Significant Differences In Collagen Research Articles

Comparison of Biological Augmentation in Rotator Cuff Repair Using Inflamed Versus Noninflamed Bursal Tissue in Rats

PurposeTo examine how augmentation of a rotator cuff repair with inflamed versus non-inflamed bursal tissue affects tendon-to-bone healing in a rat model of rotator cuff repair. Methods136 Sprague-Dawley rats were randomly assigned to an inflamed or non-inflamed bursal tissue application group. After detachment the supraspinatus tendon was re-attached with bursal tissue sewn onto the tendon-to-bone interface. The specimens were analysed biomechanically 6 and at 7 weeks and immunohistologically at 1 and at 7 weeks after surgery. ResultsImmunohistological results showed no significant difference in the percentage of collagen type II in the tendon-to-bone interface at 1 (p = 0.97) and 7 weeks (p = 0.42) when utilising autologous non-inflamed bursal tissue in comparison to inflamed bursal tissue specimens. The inflamed bursa group also showed no significant difference in collagen I to III quotient (p= 0.14) after surgery in comparison to post-surgery non-inflamed bursa groups. Biomechanical assessment showed that tendon stiffness (p = 0.87 resp. p = 0.1) and the tendon viscoelasticity (p = 0.12 resp. p = 0.07) was the same after 6 and 7 weeks comparing inflamed bursa to the non-inflamed bursa group. There was no significant difference (p = 0.8 resp. p = 0.97) in load to failure between in both inflamed and non-inflamed bursa groups after 6 and 7 weeks. ConclusionAutologous inflamed bursal tissue derived from the Achilles bursa and implanted to the tendon-to-bone interface after rotator cuff repair facilitates the same histological and biomechanical healing response as using a non-inflamed bursa interposition in rats. Clinical relevanceDuring augmentation of a rotator cuff repair, it is irrelevant whether the bursa tissue is inflamed or not.

Daunorubicin Treatment Impairs Cardiac Function in The Ts65Dn Mouse Model Of Down Syndrome

Anthracycline treatment for leukemia is effective but has toxic side effects on the cardiovascular system. Few studies regarding the impact of anthracyclines on the heart in individuals with Down syndrome have been addressed although they are at a higher risk of developing leukemia. It is unknown if established mechanisms apply to individuals with Down syndrome. We hypothesize that anthracycline cardiotoxicity will lead to the enhanced impairment of cardiac function and initiate pronounced vascular dysfunction in a mouse model of Down syndrome. We used the Ts65Dn mouse model of Down syndrome to investigate the impact of daunorubicin chemotherapy on survival, arterial stiffness and left ventricular function. Four-month-old Ts65Dn (n=12) and wild-type control Ts65Dn (WT, n=8) mice were treated with 2mg/kg of daunorubicin or saline. Following treatment, left ventricular catheterization was performed to assess cardiac function. Serum cardiac troponin levels were evaluated using ELISA. Histological analysis was conducted on the heart and aorta of mice to assess the accumulation of elastin and collagen. We found decreased end-diastolic pressure (p=0.016) and ejection fraction (p=0.029) in Ts65Dn daunorubicin-treated mice compared to WT control. However, contractility (end-systolic pressure-volume relationship, p=0.221) and diastolic stiffness coeffcient (end-diastolic pressure-volume relationship, p=0.418) remained unchanged. Ts65Dn mice had a higher cardiac troponin concentration (p=0.02). Histological assessment showed no significant difference in collagen (p=0.841) and elastin (p=0.768) content. Daunorubicin negatively impacted end-diastolic pressure and ejection fraction of the Ts65Dn mouse model of Down syndrome. We demonstrated that we could recapitulate the findings of anthracycline cardiotoxicity in Ts65Dn mice indicating this is a good model to investigate toxicity. Continued work with this model will better elucidate the mechanisms underlying anthracycline cardiotoxicity in Down syndrome. This shows the need for constant screening after anthracycline treatment which will improve the survival of this population. NIH R21 HD099573. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Paper 08: In Vitro Effects of Triamcinolone and Methylprednisolone on the Viability and Mechanics of Native Articular Cartilage

