Astroglial Hemichannels and Pannexons: The Hidden Link between Maternal Inflammation and Neurological Disorders.

Maternal inflammation during pregnancy causes later-in-life alterations of the offspring's brain structure and function. These abnormalities increase the risk of developing several psychiatric and neurological disorders, including schizophrenia, intellectual disability, bipolar disorder, autism spectrum disorder, microcephaly, and cerebral palsy. Here, we discuss how astrocytes might contribute to postnatal brain dysfunction following maternal inflammation, focusing on the signaling mediated by two families of plasma membrane channels: hemi-channels and pannexons. [Ca2+]i imbalance linked to the opening of astrocytic hemichannels and pannexons could disturb essential functions that sustain astrocytic survival and astrocyte-to-neuron support, including energy and redox homeostasis, uptake of K+ and glutamate, and the delivery of neurotrophic factors and energy-rich metabolites. Both phenomena could make neurons more susceptible to the harmful effect of prenatal inflammation and the experience of a second immune challenge during adulthood. On the other hand, maternal inflammation could cause excitotoxicity by producing the release of high amounts of gliotransmitters via astrocytic hemichannels/pannexons, eliciting further neuronal damage. Understanding how hemichannels and pannexons participate in maternal inflammation-induced brain abnormalities could be critical for developing pharmacological therapies against neurological disorders observed in the offspring.

Mucosal Exposure to Cigarette Components Induces Intestinal Inflammation and Alters Antimicrobial Response in Mice.

The main environmental risk factor associated with the development of Crohn's disease (CD) is cigarette smoking. Although the mechanism is still unknown, some studies have shown that cigarette exposure affects the intestinal barrier of the small bowel. Among the factors that may be involved in this process are Paneth cells. These specialized epithelial cells are located into the small intestine, and they are able to secrete antimicrobial peptides, having an essential role in the control of the growth of microorganisms. Alterations in its function are associated with inflammatory processes, such as CD. To study how cigarette components impact ileum homeostasis and Paneth cells integrity, we used intragastric administration of cigarette smoke condensate (CSC) in mice. Our results showed that inflammation was triggered after mucosal exposure of CSC, which induced particular alterations in Paneth cells granules, antimicrobial peptide production, and a reduction of bactericidal capacity. In fact, exposure to CSC generated an imbalance in the fecal bacterial population and increased the susceptibility of mice to develop ileal damage in response to bacterial infection. Moreover, our results obtained in mice unable to produce interleukin 10 (IL-10-/- mice) suggest that CSC treatment can induce a symptomatic enterocolitis with a pathological inflammation in genetically susceptible individuals.

The absence of interleukin 10 affects the morphology, differentiation, granule content and the production of cryptidin-4 in Paneth cells in mice.

Paneth cells (PCs) are specialized epithelial cells of the small bowel that contain multiple secretory granules filled with antimicrobial peptides and trophic factors, which are essential for the control of the microorganisms growth and maintaining intestinal integrity. Alterations in their function are associated with an imbalance of the normal microbiota, gastrointestinal infections and inflammatory processes, such as Crohn's disease (CD). One of the most common murine models for studying CD is IL-10-/- mouse. IL-10-/- mice when housed in conventional conditions and take contact with commensal microorganisms develop an acute enterocolitis mediated by a Th1 immune response. Even though, alterations in PCs function are related to CD, they had not been characterized yet in this mouse model. Here we show that in specific pathogen free conditions IL-10-/- mice have aberrant granules and a large number of immature PCs at the bottom of the crypt in the ileum of IL-10-/- mice before developing intestinal inflammation, along with a reduced expression of Indian Hedgehog. In addition, IL-10-/- Paneth cells presented a reduced expression of cryptidin-4, and a heterogeneous distribution of lysozyme+ granules. The alterations in the maturation of the PCs at the bottom of the crypt were not modified after the colonization by the conventional microbiota. On the other hand, depletion of microbiota altered the phenotype, but did not normalize PCs. Our results suggest that IL-10 could be necessary for the integrity of PCs. Moreover, our results help to explain why IL-10-/- mice develop enterocolitis in response to microorganisms.

Etiologic and clinical characterization of community acquired gastroenteritis in adult patients in a Chilean emergency room by the FilmArray GI panel.

