Publications by authors named "Gabriela Fragoso"

22 Publications

  • Page 1 of 1

Improvement of colonic healing and surgical recovery with perioperative supplementation of inulin and galacto-oligosaccharides.

Clin Nutr 2021 Jun 27;40(6):3842-3851. Epub 2021 Apr 27.

Nutrition and Microbiome Laboratory, Institut du Cancer de Montréal, Centre de recherche du Centre hospitalier de l'Université de Montréal (CRCHUM), 900 Rue Saint-Denis, Montréal, Québec, H2X 0A9, Canada; Department of Medicine, Université de Montréal, 2900 Boulevard Edouard-Montpetit, Montréal, Québec, H3T 1J4, Canada. Electronic address:

Background And Aims: Anastomotic leak (AL) is a major complication in colorectal surgery. Recent evidence suggests that the gut microbiota may affect healing and may cause or prevent AL. Butyrate is a beneficial short-chain fatty acid (SCFA) that is produced as a result of bacterial fermentation of dietary oligosaccharides and has been described as beneficial in the maintenance of colonic health. To assess the impact of oligosaccharides on colonic anastomotic healing in mice, we propose to modulate the microbiota with oligosaccharides to increase butyrate production via enhancement of butyrate-producing bacteria and, consequently, improve anastomotic healing in mice.

Methods: Animal experiments were conducted in mice that were subjected to diets supplemented with inulin, galacto-oligosaccharides (GOS) or cellulose, as a control, for two weeks before undergoing a surgical colonic anastomosis. Macroscopic and histological assessment of the anastomosis was performed. Extent of epithelial proliferation was assessed by Ki-67 immunohistochemistry. Gelatin zymography was used to evaluate the extent of matrix metalloproteinase (MMP) hydrolytic activity.

Results: Inulin and GOS diets were associated with increased butyrate production and better anastomotic healing. Histological analysis revealed an enhanced mucosal continuity, and this was associated with an increased re-epithelialization of the wound as determined by increased epithelial proliferation. Collagen concentration in peri-anastomotic tissue was higher with inulin and GOS diets and MMP activity, a marker of collagen degradation, was lower with both oligosaccharides. Inulin and GOS diets were further associated with lower bacterial translocation.

Conclusions: Dietary supplementation with inulin and GOS may improve anastomotic healing and reinforce the gut barrier in mice.
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http://dx.doi.org/10.1016/j.clnu.2021.04.032DOI Listing
June 2021

Oligosaccharides increase the genotoxic effect of colibactin produced by pks+ Escherichia coli strains.

BMC Cancer 2021 Feb 17;21(1):172. Epub 2021 Feb 17.

Nutrition and Microbiome Laboratory, Institut du cancer de Montréal, Centre de recherche du Centre hospitalier de l'Université de Montréal (CRCHUM), 900 Rue Saint Denis, Montreal, QC, H2X 0A9, Canada.

Background: Colibactin is a genotoxin that induces DNA double-strand breaks that may lead to carcinogenesis and is produced by Escherichia coli strains harboring the pks island. Human and animal studies have shown that colibactin-producing gut bacteria promote carcinogenesis and enhance the progression of colorectal cancer through cellular senescence and chromosomal abnormalities. In this study, we investigated the impact of prebiotics on the genotoxicity of colibactin-producing E. coli strains Nissle 1917 and NC101.

Methods: Bacteria were grown in medium supplemented with 20, 30 and 40 mg/mL of prebiotics inulin or galacto-oligosaccharide, and with or without 5 μM, 25 μM and 125 μM of ferrous sulfate. Colibactin expression was assessed by luciferase reporter assay for the clbA gene, essential for colibactin production, in E. coli Nissle 1917 and by RT-PCR in E. coli NC101. The human epithelial colorectal adenocarcinoma cell line, Caco-2, was used to assess colibactin-induced megalocytosis by methylene blue binding assay and genotoxicity by γ-H2AX immunofluorescence analysis.

Results: Inulin and galacto-oligosaccharide enhanced the expression of clbA in pks+ E. coli. However, the addition of 125 μM of ferrous sulfate inhibited the expression of clbA triggered by oligosaccharides. In the presence of either oligosaccharide, E. coli NC101 increased dysplasia and DNA double-strand breaks in Caco-2 cells compared to untreated cells.

Conclusion: Our results suggest that, in vitro, prebiotic oligosaccharides exacerbate DNA damage induced by colibactin-producing bacteria. Further studies are necessary to establish whether oligosaccharide supplementation may lead to increased colorectal tumorigenesis in animal models colonized with pks+ E. coli.
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http://dx.doi.org/10.1186/s12885-021-07876-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7890614PMC
February 2021

Acute invariant NKT cell activation triggers an immune response that drives prominent changes in iron homeostasis.

