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Blood glucose levels regulate pancreatic beta-cell proliferation during experimentally-induced and spontaneous autoimmune diabetes in mice.

Pechhold K, Koczwara K, Zhu X, Harrison VS, Walker G, Lee J, Harlan DM - PLoS ONE (2009)

Bottom Line: For instance, we show that when normoglycemia is restored by exogenous insulin or islet transplantation, the beta-cell proliferation rate returns towards low levels found in control animals, yet surges when hyperglycemia recurs.Rather, disease-associated alterations of BrdU-incorporation rates of delta-cells (minor decrease), and non-endocrine islet cells (slight increase) were not affected by blood glucose levels, or were inversely related to glycemia control after diabetes onset (alpha-cells).We conclude that murine beta-cells' ability to proliferate in response to metabolic need (i.e. rising blood glucose concentrations) is remarkably well preserved during severe, chronic beta-cell autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institutes of Health (NIH), Bethesda, MD, USA. klausp@intra.niddk.nih.gov

ABSTRACT

Background: Type 1 diabetes mellitus is caused by immune-mediated destruction of pancreatic beta-cells leading to insulin deficiency, impaired intermediary metabolism, and elevated blood glucose concentrations. While at autoimmune diabetes onset a limited number of beta-cells persist, the cells' regenerative potential and its regulation have remained largely unexplored. Using two mouse autoimmune diabetes models, this study examined the proliferation of pancreatic islet ss-cells and other endocrine and non-endocrine subsets, and the factors regulating that proliferation.

Methodology and principal findings: We adapted multi-parameter flow cytometry techniques (including DNA-content measurements and 5'-bromo-2'-deoxyuridine [BrdU] incorporation) to study pancreatic islet single cell suspensions. These studies demonstrate that beta-cell proliferation rapidly increases at diabetes onset, and that this proliferation is closely correlated with the diabetic animals' elevated blood glucose levels. For instance, we show that when normoglycemia is restored by exogenous insulin or islet transplantation, the beta-cell proliferation rate returns towards low levels found in control animals, yet surges when hyperglycemia recurs. In contrast, other-than-ss endocrine islet cells did not exhibit the same glucose-dependent proliferative responses. Rather, disease-associated alterations of BrdU-incorporation rates of delta-cells (minor decrease), and non-endocrine islet cells (slight increase) were not affected by blood glucose levels, or were inversely related to glycemia control after diabetes onset (alpha-cells).

Conclusion: We conclude that murine beta-cells' ability to proliferate in response to metabolic need (i.e. rising blood glucose concentrations) is remarkably well preserved during severe, chronic beta-cell autoimmunity. These data suggest that timely control of the destructive immune response after disease manifestation could allow spontaneous regeneration of sufficient beta-cell mass to restore normal glucose homeostasis.

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Glycemia regulates β-cell proliferation in spontaneously autoimmune diabetic NOD mice.A: BrdU uptake by residual β-cells of NOD mice is controlled by BG levels. Newly diabetic NOD mice were randomly treated with: no℞, n = 8; exogenous insulin (pellet) and BrdU labeled day1–3 (early), n = 3, crosshatched grey bar, or BrdU labeled day 4–7 (late), n = 6, hatched; insulin pellet treated mice that did not restore normoglycemia (prim. fail.), n = 4, hatched grey bar, and were compared to pre-diabetic NOD mice, n = 3, open bar. Results are shown as mean±SE, with significance levels: (*) prediabetic: p = 0.016, and insulin pellet (late): p = 0.0051, respectively. B: Correlation plot as in Figure 4C. Each symbol represents a single mouse, and all mice, regardless of treatment group are included. Correlation coefficient r = 0.78; 95% confidence interval (dashed lines). Note, that NOD mice that restored normoglycemia upon insulin pellet, and were immediately BrdU labeled (pellet early) maintain increased β-cell proliferation. These mice are shown in the plot (⧫) but were omitted from correlation coefficient calculation.
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pone-0004827-g006: Glycemia regulates β-cell proliferation in spontaneously autoimmune diabetic NOD mice.A: BrdU uptake by residual β-cells of NOD mice is controlled by BG levels. Newly diabetic NOD mice were randomly treated with: no℞, n = 8; exogenous insulin (pellet) and BrdU labeled day1–3 (early), n = 3, crosshatched grey bar, or BrdU labeled day 4–7 (late), n = 6, hatched; insulin pellet treated mice that did not restore normoglycemia (prim. fail.), n = 4, hatched grey bar, and were compared to pre-diabetic NOD mice, n = 3, open bar. Results are shown as mean±SE, with significance levels: (*) prediabetic: p = 0.016, and insulin pellet (late): p = 0.0051, respectively. B: Correlation plot as in Figure 4C. Each symbol represents a single mouse, and all mice, regardless of treatment group are included. Correlation coefficient r = 0.78; 95% confidence interval (dashed lines). Note, that NOD mice that restored normoglycemia upon insulin pellet, and were immediately BrdU labeled (pellet early) maintain increased β-cell proliferation. These mice are shown in the plot (⧫) but were omitted from correlation coefficient calculation.

