​Unidad Funcional de Investigación de Enfermedades Crónicas

This work introduces a novel dual-component electrode modifier comprising gold nanoparticles (AuNPs) and a hydrated form of the poly(3,4-ethylenedioxythiophene) (hPEDOT) conducting polymer to enhance biomolecule immobilization and signal amplification in electrochemical immunosensing. The thiophene–AuNP interaction promotes electropolymerization, forming a citrate-doped matrix with carboxyl groups that enable covalent, oriented antibody anchoring. Moreover, the hydration of the polymeric scaffold enhances the biocompatibility and anti-fouling properties of the sensing interface. This multifunctional polymer–hydrogel nanocomposite was used to develop a disposable sandwich-type electrochemical immunoplatform for the determination of human endostatin (HE), a biomarker of colorectal cancer (CRC). The immunoplatform achieved a detection limit of 0.04 ng mL⁻ 1 and distinguished not only healthy individuals from early-stage CRC patients but also from those with premalignant colorectal lesions. These findings underscore the potential of this new multifunctional hybrid nanocomposite to improve the performance of electrochemical immunosensing, which is crucial for timely cancer diagnosis and intervention. • Engineered multifunctional hybrid polymer-hydrogel nanocomposite as electrode modifier. • Disposable sandwich-format electrochemical immunoplatform for human endostatin determination. • Remarkable sensitivity (LOD of 0.04 ng mL −1 ), anti-fouling properties, and stability. • Early and minimally invasive colorectal cancer diagnosis through plasma analysis.

Abstract BACKGROUND Malignant gliomas (MGs), including glioblastomas, are among the most aggressive cancers, characterized by rapid progression and resistance to standard therapies such as temozolomide (TMZ) and radiotherapy. Standard treatments, such as the Stupp protocol, offer limited survival benefits and high toxicity. This study aims to optimize chemoradiotherapy (CRT) protocols using a validated mathematical model and in silico trials to improve patient outcomes. Finally, our model can be adapted to other cancers with similar resistance mechanisms, suggesting broader utility. MATERIAL AND METHODS We developed a mathematical model using ordinary differential equations to simulate MG dynamics and CRT response. The model integrates key biological processes, such as cell proliferation, tumor dormancy, and resistance mechanisms. Key features include Ki-67, the role of persister cells in the development of TMZ resistance and the impact of treatment timing on survival. We calibrated and validated the model using experimental data from a mouse subventricular zone (SVZ) model developed through retroviral expression of EGFR-wt in primary cultures of neural stem cells from mice. For calibration, we administered 3 daily sessions of 3 Gy each, beginning a week post-tumor injection, and monitored tumor progression via bioluminescence using the In Vivo Imaging System (IVIS), which tracks tumor cell viability. In addition, survival data from prior TMZ experiments were utilized. To validate the model, we compared in vivo and in silico OS from CRT treatments. Finally, we translated the model and the optimal treatment into humans. RESULTS Our in silico simulations revealed that concomitant CRT protocols, incorporating treatment breaks, resulted in a 17% improvement in median overall survival (OS) in mice compared to the standard Stupp protocol, with reduced toxicity. Metronomic CRT protocols also showed promising results. Notably, spaced TMZ monotherapy achieved survival outcomes comparable to CRT, underscoring its efficacy in this context. When translated to human scenarios, extended treatment protocols with concomitant CRT or concomitant CRT plus adjuvant TMZ separated by treatment breaks resulted in a median OS of up to 48 months, a 280% increase over the standard Stupp protocol. These results underscore the potential of optimized protocols to enhance OS and reduce side effects. CONCLUSION This study highlights the power of mathematical modeling and in silico trials in optimizing oncology, providing valuable insights into the design of more effective CRT regimens for MGs. Our findings indicate that extended treatment schedules with treatment breaks could significantly improve OS and quality of life. Clinical trials are required to confirm our findings in real-world settings.

Biomarkers known as neo‐antigens, such as collagen, are increasingly considered of interest for the diagnosis and therapeutic monitoring of oncology patients. This work reports the development of an attractive strategy for the amperometric immunodetection of Collagen type I α 1 (COL1A1), involving magnetic microbeads for the construction of the immunoplatform, a sandwich assay format using specific capture and biotinylated‐detector antibodies, horseradish peroxidase labeling, and amperometric transduction at disposable screen‐printed carbon electrodes. The developed methodology allowed the selective, rapid, simple, and reliable detection of COL1A1 in the 0.051–10 ng mL −1 linear range with a detection limit of 15 pg mL −1 . The developed immunoplatform was successfully applied to the analysis of COL1A1 in plasma and tissue extracts from healthy individuals and colorectal cancer patients. The results obtained proved the potential of this neo‐antigen as biomarker for diagnosis of this disease.

