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Silencing Mist1 Gene Expression Is Essential for Recovery from Acute Pancreatitis.

Karki A, Humphrey SE, Steele RE, Hess DA, Taparowsky EJ, Konieczny SF - PLoS ONE (2015)

Bottom Line: Despite a wealth of information concerning the broad phenotype associated with pancreatitis, little is understood regarding specific transcriptional regulatory networks that are susceptible to AP and the role these networks play in acinar cell and exocrine pancreas responses.We conclude that the transient silencing of Mist1 expression is critical for acinar cells to survive an AP episode, providing cells an opportunity to suppress their secretory function and regenerate damaged cells.The importance of MIST1 to these events suggests that modulating key pancreas transcription networks could ease clinical symptoms in patients diagnosed with pancreatitis and pancreatic cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, the Purdue Center for Cancer Research, and the Bindley Bioscience Center, Purdue University, West Lafayette, IN, 47907-2057, United States of America.

ABSTRACT
Acinar cells of the exocrine pancreas are tasked with synthesizing, packaging and secreting vast quantities of pro-digestive enzymes to maintain proper metabolic homeostasis for the organism. Because the synthesis of high levels of hydrolases is potentially dangerous, the pancreas is prone to acute pancreatitis (AP), a disease that targets acinar cells, leading to acinar-ductal metaplasia (ADM), inflammation and fibrosis-events that can transition into the earliest stages of pancreatic ductal adenocarcinoma. Despite a wealth of information concerning the broad phenotype associated with pancreatitis, little is understood regarding specific transcriptional regulatory networks that are susceptible to AP and the role these networks play in acinar cell and exocrine pancreas responses. In this study, we examined the importance of the acinar-specific maturation transcription factor MIST1 to AP damage and organ recovery. Analysis of wild-type and Mist1 conditional mice revealed that Mist1 gene transcription and protein accumulation were dramatically reduced as acinar cells underwent ADM alterations during AP episodes. To test if loss of MIST1 function was primarily responsible for the damaged status of the organ, mice harboring a Cre-inducible Mist1 transgene (iMist1) were utilized to determine if sustained MIST1 activity could alleviate AP damage responses. Unexpectedly, constitutive iMist1 expression during AP led to a dramatic increase in organ damage followed by acinar cell death. We conclude that the transient silencing of Mist1 expression is critical for acinar cells to survive an AP episode, providing cells an opportunity to suppress their secretory function and regenerate damaged cells. The importance of MIST1 to these events suggests that modulating key pancreas transcription networks could ease clinical symptoms in patients diagnosed with pancreatitis and pancreatic cancer.

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Characterization of Mist1CreERT/+ mice following acute pancreatitis.(A) Time course diagram of caerulein-induced acute pancreatitis. (B) H&E and IF analyses of Mist1CreERT/+ pancreas samples in the absence of AP treatment (control) or post-AP for the indicated times. At early times acinar cells exhibit elevated levels of Clusterin expression and Amylase+/K19+ ADM lesions. However, by 10d post-AP the majority of the tissue fully recovers. Arrows in the H&E section point out recovered acini whereas arrows in the K19/AMY panels indicate ADM lesions. (d, duct) (C) Immunoblot analysis of Mist1CreERT/+ pancreata post-AP. HSP90 was used as a loading control. Relative expression levels are indicated below each panel, normalized to the corresponding HSP90 signal. (D) RT-qPCR analysis of gene transcripts confirms the initial ADM phenotype followed by recovery at 10d post-AP. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001.
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pone.0145724.g001: Characterization of Mist1CreERT/+ mice following acute pancreatitis.(A) Time course diagram of caerulein-induced acute pancreatitis. (B) H&E and IF analyses of Mist1CreERT/+ pancreas samples in the absence of AP treatment (control) or post-AP for the indicated times. At early times acinar cells exhibit elevated levels of Clusterin expression and Amylase+/K19+ ADM lesions. However, by 10d post-AP the majority of the tissue fully recovers. Arrows in the H&E section point out recovered acini whereas arrows in the K19/AMY panels indicate ADM lesions. (d, duct) (C) Immunoblot analysis of Mist1CreERT/+ pancreata post-AP. HSP90 was used as a loading control. Relative expression levels are indicated below each panel, normalized to the corresponding HSP90 signal. (D) RT-qPCR analysis of gene transcripts confirms the initial ADM phenotype followed by recovery at 10d post-AP. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001.

