Preexercise Breakfast Ingestion versus Extended Overnight Fasting Increases Postprandial Glucose Flux after Exercise in Healthy Men

Aims To characterize postprandial glucose flux after exercise in the fed versus overnight fasted-state and to investigate potential underlying mechanisms. Methods In a randomized order, twelve men underwent breakfast-rest (BR; 3 h semi-recumbent), breakfast-exercise (BE; 2 h semi-recumbent before 60-min of cycling (50% peak power output) and overnight fasted-exercise (FE; as per BE omitting breakfast) trials. An oral glucose tolerance test (OGTT) was completed post-exercise (post-rest on BR). Dual stable isotope tracers ([U-13C] glucose ingestion and [6,6-2H2] glucose infusion) and muscle biopsies were combined to assess postprandial plasma glucose kinetics and intramuscular signaling, respectively. Plasma intestinal fatty acid binding (I-FABP) concentrations were determined as a marker of intestinal damage. Results Breakfast before exercise increased post-exercise plasma glucose disposal rates during the OGTT, from 44 g•120 min-1 in FE [35 to 53 g•120 min-1] (mean [normalized 95% CI]) to 73 g•120 min-1 in BE [55 to 90 g•120 min-1; p = 0.01]. This higher plasma glucose disposal rate was, however, offset by increased plasma glucose appearance rates (principally OGTT-derived), resulting in a glycemic response that did not differ between BE and FE (p = 0.11). Plasma I-FABP concentrations during exercise were 264 pg•mL-1 [196 to 332 pg•mL-1] lower in BE versus FE (p = 0.01). Conclusion Breakfast before exercise increases post-exercise postprandial plasma glucose disposal, which is offset (primarily) by increased appearance rates of orally-ingested glucose. Therefore, metabolic responses to fed-state exercise cannot be readily inferred from studies conducted in a fasted state

Interventions targeting glucocorticoid-Krüppel-like factor 15-branched-chain amino acid signaling improve disease phenotypes in spinal muscular atrophy mice

The circadian glucocorticoid-Krüppel-like factor 15-branched-chain amino acid (GC-KLF15-BCAA) signaling pathway is a key regulatory axis in muscle, whose imbalance has wide-reaching effects on metabolic homeostasis. Spinal muscular atrophy (SMA) is a neuromuscular disorder also characterized by intrinsic muscle pathologies, metabolic abnormalities and disrupted sleep patterns, which can influence or be influenced by circadian regulatory networks that control behavioral and metabolic rhythms. We therefore set out to investigate the contribution of the GC-KLF15-BCAA pathway in SMA pathophysiology of Taiwanese Smn−/−;SMN2 and Smn2B/− mouse models. We thus uncover substantial dysregulation of GC-KLF15-BCAA diurnal rhythmicity in serum, skeletal muscle and metabolic tissues of SMA mice. Importantly, modulating the components of the GC-KLF15-BCAA pathway via pharmacological (prednisolone), genetic (muscle-specific Klf15 overexpression) and dietary (BCAA supplementation) interventions significantly improves disease phenotypes in SMA mice. Our study highlights the GC-KLF15-BCAA pathway as a contributor to SMA pathogenesis and provides several treatment avenues to alleviate peripheral manifestations of the disease. The therapeutic potential of targeting metabolic perturbations by diet and commercially available drugs could have a broader implementation across other neuromuscular and metabolic disorders characterized by altered GC-KLF15-BCAA signaling

CD4(+)CD25(+)FOXP3(+) Regulatory T Cells Suppress Anti-Tumor Immune Responses in Patients with Colorectal Cancer

