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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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iMist1myc acinar cells exhibit apoptosis followed by regeneration of iMIST1myc-negative cells upon AP induction.(A) Acinar cells in iMist1myc pancreata undergo extensive apoptosis as detected by cleaved CASPASE 3 staining in AMY+ cells. (B) Over time the number of MIST1myc+ cells is greatly decreased as the iMist1myc pancreas recovers post-AP. (C) BrdU pulse labeling reveals that regeneration of iMist1myc pancreata following AP is due to proliferation of rare acinar cells that did not activate expression of iMist1myc during the initial Tam treatment. Arrows indicate AMY+/BrdU+/MYC- cells. ***p ≤ 0.001. N.D.—not detected.
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pone.0145724.g009: iMist1myc acinar cells exhibit apoptosis followed by regeneration of iMIST1myc-negative cells upon AP induction.(A) Acinar cells in iMist1myc pancreata undergo extensive apoptosis as detected by cleaved CASPASE 3 staining in AMY+ cells. (B) Over time the number of MIST1myc+ cells is greatly decreased as the iMist1myc pancreas recovers post-AP. (C) BrdU pulse labeling reveals that regeneration of iMist1myc pancreata following AP is due to proliferation of rare acinar cells that did not activate expression of iMist1myc during the initial Tam treatment. Arrows indicate AMY+/BrdU+/MYC- cells. ***p ≤ 0.001. N.D.—not detected.

Mentions: The inability of iMist1myc mice to recover from AP damage by 7d prompted us to examine animals at extended times. During 7d-10d post-AP, iMist1myc pancreata were grossly reduced in size (S7A Fig) with no evidence of normal acini structures. Instead, the tissue was composed of loose connective tissue containing VIMENTIN+ fibroblasts, CD45+ immune cells and areas of edema (S7A Fig). Within the remaining pancreas tissue, we observed small pockets of epithelial ADM structures that exhibited elevated levels of CLUSTERIN and retained co-expression of AMY and K19 (Fig 8A and S7A and S8 Figs). However, the number of AMY+ acinar cells greatly decreased over this time period with only small groupings of acinar cells remaining at 7d post-AP (Fig 8B and 8C). During this time frame there was a significant increase in cleaved CASPASE 3+/AMY+ epithelial cells, suggesting that cell death was primarily responsible for the vivid loss of acini structures (Fig 9A and S9 Fig). Over the ensuing 3–8 weeks post-AP iMist1myc pancreata underwent a significant recovery as healthy acinar tissue began to appear in the disrupted organs (S7B Fig). Areas of ADM were replaced with relatively normal acini that were AMY+ and CLUSTERIN negative (Fig 8A–8C). Interestingly, lineage-tracing revealed that the majority of the recovered acini were MIST1myc negative. This was particularly evident in the later (3–8 wk post-AP) times. Quantification of these tissues showed that approximately 75% of AMY+ acinar cells did not express the iMIST1myc protein (Fig 9B). The increase in AMY+/MYC- acinar cells was exclusively due to an increase in cell proliferation of the MYC- population. At 3w post-AP there was an 18.7-fold increase in BrdU-labelled cells when compared to control pancreas samples. Importantly, of the regenerating cell population >90% BrdU+ cells were MYC- (Fig 9C). At 8w post-AP pancreata also accumulated small amounts of adipose tissue that typically associated with the periphery of the organ (Fig 8A). However, the fat cells were always MYC-, demonstrating that they did not arise via an acinar cell transdifferentiation event. Taken together, these results show that sustained MIST1 protein is detrimental to AP recovery and that the iMist1myc pancreas recovers from an AP episode by relying on the small percentage of acinar cells that failed to initially activate iMist1myc expression, allowing this population to re-enter a proliferative state and repopulate the organ. We conclude that sustained Mist1 expression does not alleviate the initial AP damage and instead is detrimental to maintaining a healthy acinar cell state under AP conditions.


