The proportion of Tregs was evaluated. To elucidate possible differences in functional properties of Tregs, MFI of FoxP3 and intracellular regulatory cytokines IL-10 and TGF-beta were tested. Differences in Treg proportions and their functional properties were found between the groups. Using our gating strategy (Fig. 1) and antibodies against CD4, CD25, CD127 and FoxP3, we did not find significant differences in the proportion of Tregs in the cord blood of children of healthy and allergic mothers, although the trend towards an increased number of Tregs in the CD4+ lymphocyte population from the allergic group was obvious (P = 0·07) (Fig. 2a). A significantly

increased proportion of Tregs in cord blood of children of allergic mothers was observed when I-BET-762 purchase Tregs were considered only as CD4+CD25+ cells find more (P = 0·0117) (Fig. 2b). Different gating strategies together with using different Treg markers may account for variation among the results of different research groups. Transcription factor FoxP3 is considered to be a master marker for identifying Tregs[24] (as CD25 can be expressed on other activated CD4+ T lymphocytes and CD127 is present on various cell types). The values of MFI of

FoxP3 in cord blood of children of allergic mothers followed an opposite trend to the proportion of Tregs. A significantly higher MFI of FoxP3 (P = 0·0159) in cord blood Tregs of children of healthy mothers was detected in comparison to children tuclazepam of allergic mothers (Fig. 3). To evaluate the possible differences in functional characteristics of Tregs, the presence of regulatory cytokines IL-10 and TGF-beta was estimated by intracellular staining. A significantly

higher number of IL-10+ Tregs in cord blood of children of healthy mothers was detected in comparison to children of allergic mothers (P = 0·0012) (Fig. 4). Similarly, a significantly higher proportion of TGF-beta+ Tregs in cord blood of children of healthy mothers is documented in Fig. 5 (P = 0·0174). The importance of Tregs in immune regulations consists mainly in their role in induction of peripheral tolerance against autoantigens and harmless food and environmental antigens [25]. An insufficiency of Tregs can result in autoimmunity and allergy development [26–29]. We followed the status of newborn Tregs as a possible prognostic marker for future allergy manifestation. It is possible to assume that changes of immune regulation in allergy-prone infants can be evident prior to development of the clinical signs of allergy. We found differences in immune characteristics of Tregs in the cord blood of children of allergic mothers in comparison to children of healthy mothers. Tregs were assessed on the basis of their cell surface markers (CD4, CD25high and CD127low), typical transcription factor FoxP3 and intracellular regulatory cytokines IL-10 and TGF-beta.

Moreover, the same factors were compared between the pneumonia patients with and without leukocytosis. Mean peak cytokine and chemokine concentrations in the patients were compared between the two groups using the Mann-Whitney U test. Statistical analysis was performed with StatView software, version J-5.0. No patients required mechanical ventilation and all pneumonia patients recovered completely. Antiviral drugs were administered to 46 patients (oseltamivir; 35 patients, zanamivir; 11 patients), and steroid treatment in addition to antiviral drugs in 21 patients. Steroids were administered soon

after admission AUY-922 datasheet to hospital (after serum sample collection). As shown in Table 1, no statistical differences were observed in age, male to female ratio, sampling selleck time of the serum, and C-reactive protein concentration between the patients with and without pneumonia. SpO2 was significantly lower in patients with pneumonia than in those without pneumonia (P = 0.036), whereas white blood cell counts were significantly

higher in patients with pneumonia than in those without pneumonia (P = 0.003). Cytokine and chemokine concentrations in patients with and without pneumonia are summarized in Table 1. Expression of IL-10 (23.5 pg/mL vs 9.1 pg/mL, P = 0.027) and IL-5 (18.0 pg/mL vs 12.6 pg/mL, P = 0.014) were significantly higher in patients with pneumonia than in those without pneumonia. No statistical differences between the two groups were observed in the concentrations of the other six cytokines and five chemokines. As shown in Table 2, except for white blood cell counts, no statistical