Objectives: The chondrotoxic effects of methylprednisolone acetate (MP) and triamcinolone acetonide (TA) have been well described. Prior studies have demonstrated a dose-dependent chondrotoxic effect of intra articular (IA) steroids starting at 7 mg for MP and 18 mg for TA. However, the effects of these steroids on the mechanical properties of articular cartilage are largely unknown. The purpose of this study was to investigate the in vitro effects of a single one-hour MP or TA exposure on the viability, mechanics, and biochemical content of native articular cartilage explants. Methods: Articular cartilage explants (n=6 per group) were harvested from the femoral condyle of the bovine stifle. Explants were exposed to chondrogenic medium containing a clinical dose of MP or TA for one hour, followed by fresh medium wash and exchange. Explants in the negative control group underwent the same treatment with chondrogenic medium alone. At t=24 hours post-treatment, samples were assessed for viability (live/dead), mechanical properties (creep indentation and Instron tensile testing), biochemical (collagen and glycosaminoglycan) content, and pyridinoline cross linking via mass spectrometry. Results: Mean cell viability was significantly decreased in native explants exposed to MP (35.5%) compared to control (49.8%, p < 0.001) and TA (45.7%, p = 0.003) (Figure 1). There was significant weakening of mechanical properties of steroid-treated native explants when compared to control (Figure 2), with decreases in aggregate modulus (646.3 kPa vs 312.8 kPa [MP] and 257.0 kPa [TA), p < 0.001), shear modulus (370.1 kPa vs 191.2 kPa [MP] and 157.4 kPa [TA], p < 0.001), and ultimate tensile strength (UTS) (9.650 MPa vs 5.648 MPa [MP], p = 0.021 and 6.065 MPa [TA], p = 0.040). The Young’s modulus in TA-exposed native cartilage (7.924 MPa) was significantly lower than MP (12.35 MPa, p = 0.026) exposed explants but not significantly different compared to control (11.97 MPa, p =0.072). No significant differences in collagen and glycosaminoglycan content were found in the steroid-treated groups (Figure 3). Pyridinoline cross linking was significantly decreased in explants exposed to TA compared to control (p = 0.027). Conclusions: The results of this in vitro study suggest that a single clinical dose of MP is chondrotoxic and that a single clinical dose of MP or TA can lead to substantial early decreases in both the tensile and compressive properties of native articular cartilage. The shear modulus of steroid-exposed explants in this study dropped 48-57% compared to healthy control. In human knee cartilage, drops in shear modulus of this magnitude have been correlated with the progression from healthy knee cartilage to ICRS grade 1-2 osteoarthritis. Furthermore, decreases in Young’s modulus by 25-50% have been correlated with the same degree of osteoarthritis, and the Young’s modulus of steroid exposed cartilage dropped an average of 34% in this study. Although there were no detectable changes to the extracellular matrix composition or structure, a single one-hour steroid exposure was enough to cause chondrotoxicity and altered mechanics to native explants 24 hours after exposure, which raises the concern for risk of mechanical failure of the cartilage tissue. Clinicians should be judicious regarding use of intra-articular steroids, particularly in patients with intact healthy articular cartilage.

A Soy Protein Isolate Rich in Genistein and Daidzein and Its Effects on Plasma Isoflavone Concentrations, Platelet Aggregation, Blood Lipids and Fatty Acid Composition of Plasma Phospholipid in Normal Men

The effects of consuming a soy protein isolate beverage powder (60 g/d for 28 d) vs. a casein supplement was evaluated in 20 male subjects who were randomly allocated into the two groups. A dramatic rise in plasma isoflavone concentrations was observed after supplementation in the soy protein group, the levels reaching 907 ± 245 nmol/L for genistein (a 110-fold increase) and 498 ± 102 nmol/L for daidzein (a 150-fold increase) as measured by isotope dilution gas chromatography - mass spectrometry. These concentrations are higher than previously reported for the plasma of Japanese subjects consuming a traditional diet (276 nmol/L and 107 nmol/L, respectively). No significant differences in collagen- or 9,11-dideoxy-11α, 9α-epoxymethanoprostaglandin F2α (U46619)-induced platelet aggregation were observed in platelet-rich plasma from the two groups; the increase in plasma isoflavonoids from soy protein supplementation is not sufficient to significantly inhibit platelet aggregation ex vivo. Similarly, plasma total and HDL-cholesterol were not affected by protein supplementation, possibly because the men were normo-cholesterolemic at entry. Analysis of plasma phospholipid polyunsaturated fatty acid composition showed no differences between soy protein and casein supplementation. Previous investigations reported a significant alteration in fatty acid status in animals fed soy protein relative to those fed casein. The present studies indicate that although soy protein supplementation to a typical Western diet can increase plasma concentrations of isoflavones, this may not necessarily be sufficient to counter heart disease risk factors such as high plasma cholesterol and platelet aggregation.

Label-free Raman spectroscopy characterizes signatures of inflammation and fibrosis in the silicosis

Silicosis is an occupational disease that seriously damages the life and health of miners. Herein, we constructed a mouse model of silicosis and used label-free confocal Raman spectroscopy to analyze the biomolecular variations in lung fibrous nodules and inflammatory sites. The mice were exposed to silica particles for 1 month (SIL-1M group), 3 months (SIL-3M group), or no exposure (control tissues, NS). Raman spectra obtained from treated and untreated lung tissue were subjected to chemometric analysis to quantify biochemical composition differences in the silicosis. Simultaneously, immunohistochemistry and collagen staining were used to evaluate inflammation, fibrosis, and apoptosis. As a result, the SIL-1M and SIL-3M groups showed significant differences in cholesterol, lipids, amino acids, nucleic acids, and cytochrome C, and the collagen peaks at 1248 cm−1 and 1448 cm−1 were significantly higher than in the NS group. Notably, glycogen and phospholipid may be an inflammatory indicator consistent with NF-κB expression. In addition, significant differences in collagen and cytochrome C content in silicosis lung tissue were found using Raman spectroscopy and were verified by Masson's staining and Bax/Bcl-2 expression ratio. In summary, our findings provide a label-free technique to understand the biochemical changes in lung inflammatory and fibrosis microenvironment after exposure to silica particles and provide a valuable reference for studying the mechanism of silicosis.

