The current demographic shift toward an aging population has led to a robust increase in the prevalence of age-associated metabolic disorders. Recent studies have demonstrated that the etiology of obesity-related insulin resistance that develops with aging differs from that induced by high-calorie diets. Whereas the role of adaptive immunity in changes in energy metabolism driven by nutritional challenges has recently gained attention, its impact on aging remains mostly unknown. Here we found that the number of follicular B2 lymphocytes and expression of the B-cell-specific transcriptional coactivator OcaB increase with age in spleen and in intra-abdominal epididymal white adipose tissue (eWAT), concomitantly with higher circulating levels of IgG and impaired glucose homeostasis. Reduction of B-cell maturation and Ig production—especially that of IgG2c—by ablation of OcaB prevented age-induced glucose intolerance and insulin resistance and promoted energy expenditure by stimulating fatty acid utilization in eWAT and brown adipose tissue. Transfer of wild-type bone marrow in OcaB−/− mice replenished the eWAT B2-cell population and IgG levels, which diminished glucose tolerance, insulin sensitivity, and energy expenditure while increasing body weight gain in aged mice. Thus these findings demonstrate that upon aging, modifications in B-cell-driven adaptive immunity contribute to glucose intolerance and fat accretion.

Recent studies have stressed the importance of the adaptive immune response in the pathogenesis of insulin resistance in diet-induced obesity (DIO) (1). In white adipose tissue (WAT), adipocytes are surrounded by cells from the innate and adaptive immune systems that modulate inflammatory status (24). In DIO models, the recruitment of B lymphocytes in WAT precedes the infiltration of inflammatory CD4+ and CD8+ T cells and macrophages (5,6), leading to Th1 polarization (7,8) and the production of IgG autoantibodies by B2 cells, which are major factors contributing to the development of insulin resistance (9). In contrast, B-regulatory (10), B1-a (11) and B1-b (12) cell subtypes have, through the secretion of IgM autoantibodies, been shown to favor insulin sensitization in the context of DIO.

Aging is characterized by a redistribution and accumulation of lipids in visceral and ectopic adipose depots, reduced energy expenditure, and decreased glucose tolerance (13). However, the etiology of insulin resistance that develops with aging has been suggested to differ from that induced by DIO (14). Notably, studies have recently shown that, along with many aspects of the immune response that are altered during aging (15), the relative frequency of fat-resident T regulatory cells is increased 12-fold in visceral WAT of 11-month-old mice compared with that of 3-month-old animals (14). Blocking this accumulation prevented age- but not DIO-induced insulin resistance (14). Whether other cells from the immune system also play a role in this context is not yet established.

The B-cell-specific nuclear cofactor Oct coactivator from B cells (OcaB, or Pou2af1) is essential for transcriptional control of B-lymphocyte maturation, antibody repertoire development, and Ig production (notably IgG) (1620). Expression of OcaB upon activation of CD4+ T cells also contributes to memory generation (21). Accordingly, OcaB−/− mice are characterized by an abnormal transition of B cells and reduced antigen-stimulated Ig production (18,22). In this study we first tested the hypothesis that aging is associated with an accumulation of B2 cells in WAT. Then, because OcaB primarily affects follicular B2 cells, and because of the central role of OcaB in IgG production and the detrimental effect of IgG on insulin sensitivity, we investigated whether hampering B-lymphocyte activity through OcaB ablation would attenuate development of age-induced fat accumulation and insulin resistance.

Animal Experiments

OcaB−/− mice on a mixed C57BL/129 genetic background (The Jackson Laboratory) were described previously (18). Before the experiments described herein, mice were backcrossed 9–11 times on a pure C57BL/6J background. Breeding pairs consisted of a heterozygous (OcaB+/−) male and females to produce OcaB+/+, OcaB+/−, and OcaB−/− mice in every litter. In all experiments, OcaB+/+ littermates were used as controls for comparison with OcaB−/− animals.

All mice were housed alone and fed a standard chow diet (Harlan diet #2018), with free access to food and water. A group of OcaB+/+ animals were fed daily according to the amount of food eaten by OcaB−/− mice (pair-feeding experiment) and had free access to water. Other animals were weighed weekly, and food consumption was recorded. Mice were exposed to a 12-h dark/12-h light cycle and kept under an ambient temperature (23 ± 1°C). Animals were cared for and handled according to the Canadian Guide for the Care and Use of Laboratory Animals. The Laval University Animal Care and Use Committee approved the procedures.

Chimeras were produced in 8-week-old mice, as previously described (23). After whole-body irradiation, mice were reconstituted with bone marrow harvested from the femur of 7-week-old OcaB+/+ (wild-type [WT]) or OcaB−/− (knockout [KO]) mice by injecting 3.2 × 106 cells into the tail vein. Quantification of chimerism was evaluated as described previously (24), by amplifying the WT and the KO alleles from isolated splenic B cells (EasySep; STEMCELL Technologies, Vancouver, Canada).

Human Tissue Collection

Omental and subcutaneous WAT samples were obtained during ongoing bariatric surgery at the Institut universitaire de cardiologie et de pneumologie de Québec. The study included obese men (mean BMI 55 ± 11 kg/m2), described previously (25), aged 18.8–65.8 years; these men were divided into three age groups, with mean ages ± SEMs of 23.4 ± 0.6, 40.0 ± 0.4, and 58.9 ± 1.5 years. Approval was obtained from the ethics committee of the Institut universitaire de cardiologie et de pneumologie de Québec. All subjects provided written informed consent before their inclusion in the study.

Immune Challenge

OcaB+/+ and OcaB−/− mice were administered Methanosphaera stadtmanae intranasally for 3 weeks, exactly as previously described (26). Lung leukocytes were isolated from the right lobes and digested with collagenase. T- and B-cell frequencies were assessed by fluorescence-activated cell sorting (FACS).

FACS Analysis

Visceral epididymal WAT (eWAT), subcutaneous inguinal (iWAT), and interscapular brown adipose tissue (BAT) depots were minced and digested with collagenase II in Krebs buffer at 37°C for 45 min. The digested tissues were filtered through 40-µm cell strainers, and the stromal vascular fraction (SVF) was centrifuged to a pellet. The SVF was then washed with Krebs buffer. Splenic cells were collected by mechanical disruption, filtered, and washed with cold PBS. All cells were conserved on ice in PBS and 1% FBS until FACS.

Splenic cells were incubated in a CD19-APC, CD23-FITC, and CD93-BV421 antibody cocktail and a CD5-BV421, CD19-APC, IgM-PE, and IgD-PE Cy7 cocktail for subcharacterization. SVF cells were incubated in antibody cocktail 1 (CD5-BV421, CD19-APC, IgM-PE, IgD-PE Cy7), cocktail 2 (CD4-BV605 and CD8-PB), and cocktail 3 (F4/80-PB, CD11b-AF700, and CD11c-PE/Cy7) to characterize B and T lymphocytes and macrophages, respectively. Forward- and side-scattered signals were used to determine populations. CD19+ B lymphocytes were characterized as stage T1 (CD23CD93+), stage T2/3 (CD23+CD93+), or follicular (CD23+CD93). Among CD45+ cells, lymphocytes were characterized as B1-a (CD19+CD5+IgM+IgD), B1-b (CD19+CD5IgM+IgD), or B2 (CD19+CD5IgMIgD+). The total number of cells, obtained by manual counting using a hemacytometer, was multiplied by frequencies obtained by FACS to compute the absolute number of specific subsets. CD93-BV421 and CD5-BV421 antibodies were from BD Biosciences and the others were from BioLegend. Analyses were conducted on a FORTESSA cytometer (BD Biosciences) using FlowJo software version X (Tree Star, Inc.). Details of the gating strategy are illustrated in Supplementary Fig. 1.

