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Eight horses were allotted into pairs consisting of one cecum- and right ventral colon-fistulated animal and one cecum-fistulated animal. They were fed daily at the same level of intake either a high-fiber (HF) or a high-starch (HS) diet without or with 10 g of a Saccharomyces cerevisiae preparation, in a 4 x 4 Latin square design. The HS diet provided a starch overload (i.e., 3.4 g starch*[kg.sup.-1] BW*[meal.sup.-1] while maintaining a high amount of fiber intake (i.e., dietary NDF/ starch ratio was 1.0). A 21-d period of adaptation to the treatments occurred before cecal and colonic contents were withdrawn 4 h after the morning meal to count total anaerobic, cellulolytic, and lactic acid-utilizing bacteria, lactobacilli, and streptococci. Lactic acid, volatile fatty acids, ammonia concentrations, and pH were measured on cecal and colonic fluid samples collected hourly during the first 12-h postfeeding. When the HS diet was fed, the concentration of total anaerobic and lactic acid-utilizing bacteria increased (P < 0.001), whereas that of cellulolytic bacteria decreased (P < 0.05) in the cecum. The concentration of lactobacilli and streptococci increased (P < 0.001) in the cecal and colonic contents. These alterations of the microbial profiles were associated with decreases (P < 0.001) of pH, (acetate + butyrate)/propionate ratio and with an increase (P < 0.001) of lactic acid concentration. Supplementing the S. cerevisiae preparation increased (P < 0.01) the concentration of viable yeast cells, averaging 4.3 x [10.sup.6] and 4.5 x [10.sup.4] cfu/mL in the cecal and colonic contents, respectively. Yeast supplementation had almost no effect on microbial counts in the cecum and colon. The supplementation of S. cerevisiae appeared to modify (P < 0.05) pH, concentrations of lactic acid and ammonia, molar percentages of acetate and butyrate with the HS diet and [(acetate + butyrate)/propionate] ratio when the HF diet was fed. The effects of the S. cerevisiae preparation were greater in the cecum than in the colon, which coincided with the abundance of yeast cells. When the digestion of starch in the small intestine was saturated, the effect of the addition of a S. cerevisiae preparation appeared to limit the extent of undesirable changes in the intestinal ecosystem of the horse. Key Words: Cellulolytic Microorganisms, Horses, Large Intestine Fermentation, Saccharomyces cerevisiae, Starch Digestion

The HIV-1 encoded apoptogenic protein Vpr induces mitochondrial membrane permeabilization (MMP) via interactions with the voltage-dependent anion channel (VDAC) and the adenine nucleotide translocator (ANT). We have designed a peptide, TEAM-VP, composed of two functional domains, one a tumor blood vessel RGD-like ‘homing’ motif and the other an MMP-inducing sequence derived from Vpr. When added to isolated mitochondria, TEAM-VP interacts with ANT and VDAC, reduces oxygen consumption and overcomes Bcl-2 protection to cause inner and outer MMP. TEAM-VP specifically recognizes cell-surface expressed αVβ3 integrins, internalizes, temporarily localizes to lysosomes and progressively co-distributes with the mitochondrial compartment with no sign of lysosomal membrane permeabilization. Finally TEAM-VP reaches mitochondria of angiogenic endothelial cells to induce mitochondrial fission, dissipation of the mitochondrial transmembrane potential (ΔΨm), cytochrome c release and apoptosis hallmarks. Hence, this chimeric peptide constitutes the first example of a virus-derived mitochondriotoxic compound as a candidate to kill selectively tumor neo-endothelia.Cell Death and Differentiation (2007) 14, 422–435. doi:10.1038/sj.cdd.4402018; published online 4 August 2006 [ABSTRACT FROM AUTHOR]

A letter to the editor on the use of broad-spectrum caspase inhibitors in elucidating apoptotic cell death pathways is presented.

