The identification of the underlying mechanisms, which regulate t

The identification of the underlying mechanisms, which regulate the expression levels of the various isoforms, and the elucidation of the physiological relevance for the differential modulation of IRF3 and NF-κB activation will lead to an enhanced understanding of the diverse functions of IKKε in the context of an innate immune response. The Ab against TBK1, phospho-IRF3, phospho-p65 (Ser-536 and Ser-468 specific), and the two different Ab against IKKε (rabbit mAb D20G4 and rabbit polyclonal antiserum recognizing the C-terminus of IKKε) were purchased

from Cell Signaling Technology (Frankfurt am Main, Germany), the anti-FLAG mAb M2 was obtained from Sigma (Taufkirchen, Germany), the anti-myc mAb from Invitrogen (Karlsruhe, Germany), the IRF3 Ab from Epitomics (Burlingame, selleck compound CA, USA), and the actin Ab was purchased from Santa Cruz (Heidelberg, Germany). Poly(I:C) Afatinib cost and blasticidine were obtained from InvivoGen (San Diego, CA, USA). The purification of RNA was performed using the NucleoSpin RNA II kit from Macherey-Nagel (Düren, Germany); cDNA was generated using the First-Strand cDNA Synthesis Kit from GE-Healthcare (München, Germany). Amplification by PCR and ligation into the expression vectors pRK5, pFLAG.CMV2, and pcDNA3.1 myc-His were performed using standard protocols.

Fusion constructs of NAP1, TANK, and SINTBAD with Renilla luciferase were kindly provided by F. Randow (Cambridge, tuclazepam UK) 9. In vitro mutagenesis was performed using the QuickChange kit purchased from Stratagene (La Jolla, CA, USA), following the instructions of the manufacturer. Primers used for PCR and mutagenesis are summarized in Supporting Information Table S1. All constructs were verified by DNA sequencing. To quantify the expression of the different IKKε isoforms, PCR products were cloned into the pCR2.1-TOPO vector using the TOPO-TA cloning kit from Invitrogen. Plasmid DNA was isolated from the resulting colonies and inserts were analyzed by sequencing. HEK293T, MCF7, U937, and THP1 cells were originally obtained from ATCC, 293/TLR3

cells were obtained from InvivoGen. HEK293T, 293/TLR3, and MCF7 cells were grown in DMEM medium, U937 and THP1 cells in RPMI 1640 medium. Both media were supplemented with 10% fetal calf serum and 50 μg/mL each of streptomycin and penicillin. Briefly, 293/TLR3 cells were additionally cultivated with 10 μg/mL blasticidine. HEK293T and 293/TLR3 cells were transiently transfected by standard calcium phosphate precipitation or using FuGene HD (Roche Molecular Biochemicals, Penzberg, Germany) as suggested by the manufacturer. Human PBMC were purified from buffy coats of healthy donors using Ficoll-Hypaque and grown in RPMI 1640 medium supplemented with 10% fetal calf serum and 50 μg/mL of streptomycin/penicillin. The use of buffy coat cells for these experiments was approved by the local Ethics Commission.

Similarly, icv injections of anti-p75-saporin and sham-lesions in

Similarly, icv injections of anti-p75-saporin and sham-lesions into 3- and 12-month-old WT mice (again with a subsequent observation period of 4 months) resulted in staining patterns indistinguishable from those in the related animal groups displayed in Figure 2

(data not shown). After ChAT immunolabelling in CPN of immunotoxin-treated animals, the success of immunolesion had been checked microscopically, while only hippocampi from mice with verified immunolesion of the MS/DB (n = 21) were applied to subsequent biochemical analysis. Western blotting of TBS-solubilized hippocampal tissues from 7-month-old mice did not show significant AZD1208 differences in APP protein levels between control (n = 4) and immunolesioned

