Local administration of interleukin-1 receptor antagonist inhibits deterioration of mechanical properties of the stress-shielded patellar tendon

We previously found that interleukin (IL)-1beta is over-expressed in the fibroblasts of the stress-shielded patellar tendon using a stress-shielding model [Uchida, H., Tohyama, H., Nagashima, K., Ohba, Y., Matsumoto, H., Toyama, Y., Yasuda, K., 2005. Stress deprivation simultaneously induces over-expression of interleukin-1beta, tumor necrosis factor-alpha, and transforming growth factor-beta in fibroblasts and mechanical deterioration of the tissue in the patellar tendon. Journal of Biomechanics 38(4), 791-798.]. Therefore, IL-1beta may play a role in tendon deterioration in response to stress deprivation. This study was conducted to clarify the effects of local administration of interleukin-1 receptor antagonist (IL-1ra) on the mechanical properties of the stress-shielded patellar tendon as well as the tendon fascicles harvested from it. Twenty-six mature rabbits were equally divided into Groups IL-1ra and PBS after the right patellar tendon underwent the stress-shielding treatment, which completely released the patellar tendon from tension by stretching the flexible wire installed between the patella and the tibial tubercle. In Group IL-1ra, IL-1ra was injected between the patellar tendon and the infra-patellar fat pad. In Group PBS, phosphate-buffered saline was injected in the same manner as IL-1ra. All rabbits were evaluated at 3 weeks after the stress-shielding procedure. The tangent modulus and the tensile strength of the patellar tendons were significantly greater in Group IL-1ra than in Group PBS, while there was no significant difference in the strain at failure between Groups IL-1ra and PBS. Concerning the mechanical properties of the fascicles harvested from the patellar tendon, however, we could not detect any significant differences in the tangent modulus, tensile strength, or strain at failure between Groups IL-1ra and PBS. The present study suggested that IL-1 plays an important role in the deterioration of the mechanical properties of the patellar tendon in response to stress shielding and that IL-1 does not affect the fascicles themselves.     

Effects of administration of transforming growth factor (TGF)-beta1 and anti-TGF-beta1 antibody on the mechanical properties of the stress-shielded patellar tendon

Previous studies have shown that, in the stress-shielded patellar tendon, the mechanical properties of the tendon were dramatically reduced and TGF-beta was over-expressed in tendon fibroblasts. In the present study, therefore, we tested two supportive hypotheses using 40 rabbits: One was that an application of TGF-beta1 might significantly increase the tensile strength and the tangent modulus of the stress-shielded patellar tendon. The other one was that an administration of anti-TGF-beta1 antibody might significantly reduce the mechanical properties of the stress-shielded patellar tendon. In the results, an application of 4-ng TGF-beta1 significantly increased the tangent modulus of the stress-shielded patellar tendon at 3 weeks (p = 0.019), compared with the sham treatment. Concerning the tensile strength, the 4-ng TGF-beta1 application increased the average value, but a statistical significance was not reached. An application of 50-microg anti-TGF-beta1 antibody significantly reduced the tangent modulus and the tensile strength of the stress-shielded patellar tendon at 3 weeks (p = 0.0068 and p = 0.0355), compared with the sham treatment. Because the stress-shielding treatment used in this study dramatically reduces the tangent modulus and the tensile strength of the patellar tendon, the present study suggested that an administration of TGF-beta1 weakly but significantly inhibited the reduction of the mechanical properties of the stress-shielded patellar tendon, and that inactivation of TGF-beta1 with its antibody significantly enhanced the reduction of the mechanical properties that occurs in the stress-shielded patellar tendon. These results suggested that TGF-beta1 plays an important role in remodeling of the stress-shielded patellar tendon.     

