| 1. |
- Ahl, David, et al.
(författare)
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Turning Up the Heat : Local Temperature Control During in vivo Imaging of Immune Cells
- 2019
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Ingår i: Frontiers in Immunology. - : FRONTIERS MEDIA SA. - 1664-3224. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Intravital imaging is an invaluable tool for studying the expanding range of immune cell functions. Only in vivo can the complex and dynamic behavior of leukocytes and their interactions with their natural microenvironment be observed and quantified. While the capabilities of high-speed, high-resolution confocal and multiphoton microscopes are well-documented and steadily improving, other crucial hardware required for intravital imaging is often developed in-house and less commonly published in detail. In this report, we describe a low-cost, multipurpose, and tissue-stabilizing in vivo imaging platform that enables sensing and regulation of local tissue temperature. The effect of tissue temperature on local blood flow and leukocyte migration is demonstrated in muscle and skin. Two different models of vacuum windows are described in this report, however, the design of the vacuum window can easily be adapted to fit different organs and tissues.
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| 2. |
- Benedict, Christian, et al.
(författare)
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Acute sleep deprivation increases serum levels of neuron-specific enolase (NSE) and S100 calcium binding protein B (S-100B) in healthy young men.
- 2014
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Ingår i: Sleep. - : Oxford University Press (OUP). - 1550-9109 .- 0161-8105. ; 37:1, s. 195-8
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Tidskriftsartikel (refereegranskat)abstract
- To investigate whether total sleep deprivation (TSD) affects circulating concentrations of neuron-specific enolase (NSE) and S100 calcium binding protein B (S-100B) in humans. These factors are usually found in the cytoplasm of neurons and glia cells. Increasing concentrations of these factors in blood may be therefore indicative for either neuronal damage, impaired blood brain barrier function, or both. In addition, amyloid β (Aβ) peptides 1-42 and 1-40 were measured in plasma to calculate their ratio. A reduced plasma ratio of Aβ peptides 1-42 to 1-40 is considered an indirect measure of increased deposition of Aβ 1-42 peptide in the brain.
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| 3. |
- Chodaczek, Grzegorz, et al.
(författare)
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The effect of Toll-like receptor stimulation on the motility of regulatory T cells
- 2021
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Ingår i: Journal of Autoimmunity. - : Elsevier. - 0896-8411 .- 1095-9157. ; 116
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Tidskriftsartikel (refereegranskat)abstract
- Regulatory T cells (Tregs) have suppressive functions and play an important role in controlling inflammation and autoimmunity. The migratory capacity of Tregs determines their location and their location determines whether they inhibit the priming of naïve lymphocytes in lymphoid tissues or the effector phase of immune responses at inflamed sites. Tregs generated or expanded in vitro are currently being tested in clinics for the treatment of autoimmune disorders, however, little is known about the factors controlling their migration towards therapeutically relevant locations. In this study, we have modulated Treg dynamics using Toll-like receptor (TLR) agonists. Dynamic imaging with confocal and two-photon microscopy revealed that Tregs generated in vitro and stimulated with P3C (a TLR2 agonist) but not with R848 (a TLR7 agonist) or LPS (a TLR4 agonist) showed enhanced cell migration within splenic white pulp or draining lymph node when transferred into mice intravenously or into the footpad, respectively. In summary, our data demonstrate that Tregs are more motile in response to direct TLR stimulation in particular towards TLR2 signals. This may have implications for efficient clinical Treg induction protocols.
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| 4. |
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| 5. |
- Christoffersson, Gustaf, et al.
(författare)
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Acute sleep deprivation in healthy young men : Impact on population diversity and function of circulating neutrophils
- 2014
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Ingår i: Brain, behavior, and immunity. - : Elsevier BV. - 0889-1591 .- 1090-2139. ; 41, s. 162-172
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Tidskriftsartikel (refereegranskat)abstract
- Lack of sleep greatly affects our immune system. The present study investigates the acute effects of total sleep deprivation on blood neutrophils, the most abundant immune cell in our circulation and the first cell type recruited to sites of infection. Thus, the population diversity and function of circulating neutrophils were compared in healthy young men following one night of total sleep deprivation (TSD) or after 8 h regular sleep. We found that neutrophil counts were elevated after nocturnal wakefulness (2.0 +/- 0.2 x 10(9)/l vs. 2.6 +/- 0.2 x 10(9)/l, sleep vs. TSD, respectively) and the population contained more immature CD16(dim)/CD62L(bright) cells (0.11 +/- 0.040 x 10(9)/l [5.5 +/- 1.1%] vs. 0.26 +/- 0.020 x 10(9)/l [9.9 +/- 1.4%]). As the rise in numbers of circulating mature CD16(bright)/CD62L(bright) neutrophils was less pronounced, the fraction of this subpopulation showed a significant decrease (1.8 +/- 0.15 x 10(9)/l [88 +/- 1.8%] vs. 2.1 +/- 0.12 x 10(9)/l [82 +/- 2.8%]). The surface expression of receptors regulating mobilization of neutrophils from bone marrow was decreased (CXCR4 and CD49d on immature neutrophils; CXCR2 on mature neutrophils). The receptor CXCR2 is also involved in the production of reactive oxygen species (ROS), and in line with this, total neutrophils produced less ROS. In addition, following sleep loss, circulating neutrophils exhibited enhanced surface levels of CD11b, which indicates enhanced granular fusion and concomitant protein translocation to the membrane. Our findings demonstrate that sleep loss exerts significant effects on population diversity and function of circulating neutrophils in healthy men. To which extent these changes could explain as to why people with poor sleep patterns are more susceptible to infections warrants further investigation.
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| 6. |
- Christoffersson, Gustaf, et al.
(författare)
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Clinical and Experimental Pancreatic Islet Transplantation to Striated Muscle : Establishment of a Vascular System Similar to that in Native Islets
- 2010
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 59:10, s. 2569-2578
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Tidskriftsartikel (refereegranskat)abstract
- Objective: Curing type 1 diabetes by transplanting pancreatic islets into the liver is associated with poor long-term outcome and graft failure at least partly due to inadequate graft revascularization. The aim of the current study was to evaluate striated muscle as a potential angiogenic site for islet transplantation. Research Design and Methods: The current study presents a new experimental model which is found applicable to clinical islet transplantation. Islets were implanted into striated muscle where after intra-islet vascular density and blood flow were visualized with intravital and confocal microscopy in mice, and by magnetic resonance imaging in three auto-transplanted pancreatectomized patients. Mice were rendered neutropenic by repeated injections of Gr-1 antibody and diabetes was induced by alloxan treatment. Results: Contrary to liver-engrafted islets, islets transplanted to mouse muscle were revascularized with vessel densities and blood flow entirely comparable to islets within intact pancreas. Initiation of islet revascularization at the muscular site was dependent on neutrophils, and the function of islets transplanted to muscle was proven by curing diabetic mice. The experimental data were confirmed in auto-transplanted patients where higher plasma volumes were measured in islets engrafted in forearm muscle compared to adjacent muscle tissue through high-resolution magnetic resonance imaging. Conclusions: This study presents a novel paradigm in islet transplantation whereby recruited neutrophils are crucial for the functionally restored intra-islet blood perfusion following transplantation to striated muscle under experimental and clinical situations.
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| 7. |
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| 8. |
- Christoffersson, Gustaf, et al.