3xTg mice (n = 3; Figure 3a), whereas levels of its APP metabolites C99 and Aβ could not be detected by direct Western blotting at this time. Interestingly, protein levels of the astrocytic marker GFAP were significantly increased in immunolesioned 3xTg mice at 7 months of age (P < 0.05), when no obvious extracellular Aβ deposition is present in the hippocampal formation. Immunolesioned 3xTg mice at 16 months (n = 4) showed significantly increased levels of APP (P < 0.05) and C99 (P < 0.01) as well as monomeric Aβ (P < 0.05) for which only a faint band could be detected in the hippocampal lysates from untreated control 3xTg mice (n = 3). No differences in GFAP HM781-36B mouse levels could be detected at this time (not shown). Next, we quantified

the levels of total tau protein in SDS-soluble fractions using anti-human tau as well as antibodies directed against a variety of phospho-tau epitopes, including MC-1, pS199, CP13, AT8 and pS422. No differences in any of these tau variants could be detected in 7-month-old immunolesioned 3xTg mice compared to their age-matched Loperamide controls (Figure 4a). In contrast, 16-month-old immunolesioned 3xTg mice showed a significant increase in total tau levels (P < 0.05), as well as significantly increased protein levels using the phospho-tau specific antibodies MC-1, pS199 and CP13 (all P < 0.05). Using the antibodies AT8 and pS422, increased protein levels were detected in these mice, but failed to reach statistical significance (Figure 4b). In 16-month-old animals, immunofluorescence labelling of phospho-tau with AT8 and detection of total Aβ with a rabbit antiserum revealed strong hippocampal tau hyperphosphorylation and considerable β-amyloidosis even in tissue from naive mouse (Figure 5a), which was apparently enhanced in immunolesioned animals as exemplarily shown in Figure 5b. Additional co-staining elucidated phospho-tau-immunoreactivity for CP13 in close vicinity to hippocampal Aβ deposits in an immunolesioned animal (Figure 5c) and a naive mouse (not shown).

In such a simplified system with only two elements, i e short sy

In such a simplified system with only two elements, i.e. short synthetic peptide and CpG, there are limited means by which the B cells

could be impacting the CD8+ T-cell responses. Previous studies have demonstrated that B-cell presentation of antigen directly to CD8+ T cells could lead to aberrant T-cell responses or deletion of antigen-specific T cells altogether 24, 25, 28. It has been shown that direct antigen presentation to CD4+ helper T cells by antigen-specific B cells is important to optimal antibody responses 31. However, their role in priming CD8+ T cells is unclear. Thus, while B cells are considered professional Metformin APC because of their expression of MHC class II and other T-cell costimulatory machinery, they may be unable to properly prime cytotoxic CD8+ T cells. Selleckchem MDV3100 In our experiments, reconstitution of B cell-deficient mice with only 3×106 B cells largely restored the phenotype of WT mice. The ability of this relatively low number of cells suggests that direct antigen presentation of peptide to T cells by B cells may not be the mechanism of B-cell regulation, though this possibility cannot be ruled out entirely. Despite significantly enhanced survival of CD8+ T cells in the absence of B cells, the T cells were unable to provide protection against live P. yoelii parasite challenge (data not shown). Studies are currently underway to determine if there are defects in T-cell effector

function in the absence of B cells or if there are limitations of this immunization protocol in generating large enough numbers of T cells required

for protection in this assay. B cells could regulate CD8+ T-cell responses to peptide by responding to CpG in a manner that is detrimental D-malate dehydrogenase to effector T-cell survival 32. Indeed, B cells have been shown to proliferate 33–36 and upregulate costimulatory molecules 35, 36 in response to LPS or CpG, but they also potently produce IL-10 and TGF-β 26, 37–40. Thus, while CpG pre-treatment could induce factors that promote T-cell survival such as production of IFN-α 41, 42 and increased numbers of DC in the LN 33, it may also induce suppressive factors from B cells that drive T-cell death. There is likely a delicate balance of these factors that allows for the survival of a small number of T cells in normal mice that receive CpG and peptide. Differential kinetics of the production of enhancing and detrimental soluble factors could help to explain the positive effects of delaying antigen delivery after CpG pre-treatment. It has been proposed that B-cell responses to innate stimuli, such as CpG, contribute to immune suppression through promotion of regulatory T-cell activity 43. However, depletion of CD4+ cells did not alleviate the suppression of the CD8+ T-cell response to CpG and peptide in intact mice, suggesting that regulatory T cells were not playing a direct role (data not shown).

