Changes In Glycosaminoglycans Research Articles

Shining Light on Osteoarthritis: Spatially Offset Raman Spectroscopy as a Window into Cartilage Health.

Articular cartilage is a complex tissue, and early detection of osteoarthritis (OA) is crucial for effective treatment. However, current imaging modalities lack molecular specificity and primarily detect late-stage changes. In this study, we propose the use of spatially offset Raman spectroscopy (SORS) for noninvasive, depth-dependent, and molecular-specific diagnostics of articular cartilage. We demonstrate the potential of SORS to penetrate deep layers of cartilage, providing a comprehensive understanding of disease progression. Our SORS measurements were characterized and validated through mechanical and histological techniques, revealing strong correlations between spectroscopic measurements and both Young's modulus and depth of cartilage damage. By longitudinally monitoring enzymatically degraded condyles, we further developed a depth-dependent damage-tracking method. Our analysis revealed distinct components related to sample depth and glycosaminoglycan (GAG) changes, offering a comprehensive picture of cartilage health. Collectively, these findings highlight the potential of SORS as a valuable tool for enhancing OA management and improving patient outcomes.

A role for decorin in improving motor deficits after traumatic brain injury

Traumatic brain injury (TBI) is the leading cause of death and disability due to injury worldwide. Extracellular matrix (ECM) remodeling is known to significantly contribute to TBI pathophysiology. Glycosaminoglycans, which are long-chain, variably sulfated polysaccharides abundant within the ECM, have previously been shown to be substantially altered after TBI. In this study, we sought to delineate the dynamics of glycosaminoglycan alterations after TBI and discover the precise biologic processes responsible for observed glycosaminoglycan changes after injury. We performed state-of-the art mass spectrometry on brain tissues isolated from mice after TBI or craniotomy-alone. We observed dynamic changes in glycosaminoglycans at Day 1 and 7 post-TBI, with heparan sulfate, chondroitin sulfate, and hyaluronan remaining significantly increased after a week vis-à-vis craniotomy-alone tissues. We did not observe appreciable changes in circulating glycosaminoglycans in mice after experimental TBI compared to craniotomy-alone nor in patients with TBI and severe polytrauma compared to control patients with mild injuries, suggesting increases in injury site glycosaminoglycans are driven by local synthesis. We subsequently performed an unbiased whole genome transcriptomics analysis on mouse brain tissues 7 days post-TBI and discovered a significant induction of hyaluronan synthase 2, glypican-3, and decorin. The functional role of decorin after injury was further examined through multimodal behavioral testing comparing wild-type and Dcn−/− mice. We discovered that genetic ablation of Dcn led to an overall negative effect of TBI on function, exacerbating motor impairments after TBI. Collectively, our results provide a spatiotemporal characterization of post-TBI glycosaminoglycan alterations in the brain ECM and support an important adaptive role for decorin upregulation after TBI.

Biomaterial considerations for ovarian cancer models

Ovarian cancer is the 5th most common and the deadliest gynecological cancer, with a 5-year survival rate of less than 50 percent. Most deaths due to ovarian cancer are caused by recurrent disease, which typically corresponds to an increase in chemoresistance of the tumor cells. However, little is known about how ovarian tumor chemoresponse changes and if such changes are regulated by the tumor microenvironment (TME). Moreover, the ovarian TME, including the tissue compositions and biomechanical features, is not well-characterized primarily due to a lack of optimal models. To more effectively characterize the TME of ovarian cancer, which may help develop innovative treatment strategies, appropriate models are desperately needed. The most utilized models include mouse models with both patient-derived xenografts and mouse or human tumor cell line derivatives, and more recently microphysiological systems (MPS). While mouse models provide high levels of physiological complexity, there is virtually no control over the TME components after tumor initiation or implantation. On the other hand, MPS or organoid models permit high levels of control of initial composition but lack many features of in vivo models. Selection of appropriate components to create a TME model is paramount for generating a physiologically relevant in vitro and ex vivo systems. The importance of biomaterial or matrix selection in ovarian TME models lies in the role of these components to activate oncogenic signaling pathways either through receptor-ligand interactions or mechanotransduction. Recent studies suggest that off-target or post-target effects of chemotherapies may interfere with mechanotransductive pathways. In ovarian cancer, changes in fibrous proteins, adhesive glycoproteins, and glycosaminoglycans can remodel the mechanical environment, further altering mechanotransductive pathways. Therefore, the next-generation of ovarian tumor models should incorporate relevant biomaterials including hyaluronic acid (HA), collagens, fibrinogen, and fibronectin to investigate the link between matrix properties and mechanobiology with metastasis and chemoresistance.

