Production of Collagenase and Inhibitor (Timp) by Normal, Rheumatoid and Osteoarthritic Synovium In Vitro: Effects of Hydrocortisone and Indomethacin

1981 ◽  
Vol 61 (6) ◽  
pp. 703-710 ◽  
Author(s):  
Mary B. McGuire ◽  
Gillian Murphy ◽  
J. J. Reynolds ◽  
R. G. G. Russell
Keyword(s):  
Animal Studies ◽  
Culture Media ◽  
Active Form ◽  
Prostaglandin E ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Active Collagenase ◽  
In Vitro Effects ◽  
Destructive Processes

1. the amounts of latent and active collagenase and of collagenase inhibitor (TIMP) produced by two normal, three rheumatoid and two osteoarthritic synovial specimens in culture were compared. Normal synovia produced TIMP, but little latent enzyme. Rheumatoid synovia produced higher levels of total collagenase activity than normal, of which up to 50% in one sample was present in the medium in an active form, whereas no specific inhibitory activity due to TIMP was detectable. the amounts of collagenase and TIMP produced by osteoarthritic synovia were more variable and appeared to reflect the degree of inflammation in the tissue at the time of initiating the cultures. 2. Concentrations of TIMP were usually higher in the culture media of normal, rheumatoid and osteoarthritic synovia when hydrocortisone was present. Correspondingly, amounts of total collagenase were reduced. Production of prostaglandin E (PGE) were inhibited in a dose-dependent manner by hydrocortisone. 3. Indomethacin had no consistent effect on the production of TIMP by rheumatoid and osteoarthritic synovia, although it tended to depress production of collagenase. the production of TIMP by normal synovia was depressed by indomethacin. No PGE was detectable in the media when indomethacin was present. 4. These results are consistent with those from previous animal studies, and we conclude that the balance between production of collagenase and TIMP may be critical in determining the extent of the destructive processes in arthritis. the ability of hydrocortisone to suppress production of collagenase and to increase free TIMP concentration, as well as to inhibit synthesis of prostaglandin, may explain in part how the drug exerts its therapeutic effects in patients with rheumatoid arthritis.

scholarly journals Role of leukemia inhibitor factor (LIF) in decidualisation of murine uterine stromal cells in vitro

2004 ◽  
Vol 181 (3) ◽  
pp. 477-492 ◽  
Author(s):  
AA Fouladi Nashta ◽  
CV Andreu ◽  
N Nijjar ◽  
JK Heath ◽  
SJ Kimber
Keyword(s):  
In Vitro Culture ◽  
Stromal Cells ◽  
Control Level ◽  
Calf Serum ◽  
Prostaglandin E ◽  
Dependent Manner ◽  
Alp Activity ◽  
Dose Dependent ◽  
In Cells

Decidualisation of uterine stromal cells is a prerequisite for implantation of the embryo in mice. Here we have used an in vitro culture system in which stromal cells decidualise as indicated by a number of markers, including an increase in alkaline phosphatase (ALP) activity. The latter was used as a quantitative marker of decidualisation in the presence of low (2%) fetal calf serum. Prostaglandin E(2) (PGE(2)), which is known to induce decidualisation, increased ALP activity, and this effect was blocked in a dose-dependent manner by indomethacin. Leukemia inhibitory factor (LIF) was then examined, but it had no effect on PGE(2) secretion. However, LIF suppressed ALP activity in a dose-dependent manner in the presence of 2% serum, while an inhibitor of LIF that competes for binding to its receptor reversed the effect of LIF and increased ALP activity above the control level. In serum-free cultures, stromal cells differentiated rapidly, and no differences were observed between LIF-treated and untreated cultures. Stromal cells produce LIF during in vitro culture, and this peaked at 48 h. Freshly collected stromal cells from both day-2 and -4 pregnant mice expressed mRNA for the LIF receptor, and the transcript level was higher in cells isolated on day 4. However, no differences were observed in the relative levels of transcripts in cells from day 2 and day 4 after culture, nor were there differences between the LIF-treated cultures and controls. Therefore, in this study, we have shown that LIF suppresses decidualisation of murine uterine stromal cells in the presence of serum, this is not due to the regulation of PGE(2) secretion by stromal cells.