[16] Serum ferritin, folate or vitamin B12 levels were in normal

[16] Serum ferritin, folate or vitamin B12 levels were in normal range in all of the patients and none of the patients had a blood transfusion in the past 6 months. Therefore the RDW increase in this study seems to be related to prostate enlargement. Although not previously correlated with prostate enlargement, elevation of the RDW has been associated with other non-hematologic disease processes including PLX-4720 molecular weight liver disease, malnutrition, heart failure, cardiovascular events, and “occult” colon cancer.[4, 17, 18] None of our patients reported any of the aforementioned disorders or other disorders having chronic inflammatory

or infective processes. Although the exact pathophysiological mechanisms that underlie the association of the RDW with the aforementioned disorders are unknown, systemic factors that alter erythrocyte homeostasis, such as inflammation, likely play a role.[4-6] In BPH there is enough evidence indicating that chronic inflammation has a crucial role in the development of the disease.[10-14, 19, 20] Emans et al.[21] and Lippi et al.[8] reported a graded association of the RDW with high-sensitivity CRP and ESR independent of numerous confounding factors. In this study, the WBC and CRP were positively related to Roscovitine mw the RDW when used as indicators of inflammation, suggesting that

inflammation has a role in increasing the RDW. It has been suggested that inflammation might contribute to an increased RDW via ineffective 3-mercaptopyruvate sulfurtransferase erythrocyte production by impairing iron metabolism, by inhibiting erythropoietin and the response to erythropoietin, or by shortening erythrocyte survival rates.[22, 23] One of the inflammatory mediators, interleukin-6 (IL-6), was found to be strongly associated with an elevated RDW in various studies.[7, 24] IL-6 is a strong inducer of hepcidin gene transcription.[25] In the intestine hepcidin decreases iron absorption and inhibits iron release from reticuloendothelial stores.[26] This so-called “reticuloendothelial block” may lead

to the RDW elevation. Thus, hepcidin seems to be the possible connection between inflammation and decreased functional iron availability, leading to elevated RDW levels. Interleukin-6 is also one of the key executors of prostate enlargement. IL-6 as a potential autocrine growth factor has been shown to be the favorite executor of stromal and epithelial growth in BPH.[14, 19] Elevations in the RDW appear to reflect a state of increased inflammation and impaired iron metabolism. Findings suggest the possibility that the RDW may provide an integrated measure of these underlying processes in BPH. Nickel et al. found a relationship between LUTS and prostatic inflammation.[20] A higher IPSS in patients with an elevated RDW, which may reflect the status of inflammation, was found in this study.

Interestingly, CNS infiltrating Th1 cells kept the largest IFN-γ-

Interestingly, CNS infiltrating Th1 cells kept the largest IFN-γ-positive population, probably due to the inflammatory environment or selective enrichment. Surprisingly, Th1 cells recovered from the LN (pooled peripheral LN (pLN) and mLN) showed a consistent population of IL-17A/IFN-γ double-positive cells (9.1%). Next, we analyzed the expression of cytokines and transcription factors by quantitative real-time RT-PCR in sorted EYFP positive cells

before and after transfer and found that in accordance with the intracellular cytokine staining, tbx21 as well as ifng mRNA were highly upregulated, while the mRNA of il17a and il17f were down regulated (Fig. 1F). In contrast, we did not find a change in the expression levels of Th17-specific transcription Fulvestrant factors rorc and irf4 (Fig. 1F). This indicates that the observed plasticity and coexpression of IL-17A and IFN-γ are based on dual expression of Th1 as well as Th17 specific transcription factors. Collectively, these data clearly illustrate that Th17 cells, once expressing IL-17A and IL-17F, are able to alter their previous cytokine expression pattern in vivo. To analyze whether Th1 cells behave in a similar fashion to Th17

cells, we used a differentiation protocol in which a 2D2-Th1 population with nearly 100% IFN-γ producing cells was generated (Fig. 2A). We transferred 5×106 of these cells to RAG1−/− mice and reanalyzed their fate at the peak clinical EAE symptoms (Fig. 2B). Compared to Th17 cells, transferred 2D2-Th1 cells isolated from CNS and spleen did not shift in large numbers to express

IL-17A, but either kept or lost IFN-γ expression. Surprisingly, Th1 cells recovered from the LN (pooled pLN and mLN) showed a consistent population of IL-17A/IFN-γ double-producing cells (Fig. 2C). The redifferentiation of Th1 cells in LN correlated with a rise in expression levels of IL-17A and IL-17F Resminostat and a slight decrease of IFN-γ mRNA expression (Fig. 2D). In accordance with the upregulation of a Th17 phenotype, rorc expression was nearly 100-fold upregulated in Th1 cells recovered from mLN. In agreement with the relative stability of IFN-γ expression observed after intracellular staining, tbx21 remained stably expressed by Th1 cells (Fig. 2D). Since EAE induces peripheral changes to the immune system and cellular composition, especially in the spleen and the BM, we transferred sorted, non-encephalitogenic reporter cells (IL-17F-CreEYFP) to RAG1−/− mice. Again, we found that a major part of the transferred population lost IL-17 expression and instead, upregulated the expression of IFN-γ (Fig. 3A), showing that the plasticity of the transferred Th17 population can take place independently of EAE. In this experiment, we analyzed pLN separately from mLN (Fig. 3B).