Effects of stress shielding on the transverse mechanical properties of rabbit patellar tendons

With the aim of studying mechanisms of the remodeling of tendons and ligaments, the effects of stress shielding on the rabbit patellar tendon were studied by performing tensile and stress relaxation tests in the transverse direction. The tangent modulus, tensile strength, and strain at failure of non-treated, control patellar tendons in the transverse direction were 1272 kPa, 370 kPa, and 40.5 percent, respectively, whereas those of the tendons stress-shielded for 1 week were 299 kPa, 108 kPa, and 40.4 percent, respectively. Stress shielding markedly decreased tangent modulus and tensile strength in the transverse direction, and the decreases were larger than those in the longitudinal direction, which were determined in our previous study. For example, tensile strength in the transverse and longitudinal direction decreased to 29 and 50 percent of each control value, respectively, after 1 week stress shielding. In addition, the stress relaxation in the transverse direction of stress-shielded patellar tendons was much larger than that of nontreated, control ones. In contrast to longitudinal tensile tests for the behavior of collagen, transverse tests reflect the contributions of ground substances such as proteoglycans and mechanical interactions between collagen fibers. Ground substances provide lubrication and spacing between fibers, and also confer viscoelastic properties. Therefore, the results obtained from the present study suggest that ground substance matrix, and interfiber and fiber-matrix interactions have important roles in the remodeling response of tendons to stress.     

Antibody neutralization of TGF-beta enhances the deterioration of collagen fascicles in a tissue-cultured tendon matrix with ex vivo fibroblast infiltration

A tissue-cultured tendon matrix infiltrated with cultured fibroblasts can be regarded as an ideal tissue-engineered tendon model. To clarify the role of TGF-beta in a tissue-cultured tendon matrix during ex vivo cellular infiltration, the present ex vivo study was conducted to test the following hypothesis that antibody neutralization of TGF-beta enhances weakening of the collagen fascicles of the patellar tendon matrix in response to ex vivo fibroblast infiltration. In skeletally mature female rabbits, fibroblasts were isolated from the right patellar tendons using an explant culture technique, and the left patellar tendons underwent multiple freeze/thaw treatment with liquid nitrogen to obtain an acellular tendon matrix. Each acellular tendon was placed in a collagen gel containing cultured fibroblasts and then incubated with or without anti-TGF-beta1 antibody for 6 weeks. We found that antibody neutralization of TGF-beta enhanced the decrease in the tensile strength and tensile modulus of the collagen fascicles of the patellar tendon matrix in response to ex vivo fibroblast infiltration. The present study indicates a possibility that TGF-beta may have a role in suppressing the material deterioration of the fascicles in the tendon during ex vivo cellular infiltration.     

Stress deprivation from the patellar tendon induces apoptosis of fibroblasts in vivo with activation of mitogen-activated protein kinases

The effect of stress deprivation on the tendon tissue has been an important focus in the field of biomechanics. However, less is known about the in vivo effect of stress deprivation on fibroblast apoptosis as of yet. This study was conducted to test a hypothesis that complete stress deprivation of the patellar tendon induces fibroblast apoptosis in vivo with activation of Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38) within 24 h after treatment. A total of 35 mature rabbits were divided into stress-shielded (n=15), sham-operated (n=15), and control (n=5) groups. To completely shield the patellar tendon from stress, we used an established surgical method. Animals were sacrificed at 24 h, and 2, 4, 7, and 14 days after the treatment. Tendon specimens underwent TUNEL assay and immunohistological examinations of active caspase-3, JNK, and p38. Both the number and the ratio of TUNEL-positive and caspase-3-positive cells were significantly greater (p<0.0001) in the stress-shielded group than in the sham group at 24 h, 2, 4, and 7 days. Concerning JNK and p38, both the number and the ratio were significantly greater (p<0.0001) in the stress-shielded group than in the sham group at 24 h, 2, and 4 days. This study demonstrated that complete stress deprivation induces fibroblast apoptosis in vivo with activation of JNK and p38 within 24 h. This fact suggested that the fibroblast apoptosis caused by stress deprivation is induced via the mitogen-activated protein kinase signaling pathway.     

Effects of cyclic stress on the mechanical properties of cultured collagen fascicles from the rabbit patellar tendon

