Th17 cells promote pancreatic inflammation but only induce diabetes efficiently in lymphopenic hosts after conversion into Th1 cells
Authors: Yi-Hong Wang
Chen Dong
Seon Hee Chang
Yeonseok Chung
Natalia Martin-Orozco
Journal: European Journal of Immunology (2009)
Discipline: IMMUNOLOGY
Keywords: IDDM; Th17 cells; diabetes; NOD mice; insulitis
Affiliations: Department of Immunology, MD Anderson Cancer Center, Houston, TX 77030, USA.

Abstract:  IDDM is characterized by leukocyte invasion to the pancreatic tissues followed by immune destruction of the islets. Despite the important function of Th17 cells in other autoimmune disease models, their function in IDDM is relatively unclear. In this study, we found association of elevated Th17 cytokine expression with diabetes in NOD mice. To understand the function of Th17 cells in IDDM, we differentiated islet-reactive BDC2.5 TcR transgenic CD4(+) cells in vitro into Th17 cells and transferred them into NOD.scid and neonate NOD mice. NOD.scid recipient mice developed rapid onset of diabetes with extensive insulitic lesions, whereas in newborn NOD mice, despite extensive insulitis, most recipient mice did not develop diabetes. Surprisingly, BDC2.5(+) cells recovered from diabetic NOD.scid mice, in comparison with those from neonate NOD mice, showed predominant IFN-gamma over IL-17 expression, indicating conversion of donor cells into Th1 cells. Moreover, diabetes progression in NOD.scid recipients was dependent on IFN-gamma while anti-IL-17 treatment reduced insulitic inflammation. These results indicate that islet-reactive Th17 cells promote pancreatic inflammation, but only induce IDDM upon conversion into IFN-gamma producers. 

Retracted: No

PubPeer comments:
 In Figure 1B, the FACS for the PLN from Diabetic mice exhibited a different gating compared to all other samples. Since the manuscript performed a statistical comparison between the cell frequency, the gatings should be the same. Could the authors please explain? ![file] Also, Figure 3A showed the same problem for ILN. ![file] , 
 First of all, prior to addressing this specific question, we would like to clarify that there are no guidelines within the immunology research field that FACS gating should be kept exactly the SAME for various samples, especially for samples collected from different tissues, at different time points and or stained with different combinations of intracellular or surface markers. In fact, it is the well-accepted consensus by the research community that gating could be and should be adjusted according to factors including but not limited to controls used and intensities of isotype staining. The scientific principles underlying such adjustments include varying levels of auto-fluorescence and varying degrees of laser compensation. With the above being said, we still spend time to compose comprehensive answers to each specific question on FACS gating with the hope that the user community would witness our good will and support of the Pubpeeru2019s principles of fostering open and honest discussions on scientifically related issues of high values. In this study, we determined IL-17 and IFN-gamma production of cells from lymph nodes and spleen by intracellular staining. The tissues were harvested, cleaned and dissociated to generate single cell suspensions followed by culturing with PMA Ionomycin plus Golgi plug prior to cytokine intracellular staining. The levels of cytokine production and gating strategies for each tissue were set based on the fluorescence levels in apparently cytokine-negative cells. In many cases, the shapes of dot plots of the lymphocyte populations display different patterns among different tissues, which could result from distinct tissue features, tissue fat content, cell activation status, distinct processing procedures, and other tissue debris in the cell suspensions. Therefore, you canu2019t simply set the same gate for cells from different tissues. In addition, diabetic mice were skinny and unhealthy by the time they were euthanized, which usually resulted in cell suspensions of lower quality. Given the significant differences between these animals, we believe that the application of different gating was technically valid and comparisons between young and diabetic mice were correctly made. Similarly, for the NOD-SCID mice that received the Th17 cells, lymph nodes were smaller since the mice were lymphopenic and suffered from diabetes. Therefore, the concerns from the anonymous commenter were not founded. -Natalia Martin-Orozco and Chen Dong ,