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Hypoxia inducible factor (HIF)-1 coordinates induction of Toll-like receptors TLR2 and TLR6 during hypoxia.

Kuhlicke J, Frick JS, Morote-Garcia JC, Rosenberger P, Eltzschig HK - PLoS ONE (2007)

Bottom Line: During acute infection and inflammation, dramatic shifts in tissue metabolism are typical, thereby resulting in profound tissue hypoxia.While transcript levels of other TLRs remained unchanged, we found a robust induction of TLR2 (2.36+/-0.7-fold; P<0.05) and TLR6 (3.46+/-1.56-fold; P<0.05).Furthermore, analysis of the putative TLR2 and TLR6 promoters revealed previously unrecognized binding sites for HIF-1, which were shown by chromatin immunoprecipitation to bind the pivotal hypoxia-regulating transcription factor HIF-1alpha.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology and Intensive Care Medicine, Tübingen University Hospital, Tübingen, Germany.

ABSTRACT

Background: During acute infection and inflammation, dramatic shifts in tissue metabolism are typical, thereby resulting in profound tissue hypoxia. Therefore, we pursued the hypothesis, that tissue hypoxia may influence innate immune responses by transcriptional modulation of Toll-like receptor (TLRs) expression and function.

Methodology/principal findings: We gained first insight from transcriptional profiling of murine dendritic cells exposed to hypoxia (2% oxygen for 24 h). While transcript levels of other TLRs remained unchanged, we found a robust induction of TLR2 (2.36+/-0.7-fold; P<0.05) and TLR6 (3.46+/-1.56-fold; P<0.05). Additional studies in different cells types and cell-lines including human dendritic cells, monocytic cells (MM6), endothelia (HMEC-1) or intestinal epithelia (Caco-2) confirmed TLR2 and TLR6 induction of transcript, protein and function during hypoxia. Furthermore, analysis of the putative TLR2 and TLR6 promoters revealed previously unrecognized binding sites for HIF-1, which were shown by chromatin immunoprecipitation to bind the pivotal hypoxia-regulating transcription factor HIF-1alpha. Studies using loss and gain of function of HIF-1 confirmed a critical role of HIF-1alpha in coordinating TLR2 and TLR6 induction. Moreover, studies of murine hypoxia (8% oxygen over 6 h) showed TLR2 and TLR 6 induction in mucosal organs in vivo. In contrast, hypoxia induction of TLR2 and TLR6 was abolished in conditional HIF-1alpha mutant mice.

Conclusions/significance: Taking together, these studies reveal coordinated induction of TLR2 and TLR6 during hypoxia and suggest tissue hypoxia in transcriptional adaptation of innate immune responses during acute infection or inflammation.

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Influence of hypoxia on TLR mRNA expression in murine dendritic cells.Isolated murine bone-marrow-derived dendritic cells (BMDCs) were exposed to normoxia or hypoxia for 24 hours. Total RNA was isolated, and quantitative mRNA levels of TLR1-9 and TLR11-13 were assessed by real-time RT-PCR. Data were calculated relative to ß-actin and expressed as fold change relative to normoxia±SEM and transcript levels in normoxic BMDCs were normalized to 1. Results are derived from three different experiments (*p<0.05, significant differences from normoxia).
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pone-0001364-g001: Influence of hypoxia on TLR mRNA expression in murine dendritic cells.Isolated murine bone-marrow-derived dendritic cells (BMDCs) were exposed to normoxia or hypoxia for 24 hours. Total RNA was isolated, and quantitative mRNA levels of TLR1-9 and TLR11-13 were assessed by real-time RT-PCR. Data were calculated relative to ß-actin and expressed as fold change relative to normoxia±SEM and transcript levels in normoxic BMDCs were normalized to 1. Results are derived from three different experiments (*p<0.05, significant differences from normoxia).

Mentions: Sites of infection and inflammation are characterized by dramatic shifts in metabolic supply and demand, thereby leading to profound tissue hypoxia [5]. Given the association of hypoxia with tissue-infections and the predominant role of TLR-signaling in regulating host-cell responses during infections with human pathogens [2], we pursued transcriptional responses of TLRs during hypoxia. As first step in the line of these experiments, we examined the influence of hypoxia on expressional levels of mammalian TLRs (TLR1-9 and 11-13) in vitro, using a cellular model of murine dendritic cells. For this purpose, we isolated bone-marrow-derived dendritic cells (BMDCs) from C57BL/6×129SV mice and exposed them ex vivo to normoxia or normobaric hypoxia (2% oxygen, 24h). Using real-time RT-PCR analysis, we found selective induction of mRNA levels of TLR2 (2.360±0.3810, P<0.05) and TLR6 (3.460±0.7820, p<0.05) with hypoxia exposure in comparison to normoxia (Fig. 1). In contrast, transcript levels of other TLRs remained unchanged with hypoxia exposure. Due to the fact that TLR10 expression has not been detected in murine BMDCs [17] we studies modulation of TLR 10 transcript in human microvascular endothelia (HMEC-1) and human intestinal epithelia (CaCo-2). These studies revealed no changes of TLR10 transcript with hypoxia (data not shown). Taken together, these studies show a robust and selective induction of TLR2 and TLR6 coordinated by hypoxia.