Effects of cyclic stress on the mechanical properties of collagen fascicles were studied by in vitro tissue culture experiments. Collagen fascicles (approximately 300 microns in diameter) obtained from the rabbit patellar tendon were applied cyclic load at 4 Hz for one hour per day during culture period for one or two weeks, and then their mechanical properties were determined using a micro-tensile tester. There was a statistically significant correlation between tensile strength and applied peak stress in the range of 0 to 5 MPa, and the relation was expressed by a quadratic function. The maximum strength (19.4 MPa) was obtained at the applied peak stress of 1.8 MPa. The tensile strength of fascicles were within a range of control values, if they were cultured under peak stresses between 1.1 and 2.6 MPa. Similar results were also observed in the tangent modulus, which was maintained at control level under applied peak stresses between 0.9 and 2.8 MPa. The stress of 0.9 to 1.1 MPa is equivalent to approximately 40% of the in vivo peak stress which is developed in the intact rabbit patellar tendon by running, whereas that of 2.6 to 2.8 MPa corresponds to approximately 120% of the in vivo peak stress. Therefore, the fascicles cultured under applied peak stresses of lower than 40% and higher than 120% of the in vivo peak stress do not keep the original strength and modulus. These results indicate that the mechanical properties of cultured collagen fascicles strongly depend upon the magnitude of the stress applied during culture, which are similar to our previous results observed in stress-shielded and overstressed patellar tendons in vivo.     

Human patellar tendon stiffness is restored following graft harvest for anterior cruciate ligament surgery

Minimising post-operative donor site morbidity is an important consideration when selecting a graft for surgical reconstruction of the torn anterior cruciate ligament (ACL). One of the most common procedures, the bone-patellar tendon-bone (BPTB) graft involves removal of the central third from the tendon. However, it is unknown whether the mechanical properties of the donor site (patellar tendon) recover. The present study investigated the mechanical properties of the human patellar tendon in 12 males (mean±S.D. age: 37±14 years) who had undergone surgical reconstruction of the ACL using a BPTB graft between 1 and 10 years before the study (operated knee; OP). The uninjured contralateral knee served as a control (CTRL). Patellar tendon mechanical properties were assessed in vivo combining dynamometry with ultrasound imaging. Patellar tendon stiffness was calculated from the gradient of the tendon's force–elongation curve. Tendon stiffness was normalised to the tendon's dimensions to obtain the tendon's Young's modulus. Cross-sectional area (CSA) of OP patellar tendons was larger by 21% than CTRL tendons (P<0.01). Patellar tendon stiffness was not significantly different between OP and CTRL tendons, but the Young's modulus was lower by 24% in OP tendons (P<0.01). A compensatory enlargement of the patellar tendon CSA, presumably due to scar tissue formation, enabled a recovery of tendon stiffness in the OP tendons. The newly formed tendon tissue had inferior properties as indicated by the reduced tendon Young's modulus, but it increased to a level that enabled recovery of tendon stiffness.     

Regulation of rat pulmonary artery endothelial cell transforming growth factor-beta production by IL-1 beta and tumor necrosis factor-alpha.

Recent studies suggest that transforming growth factor-beta (TGF-beta) production is up-regulated at sites of tissue injury, inflammation and repair, or fibrosis. Endothelial cells represent a potentially important in vivo source of TGF-beta; however, the identity of endogenous modulators of TGF-beta production by these cells remains unclear. To address this issue, the effects of the cytokines, IL-1 beta, and TNF-alpha on TGF-beta production by rat pulmonary artery endothelial cells were examined. Conditioned media from cells treated with 0 to 20 ng/ml IL-1 beta and/or TNF-alpha were assayed for TGF-beta activity using a mink lung epithelial cell line. The results show that rat pulmonary artery endothelial cells secreted undetectable amounts of active TGF-beta in the absence of cytokines. However, upon acidification of the conditioned media before assay, a time-dependent increase in TGF-beta activity was noted in media from both untreated and cytokine-treated cells. However, both IL-1 beta and TNF-alpha treatment caused the secretion of significantly greater amounts of TGF-beta activity than control cells, in a dose-dependent manner, with maximal response obtained at cytokine doses of greater than 10 ng/ml. At equivalent doses of cytokine tested, the magnitude of the response was significantly greater with IL-1 beta. These responses were paralleled by increases in steady state mRNA levels for TGF-beta 1. Addition of both cytokines resulted in a synergistic response. Synergism with IL-1 beta was also noted with the fibrogenic agent bleomycin. Kinetic studies indicated that a minimum of 4 h of treatment with either IL-1 beta or TNF-alpha was required for detection of significant increases in either secreted TGF-beta activity or steady state TGF-beta 1 mRNA levels. Thus, endothelial cells could play a role in various TGF-beta-dependent processes in vivo, in situations wherein IL-1 beta and/or TNF-alpha may be present at comparable concentrations.     

