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Integrated real-time imaging of executioner caspase dynamics, apoptosis-induced proliferation, and immunogenic cell death using a stable fluorescent reporter platform
Get PDFRegulated cell death plays a central role in tissue homeostasis, disease progression, and therapeutic responses. However, tools to study these processes with high spatiotemporal resolution in physiologically relevant systems remain limited. Here, we present a fluorescent reporter cell system that enables real-time visualization of caspase-3/-7 activity via a DEVD-based biosensor, alongside a constitutive fluorescent marker for assessing successful transduction and cell presence. We generated stable cell lines expressing this reporter and adapted them to both 2D and 3D culture systems, including organoids. This platform allowed dynamic tracking of apoptotic events and viability loss at single-cell resolution. Using a proliferation dye, we also detected apoptosis-induced proliferation in neighboring cells. Furthermore, the system enabled simultaneous detection of immunogenic cell death via an endpoint measurement of surface calreticulin exposure by flow cytometry, supporting its application in studying immunogenic signaling. By measuring and integrating multiple cell death readouts by live-cell imaging, our system is well-suited for high-content screening and mechanistic dissection of different modes of cell death. When combined with complementary markers of pyroptosis and necroptosis, this platform may also be extended to investigate more complex, integrated forms of cell death
Detergent-free reconstitution of transmembrane proteins in giant liposomes of complex curvature by the Synthetic Membrane Transfer
Get PDFTransmembrane proteins perform many crucial functions in the cell, including transfer of matter and information across the membrane. Giant Unilamellar Vesicles (GUVs) are the ideal tool for studying these processes; yet functional transmembrane protein reconstitution in GUVs still represents a bottleneck in bottom-up synthetic biology. Here, we developed a novel approach that we call Synthetic Membrane Transfer (SMT), where transmembrane proteins reconstituted into styrene-maleic acid particles in a detergent-free environment can be transferred to the GUV membrane. The SMT approach is a one-step, facile and general method that works for structurally diverse proteins, and it does not require any specific lipid or buffer composition. Moreover, the SMT is fully compatible with the Synthetic Membrane Shaper (SMS) technology, which allows to deform GUVs in a controlled fashion, obtaining dumbbell-shaped GUVs exhibiting a catenoid-like geometry. The combination of SMT and SMS results in functional reconstitution of transmembrane proteins in catenoid membrane necks. Using this approach, we demonstrate that Mic10, a component of the MICOS complex, directly senses membrane curvature and localizes at the neck of dumbbells GUVs, a geometry that recapitulates the shape of mitochondria cristae junctions. This paves the way for bottom-up reconstitution of the MICOS complex on a physiological membrane geometry
MYC deregulation sensitizes cancer cells to N-myristoyltransferase inhibition
Get PDFHuman N-myristoyltransferases (NMTs) catalyze N-terminal protein N-myristoylation and are promising targets in cancer, with an emerging mechanistic rationale for targeted therapy. Here, we screened 245 cancer cell lines against IMP-1320, a potent NMT inhibitor (NMTi), and conducted pathway-level analyses to identify that deregulated MYC increases cancer cell sensitivity to NMTis. Proteomics on detergent-enriched membrane fractions in MYC or MYCN-deregulated cancer cell models revealed that cell death is associated at least in part with loss of membrane association of mitochondrial respiratory complex I. This is concurrent with loss of myristoylation and degradation of the complex I assembly factor NDUFAF4, and induction of mitochondrial dysfunction, driven by MYC or MYCN-deregulation. NMTis eliminated or suppressed MYC- and MYCN-driven tumors in vivo without overt toxicity, suggesting that this constitutive co-translational protein modification can be targeted in MYC-driven cancers
Fluorescence intensity data from HEK293 Ca(2+) spike trains
No full textThis dataset contains fluorescence intensity time series extracted from regions of interest (ROIs) of time-lapse microscopy recordings of Ca²⁺ signalling in HEK293 cells.
The data was extracted using Time Measure tool in NIS element (version 5.21.03). Acquisition intervals (ND.T) of 1 s - 4 s.