Sci Rep 2020 12 3;10(1):21026. Epub 2020 Dec 3.

Centre de Recherche du Centre Hospitalier de l'Université de Montréal (CRCHUM), Montréal, Québec, Canada.

Iron homeostasis is an essential biological process that ensures the tissue distribution of iron for various cellular processes. As the major producer of hepcidin, the liver is central to the regulation of iron metabolism. The liver is also home to many immune cells, which upon activation may greatly impact iron metabolism. Here, we focus on the role of invariant natural killer T (iNKT) cells, a subset of T lymphocytes that, in mice, is most abundant in the liver. Activation of iNKT cells with the prototypical glycosphingolipid antigen, α-galactosylceramide, resulted in immune cell proliferation and biphasic changes in iron metabolism. This involved an early phase characterized by hypoferremia, hepcidin induction and ferroportin suppression, and a second phase associated with strong suppression of hepcidin despite elevated levels of circulating and tissue iron. We further show that these changes in iron metabolism are fully dependent on iNKT cell activation. Finally, we demonstrate that the biphasic regulation of hepcidin is independent of NK and Kupffer cells, and is initially driven by the STAT3 inflammatory pathway, whereas the second phase is regulated by repression of the BMP/SMAD signaling pathway. These findings indicate that iNKT activation and the resulting cell proliferation influence iron homeostasis.
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http://dx.doi.org/10.1038/s41598-020-78037-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7713400PMC
December 2020

Validation of the prognostic value of NF-κB p65 in prostate cancer: A retrospective study using a large multi-institutional cohort of the Canadian Prostate Cancer Biomarker Network.

PLoS Med 2019 07 2;16(7):e1002847. Epub 2019 Jul 2.

Centre de recherche du Centre hospitalier de l'Université de Montréal, Centre hospitalier de l'Université de Montréal, Montreal, Quebec, Canada.

Background: The identification of patients with high-risk prostate cancer (PC) is a major challenge for clinicians, and the improvement of current prognostic parameters is an unmet clinical need. We and others have identified an association between the nuclear localization of NF-κB p65 and biochemical recurrence (BCR) in PC in small and/or single-centre cohorts of patients.

Methods And Findings: In this study, we accessed 2 different multi-centre tissue microarrays (TMAs) representing cohorts of patients (Test-TMA and Validation-TMA series) of the Canadian Prostate Cancer Biomarker Network (CPCBN) to validate the association between p65 nuclear frequency and PC outcomes. Immunohistochemical staining of p65 was performed on the Test-TMA and Validation-TMA series, which include PC tissues from patients treated by first-line radical prostatectomy (n = 250 and n = 1,262, respectively). Two independent observers evaluated the p65 nuclear frequency in digital images of cancer tissue and benign adjacent gland tissue. Kaplan-Meier curves coupled with a log-rank test and univariate and multivariate Cox regression models were used for statistical analyses of continuous values and dichotomized data (cutoff of 3%). Multivariate analysis of the Validation-TMA cohort showed that p65 nuclear frequency in cancer cells was an independent predictor of BCR using continuous (hazard ratio [HR] 1.02 [95% CI 1.00-1.03], p = 0.004) and dichotomized data (HR 1.33 [95% CI 1.09-1.62], p = 0.005). Using a cutoff of 3%, we found that this biomarker was also associated with the development of bone metastases (HR 1.82 [95% CI 1.05-3.16], p = 0.033) and PC-specific mortality (HR 2.63 [95% CI 1.30-5.31], p = 0.004), independent of clinical parameters. BCR-free survival, bone-metastasis-free survival, and PC-specific survival were shorter for patients with higher p65 nuclear frequency (p < 0.005). As the small cores on TMAs are a limitation of the study, a backward validation of whole PC tissue section will be necessary for the implementation of p65 nuclear frequency as a PC biomarker in the clinical workflow.

Conclusions: We report the first study using the pan-Canadian multi-centre cohorts of CPCBN and validate the association between increased frequency of nuclear p65 frequency and a risk of disease progression.
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http://dx.doi.org/10.1371/journal.pmed.1002847DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6605640PMC
July 2019

Curcumin induces mild anemia in a DSS-induced colitis mouse model maintained on an iron-sufficient diet.

PLoS One 2019 26;14(4):e0208677. Epub 2019 Apr 26.

Nutrition and Microbiome Laboratory, Centre de recherche du Centre hospitalier de l'Université de Montréal (CRCHUM), Institut du cancer de Montréal, Montréal, Québec, Canada.

