Metabolic dysfunction-associated steatohepatitis (MASH) is a common but frequently unrecognized complication of obesity and type 2 diabetes. The association between these conditions is multifaceted and involves complex interactions between metabolic, inflammatory, and genetic factors. Here we assess the underlying structural and molecular processes focusing on the immunological phase of MASH in the nonobese inflammation and fibrosis (NIF) mouse model and compare it to the human disease as well as other murine models. Histopathology together with synchrotron-radiation-based x-ray micro-computed tomography (SRµCT) was used to investigate structural changes within the hepatic sinusoids network in the NIF mouse in comparison to patients with different severities of MASH. A time-course, bulk RNA-sequencing analysis of liver tissue from NIF mice was performed to identify the dynamics of key processes associated with the pathogenesis. Transcriptomics profiling of the NIF mouse revealed a gradual transition from an initially reactive inflammatory response to a regenerative, pro-fibrotic inflammatory response suggesting new avenues for treatment strategies that focus on immunological targets. Despite the lack of metabolic stress induced liver phenotype, a large similarity between the NIF mouse and the immunological phase of human MASH was detected. The translational value was further supported by the comparative analyses with MASH patients and additional animal models. Finally, the impact of diets known to induce metabolic stress, was explored in the NIF mouse. An obesogenic diet was found to induce key physiological, metabolic, and histologic changes akin to those observed in human MASH.

Conclusion:In rats with biliary cirrhosis, dual alpha-v/beta-6 and alpha-v/beta-1 inhibition by PLN-75068 ameliorated portal hypertension, exerted dose-dependent anti-fibrotic effects as assessed by CPA and significantly reduced ALT levels.A multinational Phase 2a evaluation of PLN-74809 in participants with PSC is ongoing (INTEGRIS-PSC;

Sterile liver inflammation and fibrosis are associated with many liver disorders of different etiologies. Both type 1 and type 2 inflammatory responses have been reported to contribute to liver pathology. However, the mechanisms controlling the balance between these responses are largely unknown. Natural killer T (NKT) cells can be activated to rapidly secrete cytokines and chemokines associated with both type 1 and type 2 inflammatory responses. As these proteins have been reported to accumulate in different types of sterile liver inflammation, we hypothesized that these cells may play a role in this pathological process. We have found that a transgenic NKT (tgNKT) cell population produced in the immunodeficient 2,4αβNOD. Rag2 −/− mice, but not in 2,4αβNOD. Rag2 +/− control mice, promoted a type 1 inflammatory response with engagement of the NOD-, LRR- and pyrin domain-containing protein-3 (NLRP3) inflammasome. The induction of the type 1 inflammatory response was followed by an altered cytokine profile of the tgNKT cell population with a biased production of anti-inflammatory/profibrotic cytokines and development of liver fibrosis. These findings illustrate how the plasticity of NKT cells modulates the inflammatory response, suggesting a key role for the NKT cell population in the control of sterile liver inflammation.

Dementia is a devastating manifestation of Parkinson's disease (PD). This study investigates whether a common polymorphism in the PITX3 gene (rs2281983), which is of importance for the function of dopaminergic neurons, affects the risk of developing dementia in PD and whether it affects dopamine transporter (DAT) uptake. We PITX3 genotyped 133 patients with new-onset, idiopathic PD, participating in a population-based study in Sweden. Patients were followed prospectively during 6-11years with extensive investigations, including neuropsychology and DAT-imaging with 123I FP-CIT. The primary outcome was the incidence of PD dementia (PDD), diagnosed according to published criteria, studied by the Kaplan-Meier method and Cox proportional hazards. Performance in individual cognitive domains, the incidence of visual hallucinations, disease progression and striatal DAT uptake on imaging was also investigated. PD patients carrying the PITX3 C allele had an increased risk of developing PDD (hazard ratio: 2.87, 95% CI: 1.42-5.81, p=0.003), compared to the PD patients homozygous for the T-allele. Furthermore, the PITX3 C allele carriers with PD had a poorer cognitive performance in the visuospatial domain (p<0.001) and a higher incidence of visual hallucinations. A trend towards a lower striatal DAT uptake in the PITX3 C allele carriers was suggested, but could not be confirmed. Our results show that a common polymorphism in the PITX3 gene affects the risk of developing PDD and visuospatial dysfunction in idiopathic PD. If validated, these findings can provide new insights into the neurobiology and genetics of non-motor symptoms in PD.