Of particular interest in this context are recent studies on huma

Of particular interest in this context are recent studies on human endothelial cell cultures which documented that above a threshold of 135 mmol/L a stepwise increase in the sodium concentration of the incubation medium progressively increases endothelial cell stiffness, causes inhibition of endothelial NO synthase and decreases release of nitric oxide; this effect was abrogated by the mineralocorticoid receptor spironolactone.30 In addition to aldosterone, digitalis-like endogenous

inhibitors of Na+, K+-ATPase have recently been recognized as one class of agents raising blood pressure in response to sodium loads.31 Recent studies clearly documented minor increases in plasma sodium concentration in hypertensive individuals.32 Changes in plasma sodium concentration are transmitted into the cerebrospinal fluid33 triggering the release of cardiotonic steroids, selleck compound namely, analogues Everolimus cell line of digitalis such as ouabain and marinobufagenin.31 In the Dahl salt-sensitive rat, a standard hypertensive animal model with an underlying mutation of the α-1 Na+, K+-ATPase, chronic salt loading increases the excretion of marinobufagenin in the urine.34 Marinobufagenin causes vasoconstriction35 and is increased

in pathological states of sodium overload, for example uraemia and preeclampsia.35,36 The most convincing proof of a key role of sodium and specifically renal sodium handling in the genesis of hypertension has been provided by studies in which heterozygous carriers of mutations of renal sodium transporters were compared with corresponding normotensive control individuals. For instance, in the study of Fava37 in the Framingham population, heterozygous carriers of the Gitelman mutation failed to have phenomena relating to the Gitelman syndrome, but had significantly Megestrol Acetate lower systolic and diastolic pressures compared to matched controls, obviously as the result of higher renal sodium excretion with a shift in the pressure/natriuresis relationship. In summary, the evidence is overwhelming that current intakes of salt contribute in

a major fashion to the current ‘epidemic’ of hypertension. This justifies public health efforts to reduce salt intakes, particularly in commercial food items,38 since it had been shown that only 15% of current salt intakes can be controlled by the patient, whilst 85% of salt is already contained in commercial food items.39 The Author states that there is no conflict of interest regarding the material discussed in the manuscript. “
“Aim:  Podocytes provide a slit diaphragm to inhibit proteinuria, and nephrin between podocytes functions as a barrier during glomerular filtration. Hepatocyte growth factor (HGF) can improve proteinuria in rodents with various renal injuries, but little is known about the role of HGF in podocyte-based events during glomerulonephritis.

The BabA-MBS was significantly higher in the cancer than the non-

The BabA-MBS was significantly higher in the cancer than the non-cancer group (P= 0.019), but there was no significant difference for SabA-MBS. A weak correlation PLX4032 datasheet between BabA-MBS and SabA-MBS (r= 0.418) was observed, the positive correlation being higher in the cancer than the non-cancer group (r= 0.598 and 0.288, respectively). The isolates were classified into two groups: a BabA-high-binding and a BabA-low-binding group (in comparison to the average for BabA-MBS). The average SabA-MBS in the BabA-high-binding group was significantly higher than in the BabA-low-binding

group (P < 0.0001). Analysis of babA2 middle region diversity (AD1–5) revealed that AD2-type was predominant in isolates irrespective of BabA-MBS. H. pylori BabA-MBS might have an effect on SabA-MBS and relate to the severity of gastric disorders, including gastric cancer. Evaluation of MBS of the combined two adhesins would be helpful for predicting damage in the H. pylori infected stomach. H. pylori is a Gram-negative, spiral and microaerophilic bacterium that colonizes the human stomach. H. pylori infection occurs mostly in early childhood (1) and causes chronic gastritis, peptic ulcer, gastric cancer (2) and gastric mucosa-associated lymphoid tissue lymphoma (3). H. pylori begins its colonization by binding to certain adhesive molecules

