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Spermidine mitigates immune cell senescence, enhances autophagy, and boosts vaccine responses in healthy older adults
Get PDFOlder adults are particularly vulnerable to infectious diseases, and vaccines are often less effective in this population due to immunosenescence, which is characterized by diminished B and T memory responses. Autophagy is believed to underlie many facets of cellular aging, including immunosenescence. It is crucial for maintaining memory T and B cell functions but declines with age, along with the endogenous metabolite spermidine that helps maintain autophagy levels. We conducted a double-blind, randomized, placebo-controlled study in 40 volunteers over 65, administering oral spermidine after their third SARS-CoV-2 vaccine dose. Spermidine reduced immune cell senescence, evidenced by decreased p16 expression in lymphocytes. Following spermidine treatment, autophagic ux, TFEB targets, and autophagy-related genes detected by scRNA-seq were highly enriched in B cells. Spermidine signi cantly increased spike-speci c IgG secretion and memory B cells, and neutralizing antibody activity against SARS-CoV-2 strains, in vaccine non-responders that also presented with high immune cell senescence. Targeting immune senescence using spermidine may offer a practical approach to improve immune responses in vaccine non-responders, as a post- or pre-vaccination intervention. Additionally, it highlights the utility of immune senescence markers as predictive biomarkers for identifying vaccine non-responders, addressing a key challenge in vaccine development for an aging population
Serially quantifying TERT rearrangement breakpoints in circulating tumor DNA enables minimal residual disease monitoring in patients with neuroblastoma
Get PDFTelomerase is reactivated by genomic TERT rearrangements in ~30% of diagnosed high-risk neuroblastomas. Dismal patient prognosis results if the RAS/MAPK/ALK signaling transduction network also harbors mutations. We present a liquid biopsy-based monitoring strategy for this particularly vulnerable pediatric patient subgroup, for whom real-time molecular diagnostic tools are limited to date. Droplet digital PCR assays quantifying patient-individualized TERT rearrangement breakpoint copies, ALK copy numbers and allelic ALK p.R1275Q mutation frequencies were applied to longitudinally collected liquid biopsies (peripheral blood and bone marrow plasma, n=44 biosamples), the mononuclear cell fraction from bone marrow and matched tumor samples. Marker detection was compared with current gold standard diagnostics. Reanalysis of whole-genome and targeted panel sequencing data from 169 patients identified 64 TERT-rearranged neuroblastoma samples collected at initial and/or relapse diagnosis from 55 patients (254 total TERT rearrangement events). Detection and quantification of unique TERT rearrangement breakpoints in as little as 1ng of total cell-free DNA in peripheral blood plasma improved therapy response assessment and early relapse detection in individual patients. Proof-of-concept is provided for minimal residual disease detection in the bone marrow niche, from which relapses frequently arise, by analyzing unique TERT rearrangement breakpoints in bone marrow plasma-derived cell-free DNA. TERT rearrangement breakpoints, as a single marker or combined with mutations in the RAS/MAPK/ALK signaling transduction network, can serve as robust and highly sensitive biomarkers for disease activity and spatially and temporally resolve disease better than current gold standard diagnostics in individual patients with TERT-driven neuroblastoma
Ultrasound pupillometry for the detection of a relative afferent pupillary defect (RAPD): systematic evaluation in patients with optic neuritis and comparison with infrared video pupillometry
Get PDFPURPOSE: A relative afferent pupillary defect (RAPD) is a characteristic clinical sign of optic neuritis (ON). Here, we systematically evaluated ultrasound pupillometry (UP) for the detection of an RAPD in patients with ON, including a comparison with infrared video pupillometry (IVP), the gold standard for objective pupillometry. MATERIALS AND METHODS: We enrolled 40 patients with acute (n = 9) or past (n = 31) ON (ON+), 31 patients with multiple sclerosis (MS) without prior ON, and 50 healthy controls (HC) in a cross-sectional observational study. Examinations comprised the swinging flashlight test, B-mode UP, IVP, autorefraction to assess the best-corrected visual acuity, optical coherence tomography to determine peripapillary retinal nerve fiber layer thickness, and the 51-item National Eye Institute-Visual Function Questionnaire to determine the vision-related quality of life. RESULTS: While UP and IVP measurements of pupil diameter (PD) at rest correlated in ON+ eyes (n = 52, r = 0.56, 95% CI: 0.35; 0.72) and in HC eyes (n = 100, r = 0.60, 95% CI: 0.47; 0.72), PD at rest was smaller in UP than in IVP measurements (difference, mean (SD) ON+ eyes: 0.44 (0.87) mm, HC eyes: 0.69 (0.80) mm). RAPD assessment by UP