TLR-Stimulated Eosinophils Mediate Recruitment and Activation of NK Cells In Vivo.

Eosinophils like many myeloid innate immune cells can provide cytokines and chemokines for the activation of other immune cells upon TLR stimulation. When TLR-stimulated eosinophils were inoculated i.p. into wild-type mice, and NK cells were rapidly recruited and exhibited antitumour cytotoxicity. However, when mice depleted of CD11c+ cells were used, a marked decrease in the number of recruited NK cells was observed. We postulated that CpG or LPS from the injected eosinophils could be transferred to host cells, which in turn could recruit NK cells. However, by inoculating mice deficient in TLR4 or TLR9 with LPS or CpG-stimulated eosinophils respectively, NK cell recruitment was still observed alongside cytotoxicity and IFNγ production. CpG stimulation of eosinophils produced the pro-inflammatory cytokine IL-12 and the chemokine CXCL10, which are important for NK cell activation and recruitment in vivo. To demonstrate the importance of CXCL10 in NK cell recruitment, we found that CpG-stimulated eosinophils pretreated with the gut microbial metabolite butyrate had reduced expression and production of CXCL10 and IL-12 and concomitantly were poor at recruitment of NK cells and inducing IFNγ in NK cells. Therefore, eosinophils like other innate immune cells of myeloid origin can conceivably stimulate NK cell activity. In addition, products of the gut microbiota can be potential inhibitors of NK cell.

The Abl-1 Kinase is Dispensable for NK Cell Inhibitory Signalling and is not Involved in Murine NK Cell Education.

Natural killer (NK) cell responsiveness in the mouse is determined in an education process guided by inhibitory Ly49 and NKG2A receptors binding to MHC class I molecules. It has been proposed that inhibitory signalling in human NK cells involves Abl-1 (c-Abl)-mediated phosphorylation of Crk, lowering NK cell function via disruption of a signalling complex including C3G and c-Cbl, suggesting that NK cell education might involve c-Abl. Mice deficient in c-Abl expression specifically in murine NK cells displayed normal inhibitory and activating receptor repertoires. Furthermore, c-Abl-deficient NK cells fluxed Ca2+ normally after triggering of ITAM receptors, killed YAC-1 tumour cells efficiently and showed normal, or even slightly elevated, capacity to produce IFN-γ after activating receptor stimulation. Consistent with these results, c-Abl deficiency in NK cells did not affect NK cell inhibition via the receptors Ly49G2, Ly49A and NKG2A. We conclude that signalling downstream of murine inhibitory receptors does not involve c-Abl and that c-Abl plays no major role in NK cell education in the mouse.

Storage of red blood cells in a novel polyolefin blood container: a pilot in vitro study.

BACKGROUND AND OBJECTIVES: The present general plasticizer di-2-ethylhexyl-phthalate in polyvinylchloride (PVC) blood bags is only physically dispersed in PVC and will therefore leach into blood components. The objective of this study was to perform a first preliminary red blood cell (RBC) storage evaluation in a new blood bag manufactured of polyolefin without any inclusion of potentially migrating substances. STUDY DESIGN AND METHODS: This is a RBC storage study for 42 days. Blood collection was performed in a polyolefin-based PVC-free blood bag. RBCs were prepared within 8 h. Two different RBC additive solutions were used, either PAGGS-M or PAGGG-M. We weekly measured pH, K+ , glucose, lactate, haemolysis, red cell ATP and 2,3-DPG. RESULTS: RBC storage in PAGGS-M resulted in high haemolysis levels already after 21 days, exceeding the European maximum limit of 0·8%, and low ATP levels by the end of the storage period. With PAGGG-M, haemolysis exceeded 0·8% after 28 days of storage. For additional parameters, the results were comparable to those of previous studies in conventional blood bags. CONCLUSION: This is a first preliminary study of RBC storage in a new type of blood bags. PAGGG-M gave encouraging results except for its inability to prevent increased haemolysis. There will be room for further development of RBC additive solutions to address the haemolysis problems. Plasma should also be tested regarding the stability of coagulation and activation pathway variables. There may also be a potential for future use of the bag for preparation of pooled buffy-coat-derived platelets.

