Line 93: | Line 93: | ||
</div> | </div> | ||
<h3 class="ar-title"><span class="dg"> </span>Summary of Mathematical Models</h3> | <h3 class="ar-title"><span class="dg"> </span>Summary of Mathematical Models</h3> | ||
− | <h4><br>In order to further evaluate the clinical value of our system and provide a theoretical reference for the next in vivo and preclinical experiments, we also establish a systematic mathematical model for the local and overall immune environment of rheumatoid arthritis (See the Model section for details). Thus, we have a theoretical explanation of the relationships among the cytokines, different cell subpopulations and rheumatoid arthritis, which lays a solid foundation for further improvement of our research ideas regarding the subsequent in vivo and preclinical experiments</h4> | + | <h4><br>In order to further evaluate the clinical value of our system and provide a theoretical reference for the next in vivo and preclinical experiments, we also establish a systematic mathematical model for the local and overall immune environment of rheumatoid arthritis (See the Model section for details). Thus, we have a theoretical explanation of the relationships among the cytokines, different cell subpopulations and rheumatoid arthritis, which lays a solid foundation for further improvement of our research ideas regarding the subsequent in vivo and preclinical experiments.</h4> |
<h3 class="ar-title"><span class="dg"> </span>RESULTS</h3> | <h3 class="ar-title"><span class="dg"> </span>RESULTS</h3> | ||
<h2><br>1.The construction and expressing validation of SynNotch and CAR system</h2> | <h2><br>1.The construction and expressing validation of SynNotch and CAR system</h2> | ||
Line 115: | Line 115: | ||
<h4><br>Although we demonstrated the function of the SynNotch system and the CAR system in the Flag-FOXP3-Jurkat cell line, the results obtained from our experiments still have some limitations: Although Flag-FOXP3-Jurkat cells are able to stimulate the functional characteristics of T cells on the core transcription factors and its posttranslational modification, Flag-FOXP3-Jurkat cell line, as a simplified Treg model, does not fully exhibit the functional characteristics of Treg cells in other respects, such as cell surface receptor types and cytokine types. Therefore, it is only suitable for studying the mechanism of transcription, translation and post-translational modification on the upstream genes of Treg cells. But we can prove that with the cytokines IL-17A and IL-6 stimulation, our SynNotch system and CAR system can work effectively based on our experimental data. Specifically, the SynNotch system activated the expression of the USP7 gene under the stimulation of IL-17A and IL-6 and enhanced the stability of FOXP3 protein. And its intensity increased with the increase of IL-17A concentration. FOXP3 protein stability level can be equivalent to Treg function level to a large extent, so we can initially confirm that our overall idea and experimental design was correct.</h4> | <h4><br>Although we demonstrated the function of the SynNotch system and the CAR system in the Flag-FOXP3-Jurkat cell line, the results obtained from our experiments still have some limitations: Although Flag-FOXP3-Jurkat cells are able to stimulate the functional characteristics of T cells on the core transcription factors and its posttranslational modification, Flag-FOXP3-Jurkat cell line, as a simplified Treg model, does not fully exhibit the functional characteristics of Treg cells in other respects, such as cell surface receptor types and cytokine types. Therefore, it is only suitable for studying the mechanism of transcription, translation and post-translational modification on the upstream genes of Treg cells. But we can prove that with the cytokines IL-17A and IL-6 stimulation, our SynNotch system and CAR system can work effectively based on our experimental data. Specifically, the SynNotch system activated the expression of the USP7 gene under the stimulation of IL-17A and IL-6 and enhanced the stability of FOXP3 protein. And its intensity increased with the increase of IL-17A concentration. FOXP3 protein stability level can be equivalent to Treg function level to a large extent, so we can initially confirm that our overall idea and experimental design was correct.</h4> | ||
<br> | <br> | ||
− | <h4><br>In order to further determine the reliability of our experimental design, we also need to conduct a series of experiments. First of all, for the CAR system, we need to further verify its inhibition of CD20 + B lymphocytes, and the completion of these experiments requires primary T lymphocytes. The reconstructed HEAD-Tregs need to be cultivated together with primary CD20 + B lymphocytes and we also need to verify their inhibitory function by flow cytometry and ELISA. The next main problems we need to solve are the induction of primary Treg cell differentiation and primary Treg cell transfection. We will gradually optimize the experimental conditions to carry out the experiments | + | <h4><br>In order to further determine the reliability of our experimental design, we also need to conduct a series of experiments. First of all, for the CAR system, we need to further verify its inhibition of CD20 + B lymphocytes, and the completion of these experiments requires primary T lymphocytes. The reconstructed HEAD-Tregs need to be cultivated together with primary CD20 + B lymphocytes and we also need to verify their inhibitory function by flow cytometry and ELISA. The next main problems we need to solve are the induction of primary Treg cell differentiation and primary Treg cell transfection. We will gradually optimize the experimental conditions to carry out the experiments |
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + | ||
− | + |
Latest revision as of 02:52, 2 November 2017