Hypoxia inducible factor (HIF)-1 coordinates induction of Toll-like receptors TLR2 and TLR6 during hypoxia.

Kuhlicke J, Frick JS, Morote-Garcia JC, Rosenberger P, Eltzschig HK - PLoS ONE (2007)

Influence of hypoxia on TLR mRNA expression in murine dendritic cells.Isolated murine bone-marrow-derived dendritic cells (BMDCs) were exposed to normoxia or hypoxia for 24 hours. Total RNA was isolated, and quantitative mRNA levels of TLR1-9 and TLR11-13 were assessed by real-time RT-PCR. Data were calculated relative to ß-actin and expressed as fold change relative to normoxia±SEM and transcript levels in normoxic BMDCs were normalized to 1. Results are derived from three different experiments (*p<0.05, significant differences from normoxia).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2147045&req=5

pone-0001364-g001: Influence of hypoxia on TLR mRNA expression in murine dendritic cells.Isolated murine bone-marrow-derived dendritic cells (BMDCs) were exposed to normoxia or hypoxia for 24 hours. Total RNA was isolated, and quantitative mRNA levels of TLR1-9 and TLR11-13 were assessed by real-time RT-PCR. Data were calculated relative to ß-actin and expressed as fold change relative to normoxia±SEM and transcript levels in normoxic BMDCs were normalized to 1. Results are derived from three different experiments (*p<0.05, significant differences from normoxia).
Mentions: Sites of infection and inflammation are characterized by dramatic shifts in metabolic supply and demand, thereby leading to profound tissue hypoxia [5]. Given the association of hypoxia with tissue-infections and the predominant role of TLR-signaling in regulating host-cell responses during infections with human pathogens [2], we pursued transcriptional responses of TLRs during hypoxia. As first step in the line of these experiments, we examined the influence of hypoxia on expressional levels of mammalian TLRs (TLR1-9 and 11-13) in vitro, using a cellular model of murine dendritic cells. For this purpose, we isolated bone-marrow-derived dendritic cells (BMDCs) from C57BL/6×129SV mice and exposed them ex vivo to normoxia or normobaric hypoxia (2% oxygen, 24h). Using real-time RT-PCR analysis, we found selective induction of mRNA levels of TLR2 (2.360±0.3810, P<0.05) and TLR6 (3.460±0.7820, p<0.05) with hypoxia exposure in comparison to normoxia (Fig. 1). In contrast, transcript levels of other TLRs remained unchanged with hypoxia exposure. Due to the fact that TLR10 expression has not been detected in murine BMDCs [17] we studies modulation of TLR 10 transcript in human microvascular endothelia (HMEC-1) and human intestinal epithelia (CaCo-2). These studies revealed no changes of TLR10 transcript with hypoxia (data not shown). Taken together, these studies show a robust and selective induction of TLR2 and TLR6 coordinated by hypoxia.

Bottom Line: During acute infection and inflammation, dramatic shifts in tissue metabolism are typical, thereby resulting in profound tissue hypoxia.While transcript levels of other TLRs remained unchanged, we found a robust induction of TLR2 (2.36+/-0.7-fold; P<0.05) and TLR6 (3.46+/-1.56-fold; P<0.05).Furthermore, analysis of the putative TLR2 and TLR6 promoters revealed previously unrecognized binding sites for HIF-1, which were shown by chromatin immunoprecipitation to bind the pivotal hypoxia-regulating transcription factor HIF-1alpha.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology and Intensive Care Medicine, Tübingen University Hospital, Tübingen, Germany.

ABSTRACT

Background: During acute infection and inflammation, dramatic shifts in tissue metabolism are typical, thereby resulting in profound tissue hypoxia. Therefore, we pursued the hypothesis, that tissue hypoxia may influence innate immune responses by transcriptional modulation of Toll-like receptor (TLRs) expression and function.

Methodology/principal findings: We gained first insight from transcriptional profiling of murine dendritic cells exposed to hypoxia (2% oxygen for 24 h). While transcript levels of other TLRs remained unchanged, we found a robust induction of TLR2 (2.36+/-0.7-fold; P<0.05) and TLR6 (3.46+/-1.56-fold; P<0.05). Additional studies in different cells types and cell-lines including human dendritic cells, monocytic cells (MM6), endothelia (HMEC-1) or intestinal epithelia (Caco-2) confirmed TLR2 and TLR6 induction of transcript, protein and function during hypoxia. Furthermore, analysis of the putative TLR2 and TLR6 promoters revealed previously unrecognized binding sites for HIF-1, which were shown by chromatin immunoprecipitation to bind the pivotal hypoxia-regulating transcription factor HIF-1alpha. Studies using loss and gain of function of HIF-1 confirmed a critical role of HIF-1alpha in coordinating TLR2 and TLR6 induction. Moreover, studies of murine hypoxia (8% oxygen over 6 h) showed TLR2 and TLR 6 induction in mucosal organs in vivo. In contrast, hypoxia induction of TLR2 and TLR6 was abolished in conditional HIF-1alpha mutant mice.

Conclusions/significance: Taking together, these studies reveal coordinated induction of TLR2 and TLR6 during hypoxia and suggest tissue hypoxia in transcriptional adaptation of innate immune responses during acute infection or inflammation.

Show MeSH
Related in: MedlinePlus