sharply discriminated acute ON eyes (n = 9) and HC eyes (n = 100, AUC = 1, 95%CI: 1; 1). UP detected an RAPD in 5/31 (16%) patients with MS without prior ON who had not exhibited an RAPD during the swinging flashlight test. In ON+ eyes (n = 52), UP showed stronger correlations with visual acuity (r = 0.66, 95% CI: 0.50; 0.78) and vision-related quality of life (r = 0.47, 95% CI: 0.24; 0.66) than IVP (r = 0.52, 95% CI: 0.36; 0.67 and r = 0.27, 95% CI: 0.03; 0.51). CONCLUSIONS: B-mode UP allows for objective detection and quantification of an RAPD with performance characteristics similar to IVP. RAPD assessment by UP may detect subclinical optic nerve damage in patients with MS. We propose a standardized protocol for RAPD detection by UP that can be used in routine clinical evaluation of patients with ON or other optic neuropathies
Visualization of gap junction-mediated astrocyte coupling in acute mouse brain slices
Get PDFGap junctions are transmembrane protein channels that enable the exchange of small molecules such as ions, second messengers, and metabolites between adjacent cells. Gap junctions are found in various mammalian organs, including skin, endothelium, liver, pancreas, muscle, and central nervous system (CNS). In the CNS, they mediate coupling between neural cells including glial cells, and the resulting panglial networks are vital for brain homeostasis. Tracers of sufficiently small molecular mass can diffuse across gap junctions, and are used to visualize the extent of cell-to-cell coupling in situ by delivering them to a single cell through sharp electrodes or patch clamp micropipettes. Here, we describe a protocol for pre-labeling and identification of astrocytes in acute mouse forebrain slices using Sulforhodamine 101 (SR101). Fluorescent cells can then be targeted for whole-cell patch clamp, which allows for further confirmation of astroglial identity by assessing their electrophysiological properties, as well as for passive dialysis with a tracer such as biocytin. Slices can then be subjected to chemical fixation and immunostaining to detect dye-coupled networks. This protocol provides a method for the identification of astrocytes in live tissue though SR101 labeling. Alternatively, transgenic reporter mice can also be used to identify astrocytes. While we illustrate the use of this protocol for the study of glial networks in the mouse brain, the general principles are applicable to other species, tissues and cell types
Sleep and circadian rhythms in delayed sleep-wake phase disorder: phenotypic differences between patients with and without comorbid depression
No full textDelayed sleep–wake phase disorder involves chronic difficulty going to bed and waking up at conventional times and often co-occurs with depression. This study compared sleep and circadian rhythms between patients with delayed sleep–wake phase disorder with depression (DSWPD-D) and without (DSWPD-ND) comorbid depression. Clinical records of 162 patients with delayed sleep–wake phase disorder (70 DSWPD-D, 92 DSWPD-ND) were analysed, including a subset of 76 patients with circadian phase determined by the dim light melatonin onset. Variables assessed included sleep behaviour on work and free days, weekly sleep duration, social jet lag, chronotype, and phase relationships between dim light melatonin onset and sleep/wake times. Mean (SD) or median [Q1–Q3] values were compared using t-tests or Mann–Whitney. Patients with DSWPD-D showed longer sleep on workdays (DSWPD-D = 7.63 hr [1.70] versus DSWPD-ND = 6.20 hr [1.59]; p < 0.001), but not on free days. DSWPD-D also showed later sleep onset (DSWPD-D = 03:30 14;hours [02:49 hours–04:23 hours], DSWPD-ND = 02:53 hours [02:00 hours–03:41 hours]; p = 0.02) and wake times (DSWPD-D = 11:30 hours [09:30 hours–13:00 hours], DSWPD-ND = 08:45 hours [07:20 hours–11:00 hours]; p < 0.01) on workdays. Furthermore, DSWPD-D showed less social jet lag (DSWPD-D = 0.38 [0.00–1.75] versus DSWPD-ND = 2.17 [1.25–3.03]; p < 0.01), and reported higher anxiety symptoms (DSWPD-D = 71.4% versus DSWPD-ND = 45.8%; p = 0.03) and medication use (DSWPD-D = 75.0% versus DSWPD-ND = 43.8%; p = 0.01). DSWPD-D also showed wider dim light melatonin onset phase relationships with dim light melatonin onset-mid-sleep (DSWPD-D = −5.77 [1.32] versus DSWPD-ND = −4.86 [1.53]; p = 0.01) and dim light melatonin onset-waketime (DSWPD-D = −9.46 [1.82]; DSWPD-ND = −8.13 [2.08]; p = 0.01). Multivariable Poisson regression, adjusted for age and sex, showed more medication use, less social jet lag, and longer weekly sleep duration as significantly associated with DSWPD-D. These findings suggest potential biopsychosocial protective factors linked to depression in delayed sleep–wake phase disorder. Further research is required to confirm these phenotypic differences and their relevance to delayed sleep–wake phase disorder aetiology and treatment
Oncofetal reprogramming drives phenotypic plasticity in WNT-dependent colorectal cancer