Here we characterize a new animal model that spontaneously develops chronic inflammation and fibrosis in multiple organs, the non-obese diabetic inflammation and fibrosis (N-IF) mouse. In the liver, the N-IF mouse displays inflammation and fibrosis particularly evident around portal tracts and central veins and accompanied with evidence of abnormal intrahepatic bile ducts. The extensive cellular infiltration consists mainly of macrophages, granulocytes, particularly eosinophils, and mast cells. This inflammatory syndrome is mediated by a transgenic population of natural killer T cells (NKT) induced in an immunodeficient NOD genetic background. The disease is transferrable to immunodeficient recipients, while polyclonal T cells from unaffected syngeneic donors can inhibit the disease phenotype. Because of the fibrotic component, early on-set, spontaneous nature and reproducibility, this novel mouse model provides a unique tool to gain further insight into the underlying mechanisms mediating transformation of chronic inflammation into fibrosis and to evaluate intervention protocols for treating conditions of fibrotic disorders.

Background: T1D and AITD are autoimmune disorders commonly occurring in the same family and even in the same individual. The genetic contribution to these disorders is complex making uncovering of susceptibility genes very challenging. The general aim of this study was to identify loci and genes contributing to T1D/AITD susceptibility. Our strategy was to perform linkage and association studies in the relatively genetically homogenous population of northern Sweden. We performed a GWLS to find genomic regions linked to T1D/AITD in families from northern Sweden and we performed an association study in the families to test for association between T1D/ AITD and variants in previously published candidate genes as well as a novel candidate gene, CD247.Methods: DNA prepared from 459 individuals was used to perform a linkage and an association study. The ABI PRISM Linkage Mapping Set v2.5MD10 was employed for an initial 10-cM GWLS, and additional markers were added for fine mapping. Merlin was used for linkage calculations. For the association analysis, a GoldenGate Custom Panel from Illumina containing 79 SNPs of interest was used and FBAT was used for association calculations.Results: Our study revealed linkage to two previously identified chromosomal regions, 4q25 and 6p22, as well as to a novel chromosomal region, 1q23. The association study replicated association to PTPN22, HLA-DRB1, INS, IFIH1, CTLA4 and C12orf30. Evidence in favor of association was also found for SNPs in the novel susceptibility gene CD247.Conclusions: Several risk loci for T1D/AITD identified in published association studies were replicated in a family material, of modest size, from northern Sweden. This provides evidence that these loci confer disease susceptibility in this population and emphasizes that small to intermediate sized family studies in this population can be used in a cost-effective manner for the search of genes involved in complex diseases. The linkage study revealed a chromosomal region in which a novel T1D/AITD susceptibility gene, CD247, is located. The association study showed association between T1D/AITD and several variants in this gene. These results suggests that common susceptibility genes act in concert with variants of CD247 to generate genetic risk for T1D/AITD in this population.

Coreceptor CD4 and CD8αβ double-negative (DN) TCRαβ+ intraepithelial T cells, although numerous, have been greatly overlooked and their contribution to the immune response is not known. Here we used T cell receptor (TCR) sequencing of single cells combined with retrogenic expression of TCRs to study the fate and the major histocompatibility complex (MHC) restriction of DN TCRαβ+ intraepithelial T cells. The data show that commitment of thymic precursors to the DN TCRαβ+ lineage is imprinted by their TCR specificity. Moreover, the TCRs they express display a diverse and unusual pattern of MHC restriction that is nonoverlapping with that of CD4+ or CD8αβ+ T cells, indicating that they sense antigens that are not recognized by the conventional T cell subsets. The new insights indicate that DN TCRαβ+ T cells form a third lineage of TCRαβ T lymphocytes expressing a variable TCR repertoire, which serve nonredundant immune functions.

