Role of TG2, TG2-specific B cells and gluten-specific B cells in celiac disease
1 Purpose
Transglutaminase 2 (TG2) is thought to have a critical role in celiac disease, and the enzyme is also the target of disease-specific autoantibodies. The mechanisms, kinetics and location of TG2 activation, and how the antibody response to TG2 and gluten is induced, are not understood, and will be studied in this project. In particular, the involvement of small intestinal epithelial cells and epithelial-cell derived TG2 will be studied.
2 Distress
Most animals in this project will only suffer from minimal distress for short time due to the intravenous, intraperitoneal, subcuteaneous or oral administration of cells or antigen, in the presence of the adjuvants cholera toxin or CFA, as well as due to withdrawal of blood. In a previous amendment submitted April 2022, a small group of mice will suffer moderate distress due to overnight fasting. The increase in experiment duration and extra blood samples will have minimal negative affect on mouse distress.
3 Expected benefit
A better understanding is crucial for our understanding of celiac disease pathogenesis, and will enable the development of improved drugs that inhibit TG2 activity in patients. With the current changes we further develop our models to achieve these goals.
4 Number of animals, and what kind
The number of mice on this protocol will not change and remains 2311 mice in total. Wild type C57Bl/6 mice, TG2KO mice, and humanized mice expressing the celiac disease risk allele HLA-DQ2, and mice expressing celiac patient derived gluten-specific T-cell receptors, or B-cell receptors recognizing TG2 and gluten. In addition, we will make conditional KO mice for TG2 specifically in enterocytes using Villin-Cre and TG2-flox/flox transgenic mice. In the 4th Amendment (August 2022) we have added a new conditional knock in mouse to study the role of epithelial cells in the induction of immune responses in celiac disease: Vil1-rtTA*M2 TetO-TG2w18 trangenic mice.
5 How to adhere to 3R
We will minimize the amount of distress by keeping injection volumes low. Oral gavages, intravenous injections and blood sampling will be performed by well trained staff. We also use the minimal number of mice to get valid answers in our experiments.
Transglutaminase 2 (TG2) is thought to have a critical role in celiac disease, and the enzyme is also the target of disease-specific autoantibodies. The mechanisms, kinetics and location of TG2 activation, and how the antibody response to TG2 and gluten is induced, are not understood, and will be studied in this project. In particular, the involvement of small intestinal epithelial cells and epithelial-cell derived TG2 will be studied.
2 Distress
Most animals in this project will only suffer from minimal distress for short time due to the intravenous, intraperitoneal, subcuteaneous or oral administration of cells or antigen, in the presence of the adjuvants cholera toxin or CFA, as well as due to withdrawal of blood. In a previous amendment submitted April 2022, a small group of mice will suffer moderate distress due to overnight fasting. The increase in experiment duration and extra blood samples will have minimal negative affect on mouse distress.
3 Expected benefit
A better understanding is crucial for our understanding of celiac disease pathogenesis, and will enable the development of improved drugs that inhibit TG2 activity in patients. With the current changes we further develop our models to achieve these goals.
4 Number of animals, and what kind
The number of mice on this protocol will not change and remains 2311 mice in total. Wild type C57Bl/6 mice, TG2KO mice, and humanized mice expressing the celiac disease risk allele HLA-DQ2, and mice expressing celiac patient derived gluten-specific T-cell receptors, or B-cell receptors recognizing TG2 and gluten. In addition, we will make conditional KO mice for TG2 specifically in enterocytes using Villin-Cre and TG2-flox/flox transgenic mice. In the 4th Amendment (August 2022) we have added a new conditional knock in mouse to study the role of epithelial cells in the induction of immune responses in celiac disease: Vil1-rtTA*M2 TetO-TG2w18 trangenic mice.
5 How to adhere to 3R
We will minimize the amount of distress by keeping injection volumes low. Oral gavages, intravenous injections and blood sampling will be performed by well trained staff. We also use the minimal number of mice to get valid answers in our experiments.