Effect of biomolecules derived from human platelet-rich plasma on the ex vivo expansion of human adipose-derived mesenchymal stem cells for clinical applications

Mesenchymal stem cells are a tool in cell therapies but demand a large cell number per treatment, for that, suitable culture media is required which contains fetal bovine serum (FBS). However, for cell-based therapy applications, the use of FBS is problematic. Several alternatives to FBS have been explored, including human derivatives from platelet-rich plasma (hD-PRP). Although various studies have evaluated the impact of hD-PRP on MSC proliferation and differentiation, few of them have assessed their influence on processes, such as metabolism and gene expression. Here, we cultured human adipose-derived MSCs (hAD-MSCs) in media supplemented with either 10% hD-PRP (hD-PRP-SM) or 10% FBS (FBS-SM) in order to characterize them and evaluate the effect of hD-PRP on cell metabolism, gene expression of associated regenerative factors, as well as chromosome stability during cell expansion. We found that hAD-MSCs cultured in hD-PRP-SM have a greater cell elongation but express similar surface markers; in addition, hD-PRP-SM promoted a significant osteogenic differentiation in the absence of differentiation medium and increased the growth rate, maintaining chromosomal stability. In terms of cell metabolic profile, hAD-MSC behavior did not reveal any differences between both culture conditions. Conversely, significant differences in collagen I and angiopoietin 2 expression were observed between both conditions. The present results suggest that hD-PRP may influence hAD-MSC behavior.

A matrisome RNA signature from early-pregnancy mouse mammary fibroblasts predicts distant metastasis-free breast cancer survival in humans

BackgroundDuring pregnancy, the mouse mammary ductal epithelium branches and grows into the surrounding stroma, requiring extensive extracellular matrix (ECM) and tissue remodelling. It therefore shows parallels to cancer invasion. We hypothesised that similar molecular mechanisms may be utilised in both processes, and that assessment of the stromal changes during pregnancy-associated branching may depict the stromal involvement during human breast cancer progression.MethodsImmunohistochemistry (IHC) was employed to assess the alterations within the mouse mammary gland extracellular matrix during early pregnancy when lateral branching of the primary ductal epithelium is initiated. Primary mouse mammary fibroblasts from three-day pregnant and age-matched non-pregnant control mice, respectively, were 3D co-cultured with mammary epithelial cells to assess differences in their abilities to induce branching morphogenesis in vitro. Transcriptome analysis was performed to identify the underlying molecular changes. A signature of the human orthologues of the differentially expressed matrisome RNAs was analysed by Kaplan–Meier and multi-variate analysis in two large breast cancer RNA datasets (Gene expression-based Outcome for Breast cancer Online (GOBO) und Kaplan–Meier Plotter), respectively, to test for similarities in expression between early-pregnancy mouse mammary gland development and breast cancer progression.ResultsThe ECM surrounding the primary ductal network showed significant differences in collagen and basement membrane protein distribution early during pregnancy. Pregnancy-associated fibroblasts (PAFs) significantly enhanced branching initiation compared to age-matched control fibroblast. A combined signature of 64 differentially expressed RNAs, encoding matrisome proteins, was a strong prognostic indicator of distant metastasis-free survival (DMFS) independent of other clinical parameters. The prognostic power could be significantly strengthened by using only a subset of 18 RNAs (LogRank P ≤ 1.00e−13; Hazard ratio (HR) = 2.42 (1.8–3.26); p = 5.61e−09). The prognostic power was confirmed in a second breast cancer dataset, as well as in datasets from ovarian and lung cancer patients.ConclusionsOur results describe for the first time the early stromal changes that accompany pregnancy-associated branching morphogenesis in mice, specify the early pregnancy-associated molecular alterations in mouse mammary fibroblasts, and identify a matrisome signature as a strong prognostic indicator of human breast cancer progression, with particular strength in oestrogen receptor (ER)-negative breast cancers.

Bone Marrow Mesenchymal Stem Cell-Derived Tissues are Mechanically Superior to Meniscus Cells.

