Science.gov

Sample records for activity collagen synthesis

  1. Regulation of collagen synthesis by ascorbic acid.

    PubMed Central

    Murad, S; Grove, D; Lindberg, K A; Reynolds, G; Sivarajah, A; Pinnell, S R

    1981-01-01

    After prolonged exposure to ascorbate, collagen synthesis in cultured human skin fibroblasts increased approximately 8-fold with no significant change in synthesis of noncollagen protein. This effect of ascorbate appears to be unrelated to its cofactor function in collagen hydroxylation. The collagenous protein secreted in the absence of added ascorbate was normal in hydroxylysine but was mildly deficient in hydroxyproline. In parallel experiments, lysine hydroxylase (peptidyllysine, 2-oxoglutarate:oxygen 5-oxidoreductase, EC 1.14.11.4) activity increased 3-fold in response to ascorbate administration whereas proline hydroxylase (prolyl-glycyl-peptide, 2-oxoglutarate:oxygen oxidoreductase, EC 1.14.11.2) activity decreased considerably. These results suggest that collage polypeptide synthesis, posttranslational hydroxylations, and activities of the two hydroxylases are independently regulated by ascorbate. PMID:6265920

  2. In vivo determination of arterial collagen synthesis in atherosclerotic rabbits

    SciTech Connect

    Opsahl, W.P.; DeLuca, D.J.; Ehrhart, L.A.

    1986-03-01

    Collagen and non-collagen protein synthesis rates were determined in vivo in tissues from rabbits fed a control or atherogenic diet supplemented with 2% peanut oil and 0.25% cholesterol for 4 months. Rabbits received a bolus intravenous injection of L-(/sup 3/H)-proline (1.0 mCi/kg) and unlabeled L-proline (7 mmoles/kg) in 0.9% NaCl. Plasma proline specific activity decreased only 20% over 5 hr and was similar to the specific activity of free proline in tissues. Thoracic aortas from atherosclerotic rabbits exhibited raised plaques covering at least 75% of the surface. Thoracic intima plus a portion of the media (TIM) was separated from the remaining media plus adventitia (TMA). Dry delipidated weight, total collagen content, and collagen as a percent of dry weight were increased significantly in the TIM of atherosclerotic rabbits. Collagen synthesis rates and collagen synthesis as a percent of total protein synthesis were likewise increased both in the TIM and in the abdominal aortas. No differences from controls either in collagen content or collagen synthesis rates were observed in the TMA, lung or skin. These results demonstrate for the first time in vivo that formation of atherosclerotic plaques is associated with increased rates of collagen synthesis. Furthermore, as previously observed with incubations in vitro, collagen synthesis was elevated to a greater extent than noncollagen protein synthesis in atherosclerotic aortas from rabbits fed cholesterol plus peanut oil.

  3. Solid-Phase Synthesis, Characterization, and Cellular Activities of Collagen-Model Nanodiamond-Peptide Conjugates

    PubMed Central

    Knapinska, Anna M.; Tokmina-Roszyk, Dorota; Amar, Sabrina; Tokmina-Roszyk, Michal; Mochalin, Vadym N.; Gogotsi, Yury; Cosme, Patrick; Terentis, Andrew C.; Fields, Gregg B.

    2015-01-01

    Nanodiamonds (NDs) have received considerable attention as potential drug delivery vehicles. NDs are small (~5 nm diameter), can be surface modified in a controllable fashion with a variety of functional groups, and have little observed toxicity in vitro and in vivo. However, most biomedical applications of NDs utilize surface adsorption of biomolecules, as opposed to covalent attachment. Covalent modification provides reliable and reproducible ND–biomolecule ratios, and alleviates concerns over biomolecule desorption prior to delivery. The present study has outlined methods for the efficient solid-phase conjugation of ND to peptides and characterization of ND–peptide conjugates. Utilizing collagen-derived peptides, the ND was found to support or even enhance the cell adhesion and viability activities of the conjugated sequence. Thus, NDs can be incorporated into peptides and proteins in a selective manner, where the presence of the ND could potentially enhance the in vivo activities of the biomolecule it is attached to. PMID:25753561

  4. Urinary polypeptides related to collagen synthesis

    PubMed Central

    Krane, Stephen M.; Muñoz, Alberto J.; Harris, Edward D.

    1970-01-01

    Of the total urinary hydroxyproline in normal subjects and those with skeletal disorders, between 4 and 20% was nondialyzable. In some patients with Paget's disease of bone, hyperparathyroidism with osteitis fibrosa, hyperphosphatasia, and extensive fibrous dysplasia the total urinary hydroxyproline was sufficiently high to permit purification of this polypeptide hydroxyproline by gel filtration and ion exchange chromatography. The partially purified polypeptides had molecular weights between 4500 and 10,000 and amino acid compositions and physical properties resembling those of gelatin. The polypeptide fractions also contained neutral sugar and glucosamine. These fragments had been shown to be susceptible to cleavage by purified bacterial collagenase suggesting the presence of the sequence-Pro-X-Gly-Pro-Y-. After administration of proline-14C to patients with Paget's disease hydroxyproline-14C was excreted in the urine. The hydroxyproline-14C specific activity reached a peak in 2-4 hr and declined rapidly. The specific activity of the polypeptide (retentate) portion was severalfold greater than that of the raw urine and diffusate. When the labeled urines were subjected to gel filtration the hydroxyproline-14C fractions of highest molecular weight which were eluted first from the columns had the highest specific activities. Exposure of the hydroxyproline-14C-containing polypeptides to bacterial collagenase rendered them dialyzable. Four patients with hyperparathyroidism and osteitis fibrosa were studied before and after removal of a parathyroid adenoma, a period of transition from a predominance of bone collagen resorption to one of relatively increased bone collagen synthesis. The total urinary hydroxyproline fell rapidly after operation whereas the ratio of the polypeptide fraction to the total rose three- to fourfold. The results of these studies suggest that the urinary polypeptides represent fragments of collagen related to collagen synthesis. Changes in the

  5. Nicotine inhibits collagen synthesis and alkaline phosphatase activity, but stimulates DNA synthesis in osteoblast-like cells

    SciTech Connect

    Ramp, W.K.; Lenz, L.G.; Galvin, R.J. )

    1991-05-01

    Use of smokeless tobacco is associated with various oral lesions including periodontal damage and alveolar bone loss. This study was performed to test the effects of nicotine on bone-forming cells at concentrations that occur in the saliva of smokeless tobacco users. Confluent cultures of osteoblast-like cells isolated from chick embryo calvariae were incubated for 2 days with nicotine added to the culture medium (25-600 micrograms/ml). Nicotine inhibited alkaline phosphatase in the cell layer and released to the medium, whereas glycolysis (as indexed by lactate production) was unaffected or slightly elevated. The effects on medium and cell layer alkaline phosphatase were concentration dependent with maximal inhibition occurring at 600 micrograms nicotine/ml. Nicotine essentially did not affect the noncollagenous protein content of the cell layer, but did inhibit collagen synthesis (hydroxylation of ({sup 3}H)proline and collagenase-digestible protein) at 100, 300, and 600 micrograms/ml. Release of ({sup 3}H)hydroxyproline to the medium was also decreased in a dose-dependent manner, as was the collagenase-digestible protein for both the medium and cell layer. In contrast, DNA synthesis (incorporation of ({sup 3}H)thymidine) was more than doubled by the alkaloid, whereas total DNA content was slightly inhibited at 600 micrograms/ml, suggesting stimulated cell turnover. Morphologic changes occurred in nicotine-treated cells including rounding up, detachment, and the occurrence of numerous large vacuoles. These results suggest that steps to reduce the salivary concentration of nicotine in smokeless tobacco users might diminish damaging effects of this product on alveolar bone.

  6. Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonist inhibits collagen synthesis in human hypertrophic scar fibroblasts by targeting Smad3 via miR-145

    SciTech Connect

    Zhu, Hua-Yu; Li, Chao; Zheng, Zhao; Zhou, Qin; Guan, Hao; Su, Lin-Lin; Han, Jun-Tao; Zhu, Xiong-Xiang; Wang, Shu-yue; Li, Jun Hu, Da-Hai

    2015-03-27

    The transcription factor peroxisome proliferator-activated receptor-γ (PPAR-γ) functions to regulate cell differentiation and lipid metabolism. Recently, its agonist has been documented to regulate extracellular matrix production in human dermal fibroblasts. This study explored the underlying molecular mechanisms and gene interactions in hypertrophic scar fibroblasts (HSFBs) in vitro. HSFBs were cultured and treated with or without PPAR-γ agonist or antagonist for gene expression. Bioinformatical analysis predicted that miR-145 could target Smad3 expression. Luciferase assay was used to confirm such an interaction. The data showed that PPAR-γ agonist troglitazone suppressed expression of Smad3 and Col1 in HSFBs. PPAR-γ agonist induced miR-145 at the gene transcriptional level, which in turn inhibited Smad3 expression and Col1 level in HSFBs. Furthermore, ELISA data showed that Col1 level in HSFBs was controlled by a feedback regulation mechanism involved in PPAR-γ agonist and antagonist-regulated expression of miR-145 and Smad3 in HSFBs. These findings indicate that PPAR-γ-miR-145-Smad3 axis plays a role in regulation of collagen synthesis in HSFBs. - Highlights: • PPAR-γ agonist inhibits collagen synthesis in HSFBs. • Smad3 and type I collagen expression are decreased by PPAR-γ agonist. • miR-145 expression is increased by PPAR-γ agonist in HSFBs. • Increased miR-145 inhibits collagen synthesis by targeting Smad3. • miR-145 regulates collagen synthesis.

  7. Screening of medicinal and edible plants in Okinawa, Japan, for enhanced proliferative and collagen synthesis activities in NB1RGB human skin fibroblast cells.

    PubMed

    Takahashi, Makoto; Asikin, Yonathan; Takara, Kensaku; Wada, Koji

    2012-01-01

    To identify plants with bioactive potential for skin care, methanol extracts of 56 plant parts from 47 medical and edible plants cultivated in Okinawa were tested for their proliferative effects on NB1RGB skin fibroblast cells. Extracts from six plants, Bischofia javanica, Colocasia esculenta, Melaleuca alternifolia, Piper angustifolia, Jasminum sambac, and Curcuma longa, showed higher NB1RGB cell proliferation activity (>10%) than the control, at various concentrations. Among the six extracts, only the C. longa extract caused an increase in collagen synthesis in NB1RGB cells, as compared to treatment with the positive control, ascorbic acid (AsA). Expression of the collagen synthesis marker, transforming growth factor-β1, was higher after treatment with the C. longa extract than with AsA. PMID:23221723

  8. TSLP Is a Potential Initiator of Collagen Synthesis and an Activator of CXCR4/SDF-1 Axis in Keloid Pathogenesis.

    PubMed

    Shin, Jung U; Kim, Seo Hyeong; Kim, Hyeran; Noh, Ji Yeon; Jin, Shan; Park, Chang Ook; Lee, Won Jai; Lee, Dong Won; Lee, Ju Hee; Lee, Kwang Hoon

    2016-02-01

    Recently, thymic stromal lymphopoietin (TSLP), which is well studied in allergic diseases, has been reported in fibrotic diseases, including idiopathic pulmonary fibrosis and atopic dermatitis fibrosis. However, the role of TSLP in keloid is obscure. In this study, we assessed the expression of TSLP in keloid tissue and investigated the possible role of TSLP in keloid pathogenesis. We observed that TSLP expression was increased in keloid tissue compared to normal tissue. Furthermore, TSLP treatment induced increased collagen I and collagen III expression in fibroblasts via transforming growth factor-?; however, there was higher expression in keloid fibroblasts compared to normal fibroblasts. Stromal cell-derived factor-1?, which was recently reported to enhance wound healing through recruiting bone marrow-derived mesenchymal stem cells to the wound area, increased after TSLP treatment in fibroblasts and was primarily expressed in ?-smooth muscle action-positive myofibroblasts in keloid tissue. Furthermore, fibrocytes expressing CXCR4, a stromal cell-derived factor-1? receptor, were significantly increased in keloid tissue compared to normal tissue. Finally, intradermal TSLP injection on BALB/c mice increased stromal cell-derived factor-1? expression and CXCR4(+) fibrocytes infiltration. Our data suggest that TSLP is a potent inducer of collagen and transforming growth factor-? production in keloid fibroblasts. In addition, it might activate the CXCR4/stromal cell-derived factor-1 axis to increase fibrocyte infiltration into the keloid tissue. PMID:26824743

  9. A Recombinant Collagen-mRNA Platform for Controllable Protein Synthesis.

    PubMed

    Sun, Liping; Xiong, Yunjing; Bashan, Anat; Zimmerman, Ella; Shulman Daube, Shirley; Peleg, Yoav; Albeck, Shira; Unger, Tamar; Yonath, Hagith; Krupkin, Miri; Matzov, Donna; Yonath, Ada

    2015-07-01

    We have developed a collagen-mRNA platform for controllable protein production that is intended to be less prone to the problems associated with commonly used mRNA therapy as well as with collagen skin-healing procedures. A collagen mimic was constructed according to a recombinant method and was used as scaffold for translating mRNA chains into proteins. Cysteines were genetically inserted into the collagen chain at positions allowing efficient ribosome translation activity while minimizing mRNA misfolding and degradation. Enhanced green fluorescence protein (eGFP) mRNA bound to collagen was successfully translated by cell-free Escherichia coli ribosomes. This system enabled an accurate control of specific protein synthesis by monitoring expression time and level. Luciferase-mRNA was also translated on collagen scaffold by eukaryotic cell extracts. Thus we have demonstrated the feasibility of controllable protein synthesis on collagen scaffolds by ribosomal machinery.

  10. Cyclic tensile strain upregulates collagen synthesis in isolated tendon fascicles

    SciTech Connect

    Screen, Hazel R.C. . E-mail: H.R.C.Screen@qmul.ac.uk; Shelton, Julia C.; Bader, Dan L.; Lee, David A.

    2005-10-21

    Mechanical stimulation has been implicated as an important regulatory factor in tendon homeostasis. In this study, a custom-designed tensile loading system was used to apply controlled mechanical stimulation to isolated tendon fascicles, in order to examine the effects of 5% cyclic tensile strain at 1 Hz on cell proliferation and matrix synthesis. Sample viability and gross structural composition were maintained over a 24 h loading period. Data demonstrated no statistically significant differences in cell proliferation or glycosaminoglycan production, however, collagen synthesis was upregulated with the application of cyclic tensile strain over the 24 h period. Moreover, a greater proportion of the newly synthesised matrix was retained within the sample after loading. These data provide evidence of altered anabolic activity within tendon in response to mechanical stimuli, and suggest the importance of cyclic tensile loading for the maintenance of the collagen hierarchy within tendon.

  11. Laminin peptide YIGSR induces collagen synthesis in Hs27 human dermal fibroblasts

    SciTech Connect

    Yoon, Jong Hyuk; Kim, Jaeyoon; Lee, Hyeongjoo; Kim, So Young; Jang, Hwan-Hee; Ryu, Sung Ho; Kim, Beom Joon; Lee, Taehoon G.

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer We identify a function of the YIGSR peptide to enhance collagen synthesis in Hs27. Black-Right-Pointing-Pointer YIGSR peptide enhanced collagen type 1 synthesis both of gene and protein levels. Black-Right-Pointing-Pointer There were no changes in cell proliferation and MMP-1 level in YIGSR treatment. Black-Right-Pointing-Pointer The YIGSR effect on collagen synthesis mediated activation of FAK, pyk2 and ERK. Black-Right-Pointing-Pointer The YIGSR-induced FAK and ERK activation was modulated by FAK and MEK inhibitors. -- Abstract: The dermal ECM is synthesized from fibroblasts and is primarily compromised of fibrillar collagen and elastic fibers, which support the mechanical strength and resiliency of skin, respectively. Laminin, a major glycoprotein located in the basement membrane, promotes cell adhesion, cell growth, differentiation, and migration. The laminin tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide, corresponding to the 929-933 sequence of the {beta}1 chain, is known to be a functional motif with effects on the inhibition of tumor metastasis, the regulation of sensory axonal response and the inhibition of angiogenesis through high affinity to the 67 kDa laminin receptor. In this study, we identified a novel function of the YIGSR peptide to enhance collagen synthesis in human dermal fibroblasts. To elucidate this novel function regarding collagen synthesis, we treated human dermal fibroblasts with YIGSR peptide in both a time- and dose-dependent manner. According to subsequent experiments, we found that the YIGSR peptide strongly enhanced collagen type 1 synthesis without changing cell proliferation or cellular MMP-1 level. This YIGSR peptide-mediated collagen type 1 synthesis was modulated by FAK inhibitor and MEK inhibitor. This study clearly reveals that YIGSR peptide plays a novel function on the collagen type 1 synthesis of dermal fibroblasts and also suggests that YIGSR is a strong candidate

  12. Linoleate impairs collagen synthesis in primary cultures of avian chondrocytes.

    PubMed

    Watkins, B A; Xu, H; Turek, J J

    1996-06-01

    The effects of supplemental fatty acids, vitamin E (VIT E), and iron-induced oxidative stress on collagen synthesis, cellular injury, and lipid peroxidation were evaluated in primary cultures of avian epiphyseal chondrocytes. The treatments included oleic and linoleic acids (O or 50 microM) complexed with BSA and dl-alpha-tocopheryl acetate (VIT E at 0 or 100 microM). After 14 days of preculture, the chondrocytes were enriched with fatty acids for 8 days then cultured with VIT E for 2 days. The chondrocytes were then treated with ferrous sulfate (O or 20 microM) for 24 hr to induce oxidative stress. Collagen synthesis was the lowest and the activity of lactate dehydrogenase (LDH) was the highest in chondrocyte cultures treated with 50 microM linoleic acid and 0 VIT E. In contrast, VIT E supplemented at 100 microM partially restored collagen synthesis in the chondrocytes enriched with linoleic acid and lowered LDH activity in the media. The iron oxidative inducer significantly increased the values of thiobarbituric acid-reactive substances (TBARS) in the culture medium. The data showed that linoleic acid impaired chondrocyte cell function and caused cellular injury but that VIT E reversed these effects. Results from a previous study demonstrated that VIT E stimulated bone formation in chicks fed unsaturated fat, and the present findings in cultures of epiphyseal chondrocytes suggest that VIT E is important for chondrocyte function in the presence of polyunsaturated fatty acids. VIT E appears to be beneficial for growth cartilage biology and in optimizing bone growth.

  13. The BRAFV600E inhibitor, PLX4032, increases type I collagen synthesis in melanoma cells

    PubMed Central

    Jenkins, Molly H.; Croteau, Walburga; Mullins, David W.; Brinckerhoff, Constance E.

    2016-01-01

    Vertical growth phase (VGP) melanoma is frequently metastatic, a process mediated by changes in gene expression, which are directed by signal transduction pathways in the tumor cells. A prominent signaling pathway is the Ras-Raf-Mek-Erk MAPK pathway, which increases expression of genes that promote melanoma progression. Many melanomas harbor a mutation in this pathway, BRAFV600E, which constitutively activates MAPK signaling and expression of downstream target genes that facilitate tumor progression. In BRAFV600E melanoma, the small molecule inhibitor, vemurafenib (PLX4032), has revolutionized therapy for melanoma by inducing rapid tumor regression. This compound down-regulates the expression of many genes. However, in this study, we document that blocking the Ras-Raf-Mek-Erk MAPK pathway, either with an ERK (PLX4032) or a MEK (U1026) signaling inhibitor, in BRAFV600E human and murine melanoma cell lines increases collagen synthesis in vitro and collagen deposition in vivo. Since TGFβ signaling is a major mediator of collagen synthesis, we examined whether blocking TGFβ signaling with a small molecule inhibitor would block this increase in collagen. However, there was minimal reduction in collagen synthesis in response to blocking TGFβ signaling, suggesting additional mechanism(s), which may include activation of the p38 MAPK pathway. Presently, it is unclear whether this increased collagen synthesis and deposition in melanomas represent a therapeutic benefit or an unwanted “off target” effect of inhibiting the Ras-Raf-Erk-Mek pathway. PMID:25989506

  14. Collagen degrading activity associated with Mycobacterium species

    PubMed Central

    Masso, F; Paez, A; Varela, E; d Diaz; Zenteno, E; Montano, L

    1999-01-01

    BACKGROUND—The mechanism of Mycobacterium tuberculosis penetration into tissues is poorly understood but it is reasonable to assume that there is a contribution from proteases capable of disrupting the extracellular matrix of the pulmonary epithelium and the blood vessels. A study was undertaken to identify and characterise collagen degrading activity of M tuberculosis.
METHODS—Culture filtrate protein extract (CFPE) was obtained from reference mycobacterial strains and mycobacteria isolated from patients with tuberculosis. The collagen degrading activity of CFPE was determined according to the method of Johnson-Wint using 3H-type I collagen. The enzyme was identified by the Birkedal-Hansen and Taylor method and its molecular mass determined by SDS-PAGE and Sephacryl S-300 gel filtration chromatography using an electroelution purified enzyme.
RESULTS—CFPE from Mycobacterium tuberculosis strain H37Rv showed collagenolytic activity that was four times higher than that of the avirulent strain H37Ra. The 75 kDa enzyme responsible was divalent cation dependent. Other mycobacterial species and those isolated from patients with tuberculosis also had collagen degrading activity.
CONCLUSIONS—Mycobacterium species possess a metalloprotease with collagen degrading activity. The highest enzymatic activity was found in the virulent reference strain H37Rv.

 PMID:10212111

  15. Collagen synthesis and carbohydrate metabolism of rachitic bone

    PubMed Central

    Paterson, C. R.; Fourman, P.

    1968-01-01

    1. This paper reports studies on the metabolism of bone from normal chicks and from chicks with vitamin D-deficiency rickets. Both in vitro and in vivo there was an increased incorporation of [14C]proline into collagen hydroxyproline by rachitic bone. The proportion of the collagen that was soluble in cold salt solutions was greater with the rachitic bone. These results show that in rickets there is an increased synthesis of bone collagen, but they do not provide any evidence of a defect in the maturation of collagen. 2. Rachitic bone incubated aerobically in vitro consumed more glucose and released more lactate than normal bone. Bone from rachitic chicks treated with vitamin D 48hr. previously had rates of glycolysis that were nearly normal. Though we were unable to show any direct action of vitamin D in vitro, we consider that vitamin D probably has a direct action on bone, possibly related to matrix biosynthesis. PMID:5669840

  16. Effects of Panax ginseng extract on human dermal fibroblast proliferation and collagen synthesis.

    PubMed

    Lee, Geum-Young; Park, Kang-Gyun; Namgoong, Sik; Han, Seung-Kyu; Jeong, Seong-Ho; Dhong, Eun-Sang; Kim, Woo-Kyung

    2016-03-01

    Current studies of Panax ginseng (or Korean ginseng) have demonstrated that it has various biological effects, including angiogenesis, immunostimulation, antimicrobial and anti-inflammatory effects. Therefore, we hypothesised that P. ginseng may also play an important role in wound healing. However, few studies have been conducted on the wound-healing effects of P. ginseng. Thus, the purpose of this in vitro pilot study was to determine the effects of P. ginseng on the activities of fibroblasts, which are key wound-healing cells. Cultured human dermal fibroblasts were treated with one of six concentrations of P. ginseng: 0, 1, 10 and 100 ng/ml and 1 and 10 µg/ml. Cell proliferation was determined 3 days post-treatment using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay, and collagen synthesis was evaluated by the collagen type I carboxy-terminal propeptide method. Cell proliferation levels and collagen synthesis were compared among the groups. The 10 ng/ml to 1 µg/ml P. ginseng treatments significantly increased cell proliferation, and the 1 ng/ml to 1 µg/ml concentrations significantly increased collagen synthesis. The maximum effects for both parameters were observed at 10 ng/ml. P. ginseng stimulated human dermal fibroblast proliferation and collagen synthesis at an optimal concentration of 10 ng/ml.

  17. G protein-coupled receptor kinase-2 is a novel regulator of collagen synthesis in adult human cardiac fibroblasts.

    PubMed

    D'Souza, Karen M; Malhotra, Ricky; Philip, Jennifer L; Staron, Michelle L; Theccanat, Tiju; Jeevanandam, Valluvan; Akhter, Shahab A

    2011-04-29

    Cardiac fibroblasts (CF) make up 60-70% of the total cell number in the heart and play a critical role in regulating normal myocardial function and in adverse remodeling following myocardial infarction and the transition to heart failure. Recent studies have shown that increased intracellular cAMP can inhibit CF transformation and collagen synthesis in adult rat CF; however, mechanisms by which cAMP production is regulated in CF have not been elucidated. We investigated the potential role of G protein-coupled receptor kinase-2 (GRK2) in modulating collagen synthesis by adult human CF isolated from normal and failing left ventricles. Baseline collagen synthesis was elevated in failing CF and was not inhibited by β-agonist stimulation in contrast to normal controls. β-adrenergic receptor (β-AR) signaling was markedly uncoupled in the failing CF, and expression and activity of GRK2 were increased 3-fold. Overexpression of GRK2 in normal CF recapitulated a heart failure phenotype with minimal inhibition of collagen synthesis following β-agonist stimulation. In contrast, knockdown of GRK2 expression in normal CF enhanced cAMP production and led to greater β-agonist-mediated inhibition of basal and TGFβ-stimulated collagen synthesis versus control. Inhibition of GRK2 activity in failing CF by expression of the GRK2 inhibitor, GRK2ct, or siRNA-mediated knockdown restored β-agonist-stimulated inhibition of collagen synthesis and decreased collagen synthesis in response to TGFβ stimulation. GRK2 appears to play a significant role in regulating collagen synthesis in adult human CF, and increased activity of this kinase may be an important mechanism of maladaptive ventricular remodeling as mediated by cardiac fibroblasts.

  18. Prostaglandin E2 inhibits collagen synthesis in dermal fibroblasts and prevents hypertrophic scar formation in vivo.

    PubMed

    Zhao, Jingling; Shu, Bin; Chen, Lei; Tang, Jinming; Zhang, Lijun; Xie, Julin; Liu, Xusheng; Xu, Yingbin; Qi, Shaohai

    2016-08-01

    Hypertrophic scarring is a common dermal fibroproliferative disorder characterized by excessive collagen deposition. Prostaglandin E2 (PGE2 ), an important inflammatory product synthesized via the arachidonic acid cascade, has been shown to act as a fibroblast modulator and to possess antifibroblastic activity. However, the mechanism underlying the antifibrotic effect of PGE2 remains unclear. In this study, we explored the effects of PGE2 on TGF-β1-treated dermal fibroblasts in terms of collagen production and to determine the regulatory pathways involved, as well as understand the antiscarring function of PGE2 in vivo. We found that PGE2 inhibited TGF-β1-induced collagen synthesis by regulating the balance of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinase (TIMP). It did so by upregulating cAMP through the E prostanoid (EP)2 receptor. We determined that inhibition of the TGF-β1/Smad pathway by PGE2 is associated with its ability to inhibit collagen synthesis. An in vivo study further confirmed that PGE2 inhibits hypertrophic scar formation by decreasing collagen production. Our results demonstrate that the novel anti-scarring function of PGE2 is achieved by balancing MMPs/TIMP expression and decreasing collagen production. PMID:26997546

  19. Collagen synthesis in axotomized peripheral nerve: evidence against Schwann cell involvement.

    PubMed

    Eather, T F; Pollock, M

    1987-04-01

    To explore the role of Schwann cells in nerve collagen formation, fascicular collagen was measured in axotomized rat sciatic nerve after regeneration or permanent axotomy. The same increase in fascicular collagen was seen in both paradigms. Thus there is no evidence that Schwann cells unassociated with axons make an important contribution to nerve collagen synthesis after transection.

  20. Tryptase inhibitor APC 366 prevents hepatic fibrosis by inhibiting collagen synthesis induced by tryptase/protease-activated receptor 2 interactions in hepatic stellate cells.

    PubMed

    Lu, Jing; Chen, Baian; Li, Shengli; Sun, Quan

    2014-06-01

    Protease-activated receptor (PAR) 2 is a G-protein-coupled receptor that is activated by mast cell tryptase. PAR-2 activation augments profibrotic pathways through the induction of extracellular matrix proteins. PAR-2 is widely expressed in hepatic stellate cells (HSCs), but the role of tryptase/PAR-2 interaction in liver fibrosis is unclear. We studied the development of bile duct ligation (BDL)-induced hepatic fibrosis in rats treated with mast cell tryptase inhibitor APC 366, and showed that APC 366 reduced hepatic fibrosis scores, collagen content and serum biochemical parameters. Reduced fibrosis was associated with decreased expression of PAR-2 and α-smooth muscle actin (α-SMA). Our findings demonstrate that mast cell tryptase induces PAR-2 activation to augment HSC proliferation and promote hepatic fibrosis in rats. Treatment with tryptase antagonists may be a novel therapeutic approach to prevent fibrosis in patients with chronic liver disease.

  1. Tryptase inhibitor APC 366 prevents hepatic fibrosis by inhibiting collagen synthesis induced by tryptase/protease-activated receptor 2 interactions in hepatic stellate cells.

    PubMed

    Lu, Jing; Chen, Baian; Li, Shengli; Sun, Quan

    2014-06-01

    Protease-activated receptor (PAR) 2 is a G-protein-coupled receptor that is activated by mast cell tryptase. PAR-2 activation augments profibrotic pathways through the induction of extracellular matrix proteins. PAR-2 is widely expressed in hepatic stellate cells (HSCs), but the role of tryptase/PAR-2 interaction in liver fibrosis is unclear. We studied the development of bile duct ligation (BDL)-induced hepatic fibrosis in rats treated with mast cell tryptase inhibitor APC 366, and showed that APC 366 reduced hepatic fibrosis scores, collagen content and serum biochemical parameters. Reduced fibrosis was associated with decreased expression of PAR-2 and α-smooth muscle actin (α-SMA). Our findings demonstrate that mast cell tryptase induces PAR-2 activation to augment HSC proliferation and promote hepatic fibrosis in rats. Treatment with tryptase antagonists may be a novel therapeutic approach to prevent fibrosis in patients with chronic liver disease. PMID:24735816

  2. Potentiation of collagen synthesis in explants of the rabbit eye by 5 beta-dihydrocortisol

    SciTech Connect

    Southren, A.L.; Hernandez, M.R.; l'Hommedieu, D.; Gordon, G.G.; Weinstein, B.I.

    1986-12-01

    The biologic effect of 5 beta-dihydrocortisol on collagen synthesis was evaluated. The metabolite was found to potentiate subthreshold levels of dexamethasone in increasing /sup 3/H-proline incorporation in cells of the outflow region of the rabbit. Digestion of the tissue with highly purified collagenase indicated that the /sup 3/H-proline was incorporated into collagen type protein. This study demonstrates another biologic activity of 5 beta-dihydrocortisol, a metabolite found to accumulate in cells cultured from trabeculectomy specimens from patients with primary open angle glaucoma.

  3. Novel synthesis and characterization of a collagen-based biopolymer initiated by hydroxyapatite nanoparticles.

    PubMed

    Bhuiyan, D; Jablonsky, M J; Kolesov, I; Middleton, J; Wick, T M; Tannenbaum, R

    2015-03-01

    In this study, we developed a novel synthesis method to create a complex collagen-based biopolymer that promises to possess the necessary material properties for a bone graft substitute. The synthesis was carried out in several steps. In the first step, a ring-opening polymerization reaction initiated by hydroxyapatite nanoparticles was used to polymerize d,l-lactide and glycolide monomers to form poly(lactide-co-glycolide) co-polymer. In the second step, the polymerization product was coupled with succinic anhydride, and subsequently was reacted with N-hydroxysuccinimide in the presence of dicyclohexylcarbodiimide as the cross-linking agent, in order to activate the co-polymer for collagen attachment. In the third and final step, the activated co-polymer was attached to calf skin collagen type I, in hydrochloric acid/phosphate buffer solution and the precipitated co-polymer with attached collagen was isolated. The synthesis was monitored by proton nuclear magnetic resonance, infrared and Raman spectroscopies, and the products after each step were characterized by thermal and mechanical analysis. Calculations of the relative amounts of the various components, coupled with initial dynamic mechanical analysis testing of the resulting biopolymer, afforded a preliminary assessment of the structure of the complex biomaterial formed by this novel polymerization process. PMID:25481742

  4. Synthesis of glycosaminoglycans by human skin fibroblasts cultured on collagen gels.

    PubMed Central

    Gallagher, J T; Gasiunas, N; Schor, S L

    1980-01-01

    A comparison has been made of the synthesis of glycosaminoglycans by human skin fibroblasts cultured on plastic or collagen gel substrata. Confluent cultures were incubated with [3H]glucosamine and Na235SO4 for 48h. Radiolabelled glycosaminoglycans were then analysed in the spent media and trypsin extracts from cells on plastic and in the medium, trypsin and collagenase extracts from cells on collagen gels. All enzyme extracts and spent media contained hyaluronic acid, heparan sulphate and dermatan sulphate. Hyaluronic acid was the main 3H-labelled component in media and enzyme extracts from cells on both substrata, although it was distributed mainly to the media fractions. Heparan sulphate was the major [35S]sulphated glycosaminoglycan in trypsin extracts of cells on plastic, and dermatan sulphate was the minor component. In contrast, dermatan sulphate was the principal [35S]sulphated glycosaminoglycan in trypsin and collagenase extracts of cells on collagen gels. The culture substratum also influenced the amounts of [35S]sulphated glycosaminoglycans in media and enzyme extracts. With cells on plastic, the medium contained most of the heparan sulphate (75%) and dermatan sulphate (> 90%), whereas the collagenase extract was the main source of heparan sulphate (60%) and dermatan sulphate (80%) from cells on collagen gels; when cells were grown on collagen, the medium contained only 5-20% of the total [35S]sulphated glycosaminoglycans. Depletion of the medium pool was probably caused by binding of [35S]sulphated glycosaminoglycans to the network of native collagen fibres that formed the insoluble fraction of the collagen gel. Furthermore, cells on collagen showed a 3-fold increase in dermatan sulphate synthesis, which could be due to a positive-feedback mechanism activated by the accumulation of dermatan sulphate in the microenvironment of the cultured cells. For comparative structural analyses of glycosaminoglycans synthesized on different substrata labelling

  5. Eccentric rehabilitation exercise increases peritendinous type I collagen synthesis in humans with Achilles tendinosis.

    PubMed

    Langberg, H; Ellingsgaard, H; Madsen, T; Jansson, J; Magnusson, S P; Aagaard, P; Kjaer, M

    2007-02-01

    It has been shown that 12 weeks of eccentric heavy resistance training can reduce pain in runners suffering from chronic Achilles tendinosis, but the mechanism behind the effectiveness of this treatment is unknown. The present study investigates the local effect of an eccentric training regime on elite soccer players suffering from chronic Achilles tendinosis on the turnover of the peritendinous connective tissue. Twelve elite male soccer players, of whom six suffered from unilateral tendinosis and six were healthy controls, participated in this study. All participants performed 12 weeks of heavy-resistance eccentric training apart from their regular training and soccer activity. Before and after the training period the tissue concentration of indicators of collagen turnover was measured by the use of the microdialysis technique. After training, collagen synthesis was increased in the initially injured tendon (n=6; carboxyterminal propeptide of type I collagen (PICP): pre 3.9+/-2.5 microg/L to post 19.7+/-5.4 microg/L, P<0.05). The collagen synthesis was unchanged in healthy tendons in response to training (n=6; PICP: pre 8.3+/-5.2 microg/L to post 11.5+/-5.0 microg/L, P>0.05). Collagen degradation, measured as carboxyterminal telopeptide region of type I collagen (ICTP), was not affected by training neither in the injured nor in the healthy tendons. The clinical effect of the 12 weeks of eccentric training was determined by using a standardized loading procedure of the Achilles tendons showing a decrease in pain in all the chronic injured tendons (VAS before 44+/-9, after 13+/-9; P<0.05), and all subjects were back playing soccer following the eccentric training regime. The present study demonstrates that chronically injured Achilles tendons respond to 12 weeks of eccentric training by increasing collagen synthesis rate. In contrast, the collagen metabolism in healthy control tendons seems not to be affected by eccentric training. These findings could indicate a

  6. Synthesis of hydrogels based on poly(NIPAM) inserted into collagen sponge.

    PubMed

    Nistor, Manuela-Tatiana; Chiriac, Aurica P; Vasile, Cornelia; Verestiuc, Liliana; Nita, Loredana Elena

    2011-10-15

    The study presents the preparation of a semi-synthetic hydrogel based on poly(N-isopropyl acrylamide-co-diethylene glycol diacrylate) inserted onto the collagen porous membrane. The synthesis of the hydrogels was performed through radical copolymerization of N-isopropyl acrylamide (NIPAM) with diethylene glycol diacrylate (DEGDA) also as crosslinking agent, using ammonium persulfate as initiator and N,N,N',N'-tetramethylethylene diamine as activator, and it was achieved in the presence of the collagen matrix. The prepared hydrogels were characterized by Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy and Scanning Electron Microscopy. The swelling behaviour of the semi-interpenetrated polymer network related on the hydrogel composition, it was also evaluated. The pore sizes of the synthesized hydrogels, much larger than the typical mesh size of a conventional hydrogel, allow to consider the hybrid hydrogel based on the inserted poly(NIPAM-co-DEGDA) onto collagen fibrils as a super-porous hydrogel. PMID:21700433

  7. Peroxisome proliferator-activated receptor-γ agonist inhibits collagen synthesis in human keloid fibroblasts by suppression of early growth response-1 expression through upregulation of miR-543 expression

    PubMed Central

    Zhu, Hua-Yu; Bai, Wen-Dong; Wang, Hong-Tao; Xie, Song-Tao; Tao, Ke; Su, Lin-Lin; Liu, Jia-Qi; Yang, Xue-Kang; Li, Jun; Wang, Yun-Chuan; He, Ting; Han, Jun-Tao; Hu, Da-Hai

    2016-01-01

    A keloid is a benign skin tumor formed by an overgrowth of granulation tissue in affected patients. Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists were reported to be able to regulate extracellular matrix production in human dermal fibroblasts. This study explored the underlying molecular mechanism of PPAR-γ agonist troglitazone treatment for fibroblasts obtained from keloid patients. The data revealed that troglitazone treatment of keloid fibroblasts (KFs) downregulated the expression of early growth response-1 (Egr1) and collagen-1 (Col1). Level of Egr1 were closely associated with KF-induced fibrosis. The miRNA profiling data revealed that miR-543 was transcriptionally activated after troglitazone treatment. Bioinformatic analysis and experimental data showed that miR-543 was able to target Egr1. ELISA data confirmed that Col1 protein in the supernatant were modulated by the feedback regulatory axis of PPAR-γ agonist-induced miR-543 to inhibit Egr1 expression, whereas PPAR-γ antagonist treatment abolished such effect on Col1 suppression in KFs. This study demonstrated that the PPAR-γ agonist-mediated miR-543 and Egr1 signaling plays an important role in the suppression of collagen synthesis in KFs. Future in vivo studies are needed to confirm these in vitro data. PMID:27429849

  8. Peroxisome proliferator-activated receptor-γ agonist inhibits collagen synthesis in human keloid fibroblasts by suppression of early growth response-1 expression through upregulation of miR-543 expression.

    PubMed

    Zhu, Hua-Yu; Bai, Wen-Dong; Wang, Hong-Tao; Xie, Song-Tao; Tao, Ke; Su, Lin-Lin; Liu, Jia-Qi; Yang, Xue-Kang; Li, Jun; Wang, Yun-Chuan; He, Ting; Han, Jun-Tao; Hu, Da-Hai

    2016-01-01

    A keloid is a benign skin tumor formed by an overgrowth of granulation tissue in affected patients. Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists were reported to be able to regulate extracellular matrix production in human dermal fibroblasts. This study explored the underlying molecular mechanism of PPAR-γ agonist troglitazone treatment for fibroblasts obtained from keloid patients. The data revealed that troglitazone treatment of keloid fibroblasts (KFs) downregulated the expression of early growth response-1 (Egr1) and collagen-1 (Col1). Level of Egr1 were closely associated with KF-induced fibrosis. The miRNA profiling data revealed that miR-543 was transcriptionally activated after troglitazone treatment. Bioinformatic analysis and experimental data showed that miR-543 was able to target Egr1. ELISA data confirmed that Col1 protein in the supernatant were modulated by the feedback regulatory axis of PPAR-γ agonist-induced miR-543 to inhibit Egr1 expression, whereas PPAR-γ antagonist treatment abolished such effect on Col1 suppression in KFs. This study demonstrated that the PPAR-γ agonist-mediated miR-543 and Egr1 signaling plays an important role in the suppression of collagen synthesis in KFs. Future in vivo studies are needed to confirm these in vitro data. PMID:27429849

  9. Photo-active collagen systems with controlled triple helix architecture

    PubMed Central

    Tronci, Giuseppe; Russell, Stephen J.; Wood, David J.

    2016-01-01

    The design of photo-active collagen systems is presented as a basis for establishing biomimetic materials with varied network architecture and programmable macroscopic properties. Following in-house isolation of type I collagen, reaction with vinyl-bearing compounds of varied backbone rigidity, i.e. 4-vinylbenzyl chloride (4VBC) and glycidyl methacrylate (GMA), was carried out. TNBS colorimetric assay, 1H-NMR and ATR-FTIR confirmed covalent and tunable functionalization of collagen lysines. Depending on the type and extent of functionalization, controlled stability and thermal denaturation of triple helices were observed via circular dichroism (CD), whereby the hydrogen-bonding capability of introduced moieties was shown to play a major role. Full gel formation was observed following photo-activation of functionalized collagen solutions. The presence of a covalent network only slightly affected collagen triple helix conformation (as observed by WAXS and ATR-FTIR), confirming the structural organization of functionalized collagen precursors. Photo-activated hydrogels demonstrated an increased denaturation temperature (DSC) with respect to native collagen, suggesting that the formation of the covalent network successfully stabilized collagen triple helices. Moreover, biocompatibility and mechanical competence of obtained hydrogels were successfully demonstrated under physiologically-relevant conditions. These results demonstrate that this novel synthetic approach enabled the formation of biocompatible collagen systems with defined network architecture and programmable macroscopic properties, which can only partially be obtained with current synthetic methods. PMID:27398214

  10. 4-[F-18]fluoroproline: A potential tracer for collagen synthesis. Radiosynthesis and biological evaluation

    SciTech Connect

    Hamacher, K.; Herz, M.; Truckenbrodt, R.

    1996-05-01

    Proline is an important constituent of the structural protein collagen. It has been shown that its fluorinated analogs (2S,4S)- and particularly (2S,4R)-4-fluoroproline are also incorporated into collagen (Gottlieb et al., Biochemistry (1965), 4: 2507). 4-[F-18]fluoroproline is therefore a potential probe for studying abnormal collagen synthesis e. g. in tumors, lung fibrosis and liver cirrhosis. We have evaluated the two diastereomeric forms using a transplantable osteosarcoma in mice as an in vivo model for elevated collagen synthesis, and a MCF 7 mamma carcinoma cell line for monolayer incubation studies.

  11. Collagen-chitosan scaffold modified with Au and Ag nanoparticles: Synthesis and structure

    NASA Astrophysics Data System (ADS)

    Rubina, M. S.; Kamitov, E. E.; Zubavichus, Ya. V.; Peters, G. S.; Naumkin, A. V.; Suzer, S.; Vasil'kov, A. Yu.

    2016-03-01

    Nowadays, the dermal biomimetic scaffolds are widely used in regenerative medicine. Collagen-chitosan scaffold one of these materials possesses antibacterial activity, good compatibility with living tissues and has been already used as a wound-healing material. In this article, collagen-chitosan scaffolds modified with Ag and Au nanoparticles have been synthesized using novel method - the metal-vapor synthesis. The nanocomposite materials are characterized by XPS, TEM, SEM and synchrotron radiation-based X-ray techniques. According to XRD data, the mean size of the nanoparticles (NPs) is 10.5 nm and 20.2 nm in Au-Collagen-Chitosan (Au-CollCh) and Ag-Collagen-Chitosan (Ag-CollCh) scaffolds, respectively in fair agreement with the TEM data. SAXS analysis of the composites reveals an asymmetric size distribution peaked at 10 nm for Au-CollCh and 25 nm for Ag-CollCh indicative of particle's aggregation. According to SEM data, the metal-carrying scaffolds have layered structure and the nanoparticles are rather uniformly distributed on the surface material. XPS data indicate that the metallic nanoparticles are in their unoxidized/neutral states and dominantly stabilized within the chitosan-rich domains.

  12. N-Phenethyl caffeamide and photodamage: protecting skin by inhibiting type I procollagen degradation and stimulating collagen synthesis.

    PubMed

    Chiang, Hsiu-Mei; Chen, Chien-Wen; Lin, Tzu-Yu; Kuo, Yueh-Hsiung

    2014-10-01

    Skin is mainly damaged by genetic and environmental factors such as ultraviolet (UV) light and pollutants. UV light is a well-known factor that causes various types of skin damage and premature aging. Reactive oxygen species (ROS) are commonly involved in the pathogenesis of skin damage by activating the metalloproteinases that break down type I collagen. This study investigated the antioxidant and antiphotodamage activity and mechanisms of N-phenethyl caffeamide (K36) in human skin fibroblasts. The results indicated that K36 demonstrated strong 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) scavenging activity, which dose-dependently reduced the production of UVB-induced intracellular ROS in human dermal fibroblasts. K36 prevented UVB-irradiation-induced type I collagen degradation by inhibiting the expression of matrix metalloproteins-1, -3, and -9 and the phosphorylation of mitogen-activated protein (MAP) kinases. Furthermore, K36 elevated collagen synthesis in skin fibroblasts by inhibiting UVB-induced Smad7 overexpression. K36 downregulated the expression of the transcription factor, activator protein-1 (AP-1). Our results indicated that K36 exhibited antioxidant properties and prevented skin collagen degradation caused by UV exposure and the stimulation of collagen synthesis, which suggests the potential use of K36 in preventing photodamage. PMID:25019243

  13. N-Phenethyl caffeamide and photodamage: protecting skin by inhibiting type I procollagen degradation and stimulating collagen synthesis.

    PubMed

    Chiang, Hsiu-Mei; Chen, Chien-Wen; Lin, Tzu-Yu; Kuo, Yueh-Hsiung

    2014-10-01

    Skin is mainly damaged by genetic and environmental factors such as ultraviolet (UV) light and pollutants. UV light is a well-known factor that causes various types of skin damage and premature aging. Reactive oxygen species (ROS) are commonly involved in the pathogenesis of skin damage by activating the metalloproteinases that break down type I collagen. This study investigated the antioxidant and antiphotodamage activity and mechanisms of N-phenethyl caffeamide (K36) in human skin fibroblasts. The results indicated that K36 demonstrated strong 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) scavenging activity, which dose-dependently reduced the production of UVB-induced intracellular ROS in human dermal fibroblasts. K36 prevented UVB-irradiation-induced type I collagen degradation by inhibiting the expression of matrix metalloproteins-1, -3, and -9 and the phosphorylation of mitogen-activated protein (MAP) kinases. Furthermore, K36 elevated collagen synthesis in skin fibroblasts by inhibiting UVB-induced Smad7 overexpression. K36 downregulated the expression of the transcription factor, activator protein-1 (AP-1). Our results indicated that K36 exhibited antioxidant properties and prevented skin collagen degradation caused by UV exposure and the stimulation of collagen synthesis, which suggests the potential use of K36 in preventing photodamage.

  14. Epithelial sodium channel modulates platelet collagen activation.

    PubMed

    Cerecedo, Doris; Martínez-Vieyra, Ivette; Alonso-Rangel, Lea; Benítez-Cardoza, Claudia; Ortega, Arturo

    2014-03-01

    Activated platelets adhere to the exposed subendothelial extracellular matrix and undergo a rapid cytoskeletal rearrangement resulting in shape change and release of their intracellular dense and alpha granule contents to avoid hemorrhage. A central step in this process is the elevation of the intracellular Ca(2+) concentration through its release from intracellular stores and on throughout its influx from the extracellular space. The Epithelial sodium channel (ENaC) is a highly selective Na(+) channel involved in mechanosensation, nociception, fluid volume homeostasis, and control of arterial blood pressure. The present study describes the expression, distribution, and participation of ENaC in platelet migration and granule secretion using pharmacological inhibition with amiloride. Our biochemical and confocal analysis in suspended and adhered platelets suggests that ENaC is associated with Intermediate filaments (IF) and with Dystrophin-associated proteins (DAP) via α-syntrophin and β-dystroglycan. Migration assays, quantification of soluble P-selectin, and serotonin release suggest that ENaC is dispensable for migration and alpha and dense granule secretion, whereas Na(+) influx through this channel is fundamental for platelet collagen activation.

  15. Laminin peptide YIGSR induces collagen synthesis in Hs27 human dermal fibroblasts.

    PubMed

    Yoon, Jong Hyuk; Kim, Jaeyoon; Lee, Hyeongjoo; Kim, So Young; Jang, Hwan-Hee; Ryu, Sung Ho; Kim, Beom Joon; Lee, Taehoon G

    2012-11-23

    The dermal ECM is synthesized from fibroblasts and is primarily compromised of fibrillar collagen and elastic fibers, which support the mechanical strength and resiliency of skin, respectively. Laminin, a major glycoprotein located in the basement membrane, promotes cell adhesion, cell growth, differentiation, and migration. The laminin tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide, corresponding to the 929-933 sequence of the β1 chain, is known to be a functional motif with effects on the inhibition of tumor metastasis, the regulation of sensory axonal response and the inhibition of angiogenesis through high affinity to the 67kDa laminin receptor. In this study, we identified a novel function of the YIGSR peptide to enhance collagen synthesis in human dermal fibroblasts. To elucidate this novel function regarding collagen synthesis, we treated human dermal fibroblasts with YIGSR peptide in both a time- and dose-dependent manner. According to subsequent experiments, we found that the YIGSR peptide strongly enhanced collagen type 1 synthesis without changing cell proliferation or cellular MMP-1 level. This YIGSR peptide-mediated collagen type 1 synthesis was modulated by FAK inhibitor and MEK inhibitor. This study clearly reveals that YIGSR peptide plays a novel function on the collagen type 1 synthesis of dermal fibroblasts and also suggests that YIGSR is a strong candidate peptide for the treatment of skin aging and wrinkles.

  16. STAT6-Dependent Collagen Synthesis in Human Fibroblasts Is Induced by Bovine Milk.

    PubMed

    Kippenberger, Stefan; Zöller, Nadja; Kleemann, Johannes; Müller, Jutta; Kaufmann, Roland; Hofmann, Matthias; Bernd, August; Meissner, Markus; Valesky, Eva

    2015-01-01

    Since the domestication of the urus, 10.000 years ago, mankind utilizes bovine milk for different purposes. Besides usage as a nutrient also the external application of milk on skin has a long tradition going back to at least the ancient Aegypt with Cleopatra VII as a great exponent. In order to test whether milk has impact on skin physiology, cultures of human skin fibroblasts were exposed to commercial bovine milk. Our data show significant induction of proliferation by milk (max. 2,3-fold, EC50: 2,5% milk) without toxic effects. Surprisingly, bovine milk was identified as strong inducer of collagen 1A1 synthesis at both, the protein (4-fold, EC50: 0,09% milk) and promoter level. Regarding the underlying molecular pathways, we show functional activation of STAT6 in a p44/42 and p38-dependent manner. More upstream, we identified IGF-1 and insulin as key factors responsible for milk-induced collagen synthesis. These findings show that bovine milk contains bioactive molecules that act on human skin cells. Therefore, it is tempting to test the herein introduced concept in treatment of atrophic skin conditions induced e.g. by UV light or corticosteroids.

  17. STAT6-Dependent Collagen Synthesis in Human Fibroblasts Is Induced by Bovine Milk

    PubMed Central

    Kippenberger, Stefan; Zöller, Nadja; Kleemann, Johannes; Müller, Jutta; Kaufmann, Roland; Hofmann, Matthias; Bernd, August; Meissner, Markus; Valesky, Eva

    2015-01-01

    Since the domestication of the urus, 10.000 years ago, mankind utilizes bovine milk for different purposes. Besides usage as a nutrient also the external application of milk on skin has a long tradition going back to at least the ancient Aegypt with Cleopatra VII as a great exponent. In order to test whether milk has impact on skin physiology, cultures of human skin fibroblasts were exposed to commercial bovine milk. Our data show significant induction of proliferation by milk (max. 2,3-fold, EC50: 2,5% milk) without toxic effects. Surprisingly, bovine milk was identified as strong inducer of collagen 1A1 synthesis at both, the protein (4-fold, EC50: 0,09% milk) and promoter level. Regarding the underlying molecular pathways, we show functional activation of STAT6 in a p44/42 and p38-dependent manner. More upstream, we identified IGF-1 and insulin as key factors responsible for milk-induced collagen synthesis. These findings show that bovine milk contains bioactive molecules that act on human skin cells. Therefore, it is tempting to test the herein introduced concept in treatment of atrophic skin conditions induced e.g. by UV light or corticosteroids. PMID:26134630

  18. STAT6-Dependent Collagen Synthesis in Human Fibroblasts Is Induced by Bovine Milk.

    PubMed

    Kippenberger, Stefan; Zöller, Nadja; Kleemann, Johannes; Müller, Jutta; Kaufmann, Roland; Hofmann, Matthias; Bernd, August; Meissner, Markus; Valesky, Eva

    2015-01-01

    Since the domestication of the urus, 10.000 years ago, mankind utilizes bovine milk for different purposes. Besides usage as a nutrient also the external application of milk on skin has a long tradition going back to at least the ancient Aegypt with Cleopatra VII as a great exponent. In order to test whether milk has impact on skin physiology, cultures of human skin fibroblasts were exposed to commercial bovine milk. Our data show significant induction of proliferation by milk (max. 2,3-fold, EC50: 2,5% milk) without toxic effects. Surprisingly, bovine milk was identified as strong inducer of collagen 1A1 synthesis at both, the protein (4-fold, EC50: 0,09% milk) and promoter level. Regarding the underlying molecular pathways, we show functional activation of STAT6 in a p44/42 and p38-dependent manner. More upstream, we identified IGF-1 and insulin as key factors responsible for milk-induced collagen synthesis. These findings show that bovine milk contains bioactive molecules that act on human skin cells. Therefore, it is tempting to test the herein introduced concept in treatment of atrophic skin conditions induced e.g. by UV light or corticosteroids. PMID:26134630

  19. The effects of an inhibitor of diglyceride lipase on collagen-induced platelet activation.

    PubMed

    Jackson, Elke C G; Ortar, Giorgio; McNicol, Archie

    2013-12-01

    Human platelet activation by collagen occurs in a dose-dependent manner. High concentrations of collagen bind to a pair of receptors, the α2β1 integrin and glycoprotein (GP)VI/Fc-receptor γ-chain (FcRγ), which stimulate a cascade of events including Syk, LAT, Btk, Gads, and phospholipase Cγ2, leading to calcium release and protein kinase C (PKC) activation. Calcium and PKC are responsible for a range of platelet responses including exocytosis and aggregation, as well as the cytosolic phospholipase A2 (cPLA2)-mediated release of arachidonic acid, which is converted to thromboxane (Tx)A2. In contrast, low concentrations of collagen are acutely aspirin-sensitive, and calcium release and aggregation are TxA2-dependent. Under these conditions, cPLA2 is not involved and it has been suggested that phospholipase C generates 1,2-diacylglycerol (DG) from which arachidonic acid is liberated by diglyceride lipase (DGL). Here a novel DGL blocker (OMDM-188) inhibited collagen-, but not arachidonic acid-induced aggregation and TxA2 synthesis. Furthermore, OMDM-188 inhibited collagen-induced arachidonic acid release. Finally OMDM-188 inhibited collagen-induced p38(MAPK) phosphorylation, but not extracellular signal-regulated kinase (ERK) phosphorylation, with no effect on the phosphorylation of either enzyme in response to arachidonic acid. Taken together, these data suggest a role for a pathway involving phospholipase C liberating DG from membrane phospholipids in response to minimally activating concentrations of collagen. The DG serves as a substrate for DGL, potentially under the regulations of p38(MAPK), to release arachidonic acid, which is subsequently converted to TxA2, which mediates the final platelet response.

  20. Tacrolimus (FK506) prevents early stages of ethanol induced hepatic fibrosis by targeting LARP6 dependent mechanism of collagen synthesis.

    PubMed

    Manojlovic, Zarko; Blackmon, John; Stefanovic, Branko

    2013-01-01

    Tacrolimus (FK506) is a widely used immunosuppressive drug. Its effects on hepatic fibrosis have been controversial and attributed to immunosuppression. We show that in vitro FK506, inhibited synthesis of type I collagen polypeptides, without affecting expression of collagen mRNAs. In vivo, administration of FK506 at a dose of 4 mg/kg completely prevented development of alcohol/carbon tetrachloride induced liver fibrosis in rats. Activation of hepatic stellate cells (HSCs) was absent in the FK506 treated livers and expression of collagen α2(I) mRNA was at normal levels. Collagen α1(I) mRNA was increased in the FK506 treated livers, but this mRNA was not translated into α1(I) polypeptide. No significant inflammation was associated with the fibrosis model used. FK506 binding protein 3 (FKBP3) is one of cellular proteins which binds FK506 with high affinity. We discovered that FKBP3 interacts with LARP6 and LARP6 is the major regulator of translation and stability of collagen mRNAs. In the presence of FK506 the interaction between FKBP3 and LARP6 is weakened and so is the pull down of collagen mRNAs with FKBP3. We postulate that FK506 inactivates FKBP3 and that lack of interaction of LARP6 and FKBP3 results in aberrant translation of collagen mRNAs and prevention of fibrosis. This is the first report of such activity of FK506 and may renew the interest in using this drug to alleviate hepatic fibrosis.

  1. Tacrolimus (FK506) Prevents Early Stages of Ethanol Induced Hepatic Fibrosis by Targeting LARP6 Dependent Mechanism of Collagen Synthesis

    PubMed Central

    Manojlovic, Zarko; Blackmon, John; Stefanovic, Branko

    2013-01-01

    Tacrolimus (FK506) is a widely used immunosuppressive drug. Its effects on hepatic fibrosis have been controversial and attributed to immunosuppression. We show that in vitro FK506, inhibited synthesis of type I collagen polypeptides, without affecting expression of collagen mRNAs. In vivo, administration of FK506 at a dose of 4 mg/kg completely prevented development of alcohol/carbon tetrachloride induced liver fibrosis in rats. Activation of hepatic stellate cells (HSCs) was absent in the FK506 treated livers and expression of collagen α2(I) mRNA was at normal levels. Collagen α1(I) mRNA was increased in the FK506 treated livers, but this mRNA was not translated into α1(I) polypeptide. No significant inflammation was associated with the fibrosis model used. FK506 binding protein 3 (FKBP3) is one of cellular proteins which binds FK506 with high affinity. We discovered that FKBP3 interacts with LARP6 and LARP6 is the major regulator of translation and stability of collagen mRNAs. In the presence of FK506 the interaction between FKBP3 and LARP6 is weakened and so is the pull down of collagen mRNAs with FKBP3. We postulate that FK506 inactivates FKBP3 and that lack of interaction of LARP6 and FKBP3 results in aberrant translation of collagen mRNAs and prevention of fibrosis. This is the first report of such activity of FK506 and may renew the interest in using this drug to alleviate hepatic fibrosis. PMID:23755290

  2. Stimulation of DNA and Collagen Synthesis by Autologous Growth Factor in Cultured Fetal Rat Calvaria

    NASA Astrophysics Data System (ADS)

    Canalis, Ernesto; Peck, William A.; Raisz, Lawrence G.

    1980-11-01

    Conditioned medium derived from organ or cell cultures prepared from 19- to 21-day fetal rat calvaria stimulated the incorporation of [3H]proline into collagen and of [3H]thymidine into DNA in organ cultures of the same tissue. Addition of cortisol enhanced the effect on collagen but not on DNA synthesis. These effects appeared to be due to a nondialyzable and heat-stable growth factor.

  3. Type I collagen-mediated synthesis of noble metallic nanoparticles networks and the applications in Surface-Enhanced Raman Scattering and electrochemistry.

    PubMed

    Sun, Yujing; Sun, Lanlan; Zhang, Baohua; Xu, Fugang; Liu, Zhelin; Guo, Cunlan; Zhang, Yue; Li, Zhuang

    2009-08-15

    In this paper, we demonstrated an effective environmentally friendly synthesis route to prepare noble metallic (Au, Ag, Pt and Pd) nanoparticles (NPs) networks mediated by type I collagen in the absence of any seeds or surfactants. In the reactions, type I collagen served as stabilizing agent and assembly template for the synthesized metallic NPs. The hydrophobic interaction between collagen and mica interface as well as the hydrogen bonds between inter- and intra-collagen molecules play important roles in the formation of collagen-metallic NPs networks. The noble metallic NPs networks have many advantages in the applications of Surface-Enhanced Raman Scattering (SERS) and electrochemistry detection. Typically, the as-prepared Ag NPs networks reveal great Raman enhancement activity for 4-ATP, and can even be used to detect low concentration of DNA base, adenine, without any label step. Furthermore, the cyclic voltammograms showed Pt NPs networks have good electrocatalytic ability for the reduction of O(2).

  4. Salamander-Derived, Human-Optimized nAG Protein Suppresses Collagen Synthesis and Increases Collagen Degradation in Primary Human Fibroblasts

    PubMed Central

    Al-Qattan, Mohammad M.; Shier, Medhat K.; Abd-AlWahed, Mervat M.; Mawlana, Ola H.; El-Wetidy, Mohammed S.; Bagayawa, Reginald S.; Ali, Hebatallah H.; Al-Nbaheen, May S.; Aldahmash, Abdullah M.

    2013-01-01

    Unlike humans, salamanders regrow their amputated limbs. Regeneration depends on the presence of regenerating axons which upregulate the expression of newt anterior gradient (nAG) protein. We had the hypothesis that nAG might have an inhibitory effect on collagen production since excessive collagen production results in scarring, which is a major enemy to regeneration. nAG gene was designed, synthesized, and cloned. The cloned vector was then transfected into primary human fibroblasts. The results showed that the expression of nAG protein in primary human fibroblast cells suppresses the expression of collagen I and III, with or without TGF-β1 stimulation. This suppression is due to a dual effect of nAG both by decreasing collagen synthesis and by increasing collagen degradation. Furthermore, nAG had an inhibitory effect on proliferation of transfected fibroblasts. It was concluded that nAG suppresses collagen through multiple effects. PMID:24288677

  5. Synthesis of the Peptide Ac-Wahx-KTTKS and Evaluation of the Ability to Induce In Vitro Collagen Synthesis.

    PubMed

    Guglielmi, Danila A S; Martinelli, Ariane M; Rissi, Nathália C; Cilli, Eduardo M; Soares, Christiane P; Chiavacci, Leila A

    2016-01-01

    In this work, we report the synthesis of a peptide analogue of the KTTKS, termed Ac- Wahx-KTTKS and evaluate its cytotoxicity and role in biosynthesis of collagen for future application in skin aging. The peptide was obtained with purity higher than 97.5%. In the cytotoxicity assay, we observed non-toxic effects for Ac-WAhx-KTTKS at concentrations below 600 µM for HaCaT and 500 µM for HepG2 cells, respectively. After 24 and 48 h it was possible to observe significant changes in collagen synthesis in the groups treated with various concentrations of the peptide. In conclusion, the Ac-Wahx- KTTKS peptide increased collagen synthesis in fibroblasts by 80% and it is a promising candidate for improving skin aging. PMID:27184584

  6. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    PubMed Central

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-01-01

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d3-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise. PMID:27367725

  7. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running.

    PubMed

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-01-01

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d₃-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise. PMID:27367725

  8. Collagen synthesis in rat gingiva during tooth movement

    SciTech Connect

    Boisson, M.; Gianelly, A.A.

    1981-09-01

    The response of the gingiva to an increased interdental space was studied by creating a diastema between the central incisors of rats and analyzing autoradiographically the incorporation of H3 proline in the gingiva to detect increased collagen production. In addition, conventional histologic methods were used to determine changes in the gingival architecture. The results indicate that the gingiva responds to an increased space in at least two ways. One is the production of more collagen fibers. The other involves the reorientation of the existing fibers in a horizontal plane as the gingival papilla becomes flattened.

  9. Characterization of binding of LARP6 to the 5’ stem-loop of collagen mRNAs: Implications for synthesis of type I collagen

    PubMed Central

    Stefanovic, Lela; Longo, Liam; Zhang, Yujie; Stefanovic, Branko

    2014-01-01

    Type I collagen is composed of 2 polypeptides, α1(I) and α2(I), which fold into triple helix. Collagen α1(I) and α2(I) mRNAs have a conserved stem-loop structure in their 5’ UTRs, the 5’SL. LARP6 binds the 5’SL to regulate type I collagen expression. We show that 5 nucleotides within the single stranded regions of 5’SL contribute to the high affinity of LARP6 binding. Mutation of individual nucleotides abolishes the binding in gel mobility shift assay. LARP6 binding to 5’SL of collagen α2(I) mRNA is more stable than the binding to 5’SL of α1(I) mRNA, although the equilibrium binding constants are similar. The more stable binding to α2(I) mRNA may favor synthesis of the heterotrimeric type I collagen. LARP6 needs 2 domains to contact 5’SL, the La domain and the RRM. T133 in the La domain is critical for folding of the protein, while loop 3 in the RRM is critical for binding 5’SL. Loop 3 is also involved in the interaction of LARP6 and protein translocation channel SEC61. This interaction is essential for type I collagen synthesis, because LARP6 mutant which binds 5’SL but which does not interact with SEC61, suppresses collagen synthesis in a dominant negative manner. We postulate that LARP6 directly targets collagen mRNAs to the SEC61 translocons to facilitate coordinated translation of the 2 collagen mRNAs. The unique sequences of LARP6 identified in this work may have evolved to enable its role in type I collagen biosynthesis. PMID:25692237

  10. Asiaticoside induces type I collagen synthesis and osteogenic differentiation in human periodontal ligament cells.

    PubMed

    Nowwarote, Nunthawan; Osathanon, Thanaphum; Jitjaturunt, Peachaya; Manopattanasoontorn, Sukuman; Pavasant, Prasit

    2013-03-01

    Asiaticoside, an active ingredient extracted from Centella asiatica, has been widely used to promote wound healing. In this study, the effects of asiaticoside on proliferation, protein synthesis, and osteogenic differentiation in human periodontal ligament cells (HPDLs) were investigated. HPDLs were treated with asiaticoside at concentrations of 25, 50, and 100 µg/mL. Cell number was determined by MTT assay. The mRNA expression was analyzed by reverse transcription-polymerase chain reaction. Western blot analysis and immunocytochemistry were used to confirm protein synthesis. Osteogenic differentiation was determined by alkaline phosphatase activity, osteoblast marker gene expression, and in vitro mineralization. The results showed that asiaticoside treatment, ranging from 25 to 100 mg/mL, had no effect on cytotoxicity or cell proliferation. When HPDLs were treated with asiaticoside in serum-free medium, dose-dependent increases in the levels of fibronectin and collagen type I mRNA and protein were observed at 72 h. Moreover, asiaticoside attenuated matrix metalloproteinase-1 but enhanced tissue inhibitor of metalloproteinase-1 mRNA expression. The addition of asiaticoside to osteogenic medium resulted in an increase in alkaline phosphatase enzymatic activity, up-regulation of osteoblast marker gene mRNA expression, and enhancement of mineralization by HPDLs. These results suggest the potential application of asiaticoside for enhancing periodontal tissue healing.

  11. Ultrananocrystalline diamond thin films functionalized with therapeutically active collagen networks.

    SciTech Connect

    Huang, H.; Chen, M.; Bruno, P.; Lam, R.; Robinson, E.; Gruen, D.; Ho, D.; Materials Science Division; Northwestern Univ.

    2009-01-01

    The fabrication of biologically amenable interfaces in medicine bridges translational technologies with their surrounding biological environment. Functionalized nanomaterials catalyze this coalescence through the creation of biomimetic and active substrates upon which a spectrum of therapeutic elements can be delivered to adherent cells to address biomolecular processes in cancer, inflammation, etc. Here, we demonstrate the robust functionalization of ultrananocrystalline diamond (UNCD) with type I collagen and dexamethasone (Dex), an anti-inflammatory drug, to fabricate a hybrid therapeutically active substrate for localized drug delivery. UNCD oxidation coupled with a pH-mediated collagen adsorption process generated a comprehensive interface between the two materials, and subsequent Dex integration, activity, and elution were confirmed through inflammatory gene expression assays. These studies confer a translational relevance to the biofunctionalized UNCD in its role as an active therapeutic network for potent regulation of cellular activity toward applications in nanomedicine.

  12. Hydrogen peroxide is involved in collagen-induced platelet activation.

    PubMed

    Pignatelli, P; Pulcinelli, F M; Lenti, L; Gazzaniga, P P; Violi, F

    1998-01-15

    In this study, we investigated whether (1) collagen-induced platelet aggregation is associated with a burst of H2O2, (2) this oxidant species is involved in the activation of platelets, and (3) the pathways of platelet activation are stimulated by H2O2. Collagen-induced platelet aggregation was associated with production of H2O2, which was abolished by catalase, an enzyme that destroys H2O2. H2O2 production was not observed when ADP or thrombin were used as agonists. Catalase inhibited dose-dependently thromboxane A2 production, release of arachidonic acid from platelet membrane, and Inositol 1,4,5P3 (IP3) formation. In aspirin-treated platelets stimulated with high concentrations of collagen, catalase inhibited platelet aggregation, calcium mobilization, and IP3 production. This study suggests that collagen-induced platelet aggregation is associated with a burst of H2O2 that acts as a second messenger by stimulating the arachidonic acid metabolism and phospholipase C pathway.

  13. Uniform spatial distribution of collagen fibril radii within tendon implies local activation of pC-collagen at individual fibrils

    NASA Astrophysics Data System (ADS)

    Rutenberg, Andrew D.; Brown, Aidan I.; Kreplak, Laurent

    2016-08-01

    Collagen fibril cross-sectional radii show no systematic variation between the interior and the periphery of fibril bundles, indicating an effectively constant rate of collagen incorporation into fibrils throughout the bundle. Such spatially homogeneous incorporation constrains the extracellular diffusion of collagen precursors from sources at the bundle boundary to sinks at the growing fibrils. With a coarse-grained diffusion equation we determine stringent bounds, using parameters extracted from published experimental measurements of tendon development. From the lack of new fibril formation after birth, we further require that the concentration of diffusing precursors stays below the critical concentration for fibril nucleation. We find that the combination of the diffusive bound, which requires larger concentrations to ensure homogeneous fibril radii, and lack of nucleation, which requires lower concentrations, is only marginally consistent with fully processed collagen using conservative bounds. More realistic bounds may leave no consistent concentrations. Therefore, we propose that unprocessed pC-collagen diffuses from the bundle periphery followed by local C-proteinase activity and subsequent collagen incorporation at each fibril. We suggest that C-proteinase is localized within bundles, at fibril surfaces, during radial fibrillar growth. The much greater critical concentration of pC-collagen, as compared to fully processed collagen, then provides broad consistency between homogeneous fibril radii and the lack of fibril nucleation during fibril growth.

  14. The contribution of increased collagen synthesis to human glomerulosclerosis: a quantitative analysis of alpha 2IV collagen mRNA expression by competitive polymerase chain reaction

    PubMed Central

    1992-01-01

    We previously reported that one of the main components of the sclerotic material in human glomerular diseases was type IV collagen. In this study we examined the contribution of increased synthesis to this process at the gene expression level. Sufficient material has not been available to study type IV collagen synthesis by normal or sclerotic glomeruli in humans. We took advantage of the availability of nephrectomy specimens from patients with renal carcinoma, and of the observation that approximately 50% of these patients develop varying degrees of glomerulosclerosis. We microdissected glomeruli from 10 patients and analyzed them using in situ reverse transcription coupled with polymerase chain reaction (PCR) analyses (in situ RT-PCR). alpha 2IV collagen mRNA, after reverse transcription into cDNA, was detected in all patients and appeared to be increased in those with glomerulosclerosis (n = 5). A competitive PCR assay was developed to quantitate this change. There was an average 3.7-fold increase in glomerular type IV collagen cDNA in patients with significant sclerosis. This change was not due to an increased number of glomerular cells. Thus, glomerulosclerosis in humans is associated with an elevation of glomerular type IV collagen gene expression, suggesting that increased synthesis of type IV collagen may represent one component of this process. PMID:1281210

  15. Mechanical stretching stimulates collagen synthesis via down-regulating SO2/AAT1 pathway.

    PubMed

    Liu, Jia; Yu, Wen; Liu, Yan; Chen, Selena; Huang, Yaqian; Li, Xiaohui; Liu, Cuiping; Zhang, Yanqiu; Li, Zhenzhen; Du, Jie; Tang, Chaoshu; Du, Junbao; Jin, Hongfang

    2016-02-16

    The aim of the study was to investigate the role of endogenous sulfur dioxide (SO2)/ aspartate aminotransferase 1 (AAT1) pathway in stretch-induced excessive collagen expression and its mechanism. The mechanical stretch downregulated SO2/AAT1 pathway and increased collagen I and III protein expression. Importantly, AAT1 overexpression blocked the increase in collagen I and III expression, transforming growth factor-β1 (TGF- β1) expression and phosphorylation of Smad2/3 induced by stretch, but AAT1 knockdown mimicked the increase in collagen I and III expression, TGF- β1 expression and phosphorylation of Smad2/3 induced by stretch. Mechanistically, SB431542, a TGF-β1/Smad2/3 inhibitor, eliminated excessive collagen I and III accumulation induced by AAT1 knockdown, stretch or stretch plus AAT1 knockdown. In a rat model of high pulmonary blood flow-induced pulmonary vascular collagen accumulation, AAT1 expression and SO2 content in lung tissues of rat were reduced in shunt rats with high pulmonary blood flow. Supplement of SO2 derivatives inhibited activation of TGF- β1/Smad2/3 pathway and alleviated the excessive collagen accumulation in lung tissues of shunt rats. The results suggested that deficiency of endogenous SO2/AAT1 pathway mediated mechanical stretch-stimulated abnormal collagen accumulation via TGF-β1/Smad2/3 pathway.

  16. Mechanical stretching stimulates collagen synthesis via down-regulating SO2/AAT1 pathway

    PubMed Central

    Liu, Jia; Yu, Wen; Liu, Yan; Chen, Selena; Huang, Yaqian; Li, Xiaohui; Liu, Cuiping; Zhang, Yanqiu; Li, Zhenzhen; Du, Jie; Tang, Chaoshu; Du, Junbao; Jin, Hongfang

    2016-01-01

    The aim of the study was to investigate the role of endogenous sulfur dioxide (SO2)/ aspartate aminotransferase 1 (AAT1) pathway in stretch-induced excessive collagen expression and its mechanism. The mechanical stretch downregulated SO2/AAT1 pathway and increased collagen I and III protein expression. Importantly, AAT1 overexpression blocked the increase in collagen I and III expression, transforming growth factor-β1 (TGF- β1) expression and phosphorylation of Smad2/3 induced by stretch, but AAT1 knockdown mimicked the increase in collagen I and III expression, TGF- β1 expression and phosphorylation of Smad2/3 induced by stretch. Mechanistically, SB431542, a TGF-β1/Smad2/3 inhibitor, eliminated excessive collagen I and III accumulation induced by AAT1 knockdown, stretch or stretch plus AAT1 knockdown. In a rat model of high pulmonary blood flow-induced pulmonary vascular collagen accumulation, AAT1 expression and SO2 content in lung tissues of rat were reduced in shunt rats with high pulmonary blood flow. Supplement of SO2 derivatives inhibited activation of TGF- β1/Smad2/3 pathway and alleviated the excessive collagen accumulation in lung tissues of shunt rats. The results suggested that deficiency of endogenous SO2/AAT1 pathway mediated mechanical stretch-stimulated abnormal collagen accumulation via TGF-β1/Smad2/3 pathway. PMID:26880260

  17. Ethinyl oestradiol administration in women suppresses synthesis of collagen in tendon in response to exercise

    PubMed Central

    Hansen, Mette; Koskinen, Satu O; Petersen, Susanne G; Doessing, Simon; Frystyk, Jan; Flyvbjerg, Allan; Westh, Eva; Magnusson, S Peter; Kjaer, Michael; Langberg, Henning

    2008-01-01

    Women are at greater risk than men of sustaining certain kinds of injury and diseases of collagen-rich tissues. To determine whether a high level of oestradiol has an acute influence on collagen synthesis in tendons at rest and in response to exercise, one-legged kicking exercise was performed for 60 min at 67% of maximum power by healthy, young oral contraceptive (OC) users when circulating synthetic (ethinyl) oestradiol was high (n = 11, HE-OC) and compared to similar women who had never used OCs when circulating endogenous oestrogen was low (n = 12, LE-NOC). Interstitial fluid was collected 24 h post-exercise through microdialysis catheters placed anterior to the patellar tendon in both legs and subsequently analysed for the amino-terminal propeptide of type I collagen (PINP), a marker of tendon collagen synthesis. To determine the long-term effect of OC usage, patellar tendon cross-sectional area (CSA) was measured by magnetic resonance imaging (MRI). A lower exercise-induced increase in tendon collagen synthesis was observed in HE-OC than in LE-NOC (ΔPINP (mean ±s.e.m.) 1.5 ± 5.3 versus 24.2 ± 9.4 ng ml−1, P < 0.05). Furthermore, serum and the interstitial peritendinous tissue concentrations of insulin-like growth factor I (IGF-I) and IGF-binding proteins showed a reduced bioavailability in HE-OC compared with results in LE-NOC. No difference in patellar tendon CSA was observed between groups. In conclusion, the selective increase in tendon collagen synthesis in LE-NOC but not HE-OC 24 h post-exercise is consistent with the hypothesis that oestradiol inhibits exercise-induced collagen synthesis in human tendon. The mechanism behind this is either a direct effect of oestradiol, or an indirect effect via a reduction in levels of free IGF-I. However, the data did not indicate any long-term effect on tendon size associated with chronic OC use. PMID:18420709

  18. Fibromodulin Interacts with Collagen Cross-linking Sites and Activates Lysyl Oxidase*

    PubMed Central

    Bihan, Dominique; Bonna, Arkadiusz; Rubin, Kristofer; Farndale, Richard W.

    2016-01-01

    The hallmark of fibrotic disorders is a highly cross-linked and dense collagen matrix, a property driven by the oxidative action of lysyl oxidase. Other fibrosis-associated proteins also contribute to the final collagen matrix properties, one of which is fibromodulin. Its interactions with collagen affect collagen cross-linking, packing, and fibril diameter. We investigated the possibility that a specific relationship exists between fibromodulin and lysyl oxidase, potentially imparting a specific collagen matrix phenotype. We mapped the fibromodulin-collagen interaction sites using the collagen II and III Toolkit peptide libraries. Fibromodulin interacted with the peptides containing the known collagen cross-linking sites and the MMP-1 cleavage site in collagens I and II. Interestingly, the interaction sites are closely aligned within the quarter-staggered collagen fibril, suggesting a multivalent interaction between fibromodulin and several collagen helices. Furthermore, we detected an interaction between fibromodulin and lysyl oxidase (a major collagen cross-linking enzyme) and mapped the interaction site to 12 N-terminal amino acids on fibromodulin. This interaction also increases the activity of lysyl oxidase. Together, the data suggest a fibromodulin-modulated collagen cross-linking mechanism where fibromodulin binds to a specific part of the collagen domain and also forms a complex with lysyl oxidase, targeting the enzyme toward specific cross-linking sites. PMID:26893379

  19. Fibromodulin Interacts with Collagen Cross-linking Sites and Activates Lysyl Oxidase.

    PubMed

    Kalamajski, Sebastian; Bihan, Dominique; Bonna, Arkadiusz; Rubin, Kristofer; Farndale, Richard W

    2016-04-01

    The hallmark of fibrotic disorders is a highly cross-linked and dense collagen matrix, a property driven by the oxidative action of lysyl oxidase. Other fibrosis-associated proteins also contribute to the final collagen matrix properties, one of which is fibromodulin. Its interactions with collagen affect collagen cross-linking, packing, and fibril diameter. We investigated the possibility that a specific relationship exists between fibromodulin and lysyl oxidase, potentially imparting a specific collagen matrix phenotype. We mapped the fibromodulin-collagen interaction sites using the collagen II and III Toolkit peptide libraries. Fibromodulin interacted with the peptides containing the known collagen cross-linking sites and the MMP-1 cleavage site in collagens I and II. Interestingly, the interaction sites are closely aligned within the quarter-staggered collagen fibril, suggesting a multivalent interaction between fibromodulin and several collagen helices. Furthermore, we detected an interaction between fibromodulin and lysyl oxidase (a major collagen cross-linking enzyme) and mapped the interaction site to 12 N-terminal amino acids on fibromodulin. This interaction also increases the activity of lysyl oxidase. Together, the data suggest a fibromodulin-modulated collagen cross-linking mechanism where fibromodulin binds to a specific part of the collagen domain and also forms a complex with lysyl oxidase, targeting the enzyme toward specific cross-linking sites.