Glucose Metabolism

Glucose tolerance was evaluated in mice fasted for 12 h and injected intraperitoneally with 2 g/kg d-glucose. Insulin sensitivity was evaluated in mice fasted for 5 h and injected with 0.75 units/kg insulin (Novolin; Novo Nordisk). Glycemia was measured in blood from the tail vein using an Accu-Chek glucometer (Roche).

Euglycemic-hyperinsulinemic clamps were conducted exactly as previously described (27). Mice received saline-washed erythrocytes from donor mice to prevent hematocrit from decreasing ≥5% as a result of blood sampling. Glucose production and disposal were determined using the non–steady-state equations for a two-compartment model developed by Mari (28).

In Vivo Analysis of Constitutive Glucose and Palmitate Uptake

After an overnight fast, mice received an intraperitoneal bolus of 14C-bromopalmitate and [3H]-deoxyglucose (10 µCi of each), as previously described (29,30). Tissues were collected 2 h later. Specific fractional uptake data were expressed as the percentage of the injected dose per milligram of tissue.

Indirect Calorimetry

Mice were placed in metabolic chambers (AccuScan Instruments, Inc., Columbus, OH) and acclimated for 72 h with free access to food and water. Then, oxygen consumption (Vo2) and carbon dioxide production (Vco2) were measured every 15 min over 24 h. Locomotor activity was estimated using a grid of invisible infrared light beams.

DEXA

Anesthetized mice were placed in a Lunar PIXImus2 DEXA instrument (General Electric). Body composition was calculated by computerized absorptiometry.

Oxygen Bomb Calorimetry

Two- and 6-month-old mice were moved to new cages, and feces were collected over a 24-h period. Energy density of air-dried feces was determined using bomb calorimetry with a 6100 Calorimeter (Parr Instrument Company).

RNA Extraction and Real-time Quantitative PCR Analysis

Total RNA was extracted using an Aurum Total RNA Fatty and Fibrous Tissue Kit (Bio-Rad). Quantitative PCR was achieved with SYBR Green Jumpstart Taq ReadyMix (Sigma) and Rox as the reference dye using a CFX96 (Bio-Rad). Gene expression was determined by the standard curve method and normalized to the expression of a reference gene that did not differ between conditions. Supplementary Table 1 lists the PCR primers.

Histology

Small sections of iWAT, eWAT, and BAT were fixed in 4% paraformaldehyde and embedded in paraffin. Sections (4 µm thick) were stained using hematoxylin and eosin. Images were taken using an Olympus BX51 microscope. Adipocyte size was scored with Image J software on two images of the same dimensions per depot per mouse, using three mice per genotype.

Plasma Bioassays

Blood was collected from the submandibular vein into a tube containing EDTA, and plasma was stored at −80°C. Nonesterified fatty acids (NEFA), cholesterol, triglycerides, and glucose were measured by colorimetric assays (Randox Laboratories). Plasma insulin and leptin (ALPCO); IgG, IgG2c, IgE, and IgM (Bethyl Laboratories); FGF21 (BioVendor); and IL-6 (R&D Systems) levels were quantified by ELISA. Other plasma and tissue cytokines were quantified by BioPlex (Bio-Rad).

Microarray Experiment

Gene expression was evaluated in iWAT of 6-month-old mice following an overnight fast. Whole-genome gene expression was evaluated using Illumina MouseWG-6 v2.0 Expression BeadChips and a standard Illumina protocol (Génome Québec Innovation Centre). Raw data were quantile-normalized after log-2 transformation using the lumi package in R version 3.1.3 (31,32). The t tests was used to identify probes differentially expressed between groups (P value set at 1% and absolute fold change threshold set at 2). Analyses were carried out the R statistical software and the Bioconductor package (33). Biological pathways were evaluated for 95 differentially expressed genes using Ingenuity Pathway Analysis (Qiagen).

Statistical Analysis

Data are expressed as mean ± SEM. One-way, two-way, or repeated-measures ANOVA, followed by the Fisher post hoc test, if appropriate, was performed to detect statistical differences. A P value <0.05 was considered significant. Linear regression analyses were performed to identify correlations between IgG levels and glucose tolerance. Statistical analyses were performed with GraphPad Prism software (GraphPad Software, Inc.).

We first quantified immune cells by FACS in male C57BL/6J mice that were aging naturally. Between the ages of 6 and 12 months, the frequencies of B lymphocytes increased in the spleen and eWAT, but not in iWAT (Fig. 1A–C). In eWAT of 12-month-old mice, B cells represented close to 20% of all cells from the SVF (Fig. 1B). Age-induced changes were linked to a shift in B-cell maturation, with reduced frequencies of T1 (Fig. 1D) and T2/3 transitional B cells (Fig. 1E) but increased numbers of follicular B cells (Fig. 1F). The increased B-cell frequency in eWAT was largely due to a higher total number of B2 lymphocytes, and of B1-a and B1-b cells, although to a smaller extent (Fig. 1G). By contrast, aging had no effect on B1-a, B1-b, or B2 cell numbers in iWAT (Fig. 1H) or in BAT (Supplementary Fig. 2C). In addition, aging did not modify the global frequencies of WAT-infiltrated CD4+ and CD8+ T cells and macrophages (Supplementary Fig. 2A and B). These results suggest that aging is associated with a gradual augmentation of circulating mature B2 lymphocytes, which accumulate preferentially in eWAT.

Figure 1

Expansion of circulating B2 lymphocytes and accumulation in eWAT parallels age-associated insulin resistance and augmentation of IgG levels. AC: Effects of aging on the frequency of CD19+ B lymphocytes in spleen (A), eWAT (B), and iWAT (C) of male C57BL/6J mice (n = 3 mice/group). D–F: Effects of aging on transition stage T1 (CD23CD93+) (D), T2/3 (CD23+CD93+) (E), and follicular (CD23+CD93) (F) splenic B cells (n = 3 mice/group). G and H: Counts of B1-a (CD19+CD5+IgM+IgD), B1-b (CD19+CD5IgM+IgD), and B2 (CD19+CD5IgMIgD+) lymphocyte subtypes in eWAT (G) and iWAT (H) (n = 4 mice/group). I: Effects of aging on OcaB expression in WAT in men (n = 6–10/group), according to mean ± SEM ages of 23 ± 1, 40 ± 1, and 59 ± 1 years. J: Impact of aging on OcaB expression in WAT of C57BL/6J mice (n = 11–14/group). K: Irradiation (Irrad.) nullifies OcaB expression in WAT of 2-month-old C57BL/6J mice (n = 3–4/group). LO: Ig levels upon aging in male C57BL/6J mice (n = 6–10/group). P: Intraperitoneal glucose tolerance test (IPGTT) in male C57BL/6J mice at 2, 6, and 12 months old (n = 6–13 mice/group). Q: Regression analysis between blood IgG levels and glucose response (area under the curve [AUC] during the IPGTT shown in O). *P < 0.05; **P < 0.01; ***P < 0.0001. Ctrl, control; Vis, visceral adipose tissue; SC, subcutaneous adipose tissue.

Figure 1

Expansion of circulating B2 lymphocytes and accumulation in eWAT parallels age-associated insulin resistance and augmentation of IgG levels. AC: Effects of aging on the frequency of CD19+ B lymphocytes in spleen (A), eWAT (B), and iWAT (C) of male C57BL/6J mice (n = 3 mice/group). D–F: Effects of aging on transition stage T1 (CD23CD93+) (D), T2/3 (CD23+CD93+) (E), and follicular (CD23+CD93) (F) splenic B cells (n = 3 mice/group). G and H: Counts of B1-a (CD19+CD5+IgM+IgD), B1-b (CD19+CD5IgM+IgD), and B2 (CD19+CD5IgMIgD+) lymphocyte subtypes in eWAT (G) and iWAT (H) (n = 4 mice/group). I: Effects of aging on OcaB expression in WAT in men (n = 6–10/group), according to mean ± SEM ages of 23 ± 1, 40 ± 1, and 59 ± 1 years. J: Impact of aging on OcaB expression in WAT of C57BL/6J mice (n = 11–14/group). K: Irradiation (Irrad.) nullifies OcaB expression in WAT of 2-month-old C57BL/6J mice (n = 3–4/group). LO: Ig levels upon aging in male C57BL/6J mice (n = 6–10/group). P: Intraperitoneal glucose tolerance test (IPGTT) in male C57BL/6J mice at 2, 6, and 12 months old (n = 6–13 mice/group). Q: Regression analysis between blood IgG levels and glucose response (area under the curve [AUC] during the IPGTT shown in O). *P < 0.05; **P < 0.01; ***P < 0.0001. Ctrl, control; Vis, visceral adipose tissue; SC, subcutaneous adipose tissue.

