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Hydrogen sulfide acts as a mediator of inflammation in acute pancreatitis: in vitro studies using isolated mouse pancreatic acinar cells.

Tamizhselvi R, Moore PK, Bhatia M - J. Cell. Mol. Med. (2007 Mar-Apr)

Bottom Line: Caerulein increased the levels of H(2)S and CSE mRNA expression while CBS mRNA expression was decreased.Inhibition of endogenous production of H(2)S by PAG significantly suppressed SP concentration, PPT-A expression and NK1-R expression in the acini.To determine whether H(2)S itself provoked inflammation in acinar cells, the cells were treated with H(2)S donor drug, sodium hydrosulphide (NaHS), (10, 50 and 100 muM), that resulted in a significant increase in SP concentration and expression of PPT-A and NK1-R in acinar cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, National University of Singapore, Yong Loo Lin School of Medicine, Centre for the Sciences, 28 Medical Drive, Singapore 117456.

ABSTRACT
Hydrogen sulphide (H(2)S) is synthesized from L-cysteine via the action of cystathionine-gamma-lyase (CSE) and cystathionine-beta-synthase (CBS). We have earlier shown that H(2)S acts as a mediator of inflammation. However the mechanism remains unclear. In this study, we investigated the presence of H(2)S and the expression of H(2)S synthesizing enzymes, CSE and CBS, in isolated mouse pancreatic acini. Pancreatic acinar cells from mice were incubated with or without caerulein (10(-7) M for 30 and 60 min). Caerulein increased the levels of H(2)S and CSE mRNA expression while CBS mRNA expression was decreased. In addition, cells pre-treated with DL-propargylglycine (PAG, 3 mM), a CSE inhibitor, reduced the formation of H(2)S in caerulein treated cells, suggesting that CSE may be the main enzyme involved in H(2)S formation in mouse acinar cells. Furthermore, substance P (SP) concentration in the acini and expression of SP gene (preprotachykinin-A, PPT-A) and neurokinin-1 receptor (NK-1R), the primary receptor for SP, are increased in secretagogue caerulein-treated acinar cells. Inhibition of endogenous production of H(2)S by PAG significantly suppressed SP concentration, PPT-A expression and NK1-R expression in the acini. To determine whether H(2)S itself provoked inflammation in acinar cells, the cells were treated with H(2)S donor drug, sodium hydrosulphide (NaHS), (10, 50 and 100 muM), that resulted in a significant increase in SP concentration and expression of PPT-A and NK1-R in acinar cells. These results suggest that the pro-inflammatory effect of H(2)S may be mediated by SP-NK-1R related pathway in mouse pancreatic acinar cells.

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PAG treatment suppressed caerulein-induced up regulation of preprotachykinin-A (PPT-A) and neurokonin-1 receptor (NK-1R) mRNA expression in mouse pancreatic acinar cells. (A) PPT-A mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (30 min before caerulein treatment) cells. The graphs represent the optical density of the bands of PPT-A generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between control and caerulein treated cells. (B) NK-1 receptor mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (60 min before caerulein treatment) cells. The graphs represent the optical density of the bands of NK-1 receptor generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between caerulein and PAG pretreated cells.
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fig05: PAG treatment suppressed caerulein-induced up regulation of preprotachykinin-A (PPT-A) and neurokonin-1 receptor (NK-1R) mRNA expression in mouse pancreatic acinar cells. (A) PPT-A mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (30 min before caerulein treatment) cells. The graphs represent the optical density of the bands of PPT-A generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between control and caerulein treated cells. (B) NK-1 receptor mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (60 min before caerulein treatment) cells. The graphs represent the optical density of the bands of NK-1 receptor generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between caerulein and PAG pretreated cells.

Mentions: Figure 4 shows the concentration of SP in the pancreatic acinar cells. As expected, hyper stimulation by caerulein increased the levels of SP in acinar cells (P < 0.001). Pretreatment of acini with 3 mM PAG followed by stimulation with caerulein significantly suppressed the SP concentration when compared with caerulein treated cells (P < 0.05). Densitometry analysis of the PCR products on agarose gel shows that the PPT-A mRNA expression increased in caerulein induced acinar cells (P < 0.001) and pretreatment with 3 mM PAG resulted in a significant decrease in PPT-A gene expression (P < 0.001; Fig. 5A).


Hydrogen sulfide acts as a mediator of inflammation in acute pancreatitis: in vitro studies using isolated mouse pancreatic acinar cells.