  20. Synthesis and degradation rates of collagens in vivo in whole skin of rats, studied with 1802 labelling.

    PubMed Central

    Molnar, J A; Alpert, N; Burke, J F; Young, V R

    1986-01-01

    Rats of synthesis and degradation in vivo of collagens in 0.5 M-acetic acid-soluble and -insoluble extracts from skins of three growing rats were determined by using a labelling procedure involving exposure of the animals to an atmosphere of 18O2 for 36 h. For comparison, rats also received injections of [2H]proline. Serial skin biopsies were taken at frequent intervals over 392 days. Enrichment of 18O and 2H in the hydroxyproline of the collagen fractions was determined by gas chromatography-mass spectrometry. Changes in size of the soluble and insoluble collagen pools were considered in the evaluation of isotope kinetic data. The insoluble collagen fraction showed no degradation. The efflux (mean +/- S.D., expressed as mumol of hydroxyproline) from the soluble collagen pool was estimated to be 59.9 +/- 1.9 per day from the 18O data, and 25.5 +/- 7.5 per day from the 2H results. The finding indicates significant reutilization of 2H-radiolabelled proline for hydroxyproline synthesis. From these isotope data and estimates of size of the collagen pools it was determined that 55% of the collagen disappearing from the soluble pool was due to maturation into insoluble collagens and 45% of the disappearance was a result of actual degradation of soluble collagen. These results confirm the utility of 18O2 as a non-reutilizable label for studies of collagen turnover in vivo. PMID:3814092

  1. High voltage pulsed current in collagen realignment, synthesis, and angiogenesis after Achilles tendon partial rupture

    PubMed Central

    Rampazo, Érika P.; Liebano, Richard E.; Pinfildi, Carlos Eduardo; Folha, Roberta A. C.; Ferreira, Lydia M.

    2016-01-01

    ABSTRACT Objective To verify the efficacy of high voltage pulsed current in collagen realignment and synthesis and in angiogenesis after the partial rupturing of the Achilles tendon in rats. Method Forty male Wistar rats were randomized into four groups of 10 animals each: sham, cathodic stimulation, anodic stimulation, and alternating stimulation. Their Achilles tendons were submitted to direct trauma by a free-falling metal bar. Then, the treatment was administered for six consecutive days after the injury. In the simulation group, the electrodes were positioned on the animal, but the device remained off for 30 minutes. The other groups used a frequency of 120 pps, sensory threshold, and the corresponding polarity. On the seventh day, the tendons were removed and sent for histological slide preparation for birefringence and Picrosirius Red analysis and for blood vessel quantification. Results No significant difference was observed among the groups regarding collagen realignment (types I or III collagen) or quantity of blood vessels. Conclusion High voltage pulsed current for six consecutive days was not effective in collagen realignment, synthesis, or angiogenesis after the partial rupturing of the Achilles tendon in rats. PMID:27556387

  2. Use of Cis-[18F]Fluoro-Proline for Assessment of Exercise-Related Collagen Synthesis in Musculoskeletal Connective Tissue

    PubMed Central

    Skovgaard, Dorthe; Kjaer, Andreas; Heinemeier, Katja Maria; Brandt-Larsen, Malene; Madsen, Jacob; Kjaer, Michael

    2011-01-01

    Protein turnover in collagen rich tissue is influenced by exercise, but can only with difficulty be studied in vivo due to use of invasive procedure. The present study was done to investigate the possibility of applying the PET-tracer, cis-[18F]fluoro-proline (cis-Fpro), for non-invasive assessment of collagen synthesis in rat musculoskeletal tissues at rest and following short-term (3 days) treadmill running. Musculoskeletal collagen synthesis was studied in rats at rest and 24 h post-exercise. At each session, rats were PET scanned at two time points following injection of cis-FPro: (60 and 240 min p.i). SUV were calculated for Achilles tendon, calf muscle and tibial bone. The PET-derived results were compared to mRNA expression of collagen type I and III. Tibial bone had the highest SUV that increased significantly (p<0.001) from the early (60 min) to the late (240 min) PET scan, while SUV in tendon and muscle decreased (p<0.001). Exercise had no influence on SUV, which was contradicted by an increased gene expression of collagen type I and III in muscle and tendon. The clearly, visible uptake of cis-Fpro in the collagen-rich musculoskeletal tissues is promising for multi-tissue studies in vivo. The tissue-specific differences with the highest basal uptake in bone are in accordance with earlier studies relying on tissue incorporation of isotopic-labelled proline. A possible explanation of the failure to demonstrate enhanced collagen synthesis following exercise, despite augmented collagen type I and III transcription, is that SUV calculations are not sensitive enough to detect minor changes in collagen synthesis. Further studies including kinetic compartment modeling must be performed to establish whether cis-Fpro can be used for non-invasive in-vivo assessment of exercise-induced changes in musculoskeletal collagen synthesis. PMID:21347251

  3. The synthesis and coupling of photoreactive collagen-based peptides to restore integrin reactivity to an inert substrate, chemically-crosslinked collagen.

    PubMed

    Malcor, Jean-Daniel; Bax, Daniel; Hamaia, Samir W; Davidenko, Natalia; Best, Serena M; Cameron, Ruth E; Farndale, Richard W; Bihan, Dominique

    2016-04-01

    Collagen is frequently advocated as a scaffold for use in regenerative medicine. Increasing the mechanical stability of a collagen scaffold is widely achieved by cross-linking using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). However, this treatment consumes the carboxylate-containing amino acid sidechains that are crucial for recognition by the cell-surface integrins, abolishing cell adhesion. Here, we restore cell reactivity to a cross-linked type I collagen film by covalently linking synthetic triple-helical peptides (THPs), mimicking the structure of collagen. These THPs are ligands containing an active cell-recognition motif, GFOGER, a high-affinity binding site for the collagen-binding integrins. We end-stapled peptide strands containing GFOGER by coupling a short diglutamate-containing peptide to their N-terminus, improving the thermal stability of the resulting THP. A photoreactive Diazirine group was grafted onto the end-stapled THP to allow covalent linkage to the collagen film upon UV activation. Such GFOGER-derivatized collagen films showed restored affinity for the ligand-binding I domain of integrin α2β1, and increased integrin-dependent cell attachment and spreading of HT1080 and Rugli cell lines, expressing integrins α2β1 and α1β1, respectively. The method we describe has wide application, beyond collagen films or scaffolds, since the photoreactive diazirine will react with many organic carbon skeletons.

  4. The synthesis and coupling of photoreactive collagen-based peptides to restore integrin reactivity to an inert substrate, chemically-crosslinked collagen.

    PubMed

    Malcor, Jean-Daniel; Bax, Daniel; Hamaia, Samir W; Davidenko, Natalia; Best, Serena M; Cameron, Ruth E; Farndale, Richard W; Bihan, Dominique

    2016-04-01

    Collagen is frequently advocated as a scaffold for use in regenerative medicine. Increasing the mechanical stability of a collagen scaffold is widely achieved by cross-linking using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). However, this treatment consumes the carboxylate-containing amino acid sidechains that are crucial for recognition by the cell-surface integrins, abolishing cell adhesion. Here, we restore cell reactivity to a cross-linked type I collagen film by covalently linking synthetic triple-helical peptides (THPs), mimicking the structure of collagen. These THPs are ligands containing an active cell-recognition motif, GFOGER, a high-affinity binding site for the collagen-binding integrins. We end-stapled peptide strands containing GFOGER by coupling a short diglutamate-containing peptide to their N-terminus, improving the thermal stability of the resulting THP. A photoreactive Diazirine group was grafted onto the end-stapled THP to allow covalent linkage to the collagen film upon UV activation. Such GFOGER-derivatized collagen films showed restored affinity for the ligand-binding I domain of integrin α2β1, and increased integrin-dependent cell attachment and spreading of HT1080 and Rugli cell lines, expressing integrins α2β1 and α1β1, respectively. The method we describe has wide application, beyond collagen films or scaffolds, since the photoreactive diazirine will react with many organic carbon skeletons. PMID:26854392

  5. The synthesis and coupling of photoreactive collagen-based peptides to restore integrin reactivity to an inert substrate, chemically-crosslinked collagen

    PubMed Central

    Malcor, Jean-Daniel; Bax, Daniel; Hamaia, Samir W.; Davidenko, Natalia; Best, Serena M.; Cameron, Ruth E.; Farndale, Richard W.; Bihan, Dominique

    2016-01-01

    Collagen is frequently advocated as a scaffold for use in regenerative medicine. Increasing the mechanical stability of a collagen scaffold is widely achieved by cross-linking using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). However, this treatment consumes the carboxylate-containing amino acid sidechains that are crucial for recognition by the cell-surface integrins, abolishing cell adhesion. Here, we restore cell reactivity to a cross-linked type I collagen film by covalently linking synthetic triple-helical peptides (THPs), mimicking the structure of collagen. These THPs are ligands containing an active cell-recognition motif, GFOGER, a high-affinity binding site for the collagen-binding integrins. We end-stapled peptide strands containing GFOGER by coupling a short diglutamate-containing peptide to their N-terminus, improving the thermal stability of the resulting THP. A photoreactive Diazirine group was grafted onto the end-stapled THP to allow covalent linkage to the collagen film upon UV activation. Such GFOGER-derivatized collagen films showed restored affinity for the ligand-binding I domain of integrin α2β1, and increased integrin-dependent cell attachment and spreading of HT1080 and Rugli cell lines, expressing integrins α2β1 and α1β1, respectively. The method we describe has wide application, beyond collagen films or scaffolds, since the photoreactive diazirine will react with many organic carbon skeletons. PMID:26854392

  6. The effect of oxygen partial pressure on protein synthesis and collagen hydroxylation by mature periodontal tissues maintained in organ cultures

    PubMed Central

    Yen, Edwin H. K.; Sodek, Jaro; Melcher, Antony H.

    1979-01-01

    Mature periodontal tissues from adult-mouse first mandibular molars were cultured in a continuous-flow organ-culture system which allowed the regulation of both ascorbic acid concentration and pO2 (oxygen partial pressure). Protein synthesis was measured by analysing the incorporation of [3H]proline into collagenous and non-collagenous proteins during the last 24h of a 2-day culture. At low pO2 [16.0kPa (approx. 120mmHg)] approx. 60% of protein-incorporated [3H]proline was found in collagenous proteins. However, it was evident that this collagen was considerably underhydroxylated. At high pO2 [56.0kPa (approx. 420mmHg)], both the amount of collagen deposited in the tissues and the degree of hydroxylation were increased considerably. In contrast, no significant effect on non-collagenous protein was observed. Tissues cultured at low pO2 for the first 48h were unable to respond to a subsequent increase in pO2 during the last 24h. Analysis of pepsin-solubilized collagen α-chains labelled with [14C]glycine demonstrated the synthesis of both type-I and type-III collagens by explants cultured for 48h at high pO2. Type-III collagen comprised 20–30% of the radioactivity in α-chains in both the periodontal ligament and the tissues of the alveolar process. The pattern of protein synthesis in the alveolar tissues at high pO2 was similar to that observed in these tissues in vivo. However, in the cultured periodontal ligament the proportions of non-collagenous proteins and type-III collagens were increased in comparison with the tissue in vivo. PMID:454369

  7. Synthesis of type I collagen in healing wounds in humans.

    PubMed Central

    Haukipuro, K; Melkko, J; Risteli, L; Kairaluoma, M; Risteli, J

    1991-01-01

    To quantify wound healing in surgical patients, samples of wound fluid were collected through a silicone rubber tube for 7 postoperative days and their concentrations of the carboxyterminal propeptide of type I procollagen (PICP) and the aminoterminal propeptide of type III procollagen (PIIINP) were measured with specific radioimmunoassays. The mean concentration of PICP in would fluid on day 1 was 207 +/- 92 (SD) micrograms/L, and on day 2 908 +/- 469 micrograms/L (p less than 0.001, signed rank test). On day 7, the mean concentration reached was 380 times higher than that of day 1 (79,330 +/- 54,151 micrograms/L). Only one peak of PICP antigenicity, corresponding to the intact propeptide as set free during synthesis of type I procollagen, was detected on Sephacryl S-300 gel filtration analysis of wound fluid samples. The mean concentration of PIIINP was 70 +/- 61 micrograms/L on day 1, 86 +/- 88 micrograms/L on day 2, and 180 +/- 129 micrograms/L on day 3 (p less than 0.001 when compared with day 1). Finally on day 7, a 250-fold concentration (17,812 +/- 9839 micrograms/L), compared with day 1, was reached. Methods described in the present paper allow separate and repetitive quantification of the synthesis of both type I and type III procollagen during human wound healing. PMID:1985542

  8. Synthesis, characterization and bioevaluation of drug-collagen hybrid materials for biomedical applications.

    PubMed

    Voicu, Georgeta; Geanaliu-Nicolae, Ruxandra-Elena; Pîrvan, Adrian-Alexandru; Andronescu, Ecaterina; Iordache, Florin

    2016-08-30

    This work presents a study based on the preparation and characterization of drug-collagen hybrid materials. Materials used for obtaining drug-collagen hybrids were collagen type I (Coll) as matrix and fludarabine (F) and epirubicin (E) as hydrophilic active substances. After incorporation of drugs into Coll in different ratios, the obtained hybrid materials (Coll/F and Coll/E) could be used according to our results as potential drug delivery systems in medicine for the topical (local) treatment of cancerous tissues (e.g. the treatment of breast, stomach, lung, colorectal or advanced ovarian cancer). The materials were characterized considering their composition (by XRD, FT-IR and DTA-TG) and their morphology (by SEM). The delivery of drug was assessed by UV-vis. The in vitro citotoxicity demonstrates an antitumoral activity of the obtained hybrid materials and their potential use for biomedical applications as drug delivery systems in tumoral treatments.

  9. Comparative effects of biodynes, tocotrienol-rich fraction, and tocopherol in enhancing collagen synthesis and inhibiting collagen degradation in stress-induced premature senescence model of human diploid fibroblasts.

    PubMed

    Makpol, Suzana; Jam, Faidruz Azura; Khor, Shy Cian; Ismail, Zahariah; Mohd Yusof, Yasmin Anum; Ngah, Wan Zurinah Wan

    2013-01-01

    Biodynes, tocotrienol-rich fraction (TRF), and tocopherol have shown antiaging properties. However, the combined effects of these compounds on skin aging are yet to be investigated. This study aimed to elucidate the skin aging effects of biodynes, TRF, and tocopherol on stress-induced premature senescence (SIPS) model of human diploid fibroblasts (HDFs) by determining the expression of collagen and MMPs at gene and protein levels. Primary HDFs were treated with biodynes, TRF, and tocopherol prior to hydrogen peroxide (H2O2) exposure. The expression of COL1A1, COL3A1, MMP1, MMP2, MMP3, and MMP9 genes was determined by qRT-PCR. Type I and type III procollagen proteins were measured by Western blotting while the activities of MMPs were quantified by fluorometric Sensolyte MMP Kit. Our results showed that biodynes, TRF, and tocopherol upregulated collagen genes and downregulated MMP genes (P < 0.05). Type I procollagen and type III procollagen protein levels were significantly increased in response to biodynes, TRF, and tocopherol treatment (P < 0.05) with reduction in MMP-1, MMP-2, MMP-3, and MMP-9 activities (P < 0.05). These findings indicated that biodynes, TRF, and tocopherol effectively enhanced collagen synthesis and inhibited collagen degradation and therefore may protect the skin from aging.

  10. Comparative effects of biodynes, tocotrienol-rich fraction, and tocopherol in enhancing collagen synthesis and inhibiting collagen degradation in stress-induced premature senescence model of human diploid fibroblasts.

    PubMed

    Makpol, Suzana; Jam, Faidruz Azura; Khor, Shy Cian; Ismail, Zahariah; Mohd Yusof, Yasmin Anum; Ngah, Wan Zurinah Wan

    2013-01-01

    Biodynes, tocotrienol-rich fraction (TRF), and tocopherol have shown antiaging properties. However, the combined effects of these compounds on skin aging are yet to be investigated. This study aimed to elucidate the skin aging effects of biodynes, TRF, and tocopherol on stress-induced premature senescence (SIPS) model of human diploid fibroblasts (HDFs) by determining the expression of collagen and MMPs at gene and protein levels. Primary HDFs were treated with biodynes, TRF, and tocopherol prior to hydrogen peroxide (H2O2) exposure. The expression of COL1A1, COL3A1, MMP1, MMP2, MMP3, and MMP9 genes was determined by qRT-PCR. Type I and type III procollagen proteins were measured by Western blotting while the activities of MMPs were quantified by fluorometric Sensolyte MMP Kit. Our results showed that biodynes, TRF, and tocopherol upregulated collagen genes and downregulated MMP genes (P < 0.05). Type I procollagen and type III procollagen protein levels were significantly increased in response to biodynes, TRF, and tocopherol treatment (P < 0.05) with reduction in MMP-1, MMP-2, MMP-3, and MMP-9 activities (P < 0.05). These findings indicated that biodynes, TRF, and tocopherol effectively enhanced collagen synthesis and inhibited collagen degradation and therefore may protect the skin from aging. PMID:24396567

  11. Comparative Effects of Biodynes, Tocotrienol-Rich Fraction, and Tocopherol in Enhancing Collagen Synthesis and Inhibiting Collagen Degradation in Stress-Induced Premature Senescence Model of Human Diploid Fibroblasts

    PubMed Central

    Jam, Faidruz Azura; Ismail, Zahariah; Wan Ngah, Wan Zurinah

    2013-01-01

    Biodynes, tocotrienol-rich fraction (TRF), and tocopherol have shown antiaging properties. However, the combined effects of these compounds on skin aging are yet to be investigated. This study aimed to elucidate the skin aging effects of biodynes, TRF, and tocopherol on stress-induced premature senescence (SIPS) model of human diploid fibroblasts (HDFs) by determining the expression of collagen and MMPs at gene and protein levels. Primary HDFs were treated with biodynes, TRF, and tocopherol prior to hydrogen peroxide (H2O2) exposure. The expression of COL1A1, COL3A1, MMP1, MMP2, MMP3, and MMP9 genes was determined by qRT-PCR. Type I and type III procollagen proteins were measured by Western blotting while the activities of MMPs were quantified by fluorometric Sensolyte MMP Kit. Our results showed that biodynes, TRF, and tocopherol upregulated collagen genes and downregulated MMP genes (P < 0.05). Type I procollagen and type III procollagen protein levels were significantly increased in response to biodynes, TRF, and tocopherol treatment (P < 0.05) with reduction in MMP-1, MMP-2, MMP-3, and MMP-9 activities (P < 0.05). These findings indicated that biodynes, TRF, and tocopherol effectively enhanced collagen synthesis and inhibited collagen degradation and therefore may protect the skin from aging. PMID:24396567

  12. Resveratrol causes cell cycle arrest, decreased collagen synthesis, and apoptosis in rat intestinal smooth muscle cells.

    PubMed

    Garcia, Patricia; Schmiedlin-Ren, Phyllissa; Mathias, Jason S; Tang, Huaijing; Christman, Gregory M; Zimmermann, Ellen M

    2012-02-01

    One of the most difficult and treatment-resistant complications of Crohn's disease is the development of fibrotic intestinal strictures due to mesenchymal cell hyperplasia and collagen deposition. Resveratrol, a phytoalexin found in berries, peanuts, grapes, and red wine, has been shown to inhibit fibrosis in vasculature, heart, lung, kidney, liver, and esophagus in animal models. Resveratrol has also been shown to inhibit oxidation, inflammation, and cell proliferation and to decrease collagen synthesis in several cell types or animal models. The aim of this study was to determine whether resveratrol has antifibrotic effects on intestinal smooth muscle cells. Responses to resveratrol by cultured smooth muscle cells isolated from colons of untreated Lewis rats were examined; this rat strain is used in a model of Crohn's disease with prominent intestinal fibrosis. A relative decrease in cell numbers following treatment with 50 and 100 μM resveratrol was evident at 24 h (P ≤ 0.005). This effect was largely due to cell cycle arrest, with an increase in the percent of cells in S phase from 8 to 25-35% (P < 0.05). Cell viability was unchanged until 2-3 days of treatment when there was a 1.2- to 5.0-fold increase in the percent of apoptotic cells, depending on the assay (P < 0.05). Expression of collagen type I protein was decreased following treatment with resveratrol for 24 h (to 44 and 25% of control levels with 50 and 100 μM resveratrol, respectively; P < 0.05). Expression of procollagen types I and III mRNA was also decreased with resveratrol treatment. Resveratrol (50 μM) diminished the proliferative response to TGF-β₁ (P = 0.02) as well as IGF-I-stimulated collagen production (P = 0.02). Thus resveratrol decreases intestinal smooth muscle cell numbers through its effects on cell cycle arrest and apoptosis and also decreases collagen synthesis by the cells. These effects could be useful in preventing the smooth muscle cell hyperplasia and collagen

  13. Fabrication of nanocomposites by collagen templated synthesis of layered double hydroxides assisted by an acrylic silane coupling agent

    NASA Astrophysics Data System (ADS)

    Sun, Yanqing; Zhou, Yuming; Wang, Zhiqiang; Ye, Xiaoyun

    2009-02-01

    The purpose of this study was to control the fabrication of nanocomposites at the nanoscale interface by collagen templated synthesis of Zn-Al layered double hydroxides (LDHs) assisted by γ-methacryloxypropyl trimethoxy silane (KH570) with further treatment of graft polymerization. The results show that collagen directs the growth of LDHs into curved nanorods by length of 300 nm in perfect consistency with collagen chain in both the size and flexility under the essential hydrophobic environment on the solid surface provided by KH570. The nanorods are aggregated into thin curved platelets due to strong interaction between collagen molecules themselves and strong interaction between collagen and LDH sheets. By further treatment of graft polymerization, the adjacent curved platelets encircle into numerous hollows via chemical linkage, achieving polyporous nanocomposites. Nanohybrid materials with this structure are especially interesting for applications as biosensors or supported catalysis.

  14. Aryl hydrocarbon receptor-driven signals inhibit collagen synthesis in the gut.

    PubMed

    Monteleone, Ivan; Zorzi, Francesca; Marafini, Irene; Di Fusco, Davide; Dinallo, Vincenzo; Caruso, Roberta; Izzo, Roberta; Franzè, Eleonora; Colantoni, Alfredo; Pallone, Francesco; Monteleone, Giovanni

    2016-04-01

    Fibrostrictures (FS) are a major complication of Crohn's disease (CD). Pathogenesis of FS is not fully understood, but activation of fibroblasts and excessive collagen deposition are crucial in the development of FS. Here, we investigated the role of aryl hydrocarbon receptor (AhR) in intestinal fibrosis. AhR RNA and protein expression were evaluated in intestinal fibroblasts of CD patients and controls. CD fibroblasts were stimulated with TGF-β1 or TNF-α in the presence or absence of the AhR activator Ficz, an AhR antagonist CH223191, or a specific AhR-silencing RNA. In CD fibroblasts, TGF-β1 and TNF-α increased Col1A1, Col3A1 and α-SMA transcripts and collagen secretion and this effect was reduced by Ficz and upregulated by CH22319. TGF-β1 or TNF-α induced activation of p38 and ERK1/2 MAP kinases was decreased by Ficz and increased by CH223191. The inhibitory effect of Ficz on Map kinase activation and collagen induction was abolished by AhR silencing. To assess the role of AhR in vivo, mice with trinitrobenzene-sulfonic-acid induced colonic fibrosis were given Ficz or CH223191. Mice given either Ficz or CH223191 produced less or more collagen respectively as compared with control mice. Our results indicate that AhR is a negative regulator of profibrotic signals in the gut. PMID:26786786

  15. Transdermal nicotine patch enhances type I collagen synthesis in abstinent smokers.

    PubMed

    Sørensen, Lars T; Jorgensen, Lars N; Zillmer, Rikke; Vange, Jakob; Hemmingsen, Ulla; Gottrup, Finn

    2006-01-01

    Cigarette smokers deposit less collagen, expressed as hydroxyproline, in granulation tissue than nonsmokers. We studied the effect of abstinence from smoking and transdermal nicotine patches on deposition of hydroxyproline, proline, type I procollagen, and total proteins. Fifty-four healthy smokers were studied during 10 days of smoking and again from days 10 to 20 following smoking cessation. After the first 10 days of abstinence they were randomized to double-blind treatment with transdermal nicotine patches of 25 mg/day or placebo for a period of 10 days. During this period and during smoking, an expanded polytetrafluoroethylene tube was implanted into the subcutis. Following removal of the implant, total amino acids and peptides were extracted. Hydroxyproline and proline were analyzed by high-pressure liquid chromatography, type I procollagen was analyzed by enzyme-linked immunoassay, and total proteins were determined colorimetrically. In the 39 subjects who complied with the study protocol, abstinence from smoking did not affect the deposition of hydroxyproline, proline, type I procollagen, or total protein in the implants. During abstinence, the type I procollagen level increased by 18% in the transdermal nicotine patches group and decreased by 10% in the placebo group (p<0.05). We conclude that 20 days of abstinence from smoking does not affect collagen deposition in granulation tissue. However, in abstinent smokers, transdermal nicotine patches appears to increase type I collagen synthesis.

  16. Development of biomimetic tilapia collagen nanofibers for skin regeneration through inducing keratinocytes differentiation and collagen synthesis of dermal fibroblasts.

    PubMed

    Zhou, Tian; Wang, Nanping; Xue, Yang; Ding, Tingting; Liu, Xin; Mo, Xiumei; Sun, Jiao

    2015-02-11

    In this study, tilapia skin collagen sponge and electrospun nanofibers were developed for wound dressing. The collagen sponge was composed of at least two α-peptides, and its denaturation temperature was 44.99 °C. It did not change the number of spleen-derived lymphocytes in BALB/c mice, the ratio of CD4+/CD8+ lymphocytes, and the level of IgG or IgM in Sprague-Dawley rat. The contact angle, tensile strength, and weight loss temperature of collagen nanofibers were 21.2°, 6.72±0.44 MPa, and 300 °C, respectively. The nanofibers could promote the viabilities of human keratinocytes (HaCaTs) and human dermal fibroblasts (HDFs), inducing epidermal differentiation through the gene expression of involucrin, filaggrin, and type I transglutaminase of HaCaTs, and they could also accelerate migration of HaCaTs with the expression of matrix metalloproteinase-9 and transforming growth factor-β1 (TGF-β1). Besides, the nanofibers could upregulate the protien level of Col-I in HDFs both via a direct effect and TGF-β1 secreted from HaCaTs, thus facilitating the formation of collagen fibers. Furthermore, the collagen nanofibers stimulated the skin regeneration rapidly and effectively in vivo. These biological effects could be explained as the contributions from the biomimic extracellular cell matrix structure, hydrophilicity, and the multiple amino acids of the collagen nanofibers. PMID:25598076

  17. Development of biomimetic tilapia collagen nanofibers for skin regeneration through inducing keratinocytes differentiation and collagen synthesis of dermal fibroblasts.

    PubMed

    Zhou, Tian; Wang, Nanping; Xue, Yang; Ding, Tingting; Liu, Xin; Mo, Xiumei; Sun, Jiao

    2015-02-11

    In this study, tilapia skin collagen sponge and electrospun nanofibers were developed for wound dressing. The collagen sponge was composed of at least two α-peptides, and its denaturation temperature was 44.99 °C. It did not change the number of spleen-derived lymphocytes in BALB/c mice, the ratio of CD4+/CD8+ lymphocytes, and the level of IgG or IgM in Sprague-Dawley rat. The contact angle, tensile strength, and weight loss temperature of collagen nanofibers were 21.2°, 6.72±0.44 MPa, and 300 °C, respectively. The nanofibers could promote the viabilities of human keratinocytes (HaCaTs) and human dermal fibroblasts (HDFs), inducing epidermal differentiation through the gene expression of involucrin, filaggrin, and type I transglutaminase of HaCaTs, and they could also accelerate migration of HaCaTs with the expression of matrix metalloproteinase-9 and transforming growth factor-β1 (TGF-β1). Besides, the nanofibers could upregulate the protien level of Col-I in HDFs both via a direct effect and TGF-β1 secreted from HaCaTs, thus facilitating the formation of collagen fibers. Furthermore, the collagen nanofibers stimulated the skin regeneration rapidly and effectively in vivo. These biological effects could be explained as the contributions from the biomimic extracellular cell matrix structure, hydrophilicity, and the multiple amino acids of the collagen nanofibers.

  18. Effect of Cold Plasma on Cell Viability and Collagen Synthesis in Cultured Murine Fibroblasts

    NASA Astrophysics Data System (ADS)

    Shi, Xingmin; Cai, Jingfen; Xu, Guimin; Ren, Hongbin; Chen, Sile; Chang, Zhengshi; Liu, Jinren; Huang, Chongya; Zhang, Guanjun; Wu, Xili

    2016-04-01

    An argon atmospheric pressure plasma jet was employed to treat L929 murine fibroblasts cultured in vitro. Experimental results showed that, compared with the control cells, the treatment of fibroblasts with 15 s of plasma led to a significant increase of cell viability and collagen synthesis, while the treatment of 25 s plasma resulted in a remarkable decrease. Exploration of related mechanisms suggested that cold plasma could up-regulate CyclinD1 gene expression and down-regulate p27 gene expression at a low dose, while it could down-regulate CyclinD1 expression and up-regulate p27 expression at a higher dose, thus altering the cell cycle progression, and then affecting cell viability and collagen synthesis of fibroblasts. supported partly by National Natural Science Foundation of China (Nos. 81372076, 51307133 and 51221005), China National Funds for Distinguished Young Scientists (No. 51125029), the Sci-Tech Project of Shaanxi Province of China (No. 2010K16-04), and the Fundamental Research Funds for the Central Universities of China (No. xkjc2013004)

  19. Bioinspired synthesis of hydroxyapatite nanocrystals in the presence of collagen and l-arginine: Candidates for bone regeneration.

    PubMed

    Brasinika, Despoina; Tsigkou, Olga; Tsetsekou, Athena; Missirlis, Yiannis F

    2016-04-01

    This work aims at the bioinspired synthesis of hydroxyapatite (HAp) crystals in the presence of both collagen and l-arginine, in an effort to obtain a homogeneous hybrid material, having a bone-like nanostructure. Collagen (Col) is the most commonly utilized protein in most species of life, while L-arginine (Arg) encourages cell attachment, proliferation, and differentiation on HAp surfaces. Transmission electron microscopy, X-ray diffraction and Fourier transform-infrared spectroscopy were used to analyze surface morphology and structure of nanocrystals obtained under different synthesis conditions. It was shown that collagen and arginine content affect HAp crystallization. Collagen has an inhibition effect since HAp crystal size is reduced with the increase of collagen content. The presence of arginine is crucial as a critical content exists (Ca(2+):Arg = 1:1) under which HAp nanocrystals coexist with brushite. Under the optimum synthesis conditions (HAp/Col weight ratio 70/30 and Ca(2+):Arg molar ratio 1:1) HAp nanoplates of a uniform size (around 10 × 10 nm) were obtained. The biocompatibility of this hybrid powder was assessed using human bone marrow derived mesenchymal stem cells (MSCs). Cell response in terms of MSC attachment (scanning electron microscopy) and viability/proliferation (Alamar Blue) demonstrated a noncytotoxic effect of the new material. PMID:25891542

  20. Activity of capryloyl collagenic acid against bacteria involved in acne.

    PubMed

    Fourniat, J; Bourlioux, P

    1989-12-01

    Synopsis Capryloyl collagenic acid (Lipacide C8Co) has similar bacteriostatic activity in vitro to that of benzoyl peroxide towards the bacteria found in acne lesions (Staphylococcus aureus, Staphylococcus epidermidis and Propionibacterium acnes) (MIC between 1 and 4 mg ml(-1) for C8Co, and between 0.5 and 5 mg ml(-1) for benzoyl peroxide). The presence of Emulgine M8 did not affect the bacteriostatic activity of C8Co. A 4% w/v solution of C8Co (incorporating Emulgine M8) fulfilled the criteria for an antiseptic preparation as laid down by the French Pharmacopoeia (10th Edition), and had a spectrum 5 bactericidal activity according to the French Standard AFNOR NF T 72-151. The excellent cutaneous tolerance of capryloyl collagenic acid would indicate that an aqueous solution might be of value for topical treatment of the bacterial component of acne. Résumé Activité antibactérienne de l'acide capryloyl-collagénique vis à vis des bactéries impliquées dans l'etiologie de l'acné L'acide capryloyl-collagénique (Lipacide C8Co) et le peroxyde de benzoyle présentent une activité bactériostatique in-vitroéquivalente vis à vis des espèces bactériennes retrouvées au niveau des lésions acnéiques (Staphylococcus aureus, S. epidermidis et Propionibacterium acnes) (CMI comprise entre 1 et 4 mg ml(-1) pour le lipoaminoacide, et 0,5 et 5 mg ml(-1) pour le peroxyde de benzoyle). La mise en solution aqueuse de l'acide capryloyl-collagénique en présence d'Emulgine M8 ne modifie pas son activité bactériostatique. Une telle solution, à 4% m/V d'acide capryloyl-collagénique et 5% m/V d'Emulgine M8, satisfait à l'essai d'activité des préparations antiseptiques décrit à la Pharmacopée Française (Xème Ed.) (concentration minimale antiseptique: 10% v/V, pour un temps de contact de 5 min à 32 degrees C entre les germes tests et la solution diluée en eau distillée), et posséde une activité bactéricide antiseptique spectre 5 conforme à la norme AFNOR NF T

  1. Biomimetic composite microspheres of collagen/chitosan/nano-hydroxyapatite: In-situ synthesis and characterization.

    PubMed

    Teng, Shu-Hua; Liang, Mian-Hui; Wang, Peng; Luo, Yong

    2016-01-01

    The collagen/chitosan/hydroxyapatite (COL/CS/HA) composite microspheres with a good spherical form and a high dispersity were successfully obtained using an in-situ synthesis method. The FT-IR and XRD results revealed that the inorganic phase in the microspheres was crystalline HA containing carbonate ions. The morphology of the composite microspheres was dependent on the HA content, and a more desirable morphology was achieved when 20 wt.% HA was contained. The composite microspheres exhibited a narrow particle distribution, most of which ranged from 5 to 10 μm. In addition, the needle-like HA nano-particles were uniformly distributed in the composite microspheres, and their crystallinity and crystal size decreased with the HA content. PMID:26478351

  2. Biomimetic composite microspheres of collagen/chitosan/nano-hydroxyapatite: In-situ synthesis and characterization.

    PubMed

    Teng, Shu-Hua; Liang, Mian-Hui; Wang, Peng; Luo, Yong

    2016-01-01

    The collagen/chitosan/hydroxyapatite (COL/CS/HA) composite microspheres with a good spherical form and a high dispersity were successfully obtained using an in-situ synthesis method. The FT-IR and XRD results revealed that the inorganic phase in the microspheres was crystalline HA containing carbonate ions. The morphology of the composite microspheres was dependent on the HA content, and a more desirable morphology was achieved when 20 wt.% HA was contained. The composite microspheres exhibited a narrow particle distribution, most of which ranged from 5 to 10 μm. In addition, the needle-like HA nano-particles were uniformly distributed in the composite microspheres, and their crystallinity and crystal size decreased with the HA content.

  3. Overexpression of RACK1 inhibits collagen synthesis in keloid fibroblasts via inhibition of transforming growth factor-β1/Smad signaling pathway

    PubMed Central

    Zhou, Ping; Shi, Lina; Li, Qing; Lu, Di

    2015-01-01

    Keloids are benign skin tumors characterized by collagen accumulation and hyperproliferation of fibroblasts. The receptor for activated C-kinase 1 (RACK1) was involved in liver fibrosis. However, the role of RACK1 in dermal fibrosis keloids is still unclear. Therefore, in this study, we investigated the effects of RACK1 on keloid fibroblasts (KFs) and transforming growth factor-β1 (TGF-β1)-induced collagen expression and explored the underlying mechanism. We found that RACK1 was decreased in KFs, overexpression of RACK1 significantly inhibited TGF-β1-induced KFs proliferation. RACK1 also obviously inhibited the expression of TGF-β1-induced TGF-β receptor I, II, type I collagen and α-smooth muscle actin (α-SMA) in human KFs. In addition, RACK1 suppressed the expression of TGF-β1-induced Smad2 and Smad3 phosphorylation in human KFs. Taken together, our study suggested that RACK1 inhibits collagen synthesis in KFs via inhibition the TGF-β1/Smad signaling pathway, and RACK1 is a potential target for treatment of the keloid disease. PMID:26629012

  4. Repeat mild heat shock increases dermal fibroblast activity and collagen production.

    PubMed

    Mayes, Andrew E; Holyoak, Caroline D

    2008-04-01

    Repeat mild heat shock (RMHS) has been shown to have anti-aging effects on cellular and biological processes within human dermal fibroblasts. We have investigated the potential of an abridged mild heat shock regime to impact upon the functional properties of human dermal fibroblasts derived from three donors (male, 12 years; female, 22 years; female, 65 years). For each donor mild heat shock increased the rate of contraction of fibroblast-containing collagen gels and increased the de novo synthesis of collagen. Thus, hormetic mechanisms are proposed to provide functional anti-aging benefits to skin cells.

  5. Angiotensin converting enzyme inhibitors mitigate collagen synthesis induced by a single dose of radiation to the whole thorax.

    PubMed

    Kma, Lakhan; Gao, Feng; Fish, Brian L; Moulder, John E; Jacobs, Elizabeth R; Medhora, Meetha

    2012-01-01

    Our long-term goal is to use angiotensin converting enzyme (ACE) inhibitors to mitigate the increase in lung collagen synthesis that is induced by irradiation to the lung, which could result from accidental exposure or radiological terrorism. Rats (WAG/RijCmcr) were given a single dose of 13 Gy (dose rate of 1.43 Gy/min) of X-irradiation to the thorax. Three structurally-different ACE inhibitors, captopril, enalapril and fosinopril were provided in drinking water beginning 1 week after irradiation. Rats that survived acute pneumonitis (at 6-12 weeks) were evaluated monthly for synthesis of lung collagen. Other endpoints included breathing rate, wet to dry lung weight ratio, and analysis of lung structure. Treatment with captopril (145-207 mg/m(2)/day) or enalapril (19-28 mg/m(2)/day), but not fosinopril (19-28 mg/m(2)/day), decreased morbidity from acute pneumonitis. Lung collagen in the surviving irradiated rats was increased over that of controls by 7 months after irradiation. This increase in collagen synthesis was not observed in rats treated with any of the three ACE inhibitors. Analysis of the lung morphology at 7 months supports the efficacy of ACE inhibitors against radiation-induced fibrosis. The effectiveness of fosinopril against fibrosis, but not against acute pneumonitis, suggests that pulmonary fibrosis may not be a simple consequence of injury during acute pneumonitis. In summary, three structurally-different ACE inhibitors mitigate the increase in collagen synthesis 7 months following irradiation of the whole thorax and do so, even when therapy is started one week after irradiation. PMID:22302041

  6. Effect of Phosphatase and Tensin Homologue on Chromosome 10 on Angiotensin II-Mediated Proliferation, Collagen Synthesis, and Akt/P27 Signaling in Neonatal Rat Cardiac Fibroblasts

    PubMed Central

    Nie, Ling; Zhao, Jing-Hong; Wang, Jiang; Song, Rong

    2016-01-01

    Cardiac fibroblasts (CFs) play a key role in cardiac fibrosis by regulating the balance between extracellular matrix synthesis and breakdown. Although phosphatase and tensin homologue on chromosome 10 (PTEN) has been found to play an important role in cardiovascular disease, it is not clear whether PTEN is involved in functional regulation of CFs. In the present study, PTEN was overexpressed in neonatal rat CFs via recombinant adenovirus-mediated gene transfer. The effects of PTEN overexpression on cell-cycle progression and angiotensin II- (Ang II-) mediated regulation of collagen metabolism, synthesis of matrix metalloproteinases, and Akt/P27 signaling were investigated. Compared with uninfected cells and cells infected with green fluorescent protein-expressing adenovirus (Ad-GFP), cells infected with PTEN-expressing adenovirus (Ad-PTEN) significantly increased PTEN protein and mRNA levels in CFs (P < 0.05). The proportion of CFs in the G1/S cell-cycle phase was significantly higher for PTEN-overexpressing cells. In addition, Ad-PTEN decreased mRNA expression and the protein synthesis rate of collagen types I and III and antagonized Ang II-induced collagen synthesis. Overexpression of PTEN also decreased Ang II-induced matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-1 (TIMP-1) production as well as gelatinase activity. Moreover, Ad-PTEN decreased Akt expression and increased P27 expression independent of Ang II stimulation. These results suggest that PTEN could regulate its functional effects in neonatal rat CFs partially via the Akt/P27 signaling pathway. PMID:27747225

  7. Collagenous gastritis.

    PubMed

    Colletti, R B; Trainer, T D

    1989-12-01

    Subepithelial fibrosis has previously been reported in the small intestine (collagenous sprue) and colon (collagenous colitis). We report a 15-yr-old girl with chronic gastritis and subepithelial fibrosis of the gastric corpus who presented with recurrent abdominal pain and acute upper gastrointestinal bleeding. Nodularity and erythema of the gastric corpus were persistent endoscopic findings. Biopsies revealed patchy chronic active gastritis with a striking focal thick band of collagen immediately beneath the surface epithelial cells that did not extend to deeper portions of the lamina propria. Contrast radiography demonstrated an abnormal mucosa of the gastric corpus with a mosaiclike surface pattern. Numerous studies have failed to elucidate the etiology. Despite treatment with ranitidine, sucralfate, and furazolidone, there has been no clinical or pathologic improvement. The pathogenesis and prognosis of collagenous gastritis, and its relationship to collagenous sprue and collagenous colitis, remain to be defined. PMID:2583419

  8. Preparation and biological activity of quaternized carboxymethyl chitosan conjugated with collagen peptide.

    PubMed

    Zhu, Xiaoming; Zhou, Xiaoyu; Yi, Jiayan; Tong, Jun; Wu, Huan; Fan, Lihong

    2014-09-01

    Tissue repair is a spontaneous process which initiated on wounding. If this complex mechanism is disturbed or impaired, the use of biomaterials might increase the chance of successful healing. In this view, a water-soluble chitosan derivative, quaternized carboxymethyl chitosan (QCMC) was prepared and collagen peptides (COPs) were grafted to the backbone by carbodiimide method. The reaction conditions affecting the degree of substitution (DS) were studied including the mass ratio of collagen peptide to QCMC, reaction temperature and reaction time. The hydrogen peroxide-scavenging activity could be different by changing the DS, concentration and molecular weight. MTT assay was used to investigate the cell viability of the derivative. The results indicated that the introduction of collagen peptide into the QCMC improved its hydrogen peroxide-scavenging activity and cell viability with the DS and concentration increased. Therefore, QCMC conjugated with collagen peptides may prove beneficial to the process of the wound-healing. PMID:24995634

  9. Inhibition of collagen synthesis by select calcium and sodium channel blockers can be mitigated by ascorbic acid and ascorbyl palmitate.

    PubMed

    Ivanov, Vadim; Ivanova, Svetlana; Kalinovsky, Tatiana; Niedzwiecki, Aleksandra; Rath, Matthias

    2016-01-01

    Calcium, sodium and potassium channel blockers are widely prescribed medications for a variety of health problems, most frequently for cardiac arrhythmias, hypertension, angina pectoris and other disorders. However, chronic application of channel blockers is associated with numerous side effects, including worsening cardiac pathology. For example, nifedipine, a calcium-channel blocker was found to be associated with increased mortality and increased risk for myocardial infarction. In addition to the side effects mentioned above by different channel blockers, these drugs can cause arterial wall damage, thereby contributing to vascular wall structure destabilization and promoting events facilitating rupture of plaques. Collagen synthesis is regulated by ascorbic acid, which is also essential for its optimum structure as a cofactor in lysine and proline hydroxylation, a precondition for optimum crosslinking of collagen and elastin. Therefore, the main objective in this study was to evaluate effects of various types of channel blockers on intracellular accumulation and cellular functions of ascorbate, specifically in relation to formation and extracellular deposition of major collagen types relevant for vascular function. Effects of select Na- and Ca- channel blockers on collagen synthesis and deposition were evaluated in cultured human dermal fibroblasts and aortic smooth muscle cells by immunoassay. All channel blockers tested demonstrated inhibitory effects on collagen type I deposition to the ECM by fibroblasts, each to a different degree. Ascorbic acid significantly increased collagen I ECM deposition. Nifedipine (50 µM), a representative of channel blockers tested, significantly reduced ascorbic acid and ascorbyl palmitate-dependent ECM deposition of collagen type l and collagen type lV by cultured aortic smooth muscle cells. In addition, nifedipine (50 µM) significantly reduced ascorbate-dependent collagen type l and type lV synthesis by cultured aortic smooth

  10. Inhibition of collagen synthesis by select calcium and sodium channel blockers can be mitigated by ascorbic acid and ascorbyl palmitate.

    PubMed

    Ivanov, Vadim; Ivanova, Svetlana; Kalinovsky, Tatiana; Niedzwiecki, Aleksandra; Rath, Matthias

    2016-01-01

    Calcium, sodium and potassium channel blockers are widely prescribed medications for a variety of health problems, most frequently for cardiac arrhythmias, hypertension, angina pectoris and other disorders. However, chronic application of channel blockers is associated with numerous side effects, including worsening cardiac pathology. For example, nifedipine, a calcium-channel blocker was found to be associated with increased mortality and increased risk for myocardial infarction. In addition to the side effects mentioned above by different channel blockers, these drugs can cause arterial wall damage, thereby contributing to vascular wall structure destabilization and promoting events facilitating rupture of plaques. Collagen synthesis is regulated by ascorbic acid, which is also essential for its optimum structure as a cofactor in lysine and proline hydroxylation, a precondition for optimum crosslinking of collagen and elastin. Therefore, the main objective in this study was to evaluate effects of various types of channel blockers on intracellular accumulation and cellular functions of ascorbate, specifically in relation to formation and extracellular deposition of major collagen types relevant for vascular function. Effects of select Na- and Ca- channel blockers on collagen synthesis and deposition were evaluated in cultured human dermal fibroblasts and aortic smooth muscle cells by immunoassay. All channel blockers tested demonstrated inhibitory effects on collagen type I deposition to the ECM by fibroblasts, each to a different degree. Ascorbic acid significantly increased collagen I ECM deposition. Nifedipine (50 µM), a representative of channel blockers tested, significantly reduced ascorbic acid and ascorbyl palmitate-dependent ECM deposition of collagen type l and collagen type lV by cultured aortic smooth muscle cells. In addition, nifedipine (50 µM) significantly reduced ascorbate-dependent collagen type l and type lV synthesis by cultured aortic smooth

  11. Inhibition of collagen synthesis by select calcium and sodium channel blockers can be mitigated by ascorbic acid and ascorbyl palmitate

    PubMed Central

    Ivanov, Vadim; Ivanova, Svetlana; Kalinovsky, Tatiana; Niedzwiecki, Aleksandra; Rath, Matthias

    2016-01-01

    Calcium, sodium and potassium channel blockers are widely prescribed medications for a variety of health problems, most frequently for cardiac arrhythmias, hypertension, angina pectoris and other disorders. However, chronic application of channel blockers is associated with numerous side effects, including worsening cardiac pathology. For example, nifedipine, a calcium-channel blocker was found to be associated with increased mortality and increased risk for myocardial infarction. In addition to the side effects mentioned above by different channel blockers, these drugs can cause arterial wall damage, thereby contributing to vascular wall structure destabilization and promoting events facilitating rupture of plaques. Collagen synthesis is regulated by ascorbic acid, which is also essential for its optimum structure as a cofactor in lysine and proline hydroxylation, a precondition for optimum crosslinking of collagen and elastin. Therefore, the main objective in this study was to evaluate effects of various types of channel blockers on intracellular accumulation and cellular functions of ascorbate, specifically in relation to formation and extracellular deposition of major collagen types relevant for vascular function. Effects of select Na- and Ca- channel blockers on collagen synthesis and deposition were evaluated in cultured human dermal fibroblasts and aortic smooth muscle cells by immunoassay. All channel blockers tested demonstrated inhibitory effects on collagen type I deposition to the ECM by fibroblasts, each to a different degree. Ascorbic acid significantly increased collagen I ECM deposition. Nifedipine (50 µM), a representative of channel blockers tested, significantly reduced ascorbic acid and ascorbyl palmitate-dependent ECM deposition of collagen type l and collagen type lV by cultured aortic smooth muscle cells. In addition, nifedipine (50 µM) significantly reduced ascorbate-dependent collagen type l and type lV synthesis by cultured aortic smooth

  12. A role for adhesion and degranulation-promoting adapter protein in collagen-induced platelet activation mediated via integrin α2β1

    PubMed Central

    JARVIS, G. E.; BIHAN, D.; HAMAIA, S.; PUGH, N.; GHEVAERT, C. J. G.; PEARCE, A. C.; HUGHES, C. E.; WATSON, S. P.; WARE, J.; RUDD, C. E.; FARNDALE, R. W.

    2013-01-01

    Summary Background Collagen-induced platelet activation is a key step in the development of arterial thrombosis via its interaction with the receptors glycoprotein (GP)VI and integrin α2β1. Adhesion and degranulation-promoting adapter protein (ADAP) regulates αIIbβ3 in platelets and αLβ2 in T cells, and is phosphorylated in GPVI-deficient platelets activated by collagen. Objectives To determine whether ADAP plays a role in collagen-induced platelet activation and in the regulation and function of α2β1. Methods Using ADAP−/− mice and synthetic collagen peptides, we investigated the role of ADAP in platelet aggregation, adhesion, spreading, thromboxane synthesis, and tyrosine phosphorylation. Results and Conclusions Platelet aggregation and phosphorylation of phospholipase Cγ2 induced by collagen were attenuated in ADAP−/− platelets. However, aggregation and signaling induced by collagen-related peptide (CRP), a GPVI-selective agonist, were largely unaffected. Platelet adhesion to CRP was also unaffected by ADAP deficiency. Adhesion to the α2β1-selective ligand GFOGER and to a peptide (III-04), which supports adhesion that is dependent on both GPVI and α2β1, was reduced in ADAP−/− platelets. An impedance-based label-free detection technique, which measures adhesion and spreading of platelets, indicated that, in the absence of ADAP, spreading on GFOGER was also reduced. This was confirmed with non-fluorescent differential-interference contrast microscopy, which revealed reduced filpodia formation in ADAP−/− platelets adherent to GFOGER. This indicates that ADAP plays a role in mediating platelet activation via the collagen-binding integrin α2β1. In addition, we found that ADAP−/− mice, which are mildly thrombocytopenic, have enlarged spleens as compared with wild-type animals. This may reflect increased removal of platelets from the circulation. PMID:22103309

  13. Novel synthesis strategy for composite hydrogel of collagen/hydroxyapatite-microsphere originating from conversion of CaCO3 templates.

    PubMed

    Wei, Qingrong; Lu, Jian; Wang, Qiaoying; Fan, Hongsong; Zhang, Xingdong

    2015-03-20

    Inspired by coralline-derived hydroxyapatite, we designed a methodological route to synthesize carbonated-hydroxyapatite microspheres from the conversion of CaCO3 spherulite templates within a collagen matrix under mild conditions and thus constructed the composite hydrogel of collagen/hydroxyapatite-microspheres. Fourier transform infrared spectroscopy (FTIR) and x-ray diffraction (XRD) were employed to confirm the successful generation of the carbonated hydroxyapatite phase originating from CaCO3, and the ratios of calcium to phosphate were tracked over time. Variations in the weight portion of the components in the hybrid gels before and after the phase transformation of the CaCO3 templates were identified via thermogravimetric analysis (TGA). Scanning electron microscopy (SEM) shows these composite hydrogels have a unique multiscale microstructure consisting of a collagen nanofibril network and hydroxyapatite microspheres. The relationship between the hydroxyapatite nanocrystals and the collagen fibrils was revealed by transmission electron microscopy (TEM) in detail, and the selected area electron diffraction (SAED) pattern further confirmed the results of the XRD analyses which show the typical low crystallinity of the generated hydroxyapatite. This smart synthesis strategy achieved the simultaneous construction of microscale hydroxyapatite particles and collagen fibrillar hydrogel, and appears to provide a novel route to explore an advanced functional hydrogel materials with promising potentials for applications in bone tissue engineering and reconstruction medicine.

  14. Plasma membrane overgrowth causes fibrotic collagen accumulation and immune activation in Drosophila adipocytes

    PubMed Central

    Zang, Yiran; Wan, Ming; Liu, Min; Ke, Hongmei; Ma, Shuangchun; Liu, Lu-Ping; Ni, Jian-Quan; Carlos Pastor-Pareja, José

    2015-01-01

    Many chronic diseases are associated with fibrotic deposition of Collagen and other matrix proteins. Little is known about the factors that determine preferential onset of fibrosis in particular tissues. Here we show that plasma membrane (PM) overgrowth causes pericellular Collagen accumulation in Drosophila adipocytes. We found that loss of Dynamin and other endocytic components causes pericellular trapping of outgoing Collagen IV due to dramatic cortex expansion when endocytic removal of PM is prevented. Deposits also form in the absence of negative Toll immune regulator Cactus, excess PM being caused in this case by increased secretion. Finally, we show that trimeric Collagen accumulation, downstream of Toll or endocytic defects, activates a tissue damage response. Our work indicates that traffic imbalances and PM topology may contribute to fibrosis. It also places fibrotic deposits both downstream and upstream of immune signaling, consistent with the chronic character of fibrotic diseases. DOI: http://dx.doi.org/10.7554/eLife.07187.001 PMID:26090908

  15. The effect of Centella asiatica, vitamins, glycolic acid and their mixtures preparations in stimulating collagen and fibronectin synthesis in cultured human skin fibroblast.

    PubMed

    Hashim, Puziah

    2014-03-01

    Centella asiatica (Linn.) Urban is well known in promoting wound healing and provides significant benefits in skin care and therapeutic products formulation. Glycolic acid and vitamins also play a role in the enhancement of collagen and fibronectin synthesis. Here, we evaluate the specific effect of Centella asiatica (CA), vitamins, glycolic acid and their mixture preparations to stimulate collagen and fibronectin synthesis in cultured human fibroblast cells. The fibroblast cells are incubated with CA, glycolic acid, vitamins and their mixture preparations for 48 h. The cell lysates were analyzed for protein content and collagen synthesis by direct binding enzyme immunoassay. The fibronectin of the cultured supernatant was measured by sandwich enzyme immunoassay. The results showed that CA, glycolic acid, vitamins A, E and C significantly stimulate collagen and fibronectin synthesis in the fibroblast. Addition of glycolic acid and vitamins to CA further increased the levels of collagen and fibronectin synthesis to 8.55 and 23.75 μg/100 μg, respectively. CA, glycolic acid, vitamins A, E, and C, and their mixtures demonstrated stimulatory effect on both extra-cellular matrix synthesis of collagen and fibronectin in in vitro studies on human foreskin fibroblasts, which is beneficial to skin care and therapeutic products formulation.

  16. Constitutive Smad signaling and Smad-dependent collagen gene expression in mouse embryonic fibroblasts lacking peroxisome proliferator-activated receptor-{gamma}

    SciTech Connect

    Ghosh, Asish K Wei, Jun; Wu, Minghua; Varga, John

    2008-09-19

    Transforming growth factor-{beta} (TGF-{beta}), a potent inducer of collagen synthesis, is implicated in pathological fibrosis. Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is a nuclear hormone receptor that regulates adipogenesis and numerous other biological processes. Here, we demonstrate that collagen gene expression was markedly elevated in mouse embryonic fibroblasts (MEFs) lacking PPAR-{gamma} compared to heterozygous control MEFs. Treatment with the PPAR-{gamma} ligand 15d-PGJ{sub 2} failed to down-regulate collagen gene expression in PPAR-{gamma} null MEFs, whereas reconstitution of these cells with ectopic PPAR-{gamma} resulted in their normalization. Compared to control MEFs, PPAR-{gamma} null MEFs displayed elevated levels of the Type I TGF-{beta} receptor (T{beta}RI), and secreted more TGF-{beta}1 into the media. Furthermore, PPAR-{gamma} null MEFs showed constitutive phosphorylation of cellular Smad2 and Smad3, even in the absence of exogenous TGF-{beta}, which was abrogated by the ALK5 inhibitor SB431542. Constitutive Smad2/3 phosphorylation in PPAR-{gamma} null MEFs was associated with Smad3 binding to its cognate DNA recognition sequences, and interaction with coactivator p300 previously implicated in TGF-{beta} responses. Taken together, these results indicate that loss of PPAR-{gamma} in MEFs is associated with upregulation of collagen synthesis, and activation of intracellular Smad signal transduction, due, at least in part, to autocrine TGF-{beta} stimulation.

  17. A three-dimensional collagen construct to model lipopolysaccharide-induced activation of BV2 microglia

    PubMed Central

    2014-01-01

    Background We report a novel method of culturing microglia in three dimension (3D) using collagen as a substrate. By culturing microglia within a matrix, we aim to emulate the physical state of microglia embedded within parenchyma. Methods BV2 microglia cell suspensions were prepared with type I collagen and cast into culture plates. To characterise the BV2 microglia cultured in 3D, the cultures were evaluated for their viability, cell morphology and response to lipopolysaccharide (LPS) activation. Conventional monolayer cultures (grown on uncoated and collagen-coated polystyrene) were set up concurrently for comparison. Results BV2 microglia in 3D collagen matrices were viable at 48 hrs of culture and exhibit a ramified morphology with multiplanar cytoplasmic projections. Following stimulation with 1 μg/ml LPS, microglia cultured in 3D collagen gels increase their expression of nitric oxide (NO) and CD40, indicating their capacity to become activated within the matrix. Up to 97.8% of BV2 microglia grown in 3D cultures gained CD40 positivity in response to LPS, compared to approximately 60% of cells grown in a monolayer (P < .05). BV2 microglia in 3D collagen gels also showed increased mRNA and protein expression of inflammatory cytokines IL-6, TNF-α and the chemoattractant MCP-1 following LPS stimulation. Conclusions In summary, BV2 microglia cultured in 3D collagen hydrogels exhibit multiplanar cytoplasmic projections and undergo a characteristic and robust activation response to LPS. This culture system is accessible to a wide range of analyses and provides a useful new in vitro tool for research into microglial activation. PMID:25074682

  18. Preparation of polysaccharide loaded collagen membrane with anti-oxidative activity.

    PubMed

    Shu, Zibin; Ding, Shengli; He, Xiaohong; Dai, Xuemei; Xiao, Qian; Yang, Min; Leng, Xue; Ma, Yanshun; Yang, Hua

    2015-01-01

    The scavenging activity of polysaccharides from Lycium barbarum, Lentinus edodes and Ganoderma Lucidum Karst to DPPH free radicals was investigated. It was found that among the three polysaccharides, Lycium barbarum polysaccharides (LBP) exhibits the best scavenging activity. Polysaccharide loaded collagen membranes were prepared by mixing LBP with collagen, starch, glycerol, sodium carboxymethyl cellulose and glutaraldehyde. In vitro drug release from membranes was evaluated. With increasing the immersion time, the release rate first increases and then slows down. Meanwhile, the scavenging activity to DPPH radicals exhibits similar variation, in agreement with a good release effect of the membrane. The optimal formulation of collagen membrane and preparation parameters were obtained considering the overall properties and the scavenging activity to radicals. PMID:26406078

  19. Human prostatic acid phosphatase directly stimulates collagen synthesis and alkaline phosphatase content of isolated bone cells

    SciTech Connect

    Ishibe, M.; Rosier, R.N.; Puzas, J.E. )

    1991-10-01

    Human prostatic acid phosphatase (hPAP) directly enhances the differentiated characteristics of isolated bone cells in vitro. This enzyme, when added to cell cultures for 24 h in vitro stimulates collagen synthesis and the production of alkaline phosphatase. The effects are dose dependent, with statistically significant effects occurring from 0.1-100 nM hPAP. Concentrations higher than 100 nM do not evoke greater effects. The maximal effect of hPAP occurs between 12 and 24 h of exposure. The cells stimulated to the greatest degree are osteoprogenitor cells and osteoblasts. Fibroblasts isolated from the same tissue show a lesser sensitivity to hPAP. hPAP has no detectable effect on cell proliferation, as measured by radiolabeled thymidine incorporation or total DNA synthesis. None of the observations reported in this work can be attributed to contaminating proteins in the hPAP preparation. hPAP was radiolabeled with 125I and was used for affinity binding and cross-linking studies. Scatchard analysis of specific binding indicated the presence of 1.0 X 10(5) high affinity binding sites/cell, with a Kd of 6.5 nM. Cross-linking studies demonstrated the presence of one 320-kDa binding complex. The pH profile and kinetic determinations of Km and maximum velocity for hPAP were similar to those previously reported, except for the finding of positive cooperativity of the substrate with the enzyme under the conditions of our assay. We believe that the direct stimulation of bone-forming cells by hPAP may contribute to the sclerotic nature of skeletal bone around sites of neoplastic prostatic metastases and that the effect of the enzyme is probably mediated by a plasma membrane receptor.

  20. RB1CC1 Protein Suppresses Type II Collagen Synthesis in Chondrocytes and Causes Dwarfism*

    PubMed Central

    Nishimura, Ichiro; Chano, Tokuhiro; Kita, Hiroko; Matsusue, Yoshitaka; Okabe, Hidetoshi

    2011-01-01

    RB1-inducible coiled-coil 1 (RB1CC1) functions in various processes, such as cell growth, differentiation, senescence, apoptosis, and autophagy. The conditional transgenic mice with cartilage-specific RB1CC1 excess that were used in the present study were made for the first time by the Cre-loxP system. Cartilage-specific RB1CC1 excess caused dwarfism in mice without causing obvious abnormalities in endochondral ossification and subsequent skeletal development from embryo to adult. In vitro and in vivo analysis revealed that the dwarf phenotype in cartilaginous RB1CC1 excess was induced by reductions in the total amount of cartilage and the number of cartilaginous cells, following suppressions of type II collagen synthesis and Erk1/2 signals. In addition, we have demonstrated that two kinds of SNPs (T-547C and C-468T) in the human RB1CC1 promoter have significant influence on the self-transcriptional level. Accordingly, human genotypic variants of RB1CC1 that either stimulate or inhibit RB1CC1 transcription in vivo may cause body size variations. PMID:22049074

  1. Collagen-binding VEGF mimetic peptide: Structure, matrix interaction, and endothelial cell activation

    NASA Astrophysics Data System (ADS)

    Chan, Tania R.

    Long term survival of artificial tissue constructs depends greatly on proper vascularization. In nature, differentiation of endothelial cells and formation of vasculature are directed by dynamic spatio-temporal cues in the extracellular matrix that are difficult to reproduce in vitro. In this dissertation, we present a novel bifunctional peptide that mimics matrix-bound vascular endothelial growth factor (VEGF), which can be used to encode spatially controlled angiogenic signals in collagen-based scaffolds. The peptide, QKCMP, contains a collagen mimetic domain (CMP) that binds to type I collagen by a unique triple helix hybridization mechanism and a VEGF mimetic domain (QK) with pro-angiogenic activity. We demonstrate QKCMP's ability to hybridize with native and heat denatured collagens through a series of binding studies on collagen and gelatin substrates. Circular dichroism experiments show that the peptide retains the triple helical structure vital for collagen binding, and surface plasmon resonance study confirms the molecular interaction between the peptide and collagen strands. Cell culture studies demonstrate QKCMP's ability to induce endothelial cell morphogenesis and network formation as a matrix-bound factor in 2D and 3D collagen scaffolds. We also show that the peptide can be used to spatially modify collagen-based substrates to promote localized endothelial cell activation and network formation. To probe the biological events that govern these angiogenic cellular responses, we investigated the cell signaling pathways activated by collagen-bound QKCMP and determined short and long-term endothelial cell response profiles for p38, ERK1/2, and Akt signal transduction cascades. Finally, we present our efforts to translate the peptide's in vitro bioactivity to an in vivo burn injury animal model. When implanted at the wound site, QKCMP functionalized biodegradable hydrogels induce enhanced neovascularization in the granulation tissue. The results show QKCMP

  2. THE DIFFERENTIAL REGULATION OF CELL MOTILE ACTIVITY THROUGH MATRIX STIFFNESS AND POROSITY IN THREE DIMENSIONAL COLLAGEN MATRICES

    PubMed Central

    Miron-Mendoza, Miguel; Seemann, Joachim; Grinnell, Frederick

    2010-01-01

    In three dimensional collagen matrices, cell motile activity results in collagen translocation, cell spreading and cell migration. Cells can penetrate into the matrix as well as spread and migrate along its surface. In the current studies, we quantitatively characterize collagen translocation, cell spreading and cell migration in relationship to collagen matrix stiffness and porosity. Collagen matrices prepared with 1 to 4 mg/ml collagen exhibited matrix stiffness (storage modulus measured by oscillating rheometry) increasing from 4 to 60 Pa and matrix porosity (measured by scanning electron microscopy) decreasing from 4 to 1 μm2. Over this collagen concentration range, the consequences of cell motile activity changed markedly. As collagen concentration increased, cells no longer were able to cause translocation of collagen fibrils. Cell migration increased and cell spreading changed from dendritic to more flattened and polarized morphology depending on location of cells within or on the surface of the matrix. Collagen translocation appeared to depend primarily on matrix stiffness, whereas cell spreading and migration were less dependent on matrix stiffness and more dependent on collagen matrix porosity. PMID:20537378

  3. Inhibitory effects of prostaglandin E2 on collagen synthesis and cell proliferation in human stellate cells from pancreatic head adenocarcinoma

    PubMed Central

    2014-01-01

    Background Several studies have described an increased cyclooxygenase-2 (COX-2) expression in pancreatic cancer, but the role of COX-2 in tumour development and progression is not clear. The aim of the present study was to examine expression of COX-2 in cancer cells and stromal cells in pancreatic cancer specimens, and to explore the role of PGE2 in pancreatic stellate cell proliferation and collagen synthesis. Methods Immunohistochemistry and immunofluorescence was performed on slides from whole sections of tissue blocks using antibodies against COX-2 and α-smooth muscle actin (αSMA). Pancreatic stellate cells (PSC) were isolated from surgically resected tumour tissue by the outgrowth method. Cells were used between passages 4 and 8. Collagen synthesis was determined by [3H]-proline incorporation, or by enzyme immunoassay measurement of collagen C-peptide. DNA synthesis was measured by incorporation of [3H]-thymidine in DNA. Cyclic AMP (cAMP) was determined by radioimmunoassay. Collagen 1A1 mRNA was determined by RT-qPCR. Results Immunohistochemistry staining showed COX-2 in pancreatic carcinoma cells, but not in stromal cells. All tumours showed positive staining for αSMA in the fibrotic stroma. Cultured PSC expressed COX-2, which could be further induced by interleukin-1β (IL-1β), epidermal growth factor (EGF), thrombin, and PGE2, but not by transforming growth factor-β1 (TGFβ). Indirect coculture with the adenocarcinoma cell line BxPC-3, but not HPAFII or Panc-1, induced COX-2 expression in PSC. Treatment of PSC with PGE2 strongly stimulated cAMP accumulation, mediated by EP2 receptors, and also stimulated phosphorylation of extracellular signal-regulated kinase (ERK). Treatment of PSC with PGE2 or forskolin suppressed both TGFβ-stimulated collagen synthesis and PDGF-stimulated DNA synthesis. Conclusions The present results show that COX-2 is mainly produced in carcinoma cells and suggest that the cancer cells are the main source of PGE2 in pancreatic

  4. Importance of amino acid composition to improve skin collagen protein synthesis rates in UV-irradiated mice.

    PubMed

    Murakami, Hitoshi; Shimbo, Kazutaka; Inoue, Yoshiko; Takino, Yoshinobu; Kobayashi, Hisamine

    2012-06-01

    Skin collagen metabolism abnormalities induced by ultraviolet (UV) radiation are the major causes of skin photoaging. It has been shown that the one-time exposure of UV irradiation decreases procollagen mRNA expression in dermis and that chronic UV irradiation decreases collagen amounts and induces wrinkle formation. Amino acids are generally known to regulate protein metabolism. Therefore, we investigated the effects of UV irradiation and various orally administered amino acids on skin collagen synthesis rates. Groups of 4-5 male, 8-week-old HR-1 hairless mice were irradiated with UVB (66 mJ/cm2) twice every other day, then fasted for 16 h. The fractional synthesis rate (FSR; %/h) of skin tropocollagen was evaluated by incorporating L-[ring-2H5]-phenylalanine. We confirmed that the FSR of dermal tropocollagen decreased after UVB irradiation. The FSR of dermal tropocollagen was measured 30 min after a single oral administration of amino acids (1 g/kg) to groups of 5-16 UVB-irradiated mice. Branched-chain amino acids (BCAA, 1.34±0.32), arginine (Arg, 1.66±0.39), glutamine (Gln, 1.75±0.60), and proline (Pro, 1.48±0.26) did not increase the FSR of skin tropocollagen compared with distilled water, which was used as a control (1.56±0.30). However, essential amino acids mixtures (BCAA+Arg+Gln, BCAA+Gln, and BCAA+Pro) significantly increased the FSR (2.07±0.58, 2.04±0.54, 2.01±0.50 and 2.07±0.59, respectively). This result suggests that combinations of BCAA and glutamine or proline are important for restoring dermal collagen protein synthesis impaired by UV irradiation.

  5. Human Collagen Prolyl 4-Hydroxylase Is Activated by Ligands for Its Iron Center.

    PubMed

    Vasta, James D; Raines, Ronald T

    2016-06-14

    Collagen is the most abundant protein in animals. The posttranslational hydroxylation of proline residues in collagen contributes greatly to its conformational stability. Deficient hydroxylation is associated with a variety of disease states, including scurvy. The hydroxylation of proline residues in collagen is catalyzed by an Fe(II)- and α-ketoglutarate-dependent dioxygenase, collagen prolyl 4-hydroxylase (CP4H). CP4H has long been known to suffer oxidative inactivation during catalysis, and the cofactor ascorbate (vitamin C) is required to reactivate the enzyme by reducing its iron center from Fe(III) to Fe(II). Herein, we report on the discovery of the first synthetic activators of CP4H. Specifically, we find that 2,2'-bipyridine-4-carboxylate and 2,2'-bipyridine-5-carboxylate serve as ligands for the iron center in human CP4H that enhance the rate of ascorbate-dependent reactivation. This new mode of CP4H activation is available to other biheteroaryl compounds but does not necessarily extend to other prolyl 4-hydroxylases. As collagen is weakened in many indications, analogous activators of CP4H could have therapeutic benefits. PMID:27183028

  6. Multifunctional role of osteopontin in directing intrafibrillar mineralization of collagen and activation of osteoclasts

    PubMed Central

    Rodriguez, Douglas E.; Thula-Mata, Taili; Toro, Edgardo J.; Yeh, Ya-Wen; Holt, Carl; Holliday, L. Shannon; Gower, Laurie B.

    2013-01-01

    Mineralized collagen composites are of interest because they have the potential to provide a bone-like scaffold that stimulates the natural processes of resorption and remodeling. Working toward this goal, our group has previously shown that the nanostructure of bone can be reproduced using a polymer-induced liquid-precursor (PILP) process, which enables intrafibrillar mineralization of collagen with hydroxyapatite (HA) to be achieved. This prior work used polyaspartic acid (pASP), a simple mimic for acidic non-collagenous proteins (NCPs), to generate nanodroplets/nanoparticles of an amorphous mineral precursor which can infiltrate the interstices of type-I collagen fibrils. In this study we show that osteopontin (OPN) can similarly serve as a process-directing agent for the intrafibrillar mineralization of collagen, even though OPN is generally considered a mineralization inhibitor. We also found that inclusion of OPN in the mineralization process promotes the interaction of mouse marrow-derived osteoclasts with PILP-remineralized bone that was previously demineralized, as measured by actin ring formation. While osteoclast activation occurred when pASP was used as the process-directing agent, using OPN resulted in a dramatic effect on osteoclast activation, presumably because of the inherent arginine-glycine-aspartate acid (RGD) ligands of OPN. By capitalizing on the multifunctionality of OPN, these studies may lead the way to producing biomimetic bone substitutes with the capability of tailorable bioresorption rates. PMID:24140612

  7. Multifunctional role of osteopontin in directing intrafibrillar mineralization of collagen and activation of osteoclasts.

    PubMed

    Rodriguez, Douglas E; Thula-Mata, Taili; Toro, Edgardo J; Yeh, Ya-Wen; Holt, Carl; Holliday, L Shannon; Gower, Laurie B

    2014-01-01

    Mineralized collagen composites are of interest because they have the potential to provide a bone-like scaffold that stimulates the natural processes of resorption and remodeling. Working towards this goal, our group has previously shown that the nanostructure of bone can be reproduced using a polymer-induced liquid-precursor (PILP) process, which enables intrafibrillar mineralization of collagen with hydroxyapatite to be achieved. This prior work used polyaspartic acid (pASP), a simple mimic for acidic non-collagenous proteins, to generate nanodroplets/nanoparticles of an amorphous mineral precursor which can infiltrate the interstices of type-I collagen fibrils. In this study we show that osteopontin (OPN) can similarly serve as a process-directing agent for the intrafibrillar mineralization of collagen, even though OPN is generally considered a mineralization inhibitor. We also found that inclusion of OPN in the mineralization process promotes the interaction of mouse marrow-derived osteoclasts with PILP-remineralized bone that was previously demineralized, as measured by actin ring formation. While osteoclast activation occurred when pASP was used as the process-directing agent, using OPN resulted in a dramatic effect on osteoclast activation, presumably because of the inherent arginine-glycine-aspartate acid ligands of OPN. By capitalizing on the multifunctionality of OPN, these studies may lead the way to producing biomimetic bone substitutes with the capability of tailorable bioresorption rates.

  8. Properties of alkali-solubilized collagen solution crosslinked by N-hydroxysuccinimide activated adipic acid

    NASA Astrophysics Data System (ADS)

    Chen, Yihui; Zhang, Min; Liu, Wentao; Li, Guoying

    2011-03-01

    The effect of N-hydroxysuccinimide activated adipic acid (NHS-AA) on the properties of alkali-solubilized collagen solutions was examined. The residual amino group content in crosslinked collagen, determined by trinitrobenzensulfonic acid (TNBS) assay, was decreased with increasing NHS-AA concentration. The results from differential scanning calorimeter (DSC) indicated that the maximum denaturation temperature ( T d) of crosslinked collagen solution was about 4.2°C higher than that of un-crosslinked collagen solution (36.6°C). Moreover, the values of storage modulus ( G'), loss modulus ( G″) and complex viscosity ( η*), obtained by means of dynamic frequency sweeps, were increased as NHS-AA concentration added up to 1.5 mM, and then decreased slightly when further increased NHS-AA concentration. Besides, for collagen solution crosslinked with 1.5 mM NHS-AA, dynamic denaturation temperature ( T dd) was about 1.1°C lower than T d (40.8°C), and the Arrhenius-type time-temperature superposition (TTS) principle was applied to yield the activation energy to be 474.4 kJmol-1.

  9. Generation of biologically active endostatin fragments from human collagen XVIII by distinct matrix metalloproteases

    SciTech Connect

    Heljasvaara, Ritva; Nyberg, Pia; Luostarinen, Jani; Parikka, Mataleena; Heikkilae, Pia; Rehn, Marko; Sorsa, Timo; Salo, Tuula; Pihlajaniemi, Taina . E-mail: taina.pihlajaniemi@oulu.fi

    2005-07-15

    Endostatin, a potent inhibitor of endothelial cell proliferation, migration, angiogenesis and tumor growth, is proteolytically cleaved from the C-terminal noncollagenous NC1 domain of type XVIII collagen. We investigated the endostatin formation from human collagen XVIII by several MMPs in vitro. The generation of endostatin fragments differing in molecular size (24-30 kDa) and in N-terminal sequences was identified in the cases of MMP-3, -7, -9, -13 and -20. The cleavage sites were located in the protease-sensitive hinge region between the trimerization and endostatin domains of NC1. MMP-1, -2, -8 and -12 did not show any significant activity against the C-terminus of collagen XVIII. The anti-proliferative effect of the 20-kDa endostatin, three longer endostatin-containing fragments generated in vitro by distinct MMPs and the entire NC1 domain, on bFGF-stimulated human umbilical vein endothelial cells was established. The anti-migratory potential of some of these fragments was also studied. In addition, production of endostatin fragments between 24-30 kDa by human hepatoblastoma cells was shown to be due to MMP action on type XVIII collagen. Our results indicate that certain, especially cancer-related, MMP family members can generate biologically active endostatin-containing polypeptides from collagen XVIII and thus, by releasing endostatin fragments, may participate in the inhibition of endothelial cell proliferation, migration and angiogenesis.

  10. A comparative study of skin cell activities in collagen and fibrin constructs.

    PubMed

    Law, Jia Xian; Musa, Faiza; Ruszymah, Bt Hj Idrus; El Haj, Alicia J; Yang, Ying

    2016-09-01

    Collagen and fibrin are widely used in tissue engineering due to their excellent biocompatibility and bioactivities that support in vivo tissue formation. These two hydrogels naturally present in different wound healing stages with different regulatory effects on cells, and both of them are mechanically weak in the reconstructed hydrogels. We conducted a comparative study by the growth of rat dermal fibroblasts or dermal fibroblasts and epidermal keratinocytes together in collagen and fibrin constructs respectively with and without the reinforcement of electrospun poly(lactic acid) nanofiber mesh. Cell proliferation, gel contraction and elastic modulus of the constructs were measured on the same gels at multiple time points during the 22 day culturing period using multiple non-destructive techniques. The results demonstrated considerably different cellular activities within the two types of constructs. Co-culturing keratinocytes with fibroblasts in the collagen constructs reduced the fibroblast proliferation, collagen contraction and mechanical strength at late culture point regardless of the presence of nanofibers. Co-culturing keratinocytes with fibroblasts in the fibrin constructs promoted fibroblast proliferation but exerted no influence on fibrin contraction and mechanical strength. The presence of nanofibers in the collagen and fibrin constructs played a favorable role on the fibroblast proliferation when keratinocytes were absent. Thus, this study exhibited new evidence of the strong cross-talk between keratinocytes and fibroblasts, which can be used to control fibroblast proliferation and construct contraction. This cross-talk activity is extracellular matrix-dependent in terms of the fibrous network morphology, density and strength. PMID:27349492

  11. Studies on the Interaction between Collagen and a Plasma Kallikrein-Like Activity EVIDENCE FOR A SURFACE-ACTIVE ENZYME SYSTEM

    PubMed Central

    Harpel, Peter C.

    1972-01-01

    This study has demonstrated that collagen particles, after exposure to platelet-poor human plasma and subsequent washing, generate a kinin-like agent when incubated with prekinin substrate. The binding of kinin-generating activity to collagen in the plasma collagen incubation mixture occurs rapidly, whereas the loss of this activity in the incubation mixture occurs relatively slowly. The Hageman factor appeared to be necessary for the surface-bound kinin-generating activity, as this activity was absent in collagen exposed to Hageman factor-deficient plasma. These studies have partially characterized the plasma-derived enzymatic activity bound to collagen. Incubation of collagen with plasma caused a concentration-dependent reduction in the kinin-producing activity which was generated by the addition of ellagic acid, a known activator of plasma kallikrein. The kinin-inducing activity bound to collagen is inhibited by soybean trypsin inhibitor, Trasylol, serum C1̄ inactivator and the plasma α2-macroglobulin, but not by lima bean trypsin inhibitor. An eluate prepared from plasma-treated collagen, when compared with purified plasma kallikrein, shared a similar inhibitor profile. Selective chemical blockage of the free carboxyl groups on the collagen molecule, or heat denaturation, inactivated the ability of the collagen to generate kinin-like activity after incubation with plasma. Removal of the collagen telopeptides or blockage of the free amino groups failed to affect the collagen-plasma interaction. The binding of partially purified plasma kallikrein to collagen was found to have similar structural and chemical requirements. These data indicate that there is a structural and chemical specificity for the activation and binding of plasma kallikrein-like activity by collagen. These studies suggest that a plasma kallikrein may function as a surface-bound enzyme system. PMID:4338122

  12. Contact activation of blood coagulation on a defined kaolin/collagen surface in a microfluidic assay.

    PubMed

    Zhu, Shu; Diamond, Scott L

    2014-12-01

    Generation of active Factor XII (FXIIa) triggers blood clotting on artificial surfaces and may also enhance intravascular thrombosis. We developed a patterned kaolin (0 to 0.3 pg/μm(2))/type 1 collagen fibril surface for controlled microfluidic clotting assays. Perfusion of whole blood (treated only with a low level of 4 μg/mL of the XIIa inhibitor, corn trypsin inhibitor) drove platelet deposition followed by fibrin formation. At venous wall shear rate (100 s(-1)), kaolin accelerated onset of fibrin formation by ~100 sec when compared to collagen alone (250 sec vs. 350 sec), with little effect on platelet deposition. Even with kaolin present, arterial wall shear rate (1000 s(-1)) delayed and suppressed fibrin formation compared to venous wall shear rate. A comparison of surfaces for extrinsic activation (tissue factor TF/collagen) versus contact activation (kaolin/collagen) that each generated equal platelet deposition at 100 s(-1) revealed: (1) TF surfaces promoted much faster fibrin onset (at 100 sec) and more endpoint fibrin at 600 sec at either 100 s(-1) or 1000 s(-1), and (2) kaolin and TF surfaces had a similar sensitivity for reduced fibrin deposition at 1000 s(-1) (compared to fibrin formed at 100 s(-1)) despite differing coagulation triggers. Anti-platelet drugs inhibiting P2Y1, P2Y12, cyclooxygenase-1 or activating IP-receptor or guanylate cyclase reduced platelet and fibrin deposition on kaolin/collagen. Since FXIIa or FXIa inhibition may offer safe antithrombotic therapy, especially for biomaterial thrombosis, these defined collagen/kaolin surfaces may prove useful in drug screening tests or in clinical diagnostic assays of blood under flow conditions.

