Metabolic syndrome in adults with autistic traits: associated psychological, behavioral, and biological factors in females and males - a PharmLines initiative.

Background: While cardiovascular diseases is highly prevalent and an important cause of mortality in autistic adults, knowledge on their increased cardiovascular risk is limited. Hence, this study aimed to investigate psychological, behavioral, and physical factors associated with metabolic syndrome (MetS) in adults with autistic traits. Methods: In total, 17,705 adults from the Lifelines Cohort were included and categorized using Autism Spectrum Quotient-10 sum-scores. The quartiles with highest (HQ-traits-group females: n = 2,635; males: n = 1803) and lowest levels of autistic traits (LQ-traits-group, n = idem) were analyzed. Using multivariable logistic regression, the associations between MetS and (self-reported and interviewed) psychological, behavioral, and physically measured factors in these stratified groups were investigated. Results: Among females, MetS was more common in the HQ-traits-group than in the LQ-traits-group (10.0% versus 7.5%, p < 0.01), while this was not the case among males (HQ-traits-group 13.8% versus LQ-traits-group 13.1%, p = 0.52). In both the female and male HQ-traits-group, the presence of MetS was associated with poorer self-reported health, less daily physical activity, and altered leukocyte counts. Conclusion: These findings underline the relevance of adequate cardiovascular prevention in adults with higher levels of autistic traits. Future research could gain more insight into the relationship between cardiovascular risk and autistic traits in females, and into tailored cardiovascular prevention.

Breakfast partly restores the anti-inflammatory function of high-density lipoproteins from patients with type 2 diabetes mellitus.

Background and aims: High-density lipoproteins (HDL) of patients with type 2 diabetes mellitus (T2DM) have impaired anti-inflammatory activities. The anti-inflammatory activity of HDL has been determined ex vivo after isolation by different methods from blood mostly obtained after overnight fasting. We first determined the effect of the HDL isolation method, and subsequently the effect of food intake on the anti-inflammatory function of HDL from T2DM patients. Methods: Blood was collected from healthy controls and T2DM patients after an overnight fast, and from T2DM patients 3 h after breakfast (n = 17 each). HDL was isolated by a two-step density gradient ultracentrifugation in iodixanol (HDLDGUC2), by sequential salt density flotation (HDLSEQ) or by PEG precipitation (HDLPEG). The anti-inflammatory function of HDL was determined by the reduction of the TNFα-induced expression of VCAM-1 in human coronary artery endothelial cells (HCAEC) and retinal endothelial cells (REC). Results: HDL isolated by the three different methods from healthy controls inhibited TNFα-induced VCAM-1 expression in HCAEC. With apoA-I at 0.7 µM, HDLDGUC2 and HDLSEQ were similarly effective (16% versus 14% reduction; n = 3; p > 0.05) but less effective than HDLPEG (28%, p < 0.05). Since ultracentrifugation removes most of the unbound plasma proteins, we used HDLDGUC2 for further experiments. With apoA-I at 3.2 µM, HDL from fasting healthy controls and T2DM patients reduced TNFα-induced VCAM-1 expression in HCAEC by 58 ± 13% and 51 ± 20%, respectively (p = 0.35), and in REC by 42 ± 13% and 25 ± 18%, respectively (p < 0.05). Compared to preprandial HDL, postprandial HDL from T2DM patients reduced VCAM-1 expression by 56 ± 16% (paired test: p < 0.001) in HCAEC and by 34 ± 13% (paired test: p < 0.05) in REC. Conclusions: The ex vivo anti-inflammatory activity of HDL is affected by the HDL isolation method. Two-step ultracentrifugation in an iodixanol gradient is a suitable method for HDL isolation when testing HDL anti-inflammatory function. The anti-inflammatory activity of HDL from overnight fasted T2DM patients is significantly impaired in REC but not in HCAEC. The anti-inflammatory function of HDL is partly restored by food intake.

Obesity-associated T-cell and macrophage activation improve partly after a lifestyle intervention.

