Cigarette smoke and the terminal ileum: increased autophagy in murine follicle-associated epithelium and Peyer’s patches

Cigarette smoke (CS) exposure is associated with increased autophagy in several cell types, such as bronchial epithelial cells. Smoking is also an environmental risk factor in Crohn’s disease, in which impairment of the autophagy-mediated anti-bacterial pathway has been implicated. So far, it is unknown whether CS induces autophagy in the gut. Here, we examined the effect of chronic CS exposure on autophagy in the follicle-associated epithelium (FAE) of murine Peyer’s patches. Transmission electron microscopy revealed that the proportion of cell area occupied by autophagic vesicles significantly increased in the FAE after CS exposure. An increased number of autophagic vesicles was observed in the FAE, whereas the vesicle size remained unaltered. Besides enterocytes, also M-cells contain more autophagic vesicles upon CS exposure. In addition, the mRNA level of the autophagy-related protein Atg7 in the underlying Peyer’s patches is increased after CS exposure, which indicates that the autophagy-inducing effect of CS is not limited to the FAE. In conclusion, our results demonstrate that CS exposure induces autophagy in murine FAE and in the underlying immune cells of Peyer’s patches, suggesting that CS exposure increases the risk for Crohn’s disease by causing epithelial oxidative damage, which needs to be repaired by autophagy.     

IL-10 secreting CD21+ B cells are present in jejunal Peyer’s patches of sheep during fetal development

We recently reported a novel interleukin-10 (IL-10)-secreting CD21⁺ B cell population in jejunal Peyer’s patches (JPP) of sheep with a regulatory function (B_regs) suppressing Toll-like receptor 9 (TLR9)-induced cytokine responses. However, little is known about the development of these cells. Therefore, we investigate their existence in JPP cells from fetal and newborn lambs. CD21⁺ B cells were purified from JPP cells by magnetic cell sorting and subsequently stimulated with the TLR9 agonist, CpG ODN (CpG oligodeoxynucleotide). Lymphocyte proliferative responses, cytokine production (IL-10, IL-12 and interferon-γ [INF-γ]) and antibody secretion were assayed. We found that fetal and neonatal CD21⁺ B cells spontaneously secreted high levels of IL-10 regardless of CpG stimulation but that these cells did not produce any IL-12 or INF-γ upon stimulation with CpG. The observed responses are consistent with those previously reported for B_regs characterized in JPP of older lambs. Surprisingly, unlike in older lambs, fetal and neonatal JPP CD21⁺ B cells proliferated in response to CpG stimulation. Our investigations of fetal and neonatal lambs provide evidence for the development of IL-10-secreting CD21⁺ B cells in PPs prior to antigen exposure.     

Distribution of immune cells and expression of interleukin receptors in ileal Peyer’s patches of calves

Newborn calves lack a mature immune system. The immune system develops with age, but the role of the expression of cytokine receptors in the development of immune cells of Peyer’s patches (PPs) in the intestines of calves in the first 2 months has not yet been elucidated. In this study, the distribution of immune cells and the expression of interleukin (IL) receptors (R) in the ileal PPs of newborn and 2-month-old calves were investigated immunohistochemically with monoclonal antibodies against bovine CD4, CD8, IgM, γδTCR, T19, WC3, WC5, and WC6 antigens. The expression of ILRs was examined with antibodies against CD25 (IL-2Rα), IL-2Rγ, IL-4R, IL-6R, IL-10R, and IL-13R antigens. CD4⁺, CD8⁺, γδTCR⁺, T19⁺, and WC6⁺ cells were found to be more widely distributed in the ileal PPs of 2-month-old calves than in those of newborn calves. Moreover, the expression of CD25 (IL-2Rα), IL-4R, and IL-13R in the ileal PPs of 2-month-old calves was more prominent than that in newborn calves. These data suggest that the immune system of calves at 2 months of age is developed by reactions to foreign antigens and aging.     

