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Objective:To investigate the differences in gene expression levels in knee chondrocytes and chondrons in vitro.Methods:The chondrocytes and chondrons were isolated from full thickness of the 8-months ( n=5) rabbit knees cartilage. Chondrons from right knee were enzymatically isolated using 0.3% dispase and 0.2% collagenase-2 with shaking for 3 hours. Chondrocytes were isolated by 0.4% Pronase and 0.025% collagenase-2 from left knee. The mRNA levels in chondrocytes and chondrons were analyzed by quantitative real-time PCR, including matrix proteins [aggrecan(Agg), collagen(Col-2), Col-6A6, Col-10, Col-11], MMPs and inhibitors (MMP-1, MMP-3, MMP-9, MMP-13, TIMP-1, TIMP-2, TIMP-3), cytoskeletal proteins (Sox-9, vinculin, tubulin, actin), cytokines (IL-β, TNF-α). Analysis was performed using SPSS 16.0 statistical software, and the two-group comparisons were considered as significant by t-test at P<0.05. Results:Compared to the chondrocytes, the Agg [(5.78±0.90) vs (1.89±0.27), t=9.26, P<0.001], Col-2 [(6.29±0.76) vs (3.06±0.60), t=7.46, P<0.001], Col-6A6 [(0.89±0.18) vs (0.22±0.06), t=7.90, P<0.001], Col-10 [(3.83±0.76) vs (1.00±0.26), t=7.88, P<0.001] and TIMP-1 [(1.98±0.85) vs (1.03±0.34), t=2.32, P=0.049], TIMP-2[(3.46±1.50) vs (1.52±1.06), t=2.36, P=0.046], TIMP-3 [(3.96±0.50) vs (1.36±0.18), t=10.94, P<0.001], Sox-9 [(7.09±2.93) vs (3.24±0.77), t=2.84, P=0.022], vinculin [(3.42±1.69) vs (1.46±0.68), t=2.41, P=0.043], tubulin[(9.34±0.71) vs (2.35±0.80), t=14.61, P<0.001] showed higher expression in the chondrons. Compared to the chondrocytes, the MMP-1 [(1.02±0.30) vs (2.67±0.45), t=6.91, P<0.001], MMP-3 [(1.21±0.32) vs (2.52±0.79), t=3.44, P=0.009], MMP-13 [(1.23±0.34) vs (3.42±0.86), t=5.30, P=0.007], IL-1β [(1.02±0.14) vs (2.70±0.49), t=7.37, P<0.001], TNF-α [(0.99±0.08) vs (3.15±0.54), t=8.85, P<0.001] showed lower expression in the chondrons. There were no difference between chondrons and chondrocytes for Col-11, MMP-9, actin ( P>0.05). Conclusion:The gene expression of extracellular matrix components are higher and the gene expression levels of inflammatory factors and MMPs are decreased in chondrons compared with the chondrocytes, suggesting the chondrons have more multiplication potential as seeding cells for tissue-engineered cartilage.
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Objective:By studying the gait changes of patients with knee osteoarthritis (KOA), the study provided a theoretical basis for the quantitative indicators obtained from gait analysis to the diagnosis of KOA. And it provided a gait reference for the clinical diagnosis, treatment, rehabilitation, prevention and efficacy evaluation of KOA.Methods:A total of 30 patients (KOA group) with KOA hospitalized in our hospital from May 2021 to October 2021 and 30 healthy people (control group) were compared for gait changes. The t test, Mann-Whitney U test and Fisher′s exact test were used to compare differences between groups. Results:The KOA group were greater than the control group in terms of step time [(642±81) ms and (548±62) ms, t=-5.01, P<0.001], gait cycle [(1 284±168) ms and (1 076±114) ms, t=-5.61, P<0.001], double support time [(531±125) ms and (331±51) ms, t=-8.10, P<0.001], double support time period proportion (0.417±0.063 and 0.309±0.023, t=-8.50, P<0.001), total support time [(914±135) ms and (678±107) ms, t=-7.52, P<0.001], total support time period proportion (0.711±0.027 and 0.627±0.044, t=-8.87, P<0.001), and left static standing time (55.7±8.4 and 51.5±2.2, t=-2.65, P=0.012), for which the differences were statistically significant. The KOA group were lower than the control group in terms of single support time period proportion (0.287±0.030 and 0.334±0.013, t=7.80, P<0.001), right static standing time (44.3±8.4 and 48.5±2.3, t=2.65, P=0.012), step length [(36±8) cm and (52±5) cm, t=9.97, P<0.001], stride length [(70±16) cm and (103±8) cm, t=10.00, P<0.001], velocity [(0.60±0.18) m/s and (1.05±0.19) m/s, t=9.54, P<0.001], left knee range of motion [(42±17)° and (63±4) °, t=6.49, P<0.001], and right knee range of motion [(37±18) ° and (62±3)°, t=7.54, P<0.001], for which the differences were statistically significant. Conclusion:Gait analysis can quantitatively evaluate the condition of patients with KOA, making it possible to transform the diagnostic criteria of KOA from qualitative to quantitative.
