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Background: The purpose of this study was to investigate physiological phenotypes of asthma in obesity., Research Question: Do physiological responses during bronchoconstriction distinguish different groups of asthma in people with obesity, and also differentiate from responses simply related to obesity?, Study Design and Methods: Cross-sectional study of people with obesity (31 with asthma and 22 without lung disease). Participants underwent methacholine challenge testing with measurement of spirometry and respiratory system impedance by oscillometry., Results: Participants had class III obesity (BMI, 46.7 ± 6.6 kg/m 2 in control subjects and 47.2 ± 8.2 kg/m 2 in people with asthma). Most participants had significant changes in peripheral airway impedance in response to methacholine: in control subjects, resistance at 5 Hz measured by oscillometry increased by 45% ± 27% and area under the reactance curve (AX) by 268% ± 236% in response to 16 mg/mL methacholine; in people with asthma, resistance at 5 Hz measured by oscillometry increased by 52% ± 38% and AX by 361% ± 295% in response to provocation concentration producing a 20% fall in FEV 1 dose of methacholine. These responses suggest that obesity predisposes to peripheral airway reactivity. Two distinct groups of asthma emerged based on respiratory system impedance: one with lower reactance (baseline AX, 11.8; interquartile range, 9.9-23.4 cm H 2 O/L) and more concordant bronchoconstriction in central and peripheral airways; the other with high reactance (baseline AX, 46.7; interquartile range, 23.2-53.7 cm H 2 O/L) and discordant bronchoconstriction responses in central and peripheral airways. The high reactance asthma group included only women, and reported significantly more gastroesophageal reflux disease, worse chest tightness, more wheeze, and more asthma exacerbations than the low reactance group., Interpretation: Peripheral airway reactivity detected by oscillometry is common in obese control subjects and obese people with asthma. There is a subgroup of obese asthma characterized by significant peripheral airway dysfunction by oscillometry out of proportion to spirometric airway dysfunction. This peripheral dysfunction represents clinically significant respiratory disease not readily assessed by spirometry., (Copyright © 2023 American College of Chest Physicians. Published by Elsevier Inc. All rights reserved.)

Prior studies in younger adults showed that reducing the normally high intake of the saturated fatty acid, palmitic acid (PA), in the North American diet by replacing it with the monounsaturated fatty acid, oleic acid (OA), decreased blood concentrations and secretion by peripheral blood mononuclear cells (PBMCs) of interleukin (IL)-1β and IL-6 and changed brain activation in regions of the working memory network. We examined the effects of these fatty acid manipulations in the diet of older adults. Ten subjects, aged 65-75 years, participated in a randomized, cross-over trial comparing 1-week high PA versus low PA/high OA diets. We evaluated functional magnetic resonance imaging (fMRI) using an N-back test of working memory and a resting state scan, cytokine secretion by lipopolysaccharide (LPS)-stimulated PBMCs, and plasma cytokine concentrations. During the low PA compared to the high PA diet, we observed increased activation for the 2-back minus 0-back conditions in the right dorsolateral prefrontal cortex (Broadman Area (BA) 9; p < 0.005), but the effect of diet on working memory performance was not significant (p = 0.09). We observed increased connectivity between anterior regions of the salience network during the low PA/high OA diet (p < 0.001). The concentrations of IL-1β (p = 0.026), IL-8 (p = 0.013), and IL-6 (p = 0.009) in conditioned media from LPS-stimulated PBMCs were lower during the low PA/high OA diet. This study suggests that lowering the dietary intake of PA down-regulated pro-inflammatory cytokine secretion and altered working memory, task-based activation and resting state functional connectivity in older adults., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Author(s).)

