Serum-free media for articular chondrocytes in vitro expansion.

BACKGROUND: In vitro chondrocyte expansion is a major challenge in cell-based therapy for human articular cartilage repair. Classical culture conditions usually use animal serum as a medium supplement, which raises a number of undesirable questions. In the present study, two kinds of defined, serum-free media were developed to expand chondrocytes in monolayer culture for the purpose of cartilage tissue engineering. METHODS: Bovine chondrocytes were expanded in serum-free media supplemented with fibroblast growth factor-2 and platelet-derived growth factor or fibroblast growth factor-2 and insulin-like growth factor. Expansion culture in a conventional 10% fetal bovine serum (FBS) medium served as control. Fibronectin coating was used to help cell adhesion in serum-free medium. Next, in vitro three-dimensional pellet culture was used to evaluate the chondrocyte capacity. Cell pellets were expanded in different media to re-express the differentiated phenotype (re-differentiation) and to form cartilaginous tissue. The pellets were assessed by glycosaminoglycans contents, collagen II, collagen I and collagen X immunohistological staining. RESULTS: Chondrocytes cultured in serum-free media showed no proliferation difference than cells grown with 10% FBS medium. In addition, chondrocytes expanded in both serum-free media expressed more differentiated phenotypes at the end of monolayer culture, as indicated by higher gene expression ratios of collagen type II to collagen type I. Pellets derived from chondrocytes cultured in both serum-free media displayed comparable chondrogenic capacities to pellets from cells expanded in 10% FBS medium. CONCLUSION: These findings provide alternative culture approaches for chondrocytes in vitro expansion, which may benefit the clinical use of autologous chondrocytes implantation.

Protective effect of sinomenine on cartilage degradation and chondrocytes apoptosis.

Sinomenine (SIN), an alkaloid extracted from the stem of the Chinese medicinal plant sinomenium acutum, has been used for treating rheumatoid arthritis. But little is known whether SIN has a protective effect on osteoarthritis (OA). In this study, we investigated the protective effect of SIN on IL-1beta-induced proteoglycan degradation and apoptosis in rabbit articular cartilage and chondrocytes. Treatment with 10 ng/ml IL-1beta increased the level of glycosaminoglycan (GAG) released into the culture media, and up-regulated the activity and mRNA expression of matrix metalloproteinase 13 (MMP-13) and down-regulated the activity and mRNA expression of tissue inhibitor of metalloproteinase 1 (TIMP-1) in cartilage explants, as confirmed by the methods of GAG quantitation, MMP-13/TIMP-1 enzyme-linked immunosorbent assay (ELISA) and real-time quantitative RT-PCR. Treatment with 10 ng/ml IL-1beta resulted in marked apoptosis in chondrocytes, as demonstrated by decreased cell viability, occurrence of DNA laddering and increased caspase-3 activity and annexin V binding of phosphatidylserine. However, simultaneous treatment with SIN (10, 50 or 250 microM) inhibited the GAG release and the activity and mRNA expression of MMP-13, and enhanced the activity and mRNA expression of TIMP-1 in a dose-dependent manner in cartilage explants. Furthermore, DNA fragment, caspase-3 activity and apoptosis rate were down-regulated, and cell viability was up-regulated dose-dependently in chondrocytes. Thus, SIN has the protective capacity to antagonize cartilage degradation and chondrocyte apoptosis, which suggest that SIN may act as an agent for pharmacological intervention in the progress of OA.

The protective effect of tetramethylpyrazine on cartilage explants and chondrocytes.

