RESUMEN
Chronic consumption of ß-sitosterol-ß-D-glucoside (BSSG), a neurotoxin contained in cycad seeds, leads to Parkinson's disease in humans and rodents. Here, we explored whether a single intranigral administration of BSSG triggers neuroinflammation and neurotoxic A1 reactive astrocytes besides dopaminergic neurodegeneration. We injected 6 µg BSSG/1 µL DMSO or vehicle into the left substantia nigra and immunostained with antibodies against tyrosine hydroxylase (TH) together with markers of microglia (OX42), astrocytes (GFAP, S100ß, C3), and leukocytes (CD45). We also measured nitric oxide (NO), lipid peroxidation (LPX), and proinflammatory cytokines (TNF-α, IL-1ß, IL-6). The Evans blue assay was used to explore the blood-brain barrier (BBB) permeability. We found that BSSG activates NO production on days 15 and 30 and LPX on day 120. Throughout the study, high levels of TNF-α were present in BSSG-treated animals, whereas IL-1ß was induced until day 60 and IL-6 until day 30. Immunoreactivity of activated microglia (899.0 ± 80.20%) and reactive astrocytes (651.50 ± 11.28%) progressively increased until day 30 and then decreased to remain 251.2 ± 48.8% (microglia) and 91.02 ± 39.8 (astrocytes) higher over controls on day 120. C3(+) cells were also GFAP and S100ß immunoreactive, showing they were neurotoxic A1 reactive astrocytes. BBB remained permeable until day 15 when immune cell infiltration was maximum. TH immunoreactivity progressively declined, reaching 83.6 ± 1.8% reduction on day 120. Our data show that BSSG acute administration causes chronic neuroinflammation mediated by activated microglia, neurotoxic A1 reactive astrocytes, and infiltrated immune cells. The severe neuroinflammation might trigger Parkinson's disease in BSSG intoxication.
Asunto(s)
Astrocitos/efectos de los fármacos , Astrocitos/inmunología , Inflamación/etiología , Neurotoxinas/inmunología , Sitoesteroles/administración & dosificación , Sustancia Negra/efectos de los fármacos , Sustancia Negra/metabolismo , Animales , Astrocitos/metabolismo , Biomarcadores , Enfermedad Crónica , Citocinas/metabolismo , Modelos Animales de Enfermedad , Humanos , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Inflamación/patología , Mediadores de Inflamación/metabolismo , Leucocitos/inmunología , Leucocitos/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Masculino , Microglía/inmunología , Microglía/metabolismo , Neurotoxinas/efectos adversos , Estrés Oxidativo/efectos de los fármacos , Enfermedad de Parkinson/etiología , Enfermedad de Parkinson/metabolismo , Enfermedad de Parkinson/patología , Ratas , Sustancia Negra/patologíaRESUMEN
Beta-lactam resistant bacteria, which are commonly resident in tertiary hospitals, have emerged as a worldwide health problem because of ready-to-eat vegetable intake. We aimed to characterize the genes that provide resistance to beta-lactam antibiotics in Enterobacteriaceae, isolated from five commercial salad brands for human consumption in Mexico City. In total, twenty-five samples were collected, grown in blood agar plates, and the bacteria were biochemistry identified and antimicrobial susceptibility testing was done. The carried family genes were identified by endpoint PCR and the specific genes were confirmed with whole genome sequencing (WGS) by Next Generation Sequencing (NGS). Twelve positive cultures were identified and their microbiological distribution was as follows: 8.3% for Enterobacter aerogene (n = 1), 8.3% for Serratia fonticola (n = 1), 16.7% for Serratia marcesens (n = 2), 16.7% for Klebsiella pneumoniae (n = 2), and 50% (n = 6) for Enterobacter cloacae. The endpoint PCR results showed 11 colonies positive for blaBIL (91.7%), 11 for blaSHV (91.7%), 11 for blaCTX (97.7%), 12 for blaDHA (100%), four for blaVIM (33.3%), two for blaOXA (16.7%), two for blaIMP (16.7%), one for blaKPC (8.3%), and one for blaTEM (8.3%) gen; all samples were negative for blaROB, blaCMY, blaP, blaCFX and blaLAP gene. The sequencing analysis revealed a specific genotype for Enterobacter cloacae (blaSHV-12, blaCTX-M-15, blaDHA-1, blaKPC-2); Serratia marcescens (blaSHV-1, blaCTX-M-3, blaDHA-1, blaVIM-2); Klebsiella pneumoniae (blaSHV-12, blaCTX-M-15, blaDHA-1); Serratia fonticola (blaSHV-12, blaVIM-1, blaDHA-1); and, Enterobacter aerogene (blaSHV-1, blaCTX-M-1, blaDHA-1, blaVIM-2, blaOXA-9). Our results indicate that beta-lactam-resistant bacteria have acquired integrons with a different number of genes that provide pan-resistance to beta-lactam antibiotics, including penicillins, oxacillins, cefalosporins, monobactams, carbapenems, and imipenems.