scholarly journals Mechanisms underlying the antifibrotic properties of noninhibitory PAI-1 (PAI-1R) in experimental nephritis

2009 ◽  
Vol 297 (4) ◽  
pp. F1045-F1054 ◽  
Author(s):  
Yufeng Huang ◽  
Wayne A. Border ◽  
Daniel A. Lawrence ◽  
Nancy A. Noble
Keyword(s):  
Half Life ◽  
Stable Form ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Therapeutic Action ◽  
Protease Inhibition ◽  
Ecm Degradation ◽  
Dose Dependent ◽  
Pai 1

Administration of a mutant, noninhibitory PAI-1 (PAI-1R), reduces disease in experimental glomerulonephritis. Here we investigated the importance of vitronectin (Vn) binding, PAI-1 stability and protease binding in this therapeutic effect using a panel of PAI-1 mutants differing in half-life, protease binding, and Vn binding. PAI-1R binds Vn normally but does not inhibit proteases. PAI-1AK has a complete defect in Vn binding but retains full inhibitory activity, with a short half-life similar to wild-type (wt)-PAI-1. Mutant 14-lb is identical to wt-PAI-1 but with a longer half-life. PAI-1K has defective Vn binding, inhibits proteases normally, and has a long half-life. In vitro wt-PAI-1 dramatically inhibited degradation of mesangial cell ECM while the AK mutant had much less effect. Mutants 14-1b and PAI-1K, like wt-PAI-1, inhibited matrix degradation but PAI-1R failed to reverse this inhibition although PAI-1R reversed the wt-PAI-1-induced inhibition of ECM degradation in a plasmin-, time-, and dose-dependent manner. Thus the ability of PAI-1 to inhibit ECM degradation is dependent both on its antiproteinase activity and on maintaining an active conformation achieved either by Vn binding or mutation to a stable form. Administration of these PAI-1 mutants to nephritic rats confirmed the in vitro data; only PAI-1R showed therapeutic effects. PAI-1K did not bind to nephritic kidney, indicating that Vn binding is essential to the therapeutic action of PAI-1R. The ability of PAI-1R to remain bound to Vn even in a high-protease environment is very likely the key to its therapeutic efficacy. Furthermore, because both PAI-1R and 14-1b bound to the nephritic kidney in the same pattern and differ only in their ability to bind proteases, lack of protease inhibition is also keyed to PAI-1R's therapeutic action.

scholarly journals Tolerogenic Splenic IDO+Dendritic Cells from the Mice Treated with Induced-Treg Cells Suppress Collagen-Induced Arthritis

2014 ◽  
Vol 2014 ◽  
pp. 1-12 ◽  
Author(s):  
Jie Yang ◽  
Yiming Yang ◽  
Huahua Fan ◽  
Hejian Zou
Keyword(s):  
Dendritic Cells ◽  
Immune Responses ◽  
Foxp3 Expression ◽  
Treated Group ◽  
Treg Cells ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Collagen Induced Arthritis

