Nonspecific Activity Research Articles

An attenuated live strain of HY9901 mutant Δgr provides protection against Vibrio alginolyticus in pearl gentian grouper (♀Epinephelus fuscoguttatus × ♂Epinephelus lanceolatu).

Vibrio alginolyticus is a serious aquaculture bacterial pathogen, which is widely distributed in the ocean and rivers, and cause vibriosis in aquaculture. Therefore, it is imperative to develop effective vaccine to prevent vibriosis. In this study, the efficacy of gr deletion strain (Δgr) of V. alginolyticus as an attenuated live vaccine was evaluated in peal gentian grouper by intraperitoneal injection immunization, followed by a wild strain injection challenge. Real-time fluorescence quantitative PCR was used for analyzing the expression dynamics of immune-related genes. ELISA was used to assess the serum antibody titer and non-specific enzyme activities were determined by kit. The results showed that the safe dose of Δgr was 1.0 × 106 CFU/mL for pearl gentian grouper, and the immune protection rate of Δgr was 73.08 %, providing significant protection against V. alginolyticus. The expression levels of all the immune-related genes MHC-Iα, TNF-α, IL-1β, IL-16 and MyD88 were upregulated in liver, spleen, head kidney and thymus within 42 days post-vaccination. In addition, specific antibody IgM, activities of catalase, superoxide dismutase and lysozyme were also significantly increased compared with the control group. It is not only safe for groupers, but also can effectively stimulate both specific and non-specific immunity of fish. These results indicated that Δgr could be used as an effective attenuated live vaccine candidate to prevent pearl gentian grouper against V. alginolyticus in aquaculture.

A Bifunctional Lysosome-Targeting Chimera Nanoplatform for Tumor-Selective Protein Degradation and Enhanced Cancer Immunotherapy.

Lysosome-targeting chimeras (LYTACs) have recently emerged as a promising therapeutic strategy for degrading extracellular and membrane-associated pathogenic proteins by hijacking lysosome-targeting receptors. However, the antitumor performance of LYTAC is limited by its insufficient tumor accumulation and nonspecific activation. Additionally, the synergistic effects of LYTACs and other therapeutic modalities are crucial. To address these issues, a bifunctional LYTAC nanoplatform (NLTC) is developed for tumor-selective protein degradation and enhanced cancer immunotherapy. By rationally controlling the surface composition, the NLTC can effectively transport extracellular or membrane proteins into lysosomes for degradation via cation-independent mannose 6-phosphate receptors. With removable surface modification, an NLTC is obtained that efficiently accumulated in tumor tissues and avoided on-target off-tumor toxicity. Moreover, the synthesis method of NLTC is generally applicable to various enzymes. Thus, catalase (CAT) is encapsulated with NLTC to synergistically degrade cancer cell surface programmed death ligand-1 (PD-L1), relieve the immunosuppressive tumor microenvironment for effective cancer immunotherapy, and significantly inhibit tumor growth, recurrence, and metastasis in B16F10-bearing mice. This work presents a bifunctional LYTAC nanoplatform that can not only perform tissue-selective protein degradation but also integrate other therapeutic modalities, providing insights into the design of advanced LYTAC technologies for clinical applications.

Spatioselective Imaging of Noncoding RNAs in Mitochondria via an Organelle-Specific DNA Assembly Strategy.

Precise imaging of noncoding RNAs (ncRNAs) in specific organelles allows decoding of their functions at subcellular level but lacks advanced tools. Here we present a DNA-based nanobiotechnology for spatially selective imaging of ncRNA (e.g., microRNA (miRNA)) in mitochondria via an organelle-specific DNA assembly strategy. The target miRNA-initiated assembly of DNA hairpins is inhibited by the block of toehold-mediated strand displacement reaction but can be exclusively activated by a mitochondria-encoded ribosomal RNA (rRNA) for hybridization chain reaction, enabling spatial control over miRNA imaging. We demonstrate that the conditionally controlled DNA assembly technology allows for minimization of nonspecific activation and thus improves the spatial precision of miRNA detection. In addition, the strategy is adaptable to visualizing other ncRNAs such as long noncoding RNAs in mitochondria, highlighting the universality of the approach. Overall, this work provides a useful tool for spatially selective imaging of ncRNAs and investigating the functions of organelle-located RNA.