RESUMEN
Models of Parkinson's disease with neurotoxins have shown that microglial activation does not evoke a typical inflammatory response in the substantia nigra, questioning whether neuroinflammation leads to neurodegeneration. To address this issue, the archetypal inflammatory stimulus, lipopolysaccharide (LPS), was injected into the rat substantia nigra. LPS induced fever, sickness behavior, and microglial activation (OX42 immunoreactivity), followed by astrocyte activation and leukocyte infiltration (GFAP and CD45 immunoreactivities). During the acute phase of neuroinflammation, pro- and anti-inflammatory cytokines (TNF-α, IL-1ß, IL-6, IL-4, and IL-10) responded differentially at mRNA and protein level. Increased NO production and lipid peroxidation occurred at 168 h after LPS injection. At this time, evidence of neurodegeneration could be seen, entailing decreased tyrosine hydroxylase (TH) immunoreactivity, irregular body contour, and prolongation discontinuity of TH+ cells, as well as apparent phagocytosis of TH+ cells by OX42+ cells. Altogether, these results show that LPS evokes a typical inflammatory response in the substantia nigra that is followed by dopaminergic neurodegeneration.
Asunto(s)
Astrocitos/fisiología , Neuronas Dopaminérgicas/fisiología , Leucocitos Mononucleares/fisiología , Lipopolisacáridos/inmunología , Microglía/fisiología , Enfermedades Neurodegenerativas/inmunología , Inflamación Neurogénica/inmunología , Enfermedad de Parkinson/inmunología , Porción Compacta de la Sustancia Negra/inmunología , Tirosina 3-Monooxigenasa/inmunología , Enfermedad Aguda , Animales , Diferenciación Celular , Movimiento Celular , Células Cultivadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Humanos , Peroxidación de Lípido , Masculino , Ratas , Ratas WistarRESUMEN
BACKGROUND: The anti-inflammatory effect of the cerebral dopamine neurotrophic factor (CDNF) was shown recently in primary glial cell cultures, yet such effect remains unknown both in vivo and in 6-hydroxydopamine (6-OHDA) models of Parkinson's disease (PD). We addressed this issue by performing an intranigral transfection of the human CDNF (hCDNF) gene in the critical period of inflammation after a single intrastriatal 6-OHDA injection in the rat. METHODS: At day 15 after lesion, the plasmids p3xNBRE-hCDNF or p3xNBRE-EGFP, coding for enhanced green florescent protein (EGFP), were transfected into the rat substantia nigra (SN) using neurotensin (NTS)-polyplex. At day 15 post-transfection, we measured nitrite and lipoperoxide levels in the SN. We used ELISA to quantify the levels of TNF-α, IL-1ß, IL-6, endogenous rat CDNF (rCDNF) and hCDNF. We also used qRT-PCR to measure rCDNF and hCDNF transcripts, and immunofluorescence assays to evaluate iNOS, CDNF and glial cells (microglia, astrocytes and Neuron/Glial type 2 (NG2) cells). Intact SNs were additional controls. RESULTS: In the SN, 6-OHDA triggered nitrosative stress, increased inflammatory cytokines levels, and activated the multipotent progenitor NG2 cells, which convert into astrocytes to produce rCDNF. In comparison with the hemiparkinsonian rats that were transfected with the EGFP gene or without transfection, 6-OHDA treatment and p3xNBRE-hCDNF transfection increased the conversion of NG2 cells into astrocytes resulting in 4-fold increase in the rCDNF protein levels. The overexpressed CDNF reduced nitrosative stress, glial markers and IL-6 levels in the SN, but not TNF-α and IL-1ß levels. CONCLUSION: Our results show the anti-inflammatory effect of CDNF in a 6-OHDA rat of Parkinson's disease. Our results also suggest the possible participation of TNF-α, IL-1ß and IL-6 in rCDNF production by astrocytes, supporting their anti-inflammatory role.