Characterisation of maternal human leukocyte antigen class I antibodies in suspected foetal and neonatal alloimmune thrombocytopenia.

OBJECTIVES: To investigate the specificities and level of HLA class I antibodies in selected cases referred for suspected foetal and neonatal alloimmune thrombocytopenia (FNAIT). BACKGROUND: FNAIT occurs in 1 : 1-2000 live births, whereas maternal immunisation against human leukocyte antigen (HLA) class I is common. Whether HLA class I antibodies alone can cause FNAIT is debatable. MATERIAL AND METHODS: A total of 260 patient samples were referred between 2007 and 2012. Referrals with maternal HLA class I antibodies and no other cause for the neonatal thrombocytopenia were included for analysis (cases, n = 23). HPA-1a negative mothers were excluded. Control groups were screened positive mothers of healthy neonates (controls, n = 33) and female blood donors (blood donors, n = 19). LABScreen single antigen HLA class I beads was used for antibody analysis. Clinical records were reviewed for cases. RESULTS: All groups had broad antibody reactivity. Cases had more antibodies with high SFI levels compared with the controls (SFI>9999; medians 26, 6 and 0; P < 0·05) and higher overall median HLA-ABC and HLA-B SFI (P < 0·05). Many of the antibodies were reactive with rare alleles. When reviewing the clinical records, several of the cases had other contributing factors to the thrombocytopenia. There was no correlation between foetal platelet count and antibody levels. CONCLUSION: Mothers of thrombocytopenic neonates had higher levels of HLA class I antibodies compared with control groups of women with healthy children and female blood donors. However, clinical outcome and antibody response correlated poorly in the heterogeneous case group, indicating a multifactorial cause to the thrombocytopenia in the majority of cases.

Circulation stabilizing therapy and pulmonary high-resolution computed tomography in a porcine brain-dead model.

BACKGROUND: Currently 80% of donor lungs are not accepted for transplantation, often due to fluid overload. Our aim was to investigate if forced fluid infusion may be replaced by a new pharmacological therapy to stabilize circulation after brain death in an animal model, and to assess therapy effects on lung function and morphology trough blood gas parameters and state-of-the-art High-resolution CT (HRCT). METHODS: Brain death was caused by surgical decapitation. To maintain mean aortic pressure > 60 mmHg, pigs were treated with forced electrolyte solution infusion (GI; n = 6) or the pharmacological therapy (GII; n = 11). GIII (n = 11) were non-decapitated controls. Lung function was investigated with blood gases and lung morphology with HRCT. RESULTS: GI pigs became circulatory instable 4-6 h after brain death in spite of forced fluid infusion, five pigs showed moderate to severe pulmonary edema on HRCT and median final PaO2 /FiO2 was 29 kPa (Q1; Q3; range 26; 40; 17-76). GII and GIII were circulatory stable (mean aortic pressure > 80 mmHg) and median final PaO2 /FiO2 after 24 h was 72 kPa (Q1; Q3; range 64; 76; 53-91) (GII) and 66 kPa (55; 78; 43-90) (GIII). On HRCT, only two pigs in GII had mild pulmonary edema and none in GIII. More than 50% of HRCT exams revealed unexpected lung disease even in spite of PaO2 /FiO2 > 40 kPa. CONCLUSION: Pharmacological therapy but not forced fluid infusion prevented circulatory collapse and extensive HRCT verified pulmonary edema after acute brain death. HRCT was useful to evaluate lung morphology and revealed substantial occult parenchymal changes justifying efforts toward a more intense use of HRCT in the pre-transplant evaluation.

Comparability of patients with ANCA-associated vasculitis enrolled in clinical trials or in observational cohorts.

