Bacteroides thetaiotaomicron (B. theta) dominates the gut microbiome of most mammals. This strictly anaerobic gut symbiont colonizes the mucus layer of host intestinal epithelial cells in both healthy and diseased conditions. Reduced neuronal and vagal afferent innervation observed in germ-free mice was found to be normalized by colonization with B. theta. In addition to deficits in gut innervation, germ-free mice have been reported to have reduced neuronal number and neurotransmitter levels in the brain. Here, we investigated the hallmarks of Alzheimer’s disease (AD) in the brain of germ-free mice compared to mice mono-colonized with B. theta. We analyzed the number of mature neurons, neurotransmitter transporters, amyloid precursor protein processing, and inflammatory status in three brain regions: the hippocampus, prefrontal cortex (PFC), and cerebellum. The hippocampus and the PFC are regions thought to be highly susceptible to pathogenesis, whereas the cerebellum is thought to be only mildly affected. Interestingly, secretion of neuroprotective sAPPα decreased in hippocampus and remained unchanged in PFC, while levels were increased in the cerebellum in response to bacterial colonization. In addition, the number of presynaptic boutons increased in the hippocampus but remained unaffected in the cerebellum. IMPORTANCE The gut microbiome has been reported to not only contribute to diseases of the gastrointestinal tract but also to interfere with and potentially even initiate diseases of other organ systems, such as the brain. Interference with the gut microbiome has been shown to elicit cognitive changes, for example, in rodent models of AD. Colonization with the common gut microbe B. theta not only affected the brain per se in our study but also showed specific brain region-dependent effects related to AD. This implies that evaluating the impact the microbiome might have on brain disorders needs a much more detailed investigation in the future with spatial and also potentially time resolution.

Proteases are emerging biomarkers for diseases including cancer, cardiovascular disorders, and inflammation, making their detection valuable for early diagnosis and therapeutic monitoring. Translating biomarker monitoring from laboratories to personalized healthcare requires biosensors that are simple, selective, and compatible with point-of-care formats. To the best of our knowledge, we report the first peptide-based chronopotentiometric biosensor for protease detection. The sensor is fabricated on cost-effective disposable electrodes, has a straightforward design, and operates at ultra-low intrinsic sensing power (<1 nW), supporting future integration into compact sensing devices.The sensor enables direct, label-free detection of matrix metalloproteinase-9 (MMP-9) with picomolar detection limit through potential shifts induced by peptide hydrolysis. Careful peptide design provides high selectivity against other MMP family members, and the device successfully detects proteases released from cells in moderately complex media. Inhibition studies confirm that the signal originates from proteolytic activity.In parallel, the biosensing platform is validated using multi-parametric surface plasmon resonance (MP-SPR), providing an independent and robust optical method to confirm peptide–MMP-9 interactions. This work demonstrates peptide-based dual-platform sensing for highly sensitive, real-time protease detection and establishes a versatile foundation for future point-of-care and wearable biosensing applications.

Bacterial signal peptides are a large group of high-affinity ligands for formyl peptide receptors (FPRs). These receptors are highly enriched on the surface of glioblastomas and several other tumor types. In this study, we evaluated the potential of bacterial signal peptides, a yet relatively unexplored class of high affinity FPR agonist for radiopharmaceutical applications. We tested a range of bacterial signal peptides and their fluorescently labeled derivatives to identify peptide residues that allow chemical modification without drastic loss of affinity. We then developed a selective fluorescent peptide derivative with more than 1000-fold selectivity towards FPR1, which binds rapidly at low nanomolar concentrations and forms stable receptor-ligand complexes that can persist for up to 72 h. This peptide probe effectively bound to a human glioma cell line U87-MG and efficiently penetrated spheroids derived from U87-MG cells. Finally, we have designed a corresponding metal chelate-peptide conjugate with similar affinity that is taken up by FPR1-transfected HEK293T cells and naturally FPR1-expressing U87-MG cells. A pilot study with planar scintigraphy in healthy mice showed no substantial uptake or retention in healthy organs, suggesting that bacterial signal peptides could indeed be an interesting tool for the development of radiopharmaceuticals.

Changes in the composition of specific cell‐surface molecules on immune cells are important early markers of directed immune responses. Current biosensors largely focus on detecting soluble disease markers. Here, a peptide‐functionalized graphene biosensor is presented capable of platform‐independent, highly sensitive detection of specific immune receptors on living cells. Formyl peptide receptor 2 (FPR2) is targeted, a key modulator of innate immune cells, and validated the technology in two complementary formats: i) graphene‐enhanced surface plasmon resonance (SPR) as a robust optical benchmark, and ii) graphene field‐effect transistors (gFETs) as a compact, cost‐effective electrical alternative. Target specificity is first confirmed using HEK293T cells selectively overexpressing FPR2. Detection of FPR2 on primary human neutrophils is achieved with high reproducibility using fewer than 10 000 cells mL −1 , demonstrating both sensitivity and reliability. By demonstrating sensitive and reproducible detection across both optical and electrical platforms, this work bridges materials science and immunology, highlighting the potential of peptide‐functionalized graphene biosensors for point‐of‐care diagnostics, immune monitoring, and early sepsis triage.