Mentions: We also measured the β-cell proliferation rate in the commonly studied NOD mouse, a spontaneous autoimmune diabetes model. As shown in Figure 6, β-cells from newly diabetic NOD mice exhibited increased proliferation (5.5±0.9% at disease onset up from 0.9±0.3% of randomly selected non (or pre)-diabetic NOD mice, Figure 6, left panel). This mitotic activity largely decreased to 1.5±0.5% following sustained exogenous insulin-induced normoglycemia (insulin-pellet). Overall, NOD mouse β-cell proliferation rates also correlated well with pre-euthanasia BG levels (Figure 6 right panel, r = 0.78). Interestingly, while restoring normal BG concentrations in our mice with CTL-induced diabetes (EAD model) rapidly restored normal β-cell replication rates, NOD mouse β-cell proliferation displayed a delayed response to insulin treatment (Figure 6, right panel, n = 3 grey triangles). That is, NOD mouse β-cell proliferation rates did not decrease until normal blood concentrations were restored for 3–4 days (Figure 6, hatched open bar, open diamonds, n = 6). The few EAD model mice that failed treatment to normalized BG levels following exogenous insulin (pellets) and anti-CD8 antibody injections (Figure 4C, 6, and data not shown), presumably because they had lost too many β-cells prior to treatment, also maintained increased β-cell proliferation. More interestingly, short term insulin replacement therapy in diabetic NOD mice reversed hyperglycemia, yet regularly failed to promptly normalize β-cell proliferation (Figure 6, pellet, early). To restore lower β-cell proliferation rates to diabetic NOD mice required several days of insulin-induced euglycemia (Figure 6, pellet, late). Taken together, β-cell proliferation rates generally correlated well with recent BG concentrations as objectively quantified using flow cytometry analysis.


Blood glucose levels regulate pancreatic beta-cell proliferation during experimentally-induced and spontaneous autoimmune diabetes in mice.

Pechhold K, Koczwara K, Zhu X, Harrison VS, Walker G, Lee J, Harlan DM - PLoS ONE (2009)

Glycemia regulates β-cell proliferation in spontaneously autoimmune diabetic NOD mice.A: BrdU uptake by residual β-cells of NOD mice is controlled by BG levels. Newly diabetic NOD mice were randomly treated with: no℞, n = 8; exogenous insulin (pellet) and BrdU labeled day1–3 (early), n = 3, crosshatched grey bar, or BrdU labeled day 4–7 (late), n = 6, hatched; insulin pellet treated mice that did not restore normoglycemia (prim. fail.), n = 4, hatched grey bar, and were compared to pre-diabetic NOD mice, n = 3, open bar. Results are shown as mean±SE, with significance levels: (*) prediabetic: p = 0.016, and insulin pellet (late): p = 0.0051, respectively. B: Correlation plot as in Figure 4C. Each symbol represents a single mouse, and all mice, regardless of treatment group are included. Correlation coefficient r = 0.78; 95% confidence interval (dashed lines). Note, that NOD mice that restored normoglycemia upon insulin pellet, and were immediately BrdU labeled (pellet early) maintain increased β-cell proliferation. These mice are shown in the plot (⧫) but were omitted from correlation coefficient calculation.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2654100&req=5

