Mouse Plasmacytoid Cells: Long-lived Cells, Heterogeneous in Surface Phenotype and Function, that Differentiate Into CD8+ Dendritic Cells Only after Microbial Stimulus

The CD45RAhiCD11cint plasmacytoid predendritic cells (p-preDCs) of mouse lymphoid organs, when stimulated in culture with CpG or influenza virus, produce large amounts of type I interferons and transform without division into CD8+CD205− DCs. P-preDCs express CIRE, the murine equivalent of DC-specific intercellular adhesion molecule 3 grabbing nonintegrin (DC-SIGN). P-preDCs are divisible by CD4 expression into two subgroups differing in turnover rate and in response to Staphylococcus aureus. The kinetics of bromodeoxyuridine labeling and the results of transfer to normal recipient mice indicate that CD4− p-preDCs are the immediate precursors of CD4+ p-preDCs. Similar experiments indicate that p-preDCs are normally long lived and are not the precursors of the short-lived steady-state conventional DCs. However, in line with the culture studies on transfer to influenza virus-stimulated mice the p-preDCs transform into CD8+CD205− DCs, distinct from conventional CD8+CD205+ DCs. Hence as well as activating preexistant DCs, microbial infection induces a wave of production of a new DC subtype. The functional implications of this shift in the DC network remain to be determined

Transcriptional Profiling of Chondrodysplasia Growth Plate Cartilage Reveals Adaptive ER-Stress Networks That Allow Survival but Disrupt Hypertrophy

Metaphyseal chondrodysplasia, Schmid type (MCDS) is characterized by mild short stature and growth plate hypertrophic zone expansion, and caused by collagen X mutations. We recently demonstrated the central importance of ER stress in the pathology of MCDS by recapitulating the disease phenotype by expressing misfolding forms of collagen X (Schmid) or thyroglobulin (Cog) in the hypertrophic zone. Here we characterize the Schmid and Cog ER stress signaling networks by transcriptional profiling of microdissected mutant and wildtype hypertrophic zones. Both models displayed similar unfolded protein responses (UPRs), involving activation of canonical ER stress sensors and upregulation of their downstream targets, including molecular chaperones, foldases, and ER-associated degradation machinery. Also upregulated were the emerging UPR regulators Wfs1 and Syvn1, recently identified UPR components including Armet and Creld2, and genes not previously implicated in ER stress such as Steap1 and Fgf21. Despite upregulation of the Chop/Cebpb pathway, apoptosis was not increased in mutant hypertrophic zones. Ultrastructural analysis of mutant growth plates revealed ER stress and disrupted chondrocyte maturation throughout mutant hypertrophic zones. This disruption was defined by profiling the expression of wildtype growth plate zone gene signatures in the mutant hypertrophic zones. Hypertrophic zone gene upregulation and proliferative zone gene downregulation were both inhibited in Schmid hypertrophic zones, resulting in the persistence of a proliferative chondrocyte-like expression profile in ER-stressed Schmid chondrocytes. Our findings provide a transcriptional map of two chondrocyte UPR gene networks in vivo, and define the consequences of UPR activation for the adaptation, differentiation, and survival of chondrocytes experiencing ER stress during hypertrophy. Thus they provide important insights into ER stress signaling and its impact on cartilage pathophysiology

Involution of the sheep mammary gland

Changes in the ovine mammary gland epithelium during initiated involution were studied by light and electron microscopy. Apoptosis of the duct and alveolar epithelial cells was first identified at 2 d after weaning, reached a peak at 4 d and then progressed gradually thereafter. Apoptotic cells were phagocytosed by intraepithelial macrophages and alveolar epithelial cells. Occasional apoptotic epithelial cells were observed in the alveolar and duct lumina. The highly vacuolated cells in the alveolar and duct lumina were confirmed to be macrophages as they were CD45(+), MHC class II(+). Changes in myoepithelial cells involved shrinkage and extension of cytoplasmic processes into the underlying stroma and no apoptosis was observed. Regression of the blood capillaries was also by apoptosis. The resulting apoptotic bodies were either taken up by adjacent endothelial cells or were shed into the capillary lumen to be phagocytosed later by mural endothelial cells or blood monocytes. The mammary glands were completely involuted by 30 d after weaning. It was concluded that the mammary gland involutes by apoptosis, a process which allows deletion of cells without the loss of the basic architecture and the integrity of the epithelial lining of the gland

Leucocyte phenotypes in involuting and fully involuted mammary glandular tissues and secretions of sheep

Mammary glandular tissues and mammary secretions were obtained from sheep at 2–60 d after weaning to study the leucocyte phenotypes associated with mammary involution. From 2–4 d after weaning, neutrophils were the predominant leucocytes in the alveolar and ductal lumina. Lymphocytes were present in the alveolar and ductal epithelium, interalveolar and periductal areas. Most of the lymphocytes in the alveolar and ductal epithelium (IEL) were CD8(+), some were CD45R(+) and few were CD4(+). In the periductal clusters and in the interalveolar areas most of the lymphocytes were CD4(+). There was a significant increase (P < 0.05) in the percentages of CD45R(+) granulated IEL from 2 to 7 d after weaning, and this paralleled the increase in the percentages of apoptotic cells in the glandular epithelium. By 7–60 d after weaning, most cells within the alveolar and ductal lumina were macrophages followed by predominantly CD8(+) lymphocytes. CD8(+) lymphocytes were still predominant in the alveolar and ductal epithelium while CD4(+) cells were predominant in the interalveolar areas. Very few γδ(+) T cells were observed at all the stages examined. The cells in the mammary secretions correlated with those observed in the alveolar and ductal lumina. At the early stages of involution, the neutrophils and macrophages were heavily laden with lipid droplets, casein and cellular debris. The most interesting feature was the presence of cells either with extensive cytoplasmic processes (LCA(+) MHC class II(+)) or cytoplasmic veils (LCA(+) MHC class II(+)CD1(+)), probably dendritic cells. It is concluded that the cellular constituents of the mammary gland at the latter part of involution may afford the mammary gland more resistance to infection than the lactating gland and the gland at early stages of involution. The CD45R(+) IEL may trigger apoptotic cell death in the mammary glandular epithelium during mammary involution