Formyl peptide receptors (FPRs) are pattern recognition receptors well-known for bacterial pathogen sensing. We here identified activator and inhibitor motifs for FPRs that are present on surface proteins of various viral pathogens. Peptides containing these motifs interact with all FPR family members and modulate various important immune functions in innate immune cells. Viral breakdown products comprising these motifs were found in patients with COVID-19. In the spike protein, many activators are found in highly mutagenic regions, whereas the inhibitor motif is located in a conserved domain that also exists in further unrelated viruses. The physiochemical properties of FPR1 activators correlate with the occurrence of protein aggregation hotspots. Such hotspots are present on various surface proteins of unrelated viruses that can also activate FPRs. This points toward a general contribution of FPRs in modulating antiviral immune responses during many distinct viral infections.

The detection of cells and viruses is essential for research and clinical applications, creating a demand for high-performance biosensors. Surface plasmon resonance (SPR) enables label-free, real-time detection and is highly promising for healthcare, including point-of-care diagnostics. However, its performance is often limited in complex biological systems. Integrating two-dimensional (2D) materials such as graphene into SPR sensors has been proposed as a strategy to improve sensitivity, but experimental evidence remains scarce. Here, we investigate the influence of graphene on SPR biosensors using several relevant biological examples, including antibody-virus and peptide-cell interactions. Compared to gold sensors, graphene integration produced reproducible signal enhancement of up to 600%, far exceeding previous reports.  Importantly, graphene-enhanced SPR enabled discrimination between different cell types, a capability not observed with gold alone. Our findings demonstrate that graphene provides substantially greater enhancement than predicted and can be applied across diverse biological systems. This establishes graphene-enhanced SPR as a powerful platform for advancing biosensor performance, with broad potential in biomedical research, diagnostics, and gene therapy.

Matrix metalloproteinase-9 (MMP-9) is a key biomarker targeted in biosensing applications due to its involvement not only in maintaining good health but also in triggering various diseases such as cancer. While quantitative detection of MMP-9 is widely performed using bioanalytical detection kits such as enzyme-linked immunosorbent assay (ELISA), faster, label-free and real-time monitoring of MMP-9 activity would lead to improved disease diagnosis with better understanding of its role in underlying disease progression and development of therapeutic strategies. In this work, multi-parametric surface plasmon resonance spectroscopy (MP-SPR) is used to develop a highly sensitive MMP-9 sensor using immobilized synthetic peptides as MMP-9 substrates. Upon binding to MMP-9, the MMP-9 specific peptide is hydrolyzed between two sites of the amino acid sequence (P1 Gly and P1' Met), resulting in a decrease in the SPR signal response. The sensor detects different concentrations of MMP-9 in buffer and cell culture medium (RPMI-1640), indicating that it can be used under physiological conditions. The limit of detection (LOD) for MMP-9 in buffer is 0.34 pM and the linear detection range is between 5 pM and 9 nM, covering the clinically relevant detection range of MMP-9. To our knowledge, this is the first short synthetic peptide-based MP-SPR biosensor for monitoring MMP-9 activity. The sensor is faster than ELISA (minutes vs. hours) and provides real-time detection with access to binding kinetics information. The use of MP-SPR provides information on surface coverage and peptide thickness before and after cleavage, which is unique compared to other detection methods.

Proteases have been proposed as potential biomarkers for several pathological conditions including cancers, multiple sclerosis and cardiovascular diseases, due to their ability to break down the components of extracellular matrix and basement membrane. The development of protease biosensors opened up the possibility to investigate the proteolytic activity of dysregulated proteases with higher efficiency over the traditional detection assays due to their quick detection capability, high sensitivity and selectivity, simple instrumentation and cost-effective fabrication processes. In contrast to the recently published review papers that primarily focused on one specific class of proteases or one specific detection method, this review article presents different optical and electrochemical detection methods that can be used to design biosensors for all major protease families. The benefits and drawbacks of various transducer techniques integrated into protease biosensing platforms are analyzed and compared. The main focus is on activity-based biosensors that use peptides as biorecognition elements. The effects of nanomaterials on biosensor performance are also discussed. This review should help readers to select the biosensor that best fits their needs, and contribute to the further development of this research field. Protease biosensors may allow better comprehension of protease overexperession and potentially enable novel devices for point-of-care testing.

