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. 2024 May 18;15:4232. doi: 10.1038/s41467-024-48611-8

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© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — SPF Rag1^-/- and GF Rag1^-/- B6 mice were recipients for transfer of immune cells or/and gut microbiota from KO and Ctr donor mice. A The experimental procedure of immune cell adoptive transfer to SPF Rag1^-/- B6 mice. SPF Rag1^-/- B6 mice (4 weeks of age) were adoptively transferred (i.v.) 1 × 10⁷ total splenocytes from Tlr9^fl/fl/Cd19Cre^+/- (KO) and Tlr9^fl/fl/Cd19Cre^-/-B6 (Ctr) male donor mice (12–16 weeks post HFD). Two weeks after the immune cell transfer, the recipient Rag1^-/- B6 mice (6 wks of age) were started with high fat diet. IPGTT and ITT were assessed when the mice were at 13 and 14 weeks of age, respectively. Graph was created with BioRender.com. B, C Body weight change and net body weight gain of the SPF Rag1^-/- B6 mice transferred with the immune cells from KO and Ctr donors (Rag_KO: n = 7 mice, Rag_Ctr: n = 8 mice). D Fasting glucose of the SPF Rag1^-/- B6 recipient mice transferred with the immune cells from KO and control mice (Rag_KO: n = 9 mice, Rag_Ctr: n = 6 mice). E IPGTT result of the SPF Rag1^-/- B6 recipient mice transferred with immune cells from KO and Ctr donor mice (Rag_KO: n = 9 mice, Rag_Ctr: n = 6 mice). F Area under the curve (AUC) of IPGTT results from D. G ITT result of the SPF Rag1^-/- recipient B6 mice transferred with immune cells from KO and control donors (Rag_KO: n = 4 mice, Rag_Ctr: n = 6 mice). H Area under the curve (AUC) of the ITT of the results from F. I The experimental procedure of immune cell adoptive transfer and fecal microbiota transfer (FMT) to germ free Rag1^-/- B6 mice. Germ free Rag1^-/- B6 mice (4 weeks of age) were adoptively transferred (i.v.) 1 × 10⁷ total splenocytes and gavaged with 1 × 10⁸ gut microbiota from the pooled stool samples from Tlr9^fl/fl/Cd19Cre^+/- (KO) or Tlr9^fl/fl/Cd19Cre^-/-B6 (Ctr) male donor mice (12–16 weeks post HFD). The recipient mice were maintained on normal diet. IPGTT was assessed 4 weeks after the FMT and immune cell transfer. The experiment was terminated 5 weeks after the FMT and immune cell transfer. Graph was created with BioRender.com. J IPGTT result of GF Rag1^-/- B6 mice transferred with the immune cells from KO and Ctr donors and colonized with the bacteria from the same KO and Ctr donor mice (n = 5 mice per group). K Area under the curve (AUC) of the IPGTT of the results from J. All the experiments were repeated twice with similar results. B, C, E, G, J were analyzed using two-way ANOVA. K was analyzed using one-way ANOVA. D, F, H were analyzed using two-tailed Student’s t-test. The data are presented as mean ± SD.