Anemia is frequently encountered in patients with inflammatory bowel disease (IBD), decreasing the quality of life and significantly worsening the prognosis of the disease. The pathogenesis of anemia in IBD is multifactorial and results mainly from intestinal blood loss in inflamed mucosa and impaired dietary iron absorption. Multiple studies have proposed the use of the polyphenolic compound curcumin to counteract IBD pathogenesis since it has significant preventive and therapeutic properties as an anti-inflammatory agent and very low toxicity, even at high dosages. However, curcumin has been shown to possess properties consistent with those of an iron-chelator, such as the ability to modulate proteins of iron metabolism and decrease spleen and liver iron content. Thus, this property may further contribute to the development and severity of anemia of inflammation and iron deficiency in IBD. Herein, we evaluate the effects of curcumin on systemic iron balance in the dextran sodium sulfate (DSS) model of colitis in C57Bl/6 and BALB/c mouse strains that were fed an iron-sufficient diet. In these conditions, curcumin supplementation caused mild anemia, lowered iron stores, worsened colitis and significantly decreased overall survival, independent of the mouse strain. These findings suggest that curcumin usage as an anti-inflammatory supplement should be accompanied by monitoring of erythroid parameters to avoid exacerbation of iron deficiency anemia in IBD.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0208677PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6485613PMC
January 2020

MyD88 Adaptor Protein Is Required for Appropriate Hepcidin Induction in Response to Dietary Iron Overload in Mice.

Front Physiol 2018 5;9:159. Epub 2018 Mar 5.

Nutrition and Microbiome Laboratory, Centre de Recherche du Centre Hospitalier de l'Université de Montréal, Montréal, QC, Canada.

Iron homeostasis is tightly regulated to provide virtually all cells in the body, particularly red blood cells, with this essential element while defending against its toxicity. The peptide hormone hepcidin is central to the control of the amount of iron absorbed from the diet and iron recycling from macrophages. Previously, we have shown that hepcidin induction in macrophages following Toll-like receptor (TLR) stimulation depends on the presence of myeloid differentiation primary response gene 88 (MyD88). In this study, we analyzed the regulation of iron metabolism in mice to further investigate MyD88 involvement in iron sensing and hepcidin induction. We show that mice lacking MyD88 accumulate significantly more iron in their livers than wild-type counterparts in response to dietary iron loading as they are unable to appropriately control hepcidin levels. The defect was associated with inappropriately low levels of Smad4 protein and Smad1/5/8 phosphorylation in liver samples found in the mice compared to wild-type mice. In conclusion, our results reveal a previously unknown link between MyD88 and iron homeostasis, and provide new insights into the regulation of hepcidin through the iron-sensing pathway.
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http://dx.doi.org/10.3389/fphys.2018.00159DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5845127PMC
March 2018

Dietary Heme Induces Gut Dysbiosis, Aggravates Colitis, and Potentiates the Development of Adenomas in Mice.

Front Microbiol 2017 21;8:1809. Epub 2017 Sep 21.

Département de Médecine, Université de Montréal, MontréalQC, Canada.

Dietary heme can be used by colonic bacteria equipped with heme-uptake systems as a growth factor and thereby impact on the microbial community structure. The impact of heme on the gut microbiota composition may be particularly pertinent in chronic inflammation such as in inflammatory bowel disease (IBD), where a strong association with gut dysbiosis has been consistently reported. In this study we investigated the influence of dietary heme on the gut microbiota and inferred metagenomic composition, and on chemically induced colitis and colitis-associated adenoma development in mice. Using 16S rRNA gene sequencing, we found that mice fed a diet supplemented with heme significantly altered their microbiota composition, characterized by a decrease in α-diversity, a reduction of and an increase of , particularly . These changes were similar to shifts seen in dextran sodium sulfate (DSS)-treated mice to induce colitis. In addition, dietary heme, but not systemically delivered heme, contributed to the exacerbation of DSS-induced colitis and facilitated adenoma formation in the azoxymethane/DSS colorectal cancer (CRC) mouse model. Using inferred metagenomics, we found that the microbiota alterations elicited by dietary heme resulted in non-beneficial functional shifts, which were also characteristic of DSS-induced colitis. Furthermore, a reduction in fecal butyrate levels was found in mice fed the heme supplemented diet compared to mice fed the control diet. Iron metabolism genes known to contribute to heme release from red blood cells, heme uptake, and heme exporter proteins, were significantly enriched, indicating a shift toward favoring the growth of bacteria able to uptake heme and protect against its toxicity. In conclusion, our data suggest that luminal heme, originating from dietary components or gastrointestinal bleeding in IBD and, to lesser extent in CRC, directly contributes to microbiota dysbiosis. Thus, luminal heme levels may further exacerbate colitis through the modulation of the gut microbiota and its metagenomic functional composition. Our data may have implications in the development of novel targets for therapeutic approaches aimed at lowering gastrointestinal heme levels through heme chelation or degradation using probiotics and nutritional interventions.
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http://dx.doi.org/10.3389/fmicb.2017.01809DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5613120PMC
September 2017

Iron Supplements Modulate Colon Microbiota Composition and Potentiate the Protective Effects of Probiotics in Dextran Sodium Sulfate-induced Colitis.

