regimen in SBDTT-HTO. METHODS: We conducted a retrospective case-control study including 54 patients who underwent SBDTT-HTO. The single-dose group (n = 18) received 1 g of intravenous TXA 15-30 min before surgery, the two-dose group (n = 18) received an additional 1 g of intravenous TXA 6 h after surgery, and the multiple-dose group (n = 18) received an additional 1 g intravenous TXA per-day until discharge. Blood loss, hemoglobin levels, occurrence of any adverse events,functional analysis, quality of life, and pain assessmentswere compared among the three groups. RESULTS: The total blood loss, hidden blood loss, drainage volumes, and haemoglobin level in the multiple-dose group all occupy a significant advantage.(p < 0.05). In addition, better quality of life were observed in patients belonging to the multiple-dose group then single-dose group.(p < 0.05). CONCLUSIONS: Based on our results, for patients undergoing SBDTT-HTO, sequential intravenous TXA administration can effectively and safely reduce blood loss,maintain postoperative Hb levels,and with the advantage of accelerating recovery.

Modified negative-pressure wound therapy for linear blister formation prevention around foam dressings: technical note and case series.

BACKGROUND: Linear blisters (LBs) often occur around dressings when negative-pressure wound therapy (NPWT) is used to cover open wounds. Tension blisters may increase the wound infection incidence rate, delay the start of operation, and prolong the duration of hospital stay. Currently, there are no established methods for the prevention of LB formation around dressings, which remains to be a major concern in clinical applications. Therefore, we developed a novel, simple, reproducible, and convenient method for preventing LB formation around NPWT dressings. METHOD: Fifty-three cases of Gustilo type II and III open fractures under NPWT were considered. NPWT was used on every wound after debridement. All patients were divided into a conventional group (27 cases, 33 wounds) and a novel group (26 cases, 27 wounds) based on the difference in the NPWT dressing appearance. A healthy volunteer with intact skin was also included to perform the detailed process of NPWT. LBs occurring on intact skin around the dressings were observed and recorded when the dressing was removed 3 days after the operation. The occurrence of LB formation and wound infection was considered as categorical data and compared between the two groups using a chi-square test. The duration of hospital stay was considered as numerical data and compared between the two groups using two independent t tests. RESULTS: The percentage of occurrence of LB formation around dressings in the conventional group was 27.3%, whereas it was merely 3.7% in the novel group (P = 0.037). The infection incidence rate in the conventional group was 30.3%, whereas that in the novel group was 25.9%; however, no statistical difference was observed between the two groups (P = 0.708). The average duration of hospital stay in the conventional group was 14.39 ± 4.55 days, whereas that in the novel group was 11.04 ± 3.47 days (P = 0.003). CONCLUSION: Thus, changing the NPWT dressing appearance can prevent LB formation around dressings, providing an effective method to improve NPWT application. Modified NPWT dressings also shorten the duration of hospital stay, but do not significantly decrease the incidence of wound infection.

A New Technique for Placement of Blocking Screws and its Mechanical Effect on Stability of Tibia Fractures with Distal Fragments after Insertion of Small-Diameter Intramedullary Nails.

OBJECTIVE: To design a novel blocking screws (BSs) geometry and insertion method to treat distal tibia fracture with nailing and comparison of mechanical properties of novel and traditional screws. METHODS: Twenty-one synthetic left tibiae were sectioned to obtain 21 distal segments measuring 55 mm. Intramedullary (IM) 9-mm tibial nails were advanced to 6 mm from the ankle joint. Two transverse and one anterior-posterior (AP) locking screws were inserted. Both medial-lateral (ML) BSs were placed 10 mm from the topmost interlocking screw. A custom-made jig assisted in placing the novel and traditional BSs. The time spent in placing each BS was recorded. All the samples were repaired with an IM nail and without BSs, with two traditional BSs, and with two novel BSs. An initial loading from -150 to +150 N was applied to specimens in the ML direction at 185 mm from the nail end, followed by cyclic loading of the same for 10,000 cycles with failure-to-test loading of 350 N in the ML direction. The maximum displacement was measured at 80 mm from the nail end and recorded under initial loading. The damage of two kinds of BSs to the nail was recorded. RESULTS: Compared with average 5.21 min of the time of placing a traditional BS, the time spent in positioning a novel BS on the fracture model was 2.53 min. In the distal bone-implant constructs (BICs), the addition of traditional BSs decreased the maximum displacement of the BICs by 26.2%. The addition of the novel BSs decreased the displacement by 28.9%. All constructs survived 10,000 cycles without hardware deformation. The failure rate of the control group was significantly greater than that of the traditional group; however, the novel group was similar to the traditional group. The damage of the traditional BS to the nail was greater than that of the novel one. CONCLUSIONS: The novel and traditional BSs are comparably effective for increasing the primary mechanical stability of distal metaphyseal fractures after nailin. However, compared to the placement of a traditional BS, implanting a novel BS took more less time and caused less damage to the nail. Additionally, the most obvious advantage of the novel BS design and insertion technology was that the pressure and distance between it and the IM nail could be controlled by rotating the screw. These advantages of the novel BS will be beneficial for clinical application.