Uptake and transport of foreign particles in Peyer’s patches of both distal ileum and jejunum of calves

To investigate the uptake and transport patterns of variously sized particles in Peyer’s patches (PPs) of calves, intestinal loops were created in four newborn and two 2-month-old calves, and the loops were inoculated with various particles, including carbon black, fluorescein isothiocyanate (FITC)-labeled latex, FITC-labeled dextran, bovine serum, and recombinant mouse prion protein (rMPrP). The intestinal loops were recovered at 3, 6, 9, and 24 h in newborn calves and at 24 h in 2-month-old calves after inoculation, and the transport of the particles was examined by histological and immunohistochemical means. The uptake of the particles was quantified by estimation of signal intensities. A greater intensity was found in newborn calves compared with the 2-month-old calves. The peak uptake of carbon black, FITC-labeled latex, and rMPrP in the PPs of the distal ileum occurred at 6 h after inoculation in newborn calves and then progressively decreased with time. Uptake was also dependent on the site within the small intestine and the size of the particle studied. The transport of carbon black, FITC-labeled latex, and FITC-labeled dextran occurred via the bloodstream, the mesenteric lymph nodes, and the liver of newborn calves. rMPrP was found primarily in the interfollicular regions of the submucosa of the distal ileum of newborn calves. Thus, distal ileal PPs are probably more effective at particle absorption than the jejunal PPs, and the peak uptake of the PPs within the newborn calf occurs at 6 h after inoculation. This study was partly supported by a grant for BSE research from the Ministry of Health, Labour, and Welfare, Japan (17270701) and Grant-in-Aid (no. 17380180) from the Ministry of Education, Culture, Sports, Science, and Technology, Japan.     

Peyer’s Patches and the Follicle-Associated Epithelium in Diarrheic Calves

Twelve 2–5-week-old calves affected with a spontaneous intestinal disorder were examined; 8 had diarrhea and 4 were convalescents. In all the affected calves the “pseudovilli” (syn. domes or lymphoid villi) over Peyer’s patches seemed atrophic and appeared enclosed within the mucosa, owing to fusion of ordinary villi with “pseudovilli”. Morphometric examination showed a decrease of lymphoid follicle length in the affected calves as compared with controls (P < 0.01). Convalescents showed longer follicles than diarrheic calves (P<0.05). Often cytoplasmic acid phosphatase of the follicle-associated epithelium (FAE) in affected calves did not show the marked basal-apical decrease along “pseudovillus”, typical of the controls. Scanning electron microscopy revealed sparse development of concentric folds in the luminal plasma membrane of the enclosed FAE, contrasting with their abundance in the normal FAE. Transmission electron microscopy showed that the “pseudovilli” had increased numbers of ordinary villous epithelial cells. Affinity of chlamydia for FAE was shown. It is suggested that the sparse occurrence of surface folds in the FAE and the change in acid phosphatase distribution indicate diminished endocytosis of antigenic material, probably resulting from the enclosure of “pseudovilli”. The atrophy of lymphoid follicles may be another expression of the probable decreased contact with the intestinal contents.     

Do small and large luteal cells of the sheep interact in the production of progesterone?

Corpora lutea from cyclic ewes were dissociated by collagenase and trypsin/EGTA treatments, and enriched fractions of small and large luteal cells were prepared on gradients of Ficoll. These fractions were incubated separately or remixed before incubation. Colchicine, cytochalasin B and the calcium channel-blocker verapamil significantly reduced progesterone production by both small and large luteal cell fractions, while isoprenaline stimulated an increase in progesterone production by large luteal cell fractions only. When fractions of small and large luteal cells were remixed, no more and no less progesterone was produced than would have been predicted from equivalent fractions incubated separately. There was therefore no evidence of synergism between small and large luteal cells in the production of progesterone. Prostaglandin F-2 alpha, which can inhibit LH-stimulated progesterone production by ovine luteal tissue in vitro, had no effect on LH-stimulated progesterone production by small luteal cell fractions, but significantly inhibited that by enriched fractions of large luteal cells. Since large luteal cell fractions were contaminated with small luteal cells, which are probably responsible for the progesterone-secretory response of these fractions to LH, it was concluded that the inhibition of LH-stimulated progesterone production by small luteal cells is dependent on the presence of large luteal cells. Oxytocin added to large and small luteal cell fractions did not affect progesterone production by either fraction. It was therefore concluded that the inhibitory action of PGF-2 alpha on LH-stimulated progesterone production may require the interaction of large and small luteal cells, but that oxytocin is not likely to be an intermediary in this interaction.     