OBJECTIVES: To analyse the differences between patients with granulomatosis with polyangiitis (GPA) or microscopic polyangiitis (MPA) entered into randomised clinical trials (RCTs) and those followed in large observational cohorts. METHODS: The main characteristics and outcomes of patients with generalised and/or severe GPA or MPA with a five-factor score ≥ 1 enrolled in the French Vasculitis Study Group (FVSG) or the US-Canadian-based Vasculitis Clinical Research Consortium cohorts were compared to those enrolled in one of 2 FVSG clinical RCTs (WEG91, WEGENT) or 3 European Vasculitis Society clinical trials (CYCLOPS, CYCAZAREM, IMPROVE). RESULTS: 657 patients (65.3% with GPA) in RCTs were compared to 437 in cohorts (90.6% with GPA). RCT patients were older at diagnosis than the cohort patients (56.6 ± 13.9 vs. 46.8 ± 17.3 years), had higher Birmingham vasculitis activity score (19.5 ± 9.1 vs. 16.9 ± 7.4), and more frequent kidney disease (84.0% vs. 54.9%) but fewer ear, nose, and throat symptoms (56.8% vs. 72.2%). At 56 months post-diagnosis, mortality and relapse rates, adjusted for age and renal function, were higher for patients with GPA in RCTs vs. cohorts (10.7% vs. 2.5% [p=0.001] and 22.5% vs. 15.6% [p=0.03], respectively) but similar for patients with MPA (6.2% vs. 6.6% [p=0.92] and 16.6% vs. 10.1% [p=0.39], respectively). CONCLUSIONS: Patients with GPA or MPA in RCTs and those in observational cohorts show important differences that should be remembered when interpreting results based on these study populations.

Gender aspects of survival after surgical treatment for rectal cancer.

AIM: Longer survival in women than men after rectal cancer surgery has been reported. Our hypothesis was that after correction for their longer life expectancy a survival benefit for women would still remain. METHOD: We studied 2792 patients diagnosed with rectal cancer in the southern part of Sweden between 1996 and 2006. The following parameters were included in a prespecified multivariable Cox regression analysis: age at diagnosis, gender, preoperative radiotherapy, stage, year and type of surgery. In addition to overall survival, relative survival was calculated using the Hakulinen approach utilizing an age-, gender- and calendar year-matched Swedish control cohort. RESULTS: Female patients were significantly older, received neoadjuvant treatment less often and were more often operated on by local excision. Overall survival was significantly longer in women. In the multivariable analysis of relative survival, controlling for neoadjuvant treatment, Dukes stage and year and type of surgery, no significant effect of gender [hazard ratio (HR) 1.10 for men, P = 0.114] was found, whereas an improved relative survival with increased age (HR 0.96 per year, P < 0.001) was seen. In contrast, using the same multivariable model with no correction for underlying mortality in the population, male gender (HR 1.38, P < 0.001) and greater age (HR 1.05 per year, P < 0.001) increased the risk of death. CONCLUSION: The results show that after correction for the underlying longer survival in women and some known confounders, survival after surgical treatment for rectal cancer appears to be gender neutral.

Are 25 SNPs from the CARDIoGRAM study associated with ischaemic stroke?

BACKGROUND AND PURPOSE: The Coronary Artery Disease Genome-Wide Replication and Meta-Analysis Study (CARDIoGRAM) reported 25 single-nucleotide polymorphisms (SNPs) on 15 chromosomes to be associated with coronary artery disease (CAD) risk. Because common vascular risk factors are shared between CAD and ischaemic stroke (IS), these SNPs may also be related to IS overall or one or more of its pathogenetic subtypes. METHODS: We performed a candidate gene study comprising 3986 patients with IS and 2459 control subjects. The 25 CAD-associated SNPs reported by CARDIoGRAM were examined by allelic association analysis including logistic regression. Weighted and unweighted genetic risk scores (GRSs) were also compiled and likewise analysed against IS. We furthermore considered the IS main subtypes large-vessel disease (LVD), small-vessel disease and cardioembolic stroke [according to Trial of Org 10172 in Acute Stroke Treatment (TOAST)] separately. RESULTS: SNP rs4977574 on chromosome 9p21.3 was associated with overall IS [odds ratio (OR) = 1.12; 95% confidence interval (CI): 1.04-1.20; P = 0.002] as well as LVD (OR = 1.36; 95% CI: 1.13-1.64; P = 0.001). No other SNP was significantly associated with IS or any of its main subtypes. Analogously, the GRSs did not show any noticeable effect. CONCLUSIONS: Besides the previously reported association with SNPs on chromosome 9p21, this study did not detect any significant association between IS and CAD-susceptible genetic variants. Also, GRSs compiled from these variants did not predict IS or any pathogenetic IS subtype, despite a total sample size of 6445 participants.