Effect of cyclosporin A on the survival and ultrastructure of Echinococcus granulosus protoscoleces in vitro

Surgical treatment of human hydatidsosis involves the use of various scolicidal agents to kill infective Echinococcus granulosus protoscoleces that may disseminate into the peritoneal cavity during surgery and potentially re-infect the patient. Currently, no scolicidal agent is completely effective in killing intracystic protoscoleces in humans. Cyclosporin A (CsA) has previously been found to be lethal for E. granulosus protoscoleces in vitro. In this study, we further assessed the effectiveness of CsA as a scolicidal agent by testing the toxic effect of CsA at higher doses over various time-periods. Experiments were performed on activated and unactivated protoscoleces cultured in nutrient medium or sheep hydatid cyst fluid. All activated protoscoleces were killed following culture in 100 μg/ml of CsA for 3 days and 50 or 20 μg/ml for 5 days. The lethal effect of CsA on unactivated protoscoleces varied but reached 100% over 15 days in culture with 100 or 50 μg/ml of CsA. Pulse treatment of protoscoleces with 50, 20 or 10 μg/ml of CsA for 5 min or 72 h killed all parasites by day 10 and day 5 respectively. Untreated protoscoleces remained greater than 95% viable for the duration of experiments. Changes in protoscolex ultrastructure induced by treatment with 10 μg/ml of CsA over 10 days in in vitro culture was assessed by TEM. Protoscolex alterations observed in treated parasites included an increase in cellular vacuolization, swelling of mitochondria, rounding of cells, damage to the tegument, decrease in glycogen, a breakdown of the extracellular matrix and an increase in lipid globules. The untreated protoscoleces, by comparison, had few changes during the 10-day culture period with the exception of large amounts of extracellular glycogen observed in the protoscoleces at culture days 7 and 10. From these results, CsA is clearly an effective scolicidal agent in vitro that may have potential application as a new therapeutic agent in the treatment of human hydatid disease

Identification of novel osteochondrosis Associated genes

During the early stages of articular osteochondrosis, cartilage is retained in subchondral bone, but the pathophysiology of this condition of growing humans and domestic animals is poorly understood. A subtractive hybridization study was undertaken to compare gene expression between the cartilage of early experimentally induced equine osteochondrosis lesions and control cartilage. Of the many putative differentially expressed genes identified, eight were confirmed by quantitative PCR analysis as differentially expressed, in addition to those already known to be associated with early lesions. Genes encoding vacuolar H(+)-ATPase V0 subunit d2 (ATP6V0D2), cathepsin K, integrin-binding sialoprotein, integrin αV, low density lipoprotein receptor-related protein 4, lumican, osteopontin, and thymosin β4 (TMSB4) were expressed at higher levels in lesions than in control cartilage. These genes included 34 genes not previously identified in cartilage. Some genes identified as associated with early lesions are known chondrocyte hypertrophy-associated genes, and in transmission electron microscopy studies normal hypertrophic chondrocytes were observed in lesions. Differential expression of ATP6V0D2 and TMSB4 in the cartilage of early naturally occurring osteochondrosis lesions was confirmed by immunohistochemistry. These results identify novel osteochondrosis-associated genes and provide evidence that articular osteochondrosis does not necessarily result from failure of chondrocytes to undergo hypertrophy

Eimeria taggarti n. sp., a novel coccidian (Apicomplexa: Eimeriorina) in the prostate of an Antechinus flavipes

A novel coccidian species was discovered in the prostate of an Antechinus flavipes (yellow-footed antechinus) in South Australia, during the period of post-mating male antechinus immunosuppression and mortality. This novel coccidian is unusual because it develops extra-intestinally and sporulates endogenously within the prostate gland of its mammalian host. Histological examination of prostatic tissue revealed dense aggregations of spherical and thin-walled tetrasporocystic, dizoic sporulated coccidian oocysts within tubular lumina, with unsporulated oocysts and gamogonic stages within the cytoplasm of glandular epithelial cells. This coccidian was observed occurring concurrently with dasyurid herpesvirus 1 infection of the antechinus' prostate. Eimeria-specific 18S small subunit ribosomal DNA PCR amplification was used to obtain a partial 18S rDNA nucleotide sequence from the antechinus coccidian. Bayesian phylogenetic analysis based on 18S rDNA gene sequences revealed that the novel coccidian clusters with reptile-host coccidians, forming an ancestral basal lineage of the eimeriid clade. The species has been named Eimeria taggarti n. sp., based on both sporulated oocyst morphology and molecular characterization. It is suspected that E. taggarti is sexually transmitted via excretion of sporulated oocysts and/or free sporocysts with prostatic secretions in semen.J. Amery-Gale, J.M. Devlin, L. Tatarczuch, D.A. Taggart, D.J. Schultz, J.A. Charles and I. Beveridg