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1.
    
We investigated whether immature allysine‐derived cross‐links provide mechanically labile linkages by exploring the effects of immature cross‐link stabilization at three levels of collagen hierarchy: damaged fibril morphology, whole tendon mechanics, and molecular stability. Tendons from the tails of young adult steers were either treated with sodium borohydride (NaBH4) to stabilize labile cross‐links, exposed only to the buffer used during stabilization treatment, or maintained as untreated controls. One‐half of each tendon was then subjected to five cycles of subrupture overload. Morphologic changes to collagen fibrils resulting from overload were investigated using scanning electron microscopy, and changes in the hydrothermal stability of collagen molecules were assessed using hydrothermal isometric tension testing. NaBH4 cross‐link stabilization did not affect the response of tendon collagen to tensile overload at any of the three levels of hierarchy studied. Cross‐link stabilization did not prevent the characteristic overload‐induced mode of fibril damage that we term discrete plasticity. Similarly, stabilization did not alter the mechanical response of whole tendons to overload, and did not prevent an overload‐induced thermal destabilization of collagen molecules. Our results indicate that hydrothermally labile cross‐links may not be as mechanically labile as was previously thought. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31:1907–1913, 2013  相似文献   

2.
    
Manipulations in cell culture and mouse models have demonstrated that reduction of collagen V results in altered fibril structure and matrix assembly. A tissue‐dependent role for collagen V in determining mechanical function was recently established, but its role in determining regional properties has not been addressed. The objective of this study was to define the role(s) of collagen V expression in establishing the site‐specific properties of the supraspinatus tendon. The insertion and midsubstance of tendons from wild type, heterozygous and tendon/ligament‐specific null mice were assessed for crimp morphology, fibril morphology, cell morphology, as well as total collagen and pyridinoline cross‐link (PYD) content. Fibril morphology was altered at the midsubstance of both groups with larger, but fewer, fibrils and no change in cell morphology or collagen compared to the wild type controls. In contrast, a significant disruption of fibril assembly was observed at the insertion site of the null group with the presence of structurally aberrant fibrils. Alterations were also present in cell density and PYD content. Altogether, these results demonstrate that collagen V plays a crucial role in determining region‐specific differences in mouse supraspinatus tendon structure. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:2154–2161, 2016.  相似文献   

3.
    
The cause of the high failure rates often observed following rotator cuff tendon repairs, particularly massive tears, is not fully understood. Collagen structural changes have been shown to alter tendon thermal and mechanical properties. This study aimed to form a quantitative rather than qualitative assessment, of whether differences in collagen structure and integrity existed between small biopsies of normal, small, and massive rotator cuff tears using differential scanning calorimetry. Thermal properties were measured for 28 human biopsies taken intra‐operatively from normal, small, and massive rotator cuff tendon tears in this powered study. Denaturation temperatures are represented by Tonset (°C) and Tpeak (°C). The Tonset is proposed to represent water‐amide hydrogen bond breakage and resulting protein backbone mobility. Tpeak reportedly corresponds to the temperature at which the majority of proteins fall out of solution. Denaturation enthalpy (ΔH) should correlate with the amount of triple helical structure that is denatured. Fluorescence and confocal microscopy allowed quantitative validation. Small and massive rotator cuff tears had significantly higher Tonset, Tpeak, and ΔH compared to controls. Polarized light microscopy of torn tendons confirmed greater collagen structural disruption compared to controls. These novel findings suggest greater quantifiable collagen structural disruption in rotator cuff tears, compared to controls. This study offers insight into possible mechanisms for the reduced strength of torn tendons and may explain why repaired tendons fail to heal. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1938–1943, 2011  相似文献   

4.
    
