In this study we have addressed the potential utility of immunotherapy PI3K inhibitor using regulatory T cells (Treg) to treat murine autoimmune cholangitis. In particular, we have taken advantage of our ability to produce portal inflammation and bile duct cell loss by transfer of CD8+ T cells from the dominant negative form of transforming growth factor beta receptor type II (dnTGF-βRII) mice to recombination-activating gene (Rag)1–/– recipients. We then used this robust established adoptive transfer system and co-transferred CD8+ T cells from dnTGF-βRII mice with either C57BL/6 or dnTGF-βRII forkhead box protein 3 (FoxP3+) T cells. Recipient mice were monitored for histology,

including portal inflammation and intralobular biliary cell damage, and also included a study of the phenotypical changes in recipient lymphoid populations and local and systemic cytokine production. Importantly, we report herein that adoptive transfer of Treg from C57BL/6 but not dnTGF-βRII

mice significantly reduced the pathology of autoimmune cholangitis, including decreased portal inflammation and bile duct damage as well as down-regulation of the secondary inflammatory response. Further, to define the mechanism of selleckchem action that explains the differential ability of C57BL/6 Treg versus dnTGF-βRII Treg on the ability to down-regulate autoimmune cholangitis, we noted significant differential expression of glycoprotein A repetitions predominant (GARP), CD73, CD101 and CD103 and a functionally significant increase in interleukin (IL)-10 in Treg from C57BL/6 compared to dnTGF-βRII mice. Our data reflect the therapeutic potential of wild-type CD4+ FoxP3+ Treg in reducing the excessive T cell responses of autoimmune cholangitis, which has significance for the potential immunotherapy of PBC. “
“Cryptosporidium parvum infects intestinal Buspirone HCl epithelial cells and is commonly the parasite

species involved in mammalian cryptosporidiosis, a major health problem for humans and neonatal livestock. In mice, immunologically mediated elimination of C. parvum requires CD4+ T cells and IFN-γ. However, innate immune responses also have a significant protective role in both adult and neonatal mice. NK cells and IFN-γ have been shown to be important components in immunity in T and B cell-deficient mice, but IFN-γ-dependent resistance has also been demonstrated in alymphocytic mice. Epithelial cells may play a vital role in immunity as once infected these cells have increased expression of inflammatory chemokines and cytokines and demonstrate antimicrobial killing mechanisms, including production of NO and antimicrobial peptides. Toll-like receptors facilitate the establishment of immunity in mice and are involved in the development of inflammatory responses of infected epithelial cells and also dendritic cells. Around 20 recognized species of the apicomplexan Cryptosporidium infect the gastro-intestinal tract of vertebrates.

In contrast, increased lung neutrophils were seen in the Nod2−/− animals at 24 h. BTK inhibitor Analysis of cytokine production at 4 h post infection revealed a significant decrease in proinflammatory cytokines in the Nod1−/− animals when compared to WT animals. In contrast, increased 4-h proinflammatory cytokines were seen in the Nod2−/− animals. Furthermore, the lungs of both Nod1−/− and Nod2−/− mice had significantly increased pro-inflammatory

cytokine levels at 24 h, suggesting possible suppressive roles for later stages of infection. Together, our data suggest that although both NOD1 and NOD2 can detect Legionella, these receptors modulate the in vivo pulmonary immune response differently. The immune response to intracellular pathogens in the lung initially involves detection of the organisms through a set of receptors located on the cell surface or endosomal compartment (Toll-like receptors (TLR)) or in the cytoplasm (Nod-like receptors (NLR)) and retinoic acid inducible gene I-like receptors (RLR). Based on the type of foreign material (dsRNA, peptidoglycan, lipopolysaccharide)

and location (extracellular, endosomal, cytoplasm), pathogens stimulate distinct sets of receptors to activate the immune response. Legionella pneumophila (Lp), an organism known to persist within water-borne amoeba, usually infects humans Angiogenesis inhibitor as a terminal host after exposure to contaminated water systems 1. Lp replicates within the phagolysosome of the macrophage and secretes bacterial products into the cytosol of the cell through a type IV secretion system (T4SS) which is known to translocate both DNA and proteins that impair the destruction of the organism 2. Previous Sinomenine work has identified several innate immune receptors that are responsible for the detection of Lp in the murine model of infection. NAIP5 (Baculoviral inhibitor of apoptosis repeat-containing 1e protein (Birc1e)), NLRC4 (IL-1β converting enzyme-protease activating factor (Ipaf)), and caspase-1 have been shown to be important in restriction of Lp replication both in vivo and in vitro3–6. TLR5, TLR2, and TLR9