BACKGROUND: The relation between low-grade inflammation and metabolic dysfunction in obesity is not fully explored. OBJECTIVE: To evaluate immune parameters in the obese state and after a lifestyle intervention program. METHODS: Patients with obesity (n = 87) from an academic obesity clinic were compared with controls with regard to macrophage and T-cell activation (reflected by serum levels of soluble CD163 (sCD163) and soluble IL-2 receptor (sIL-2R), respectively), and an array of cytokines, chemokines, and growth factors. In addition, these parameters and regulatory T-cells (Treg), were studied in 27 patients who followed a 75-week lifestyle intervention (dietary advice, exercise, and psychoeducation). RESULTS: Mean sIL-2R and sCD163 levels were higher in patients than controls (sIL-2R:2884 ± 936 pg/ml vs. 2207 ± 813 pg/ml, p = 0.001; sCD163:1279 ± 580 pg/ml vs. 661 ± 271 pg/ml, p < 0.0001 respectively). Patients with metabolic syndrome (MetS) had higher sCD163 than those without (1467 ± 656 pg/ml vs. 1103 ± 438 pg/ml). Patients had higher IL-1ß, IL-1RA, IL-2, IL-4, IL-5, IL-6, IL-8, IL-9, IL-10, IL-15, IL-17A, MCP-1/CCL2, MIP-1α/CCL3, MIP-1ß/CCL4, G-CSF, GM-CSF, FGF, IFN-γ, and TNF-α than controls, whereas VEGF-A, PDGF-BB, and eotaxin were lower. Upon intervention, sIL-2R decreased while peripheral Treg frequencies increased within the reference range (p = 0.042 and p = 0.005 respectively). The sIL-2R decrease correlated to a decrease in waist circumference (rho = 0.388, p = 0.045) and in trend to a decrease in MetS components (rho = 0.345, p = 0.078). The Treg increase was unrelated to weight loss or metabolic improvement. Mean sCD163 did not change significantly upon intervention, nor did the cytokines, chemokines, and growth factors (except IP-10/CXCL10). CONCLUSION: In obesity, T-cell homeostasis improves after a lifestyle intervention. Immunologic alterations can occur independently of metabolic improvement.

CECR1-mediated cross talk between macrophages and vascular mural cells promotes neovascularization in malignant glioma.

Glioblastomas (glioblastoma multiforme, GBM) are most malignant brain tumors characterized by profound vascularization. The activation of macrophages strongly contributes to tumor angiogenesis during GBM development. Previously, we showed that extracellular adenosine deaminase protein Cat Eye Syndrome Critical Region Protein 1 (CECR1) is highly expressed by M2-like macrophages in GBM where it defines macrophage M2 polarization and contributes to tumor expansion. In this study, the effect of CECR1 in macrophages on tumor angiogenesis was investigated. Immunohistochemical evaluation of GBM tissue samples showed that the expression of CECR1 correlates with microvascular density in the tumors, confirming data from the TCGA set. In a three-dimensional co-culture system consisting of human pericytes, human umbilical vein endothelial cells and THP1-derived macrophages, CECR1 knockdown by siRNA and CECR1 stimulation of macrophages inhibited and promoted new vessel formation, respectively. Loss and gain of function studies demonstrated that PDGFB mRNA and protein levels in macrophages are modulated by CECR1. The proangiogenic properties of CECR1 in macrophages were partially mediated via paracrine activation of pericytes by PDGFB-PDGFRß signaling. CECR1-PDGFB-PDGFRß cross-activation between macrophages and pericytes promoted pericyte migration, shown by transwell migration assay, and enhanced expression and deposition of periostin, a matrix component with proangiogenic properties. CECR1 function in (M2-like) macrophages mediates cross talk between macrophages and pericytes in GBM via paracrine PDGFB-PDGFRß signaling, promoting pericyte recruitment and migration, and tumor angiogenesis. Therefore, CECR1 offers a new portent target for anti-angiogenic therapy in GBM via immune modulation.

Interaction of mouse splenocytes and macrophages with bacterial strains in vitro: the effect of age in the immune response.