One proposed mechanism of tendon lengthening is the “sliding fibril” hypothesis, in which tendons lengthen through the sliding of discontinuous fibrils after release of decorin‐fibronectin interfibrillar bonds. The pentapeptide NKISK has been reported to inhibit the binding of decorin, a proteoglycan on the surface of collagen fibrils, to fibronectin, a tissue adhesion molecule, which are thought to play a role in interfibrillar binding. Prior investigations have demonstrated that NKISK produces in vivo tendon lengthening. This study investigates the effect of potential clinically applicable NKISK injection regimens in an in vivo model. One hundred and thirteen male Sprague‐Dawley rats were divided into 15 different treatment groups, each receiving percutaneous patellar tendon injections of NKISK, QKTSK (a “nonsense” pentapeptide), or PBS of varying volumes, concentrations, and injection schedules. Following sacrifice, the patellar tendon lengths were measured in all groups, and biomechanical testing was performed with comparisons made to the contralateral, untreated control limbs. Tendon lengthening was significantly greater (p ≤ 0.05) in nearly all NKISK‐treated tendons as compared to PBS‐ and QKTSK‐treated tendons and was dose‐dependent. Measured lengthening was less in rats whose sacrifice was delayed following the final injection of NKISK, which likely indicates recontraction of lengthened tendons, but they remained significantly longer than the controls. Biomechanical testing did not reveal significant differences in ultimate load, modulus, stiffness, or displacement. This study demonstrates that NKISK given in clinically plausible dosing regimens produces dose‐dependent tendon lengthening in an in vivo setting with minimal effects on the mechanical properties of the treated tendons. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:971–976, 2008  相似文献   

5.
    
Using functional tissue engineering principles, our laboratory has produced tendon repair tissue which matches the normal patellar tendon force‐displacement curve up to 32% of failure. This repair tissue will need to withstand more strenuous activities, which can reach or even exceed 40% of failure force. To improve the linear stiffness of our tissue engineered constructs (TECs) and tissue engineered repairs, our lab is incorporating the glycosaminoglycan chondroitin‐6‐sulfate (C6S) into a type I collagen scaffold. In this study, we examined the effect of C6S incorporation and mechanical stimulation cycle number on linear stiffness and mRNA expression (collagen types I and III, decorin and fibronectin) for mesenchymal stem cell (MSC)‐collagen sponge TECs. The TECs were fabricated by inoculating MSCs at a density of 0.14 × 106 cells/construct onto pre‐cut scaffolds. Primarily type I collagen scaffold materials, with or without C6S, were cultured using mechanical stimulation with three different cycle numbers (0, 100, or 3,000 cycles/day). After 2 weeks in culture, TECs were evaluated for linear stiffness and mRNA expression. C6S incorporation and cycle number each played an important role in gene expression, but only the interaction of C6S incorporation and cycle number produced a benefit for TEC linear stiffness. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1092–1099, 2010  相似文献   

6.
  总被引:5,自引:0,他引:5  
Porcine small intestinal submucosa (SIS) was previously shown to enhance the mechanical properties of healing medial collateral ligaments (MCL), and the histomorphological appearance and collagen type V/I ratio were found to be close to those of normal MCL. We hypothesized that at a longer term, 26 weeks, SIS could guide a better organized neo-ligament formation, increasing mechanical properties and increasing collagen fibril diameters mediated by a reduction in collagen type V. A 6 mm gap injury in the right MCL was surgically created in 38 rabbits, while the contralateral intact MCL served as a sham-operated control. In half the animals, a strip of SIS was sutured onto the severed ends. In the other half, no SIS was applied. The cross-sectional area (CSA) was determined with a laser micrometer system. The femur-MCL-tibia complex was mechanically tested in uniaxial tension. Histomorphology was determined through H&E and immunofluorescent staining and transmission electron microscopy (TEM). Sodium-dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was used to determine collagen type V/I ratio. SIS-treated MCLs displayed a 28% reduction in CSA, a 33% increase in tangent modulus, and a 50% increase in tensile strength compared with the nontreated group (p < 0.05). TEM showed groups of collagen fibrils with larger diameters in the SIS-treated ligaments in comparison with uniformly small fibrils for the nontreated group. H&E staining showed more densely stained collagen fibers in the SIS-treated group aligned along the longitudinal axis with more interspersed spindle-shaped cells. Immunofluorescent staining showed less collagen type V signals, confirmed by a 5% lower ratio of collagen type V/I compared with the nontreated controls (p < 0.05). The findings extend the shorter term 12-week results, and support the potential of porcine SIS as a bioscaffold to enhance ligament healing.  相似文献   

7.
    