A first group of three horses was given diet 1 (D1) allowing 1180 g per 100 kg body weight (BW) of a pelleted food rich in fibre (P1) and 556 g per 100 kg BW of straw during a 20-day period to allow for adaptation. A second group of four horses were given diet 2 (D2) allowing 1180 g per 100 kg BW of a pelleted food rich in cereals (P2) and 1000 g per 100 kg BW of meadow hay during the same period. Digesta was collected from the stomach, duodenum, jejunum, ileum, caecum, right ventral colon, left ventral colon, left dorsal colon, right dorsal colon, and small colon, and faeces were collected under general anaesthesia 2.5 h after the ingestion of the morning pelleted meal. The concentration of total anaerobic, cellulolytic and lactic acid-utilizing bacteria, lactobacilli and streptococci were determined in all these segments except for the duodenum, left ventral colon, right dorsal colon and small colon. D-/L-lactic acid, volatile fatty acids and pH were measured in all anatomic segments of the digestive tract (from stomach to small colon). The caecal concentration of total anaerobic bacteria was the lowest (7.9 x 10[sup7] colony-forming units (c. f. u. ) per ml), whereas that of the stomach was the highest (1.4 x 10[sup9] c. f. u. per ml) (P < 0.001). Cellulolytic bacteria did not exceed 3.0 x 10² c. f. u. per ml in the ante-caecal segments whereas in the hindgut the average concentration was 5.3 x 10[sup5] c. f. u. per ml (P < 0.001). Likewise, VFA concentrations were also greater in the large intestine (on average, 96.3 mmol/l v. 8.8 mmol/l in the ante-caecal segments) (P <... [ABSTRACT FROM AUTHOR]

Mitochondrial membrane permeabilization can be a rate limiting step of apoptotic as well as necrotic cell death. Permeabilization of the outer mitochondrial membrane (OM) and/or inner membrane (IM) is, at least in part, mediated by the permeability transition pore complex (PTPC). The PTPC is formed in the IM/OM contact site and contains the two most abundant IM and OM proteins, adenine nucleotide translocator (ANT, in the IM) and voltage-dependent anion channel (VDAC, in the OM), the matrix protein cyclophilin D, which can interact with ANT, as well as apoptosis-regulatory proteins from the Bax/Bcl-2 family. Here we discuss that ANT has two opposite functions. On the one hand, ANT is a vital, specific antiporter which accounts for the exchange of ATP and ADP on IM. On the other hand, ANT can form a non-specific pore, as this has been shown by electrophysiological characterization of purified ANT reconstituted into synthetic lipid bilayers or by measuring the permeabilization of proteoliposomes containing ANT. Pore formation by ANT is induced by a variety of different agents (e.g. Ca[sup 2+], atractyloside, thiol oxidation, the pro-apoptotic HIV-1 protein Vpr, etc.) and is enhanced by Bax and inhibited by Bcl-2, as well as by ADP. In isolated mitochondria, pore formation by ANT leads to an increase in IM permeability to solutes up to 1500 Da, swelling of the mitochondrial matrix, and OM permeabilization, presumably due to physical rupture of OM. Although alternative mechanisms of mitochondrial membrane permeabilization may exist, ANT emerges as a major player in the regulation of cell death. [ABSTRACT FROM AUTHOR]

Mitochondrial membrane permeabilization is a critical event in the process leading to physiologic or chemotherapy-induced apoptosis (programmed cell death). This permeabilization event is, at least in part, under the control of the permeability transition pore complex (PTPC). Oncoproteins from the Bcl-2 family and tumor suppressor proteins from the Bax family interact with PTPC to inhibit or facilitate membrane permeabilization, respectively. Conventional chemotherapeutic agents elicit mitochondrial permeabilization in an indirect fashion by induction of endogenous effectors that are involved in the physiologic control of apoptosis. However, an increasing number of experimental anticancer drugs, including lonidamine, arsenite, betulinic acid, CD437, and several amphipathic cationic alpha-helical peptides, act directly on mitochondrial membranes and/or on the PTPC. Such agents may induce apoptosis in circumstances in which conventional drugs fail to act because endogenous apoptosis induction pathways, such as those involving p53, death receptors, or apical caspase activation, are disrupted. However, stabilization of the mitochondrial membrane by antiapoptotic Bcl-2-like proteins reduces the cytotoxic potential of most of these drugs. Targeting of specific PTPC components may overcome this Bcl-2-mediated apoptosis inhibition. One strategy involves cross-linking of critical redox-sensitive thiol groups within the PTPC; another involves the use of ligands to the mitochondrial benzodiazepine receptor. Thus, the design of mitochondrion-targeted cytotoxic drugs may constitute a novel strategy for overcoming apoptosis resistance. [ABSTRACT FROM AUTHOR]

Examines the capacity of HIV-1-envelope glycoprotein complex to induce apoptosis and karyogamy in syncytia. Mechanisms accounting for HIV-1-induced lypmhodepletion; Capacity of the envelope glycoprotein complex expressed on HIV-1 infected cells to interact with CD4 and a suitable coreceptor; Functional relationship between syncitium formation, karyogamy and apoptosis.

