Aryane Cruz Oliveira Pinho

Background/Objectives: Epicardial adipose tissue (EAT) is metabolically active and is in dynamic crosstalk with the surrounding cardiomyocytes, modulating their function and metabolism. Oxidative stress is a key contributor to cell death and cardiac remodeling, is a hallmark of diabetes (DM) and cardiovascular disease, such as coronary artery disease (CAD). However, little is known about these processes in EAT from patients undergoing cardiac surgery. This study investigates changes in mitochondrial dynamics, reactive oxygen species (ROS) production, and antioxidant defense levels in EAT compared to subcutaneous adipose tissue (SAT) in patients undergoing cardiac surgery, with a focus on the impact of DM and CAD. Methods: Adipose tissue biopsies were collected from 128 patients undergoing surgical cardiac intervention. Mitochondrial dynamics and oxidative stress markers were analyzed. Results: EAT exhibited increased expression of mitochondrial fusion markers [mitofusin 1 (p ≤ 0.001), mitofusin 2 (p = 0.038), and optic atrophy 1 (p ≤ 0.001)], as well as fission markers [fission 1 (p ≤ 0.001) and dynamin-related protein 1 (p ≤ 0.001)] relative to SAT. Additionally, ROS levels (dihydroethidium, p = 0.004) were elevated, while lipid peroxidation (malondialdehyde, p ≤ 0.001) was reduced in EAT compared to SAT. Reduced glutathione (GSH) levels (p ≤ 0.001) and the redox buffer ratio between reduced and oxidized glutathione (GSH/GSSG, p ≤ 0.001) were significantly increased in EAT. Interestingly, glutathione peroxidase activity (p ≤ 0.001) and the antioxidant defense markers catalase (p ≤ 0.001) and superoxide dismutase 2 (p = 0.001) were significantly reduced in EAT compared to SAT. Conclusions: The findings provide a unique molecular insight into the mitochondrial dynamics and oxidative stress profiles of EAT, highlighting potential avenues for a novel diagnostic method and therapeutic strategies for cardiac disease.

Obesity is associated with insulin resistance (IR) development, a risk factor for type 2 diabetes (T2D). How mitochondrial bioenergetics, in adipose tissue (AT), differs according to distinct metabolic profiles (i.e. insulin sensitive (IS), IR normoglycaemic (IR‐NG), pre‐diabetes (PD) and T2D) is still poorly understood. The purpose of this study was to evaluate and compare bioenergetics and energy substrate preference by omental AT (OAT) and subcutaneous AT (SAT) from subjects with obesity (OB, n = 40) at distinct metabolic stages. Furthermore, AT bioenergetics was also evaluated pre‐ and post‐bariatric/metabolic surgery (BMS). High‐resolution respirometry (HRR) was used to measure the real‐time oxidative phosphorylation (OXPHOS) capacity and mitochondrial substrate preferences in both tissues. Substrate‐uncoupler‐inhibitor titration protocols were used: SUIT‐P1 (complex I and II‐linked mitochondrial respiration) and SUIT‐P2 (fatty acid oxidation (FAO)‐linked mitochondrial respiration). Flux control ratios (FCRs) were calculated. In SUIT‐P1, lower OXPHOS capacity was observed in AT, particularly in SAT, during the establishment of IR (OB‐IR‐NG) and in the T2D group, due to alterations of mitochondrial coupling, evaluated by FCRs. In SUIT‐P2, the OXPHOS coupling efficiency was highest in the OB‐IR‐NG group. AT from OB‐IS, OB‐IR‐NG and OB‐IR‐PD preferred pyruvate, malate and glutamate oxidation and/or FAO during OXPHOS, whereas AT from T2D preferred succinate oxidation. BMS enhanced mitochondrial respiration in OAT, even under poor OXPHOS coupling efficiency. In conclusion, real‐time OXPHOS analysis by HRR may be a sensitive biomarker of mitochondrial fitness, particularly in AT. Interventions based on modulating energetic substrate availability may become a good tool for obesity treatment stratification. image Key points: Omental adipose tissue shows higher oxidative phosphorylation (OXPHOS) capacity compared to subcutaneous adipose tissue in paired explants from subjects with obesity. The OXPHOS capacity of adipose tissue differs through the progression of metabolic disease. Subjects with obesity and diabetes have the lowest OXPHOS capacity in paired explants of subcutaneous and omental adipose tissues. Bariatric surgery enhanced the OXPHOS capacity in omental adipose tissue, even under poor OXPHOS coupling efficiency. Assessment of the oxidative capacity in fresh adipose tissue explants could be a sensitive tool for early diagnosis of metabolic disease.