detect Lp and regulate the in vivo immune response to Lp 7–10. Mice deficient in myeloid differentiation factor 88 (Myd88), an adaptor protein for many TLR, are highly susceptible to in vivo Lp infection with lack of an early immune response, inability to control bacterial replication, and enhanced mortality 8, 10. More recently, receptor-interacting serine-threonine kinase (RIP2), an adaptor for nucleotide-binding oligomerization domain-1 (NOD1) and nucleotide-binding oligomerization domain-2 (NOD2), was found to regulate Lp replication in the lung, but only on a Myd88−/− genetic background 11. Since Lp is known to replicate intracellularly and can translocate substances to the cytosol via its type IV secretion system, we hypothesized that the cytosolic NLR may be important in control of the innate immune response to Lp.

2006AA02A109. 2006AA02A115); the National Natural Science Foundation of China (no. 30570771); the Beijing Ministry of Science see more and Technology (no. D07050701350701) and the Cheung Kong Scholars programme. All disclosures were provided in the ‘Acknowledgements’ section. “
“Thomas Jefferson University, Philadelphia, PA, USA Vaccinia virus (VV) has been used globally as a vaccine to eradicate smallpox. Widespread use of this viral vaccine has been tempered in recent years

because of its immuno-evasive properties, with restrictions prohibiting VV inoculation of individuals with immune deficiencies or atopic skin diseases. VV infection is known to perturb several pathways for immune recognition including MHC class II (MHCII) and CD1d-restricted antigen presentation. MHCII and CD1d molecules associate with a conserved intracellular chaperone, CD74, also known as invariant chain. Upon VV infection, cellular Dabrafenib CD74 levels are significantly reduced in antigen-presenting cells, consistent with the observed destabilization of MHCII molecules. In the current study, the ability of sustained CD74 expression to overcome VV-induced suppression of antigen presentation was investigated. Viral inhibition of MHCII antigen presentation could be partially ameliorated by ectopic expression of CD74 or by infection of cells with a recombinant VV encoding murine CD74 (mCD74-VV). In contrast,

virus-induced disruptions in CD1d-mediated antigen presentation persisted even with sustained CD74 expression. Mice immunized with the recombinant mCD74-VV displayed greater protection during VV challenge and more robust anti-VV antibody responses. Together, these

observations suggest that recombinant VV vaccines encoding CD74 may be useful tools to improve CD4+ T-cell responses to viral and tumour antigens. “
“TCR repertoire diversity can influence the efficacy of CD8+ T-cell populations, with greater breadth eliciting better protection. We analyzed TCRβ diversity and functional capacity for influenza-specific CD8+ T cells expressing a single TCRα chain. Mice (A7) transgenic Cyclin-dependent kinase 3 for the H2KbOVA257–264-specific Vα2.7 TCR were challenged with influenza to determine how fixing this “irrelevant” TCRα affects the “public” and restricted DbNPCD8+versus the “private” and diverse DbPACD8+ responses. Though both DbNPCD8+ and DbPACD8+ sets are generated in virus-primed A7 mice, the constrained DbNPCD8+ population lacked the characteristic, public TCRVβ8.3, and consequently was reduced in magnitude and pMHC-I avidity. For the more diverse DbPACD8+ T cells, this particular forcing led to a narrowing and higher TCRβ conservation of the dominant Vβ7, though the responses were of comparable magnitude to C57BL/6J controls. Interestingly, although both the TCRβ diversity and the cytokine profiles were reduced for the DbNPCD8+ and DbPACD8+ sets in spleen, the latter measure of polyfunctionality was comparable for T cells recovered from the infected lungs of A7 and control mice.