Background/Objectives: Lactoferrin, a protein involved in the immune response, plays a significant role in the etiopathogenesis of periodontitis in the general population. This cross-sectional study aims to determine the salivary concentration of lactoferrin in Down syndrome individuals with periodontitis. Methods: A convenience cohort of 76 Down syndrome individuals was established, including 34 with periodontitis, 29 with gingivitis, and 13 with healthy gums. Unstimulated saliva samples were collected and processed to quantify the lactoferrin concentration using the Human Lactoferrin ELISA kit, the total protein concentration (bicinchoninic acid assay [BCA]) using the BCA Protein Assay Kit (Pierce, Rockford, IL, USA), and the lactoferrin/BCA ratio. Additionally, the Lf rs1126478 (140A/G in exon 2, Lys/Arg) genotypes were determined via PCR with restriction fragment length polymorphism (RFLP) analysis using the Earl enzyme. Results: The lactoferrin levels were comparable across patients with periodontitis, gingivitis, and healthy gums (median = 8.20, 6.57, and 7.61 µg/mL, respectively). There were no differences in the BCA levels between the three groups (median = 2.21, 3.17, and 2.08 µg/µL, respectively) nor in the lactoferrin/BCA ratios. The distribution of the Lf 140A/G polymorphism did not show differences concerning periodontal health status. Conclusions: In Down syndrome individuals, salivary lactoferrin and BCA levels are not influenced by the periodontal health condition. Additionally, no significant genetic associations were found with the rs1126478 polymorphism in Down syndrome individuals with and without periodontitis. Lactoferrin production in Down syndrome may not be upregulated in response to periodontal pathogens, which could be indicative of an immune system dysregulation contributing to the early onset and severity of periodontitis in this population.

A multifunctional cerium oxide nanoparticles (CeO2NPs)-based nanolabel is exploited to implement an electrochemical sandwich-type immunoplatform for the determination of T-cell immunoglobulin and mucin domain 1 (TIM-1) biomarker, a mucin-like class I membrane glycoprotein associated with cancer angiogenesis. The immunoplatform is constructed using screen-printed electrodes where capture antibody is immobilized through the chemistry of diazonium salts. CeO2NPs exhibit robust pseudo-peroxidase activity even at high substrate concentrations. They are covalently functionalized in a simple manner after carboxylation with a detector antibody (dAb), acting dually as a nanozyme and nanocarrier for sensing bioreceptors. This allows the development of immunoplatforms with improved robustness and performance (in terms of a moderate enhancement in sensitivity, a significant expansion in the linear range, and a reduction in the background current) compared with the immunoplatforms prepared using nanolabels also decorated with the natural enzyme (horseradish peroxidase, HRP) or the conventional enzymatic labeling involving the dAb and an HRP-secondary antibody. Under the optimized experimental conditions, the developed electrochemical immunoplatform allows the highly sensitive detection of the TIM-1 glycoprotein, with a detection limit of 9.9 pg mL−1 and a linear working range of 33–600 pg mL−1. This performance permits biomarker quantification within clinically relevant ranges. This innovative configuration enables the precise diagnosis and stratification of colorectal cancer patients by analyzing plasma samples without pretreatment beyond a sample dilution and allows establishment of the first cut-off values reported for this purpose.Graphical abstractSupplementary InformationThe online version contains supplementary material available at 10.1007/s00604-025-07021-3.

The Charcot-Marie-Tooth-1E (CMT1E) disease is typically described as a peripheral neuropathy in humans, causing decreased nerve conduction, spastic paralysis, and tremor. The Trembler-J (TrJ) mice serve as a high fidelity model of this disease. Here, we use functional ultrasound (fUS) and functional connectivity (FC) to analyze TrJ mice’s brain activity during sensory stimulation and resting state experiments against wild type (WT) mice - the healthy counterpart. fUS is an imaging technique that measures cerebral blood volume (CBV) temporal changes. We study these changes in the primary somatosensory cortex barrel field (S1BF) of both mice populations during periodic vibrissae stimulation, measuring the number of pixels that correlate to the stimulation (i.e., the size of the activation area), the average correlation of these pixels (i.e., the response strength), and the CBV’s rate of change for each stimulation (i.e., the hemodynamic response). Then, we construct a FC matrix for each genotype and experiment by correlating the CBV signals from the eight cortical regions defined by the Paxinos and Franklin atlas. Our results show that TrJ mice have significantly diminished neurovascular responses and altered brain connectivity with respect to WT mice, pointing to central nervous system effects that could shift our understanding of the CMT1E disease.

