1,25-dihydroxyvitamin D3 induces stable and reproducible therapeutic tolerogenic dendritic cells with specific epigenetic modifications.

Autologous, antigen-specific, tolerogenic dendritic cells (tolDCs) are presently assessed to reverse and possibly cure autoimmune diseases such as type 1 diabetes (T1D). Good Manufacturing Practice production and clinical implementation of such cell therapies critically depend on their stability and reproducible production from healthy donors and, more importantly, patient-derived monocytes. Here the authors demonstrate that tolDCs (modulated using 1,25-dihydroxyvitamin D3 and dexamethasone) displayed similar features, including protein, transcriptome and epigenome profiles, between two international clinical centers and between T1D and healthy donors, validating reproducible production. In addition, neither phenotype nor function of tolDCs was affected by repeated stimulation with inflammatory stimuli, underscoring their stability as semi-mature DCs. Furthermore, tolDCs exhibited differential DNA methylation profiles compared with inflammatory mature DCs (mDCs), and this was already largely established prior to maturation, indicating that tolDCs are locked into an immature state. Finally, approximately 80% of differentially expressed known T1D risk genes displayed a corresponding differential DNA methylome in tolDCs versus mDCs and metabolic and immune pathway genes were also differentially methylated and expressed. In summary, tolDCs are reproducible and stable clinical cell products unaffected by the T1D status of donors. The observed stable, semi-mature phenotype and function of tolDCs are exemplified by epigenetic modifications representative of immature-stage cells. Together, the authors' data provide a strong basis for the production and clinical implementation of tolDCs in the treatment of autoimmune diseases such as T1D.

Relapsing/remitting type 1 diabetes.

AIMS/HYPOTHESIS: Type 1 diabetes is believed to be an autoimmune disease associated with irreversible loss of insulin secretory function that follows a chronic progressive course. However, it has been speculated that relapsing/remitting disease progression may occur in type 1 diabetes. METHODS: We report the case of an 18-year-old girl with Graves' disease, chronic inflammatory demyelinating polyneuropathy (CIDP) and multiple islet autoantibodies, presenting with relapsing/remitting hyperglycaemia. Peripheral blood mononuclear cells were analysed for islet autoimmunity. RESULTS: There were two instances of hyperglycaemia relapse during CIDP flare-ups that required insulin therapy and remitted after i.v. immunoglobulin (IVIG) therapy improving neurological symptoms. A diagnosis of type 1 diabetes was assigned on the basis of insulin need, HbA1c and islet autoantibodies. Insulin requirements disappeared following IVIG treatment and peaked during CIDP flare-ups. Pro- and anti-inflammatory cytokine responses were noted against islet autoantigens. CONCLUSIONS/INTERPRETATION: We provide clinical evidence of relapsing/remitting type 1 diabetes associated with IVIG treatment and the regulation of islet autoimmunity. Despite sufficient residual beta cell mass, individuals can experience episodes of impaired glycaemia control. This disconnect between beta cell mass and function highlighted by our case may have implications for the use of beta cell function as the primary endpoint for immune intervention trials aiming to protect beta cell mass rather than function. Immune modulation may restore beta cell function and glycaemic control.

[Gastric rupture as a complication of anorexia nervosa]. / Maagruptuur als complicatie van anorexia nervosa.

BACKGROUND: Anorexia nervosa may cause several gastro-intestinal complications. CASE DESCRIPTION: A 21-year-old woman presented herself with abdominal pain and vomiting a day after her first binge-eating episode. In her recent history she had lost 40 kg in weight and her BMI was 15 at presentation. Imaging showed gastric distension and ascites in all quadrants, suspicious of a gastric rupture. A blow-out of the stomach was seen during a diagnostic laparotomy, which was treated with a gastric sleeve procedure. Following surgery, the patient was diagnosed with anorexia nervosa of the restrictive type with a first episode of binge-eating. CONCLUSION: Gastric rupture is a rare and unknown complication of anorexia nervosa with a high mortality.

Activated Mesenchymal Stromal Cells Process and Present Antigens Regulating Adaptive Immunity.

