CXCL10 is upregulated in synovium and cartilage following articular fracture

The objective of this study was to investigate the expression of the chemokine CXCL10 and its role in joint tissues following articular fracture. We hypothesized that CXCL10 is upregulated following articular fracture and contributes to cartilage degradation associated with post‐traumatic arthritis (PTA). To evaluate CXCL10 expression following articular fracture, gene expression was quantified in synovial tissue from knee joints of C57BL/6 mice that develop PTA following articular fracture, and MRL/MpJ mice that are protected from PTA. CXCL10 protein expression was assessed in human cartilage in normal, osteoarthritic (OA), and post‐traumatic tissue using immunohistochemistry. The effects of exogenous CXCL10, alone and in combination with IL‐1, on porcine cartilage explants were assessed by quantifying the release of catabolic mediators. Synovial tissue gene expression of CXCL10 was upregulated by joint trauma, peaking one day in C57BL/6 mice (25‐fold) versus 3 days post‐fracture in MRL/MpJ mice (15‐fold). CXCL10 protein in articular cartilage was most highly expressed following trauma compared with normal and OA tissue. In a dose dependent manner, exogenous CXCL10 significantly reduced total matrix metalloproteinase (MMP) and aggrecanase activity of culture media from cartilage explants. CXCL10 also trended toward a reduction in IL‐1α‐stimulated total MMP activity (p = 0.09) and S‐GAG (p = 0.09), but not NO release. In conclusion, CXCL10 was upregulated in synovium and chondrocytes following trauma. However, exogenous CXCL10 did not induce a catabolic response in cartilage. CXCL10 may play a role in modulating the chondrocyte response to inflammatory stimuli associated with joint injury and the progression of PTA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1220–1227, 2018.

     

Cartilage viability and catabolism in the intact porcine knee following transarticular impact loading with and without articular fracture

Posttraumatic arthritis commonly develops following articular fracture. The objective of this study was to develop a closed joint model of transarticular impact with and without creation of an articular fracture that maintains the physiologic environment during loading. Fresh intact porcine knees were preloaded and impacted at 294 J via a drop track. Osteochondral cores were obtained from the medial and lateral aspects of the femoral condyles and tibial plateau. Chondrocyte viability was assessed at days 0, 3, and 5 postimpact in sham, impacted nonfractured, and impacted fractured joints. Total matrix metalloproteinase (MMP) activity, aggrecanase (ADAMTS‐4) activity, and sulfated glycosaminoglycan (S‐GAG) release were measured in culture media from days 3 and 5 posttrauma. No differences were observed in chondrocyte viability of impacted nonfractured joints (95.9 ± 6.9%) when compared to sham joints (93.8 ± 7.7%). In impacted fractured joints, viability of the fractured edge was 40.5 ± 27.6% and significantly lower than all other sites, including cartilage adjacent to the fractured edge (p < 0.001). MMP and aggrecanase activity and S‐GAG release were significantly increased in specimens from the fractured edge. This study showed that joint impact resulting in articular fracture significantly decreased chondrocyte viability, increased production of MMPs and aggrecanases, and enhanced S‐GAG release, whereas the same level of impact without fracture did not cause such changes. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:501–510, 2011     

Cytoskeletal dissolution blocks oxidant release and cell death in injured cartilage

The mechanisms by which articular surface impact causes post‐traumatic osteoarthritis are not well understood, but studies of cartilage explants implicate the mitochondrial electron transport chain as a source of oxidants that cause chondrocyte death from mechanical injury. The linkage of mitochondria to the cytoskeleton suggests that they might release oxidants in response to mechanical strain, an effect that disrupting the cytoskeleton would prevent. To test this we investigated the effects of agents that promote the dissolution of microfilaments (cytochalasin B) or microtubules (nocodazole) on oxidant production and chondrocyte death following impact injury. Osteochondral explants treated with cytochalasin B or nocodazole for 4 h were impacted (7 J/cm²) and stained for oxidant production directly after impact and for cell viability 24 h after impact. Surfaces within and outside impact sites were then imaged by confocal microscopy. Both agents significantly reduced impact‐induced oxidant release (p < 0.05); however, cytochalasin B was more effective than nocodazole (>60% reduction vs. 40% reduction, respectively). Both agents also prevented impact induced cell death. Dissolution of the cytoskeleton by both drugs was confirmed by phalloidin staining and confocal microscopy. These findings show that chondrocyte mortality from impact injury depends substantially on mitochondrial–cytoskeletal linkage, suggesting new approaches to stem mechanically induced cartilage degeneration. © 2011 Orthopaedic Research Society. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:593–598, 2012     