Macrophages are important mediators of skeletal muscle function in both healthy and diseased states. In vivo specific depletion of macrophages provides an experimental method to understand physiological and pathophysiological effects of macrophages. Systemic depletion of macrophages can deplete skeletal muscle macrophages but also alters systemic inflammatory responses and metabolism, which confounds the muscle specific effects of macrophage depletion. The primary aim of this manuscript is to evaluate two methods of murine intramuscular macrophage depletion in an acute lung injury-associated indirect skeletal muscle wasting mouse model. Adult C57BL/6 (WT) and Macrophage Fas-Induced Apoptosis (MaFIA, C57BL/6-Tg) mice received clodronate liposomes or the dimerization drug AP20187 through intramuscular injection of the tibialis anterior muscle compartment, respectively. Vehicle control was injected in the contralateral muscle. We demonstrate intramuscular AP20187 in the MaFIA mouse depletes macrophages but causes an infiltration of CD45 intermediate neutrophils. In contrast, intramuscular clodronate liposomes successfully depletes macrophages without an associated increase in CD45 intermediate cells. In conclusion, intramuscular clodronate is effective for selective depletion of muscle macrophages without eliciting acute inflammation seen with AP20187 in MaFIA mice. This technique is an important tool to study the functional roles of macrophages in skeletal muscle., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest in this work., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The NADPH oxidase DUOX1 contributes to epithelial production of alarmins, including interleukin (IL)-33, in response to injurious triggers such as airborne protease allergens, and mediates development of mucus metaplasia and airway remodeling in chronic allergic airways diseases. DUOX1 is also expressed in non-epithelial lung cell types, including macrophages that play an important role in airway remodeling during chronic lung disease. We therefore conditionally deleted DUOX1 in either lung epithelial or monocyte/macrophage lineages to address its cell-specific actions in innate airway responses to acute airway challenge with house dust mite (HDM) allergen, and in chronic HDM-driven allergic airway inflammation. As expected, acute responses to airway challenge with HDM, as well as type 2 inflammation and related features of airway remodeling during chronic HDM-induced allergic inflammation, were largely driven by DUOX1 with the respiratory epithelium. However, in the context of chronic HDM-driven inflammation, DUOX1 deletion in macrophages also significantly impaired type 2 cytokine production and indices of mucus metaplasia. Further studies revealed a contribution of macrophage-intrinsic DUOX1 in macrophage recruitment upon chronic HDM challenge, as well as features of macrophage activation that impact on type 2 inflammation and remodeling., (© 2022. The Author(s), under exclusive licence to Society for Mucosal Immunology.)