AIMS OF STUDY: Ligusticum wallichi Franchat (chuanxiong) is a very common traditional Chinese herbal medicine in China. Tetramethylpyrazine (TMP) is a major active ingredient extracted from Ligusticum wallichi Franchat. We investigated the protective effect of TMP on interleukin-1ß (IL-1ß) induced proteoglycan (PG) degradation and apoptosis in rabbit articular cartilage and chondrocytes. MATERIALS AND METHODS: Rabbit articular cartilage explants and chondrocytes were cultured with 10 ng/ml IL-1ß for 72 h in the absence or presence of various concentrations of TMP (50, 100 or 200 µM). Cartilage and chondroprotective effects of TMP were determined by evaluating (1) the degree of PG degradation by measuring the amount of glycosaminoglycan (GAG) released into the culture media with 1,9-dimethylmethylene blue (DMMB) assay in cartilage explants; (2) gene expression of MMP-3 and TIMP-1 by real-time quantitative reverse transcription-polymerase chain reaction analysis in cartilage explants; (3) chondrocytes viability with MTT assay; (4) the production of intracellular reactive oxygen species (ROS) with laser scanning confocal microscopy (LSCM). Anti-apoptotic effects of TMP were determined by measuring (1) apoptosis with flow cytometric analysis; (2) mitochondrial membrane potential assay with LSCM; (3) caspase-3 activity with special assay kit. RESULTS: IL-1ß treatment increased the level of GAG released into the culture media, and induced the gene expression of MMP-3 and inhibited the gene expression of TIMP-1 in cartilage explants. Moreover, IL-1ß treatment decreased the cell viability and mitochondrial membrane potential, and enhanced the level of intracellular ROS, apoptosis rate, and caspase-3 activity in chondrocytes. However, simultaneous treatment with TMP attenuated the IL-1ß-induced cartilage and chondrocyte destruction in a dose-dependent manner. TMP showed the decrease of GAG degradation and MMP-3 mRNA production, and the enhancement of TIMP-1 mRNA production in cartilage explants. TMP also increased the cell viability in chondrocytes. Furthermore, TMP inhibited the chondrocytes apoptosis through suppression of ROS production, maintaining of mitochondrial membrane potential and downregulation of caspase-3 activity. CONCLUSION: These results demonstrate that TMP has the cartilage and chondroprotective effect, which suggest that TMP could act as an agent for pharmacological intervention in the progress of OA.

Arthroscopic treatment for osteochondral lesions of the talus: analysis of outcome predictors.

BACKGROUND: Compared with traditional arthrotomy procedures, arthroscopic treatment for osteochondral lesions of the talus has some advantages. However, there has been considerable debate about the outcome predictors for this surgical technique. This study aimed to investigate the outcomes of arthroscopic treatment for osteochondral lesions of the talus, and analyze its outcome predictors. METHODS: Clinical data of 48 patients with osteochondral lesions of the talus who underwent ankle arthroscopy were studied. Arthroscopic debridement was performed on all patients, and microfracture was also performed in 36 cases. Scores on a subjective satisfaction questionnaire, visual analog scale (VAS) for pain, and the American Orthopedic Foot & Ankle Society (AOFAS) ankle and hindfoot scores were obtained before and after surgery. RESULTS: Five patients lost to follow up. The other forty-three patients, 8 of whom were athletes, were followed up for an average of 23.9 months. The average AOFAS post-operative score was 90.16 +/- 9.96, compared with 70.81 +/- 6.96 before surgery (t = 9.353, P < 0.001). The VAS pain score after the operation (2.51 +/- 2.45) was significantly lower than that before the operation (6.95 +/- 1.40) (t = 8.647, P < 0.001). Of the 43 patients, 35 (81.4%) had good or excellent results. There was no significant difference in outcome between the medial and lateral groups (z = 0.205, P = 0.838), while a better outcome was found with lesions smaller than 10 mm than those with larger lesions (z = 2.199, P = 0.028). Age, sex, athletic profession and location of the lesion did not significantly correlate with outcomes. CONCLUSIONS: Arthroscopic treatment is effective and safe for osteochondral lesions of the talus. A strong correlation was found between the size of the lesion and successful outcome.

[Anatomic and clinical study on 4-bone-tunnel double-bundle anterior cruciate ligament].

OBJECTIVE: To conduct a primary clinical analysis on 411 cases of arthroscopic 4 bone-tunnel double-bundle anterior cruciate ligament (ACL) reconstruction done by one operator on the basis of anatomic measurements of cadaver knees. METHODS: Twenty-two adult male Chinese cadaver knees of 25-45 years old were dissected. The sizes, distributions and positions of femoral and tibia footprints of anterior medial bundle (AMB) and posterior lateral bundle (PLB) of ACL and their fiber's orientations were measured and recorded. The anatomy and measurement results were used by one operator in 413 double-bundle ACL reconstruction knees of 411 cases with arthroscopic 4 bone-tunnel technique. Among them, 297 males and 114 females, there were 222 left knees and 191 right knees. The average age was 27.61 +/- 7.23 years old. RESULTS: The anatomy results showed the locations and sizes of AMB and PLB footprints in both femoral and tibia sides were different for different individuals. It was suggested to locate AMB femoral tunnel 7.95 +/- 1.40 mm inferior to 12:00 o'clock position on the posterior edge of the lateral wall of intercondylar fossa with 1 mm thick of the posterior wall of finished AMB femoral tunnel. With the knee flexed to 90 degrees, the PLB tunnel was 5.05 +/- 0.76 mm superior to the lower cartilage edge of the lateral wall of intercondylar fossa, 8.60 +/- 1.52 mm away from the anterior cartilage edge and 8.65 +/- 1.54 mm from the posterior edge. The anatomic orientations of tibia and femoral tunnels were also measured. The study results were used in arthroscopic double-bundle ACL reconstructions of 413 knees. In double-bundle ACL reconstruction operations, the grafts were used in the way from 2-strip double-bundle to 9-strip double-bundle. For most cases, Endo-Button was used for femoral fixation and bio-absorbable interference screw and staple were used for tibia fixation. For the patients adopting autologous semitendinosus and gracilis tendons (STG), the diameter of PLB bone tunnels was 5-6 mm in 95.9% cases, while that of AMB bone tunnel 5-7 mm in 72.4% cases. After a 28-month follow-up in 75 cases, the results showed that double-bundle ACL reconstruction could better reconstruct the stability of knees. CONCLUSION: In 4-bone-tunnel double-bundle arthroscopic ACL reconstruction, both bone tunnel positions and their orientations should be determined according to the anatomic measurement results. Since the PLB diameters of 95.9% cases were 5-6mm, considering the operative outcome of ACL revision, the double bundle ACL reconstruction technique was safe. A better knee stability could be reconstructed by this technique.

