The Neurotoxic Effect of Ochratoxin-A on the Hippocampal Neurogenic Niche of Adult Mouse Brain.

Ochratoxin A (OTA) is a common secondary metabolite of Aspergillus ochraceus, A. carbonarius, and Penicillium verrucosum. This mycotoxin is largely present as a contaminant in several cereal crops and human foodstuffs, including grapes, corn, nuts, and figs, among others. Preclinical studies have reported the involvement of OTA in metabolic, physiologic, and immunologic disturbances as well as in carcinogenesis. More recently, it has also been suggested that OTA may impair hippocampal neurogenesis in vivo and that this might be associated with learning and memory deficits. Furthermore, aside from its widely proven toxicity in tissues other than the brain, there is reason to believe that OTA contributes to neurodegenerative disorders. Thus, in this present in vivo study, we investigated this possibility by intraperitoneally (i.p.) administering 3.5 mg OTA/kg body weight to adult male mice to assess whether chronic exposure to this mycotoxin negatively affects cell viability in the dentate gyrus of the hippocampus. Immunohistochemistry assays showed that doses of 3.5 mg/kg caused a significant and dose-dependent reduction in repetitive cell division and branching (from 12% to 62%). Moreover, the number of countable astrocytes (p < 0.001), young neurons (p < 0.001), and mature neurons (p < 0.001) negatively correlated with the number of i.p. OTA injections administered (one, two, three, or six repeated doses). Our results show that OTA induced adverse effects in the hippocampus cells of adult mice brain tissue when administered in cumulative doses.

The effect of a mindfulness-based therapy on different biomarkers among patients with inflammatory bowel disease: a randomised controlled trial.

Mindfulness-based interventions have shown some efficacy in decreasing stress levels and improving quality of life. However, so far, only a few studies have studied this type of intervention among patients with inflammatory bowel disease and none of them have studied their effects on inflammatory biomarkers. This current study was a two-armed, single-centre, randomised (2:1 ratio) controlled trial used to evaluate the effects of a mindfulness-based intervention (n = 37) compared to standard medical therapy (n = 20) in patients with Crohn's disease or ulcerative colitis. The mindfulness intervention blended four internet-based therapy modules with four face-to-face support sessions. The outcomes we assessed were faecal calprotectin (primary outcome), C-reactive protein, and cortisol levels measured in hair samples at several timepoints. The between-group analysis highlighted significant decreases in faecal calprotectin and in C-reactive protein levels in the mindfulness-based intervention group compared to the standard medical therapy group at the six-month follow-up (faecal calprotectin: -367, [95% CI: -705, -29], P = 0.03; C-reactive protein: -2.82, [95% CI: -5.70, 0.08], P = 0.05), with moderate to large effect sizes (faecal calprotectin: ηp2 = 0.085; C-reactive protein: ηp2 = 0.066). We concluded that mindfulness-based therapy administered as part of standard clinical practice effectively improves inflammatory biomarkers in patients diagnosed with inflammatory bowel disease.

Assessment of Toxic Effects of Ochratoxin A in Human Embryonic Stem Cells.

Ochratoxin A (OTA) is a mycotoxin produced by different Aspergillus and Penicillium species, and it is considered a common contaminant in food and animal feed worldwide. On the other hand, human embryonic stem cells (hESCs) have been suggested as a valuable model for evaluating drug embryotoxicity. In this study, we have evaluated potentially toxic effects of OTA in hESCs. By using in vitro culture techniques, specific cellular markers, and molecular biology procedures, we found that OTA produces mild cytotoxic effects in hESCs by inhibiting cell attachment, survival, and proliferation in a dose-dependent manner. Thus, we suggest that hESCs provide a valuable human and cellular model for toxicological studies regarding preimplantation stage of human fetal development.

Progesterone anti-inflammatory properties in hereditary retinal degeneration.