Because OcaB is closely related to B2-lymphocyte maturation and Ig-producing activity, we measured its levels upon aging. It is interesting to note that, in both mice and humans, aging was concomitant with an upward modulation of OcaB mRNA levels in WAT (Fig. 1I and J). Increased OcaB expression was likely due to changes occurring in B cells, because irradiation prevented the detection of OcaB in iWAT and eWAT (Fig. 1K). Moreover, OcaB expression seems to be undetectable in cultured mouse 3T3-L1 and human SW872 adipocytes (data not shown).

Consistent with the expansion of follicular B2 cells upon aging, plasma levels of IgG and IgG2c, but not of IgM or IgE, increased with age (Fig. 1L–O). The robust glucose intolerance developed upon aging correlated with circulating IgG concentrations (Fig. 1P and Q). Taken together, these findings indicate that age-induced insulin resistance is linked with an adaptive immune phenotype skewed toward IgG-producing B2 cells both in the circulation and within eWAT.

We then tested the hypothesis that altering B2-lymphocyte activity by ablating the OcaB gene would attenuate the impact of aging on insulin resistance. It is important to note that backcrossing the KO line on a pure C57BL/6J genetic background resulted in the expected defects in B cells, as OcaB−/− mice failed to respond appropriately to a chronic M. stadtmanae immune challenge; they were incompetent with increasing B-cell numbers at the site of infection (Supplementary Fig. 3). In unchallenged 6-month-old OcaB−/− mice, we observed an age-associated reduction in splenic transitional and follicular B cells, but a small relative increase in marginal zone B cells compared with those of their OcaB+/+ littermates (Fig. 2A). Frequencies of CD4+ and CD8+ T cells and macrophages in WAT were similar between OcaB+/+ and OcaB−/− mice (Supplementary Fig. 2D and E). Ablation of OcaB had no major impact on the accumulation of B1-a or B1-b cells in either WAT depots, whereas it resulted in a large reduction in B2 cells in iWAT (Fig. 2B), eWAT (Fig. 2C), and BAT (Supplementary Fig. 2F). In iWAT, these B cells both surrounded adipocytes and were distributed within clusters found ubiquitously in the depot (Fig. 2D). The absence of OcaB also resulted in reduced basal systemic levels of IgE and IgM, and a slight diminution of IgG levels (Fig. 2E–G). OcaB−/− mice had no detectable circulating IgG2c (Fig. 2H).

Figure 2

Lack of OcaB impairs age-associated B-lymphocyte accumulation in WAT. AC: Transitional state of splenic B lymphocytes (A) and accumulation of B cells in iWAT (B) and eWAT (C) in 6-month-old OcaB+/+ and OcaB−/− mice (n = 3/group). D: Presence of CD19+ B cells (arrows) in iWAT (magnification ×200) and in one lymph node (magnification ×40) present within iWAT of OcaB+/+ mice. EH: Blood levels of Igs in 6-month-old OcaB+/+ and OcaB−/− mice (n = 5/group). *P < 0.05; **P < 0.01; ***P < 0.0001. Foll, follicular; KO, knockout (OcaB−/−); MZ, marginal zone; N.D., none detected; NS, not significant; WT, wild type (OcaB+/+ littermates).

Figure 2

Lack of OcaB impairs age-associated B-lymphocyte accumulation in WAT. AC: Transitional state of splenic B lymphocytes (A) and accumulation of B cells in iWAT (B) and eWAT (C) in 6-month-old OcaB+/+ and OcaB−/− mice (n = 3/group). D: Presence of CD19+ B cells (arrows) in iWAT (magnification ×200) and in one lymph node (magnification ×40) present within iWAT of OcaB+/+ mice. EH: Blood levels of Igs in 6-month-old OcaB+/+ and OcaB−/− mice (n = 5/group). *P < 0.05; **P < 0.01; ***P < 0.0001. Foll, follicular; KO, knockout (OcaB−/−); MZ, marginal zone; N.D., none detected; NS, not significant; WT, wild type (OcaB+/+ littermates).

At the age of 2 months, OcaB−/− mice had glucose tolerance comparable to that of their OcaB+/+ littermates (Fig. 3A). It is striking that 6-month-old OcaB−/− mice did not develop age-associated glucose intolerance (Fig. 3B) and insulin resistance (Fig. 3C and D). Consistent with this, 6-month-old OcaB−/− mice had lower fasting glycemia and insulinemia than their OcaB+/+ littermates (Table 1). Moreover, hyperinsulinemic-euglycemic clamp experiments revealed that OcaB−/− mice were highly sensitive to insulin (Fig. 3E). OcaB−/− animals showed at least a twofold higher insulin-stimulated glucose disposal rate (Fig. 3F), despite similar hepatic glucose production (Fig. 3G), than that of their age-matched OcaB+/+ littermates. This enhanced insulin-stimulated glucose uptake notably manifested in iWAT and skeletal muscle (Fig. 3H). Taken together, these findings strongly suggest that the absence of OcaB blocks the development of age-induced insulin resistance.

Figure 3

Absence of OcaB protects against age-induced insulin resistance. A and B: Intraperitoneal glucose tolerance test in 2-month-old (A) and 6-month-old (B) OcaB+/+ and OcaB−/− mice (n = 11–13/group). C and D: Insulin tolerance test in 2-month-old (C) and 6-month-old (D) OcaB+/+ and OcaB−/− mice (n = 11–13/group). E–H: Rates of glucose infusion (E), glucose disposal (Rd) (F), glucose appearance (Ra) (G), and glucose uptake (H) in 6-month-old OcaB+/+ and OcaB−/− mice (n = 5–6/group) during a euglycemic-hyperinsulinemic clamp consisting of a 5-μCi bolus of D-[3-3H]glucose given at the beginning, followed by infusion of 0.05 μCi/min during a 90-min tracer equilibration period. The clamp then began with a primed continuous infusion of human insulin (a bolus of 16 mU/kg body weight followed by 4 mU/kg body weight/min). The D-[3-3H]glucose infusion was increased to 0.2 μCi/min for the rest of the 120-min experimental period. Euglycemia was achieved by infusing a 20% dextrose solution as necessary. *P < 0.05; ***P < 0.0001. GIR, glucose infusion rate; KO, knockout (OcaB−/−); WT, wild type (OcaB+/+ littermates).

Figure 3

Absence of OcaB protects against age-induced insulin resistance. A and B: Intraperitoneal glucose tolerance test in 2-month-old (A) and 6-month-old (B) OcaB+/+ and OcaB−/− mice (n = 11–13/group). C and D: Insulin tolerance test in 2-month-old (C) and 6-month-old (D) OcaB+/+ and OcaB−/− mice (n = 11–13/group). E–H: Rates of glucose infusion (E), glucose disposal (Rd) (F), glucose appearance (Ra) (G), and glucose uptake (H) in 6-month-old OcaB+/+ and OcaB−/− mice (n = 5–6/group) during a euglycemic-hyperinsulinemic clamp consisting of a 5-μCi bolus of D-[3-3H]glucose given at the beginning, followed by infusion of 0.05 μCi/min during a 90-min tracer equilibration period. The clamp then began with a primed continuous infusion of human insulin (a bolus of 16 mU/kg body weight followed by 4 mU/kg body weight/min). The D-[3-3H]glucose infusion was increased to 0.2 μCi/min for the rest of the 120-min experimental period. Euglycemia was achieved by infusing a 20% dextrose solution as necessary. *P < 0.05; ***P < 0.0001. GIR, glucose infusion rate; KO, knockout (OcaB−/−); WT, wild type (OcaB+/+ littermates).