Modulating effects of chlorogenic acid on lipids and glucose metabolism and expression of hepatic peroxisome proliferator-activated receptor-alpha in golden hamsters fed on high fat diet.

OBJECTIVE: To examine the effects of chlorogenic acid (CGA) on lipid and glucose metabolism under a high dietary fat burden and to explore the possible role of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) in these effects. METHODS: Twenty male golden hamsters were randomly divided into CGA treatment group (n=10, given peritoneal injection of CGA solution prepared with PBS, 80 mg CGA/kg body weight daily), and control group (n=10, given PBS i.p. at the average volume of the treatment group). Animals in both groups were given 15% high fat diet. Eight weeks after treatment with CGA, the level of biochemical parameters in fasting serum and tissues and the expression of hepatic mRNA and protein PPAR-alpha were determined. RESULTS: Eight weeks after treatment with CGA, the levels of fasting serum triglyceride (TG), free fatty acid (FFA), total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), glucose (FSG), and insulin (FSI) were significantly lower in the GGA treatment group than in the control group. CGA also led to higher activity of hepatic lipase (HL), lower contents of TG and FFA in liver, and lower activity of lipoprotein lipase (LPL) in skeletal muscle. Furthermore, CGA significantly elevated significantly elevated the expression level of mRNA and protein expression in hepatic PPAR-alpha. CONCLUSION: CGA can modify lipids and glucose metabolism, which may be attributed to PPAR-alpha facilitated lipid clearance in liver and improved insulin sensitivity.

An anatomical and histological study of human meniscal horn bony insertions and peri-meniscal attachments as a basis for meniscal transplantation.

BACKGROUND: Allograft meniscal transplantation is an increasingly popular treatment option for the symptomatic young patients with meniscus deficiency. However, many questions still surround it. In this research, we studied the anatomical location and histological structure of human meniscal horn bony insertions and to observe the anatomical morphology and histomorphology of peri-meniscal attachments based on meniscal allograft transplantation. METHODS: Twenty-two fresh-frozen adult cadaver knees were dissected. The locations of meniscal anterior and posterior horn bony insertions to tibia were measured. The anatomical morphology of peri-meniscal attachments was observed and the histological structure of meniscal horn bony insertions and peri-meniscal attachment were studied by HE staining. RESULTS: The anterior horn bony insertion of medial meniscus was (9.19 +/- 1.83) mm inferior to the corresponding anterior border of tibial plateau, and (7.81 +/- 2.25) mm lateral to the axial line of the medial intercondylar eminence. The posterior horn bony insertion of medial meniscus was in the posterior intercondylar fossa of tibia, located between the anterior fibers of the posterior cruciate ligament (PCL) tibial insertion and anterior border of the tibial posterior intercondylar fossa, and was (5.05 +/- 1.18) mm lateral to the axial line of the medial intercondylar eminence. The distance between anterior and posterior horn bony insertions of the lateral meniscus was (13.68 +/- 2.19) mm. Anterior horn bony insertion of the lateral meniscus was (3.99 +/- 1.27) mm medial to the axial line of the lateral intercondylar eminence, and the posterior horn bony insertion of the lateral meniscus was (5.80 +/- 1.36) mm medial to the axial line of the lateral intercondylar eminence. Except for the meniscal horn bony insertions, which is the typical enthesis, we call the attachment of the other parts of menisci as 'peri-meniscal attachment'. The morphological and histological study showed that the main peri-meniscal attachment was the meniscotibial ligament, through which the meniscus attached to the tibia with enthesis structure, and there was only loose connective tissue between menisci and capsule. CONCLUSIONS: In meniscal allograft transplantation, the traditional meniscal size-matching method which take medial and lateral intercondylar eminences as references is not as accurate as expected. Attention should be taken to locate both anterior and posterior horn tunnels of medial meniscal allograft inferior to the tibia plateau, and to locate anterior and posterior horn tunnels of lateral meniscus close enough (mean 13.68 mm). The best way to reconstruct the peri-meniscal attachment is to suture the allograft to the preserved outer remnant of the original meniscus.