TGF-β-induced regulatory T cells (iTregs) retain Foxp3 expression and immune-suppressive activity in collagen-induced arthritis (CIA). However, the mechanisms whereby transferred iTregs suppress immune responses, particularly the interplay between iTregs and dendritic cells (DCs)in vivo, remain incompletely understood. In this study, we found that after treatment with iTregs, splenic CD11c+DCs, termed “DCiTreg,” expressed tolerogenic phenotypes, secreted high levels of IL-10, TGF-β, and IDO, and showed potent immunosuppressive activityin vitro. After reinfusion with DCiTreg, marked antiarthritic activity improved clinical scores and histological end-points were observed. The serological levels of inflammatory cytokines and anti-CII antibodies were low and TGF-βproduction was high in the DCiTreg-treated group. DCiTregalso induced new iTregsin vivo. Moreover, the inhibitory activity of DCiTregon CIA was lost following pretreatment with the inhibitor of indoleamine 2,3-dioxygenase (IDO). Collectively, these findings suggest that transferred iTregs could induce tolerogenic characteristics in splenic DCs and these cells could effectively dampen CIA in an IDO-dependent manner. Thus, the potential therapeutic effects of iTregs in CIA are likely maintained through the generation of tolerogenic DCsin vivo.

scholarly journals Induction of PI3K/Akt-Mediated Apoptosis in Osteoclasts Is a Key Approach for Buxue Tongluo Pills to Treat Osteonecrosis of the Femoral Head

2021 ◽  
Vol 12 ◽  
Author(s):  
Dan Wang ◽  
Yicheng Liu ◽  
Dandan Tang ◽  
Shujun Wei ◽  
Jiayi Sun ◽  
...  
Keyword(s):  
Femoral Head ◽  
Functional Annotation ◽  
Caspase 3 ◽  
Differentially Expressed Gene ◽  
Pathway Enrichment Analysis ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Clinical Practices ◽  
Key Genes

The Buxue Tongluo pill (BTP) is a self-made pill with the functions of nourishing blood, promoting blood circulation, dredging collaterals, and relieving pain. It consists of Angelica sinensis (Oliv.) Diels, Pheretima aspergillum (E.Perrier), Panax notoginseng (Burk.) F. H. Chen, Astragalus membranaceus (Fisch.) Bge, and Glycyrrhiza uralensis Fisch. Various clinical practices have confirmed the therapeutic effect of BTP on osteonecrosis of the femoral head (ONFH), but little attention has been paid to the study of its bioactive ingredients and related mechanisms of action. In this study, UPLC/MS-MS combined with GEO data mining was used to construct a bioactive ingredient library of BTP and a differentially expressed gene (DEG) library for ONFH. Subsequently, Cytoscape (3.7.2) software was used to analyze the protein–protein interaction between BTP and DEGs of ONFH to screen the key targets, and functional annotation analysis and pathway enrichment analysis were carried out. Finally, 34 bioactive compounds were screened, which acted on 1,232 targets. A total of 178 DEGs were collected, and 17 key genes were obtained after two screenings. By bioinformatics annotation on these key genes, a total of 354 gene ontology (GO) functional annotation analyses and 42 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were obtained. The present study found that GO and KEGG enrichment were mainly related to apoptosis, suggesting that BTP may exert an anti-ONFH effect by promoting osteoclast apoptosis. Experiments in vitro demonstrated that BTP could increase the mitochondrial membrane potential (MMP) and induce remarkable apoptosis in osteoclasts. Furthermore, we determined the apoptosis marker of cleaved(C)-caspase-3, bcl-2, and bax and found that BTP could upregulate the C-caspase-3 and bax expression in osteoclasts and decrease the expression of bcl-2, p-Akt, and p-PI3K in a dose-dependent manner, indicating that BTP could induce PI3K/Akt-mediated apoptosis in osteoclasts to treat ONFH. This study explored the pharmacodynamic basis and mechanism of BTP against ONFH from the perspective of systemic pharmacology, laying a foundation for further elucidating the therapeutic effects of BTP against ONFH.

scholarly journals The effect of calcium antagonists on the growth of Entamoeba histolytica in vitro