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)

iMist1myc acinar cells exhibit apoptosis followed by regeneration of iMIST1myc-negative cells upon AP induction.(A) Acinar cells in iMist1myc pancreata undergo extensive apoptosis as detected by cleaved CASPASE 3 staining in AMY+ cells. (B) Over time the number of MIST1myc+ cells is greatly decreased as the iMist1myc pancreas recovers post-AP. (C) BrdU pulse labeling reveals that regeneration of iMist1myc pancreata following AP is due to proliferation of rare acinar cells that did not activate expression of iMist1myc during the initial Tam treatment. Arrows indicate AMY+/BrdU+/MYC- cells. ***p ≤ 0.001. N.D.—not detected.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4696804&req=5

pone.0145724.g009: iMist1myc acinar cells exhibit apoptosis followed by regeneration of iMIST1myc-negative cells upon AP induction.(A) Acinar cells in iMist1myc pancreata undergo extensive apoptosis as detected by cleaved CASPASE 3 staining in AMY+ cells. (B) Over time the number of MIST1myc+ cells is greatly decreased as the iMist1myc pancreas recovers post-AP. (C) BrdU pulse labeling reveals that regeneration of iMist1myc pancreata following AP is due to proliferation of rare acinar cells that did not activate expression of iMist1myc during the initial Tam treatment. Arrows indicate AMY+/BrdU+/MYC- cells. ***p ≤ 0.001. N.D.—not detected.
Mentions: The inability of iMist1myc mice to recover from AP damage by 7d prompted us to examine animals at extended times. During 7d-10d post-AP, iMist1myc pancreata were grossly reduced in size (S7A Fig) with no evidence of normal acini structures. Instead, the tissue was composed of loose connective tissue containing VIMENTIN+ fibroblasts, CD45+ immune cells and areas of edema (S7A Fig). Within the remaining pancreas tissue, we observed small pockets of epithelial ADM structures that exhibited elevated levels of CLUSTERIN and retained co-expression of AMY and K19 (Fig 8A and S7A and S8 Figs). However, the number of AMY+ acinar cells greatly decreased over this time period with only small groupings of acinar cells remaining at 7d post-AP (Fig 8B and 8C). During this time frame there was a significant increase in cleaved CASPASE 3+/AMY+ epithelial cells, suggesting that cell death was primarily responsible for the vivid loss of acini structures (Fig 9A and S9 Fig). Over the ensuing 3–8 weeks post-AP iMist1myc pancreata underwent a significant recovery as healthy acinar tissue began to appear in the disrupted organs (S7B Fig). Areas of ADM were replaced with relatively normal acini that were AMY+ and CLUSTERIN negative (Fig 8A–8C). Interestingly, lineage-tracing revealed that the majority of the recovered acini were MIST1myc negative. This was particularly evident in the later (3–8 wk post-AP) times. Quantification of these tissues showed that approximately 75% of AMY+ acinar cells did not express the iMIST1myc protein (Fig 9B). The increase in AMY+/MYC- acinar cells was exclusively due to an increase in cell proliferation of the MYC- population. At 3w post-AP there was an 18.7-fold increase in BrdU-labelled cells when compared to control pancreas samples. Importantly, of the regenerating cell population >90% BrdU+ cells were MYC- (Fig 9C). At 8w post-AP pancreata also accumulated small amounts of adipose tissue that typically associated with the periphery of the organ (Fig 8A). However, the fat cells were always MYC-, demonstrating that they did not arise via an acinar cell transdifferentiation event. Taken together, these results show that sustained MIST1 protein is detrimental to AP recovery and that the iMist1myc pancreas recovers from an AP episode by relying on the small percentage of acinar cells that failed to initially activate iMist1myc expression, allowing this population to re-enter a proliferative state and repopulate the organ. We conclude that sustained Mist1 expression does not alleviate the initial AP damage and instead is detrimental to maintaining a healthy acinar cell state under AP conditions.

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