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Expression of wild-type and mutant S20G hIAPP in physiologic knock-in mouse models fails to induce islet amyloid formation, but induces mild glucose intolerance.

Hiddinga HJ, Sakagashira S, Ishigame M, Madde P, Sanke T, Nanjo K, Kudva YC, Lee JJ, van Deursen J, Eberhardt NL - J Diabetes Investig (2012)

Bottom Line: WW and GW mice maintained on both diets lacked intraislet amyloid at all ages.On both diets relative to MM controls WW and GW mice exhibit glucose intolerance (P < 0.008) with no differences in insulin secretion.However, GW mice secreted significantly more insulin (P < 0.03 that WW mice on both diets throughout the study.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine/Divisions of Endocrinology.

ABSTRACT

Unlabelled: Aims/Introduction:  Human islet polypeptide S20G mutation (hIAPP(S20G)) is associated with earlier onset type 2 diabetes and increased amyloidogenicity and cytotoxicity in vitro vs wild-type hIAPP (hIAPP(WT)), suggesting that amyloidogenesis may be pathogenic for type 2 diabetes. We compared the contributions of hIAPP(S20G) and hIAPP(WT) toward intra islet amyloid formation and development of type 2 diabetes in a unique physiologic knock-in mouse model.

Materials and methods:   We replaced the mouse IAPP gene (M allele) with hIAPP(WT) (W allele) and hIAPP(S20G) (G allele) via homologous recombination and backbred transgenic mice against C57Bl/6 strain 5 generations to minimize genetic variation. Mice (3 month old) were maintained on control (CD) or high fat diet (HFD) for 15 months and studied at 3 month intervals by oral glucose tolerance testing (OGTT) and pancreas histology to assess glucose homeostastis, amyloidogeneisis, islet mass, β cell replication, and apoptosis.

Results:   IAPP blood levels were indistinguishable in all mice. WW and GW mice maintained on both diets lacked intraislet amyloid at all ages. On both diets relative to MM controls WW and GW mice exhibit glucose intolerance (P < 0.008) with no differences in insulin secretion. However, GW mice secreted significantly more insulin (P < 0.03 that WW mice on both diets throughout the study. By 12 months on the high fat diet all mice increased their β cell mass about 3-fold and were indistinguishable.

Conclusions:   Physiologic expression of hIAPP(WT) and hIAPP(S20G) in C57Bl/6 mice produces mild glucose intolerance with inappropriately normal insulin secretion that is independent of intraislet amyloid formation. (J Diabetes Invest, doi: 10.1111/j.2040-1124.2011.00166.x, 2011).

No MeSH data available.


Related in: MedlinePlus

 Characterization of hIAPP expression in WW mice. (a) Distribution of hIAPPWT mRNA in various tissues from the W/W knock in mouse via RT‐PCR using IAPP (upper panel), insulin (middle panel) and β‐actin (lower panel) primers. (b) Sequences of mouse IAPP, hIAPPWT, and hIAPPS20G mRNAs derived from cDNA sequence of RT‐PCR products of WW and GG mouse pancreas RNAs. Sequences within the boxed region represent the sequences engineered into the mouse genomic IAPP locus. (c) Circulating levels of randomly selected wild‐type and S20G mutant hIAPP and mIAPP in MM, WW and GG mice that were maintained on control diet. The striped area represents the typical range of values from the literature.33–35 Error bars represent S.D.
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f1:  Characterization of hIAPP expression in WW mice. (a) Distribution of hIAPPWT mRNA in various tissues from the W/W knock in mouse via RT‐PCR using IAPP (upper panel), insulin (middle panel) and β‐actin (lower panel) primers. (b) Sequences of mouse IAPP, hIAPPWT, and hIAPPS20G mRNAs derived from cDNA sequence of RT‐PCR products of WW and GG mouse pancreas RNAs. Sequences within the boxed region represent the sequences engineered into the mouse genomic IAPP locus. (c) Circulating levels of randomly selected wild‐type and S20G mutant hIAPP and mIAPP in MM, WW and GG mice that were maintained on control diet. The striped area represents the typical range of values from the literature.33–35 Error bars represent S.D.

Mentions: RNA from various tissues obtained from the WW mice were analyzed by RT‐PCR for the presence of hIAPP and mouse insulin mRNA (Figure 1a). Human IAPPWT and mouse insulin were detected only in the pancreas of WW mice. Identical results were obtained with RT‐PCR analyses of RNA from GG‐KI mice (not shown). We verified the sequences of the human IAPPWT and hIAPPS20G mRNAs via RT‐PCR and cDNA sequencing in the WW and GG mice (Figure 1b). These data confirm the appropriate genotypic and tissue‐specific expression patterns of the WW and GG mice. Using an IAPP‐specific ELISA, which detects rat IAPP (identical to mouse IAPP) and both hIAPPWT and hIAPPS20G equally well13, circulating hIAPP levels were determined in 6 month old male and female animals in the fed state (Figure 1c). No significant differences in circulating IAPP levels were observed in the MM, WW and GG mice.