Tamizhselvi R, Moore PK, Bhatia M - J. Cell. Mol. Med. (2007 Mar-Apr)

PAG treatment suppressed caerulein-induced up regulation of preprotachykinin-A (PPT-A) and neurokonin-1 receptor (NK-1R) mRNA expression in mouse pancreatic acinar cells. (A) PPT-A mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (30 min before caerulein treatment) cells. The graphs represent the optical density of the bands of PPT-A generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between control and caerulein treated cells. (B) NK-1 receptor mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (60 min before caerulein treatment) cells. The graphs represent the optical density of the bands of NK-1 receptor generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between caerulein and PAG pretreated cells.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3822830&req=5

fig05: PAG treatment suppressed caerulein-induced up regulation of preprotachykinin-A (PPT-A) and neurokonin-1 receptor (NK-1R) mRNA expression in mouse pancreatic acinar cells. (A) PPT-A mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (30 min before caerulein treatment) cells. The graphs represent the optical density of the bands of PPT-A generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between control and caerulein treated cells. (B) NK-1 receptor mRNA expression in control, caerulein-treated (10−7 M for 60 min) and PAG pretreated (3 mM) (60 min before caerulein treatment) cells. The graphs represent the optical density of the bands of NK-1 receptor generated from six independent experiments that were normalized with the expression of 18S. ***P < 0.001 between caerulein and PAG pretreated cells.
Mentions: Figure 4 shows the concentration of SP in the pancreatic acinar cells. As expected, hyper stimulation by caerulein increased the levels of SP in acinar cells (P < 0.001). Pretreatment of acini with 3 mM PAG followed by stimulation with caerulein significantly suppressed the SP concentration when compared with caerulein treated cells (P < 0.05). Densitometry analysis of the PCR products on agarose gel shows that the PPT-A mRNA expression increased in caerulein induced acinar cells (P < 0.001) and pretreatment with 3 mM PAG resulted in a significant decrease in PPT-A gene expression (P < 0.001; Fig. 5A).

Bottom Line: Caerulein increased the levels of H(2)S and CSE mRNA expression while CBS mRNA expression was decreased.Inhibition of endogenous production of H(2)S by PAG significantly suppressed SP concentration, PPT-A expression and NK1-R expression in the acini.To determine whether H(2)S itself provoked inflammation in acinar cells, the cells were treated with H(2)S donor drug, sodium hydrosulphide (NaHS), (10, 50 and 100 muM), that resulted in a significant increase in SP concentration and expression of PPT-A and NK1-R in acinar cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, National University of Singapore, Yong Loo Lin School of Medicine, Centre for the Sciences, 28 Medical Drive, Singapore 117456.

ABSTRACT
Hydrogen sulphide (H(2)S) is synthesized from L-cysteine via the action of cystathionine-gamma-lyase (CSE) and cystathionine-beta-synthase (CBS). We have earlier shown that H(2)S acts as a mediator of inflammation. However the mechanism remains unclear. In this study, we investigated the presence of H(2)S and the expression of H(2)S synthesizing enzymes, CSE and CBS, in isolated mouse pancreatic acini. Pancreatic acinar cells from mice were incubated with or without caerulein (10(-7) M for 30 and 60 min). Caerulein increased the levels of H(2)S and CSE mRNA expression while CBS mRNA expression was decreased. In addition, cells pre-treated with DL-propargylglycine (PAG, 3 mM), a CSE inhibitor, reduced the formation of H(2)S in caerulein treated cells, suggesting that CSE may be the main enzyme involved in H(2)S formation in mouse acinar cells. Furthermore, substance P (SP) concentration in the acini and expression of SP gene (preprotachykinin-A, PPT-A) and neurokinin-1 receptor (NK-1R), the primary receptor for SP, are increased in secretagogue caerulein-treated acinar cells. Inhibition of endogenous production of H(2)S by PAG significantly suppressed SP concentration, PPT-A expression and NK1-R expression in the acini. To determine whether H(2)S itself provoked inflammation in acinar cells, the cells were treated with H(2)S donor drug, sodium hydrosulphide (NaHS), (10, 50 and 100 muM), that resulted in a significant increase in SP concentration and expression of PPT-A and NK1-R in acinar cells. These results suggest that the pro-inflammatory effect of H(2)S may be mediated by SP-NK-1R related pathway in mouse pancreatic acinar cells.

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