Epigenetic Threads of Neurodegeneration: TFAM's Intricate Role in Mitochondrial Transcription.

There is a myriad of activities that involve mitochondria that are crucial for maintaining cellular equilibrium and genetic stability. In the pathophysiology of neurodegenerative illnesses, mitochondrial transcription influences mitochondrial equilibrium, which in turn affects their biogenesis and integrity. Among the crucial proteins for keeping the genome in optimal repair is mitochondrial transcription factor A, more commonly termed TFAM. TFAM's non-specific DNA binding activity demonstrates its involvement in the control of mitochondrial DNA (mtDNA) transcription. The role of TFAM in controlling packing, stability, and replication when assessing the quantity of the mitochondrial genome is well recognised. Despite mounting evidence linking lower mtDNA copy numbers to various age-related diseases, the correlation between TFAM abundance and neurodegenerative disease remains insufficient. This review delves into the link between neurodegeneration and mitochondrial dysfunction caused by oxidative stress. Additionally, the article will go into detail about how TFAM controls mitochondrial transcription, which is responsible for encoding key components of the oxidative phosphorylation (OXPHOS) system.

AlphaFold 3 sheds insights into chemical enhancer-induced structural changes in Cas12a RNPs

BackgroundThe CRISPR/Cas12a system has revolutionized nucleic acid detection through trigger-activated nonspecific trans-cleavage activity. Enhancing this activity is vital for improving the sensitivity of CRISPR/Cas biosensing systems. Although various chemical enhancers have been shown to affect Cas12a ribonucleoprotein (RNP), the underlying mechanisms remain poorly understood. Investigating how these enhancers alter the structure of Cas12a RNPs is essential for elucidating the enhancement mechanisms involved.ResultsThis study focuses on elucidating the structural changes in Cas12a RNPs induced by various chemical enhancers via AlphaFold 3, an emerging and powerful tool for analyzing structural changes in protein‒nucleic acid complexes. We validated the ability of AlphaFold 3 to simulate structural changes in Cas12a RNPs activated by triggers and subsequently analyzed the effects of specific enhancers, such as reducing agents (e.g., Dithiothreitol, namely DTT), divalent cations (e.g., Mg2+, Mn2+), and bovine serum albumin (BSA). Our findings revealed that DTT, simulated with a cysteine-to-serine Cas12a mutant, caused significant structural changes in Cas12a RNP, as evidenced by notable shifts in the distance between key residues (Val377 to Gln1136) and a high root mean square deviation (RMSD)(RMSD > 2). Conversely, divalent cations and BSA did not cause substantial structural changes, resulting in only minor shifts in residue distance and a low RMSD (RMSD < 2).ConclusionsThese results demonstrate that DTT enhances Cas12a activity by inducing significant structural rearrangements, whereas divalent cations and BSA-induced enhancements do not involve substantial structural modifications.Graphical Summary of structural changes from inactivated Cas12a RNP to activated Cas12a RNP and from activated Cas12a RNP to activated Cas12a RNP with an enhancer. The units of both the RMSD and distance are Å.

Effects of immersion bathing in Lactobacillus plantarum CLY-05 on the growth performance, non-specific immune enzyme activities and gut microbiota of Apostichopus japonicus.