(författare)
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Interference with pancreatic sympathetic signaling halts the onset of diabetes in mice
- 2020
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Ingår i: Science Advances. - : American Association for the Advancement of Science (AAAS). - 2375-2548. ; 6:35
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Tidskriftsartikel (refereegranskat)abstract
- The notably lobular distribution of immune lesions in type 1 diabetes (T1D) has been hypothesized to be the result of innervation within the pancreas. To investigate whether neuroimmune interactions could explain this phenomenon, we explored the impact of sympathetic signaling in the RIP-LCMV-GP mouse model of autoimmune diabetes. In this model, the CD8(+) T cell attack on beta cells replicates a key pathogenic feature of human T1D. We found that inhibition of alpha(1) adrenoceptors, ablation of sympathetic nerves, and surgical denervation all had a protective effect in this model, without affecting the systemic presence of beta cell-reactive CD8(+) T cells. In vivo multiphoton imaging revealed a local effect within pancreatic islets including limited infiltration of both macrophages and beta cell-specific CD8(+) T cells. Islet-resident macrophages expressed adrenoceptors and were responsive to catecholamines. Islet macrophages may therefore constitute a pivotal neuroimmune signaling relay and could be a target for future interventions in T1D.
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| 9. |
- Christoffersson, Gustaf, et al.
(författare)
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Intramuscular islet transplantation promotes restored islet vascularity
- 2011
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Ingår i: Islets. - : Informa UK Limited. - 1938-2014 .- 1938-2022. ; 3:2, s. 69-71
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- In a recent publication, we reported that islets transplanted to mouse striated muscle became revascularized with intra-islet vessel densities comparable to native islets. Revascularization of islet grafts was completely dependent on recruited Gr-1+ leukocytes. Diabetic mice cured by transplantation of 300 islets into muscle handled glucose tolerance tests as healthy controls, whereas mice cured by intraportal islet transplantation into the liver had increased blood glucose values during the load. The translational impact of these observations were confirmed by magnetic resonance imaging of autotransplanted islets in the forearm muscle of pancreactomized patients, and higher blood perfusion of the grafts compared to adjacent muscle were found. In summary, the striated muscle is a promising site for islet transplantation which promotes full revascularization of implanted grafts. The proangiogenic role of recruited leukocytes during engraftment needs to be further characterized, and considered for immune suppression treatments.
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| 10. |
- Christoffersson, Gustaf
(författare)
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Leukocytes in Angiogenesis : Learning from Transplanted Pancreatic Islets
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Angiogenesis, the growth of new blood vessels, is a complex process involving several cell types and molecular signals. Excessive vascular growth is a problem in tumors, and insufficient vascularization hampers the function of transplanted insulin-producing pancreatic islets. Understanding the mechanisms behind blood vessel growth generates increased means to control angiogenesis. In this thesis a model of pancreatic islet transplantation to muscle has been used to study the involvement of leukocytes in the development of new vasculature.Transplantation of isolated islets of Langerhans into mouse muscle promoted revascularization of the grafts to a level comparable to native islets in the pancreas. The complete and functional vascular restoration resulted in improved blood glucose control compared to the clinical standard implantation site, the liver. This proved muscle as a transplantation site to be a clinically relevant option for the treatment of type 1 diabetes.The rapid islet revascularization process was found to be dependent on a distinct subset of neutrophils characterized by high expression of the chemokine receptor CXCR4 and the enzyme matrix metalloproteinase 9 (MMP-9). These cells were recruited to recently transplanted and hypoxic grafts by islet-secreted vascular endothelial growth factor A (VEGF-A). Leukocyte migration and interactions in the engraftment area were monitored using a high-speed confocal microscope followed by software tracking. New software was developed to visualize migration statistics. This tool revealed areas around the islet graft where neutrophil gathering coincided with sites of angiogenesis. Macrophages in the engraftment area positioned themselves close to the newly formed vasculature and were shown to have a stabilizing effect on the vessels. When macrophages were removed, no pericytes were recruited to the forming vasculature. The perivascular macrophages also began to express a pericyte marker when in the graft, suggesting a close relationship between these cell types or macrophage plasticity.In conclusion, this thesis presents muscle as a proangiogenic transplantation site for pancreatic islets for the treatment of type 1 diabetes, where the revascularization of the grafts was dependent on the recruitment and actions of specialized immune cells.
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| 11. |
- Christoffersson, Gustaf, et al.
(författare)
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Matrix Metalloproteinase-9 Is Essential for Physiological Beta Cell Function and Islet Vascularization in Adult Mice
- 2015
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Ingår i: American Journal of Pathology. - : Elsevier BV. - 0002-9440 .- 1525-2191. ; 185:4, s. 1094-1103
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Tidskriftsartikel (refereegranskat)abstract
- The availability of paracrine factors in the islets of Langerhans, and the constitution of the beta cell basement membrane can both be affected by proteolytic enzymes. This study aimed to investigate the effects of the extraceaular matrix-degrading enzyme gelatinase B/matrix metalloproteinase-9 (Mmp-9) on islet function in mice. Islet function of Mmp9-deficient (Mmp9(-/-)) mice and their wild-type Littermates was evaluated both in vivo and in vitro. The pancreata of Mmp9(-/-) mice did not differ from wild type in islet mass or distribution. However, Mmp9(-/-) mice had an impaired response to a glucose toad in vivo, with lower serum insulin levels. The glucose-stimulated insulin secretion was reduced also in vitro in isolated Mmp9(-/-) islets. The vascular density of Mmp9(-/-) islets was lower, and the capillaries had fewer fenestrations, whereas the islet blood flow was threefold higher. These alterations could partly be explained by compensatory changes in the expression of matrix-related proteins. This in-depth investigation of the effects of the loss of MMP9(-/-) function on pancreatic islets uncovers a deteriorated beta cell function that is primarily due to a shift in the beta cell phenotype, but also due to islet vascular aberrations. This likely reflects the importance of a normal islet matrix turnover exerted by MMP-9, and concomitant release of paracrine factors sequestered on the matrix.
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| 12. |
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| 13. |
- Christoffersson, Gustaf, et al.
(författare)
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Regulatory Immune Mechanisms beyond Regulatory T Cells
- 2019
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Ingår i: Trends in immunology. - : Elsevier BV. - 1471-4906 .- 1471-4981. ; 40:6, s. 482-491
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Forskningsöversikt (refereegranskat)abstract
- In autoimmunity, aggressive immune responses are counteracted by suppressive rejoinders. For instance, FOXP3-expressing regulatory T cells (Tregs), have shown remarkable effects in limiting autoimmunity in preclinical models. However, early results from human Treg trials have not been as positive. Here, we highlight questions surrounding Treg transfers as putative treatments for autoimmunity. We discuss whether lack of antigenic recognition might be key to shifting cells from contributing to an aggressive autoresponse, to being part of a regulatory network. Moreover, we argue that identifying the physiological range of immunosuppression of Tregs might help potentiate their efficacy. We propose widening the view on immunoregulation by considering the participation of CD8(+) Tregs in this process, which could have major implications in autoimmunity.
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| 14. |
- Christoffersson, Gustaf, et al.
(författare)
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The neutrophil : one cell on many missions or many cells with different agendas?