Samples were read on a FACSCanto (BD Biosciences) and analyzed using FlowJo Software Version 8.7. Gates for FOXP3+ cells were set based on fluorescence minus one controls 16 and for cytokines on unstimulated, but stained, samples. The production of lentivirus and transduction of T cells has been previously described 16. Control ΔNGFR+-transduced T cells and FOXP3-transduced T cells were purified (>90% based on surface NGFR expression) and expanded in rhIL-2-containing media (100 U/mL, Chiron) 16. T cells in the resting Microtubule Associated inhibitor phase (10–13 days after activation) were washed and rested in IL-2 free media overnight, and stimulated with αCD3/αCD28-coated beads at a 1:8 cell:bead

ratio for 72 h. The CXCL8 promoter (region −1793 to +49; 1,842 bp) was amplified from human genomic DNA and cloned into pGL3. Jurkat cells were transiently transfected as described 27 with pGL3 or pGL3-CXCL8 and a renilla luciferase reporter vector (pRL-TK), in the presence or absence of FOXP3. After 24 h, cells were stimulated with PMA (10 ng/mL) and Ca2+ ionophore (500 ng/mL) for 6 h. Luciferase

activity was measured using a luminometer (EG&G Burthold) and a Dual Luciferase Reporter Assay System (Promega). All values were normalized to renilla luciferase activity and expressed relative to unstimulated controls. Supernatants (235 μL) from FACS-sorted CD4+CD25− Tconv and CD4+CD25hi Tregs cultured at 1×106/mL for 72 h with αCD3/αCD28-coated beads at a 1:8 cell:bead ratio in complete medium, but with serum replaced by 1% human serum albumin, were added to the lower chamber of a transwell plate (Corning this website HTS 96 well transwell, 3.0 μm pore size). Neutrophils were isolated using a Ficoll separation followed by a 6% dextran gradient, and 100 000 cells were added to the upper chamber of the transwell plate. In some cases, anti-CXCL8 mAb (2A2, 150 μg/mL, BD Biosciences) was added to the lower chamber for 1 h at 37°C prior to neutrophil addition. This amount of mAb neutralized migration in response to at least 8 ng/mL of CXCL8

(data not shown). Dilutions ranging from Rutecarpine 200 pg/mL to 100 ng/mL of rhCXCL8 (eBiosciences) were added to the lower chamber as a positive control. After 30 min of incubation at 37°C, 50 000 surfactant-free white sulfate latex beads (4.9 μm, Dynamics) were added to lower chamber supernatants, and the number of neutrophils which had migrated to the lower chamber per 10 000 beads were counted by flow cytometry based on FSC and SSC parameters. All analysis for statistically significant differences was performed using the Student’s paired t-test. p-Values less than 0.05 (indicated by *) were considered significant. All cultures were performed in triplicate and error bars represent the SD unless otherwise indicated. Supported by the Canadian Institutes of Health Research (MOP 57834 to M. K. L.), a CIHR New Emerging Team grant in Immunoregulation and IBD (IIN84037 to C. P., T. S. S. and M. K. L.), and Stem Cell Technologies Inc.

Underlying mechanisms would include the

cleavage by calpains of several focal adhesion components leading to the turnover of integrin-dependent cell–matrix adhesions that is required for cell movement 17 and of proteins linked to actin bundles and integrins, such as α-actinin 26, 27. In addition, in vivo, endothelial cell calpains could be implicated in lymphocyte transendothelial migration, as they participate in the assembly of docking structures involved in diapedesis process 27. Thus, evidence is accumulating to suggest that the calpain check details inhibition by calpastatin is sufficient to limit lymphocyte recruitment, as we previously demonstrated in a model of peritonitis 13. Besides the observed decrease in T-cell migration, mechanisms underlying delayed rejection could involve reduced proliferative responses. But in vitro experiments showed conclusively that the calpain inhibition by calpastatin transgene rather increased T-cell proliferation. One possible explanation would be that calpastatin prevented the proteolytic cleavage of the γc chain in IL-2 receptor, thereby amplifying

IL-2-dependent proliferative responses 18, 19. Consistent with this model, we found an increase in IL-2-induced STAT5 phosphorylation in T cells from CalpTG as compared with WT mice (data not shown). However, it is not yet clear whether this mechanism occurs in vivo, as 1 IL-2 expression is limited in CalpTG mice and 2 γc overexpression would increase T-cell response to several cytokines MK-8669 nmr sharing this common receptor (e.g. IL-4, IL-9, IL-21 in addition to IL-2). TH phenotype is believed to control allograft rejection, each phenotype producing its own set of cytokines 1. Hence, one supplementary explanation for the observed delay in skin allograft rejection could