Infectious diarrhea can be caused by a large number of microorganisms including bacteria virus and parasites. The clinical syndromic approach has been traditionally used to guide therapy. The aim of this study was to characterize the etiology of acute diarrhea by the FilmArray GI panel and to correlate it with its clinical presentation in an adult population presenting to the emergency room in a developing country. MATERIAL AND METHODS: Adult patients attending the ER due to acute diarrhea were selected. All patients included had a FilmArray GI panel performed and the clinical characteristics were recorded. RESULTS: One hundred and ninety-nine patients were included. One hundred and eighteen (59.3%) were females. The mean age was 43 years old. Thirty three percent of the patients presented dysentery, 36.7% fever, 54.8% referred nauseas and 35.7% vomiting. Sixty three percent of the patients presented some degree of dehydration. In total, 221 microorganisms were detected of which 71.5% corresponded to bacteria (158/221), 19.9% to virus (44/221) and 8.6% to parasites (19/221). In 133 (67.0%) of 199 patients at least one microorganism was identified. Infections with more than one microorganism were detected in 27.1% of the patients. Polimicrobial infections were associated with a higher frequency of nausea (50.0% vs 32.0%, p 0.046), abdominal pain (87.0% vs 44.0%, p<0.0001) and travel history (20.0% vs 5.0%, p 0.0102). Bacterial infections occurred without a seasonal distribution with the exception of Salmonella sp whereas viral infections predominated during the autumn-winter months. Diarreicogenic E. coli were present in the context of a co-infection in more than 80.0% of the cases. DISCUSSION: The use of multiplex panels has given us invaluable information regarding the epidemiology of acute diarrhea in adult. It highlighted the importance of polimicrobial infections and the frequency of diarreicogenic E. coli infections. Nevertheless, the lack of severity compared to monomicrobial infections and the usual association with other microorganisms in the latter make their clinical importance debatable.

Prenatal nicotine exposure enhances Cx43 and Panx1 unopposed channel activity in brain cells of adult offspring mice fed a high-fat/cholesterol diet.

Nicotine, the most important neuroteratogen of tobacco smoke, can reproduce brain and cognitive disturbances per se when administered prenatally. However, it is still unknown if paracrine signaling among brain cells participates in prenatal nicotine-induced brain impairment of adult offspring. Paracrine signaling is partly mediated by unopposed channels formed by connexins hemichannels (HCs) and pannexins serving as aqueous pores permeable to ions and small signaling molecules, allowing exchange between the intra- and extracellular milieus. Our aim was to address whether prenatal nicotine exposure changes the activity of those channels in adult mice offspring under control conditions or subjected to a second challenge during young ages: high-fat/cholesterol (HFC) diet. To induce prenatal exposure to nicotine, osmotic minipumps were implanted in CF1 pregnant mice at gestational day 5 to deliver nicotine bitartrate or saline (control) solutions. After weaning, offspring of nicotine-treated or untreated pregnant mice were fed ad libitum with chow or HFC diets for 8 weeks. The functional state of connexin 43 (Cx43) and pannexin 1 (Panx1) unopposed channels was evaluated by dye uptake experiments in hippocampal slices from 11-week-old mice. We found that prenatal nicotine increased the opening of Cx43 HCs in astrocytes, and Panx1 channels in microglia and neurons only if offspring mice were fed with HFC diet. Blockade of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2) and prostaglandin E receptor 1 (EP1), ionotropic ATP receptor type 7 (P2X7) and NMDA receptors, showed differential inhibition of prenatal nicotine-induced channel opening in glial cells and neurons. Importantly, inhibition of the above mentioned enzymes and receptors, or blockade of Cx43 and Panx1 unopposed channels greatly reduced adenosine triphosphate (ATP) and glutamate release from hippocampal slices of prenatally nicotine-exposed offspring. We propose that unregulated gliotransmitter release through Cx43 and Panx1 unopposed channels may participate in brain alterations observed in offspring of mothers exposed to tobacco smoke during pregnancy.

Age-dependent changes on TGFß1 Smad3 pathway modify the pattern of microglial cell activation.