pone-0004827-g006: Glycemia regulates β-cell proliferation in spontaneously autoimmune diabetic NOD mice.A: BrdU uptake by residual β-cells of NOD mice is controlled by BG levels. Newly diabetic NOD mice were randomly treated with: no℞, n = 8; exogenous insulin (pellet) and BrdU labeled day1–3 (early), n = 3, crosshatched grey bar, or BrdU labeled day 4–7 (late), n = 6, hatched; insulin pellet treated mice that did not restore normoglycemia (prim. fail.), n = 4, hatched grey bar, and were compared to pre-diabetic NOD mice, n = 3, open bar. Results are shown as mean±SE, with significance levels: (*) prediabetic: p = 0.016, and insulin pellet (late): p = 0.0051, respectively. B: Correlation plot as in Figure 4C. Each symbol represents a single mouse, and all mice, regardless of treatment group are included. Correlation coefficient r = 0.78; 95% confidence interval (dashed lines). Note, that NOD mice that restored normoglycemia upon insulin pellet, and were immediately BrdU labeled (pellet early) maintain increased β-cell proliferation. These mice are shown in the plot (⧫) but were omitted from correlation coefficient calculation.
Mentions: We also measured the β-cell proliferation rate in the commonly studied NOD mouse, a spontaneous autoimmune diabetes model. As shown in Figure 6, β-cells from newly diabetic NOD mice exhibited increased proliferation (5.5±0.9% at disease onset up from 0.9±0.3% of randomly selected non (or pre)-diabetic NOD mice, Figure 6, left panel). This mitotic activity largely decreased to 1.5±0.5% following sustained exogenous insulin-induced normoglycemia (insulin-pellet). Overall, NOD mouse β-cell proliferation rates also correlated well with pre-euthanasia BG levels (Figure 6 right panel, r = 0.78). Interestingly, while restoring normal BG concentrations in our mice with CTL-induced diabetes (EAD model) rapidly restored normal β-cell replication rates, NOD mouse β-cell proliferation displayed a delayed response to insulin treatment (Figure 6, right panel, n = 3 grey triangles). That is, NOD mouse β-cell proliferation rates did not decrease until normal blood concentrations were restored for 3–4 days (Figure 6, hatched open bar, open diamonds, n = 6). The few EAD model mice that failed treatment to normalized BG levels following exogenous insulin (pellets) and anti-CD8 antibody injections (Figure 4C, 6, and data not shown), presumably because they had lost too many β-cells prior to treatment, also maintained increased β-cell proliferation. More interestingly, short term insulin replacement therapy in diabetic NOD mice reversed hyperglycemia, yet regularly failed to promptly normalize β-cell proliferation (Figure 6, pellet, early). To restore lower β-cell proliferation rates to diabetic NOD mice required several days of insulin-induced euglycemia (Figure 6, pellet, late). Taken together, β-cell proliferation rates generally correlated well with recent BG concentrations as objectively quantified using flow cytometry analysis.

Bottom Line: For instance, we show that when normoglycemia is restored by exogenous insulin or islet transplantation, the beta-cell proliferation rate returns towards low levels found in control animals, yet surges when hyperglycemia recurs.Rather, disease-associated alterations of BrdU-incorporation rates of delta-cells (minor decrease), and non-endocrine islet cells (slight increase) were not affected by blood glucose levels, or were inversely related to glycemia control after diabetes onset (alpha-cells).We conclude that murine beta-cells' ability to proliferate in response to metabolic need (i.e. rising blood glucose concentrations) is remarkably well preserved during severe, chronic beta-cell autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institutes of Health (NIH), Bethesda, MD, USA. klausp@intra.niddk.nih.gov

ABSTRACT

Background: Type 1 diabetes mellitus is caused by immune-mediated destruction of pancreatic beta-cells leading to insulin deficiency, impaired intermediary metabolism, and elevated blood glucose concentrations. While at autoimmune diabetes onset a limited number of beta-cells persist, the cells' regenerative potential and its regulation have remained largely unexplored. Using two mouse autoimmune diabetes models, this study examined the proliferation of pancreatic islet ss-cells and other endocrine and non-endocrine subsets, and the factors regulating that proliferation.

Methodology and principal findings: We adapted multi-parameter flow cytometry techniques (including DNA-content measurements and 5'-bromo-2'-deoxyuridine [BrdU] incorporation) to study pancreatic islet single cell suspensions. These studies demonstrate that beta-cell proliferation rapidly increases at diabetes onset, and that this proliferation is closely correlated with the diabetic animals' elevated blood glucose levels. For instance, we show that when normoglycemia is restored by exogenous insulin or islet transplantation, the beta-cell proliferation rate returns towards low levels found in control animals, yet surges when hyperglycemia recurs. In contrast, other-than-ss endocrine islet cells did not exhibit the same glucose-dependent proliferative responses. Rather, disease-associated alterations of BrdU-incorporation rates of delta-cells (minor decrease), and non-endocrine islet cells (slight increase) were not affected by blood glucose levels, or were inversely related to glycemia control after diabetes onset (alpha-cells).

Conclusion: We conclude that murine beta-cells' ability to proliferate in response to metabolic need (i.e. rising blood glucose concentrations) is remarkably well preserved during severe, chronic beta-cell autoimmunity. These data suggest that timely control of the destructive immune response after disease manifestation could allow spontaneous regeneration of sufficient beta-cell mass to restore normal glucose homeostasis.

Show MeSH
Related in: MedlinePlus