differences were observed in the other variables assessed, including the detection rate of bacteria in throat swabs from pneumonia patients with and without leukocytosis. As shown in Table 2, neutrophilia contributed exclusively to leukocytosis. Cytokine and chemokine concentrations in these patients are summarized in Table many 2. Serum concentrations of IFN-γ (35.7 pg/mL vs 62.8 pg/mL, P = 0.009), TNF-α (9.6 pg/mL vs 18.2 pg/mL, P = 0.01), IL-4 (22.5 pg/mL vs 30.5 pg/mL, P = 0.024), and IL-2 (9.0 pg/mL vs 18.1 pg/mL, P = 0.012) were significantly lower in the pneumonia patients with leukocytosis than in those without leukocytosis. Of the five serum chemokine concentrations assessed, only IL-8 was significantly lower in pneumonia patients with leukocytosis than in those without leukocytosis (16.2 pg/mL vs 181.1 pg/mL, P = 0.001). As reported and discussed in previous studies (3, 4, 8), high concentrations of IL-10, an immunomodulatory cytokine, have been associated with severe cases of pandemic A/H1N1/2009 influenza virus infection and appear to reflect regulation of excessive immune responses due to lung injury in patients with pneumonia. In addition to IL-10, the IL-5 concentration was also significantly higher in patients with pneumonia than in those without pneumonia.

[3] Finally, activation of iNKT cells with αGalCer caused rapid weight loss, and reversal of glucose and insulin sensitivity without hypoglycaemia.[3, 39] Hence, the scenario appears that iNKT cells normally reside in adipose tissue, produce mainly Th2 and regulatory cytokines and positively regulate anti-inflammatory macrophages

and adipocyte function. In an obese setting, adipose iNKT cells are depleted, representing the loss of an important regulatory population and at the same time, adipose tissue becomes an inflammatory environment due to an accumulation of pro-inflammatory macrophages (Fig. 2). Although the exact Regorafenib clinical trial pathway of iNKT cell regulation is not yet clear, it appears that adipose iNKT cells can directly regulate macrophage levels and phenotype, and therefore inflammation. However, the role of iNKT cells in the protection against obesity, weight gain and metabolic disorder has been somewhat controversial. The similarities and differences selleck kinase inhibitor between these studies are summarized below. To study the effects of

iNKT cells on obesity and metabolism control, there are a number of methods that have been applied. Most research groups have used models of iNKT cell deficiency, namely CD1d−/− and Jα18−/− mice. Mice lacking CD1d, which is essential for iNKT cell development, do not develop iNKT cells. However, these mice not only lack type I NKT cells but also type II NKT cells, Etoposide as well as CD1d itself, which is expressed on adipocytes and other non-hepatopoietic cells and so may be an important molecule in metabolism. Jα18−/− mice have

a specific deficiency in the invariant chain of the NKT TCR, and specifically lack iNKT cells, but it has recently come to light that Jα18−/− mice have lower TCR diversity than was first thought,[59] which could potentially contribute to any phenotype observed. Loss or gain of function after birth in wild-type mice may be a more appropriate method to study iNKT cell function in obesity. Mice can develop with a normal T-cell repertoire, and then iNKT cells can be depleted or adoptively transferred into mice to measure the effect on weight and metabolism. However, there is currently no way to specifically deplete iNKT cells in vivo. The common method is to use anti-NK1.1 antibody; however, this also depletes NK cells, which often outnumber iNKT cells. This method also would not deplete iNKT cells lacking the NK1.1 receptor, which is a substantial proportion of adipose iNKT cells. We, and others, have performed gain of function studies, by adoptively transferring iNKT cells into obese wild-type and iNKT-deficient mice, as well as specifically activating them by injection of αGalCer. In the recent studies that aimed to determine the role, if any, for iNKT cells in obesity, the main discrepancies between laboratories were seen in the mouse models of iNKT cell deficiency. On one side of the argument, Ohmura et al.

Similar findings later revealed that macrophages ingest and kill the parasites [98, 99]. Protective

immunity depended on macrophage activation as well as antibody. Protection against P. chabaudi adami was found to be independent of antibody but critically dependent on spleen T cells [100], and TNF released from endotoxin-induced macrophage activation was shown to be damaging to both lethal and Selleck Doxorubicin nonlethal P. yoelii parasites [101, 102]. T cells from mice successfully vaccinated against lethal P. yoelii reacted strongly to parasite antigens, with infiltration of mononuclear cells in antigen-challenged pinnae [28]; when the mice were challenged with live parasites, increased homing of mononuclear cells and parasites to the spleen and liver occurred [78]. This T-cell-dependent shift in cell traffic was interpreted to be a cell-mediated immune response against the parasites as they migrated to the

