Unexpected effects of the MIP-Cre<sup>ER</sup>transgene and tamoxifen on<i>β</i>-cell growth in C57Bl6/J male mice

Carboneau, Lindsey; Le, Thao D. V.; Dunn, Jennifer C.; Gannon, Maureen

doi:10.14814/phy2.12863

Cited by 38 publications

(31 citation statements)

References 30 publications

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“…As all mice in our study were on the same C57Bl/6N background, the difference between Cre-expressing (MIP-CreERT and bPKD1KO) and non-Cre-expressing (FL and WT) mice is likely due to the human growth hormone minigene included in the MIP-CreERT transgene (14). In a previous study (14), we observed normal glucose tolerance in MIP-CreERT mice of similar age (equivalent to week 0 in this study) on the C57Bl/6J background-an observation subsequently confirmed by Carboneau et al (19). We attribute this apparent discrepancy (at weeks 8 and 12 in this study) to the fact that both of these previous studies used MIP-CreERT mice on a C57Bl/6J background, while ours were on a C57Bl/6N background.…”

Section: Discussionsupporting

confidence: 88%

Deletion of Protein Kinase D1 in Pancreatic β-Cells Impairs Insulin Secretion in High-Fat Diet–Fed Mice

et al. 2017

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Ββ-Cell adaptation to insulin resistance is necessary to maintain glucose homeostasis in obesity. Failure of this mechanism is a hallmark of type 2 diabetes (T2D). Hence, factors controlling functional β-cell compensation are potentially important targets for the treatment of T2D. Protein kinase D1 (PKD1) integrates diverse signals in the β-cell and plays a critical role in the control of insulin secretion. However, the role of β-cell PKD1 in glucose homeostasis in vivo is essentially unknown. Using β-cell-specific, inducible PKD1 knockout mice (βPKD1KO), we examined the role of β-cell PKD1 under basal conditions and during high-fat feeding. βPKD1KO mice under a chow diet presented no significant difference in glucose tolerance or insulin secretion compared with mice expressing the Cre transgene alone; however, when compared with wild-type mice, both groups developed glucose intolerance. Under a high-fat diet, deletion of PKD1 in β-cells worsened hyperglycemia, hyperinsulinemia, and glucose intolerance. This was accompanied by impaired glucose-induced insulin secretion both in vivo in hyperglycemic clamps and ex vivo in isolated islets from high-fat diet-fed βPKD1KO mice without changes in islet mass. This study demonstrates an essential role for PKD1 in the β-cell adaptive secretory response to high-fat feeding in mice.

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Section: Discussionsupporting

confidence: 88%

Deletion of Protein Kinase D1 in Pancreatic β-Cells Impairs Insulin Secretion in High-Fat Diet–Fed Mice

et al. 2017

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“…To date, the field has relied on transgenic mouse lines to deliver Cre and inducible CreER proteins to cells of interest. Many Cre driver lines face caveats of off-target recombination [6][7][8] and constitutive expression of Cre may have detrimental effects on β-cell development and function 4,5,[13][14][15] . Additionally, inducible CreER lines are complicated by GH minigene inclusions within transgenes, leaky recombination before tamoxifen delivery, potential β-cell defects, and tamoxifen toxicity.…”

Section: Discussionmentioning

confidence: 99%

“…Further, inducible ER conjugated Cre models like the Ins2-cre/Esr1 mouse can have tamoxifen-independent recombination 11 but this can be limited by use of the mutated ER in Cre-ER T212 . However, it is notable that delivery of tamoxifen itself (to induce recombination) can alter glucose homeostasis and impair β-cell proliferation 13 . Additionally, inclusion of a growth hormone (GH) minigene in many Cre transgenic driver lines (including Ins2-Cre, Pdx1-Cre Late , Ins1-Cre and others) is also a source of β-cell dysfunction via local activation of prolactin receptors 14 that can induce β-cell proliferation 15 .…”

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confidence: 99%

AAV8 Ins1-Cre can produce efficient β-cell recombination but requires consideration of off-target effects