In order to study the optimal use of Lactobacillus plantarum in sea cucumber (Apostichopus japonicus), 49 days feeding trial was conducted to determine the influence of immersion bathing in different concentrations of Lactobacillus plantarum CLY-05 on body weight gain rate and non-specific immune activities. The potential effect of CLY-05 on gut microbiota was also analyzed during the immersion bathing at the optimum concentration. The results showed that the body weight growth rate of all bathing groups was higher than that of control. The highest specific growth rate (4.58%) and weight gain rate (25.35%) was achieved at the bacterial concentration of 1×103 CFU/mL. The activities of non-specific immune enzymes (ACP, AKP, SOD and LZM) of all bathing groups increased after immersion bathing, and the enzyme activities of groups bathed with the bacterium at 1×103 and 1×104 CFU/mL reached the highest. Therefore, 1×103 CFU/mL was considered as the optimum concentration of L. plantarum CLY-05 for A. japonicus pond culture. The results of gut microbiota analysis showed that the gut microbiota changed with the addition of L. plantarum CLY-05, and the richness and diversity of the gut microbiota peaked on day 14 and day 21, respectively. The correlation analysis revealed that the non-specific immune enzyme activities were significantly correlated to some gut bacteria (in the phyla Proteobacteria, Firmicutes) after immersion bathing in L. plantarum CLY-05. These findings provide the theoretical foundation for probiotic application in sea cucumber farming.

Effects of Saprolegnia parasitica on pathological damage and metabolism of Epithelioma papulosum cyprini cell.

Saprolegniasis is a common fungal disease in aquaculture. It will form white flocculent hyphae on the skin of fish, and the hyphae may grow inward and penetrate into muscle tissue, which will reduce the immunity of the body and eventually lead to death. However, there are still some gaps in the mechanism of the fish body surface against the invasion of Saprolegnia. This study explored the defense mechanism of Epithelioma papulosum cyprini cell (EPC) in the process of Saprolegnia parasitica infection from the perspective of pathogenic bacteria and host cells, so as to provide a theoretical basis for further exploring the mechanism of host resistance to S. parasitica invasion. The EPC cell was used as the research object. The EPC cells were treated with 1×106 CFU/mL of S. parasitica for 0, 6, 12, 24, 48 and 72 h. Cell viability and cell membrane damage were detected, and the non-specific immune enzyme activity in the cells was detected. Based on the above research, the apoptosis genes and antioxidant genes in the cells were detected to analyze the effect of S. parasitica on the metabolism of the EPC cells. The results showed that with the prolongation of the co-culture time of S. parasitica and cells, the cell viability gradually decreased and the cell membrane integrity was destroyed, but at the same time, the activity of non-specific immune enzymes increased to resist the infection of S. parasitica. In addition, the detection of EPC apoptosis gene casp3a and CTSD showed that the relative content of casp3a gene increased significantly at 24 h and reached the maximum value of the culture time (P<0.05). The content of CTSD gene increased significantly at 12 h and reached the maximum value (P<0.05). The results of antioxidant immune genes serpinh1a and gpx1a were opposite to the structure of apoptotic genes. The content of serpinh1a and gpx1a genes decreased significantly at 12 h (P<0.05), but with the prolongation of culture time, the content increased significantly at 24 h and 48 h (P<0.05). After stimulation of EPC cells by S. parasitica, the differential metabolites were mainly concentrated in Lipids, Compounds with biological roles and Phytochemical compounds. The KEGG pathway mainly focused on ABC transporters, Glycerophospholipid metabolism, Cysteine and methionine metabolism, Glycine, serine and threonine metabolism, Purine metabolism. In general, S. parasitica can affect cell activity, destroy the cell membrane of EPC cells, and cause apoptosis. However, EPC cells can also resist the invasion of S. parasitica by regulating their own non-specific immunity and their own metabolites, thereby protecting the body from the infection of S. parasitica.

Application and Prospective of CRISPR-Cas in Cancer Diagnosis and Therapy

Cancer, a leading cause of mortality worldwide, poses a significant challenge to public health. The advent of CRISPR-Cas systems has introduced a novel and promising approach to overcoming the limitations of conventional cancer therapies. This review delves into the burgeoning role of CRISPR-Cas in the oncology field, highlighting its utility in both cancer diagnosis and therapy. In the diagnostic realm, CRISPR-based technologies, such as DETECTR and SHERLOCK, leverage the non-specific collateral cleavage activity of Cas12 and Cas13 proteins for highly sensitive detection of nucleic acid sequences, offering rapid and sensitive diagnostic capabilities. Therapeutically, CRISPR-Cas systems inactivate oncogenes and enhance treatment outcomes through engineered variants and innovative approaches. Variants such as nCas9 and dCas9 have been engineered to enhance specificity and transcriptional regulation, respectively. Challenges including the need for high specificity to avoid off-target effects, long-term safety assessments and ethical considerations persist, yet ongoing research aims to refine these technologies for enhanced efficacy and safety. Overall, CRISPR-Cas systems herald a new era in precision medicine for cancer patients.