Cartilage‐on‐cartilage versus metal‐on‐cartilage impact characteristics and responses

A common in vitro model for studying acute mechanical damage in cartilage is to impact an isolated osteochondral or cartilage specimen with a metallic impactor. The mechanics of a cartilage‐on‐cartilage (COC) impact, as encountered in vivo, are likely different than those of a metal‐on‐cartilage (MOC) impact. The hypothesis of this study was that impacted in vitro COC and MOC specimens would differ in their impact behavior, mechanical properties, chondrocyte viability, cell metabolism, and histologic structural damage. Osteochondral specimens were impacted with either an osteochondral plug or a metallic cylinder at the same delivered impact energy per unit area, and processed after 14 days in culture. The COC impacts resulted in about half of the impact maximum stress and a quarter of the impact maximum stress rate of change, as compared to the MOC impacts. The impacted COC specimens had smaller changes in mechanical properties, smaller decreases in chondrocyte viability, higher total proteoglycan content, and less histologic structural damage, as compared to the impacted MOC specimens. If MOC impact conditions are to be used for modeling of articular injuries and post‐traumatic osteoarthritis, the differences between COC and MOC impacts must be kept in mind. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 887–893, 2013     

Inhibition of cell‐matrix adhesions prevents cartilage chondrocyte death following impact injury

Focal adhesions are transmembrane protein complexes that attach chondrocytes to the pericellular cartilage matrix and in turn, are linked to intracellular organelles via cytoskeleton. We previously found that excessive compression of articular cartilage leads to cytoskeleton‐dependent chondrocyte death. Here we tested the hypothesis that this process also requires integrin activation and signaling via focal adhesion kinase (FAK) and Src family kinase (SFK). Osteochondral explants were treated with FAK and SFK inhibitors (FAKi, SFKi, respectively) for 2 h and then subjected to a death‐inducing impact load. Chondrocyte viability was assessed by confocal microscopy immediately and at 24 h post‐impact. With no treatment immediate post‐impact viability was 59%. Treatment with 10 µM SFKi, 10 μM, or 100 µM FAKi improved viability to 80%, 77%, and 82%, respectively (p < 0.05). After 24 h viability declined to 34% in controls, 48% with 10 µM SFKi, 45% with 10 µM FAKi, and 56% with 100 µM FAKi (p < 0.01) treatment. These results confirmed that most of the acute chondrocyte mortality was FAK‐ and SFK‐dependent, which implicates integrin‐cytoskeleton interactions in the death signaling pathway. Together with previous findings, these data support the hypothesis that the excessive tissue strains accompanying impact loading induce death via a pathway initiated by strain on cell adhesion receptors. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:448–454, 2014.     

Brazilin blocks catabolic processes in human osteoarthritic chondrocytes via inhibition of NFKB1/p50

This study aimed to evaluate the chondroprotective and anti‐inflammatory activity of brazilin in human osteoarthritic (OA) cartilage and chondrocytes with particular focus on the nuclear factor‐kappa B (NF‐κB) pathway. Therefore, brazilin was isolated from Caesalpinia sappan and identified using high performance liquid chromatography (HPLC). The effect of brazilin was assessed in cartilage explants treated with 10 ng/ml interleukin (IL)‐1β and 10 ng/ml tumor necrosis factor (TNF)‐α using histological and biochemical glycosaminoglycan (GAG) analyses and in primary chondrocytes treated with 10 ng/ml IL‐1β using RT‐qPCR, ELISA, and Western blot. The involvement of NF‐κB signaling was examined using a human NF‐κB signaling array and in silico pathway analysis. Brazilin was found to reduce the GAG loss from cartilage explants stimulated with IL‐1β and TNF‐α. NF‐κB pathway analysis in chondrocytes revealed NFKB1/p50 as a central player regulating the anti‐inflammatory activities of brazilin. Brazilin suppressed the IL‐1β‐mediated up‐regulation of OA markers and the induction of NFKB1/p50 in chondrocytes. In conclusion, brazilin effectively attenuates catabolic processes in human OA cartilage and chondrocytes—at least in part due to the inhibition of NFKB1/p50—which indicates a chondroprotective potential of brazilin in OA. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2431–2438, 2018.