Inflamm Bowel Dis 2017 05;23(5):753-766

*Département de Médecine, Université de Montréal, Montréal, Canada; and†Centre de recherche, Centre hospitalier de l'Université de Montréal (CHUM), Institut du cancer de Montréal, and Université de Montréal, Montréal, Canada.

Background: Iron is an important nutrient for both the host and colonizing bacteria. Oral iron supplementation may impact the composition of the microbiota and can be particularly damaging to patients suffering from inflammatory bowel disease (IBD). However, patients with IBD may require iron supplementation to treat their anemia.

Methods: We fed mice with diets supplemented with ferrous sulfate at different doses (5, 50, and 500 mg of iron/kg chow) and with different iron formulations (ferrous sulfate, ferrous bisglycinate and ferric ethylenediaminetetraacetic acid [FEDTA]), and analyzed the effects on the composition of the gut microbiota by 16S ribosomal RNA gene sequencing. Using the dextran sodium sulfate (DSS)-induced colitis mouse model, we investigated the effects of iron supplementation in colitis severity, as well as the use of the probiotic Escherichia coli Nissle 1917 (EcN) in combination with iron supplementation.

Results: Iron supplementation at different doses induced shifts in the gut microbial communities and inferred metabolic pathways. However, depending on the iron formulation used in the diets, iron supplementation during dextran sodium sulfate-induced colitis was either beneficial (ferrous bisglycinate) or highly detrimental (FEDTA). Finally, the beneficial effect of the probiotic EcN in the dextran sodium sulfate-induced colitis model was potentiated by oral iron supplementation with ferrous sulfate.

Conclusions: These results show that the iron formulations used to treat iron deficiency influence the gut microbiota and colitis in mice and suggest that distinct iron compounds may be of particular relevance to patients with IBD. In addition, the beneficial action of probiotics in IBD may be enhanced by oral iron supplementation.
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http://dx.doi.org/10.1097/MIB.0000000000001089DOI Listing
May 2017

Mice are poor heme absorbers and do not require intestinal Hmox1 for dietary heme iron assimilation.

Haematologica 2015 Sep 14;100(9):e334-7. Epub 2015 May 14.

Department of Medicine, McGill University, and Lady Davis Institute for Medical Research, Jewish General Hospital, Montreal, Quebec, Canada

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http://dx.doi.org/10.3324/haematol.2015.126870DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4800685PMC
September 2015

Agonist-induced down-regulation of AMPA receptors in oligodendrocyte progenitors.

Neuropharmacology 2014 Apr 9;79:506-14. Epub 2014 Jan 9.

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada. Electronic address:

Prolonged exposure of oligodendrocyte progenitor cultures to non-toxic concentrations of glutamate receptor agonists for 24 h decreased cellular proliferation mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. Since prolonged agonist stimulation can regulate the expression of various families of receptors, we examined this possibility. Pretreatment of progenitor cultures with 100 μM kainic acid (KA) for 1-24 h caused a time-dependent decrease in AMPA receptor activity, determined by agonist-induced (45)Ca(2+) uptake. The maximum effect (70-80% decrease), observed in the 24 h-pretreated cells, was accompanied by a significant reduction in AMPA receptor subunits, as determined by Western blotting. GluR2/3 and GluR4 subunits were the most affected. Receptor down-regulation and (45)Ca(2+) uptake were only partially reversible upon KA removal. Furthermore, 24 h co-treatment of cultures with CNQX blocked the KA-induced decreases in calcium uptake. To address whether calpain, a calcium-activated protease, was implicated in the regulation of the AMPA receptor subunits, cultures were treated with the specific inhibitor PD150606 alone or in combination with KA for 24 h. Calpain inhibition significantly increased GluR1 in both conditions and partly reversed downregulation of GluR4 by KA. Collectively, these results indicate that calpain is not involved in the agonist-induced down-regulation of AMPA receptors subunits 2/3 in oligodendrocyte progenitors, while it downregulates GluR1 and GluR4.
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http://dx.doi.org/10.1016/j.neuropharm.2013.12.020DOI Listing
April 2014

Response of human oligodendrocyte progenitors to growth factors and axon signals.

J Neuropathol Exp Neurol 2010 Sep;69(9):930-44

Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.

