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Genetic deficiency of Itgb2 or ItgaL prevents autoimmune diabetes through distinctly different mechanisms in NOD/LtJ mice.

Glawe JD, Patrick DR, Huang M, Sharp CD, Barlow SC, Kevil CG - Diabetes (2009)

Bottom Line: However, ItgaL deficiency did not alter NOD T-cell adhesion to or transmigration across islet endothelial cells.Adoptive transfer of ItgaL-deficient splenocytes into NOD/Rag-1 mice did not result in development of diabetes, suggesting a role for ItgaL in NOD/LtJ T-cell activation.Together, these data demonstrate that genetic deficiency of Itgb2 or ItgaL confers protection against autoimmune diabetes through distinctly different mechanisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Louisiana State University Health Sciences Center Shreveport, Shreveport, Louisiana, USA.

ABSTRACT

Objective: Insulitis is an important pathological feature of autoimmune diabetes; however, mechanisms governing the recruitment of diabetogenic T-cells into pancreatic islets are poorly understood. Here, we determined the importance of leukocyte integrins beta(2)(Itgb2) and alphaL (ItgaL) in developing insulitis and frank diabetes.

Research design and methods: Gene-targeted mutations of either Itgb2 or ItgaL were established on the NOD/LtJ mouse strain. Experiments were performed to measure insulitis and diabetes development. Studies were also performed measuring mutant T-cell adhesion to islet microvascular endothelial cells under hydrodynamic flow conditions. T-cell adhesion molecule profiles and adoptive transfer studies were also performed.

Results: Genetic deficiency of either Itgb2 or ItgaL completely prevented the development of hyperglycemia and frank diabetes in NOD mice. Loss of Itgb2 or ItgaL prevented insulitis with Itgb2 deficiency conferring complete protection. In vitro hydrodynamic flow adhesion studies also showed that loss of Itgb2 completely abrogated T-cell adhesion. However, ItgaL deficiency did not alter NOD T-cell adhesion to or transmigration across islet endothelial cells. Adoptive transfer of ItgaL-deficient splenocytes into NOD/Rag-1 mice did not result in development of diabetes, suggesting a role for ItgaL in NOD/LtJ T-cell activation.

Conclusions: Together, these data demonstrate that genetic deficiency of Itgb2 or ItgaL confers protection against autoimmune diabetes through distinctly different mechanisms.

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Related in: MedlinePlus

Genetic deficiency of Itgb2 or ItgaL alters the phenotype of NOD/LtJ CD3 T-cells. Whole splenocytes were isolated from wild-type pre-diabetic (12 weeks old), wild-type diabetic (18 weeks old), Itgb2−/−  (18 weeks old), and ItgaL−/−  (18 weeks old) NOD/LtJ mice and stained for CD3 and individual adhesion molecules to determine the percentage of cells expressing various molecules. A: The percent of CD3 T-cells that are positive for CD11a expression. B: The percent of CD3 T-cells that are positive for CD18 expression. C: The percent of CD3 T-cells that are positive for CD49d expression. D: The percent of CD3 T-cells that are positive for CD29 expression. E: The percent of CD3 T-cells that are positive for CD62L expression. F: The percent of CD3 T-cells that are positive for LPAM-1 expression. *P < 0.01 vs. wild-type diabetic mice; # P < 0.01 Itgb2  vs. ItgaL  mice, n = 6–7 animals per genotype.
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Figure 2: Genetic deficiency of Itgb2 or ItgaL alters the phenotype of NOD/LtJ CD3 T-cells. Whole splenocytes were isolated from wild-type pre-diabetic (12 weeks old), wild-type diabetic (18 weeks old), Itgb2−/− (18 weeks old), and ItgaL−/− (18 weeks old) NOD/LtJ mice and stained for CD3 and individual adhesion molecules to determine the percentage of cells expressing various molecules. A: The percent of CD3 T-cells that are positive for CD11a expression. B: The percent of CD3 T-cells that are positive for CD18 expression. C: The percent of CD3 T-cells that are positive for CD49d expression. D: The percent of CD3 T-cells that are positive for CD29 expression. E: The percent of CD3 T-cells that are positive for CD62L expression. F: The percent of CD3 T-cells that are positive for LPAM-1 expression. *P < 0.01 vs. wild-type diabetic mice; # P < 0.01 Itgb2 vs. ItgaL mice, n = 6–7 animals per genotype.