The purpose of this study was to investigate the existence of sex‐based differences in the ultrastructural characteristics of the human anterior cruciate ligament (ACL) as the underlying cause of differences in the structural and mechanical properties between sexes. The ACL of six male and six female cadaveric donors were randomly chosen from a pool of 10 male and 10 female ACLs that had previously been tested for their structural and mechanical properties. Eighteen tissue samples from the distal, proximal, and middle sections of the anteromedial and posterolateral bundles were analyzed by transmission electron microscopy. Female ACLs exhibited both lower fibril concentration and lower percent area occupied by collagen fibrils (p < 0.05) compared to males. There was also a difference in the fibril diameters (p < 0.05); donor age, height, body mass, and body mass index contributed significantly to this difference. In females, ACL stiffness and modulus of elasticity were highly correlated to fibril concentration (r = 0.96 and 0.97, respectively); in males ACL failure load and strength were highly correlated to percent area occupied by collagen (r = 0.96 and 0.96, respectively). These differences in ultrastructure may underlie differences in ACL properties between sexes. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:945–950, 2008  相似文献   

8.
  总被引:3,自引:0,他引:3  
The purpose of this study was to assess the effects of stress shielding on the microstructure and ultrastructure of the patellar tendon using 40 mature female Japanese white rabbits. The patellar tendon was completely released from stress by drawing the patella toward the tibial tubercle with a stainless steel wire installed between them. Microstructurally, stress shielding for 3 and 6 weeks increased the number of cells approximately fivefold, to that of the control tendon. Collagen bundles were less well oriented in the stress-shielded tendon than in the control. Ultrastructurally, small collagen fibrils with a diameter of less than 90nm increased in the stress-shielded tendon. The median collagen fibril diameter in 6-week stress-shielded tendon was significantly smaller (P 0.05) than in the control tendon (58.8% of control). The ratio of the total area of collagen fibrils to the whole visualized area in the stress-shielded patellar tendon was significantly smaller at 3 and 6 weeks than that in the control. This study demonstrated that complete stress shielding significantly affects the microstructure and ultrastructure of the patellar tendon  相似文献   

9.
    
We investigated the effects of exercise‐induced loading on the collagen network of equine articular cartilage. Collagen fibril architecture at a site (1) subjected to intermittent high‐intensity loading was compared with that of an adjacent site (2) sustaining continuous low‐level load. From horses exposed to forced exercise (CONDEX group) or not (PASTEX group), the spatial parallelism of fibrils and the orientation angle between fibrils and the surface at depths 9 µm apart through cartilage from surface to tidemark were determined using polarized light microscopy, and expressed as parallelism index (PI) and orientation index (OI). PI was significantly higher in site 2 than 1 in CONDEX and PASTEX groups. PI was significantly higher in forced exercised horses at site 2 but not site 1. OI was significantly greater (more perpendicular to the surface) in the superficial and deep cartilage of site 2 than 1 in both CONDEX and PASTEX groups. Superficial zone OI was higher in exercised horses at site 1 but not at site 2. Exercise increased collagen parallelism and affected orientation. The site differences in OI indicate that Benninghoff's classic predominantly perpendicular arcades appear not to be a consistent architectural feature, but adapt to local forces sustained. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res  相似文献   

10.
Tendon rupture has been linked with anabolic steroid abuse on the basis of a small number of published case reports. Although experimental data from animal models suggest steroids alter the biomechanical properties of tendon, ultrastructural evidence to support this theory is lacking. Indeed, microscopic analysis of human tendon from steroid users has not previously been reported. In this study, specimens of ruptured human tendon from four patients were biopsied during surgical repair. Two of the subjects were anabolic steroid users, and two subjects were used as nonsteroid-user controls. Ruptured tendon from both groups was examined using electron microscopy. No differences in collagen fibril ultrastructure were seen. We conclude that anabolic steroids did not induce ultrastructural collagen changes that might predispose to tendon rupture in humans.  相似文献   

11.
    