BACKGROUND: A wealth of evidence obtained using mouse models indicates that CD4(+)CD25(+)FOXP3(+) regulatory T cells (Treg) maintain peripheral tolerance to self-antigens and also inhibit anti-tumor immune responses. To date there is limited information about CD4(+) T cell responses in patients with colorectal cancer (CRC). We set out to measure T cell responses to a tumor-associated antigen and examine whether Treg impinge on those anti-tumor immune responses in CRC patients. METHODOLOGY AND PRINCIPAL FINDINGS: Treg were identified and characterized as CD4(+)CD25(+)FOXP3(+) using flow cytometry. An increased frequency of Treg was demonstrated in both peripheral blood and mesenteric lymph nodes of patients with colorectal cancer (CRC) compared with either healthy controls or patients with inflammatory bowel disease (IBD). Depletion of Treg from peripheral blood mononuclear cells (PBMC) of CRC patients unmasked CD4(+) T cell responses, as observed by IFNγ release, to the tumor associated antigen 5T4, whereas no effect was observed in a healthy age-matched control group. CONCLUSIONS/SIGNIFICANCE: Collectively, these data demonstrate that Treg capable of inhibiting tumor associated antigen-specific immune responses are enriched in patients with CRC. These results support a rationale for manipulating Treg to enhance cancer immunotherapy

Lipid Metabolism Links Nutrient-Exercise Timing to Insulin Sensitivity in Men Classified as Overweight or Obese

Context Pre-exercise nutrient availability alters acute metabolic responses to exercise, which could modulate training responsiveness. Objective To assess acute and chronic effects of exercise performed before versus after nutrient ingestion on whole-body and intramuscular lipid utilization and postprandial glucose metabolism. Design (1) Acute, randomized, crossover design (Acute Study); (2) 6-week, randomized, controlled design (Training Study). Setting General community. Participants Men with overweight/obesity (mean ± standard deviation, body mass index: 30.2 ± 3.5 kg⋅m-2 for Acute Study, 30.9 ± 4.5 kg⋅m-2 for Training Study). Interventions Moderate-intensity cycling performed before versus after mixed-macronutrient breakfast (Acute Study) or carbohydrate (Training Study) ingestion. Results Acute Study—exercise before versus after breakfast consumption increased net intramuscular lipid utilization in type I (net change: –3.44 ± 2.63% versus 1.44 ± 4.18% area lipid staining, P 0.05). However, postprandial insulinemia was reduced with exercise training performed before but not after carbohydrate ingestion (P = 0.03). This resulted in increased oral glucose insulin sensitivity (25 ± 38 vs –21 ± 32 mL⋅min-1⋅m-2; P = 0.01), associated with increased lipid utilization during exercise (r = 0.50, P = 0.02). Regular exercise before nutrient provision also augmented remodeling of skeletal muscle phospholipids and protein content of the glucose transport protein GLUT4 (P < 0.05). Conclusions Experiments investigating exercise training and metabolic health should consider nutrient-exercise timing, and exercise performed before versus after nutrient intake (ie, in the fasted state) may exert beneficial effects on lipid utilization and reduce postprandial insulinemia

Multiorgan MRI findings after hospitalisation with COVID-19 in the UK (C-MORE): a prospective, multicentre, observational cohort study