Clinical evaluation of double-bundle anterior cruciate ligament reconstruction procedure using hamstring tendon grafts: a prospective, randomized and controlled study.

BACKGROUND: In clinical studies there is still a lot of controversy about the increased anterior and rotational stability between double-bundle (DB) and single-bundle (SB) anterior cruciate ligament (ACL) reconstruction. The aim of this study was to evaluate the clinical results of four-tunnel DB ACL reconstruction. METHODS: Sixty-four consecutive patients with ACL ruptures from May 2005 to May 2006 were randomly assigned into two groups: 32 cases for SB ACL reconstruction and 32 cases for DB ACL reconstruction. Clinical data, including KT 2000, Biodex test, Lysholm score, Tegner score and IKDC score, were prospectively collected until at least 10 months post-operative. RESULTS: The average values of KT 2000 were (1.47 +/- 1.17) mm and (1.68 +/- 1.14) mm for the SB and DB ACL reconstruction groups at 30 degrees of knee flexion (P > 0.05), and were (1.04 +/- 0.98) mm and (1.13 +/- 0.98) mm at 90 degrees of knee flexion (P > 0.05). There were also no significant differences in Lysholm score, Tegner score, IKDC score and Biodex test scores between the two groups (P > 0.05). The operation time of DB ACL reconstruction was 20 minutes longer than the SB ACL reconstruction (P < 0.05). CONCLUSION: Double bundle ACL reconstructions have no obvious clinical advantages over single bundle ACL reconstructions.

MRI signal changes in completely healed meniscus confirmed by second-look arthroscopy after meniscal repair with bioabsorbable arrows.

This study evaluated the MRI signal characteristics and MRI diagnostic accuracy in identifying completely healed menisci repaired with bioabsorbable arrows. A total of 34 patients (38 menisci), with a mean age of 26.0 years, underwent arthroscopic meniscal repair with bioabsorbable arrows and concomitant anterior cruciate ligament (ACL) reconstruction. Of the 34 patients, 27 were male and 7 were female. Of the 38 menisci, 27 were medial and 11 were lateral. Second-look arthroscopy was performed for each patient while taking out the hardware for ACL reconstruction of the tibial side to evaluate the healing status of the repaired menisci. Postoperative MRI was done 2 days before or after second-look arthroscopy. Sagittal T1, T2 and PD images and coronal T2 and PD images were used as the main diagnostic serials. Second-look arthroscopy showed that surfaces of the repaired sites of all 38 menici were almost smooth. In all 38 cases the tail ends of meniscus arrows disappeared and in four patients new overlying injury of compartmental cartilage at the repaired side was detected. MRI results revealed that different serials had different diagnostic accuracy. Sagittal: T1 28.9%, PD 34.2%, T2 60.5%. Coronal: PD 36.8%, T2 65.8%. The double sides Grade 3 signal had a higher proportion in saggital T1 and PD serials, 47.4 and 39.5%, respectively, while lower in sagittal and coronal T2 serials, both 5.3%. MRI diagnostic accuracy was correlated positively with the follow-up time (P < 0.05). MRI has its limitation in evaluating the status of menisci repaired with bioabsorbable arrows, especially for PD and T1 serials. T2 serials have higher diagnostic accuracy than other serials. MRI diagnostic accuracy can be improved by prolonging follow-up time and might be improved by further classifying Grade 3 signal in terms of signal intensity and the shape of the signal margin.

Factors affecting proprioceptive recovery after anterior cruciate ligament reconstruction.