Table 1

Fasting blood parameters in 6-month-old OcaB−/− male mice and their OcaB+/+ littermates

ParameterOcaB+/+OcaB−/−P value
Glucose (mmol/L) 10.5 ± 0.82 8.9 ± 1.46 0.003 
Insulin (ng/L) 1.41 ± 0.51 0.53 ± 0.26 0.004 
NEFA (mmol/L) 0.77 ± 0.33 0.68 ± 0.23 0.442 
Triglycerides (mmol/L) 0.92 ± 0.28 0.85 ± 0.29 0.561 
Cholesterol (mmol/L) 3.26 ± 0.84 3.24 ± 0.70 0.953 
Leptin (pg/mL) 903 ± 590.9 71.8 ± 27.1 0.006 
IL-1α (pg/mL) 4.74 ± 2.66 5.02 ± 2.05 0.845 
IL-3 (pg/mL) 13.52 ± 11.84 4.335 ± 1.18 0.088 
IL-10 (pg/mL) 50.3 ± 27.88 40.6 ± 34.16 0.571 
IL-13 (pg/mL) 115.5 ± 70.18 73.70 ± 15.03 0.182 
MIP-1β (pg/mL) 15.88 ± 10.59 9.14 ± 1.44 0.152 
ParameterOcaB+/+OcaB−/−P value
Glucose (mmol/L) 10.5 ± 0.82 8.9 ± 1.46 0.003 
Insulin (ng/L) 1.41 ± 0.51 0.53 ± 0.26 0.004 
NEFA (mmol/L) 0.77 ± 0.33 0.68 ± 0.23 0.442 
Triglycerides (mmol/L) 0.92 ± 0.28 0.85 ± 0.29 0.561 
Cholesterol (mmol/L) 3.26 ± 0.84 3.24 ± 0.70 0.953 
Leptin (pg/mL) 903 ± 590.9 71.8 ± 27.1 0.006 
IL-1α (pg/mL) 4.74 ± 2.66 5.02 ± 2.05 0.845 
IL-3 (pg/mL) 13.52 ± 11.84 4.335 ± 1.18 0.088 
IL-10 (pg/mL) 50.3 ± 27.88 40.6 ± 34.16 0.571 
IL-13 (pg/mL) 115.5 ± 70.18 73.70 ± 15.03 0.182 
MIP-1β (pg/mL) 15.88 ± 10.59 9.14 ± 1.44 0.152 

Data are mean ± SD. Each group contained six mice.

Because glucose homeostasis is highly sensitive to changes in body composition, we investigated the impact of OcaB deletion on energy balance. We observed that OcaB−/− mice had a lower body weight than their OcaB+/+ littermates and showed resistance to age-induced weight gain (Fig. 4A) and fat accretion (Fig. 4B and C). OcaB−/− mice consumed less food than (Fig. 4D), but had fecal energy density (Fig. 4E) and physical activity (Fig. 4F) levels similar to, the levels observed in OcaB+/+ littermates. The lean phenotype of OcaB−/− animals was reflected by low leptin levels (Table 1). By contrast, circulating levels of NEFA, triglycerides, and cholesterol did not significantly differ between OcaB+/+ and OcaB−/− mice (Table 1). It is remarkable that OcaB−/− mice still gained less body weight than their pair-fed OcaB+/+ littermates (Fig. 4G). This resulted in significantly lower food efficiency than that of pair-fed OcaB+/+ animals (Fig. 4H). These findings suggest that the low body weight gain caused by the absence of OcaB is partly due to enhanced energy expenditure.

Figure 4

Absence of OcaB protects against age-induced fat accretion. The graphs show body weight (A), fat mass (B), tissue weight (C), food intake (D), fecal energy density (E), and locomotor activity (F) in 2- and 6-month-old male OcaB+/+ and OcaB−/− mice (n = 10–19/group). G and H: Age-dependent body weight gain (G) and food efficiency (H) in OcaB+/+ mice. *P < 0.05; **P < 0.01; ***P < 0.0001. KO, knockout (OcaB−/−); PF, pair-fed; WT, wild type (OcaB+/+ littermates).

Figure 4

Absence of OcaB protects against age-induced fat accretion. The graphs show body weight (A), fat mass (B), tissue weight (C), food intake (D), fecal energy density (E), and locomotor activity (F) in 2- and 6-month-old male OcaB+/+ and OcaB−/− mice (n = 10–19/group). G and H: Age-dependent body weight gain (G) and food efficiency (H) in OcaB+/+ mice. *P < 0.05; **P < 0.01; ***P < 0.0001. KO, knockout (OcaB−/−); PF, pair-fed; WT, wild type (OcaB+/+ littermates).

Indirect calorimetry experiments were performed to test this possibility further. Six-month-old OcaB−/− mice had higher oxygen consumption (Fig. 5A), higher respiratory quotient (P = 0.03; data not shown), and higher core body temperature than OcaB+/+ mice (Fig. 5B). In addition, we found evidence of enhanced metabolic activity in adipose depots of aged OcaB−/− mice, including macroscopically darker BAT and iWAT (Fig. 5D), smaller adipocyte size in eWAT and iWAT (Fig. 5D), and the occurrence of multilocular-like adipocytes in iWAT (Fig. 5D). In line with this, OcaB−/− mice were characterized by higher constitutive uptakes of bromopalmitate in eWAT and BAT and of glucose in eWAT (Fig. 5E and F) than those of their OcaB+/+ littermates. The constitutive uptake of both energy substrates in skeletal muscle was similar between genotypes (data not shown). The metabolic and histological modifications in iWAT of OcaB−/− mice were paralleled by significant changes in the gene expression profile toward energy production and browning—notably the gene coding for nicotinamide N-methyltransferase (Nnmt), which was downregulated 53-fold in OcaB−/− mice (Supplementary Fig. 4A). Nnmt expression was also significantly lower (a 59% reduction) in eWAT of OcaB−/− mice than in that of OcaB+/+ animals (Supplementary Fig. 4B). Taken together, these findings suggest that impaired B-cell activity due to a lack of OcaB prompts energy expenditure by increasing energy uptake and utilization in adipose depots.

Figure 5

OcaB−/− mice show stimulated energy expenditure, as indicated by indirect calorimetry (A), random body temperature (B), tissue appearance (C) and histology (D), and tissue uptake of bromo-palmitate (E) and glucose (F) in 6-month-old male OcaB+/+ and OcaB−/− mice (n = 5–8/group). To determine adipocyte size, 622 and 1,091 adipocytes in iWAT and 485 and 634 adipocytes in eWAT were analyzed within histological slides of three OcaB+/+ and three OcaB−/− mice. In panel D, magnification is ×200. *P < 0.05; **P < 0.01. KO, knockout (OcaB−/−); WT, wild type (OcaB+/+ littermates).

Figure 5

OcaB−/− mice show stimulated energy expenditure, as indicated by indirect calorimetry (A), random body temperature (B), tissue appearance (C) and histology (D), and tissue uptake of bromo-palmitate (E) and glucose (F) in 6-month-old male OcaB+/+ and OcaB−/− mice (n = 5–8/group). To determine adipocyte size, 622 and 1,091 adipocytes in iWAT and 485 and 634 adipocytes in eWAT were analyzed within histological slides of three OcaB+/+ and three OcaB−/− mice. In panel D, magnification is ×200. *P < 0.05; **P < 0.01. KO, knockout (OcaB−/−); WT, wild type (OcaB+/+ littermates).

