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Extracellular acidification stimulates GPR68 mediated IL-8 production in human pancreatic β cells.

Chandra V, Karamitri A, Richards P, Cormier F, Ramond C, Jockers R, Armanet M, Albagli-Curiel O, Scharfmann R - Sci Rep (2016)

Bottom Line: However, the mechanisms by which human β-cells sense and respond to acidic pH remain elusive.Using gain- and loss-of-function studies, we provide evidence that the β-cell enriched transcription factor RFX6 is a major regulator of GPR68.Further, we show that acidic pH stimulates the production and secretion of the chemokine IL-8 by β-cells through NF-кB activation.

View Article: PubMed Central - PubMed

Affiliation: INSERM, U1016, Institut Cochin, Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, 75014, France.

ABSTRACT
Acute or chronic metabolic complications such as diabetic ketoacidosis are often associated with extracellular acidification and pancreatic β-cell dysfunction. However, the mechanisms by which human β-cells sense and respond to acidic pH remain elusive. In this study, using the recently developed human β-cell line EndoC-βH2, we demonstrate that β-cells respond to extracellular acidification through GPR68, which is the predominant proton sensing receptor of human β-cells. Using gain- and loss-of-function studies, we provide evidence that the β-cell enriched transcription factor RFX6 is a major regulator of GPR68. Further, we show that acidic pH stimulates the production and secretion of the chemokine IL-8 by β-cells through NF-кB activation. Blocking of GPR68 or NF-кB activity severely attenuated acidification induced IL-8 production. Thus, we provide mechanistic insights into GPR68 mediated β-cell response to acidic microenvironment, which could be a new target to protect β-cell against acidosis induced inflammation.

No MeSH data available.


Related in: MedlinePlus

Expression of proton sensing GPCRs in EndoC-βH2, SKPC and human islets.Transcript levels of proton sensing GPCRs (GPR68, GPR4, GPR65 and GPR132) determined by RT-qPCR in (a) EndoC-βH2 cells compared with ductal carcinoma SKPC cell-line; (b) human islet preparation. Data represented as mean values ± SEM of at least 3 independent experiments. ***p < 0.001; (one-way ANOVA, followed by a Tukey’s multiple comparisons post-test).
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f1: Expression of proton sensing GPCRs in EndoC-βH2, SKPC and human islets.Transcript levels of proton sensing GPCRs (GPR68, GPR4, GPR65 and GPR132) determined by RT-qPCR in (a) EndoC-βH2 cells compared with ductal carcinoma SKPC cell-line; (b) human islet preparation. Data represented as mean values ± SEM of at least 3 independent experiments. ***p < 0.001; (one-way ANOVA, followed by a Tukey’s multiple comparisons post-test).

Mentions: Our previously published transcriptomic analyses (GEO No: GSE48101) indicated that EndoC-βH2 cells express mRNA coding for the proton-sensing receptor GPR6824. We validated these data by Real-Time-quantitative PCR (RT-qPCR) that indicated that GPR68 mRNA expression was enriched in EndoC-βH2 cells compared to the duct cell line SKPC (Fig. 1a). Transient transfection of EGFP tagged human GPR68 construct in EndoC-βH2 cells showed its predominant localization on the plasma membrane (Supplementary Fig. 1). GPR68 was almost the sole proton sensing GPCR expressed in EndoC-βH2 cells, the other ones (GPR4, GPR65, GPR132) being expressed at nearly undetectable levels (Fig. 1a). Similar data were obtained using human islet preparations that expressed GPR68, but not GPR65 and GPR132 (Fig. 1b). Of note, GPR4 was detected in human islets and not in EndoC-βH2 cells (Fig. 1a), which could be due to its expression by non β-cells present in human islet preparations like endothelial cells2526.


Extracellular acidification stimulates GPR68 mediated IL-8 production in human pancreatic β cells.

Chandra V, Karamitri A, Richards P, Cormier F, Ramond C, Jockers R, Armanet M, Albagli-Curiel O, Scharfmann R - Sci Rep (2016)

Expression of proton sensing GPCRs in EndoC-βH2, SKPC and human islets.Transcript levels of proton sensing GPCRs (GPR68, GPR4, GPR65 and GPR132) determined by RT-qPCR in (a) EndoC-βH2 cells compared with ductal carcinoma SKPC cell-line; (b) human islet preparation. Data represented as mean values ± SEM of at least 3 independent experiments. ***p < 0.001; (one-way ANOVA, followed by a Tukey’s multiple comparisons post-test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4863151&req=5

f1: Expression of proton sensing GPCRs in EndoC-βH2, SKPC and human islets.Transcript levels of proton sensing GPCRs (GPR68, GPR4, GPR65 and GPR132) determined by RT-qPCR in (a) EndoC-βH2 cells compared with ductal carcinoma SKPC cell-line; (b) human islet preparation. Data represented as mean values ± SEM of at least 3 independent experiments. ***p < 0.001; (one-way ANOVA, followed by a Tukey’s multiple comparisons post-test).
Mentions: Our previously published transcriptomic analyses (GEO No: GSE48101) indicated that EndoC-βH2 cells express mRNA coding for the proton-sensing receptor GPR6824. We validated these data by Real-Time-quantitative PCR (RT-qPCR) that indicated that GPR68 mRNA expression was enriched in EndoC-βH2 cells compared to the duct cell line SKPC (Fig. 1a). Transient transfection of EGFP tagged human GPR68 construct in EndoC-βH2 cells showed its predominant localization on the plasma membrane (Supplementary Fig. 1). GPR68 was almost the sole proton sensing GPCR expressed in EndoC-βH2 cells, the other ones (GPR4, GPR65, GPR132) being expressed at nearly undetectable levels (Fig. 1a). Similar data were obtained using human islet preparations that expressed GPR68, but not GPR65 and GPR132 (Fig. 1b). Of note, GPR4 was detected in human islets and not in EndoC-βH2 cells (Fig. 1a), which could be due to its expression by non β-cells present in human islet preparations like endothelial cells2526.

Bottom Line: However, the mechanisms by which human β-cells sense and respond to acidic pH remain elusive.Using gain- and loss-of-function studies, we provide evidence that the β-cell enriched transcription factor RFX6 is a major regulator of GPR68.Further, we show that acidic pH stimulates the production and secretion of the chemokine IL-8 by β-cells through NF-кB activation.

View Article: PubMed Central - PubMed

Affiliation: INSERM, U1016, Institut Cochin, Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, 75014, France.

ABSTRACT
Acute or chronic metabolic complications such as diabetic ketoacidosis are often associated with extracellular acidification and pancreatic β-cell dysfunction. However, the mechanisms by which human β-cells sense and respond to acidic pH remain elusive. In this study, using the recently developed human β-cell line EndoC-βH2, we demonstrate that β-cells respond to extracellular acidification through GPR68, which is the predominant proton sensing receptor of human β-cells. Using gain- and loss-of-function studies, we provide evidence that the β-cell enriched transcription factor RFX6 is a major regulator of GPR68. Further, we show that acidic pH stimulates the production and secretion of the chemokine IL-8 by β-cells through NF-кB activation. Blocking of GPR68 or NF-кB activity severely attenuated acidification induced IL-8 production. Thus, we provide mechanistic insights into GPR68 mediated β-cell response to acidic microenvironment, which could be a new target to protect β-cell against acidosis induced inflammation.

No MeSH data available.


Related in: MedlinePlus