Mesenchymal stromal cells (MSCs) are inherently immunomodulatory through production of inhibiting soluble factors and expression of immunosuppressive cell surface markers. We tested whether activated MSCs qualify for the induction of antigen-specific immune regulation. Bone marrow derived human MSCs were activated by interferon-γ and analyzed for antigen uptake and processing and immune regulatory features including phenotype, immunosuppressive capacity, and metabolic activity. To assess whether activated MSC can modulate adaptive immunity, MSCs were pulsed with islet auto-antigen (GAD65) peptide to stimulate GAD65-specific T-cells. We confirm that inflammatory activation of MSCs increased HLA class II, PD-L1, and intracellular IDO expression, whereas co-stimulatory molecules including CD86 remained absent. MSCs remained locked in their metabolic phenotype, as activation did not alter glycolytic function or mitochondrial respiration. MSCs were able to uptake and process protein. Activated HLA-DR3-expressing MSCs pulsed with GAD65 peptide inhibited proliferation of HLA-DR3-restricted GAD65-specific T-cells, while this HLA class II expression did not induce cellular alloreactivity. Conditioning of antigen-specific T-cells by activated and antigen-pulsed MSCs prevented T-cells to proliferate upon subsequent activation by dendritic cells, even after removal of the MSCs. In sum, activation of MSCs with inflammatory stimuli turns these cells into suppressive cells capable of mediating adaptive regulation of proinflammatory pathogenic T-cells.

Intra-pancreatic tissue-derived mesenchymal stromal cells: a promising therapeutic potential with anti-inflammatory and pro-angiogenic profiles.

BACKGROUND: Human pancreata contain many types of cells, such as endocrine islets, acinar, ductal, fat, and mesenchymal stromal cells (MSCs). MSCs are important and shown to have a promising therapeutic potential to treat various disease conditions. METHODS: We investigated intra-pancreatic tissue-derived (IPTD) MSCs isolated from tissue fractions that are routinely discarded during pancreatic islet isolation of human cadaveric donors. Furthermore, whether pro-angiogenic and anti-inflammatory properties of these cells could be enhanced was investigated. RESULTS: IPTD-MSCs were expanded in GMP-compatible CMRL-1066 medium supplemented with 5% human platelet lysate (hPL). IPTD-MSCs were found to be highly pure, with > 95% positive for CD90, CD105, and CD73, and negative for CD45, CD34, CD14, and HLA-DR. Immunofluorescence staining of pancreas tissue demonstrated the presence of CD105+ cells in the vicinity of islets. IPTD-MSCs were capable of differentiation into adipocytes, chondrocytes, and osteoblasts in vitro, underscoring their multipotent features. When these cells were cultured in the presence of a low dose of TNF-α, gene expression of tumor necrosis factor alpha-stimulated gene-6 (TSG-6) was significantly increased, compared to control. In contrast, treating cells with dimethyloxallyl glycine (DMOG) (a prolyl 4-hydroxylase inhibitor) enhanced mRNA levels of nuclear factor erythroid 2-related factor 2 (NRF2) and vascular endothelial growth factor (VEGF). Interestingly, a combination of TNF-α and DMOG stimulated the optimal expression of all three genes in IPTD-MSCs. Conditioned medium of IPTD-MSCs treated with a combination of DMOG and TNF-α contained higher levels of pro-angiogenic (VEGF, IL-6, and IL-8) compared to controls, promoting angiogenesis of human endothelial cells in vitro. In contrast, levels of MCP-1, a pro-inflammatory cytokine, were reduced in the conditioned medium of IPTD-MSCs treated with a combination of DMOG and TNF-α. CONCLUSIONS: The results demonstrate that IPTD-MSCs reside within the pancreas and can be separated as part of a standard islet-isolation protocol. These IPTD-MSCs can be expanded and potentiated ex vivo to enhance their anti-inflammatory and pro-angiogenic profiles. The fact that IPTD-MSCs are generated in a GMP-compatible procedure implicates a direct clinical application.