Compared with OcaB+/+ animals, OcaB−/− mice had chronically elevated circulating levels of interleukin (IL)-6 (Fig. 6A), in part due to higher mRNA expression in skeletal muscle (Fig. 6B). OcaB−/− mice also had lower plasma levels of the proinflammatory cytokines tumor necrosis factor (TNF)-α, keratinocyte-derived chemokine (KC), and macrophage inflammatory protein–related protein (MIP)-1α than their OcaB+/+ littermates (Fig. 6C–E). OcaB−/− mice had levels of circulating FGF21 similar to those of OcaB +/+ mice (Fig. 6F). Finally, OcaB−/− mice had a higher protein level of IL-10 locally in eWAT (2.61 ± 0.30 pg/mL, vs. 1.53 ± 0.10 pg/mL in OcaB+/+ mice; P = 0.018), although the circulating concentration remained unchanged (Table 1).

Figure 6

Ablation of OcaB robustly modulates cytokine levels in vivo. A and B: Levels of IL-6 in 6-month-old OcaB+/+ and OcaB−/− mice (n = 5) in plasma (A) and in tissues (B). C: Plasma levels of TNF-α, KC, and MIP-1α in 6-month-old OcaB +/+ and OcaB −/− mice (n = 6). D: Plasma FGF21 levels in 6-month-old OcaB +/+ and OcaB −/− mice (n = 10). *P < 0.05. KO, knockout (OcaB−/−); NS, not significant; WT, wild type (OcaB+/+ littermates).

Figure 6

Ablation of OcaB robustly modulates cytokine levels in vivo. A and B: Levels of IL-6 in 6-month-old OcaB+/+ and OcaB−/− mice (n = 5) in plasma (A) and in tissues (B). C: Plasma levels of TNF-α, KC, and MIP-1α in 6-month-old OcaB +/+ and OcaB −/− mice (n = 6). D: Plasma FGF21 levels in 6-month-old OcaB +/+ and OcaB −/− mice (n = 10). *P < 0.05. KO, knockout (OcaB−/−); NS, not significant; WT, wild type (OcaB+/+ littermates).

To validate further the concept that a lack of OcaB affects energy balance and insulin sensitivity through B cells, bone marrow was transferred between irradiated 2-month-old OcaB+/+ and OcaB−/− mice (Fig. 7A). Our irradiation protocol resulted in almost complete destruction of CD19+ B cells in blood and adipose depots (Supplementary Fig. 5). The transfer of bone marrow resulted in at least 86% chimerism at the age of 6 months (Fig. 7B). As expected, the low B-cell counts in OcaB−/− animals were restored to normal by transfer of OcaB+/+ bone marrow (Fig. 7C and Supplementary Fig. 6A), which also reinstated the typical B-cell transitional state (Fig. 7D).

Figure 7

Transfer of WT (OcaB+/+) bone marrow to OcaB−/− (KO) irradiated mice attenuates energy expenditure, triggering weight gain and insulin resistance. A: OcaB+/+ and OcaB−/− mice were irradiated at the age of 2 months, and bone marrow was transferred from donor WT or KO, resulting in four groups, indicated in pairs as recipient genotype/transferred bone marrow genotype. B and C: Evaluation of chimerism (B) and splenic total (C) and T2/3 (D) B lymphocytes in the 6-month-old chimeric mice described in A. E–J: Weight gain (E), oxygen consumption (F and G), food intake (H), and insulin sensitivity (I and J) in the 6-month-old chimeric mice described in A. K and L: Infiltration of B1-a, B1-b, and B2 lymphocytes in eWAT (K) and iWAT (L) in the 6-month-old chimeric mice described in A. MO: Blood levels of Igs in the 6-month-old mice described in A. P: Regression analysis between blood IgG levels and glucose response (area under the curve [AUC] during the insulin tolerance test [ITT] shown in I) (n = 5 mice/group). *P < 0.05; **P < 0.01. Different lowercase letters above the bars indicate statistically different values. BMT, bone marrow transfer.

Figure 7

Transfer of WT (OcaB+/+) bone marrow to OcaB−/− (KO) irradiated mice attenuates energy expenditure, triggering weight gain and insulin resistance. A: OcaB+/+ and OcaB−/− mice were irradiated at the age of 2 months, and bone marrow was transferred from donor WT or KO, resulting in four groups, indicated in pairs as recipient genotype/transferred bone marrow genotype. B and C: Evaluation of chimerism (B) and splenic total (C) and T2/3 (D) B lymphocytes in the 6-month-old chimeric mice described in A. E–J: Weight gain (E), oxygen consumption (F and G), food intake (H), and insulin sensitivity (I and J) in the 6-month-old chimeric mice described in A. K and L: Infiltration of B1-a, B1-b, and B2 lymphocytes in eWAT (K) and iWAT (L) in the 6-month-old chimeric mice described in A. MO: Blood levels of Igs in the 6-month-old mice described in A. P: Regression analysis between blood IgG levels and glucose response (area under the curve [AUC] during the insulin tolerance test [ITT] shown in I) (n = 5 mice/group). *P < 0.05; **P < 0.01. Different lowercase letters above the bars indicate statistically different values. BMT, bone marrow transfer.

After transfer, weight gain was similar in OcaB+/+ (WT) mice harboring either WT (WT recipient/WT marrow) or OcaB−/− (WT/KO) bone marrow (Fig. 7E). By contrast, OcaB−/− mice with OcaB+/+ bone marrow (KO/WT) gained significantly more weight (Fig. 7E). This phenotype was associated with a reduction in energy expenditure to levels similar to those of OcaB+/+ animals (Fig. 7F and G). However, food intake in OcaB−/− mice was comparable after transfer of either OcaB+/+ (KO/WT) or OcaB−/− (KO/KO) marrow (Fig. 7H). Consistent with the changes in body weight, insulin sensitivity was significantly decreased after transfer of OcaB+/+ bone marrow to OcaB−/− animals (Fig. 7I and J), whereas it remained similar in OcaB+/+ mice harboring either OcaB+/+ or OcaB−/− bone marrow (data not shown).

The increased insulin resistance in KO/WT mice compared with that in KO/KO animals was associated with higher levels of infiltrated B2 cells in eWAT and iWAT after irradiation, whereas no significant change was observed in levels of infiltrated B1-a or B1-b cells (Fig. 7K and L and Supplementary Fig. 6B). These responses were associated with a normalization of circulating IgG and IgE concentrations, whereas levels of IgM increased only slightly (Fig. 7M–O). Changes in IgG levels in OcaB−/− mice with OcaB+/+ bone marrow paralleled the changes in insulin sensitivity (Fig. 7P). Thus normalization of OcaB activity in B cells through bone marrow transfer restored age-induced weight gain and insulin resistance.

Whereas the roles of B lymphocytes in the modulation of energy metabolism upon DIO have been partially characterized, their involvement in age-induced metabolic disorders is not well understood. This is of importance because aging and obesity—despite sharing many similar outcomes—display distinct genomic fingerprints and etiology (13,14). In this study we showed that aging induces the accumulation of B2 lymphocytes in visceral WAT depots. Blocking the activity of B lymphocytes through genetic KO of OcaB prevented the development of age-associated fat accumulation and glucose intolerance, whereas restoring their activity through bone marrow transfer triggered fat accretion and insulin resistance in aged mice. Thus our observations lead us to propose that B2 lymphocytes affect energy balance and glucose homeostasis upon aging, in part through changes in OcaB activity.