The interactions between steroid gonadal hormones and the retina (a part of the visual system and the central nervous system (CNS)) have received limited attention and beneficial effects of these hormones in retinal diseases is controversial. Retinitis pigmentosa (RP) is the most common cause of retinal hereditary blindness and to date no treatment is available. However, results regarding the effects of progesterone on the progression of RP are promising. With the idea of demonstrating if the progesterone retinal protection in RP is related to its possible anti-inflammatory properties, we have administered orally progesterone to rd10 mice, an animal model of RP. We observed that progesterone decreased photoreceptors cell death, reactive gliosis and the increase in microglial cells caused by RP. We also examined the expression of neuronal and inducible nitric oxide synthase (nNOS and iNOS), the enzyme responsible for NO production. The results demonstrated a decrease in nNOS expression only in control mice treated with progesterone. Inflammation has been related with an increase in lipid peroxidation. Noticeably progesterone administration was able to diminish retinal malondialdehyde (MDA, a lipid peroxidation product) concentrations in rd10 mice. Altogether, we can conclude that progesterone could be a good therapeutic option not only in RP but also for other retinal diseases that have been associated with inflammation and lipid peroxidation.

Neuroprotection of Brain Cells by Lipoic Acid Treatment after Cellular Stress.

We have previously observed that in vivo lipoic acid (LA) treatment induced a protective effect onto primary cortical neurons after brain injury. In an effort to better understand LA action mechanism in the brain, in the present study, we stressed brain cells in vitro and ex vivo and then analyzed by inmmunocytochemistry and biochemical assays, the changes induced by LA on cell survival and on the concentration of oxidative stress markers, such as glutathione (GSH), oxidized glutathione (GSSG), and malondialdehyde (MDA). The stressors used were lipopolysaccharide (LPS), dopamine, and l-buthionine-S,R-sulfoximine (BSO). Our results showed that LA decreased cell death and increased GSH/GSSG ratio in cells stressed by LPS + dopamine, suggesting that the mechanism underlying LA action is regeneration of GSSG to GSH. When cells were stressed by BSO, LA diminished cell death and decreased GSH/GSSG ratio. In this case, it could be concluded that, due to the low GSH basal levels, GSSG reduction is not possible and therefore it might be thought that cell death prevention might be mediated through other mechanisms. Finally, we induced chemical oxidative damage in brain homogenate. After LA treatment, GSH and GSH/GSSG ratio increased and MDA concentration decreased, demonstrating again that LA was not able to increase de novo GSH synthesis but is able to increase GSSG conversion to GSH.

Combined application of polyacrylate scaffold and lipoic acid treatment promotes neural tissue reparation after brain injury.

PRIMARY OBJECTIVE: The aim of this study was to investigate the reparative potential of a polymeric scaffold designed for brain tissue repair in combination with lipoic acid. RESEARCH DESIGN: Histological, cytological and structural analysis of a combined treatment after a brain cryo-injury model in rats. METHODS AND PROCEDURES: Adult Wistar rats were subjected to cryogenic brain injury. A channelled-porous scaffold of ethyl acrylate and hydroxyethylacrylate, p(EA-co-HEA) was grafted into cerebral penumbra alone or combined with intraperitoneal LA administration. Histological and cytological evaluation was performed after 15 and 60 days and structural magnetic resonance (MRI) assessment was performed at 2 and 6 months after the surgery. MAIN OUTCOMES AND RESULTS: The scaffold was suitable for the establishment of different cellular types. The results obtained suggest that this strategy promotes blood vessels formation, decreased microglial response and neuron migration, particularly when LA was administrated. CONCLUSIONS: These evidences demonstrated that the combination of a channelled polymer scaffold with LA administration may represent a potential treatment for neural tissue repair after brain injury.

Evaluation of the pathogenic potential, antimicrobial susceptibility, and genomic relations of Yersinia enterocolitica strains from food and human origin.

