A reliable, reproducible flow cytometry protocol for immune cell quantification in human adipose tissue.

The ability to accurately identify and quantify immune cell populations within adipose tissue is important in understanding the role of immune cells in metabolic disease risk. Flow cytometry is the gold standard method for immune cell quantification. However, quantification of immune cells from adipose tissue presents a number of challenges because of the complexities of working with an oily substance and the rapid deterioration of immune cell viability before analysis can be performed. Here we present a highly reproducible flow cytometry protocol for the quantification of immune cells in human adipose tissue, which overcomes these issues.

Substrate Oxidation Is Altered by Obesity During Submaximal Cycling in Prepubertal and Early Pubertal Children: A Quality Study.

BACKGROUND: To examine substrate oxidation in prepubertal and early pubertal children as a function of body weight, body composition, and sex during an exhaustive cycling test. METHODS: This study included 320 children in prepubertal and early puberty (Tanner stage 1 or 2; n = 188 males) who completed a minimum of 4 stages (2-5 min/stage) of an adapted version of the McMaster exhaustive exercise protocol on an upright cycle ergometer. Substrate utilization, relative to individual VO2peak, was determined using VO2 and VCO2 data, obtained with breath-by-breath gas analysis during exercise. RESULTS: Both peak (mg/kg lean body mass·min) and submaximal lipid oxidation (mg/kg lean body mass·min) were highest (P < .01) in children with healthy weight (HW), then overweight, and lowest in obese (OB). Both females with HW (compared with males with HW) and females with OB (compared with males with OB) had higher (P < .01) peak and submaximal lipid oxidation. In children with OB, fat-free mass correlated positively (P < .01) with submaximal lipid oxidation (r = .50). In contrast, in children with HW and overweight, fat-free mass correlated positively (P < .01) with carbohydrate oxidation (r = .52 and r = .47, respectively). CONCLUSION: Obesity during childhood may alter substrate oxidation during exercise. These results may have implications in the implementation of exercise programs in prepubertal or early puberty to control adiposity.

Sex differences in regional adipose tissue depots pose different threats for the development of Type 2 diabetes in males and females.

Type 2 diabetes mellitus (T2DM) affects males and females disproportionately. In midlife, more males have T2DM than females. The sex difference in T2DM prevalence is, in part, explained by differences in regional adipose tissue characteristics. With obesity, changes to regional adipokine and cytokine release increases the risk of T2DM in both males and females with males having greater levels of TNFα and females having greater levels of leptin, CRP, and adiponectin. Regional immune cell infiltration appears to be pathogenic in both sexes via different routes as males with obesity have greater VAT ATM and a decrease in the protective Treg cells, whereas females have greater SAT ATM and T cells. Lastly, the ability of female adipose tissue to expand all regions through hyperplasia, rather than hypertrophy, protects them against the development of large insulin-resistant adipocytes that dominate male adipose tissue. The objective of this review is to discuss how sex may affect regional differences in adipose tissue characteristics and how these differences may distinguish the development of T2DM in males and females. In doing so, we will show that the origins of T2DM development differ between males and females.

Teaching Culturally Safe Care in Simulated Cultural Communication Scenarios During the COVID-19 Pandemic: Virtual Visits with Indigenous Animators.

Clinical learning activities involving Indigenous patient actors that specifically address the development of culturally safe care skills among medical students are important in order to improve health care for Indigenous people. In 2020, the COVID-19 pandemic led to strict physical distancing regulations and regional lockdowns that made the in-person delivery of Simulated Cultural Communication Scenarios (SCCS) with Indigenous patient actors impossible due to the disproportionate risk that public health emergencies pose for Indigenous communities. As the pandemic continued in 2021, we co-created a Virtual Visit approach to SCCS for the education of culturally safe care to pre-clerkship medical students. We report on student and tutor evaluation of these virtual sessions and contextualize our findings with our previous results delivering In-Person SCCSs. We found that Virtual Visit SCCS were highly effective in providing authentic exposure to and feedback from Indigenous patients. However, students rated their learning outcomes with Virtual Visit lower than the In-person approach to SCCS. We recommend formal training on interacting with patients in virtual care scenarios prior to Virtual Visit SCCS. We also found that exposure to SCCS with Indigenous animators has the potential to conjure up a diverse spectrum of sometimes unresolved negative feelings related to colonialism among students and tutors including discomfort, embarrassment, and anxiety. Our findings underscore the importance of resolving these sentiments within the safe environment of a classroom. To prepare Indigenous as well as non-Indigenous students and tutors adequately, it is important to acknowledge and critically deconstruct the embodiment of colonialism and Indigenous-settler relations when teaching physicians, as well as future physicians, preparedness for culturally safe care of Indigenous peoples.