  13. Contact activation of blood coagulation on a defined kaolin/collagen surface in a microfluidic assay

    PubMed Central

    Zhu, Shu; Diamond, Scott L.

    2014-01-01

    Generation of active Factor XII (FXIIa) triggers blood clotting on artificial surfaces and may also enhance intravascular thrombosis. We developed a patterned kaolin (0 to 0.3 pg/μm2)/type 1 collagen fibril surface for controlled microfluidic clotting assays. Perfusion of whole blood (treated only with a low level of 4 μg/mL of the XIIa inhibitor, corn trypsin inhibitor) drove platelet deposition followed by fibrin formation. At venous wall shear rate (100 s−1), kaolin accelerated onset of fibrin formation by ~100 sec when compared to collagen alone (250 sec vs. 350 sec), with little effect on platelet deposition. Even with kaolin present, arterial wall shear rate (1000 s−1) delayed and suppressed fibrin formation compared to venous wall shear rate. A comparison of surfaces for extrinsic activation (tissue factor TF/collagen) versus contact activation (kaolin/collagen) that each generated equal platelet deposition at 100 s−1 revealed: (1) TF surfaces promoted much faster fibrin onset (at 100 sec) and more endpoint fibrin at 600 sec at either 100 s−1 or 1000 s−1, and (2) kaolin and TF surfaces had a similar sensitivity for reduced fibrin deposition at 1000 s−1 (compared to fibrin formed at 100 s−1) despite differing coagulation triggers. Anti-platelet drugs inhibiting P2Y1, P2Y12, cyclooxygenase-1 or activating IP-receptor or guanylate cyclase reduced platelet and fibrin deposition on kaolin/collagen. Since FXIIa or FXIa inhibition may offer safe antithrombotic therapy, especially for biomaterial thrombosis, these defined collagen/kaolin surfaces may prove useful in drug screening tests or in clinical diagnostic assays of blood under flow conditions. PMID:25303860

  14. Contact activation of blood coagulation on a defined kaolin/collagen surface in a microfluidic assay.

    PubMed

    Zhu, Shu; Diamond, Scott L

    2014-12-01

    Generation of active Factor XII (FXIIa) triggers blood clotting on artificial surfaces and may also enhance intravascular thrombosis. We developed a patterned kaolin (0 to 0.3 pg/μm(2))/type 1 collagen fibril surface for controlled microfluidic clotting assays. Perfusion of whole blood (treated only with a low level of 4 μg/mL of the XIIa inhibitor, corn trypsin inhibitor) drove platelet deposition followed by fibrin formation. At venous wall shear rate (100 s(-1)), kaolin accelerated onset of fibrin formation by ~100 sec when compared to collagen alone (250 sec vs. 350 sec), with little effect on platelet deposition. Even with kaolin present, arterial wall shear rate (1000 s(-1)) delayed and suppressed fibrin formation compared to venous wall shear rate. A comparison of surfaces for extrinsic activation (tissue factor TF/collagen) versus contact activation (kaolin/collagen) that each generated equal platelet deposition at 100 s(-1) revealed: (1) TF surfaces promoted much faster fibrin onset (at 100 sec) and more endpoint fibrin at 600 sec at either 100 s(-1) or 1000 s(-1), and (2) kaolin and TF surfaces had a similar sensitivity for reduced fibrin deposition at 1000 s(-1) (compared to fibrin formed at 100 s(-1)) despite differing coagulation triggers. Anti-platelet drugs inhibiting P2Y1, P2Y12, cyclooxygenase-1 or activating IP-receptor or guanylate cyclase reduced platelet and fibrin deposition on kaolin/collagen. Since FXIIa or FXIa inhibition may offer safe antithrombotic therapy, especially for biomaterial thrombosis, these defined collagen/kaolin surfaces may prove useful in drug screening tests or in clinical diagnostic assays of blood under flow conditions. PMID:25303860

  15. Fibroblast Activation Protein (FAP) Accelerates Collagen Degradation and Clearance from Lungs in Mice.

    PubMed

    Fan, Ming-Hui; Zhu, Qiang; Li, Hui-Hua; Ra, Hyun-Jeong; Majumdar, Sonali; Gulick, Dexter L; Jerome, Jacob A; Madsen, Daniel H; Christofidou-Solomidou, Melpo; Speicher, David W; Bachovchin, William W; Feghali-Bostwick, Carol; Puré, Ellen

    2016-04-01

    Idiopathic pulmonary fibrosis is a disease characterized by progressive, unrelenting lung scarring, with death from respiratory failure within 2-4 years unless lung transplantation is performed. New effective therapies are clearly needed. Fibroblast activation protein (FAP) is a cell surface-associated serine protease up-regulated in the lungs of patients with idiopathic pulmonary fibrosis as well as in wound healing and cancer. We postulate that FAP is not only a marker of disease but influences the development of pulmonary fibrosis after lung injury. In two different models of pulmonary fibrosis, intratracheal bleomycin instillation and thoracic irradiation, we find increased mortality and increased lung fibrosis in FAP-deficient mice compared with wild-type mice. Lung extracellular matrix analysis reveals accumulation of intermediate-sized collagen fragments in FAP-deficient mouse lungs, consistent within vitrostudies showing that FAP mediates ordered proteolytic processing of matrix metalloproteinase (MMP)-derived collagen cleavage products. FAP-mediated collagen processing leads to increased collagen internalization without altering expression of the endocytic collagen receptor, Endo180. Pharmacologic FAP inhibition decreases collagen internalization as expected. Conversely, restoration of FAP expression in the lungs of FAP-deficient mice decreases lung hydroxyproline content after intratracheal bleomycin to levels comparable with that of wild-type controls. Our findings indicate that FAP participates directly, in concert with MMPs, in collagen catabolism and clearance and is an important factor in resolving scar after injury and restoring lung homeostasis. Our study identifies FAP as a novel endogenous regulator of fibrosis and is the first to show FAP's protective effects in the lung.

  16. Loss of fibulin-4 disrupts collagen synthesis and maturation: implications for pathology resulting from EFEMP2 mutations.

    PubMed

    Papke, Christina L; Tsunezumi, Jun; Ringuette, Léa-Jeanne; Nagaoka, Hideaki; Terajima, Masahiko; Yamashiro, Yoshito; Urquhart, Greg; Yamauchi, Mitsuo; Davis, Elaine C; Yanagisawa, Hiromi

    2015-10-15

    Homozygous recessive mutations in either EFEMP2 (encoding fibulin-4) or FBLN5 (encoding fibulin-5), critical genes for elastogenesis, lead to autosomal recessive cutis laxa types 1B and 1A, respectively. Previously, fibulin-4 was shown to bind lysyl oxidase (LOX), an elastin/collagen cross-linking enzyme, in vitro. Consistently, reported defects in humans with EFEMP2 mutations are more severe and broad in range than those due to FBLN5 mutations and encompass both elastin-rich and collagen-rich tissues. However, the underlying disease mechanism in EFEMP2 mutations has not been fully addressed. Here, we show that fibulin-4 is important for the integrity of aortic collagen in addition to elastin. Smooth muscle-specific Efemp2 loss in mouse (termed SMKO) resulted in altered fibrillar collagen localization with larger, poorly organized fibrils. LOX activity was decreased in Efemp2-null cells, and collagen cross-linking was diminished in SMKO aortas; however, elastin cross-linking was unaffected and the level of mature LOX was maintained to that of wild-type aortas. Proteomic screening identified multiple proteins involved in procollagen processing and maturation as potential fibulin-4-binding partners. We showed that fibulin-4 binds procollagen C-endopeptidase enhancer 1 (Pcolce), which enhances proteolytic cleavage of the procollagen C-terminal propeptide during procollagen processing. Interestingly, however, procollagen cleavage was not affected by the presence or absence of fibulin-4 in vitro. Thus, our data indicate that fibulin-4 serves as a potential scaffolding protein during collagen maturation in the extracellular space. Analysis of collagen in other tissues affected by fibulin-4 loss should further increase our understanding of underlying pathologic mechanisms in patients with EFEMP2 mutations.

  17. Na/K-ATPase signaling regulates collagen synthesis through microRNA-29b-3p in cardiac fibroblasts.

    PubMed

    Drummond, Christopher A; Hill, Michael C; Shi, Huilin; Fan, Xiaoming; Xie, Jeffrey X; Haller, Steven T; Kennedy, David J; Liu, Jiang; Garrett, Michael R; Xie, Zijian; Cooper, Christopher J; Shapiro, Joseph I; Tian, Jiang

    2016-03-01

    Chronic kidney disease (CKD) is accompanied by cardiac fibrosis, hypertrophy, and dysfunction, which are commonly referred to as uremic cardiomyopathy. Our previous studies found that Na/K-ATPase ligands or 5/6th partial nephrectomy (PNx) induces cardiac fibrosis in rats and mice. The current study used in vitro and in vivo models to explore novel roles for microRNA in this mechanism of cardiac fibrosis formation. To accomplish this, we performed microRNA profiling with RT-qPCR based arrays on cardiac tissue from rats subjected to marinobufagenin (MBG) infusion or PNx. The analysis showed that a series of fibrosis-related microRNAs were dysregulated. Among the dysregulated microRNAs, microRNA (miR)-29b-3p, which directly targets mRNA of collagen, was consistently reduced in both PNx and MBG-infused animals. In vitro experiments demonstrated that treatment of primary cultures of adult rat cardiac fibroblasts with Na/K-ATPase ligands induced significant increases in the fibrosis marker, collagen protein, and mRNA expression compared with controls, whereas miR-29b-3p expression decreased >50%. Transfection of miR-29b-3p mimics into cardiac fibroblasts inhibited cardiotonic steroids-induced collagen synthesis. Moreover, a specific Na/K-ATPase signaling antagonist, pNaKtide, prevented ouabain-induced increases in collagen synthesis and decreases in miR-29b-3p expression in these cells. In conclusion, these data are the first to indicate that signaling through Na/K-ATPase regulates miRNAs and specifically, miR-29b-3p expression both in vivo and in vitro. Additionally, these data indicate that miR-29b-3p expression plays an important role in the formation of cardiac fibrosis in CKD. PMID:26702050

  18. Cloning, expression and antioxidant activity of a novel collagen from Pelodiscus sinensis.

    PubMed

    Xu, Ran; Li, Dengfeng; Peng, Jiao; Fang, Jing; Zhang, Liping; Liu, Lianguo

    2016-06-01

    Collagen is the main structural protein of various connective tissues in animals and naturally plays an important role within the body. It is increasingly used within certain areas, such as medicine, citology and cosmetology. The soft-shelled turtle (Pelodiscus sinensis) is a commercially important aquatic species rich in collagen. In this study, a novel collagen gene fragment of 756 bp, which encodes 252 deduced amino acid residues, including 25 conserved Gly-X-Y motifs, was cloned from a soft-shelled turtle. Recombinant soft-shelled turtle collagen (rSTC) was stably expressed in Escherichia coli Rosetta and purified by His GraviTrap affinity columns. The antioxidant activities of rSTC were measured using hydroxyl and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals. The results showed that rSTC quenched the free radicals in a dose-dependent manner. The hydroxyl radical scavenging activity (HRSA) of rSTC was 98.9 % at a concentration of 3 mg/mL. At a concentration of 5 mg/mL, rSTC exhibited a DPPH radical scavenging activity of 32.7 %. At the tested concentrations, rSTC exhibited higher HRSA and lower DPPH radical scavenging activity. PMID:27116966

  19. Collagen Homeostasis and Metabolism.

    PubMed

    Magnusson, S Peter; Heinemeier, Katja M; Kjaer, Michael

    2016-01-01

    The musculoskeletal system and its collagen rich tissue is important for ensuring architecture of skeletal muscle, energy storage in tendon and ligaments, joint surface protection, and for ensuring the transfer of muscular forces into resulting limb movement. Structure of tendon is stable and the metabolic activity is low, but mechanical loading and subsequent mechanotransduction and molecular anabolic signaling can result in some adaptation of the tendon especially during youth and adolescence. Within short time, tendon will get stiffer with training and lack of mechanical tissue loading through inactivity or immobilization of the human body will conversely result in a dramatic loss in tendon stiffness and collagen synthesis. This illustrates the importance of regular mechanical load in order to preserve the stabilizing role of the connective tissue for the overall function of the musculoskeletal system in both daily activity and exercise. Adaptive responses may vary along the tendon, and differ between mid-substance and insertional areas of the tendon. PMID:27535245

  20. Arterial Tortuosity Syndrome reveals function of dehydroascorbic acid in collagen and elastin synthesis: Implications for skin care.

    PubMed

    Kitt, Douglas Q

    2016-02-01

    Some investigations in Arterial Tortuosity Syndrome (ATS) suggest that impaired intracellular transport of the oxidized form of vitamin C (dehydroascorbic acid, DHAA) is at the core of the pathogenesis. Lack of vitamin C for lysyl- and prolyl-hydroxylase activity may explain the defects in collagen and elastin formation found in ATS, and draws strong parallels between ATS and scurvy. Topically applied vitamin C has a well-established basis in the field of skin care, and part of its benefit is attributed to proper collagen formation in the skin. The ATS studies suggest that DHAA transport is necessary for normal skin collagen formation, and this has implications as to the forms of vitamin C best-suited for topical skin care. PMID:26826631

  1. Overexpression of collagen XIII in extraocular fat affected by active thyroid-associated ophthalmopathy: A crucial piece of the puzzle?

    PubMed

    Morris, Olivia Claire; Schebitz Walter, Kirsten; Telemo, Esbjörn; Hintschich, Christoph

    2016-08-01

    Thyroid-associated ophthalmopathy (TAO) causes irreversible increase in extraocular fat volume that contributes to the risk of exophthalmos and compressive optic neuropathy. Collagen XIII is implicated in uncontrolled cell growth in some tumours, but we are not aware of any studies of collagen XIII in TAO-affected solid tissue to date. We conducted immunohistochemical staining for collagen XIII alpha 1 (COL13A1), present in both the transmembrane and cleaved forms of collagen XIII, in consecutive prospectively collected human extraocular tissue specimens from patients with TAO and controls. We identified overexpression of collagen XIII in active TAO-affected fat. We discuss how species and cell-type specific responses of collagen XIII to stressors may help explain the different phenotypes of TAO. PMID:27245701

  2. Cathepsin B Regulates Collagen Expression by Fibroblasts via Prolonging TLR2/NF-κB Activation

    PubMed Central

    Li, Xue; Ni, Junjun; Meng, Jie; Yu, Weixian; Nakanishi, Hiroshi

    2016-01-01

    Fibroblasts are essential for tissue repair due to producing collagens, and lysosomal proteinase cathepsin B (CatB) is involved in promoting chronic inflammation. We herein report that CatB regulates the expression of collagens III and IV by fibroblasts in response to a TLR2 agonist, lipopolysaccharide from Porphyromonas gingivalis (P.g. LPS). In cultured human BJ fibroblasts, mRNA expression of CatB was significantly increased, while that of collagens III and IV was significantly decreased at 24 h after challenge with P.g. LPS (1 μg/mL). The P.g. LPS-decreased collagen expression was completely inhibited by CA-074Me, the specific inhibitor of CatB. Surprisingly, expression of collagens III and IV was significantly increased in the primary fibroblasts from CatB-deficient mice after challenge with P.g. LPS. The increase of CatB was accompanied with an increase of 8-hydroxy-2′-deoxyguanosine (8-OHdG) and a decrease of IκBα. Furthermore, the P.g. LPS-increased 8-OHdG and decreased IκBα were restored by CA-074Me. Moreover, 87% of CatB and 86% of 8-OHdG were colocalized with gingival fibroblasts of chronic periodontitis patients. The findings indicate the critical role of CatB in regulating the expression of collagens III and IV by fibroblasts via prolonging TLR2/NF-κB activation and oxidative stress. CatB-specific inhibitors may therefore improve chronic inflammation-delayed tissue repair. PMID:27648120

  3. Cathepsin B Regulates Collagen Expression by Fibroblasts via Prolonging TLR2/NF-κB Activation

    PubMed Central

    Li, Xue; Ni, Junjun; Meng, Jie; Yu, Weixian; Nakanishi, Hiroshi

    2016-01-01

    Fibroblasts are essential for tissue repair due to producing collagens, and lysosomal proteinase cathepsin B (CatB) is involved in promoting chronic inflammation. We herein report that CatB regulates the expression of collagens III and IV by fibroblasts in response to a TLR2 agonist, lipopolysaccharide from Porphyromonas gingivalis (P.g. LPS). In cultured human BJ fibroblasts, mRNA expression of CatB was significantly increased, while that of collagens III and IV was significantly decreased at 24 h after challenge with P.g. LPS (1 μg/mL). The P.g. LPS-decreased collagen expression was completely inhibited by CA-074Me, the specific inhibitor of CatB. Surprisingly, expression of collagens III and IV was significantly increased in the primary fibroblasts from CatB-deficient mice after challenge with P.g. LPS. The increase of CatB was accompanied with an increase of 8-hydroxy-2′-deoxyguanosine (8-OHdG) and a decrease of IκBα. Furthermore, the P.g. LPS-increased 8-OHdG and decreased IκBα were restored by CA-074Me. Moreover, 87% of CatB and 86% of 8-OHdG were colocalized with gingival fibroblasts of chronic periodontitis patients. The findings indicate the critical role of CatB in regulating the expression of collagens III and IV by fibroblasts via prolonging TLR2/NF-κB activation and oxidative stress. CatB-specific inhibitors may therefore improve chronic inflammation-delayed tissue repair.

  4. Cathepsin B Regulates Collagen Expression by Fibroblasts via Prolonging TLR2/NF-κB Activation.

    PubMed

    Li, Xue; Wu, Zhou; Ni, Junjun; Liu, Yicong; Meng, Jie; Yu, Weixian; Nakanishi, Hiroshi; Zhou, Yanmin

    2016-01-01

    Fibroblasts are essential for tissue repair due to producing collagens, and lysosomal proteinase cathepsin B (CatB) is involved in promoting chronic inflammation. We herein report that CatB regulates the expression of collagens III and IV by fibroblasts in response to a TLR2 agonist, lipopolysaccharide from Porphyromonas gingivalis (P.g. LPS). In cultured human BJ fibroblasts, mRNA expression of CatB was significantly increased, while that of collagens III and IV was significantly decreased at 24 h after challenge with P.g. LPS (1 μg/mL). The P.g. LPS-decreased collagen expression was completely inhibited by CA-074Me, the specific inhibitor of CatB. Surprisingly, expression of collagens III and IV was significantly increased in the primary fibroblasts from CatB-deficient mice after challenge with P.g. LPS. The increase of CatB was accompanied with an increase of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and a decrease of IκBα. Furthermore, the P.g. LPS-increased 8-OHdG and decreased IκBα were restored by CA-074Me. Moreover, 87% of CatB and 86% of 8-OHdG were colocalized with gingival fibroblasts of chronic periodontitis patients. The findings indicate the critical role of CatB in regulating the expression of collagens III and IV by fibroblasts via prolonging TLR2/NF-κB activation and oxidative stress. CatB-specific inhibitors may therefore improve chronic inflammation-delayed tissue repair. PMID:27648120

  5. Effects of collagen deposition on passive and active mechanical properties of large pulmonary arteries in hypoxic pulmonary hypertension.

    PubMed

    Wang, Zhijie; Lakes, Roderic S; Eickhoff, Jens C; Chesler, Naomi C

    2013-11-01

    Proximal pulmonary artery (PA) stiffening is a strong predictor of mortality in pulmonary hypertension. Collagen accumulation is mainly responsible for PA stiffening in hypoxia-induced pulmonary hypertension (HPH) in mouse models. We hypothesized that collagen cross-linking and the type I isoform are the main determinants of large PA mechanical changes during HPH, which we tested by exposing mice that resist type I collagen degradation (Col1a1[Formula: see text] and littermate controls (Col1a1[Formula: see text] to hypoxia for 10 days with or without [Formula: see text]-aminopropionitrile (BAPN) treatment to prevent cross-link formation. Static and dynamic mechanical tests were performed on isolated PAs with smooth muscle cells (SMC) in passive and active states. Percentages of type I and III collagen were quantified by histology; total collagen content and cross-linking were measured biochemically. In the SMC passive state, for both genotypes, hypoxia tended to increase PA stiffness and damping capacity, and BAPN treatment limited these increases. These changes were correlated with collagen cross-linking ([Formula: see text]). In the SMC active state, hypoxia increased PA dynamic stiffness and BAPN had no effect in Col1a1[Formula: see text] mice ([Formula: see text]). PA stiffness did not change in Col1a1[Formula: see text] mice. Similarly, damping capacity did not change for either genotype. Type I collagen accumulated more in Col1a1[Formula: see text] mice, whereas type III collagen increased more in Col1a1[Formula: see text] mice during HPH. In summary, PA passive mechanical properties (both static and dynamic) are related to collagen cross-linking. Type I collagen turnover is critical to large PA remodeling during HPH when collagen metabolism is not mutated and type III collagen may serve as a reserve. PMID:23377784

  6. Collagen degradation and MMP9 activation by Enterococcus faecalis contributes to intestinal anastomotic leak

    PubMed Central

    Shogan, B. D.; Belogortseva, N.; Luong, P. M.; Zaborin, A.; Lax, S.; Bethel, Cindy; Ward, M.; Muldoon, J. P.; Singer, M.; An, G.; Umanskiy, K.; Konda, V.; Shakhsheer, B.; Luo, J.; Klabbers, R.; Hancock, L. E.; Gilbert, J.; Zaborina, O.; Alverdy, J. C.

    2016-01-01

    Even under the most expert care, a properly constructed intestinal anastomosis can fail to heal resulting in leakage of its contents, peritonitis and sepsis. The cause of anastomotic leak remains unknown and its incidence has not changed in decades. Here, we demonstrate that the commensal bacterium Enterococcus faecalis contributes to the pathogenesis of anastomotic leak through its capacity to degrade collagen and to activate tissue matrix metalloprotease-9 (MMP9) in host intestinal tissues. We demonstrate in rats that leaking anastomotic tissues were colonized by E. faecalis strains that showed an increased collagen-degrading activity and also an increased ability to activate host MMP9, both of which contributed to anastomotic leakage. We demonstrate that the E. faecalis genes gelE and sprE were required for E. faecalis-mediated MMP9 activation. Either elimination of E. faecalis strains through direct topical antibiotics applied to rat intestinal tissues or pharmacological suppression of intestinal MMP9 activation prevented anastomotic leak in rats. In contrast, the standard recommended intravenous antibiotics used in patients undergoing colorectal surgery did not eliminate E. faecalis at anastomotic tissues nor did they prevent leak in our rat model. Finally, we show in humans undergoing colon surgery and treated with the standard recommended intravenous antibiotics, that their anastomotic tissues still contained E. faecalis and other bacterial strains with collagen-degrading/MMP9 activity. We suggest that intestinal microbes with the capacity to produce collagenases and to activate host metalloproteinase MMP9 may break down collagen in the gut tissue contributing to anastomotic leak. PMID:25947163

  7. Suboptimal Activation of Protease-activated Receptors Enhances α2β1 Integrin-mediated Platelet Adhesion to Collagen*

    PubMed Central

    Marjoram, Robin J.; Voss, Bryan; Pan, Yumei; Dickeson, S. Kent; Zutter, Mary M.; Hamm, Heidi E.; Santoro, Samuel A.

    2009-01-01

    Thrombin and fibrillar collagen are potent activators of platelets at sites of vascular injury. Both agonists cause platelet shape change, granule secretion, and aggregation to form the primary hemostatic plug. Human platelets express two thrombin receptors, protease-activated receptors 1 and 4 (PAR1 and PAR4) and two collagen receptors, the α2β1 integrin (α2β1) and the glycoprotein VI (GPVI)/FcRγ chain complex. Although these receptors and their signaling mechanisms have been intensely studied, it is not known whether and how these receptors cooperate in the hemostatic function of platelets. This study examined cooperation between the thrombin and collagen receptors in platelet adhesion by utilizing a collagen-related peptide (α2-CRP) containing the α2β1-specific binding motif, GFOGER, in conjunction with PAR-activating peptides. We demonstrate that platelet adhesion to α2-CRP is substantially enhanced by suboptimal PAR activation (agonist concentrations that do not stimulate platelet aggregation) using the PAR4 agonist peptide and thrombin. The enhanced adhesion induced by suboptimal PAR4 activation was α2β1-dependent and GPVI/FcRγ-independent as revealed in experiments with α2β1- or FcRγ-deficient mouse platelets. We further show that suboptimal activation of other platelet Gq-linked G protein-coupled receptors (GPCRs) produces enhanced platelet adhesion to α2-CRP. The enhanced α2β1-mediated platelet adhesion is controlled by phospholipase C (PLC), but is not dependent on granule secretion, activation of αIIbβ3 integrin, or on phosphoinositol-3 kinase (PI3K) activity. In conclusion, we demonstrate a platelet priming mechanism initiated by suboptimal activation of PAR4 or other platelet Gq-linked GPCRs through a PLC-dependent signaling cascade that promotes enhanced α2β1 binding to collagens containing GFOGER sites. PMID:19815553

  8. Differential effect of water-soluble chitin on collagen synthesis of human bone marrow stem cells and human periodontal ligament stem cells.

    PubMed

    Park, So-Yon; Park, Jung-Chul; Kim, Min-Soo; Lee, Sung-Eun; Kim, Ki-Joon; Jung, Byung-Joo; Park, Wonse; Jeon, Dong-Won; Cho, Kyoo-Sung; Kim, Chang-Sung

    2015-02-01

    Human bone marrow stem cells (hBMSCs) represent a promising regenerative material because of their mutipotency, including their ability to regenerate collagenous soft tissues. We previously found that water-soluble chitin (WSC) enhances the ability of human periodontal ligament stem cells (hPDLSCs) to synthesize collagen tissue. The aim of this study was to determine the effects of WSC on hBMSCs and hPDLSCs for the collagen synthesis both in vitro and in vivo. hBMSCs and hPDLSCs were isolated and expanded with or without 0.3 mg/mL WSC. A series of in vitro and in vivo analyses were performed to evaluate their characteristics as stem cell populations. Then, collagen and hydroxyproline assays were conducted using both in vitro and in vivo assay models, and the real-time polymerase chain reaction was performed to analyze the expression of collagen-related markers. WSC-treated and nontreated hBMSCs and hPDLSCs were transplanted into immunocompromised mice, and histology and immunohistochemistry analyses were conducted after 8 weeks. The in vitro results showed that those cells possessed the characteristics of mesenchymal stem cells. The amount of soluble collagen synthesized was significantly greater in WSC-treated hBMSCs than in the nontreated group; conversely, treatment of hPDLSCs with WSC decreased the formation of soluble collagen. The amount of insoluble collagen synthesized was greater in the WSC-treated groups than in the nontreated groups for both hBMSCs and hPDLSCs. The hydroxyproline contents of the regenerated soluble and insoluble collagens were similar. The expressions of mRNA for collagen types I-V, hyaluronic acid synthase 1 (HAS1), HAS2, and HAS3, and the LOX family were higher in WSC-treated hPDLSCs than in the nontreated group, whereas WSC increased the expression of collagen type III and decreased that of collagen type I in hBMSCs. The histology and immunohistochemistry results revealed that WSC significantly increased the amount of collagen

  9. Arf6 plays an early role in platelet activation by collagen and convulxin.

    PubMed

    Choi, Wangsun; Karim, Zubair A; Whiteheart, Sidney W

    2006-04-15

    Small GTPases play critical roles in hemostasis, though the roster of such molecules in platelets is not complete. In this study, we report the presence of Ras-related GTPases of the ADP-ribosylation factor (Arf) family. Platelets contain Arf1 or 3 and Arf6, with the latter being predominantly membrane associated. Using effector domain pull-down assays, we show, counter to other GTPases, that Arf6-GTP is present in resting platelets and decreases rapidly upon activation with collagen or convulxin. This decrease does not completely rely on secondary agonists (ADP and thromboxane A2) or require integrin signaling. The decrease in free Arf6-GTP temporally precedes activation of Rho family GTPases (RhoA, Cdc42, and Rac1). Using a membrane-permeant, myristoylated peptide, which mimics the N-terminus of Arf6, we show that the Arf6-GTP decrease is essential for collagen- and convulxin-induced aggregation, platelet adherence, and spreading on collagen-coated glass. Treatment with this peptide also affects the activation of Rho family GTPases, but has little effect on RalA and Rap1 or on agonist-induced calcium mobilization. These data show that Arf6 is a key element in activation through GPVI, and is required for activation of the Rho family GTPases and the subsequent cytoskeletal rearrangements needed for full platelet function. PMID:16352809

  10. Water-filtered near-infrared influences collagen synthesis of keloid-fibroblasts in contrast to normal foreskin fibroblasts.

    PubMed

    Zöller, Nadja; König, Anke; Butting, Manuel; Kaufmann, Roland; Bernd, August; Valesky, Eva; Kippenberger, Stefan

    2016-10-01

    Hypertrophic scar development is associated to impaired wound healing, imbalanced fibroblast proliferation and extracellular matrix synthesis. Stigmatization, physical restrictions and high recurrence rates are only some aspects that illustrate the severe influence impaired wound healing can have on patients' life. The treatment of hypertrophic scars especially keloids is still a challenge. In recent years water-filtered near-infrared irradiation (wIRA) composed of near-infrared (NIR) and a thermal component is applied for an increasing penal of clinical purposes. It is described to beneficially influence e.g. wound healing. But discrimination between the thermal and the NIR dependent components of these effects has not been conclusively elucidated. Aim of our study was therefore to investigate the influence of the light fraction on the thermal impact of wIRA irradiation in dermal cells. We concentrated our analysis on morphological properties and collagen synthesis. Foreskin fibroblasts and the keloid fibroblast cell line KF111 were exposed to temperatures between 37°C and 46°C with or without additional irradiation with 360J/cm(2) NIR. Our results show that viability was not influenced by irradiation. Independent of the analysed fibroblast species temperature dependent occurrence of spheric cells could be observed. These morphological changes were clearly counteracted by additional light exposure. Convective heat reduced collagen type I synthesis in both cell species depending on the applied temperature. Co-treatment with NIR significantly reversed this effect in keloid fibroblast cultures treated at 46°C whereas no difference could be observed in the foreskin fibroblasts. The observed influence on collagen type I synthesis was associated to a temperature dependent TGF-β1 secretion reduction. Co-stimulation of keloid cultures with NIR at 46°C completely abolished the temperature dependent TGF-β1 secretion reduction. In foreskin fibroblast cultures co

  11. Water-filtered near-infrared influences collagen synthesis of keloid-fibroblasts in contrast to normal foreskin fibroblasts.

    PubMed

    Zöller, Nadja; König, Anke; Butting, Manuel; Kaufmann, Roland; Bernd, August; Valesky, Eva; Kippenberger, Stefan

    2016-10-01

    Hypertrophic scar development is associated to impaired wound healing, imbalanced fibroblast proliferation and extracellular matrix synthesis. Stigmatization, physical restrictions and high recurrence rates are only some aspects that illustrate the severe influence impaired wound healing can have on patients' life. The treatment of hypertrophic scars especially keloids is still a challenge. In recent years water-filtered near-infrared irradiation (wIRA) composed of near-infrared (NIR) and a thermal component is applied for an increasing penal of clinical purposes. It is described to beneficially influence e.g. wound healing. But discrimination between the thermal and the NIR dependent components of these effects has not been conclusively elucidated. Aim of our study was therefore to investigate the influence of the light fraction on the thermal impact of wIRA irradiation in dermal cells. We concentrated our analysis on morphological properties and collagen synthesis. Foreskin fibroblasts and the keloid fibroblast cell line KF111 were exposed to temperatures between 37°C and 46°C with or without additional irradiation with 360J/cm(2) NIR. Our results show that viability was not influenced by irradiation. Independent of the analysed fibroblast species temperature dependent occurrence of spheric cells could be observed. These morphological changes were clearly counteracted by additional light exposure. Convective heat reduced collagen type I synthesis in both cell species depending on the applied temperature. Co-treatment with NIR significantly reversed this effect in keloid fibroblast cultures treated at 46°C whereas no difference could be observed in the foreskin fibroblasts. The observed influence on collagen type I synthesis was associated to a temperature dependent TGF-β1 secretion reduction. Co-stimulation of keloid cultures with NIR at 46°C completely abolished the temperature dependent TGF-β1 secretion reduction. In foreskin fibroblast cultures co

  12. Colloidal Gold--Collagen Protein Core--Shell Nanoconjugate: One-Step Biomimetic Synthesis, Layer-by-Layer Assembled Film, and Controlled Cell Growth.

    PubMed

    Xing, Ruirui; Jiao, Tifeng; Yan, Linyin; Ma, Guanghui; Liu, Lei; Dai, Luru; Li, Junbai; Möhwald, Helmuth; Yan, Xuehai

    2015-11-11

    The biogenic synthesis of biomolecule-gold nanoconjugates is of key importance for a broad range of biomedical applications. In this work, a one-step, green, and condition-gentle strategy is presented to synthesize stable colloidal gold-collagen core-shell nanoconjugates in an aqueous solution at room temperature, without use of any reducing agents and stabilizing agents. It is discovered that electrostatic binding between gold ions and collagen proteins and concomitant in situ reduction by hydroxyproline residues are critically responsible for the formation of the core-shell nanoconjugates. The film formed by layer-by-layer assembly of such colloidal gold-collagen nanoconjugates can notably improve the mechanical properties and promote cell adhesion, growth, and differentiation. Thus, the colloidal gold-collagen nanoconjugates synthesized by such a straightforward and clean manner, analogous to a biomineralization pathway, provide new alternatives for developing biologically based hybrid biomaterials toward a range of therapeutic and diagnostic applications.

  13. Mechanically improved electrospun PCL biocomposites reinforced with a collagen coating process: preparation, physical properties, and cellular activity.

    PubMed

    Hong, Soon Gee; Kim, Geun Hyung

    2013-02-01

    In this work, we fabricated highly aligned electrospun poly(ε-caprolactone)(PCL)/collagen biocomposites, which were consisted of multi-layered structure. The aligned directions of the composites were controlled with two rotating collectors, and various weight fractions (1, 2, 3 wt%) of collagen were embedded between the mat of PCL microfibers to improve the mechanical property and biological activities of osteoblast-like cells (MG63). The PCL/collagen biocomposite showed nine times of increment in mechanical strength of random PCL/collagen composite. An increase in collagen content in the biocomposites displayed significant increase of mechanical properties, hydrophilic property, water-absorption ability, and even cell viability of osteoblast-like cells (MG63). PMID:22763777

  14. Icaritin Inhibits Collagen Degradation-Related Factors and Facilitates Collagen Accumulation in Atherosclerotic Lesions: A Potential Action for Plaque Stabilization.

    PubMed

    Zhang, Zong-Kang; Li, Jie; Yan, De-Xin; Leung, Wing-Nang; Zhang, Bao-Ting

    2016-01-28

    Most acute coronary syndromes result from rupture of vulnerable atherosclerotic plaques. The collagen content of plaques may critically affect plaque stability. This study tested whether Icaritin (ICT), an intestinal metabolite of Epimedium-derived flavonoids, could alter the collagen synthesis/degradation balance in atherosclerotic lesions. Rabbits were fed with an atherogenic diet for four months. Oral administration of ICT (10 mg·kg(-1)·day(-1)) was started after two months of an atherogenic diet and lasted for two months. The collagen degradation-related parameters, including macrophages accumulation, content and activity of interstitial collagenase-1 (MMP-1), and the collagen synthesis-related parameters, including amount and distribution of smooth muscle cells (SMC) and collagen mRNA/protein levels, were evaluated in the aorta. ICT reduced plasma lipid levels, inhibited macrophage accumulation, lowered MMP-1 mRNA and protein expression, and suppressed proteolytic activity of pro-MMP-1 and MMP-1 in the aorta. ICT changed the distribution of the SMCs towards the fibrous cap of lesions without increasing the amount of SMCs. Higher collagen protein content in lesions and aorta homogenates was observed with ICT treatment compared with the atherogenic diet only, without altered collagen mRNA level. These results suggest that ICT could inhibit the collagen degradation-related factors and facilitate collagen accumulation in atherosclerotic lesions, indicating a new potential of ICT in atherosclerotic plaques.

  15. Glucose and collagen regulate human platelet activity through aldose reductase induction of thromboxane.

    PubMed

    Tang, Wai Ho; Stitham, Jeremiah; Gleim, Scott; Di Febbo, Concetta; Porreca, Ettore; Fava, Cristiano; Tacconelli, Stefania; Capone, Marta; Evangelista, Virgilio; Levantesi, Giacomo; Wen, Li; Martin, Kathleen; Minuz, Pietro; Rade, Jeffrey; Patrignani, Paola; Hwa, John

    2011-11-01

    Diabetes mellitus is associated with platelet hyperactivity, which leads to increased morbidity and mortality from cardiovascular disease. This is coupled with enhanced levels of thromboxane (TX), an eicosanoid that facilitates platelet aggregation. Although intensely studied, the mechanism underlying the relationship among hyperglycemia, TX generation, and platelet hyperactivity remains unclear. We sought to identify key signaling components that connect high levels of glucose to TX generation and to examine their clinical relevance. In human platelets, aldose reductase synergistically modulated platelet response to both hyperglycemia and collagen exposure through a pathway involving ROS/PLCγ2/PKC/p38α MAPK. In clinical patients with platelet activation (deep vein thrombosis; saphenous vein graft occlusion after coronary bypass surgery), and particularly those with diabetes, urinary levels of a major enzymatic metabolite of TX (11-dehydro-TXB2 [TX-M]) were substantially increased. Elevated TX-M persisted in diabetic patients taking low-dose aspirin (acetylsalicylic acid, ASA), suggesting that such patients may have underlying endothelial damage, collagen exposure, and thrombovascular disease. Thus, our study has identified multiple potential signaling targets for designing combination chemotherapies that could inhibit the synergistic activation of platelets by hyperglycemia and collagen exposure. PMID:22005299

  16. Role for high-glucose-induced protein O-GlcNAcylation in stimulating cardiac fibroblast collagen synthesis

    PubMed Central

    Aguilar, Hugo; Fricovsky, Eduardo; Ihm, Sang; Schimke, Magdalena; Maya-Ramos, Lisandro; Aroonsakool, Nakon; Ceballos, Guillermo; Dillmann, Wolfgang; Villarreal, Francisco

    2014-01-01

    Excess enzyme-mediated protein O-GlcNAcylation is known to occur with diabetes mellitus. A characteristic of diabetic cardiomyopathy is the development of myocardial fibrosis. The role that enhanced protein O-GlcNAcylation plays in modulating the phenotype of cardiac fibroblasts (CF) is unknown. To address this issue, rat CF were cultured in normal glucose (NG; 5 mM glucose) or high-glucose (HG; 25 mM) media for 48 h. Results demonstrate that CF cultured in HG have higher levels (∼50%) of overall protein O-GlcNAcylation vs. NG cells. Key regulators of collagen synthesis such as transforming-growth factor-β1 (TGF-β1), SMADs 2/3, and SMAD 7 protein levels, including those of arginase I and II, were altered, leading to increases in collagen levels. The nuclear transcription factor Sp1 and arginase II evidence excess O-GlcNAcylation in HG cells. Expression in CF of an adenovirus coding for the enzyme N-acetylglucosaminidase, which removes O-GlcNAc moieties from proteins, decreased Sp1 and arginase II O-GlcNAcylation and restored HG-induced perturbations in CF back to NG levels. These findings may have important pathophysiological implications for the development of diabetes-induced cardiac fibrosis. PMID:24553187

  17. Synthesis of highly interconnected 3D scaffold from Arothron stellatus skin collagen for tissue engineering application.

    PubMed

    Ramanathan, Giriprasath; Singaravelu, Sivakumar; Raja, M D; Sivagnanam, Uma Tiruchirapalli

    2015-11-01

    The substrate which is avidly used for tissue engineering applications should have good mechanical and biocompatible properties, and all these parameters are often considered as essential for dermal reformation. Highly interconnected three dimensional (3D) wound dressing material with enhanced structural integrity was synthesized from Arothron stellatus fish skin (AsFS) collagen for tissue engineering applications. The synthesized 3D collagen sponge (COL-SPG) was further characterized by different physicochemical methods. The scanning electron microscopy analysis of the material demonstrated that well interconnected pores with homogeneous microstructure on the surface aids higher swelling index and that the material also possessed good mechanical properties with a Young's modulus of 0.89±0.2 MPa. Biocompatibility of the 3D COL-SPG showed 92% growth for both NIH 3T3 fibroblasts and keratinocytes. Overall, the study revealed that synthesized 3D COL-SPG from fish skin will act as a promising wound dressing in skin tissue engineering.

  18. Poly-L-arginine based materials as instructive substrates for fibroblast synthesis of collagen.

    PubMed

    Bygd, Hannah C; Akilbekova, Dana; Muñoz, Adam; Forsmark, Kiva D; Bratlie, Kaitlin M

    2015-09-01

    The interactions of cells and surrounding tissues with biomaterials used in tissue engineering, wound healing, and artificial organs ultimately determine their fate in vivo. We have demonstrated the ability to tune fibroblast responses with the use of varied material chemistries. In particular, we examined cell morphology, cytokine production, and collagen fiber deposition angles in response to a library of arginine-based polymeric materials. The data presented here shows a large range of vascular endothelial growth factor (VEGF) secretion (0.637 ng/10(6) cells/day to 3.25 ng/10(6) cells/day), cell migration (∼15 min < persistence time < 120 min, 0.11 μm/min < speed < 0.23 μm/min), and cell morphology (0.039 < form factor (FF) < 0.107). Collagen orientation, quantified by shape descriptor (D) values that ranges from 0 to 1, representing completely random (D = 0) to aligned (D = 1) fibers, exhibited large variation both in vitro and in vivo (0.167 < D < 0.36 and 0.17 < D < 0.52, respectively). These findings demonstrate the ability to exert a certain level of control over cellular responses with biomaterials and the potential to attain a desired cellular response such as, increased VEGF production or isotropic collagen deposition upon exposure to these materials in wound healing and tissue engineering applications.

  19. Delphinidin prevents high glucose-induced cell proliferation and collagen synthesis by inhibition of NOX-1 and mitochondrial superoxide in mesangial cells.

    PubMed

    Song, Seung Eun; Jo, Hye Jun; Kim, Yong-Woon; Cho, Young-Je; Kim, Jae-Ryong; Park, So-Young

    2016-04-01

    This study examined the effect of delphinidin on high glucose-induced cell proliferation and collagen synthesis in mesangial cells. Glucose dose-dependently (5.6-25 mM) increased cell proliferation and collagen I and IV mRNA levels, whereas pretreatment with delphinidin (50 μM) prevented cell proliferation and the increased collagen mRNA levels induced by high glucose (25 mM). High glucose increased reactive oxygen species (ROS) generation, and this was suppressed by pretreating delphinidin or the antioxidant N-acetyl cysteine. NADPH oxidase (NOX) 1 was upregulated by high glucose, but pretreatment with delphinidin abrogated this upregulation. Increased mitochondrial superoxide by 25 mM glucose was also suppressed by delphinidin. The NOX inhibitor apocynin and mitochondria-targeted antioxidant Mito TEMPO inhibited ROS generation and cell proliferation induced by high glucose. Phosphorylation of extracellular signal regulated kinase (ERK)1/2 was increased by high glucose, which was suppressed by delphinidin, apocynin or Mito TEMPO. Furthermore, PD98059 (an ERK1/2 inhibitor) prevented the high glucose-induced cell proliferation and increased collagen mRNA levels. Transforming growth factor (TGF)-β protein levels were elevated by high glucose, and pretreatment with delphinidin or PD98059 prevented this augmentation. These results suggest that delphinidin prevents high glucose-induced cell proliferation and collagen synthesis by inhibition of NOX-1 and mitochondrial superoxide in mesangial cells.

  20. Effects of integrin ανβ3 on differentiation and collagen synthesis induced by connective tissue growth factor in human hypertrophic scar fibroblasts.

    PubMed

    Hu, Xiaolong; Li, Na; Tao, Ke; Fang, Xiaobing; Liu, Jiaqi; Wang, Yaojun; Wang, Hongtao; Shi, Jihong; Wang, Yunchuan; Ji, Peng; Cai, Weixia; Bai, Xiaozhi; Zhu, Xiongxiang; Han, Juntao; Hu, Dahai

    2014-11-01

    CCN2 is a matricellular protein that appears to be important in scar formation. CCN2 mediates the pro-fibrotic effects in hypertrophic scars (HTSs) through an unknown mechanism. However, many activities of CCN2 protein are known to be mediated by direct binding to integrin receptors. In this study, we investigated the role of integrin α(ν)β(3) in the differentiation of hypertrophic scar fibroblasts (HTSFs) induced by CCN2. The levels of integrin α(ν)β(3) between normal skin and hypertrophic scar (HTS) tissues were compared, and integrin α(ν)β(3) was found to be upregulated in HTS. CCN2 was shown to induce HTSF differentiation and collagen (COL) synthesis at the mRNA and protein levels. Based on these results, the expression of integrin α(ν)β(3) was upregulated by CCN2 stimulation during HTSF differentiation. Blockade of integrin α(ν)β(3) prevented CCN2-induced HTSF differentiation and COL synthesis. Furthermore, the CCN2-induced increase in contractility of the HTSF in COL lattices was inhibited by integrin α(ν)β(3) blocking antibodies. HTSs were established in a rabbit ear model, and the inhibitor of integrin α(ν)β(3) significantly improved the architecture of the rabbit ear scar. Results of the present study showed that integrin α(ν)β(3) contributes to pro-fibrotic CCN2 signaling. Blocking this pathway may therefore be beneficial for the treatment of HTS.

  1. Absence of the ER Cation Channel TMEM38B/TRIC-B Disrupts Intracellular Calcium Homeostasis and Dysregulates Collagen Synthesis in Recessive Osteogenesis Imperfecta

    PubMed Central

    Cabral, Wayne A.; Ishikawa, Masaki; Garten, Matthias; Makareeva, Elena N.; Sargent, Brandi M.; Weis, MaryAnn; Barnes, Aileen M.; Webb, Emma A.; Shaw, Nicholas J.; Ala-Kokko, Leena; Lacbawan, Felicitas L.; Högler, Wolfgang; Leikin, Sergey; Blank, Paul S.; Zimmerberg, Joshua; Eyre, David R.; Yamada, Yoshihiko; Marini, Joan C.

    2016-01-01

    Recessive osteogenesis imperfecta (OI) is caused by defects in proteins involved in post-translational interactions with type I collagen. Recently, a novel form of moderately severe OI caused by null mutations in TMEM38B was identified. TMEM38B encodes the ER membrane monovalent cation channel, TRIC-B, proposed to counterbalance IP3R-mediated Ca2+ release from intracellular stores. The molecular mechanisms by which TMEM38B mutations cause OI are unknown. We identified 3 probands with recessive defects in TMEM38B. TRIC-B protein is undetectable in proband fibroblasts and osteoblasts, although reduced TMEM38B transcripts are present. TRIC-B deficiency causes impaired release of ER luminal Ca2+, associated with deficient store-operated calcium entry, although SERCA and IP3R have normal stability. Notably, steady state ER Ca2+ is unchanged in TRIC-B deficiency, supporting a role for TRIC-B in the kinetics of ER calcium depletion and recovery. The disturbed Ca2+ flux causes ER stress and increased BiP, and dysregulates synthesis of proband type I collagen at multiple steps. Collagen helical lysine hydroxylation is reduced, while telopeptide hydroxylation is increased, despite increased LH1 and decreased Ca2+-dependent FKBP65, respectively. Although PDI levels are maintained, procollagen chain assembly is delayed in proband cells. The resulting misfolded collagen is substantially retained in TRIC-B null cells, consistent with a 50–70% reduction in secreted collagen. Lower-stability forms of collagen that elude proteasomal degradation are not incorporated into extracellular matrix, which contains only normal stability collagen, resulting in matrix insufficiency. These data support a role for TRIC-B in intracellular Ca2+ homeostasis, and demonstrate that absence of TMEM38B causes OI by dysregulation of calcium flux kinetics in the ER, impacting multiple collagen-specific chaperones and modifying enzymes. PMID:27441836

  2. Diallylsulfide attenuates excessive collagen production and apoptosis in a rat model of bleomycin induced pulmonary fibrosis through the involvement of protease activated receptor-2

    SciTech Connect

    Kalayarasan, Srinivasan Sriram, Narayanan; Soumyakrishnan, Syamala; Sudhandiran, Ganapasam

    2013-09-01

    Pulmonary fibrosis (PF) can be a devastating lung disease. It is primarily caused by inflammation leading to severe damage of the alveolar epithelial cells. The pathophysiology of PF is not yet been clearly defined, but studying lung parenchymal injury by involving reactive oxygen species (ROS) through the activation of protease activated receptor-2 (PAR-2) may provide promising results. PAR-2 is a G-protein coupled receptor is known to play an important role in the development of PF. In this study, we investigated the inhibitory role of diallylsulfide (DAS) against ROS mediated activation of PAR-2 and collagen production accompanied by epithelial cell apoptosis. Bleomycin induced ROS levels may prompt to induce the expression of PAR-2 as well as extracellular matrix proteins (ECM), such as MMP 2 and 9, collagen specific proteins HSP-47, α-SMA, and cytokines IL-6, and IL-8RA. Importantly DAS treatment effectively decreased the expression of all these proteins. The inhibitory effect of DAS on profibrotic molecules is mediated by blocking the ROS level. To identify apoptotic signaling as a mediator of PF induction, we performed apoptotic protein expression, DNA fragmentation analysis and ultrastructural details of the lung tissue were performed. DAS treatment restored all these changes to near normalcy. In conclusion, treatment of PF bearing rats with DAS results in amelioration of the ROS production, PAR-2 activation, ECM production, collagen synthesis and alveolar epithelial cell apoptosis during bleomycin induction. We attained the first evidence that treatment of DAS decreases the ROS levels and may provide a potential therapeutic effect attenuating bleomycin induced PF. - Highlights: • DAS inhibits PAR-2 activity; bleomycin stimulates PAR-2 activity. • Increase in PAR-2 activity is correlated with pulmonary fibrosis • DAS reduces pro-inflammatory activity linked to facilitating pulmonary fibrosis. • DAS inhibits apoptosis of alveolar epithelial cells.

  3. MORPHOLOGICAL AND CHEMICAL STUDIES OF COLLAGEN FORMATION

    PubMed Central

    Lowther, D. A.; Green, N. M.; Chapman, J. A.

    1961-01-01

    Electron micrographs of thin sections of nuclear, microsomal, and mitochondrial fractions obtained from a carrageenin-induced granuloma showed considerable contamination of the heavier by the lighter fractions. Striated collagen fibrils could be identified in the nuclei + debris fraction. Only a few striated fibrils occurred in the mitochondrial fraction; very fine filaments (diameter 50 A) could be seen in this fraction, but could not be distinguished with certainty from fibrillar material derived from broken nuclei. 35 per cent of the mitochondrial and 80 per cent of the microsomal collagen was extractable by 0.2 M NaCl and could be purified by the standard methods of solution and reprecipitation. The amino acid composition of these collagen fractions determined by ion exchange chromatography was within the range normally found for collagen and gelatin from other mammalian species, allowing for 10 to 20 per cent of some non-collagenous contaminant of the microsomal collagen. Hydroxyproline and proline were isolated by chromatography on paper from hydrolysates of the nuclear, mitochondrial, and microsomal collagen fractions, after incubation of tissue slices with L-14C-proline. The specific activities of the hydroxyproline from these collagens were in the approximate ratio 1:2:6, while that of bound hydroxyproline derived from the supernatant was only 1, indicating primary synthesis of collagen in the microsomes. Attempts to demonstrate incorporation of L-14C-proline into collagen or into free hydroxyproline in cell free systems were unsuccessful, nor was it possible to demonstrate non-specific incorporation of L-14C-valine into TCA-insoluble material by various combinations of subcellular fractions. PMID:13763869

  4. Synthesis and biocompatibility of porous nano-hydroxyapatite/collagen/alginate composite.

    PubMed

    Zhang, S M; Cui, F Z; Liao, S S; Zhu, Y; Han, L

    2003-07-01

    Porous nano-hydroxyapatite/collagen/alginate (nHAC/Alginate) composite containing nHAC and Ca-crosslinked alginate is synthesized biomimetically. This composite shows a significant improvement in mechanical properties over nHAC material. Mechanical test results show that the compressive modulus and yield strength of this composite are in direct proportion to the percentage of Ca-crosslinked alginate in the composite. Primary biocompatibility experiments in vitro including fibroblasts and osteoblasts co-culture with nHAC/alginate composite indicated the high biocompatibility of this composite. Therefore the composite can be a promising candidate of scaffold material for bone tissue engineering.

  5. Biochemical and immunohistochemical comparison of collagen in granuloma annulare and skin sarcoidosis.

    PubMed

    Oikarinen, A; Kinnunen, T; Kallioinen, M

    1989-01-01

    Collagen was studied by biochemical and immunohistochemical means in 5 patients with granuloma annulare (GA) and 3 with cutaneous sarcoidosis (SA). The solubility of collagen from the lesional skin in acetic acid was higher than that of collagen from unaffected skin from both patients and control subjects. Collagen concentration in the skin lesions, measured in terms of hydroxyproline content, was reduced in 3 patients with granuloma annulare and one with sarcoidosis, but the ratio of type III/I collagen was unchanged vis-à-vis non-affected skin. The collagen concentration in non-affected skin of both GA and SA-patients was also lower than in controls. The most typical immunohistochemical finding was the association of type III procollagen and fibronectin with granulomas in the lesional skin of both GA and SA cases. The activity of prolyl hydroxylase, a key enzyme in collagen biosynthesis, was markedly increased in the lesional skin, indicating that collagen synthesis in vivo was also increased. Surprisingly, collagen synthesis was not increased in cell culture studies. This could be due to cell selection as observed previously in scleroderma. Another possibility could be that various mediators released in vivo from inflammatory cells activate fibroblasts. However, when cells are subcultivated, this effect is not maintained. In conclusion, marked changes in collagen could be observed in granuloma annulare and skin sarcoidosis, reflecting increased turnover of collagen in vivo.

  6. Double-layered collagen gel hemisphere for cell invasion assay: successful visualization and quantification of cell invasion activity.

    PubMed

    Takata, Masahiko; Maniwa, Yoshimasa; Doi, Takefumi; Tanaka, Yugo; Okada, Kenji; Nishio, Wataru; Ohbayashi, Chiho; Yoshimura, Masahiro; Hayashi, Yoshitake; Okita, Yutaka

    2007-10-01

    Although various methods for collagen gel-based cell invasion assays have been described, there continues to be a need for a simpler and more objective assay. Here, we describe an easy-to-prepare double-layered collagen gel hemisphere (DL-CGH) system that satisfies these requirements, and we demonstrate the advantages of this new system for visualizing cell movements during invasion. DL-CGH consists of a central core collagen layer surrounded by an outer cover collagen layer. A droplet of collagen I solution (containing cells to be examined) naturally forms a small hemisphere on the bottom of the culture dish. After this central core layer gels, a second droplet is placed atop the first gel, encapsulating it completely. The hemisphere is submerged in the medium and cultured. The invasive activity of cells that infiltrate from the inner to the outer layer can be evaluated optically. Using this in vitro system, we measured the inhibitory effect of E-cadherin expression on cancer cell invasion. DL-CGH also allowed visualization of interactions between invading cancer cells and the stroma. Cancer cells, which lack the proteases required for direct entrance into the three-dimensional collagen matrix, were seen to slip like amoebas through matrix gaps generated by the pericellular proteolytic activity of fibroblasts. [Supplementary materials are available for this article. Go to the publisher's online edition of Cell Communication and Adhesion for the following free supplemental resources: Movies 1-3; 4a and b]. PMID:17957531

  7. Type IV collagen is an activating ligand for the adhesion G protein-coupled receptor GPR126.

    PubMed

    Paavola, Kevin J; Sidik, Harwin; Zuchero, J Bradley; Eckart, Michael; Talbot, William S

    2014-08-12

    GPR126 is an orphan heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptor (GPCR) that is essential for the development of diverse organs. We found that type IV collagen, a major constituent of the basement membrane, binds to Gpr126 and activates its signaling function. Type IV collagen stimulated the production of cyclic adenosine monophosphate in rodent Schwann cells, which require Gpr126 activity to differentiate, and in human embryonic kidney (HEK) 293 cells expressing exogenous Gpr126. Type IV collagen specifically bound to the extracellular amino-terminal region of Gpr126 containing the CUB (complement, Uegf, Bmp1) and pentraxin domains. Gpr126 derivatives lacking the entire amino-terminal region were constitutively active, suggesting that this region inhibits signaling and that ligand binding relieves this inhibition to stimulate receptor activity. A new zebrafish mutation that truncates Gpr126 after the CUB and pentraxin domains disrupted development of peripheral nerves and the inner ear. Thus, our findings identify type IV collagen as an activating ligand for GPR126, define its mechanism of activation, and highlight a previously unrecognized signaling function of type IV collagen in basement membranes. PMID:25118328

  8. Synthesis, development, characterization and effectiveness of bovine pure platelet gel-collagen-polydioxanone bioactive graft on tendon healing

    PubMed Central

    Moshiri, Ali; Oryan, Ahmad; Meimandi-Parizi, Abdolhamid

    2015-01-01

    Bovine platelet gel (BPG) is an accessible and cost-effective source of growth factors which may have a value in tendon regenerative medicine. We produced a collagen implant (CI) as a tendon proper, covered it with polydioxanone (PDS) sheath to simulate paratenon and finally embedded the BPG as an active source of growth factor within the bioimplant to test whether BPG would be able to accelerate and enhance tendon regeneration and repair. After in vitro characterization of the bioactive grafts, the grafts were implanted in rabbit large tendon defect model. Untreated tendons and tendons treated with either CI or CI-PDS were served as controls for the CI-PDS-BPG. The animals were investigated clinically, ultrasonographically and haematologically for 120 days. After euthanasia, dry matter content, water uptake and delivery characteristics and also gross morphological, histopathological and scanning electron microscopic features of the healing tendons were assessed. In vitro, the activated platelets in the scaffold, released their growth factors significantly more than the controls. BPG also increased cell viability, and enhanced cellular differentiation, maturation and proliferation inside the CI-PDS compared with the controls. In vivo, the BPG modulated inflammation, increased quality and rate of fibroplasia and produced a remodelled tendon that had significantly higher collagen content and superior collagen fibril and fibre differentiation than controls. Treatment also significantly improved tendon water uptake and delivery characteristics, animals’ serum PDGF level, CI-PDS biocompatibility and biodegradability and reduced peritendinous adhesions, muscle fibrosis and atrophy. BPG was effective on tendon healing and CI-PDS-BPG may be a valuable bioscaffold in tendon reconstructive surgery. PMID:25702535

  9. Synthesis, development, characterization and effectiveness of bovine pure platelet gel-collagen-polydioxanone bioactive graft on tendon healing.

    PubMed

    Moshiri, Ali; Oryan, Ahmad; Meimandi-Parizi, Abdolhamid

    2015-06-01

    Bovine platelet gel (BPG) is an accessible and cost-effective source of growth factors which may have a value in tendon regenerative medicine. We produced a collagen implant (CI) as a tendon proper, covered it with polydioxanone (PDS) sheath to simulate paratenon and finally embedded the BPG as an active source of growth factor within the bioimplant to test whether BPG would be able to accelerate and enhance tendon regeneration and repair. After in vitro characterization of the bioactive grafts, the grafts were implanted in rabbit large tendon defect model. Untreated tendons and tendons treated with either CI or CI-PDS were served as controls for the CI-PDS-BPG. The animals were investigated clinically, ultrasonographically and haematologically for 120 days. After euthanasia, dry matter content, water uptake and delivery characteristics and also gross morphological, histopathological and scanning electron microscopic features of the healing tendons were assessed. In vitro, the activated platelets in the scaffold, released their growth factors significantly more than the controls. BPG also increased cell viability, and enhanced cellular differentiation, maturation and proliferation inside the CI-PDS compared with the controls. In vivo, the BPG modulated inflammation, increased quality and rate of fibroplasia and produced a remodelled tendon that had significantly higher collagen content and superior collagen fibril and fibre differentiation than controls. Treatment also significantly improved tendon water uptake and delivery characteristics, animals' serum PDGF level, CI-PDS biocompatibility and biodegradability and reduced peritendinous adhesions, muscle fibrosis and atrophy. BPG was effective on tendon healing and CI-PDS-BPG may be a valuable bioscaffold in tendon reconstructive surgery.

  10. Collagen type I and III synthesis by Tenon's capsule fibroblasts in culture: individual patient characteristics and response to mitomycin C, 5-fluorouracil, and ascorbic acid.

    PubMed Central

    Gross, R L

    1999-01-01

    PURPOSE: This study was performed to better understand the differences between patients in specific components of wound healing as it may pertain to glaucoma filtration surgery, including the use of antimetabolites. METHODS: Human Tenon's capsule fibroblasts were obtained at the time of glaucoma filtering surgery and established in individual cell cultures from 35 glaucoma patients. The dose-response to 5-fluorouracil (5FU) and mitomycin C (MMC) was determined. The individual cell lines were exposed to the antimetabolites and ascorbic acid with measurement of collagen type I and III production by an ELISA-type dot blot assay. These results were then statistically compared to the individual patient characteristics including age, race, previous surgery and medications, and type of glaucoma. RESULTS: 5-FU had little effect on collagen type I and III production or protein synthesis. MMC had an inhibitory effect on collagen secretion and total protein synthesis with increasing concentration. Photomicrographs of the cells after each treatment condition revealed characteristic morphologic changes when compared to controls. There was a large range of collagen type I and III production with correlation between the amounts of each collagen type secreted in response to the antimetabolites. However, there was no correlation with accepted risk factors for filtration failure. CONCLUSION: These antimetabolites act similarly on different cell lines in a nonspecific manner. The results suggest that the increased risk of filtration failure due to age, race, diagnosis, and previous conjunctival surgery is not due to differences in secretion of collagen types I and III by Tenon's capsule fibroblasts. Images FIGURE 3 PMID:10703140

  11. Synthesis and antimicrobial activity of squalamine analogue.

    PubMed

    Kim, H S; Choi, B S; Kwon, K C; Lee, S O; Kwak, H J; Lee, C H

    2000-08-01

    Synthesis and antimicrobial activity of squalamine analogue 2 are reported. The synthesis of 2 was accomplished from bisnoralcohol 3. The spermidine moiety was introduced via reductive amination of an appropriately functionalized 3beta-aminosterol with spermidinyl aldehyde 17 utilizing sodium triacetoxyborohydride as the reducing agent. Compound 2 shows weaker antimicrobial activity than squalamine. PMID:11003150

  12. Green Tea and Glycine Modulate the Activity of Metalloproteinases and Collagen in the Tendinitis of the Myotendinous Junction of the Achilles Tendon.

    PubMed

    Vieira, Cristiano Pedrozo; De Oliveira, LetÍCia Prado; Da Ré Guerra, Flávia; Marcondes, Maria Cristina Cintra; Pimentel, Edson Rosa

    2016-07-01

    The myotendinous junction (MTJ) is the weakest element in the muscle-tendon unit of the heel, and thus the most susceptible to injuries. The scarcity of adequate treatments means that tendinitis is a major concern to athletes and other groups who depend on their physical fitness, although green tea and glycine have both been shown to have beneficial effects on the inflammation. The present study investigated the remodeling effects of green tea and glycine in the MTJ of rats with tendinitis. For this, male Wistar rats were divided into five groups: animals without tendinitis and animals with tendinitis; animals with tendinitis supplied with green tea; animals with tendinitis supplied with a glycine diet; animals with tendinitis supplied with a green tea and glycine diet. Tendinitis was induced and the treatment with green tea (700 mg/kg/day) and a 5% glycine diet lasted 7 days. The treatments regulated the activity of metalloproteinases (MMP)-2, -8, and -9, and induced the synthesis of type I collagen, glycosaminoglycans, and non-collagenous proteins. Changes were also noted in the compaction of the collagen molecules and the amount of tenocytes. When combined, green tea and glycine modulated the inflammatory process and induced the synthesis of the elements involved in the post-lesion recovery of the tissue. The data from the MTJ were different when compared with results already published using the whole Achilles tendon. These data indicate that each region of the inflamed tendon can exhibit different responses during the treatment and therefore, modify its extracellular matrix components to facilitate recovery and repair. Anat Rec, 299:918-928, 2016. © 2016 Wiley Periodicals, Inc. PMID:27121758

  13. Lysyl oxidase activity contributes to collagen stabilization during liver fibrosis progression and limits spontaneous fibrosis reversal in mice.

    PubMed

    Liu, Susan B; Ikenaga, Naoki; Peng, Zhen-Wei; Sverdlov, Deanna Y; Greenstein, Andrew; Smith, Victoria; Schuppan, Detlef; Popov, Yury

    2016-04-01

    Collagen stabilization through irreversible cross-linking is thought to promote hepatic fibrosis progression and limit its reversibility. However, the mechanism of this process remains poorly defined. We studied the functional contribution of lysyl oxidase (LOX) to collagen stabilization and hepatic fibrosis progression/reversalin vivousing chronic administration of irreversible LOX inhibitor β-aminopropionitrile (BAPN, or vehicle as control) in C57Bl/6J mice with carbon tetrachloride (CCl4)-induced fibrosis. Fibrotic matrix stability was directly assessed using a stepwise collagen extraction assay and fibrotic septae morphometry. Liver cells and fibrosis were studied by histologic, biochemical methods and quantitative real-time reverse-transcription PCR. During fibrosis progression, BAPN administration suppressed accumulation of cross-linked collagens, and fibrotic septae showed widening and collagen fibrils splitting, reminiscent of remodeling signs observed during fibrosis reversal. LOX inhibition attenuated hepatic stellate cell activation markers and promoted F4/80-positive scar-associated macrophage infiltration without an increase in liver injury. In reversal experiments, BAPN-treated fibrotic mice demonstrated accelerated fibrosis reversal after CCl4withdrawal. Our findings demonstrate for the first time that LOX contributes significantly to collagen stabilization in liver fibrosis, promotes fibrogenic activation of attenuated hepatic stellate cells, and limits fibrosis reversal. Our data support the concept of pharmacologic targeting of LOX pathway to inhibit liver fibrosis and promote its resolution.-Liu, S. B., Ikenaga, N., Peng, Z.-W., Sverdlov, D. Y., Greenstein, A., Smith, V., Schuppan, D., Popov, Y. Lysyl oxidase activity contributes to collagen stabilization during liver fibrosis progression and limits spontaneous fibrosis reversal in mice.

  14. Photoactivated rose bengal functionalized chitosan nanoparticles produce antibacterial/biofilm activity and stabilize dentin-collagen

    PubMed Central

    Shrestha, Annie; Hamblin, Michael R.; Kishen, Anil

    2014-01-01

    Treatment of infected teeth presents two major challenges: persistence of the bacterial-biofilm within root canals after treatment and compromised structural integrity of the dentin hard-tissue. In this study bioactive polymeric chitosan nanoparticles functionalized with rose-bengal, CSRBnp was developed to produce antibiofilm effects as well as stabilize structural-integrity by photocrosslinking dentin-collagen. CSRBnp was less toxic to fibroblasts and had significant antibacterial activity even in the presence of bovine serum albumin. CSRBnp exerted antibacterial mechanism by adhering to bacterial cell surface, permeabilizing the membrane and lysing the cells subsequent to photodynamic treatment. Photoactivated CSRBnp resulted in reduced viability of Enterococcus faecalis biofilms and disruption of biofilm structure. Incorporation of CSRBnp and photocrosslinking significantly improved resistance to degradation and mechanical strength of dentin-collagen (p<0.05). The functionalized chitosan nanoparticles provided a single-step treatment of infected root dentin by combining the properties of chitosan and that of photosensitizer to eliminate bacterial-biofilms and stabilize dentin-matrix. PMID:24200522

  15. Diallylsulfide attenuates excessive collagen production and apoptosis in a rat model of bleomycin induced pulmonary fibrosis through the involvement of protease activated receptor-2.

    PubMed

    Kalayarasan, Srinivasan; Sriram, Narayanan; Soumyakrishnan, Syamala; Sudhandiran, Ganapasam

    2013-09-01

    Pulmonary fibrosis (PF) can be a devastating lung disease. It is primarily caused by inflammation leading to severe damage of the alveolar epithelial cells. The pathophysiology of PF is not yet been clearly defined, but studying lung parenchymal injury by involving reactive oxygen species (ROS) through the activation of protease activated receptor-2 (PAR-2) may provide promising results. PAR-2 is a G-protein coupled receptor is known to play an important role in the development of PF. In this study, we investigated the inhibitory role of diallylsulfide (DAS) against ROS mediated activation of PAR-2 and collagen production accompanied by epithelial cell apoptosis. Bleomycin induced ROS levels may prompt to induce the expression of PAR-2 as well as extracellular matrix proteins (ECM), such as MMP 2 and 9, collagen specific proteins HSP-47, α-SMA, and cytokines IL-6, and IL-8RA. Importantly DAS treatment effectively decreased the expression of all these proteins. The inhibitory effect of DAS on profibrotic molecules is mediated by blocking the ROS level. To identify apoptotic signaling as a mediator of PF induction, we performed apoptotic protein expression, DNA fragmentation analysis and ultrastructural details of the lung tissue were performed. DAS treatment restored all these changes to near normalcy. In conclusion, treatment of PF bearing rats with DAS results in amelioration of the ROS production, PAR-2 activation, ECM production, collagen synthesis and alveolar epithelial cell apoptosis during bleomycin induction. We attained the first evidence that treatment of DAS decreases the ROS levels and may provide a potential therapeutic effect attenuating bleomycin induced PF.

  16. The pro-fibrotic properties of transforming growth factor on human fibroblasts are counteracted by caffeic acid by inhibiting myofibroblast formation and collagen synthesis.

    PubMed

    Mia, Masum M; Bank, Ruud A

    2016-03-01

    Fibrosis is a chronic disorder affecting many organs. A universal process in fibrosis is the formation of myofibroblasts and the subsequent collagen deposition by these cells. Transforming growth factor beta1 (TGFβ1) plays a major role in the formation of myofibroblasts, e.g. by activating fibroblasts. Currently, no treatments are available to circumvent fibrosis. Caffeic acid phenethyl ester (CAPE) shows a broad spectrum of biological activities, including anti-fibrotic properties in vivo in mice and rats. However, little is known about the direct effects of CAPE on fibroblasts. We have tested whether CAPE is able to suppress myofibroblast formation and collagen formation of human dermal and lung fibroblasts exposed to TGFβ1, and found that this was indeed the case. In fact, the formation of myofibroblasts by TGFβ1 and subsequent collagen formation was completely abolished by CAPE. The same was observed for fibronectin and tenascin C. The lack of myofibroblast formation is likely due to the suppression of GLI1 and GLI2 expression by CAPE because of diminished nuclear SMAD2/3 levels. Post-treatment with CAPE after myofibroblast formation even resulted in a partial reversal of myofibroblasts into fibroblasts and/or reduction in collagen formation. Major discrepancies were seen between mRNA levels of collagen type I and cells stained positive for collagen, underlining the need for protein data in fibrosis studies to make reliable conclusions. PMID:26453399

  17. The Fc receptor gamma-chain and the tyrosine kinase Syk are essential for activation of mouse platelets by collagen.

    PubMed Central

    Poole, A; Gibbins, J M; Turner, M; van Vugt, M J; van de Winkel, J G; Saito, T; Tybulewicz, V L; Watson, S P

    1997-01-01

    Activation of mouse platelets by collagen is associated with tyrosine phosphorylation of multiple proteins including the Fc receptor gamma-chain, the tyrosine kinase Syk and phospholipase Cgamma2, suggesting that collagen signals in a manner similar to that of immune receptors. This hypothesis has been tested using platelets from mice lacking the Fc receptor gamma-chain or Syk. Tyrosine phosphorylation of Syk and phospholipase Cgamma2 by collagen stimulation is absent in mice lacking the Fc receptor gamma-chain. Tyrosine phosphorylation of phospholipase Cgamma2 by collagen stimulation is also absent in mice platelets which lack Syk, although phosphorylation of the Fc receptor gamma-chain is maintained. In contrast, tyrosine phosphorylation of platelet proteins by the G protein-coupled receptor agonist thrombin is maintained in mouse platelets deficient in Fc receptor gamma-chain or Syk. The absence of Fc receptor gamma-chain or Syk is accompanied by a loss of secretion and aggregation responses in collagen- but not thrombin-stimulated platelets. These observations provide the first direct evidence of an essential role for the immunoreceptor tyrosine-based activation motif (ITAM) in signalling by a non-immune receptor stimulus. PMID:9171347

  18. Disturbed synthesis of type II collagen interferes with rate of bone formation and growth and increases bone resorption in transgenic mice.

    PubMed

    Nieminen, Jyrki; Sahlman, Janne; Hirvonen, Teemu; Lapveteläinen, Tuomo; Miettinen, Markku; Arnala, Ilkka; Malluche, Hartmut H; Helminen, Heikki J

    2008-03-01

    Transgenic mice carrying an internally deleted human type II collagen gene (COL2A1) were used to study bone growth and development. This mutation has previously been shown to disturb the development of collagen fibrils in articular cartilage, causing chondrodysplasia and osteoarthritis. Type II collagen expression in bones was investigated with immunohistochemistry. The development and mineralization of the skeleton and anthropometric measurements on bones were evaluated using X-rays and dynamic histomorphometry. Type II collagen was expressed in the cartilage of developing bones. The bones of transgenic mice were smaller compared with the controls. The bone mass remained almost unchanged in transgenic mice after 1 month of age, leading to differences of 47% in trabecular bone volume (P = 0.012) and 40% in trabecular thickness (P < 0.01) at the age of 3 months compared with controls. At the age of 3 months the eroded surface per bone volume was 31% greater in transgenic mice compared with controls (P < 0.05). Trabecular thickness correlated positively with body weight (R = 0.71, P < 0.001). Interestingly, body weight correlated with bone volume in control mice (R = 0.27, P < 0.01), but no correlation was observed in transgenic mice. The disturbed synthesis of cartilage-specific type II collagen in growing transgenic mice retarded bone development, increased bone resorption, and altered tissue properties. This led to a negative net bone balance and small bone size. The results support the idea that an altered synthesis of cartilage-specific molecule(s) can disturb postnatal bone development and growth.

  19. Lysyl Oxidase Activity Is Required for Ordered Collagen Fibrillogenesis by Tendon Cells.

    PubMed

    Herchenhan, Andreas; Uhlenbrock, Franziska; Eliasson, Pernilla; Weis, MaryAnn; Eyre, David; Kadler, Karl E; Magnusson, S Peter; Kjaer, Michael

    2015-06-26

    Lysyl oxidases (LOXs) are a family of copper-dependent oxido-deaminases that can modify the side chain of lysyl residues in collagen and elastin, thereby leading to the spontaneous formation of non-reducible aldehyde-derived interpolypeptide chain cross-links. The consequences of LOX inhibition in producing lathyrism are well documented, but the consequences on collagen fibril formation are less clear. Here we used β-aminoproprionitrile (BAPN) to inhibit LOX in tendon-like constructs (prepared from human tenocytes), which are an experimental model of cell-mediated collagen fibril formation. The improvement in structure and strength seen with time in control constructs was absent in constructs treated with BAPN. As expected, BAPN inhibited the formation of aldimine-derived cross-links in collagen, and the constructs were mechanically weak. However, an unexpected finding was that BAPN treatment led to structurally abnormal collagen fibrils with irregular profiles and widely dispersed diameters. Of special interest, the abnormal fibril profiles resembled those seen in some Ehlers-Danlos Syndrome phenotypes. Importantly, the total collagen content developed normally, and there was no difference in COL1A1 gene expression. Collagen type V, decorin, fibromodulin, and tenascin-X proteins were unaffected by the cross-link inhibition, suggesting that LOX regulates fibrillogenesis independently of these molecules. Collectively, the data show the importance of LOX for the mechanical development of early collagenous tissues and that LOX is essential for correct collagen fibril shape formation. PMID:25979340

  20. Activated hepatic stellate cells are dependent on self-collagen, cleaved by membrane type 1 matrix metalloproteinase for their growth.

    PubMed

    Birukawa, Naoko Kubo; Murase, Kazuyuki; Sato, Yasushi; Kosaka, Akemi; Yoneda, Akihiro; Nishita, Hiroki; Fujita, Ryosuke; Nishimura, Miyuki; Ninomiya, Takafumi; Kajiwara, Keiko; Miyazaki, Miyono; Nakashima, Yusuke; Ota, Sigenori; Murakami, Yuya; Tanaka, Yasunobu; Minomi, Kenjiro; Tamura, Yasuaki; Niitsu, Yoshiro

    2014-07-18

    Stellate cells are distributed throughout organs, where, upon chronic damage, they become activated and proliferate to secrete collagen, which results in organ fibrosis. An intriguing property of hepatic stellate cells (HSCs) is that they undergo apoptosis when collagen is resolved by stopping tissue damage or by treatment, even though the mechanisms are unknown. Here we disclose the fact that HSCs, normal diploid cells, acquired dependence on collagen for their growth during the transition from quiescent to active states. The intramolecular RGD motifs of collagen were exposed by cleavage with their own membrane type 1 matrix metalloproteinase (MT1-MMP). The following evidence supports this conclusion. When rat activated HSCs (aHSCs) were transduced with siRNA against the collagen-specific chaperone gp46 to inhibit collagen secretion, the cells underwent autophagy followed by apoptosis. Concomitantly, the growth of aHSCs was suppressed, whereas that of quiescent HSCs was not. These in vitro results are compatible with the in vivo observation that apoptosis of aHSCs was induced in cirrhotic livers of rats treated with siRNAgp46. siRNA against MT1-MMP and addition of tissue inhibitor of metalloproteinase 2 (TIMP-2), which mainly inhibits MT1-MMP, also significantly suppressed the growth of aHSCs in vitro. The RGD inhibitors echistatin and GRGDS peptide and siRNA against the RGD receptor αVβ1 resulted in the inhibition of aHSCs growth. Transduction of siRNAs against gp46, αVβ1, and MT1-MMP to aHSCs inhibited the survival signal of PI3K/AKT/IκB. These results could provide novel antifibrosis strategies.

  1. Sulphatides trigger polymorphonuclear granulocyte spreading on collagen-coated surfaces and inhibit subsequent activation of 5-lipoxygenase.

    PubMed Central

    Sud'ina, G F; Brock, T G; Pushkareva, M A; Galkina, S I; Turutin, D V; Peters-Golden, M; Ullrich, V

    2001-01-01

    Sulphatides are sulphate esters of galactocerebrosides that are present on the surfaces of many cell types and act as specific ligands to selectins. The present study was undertaken to investigate the effect of sulphatides on polymorphonuclear granulocyte (PMN) attachment, spreading and 5-lipoxygenase (5-LO) metabolism. Sulphatides, but not non-sulphated galactocerebrosides, dose-dependently enhanced attachment to collagen, as measured by the myeloperoxidase assay. Studies with blocking antibodies indicated that the increased attachment was mediated by CD11b/CD18 (Mac-1) beta 2 integrin. Scanning electron microscopy indicated that sulphatides also greatly enhanced the degree of cell spreading. In PMNs treated in suspension, sulphatides had no effect on the ionophore A23187-stimulated release of arachidonic acid and the synthesis of 5-LO metabolites. In contrast, in PMNs attached to collagen, the enzymic conversion of arachidonic acid by 5-LO was inhibited by sulphatides. Inhibition of 5-LO metabolism by sulphatides was observed even in the presence of exogenous substrate, suggesting that sulphatides directly inhibited 5-LO action. Consistent with this, sulphatides interfered with ionophore-induced translocation of the 5-LO to the nuclear envelope. Substances competing with sulphatide binding to cells, like dextran sulphate, or a strong inhibitor of cell spreading, like the actin-polymerizing agent jasplakinolide, prevented the effects of sulphatides on PMN attachment and spreading and leukotriene synthesis. We conclude that shape changes occurring in response to sulphatides specifically impair PMN leukotriene synthesis by inhibiting translocation of 5-LO. PMID:11672437

  2. Electrodeposition of platinum nanoclusters on type I collagen modified electrode and its electrocatalytic activity for methanol oxidation

    NASA Astrophysics Data System (ADS)

    Sun, Yujing; Sun, Lanlan; Xu, Fugang; Guo, Cunlan; Liu, Zhelin; Zhang, Yue; Yang, Tao; Li, Zhuang

    2009-05-01

    We firstly reported a novel polymer matrix fabricated by type I collagen and polymers, and this matrix can be used as nanoreactors for electrodepositing platinum nanoclusters (PNCs). The type I collagen film has a significant effect on the growth of PNCs. The size of the platinum nanoparticles could be readily tuned by adjusting deposition time, potential and the concentration of electrolyte, which have been verified by field-emitted scanning electron microscopy (FE-SEM). Furthermore, cyclic voltammetry (CV) has demonstrated that the as-prepared PNCs can catalyze methanol directly with higher activity than that prepared on PSS/PDDA film, and with better tolerance to poisoning than the commercial E-TEK catalyst. The collagen-polymer matrix can be used as a general reactor to electrodeposit other metal nanostructures.

  3. Pepsin-solubilised collagen (PSC) from Red Sea cucumber (Stichopus japonicus) regulates cell cycle and the fibronectin synthesis in HaCaT cell migration.