Yersinia enterocolitica is a food-borne pathogen that causes gastroenteritis with occasional postinfection sequels. This study was aimed to determinate the pathogenic potential, antimicrobial susceptibility, and genomic relationships of Y. enterocolitica strains of different bioserotypes (B/O) isolated from foods and human samples in San Luis, Argentina. Strains obtained by culture were bioserotyped and characterized by phenotypic and genotypic virulence markers, antimicrobial susceptibility, and pulsed-field gel electrophoresis (PFGE). Yersinia enterocolitica was detected in 9.2% of 380 samples, with a distribution of 10.6% (30/284) for food products and 5.2% (5/96) for human samples. Regarding the pathogenic potential, B1A strains of different serotypes were virF(-) ail(-), of which 72.0% (13/18) were ystB(+) with virulence-related phenotypic characteristics. Among B2/O:9 isolates, 75.0% (9/12) exhibited the genotype virF(+) ail(+) ystB(-) along with phenotypic traits associated with virulence; the same genotype was observed in 80.0% (4/5) of B3/O:3 and B3/O:5 strains. By PFGE, it was possible to separate Y. enterocolitica biotypes into 4 clonal groups (A to D) with 23 genomic types, generating a discriminatory index of 0.96. All isolates were susceptible to antimicrobials used for clinical treatment. This study highlights the presence of pathogenic bioserotypes and the high genomic diversity of the Y. enterocolitica strains isolated in our region.

Neural tissue regeneration in experimental brain injury model with channeled scaffolds of acrylate copolymers.

The objective of the present study was to evaluate the biocompatibility and cell hosting ability of a copolymer scaffold based on ethyl acrylate (EA) and hydroxyl ethyl acrylate (HEA) in vivo after an experimental brain injury. Wistar rats were subjected to cryogenic traumatic brain injury. We evaluated the tissue response to the implanted materials after 8 weeks. The materials were implanted devoid of cells; they provoked a minimal scar response by the host tissue and permitted the invasion of neurons and glia inside them. We also found new blood vessels surrounding and inside the implant. Thus, the copolymer scaffold proves to offer a suitable environment producing a cellular network potentially useful in brain repair after brain injury.

Neurotoxic effects of ochratoxin A on the subventricular zone of adult mouse brain.

Ochratoxin A (OTA), a mycotoxin that was discovered as a secondary metabolite of the fungal species Aspergillus and Penicillium, is a common contaminant in food and animal feed. This mycotoxin has been described as teratogenic, carcinogenic, genotoxic, immunotoxic and has been proven a potent neurotoxin. Other authors have previously reported the effects of OTA in different structures of the central nervous system as well as in some neurogenic regions. However, the impact of OTA exposure in the subventricular zone (SVZ) has not been assessed yet. To elucidate whether OTA affects neural precursors of the mouse SVZ we investigated, in vitro and in vivo, the effects of OTA exposure on the SVZ and on the neural precursors obtained from this neurogenic niche. In this work, we prove the cumulative effect of OTA exposure on proliferation, differentiation and depletion of neural stem cells cultured from the SVZ. In addition, we corroborated these results in vivo by immunohistochemistry and electron microscopy. As a result, we found a significant alteration in the proliferation process, which was evidenced by a decrease in the number of 5-bromo-2-deoxyuridine-positive cells and glial cells, as well as, a significant decrease in the number of neuroblasts in the SVZ. To summarize, in this study we demonstrate how OTA could be a threat to the developing and the adult SVZ through its impact in cell viability, proliferation and differentiation in a dose-dependent manner.

Lipoic acid and bone marrow derived cells therapy induce angiogenesis and cell proliferation after focal brain injury.

UNLABELLED: Abstract Introduction: Traumatic brain injury is a main cause of disability and death in developed countries, above all among children and adolescents. The intrinsic inability of the central nervous system to efficiently repair traumatic injuries renders transplantation of bone marrow-derived cells (BMDC) a promising approach towards repair of brain lesions. On the other hand, many studies have reported the beneficial effect of Lipoic acid (LA), a potent antioxidant promoting cell survival, angiogenesis and neuroregeneration. METHODS: In this study, the cortex of adult mice was cryo-injured in order to mimic local traumatic brain injury. Vehicle or freshly prepared BMDC were grafted in the cerebral penumbra area 24 hours after unilateral local injury alone or combined with intra-peritoneal LA administration as a new regenerative strategy. RESULTS: Differences were found in the process of cell proliferation, angiogenesis and glial scar formation after local injury depending of the applied treatment, either LA or BMDC alone or in combination. CONCLUSION: The data presented here suggest that transplantation of BMDC is a good alternative and valid strategy to treat a focal brain injury when LA could not be prescribed due to its non-desired secondary effects.