Stimulation per experiment:
CChCB_2024_03_07_E1.txt / CChCB_2024_03_12_E1.txt / CChCB_2024_03_14_E2.txt / CChCB_2024_03_19_E1.txt
t = 600 s - 5400 s , CCh 5 µM
t = 5400 s - 8400 s , CCh 10 µM
t = 8400 s - end , CCh 15 µM
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CChCB_2024_03_28_E1.txt
t = 600 s - 1200 s , CCh 5 µM
t = 1200 s - end , CCh 30 µM
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CChCB_2024_04_03_E1.txt / CChCB_2024_04_03_E2.txt / CChCB_2024_04_03_E3.txt
t = 300 s - 1020 s , CCh 5 µM
t = 1020 s - end , CCh 30 µM
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CChCB_2024_06_05_E1.txt
t = 600 s - end , CCh 30 µM
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CChCB_2024_07_11_E1.txt / CChCB_2024_07_18_E1.txt / CChCB_2024_08_08_E1.txt / CChCB_2024_08_08_E2.txt / CChCB_2024_08_09_E1.txt / CChCB_2024_08_09_E2.txt / CChCB_2024_08_09_E3.txt
t = 900 s - end , CCh 15 µM
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CChCB_2024_08_21_E1.txt / CChCB_2024_08_22_E1.txt / CChCB_2024_08_22_E2.txt / CChCB_2024_08_23_E1.txt / CChCB_2024_08_23_E2.txt
t = 900 s - end , CCh 3 µM
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CChCB_2024_09_17_E1.txt / CChCB_2024_09_24_E1.txt
t = 600 s - end , CCh 5 µ
Serum metabolic profiling in rheumatic heart disease and degenerative aortic stenosis
Get PDFMetabolomics enables the characterisation of pathogenic and diagnostic biomarkers in cardiovascular disease (CVD). Given the high prevalence of rheumatic heart disease (RHD) and degenerative aortic stenosis (AS) in Africa, we investigated potential discriminant metabolic biomarkers in individuals with severe RHD and AS undergoing valve replacement and compared them to those in matched controls. Untargeted metabolomics of serum samples showed that seven metabolites that were differentially expressed in RHD patients were independent of the patients’ baseline characteristics (covariates) and could differentiate RHD patients from healthy controls (AUC > 0.7). Four metabolites could differentiate AS patients from controls (AUC ≥ 0.7). Of the perturbed metabolites in RHD and AS, 7-HOCA and deoxycholate showed a moderate association with left ventricular ejection fraction. Furthermore, acylcarnitine and ketone body levels were correlated with the left ventricular mass index in RHD and left atrial area in AS patients. Elevated levels of cortisol were associated with the presence of valve calcification in RHD and AS patients. This is a pilot study on rarely studied CVD in sub-Saharan Africa. This study suggested that the metabolites altered in RHD and degenerative AS are not only associated with cardiac remodelling but also involved in major energetic pathways, amino acid metabolism, and inflammation regulation processes
A multicenter study on the comparability of myocardial strain values acquired with different CMR scanners and analyzed with different post-processing software: insights into the "Traveling Volunteers" study
Get PDFPURPOSE: Strain quantifies myocardial deformation. Despite its high diagnostic value, strain analyses using cardiovascular magnetic resonance (CMR) feature tracking (FT) have not been fully implemented into clinical routine due to lack of information on reproducibility. The purpose of this study was to assess the comparability of cardiovascular magnetic resonance CMR FT strain and ejection fraction (EF) measurements, obtained from different MR scanners and analyzed using different software platforms. METHODS: CMR examinations were performed in 15 healthy volunteers using three different scanners (German Heart Center of the Charité, Charité Campus Berlin Buch, and Theresien-Hospital Mannheim). FT was performed using Medis Suite and Circle CVI. Inter-software/scanner agreement was determined using Bland-Altman plots, Wilcoxon test, and paired Student's t test. Intra-/inter-observer reproducibility was evaluated using intraclass correlation coefficients. RESULTS: Left ventricular (LV) global longitudinal (GLS) and circumferential (GCS) strain did not differ between the three centers (small bias of - 1.27 to 1.32% for GLS and 0.91 to 0.69% for GCS). Inter-scanner agreement was lower regarding LV global radial strain (GRS) (bias of - 2.29 to 4.53%) and good for LV EF (bias of - 0.59 to 0.94%). Inter-software agreement was low for GLS (bias of - 5.72 to - 4.59%), GCS (- 1.13 to - 1.55%), and GRS (18.34 to 19.83%), with higher GLS/GCS and lower GRS values in CVI than Medis. LV EF showed better inter-software agreement (biases of - 0.07 to 0.06%). Intra- and inter-observer reproducibility was good for strain measurements across all scanners (bias of - 0.01 to 2.05 and 0.22 to 1.92, respectively) and software packages (ICC 0.70 to 0.90 and 0.51 to 0.89, respectively). CONCLUSION: Inter-scanner reproducibility for CMR FT measurements was high for GLS and GCS, suggesting potential use in routine CMR examinations. However, strain values between the two software vendors (CVI and Medis) were significantly different, indicating the need for standardization and implementation of software-specific cutoff values
