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DAMP production by human islets under low oxygen and nutrients in the presence or absence of an immunoisolating-capsule and necrostatin-1.

Paredes-Juarez GA, Sahasrabudhe NM, Tjoelker RS, de Haan BJ, Engelse MA, de Koning EJ, Faas MM, de Vos P - Sci Rep (2015)

Bottom Line: Immunoisolation in immunoprotective membranes reduced DAMP release and immune activation via retention of the relative large DAMPs in the capsules.Another effective strategy was suppressing necroptosis using the inhibitor nec-1.DAMP release can be reduced in vitro by immunoisolation or intervention with nec-1.

View Article: PubMed Central - PubMed

Affiliation: University of Groningen, University Medical Center Groningen, Department of Pathology and Medical Biology, Section of Immunoendocrinology, Groningen, 9713 GZ, The Netherlands.

ABSTRACT
In between the period of transplantation and revascularization, pancreatic islets are exposed to low-oxygen and low-nutrient conditions. In the present study we mimicked those conditions in vitro to study the involvement of different cell death processes, release of danger-associated molecular patterns (DAMP), and associated in vitro immune activation. Under low-oxygen and low-nutrient conditions, apoptosis, autophagy and necroptosis occur in human islets. Necroptosis is responsible for DAMP-release such as dsDNA, uric acid, and HMGB1. The sensors of the innate immune system able to recognize these DAMPs are mainly TLR, NOD receptors, and C-type lectins. By using cell-lines with a non-functional adaptor molecule MyD88, we were able to show that the islet-derived DAMPs signal mainly via TLR. Immunoisolation in immunoprotective membranes reduced DAMP release and immune activation via retention of the relative large DAMPs in the capsules. Another effective strategy was suppressing necroptosis using the inhibitor nec-1. Although the effect on cell-survival was minor, nec-1 was able to reduce the release of HMGB1 and its associated immune activation. Our data demonstrate that in the immediate post-transplant period islets release DAMPs that in vitro enhance responses of innate immune cells. DAMP release can be reduced in vitro by immunoisolation or intervention with nec-1.

No MeSH data available.


Related in: MedlinePlus

Reduction of danger-associated molecular patterns (DAMPs) from immunoisolated islets in alginate-based microcapsules.Supernatant of encapsulated pancreatic human islets incubated for 1 (a,c) and 7 (b,d) days under control and low nutrients conditions at 20% and 1% of oxygen. DAMPs found were double stranded DNA (dsDNA; (a,b)), and uric acid (c,d). High mobility group protein B1 (HMGB1) was not found in encapsulated islets and therefore not shown in the graph. Values are presented as median ± IQR (n = 4 separate batches of human islets). a p < 0.05 was consider statistical significant (*p < 0.05; **p < 0.01).
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f5: Reduction of danger-associated molecular patterns (DAMPs) from immunoisolated islets in alginate-based microcapsules.Supernatant of encapsulated pancreatic human islets incubated for 1 (a,c) and 7 (b,d) days under control and low nutrients conditions at 20% and 1% of oxygen. DAMPs found were double stranded DNA (dsDNA; (a,b)), and uric acid (c,d). High mobility group protein B1 (HMGB1) was not found in encapsulated islets and therefore not shown in the graph. Values are presented as median ± IQR (n = 4 separate batches of human islets). a p < 0.05 was consider statistical significant (*p < 0.05; **p < 0.01).

Mentions: Immunoisolation as a mean to prevent rejection of islets is gaining much attention by the scientific community79. DAMPs are relatively large structures that should be retained by the membranes. To confirm this, human islets were encapsulated in alginate-PLL-alginate (APA) capsules and cultured under the same low-nutrient and hypoxic conditions. Supernatant was subsequently incubated with THP1-XBlue™-MD2-CD14. The induced responses were up to tenfold lower when compared to free islets after 1 day of incubation under all the different conditions (supplemental Fig. S2) indicating that encapsulation prevents the release of some DAMPs. To confirm that, we quantified the release of DAMPs by encapsulated islets after incubation. Interestingly, HMGB1 was absent under all incubation conditions. We detected dsDNA under low (Fig. 5a) or normal nutrient conditions (Fig. 5b), and under hypoxia or normoxia, with no statistical significant differences. Uric acid was statistically significant increased (p < 0.05) on day 1 and 7 under all the conditions with low nutrient availability (Fig. 5c,d).