Mutations of the Down-regulated in adenoma (DRA) gene cause congenital chloride diarrhoea.

A major transport function of the human intestine involves the absorption of chloride in exchange for bicarbonate. We have studied a recessively inherited defect of this exchange, congenital chloride diarrhoea (CLD; MIM 214700). The clinical presentation of CLD is a lifetime, potentially fatal diarrhoea with a high chloride content. The CLD locus was previously mapped to 7q3 adjacent to the cystic fibrosis gene (CFTR). By refined genetic and physical mapping, a cloned gene having anion transport function, Down-regulated in adenoma (DRA), was implicated as a positional and functional candidate for CLD. In this study, we report segregation of two missense mutations, delta V317 and H124L, and one frameshift mutation, 344delT, of DRA in 32 Finnish and four Polish CLD patients. The disease-causing nature of delta V317 is supported by genetic data in relation to the population history of Finland. By mRNA in situ hybridization, we demonstrate that the expression of DRA occurs preferentially in highly differentiated colonic epithelial cells, is unchanged in Finnish CLD patients with delta V317, and is low in undifferentiated (including neoplastic) cells. We conclude that DRA is an intestinal anion transport molecule that causes chloride diarrhoea when mutated.

Clinical implementation of systematic medication reconciliation and review as part of the Lund Integrated Medicines Management model--impact on all-cause emergency department revisits.

WHAT IS KNOWN AND OBJECTIVE: Interventions involving medication reconciliation and review by clinical pharmacists can reduce drug-related problems and improve therapeutic outcomes. The objective of this study was to examine the impact of routine admission medication reconciliation and inpatient medication review on emergency department (ED) revisits after discharge. Secondary outcomes included the combined rate of post-discharge hospital revisits or death. METHODS: This prospective, controlled study included all patients hospitalized in three internal medicine wards in a university hospital, between 1 January 2006 and 31 May 2008. Medication reconciliation on admission and inpatient medication review, conducted by clinical pharmacists in a multiprofessional team, were implemented in these wards at different times during 2007 and 2008 (intervention periods). A discharge medication reconciliation was undertaken in all the study wards, during both control and intervention periods. Patients were included in the intervention group (n = 1216) if they attended a ward with medication reconciliation and review, whether they had received the intervention or not. Control patients (n = 2758) attended the wards before implementation of the intervention. RESULTS AND DISCUSSION: No impact of medication reconciliation and reviews on ED revisits [hazard ratio (HR), 0.95; 95% confidence interval (CI), 0.86-1.04]or event-free survival (HR, 0.96; 95% CI, 0.88-1.04) was demonstrated. In the intervention group, 594 patients (48.8%) visited the ED, compared with 1416 (51.3%) control patients. In total, 716 intervention (58.9%) and 1688 (61.2%) control patients experienced any event (ED visit, hospitalization or death). Because the time to a subsequent ED visit was longer for the control as well as the intervention groups in 2007 than in 2006 (P < 0.05), we re-examined this cohort of patients; the proportion of patients revisiting the ED was similar in both groups in 2007 (P = 0.608). WHAT IS NEW AND CONCLUSION: Routine implementation of medication reconciliation and reviews on admission and during the hospital stay did not appear to have any impact on ED revisits, re-hospitalizations or mortality over 6-month follow-up.

Time trends in risk and risk determinants of non-Hodgkin lymphoma in solid organ transplant recipients.