BACKGROUND: Proprioception plays an important role in knee movements. Since there are controversies surrounding the overall recovery time of proprioception following surgery, it is necessary to define the factors affecting proprioceptive recovery after anterior cruciate ligament (ACL) reconstruction and to investigate the relationship between proprioception and muscle strength. METHODS: A total of 36 patients who had their ACL reconstructed with a semitendinosus/gracilis graft (reconstructed group: 6 months post-surgery) and 13 healthy adults without any knee injury (control group) were included in the study. Knee proprioception was evaluated with a passive reproduction test. Isokinetic strength was measured using the Biodex System. Statistical analysis was used to compare proprioception of the reconstructed group versus the control group, and to define causal factors, including sex, hamstring/quadriceps ratio, and the course of injury before reconstruction. We also investigated the correlation between the passive reproduction error and quadriceps index. RESULTS: There was a significant difference in proprioception between the reconstructed and control groups (P < 0.05). When the course of injury before reconstruction was less than 4 months, there was a linear correlation with proprioception 6 months after the operation (r = 0.713, P < 0.05). There was a positive correlation between post-surgery proprioception and the quadriceps index at 6 months post-surgery. CONCLUSIONS: Impaired knee proprioception is observed 6 months after ACL reconstruction. Within 4 months of injury, early undertaking of reconstruction is associated with better proprioception outcome. Patients with enhanced proprioception have a better quadriceps index.

[Arthroscopic treatment of recurrent anterior shoulder dislocation].

OBJECTIVE: To investigate the effect of the arthroscopic procedure on the patients with recurrent anterior shoulder dislocation. METHODS: From January 2001 to March 2006,52 patients with recurrent anterior shoulder dislocation were treated by arthroscopy. Among them 44 patients were followed up for 12 to 54 months(on average of 26 months). The data of the 44 patients was reviewed. Three evaluation systems, University of California at Los Angeles Shoulder Scores (UCLA), Simple Shoulder Test (SST) and Dawson, were used. The study evaluated the effect based on the Dawson system by the factors as: age, course of the disease, frequency of dislocation and relocation methods and the range of shoulder movement. RESULTS: The ratio of recurrent dislocation after operation was 4.5%. Assessing through 3 evaluation systems, UCLA, SST and Dawson, results were similar: the follow-up evaluation were extraordinarily different from preoperative assessment, and the rating of good or excellent at the time of the final follow-up reached 91% higher. Based on the Dawson system, the evaluation results had no statistic diversity according to such factors as: age, course of the disease, frequency of dislocation and relocation methods. CONCLUSIONS: The effects of arthroscopic surgery to the recurrent anterior shoulder dislocation are satisfied and evidence-proved. It is a good option for both common patients and athletes.

Comparative study on early period of recovery between minimally invasive surgery total knee arthroplasty and minimally invasive surgery-quadriceps sparing total knee arthroplasty in Chinese patients.

BACKGROUND: Different kinds of minimally invasive surgery (MIS) procedures have now been used in total knee arthroplasty (TKA). Compared with traditional TKA procedure with a long skin incision, clinical studies showed MIS procedures had some advantages. Quadriceps sparing (QS) procedures are the most minimally invasive MIS procedure until now. This study was aimed to find the insertion types for Chinese patients' vastus medialis and if the QS procedure had some advantages in patients' early recovery. METHODS: Between February 2006 and May 2007, 120 consecutive patients underwent unilateral primary TKA under general anesthesia, among whom 14 patients were lost to follow-up, the remaining 106 cases were enrolled in this study. Among the 106 cases there were 85 right knees, 21 left knees (15 men and 91 women, with a mean age of 65.1+/-7.4 years); osteoarthritis in 97 patients (91.5%) and rheumatoid arthritis in 9 patients (8.5%). MIS TKA was performed in 49 cases (MIS TKA group), while MIS-QS TKA in 57 cases (MIS-QS TKA group). During the operation, the type I, II and III insertions of the vastus medialis for all patients were recorded. Each knee was rated post-operatively according to the Hospital of Special Surgery (HSS) scoring system. Clinical follow-up was undertaken at 1 week, 2, 6, 12 and 24 weeks. Operating time and complications were recorded. RESULTS: There was no statistically significant difference between the two groups for gender distribution, age, left or right knee incidence, pre-operative diagnosis, incidence of varus or valgus deformity. Of the 106 cases there was 1 (0.9%) case with a type I insertion of the vastus medialis, 4 (3.8%) cases with type II insertions, 101 (95.3%) cases with type III insertions. The HSS scoring was significantly different between the MIS-QS TKA group and MIS TKA group within the first two weeks post operation. From 2 weeks later to 24 weeks, no significant difference was found. The average operating time was (53.3+/-12.4) minutes in the MIS TKA group and (64.1+/-15.1) minutes in the MIS-QS TKA group (P<0.001). In the MIS-QS TKA group, 1 patient had delayed healing of the partial skin incision (1.8%). No other complications were found in either group. CONCLUSIONS: Although most of the Chinese patients had type III insertions of the vastus medialis, the MIS-QS TKA procedure showed less injury to the quadriceps than the standard MIS TKA and this could contribute to the earlier recovery of the patients. But a shorter skin incision and more tension on the skin may also lead to more skin complications.