Introduction: The multiorgan impact of moderate to severe coronavirus infections in the post-acute phase is still poorly understood. We aimed to evaluate the excess burden of multiorgan abnormalities after hospitalisation with COVID-19, evaluate their determinants, and explore associations with patient-related outcome measures. Methods: In a prospective, UK-wide, multicentre MRI follow-up study (C-MORE), adults (aged ≥18 years) discharged from hospital following COVID-19 who were included in Tier 2 of the Post-hospitalisation COVID-19 study (PHOSP-COVID) and contemporary controls with no evidence of previous COVID-19 (SARS-CoV-2 nucleocapsid antibody negative) underwent multiorgan MRI (lungs, heart, brain, liver, and kidneys) with quantitative and qualitative assessment of images and clinical adjudication when relevant. Individuals with end-stage renal failure or contraindications to MRI were excluded. Participants also underwent detailed recording of symptoms, and physiological and biochemical tests. The primary outcome was the excess burden of multiorgan abnormalities (two or more organs) relative to controls, with further adjustments for potential confounders. The C-MORE study is ongoing and is registered with ClinicalTrials.gov, NCT04510025. Findings: Of 2710 participants in Tier 2 of PHOSP-COVID, 531 were recruited across 13 UK-wide C-MORE sites. After exclusions, 259 C-MORE patients (mean age 57 years [SD 12]; 158 [61%] male and 101 [39%] female) who were discharged from hospital with PCR-confirmed or clinically diagnosed COVID-19 between March 1, 2020, and Nov 1, 2021, and 52 non-COVID-19 controls from the community (mean age 49 years [SD 14]; 30 [58%] male and 22 [42%] female) were included in the analysis. Patients were assessed at a median of 5·0 months (IQR 4·2–6·3) after hospital discharge. Compared with non-COVID-19 controls, patients were older, living with more obesity, and had more comorbidities. Multiorgan abnormalities on MRI were more frequent in patients than in controls (157 [61%] of 259 vs 14 [27%] of 52; p&lt;0·0001) and independently associated with COVID-19 status (odds ratio [OR] 2·9 [95% CI 1·5–5·8]; padjusted=0·0023) after adjusting for relevant confounders. Compared with controls, patients were more likely to have MRI evidence of lung abnormalities (p=0·0001; parenchymal abnormalities), brain abnormalities (p&lt;0·0001; more white matter hyperintensities and regional brain volume reduction), and kidney abnormalities (p=0·014; lower medullary T1 and loss of corticomedullary differentiation), whereas cardiac and liver MRI abnormalities were similar between patients and controls. Patients with multiorgan abnormalities were older (difference in mean age 7 years [95% CI 4–10]; mean age of 59·8 years [SD 11·7] with multiorgan abnormalities vs mean age of 52·8 years [11·9] without multiorgan abnormalities; p&lt;0·0001), more likely to have three or more comorbidities (OR 2·47 [1·32–4·82]; padjusted=0·0059), and more likely to have a more severe acute infection (acute CRP &gt;5mg/L, OR 3·55 [1·23–11·88]; padjusted=0·025) than those without multiorgan abnormalities. Presence of lung MRI abnormalities was associated with a two-fold higher risk of chest tightness, and multiorgan MRI abnormalities were associated with severe and very severe persistent physical and mental health impairment (PHOSP-COVID symptom clusters) after hospitalisation. Interpretation: After hospitalisation for COVID-19, people are at risk of multiorgan abnormalities in the medium term. Our findings emphasise the need for proactive multidisciplinary care pathways, with the potential for imaging to guide surveillance frequency and therapeutic stratification

Transiently activated human regulatory T cells upregulate BCL-XL expression and acquire a functional advantage in vivo

Regulatory T cells (Tregs) can control excessive or undesirable immune responses toward autoantigens, alloantigens, and pathogens. In transplantation, host immune responses against the allograft are suppressed through the use of immunosuppressive drugs, however this often results in life-threatening side effects including nephrotoxicity and an increased incidence of cancer and opportunistic infections. Tregs can control graft-vs.-host disease and transplant rejection in experimental models, providing impetus for the use of Tregs as a cellular therapy in clinical transplantation. One of the major barriers to the widespread use of Treg cellular therapy is the requirement to expand cells ex vivo to large numbers in order to alter the overall balance between regulatory and effector cells. Methods that enhance suppressive capacity thereby reducing the need for expansion are therefore of interest. Here, we have compared the function of freshly-isolated and ex vivo-manipulated human Tregs in a pre-clinical humanized mouse model of skin transplantation. Sorted human CD127loCD25+CD4+ Tregs were assessed in three different conditions: freshly-isolated, following transient in vitro activation with antiCD3/antiCD28 beads or after ex vivo-expansion for 2 weeks in the presence of antiCD3/antiCD28 beads and recombinant human IL2. While ex vivo-expansion of human Tregs increased their suppressive function moderately, transient in vitro-activation of freshly isolated Tregs resulted in a powerful enhancement of Treg activity sufficient to promote long-term graft survival of all transplants in vivo. In order to investigate the mechanisms responsible for these effects, we measured the expression of Treg-associated markers and susceptibility to apoptosis in activated Tregs. Transiently activated Tregs displayed enhanced survival and proliferation in vitro and in vivo. On a molecular level, Treg activation resulted in an increased expression of anti-apoptotic BCL2L1 (encoding BCL-XL) which may be at least partially responsible for the observed enhancement in function. Our results suggest that in vitro activation of human Tregs arms them with superior proliferative and survival abilities, enabling them to more effectively control alloresponses. Importantly, this transient activation results in a rapid functional enhancement of freshly-isolated Tregs, thereby providing an opportunity to eliminate the need for in vitro expansion in select circumstances. A protocol employing this technique would therefore benefit from a reduced requirement for large cell numbers for effective therapy