Background: The purpose of this study was to investigate how 8-isoprostanes, used as a marker of airway oxidative stress, were related to sinus disease and asthma., Methods: We analyzed samples and data from two separate studies, one investigating sinonasal disease in asthma, the other investigating the effect of BMI on airway disease. We measured airway (nasal lavage) 8-isoprostanes and investigated the relationship with measures of sinus and asthma symptoms, asthma control and lung function., Results: The study of people with sinonasal disease and poorly controlled asthma included 48 obese, 31 overweight and 23 lean participants. In multivariate analysis, nasal lavage 8-isoprostane levels increased with increasing BMI (p < 0.01), and were higher in Caucasian than African American participants (p = 0.01). Sinus symptoms were inversely related to nasal 8-isoprostanes (p = 0.02) independent of BMI and Race. In the study investigating the effect of BMI on airway disease, we enrolled 13 controls with obesity and 21 people with obesity and asthma: 8-isoprostane levels were higher in obese controls than in obese people with asthma (p < 0.01), and levels were inversely related to sinus symptoms (p = 0.02) and asthma control (p < 0.01)., Interpretation: 8-isoprostanes in nasal lavage are increased in obesity, and increased in Caucasians compared with African Americans. However, levels are higher in obese controls than obese people with asthma, and appear inversely related to symptoms of airway disease., Clinical Implication: Airway 8-isoprostanes likely reflect complex oxidative signaling pathways, which are altered in obesity and those of different race, rather than being a simple marker of airway oxidative injury., Capsule Summary: Increased airway oxidative signaling (8-isoprostanes), may reflect normal physiology in the setting of obesity, as decreased levels are associated with disease activity in people with chronic sinonasal disease and asthma., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background: Pharmacokinetics (PK) of pharmaceuticals and pharmaconutrients are poorly understood in critically ill patients, and dosing is often based on healthy subject data. This might be particularly problematic with enteral medications due to metabolic abnormalities and impaired gastrointestinal tract absorption common in critically ill patients. Utilizing enteral fish oil, this study was undertaken to better understand and define PK of enteral omega-3 fatty acids (eicospentaenoic acid [EPA] and docosahexaenoic acid [DHA]) in critically ill patients with severe sepsis., Materials and Methods: Healthy volunteers (n = 15) and mechanically ventilated (MV) adults with severe sepsis (n = 10) were recruited and received 9.75 g EPA and 6.75 g DHA daily in two divided enteral doses of fish oil for 7 days. Volunteers continued their normal diet without other sources of fish oil, and sepsis patients received standard enteral feeding. Blood was collected at frequent intervals during the 14-day study period. Peripheral blood mononuclear cells (PMBCs) and neutrophils were isolated and analyzed for membrane fatty acid (FA) content. Mixed linear models and t-tests were used to analyze changes in FA levels over time and FA levels at individual time points, respectively. PK parameters were obtained based on single compartment models of EPA and DHA kinetics., Results: Healthy volunteers were 41.1 ± 10.3 years; 67% were women. In patients with severe sepsis (55.6 ± 13.4 years, 50% women), acute physiologic and chronic health evaluation (APACHE) II score was 27.2 ± 8.8 at ICU admission and median MV duration was 10.5 days. Serum EPA and DHA were significantly lower in sepsis vs. healthy subjects over time. PBMC EPA concentrations were generally not different between groups over time, while PBMC DHA was higher in sepsis patients. Neutrophil EPA and DHA concentrations were similar between groups. The half-life of EPA in serum and neutrophils was significantly shorter in sepsis patients, whereas other half-life parameters did not vary significantly between healthy volunteers and sepsis patients., Conclusions: While incorporation of n-3 FAs into PBMC and neutrophil membranes was relatively similar between healthy volunteers and sepsis patients receiving identical high doses of fish oil for one week, serum EPA and DHA were significantly lower in sepsis patients. These findings imply that serum concentrations and EPA and DHA may not be the dominant driver of leukocyte membrane incorporation of EPA and DHA. Furthermore, lower serum EPA and DHA concentrations suggest that either these n-3 FAs were being metabolized rapidly in sepsis patients or that absorption of enteral medications and pharmaconutrients, including fish oil, may be impaired in sepsis patients. If enteral absorption is impaired, doses of enteral medications administered to critically ill patients may be suboptimal., (Copyright © 2019 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.)

Palmitic acid (PA) is associated with higher blood concentrations of medium-chain acylcarnitines (MCACs), and we hypothesized that PA may inhibit progression of FA β-oxidation. Using a cross-over design, 17 adults were fed high PA (HPA) and low PA/high oleic acid (HOA) diets, each for 3 weeks. The [1-(13)C]PA and [13-(13)C]PA tracers were administered with food in random order with each diet, and we assessed PA oxidation (PA OX) and serum AC concentration to determine whether a higher PA intake promoted incomplete PA OX. Dietary PA was completely oxidized during the HOA diet, but only about 40% was oxidized during the HPA diet. The [13-(13)C]PA/[1-(13)C]PA ratio of PA OX had an approximate value of 1.0 for either diet, but the ratio of the serum concentrations of MCACs to long-chain ACs (LCACs) was significantly higher during the HPA diet. Thus, direct measurement of PA OX did not confirm that the HPA diet caused incomplete PA OX, despite the modest, but statistically significant, increase in the ratio of MCACs to LCACs in blood., (Copyright © 2015 by the American Society for Biochemistry and Molecular Biology, Inc.)