Ramzy

Tudurí

Glavas

et al. 2020

Sci Rep

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In vivo genetic manipulation is used to study the impact of gene deletion or re-expression on β-cell function and organism physiology. Cre-LoxP is a system wherein LoxP sites flanking a gene are recognized by Cre recombinase. Cre transgenic mice are the most prevalent technology used to deliver Cre but many models have caveats of off-target recombination, impaired β-cell function, and high cost of animal production. Inducible estrogen receptor conjugated Cre models face leaky recombination and confounding effects of tamoxifen. As an alternative, we characterize an adeno associated virus (AAV) with a rat insulin 1 promoter driving Cre recombinase (AAV8 Ins1-Cre) that is economical and rapid to implement, and has limited caveats. Intraperitoneal AAV8 Ins1-Cre produced efficient β-cell recombination, alongside some hepatic, exocrine pancreas, α-cell, δ-cell, and hypothalamic recombination. Delivery of lower doses via the pancreatic duct retained good rates of β-cell recombination and limited rates of off-target recombination. Unlike inducible Cre in transgenic mice, AAV8 Ins1-Cre required no tamoxifen and premature recombination was avoided. We demonstrate the utility of this technology by inducing hyperglycemia in inducible insulin knockout mice (Ins1 −/− ;Ins2 f/f). AAV-mediated expression of Cre in β-cells provides an effective alternative to transgenic approaches for inducible knockout studies. Diabetes is a chronic condition affecting over 400 million worldwide and is characterized by a relative or absolute insulin insufficiency associated with a loss of insulin secreting pancreatic β-cells. As such, the development and function of β-cells is a major focus of diabetes research. Specific in vivo genetic modification is a useful way to assess the impact of genes on relevant physiological processes and one of the most useful genetic tools to study the role of specific genes is the Cre-LoxP system. Cre recombinase is an enzyme that recognizes LoxP sites in the genome and based on orientation and location, can excise, flip, or translocate targets. By using a tissue specific promoter for Cre recombinase, LoxP flanked sites can be deleted in a tissue-specific manner 1. Additionally, recombination can be temporally controlled by conjugating Cre to a modified estrogen receptor (ER). By fusing Cre to an ER, Cre is retained in the cytoplasm and is thus unable to bind DNA until the tamoxifen metabolic products endoxifen or 4-hydroxytamoxifen bind the ER and translocate Cre to the nucleus. Many Cre driver mouse lines have been generated, including dozens that are specific for the pancreas or certain pancreatic cell lineages 2. These tools have been used extensively with great success, but this approach faces important caveats. One of the earliest pancreatic Cre driver mice developed uses 668 bp of the rat insulin 2 (Ins2) promoter to drive Cre, often called the "RIP-Cre" mouse 3. This mouse model has been used in many studies, but findings have been challenged because of a lack of appropriate controls in many studies. There...

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“…For instance, tamoxifen can strongly attenuate CCl4‐induced hepatotoxicity in male C57BL/6N mice, even after a 10‐day tamoxifen exposure‐free period . It also interferes with fibrogenesis in obstructive nephropathy and impairs the induction of pancreatic β‐cell proliferation during embryonic development . Therefore, the implementation of an optimized and adjusted tamoxifen treatment into the overall experimental scheme is required to achieve an efficient Cre recombination while minimizing ‘off‐target’ effects.…”

Section: Discussionmentioning

confidence: 99%

“…This is followed by varied drug‐free days after the last tamoxifen application in order to allow for optimal recombination before further interventions . However, recent studies indicated that tamoxifen not only exerts the role of activating Cre recombinase, but also has various side effects in different genetically engineered mouse models .…”

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confidence: 99%

Tamoxifen affects chronic pancreatitis‐related fibrogenesis in an experimental mouse model: an effect beyond Cre recombination

Clappier

Kleiter

et al. 2019

FEBS Open Bio

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Tamoxifen is very successfully used for the induction of CreERT‐mediated genomic recombination in conditional mouse models. Recent studies, however, indicated that tamoxifen might also affect the fibrotic response in several disease models following administration, both in vitro and in vivo. In order to investigate a possible effect of tamoxifen on pancreatic fibrogenesis and to evaluate an optimal treatment scheme in an experimental pancreatitis mouse model, we administered tamoxifen by oral gavage to both male and female C57BL/6J mice and then waited for different periods of time before inducing chronic pancreatitis by cerulein. We observed a sex‐specific and time‐dependent effect of tamoxifen on the fibrotic response as measured by collagen deposition and the number of myofibroblasts and macrophages. The findings of in vitro studies, in which cerulein was administrated with or without 4‐hydroxytamoxifen to stimulate primary murine female and male pancreatic stellate cells, supported our in vivo observations. Real‐time PCR also indicated that this effect may be related to differences in ERα expression between female and male stellate cells. Our data demonstrate that tamoxifen administration has unignorable side effects, which affect the experimental outcome in a cerulein‐based model of chronic pancreatitis in mice. We suggest a 2‐week waiting period before cerulein administration to reduce side effects to a minimum for the described fibrosis model in female mice.

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Unexpected effects of the MIP-Cre^ERtransgene and tamoxifen onβ-cell growth in C57Bl6/J male mice

Cited by 38 publications

References 30 publications

Deletion of Protein Kinase D1 in Pancreatic β-Cells Impairs Insulin Secretion in High-Fat Diet–Fed Mice

Deletion of Protein Kinase D1 in Pancreatic β-Cells Impairs Insulin Secretion in High-Fat Diet–Fed Mice

AAV8 Ins1-Cre can produce efficient β-cell recombination but requires consideration of off-target effects

Tamoxifen affects chronic pancreatitis‐related fibrogenesis in an experimental mouse model: an effect beyond Cre recombination

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Unexpected effects of the MIP-CreERtransgene and tamoxifen onβ-cell growth in C57Bl6/J male mice

Cited by 38 publications

References 30 publications

Deletion of Protein Kinase D1 in Pancreatic β-Cells Impairs Insulin Secretion in High-Fat Diet–Fed Mice

Deletion of Protein Kinase D1 in Pancreatic β-Cells Impairs Insulin Secretion in High-Fat Diet–Fed Mice

AAV8 Ins1-Cre can produce efficient β-cell recombination but requires consideration of off-target effects

Tamoxifen affects chronic pancreatitis‐related fibrogenesis in an experimental mouse model: an effect beyond Cre recombination

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Unexpected effects of the MIP-Cre^ERtransgene and tamoxifen onβ-cell growth in C57Bl6/J male mice