This is difficult but informative research article on the role of obesity and macrophage activity (white blood cells) in inflammation. Depending upon how macrophages are activated, they can play a highly inflammatory role which can be destructive if sustained, or they can play a surprising role in metabolic homeostasis. Obesity appears to be a primary instigater of the classical activation state.
Profiling studies of visceral fat from lean and obese animals showed that rather than a passive energy repository, adipose tissue teems with hematopoietic cells whose activation states and functionalities vary with the nutritional status of the organism (Hotamisligil 2006). However, despite the presence of T-cells and B-cells, NK cells, DCs, NKT cells, eosinophils, mast cells, and basophils in obese adipose tissue, macrophages are numerically and functionally dominant. Macrophages comprise ∼10%–15% of all cells within lean visceral adipose tissue and expand tremendously with obesity, where they account for a staggering 45%–60% of all cells (Weisberg et al. 2003; Xu et al. 2003). Importantly, this numerical expansion is accompanied by a marked phenotypic transition as well: adipose tissue–associated macrophages isolated from lean animals express a distinct bias toward alternative activation, which is swapped for a pro-inflammatory classical bent with the acquisition of obesity (Lumeng et al. 2007a; Odegaard et al. 2007). Similarly, liver-associated macrophages—Kupffer cells—isolated from lean animals are markedly alternatively biased, whereas those from obese animals are polarized in the opposite direction of classical activation (Odegaard et al. 2008). To appreciate the significance of this transition, we must first briefly discuss the physiological implications of the classical and alternative activation phenotypes.
Macrophages, in their canonical role as sentinels of the innate immune system, are responsible for sensing, integrating, and responding appropriately to myriad stimuli in their tissue milieux. Despite the protean nature of these stimuli, macrophage activity is channeled through two distinct response patterns designated as classical (M1) and alternative (M2) activation (Gordon 2003). These programs constitute the stereotyped response to bacterial and parasitic infection, respectively:
Classical activation results in short-lived, highly inflammatory macrophages with potent bactericidal potential, whereas alternative activation is associated with enduring anti-parasitic and regulatory/reparative responses (Martinez et al. 2009; Odegaard and Chawla 2011).
Classically activated macrophages secrete large amounts of pro-inflammatory cytokines (e.g., IL-1β, IL-6, IL-8, IL-12, TNFα), express high levels of costimulatory molecules important in T-cell activation (e.g., MHC, CD40, CD86), and produce bactericidal mediators, such as nitric oxide, via Nos2 (Gordon 2003). Conversely, alternatively activated cells have a distinct secretory phenotype (e.g., IL-10, TGFβ, Chi3l3, Retnla), express numerous pattern recognition receptors (e.g., mannose receptor, dectin, CD301), and metabolize arginine to produce biosynthetic precursors (e.g., polyamines, proline) via arginase 1 (Martinez et al. 2009; Odegaard and Chawla 2011).
Although these activation state definitions rest on the macrophage’s role in host defense, further study has defined their critical function in non-immunological contexts as well. For example, the classical phenotype is implicated in numerous inflammatory and metabolic diseases, whereas alternative activation is associated with wound healing, tissue remodeling, metabolic homeostasis, and atopic disease (Gordon 2003; Odegaard and Chawla 2011). Although the classical–alternative dichotomy has been studied in numerous contexts, few instances are as instructive as the tissue–macrophage relationship in metabolic disease.
Adipose Tissue Macrophages, Inflammation, and Insulin Resistance
Aside from the observational data presented above, at least four distinct lines of evidence implicate the macrophage as the nexus of inflammatory insulin resistance. First, ablation of CD11c+ inflammatory macrophages using CD11c-DTR transgenic mice improves insulin sensitivity without altering adipose tissue mass in obese animals (Patsouris et al. 2008). Second, interference with inflammatory macrophage recruitment through genetic or pharmacologic disruption of C–C motif chemokine receptor-2 (CCR2) results in protection against obesity-related insulin resistance and hepatic steatosis (Weisberg et al. 2006; Ito et al. 2008). Third, transgenic mice expressing Ccl2, a ligand for CCR2, in adipocytes have macrophage infiltration of their adipose tissue that is associated with increased insulin resistance (Kamei et al. 2006; Kanda et al. 2006). Lastly, hematopoietic-specific loss of JNK1 and myeloid-specific loss of IKKβ, both interventions that render macrophages inflammatorily effete, preclude the development of obesity-related insulin resistance (Arkan et al. 2005; Solinas et al. 2007).