Aging is the main risk factor for Alzheimer's disease. Among other characteristics, it shows changes in inflammatory signaling that could affect the regulation of glial cell activation. We have shown that astrocytes prevent microglial cell cytotoxicity by mechanisms mediated by TGFß1. However, whereas TGFß1 is increased, glial cell activation persists in aging. To understand this apparent contradiction, we studied TGFß1-Smad3 signaling during aging and their effect on microglial cell function. TGFß1 induction and activation of Smad3 signaling in the hippocampus by inflammatory stimulation was greatly reduced in adult mice. We evaluated the effect of TGFß1-Smad3 pathway on the regulation of nitric oxide (NO) and reactive oxygen species (ROS) secretion, and phagocytosis of microglia from mice at different ages with and without in vivo treatment with lipopolysaccharide (LPS) to induce an inflammatory status. NO secretion was only induced on microglia from young mice exposed to LPS, and was potentiated by inflammatory preconditioning, whereas in adult mice the induction of ROS was predominant. TGFß1 modulated induction of NO and ROS production in young and adult microglia, respectively. Modulation was partially dependent on Smad3 pathway and was impaired by inflammatory preconditioning. Phagocytosis was induced by inflammation and TGFß1 only in microglia cultures from young mice. Induction by TGFß1 was also prevented by Smad3 inhibition. Our findings suggest that activation of the TGFß1-Smad3 pathway is impaired in aging. Age-related impairment of TGFß1-Smad3 can reduce protective activation while facilitating cytotoxic activation of microglia, potentiating microglia-mediated neurodegeneration.

ATP is required and advances cytokine-induced gap junction formation in microglia in vitro.

Microglia are the immune cells in the central nervous system. After injury microglia release bioactive molecules, including cytokines and ATP, which modify the functional state of hemichannels (HCs) and gap junction channels (GJCs), affecting the intercellular communication via extracellular and intracellular compartments, respectively. Here, we studied the role of extracellular ATP and several cytokines as modulators of the functional state of microglial HCs and GJCs using dye uptake and dye coupling techniques, respectively. In microglia and the microglia cell line EOC20, ATP advanced the TNF-α/IFN-γ-induced dye coupling, probably through the induction of IL-1ß release. Moreover, TNF-α/IFN-γ, but not TNF-α plus ATP, increased dye uptake in EOC20 cells. Blockade of Cx43 and Panx1 HCs prevented dye coupling induced by TNF-α/IFN-γ, but not TNF-α plus ATP. In addition, IL-6 prevented the induction of dye coupling and HC activity induced by TNF-α/IFN-γ in EOC20 cells. Our data support the notion that extracellular ATP affects the cellular communication between microglia through autocrine and paracrine mechanisms, which might affect the timing of immune response under neuroinflammatory conditions.

Proinflammatory stimuli are needed for induction of microglial cell-mediated AbetaPP_{244-C} and Abeta-neurotoxicity in hippocampal cultures.

Amyloid-beta plaques and neurodegeneration are hallmarks of Alzheimer's disease, where glial cells are responsible for sustained neuroinflammation. Here we show that hippocampal-microglia co-cultures exposed to proinflammatory mediators, amyloid-beta- and amyloid-beta protein precursor construct-conjugated beads increased their production of nitrites. In contrast, inflammation was unable to significantly induce cell death by itself, whereas inflammation plus amyloid-beta or amyloid-beta protein precursor induced a significant increment of cell death and a 6-fold increase of production of Interleukin 1beta. Those effects were not observed in the absence of microglia or when hippocampal cells were co-cultured with microglia for one day. In contrast, a 2-fold increase of transforming growth factor beta1 was observed in hippocampal cultures exposed to inflammatory stimuli for 4 days, whereas induction of transforming growth factor beta1 by inflammation plus amyloid-beta and amyloid-beta protein precursor was nearly abolished by microglia. Our results indicate that neurotoxicity induced by amyloid-beta or amyloid-beta protein precursor was a slow process depending on activated microglia and additional stimuli. The observed cytotoxicity could be consequence of a vicious cycle in which elevated concentrations of Interleukin 1beta and radical species along with decreased secretion of neuroprotective cytokines such as transforming growth factor beta1 support persistent activation of glial cells and cell damage.

Pro-inflammatory conditions promote neuronal damage mediated by Amyloid Precursor Protein and decrease its phagocytosis and degradation by microglial cells in culture.