SummaryThe impact of mechanical cues on cell behavior is increasingly being recognized, rendering hydrogel platforms that mimic the extracellular matrix indispensable inin vitrocell biology research. Here, we present a step-by-step protocol for synthesis and rheological characterization of polyacrylamide (PAAm) hydrogels with varying stiffnesses, produced as large circular unattached gels customizable in shape and size. We outline methods for their use in cell culture and downstream applications involving secretome or cell analysis, and protein visualization by fluorescence microscopy.This protocol is based on the recent work of Shi & Janmey who describe a novel and straightforward method for the production of large PAAm hydrogels for bulk cell culture and mechanobiology studies.1Their procedure results in one large gel that is not attached to a supporting surface and therefore can be transferred and/or stamped to generate PAAm gels of custom shapes and sizes. The aim of this step-by-step procedure is therefore not to improve the reported protocol, but to create a clearly outlined and repeatable protocol that enables a smooth implementation in any lab for a diverse audience. In addition, our protocol describes besides harvesting of cells also the collection of secretome for downstream biochemical analyses, as well as immunofluorescence labeling using antibodies that can readily be multiplexed for optimization of labeling conditions.Graphical abstract

Human prion diseases are rare, transmissible and often rapidly progressive dementias. The most common type, sporadic Creutzfeldt-Jakob disease (sCJD), is highly variable in clinical duration and age at onset. Genetic determinants of late onset or slower progression might suggest new targets for research and therapeutics. We assembled and array genotyped sCJD cases diagnosed in life or at autopsy. Clinical duration (median:4, interquartile range (IQR):2.5-9 (months)) was available in 3,773 and age at onset (median:67, IQR:61-73 (years)) in 3,767 cases. Phenotypes were successfully transformed to approximate normal distributions allowing genome-wide analysis without statistical inflation. 53 SNPs achieved genome-wide significance for the clinical duration phenotype; all of which were located at chromosome 20 (top SNP rs1799990, pvalue = 3.45x10-36, beta = 0.34 for an additive model; rs1799990, pvalue = 9.92x10-67, beta = 0.84 for a heterozygous model). Fine mapping, conditional and expression analysis suggests that the well-known non-synonymous variant at codon 129 is the obvious outstanding genome-wide determinant of clinical duration. Pathway analysis and suggestive loci are described. No genome-wide significant SNP determinants of age at onset were found, but the HS6ST3 gene was significant (pvalue = 1.93 x 10-6) in a gene-based test. We found no evidence of genome-wide genetic correlation between case-control (disease risk factors) and case-only (determinants of phenotypes) studies. Relative to other common genetic variants, PRNP codon 129 is by far the outstanding modifier of CJD survival suggesting only modest or rare variant effects at other genetic loci.

Curcumin is a polyphenol extracted from Curcuma longa’s roots. Low doses of curcumin are related to anti-inflammatory, antioxidant, and neuroprotective effects, while high doses are used for their lethality. This diversity of behaviors allows us to understand curcumin as a compound with hormetic action. Due to its strongly hydrophobic character, curcumin is often solubilized in organic compounds. In this way, we have recently reported the undesirable and occasionally irreversible effects of alcohol and DMSO on the viability of primary Schwann cell cultures. In this scenario, the use of nanoparticles as delivery systems has become a successful alternative strategy for these compounds. In the present work, we describe the structure of Polydopamine (PDA) nanoparticles, loaded with a low dose of curcumin (Curc-PDA) without the use of additional organic solvents. We analyzed the curcumin released, and we found two different forms of curcumin. Small increased cell viability and proliferation were observed in endoneurial fibroblast and Schwann cell primary cultures when Curc-PDA was steadily supplied for 5 days. The increased bioavailability of this natural compound and the impact on cells in culture not only confirm the properties of curcumin at very low doses but also provide a glimpse of a possible therapeutic alternative for PNS conditions in which SCs are involved.