Expression of wild-type and mutant S20G hIAPP in physiologic knock-in mouse models fails to induce islet amyloid formation, but induces mild glucose intolerance.

Hiddinga HJ, Sakagashira S, Ishigame M, Madde P, Sanke T, Nanjo K, Kudva YC, Lee JJ, van Deursen J, Eberhardt NL - J Diabetes Investig (2012)

 Characterization of hIAPP expression in WW mice. (a) Distribution of hIAPPWT mRNA in various tissues from the W/W knock in mouse via RT‐PCR using IAPP (upper panel), insulin (middle panel) and β‐actin (lower panel) primers. (b) Sequences of mouse IAPP, hIAPPWT, and hIAPPS20G mRNAs derived from cDNA sequence of RT‐PCR products of WW and GG mouse pancreas RNAs. Sequences within the boxed region represent the sequences engineered into the mouse genomic IAPP locus. (c) Circulating levels of randomly selected wild‐type and S20G mutant hIAPP and mIAPP in MM, WW and GG mice that were maintained on control diet. The striped area represents the typical range of values from the literature.33–35 Error bars represent S.D.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4020731&req=5

f1:  Characterization of hIAPP expression in WW mice. (a) Distribution of hIAPPWT mRNA in various tissues from the W/W knock in mouse via RT‐PCR using IAPP (upper panel), insulin (middle panel) and β‐actin (lower panel) primers. (b) Sequences of mouse IAPP, hIAPPWT, and hIAPPS20G mRNAs derived from cDNA sequence of RT‐PCR products of WW and GG mouse pancreas RNAs. Sequences within the boxed region represent the sequences engineered into the mouse genomic IAPP locus. (c) Circulating levels of randomly selected wild‐type and S20G mutant hIAPP and mIAPP in MM, WW and GG mice that were maintained on control diet. The striped area represents the typical range of values from the literature.33–35 Error bars represent S.D.
Mentions: RNA from various tissues obtained from the WW mice were analyzed by RT‐PCR for the presence of hIAPP and mouse insulin mRNA (Figure 1a). Human IAPPWT and mouse insulin were detected only in the pancreas of WW mice. Identical results were obtained with RT‐PCR analyses of RNA from GG‐KI mice (not shown). We verified the sequences of the human IAPPWT and hIAPPS20G mRNAs via RT‐PCR and cDNA sequencing in the WW and GG mice (Figure 1b). These data confirm the appropriate genotypic and tissue‐specific expression patterns of the WW and GG mice. Using an IAPP‐specific ELISA, which detects rat IAPP (identical to mouse IAPP) and both hIAPPWT and hIAPPS20G equally well13, circulating hIAPP levels were determined in 6 month old male and female animals in the fed state (Figure 1c). No significant differences in circulating IAPP levels were observed in the MM, WW and GG mice.

Bottom Line: WW and GW mice maintained on both diets lacked intraislet amyloid at all ages.On both diets relative to MM controls WW and GW mice exhibit glucose intolerance (P < 0.008) with no differences in insulin secretion.However, GW mice secreted significantly more insulin (P < 0.03 that WW mice on both diets throughout the study.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine/Divisions of Endocrinology.

ABSTRACT

Unlabelled: Aims/Introduction:  Human islet polypeptide S20G mutation (hIAPP(S20G)) is associated with earlier onset type 2 diabetes and increased amyloidogenicity and cytotoxicity in vitro vs wild-type hIAPP (hIAPP(WT)), suggesting that amyloidogenesis may be pathogenic for type 2 diabetes. We compared the contributions of hIAPP(S20G) and hIAPP(WT) toward intra islet amyloid formation and development of type 2 diabetes in a unique physiologic knock-in mouse model.

Materials and methods:   We replaced the mouse IAPP gene (M allele) with hIAPP(WT) (W allele) and hIAPP(S20G) (G allele) via homologous recombination and backbred transgenic mice against C57Bl/6 strain 5 generations to minimize genetic variation. Mice (3 month old) were maintained on control (CD) or high fat diet (HFD) for 15 months and studied at 3 month intervals by oral glucose tolerance testing (OGTT) and pancreas histology to assess glucose homeostastis, amyloidogeneisis, islet mass, β cell replication, and apoptosis.

Results:   IAPP blood levels were indistinguishable in all mice. WW and GW mice maintained on both diets lacked intraislet amyloid at all ages. On both diets relative to MM controls WW and GW mice exhibit glucose intolerance (P < 0.008) with no differences in insulin secretion. However, GW mice secreted significantly more insulin (P < 0.03 that WW mice on both diets throughout the study. By 12 months on the high fat diet all mice increased their β cell mass about 3-fold and were indistinguishable.

Conclusions:   Physiologic expression of hIAPP(WT) and hIAPP(S20G) in C57Bl/6 mice produces mild glucose intolerance with inappropriately normal insulin secretion that is independent of intraislet amyloid formation. (J Diabetes Invest, doi: 10.1111/j.2040-1124.2011.00166.x, 2011).

No MeSH data available.


Related in: MedlinePlus