Aberrant handling of Amyloid Precursor Protein (APP) and beta-amyloid (Abeta), glial activation and inflammation are key events in Alzheimer's disease. We set out to determine the role of inflammation on microglial reactivity against APP. We studied microglia-mediated neurotoxicity, uptake and degradation of a biotinylated APP construct (biotin-APP-C-244). APP, in contrast to Abeta, only induced mild activation of glial cells. However, under pro-inflammatory conditions, APP induced microglial-mediated cytotoxicity. Biotin-APP-C-244 or lipopolysaccharide and interferon-gamma (LPS+IFNgamma), administered separately, did not change reduction metabolism of microglia. However, biotin-APP-C-244+(LPS+IFNgamma) increased microglial reactivity and decreased reduction metabolism by 75% (P<0.001). Biotin-APP-C-244 was readily taken up by microglial cells; 80% was phagocytosed at 2 h. In the presence of LPS+IFNgamma, phagocytosis of biotin-APP-C-244 was reduced at 2 h; and cell damage was evident after 4 h. Our results support our hypothesis that, in neuroinflammation, microglial scavenger function is impaired and reactivity against APP enhanced as an initial step for neurodegeneration.

Protection of rat primary hippocampal cultures from A beta cytotoxicity by pro-inflammatory molecules is mediated by astrocytes.

The brain of Alzheimer's disease patients shows abundant dystrophic neurites in close proximity to fibrillar beta-amyloid (A beta) plaques, and activated glial cells. We evaluated the influence of pro-inflammatory molecules (LPS + IFN-gamma) on A beta(1-42) neurotoxicity. 2 microM A beta(1-42) induced apoptosis of hippocampal cells and LPS + IFN-gamma reduced the apoptosis induced by A beta. However, LPS + IFN-gamma prevented apoptosis only in hippocampal cultures containing astrocytes. Also, LPS + IFN-gamma induced the secretion of TGF beta, a cytokine having neuroprotective effects, only in hippocampal cultures that contained astrocytes. Astrocytes had a regulatory effect over microglial and neuronal responses to A beta. The results suggest that LPS + IFN-gamma, traditionally considered as pro-apoptotic, reduced apoptosis induced by A beta through the activation of neuroprotective mechanisms mediated by astrocytes. We propose that astrocytes are pivotal in the modulation of inflammation of the CNS. The impairment of the regulatory functions performed by activated astrocytes could represent an important pathogenic mechanism for neurodegenerative diseases.

Oligodendrocytes damage in Alzheimer's disease: beta amyloid toxicity and inflammation.

Research on Alzheimer's disease (AD) focuses mainly on neuronal death and synaptic impairment induced by beta-Amyloid peptide (Abeta), events at least partially mediated by astrocyte and microglia activation. However, substantial white matter damage and its consequences on brain function warrant the study of oligodendrocytes participation in the pathogenesis and progression of AD. Here, we analyze reports on oligodendrocytes' compromise in AD and discuss some experimental data indicative of Abeta toxicity in culture. We observed that 1 microM of fibrilogenic Abeta peptide damages oligodendrocytes in vitro: while pro-inflammatory molecules (1 microg/ ml LPS + 1 ng/ml IFNgamma) or the presence of astrocytes reduced the Abeta-induced damage. This agrees with our previous results showing an astrocyte-mediated protective effect over Abeta-induced damage on hippocampal cells and modulation of the activation of microglial cells in culture. Oligodendrocytes protection by astrocytes could be, either by reduction of Abeta fibrilogenesis/deposition or prevention of oxidative damage. Likewise, the decrease of Abeta-induced damage by proinflammatory molecules could reflect the production of trophic factors by activated oligodendrocytes and/or a metabolic activation as observed during myelination. Considering the association of inflammation with neurodegenerative diseases. oligodendrocytes impairment in AD patients could potentiate cell damage under pathological conditions.

Oligodendrocytes damage in Alzheimer's disease: Beta amyloid toxicity and inflammation

Research on Alzheimer's disease (AD) focuses mainly on neuronal death and synaptic impairment induced by â-Amyloid peptide (Aâ), events at least partially mediated by astrocyte and microglia activation. However, substantial white matter damage and its consequences on brain function warrant the study of oligodendrocytes participation in the pathogenesis and progression of AD. Here, we analyze reports on oligodendrocytes' compromise in AD and discuss some experimental data indicative of Aâ toxicity in culture. We observed that 1 ìM of fibrilogenic Aâ peptide damages oligodendrocytes in vitro; while pro-inflammatory molecules (1 ìg/ml LPS + 1 ng/ml IFNã) or the presence of astrocytes reduced the Ab-induced damage. This agrees with our previous results showing an astrocyte-mediated protective effect over Aâ-induced damage on hippocampal cells and modulation of the activation of microglial cells in culture. Oligodendrocytes protection by astrocytes could be, either by reduction of Aâ fibrilogenesis/deposition or prevention of oxidative damage. Likewise, the decrease of Aâ-induced damage by proinflammatory molecules could reflect the production of trophic factors by activated oligodendrocytes and/or a metabolic activation as observed during myelination. Considering the association of inflammation with neurodegenerative diseases, oligodendrocytes impairment in AD patients could potentiate cell damage under pathological conditions.