PECULIARITIES OF CONNECTIVE TISSUE METABOLISM IN PATIENTS WITH POST-INFARCTION REMODELING AND REPERFUSION SYNDROME

Early restoration of coronary blood flow in acute myocardial infarction with elevation helps reduce the area of myocardial necrosis, prevent dilatation of the ventricular cavity, and reduce the development of arrhythmias and mortality. However, despite modern technologies in STEMI treatment, the development of reperfusion syndrome, in particular, arrhythmias, is possible after the restoration of blood flow in the infarct-related artery, which is a consequence of acute cellular, metabolic and local electrophysiological changes in the myocardium.
 The aim: is to investigate the long-term effects of reperfusion syndrome on the catabolic/anabolic activity of connective tissue and the processes of connective tissue framework formation in the post-infarction period in patients with myocardial infarction.
 Materials and methods. The study involved 156 patients with acute myocardial infarction with ST segment elevation (STEMI), and 15 apparently healthy individuals (control of laboratory parameters). The average age of the patients was (57.8±1.2) years. The percent of male patients prevailed (76 %).
 All MI patients were divided into two groups: Group I (98 people) – MI patients without clinical signs of reperfusion syndrome during revascularization, Group II (58 people) – MI patients with the signs of reperfusion syndrome in the form of rhythm and conductivity disturbances during revascularization of coronary arteries by percutaneous coronarography intervention (PCI) with prolonged stenting of the infarct-dependent coronary artery.
 Peculiarities of connective tissue metabolism were studied by the content of free and bound oxyproline, glycosaminoglycans, and antibodies to atypical collagens (Ig M and Ig G) in blood serum.
 Results. The recovery period after myocardial infarction showed an increase in collagen catabolism over collagen synthesizing processes in patients with MI who had reperfusion syndrome. The described patterns were also established by changes in glycosaminoglycans. The study showed an increase in the level of antibodies to atypical collagens. Particularly, the level of Ig M, which was within the norm (0.43±0.18) ng/ml, in patients with MI without reperfusion manifestations was three times higher (p<0.001), and in cases of combination of MI and reperfusion syndrome – the highest (p<0.001).
 Conclusions:
 
 Changes in collagen-synthesizing processes are observed in patients with MI, with a tendency to increase the processes of catabolism of the connective tissue matrix.
 The presence of reperfusion syndrome manifestations in MI patients after revascularization aggravates the processes of catabolism, even in the recovery period after MI.
 Increase in the activity of catabolic processes stimulates fibrosis and formation of rigid matrix and atypical collagen, to which antibodies are synthesized, thus blocking the pathological circuit of restructuring of the heart connective tissue framework, and accordingly its pathological remodeling.
 The obtained study results contribute to the identification of more sophisticated approaches to revascularization techniques to avoid negative effects of reperfusion syndrome, as well as new effective medications for collagen synthesis correction, which can increase the effectiveness of rehabilitative management of such

Biomechanisms of cervical remodeling and current approaches to maturity assessment

The cervical remodeling process is unique and consists of softening, ripening, dilation at fetal birth, and repair to the original state, which is a dense long tubular structure. In this review, the problem of impaired cervical remodeling is discussed in both preterm birth and delayed transformation, which leads to the unpreparedness of the cervix for childbirth and prolongation of pregnancy. Histological, immunological and structural dynamic changes in the cervix begin long before delivery and are noted as early as the first trimester of gestation. There are a few ways to assess the maturity of the cervix. In the second and third trimester, in order to predict preterm birth, these are ultrasound cervicometry and a cervical phosphorylated insulin-like growth factor binding protein-1 test. At full term, in order to determine its readiness for delivery, this is a palpation assessment. Inadequate assessment of the cervical characteristics is one of the factors of untimely prevention of preterm birth, and at full term leads to inappropriate choice of method of preparation for labor. It is necessary to develop new approaches to the comprehensive assessment of the cervix, using existing methods, and to discover new ways to assess its maturity.
 In this review, the problem of cervical maturation diagnosis is considered based on literature data from such databases as PubMed, ResearchGate, and Google Scholar, as well as from electronic resources of the M. Gorky Scientific Library (St. Petersburg State University, Russia). This review analyzes data on molecular, biochemical and histophysiological processes occurring during cervical maturation at all stages of gestation.
 It is generally accepted that the main role in cervical changes at all stages of gestation is played by: collagen fiber restructuring / desorganization, decreased concentrations of collagen and elastin, high molecular weight hylauronic acid cleavage, increased aquaparin level and tissue hydrophilicity, increased cervical vascularization, as well as changes in glycosaminoglycan and matrix metalloproteinase content. Palpatory technique and ultrasound cervicometry are the most common methods of determining the cervical length, which have insufficient sensitivity, probably because they do not cover all pathogenetic pathways of remodeling and cannot assess all cervical characteristics. Improvement of efficiency is possible through the introduction of combined techniques and the use of promising methods such as elastography, ultrasound diagnosis of the cervix with Doppler assessment of its vessels, determination of a disintegrin and metalloprotease with thrombospondin-like repeats-1 and placental 1-microglobulin in cervical secretion, and relaxin in maternal blood.
 Understanding the molecular, biochemical and histophysiological processes that occur during cervical remodeling is crucial for predicting preterm birth, diagnosing isthmic-cervical insufficiency, understanding the lack of timely cervical readiness, and choosing tactics the method of preinduction and induction of labor if necessary. The lack of clinical methods and their lack of objectivity require a combined approach and the search for new prognostic markers of cervical maturation.