Infectious pneumonias are the leading cause of death worldwide, particularly among immunocompromised patients. Therapeutic stimulation of the lungs' intrinsic defenses with a unique combination of inhaled Toll-like receptor (TLR) agonists broadly protects mice against otherwise lethal pneumonias. As the survival benefit persists despite cytotoxic chemotherapy-related neutropenia, the cells required for protection were investigated. The inducibility of resistance was tested in mice with deficiencies of leukocyte lineages due to genetic deletions and in wild-type mice with leukocyte populations significantly reduced by antibodies or toxins. Surprisingly, these serial reductions in leukocyte lineages did not appreciably impair inducible resistance, but targeted disruption of TLR signaling in the lung epithelium resulted in complete abrogation of the protective effect. Isolated lung epithelial cells were also induced to kill pathogens in the absence of leukocytes. Proteomic and gene expression analyses of isolated epithelial cells and whole lungs revealed highly congruent antimicrobial responses. Taken together, these data indicate that lung epithelial cells are necessary and sufficient effectors of inducible resistance. These findings challenge conventional paradigms about the role of epithelia in antimicrobial defense and offer a novel potential intervention to protect patients with impaired leukocyte-mediated immunity from fatal pneumonias.

The histamine H(1) receptor (Hrh1/H(1)R) was identified as an autoimmune disease gene in experimental allergic encephalomyelitis (EAE), the principal autoimmune model of multiple sclerosis (MS). Previously, we showed that selective re-expression of H(1)R by endothelial cells or T cells in H(1)RKO mice significantly reduced or complemented EAE severity and cytokine responses, respectively. H(1)R regulates innate immune cells, which in turn influences peripheral and central nervous system CD4(+) T cell effector responses. Therefore, we selectively re-expressed H(1)R in CD11b(+) cells of H(1)RKO mice to test the hypothesis that H(1)R signaling in these cells contributes to EAE susceptibility. We demonstrate that transgenic re-expression of H(1)R by H(1)RKO-CD11b(+) cells neither complements EAE susceptibility nor T cell cytokine responses highlighting the cell-specific effects of Hrh1 in the pathogenesis of EAE and MS, and the need for cell-specific targeting in optimizing therapeutic interventions based on such genes., (Published by Elsevier Inc.)

Histamine, a biogenic amine with both neurotransmitter and vasoactive properties, is well recognized as an immunomodulatory agent in allergic and inflammatory reactions. It also plays a regulatory role in the development of antigen-specific immune responses. CD4+ T-cells from histamine H1 receptor (H1R)-deficient (H1RKO) mice produce significantly less interferon-gamma and more interleukin (IL)-4 in in vitro recall assays compared to wild-type controls. H1RKO mice are also less susceptible to acute early-phase experimental allergic encephalomyelitis indicating that H1R signaling in CD4+ T cells plays a central role in regulating pathogenic T-cell responses. In this study, we show that mice lacking histamine H2 receptor (H2RKO) are similar to H1RKO mice in that they develop encephalitogen-specific T-cell responses as assessed by proliferation and IL-2 production and present with less severe acute early-phase experimental allergic encephalomyelitis. However, unlike T cells from H1RKO mice, which exhibit a strong Th2 bias, T cells from H2RKO mice do not. Rather, they are uniquely characterized by a significant inhibition of Th1 effector cell responses. Given that both histamine and adjuvants such as pertussis toxin modulate antigen-presenting cell (APC) maturation and function, including T-cell-polarizing activity, we analyzed the cytokines/chemokines secreted by APCs from wild-type, H1RKO, and H2RKO mice. Significant differences in cytokine/chemokine production by APCs from unimmunized and immunized mice were delineated. APCs from H2RKO mice produce significantly less IL-12 and IL-6 and markedly greater amounts of MCP-1 compared to wild-type and H1RKO mice. Because MCP-1 is known to inhibit IL-12 production, the failure of H2RKO mice to generate encephalitogenic Th1 effector cell responses is consistent with inhibition of negative regulation of MCP-1 secretion by H2R signaling in APCs.