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Objective:To explore the effects of Alpha-2-macroglobulin-rich serum (A2MRS) on knee post-traumatic osteoarthritis (PTOA).Methods:The knee PTOA models were constructed by transection of the anterior cruciate ligament (ACL) in 80 SD male rats, aged 2 months and weighing from 250 to 300 g, which were randomized into 4 groups ( n=20): a high dose group (A2MRS containing 20 μg/μL A2M administered), a low dose group (A2MRS containing 10 μg/μL A2M administered), a positive control group (normal saline administered), and a blank control group (the knee joint cut pseudooperatively and normal saline administered). HE, toluidine blue staining, safranine O staining, modified Mankin scoring and Osteoarthritis Research Society International (OARSI) scoring were conducted to evaluate and compare the therapeutic effects of A2MRS on the knee PTOA among the 4 groups. Results:The rat cartilage was thinner with patchy and cracked surface, and the chondrocytes were reduced and distributed unevenly in the positive control group, compared with the blank control group. The modified Mankin score (3.89±0.93) and OARSI score (10.05±0.72) in the positive control group were significantly higher than those in the blank control group (0.67±0.07 and 3.10±0.29) ( P<0.05). The rat cartilage was thicker with basically complete and crack-free surface, and the chondrocytes were increased and distributed more evenly in the high dose group and the low dose group, compared with the positive control group. The modified Mankin scores (1.33±0.50 and 1.56±0.53) and OARSI scores (6.30±0.64 and 4.75±0.66) in the high dose group and the low dose group were significantly lower than those in the positive control group ( P<0.05). However, there were no such differences between the high dose group and the low dose group ( P>0.05). Conclusion:A2MRS effectively delays the pathological process of knee PTOA.
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Objective To investigate the effect of POSTN protein on the proliferation of chondrocytes of tibial plateau in old rats.Methods Cartilage cells collected from the tibial plateau of old rats were cultured in vitro to the third generation.Then the cells were divided into 3 groups:POSTN group,PBS group and POSTN antibody group.The proliferations of the three groups at 24 h,48 h and 72 h were determined by EDU method.The expression of Notch1 protein was detected by immunohistochemistry in all groups at the same time.Female 20-month-old Sprague-Dawley rats were randomly divided into 3 groups:POSTN protein injection group,Phosphate Buffered Saline (PBS) injection group and POSTN antibody injection group.Twelve weeks after the operation,related reagents were injected 3 times consecutively at day 1,day 3,day 5 and EDU was injected into joints at day 1.At 2 weeks after injection,the rats were killed and the knee tibial plateau was taken to observe the proliferation of the cartilage cells.Results At 24 h,there were differences between the three groups O(F=27.32,P=0.017).The proliferation rates of POSTN group [(23±8)％] and PBS group [(21±10)％] were higher than that of POSTN antibody group (16±5)(P=0.003,P=0.011).At 48 h,there were differences between the three groups (F=35.34,P＜0.01).The proliferation rate of POSTN group [(36±11)％] was higher than that of the other groups [(22±6)％],(18±6)％(P=0.021,P＜0.01).At 72h,there were differences between the three groups (F=52.62,P=0.000).The proliferation rate of POSTN group [(56±17)％] was the highest one,the proliferation rate of PBS group [(31±8)％] was the medium,and the POSTN antibody group [(26±7)％] was the lowest one (all P＜0.05).As for Notch1 protein expression in chondrocytes,there were differences between the three groups (F=26.72,P＜0.01).The Notch1 protein was the most frequently expressed in POSTN protein-injection group and the least in the anti-POSTN group.In rats,the proliferation rate of the chondrocytes in the medial tibia plateau of the knee of POSTN protein injection group [(36±14)％],which was the highest,and that of the POSTN antibody injection group [(10 ±4)％] was the lowest (all P＜0.05).Conclusion POSTN protein can promote the proliferation of chondrocytes knee OA rats.POSTN antibody injection has been shown to induce the proliferation of chondrocytes.The POSTN protein may promote the proliferation of chondrocytes by activating the Notch signaling pathway.