Probiotics influence the immune system, both at the local and systemic level. Recent findings suggest the relation between microbiota and the immune system alters with age. Our objective was to address direct effects of six bacterial strains on immune cells from young and aged mice: Lactobacillus plantarum WCFS1, Lactobacillus casei BL23, Lactococcus lactis MG1363, Bifidobacterium breve ATCC15700, Bifidobacterium infantis ATCC15697, and Akkermansia muciniphila ATCC BAA-835. We used splenocytes and naïve or interferon-γ-stimulated bone marrow-derived macrophages (BMDM) as responder populations. All tested bacterial strains induced phenotypic and cytokine responses in splenocytes and BMDM. Based on magnitude of the cellular inflammatory response and cytokine profiles, two subgroups of bacteria were identified, i.e. L. plantarum and L. casei versus B. breve, B. infantis, and A. muciniphila. The latter group of bacteria induced high levels of cytokines produced under inflammatory conditions, including tumour necrosis factor (TNF), interleukin (IL)-6 and IL-10. Responses to L. lactis showed features of both subgroups. In addition, we compared responses by splenocytes and BMDM derived from young mice to those of aged mice, and found that splenocytes and BMDM derived from aged mice had an increased IL-10 production and dysregulated IL-6 and TNF production compared to young immune cells. Overall, our study shows differential inflammatory responses to distinct bacterial strains, and profound age-dependent effects. These findings, moreover, support the view that immune environment importantly influences bacterial immune effects.

Angiogenic murine endothelial progenitor cells are derived from a myeloid bone marrow fraction and can be identified by endothelial NO synthase expression.

OBJECTIVE: Endothelial progenitor cells (EPCs) contribute to postnatal neovascularization and are therefore of great interest for autologous cell therapies to treat ischemic vascular disease. However, the origin and functional properties of these EPCs are still in debate. METHODS AND RESULTS: Here, ex vivo expanded murine EPCs were characterized in terms of phenotype, lineage potential, differentiation from bone marrow (BM) precursors, and their functional properties using endothelial NO synthase (eNOS)-green fluorescent protein transgenic mice. Despite high phenotypic overlap with macrophages and dendritic cells, EPCs displayed unique eNOS expression, endothelial lineage potential in colony assays, and angiogenic characteristics, but also immunologic properties such as interleukin-12p70 production and low levels of T-cell stimulation. The majority of EPCs developed from an immature, CD31(+)Ly6C+ myeloid progenitor fraction in the BM. Addition of myeloid growth factors such as macrophage-colony-stimulating factor (M-CSF) and granulocyte/macrophage (GM)-CSF stimulated the expansion of spleen-derived EPCs but not BM-derived EPCs. CONCLUSIONS: The close relationship between EPCs and other myeloid lineages may add to the complexity of using them in cell therapy. Our mouse model could be a highly useful tool to characterize EPCs functionally and phenotypically, to explore the origin and optimize the isolation of EPC fractions for therapeutic neovascularization.

Macrophages in the murine pancreas and their involvement in fetal endocrine development in vitro.

Macrophages are a heterogeneous population of cells that belong to the mononuclear phagocyte system. They play an important role in tissue homeostasis and remodeling and are also potent immune regulators. Pancreatic macrophages are critically involved in the development and pathogenesis of autoimmune diabetes. To elucidate the ontogeny of pancreatic macrophages, we characterized in this study the macrophages present in the adult and developing fetal pancreas of normal mice. We additionally examined the presence of local macrophage precursors and the involvement of macrophages in the growth of endocrine tissue in the fetal pancreas. We identified two phenotypically distinct macrophage subsets in the adult pancreas. The majority of macrophages was CD45(+)ER-MP23(+)MOMA-1(+). Under noninflammatory conditions, only a minority ( approximately 5%) of the pancreatic macrophages additionally expressed the macrophage marker F4/80. In contrast, in the fetal pancreas, phenotypically, mature macrophages were identified exclusively by their expression of F4/80 and lacked detectable staining with ER-MP23 and MOMA-1 antibodies. In fetal pancreas organ cultures, we could show that macrophages develop from pre-existing precursors, which are present in the fetal pancreas at embryonic age 12.5. Moreover, the number of macrophages increased significantly when macrophage-colony stimulating factor was added to these cultures. It is important that this increase of F4/80-positive cells was paralleled by an increase in the number of insulin-producing cells, suggesting that macrophages support the growth of these endocrine cells.

Cellular composition of pancreas-associated lymphoid tissue during human fetal pancreatic development.