Bone marrow particle enrichment analysis for the laboratory diagnosis of multiple myeloma: A case study.

BACKGROUND: Bone marrow smear and biopsy are the main methods for the diagnosis of multiple myeloma (MM), bone marrow infiltration, and metastasis in lymphoma and cancer. However, several factors, including the focal growth of tumor cells, inappropriate puncture sites, and hemodilution of bone marrow aspirates, lower the rate of target cell detection. To solve this problem, we developed a novel method-bone marrow particle enrichment analysis-and here, we describe this procedure and its use in the diagnosis of a rare case of MM. METHODS: An 88-year-old man with primary gastric gamma delta T-cell lymphoma (γδTCL) was found to have anemia. As the cause of anemia could not be determined, hemodilution was suspected, warranting the re-examination of the bone marrow aspirate. Re-puncture could not be performed because of the patient's age and unwillingness to undergo this procedure. Hence, we used a novel approach to enrich bone marrow particles and isolate marrow cells, and subsequently performed morphological and flow cytometric analysis. RESULTS: Examinations performed after bone marrow particle enrichment revealed the presence of myeloma cells, and the patient was diagnosed with primary gastric γδTCL accompanied by MM. CONCLUSIONS: Bone marrow particle enrichment analysis may be applied to overcome the problems caused by hemodilution of bone marrow aspirates and to improve the rate of tumor cell detection. The application of this method for the diagnosis of hematological disorders should be explored further.

Overexpression of p62 is associated with poor prognosis and aggressive phenotypes in osteosarcoma.

p62 (also known as sequestosome 1) protein, is a small regulatory protein that accumulates in autophagy-defective cells that has been demonstrated to be involved in the prognosis and survival of patients with several types of cancer. However, to the best of our knowledge, there have been no such studies for osteosarcoma (OS). In the present study, the expression of p62 in 70 OS samples was determined using immunohistochemistry and its association with various clinicopathological factors was assessed. The results demonstrated that the overexpression of p62 protein was detected in 77.1% (54/70) samples, and the expression levels were significantly associated with tumor size (P=0.001), metastasis (P=0.036), clinical staging (P=0.003) and poor prognosis (P=0.0058). Furthermore, suppression of the p62 expression by short hairpin RNA interference in F5M2 and F4 cells lines led to decreased cell proliferation, migration and invasion in vitro. These results suggested that increased expression of p62 may be involved in OS progression, and therefore the excess expression of p62 may serve as a novel prognostic biomarker for patients with OS.

Cartilage oligomeric matrix protein and hyaluronic acid are sensitive serum biomarkers for early cartilage lesions in the knee joint.

The purpose of this study was to evaluate the relationship between five previously established serum osteoarthritis biomarkers and the severity of cartilage lesions in the knee. Cartilage damage (classified according to the Outerbridge scoring system) and serum concentrations of cartilage oligomeric matrix protein (COMP), collagen type II C-telopeptide (CTX-II), matrix metalloproteinase-3 (MMP-3), collagen type III N-propeptide, (PIIINP), and hyaluronic acid (HA) were determined in 79 patients who underwent knee arthroscopy or total knee replacement. HA and COMP concentrations were significantly higher in the Outerbridge score 1 and 2 groups, respectively. These results suggest that serum COMP and HA concentrations can be used to predict early cartilage lesions in the knee.

Decreased histone deacetylase 4 is associated with human osteoarthritis cartilage degeneration by releasing histone deacetylase 4 inhibition of runt-related transcription factor-2 and increasing osteoarthritis-related genes: a novel mechanism of human osteoarthritis cartilage degeneration.