Bone marrow-derived mesenchymal stem cells (BMSCs) have the potential to form the mechanically responsive matrices of joint tissues, including the menisci of the knee joint. The purpose of this study is to assess BMSC's potential to engineer meniscus-like tissue relative to meniscus fibrochondrocytes (MFCs). MFCs were isolated from castoffs of partial meniscectomy from nonosteoarthritic knees. BMSCs were developed from bone marrow aspirates of the iliac crest. All cells were of human origin. Cells were cultured in type I collagen scaffolds under normoxia (21% O2) for 2 weeks followed by hypoxia (3% O2) for 3 weeks. The structural and functional assessment of the generated meniscus constructs were based on glycosaminoglycan (GAG) content, histological appearance, gene expression, and mechanical properties. The tissues formed by both cell types were histologically positive for Safranin O stain and appeared more intense in the BMSC constructs. This observation was confirmed by a 2.7-fold higher GAG content. However, there was no significant difference in collagen I (COL1A2) expression in BMSC- and MFC-based constructs (p = 0.17). The expression of collagen II (COL2A1) and aggrecan (ACAN) were significantly higher in BMSCs than MFC (p ≤ 0.05). Also, the gene expression of the hypertrophic marker collagen X (COL10A1) was 199-fold higher in BMSCs than MFC (p < 0.001). Moreover, relaxation moduli were significantly higher in BMSC-based constructs at 10-20% strain step than MFC-based constructs. BMSC-based constructs expressed higher COL2A1, ACAN, COL10A1, contained higher GAG content, and exhibited higher relaxation moduli at 10-20% strain than MFC-based construct. Impact statement Cell-based tissue engineering (TE) has the potential to produce functional tissue replacements for irreparably damaged knee meniscus. But the source of cells for the fabrication of the tissue replacements is currently unknown and of research interest in orthopedic TE. In this study, we fabricated tissue-engineered constructs using type I collagen scaffolds and two candidate cell sources in meniscus TE. We compared the mechanical properties of the tissues formed from human meniscus fibrochondrocytes and bone marrow-derived mesenchymal stem cells (BMSCs). Our data show that the tissues engineered from the BMSC are mechanically superior in relaxation modulus.

905 Sex differences in the molecular cause of perioral skin wrinkling

Wrinkling is the hallmark of skin aging. We have previously reported that perioral wrinkling is more severe in females; however the molecular basis for this is unknown. We enrolled 12 subjects (n=6 male/female) age 54-86 with Griffith’s photoaging severity grade between 4 and 8 (0=none, 8=severe) and took biopsies from both the perioral and periocular region. Using qPCR, we assessed RNA expression levels of collagen I, collagen III, cysteine-rich angiogenic inducer 61 (CYR61), and insulin like growth factor 1 (IGF-1). CYR61 causes dermal fibroblast senescence in response to photodamage, leading to the decreased collagen production linked to skin wrinkling. IGF-1 is produced by dermal fibroblasts and plays a role in regulating the hair cycle and promoting oxidative stress. While there was no difference between females’ and males’ Griffith’s grade (6.67 and 5.67 respectively, p=0.096) or periocular wrinkling grade (3.2 and 2.6 respectively, p=0.421), females did have a significantly more severe perioral wrinkling grade when compared with males (6.2 and 2.8 respectively, p=.0.035). Consistent with the increase in severity in perioral wrinkling, females also expressed significantly more CYR61 (p=0.018) than males, however, they also expressed more collagen III (p=0.016). In this location, there was no significant difference in collagen I (p=0.115) or IGF-1 (p=0.124) expression between males and females, although females trended towards higher expression of both, with the latter marker’s role in oxidative stress potentially contributing to the increased perioral wrinkling in females. In the periocular region, there were no significant differences between males and females in the expression levels of all four markers. While both sexes had similar overall photoaging and gene expression in the periocular region, the perioral region expressed significant molecular differences between the sexes, which may contribute to the greater perioral skin wrinkling seen clinically in females. It also highlights the complex process of skin aging and the influence of specific anatomic milieu in this process.

Phenogenomics of the gingival fibroblast interaction with implant materials- a standardized system to determine material/surface potential for connective tissue attachment

