biotin-streptavidine-PC5

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Presentation transcript:

biotin-streptavidine-PC5 Supplementary Table 1: antibodies used for flow cytometry Analyze of hematopoietic cells: Target protein label Clone Laboratory CD45 Horizon V450 30F11 BD Pharmingen CD11c Biotin-PE-Texas Red N418 eBiosciences CD11b APC-Cy7 M1/70 anti-MHC-II IA/IE Alexa 700 M5:114 CD115 PE-YG AFS98 mPDCA APC JF05-1C2.4.1 Miltenyi Gr1 PerCy5.5 RB6-8C5 CD8a PE-Cy5 53-6.7 CD3 Biotin-PE-Texas red 145-2C11 CD4 FITC RM4-5 CD44 IM7 FR4 12A5 CD62-L MEL-14 NK1.1 biotin-streptavidine-PC5 PK136 Analyze of adipose tissue stromal vascular fraction (SVF) cells: Target protein label Clone Laboratory CD45 Horizon V450 30F11 BD Pharmingen CD11c Biotin-PE-Texas Red N418 eBiosciences CD11b APC-Cy7 M1/70 anti-MHC-II IA/IE Alexa 700 M5:114 F4/80 PE-Cy5 BM8 Mgl1 (CD301) ER-MP23 Serotec

A B C * D Adipocyte size WT P<0.001 *** CX3CR1-/- WT CX3CR1-/- Adipocyte size (µm) CX3CR1-/- WT CX3CR1-/- B C * p=0.01 OGTT CX3CR1 -/- Control Food intake WT CX3CR1-/- g of food/mice/day ** Glucose (mg/dL) Adipose tissue weight (g) Total WAT SC WAT perigonadal WAT CX3CR1-/- donor WT donor * D Supplemental Figure 1: Food intake, OGTT and adipocyte size in female mice CX3CR1-/- and adipose tissue weights in female transplanted with BM of CX3CR1-/- mice (A) Adipocyte size in female mice CX3CR1-/- and their controls C57Bl/6 fed a HFD (60%fat) for 30 weeks (n=5 mice/genotype) (B) Oral Glucose Tolerance Test (OGTT) in female mice CX3CR1-/- and their controls C57Bl/6 fed a HFD (60%fat) for 26 weeks (n=5 mice/genotype) (C) Food intake (g of food per mice and per day) in female mice CX3CR1-/- and their controls C57Bl/6 fed a HFD (60%fat) (n=6 mice/genotype) (D): AT weights of recipient mice of CX3CR1-/- BM or WT BM after 22 weeks of HFD with 60%Kcal from fat. *, p<0.05. **, p<0.01. ***, p<0.001. Data are expressed as means ±SD.

A B C Gr1low monocytes (%CD45) Bodyweight (g) r2=0.24 p=0.03 r2=0.53 p=0.0032 r2=0.41 p<0.0001 Bodyweight (g) Bodyweight (g) Gr1low monocytes (%CD45) Gr1low monocytes (%CD45) Supplemental Figure 2: Correlation between the rate of Gr1low monocytes and bodyweight in mice totally deficient for CX3CR1 or deficient for CX3CR1 in the hematopoietic compartment A: Correlation between BW and the rate of circulating Gr1low monocytes (%CD45) in mice deficient for CX3CR1 in the hematopoietic compartment (recipients of CX3CR1-/- BM) and their controls (recipients of WT BM) fed a HFD (n=18 mice). B: Correlation between BW and the rate of circulating Gr1low monocytes (%CD45) in mice totally deficient for CX3CR1 (CX3CR1-/- mice) and their controls fed a HFD (n=14 mice). C: Correlation between BW and the rate of circulating Gr1low monocytes (%CD45) in all mice deficient for CX3CR1 (totally or in the hematopoietic compartment) and their controls fed a HFD (n=32 mice).

Gr1high monocytes (%CD45) Bodyweight (g) r=0.2 p=0.1 Gr1high monocytes (%CD45) Supplemental Figure 3: Correlation between bodyweight and the rate of circulating Gr1high monocytes (% CD45) in mice deficient for CX3CR1 (CX3CR1-/- and recipients of CX3CR1-/- BM) and their controls (respectively C57Bl/6 and controls BM) fed a HFD (r=0.2, p=0.1, n=31 mice).

A B *** WT donor CD11c-hBcl2 donor Adipose tissue weight (g) Adipocyte size (µm) WT donor CD11c-hBcl2 A *** Adipose tissue weight (g) Total WAT SC WAT perigonadal WAT CD11c-hBcl2 donor WT donor B Supplemental Figure 4: Adipose tissue weights of mice transplanted with CX3CR1-/- BM or CD11c-hBcl2 BM and adipocyte size of mice transplanted with CD11c-hBcl2 BM (A): AT weights of recipient mice of CX3CR1-/- BM or WT BM after 22 weeks of HFD with 60%Kcal from fat. (B): AT weights of recipient mice of CD11c-hBcl2 BM or WT BM after 22 weeks of HFD with 60%Kcal from fat. (C): Adipocyte size in recipient mice of CD11c-hBcl2 BM fed a HFD for 12 weeks; *** p<0.001. WAT= white adipose tissue, SC= subcuteanous.

A B C D E CX3CR1 -/- Control Supplemental Figure 5: Other circulating cells in CX3CR1-/- mice (CD4 T cells, activated CD4 T cells, NK cells, dendritic cells and neutrophils) (A) The frequency of T lymphocytes (CD3+, CD4+) was determined by flow cytometry in C57Bl/6 (white histograms) or CX3CR1-/- mice (black histograms) fed a HFD containing 60 % kcal from fat for 30 weeks, as in Figure 1. **, p<0.01. n =9 mice in C57Bl/6 and n= 12 CX3CR1 -/-. (B) The frequency of activated T lymphocytes (CD3+, CD4+,CD62- and CD44high) was determined in the same mice than in A. (C) The frequency of circulating dendritic cells (CD11chigh, CMH-II+, and CD115-) was determined in the same mice than in A. (D) The frequency of circulating NK cells (NK1.1+, CD45+)) was determined in the same mice than in A. (E) The frequency of circulating neutrophils (CD11b+, Gr1high and CD115-) was determined in the same mice than in A. Plots show the mean values (±SD).

Supplemental Figure 6: Gating strategy to isolate the subpopulation of Gr1high and Gr1low monocytes in the blood of female mice upon HFD diet.