Organ transplantation increases risk of non-Hodgkin lymphoma (NHL), but long-term risk and time trends have seldom been evaluated. Immunosuppressive drug load is an important risk determinant, but the details are unclear. We studied NHL risk in a nationwide Swedish cohort of 11 081 graft recipients transplanted 1970-2008. Relative risks (RRs) were estimated within the cohort and versus the general population by age, sex, follow-up time and calendar period. NHL risk was also assessed by cumulative and average doses of immunosuppressive treatments in a nested case-control design throughout 1997 using conditional logistic regression. We observed 153 NHL cases during 97 853 years of follow-up. Compared with the general population, NHL risk was eightfold increased (RR 7.9; 95% confidence interval [CI] 6.6-9.4), and increased risks persisted after ≥15 years of follow-up among kidney (6.1; 95% CI 3.5-10) and nonkidney recipients (44; 14-103). Among nonkidney recipients, NHL risk was lower in the 2000s compared with the 1990s (0.5; 95% CI 0.3-1.0; p = 0.04). A high average dose of antithymocyte immunoglobulin (ATG) conferred an eightfold increased risk of NHL (OR 8.5; 95% CI 1.9-38). To conclude, posttransplant NHL risk decreased during the last decade among nonkidney recipients, possibly because of a more careful use of ATG, the introduction of new drugs, or both.

Incidence of malignancy in patients treated for antineutrophil cytoplasm antibody-associated vasculitis: follow-up data from European Vasculitis Study Group clinical trials.

OBJECTIVES: Because standard immunosuppressive treatment for antineutrophil cytoplasm antibody-associated vasculitis (AAV) (granulomatosis with polyangiitis (Wegener's) (GPA) and microscopic polyangiitis (MPA)) has been associated with a significant risk of developing cancer, the cancer incidence of treated AAV patients was assessed. METHODS: This analysis concerned 535 patients with newly diagnosed AAV from 15 countries who had been enrolled between 1995 and 2002 in four European clinical trials. Over the period 2004-7, study participants' follow-up events were updated, including cancers diagnosed. Age, sex and area-standardised incidence ratios (SIR) and their 95% CI were calculated by linkage to five national cancer databases. RESULTS: During the 2650 person-years' observation period, 50 cancers were diagnosed in 46 patients. SIR (95% CI) were 1.58 (1.17 to 2.08) for cancers at all sites, 1.30 (0.90 to 1.80) for cancers at all sites excluding non-melanoma skin cancer (NMSC), 2.41 (0.66 to 6.17) for bladder cancer, 3.23 (0.39 to 11.65) for leukaemia, 1.11 (0.03 to 6.19) for lymphoma and 2.78 (1.56 to 4.59) for NMSC. Subgroup SIR for cancers at all sites were 1.92 (1.31 to 2.71) for GPA and 1.20 (0.71 to 1.89) for MPA. CONCLUSIONS: Cancer rates for AAV patients treated with conventional immunosuppressive therapy exceeded those expected for the general population. This cancer excess was largely driven by an increased incidence of NMSC. The smaller cancer risk magnitude in this cohort, compared with previous studies, might reflect less extensive use of cyclophosphamide in current treatment protocols. Longer follow-up data are warranted to appraise the risk of developing cancers later during the course of AAV.

The CD8+ T cell repertoire in beta 2-microglobulin-deficient mice is biased towards reactivity against self-major histocompatibility class I.

Beta 2-Microglobulin-deficient (beta 2m -/-) mice are reported to lack cell surface expression of major histocompatibility complex (MHC) class I molecules, CD8+ T cells, and the ability to mount MHC class I-specific T cell responses. We have observed that beta 2m -/- mice possess CD8+ T cells that can be induced to perform strong allospecific cytotoxic responses against nonself-MHC class I by in vivo priming. We report that these beta 2m -/- cytotoxic T lymphocyte (CTL) differ from those induced in beta 2m-positive littermates in that they cross-react and kill cells expressing self-MHC class I at normal ligand density with beta 2m. beta 2m -/- CTL could even be induced in primary mixed lymphocyte culture by self-MHC class I expressing stimulator cells, whereas allogeneic stimulator cells failed to elicit a response under similar conditions. Cells with a reduced cell surface MHC class I expression were less sensitive, while syngeneic beta 2m -/- cells were resistant to the beta 2m -/- CTL. This antiself-MHC reactivity could not be induced when beta 2m -/- T cells matured in an environment with normal MHC class I expression in bone marrow chimeric mice. Antiself-MHC reactivity was also observed against human peptide loading-deficient cells expressing the appropriate murine class I molecules, suggesting that affinity to self-MHC class I may occur irrespective of peptide content. The results fit with a model where positive and negative selection of CD8+ T cells in beta 2m -/- mice is mediated by low levels of MHC class I free heavy chains. In this model, low ligand density on selecting cells leads to positive selection of rare T cells that bind to low levels of MHC class I free heavy chains, resulting in a very small peripheral CD8+ compartment. Due to low density of the selecting ligand, negative selection does not remove T cells recognizing beta 2m-positive cells expressing self-MHC class I at normal ligand density, which generates a T cell repertoire that would be autoreactive in a beta 2m-positive littermate. The first "MHC deficient" animals thus paradoxically provide a tool for direct demonstration and analysis of self MHC bias in the T cell repertoire.