Guillain-Barré syndrome and related neuropathies post-bariatric surgery: a systematic review of clinical cases and outcomes
No full textBariatric surgery is a widely used treatment for morbid obesity, but it can lead to neurological complications, including Guillain–Barré Syndrome (GBS), a rare immune-mediated polyneuropathy. The mechanisms underlying post-bariatric GBS remain unclear, and no comprehensive review has evaluated its incidence, risk factors, and outcomes. This systematic review synthesizes available clinical data to assess the relationship between bariatric surgery and GBS, focusing on its presentation, diagnosis, and management. A systematic literature search was conducted in Web of Science, PubMed, and Scopus following PRISMA guidelines. Studies reporting GBS following bariatric surgery were included. Data extraction focused on demographics, clinical presentation, laboratory findings, treatment approaches, and outcomes. Study quality was assessed using the Joanna Briggs Institute (JBI) tool. GBS cases were predominantly reported in female patients, with symptom onset occurring within weeks to months post-surgery. The most common clinical features included ascending motor weakness, sensory disturbances, hyporeflexia, and autonomic dysfunction. Laboratory findings showed elevated cerebrospinal fluid protein and abnormal nerve conduction studies. Patients were treated with intravenous immunoglobulin (IVIG), plasmapheresis, and nutritional supplementation, particularly thiamine and vitamin B12. Recovery varied; some patients regained full function, while others experienced persistent neurological deficits or severe complications such as respiratory failure. GBS is a rare but serious post-operative complication of bariatric surgery, potentially linked to immune dysregulation and nutritional deficiencies. Early recognition and prompt intervention are critical. Further research is needed to clarify underlying mechanisms, optimize treatment, and improve preventive strategies
TFAP2A orchestrates gene regulatory networks and tubular architecture in kidney outer medullary collecting ducts
Get PDFMutations in the transcription factor TFAP2A are linked to congenital anomalies of the kidney and urinary tract in humans. While Tfap2a knockout (KO) in mouse collecting ducts leads to tubular epithelial abnormalities, its precise molecular functions in kidney tubules remain unclear. To investigate Tfap2a-dependent gene regulatory networks in the mouse kidney collecting ducts, we employed conditional knockout (Hoxb7-Cre; Tfap2aflox/flox) models combined with transcriptomics. Histomorphological and physiological assessments of Tfap2a knockout mice revealed progressive postnatal dilation of the outer medullary collecting ducts. Integrating bulk and single-nucleus RNA sequencing with in silico motif mapping in ATAC-seq datasets demonstrated that Tfap2a is highly expressed and active in normal collecting duct principal cells. Comparative transcriptomics between 3-month-old Tfap2a KO and control mice identified dysregulated genes associated with cell adhesion and WNT signaling, including Alcam and Wnt9b. These alterations were confirmed by in situ hybridization. Our findings reveal that Tfap2a regulates medullary collecting duct diameter by orchestrating a transcriptional network involving Wnt9b and Alcam, providing new insights into its role in kidney structural integrity
Future treatments for myelin oligodendrocyte glycoprotein antibody-associated disease: the clinical trial landscape
No full textINTRODUCTION: Myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) is an emerging autoimmune demyelinating disorder distinct from multiple sclerosis and AQP4-IgG-positive neuromyelitis optica. Despite increasing recognition, no therapies are currently approved for MOGAD, and treatment remains empirical, with significant variability in clinical response and access to care. AREAS COVERED: This review explores the evolving treatment landscape of adult MOGAD, with a focus on immunotherapies under active clinical investigation: azathioprine, tocilizumab, satralizumab, and rozanolixizumab. For each agent, we discuss mechanisms of action, pharmacokinetics, dosing, safety, and efficacy based on clinical trials and observational data. Literature was identified through PubMed and ClinicalTrials.gov, including ongoing phase 2/3 studies (MOGwAI, TOMATO, METEOROID, and cosMOG). EXPERT OPINION: Targeted immunotherapies have the potential to transform MOGAD management. In the next five years, one or more of these agents may achieve regulatory approval, particularly if biomarker-driven strategies and trial designs are refined. Addressing unmet needs in pediatric populations and low-resource settings will be essential to ensure equitable, personalized treatment