Despite their low quantity and abundance, the cellulolytic bacteria that inhabit the equine large intestine are vital to their host, as they enable the crucial use of forage-based diets. Fibrobacter succinogenes is one of the most important intestinal cellulolytic bacteria. In this study, Fibrobacter sp. HC4, one cellulolytic strain newly isolated from the horse cecum, was characterized for its ability to utilize plant cell wall fibers. Fibrobacter sp. HC4 consumed only cellulose, cellobiose, and glucose and produced succinate and acetate in equal amounts. Among genes coding for CAZymes, 26% of the detected glycoside hydrolases (GHs) were involved in cellulolysis. These cellulases belong to the GH5, GH8, GH9, GH44, GH45, and GH51 families. Both carboxymethyl cellulase and xylanase activities of Fibrobacter sp. HC4 were detected using the Congo red method and were higher than those of F. succinogenes S85, the type strain. The in vitro addition of Fibrobacter sp. HC4 to a fecal microbial ecosystem of horses with large intestinal acidosis significantly enhanced fibrolytic activity as measured by the increase in gas and volatile fatty acids production during the first 48 h. According to this, the pH decreased and the disappearance of dry matter increased at a faster rate with Fibrobacter sp. HC4. Our data suggest a high specialization of the new strain in cellulose degradation. Such a strain could be of interest for future exploitation of its probiotic potential, which needs to be further determined by in vivo studies.IMPORTANCECellulose is the most abundant of plant cell wall fiber and can only be degraded by the large intestine microbiota, resulting in the production of volatile fatty acids that are essential for the host nutrition and health. Consequently, cellulolytic bacteria are of major importance to herbivores. However, these bacteria are challenged by various factors, such as high starch diets, which acidify the ecosystem and reduce their numbers and activity. This can lead to an imbalance in the gut microbiota and digestive problems such as colic, a major cause of mortality in horses. In this work, we characterized a newly isolated cellulolytic strain, Fibrobacter sp. HC4, from the equine intestinal microbiota. Due to its high cellulolytic capacity, reintroduction of this strain into an equine fecal ecosystem stimulates hay fermentation in vitro . Isolating and describing cellulolytic bacteria is a prerequisite for using them as probiotics to restore intestinal balance., Competing Interests: The authors declare no conflict of interest.

High-fat diets alter gut barrier integrity, leading to endotoxemia by impacting epithelial functions and inducing endoplasmic reticulum (ER) stress in intestinal secretory goblet cells. Indeed, ER stress, which is an important contributor to many chronic diseases such as obesity and obesity-related disorders, leads to altered synthesis and secretion of mucins that form the protective mucus barrier. In the present study, we investigated the relative contribution of omega-3 polyunsaturated fatty acid (PUFAs)-modified microbiota to alleviating alterations in intestinal mucus layer thickness and preserving gut barrier integrity. Male fat-1 transgenic mice (exhibiting endogenous omega-3 PUFAs tissue enrichment) and wild-type (WT) littermates were fed either an obesogenic high-fat diet (HFD) or a control diet. Unlike WT mice, HFD-fed fat-1 mice were protected against mucus layer alterations as well as an ER stress-mediated decrease in mucin expression. Moreover, cecal microbiota transferred from fat-1 to WT mice prevented changes in the colonic mucus layer mainly through colonic ER stress downregulation. These findings highlight a novel feature of the preventive effects of omega-3 fatty acids against intestinal permeability in obesity-related conditions.

Fibrobacter succinogenes is a major cellulolytic bacterial species living in the large intestines of herbivores. This study reports the genome sequencing, assembly, and annotation of F. succinogenes HC4 (DSM 33656), a strain isolated from horse cecal contents. The genome comprised a total of 3.74 Mbp, with a G+C content of 48.96%.