Lipoic acid treatment after brain injury: study of the glial reaction.

After trauma brain injury, oxidative substances released to the medium provoke an enlargement of the initial lesion, increasing glial cell activation and, occasionally, an influx of immune cells into the central nervous system, developing the secondary damage. In response to these stimuli, microglia are activated to perform upregulation of intracellular enzymes and cell surface markers to propagate the immune response and phagocytosis of cellular debris. The phagocytosis of debris and dead cells is essential to limit the inflammatory reaction and potentially prevent extension of the damage to noninjured regions. Lipoic acid has been reported as a neuroprotectant by acting as an antioxidant and anti-inflammatory agent. Furthermore, angiogenic effect promoted by lipoic acid has been recently shown by our group as a crucial process for neural regeneration after brain injury. In this work, we focus our attention on the lipoic acid effect on astroglial and microglial response after brain injury.

Biomaterials coated by dental pulp cells as substrate for neural stem cell differentiation.

This study is focused on the development of an in vitro hybrid system, consisting in a polymeric biomaterial covered by a dental pulp cellular stroma that acts as a scaffold offering a neurotrophic support for the subsequent survival and differentiation of neural stem cells. In the first place, the behavior of dental pulp stroma on the polymeric biomaterial based on ethyl acrylate and hydroxy ethyl acrylate copolymer was studied. For this purpose, cells from normal human third molars were grown onto 0.5-mm-diameter biomaterial discs. After cell culture, quantification of neurotrophic factors generated by the stromal cells was performed by means of an ELISA assay. In the second place, survival and differentiation of adult murine neural stem cells on the polymeric biomaterials covered by dental pulp stromal cells was studied. The results show the capacity of dental pulp cells to uniformly coat the majority of the material's surface and to secrete neurotrophic factors that become crucial for a subsequent differentiation of neural stem cells. The use of stromal cells cultured on scaffolding biomaterials provides neurotrophic pumps that may suggest new criteria for the design of cell therapy experiments in animal models to assist the repair of lesions in Central Nervous System.

Three-dimensional scaffolds as a model system for neural and endothelial 'in vitro' culture.

Biomaterials based on the hydrophobic homopolymer poly(ethyl acrylate), PEA, and its copolymers with hydroxyethyl acrylate, p(EA-co-HEA) and methacrylic acid, p(EA-co-MAAc) were prepared as polymeric scaffolds with interconnected pores of 90 microns and tested in vitro as culture substrates and compared for their impact on the differentiation of neural stem cells (NSC) obtained from the subventricular zone (SVZ) of postnatal rats and human endothelial cells (HUVEC). Immunocytochemical staining assay for specific markers show that p(EA-co-MAAc) scaffolds were suitable substrates to promote cell attachment and differentiation of adult NSC and HUVEC cells.

Differentiation of postnatal neural stem cells into glia and functional neurons on laminin-coated polymeric substrates.

A series of polymeric biomaterials, including poly(methyl acrylate), chitosan, poly(ethyl acrylate) (PEA), poly(hydroxyethyl acrylate) (PHEA), and a series of random copolymers containing ethyl acrylate, hydroxyethyl acrylate, and methyl acrylate were tested in vitro as culture substrates and compared for their effect on the differentiation of neural stem cells (NSCs) obtained from the subventricular zone of postnatal rats. Immunocytochemical assay for specific markers and scanning electron microscopy techniques were employed to determine the adhesion of the cultured NSCs to the different biomaterials and the respective neuronal differentiation. The functional properties and the membrane excitability of differentiated NSCs were investigated using a patch-clamp. The results show that the substrate's surface chemistry influences cell attachment and neuronal differentiation, probably through its influence on adsorbed laminin, and that copolymers based on PEA and PHEA in a narrow composition window are suitable substrates to promote cell attachment and differentiation of adult NSCs into functional neurons and glia.

Substrate chemistry-dependent conformations of single laminin molecules on polymer surfaces are revealed by the phase signal of atomic force microscopy.