Biomechanical studies of the rabbit patellar tendon after removal of its one-fourth or a half.

Effects of the overstressing induced by the harvest of grafts from the patellar tendon on the mechanical properties and morphometry of remaining tendon were studied using a rabbit model. The width of the patellar tendon was reduced by one-fourth or one-half equally removing the medial and lateral portions; by this surgery, the cross-sectional area was decreased by 25 or 50 percent from the original area. After all the rabbits were allowed unrestricted activities in cages for 3 to 12 weeks, their patellar tendons were harvested for mechanical and histological studies. The one-fourth removal induced no significant changes in the mechanical properties, but significantly increased the cross-sectional area. In the case of one-half removal, tensile strength and tangent modulus did not change in some tendons, although the cross-sectional area increased significantly. In the other central half tendons, mechanical strength decreased markedly, while the cross-sectional area increased; hypercellular areas and breakage of collagen bundles were observed in these tendons. These results indicate that the patellar tendon has an ability of functionally adapting to overstressing by changing the cross-sectional area, while keeping the mechanical properties unchanged, if the extent of overstressing is less than 30 percent.     

Flexor tendon wound healing in vitro: lactate up-regulation of TGF-beta expression and functional activity

Flexor tendon wound healing in zone II is complicated by adhesions to the surrounding fibro-osseous sheath. These adhesions can significantly alter tendon gliding and ultimately hand function. Lactate and transforming growth factor-beta (TGF-beta) are two important mediators of wound healing that have been demonstrated to independently increase collagen production by cells of the tendon sheath, epitenon, and endotenon. This study examined the effects of lactate on TGF-beta peptide and receptor production by flexor tendon cells. Tendon sheath fibroblasts, epitenon tenocytes, and endotenon tenocytes were isolated from rabbit flexor tendons and cultured separately. Cell cultures were supplemented with 50 mM lactate, and the expression of three TGF-beta peptide isoforms (beta1, beta2, and beta3) and three receptor isoforms (R1, R2, and R3) was quantified with enzyme-linked immunosorbent assays. TGF-beta functional activity was also assessed with the addition of tendon cell conditioned media to mink lung epithelial cells transfected with a luciferase reporter gene expression construct responsive to TGF-beta. Supplementation of the cell culture medium with lactate significantly (p < 0.05) increased the expression of all TGF-beta peptide and receptor isoforms in all three cell lines. Tendon sheath fibroblasts exhibited the greatest increases in beta1 and beta2 peptide isoform expression (30 and 23 percent, respectively), whereas endotenon tenocytes demonstrated the greatest increase in beta3 peptide expression (32 percent). Epitenon tenocytes exhibited the greatest increases in receptor isoform R1 and R2 expression (17 and 19 percent, respectively). All three tendon cell types demonstrated significant (p < 0.05) increases in TGF-beta functional activity when exposed to lactate. Epitenon tenocytes demonstrated the greatest increase in activity (>4 times control values), whereas tendon sheath fibroblasts demonstrated the highest overall levels of total TGF-beta functional activity. Lactate significantly increased TGF-beta peptide (beta1, beta2, and beta3) expression, receptor (R1, R2, and R3) expression, and functional activity, suggesting a common pathway regulating tendon cell collagen production. Modulation of lactate and TGF-beta levels may provide a means of modulating the effects of TGF-beta on adhesion formation in flexor tendon wound healing.     

ATP modulates load-inducible IL-1beta,COX 2, and MMP-3 gene expression in human tendon cells