Obesity results in recruitment of macrophages into adipose tissue, which promotes adipose tissue inflammation and insulin resistance. Obesity results in increased levels of circulating saturated fatty acids, which activate Toll-like receptors 2/4 (TLR2/4) …
Triggers for Classical Activation
Several lines of evidence link well-described events early in obesity with classical macrophage activation. Obesity is characterized by the expansion of adipose tissue: Adipose tissue depots expand approximately fivefold to 10-fold over their lean mass. This massive hypertrophy induces necrosis of adipocytes due to excessive endoplasmic reticulum (ER) stress and hypoxia, as the expanding tissue outgrows its vascular supply (Hotamisligil 2006; Rutkowski et al. 2009), providing a potent inflammatory stimuli for macrophages (Savill and Fadok 2000; Savill et al. 2002). Indeed, temporal analyses of obesity reveal a correlation between the appearance of necrotic adipocytes surrounded by inflammatory CD11c+ macrophages and the onset of clinical insulin resistance (Cinti et al. 2005; Strissel et al. 2007).
Another hallmark of developing obesity is dysregulation of fatty acid homeostasis, which is usually associated with high-risk dietary patterns (e.g., refined sugars and saturated fatty acid consumption).
Interestingly, the same saturated fatty acids that come to dominate the obese lipid milieu are capable of activating Toll-like receptor 4 (TLR4), a sensor tuned to structurally similar bacterial lipids such as lipopolysaccharide, to produce an inflammatory response (Konner and Bruning 2011). Indeed, (Shi et al. 2006), whereas genetic deletion of TLR4 in experimental animals protects against diet-induced insulin resistance (Saberi et al. 2009). Accompanying this shift in fatty acid composition, escalating ceramide biosynthesis also contributes to inflammatory activation (Vandanmagsar et al. 2011).
Many inflammatory signaling cascades, including those provoked by ER stress, hypoxia, necrotic cellular debris, and ceramide dysregulation, converge on the inflammasome, a multiprotein complex critical for the production and secretion of IL-1β and IL-18 (Petrilli et al. 2007; Chen and Nunez 2010). Indeed, adipose tissue inflammasome activation, both in adipocytes and macrophages, parallels the development of obesity, resulting in IL-1β-mediated inflammatory insulin resistance, whereas its inhibition is effective in control of obesity-related metabolic pathology (Stienstra et al. 2010; Zhou et al. 2010; Vandanmagsar et al. 2011). Interestingly, as obesity progresses and peripheral insulin resistance builds, inflammasome activation is also demonstrable within islet-infiltrating macrophages and the β-cell itself and is correlated with IL-1β production (Boni-Schnetzler et al. 2008; Maedler et al. 2008; Masters et al. 2010). In keeping with this, treatment of type 2 diabetic patients with anakinra, a competitive IL-1R antagonist, decreases markers of systemic inflammation and improves glycemic control and secretory function of the β-cells (Larsen et al. 2007).
In addition to the acquisition of an inflammatory phenotype, obesity is also accompanied by a marked increase in macrophage representation, which, in the context of increasing adipose tissue mass, represents de novo recruitment (Weisberg et al. 2003). Among the recruitment factors, CCL2-mediated ingress of Ly6ChiCCR2+ monocytes, which selectively differentiate into classically activated macrophages, plays a dominant role in trafficking of adipose tissue macrophages (Kamei et al. 2006; Kanda et al. 2006; Weisberg et al. 2006). Congruent with this view, disruption of either CCL2 or CCR2, as mentioned previously, protects against diet-related insulin resistance.