Asunto(s)
Factores de Crecimiento Nervioso/biosíntesis , Factores de Crecimiento Nervioso/genética , Oxidopamina/toxicidad , Sustancia Negra/efectos de los fármacos , Sustancia Negra/metabolismo , Transfección , Animales , Humanos , Inflamación/inducido químicamente , Inflamación/genética , Inflamación/metabolismo , Inyecciones Intraventriculares , Masculino , Oxidopamina/administración & dosificación , Ratas , Ratas WistarRESUMEN
BACKGROUND AND AIMS: Glutathione peroxidase 3 (GPx3) plays a main role in removing hydro- and lipoperoxides from the body. Changes in concentration and several single-nucleotide polymorphisms (SNP) at the GPX3 gene have been associated with vascular diseases, but the relationship of GPx3 with metabolic syndrome (MetS) remains unexplored. We undertook this study to determine the association of GPx3 serum levels and several GPX3 SNPs with the presence of MetS in Mexican subjects. METHODS: Clinical, biochemical, and anthropometric evaluation were conducted in 426 subjects assigned to three groups: control (n = 42); risk group (RG, n = 200), and MetS group (n = 184). Insulin sensitivity (IS) and cardiovascular risk were determined by the QUICKI and TG/HDL-C index, respectively. Serum GPx3 was determined by enzyme immunoassay and polymorphisms within GPX3 gene were identified by nucleotide sequencing. RESULTS: MetS group showed low IS and increased cardiovascular risk with respect to controls as well as higher GPx3 serum levels (172.9 ± 32.2 vs. 145.6 ± 24.8 ng/dL; p <0.05). Only three of the ten GPX3 SNPs screened were polymorphic with two haplotypes observed (CCT and TTA-rs8177404, rs8177406, and rs8177409), indicating tight linkage disequilibrium in this genetic region. No differences for either genotype or allele frequencies among groups were observed, but rs8177409 (allele T) was associated with cardiovascular risk (odds ratio [OR], 4.5; p = 0.0125). CONCLUSION: This study shows that serum levels of GPx3 are increased in subjects with MetS and that rs8177409 SNP was associated with cardiovascular risk in a Mexican population.
Asunto(s)
Glutatión Peroxidasa/sangre , Glutatión Peroxidasa/genética , Síndrome Metabólico/sangre , Síndrome Metabólico/genética , Polimorfismo de Nucleótido Simple , Adulto , Biomarcadores/sangre , Enfermedades Cardiovasculares/etiología , Estudios de Casos y Controles , Femenino , Frecuencia de los Genes , Marcadores Genéticos , Haplotipos , Humanos , Resistencia a la Insulina/genética , Desequilibrio de Ligamiento , Modelos Logísticos , Masculino , Síndrome Metabólico/complicaciones , Síndrome Metabólico/diagnóstico , México , Persona de Mediana Edad , Oportunidad Relativa , Reacción en Cadena de la Polimerasa , Medición de Riesgo , Factores de Riesgo , Análisis de Secuencia de ADNRESUMEN
High- and low-yawning rats (HY and LY) were selectively bred as a function of their spontaneous yawning frequency with the LY subline about 2 yawns/hr and the HY 20 yawns/hr. The HY rats have more grooming bouts and travel longer distances in an open field. HY dams spent less time in the nest, retrieved their pups faster, and show a longer latency to licking and mouthing the pups than the LY or outbred Sprague-Dawley (SD) animals. The percentage of HY dams that had atypical retrieving was higher, with a lower nest quality, and produced offspring whose weights were lower than those from the LY subline. We also showed that the pregnant HY dams have fewer pups and the percentage that had lost at least three pups during lactation was higher than the SD and LY dams. In conclusion, HY dams are motivated to take care of their pups, but the "fine tuning" of maternal care is disturbed.