Assessing the prognostic value of preoperative carcinoembryonic antigen-specific T-cell responses in colorectal cancer

Current dogma suggests that tumor-reactive IFN-γ–producing (TH1-type) T-cells are beneficial to patient outcome; however, the clinical consequence of these responses with respect to long-term prognosis in colorectal cancer (CRC) is not understood. Here, we compared the utility of preoperative, peripheral blood–derived IFN-γ+ T-cell responses specific to carcinoembryonic antigen (CEA), 5T4, or control antigens (n = 64) with tumor staging and clinical details (n = 87) in predicting five-year outcome of CRC patients who underwent resection with curative intent. Although disease recurrence was more likely in patients with stage III tumors, the presence of preoperative, CEA-specific IFN-γ–producing T-cells identified patients at a statistically significantly greater risk of tumor recurrence following surgical resection, irrespective of tumor stage (odds ratio = 5.00, 95% confidence interval = 1.96 to 12.77, two-sided P <.001). Responses to other antigens, including 5T4, did not reflect outcome. Whilst these results initially appear surprising, they could improve prognostication and help redirect adjuvant treatments

CD59 blockade enhances antigen-specific CD4+ T cell responses in humans: a new target for cancer immunotherapy?

CD59, a broadly expressed GPI-anchored molecule, regulates formation of the membrane attack complex of the complement cascade. We previously demonstrated that mouse CD59 also down-modulates CD4+ T cell activity in vivo. In this study, we explored the role of CD59 on human CD4+ T cells. Our data demonstrate that CD59 is up-regulated on activated CD4+ T cells and serves to down-modulate their activity in response to polyclonal and Ag- specific stimulation. The therapeutic potential of this finding was explored using T cells isolated from colorectal cancer patients. The findings were striking and indicated that blockade of CD59 significantly enhanced the CD4+ T cell response to two different tumor Ags. These data highlight the potential for manipulating CD59 expression on T cells for boosting weak immune responses, such as those found in individuals with cancer

Effect of cholera toxin on BCG immunogenicity and <i>M.tb</i> challenge outcome.

<p>(<b>A</b>) Cytokine gating in spleens and lungs, unstimulated vs. Ag85A peptide pool-stimulated. (<b>B</b>) Balb/c mice received 4×10⁵ CFU BCG <i>i.n.</i> or <i>i.d.</i> followed 10 weeks later by 1×10⁶ PFU MVA85A <i>i.d.</i> At weeks 10, 11 and 14 post-BCG, lungs were examined for cytokine production by ICS following Ag85A peptide pool stimulation in the presence of Brefeldin A and GolgiStop. (<b>C</b> & <b>D</b>) Balb/c mice received 4×10⁵ CFU BCG±2 µg CT <i>i.n</i>. After 10 weeks, lungs and spleen were dissected and stimulated with PPD in the presence of Brefeldin A and GolgiStop. Percentages of CD4⁺ T cells producing IFN-γ and IL-17 were calculated following ICS on lung cells (<b>C</b>) and splenocytes (<b>D</b>). Mice receiving BCG only are plotted with closed circles and those receiving CT have open circles. P values were calculated using a Mann Whitney test (n = 5). (<b>E</b>) Ten weeks post-BCG, mice received ∼100 CFU <i>M.tb</i> via aerosol and four weeks later lungs and spleen were homogenised and plated out for CFU quantitation. Statistical analysis was performed using a one way ANOVA and post-hoc tests on the vaccinated groups (n = 8).</p