Get PDFTargeting cancer stem cells (CSCs) is crucial for effective cancer treatment, yet resistance mechanisms to LGR5(+) CSC depletion in WNT-driven colorectal cancer (CRC) remain elusive. In the present study, we revealed that mutant intestinal stem cells (SCs) depart from their canonical identity, traversing a dynamic phenotypic spectrum. This enhanced plasticity is initiated by oncofetal (OnF) reprogramming, driven by YAP and AP-1, with subsequent AP-1 hyperactivation promoting lineage infidelity. The retinoid X receptor serves as a gatekeeper of OnF reprogramming and its deregulation after adenomatous polyposis coli (APC) loss of function establishes an OnF ‘memory’ sustained by YAP and AP-1. Notably, the clinical significance of OnF and LGR5(+) states in isolation is constrained by their functional redundancy. Although the canonical LGR5(+) state is sensitive to the FOLFIRI regimen, an active OnF program correlates with resistance, supporting its role in driving drug-tolerant states. Targeting this program in combination with the current standard of care is pivotal for achieving effective and durable CRC treatment
Clonal driver neoantigen loss under EGFR TKI and immune selection pressures
Get PDFNeoantigen vaccines are under investigation for various cancers, including epidermal growth factor receptor (EGFR)-driven lung cancers. We tracked the phylogenetic history of an EGFR mutant lung cancer treated with erlotinib, osimertinib, radiotherapy and a personalized neopeptide vaccine (NPV) targeting ten somatic mutations, including EGFR exon 19 deletion (ex19del). The ex19del mutation was clonal, but is likely to have appeared after a whole-genome doubling (WGD) event. Following osimertinib and NPV treatment, loss of the ex19del mutation was identified in a progressing small-cell-transformed liver metastasis. Circulating tumour DNA analyses tracking 467 somatic variants revealed the presence of this EGFR wild-type clone before vaccination and its expansion during osimertinib/NPV therapy. Despite systemic T cell reactivity to the vaccine-targeted ex19del neoantigen, the NPV failed to halt disease progression. The liver metastasis lost vaccine-targeted neoantigens through chromosomal instability and exhibited a hostile microenvironment, characterized by limited immune infiltration, low CXCL9 and elevated M2 macrophage levels. Neoantigens arising post-WGD were more likely to be absent in the progressing liver metastasis than those occurring pre-WGD, suggesting that prioritizing pre-WGD neoantigens may improve vaccine design. Data from the TRACERx 421 cohort provide evidence that pre-WGD mutations better represent clonal variants, and owing to their presence at multiple copy numbers, are less likely to be lost in metastatic transition. These data highlight the power of phylogenetic disease tracking and functional T cell profiling to understand mechanisms of immune escape during combination therapies
Determinants of ascending aortic morphology: cross-sectional deep learning-based analysis on 25 073 non-contrast-enhanced NAKO MRI studies
No full textAIMS: Understanding determinants of thoracic aortic morphology is crucial for precise diagnostics and therapeutic approaches. This study aimed to automatically characterize ascending aortic morphology based on 3D non-contrast-enhanced magnetic resonance angiography (NC-MRA) data from the epidemiological cross-sectional German National Cohort (NAKO) and to investigate possible determinants of mid-ascending aortic diameter (mid-AAoD). METHODS AND RESULTS: Deep learning (DL) automatically segmented the thoracic aorta and ascending aortic length, volume, and diameter was extracted from 25,073 NC-MRAs. Statistical analyses investigated relationships between mid-AAoD and demographic factors, hypertension, diabetes, alcohol, and tobacco consumption. Males exhibited significantly larger mid-AAoD than females (M:35.5±4.8mm, F:33.3±4.5mm). Age and body surface area (BSA) were positively correlated with mid-AAoD (age: male: r(2)=0.20, p<0.001, female: r(2)=0.16, p<0.001; BSA: male: r(2)=0.08, p<0.001, female: r(2)=0.05, p<0.001). Hypertensive and diabetic subjects showed higher mid-AAoD (Δ(Hypertension) = 2.9 ± 0.5mm; Δ(Diabetes) = 1.5 ± 0.6mm). Hypertension was linked to higher mid-AAoD regardless of age and BSA, while diabetes and mid-AAoD were uncorrelated across age-stratified subgroups. Daily alcohol consumption (male: 37.4±5.1mm, female: 35.0±4.8mm) and smoking history exceeding 16.5 pack-years (male: 36.6±5.0mm, female: 33.9±4.3mm) exhibited highest mid-AAoD. Causal analysis (Peter-Clark algorithm) suggested that age, BSA, hypertension, and alcohol consumption are possibly causally related to mid-AAoD, while diabetes and smoking are likely spuriously correlated. CONCLUSION: This study demonstrates the potential of DL and causal analysis for understanding ascending aortic morphology. By disentangling observed correlations using causal analysis, this approach identifies possible causal determinants, such as age, BSA, hypertension, and alcohol consumption. These findings can inform targeted diagnostics and preventive strategies, supporting clinical decision-making for cardiovascular health