Event Abstract Back to Event Phenogenomics of the gingival fibroblast interaction with implant materials- a standardized system to determine material/surface potential for connective tissue attachment Gethin Owen1, Ping Huang1, Sadaf G. Kashani1, Lari Hakkinen1 and Hannu Larjava1 1 University of British Columbia, Oral, Biol & Med Sci, Canada The health of the soft-tissue surrounding a dental implant is an integral part of successful dental implant treatment. Human and animal models have shown that the peri-implant connective tissue contains relatively low numbers of fibroblasts, little or no blood vessels, similar to scar-like tissue. Such a tissue forms a poorly functional attachment to the implant surface unlike the gingival attachment to teeth. We report on an in vitro test at the phenotype and genotype level to quickly assess the scar forming potential of implant materials/surfaces. To test our methodology 3 titanium surfaces were used, namely pickled (PT) (Sa =0.3 µm), sandblasted (S) and sand blasted acid etched (SLA) (both Sa =1.2 µm). The surfaces were characterized by scanning electron microscopy (SEM). The initial gingival fibroblast (GF) response was quantified comparing cell adhesion, cell proliferation, immunolabelling and area of cell-implant contact with a focused ion beam/scanning electron microscope (FIB/SEM). Extracellular matrix protein (ECM) synthesis, angiogenesis potential and gene expression on each material surface were also quantified at 1 and 7 days. GFs cultured on all surfaces had distinct morphology after 1 day. On PT they were well spread. On S some cells were well spread and others had a stellate shape. All cells on SLA were stellate. Mature focal adhesions (FA) were visible on PT and S. Stellate shaped GFs cultured on S and SLA had small FA on the edges of surface features. Cross-sections of cell/implant with FIB/SEM confirmed this finding. FA area of adhesion was greatest on SLA which resulted in stronger cell adhesion as assessed by a cell detachment assay. After 1 day cell proliferation on SLA was significantly lower than PT and S but there was no difference after 7 days. Significant differences in expression of wound healing genes were seen relative to tissue culture plastic. Increased roughness caused a decrease in collagen I, III and decorin genes whereas matrix metalloproteinase (MMP)-1 increased after 1d. MMP-3 expression increased only in SLA after 7d. VEGF was highly expressed in SLA but the tube formation assay with HUVEC cells showed no angiogenic effect. ECM synthesis quantification showed significant differences in collagen I, decorin and fibronectin-EDA production. Production was highest on PT and the ECM had orientated with the topography in PT and S but not with SLA. This phenogenomic test suggests that surface topography is effective in modifying wound healing behavior at the GF phenotype, proteome and gene level. The investigation was funded by ITI Research Grant No.964-2013 Keywords: Implant, Cell response, tissue compatibility, surface topolography Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: General Session Oral Topic: Biomaterials in dental applications Citation: Owen G, Huang P, Kashani SG, Hakkinen L and Larjava H (2016). Phenogenomics of the gingival fibroblast interaction with implant materials- a standardized system to determine material/surface potential for connective tissue attachment. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.02438 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2016; Published Online: 30 Mar 2016. Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Gethin Owen Ping Huang Sadaf G Kashani Lari Hakkinen Hannu Larjava Google Gethin Owen Ping Huang Sadaf G Kashani Lari Hakkinen Hannu Larjava Google Scholar Gethin Owen Ping Huang Sadaf G Kashani Lari Hakkinen Hannu Larjava PubMed Gethin Owen Ping Huang Sadaf G Kashani Lari Hakkinen Hannu Larjava Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Extracellular Matrix of Collagen Modulates Intracellular Calcium Handling and Electrophysiological Characteristics of HL-1 Cardiomyocytes With Activation of Angiotensin II Type 1 Receptor

Background Myocardial fibrosis plays a critical role in heart failure, resulting in cardiac structural and electrical remodeling which can induce atrial arrhythmias. Collagen is the major element of fibrosis. However, it is not clear whether collagen can directly regulate the calcium homeostasis and the electrophysiologic characteristics of cardiomyocytes. The aim of this study was to determine the effects of collagen on calcium homeostasis and the electrical properties of atrial cardiomyocytes. Methods and Results HL-1 cardiomyocytes were cultured with and without collagen type I (1 or 10 μg/mL) or losartan (10 μmol/L). Whole-cell clamp, indo-1 fluorescence, and Western blotting were used to evaluate the action potential (AP) and ionic currents, intracellular calcium homeostasis, and calcium regulatory proteins. Compared with the control samples, there was no significant difference in collagen (1 μg/mL)–treated HL-1 cardiomyocytes. However, collagen (10 μg/mL)–treated HL-1 cardiomyocytes exhibited larger intracellular calcium ([Ca 2+] i) transients by 113% and a larger sarcoplasmic reticulum calcium content by 86%. Collagen (10 μg/mL)–treated HL-1 cardiomyocytes had higher expression of sarcoplasmic reticulum ATPase (SERCA2a) and Thr17-phosphorylated phospholamban but similar protein expressions of the Na +/Ca 2+ exchanger and ryanodine receptor. Collagen (10 μg/mL)–treated HL-1 cardiomyocytes (n = 11) had larger AP amplitude (104 ± 5 vs 83 ± 7 mV; P < .05), and shorter 90% of AP duration (25 ± 2 vs 33 ± 2 ms, P < .05) than control cells (n = 11). Moreover, collagen (10 μg/mL)–treated HL-1 cells had larger I to and I Ksus values than control cells. The administration of losartan (10 μmol/L) attenuated collagen-induced changes in [Ca 2+] i transients, [Ca 2+] i stores, AP morphology, ionic currents, SERCA2a, and Thr17-phosphorylated phospholamban expressions. Conclusions This study demonstrates that collagen can directly modulate the calcium dynamics and electrical activities of atrial cardiomyocytes, which are associated with the renin-angiotensin system. These findings suggest a critical role of collagen in electrical remodeling during fibrosis.