Mentions: Figure 2 reports the percent of CD3 T-cells that express various surface adhesion molecules between wild-type, Itgb2 , and ItgaL NOD/LtJ mice. Adhesion molecule expression analysis was performed using pre-diabetic wild-type mice at 12 weeks of age and diabetic wild-type and mutant mice at 18 weeks of age. As expected, genetic deficiency of Itgb2 eliminates CD18 and CD11a surface expression (Fig. 2A and B). Interestingly, genetic deficiency of ItgaL does not completely abolish CD18 surface expression, indicating the presence of other β2integrins on the T-cell surface (Fig. 2B). This is an important observation because previous studies have assumed that the only relevant Itgb2 expressed on the T-cell surface is CD18/CD11a and that genetic deficiency of CD18 serves as a surrogate for knockout of CD11a (23,24). Our data demonstrate that this is clearly not the case and that genetic disruption of ItgaL still results in a Itgb2 phenotype in NOD/LtJ mice, highlighting that these molecules are not interchangeable. Deficiency of ItgaL or Itgb2 did not significantly alter CD49d positivity (Fig. 2C); however, deficiency of Itgb2 did significantly increase CD29 (β1 integrin) positivity (Fig. 2D). Interestingly, genetic deficiency of Itgb2 significantly decreased the number of CD3 T-cells positive for CD62L (Fig. 2E), suggesting differential regulation of adhesion molecule expression between ItgaL and Itgb2.


Genetic deficiency of Itgb2 or ItgaL prevents autoimmune diabetes through distinctly different mechanisms in NOD/LtJ mice.

Glawe JD, Patrick DR, Huang M, Sharp CD, Barlow SC, Kevil CG - Diabetes (2009)