Construction of recombinant adenovirus co-expression vector carrying the human transforming growth factor-beta1 and vascular endothelial growth factor genes and its effect on anterior cruciate ligament fibroblasts.

BACKGROUND: Remodeling of the anterior cruciate ligament (ACL) graft usually takes longer than expected. Gene therapy offers a radical different approach to remodeling of the graft. In this study, the internal ribosome entry site (IRES) sequence was used to construct a new recombinant adenovirus which permits co-expression of transforming growth factor-beta1 (TGFbeta1) and vascular endothelial growth factor 165 (VEGF165) genes (named Ad-VEGF165-IRES-TGFbeta1). We investigated the effects of the new adenovirus on the migration of and matrix synthesis by ACL fibroblasts. METHODS: Adenoviral vector containing TGFbeta1 and VEGF165 genes was constructed. ACL fibroblasts were obtained from New Zealand white rabbits. After ACL fibroblasts were exposed to Ad-VEGF165-IRES-TGFbeta1, the expression of VEGF165 and TGFbeta1 proteins were assessed by enzyme-linked immunosorbent assay (ELISA) and Western blotting analysis. Bioassay of VEGF165 and TGFbeta1 proteins were assessed by Western blotting analysis. Proliferation and migration of ACL fibroblasts were assessed by in vitro wound closure assay. Gene expression of collagen type I, collagen type III, and fibronectin mRNA among matrix markers were assessed by real-time PCR. RESULTS: The results showed the successful construction of a recombinant co-expression adenovirus vector containing TGFbeta1 and VEGF165 genes. Co-expression of TGFbeta1 and VEGF165 can induce relatively rapid and continuous proliferation of ACL fibroblasts and high gene expression of collagen type I, collagen type III, and fibronectin mRNA among matrix markers. CONCLUSION: Co-expression of TGFbeta1 and VEGF165 genes has more powerful and efficient effects on the migration of and matrix synthesis by ACL fibroblasts.

[Diagnosis and arthroscopic treatment of osteochondral lesions of the talus].

OBJECTIVE: To summarize and analyze the diagnosis and arthroscopic treatment of osteochondral lesion of talus (OLT). METHODS: From 2000 to 2005 the data of 34 patients of OLT of the talus were retrospectively studied, including the symptom, physical examination, image, arthroscopic treatment All patients took X-ray and MRI examination before the arthroscopic surgery. Arthroscopic debridement was performed for all patients, in addition to drilling in 5 cases, and microfracture in 18 cases. Before operation, ankle-hindfoot score of American Orthopaedic Foot and Ankle Society (AOFAS) was 71 +/- 8, and the score of pain (visual analogue scale, VAS) was 7.5 +/- 1.3. RESULTS: Weight-bearing pain of the ankle joint aggravated after exercise was the predominant complaint of OLT. X-ray examination was negative in 13 cases, and all lesions were detected by MRI, which was significantly better than X-ray (chi2 = 16.07, P < 0. 001). Thirty-one patients were followed up for an average of 28 months. The average post-operative AOFAS was 91 +/- 9 (t = 9.147, P < 0.001); And VAS was 2.4 +/- 2. 3, which was significantly lower than that in pre-operation (t = 10.853, P < 0.001). Of the 31 patients, 27 (87.1%) had good or excellent results. CONCLUSIONS: MRI could improve the accuracy of diagnosis. The results of arthroscopic treatment for OLT are satisfactory.

Relationship between different skin incisions and the injury of the infrapatellar branch of the saphenous nerve during anterior cruciate ligament reconstruction.

BACKGROUND: After anterior cruciate ligament (ACL) reconstruction, some patients suffered from sensory disturbance around the surgical incision of the leg. This research was aimed to investigate the relationship between the different skin incisions and the injury of the infrapatellar branch of the saphenous nerve (IPBSN) post ACL reconstruction. METHODS: ACL reconstructions were performed with quadruple hamstring tendon for 60 patients. Sensory disturbance around the skin incision was followed up at an average of 14.5 +/- 4.7 months post operation. Among the 60 patients, vertical incision for 35 patients and oblique incision for 25 patients were used for graft taking during ACL reconstruction. The lengths of the incisions were measured. The patients were asked to mark the sensory disturbance zone at follow up time, and then the marked area was measured. The IPBSN of 15 cadaver knees were anatomized. The distance between the IPBSN and the upper edge of the pes anserinus tendon at the middle point of the incision was measured. Independent-samples t-test, chi-square and Mann-Whitney tests were used for statistical analyses. RESULTS: The patients' age (P = 0.329), the follow-up time (P = 0.681), and the incision length (P = 0.732) between the two groups had no significant difference. Twenty-three patients (65.7%) in the vertical incision group had IPBSN injury compared with 6 patients (24.0%) in oblique incision group (P = 0.002). The average sensory disturbance area in vertical incision group ((48.0 +/- 75.3) cm(2)) was significantly larger (P = 0.004) than that in the oblique group ((8.4 +/- 19.4) cm(2)). The anatomy measurement showed the average distance between IPBSN and the upper edge of the pes anserinus tendon was 0.6 cm at the incision. CONCLUSIONS: Oblique incision with less risk of damage for IPBSN may be better for graft harvesting in ACL reconstruction. As the IPBSN is so near and parallel to the hamstring tendons, damage to the IPBSN is one of the potential complications for graft harvesting, regardless of the incision used. That's why even in the oblique incision group, 24% patients also had sensory disturbance complication.