spleen and liver [79]. Macrophage activation was strongest in infections with the nonlethal P. yoelii and P. chabaudi, and significantly weaker in the lethal P. yoelii and P. berghei infections, and vaccination increased activation more with the former than the latter. It appeared that the attraction and stimulation of cytotoxic myeloid cells by T cells play an important part in the protection of vaccinated mice. This DTH T-cell

response was confirmed by the observation that the trapping of parasites, and of both buy Galunisertib lymphoid and myeloid cells, in the spleens and livers of vaccinated mice increased shortly after infection [79]. Studies in vitro revealed that spleen- and liver-derived myeloid cells killed lethal P. yoelii parasites in vitro through TNF and hydrogen peroxide-dependent killing mechanisms [103], and macrophages from mice infected with nonlethal parasites and those from vaccinated mice were more cytotoxic against L929 cells than macrophages from lethal infections [104]. Stevenson and colleagues showed that depletion of macrophages in mice treated with silica significantly impaired over their ability to control P. chabaudi parasites [105]. The susceptibility of A/J mice to P. chabaudi infection also correlated with impaired macrophage activation and effector functions, including impaired expression of MHC-II, decreased production of IL-12 p70 and decreased strength of the respiratory burst [106]. Monocytes and macrophages played a significant role in the control of the early parasitaemia of both lethal and nonlethal P. yoelii infections [107]. Mice given clodronate-encapsulated liposomes to deplete them of macrophages failed to control nonlethal P. yoelii infections, suggesting that a strong early innate response was needed to control infection (107).

Occupational allergies, drug allergies and allergies to stings (occasionally fatal) add further complexity and concerns. Finally, new types of allergic diseases and allergies against previously non-allergenic substances are increasingly RG7204 being reported; however, the fact that more patients are affected

and that allergic conditions are nowadays more severe and complicated are not the only issues which make these diseases a matter of concern – the actual burden for patients and for society as a whole is very high. The quality of life is severely affected in allergic patients. Although some allergic conditions are considered non-severe, others such as asthma or anaphylaxis can be life threatening. Allergic patients have increased disadvantages affecting their personal development, career progression and lifestyle choices. Allergic children demonstrate difficulty in coping at school and they develop associated learning difficulties and sleeping problems. As a result, it has been observed that sleepiness and mood swings frequently lead children to be isolated and even bullied by their peers. Allergic rhinitis in students increases by 40%, the chance of dropping a grade in summer examinations,

www.selleckchem.com/products/Adriamycin.html while taking a sedating drug may further increase this to 70% 5. Young adult patients also face a significantly higher amount of problems in their work place due to increased numbers of sick days and a reduction in productivity. Many allergic patients report problems in their personal

relationships. Finally, several studies have shown that allergic individuals have a higher risk of developing depression 6. The impact of allergies on the quality of life can be as high, or higher, than diseases that are considered more ‘serious’ (i.e. diabetes). The impact of allergy on health economics and macroeconomics is equally high. The associated reduction in productivity and the rising number of sick days taken by patients represent some of the biggest negative outputs recorded impacting national, business and health economies in Europe. Allergy incidents and their increase have an adverse effect on the European economy due to both direct costs (e.g. for asthma alone, the pharmaceutical cost stands at € 3.6 billion per year and the cost Amoxicillin of health care services at € 4.3 billion per year) 7 and, perhaps to an even greater degree, indirect costs. In total, 15% of the population receiving long-term treatment in Europe is due to allergies and asthma, making them the most common reasons for treatment among the young age group 8. Among the direct medical costs, diagnostic tests, consultations and medication represent the primary components, while a major cost item is hospitalisation, usually associated with severe exacerbations of asthma or severe anaphylactic reactions. Moreover, performance deficits, loss of productivity and absenteeism are closely linked to allergy suffering and have a major effect on macro-economics.

Mice that received pretreatment with Con-A or PBS were infected with C. albicans and sacrificed after 30 min, 2, 6, 18, and 24 h, and their peritoneal exudates were collected with 2 mL RPMI 1640 medium (Sigma-Aldrich) pH 7.0 with 14 μg gentamicin. For cytokine analysis, the supernatants were