Aldosterone receptor inhibition alters the viscoelastic biomechanical behavior of the aortic wall

Dynamic mechanical disturbances in the aortic wall may lead to progressive aortic dilation and possibly aneurysmal formation. Here, we investigated the previously unexplored effects of aldosterone inhibition on aortic wall viscoelastic properties in hyperlipidemic rabbits. Thirty-six New Zealand male rabbits, fed a standard diet for four weeks, were separated into three groups: control (C; n = 10), standard diet; eplerenone (A; n = 12), hyperlipidemic diet plus 100 mg/kg/d eplerenone (last 4 weeks); and vehicle (V; n = 14), hyperlipidemic diet (no eplerenone). After eight weeks, animals were sacrificed and rectangular strips from the aortic wall, cut in radial and axial orientations, were prepared. Fresh, saline-wetted strips at 37 degrees C were subjected to cyclic sinusoidal elongation from zero to 20% of the resting length at a frequency of 1 Hz. The dynamic biomechanical viscoelastic characteristics, 'elastin phase' low modulus (E(L)), 'collagen phase' high modulus (E(H)) and dissipated-energy index, were determined. Aortic tissue preparations were also examined histologically. Eplerenone increased aldosterone concentrations but did not affect blood pressure, cholesterol or potassium concentrations. There was a significant reduction of E(H) (from 3.40 to 1.80 MPa; P < 0.01) and E(L) (from 0.46 to 0.27 MPa; P < 0.05) in group A in the radial direction compared with group C. In the axial direction E(L) significantly increased in group A compared with group V (from 0.42 to 1.11 MPa; P < 0.01). Energy dissipation was not significantly different among groups, although there was a trend toward higher values in group A for both directions. Histological assessments revealed no significant differences in collagen or elastic fibers among groups. In conclusion, aldosterone receptor inhibition altered the viscoelastic properties of the aortic wall in hyperlipidemic rabbits without detectable microscopic changes in elastic or collagen fibers, an effect that progressively might predispose to dilation and/or aneurysmal formation.

Airway hyperresponsiveness is dissociated from airway wall structural remodeling

Nonasthmatic eosinophilic bronchitis (EB) has emerged as a useful tool to study the structural and inflammatory mechanisms of airway hyperresponsiveness (AHR) in asthma. We have previously shown that vascular remodeling and reticular basement membrane (RBM) thickening are present in EB. However, it is not known whether other features of structural remodeling including increased airway smooth muscle (ASM) mass, matrix deposition, and glandular hyperplasia are also present in EB. We sought to determine whether structural remodeling occurs in EB and is associated with AHR and airflow limitation. Forty-two patients with asthma, 21 patients with EB, and 19 healthy volunteers were recruited. ASM area, RBM thickness, collagen 3 deposition, glandular area, mast cells, and granulocytes were assessed in bronchial biopsy samples. Nonasthmatic eosinophilic bronchitis and asthma were associated with a significant increase in ASM mass and RBM thickness compared with healthy subjects. In contrast, we did not observe any significant differences in collagen 3 deposition in the lamina propria and ASM or the % area of glands in the lamina propria. Univariate analysis demonstrated that mast cell numbers in the ASM were the only feature of remodeling associated with AHR (beta = -0.51; P = .004). Stepwise linear regression revealed that a combination of mast cell numbers in the ASM (beta = -0.43) and disease duration (beta = -0.25; model-adjusted R(2) = 0.26; P = .027) best modeled AHR. Mast cell localization to the ASM bundle, but not structural remodeling of the airway wall, is associated with AHR in asthma.

The Effect of Serial Monolayer Passaging on the Collagen Expression Profile of Outer and Inner Anulus Fibrosus Cells

Sheep outer and inner anulus fibrosus cells were isolated and analyzed to determine the effect of serial monolayer passaging on their phenotype. To characterize the effect of sequential serial passage on outer and inner anulus cells to determine at which point passaged cells are significantly different from freshly isolated cells. Previous studies show that chondrocytic cells lose their differentiated phenotype with sequential monolayer passage. Although intervertebral disc cells are similar, to our knowledge, a complete characterization of passage effects has not been performed. Sheep outer and inner anulus cells were isolated, serially passaged, and evaluated for changes in cellular morphology, collagen I and II gene expression and protein elaboration, and total protein and deoxyribonucleic acid content. Outer anulus cells displayed an elongated morphology, while inner anulus cells were initially polygonal and became more fibroblast-like with passage. At low passage, outer anulus cells showed higher collagen I expression, while inner anulus cells indicated higher collagen II expression. At high passage, collagen I expression increased for inner anulus cells and decreased for outer anulus cells, whereas collagen II expression decreased for both cell types. Immunohistochemical staining confirmed gene expression results. The differences in expression profiles of outer and inner anulus cells support previous findings that zonal differences exist between the cell types. Up to passage 2, both cell types were not significantly different from freshly isolated cells and maintained distinct phenotypic characteristics. However, after 6 sequential passages, outer and inner anulus cells became morphologically indistinguishable, and displayed no significant differences in collagen I gene and protein expression, thus becoming a more homogeneous population. As such, serial monolayer passaging has a marked effect on disc cell behavior, and is an important factor to consider when designing and evaluating in vitro studies and for potential cell-based therapies for disc repair.