Effect of high glucose on glycosaminoglycans in cultured retinal endothelial cells and rat retina.

The endothelial glycocalyx regulates vascular permeability, inflammation, and coagulation, and acts as a mechanosensor. The loss of glycocalyx can cause endothelial injury and contribute to several microvascular complications and, therefore, may promote diabetic retinopathy. Studies have shown a partial loss of retinal glycocalyx in diabetes, but with few molecular details of the changes in glycosaminoglycan (GAG) composition. Therefore, the purpose of our study was to investigate the effect of hyperglycemia on GAGs of the retinal endothelial glycocalyx. GAGs were isolated from rat retinal microvascular endothelial cells (RRMECs), media, and retinas, followed by liquid chromatography-mass spectrometry assays. Quantitative real-time polymerase chain reaction was used to study mRNA transcripts of the enzymes involved in GAG biosynthesis. Hyperglycemia significantly increased the shedding of heparan sulfate (HS), chondroitin sulfate (CS), and hyaluronic acid (HA). There were no changes to the levels of HS in RRMEC monolayers grown in high-glucose media, but the levels of CS and HA decreased dramatically. Similarly, while HA decreased in the retinas of diabetic rats, the total GAG and CS levels increased. Hyperglycemia in RRMECs caused a significant increase in the mRNA levels of the enzymes involved in GAG biosynthesis (including EXTL-1,2,3, EXT-1,2, ChSY-1,3, and HAS-2,3), with these increases potentially being compensatory responses to overall glycocalyx loss. Both RRMECs and retinas of diabetic rats exhibited glucose-induced alterations in the disaccharide compositions and sulfation of HS and CS, with the changes in sulfation including N,6-O-sulfation on HS and 4-O-sulfation on CS.

3D Imaging of the Liver Extracellular Matrix in a Mouse Model of Non-Alcoholic Steatohepatitis.

Non-alcoholic steatohepatitis (NASH) is the most common chronic liver disease in the United States, affecting more than 70 million Americans. NASH can progress to fibrosis and eventually to cirrhosis, a significant risk factor for hepatocellular carcinoma. The extracellular matrix (ECM) provides structural support and maintains liver homeostasis via matricellular signals. Liver fibrosis results from an imbalance in the dynamic ECM remodeling process and is characterized by excessive accumulation of structural elements and associated changes in glycosaminoglycans. The typical fibrosis pattern of NASH is called "chicken wire," which usually consists of zone 3 perisinusoidal/pericellular fibrosis, based on features observed by Masson's trichrome stain and Picrosirius Red stains. However, these traditional thin two-dimensional (2D) tissue slide-based imaging techniques cannot demonstrate the detailed three-dimensional (3D) ECM structural changes, limiting the understanding of the dynamic ECM remodeling in liver fibrosis. The current work optimized a fast and efficient protocol to image the native ECM structure in the liver via decellularization to address the above challenges. Mice were fed either with chow or fast-food diet for 14 weeks. Decellularization was performed after in situ portal vein perfusion, and the two-photon microscopy techniques were applied to image and analyze changes in the native ECM. The 3D images of the normal and NASH livers were reconstituted and analyzed. Performing in situ perfusion decellularization and analyzing the scaffold by two-photon microscopy provided a practical and reliable platform to visualize the dynamic ECM remodeling in the liver.