Molecular Imaging of Tumor-Infiltrating Lymphocytes in Living Animals Using a Novel mCD3 Fibronectin Scaffold.

The interaction between cancer cells and immune cells in the tumor microenvironment (TME) plays a crucial role in determining tumor growth, metastasis, and response to treatment. Tumor-infiltrating lymphocytes (TILs) in TME could be a predictive marker for treatment response in various therapeutic interventions, including chemotherapy and immunotherapy. Thus, imaging the tumor immune microenvironment is important for selecting the optimal treatment strategies in cancer therapy. The CD3 protein represents a promising target for diagnostic imaging of TILs in vivo to assess the immune state of the TME. Although many anti-CD3 antibodies have been explored for this application, the nonspecific immune activation by these antibodies limits their applications. To overcome this issue, we engineered a novel fibronectin III domain (FN3) protein binder (mCD3-FN3;11.8 kDa) against mouse CD3 antigen protein using a yeast display library to image TILs homing in vivo into the TME. We performed in vitro and in vivo assays to test the mCD3-FN3 binder purity as well as in vivo targetability in mouse models of syngeneic tumors. We used near-infrared 800 dye conjugated with mCD3-FN3 (IR800-mCD3-FN3) for in vivo tracking of TILs via optical imaging. We used three different syngeneic tumors in mice (mCD3+ EL4 tumor in C57BL/6 mice, mCD3- CT26 colon tumor, and mCD3- 4T1 breast tumor in BALB/c mice) for imaging TILs in vivo. C57BL/6 mice bearing EL4 tumors were separated into two groups (blocking [Blk] and nonblocking [Nblk]; n = 3 per group) and used for in vivo imaging. Blocking groups received 200 μg of unlabeled mCD3-FN3 2 h prior to the administration of IR800-mCD3-FN3 binder. Each mouse was administered with 25 μg of the IR800-mCD3-FN3 binder and tracked using an IVIS optical imaging system over time. C57BL/6/EL4 mice were imaged at 4 and 24 h post injection of the IR800-mCD3-FN3 binder, and mouse organs were collected at 24 h after final imaging and used for ex vivo histological imaging. In CT26 and 4T1 tumor models, TILs in TME were imaged 4, 24, and 48 h after binder injection. The NIR imaging of EL4 tumors showed that IR800-mCD3-FN3 can detect both TILs within the tumor and the tumor cells with a high signal-to-background ratio 24 h after initial binder injection with a total radiant efficiency (mean TRE ± SD) of 6.5 × 1010 ± 1.5 × 1010 [photons/s]/[μW/cm2]. The animals received preinjection of unlabeled mCD3-FN3(Blk) prior to IR800-mCD3-FN3 binder administration and showed a significant level of fluorescence signal reduction (mean TRE ± SD: 1.6 × 1010 ± 4.1 × 109) in the tumor when compared to the EL4-Nblk tumors (p = 0.006). The mouse group with CT26 and 4T1 tumors where the probe can only bind to TILs within the tumor showed a specific imaging signal (mean TRE ± SD) of 1.1 × 1011 ± 5.2 × 1010 and 9.5 × 1010 ± 4.6 × 1010, respectively, at 48 h p.i. For these groups, the ex vivo tumor-to-muscle ratios were 20- and 27-fold for CT26 and 4T1 tumors, respectively. These results clearly demonstrate the in vivo binding ability of the mCD3-FN3 binder to mCD3 marker expressed by T cells in the TME. The ex vivo histological analysis of tumors, and the organs of animals with EL4 tumors, and TILs imaging of CT26, and 4T1 tumors (at 48 p.i.) confirmed that the IR800-mCD3-FN3 probe was able to specifically bind to CD3 markers expressed by the T cells. In summary, both in vitro and in vivo data indicated that the engineered mCD3-FN3 binder by this study is a promising ligand for diagnostic imaging of tumors in vivo for the assessment of mCD3 expressing TILs in the TME. This can be used as a prognostic marker in evaluating tumor response to therapeutic intervention as well as a diagnostic marker in imaging tumor response to immune checkpoint blockade cancer therapies.