    PubMed

    Park, Soo-Yeong; Lim, Hee Kyoung; Lee, Seogjae; Hwang, Hyeong Cheol; Cho, Somi K; Cho, Moonjae

    2012-05-01

    Pepsin-solubilised collagen (PSC) from Red Sea cucumber (Stichopus japonicus) was studied with respect to its wound-healing effects on a human keratinocyte (HaCaT) cell line. Disaggregated collagen fibres were treated with 0.1M NaOH for 24h and digested with pepsin for 72h to reach maximum yield of 26.6%. The results of an in vitro wound-healing test showed that migration of HaCaT cells was 1.5-fold faster on PSC-coated plates than on untreated plates. The migration rate of sea cucumber PSC was similar to that of rat PSC, but five times higher than that of bovine gelatin. HaCaT cells grown on PSC-coated plates revealed increased fibronectin synthesis (6-fold and 3-fold compared to gelatin and rat PSC, respectively). Additionally, sea cucumber PSCs induced HaCaT cell proliferation by decreasing the G1 phase by 5% and maintaining a larger population (8%) of cells in mitosis. Collagen from Red Sea cucumber might be useful as an alternative to mammalian collagen in the nutraceutical and pharmaceutical industries.

  4. B lymphocytes and B-cell activating factor promote collagen and profibrotic markers expression by dermal fibroblasts in systemic sclerosis

    PubMed Central

    2013-01-01

    Introduction B lymphocytes might play a pathogenic role in dermal fibrosis in systemic sclerosis (SSc). B-cell activating factor (BAFF), a key cytokine for B-cell activation, is increased in the serum and the skin of patients with SSc. However, the ability of B cells directly to stimulate dermal fibroblasts and the role of BAFF are not fully understood. We therefore investigated the involvement of B cells and BAFF in the expression of collagen and profibrotic markers by dermal fibroblasts. Methods Cocultures of blood B cells from healthy blood donors and normal or SSc dermal fibroblasts stimulated with anti-IgM and BAFF were performed. Alpha-SMA, TIMP1, MMP9, COL1A1, COL1A2, and COL3A1 mRNA expression were determined by quantitative RT-PCR. Soluble collagen, BAFF, IL-6, IL-1β, TGF-β1, and CCL2 protein secretion were assessed. Results Coculture of blood B cells and dermal fibroblasts isolated from SSc patients induced IL-6, TGF-β1, CCL2, and collagen secretion, as well as Alpha-SMA, TIMP1, and MMP9 expression in dermal fibroblasts. Transwell assays demonstrated that this induction was dependent on cell-cell contact. Addition of anti-IgM and BAFF to the coculture increased IL-6, CCL2, TGF-β1, and collagen secretion. B cell- and BAFF-induced collagen secretion was highly reduced by anti-TGF-β1 antibodies. Conclusions Our results showed for the first time a direct role of B cells on the production of collagen by dermal fibroblasts, which is further enhanced by BAFF. Thus, these results demonstrate a new pathogenic role of B cells and BAFF in fibrosis and systemic sclerosis. PMID:24289101

  5. Collagenous gastroduodenitis.

    PubMed

    Rustagi, Tarun; Rai, Mridula; Scholes, John V

    2011-10-01

    Collagenous gastroduodenitis is a rare histopathologic entity characterized by marked subepithelial collagen deposition with associated mucosal inflammatory infiltrate. Only 4 cases have been reported, of which 3 had associated collagenous colitis. Collagenous gastroduodenitis without colonic involvement is exceptionally rare with only 1 case reported so far in the literature. We present a case of a 68-year-old woman with dyspepsia and mild anemia, who was found to have nodular gastric and duodenal mucosa on endoscopic examination. Histopathology showed collagenous gastroduodenitis. To the best of our knowledge, this is the second (and first in English literature) reported case of isolated collagenous gastroduodenitis.

  6. Collagenous gastroduodenitis.

    PubMed

    Rustagi, Tarun; Rai, Mridula; Scholes, John V

    2011-10-01

    Collagenous gastroduodenitis is a rare histopathologic entity characterized by marked subepithelial collagen deposition with associated mucosal inflammatory infiltrate. Only 4 cases have been reported, of which 3 had associated collagenous colitis. Collagenous gastroduodenitis without colonic involvement is exceptionally rare with only 1 case reported so far in the literature. We present a case of a 68-year-old woman with dyspepsia and mild anemia, who was found to have nodular gastric and duodenal mucosa on endoscopic examination. Histopathology showed collagenous gastroduodenitis. To the best of our knowledge, this is the second (and first in English literature) reported case of isolated collagenous gastroduodenitis. PMID:21346601

  7. Effect of retinoic acid on protein synthesis by foetal bovine chondrocytes in high-density culture: down-regulation of the glucose-regulated protein, GRP-78, and type II collagen.

    PubMed Central

    Freyria, A M; Ronzière, M C; Boutillon, M M; Herbage, D

    1995-01-01

    The effect of 0.1-10 microM retinoic acid (RA) on foetal bovine chondrocytes was investigated in high-density cultures (0.6 x 10(6) cells/cm2). After 5 days of culture in ascorbate-free medium, control chondrocytes presented a typical rounded shape and synthesized type II, IX, XI and III collagens. After RA treatment on days 2-5 of culture, the cells exhibited a fibroblast-like shape and decreased synthesis of total protein (48%) and pepsinresistant proteins (60%) as determined by [35S]methionine labelling. Addition of RA was not followed by the expression of type I collagen, but induced quantitative changes in the synthesis of cartilage-specific collagens (II, IX and XI) as measured by direct autoradiography of the corresponding bands after SDS/PAGE. The main change was in type II collagen synthesis, with a 80% decrease in the cell-layer fraction and a 89% decrease in culture-medium fraction; inhibition of type IX and XI collagen synthesis was limited to 25 and 31% respectively. Modifications to intracellular proteins induced by RA were determined by using two-dimensional electrophoresis associated with a computerized imaging system. Synthesis of one of the more abundant proteins (pI 4.8; 78 kDa) was decreased by 75% after RA treatment. This protein was characterized by micro-sequencing as the glucose-regulated protein 78 (GRP 78). It was reported previously to bind denatured collagen and mutated type I procollagen molecule and to function as a molecular chaperone for collagen molecules. It remains to demonstrate whether the parallel down-regulation of GRP 78 and type II collagen observed here corresponds to a co-ordinate regulation of these two proteins. Images Figure 1 Figure 2 Figure 3 PMID:7832751

  8. Identification of Domains and Amino Acids Essential to the Collagen Galactosyltransferase Activity of GLT25D1

    PubMed Central

    Perrin-Tricaud, Claire; Rutschmann, Christoph; Hennet, Thierry

    2011-01-01

    Collagen is modified by hydroxylation and glycosylation of hydroxylysine residues. This glycosylation is initiated by the β1,O galactosyltransferases GLT25D1 and GLT25D2. The structurally similar protein cerebral endothelial cell adhesion molecule CEECAM1 was previously reported to be inactive when assayed for collagen glycosyltransferase activity. To address the cause of the absent galactosyltransferase activity, we have generated several chimeric constructs between the active human GLT25D1 and inactive human CEECAM1 proteins. The assay of these chimeric constructs pointed to a short central region and a large C-terminal region of CEECAM1 leading to the loss of collagen galactosyltransferase activity. Examination of the three DXD motifs of the active GLT25D1 by site-directed mutagenesis confirmed the importance of the first (amino acids 166–168) and second motif (amino acids 461–463) for enzymatic activity, whereas the third one was dispensable. Since the second DXD motif is incomplete in CEECAM1, we have restored the motif by introducing the substitution S461D. This change did not restore the activity of the C-terminal region, thereby showing that additional amino acids were required in this C-terminal region to confer enzymatic activity. Finally, we have introduced the substitution Q471R-V472M-N473Q-P474V in the CEECAM1-C-terminal construct, which is found in most animal GLT25D1 and GLT25D2 isoforms but not in CEECAM1. This substitution was shown to partially restore collagen galactosyltransferase activity, underlining its importance for catalytic activity in the C-terminal domain. Because multiple mutations in different regions of CEECAM1 contribute to the lack of galactosyltransferase activity, we deduced that CEECAM1 is functionally different from the related GLT25D1 protein. PMID:22216269

  9. Stimulation of collagen synthesis in fibroblast cultures by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+.

    PubMed

    Maquart, F X; Pickart, L; Laurent, M; Gillery, P; Monboisse, J C; Borel, J P

    1988-10-10

    Glycyl-L-histidyl-L-lysine (GHK) is a tripeptide with affinity for copper(II) ions and was isolated from human plasma. This peptide appears to play a physiological role in wound healing. We report the stimulating effect of GHK-Cu on collagen synthesis by fibroblasts. The stimulation began between 10(-12) and 10(-11) M, maximized at 10(-9) M, and was independent of any change in cell number. The presence of a GHK triplet in the alpha 2(I) chain of type I collagen suggests that the tripeptide might be liberated by proteases at the site of a wound and exert in situ healing effects. PMID:3169264

  10. Yeast hydrolysate protects cartilage via stimulation of type II collagen synthesis and suppression of MMP-13 production.

    PubMed

    Lee, Hyun-Sun; Park, So Yeon; Park, Yooheon; Bae, Song Hwan; Suh, Hyung Joo

    2013-09-01

    Type II collagen (COL II) is one of the primary components of hyaline cartilage and plays a key role in maintaining chondrocyte function. COL II is the principal target of destruction, and matrix metalloproteases (MMPs) have a major role in arthritis. In the present study, we investigated the chondroctye protection effects of specific fraction of yeast hydrolysate ((10-30 kDa molecular weight peptides). The mRNA expression of COL II was significantly increased in the YH-treated group compared to the control at concentrations above 50 µg/ml, respectively. The 200 µg/ml YH-treated group (3.43 ± 0.23 µg/ml) showed significantly reduced glycosaminoglycan (GAG) degradation relative to that in the interleukin-1β (IL-1β)-treated control group (4.72 ± 0.05 µg/ml). In the YH-treated group, MMP-13 level was significantly decreased in a dose-dependent manner compared to the IL-1β-treated group without YH treatment. However, MMP-1 and MMP-3 level were not different from that of control. Under the same conditions, we also examined mRNA levels of COL II. The mRNA expression of COL II was significantly higher in the YH-treated group than in the IL-1β-treated control group at concentrations above 100 µg/ml. In conclusion, YH stimulated COL II synthesis and significantly inhibited MMP-13 and GAG degradation caused by IL-1β treatment. PMID:23070893

  11. Collagen fillers.

    PubMed

    Baumann, Leslie; Kaufman, Joely; Saghari, Sogol

    2006-01-01

    Collagen implants, both animal and human derived, have been used for soft tissue augmentation for many years. Bovine collagen fillers were the most popular injectable implants for nearly two decades in the United States. Since then, human bioengineered collagen products have been available in addition to hyaluronic acid-containing fillers. This article outlines the different types of injectable collagen implants, injection techniques, preferred methods of treatment, and possible adverse reactions to the injectable materials.

  12. Greener synthesis of electrospun collagen/hydroxyapatite composite fibers with an excellent microstructure for bone tissue engineering

    PubMed Central

    Zhou, Yuanyuan; Yao, Hongchang; Wang, Jianshe; Wang, Dalu; Liu, Qian; Li, Zhongjun

    2015-01-01

    In bone tissue engineering, collagen/hydroxyapatite (HAP) fibrous composite obtained via electrospinning method has been demonstrated to support the cells’ adhesion and bone regeneration. However, electrospinning of natural collagen often requires the use of cytotoxic organic solvents, and the HAP crystals were usually aggregated and randomly distributed within a fibrous matrix of collagen, limiting their clinical potential. Here, an effective and greener method for the preparation of collagen/HAP composite fibers was developed for the first time, and this green product not only had 40 times higher mechanical properties than that previously reported, but also had an excellent microstructure similar to that of natural bone. By dissolving type I collagen in environmentally friendly phosphate buffered saline/ethanol solution instead of the frequently-used cytotoxic organic solvents, followed with the key step of desalination, co-electrospinning the collagen solution with the HAP sol, generates a collagen/HAP composite with a uniform and continuous fibrous morphology. Interestingly, the nano-HAP needles were found to preferentially orient along the longitudinal direction of the collagen fibers, which mimicked the nanostructure of natural bones. Based on the characterization of the related products, the formation mechanism for this novel phenomenon was proposed. After cross-linking with 1-ethyl-3-(3-dimethyl-aminopropyl)-1-carbodiimide hydrochloride/N-hydroxysuccinimide, the obtained composite exhibited a significant enhancement in mechanical properties. In addition, the biocompatibility of the obtained composite fibers was evaluated by in vitro culture of the human myeloma cells (U2-OS). Taken together, the process outlined herein provides an effective, non-toxic approach for the fabrication of collagen/HAP composite nanofibers that could be good candidates for bone tissue engineering. PMID:25995630

  13. Greener synthesis of electrospun collagen/hydroxyapatite composite fibers with an excellent microstructure for bone tissue engineering.

    PubMed

    Zhou, Yuanyuan; Yao, Hongchang; Wang, Jianshe; Wang, Dalu; Liu, Qian; Li, Zhongjun

    2015-01-01

    In bone tissue engineering, collagen/hydroxyapatite (HAP) fibrous composite obtained via electrospinning method has been demonstrated to support the cells' adhesion and bone regeneration. However, electrospinning of natural collagen often requires the use of cytotoxic organic solvents, and the HAP crystals were usually aggregated and randomly distributed within a fibrous matrix of collagen, limiting their clinical potential. Here, an effective and greener method for the preparation of collagen/HAP composite fibers was developed for the first time, and this green product not only had 40 times higher mechanical properties than that previously reported, but also had an excellent microstructure similar to that of natural bone. By dissolving type I collagen in environmentally friendly phosphate buffered saline/ethanol solution instead of the frequently-used cytotoxic organic solvents, followed with the key step of desalination, co-electrospinning the collagen solution with the HAP sol, generates a collagen/HAP composite with a uniform and continuous fibrous morphology. Interestingly, the nano-HAP needles were found to preferentially orient along the longitudinal direction of the collagen fibers, which mimicked the nanostructure of natural bones. Based on the characterization of the related products, the formation mechanism for this novel phenomenon was proposed. After cross-linking with 1-ethyl-3-(3-dimethyl-aminopropyl)-1-carbodiimide hydrochloride/N-hydroxysuccinimide, the obtained composite exhibited a significant enhancement in mechanical properties. In addition, the biocompatibility of the obtained composite fibers was evaluated by in vitro culture of the human myeloma cells (U2-OS). Taken together, the process outlined herein provides an effective, non-toxic approach for the fabrication of collagen/HAP composite nanofibers that could be good candidates for bone tissue engineering.

  14. Greener synthesis of electrospun collagen/hydroxyapatite composite fibers with an excellent microstructure for bone tissue engineering.

    PubMed

    Zhou, Yuanyuan; Yao, Hongchang; Wang, Jianshe; Wang, Dalu; Liu, Qian; Li, Zhongjun

    2015-01-01

    In bone tissue engineering, collagen/hydroxyapatite (HAP) fibrous composite obtained via electrospinning method has been demonstrated to support the cells' adhesion and bone regeneration. However, electrospinning of natural collagen often requires the use of cytotoxic organic solvents, and the HAP crystals were usually aggregated and randomly distributed within a fibrous matrix of collagen, limiting their clinical potential. Here, an effective and greener method for the preparation of collagen/HAP composite fibers was developed for the first time, and this green product not only had 40 times higher mechanical properties than that previously reported, but also had an excellent microstructure similar to that of natural bone. By dissolving type I collagen in environmentally friendly phosphate buffered saline/ethanol solution instead of the frequently-used cytotoxic organic solvents, followed with the key step of desalination, co-electrospinning the collagen solution with the HAP sol, generates a collagen/HAP composite with a uniform and continuous fibrous morphology. Interestingly, the nano-HAP needles were found to preferentially orient along the longitudinal direction of the collagen fibers, which mimicked the nanostructure of natural bones. Based on the characterization of the related products, the formation mechanism for this novel phenomenon was proposed. After cross-linking with 1-ethyl-3-(3-dimethyl-aminopropyl)-1-carbodiimide hydrochloride/N-hydroxysuccinimide, the obtained composite exhibited a significant enhancement in mechanical properties. In addition, the biocompatibility of the obtained composite fibers was evaluated by in vitro culture of the human myeloma cells (U2-OS). Taken together, the process outlined herein provides an effective, non-toxic approach for the fabrication of collagen/HAP composite nanofibers that could be good candidates for bone tissue engineering. PMID:25995630

  15. Method to evaluate the proliferation of activated lymphocytes in a three-dimensional collagen matrix.

    PubMed

    Davidova, Natalya V; Gorlina, Natalia K; Sharova, Svetlana V; Cheredeev, Anatoly N; Kozlov, Ivan G

    2002-12-01

    It is well known that the enhancement of the cell-matrix interactions represents one of the early steps in the process of lymphocyte activation. However, the information regarding the role of these interactions in the late stages of lymphocyte activation (in particular, the proliferation) is still controversial. This is basically due to the absence of adequate experimental models. In the present work we carried out a step-by-step modification of a well-studied model of mitogen-stimulated lymphocyte activation, adjusting it to the conditions of a three-dimensional collagen matrix (3D-CM). All the changes added to the standard procedure in the process of this modification were rigorously controlled using various experimental models. The final version of the method includes the following steps: (i) 24-h lymphocyte (lymphocyte fraction from mouse spleen) preincubation with mitogens (Con A or LPS) with a subsequent cell wash (parameters being controlled: irreversible lymphocyte activation, independence of the proliferation from cell-cell interactions); (ii) transfer of the activated lymphocytes to (3)H-thymidine containing 3D-CM and incubation for 48 h (controlled parameters: distribution of the radioactive label within the 3D-CM and its biological accessibility to lymphocytes); (iii) degradation of the 3D-CM with bacterial collagenase and cell transfer onto glass fiber filters (controlled parameters: cell viability after cultivation in the 3D-CM and treatment with the collagenase). With this method we found that the proliferation of the Con A- and LPS-stimulated lymphocytes in 3D-CM was dramatically inhibited (by 66.5 +/- 14.9% and by 88.1 +/- 10.2%, respectively). The discovered inhibition of the lymphocyte proliferation was not a consequence of either the ineffectiveness of the mitogens, the disruption of the cell-cell interactions, an insufficient inclusion of the radioactive label into cells, or of a decreased cell viability.

  16. Characterization and antioxidant activities of marine pepsin soluble collagen from the skin of yellow goosefish Lophius litulon

    NASA Astrophysics Data System (ADS)

    Zheng, Bin; Xiang, Xingwei; Zhou, Yufang; Yang, Huicheng; Luo, Hongyu; Liao, Miaofei; Wen, Zhengshun

    2016-06-01

    Characteristics and antioxidant activities of pepsin-soluble collagen (PSC) from yellow goosefish (Lophius litulon) skins were investigated. PSC was characterized as a type I collagen, and its imino acid content was 193 residues/1 000 residues. PSC's denaturation temperature was ~17.56°C and Fourier transform infrared spectra confirmed the presence of triple helices. Solubility analysis showed good solubility at acidic pH (1-6) or low NaCl concentrations (≤2%). PSC showed scavenging activity against hydroxyl radicals and superoxide anions in a concentration-dependent manner. Furthermore, PSC could protect D-galactose-induced skin aging by significantly controlling malondialdehyde formation and improving the activity of superoxide dismutase, glutathione peroxidase, catalase, glutathione, and hydroxyproline. PSC may be a promising antioxidant in appropriate applications.

  17. Antifreeze and cryoprotective activities of ice-binding collagen peptides from pig skin.

    PubMed

    Cao, Hui; Zhao, Ying; Zhu, Yu Bing; Xu, Fei; Yu, Jing Song; Yuan, Min

    2016-03-01

    A novel "hyperactive" ice-binding peptide from porcine collagen was prepared by alkaline protease hydrolysis and a series of column chromatography separations, and then its antifreeze and cryoprotective properties were reported. Using differential scanning calorimetry (DSC), the thermal hysteresis (TH) of ice-binding collagen peptides was closely related to their concentration and crystal fraction. Collagen hydrolysates with maximal TH were obtained by hydrolysis at pH 8.0, DH 15.0%, and 5% alkaline protease at 55°C. After purification by column chromatography, the AP-3 ice-binding collagen peptide (GLLGPLGPRGLL) with 1162.8Da molecular weights exhibited the highest TH (5.28°C), which can be classified as "hyperactive". Recrystallisation and melt-resistance of ice cream were improved by AP-3 ice-binding collagen peptide at 0.2% (w/v) in a similar manner to natural antifreeze proteins. Moreover, the addition of AP-3 collagen peptides in ice cream greatly elevated the glass transition temperature (Tg) to -17.64°C.

  18. Antifreeze and cryoprotective activities of ice-binding collagen peptides from pig skin.

    PubMed

    Cao, Hui; Zhao, Ying; Zhu, Yu Bing; Xu, Fei; Yu, Jing Song; Yuan, Min

    2016-03-01

    A novel "hyperactive" ice-binding peptide from porcine collagen was prepared by alkaline protease hydrolysis and a series of column chromatography separations, and then its antifreeze and cryoprotective properties were reported. Using differential scanning calorimetry (DSC), the thermal hysteresis (TH) of ice-binding collagen peptides was closely related to their concentration and crystal fraction. Collagen hydrolysates with maximal TH were obtained by hydrolysis at pH 8.0, DH 15.0%, and 5% alkaline protease at 55°C. After purification by column chromatography, the AP-3 ice-binding collagen peptide (GLLGPLGPRGLL) with 1162.8Da molecular weights exhibited the highest TH (5.28°C), which can be classified as "hyperactive". Recrystallisation and melt-resistance of ice cream were improved by AP-3 ice-binding collagen peptide at 0.2% (w/v) in a similar manner to natural antifreeze proteins. Moreover, the addition of AP-3 collagen peptides in ice cream greatly elevated the glass transition temperature (Tg) to -17.64°C. PMID:26471678

  19. Transcript-activated collagen matrix as sustained mRNA delivery system for bone regeneration.

    PubMed

    Badieyan, Zohreh Sadat; Berezhanskyy, Taras; Utzinger, Maximilian; Aneja, Manish Kumar; Emrich, Daniela; Erben, Reinhold; Schüler, Christiane; Altpeter, Philipp; Ferizi, Mehrije; Hasenpusch, Günther; Rudolph, Carsten; Plank, Christian

    2016-10-10

    Transcript therapies using chemically modified messenger RNAs (cmRNAs) are emerging as safe and promising alternatives for gene and recombinant protein therapies. However, their applications have been limited due to transient translation and relatively low stability of cmRNAs compared to DNA. Here we show that vacuum-dried cmRNA-loaded collagen sponges, termed transcript activated matrices (TAMs), can serve as depots for sustained delivery of cmRNA. TAMs provide steady state protein production for up to six days, and substantial residual expression until 11days post transfection. Another advantage of this technology was nearly 100% transfection efficiency as well as low toxicity in vitro. TAMs were stable for at least 6months at room temperature. Human BMP-2-encoding TAMs induced osteogenic differentiation of MC3T3-E1 cells in vitro and bone regeneration in a non-critical rat femoral bone defect model in vivo. In summary, TAMs are a promising tool for bone regeneration and potentially also for other applications in regenerative medicine and tissue engineering. PMID:27586186

  20. Collagen Specific T-Cell Repertoire and HLA-DR Alleles: Biomarkers of Active Refractory Rheumatoid Arthritis

    PubMed Central

    Di Sante, Gabriele; Tolusso, Barbara; Fedele, Anna Laura; Gremese, Elisa; Alivernini, Stefano; Nicolò, Chiara; Ria, Francesco; Ferraccioli, Gianfranco

    2015-01-01

    Rheumatoid arthritis (RA) is characterized by chronic joint inflammation and associates with HLA-DRB1*04. The Collagen IIp261-273-specific T cell repertoire in the peripheral blood of DR4 + patients at the onset of the disease shows a restricted TCR-beta chain usage among which the most frequent is TRBV25. To define whether this group of DR4-restricted collagen-specific shared T cell could represent markers of active-severe disease and response to therapy, 90 subjects affected by early-RA were enrolled in the study; peripheral blood mononuclear cells were cultured with or without the human collagen II peptide p261-273 and were examined by immunoscope analysis for the usage of the previously identified shared TCR-beta chains. We report that the presence of T cells carrying rearrangement TRBV25 associated with HLA-DR haplotype and disease activity. HLA-DRB1* haplotypes 04–04, 04–01 and 04–11 were significantly associated with usage of TRBV25, higher disease activity at the onset of disease and poor response to DMARDs. Finally, the HLA-DRB1* haplotype appeared complementary with current serologic tools to predict good and poor responders in a treat to target strategy. The data reported here offer clues to predict the course of the disease and to foresee personalized treatments in RA patients. PMID:26844284

  1. Collagen Specific T-Cell Repertoire and HLA-DR Alleles: Biomarkers of Active Refractory Rheumatoid Arthritis.

    PubMed

    Di Sante, Gabriele; Tolusso, Barbara; Fedele, Anna Laura; Gremese, Elisa; Alivernini, Stefano; Nicolò, Chiara; Ria, Francesco; Ferraccioli, Gianfranco

    2015-12-01

    Rheumatoid arthritis (RA) is characterized by chronic joint inflammation and associates with HLA-DRB1*04. The Collagen IIp261-273-specific T cell repertoire in the peripheral blood of DR4 + patients at the onset of the disease shows a restricted TCR-beta chain usage among which the most frequent is TRBV25. To define whether this group of DR4-restricted collagen-specific shared T cell could represent markers of active-severe disease and response to therapy, 90 subjects affected by early-RA were enrolled in the study; peripheral blood mononuclear cells were cultured with or without the human collagen II peptide p261-273 and were examined by immunoscope analysis for the usage of the previously identified shared TCR-beta chains. We report that the presence of T cells carrying rearrangement TRBV25 associated with HLA-DR haplotype and disease activity. HLA-DRB1* haplotypes 04-04, 04-01 and 04-11 were significantly associated with usage of TRBV25, higher disease activity at the onset of disease and poor response to DMARDs. Finally, the HLA-DRB1* haplotype appeared complementary with current serologic tools to predict good and poor responders in a treat to target strategy. The data reported here offer clues to predict the course of the disease and to foresee personalized treatments in RA patients.

  2. The 45 kDa collagen-binding fragment of fibronectin induces matrix metalloproteinase-13 synthesis by chondrocytes and aggrecan degradation by aggrecanases.

    PubMed Central

    Stanton, Heather; Ung, Linh; Fosang, Amanda J

    2002-01-01

    Fragments of fibronectin occur naturally in vivo and are increased in the synovial fluid of arthritis patients. We have studied the 45 kDa fragment (Fn-f 45), representing the N-terminal collagen-binding domain of fibronectin, for its ability to modulate the expression of metalloproteinases by porcine articular chondrocytes in vitro. We report that stimulation of cultured chondrocytes, or cartilage explants, with Fn-f 45 increased the levels of matrix metalloproteinase-13 (MMP-13; collagenase-3) released into the conditioned medium in a dose-dependent manner. Increased levels of MMP-13 were due to stimulation of MMP-13 synthesis, rather than release of MMP-13 from accumulated matrix stores. Fn-f 45 also stimulated the synthesis of MMP-3 (stromelysin-1) from cultured chondrocytes and cartilage cultures. The Fn-f 45-induced increase in MMP-3 and MMP-13 synthesis occurred via an interleukin 1-independent mechanism, since the receptor antagonist of interleukin-1 was unable to block the increased synthesis. The gelatinases, MMP-2 and MMP-9, were not modulated by Fn-f 45 in these culture systems. Fn-f 45 also stimulated the release of aggrecan from cartilage explants into conditioned medium. Neoepitope antibodies specific for aggrecan fragments generated by MMPs or aggrecanases showed that the Fn-f 45-induced aggrecan loss was mediated by aggrecanases, and not by MMPs. Extracts of cultured cartilage contained elevated levels of the aggrecanase-derived ITEGE(373)-G1 domain, whereas levels of the matrix metalloproteinase-derived DIPEN(341)-G1 domain were unchanged. These studies show that Fn-f 45 can induce a catabolic phenotype in articular chondrocytes by up-regulating the expression of metalloproteinases specific for the degradation of collagen and aggrecan. PMID:11988091

  3. Collagen in organ development

    NASA Technical Reports Server (NTRS)

    Hardman, P.; Spooner, B. S.

    1992-01-01

    It is important to know whether microgravity will adversely affect developmental processes. Collagens are macromolecular structural components of the extracellular matrix (ECM) which may be altered by perturbations in gravity. Interstitial collagens have been shown to be necessary for normal growth and morphogenesis in some embryonic organs, and in the mouse salivary gland, the biosynthetic pattern of these molecules changes during development. Determination of the effects of microgravity on epithelial organ development must be preceded by crucial ground-based studies. These will define control of normal synthesis, secretion, and deposition of ECM macromolecules and the relationship of these processes to morphogenesis.

  4. Collagenous gastritis.

    PubMed

    Jain, Richa; Chetty, Runjan

    2010-12-01

    A 25-year-old patient presented with epigastric pain, which on gastric biopsy revealed the characteristic appearance of collagenous gastritis. There was a thick prominent subepithelial band that was confirmed to be collagen with a Masson's trichrome stain. There was associated Helicobacter pylori gastritis but no evidence of a lymphocytic gastritis. The patient did not have watery diarrhea. Collagenous gastritis can occur in young patients, be restricted to the stomach, and can be associated with celiac disease. PMID:19103610

  5. Correlation of collagen synthesis with polarization-sensitive optical coherence tomography imaging of in vitro human atherosclerosis

    NASA Astrophysics Data System (ADS)

    Kuo, Wen-Chuan; Shyu, Jeou-Jong; Chou, Nai-Kuan; Lai, Chih-Ming; Tien, En-Kuang; Huang, Huan-Jang; Chou, Chien; Jan, Gwo-Jen

    2005-04-01

    Atherosclerosis is unquestionably the leading cause of morbidity and mortality in developed countries. In the mean time, the worldwide importance of acute vascular syndromes is increasing. Because collagen fiber is a critical component of atherosclerotic lesions; it constitutes up to 60% of the total atherosclerotic plaque protein. The uncontrolled collagen accumulation leads to arterial stenosis, whereas excessive collagen breakdown weakens plaques thereby making them prone to rupture finally. Thus, in this study, we present the first application, to our knowledge, of using polarization-sensitive optical coherence tomography (PS-OCT) in human atherosclerosis. We demonstrate this technique for imaging of intensity, birefringence, and fast-axis orientation simultaneously in atherosclerotic plaques. This in vitro study suggests that the birefringence change in plaque is due to the prominent deposition of collagen according to the correlation of PS-OCT images with histological counterpart. Moreover, we can acquire quantitative criteria based on the change of polarization of incident beam to estimate whether the collagen synthesized is "too much" or "not enough". Thus by combining of high resolution intensity imaging and birefringence detection makes PS-OCT could be a potentially powerful tool for early assessment of atherosclerosis appearance and the prediction of plaque rupture in clinic.

  6. In situ quantitative evaluation of osteoblastic collagen synthesis under cyclic strain by using second-harmonic-generation microscope

    NASA Astrophysics Data System (ADS)

    Matsubara, Oki; Hase, Eiji; Minamikawa, Takeo; Yasui, Takeshi; Sato, Katsuya

    2016-03-01

    Osteoblast-produced collagen matrix in bone is influenced by the mechanical stimulus from their surroundings. However, it has been still unclear how mechanical stimulus affects collagen production by osteoblasts. Therefore, it is strongly required to investigate the characteristics of osteoblastic bone regenerative tissue engineering. Recently, second-harmonic-generation (SHG) microscope has attracted attention for in situ visualization of collagen fiber because of less invasiveness, unstaining and no fixation, as well as high spatial resolution and 3D imaging. Using SHG microscopy, one can track the temporal dynamics of collagen fiber during the cultured period of the sample. We applied cyclic stretch strain to osteoblasts (MC3T3-E1) by using originally developed cell stretching device. The stimulation time was set to 5min or 3hours with same strain 5% and same frequency 0.5Hz. Cells were seeded onto the PDMS (polydimethylsiloxane) rubber chamber at a density of 50,000 cells/cm2 and cultured in α-MEM with 10% FBS, 1% P/S, 1% Ascorbic acid, 0.2% hydrocortisone and 2% β-Glycerophosphate. SHG imaging was carried out every 7 days. As a result, we confirmed from SHG image that the collagen production was enhanced by the cyclic stretch strain, stretch stimulation time and stretch application term.

  7. Trehalose glycolipids--synthesis and biological activities.

    PubMed

    Khan, Ashna A; Stocker, Bridget L; Timmer, Mattie S M

    2012-07-15

    A variety of trehalose glycolipids have been isolated from natural sources, and several of these glycolipids exhibit important biological properties. These molecules also represent challenging synthetic targets due to their highly amphiphilic character, their large number of functional groups and additional chiral centres. This review highlights some of the recent advances made in the synthesis of trehalose glycolipids, and their associated biological activities.

  8. Multi-scale mechanical characterization of highly swollen photo-activated collagen hydrogels.

    PubMed

    Tronci, Giuseppe; Grant, Colin A; Thomson, Neil H; Russell, Stephen J; Wood, David J

    2015-01-01

    Biological hydrogels have been increasingly sought after as wound dressings or scaffolds for regenerative medicine, owing to their inherent biofunctionality in biological environments. Especially in moist wound healing, the ideal material should absorb large amounts of wound exudate while remaining mechanically competent in situ. Despite their large hydration, however, current biological hydrogels still leave much to be desired in terms of mechanical properties in physiological conditions. To address this challenge, a multi-scale approach is presented for the synthetic design of cyto-compatible collagen hydrogels with tunable mechanical properties (from the nano- up to the macro-scale), uniquely high swelling ratios and retained (more than 70%) triple helical features. Type I collagen was covalently functionalized with three different monomers, i.e. 4-vinylbenzyl chloride, glycidyl methacrylate and methacrylic anhydride, respectively. Backbone rigidity, hydrogen-bonding capability and degree of functionalization (F: 16 ± 12-91 ± 7 mol%) of introduced moieties governed the structure-property relationships in resulting collagen networks, so that the swelling ratio (SR: 707 ± 51-1996 ± 182 wt%), bulk compressive modulus (Ec: 30 ± 7-168 ± 40 kPa) and atomic force microscopy elastic modulus (EAFM: 16 ± 2-387 ± 66 kPa) were readily adjusted. Because of their remarkably high swelling and mechanical properties, these tunable collagen hydrogels may be further exploited for the design of advanced dressings for chronic wound care. PMID:25411409

  9. Multi-scale mechanical characterization of highly swollen photo-activated collagen hydrogels

    PubMed Central

    Tronci, Giuseppe; Grant, Colin A.; Thomson, Neil H.; Russell, Stephen J.; Wood, David J.

    2015-01-01

    Biological hydrogels have been increasingly sought after as wound dressings or scaffolds for regenerative medicine, owing to their inherent biofunctionality in biological environments. Especially in moist wound healing, the ideal material should absorb large amounts of wound exudate while remaining mechanically competent in situ. Despite their large hydration, however, current biological hydrogels still leave much to be desired in terms of mechanical properties in physiological conditions. To address this challenge, a multi-scale approach is presented for the synthetic design of cyto-compatible collagen hydrogels with tunable mechanical properties (from the nano- up to the macro-scale), uniquely high swelling ratios and retained (more than 70%) triple helical features. Type I collagen was covalently functionalized with three different monomers, i.e. 4-vinylbenzyl chloride, glycidyl methacrylate and methacrylic anhydride, respectively. Backbone rigidity, hydrogen-bonding capability and degree of functionalization (F: 16 ± 12–91 ± 7 mol%) of introduced moieties governed the structure–property relationships in resulting collagen networks, so that the swelling ratio (SR: 707 ± 51–1996 ± 182 wt%), bulk compressive modulus (Ec: 30 ± 7–168 ± 40 kPa) and atomic force microscopy elastic modulus (EAFM: 16 ± 2–387 ± 66 kPa) were readily adjusted. Because of their remarkably high swelling and mechanical properties, these tunable collagen hydrogels may be further exploited for the design of advanced dressings for chronic wound care. PMID:25411409

  10. Crystal structure and mapping by site-directed mutagenesis of the collagen-binding epitope of an activated form of BM-40/SPARC/osteonectin.

    PubMed Central

    Sasaki, T; Hohenester, E; Göhring, W; Timpl, R

    1998-01-01

    The extracellular calcium-binding domain (positions 138-286) of the matrix protein BM-40 possesses a binding epitope of moderate affinity for several collagen types. This epitope was predicted to reside in helix alphaA and to be partially masked by helix alphaC. Here we show that deletion of helix alphaC produces a 10-fold increase in collagen affinity similar to that seen after proteolytic cleavage of this helix. The predicted removal of the steric constraint was clearly demonstrated by the crystal structure of the mutant at 2.8 A resolution. This constitutively activated mutant was used to map the collagen-binding site following alanine mutagenesis at 13 positions. Five residues were crucial for binding, R149 and N156 in helix alphaA, and L242, M245 and E246 in a loop region connecting the two EF hands of BM-40. These residues are spatially close and form a flat ring of 15 A diameter which matches the diameter of a triple-helical collagen domain. The mutations showed similar effects on binding to collagens I and IV, indicating nearly identical binding sites on both collagens. Selected mutations in the non-activated mutant DeltaI also reduced collagen binding, consistent with the same location of the epitope but in a more cryptic form in intact BM-40. PMID:9501084

  11. Bioengineered collagens

    PubMed Central

    Ramshaw, John AM; Werkmeister, Jerome A; Dumsday, Geoff J

    2014-01-01

    Mammalian collagen has been widely used as a biomedical material. Nevertheless, there are still concerns about the variability between preparations, particularly with the possibility that the products may transmit animal-based diseases. Many groups have examined the possible application of bioengineered mammalian collagens. However, translating laboratory studies into large-scale manufacturing has often proved difficult, although certain yeast and plant systems seem effective. Production of full-length mammalian collagens, with the required secondary modification to give proline hydroxylation, has proved difficult in E. coli. However, recently, a new group of collagens, which have the characteristic triple helical structure of collagen, has been identified in bacteria. These proteins are stable without the need for hydroxyproline and are able to be produced and purified from E. coli in high yield. Initial studies indicate that they would be suitable for biomedical applications. PMID:24717980

  12. Efficacy of Annona squamosa L in the synthesis of glycosaminoglycans and collagen during wound repair in streptozotocin induced diabetic rats.

    PubMed

    Ponrasu, Thangavel; Suguna, Lonchin

    2014-01-01

    The aim of this work was to find out the effects of Annona squamosa on the formation of glycosaminoglycans and collagen during wound healing in normal and diabetic rats. Diabetes induced rats were segregated into 4 groups, each containing six animals. Groups I and III served as the normal and diabetic control while groups II and IV served as normal and diabetic treated. The animals were treated with 200 μL of Annona squamosa extract topically. The granulation tissues formed were removed on the 8th day and the amount of glycosaminoglycans (GAGs) and collagen formed was evaluated by sequential extraction and SDSPAGE, respectively. Histological evaluation was also carried out using Masson's trichrome stain. In vitro wound healing efficacy of A. squamosa in human dermal fibroblast culture (HDF) was also carried out. The fibroblasts treated with varying concentrations of A. squamosa were examined for proliferation and closure of the wound area and photographed. A. squamosa increased cellular proliferation in HDF culture. The granulation tissues of treated wounds showed increased levels of glycosaminoglycans (P < 0.05) and collagen which were also confirmed by histopathology. The results strongly substantiate the beneficial effects of A. squamosa on the formation of glycosaminoglycans and collagen during wound healing.

  13. Type XIV Collagen Regulates Fibrillogenesis

    PubMed Central

    Ansorge, Heather L.; Meng, Xianmin; Zhang, Guiyun; Veit, Guido; Sun, Mei; Klement, John F.; Beason, David P.; Soslowsky, Louis J.; Koch, Manuel; Birk, David E.

    2009-01-01

    Type XIV collagen is a fibril-associated collagen with an interrupted triple helix. This collagen interacts with the fibril surface and has been implicated as a regulator of fibrillogenesis; however, a specific role has not been elucidated. Functional roles for type XIV collagen were defined utilizing a new type XIV collagen-deficient mouse line. This line was produced using a conventional targeted knock-out approach. Col14a1(–/–) mice were devoid of type XIV collagen, whereas heterozygous mice had reduced synthesis. Both mutant Col14a1 genotypes were viable with a grossly normal phenotype; however, mature skin exhibited altered mechanical properties. Prior to evaluating tendon fibrillogenesis in type XIV collagen-deficient mice, the developmental expression patterns were analyzed in wild-type flexor digitorum longus (FDL) tendons. Analyses of mRNA and protein expression indicated tissue-specific temporal expression that was associated with the early stages in fibrillogenesis. Ultrastructural analyses of wild-type and null tendons demonstrated premature fibril growth and larger fibril diameters in tendons from null mice at postnatal day 4 (P4). However, fibril structure in mature tendons was normal. Biomechanical studies established a direct structure/function relationship with reduced strength in P7-null tendons. However, the biomechanical properties in P60 tendons were comparable in null and wild-type mice. Our results indicate a regulatory function for type XIV collagen in early stages of collagen fibrillogenesis with tissue differences. PMID:19136672

  14. Peroxisome proliferator-activated receptor-gamma abrogates Smad-dependent collagen stimulation by targeting the p300 transcriptional coactivator.

    PubMed

    Ghosh, Asish K; Bhattacharyya, Swati; Wei, Jun; Kim, Suyeon; Barak, Yaacov; Mori, Yasuji; Varga, John

    2009-09-01

    Ligands of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) abrogate the stimulation of collagen gene transcription induced by transforming growth factor-beta (TGF-beta). Here, we delineate the mechanisms underlying this important novel physiological function for PPAR-gamma in connective tissue homeostasis. First, we demonstrated that antagonistic regulation of TGF-beta activity by PPAR-gamma ligands involves cellular PPAR-gamma, since 15-deoxy-Delta12,14-prostaglandin J(2) (15d-PGJ(2)) failed to block TGF-beta-induced responses in either primary cultures of PPAR-gamma-null murine embryonic fibroblasts, or in normal human skin fibroblasts with RNAi-mediated knockdown of PPAR-gamma. Next, we examined the molecular basis underlying the abrogation of TGF-beta signaling by PPAR-gamma in normal human fibroblasts in culture. The results demonstrated that Smad-dependent transcriptional responses were blocked by PPAR-gamma without preventing Smad2/3 activation. In contrast, the interaction between activated Smad2/3 and the transcriptional coactivator and histone acetyltransferase p300 induced by TGF-beta, and the accumulation of p300 on consensus Smad-binding DNA sequences and histone H4 hyperacetylation at the COL1A2 locus, were all prevented by PPAR-gamma. Wild-type p300, but not a mutant form of p300 lacking functional histone acetyltransferase, was able to restore TGF-beta-induced stimulation of COL1A2 in the presence of PPAR-gamma ligands. Collectively, these results indicate that PPAR-gamma blocked Smad-mediated transcriptional responses by preventing p300 recruitment and histone H4 hyperacetylation, resulting in the inhibition of TGF-beta-induced collagen gene expression. Pharmacological activation of PPAR-gamma thus may represent a novel therapeutic approach to target p300-dependent TGF-beta profibrotic responses such as stimulation of collagen gene expression.

  15. Effects of ultraviolet radiation on the type-I collagen protein triple helical structure: a method for measuring structural changes through optical activity.

    PubMed

    Majewski, Alexander J; Sanzari, Martin; Cui, Hong-Liang; Torzilli, Peter

    2002-03-01

    A detailed study of the effects of ultraviolet radiation on type-I collagen has been conducted. We have confirmed that exposure to ultraviolet radiation lowers the denaturation temperature of type-I collagen and that the triple helical state is destroyed provided that the radiation dose exceeds a threshold level, which is defined as the incident radiation dose that raises the sample temperature above the (lower) denaturation temperature. For incident radiation doses below threshold, the collagen molecule remains in a triple helical state. Denaturation is determined by changes in the optical activity of the collagen solution. Furthermore, a new instrument has been developed and tested to measure the optical rotatory dispersion properties of chiral molecules. The advantage of this instrument is that it enables a real-time measurement of the optical activity of chiral macromolecules while exposing samples to ultraviolet radiation and requiring no special sample preparation techniques. Using a differential measurement scheme, system errors have been minimized.

  16. Identification and Evaluation of Cryoprotective Peptides from Chicken Collagen: Ice-Growth Inhibition Activity Compared to That of Type I Antifreeze Proteins in Sucrose Model Systems.

    PubMed

    Du, Lihui; Betti, Mirko

    2016-06-29

    The ability of chicken collagen peptides to inhibit the growth of ice crystals was evaluated and compared to that of fish antifreeze proteins (AFPs). This ice inhibition activity was assessed using a polarized microscope by measuring ice crystal dimensions in a sucrose model system with and without collagen peptides after seven thermal cycles. The system was stabilized at -25 °C and cycled between -16 and -12 °C. Five candidate peptides with ice inhibition activity were identified using liquid chromatography and tandem mass spectrometry and were then synthesized. Their ice inhibition capacity was compared to that of type I AFPs in a 23% sucrose model system. Specific collagen peptides with certain amino acid sequences reduced the extent of ice growth by approximately 70% at a relatively low concentration (1 mg/mL). These results suggest that specific collagen peptides may act in a noncolligative manner, inhibiting ice crystal growth like type I AFPs, but less efficiently. PMID:27293017

  17. Collagenous gastroduodenitis on collagenous colitis.

    PubMed

    Stolte, M; Ritter, M; Borchard, F; Koch-Scherrer, G

    1990-07-01

    We report on a case of collagenous gastroduodenitis with concomitant collagenous colitis in a 75-year-old woman with watery diarrhea of approximately six months' standing. The step biopsy material obtained from the colon revealed continuous collagenous colitis with thickening of the basal membrane to 30 microns. The biopsies taken from the stomach and duodenum also revealed a band-like deposition of collagen in the duodenum (bulb and proximal portion of the descending portion) along the basal membrane of the lining epithelium, associated with partial atrophy of the villi. In the stomach, this band of collagen was located, parallel to the mucosal surface, at the level of the floor of the foveolae. PMID:2209504

  18. Upregulation of Collagen Expression via PPARβ/δ Activation in Aged Skin by Magnesium Lithospermate B from Salvia miltiorrhiza.

    PubMed

    Jung, Yu Ri; Lee, Eun Kyeong; Kim, Dae Hyun; Park, Chan Hum; Park, Min Hi; Jeong, Hyoung Oh; Yokozawa, Takako; Tanaka, Takashi; Im, Dong Soon; Kim, Nam Deuk; Yu, Byung Pal; Mo, Sang Hyun; Chung, Hae Young

    2015-08-28

    This study investigated the agonistic activity of magnesium lithospermate B (1), isolated from Salvia miltiorrhiza, on peroxisome proliferator-activated receptor (PPARβ/δ) and the expressions of collagen genes (COL1A1 and COL3A1) and transforming growth factor-β1 (TGF-β1) in models of skin aging. The action of compound 1 as a PPARβ/δ agonist was determined by reporter gene assay, immunostaining, and Western blotting. To determine the antiaging effects of compound 1 on skin, aged Sprague-Dawley rat skin and ultraviolet B (UVB)-irradiated human skin fibroblasts were used. The results show that 1 presented a marked enhancement of both nuclear protein levels and activity of PPARβ/δ in fibroblasts. In addition, 1 prevented downregulation of PPARβ/δ activity in aged rat skin and UVB-induced fibroblasts. Furthermore, 1 increased the expressions of COL1A1, COL3A1, and TGF-β1 in vivo and in a cell culture system. Therefore, the present study shows that compound 1 prevents collagen degradation in aged rat skin and UVB-exposed fibroblasts through PPARβ/δ activation. The therapeutic and cosmetic applications of compound 1 need further investigation. PMID:26280594

  19. Characterization of a non-fibrillar-related collagen in the mollusc Haliotis tuberculata and its biological activity on human dermal fibroblasts.

    PubMed

    Fleury, Christophe; Serpentini, Antoine; Kypriotou, Magdalini; Renard, Emmanuelle; Galéra, Philippe; Lebel, Jean-Marc

    2011-10-01

    In invertebrates, members of the collagen family have been found in various phyla. Surprisingly, in mollusc, little is known about such molecules. In this study, we characterize the full-length abalone type IV collagen and we analysed its biological effects on human fibroblast in order to gain insights about this molecule in molluscs and particularly clues about its roles. We screened a cDNA library of Haliotis tuberculata hemocytes. The expression pattern of the transcript is determined using real-time polymerase chain reaction and in situ hybridization. The close identity between α1(IV) C-terminal domain and the vertebrate homologue led us to produce, purify and test in vitro a recombinant protein corresponding to this region using human dermal fibroblasts cell culture. The biological effects were evaluated on proliferation and on differentiation. We found that the 5,334-bp open reading frame transcript encodes a protein of 1,777 amino acids, including an interrupted 1,502-residue collagenous domain and a 232-residue C-terminal non-collagenous domain. The expression pattern of this transcript is mainly found in the mantle and hemocytes. The recombinant protein corresponding α1(IV) C-terminal domain increased fibroblast proliferation by 69% and doubled collagen synthesis produced in primary cultures. This work provides the first complete primary structure of a mollusc non-fibrillar collagen chain and the biological effects of its C-terminal domain on human cells. In this study, we prove that the NC1 domain from a molluscan collagen can improve human fibroblast proliferation as well as differentiation.

  20. Collagencin, an antibacterial peptide from fish collagen: Activity, structure and interaction dynamics with membrane.

    PubMed

    Ennaas, Nadia; Hammami, Riadh; Gomaa, Ahmed; Bédard, François; Biron, Éric; Subirade, Muriel; Beaulieu, Lucie; Fliss, Ismail

    2016-04-29

    In this study, we first report characterization of collagencin, an antimicrobial peptide identified from fish collagen hydrolysate. The peptide completely inhibited the growth of Staphylococcus aureus at 1.88 mM. Although non-toxic up to 470 μM, collagencin was hemolytic at higher concentrations. The secondary structure of collagencin was mainly composed by β-sheet and β-turn as determined by CD measurements and molecular dynamics. The peptide is likely to form β-sheet structure under hydrophobic environments and interacts with both anionic (phosphatidylglycerol) and zwitterionic (phosphoethanolamine and phosphatidylcholine) lipids as shown with CD spectroscopy and molecular dynamics. The peptide formed several hydrogen bonds with both POPG and POPE lipids and remained at membrane-water interface, suggesting that collagencin antibacterial action follows a carpet mechanism. Collagenous fish wastes could be processed by enzymatic hydrolysis and transformed into products of high value having functional or biological properties. Marine collagens are a promising source of antimicrobial peptides with new implications in food safety and human health. PMID:27038545

  1. Marine Natural Meroterpenes: Synthesis and Antiproliferative Activity

    PubMed Central

    Simon-Levert, Annabel; Menniti, Christophe; Soulère, Laurent; Genevière, Anne-Marie; Barthomeuf, Chantal; Banaigs, Bernard; Witczak, Anne

    2010-01-01

    Meroterpenes are compounds of mixed biogenesis, isolated from plants, microorganisms and marine invertebrates. We have previously isolated and determined the structure for a series of meroterpenes extracted from the ascidian Aplidium aff. densum. Here, we demonstrate the chemical synthesis of three of them and their derivatives, and evaluate their biological activity on two bacterial strains, on sea urchin eggs, and on cancerous and healthy human cells. PMID:20390109

  2. Plasminogen Activator Inhibitor-1 and Vitronectin Expression Level and Stoichiometry Regulate Vascular Smooth Muscle Cell Migration through Physiological Collagen Matrices

    PubMed Central

    Garg, N.; Goyal, N.; Strawn, T. L.; Wu, J.; Mann, K. M.; Lawrence, D. A.; Fay, W. P.

    2010-01-01

    Summary Background Vascular smooth muscle cell (VSMC) migration is a critical process in arterial remodeling. Purified plasminogen activator inhibitor-1 (PAI-1) is reported to both promote and inhibit VSMC migration on 2-dimensional (D) surfaces. Objective To determine the effects of PAI-1 and vitronectin (VN) expressed by VSMC themselves on migration through physiological collagen matrices. Methods We studied migration of wild-type (WT), PAI-1-deficient, VN-deficient, PAI-1/VN doubly-deficient (DKO), and PAI-1-transgenic (Tg) VSMC through 3-D collagen gels. Results WT VSMC migrated significantly slower than PAI-1- and VN-deficient VSMC, but significantly faster than DKO VSMC. Experiments with recombinant PAI-1 suggested that basal VSMC PAI-1 expression inhibits migration by binding VN, which is secreted by VSMC and binds collagen. However, PAI-1-over-expressing Tg VSMC migrated faster than WT VSMC. Reconstitution experiments with recombinant PAI-1 mutants suggested that the pro-migratory effect of PAI-1 over-expression required its anti-plasminogen activator (PA) and LDL receptor-related protein (LRP) binding functions, but not VN binding. While promoting VSMC migration in the absence of PAI-1, VN inhibited the pro-migratory effect of active PAI-1. Conclusions In isolation, VN and PAI-1 are each pro-migratory. However, via formation of a high-affinity, non-motogenic complex, PAI-1 and VN each buffers the other's pro-migratory effect. The level of PAI-1 expression by VSMC and the concentration of VN in extracellular matrix are critical determinants of whether PAI-1 and VN promote or inhibit migration. These findings help to rectify previously conflicting reports and suggest that PAI-1/VN stoichiometry plays an important role in VSMC migration and vascular remodeling. PMID:20492459

  3. Cotransfected human chondrocytes: over-expression of IGF-I and SOX9 enhances the synthesis of cartilage matrix components collagen-II and glycosaminoglycans.

    PubMed

    Simental-Mendía, M; Lara-Arias, J; Álvarez-Lozano, E; Said-Fernández, S; Soto-Domínguez, A; Padilla-Rivas, G R; Martínez-Rodríguez, H G

    2015-12-01

    Damage to cartilage causes a loss of type II collagen (Col-II) and glycosaminoglycans (GAG). To restore the original cartilage architecture, cell factors that stimulate Col-II and GAG production are needed. Insulin-like growth factor I (IGF-I) and transcription factor SOX9are essential for the synthesis of cartilage matrix, chondrocyte proliferation, and phenotype maintenance. We evaluated the combined effect of IGF-I and SOX9 transgene expression on Col-II and GAG production by cultured human articular chondrocytes. Transient transfection and cotransfection were performed using two mammalian expression plasmids (pCMV-SPORT6), one for each transgene. At day 9 post-transfection, the chondrocytes that were over-expressing IGF-I/SOX9 showed 2-fold increased mRNA expression of the Col-II gene, as well as a 57% increase in Col-II protein, whereas type I collagen expression (Col-I) was decreased by 59.3% compared with controls. The production of GAG by these cells increased significantly compared with the controls at day 9 (3.3- vs 1.8-times, an increase of almost 83%). Thus, IGF-I/SOX9 cotransfected chondrocytes may be useful for cell-based articular cartilage therapies.

  4. Collagen structure: new tricks from a very old dog.

    PubMed

    Bella, Jordi

    2016-04-15

    The main features of the triple helical structure of collagen were deduced in the mid-1950s from fibre X-ray diffraction of tendons. Yet, the resulting models only could offer an average description of the molecular conformation. A critical advance came about 20 years later with the chemical synthesis of sufficiently long and homogeneous peptides with collagen-like sequences. The availability of these collagen model peptides resulted in a large number of biochemical, crystallographic and NMR studies that have revolutionized our understanding of collagen structure. High-resolution crystal structures from collagen model peptides have provided a wealth of data on collagen conformational variability, interaction with water, collagen stability or the effects of interruptions. Furthermore, a large increase in the number of structures of collagen model peptides in complex with domains from receptors or collagen-binding proteins has shed light on the mechanisms of collagen recognition. In recent years, collagen biochemistry has escaped the boundaries of natural collagen sequences. Detailed knowledge of collagen structure has opened the field for protein engineers who have used chemical biology approaches to produce hyperstable collagens with unnatural residues, rationally designed collagen heterotrimers, self-assembling collagen peptides, etc. This review summarizes our current understanding of the structure of the collagen triple helical domain (COL×3) and gives an overview of some of the new developments in collagen molecular engineering aiming to produce novel collagen-based materials with superior properties.

  5. Coumarin heterocyclic derivatives: chemical synthesis and biological activity.

    PubMed

    Medina, Fernanda G; Marrero, Joaquín G; Macías-Alonso, Mariana; González, Magdalena C; Córdova-Guerrero, Iván; Teissier García, Ariana G; Osegueda-Robles, Soraya

    2015-09-23

    This review highlights the broad range of science that has arisen from the synthesis of coumarin-linked and fused heterocycle derivatives. Specific topics include their synthesis and biological activity.

  6. Investigation of the effect of stress on the chemistry, metabolism, and biophysics of collagen

    NASA Technical Reports Server (NTRS)

    Houck, J. C.

    1973-01-01

    The research is reported concerning the effect of stress on the chemistry in the connective tissue of the rat. It was found that within a day after administration of cortisol (stress harmone), a significant amount of the insoluble collagen disappeared from the skin. It is concluded that the abrupt catabolism of cutaneous collagen releases peptides. These peptides are rapidly degraded to free amino acids which are active in effecting liver glycogen synthesis, and provide a major energy source to assist the animal in the fight or flight reaction. It is proposed that cutaneous collagen represents a reserve energy pool, which can be mobilized via stress harmones.

  7. The Activation of β1-integrin by Type I Collagen Coupling with the Hedgehog Pathway Promotes the Epithelial-Mesenchymal Transition in Pancreatic Cancer.

    PubMed

    Duan, Wanxing; Ma, Jiguang; Ma, Qingyong; Xu, Qinhong; Lei, Jianjun; Han, Liang; Li, Xuqi; Wang, Zheng; Wu, Zheng; Lv, Shifang; Ma, Zhenhua; Liu, Mouzhu; Wang, Fengfei; Wu, Erxi

    2014-01-01

    Pancreatic ductal adenocarcinoma (PDAC) is characterized by the excessive deposition of extracellular matrix (ECM), which is thought to contribute to this tumor's malignant behavior. However, the detailed mechanism and the contribution of excessive deposition of ECM in PDAC progression remain unclear. A better understanding of the mechanism involved in this process is essential for the design of new effective therapies. In this study, we demonstrated that pancreatic cancer cells exhibited increased proliferation and decreased apoptosis in response to type I collagen. In addition, PDAC cells exposed to type I collagen lost the expression of E-cadherin and increased expression of mesenchymal markers, including N-cadherin and vimentin. This epithelial- mesenchymal transition (EMT) was correlated with enhanced cell migration and invasiveness. Knockdown of β1-integrin abolished the effects induced by type I collagen, and further investigation revealed that type I collagen activates β1-integrin (marked by phosphorylation of β1 integrin downstream effectors, focal adhesion kinase [FAK], AKT, and ERK) accompanied by markedly up-regulation of Gli-1, a component of the Hedgehog (HH) pathway. Knockdown of Gli-1 reversed the effects of type I collagen on PDAC invasion and EMT. These results suggest that there is cross-talk between the β1-integrin signaling pathway and the HH pathway in pancreatic cancer and that activation of the HH pathway plays a key role in the type I collagen-induced effects on pancreatic cancer. PMID:24720337

  8. Perylenequinones: Isolation, Synthesis, and Biological Activity

    PubMed Central

    Mulrooey, Carol A.; O'Brien, Erin M.; Morgan, Barbara J.

    2013-01-01

    The perylenequinones are a novel class of natural products characterized by pentacyclic conjugated chromophore giving rise to photoactivity. Potentially useful light-activated biological activity, targeting protein kinase C (PKC), has been identified for several of the natural products. Recently discovered new members of this class of compound, as well as several related phenanthroperylenequinones, are reviewed. Natural product modifications that improve biological profiles, and avenues for the total synthesis of analogs, which are not available from the natural product series, are outlined. An overview of structure/function relationships is provided. PMID:24039544

  9. Biosynthesis, Synthesis and Biological Activities of Pyrrolobenzodiazepines

    PubMed Central

    Gerratana, Barbara

    2014-01-01

    Pyrrolobenzodiazepines (PBDs) are sequence selective DNA alkylating agents with remarkable antineoplastic activity. They are either naturally produced by actinomycetes or synthetically produced. The remarkable broad spectrum of activities of the naturally produced PBDs encouraged the synthesis of several PBDs, including dimeric and hybrid PBDs yielding to an improvement in the DNA binding sequence specificity and in the potency of this class of compounds. However, limitation in the chemical synthesis prevented the testing of one of the most potent PBDs, sibiromycin, a naturally produced glycosylated PBDs. Only recently the biosynthetic gene clusters for PBDs have been identified opening the doors to the production of glycosylated PBDs by mutasynthesis and biosynthetic engineering. The present review describes the recent studies on the biosynthesis of naturally produced pyrrolobenzodiazepines. In addition, it provides an overview on the isolation and characterization of naturally produced PBDs, on the chemical synthesis of PBDs, on the mechanism of DNA alkylation, and on the DNA binding affinity and cytotoxic properties of both naturally produced and synthetic pyrrolobenzodiazepines. PMID:20544978

  10. Collagen regulates transforming growth factor-β receptors of HL-1 cardiomyocytes through activation of stretch and integrin signaling.

    PubMed

    Lu, Yen-Yu; Lin, Yung-Kuo; Kao, Yu-Hsun; Chung, Cheng-Chih; Yeh, Yung-Hsin; Chen, Shih-Ann; Chen, Yi-Jen

    2016-10-01

    The extracellular matrix (ECM) and transforming growth factor-β (TGF)-β are important in cardiac fibrosis, however, the effects of the ECM on TGF‑β signaling remain to be fully elucidated. The aims of the present study were to evaluate the role of collagen in TGF‑β signaling and examine the underlying mechanisms. In the present study, western blot analysis was used to examine TGF‑β signaling in HL‑1 cells treated with and without (control) type I collagen (10 µg/ml), which was co‑administered with either an anti‑β1 integrin antibody (10 µg/ml) or a stretch‑activated channel inhibitor (gadolinium; 50 µM). Cell proliferation and adhesion assays were used to investigate the roles of integrin, mechanical stretch and mitogen‑activated protein kinases (MAPKs) on cell proliferation and adhesion. The type I collagen (10 µg/ml)‑treated HL‑1 cells were incubated with or without anti‑β1 integrin antibody (10 µg/ml), gadolinium (50 µM) or inhibitors of p38 (SB203580; 3 µM), extracellular signal‑regulated kinase (ERK; PD98059; 50 µM) and c‑Jun N‑terminal kinase (JNK; SP600125; 50 µM). Compared with the control cells, the collagen‑treated HL‑1 cells had lower expression levels of type I and type II TGF‑β receptors (TGFβRI and TGFβRII), with an increase in phosphorylated focal adhesion kinase (FAK), p38 and ERK1/2, and a decrease in JNK. Incubation with the anti‑β1 integrin antibody reversed the collagen‑induced downregulation of the expression of TGFβRII and phosphorylated FAK. Gadolinium downregulated the expression levels of TGFβRI and small mothers against decapentaplegic (Smad)2/3, and decreased the levels of phosphorylated p38, ERK1/2 and JNK. In addition, gadolinium reversed the collagen‑induced activation of p38 and ERK1/2. In the presence of gadolinium and anti‑β1 integrin antibody, collagen regulated the expression levels of TGFβRI, TGFβRII and Smad2/3, but did not alter the phosphorylation

  11. Mechanism for Adhesion G Protein-Coupled Receptor GPR56-Mediated RhoA Activation Induced By Collagen III Stimulation

    PubMed Central

    Luo, Rong; Jeong, Sung-Jin; Yang, Annie; Wen, Miaoyun; Saslowsky, David E.; Lencer, Wayne I.; Araç, Demet; Piao, Xianhua

    2014-01-01

    GPR56 is a member of the adhesion G protein-coupled receptor (GPCR) family. Despite the importance of GPR56 in brain development, where mutations cause a devastating human brain malformation called bilateral frontoparietal polymicrogyria (BFPP), the signaling mechanism(s) remain largely unknown. Like many other adhesion GPCRs, GPR56 is cleaved via a GPCR autoproteolysis-inducing (GAIN) domain into N- and C-terminal fragments (GPR56N and GPR56C); however, the biological significance of this cleavage is elusive. Taking advantage of the recent identification of a GPR56 ligand and the presence of BFPP-associated mutations, we investigated the molecular mechanism of GPR56 signaling. We demonstrate that ligand binding releases GPR56N from the membrane-bound GPR56C and triggers the association of GPR56C with lipid rafts and RhoA activation. Furthermore, one of the BFPP-associated mutations, L640R, does not affect collagen III-induced lipid raft association of GPR56. Instead, it specifically abolishes collagen III-mediated RhoA activation. Together, these findings reveal a novel signaling mechanism that may apply to other members of the adhesion GPCR family. PMID:24949629

  12. One-step synthesis of collagen hybrid gold nanoparticles and formation on Egyptian-like gold-plated archaeological ivory.

    PubMed

    Spadavecchia, Jolanda; Apchain, Emilande; Albéric, Marie; Fontan, Elisabeth; Reiche, Ina

    2014-08-01

    A one-step method is reported to synthesize hybrid gold nanoparticles (AuNPs) by reduction of HAuCl4 in acetic solution in the presence of collagen (Col), dicarboxylic acid-terminated polyethylene glycol (PEG), and cetyltetrammonium bromide (CTAB) mixed with hydoxyapatite (HAP) as surfactants. Such formation process of AuNPs was shown to be responsible for purple stains naturally formed on Egyptianizing archaeological gilded ivories from 8th BC Syria. The understanding of this formation mechanism, which most likely involves a step with hybrid AuNPs, allows the establishing of an authenticity marker of ancient gold-plated ivories.

  13. A therapeutic approach for diabetic wound healing using biotinylated GHK incorporated collagen matrices.

    PubMed

    Arul, Vadivel; Kartha, Reena; Jayakumar, Rajadas

    2007-01-01

    Chronically elevated blood glucose levels result in reduced leukocyte function and cell malnutrition, which contribute to a high rate of wound infection and associated healing problems in diabetic patients. In the present study, the role of biotinylated GHK peptide (BioGHK) incorporated collagen biomaterial was tested for wound healing in diabetic rats. The rate of wound contraction and the levels of collagen, uronic acid, protein and DNA in the granulation tissue were determined. Further, the concentration of nitric oxide and other skin antioxidants was also monitored during the study. In diabetic rats treated with BioGHK incorporated collagen (Peptide Incorporated Collagen--PIC), the healing process was hastened with an increased rate of wound contraction. Glutathione (GSH) and ascorbic acid levels in the skin of streptozotocin-induced diabetic rats were higher in the PIC group as compared to control (Untreated) and collagen (Collagen Film--CF) treated groups. Superoxide dismutase (SOD) and catalase (CAT) activity was altered in all the groups. In vitro fibroblast cell culture studies suggest that PIC promotes fibroblast growth. Histological evaluation by haematoxylin-eosin and Masson's trichrome method revealed epithelialization, increased synthesis of collagen and activation of fibroblasts and mast cells in the PIC group. This study provides a rationale for the topical application of BioGHK incorporated collagen as a feasible and productive approach to support diabetic wound healing. PMID:17049946

  14. Adenoviral overexpression and small interfering RNA suppression demonstrate that plasminogen activator inhibitor-1 produces elevated collagen accumulation in normal and keloid fibroblasts.

    PubMed

    Tuan, Tai-Lan; Hwu, Paul; Ho, Wendy; Yiu, Peter; Chang, Richard; Wysocki, Annette; Benya, Paul D

    2008-11-01

    Keloids are tumor-like skin scars that grow as a result of the aberrant healing of skin injuries, with no effective treatment. We provide new evidence that both overexpression of plasminogen activator inhibitor-1 (PAI-1) and elevated collagen accumulation are intrinsic features of keloid fibroblasts and that these characteristics are causally linked. Using seven strains each of early passage normal and keloid fibroblasts, the keloid strains exhibited inherently elevated collagen accumulation and PAI-1 expression in serum-free, 0.1% ITS+ culture; larger increases in these parameters occurred when cells were cultured in 3% serum. To demonstrate a causal relationship between PAI-1 overexpression and collagen accumulation, normal fibroblasts were infected with PAI-1-expressing adenovirus. Such cells exhibited a two- to fourfold increase in the accumulation of newly synthesized collagen in a viral dose-dependent fashion in both monolayers and fibrin gel, provisional matrix-like cultures. Three different PAI-1-targeted small interfering RNAs, alone or in combination, produced greater than an 80% PAI-1 knockdown and reduced collagen accumulation in PAI-1-overexpressing normal or keloid fibroblasts. A vitronectin-binding mutant of PAI-1 was equipotent with wild-type PAI-1 in inducing collagen accumulation, whereas a complete protease inhibitor mutant retained approximately 50% activity. Thus, PAI-1 may use more than its protease inhibitory activity to control keloid collagen accumulation. PAI-1-targeted interventions, such as small interfering RNA and lentiviral short hairpin RNA-containing microRNA sequence suppression reported here, may have therapeutic utility in the prevention of keloid scarring.

  15. Inhibition of collagen-induced platelet aggregation by antibodies to distinct types of collagens.

    PubMed Central

    Balleisen, L; Nowack, H; Gay, S; Timpl, R

    1979-01-01

    Aggregation of platelets by fibrils formed from collagens type I, II and III could be inhibited by coating the fibrils with anti-collagen antibodies or Fab fragments. Similar results were obtained in a clot-retraction assay. Inhibition was achieved with stoichiometric amounts of antibodies and was specific for each type of collagen. Aggregation caused by a mixture of type-I and -III collagens could only be inhibited by a mixture of antibodies against both collagens. The data show that each interstitial collagen is capable of interacting with platelets and do not support the concept of an outstanding activity of type-III collagen. Images PLATE 1 PMID:395952

  16. Collagenous gastritis.

    PubMed

    Jin, Xiaoyi; Koike, Tomoyuki; Chiba, Takashi; Kondo, Yutaka; Ara, Nobuyuki; Uno, Kaname; Asano, Naoki; Iijima, Katsunori; Imatani, Akira; Watanabe, Mika; Shirane, Akio; Shimosegawa, Tooru

    2013-09-01

    In the present paper, we report a case of rare collagenous gastritis. The patient was a 25-year-old man who had experienced nausea, abdominal distention and epigastralgia since 2005. Esophagogastroduodenoscopy (EGD) carried out at initial examination by the patient's local doctor revealed an extensively discolored depression from the upper gastric body to the lower gastric body, mainly including the greater curvature, accompanied by residual mucosa with multiple islands and nodularity with a cobblestone appearance. Initial biopsies sampled from the nodules and accompanying atrophic mucosa were diagnosed as chronic gastritis. In August, 2011, the patient was referred to Tohoku University Hospital for observation and treatment. EGD at our hospital showed the same findings as those by the patient's local doctor. Pathological findings included a membranous collagen band in the superficial layer area of the gastric mucosa, which led to a diagnosis of collagenous gastritis. Collagenous gastritis is an extremely rare disease, but it is important to recognize its characteristic endoscopic findings to make a diagnosis. PMID:23363075

  17. Expression of receptor activator of nuclear factor-κB ligand is related to sex differences in collagen-induced arthritis.

    PubMed

    Ding, Zheng; Wang, Qi; Pan, Xiongxiong; Zhu, Qin; Lu, Hao; Wang, Kunpeng; Ni, Xuhao; Lu, Yunjie; Gu, Jian

    2015-10-01

    Osteoclasts are responsible for bone destruction in rheumatoid arthritis, and women show greater disease activity and functional disability than men. This study aimed to examine differences in the pathogenesis of collagen-induced arthritis and osteoclastogenesis between female and male mice in vivo and in vitro. Female mice exhibited worse disease progression and increased osteoclastogenesis, as measured by tartrate-resistant acid phosphatase (TRAP) staining than male mice. Significantly higher levels of CD11b(+) cells were detected in the bone marrow of female mice than that of male mice. Furthermore, the mRNA expression of receptor activator of nuclear factor-κB ligand was higher in female mice that were immunized with or without collagen II. These findings highlighted sex differences in arthritis morbidity and suggested that female mice are more likely to develop arthritis than male mice. Further studies are needed to investigate the mechanisms of sex differences in collagen-induced arthritis. PMID:25863233

  18. Betulin Phosphonates; Synthesis, Structure, and Cytotoxic Activity.

    PubMed

    Chrobak, Elwira; Bębenek, Ewa; Kadela-Tomanek, Monika; Latocha, Małgorzata; Jelsch, Christian; Wenger, Emmanuel; Boryczka, Stanisław

    2016-01-01

    Betulin derivatives are a widely studied group of compounds of natural origin due to their wide spectrum of biological activities. This paper describes new betulin derivatives, containing a phosphonate group. The allyl-vinyl isomerization and synthesis of acetylenic derivatives have been reported. Structural identification of products as E and Z isomers has been carried out using ¹H-, (13)C-, (31)P-NMR, and crystallographic analysis. The crystal structure in the orthorhombic space group and analysis of crystal packing contacts for 29-diethoxyphosphoryl-28-cyclopropylpropynoyloxy-lup-20E(29)-en-3β-ol 8a are reported. All new compounds were tested in vitro for their antiproliferative activity against human T47D (breast cancer), SNB-19 (glioblastoma), and C32 (melanoma) cell lines. PMID:27571057

  19. Anti-inflammatory activity of lycopene isolated from Chlorella marina on type II collagen induced arthritis in Sprague Dawley rats.

    PubMed

    Renju, G L; Muraleedhara Kurup, G; Saritha Kumari, C H

    2013-04-01

    The role of commercially available lycopene (all-trans) from tomato in controlling arthritis has been reported. Even though many reports are available that the cis form of lycopene is more biologically active, no report seems to be available on lycopene (cis and trans) isolated from an easily available and culturable sources. In the present study, the anti-arthritic effect of lycopene (cis and trans) from the algae Chlorella marina (AL) has been compared with lycopene (all-trans) from tomato (TL) and indomethacin (Indo). Arthritis (CIA) was developed in male Sprague dawley rats by collagen and the following parameters were studied. The activities of inflammatory marker enzymes like cyclooxygenase (COX), lipoxygenase (LOX) and myeloperoxidase (MPO) were found to be decreased on treatment with AL when compared to TL and Indo. Changes in Erythrocyte sedimentation rate (ESR), white blood cell (WBC) count, red blood cells (RBC) count, hemoglobin (Hb), C-reactive protein (CRP), rheumatoid factor (RF), and ceruloplasmin levels observed in the blood of arthritic animals were brought back to normal by AL when compared to TL and Indo. Histopathology of paw and joint tissues showed marked reduction in edema on supplementation of AL. Thus these results indicate the potential beneficiary effect of algal lycopene on collagen induced arthritis in rats when compared to TL and even to the commonly used anti-inflammatory drug indomethacin. Therefore lycopene from C. marina would be recommended as a better natural source with increased activity and without side effects in the treatment of anti-inflammatory diseases. PMID:23237458

  20. Platelet 12-lipoxygenase activation via glycoprotein VI: involvement of multiple signaling pathways in agonist control of H(P)ETE synthesis.

    PubMed

    Coffey, Marcus J; Jarvis, Gavin E; Gibbins, Jonathan M; Coles, Barbara; Barrett, Natasha E; Wylie, Oliver R E; O'Donnell, Valerie B

    2004-06-25

    Lipoxygenases (LOX) contribute to vascular disease and inflammation through generation of bioactive lipids, including 12-hydro(pero)xyeicosatetraenoic acid (12-H(P)ETE). The physiological mechanisms that acutely control LOX product generation in mammalian cells are uncharacterized. Human platelets that contain a 12-LOX isoform (p12-LOX) were used to define pathways that activate H(P)ETE synthesis in the vasculature. Collagen and collagen-related peptide (CRP) (1 to 10 microg/mL) acutely induced platelet 12-H(P)ETE synthesis. This implicated the collagen receptor glycoprotein VI (GPVI), which signals via the immunoreceptor-based activatory motif (ITAM)-containing FcRgamma chain. Conversely, thrombin only activated at high concentrations (> 0.2 U/mL), whereas U46619 and ADP alone were ineffective. Collagen or CRP-stimulated 12-H(P)ETE generation was inhibited by staurosporine, PP2, wortmannin, BAPTA/AM, EGTA, and L-655238, implicating src-tyrosine kinases, PI3-kinase, Ca2+ mobilization, and p12-LOX translocation. In contrast, protein kinase C (PKC) inhibition potentiated 12-H(P)ETE generation. Finally, activation of the immunoreceptor tyrosine-based inhibitory motif (ITIM)-containing platelet endothelial cell adhesion molecule (PECAM-1) inhibited p12-LOX product generation. This study characterizes a receptor-dependent pathway for 12-H(P)ETE synthesis via the collagen receptor GPVI, which is negatively regulated by PECAM-1 and PKC, and demonstrates a novel link between immune receptor signaling and lipid mediator generation in the vasculature. PMID:15142951

  1. Sc65-Null Mice Provide Evidence for a Novel Endoplasmic Reticulum Complex Regulating Collagen Lysyl Hydroxylation

    PubMed Central

    Weis, MaryAnn; Rai, Jyoti; Hudson, David M.; Dimori, Milena; Zimmerman, Sarah M.; Hogue, William R.; Swain, Frances L.; Burdine, Marie S.; Mackintosh, Samuel G.; Tackett, Alan J.; Suva, Larry J.; Eyre, David R.

    2016-01-01

    Collagen is a major component of the extracellular matrix and its integrity is essential for connective tissue and organ function. The importance of proteins involved in intracellular collagen post-translational modification, folding and transport was recently highlighted from studies on recessive forms of osteogenesis imperfecta (OI). Here we describe the critical role of SC65 (Synaptonemal Complex 65, P3H4), a leprecan-family member, as part of an endoplasmic reticulum (ER) complex with prolyl 3-hydroxylase 3. This complex affects the activity of lysyl-hydroxylase 1 potentially through interactions with the enzyme and/or cyclophilin B. Loss of Sc65 in the mouse results in instability of this complex, altered collagen lysine hydroxylation and cross-linking leading to connective tissue defects that include low bone mass and skin fragility. This is the first indication of a prolyl-hydroxylase complex in the ER controlling lysyl-hydroxylase activity during collagen synthesis. PMID:27119146

  2. Sc65-Null Mice Provide Evidence for a Novel Endoplasmic Reticulum Complex Regulating Collagen Lysyl Hydroxylation.

    PubMed

    Heard, Melissa E; Besio, Roberta; Weis, MaryAnn; Rai, Jyoti; Hudson, David M; Dimori, Milena; Zimmerman, Sarah M; Kamykowski, Jeffrey A; Hogue, William R; Swain, Frances L; Burdine, Marie S; Mackintosh, Samuel G; Tackett, Alan J; Suva, Larry J; Eyre, David R; Morello, Roy

    2016-04-01

    Collagen is a major component of the extracellular matrix and its integrity is essential for connective tissue and organ function. The importance of proteins involved in intracellular collagen post-translational modification, folding and transport was recently highlighted from studies on recessive forms of osteogenesis imperfecta (OI). Here we describe the critical role of SC65 (Synaptonemal Complex 65, P3H4), a leprecan-family member, as part of an endoplasmic reticulum (ER) complex with prolyl 3-hydroxylase 3. This complex affects the activity of lysyl-hydroxylase 1 potentially through interactions with the enzyme and/or cyclophilin B. Loss of Sc65 in the mouse results in instability of this complex, altered collagen lysine hydroxylation and cross-linking leading to connective tissue defects that include low bone mass and skin fragility. This is the first indication of a prolyl-hydroxylase complex in the ER controlling lysyl-hydroxylase activity during collagen synthesis.

  3. Withaferin-A Reduces Type I Collagen Expression In Vitro and Inhibits Development of Myocardial Fibrosis In Vivo

    PubMed Central

    Challa, Azariyas A.; Vukmirovic, Milica; Blackmon, John; Stefanovic, Branko

    2012-01-01

    Type I collagen is the most abundant protein in the human body. Its excessive synthesis results in fibrosis of various organs. Fibrosis is a major medical problem without an existing cure. Excessive synthesis of type I collagen in fibrosis is primarily due to stabilization of collagen mRNAs. We recently reported that intermediate filaments composed of vimentin regulate collagen synthesis by stabilizing collagen mRNAs. Vimentin is a primary target of Withaferin-A (WF-A). Therefore, we hypothesized that WF-A may reduce type I collagen production by disrupting vimentin filaments and decreasing the stability of collagen mRNAs. This study is to determine if WF-A exhibits anti-fibrotic properties in vitro and in vivo and to elucidate the molecular mechanisms of its action. In lung, skin and heart fibroblasts WF-A disrupted vimentin filaments at concentrations of 0.5–1.5 µM and reduced 3 fold the half-lives of collagen α1(I) and α2(I) mRNAs and protein expression. In addition, WF-A inhibited TGF-β1 induced phosphorylation of TGF-β1 receptor I, Smad3 phosphorylation and transcription of collagen genes. WF-A also inhibited in vitro activation of primary hepatic stellate cells and decreased their type I collagen expression. In mice, administration of 4 mg/kg WF-A daily for 2 weeks reduced isoproterenol-induced myocardial fibrosis by 50%. Our findings provide strong evidence that Withaferin-A could act as an anti-fibrotic compound against fibroproliferative diseases, including, but not limited to, cardiac interstitial fibrosis. PMID:22900077

  4. Adipose tissue macrophages in insulin-resistant subjects are associated with collagen VI and fibrosis and demonstrate alternative activation.