3D Imaging of the Liver Extracellular Matrix in a Mouse Model of Non-Alcoholic Steatohepatitis

Non-alcoholic steatohepatitis (NASH) is the most common chronic liver disease in the United States, affecting more than 70 million Americans. NASH can progress to fibrosis and eventually to cirrhosis, a significant risk factor for hepatocellular carcinoma. The extracellular matrix (ECM) provides structural support and maintains liver homeostasis via matricellular signals. Liver fibrosis results from an imbalance in the dynamic ECM remodeling process and is characterized by excessive accumulation of structural elements and associated changes in glycosaminoglycans. The typical fibrosis pattern of NASH is called "chicken wire," which usually consists of zone 3 perisinusoidal/pericellular fibrosis, based on features observed by Masson's trichrome stain and Picrosirius Red stains. However, these traditional thin two-dimensional (2D) tissue slide-based imaging techniques cannot demonstrate the detailed three-dimensional (3D) ECM structural changes, limiting the understanding of the dynamic ECM remodeling in liver fibrosis. The current work optimized a fast and efficient protocol to image the native ECM structure in the liver via decellularization to address the above challenges. Mice were fed either with chow or fast-food diet for 14 weeks. Decellularization was performed after in situ portal vein perfusion, and the two-photon microscopy techniques were applied to image and analyze changes in the native ECM. The 3D images of the normal and NASH livers were reconstituted and analyzed. Performing in situ perfusion decellularization and analyzing the scaffold by two-photon microscopy provided a practical and reliable platform to visualize the dynamic ECM remodeling in the liver.

Bio-3D printing iPSC-derived human chondrocytes for articular cartilage regeneration

Osteoarthritis is a leading cause of pain and joint immobility, the incidence of which is increasing worldwide. Currently, total joint replacement is the only treatment for end-stage disease. Scaffold-based tissue engineering is a promising alternative approach for joint repair but is subject to limitations such as poor cytocompatibility and degradation-associated toxicity. To overcome these limitations, a completely scaffold-free Kenzan method for bio-3D printing was used to fabricate cartilage constructs feasible for repairing large chondral defects. Human induced pluripotent stem cell (iPSC)-derived neural crest cells with high potential to undergo chondrogenesis through mesenchymal stem cell differentiation were used to fabricate the cartilage. Unified, self-sufficient, and functional cartilaginous constructs up to 6 cm2 in size were assembled by optimizing fabrication time during chondrogenic induction. Maturation for 3 weeks facilitated the self-organisation of the cells, which improved the construct’s mechanical strength (compressive and tensile properties) and induced changes in glycosaminoglycan and type II collagen expression, resulting in improved tissue function. The compressive modulus of the construct reached the native cartilage range of 0.88 MPa in the 5th week of maturation. This paper reports the fabrication of anatomically sized and shaped cartilage constructs, achieved by combining novel iPSCs and bio-3D printers using a Kenzan needle array technology, which may facilitate chondral resurfacing of articular cartilage defects.

Diabetic Foot: The Role of Fasciae, a Narrative Review.

Wound healing is an intricate, dynamic process, in which various elements such as hyperglycemia, neuropathy, blood supply, matrix turnover, wound contraction, and the microbiome all have a role in this "out of tune" diabetic complex symphony, particularly noticeable in the complications of diabetic foot. Recently it was demonstrated that the fasciae have a crucial role in proprioception, muscular force transmission, skin vascularization and tropism, and wound healing. Indeed, the fasciae are a dynamic multifaceted meshwork of connective tissue comprised of diverse cells settled down in the extracellular matrix and nervous fibers; each constituent plays a particular role in the fasciae adapting in various ways to the diverse stimuli. This review intends to deepen the discussion on the possible fascial role in diabetic wounds. In diabetes, the thickening of collagen, the fragmentation of elastic fibers, and the changes in glycosaminoglycans, in particular hyaluronan, leads to changes in the stiffness, gliding, and the distribution of force transmission in the fasciae, with cascading repercussions at the cellular and molecular levels, consequently feeding a vicious pathophysiological circle. A clear scientific perception of fascial role from microscopic and macroscopic points of view can facilitate the identification of appropriate treatment strategies for wounds in diabetes and create new perspectives of treatment.