Pt(IV) Prodrug Photoactivation: A Promising Strategy for Cancer Therapy.

Platinum (II) drugs, including cisplatin, carboplatin, and oxaliplatin, have achieved significant clinical success in cancer treatment. However, their clinical application has been greatly hindered by various adverse factors, such as non-specific activation and drug resistance. Compared with Pt(II) drugs, the axial ligands within Pt(IV) compounds can improve the pharmacokinetic properties, selectivity, and biological activity, implementing alternative cytotoxic mechanisms beyond DNA cross-linking and partially overcoming drug resistance. The controlled conversion of Pt(IV) prodrugs into Pt(II) agents at the tumor site has been extensively explored internationally. In this review, Pt(IV) prodrug modification strategies are first summarized, and the development of the predominant external and internal photosensitizers is listed. Finally, three representative photoreduction mechanisms and strategies for developing corresponding Pt(IV) prodrugs are discussed. This work provides constructive instruction for the subsequent molecular design of Pt(IV) prodrugs.

Dual Role of TRPV1 Channels in Cerebral Stroke: An Exploration from a Mechanistic and Therapeutic Perspective.

Transient receptor potential vanilloid subfamily member 1 (TRPV1) has been strongly implicated in the pathophysiology of cerebral stroke. However, the exact role and mechanism remain elusive. TPRV1 channels are exclusively present in the neurovascular system and involve many neuronal processes. Numerous experimental investigations have demonstrated that TRPV1 channel blockers or the lack of TRPV1 channels may prevent harmful inflammatory responses during ischemia-reperfusion injury, hence conferring neuroprotection. However, TRPV1 agonists such as capsaicin and some other non-specific TRPV1 activators may induce transient/slight degree of TRPV1 channel activation to confer neuroprotection through a variety of mechanisms, including hypothermia induction, improving vascular functions, inducing autophagy, preventing neuronal death, improving memory deficits, and inhibiting inflammation. Another factor in capsaicin-mediated neuroprotection could be the desensitization of TRPV1 channels. Based on the summarized evidence, it may be plausible to suggest that TPRV1 channels have a dual role in ischemia-reperfusion-induced cerebral injury, and thus, both agonists and antagonists may produce neuroprotection depending upon the dose and duration. The current review summarizes the dual function of TRPV1 in ischemia-reperfusion-induced cerebral injury models, explains its mechanism, and predicts the future.

Cognate antigen engagement induces HIV-1 expression in latently infected CD4+ Tcells from people on long-term antiretroviral therapy.

Despite antiretroviral therapy (ART), HIV-1 persists in latently infected CD4+ Tcells, preventing a cure. Antigens drive the proliferation of infected cells, precluding latent reservoir decay. However, the relationship between antigen recognition and HIV-1 gene expression is poorly understood because most studies of latency reversal use agents that induce non-specific global Tcell activation. Here, we isolated rare CD4+ Tcells responding to cytomegalovirus (CMV) or HIV-1 Gag antigens from people living with HIV-1 on long-term ART and assessed Tcell activation and HIV-1 RNA expression upon coculture with autologous dendritic cells (DCs) presenting cognate antigens. Presentation of cognate antigens exvivo induced broad Tcell activation (median 42-fold increase in CD154+CD69+ cells) and significantly increased HIV-1 transcription (median 4-fold), mostly through the induction of rare cells with higher viral expression. Thus, despite low proviral inducibility, antigen recognition can promote HIV-1 expression, potentially contributing to spontaneous reservoir activity and viral rebound upon ART interruption.

Mechanisms governing bystander activation of T cells.