Association between rs174537 FADS1 polymorphism and immune cell profiles in abdominal and femoral subcutaneous adipose tissue: an exploratory study in adults with obesity.

Fatty acid desaturase 1 (FADS1) polymorphisms alter fatty acid content in subcutaneous adipose tissue (SAT); however, existing evidence is limited and conflicting regarding the association between FADS1 variants and SAT inflammatory status. To advance this area, we conducted an exploratory study to investigate whether the common rs174537 polymorphism in FADS1 was associated with immune cell profiles in abdominal and femoral SAT in individuals with obesity. FADS1 gene expression and immune cell profiles in SAT depots were assessed by qPCR and flow cytometry, respectively. Although FADS1 gene expression was associated with genotype, no associations were observed with immune cell profiles in either depot. Our study provides additional evidence that rs174537 in FADS1 has minimal impact on inflammatory status in obese SAT.

Sex Affects Regional Variations in Subcutaneous Adipose Tissue T Cells but not Macrophages in Adults with Obesity.

OBJECTIVE: The inflammatory environment in lower-body subcutaneous adipose tissue (SAT) has been largely unexplored. This study aimed to examine the effects of region (upper body vs. lower body) and sex on SAT immune cell profiles in young adults with obesity. METHODS: Abdominal (AB) and femoral (FEM) SAT was collected from 12 males (mean [SEM] age = 30.8 [1.4] years; mean [SEM] BMI = 34.1 [1.1] kg/m2 ) and 22 females (mean [SEM] age = 30.6 [0.6] years; mean [SEM] BMI = 34.0 [0.7] kg/m2 ) with obesity via needle aspiration. Flow cytometry was used to quantify macrophage (CD68+) and T-cell (CD3+) subpopulations in the stromovascular fraction of each SAT region. RESULTS: Females had a greater proportion of most T-cell types (CD3+CD4+CD45RA+, CD3+CD4+CD45RA-, and CD3+CD8+CD45RA+) in FEM compared with AB SAT, while males had similar proportions in both regions. Regardless of sex, the M1-like macrophage population (CD68+CD206-) was proportionally higher in AB SAT than in FEM SAT. CONCLUSIONS: Results showed that T-cell populations vary by SAT region in females but not males. Both sexes, however, have proportionately more proinflammatory macrophages in upper-body than in lower-body SAT. It remains to be seen how these unique immune cell profiles in males and females with obesity contribute to adipose tissue inflammation and metabolic disease risk.

Regional adiposity and markers of inflammation in pre-school age children.

In adults, upper body fat partially increases metabolic disease risk through increasing systemic inflammation. Our objective was to determine if this relationship exists in preschool-aged children. A subset of children (n = 71, 35 males), 3.7 ± 1.0 y, were studied from n = 515 children recruited from randomly selected daycares in Montréal, QC. According to WHO charts for 2-5 y, 49 children were healthy weight (HW) and 21 were overweight (OW). Adiposity was determined through dual-energy x-ray absorptiometry. Blood concentrations of C-reactive protein (CRP) and tumour necrosis factor alpha (TNFα) were determined via enzyme-linked immunosorbent and multiplex assays, respectively. OW children had higher (p = 0.03) android:gynoid ratio 0.50 ± 0.09 compared to HW children 0.56 ± 0.12, indicating excess fat was predominantly stored in the abdominal depot. CRP was higher (p = 0.01) in OW children 1.45 ± 2.02 mg/L compared to HW 0.74 ± 1.38 mg/L. Percent fat was correlated with CRP (r = 0.32; p < 0.01) and TNFα (r = 0.25; p = 0.04) concentrations. CRP also correlated with android adiposity (r = 0.24; p = 0.04) and TNFα correlated with gynoid adiposity (r = 0.24; p = 0.04). We observed that greater adiposity is associated with higher systemic inflammation in pre-school aged children. Future longitudinal studies are needed to understand the long term consequences of excess total and regional body fat in young children.