Autoimmune diseases result from a breakdown in self-tolerance, with a major contribution of pathogenic self-reactive T cells. Individuals with a particular autoimmune disease are often at risk of developing additional autoimmunity directed towards other target tissues. These observations suggest that there might be a mechanism leading to a general defect in self-tolerance, resulting in increased susceptibility to autoimmune disease. Tolerance towards self is initially established in the thymus during the process of thymic selection. Developing thymocytes are first immature CD4 and CD8 double negative thymocytes. Cells with a productive TCRβ gene rearrangement will express a pre-TCR/CD3 complex on the cell surface by combining a TCRβ chain with the pTα chain. Signaling through the pre-TCR, β-selection, plays a crucial role in the generation αβT cells, by enabling further maturation to the CD4 and CD8 double positive stage. The crucial role of pTα at β-selection together with the genetic localization of the ptcra gene and the fact that variations in pTα will affect all developing T cells, suggests that pTα could be a common factor leading to a general defect in self-tolerance. We have discovered differences in the isoform expression pattern and amino acid sequence of pTa between NOD and B6 mice. These findings are consistent with the hypothesis that alterations in the pre-TCR might affect selection, giving a modified TCR repertoire and increased susceptibility to autoimmunity.

T lymphocytes develop in the thymus into two major lineages characterized by expression of ab and gd TCR. Each lineage can be further subdivided into distinct subsets that differ in TCR specificity, phenotype and function. CD8aa TCRab intraepithelial lymphocytes (T-IEL) located in the epithelium of the small intestine is phenotypically different from conventional T cells. CD8aa T-IEL have been studied for years however the question of their MHC restriction has not been elucidated. Therefore, we decided to clone TCR isolated from naturally arising CD8aa T-IEL and retrovirally express these TCRs in BM chimera. First, we cloned four TCRab that were expressed retrovirally in RagKO BM chimera. Analysis of the chimera showed that all TCR clones gave rise to T cells. The T cell that developed were CD4-CD8b- but CD8a+ and phenotypically identical to CD8aa T-IEL isolated from wild type mice. In addition, they were preferentially found in the gut. In order to define the MHC restriction of these particular TCRs, BM chimeras in various MHC-deficient backgrounds were generated. Subsequent analysis of the chimera for the presence of CD8aaTCRab IEL demonstrated that they are dependent either on Kb or Db MHC I or b2m-dependent MHC I aside from KbDb. In conclusion, our results indicate that CD8aa T-IEL develops from precursors expressing particular TCRs that allow their engagement in this unique T cell lineage. In addition, CD8aa T-IEL can harbor clones selected on different MHC molecules.

Macrophages are multifunctional immune cells that may either drive or modulate disease pathogenesis depending on their activation phenotype. Autoimmune type 1 diabetes (T1D) is a chronic proinflammatory condition characterized by unresolved destruction of pancreatic islets. Adoptive cell transfer of macrophages with immunosuppressive properties represents a novel immunotherapy for treatment of such chronic autoimmune diseases. We used a panel of cytokines and other stimuli to discern the most effective regimen for in vitro induction of immunosuppressive macrophages (M2r) and determined interleukin (IL)-4/IL-10/transforming growth factor-β (TGF-β) to be optimal. M2r cells expressed programmed cell death 1 ligand-2, fragment crystallizable region γ receptor IIb, IL-10, and TGF-β, had a potent deactivating effect on proinflammatory lipopolysaccharide/interferon-γ–stimulated macrophages, and significantly suppressed T-cell proliferation. Clinical therapeutic efficacy was assessed after adoptive transfer in NOD T1D mice, and after a single transfer of M2r macrophages, >80% of treated NOD mice were protected against T1D for at least 3 months, even when transfer was conducted just prior to clinical onset. Fluorescent imaging analyses revealed that adoptively transferred M2r macrophages specifically homed to the inflamed pancreas, promoting β-cell survival. We suggest that M2r macrophage therapy represents a novel intervention that stops ongoing autoimmune T1D and may have relevance in a clinical setting.