Anthrax lethal toxin (LT) is the principal virulence factor associated with lethal pathologies following infection with Bacillus anthracis. Macrophages are the primary effector cells mediating lethality since macrophage-depleted mice are resistant to LT challenge. Recently, Ltxs1, the gene controlling differential susceptibility of murine macrophages to cytolysis following in vitro exposure to LT, was identified as Kif1c. To directly assess the in vivo role of Kif1c alleles in mortality, we studied a panel of interval-specific recombinant congenic lines carrying various segments of central chromosome 11 derived from LT-resistant DBA/2 mice on the LT-susceptible BALB/c background. The results of this study reveal that mortality is controlled by three linked quantitative trait loci (QTL): Ltxs1/Kif1c (42-43 cM), Ltxs2 (35-37 cM), and Ltxs3 (45-47 cM). The Ltxs3 interval encompasses Nos2, which is an attractive candidate gene for Ltxs3. In this regard, we demonstrate that selective, pharmacologically based inhibition of Nos2 activity in vivo partially overrides genetic resistance to LT and that Nos2 expression as determined by reverse transcription-polymerase chain reaction differs significantly between DBA/2 and BALB/c macrophages. Additionally, to recapitulate dominant resistance to mortality as seen in (BALB/c x DBA/2) F(1) hybrids, DBA/2 alleles are required at all three QTL.

Bronchiolar epithelium is postulated to play a critical role in the orchestration of responses to inhaled allergens, and may contribute to the pathogenesis of asthma. Using a murine model of allergic airway inflammation and hyperresponsiveness, we demonstrate in mice sensitized with ovalbumin (OVA) that following a single challenge with nebulized OVA, a rapid and protracted activation of inhibitor of kappa B kinase (IKK) occurred in lung tissue. IKK activation was followed by nuclear localization of nuclear factor (NF)-kappaB within the bronchiolar epithelium and increased luciferase activity in lungs of mice containing a NF-kappaB-dependent reporter gene. Challenge of sensitized mice with OVA also induced mRNA expression of the chemokines, macrophage inflammatory protein-2 (MIP-2) and eotaxin in lung tissue, which corresponded temporally with the observed influx of neutrophils and eosinophils, respectively, into the airspaces. Using laser capture microdissection and quantitative polymerase chain reaction, we demonstrated that MIP-2 and eotaxin were predominantly expressed in bronchiolar epithelium, in contrast to distal regions of the lungs, which expressed lower or undetectable levels of these mRNAs. These studies strengthen the potential importance of the bronchiolar epithelial cell as a source of production of NF-kappaB-dependent mediators that play a role in asthma.

Alterations in transcription factor activities in aged mice may lead to the production of many inflammatory molecules in the absence of exogenous stimulation. Splenocytes from 22-month-old female C57BL/6 mice are dysregulated in their capacity to control the inducible nitric oxide synthase gene as a result of elevations in the endogenous levels and activity of interferon (IFN)-gamma. Splenocytes from aged mice produced high levels of IFN-gamma in vitro and active STAT-1 was found in nuclear extracts from these splenocytes. Administration to aged mice of neutralizing antibodies against IFN-gamma imposed appropriate regulation over nitric oxide production by stimulated splenocytes. Reestablishment of normal redox balance following dietary supplementation of aged mice with activators of the peroxisome proliferator-activated receptor alpha or the antioxidant alpha-tocopherol (vitamin E) restored appropriate regulation over both the production of IFN-gamma and the secretion of nitric oxide., (Copyright 1999 Academic Press.)

Interleukin (IL)-12 or IL-4 produced early in an immune response directs the differentiation of naive antigen-activated CD4+ T cells down a Th1 or Th2 pathway. The NK1.1+ subset of T cells promptly produces IL-4 following activation in vivo. We demonstrate here that NK1.1+ T cells can be directly induced to produce IL-4 in vitro when activated under serum-free culture conditions. Platelet-derived growth factor in cell culture medium was inhibitory to the production of IL-4 by NK1.1+ T cells in vitro. Lymphocytes obtained from secondary lymphoid organs of aged mice produced greater quantities of IL-4 following stimulation than lymphocytes from mature adult animals. Aged mice expressed elevated percentages of NK1.1+ T cells in their secondary lymphoid organs and peripheral blood. While this cell type was responsible for the total early IL-4 produced by lymphocytes from mature adult mice, both NK1.1+ and memory phenotype (CD44high, CD45RBlow, NK1.1-) T cells from aged donors produced IL-4 following polyclonal T cell activation.