We examined the effects of growth factors and axonal signals on the differentiation of human fetal and adult oligodendrocyte progenitor cells (OPCs) and determined whether these effects translated into enhanced axonal ensheathment. Only small numbers of fetal OPCs grown in defined medium expressed the oligodendroglial lineage markers Olig2 and O4. The combination of platelet-derived growth factor-AA and basic fibroblast growth factor enhanced proliferation of Olig2-positive and O4-positive cells; a combination of brain-derived neurotrophic factor and insulin-like growth factor 1 promoted O4-positive cell differentiation, galactocerebroside expression, and morphological complexity. Coculturing with rodent dorsal root ganglion neurons in defined medium alone enhanced OPC differentiation and myelin basic protein expression. The addition of brain-derived neurotrophic factor/insulin-like growth factor 1 further enhanced differentiation, axonal attachment and ensheathment, and clustering of the contactin-associated protein Caspr and Na+ channels. By contrast, most adult OPCs were O4 positive and Olig2 positive in defined medium; both brain-derived neurotrophic factor/insulin-like growth factor 1 and platelet-derived growth factor-AA/basic fibroblast growth factor promoted their myelin basic protein expression and membrane sheet formation; coculture with dorsal root ganglion neurons further increased myelin basic protein expression. Growth factors also enhanced attachment of adult OPCs to axons, but their capacity to ensheath axons was lower than that of fetal OPCs. These results demonstrate that fetal and adult OPCs show measurable responses to selected growth factors and axon signals that correlate with their capacity for axon ensheathment. The distinct properties of fetal and adult OPCs may be related to differences in their chronological age and initial differentiation states.
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http://dx.doi.org/10.1097/NEN.0b013e3181ef3be4DOI Listing
September 2010

Regulation of peripheral myelination by Src-like kinases.

Exp Neurol 2010 Nov 7;226(1):47-57. Epub 2010 Aug 7.

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.

Fyn, a nonreceptor Src-like tyrosine kinase (SLK), plays an important role in oligodendrocyte differentiation and myelination in the brain. However, its role in myelination of peripheral nerves remains undefined. Here we report that selective inhibitors of SLKs (PP2 and SU6656) caused a dose-dependent decrease in the accumulation of several myelin proteins, including myelin basic protein (MBP), protein zero (P0) and myelin-associated glycoprotein (MAG) in rat Schwann cell-dorsal root ganglion neuron (SC-DRGN) co-cultures. Interestingly, SLK inhibition was insufficient to completely abrogate myelin synthesis, as removal of PP2 after several days of treatment permitted a partial recovery of myelin proteins expression. Furthermore, fewer and shorter myelinated segments formed in the continuous presence of PP2, although the myelin formed was normally compacted. PP2 also decreased the number of SCs expressing Krox-20, a master-regulatory transcription factor expressed by myelinating SCs, by 50%. These results were corroborated by selective knockdown of Fyn and Lyn kinases using siRNA. Extracellular matrix is important to SC differentiation and peripheral myelination. Using phospho-specific antibodies, we showed that addition of extracellular matrix extracts to SC-DRGN co-cultures resulted in the activation of ERK, Akt and p38 MAPK, three protein kinases involved in SC proliferation, differentiation and peripheral myelination. PP2 blocked the phosphorylation of all three kinases. Our results support a role for SLKs in the initiation of peripheral myelination via the activation of p38, Akt and ERK, which regulate Krox-20 expression and peripheral myelination.
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http://dx.doi.org/10.1016/j.expneurol.2010.08.002DOI Listing
November 2010

Amyloid beta-induced nerve growth factor dysmetabolism in Alzheimer disease.

J Neuropathol Exp Neurol 2009 Aug;68(8):857-69

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.

We previously reported that the precursor form of nerve growth factor (pro-NGF) and not mature NGF is liberated in the CNS in an activity-dependent manner, and that its maturation and degradation occur in the extracellular space by the coordinated action of proteases.Here, we present evidence of diminished conversion of pro-NGF to its mature form and of greater NGF degradation in Alzheimer disease (AD) brain samples compared with controls. These alterations of the NGF metabolic pathway likely resulted in the increased pro-NGF levels. The pro-NGF was largely in a peroxynitrited form in the AD samples. Intrahippocampal injection of amyloid-beta oligomers provoked similar upregulation of pro-NGF in naive rats that was accompanied by evidence of microglial activation (CD40), increased levels of inducible nitric oxide synthase, and increased activity of the NGF-degrading enzyme matrix metalloproteinase 9. The elevated inducible nitric oxide synthase provoked the generation of biologically inactive, peroxynitrite-modified pro-NGF in amyloid-beta oligomer-injected rats. These parameters were corrected by minocycline treatment. Minocycline also diminished altered matrix metalloproteinase 9, inducible nitric oxide synthase, and microglial activation (CD40); improved cognitive behavior; and normalized pro-NGF levels in a transgenic mouse AD model. The effects of amyloid-beta amyloid CNS burden on NGF metabolism may explain the paradoxical upregulation of pro-NGF in AD accompanied by atrophy of forebrain cholinergic neurons.
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http://dx.doi.org/10.1097/NEN.0b013e3181aed9e6DOI Listing
August 2009

The QKI-6 and QKI-7 RNA binding proteins block proliferation and promote Schwann cell myelination.