Alteration of the natural killer repertoire in H-2 transgenic mice: specificity of rapid lymphoma cell clearance determined by the H-2 phenotype of the target.

The mechanism behind natural tumor resistance conveyed by a H-2Dd transgene to C57Bl/6 (B6) mice was investigated. Transgenic D8 mice were more efficient than control mice in natural killer (NK) cell mediated rapid elimination of intravenously inoculated radiolabeled lymphoma cells of B6 origin, such as RBL-5. There was no difference between D8 and B6 mice when elimination of YAC-1 targets was monitored. The effect of the transgene on the NK repertoire was related to the H-2 phenotype of the target: the differential elimination of RBL-5 lymphoma cells in D8 and B6 mice was not seen when a H-2 deficient variant of this line was used (efficiently eliminated in both genotypes), nor was it seen with a H-2Dd transfectant (surviving in both genotypes). The data show that a MHC class I transgene can directly control natural killing in vivo by altering the repertoire rather than the general levels of NK activity. Since the NK mediated elimination seen after introduction of a novel gene in the host was neutralized by introducing the same gene (H-2Dd), but not an unrelated class I gene (H-2Dp), in the tumor, the data support the concept of NK surveillance against missing self. This combined transgenic/transfectant system may serve as a tool for a molecular dissection of the interactions between NK cells and their targets in vivo.

Natural killer cell tolerance in mice with mosaic expression of major histocompatibility complex class I transgene.

We have studied natural killer (NK) cell tolerance in a major histocompatibility complex (MHC) class I transgenic line, DL6, in which the transgene product was expressed on only a fraction of blood cells. In contrast with transgenic mice expressing the same transgene in all cells, NK cells from mosaic mice failed to reject transgene-negative bone marrow or lymphoma grafts. However, they retained the capability to reject cells with a total missing-self phenotype, i.e., cells lacking also wild-type MHC class I molecules. Tolerance against transgene-negative cells was demonstrated also in vitro, and could be broken if transgene-positive spleen cells of mosaic mice were separated from negative cells before, or after 4 d of culture in interleukin-2. The results provide support for selective NK cell tolerance to one particular missing-self phenotype but not to another. We suggest that this tolerance is determined by NK cell interactions with multiple cells in the environment, and that it is dominantly controlled by the presence of cells lacking a specific MHC class I ligand. Furthermore, the tolerant NK cells could be reactivated in vitro, which suggests that the tolerance occurs without deletion of the potentially autoreactive NK cell subset(s), and that it may be dependent upon the continuous presence of tolerizing cells.

Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) regulates the unfolding of autoimmune diabetes.

Evidence has been accumulating that shows that insulin-dependent diabetes is subject to immunoregulation. To determine whether cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) is involved, we injected anti-CTLA-4 mAb into a TCR transgenic model of diabetes at different stages of disease. When injected into young mice, months before they would normally become diabetic, anti-CTLA-4 induced diabetes rapidly and essentially universally; this was not the result of a global activation of T lymphocytes, but did reflect a much more aggressive T cell infiltrate in the pancreatic islets. These effects were only observed if anti-CTLA-4 was injected during a narrow time window, before the initiation of insulitis. Thus, engagement of CTLA-4 at the time when potentially diabetogenic T cells are first activated is a pivotal event; if engagement is permitted, invasion of the islets occurs, but remains quite innocuous for months, if not, insulitis is much more aggressive, and diabetes quickly ensues.