    PubMed

    Spencer, Michael; Yao-Borengasser, Aiwei; Unal, Resat; Rasouli, Neda; Gurley, Catherine M; Zhu, Beibei; Peterson, Charlotte A; Kern, Philip A

    2010-12-01

    Adipose tissue macrophages are associated with insulin resistance and are linked to changes in the extracellular matrix. To better characterize adipose macrophages, the extracellular matrix, and adipocyte-macrophage interactions, gene expression from adipose tissue and the stromal vascular fraction was assessed for markers of inflammation and fibrosis, and macrophages from obese and lean subjects were counted and characterized immunohistochemically. Coculture experiments examined the effects of adipocyte-macrophage interaction. Collagen VI gene expression was associated with insulin sensitivity and CD68 (r = -0.56 and 0.60, P < 0.0001) and with other markers of inflammation and fibrosis. Compared with adipose tissue from lean subjects, adipose tissue from obese subjects contained increased areas of fibrosis, which correlated inversely with insulin sensitivity (r = -0.58, P < 0.02) and positively with macrophage number (r = 0.70, P < 0.01). Although macrophages in crownlike structures (CLS) were more abundant in obese adipose tissue, the majority of macrophages were associated with fibrosis and were not organized in CLS. Macrophages in CLS were predominantly M1, but most other macrophages, particularly those in fibrotic areas, were M2 and also expressed CD150, a marker of M2c macrophages. Coculture of THP-1 macrophages with adipocytes promoted the M2 phenotype, with a lower level of IL-1 expression and a higher ratio of IL-10 to IL-12. Transforming growth factor-β (TGF-β) was more abundant in M2 macrophages and was further increased by coculture with adipocytes. Downstream effectors of TGF-β, such as plasminogen activator inhibitor-1, collagen VI, and phosphorylated Smad, were increased in macrophages and adipocytes. Thus adipose tissue of insulin-resistant humans demonstrated increased fibrosis, M2 macrophage abundance, and TGF-β activity.

  5. Collagen binding to OSCAR: the odd couple.

    PubMed

    An, Bo; Brodsky, Barbara

    2016-02-01

    In this issue of Blood, Zhou et al reported the high-resolution structure of the collagen-activated osteoclast-associated receptor (OSCAR) bound to a collagen model peptide. Together with binding studies, the results confirm a novel recognition mechanism for collagen by immunoglobulin-like motifs. PMID:26847065

  6. Synthesis and antiviral activity of substituted quercetins.

    PubMed

    Thapa, Mahendra; Kim, Yunjeong; Desper, John; Chang, Kyeong-Ok; Hua, Duy H

    2012-01-01

    Influenza viruses are important pathogens that cause respiratory infections in humans and animals. In addition to vaccination, antiviral drugs against influenza virus play a significant role in controlling viral infections by reducing disease progression and virus transmission. Plant derived polyphenols are associated with antioxidant activity, anti-carcinogenic, and cardio- and neuro-protective actions. Some polyphenols, such as resveratrol and epigallocatechin gallate (EGCG), showed significant anti-influenza activity in vitro and/or in vivo. Recently we showed that quercetin and isoquercetin (quercetin-3-β-d-glucoside), a glucoside form of quercetin, significantly reduced the replication of influenza viruses in vitro and in vivo (isoquercetin). The antiviral effects of isoquercetin were greater than that of quercetin with lower IC(50) values and higher in vitro therapeutic index. Thus, we investigated the synthesis and antiviral activities of various quercetin derivatives with substitution of C3, C3', and C5 hydroxyl functions with various phenolic ester, alkoxy, and aminoalkoxy moieties. Among newly synthesized compounds, quercetin-3-gallate which is structurally related to EGCG showed comparable antiviral activity against influenza virus (porcine H1N1 strain) to that of EGCG with improved in vitro therapeutic index.

  7. : Synthesis, Characterization, and Enhanced Photocatalytic Activity

    NASA Astrophysics Data System (ADS)

    Gao, Xiaoming; Fu, Feng; Li, Wenhong

    2014-12-01

    3D hierarchical microspheres of Cu-loaded Bi2WO6 are successfully prepared by the hydrothermal synthesis method on a large scale. The as-prepared samples are characterized by UV-Vis DRS, BET, XRD, XPS, and SEM. The results reveal that the light absorption of Cu-loaded Bi2WO6 has higher intensity in the visible range and a bathochromic shift of the absorption edge compared to that of pure Bi2WO6. The photocatalytic activity is evaluated by phenol removal from aqueous solution under visible-light irradiation. The results demonstrate that loaded Cu significantly enhances the photocatalytic activity of Bi2WO6, for the loaded Cu acts as the electron receptor on the surface of Bi2WO6, and inhibits the recombination of photogenerated electron-hole. The content of loaded Cu has an impact on the catalytic activity, and the 1.0 wt.% Cu-loaded Bi2WO6 exhibits the best photocatalytic activity in the degradation of phenol. Furthermore, the reaction kinetics of phenol removal from aqueous solution over the Cu-loaded Bi2WO6 is established by the way of the Langmuir-Hinshelwood model. The results indicate that the process of photodegradation of phenol on Cu-loaded Bi2WO6 match the Langmuir-Hinshelwood kinetic model.

  8. Regulation of collagen biosynthesis in cultured bovine aortic smooth muscle cells

    SciTech Connect

    Stepp, M.A.

    1986-01-01

    Aortic smooth muscles cells have been implicated in the etiology of lesions which occur in atherosclerosis and hypertension. Both diseases involve proliferation of smooth muscle cells and accumulation of excessive amounts of extracellular matrix proteins, including collagen type I and type III produced by the smooth muscle cells. To better understand the sites of regulation of collagen biosynthesis and to correlate these with the growth rate of the cells, cultured bovine aortic smooth muscle cells were studied as a function of the number of days (3 to 14) in second passage. Cells grew rapidly up to day 6 when confluence was reached. The total incorporation of (/sup 3/H)-proline into proteins was highest at day 3 and decreased to a constant level after the cultures reached confluence. In contrast, collagen protein production was lowest before confluence and continued to increase over the entire time course of the experiments. cDNA clones for the ..cap alpha..1 and ..cap alpha..2 chains of type I and the ..cap alpha..1 chain of type III collagen were used to quantitate the steady state level of collagen mRNAs. RNA was tested in a cell-free translation system. Changes in the translational activity of collagen mRNAs parallelled the observed increases in collagen protein production. Thus, at later time points, collagen mRNAs are more active in directing synthesis of preprocollagens, even though less collagen mRNA is present. The conclusion is that the site of regulation of the expression of collagen genes is a function of the growth rate of cultured smooth muscle cells.

  9. Synthesis of α-collagen fragments and research of their influence on the degree of hydration of a model of epidermis

    PubMed Central

    Grobelna, Beata; Maćkiewicz, Zbigniew

    2013-01-01

    Introduction In recent years the interest into areas of science, such as cosmetology, dermatology, pharmacology or aesthetic medicine has increased significantly. Scientists are more frequently looking for ingredients that affect the skin's condition and slow down the aging process. Practically every year, the scientists discover a number of new chemical substances (both natural and synthetic) that can be potentially used to manufacture cosmetics. Aim To evaluate the influence of selected peptides derived from α-collagen fragments on the degree of hydration of a model of epidermis isolated from a pig. Material and methods The synthesis of selected cosmetic oligopeptides were performed manually, on the solid medium, using procedure of SPPS (solid phase peptide synthesis). Following components: aqua, carbomer, glycerine, phenonip, D-panthenol, dimethicone and triethanolamine were used to prepare a reference hydrogel masks. Both the number of components and the composition of hydrogels have been developed individually for the purposes of this research. For this study the skin from a domestic pig was used. The degree of the skin hydration was measured with the SKINTEST plus camera, which uses the latest semiconductor technology. Results During the study the absorption of hydrogels with peptides was faster than that of the reference hydrogel mask. The combination of hydrophilic properties of the peptide with hydrophobic properties of Palm enabled receiving an amphiphilic structure. Such molecules are considered to be able to penetrate the corneum barrier with the greatest ease. Conclusions The results showed that the modified compounds have contributed to water retention in the cells, thereby increasing the degree of hydration of the biological material. PMID:24278040

  10. Anti-inflammatory activities of light emitting diode irradiation on collagen-induced arthritis in mice (a secondary publication)

    PubMed Central

    Ohta, Mitsuhiro; Sato, Yusuke; Abiko, Yoshimitsu

    2014-01-01

    Background and aims: Rheumatoid arthritis (RA) is an auto-immune disease afflicting multiple joints of the body, where as a result of the increase in inflammatory cytokines and tissue destructive factors such as matrix metalloproteinase (MMP)-3, deterioration of the bones and cartilages of the joints occurs. The present investigation was carried out to study the anti-inflammatory activities of light emitting diode (LED) irradiation on hind paw inflammation in collagen-induced arthritis (CIA) mice models. Materials and method: RA in the CIA mouse model was induced by immunization of DBA/1J mice with intradermal injections of an emulsion of bovine type II collagen and complete Freund's adjuvant. A total of 20 CIA mice were subdivided into the following groups: control group, CIA group and 2 groups of LED irradiated CIA mice (LED groups) (n=5 per group). The mouse knee joint area in the LED groups (the 570 nm and 940 nm groups) was irradiated with LED energy, three times a week for 500 s per session over 8 weeks at a dose of 5 J/cm2. The hind paw swelling was assessed by the increase in hind paw thickness. The serum levels of the inflammatory cytokines and arthritic factor MMP-3 were determined with an enzyme-linked immunosorbent assay (ELISA). Results: In the LED-570 and LED-940 groups at 4 weeks after arthritis induction, the swelling inhibition index was 18.1±4.9 and 29.3±4.0 respectively. Interleukin (IL)-1β, IL-6 and MMP-3 serum levels were significantly lower in the LED-940 group. Conclusions: LED irradiation, particularly in the near-infrared was effective for inhibition of the inflammatory reactions caused by RA. PMID:25368445

  11. Optimizing an Intermittent Stretch Paradigm Using ERK1/2 Phosphorylation Results in Increased Collagen Synthesis in Engineered Ligaments

    PubMed Central

    Paxton, Jennifer Z.; Hagerty, Paul; Andrick, Jonathan J.

    2012-01-01

    Dynamic mechanical input is believed to play a critical role in the development of functional musculoskeletal tissues. To study this phenomenon, cyclic uniaxial mechanical stretch was applied to engineered ligaments using a custom-built bioreactor and the effects of different stretch frequency, amplitude, and duration were determined. Stretch acutely increased the phosphorylation of p38 (3.5±0.74-fold), S6K1 (3.9±0.19-fold), and ERK1/2 (2.45±0.32-fold). The phosphorylation of ERK1/2 was dependent on time, rather than on frequency or amplitude, within these constructs. ERK1/2 phosphorylation was similar following stretch at frequencies from 0.1 to 1 Hz and amplitudes from 2.5% to 15%, whereas phosphorylation reached maximal levels at 10 min of stretch and returned toward basal within 60 min of stretch. Following a single 10-min bout of cyclic stretch, the cells remained refractory to a second stretch for up to 6 h. Using the phosphorylation of ERK1/2 as a guide, the optimum stretch paradigm was hypothesized to be 10 min of stretch at 2.5% of resting length repeated every 6 h. Consistent with this hypothesis, 7 days of stretch using this optimized intermittent stretch program increased the collagen content of the grafts more than a continuous stretch program (CTL=3.1%±0.44%; CONT=4.8%±0.30%; and INT=5.9%±0.56%). These results suggest that short infrequent bouts of loading are optimal for improving engineered tendon and ligament physiology. PMID:21902469

  12. Cystamine immobilization on TiO 2 film surfaces and the influence on inhibition of collagen-induced platelet activation

    NASA Astrophysics Data System (ADS)

    Zhou, Yujuan; Weng, Yajun; Zhang, Liping; Jing, Fengjuan; Huang, Nan; Chen, Junying

    2011-12-01

    Poor haemocompatibility is a main issue of artificial cardiovascular materials in clinical application. Nitric oxide (NO), produced by vascular endothelial cells, is a well known inhibitor of platelet adhesion and activation. Thus, NO-releasing biomaterials are beneficial for improving haemocompatibility of blood-contacting biomedical devices. In this paper, a novel method was developed for enhancement of haemocompatibility by exploiting endogenous NO donors. TiO 2 films were firstly synthesized on Si (1 0 0) wafers via unbalanced magnetron sputtering technology, and then polydopamine was grafted on TiO 2 films and used as a linker for further immobilization of cystamine. The obtained surfaces were characterized by scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) analysis. NO generation is evaluated by saville-griess reagents, and it shows that cystamine immobilized samples are able to catalytically generate NO by decomposing endogenous S-nitrosothiols (RSNO). In vitro platelet adhesion results reveal that cystamine modified surfaces can inhibit collagen-induced platelet activation. ELISA analysis reveals that cGMP in platelets obviously increases on cystamine immobilized surface, which suggests the reducing of platelet activation is through NO/cGMP signal channel. It can be concluded that cystamine immobilized surface shows better blood compatibility by catalyzing NO release from the endogenous NO donor. It may be a promising method for improvement of haemocompatibility of blood-contacting implants.

  13. Exosomes from hypoxic endothelial cells have increased collagen crosslinking activity through up-regulation of lysyl oxidase-like 2.

    PubMed

    de Jong, Olivier G; van Balkom, Bas W M; Gremmels, Hendrik; Verhaar, Marianne C

    2016-02-01

    Exosomes are important mediators of intercellular communication. Additionally, they contain a variety of components capable of interacting with the extracellular matrix (ECM), including integrins, matrix metalloproteinases and members of the immunoglobin superfamily. Despite these observations, research on exosome-ECM interactions is limited. Here, we investigate whether the exosome-associated lysyl oxidase family member lysyl oxidase-like 2 (LOXL2) is involved in ECM remodelling. We found that LOXL2 is present on the exterior of endothelial cell (EC)-derived exosomes, placing it in direct vicinity of the ECM. It is up-regulated twofold in EC-derived exosomes cultured under hypoxic conditions. Intact exosomes from hypoxic EC and LOXL2 overexpressing EC show increased activity in a fluorometric lysyl oxidase enzymatic activity assay as well as in a collagen gel contraction assay. Concordantly, knockdown of LOXL2 in exosome-producing EC in both normal and hypoxic conditions reduces activity of exosomes in both assays. Our findings show for the first time that ECM crosslinking by EC-derived exosomes is mediated by LOXL2 under the regulation of hypoxia, and implicate a role for exosomes in hypoxia-regulated focal ECM remodelling, a key process in both fibrosis and wound healing.

  14. Positive impact of IGF-1-coupled nanoparticles on the differentiation potential of human chondrocytes cultured on collagen scaffolds

    PubMed Central

    Pasold, Juliane; Zander, Kathleen; Heskamp, Benjamin; Grüttner, Cordula; Lüthen, Frank; Tischer, Thomas; Jonitz-Heincke, Anika; Bader, Rainer

    2015-01-01

    Purpose In the present study, silica nanoparticles (sNP) coupled with insulin-like growth factor 1 (IGF-1) were loaded on a collagen-based scaffold intended for cartilage repair, and the influence on the viability, proliferation, and differentiation potential of human primary articular chondrocytes was examined. Methods Human chondrocytes were isolated from the hyaline cartilage of patients (n=4, female, mean age: 73±5.1 years) undergoing primary total knee joint replacement. Cells were dedifferentiated and then cultivated on a bioresorbable collagen matrix supplemented with fluorescent sNP coupled with IGF-1 (sNP–IGF-1). After 3, 7, and 14 days of cultivation, cell viability and integrity into the collagen scaffold as well as metabolic cell activity and synthesis rate of matrix proteins (collagen type I and II) were analyzed. Results The number of vital cells increased over 14 days of cultivation, and the cells were able to infiltrate the collagen matrix (up to 120 μm by day 7). Chondrocytes cultured on the collagen scaffold supplemented with sNP–IGF-1 showed an increase in metabolic activity (5.98-fold), and reduced collagen type I (1.58-fold), but significantly increased collagen type II expression levels (1.53-fold; P=0.02) after 7 days of cultivation compared to 3 days. In contrast, chondrocytes grown in a monolayer on plastic supplemented with sNP-IGF-1 had significantly lower metabolic activity (1.32-fold; P=0.007), a consistent amount of collagen type I, and significantly reduced collagen type II protein expression (1.86-fold; P=0.001) after 7 days compared to 3 days. Conclusion Collagen-based scaffolds enriched with growth factors, such as IGF-1 coupled to nanoparticles, represent an improved therapeutic intervention for the targeted and controlled treatment of articular cartilage lesions. PMID:25709437

  15. Syk interacts with tyrosine-phosphorylated proteins in human platelets activated by collagen and cross-linking of the Fc gamma-IIA receptor.

    PubMed Central

    Yanaga, F; Poole, A; Asselin, J; Blake, R; Schieven, G L; Clark, E A; Law, C L; Watson, S P

    1995-01-01

    Activation of human platelets by cross-linking of the platelet low-affinity IgG receptor, the Fc gamma receptor IIA (Fc gamma-RIIA), or by collagen is associated with rapid phosphorylation on tyrosine of the non-receptor tyrosine kinase syk. Phosphorylation is still observed, albeit sometimes reduced, in the presence of a combination of a protein kinase C inhibitor, Ro 31-8220, and the intracellular calcium chelator, BAPTA-AM, demonstrating independence from phosphoinositide-specific phospholipase C (PLC) activity. In contrast, the combination of Ro 31-8220 and BAPTA-AM completely inhibits phosphorylation of syk in thrombin-stimulated platelets. Phosphorylation of syk increases its autophosphorylation activity measured in a kinase assay performed on syk immunoprecipitates. Fc gamma-RIIA also undergoes phosphorylation in syk immunoprecipitates from platelets activated by cross-linking of Fc gamma-RIIA but not by collagen, suggesting that it associates with the kinase. Consistent with this, tyrosine-phosphorylated Fc gamma-RIIA is precipitated by a glutathione S-transferase (GST) fusion protein containing the tandem src homology (SH2) domains of syk from Fc gamma-RIIA- but not collagen-activated cells. Two uncharacterized tyrosine-phosphorylated proteins of 40 and 65 kDa are uniquely precipitated by a GST fusion protein containing the tandem syk-SH2 domains in collagen-stimulated platelets. A peptide based on the antigen recognition activation motif (ARAM) of Fc gamma-RIIA, and phosphorylated on the two tyrosine residues found within this region, selectively binds syk from lysates of resting platelets; this interaction is not seen with a non-phosphorylated peptide. Kinase assays on Fc gamma-RIIA immunoprecipitates reveal the constitutive association of an unidentified kinase activity in resting cells which phosphorylates a 67 kDa protein. Syk is not detected in Fc gamma-RIIA immunoprecipitates from resting cells but associates with the receptor following activation

  16. Study of HMG-CoA Reductase Inhibition Activity of the Hydrolyzed Product of Snakehead Fish (Channa striata) Skin Collagen with 50 kDa Collagenase from Bacillus licheniformis F11.4.

    PubMed

    Virginia, Agnes; Rachmawati, Heni; Riani, Catur; Retnoningrum, Debbie S

    2016-01-01

    Bioactive peptides produced from enzymatic hydrolysis fibrous protein have been proven to have several biological activities. Previous study showed that the hydrolysis product of snakehead fish skin collagen with 26 kDa collagenase from Bacillus licheniformis F11.4 showed HMG-CoA (HMGR) inhibition activity. The aim of this research was to determine the ability of the hydrolysis product produced from snakehead fish skin collagen hydrolysed by 50 kDa collagenase from B. licheniformis F11.4 in inhibiting HMGR activity. Snakehead fish skin collagen was extracted using an acid method and collagenase was produced from B. licheniformis F11.4 using half-strength Luria Bertani (LB) medium containing 5% collagen. Crude collagenase was concentrated and fractionated using the DEAE Sephadex A-25 column eluted with increasing gradient concentrations of NaCl. Collagen, collagenase, and fractions were analyzed using SDS-PAGE and collagenolytic activity was analyzed by the zymography method. Collagenase with 50 kDa molecular weight presented in fraction one was used to hydrolyze the collagen. The reaction was done in 18 hours at 50°C. The hydrolysis product using 3.51 μg collagen and 9 ng collagenase showed 25.8% inhibition activity against pravastatin. This work shows for the first time that the hydrolysis product of snakehead fish skin collagen and 50 kDa collagenase from B. licheniformis F11.4 has potential as an anticholesterol agent. PMID:27110500

  17. Study of HMG-CoA Reductase Inhibition Activity of the Hydrolyzed Product of Snakehead Fish (Channa striata) Skin Collagen with 50 kDa Collagenase from Bacillus licheniformis F11.4

    PubMed Central

    Virginia, Agnes; Rachmawati, Heni; Riani, Catur; Retnoningrum, Debbie S.

    2016-01-01

    Bioactive peptides produced from enzymatic hydrolysis fibrous protein have been proven to have several biological activities. Previous study showed that the hydrolysis product of snakehead fish skin collagen with 26 kDa collagenase from Bacillus licheniformis F11.4 showed HMG-CoA (HMGR) inhibition activity. The aim of this research was to determine the ability of the hydrolysis product produced from snakehead fish skin collagen hydrolysed by 50 kDa collagenase from B. licheniformis F11.4 in inhibiting HMGR activity. Snakehead fish skin collagen was extracted using an acid method and collagenase was produced from B. licheniformis F11.4 using half-strength Luria Bertani (LB) medium containing 5% collagen. Crude collagenase was concentrated and fractionated using the DEAE Sephadex A-25 column eluted with increasing gradient concentrations of NaCl. Collagen, collagenase, and fractions were analyzed using SDS-PAGE and collagenolytic activity was analyzed by the zymography method. Collagenase with 50 kDa molecular weight presented in fraction one was used to hydrolyze the collagen. The reaction was done in 18 hours at 50°C. The hydrolysis product using 3.51 μg collagen and 9 ng collagenase showed 25.8% inhibition activity against pravastatin. This work shows for the first time that the hydrolysis product of snakehead fish skin collagen and 50 kDa collagenase from B. licheniformis F11.4 has potential as an anticholesterol agent. PMID:27110500

  18. Study of HMG-CoA Reductase Inhibition Activity of the Hydrolyzed Product of Snakehead Fish (Channa striata) Skin Collagen with 50 kDa Collagenase from Bacillus licheniformis F11.4.

    PubMed

    Virginia, Agnes; Rachmawati, Heni; Riani, Catur; Retnoningrum, Debbie S

    2016-01-01

    Bioactive peptides produced from enzymatic hydrolysis fibrous protein have been proven to have several biological activities. Previous study showed that the hydrolysis product of snakehead fish skin collagen with 26 kDa collagenase from Bacillus licheniformis F11.4 showed HMG-CoA (HMGR) inhibition activity. The aim of this research was to determine the ability of the hydrolysis product produced from snakehead fish skin collagen hydrolysed by 50 kDa collagenase from B. licheniformis F11.4 in inhibiting HMGR activity. Snakehead fish skin collagen was extracted using an acid method and collagenase was produced from B. licheniformis F11.4 using half-strength Luria Bertani (LB) medium containing 5% collagen. Crude collagenase was concentrated and fractionated using the DEAE Sephadex A-25 column eluted with increasing gradient concentrations of NaCl. Collagen, collagenase, and fractions were analyzed using SDS-PAGE and collagenolytic activity was analyzed by the zymography method. Collagenase with 50 kDa molecular weight presented in fraction one was used to hydrolyze the collagen. The reaction was done in 18 hours at 50°C. The hydrolysis product using 3.51 μg collagen and 9 ng collagenase showed 25.8% inhibition activity against pravastatin. This work shows for the first time that the hydrolysis product of snakehead fish skin collagen and 50 kDa collagenase from B. licheniformis F11.4 has potential as an anticholesterol agent.

  19. MT1-MMP promotes cell growth and ERK activation through c-Src and paxillin in three-dimensional collagen matrix

    SciTech Connect

    Takino, Takahisa; Tsuge, Hisashi; Ozawa, Terumasa; Sato, Hiroshi

    2010-06-11

    Membrane-type 1 matrix metalloproteinase (MT1-MMP) is essential for tumor invasion and growth. We show here that MT1-MMP induces extracellular signal-regulated kinase (ERK) activation in cancer cells cultured in collagen gel, which is indispensable for their proliferation. Inhibition of MT1-MMP by MMP inhibitor or small interfering RNA suppressed activation of focal adhesion kinase (FAK) and ERK in MT1-MMP-expressing cancer cells, which resulted in up-regulation of p21{sup WAF1} and suppression of cell growth in collagen gel. Cell proliferation was also abrogated by the inhibitor against ERK pathway without affecting FAK phosphorylation. MT1-MMP and integrin {alpha}{sub v}{beta}{sub 3} were shown to be involved in c-Src activation, which induced FAK and ERK activation in collagen gel. These MT1-MMP-mediated signal transductions were paxillin dependent, as knockdown of paxillin reduced cell growth and ERK activation, and co-expression of MT1-MMP with paxillin induced ERK activation. The results suggest that MT1-MMP contributes to proliferation of cancer cells in the extracellular matrix by activating ERK through c-Src and paxillin.

  20. Alternative activation of macrophages and pulmonary fibrosis are modulated by scavenger receptor, macrophage receptor with collagenous structure.

    PubMed

    Murthy, Shubha; Larson-Casey, Jennifer L; Ryan, Alan J; He, Chao; Kobzik, Lester; Carter, A Brent

    2015-08-01

    Alternative activation of alveolar macrophages is linked to fibrosis following exposure to asbestos. The scavenger receptor, macrophage receptor with collagenous structure (MARCO), provides innate immune defense against inhaled particles and pathogens; however, a receptor for asbestos has not been identified. We hypothesized that MARCO acts as an initial signaling receptor for asbestos, polarizes macrophages to a profibrotic M2 phenotype, and is required for the development of asbestos-induced fibrosis. Compared with normal subjects, alveolar macrophages isolated from patients with asbestosis express higher amounts of MARCO and have greater profibrotic polarization. Arginase 1 (40-fold) and IL-10 (265-fold) were higher in patients. In vivo, the genetic deletion of MARCO attenuated the profibrotic environment and pulmonary fibrosis in mice exposed to chrysotile. Moreover, alveolar macrophages from MARCO(-/-) mice polarize to an M1 phenotype, whereas wild-type mice have higher Ym1 (>3.0-fold) and nearly 7-fold more active TGF-β1 in bronchoalveolar lavage (BAL) fluid (BALF). Arg(432) and Arg(434) in domain V of MARCO are required for the polarization of macrophages to a profibrotic phenotype as mutation of these residues reduced FIZZ1 expression (17-fold) compared with cells expressing MARCO. These observations demonstrate that a macrophage membrane protein regulates the fibrotic response to lung injury and suggest a novel target for therapeutic intervention.

  1. EhPAK2, a novel p21-activated kinase, is required for collagen invasion and capping in Entamoeba histolytica.

    PubMed

    Arias-Romero, Luis Enrique; de Jesús Almáraz-Barrera, Ma; Díaz-Valencia, Juan Daniel; Rojo-Domínguez, Arturo; Hernandez-Rivas, Rosaura; Vargas, Miguel

    2006-09-01

    p21-activated kinases (PAKs) are a highly conserved family of enzymes that are activated by Rho GTPases. All PAKs contain an N-terminal Cdc42/Rac interacting binding (CRIB) domain, which confers binding to these GTPases, and a C-terminal kinase domain. In addition, some PAKs such as Cla4p, Skm1p and Pak2p contain an N-terminal pleckstrin homology (PH) domain and form a distinct group of PAK proteins involved in cell morphology, cell-cycle and gene transcription. Here, we describe a novel p21-activated kinase, denominated EhPAK2, on the parasitic protozoan Entamoeba histolytica. This is the first reported Entamoeba PAK member that contains a N-terminal PH domain and a highly conserved CRIB domain. EhPAK2 CRIB domain shares 29% of amino acid identity and 53% of amino acid homology with these of DdPAKC from Dictyostelium discoideum and Cla4p from Saccharomyces cerevisiae and binds in vitro and in vivo to EhRacA GTPase. This domain also possesses the conserved residues His123, Phe134 and Trp141, which are important for the interaction with the effector loop and strand beta2 of the GTPase; and the residues Met121 and Phe145, which are specific for the interaction of EhPAK2 with EhRacA. Functional studies of EhPAK2 showed that its C-terminal kinase domain had activity toward myelin basic protein. Cellular studies showed that Entamoeba trophozoites transfected with the vector pExEhNeo/kinase-myc, had a 90% decrease in the ability to invade a collagen matrix as well as severe defects in capping, suggesting the involvement of EhPAK2 in these cellular processes.

  2. Collagenous gastritis and collagenous colitis: a report with sequential histological and ultrastructural findings.

    PubMed

    Pulimood, A B; Ramakrishna, B S; Mathan, M M

    1999-06-01

    The case is reported of a young adult man with collagenous gastritis, an extremely rare disorder with only three case reports in the English literature, who subsequently presented with collagenous colitis. Sequential gastric biopsies showed a notable increase in thickness of the subepithelial collagen band. Ultrastructural study of gastric and rectal mucosa showed the characteristic subepithelial band composed of haphazardly arranged collagen fibres, prominent degranulating eosinophils, and activated pericryptal fibroblasts. PMID:10323893

  3. Measurement of whole body cellular and collagen nitrogen, potassium, and other elements by neutron activation and whole body counting

    SciTech Connect

    James, H.M.; Fabricius, P.J.; Dykes, P.W.

    1987-09-01

    Whole body nitrogen can be measured by neutron activation analysis with an acceptable radiation dose; it is an index of body protein which, in normal subjects, is 65% cellular protein and 35% extracellular connective collagen. Whole body potassium can be measured by whole body counting without irradiating the subject; it is an index of body cell mass. We measured whole body nitrogen, potassium, extracellular water, intracellular water, and fat-folds. The differences between 37 malnourished patients and five normal subjects suggested that the patients had 9 kg less cell mass than normal, but no difference in extracellular mass. Measurements were made on eight patients before and after 14 days of total parenteral nutrition; balance of nitrogen intake and excretion also was measured. The changes were consistent with mean increases of 3 kg of cellular mass and 3 kg of fat with no change of extracellular mass. The accuracy and sensitivity of the whole body measurements need further confirmation for use in patients with changing body composition. Where tissue wasting is largely from the cellular compartment, potassium could be a more sensitive index of wasting than nitrogen. Multielement analysis of nitrogen, potassium, chlorine, and carbon will probably be valuable in elucidating body composition in malnutrition.

  4. Synthesis of (-)-arctigenin derivatives and their anticancer activity.

    PubMed

    Gui-Rong, Chen; Li-Ping, Cai; De-Qiang, Dou; Ting-Guo, Kang; Hong-Fu, Li; Fu-Rui, Li; Ning, Jiang

    2012-01-01

    The natural dibenzylbutyrolactone type lignanolide (-)-arctigenin, which was prepared from fructus arctii, showed obvious anticancer activity. The synthesis of four new (-)-arctigenin derivatives and their anticancer bioactivities were examined. The structures of the four new synthetic derivatives were elucidated.

  5. Synthesis and in vitro antiproliferative activities of quinoline derivatives.

    PubMed

    Broch, Sidonie; Aboab, Bettina; Anizon, Fabrice; Moreau, Pascale

    2010-04-01

    The synthesis of new di- and trimeric quinoline derivatives is described as well as their in vitro antiproliferative activities toward a human fibroblast primary culture and two human solid cancer cell lines (MCF-7 and PA 1).

  6. Type I Collagen and Collagen Mimetics as Angiogenesis Promoting Superpolymers

    SciTech Connect

    Twardowski, T.; Fertala, A.; Orgel, J.P.R.O.; San Antonio, J.D.

    2008-07-18

    Angiogenesis, the development of blood vessels from the pre-existing vasculature, is a key component of embryogenesis and tissue regeneration. Angiogenesis also drives pathologies such as tumor growth and metastasis, and hemangioma development in newborns. On the other hand, promotion of angiogenesis is needed in tissues with vascular insufficiencies, and in bioengineering, to endow tissue substitutes with appropriate microvasculatures. Therefore, much research has focused on defining mechanisms of angiogenesis, and identifying pro- and anti-angiogenic molecules. Type I collagen, the most abundant protein in humans, potently stimulates angiogenesis in vitro and in vivo. Crucial to its angiogenic activity appears to be ligation and possibly clustering of endothelial cell (EC) surface {alpha}1{beta}1/{alpha}2{beta}1 integrin receptors by the GFPGER502-507 sequence of the collagen fibril. However, additional aspects of collagen structure and function that may modulate its angiogenic properties are discussed. Moreover, type I collagen and fibrin, another angiogenic polymer, share several structural features. These observations suggest strategies for creating 'angiogenic superpolymers', including: modifying type I collagen to influence its biological half-life, immunogenicity, and integrin binding capacity; genetically engineering fibrillar collagens to include additional integrin binding sites or angiogenic determinants, and remove unnecessary or deleterious sequences without compromising fibril integrity; and exploring the suitability of poly(ortho ester), PEG-lysine copolymer, tubulin, and cholesteric cuticle as collagen mimetics, and suggesting means of modifying them to display ideal angiogenic properties. The collagenous and collagen mimetic angiogenic superpolymers described here may someday prove useful for many applications in tissue engineering and human medicine.

  7. Non Insulin Dependent Diabetes in Sand Rat (Psammomys obesus) and Production of Collagen in Cultured Aortic Smooth Muscle Cells. Influence of Insulin

    PubMed Central

    Bourdillon, M. C.; Dahmani, Y.; Bekkhoucha, F.

    2001-01-01

    In this report, we have shown that the standard laboratory diet administered to Psammomys obesus (sand rat) from Beni Abbes in Algeria, induced a non-insulin dependant diabetes, characterised by increase of body weight (p<0.001) as well as hyperinsulinemia, hyperglycemia and hypercholesterolemia. In cultured aortic smooth muscle cells (SMC) of sand rats, type I and type III collagen biosynthesis and insulin effects, at low dose, on these parameters were investigated. In all experimental conditions of cultured SMC study, The α chains of type I collagen were analysed by immunoblotting in media and cells. Metabolic radiolabelling and Immunochemical procedures revealed that, in diabetic state, synthetic SMC (SMCs) actively produce type I and III collagen which are synthesised in the cells and secreted in the medium; type I collagen was predominant as compared with type III collagen. Diabetes enhanced the collagen synthesis. Low dose of Insulin added to the medium, during 48h of incubation, induced a marked reduction in the synthesis of collagen types, especially type I collagen. PMID:12369725

  8. The effect of anisotropic collagen-GAG scaffolds and growth factor supplementation on tendon cell recruitment, alignment, and metabolic activity

    PubMed Central

    Caliari, Steven R.; Harley, Brendan A.C.

    2014-01-01

    Current surgical and tissue engineering approaches for treating tendon injuries have shown limited success, suggesting the need for new biomaterial strategies. Here we describe the development of an anisotropic collagen-glycosaminoglycan (CG) scaffold and use of growth factor supplementation strategies to create a 3D platform for tendon tissue engineering. We fabricated cylindrical CG scaffolds with aligned tracks of ellipsoidal pores that mimic the native physiology of tendon by incorporating a directional solidification step into a conventional lyophilization strategy. By modifying the freezing temperature, we created a homologous series of aligned CG scaffolds with constant relative density and degree of anisotropy but a range of pore sizes (55–243 μm). Equine tendon cells showed greater levels of attachment, metabolic activity, and alignment as well as less cell-mediated scaffold contraction, when cultured in anisotropic scaffolds compared to an isotropic CG scaffold control. The anisotropic CG scaffolds also provided critical contact guidance cues for cell alignment. While tendon cells were randomly oriented in the isotropic control scaffold and the transverse (unaligned) plane of the anisotropic scaffolds, significant cell alignment was observed in the direction of the contact guidance cues in the longitudinal plane of the anisotropic scaffolds. Scaffold pore size was found to significantly influence tendon cell viability, proliferation, penetration into the scaffold, and metabolic activity in a manner predicted by cellular solids arguments. Finally, the addition of the growth factors PDGF-BB and IGF-1 to aligned CG scaffolds was found to enhance tendon cell motility, viability, and metabolic activity in dose-dependent manners. This work suggests a composite strategy for developing bioactive, 3D material systems for tendon tissue engineering. PMID:21550653

  9. Imaging Denatured Collagen Strands In vivo and Ex vivo via Photo-triggered Hybridization of Caged Collagen Mimetic Peptides

    PubMed Central

    Li, Yang; Foss, Catherine A.; Pomper, Martin G.; Yu, S. Michael

    2014-01-01

    Collagen is a major structural component of the extracellular matrix that supports tissue formation and maintenance. Although collagen remodeling is an integral part of normal tissue renewal, excessive amount of remodeling activity is involved in tumors, arthritis, and many other pathological conditions. During collagen remodeling, the triple helical structure of collagen molecules is disrupted by proteases in the extracellular environment. In addition, collagens present in many histological tissue samples are partially denatured by the fixation and preservation processes. Therefore, these denatured collagen strands can serve as effective targets for biological imaging. We previously developed a caged collagen mimetic peptide (CMP) that can be photo-triggered to hybridize with denatured collagen strands by forming triple helical structure, which is unique to collagens. The overall goals of this procedure are i) to image denatured collagen strands resulting from normal remodeling activities in vivo, and ii) to visualize collagens in ex vivo tissue sections using the photo-triggered caged CMPs. To achieve effective hybridization and successful in vivo and ex vivo imaging, fluorescently labeled caged CMPs are either photo-activated immediately before intravenous injection, or are directly activated on tissue sections. Normal skeletal collagen remolding in nude mice and collagens in prefixed mouse cornea tissue sections are imaged in this procedure. The imaging method based on the CMP-collagen hybridization technology presented here could lead to deeper understanding of the tissue remodeling process, as well as allow development of new diagnostics for diseases associated with high collagen remodeling activity. PMID:24513868

  10. Targeting and mimicking collagens via triple helical peptide assembly

    PubMed Central

    Li, Yang; Yu, S. Michael

    2013-01-01

    As the major structural component of the extracellular matrix, collagen plays a crucial role in tissue development and regeneration. Since structural and metabolic abnormalities of collagen are associated with numerous debilitating diseases and pathologic conditions, the ability to target collagens of diseased tissues could lead to new diagnostics and therapeutics. Collagen is also a natural biomaterial widely used in drug delivery and tissue engineering, and construction of synthetic collagen-like materials is gaining interests in the biomaterials community. The unique triple helical structure of collagen has been explored for targeting collagen strands, and for engineering collagen-like functional assemblies and conjugates. This review focuses on the forefront of research activities in the use of the collagen mimetic peptide for both targeting and mimicking collagens via its triple helix mediated strand hybridization and higher order assembly. PMID:24210894

  11. Posttranscriptional regulation of collagen alpha1(I) mRNA in hepatic stellate cells.

    PubMed Central

    Stefanovic, B; Hellerbrand, C; Holcik, M; Briendl, M; Aliebhaber, S; Brenner, D A

    1997-01-01

    The hepatic stellate cell (HSC) is the primary cell responsible for the dramatic increase in the synthesis of type I collagen in the cirrhotic liver. Quiescent HSCs contain a low level of collagen alpha1(I) mRNA, while activated HSCs contain about 60- to 70-fold more of this mRNA. The transcription rate of the collagen alpha1(I) gene is only two fold higher in activated HSCs than in quiescent HSCs. In assays using actinomycin D or 5,6-dichlorobenzimidazole riboside collagen alpha1(I) mRNA has estimated half-lives of 1.5 h in quiescent HSCs and 24 h in activated HSCs. Thus, this 16-fold change in mRNA stability is primarily responsible for the increase in collagen alpha1(I) mRNA steady-state level in activated HSCs. We have identified a novel RNA-protein interaction targeted to the C-rich sequence in the collagen alpha1(I) mRNA 3' untranslated region (UTR). This sequence is localized 24 nucleotides 3' to the stop codon. In transient transfection experiments, mutation of this sequence diminished accumulation of an mRNA transcribed from a collagen alpha1(I) minigene and in stable transfections decreased the half-life of collagen alpha1(I) minigene mRNA. Binding to the collagen alpha1(I) 3' UTR is present in cytoplasmic extracts of activated but not quiescent HSCs. It contains as a subunit alphaCP, which is also found in the complex involved in stabilization of alpha-globin mRNA. The auxiliary factors necessary to promote binding of alphaCP to the collagen 3' UTR are distinct from the factors necessary for binding to the alpha-globin sequence. Since alphaCP is expressed in both quiescent and activated HSCs, these auxiliary factors are responsible for the differentially expressed RNA-protein interaction at the collagen alpha1(I) mRNA 3' UTR. PMID:9271398

  12. Amiloride inhibits rat mucosal ornithine decarboxylase activity and DNA synthesis

    SciTech Connect

    Ulrich-Baker, M.G.; Wang, P.; Fitzpatrick, L.; Johnson, L.R. )

    1988-03-01

    Refeeding fasted rats induces a dramatic trophic response in gastrointestinal mucosa and is associated with elevations in both rate of DNA synthesis and ornithine decarboxylase (ODC) activity. The signal for these increases is unknown. Amiloride prevents cell alkalinization by blocking Na{sup +}-H{sup +} exchange at apical epithelial cell membranes. In study 1, rats were fasted 48 h, treated with amiloride (0.5 to 500 mg/kg), and refed for 4 h. Refeeding increased ODC activities in the jejunal mucosa (X8) and liver (X19) but not in the oxyntic gland mucosa. In the jejunum, but not the liver, the activation of ODC was completely abolished by 100 mg/kg amiloride. In study 2, the rate of DNA synthesis was determine by measuring the rate of ({sup 3}H)thymidine incorporation 16 h after refeeding. Refeeding resulted in significantly increased rates of DNA synthesis over fasted levels, and amiloride at 100 mg/kg significantly reduced the elevations in the jejenum and liver. In conclusion, amiloride inhibits the postprandial increases in jejunal ODC activity and DNA synthesis in the jejunum and liver. The results indicate that (1) the Na{sup +}-H{sup +} antiport is essential to the increased ODC activity in the jejunum and liver after a meal and (2) increases in DNA synthesis and their suppression by amiloride are not necessary linked to ODC activity.

  13. [Collagenous crystalloids and collagenous spherules in salivary gland tumors. A light microscopy and immunohistochemistry study].

    PubMed

    Skálová, A; Michal, M; Leivo, I

    1993-04-01

    In a series of 354 salivary gland tumors, the morphological and immunohistochemical study of two distinctive types of extracellular matrix deposits was carried out. First, collagenous crystalloids, distinct spherical crystalloids composed of radially arranged needle-shaped collagen fibres, were found in twelve cases of benign salivary gland tumors. Second, collagenous spherules, globoid structures often showing concentric lamellar or radial pattern, were found in 46 cases of both benign and malignant salivary gland tumors. Immunohistochemically, collagenous crystalloids and collagenous spherules contain varying amounts of type I and III collagens, proteoglycans and elastic fibres but not collagen types II and VI. Strong linear deposition of basement membrane proteins, collagen type IV and laminin, surrounded collagenous spherules. Discontinuous patchy deposits of both proteins were, however, found near collagenous crystalloids. The cells surrounding these collagenous crystalloids and collagenous spherules showed immunohistochemical and morphological features of modified myoepithelial cells. Our observations may improve a terminology of the structures in question. Proposed active role of modified myoepithelial cells in the origin of these extracellular deposits still remains open for discussion.

  14. Three-dimensional, bioactive, biodegradable, polymer-bioactive glass composite scaffolds with improved mechanical properties support collagen synthesis and mineralization of human osteoblast-like cells in vitro.

    PubMed

    Lu, Helen H; El-Amin, Saadiq F; Scott, Kimberli D; Laurencin, Cato T

    2003-03-01

    In the past decade, tissue engineering-based bone grafting has emerged as a viable alternative to biological and synthetic grafts. The biomaterial component is a critical determinant of the ultimate success of the tissue-engineered graft. Because no single existing material possesses all the necessary properties required in an ideal bone graft, our approach has been to develop a three dimensional (3-D), porous composite of polylactide-co-glycolide (PLAGA) and 45S5 bioactive glass (BG) that is biodegradable, bioactive, and suitable as a scaffold for bone tissue engineering (PLAGA-BG composite). The objectives of this study were to examine the mechanical properties of a PLAGA-BG matrix, to evaluate the response of human osteoblast-like cells to the PLAGA-BG composite, and to evaluate the ability of the composite to form a surface calcium phosphate layer in vitro. Structural and mechanical properties of PLAGA-BG were measured, and the formation of a surface calcium phosphate layer was evaluated by surface analysis methods. The growth and differentiation of human osteoblast-like cells on PLAGA-BG were also examined. A hypothesis was that the combination of PLAGA with BG would result in a biocompatible and bioactive composite, capable of supporting osteoblast adhesion, growth and differentiation, with mechanical properties superior to PLAGA alone. The addition of bioactive glass granules to the PLAGA matrix resulted in a structure with higher compressive modulus than PLAGA alone. Moreover, the PLAGA-BA composite was found to be a bioactive material, as it formed surface calcium phosphate deposits in a simulated body fluid (SBF), and in the presence of cells and serum proteins. The composite supported osteoblast-like morphology, stained positively for alkaline phosphatase, and supported higher levels of Type I collagen synthesis than tissue culture polystyrene controls. We have successfully developed a degradable, porous, polymer bioactive glass composite possessing

  15. Reconstruction of #7 facial cleft with distraction-assisted in situ osteogenesis (DISO): role of recombinant human bone morphogenetic protein-2 with Helistat-activated collagen implant.

    PubMed

    Carstens, Michael H; Chin, Martin; Ng, Theodore; Tom, William K

    2005-11-01

    A case involving concomitant presentation of a #7 lateral facial cleft with a complete cleft of the ipsilateral lip, alveolus, and palate is presented. The mandibular defect was Pruzansky III with a foreshortened body, absent ramus and absent masseter. Taking advantage of developmental field theory, reconstruction of the osseous defect was undertaken using the autogenous periosteum as a source of mesenchymal stem cells. Expansion of the periosteum was followed by implantation of Helistat (Integra Life Sciences, Plainsboro, NJ) collagen sponge saturated with recombinant human bone morphogenetic protein-2. Stimulation of this distraction-induced envelope by rhBMP-2 resulted in abundant production of bicortical membranous bone in situ within 12 weeks. The neoramus was subsequently suspended from the cranial base, and a temporalis muscle transfer was used to provide motor control of the jaw. Synthesis of bone in this manner is termed DISO (distraction-assisted in situ osteogenesis). The biologic rationale and clinical implications of DISO are discussed.

  16. Collagen metabolism in mouse lung after X irradiation

    SciTech Connect

    Murray, J.C.; Parkins, C.S.

    1987-09-01

    Collagen and total protein synthesis rates have been determined in the lungs of CBA mice irradiated with single doses of X rays between 8 and 16 Gy. Mice were injected with (/sup 3/H)proline accompanied by a large dose of unlabeled proline, and synthesis rates were measured at 2-month intervals from 8 to 31 weeks after irradiation. At 2 months after radiation treatment, collagen and total protein synthesis rates were significantly depressed but they had recovered by 4 months. By 6 months collagen synthesis rates had increased above control in a dose-dependent manner, so that in the 14-Gy dose group the fractional synthesis rate for collagen was 4.6 times higher than in control mice as measured by incorporation of (/sup 3/H)proline. However, a significant net accumulation of collagen was seen only in the lungs of the highest dose group at 31 weeks, as indicated by total hydroxyproline measurements. There was a slight increase in the ratio of types I and III collagen. Late radiation damage in the CBA mouse lung is characterized by increased collagen metabolism, which may or may not lead to a net accumulation of collagen.

  17. Optimal cooperative control synthesis of active displays

    NASA Technical Reports Server (NTRS)

    Garg, S.; Schmidt, D. K.

    1986-01-01

    The utility of augmenting displays to aid the human operator in controlling high order complex systems is well known. Analytical evaluation of various display designs for a simple k/s sup 2 plant in a compensatory tracking task using an optimal Control Model (OCM) of human behavior is carried out. This analysis reveals that significant improvement in performance should be obtained by skillful integration of key information into the display dynamics. The cooperative control synthesis technique previously developed to design pilot-optimal control augmentation is extended to incorporate the simultaneous design of performance enhancing augmented displays. The application of the cooperative control synthesis technique to the design of augmented displays is discussed for the simple k/s sup 2 plant. This technique is intended to provide a systematic approach to design optimally augmented displays tailored for specific tasks.

  18. Optimal cooperative control synthesis of active displays

    NASA Technical Reports Server (NTRS)

    Garg, S.; Schmidt, D. K.

    1985-01-01

    The utility of augmenting displays to aid the human operator in controlling high order complex systems is well known. Analytical evaluations of various display designs for a simple k/s-squared plant in a compensatory tracking task using an Optimal Control Model (OCM) of human behavior is carried out. This analysis reveals that significant improvement in performance should be obtained by skillful integration of key information into the display dynamics. The cooperative control synthesis technique previously developed to design pilot-optimal control augmentation is extended to incorporate the simultaneous design of performance enhancing augmented displays. The application of the cooperative control synthesis technique to the design of augmented displays is discussed for the simple k/s-squared plant. This technique is intended to provide a systematic approach to design optimally augmented displays tailored for specific tasks.

  19. Activation of catalysts for synthesizing methanol from synthesis gas

    DOEpatents

    Blum, David B.; Gelbein, Abraham P.

    1985-01-01

    A method for activating a methanol synthesis catalyst is disclosed. In this method, the catalyst is slurried in an inert liquid and is activated by a reducing gas stream. The activation step occurs in-situ. That is, it is conducted in the same reactor as is the subsequent step of synthesizing methanol from a methanol gas stream catalyzed by the activated catalyst still dispersed in a slurry.

  20. Vegetable peptones increase production of type I collagen in human fibroblasts by inducing the RSK-CCAAT/enhancer binding protein-β phosphorylation pathway.

    PubMed

    Jung, Eunsun; Cho, Jae Youl; Park, Deokhoon; Kim, Min Hee; Park, Beomseok; Lee, Sang Yeol; Lee, Jongsung

    2015-02-01

    Skin aging appears to be principally attributed to a decrease in type I collagen level and the regeneration ability of dermal fibroblasts. We hypothesized that vegetable peptones promote cell proliferation and production of type I collagen in human dermal fibroblasts. Therefore, we investigated the effects of vegetable peptones on cell proliferation and type I collagen production and their possible mechanisms in human dermal fibroblasts. Vegetable peptones significantly promoted cell proliferation in a concentration-dependent manner. In addition, the human luciferase type I collagen α2 promoter and type I procollagen synthesis assays showed that the vegetable peptones induced type I procollagen production by activating the type I collagen α2 promoter. Moreover, the vegetable peptones activated p90 ribosomal s6 kinase, which was mediated by activating the Raf-p44/42 mitogen-activated protein kinase signaling pathway. Furthermore, the vegetable peptone-induced increase in cell proliferation and type I collagen production decreased upon treatment with the ERK inhibitor PD98059. Taken together, these findings suggest that increased proliferation of human dermal fibroblasts and enhanced production of type I collagen by vegetable peptones occur primarily by inducing the p90 ribosomal s6 kinase-CCAAT/enhancer binding protein β phosphorylation pathway, which is mediated by activating Raf-ERK signaling.

  1. The Synthesis and Antimicrobial Activity of Heterocyclic Derivatives of Totarol

    PubMed Central

    2012-01-01

    The synthesis and antimicrobial activity of heterocyclic analogues of the diterpenoid totarol are described. An advanced synthetic intermediate with a ketone on the A-ring is used to attach fused heterocycles, and a carbon-to-nitrogen atom replacement is made on the B-ring by de novo synthesis. A-ring analogues with an indole attached exhibit, for the first time, enhanced antimicrobial activity relative to the parent natural product. Preliminary experiments demonstrate that the indole analogues do not target the bacterial cell division protein FtsZ as had been hypothesized for totarol. PMID:23119123

  2. Synthesis and antitumour activity of 4-aminoquinazoline derivatives

    NASA Astrophysics Data System (ADS)

    Lipunova, G. N.; Nosova, E. V.; Charushin, V. N.; Chupakhin, O. N.

    2016-07-01

    Pieces of data on the synthesis and antitumour activity of 4-aminoquinazolines are summarized and analyzed. Key methods for the synthesis of these compounds are considered, primarily cyclocondensation of carboxylic acid derivatives, as well as the oxidation of quinazolines and the cyclization of disubstituted thioureas. Improvements of synthetic schemes for erlotinib, gefitinib and lapatinib, which are the best-known pharmaceuticals based on compounds of the title class, are also considered. Synthetic strategies and biological activities for new 4-aminoquinazoline derivatives that are EGFR-tyrosine kinase inhibitors, multiactive compounds, and labelled compounds for use as positron emission tomography (PET) imaging agents are discussed. The bibliography includes 263 references.

  3. Curcuminoids Extract, Hydrolyzed Collagen and Green Tea Extract Synergically Inhibit Inflammatory and Catabolic Mediator’s Synthesis by Normal Bovine and Osteoarthritic Human Chondrocytes in Monolayer

    PubMed Central

    Comblain, Fanny; Sanchez, Christelle; Lesponne, Isabelle; Balligand, Marc; Serisier, Samuel; Henrotin, Yves

    2015-01-01

    The main objective of this study was to assess the in vitro effects of curcuminoids extract, hydrolyzed collagen and green tea extract in normal bovine chondrocytes and osteoarthritic human chondrocytes cultured in monolayer. This study also investigated the synergic or additive effects of these compounds. Enzymatically isolated primary bovine or human chondrocytes were cultured in monolayer until confluence and then incubated for 24 hours or 48 hours in the absence or in the presence of interleukin-1β and with or without curcuminoids extract, hydrolyzed collagen or green tea extract, added alone or in combination, at different concentrations. Cell viability was neither affected by these compounds, nor by interleukin 1β. In the absence of interleukin-1β, compounds did not significantly affect bovine chondrocytes metabolism. In human chondrocytes and in the absence of interleukin 1β, curcuminoids extract alone or in combination with hydrolyzed collagen and green tea extract significantly inhibited matrix metalloproteinase-3 production. In interleukin-1β-stimulated bovine chondrocytes, interleukin-6, inducible nitric oxide synthase, cyclooxygenase2, matrix metalloproteinase 3, a disintegrin and metalloproteinase with thrombospondin type I motifs 4 and a disintegrin and metalloproteinase with thrombospondin type I motifs 5 expressions were decreased by curcuminoids extract alone or in combination with hydrolyzed collagen and green tea extract. The combination of the three compounds was significantly more efficient to inhibit interleukin-1β stimulated matrix metalloproteinase-3 expression than curcuminoids extract alone. In interleukin-1β-stimulated human chondrocytes, nitric oxide, interleukin-6 and matrix metalloproteinase 3 productions were significantly reduced by curcuminoids extract alone or in combination with hydrolyzed collagen and green tea extract. These findings indicate that a mixture of curcuminoids extract, hydrolyzed collagen and green tea

  4. Curcuminoids extract, hydrolyzed collagen and green tea extract synergically inhibit inflammatory and catabolic mediator's synthesis by normal bovine and osteoarthritic human chondrocytes in monolayer.

    PubMed

    Comblain, Fanny; Sanchez, Christelle; Lesponne, Isabelle; Balligand, Marc; Serisier, Samuel; Henrotin, Yves

    2015-01-01

    The main objective of this study was to assess the in vitro effects of curcuminoids extract, hydrolyzed collagen and green tea extract in normal bovine chondrocytes and osteoarthritic human chondrocytes cultured in monolayer. This study also investigated the synergic or additive effects of these compounds. Enzymatically isolated primary bovine or human chondrocytes were cultured in monolayer until confluence and then incubated for 24 hours or 48 hours in the absence or in the presence of interleukin-1β and with or without curcuminoids extract, hydrolyzed collagen or green tea extract, added alone or in combination, at different concentrations. Cell viability was neither affected by these compounds, nor by interleukin 1β. In the absence of interleukin-1β, compounds did not significantly affect bovine chondrocytes metabolism. In human chondrocytes and in the absence of interleukin 1β, curcuminoids extract alone or in combination with hydrolyzed collagen and green tea extract significantly inhibited matrix metalloproteinase-3 production. In interleukin-1β-stimulated bovine chondrocytes, interleukin-6, inducible nitric oxide synthase, cyclooxygenase2, matrix metalloproteinase 3, a disintegrin and metalloproteinase with thrombospondin type I motifs 4 and a disintegrin and metalloproteinase with thrombospondin type I motifs 5 expressions were decreased by curcuminoids extract alone or in combination with hydrolyzed collagen and green tea extract. The combination of the three compounds was significantly more efficient to inhibit interleukin-1β stimulated matrix metalloproteinase-3 expression than curcuminoids extract alone. In interleukin-1β-stimulated human chondrocytes, nitric oxide, interleukin-6 and matrix metalloproteinase 3 productions were significantly reduced by curcuminoids extract alone or in combination with hydrolyzed collagen and green tea extract. These findings indicate that a mixture of curcuminoids extract, hydrolyzed collagen and green tea

  5. Chitosan nanoparticles enhance the antibacterial activity of chlorhexidine in collagen membranes used for periapical guided tissue regeneration.

    PubMed

    Barreras, Uriel Soto; Méndez, Fernando Torres; Martínez, Rita Elizabeth Martínez; Valencia, Carolina Samano; Rodríguez, Panfilo Raymundo Martinez; Rodríguez, Juan Pablo Loyola

    2016-01-01

    Endodontic failure is mainly associated with the persistence of microbial infection in the root canal system and/or the periradicular area. Microorganisms and their toxins located in the root canal system may trigger apical periodontitis and tissue destruction. Tissue regeneration in periapical surgery by using membrane barriers has shown an improved healing and bone closure. However, bacterial membrane contamination is a main reason of failure. In this in vitro study, different brands of chlorhexidine, a combination of chitosan nanoparticles containing chlorhexidine were tested against Enterococcus faecalis on agar plate's cultures and infected collagen membranes. Our results indicated that chitosan nanoparticles acted synergistically with chlorhexidine, inhibiting and eliminating significantly a greater amount of colony former units in both BHI-agar cultures and infected collagen membranes. These results suggested that chitosan nanoparticles could be used to improve regenerative procedures in periapical surgery. PMID:26478419

  6. SdrX, a serine-aspartate repeat protein expressed by Staphylococcus capitis with collagen VI binding activity.

    PubMed

    Liu, Yule; Ames, Brenda; Gorovits, Elena; Prater, Bradley D; Syribeys, Peter; Vernachio, John H; Patti, Joseph M

    2004-11-01

    Staphylococcus capitis (S. capitis) has been implicated in a large proportion of coagulase-negative staphylococcal infections in very-low-birth-weight infants. To identify potential therapeutic targets, the S. capitis genome was probed for the presence of genes encoding microbial surface components recognizing adhesive matrix molecules (MSCRAMM). By using Southern blot analysis, an S. capitis gene, designated sdrX, that contained sequence motifs consistent with the Sdr family of MSCRAMM proteins was identified. By using monospecific antisera in Western blot and flow cytometry, SdrX was demonstrated to be expressed on the surface of S. capitis. Human collagen type VI was found to bind both the recombinant A domain of SdrX and viable S. capitis expressing SdrX. SdrX is the first collagen-binding Sdr protein described and is the first MSCRAMM protein identified in S. capitis. PMID:15501749

  7. Chitosan nanoparticles enhance the antibacterial activity of chlorhexidine in collagen membranes used for periapical guided tissue regeneration.

    PubMed

    Barreras, Uriel Soto; Méndez, Fernando Torres; Martínez, Rita Elizabeth Martínez; Valencia, Carolina Samano; Rodríguez, Panfilo Raymundo Martinez; Rodríguez, Juan Pablo Loyola

    2016-01-01

    Endodontic failure is mainly associated with the persistence of microbial infection in the root canal system and/or the periradicular area. Microorganisms and their toxins located in the root canal system may trigger apical periodontitis and tissue destruction. Tissue regeneration in periapical surgery by using membrane barriers has shown an improved healing and bone closure. However, bacterial membrane contamination is a main reason of failure. In this in vitro study, different brands of chlorhexidine, a combination of chitosan nanoparticles containing chlorhexidine were tested against Enterococcus faecalis on agar plate's cultures and infected collagen membranes. Our results indicated that chitosan nanoparticles acted synergistically with chlorhexidine, inhibiting and eliminating significantly a greater amount of colony former units in both BHI-agar cultures and infected collagen membranes. These results suggested that chitosan nanoparticles could be used to improve regenerative procedures in periapical surgery.

  8. Adopting the principles of collagen biomineralization for intrafibrillar infiltration of yttria-stabilized zirconia into three-dimensional collagen scaffolds

    PubMed Central

    Zhou, Bin; Niu, Li-na; Shi, Wei; Zhang, Wei; Arola, Dwayne D.; Breschi, Lorenzo; Mao, Jing; Pashley, David H.

    2014-01-01

    In this paper, we report a process for generating collagen-yttria-stabilized amorphous zirconia hybrid scaffolds by introducing acetylacetone-inhibited zirconia precursor nanodroplets into a poly(allylamine)-coated collagen matrix. This polyelectrolyte coating triggers intrafibrillar condensation of the precursors into amorphous zirconia, which is subsequently transformed into tetragonal yttria-stabilized zirconia after calcination. Our findings represent a new paradigm in the synthesis of non-naturally occurring collagen-based hybrid scaffolds under alcoholic mineralizing conditions. PMID:25477773

  9. Phytochrome activation of two nuclear genes requires cytoplasmic protein synthesis.

    PubMed Central

    Lam, E; Green, P J; Wong, M; Chua, N H

    1989-01-01

    We have investigated the effects of protein synthesis inhibitors on light-induced expression of two plant nuclear genes, Cab and rbcS, in wheat, pea and transgenic tobacco. Light activation of these two genes is very sensitive to cycloheximide, an inhibitor of cytoplasmic protein synthesis but not to chloramphenicol, an inhibitor of organellar protein synthesis. Studies with chimeric gene constructs in transgenic tobacco seedlings show that cycloheximide exerts its effect at the transcriptional level. As a control, we show that the expression of the cauliflower mosaic virus (CaMV) 35S promoter is enhanced by cycloheximide treatment, irrespective of the coding sequence used. Escape-time analyses with green wheat seedlings show that the cycloheximide block for Cab gene expression is after the primary signal transduction step linked to phytochrome photoconversion. Our results suggest that phytochrome activation of Cab and rbcS is mediated by a labile protein factor(s) synthesized on cytoplasmic ribosomes. Images PMID:2583082

  10. Collagen vascular disease

    MedlinePlus

    ... this page: //medlineplus.gov/ency/article/001223.htm Collagen vascular disease To use the sharing features on ... were previously said to have "connective tissue" or "collagen vascular" disease. We now have names for many ...

  11. Platelet-activating factor: mediator of the third pathway of platelet aggregation? A study in three patients with deficient platelet-activating factor synthesis.

    PubMed Central

    Sturk, A; Schaap, M C; ten Cate, J W; Heymans, H S; Schutgens, R B; Przyrembel, H; Borst, P

    1987-01-01

    Thrombin, collagen, and Ca2+-ionophore A23187 aggregate platelets in the presence of inhibitors of the first (ADP-mediated) and second (cyclooxygenase-dependent) pathway of platelet activation. This aggregation, via a third pathway, was hypothesized to be mediated by the alkoxyether lipid platelet-activating factor (PAF). We recently demonstrated virtual absence of plasmalogen-type alkoxyether lipids and deficiency in key enzymes of their biosynthesis in Zellweger patients. We hypothesized that PAF synthesis might also be impaired. We report two Zellweger patients with an undetectable A23187-induced PAF synthesis of leukocytes (patients, less than 3 pmol PAF/10(8) granulocytes (PMN); four age-matched controls, 249-2,757 pmol PAF/10(8) PMN; five adult controls, 291-5,433 pmol PAF/10(8) PMN). In a third patient, residual PAF synthesis was detected. However in all patients the thrombin-induced third mechanism of platelet aggregation was present. We therefore conclude that PAF may not be the mediator of the third pathway. PMID:3805272

  12. Optimal cooperative control synthesis of active displays

    NASA Technical Reports Server (NTRS)

    Gary, Sanjay; Schmidt, David K.

    1987-01-01

    A technique is developed that is intended to provide a systematic approach to synthesizing display augmentation for optimal manual control in complex, closed-loop tasks. A cooperative control synthesis technique, previously developed to design pilot-optimal control augmentation for the plant, is extended to incorporate the simultaneous design of performance enhancing displays. The technique utilizes an optimal control model of the man in the loop. It is applied to the design of a quickening control law for a display and a simple K/(s squared) plant, and then to an F-15 type aircraft in a multichannel task. Utilizing the closed-loop modeling and analysis procedures, the results from the display design algorithm are evaluated and an analytical validation is performed. Experimental validation is recommended for future efforts.

  13. Optimal cooperative control synthesis of active displays

    NASA Technical Reports Server (NTRS)

    Garg, S.; Schmidt, D. K.

    1985-01-01

    A technique is developed that is intended to provide a systematic approach to synthesizing display augmentation for optimal manual control in complex, closed-loop tasks. A cooperative control synthesis technique, previously developed to design pilot-optimal control augmentation for the plant, is extended to incorporate the simultaneous design of performance enhancing displays. The technique utilizes an optimal control model of the man in the loop. It is applied to the design of a quickening control law for a display and a simple K/s(2) plant, and then to an F-15 type aircraft in a multi-channel task. Utilizing the closed loop modeling and analysis procedures, the results from the display design algorithm are evaluated and an analytical validation is performed. Experimental validation is recommended for future efforts.

  14. The simultaneous release by bone explants in culture and the parallel activation of procollagenase and of a latent neutral proteinase that degrades cartilage proteoglycans and denatured collagen.

    PubMed Central

    Vaes, G; Eeckhout, Y; Lenaers-Claeys, G; François-Gillet, C; Druetz, J E

    1978-01-01

    1. A latent neutral proteinase was found in culture media of mouse bone explants. Its accumulation during the cultures is closely parallel to that of procollagenase; both require the presence of heparin in the media. 2. Latent neutral proteinase was activated by several treatments of the media known to activate procollagenase, such as limited proteolysis by trypsin, chymotrypsin, plasmin or kallikrein, dialysis against 3 M-NaSCN at 4 degrees C and prolonged preincubation at 25 degrees C. Its activation often followed that of the procollagenase present in the same media. 3. Activation of neutral proteinase (as does that of procollagenase) by trypsin or plasmin involved two successive steps: the activation of a latent endogenous activator present in the media followed by the activation of neutral proteinase itself by that activator. 4. The proteinase degrades cartilage proteoglycans, denatured collagen (Azocoll) and casein at neutral pH; it is inhibited by EDTA, cysteine or serum. Collagenase is not inhibited by casein or Azocoll and is less resistant to heat or to trypsin than is the proteinase. Partial separation of the two enzymes was achieved by gel filtration of the media but not by fractional (NH4)2SO4 precipitation, by ion exchange or by affinity chromatography on Sepharose-collagen. These fractionations did not activate latent enzymes. 5. Trypsin activation decreases the molecular weight of both latent enzymes (60 000-70 000) by 20 000-30 000, as determined by gel filtration of media after removal of heparin. 6. The latency of both enzymes could be due either to a zymogen or to an enzyme-inhibitor complex. A thermostable inhibitor of both enzymes was found in some media. However, combinations of either enzyme with that inhibitor were not reactivated by trypsin, indicating that this inhibitor is unlikely to be the cause of the latency. PMID:208518

  15. A nanostructured synthetic collagen mimic for hemostasis.

    PubMed

    Kumar, Vivek A; Taylor, Nichole L; Jalan, Abhishek A; Hwang, Lyahn K; Wang, Benjamin K; Hartgerink, Jeffery D

    2014-04-14

    Collagen is a major component of the extracellular matrix and plays a wide variety of important roles in blood clotting, healing, and tissue remodeling. Natural, animal derived, collagen is used in many clinical applications but concerns exist with respect to its role in inflammation, batch-to-batch variability, and possible disease transfection. Therefore, development of synthetic nanomaterials that can mimic the nanostructure and properties of natural collagen has been a heavily pursued goal in biomaterials. Previously, we reported on the design and multihierarchial self-assembly of a 36 amino acid collagen mimetic peptide (KOD) that forms nanofibrous triple helices that entangle to form a hydrogel. In this report, we utilize this nanofiber forming collagen mimetic peptide as a synthetic biomimetic matrix useful in thrombosis. We demonstrate that nanofibrous KOD synthetic collagen matrices adhere platelets, activate them (indicated by soluble P-selectin secretion), and clot plasma and blood similar to animal derived collagen and control surfaces. In addition to the thrombotic potential, THP-1 monocytes incubated with our KOD collagen mimetic showed minimal proinflammatory cytokine (TNF-α or IL-1β) production. Together, the data presented demonstrates the potential of a novel synthetic collagen mimetic as a hemostat.

  16. A nanostructured synthetic collagen mimic for hemostasis.

    PubMed

    Kumar, Vivek A; Taylor, Nichole L; Jalan, Abhishek A; Hwang, Lyahn K; Wang, Benjamin K; Hartgerink, Jeffery D

    2014-04-14

    Collagen is a major component of the extracellular matrix and plays a wide variety of important roles in blood clotting, healing, and tissue remodeling. Natural, animal derived, collagen is used in many clinical applications but concerns exist with respect to its role in inflammation, batch-to-batch variability, and possible disease transfection. Therefore, development of synthetic nanomaterials that can mimic the nanostructure and properties of natural collagen has been a heavily pursued goal in biomaterials. Previously, we reported on the design and multihierarchial self-assembly of a 36 amino acid collagen mimetic peptide (KOD) that forms nanofibrous triple helices that entangle to form a hydrogel. In this report, we utilize this nanofiber forming collagen mimetic peptide as a synthetic biomimetic matrix useful in thrombosis. We demonstrate that nanofibrous KOD synthetic collagen matrices adhere platelets, activate them (indicated by soluble P-selectin secretion), and clot plasma and blood similar to animal derived collagen and control surfaces. In addition to the thrombotic potential, THP-1 monocytes incubated with our KOD collagen mimetic showed minimal proinflammatory cytokine (TNF-α or IL-1β) production. Together, the data presented demonstrates the potential of a novel synthetic collagen mimetic as a hemostat. PMID:24694012

  17. (+)- and (-)-mutisianthol: first total synthesis, absolute configuration, and antitumor activity.

    PubMed

    Bianco, Graziela G; Ferraz, Helena M C; Costa, Arinice M; Costa-Lotufo, Letícia V; Pessoa, Cláudia; de Moraes, Manoel O; Schrems, Marcus G; Pfaltz, Andreas; Silva, Luiz F

    2009-03-20

    The first synthesis of the natural product (+)-mutisianthol was accomplished in 11 steps and in 21% overall yield from 2-methylanisole. The synthesis of its enantiomer was also performed in a similar overall yield. The absolute configuration of the sesquiterpene (+)-mutisianthol was assigned as (1S,3R). Key steps in the route are the asymmetric hydrogenation of a nonfunctionalized olefin using chiral iridium catalysts and the ring contraction of 1,2-dihydronaphthalenes using thallium(III) or iodine(III). The target molecules show moderate activity against the human tumor cell lines SF-295, HCT-8, and MDA-MB-435.

  18. Intracellular collagen fibrils: evidence of an intracellular source from experiments with tendon fibroblasts and fibroblastic tumour cells.

    PubMed Central

    Michna, H

    1988-01-01

    This study was designed to substantiate one or both of the two hypotheses for the explanation of intracellular collagen fibrils in collagen-producing cells. The more obvious is the phagocytosis of extracellular collagen fibrils by the cell and the other is a form of autophagocytosis of newly synthesised collagenous products. Information was collected on fibroblasts from murine tendons after exercise and simultaneously stimulating collagen synthesis by treatment with an anabolic steroid hormone. Moreover, in vivo and in vitro fibroblastic tumour cells which demonstrate enhanced protein synthesis were also treated with the anabolic steroid. The findings of intracellular collagen fibrils in tendon fibroblasts and the sarcoma cells after experimentally stimulating collagen synthesis are discussed in the light of the hypothesis that the findings may represent steps of autophagocytosis of newly synthesised collagenous products in the absence of a control mechanism to remove collagenous products which cannot be secreted. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:3225213

  19. Synthesis and antiplasmodial activity of lycorine derivatives.

    PubMed

    Cedrón, Juan C; Gutiérrez, David; Flores, Ninoska; Ravelo, Angel G; Estévez-Braun, Ana

    2010-07-01

    Twenty seven lycorine derivatives were prepared and evaluated for their in vitro antimalarial activity against chloroquine-sensitive strains of Plasmodium falciparum. The best antiplasmodial activities were achieved with lycorine derivatives that present free hydroxyl groups at C-1 and C-2 or esterified as acetates or isobutyrates. The double bond C-2-C-3 is also important for the activity. Concerning to the antiplasmodial activity of the secolycorines, the higher values were obtained with the replacement of the methylenedioxy moiety by hydroxyl or acetate groups and with methyl substituent attached to the nitrogen atom.

  20. Total synthesis and biological activity of natural product Urukthapelstatin A.

    PubMed

    Lin, Chun-Chieh; Tantisantisom, Worawan; McAlpine, Shelli R

    2013-07-19

    Herein we report the first total synthesis of the natural product Urkuthaplestatin A (Ustat A) utilizing a convergent synthetic strategy. The characterization and biological activity match those of the previously published natural product. Interestingly, several intermediates, including the linear and serine cyclized precursors, show a 100-fold decrease in cytotoxicity, with IC50's in the low micromolar range. These data indicate that the rigidity and the consecutive aromatic heterocyclic system are responsible for the biological activity. PMID:23819711

  1. Synthesis and anti-inflammatory activity of aromatic glucosinolates.

    PubMed

    Vo, Quan V; Trenerry, Craige; Rochfort, Simone; Wadeson, Jenny; Leyton, Carolina; Hughes, Andrew B

    2013-10-01

    Aromatic GLs are important members of the glucosinolate family of compounds because of their potential biological activity and medicinal properties. This study has shown success in the high yielding synthesis of some important aromatic GLs as well as the results of testing for anti-inflammatory properties of the synthetic GLs. 3,4-Dimethoxyphenylglucosinolate was found to be the most active anti-inflammatory of the seven glucosinolates assayed. PMID:23978357

  2. Complications of collagenous colitis.

    PubMed

    Freeman, Hugh-James

    2008-03-21

    Microscopic forms of colitis have been described, including collagenous colitis. This disorder generally has an apparently benign clinical course. However, a number of gastric and intestinal complications, possibly coincidental, may develop with collagenous colitis. Distinctive inflammatory disorders of the gastric mucosa have been described, including lymphocytic gastritis and collagenous gastritis. Celiac disease and collagenous sprue (or collagenous enteritis) may occur. Colonic ulceration has been associated with use of nonsteroidal anti-inflammatory drugs, while other forms of inflammatory bowel disease, including ulcerative colitis and Crohn's disease, may evolve from collagenous colitis. Submucosal "dissection", colonic fractures or mucosal tears and perforation from air insufflation during colonoscopy may occur and has been hypothesized to be due to compromise of the colonic wall from submucosal collagen deposition. Similar changes may result from increased intraluminal pressure during barium enema contrast studies. Finally, malignant disorders have also been reported, including carcinoma and lymphoproliferative disease. PMID:18350593

  3. Design, Synthesis and Evaluation of Antiproliferative Activity of New Benzimidazolehydrazones.

    PubMed

    Onnis, Valentina; Demurtas, Monica; Deplano, Alessandro; Balboni, Gianfranco; Baldisserotto, Anna; Manfredini, Stefano; Pacifico, Salvatore; Liekens, Sandra; Balzarini, Jan

    2016-01-01

    The synthesis and antiproliferative activity of new benzimidazole derivatives bearing an hydrazone mojety at the 2-position is described. The new N'-(4-arylidene)-1H-benzo[d]imidazole-2-carbohydrazides were evaluated for their cytostatic activity toward the murine leukemia (L1210), human T-cell leukemia (CEM), human cervix carcinoma (HeLa) and human pancreas carcinoma cells (Mia Paca-2). A preliminary structure-activity relationship could be defined. Some of the compounds possess encouraging and consistent antiproliferative activity, having IC50 values in the low micromolar range. PMID:27144551

  4. Cyclobutane-Containing Alkaloids: Origin, Synthesis, and Biological Activities

    PubMed Central

    Sergeiko, Anastasia; Poroikov, Vladimir V; Hanuš, Lumir O; Dembitsky, Valery M

    2008-01-01

    Present review describes research on novel natural cyclobutane-containing alkaloids isolated from terrestrial and marine species. More than 60 biological active compounds have been confirmed to have antimicrobial, antibacterial, antitumor, and other activities. The structures, synthesis, origins, and biological activities of a selection of cyclobutane-containing alkaloids are reviewed. With the computer program PASS some additional biological activities are also predicted, which point toward new possible applications of these compounds. This review emphasizes the role of cyclobutane-containing alkaloids as an important source of leads for drug discovery. PMID:19696873

  5. Efficient delivery to human lung fibroblasts (WI-38) of pirfenidone incorporated into liposomes modified with truncated basic fibroblast growth factor and its inhibitory effect on collagen synthesis in idiopathic pulmonary fibrosis.

    PubMed

    Togami, Kohei; Miyao, Aki; Miyakoshi, Kei; Kanehira, Yukimune; Tada, Hitoshi; Chono, Sumio

    2015-01-01

    In the present in vitro study, we assessed the delivery of pirfenidone incorporated into liposomes modified with truncated basic fibroblast growth factor (tbFGF) to lung fibroblasts and investigated the anti-fibrotic effect of the drug. The tbFGF peptide, KRTGQYKLC, was used to modify the surface of liposomes (tbFGF-liposomes). We used the thin-layer evaporation method, followed by sonication, to prepare tbFGF-liposomes containing pirfenidone. The cellular accumulation of tbFGF-liposomes was 1.7-fold greater than that of non-modified liposomes in WI-38 cells used as a model of lung fibroblasts. Confocal laser scanning microscopy showed that tbFGF-liposomes were widely localized in WI-38 cells. The inhibitory effects of pirfenidone incorporated into tbFGF-liposomes on transforming growth factor-β1 (TGF-β1)-induced collagen synthesis in WI-38 cells were evaluated by measuring the level of intracellular hydroxyproline, a major component of the protein collagen. Pirfenidone incorporated into tbFGF-liposomes at concentrations of 10, 30, and 100 µM significantly decreased the TGF-β1-induced hydroxyproline content in WI-38 cells. The anti-fibrotic effect of pirfenidone incorporated into tbFGF-liposomes was enhanced compared with that of pirfenidone solution. These results indicate that tbFGF-liposomes are a useful drug delivery system of anti-fibrotic drugs to lung fibroblasts for the treatment of idiopathic pulmonary fibrosis.

  6. Xenicane Natural Products: Biological Activity and Total Synthesis.

    PubMed

    Betschart, Leo; Altmann, Karl-Heinz

    2015-01-01

    The xenicanes are a large class of mostly bicyclic marine diterpenoids featuring a cyclononane ring as a common structural denominator. After a brief introduction into the characteristic structural features of xenicanes and some biogenetic considerations, the major focus of this review will be on the various biological activities that have been reported for xenicanes and on efforts towards the total synthesis of these structures. Several xenicanes have been shown to be potent antiproliferative agents in vitro, but activities have also been reported in relation to inflammatory processes. However, so far, data on the possible in vivo activity of xenicanes are lacking. The major challenge in the total synthesis of xenicanes is the construction of the nine-membered ring. Different strategies have been pursued to establish this crucial substructure, including Grob fragmentation, ring-closing olefin metathesis, or Suzuki cross coupling as the enabling transformations. PMID:26429717

  7. Synthesis and antimalarial activity of prodigiosenes.

    PubMed

    Marchal, Estelle; Smithen, Deborah A; Uddin, Md Imam; Robertson, Andrew W; Jakeman, David L; Mollard, Vanessa; Goodman, Christopher D; MacDougall, Kristopher S; McFarland, Sherri A; McFadden, Geoffrey I; Thompson, Alison

    2014-06-28

    Several analogues of the natural compound prodigiosin with modified A- and C-rings were synthesised as were some of their tin, cobalt, boron and zinc complexes. The antimalarial activity of these prodigiosenes was evaluated in vitro using the 3D7 Plasmodium falciparum strain. The presence of a nitrogen atom in the A-ring is needed for antimalarial activity but the presence of an alkyl group at the β'-position of the C-ring seems detrimental. Dibutyl tin complexes exhibit IC50 values mostly in the nanomolar range with equal or improved activity compared to the free-base prodigiosene ligand, despite the fact that the general toxicity of such tin complexes is demonstrably lower than that of the free-bases.

  8. Synthesis and antiproliferative activity of 6-phenylaminopurines.

    PubMed

    Canela, María-Dolores; Liekens, Sandra; Camarasa, María-José; Priego, Eva María; Pérez-Pérez, María-Jesús

    2014-11-24

    A series of novel 6-phenylaminopurines have been efficiently synthesized in 3 steps exploring different groups at positions 2, 8 and 9 of the purine ring and at the exocyclic nitrogen atom at position 6. Among the newly described purines, five compounds showed antiproliferative activity with IC50 values below 10 μM, the tetrahydroquinoline derivative at position 6 of phenylaminopurine being the most active of the series in the six cell lines tested. Moreover, the compounds induced G2/M phase arrest in human cervical carcinoma HeLa cells as reported for tubulin depolymerizing agents.

