| 1. |
- Edlund, Anna, et al.
(författare)
-
Defective exocytosis and processing of insulin in a cystic fibrosis mouse model
- 2019
-
Ingår i: Journal of Endocrinology. - 1479-6805. ; 241:1, s. 45-57
-
Tidskriftsartikel (refereegranskat)abstract
- Cystic fibrosis-related diabetes (CFRD) is a common complication for patients with cystic fibrosis (CF), a disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR). The cause of CFRD is unclear, but a commonly observed reduction in first-phase insulin secretion suggests defects at the beta cell level. Here we aimed to examine beta- and alpha-cell function in the Cftrtm1EUR/F508del mouse model (C57BL/6J), which carries the most common human mutation in CFTR, the F508del mutation. CFTR expression, beta cell mass, insulin granule distribution, hormone secretion and single cell capacitance changes were evaluated using islets (or beta cells) from F508del mice and age-matched wild-type mice aged 7-10 weeks. Granular pH was measured with DND-189 fluorescence. Serum glucose, insulin and glucagon levels were measured in vivo, and glucose tolerance was assessed using IPGTT. We show increased secretion of proinsulin and concomitant reduced secretion of C-peptide in islets from F508del mice compared to WT mice. Exocytosis and number of docked granules was reduced. We confirmed reduced granular pH by CFTR stimulation. We detected decreased pancreatic beta cell area, but unchanged beta cell number. Moreover, the F508del mutation caused failure to suppress glucagon secretion leading to hyperglucagonemia. In conclusion, F508del mice have beta cell defects resulting in 1) reduced number of docked insulin granules and reduced exocytosis, and 2) potential defective proinsulin cleavage and secretion of immature insulin. These observations provide insight into the functional role of CFTR in pancreatic islets and contribute to increased understanding of the pathogenesis of CFRD.
|
|
| 2. |
- Forkel, Marianne, et al.
(författare)
-
Composition and functionality of the intrahepatic innate lymphoid cell-compartment in human nonfibrotic and fibrotic livers
- 2017
-
Ingår i: European Journal of Immunology. - : Wiley. - 0014-2980 .- 1521-4141. ; 47:8, s. 1280-1294
-
Tidskriftsartikel (refereegranskat)abstract
- Human innate lymphoid cells have been described to exist in different organs, with functional deregulation of these cells contributing to several disease states. Here, we performed the first detailed characterization of the phenotype, tissue-residency properties, and functionality of ILC1s, ILC2s, and ILC3s in the human adult and fetal liver. In addition, we investigated changes in the ILC compartment in liver fibrosis. A unique composition of tissue-resident ILCs was observed in nonfibrotic livers as compared with that in mucosal tissues, with NKp44− ILC3s accounting for the majority of total intrahepatic ILCs. The frequency of ILC2s, representing a small fraction of ILCs in nonfibrotic livers, increased in liver fibrosis and correlated directly with the severity of the disease. Notably, intrahepatic ILC2s secreted the profibrotic cytokine IL-13 when exposed to IL-33 and thymic stromal lymphopoetin (TSLP); these cytokines were produced by hepatocytes, hepatic stellate cells (HSCs), and Kupffer cells in response to TLR-3 stimulation. In summary, the present results provide the first detailed characterization of intrahepatic ILCs in human adult and fetal liver. The results indicate a role for ILC2s in human liver fibrosis, implying that targeting ILC2s might be a novel therapeutic strategy for its treatment.
|
|
| 3. |
- Pedersen, Morten Gram, et al.
(författare)
-
Calcium Current Inactivation Rather than Pool Depletion Explains Reduced Exocytotic Rate with Prolonged Stimulation in Insulin-Secreting INS-1 832/13 Cells.
- 2014
-
Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 9:8
-
Tidskriftsartikel (refereegranskat)abstract
- Impairment in beta-cell exocytosis is associated with reduced insulin secretion and diabetes. Here we aimed to investigate the dynamics of Ca2+-dependent insulin exocytosis with respect to pool depletion and Ca2+-current inactivation. We studied exocytosis, measured as increase in membrane capacitance (ΔCm), as a function of calcium entry (Q) in insulin secreting INS-1 832/13 cells using patch clamp and mixed-effects statistical analysis. The observed linear relationship between ΔCm and Q suggests that Ca2+-channel inactivation rather than granule pool restrictions is responsible for the decline in exocytosis observed at longer depolarizations. INS-1 832/13 cells possess an immediately releasable pool (IRP) of ∼10 granules and most exocytosis of granules occurs from a large pool. The latter is attenuated by the calcium-buffer EGTA, while IRP is unaffected. These findings suggest that most insulin release occurs away from Ca2+-channels, and that pool depletion plays a minor role in the decline of exocytosis upon prolonged stimulation.
|
|
| 4. |
|
|
| 5. |
- Richardson, Sarah J., et al.