PLoS One 2009 Jun 11;4(6):e5867. Epub 2009 Jun 11.

Terry Fox Molecular Oncology Group and the Bloomfield Center for Research on Aging, Lady Davis Institute for Medical Research, Sir Mortimer B Davis Jewish General Hospital, Department of Oncology and Medicine, McGill University, Montréal, Québec, Canada.

Background: The quaking viable (qk(v)) mice have uncompacted myelin in their central and peripheral nervous system (CNS, PNS). The qk gene encodes 3 major alternatively spliced isoforms that contain unique sequence at their C-terminus dictating their cellular localization. QKI-5 is a nuclear isoform, whereas QKI-6 and QKI-7 are cytoplasmic isoforms. The qk(v) mice harbor an enhancer/promoter deletion that prevents the expression of isoforms QKI-6 and QKI-7 in myelinating cells resulting in a dysmyelination phenotype. It was shown that QKI regulates the differentiation of oligodendrocytes, the myelinating cells of the CNS, however, little is known about the role of the QKI proteins, or RNA binding proteins in PNS myelination.

Methodology/principal Findings: To define the role of the QKI proteins in PNS myelination, we ectopically expressed QKI-6 and QKI-7 in primary rat Schwann cell/neuron from dorsal root ganglia cocultures. We show that the QKI isoforms blocked proliferation and promoted Schwann cell differentiation and myelination. In addition, these events were coordinated with elevated proteins levels of p27(KIP1) and myelin basic protein (MBP), markers of Schwann cell differentiation. QKI-6 and QKI-7 expressing co-cultures contained myelinated fibers that had directionality and contained significantly thicker myelin, as assessed by electron microscopy. Moreover, QKI-deficient Schwann cells had reduced levels of MBP, p27(KIP1) and Krox-20 mRNAs, as assessed by quantitative RT-PCR.

Conclusions/significance: Our findings suggest that the QKI-6 and QKI-7 RNA binding proteins are positive regulators of PNS myelination and show that the QKI RNA binding proteins play a key role in Schwann cell differentiation and myelination.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0005867PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2690695PMC
June 2009

p38 Mitogen-activated protein kinase regulates myelination.

J Mol Neurosci 2008 May 10;35(1):23-33. Epub 2007 Nov 10.

Department of Pharmacology and Therapeutics, McGill University, 3655 Promenade Sir William Osler, Montreal, QC H3G 1Y6, Canada.

The p38 mitogen-activated protein kinase family is emerging as a crucial signaling molecule for a vast number of cellular functions including cell migration, proliferation, and differentiation. The function of p38 in myelination has only been recently addressed. Using pyridinyl imidazole-based p38 alpha/beta selective inhibitors, we have reported a critical role for this kinase in the regulation of myelination, specifically, in controlling the differentiation of Schwann cells, and oligodendrocytes, the myelinating glia of the peripheral and central nervous systems, respectively. These compounds inhibited the accumulation of myelin-cell-specific markers, including myelin-specific glycosphingolipids, myelin-associated glycoprotein, and myelin basic protein. More significantly, myelination of dorsal root ganglia neurons by oligodendrocytes was irreversibly blocked by p38 inhibitors. Our current studies are focusing on the molecular mechanisms by which p38 regulates oligodendrocyte and Schwann cell differentiation and its role in models of myelination and remyelination.
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http://dx.doi.org/10.1007/s12031-007-9011-0DOI Listing
May 2008

p38 mitogen-activated protein kinase is required for central nervous system myelination.

Glia 2007 Nov;55(15):1531-41

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.