Tendon cells receive mechanical signals from the load bearing matrices. The response to mechanical stimulation is crucial for tendon function. However, overloading tendon cells may deteriorate extracellular matrix integrity by activating intrinsic factors such as matrix metalloproteinases (MMPs) that trigger matrix destruction. We hypothesized that mechanical loading might induce interleukin-1beta (IL-1beta) in tendon cells, which can induce MMPs, and that extracellular ATP might inhibit the load-inducible gene expression. Human tendon cells isolated from flexor digitorum profundus tendons (FDPs) of four patients were made quiescent and treated with ATP (10 or 100 microM) for 5 min, then stretched equibiaxially (1 Hz, 3.5% elongation) for 2 h followed by an 18-h-rest period. Stretching induced IL-1beta, cyclooxygenase 2 (COX 2), and MMP-3 genes but not MMP-1. ATP reduced the load-inducible gene expression but had no effect alone. A medium change caused tendon cells to secrete ATP into the medium, as did exogenous UTP. The data demonstrate that mechanical loading induces ATP release in tendon cells and stimulates expression of IL-1beta, COX 2, and MMP-3. Load-induced endogenous IL-1beta may trigger matrix remodeling or a more destructive pathway(s) involving IL-1beta, COX 2, and MMP-3. Concomitant autocrine and paracrine release of ATP may serve as a negative feedback mechanism to limit activation of such an injurious pathway. Attenuation or failure of this negative feedback mechanism may result in the progression to tendinosis.     

TGF-β1, TGF-β3, and PGE<Subscript>2</Subscript> regulate contraction of human patellar tendon fibroblasts

To understand the role of tendon fibroblast contraction in tendon healing, we investigated the contraction of human patellar tendon fibroblasts (HPTFs) and its regulation by transforming growth factor-beta1 (TGF-beta1), TGF-beta3, and prostaglandin E(2) (PGE(2)). HPTFs were found to wrinkle the underlying thin silicone membranes, demonstrating that these tendon fibroblasts are contractile. Using fibroblast populated collagen gels (FPCGs), exogenous addition of TGF-beta1 or TGF-beta3 was found to increase fibroblast contraction compared to non-treated fibroblasts in serum-free medium, whereas PGE(2) was found to decrease the tendon fibroblast contraction. Moreover, the tendon fibroblasts in collagen gels treated with TGF-beta1 contracted to a greater degree than those treated with TGF-beta3. Since the extent of fibroblast contraction is related to scar tissue formation, this differential effect of TGF-beta1 and TGF-beta3 on HPTF contraction supports the previous finding that TGF-beta1 induces scar tissue formation, whereas TGF-beta3 reduces its formation. Further, the reduced tendon fibroblast contraction by PGE(2) suggests that excessive presence of this inflammatory mediator in the wound site might retard tendon healing. Taken together, the results of this study suggest that regulation of human tendon fibroblast contraction may reduce scar tissue formation and therefore improve the mechanical properties of healing tendons.     

Effects of stress shielding and subsequent restressing on mechanical properties of regenerated and residual tissues in rabbit patellar tendon after resection of its central one-third

Central third of patellar tendon (PT) is used as an autograft for anterior cruciate ligament (ACL) reconstruction. Previous studies investigated temporal changes in material properties of healing tissues in PT after resection of the central third. However, no study has been performed on effects of stress shielding (SS) and restressing (RS) on the properties of healing tissues. The present study hypothesised that SS adversely affects the mechanical integrity of healing tissues, which is recovered by subsequent RS. An entire rectangular defect was created in the central third of rabbit PT. Operated PTs were subjected to either SS or no stress shielding (NSS). A subgroup of stress-shielded PTs was followed by the resumption of normal loading, namely RS. Tensile properties of tissues regenerated in the defect and residual tendons were evaluated. Regenerated tissues of SS for 3 weeks resulted in significantly lower strength than NSS, which was recovered to NSS level by 3 weeks of RS. Strength of residual tissues in RS reversed SS effects, leading to the strength at NSS level after 12 weeks. However, tangent modulus of residual tissues in RS was still significantly lower than that of NSS at 12 weeks. Therefore, SS induces detrimental effects on the mechanical integrity of healing PTs, and the response to RS was different between regenerate and residual tissues, the latter of which took longer period to reach NSS level.     