2010 ◽  
Vol 4 (1) ◽  
pp. 5-10
Author(s):  
Zahra’a Abdul-Raheem Ahmed ◽  
Ali H. Ad’hiah ◽  
Amna N. Jasim
Keyword(s):  
Entamoeba Histolytica ◽  
Calcium Antagonists ◽  
Culture Media ◽  
Agar Medium ◽  
Reproduction Rate ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Infusion Agar ◽  
Ethylene Diaminetetraacetic Acid

he E. histolytica parasite was maintained in vitro using Locke-egg medium (LEM) and Liver infusion agar medium (LIAM). The effect of two calcium antagonists (Nifedipine and Ethylene-diaminetetraacetic acid EDTA) on the growth and activity of the parasite in the two culture media was investigated. The calcium antagonists Nifedipine and EDTA inhibited the reproduction rate of E. histolytica in a concentration-dependent manner. For Nifedipine, a concentration of 41.6 mg/ml inhibited the reproduction rate to 99.7% in both media. The EDTA had an approximate effect (98.2 and 95.8)% at a concentration of 0.83 mg/ml in LEM and LIAM media, respectively. Additionally, some cases of a parasite encystment were observed in LEM medium that was treated with Nifedipine.

Plasminogen: an enigmatic zymogen

Blood ◽  
2021 ◽  
Author(s):  
Charithani B Keragala ◽  
Robert L Medcalf
Keyword(s):  
Animal Studies ◽  
Wound Repair ◽  
Active Form ◽  
Cell Behaviour ◽  
Surface Receptors ◽  
Enzymatic Cascades ◽  
Inflammatory Processes ◽  
Over 40 Years

Plasminogen is an abundant plasma protein that exists in various zymogenic forms. Plasmin, the proteolytically active form of plasminogen, is known for its essential role in fibrinolysis. The therapeutic targeting of the fibrinolytic system to date has been for two purposes: to promote plasmin generation for thromboembolic conditions, or to stop plasmin to reduce bleeding. However, both plasmin and plasminogen serve other important functions, some of which are unrelated to fibrin removal. Indeed, for over 40 years, the anti-fibrinolytic agent, tranexamic acid, has been administered for its serendipitously discovered skin whitening properties. Plasmin also plays an important role in the removal of misfolded/aggregated proteins and can trigger other enzymatic cascades including complement. In addition, plasminogen, via binding to one of its dozen cell-surface receptors, can modulate cell behaviour and further influence immune and inflammatory processes. Plasminogen administration itself has been reported to improve thrombolysis and to accelerate wound repair. While many of these more recent findings have been derived from in vitro or animal studies, the use of anti-fibrinolytics to reduce bleeding in humans has revealed additional clinically relevant consequences, particularly in relation to reducing infection risk that is independent of its haemostatic effects. The finding that many viruses harness the host plasminogen to aid infectivity has suggested that anti-fibrinolytic agents may have anti-viral benefits. Here we review the broadening role of the plasminogen activating system in physiology and pathophysiology and how manipulation of this system may be harnessed for benefits unrelated to its conventional application in thrombosis and haemostasis.

Abstract P019: Exosomes from Human CD34+ Cells: Critical Mediators of Proangiogenic Paracrine Effect of EPCs

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Susmita Sahoo ◽  
Sol Misener ◽  
Tina Thorne ◽  
Meredith Millay ◽  
Kathryn M Schultz ◽  
...  
Keyword(s):  
Cell Transplantation ◽  
Human Peripheral Blood ◽  
Limb Ischemia ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Hematopoietic Stem ◽  
Paracrine Effect ◽  
Human Cd34 ◽  
Cd34 Cells