     

Viability and volume of in situ bovine articular chondrocytes-changes following a single impact and effects of medium osmolarity

OBJECTIVE: Mechanical stress above the physiological range can profoundly influence articular cartilage causing matrix damage, changes to chondrocyte metabolism and cell injury/death. It has also been implicated as a risk factor in the development of osteoarthritis (OA). The mechanism of cell damage is not understood, but chondrocyte volume could be a determinant of the sensitivity and subsequent response to load. For example, in OA, it is possible that the chondrocyte swelling that occurs renders the cells more sensitive to the damaging effects of mechanical stress. This study had two aims: (1) to investigate the changes to the volume and viability of in situ chondrocytes near an injury to cartilage resulting from a single blunt impact, and (2) to determine if alterations to chondrocyte volume at the time of impact influenced cell viability. METHODS: Explants of bovine articular cartilage were incubated with the fluorescent indicators calcein-AM and propidium iodide permitting the measurement of cell volume and viability, respectively, using confocal laser scanning microscopy (CLSM). Cartilage was then subjected to a single impact (optimally 100g from 10 cm) delivered from a drop tower which caused areas of chondrocyte injury/death within the superficial zone (SZ). The presence of lactate dehydrogenase (LDH; an enzyme released following cell injury) was used to determine the effects of medium osmolarity on the response of chondrocytes to a single impact. RESULTS: A single impact caused discrete areas of chondrocyte injury/death which were almost exclusively within the SZ of cartilage. There appeared to be two phases of cell death, a rapid phase lasting approximately 3 min, followed by a slower progressive 'wave of cell death' away from the initial area lasting for approximately 20 min. The volume of the majority (88.1+/-5.99% (n=7) of the viable chondrocytes in this region decreased significantly (P<0.006). By monitoring LDH release, a single impact 5 min after changing the culture medium to hyper-, or hypo-osmolarity, reduced or stimulated chondrocyte injury, respectively. CONCLUSIONS: A single impact caused temporal and spatial changes to in situ chondrocyte viability with cell shrinkage occurring in the majority of cells. However, chondrocyte shrinkage by raising medium osmolarity at the time of impact protected the cells from injury, whereas swollen chondrocytes were markedly more sensitive. These data showed that chondrocyte volume could be an important determinant of the sensitivity and response of in situ chondrocytes to mechanical stress.     

Decreased osteogenesis in mesenchymal stem cells derived from the aged mouse is associated with enhanced NF‐κB activity

Aging is associated with significant bone loss and delayed fracture healing. NF‐κB activation is highly correlated with inflammatory‐associated bone diseases including infection, wear particle exposure, and chronic inflammation during natural aging processes. The critical roles of NF‐κB in both the pro‐inflammatory response and osteoclast‐mediated bone resorption have been well defined. However, the biological effects of NF‐κB activation in mesenchymal stem cell (MSC)‐mediated bone formation remain largely unknown. In the current study, bone marrow‐MSCs were isolated from young (8 weeks old) and aged (72 weeks old) mice. NF‐κB activity in MSCs at basal levels and under different biological conditions were determined by our recently established lentiviral vector‐based luciferase reporter assay. We found that NF‐κB activity was increased in aged MSCs at basal levels or when exposed to low dose (10 or 100 ng/ml) lipopolysaccharide (LPS); this effect was not seen when the cells were exposed to higher dose (1 μg/ml) LPS. During osteogenesis, NF‐κB activity was increased in aged MSCs at weeks 1 and 2, but showed no significant difference at week 3. Both Smurf2 and TAZ, the NF‐κB target genes that regulate osteogenic differentiation, were increased in aged MSCs. In addition, the expression of RANKL was dramatically increased, and OPG was decreased in aged MSCs. Our findings suggest that targeting NF‐κB activity in MSCs has the potential to modulate aging‐associated bone loss, or enhance bone‐healing in aged patients. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:281–288, 2017.