- 2018
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Ingår i: Cell and Tissue Research. - : Springer. - 0302-766X .- 1432-0878. ; 371:3, s. 415-423
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Forskningsöversikt (refereegranskat)abstract
- The unique role of neutrophils in host defense is not only based on their abilities to kill bacteria but is also due to their abundance in circulation and their ability to quickly migrate and accumulate in great numbers at afflicted sites. The high number of circulating neutrophils is the result of regulated release of new neutrophils from bone marrow as well as from marginated pools to balance their recruitment to tissue. Marginated pools, such as the spleen and lung, have previously been attributed to passively delay neutrophil transit time due to their large capillary network, but recent reports demonstrate that they are comprised of neutrophils with specific functions. The spleen, for instance, holds neutrophil subpopulations at different anatomical locations with distinct functions important for, e.g., bacterial eradication, and the lung was recently shown to re-educate neutrophils that had trafficked from a site of sterile injury to home back to bone marrow for elimination. Further, recent reports demonstrate subpopulations of neutrophils with different actions during homeostasis, infection, tissue restitution and cancer. It is becoming increasingly clear that this cannot be due to different stages of neutrophil activation during their life span but instead points towards distinct subpopulations of neutrophils with different effector functions. Whether these cellular distinctions are due to different education or origin is, however, not yet known. Together, the accumulating information about the heterogeneous neutrophils presents important insights into their role in development of pathologies, as well as revealing novel targets in the form of certain subpopulations to treat disease.
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| 15. |
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| 16. |
- Christoffersson, Gustaf, et al.
(författare)
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VEGF-A recruits a proangiogenic MMP-9-delivering neutrophil subset that induces angiogenesis in transplanted hypoxic tissue
- 2012
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Ingår i: Blood. - : American Society of Hematology. - 0006-4971 .- 1528-0020. ; 120:23, s. 4653-4662
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Tidskriftsartikel (refereegranskat)abstract
- Recruitment and retention of leukocytes at a site of blood vessel growth are crucial for proper angiogenesis and subsequent tissue perfusion. Although critical for many aspects of regenerative medicine, the mechanisms of leukocyte recruitment to and actions at sites of angiogenesis are not fully understood. In this study, we investigated the signals attracting leukocytes to avascular transplanted pancreatic islets and leukocyte actions at the engraftment site. Expression of the angiogenic stimulus VEGF-A by mouse pancreatic islets was elevated shortly after syngeneic transplantation to muscle. High levels of leukocytes, predominantly CD11b+/Gr-1+/CXCR4hi neutrophils, were observed at the site of engraftment, whereas VEGF-A–deficient islets recruited only half of the amount of leukocytes when transplanted. Acute VEGF-A exposure of muscle increased leukocyte extravasation but not the levels of SDF-1α. VEGF-A–recruited neutrophils expressed 10 times higher amounts of MMP-9 than neutrophils recruited to an inflammatory stimulus. Revascularization of islets transplanted to MMP-9–deficient mice was impaired because blood vessels initially failed to penetrate grafts, and after 2 weeks vascularity was still disturbed. This study demonstrates that VEGF-A recruits a proangiogenic circulating subset of CD11b+/Gr-1+ neutrophils that are CXCR4hi and deliver large amounts of the effector protein MMP-9, required for islet revascularization and functional integration after transplantation.
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| 17. |
- Espes, Daniel, et al.
(författare)
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Rapid Restoration of Vascularity and Oxygenation in Mouse and Human Islets Transplanted to Omentum May Contribute to Their Superior Function Compared to Intraportally Transplanted Islets
- 2016
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Ingår i: American Journal of Transplantation. - : Elsevier BV. - 1600-6135 .- 1600-6143. ; 16:11, s. 3246-3254
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Tidskriftsartikel (refereegranskat)abstract
- Transplantation of islets into the liver confers several site-specific challenges, including a delayed vascularization and prevailing hypoxia. The greater omentum has in several experimental studies been suggested as an alternative implantation site for clinical use, but there has been no direct functional comparison to the liver. In this experimental study in mice, we characterized the engraftment of mouse and human islets in the omentum and compared engraftment and functional outcome with those in the intraportal site. The vascularization and innervation of the islets transplanted into the omentum were restored within the first month by paralleled ingrowth of capillaries and nerves. The hypoxic conditions in the islets early posttransplantation were transient and restricted to the first days. Newly formed blood vessels were fully functional, and the blood perfusion and oxygenation of the islets became similar to that of endogenous islets. Furthermore, islet grafts in the omentum showed at 1 month posttransplantation functional superiority to intraportally transplanted grafts. We conclude that in contrast to the liver the omentum provides excellent engraftment conditions for transplanted islets. Future studies in humans will be of great interest to investigate the capability of this site to also harbor larger grafts without interfering with islet functionality.
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| 18. |
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| 19. |
- Grapensparr, Liza, et al.
(författare)
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Bioengineering with Endothelial Progenitor Cells Improves the Vascular Engraftment of Transplanted Human Islets
- 2018
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Ingår i: Cell Transplantation. - : SAGE Publications. - 0963-6897 .- 1555-3892. ; 27:6, s. 948-956
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Tidskriftsartikel (refereegranskat)abstract
- Pancreatic islets isolated for transplantation are disconnected from their vascular supply and need to establish a new functional network posttransplantation. Due to poor revascularization, prevailing hypoxia with correlating increased apoptosis rates in experimental studies can be observed for months posttransplantation. Endothelial progenitor cells (EPCs) are bone marrow-derived cells that promote neovascularization. The present study tested the hypothesis that EPCs, isolated from human umbilical cord blood, could be coated to human islet surfaces and be used to promote islet vascular engraftment. Control or EPC bioengineered human islets were transplanted into the renal subcapsular space of nonobese diabetic/severe combined immunodeficiency mice. Four weeks posttransplantation, graft blood perfusion and oxygen tension were measured using laser Doppler flowmetry and Clark microelectrodes, respectively. Vessel functionality was also assessed by in vivo confocal imaging. The vascular density and the respective contribution of human and recipient endothelium were assessed immunohistochemically by staining for human and mouse CD31. Islet grafts with EPCs had substantially higher blood perfusion and oxygen tension than control transplants. Furthermore, analysis of the vascular network of the grafts revealed that grafts containing EPC bioengineered islets had a superior vascular density compared with control grafts, with functional chimeric blood vessels. We conclude that a simple procedure of surface coating with EPCs provides a possibility to improve the vascular engraftment of transplanted human islets. Established protocols are also easily applicable for intraportal islet transplantation in order to obtain a novel directed cellular therapy at the site of implantation in the liver.
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| 20. |
- Jin, Zhe, et al.
(författare)
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GABA-mediated inhibition of human CD4+ T cell functions is enhanced by insulin but impaired by high glucose levels
- 2024
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Ingår i: EBioMedicine. - : Elsevier. - 2352-3964. ; 105
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: γ-aminobutyric acid (GABA), known as the main inhibitory neurotransmitter in the brain, exerts immunomodulatory functions by interaction with immune cells, including T cells. Metabolic programs of T cells are closely linked to their effector functions including proliferation, differentiation, and cytokine production. The physiological molecules glucose and insulin may provide environmental cues and guidance, but whether they coordinate to regulate GABA-mediated T cell immunomodulation is still being examined.METHODS: CD4+ T cells that were isolated from blood samples from healthy individuals and from patients with type 1 diabetes (T1D) were activated in vitro. We carried out metabolic assays, multiple proximity extension assay (PEA), ELISA, qPCR, immunoblotting, immunofluorescence staining, flow cytometry analysis, MS-based proteomics, as well as electrophysiology and live-cell Ca2+ imaging.FINDINGS: We demonstrate that GABA-mediated reduction of metabolic activity and the release of inflammatory proteins, including IFNγ and IL-10, were abolished in human CD4+ T cells from healthy individuals and patients with T1D when the glucose concentration was elevated above levels typically observed in healthy people. Insulin increased GABAA receptor-subunit ρ2 expression, enhanced the GABAA receptors-mediated currents and Ca2+ influx. GABA decreased, whereas insulin sustained, hexokinase activity and glycolysis in a glucose concentration-dependent manner.INTERPRETATION: These findings support that metabolic factors, such as glucose and insulin, influence the GABA-mediated immunomodulation of human primary T cells effector functions.FUNDING: The Swedish Children's Diabetes Foundation, The Swedish Diabetes Foundation, The Swedish Research Council 2018-02952, EXODIAB, The Ernfors Foundation, The Thurings Foundation and the Science for Life Laboratory.