Musculoskeletal soft tissue injuries are very common, yet poorly understood. We investigated molecular‐level changes in collagen caused by tensile overload of bovine tail tendons in vitro. Previous investigators concluded that tensile tendon rupture resulted in collagen denaturation, but our study suggests otherwise. Based on contemporary collagen biophysics, we hypothesized that tensile overload would lead to reduced thermal stability without change in the nativity of the molecular conformation. The thermal behavior of collagen from tail tendons ruptured in vitro at two strain rates (0.01 s?1 and 10 s?1) was measured by differential scanning calorimetry (DSC). The 1,000‐fold difference in strain rate was used since molecular mechanisms that determine mechanical behavior are thought to be strain rate‐dependent. DSC revealed that the collagen in tensile overloaded tendons was less thermally stable by 3° to 5°C relative to undamaged controls and was not denatured since there was no change in enthalpy of denaturation. The decrease in thermal stability occurred throughout the overloaded regions, independent of rupture site, and was greater in specimens ruptured at the lower strain rate. The deformation mechanism apparently involves disruption of the lattice structure of the collagen fibrils and greatly increases the molecular freedom of the collagen molecules, leading to reduced thermal molecular stability and the previously reported increased proteolysis. This has important implications for understanding soft tissue injuries, disease etiology and treatment, and for developing tissue engineered products with improved durability. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res  相似文献   

12.
    
Dynamic mechanical culture systems are a widely studied approach for improving the functional mechanical properties of tissue engineering constructs intended for loading‐bearing orthopedic applications such as tendon/ligament reconstruction. The design of effective mechanical stimulation regimes requires a fundamental understanding of the effects of cyclic strain parameters on the resulting construct properties. Toward this end, these studies employed a modular cyclic strain bioreactor system and fibroblast‐seeded, porous polyurethane substrates to systematically investigate the effect of varying cyclic strain amplitude, rate, frequency, and daily cycle number on construct mechanical properties. Significant differences were observed in response to variation of all four loading parameters tested. In general, the highest values of elastic modulus within each experimental group were observed at low to intermediate values of the experimental variables tested, corresponding to the low to subphysiological range (2.5% strain amplitude, 25%/s strain rate, 0.1–0.5 Hz frequency, and 7,200–28,800 cycles/day). These studies demonstrate that fibroblasts are sensitive and responsive to multiple characteristics of their mechanical environment, and suggest that systematic optimization of dynamic culture conditions may be useful for the acceleration of construct maturation and mechanical function. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:1105–1113, 2008  相似文献   

13.
    
BACKGROUND AND OBJECTIVE: Deep coagulation of skin collagen by Er:YAG laser repetitive pulses has been predicted by previous theoretical models and later demonstrated on animal skin. The goal of this study was to determine the effect of repetitive Er:YAG laser pulses on human skin and its response to this treatment. STUDY DESIGN/MATERIALS AND METHODS: Lid skin of six female volunteers with blepharochalasis has been treated with laser at day 0, 7, and 21 before elective surgery-blepharoplasty. The treated skin was excised as part of the procedure and prepared for further histological examination. We used a 2,940 nm Er:YAG laser (Fidelis M320A by Fotona) with 'smooth' mode parameters: fluence from 0.50 to 2.00 J/cm2; six pulses per packet; 550 microsecond/pulse, 250 millisecond/packet; single pass, no overlapping; spot size 5 mm; repetition rate 20 Hz. RESULTS: We observed deep collagen denaturation at laser fluences of 1.25 J/cm2 and over; epidermal damage was proportional to fluence with total coagulation of the epidermal layer at fluences of 1.75 J/cm2 and over. At day 7 after laser treatment we observed a complete regeneration of the epidermal layer and a regeneration zone within the dermis with prominent infiltration of CD68+ monocytes/macrophages. At day 21 after laser treatment we observed collagen remodeling and (myo-)fibroblast proliferation at tissue depths of up to 240 microm. CONCLUSIONS: Repetitive Er:YAG laser irradiation is effective in deep denaturation and remodeling of human skin collagen in vivo, with less epidermal damage compared to standard Er:YAG laser skin resurfacing.  相似文献   

14.
    