     

Mitoprotective therapy preserves chondrocyte viability and prevents cartilage degeneration in an ex vivo model of posttraumatic osteoarthritis

No disease‐modifying osteoarthritis (OA) drugs are available to prevent posttraumatic osteoarthritis (PTOA). Mitochondria (MT) mediate the pathogenesis of many degenerative diseases, and recent evidence indicates that MT dysfunction is a peracute (within minutes to hours) response of cartilage to mechanical injury. The goal of this study was to investigate cardiolipin‐targeted mitoprotection as a new strategy to prevent chondrocyte death and cartilage degeneration after injury. Cartilage was harvested from bovine knee joints and subjected to a single, rapid impact injury (24.0 ±1.4 MPa, 53.8 ± 5.3 GPa/s). Explants were then treated with a mitoprotective peptide, SS‐31 (1µM), immediately post‐impact, or at 1, 6, or 12 h after injury, and then cultured for up to 7 days. Chondrocyte viability and apoptosis were quantified in situ using confocal microscopy. Cell membrane damage (lactate dehydrogenase activity) and cartilage matrix degradation (glycosaminoglycan loss) were quantified in cartilage‐conditioned media. SS‐31 treatment at all time points after impact resulted in chondrocyte viability similar to that of un‐injured controls. This effect was sustained for up to a week in culture. Further, SS‐31 prevented impact‐induced chondrocyte apoptosis, cell membrane damage, and cartilage matrix degeneration. Clinical Significance: This study is the first investigation of cardiolipin‐targeted mitoprotective therapy in cartilage. These results suggest that even when treatment is delayed by up to 12 h after injury, mitoprotection may be a useful strategy in the prevention of PTOA. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2147–2156, 2018.

     

How severe must repetitive loading be to kill chondrocytes in articular cartilage?

OBJECTIVE: Little is known about the effects of severe repetitive loading on articular cartilage chondrocytes, even though epidemiological studies associate this type of loading with osteoarthritis. We hypothesize that repetitive loading can kill cartilage chondrocytes in a dose-related manner. DESIGN: Large cartilage-on-bone specimens were cut from the patella groove of bovine knees obtained directly from a slaughterhouse. Cartilage was loaded using a flat impermeable indenter in such a manner that the loaded region was supported naturally by surrounding cartilage and subchondral bone. Specimens received 3600 cycles of compressive loading at 1 Hz, with the peak load lying in the range 1-70% of the force required to damage cartilage in a single loading cycle (35 MPa). Cell viability was assessed in thick sections of loaded and control cartilage using a paravital staining method: fluorescein diacetate stained live cells green, and propidium iodide stained dead cells red. The assay was validated on cartilage which had been subjected to repeated freeze-thaw cycles to kill the chondrocytes. RESULTS: Paravital staining revealed 100% cell death after one freeze-thaw cycle at -196 degrees C and three cycles at -20 degrees C. Baseline chondrocyte viability was 80% in unloaded cartilage, and viability decreased when applied compressive loading exceeded 6 MPa. Above this threshold, cell viability was inversely proportional to applied stress. When gross damage to the cartilage surface first became evident, above 14 MPa, 40% of cells remained viable. Load-induced chondrocyte death was greatest in the surface zone, and extended beyond the loaded area. Electron micrographs indicated that some cells were dying by apoptosis. CONCLUSIONS: Some chondrocytes are much more vulnerable to repetitive mechanical loading than others, suggesting that vigorous activity may lead to cell death in articular cartilage.     

Genipin crosslinking of cartilage enhances resistance to biochemical degradation and mechanical wear

Collagen crosslinking enhances many beneficial properties of articular cartilage, including resistance to chemical degradation and mechanical wear, but many crosslinking agents are cytotoxic. The purpose of this study was to evaluate the effectiveness of genipin, a crosslinking agent with favorable biocompatibility and cytotoxicity, as a potential treatment to prevent the degradation and wear of articular cartilage. First, the impact of genipin concentration and treatment duration on the viscoelastic properties of bovine articular cartilage was quantified. Next, two short‐term (15 min) genipin crosslinking treatments were chosen, and the change in collagenase digestion, cartilage wear, and the friction coefficient of the tissue with these treatments was measured. Finally, chondrocyte viability after exposure to these genipin treatments was assessed. Genipin treatment increased the stiffness of healthy, intact cartilage in a dose‐dependent manner. The 15‐min crosslinking treatments improved cartilage's resistance to both chemical degradation, particularly at the articular surface, and to damage due to mechanical wear. These enhancements were achieved without sacrificing the low coefficient of friction of the tissue and at a genipin dose that preserved chondrocyte viability. The results of this study suggest that collagen crosslinking via genipin may be a promising preventative treatment to slow the degradation of cartilage. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 33:1571–1579, 2015.     