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| 21. |
- Lauren, Ida, et al.
(författare)
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Long-term SARS-CoV-2-specific and cross-reactive cellular immune responses correlate with humoral responses, disease severity, and symptomatology
- 2022
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Ingår i: Immunity, Inflammation and Disease. - : John Wiley & Sons. - 2050-4527. ; 10:4
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Tidskriftsartikel (refereegranskat)abstract
- Background: Cellular immune memory responses post coronavirus disease 2019 (COVID-19) have been difficult to assess due to the risks of contaminating the immune response readout with memory responses stemming from previous exposure to endemic coronaviruses. The work herein presents a large-scale long-term follow-up study investigating the correlation between symptomology and cellular immune responses four to five months post seroconversion based on a unique severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific peptide pool that contains no overlapping peptides with endemic human coronaviruses. Methods: Peptide stimulated memory T cell responses were assessed with dual interferon-gamma (IFN gamma) and interleukin (IL)-2 Fluorospot. Serological analyses were performed using a multiplex antigen bead array. Results: Our work demonstrates that long-term SARS-CoV-2-specific memory T cell responses feature dual IFN gamma and IL-2 responses, whereas cross-reactive memory T cell responses primarily generate IFN gamma in response to SARS-CoV-2 peptide stimulation. T cell responses correlated to long-term humoral immune responses. Disease severity as well as specific COVID-19 symptoms correlated with the magnitude of the SARS-CoV-2-specific memory T cell response four to five months post seroconversion. Conclusion: Using a large cohort and a SARS-CoV-2-specific peptide pool we were able to substantiate that initial disease severity and symptoms correlate with the magnitude of the SARS-CoV-2-specific memory T cell responses.
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| 22. |
- Lomei, Jalal, 1980-, et al.
(författare)
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Characterization of pro-angiogenic neutrophils
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The roles of neutrophils in immune defense have been investigated for decades. These cells are well equipped to protect the body in several ways against invaders such as microorganism. Recently it has been reported that neutrophils also contribute to angiogenesis; they are recruited to the site of hypoxia where they can promote blood vessel formation, as demonstrated both in vivo and in vitro. We found that these neutrophils with proangiogenic actions form a specific subset of the circulating neutrophils. The proangiogenic neutrophils (PANs) exclusively express the adhesion molecule CD49d and vascular endothelial growth factor receptor 1 (VEGFR1), and contribute to angiogenesis by delivering MMP-9 (matrix metalloproteinase 9). In this study, PANs were compared to classic neutrophils in respect to physical features as well as functionality. We found that PANs in humans were smaller and in human and mice PANs had higher granularity compared to the classic neutrophils. Moreover, they were more efficient phagocytes than classic neutrophils. In the aortic ring model of angiogenesis, vessel neo-formation was increased by the presence of pro-angiogenic neutrophils. Finally, by using neutrophils from mice with impaired VEGFR1 receptor (Flt-1 tk-/- mice) we demonstrated the role of VEGFR1 in neutrophil recruitment towards angiogenic endothelium. Together these results show clear differences between the pro-angiogenic subpopulation and the classic neutrophils, which further solidify the conclusion of a specific neutrophil subpopulation.
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| 23. |
- Lomei, Jalal, 1980-
(författare)
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Functional characterization of pro-angiogenic neutrophils
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The vascular system stretches throughout the body to provide oxygen, nutrition, and to remove waste products from cell metabolism. Angiogenesis, formation of new blood vessels by sprouting from pre-existing vessels, is one of the main mechanisms involved in blood vessel formation. A successful angiogenic process is dependent on the timely involvement of several parameters; from different cell types to anti- and pro-angiogenic soluble factors.White blood cells are mostly famous for being involved in host defense against pathogens, through rapid reactions by innate immunity and delayed but specific responses through adaptive immunity. The neutrophils are innate immune cells, which are the most abundant white blood cells in the circulation. In addition to their classical roles in defense against invading microorganisms, it was recently revealed that neutrophils actively contribute to angiogenesis. Neutrophils that are involved in angiogenesis comprise a specific population, namely pro-angiogenic neutrophils (PANs), that are recruited to sites of hypoxia by using different adhesion molecules compared to when they chemotax towards infection.The present investigations focus on characterization of PANs and comparing them to the classic neutrophil population in terms of their physical features and their functions. By modifying and applying new in vivo and in vitro models of angiogenesis, interactions between growing endothelial cells and neutrophils have been further revealed, as well as neutrophil recruitment and movement towards an active site of angiogenesis. We found that the main neutrophil contribution to angiogenesis at site of islet transplantation occurs at early stages of revascularization to establish new vessels, where after the neutrophils leave the site. Neutrophil recruitment to a site of infection relies to a large extent on macrophage signals, but this was not the case when they were recruited to sites of hypoxia. PANs are a specific sub-population of neutrophils that significantly differ from the rest of the neutrophil population not only in terms of their active contribution to angiogenesis, but also in terms of their physical features and their phagocytosis abilities. The role of vascular endothelial growth factor receptor (VEGFR1) and also chemokine CXCL12 (CXCR4/CXCL12 signaling) in neutrophil recruitment has been further revealed by our in vitro model; neutrophil migration to sprouting endothelium is directed by CXCL12 and VEGFR1. Furthermore we found that hypoxic condition boosted pro-angiogenic activities of PANs. Moreover, how vascular permeability affects neutrophil recruitment was studied; vascular permeability regulates inflammation by increasing chemokine transport into the blood vessels and thereby promotes neutrophil recruitment.In conclusion, functional characterization of pro-angiogenic neutrophils performed in this thesis demonstrates differences beyond marker expression when compared to classic neutrophils. Moreover, intricate interactions necessary for the formation of new blood vessels between neutrophils and the growing vasculature were shown. Increased understanding of the contribution of neutrophils to blood vessel formation in hypoxic environment or/and tumors could be exploited further to develop potential therapies.
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| 24. |
- Lomei, Jalal, 1980-, et al.
(författare)
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Pro-angiogenic neutrophils are potentiated by hypoxia
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- AbstractHypoxia, shortage of oxygen in tissues, is closely related to injury, inflammation and tissue damage. One way to overcome this issue is increasing angiogenesis, growing new blood vessels from preexist-ing ones, at the site of hypoxia. Considerable number of cells, factors and signaling pathways are involved in regulating angiogenesis.Neutrophils have been detected at the site of hypoxia and it has been shown that a subpopulation of these cells, pro-angiogenic neutrophils, PANs is actively involved in increasing angiogenesis. In this study, the effect of hypoxia on PANs was studied by co-culturing PANs with growing endothelial cells using in vitro angiogenesis assay and hypoxic and normoxic incubator. Moreover, life spans of neutrophils and PANs, as well as expression of PANs specific markers have been investigated under hypoxia and normoxia. Our data shows that the ability of PANs, to induce angiogenesis was increased under hypoxic conditions. Moreover larger number of PANs survived while co-culturing with active growing endothelial cells. We thereby conclude that the hypoxic microenvironment primes pro-angiogenic neutrophils increase their pro-angiogenic ability.