Identification of intestinal M cells in isolated lymphoid follicles and Peyer’s patches of the Angora rabbit

The presence, distribution, and localization of M cells in isolated lymphoid follicles (ILF) and in follicle-associated epithelia (FAE) covering Peyer’s patches (PP) in Angora rabbits were investigated by immunohistochemistry and electron microscopy. Although PP could macroscopically be identified along the length of the mucosal and serosal surfaces of jejunum and ileum, the presence of ILF could only be located microscopically. Typical M cells in FAE were detected within the periphery of the dome regions of the PP, and immature columnar M cells in the FAE resided in the vicinity of the crypts. M cells in the FAE of both ILF and PP showed vimentin-positive reaction. M cells in the FAE of ILF were morphologically similar to the immature M cells found in the FAE of PP. Typical mature M cells were also observed in the FAE of a few ILF. In contrast to FAE of PP, numerous goblet cells were observed in the FAE of many ILF. Moreover, among intestinal villi, we noticed villi-like solitary lymphoid structures that showed abundant lymphocytes in their lamina propria and that were surrounded with vimentin-positive cells and goblet cells. Thus, the occurrence of copious immature M cells and goblet cells, in addition to the detection of villi-like solitary lymphoid structures full of lymphocytes in the FAE of many ILF, indicate that ILF do not complete their immunological maturation in contrast to PP. Various antigenic stimulations conceivably induce the formation and maturation of ILF along the length of the small intestine. The morphological resemblance between ILF M cells and PP M cells suggests that these two types of cells perform similar or the same immunological functions.     

Loss of Function of the Mouse Sharpin Gene Results in Peyer’s Patch Regression

Peyer’s patches (PP) are an important component in the immune response against intestinal pathogens. Two independent, spontaneous mutations in the mouse Sharpin gene (Sharpin^cpdm and Sharpin^cpdm-Dem) result in the absence of PP and disrupted splenic white pulp in adult mice, although a full complement of lymph nodes is present. Here we report that rudimentary PP begin to develop in Sharpin^cpdm mice during embryogenesis, but lack the organizational patterns that are typical of this tissue. In the present study, small intestines examined at weekly intervals from birth to maturity showed spontaneous regression of PP in mutant mice with concurrent infiltration of granulocytes. At 5 to 6 weeks of age, only indistinct remnants of granulocytic accumulations remain. Transplantation of normal bone marrow into Sharpin^cpdm mice at 7 days of age did not prevent regression of PP in bone marrow chimeras examined at 7 to 8 weeks of age. These findings indicate that SHARPIN expression is required for the normal development and maintenance, but not initiation, of PP.     

Gene expression profiling of jejunal Peyer’s patches in juvenile and adult pigs

Peyer's patches are organized lymphoid tissues of the small intestine that play a critical role in disease resistance and oral tolerance. Peyer's patches in the jejunum contain lymphocytes, dendritic cells, macrophages, villous epithelium, and specialized follicle-associated epithelium. Little is known about the mechanisms and processes by which cells of the Peyer's patches discriminate food nutrients and commensal microflora from pathogenic microbiota. We hypothesize that the jejunal Peyer's patches express genes that mediate and regulate its essential functions. Expression patterns of approximately 2600 cDNAs from a porcine Peyer's patch subtracted library were examined by microarray profiling. Individual mRNAs of interest were further examined by quantitative RT-PCR. Innate immunity-associated genes, including complement 3 and lysozyme, and the genes for epithelial chloride channel and trappin 1 were highly expressed by jejunal Peyer's patch in both juvenile and adult pigs. The growth- and apoptosis-associated genes CIDE-B, GW112, and PSP/Reg I (pancreatic stone protein or regenerating gene) were differentially expressed in juvenile pig Peyer's patches. Many sequences which were highly expressed in jejunal Peyer's patches have previously been described with functions in epithelial cells. Animal-to-animal variation in basal jejunal Peyer's patch gene expression was considerable and reflects the dynamic physiological environment of the gut in addition to genetic, epigenetic, and microbiological variation in the small intestine.     

Is a mutator analogous to the Ig hypermutator of the sheep ileal Peyer's patch active on MHC class I genes in the germ line?

 Polymorphic sequence variation in the peptide-binding domains of MHC class I molecules appears to have been driven largely by the constructive action of natural selection on the specificity of the peptide-binding groove. Similar features are displayed by the variable domains of immunoglobulins generated in the sheep ileal Peyer's patch, but in this case there is evidence that the action of a targeted hypermutator acting on a selected substrate rather than antigen-driven selection is responsible for the pattern of variation in the system. Such a hypermutator acting in the germ line would not only mimic the action of natural selection but also, by convergent mutation, generate similar patterns of variation in unrelated alleles that could be interpreted as evidence for short-tract gene conversion. We analyzed human class I MHC alleles in the light of these data, but failed to find evidence of the action of a similar hypermutator. A search for other mutationally driven patterns of variation also failed, even in hypervariable residues from parsimonious phylogenies. Single-nucleotide variation at these residues is also frequent in recent allelic variants, but the data are as consistent with short-tract gene conversion as with base mutation. We conclude that the patterns of allelic variation in MHC molecules are not driven by mutational pressure, but rather by conventional mutational processes, accompanied by short-tract gene conversion and intense natural selection. Received: 6 October 1999 / Revised: 30 December 1999     