Single high‐energy impact load causes posttraumatic OA in young rabbits via a decrease in cellular metabolism

Articular cartilage deterioration commonly occurs following traumatic joint injury. Patients with posttraumatic osteoarthritis (PTA) experience pain and stiffness in the involved joint causing limited mobility and function. The mechanism by which PTA occurs has not been fully delineated. The goal of this study was to determine if a single high‐energy impact load could cause the development of PTA in 3‐month‐old NZ White rabbits. Each rabbit underwent the application of a single, rapid, high‐energy impact load to the posterior aspect of their right medial femoral condyle using a previously validated mechanism. At regular intervals (0, 1, 6 months) the injured cartilage was harvested and analyzed for the presence of PTA. Each specimen was assessed histologically for cell and tissue morphology and chondrocyte metabolism, including BMP‐2 production and synthesis of extracellular matrix (type II procollagen mRNA). Cartilage from the contralateral sham limb, as well as uninjured cartilage from the experimental limb served as internal controls for each animal. Significant changes were found in the morphology of the cartilage including proteoglycan loss along with decreased BMP‐2 and type II procollagen mRNA staining. These findings confirm that a single high‐energy impact load can cause the development of PTA by disrupting the extracellular matrix and by causing a decrease in chondrocyte metabolism. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:347–352, 2009     

Diminished cartilage creep properties and increased trabecular bone density following a single,sub‐fracture impact of the rabbit femoral condyle

Traumatic injury to articular cartilage can lead to post‐traumatic arthritis. We used a custom pendulum device to deliver a single, near‐fracture impact to the medial femoral condyles of rabbits. Impact was localized to a region ～3 mm in diameter, and impact stress averaged ～100 MPa. Animals were euthanized at 0, 1, and 6 months after impact. Cartilage mechanical properties from impacted and sham knees were evaluated by creep‐indentation testing, and periarticular trabecular bone was evaluated by microCT and histomorphometry. Impact caused immediate and statistically significant loss of cartilage thickness (?40% vs. sham) and led to a greater than twofold increase in creep strain. From 0 to 6 months after impact, the ability of cartilage to recover from creep deformation became significantly impaired (percent recovery different from control at 1 and 6 months). At 1 month, there was a 33% increase in the trabecular bone volume fraction of the epiphysis beneath the site of impact compared to control, and increased bone formation was observed histologically. Taken together, these findings demonstrate that a single, high‐energy impact below the fracture threshold leads to acute deleterious changes in the viscoelastic properties of articular cartilage that worsen with time, while at the same time stimulating increased bone formation beneath the impact site. Published by Wiley Periodicals, Inc. J Orthop Res 28:1307–1314, 2010     

Rotenone prevents impact‐induced chondrocyte death

Mechanical insult to articular cartilage kills chondrocytes, an event that may increase the risk of posttraumatic osteoarthritis. Recent reports indicate that antioxidants decrease impact‐induced chondrocyte death, but the source(s) of oxidants, the time course of oxidant release, and the identity of the oxidative species generated in response to injury are unknown. A better understanding of these processes could lead to new treatments of acute joint injuries. To that end, we studied the kinetics and distribution of oxidant production in osteochondral explants subjected to a single, blunt‐impact injury. We followed superoxide production by measuring the time‐dependent accumulation of chondrocyte nuclei stained with the superoxide‐sensitive probe dihydroethidium. The percentage of chondrocytes that were dihydroethidium‐positive was 35% above baseline 10 min after impact, and 65% above baseline 60 min after impact. Most positive cells were found within and near areas contacted directly by the impact platen. Rotenone, an electron transport chain inhibitor, was used to test the hypothesis that mitochondria contribute to superoxide release. Rotenone treatment significantly reduced dihydroethidium staining, which remained steady at 15% above baseline for up to 60 min postimpact. Moreover, rotenone reduced chondrocyte death in impact sites by more than 40%, even when administered 2 h after injury (p < 0.001). These data show that much of the acute chondrocyte mortality caused by in vitro impact injuries results from superoxide release from mitochondria, and suggest that brief exposure to free radical scavengers could significantly improve chondrocyte viability following joint injury. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1057–1063, 2010     

Mechanical response of bovine articular cartilage under dynamic unconfined compression loading at physiological stress levels