ALTERNATIVELY ACTIVATED MACROPHAGES AND INSULIN SENSITIVITY
Although much of the last two decades have been spent maligning the role of innate immunity in metabolic homeostasis, evidence of a positive influence is also emerging. The foremost example of innate immunity’s beneficent potential is found in the tissue macrophage. In our rush to demonize this cell for its central role in insulin resistance, we neglect the key observation that the adipose tissue and liver of lean, insulin-sensitive individuals are rife with macrophages—∼10%–15% of all healthy liver and adipose tissue cells are macrophages (Gordon et al. 1992; Weisberg et al. 2003; Lumeng et al. 2007a). Adipose tissue macrophages from lean adipose tissue, rather than being simply not inflammatory, are significantly activated along the alternative pathway (Lumeng et al. 2007a; Odegaard et al. 2007). Similarly, Kupffer cells from lean animals express markers of alternative activation, which they swap for an inflammatory profile in obesity (Odegaard et al. 2008).
This hitherto unexpected activation-state diversity raises an important question: What are large numbers of alternatively activated macrophages doing in non-inflamed liver and fat?
Transcriptional Regulation of Alternative Macrophage Activation
The first experiments addressing this question were derived from early studies of transcriptional determinants of macrophage alternative activation. These studies showed that although STAT6 was the dominant transcriptional initiator of alternative activation, peroxisome proliferator-activated receptors PPARγ and PPARδ—nuclear receptors hitherto recognized only as the body’s fatty acid sensors—were identified as critical for sustaining the response (Odegaard et al. 2007, 2008; Kang et al. 2008). In the absence of PPARγ, murine macrophages are incapable of affecting the metabolic shift required for sustained alternative response (Vats et al. 2006; Odegaard et al. 2007), whereas PPARδ deficiency disrupts the macrophage’s ability to coordinate and sustain the immunologic phenotype of alternative activation (Kang et al. 2008; Odegaard et al. 2008). Either’s absence is then sufficient to abrogate any durable alternative response in vitro. In vivo, however, a surprising phenotypical nuance was uncovered: macrophage-specific PPARγ deficiency preferentially abolishes alternative activation in visceral adipose tissue macrophages, but loss of PPARδ in the hematopoietic system appears to more potently affect Kupffer cell alternative activation capacity (Kang et al. 2008; Odegaard et al. 2008). Importantly, both strains of mice show similar and marked decreases in insulin sensitivity and increased weight gain when challenged with a high-fat diet (Hevener et al. 2007; Odegaard et al. 2007, 2008; Kang et al. 2008), showing that the absence of alternatively activated macrophages impairs insulin signaling even in the absence of established inflammation.
Eosinophils: IL-4-Producing Cells in Adipose Tissue
The unexpected, Janus-like ability of macrophages to promote insulin sensitivity as well as resistance established a paradigm under which other immunological determinants of insulin sensitivity were actively sought. To this end, differential profiling of other hematopoietic subsets in adipose tissue from lean and obese animals uncovered a striking correlation between eosinophil representation, alternative macrophage activation in adipose tissue, and the obesity–insulin resistance disease axis.
Indeed, eosinophil number shows a tight correlation to macrophage activation status and is inversely related to body mass, insulin resistance, and inflammatory markers (Wu et al. 2011). Furthermore, eosinophil-deficient mice show an alarming propensity to insulin resistance, whereas those with a surfeit of eosinophils (e.g., the IL-5 transgenic mouse) are resistant to diet-induced obesity and insulin resistance (Wu et al. 2011).
Health-e-Solutions comment: Obesity is a significant contributor to the development of chronic disease. This study demonstrates the impact of obesity on adipose tissue. Losing weight may improve the number of alternatively-activated macrophages and thus improve insulin sensitivity.
In our boys, the Health-e-Solutions lifestyle usually results in a lower total white blood cell count. Our doctors explained it is due to the “clean” nature of the diet that put less debris into the blood stream, thus requiring less white blood cell activity. Macrophages (Monos) usually make up 2% to 8% of WBCs. They fight infections by “eating” germs and telling the immune system what germs they have found. Monocytes circulate in the blood. When monocytes settle in various tissues they are called macrophages. A high count usually indicates a bacterial infection. A low count would indicate the opposite.
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