Genetic deficiency of Itgb2 or ItgaL alters the phenotype of NOD/LtJ CD3 T-cells. Whole splenocytes were isolated from wild-type pre-diabetic (12 weeks old), wild-type diabetic (18 weeks old), Itgb2−/−  (18 weeks old), and ItgaL−/−  (18 weeks old) NOD/LtJ mice and stained for CD3 and individual adhesion molecules to determine the percentage of cells expressing various molecules. A: The percent of CD3 T-cells that are positive for CD11a expression. B: The percent of CD3 T-cells that are positive for CD18 expression. C: The percent of CD3 T-cells that are positive for CD49d expression. D: The percent of CD3 T-cells that are positive for CD29 expression. E: The percent of CD3 T-cells that are positive for CD62L expression. F: The percent of CD3 T-cells that are positive for LPAM-1 expression. *P < 0.01 vs. wild-type diabetic mice; # P < 0.01 Itgb2  vs. ItgaL  mice, n = 6–7 animals per genotype.
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Figure 2: Genetic deficiency of Itgb2 or ItgaL alters the phenotype of NOD/LtJ CD3 T-cells. Whole splenocytes were isolated from wild-type pre-diabetic (12 weeks old), wild-type diabetic (18 weeks old), Itgb2−/− (18 weeks old), and ItgaL−/− (18 weeks old) NOD/LtJ mice and stained for CD3 and individual adhesion molecules to determine the percentage of cells expressing various molecules. A: The percent of CD3 T-cells that are positive for CD11a expression. B: The percent of CD3 T-cells that are positive for CD18 expression. C: The percent of CD3 T-cells that are positive for CD49d expression. D: The percent of CD3 T-cells that are positive for CD29 expression. E: The percent of CD3 T-cells that are positive for CD62L expression. F: The percent of CD3 T-cells that are positive for LPAM-1 expression. *P < 0.01 vs. wild-type diabetic mice; # P < 0.01 Itgb2 vs. ItgaL mice, n = 6–7 animals per genotype.
Mentions: Figure 2 reports the percent of CD3 T-cells that express various surface adhesion molecules between wild-type, Itgb2 , and ItgaL NOD/LtJ mice. Adhesion molecule expression analysis was performed using pre-diabetic wild-type mice at 12 weeks of age and diabetic wild-type and mutant mice at 18 weeks of age. As expected, genetic deficiency of Itgb2 eliminates CD18 and CD11a surface expression (Fig. 2A and B). Interestingly, genetic deficiency of ItgaL does not completely abolish CD18 surface expression, indicating the presence of other β2integrins on the T-cell surface (Fig. 2B). This is an important observation because previous studies have assumed that the only relevant Itgb2 expressed on the T-cell surface is CD18/CD11a and that genetic deficiency of CD18 serves as a surrogate for knockout of CD11a (23,24). Our data demonstrate that this is clearly not the case and that genetic disruption of ItgaL still results in a Itgb2 phenotype in NOD/LtJ mice, highlighting that these molecules are not interchangeable. Deficiency of ItgaL or Itgb2 did not significantly alter CD49d positivity (Fig. 2C); however, deficiency of Itgb2 did significantly increase CD29 (β1 integrin) positivity (Fig. 2D). Interestingly, genetic deficiency of Itgb2 significantly decreased the number of CD3 T-cells positive for CD62L (Fig. 2E), suggesting differential regulation of adhesion molecule expression between ItgaL and Itgb2.

Bottom Line: However, ItgaL deficiency did not alter NOD T-cell adhesion to or transmigration across islet endothelial cells.Adoptive transfer of ItgaL-deficient splenocytes into NOD/Rag-1 mice did not result in development of diabetes, suggesting a role for ItgaL in NOD/LtJ T-cell activation.Together, these data demonstrate that genetic deficiency of Itgb2 or ItgaL confers protection against autoimmune diabetes through distinctly different mechanisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Louisiana State University Health Sciences Center Shreveport, Shreveport, Louisiana, USA.

ABSTRACT

Objective: Insulitis is an important pathological feature of autoimmune diabetes; however, mechanisms governing the recruitment of diabetogenic T-cells into pancreatic islets are poorly understood. Here, we determined the importance of leukocyte integrins beta(2)(Itgb2) and alphaL (ItgaL) in developing insulitis and frank diabetes.

Research design and methods: Gene-targeted mutations of either Itgb2 or ItgaL were established on the NOD/LtJ mouse strain. Experiments were performed to measure insulitis and diabetes development. Studies were also performed measuring mutant T-cell adhesion to islet microvascular endothelial cells under hydrodynamic flow conditions. T-cell adhesion molecule profiles and adoptive transfer studies were also performed.

Results: Genetic deficiency of either Itgb2 or ItgaL completely prevented the development of hyperglycemia and frank diabetes in NOD mice. Loss of Itgb2 or ItgaL prevented insulitis with Itgb2 deficiency conferring complete protection. In vitro hydrodynamic flow adhesion studies also showed that loss of Itgb2 completely abrogated T-cell adhesion. However, ItgaL deficiency did not alter NOD T-cell adhesion to or transmigration across islet endothelial cells. Adoptive transfer of ItgaL-deficient splenocytes into NOD/Rag-1 mice did not result in development of diabetes, suggesting a role for ItgaL in NOD/LtJ T-cell activation.

Conclusions: Together, these data demonstrate that genetic deficiency of Itgb2 or ItgaL confers protection against autoimmune diabetes through distinctly different mechanisms.

Show MeSH
Related in: MedlinePlus