[Posterior and superior compression test and weight-bearing on elbow test for the diagnosis of superior labrum anterior to posterior lesion].

OBJECTIVE: To evaluate the posterior and superior compression test (PSCT) and weight-bearing on elbow test (WBE) on the diagnosis of superior labrum anterior to posterior (SLAP) lesion. METHODS: From July 2000 to March 2007, 4 clinical tests including O'Brien test, Crank test, PSCT and WBE were randomly performed on 207 patients (209 shoulders). Among the patients, 125 were males and 82 were females with the mean age of 39 years. All the patients underwent arthroscopic treatment by the same doctor. The arthroscopic diagnosis was considered as the golden standard. For each test, the parameters on clinical epidemiology were calculated, and the differences of detection rate between the physical tests and the arthroscopic examination were compared. RESULTS: The parameters on clinical epidemiology of O'Brien test, Crank test, PSCT and WBE were comparable. The difference of detection rate between the physical tests and the arthroscopic examination was not statistically significant (P > 0.05). The detection rates of PSCT and WEB in the group of injury of posterior and superior labrum were statistically higher than the group of injury of anterior and superior labrum. The detection rate of PSCT in the group of simple SLAP lesion was relatively higher. CONCLUSIONS: Compared with O'Brien test and Crank test, the new PSCT and WBE have not only comparable clinical value in the diagnosis of SLAP lesion, but also the advantage in predicting the location of SLAP lesion. The detection rate of PSCT in the group of simple SLAP lesion is relatively higher, but the mechanism is to be determined.

Rat adipose-derived stromal cells expressing BMP4 induce ectopic bone formation in vitro and in vivo.

AIM: Bone morphogenetic protein 4 (BMP4) is one of the main local contributing factors in callus formation in the early phase of fracture healing. Adipose-derived stromal cells (ADSC) are multipotent cells. The present study was conducted to investigate the osteogenic potential of ADSC when exposed to adenovirus containing BMP4 cDNA (Ad-BMP4). METHODS: ADSC were harvested from Sprague-Dawley rats. After exposure to Ad-BMP4, ADSC were assessed by alkaline phosphatase activity (ALP) assay, RT-PCR and von Kossa staining. BMP4 expression was assessed by RT-PCR, immunofluorescence and Western blot analysis. ADSC transduced with Ad-BMP4 were directly injected into the hind limb muscles of athymic mice. ADSC Ad-EGFP(enhanced green fluorescence protein) served as controls. All animals were examined by X-ray film and histological analysis. RESULTS: The expression of BMP4 was confirmed at both mRNA and protein levels. The expression of the osteoblastic gene, ALP activity and von Kossa staining confirmed that ADSC transduced with Ad-BMP4 underwent rapid and marked osteoblast differentiation, whereas ADSC transduced with Ad-EGFP and cells left alone displayed no osteogenic differentiation. X-ray and histological examination confirmed new bone formation in athymic mice transplanted with ADSC transduced with Ad-BMP4. CONCLUSION: Our data demonstrated successful osteogenic differentiation of ADSC transduced with Ad-BMP4 in vitro and in vivo. ADSC may be an ideal source of mesenchyme lineage stem cells for gene therapy and tissue engineering.

Suppression of experimental osteoarthritis by adenovirus-mediated double gene transfer.