Neuroimmune connections in jejunal and ileal Peyer’s patches at various bovine ages: potential sites for prion neuroinvasion

During preclinical stages of cattle orally infected with bovine spongiform encephalopathy (BSE), the responsible agent is confined to ileal Peyer’s patches (IPP), namely in nerve fibers and in lymph follicles, before reaching the peripheral and central nervous systems. No infectivity has been reported in other bovine lymphoid organs, including jejunal Peyer’s patches (JPP). To determine the potential sites for prion neuroinvasion in IPP, we analyzed the mucosal innervation and the interface between nerve fibers and follicular dendritic cells (FDC), two dramatic influences on neuroinvasion. Bovine IPP were studied at three ages, viz., newborn calves, calves less than 12 months old, and bovines older than 24 months, and the parameters obtained were compared with those of JPP. No differences in innervation patterns between IPP and JPP were found. The major difference observed was that, in calves of less than 12 months, IPP were the major mucosal-associated lymphoid organ that possessed a large number of follicles with extended FDC networks. Using a panel of antibodies, we showed that PP in 24-month-old bovines were highly innervated at various strategic sites assumed to be involved in the invasion and replication of the BSE pathogen: the suprafollicular dome, T cell area, and germinal centers. In PP in calves of less than 12 months old, no nerve fibers positive for the neurofilament markers NF-L (70 kDa) and NF-H (200 kDa) were observed in contact with FDC. Thus, in view of the proportion of these protein subunits present in neurofilaments, the innervation of the germinal centers can be said to be an age-dependent dynamic process. This variation in innervation might influence the path of neuroinvasion and, thus, the susceptibility of bovines to the BSE agent. This work was supported by the Région Wallonne and by the Leon Frederiq Foundation.     

The distribution of α1-antichymotrypsin and amyloid production in the brain in Alzheimer’s disease

In this immunohistopathological study α₁-antichymotrypsin, which is barely demonstrable in the normal brain, was found in amyloid fibrils, endothelial cells and the cytoplasm of astroglial cells in brains from patients with Alzheimer’s disease. Amyloid precursors stained with methenamine silver were arrayed mainly along the membranes, and amyloid fibrils, which stained densely with anti-α₁-antichymotrypsin, were in direct contact with the fibrous structures connecting with the membranes of vascular feet or astrocytic processes. From the above findings, α₁-antichymotrypsin seems to play a role in the production of amyloid fibrils in Alzheimer’s disease.     

Sodium glucose-linked transport in the ruminal epithelium of fallow deer – comparison to sheep

A higher expression of the sodium glucose-linked transporter, SGLT-1, has been previously demonstrated in the intestine of adult fallow deer compared to sheep. The functional presence of SGLT-1 in the rumen of both species was investigated in the present study. Isolated ruminal epithelia were incubated in Ussing chambers. D-Glucose or 3-O-methyl-alpha-D-glucose (3-OMG) were applied at 10 mmol x l-1 to the mucosal side of tissues and the changes in short-circuit current (Delta I(sc)) were recorded. Alternatively, apical uptake of [14C]-D-glucose (200 micro mol x l-1) was determined in the presence or absence of phlorizin (100 micro mol x l-1) or Na+ (115 mmol x l-1). In both species, mucosal D-glucose addition induced a larger ( P<0.05) Delta I(sc) than the mucosal addition of 3-OMG. When comparing the effects of D-glucose or 3-OMG between species, the Delta I(sc) was two-fold larger ( P<0.01) in sheep compared to fallow deer. Accordingly, phlorizin-sensitive D-glucose uptake was approximately 50% smaller ( P<0.05) in fallow deer. It is concluded that D-glucose can be absorbed via SGLT-1 from the forestomachs of both species. However, the intermediate-mixed feeding habit of fallow deer does not seem to coincide with a higher activity of the ruminal SGLT-1.     