collected by centrifugation (800 g × 4 min at 4 °C). To evaluate the population of collected cells from the peritoneal exudate cells, the pellet XAV 939 was resuspended in 1 mL RPMI medium pH 7.0 supplemented with 5% inactivated fetal calf serum (FCS) plus 7 μg gentamicin. The peritoneal exudate cells collected following infection were adhered to coverslips (0.2 mL per coverslip) for 30 min at 37 °C. The following tests were applied to the cells: (1) staining by May-Grunwald-Giemsa (Merck, Darmstadt, Germany) and analysis by light microscopy to evaluate the populations of macrophages and neutrophils, percentage selleck screening library of phagocytosing cells by counting 20 fields; (2) staining with propidium iodide plus 6-carboxyfluorescein

diacetate (6-CFDA; Sigma-Aldrich) to evaluate the presence of necrotic and viable cells, as described by Gasparoto et al. (2004). Analysis of fluorescence labeling was performed in a fluorescent microscope (Zeiss) and photographed (blue-violet irradiation: BG-38 and BG-12 excitation filters and 530-barrier filter), and (3) the cells were fixed in 2.5% glutaraldehyde (Merck) buffered with phosphate 0.1 M for 2 h at room temperature

followed by postfixation in osmium tetroxide 1% for 1 h. Following dehydration in ethanol, the samples were dried by the critical-point method, coated with a thin layer of gold and examined in a scanning microscopy (Shimadzu 550 SS) after 2 h of phagocytic assays in vitro. The mice received intraperitoneally Con-A 250 μg per 250 μL PBS or PBS alone 72 h before phagocytic assays. To study Dectin-1, monolayers of peritoneal macrophages 4 × 105 were preincubated with laminarin (Sigma-Aldrich) 100 μg mL−1 PBS for 30 min at 37 °C (Gantner et al., 2005). To study mannose receptors, macrophages were preincubated with mannan (Sigma-Aldrich; 100 μg mL−1 PBS; Gaziri et al., 1999). Fresh laminarin or mannan were TCL added to C. albicans 2 × 106 and coincubated with macrophages at 37 °C for 30 min. A total of 200 phagocytes were analyzed for each preparation, and the percentage of phagocytes that phagocytosed Candida was determined following staining of the cells with May-Grumwald-Giemsa (Merck). Supernatants were collected after centrifugation of peritoneal exudates obtained from each mouse from all the experimental groups and submitted to capture ELISA (eBioscience, San Diego, CA) to determine the concentrations of IL-17, TGF-β, IL-1β, IL-6, IL-12, IFN-γ and TNF-α. Cytokines assays were performed according to the manufacturer’s instructions. The differences between groups were analyzed by the Student’s t-test. P < 0.

Although free-living species display a high propensity for symbioses spanning the spectrum from commensalism to parasitism, there is strong evidence that the major parasitic lineages form a monophyletic group, demonstrating that obligate parasitism arose only once during the course of flatworm evolution (11). This was associated with a major developmental shift involving the separation of ontogenetically distinct

larval and adult stages, with replacement of the larval epidermis by a syncytial tegument. Within this clade, we now recognize four independent lineages: the cestodes (tapeworms), digeneans (flukes) and monopisthocotylean and polyopisthocotylean ‘monogeneans’. Interrelationships of these lineages remain controversial, but have begun to point toward a sister relationship between cestodes and digeneans, Selleck BMS-777607 and paraphyly of the ‘Monogenea’ (11,14,15), in contrast to previous hypotheses (and classifications) that considered ‘monogeneans’ to be both monophyletic and the sister group to tapeworms. The main implications of the molecular-based hypotheses are a common origin of both enteric parasitism and complex life cycles in tapeworms Selleckchem AZD1208 and flukes despite major differences in their life histories, and that the first neodermatan flatworms were nonenteric and direct-developing, as seen in contemporary monopisthocotylean

and polyopisthocotylean parasites. Only in the last two decades Liothyronine Sodium has our understanding of tapeworm interrelationships begun to stabilize, thanks to a more concerted effort on the part of cestodologists (16) and the wide application of molecular phylogenetic techniques (14). Circumscription of even the primary tapeworm lineages has required major revisions to reflect new insights into their affinities, resulting in the proposal of three new tapeworm orders since 2008 (17,18). Interrelationships of the 15 or more natural (i.e. monophyletic) groups of tapeworms

have yet to be resolved satisfactorily, but it is clear that early branching lineages colonized a wide spectrum of cartilaginous and bony fishes before subsequent diversification led to the colonization of homeothermic hosts (e.g. birds, mammals) (19–21). Among the early branching groups, only the Diphyllobothriidea [n.b. formally classified as a family of Pseudophyllidea (18)] radiated into homeotherms, but retained its association with fishes (which became 2nd intermediate hosts) and transmission via aquatic life cycles (22). There was thus a single primary colonization of homeothermic hosts coincident with the adoption of fully terrestrial life cycles that gave rise to the most speciose contemporary group, the Cyclophyllidea. The extent to which tapeworm–host associations were shaped by the unique adaptive immunity of the mammalian host is not clear from an evolutionary perspective.