CD20⁺ T cells constitute a small subset of T cells. These are found among CD4⁺, CD8⁺, CD4⁺CD8⁺, CD4⁻CD8⁻ T, and TCRγδ⁺ T cells, and have been poorly characterized. The aim of this study was to characterize peripheral blood (PB) CD20⁺ T cells and compare them to their PB CD20⁻ T cell counterparts. PB from 17 healthy individuals was collected. The distribution of CD20⁺ T cells among maturation‐associated T cells compartments (naïve, central memory, transitional memory, effector memory, and effector T cells), their polarization, activation status, and expression of immune‐regulatory proteins were evaluated by flow cytometry. Their function was also assessed, by measuring IFN‐γ, TNF‐α, and IL‐17 production. Compared with CD20⁻ T cells, CD20⁺ T cells represent a higher proportion of transitional memory cells. Furthermore, CD20⁺ T cells display a proinflammatory phenotype, characterized by the expansion of Th1, Th1/17, and Tc1 cell subsets , associated to a high expression of activation (CD25) and exhaustion (PD‐1) markers. In addition, the simultaneous production of the proinflammatory cytokines IFN‐γ, TNF‐α, and IL‐17 was also detected in CD4⁺CD20⁺ T cells. Our results show that CD20⁺ T cells are phenotypically and functionally different from CD20⁻ T cells, suggesting that these cells are a distinct subset of T cells.

Low-grade inflammation is closely linked to obesity and obesity-related comorbidities; therefore, immune cells have become an important topic in obesity research. Here, we performed a deep phenotypic characterization of circulating T cells in people with obesity, using flow cytometry. Forty-one individuals with obesity (OB) and clinical criteria for bariatric surgery were enrolled in this study. We identified and quantified 44 different circulating T cell subsets and assessed their activation status and the expression of immune-checkpoint molecules, immediately before (T1) and 7–18 months after (T2) the bariatric surgery. Twelve age- and sex-matched healthy individuals (nOB) were also recruited. The OB participants showed higher leukocyte counts and a higher percentage of neutrophils. The percentage of circulating Th1 cells were negatively correlated to HbA1c and insulin levels. OB Th1 cells displayed a higher activation status and lower PD-1 expression. The percentage of Th17 and Th1/17 cells were increased in OB, whereas the CD4⁺ Tregs’ percentage was decreased. Interestingly, a higher proportion of OB CD4⁺ Tregs were polarized toward Th1- and Th1/17-like cells and expressed higher levels of CCR5. Bariatric surgery induced the recovery of CD4⁺ Treg cell levels and the expansion and activation of Tfh and B cells. Our results show alterations in the distribution and phenotype of circulating T cells from OB people, including activation markers and immune-checkpoint proteins, demonstrating that different metabolic profiles are associated to distinct immune profiles, and both are modulated by bariatric surgery.

Despite the known link between obesity and insulin resistance (IR) to chronic low-grade inflammation, new markers capable of early IR detection are needed. Immune cells are components of adipose tissue’s (AT) stromal vascular fraction (SVF) that regulate AT homeostasis. The altered phenotype and function of AT-infiltrating immune cells may contribute to the development and maintenance of local AT inflammation observed under obesity-induced IR conditions. Impaired AT-specific immunometabolic function may influence the whole organism. Therefore, AT-infiltrating immune cells may be important players in the development of obesity-related metabolic complications, such as type 2 diabetes mellitus (T2DM). B and T cells, particularly CD20⁺ T cells, play important roles in human pathology, such as autoimmune disease and cancer. However, the question remains as to whether CD20⁺ T cells have an important contribution to the development of obesity-related IR. While circulating CD20⁺ T cells are mostly of the central memory phenotype (i.e. antigen-experienced T cells with the ability to home to secondary lymphoid organs), tissues-infiltrated CD20⁺ T cells are predominantly of the effector memory phenotype (i.e. antigen-experienced T cells that preferentially infiltrate peripheral tissues). The latter produce pro-inflammatory cytokines, such as IFN-γ and IL-17, which play a role in obesity-related IR development. This review describes the CD20 molecule and its presence in both B and T cells, shedding light on its ontogeny and function, in health and disease, with emphasis on AT. The link between CD20⁺ T cell dysregulation, obesity, and IR development supports the role of CD20⁺ T cells as markers of adipose tissue dysmetabolism.

Introduction: Diabetes mellitus (DM) impairs wound healing. The aim was to determine whether DM influences mitochondrial respiration in wounded skin (WS) and non‐wounded skin (NWS), in a pre‐clinical wound healing model of streptozotocin (STZ)‐induced diabetes.Methods: Six weeks after diabetes induction, two wounds were created in the back of C57BL/J6 mice. Using high‐resolution respirometry (HRR), oxygen flux was measured, in WS and NWS, using two substrate‐uncoupler‐inhibitor titration protocols, at baseline (day 0), day 3 and 10 post‐wounding, in STZ‐DM and non‐diabetic (NDM) mice. Flux control ratios for the oxidative phosphorylation (OXPHOS) capacity were calculated.Results: A significant increase in mitochondrial respiration was observed in STZ‐DM skin compared to control skin at baseline. The OXPHOS capacity was decreased in WS under diabetes at day 3 post‐wounding (inflammation phase). However, at day 10 post‐wounding (remodeling phase), the OXPHOS capacity was higher in WS from STZ‐DM compared to NDM mice, and compared to NWS from STZ‐DM mice. A significant relative contribution of pyruvate, malate and glutamate (PMG) oxidation to the OXPHOS capacity was observed in WS compared to NWS from STZ‐DM mice, at day 10, while the relative contribution of fatty acid oxidation to the OXPHOS capacity was higher in NWS. The OXPHOS capacity is altered in WS from STZ‐DM compared to NDM mice across the healing process, and so is the substrate contribution in WS and NWS from STZ‐DM mice, at each time point.Conclusion: HRR may be a sensitive tool to evaluate the underlying mechanisms of tissue repair during wound healing.