AIM: To study human fetal pancreatic tissue between 15 weeks of gestation and term, analysing the development of pancreatic lymphoid tissue and focusing on the presence and maturational status of dendritic cells (DCs). During normal human fetal pancreatic development lymphoid tissue arises in and around the pancreas. DCs are antigen-presenting cells which are capable of initiating immunity, but are also essential in inducing and maintaining T-cell tolerance. METHODS AND RESULTS: First, the presence and general composition of intra- and peripancreatic lymphoid tissue was investigated by histology and immunohistochemistry with antibodies to CD3, CD4, CD8, CD14, CD20, CD68, and CD79. Intrapancreatic lymphoid tissue (IPLT) appeared to be present only from 29 weeks of gestation onwards, and had a similar composition to peripancreatic lymphoid tissue (PPLT), which was found in all 23 specimens examined. Both forms of lymphoid tissue had an architecture similar to lymph nodes, with separate B- and T-lymphocyte areas and scattered macrophages. DCs were investigated in detail by immunohistochemistry for CD1a, CD83, CD86, CD123, Langerin, and DC-LAMP. Both Langerin, a marker for immature DCs, as well as DC-LAMP, a marker for mature DCs, were expressed by cells in both the IPLT and PPLT at all ages examined. CONCLUSION: The presence of DCs at all developmental stages, expressing various maturation-related markers, in addition to the general composition of the human fetal PPLT and IPLT suggests that this is fully functional and has a function comparable to peripheral lymph nodes.

Immature macrophages derived from mouse bone marrow produce large amounts of IL-12p40 after LPS stimulation.

Production of IL-12 is an important indicator of the macrophage's ability to regulate immune responses. In this study, we investigated the IL-12 production by macrophages in different developmental stages. To this end, macrophages were generated in vitro from precursors stimulated with M-CSF, GM-CSF or IL-3. Density separation yielded populations enriched in different maturation stages. Invariably, only cells banding at the 40-50% Percoll interface produced large amounts of IL-12p40 when stimulated with LPS, whereas only low levels of IL-12p70 were produced. These cells represented immature macrophages, as indicated by the absence of precursor markers CD31/ER-MP12, Ly-6C/ER-MP20 and ER-MP58, and by the low level of expression of mature-cell markers like ER-HR3, scavenger receptor and CD11b/Mac-1. Upon further maturation, the macrophages' ability to produce IL-12p40 decreased, coinciding with increased nitric oxide production upon LPS stimulation. These results show that immature macrophages produce high levels of IL-12p40 and thus may either contribute to IL-12p70 production or regulate it.

Abnormalities in dendritic cell and macrophage accumulation in the pancreas of nonobese diabetic (NOD) mice during the early neonatal period.

Dendritic cell (DC), macrophage (Mphi) and lymphocyte infiltrations have been observed in normal human perinatal pancreata, but have never been investigated so early in control mice. In type 1 diabetes-prone NOD mice, these cells are thought to infiltrate first the periphery of the islets of Langerhans around weaning before further islet infiltration and beta-cell destruction. We quantified, during the first month of life, the numbers of DC (characterized by CD11c positivity and dendritic morphology), histiocyte-like Mphi (characterized by ER-MP23 positivity) and Mphi with scavenging potential (characterized by BM8 positivity) in C57BL/6, DBA/2 and BALB/c control, and NOD and lymphocyte-deficient NODscid mouse pancreata. First, CD11c+ DC were present at low densities from birth onwards in control pancreata, while densities were higher in NOD and NODscid. Second, high numbers of BM8+ and ER-MP23+ Mphi were observed at birth in all strains investigated. After birth, particularly BM8+ cells disappeared progressively in control strains, but not in NOD and NODscid. Third, NOD mice also had more ER-MP23+ Mphi at birth compared to controls. Finally, DC and Mphi localizations were similar in all strains, i.e., mostly as dispersed cells in perivascular, periductular, peri-islet areas and interlobular septa. The most remarkable finding was that particularly BM8+ Mphi, were seen at sites of islet neogenesis and predominantly at the duct-islet interface. Our data showed that different types of APC were present in the pancreas during postnatal development in various control mouse strains and some differences were observed in NOD and NODscid mice from birth onwards.