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| 25. |
- Mangsbo, Sara, 1981-, et al.
(författare)
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An evaluation of a FluoroSpot assay as a diagnostic tool to determine SARS-CoV-2 specific T cell responses
- 2021
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 16:9
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Tidskriftsartikel (refereegranskat)abstract
- Numerous assays evaluating serological and cellular responses have been developed to characterize immune responses against SARS-CoV-2. Serological assays are both cost- and time-effective compared to cellular assays, but cellular immune responses may provide a diagnostic value to determine previous SARS-CoV-2 infection in seronegative individuals. However, potential cross-reactive T cell responses stemming from prior encounters with human coronaviruses (HCoVs) may affect assay specificity. In this study, we evaluated the specificity and sensitivity of a SARS-CoV-2 IFN-gamma Release Assay (IGRA) based on the FluoroSpot method employing commercially available SARS-CoV-2-specific peptide pools, as well as an in-house designed SARS-CoV-2 peptide pool restricted to 5 amino acid stretches or less aligning with endemic HCoVs. Blood samples were obtained from healthcare workers (HCW) 5-6 months post SARS-CoV-2 spike (S) IgG and nucleocapsid (N) IgG dual seroconversion (n = 187) and HCW who had been S IgG and N IgG dual seronegative at repeated occasions, including the current sampling time point (n = 102). In addition, samples were obtained 4 to 5 months post infection from 55 polymerase chain reaction (PCR)-confirmed COVID-19 patients. Assay specificity and sensitivity were calculated with serology as a reference standard for HCW. The in-house generated peptide pool displayed a specificity of 96.1%, while the commercially available peptide pools displayed specificities of 80.4% and 85.3%, respectively. Sensitivity was higher in a cohort of previously hospitalized COVID-19 patients (96.4% and 84.0% for the commercially available peptide pools and 92.7% for the in-house generated peptide pool) compared to the HCW cohort (92.0% and 66.8% for the commercially available peptide pools and 76.0% for the in-house generated peptide pool). Based on these findings, the individual diagnostic value of T cell immune responses against SARS-CoV-2 currently appears to be limited but remain an important research tool ahead.
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| 26. |
- Massena, Sara, et al.
(författare)
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A chemotactic gradient sequestered on endothelial heparan sulfate induces directional intraluminal crawling of neutrophils
- 2010
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Ingår i: Blood. - : American Society of Hematology. - 0006-4971 .- 1528-0020. ; 116:11, s. 1924-1931
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Tidskriftsartikel (refereegranskat)abstract
- During infection, chemokines sequestered on endothelium induce recruitment of circulating leukocytes into the tissue where they chemotax along chemokine gradients toward the afflicted site. The aim of this in vivo study was to determine whether a chemokine gradient was formed intravascularly and influenced intraluminal neutrophil crawling and transmigration. A chemokine gradient was induced by placing a macrophage inflammatory protein-2 (MIP-2)-containing (CXCL2) gel on the cremaster muscle of anesthetized wild-type mice or heparanase-overexpressing transgenic mice (hpa-tg) with truncated heparan sulfate (HS) side chains. Neutrophil-endothelial interactions were visualized by intravital microscopy and chemokine gradients detected by confocal microscopy. Localized extravascular chemokine release (MIP-2 gel) induced directed neutrophil crawling along a chemotactic gradient immobilized on the endothelium and accelerated their recruitment into the target tissue compared with homogeneous extravascular chemokine concentration (MIP-2 super-fusion). Endothelial chemokine sequestration occurred exclusively in venules and was HS-dependent, and neutrophils in hpa-tg mice exhibited random crawling. Despite similar numbers of adherent neutrophils in hpa-tg and wild-type mice, the altered crawling in hpa-tg mice was translated into decreased number of emigrated neutrophils and ultimately decreased the ability to clear bacterial infections. In conclusion, an intravascular chemokine gradient sequestered by endothelial HS effectively directs crawling leukocytes toward transmigration loci close to the infection site.
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| 27. |
- Massena, Sara, et al.
(författare)
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Identification and characterization of VEGF-A-responsive neutrophils expressing CD49d, VEGFR1, and CXCR4 in mice and humans
- 2015
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Ingår i: Blood. - : American Society of Hematology. - 0006-4971 .- 1528-0020. ; 126:17, s. 2016-2026
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Tidskriftsartikel (refereegranskat)abstract
- Vascular endothelial growth factor A (VEGF-A) is upregulated during hypoxia and is the major regulator of angiogenesis. VEGF-A expression has also been found to recruit myeloid cells to ischemic tissues where they contribute to angiogenesis. This study investigates the mechanisms underlying neutrophil recruitment to VEGF-A as well as the characteristics of these neutrophils. A previously undefined circulating subset of neutrophils shown to be CD49d(+)VEGFR1(high)CXCR4(high) was identified in mice and humans. By using chimeric mice with impaired VEGF receptor 1 (VEGFR1) or VEGFR2 signaling (Flt-1tk(-/-), tsad(-/-)), we found that parallel activation of VEGFR1 on neutrophils and VEGFR2 on endothelial cells was required for VEGF-A-induced recruitment of circulating neutrophils to tissue. Intravital microscopy of mouse microcirculation revealed that neutrophil recruitment by VEGF-A versus by the chemokine macrophage inflammatory protein 2 (MIP-2 [CXCL2]) involved the same steps of the recruitment cascade but that an additional neutrophil integrin (eg, VLA-4 [CD49d/CD29]) played a crucial role in neutrophil crawling and emigration to VEGF-A. Isolated CD49d(+) neutrophils featured increased chemokinesis but not chemotaxis compared with CD49d(-) neutrophils in the presence of VEGF-A. Finally, by targeting the integrin α4 subunit (CD49d) in a transplantation-based angiogenesis model that used avascular pancreatic islets transplanted to striated muscle, we demonstrated that inhibiting the recruitment of circulating proangiogenic neutrophils to hypoxic tissue impairs vessel neoformation. Thus, angiogenesis can be modulated by targeting cell-surface receptors specifically involved in VEGF-A-dependent recruitment of proangiogenic neutrophils without compromising recruitment of the neutrophil population involved in the immune response to pathogens.
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| 28. |
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| 29. |
- Muntjewerff, Elke M., et al.