To determine if the extensive infiltration of eosinophils in thyroids of IFN-γ−/− mice contributes to thyroid damage and/or early resolution of G-EAT, anti-IL-5 was used to inhibit migration of eosinophils to thyroids. G-EAT severity was compared at day 20 and day 40–50 in IFN-γ−/− recipients

given anti-IL-5 or control immunoglobulin G (IgG). Thyroids of anti-IL-5-treated IFN-γ−/− mice had few eosinophils and more neutrophils at day 20, but G-EAT severity scores were comparable to those of control IgG-treated mice at both day 20 and day 40–50. Expression of chemokine (C-X-C motif) ligand 1 (CXCL1) mRNA was higher and that of chemokine (C-C motif) ligand 11 (CCL11) mRNA was lower in thyroids of anti-IL-5-treated IFN-γ−/− mice. IL-5 neutralization did not influence mRNA expression of most cytokines in IFN-γ−/− mice. Thus, inhibiting eosinophil migration Protease Inhibitor Library nmr to thyroids did not affect G-EAT severity or resolution in IFN-γ−/− mice, suggesting that eosinophil infiltration of thyroids occurs as a consequence of IFN-γ deficiency, but these cells have no apparent pathogenic role in G-EAT. Granulomatous experimental autoimmune thyroiditis (G-EAT) is induced by adoptive transfer of mouse thyroglobulin (MTg)-primed splenocytes activated in vitro with MTg

and interleukin (IL)-12.1–4 The adoptive transfer model of G-EAT is an excellent experimental model with which to study the contributions of various cells and inflammatory mediators in induction and resolution of autoimmune inflammation.1–6 CHIR-99021 research buy Our previous studies showed that G-EAT lesions in recipients of activated splenocytes reach

maximal severity 20 days after cell transfer and evolve over time to two distinct outcomes: either inflammation resolves, or there is continuing inflammation with development of fibrosis.6–8 When interferon (IFN)-γ−/− or wild-type (WT) DBA/1 mice are used as donors and recipients, both develop G-EAT with similar severity scores 20 days after cell transfer.6–8 However, thyroid lesions in IFN-γ−/− mice have many eosinophils and almost no neutrophils, while those in WT mice have many neutrophils and very few eosinophils, with fibrosis and necrosis.6–8 Thyroid lesions in IFN-γ−/− mice consistently resolve by day Erlotinib clinical trial 40–50, whereas those in WT mice have ongoing inflammation and fibrosis that persists for more than 60 days.6–8 These results suggest that differential infiltration of neutrophils versus eosinophils could contribute to the different outcomes of G-EAT in WT versus IFN-γ−/− mice. Eosinophils are multifunctional leucocytes that play important roles in asthma and several other inflammatory processes.9 Eosinophils are frequently associated with tissue remodelling and fibrosis in allergy as well as other diseases, including Riedel’s thyroiditis and pulmonary fibrosis.10–14 IL-5 regulates the activation, differentiation, recruitment and survival of eosinophils.

While the mechanisms that control T. retortaeformis and G. strigosum abundance remain obscure, our findings support the hypothesis of a Th2-mediated antibody and eosinophil clearance of primary infections to the former species but not the latter nematode (47–50). Our recent modelling of the immune response network to T. retortaeformis, based on this study, was consistent

with a Th2-mediated antibody/eosinophil clearance and an IL-4 anti-inflammatory Selleck INCB024360 protection against this nematode (Takar et al., in preparation). However, additional experiments are necessary to confirm these conclusions. In this respect, the evidence that IL-4 can induce Foxp3-expressing Treg and the potential for parasite tolerance (51) raises the question of whether the persistence of G. strigosum in the presence of high IL-4 mucosa expression