Local transplantation of human CD34+ hematopoietic stem cells has been shown to promote neovascularization in pre-clinical studies in models of myocardial and limb ischemia. In early phase clinical trials, transplantation of CD34+ cells has been associated with reduced angina, improved exercise time and reduced amputation rates. Several studies have suggested that paracrine effects by these pro-angiogenic cells mediate the effects induced by cell transplantation. We hypothesized that CD34+ cells secrete exosomes (Exo), which mediate at least a part of the therapeutic function of the cells. Methods and Results: We isolated Exo from the conditioned media of adult human peripheral blood (PB) CD34+ cells. The angiogenic and therapeutic potency of CD34+ Exo was compared with the intact CD34+ cells and also with PB mononuclear cell (MNC) Exo. Exo from both CD34+ cells and MNC are 50–90nm in size, have cup shaped morphology, and carry known Exo-marker proteins such as CD63, TSG101 and Annexin V as shown by electron microscopy, Western blot and flow cytometry. Compared to CD34+ cells or MNC Exo, CD34+ Exo significantly induces in vitro angiogenic activities such as viability, proliferation and tube formation of HUVECs on matrigel- in a dose dependent manner. In vivo, CD34+ Exo stimulated significant neovascularization in mouse corneal angiogenesis assay (14±4 mm v MNC Exo, 4±1 mm, p<0.01) and incorporation of endothelial (CD31+) cells in mouse matrigel-plug assay (6±1.7% v CD34+ cells, 2±0.8%, p<0.01). Finally, in a mouse model of hind limb ischemia (HLI), CD34+ Exo significantly improved perfusion (ratio: 1.01±0.04 v 0.57±0.1, P<0.05), increased capillary density (1.8±0.3/HPF v 0.9±0.1/HPF, p<0.001) and prevented ischemic leg amputation (16% v 100%), as compared with MNC Exo. Conclusions: These data demonstrate that CD34+ Exo induce angiogenic activity and ischemic tissue repair in the absence of CD34+ cells, and suggest that Exo represent important mediators of the therapeutic effects associated with CD34+ cell therapy. We speculate that Exo derived from CD34+ cells may represent a significant component of the paracrine effect of progenitor-cell transplantation for therapeutic angiogenesis.

scholarly journals In Vitro Effects of Propofol on Gravid Human Myometrium

2008 ◽  
Vol 36 (6) ◽  
pp. 802-806 ◽  
Author(s):  
A. S. Thind ◽  
R. J. Turner
Keyword(s):  
Isometric Tension ◽  
Human Myometrium ◽  
Organ Bath ◽  
Dependent Manner ◽  
Uterine Contractility ◽  
Time Curve ◽  
Lower Segment Caesarean Section ◽  
Uterine Muscle ◽  
In Vitro Effects

The aim of this study was to evaluate the direct effect of propofol (di-isopropyl phenol) on the contractile properties of gravid human uterine muscle. Six specimens of uterine muscle were obtained from term parturients undergoing elective lower segment caesarean section. Small strips (1 × 2 x 12 mm) of muscle were prepared and suspended in an organ bath containing oxygenated Kreb's solution at 36.5°C. Following preparation, spontaneous regular contractions developed at a rate of one contraction every six to 10 minutes. Force of contraction was recorded continuously using an isometric tension transducer. Following baseline measurements, propofol was introduced into the bath at concentrations corresponding to 2 /μg/ml, 5 /μg/ml and 8 /μg/ml. The specimens were also exposed to intralipid in concentrations equivalent to that found in the 8 μ/ml solution of propofol to determine whether this additive influenced uterine contractility. Contractility (defined as area under the tension/time curve) was decreased to 89 ± 6.5% of control at 2 μg/ml 53±4.3% at 5 μ/ml and 45 ± 4.1% at 8 μg/ml. This decrease in contractility was statistically significant at concentrations >2 μg/ml. Intralipid did not significantly affect uterine contractility. The results of this study show that propofol decreases isolated human uterine muscle contractility in a dose-dependent manner.

scholarly journals Effect of the CCL5-Releasing Fibrin Gel for Intervertebral Disc Regeneration