Supplementation of female rats with alpha-linolenic acid or docosahexaenoic acid leads to the same omega-6/omega-3 LC-PUFA accretion in mother tissues and in fetal and newborn brains.

BACKGROUND: Maternal omega-3 fatty acid supplementation has been suggested to provide docosahexaenoic acid (DHA) for the normal brain development during gestation. DHA can be given as such (preformed) or through the omega-3 precursor alpha-linolenic acid (LNA) which is transformed into DHA by elongation and desaturation reactions. Western diet provides low amounts of LNA and DHA; therefore, supplementation with these omega-3 fatty acids has been suggested for pregnant women. However, the bioequivalence of LNA ingestion to DHA supplementation has not been established. METHODS: Recently weaning female Wistar rats were fed a diet containing a small amount of LNA and no DHA. The animals were daily supplemented 40 days before mating, during pregnancy, and until delivery with 60 mg/kg of LNA or 6 mg/kg of DHA dissolved in coconut oil. Fatty acids were given as ethyl ester derivatives. Controls received coconut oil. The fatty acid composition of blood plasma, erythrocytes, liver, visceral adipose tissue, and brain segments (frontal cortex, hippocampus, and cerebellum) was analyzed. Brain segments obtained from 16- and 19-day-old fetuses and from 2- and 21-day-old rats were also analyzed for fatty acid composition. RESULTS: Supplementation with LNA and DHA induced a similar accretion of DHA in plasma, erythrocytes, liver, and brain segments of the mothers. The adipose tissue showed a higher DHA accretion after DHA-supplementation. The DHA accretion in frontal cortex, hippocampus, and cerebellum obtained from the fetuses and the newborn rats was similar when the mothers were supplemented with LNA and DHA. Our results show that under our experimental conditions a similar accretion of DHA in the different tissues of the mothers and in the brain segments of fetuses and newborn rats is obtained after LNA and DHA supplementation. CONCLUSION: LNA and DHA, at the amounts given in this study, show a similar bioequivalence for DHA accretion in different tissues of the mother and in brain segments of fetuses and newborn rats.

Acetylcholinesterase induces the expression of the beta-amyloid precursor protein in glia and activates glial cells in culture.

Acetylcholinesterase (AChE) activities in CNS physiopathology are increasingly diverse and range from neuritogenesis, through synaptogenesis, to enhancement of amyloid fiber assembly. In Alzheimer's disease, senile plaques and neurodegeneration specially affect regions enriched for cholinergic synapses. In this study we show an effect of AChE that could contribute to the increased deposition of Abeta in certain regions. Affinity-purified AChE induced the expression of amyloid-beta-precursor protein (beta-APP) in glial cells in a concentration-dependent manner up to 5 nM. In glia, AChE also increased inducible nitric oxide synthase (iNOS) assessed by immunocytochemistry and decreased reductive metabolism as evidence of cell activation. AChE could increase the expression of beta-APP in astrocytes and microglia as result of the activation of glial cells. As a whole, we found that AChE has additional effects that could result in an increased synthesis of Abeta, both by increasing beta-APP expression of astrocytes and by further activating glial cells.

Microglia - astrocyte interaction in Alzheimer's disease: friends or foes for the nervous system?

Brain glial cells secrete several molecules that can modulate the survival of neurons after various types of damage to the CNS. Activated microglia and astrocytes closely associate to amyloid plaques in Alzheimer Disease (AD). They could have a role in the neurotoxicity observed in AD because of the inflammatory reaction they generate. There is controversy regarding the individual part played by the different glial cells, and the interrelationships between them. Both astrocytes and microglia produce several cytokines involved in the inflammatory reaction. Moreover, the same cytokines may have different effects, depending on their concentration and the type of cells in the vicinity. In turn, the events occurring in response to injury may lead to changes in the nature and relative concentration of the various factors involved. To learn about these putative glial interrelationships, we examined some effects of astrocytes on microglial activation.