Comparative analysis of the penis corpora cavernosa in controls and patients with erectile dysfunction

To evaluate the presence of structural disorders of the corpora cavernosa in patients with erectile dysfunction (ED), as despite new drugs being effective in many men with ED, some aspects of structural disorders of the corpora cavernosa remain unknown. Biopsy specimens were taken from the corpora cavernosa of seven patients (mean age 57.8 years, range 51-72) with severe ED who had a penile prosthesis implanted. The controls tissues were fragments of corpora cavernosa obtained from autopsies of six men (mean age 52.3 years, range 40-66) who died from causes unrelated to the urogenital system. For light microscopy, the specimens were processed routinely to paraffin wax, and by immunohistochemistry to evaluate elastic fibres, and by Masson's trichrome to analyse collagen and smooth muscle fibres. Stereological methods were used to quantitatively evaluate the different elements (as a percentage). The percentages of the different elements in the human penis of controls and men with ED, respectively, were: elastic fibres 13.2% and 9.1%; collagen fibres 40.8% and 41.6%; and smooth muscle, 40.4% and 42%. In patients with ED there was a statistically significant reduction in the percentage of elastic fibres, but no statistically significant difference in collagen and smooth muscle fibres, and no appreciable differences in collagen distribution between the groups.

106 MMP-1, TIMP-1, collagen I and collagen III genes expression in myocardium from patients with obstructive HCM

Purpose: To estimate matrix metalloproteinase 1 (MMP-1), tissue inhibitor of matrix metalloproteinases type 1 (TIMP-1), collagen I and collagen Ill genes expression in intraventricular septum (IVS) from patients with obstructive hypertrophic cardiomyopathy (OHCM). Materials and Methods: We examined 22 patients with OHCM. The mean age was 33,04-14,4; male/female ratio was 1/1. We used 8 patients with essential hypertension (EH) stage llI as controls. We used myocardial samples obtained as a result of cardiosurgery in OHCM group and as a result of express-autopsy in EH group for RNA isolation with subsequent RT-PCR. Results: We did not reveal any significant differences in collagen I gene expression between OHCM and EH groups. Collagen llI gene expression in IVS area of hypertrophy was significantly higher in OHCM group (0,214-0,05) to compare with EH group (0,104-0,06), p<0,05. MMP1 gene expression in IVS area of hypertrophy was 5,6 times higher in OHCM group (0,84-0,22) to compare with EH group (0,144-0,08), p<0,05. We revealed a negative correlation between MMP-1 and TIMP1 genes expression in OHCM group(R=-0,48;p<0,01). Conclusion: We revealed a disproportional increase of collagen Ill and TIMP-1 genes expression in the area of asymmetrical IVS hypertrophy in patients with OHCM.

IGF-I and TGF-beta 1 incorporated in a poly(D,L-lactide) implant coating stimulates osteoblast differentiation and collagen-1 production but reduces osteoblast proliferation in cell culture.

Previous in vivo studies revealed a stimulating effect of locally applied IGF-I and TGF-beta1 released from poly(D,L-lactide)-coated titanium implants on rat and porcine fracture healing. The purpose of the present study was to evaluate the effect of IGF-I (5% w/w) and TGF-beta1 (1% w/w) and the carrier PDLLA on osteoblasts in cell culture to improve the understanding of these growth factors. The well-characterized human osteoblast cell line hFOB 1.19 was used in the study. The implants and cells were cocultured in a noncontact manner. The cells were incubated for 10 days in total, and the implants (n = 6 each group and time point) were added for 1 h, 12 h, 24 h, 2 d, 4 d, or 10 d. To analyze a possible effect of the growth factors or the coating, cell proliferation, metabolism, and differentiation were investigated. As an indicator for differentiation the production of collagen I was chosen. All experimental groups showed comparable cell vitality. No change in the pH of the medium was detectable between the analyzed groups. When the effect of the titanium implant and the PDLLA coating were compared with the control culture, no differences in proliferation, metabolic activity, and collagen I production were detectable. The osteoblasts treated with IGF-I and TGF-beta1 released from PDLLA revealed a significantly enhanced collagen I production with a decrease in proliferation and metabolic activity compared to the other groups. No significant differences in collagen I production were seen due to the incubation time points. None of the experimental groups evoked an immunological response on mouse macrophages. In conclusion, the PDLLA-carrier showed no negative effect on osteoblasts, whereas the incorporated growth factors stimulated osteoblast differentiation.