Mentions: To evaluate the importance of MIST1 during acinar metaplasia, we characterized Mist1 expression during the damage and subsequent recovery phases of AP, a known driver of PDAC tumor development [15, 18, 19, 21]. For these studies, Mist1CreERT/+ mice were used as controls as all subsequent mouse lines contained the Mist1CreERT knock-in allele [26, 58]. Standard caerulein treatment (Fig 1A) of 8 week Mist1CreERT/+ mice led to significant and rapid damage to the exocrine acinar cells. As early as 6h post-AP, acinar lumens were distended and zymogen granules were rapidly lost (Fig 1B and S1A Fig). By 1d post-AP, significant increases in edema and inflammatory cell infiltrates were observed, accompanied by extensive formation of KERATIN19 (K19)+/AMYLASE (AMY)+ ADM lesions. Expression of CLUSTERIN, a known marker of acinar cell damage [67, 68], also was greatly elevated at 6h post-AP (Fig 1B,1C and S1A,S1B Fig). Transcript and protein levels of acinar cell markers, including Amylase (Amy), Trypsinogen (Tryp) and Carboxypeptidase (Cpa), were significantly reduced over the 6h-2d post-AP period (Fig 1C,1D and S1B Fig). In contrast, ductal markers (K19, SOX9) were greatly elevated, confirming the formation of extensive ADM (Fig 1B–1D, S1B Fig). Identical ADM responses were obtained with caerulein-treated wild-type mice (data not shown). Despite significant development of ADM lesions upon AP induction, AP metaplasia was transient as lesions resolved 4d-10d post-AP. In all cases, Clusterin, K19 and Sox9 transcript and protein levels returned to their low control states while acinar markers (Amylase, Trypsinogen, Carboxypeptidase) re-established high expression thresholds (Fig 1B–1D and S1A,S1B Fig).


Silencing Mist1 Gene Expression Is Essential for Recovery from Acute Pancreatitis.

Karki A, Humphrey SE, Steele RE, Hess DA, Taparowsky EJ, Konieczny SF - PLoS ONE (2015)

Characterization of Mist1CreERT/+ mice following acute pancreatitis.(A) Time course diagram of caerulein-induced acute pancreatitis. (B) H&E and IF analyses of Mist1CreERT/+ pancreas samples in the absence of AP treatment (control) or post-AP for the indicated times. At early times acinar cells exhibit elevated levels of Clusterin expression and Amylase+/K19+ ADM lesions. However, by 10d post-AP the majority of the tissue fully recovers. Arrows in the H&E section point out recovered acini whereas arrows in the K19/AMY panels indicate ADM lesions. (d, duct) (C) Immunoblot analysis of Mist1CreERT/+ pancreata post-AP. HSP90 was used as a loading control. Relative expression levels are indicated below each panel, normalized to the corresponding HSP90 signal. (D) RT-qPCR analysis of gene transcripts confirms the initial ADM phenotype followed by recovery at 10d post-AP. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4696804&req=5