The conformation of single laminin molecules adsorbed on synthetic substrates is directly observed making use of the phase magnitude in tapping mode atomic force microscopy (AFM). With AFM, it is not possible to differentiate the proteins on the substrate if use is made of the height signal, since the roughness of the material becomes of the same order of magnitude as the adsorbed protein, typically 10 nm height. This work shows how AFM can be exploited to reveal protein conformation on polymer materials. Different laminin morphologies are observed on a series of different copolymers based on ethyl acrylate and hydroxyethyl acrylate as a function of the surface density of -OH groups: from globular to completely extended morphologies of the protein molecules are obtained, and the onset of laminin network formation on some substrates can be clearly identified. The results stress the importance of the underlying synthetic substrate's surface chemistry for the biofunctional conformation of adsorbed proteins.

Survival and differentiation of embryonic neural explants on different biomaterials.

Biomaterials prepared from polyacrylamide, ethyl acrylate (EA), and hydroxyethyl acrylate (HEA) in various blend ratios, methyl acrylate and chitosan, were tested in vitro as culture substrates and compared for their ability to be colonized by the cells migrating from embryonic brain explants. Neural explants were isolated from proliferative areas of the medial ganglionic eminence and the cortical ventricular zone of embryonic rat brains and cultured in vitro on the different biomaterials. Chitosan, poly(methyl acrylate), and the 50% wt copolymer of EA and HEA were the most suitable substrates to promote cell attachment and differentiation of the neural cells among those tested. Immunofluorescence microscopy analysis showed that progenitor cells had undergone differentiation and that the resulting glial and neuronal cells expressed their intrinsic morphological characteristics in culture.

Defective postnatal neurogenesis and disorganization of the rostral migratory stream in absence of the Vax1 homeobox gene.

The subventricular zone (SVZ) is one of the sources of adult neural stem cells (ANSCs) in the mouse brain. Precursor cells proliferate in the SVZ and migrate through the rostral migratory stream (RMS) to the olfactory bulb (OB), where they differentiate into granule and periglomerular cells. Few transcription factors are known to be responsible for regulating NSC proliferation, migration, and differentiation processes; even fewer have been found to be responsible for the organization of the SVZ and RMS. For this reason, we studied the ventral anterior homeobox (Vax1) gene in NSC proliferation and in SVZ organization. We found that Vax1 is strongly expressed in the SVZ and in the RMS and that, in the absence of Vax1, embryonic precursor cells proliferate 100 times more than wild-type controls, in vitro. The SVZ of Vax1(-/-) brains is hyperplastic and mostly disorganized, and the RMS is missing, causing a failure of precursor cell migration to the OBs, which as a result are severely hypoplastic. Moreover, we found that Vax1 is essential for the correct differentiation of ependyma and astrocytes. Together, these data indicate that Vax1 is a potent regulator of SVZ organization and NSC proliferation, with important consequences on postnatal neurogenesis.

Compromised generation of GABAergic interneurons in the brains of Vax1-/- mice.

The subcortical telencephalon is the major source of GABAergic interneurons that, during development, tangentially migrate to the cerebral cortex, where they modulate the glutamatergic excitatory action of pyramidal cells. The transcription factor Vax1, an intracellular mediator of both Shh and Fgf signaling, is expressed at high levels in the medial and lateral ganglionic eminences (MGE and LGE, respectively), in the septal area (SA), in the anterior entopeduncular area (AEP) and in the preoptic area (POA). We show that Vax1 expression in the neuroepithelium is graded: low in the ventricular zone (VZ) and high in the subventricular zone (SVZ), in a pattern that closely reproduces that of several members of the Dlx and Gsh family of homeobox transcription factors. We provide evidence that Vax1 plays an important role in proliferation and differentiation of MGE, POA/AEP and septum, and that the last structure is completely absent in Vax1-/- mice. We show that the absence of Vax1 causes a severe depletion of GABAergic neurons in the neocortex, ranging from 30% to 44%, depending on the cortical areas considered. Taken together, our data indicate that a loss of function mutation in the Vax1 gene generates abnormalities in basal ganglia subventricular zone development and that it prevents the formation of the septum, impairing GABAergic interneuron generation.