DAMP production by human islets under low oxygen and nutrients in the presence or absence of an immunoisolating-capsule and necrostatin-1.

Paredes-Juarez GA, Sahasrabudhe NM, Tjoelker RS, de Haan BJ, Engelse MA, de Koning EJ, Faas MM, de Vos P - Sci Rep (2015)

Reduction of danger-associated molecular patterns (DAMPs) from immunoisolated islets in alginate-based microcapsules.Supernatant of encapsulated pancreatic human islets incubated for 1 (a,c) and 7 (b,d) days under control and low nutrients conditions at 20% and 1% of oxygen. DAMPs found were double stranded DNA (dsDNA; (a,b)), and uric acid (c,d). High mobility group protein B1 (HMGB1) was not found in encapsulated islets and therefore not shown in the graph. Values are presented as median ± IQR (n = 4 separate batches of human islets). a p < 0.05 was consider statistical significant (*p < 0.05; **p < 0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4588515&req=5

f5: Reduction of danger-associated molecular patterns (DAMPs) from immunoisolated islets in alginate-based microcapsules.Supernatant of encapsulated pancreatic human islets incubated for 1 (a,c) and 7 (b,d) days under control and low nutrients conditions at 20% and 1% of oxygen. DAMPs found were double stranded DNA (dsDNA; (a,b)), and uric acid (c,d). High mobility group protein B1 (HMGB1) was not found in encapsulated islets and therefore not shown in the graph. Values are presented as median ± IQR (n = 4 separate batches of human islets). a p < 0.05 was consider statistical significant (*p < 0.05; **p < 0.01).
Mentions: Immunoisolation as a mean to prevent rejection of islets is gaining much attention by the scientific community79. DAMPs are relatively large structures that should be retained by the membranes. To confirm this, human islets were encapsulated in alginate-PLL-alginate (APA) capsules and cultured under the same low-nutrient and hypoxic conditions. Supernatant was subsequently incubated with THP1-XBlue™-MD2-CD14. The induced responses were up to tenfold lower when compared to free islets after 1 day of incubation under all the different conditions (supplemental Fig. S2) indicating that encapsulation prevents the release of some DAMPs. To confirm that, we quantified the release of DAMPs by encapsulated islets after incubation. Interestingly, HMGB1 was absent under all incubation conditions. We detected dsDNA under low (Fig. 5a) or normal nutrient conditions (Fig. 5b), and under hypoxia or normoxia, with no statistical significant differences. Uric acid was statistically significant increased (p < 0.05) on day 1 and 7 under all the conditions with low nutrient availability (Fig. 5c,d).

Bottom Line: Immunoisolation in immunoprotective membranes reduced DAMP release and immune activation via retention of the relative large DAMPs in the capsules.Another effective strategy was suppressing necroptosis using the inhibitor nec-1.DAMP release can be reduced in vitro by immunoisolation or intervention with nec-1.

View Article: PubMed Central - PubMed

Affiliation: University of Groningen, University Medical Center Groningen, Department of Pathology and Medical Biology, Section of Immunoendocrinology, Groningen, 9713 GZ, The Netherlands.

ABSTRACT
In between the period of transplantation and revascularization, pancreatic islets are exposed to low-oxygen and low-nutrient conditions. In the present study we mimicked those conditions in vitro to study the involvement of different cell death processes, release of danger-associated molecular patterns (DAMP), and associated in vitro immune activation. Under low-oxygen and low-nutrient conditions, apoptosis, autophagy and necroptosis occur in human islets. Necroptosis is responsible for DAMP-release such as dsDNA, uric acid, and HMGB1. The sensors of the innate immune system able to recognize these DAMPs are mainly TLR, NOD receptors, and C-type lectins. By using cell-lines with a non-functional adaptor molecule MyD88, we were able to show that the islet-derived DAMPs signal mainly via TLR. Immunoisolation in immunoprotective membranes reduced DAMP release and immune activation via retention of the relative large DAMPs in the capsules. Another effective strategy was suppressing necroptosis using the inhibitor nec-1. Although the effect on cell-survival was minor, nec-1 was able to reduce the release of HMGB1 and its associated immune activation. Our data demonstrate that in the immediate post-transplant period islets release DAMPs that in vitro enhance responses of innate immune cells. DAMP release can be reduced in vitro by immunoisolation or intervention with nec-1.

No MeSH data available.


Related in: MedlinePlus