Our first finding was the accumulation of B2 cells upon aging in visceral but not in subcutaneous WAT. In 4-month-old mice, eWAT comprised 8% B cells, similar to the value in 2-month-old mice described in a previous report (34), and then doubled at the age of 1 y. We found no age-associated changes in the frequencies of total T cells or macrophages, although we did not discriminate between their different subtypes. This was consistent with unaltered total numbers of adipocyte-surrounding macrophages in aging mice (35) despite a functional shift toward a proinflammatory M1 status (36). The increase in B cells in visceral WAT was mainly due to accumulation of the B2 subtype, mirroring a general increase in circulating IgG and the number of mature lymphocytes. That B-cell numbers in subcutaneous WAT did not change upon aging possibly reflects that either a plateau was attained or the effects of mechanisms to retain infiltrated B cells were lessened. In line with increased B cells in eWAT of aged mice, we found that OcaB levels also increased in WAT upon aging. It is still unclear whether this increase resulted from augmented expression on a per B cell basis or from an increase in infiltrated B cells (37).

B2 cells from DIO mice produce a pathogenic phenotype that partly depends on the stimulation of leukotriene LTB4 signaling (7), activation of CD4+ cells, and production of IgG (1). Previous studies have established that OcaB modulates at least the two latter processes (21,38). Therefore, we postulated that blunting B-cell activation through deletion of OcaB would prevent metabolic alterations upon aging. Consistent with previous reports, OcaB−/− mice on a pure C57BL/6J genetic background were capable of B-cell differentiation and Ig secretion (18,38), albeit at lower levels than OcaB+/+ mice. The production of Igs, and strikingly that of IgG2c, was impaired by lack of OcaB. In these settings, we found that despite having apparent normal glucose homeostasis at the age of 2 months, OcaB−/− mice at 6 months of age were considerably more tolerant to glucose and sensitive to insulin than their OcaB+/+ littermates. This was due to large differences in their ability to take up glucose in tissues such as iWAT and skeletal muscle, and not because of reduced hepatic glucose production. Because we observed no modification in B-cell numbers in iWAT upon normal aging, the observation that changes in B2 cells in eWAT caused insulin sensitization in other depots and tissues suggests communication through endocrine cross talk, likely including IgG. In this view, lack of IgG2c in OcaB−/− mice might contribute to the mechanisms by which they are resistant to the detrimental effects of aging on insulin sensitivity. During obesity, B cells are activated through lipid-induced Toll-like receptor signaling to produce IgG2c (auto)antibodies that promote WAT inflammation and insulin resistance (9,39). This would also be in agreement with the observation that treatment of B-cell-deficient muMt mice with IgG isolated from DIO mice results in reduced insulin sensitivity (9). However, our attempts to directly modulate subcutaneous adipocytes with B cells in coculture experiments or with IgG2c failed to provide a direct mechanistic link (data not shown). Rather, the recent discovery that endothelial cells are necessary to relay IgG messages to adipocytes (40), together with our own results, indicates an intricate and indirect mechanism whereby B cells influence adipocyte biology and whole-body insulin sensitivity. Finally, our findings support and extend previous reports that reduction of B-cell activity improves whole-body insulin sensitivity in DIO mice (7,9). In addition, these results indicate a genuine contribution of OcaB in the age-related impairment of glucose homeostasis.

From birth, OcaB−/− mice had a lower body weight than their OcaB+/+ littermates, a phenotype also noted in a previous report (18). These mice showed a low daily food intake that likely contributed to their lower body weight and smaller WAT mass. However, OcaB−/− animals were characterized by a very low food efficiency that persisted upon aging, suggesting higher energy expenditure that was confirmed by studying body weight gain in comparison with a group of pair-fed OcaB+/+ mice. Because fecal energy content was similar between OcaB+/+ and OcaB−/− mice, and considering the important role of B cells in the gut, it is possible that OcaB−/− animals experienced intestinal malabsorption that might have contributed to their lower food efficiency. Thus, the additive effects of OcaB on both ends of the energy balance equation resulted in the robust inhibition of age-induced fat accretion. The small amount of body fat probably also favored insulin sensitivity in OcaB−/− animals. Because B2 and B1-a cells modulate insulin sensitivity without changes in body weight in murine DIO models (9,11), our results further suggest that OcaB might influence body weight regulation beyond its typical impact on B cells. In this regard, it is important to take into account the potential contributions of OcaB in activated T cells (21) and in nonhematopoietic cells such as those in the airway epithelium (41), which can only be resolved using tissue-specific and inducible models.

Physical activity was not the main source of increased energy expenditure in OcaB-deficient mice. Instead, their elevated oxygen consumption was associated with higher body temperature and stimulated metabolic activity of BAT and WAT depots. Browning of subcutaneous iWAT in OcaB−/− mice was notably related to modifications in gene expression, including that of Cidea, and extensive downregulation of Nnmt. It is interesting to note that, mice with Nnmt knockdown are protected against DIO in part because of increased energy expenditure in adipocytes (42). Because Nnmt and OcaB are expressed in adipocytes and B cells, respectively, this finding supports the concepts that infiltrated B cells could exert paracrine cross talk toward adipocytes and that depletion of OcaB strongly alters this communication.

The absence of OcaB also affected levels of insulin-sensitizing cytokines. In this regard, we found that IL-10 levels were locally increased in eWAT of OcaB−/− mice. Although whole-body IL-10 ablation enhances glucose tolerance and browning of iWAT in the context of DIO and aging (43), B-cell-specific deletion of IL-10 causes WAT inflammation and insulin resistance in mice with DIO (10). Lack of OcaB could modify a specific age-associated B-cell repertoire in concert with other immune cells, such as pools of fat-resident T regulatory cells (14), resident macrophages (4), or IL-10-secreting, innate-like type 2 lymphoid cells (44). Furthermore, in adipose tissue, feedback interactions with the sympathetic nervous system are also plausible because β-adrenergic receptors are highly expressed in B lymphocytes (45). This hypothesis is especially intriguing considering a recent report that macrophages from old mice exacerbate catecholamine degradation within adipose tissue (46), thereby in theory reducing the sympathetic drive to surrounding cells. Finally, OcaB−/− mice displayed favorable systemic reduction of proinflammatory TNF-α, KC, and MIP-1α cytokines, which also likely contributed to insulin sensitization. These possibilities must yet be experimentally evaluated.

It is also interesting that OcaB−/− mice showed robustly elevated IL-6 levels. This effect was likely mediated through indirect mechanisms, because OcaB positively controls IL-6 transcription in B cells (47). However, because IL-6 is important for germinal centers (47), the elevated IL-6 levels in OcaB−/− animals may represent a compensatory response to the impaired formation of germinal centers present in these mice. As a secondary impact, high levels of muscle-derived IL-6 in mice lacking OcaB could have contributed to their greater insulin sensitivity (27), their lower food intake (48), and the enhanced browning of iWAT, as observed in cachexia (49) or burn injury (50).

It is important to note that transfer of OcaB+/+ bone marrow to OcaB−/− mice enhanced weight gain and induced insulin resistance. However, weight gain and insulin sensitivity after the transfer were unexpectedly similar in OcaB+/+ mice harboring either OcaB+/+ or OcaB−/− bone marrow. This lack of effect was possibly due to remaining hematopoietic cells found in radio-insensitive compartments (23), as total splenic B-cell counts remained comparable between the two groups, or to the expression of OcaB in nonhematopoietic cells (41). However, replenishment with OcaB+/+ bone marrow in OcaB−/− mice notably reduced energy expenditure to levels observed in OcaB+/+ animals. This strongly suggests that the changes in energy balance and metabolism found in OcaB−/− mice are primarily brought about by modifications in B lymphocytes. Again, the intrinsic nature of bone marrow transfer prevents definite conclusions excluding possible roles for OcaB in activated T cells (21). In addition, because OcaB−/− mice with OcaB+/+ marrow displayed circulating IgG and IgE levels comparable to those of OcaB+/+ mice, it is plausible that these Igs at least partially mediate the impact of OcaB+/+ B lymphocytes in OcaB−/− animals. Because introduction of OcaB−/− bone marrow in OcaB+/+ mice robustly reduced IgE levels without ultimately modulating energy metabolism, it is probable that the role of IgG overwhelms that of IgE in insulin sensitivity. This suggests that alterations in IgE levels in OcaB−/− mice were related more to intrinsic B2 deficiency due to the lack of OcaB than to age-associated changes in insulin resistance per se (18,38).