be a change in the level of IFN-γ, IL-4/IL-10, second and IL-17 produced by TH1, TH2, and TH17 cells, respectively. In fact, in vitro experiments showed that the calpain inhibition by calpastatin transgene affected mainly the IL-17 expression. One possible explanation for this finding is again that calpastatin limited proteolytic cleavage of the γc chain in IL-2 receptor, thereby amplifying IL-2-dependent inhibition of TH17 generation. A proper role of IL-17 in allograft rejection has recently been proposed 28. Nevertheless, its importance would be limited to rejection responses in older transplant recipients 29 and in case of minor antigen disparity 30. Thus, the limited TH17 response in CalpTG mice confirms strongly our finding of a reduced cleavage of the γc chain in IL-2 receptor but does not provide an additional explanation for delayed allograft rejection. Finally, our findings do not exclude effects of calpastatin transgene expression on T-cell functions other than their recruitment and differentiation. Interestingly, our data demonstrate a marked decrease in specific cytolytic capacity of alloreactif lymphocytes in CalpTG mice as compared with WT mice.

e. in nonstressed females), prolonged exposure to chronic stress results in an attenuated CORT response to stimuli, which predisposes to higher susceptibility to pathogenic autoimmunity. A comprehensive and widely accepted biological model linking stress, CORT and autoimmune diseases is currently lacking. Although numerous studies demonstrated that CORT suppresses autoimmune diseases in humans and in animal models [15, 35, 36], other studies indicate that low levels of CORT or certain stress

paradigms may skew to proinflammatory conditions [14, 18, 19, 37-42]. In the present study we found that CVS exacerbated EAE in female mice despite the overall stress-induced increase in CORT levels, which was also reported previously [32, 43, 44]. The elevated urine CORT levels selleck in females were, however, significantly lower on the fourth week of stress and reached those of nonstressed females. In addition, CORT MLN0128 cell line levels failed to increase toward disease onset (9 days postimmunization) in stressed as compared with nonstressed mice. Following the disease onset (14 and 21 days postimmunization) CORT levels in stressed mice markedly increased to levels higher than those observed during stress, and remained similar to those observed in nonstressed mice throughout the course of the disease. These results suggest that the temporarily decreased functionality of the HPA axis in stressed female mice, which resulted in a

delayed CORT response to MOG35-55 immunization, could at least partially account for the initial exacerbation of the disease over that induced in nonstressed mice. An important Adenosine finding in our study was that although stressed male mice demonstrated decreased weight gain and increased

anxiety index similar to females, they showed significantly lower levels of urine CORT under basal, stress and EAE conditions. Although to a less extent, blood CORT levels were also lower in male than in female mice. However, whereas primarily free CORT was observed in the urine, only a small fraction (less than 10%) of the blood CORT was free, with levels similar between male and female mice, while the rest was presumably bound to CORT-binding globulin [45]. Higher CORT levels were previously documented in female compared with male Sprague–Dawley rats [46]. Furthermore, CORT secretion has been previously shown to attenuate EAE severity, suggesting that the HPA axis suppresses autoimmune disease progression [47-49]. Taking together, it is reasonable to assume that although similar levels of free CORT were observed in male and female mice, the overall higher basal levels of CORT in nonstressed females attenuated their EAE severity. The role of free versus bound CORT in gender-related EAE susceptibility should be further investigated. Given the antiinflammatory properties of CORT, we asked why CVS generally exacerbated EAE in female mice.

In addition, in the normally formed glomeruli there was a significant increase in size, indicative of glomerular hypertrophy and thus hyperfiltration. The variability in the proportion of abnormal glomeruli in the outer renal cortex between preterm infants suggests that there may be differences in haemodynamics, and/or other factors in the postnatal environment of the infant (such as

exposure to Opaganib nephrotoxic drugs, oxygen supplementation, mechanical ventilation and co-morbidities) which may be negatively impacting glomerulogenesis[3] (Fig. 1). In this regard, there is a major haemodynamic transition at the time of birth when blood pressure and heart rate are markedly elevated[10] and blood flow to the kidneys is increased.[11] Hence, it is possible that the developing capillaries of immature glomeruli are not prepared for the haemodynamic transition at birth and their formation is adversely affected. Indeed, we have recently shown that there is injury to the wall of the aorta as a result of preterm delivery.[12] LY2109761 datasheet In future studies, it is imperative to determine the cause of the glomerular abnormalities in the preterm kidney, in order to maximize the number of functional nephrons at the beginning of life; this will likely lead to short-term and long-term benefits to renal health. “
“We recommend that all candidates

for kidney transplant are screened for cardiovascular risk factors (1B). Indicators of high risk include (1B): Older age. Diabetes mellitus. Abnormal echocardiogram (ECG). Previous ischaemic heart disease or congestive heart