Large intestine barrier disturbances can have serious consequences for the health of horses. The loss of mucosal integrity that leads to increased intestinal permeability may result from a local inflammatory immune response following alterations of the microbiota, known as dysbiosis. Therefore, our research aimed to identify noninvasive biomarkers for studying the intestinal permeability and the local inflammatory immune response in horses. Regarding the biomarkers used in other mammalian species, we measured the concentrations of lipopolysaccharides (LPS), reflected by 3-OH C14, C16, and C18 fatty acids, in blood, and fecal secretory immunoglobulin-A (SIgA). These biomarkers were evaluated in two trials including 9 and 12 healthy horses, which developed large intestinal dysbiosis experimentally induced by 5 d of antibiotic administration (trimethoprim sulfadiazine [TMS]) or 5 d of abrupt introduction of high starch levels (barley) into the diet. Horses were either control or supplemented with Lactobacillus acidophilus, Ligilactobacillus salivarius, and Bifidobacterium lactis. Correlations were performed between biomarkers and fecal bacterial diversity, composition, and function. No significant interaction between day and supplementation, or supplementation effect were observed for each biomarker. However, with the dietary stressor, a significant increase in blood concentrations of 3-OH C16 (P = 0.0125) and C14 (P = 0.0252) fatty acids was measured 2 d after the cessation of barley administration. Furthermore, with the antibiotic stressor, blood levels of 3-OH C16 progressively increased (P = 0.0114) from the first day to 2 d after the end of TMS administration. No significant day effect was observed for fecal SIgA concentrations for both stressors. These results indicate that both antibiotic- and diet-induced dysbiosis resulted in a local translocation of LPS 2 d after the cessation of the stressor treatments, suggesting an impairment of intestinal permeability, without detectable local inflammation. Blood LPS and fecal SIgA concentrations were significantly correlated with several bacterial variations in the large intestine, which are features of antibiotic- and diet-induced dysbiosis. These findings support the hypothesis that a relationship exists between dysbiosis and the loss of mucosal integrity in the large intestine of horses., (© The Author(s) 2022. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

NAD + depletion is a common phenomenon in neurodegenerative pathologies. Excitotoxicity occurs in multiple neurologic disorders and NAD + was shown to prevent neuronal degeneration in this process through mechanisms that remained to be determined. The activity of nicotinamide riboside (NR) in neuroprotective models and the recent description of extracellular conversion of NAD + to NR prompted us to probe the effects of NAD + and NR in protection against excitotoxicity. Here, we show that intracortical administration of NR but not NAD + reduces brain damage induced by NMDA injection. Using cortical neurons, we found that provision of extracellular NR delays NMDA-induced axonal degeneration (AxD) much more strongly than extracellular NAD + Moreover, the stronger effect of NR compared to NAD + depends of axonal stress since in AxD induced by pharmacological inhibition of nicotinamide salvage, both NAD + and NR prevent neuronal death and AxD in a manner that depends on internalization of NR. Taken together, our findings demonstrate that NR is a better neuroprotective agent than NAD + in excitotoxicity-induced AxD and that axonal protection involves defending intracellular NAD + homeostasis.-Vaur, P., Brugg, B., Mericskay, M., Li, Z., Schmidt, M. S., Vivien, D., Orset, C., Jacotot, E., Brenner, C., Duplus, E. Nicotinamide riboside, a form of vitamin B 3 , protects against excitotoxicity-induced axonal degeneration., (© FASEB.)

