Application of calcium phosphates and fibronectin as complementary treatment for osteoporotic bone fractures.

INTRODUCTION: The gradual aging of the population results in increased incidence of osteoporotic bone fractures. In a good quality bone, the fixation with the usual methods is adequate, but not in osteoporotic bone, in which consolidation delays and other complications are common, with failure rates for screws up to 25%. OBJECTIVE: To test fibronectin loaded hydroxyapatite as a complementary treatment for osteoporotic fractures. MATERIAL AND METHODS: This study was performed in a vivo model; 42 female osteoporotic adult rabbits 4-5kg (White New Zealand) were used. Two groups (hydroxyapatite and fibronectin loaded hydroxyapatite) and a control group were tested. 3 time points 24h, 48h and 5days were studied. Defects were created in both femurs, in one of them, a cannulated screw (4mm) and a biocompatible material were placed; in the other femur a screw was inserted without supplemented material forming the control group. Osteoporosis was induced from models already known throughout administration of steroids. Samples were analyzed histologically and through imaging (micro Ct). RESULTS: Basal levels of BMD are observed below to normal when compared to other studies (0.25/0.3 instead of 0.4). Global and dependent of time analysis of samples, show no significant differences for samples analyzed. However, an important trend was noted for variables that define the trabecular bone microarchitecture. Indices that define trabecular microarchitecture in the comparative analysis found to have statistical differences (p<0.01). DISCUSSION: Osteosynthesis in an osteoporotic bone is a challenge for the surgeon, due to a reduced bone mineral density and different bone architecture. The main finding was the verification of the hypothesis that the trabecular bone parameters increases with our augmentation material in weak rabbit bone quality. Also, the histological analyses of samples show an increase of non inflammatory cells in protein samples (OHAp-Fn) from the first 24hours. CONCLUSION: An early response of rabbit osteroporotic bone to a complementary treatment with fibronectin loaded hydroxyapatite has been observed. This response is reflected in greater values for indices that define the trabecular bone microarchitecture, thickness and separation, a greater non-inflammatory cellularity after only 24hours and an increased amount of connective tissue observed at 48hours.

Time-course investigation of the gene expression profile during Fasciola hepatica infection: A microarray-based study.

Fasciolosis is listed as one of the most important neglected tropical diseases according with the World Health Organization and is also considered as a reemerging disease in the human beings. Despite there are several studies describing the immune response induced by Fasciola hepatica in the mammalian host, investigations aimed at identifying the expression profile of genes involved in inducing hepatic injury are currently scarce. Data presented here belong to a time-course investigation of the gene expression profile in the liver of BALB/c mice infected with F. hepatica metacercariae at 7 and 21 days after experimental infection. The data published here have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO Series accession number GSE69588, previously published by Rojas-Caraballo et al. (2015) in PLoS One [1].

Gene Expression Profile in the Liver of BALB/c Mice Infected with Fasciola hepatica.

BACKGROUND: Fasciola hepatica infection still remains one of the helminthic neglected tropical diseases (NTDs). It has a huge worldwide distribution, affecting mainly cattle and, sometimes, human beings. In addition to data reported about the immunological response induced by helminthic infections and that induced by Fasciola hepatica, little is known about the gene expression profile in its organ target, the liver, which is where adult worms are established and live for long periods of time, causing its characteristic pathology. In the present work, we study both the early and late gene expression profiles in the livers of mice infected with F. hepatica metacercariae using a microarray-based methodology. METHODOLOGY: A total of 9 female-6-week-old BALB/c mice (Charles River Laboratories, Barcelona, Spain) weighing 20 to 35 g were used for the experiments. Two groups of BALB/c mice were orally infected with seven F. hepatica metacercariae, and the other group remained untreated and served as a control. Mice were humanely euthanized and necropsied for liver recovery, histological assessment of hepatic damage, RNA isolation, microarray design and gene expression analysis on the day of infection (t0), seven days post-infection (t7) and twenty-one days post-infection (t21). RESULTS: We found that F. hepatica infection induces the differential expression of 128 genes in the liver in the early stage of infection and 308 genes in the late stage, and most of them are up-regulated. The Ingenuity Pathway Analysis revealed significant changes in the pathways related to metabolism, biosynthesis and signaling as well as genes implicated in inducing liver-toxicity, injury and death. CONCLUSION: The present study provides us insights at the molecular level about the underlying mechanisms used by F. hepatica, leading to liver damage and its subsequent pathophysiology. The expression pattern obtained here could also be used to explain the lack of association between infection with F. hepatica and cholangiocarcinoma. However, more studies should be performed to confirm this hypothesis.

Evaluation of N-acetylcysteine treatment in acute pancreatitis-induced lung injury.