Inflammatory cytokines augments TGF-beta1-induced epithelial-mesenchymal transition in A549 cells by up-regulating TbetaR-I

Epithelial-mesenchymal transition (EMT) is believed to play an important role in fibrosis and tumor invasion. EMT can be induced in vitro cell culture by various stimuli including growth factors and matrix metalloproteinases. In this study, we report that cytomix (a mixture of IL-1beta, TNF-alpha and IFN-gamma) significantly enhances TGF-beta1-induced EMT in A549 cells as evidenced by acquisition of fibroblast-like cell shape, loss of E-cadherin, and reorganization of F-actin. IL-1beta or TNF-alpha alone can also augment TGF-beta1-induced EMT. However, a combination of IL-1beta and TNF-alpha or the cytomix is more potent to induce EMT. Cytomix, but not individual cytokine of IL-1beta, TNF-alpha or IFN-gamma, significantly up-regulates expression of TGF-beta receptor type I (TbetaR-I). Suppression of TbetaR-I, Smad2 or Smad3 by siRNA partially blocks EMT induction by cytomix plus TGF-beta1, indicating cytomix augments TGF-beta1-induced EMT through enhancing TbetaR-I and Smad signaling. These results indicate that inflammatory cytokines together with TGF-beta1 may play an important role in the development of fibrosis and tumor progress via the mechanism of epithelial-mesenchymal transition.     

Orientation of tendons in vivo with active and passive knee muscles

Tendon orientations in knee models are often taken from cadaver studies. The aim of this study was to investigate the effect of muscle activation on tendon orientation in vivo. Magnetic resonance imaging (MRI) images of the knee were made during relaxation and isometric knee extensions and flexions with 0 degrees , 15 degrees and 30 degrees of knee joint flexion. For six tendons, the orientation angles in sagittal and frontal plane were calculated. In the sagittal plane, muscle activation pulled the patellar tendon to a more vertical orientation and the semitendinosus and sartorius tendons to a more posterior orientation. In the frontal plane, the semitendinosus had a less lateral orientation, the biceps femoris a more medial orientation and the patellar tendon less medial orientation in loaded compared to unloaded conditions. The knee joint angle also influenced the tendon orientations. In the sagittal plane, the patellar tendon had a more anterior orientation near full extension and the biceps femoris had an anterior orientation with 0 degrees and 15 degrees flexions and neutral with 30 degrees flexions. Within 0 degrees to 30 degrees of flexion, the biceps femoris cannot produce a posterior shear force and the anterior angle of the patellar tendon is always larger than the hamstring tendons. Therefore, co-contraction of the hamstring and quadriceps is unlikely to reduce anterior shear forces in knee angles up to 30 degrees . Finally, inter-individual variation in tendon angles was large. This suggests that the amount of shear force produced and the potential to counteract shear forces by co-contraction is subject-specific.     

Ethanol induces the production of cytokines via the Ca2+, MAP kinase, HIF-1alpha, and NF-kappaB pathway

In the present study, we sought to investigate the signal transduction pathways of expression of cytokines in the ethanol-stimulated human mast cell line, HMC-1. Ethanol significantly increased the intracellular calcium level in HMC-1. Ethanol also significantly enhanced IL-6, TNF-alpha, and TGF-beta1 production compared with media control, but did not significantly affect the IL-1beta production. After 8 h of stimulation, ethanol increased mRNA and protein expression levels of TNF-alpha and TGF-beta1 in HMC-1. The increased cytokine level was significantly inhibited by BAPTA-AM, PD98059, and SB203580. These inhibitors also inhibited ethanol-induced ERK and p38 MAPK phosphorylation. Ethanol resulted in a great increase in protein levels and promoter activity driving luciferase expression of HIF-1alpha and NF-kappaB in HMC-1 cells, but it did not affect on HIF-1alpha mRNA expression. Our observations show that calcium, MAPK activation, HIF-1alpha, and NF-kappaB are necessary for ethanol-induced TNF-alpha and TGF-beta1 expression. These results may have important implications for the study of alcohol-related diseases.     

Non-uniformity in the healthy patellar tendon is greater in males and similar in different age groups