Identification of broiler chicken meat using a visible/near-infrared spectroscopic technique

A near-infrared spectroscopic method was developed with a dummy regression technique to differentiate meat originating from broilers and Chinese local chickens. Best classification accuracies of 100%, 92%, 96% and 92% were achieved for minced thigh meat, minced breast meat, breast cut without skin and breast cut with skin respectively. Comparison among the regression models of MLR, PCR, PLS and mPLS did not show obvious differences in classification accuracy. Scatter correction and derivative treatment of the spectral data before discriminant analysis often improved the classification accuracy for minced meat, while for meat cuts, spectra without pretreatment produced better classification. In general, using the full spectrum of 400–2500 nm produced satisfactory classification. The spectrum in the visible region of 400–750 nm, the short-wavelength NIR region of 750–1100 nm or the long-wavelength NIR region of 1100–2500 nm can also produce satisfactory classification depending on sample presentation methods and regression models. The spectroscopic classification was supported by physical and chemical properties of meat samples, which showed significant differences in collagen and fat contents and pH and chromatic values between the two groups of chickens. The results of the present study indicate that NIR spectroscopy can be used to identify broiler meat or carcass from those of local chickens. © 1999 Society of Chemical Industry

Abnormalities of collagen cross-linkage in posterior communicating artery aneurysms: a preliminary study.

The collagen of tissues submitted to mechanical load (i.e. arterial wall) is characterised by the presence of intermolecular covalent cross-links (hydroxylysyl pyridinoline or pyridinoline: PYD; and lysyl-pyridinoline or deoxypyridinoline: DPD) which stabilise the molecular structure. In this preliminary study we look for quantitative or qualitative alterations of collagen cross-linkage in intracranial aneurysms of posterior communicating artery (PCoA) which may be considered a site of elective weakening in the intracranial arterial circulation, being one of the most frequent locations of intracranial aneurysms and of so called "infundibular widening'. We analysed the collagen cross-linkage in 6 autopsy samples of intracranial arterial segments of the Circle of Willis which were removed from patients whose cause of death was other than cerebral hemorrhage, and in 6 samples of intracranial PCoA aneurysms obtained at surgery. The analysis of cross-links showed that there was no significant difference in collagen and cross-link content between autopic and aneurysmatic samples except for PCoA. In autoptic nonaneurysmatic samples of PCoA a lower content of PYD than in internal carotid artery (ICA) and a lower content of DPD than in all other arterial segments has been demonstrated; moreover a lower content of cross-links (DPD + PYD/ Collagen) was evident in nonaneurysmatic PCoA samples when compared to other segments (ICA and Anterior Communicating Artery). On the other hand, the mean content of DPD was significantly lower in PCoA aneurysms than in nonaneurysmatic samples of the artery and moreover, a significantly low content of cross-links (DPD + PYD/Collagen) is overemphasised in PCoA aneurysms, suggesting that the peculiar lower content of DPD in PCoA arteries may be considered the expression of minor resistance of the arterial wall at this site, and may be related to the higher incidence of aneurysms or infundibular widening of this arterial segment.

Spectroscopic changes in human dentine exposed to various storage solutions — short term

Objectives: Tooth storage conditions in studies of dentine have not been standardized. The objective of this investigation was to determine the effects of storage solution and time on dentine, using spectroscopic methods. Methods: Twenty-seven dentine disks (0.75 mm thick) were prepared from freshly extracted third molars with documented histories. The teeth were sectioned parallel to the occlusal plane using a diamond saw. Specimens were analysed non-destructively for surface chemical changes using specular reflectance infra-red Fourier transform spectroscopy (SRIFTS), and changes in optical properties were measured in the ultraviolet, visible and near-infra-red spectral ranges with an integrating sphere spectrophotometer (UV/VIS/NIR) before and after storage. A minimum of five specimens were stored in distilled water with 0.02% thymol, purified and filtered water, phosphate buffered saline with 0.02% thymol, 70% ethanol or 10% buffered formalin and evaluated at 0, 1, 2, 7, 14, 21 and 28 days. After each measurement, specimens were stored in fresh solutions. IR spectra were obtained from 4400 to 400 wave-numbers (cm −1). Optical properties of reflectance and transmittance were determined from 200 to 2500 nm, and absorbance was calculated. Results: Analysis of the IR spectra indicated no significant differences in collagen or mineral peak positions for any solution at any time period. There were significant differences in peak intensities of dentine stored in distilled water, purified and filtered water and phosphate-buffered saline over time in the IR spectra. Furthermore, differences in optical properties of dentine disks were detected by UV/VIS/NIR. Conclusion: It was concluded that changes in surface chemistry and optical properties of dentine occur as a function of storage solution and time, which must be considered when studying dentine.