BACKGROUND AND OBJECTIVES: Low energy laser therapy has been shown to enhance mechanical strength of healing medial collateral ligament (MCL) in rats. The present study investigated its effects on the ultrastructural morphology and collagen fibril profile of healing MCL in rats. STUDY DESIGN/MATERIALS AND METHODS: Thirty-two mature male Sprague-Dawley (SD) rats were used. Twenty-four underwent surgical transection to their right MCLs and eight received only skin wound. Immediately after surgery, eight of the MCL transected rats were treated with a single dose of laser therapy at 63.2 J cm(-2), eight were treated with a single dose of laser therapy at 31.6 J cm(-2), the rest had no treatment and served as control. At 3 and 6 weeks after surgery, the MCLs were harvested and examined with electron microscopy for collagen fibril size, distribution, and alignment. RESULTS: Significant differences (P < 0.001) were found in fibril diameters from the same anatomical site and time period among different groups. The mass-averaged diameters of the laser-treated (64.99-186.29 nm) and sham (64.74-204.34 nm) groups were larger than the control group (58.66-85.89 nm). The collagen fibrils occupied 42.55-59.78, 42.63-53.94, and 36.92-71.64% of the total cross-sectional areas in the laser-treated, control and sham groups, respectively. Mode obliquity was 0.53-0.84 among the three groups. CONCLUSIONS: Single application of low energy laser therapy increases the collagen fibril size of healing MCLs in rats.  相似文献   

15.
16.
目的 观察枝化状聚乙二醇对牛跟腱Ⅰ型胶原结构与构象稳定性的影响.方法 以牛跟腱Ⅰ型胶原为原料,以枝化状聚乙二醇为交联剂,在有效交联的前提下,通过测定羟脯氨酸含量观察聚乙二醇交联对胶原酶解稳定性的影响,采用示差扫描量热法(DSC)观察胶原热变性温度,采用圆二色光谱法(CD)观察聚乙二醇对胶原三螺旋构象稳定性的影响.结果 枝化状聚乙二醇交联组24 h降解率为26.79%,而对照组则达97.39%,两者比较差异有统计学意义(P<0.05).交联组热变性温度为(79.1±1.2)℃,与对照组(68.6±1.9)℃比较,差异有统计学意义(P<0.05);CD谱测定特征性吸收峰差异无统计学意义(P>0.05).结论 枝化状聚乙二醇可显著提高其热变形温度和抗酶性分解能力,而对其三螺旋结构与构象无明显不利影响,是较理想的生物高分子交联剂,可有望用于胶原分子的交联改性.
Abstract:
Objective The triple helix of collagen is the basis of its biological function, such as cell adhesion and tissue remodeling. Crosslinking of collagen with chemical agent will improve the biomechanical properties. However, the effects of differernt crosslinking agents on the structure and conformation stability of type Ⅰ collagen are rarely investigated. Methods The branched polyethylene glycol ( PEG)derivative ( MW 12 kD) was used as crosslinking agent, and allowed to react with bovine achilles' s tendon type Ⅰ collagen modified by succinimidylacetylthioacetate (SATA). The ability of resistance of crosslinked collagen to enzymatic degradation was investigated by measuring the release of hydroxyproline, and differential scanning calorimeter ( DSC ) was taken to determine the thermal denaturation temperature. The effect of PEG on the riple helix of collagen was studied by circular dichroism ( CD) . Results The resistance ability of PEG crosslinked collagen was strongly enhanced when compared with that of control group ( P <0. 05 ) , and the thermal denaturation temperature was also significantly rised. CD demonstrated that PEG crosslinking did not result in the destruction of the triple helix conformation of type Ⅰ collagen. Conclusion Branched PEG derivative used in this study is a promising polymer crosslinking agent that may be utilized in the modification of type Ⅰ collagen.  相似文献   

17.
18.
    