There is increasing evidence that tendons are heterogeneous and take advantage of structural mechanisms to enhance performance and reduce injury. Fascicle-sliding, for example, is used by energy-storing tendons to enable them to undergo large extensions while protecting the fascicles from damage. Reductions in fascicle-sliding capacity may thus predispose certain populations to tendinopathy. Evidence from the Achilles tendon of significant superficial-to-deep non-uniformity that is reduced with age supports this theory. Similar patellar tendon non-uniformity has been observed, but the effects of age and sex have yet to be assessed. Healthy adults (n = 50, 25M/25F) from a broad range of ages (23–80) were recruited and non-uniformity was quantified using ultrasound speckle-tracking during passive knee extension. Significant superficial-to-deep non-uniformity and proximal/distal variations were observed. No effect of age was found, but males exhibited significantly greater non-uniformity than females (p < 0.05). The results contrast with previous findings in the Achilles tendon; in this study, tendons and tendon regions at high risk for tendinopathy (i.e. males and proximal regions, respectively) exhibited greater non-uniformity, whereas high-risk Achilles tendons (i.e. older adults) previously showed reduced non-uniformity. This suggests that non-uniformity may be dominated by factors other than fascicle-sliding. Anatomically, the varied proximal attachment of the patellar tendon may influence non-uniformity, with quadriceps passive resistance limiting superficial tendon movement, thus linking flexibility, non-uniformity and injury risk. This study also provides evidence of a differential effect of aging on the patellar tendon compared with evidence from prior studies on other tendons necessitating further study to elucidate links between non-uniformity and injury.     

Influence of decorin and biglycan on mechanical properties of multiple tendons in knockout mice

Evaluations of tendon mechanical behavior based on biochemical and structural arrangement have implications for designing tendon specific treatment modalities or replacement strategies. In addition to the well studied type I collagen, other important constituents of tendon are the small proteoglycans (PGs). PGs have been shown to vary in concentration within differently loaded areas of tendon, implicating them in specific tendon function. This study measured the mechanical properties of multiple tendon tissues from normal mice and from mice with knock-outs of the PGs decorin or biglycan. Tail tendon fascicles, patellar tendons (PT), and flexor digitorum longus tendons (FDL), three tissues representing different in vivo loading environments, were characterized from the three groups of mice. It was hypothesized that the absence of decorin or biglycan would have individual effects on each type of tendon tissue. Surprisingly, no change in mechanical properties was observed for the tail tendon fascicles due to the PG knockouts. The loss of decorin affected the PT causing an increase in modulus and stress relaxation, but had little effect on the FDL. Conversely, the loss of biglycan did not significantly affect the PT, but caused a reduction in both the maximum stress and modulus of the FDL. These results give mechanical support to previous biochemical data that tendons likely are uniquely tailored to their specific location and function. Variances such as those presented here need to be further characterized and taken into account when designing therapies or replacements for any one particular tendon.     

Enhancement by transforming growth factor-beta 1 (TGF-beta 1) of the proliferation of leukemic blast progenitors stimulated with IL-3.

We studied the effect of transforming growth factor-beta 1 (TGF-beta 1) on colony formation of leukemic blast progenitors from ten acute myeloblastic leukemia (AML) patients stimulated with granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), interleukin-6 (IL-6), or interleukin-1 beta (IL-1 beta). These CSFs and interleukins by themselves stimulated the proliferation of leukemic blast progenitors without adding TGF-beta 1. G-CSF, GM-CSF, and IL-3 stimulated blast colony formation in nine patients, IL-6 stimulated it in five, and IL-1 beta stimulated in four. TGF-beta 1 significantly reduced blast colony formation stimulated by G-CSF, GM-CSF, or IL-6 in all patients. In contrast, TGF-beta 1 enhanced the stimulatory effect of IL-3 on blast progenitors from three cases, while in the other seven patients TGF-beta 1 reduced blast colony formation in the presence of IL-3. To study the mechanism by which TGF-beta 1 enhanced the stimulatory effect of IL-3 on blast progenitors, we carried out the following experiments in the three patients in which it occurred. First, the media conditioned by leukemic cells in the presence of TGF-beta 1 stimulated the growth of leukemic blast progenitors, but such effect was completely abolished by anti-IL-1 beta antibody. Second, the addition of IL-1 beta in the culture significantly enhanced the growth of blast progenitors stimulated with IL-3. Third, leukemic cells of the two patients studied were revealed to secrete IL-1 beta and tumor necrosis factor-alpha (TNF-alpha) constitutively; the production by leukemic cells of IL-1 beta and TNF-alpha was significantly promoted by TGF-beta 1. Furthermore, the growth enhancing effect of TGF-beta 1 in the presence of IL-3 was fully neutralized by anti-IL-1 beta antibody. These findings suggest that TGF-beta 1 stimulated the growth of blast progenitors through the production and secretion of IL-1 beta by leukemic cells.