OBJECTIVE: Pulmonary complications are frequent during acute pancreatitis (AP). We investigate the effects of N-acetylcysteine (NAC) on lung injury in mild and severe AP. ANIMALS AND TREATMENT: Mild and severe AP was induced in rats by bile-pancreatic duct obstruction (BPDO) and infusion of 3.5 % sodium taurocholate (NaTc) into the bile-pancreatic duct, respectively. NAC (50 mg/kg) was given 1 h before and 1 h after AP. METHODS: Amylase activity was measured in plasma. Lungs were harvested for mRNA expression analysis of monocyte chemoattractant protein-1 (MCP-1), cytokine-induced neutrophil chemoattractant (CINC), P-selectin and intercellular adhesion molecule-1 (ICAM-1), myeloperoxidase (MPO) activity and histological examination. RESULTS: Hyperamylasemia was reduced by NAC in both AP models. NAC down-regulated MCP-1, CINC and P-selectin in BPDO- but not in NaTc-induced AP. Pulmonary insults did not vary in mild AP and were exacerbated in severe AP by NAC treatment. NAC reduced lung MPO activity in mild but not in severe AP. CONCLUSIONS: Although NAC treatment down-regulated inflammatory mediators in lungs during AP it did not prevent leukocyte infiltration, which could be responsible for maintaining the lung injury. As a result, NAC aggravated the lung damage in severe AP and failed to exert beneficial effects in the mild disease model.

Novel anti-inflammatory action of edelfosine lacking toxicity with protective effect in experimental colitis.

Edelfosine (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine; ET-18-OCH(3)) is an antitumor alkyl-lysophospholipid analog that binds lipid rafts, altering their protein composition (J Exp Med 200:353-365). Because L-selectin locates in lipid rafts and plays a crucial role in the recruitment of leukocytes into inflamed tissues, we hypothesized that edelfosine might affect inflammation by modulating L-selectin and inflammatory cell migration. Here, we have found that edelfosine inhibited neutrophil-endothelium interaction through L-selectin shedding. Oral treatment of edelfosine diminished inflammation in two murine animal models. Edelfosine showed a higher antiinflammatory effect than the nonsteroidal anti-inflammatory drug (NSAID) indomethacin in the bentonite mouse-paw edema model. Using a rat model of experimental colitis, edelfosine oral administration ameliorated the clinical and histopathologic severity of the inflammatory colitis with a dramatic decrease in mucosal damage and neutrophil infiltration. Colon sections from edelfosine-treated rats showed a remarkable reduction in ulcer formation, edema, and inflammatory cell infiltration. Edelfosine enhanced lipopolysaccharide-induced expression of anti-inflammatory interleukin-10 in mouse macrophages. Edelfosine oral treatment in rats, at doses 8-fold higher than those displaying anti-inflammatory action, lacked toxicity. Edelfosine treatment showed no any significant cardiotoxicity, hepatotoxicity or renal toxicity. Unlike NSAIDs, edelfosine did not inhibit prostaglandin E(2) synthesis in gastrointestinal mucosal biopsies, and no histologic alteration in gastrointestinal tract was detected after drug treatment. Thus, edelfosine shows a potent in vitro and in vivo anti-inflammatory activity while sparing gastric mucosa. Our data identify edelfosine as a novel anti-inflammatory drug by abating neutrophil infiltration through L-selectin shedding and may provide a new therapeutic approach for inflammatory bowel disease free from toxicity.

Injectable acrylic bone cements for vertebroplasty based on a radiopaque hydroxyapatite. Bioactivity and biocompatibility.

Radiopaque bone cements have been formulated to provide injectable pastes with improved bioactivity to be applied in vertebroplasty and kyphoplasty techniques. The bioactive compound was strontium containing hydroxyapatite salt, which was introduced as obtained (SrHA) or after treatment with MMA monomer (SrHA-t). The in vitro bioactivity of the cements was tested in cement films or in cement pastes introduced directly in a simulated body fluid (SBF) solution at 37 degrees C to mimic the in vivo conditions. Precipitation of an apatite-like layer was observed for the 20 wt %-SrHA-t containing cement in the first experiments, and in all formulations in the second ones. The deposited particles were characterized by FTIR spectroscopy and by EDAX analysis. Radiopacity of cements after immersion in SBF was confirmed. The biocompatibility exhibited by the SrHA containing cements was, in some cases, superior to that shown by a formulation with 10 wt % of BaSO(4). The new formulations prepared with the treated filler exhibited the lowest cytotoxicity and enhanced cellular proliferation. The in vivo biocompatibility tested by an intramuscular model in rats indicated the formation of a membrane formed by collagen fibers containing fibroblasts with no inflammatory cells, such as macrophages, giant cells or lymphocytes in all formulations.

Self-curing acrylic formulations with applications in intervertebral disk restoration: drug release and biological behaviour.