Here we report that aging triggers modifications in the adaptive immune system, which contribute to age-associated metabolic alterations. Mice with impaired B lymphocytes due to OcaB invalidation were resistant to age-induced defects in glucose tolerance and energy expenditure, both of which were abrogated by transfer of OcaB+/+ hematopoietic cells. How B lymphocytes sense and respond to metabolic stimuli during the life span remains to be thoroughly investigated.

Acknowledgments. The authors thank Serge Rivest (Université Laval) for help with the irradiation protocol, Christine Dion (Centre de recherche de l’Institut universitaire de cardiologie et de pneumologie de Québec) for surgical preparation, and Bruno Marcotte (Centre de recherche de l’Institut universitaire de cardiologie et de pneumologie de Québec) for technical assistance with cytokine measurements. The authors also thank Nataly Laflamme (Centre de recherche du CHU de Québec) and Marie-Michèle Plante (Centre de recherche du CHU de Québec) for technical assistance with bone marrow extraction, and Pierre Samson (Centre de recherche de l’Institut universitaire de cardiologie et de pneumologie de Québec) for his technical expertise with indirect calorimetry and DEXA scans. The authors also acknowledge the support of the Quebec Network of Research on Aging, which allowed access to its colony of aging mice.

Funding. This work was funded by grants from the Canadian Institutes of Health Research (CIHR) (FRN-14813 to A.M. and MOP-110992 to F.P.). S.C. was the recipient of a PhD studentship award from the Fonds de Recherche du Québec-Santé (FRQS). A.C. and J.L. are Canadian Diabetes Association postdoctoral fellows. M.F. holds the Canada Research Chair in Bone and Energy Metabolism. Y.B. holds a Canada Research Chair in Genomics of Heart and Lung Diseases. D.M. is a FRQS Junior 1 Scholar. F.P. has received an FRQS Senior Scholarship.

Duality of Interest. No potential conflicts of interest relevant to this article were reported.

Author Contributions. S.C. and F.P. conceived the study and wrote the manuscript. S.C., A.C., P.S.-P., F.F.A., P.B.-L., M.-C.D., and J.L. performed mouse experiments under the supervision of Y.D., J.C., M.L., M.F., A.M., D.R., D.M., and F.P. S.M., S.S.-L., and P.B.-L. performed FACS analyses under the supervision of J.S.L., D.M., and F.P. S.C., S.M., and S.S.-L. conducted ELISA. S.C., S.M., and A.C. carried out quantitative PCR. S.C. performed the chimera protocol. E.L.-C. and Y.B. conceived and analyzed the microarray experiment. All authors analyzed and discussed the data and revised the manuscript. F.P. is the guarantor of this work and, as such, had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.

Prior Presentation. Parts of this study were presented at the Keystone Symposia on Obesity, Keystone, CO, 12–17 January 2011; the 1st International Conference on ImmunoMetabolism, Crete, Greece, 18–23 September 2011; and the Benzon Symposium on Adipose Tissue in Health and Disease, Copenhagen, Denmark, 27–30 August 2012.