  9. Biomimetic synthesis of active isorhapontigenin dimers.

    PubMed

    Wang, Xian-Fen; Zhang, Yuan; Lin, Ming-Bao; Hou, Qi; Yao, Chun-Suo; Shi, Jian-Gong

    2014-01-01

    Synthetic isorhapontigenin was treated with several kinds of inorganic reagents and peroxidase so as to prepare active stilbene dimers. Among them, silver acetate in methanol gave two new isorhapontigenin dimers 4 and 5, together with four known natural stilbene dimers 2, 3, 6, and 7. Their structures and relative configurations were determined on the basis of spectral analysis, and their possible formation mechanisms were discussed, respectively. Compounds 2, 6, and 7 were artificially synthesized for the first time. All the products were evaluated for anti-inflammatory activities.

  10. Impaired intestinal wound healing in Fhl2-deficient mice is due to disturbed collagen metabolism

    SciTech Connect

    Kirfel, Jutta Pantelis, Dimitrios; Kabba, Mustapha; Kahl, Philip; Roeper, Anke; Kalff, Joerg C.; Buettner, Reinhard

    2008-12-10

    Four and one half LIM domain protein FHL2 participates in many cellular processes involved in tissue repair such as regulation of gene expression, cytoarchitecture, cell adhesion, migration and signal transduction. The repair process after wounding is initiated by the release of peptides and bioactive lipids. These molecules induce synthesis and deposition of a provisional extracellular matrix. We showed previously that sphingosine-1-phosphate (S1P) triggers a signal transduction cascade mediating nuclear translocation of FHL2 in response to activation of the RhoA GTPase. Our present study shows that FHL2 is an important signal transducer influencing the outcome of intestinal anastomotic healing. Early wound healing is accompanied by reconstitution and remodelling of the extracellular matrix and collagen is primarily responsible for wound strength. Our results show that impaired intestinal wound healing in Fhl2-deficient mice is due to disturbed collagen III metabolism. Impaired collagen III synthesis reduced the mechanical stability of the anastomoses and led to lower bursting pressure in Fhl2-deficient mice after surgery. Our data confirm that FHL2 is an important factor regulating collagen expression in the early phase of wound healing, and thereby is critically involved in the physiologic process of anastomosis healing after bowel surgery and thus may represent a new therapeutic target.

  11. Curcumin improves wound healing by modulating collagen and decreasing reactive oxygen species.

    PubMed

    Panchatcharam, Manikandan; Miriyala, Sumitra; Gayathri, Vinaya Subramani; Suguna, Lonchin

    2006-10-01

    Wound healing consists of an orderly progression of events that re-establish the integrity of the damaged tissue. Several natural products have been shown to accelerate the healing process. The present investigation was undertaken to determine the role of curcumin on changes in collagen characteristics and antioxidant property during cutaneous wound healing in rats. Full-thickness excision wounds were made on the back of rat and curcumin was administered topically. The wound tissues removed on 4th, 8th and 12th day (post-wound) were used to analyse biochemical and pathological changes. Curcumin increased cellular proliferation and collagen synthesis at the wound site, as evidenced by increase in DNA, total protein and type III collagen content of wound tissues. Curcumin treated wounds were found to heal much faster as indicated by improved rates of epithelialisation, wound contraction and increased tensile strength which were also confirmed by histopathological examinations. Curcumin treatment was shown to decrease the levels of lipid peroxides (LPs), while the levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), activities were significantly increased exhibiting the antioxidant properties of curcumin in accelerating wound healing. Better maturation and cross linking of collagen were observed in the curcumin treated rats, by increased stability of acid-soluble collagen, aldehyde content, shrinkage temperature and tensile strength. The results clearly substantiate the beneficial effects of the topical application of curcumin in the acceleration of wound healing and its antioxidant effect. PMID:16770527

  12. ETP-46321, a dual p110α/δ class IA phosphoinositide 3-kinase inhibitor modulates T lymphocyte activation and collagen-induced arthritis.

    PubMed

    Aragoneses-Fenoll, L; Montes-Casado, M; Ojeda, G; Acosta, Y Y; Herranz, J; Martínez, S; Blanco-Aparicio, C; Criado, G; Pastor, J; Dianzani, U; Portolés, P; Rojo, J M

    2016-04-15

    Class IA phosphoinositide 3-kinases (PI3Ks) are essential to function of normal and tumor cells, and to modulate immune responses. T lymphocytes express high levels of p110α and p110δ class IA PI3K. Whereas the functioning of PI3K p110δ in immune and autoimmune reactions is well established, the role of p110α is less well understood. Here, a novel dual p110α/δ inhibitor (ETP-46321) and highly specific p110α (A66) or p110δ (IC87114) inhibitors have been compared concerning T cell activation in vitro, as well as the effect on responses to protein antigen and collagen-induced arthritis in vivo. In vitro activation of naive CD4(+) T lymphocytes by anti-CD3 and anti-CD28 was inhibited more effectively by the p110δ inhibitor than by the p110α inhibitor as measured by cytokine secretion (IL-2, IL-10, and IFN-γ), T-bet expression and NFAT activation. In activated CD4(+) T cells re-stimulated through CD3 and ICOS, IC87114 inhibited Akt and Erk activation, and the secretion of IL-2, IL-4, IL-17A, and IFN-γ better than A66. The p110α/δ inhibitor ETP-46321, or p110α plus p110δ inhibitors also inhibited IL-21 secretion by differentiated CD4(+) T follicular (Tfh) or IL-17-producing (Th17) helper cells. In vivo, therapeutic administration of ETP-46321 significantly inhibited responses to protein antigen as well as collagen-induced arthritis, as measured by antigen-specific antibody responses, secretion of IL-10, IL-17A or IFN-γ, or clinical symptoms. Hence, p110α as well as p110δ Class IA PI3Ks are important to immune regulation; inhibition of both subunits may be an effective therapeutic approach in inflammatory autoimmune diseases like rheumatoid arthritis.

  13. Induction of an Epithelial Integrin αvβ6 in Human Cytomegalovirus-Infected Endothelial Cells Leads to Activation of Transforming Growth Factor-β1 and Increased Collagen Production

    PubMed Central

    Tabata, Takako; Kawakatsu, Hisaaki; Maidji, Ekaterina; Sakai, Takao; Sakai, Keiko; Fang-Hoover, June; Aiba, Motohiko; Sheppard, Dean; Pereira, Lenore

    2008-01-01

    Human cytomegalovirus (CMV) infection is a major cause of morbidity in immunosuppressed individuals, and congenital CMV infection is a leading cause of birth defects in newborns. Infection with pathogenic viral strains alters cell-cell and cell-matrix interactions, affecting extracellular matrix remodeling and endothelial cell migration. The multifunctional cytokine transforming growth factor (TGF)-β1 regulates cell proliferation, differentiation, and extracellular matrix remodeling. Secreted as a latent protein complex, TGF-β1 requires activation before binding to receptors that phosphorylate intracellular effectors. TGF-β1 is activated by integrin αvβ6, which is strongly induced in the epithelium by injury and inflammation but has not previously been found in endothelial cells. Here, we report that CMV infection induces integrin αvβ6 expression in endothelial cells, leading to activation of TGF-β1, signaling through its receptor ALK5, and phosphorylation of its intracellular effector Smad3. Infection of endothelial cells was also found to stimulate collagen synthesis through a mechanism dependent on both TGF-β1 and integrin αvβ6. Immunohistochemical analysis showed integrin αvβ6 up-regulation in capillaries proximal to foci of CMV infection in lungs, salivary glands, uterine decidua, and injured chorionic villi of the placenta, demonstrating both its induction in endothelium and up-regulation in epithelium in vivo. Our results suggest that activation of TGF-β1 by integrin αvβ6 contributes to pathological changes and may impair endothelial cell functions in tissues that are chronically infected with CMV. PMID:18349127

  14. Aqueous Extract of Chrysanthemum morifolium (菊花 Jú Huā) Enhances the Antimelanogenic and Antioxidative Activities of the Mixture of Soy Peptide and Collagen Peptide

    PubMed Central

    Gui, Min; Du, Jun; Guo, Jianmin; Xiao, Baiquan; Yang, Wei; Li, Minjie

    2014-01-01

    The possible synergistic effect between the aqueous extract of Chrysanthemum morifolium (菊花 Jú Huā) (AECM) and the peptide mixture (PM) containing soy peptide and collagen peptide was investigated in an ultraviolet (UV) irradiation–induced skin damage mouse model. The irradiated mice were treated with the PM or PM + AECM (containing PM and AECM), respectively. Both PM and PM + AECM groups displayed an apparent photoprotective effect on the UV-irradiated skin damage of mice. Histological evaluation demonstrated that the epidermal hyperplasia and melanocytes in the basal epidermal layer of the UV-irradiated skin in mice decreased when treated with either PM or PM + AECM. Further study showed that soy peptide, collagen peptide, and AECM also inhibited the activities of mushroom tyrosinase with IC50 values of 82.3, 28.2, and 1.6 μg/ml, respectively. Additionally, PM + AECM reduced melanogenesis by 46.2% at the concentration of 10 mg/ml in B16 mouse melanoma cells. Meanwhile, the UV-induced increase of antioxidative indicators, including glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and malondialdehyde (MDA), was reduced significantly after treatment with 1.83 g/kg/dbw of PM + AECM. This evidence supported the synergistic antioxidative effect of AECM with PM. These results demonstrated that oral intake of PM and AECM had synergistic antimelanogenic and antioxidative effects in UV-irradiated mice. PMID:25161922

  15. Aqueous Extract of Chrysanthemum morifolium ( Jú Huā) Enhances the Antimelanogenic and Antioxidative Activities of the Mixture of Soy Peptide and Collagen Peptide.

    PubMed

    Gui, Min; Du, Jun; Guo, Jianmin; Xiao, Baiquan; Yang, Wei; Li, Minjie

    2014-07-01

    The possible synergistic effect between the aqueous extract of Chrysanthemum morifolium ( Jú Huā) (AECM) and the peptide mixture (PM) containing soy peptide and collagen peptide was investigated in an ultraviolet (UV) irradiation-induced skin damage mouse model. The irradiated mice were treated with the PM or PM + AECM (containing PM and AECM), respectively. Both PM and PM + AECM groups displayed an apparent photoprotective effect on the UV-irradiated skin damage of mice. Histological evaluation demonstrated that the epidermal hyperplasia and melanocytes in the basal epidermal layer of the UV-irradiated skin in mice decreased when treated with either PM or PM + AECM. Further study showed that soy peptide, collagen peptide, and AECM also inhibited the activities of mushroom tyrosinase with IC50 values of 82.3, 28.2, and 1.6 μg/ml, respectively. Additionally, PM + AECM reduced melanogenesis by 46.2% at the concentration of 10 mg/ml in B16 mouse melanoma cells. Meanwhile, the UV-induced increase of antioxidative indicators, including glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and malondialdehyde (MDA), was reduced significantly after treatment with 1.83 g/kg/dbw of PM + AECM. This evidence supported the synergistic antioxidative effect of AECM with PM. These results demonstrated that oral intake of PM and AECM had synergistic antimelanogenic and antioxidative effects in UV-irradiated mice.

  16. Aminoderivatives of cycloalkanespirohydantoins: synthesis and biological activity.

    PubMed

    Naydenova, Emilia; Pencheva, Nevena; Popova, Julita; Stoyanov, Neyko; Lazarova, Maria; Aleksiev, Boris

    2002-03-01

    3-Aminocycloalkanespiro-5-hydantoins were synthesized and their biological activity was studied. In contrast to hydantoins, these compounds failed to induce either anticonvulsive effects in the central nervous system or inhibitory effects on cholinergic contractions in the enteric nervous system. However, they exerted well pronounced, atropinsensitive, contractile effects on the guinea-pig ileum longitudinal muscle preparations. Structure-activity relationships established allow the assumption that: (i) the reduction of the ring size in the molecule of the spirohydantoins leads to an increase in the potency of the respective analogue to induce contractile effect; (ii) the introduction of -NH2 in position 3 increases the ability of all the compounds studied to exert contractions; (iii) the enlargement of the ring leads to: (1) an increase of the degree of desensitization of the preparations; and (2) a decrease (except 1a) of the potency of the analogues to exert contractile effects.

  17. Synthesis and antiviral activity of 5'-deoxypyrazofurin.

    PubMed

    Chen, X; Schneller, S W; Ikeda, S; Snoeck, R; Andrei, G; Balzarini, J; De Clercq, E

    1993-11-12

    In searching for derivatives of pyrazofurin that could display antiviral properties by means that do not require C-5' phosphorylation, 5'-deoxypyrazofurin (3) has been synthesized in six steps from methyl5-deoxy-2,3-O-isopropylidene-beta-D-ribofuranoside (4). Compound 3 was evaluated for antiviral activity against a large number of viruses including herpes-, pox-, myxo-, toga-, arena-, rhabdo-, picorna-,reo-, and retroviruses. Compound 3 proved active against respiratory syncytial virus (in HeLa cells), vaccinia virus (in embryonic skin-muscle fibroblast cells), vesicular stomatitis virus (in HeLa cells), and influenza A virus (in Madin-Darby canine kidney cells) at concentrations (ranging from 4 to 20 micrograms/mL) that were nontoxic to the confluent host cell cultures. PMID:8246242

  18. Synthesis and trypanocidal activity of novel benzimidazole derivatives.

    PubMed

    Velázquez-López, José Miguel; Hernández-Campos, Alicia; Yépez-Mulia, Lilián; Téllez-Valencia, Alfredo; Flores-Carrillo, Paulina; Nieto-Meneses, Rocío; Castillo, Rafael

    2016-09-01

    The present work reports the synthesis and biological activity of a series of 14 benzimidazole derivatives designed to act on the enzyme triosephosphate isomerase of Trypanosoma cruzi (TcTIM). This enzyme is involved in the metabolism of glucose, the only source of energy for the parasite. In this study, we found four compounds that inhibit TcTIM moderately and lack inhibitory activity against human TIM (HsTIM). In vitro studies against T. cruzi epimastigotes showed two compounds that were more active than the reference drug nifurtimox, and these presented a low cytotoxic effect in mouse macrophages (J744 cell line). PMID:27503677

  19. Synthesis and trypanocidal activity of novel benzimidazole derivatives.

    PubMed

    Velázquez-López, José Miguel; Hernández-Campos, Alicia; Yépez-Mulia, Lilián; Téllez-Valencia, Alfredo; Flores-Carrillo, Paulina; Nieto-Meneses, Rocío; Castillo, Rafael

    2016-09-01

    The present work reports the synthesis and biological activity of a series of 14 benzimidazole derivatives designed to act on the enzyme triosephosphate isomerase of Trypanosoma cruzi (TcTIM). This enzyme is involved in the metabolism of glucose, the only source of energy for the parasite. In this study, we found four compounds that inhibit TcTIM moderately and lack inhibitory activity against human TIM (HsTIM). In vitro studies against T. cruzi epimastigotes showed two compounds that were more active than the reference drug nifurtimox, and these presented a low cytotoxic effect in mouse macrophages (J744 cell line).

  20. Synthesis and Potent Antimalarial Activity of Kalihinol B

    PubMed Central

    2016-01-01

    Of the 50+ kalihinane diterpenoids reported to date, only five had been tested for antimalarial activity, in spite of the fact that kalihinol A is the most potent among the members of the larger family of antimalarial isocyanoterpenes. We have validated a strategy designed to access many of the kalihinanes with a 12-step enantioselective synthesis of kalihinol B, the tetrahydrofuran isomer of kalihinol A (a tetrahydropyran). Kalihinol B shows similarly high potency against chloroquine-resistant Plasmodium falciparum. PMID:25815413

  1. Alternative complement pathway activation is essential for inflammation and joint destruction in the passive transfer model of collagen-induced arthritis.

    PubMed

    Banda, Nirmal K; Thurman, Joshua M; Kraus, Damian; Wood, Allyson; Carroll, Michael C; Arend, William P; Holers, V Michael

    2006-08-01

    Activation of each complement initiation pathway (classical, alternative, and lectin) can lead to the generation of bioactive fragments with resulting inflammation in target organs. The objective of the current study was to determine the role of specific complement activation pathways in the pathogenesis of experimental anti-type II collagen mAb-passive transfer arthritis. C57BL/6 mice were used that were genetically deficient in either the alternative pathway protein factor B (Bf(-/-)) or in the classical pathway component C4 (C4(-/-)). Clinical disease activity was markedly decreased in Bf(-/-) compared with wild-type (WT) mice (0.5 +/- 0.22 (n = 6) in Bf(-/-) vs 8.83 +/- 0.41 (n = 6) in WT mice (p < 0.0001)). Disease activity scores were not different between C4(-/-) and WT mice. Analyses of joints showed that C3 deposition, inflammation, pannus, cartilage, and bone damage scores were all significantly less in Bf(-/-) as compared with WT mice. There were significant decreases in mRNA levels of C3, C4, CR2, CR3, C3aR, and C5aR in the knees of Bf(-/-) as compared with C4(-/-) and WT mice with arthritis; mRNA levels for complement regulatory proteins did not differ between the three strains. These results indicate that the alternative pathway is absolutely required for the induction of arthritis following injection of anti-collagen Abs. The mechanisms by which these target organ-specific mAbs bypass the requirements for engagement of the classical pathway remain to be defined but do not appear to involve a lack of alternative pathway regulatory proteins. PMID:16849503

  2. Effect of the tripeptide glycyl-L-histidyl-L-lysine on the proliferation and synthetic activity of chick embryo chondrocytes.

    PubMed

    Pesáková, V; Novotná, J; Adam, M

    1995-08-01

    Under certain conditions chondrocytes form lattices with cartilage collagens, which may serve as cartilage implants. It is necessary to find the optimal conditions for culturing chondrocytes. Three different supports are compared: (a) plastic; (b) cartilage collagens; and (c) insoluble skin collagen solubilized under denaturing conditions (ISC-40). The effect of culture medium supplementation with the tripeptide (Gly-His-Lys)2.Cu.2H2O.2NaCl (GHK) on chondrocyte proliferation and synthetic activity is studied, with particular attention paid to collagen types I, II and III. The collagen supports stimulated chondrocyte proliferation, but on the ISC-40 support they started to dedifferentiate rather early. In the primary culture, chondrocytes on all three supports synthesized mainly collagen type II, and only small amounts of types I and III. In the first passage the synthesis of these two collagen types increased, relative to collagen type II, at least on the cartilage collagen support. Supplementation of culture medium with GHK stimulated chondrocyte proliferation in the primary structure mostly on the ISC-40 support. On the other two types of supports the stimulatory effect of GHK was expressed mostly in the first passages. The collagen synthetic rate was increased by GHK on both of the collagen supports; on the cartilage collagen support collagen type II was synthesized predominantly and on the ISC-40 support types I and III were mostly formed. It is suggested that supplementation of culture medium with GHK may be useful in the preparation of cartilage implants. PMID:8562779

  3. Temporal variation in the deposition of different types of collagen within a porous biomaterial implant.

    PubMed

    White, Jacinta F; Werkmeister, Jerome A; Bisucci, Teresa; Darby, Ian A; Ramshaw, John A M

    2014-10-01

    The deposition of new collagen in association with a medical implant has been studied using expanded polytetrafluoroethylene vascular replacement samples implanted subcutaneously in sheep, for up to 28 days. New type I collagen mRNA synthesis was followed by in situ hybridization, while the accumulation of new collagen types III, V, VI, XII, and XIV was followed by immunohistochemistry. All the collagen detected in the pores of the implant were newly deposited at various times after implantation and were not due to any pre-existing dermal collagen that may have been present around the implant. Collagen deposition was seen initially surrounding the implant and, with time, was seen to infiltrate within its pores. In situ hybridization showed that the majority of infiltrating cells had switched on mRNA that coded for type I collagen production. Histology showed that cellular infiltration increased with time, accompanied by increasing collagen deposition. The deposition of different collagen types happened at different rates. The type V and VI collagens preceded the major interstitial collagens in the newly deposited tissue, although at longer time points, detection of type V collagen appeared to decrease. After disruption of the interstitial collagens with enzyme, the "masked" type V collagen was clearly still visible by immunohistochemistry. Little type XII collagen could be seen within the porous mesh, although it was seen in the surrounding tissues. By contrast, type XIV was seen throughout the porous structure of the implanted mesh, with less being visible outside the material where type XII was more abundant.

  4. Enigmatic insight into collagen.

    PubMed

    Deshmukh, Shrutal Narendra; Dive, Alka M; Moharil, Rohit; Munde, Prashant

    2016-01-01

    Collagen is a unique, triple helical molecule which forms the major part of extracellular matrix. It is the most abundant protein in the human body, representing 30% of its dry weight. It is the fibrous structural protein that makes up the white fibers (collagen fibers) of skin, tendons, bones, cartilage and all other connective tissues. Collagens are not only essential for the mechanical resistance and resilience of multicellular organisms, but are also signaling molecules defining cellular shape and behavior. The human body has at least 16 types of collagen, but the most prominent types are I, II and III. Collagens are produced by several cell types and are distinguishable by their molecular compositions, morphologic characteristics, distribution, functions and pathogenesis. This is the major fibrous glycoprotein present in the extracellular matrix and in connective tissue and helps in maintaining the structural integrity of these tissues. It has a triple helical structure. Various studies have proved that mutations that modify folding of the triple helix result in identifiable genetic disorders. Collagen diseases share certain similarities with autoimmune diseases, because autoantibodies specific to each collagen disease are produced. Therefore, this review highlights the role of collagen in normal health and also the disorders associated with structural and functional defects in collagen. PMID:27601823

  5. Enigmatic insight into collagen

    PubMed Central

    Deshmukh, Shrutal Narendra; Dive, Alka M; Moharil, Rohit; Munde, Prashant

    2016-01-01

    Collagen is a unique, triple helical molecule which forms the major part of extracellular matrix. It is the most abundant protein in the human body, representing 30% of its dry weight. It is the fibrous structural protein that makes up the white fibers (collagen fibers) of skin, tendons, bones, cartilage and all other connective tissues. Collagens are not only essential for the mechanical resistance and resilience of multicellular organisms, but are also signaling molecules defining cellular shape and behavior. The human body has at least 16 types of collagen, but the most prominent types are I, II and III. Collagens are produced by several cell types and are distinguishable by their molecular compositions, morphologic characteristics, distribution, functions and pathogenesis. This is the major fibrous glycoprotein present in the extracellular matrix and in connective tissue and helps in maintaining the structural integrity of these tissues. It has a triple helical structure. Various studies have proved that mutations that modify folding of the triple helix result in identifiable genetic disorders. Collagen diseases share certain similarities with autoimmune diseases, because autoantibodies specific to each collagen disease are produced. Therefore, this review highlights the role of collagen in normal health and also the disorders associated with structural and functional defects in collagen.

  6. Enigmatic insight into collagen

    PubMed Central

    Deshmukh, Shrutal Narendra; Dive, Alka M; Moharil, Rohit; Munde, Prashant

    2016-01-01

    Collagen is a unique, triple helical molecule which forms the major part of extracellular matrix. It is the most abundant protein in the human body, representing 30% of its dry weight. It is the fibrous structural protein that makes up the white fibers (collagen fibers) of skin, tendons, bones, cartilage and all other connective tissues. Collagens are not only essential for the mechanical resistance and resilience of multicellular organisms, but are also signaling molecules defining cellular shape and behavior. The human body has at least 16 types of collagen, but the most prominent types are I, II and III. Collagens are produced by several cell types and are distinguishable by their molecular compositions, morphologic characteristics, distribution, functions and pathogenesis. This is the major fibrous glycoprotein present in the extracellular matrix and in connective tissue and helps in maintaining the structural integrity of these tissues. It has a triple helical structure. Various studies have proved that mutations that modify folding of the triple helix result in identifiable genetic disorders. Collagen diseases share certain similarities with autoimmune diseases, because autoantibodies specific to each collagen disease are produced. Therefore, this review highlights the role of collagen in normal health and also the disorders associated with structural and functional defects in collagen. PMID:27601823

  7. Angiotensin II enhances adenylyl cyclase signaling via Ca2+/calmodulin. Gq-Gs cross-talk regulates collagen production in cardiac fibroblasts.

    PubMed

    Ostrom, Rennolds S; Naugle, Jennifer E; Hase, Miki; Gregorian, Caroline; Swaney, James S; Insel, Paul A; Brunton, Laurence L; Meszaros, J Gary

    2003-07-01

    Cardiac fibroblasts regulate formation of extracellular matrix in the heart, playing key roles in cardiac remodeling and hypertrophy. In this study, we sought to characterize cross-talk between Gq and Gs signaling pathways and its impact on modulating collagen synthesis by cardiac fibroblasts. Angiotensin II (ANG II) activates cell proliferation and collagen synthesis but also potentiates cyclic AMP (cAMP) production stimulated by beta-adrenergic receptors (beta-AR). The potentiation of beta-AR-stimulated cAMP production by ANG II is reduced by phospholipase C inhibition and enhanced by overexpression of Gq. Ionomycin and thapsigargin increased intracellular Ca2+ levels and potentiated isoproterenol- and forskolin-stimulated cAMP production, whereas chelation of Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid/AM inhibited such potentiation. Inhibitors of tyrosine kinases, protein kinase C, or Gbetagamma did not alter this cross-talk. Immunoblot analyses showed prominent expression of adenylyl cyclase 3 (AC3), a Ca2+-activated isoform, along with AC2, AC4, AC5, AC6, and AC7. Of those isoforms, only AC3 and AC5/6 proteins were detected in caveolin-rich fractions. Overexpression of AC6 increased betaAR-stimulated cAMP accumulation but did not alter the size of the ANG II potentiation, suggesting that the cross-talk is AC isoform-specific. Isoproterenol-mediated inhibition of serum-stimulated collagen synthesis increased from 31 to 48% in the presence of ANG II, indicating that betaAR-regulated collagen synthesis increased in the presence of ANG II. These data indicate that ANG II potentiates cAMP formation via Ca2+-dependent activation of AC activity, which in turn attenuates collagen synthesis and demonstrates one functional consequence of cross-talk between Gq and Gs signaling pathways in cardiac fibroblasts. PMID:12711600

  8. Angiotensin II enhances adenylyl cyclase signaling via Ca2+/calmodulin. Gq-Gs cross-talk regulates collagen production in cardiac fibroblasts.

    PubMed

    Ostrom, Rennolds S; Naugle, Jennifer E; Hase, Miki; Gregorian, Caroline; Swaney, James S; Insel, Paul A; Brunton, Laurence L; Meszaros, J Gary

    2003-07-01

    Cardiac fibroblasts regulate formation of extracellular matrix in the heart, playing key roles in cardiac remodeling and hypertrophy. In this study, we sought to characterize cross-talk between Gq and Gs signaling pathways and its impact on modulating collagen synthesis by cardiac fibroblasts. Angiotensin II (ANG II) activates cell proliferation and collagen synthesis but also potentiates cyclic AMP (cAMP) production stimulated by beta-adrenergic receptors (beta-AR). The potentiation of beta-AR-stimulated cAMP production by ANG II is reduced by phospholipase C inhibition and enhanced by overexpression of Gq. Ionomycin and thapsigargin increased intracellular Ca2+ levels and potentiated isoproterenol- and forskolin-stimulated cAMP production, whereas chelation of Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid/AM inhibited such potentiation. Inhibitors of tyrosine kinases, protein kinase C, or Gbetagamma did not alter this cross-talk. Immunoblot analyses showed prominent expression of adenylyl cyclase 3 (AC3), a Ca2+-activated isoform, along with AC2, AC4, AC5, AC6, and AC7. Of those isoforms, only AC3 and AC5/6 proteins were detected in caveolin-rich fractions. Overexpression of AC6 increased betaAR-stimulated cAMP accumulation but did not alter the size of the ANG II potentiation, suggesting that the cross-talk is AC isoform-specific. Isoproterenol-mediated inhibition of serum-stimulated collagen synthesis increased from 31 to 48% in the presence of ANG II, indicating that betaAR-regulated collagen synthesis increased in the presence of ANG II. These data indicate that ANG II potentiates cAMP formation via Ca2+-dependent activation of AC activity, which in turn attenuates collagen synthesis and demonstrates one functional consequence of cross-talk between Gq and Gs signaling pathways in cardiac fibroblasts.

  9. Synthesis and antiproliferative activity of novel steroidal dendrimer conjugates.

    PubMed

    Magaña-Vergara, Nancy E; Rárová, Lucie; Soto-Castro, Delia; Farfán, Norberto; Strnad, Miroslav; Santillan, Rosa

    2013-12-11

    We describe the synthesis of steroidal dendrimer conjugates of first and second generation with tetramethylene core and 5-hydroxy-isophtalic acid dimethyl ester as branching unit modified to incorporate ethynylestradiol or 17α-estradiol as terminal units. The steroidal dendrimer conjugates, the free drug (steroids) and dendrimer were tested against a panel of cancer cell lines (CEM, MCF7, HeLa) and normal human fibroblast (BJ). The steroidal dendrimer conjugates of first generation exhibited cytotoxic activity and induced apoptosis in chronic leukemia (CEM) as resultant activation of caspase cascade which is mainly provoked in G2/M arrested cells.

  10. Synthesis and antitumor activity of tetrahydrocarbazole hybridized with dithioate derivatives.

    PubMed

    El-Nassan, Hala Bakr

    2015-04-01

    The present study reported the synthesis of tetrahydrocarbazoles hybridized with dithioate derivatives. Three series were synthesized namely alkyl dithiocarbonates (4a-d), heterocyclic dithiocarbamates (6a-g) and dialkyl dithiocarbamate (7). The synthesized compounds were tested in vitro on human breast adenocarcinoma cell line (MCF7) and the human colon tumor cell line (HCT116). Most of the synthesized compounds exploited potent antitumor activity, especially compound 6f [4-chlorophenylpiperazine derivative], which showed cytotoxic activity against MCF7 superior to doxorubicin with IC50 value of 7.24 nM/mL. PMID:24899376

  11. Photodynamically crosslinked and chitosan-incorporated dentin collagen.

    PubMed

    Shrestha, A; Friedman, S; Kishen, A

    2011-11-01

    A lingering concern with restored root-filled teeth is the loss of structural integrity of the dentin and dentin-sealer interface over time. We hypothesized that crosslinking of dentin collagen with simultaneous incorporation of a biopolymer into collagen matrix would improve its structural stability. This study aimed to investigate the effects of combining chemical/photodynamic crosslinking of dentin collagen with the incorporation of carboxymethyl-chitosan (CMCS) on the resistance to enzymatic degradation and mechanical properties of dentin collagen. Ninety-six demineralized dentin collagen specimens (human, n = 72; and bovine, n = 24) were prepared and crosslinked chemically/ photodynamically, with/without CMCS. Glutaraldehyde and carbodiimides were used for chemical crosslinking, while rose Bengal activated with a non-coherent light (540 nm) at 20 J/cm(2) was applied for photodynamic crosslinking. The crosslinked human dentin collagen was subjected to chemical characterization, 7 days enzymatic degradation, and transmission electron microscopy (TEM), while the bovine dentin collagen was used for tensile-testing. Crosslinked collagen showed significantly higher resistance to enzymatic degradation (p < 0.01), stable ultrastructure, and increased tensile strength (p < 0.05). Crosslinking CMCS with collagen matrix as observed in the TEM further improved the mechanical properties of dentin collagen (p < 0.01). This study highlighted the possibility of improving the resistance and toughness of dentin collagen by chemically/photodynamically crosslinking collagen matrix with CMCS.

  12. Aspirin suppresses cardiac fibroblast proliferation and collagen formation through downregulation of angiotensin type 1 receptor transcription

    SciTech Connect

    Wang, Xianwei Lu, Jingjun; Khaidakov, Magomed; Mitra, Sona; Ding, Zufeng; Raina, Sameer; Goyal, Tanu; Mehta, Jawahar L.

    2012-03-15

    Aspirin (acetyl salicylic acid, ASA) is a common drug used for its analgesic and antipyretic effects. Recent studies show that ASA not only blocks cyclooxygenase, but also inhibits NADPH oxidase and resultant reactive oxygen species (ROS) generation, a pathway that underlies pathogenesis of several ailments, including hypertension and tissue remodeling after injury. In these disease states, angiotensin II (Ang II) activates NADPH oxidase via its type 1 receptor (AT1R) and leads to fibroblast growth and collagen synthesis. In this study, we examined if ASA would inhibit NADPH oxidase activation, upregulation of AT1R transcription, and subsequent collagen generation in mouse cardiac fibroblasts challenged with Ang II. Mouse heart fibroblasts were isolated and treated with Ang II with or without ASA. As expected, Ang II induced AT1R expression, and stimulated cardiac fibroblast growth and collagen synthesis. The AT1R blocker losartan attenuated these effects of Ang II. Similarly to losartan, ASA, and its SA moiety suppressed Ang II-mediated AT1R transcription and fibroblast proliferation as well as expression of collagens and MMPs. ASA also suppressed the expression of NADPH oxidase subunits (p22{sup phox}, p47{sup phox}, p67{sup phox}, NOX2 and NOX4) and ROS generation. ASA did not affect total NF-κB p65, but inhibited its phosphorylation and activation. These observations suggest that ASA inhibits Ang II-induced NADPH oxidase expression, NF-κB activation and AT1R transcription in cardiac fibroblasts, and fibroblast proliferation and collagen expression. The critical role of NADPH oxidase activity in stimulation of AT1R transcription became apparent in experiments where ASA also inhibited AT1R transcription in cardiac fibroblasts challenged with H{sub 2}O{sub 2}. Since SA had similar effect as ASA on AT1R expression, we suggest that ASA's effect is mediated by its SA moiety. -- Highlights: ► Aspirin in therapeutic concentrations decreases mouse cardiac fibroblast

  13. Synthesis and cytotoxic activities of semisynthetic zearalenone analogues.

    PubMed

    Tadpetch, Kwanruthai; Kaewmee, Benyapa; Chantakaew, Kittisak; Kantee, Kawalee; Rukachaisirikul, Vatcharin; Phongpaichit, Souwalak

    2016-08-01

    Zearalenone is a β-resorcylic acid macrolide with various biological activities. Herein we report the synthesis and cytotoxic activities of 34 zearalenone analogues against human oral epidermoid carcinoma (KB) and human breast adenocarcinoma (MCF-7) cells as well as noncancerous Vero cells. Some zearalenone analogues showed moderately enhanced cytotoxic activities against the two cancer cell lines. We have discovered the potential lead compounds with diminished or no cytotoxicity to Vero cells. Preliminary structure-activity relationship studies revealed that the double bond at the 1' and 2' positions of zearalenone core was crucial for cytotoxic activities on both cell lines. In addition, for zearalenol analogues, the unprotected hydroxyl group at C-2 and an alkoxy substituent at C-4 played key roles on cytotoxic effects of both cell lines.

  14. Immunosuppressive activity of deer antler extracts of Cervus korean TEMMINCK var. mantchuricus Swinhoe, on type II collagen-induced arthritis.

    PubMed

    Kang, Sung-Koo; Kim, Kap-Sung; Kim, Sung-Il; Chung, Kang-Hyun; Lee, In-Seon; Kim, Cheorl-Ho

    2006-01-01

    Unossified horn or pilose antler cut from deer, which belong to the Cervidae generally is termed Nokyong. Nokyong is one of the most famous Korean traditional medicines and has been considered to possess sexual-reinforcing and antiaging actions. In this study, water extract of deer antler extract (DAA) prepared from the growing antler of Cervus korean TEMMINCK var. mantchuricus Swinhoe was used to investigate the efficacy of the DAA on the development of type II collagen (CII)-induced arthritis (CIA) in rats. Male rats were immunized with an emulsion of 200 microg of CII and complete Freund's adjuvant (CFA). The rats then were administered by injection a suspension of DAA or phosphate-buffered saline. The effect of DAA on cellular responses to CII was examined. The injection of DAA suppressed the CII-specific secretion of interferon (IFN)-gamma from splenocytes ex vivo. The influence of DAA also was evaluated on the incidence and development of arthritis in rat CIA. Rats were immunized twice at a 3-wk interval with bovine CII, with DAA being given by injection once a d for 14 d with four different regimens. A 14-d course of DAA treatment at a daily dose of 100 microg/kg, which began on the d of the first CII immunization, suppressed the development of arthritis, as well as antibody formation and delayed-type hypersensitivity to CII. Treatment with DAA resulted in inhibition of development of arthritis and immune responses to CII.

  15. Unusual Glycosaminoglycans from a Deep Sea Hydrothermal Bacterium Improve Fibrillar Collagen Structuring and Fibroblast Activities in Engineered Connective Tissues

    PubMed Central

    Senni, Karim; Gueniche, Farida; Changotade, Sylvie; Septier, Dominique; Sinquin, Corinne; Ratiskol, Jacqueline; Lutomski, Didier; Godeau, Gaston; Guezennec, Jean; Colliec-Jouault, Sylvia

    2013-01-01

    Biopolymers produced by marine organisms can offer useful tools for regenerative medicine. Particularly, HE800 exopolysaccharide (HE800 EPS) secreted by a deep-sea hydrothermal bacterium displays an interesting glycosaminoglycan-like feature resembling hyaluronan. Previous studies demonstrated its effectiveness to enhance in vivo bone regeneration and to support osteoblastic cell metabolism in culture. Thus, in order to assess the usefulness of this high-molecular weight polymer in tissue engineering and tissue repair, in vitro reconstructed connective tissues containing HE800 EPS were performed. We showed that this polysaccharide promotes both collagen structuring and extracellular matrix settle by dermal fibroblasts. Furthermore, from the native HE800 EPS, a low-molecular weight sulfated derivative (HE800 DROS) displaying chemical analogy with heparan-sulfate, was designed. Thus, it was demonstrated that HE800 DROS mimics some properties of heparan-sulfate, such as promotion of fibroblast proliferation and inhibition of matrix metalloproteinase (MMP) secretion. Therefore, we suggest that the HE800EPS family can be considered as an innovative biotechnological source of glycosaminoglycan-like compounds useful to design biomaterials and drugs for tissue engineering and repair. PMID:23612369

  16. Collagenous gastritis: Review

    PubMed Central

    Kamimura, Kenya; Kobayashi, Masaaki; Sato, Yuichi; Aoyagi, Yutaka; Terai, Shuji

    2015-01-01

    Collagenous gastritis is a rare disease characterized by the subepithelial deposition of collagen bands thicker than 10 μm and the infiltration of inflammatory mononuclear cells in the lamina propria. Collagenous colitis and collagenous sprue have similar histological characteristics to collagenous gastritis and are thought to be part of the same disease entity. However, while collagenous colitis has become more common in the field of gastroenterology, presenting with clinical symptoms of chronic diarrhea in older patients, collagenous gastritis is rare. Since the disease was first reported in 1989, only 60 cases have been documented in the English literature. No safe and effective treatments have been identified from randomized, controlled trials. Therefore, better understanding of the disease and the reporting of more cases will help to establish diagnostic criteria and to develop therapeutic strategies. Therefore, here we review the clinical characteristics, endoscopic and histological findings, treatment, and clinical outcomes from case reports and case series published to date, and provide a summary of the latest information on the disease. This information will contribute to improved knowledge of collagenous gastritis so physicians can recognize and correctly diagnose the disease, and will help to develop a standard therapeutic strategy for future clinical trials. PMID:25789098

  17. Collagenous gastritis: Review.

    PubMed

    Kamimura, Kenya; Kobayashi, Masaaki; Sato, Yuichi; Aoyagi, Yutaka; Terai, Shuji

    2015-03-16

    Collagenous gastritis is a rare disease characterized by the subepithelial deposition of collagen bands thicker than 10 μm and the infiltration of inflammatory mononuclear cells in the lamina propria. Collagenous colitis and collagenous sprue have similar histological characteristics to collagenous gastritis and are thought to be part of the same disease entity. However, while collagenous colitis has become more common in the field of gastroenterology, presenting with clinical symptoms of chronic diarrhea in older patients, collagenous gastritis is rare. Since the disease was first reported in 1989, only 60 cases have been documented in the English literature. No safe and effective treatments have been identified from randomized, controlled trials. Therefore, better understanding of the disease and the reporting of more cases will help to establish diagnostic criteria and to develop therapeutic strategies. Therefore, here we review the clinical characteristics, endoscopic and histological findings, treatment, and clinical outcomes from case reports and case series published to date, and provide a summary of the latest information on the disease. This information will contribute to improved knowledge of collagenous gastritis so physicians can recognize and correctly diagnose the disease, and will help to develop a standard therapeutic strategy for future clinical trials. PMID:25789098

  18. The exercise-induced biochemical milieu enhances collagen content and tensile strength of engineered ligaments.

    PubMed

    West, Daniel W D; Lee-Barthel, Ann; McIntyre, Todd; Shamim, Baubak; Lee, Cassandra A; Baar, Keith

    2015-10-15

    Exercise stimulates a dramatic change in the concentration of circulating hormones, such as growth hormone (GH), but the biological functions of this response are unclear. Pharmacological GH administration stimulates collagen synthesis; however, whether the post-exercise systemic milieu has a similar action is unknown. We aimed to determine whether the collagen content and tensile strength of tissue-engineered ligaments is enhanced by serum obtained post-exercise. Primary cells from a human anterior cruciate ligament (ACL) were used to engineer ligament constructs in vitro. Blood obtained from 12 healthy young men 15 min after resistance exercise contained GH concentrations that were ∼7-fold greater than resting serum (P < 0.001), whereas IGF-1 was not elevated at this time point (P = 0.21 vs. rest). Ligament constructs were treated for 7 days with medium supplemented with serum obtained at rest (RestTx) or 15 min post-exercise (ExTx), before tensile testing and collagen content analysis. Compared with RestTx, ExTx enhanced collagen content (+19%; 181 ± 33 vs. 215 ± 40 μg per construct P = 0.001) and ligament mechanical properties - maximal tensile load (+17%, P = 0.03 vs. RestTx) and ultimate tensile strength (+10%, P = 0.15 vs. RestTx). In a separate set of engineered ligaments, recombinant IGF-1, but not GH, enhanced collagen content and mechanics. Bioassays in 2D culture revealed that acute treatment with post-exercise serum activated mTORC1 and ERK1/2. In conclusion, the post-exercise biochemical milieu, but not recombinant GH, enhances collagen content and tensile strength of engineered ligaments, in association with mTORC1 and ERK1/2 activation.

  19. Collagen: Biochemistry, biomechanics, biotechnology

    SciTech Connect

    Nimni, M.E.

    1988-01-01

    This book is an up-to-date reference for new ideas, information, and concepts in collagen research. The first volume emphasizes the relationship between the molecular structure and function of collagen, including descriptions of collagen types which exist in tissues as well as how these molecules organize into fibrils and the nature of the chemical crosslinks which stabilize them. In Volume II the biomechanical behavior of various specialized tissues, abnormal accumulation of collagen in the form of scars of fibrous infiltration are examined/and wound healing, tissue regulation and repair are covered in detail. Volume III explores the increasing application of collagen technology to the field of bioprosthesis, including the production of heart valve bioprosthesis, blood vessels, ligament substitutes, and bone substitutes.

  20. Backbone dynamics in collagen

    NASA Astrophysics Data System (ADS)

    Aliev, Abil E.

    2004-11-01

    Peptide backbone motions of collagen have been extensively studied in the past. The experimental results were interpreted using a model of a collagen rod librating about its helix axis. Considering the size of the collagen molecule and the presence of cross-linked molecules, motional amplitudes derived for the helix axis libration were unusually high. Using solid-state NMR 13C chemical shift anisotropy and 2H quadrupolar lineshape analysis for five different isotope labelled collagens we show that motional averaging of the NMR interactions occurs primarily via small-angle librations about internal bond directions. This type of dynamics is compatible with both the presence of cross-links in collagen and the X-ray data, as well as dynamic models used for other proteins.

  1. Vitamins E and C may increase collagen turnover by intramuscular fibroblasts. Potential for improved meat quality.

    PubMed

    Archile-Contreras, Anangelina C; Cha, Ming C; Mandell, Ira B; Miller, Stephen P; Purslow, Peter P

    2011-01-26

    Vitamins influence collagen metabolism in animals grown for meat. This study investigated whether vitamins E and C regulate collagen turnover in muscle by the balance of effects on the synthesis of collagen and its degradation by secretion of matrix metalloproteinases (MMPs) by bovine intramuscular fibroblasts. Fibroblasts isolated from longissimus dorsi (LD) and semitendinosus (ST) muscle were treated with different concentrations of vitamins. Pro-MMP-2, MMP-2, and total soluble collagen (TSC) synthesis were determined. Vitamins E and C each preferentially increased (P < 0.05) MMP-2 in cells derived from LD relative to those derived from ST. Higher TSC values (P < 0.05) were found for ST cells than for LD cells. Both vitamins may increase collagen turnover exerted by intramuscular connective tissue fibroblasts. These results may have implications in vivo on animal production, as a high rate of collagen turnover may lead to increased collagen solubility in muscles, which can affect meat tenderness.

  2. Activation of Melanin Synthesis in Alternaria infectoria by Antifungal Drugs

    PubMed Central

    Fernandes, Chantal; Prados-Rosales, Rafael; Silva, Branca M. A.; Nakouzi-Naranjo, Antonio; Zuzarte, Mónica; Chatterjee, Subhasish; Stark, Ruth E.; Casadevall, Arturo

    2015-01-01

    The importance of Alternaria species fungi to human health ranges from their role as etiological agents of serious infections with poor prognoses in immunosuppressed individuals to their association with respiratory allergic diseases. The present work focuses on Alternaria infectoria, which was used as a model organism of the genus, and was designed to unravel melanin production in response to antifungals. After we characterized the pigment produced by A. infectoria, we studied the dynamics of 1,8-dihydroxynaphthalene (DHN)-melanin production during growth, the degree of melanization in response to antifungals, and how melanization affected susceptibility to several classes of therapeutic drugs. We demonstrate that A. infectoria increased melanin deposition in cell walls in response to nikkomycin Z, caspofungin, and itraconazole but not in response to fluconazole or amphotericin B. These results indicate that A. infectoria activates DHN-melanin synthesis in response to certain antifungal drugs, possibly as a protective mechanism against these drugs. Inhibition of DHN-melanin synthesis by pyroquilon resulted in a lower minimum effective concentration (MEC) of caspofungin and enhanced morphological changes (increased hyphal balloon size), characterized by thinner and less organized A. infectoria cell walls. In summary, A. infectoria synthesizes melanin in response to certain antifungal drugs, and its susceptibility is influenced by melanization, suggesting the therapeutic potential of drug combinations that affect melanin synthesis. PMID:26711773

  3. Abnormal collagen binding activity of 2A von Willebrand factor: evidence that the defect depends only on the lack of large multimers.

    PubMed

    Casonato, A; Pontara, E; Bertomoro, A; Zucchetto, S; Zerbinati, P; Girolami, A

    1997-02-01

    It is well established that the large von Willebrand factor (vWf) multimers bind with high affinity to the extracellular matrix. To explore the different roles of intermediate and large vWf multimers, we studied the collagen-binding activity (vWf:CBA) of 2A vWf under nonflowing conditions in relation to the multimer organization of the molecule. Regardless of the anticoagulant used for blood collection, vWf:CBA was significantly decreased, in 4 patients with 2A von Willebrand's disease (vWd), in accordance with the lack of high and intermediate vWf multimers. After 1-deamino-8-D-arginine vasopressin (DDAVP) infusion, the appearance of circulating large and unusually large vWf multimers, in samples collected in the presence of protease inhibitors, induced a complete normalization of vWf:CBA. The peak was observed 15 minutes after DDAVP, when large and unusually large multimers were maximally represented. These effects were transient because vWf:CBA decreased after 60 minutes, even though values were still significantly higher than pre-DDAVP figures; at the same time, large vWf multimers appeared to be decreased. In contrast, samples anticoagulated with sodium citrate after DDAVP did not show a normalized vWf multimer pattern and were characterized by a persistently decreased vWf:CBA. Moreover, in all of the patients studied, platelet vWf presented normal vWf:CBA values in accordance with the normal levels and multimer organization of the vWf molecule. Our findings indicate that the collagen-binding defect displayed in vitro by type 2A vWf depends only on the lack of circulating large vWf multimers. Moreover, the observation of normal platelet vWf:CBA seems to indicate a primary role of plasma rather than platelet vWf in assuring platelet plug formation.

  4. The collagen binding domain of gelatinase A modulates degradation of collagen IV by gelatinase B.

    PubMed

    Gioia, Magda; Monaco, Susanna; Van Den Steen, Philippe E; Sbardella, Diego; Grasso, Giuseppe; Marini, Stefano; Overall, Christopher M; Opdenakker, Ghislain; Coletta, Massimo

    2009-02-20

    Type IV collagen remodeling plays a critical role in inflammatory responses, angiogenesis and metastasis. Its remodeling is executed by a family of matrix metalloproteinases (MMPs), of which the constitutive gelatinase A (MMP2) and the inducible gelatinase B (MMP9) are key examples. Thus, in many pathological conditions, both gelatinases act together. Kinetic data are reported for the enzymatic processing at 37 degrees C of type IV collagen from human placenta by MMP9 and its modulation by the fibronectin-like collagen binding domain (CBD) of MMP2. The alpha1 and alpha2 chain components of type IV collagen were cleaved by gelatinases and identified by mass spectrometry as well as Edman sequencing. Surface plasmon resonance interaction assays showed that CBD bound type IV collagen at two topologically distinct sites. On the basis of linked-function analysis, we demonstrated that CBD of MMP2 tuned the cleavage of collagen IV by MMP9, presumably by inducing a ligand-linked structural change on the type IV collagen. At low concentrations, the CBD bound the first site and thereby allosterically modulated the binding of MMP9 to collagen IV, thus enhancing the collagenolytic activity of MMP9. At high concentrations, CBD binding to the second site interfered with MMP9 binding to collagen IV, acting as a competitive inhibitor. Interestingly, modulation of collagen IV degradation by inactive forms of MMP2 also occurred in a cell-based system, revealing that this interrelationship affected neutrophil migration across a collagen IV membrane. The regulation of the proteolytic processing by a catalytically inactive domain (i.e., CBD) suggests that the two gelatinases might cooperate in degrading substrates even when either one is inactive. This observation reinforces the idea of exosite targets for MMP inhibitors, which should include all macromolecular substrate recognition sites.

  5. Long-term overproduction of collagen in radiation-induced fibrosis

    SciTech Connect

    Remy, J.; Wegrowski, J.; Crechet, F.; Martin, M.; Daburon, F. )

    1991-01-01

    Collagen metabolism was investigated in the fibrotic tissue which developed in pig thigh muscle 6 to 15 months after acute gamma irradiation. During this period, total collagen deposits in the fibrotic tissue increased 10-fold compared to the healthy muscle tissue. These deposits were composed mainly of type I and III collagen, and the type I/type III ratio was lower in the fibrotic than in the muscle tissue. Small pieces of both fibrotic and muscle tissue were incubated with (14C)proline. The (14C)hydroxyproline content of the fibrotic tissue reflected large concomitant increases in the synthesis of total collagen, mainly of types I and III, which rose 14- and 17-fold, respectively. Similarly, the level of type I and type III procollagen mRNAs rose 9- and 5-fold, respectively, in the fibrotic tissue versus the muscle tissue. These results suggest that procollagen gene transcription or RNA maturation in the cell nuclei is activated in the fibrotic tissue. The possibility that such activation is due to the long-term inflammatory state of this tissue is discussed.

  6. ETP-46321, a dual p110α/δ class IA phosphoinositide 3-kinase inhibitor modulates T lymphocyte activation and collagen-induced arthritis.

    PubMed

    Aragoneses-Fenoll, L; Montes-Casado, M; Ojeda, G; Acosta, Y Y; Herranz, J; Martínez, S; Blanco-Aparicio, C; Criado, G; Pastor, J; Dianzani, U; Portolés, P; Rojo, J M

    2016-04-15

    Class IA phosphoinositide 3-kinases (PI3Ks) are essential to function of normal and tumor cells, and to modulate immune responses. T lymphocytes express high levels of p110α and p110δ class IA PI3K. Whereas the functioning of PI3K p110δ in immune and autoimmune reactions is well established, the role of p110α is less well understood. Here, a novel dual p110α/δ inhibitor (ETP-46321) and highly specific p110α (A66) or p110δ (IC87114) inhibitors have been compared concerning T cell activation in vitro, as well as the effect on responses to protein antigen and collagen-induced arthritis in vivo. In vitro activation of naive CD4(+) T lymphocytes by anti-CD3 and anti-CD28 was inhibited more effectively by the p110δ inhibitor than by the p110α inhibitor as measured by cytokine secretion (IL-2, IL-10, and IFN-γ), T-bet expression and NFAT activation. In activated CD4(+) T cells re-stimulated through CD3 and ICOS, IC87114 inhibited Akt and Erk activation, and the secretion of IL-2, IL-4, IL-17A, and IFN-γ better than A66. The p110α/δ inhibitor ETP-46321, or p110α plus p110δ inhibitors also inhibited IL-21 secretion by differentiated CD4(+) T follicular (Tfh) or IL-17-producing (Th17) helper cells. In vivo, therapeutic administration of ETP-46321 significantly inhibited responses to protein antigen as well as collagen-induced arthritis, as measured by antigen-specific antibody responses, secretion of IL-10, IL-17A or IFN-γ, or clinical symptoms. Hence, p110α as well as p110δ Class IA PI3Ks are important to immune regulation; inhibition of both subunits may be an effective therapeutic approach in inflammatory autoimmune diseases like rheumatoid arthritis. PMID:26883061

  7. Effect of Allium sativum and fish collagen on the proteolytic and angiotensin-I converting enzyme-inhibitory activities in cheese and yogurt.

    PubMed

    Shori, A B; Baba, A S; Keow, J N

    2012-12-15

    There is an increasing demand of functional foods in developed countries. Yogurt plays an important role in the management of blood pressure. Several bioactive peptides isolated from Allium sativum or fish collagen have shown antihypertensive activity. Thus, in the present study the effects of A. sativum and/or Fish Collagen (FC) on proteolysis and ACE inhibitory activity in yogurt (0, 7 and 14 day) and cheese (0, 14 and 28 day) were investigated. Proteolytic activities were the highest on day 7 of refrigerated storage in A. sativum-FC-yogurt (337.0 +/- 5.3 microg g(-1)) followed by FC-yogurt (275.3 +/- 2.0 microg g(-1)), A. sativum-yogurt (245.8 +/- 4.2 microg g(-1)) and plain-yogurt (40.4 +/- 1.2 microg g(-1)). On the other hand, proteolytic activities in cheese ripening were the highest (p < 0.05) on day 14 of storage for plain and A. sativum-cheeses (411.4 +/- 4.3 and 528.7 +/- 1.6 microg g(-1), respectively). However, the presence of FC increased the proteolysis to the highest level on day 28 of storage for FC- and A. sativum-FC cheeses (641.2 +/- 0.1 and 1128.4 +/- 4.5 microg g(-1), respectively). In addition, plain- and A. sativum-yogurts with or without FC showed maximal inhibition of ACE on day 7 of storage. Fresh plain- and A. sativum-cheeses showed ACE inhibition (72.3 +/- 7.8 and 50.4 +/- 1.6 % respectively), the presence of FC in both type of cheeses reduced the ACE inhibition to 62.9 +/- 0.8 and 44.5 +/- 5.0%, respectively. However, refrigerated storage increased ACE inhibition in cheeses (p < 0.05 on day 28) in the presence of FC more than in the absence. In conclusion, the presence of FC in A. sativum-yogurt or cheese enhanced the proteolytic activity. Thus, it has potential in the development of an effective dietary strategy for hypertension associated cardiovascular diseases.

  8. Synthesis and biological activity of novel deoxycholic acid derivatives.

    PubMed

    Popadyuk, Irina I; Markov, Andrey V; Salomatina, Oksana V; Logashenko, Evgeniya B; Shernyukov, Andrey V; Zenkova, Marina A; Salakhutdinov, Nariman F

    2015-08-01

    We report the synthesis and biological activity of new semi-synthetic derivatives of naturally occurring deoxycholic acid (DCA) bearing 2-cyano-3-oxo-1-ene, 3-oxo-1(2)-ene or 3-oxo-4(5)-ene moieties in ring A and 12-oxo or 12-oxo-9(11)-ene moieties in ring C. Bioassays using murine macrophage-like cells and tumour cells show that the presence of the 9(11)-double bond associated with the increased polarity of ring A or with isoxazole ring joined to ring A, improves the ability of the compounds to inhibit cancer cell growth. PMID:26037611

  9. Synthesis and antituberculosis activity of new fatty acid amides.

    PubMed

    D'Oca, Caroline Da Ros Montes; Coelho, Tatiane; Marinho, Tamara Germani; Hack, Carolina Rosa Lopes; Duarte, Rodrigo da Costa; da Silva, Pedro Almeida; D'Oca, Marcelo Gonçalves Montes

    2010-09-01

    This work reports the synthesis of new fatty acid amides from C16:0, 18:0, 18:1, 18:1 (OH), and 18:2 fatty acids families with cyclic and acyclic amines and demonstrate for the first time the activity of these compounds as antituberculosis agents against Mycobacterium tuberculosis H(37)Rv, M. tuberculosis rifampicin resistance (ATCC 35338), and M. tuberculosis isoniazid resistance (ATCC 35822). The fatty acid amides derivate from ricinoleic acid were the most potent one among a series of tested compounds, with a MIC 6.25 microg/mL for resistance strains.

  10. Synthesis and biological activity of novel deoxycholic acid derivatives.

    PubMed

    Popadyuk, Irina I; Markov, Andrey V; Salomatina, Oksana V; Logashenko, Evgeniya B; Shernyukov, Andrey V; Zenkova, Marina A; Salakhutdinov, Nariman F

    2015-08-01

    We report the synthesis and biological activity of new semi-synthetic derivatives of naturally occurring deoxycholic acid (DCA) bearing 2-cyano-3-oxo-1-ene, 3-oxo-1(2)-ene or 3-oxo-4(5)-ene moieties in ring A and 12-oxo or 12-oxo-9(11)-ene moieties in ring C. Bioassays using murine macrophage-like cells and tumour cells show that the presence of the 9(11)-double bond associated with the increased polarity of ring A or with isoxazole ring joined to ring A, improves the ability of the compounds to inhibit cancer cell growth.

  11. Optimal placement of active elements in control augmented structural synthesis

    NASA Technical Reports Server (NTRS)

    Sepulveda, A. E.; Jin, I. M.; Schmit, L. A., Jr.

    1992-01-01

    A methodology for structural/control synthesis is presented in which the optimal location of active members is treated in terms of (0,1) variables. Structural member sizes, control gains and (0,1) placement variables are treated simultaneously as design variables. Optimization is carried out by generating and solving a sequence of explicit approximate problems using a branch and bound strategy. Intermediate design variable and intermediate response quantity concepts are used to enhance the quality of the approximate design problems. Numerical results for example problems are presented to illustrate the efficacy of the design procedure set forth.

  12. Antifungal activity of silver nanoparticles obtained by green synthesis.

    PubMed

    Mallmann, Eduardo José J; Cunha, Francisco Afrânio; Castro, Bruno N M F; Maciel, Auberson Martins; Menezes, Everardo Albuquerque; Fechine, Pierre Basílio Almeida

    2015-01-01

    Silver nanoparticles (AgNPs) are metal structures at the nanoscale. AgNPs have exhibited antimicrobial activities against fungi and bacteria; however synthesis of AgNPs can generate toxic waste during the reaction process. Accordingly, new routes using non-toxic compounds have been researched. The proposal of the present study was to synthesize AgNPs using ribose as a reducing agent and sodium dodecyl sulfate (SDS) as a stabilizer. The antifungal activity of these particles against C. albicans and C. tropicalis was also evaluated. Stable nanoparticles 12.5 ± 4.9 nm (mean ± SD) in size were obtained, which showed high activity against Candida spp. and could represent an alternative for fungal infection treatment. PMID:25923897

  13. ANTIFUNGAL ACTIVITY OF SILVER NANOPARTICLES OBTAINED BY GREEN SYNTHESIS

    PubMed Central

    MALLMANN, Eduardo José J.; CUNHA, Francisco Afrânio; CASTRO, Bruno N.M.F.; MACIEL, Auberson Martins; MENEZES, Everardo Albuquerque; FECHINE, Pierre Basílio Almeida

    2015-01-01

    Silver nanoparticles (AgNPs) are metal structures at the nanoscale. AgNPs have exhibited antimicrobial activities against fungi and bacteria; however synthesis of AgNPs can generate toxic waste during the reaction process. Accordingly, new routes using non-toxic compounds have been researched. The proposal of the present study was to synthesize AgNPs using ribose as a reducing agent and sodium dodecyl sulfate (SDS) as a stabilizer. The antifungal activity of these particles against C. albicans and C. tropicalis was also evaluated. Stable nanoparticles 12.5 ± 4.9 nm (mean ± SD) in size were obtained, which showed high activity against Candida spp. and could represent an alternative for fungal infection treatment. PMID:25923897

  14. Facile synthesis and antibacterial activity of naturally occurring 5-methoxyfuroflavone.

    PubMed

    Alam, Mohammad Sayed; Lee, Dong-Ung

    2010-12-01

    A convenient synthesis of 5-methoxyfuroflavone (6, pongaglabol methyl ether), a constituent of some Pongamia or Millettia genus, was achieved by starting from 2,4-dihydroxy-6-methoxyacetophenone via a chalcone precursor, followed by treatment with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ). This five-step reaction (total yield: 21.6%) is more facile with that of previously utilized procedures using each different starting material. Antibacterial activities of the above compound and its precursor chalcones, which also belongs to the class of furoflavonoids, were tested by the disc diffusion method against Shigella dysenteriae, Salmonella typhi, Streptococcus-β-haemolyticus, and Staphylococcus aureus. 5-Methoxyfuroflavone showed moderate bactericidal activity against all tested bacterial strains, whereas its corresponding chalcone compound revealed a selective activity. PMID:21139271

  15. Ecological function of myroilysin, a novel bacterial M12 metalloprotease with elastinolytic activity and a synergistic role in collagen hydrolysis, in biodegradation of deep-sea high-molecular-weight organic nitrogen.

    PubMed

    Chen, Xiu-Lan; Xie, Bin-Bin; Bian, Fei; Zhao, Guo-Yan; Zhao, Hui-Lin; He, Hai-Lun; Zhou, Bai-Cheng; Zhang, Yu-Zhong

    2009-04-01

    Nearly all high-molecular-weight (HMW) dissolved organic nitrogen and part of the particulate organic nitrogen in the deep sea are present in hydrolysis-resistant amides, and so far the mechanisms of biodegradation of these types of nitrogen have not been resolved. The M12 family is the second largest family in subclan MA(M) of Zn-containing metalloproteases and includes most enzymes from animals and only one enzyme (flavastacin) from a human-pathogenic bacterium (Flavobacterium meningosepticum). Here, we characterized the novel M12 protease myroilysin with elastinolytic activity and collagen-swelling ability from the newly described deep-sea bacterium Myroides profundi D25. Myroilysin is a monomer enzyme with 205 amino acid residues and a molecular mass of 22,936 Da. It has the same conserved residues at the four zinc ligands as astacin and very low levels of identity (activity than the bacterial elastinase pseudolysin. To our knowledge, it is the first reported elastase in the M12 family. Although it displayed very low activity with collagen, myroilysin had strong collagen-swelling ability and played a synergistic role with collagenase in collagen hydrolysis. It can be speculated that myroilysin synergistically interacts with other enzymes in its in situ biotic assemblage and that it may play an important role in the degradation of deep-sea HMW organic nitrogen.

  16. Plasma clot-promoting effect of collagen in relation to collagen-platelet interaction

    SciTech Connect

    Gentry, P.A.; Schneider, M.D.; Miller, J.K.

    1981-01-01

    The hemostatic function of several acid-soluble collagen preparations and a fibrillar-form collagen preparation have been compared. Pepsin-treated acid-soluble collagen isolated from burro and horse aortic tissue and acid-soluble colagen isolated from human umbilical cord readily promoted platelet aggregation, but failed to activate the coagulation mechanism even after prolonged incubation with plasma at 37 C. By contrast, fibrillar-form collagen isolated from burro aorta was both an efficient stimulant for the induction of platelet aggregation and a potent clot-promoting agent. Similar results were found for all the collagen preparations irrespective of whether the studies were conducted with sheep or with burro plasma. Heat denaturation studies showed that the hemostatic functon of the fibrillar-form colagen was dependent on an intact tirple-helical structure.

  17. The fibrillar collagen family.

    PubMed

    Exposito, Jean-Yves; Valcourt, Ulrich; Cluzel, Caroline; Lethias, Claire

    2010-01-01

    Collagens, or more precisely collagen-based extracellular matrices, are often considered as a metazoan hallmark. Among the collagens, fibrillar collagens are present from sponges to humans, and are involved in the formation of the well-known striated fibrils. In this review we discuss the different steps in the evolution of this protein family, from the formation of an ancestral fibrillar collagen gene to the formation of different clades. Genomic data from the choanoflagellate (sister group of Metazoa) Monosiga brevicollis, and from diploblast animals, have suggested that the formation of an ancestral alpha chain occurred before the metazoan radiation. Phylogenetic studies have suggested an early emergence of the three clades that were first described in mammals. Hence the duplication events leading to the formation of the A, B and C clades occurred before the eumetazoan radiation. Another important event has been the two rounds of "whole genome duplication" leading to the amplification of fibrillar collagen gene numbers, and the importance of this diversification in developmental processes. We will also discuss some other aspects of fibrillar collagen evolution such as the development of the molecular mechanisms involved in the formation of procollagen molecules and of striated fibrils. PMID:20386646

  18. Collagen matrix as a tool in studying fibroblastic cell behavior.

    PubMed

    Kanta, Jiří

    2015-01-01

    Type I collagen is a fibrillar protein, a member of a large family of collagen proteins. It is present in most body tissues, usually in combination with other collagens and other components of extracellular matrix. Its synthesis is increased in various pathological situations, in healing wounds, in fibrotic tissues and in many tumors. After extraction from collagen-rich tissues it is widely used in studies of cell behavior, especially those of fibroblasts and myofibroblasts. Cells cultured in a classical way, on planar plastic dishes, lack the third dimension that is characteristic of body tissues. Collagen I forms gel at neutral pH and may become a basis of a 3D matrix that better mimics conditions in tissue than plastic dishes.

  19. Collagen matrix as a tool in studying fibroblastic cell behavior.

    PubMed

    Kanta, Jiří

    2015-01-01

    Type I collagen is a fibrillar protein, a member of a large family of collagen proteins. It is present in most body tissues, usually in combination with other collagens and other components of extracellular matrix. Its synthesis is increased in various pathological situations, in healing wounds, in fibrotic tissues and in many tumors. After extraction from collagen-rich tissues it is widely used in studies of cell behavior, especially those of fibroblasts and myofibroblasts. Cells cultured in a classical way, on planar plastic dishes, lack the third dimension that is characteristic of body tissues. Collagen I forms gel at neutral pH and may become a basis of a 3D matrix that better mimics conditions in tissue than plastic dishes. PMID:25734486

  20. Synthesis and antiangiogenic activity of new silybin galloyl esters.

    PubMed

    Gažák, Radek; Valentová, Kateřina; Fuksová, Kateřina; Marhol, Petr; Kuzma, Marek; Medina, Miguel Ángel; Oborná, Ivana; Ulrichová, Jitka; Křen, Vladimír

    2011-10-27

    The synthesis of various silybin monogalloyl esters was developed, and their antiangiogenic activities were evaluated in a variety of in vitro tests with human umbilical vein endothelial cells (HUVECs). A structure-activity relationship (SAR) study found the regioselectivity of the silybin galloylation to be highly significant. Silybin (as an equimolar mixture of two diastereomers A and B) exhibited quite poor antiangiogenic activities, whereas its B stereoisomer is more active than silybin A. The galloylation of phenolic OH groups of natural silybin (a mixture of both isomers) leads to increases in their antiangiogenic activities, which is more apparent with the 7-OH than the 20-OH. In contrast, gallates at aliphatic OH groups either had a comparable activity to the parent compound or are even worse than silybin, which was observed in the case of 3-O-galloylsilybin. The most effective compound from this series (7-O-galloylsilybin) has also been prepared from stereochemically pure silybins A and B to evaluate the effect of stereochemistry on the activity. As with silybin itself, the B isomer of 7-O-galloylsilybin was more active than the A isomer.

  1. A collagen-binding EGFR antibody fragment targeting tumors with a collagen-rich extracellular matrix

    PubMed Central

    Liang, Hui; Li, Xiaoran; Wang, Bin; Chen, Bing; Zhao, Yannan; Sun, Jie; Zhuang, Yan; Shi, Jiajia; Shen, He; Zhang, Zhijun; Dai, Jianwu

    2016-01-01

    Many tumors over-express collagen, which constitutes the physical scaffold of tumor microenvironment. Collagen has been considered to be a target for cancer therapy. The collagen-binding domain (CBD) is a short peptide, which could bind to collagen and achieve the sustained release of CBD-fused proteins in collagen scaffold. Here, a collagen-binding EGFR antibody fragment was designed and expressed for targeting the collagen-rich extracellular matrix in tumors. The antibody fragment (Fab) of cetuximab was fused with CBD (CBD-Fab) and expressed in Pichia pastoris. CBD-Fab maintained antigen binding and anti-tumor activity of cetuximab and obtained a collagen-binding ability in vitro. The results also showed CBD-Fab was mainly enriched in tumors and had longer retention time in tumors in A431 s.c. xenografts. Furthermore, CBD-Fab showed a similar therapeutic efficacy as cetuximab in A431 xenografts. Although CBD-Fab hasn’t showed better therapeutic effects than cetuximab, its smaller molecular and special target may be applicable as antibody–drug conjugates (ADC) or immunotoxins. PMID:26883295

  2. Synthesis and antiproliferative activity of new tonantzitlolone-derived diterpene derivatives.

    PubMed

    Busch, Torsten; Dräger, Gerald; Kunst, Eike; Benson, Hannah; Sasse, Florenz; Siems, Karsten; Kirschning, Andreas

    2016-10-14

    The synthesis of the diterpene (+)-tonantzitlolone A and a series of derivatives is reported. The study includes the determination of their antiproliferative activities against selected cancer cell lines.

  3. Co-Culture of Tumor Spheroids and Fibroblasts in a Collagen Matrix-Incorporated Microfluidic Chip Mimics Reciprocal Activation in Solid Tumor Microenvironment

    PubMed Central

    Jeong, Su-Yeong; Lee, Ji-Hyun; Shin, Yoojin; Chung, Seok; Kuh, Hyo-Jeong

    2016-01-01

    Multicellular 3D culture and interaction with stromal components are considered essential elements in establishing a ‘more clinically relevant’ tumor model. Matrix-embedded 3D cultures using a microfluidic chip platform can recapitulate the microscale interaction within tumor microenvironments. As a major component of tumor microenvironment, cancer-associated fibroblasts (CAFs) play a role in cancer progression and drug resistance. Here, we present a microfluidic chip-based tumor tissue culture model that integrates 3D tumor spheroids (TSs) with CAF in proximity within a hydrogel scaffold. HT-29 human colorectal carcinoma cells grew into 3D TSs and the growth was stimulated when co-cultured with fibroblasts as shown by 1.5-folds increase of % changes in diameter over 5 days. TS cultured for 6 days showed a reduced expression of Ki-67 along with increased expression of fibronectin when co-cultured with fibroblasts compared to mono-cultured TSs. Fibroblasts were activated under co-culture conditions, as demonstrated by increases in α-SMA expression and migratory activity. When exposed to paclitaxel, a survival advantage was observed in TSs co-cultured with activated fibroblasts. Overall, we demonstrated the reciprocal interaction between TSs and fibroblasts in our 7-channel microfluidic chip. The co-culture of 3D TS-CAF in a collagen matrix-incorporated microfluidic chip may be useful to study the tumor microenvironment and for evaluation of drug screening and evaluation. PMID:27391808

  4. Ciprofloxacin-Collagen Conjugate in the Wound Healing Treatment

    PubMed Central

    Puoci, Francesco; Piangiolino, Cristiana; Givigliano, Francesco; Parisi, Ortensia Ilaria; Cassano, Roberta; Trombino, Sonia; Curcio, Manuela; Iemma, Francesca; Cirillo, Giuseppe; Spizzirri, Umile Gianfranco; Restuccia, Donatella; Muzzalupo, Rita; Picci, Nevio

    2012-01-01

    The synthesis of a novel functional biomaterial for wound healing treatment was carried out by adopting a free-radical grafting procedure in aqueous media. With this aim, ciprofloxacin (CFX) was covalently incorporated into collagen (T1C) chains employing an ascorbic acid/hydrogen peroxide redox pair as biocompatible initiator system. The covalent insertion of CFX in the polymeric chains was confirmed by FT-IR and UV analyses, while an antibacterial assay demonstrated the activity of the synthesized conjugate against Staphylococcus aureus and Escherichia coli, microorganisms that commonly infect wounds. A catechin blended conjugate was also tested in order to evaluate the ability to influence fibroblast cell growth. The observed antibacterial activity and stimulation of fibroblast growth support the applicability of CFX-T1C conjugate in wound treatment encouraging the healing process. PMID:24955537

  5. Immunostimulation effect of jellyfish collagen.

    PubMed

    Sugahara, Takuya; Ueno, Masashi; Goto, Yoko; Shiraishi, Ryusuke; Doi, Mikiharu; Akiyama, Koichi; Yamauchi, Satoshi

    2006-09-01

    Certain edible large jellyfishes belonging to the order Rhizostomeae are consumed in large quantities in China and Japan. The exumbrella part of the edible jellyfish Stomolophus nomurai was cut and soaked in dilute hydrochloric acid solution (pH 3.0) for 12 h, and heated at 121 degrees C for 20 min. The immunostimulation effects of the jellyfish extract were examined. The jellyfish extract enhanced IgM production of human hybridoma HB4C5 cells 34-fold. IgM and IgG production of human peripheral blood lymphocytes (PBL) were also accelerated, 2.8- and 1.4-fold respectively. Moreover, production of interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha by human PBL was stimulated 100- and 17-fold respectively. Collagenase treatment inactivated the immunostimulation activity of the jellyfish extract. In addition, purified collagen from bovine Achilles' tendon accelerated IgM production of hybridoma cells. These facts mean that collagen has an immunostimulation effect, and that the active substance in jellyfish extract is collagen.

  6. Immunostimulation effect of jellyfish collagen.

    PubMed

    Sugahara, Takuya; Ueno, Masashi; Goto, Yoko; Shiraishi, Ryusuke; Doi, Mikiharu; Akiyama, Koichi; Yamauchi, Satoshi

    2006-09-01

    Certain edible large jellyfishes belonging to the order Rhizostomeae are consumed in large quantities in China and Japan. The exumbrella part of the edible jellyfish Stomolophus nomurai was cut and soaked in dilute hydrochloric acid solution (pH 3.0) for 12 h, and heated at 121 degrees C for 20 min. The immunostimulation effects of the jellyfish extract were examined. The jellyfish extract enhanced IgM production of human hybridoma HB4C5 cells 34-fold. IgM and IgG production of human peripheral blood lymphocytes (PBL) were also accelerated, 2.8- and 1.4-fold respectively. Moreover, production of interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha by human PBL was stimulated 100- and 17-fold respectively. Collagenase treatment inactivated the immunostimulation activity of the jellyfish extract. In addition, purified collagen from bovine Achilles' tendon accelerated IgM production of hybridoma cells. These facts mean that collagen has an immunostimulation effect, and that the active substance in jellyfish extract is collagen. PMID:16960386

  7. Synthesis, characteristics and antimicrobial activity of ZnO nanoparticles.

    PubMed

    Janaki, A Chinnammal; Sailatha, E; Gunasekaran, S

    2015-06-01

    The utilization of various plant resources for the bio synthesis of metallic nano particles is called green technology and it does not utilize any harmful protocols. Present study focuses on the green synthesis of ZnO nano particles by Zinc Carbonate and utilizing the bio-components of powder extract of dry ginger rhizome (Zingiber officinale). The ZnO nano crystallites of average size range of 23-26 nm have been synthesized by rapid, simple and eco friendly method. Zinc oxide nano particles were characterized by using X-ray diffraction (XRD), Scanning Electron Microscope (SEM), Energy Dispersive X-ray spectroscopy (EDX). FTIR spectra confirmed the adsorption of surfactant molecules at the surface of ZnO nanoparticles and the presence of ZnO bonding. Antimicrobial activity of ZnO nano particles was done by well diffusion method against pathogenic organisms like Klebsiella pneumonia, Staphylococcus aureus and Candida albicans and Penicillium notatum. It is observed that the ZnO synthesized in the process has the efficient antimicrobial activity. PMID:25748589

  8. Synthesis, characteristics and antimicrobial activity of ZnO nanoparticles

    NASA Astrophysics Data System (ADS)

    Janaki, A. Chinnammal; Sailatha, E.; Gunasekaran, S.

    2015-06-01

    The utilization of various plant resources for the bio synthesis of metallic nano particles is called green technology and it does not utilize any harmful protocols. Present study focuses on the green synthesis of ZnO nano particles by Zinc Carbonate and utilizing the bio-components of powder extract of dry ginger rhizome (Zingiber officinale). The ZnO nano crystallites of average size range of 23-26 nm have been synthesized by rapid, simple and eco friendly method. Zinc oxide nano particles were characterized by using X-ray diffraction (XRD), Scanning Electron Microscope (SEM), Energy Dispersive X-ray spectroscopy (EDX). FTIR spectra confirmed the adsorption of surfactant molecules at the surface of ZnO nanoparticles and the presence of ZnO bonding. Antimicrobial activity of ZnO nano particles was done by well diffusion method against pathogenic organisms like Klebsiella pneumonia, Staphylococcus aureus and Candida albicans and Penicillium notatum. It is observed that the ZnO synthesized in the process has the efficient antimicrobial activity.

  9. d-alpha-tocopherol inhibits collagen alpha 1(I) gene expression in cultured human fibroblasts. Modulation of constitutive collagen gene expression by lipid peroxidation.

    PubMed Central

    Houglum, K; Brenner, D A; Chojkier, M

    1991-01-01

    Ascorbic acid stimulates collagen gene transcription in cultured fibroblasts, and this effect is mediated through the induction of lipid peroxidation by ascorbic acid. Quiescent cultured fibroblasts in the absence of ascorbic acid have a high constitutive level of collagen production, but the mechanisms of collagen gene regulation in this unstimulated state are not known. Because lipid peroxidation also occurs in normal cells, we wondered if lipid peroxidation plays a role in the regulation of basal collagen gene expression. Inhibition of lipid peroxidation in cultured human fibroblasts with d-alpha-tocopherol or methylene blue decreased the synthesis of collagen, the steady-state levels of procollagen alpha 1(I) mRNA and the transcription of the procollagen alpha 1(I) gene. This effect on collagen gene expression was selective and not associated with cellular toxicity. Thus, these experiments suggest a role for lipid peroxidation in the modulation of constitutive collagen gene expression. Images PMID:2040703

  10. Molecular Crowding of Collagen: A Pathway to Produce Highly-Organized Collagenous Structures

    PubMed Central

    Saeidi, Nima; Karmelek, Kathryn N.; Paten, Jeffrey. A; Zareian, Ramin; DiMasi, Elaine

    2013-01-01

    Collagen in vertebrate animals is often arranged in alternating lamellae or in bundles of aligned fibrils which are designed to withstand in vivo mechanical loads. The formation of these organized structures is thought to result from a complex, large-area integration of individual cell motion and locally-controlled synthesis of fibrillar arrays via cell-surface fibripositors (direct matrix printing). The difficulty of reproducing such a process in vitro has prevented tissue engineers from constructing clinically useful load-bearing connective tissue directly from collagen. However, we and others have taken the view that long-range organizational information is potentially encoded into the structure of the collagen molecule itself, allowing the control of fibril organization to extend far from cell (or bounding) surfaces. We here demonstrate a simple, fast, cell-free method capable of producing highly-organized, anistropic collagen fibrillar lamellae de novo which persist over relatively long-distances (tens to hundreds of microns). Our approach to nanoscale organizational control takes advantage of the intrinsic physiochemical properties of collagen molecules by inducing collagen association through molecular crowding and geometric confinement. To mimic biological tissues which comprise planar, aligned collagen lamellae (e.g. cornea, lamellar bone or annulus fibrosus), type I collagen was confined to a thin, planar geometry, concentrated through molecular crowding and polymerized. The resulting fibrillar lamellae show a striking resemblance to native load-bearing lamellae in that the fibrils are small, generally aligned in the plane of the confining space and change direction en masse throughout the thickness of the construct. The process of organizational control is consistent with embryonic development where the bounded planar cell sheets produced by fibroblasts suggest a similar confinement/concentration strategy. Such a simple approach to nanoscale

  11. Molecular crowding of collagen: a pathway to produce highly-organized collagenous structures.

    PubMed

    Saeidi, Nima; Karmelek, Kathryn P; Paten, Jeffrey A; Zareian, Ramin; DiMasi, Elaine; Ruberti, Jeffrey W

    2012-10-01

    Collagen in vertebrate animals is often arranged in alternating lamellae or in bundles of aligned fibrils which are designed to withstand in vivo mechanical loads. The formation of these organized structures is thought to result from a complex, large-area integration of individual cell motion and locally-controlled synthesis of fibrillar arrays via cell-surface fibripositors (direct matrix printing). The difficulty of reproducing such a process in vitro has prevented tissue engineers from constructing clinically useful load-bearing connective tissue directly from collagen. However, we and others have taken the view that long-range organizational information is potentially encoded into the structure of the collagen molecule itself, allowing the control of fibril organization to extend far from cell (or bounding) surfaces. We here demonstrate a simple, fast, cell-free method capable of producing highly-organized, anistropic collagen fibrillar lamellae de novo which persist over relatively long-distances (tens to hundreds of microns). Our approach to nanoscale organizational control takes advantage of the intrinsic physiochemical properties of collagen molecules by inducing collagen association through molecular crowding and geometric confinement. To mimic biological tissues which comprise planar, aligned collagen lamellae (e.g. cornea, lamellar bone or annulus fibrosus), type I collagen was confined to a thin, planar geometry, concentrated through molecular crowding and polymerized. The resulting fibrillar lamellae show a striking resemblance to native load-bearing lamellae in that the fibrils are small, generally aligned in the plane of the confining space and change direction en masse throughout the thickness of the construct. The process of organizational control is consistent with embryonic development where the bounded planar cell sheets produced by fibroblasts suggest a similar confinement/concentration strategy. Such a simple approach to nanoscale

  12. Synthesis of nano-sized amorphous boron powders through active dilution self-propagating high-temperature synthesis method

    SciTech Connect

    Wang, Jilin; Gu, Yunle; Li, Zili; Wang, Weimin; Fu, Zhengyi

    2013-06-01

    Graphical abstract: Nano-sized amorphous boron powders were synthesized by active dilution self-propagating high-temperature synthesis (SHS) method. The effects of endothermic reaction rate, the possible chemical reaction mechanism and active dilution model for synthesis of the product were also discussed. Highlights: ► Nano-sized amorphous boron powders were synthesized by active dilution self-propagating high-temperature synthesis method. ► The morphology, particle size and purity of the samples could be effectively controlled via changing the endothermic rate. ► The diluter KBH{sub 4} played an important role in active dilution synthesis of amorphous nano-sized boron powders. ► The active dilution method could be further popularized and become a common approach to prepare various inorganic materials. - Abstract: Nano-sized amorphous boron powders were synthesized by active dilution self-propagating high-temperature synthesis (SHS) method at temperatures ranging from 700 °C to 850 °C in a SHS furnace using Mg, B{sub 2}O{sub 3} and KBH{sub 4} as raw materials. Samples were characterized by X-ray powder diffraction (XRD), Laser particle size analyzer, Fourier transform infrared spectra (FTIR), X-ray energy dispersive spectroscopy (EDX), scanning electron microscopy (SEM), transmission electron microscopy (TEM) and high-resolution transmission TEM (HRTEM). The boron powders demonstrated an average particle size of 50 nm with a purity of 95.64 wt.%. The diluter KBH{sub 4} played an important role in the active dilution synthesis of amorphous nano-sized boron powders. The effects of endothermic reaction rate, the possible chemical reaction mechanism and active dilution model for synthesis of the product were also discussed.