BACKGROUND: Osteoarthritis (OA) is a chronic and incurable disease, lacking effective treatment. Gene therapy offers a radical different approach to the treatment of arthritis. Even though the etiology of OA remains unclear, there is now considerable evidence to suggest that interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-alpha) are the main mediators in the pathogenesis of OA. The goal of this study was to determine the efficacy of local expression of interleukin-1 receptor antagonist (IL-1Ra) and soluble tumor necrosis factor-alpha receptor type I (sTNF-RI) by direct adenoviral-mediated intra-articular gene delivery in the rabbit model of osteoarthritis. METHODS: Adenoviral vectors containing IL-1Ra or sTNF-RI genes were constructed. OA was induced in both hind knees of 12 New Zealand white rabbits by the excision of the medial collateral ligament plus medial meniscectomy. Five days after surgery, approximately 1 x 10(8) plaque-forming units (pfu) of adenovirus were injected into the joint space of the knee through the patellar tendon. A total of 12 operated rabbits were divided into four groups. Three experimental rabbit groups received 1 x 10(8) pfu of adenovirus encoding either IL-1Ra (3 rabbits), sTNF-RI (3 rabbits) or IL-1Ra and sTNF-RI in combination (3 rabbits), into both knee joints respectively. An inflamed control group of 3 rabbits received approximately 1 x 10(8) pfu of Ad-GFP into both joints. Three days after injection of the adenovirus, both knees of each rabbit were lavaged with 1 ml of saline solution through the patellar tendon. At day 7, the rabbits were sacrificed, and the knees were lavaged, dissected and analyzed for effects of transgene expression. Levels of IL-1Ra and sTNF-RI expression in recovered lavage fluids were measured using a cytokine ELISA kit. Cartilage from the lesion areas of medial femoral condyle and synovium were fixed, embedded, sectioned and stained with hematoxylin and eosin (cartilage and synovium) and toluidine blue (cartilage). The samples were examined by light microscopy and quantitatively evaluated. RESULTS: Intra-articular delivery of IL-1Ra resulted in a significant inhibition of cartilage degradation, but did not affect synovial changes. In contrast, rabbit knee joints receiving sTNF-RI alone showed no detectable reduction in cartilage degradation. However, double gene transfer of IL-1Ra and sTNF-RI resulted in a higher suppression of the cartilage degradation and an observable reduction in synovitis. These data add to and confirm that IL-1Ra has good chondroprotective properties, but TNF-alpha blockade has little effect on joint destruction. CONCLUSION: The enhanced therapeutic effects of both antagonists in combination suggest inhibition of multiple inflammatory cytokines may be more efficacious than blockade of either cytokine alone in treating OA.

Transfection of primary chondrocytes using chitosan-pEGFP nanoparticles.

Chitosan-pEGFP nanoparticles were synthesized through the complex coacervation of the cationic polymer with pEGFP, in order to examine the potential of chitosan as a non-viral gene delivery vector to transfer exogenous gene into primary chondrocytes for the treatment of joint diseases. The nanoparticles were prepared at an N/P ratio of 3.8 and showed a spherical or irregular shape. The mean particle size and zeta potential of the nanoparticles freshly prepared with chitosan of different molecular weight were in the range of 100-300 nm and varied from +1 to +23 mV, respectively. Both the particle size and the zeta potential altered in DMEM of different pH. The transfection of primary chondrocytes was performed in different conditions by varying pH of transfection medium, molecular weight of chitosan and different plasmid dosage. Analysis of FACS demonstrated that the transfection efficiency could reach a much high level and the percentage of positive cells could exceed 50% in certain condition. These results suggest that chitosan-DNA nanoparticles have favorable characteristics for non-viral gene delivery to primary chondrocytes, and have the potential to deliver therapeutic genes directly into joint.

[Biomechanical study of four-bundle semitendinosus tendon autograft used for anterior cruciate ligament reconstruction in rabbits model].

OBJECTIVE: To observe the biomechanical change of the remodeling process of a four bundles model using semitendinosus tendon for the reconstruction of anterior cruciate ligament (ACL) in rabbits, and to explore the possibility of using braiding technique for improving the outcome for reconstruction of ACL. METHODS: Two techniques were employed for the ACL reconstruction, one group with the implant of parallel bundles and the other with the braided bundles. Specimen were collected at 26 and 52 weeks after surgery respectively for biomechanical test. Gross morphology was also observed. RESULTS: The group with the implant of the braided bundles showed a more fused rate. The biomechanical test showed that the strength of the reconstructed ACL with the unbraided bundles was 35.87% of the controlled normal ACL, while the braided bundles reached 67.20%. The strength of the graft of the 26 and 52 weeks with the braided bundles were significantly higher than that of the unbraided group, while the stiffness of the 52 weeks graft of the braided group was significantly higher than that of the unbraided group also. CONCLUSIONS: Braiding each bundle to become a single implant for reconstruction of ACL might have better outcome since it will decrease the movement between each bundles and improve the biomechanical properties.