1.
Winer
DA
,
Winer
S
,
Chng
MH
,
Shen
L
,
Engleman
EG
.
B Lymphocytes in obesity-related adipose tissue inflammation and insulin resistance
.
Cell Mol Life Sci
2014
;
71
:
1033
1043
[PubMed]
2.
Feuerer
M
,
Herrero
L
,
Cipolletta
D
, et al
.
Lean, but not obese, fat is enriched for a unique population of regulatory T cells that affect metabolic parameters
.
Nat Med
2009
;
15
:
930
939
[PubMed]
3.
Wu
D
,
Molofsky
AB
,
Liang
HE
, et al
.
Eosinophils sustain adipose alternatively activated macrophages associated with glucose homeostasis
.
Science
2011
;
332
:
243
247
[PubMed]
4.
Qiu
Y
,
Nguyen
KD
,
Odegaard
JI
, et al
.
Eosinophils and type 2 cytokine signaling in macrophages orchestrate development of functional beige fat
.
Cell
2014
;
157
:
1292
1308
[PubMed]
5.
McDonnell
ME
,
Ganley-Leal
LM
,
Mehta
A
, et al
.
B lymphocytes in human subcutaneous adipose crown-like structures
.
Obesity (Silver Spring)
2012
;
20
:
1372
1378
[PubMed]
6.
Duffaut
C
,
Galitzky
J
,
Lafontan
M
,
Bouloumié
A
.
Unexpected trafficking of immune cells within the adipose tissue during the onset of obesity
.
Biochem Biophys Res Commun
2009
;
384
:
482
485
[PubMed]
7.
Ying
W
,
Wollam
J
,
Ofrecio
JM
, et al
.
Adipose tissue B2 cells promote insulin resistance through leukotriene LTB4/LTB4R1 signaling
.
J Clin Invest
2017
;
127
:
1019
1030
[PubMed]
8.
DeFuria
J
,
Belkina
AC
,
Jagannathan-Bogdan
M
, et al
.
B cells promote inflammation in obesity and type 2 diabetes through regulation of T-cell function and an inflammatory cytokine profile
.
Proc Natl Acad Sci U S A
2013
;
110
:
5133
5138
[PubMed]
9.
Winer
DA
,
Winer
S
,
Shen
L
, et al
.
B cells promote insulin resistance through modulation of T cells and production of pathogenic IgG antibodies
.
Nat Med
2011
;
17
:
610
617
[PubMed]
10.
Nishimura
S
,
Manabe
I
,
Takaki
S
, et al
.
Adipose natural regulatory B cells negatively control adipose tissue inflammation
.
Cell Metab
2013
;
18
:
759
766
[PubMed]
11.
Shen
L
,
Chng
MH
,
Alonso
MN
,
Yuan
R
,
Winer
DA
,
Engleman
EG
.
B-1a lymphocytes attenuate insulin resistance
.
Diabetes
2015
;
64
:
593
603
[PubMed]
12.
Harmon
DB
,
Srikakulapu
P
,
Kaplan
JL
, et al
.
Protective role for B-1b B cells and IgM in obesity-associated inflammation, glucose intolerance, and insulin resistance
.
Arterioscler Thromb Vasc Biol
2016
;
36
:
682
691
[PubMed]
13.
Carter
S
,
Caron
A
,
Richard
D
,
Picard
F
.
Role of leptin resistance in the development of obesity in older patients
.
Clin Interv Aging
2013
;
8
:
829
844
[PubMed]
14.
Bapat
SP
,
Myoung Suh
J
,
Fang
S
, et al
.
Depletion of fat-resident Treg cells prevents age-associated insulin resistance
.
Nature
2015
;
528
:
137
141
[PubMed]
15.
Mehr
R
,
Melamed
D
.
Reversing B cell aging
.
Aging (Albany NY)
2011
;
3
:
438
443
[PubMed]
16.
Jankovic
M
,
Nussenzweig
MC
.
OcaB regulates transitional B cell selection
.
Int Immunol
2003
;
15
:
1099
1104
[PubMed]
17.
Qin
XF
,
Reichlin
A
,
Luo
Y
,
Roeder
RG
,
Nussenzweig
MC
.
OCA-B integrates B cell antigen receptor-, CD40L- and IL 4-mediated signals for the germinal center pathway of B cell development
.
EMBO J
1998
;
17
:
5066
5075
[PubMed]
18.
Kim
U
,
Qin
XF
,
Gong
S
, et al
.
The B-cell-specific transcription coactivator OCA-B/OBF-1/Bob-1 is essential for normal production of immunoglobulin isotypes
.
Nature
1996
;
383
:
542
547
[PubMed]
19.
Strubin
M
,
Newell
JW
,
Matthias
P
.
OBF-1, a novel B cell-specific coactivator that stimulates immunoglobulin promoter activity through association with octamer-binding proteins
.
Cell
1995
;
80
:
497
506
[PubMed]
20.
Luo
Y
,
Fujii
H
,
Gerster
T
,
Roeder
RG
.
A novel B cell-derived coactivator potentiates the activation of immunoglobulin promoters by octamer-binding transcription factors
.
Cell
1992
;
71
:
231
241
[PubMed]
21.
Shakya
A
,
Goren
A
,
Shalek
A
, et al
.
Oct1 and OCA-B are selectively required for CD4 memory T cell function
.
J Exp Med
2015
;
212
:
2115
2131
[PubMed]
22.
Hess
J
,
Nielsen
PJ
,
Fischer
KD
,
Bujard
H
,
Wirth
T
.
The B lymphocyte-specific coactivator BOB.1/OBF.1 is required at multiple stages of B-cell development
.
Mol Cell Biol
2001
;
21
:
1531
1539
[PubMed]
23.
Lampron
A
,
Lessard
M
,
Rivest
S
.
Effects of myeloablation, peripheral chimerism, and whole-body irradiation on the entry of bone marrow-derived cells into the brain
.
Cell Transplant
2012
;
21
:
1149
1159
[PubMed]
24.
Liesveld
JL
,
Rothberg
PG
.
Mixed chimerism in SCT: conflict or peaceful coexistence
?
Bone Marrow Transplant
2008
;
42
:
297
310
[PubMed]
25.
Li
Z
,
Picard
F
.
Modulation of IGFBP2 mRNA expression in white adipose tissue upon aging and obesity
.
Horm Metab Res
2010
;
42
:
787
791
[PubMed]
26.
Blais Lecours
P
,
Duchaine
C
,
Taillefer
M
, et al
.
Immunogenic properties of archaeal species found in bioaerosols
.
PLoS One
2011
;
6
:
e23326
[PubMed]
27.
White
PJ
,
St-Pierre
P
,
Charbonneau
A
, et al
.
Protectin DX alleviates insulin resistance by activating a myokine-liver glucoregulatory axis
.
Nat Med
2014
;
20
:
664
669
[PubMed]
28.
Mari
A
.
Estimation of the rate of appearance in the non-steady state with a two-compartment model
.
Am J Physiol
1992
;
263
:
E400
E415
[PubMed]
29.
Labbé
SM
,
Caron
A
,
Bakan
I
, et al
.
In vivo measurement of energy substrate contribution to cold-induced brown adipose tissue thermogenesis
.
FASEB J
2015
;
29
:
2046
2058
[PubMed]
30.
Ménard
,
SL
,
Ci
X
,
Frisch
F
, et al
.
Mechanism of reduced myocardial glucose utilization during acute hypertriglyceridemia in rats
.
Mol Imaging Biol
2009
;
11
:
6
14
.
31.
Bolstad
BM
,
Irizarry
RA
,
Astrand
M
,
Speed
TP
.
A comparison of normalization methods for high density oligonucleotide array data based on variance and bias
.
Bioinformatics
2003
;
19
:
185
193
[PubMed]
32.
Du
P
,
Kibbe
WA
,
Lin
SM
.
lumi: a pipeline for processing Illumina microarray
.
Bioinformatics
2008
;
24
:
1547
1548
[PubMed]
33.
Gentleman
RC
,
Carey
VJ
,
Bates
DM
, et al
.
Bioconductor: open software development for computational biology and bioinformatics
.
Genome Biol
2004
;
5
:
R80
[PubMed]
34.
Caspar-Bauguil
S
,
Cousin
B
,
Galinier
A
, et al
.
Adipose tissues as an ancestral immune organ: site-specific change in obesity
.
FEBS Lett
2005
;
579
:
3487
3492
[PubMed]
35.
Wu
D
,
Ren
Z
,
Pae
M
, et al
.
Aging up-regulates expression of inflammatory mediators in mouse adipose tissue
.
J Immunol
2007
;
179
:
4829
4839
[PubMed]
36.
Lumeng
CN
,
Liu
J
,
Geletka
L
, et al
.
Aging is associated with an increase in T cells and inflammatory macrophages in visceral adipose tissue
.
J Immunol
2011
;
187
:
6208
6216
[PubMed]
37.
Martinez-Jimenez
CP
,
Eling
N
,
Chen
HC
, et al
.
Aging increases cell-to-cell transcriptional variability upon immune stimulation
.
Science
2017
;
355
:
1433
1436
[PubMed]
38.
Schubart
DB
,
Rolink
A
,
Kosco-Vilbois
MH
,
Botteri
F
,
Matthias
P
.
B-cell-specific coactivator OBF-1/OCA-B/Bob1 required for immune response and germinal centre formation
.
Nature
1996
;
383
:
538
542
[PubMed]
39.
Seijkens
T
,
Kusters
P
,
Chatzigeorgiou
A
,
Chavakis
T
,
Lutgens
E
.
Immune cell crosstalk in obesity: a key role for costimulation?
Diabetes
2014
;
63
:
3982
3991
[PubMed]
40.
Tanigaki
K
,
Sacharidou
A
,
Peng
J
, et al
.
Hyposialylated IgG activates endothelial IgG receptor FcγRIIB to promote obesity-induced insulin resistance
.
J Clin Invest
2018
;
128
:
309
322
[PubMed]
41.
Zhou
H
,
Brekman
A
,
Zuo
WL
, et al
.
POU2AF1 functions in the human airway epithelium to regulate expression of host defense genes
.
J Immunol
2016
;
196
:
3159
3167
[PubMed]
42.
Kraus
D
,
Yang
Q
,
Kong
D
, et al
.
Nicotinamide N-methyltransferase knockdown protects against diet-induced obesity
.
Nature
2014
;
508
:
258
262
[PubMed]
43.
Rajbhandari
P
,
Thomas
BJ
,
Feng
AC
, et al
.
IL-10 signaling remodels adipose chromatin architecture to limit thermogenesis and energy expenditure
.
Cell
2018
;
172
:
218
233.e17
[PubMed]
44.
Lee
MW
,
Odegaard
JI
,
Mukundan
L
, et al
.
Activated type 2 innate lymphoid cells regulate beige fat biogenesis
.
Cell
2015
;
160
:
74
87
[PubMed]
45.
Sanders
VM
.
The role of norepinephrine and beta-2-adrenergic receptor stimulation in the modulation of Th1, Th2, and B lymphocyte function
.
Adv Exp Med Biol
1998
;
437
:
269
278
[PubMed]
46.
Camell
CD
,
Sander
J
,
Spadaro
O
, et al
.
Inflammasome-driven catecholamine catabolism in macrophages blunts lipolysis during ageing
.
Nature
2017
;
550
:
119
123
[PubMed]
47.
Karnowski
A
,
Chevrier
S
,
Belz
GT
, et al
.
B and T cells collaborate in antiviral responses via IL-6, IL-21, and transcriptional activator and coactivator, Oct2 and OBF-1
.
J Exp Med
2012
;
209
:
2049
2064
[PubMed]
48.
Timper
K
,
Denson
JL
,
Steculorum
SM
, et al
.
IL-6 improves energy and glucose homeostasis in obesity via enhanced central IL-6 trans-signaling
.
Cell Rep
2017
;
19
:
267
280
[PubMed]
49.
Petruzzelli
M
,
Schweiger
M
,
Schreiber
R
, et al
.
A switch from white to brown fat increases energy expenditure in cancer-associated cachexia
.
Cell Metab
2014
;
20
:
433
447
[PubMed]
50.
Patsouris
D
,
Qi
P
,
Abdullahi
A
, et al
.
Burn induces browning of the subcutaneous white adipose tissue in mice and humans
.
Cell Reports
2015
;
13
:
1538
1544
[PubMed]
Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered. More information is available at http://www.diabetesjournals.org/content/license.

Supplementary data