failure. Increased duration of dialysis. Smoker. We suggest that kidney transplant candidates with a low clinical risk of cardiovascular disease do not require stress testing for coronary artery disease (2B). We suggest that kidney transplant candidates with a moderate or high clinical risk of Liothyronine Sodium cardiovascular disease undergo cardiac stress testing prior to transplantation (2B). The following should be noted in relation to cardiac stress testing in dialysis patients: Exercise ECG has a poor predictive value in patients on dialysis (2B). The use of a cardiac stress test such as dipyridamole thallium testing or stress echocardiography is predictive of significant coronary artery disease and major cardiac events in patients with higher clinical risk. Where possible we recommend that this testing should be performed without concurrent β-blocker therapy (1B). As the prognostic accuracy of cardiac stress testing in dialysis patients is of limited duration, it is suggested that testing be repeated in high risk patients. The interval at which testing should take place has not been well defined; however, the predictive value of a positive test diminishes after 24 months (2C). We recommend that coronary angiography be considered for kidney transplant candidates with abnormalities on screening procedures (1B). We suggest that the benefit of revascularization prior to transplantation be reviewed on an individual basis (2C).

Therefore, it is not surprising that, at least for the present, an earlier start of long-term Inhibitor Library research buy dialysis than currently applied is not encouraged in Taiwan. Whether this may change will have to await the completion of a multicentre patient-directed randomized study currently underway in Taiwan to compare clinical outcome with respect to renal function at initiation. Despite the absence of high level evidence, a number of expert groups have developed clinical practice guidelines about when to initiate dialysis. These groups include CARI,5 Kidney Disease Outcomes Quality Initiative (K/DOQI) and Canadian Society of Nephrology and European Best Practice

Guidelines. Their recommendations are similar. CARI recommends that dialysis should be initiated before the development of uraemic symptoms and complications including malnutrition; that quality of life should be taken into consideration; and that in an otherwise well patient dialysis preparation should commence at a GFR of 10 mL/min and dialysis be initiated by a GFR of 5 mL/min (Table 1). In addition, individual countries have developed regulations

or guidelines about dialysis initiation for local application. For example, in Taiwan the Bureau of National Health Policy has set the following regulations for initiating dialysis: (i) absolute, CCr less than 5 mL/min or serum creatinine more than 10 mg/dL

(884 µmol/L); and (ii) relative, CCr less than 15 mL/min Acalabrutinib datasheet or serum creatinine more than 6 mg/dL (530 µmol/L), plus the presence of fluid overload or other uraemic emergency. According to the Taiwan dialysis registry data (during 2001 and 2004), 90% of the incident ESKD patients started long-term Exoribonuclease dialysis according to absolute indications, while 10% followed relative indications. Following a study endorsed by its Ministry of Health and Welfare,13 Japan introduced recommendations for initiation of haemodialysis almost 20 years ago (Table 2). The recommendations were based on scores for uraemic symptoms, level of renal function, activity and age; with a score exceeding 60, initiation of haemodialysis was recommended. These recommendations appeared to change clinical practice because the percentage of patients commencing haemodialysis with a score less than 60 rose from 3% in 1994 to 22% in 2006, and mean serum creatinine level at initiation fell from 10.6 ± 3.7 to 8.4 ± 3.6 mg/dL (937 ± 327 to 743 ± 318 µmol/L, respectively).14 These observations are confounded by changes in mean age (57 vs 66 years) and incidence of diabetes as the cause of ESKD (29% vs 43%) at initiation in 1994 versus 2006. It is likely that the recommendations about when to initiate haemodialysis will be modified.

7% of the FFRs showed abnormal cystometry, characterized primarily by increased phasic contractions.30 Hypercholesterolemia is a component of metabolic syndrome. The diagnostic criteria for metabolic syndrome are defined differently by various organizations, but all definitions of metabolic syndrome include dyslipidemia.31–35 According to the most commonly used Adult Treatment Panel III (ATP III) definition, metabolic syndrome is characterized by the presence

of three or more of the following five characteristics: (i) waist circumference greater than 102 cm for male 5-Fluoracil mouse or greater than 88 cm for female; (ii) triglycerides 150 mg/dL or greater; (iii) HDL cholesterol less than 40 mg/dL for male or less than 50 mg/dL