INTRODUCTION: To investigate if decreased histone deacetylase 4 (HDAC4) is associated with human osteoarthritis (OA) cartilage degeneration by releasing HDAC4 inhibition of runt-related transcription factor-2 (Runx2) resulting in increase of OA cartilage degeneration-related genes. METHODS: The mRNA and protein levels of HDAC4, Runx2, matrix metalloproteinase (MMP)-13, Indian hedgehog (Ihh) and type X collagen were detected by performing real-time PCR (RT-PCR), western blotting and immunohistochemistry on specimens from human OA and normal cartilage. To further explore the mechanism of regulation of Runx2 and OA-related genes by HDAC4, changes in these OA-related genes were further quantified by RT-PCR after overexpression of HDAC4 and knockdown of HDAC4 by siRNA. Runx2 and MMP-13 promoter activities were measured by dual luciferase assays. RESULTS: The levels of HDAC4 in the cartilage from OA patients and healthy 40- to 60-year-old donors were decreased to 31% and 65% compared with specimens from 20- to 40-year-old healthy donors, respectively (P <0.05). Decreased HDAC4 was associated with increased Runx2 and other OA-related genes in human OA cartilage, specifically: MMP-13, Ihh and type X collagen. Exogenous HDAC4 decreased the mRNA levels of Runx2, MMP1, MMP3, MMP-13, type X collagen, Ihh, ADAMTS-4 and -5, and increased the mRNA of type II collagen. In addition, the data also shows that overexpression of HDAC4 not only decreased the expression of interleukin (IL)-1ß, Cox2 and iNos and increased the expression of aggrecan, but also partially blocked the effect of IL-1ß on expression of catabolic events in human OA chondrocytes. HDAC4 also inhibited Runx2 promoter activity and MMP13 promotor activity in a dose-dependent manner. In contrast, inhibition of HDAC4 by TSA drug had an opposite effect. CONCLUSIONS: Our study is the first to demonstrate that decreased HDAC4 contributes, at least in part, to the pathogenesis of OA cartilage degeneration. Thus, HDAC4 may have chondroprotective properties by inhibiting Runx2 and OA-related genes.

Indian hedgehog in synovial fluid is a novel marker for early cartilage lesions in human knee joint.

To determine whether there is a correlation between the concentration of Indian hedgehog (Ihh) in synovial fluid (SF) and the severity of cartilage damage in the human knee joints, the knee cartilages from patients were classified using the Outer-bridge scoring system and graded using the Modified Mankin score. Expression of Ihh in cartilage and SF samples were analyzed with immunohistochemistry (IHC), western blot, and enzyme-linked immunosorbent assay (ELISA). Furthermore, we detected and compared Ihh protein levels in rat and mice cartilages between normal control and surgery-induced osteoarthritis (OA) group by IHC and fluorescence molecular tomography in vivo respectively. Ihh expression was increased 5.2-fold in OA cartilage, 3.1-fold in relative normal OA cartilage, and 1.71-fold in OA SF compared to normal control samples. The concentrations of Ihh in cartilage and SF samples was significantly increased in early-stage OA samples when compared to normal samples (r = 0.556; p < 0.001); however, there were no significant differences between normal samples and late-stage OA samples. Up-regulation of Ihh protein was also an early event in the surgery-induced OA models. Increased Ihh is associated with the severity of OA cartilage damage. Elevated Ihh content in human knee joint synovial fluid correlates with early cartilage lesions.

[Changes of nucleus pulposus after culture of whole intervertebral discs].

OBJECTIVE: To explore a practical method of culturing discs organ system by observing the changes of the nucleus pulposus after the whole intervertebral discs (including cartilage end-plate, nucleus pulposus, and annulus fibrous) were cultivated. METHODS: A total of 335 intervertebral discs were taken out completely from 60 healthy SD rats (about 150 g) aged 5-6 weeks of clear grade and rinsed by high osmotic saline solution containing heparin, then put to the culture plate after being divided into 5 groups randomly. The whole intervertebral discs were cultured with high osmotic (410 mOsmol/kg) culture medium and changed the medium once every day, then the cell viability (n=15), HE staining (n=15), Safranin O staining (n=15), and immunohistochemistry staining (n=2) were observed at 0, 3, 7, 14, and 21 days; RT-PCR result (n=5) was observed at 0, 3, 7, and 14 days. RESULTS: The cell viability was not changed significantly within 14 days (P > 0.05) and was significantly lower at 21 days than at other time points (P < 0.01). The immunohistochemistry staining results for collagen type II were positive in nucleus pulposus cells at every time point. HE staining showed that the tissue integrity and morphology of the whole intervertebral discs were not changed within 14 days. Safranin O staining showed no significant difference in the matrix grey scale within 14 days (P > 0.05) and significant differences between 21 days and 0-14 days (P < 0.05). RT-PCR results showed that the mRNA expression of collagen type I increased with time, but the expressions of collagen type II, aggrecan, and decorin decreased, showing significant differences in the mRNA expressions of the matrix protein at each time point (P < 0.05). CONCLUSION: High osmotic saline solution containing heparin could be used to cultivate the whole intervertebral discs, it is an ideal model for further studies on physiology and pathology of intervertebral discs.