The p38 MAPKs are a family of kinases that regulate a number of cellular functions including cell migration, proliferation, and differentiation. Here, we report that p38 regulates oligodendrocyte differentiation. Inhibition of p38 with PD169316 and SB203580 prevented accumulation of protein and mRNA of cell-stage specific markers characteristic of differentiated oligodendrocytes, including myelin basic protein, myelin-associated glycoprotein, and the glycosphingolipids, galactosylceramide and sulfatide. In addition, the cell cycle regulator p27(kip1) and the transcription factor Sox10 were also significantly reduced. Most significantly, p38 inhibitors completely and irreversibly blocked myelination of dorsal root ganglion neurons by oligodendrocytes and prevented the axolemmal organization of the axo-glial adhesion molecule Caspr. Our results suggest a role(s) for this kinase in key regulatory steps in the maturation of OLGs and initiation of myelination.
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http://dx.doi.org/10.1002/glia.20567DOI Listing
November 2007

Human coronavirus OC43 infection induces chronic encephalitis leading to disabilities in BALB/C mice.

Virology 2006 Jun 9;349(2):335-46. Epub 2006 Mar 9.

Laboratory of Neuroimmunovirology, INRS-Institut Armand-Frappier, 531 Boulevard des Prairies, Laval, Québec, Canada H7V 1B7.

The notion that an infectious respiratory pathogen can damage the central nervous system (CNS) and lead to neurological disease was tested using a human respiratory coronavirus, the OC43 strain of human coronavirus (HCoV-OC43). First, primary cell cultures were used to determine the susceptibility of each type of neural cells to virus infection. Neurons were the target cells, undergoing degeneration during infection, in part due to apoptosis. Second, neuropathogenicity was investigated in susceptible mice. Intracerebral inoculation of HCoV-OC43 into BALB/c mice led to an acute encephalitis with neuronal cell death by necrosis and apoptosis. Infectious virus was apparently cleared from surviving animals, whereas viral RNA persisted for several months. Some of the animals surviving to acute encephalitis presented an abnormal limb clasping reflex and a decrease in motor activity starting several months post-infection. These results suggest that viral persistence could be associated with an increased neuronal degeneration leading to neuropathology and motor deficits in susceptible individuals.
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http://dx.doi.org/10.1016/j.virol.2006.01.049DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7111850PMC
June 2006

Developmental differences in HO-induced oligodendrocyte cell death: role of glutathione, mitogen-activated protein kinases and caspase 3.

J Neurochem 2004 Jul;90(2):392-404

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.

The molecular mechanisms underlying H(2)O(2)-induced toxicity were characterized in rat oligodendrocyte cultures. While progenitor cells were more sensitive than mature oligodendrocytes to H(2)O(2), the antioxidant, N-acetyl-L-cysteine, blocked toxicity at both stages of development. Differentiated oligodendrocytes contained more glutathione than did progenitors and were less susceptible to decreases in glutathione concentration induced by H(2)O(2) stress. As free radicals have been considered to serve as second messengers, we examined the effect of H(2)O(2) on activation of the mitogen-activated protein kinases (MAPK), extracellular signal-regulated kinases (ERK) 1/2 and p38. H(2)O(2) caused a time- and concentration-dependent increase in MAPK phosphorylation, an effect that was totally blocked by N-acetyl-L-cysteine. Further exploration of potential mechanisms involved in oligodendrocyte cell death showed that H(2)O(2) treatment caused DNA condensation and fragmentation at both stages of development, whereas caspase 3 activation and poly (ADP-ribose) polymerase cleavage were significantly increased only in oligodendrocyte progenitors. The pan-caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp fluoromethyl ketone, blocked DNA fragmentation in progenitors and produced a small but significant level of protection from H(2)O(2) toxicity in progenitors and mature oligodendrocytes. In contrast, inhibitors of both p38 and MEK reduced H(2)O(2)-induced death most significantly in oligodendrocytes. The poly (ADP-ribose) polymerase inhibitor, PJ34, reduced H(2)O(2)-induced toxicity on its own but was most effective when combined with benzyloxycarbonyl-Val-Ala-Asp fluoromethyl ketone or PD169316. The finding that molecular mechanisms conferring resistance to reactive oxygen species toxicity are regulated during oligodendrocyte differentiation may be of importance in designing therapies for certain neurological diseases affecting white matter.
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http://dx.doi.org/10.1111/j.1471-4159.2004.02488.xDOI Listing
July 2004

Cytotoxicity of the E(2)-isoprostane 15-E(2t)-IsoP on oligodendrocyte progenitors.

Free Radic Biol Med 2004 Aug;37(3):358-66

Research Center of Hôpital Sainte-Justine, Department of Pediatrics and Pharmacology, Université de Montréal, Québec, Canada.