for female; (iv) systolic blood pressure of 130 mm Hg or greater, or diastolic blood pressure of 85 mmHg or greater; (v) fasting glucose of 110 mg/dL or more.33 High-fat diet rats used in the aforementioned studies had not only hypercholesterolemia but also other components of metabolic syndrome, such as obesity, hypertension and insulin resistance. In the report by Son et al.10, the mean body weight in the cholesterol group was significantly higher than that in the control Bortezomib manufacturer group. Hyperlipidemic rats in the study by Rahman et al.9 also had a significantly higher mean body weight than the control rats, in addition to a higher mean arterial blood pressure, though without statistical significance. In the report

by Huang et heptaminol al.11, the mean body weight and level of fasting glucose were elevated in high-fat diet rats. Furthermore, high-fat diets have been used to model obesity, dyslipidemia and insulin resistance in rodents for many decades because the complications developed by high-fat diets resemble the human metabolic syndrome.36 Therefore, the DO in high-fat diet rats cannot be said to have been affected by a single factor like hypercholesterolemia; rather, it is more reasonable to say that all components of metabolic syndrome have an effect on the occurrence of DO. Metabolic syndrome is known to cause autonomic sympathetic overactivity through complex and incompletely elucidated mechanisms.37 Hyperinsulinemia, a key concept of metabolic syndrome, is associated with increased sympathetic activity via enhanced glucose metabolism in ventromedial hypothalamic neurons.38 Increased activation of the α-adrenergic pathway increases smooth muscle contraction throughout male genitourinary tract structures, including the prostate, bladder neck, and urethra.39 Therefore, ANS overactivity may contribute to DO. An association was also shown between ANS overactivity and voiding dysfunction in a spontaneously hypertensive rat (SHR) model. Steers et al.40 reported that SHRs voided three times more frequently than normotensive rats and that such frequency can be reduced by alpha-adrenoceptor antagonists.

Polyethylene catheters, filled with heparinized saline (100 U/ml), were www.selleckchem.com/products/epacadostat-incb024360.html inserted into the ascending aorta, via the right carotid artery, and into the left femoral artery. The former catheter was connected to a pressure

transducer (PDCR 75/1; Druck Ltd., Groby, UK). When the blood pressure had remained stable for at least 20 min, the arterial blood perfusion of the whole pancreas, islets, duodenum, colon, adrenal glands and kidneys was measured with a microsphere technique [14]. Briefly, a total of 1.5–2.0 × 105 non-radioactive microspheres (EZ-Trac™; Triton Microspheres, San Diego, CA USA), with a diameter of 10 μm, were injected via the catheter with its tip in the ascending aorta during 10 s. Starting 5 s before the microsphere injection, and continuing for a total of 60 s, an CH5424802 concentration arterial blood sample

was collected by free flow from the catheter in the femoral artery at a rate of approximately 0.4 ml/min. The exact withdrawal rate was confirmed in each experiment by weighing the sample. Arterial blood was collected from the carotid catheter for determination of blood glucose and serum insulin concentrations as given below. The animals were then killed, and the pancreas and adrenal glands were removed in toto, blotted, weighed and treated with a freeze-thawing technique, which visualized the pancreatic islets and microspheres [15]. Approximately 100 mg each of the duodenum, colon and left kidney were also removed and treated in the same way. The microspheres in the organs were then counted in a microscope equipped with both bright- and dark-field illumination (Wild M3Z; Wild Heerbrugg Ltd., Heerbrugg, Switzerland). The blood flow values were calculated according to the formula Qorg = Qref × Norg/Nref where Qorg is organ blood flow (ml/min), Qref is withdrawal rate of the reference sample, Norg is number of microspheres present in the organ and Nref is number of microspheres in the reference sample. The number of microspheres in the arterial reference sample was

determined by sonicating the blood, and then transferring samples to glass microfibre filters (pore size <0.2 μm), and then counting the number Thalidomide of microspheres in the microscope referred to above. Pancreatic-duodenal transplantations.  This procedure has been described in detail elsewhere [16]. Briefly, the donor was anaesthetized with an intraperitoneal injection of ekviticine (chloral hydrate and pentobarbital; Apoteksbolaget, Umeå, Sweden) and placed on a heated operating table. The whole pancreas, together with approximately 5 cm (1 g) of the duodenum, was dissected free from surrounding tissues. Through a catheter in the abdominal aorta, the preparation was flushed with 5–7 ml of cold (4 °C) UW-solution (Via-Span™; Du Pont Pharmaceuticals Inc., Wilmington, DE, USA) at a pressure of approximately 100 cm H2O. The warm ischaemia time was <2 min.