Asunto(s)
Conducta Materna/fisiología , Bostezo/fisiología , Análisis de Varianza , Animales , Conducta Animal/fisiología , Femenino , Tamaño de la Camada/fisiología , Embarazo , Ratas , Ratas Sprague-Dawley , Tiempo de Reacción/fisiología , Especificidad de la Especie , Estadísticas no ParamétricasRESUMEN
The transient occlusion of cerebral arteries causes an increase in zinc levels in the brain, which is associated with a production of nitric oxide (NO). The types of zinc transporters (ZnT) involved in zinc homeostasis in the cerebral cortex after hypoxia-ischemia are not completely known. We studied the effect of the transient occlusion (10 min) of the common carotid artery (CCA) on NO-induced zinc levels, ZnT mRNA expression, and cell-death markers in the cerebral cortex-hippocampus of the rat. Nitrites, zinc, and lipoperoxidation were quantified by colorimetric methods, ZnT expression was determined by RT-PCR, caspase-3 by ELISA and immunohistochemistry, and histopathological alterations by H&E staining. After restoration of the blood flow, the basal levels of NO and zinc increased in a biphasic manner over time, but the peaks of NO levels appeared earlier (2 h and 24 h) than those of zinc (6 h and 36 h). Upregulation of ZnT1, ZnT2, and ZnT4 mRNAs was determined after 8-h postreperfusion, but ZnT3 RNA levels were unaffected. Lipoperoxidation and caspase-3 levels were also increased, and necrosis and apoptosis were present at 24 h postreperfusion. All the effects determined were prevented by l-nitro-arginine methyl ester injected 1 h before the occlusion of the CCA. Our results suggest that the upregulation of ZnT1, ZnT2, and ZnT4 was to decrease the cytosolic zinc levels caused by NO after transient occlusion of the CCA, although this was unable to lead to physiological levels of zinc and to prevent cell damage in the cerebral cortex-hippocampus of the rat.
Asunto(s)
Isquemia Encefálica/metabolismo , Corteza Cerebral/metabolismo , Proteínas de la Membrana/genética , Óxido Nítrico/metabolismo , Daño por Reperfusión/metabolismo , Zinc/metabolismo , Animales , Isquemia Encefálica/fisiopatología , Proteínas Portadoras/genética , Caspasa 3/metabolismo , Proteínas de Transporte de Catión/genética , Muerte Celular/efectos de los fármacos , Muerte Celular/fisiología , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/fisiopatología , Citosol/efectos de los fármacos , Citosol/metabolismo , Inhibidores Enzimáticos/farmacología , Peroxidación de Lípido/efectos de los fármacos , Peroxidación de Lípido/fisiología , Masculino , Proteínas de Transporte de Membrana , Óxido Nítrico/farmacología , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Isoformas de Proteínas/genética , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Daño por Reperfusión/fisiopatología , Regulación hacia Arriba/fisiologíaRESUMEN
Cultured glial cells from the cerebellum of 15-day-old taiep rats produced NO, increased iNOS levels, up-regulated iNOS expression and promoted TNF release when stimulated with LPS and IFNgamma. These responses were much greater than in control cells. In taiep glial cells, NO production and iNOS levels and expression induced by the co-stimulatory signal were resistant to the inhibitory effect of TGFbeta1. The glial cell priming might have been generated by oligodendrocyte alteration in taiep rats.
Asunto(s)
Enfermedades Desmielinizantes/genética , Enfermedades Desmielinizantes/metabolismo , Neuroglía/metabolismo , Óxido Nítrico/metabolismo , Animales , Células Cultivadas , Enfermedades Desmielinizantes/inmunología , Neuroglía/efectos de los fármacos , Neuroglía/inmunología , Óxido Nítrico/inmunología , Ratas , Ratas Mutantes , Ratas Sprague-DawleyRESUMEN
The effect of transient uteroplacental ischemia on nitric oxide (NO) levels, enzymatic activity, and expression of NO synthase (NOS) isoforms was studied in fetal rat brains. Fetuses were subjected to ischemia by clamping the uterine arteries for 5 min on gestational day 17 (GD17). At different times after ischemia, fetuses were delivered by Cesarean section under anesthesia to obtain the brains. Transient uteroplacental ischemia produced a time dependent increase in nitrite levels in the brain, reaching a maximum value (300 +/- 25% of baseline) 24 h after uterine artery occlusion and remaining elevated as long as 48 h. Significantly increased nitrite levels were found in the cerebral cortex but not in the mesencephalon and cerebellum. The ischemia-induced increment in nitrite levels was totally blocked by either L-NAME (10 mg/kg) or AMT (0.65 mg/kg) administered i.p. 1 h before uterine artery occlusion. Both Ca(2+)-dependent and Ca(2+)-independent NOS activities in the cerebral cortex remained significantly increased with respect to controls after 24 h following the ischemia. Reverse transcriptase-polymerase chain reaction showed augmented levels of mRNAs for both nNOS and iNOS when compared with controls at 8 h after ischemia. At 36 h, nNOS mRNA returned to basal levels whereas eNOS mRNA levels increased and iNOS mRNA remained elevated. Our results show that the three NOS isoforms participate in increasing NO levels after transient ischemia and suggest a biphasic and differential regulation of the expression of constitutive NOS isoforms in the rat cerebral cortex.