on the epithelial cells via H. pylori outer membrane proteins such as BabA, SabA, AlpA, AlpB and HopZ, leading to persistent infection and tissue damage (4–7). Two glycoconjugates, OSI 906 fucosylated Lewis b blood group (Leb) and the sialic acid antigens (sLex and sLea), have been identified as cognate substrate molecules of the H. pylori adhesins, BabA and SabA, respectively (4, 5). BabA and SabA are Etofibrate encoded by the babA2 and sabA genes, respectively, which mediate the attachment of H. pylori to human gastric epithelial cells (4, 5, 8). The relationship between the detection of these genes, babA2 and sabA, with PCR and clinical manifestations has been investigated (9–14).

There is no apparent relationship between the prevalence of sabA and gastric disease types (9). However, the sabA-negative genotype may be attributable to false negative PCR due to subtle mutations in the primer regions. On the other hand, the presence of babA2 has been shown to be associated with chronic gastritis (10), intestinal metaplasia (13) and duodenal ulcer (11), whereas several reports have shown no significant association between babA2 status and clinical manifestations in some countries, including Japan (12, 15, 16). In particular, the babA gene possesses high homologous sequences with minor diversity between babA1, babA2 and babB genes within a microorganism and among individual strains. These suggest that use of several primer pairs in PCR based-detection somewhat mitigates that risk and provides reliable findings.

0 cm radius of the image A behavior was considered to have ended

0 cm radius of the image. A behavior was considered to have ended when an infant looked away, initiated a different type of manual behavior, changed hands, or removed the hand (or hands). Uninterrupted repetitions of a given gesture type were counted as one instance of that categorical type of behavior. Thus, several uninterrupted repetitions of the same manual action were conservatively scored as a single behavior. We evaluated selleck the qualitative (“categorical”) types of manual exploration behaviors as well as the total number of behavior changes initiated in

sequence (“sequential”) for each display. In the Categorical level of analysis, infants’ manual gestures were classified as one of five gross categories of reaching behavior (e.g., touching, grasping, rubbing,

scratching, or patting). These qualitatively different types of reaching behaviors were recorded and tallied for each display. At the categorical level, infants could potentially receive a score between 0 and 5 representing the number of qualitatively different types of manual gestures initiated toward each display. In the Sequential level of analysis, a finer grain assessment of successive actions was reviewed. The total quantity of gesture changes that occurred in sequence were recorded and tallied for each display. Natural Product Library chemical structure For example, if an infant was observed rubbing a picture display with one hand followed by tapping with both hands, followed by rubbing with one hand, then those manual behaviors would be recorded as two categorical gestures and three Idelalisib cell line sequential gestures. For both measures of manual exploration, an impossible preference score was calculated for each infant by computing the total number of behaviors initiated toward the impossible cube divided by the sum of gestures

initiated to both the possible and impossible cube displays. Preference scores were then compared with 50/50 chance. We also documented the frequency of social referencing, vocalizations, and mouthing behaviors as independent and complementary measures of infants’ differential responses toward each type of display. Social referencing was defined as an occurrence of the infant looking to the parent or the experimenter only after the child had initially visually inspected the display at least once. Instances of social referencing were logged each time the child referred back to the parent/experimenter after viewing and/or touching the stimulus display. Social referencing behavior has been a useful indicator of infants’ perceptual judgments and impending actions during an ambiguous, uncertain situation involving novel or unusual stimuli (Klinnert, Emde, Butterfield, & Campos, 1986; Walden & Kim, 2005).

Chemokines produced by neutrophils can direct T lymphocyte matura

Chemokines produced by neutrophils can direct T lymphocyte maturation find more and specifically attract Th17 cells (Pelletier et al., 2010; Lowe et al., 2012). To find whether the infected neutrophil secretions have the capacity to stimulate T helper cells, the expression of CD69 (an activation marker) on T cells was analyzed. The supernatants

from H37Rv-infected neutrophils increased CD69 expression on T cells suggesting modulation of T helper cells through neutrophil-mediated signaling. This is in accordance with a previous study, where increased expression of CD69 was observed on T cells from patients with TB (Wanchu et al., 2009). It has been reported that expression of CXCR3 was increased on naïve T cells following activation and preferentially remains highly expressed on Th1 cells (Qin et al., 1998). In this study, even though there was increased expression of the activation marker CD69, we did not find any modulation in CXCR3 expression on T cells when stimulated see more with infected neutrophil supernatants. To conclude, the present study clearly indicates that H37Rv modulates neutrophils to