  13. Design, synthesis and structure-activity relationship of phthalimides endowed with dual antiproliferative and immunomodulatory activities.

    PubMed

    Cardoso, Marcos Veríssimo de Oliveira; Moreira, Diogo Rodrigo Magalhães; Oliveira Filho, Gevanio Bezerra; Cavalcanti, Suellen Melo Tiburcio; Coelho, Lucas Cunha Duarte; Espíndola, José Wanderlan Pontes; Gonzalez, Laura Rubio; Rabello, Marcelo Montenegro; Hernandes, Marcelo Zaldini; Ferreira, Paulo Michel Pinheiro; Pessoa, Cláudia; Alberto de Simone, Carlos; Guimarães, Elisalva Teixeira; Soares, Milena Botelho Pereira; Leite, Ana Cristina Lima

    2015-01-01

    The present work reports the synthesis and evaluation of the antitumour and immunomodulatory properties of new phthalimides derivatives designed to explore molecular hybridization and bioisosterism approaches between thalidomide, thiosemicarbazone, thiazolidinone and thiazole series. Twenty-seven new molecules were assessed for their immunosuppressive effect toward TNFα, IFNγ, IL-2 and IL-6 production and antiproliferative activity. The best activity profile was observed for the (6a-f) series, which presents phthalyl and thiazolidinone groups.

  14. Synthesis and biological activity of Wuweizisu C and analogs.

    PubMed

    Chang, J B; Wang, Q; Li, Y F

    2009-01-01

    Lignans are widely distributed in nature. The earliest recorded medicinal use of lignans dated back to over 1000 years ago. Lignan-rich plant products were also active ingredients in Chinese and Japanese folk medicines for the treatment of various diseases. The dried root and stem of this plant are listed in the Chinese pharmacopoeia for the treatment of rheumatoid arthritis, gastric, duodenal ulcers and many other diseases. This review highlights synthetic strategies for the Wuweizisu C analogs and the important pharmacological activities as well as therapeutic findings related to the treatment of HBV and other diseases. Notably a significant and ongoing project on Wuweizisu C and its analogs has led to the discovery and development of two potent derivatives alpha-DDB and BICYCLOL which are currently in clinical trials against HBV, especially in lowering elevated SGPT levels. Further design, synthesis, and evaluation of Wuweizisu C analogs are discussed.

  15. Patenting activity in synthesis of lipid nanotubes and peptide nanotubes.

    PubMed

    Zhou, Yong

    2007-01-01

    Lipid nanotubes (LNTs) and peptide nanotubes (PNTs) are especially intriguing and noncovalent self-assemblies of amphiphiles. They have hydrophilically internal and external membrane surfaces, and can provide the wide scope for chemical modifications, in sharp contrast to carbon nanotubes. These unique properties make themselves as ideal candidates for a variety of applications in chemistry, biochemistry, materials science and medicine. Patenting the LNTs and PNTs is quite active recently. This mini-review provides a brief outline of patenting activity in synthesis of the LNTs and PNTs since 1980s. The key point of the present review aims to create an optimistic circulation between the basic research achievement and potential application of this sub-field of nanotechnology, promoting each other in their future development.

  16. Glycosphingolipid synthesis inhibition limits osteoclast activation and myeloma bone disease

    PubMed Central

    Ersek, Adel; Xu, Ke; Antonopoulos, Aristotelis; Butters, Terry D.; Santo, Ana Espirito; Vattakuzhi, Youridies; Williams, Lynn M.; Goudevenou, Katerina; Danks, Lynett; Freidin, Andrew; Spanoudakis, Emmanouil; Parry, Simon; Papaioannou, Maria; Hatjiharissi, Evdoxia; Chaidos, Aristeidis; Alonzi, Dominic S.; Twigg, Gabriele; Hu, Ming; Dwek, Raymond A.; Haslam, Stuart M.; Roberts, Irene; Dell, Anne; Rahemtulla, Amin; Horwood, Nicole J.; Karadimitris, Anastasios

    2015-01-01

    Glycosphingolipids (GSLs) are essential constituents of cell membranes and lipid rafts and can modulate signal transduction events. The contribution of GSLs in osteoclast (OC) activation and osteolytic bone diseases in malignancies such as the plasma cell dyscrasia multiple myeloma (MM) is not known. Here, we tested the hypothesis that pathological activation of OCs in MM requires de novo GSL synthesis and is further enhanced by myeloma cell–derived GSLs. Glucosylceramide synthase (GCS) inhibitors, including the clinically approved agent N-butyl-deoxynojirimycin (NB-DNJ), prevented OC development and activation by disrupting RANKL-induced localization of TRAF6 and c-SRC into lipid rafts and preventing nuclear accumulation of transcriptional activator NFATc1. GM3 was the prevailing GSL produced by patient-derived myeloma cells and MM cell lines, and exogenous addition of GM3 synergistically enhanced the ability of the pro-osteoclastogenic factors RANKL and insulin-like growth factor 1 (IGF-1) to induce osteoclastogenesis in precursors. In WT mice, administration of GM3 increased OC numbers and activity, an effect that was reversed by treatment with NB-DNJ. In a murine MM model, treatment with NB-DNJ markedly improved osteolytic bone disease symptoms. Together, these data demonstrate that both tumor-derived and de novo synthesized GSLs influence osteoclastogenesis and suggest that NB-DNJ may reduce pathological OC activation and bone destruction associated with MM. PMID:25915583

  17. Protein kinase D activity controls endothelial nitric oxide synthesis.

    PubMed

    Aicart-Ramos, Clara; Sánchez-Ruiloba, Lucía; Gómez-Parrizas, Mónica; Zaragoza, Carlos; Iglesias, Teresa; Rodríguez-Crespo, Ignacio

    2014-08-01

    Vascular endothelial growth factor (VEGF) regulates key functions of the endothelium, such as angiogenesis or vessel repair in processes involving endothelial nitric oxide synthase (eNOS) activation. One of the effector kinases that become activated in endothelial cells upon VEGF treatment is protein kinase D (PKD). Here, we show that PKD phosphorylates eNOS, leading to its activation and a concomitant increase in NO synthesis. Using mass spectrometry, we show that the purified active kinase specifically phosphorylates recombinant eNOS on Ser1179. Treatment of endothelial cells with VEGF or phorbol 12,13-dibutyrate (PDBu) activates PKD and increases eNOS Ser1179 phosphorylation. In addition, pharmacological inhibition of PKD and gene silencing of both PKD1 and PKD2 abrogate VEGF signaling, resulting in a clear diminished migration of endothelial cells in a wound healing assay. Finally, inhibition of PKD in mice results in an almost complete disappearance of the VEGF-induced vasodilatation, as monitored through determination of the diameter of the carotid artery. Hence, our data indicate that PKD is a new regulatory kinase of eNOS in endothelial cells whose activity orchestrates mammalian vascular tone. PMID:24928905

  18. The adaptor molecule signaling lymphocytic activation molecule (SLAM)-associated protein (SAP) is essential in mechanisms involving the Fyn tyrosine kinase for induction and progression of collagen-induced arthritis.

    PubMed

    Zhong, Ming-Chao; Veillette, André

    2013-11-01

    Signaling lymphocytic activation molecule-associated protein (SAP) is an Src homology 2 domain-only adaptor involved in multiple immune cell functions. It has also been linked to immunodeficiencies and autoimmune diseases, such as systemic lupus erythematosus. Here, we examined the role and mechanism of action of SAP in autoimmunity using a mouse model of autoimmune arthritis, collagen-induced arthritis (CIA). We found that SAP was essential for development of CIA in response to collagen immunization. It was also required for production of collagen-specific antibodies, which play a key role in disease pathogenesis. These effects required SAP expression in T cells, not in B cells. In mice immunized with a high dose of collagen, the activity of SAP was nearly independent of its ability to bind the protein tyrosine kinase Fyn and correlated with the capacity of SAP to promote full differentiation of follicular T helper (TFH) cells. However, with a lower dose of collagen, the role of SAP was more dependent on Fyn binding, suggesting that additional mechanisms other than TFH cell differentiation were involved. Further studies suggested that this might be due to a role of the SAP-Fyn interaction in natural killer T cell development through the ability of SAP-Fyn to promote Vav-1 activation. We also found that removal of SAP expression during progression of CIA attenuated disease severity. However, it had no effect on disease when CIA was clinically established. Together, these results indicate that SAP plays an essential role in CIA because of Fyn-independent and Fyn-dependent effects on TFH cells and, possibly, other T cell types.

  19. Complications of collagen fillers.

    PubMed

    Lucey, Patricia; Goldberg, David J

    2014-12-01

    As the skin ages, a deficiency in collagen occurs, thus injectable collagen products have become a sensible and popular option for dermal filling and volume enhancement. Several types of collagen have been developed over the years, including animal sources such as bovine and porcine collagen, as well as human-based sources derived from pieces of the patient's own skin, cadaver skin, and later cultured from human dermal fibroblasts. While collagen overall has a relatively safe, side effect profile, there are several complications, both early and late onset, that practitioners and patients should be aware of. Early complications, occurring within days of the procedure, can be divided into non-hypersensitivity and hypersensitivity reactions. The non-hypersensitive reactions include injection site reactions, discoloration, maldistribution, infection, skin necrosis, and the very rare but dreaded risk of vision loss, whereas the hypersensitivity reactions present usually as delayed type IV reactions, but can also rarely present as an immediate type I reaction. Late complications, occurring within weeks to even years after injection, include granuloma formation, foreign body reactions, and infection secondary to atypical mycobacteria or biofilms. This review will give a detailed overview of the complications secondary to cutaneous collagen injections.

  20. Nanomechanics of collagen microfibrils

    PubMed Central

    Vesentini, Simone; Redaelli, Alberto; Gautieri, Alfonso

    2013-01-01

    Summary Collagen constitutes one third of the human proteome, providing mechanical stability, elasticity and strength to organisms and is thus the prime construction material in biology. Collagen is also the dominating material in the extracellular matrix where its stiffness controls cell differentiation, growth and pathology. We use atomistic-based hierarchical multiscale modeling to describe this complex biological material from the bottom up. This includes the use and development of large-scale computational modeling tools to investigate several aspects related to collagen-based tissues, including source of visco-elasticity and deformation mechanisms at the nanoscale level. The key innovation of this research is that until now, collagen materials have primarily been described at macroscopic scales, without explicitly understanding the mechanical contributions at the molecular and fibrillar levels. The major impact of this research will be the development of fundamental models of collagenous tissues, important to the design of new scaffolding biomaterials for regenerative medicine as well as for the understanding of collagen-related diseases. PMID:23885342

  1. Rheb Inhibits Protein Synthesis by Activating the PERK-eIF2α Signaling Cascade

    PubMed Central

    Tyagi, Richa; Shahani, Neelam; Gorgen, Lindsay; Ferretti, Max; Pryor, William; Chen, Po Yu; Swarnkar, Supriya; Worley, Paul F.; Karbstein, Katrin; Snyder, Solomon H.; Subramaniam, Srinivasa

    2015-01-01

    SUMMARY Rheb, a ubiquitous small GTPase, is well known to bind and activate mTOR, which augments protein synthesis. Inhibition of protein synthesis is also physiologically regulated. Thus, with cell stress the unfolded protein response system leads to phosphorylation of the initiation factor eIF2α and arrest of protein synthesis. We now demonstrate a major role for Rheb in inhibiting protein synthesis through enhancing the phosphorylation of eIF2α by protein kinase-like endoplasmic reticulum kinase (PERK). Interplay between the stimulatory and inhibitory roles of Rheb may enable cells to modulate protein synthesis in response to varying environmental stresses. PMID:25660019

  2. Recombinant Collagen Engineered to Bind to Discoidin Domain Receptor Functions as a Receptor Inhibitor*

    PubMed Central

    An, Bo; Abbonante, Vittorio; Xu, Huifang; Gavriilidou, Despoina; Yoshizumi, Ayumi; Bihan, Dominique; Farndale, Richard W.; Kaplan, David L.; Balduini, Alessandra; Leitinger, Birgit; Brodsky, Barbara

    2016-01-01

    A bacterial collagen-like protein Scl2 has been developed as a recombinant collagen model system to host human collagen ligand-binding sequences, with the goal of generating biomaterials with selective collagen bioactivities. Defined binding sites in human collagen for integrins, fibronectin, heparin, and MMP-1 have been introduced into the triple-helical domain of the bacterial collagen and led to the expected biological activities. The modular insertion of activities is extended here to the discoidin domain receptors (DDRs), which are collagen-activated receptor tyrosine kinases. Insertion of the DDR-binding sequence from human collagen III into bacterial collagen led to specific receptor binding. However, even at the highest testable concentrations, the construct was unable to stimulate DDR autophosphorylation. The recombinant collagen expressed in Escherichia coli does not contain hydroxyproline (Hyp), and complementary synthetic peptide studies showed that replacement of Hyp by Pro at the critical Gly-Val-Met-Gly-Phe-Hyp position decreased the DDR-binding affinity and consequently required a higher concentration for the induction of receptor activation. The ability of the recombinant bacterial collagen to bind the DDRs without inducing kinase activation suggested it could interfere with the interactions between animal collagen and the DDRs, and such an inhibitory role was confirmed in vitro and with a cell migration assay. This study illustrates that recombinant collagen can complement synthetic peptides in investigating structure-activity relationships, and this system has the potential for the introduction or inhibition of specific biological activities. PMID:26702058

  3. Recombinant Collagen Engineered to Bind to Discoidin Domain Receptor Functions as a Receptor Inhibitor.

    PubMed

    An, Bo; Abbonante, Vittorio; Xu, Huifang; Gavriilidou, Despoina; Yoshizumi, Ayumi; Bihan, Dominique; Farndale, Richard W; Kaplan, David L; Balduini, Alessandra; Leitinger, Birgit; Brodsky, Barbara

    2016-02-26

    A bacterial collagen-like protein Scl2 has been developed as a recombinant collagen model system to host human collagen ligand-binding sequences, with the goal of generating biomaterials with selective collagen bioactivities. Defined binding sites in human collagen for integrins, fibronectin, heparin, and MMP-1 have been introduced into the triple-helical domain of the bacterial collagen and led to the expected biological activities. The modular insertion of activities is extended here to the discoidin domain receptors (DDRs), which are collagen-activated receptor tyrosine kinases. Insertion of the DDR-binding sequence from human collagen III into bacterial collagen led to specific receptor binding. However, even at the highest testable concentrations, the construct was unable to stimulate DDR autophosphorylation. The recombinant collagen expressed in Escherichia coli does not contain hydroxyproline (Hyp), and complementary synthetic peptide studies showed that replacement of Hyp by Pro at the critical Gly-Val-Met-Gly-Phe-Hyp position decreased the DDR-binding affinity and consequently required a higher concentration for the induction of receptor activation. The ability of the recombinant bacterial collagen to bind the DDRs without inducing kinase activation suggested it could interfere with the interactions between animal collagen and the DDRs, and such an inhibitory role was confirmed in vitro and with a cell migration assay. This study illustrates that recombinant collagen can complement synthetic peptides in investigating structure-activity relationships, and this system has the potential for the introduction or inhibition of specific biological activities.

  4. Synthesis, structure characterization and catalytic activity of nickel tungstate nanoparticles

    NASA Astrophysics Data System (ADS)

    Pourmortazavi, Seied Mahdi; Rahimi-Nasrabadi, Mehdi; Khalilian-Shalamzari, Morteza; Zahedi, Mir Mahdi; Hajimirsadeghi, Seiedeh Somayyeh; Omrani, Ismail

    2012-12-01

    Taguchi robust design was applied to optimize experimental parameters for controllable, simple and fast synthesis of nickel tungstate nanoparticles. NiWO4 nanoparticles were synthesized by precipitation reaction involving addition of nickel ion solution to the tungstate aqueous reagent and then formation of nickel tungstate nucleolus which are insoluble in aqueous media. Effects of various parameters such as nickel and tungstate concentrations, flow rate of reagent addition and reactor temperature on diameter of synthesized nickel tungstate nanoparticles were investigated experimentally by the aid of orthogonal array design. The results for analysis of variance (ANOVA) showed that particle size of nickel tungstate can be effectively tuned by controlling significant variables involving nickel and tungstate concentrations and flow rate; while, temperature of the reactor has a no considerable effect on the size of NiWO4 particles. The ANOVA results proposed the optimum conditions for synthesis of nickel tungstate nanoparticles via this technique. Also, under optimum condition nanoparticles of NiWO4 were prepared and their structure and chemical composition were characterized by means of EDAX, XRD, SEM, FT-IR spectroscopy, UV-vis spectroscopy, and photoluminescence. Finally, catalytic activity of the nanoparticles in a cycloaddition reaction was examined.

  5. Replication stress activates DNA repair synthesis in mitosis.

    PubMed

    Minocherhomji, Sheroy; Ying, Songmin; Bjerregaard, Victoria A; Bursomanno, Sara; Aleliunaite, Aiste; Wu, Wei; Mankouri, Hocine W; Shen, Huahao; Liu, Ying; Hickson, Ian D

    2015-12-10

    Oncogene-induced DNA replication stress has been implicated as a driver of tumorigenesis. Many chromosomal rearrangements characteristic of human cancers originate from specific regions of the genome called common fragile sites (CFSs). CFSs are difficult-to-replicate loci that manifest as gaps or breaks on metaphase chromosomes (termed CFS 'expression'), particularly when cells have been exposed to replicative stress. The MUS81-EME1 structure-specific endonuclease promotes the appearance of chromosome gaps or breaks at CFSs following replicative stress. Here we show that entry of cells into mitotic prophase triggers the recruitment of MUS81 to CFSs. The nuclease activity of MUS81 then promotes POLD3-dependent DNA synthesis at CFSs, which serves to minimize chromosome mis-segregation and non-disjunction. We propose that the attempted condensation of incompletely duplicated loci in early mitosis serves as the trigger for completion of DNA replication at CFS loci in human cells. Given that this POLD3-dependent mitotic DNA synthesis is enhanced in aneuploid cancer cells that exhibit intrinsically high levels of chromosomal instability (CIN(+)) and replicative stress, we suggest that targeting this pathway could represent a new therapeutic approach.

  6. Replication stress activates DNA repair synthesis in mitosis.

    PubMed

    Minocherhomji, Sheroy; Ying, Songmin; Bjerregaard, Victoria A; Bursomanno, Sara; Aleliunaite, Aiste; Wu, Wei; Mankouri, Hocine W; Shen, Huahao; Liu, Ying; Hickson, Ian D

    2015-12-10

    Oncogene-induced DNA replication stress has been implicated as a driver of tumorigenesis. Many chromosomal rearrangements characteristic of human cancers originate from specific regions of the genome called common fragile sites (CFSs). CFSs are difficult-to-replicate loci that manifest as gaps or breaks on metaphase chromosomes (termed CFS 'expression'), particularly when cells have been exposed to replicative stress. The MUS81-EME1 structure-specific endonuclease promotes the appearance of chromosome gaps or breaks at CFSs following replicative stress. Here we show that entry of cells into mitotic prophase triggers the recruitment of MUS81 to CFSs. The nuclease activity of MUS81 then promotes POLD3-dependent DNA synthesis at CFSs, which serves to minimize chromosome mis-segregation and non-disjunction. We propose that the attempted condensation of incompletely duplicated loci in early mitosis serves as the trigger for completion of DNA replication at CFS loci in human cells. Given that this POLD3-dependent mitotic DNA synthesis is enhanced in aneuploid cancer cells that exhibit intrinsically high levels of chromosomal instability (CIN(+)) and replicative stress, we suggest that targeting this pathway could represent a new therapeutic approach. PMID:26633632

  7. Photocatalytically active titanium dioxide nanopowders: Synthesis, photoactivity and magnetic separation

    NASA Astrophysics Data System (ADS)

    Nikkanen, J.-P.; Heinonen, S.; Huttunen Saarivirta, E.; Honkanen, M.; Levänen, E.

    2013-12-01

    Two approaches were used to obtain nanocrystalline titanium dioxide (TiO2) photocatalyst powders. Firstly, low-temperature synthesis method and secondly liquid flame spraying. The structural properties of the produced powders were determined with X-ray diffraction, transmission electron microscopy and nitrogen adsorption tests. The photocatalytic properties of the powders were studied with methylene blue (MB) discoloration tests. After discolorations tests, TiO2 was coagulated with magnetite particles using FeCl3·6 H2O at a fixed pH value. Magnetic separation of coagulated TiO2 and magnetite was carried out by a permanent magnet. The obtained results showed that the particle size of the powders synthesized at low-temperature was very small and the specific surface area high. The phase content of the powder was also shown to depend greatly on the acidity of the synthesis solution. Powder synthesized by liquid flame spraying was mixture of anatase and rutile phases with essentially larger particle size and lower specific surface area than those of low-temperature synthesized powders. The MB discoloration test showed that photocatalytic activity depends on the phase structure as well as the specific surface area of the synthesized TiO2 powder. The magnetic separation of TiO2-magnetite coagulate from solution proved to be efficient around pH:8.

  8. Type V collagen controls the initiation of collagen fibril assembly.

    PubMed

    Wenstrup, Richard J; Florer, Jane B; Brunskill, Eric W; Bell, Sheila M; Chervoneva, Inna; Birk, David E

    2004-12-17

    Vertebrate collagen fibrils are heterotypically composed of a quantitatively major and minor fibril collagen. In non-cartilaginous tissues, type I collagen accounts for the majority of the collagen mass, and collagen type V, the functions of which are poorly understood, is a minor component. Type V collagen has been implicated in the regulation of fibril diameter, and we reported recently preliminary evidence that type V collagen is required for collagen fibril nucleation (Wenstrup, R. J., Florer, J. B., Cole, W. G., Willing, M. C., and Birk, D. E. (2004) J. Cell. Biochem. 92, 113-124). The purpose of this study was to define the roles of type V collagen in the regulation of collagen fibrillogenesis and matrix assembly. Mouse embryos completely deficient in pro-alpha1(V) chains were created by homologous recombination. The col5a1-/- animals die in early embryogenesis, at approximately embryonic day 10. The type V collagen-deficient mice demonstrate a virtual lack of collagen fibril formation. In contrast, the col5a1+/- animals are viable. The reduced type V collagen content is associated with a 50% reduction in fibril number and dermal collagen content. In addition, relatively normal, cylindrical fibrils are assembled with a second population of large, structurally abnormal collagen fibrils. The structural properties of the abnormal matrix are decreased relative to the wild type control animals. These data indicate a central role for the evolutionary, ancient type V collagen in the regulation of fibrillogenesis. The complete dependence of fibril formation on type V collagen is indicative of the critical role of the latter in early fibril initiation. In addition, this fibril collagen is important in the determination of fibril structure and matrix organization. PMID:15383546

  9. Turnover Rates of Hepatic Collagen and Circulating Collagen-Associated Proteins in Humans with Chronic Liver Disease

    PubMed Central

    Li, Kelvin; Gatmaitan, Michelle; Luo, Flora; Cattin, Jerome; Nakamura, Corelle; Holmes, William E.; Angel, Thomas E.; Peters, Marion G.; Turner, Scott M.; Hellerstein, Marc K.

    2015-01-01

    Accumulation and degradation of scar tissue in fibrotic liver disease occur slowly, typically over many years. Direct measurement of fibrogenesis, the rate of scar tissue deposition, may provide valuable therapeutic and prognostic information. We describe here results from a pilot study utilizing in vivo metabolic labeling to measure the turnover rate of hepatic collagen and collagen-associated proteins in plasma for the first time in human subjects. Eight subjects with chronic liver disease were labeled with daily oral doses of 2H2O for up to 8 weeks prior to diagnostic liver biopsy and plasma collection. Tandem mass spectrometry was used to measure the abundance and fractional synthesis rate (FSR) of proteins in liver and blood. Relative protein abundance and FSR data in liver revealed marked differences among subjects. FSRs of hepatic type I and III collagen ranged from 0.2–0.6% per day (half-lives of 4 months to a year) and correlated significantly with worsening histologic fibrosis. Analysis of plasma protein turnover revealed two collagen-associated proteins, lumican and transforming growth factor beta-induced-protein (TGFBI), exhibiting FSRs that correlated significantly with FSRs of hepatic collagen. In summary, this is the first direct measurement of liver collagen turnover in vivo in humans and suggests a high rate of collagen remodeling in advanced fibrosis. In addition, the FSRs of collagen-associated proteins in plasma are measurable and may provide a novel strategy for monitoring hepatic fibrogenesis rates. PMID:25909381

  10. Synthesis, antimicrobial, and antiproliferative activities of substituted phenylfuranylnicotinamidines

    PubMed Central

    Youssef, Magdy M; Arafa, Reem K; Ismail, Mohamed A

    2016-01-01

    This research work deals with the design and synthesis of a series of substituted phenylfuranylnicotinamidines 4a–i. Facile preparation of the target compounds was achieved by Suzuki coupling-based synthesis of the nitrile precursors 3a–i, followed by their conversion to the corresponding nicotinamidines 4a–i utilizing LiN(TMS)2. The antimicrobial activities of the newly synthesized nicotinamidine derivatives were evaluated against the Gram-negative bacterial strains Escherichia coli and Pseudomonas aeruginosa as well as the Gram-positive bacterial strains Staphylococcus aureus and Bacillus megaterium. The minimum inhibitory concentration values of nicotinamidines against all tested microorganisms were in the range of 10–20 μM. In specific, compounds 4a and 4b showed excellent minimum inhibitory concentration values of 10 μM against Staphylococcus aureus bacterial strain and were similar to ampicillin as an antibacterial reference. On the other hand, selected nicotinamidine derivatives were biologically screened for their cytotoxic activities against a panel of 60 cell lines representing nine types of human cancer at a single high dose at National Cancer Institute, Bethesda, MD, USA. Nicotinamidines showing promising activities were further assessed in a five-dose screening assay to determine their compound concentration causing 50% growth inhibition of tested cell (GI50), compound concentration causing 100% growth inhibition of tested cell (TGI), and compound concentration causing 50% lethality of tested cell (LC50) values. Structure-activity relationship studies demonstrated that the activity of members of this series can be modulated from cytostatic to cytotoxic based on the substitution pattern/nature on the terminal phenyl ring. The most active compound was found to be 4e displaying a submicromolar GI50 value of 0.83 μM, with TGI and LC50 values of 2.51 and 100 μM, respectively. Finally, the possible underlying mechanism of action of this series of

  11. Dihydrobenzofuran Neolignanamides: Laccase-Mediated Biomimetic Synthesis and Antiproliferative Activity.

    PubMed

    Cardullo, Nunzio; Pulvirenti, Luana; Spatafora, Carmela; Musso, Nicolò; Barresi, Vincenza; Condorelli, Daniele Filippo; Tringali, Corrado

    2016-08-26

    The biomimetic synthesis of a small library of dihydrobenzofuran neolignanamides (the natural trans-grossamide (4) and the related compounds 21-28) has been carried out through an eco-friendly oxidative coupling reaction mediated by Trametes versicolor laccase. These products, after complete spectroscopic characterization, were evaluated for their antiproliferative activity against Caco-2 (colon carcinoma), MCF-7 (mammary adenocarcinoma), and PC-3 (prostate cancer) human cells, using an MTT bioassay. The racemic neolignamides (±)-21 and (±)-27, in being the most lipophilic in the series, were potently active, with GI50 values comparable to or even lower than that of the positive control 5-FU. The racemates were resolved through chiral HPLC, and the pure enantiomers were subjected to ECD measurements to establish their absolute configurations at C-2 and C-3. All enantiomers showed potent antiproliferative activity, with, in particular, a GI50 value of 1.1 μM obtained for (2R,3R)-21. The effect of (±)-21 on the Caco-2 cell cycle was evaluated by flow cytometry, and it was demonstrated that (±)-21 exerts its antiproliferative activity by inducing cell cycle arrest and apoptosis. PMID:27504537

  12. Synthesis and biological activity of salinomycin conjugates with floxuridine.

    PubMed

    Huczyński, Adam; Antoszczak, Michał; Kleczewska, Natalia; Lewandowska, Marta; Maj, Ewa; Stefańska, Joanna; Wietrzyk, Joanna; Janczak, Jan; Celewicz, Lech

    2015-03-26

    As part of our program to develop anticancer agents, we have synthesized new compounds, which are conjugates between well-known anticancer drug, floxuridine and salinomycin which is able to selectivity kill cancer stem cells. The conjugates were obtained in two ways i.e. by copper(I) catalysed click Huisgen cycloaddition reaction performed between 3'-azido-2',3'-dideoxy-5-fluorouridine and salinomycin propargyl amide, and by the ester synthesis starting from salinomycin and floxuridine under mild condition. The compounds obtained were characterized by spectroscopic methods and evaluated for their in vitro cytotoxicity against seven human cancer cell lines as well as antibacterial activity against clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus epidermidis (MRSE). The conjugate obtained by esterification reaction showed a significantly higher antiproliferative activity against the drug-resistant cancer cells and lower toxicity than those of salinomycin and floxuridine towards normal cells, as well as standard anticancer drugs, such as cisplatin and doxorubicin. The conjugate compound revealed also moderate activity against MRSA and MRSE bacterial strains. Very high activity of floxuridine and 5-fluorouracil against MRSA and MRSE has been also observed.

  13. Abnormal Collagen Metabolism in Cultured Skin Fibroblasts from Patients with Duchenne Muscular Dystrophy

    NASA Astrophysics Data System (ADS)

    Rodemann, H. Peter; Bayreuther, Klaus

    1984-08-01

    Total collagen synthesis is decreased by about 29% (P < 0.01) in skin fibroblasts established in vitro from male patients with Duchenne muscular dystrophy (DMD) as compared with that in normal male skin fibroblasts in vitro. The reduction in collagen synthesis is associated with an approximately 2-fold increase in collagen degradation in DMD fibroblasts. Correlated to these alterations in the metabolism of collagen, DMD fibroblasts express a significantly higher hydroxyproline/proline ratio (DMD: 1.36-1.45; P < 0.01) than do normal fibroblasts (controls: 0.86-0.89). The increased hydroxylation of proline residues of collagen (composed of type I and type III) could be the cause for the enhanced degradation of collagen in DMD fibroblasts.

  14. Synthesis and Structural Activity Relationship Study of Antitubercular Carboxamides

    PubMed Central

    Ugwu, D. I.; Ezema, B. E.; Eze, F. U.; Ugwuja, D. I.

    2014-01-01

    The unusual structure and chemical composition of the mycobacterial cell wall, the tedious duration of therapy, and resistance developed by the microorganism have made the recurrence of the disease multidrug resistance and extensive or extreme drug resistance. The prevalence of tuberculosis in synergy with HIV/AIDS epidemic augments the risk of developing the disease by 100-fold. The need to synthesize new drugs that will shorten the total duration of effective treatment and/or significantly reduce the dosage taken under DOTS supervision, improve on the treatment of multidrug-resistant tuberculosis which defies the treatment with isoniazid and rifampicin, and provide effective treatment for latent TB infections which is essential for eliminating tuberculosis prompted this review. In this review, we considered the synthesis and structure activity relationship study of carboxamide derivatives with antitubercular potential. PMID:25610646

  15. Highly Selective Synthesis of Catalytically Active Monodisperse Rhodium Nanocubes

    SciTech Connect

    Zhang, Y.; Grass, M.E.; Kuhn, J.N.; Tao, F.; Habas, S.E.; Huang, W.; Yang, P.; Somorjai, G.A.

    2009-02-21

    Synthesis of monodisperse and shape-controlled colloidal inorganic nanocrystals (NCs) is of increasing scientific interest and technological significance. Recently, shape control of Pt, Pd, Ag, Au, and Rh NCs has been obtained by tuning growth kinetics in various solution-phase approaches, including modified polyol methods, seeded growth by polyol reduction, thermolysis of organometallics, and micelle techniques. Control of reduction kinetics of the noble metal precursors and regulation of the relative growth rates of low-index planes (i.e. {l_brace}100{r_brace} and {l_brace}111{r_brace}) via selective adsorption of selected chemical species are two keys for achieving shape modification of noble metal NCs. One application for noble metal NCs of well-defined shape is in understanding how NC faceting (determines which crystallographic planes are exposed) affects catalytic performance. Rh NCs are used in many catalytic reactions, including hydrogenation, hydroformylation, hydrocarbonylation, and combustion reactions. Shape manipulation of Rh NCs may be important in understanding how faceting on the nanoscale affects catalytic properties, but such control is challenging and there are fewer reports on the shape control of Rh NCs compared to other noble metals. Xia and coworkers obtained Rh multipods exhibiting interesting surface plasmonic properties by a polyol approach. The Somorjai and Tilley groups synthesized crystalline Rh multipods, cubes, horns and cuboctahedra, via polyol seeded growth. Son and colleagues prepared catalytically active monodisperse oleylamine-capped tetrahedral Rh NCs for the hydrogenation of arenes via an organometallic route. More recently, the Somorjai group synthesized sizetunable monodisperse Rh NCs using a one-step polyol technique. In this Communication, we report the highly selective synthesis of catalytically active, monodisperse Rh nanocubes of < 10 nm by a seedless polyol method. In this approach, Br{sup -} ions from trimethyl

  16. Silver nanoparticles: green synthesis and their antimicrobial activities.

    PubMed

    Sharma, Virender K; Yngard, Ria A; Lin, Yekaterina

    2009-01-30

    This review presents an overview of silver nanoparticles (Ag NPs) preparation by green synthesis approaches that have advantages over conventional methods involving chemical agents associated with environmental toxicity. Green synthetic methods include mixed-valence polyoxometallates, polysaccharide, Tollens, irradiation, and biological. The mixed-valence polyoxometallates method was carried out in water, an environmentally-friendly solvent. Solutions of AgNO(3) containing glucose and starch in water gave starch-protected Ag NPs, which could be integrated into medical applications. Tollens process involves the reduction of Ag(NH(3))(2)(+) by saccharides forming Ag NP films with particle sizes from 50-200 nm, Ag hydrosols with particles in the order of 20-50 nm, and Ag colloid particles of different shapes. The reduction of Ag(NH(3))(2)(+) by HTAB (n-hexadecyltrimethylammonium bromide) gave Ag NPs of different morphologies: cubes, triangles, wires, and aligned wires. Ag NPs synthesis by irradiation of Ag(+) ions does not involve a reducing agent and is an appealing procedure. Eco-friendly bio-organisms in plant extracts contain proteins, which act as both reducing and capping agents forming stable and shape-controlled Ag NPs. The synthetic procedures of polymer-Ag and TiO(2)-Ag NPs are also given. Both Ag NPs and Ag NPs modified by surfactants or polymers showed high antimicrobial activity against gram-positive and gram-negative bacteria. The mechanism of the Ag NP bactericidal activity is discussed in terms of Ag NP interaction with the cell membranes of bacteria. Silver-containing filters are shown to have antibacterial properties in water and air purification. Finally, human and environmental implications of Ag NPs to the ecology of aquatic environment are briefly discussed.

  17. Transition metal bimetallic oxycarbides: Synthesis, characterization, and activity studies

    SciTech Connect

    Oyama, S.T.; Yu, C.C.; Ramanathan, S.

    1999-06-10

    A new family of bimetallic oxycarbide compounds M{sup I}-M{sup II}-O-C (M{sup I} = Mo, W; M{sup II} = V, Nb, Cr, Fe, Co, Ni) has been synthesized by carburizing bimetallic oxide precursors using a temperature-programmed method. The oxide precursors are prepared by conventional solid-state reaction between two appropriate monometallic oxides. The synthesis involves passing a 20 mol% CH{sub 4} in H{sub 2} mixture over the oxide precursors while raising the temperature at a linear rate of 8.3 {times} 10{sup {minus}2} K/s (5 K/min) to a final temperature (T{sub max}) which is held for a period of time (t{sub hold}). The synthesis, chemisorption properties, and reactivation of the materials indicate that the compounds can be divided into two groups of different reducibility (high and low). Their surface activity and surface area are evaluated based on CO chemisorption and N{sub 2} physisorption measurements. It is found that the CO number density correlates with the reducibility of the compounds. The catalysts were evaluated for hydroprocessing in a three-phase trickle-bed reactor operated at 3.1 MPa and 643 K. The feed was a model liquid mixture containing 3000 ppm sulfur (dibenzothiophene), 2000 ppm nitrogen (quinoline), 500 ppm oxygen (benzofuran), 20 wt% aromatics (tetralin), and balance aliphatics (tetradecane). The bimetallic oxycarbides had moderate activity for HDN of quinoline, with Nb-Mo-O-C showing higher HDN than a commercial sulfided Ni-Mo/Al{sub 2}O{sub 3} catalyst tested at the same conditions. X-ray diffraction of the spent catalysts indicated that the oxycarbides of the early transition metals were tolerant of sulfur, while those involving the late transition metals showed bulk sulfide phases.

  18. Collagenous gastritis: a case report, morphologic evaluation, and review.

    PubMed

    Vesoulis, Z; Lozanski, G; Ravichandran, P; Esber, E

    2000-05-01

    Collagenous gastritis is rare; there are only four previous case reports. Histologic features seem to overlap with the other "collagenous enterocolitides"; however, pathologic criteria are not yet established for the diagnosis of collagenous gastritis. We describe an additional case of ostensible collagenous gastritis in a patient who initially presented with celiac sprue and subsequently developed colonic manifestations of mucosal ulcerative colitis. Endoscopic biopsies of the stomach revealed deposition of patchy, very thick bandlike subepithelial collagen in gastric antral mucosa, focal superficial epithelial degeneration, numerous intraepithelial lymphocytes, and a dense lamina propria lymphoplasmacytic infiltrate. Image analysis evaluation of gastric antral biopsies demonstrated a mean thickness of subepithelial collagen of 27.07 micron. Morphologic comparison was made with age-matched control groups of 10 patients who had normal gastric mucosal biopsies and 10 patients who had "chronic" gastritis, which revealed mean subepithelial collagen measures of 1.37 micron and 1.19 micron, respectively. We compared these morphologic findings with those of all previous case reports of collagenous gastritis and propose a pathologic definition based on the limited combined data. It seems that subepithelial collagen is dramatically thickened in reported cases of collagenous gastritis, with a cumulative mean measure of 36.9 micron. It is also apparent from this and previous reports that the thickened subepithelial collagen is accompanied by a chronic or chronic active gastritis and sometimes intraepithelial lymphocytes and surface epithelial damage. Recently described associations of lymphocytic gastritis, sprue, and lymphocytic colitis as well as collagenous and lymphocytic colitis suggest a common pathogenesis that empirically may include collagenous gastritis in the same disease spectrum. We propose that collagenous gastritis can be confidently identified by using

  19. LARP6 Meets Collagen mRNA: Specific Regulation of Type I Collagen Expression

    PubMed Central

    Zhang, Yujie; Stefanovic, Branko

    2016-01-01

    Type I collagen is the most abundant structural protein in all vertebrates, but its constitutive rate of synthesis is low due to long half-life of the protein (60–70 days). However, several hundred fold increased production of type I collagen is often seen in reparative or reactive fibrosis. The mechanism which is responsible for this dramatic upregulation is complex, including multiple levels of regulation. However, posttranscriptional regulation evidently plays a predominant role. Posttranscriptional regulation comprises processing, transport, stabilization and translation of mRNAs and is executed by RNA binding proteins. There are about 800 RNA binding proteins, but only one, La ribonucleoprotein domain family member 6 (LARP6), is specifically involved in type I collagen regulation. In the 5′untranslated region (5’UTR) of mRNAs encoding for type I and type III collagens there is an evolutionally conserved stem-loop (SL) structure; this structure is not found in any other mRNA, including any other collagen mRNA. LARP6 binds to the 5′SL in sequence specific manner to regulate stability of collagen mRNAs and their translatability. Here, we will review current understanding of how is LARP6 involved in posttranscriptional regulation of collagen mRNAs. We will also discuss how other proteins recruited by LARP6, including nonmuscle myosin, vimentin, serine threonine kinase receptor associated protein (STRAP), 25 kD FK506 binding protein (FKBP25) and RNA helicase A (RHA), contribute to this process. PMID:27011170

  20. Synthesis, antibacterial and cytotoxic activity evaluation of hydroxyurea derivatives.

    PubMed

    Kos, Ivan; Jadrijević-Mladar, Milena; Butula, Ivan; Biruš, Mladen; Maravić-Vlahoviček, Gordana; Dabelić, Sanja

    2013-06-01

    5 Synthesis and biological evaluation of a series (N = 16) of cyclic and acyclic hydroxyurea derivatives, including benzotriazole-, isocyanuric acid- and biuret-containing compounds, are disclosed. 1-N-(benzyloxycarbamoyl)benzotriazole was used as a benzyloxyisocyanate donor, a useful intermediate in the preparation of substituted hydroxyurea. Antibacterial activities of synthesized hydroxyurea derivatives were tested on three E. coli strains, i.e., a strain susceptible to antibiotics, a strain resistant to macrolide antibiotics and a strain resistant to aminoglycoside antibiotics. Six compounds (three acyclic and three cyclic hydroxyureas) showed growth inhibition of the tested E. coli strains, with different specificity toward each strain. Results of the cytotoxic activity evaluation revealed that twelve out of sixteen test compounds were cytotoxic to human acute monocytic leukemia THP-1 and/or human acute T cell leukemia Jurkat cell line. 1-(N-hydroxycarbamoyl) benzotriazole () increased the metabolic activity of both cell lines. Two compounds, 1-(N-hydroxycarbamoyl) benzotriazole (5) and N,N',N''-trihydroxybiuret (15), were identified as potential NO donors.

  1. Resonant active sites in catalytic ammonia synthesis: A structural model

    NASA Astrophysics Data System (ADS)

    Cholach, Alexander R.; Bryliakova, Anna A.; Matveev, Andrey V.; Bulgakov, Nikolai N.

    2016-03-01

    Adsorption sites Mn consisted of n adjacent atoms M, each bound to the adsorbed species, are considered within a realistic model. The sum of bonds Σ lost by atoms in a site in comparison with the bulk atoms was used for evaluation of the local surface imperfection, while the reaction enthalpy at that site was used as a measure of activity. The comparative study of Mn sites (n = 1-5) at basal planes of Pt, Rh, Ir, Fe, Re and Ru with respect to heat of N2 dissociative adsorption QN and heat of Nad + Had → NHad reaction QNH was performed using semi-empirical calculations. Linear QN(Σ) increase and QNH(Σ) decrease allowed to specify the resonant Σ for each surface in catalytic ammonia synthesis at equilibrium Nad coverage. Optimal Σ are realizable for Ru2, Re2 and Ir4 only, whereas other centers meet steric inhibition or unreal crystal structure. Relative activity of the most active sites in proportion 5.0 × 10- 5: 4.5 × 10- 3: 1: 2.5: 3.0: 1080: 2270 for a sequence of Pt4, Rh4, Fe4(fcc), Ir4, Fe2-5(bcc), Ru2, Re2, respectively, is in agreement with relevant experimental data. Similar approach can be applied to other adsorption or catalytic processes exhibiting structure sensitivity.

  2. Proximal and distal changes in collagen content of peripheral nerve that follow transection and crush lesions.

    PubMed

    Eather, T F; Pollock, M; Myers, D B

    1986-05-01

    Collagen content of rat sciatic nerve was measured 10 weeks after either nerve transection or nerve crush. Nerve transection led to a significant increase in fascicular collagen in nerve segments 2.5 mm proximal and distal to the injury site. Remote from the transection, fascicular collagen was also significantly increased, this effect being most marked distally. Nerve crush by comparison resulted in only a small increase in fascicular collagen, significantly less than after transection. The greater amount of fascicular collagen far distal to the nerve injury could relate to a predominantly caudal endoneurial flow of inflammatory or growth factors. Differences in the amount of fascicular collagen formed after nerve transection compared with nerve crush are clearly due to factors other than axonal degeneration, and may relate to collagen synthesis by denervated Schwann cells or to the severity of the nerve injury.

  3. Collagen Fibrils: Nanoscale Ropes

    PubMed Central

    Bozec, Laurent; van der Heijden, Gert; Horton, Michael

    2007-01-01

    The formation of collagen fibrils from staggered repeats of individual molecules has become “accepted” wisdom. However, for over thirty years now, such a model has failed to resolve several structural and functional questions. In a novel approach, it was found, using atomic force microscopy, that tendon collagen fibrils are composed of subcomponents in a spiral disposition—that is, their structure is similar to that of macroscale ropes. Consequently, this arrangement was modeled and confirmed using elastic rod theory. This work provides new insight into collagen fibril structure and will have wide application—from the design of scaffolds for tissue engineering and a better understanding of pathogenesis of diseases of bone and tendon, to the conservation of irreplaceable parchment-based museum exhibits. PMID:17028135

  4. Degradation of type IV collagen by neoplastic human skin fibroblasts

    SciTech Connect

    Sheela, S.; Barrett, J.C.

    1985-02-01

    An assay for the degradation of type IV (basement membrane) collagen was developed as a biochemical marker for neoplastic cells from chemically transformed human skin fibroblasts. Type IV collagen was isolated from basement membrane of Syrian hamster lung and type I collagen was isolated from rat tails; the collagens were radioactively labelled by reductive alkylation. The abilities of normal (KD) and chemically transformed (Hut-11A) human skin fibroblasts to degrade the collagens were studied. A cell-associated assay was performed by growing either normal or transformed cells in the presence of radioactively labelled type IV collagen and measuring the released soluble peptides in the medium. This assay also demonstrated that KD cells failed to synthesize an activity capable of degrading type IV collagen whereas Hut-11A cells degraded type IV collagen in a linear manner for up to 4 h. Human serum at very low concentrations, EDTA and L-cysteine inhibited the enzyme activity, whereas protease inhibitors like phenylmethyl sulfonyl fluoride, N-ethyl maleimide or soybean trypsin inhibitor did not inhibit the enzyme from Hut-11A cells. These results suggest that the ability to degrade specifically type IV collagen may be an important marker for neoplastic human fibroblasts and supports a role for this collagenase in tumor cell invasion.

  5. Development of a recombinant human collagen-type III based hemostat.

    PubMed

    Yang, C; Hillas, P; Tang, J; Balan, J; Notbohm, H; Polarek, J

    2004-04-15

    Animal-tissue-derived collagen, containing mostly type I collagen with a minor amount of type III collagen, has been widely used in the production of hemostats for many decades, although it has been known for a long time that type III collagen is more likely to induce platelet aggregation in vitro. Because it is hard to purify type III from animal tissue, it has not been possible to correlate this finding with in vivo data. In this report, it is demonstrated that recombinant human collagen III fibrils are more capable of inducing platelet aggregation in vitro than those comprised of bovine collagen I, in agreement with previously published data on tissue-derived type III collagen. When formed into three-dimensional matrices, the use of type III collagen results in formulations with better mechanical integrity, larger surface area, and higher hemostatic activity in a rabbit spleen injury model as compared with commercially available hemostats formed from bovine type I collagen.

  6. Tenascin-x deficiency mimics ehlers-danlos syndrome in mice through alteration of collagen deposition

    SciTech Connect

    Mao, J.R.; Taylor, G.; Dean, W.B.; Wagner, D.R.; Afzal, V.; Lotz, J.C.; Rubin, E.M.; Bristow, J.

    2002-03-01

    Tenascin-X is a large extracellular matrix protein of unknown function1-3. Tenascin-X deficiency in humans is associated with Ehlers-Danlos syndrome4,5, a generalized connective tissue disorder resulting from altered metabolism of the fibrillar collagens6. Because TNXB is the first Ehlers-Danlos syndrome gene that does not encode a fibrillar collagen or collagen-modifying enzyme7-14, we suggested that tenascin-X might regulate collagen synthesis or deposition15. To test this hypothesis, we inactivated Tnxb in mice. Tnxb-/- mice showed progressive skin hyperextensibility, similar to individuals with Ehlers-Danlos syndrome. Biomechanical testing confirmed increased deformability and reduced tensile strength of their skin. The skin of Tnxb-/- mice was histologically normal, but its collagen content was significantly reduced. At the ultrastructural level, collagen fibrils of Tnxb-/- mice were of normal size and shape, but the density of fibrils in their skin was reduced, commensurate with the reduction in collagen content. Studies of cultured dermal fibroblasts showed that although synthesis of collagen I by Tnxb-/- and wildtype cells was similar, Tnxb-/- fibroblasts failed to deposit collagen I into cell-associated matrix. This study confirms a causative role for TNXB in human Ehlers-Danlos syndrome and suggests that tenascin-X is an essential regulator of collagen deposition by dermal fibroblasts.

  7. Synthesis and Antiproliferative Activities of Quebecol and Its Analogs

    PubMed Central

    Pericherla, Kasiviswanadharaju; Shirazi, Amir Nasrolahi; Rao, V. Kameshwara; Tiwari, Rakesh K.; DaSilva, Nicholas; McCaffrey, Kellen T.; Beni, Yousef A.; González-Sarrías, Antonio; Seeram, Navindra P.; Parang, Keykavous; Kumar, Anil

    2013-01-01

    Simple and efficient synthesis of quebecol and a number of its analogs was accomplished in five steps. The synthesized compounds were evaluated for antiproliferative activities against human cervix adenocarcinoma (HeLa), human ovarian carcinoma (SKOV-3), human colon carcinoma (HT-29), and human breast adenocarcinoma (MCF-7) cancer cell lines. Among all the compounds, 7c, 7d, 7f, and 8f exhibited antiproliferative activities against four tested cell lines with inhibition over 80% at 75 μM after 72 h, whereas, compound 7b and 7g were more selective towards MCF-7 cell line. The IC50 values for compounds 7c, 7d, and 7f were 85.1 μM, 78.7 μM, and 80.6 μM against MCF-7 cell line, respectively, showing slightly higher antiproliferactive activtiy than the synthesized and isolated quebecol with an IC50 value of 104.2 μM against MCF-7. PMID:23953195

  8. Synthesis and photocatalytic activity of titania microspheres with hierarchical structures

    SciTech Connect

    Cheng, Qian-Qian; Cao, Ying; Yang, Lin; Zhang, Pei-Pei; Wang, Kui; Wang, Hua-Jie

    2011-03-15

    Research highlights: {yields} Lauryl alcohol directs the formation of titania with hierarchical structures. {yields} Hierarchical structures endow the higher specific area to titania. {yields} Titania had a higher photocatalytic activity than commercial Degussa P25. {yields} The synthesis process of the target product is low-cost. -- Abstract: A combined sol-gel and solvothermal process was introduced to fabricate the titania microspheres with hierarchical structures by using lauryl alcohol as the structure-directing agent. Scanning electron microscope, high-resolution transmission electron microscope, Fourier transform infrared spectrograph and powder X-ray powder diffraction indicated that the molar ratio of lauryl alcohol, water and tetra-n-butyl titanate was the key factor for the formation of the mono-dispersed titania with anatase phase and the optimal ratio was 1.2:4:1. The diameter of the end-product was 523 {+-} 74 nm and it was composed of smaller nanoparticles with about 6.8 nm size in diameter. Photocatalytic activity of the end-product was investigated by employing Rhodamine B and Methylene blue as the model compounds. The target microspheres exhibited the higher photocatalytic efficiency compared with commercial Degussa P25 titania and this result might be due to the hierarchical structures of microspheres according to the analysis of Brunauer-Emmett-Teller specific surface areas.

  9. Synthesis of marmycin A and investigation into its cellular activity

    NASA Astrophysics Data System (ADS)

    Cañeque, Tatiana; Gomes, Filipe; Mai, Trang Thi; Maestri, Giovanni; Malacria, Max; Rodriguez, Raphaël

    2015-09-01

    Anthracyclines such as doxorubicin are used extensively in the treatment of cancers. Anthraquinone-related angucyclines also exhibit antiproliferative properties and have been proposed to operate via similar mechanisms, including direct genome targeting. Here, we report the chemical synthesis of marmycin A and the study of its cellular activity. The aromatic core was constructed by means of a one-pot multistep reaction comprising a regioselective Diels-Alder cycloaddition, and the complex sugar backbone was introduced through a copper-catalysed Ullmann cross-coupling, followed by a challenging Friedel-Crafts cyclization. Remarkably, fluorescence microscopy revealed that marmycin A does not target the nucleus but instead accumulates in lysosomes, thereby promoting cell death independently of genome targeting. Furthermore, a synthetic dimer of marmycin A and the lysosome-targeting agent artesunate exhibited a synergistic activity against the invasive MDA-MB-231 cancer cell line. These findings shed light on the elusive pathways through which anthraquinone derivatives act in cells, pointing towards unanticipated biological and therapeutic applications.

  10. Synthesis and Antiangiogenic Activity of N-Alkylated Levamisole Derivatives

    PubMed Central

    Sylvest, Lene; Friis, Tina; Staerk, Dan; Jørgensen, Flemming Steen; Olsen, Christian A.; Houen, Gunnar

    2012-01-01

    Inhibition of angiogenesis is a promising addition to current cancer treatment strategies. Neutralization of vascular endothelial growth factor by monoclonal antibodies is clinically effective but may cause side effects due to thrombosis. Low molecular weight angiogenesis inhibitors are currently less effective than antibody treatment and are also associated with serious side effects. The discovery of new chemotypes with efficient antiangiogenic activity is therefore of pertinent interest. (S)-Levamisole hydrochloride, an anthelminthic drug approved for human use and with a known clinical profile, was recently shown to be an inhibitor of angiogenesis in vitro and exhibited tumor growth inhibition in mice. Here we describe the synthesis and in vitro evaluation of a series of N-alkylated analogues of levamisole with the aim of characterizing structure–activity relationships with regard to inhibition of angiogenesis. N-Methyllevamisole and p-bromolevamisole proved more effective than the parent compound, (S)-levamisole hydrochloride, with respect to inhibition of angiogenesis and induction of undifferentiated cluster morphology in human umbilical vein endothelial cells grown in co-culture with normal human dermal fibroblasts. Interestingly, the cluster morphology caused by N-methyllevamisole was different than the clusters observed for levamisole, and a third “cord-like” morphology resembling that of the known drug suramin was observed for an aniline-containing derivative. New chemotypes exhibiting antiangiogenic effects in vitro are thus described, and further investigation of their underlying mechanism of action is warranted. PMID:23024819

  11. Structure and function of collagen types

    SciTech Connect

    Mayne, R.; Burgeson, R.E.

    1987-01-01

    This book contains 10 chapters. Some of the chapter titles are: The Classical Collagens: Types I, II, and III; Type IV Collagen; Type IX Collagen; and Analysis of Collagen Structure by Molecular Biology Techniques.

  12. In vitro collagen fibril alignment via incorporation of nanocrystalline cellulose.

    PubMed

    Rudisill, Stephen G; DiVito, Michael D; Hubel, Allison; Stein, Andreas

    2015-01-01

    This study demonstrates a method for producing ordered collagen fibrils on a similar length scale to those in the cornea, using a one-pot liquid-phase synthesis. The alignment persists throughout samples on the mm scale. The addition of nanocrystalline cellulose (NCC), a biocompatible and widely available material, to collagen prior to gelation causes the fibrils to align and achieve a narrow size distribution (36±8nm). The effects of NCC loading in the composites on microstructure, transparency and biocompatibility are studied by scanning electron microscopy, ultraviolet-visible spectroscopy and cell growth experiments. A 2% loading of NCC increases the transparency of collagen while producing an ordered microstructure. A mechanism is proposed for the ordering behavior on the basis of enhanced hydrogen bonding during collagen gel formation.

  13. Expeditious oligosaccharide synthesis via selective, semi-orthogonal, and orthogonal activation

    PubMed Central

    Kaeothip, Sophon; Demchenko, Alexei V.

    2011-01-01

    Traditional strategies for oligosaccharide synthesis often require extensive protecting and/or leaving group manipulations between each glycosylation step, thereby increasing the total number of synthetic steps while decreasing the efficiency of the synthesis. In contrast, expeditious strategies allow for the rapid chemical synthesis of complex carbohydrates by minimizing extraneous chemical manipulations. Oligosaccharide synthesis by selective activation of one leaving group over another is one such expeditious strategy. Herein, the significant improvements that have recently emerged in the area of the selective activation are discussed. The development of orthogonal strategy further expands the scope of the selective activation methodology. Surveyed in this article, are representative examples wherein these excellent innovations have been applied to the synthesis of various oligosaccharide sequences. PMID:21663897

  14. Relationship of lipogenic enzyme activities to the rate of rat liver fatty acid synthesis

    SciTech Connect

    Nelson, G.; Kelley, D.; Schmidt, P.; Virk, S.; Serrato, C.

    1986-05-01

    The mechanism by which diet regulates liver lipogenesis is unclear. Here the authors report how dietary alterations effect the activities of key enzymes of fatty acid (FA) synthesis. Male Sprague-Dawley rats, 400-500 g, were fasted for 48h and then refed a fat-free, high carbohydrate (HC) diet (75% cal. from sucrose) for 0,3,9,24 and 48h, or refed a HC diet for 48h, then fed a high-fat (HF) diet (44% cal. from corn oil) for 3,9,24 and 48h. The FA synthesis rate and the activities of acetyl CoA carboxylase (AC), fatty acid synthase (FAS), ATP citrate lyase (CL), and glucose 6-phosphate dehydrogenase (G6PDH) were determined in the livers. FA synthesis was assayed with /sup 3/H/sub 2/O, enzyme activities were measured spectrophotometrically except for AC which was assayed with /sup 14/C-bicarbonate. There was no change in the activity of AC during fasting or on the HC diet. Fasting decreased the rate of FA synthesis by 25% and the activities of FAS and CL by 50%; refeeding the HC diet induced parallel changes in FA synthesis and the activities of FAS, CL, and G6PDH. After 9h on the HF diet, FA synthesis had decreased sharply, AC activity increased significantly while no changes were detected in the other activities. Subsequently FA synthesis did not change while the activities of the enzymes decreased slowly. These enzymes did not appear to regulate FA synthesis during inhibition of lipogenesis, but FAS, CL or G6PDH may be rate limiting in the induction phase. Other key factors may regulate FA synthesis during dietary alterations.

  15. Genetic disorders of collagen.

    PubMed Central

    Tsipouras, P; Ramirez, F

    1987-01-01

    Osteogenesis imperfecta, Ehlers-Danlos syndrome, and Marfan syndrome form a group of genetic disorders of connective tissue. These disorders exhibit remarkable clinical heterogeneity which reflects their underlying biochemical and molecular differences. Defects in collagen types I and III have been found in all three syndromes. PMID:3543367

  16. Mechanical properties, biological activity and protein controlled release by poly(vinyl alcohol)-bioglass/chitosan-collagen composite scaffolds: a bone tissue engineering applications.

    PubMed

    Pon-On, Weeraphat; Charoenphandhu, Narattaphol; Teerapornpuntakit, Jarinthorn; Thongbunchoo, Jirawan; Krishnamra, Nateetip; Tang, I-Ming

    2014-05-01

    In the present study, composite scaffolds made with different weight ratios (0.5:1, 1:1 and 2:1) of bioactive glass (15Ca:80Si:5P) (BG)/polyvinyl alcohol (PVA) (PVABG) and chitosan (Chi)/collagen (Col) (ChiCol) were prepared by three mechanical freeze-thaw followed by freeze-drying to obtain the porous scaffolds. The mechanical properties and the in vitro biocompatibility of the composite scaffolds to simulated body fluid (SBF) and to rat osteoblast-like UMR-106 cells were investigated. The results from the studies indicated that the porosity and compressive strength were controlled by the weight ratio of PVABG:ChiCol. The highest compressive modulus of the composites made was 214.64 MPa which was for the 1:1 weight ratio PVABG:ChiCol. Mineralization study in SBF showed the formation of apatite crystals on the PVABG:ChiCol surface after 7 days of incubation. In vitro cell availability and proliferation tests confirmed the osteoblast attachment and growth on the PVABG:ChiCol surface. MTT and ALP tests on the 1:1 weight ratio PVABG:ChiCol composite indicated that the UMR-106 cells were viable. Alkaline phosphatase activity was found to increase with increasing culturing time. In addition, we showed the potential of PVABG:ChiCol drug delivery through PBS solution studies. 81.14% of BSA loading had been achieved and controlled release for over four weeks was observed. Our results indicated that the PVABG:ChiCol composites, especially the 1:1 weight ratio composite exhibited significantly improved mechanical, mineral deposition, biological properties and controlled release. This made them potential candidates for bone tissue engineering applications.

  17. Superelectrophilic activation in superacid HF/SbF(5) and synthesis of benzofused sultams.

    PubMed

    Liu, Fei; Martin-Mingot, Agnès; Jouannetaud, Marie-Paule; Zunino, Fabien; Thibaudeau, Sébastien

    2010-02-19

    The synthesis of benzofused sultams and/or fluorinated sulfonamides, starting from N-allylic sulfonamides, was performed in superacid HF/SbF(5), through superelectrophilic activation. A dramatic effect of superacid composition on the selectivity of the reaction was shown and applied to the synthesis of cyclic 4-aminobenzenesulfonamides. PMID:20078083

  18. Collagen hydrolysate based collagen/hydroxyapatite composite materials

    NASA Astrophysics Data System (ADS)

    Ficai, Anton; Albu, Madalina Georgiana; Birsan, Mihaela; Sonmez, Maria; Ficai, Denisa; Trandafir, Viorica; Andronescu, Ecaterina

    2013-04-01

    The aim of this study was to study the influence of collagen hydrolysate (HAS) on the formation of ternary collagen-hydrolysate/hydroxyapatite composite materials (COLL-HAS/HA). During the precipitation process of HA, a large amount of brushite is resulted at pH = 7 but, practically pure HA is obtained at pH ⩾ 8. The FTIR data reveal the duplication of the most important collagen absorption bands due to the presence of the collagen hydrolysate. The presence of collagen hydrolysate is beneficial for the management of bone and joint disorders such as osteoarthritis and osteoporosis.

  19. Dynamics of Structural Parameters and Accumulation of Collagen Fibrils in Rat Lung after Inhalations of Surfactant-BL at Various Terms of Bleomycin-Induced Alveolitis.

    PubMed

    Volchkov, V A; Dubrovskaya, V F; Valkovich, A A; Klestova, O V; Serzhanina, V A; Zhuikov, A G; Seiliev, A A; Rosenberg, O A

    2016-08-01

    Rats were subjected to surfactant-BL inhalations at the early and late phases of bleomycininduced alveolitis. In both regimens, the drug reduced the severity of inflammation. In the acute phase of alveolitis, the therapeutic effect of inhalation was accompanied by activation of the synthesis of fine lose collagen fibrils. In the late phase of alveolitis, inhalation of surfactant-BL thickened the fibrils and diminished their population in alveolar walls. PMID:27591866

  20. Rescue of type I collagen-deficient phenotype by retroviral-vector-mediated transfer of human pro alpha 1(I) collagen gene into Mov-13 cells.

    PubMed Central

    Stacey, A; Mulligan, R; Jaenisch, R

    1987-01-01

    A full-length cDNA clone corresponding to the human pro alpha 1(I) collagen gene was isolated and inserted into a retrovirus vector. Cell lines were obtained which produced recombinant viruses transducing the collagen cDNA (HUC virus). To test whether the transduced cDNA was functional, Mov-13 mouse cells were infected with the virus. These cells do not produce any type I collagen due to an insertional mutation of the pro alpha 1(I) gene which blocks transcription. While normal amounts of pro alpha 2(I) RNA were synthesized, no alpha 2(I) collagen chains were detectable in the mutant Mov-13 cells. Infection with HUC virus, however, resulted in the production of stable type I collagen, which was secreted into the medium. Analysis of pepsin-resistant proteins indicated that interspecies heterotrimers consisting of human alpha 1(I) and mouse alpha 2(I) collagen chains were secreted by the infected Mov-13 cells. Our results show that pro alpha (I) collagen chains from species as distant as human and mouse can associate to form stable type I collagen. The availability of a retrovirus vector transducing a functional pro alpha 1(I) collagen gene combined with the Mov-13 mutant system should enable us to study the effect of specific mutations on the synthesis, assembly, and function of type I collagen, not only in tissue culture but also in the animal. Images PMID:3599181

  1. Role of Flightless-I (Drosophila) homolog in the transcription activation of type I collagen gene mediated by transforming growth factor beta

    SciTech Connect

    Lim, Mi-Sun; Jeong, Kwang Won

    2014-11-21

    Highlights: • FLII activates TGFβ-mediated expression of COL1A2 gene. • TGFβ induces the association of FLII with SMAD3 and BRG1 in A549 cells. • FLII is required for the recruitment of SWI/SNF complex and chromatin accessibility to COL1A2 promoter. - Abstract: Flightless-I (Drosophila) homolog (FLII) is a nuclear receptor coactivator that is known to interact with other transcriptional regulators such as the SWI/SNF complex, an ATP-dependent chromatin-remodeling complex, at the promoter or enhancer region of estrogen receptor (ER)-α target genes. However, little is known about the role of FLII during transcription initiation in the transforming growth factor beta (TGFβ)/SMAD-dependent signaling pathway. Here, we demonstrate that FLII functions as a coactivator in the expression of type I collagen gene induced by TGFβ in A549 cells. FLII activates the reporter gene driven by COL1A2 promoter in a dose-dependent manner. Co-expression of GRIP1, CARM1, or p300 did not show any synergistic activation of transcription. Furthermore, the level of COL1A2 expression correlated with the endogenous level of FLII mRNA level. Depletion of FLII resulted in a reduction of TGFβ-induced expression of COL1A2 gene. In contrast, over-expression of FLII caused an increase in the endogenous expression of COL1A2. We also showed that FLII is associated with Brahma-related gene 1 (BRG1) as well as SMAD in A549 cells. Notably, the recruitment of BRG1 to the COL1A2 promoter region was decreased in FLII-depleted A549 cells, suggesting that FLII is required for TGFβ-induced chromatin remodeling, which is carried out by the SWI/SNF complex. Furthermore, formaldehyde-assisted isolation of regulatory elements (FAIRE)-quantitative polymerase chain reaction (qPCR) experiments revealed that depletion of FLII caused a reduction in chromatin accessibility at the COL1A2 promoter. These results suggest that FLII plays a critical role in TGFβ/SMAD-mediated transcription of the COL1A2 gene

  2. Plant polyphenols: chemical properties, biological activities, and synthesis.

    PubMed

    Quideau, Stéphane; Deffieux, Denis; Douat-Casassus, Céline; Pouységu, Laurent

    2011-01-17

    Eating five servings of fruits and vegetables per day! This is what is highly recommended and heavily advertised nowadays to the general public to stay fit and healthy! Drinking green tea on a regular basis, eating chocolate from time to time, as well as savoring a couple of glasses of red wine per day have been claimed to increase life expectancy even further! Why? The answer is in fact still under scientific scrutiny, but a particular class of compounds naturally occurring in fruits and vegetables is considered to be crucial for the expression of such human health benefits: the polyphenols! What are these plant products really? What are their physicochemical properties? How do they express their biological activity? Are they really valuable for disease prevention? Can they be used to develop new pharmaceutical drugs? What recent progress has been made toward their preparation by organic synthesis? This Review gives answers from a chemical perspective, summarizes the state of the art, and highlights the most significant advances in the field of polyphenol research.

  3. Mechanochemical Synthesis and Antioxidant Activity of Curcumin-Templated Azoles.

    PubMed

    Sherin, Daisy R; Rajasekharan, Kallikat N

    2015-12-01

    A solvent-free, mechanochemical method for the synthesis of curcumin (1) derived 3,5-bis(styryl)pyrazoles and 3,5-bis(styryl)isoxazole (2a-g) at room temperature, with very short reaction time, is reported. Such earlier structural modifications of curcumin, at its β-diketone unit by transforming it into an isosteric pyrazole or isoxazole unit, required prolonged heating. The evaluation of the antioxidant activity of these compounds, based on DPPH, FRAP, and β-carotene bleaching assays, showed that several of these azoles are better antioxidants than curcumin, with the isoxazole derivative 2g being overall the best. Typically, the inhibition of 2,2-diphenyl-1-picrylhydrazyl (10(-2) mmol), expressed as EC50 values, by curcumin (1), 3,5-bis(4-hydroxy-3-methoxystyryl)pyrazole (2a), and 3,5-bis(4-hydroxy-3-methoxystyryl)isoxazole (2g) are 40 ± 0.06, 14 ± 0.18, and 8 ± 0.11 μmol, respectively. Moreover, the reported method is useful in accessing 3,5-bis(4-hydroxy-3-methoxystyryl)-1-phenylpyrazole (2b), which is important in studies related to neuroprotection and Alzheimer's disease, and 2a and 2g, which are inhibitors of protein kinases involved in neuronal excitotoxicity.

  4. Synthesis and antifungal activities of novel polyheterocyclic spirooxindole derivatives.

    PubMed

    Wu, Jia-Shou; Zhang, Xue; Zhang, Ying-Lao; Xie, Jian-Wu

    2015-05-01

    A series of spirooxindole tetrahydrofuran derivatives 3 were obtained in moderate to good yields via oxindole derivatives 1 and β-arylacrylonitrile derivatives 2via base-mediated cascade [3 + 2] double Michael reactions under mild conditions and the application of this method in the synthesis of bioactive analogues, such as functionalized spirooxindole octahydrofuro[3,4-c]pyridine derivatives 4 which contain two new heterocyclic rings and two quaternary carbon centers, has also been developed. Subsequently, antifungal activities of all of the synthesized compounds were evaluated against five phytopathogenic fungi (Rhizoctonia solani, Fusarium semitectum, Alternaria solani, Valsa mali and Fusarium graminearum) using the mycelium growth rate method. The preliminary results showed that the spirooxindole octahydrofuro[3,4-c]pyridine derivative 4 showed higher growth inhibition of Valsa mali and Fusarium graminearum, than spirooxindole tetrahydrofuran derivatives 3. For example, spirooxindole octahydrofuro[3,4-c]pyridine derivative 4ab, having a bromine atom at the meta position of the benzene ring, was the best compound in inhibiting F. g. with an IC50 value of 3.31, in particular with inhibition of 4ab on F. g. being similar to that of the control cycloheximide (IC50 = 3.3 μg mL(-1)). PMID:25820179

  5. Diastereoselective Synthesis of Biologically Active Cyclopenta[b]indoles.

    PubMed

    Santos, Marilia S; Fernandes, Daniara C; Rodrigues, Manoel T; Regiani, Thais; Andricopulo, Adriano D; Ruiz, Ana Lúcia T G; Vendramini-Costa, Débora B; de Carvalho, João E; Eberlin, Marcos N; Coelho, Fernando

    2016-08-01

    The cyclopenta[b]indole motif is present in several natural and synthetic biologically active compounds, being directly responsible for the biological effects some of them present. We described herein a three step sequence for the synthesis of cyclopenta[b]indoles with a great structural diversity. The method is based on an oxidative Michael addition of suitable indoles on the double bond of Morita-Baylis-Hillman adducts mediated by a hypervalent iodine reagent (IBX) to form β-ketoesters, which were chemoselectively reduced with NaBH4 in THF to give the corresponding β-hydroxy-esters. The diastereoisomeric mixture was then treated with a catalytic amount of triflic acid (20 mol %) to give cyclopenta[b]indoles with overall yields ranging from 8 to 73% (for 2 steps). The acid-catalyzed cyclization step gave the required heterocycles, via an intramolecular Friedel-Crafts reaction, with high diastereoselectivity, where only the trans product was observed. A mechanistic study monitored by ESI-(+)-MS was also conducted to collect evidence about the mechanism of this reaction. The new molecules herein synthesized were also evaluated against a panel of human cancer cells demonstrating a promising antitumoral profile. PMID:27403650

  6. Synthesis of Indoxyl-glycosides for Detection of Glycosidase Activities.

    PubMed

    Böttcher, Stephan; Thiem, Joachim

    2015-01-01

    Indoxyl glycosides proved to be valuable and versatile tools for monitoring glycosidase activities. Indoxyls are released by enzymatic hydrolysis and are rapidly oxidized, for example by atmospheric oxygen, to indigo type dyes. This reaction enables fast and easy screening in vivo without isolation or purification of enzymes, as well as rapid tests on agar plates or in solution (e.g., blue-white screening, micro-wells) and is used in biochemistry, histochemistry, bacteriology and molecular biology. Unfortunately the synthesis of such substrates proved to be difficult, due to various side reactions and the low reactivity of the indoxyl hydroxyl function. Especially for glucose type structures low yields were observed. Our novel approach employs indoxylic acid ester as key intermediates. Indoxylic acid esters with varied substitution patterns were prepared on scalable pathways. Phase transfer glycosylations with those acceptors and peracetylated glycosyl halides can be performed under common conditions in high yields. Ester cleavage and subsequent mild silver mediated glycosylation yields the peracetylated indoxyl glycosides in high yields. Finally deprotection is performed according to Zemplén. PMID:26068577

  7. Plant polyphenols: chemical properties, biological activities, and synthesis.

    PubMed

    Quideau, Stéphane; Deffieux, Denis; Douat-Casassus, Céline; Pouységu, Laurent

    2011-01-17

    Eating five servings of fruits and vegetables per day! This is what is highly recommended and heavily advertised nowadays to the general public to stay fit and healthy! Drinking green tea on a regular basis, eating chocolate from time to time, as well as savoring a couple of glasses of red wine per day have been claimed to increase life expectancy even further! Why? The answer is in fact still under scientific scrutiny, but a particular class of compounds naturally occurring in fruits and vegetables is considered to be crucial for the expression of such human health benefits: the polyphenols! What are these plant products really? What are their physicochemical properties? How do they express their biological activity? Are they really valuable for disease prevention? Can they be used to develop new pharmaceutical drugs? What recent progress has been made toward their preparation by organic synthesis? This Review gives answers from a chemical perspective, summarizes the state of the art, and highlights the most significant advances in the field of polyphenol research. PMID:21226137

  8. Synthesis and biological evaluation of novel peripherally active morphiceptin analogs.

    PubMed

    Gach, Katarzyna; do-Rego, Jean Claude; Fichna, Jakub; Storr, Martin; Delbro, Dick; Toth, Geza; Janecka, Anna

    2010-08-01

    Morphiceptin (Tyr-Pro-Phe-Pro-NH(2)), a tetrapeptide present in the enzymatic digest of bovine beta-casein, is a selective ligand of the mu-opioid receptor. In the present study, we describe the synthesis of a series of novel morphiceptin analogs modified in positions 1-3. Two of the obtained analogs, [Dmt(1), D-Ala(2), D-1-Nal(3)]morphiceptin and [Dmt(1), D-NMeAla(2), D-1-Nal(3)]morphiceptin (Dmt-2',6'-dimethyltyrosine and d-1-Nal-3-(1-naphthyl)-D-alanine)) displayed very high mu-receptor affinity, resistance to enzymatic degradation, and remarkable supraspinally mediated analgesia, as shown in the hot-plate test after intracerebroventricular but not intravenous administration, which indicated that they could not cross the blood-brain barrier. Therefore, these two analogs were further tested in vitro and in vivo towards their possible peripheral analgesic activity and inhibitory effect on gastrointestinal (GI) motility. We report that both peptides showed strong antinociceptive effect in the writhing test after intraperitoneal administration, inhibited smooth muscle contractility in vitro and GI motility in vivo. Taken together, these findings indicate that the novel morphiceptin analogs which induce peripheral, but not central antinociception, inhibit GI transit, and possess exceptional metabolic stability, may provide an interesting approach to the development of peripherally restricted agents for the treatment of GI motility disorders, such as diarrhea or diarrhea-predominant irritable bowel syndrome.

  9. Diastereoselective Synthesis of Biologically Active Cyclopenta[b]indoles.

    PubMed

    Santos, Marilia S; Fernandes, Daniara C; Rodrigues, Manoel T; Regiani, Thais; Andricopulo, Adriano D; Ruiz, Ana Lúcia T G; Vendramini-Costa, Débora B; de Carvalho, João E; Eberlin, Marcos N; Coelho, Fernando

    2016-08-01

    The cyclopenta[b]indole motif is present in several natural and synthetic biologically active compounds, being directly responsible for the biological effects some of them present. We described herein a three step sequence for the synthesis of cyclopenta[b]indoles with a great structural diversity. The method is based on an oxidative Michael addition of suitable indoles on the double bond of Morita-Baylis-Hillman adducts mediated by a hypervalent iodine reagent (IBX) to form β-ketoesters, which were chemoselectively reduced with NaBH4 in THF to give the corresponding β-hydroxy-esters. The diastereoisomeric mixture was then treated with a catalytic amount of triflic acid (20 mol %) to give cyclopenta[b]indoles with overall yields ranging from 8 to 73% (for 2 steps). The acid-catalyzed cyclization step gave the required heterocycles, via an intramolecular Friedel-Crafts reaction, with high diastereoselectivity, where only the trans product was observed. A mechanistic study monitored by ESI-(+)-MS was also conducted to collect evidence about the mechanism of this reaction. The new molecules herein synthesized were also evaluated against a panel of human cancer cells demonstrating a promising antitumoral profile.

  10. The effect of various avocado oils on skin collagen metabolism.

    PubMed

    Werman, M J; Mokady, S; Nimni, M E; Neeman, I

    1991-01-01

    The effects of various avocado oils on collagen metabolism in skin were studied in growing rats fed diets containing 10% (w/w) of the tested oils. Rats fed the unrefined avocado oil extracted with hexane from the intact fruit, its unsaponifiables or the avocado seed oil, showed significant increases in soluble collagen content in skin, though total collagen content was not affected. The increased soluble collagen content appears to be a consequence of the inhibition of lysyl oxidase activity. The active factor was found to be present in the unrefined avocado oil and probably originated from the avocado seed, since collagen metabolism was affected only by fractions which contained lipids fraction from the seed. In comparison rats fed the refined or unrefined soybean oils showed no effects.