Introduction: Thoracic perivascular adipose tissue (tPVAT) has a phenotype resembling brown AT. Dysfunctional tPVAT appears to be linked to vascular dysfunction. Methods: We evaluated uncoupling protein (UCP)-1 expression by western blot, oxidative stress by measuring lipid peroxidation and the antioxidant capacity by high-performance liquid chromatography and spectrophotometry, and mitochondrial respiration by high-resolution respirometry (HRR) in tPVAT, compared to inguinal white AT (iWAT), obtained from non-diabetic (NDM) and streptozocin-induced diabetic (STZ-DM) mice. Mitochondrial respiration was assessed by HRR using protocol 1: Complex I and II oxidative phosphorylation (OXPHOS) and protocol 2: Fatty acid oxidation (FAO) OXPHOS. OXPHOS capacity in tPVAT was also evaluated after UCP-1 inhibition by guanosine 5'-diphosphate (GDP). Results: UCP-1 expression was higher in tPVAT when compared with iWAT in both NDM and STZ-DM mice. The malondialdehyde concentration was elevated in tPVAT from STZ-DM compared to NDM mice. Glutathione peroxidase and reductase activities, as well as reduced glutathione levels, were not different between tPVAT from NDM and STZ-DM mice but were lower compared to iWAT of STZ-DM mice. OXPHOS capacity of tPVAT was significantly decreased under UCP-1 inhibition by GDP in protocol 1. While there were no differences in the OXPHOS capacity between NDM and STZ-DM mice in protocol 1, it was increased in STZ-DM compared to NDM mice in protocol 2. Moreover, complex II- and FAO- linked respiration were elevated in STZ-DM mice under UCP-1 inhibition. Conclusions: Pharmacological therapies could be targeted to modulate UCP-1 activity with a significant impact in the uncoupling of mitochondrial bioenergetics in tPVAT.

A broad understanding on how SARS-CoV-2 infection and vaccination mobilize the immune system is necessary to find the best predictors of long-term protection and identify individuals that would benefit from additional vaccine doses. This study aims to understand the effect of a single dose of Pfizer-BioNTech BNT162b2 COVID-19 vaccine, in individuals recovered from SARS-CoV-2 infection, on circulating CD4⁺ T follicular helper (Tfh)-cells, Spike-specific T-cells and IgG/IgA antibodies. For that, peripheral blood samples from 50 healthcare professionals, recovered from SARS-CoV-2 infection, collected immediately before (T1) and 15 days after (T2) vaccine administration, were used to analyze the frequency and numbers of Tfh-cells and their subsets, serum titers of SARS-CoV-2-specific antibodies, and SARS-CoV-2-specific T-cells. Six months after infection (T1), 96% of recovered participants presented either IgG or T-cells specific for Spike, however, Spike-specific T-cells were missing in 16% of them. These individuals presented lower levels of Spike-specific IgG (T1 and T2), IgA (T1), and Spike-specific T-cells (T2). Vaccination increased the percentage of participants reactive for Spike-specific T-cells (from 64 to 98%), IgG (from 90 to 100%) and IgA (from 48 to 98%). It also mobilized circulating Tfh-cells, increasing their frequency and activation, and promoting Tfh17 polarization, restoring the decreased numbers of Tfh-cells (especially Tfh17) observed in recovered participants. Interestingly, Tfh percentage correlated with Spike-specific IgG levels. Our data showed that a single dose of vaccine efficiently restored Spike-specific T-cells, and IgG and IgA antibodies. Mobilization of Tfh-cells, and their correlation with IgG levels, suggest that vaccination induced a functional Tfh cell response.

Research in pharmacological therapy has led to the availability of many antidiabetic agents. New recommendations for precision medicine and particularly precision nutrition may greatly contribute to the control and especially to the prevention of diabetes. This scenario greatly encourages the search for novel non-pharmaceutical molecules. In line with this, the daily and long-term consumption of diets rich in phenolic compounds, together with a healthy lifestyle, may have a protective role against the development of type 2 diabetes. In the framework of the described studies, there is clear evidence that the bio accessibility, bioavailability, and the gut microbiota are indeed affected by: the way phenolic compounds are consumed (acutely or chronically; as pure compounds, extracts, or in-side a whole meal) and the amount and the type of phenolic compounds (ex-tractable or non-extractable/macromolecular antioxidants, including non-bioavailable polyphenols and plant matrix complexed structures). In this review, we report possible effects of important, commonly consumed, phenolic-based nutraceuticals in pre-clinical and clinical diabetes studies. We highlight their mechanisms of action and their potential effects in health promotion. Translation of this nutraceutical-based approach still requires more and larger clinical trials for better elucidation of the mechanism of action toward clinical applications.