New injectable acrylic formulations have been prepared to be applied in restoration processes for intervertebral disks (IVDs). The solid phase of the formulations is composed of poly(methyl methacrylate) (PMMA), incorporating in some cases chondroitin sulphate (CS) as a regenerative bioactive molecule, whereas the liquid phase is constituted by an amphiphilic macromonomer (MT), 2-hydroxyethyl methacrylate (HEMA) and, in some formulations, acrylic acid (AA). The curing parameters and the mechanical properties of the IVD formulations make them excellent candidates for intervertebral application. In vitro and in vivo evaluation of the prepared IVD formulations is described in terms of CS release, surface analysis after immersion in SBF solutions, and biocompatibility studies based on MTT assay and Alamar blue test, as well as in vivo implantation in female Wistar rats, by injection of the IVD formulations followed by histological evaluations to assess tissue response.

Preparation of acrylic bone cements for vertebroplasty with bismuth salicylate as radiopaque agent.

One of the problems of percutaneous vertebroplasty attributed to the use of acrylic cements is related to the radiopacity of the formulation. The use of bismuth salicylate as the radiopaque agent is proposed in this work, taking into account the high radiopacity of organobismuth compounds used in dental applications and the possible analgesic effect of salicylic acid. Various cements formulated with this compound (some of them modified with polyethylene oxide) were examined. Setting parameters, mechanical properties, rheological behaviour, injectability, radiopacity and biocompatibility were studied for a variety of formulations, showing that the cement formulations containing bismuth salicylate have a higher radiopacity and better injection properties than commercial bone cement preparations and present good mechanical properties.

Injectable and self-curing composites of acrylic/bioactive glass and drug systems. A histomorphometric analysis of the behaviour in rabbits.

Injectable self-curing systems based on PMMA, phosphate-free bioactive glasses and the drug fosfosal, a phosphate derivative of salicylic acid with analgesic and moderate anti-inflammatory properties, have been tested in vivo to evaluate their biocompatibility. The model consisted of the injection of dough of cement into a defect created in the femur of rabbits, and the cure of the cement in situ after implantation. The biological response was studied in the short and long terms by macroscopic, radiological and histopathological examination, and quantitatively by histomorphometric and statistical analysis considering the most representative variables at the bone-cement interface: cement, bone marrow, newly formed bone and connective tissue. All bioactive formulations presented resorption of the cement at the end of the experiment in contrast to the control of PMMA, due to the presence of resorbable components. The presence or absence of the phosphate group added by the drug fosfosal influenced mainly on the new bone formation process. The cement formulated with bioactive glasses and in absence of fosfosal produced the maximum amount of neoformed bone at 2 weeks, and then it resorbed at 4 weeks to give a higher amount of neoformed bone at the end of the experiment, compared with the formulation containing only fosfosal. The presence of fosfosal and bioactive glass together affected the ossification process strongly. The osseous tissue was produced more gradually but it continuously increased giving rise to a more stable bone at the end of the experiment.

Relationships between NOS2 and HO-1 in liver of rats with chronic bile duct ligation.

An increased expression and activity of the heme oxygenase-1 (HO-1) in the liver has been observed in models of hepatic damage. Nitric oxide (NO) seems to be involved in HO-1 regulation. The aim of this work is to assess HO-1 induction and heme oxygenase (HO) activity in rats with bile duct ligation (BDL). We have assessed the effect of chronic inhibition of the NO synthesis by N(G)-nitro-l-arginine methyl ester (l-NAME) on HO-1 induction and HO activity. In the BDL animals, compared with sham-operated ones, we found an increased plasma nitrite and bilirubin concentration, and a marked liver expression of inducible nitric oxide synthase and HO-1, assessed by both Western blot and immunohistochemistry. Chronic l-NAME treatment prevented plasma nitrite increase in animals subjected to BDL. BDL animals treated with l-NAME, compared with untreated BDL rats, showed an important decrease in HO-1 expression and in HO activity (assessed as a decreased plasma bilirubin and bilirubin excretion). In conclusion, our experiments show parallel changes in expression and activity of HO-1 and NOS2 activity in the BDL model of liver damage and suggest that increased NO production is involved in HO-1 overexpression.

In vivo inhibition of inducible nitric oxide synthase decreases lung injury induced by Toxocara canis in experimentally infected rats.

The direct effect of nitric oxide (NO) on the viability of Toxocara canis larvae was studied. We observed that the nitric oxide donors, SIN-1 and SNOG, exert no cytotoxic effect on the in vitro viability of T. canis larvae. In addition, we developed a model in rats to elucidate the role of NO during T. canis infection. We evaluated different indicators in four experimental groups: morphological parameters, the total number cells and cell types recovered, nitrite and protein concentration, lactate dehydrogenase and alkaline phosphatase enzymatic activity in the bronchoalveolar lavage fluid, lung index and detection of anti-T. canis specific antibodies. We observed significant differences between non-infected and infected groups. The infected animals treated with the inducible nitric oxide synthase (iNOS) inhibitor aminoguanidine were less damaged than infected, non-treated animals. Our results suggest that the in vivo inhibition of the synthesis of NO triggered by iNOS diminishes the deleterious effects of the parasite upon the host, especially the vascular alterations in the lungs. We could show that in vivo production of NO induced by infection with T. canis results in direct host damage. Thus, this induction may constitute an evasion/adaptation mechanism of the parasite.