Tendon injuries are major orthopedic problems that worsen as the population ages. Type‐I (Col1) and type‐II (Col2) collagens play important roles in tendon midsubstance and tendon‐to‐bone insertion healing, respectively. Using double transgenic mice, this study aims to spatiotemporally monitor Col1 and Col2 gene expression, histology, and biomechanics up to 8 weeks following a full‐length patellar tendon injury. Gene expression and histology were analyzed weekly for up to 5 weeks while mechanical properties were measured at 1, 2, 5, and 8 weeks. At week 1, the healing region displayed loose granulation tissue with little Col1 expression. Col1 expression peaked at 2 weeks, but the ECM was highly disorganized and hypercellular. By 3 weeks, Col1 expression had reduced and by 5 weeks, the ECM was generally aligned along the tendon axis. Col2 expression was not seen in the healing midsubstance or insertion at any time point. The biomechanics of the healing tissue was inadequate at all time points, achieving ultimate loads and stiffnesses of 48% and 63% of normal values by 8 weeks. Future studies will further characterize the cells within the healing midsubstance and insertion using tenogenic markers and compare these results to those of tendon cells during normal development. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:28–36, 2012  相似文献   

19.
    
Type I collagen fibrils in tendons and ligaments assume a sinusoidal wave shape, or crimp, which straightens only with tensile load. The load response of crimp has been studied primarily in isolated subunits and not in complex, intact structures. The purpose of our study was to determine if freeze substitution fixation of an entire ligament could preserve changes in crimp morphology induced by functionally relevant loading conditions. We hypothesized that, in ligaments prepared by freeze-substitution fixation under load, crimp would progressively extinguish with increasing loads, and nonuniform strain following partial section could be detected from crimp morphology. Tensile loads ranging from 0 to 220 N were applied to patellar ligaments of 16 fresh rabbit stifle joints using simulated isometric quadriceps pull through the patella. The loaded joints were flash frozen with isopentane cooled in liquid nitrogen, then fixed using freeze substitution. Another six ligaments were loaded to 150 N following incision of the anterior third and evaluated under polarized light microscopy for crimp distribution. Ligaments with no or low loads could be identified by the presence of crimp on mid-sagittal sections. Strain distribution was inhomogeneous, in that the ligament displayed a consistent pattern of collagen fiber recruitment among three morphologically distinct bands seen on coronal sections. At very low loads (about 18 N), the fibers in a central band were uncrimped; anterior and deep bands uncrimped at higher loads. The crimp in the entire specimen was extinguished at about 67 N, which correlates closely with the previously reported toe-region of the stress-strain curve of the rabbit patellar ligament. When the anterior third was transected, fibers within that segment retained a crimp in ligaments prepared under loads that ordinarily would ablate all crimp. These findings suggest that freeze fixation could be used to map the functional microstructure of ligaments or tendons.  相似文献   

20.
    
The purpose of this study was to develop and validate an in vivo mouse model of tendon fatigue and use this model to investigate and quantify the physical manifestations of fatigue damage in mouse tendon. Patellar tendons of C57BL/6J mice were fatigue loaded at 2 Hz to three endpoints (4 N peak force per cycle for 1 h, 6 N for 1 h, and 4 N for 2 h), during which hysteresis, tangent stiffness, and peak strain of each cycle were measured. Damage accumulation was then quantified using in situ histology, and each tendon was loaded monotonically to failure. Histological damage increased significantly in all three groups (≥2-fold), and monotonic stiffness decreased significantly in the 6 N, 1 h and 4 N, 2-h groups (~25%), suggesting that damage initially manifests as changes to the collagen structure of the tendon and subsequently as changes to the function. For the fatigue loading protocols used in this study, none of the evaluated real-time parameters from fatigue loading correlated with damage area fraction measured structural damage or monotonic stiffness, suggesting that they are not suited to serve as proxies for damage accumulation. In future studies, this model will be used to compare the biological response of mouse tendon to fatigue damage across genetic strains.  相似文献   

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