involves some tolerance mechanisms activated by the rabbit or whether this is an intrinsic property of the stomach to avoid immuno-pathology. Closely related helminth CH5424802 infections of other herbivores such as sheep and cattle have highlighted the less effective immune response to the abomasal parasites Teladorsagia circumcincta, Haemonchus contortus, Ostertagia ostertagi and Haemonchus placei, compared to the more efficient response against the intestinal nematodes Trichostrongylus colubriformis, Trichostrongylus vitrinus and Cooperia spp. Our study is consistent with these general findings, specifically stomach and small intestine Thalidomide are distinct environments with different immune properties (52) and colonized by helminths with contrasting life history traits (53,54). Based on these systems, helminths in the stomach/abomasal, such as G. strigosum,

tend to have larger body size, slower development and higher fecundity. They also appear to stimulate an immune response either that is slow to develop or has higher tolerance to infections, or can be more easily immuno-suppressed by the helminth. Helminths in the small intestine, e.g. T. retortaeformis, have the opposite of these life history features, probably as a response to a more effective immune response. The co-evolution of the host immune system and the helminth life history traits in the stomach and small intestine appear to have followed different strategies. Nevertheless, in our rabbit–nematode system, the outcome has been equally successful as these parasites cause persistent chronic infections. In conclusion, we have shown that T. retortaeformis and G. strigosum exhibited different immuno-parasitological characteristics during primary infections of naïve rabbits. These nematodes appear to elicit an unequivocal Th-2-biased immune response.

2; in Braak stages V-VI, small numbers of UBL immunoreactive pyramidal cells remaining in the CA1 precluded optical density analyses). The ratio was slightly, but non-significantly, elevated in the CA2/3 field from Braak stage groups III-IV and V-VI when compared to Braak stage group 0-I-II, and a similar trend was observed in the CA4 field (Fig. 2). Optical density measurements in the nucleoplasm and cytoplasm

correlated directly across all Braak staged groups in CA2/3 as well as in CA4, but did not correlate in the CA1 field (data not shown). We detected statistically significant (Spearman r = 0.7, Gefitinib clinical trial P = 0.01) correlation between more advanced age and higher nucleoplasm/cytoplasm UBL immunoreactivity optical density ratio values in CA1, but not CA2/3 or CA4. The relationship between UBL protein and a marker of advanced stage NFT including extracellular “ghost NFT” (X-34) or an antibody that also recognizes pre/early NFT (AT8) was examined

using multiple-label fluorescence confocal microscopy (Figs 3, 4). The pattern of UBL immunofluorescence was consistent with our observations using the same antibody and chromogen-based immunohistochemistry with light microscopy (Fig. 3). In multiple-labeled (UBL, AT8, DAPI, X-34) sections from Braak stage 0-I-II cases, we observed pyramidal neurons with UBL immunofluorescence in the cytoplasm and nucleoplasm, the latter co-labeled with DAPI (Fig. 3A–D). Braak 0-I-II cases had no AT8- or X-34-positive NFT in the hippocampus, although sparse, scattered AT8 immunofluorescent neuritic elements were observed in the CA fields (Fig. 3E–H). In Braak stages III-IV and V-VI cases, we observed a complex pattern of UBL/AT8 or UBL/X-34 co-localization

Selleckchem SB203580 in CA fields. Neurons with light cytoplasmic and prominent nucleoplasmic UBL immunofluorescence co-localized AT8, but had little or no X-34, (Fig. 3I–L,M–P,M′–P′). The majority of UBL-immunofluorescent Phosphatidylinositol diacylglycerol-lyase pyramidal neurons in the CA2/3 region were AT8- and X34-negative, yet surrounded by numerous AT8-immunofluorescent neurites (Fig. 3I–L). Pyramidal neurons in CA1 and subiculum of Braak stages V-VI cases had UBL immunofluorescence co-localized with X-34, and very little or no AT8 immunofluorescence and no DAPI labeling, indicative of extracellular “ghost” NFT (eNFT, Fig. 3M–P,M″–P″). UBL immunoreactive neuritic elements were also detected within X-34 labeled amyloid plaques in the CA1 and DG molecular layer (not shown). A small number of AT8-positive neurons lacking UBL immunofluorescence were observed in the CA1 region of Braak V-VI cases. The overall pattern of UBL/AT8/X-34 immunofluorescence in a representative Braak stage VI case is illustrated diagrammatically in Figure 4. The present study investigated UBL immunoreactivity in the hippocampus from non-AD and clinically diagnosed AD cases stratified by Braak stages, in relation to markers of primarily advanced stage NFT (the pan-amyloid marker X-34) and the antibody clone AT8 which also recognizes pre/early NFT.