Introduction: Recent histopathological studies have shown that neurodegenerative processes in Alzheimer's and Parkinson's Disease develop along neuronal networks and that hallmarks could propagate trans-synaptically through neuronal pathways. The underlying molecular mechanisms are still unknown, and investigations have been impeded by the complexity of brain connectivity and the need for experimental models allowing a fine manipulation of the local microenvironment at the subcellular level., Results: In this study, we have grown primary cortical mouse neurons in microfluidic (μFD) devices to separate soma from axonal projections in fluidically isolated microenvironments, and applied β-amyloid (Aβ) peptides locally to the different cellular compartments. We observed that Aβ application to the somato-dendritic compartment triggers a "dying-back" process, involving caspase and NAD(+) signalling pathways, whereas exposure of the axonal/distal compartment to Aβ deposits did not induce axonal degeneration. In contrast, co-treatment with somatic sub-toxic glutamate and axonal Aβ peptide triggered axonal degeneration. To study the consequences of such subcellular/local Aβ stress at the network level we developed new μFD multi-chamber devices containing funnel-shaped micro-channels which force unidirectional axon growth and used them to recreate in vitro an oriented cortico-hippocampal pathway. Aβ application to the cortical somato-dendritic chamber leads to a rapid cortical pre-synaptic loss. This happens concomitantly with a post-synaptic hippocampal tau-phosphorylation which could be prevented by the NMDA-receptor antagonist, MK-801, before any sign of axonal and somato-dendritic cortical alteration., Conclusion: Thanks to μFD-based reconstructed neuronal networks we evaluated the distant effects of local Aβ stress on neuronal subcompartments and networks. Our data indicates that distant neurotransmission modifications actively take part in the early steps of the abnormal mechanisms leading to pathology progression independently of local Aβ production. This offers new tools to decipher mechanisms underlying Braak's staging. Our data suggests that local Aβ can play a role in remote tauopathy by distant disturbance of neurotransmission, providing a putative mechanism underlying the spatiotemporal appearance of pretangles.

This study assessed the time-scale variability of bacterial community structure in foal feces from birth to 365 days of age using Automated Ribosomal Intergenic Spacer Analysis (ARISA). Fecal samples were collected from five foals 2 h after birth (meconium) and in the morning at days 1, 2, 5, 10, 30, 60, 120, 179, 183, 194 and 365. The ARISA profiles were compared using an analysis of similarity (ANOSIM). Although both the age effect and the foal effect were highly significant (P < 0.010), the R-ANOSIM value for the foal effect was very low (R-ANOSIM = 0.089), while that of the age effect was much higher (R-ANOSIM = 0.309). Significant age-related changes were detected between days 0 and 2 (R-ANOSIM = 0.500), days 2 and 10 (R-ANOSIM = 0.475) and days 10 and 30 (R-ANOSIM = 0.519). No further shifts between consecutive times of sampling were detected in the bacterial community after day 30 and no changes were observed at weaning (day 180). These results show that the establishment of the intestinal bacterial community in foals is a sequential process, which reaches its climax state at around one month of age. Further studies using new generation sequencing based methods could be conducted to identify which bacterial genera are establishing in foals during the first month of life., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Microglia mediate multiple facets of neuroinflammation, including cytotoxicity, repair, regeneration, and immunosuppression due to their ability to acquire diverse activation states, or phenotypes. Modulation of microglial phenotype is an appealing neurotherapeutic strategy but a comprehensive study of classical and more novel microglial phenotypic markers in vitro is lacking. The aim of this study was to outline the temporal expression of a battery of phenotype markers from polarised microglia to generate an in vitro tool for screening the immunomodulatory potential of novel compounds. We characterised expression of thirty-one macrophage/microglial phenotype markers in primary microglia over time (4, 12, 36, and 72 h), using RT-qPCR or multiplex protein assay. Firstly, we selected Interleukin-4 (IL-4) and lipopolysaccharide (LPS) as the strongest M1-M2 polarising stimuli, from six stimuli tested. At each time point, markers useful to identify that microglia were M1 included iNOS, Cox-2 and IL-6 and a loss of M2a markers. Markers useful for quantifying M2b-immunomodulatory microglia included, increased IL-1RA and SOCS3 and for M2a-repair and regeneration, included increased arginase-1, and a loss of the M1 and M2b markers were discriminatory. Additional markers were regulated at fewer time points, but are still likely important to monitor when assessing the immunomodulatory potential of novel therapies. Further, to facilitate identification of how novel immunomodulatory treatments alter the functional affects of microglia, we characterised how the soluble products from polarised microglia affected the type and rate of neuronal death; M1/2b induced increasing and M2a-induced decreasing neuronal loss. We also assessed any effects of prior activation state, to provide a way to identify how a novel compound may alter phenotype depending on the stage of injury/insult progression. We identified generally that a prior M1/2b reduced the ability of microglia to switch to M2a. Altogether, we have characterised a profile of phenotype markers and a mechanism of assessing functional outcome that we can use as a reference guide for first-line screening of novel immunomodulatory therapies in vitro in the search for viable neuroprotectants., (Copyright © 2013 Elsevier Inc. All rights reserved.)