Cartilage ◽  
2018 ◽  
Vol 11 (2) ◽  
pp. 169-180 ◽  
Author(s):  
Zhiyu Zhou ◽  
Stephan Zeiter ◽  
Tanja Schmid ◽  
Daisuke Sakai ◽  
James C. Iatridis ◽  
...  
Keyword(s):  
Animal Study ◽  
Culture Media ◽  
Histopathological Examination ◽  
In Vitro Release ◽  
Dependent Manner ◽  
Fibrin Gel ◽  
Defect Sites ◽  
Intervertebral Disc Regeneration ◽  
Chemotactic Effect

Objective To explore if chemokine (C-C motif) ligand 5 (CCL5) delivery could recruit annulus fibrosus (AF) cells to the injury sites and facilitate the repair of ruptured AF. Design The effects of CCL5 on bovine AF cells in vitro were tested by transwell assay and quantitative real-time polymerase chain reaction. Fibrin gel containing CCL5 was used to treat annulotomized bovine caudal discs cultured under dynamic loading conditions. After 14 days of loading, the samples were collected for histological examination. A pilot animal study was performed using sheep cervical discs to investigate the effect of fibrin gel encapsulated with CCL5 for the treatment of ruptured AF. After 14 weeks, the animals were sacrificed, and the discs were scanned with magnetic resonance imaging before histopathological examination. Results CCL5 showed a chemotactic effect on AF cells in a dose-dependent manner. AF cells cultured with CCL5 in vitro did not show any change of the gene expression of CCL5 receptors, catabolic and proinflammatory markers. In vitro release study showed that CCL5 exhibited sustained release from the fibrin gel into the culture media; however, in the organ culture study CCL5 did not stimulate homing of AF cells toward the defect sites. The pilot animal study did not show any repair effect of CCL5. Conclusions CCL5 has a chemotactic effect on AF cells in vitro, but no ex vivo or in vivo regenerative effect when delivered within fibrin gel. Further study with a stronger chemotactic agent and/or an alternate biomaterial that is more conductive of cell migration is warranted.

scholarly journals Different effect of benzylacyclouridine on the toxic and therapeutic effects of azidothymidine in mice

Blood ◽  
1990 ◽  
Vol 76 (11) ◽  
pp. 2216-2221 ◽  
Author(s):  
A Falcone ◽  
JW Darnowski ◽  
RM Ruprecht ◽  
SH Chu ◽  
I Brunetti ◽  
...  
Keyword(s):  
Leukemia Virus ◽  
Hematologic Toxicity ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Aids Related Complex ◽  
Effective Doses ◽  
Acquired Immunodeficiency ◽  
Immunodeficiency Syndrome ◽  
Oral Doses

Abstract It has been reported that in vitro uridine (Urd) can reverse azidothymidine (AZT) cytotoxicity without decreasing anti-human immunodeficiency virus (HIV) activity. Our studies in mice have shown that daily oral doses of benzylacyclouridine (BAU), an inhibitor of Urd breakdown, also reduces AZT hematologic toxicity, presumably by elevating the plasma concentration of Urd. We now extend these murine studies and report the effect of various doses of exogenous Urd, various doses of BAU, or the combination of BAU and Urd, administered daily, on AZT-induced toxicity. In mice receiving concomitant AZT, daily doses of Urd of 1,000 to 2,000 mg/kg increase peripheral reticulocytes and slightly reduce AZT-induced hematologic toxicity. However, the range of effective doses is narrow, and higher doses of Urd (greater than 3,000 mg/kg/d) significantly enhance hematologic toxicity. At its most effective dose, (2,000 mg/kg/d), Urd produces 28% mortality. In contrast, BAU doses up to 300 mg/kg/d reduced AZT-related hematologic toxicity in a dose-dependent manner without mortality. Higher daily doses of BAU and the combination of BAU with low doses of Urd were not more effective. Studies conducted in mice infected with the Rauscher murine leukemia virus (RLV) indicate that BAU does not impair the antiretroviral effect of AZT when administered at doses that reduce AZT-induced anemia and leukopenia. These findings may be significant for the treatment of patients with acquired immunodeficiency syndrome (AIDS) and AIDS-related complex.

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