OBJECTIVE: The objective of this study was to characterize the dynamic modulus and compressive strain magnitudes of bovine articular cartilage at physiological compressive stress levels and loading frequencies. DESIGN: Twelve distal femoral cartilage plugs (3mm in diameter) were loaded in a custom apparatus under load control, with a load amplitude up to 40 N and loading frequencies of 0.1, 1, 10 and 40 Hz, resulting in peak Cauchy stress amplitudes of 4.8 MPa (engineering stress 5.7 MPa). RESULTS: The equilibrium Young's modulus under a tare load of 0.4N was 0.49+/-0.10 MPa. In the limit of zero applied stress, the incremental dynamic modulus derived from the slope of the stress-strain curve increased from 14.6+/-6.9 MPa at 0.1 Hz to 28.7+/-7.8 MPa at 40 Hz. At 4 MPa of applied stress, the corresponding increase was from 48.2+/-13.5 MPa at 0.1 Hz to 64.8+/-13.0 MPa at 40 Hz. Peak compressive strain amplitudes varied from 15.8+/-3.4% at 0.1 Hz to 8.7+/-1.8% at 40 Hz. The phase angle decreased from 28.8 degrees +/-6.7 degrees at 0.1 Hz to-0.5 degrees +/-3.8 degrees at 40 Hz. DISCUSSION: These results are representative of the functional properties of articular cartilage under physiological load magnitudes and frequencies. The viscoelasticity and nonlinearity of the tissue helps to maintain the compressive strains below 20% under the physiological compressive stresses achieved in this study. These findings have implications for our understanding of cartilage metabolism and chondrocyte viability under various loading regimes. They also help establish guidelines for cartilage functional tissue engineering studies.     

The development of posttraumatic arthritis after articular fracture

Posttraumatic arthritis (PTA) is one of the most frequent causes of disability after trauma involving weight-bearing joints and is estimated to be responsible for approximately 10% of the 21 million Americans who have osteoarthritis. Despite a number of similarities in the pathology and end-stage disease of PTA with primary osteoarthritis, the mechanisms involved in the onset and progression of joint degeneration after articular fracture are poorly understood. The largest area of study regarding articular fractures and the development of arthritic changes has focused on the role of adequate surgical reduction of the articular surfaces. However, it is now apparent that a number of complex and interacting biomechanical, biochemical, and, possibly, genetic factors contribute to the development of osteoarthritic changes in the joint after joint trauma, ranging from the cell and molecular level to the joint and systemic level. In this paper, we discuss the potential roles of the initial impact and fracture as well as the subsequent alterations in joint loading, biomechanical and metabolic properties of the cartilage, local and systemic inflammatory cytokines, and viability of chondrocytes in the progression of PTA. An improved understanding of the mechanisms involved in the development of PTA will hopefully lead to the improvement of surgical and nonsurgical therapies for this disease.     

冷冻保存对软骨细胞存活率及代谢活性的影响

目的了解各种冷冻保存方法对软骨细胞的存活率和代谢活性的影响，寻找满意的软骨组织冻存方法。方法采用梯度慢速降温法和连续慢速降温法对兔关节软骨进行低温冷冻保存处理，通过荧光染色及^35SO4摄入率了解冻存后软骨细胞的存活率和代谢活性。结果采用梯度降温法的软骨细胞存活率为61％，显著高于连续降温法；冷冻保存对软骨细胞的代谢活性有一定的影响．但与对照组的差异并无统计学意义。结论梯度降温法较传统保存方法能显著提高冷冻保存后软骨细胞的存活率，并能维持软骨细胞的代谢活性，是理想的关节软骨冷冻保存方法。     

Survival of articular cartilage after controlled impact

Survival characteristics of forty-three specimens of living human bone and articular cartilage from the knees of eight renal-transplant donors were studied, using a drop-tower device. Autoradiography and light and scanning electron microscopy revealed no evidence of chondrocyte death or structural damage until stress levels of twenty-five newtons per square millimeter were reached, corresponding to strains on the order of 20 to 30 per cent and involving energy absorption of one millijoule per cubic millimeter. The data for strain rates of 500 and 1000 s-1 suggest that impact loads sufficient to fracture a femoral shaft of an automobile occupant are nearly sufficient to cause chondrocyte death and fissuring in the articular cartilage of either the knee or the hip if the load-bearing areas measure less than 500 square millimeters.