High cholesterol-supplemented diet during gestation and lactation alters liver glycosaminoglycans and associated lipoprotein receptors and results in fat accumulation in adulthood

In utero insults to growing fetus impact its health in adulthood. Glycosaminoglycans (GAGs) are involved in lipoprotein metabolism in the liver and vary both quantitively and qualitatively on feeding adult rats a diet rich in cholesterol. However, no reports are available to show the modulation of GAGs when the offspring are subjected to a high cholesterol diet in gestation and lactation stages. Hypercholesterolemia in pregnant rats was induced by feeding an AIN-93 diet supplemented with 0.5% cholesterol. The pups born to mothers fed with high cholesterol diet showed a significant increase in cholesterol and triglycerides accumulation in the liver. Quantitative changes in sulfated glycosaminoglycans (sGAGs), in particular of heparan sulfate, were observed across the developmental stages. Other players involved in lipoprotein metabolism, namely low-density lipoprotein receptor-related protein 1, apolipoprotein E, and low-density lipoprotein receptor expression levels, also showed differential changes across developmental stages. Interestingly, when pups from hypercholesterolemic mothers were fed a normal diet after weaning until adulthood, a considerable amount of fat accumulation in the liver was observed, implicating fetal exposure to early high cholesterol exposure on long term health.

Pilot study quantifying muscle glycosaminoglycan using bi-exponential T1\u03c1 mapping in patients with muscle stiffness after stroke

Post stroke muscle stiffness is a common problem, which left untreated can lead to disabling muscle contractures. The purpose of this pilot study was to evaluate the feasibility of bi-exponential T1ρ mapping in patients with arm muscle stiffness after stroke and its ability to measure treatment related changes in muscle glycosaminoglycans (GAGs). Five patients with muscle stiffness after stroke and 5 healthy controls were recruited for imaging of the upper arm with 3D-T1ρ mapping. Patients were scanned before and after treatment with hyaluronidase injections, whereas the controls were scanned once. Wilcoxon Mann–Whitney tests compared patients vs. controls and patients pre-treatment vs. post-treatment. With bi-exponential modeling, the long component, T1ρl was significantly longer in the patients (biceps P = 0.01; triceps P = 0.004) compared to controls. There was also a significant difference in the signal fractions of the long and short components (biceps P = 0.03, triceps P = 0.04). The results suggest that muscle stiffness is characterized by increased muscle free water and GAG content. Post-treatment, the T1ρ parameters shifted toward control values. This pilot study demonstrates the application of bi-exponential T1ρ mapping as a marker for GAG content in muscle and as a potential treatment monitoring tool for patients with muscle stiffness after stroke.

Study of Urinary Glycosaminoglycans among Essential Hypertensive Patients

Introduction: Essential hypertension is a systemic disease which affects endothelial basement membrane. Changes in Glycosaminoglycans (GAG) distribution pattern on glomerular basement membrane has been noted in hypertension. Hence it seems likely that an increase excretion of GAG levels may be an indicator of reduction in renal function. The qualitative or quantitative determination of urinary GAGs may be of value and could constitute a non invasive marker to assess the renal damage in essential hypertension. Aim: Differential analysis of urinary GAGs in essential hypertensive patients. Materials and Methods: This analytical case-control observational study which was conducted from November 2014 to June 2016 in Department of Biochemistry, SDM College of Medical Sciences and Hospital, Dharwad, Karnataka, India. The study group included 50 male patients with age group of 30-60 years, clinically diagnosed with essential hypertension. Control group included 50 healthy male individuals with age group of 30-60 years, blood donors visiting to hospital blood bank. Random urine sample was collected. Urine creatinine was estimated by Jaffe’s method, urine GAGs by Dimethyl methylene blue (DMMB) dye method and urine microalbumin by particle enhanced Turbidimetric inhibition immunoassay method. Correlation was tested with Spearman’s correlation coefficient. Level of significance was set for p-value <0.05 with confidence interval of 95%. Results: In present study, urinary GAGs levels in essential hypertensive study group was 14.57±10.16 mg/dL and in control group was 10.09±6.04 mg/dL. Urinary GAGs levels in essential hypertensive group was significantly high (p-value=0.890) when compared with normotensive control group. But no statistical significant correlation was found between urine GAGs and urine microalbumin in hypertensive study group. Conclusion: Estimation of urine glycosaminoglycans in essential hypertensive patients is a simple, rapid and cost effective test which asseses the glomerular function. It can be used as one of the early marker for diagnosis of nephropathy before microalbuminuria sets in.

Age-related functional changes of total thyroid hormones and glycosaminoglycans in growing calves.