Oxidant stress plays a significant role in the pathogenesis of periventricular leukomalacia (PVL). Isoprostanes (IsoPs) are bioactive products of lipid peroxidation abundantly generated during hypoxic-ischemic injuries. Because loss of oligodendrocytes (OLs) occurs early in PVL, we hypothesized that IsoPs could induce progenitor OL death. 15-E(2t)-IsoP but not 15-F(2t)-IsoP elicited a concentration-dependent death of progenitor OLs by oncosis and not by apoptosis, but exerted minimal effects on mature OLs. 15-E(2t)-IsoP-induced cytotoxicity could not be explained by its conversion into cyclopentenones, because PGA(2) was hardly cytotoxic. On the other hand, thromboxane A(2) (TxA(2)) synthase inhibitor CGS12970 and cyclooxygenase inhibitor ibuprofen attenuated 15-E(2t)-IsoP-induced cytotoxicity. Susceptibility of progenitor OLs was independent of TxA(2) receptor (TP) expression, which was far less in progenitor than in mature OLs. However, TxA(2) synthase was detected in precursor but not in mature OLs, and TxA(2) mimetic U46619 induced hydroperoxides generation and progenitor OL death. The glutathione synthesis enhancer N-acetylcysteine prevented 15-E(2t)-IsoP-induced progenitor cell death. Depletion of glutathione in mature OLs with buthionine sulfoximine rendered them susceptible to cytotoxicity of 15-E(2t)-IsoP. These novel data implicate 15-E(2t)-IsoP as a product of oxidative stress that may contribute in the genesis of PVL.
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http://dx.doi.org/10.1016/j.freeradbiomed.2004.05.007DOI Listing
August 2004

Inhibition of p38 mitogen-activated protein kinase interferes with cell shape changes and gene expression associated with Schwann cell myelination.

Exp Neurol 2003 Sep;183(1):34-46

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.

In the present study we demonstrate that p38, a member of the mitogen-activated protein kinase (MAPK) family, is essential for ascorbate- and laminin-induced myelination in Schwann cell-dorsal root ganglion neuron cocultures. The inhibitory effect of the specific p38 blockers, PD 169316 and SB 203580, on ascorbate-induced myelination was exerted during the early stages (1-2 days) of ascorbate treatment. Inhibition of p38 was further shown to prevent the alignment of Schwann cells along axons in laminin-treated cocultures. The addition of laminin to Schwann cell-dorsal root ganglion neuron cocultures stimulated phosphorylation of p38, thereby demonstrating a link between laminin-induced myelination and p38 activation. Similarly, the small heat shock protein, Hsp27, which is phosphorylated by MAPKAPK2, a downstream substrate of p38, was phosphorylated in response to the addition of laminin to the cocultures. The p38 inhibitors did not affect the proliferation or survival of Schwann cells in the cocultures as assessed by BrdU incorporation and total cell counts. However, p38 inhibition interfered with an early stage in myelination, thereby preventing ascorbate-induced increases in the levels of mRNAs encoding MBP, MAG, and P(0) and reducing laminin deposition. These results indicate that activation of p38 by a signaling pathway(s) involving laminin and appropriate integrin receptor(s) is required for the alignment of Schwann cells with axons that precedes myelination.
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http://dx.doi.org/10.1016/s0014-4886(03)00101-8DOI Listing
September 2003

Catecholamine-induced oligodendrocyte cell death in culture is developmentally regulated and involves free radical generation and differential activation of caspase-3.

Glia 2002 Dec;40(3):283-99

Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.

Oligodendrocyte cultures were used to study the toxic effects of catecholamines. Our results showed that catecholamine-induced toxicity was dependent on the dose of dopamine or norepinephrine used and on the developmental stage of the cultures, with oligodendrocyte progenitors being more vulnerable. A role for oxidative stress and apoptosis on the mechanism of action of catecholamines on oligodendrocyte cell death was next assessed. Catecholamines caused a reduction in intracellular glutathione levels, an accumulation in reactive oxygen species and in heme oxygenase-1, the 32 kDa stress-induced protein. All these changes were prevented by N-acetyl-L-cysteine, a thiocompound with antioxidant activity and a precursor of glutathione, and were more pronounced in progenitors than mature cells, which could contribute to their higher susceptibility. Apoptotic cell death, as assessed by activation of caspase-9 and -3 and cleavage of poly(ADP-ribose) polymerase (a substrate of caspase-3), was only observed in oligodendrocyte progenitors. Pretreatment with zVAD, a general caspase inhibitor, prevented activation of caspase-9 and -3, DNA fragmentation, and decreased progenitors cell death. Furthermore, the expression levels of procaspase-3 and the ratio of the proapoptotic protein bax to antiapoptotic protein bcl-xl were several folds higher in immature than mature oligodendrocytes. Taken together, these results strongly suggest that the catecholamine-induced cytotoxicity in oligodendrocytes is developmentally regulated, mediated by oxidative stress, and have characteristics of apoptosis in progenitor cells.
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http://dx.doi.org/10.1002/glia.10123DOI Listing
December 2002
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