Cytokine production induced by binding and processing of calcium oxalate crystals in cultured macrophages.

Deposition of calcium oxalate (CaOx) crystals in the renal interstitium is common in humans with primary oxalosis and secondary hyperoxaluria, as well as in kidneys of rats with CaOx nephrolithiasis. In vivo, macrophages and multinucleated giant cells mostly encapsulate these crystals. To investigate whether macrophages are able to dispose of CaOx crystals after phagocytosis, we used a nontransformed macrophage cell line derived from mouse spleen progenitors. Cytokine assays showed that in response to crystal binding and phagocytosis, these macrophages release tumor necrosis factor-alpha. This release was evident at 8 hours, maximal at 24 hours, and decreased to control values after 48 hours of incubation with crystals. A very low but significant release of interleukin-6 into the culture medium was only noticed after 32 hours. Radiochemical experiments showed that these cells bind 38.8% of the CaOx crystals added. After 4 days, all internalized crystals had been dissolved and their molecular constituents released into the extracellular environment. Confocal laser scanning microscopy followed by morphometrical analyses confirmed these results. Long-term (survival) analyses showed that in the interval under study and at the crystal doses used, cell viability was not significantly affected. These findings support the view that properly functioning macrophages are able to remove CaOx deposits from the renal interstitium and that these cells produce inflammatory cytokines before crystal dissolution.

Differential ultraviolet-B-induced immunomodulation in XPA, XPC, and CSB DNA repair-deficient mice.

Ultraviolet B irradiation has serious consequences for cellular immunity and can suppress the rejection of skin tumors and the resistance to infectious diseases. DNA damage plays a crucial role in these immunomodulatory effects of ultraviolet B, as impaired repair of ultraviolet-B-induced DNA damage has been shown to cause suppression of cellular immunity. Ultraviolet-B-induced DNA damage is repaired by the nucleotide excision repair mechanism very efficiently. Nucleotide excision repair comprises two subpathways: transcription-coupled and global genome repair. In this study the immunologic consequences of specific nucleotide excision repair defects in three mouse models, XPA, XPC, and CSB mutant mice, were investigated. XPA mice carry a total nucleotide excision repair defect, whereas XPC and CSB mice only lack global genome and transcription-coupled nucleotide excision repair, respectively. Our data demonstrate that cellular immune parameters in XPA, XPC, and CSB mice are normal compared with their wild-type (control) littermates. This may indicate that the reported altered cellular responses in xeroderma pigmentosum patients are not constitutive but could be due to external factors, such as ultraviolet B. Upon exposure to ultraviolet B, only XPA mice are very sensitive to ultraviolet-B-induced inhibition of Th1-mediated contact hypersensitivity responses and interferon-gamma production in skin draining lymph nodes. Lipopolysaccharide-stimulated tumor necrosis factor alpha and interleukin-10 production are significantly augmented in both XPA and CSB mice after ultraviolet B exposure. Lymph node cell numbers were increased very significantly in XPA, mildly increased in CSB, and not in XPC mice. In general XPC mice do not exhibit any indication of enhanced ultraviolet B susceptibility with regard to the immune parameters analyzed. These data suggest that both global genome repair and transcription-coupled repair are needed to prevent immunomodulation by ultraviolet B, whereas transcription-coupled repair is the major DNA repair subpathway of nucleotide excision repair that prevents the acute ultraviolet-B-induced effects such as erythema.

Sex steroids influence pancreatic islet hypertrophy and subsequent autoimmune infiltration in nonobese diabetic (NOD) and NODscid mice.

Female nonobese diabetic (NOD) mice more frequently develop autoimmune diabetes than NOD males. Orchidectomy of the latter aggravates insulitis and diabetes. Because clear differences in immune function have not been observed between prediabetic females and males, before or after castration, we hypothesized that sex-related differences in diabetes incidence are related to target organ-specific actions of sex steroids. Previously, we showed that prediabetic NOD females develop hyperinsulinemia and subsequently mega-islets. Infiltration of the first inflammatory leukocytes is predominantly associated with these mega-islets. Here, we determined the relationship between sex hormones, mega-islet formation, and infiltrating cells in NOD and nonobese diabetic/severe combined immune-deficient (NODscid) mice. Mega-islet formation was reduced in NOD males compared with NOD females, and orchidectomy increased it, indicating a relationship between androgen levels and mega-islet formation. Moreover, enhanced mega-islet formation in castrated NOD males was associated with increased numbers of infiltrating leukocytes. Castrated NODscid males also exhibited increased mega-islet formation and dendritic cell infiltration, indicating that lymphocytes are not required for castration-induced effects. In conclusion, we show that androgens influence pancreatic islets and autoimmune infiltration in NOD and NODscid mice. This suggests that the gender difference in diabetes incidence in NOD mice is related to target organ-specific androgen effects.