Background and Aim:During the physiological growing, thyroid and proteoglycan glycosaminoglycan (GAG) changes dynamically occur, according to genetic and non-genetic factors. The purpose of this research was to compare the effects of early postnatal development (10 days) until 210 days of life on the triiodothyronine (T3), thyroxine (T4), the relative T4:T3 ratio, and GAGs profile, and to define the different reference intervals of the calf’s development through the various growing phases.Materials and Methods:The effect of growing on total thyroid hormones and GAG profiles was studied from 10 days to 210days of age in 64 clinically healthy Brown calves, 30males and 34females. Blood samples were collected at 10, 20, 30, 60, 90, 120, 150, 180, and 210days of age.Results:The results showed a significant effect of a calf’s growth on T3, T4, and GAG values (p<0.0001). Significant correlations between T3 and T4 were observed. Compared to the previous time point, T3 showed a significant decrease at 20days and at 60days (p<0.01), while a significant increase was observed at 90days and 210days (p<0.05); T4 showed a significant decrease at 20days (p<0.01), while significant increases were observed at both 180days and 210days (p<0.05); GAGs showed a significant increase at 120days and 210days (p<0.05). Positive and significant correlations between BW and GAGs in both males (p<0.0057) and females (p<0.0059) were observed.Conclusion:It can be concluded that the highest T3 and T4 concentrations have been associated with the early growing process (10days), with an increasing trend also at 210days, it is possible to hypothesize a probable metabolic effect of thyroid function in anabolic and/or catabolic directions during the calves’ development. Likewise, it can be reasonably inferred that the highest plasma GAGs at 210days may be due to their metabolic role during the development of growing calves. Taken together, these findings suggest the potential and relative contribution made by thyroid and GAGs effects on the dynamics of growing calves.

Safety Study of Sodium Pentosan Polysulfate for Adult Patients with Mucopolysaccharidosis Type II.

Current therapies for the mucopolysaccharidoses (MPS) do not effectively address skeletal and neurological manifestations. Pentosan polysulfate (PPS) is an alternative treatment strategy that has been shown to improve bone architecture, mobility, and neuroinflammation in MPS animals. The aims of this study were to a) primarily establish the safety of weekly PPS injections in attenuated MPS II, b) assess the efficacy of treatment on MPS pathology, and c) define appropriate clinical endpoints and biomarkers for future clinical trials. Subcutaneous injections were administered to three male Japanese patients for 12 weeks. Enzyme replacement therapy was continued in two of the patients while they received PPS and halted for two months in one patient before starting PPS. During treatment, one patient experienced an elevation of alanine transaminase, and another patient experienced convulsions; however, these incidences were non-cumulative and unrelated to PPS administration, respectively. Overall, the drug was well-tolerated in all patients, and no serious drug-related adverse events were noted. Generally, PPS treatment led to an increase in several parameters of shoulder range of motion and decrease of the inflammatory cytokines, MIF and TNF-α, which are potential clinical endpoints and biomarkers, respectively. Changes in urine and serum glycosaminoglycans were inconclusive. Overall, this study demonstrates the safety of using PPS in adults with MPS II and suggests the efficacy of PPS on MPS pathology with the identification of potential clinical endpoints and biomarkers.

Compressive fatigue and endurance of juvenile bovine articular cartilage explants

Articular cartilage is an enduring tissue. For most individuals, articular cartilage facilitates a lifetime of pain-free ambulation, supporting millions of loading cycles from activities of daily living. Although early studies into osteoarthritis focused on the role of mechanical fatigue in articular cartilage degeneration, much is still unknown regarding its strength and endurance characteristics. The compressive strength of juvenile, bovine articular cartilage explants was determined to be loading rate-dependent, reaching a maximum strength of 29.5 ± 4.8 MPa at a strain rate of 0.10 %/sec. The fatigue and endurance properties of articular cartilage were characterized utilizing a material testing system, as well as a custom, validated instrument termed the two degrees-of-freedom endurance meter (endurometer). These instruments characterized fatigue in articular cartilage explants at loading levels ranging from 10 to 80 % strength (%S), up to 100,000 cycles. Cartilage explants displayed characteristics of fatigue – fatigue life increased as the loading magnitude decreased. All explants failed within 14,000 cycles at loading levels between 50 and 80 %S. At 10 and 20 %S, all explants endured 100,000 loading cycles. There was no significant difference in equilibrium compressive modulus between run-out explants and unloaded controls, although the pooled modulus increased in response to testing. Histological staining and biochemical assays revealed no material changes in collagen, sulfated glycosaminoglycan, or hydration content between unloaded controls and explants cyclically loaded to run-out. These results suggest articular cartilage may have a putative endurance limit of 20 %S (5.86 MPa), with implications for articular cartilage biomechanics and mechanobiology.