Mature microRNAs (miRNAs), derived through cleavage of pre-miRNAs by the Dicer1 enzyme, regulate protein expression in many cell-types including cells in the pancreatic islets of Langerhans. To investigate the importance of miRNAs in mouse insulin secreting β-cells, we have generated mice with a β-cells specific disruption of the Dicer1 gene using the Cre-lox system controlled by the rat insulin promoter (RIP). In contrast to their normoglycaemic control littermates (RIP-Cre(+/-) Dicer1(Δ/wt)), RIP-Cre(+/-)Dicer1(flox/flox) mice (RIP-Cre Dicer1(Δ/Δ)) developed progressive hyperglycaemia and full-blown diabetes mellitus in adulthood that recapitulated the natural history of the spontaneous disease in mice. Reduced insulin gene expression and concomitant reduced insulin secretion preceded the hyperglycaemic state and diabetes development. Immunohistochemical, flow cytometric and ultrastructural analyses revealed altered islet morphology, marked decreased β-cell mass, reduced numbers of granules within the β-cells and reduced granule docking in adult RIP-Cre Dicer1(Δ/Δ) mice. β-cell specific Dicer1 deletion did not appear to disrupt fetal and neonatal β-cell development as 2-week old RIP-Cre Dicer1(Δ/Δ) mice showed ultrastructurally normal β-cells and intact insulin secretion. In conclusion, we have demonstrated that a β-cell specific disruption of the miRNAs network, although allowing for apparently normal β-cell development, leads to progressive impairment of insulin secretion, glucose homeostasis and diabetes development.

OBJECTIVE The aim of this study was to refine the information regarding the quantitative and spatial dynamics of infiltrating lymphocytes and remaining β-cell volume during the progression of type 1 diabetes in the nonobese diabetic (NOD) mouse model of the disease. RESEARCH DESIGN AND METHODS Using an ex vivo technique, optical projection tomography (OPT), we quantified and assessed the three-dimensional spatial development and progression of insulitis and β-cell destruction in pancreata from diabetes-prone NOD and non–diabetes-prone congenic NOD.H-2b mice between 3 and 16 weeks of age. RESULTS Together with results showing the spatial dynamics of the insulitis process, we provide data of β-cell volume distributions down to the level of the individual islets and throughout the pancreas during the development and progression of type 1 diabetes. Our data provide evidence for a compensatory growth potential of the larger insulin+ islets during the later stages of the disease around the time point for development of clinical diabetes. This is in contrast to smaller islets, which appear less resistant to the autoimmune attack. We also provide new information on the spatial dynamics of the insulitis process itself, including its apparently random distribution at onset, the local variations during its further development, and the formation of structures resembling tertiary lymphoid organs at later phases of insulitis progression. CONCLUSIONS Our data provide a powerful tool for phenotypic analysis of genetic and environmental effects on type 1 diabetes etiology as well as for evaluating the potential effect of therapeutic regimes.

Tuning of TCR-mediated activation was demonstrated to be critical for lineage fate in T cell development, as well as in the control of autoimmunity. In this study, we identify a novel diabetes susceptibility gene, Idd28, in the NOD mouse and provide evidence that Cd3ζ (Cd247) constitutes a prime candidate gene for this locus. Moreover, we show that the allele of the Cd3ζ gene expressed in NOD and DBA/2 mouse strains confers lower levels of T cell activation compared with the allele expressed by C57BL/6 (B6), BALB/c, and C3H/HeJ mice. These results support a model in which the development of autoimmune diabetes is dependent on a TCR signal mediated by a less-efficient NOD allele of the Cd3ζ gene.