(författare)
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Chromogranin A regulates gut permeability via the antagonistic actions of its proteolytic peptides
- 2021
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Ingår i: Acta Physiologica. - : John Wiley & Sons. - 1748-1708 .- 1748-1716. ; 232:2
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Tidskriftsartikel (refereegranskat)abstract
- Aim A "leaky" gut barrier has been implicated in the initiation and progression of a multitude of diseases, for example, inflammatory bowel disease (IBD), irritable bowel syndrome and celiac disease. Here we show how pro-hormone Chromogranin A (CgA), produced by the enteroendocrine cells, and Catestatin (CST: hCgA(352-372)), the most abundant CgA-derived proteolytic peptide, affect the gut barrier. Methods Colon tissues from region-specific CST-knockout (CST-KO) mice, CgA-knockout (CgA-KO) and WT mice were analysed by immunohistochemistry, western blot, ultrastructural and flowcytometry studies. FITC-dextran assays were used to measure intestinal barrier function. Mice were supplemented with CST or CgA fragment pancreastatin (PST: CgA(250-301)). The microbial composition of cecum was determined. CgA and CST levels were measured in blood of IBD patients. Results Plasma levels of CST were elevated in IBD patients. CST-KO mice displayed (a) elongated tight, adherens junctions and desmosomes similar to IBD patients, (b) elevated expression of Claudin 2, and (c) gut inflammation. Plasma FITC-dextran measurements showed increased intestinal paracellular permeability in the CST-KO mice. This correlated with a higher ratio of Firmicutes to Bacteroidetes, a dysbiotic pattern commonly encountered in various diseases. Supplementation of CST-KO mice with recombinant CST restored paracellular permeability and reversed inflammation, whereas CgA-KO mice supplementation with CST and/or PST in CgA-KO mice showed that intestinal paracellular permeability is regulated by the antagonistic roles of these two peptides: CST reduces and PST increases permeability. Conclusion The pro-hormone CgA regulates the intestinal paracellular permeability. CST is both necessary and sufficient to reduce permeability and primarily acts by antagonizing PST.
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| 30. |
- Muntjewerff, Elke M., et al.
(författare)
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Putative regulation of macrophage-mediated inflammation by catestatin
- 2022
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Ingår i: Trends in immunology. - : Elsevier. - 1471-4906 .- 1471-4981. ; 43:1, s. 41-50
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Forskningsöversikt (refereegranskat)abstract
- Catestatin (CST) is a bioactive cleavage product of the neuroendocrine prohormone chromogranin A (CgA). Recent findings show that CST can exert anti-inflammatory and antiadrenergic effects by suppressing the inflammatory actions of mammalian macrophages. However, recent findings also suggest that macrophages themselves are major CST producers. Here, we hypothesize that macrophages produce CST in an inflammation-dependent manner and thereby might self-regulate inflammation in an autocrine fashion. CST is associated with pathological conditions hallmarked by chronic inflammation, including autoimmune, cardiovascular, and metabolic disorders. Since intraperitoneal injection of CST in mouse models of diabetes and inflammatory bowel disease has been reported to be beneficial formitigating disease, we posit that CST should be further investigated as a candidate target for treating certain inflammatory diseases.
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| 31. |
- Muntjewerff, Elke M., et al.
(författare)
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The anti-inflammatory peptide Catestatin blocks chemotaxis
- 2022
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Ingår i: Journal of Leukocyte Biology. - : John Wiley & Sons. - 0741-5400 .- 1938-3673. ; 112:2, s. 273-278
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Tidskriftsartikel (refereegranskat)abstract
- Increased levels of the anti-inflammatory peptide Catestatin (CST), a cleavage product of the pro-hormone chromogranin A, correlate with less severe outcomes in hypertension, colitis, and diabetes. However, it is unknown how CST reduces the infiltration of monocytes and macrophages (M phi s) in inflamed tissues. Here, it is reported that CST blocks leukocyte migration toward inflammatory chemokines. By in vitro and in vivo migration assays, it is shown that although CST itself is chemotactic, it blocks migration of monocytes and neutrophils to inflammatory attracting factor CC-chemokine ligand 2 (CCL2) and C-X-C motif chemokine ligand 2 (CXCL2). Moreover, it directs CX(3)CR1(+) M phi s away from pancreatic islets. These findings suggest that the anti-inflammatory actions of CST are partly caused by its regulation of chemotaxis.
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| 32. |
- Muntjewerff, Elke M., et al.
(författare)
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Three-dimensional Co-culture Model for Live Imaging of Pancreatic Islets, Immune Cells, and Neurons in Agarose Gel
- 2023
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Ingår i: Bio-protocol. - : Bio-Protocol LLC. - 2331-8325. ; 13:20
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Tidskriftsartikel (refereegranskat)abstract
- During the onset of autoimmune diabetes, nerve-immune cell interactions seem to play an important role; however, there are currently no models to follow and interfere with these interactions over time in vivo or in vitro. Twodimensional in vitro models provide insufficient information and microfluidics or organs on a chip are usually challenging to work with. We present here what we believe to be the first simple model that provides the opportunity to co-culture pancreatic islets with sympathetic nerves and immune cells. This model is based on our stamping device that can be 3D printed (STL file provided). Due to the imprint in the agarose gel, sympathetic neurons, pancreatic islets, and macrophages can be seeded in specific locations at a level that allows for confocal live-cell imaging. In this protocol, we provide the instructions to construct and perform live cell imaging experiments in our co-culture model, including: 1) design for the stamping device to make the imprint in the gel, 2) isolation of sympathetic neurons, pancreatic islets, and macrophages, 3) co-culture conditions, 4) how this can be used for live cell imaging, and 5) possibilities for wider use of the model. In summary, we developed an easy-to-use co-culture model that allows manipulation and imaging of interactions between sympathetic nerves, pancreatic islets, and macrophages. This new co-culture model is useful to study nerve-immune cell-islet interactions and will help to identify the functional relevance of neuro-immune interactions in the pancreas.
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| 33. |
- Nezhyva, Mariya
(författare)
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Integrative Approaches in Shotgun Proteomics : From sample preparation to multifaceted data analysis
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Bottom-up proteomics mass-spectrometry gives an opportunity to shed light on var-ious biological aspects/characteristics such as protein composition, its modifications,interactions, and dynamics. Its applications span a wide range of biological sciencesaiming to eventually answer fundamental disease related questions or evaluate drugperformance and predict adverse effects.Bottom-up proteomics experiments are worth the time and effort because it is a comprehensive and multi-level approach to address research questions from a single dataset. Hence, we can gain a thorough understanding of the processes in the studied system and their interconnected changes, thereby confirming results from different perspectives. While the majority of research is focused on the improvement of analytical techniques, it remains challenging to derive meaningful biological insights from the data.This Ph.D. thesis contributes to various proteomics challenges, including the development of a high-throughput micro-scale proteomic workflow and comprehensive multifaceted analysis. In particular, the data on the thermal proteome profiling of melanocyte-stimulating hormone interactions with melanocortin receptors was analyzed using a newly developed workflow. This workflow uniquely combines thermal stabilization data, inferred transcription factor activity, and pathway analysis. Additionally, this thesis integrates omics approaches such as metabolomics and proteomics. Another dimension to the dataset was added by combining metabolic mass spectrometry imaging with region-specific proteomics for the characterization of cocaine’s neurotoxicity.