Non-Invasive Monitoring of Stromal Biophysics with Targeted Depletion of Hyaluronan in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDA) is characterized by a pronounced fibroinflammatory stromal reaction consisting of inordinate levels of hyaluronan (HA), collagen, immune cells, and activated fibroblasts that work in concert to generate a robust physical barrier to the perfusion and diffusion of small molecule therapeutics. The targeted depletion of hyaluronan with a PEGylated recombinant human hyaluronidase (PEGPH20) lowers interstitial gel–fluid pressures and re-expands collapsed intratumoral vasculature, improving the delivery of concurrently administered agents. Here we report a non-invasive means of assessing biophysical responses to stromal intervention with quantitative multiparametric magnetic resonance imaging (MRI) at 14 Tesla (T). We found that spin-spin relaxation time T2 values and glycosaminoglycan chemical exchange saturation transfer (GagCEST) values decreased at 24 h, reflecting depletion of intratumoral HA content, and that these parameters recovered at 7 days concurrent with replenishment of intratumoral HA. This was also reflected in an increase in low-b apparent diffusion coefficient (ADC) at 24 h, consistent with improved tumor perfusion that again normalized at 7 days after treatment. Phantom imaging suggests that the GagCEST signal is driven by changes in HA versus other glycosaminoglycans. Thus, multiparametric magnetic resonance imaging (MRI) can be used as a non-invasive tool to assess therapeutic responses to targeted stromal therapy in PDA and likely other stroma-rich solid tumors that have high levels of hyaluronan and collagen.

THE RISE AND FALL OF DGEMRIC EVALUATION IN THE ADOLESCENT HIP

BACKGROUND: Delayed Gadolinium Enhanced Magnetic Resonance Imaging of Cartilage (dGEMRIC) was first introduced in 1996 by Bashir et al. as a method of indirectly evaluating glycosaminoglycan (GAG) changes in cartilage microstructure. Following publications originating from the Boston Children’s research group describing the association of early joint failure following periacetabular osteotomy (PAO) in patients with diminished preoperative cartilage T1 relaxation values, the interest in this imaging technique spread. dGEMRIC imaging was introduced into our clinical practice in 2013 with enrollment of both pediatric and adult patients. However, since the program’s apex in 2015, a marked decline in referral of adolescent patients for preoperative dGEMRIC imaging has occurred. Using data acquired during a quality analysis of the dGEMRIC program, this project was designed to investigate the utility of the dGEMRIC exam in our adolescent imaging cohort from 2013-2015 and to identify potential factors leading to decreased utilization from 2015-2018. METHODS: IRB approval was obtained for a HIPAA-compliant, retrospective cohort study which included all patients referred for dGEMRIC from May 2013- June 2015. 108 patients/112 exams/122 hips were identified. The study was designed in 4 parts including a. Blinded quantitative imaging review x 2 performed by 4 radiologists, b. Surgical review with 2 year post-operative follow-up completed July 2017, c. Patient phone interview, and d. Financial billing and reimbursement analysis. Subsequently, dGEMRIC utilization data from July 2015-May 2018 was compiled for comparative analysis of adolescent referral patterns. RESULTS: Of the 108 patients included in the comprehensive retrospective analysis, 56 were &lt; 19 years of age with 43% of the adolescent patients participating in sports at time of presentation. 4 patients were excluded as the dGEMRIC data was either incomplete or could not be reprocessed. 61 hips were evaluated in the remaining 52 patients. The following summary data was compiled during the 4 part analysis. From the radiologist review: Siemens MapIt® T1 measurement software performed well at both 1.5 T and 3 T with good to excellent within rater ICC (0.79-0.92) but the between rater agreement was lacking particularly at 3.0 T (0.45) compared to 1.5 T (0.77) (p&lt;0.05). Variability between sequential T1 measurements performed by the same radiologist at 2 different time points existed averaging 36-41 ms at 1.5 T and 76-84 ms at 3 T. From the surgical perspective: 17 hips in 16 patients were determined to have abnormally low T1 relaxation values when reprocessed by the 4 radiologists. 1 patient’s T1 values differed significantly after blinded review compared to the original interpretation. Of the 16 abnormal patients, 5 were referred for total hip arthroplasty. 7 patients underwent surgical intervention (2 PAO, 3 arthroscopic debridement/rim trim/chondroplasty/labral repair, 2 femoral/acetabular osteotomy). 1 patient was referred to rheumatology and 3 were lost to follow-up. After 2 year follow-up, all surgical patients reported decreased pain and improved function compared to presentation. From the patient perspective: 62 of the original 108 patients were contacted by phone with 60 (56%) participating in the phone survey. Patients described the dGEMRIC exam as a difficult procedure. The total time expenditure for the dGEMRIC exam averaged 3.5 hours in hospital with an average time in magnet of 107 minutes. 40% of patients reported significant pain with the pre-imaging exercise and 33% reported significant pain while in the MRI. Of 49 respondents who had undergone previous hip injections, 28 (57%) stated the dGEMRIC exam was as or more difficult than the injection. However, 50/57 (88%) stated they would undergo dGEMRIC again if the surgeon felt the information would be of added value in decision making. From the financial review, a lack of standardization of CPT codes billed was discovered resulting in discrepant billed charges varying by a factor &gt;10 ($2300-$24,000). Following completion of the quality analysis and review of the results with the referring orthopedists, only 17 dGEMRIC exams have been performed in the adolescent age group in the subsequent 36 months. CONCLUSIONS: While the dGEMRIC exam holds promise as a prognosticating tool to guide surgical planning, in the adolescent practice environment, our historical data reveals that even when faced with abnormal T1 relaxation values, surgeons may opt to proceed with hip sparing surgical procedures simply because of the young age of the patient. From the radiologist’s perspective, dGEMRIC reporting is clouded by measurement variability. Inherent variability in the numbers reported brings to question the wisdom of using absolute T1 numeric thresholds to prognosticate joint failure risk. From the patient’s perspective, this imaging test comes with significant cost measured by time, pain, and financial burden. For all these reasons, utilization of this imaging study should be limited to the select few adolescent patients in whom documentation of an abnormal cartilage T1 value alone could substantially impact the surgical plan.