pone.0145724.g001: Characterization of Mist1CreERT/+ mice following acute pancreatitis.(A) Time course diagram of caerulein-induced acute pancreatitis. (B) H&E and IF analyses of Mist1CreERT/+ pancreas samples in the absence of AP treatment (control) or post-AP for the indicated times. At early times acinar cells exhibit elevated levels of Clusterin expression and Amylase+/K19+ ADM lesions. However, by 10d post-AP the majority of the tissue fully recovers. Arrows in the H&E section point out recovered acini whereas arrows in the K19/AMY panels indicate ADM lesions. (d, duct) (C) Immunoblot analysis of Mist1CreERT/+ pancreata post-AP. HSP90 was used as a loading control. Relative expression levels are indicated below each panel, normalized to the corresponding HSP90 signal. (D) RT-qPCR analysis of gene transcripts confirms the initial ADM phenotype followed by recovery at 10d post-AP. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001.
Mentions: To evaluate the importance of MIST1 during acinar metaplasia, we characterized Mist1 expression during the damage and subsequent recovery phases of AP, a known driver of PDAC tumor development [15, 18, 19, 21]. For these studies, Mist1CreERT/+ mice were used as controls as all subsequent mouse lines contained the Mist1CreERT knock-in allele [26, 58]. Standard caerulein treatment (Fig 1A) of 8 week Mist1CreERT/+ mice led to significant and rapid damage to the exocrine acinar cells. As early as 6h post-AP, acinar lumens were distended and zymogen granules were rapidly lost (Fig 1B and S1A Fig). By 1d post-AP, significant increases in edema and inflammatory cell infiltrates were observed, accompanied by extensive formation of KERATIN19 (K19)+/AMYLASE (AMY)+ ADM lesions. Expression of CLUSTERIN, a known marker of acinar cell damage [67, 68], also was greatly elevated at 6h post-AP (Fig 1B,1C and S1A,S1B Fig). Transcript and protein levels of acinar cell markers, including Amylase (Amy), Trypsinogen (Tryp) and Carboxypeptidase (Cpa), were significantly reduced over the 6h-2d post-AP period (Fig 1C,1D and S1B Fig). In contrast, ductal markers (K19, SOX9) were greatly elevated, confirming the formation of extensive ADM (Fig 1B–1D, S1B Fig). Identical ADM responses were obtained with caerulein-treated wild-type mice (data not shown). Despite significant development of ADM lesions upon AP induction, AP metaplasia was transient as lesions resolved 4d-10d post-AP. In all cases, Clusterin, K19 and Sox9 transcript and protein levels returned to their low control states while acinar markers (Amylase, Trypsinogen, Carboxypeptidase) re-established high expression thresholds (Fig 1B–1D and S1A,S1B Fig).

Bottom Line: Despite a wealth of information concerning the broad phenotype associated with pancreatitis, little is understood regarding specific transcriptional regulatory networks that are susceptible to AP and the role these networks play in acinar cell and exocrine pancreas responses.We conclude that the transient silencing of Mist1 expression is critical for acinar cells to survive an AP episode, providing cells an opportunity to suppress their secretory function and regenerate damaged cells.The importance of MIST1 to these events suggests that modulating key pancreas transcription networks could ease clinical symptoms in patients diagnosed with pancreatitis and pancreatic cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, the Purdue Center for Cancer Research, and the Bindley Bioscience Center, Purdue University, West Lafayette, IN, 47907-2057, United States of America.

ABSTRACT
Acinar cells of the exocrine pancreas are tasked with synthesizing, packaging and secreting vast quantities of pro-digestive enzymes to maintain proper metabolic homeostasis for the organism. Because the synthesis of high levels of hydrolases is potentially dangerous, the pancreas is prone to acute pancreatitis (AP), a disease that targets acinar cells, leading to acinar-ductal metaplasia (ADM), inflammation and fibrosis-events that can transition into the earliest stages of pancreatic ductal adenocarcinoma. Despite a wealth of information concerning the broad phenotype associated with pancreatitis, little is understood regarding specific transcriptional regulatory networks that are susceptible to AP and the role these networks play in acinar cell and exocrine pancreas responses. In this study, we examined the importance of the acinar-specific maturation transcription factor MIST1 to AP damage and organ recovery. Analysis of wild-type and Mist1 conditional mice revealed that Mist1 gene transcription and protein accumulation were dramatically reduced as acinar cells underwent ADM alterations during AP episodes. To test if loss of MIST1 function was primarily responsible for the damaged status of the organ, mice harboring a Cre-inducible Mist1 transgene (iMist1) were utilized to determine if sustained MIST1 activity could alleviate AP damage responses. Unexpectedly, constitutive iMist1 expression during AP led to a dramatic increase in organ damage followed by acinar cell death. We conclude that the transient silencing of Mist1 expression is critical for acinar cells to survive an AP episode, providing cells an opportunity to suppress their secretory function and regenerate damaged cells. The importance of MIST1 to these events suggests that modulating key pancreas transcription networks could ease clinical symptoms in patients diagnosed with pancreatitis and pancreatic cancer.

Show MeSH
Related in: MedlinePlus