(författare)
-
Evaluation of the fidelity of immunolabelling obtained with clone 5D8/1, a monoclonal antibody directed against the enteroviral capsid protein, VP1, in human pancreas
- 2014
-
Ingår i: Diabetologia. - : Springer Science and Business Media LLC. - 0012-186X .- 1432-0428. ; 57:2, s. 392-401
-
Tidskriftsartikel (refereegranskat)abstract
- Aims/hypothesisEnteroviral infection has been implicated in the development of islet autoimmunity in type 1 diabetes and enteroviral antigen expression has been detected by immunohistochemistry in the pancreatic beta cells of patients with recent-onset type 1 diabetes. However, the immunohistochemical evidence relies heavily on the use of a monoclonal antibody, clone 5D8/1, raised against an enteroviral capsid protein, VP1. Recent data suggest that the clone 5D8/1 may also recognise non-viral antigens; in particular, a component of the mitochondrial ATP synthase (ATP5B) and an isoform of creatine kinase (CKB). Therefore, we evaluated the fidelity of immunolabelling by clone 5D8/1 in the islets of patients with type 1 diabetes.MethodsEnteroviral VP1, CKB and ATP5B expression were analysed by western blotting, RT-PCR and immunocytochemistry in a range of cultured cell lines, isolated human islets and human tissue.ResultsClone 5D8/1 labelled CKB, but not ATP5B, on western blots performed under denaturing conditions. In cultured human cell lines, isolated human islets and pancreas sections from patients with type 1 diabetes, the immunolabelling of ATP5B, CKB and VP1 by 5D8/1 was readily distinguishable. Moreover, in a human tissue microarray displaying more than 80 different cells and tissues, only two (stomach and colon; both of which are potential sites of enterovirus infection) were immunopositive when stained with clone 5D8/1.Conclusions/interpretationWhen used under carefully optimised conditions, the immunolabelling pattern detected in sections of human pancreas with clone 5D8/1 did not reflect cross-reactivity with either ATP5B or CKB. Rather, 5D8/1 is likely to be representative of enteroviral antigen expression.
|
|
| 6. |
- Svedin, Emma
(författare)
-
Enterovirus infections in type 1 diabetes and cystic fibrosis : antiviral defence and viral immune evasion strategies
- 2017
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Enteroviruses are common viruses which cause infections in humans that usually result in mild flu-like symptoms before viral clearance. However, in some cases these infections can progress to more severe diseases such as myocarditis, pancreatitis and hepatitis. Coxsackievirus induced hepatitis in infants may become so severe that the outcome is fatal. In addition, infections with enteroviruses belonging to the group B Coxsackieviruses have been implicated in the etiology of type 1 diabetes. Enterovirus infections are also commonly observed in patients with cystic fibrosis, resulting in lung exacerbations and morbidity. Proper antiviral response mechanisms are crucial for the prevention of viral replication and spread, as well as the inhibition of virus induced cellular damage. Recently a novel group of interferons, called type III interferons, were discovered and shown to have antiviral properties predominantly in cells of epithelial origin. In Paper I we show that type III interferons protect primary human hepatocytes from Coxsackievirus infection. Given the importance of interferons in preventing early viral replication, many viruses have developed mechanisms to inhibit their induction. In Paper II, we showed that Coxsackieviruses inhibit the induction of type III interferons in infected cells. In addition, we demonstrated that this inhibition was caused by the proteolytic activity of the viral protease 2Apro. The exact role of enterovirus infections in type 1 diabetes development is still under speculation. Coxsackieviruses encodes several viral proteins that have been shown to interfere with cellular function and signaling pathways. In Paper III, we used primary human pancreatic islets and an insulin-secreting cell line to identify mechanisms by which Coxsackeiviruses can cause beta cell dysfunction. We found that the viral proteins 2Apro, 3A and 3Cpro could, independently of one other, affect exocytosis with 2Apro and 3Cpro targeting calcium influx while 3A inhibited exocytosis via a calcium independent mechanism. An impaired antimicrobial defense has been observed in patients with cystic fibrosis. This could explain why common respiratory infections are often prolonged and more severe in these patients. By using a mouse model for cystic fibrosis, we showed in Paper IV that the most common mutation resulting in cystic fibrosis, F508del, caused an impaired adaptive immune response with a delayed production of neutralizing antibodies to Coxsackievirus. In conclusion, the studies performed in this thesis add to our understanding of innate and adaptive immune response mechanisms during Coxackievirus infections. In addition they demonstrate a role for viral proteins in circumventing host antiviral immune responses, thus causing cellular damage, which could contribute to disease pathology. Increasing the knowledge of host-pathogen interactions may help to develop new treatments that could prevent severe Coxsackievirus infections.
|
|