Effects of electrospun scaffolds of di-O-butyrylchitin and poly-(ε-caprolactone) on wound healing.

We sought to determine the usefulness of electrospun dibutyrylchitin (DBC) or poly-(ε-caprolactone [PCL]), in wound treatment. We investigated the mechanisms of action of these polymers on wound healing. We synthesized DBC, a newly identified ester derivative of chitin, using a patented method comprising the substitution of butyryl groups at positions C-3 and C-6 in chitin molecules. We confirmed the double substitution by the butyric groups using infrared spectrometry. The fibrous scaffolds were obtained using the electrospinning method. A polypropylene net was implanted subcutaneously in the rat and served as a wound model. Both DBC and PCL increased granulation tissue weight in the wound. In contrast to PCL, DBC did not abolish glycosaminoglycan changes in wounds. The tested samples did not impair total collagen synthesis or induce excessive fibrosis. In both PCL- and DBC-treated wounds, we observed a lower level of soluble collagen (compared with controls). The results show better hydration of the wounds in both the DBC and PCL groups. No induction of large edema formation by the tested materials was observed. These polymers induced almost identical macrophage-mediated reactions to foreign-body implantation. The implants increased the blood vessel number in a wound. Both PCL and DBC could be used as scaffolds or dressings for wound treatment. The materials were safe and well tolerated by animals. As DBC did not disturb glycosaminoglycan accumulation in wounds and absorbed twice as much liquid as PCL, it can be considered superior.

Changes in glycosaminoglycan characteristics during progression of a human gingival carcinoma xenograft line in nude mice.

We investigated changes in the glycosaminoglycans (GAGs) during progression of a human gingival carcinoma xenograft line, GK -1, in nude mice. The GAGs extracted from cancers 3, 5, 7, 10 and 15 weeks after transplantation consisted of hyaluronic acid (HA), chondroitin sulfate (CS) and heparan sulfate (HS) as major components, and dermatan sulfate (DS) as a trace component for all cancers. HPLC analysis revealed that the HA content per defatted tissue dry weight increased in the cancers 5 weeks after transplantation compared to those of 3 weeks (p < 0.05), while CS for cancers at 10 weeks decreased compared with 7 weeks (p < 0.05). However, HS showed no significant change. Both the CS and DS contained primarily 4-sulfated disaccharide units. Immunohistochemical staining with antibody 2-B-6 for the PGs having delta DI-4S produced by chondroitinase ABC digestion showed that CS is located in the tissue surrounding the cancer nests and mass. These results indicate that the location of accumulation of CS, which primarily contains 4-sulfated disaccharide units, plays an important role in cancer progression.