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| 34. |
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| 35. |
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| 36. |
- Parv, Kristel
(författare)
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Macrophages and neutrophils in tissue homeostasis and recovery from ischemic injury
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Neutrophils and macrophages have functions beyond protection against pathogens. The overall aim of the work presented in this thesis was to identify novel tasks for these innate immune cells in maintaining homeostasis. In the studies presented here, we explored macrophage roles in tissue recovery from ischemic injury and post-natal tissue development, and the origin and recruitment mechanisms of pro-angiogenic neutrophils (PANs) to the site of ischemic injury.In Study I, it was shown that perivascular macrophages at sites of ischemic injury adopt characteristics of mural cell identity. Combining genetic heritable fate mapping of macrophages with single-cell RNA-sequencing, we were able to demonstrate that macrophages downregulated the expression of myeloid markers, and upregulated those of mural cells. Depletion of macrophages during tissue healing demonstrated that macrophages also adopt important mural cell functions that are crucial for blood vessel maturation after ischemic injury. In Study II, it was shown that perivascular macrophages form cuff-like structures around vessels in the ischemic muscle following ischemia. Using genetically modified mouse models, we showed that these macrophages regulate bloodflow in an inducible NO Synthase (iNOS)-dependent manner, and this could be therapeutically targeted to improve tissue healing through local delivery of plasmid-encoded C-X-C Motif Chemokine Ligand 12 (CXCL12). In Study III, it was shown that recruitment of PANs to the site of ischemic injury is dependent on CD49d signalling, and the spleen contains a peripheral reservoir of PANs which is crucial for achieving adequate accumulation of PANs at the site of ischemic injury. We also showed that the release of splenic PANs in response to ischemic injury relies on sympathetic signalling and downregulation of CXCL12α in the splenic red pulp.In Study IV, it was shown that pancreatic tissue-resident macrophages are important for post-natal islet development, as depletion of macrophages using clodronate liposomes led to impaired glucose tolerance in adult mice. Further, neonatal infection with S. aureus led to reduced number of pancreatic macrophages and interfered with normal post-natal β cell development, leading to impaired glucose tolerance. In summary, the work presented here expands our understanding on the various roles of macrophages during ischemic injury and tissue development, and significantly advances our understanding of the origins and recruitment mechanisms of pro-angiogenic neutrophils during ischemic injury.
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| 37. |
- Parv, Kristel, et al.
(författare)
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Phagocytosis and Efferocytosis by Resident Macrophages in the Mouse Pancreas
- 2021
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Ingår i: Frontiers in Endocrinology. - : Frontiers Media S.A.. - 1664-2392. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- The tissue microenvironment in the mouse pancreas has been shown to promote very different polarizations of resident macrophages with islet-resident macrophages displaying an inflammatory “M1” profile and macrophages in the exocrine tissue mostly displaying an alternatively activated “M2” profile. The impact of this polarization on tissue homeostasis and diabetes development is unclear. In this study, the ability of pancreas-resident macrophages to phagocyte bacterial and endogenous debris was investigated. Mouse endocrine and exocrine tissues were separated, and tissue-resident macrophages were isolated by magnetic immunolabeling. Isolated macrophages were subjected to flow cytometry for polarization markers and qPCR for phagocytosis-related genes. Functional in vitro investigations included phagocytosis and efferocytosis assays using pH-sensitive fluorescent bacterial particles and dead fluorescent neutrophils, respectively. Intravital confocal imaging of in situ phagocytosis and efferocytosis in the pancreas was used to confirm findings in vivo. Gene expression analysis revealed no significant overall difference in expression of most phagocytosis-related genes in islet-resident vs. exocrine-resident macrophages included in the analysis. In this study, pancreas-resident macrophages were shown to differ in their ability to phagocyte bacterial and endogenous debris depending on their microenvironment. This difference in abilities may be one of the factors polarizing islet-resident macrophages to an inflammatory state since phagocytosis has been found to imprint macrophage heterogeneity. It remains unclear if this difference has any implications in the development of islet dysfunction or autoimmunity.
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| 38. |
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| 39. |
- Petersson, Joel, et al.
(författare)
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Gastroprotective and blood pressure lowering effects of dietary nitrate are abolished by an antiseptic mouthwash
- 2009
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Ingår i: Free Radical Biology & Medicine. - : Elsevier BV. - 0891-5849 .- 1873-4596. ; 46:8, s. 1068-1075
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Tidskriftsartikel (refereegranskat)abstract
- Recently, it has been suggested that the supposedly inert nitrite anion is reduced in vivo to form bioactive nitric oxide with physiological and therapeutic implications in the gastrointestinal and cardiovascular systems. Intake of nitrate-rich food such as vegetables results in increased levels of circulating nitrite in a process suggested to involve nitrate-reducing bacteria in the oral cavity. Here we investigated the importance of the oral microflora and dietary nitrate in regulation of gastric mucosal defense and blood pressure. Rats were treated twice daily with a commercial antiseptic mouthwash while they were given nitrate-supplemented drinking water. The mouthwash greatly reduced the number of nitrate-reducing oral bacteria and as a consequence, nitrate-induced increases in gastric NO and circulating nitrite levels were markedly reduced. With the mouthwash the observed nitrate-induced increase in gastric mucus thickness was attenuated and the gastroprotective effect against an ulcerogenic compound was lost. Furthermore, the decrease in systemic blood pressure seen during nitrate supplementation was now absent. These results suggest that oral symbiotic bacteria modulate gastrointestinal and cardiovascular function via bioactivation of salivary nitrate. Excessive use of antiseptic mouthwashes may attenuate the bioactivity of dietary nitrate.
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| 40. |
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| 41. |
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| 42. |
- Pettersson, Ulrika Sofia, et al.
(författare)
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Increased Recruitment but Impaired Function of Leukocytes during Inflammation in Mouse Models of Type 1 and Type 2 Diabetes
- 2011
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:7, s. e22480-
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundPatients suffering from diabetes show defective bacterial clearance. This study investigates the effects of elevated plasma glucose levels during diabetes on leukocyte recruitment and function in established models of inflammation.Methodology/Principal FindingsDiabetes was induced in C57Bl/6 mice by intravenous alloxan (causing severe hyperglycemia), or by high fat diet (moderate hyperglycemia). Leukocyte recruitment was studied in anaesthetized mice using intravital microscopy of exposed cremaster muscles, where numbers of rolling, adherent and emigrated leukocytes were quantified before and during exposure to the inflammatory chemokine MIP-2 (0.5 nM). During basal conditions, prior to addition of chemokine, the adherent and emigrated leukocytes were increased in both alloxan- (62±18% and 85±21%, respectively) and high fat diet-induced (77±25% and 86±17%, respectively) diabetes compared to control mice. MIP-2 induced leukocyte emigration in all groups, albeit significantly more cells emigrated in alloxan-treated mice (15.3±1.0) compared to control (8.0±1.1) mice. Bacterial clearance was followed for 10 days after subcutaneous injection of bioluminescent S. aureus using non-invasive IVIS imaging, and the inflammatory response was assessed by Myeloperoxidase-ELISA and confocal imaging. The phagocytic ability of leukocytes was assessed using LPS-coated fluorescent beads and flow cytometry. Despite efficient leukocyte recruitment, alloxan-treated mice demonstrated an impaired ability to clear bacterial infection, which we found correlated to a 50% decreased phagocytic ability of leukocytes in diabetic mice.Conclusions/SignificanceThese results indicate that reduced ability to clear bacterial infections observed during experimentally induced diabetes is not due to reduced leukocyte recruitment since sustained hyperglycemia results in increased levels of adherent and emigrated leukocytes in mouse models of type 1 and type 2 diabetes. Instead, decreased phagocytic ability observed for leukocytes isolated from diabetic mice might account for the impaired bacterial clearance.
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| 43. |
- Rodriguez-Calvo, Teresa, et al.
(författare)
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Means, Motive, and Opportunity : Do Non-Islet-Reactive Infiltrating T Cells Contribute to Autoimmunity in Type 1 Diabetes?