  11. Topographic mapping of collagenous gastritis.

    PubMed

    Freeman, H J

    2001-07-01

    A 74-year-old woman was investigated for abdominal pain and diarrhea. Endoscopic examinations including biopsies of the stomach and colon demonstrated the typical subepithelial deposits characteristic of collagenous gastritis and collagenous colitis. Histochemical and ultrastructural methods confirmed the presence of collagen in the subepithelial deposits. The topographic distribution of these collagen deposits and their relationship to the inflammatory process in the stomach were then defined by endoscopic mapping and multiple site biopsies of the mucosa in the gastric body and antrum. These studies indicate that collagenous gastritis not only is distinctive, but also is a far more extensive and diffuse inflammatory process than has previously been appreciated. PMID:11493952

  12. RNA protein interactions governing expression of the most abundant protein in human body, type I collagen.

    PubMed

    Stefanovic, Branko

    2013-01-01

    Type I collagen is the most abundant protein in human body. The protein turns over slowly and its replacement synthesis is low. However, in wound healing or in pathological fibrosis the cells can increase production of type I collagen several hundred fold. This increase is predominantly due to posttranscriptional regulation, including increased half-life of collagen messenger RNAs (mRNAs) and their increased translatability. Type I collagen is composed of two α1 and one α2 polypeptides that fold into a triple helix. This stoichiometry is strictly regulated to prevent detrimental synthesis of α1 homotrimers. Collagen polypeptides are co-translationally modified and the rate of modifications is in dynamic equilibrium with the rate of folding, suggesting coordinated translation of collagen α1(I) and α2(I) polypeptides. Collagen α1(I) mRNA has in the 3' untranslated region (UTR) a C-rich sequence that binds protein αCP, this binding stabilizes the mRNA in collagen producing cells. In the 5' UTR both collagen mRNAs have a conserved stem-loop (5' SL) structure. The 5' SL is critical for high collagen expression, knock in mice with disruption of the 5' SL are resistant to liver fibrosis. the 5' SL binds protein LARP6 with strict sequence specificity and high affinity. LARP6 recruits RNA helicase A to facilitate translation initiation and associates collagen mRNAs with vimentin and nonmuscle myosin filaments. Binding to vimentin stabilizes collagen mRNAs, while nonmuscle myosin regulates coordinated translation of α1(I) and α2(I) mRNAs. When nonmuscle myosin filaments are disrupted the cells secrete only α1 homotrimers. Thus, the mechanism governing high collagen expression involves two RNA binding proteins and development of cytoskeletal filaments.

  13. Ovine-Based Collagen Matrix Dressing: Next-Generation Collagen Dressing for Wound Care

    PubMed Central

    Bohn, Gregory; Liden, Brock; Schultz, Gregory; Yang, Qingping; Gibson, Daniel J.

    2016-01-01

    Significance: Broad-spectrum metalloproteinase (MMP) reduction along with inherent aspects of an extracellular matrix (ECM) dressing can bring about improved wound healing outcomes and shorter treatment duration. Initial reports of clinical effectiveness of a new ovine-based collagen extracellular matrix (CECM) dressing demonstrate benefits in chronic wound healing. Recent Advances: CECM dressings are processed differently than oxidized regenerated cellulose/collagen dressings. CECM dressings consist primarily of collagens I and III arranged as native fibers that retain the three-dimensional architecture present in tissue ECM. As such, ovine-based ECM dressings represent a new generation of collagen dressings capable of impacting a broad spectrum of MMP excess known to be present in chronic wounds. Critical Issues: While MMPs are essential in normal healing, elevated presence of MMPs has been linked to wound failure. Collagen has been shown to reduce levels of MMPs, acting as a sacrificial substrate for excessive proteases in a chronic wound. Preserving collagen dressings in a more native state enhances bioactivity in terms of the ability to affect the chronic wound environment. Clinical observation and assessment may not be sufficient to identify a wound with elevated protease activity that can break down ECM, affect wound fibroblasts, and impair growth factor response. Future Directions: Collagen dressings that target broad-spectrum excessive MMP levels and can be applied early in the course of care may positively impact healing rates in difficult wounds. Next-generation collagen dressings offer broader MMP reduction capacity while providing a provisional dermal matrix or ECM. PMID:26858910

  14. The Synthesis of Active Metabolites and Analogues of Vitamin D3

    NASA Astrophysics Data System (ADS)

    Yakhimovich, R. I.

    1980-04-01

    The literature date on the synthesis of the active metabolites and analogues of vitamin D3 (cholecalciferol), which play an important role in regulating the homeostatis of calcium in the organism, are reviewed. The bibliography includes 150 references.

  15. Synthesis and characterization of vanadium nanoparticles on activated carbon and their catalytic activity in thiophene hydrodesulphurization

    NASA Astrophysics Data System (ADS)

    Pinto, Susana; D'Ornelas, Lindora; Betancourt, Paulino

    2008-06-01

    Vanadium nanoparticles (˜7 nm) stabilized on activated carbon were synthesized by the reduction of VCl 3·3THF with K[BEt 3H]. This material was characterized by inductive coupled plasma-atomic emission spectroscopy (ICP-AES), high-resolution transmission electron microscopy (HRTEM) and X-ray photoelectron spectroscopy (XPS) analyses. The catalytic performance of the carbon-supported vanadium was studied using thiophene hydrodesulfurization (HDS) as model reaction at 300 °C and P = 1 atm. The catalytic activity of the vanadium carbide phase on the activated carbon carrier was more significant than that of the reference catalysts, alumina supported NiMoS. The method proposed for the synthesis of such a catalyst led to an excellent performance of the HDS process.

  16. Isolation and synthesis of polyoxygenated dibenzofurans possessing biological activity.

    PubMed

    Love, Brian E

    2015-06-01

    Reports from the past ten years describing the isolation and/or synthesis of bioactive dibenzofurans possessing three or more oxygen-containing substituents are reviewed. Dibenzofuranoquinones are included in the review.

  17. Heparin fragments modulate the collagen phenotype of fibroblasts from radiation-induced subcutaneous fibrosis

    SciTech Connect

    el Nabout, R.; Martin, M.; Remy, J.; Robert, L.; Lafuma, C. )

    1989-10-01

    Acute local gamma irradiation of porcine skin induces, as in human skin, an extensive and mutilating sclerosis characterized by continuous expansion of the fibrosis invading the adjacent muscle and by accumulation of the macromolecular components of the extracellular matrix. Collagen synthesis, content, and types were studied in the presence of heparin fragments (100 micrograms/10(6) cells) in the culture medium, by measuring the incorporation of the radiolabeled precursor (3H)proline into confluent primary cultures of porcine fibroblasts obtained from normal and irradiated fibrotic dermis. Enhancement in collagen biosynthesis and deposition and preferential increase in collagen type III synthesis were observed in fibrotic fibroblast cultures when compared to those in normal dermis fibroblasts. The total collagen synthesis and the rate of collagen hydroxylation appear unmodified by heparin fragments both in normal and in fibrotic fibroblast cultures. But heparin fragments induce a 10- and 2-fold decrease, respectively, in collagen type III and type V syntheses by fibrosis fibroblasts. As only minor effects upon collagen type III and V are observed in cultures of normal dermis fibroblasts, these results highly suggest that heparin fragments are capable of specifically modulating the collagen phenotype of fibroblasts derived from radiation-induced dermis fibrosis and thus are able to regulate the fibrotic process.

  18. Stereoselective synthesis and antitumoral activity of Z-enyne pseudoglycosides.

    PubMed

    Dantas, Claudio R; de Freitas, Jucleiton J R; Barbosa, Queila P S; Militão, Gardenia C G; Silva, Thiago D S; da Silva, Teresinha G; Paulino, Antônio A S; Freitas, Juliano C R; Oliveira, Roberta A; Menezes, Paulo H

    2016-07-12

    An efficient approach for the synthesis of Z-1,3-enynes based on the coupling reaction of Z-vinyl tellurides and alkynes containing a pseudoglycoside moiety is described. The products were obtained in good yields via a stereoselective way. Preliminary screening against three tumor cell lines indicated that the synthesized compounds are promising intermediates for the synthesis of an array of more potent target structures.

  19. Autocrine activities of basic fibroblast growth factor: regulation of endothelial cell movement, plasminogen activator synthesis, and DNA synthesis

    PubMed Central

    1988-01-01

    We have found that the spontaneous migration of bovine aortic endothelial cells from the edge of a denuded area in a confluent monolayer is dependent upon the release of endogenous basic fibroblast growth factor (bFGF). Cell movement is blocked by purified polyclonal rabbit IgG to bFGF as well as affinity purified anti-bFGF IgG and anti- bFGF F(ab')2 fragments. The inhibitory effect of the immunoglobulins is dependent upon antibody concentration, is reversible, is overcome by the addition of recombinant bFGF, and is removed by affinity chromatography of the antiserum through a column of bFGF-Sepharose. Cell movement is also reversibly inhibited by the addition of protamine sulfate and suramin; two agents reported to block bFGF binding to its receptor. The addition of recombinant bFGF to wounded monolayers accelerates the movement of cells into the denuded area. Transforming growth factor beta which has been shown to antagonize several other effects of bFGF also inhibits cell movement. The anti-bFGF IgG prevents the movement of bovine capillary endothelial cells, BHK-21, NIH 3T3, and human skin fibroblasts into a denuded area. Antibodies to bFGF, as well as suramin and protamine sulfate also suppress the basal levels of plasminogen activator and DNA synthesis in bovine aortic endothelial cells. PMID:3417781

  20. Synthesis and Biological Activities of Organotin(IV) Complexes as Antitumoral and Antimicrobial Agents. A Review.

    PubMed

    Shah, Syed Shoaib Ahmad; Ashfaq, Muhammad; Waseem, Amir; Ahmed, M Mehboob; Najam, Tayyaba; Shaheen, Salma; Rivera, Gildardo

    2015-01-01

    Advances in the use of organotin(IV) compounds have gained relevant interest in both the chemical and pharmaceutical industry. Tin(IV) form stable complexes with a unique structure and physicochemical properties that are used in organic synthesis as heat stabilizers and catalysts, in drug development as biologically active agents, and in other areas. This review focuses on recent progress in the classical and convenient synthesis procedure, on their mechanism of action, and biological activities as antitumoral and antimicrobial agents.

  1. Laccase Catalyzed Synthesis of Iodinated Phenolic Compounds with Antifungal Activity

    PubMed Central

    Ihssen, Julian; Schubert, Mark; Thöny-Meyer, Linda; Richter, Michael

    2014-01-01

    Iodine is a well known antimicrobial compound. Laccase, an oxidoreductase which couples the one electron oxidation of diverse phenolic and non-phenolic substrates to the reduction of oxygen to water, is capable of oxidizing unreactive iodide to reactive iodine. We have shown previously that laccase-iodide treatment of spruce wood results in a wash-out resistant antimicrobial surface. In this study, we investigated whether phenolic compounds such as vanillin, which resembles sub-structures of softwood lignin, can be directly iodinated by reacting with laccase and iodide, resulting in compounds with antifungal activity. HPLC-MS analysis showed that vanillin was converted to iodovanillin by laccase catalysis at an excess of potassium iodide. No conversion of vanillin occurred in the absence of enzyme. The addition of redox mediators in catalytic concentrations increased the rate of iodide oxidation ten-fold and the yield of iodovanillin by 50%. Iodinated phenolic products were also detected when o-vanillin, ethyl vanillin, acetovanillone and methyl vanillate were incubated with laccase and iodide. At an increased educt concentration of 0.1 M an almost one to one molar ratio of iodide to vanillin could be used without compromising conversion rate, and the insoluble iodovanillin product could be recovered by simple centrifugation. The novel enzymatic synthesis procedure fulfills key criteria of green chemistry. Biocatalytically produced iodovanillin and iodo-ethyl vanillin had significant growth inhibitory effects on several wood degrading fungal species. For Trametes versicolor, a species causing white rot of wood, almost complete growth inhibition and a partial biocidal effect was observed on agar plates. Enzymatic tests indicated that the iodinated compounds acted as enzyme responsive, antimicrobial materials. PMID:24594755

  2. Analysis of collagen types synthesized by rabbit ear cartilage chondrocytes in vivo and in vitro.

    PubMed Central

    Madsen, K; von der Mark, K; van Menxel, M; Friberg, U

    1984-01-01

    type I collagen, the same collagen types in vivo, but change their cellular phenotype more rapidly after transfer to monolayer culture, as indicated by the prompt onset of type III collagen synthesis. Images Fig. 1. Fig. 2. Fig. 4. Fig. 6. Fig. 7. PMID:6380497

  3. Hydrogen Sulfide Alleviates Myocardial Collagen Remodeling in Association with Inhibition of TGF-β/Smad Signaling Pathway in Spontaneously Hypertensive Rats

    PubMed Central

    Sun, Lili; Jin, Hongfang; Sun, Lujing; Chen, Siyao; Huang, Yaqian; Liu, Jia; Li, Zhenzhen; Zhao, Manman; Sun, Yan; Tang, Chaoshu; Zhao, Bin; Du, Junbao

    2014-01-01

    The study was designed to explore the role and possible mechanisms of hydrogen sulfide (H2S) in the regulation of myocardial collagen remodeling in spontaneously hypertensive rats (SHRs). We treated nine-week-old male SHRs and age- and sex-matched Wistar–Kyoto rats (WKYs) with NaHS (90 μmol/kg−1·day−1) for 9 wks. At 18 wks, plasma H2S, tail arterial pressure, morphology of the heart, myocardial ultrastructure and collagen volume fraction (CVF), myocardial expressions of collagen I and III protein and procollagen I and III mRNA, transforming growth factor-β1 (TGF-β1), TGF-β type I receptor (TβR-I), type II receptor (TβR-II), p-Smad2 and 3, matrix metalloproteinase (MMP)-13 and tissue inhibitors of MMP (TIMP)-1 proteins were determined. TGF-β1-stimulated cultured cardiac fibroblasts (CFs) were used to further study the mechanisms. The results showed that compared with WKYs, SHRs showed a reduced plasma H2S, elevated tail artery pressure and increased myocardial collagen, TGF-β1, TβR-II, p-Smad2 and p-Smad3 expressions. However, NaHS markedly decreased tail artery pressure and inhibited myocardial collagen, TGF-β1, TβR-II, p-Smad2 and p-Smad3 protein expressions, but H2S had no effect on the expressions of MMP-13 and TIMP-1. Hydralazine reduced blood pressure but had no effect on myocardial collagen, MMP-13 and TIMP-1 expressions and TGF-β1/Smad signaling pathway. H2S prevented activation of the TGF-β1/Smad signaling pathway and abnormal collagen synthesis in CFs. In conclusion, the results suggested that H2S could prevent myocardial collagen remodeling in SHR. The mechanism might be associated with inhibition of collagen synthesis via TGF-β1/Smad signaling pathway. PMID:25222913

  4. Always cleave up your mess: targeting collagen degradation to treat tissue fibrosis

    PubMed Central

    McKleroy, William; Lee, Ting-Hein

    2013-01-01

    Pulmonary fibrosis is a vexing clinical problem with no proven therapeutic options. In the normal lung there is continuous collagen synthesis and collagen degradation, and these two processes are precisely balanced to maintain normal tissue architecture. With lung injury there is an increase in the rate of both collagen production and collagen degradation. The increase in collagen degradation is critical in preventing the formation of permanent scar tissue each time the lung is exposed to injury. In pulmonary fibrosis, collagen degradation does not keep pace with collagen production, resulting in extracellular accumulation of fibrillar collagen. Collagen degradation occurs through both extracellular and intracellular pathways. The extracellular pathway involves cleavage of collagen fibrils by proteolytic enzyme including the metalloproteinases. The less-well-described intracellular pathway involves binding and uptake of collagen fragments by fibroblasts and macrophages for lysosomal degradation. The relationship between these two pathways and their relevance to the development of fibrosis is complex. Fibrosis in the lung, liver, and skin has been associated with an impaired degradative environment. Much of the current scientific effort in fibrosis is focused on understanding the pathways that regulate increased collagen production. However, recent reports suggest an important role for collagen turnover and degradation in regulating the severity of tissue fibrosis. The objective of this review is to evaluate the roles of the extracellular and intracellular collagen degradation pathways in the development of fibrosis and to examine whether pulmonary fibrosis can be viewed as a disease of impaired matrix degradation rather than a disease of increased matrix production. PMID:23564511

  5. Stability and cellular responses to fluorapatite-collagen composites.

    PubMed

    Yoon, Byung-Ho; Kim, Hae-Won; Lee, Su-Hee; Bae, Chang-Jun; Koh, Young-Hag; Kong, Young-Min; Kim, Hyoun-Ee

    2005-06-01

    Fluorapatite (FA)-collagen composites were synthesized via a biomimetic coprecipitation method in order to improve the structural stability and cellular responses. Different amounts of ammonium fluoride (NH4F), acting as a fluorine source for FA, were added to the precipitation of the composites. The precipitated composites were freeze-dried and isostatically pressed in a dense body. The added fluorine was incorporated nearly fully into the apatite structure (fluoridation), and a near stoichiometric FA-collagen composite was obtained with complete fluoridation. The freeze-dried composites had a typical biomimetic network, consisting of collagen fibers and precipitates of nano-sized apatite crystals. The human osteoblast-like cells on the FA-collagen composites exhibited significantly higher proliferation and differentiation (according to alkaline phosphatase activity) than those on the hydroxyapatite-collagen composite. These enhanced osteoblastic cell responses were attributed to the fluorine release and the reduced dissolution rate.

  6. Human leucocyte neutral proteases, with special reference to collagen metabolism.

    PubMed

    Kobayashi, S; Nagai, Y

    1978-09-01

    Three different types of neutral proteases related to collagen metabolism have been found in the granule fraction of human leucocytes from normal adults, using collagen, gelatin, and synthetic peptides as substrates. These are collagenase, an enzyme showing a potent hydrolytic activity against gelatin but little against native collagen, and one splitting the cross-links region of collagen. Their molecular weights were estimated to be about 75,000 150,000, and 25,000, respectively, by gel chromatography. The former two enzymes were inhibited by a alpha2-macroglobulin and ethylenediaminetetraacetate, but not by alpha1-proteinase inhibitor (alpha1-antitrypsin) or phenylmethylsulfonylfluoride, while the latter enzyme, associated in behavior with an enzyme hydrolyzing succinyl-(l-alanyl)3-p-nitroanilide, was inhibited by alpha1-proteinase inhibitor, alpha2-macroglobulin, and phenylmethylsulfonylfluoride, but not by ethylenediaminetetraacetate. A possible cooperative function of these enzymes in collagen catabolism is discussed.

  7. Collagenous gastritis revealed by severe anemia in a child.

    PubMed

    Côté, J F; Hankard, G F; Faure, C; Mougenot, J F; Holvoet, L; Cézard, J P; Navarro, J; Peuchmaur, M

    1998-08-01

    Collagenous gastritis is a rare histopathological disorder of unknown origin, characterized by a subepithelial collagen deposit greater than 10 microm thick, associated with an inflammatory infiltrate of the gastric mucosa. This report describes a second pediatric case of collagenous gastritis, revealed by severe anemia caused by gastric bleeding, as was the first case. Unlike the adult cases of collagenous gastritis, lesions were limited to the stomach, and remained unchanged on six series of biopsies taken during a 30 month follow-up, despite treatment with omeprazole, sucralfate and corticosteroids. An immunohistochemical study showed signs of local immune activation on all biopsy specimens, including overexpression of HLA-DR by epithelial cells, increased numbers of CD3+ intraepithelial lymphocytes, and CD25+ cells in the lamina propria. Although the cause of the disease remains unclear, our findings suggest that the histopathological lesions of collagenous gastritis may result from a local immune process. PMID:9712433

  8. Robust controllers for the Middeck Active Control Experiment using Popov controller synthesis

    NASA Technical Reports Server (NTRS)

    How, Jonathan P.; Hall, Steven R.

    1993-01-01

    Recent work in robust control with real parameter uncertainties has focused on absolute stability and its connections to real mu theory. In particular, the research has investigated the Popov stability criterion and its associated Lur'e-Postnikov Liapunov functions. State space representations of this Popov stability analysis tests are included in an H2 design formulation to provide a powerful technique for robust controller synthesis. This synthesis approach uses a state space optimization procedure to design controllers that minimize an overbound of an H2 cost functional and satisfy stability analysis tests based on the Popov multiplier. The controller and stability multiplier coefficients are optimized simultaneously, which avoids the iteration and curve-fitting procedures required by the D-K algorithm of mu synthesis. While previous work has demonstrated this synthesis approach on benchmark control problems, the purpose of this paper is to use Popov controller synthesis to design robust compensators for the Middeck Active Control Experiment (MACE).

  9. Potency of Fish Collagen as a Scaffold for Regenerative Medicine

    PubMed Central

    Yamamoto, Kohei; Yanagiguchi, Kajiro

    2014-01-01

    Cells, growth factors, and scaffold are the crucial factors for tissue engineering. Recently, scaffolds consisting of natural polymers, such as collagen and gelatin, bioabsorbable synthetic polymers, such as polylactic acid and polyglycolic acid, and inorganic materials, such as hydroxyapatite, as well as composite materials have been rapidly developed. In particular, collagen is the most promising material for tissue engineering due to its biocompatibility and biodegradability. Collagen contains specific cell adhesion domains, including the arginine-glycine-aspartic acid (RGD) motif. After the integrin receptor on the cell surface binds to the RGD motif on the collagen molecule, cell adhesion is actively induced. This interaction contributes to the promotion of cell growth and differentiation and the regulation of various cell functions. However, it is difficult to use a pure collagen scaffold as a tissue engineering material due to its low mechanical strength. In order to make up for this disadvantage, collagen scaffolds are often modified using a cross-linker, such as gamma irradiation and carbodiimide. Taking into account the possibility of zoonosis, a variety of recent reports have been documented using fish collagen scaffolds. We herein review the potency of fish collagen scaffolds as well as associated problems to be addressed for use in regenerative medicine. PMID:24982861

  10. Heterogeneity of collagens in rabbit cornea: type III collagen

    SciTech Connect

    Cintron, C.; Hong, B.S.; Covington, H.I.; Macarak, E.J.

    1988-05-01

    Whole neonate rabbit corneas and adult corneas containing 2-week-old scars were incubated in the presence of (/sup 14/C) glycine. Radiolabeled collagen extracted from the corneas and scar tissue were analyzed by sodium dodecylsulfate/polyacrylamide gel electrophoresis and fluorography to determine the types and relative quantity of collagen polypeptides present and synthesized by these tissues. In addition to other collagen types, type III was found in both neonate cornea and scar tissue from adult cornea, albeit in relatively small quantities. Type III collagen in normal cornea was associated with the residue after pepsin digestion and formic acid extraction of the tissue, and the same type of collagen was extracted from scar tissue after similar treatment. Type III collagen-specific monoclonal antibody bound to developing normal corneas and healing adult tissue sections, as determined by immunofluorescence. Antibody binding was localized to the endothelium and growing Descemet's membrane in fetal and neonate corneas, and restricted to the most posterior region of the corneal scar tissue. Although monoclonal antibody to keratan sulfate, used as a marker for stromal fibroblasts, bound to most of the scar tissue, the antibody failed to bind to the posterior scar tissue positive for type III collagen. We conclude that endothelial cells from fetal and neonate rabbit cornea and endothelium-derived fibroblasts from healing wounds of adult cornea synthesize and deposit type III collagen. Moreover, this collagen appears to be incorporated into the growing Descemet's membrane of normal corneas and narrow posterior portion of the scar tissue.

  11. Phlorizin, an Active Ingredient of Eleutherococcus senticosus, Increases Proliferative Potential of Keratinocytes with Inhibition of MiR135b and Increased Expression of Type IV Collagen.

    PubMed

    Choi, Hye-Ryung; Nam, Kyung-Mi; Lee, Hyun-Sun; Yang, Seung-Hye; Kim, Young-Soo; Lee, Jongsung; Date, Akira; Toyama, Kazumi; Park, Kyoung-Chan

    2016-01-01

    E. senticosus extract (ESE), known as antioxidant, has diverse pharmacologic effects. It is also used as an antiaging agent for the skin and phlorizin (PZ) is identified as a main ingredient. In this study, the effects of PZ on epidermal stem cells were investigated. Cultured normal human keratinocytes and skin equivalents are used to test whether PZ affects proliferative potential of keratinocytes and how it regulates these effects. Skin equivalents (SEs) were treated with ESE and the results showed that the epidermis became slightly thickened on addition of 0.002% ESE. The staining intensity of p63 as well as proliferating cell nuclear antigen (PCNA) is increased, and integrin α6 was upregulated. Analysis of ESE confirmed that PZ is the main ingredient. When SEs were treated with PZ, similar findings were observed. In particular, the expression of integrin α6, integrin β1, and type IV collagen was increased. Levels of mRNA for type IV collagen were increased and levels of miR135b were downregulated. All these findings suggested that PZ can affect the proliferative potential of epidermal cells in part by microenvironment changes via miR135b downregulation and following increased expression of type IV collagen. PMID:27042261

  12. Phlorizin, an Active Ingredient of Eleutherococcus senticosus, Increases Proliferative Potential of Keratinocytes with Inhibition of MiR135b and Increased Expression of Type IV Collagen

    PubMed Central

    Choi, Hye-Ryung; Nam, Kyung-Mi; Lee, Hyun-Sun; Yang, Seung-Hye; Kim, Young-Soo; Lee, Jongsung; Date, Akira; Toyama, Kazumi; Park, Kyoung-Chan

    2016-01-01

    E. senticosus extract (ESE), known as antioxidant, has diverse pharmacologic effects. It is also used as an antiaging agent for the skin and phlorizin (PZ) is identified as a main ingredient. In this study, the effects of PZ on epidermal stem cells were investigated. Cultured normal human keratinocytes and skin equivalents are used to test whether PZ affects proliferative potential of keratinocytes and how it regulates these effects. Skin equivalents (SEs) were treated with ESE and the results showed that the epidermis became slightly thickened on addition of 0.002% ESE. The staining intensity of p63 as well as proliferating cell nuclear antigen (PCNA) is increased, and integrin α6 was upregulated. Analysis of ESE confirmed that PZ is the main ingredient. When SEs were treated with PZ, similar findings were observed. In particular, the expression of integrin α6, integrin β1, and type IV collagen was increased. Levels of mRNA for type IV collagen were increased and levels of miR135b were downregulated. All these findings suggested that PZ can affect the proliferative potential of epidermal cells in part by microenvironment changes via miR135b downregulation and following increased expression of type IV collagen. PMID:27042261

  13. Synthesis, Characterization and Antitumour Activity of Some Butyltin(IV) Cysteaminates and N,N-Dimethylcysteaminates

    PubMed Central

    Gielen, Marcel; Hollein, Martin; de Vos, Dick

    2000-01-01

    The synthesis and characterization of four di- and tri-n-butyltin cysteaminates and N,N-dimethylcysteaminates and three protonated / quaternized derivatives are reported. They all exhibit moderate or high in vitro cytotoxic activity. Six of seven compounds presented in this work are more active than cisplatin, etoposide and 5-fluorouracil, but less active than methotrexate and doxorubicin. PMID:18475950

  14. Collagen XII Contributes to Epicardial and Connective Tissues in the Zebrafish Heart during Ontogenesis and Regeneration

    PubMed Central

    Marro, Jan; Pfefferli, Catherine; de Preux Charles, Anne-Sophie; Bise, Thomas

    2016-01-01

    Zebrafish heart regeneration depends on cardiac cell proliferation, epicardium activation and transient reparative tissue deposition. The contribution and the regulation of specific collagen types during the regenerative process, however, remain poorly characterized. Here, we identified that the non-fibrillar type XII collagen, which serves as a matrix-bridging component, is expressed in the epicardium of the zebrafish heart, and is boosted after cryoinjury-induced ventricular damage. During heart regeneration, an intense deposition of Collagen XII covers the outer epicardial cap and the interstitial reparative tissue. Analysis of the activated epicardium and fibroblast markers revealed a heterogeneous cellular origin of Collagen XII. Interestingly, this matrix-bridging collagen co-localized with fibrillar type I collagen and several glycoproteins in the post-injury zone, suggesting its role in tissue cohesion. Using SB431542, a selective inhibitor of the TGF-β receptor, we showed that while the inhibitor treatment did not affect the expression of collagen 12 and collagen 1a2 in the epicardium, it completely suppressed the induction of both genes in the fibrotic tissue. This suggests that distinct mechanisms might regulate collagen expression in the outer heart layer and the inner injury zone. On the basis of this study, we postulate that the TGF-β signaling pathway induces and coordinates formation of a transient collagenous network that comprises fibril-forming Collagen I and fiber-associated Collagen XII, both of which contribute to the reparative matrix of the regenerating zebrafish heart. PMID:27783651

  15. Tunability of collagen matrix mechanical properties via multiple modes of mineralization.

    PubMed

    Smith, Lester J; Deymier, Alix C; Boyle, John J; Li, Zhen; Linderman, Stephen W; Pasteris, Jill D; Xia, Younan; Genin, Guy M; Thomopoulos, Stavros

    2016-02-01

    Functionally graded, mineralized collagen tissues exist at soft-to-hard material attachments throughout the body. However, the details of how collagen and hydroxyapatite mineral (HA) interact are not fully understood, hampering efforts to develop tissue-engineered constructs that can assist with repair of injuries at the attachments of soft tissues to bone. In this study, spatial control of mineralization was achieved in collagen matrices using simulated body fluids (SBFs). Based upon previous observations of poor bonding between reconstituted collagen and HA deposited using SBF, we hypothesized that mineralizing collagen in the presence of fetuin (which inhibits surface mineralization) would lead to more mineral deposition within the scaffold and therefore a greater increase in stiffness and toughness compared with collagen mineralized without fetuin. We tested this hypothesis through integrated synthesis, mechanical testing and modelling of graded, mineralized reconstituted collagen constructs. Results supported the hypothesis, and further suggested that mineralization on the interior of reconstituted collagen constructs, as promoted by fetuin, led to superior bonding between HA and collagen. The results provide us guidance for the development of mineralized collagen scaffolds, with implications for bone and tendon-to-bone tissue engineering.

  16. Laminin modified infection-preventing collagen membrane containing silver sulfadiazine-hyaluronan microparticles.

    PubMed

    Lee, Jong-Eun; Park, Jong-Chul; Lee, Kwang Hoon; Oh, Sang Ho; Suh, Hwal

    2002-06-01

    The newly developed laminin modified infection-preventing collagen membrane consists of a 3 component laminate, comprising 2 outer collagen layers and a central laminin layer. The 2 outer collagen layers (dense and porous layers) were fabricated by air-drying and freeze drying, respectively, and the laminin layer was formed by a straightforward liquid coating method. In addition, hyaluronan based microparticles containing silver sulfadiazine (AgSD) were incorporated into the 2 collagen layers (AgSD content 50 microg/cm2). Laminin coated collagen surfaces did not promote fibroblast attachment but showed a retarded fibroblast proliferation rate and an increased rate of collagen synthesis versus pure collagen surfaces. In an animal study, a laminin coating on a nonmedicated collagen membrane significantly increased both wound size reduction and vessel proliferation 7 days after application versus polyurethane film. Interestingly, the laminin coated AgSD medicated collagen membrane demonstrated higher wound size reduction and vessel proliferation and lower inflammation than the polyurethane control, suggesting that the laminin AgSD medicated collagen membrane substantially improves dermal wound healing.

  17. Measurement of Microbial Activity and Growth in the Ocean by Rates of Stable Ribonucleic Acid Synthesis

    PubMed Central

    Karl, David M.

    1979-01-01

    A relatively simple and extremely sensitive technique for measuring rates of stable ribonucleic acid (RNA) synthesis was devised and applied to bacterial cultures and seawater samples. The procedure is based upon the uptake and incorporation of exogenous radiolabeled adenine into cellular RNA. To calculate absolute rates of synthesis, measurements of the specific radioactivity of the intracellular adenosine 5′-triphosphate pools (precursor to RNA) and of the total amount of radioactivity incorporated into stable cellular RNA per unit time are required. Since the rate of RNA synthesis is positively correlated with growth rate, measurements of RNA synthesis should be extremely useful for estimating and comparing the productivities of microbial assemblages in nature. Adenosine 5′-triphosphate, adenylate energy charge, and rates of stable RNA synthesis have been measured at a station located in the Columbian Basin of the Caribbean Sea. A subsurface peak in RNA synthesis (and therefore growth) was located within the dissolved oxygen minimum zone (450 m), suggesting in situ microbiological utilization of dissolved molecular oxygen. Calculations of the specific rates of RNA synthesis (i.e., RNA synthesis per unit of biomass) revealed that the middepth maximum corresponded to the highest specific rate of growth (420 pmol of adenine incorporated into RNA·day−1) of all depths sampled, including the euphotic zone. The existence of an intermediate depth zone of active microbial growth may be an important site for nutrient regeneration and may serve as a source of reduced carbon for mesopelagic and deep sea environments. PMID:16345461

  18. Collagen macromolecular drug delivery systems

    SciTech Connect

    Gilbert, D.L.

    1988-01-01

    The objective of this study was to examine collagen for use as a macromolecular drug delivery system by determining the mechanism of release through a matrix. Collagen membranes varying in porosity, crosslinking density, structure and crosslinker were fabricated. Collagen characterized by infrared spectroscopy and solution viscosity was determined to be pure and native. The collagen membranes were determined to possess native vs. non-native quaternary structure and porous vs. dense aggregate membranes by electron microscopy. Collagen monolithic devices containing a model macromolecule (inulin) were fabricated. In vitro release rates were found to be linear with respect to t{sup {1/2}} and were affected by crosslinking density, crosslinker and structure. The biodegradation of the collagen matrix was also examined. In vivo biocompatibility, degradation and {sup 14}C-inulin release rates were evaluated subcutaneously in rats.

  19. Altered Matrix Metalloproteinase-2 and -9 Expression/Activity Links Placental Ischemia and Anti-angiogenic sFlt-1 to Uteroplacental and Vascular Remodeling and Collagen Deposition in Hypertensive Pregnancy

    PubMed Central

    Li, Wei; Mata, Karina M.; Mazzuca, Marc Q.; Khalil, Raouf A.

    2014-01-01

    Preeclampsia is a complication of pregnancy manifested as maternal hypertension and often fetal growth restriction. Placental ischemia could be an initiating event, but the linking mechanisms leading to hypertension and growth restriction are unclear. We have shown an upregulation of matrix metalloproteinases (MMPs) during normal pregnancy (Norm-Preg). To test the role of MMPs in hypertensive-pregnancy (HTN-Preg), maternal and fetal parameters, MMPs expression, activity and distribution, and collagen and elastin content were measured in uterus, placenta and aorta of Norm-Preg rats and in rat model of reduced uteroplacental perfusion pressure (RUPP). Maternal blood pressure was higher, and uterine, placental and aortic weight, and the litter size and pup weight were less in RUPP than Norm-Preg rats. Western blots and gelatin zymography revealed decreases in amount and gelatinase activity of MMP-2 and MMP-9 in uterus, placenta and aorta of RUPP compared with Norm-Preg rats. Immunohistochemistry confirmed reduced MMPs in uterus, placenta and aortic media of RUPP rats. Collagen, but not elastin, was more abundant in uterus, placenta and aorta of RUPP than Norm-Preg rats. The anti-angiogenic factor soluble fms-like tyrosine kinase-1 (sFlt-1) decreased MMPs in uterus, placenta and aorta of Norm-Preg rats, and vascular endothelial growth factor (VEGF) reversed the decreases in MMPs in tissues of RUPP rats. Thus placental ischemia and anti-angiogenic sFlt-1 decrease uterine, placental and vascular MMP-2 and MMP-9, leading to increased uteroplacental and vascular collagen, and growth-restrictive remodeling in HTN-Preg. Angiogenic factors and MMP activators may reverse the decrease in MMPs and enhance growth-permissive remodeling in preeclampsia. PMID:24704473

  20. Tetrandrine reverses human cardiac myofibroblast activation and myocardial fibrosis.

    PubMed

    Teng, Guoqi; Svystonyuk, Daniyil; Mewhort, Holly E M; Turnbull, Jeannine D; Belke, Darrell D; Duff, Henry J; Fedak, Paul W M

    2015-06-15

    Tetrandrine (TTD) is a calcium channel blocker with documented antifibrotic actions. In this study, for the first time, we identified that TTD can directly prevent in vitro human cardiac myofibroblast activation and limit in vivo myocardial fibrosis. In vitro, cardiac myofibroblasts from human atrial biopsies (N = 10) were seeded in three-dimensional collagen matrices. Cell-collagen constructs were exposed to transforming growth factor-β1 (10 ng/ml), with or without TTD (1 and 5 μM) for 48 h. Collagen gel contraction, myofibroblast activation (α-smooth muscle actin expression), expression of profibrotic mRNAs, and rate of collagen protein synthesis were compared. TTD decreased collagen gel contraction (79.7 ± 1.3 vs 60.1 ± 8.9%, P < 0.01), α-smooth muscle actin expression (flow cytometry), collagen synthesis ([(3)H]proline incorporation), and collagen mRNA expression. Cell viability was similar between groups (annexin positive cells: 1.7 vs. 1.4%). TTD inhibited collagen gel contraction in the presence of T-type and L-type calcium channel blockers, and the intracellular calcium chelator BAPTA-AM (15 μM), suggesting that the observed effects are not mediated by calcium homeostasis. In vivo, Dahl salt-sensitive hypertensive rats were treated with variable doses of TTD (by intraperitoneal injection over 4 wk) and compared with untreated controls (N = 12). Systemic blood pressure was monitored by tail cuff. Myocardial fibrosis and left ventricular compliance were assessed by histology and passive pressure-volume analysis. Myocardial fibrosis was attenuated compared with untreated controls (%collagen area: 9.4 ± 7.3 vs 2.1 ± 1.0%, P < 0.01). Left ventricular compliance was preserved. In conclusion, TTD reverses human cardiac myofibroblast activation and myocardial fibrosis, independent of calcium channel blockade. PMID:25862829

  1. Arterial calcification: Conscripted by collagen

    NASA Astrophysics Data System (ADS)

    Miller, Jordan D.

    2016-03-01

    In atherosclerotic plaques, patterns of calcification -- which have profound implications for plaque stability and vulnerability to rupture -- are determined by the collagen's content and patterning throughout the plaque.

  2. c-Fos activated phospholipid synthesis is required for neurite elongation in differentiating PC12 cells.

    PubMed

    Gil, Germán A; Bussolino, Daniela F; Portal, Maximiliano M; Alfonso Pecchio, Adolfo; Renner, Marianne L; Borioli, Graciela A; Guido, Mario E; Caputto, Beatriz L

    2004-04-01

    We have previously shown that c-Fos activates phospholipid synthesis through a mechanism independent of its genomic AP-1 activity. Herein, using PC12 cells induced to differentiate by nerve growth factor, the genomic effect of c-Fos in initiating neurite outgrowth is shown as distinct from its nongenomic effect of activating phospholipid synthesis and sustaining neurite elongation. Blocking c-Fos expression inhibited differentiation, phospholipid synthesis activation, and neuritogenesis. In cells primed to grow, blocking c-Fos expression determined neurite retraction. However, transfected cells expressing c-Fos or c-Fos deletion mutants with capacity to activate phospholipid synthesis sustain neurite outgrowth and elongation in the absence of nerve growth factor. Results disclose a dual function of c-Fos: it first releases the genomic program for differentiation and then associates to the endoplasmic reticulum and activates phospholipid synthesis. Because phospholipids are key membrane components, we hypothesize this latter phenomenon as crucial to support membrane genesis demands required for cell growth and neurite elongation. PMID:14767061

  3. The rate of synthesis and decomposition of tissue proteins in hypokinesia and increased muscular activity

    NASA Technical Reports Server (NTRS)

    Fedorov, I. V.; Chernyy, A. V.; Fedorov, A. I.

    1978-01-01

    During hypokinesia and physical loading (swimming) of rats, the radioactivity of skeletal muscle, liver, kidney, heart, and blood proteins was determined after administration of radioactive amino acids. Tissue protein synthesis decreased during hypokinesia, and decomposition increased. Both synthesis and decomposition increased during physical loading, but anabolic processes predominated in the total tissue balance. The weights of the animals decreased in hypokinesia and increased during increased muscle activity.

  4. Peroxidase enzymes regulate collagen extracellular matrix biosynthesis.

    PubMed

    DeNichilo, Mark O; Panagopoulos, Vasilios; Rayner, Timothy E; Borowicz, Romana A; Greenwood, John E; Evdokiou, Andreas

    2015-05-01

    Myeloperoxidase and eosinophil peroxidase are heme-containing enzymes often physically associated with fibrotic tissue and cancer in various organs, without any direct involvement in promoting fibroblast recruitment and extracellular matrix (ECM) biosynthesis at these sites. We report herein novel findings that show peroxidase enzymes possess a well-conserved profibrogenic capacity to stimulate the migration of fibroblastic cells and promote their ability to secrete collagenous proteins to generate a functional ECM both in vitro and in vivo. Mechanistic studies conducted using cultured fibroblasts show that these cells are capable of rapidly binding and internalizing both myeloperoxidase and eosinophil peroxidase. Peroxidase enzymes stimulate collagen biosynthesis at a post-translational level in a prolyl 4-hydroxylase-dependent manner that does not require ascorbic acid. This response was blocked by the irreversible myeloperoxidase inhibitor 4-amino-benzoic acid hydrazide, indicating peroxidase catalytic activity is essential for collagen biosynthesis. These results suggest that peroxidase enzymes, such as myeloperoxidase and eosinophil peroxidase, may play a fundamental role in regulating the recruitment of fibroblast and the biosynthesis of collagen ECM at sites of normal tissue repair and fibrosis, with enormous implications for many disease states where infiltrating inflammatory cells deposit peroxidases.

  5. Cryptic Peptides from Collagen: A Critical Review.

    PubMed

    Banerjee, Pradipta; Shanthi, C

    2016-01-01

    Collagen, a predominant structural protein in extracellular matrix (ECM), is now considered to have probable roles in many biological activities and hence, in different forms have found application as nutraceutical or pharmaceutical therapy option. Many of the biological properties are believed to be due to small hidden peptide residues in the collagen molecules, which come into play after the biodegradation or biosorption of the parent molecule. These peptide regions are called cryptic peptides or by some, as cryptides. The proteolytic hydrolysis of the ECM protein releases the cryptic peptides with many novel biological activities not exhibited directly by the parental protein which include angiogenic, antimicrobial, mitogenic and chemotactic properties. The research for understanding the role of these cryptic peptide regions and making use of them in medical field is very active. Such an understanding could lead to the development of peptide supplements for many biomedical applications. The prolific research in this area is reviewed in this paper. PMID:27173646

  6. Chemical and biological evaluation of Egyptian Nile Tilapia (Oreochromis niloticas) fish scale collagen.

    PubMed

    El-Rashidy, Aiah A; Gad, Ahmed; Abu-Hussein, Abd El-Hay G; Habib, Shaymaa I; Badr, Nadia A; Hashem, Azza A

    2015-08-01

    Collagen is considered to be one of the most useful biomaterials with different medical applications. However, collagen properties differ from one source to another. The aim of this study was to extract, purify, characterize and perform preliminary biological evaluation of type I collagen from scales of Egyptian Nile Tilapia. Pepsin-solubilized collagen (PSC) was successfully prepared from Nile Tilapia fish scale waste. Lyophilized collagen was dissolved in dilute HCl to form acidic collagen solutions (ACS) which was neutralized to form gel. To confirm the biocompatibility of the produced gel, baby hamster kidney (BHK-21) fibroblast cells were seeded onto a 3D collagen gel (0.3% and 0.5%, w/v). The results of an SDS-PAGE test showed that the extracted collagens were type I collagen, with α chain composition of (α1)2α2. Thermal analysis showed that the denaturation temperature was 32 °C. X-ray diffraction (XRD) analysis and Fourier-transform infrared spectra (FTIR) showed that the extracted collagen had a triple helix structure. Active proliferation of BHK-21 cells with no signs of toxicity was evident with both collagen gel concentrations tested. The results show that Nile Tilapia scales can be an effective source of collagen extraction that could be used as a potential biomaterial in biomedical applications.

  7. Fibrocytes Are Not an Essential Source of Type I Collagen During Lung Fibrosis1

    PubMed Central

    Kleaveland, Kathryn R.; Velikoff, Miranda; Yang, Jibing; Agarwal, Manisha; Rippe, Richard A.; Moore, Bethany B.; Kim, Kevin K.

    2014-01-01

    Progressive fibrosis involves accumulation of activated collagen producing mesenchymal cells. Fibrocytes are hematopoietic-derived cells with mesenchymal features that potentially have a unique and critical function during fibrosis. Fibrocytes have been proposed as an important direct contributor of type I collagen deposition during fibrosis based largely on fate-mapping studies. To determine the functional contribution of hematopoietic cell-derived type I collagen to fibrogenesis we utilize a double transgenic system to specifically delete the type I collagen gene across a broad population of hematopoietic cells. These mice develop a robust fibrotic response similar to littermate genotype control mice injured with bleomycin indicating that fibrocytes are not a necessary source of type I collagen. Using collagen-promoter GFP mice we find that fibrocytes express type I collagen. However, fibrocytes with confirmed deletion of the type I collagen gene have readily detectable intracellular type I collagen indicating that uptake of collagen from neighboring cells account for much of the fibrocyte collagen. Collectively these results clarify several seemingly conflicting reports regarding the direct contribution of fibrocytes to collagen deposition. PMID:25281715

  8. A biomaterial composed of collagen and solubilized elastin enhances angiogenesis and elastic fiber formation without calcification.

    PubMed

    Daamen, Willeke F; Nillesen, Suzan T M; Wismans, Ronnie G; Reinhardt, Dieter P; Hafmans, Theo; Veerkamp, Jacques H; van Kuppevelt, Toin H

    2008-03-01

    Elastin is the prime protein in elastic tissues that contributes to elasticity of, for example, lung, aorta, and skin. Upon injury, elastic fibers are not readily replaced, which hampers tissue regeneration. Incorporation of solubilized elastin (hydrolyzed insoluble elastin fibers or elastin peptides) in biomaterials may improve regeneration, because solubilized elastin is able to promote proliferation as well as elastin synthesis. Porous biomaterials composed of highly purified collagen without and without elastin fibers or solubilized elastin were prepared by freezing and lyophilization. Solubilized elastin formed spherical structures that were incorporated in the collagenous part of the scaffolds and that persisted after chemical crosslinking of the scaffolds. Crosslinked scaffolds were subcutaneously implanted in young Sprague Dawley rats. Collagen-solubilized elastin and collagen scaffolds showed no calcification in this sensitive calcification model, in contrast to scaffolds containing elastin fibers. Collagen-solubilized elastin scaffolds also induced angiogenesis, as revealed by type IV collagen staining, and promoted elastic fiber synthesis, as shown with antibodies against rat elastin and fibrillin-1. It is concluded that scaffolds produced from collagen and solubilized elastin present a non-calcifying biomaterial with a capacity for soft-tissue regeneration, especially in relation to elastic fiber synthesis.

  9. Akt mediated phosphorylation of LARP6; critical step in biosynthesis of type I collagen

    PubMed Central

    Zhang, Yujie; Stefanovic, Branko

    2016-01-01

    La ribonucleoprotein domain family, member 6 (LARP6) is the RNA binding protein, which regulates translation of collagen mRNAs and synthesis of type I collagen. Posttranslational modifications of LARP6 and how they affect type I collagen synthesis have not been studied. We show that in lung fibroblasts LARP6 is phosphorylated at 8 serines, 6 of which are located within C-terminal domain. Phosphorylation of LARP6 follows a hierarchical order; S451 phosphorylation being a prerequisite for phosphorylations of other serines. Inhibition of PI3K/Akt pathway reduced the phosphorylation of LARP6, but had no effect on the S451A mutant, suggesting that PI3K/Akt pathway targets S451 and we have identified Akt as the responsible kinase. Overexpression of S451A mutant had dominant negative effect on collagen biosynthesis; drastically reduced secretion of collagen and induced hyper-modifications of collagen α2 (I) polypeptides. This indicates that LARP6 phosphorylation at S451 is critical for regulating translation and folding of collagen polypeptides. Akt inhibitor, GSK-2141795, which is in clinical trials for treatment of solid tumors, reduced collagen production by human lung fibroblasts with EC50 of 150 nM. This effect can be explained by inhibition of LARP6 phosphorylation and suggests that Akt inhibitors may be effective in treatment of various forms of fibrosis. PMID:26932461

  10. A complement C3 inhibitor specifically targeted to sites of complement activation effectively ameliorates collagen-induced arthritis in DBA/1J mice.

    PubMed

    Song, Hongbin; Qiao, Fei; Atkinson, Carl; Holers, V Michael; Tomlinson, Stephen

    2007-12-01

    Collagen-induced arthritis (CIA) represents an animal model of autoimmune polyarthritis with similarities to human rheumatoid arthritis, and therapy with various systemic complement-inhibitory proteins has been investigated in this model with varying results. We investigated the use of complement receptor 2 (CR2)-Crry, a complement inhibitor with the ability to target C3 breakdown products deposited in a rheumatic joint. Following induction of CIA in DBA/1J mice, animals were treated with either PBS or CR2-Crry (every other day, every 4 days, or with a single injection). The severity of clinical disease was significantly reduced in all CR2-Crry-treated groups compared with controls. Joints from mice receiving multiple doses of CR2-Crry showed significantly decreased inflammatory cell infiltrate, cartilage damage, pannus formation, and bone damage. CR2-Crry treatment also significantly decreased production of anti-collagen IgG and the inflammatory cytokines TNF-alpha and IL-1beta. IL-10 and IL-1Ra levels were increased in CR2-Crry-treated mice. CR2-Crry localized preferentially in the joints of mice with CIA. Analysis of IgG and C3 deposition in the joints of treated animals indicated that both complement regulation and the modulation of anti-collagen Ab production contributed to the protective effects of CR2-Crry. Of interest, a previous study reported that Crry-Ig, an untargeted counterpart of CR2-Crry, had minimal effect on disease, even when administered at a sufficiently high dose to maintain chronic complement inhibition. PMID:18025232

  11. Modulation of secreted proteins of mouse mammary epithelial cells by the collagenous substrata

    SciTech Connect

    Lee, E.Y.H.; Parry, G.; Bissell, M.J.

    1984-01-01

    It has been shown previously that cultures of mouse mammary epithelial cells retain their characteristic morphology and their ability to produce ..gamma..-casein, a member of the casein gene family, only if they are maintained on floating collagen gels. In this paper we show: (a) Cells on floating collagen gels secrete not only ..gamma..-casein but also ..cap alpha../sub 1/-, ..cap alpha../sub 2/-, and ..beta..-caseins. These are not secreted by cells on plastic and are secreted to only a very limited extent by cells on attached collagen gels. (b) The floating collagen gel regulates at the level of synthesis and/or stabilization of the caseins rather than at the level of secretion alone. Contraction of the floating gel is important in that cells cultured on floating glutaraldehyde cross-linked gels do not secrete any of the caseins. (c) The secretion of an 80,000-mol-wt protein, most probably transferrin, and a 67,000-mol-wt protein, probably butyrophilin, a major protein of the milk fat globule membrane, are partially modulated by substrata. However, in contrast to the caseins, these are always detectable in media from cells cultured on plastic and attached gels. (d) Whey acidic protein, a major whey protein, is actively secreted by freshly isolated cells but is secreted in extremely limited quantities in cultured cells regardless of the nature of the substratum used. Lactalbumin secretion is also decreased significantly in cultured cells. (e) A previously unreported set of proteins, which may be minor milk proteins, are prominently secreted by the mammary cells on all substrata tested. We conclude that while the substratum profoundly influences the secretion of the caseins, it does not regulate the expression of every milk-specific protein in the same way. The mechanistic implications of these findings are discussed.

  12. Ribulose diphosphate carboxylase synthesis in euglena: increased enzyme activity after transferring regreening cells to darkness.

    PubMed

    Lord, J M; Merrett, M J

    1975-05-01

    The transfer of dark-grown cultures of Euglena gracilis Klebs strain Z regreening in the light back into darkness resulted in a dramatic increase in ribulose diphosphate carboxylase activity. On a culture volume basis activity increased 4-fold over a 24-hour dark period, although on a protein basis activity declined because of rapid cell division. Mixed assays with light- and dark-growing cell extracts provided no evidence for the removal of an inhibitor of ribulose diphosphate carboxylase upon transferring regreening cells back to darkness. Although ribulose diphosphate carboxylase activity increased over a 24-hour dark period, there was no concomitant increase in the potential of the cells for photosynthetic carbon dioxide fixation.Higher light intensities than the optimum for ribulose diphosphate carboxylase synthesis during regreening resulted in a greater relative rate of synthesis on transfer to darkness so that the maximum activity of ribulose diphosphate carboxylase reached in the dark was constant, regardless of light intensity during regreening. A tentative hypothesis to explain these results is that the synthesis of the large and small subunits of ribulose diphosphate carboxylase occur at different stages of cell development, light being necessary for the synthesis of the large subunit and also for regulating the synthesis of the small subunit.

  13. Collagen for bone tissue regeneration.

    PubMed

    Ferreira, Ana Marina; Gentile, Piergiorgio; Chiono, Valeria; Ciardelli, Gianluca

    2012-09-01

    In the last decades, increased knowledge about the organization, structure and properties of collagen (particularly concerning interactions between cells and collagen-based materials) has inspired scientists and engineers to design innovative collagen-based biomaterials and to develop novel tissue-engineering products. The design of resorbable collagen-based medical implants requires understanding the tissue/organ anatomy and biological function as well as the role of collagen's physicochemical properties and structure in tissue/organ regeneration. Bone is a complex tissue that plays a critical role in diverse metabolic processes mediated by calcium delivery as well as in hematopoiesis whilst maintaining skeleton strength. A wide variety of collagen-based scaffolds have been proposed for different tissue engineering applications. These scaffolds are designed to promote a biological response, such as cell interaction, and to work as artificial biomimetic extracellular matrices that guide tissue regeneration. This paper critically reviews the current understanding of the complex hierarchical structure and properties of native collagen molecules, and describes the scientific challenge of manufacturing collagen-based materials with suitable properties and shapes for specific biomedical applications, with special emphasis on bone tissue engineering. The analysis of the state of the art in the field reveals the presence of innovative techniques for scaffold and material manufacturing that are currently opening the way to the preparation of biomimetic substrates that modulate cell interaction for improved substitution, restoration, retention or enhancement of bone tissue function. PMID:22705634

  14. [Total Synthesis of Biologically Active Natural Products toward Elucidation of the Mode of Action].

    PubMed

    Yoshida, Masahito

    2015-01-01

    Total synthesis of biologically active cyclodepsipeptide destruxin E using solid- and solution-phase synthesis is described. The solid-phase synthesis of destruxin E was initially investigated for the efficient synthesis of destruxin analogues. Peptide elongation from polymer-supported β-alanine was efficiently performed using DIC/HOBt or PyBroP/DIEA, and subsequent cleavage from the polymer-support under weakly acidic conditions furnished a cyclization precursor in moderate yield. Macrolactonization of the cyclization precursor was smoothly performed using 2-methyl-6-nitrobenzoic anhydride (MNBA)/4-(dimethylamino)pyridine N-oxide (DMAPO) to afford macrolactone in moderate yield. Finally, formation of the epoxide in the side chain via three steps provided destruxin E, and the stereochemistry of the epoxide was determined to be S. Its diastereomer, epi-destruxin E, was also synthesized in the same manner used to synthesize the natural product. The stereochemistry of the epoxide was critical for the V-ATPase inhibition; natural product destruxin E exhibited 10-fold more potent V-ATPase inhibition than epi-destruxin E. Next, the scalable synthesis of destruxin E for in vivo study was also performed via solution-phase synthesis. The scalable synthesis of a key component, (S)-HA-Pro-OH, was achieved using osmium-catalyzed diastereoselective dihydroxylation with (DHQD)2PHAL as a chiral ligand; peptide synthesis using Cbz-protected amino acid derivatives furnished the cyclization precursor on a gram-scale. Macrolactonization smoothly provided the macrolactone without forming a dimerized product, even at 6 mM, and the synthesis of destruxin E was achieved via three steps on a gram scale in high purity (>98%). PMID:26423864

  15. Inositol synthesis regulates the activation of GSK-3α in neuronal cells.

    PubMed

    Ye, Cunqi; Greenberg, Miriam L

    2015-04-01

    The synthesis of inositol provides precursors of inositol lipids and inositol phosphates that are pivotal for cell signaling. Mood stabilizers lithium and valproic acid, used for treating bipolar disorder, cause cellular inositol depletion, which has been proposed as a therapeutic mechanism of action of both drugs. Despite the importance of inositol, the requirement for inositol synthesis in neuronal cells is not well understood. Here, we examined inositol effects on proliferation of SK-N-SH neuroblastoma cells. The essential role of inositol synthesis in proliferation is underscored by the findings that exogenous inositol was dispensable for proliferation, and inhibition of inositol synthesis decreased proliferation. Interestingly, the inhibition of inositol synthesis by knocking down INO1, which encodes inositol-3-phosphate synthase, the rate-limiting enzyme of inositol synthesis, led to the inactivation of GSK-3α by increasing the inhibitory phosphorylation of this kinase. Similarly, the mood stabilizer valproic acid effected transient decreases in intracellular inositol, leading to inactivation of GSK-3α. As GSK-3 inhibition has been proposed as a likely therapeutic mechanism of action, the finding that inhibition of inositol synthesis results in the inactivation of GSK-3α suggests a unifying hypothesis for mechanism of mood-stabilizing drugs. Inositol is an essential metabolite that serves as a precursor for inositol lipids and inositol phosphates. We report that inhibition of the rate-limiting enzyme of inositol synthesis leads to the inactivation of glycogen synthase kinase (GSK) 3α by increasing inhibitory phosphorylation of this kinase. These findings have implications for the therapeutic mechanisms of mood stabilizers and suggest that inositol synthesis and GSK 3α activity are intrinsically related.

  16. 1,25-dihydroxyvitamin D3 and its nuclear receptor repress human α1(I) collagen expression and type I collagen formation

    PubMed Central

    Potter, James J.; Liu, Xiaopu; Koteish, Ayman; Mezey, Esteban

    2013-01-01

    Background Vitamin D deficiency is common in chronic liver disease particularly in those with severe liver fibrosis. Aims: To determine the effect of 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) on the human α1(I) collagen promoter and collagen formation by human stellate LX-2 cells and the mechanism of the effect of the vitamin D receptor (VDR) on the promoter. Methods Type I collagen was assessed by measurements of collagen mRNA and collagen protein and by transfection experiments. Binding of VDR to the α1(I) collagen promoter was determined by EMSA and ChIP assays. Results 1,25-(OH)2D3 decreased human α1(I) collagen mRNA and protein and the secretion of type I collagen by stellate cells after exposure to TGFβ1. Furthermore, 1,25-(OH)2D3 inhibited TGFβ1–induced activation of the α1(I) collagen promoter in transfected LX-2 cells. The effect of 1, 25-(OH)2D3 is mediated by the VDR, which binds at a proximal Sp1 site and also at a newly identified distal site on the collagen promoter. A VDR expression vector reduced the activities of the collagen promoter in transfected LX-2 cells. Conclusions 1,25-(OH)2D3 inhibits type I collagen formation in human stellate cells. The effect of 1,25-(OH)2D3 is mediated by its receptor which binds at a proximal Sp1.1 site and at a newly identified distal site on the collagen promoter. Correction of vitamin D deficiency in patients with chronic liver disease is a potential therapy to inhibit progression of fibrosis. PMID:23413886

  17. Regulation of immune reactivity to collagen in human beings

    SciTech Connect

    Solinger, A.M.; Stobo, J.D.

    1981-08-01

    Denaturated beef collagen was tested for its ability to induce the production of leukocyte inhibition factor among the peripheral blood mononuclear cells from patients with rheumatoid arthritis and normal individuals. Responsiveness, defined as the production of leukocyte inhibition factor sufficient to cause greater than 20% inhibition of leukocyte migration, was significantly (P less than 0.001, X2 . 31.1) associated with HLA-DR4. All HLA-DR4 positive individuals, including subjects without any evidence of synovitis, were collagen responders. There was no significant (P . 0.3) difference in the absolute reactivity of HLA-DR4+ versus HLA-DR4- individuals to respond to another antigen, Candida albicans. Collagen reactivity required interactions between macrophages and T cells and was directed against determinants inherent in the linear polypeptide, (Gly-Pro)n. In 5 normal HLA-DR4- nonresponders tested, absence of discernable reactivity to collagen was associated with the presence of antigen-specific, radiosensitive suppressive T cells. These studies suggest that during the physiologic metabolism of collagen all individuals are exposed to Gly-Pro determinants normally buried in the interstices of the collagen triple helix. In individuals whose major histocompatibility complex contains genes linked to those coding for HLA-DR4, this results in the activation of reactive T cells. Conversely, in individuals lacking these genes, collagen-specific suppressive cells predominate.

  18. Synthetic collagen fascicles for the regeneration of tendon tissue.

    PubMed

    Kew, S J; Gwynne, J H; Enea, D; Brookes, R; Rushton, N; Best, S M; Cameron, R E

    2012-10-01

    The structure of an ideal scaffold for tendon regeneration must be designed to provide a mechanical, structural and chemotactic microenvironment for native cellular activity to synthesize functional (i.e. load bearing) tissue. Collagen fibre scaffolds for this application have shown some promise to date, although the microstructural control required to mimic the native tendon environment has yet to be achieved allowing for minimal control of critical in vivo properties such as degradation rate and mass transport. In this report we describe the fabrication of a novel multi-fibre collagen fascicle structure, based on type-I collagen with failure stress of 25-49 MPa, approximating the strength and structure of native tendon tissue. We demonstrate a microscopic fabrication process based on the automated assembly of type-I collagen fibres with the ability to produce a controllable fascicle-like, structural motif allowing variable numbers of fibres per fascicle. We have confirmed that the resulting post-fabrication type-I collagen structure retains the essential phase behaviour, alignment and spectral characteristics of aligned native type-I collagen. We have also shown that both ovine tendon fibroblasts and human white blood cells in whole blood readily infiltrate the matrix on a macroscopic scale and that these cells adhere to the fibre surface after seven days in culture. The study has indicated that the synthetic collagen fascicle system may be a suitable biomaterial scaffold to provide a rationally designed implantable matrix material to mediate tendon repair and regeneration.

  19. Lymphocytic and Collagenous Colitis.

    PubMed

    Cruz-Correa; Giardiello

    2000-06-01

    Patients with symptomatic collagenous-lymphocytic colitis should eliminate dietary secretagogues such as caffeine- or lactose-containing food from their diet. When possible, use of nonsteroidal anti-inflammatory drugs should be discontinued. If steatorrhea is documented, a low-fat diet may be helpful. In the presence of bile salt malabsorption, binding resins such as cholestyramine might be useful. Nonspecific diarrheal agents such as loperamide hydrochloride, diphenoxylate hydrochloride and atropine, deodorized tincture of opium, or codeine might prove effective in some patients. Antibacterial agents such as bismuth subsalicylate (8 chewable 262-mg tablets daily) have been effective in symptom control. Metronidazole and erythromycin achieve response rates of 60%. Sulfasalazine, at the usual dose of 2 to 4 g daily, used in collagenous-lymphocytic colitis, demonstrated cessation of diarrhea in 1 to 2 weeks for 50% of patients. Other 5-aminosalicylic (5-ASA) compounds are preferred for patients with a history of sulfa allergy, and those who experience adverse reactions to sulfasalazine. Adrenocorticoid medication is reserved for patients whose conventional treatment with sulfasalazine or 5-ASA has failed. Resolution of diarrhea has been documented in 80% to 90% of patients within 1 week of treatment, however, in most patients, long-term therapy is required. Surgical management is reserved for those patients with disease refractory to medical therapy. Colectomy with ileostomy resulted in clinical and histologic resolution in small case series. If there is no abatement of symptoms, rule out other etiologies of diarrhea such as thyroid dysfunction, celiac disease, or bacterial overgrowth. PMID:11097741

  20. Cobalt Molybdenum Oxynitrides: Synthesis, Structural, Characterization, and Catalytic Activity for the Oxygen Reduction Reaction

    SciTech Connect

    Cao, Bingfei; Veith, Gabriel M; Diaz, Rosa; Liu, Jue; Stach, Eric; Adzic, Radoslav R.; Khalifah, P.

    2013-01-01

    Here, we report the synthesis and characterization of CoxMo1 xOyNz compounds supported on carbon black as potential cathode catalysts for ORR. They were prepared by a conventional impregnation method. Their ORR activities in both acid and alkaline electrolytes were evaluated via half-cell measurements. The synthesis temperature and sample composition both strongly impacted their physical and chemical properties. Factors influencing their crystal structures, morphologies and ORR activities will be discussed based on the results of structural and spectroscopic studies.

  1. Efficient one-pot synthesis of tigogenin saponins and their antitumor activities.

    PubMed

    Gu, Guofeng; An, Lian; Fang, Min; Guo, Zhongwu

    2014-01-13

    An efficient synthesis of naturally occurring tigogenin triglycoside 1a and its three derivatives 1b-d bearing different carbohydrate moieties, as well as their antitumor activities, is described. Partially protected thiogalactosides bearing unprotected 2,4-OH or 4-OH groups were used to facilitate regioselective reactions for one-pot sequential multi-step glycosylation, which has significantly simplified the target molecule synthesis. The synthetic saponins 1a-d exhibited much higher anti-tumor activities than the positive control cisplatin against the human epithelial cervical cancer cell (HeLa) as evaluated by CCK-8 assay.

  2. Design, diversity-oriented synthesis and structure activity relationship studies of quinolinyl heterocycles as antimycobacterial agents.

    PubMed

    Rachakonda, Venkatesham; Alla, Manjula; Kotipalli, Sudha Sravanti; Ummani, Ramesh

    2013-01-01

    The current study reports design and diversity oriented synthesis of novel bis heterocycles with a common 2-methyl, C-4 unsubstituted quinoline moiety as the central key heterocycle. Employing reagent based skeletal diversity approach; a facile synthesis of bis heterocycles with different heterocyclic rings at C-3 position of the quinoline moiety has been accomplished. A broad range of heterocyclic frameworks thus obtained were evaluated for their antimycobacterial activity. The active scaffolds were further explored by a parallel library generation in order to establish SAR. Further, low cytotoxicity against A549 cell line enhances the potential of the synthesized molecules as promising antimycobacterial agents. PMID:24189497

  3. Measurement of the Mechanical Properties of Intact Collagen Fibrils

    NASA Astrophysics Data System (ADS)

    Mercedes, H.; Heim, A.; Matthews, W. G.; Koob, T.

    2006-03-01

    Motivated by the genetic disorder Ehlers-Danlos syndrome (EDS), in which proper collagen synthesis is interrupted, we are investigating the structural and mechanical properties of collagen fibrils. The fibrous glycoprotein collagen is the most abundant protein found in the human body and plays a key role in the extracellular matrix of the connective tissue, the properties of which are altered in EDS. We have selected as our model system the collagen fibrils of the sea cucumber dermis, a naturally mutable tissue. This system allows us to work with native fibrils which have their proteoglycan complement intact, something that is not possible with reconstituted mammalian collagen fibrils. Using atomic force microscopy, we measure, as a function of the concentration of divalent cations, the fibril diameter, its response to force loading, and the changes in its rigidity. Through these experiments, we will shed light on the mechanisms which control the properties of the sea cucumber dermis and hope to help explain the altered connective tissue extracellular matrix properties associated with EDS.

  4. Biologically Active Chorionic Gonadotropin: Synthesis by the Human Fetus

    NASA Astrophysics Data System (ADS)

    McGregor, W. G.; Kuhn, R. W.; Jaffe, R. B.

    1983-04-01

    The kidney, and to a slight extent the liver, of human fetuses were found to synthesize and secrete the α subunit common to glycoprotein hormones. Fetal lung and muscle did not synthesize this protein. Since fetal kidney and liver were previously found to synthesize β chorionic gonadotropin, their ability to synthesize bioactive chorionic gonadotropin was also determined. The newly synthesized hormone bound to mouse Leydig cells and elicited a biological response: namely, the synthesis of testosterone. These results suggest that the human fetus may participate in metabolic homeostasis during its development.

  5. Collagen-like antimicrobial peptides.

    PubMed

    Masuda, Ryo; Kudo, Masakazu; Dazai, Yui; Mima, Takehiko; Koide, Takaki

    2016-11-01

    Combinatorial library composed of rigid rod-like peptides with a triple-helical scaffold was constructed. The component peptides were designed to have various combinations of basic and neutral (or hydrophobic) amino acid residues based on collagen-like (Gly-Pro-Yaa)-repeating sequences, inspired from the basic and amphiphilic nature of naturally occurring antimicrobial peptides. Screening of the peptide pools resulted in identification of antimicrobial peptides. A structure-activity relationship study revealed that the position of Arg-cluster at N-terminus and cystine knots at C-terminus in the triple helix significantly contributed to the antimicrobial activity. The most potent peptide RO-A showed activity against Gram-negative Escherichia coli and Gram-positive Bacillus subtilis. In addition, Escherichia coli exposed to RO-A resulted in abnormal elongation of the cells. RO-A was also shown to have remarkable stability in human serum and low cytotoxicity to mammalian cells. © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 453-459, 2016. PMID:27271210

  6. Chemical synthesis of echistatin, a potent inhibitor of platelet aggregation from Echis carinatus: synthesis and biological activity of selected analogs.

    PubMed

    Garsky, V M; Lumma, P K; Freidinger, R M; Pitzenberger, S M; Randall, W C; Veber, D F; Gould, R J; Friedman, P A

    1989-06-01

    Echistatin, a polypeptide from the venom of the saw-scaled viper, Echis carinatus, containing 49 amino acids and 4 cystine bridges was synthesized by solid-phase methodology in 4% yield. In the final step, air oxidation of the octahydroderivative was found to be optimal at pH 8. The synthetic product was shown to be physically and biologically indistinguishable from native material. It inhibits fibrinogen-dependent platelet aggregation stimulated by ADP with IC50 = 3.3 x 10(-8) M and also prevents aggregation initiated by thrombin, epinephrine, collagen, or platelet-activating factor. Reduction of purified synthetic echistatin to octahydroechistatin with dithiothreitol followed by air oxidation regenerated homogeneous echistatin in quantitative yield. This highly specific refolding strongly suggests that the linear sequence of octahydroechistatin contains all of the information that is required for the proper folding of the peptide. The sequence Arg24-Gly-Asp of echistatin occurs also in adhesive glycoproteins that bind to the platelet fibrinogen receptor--a heterodimeric complex composed of glycoproteins IIb and IIIa. In an effort to evaluate the role of this putative binding site we have synthesized analogs of echistatin with substitution of Arg-24. Replacement with ornithine-24 (Orn-24) resulted in an analog having a platelet aggregation inhibitory activity with IC50 = 1.05 x 10(-7) M. Substitution with Ala-24 gave IC50 = 6.1 x 10(-7) M. The inhibitory activity of the corresponding short sequence analogs Arg-Gly-Asp-Phe (IC50 = 6 x 10(-6) M), Orn-Gly-Asp-Phe (IC50 = 1.3 x 10(-4) M), and Ala-Gly-Asp-Phe (IC50 = 5.0 x 10(-4) M) was also determined. These results suggest that arginine plays a more important role in the binding of the tetrapeptide than in that of echistatin. PMID:2726764

  7. Collagen fibril formation during development

    SciTech Connect

    Fleischmajer, R.; Perlish, J.S.; Timpl, R.; Olsen, B.R.

    1987-05-01

    Studies with embryonic skin and bone suggested that the aminopropeptide (AP) and carboxylpropeptide (CP) of type I pro-callagen (pro-col) play a role in fibril formation. Chick leg metatarsal tendons were studied by electron microscopy. AP and CP of type I pro-col were purified from chick leg tendons; antibodies developed in rabbits and purity tested by radioimmunoassays. Antibodies were used for immunofluorescence microscopy (IFM) and immunoblotting (IB). The peritendineum, consisting of thin 20-30 nm fibrils, revealed the AP of type I and type III procol. In the tendon area, collagen fibrils were arranged within small compartments and were of uniform diameter at 10d, 14d and 18d. However, beyond 21d, there was confluency of the compartments and a wide range of fibril diameters. IFM revealed fine streaks of collagen, staining with the AP of type I throughout the tendon. The CP was mainly intracellular with only a small amount present in the extracellular space. IB revealed procollagen, pN-collagen (AP+collagen) and pC-collagen, (CP+collagen) at all stages of development. Ratios of pN/pC collagen, determined by spectrophotometric scanning of autoradiographs, correlated well with the distribution of fibril diameter. This study suggests the hypothesis that AP initiates fibrillogenesis while CP may regulate additional fibril growth.

  8. Patterns of autoreactivity to collagen type II in autoimmune MRL/l mice.

    PubMed Central

    Tarkowski, A; Holmdahl, R; Rubin, K; Klareskog, L; Nilsson, L A; Gunnarsson, K

    1986-01-01

    The kinetics and mechanisms for secretion of antibodies against native and denatured collagen type II have been studied in spontaneously arthritic MRL/l mice. Circulating antibodies were quantified by an ELISA assay and frequencies of specific antibody secreting spleen cells by an ELISPOT assay. The degree of humoral immunity to collagen type II increased at late stages of the disease (6 months of age) whereas severe synovitis was seen earlier (5 months of age). Both the appearance of anti-collagen II producing cells and development of synovitis was preceded by and not correlated with a general state of polyclonal B cell activation. In MRL/l mice, collagen II specific antibodies appeared spontaneously and titres were largely unaffected by collagen II immunization. The levels of circulating anti-collagen II antibodies in MRL/l mice were lower, and the antibodies displayed lower avidities and different specificities as compared with the antibodies generated in collagen II high responder DBA/l mice after immunization with collagen II. It is suggested that the antibody response in MRL/l mice against collagen type II does not need MHC-restricted T cell help and that induction of antibody production to collagen II in MRL/l mice is triggered by joint cartilage destruction and subsequent collagen II release. PMID:3516469

  9. Curcumin cross-linked collagen aerogels with controlled anti-proteolytic and pro-angiogenic efficacy.

    PubMed

    Dharunya, G; Duraipandy, N; Lakra, Rachita; Korapatti, Purna Sai; Jayavel, R; Kiran, Manikantan Syamala

    2016-01-01

    This paper elucidates the development of a curcumin cross-linked collagen aerogel system with controlled anti-proteolytic activity and pro-angiogenic efficacy. The results of this study showed that in situ cross-linking of curcumin with collagen leads to the development of aerogels with enhanced physical and mechanical properties. The integrity of collagen after cross-linking with curcumin was studied via FTIR spectroscopy. The results confirmed that the cross-linking with curcumin did not induce any structural changes in the collagen. The curcumin cross-linked collagen aerogels exhibited potent anti-proteolytic and anti-microbial activity. Scanning electron and atomic force microscopic analysis of curcumin cross-linked collagen aerogels showed a 3D microstructure that enhanced the adhesion and proliferation of cells. The highly organized geometry of collagen-curcumin aerogels enhanced the permeability and water-retaining ability required for the diffusion of nutrients that aid cellular growth. The pro-angiogenic properties of collagen-curcumin aerogels were ascribed to the cumulative effect of the nutraceutical and the collagen molecule, which augmented the restoration of damaged tissue. Further, these aerogels exhibited controlled anti-proteolytic activity, which makes them suitable 3D scaffolds for biomedical applications. This study provides scope for the development of biocompatible and bioresorbable collagen aerogel systems that use a nutraceutical as a cross-linker for biomedical applications. PMID:27509047

  10. Type I collagen aging impairs discoidin domain receptor 2-mediated tumor cell growth suppression

    PubMed Central

    Saby, Charles; Buache, Emilie; Brassart-Pasco, Sylvie; El Btaouri, Hassan; Courageot, Marie-Pierre; Van Gulick, Laurence; Garnotel, Roselyne; Jeannesson, Pierre; Morjani, Hamid

    2016-01-01

    Tumor cells are confronted to a type I collagen rich environment which regulates cell proliferation and invasion. Biological aging has been associated with structural changes of type I collagen. Here, we address the effect of collagen aging on cell proliferation in a three-dimensional context (3D). We provide evidence for an inhibitory effect of adult collagen, but not of the old one, on proliferation of human fibrosarcoma HT-1080 cells. This effect involves both the activation of the tyrosine kinase Discoidin Domain Receptor 2 (DDR2) and the tyrosine phosphatase SHP-2. DDR2 and SHP-2 were less activated in old collagen. DDR2 inhibition decreased SHP-2 phosphorylation in adult collagen and increased cell proliferation to a level similar to that observed in old collagen. In the presence of old collagen, a high level of JAK2 and ERK1/2 phosphorylation was observed while expression of the cell cycle negative regulator p21CIP1 was decreased. Inhibition of DDR2 kinase function also led to an increase in ERK1/2 phosphorylation and a decrease in p21CIP1 expression. Similar signaling profile was observed when DDR2 was inhibited in adult collagen. Altogether, these data suggest that biological collagen aging could increase tumor cell proliferation by reducingthe activation of the key matrix sensor DDR2. PMID:27121132

  11. Type I collagen aging impairs discoidin domain receptor 2-mediated tumor cell growth suppression.

    PubMed

    Saby, Charles; Buache, Emilie; Brassart-Pasco, Sylvie; El Btaouri, Hassan; Courageot, Marie-Pierre; Van Gulick, Laurence; Garnotel, Roselyne; Jeannesson, Pierre; Morjani, Hamid

    2016-05-01

    Tumor cells are confronted to a type I collagen rich environment which regulates cell proliferation and invasion. Biological aging has been associated with structural changes of type I collagen. Here, we address the effect of collagen aging on cell proliferation in a three-dimensional context (3D).We provide evidence for an inhibitory effect of adult collagen, but not of the old one, on proliferation of human fibrosarcoma HT-1080 cells. This effect involves both the activation of the tyrosine kinase Discoidin Domain Receptor 2 (DDR2) and the tyrosine phosphatase SHP-2. DDR2 and SHP-2 were less activated in old collagen. DDR2 inhibition decreased SHP-2 phosphorylation in adult collagen and increased cell proliferation to a level similar to that observed in old collagen.In the presence of old collagen, a high level of JAK2 and ERK1/2 phosphorylation was observed while expression of the cell cycle negative regulator p21CIP1 was decreased. Inhibition of DDR2 kinase function also led to an increase in ERK1/2 phosphorylation and a decrease in p21CIP1 expression. Similar signaling profile was observed when DDR2 was inhibited in adult collagen. Altogether, these data suggest that biological collagen aging could increase tumor cell proliferation by reducingthe activation of the key matrix sensor DDR2. PMID:27121132

  12. Helminth Antigens Enable CpG-Activated Dendritic Cells to Inhibit the Symptoms of Collagen-induced Arthritis through Foxp3+ Regulatory T Cells

    PubMed Central

    Carranza, Franco; Falcón, Cristian Roberto; Nuñez, Nicolás; Knubel, Carolina; Correa, Silvia Graciela; Bianco, Ismael; Maccioni, Mariana; Fretes, Ricardo; Triquell, María Fernanda; Motrán, Claudia Cristina; Cervi, Laura

    2012-01-01

    Dendritic cells (DC) have the potential to control the outcome of autoimmunity by modulating the immune response. In this study, we tested the ability of Fasciola hepatica total extract (TE) to induce tolerogenic properties in CpG-ODN (CpG) maturated DC, to then evaluate the therapeutic potential of these cells to diminish the inflammatory response in collagen induced arthritis (CIA). DBA/1J mice were injected with TE plus CpG treated DC (T/C-DC) pulsed with bovine collagen II (CII) between two immunizations with CII and clinical scores CIA were determined. The levels of CII-specific IgG2 and IgG1 in sera, the histological analyses in the joints, the cytokine profile in the draining lymph node (DLN) cells and in the joints, and the number, and functionality of CD4+CD25+Foxp3+ T cells (Treg) were evaluated. Vaccination of mice with CII pulsed T/C-DC diminished the severity and incidence of CIA symptoms and the production of the inflammatory cytokine, while induced the production of anti-inflammatory cytokines. The therapeutic effect was mediated by Treg cells, since the adoptive transfer of CD4+CD25+ T cells, inhibited the inflammatory symptoms in CIA. The in vitro blockage of TGF-β in cultures of DLN cells plus CII pulsed T/C-DC inhibited the expansion of Treg cells. Vaccination with CII pulsed T/C-DC seems to be a very efficient approach to diminish exacerbated immune response in CIA, by inducing the development of Treg cells, and it is therefore an interesting candidate for a cell-based therapy for rheumatoid arthritis (RA). PMID:22848374

  13. Objective Assessment of Endogenous Collagen In Vivo during Tissue Repair by Laser Induced Fluorescence

    PubMed Central

    Prabhu, Vijendra; Rao, Satish B. S.; Fernandes, Edward Mark; Rao, Anuradha C. K.; Prasad, Keerthana; Mahato, Krishna K.

    2014-01-01

    Collagen, a triple helical protein with the primary role of mechanical function, provides tensile strength to the skin, and plays a pivotal task in tissue repair. During tissue regeneration, collagen level increases gradually and therefore, monitoring of such changes in vivo by laser induced fluorescence was the main objective behind the present study. In order to accomplish this, 15 mm diameter excisional wounds were created on six to eight week old Swiss albino mice. The collagen deposition accelerated upon irradiation of single exposure of 2 J/cm2 He-Ne laser dose immediately after wounding was recorded by laser induced autofluorescence in vivo along with un-illuminated and un-wounded controls. Autofluorescence spectra were recorded for each animal of the experimental groups on 0, 5, 10, 30, 45 and 60 days post-wounding, by exciting the granulation tissue/skin with 325 nm He-Cd laser. The variations in the average collagen intensities from the granulation tissue/skin of mice were inspected as a function of age and gender. Further, the spectral findings of the collagen synthesis in wound granulation tissue/un-wounded skin tissues were validated by Picro-Sirius red- polarized light microscopy in a blinded manner through image analysis of the respective collagen birefringence. The in vivo autofluorescence studies have shown a significant increase in collagen synthesis in laser treated animals as compared to the un-illuminated controls. Image analysis of the collagen birefringence further authenticated the ability of autofluorescence in the objective monitoring of collagen in vivo. Our results clearly demonstrate the potential of laser induced autofluorescence in the monitoring of collegen synthesis during tissue regeneration, which may have clinical implications. PMID:24874229

  14. Objective assessment of endogen