Leukocyte-facilitated entry of intracellular pathogens into the central nervous system.

Microbes use numerous strategies to invade the central nervous system. Leukocyte-facilitated entry is one such mechanism whereby intracellular pathogens establish infection by taking advantage of leukocyte trafficking to the central nervous system. Key components of this process include peripheral infection and activation of leukocytes, activation of cerebral endothelial cells with or without concomitant infection, and trafficking of infected leukocytes to and through the blood-brain or blood-cerebrospinal fluid barrier.

UVB irradiation modulates systemic immune responses by affecting cytokine production of antigen-presenting cells.

The immunosuppressive effects of UVB irradiation have been well documented. The production of cytokines by keratinocytes is considered to play a major role in the induction of local as well as systemic immunosuppression. It is thought that partly due to the interaction of locally produced cytokines with antigen-presenting cells (APC) systemic effects, like antigen-specific tolerance, can be induced. In this study we examined the effect of UVB irradiation on cytokine profiles of peripheral APC as well as the functional consequences. Our results indicate that UVB irradiation impairs T(h)1-mediated immune responses in vivo by suppression of the systemic IL-12p70 production. Splenic APC from UVB-exposed mice showed an enhanced production of prostaglandin E(2), IL-1, IL-6 and tumor necrosis factor-alpha after in vitro stimulation. Also, spleen cells from UVB irradiated IL-4(-/-) mice showed increased IL-6 levels. These APC were less efficient in inducing IFN-gamma production by CD4(+) T cells and suppressed IgM production by B cells. We conclude that the altered cytokine profile of peripheral APC can be responsible for the systemic effects of UVB irradiation on the T(h)1/T(h)2 balance as well as on B cell responses.

Islet abnormalities associated with an early influx of dendritic cells and macrophages in NOD and NODscid mice.

In the nonobese diabetic (NOD) mouse model for type 1 diabetes, the inflammatory infiltration of islets starts with an influx of dendritic cells (DC) and macrophages (Mphi) at approximately 4 weeks of age. Around this time, NOD mice show endocrine abnormalities, indicated by a transient hyperinsulinemia that lasts until 8 weeks of age. Subsequently, they develop abnormally large islets of Langerhans, here designated as "mega-islets." NODscid mice, which lack functional lymphocytes, also exhibit transient hyperinsulinemia, but to a lesser extent. First, to determine the role of lymphocytes in the morphological islet abnormalities, we compared 6-week-old (prediabetic) NOD and NODscid females regarding mega-islet development and accumulation of antigen-presenting cells (APC), particularly CD11c+ DC and ERMP23+ Mphi. In NODscid mice, early APC infiltration and mega-islets were present, but less marked compared with NOD mice, thus suggesting a role of lymphocytes in mega-islet formation. In both NOD and NODscid mice, the APC infiltration was predominantly found around the mega-islets, suggesting a relationship between both parameters. Second, to analyze the role of beta-cell hyperactivity in mega-islet formation, we studied the effect of short-term prophylactic insulin treatment on these parameters. Prophylactic insulin treatment decreased the percentages of mega-islets in both NOD and NODscid mice, indicating that beta-cell hyperactivity is also involved in mega-islet formation. In conclusion, mega-islet formation in mice with the NOD genetic background takes place under the influence of both beta-cell hyperactivity and leukocytes.

Subsets of macrophages and dendritic cells in nonobese diabetic mouse pancreatic inflammatory infiltrates: correlation with the development of diabetes.