Oral scrapie infection modifies the homeostasis of Peyer’s patches’ dendritic cells

In transmitted prion diseases the immune system supports the replication and the propagation of the pathogenic agent (PrPSc). DCs, which are mobile cells present in large numbers within lymph organs, are suspected to carry prions through the lymphoid system and to transfer them towards the peripheral nervous system. In this study, C57Bl/6 mice were orally inoculated with PrPSc (scrapie strain 139A) and sacrificed at the preclinical stages of the disease. Immunolabelled cryosections of Peyer’s patches were analysed by confocal microscopy. Membrane prion protein expression was studied by flow cytometry. In Peyer’s patches (PP), dissected at day one and day 105 after oral exposure to scrapie, we observed an increased population of DCs localised in the follicular-associated epithelium. On day 105, PrPSc was found in the follicles inside the PP of prion-infected mice. A subset of Peyer’s patches DCs, which did not express cellular prion protein on their surface in non-infected mice conditions, was prion-positive in scrapie conditions. Within Peyer’s patches oral scrapie exposure thus induced modifications of the homeostasis of DCs at the preclinical stages of the disease. These results give new arguments in favour of the implication of DCs in prion diseases.     

Structural and Functional Features of the Wheat Embryo Sac’s Antipodal Cells during Differentiation

Russian Journal of Developmental Biology - A detailed study into the structural features of the multilevel antipodal complex of wheat Triticum aestivum L. embryo sac was performed at different...     

Control of the sheep blowfly in Australia and New Zealand – are we there yet?

The last 50 years of research into infections in Australia and New Zealand caused by larvae of the sheep blowfly, Lucilia cuprina, have significantly advanced our understanding of this blowfly and its primary host, the sheep. However, apart from some highly effective drugs it could be argued that no new control methodologies have resulted. This review addresses the major areas of sheep blowfly research over this period describing the significant outcomes and analyses, and what is still required to produce new commercial control technologies. The use of drugs against this fly species has been very successful but resistance has developed to almost all current compounds. Integrated pest management is becoming basic to control, especially in the absence of mulesing, and has clearly benefited from computer-aided technologies. Biological control has more challenges but natural and perhaps transformed biopesticides offer possibilities for the future. Experimental vaccines have been developed but require further analysis of antigens and formulations to boost protection. Genetic technologies may provide potential for long-term control through more rapid indirect selection of sheep less prone to flystrike. Finally in the future, genetic analysis of the fly may allow suppression and perhaps eradication of blowfly populations or identification of new and more viable targets for drug and vaccine intervention. Clearly all these areas of research offer potential new controls but commercial development is perhaps inhibited by the success of current chemical insecticides and certainly requires a significant additional injection of resources.     

Role of zinc and copper ions in the pathogenetic mechanisms of Alzheimer’s and Parkinson’s diseases

Disbalance of zinc (Zn²⁺) and copper (Cu²⁺) ions in the central nervous system is involved in the pathogenesis of numerous neurodegenerative disorders such as multisystem atrophy, amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, Wilson-Konovalov disease, Alzheimer’s disease, and Parkinson’s disease. Among these, Alzheimer’s disease (AD) and Parkinson’s disease (PD) are the most frequent age-related neurodegenerative pathologies with disorders in Zn²⁺ and Cu²⁺ homeostasis playing a pivotal role in the mechanisms of pathogenesis. In this review we generalized and systematized current literature data concerning this problem. The interactions of Zn²⁺ and Cu²⁺ with amyloid precursor protein (APP), β-amyloid (Abeta), tau-protein, metallothioneins, and GSK3β are considered, as well as the role of these interactions in the generation of free radicals in AD and PD. Analysis of the literature suggests that the main factors of AD and PD pathogenesis (oxidative stress, structural disorders and aggregation of proteins, mitochondrial dysfunction, energy deficiency) that initiate a cascade of events resulting finally in the dysfunction of neuronal networks are mediated by the disbalance of Zn²⁺ and Cu²⁺.     

Experimental determination of the spatial scale of a prey patch from the predator’s perspective

Foraging theory predicts that predators should prefer foraging in habitat patches with higher prey densities. However, density depends on the spatial scale at which a “patch” is defined by an observer. Ecologists strive to measure prey densities at the same scale that predators do, but many natural landscapes lack obvious, well-defined prey patches. Thus one must determine the scale at which predators define patches of prey. We estimated the scale at which guppies, Poecilia reticulata, selected patches of zooplankton prey using a behavioral assay. Guppies could choose between two prey arrays, each manipulated to have a density that depended on the spatial scale at which density was calculated. We estimated the scale of guppy foraging by comparing guppy preferences across a series of trials in which we systematically varied the scale associated with “high” prey density. This approach enables the application of foraging theory to non-discrete habitats and prey landscapes.