>Formation and deposition of amyloid-beta(Aβ) are considered one of the main drivers of Alzheimer's disease(AD). For more than 30 years, Aβ has challenged researchers through its complex physicochemical properties and multiple peptide processing steps that involve several proteases(Andreasson et al., 2007),

Recent advances have placed the pro-inflammatory activity of amyloid β (Aβ) on microglia cells as the focus of research on Alzheimer's Disease (AD). Researchers are confronted with an astonishing spectrum of over 100 different Aβ variants with variable length and chemical modifications. With the exception of Aβ1-42 and Aβ1-40, the biological significance of most peptides for AD is as yet insufficiently understood. We therefore aim to provide a comprehensive overview of the contributions of these neglected Aβ variants to microglia activation. First, the impact of Aβ receptors, signaling cascades, scavenger mechanisms, and genetic variations on the physiological responses towards various Aβ species is described. Furthermore, we discuss the importance of different types of amyloid precursor protein processing for the generation of these Aβ variants in microglia, astrocytes, oligodendrocytes, and neurons, and highlight how alterations in secondary structures and oligomerization affect Aβ neurotoxicity. In sum, the data indicate that gene polymorphisms in Aβ-driven signaling pathways in combination with the production and activity of different Aβ variants might be crucial factors for the initiation and progression of different forms of AD. A deeper assessment of their interplay with glial cells may pave the way towards novel therapeutic strategies for individualized medicine.

Formyl peptide receptors (FPRs) may contribute to inflammation in Alzheimer's disease through interactions with neuropathological Amyloid beta (Aβ) peptides. Previous studies reported activation of FPR2 by Aβ1-42, but further investigation of other FPRs and Aβ variants is needed. This study provides a comprehensive overview of the interactions of mouse and human FPRs with different physiologically relevant Aβ-peptides using transiently transfected cells in combination with calcium imaging. We observed that, in addition to hFPR2, all other hFPRs also responded to Aβ1-42, Aβ1-40, and the naturally occurring variants Aβ11-40 and Aβ17-40. Notably, Aβ11-40 and Aβ17-40 are very potent activators of mouse and human FPR1, acting at nanomolar concentrations. Buffer composition and aggregation state are extremely crucial factors that critically affect the interaction of Aβ with different FPR subtypes. To investigate the physiological relevance of these findings, we examined the effects of Aβ11-40 and Aβ17-40 on the human glial cell line U87. Both peptides induced a strong calcium flux at concentrations that are very similar to those obtained in experiments for hFPR1 in HEK cells. Further immunocytochemistry, qPCR, and pharmacological experiments verified that these responses were primarily mediated through hFPR1. Chemotaxis experiments revealed that Aβ11-40 but not Aβ17-40 evoked cell migration, which argues for a functional selectivity of different Aβ peptides. Together, these findings provide the first evidence that not only hFPR2 but also hFPR1 and hFPR3 may contribute to neuroinflammation in Alzheimer's disease through an interaction with different Aβ variants.

Abstract Inflammation is a central element of many neurodegenerative diseases. Formyl peptide receptors (FPRs) can trigger several receptor-dependent signal transduction pathways that play a key role in neuroinflammation and neurodegeneration. They are chemotactic receptors that help to regulate pro- and anti-inflammatory responses in most mammals. FPRs are primarily expressed in the immune and nervous systems where they interact with a complex pattern of pathogen-derived and host-endogenous molecules. Mounting evidence points towards a contribution of FPRs – via neuropathological ligands such as Amyloid beta, and neuroprotective ligands such as Humanin, Lipoxin A4, and Annexin A1 – to multiple pathological aspects of neurodegenerative diseases. In this review, we aim to summarize the interplay of FPRs with neuropathological and neuroprotective ligands. Next, we depict their capability to trigger a number of ligand-dependent cell signaling pathways and their potential to interact with additional intracellular cofactors. Moreover, we highlight first studies, demonstrating that a pharmacological inhibition of FPRs helps to ameliorate neuroinflammation, which may pave the way towards novel therapeutic strategies.

Mucociliary clearance through coordinated ciliary beating is a major innate defense removing pathogens from the lower airways, but the pathogen sensing and downstream signaling mechanisms remain unclear. We identified virulence-associated formylated bacterial peptides that potently stimulated ciliary-driven transport in the mouse trachea. This innate response was independent of formyl peptide and taste receptors but depended on key taste transduction genes. Tracheal cholinergic chemosensory cells expressed these genes, and genetic ablation of these cells abrogated peptide-driven stimulation of mucociliary clearance. Trpm5-deficient mice were more susceptible to infection with a natural pathogen, and formylated bacterial peptides were detected in patients with chronic obstructive pulmonary disease. Optogenetics and peptide stimulation revealed that ciliary beating was driven by paracrine cholinergic signaling from chemosensory to ciliated cells operating through muscarinic M3 receptors independently of nerves. We provide a cellular and molecular framework that defines how tracheal chemosensory cells integrate chemosensation with innate defense.