Islet-specific T cells are essential in the development of type I diabetes. The role of non-lymphoid cells is relatively unclear, although infiltration of dendritic cells and macrophages is the first sign of islet autoimmunity in diabetes-prone nonobese diabetic (NOD) mice. BDC2.5 is one of the autoreactive T cell clones isolated from NOD mice. Transfer of BDC2.5 T cells into young NOD mice accelerates diabetes development, whereas transgenic expression of the BDC2.5 T cell receptor on NOD T cells (BDC2.5 TCR-Tg NOD) markedly reduces diabetes development. We show that, although the same antigen-specificity is involved, both models differ significantly in insulitis. BDC2.5 TCR-Tg NOD mice develop an extensive, but non-aggressive, peri-insulitis by 3 weeks of age. In these large peri-islet infiltrates, resembling secondary lymphoid tissue, BM8+ macrophages (Mphi) are virtually absent. In contrast, BDC2.5 T cell clone transfer results in an aggressive insulitis with small infiltrates, but relatively large numbers of BM8 Mphi. Infiltration of BM8+ Mphi therefore correlates with islet destruction. This is, however, not observed for all Mphi; Monts-4+ Mphi follow a reverse pattern and are present in higher numbers in BDC2.5 TCR-Tg than in transferred mice. ER-MP23+ Mphi are reduced in both transferred and transgenic mice compared with wild-type NOD. Thus, this study underlines and extends previous data suggesting that Mphi are implicated in both early and late phases in diabetes development. Furthermore, our data imply that subsets of non-lymphoid cells have different roles in diabetes development. It is, therefore, important to recognize this heterogeneity when interpreting both in vivo and in vitro studies concerning non-lymphoid cells in diabetes.

Splenic dendritic cells from the non-obese diabetic mouse induce a prolonged proliferation of syngeneic T cells. A role for an impaired apoptosis of NOD T cells?

In this study we have tried to detect abnormalities in the immunophenotype and/or function of dendritic cells from the non-obese diabetic mouse (NOD DC), that might be related to islet autoimmunity. The immunophenotype of NOD splenic DC did not show significant abnormalities as compared with the immunophenotype of splenic DC from C57BL/10 mice. Furthermore, NOD splenic and lymph node DC stimulated proliferation of syngeneic T cells as efficiently as DC from C57BL/10 and BALB/c mice. The allogeneic response induced by NOD DC was similar to or only slightly lower than the response induced by C57BL/10 DC. Both a normal immunophenotype of NOD DC and efficient T cell stimulation were observed regardless of the stage of diabetes development. However, the syngeneic T cell proliferation induced by NOD splenic DC, but not by C57BL/10 splenic DC, was significantly prolonged, and it was accompanied by an increased proportion of activated/memory CD4(+)cells. We demonstrated that during the interaction of NOD cells fewer apoptotic cells were generated as compared with the interaction of C57BL/10 cells. Thus, the prolonged T cell response during the syngeneic interaction between NOD DC and T cells might be due to an impaired apoptosis induction. The impaired apoptosis might be of critical importance in the development of islet autoimmunity in the NOD mouse.

Surface interleukin-10 inhibits listericidal activity by primary macrophages.

Interleukin-10 (IL-10) down-regulates multiple functions of monocytes and macrophages, including the ability of macrophages to kill many intracellular microorganisms. The experiments presented here test the hypothesis that IL-10 expressed on the cell surface inhibits the ability of primary mouse macrophages to kill the facultative, intracellular bacterium Listeria monocytogenes. We show that, in contrast to macrophages from normal mice, both bone marrow-derived macrophages (BMDM) and thioglycollate-elicited macrophages obtained from IL-10-/- mice can kill L. monocytogenes. Treatment with anti-IL-10 monoclonal antibody (mAb) enables BMDM from normal mice and thioglycollate-elicited macrophages from RAG-2-/- mice (which lack T or B cell-derived IL-10) to kill L. monocytogenes, and concurrently down-regulates the expression of surface IL-10. Surface IL-10 on paraformaldehyde-fixed cells can inhibit nitric oxide (NO) production by interferon-gamma (IFN-gamma)-stimulated macrophages from IL-10-/- mice, thus directly showing functional activity of surface IL-10. Taken together, these studies indicate that macrophage surface IL-10 is biologically active and down-regulates macrophage bactericidal activity.