the maximum followed by BCG, whereas Mw does not show any influence on the studied neutrophil parameters. This is evidenced from the upregulation in the expression of CD32, CD64, TLR4, and CXCR3; increased TNF-α secretion, and downregulation of early apoptosis in H37Rv-infected neutrophils,

whereas only CD32 expression was increased in BCG-infected neutrophils. Also, secretory products from infected neutrophils were able to modulate T helper cells and monocytes to different extents. Further studies are required to understand whether these varied phenotypical changes induced by H37Rv and BCG on ADAMTS5 neutrophils are related to pathophysiology of these strains. The first author thanks University Grants Commission (UGC) for providing Junior Research Fellowship. Help rendered by the volunteers who donated their blood is greatly acknowledged. The authors declare that there is no conflict of interest. “
“Estrogens act upon nuclear estrogen receptors (ER) to ameliorate cell-mediated autoimmune disease. As most immunomodulatory effects of estrogens in EAE have been attributed to the function of ER-α, we previously demonstrated that ER-β ligand treatment reduced disease severity without affecting peripheral cytokine production or levels of CNS inflammation, suggesting a direct neuroprotective effect; however, the effect of ER-β treatment on the function of immune cells within the target organ remained unknown. Here, we used adoptive transfer studies to show that ER-β ligand treatment was protective in the effector, but not the induction phase of EAE, as shown by decreased clinical disease severity with the preservation of axons and myelin in spinal cords.

After centrifugation at 5000 g 10 min, supernatants were frozen a

After centrifugation at 5000 g 10 min, supernatants were frozen at −80°C until used. Extracts (50 µg protein/lane) subjected to 10% SDS-PAGE were immunoblotted with antibodies to total Bad, phosphorylated Bad (Santa Cruz Biotechnology) and revealed by enhanced chemiluminescence (ECL) detection system (Pierce). Densitometric analysis of protein levels was performed with ImageQuant software. The frequency of

apoptotic acini cells was also assessed by flow cytometry analysis with Annexin V/IP double staining following the manufacturer’s recommendations (BD). Flow cytometry data were acquired in a FACSAria cytometer® and results analysed using WinMDI software®. For bax expression assays, acinar cells were homogenized either freshly or after induction with TNF-α and RT–PCR experiments were carried out as indicated click here above and previously [16]. Statistical significance of differences was determined by the two-tailed t-test Smoothened Agonist for independent populations. When multiple comparisons were necessary, the Student–Newman–Keuls test was used after analysis of variance. Differences between groups were considered significant at P < 0·05. Figure 1a shows the expression kinetics of VIP and their receptors in submandibular

glands isolated from NOD mice of different ages from postnatal day 2 to 20 weeks of age. Compared to normal mice, NOD mice showed the highest level of VIP expression at 4 weeks of age and decreased thereafter. The progressive decrease in VIP expression from the fourth week takes place with no changes in VPAC1 and VPAC2 receptors. A clear reduction

of VIP levels was evident in NOD submandibular glands at 16 weeks (-)-p-Bromotetramisole Oxalate of age (Fig. 1a), which was confirmed by qRT–PCR (Fig. 1b). The decline in VIP/VPAC1 relative expression over time is similar to the kinetics of neural nitric oxide synthase (nNOS) activity and salivary secretion loss shown previously [12]. NF-κB appears as an intracellular pivotal determining factor that conditions the apoptotic or survival fate of TNF-α-stimulated cells [28]. Thus, we analysed NF-κB activation and apoptosis in response to TNF-α in NOD acinar cells. As shown in Fig. 2a, acinar cells from NOD glands present a basal translocation of p65 of NF-κB to the nucleus (merge image with PI-stained nuclei) that is not seen in normal BALB/c mice. Consistent with this, WB analysis of I-κB in the cytosolic fraction or p65 in the nuclear fraction revealed that p65 appeared located to the nucleus, while I-κB expression was increased in cytosol of acini in basal conditions (Fig. 2b). Moreover, when treated in vitro with TNF-α, NOD mice acinar cells showed an abnormal NF-κB activation kinetics compared with BALB/c acinar cells (Fig. 2a,b).