- 2021
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Ingår i: Frontiers in Immunology. - : Frontiers Media S.A.. - 1664-3224. ; 12
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Forskningsöversikt (refereegranskat)abstract
- In human type 1 diabetes and animal models of the disease, a diverse assortment of immune cells infiltrates the pancreatic islets. CD8+ T cells are well represented within infiltrates and HLA multimer staining of pancreas sections provides clear evidence that islet epitope reactive T cells are present within autoimmune lesions. These bona fide effectors have been a key research focus because these cells represent an intellectually attractive culprit for β cell destruction. However, T cell receptors are highly diverse in human insulitis. This suggests correspondingly broad antigen specificity, which includes a majority of T cells for which there is no evidence of islet-specific reactivity. The presence of “non-cognate” T cells in insulitis raises suspicion that their role could be beyond that of an innocent bystander. In this perspective, we consider the potential pathogenic contribution of non-islet-reactive T cells. Our intellectual framework will be that of a criminal investigation. Having arraigned islet-specific CD8+ T cells for the murder of pancreatic β cells, we then turn our attention to the non-target immune cells present in human insulitis and consider the possible regulatory, benign, or effector roles that they may play in disease. Considering available evidence, we overview the case that can be made that non-islet-reactive infiltrating T cells should be suspected as co-conspirators or accessories to the crime and suggest some possible routes forward for reaching a better understanding of their role in disease.
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| 44. |
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| 45. |
- Trolle, Carl, 1985-, et al.
(författare)
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Boundary cap neural crest stem cells promote angiogenesis after transplantation to avulsed dorsal roots in mice and induce migration of endothelial cells in 3D printed scaffolds
- 2024
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Ingår i: Neuroscience Letters. - : Elsevier. - 0304-3940 .- 1872-7972. ; 826
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Tidskriftsartikel (refereegranskat)abstract
- Dorsal root avulsion injuries lead to loss of sensation and to reorganization of blood vessels (BVs) in the injured area. The inability of injured sensory axons to re-enter the spinal cord results in permanent loss of sensation, and often also leads to the development of neuropathic pain. Approaches that restore connection between peripheral sensory axons and their CNS targets are thus urgently need. Previous research has shown that sensory axons from peripherally grafted human sensory neurons are able to enter the spinal cord by growing along BVs which penetrate the CNS from the spinal cord surface. In this study we analysed the distribution of BVs after avulsion injury and how their pattern is affected by implantation at the injury site of boundary cap neural crest stem cells (bNCSCs), a transient cluster of cells, which are located at the boundary between the spinal cord and peripheral nervous system and assist the growth of sensory axons from periphery into the spinal cord during development. The superficial dorsal spinal cord vasculature was examined using intravital microscopy and intravascular BV labelling. bNCSC transplantation increased vascular volume in a non-dose responsive manner, whereas dorsal root avulsion alone did not decrease the vascular volume. To determine whether bNCSC are endowed with angiogenic properties we prepared 3D printed scaffolds, containing bNCSCs together with rings prepared from mouse aorta. We show that bNCSC do induce migration and assembly of endothelial cells in this system. These findings suggest that bNCSC transplant can promote vascularization in vivo and contribute to BV formation in 3D printed scaffolds.
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| 46. |
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| 47. |
- von Herrath, Matthias, et al.
(författare)
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Case Reports of Pre-clinical Replication Studies in Metabolism and Diabetes
- 2019
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Ingår i: Cell Metabolism. - : CELL PRESS. - 1550-4131 .- 1932-7420. ; 29:4, s. 795-802
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Forskningsöversikt (refereegranskat)abstract
- Recent articles have highlighted the lack of reproducibility of data from scientific publications. Here we would argue that a better way to describe and also tackle this matter is to use the term "lack of robustness," since it points toward potential solutions. Presenting several case reports, we highlight examples with common underlying issues from Novo Nordisk's experience: animal model variability, reagent quality, and inter-lao variability. We discuss means to prevent these issues and argue for increased collaborative work and transparent manuscript revision procedures. Collectively, we believe these measures will help promote a more rapid and efficient self-corrective process in diabetes drug target research.
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| 48. |
- Vyakaranam, Achyut Ram, et al.
(författare)
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Non-invasive imaging of sympathetic innervation of the pancreas in individuals with type 2 diabetes
- 2024
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Ingår i: Diabetologia. - : Springer. - 0012-186X .- 1432-0428. ; 67:1, s. 199-208
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Tidskriftsartikel (refereegranskat)abstract
- Aims/hypothesisCompromised pancreatic sympathetic innervation has been suggested as a factor involved in both immune-mediated beta cell destruction and endocrine dysregulation of pancreatic islets. To further explore these intriguing findings, new techniques for in vivo assessment of pancreatic innervation are required. This is a retrospective study that aimed to investigate whether the noradrenaline (norepinephrine) analogue 11C-hydroxy ephedrine (11C-HED) could be used for quantitative positron emission tomography (PET) imaging of the sympathetic innervation of the human pancreas.MethodsIn 25 individuals with type 2 diabetes and 64 individuals without diabetes, all of whom had previously undergone 11C-HED-PET/CT because of pheochromocytoma or paraganglioma (or suspicion thereof), the 11C-HED standardised uptake value (SUVmean), 11C-HED specific binding index (SBI), pancreatic functional volume (FV, in ml), functional neuronal volume (FNV, calculated as SUVmean × FV), specific binding index with functional volume (SBI FV, calculated as SBI × FV) and attenuation on CT (HU) were investigated in the entire pancreas, and additionally in six separate anatomical pancreatic regions.ResultsGenerally, 11C-HED uptake in the pancreas was high, with marked individual variation, suggesting variability in sympathetic innervation. Moreover, pancreatic CT attenuation (HU) (p<0.001), 11C-HED SBI (p=0.0049) and SBI FV (p=0.0142) were lower in individuals with type 2 diabetes than in individuals without diabetes, whereas 11C-HED SUVmean (p=0.15), FV (p=0.73) and FNV (p=0.30) were similar.Conclusions/interpretationWe demonstrate the feasibility of using 11C-HED-PET for non-invasive assessment of pancreatic sympathetic innervation in humans. These findings warrant further prospective evaluation, especially in individuals with theoretical defects in pancreatic sympathetic innervation, such as those with type 1 diabetes.
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| 49. |
- Vågesjö, Evelina, et al.
(författare)
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Immunological Shielding by Induced Recruitment of Regulatory T-Lymphocytes Delays Rejection of Islets Transplanted in Muscle
- 2015
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Ingår i: Cell Transplantation. - 0963-6897 .- 1555-3892. ; 24:2, s. 263-276
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Tidskriftsartikel (refereegranskat)abstract
- The only clinically available curative treatment of type 1 diabetes mellitus is replacement of the pancreatic islets by allogeneic transplantation, which requires immunosuppressive therapies. Regimens used today are associated with serious adverse effects and impaired islet engraftment and function. The aim of the current study was to induce local immune privilege by accumulating immune-suppressive regulatory T-lymphocytes (Tregs) at the site of intramuscular islet transplantation to reduce the need of irnmunosuppressive therapy during engraftment. Islets were cotransplanted with a plasmid encoding the chemokine CCL22 into the muscle of MHC-mismatched mice, after which pCCL22 expression and leukocyte recruitment were studied in parallel with graft functionality. Myocyte pCCL22 expression and secretion resulted in local accumulation of Tregs. When islets were cotransplanted with pCCL22, significantly fewer effector T-lymphocytes were observed in close proximity to the islets, leading to delayed graft rejection. As a result, diabetic recipients cotransplanted with islets and pCCL22 intramuscularly became normoglycemic for 10 consecutive days, while grafts cotransplanted with control plasmid were rejected immediately, leaving recipients severely hyperglycemic. Here we propose a simple method to initially shield MHC-mismatched islets by the recruitment of endogenous Tregs during engraftment in order to improve early islet survival. Using this approach, the very high doses of systemic immunosuppression used initially following transplantation can thereby be avoided.
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| 50. |
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