Initiation of autoimmune diabetes by developmentally regulated presentation of islet cell antigens in the pancreatic lymph nodes.

Little is known about the events triggering lymphocyte invasion of the pancreatic islets in prelude to autoimmune diabetes. For example, where islet-reactive T cells first encounter antigen has not been identified. We addressed this issue using BDC2.5 T cell receptor transgenic mice, which express a receptor recognizing a natural islet beta cell antigen. In BDC2.5 animals, activated T cells were found only in the islets and the lymph nodes draining them, and there was a close temporal correlation between lymph node T cell activation and islet infiltration. When naive BDC2.5 T cells were transferred into nontransgenic recipients, proliferating cells were observed only in pancreatic lymph nodes, and this occurred significantly before insulitis was detectable. Surprisingly, proliferation was not seen in 10-day-old recipients. This age-dependent dichotomy was reproduced in a second transfer system based on an unrelated antigen artificially expressed on beta cells. We conclude that beta cell antigens are transported specifically to pancreatic lymph nodes, where they trigger reactive T cells to invade the islets. Systemic or extrapancreatic T cell priming, indicative of activation via molecular mimicry or superantigens, was not seen. Compromised presentation of beta cell antigens in the pancreatic lymph nodes of juvenile animals may be the root of a first "checkpoint" in diabetes progression.

Natural resistance against lymphoma grafts conveyed by H-2Dd transgene to C57BL mice.

The H-2Dd transgenic strain D8 on C57BL background was more resistant to subcutaneous challenge of RBL-5 lymphoma cells than B6 controls. The direct role of the H-2Dd antigen was investigated by the use of (D8 x B6)F1 crosses and (D8 B6) x B6 backcrosses. The latter showed cosegregation with regard to Dd antigen expression and lymphoma resistance, both of which were inherited in a pattern consistent with control by a single dominant gene. The rejection potential in (D8 x B6)F1 mice appeared as strong as that seen in crosses between B6 and MHC congenic mice (on B10 background) carrying H-2Dd. The lymphoma resistance could be abrogated by treatment with anti-asialo GM1 antiserum or anti-NK 1.1 mAb, indicating a role for NK cells.

Acquisition of external major histocompatibility complex class I molecules by natural killer cells expressing inhibitory Ly49 receptors.

Murine natural killer (NK) cells express inhibitory Ly49 receptors specific for major histocompatibility complex (MHC) class I molecules. We report that during interactions with cells in the environment, NK cells acquired MHC class I ligands from surrounding cells in a Ly49-specific fashion and displayed them at the cell surface. Ligand acquisition sometimes reached 20% of the MHC class I expression on surrounding cells, involved transfer of the entire MHC class I protein to the NK cell, and was independent of whether or not the NK cell expressed the MHC class I ligand itself. We also present indirect evidence for spontaneous MHC class I acquisition in vivo, as well as describe an in vitro coculture system with transfected cells in which the same phenomenon occurred. Functional studies in the latter model showed that uptake of H-2D(d) by Ly49A+ NK cells was accompanied by a partial inactivation of cytotoxic activity in the NK cell, as tested against H-2D(d)-negative target cells. In addition, ligand acquisition did not abrogate the ability of Ly49A+ NK cells to receive inhibitory signals from external H-2D(d) molecules. This study is the first to describe ligand acquisition by NK cells, which parallels recently described phenomena in T and B cells.

Prevention of allogeneic bone marrow graft rejection by H-2 transgene in donor mice.

Rejection of bone marrow grafts in irradiated mice is mediated by natural killer (NK) cells and is controlled by genes linked to the major histocompatibility complex (MHC). It has, however, not been possible to identify the genes or their products. An MHC class I (Dd) transgene introduced in C57BL donors prevented the rejection of their bone marrow by NK cells in irradiated allogeneic and F1 hybrid mice expressing the Dd gene. Conversely, H-2Dd transgenic C57BL recipients acquired the ability to reject bone marrow from C57BL donors but not from H-2Dd transgenic C57BL donors. These results provide formal evidence that NK cells are part of a system capable of rejecting cells because they lack normal genes of the host type, in contrast to T cells, which recognize cells that contain abnormal or novel sequences of non-host type.