The immune system is endowed with the capacity to distinguish between self and non-self, so-called immune tolerance or "consciousness of the immune system." This type of awareness is designed to achieve host protection by eliminating cells expressing a wide range of non-self antigens including microbial-derived peptides. Such a successful immune response is associated with the secretion of a whole spectrum of soluble mediators, e.g., cytokines and chemokines, which not only contribute to the clearance of infected host cells but also activate T cells that are not specific to the original cognate antigen. This kind of non-specific T-cell activation is called "bystander activation." Although it is well-established that this phenomenon is cytokine-dependent, there is evidence in the literature showing the involvement of peptide/MHC recognition depending on the type of T-cell subset (naive vs. memory). Here, we will summarize our current understanding of the mechanism(s) of bystander T-cell activation as well as its biological significance in a wide range of diseases including microbial infections, cancer, auto- and alloimmunity, and chronic inflammatory diseases such as atherosclerosis.

Supplementation of Yupingfeng polysaccharides in low fishmeal diets enhances intestinal health through influencing the intestinal barrier, immunity, and microflora in Macrobrachium rosenbergii.

This study aimed to investigate the effects of a low-fishmeal diet (LF, substituting soybean meal for 40% fish meal) and the supplementation of 500 mg/kg and 1000 mg/kg Yu Ping Feng (YPF) polysaccharides on the growth performance, antioxidant enzyme activities, intestinal ultrastructure, non-specific immunity, and microbiota of Macrobrachium rosenbergii. The study involved the administration of different diets to M. rosenbergii, including a control diet, a low-fishmeal diet (LF), and LF diets supplemented with 500 mg/kg and 1000 mg/kg YPF polysaccharides. Growth performance, antioxidant enzyme activities, intestinal ultrastructure, non-specific immunity, and microbiota were assessed. The LF diet significantly reduced growth performance parameters compared to the control group. However, YPF supplementation notably improved these parameters, with the greatest improvement observed at a 1000 mg/kg dosage. Antioxidant enzyme activities (SOD, GSH-PX) were diminished in the LF group, accompanied by elevated MDA levels, whereas YPF supplementation restored these activities and reduced MDA levels. Ultrastructural analysis revealed that the LF diet caused intestinal villi detachment and peritrophic matrix (PM) shedding, which were alleviated by YPF. Gene expression related to PM formation (GS, CHS, EcPT) was downregulated in the LF group but significantly upregulated in the 1000P group. Non-specific immune gene expressions (IMD, Relish, IκBα) and enzyme activities (NO, iNOS) were suppressed in the LF group but enhanced by YPF supplementation. Microbial community analysis showed reduced diversity and altered composition in the LF group, with increased Proteobacteria and decreased Firmicutes, which were partially restored by YPF. Correlation analysis revealed that Lactobacillus and Chitinibacter play pivotal roles in regulating intestinal health. Lactobacillus exhibited a positive relationship with the intestinal PM and immune-related indicators, whereas Chitinibacter was negatively associated with these factors. These results highlight the adverse impacts of a low-fishmeal diet on the intestinal health of M. rosenbergii and demonstrate the beneficial effects of YPF polysaccharides in alleviating these negative consequences through various mechanisms, including improved growth performance, enhanced antioxidant enzyme activities, restored intestinal ultrastructure, and modulated immune responses. The findings suggest that YPF supplementation could be a valuable strategy for mitigating the negative effects of low-fishmeal diets in aaquaculture.

Innervation density governs crosstalk of GPCR-based norepinephrine and dopamine sensors.

GPCR-based fluorescent sensors are widely used to correlate neuromodulatory signaling with brain function. While experiments in transfected cells often reveal selectivity for individual neurotransmitters, sensor specificity in the brain frequently remains uncertain. Pursuing experiments in brain slices and in vivo, we find that norepinephrine and dopamine cross-activate the respective sensors. Non-specific activation occurred when innervation of the cross-reacting transmitter was high, and silencing of specific innervation was indispensable for interpreting sensor fluorescence.

A Single-Tube, Single-Enzyme Clustered Regularly Interspaced Short Palindromic Repeats System (UNISON) with Internal Controls for Accurate Nucleic Acid Detection.

Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins have been widely applied in molecular diagnostics. Unlike the Ct value quantification method of PCR, the CRISPR system mainly relies on the rise of the rate of the fluorescence signal to indicate the concentration of the target nucleic acid, which is susceptible to system errors caused by various factors, such as reaction conditions and instrument performance. Therefore, establishing internal controls is essential to improve the accuracy, reliability, and commercial feasibility of the CRISPR system. However, the nonspecific trans-cleavage activity of Cas proteins presents a challenge in establishing internal controls. In this study, we developed unified nucleic acid detection with a single-tube, one-enzyme system (UNISON) for accurate nucleic acid detection with internal controls. By extending the crRNA and modifying it with different fluorophores and quenchers, we achieved that the specific target can only specifically cleave the corresponding folded crRNA and generate a corresponding fluorescence signal. With this design, we established an internal control, achieving accurate and reliable detection of clinical samples of the hepatitis B virus. Integrating internal controls into the CRISPR/Cas system demonstrates significant potential in medical diagnostics and virus monitoring.

A Tandem-Locked Fluorescent Probe Activated by Hypoxia and a Viscous Environment for Precise Intraoperative Imaging of Tumor and Instant Assessment of Ferroptosis-Mediated Therapy.

Noninvasive fluorescence detection of tumor-associated biomarker dynamics provides immediate insights into tumor biology, which are essential for assessing the efficacy of therapeutic interventions, adapting treatment strategies, and achieving personalized diagnosis and therapy evaluation. However, due to the absence of a single biomarker that effectively reflects tumor development and progression, the currently available optical diagnostic agents that rely on "always-on" or single pathological activation frequently show nonspecific fluorescence responses and limited tumor accumulation, which inevitably compromises the accuracy and reliability of tumor imaging. Herein, based on intramolecular charge transfer (ICT) and twisted intramolecular charge-transfer (TICT) hybrid mechanisms, we report a tandem-locked probe, NTVI-Biotin, for simultaneously specific imaging-guided tumor resection and ferroptosis-mediated tumor ablation evaluation under the coactivation of nitro reductase (NTR)/viscosity. The dual-stimulus-responsive design strategy ensures that NTVI-Biotin exclusively activates near-infrared (NIR) fluorescence signals upon interaction with both NTR and elevated viscosity levels through triggering ICT on while inhibiting the TICT process. Meanwhile, functionalization with a tumor-targeting hydrophilic biotin-poly(ethylene glycol) moiety enhances tumor accumulation. The probe's dual-response and tumor-targeting design minimizes nonspecific tissue activation, allowing for precise tumor identification and lesion removal with a superior tumor-to-normal tissue (T/N > 6) ratio. More importantly, NTVI-Biotin was capable of evaluating ferroptosis-mediated chemotherapeutics by real-time monitoring of the alternations of NTR/viscosity levels. The results reveal that the increased tumor signals of NTVI-Biotin following the combination of ferroptosis and chemotherapy correlate well with the tumor growth inhibition, demonstrating the potential of NTVI-Biotin to assess therapeutic efficacy.

Tunable multi-enzyme activities of platinum nanoclusters for enhanced specificity and sensitivity in biosensing

Nanozymes have gained prominence for their utility in biosensing and disease diagnostics. However, challenges arise from complex sample matrices and nonspecific enzyme activities that contribute to false signals. This study introduces multifunctional platinum nanoclusters (Pt NCs) exhibiting peroxidase-like (POD-like), oxidase-like (OXD-like), and laccase-like activities tailored for enhanced biosensing capabilities. By adjusting pH, we optimized the conditions to achieve distinct POD-like and OXD-like responses, thereby reducing background signals and improving detection accuracy. The addition of ATP further amplified the POD-like activity while minimizing interference from OXD-like activity. This combined strategy substantially enhanced biomarker detection selectivity, demonstrated through glucose detection in human serum samples. Moreover, thiol inhibition of laccase-like activity in Pt NCs was leveraged for thiol-based antioxidant assessment, revealing their application in quantifying total antioxidant capacity (TAC) in human liver cancer cells, accounting for 44 % of TAC. The Pt NCs demonstrated robust sensitivity and reusability, offering a novel multi-enzyme nanomaterial with potential for precise and interference-free biosensing applications. These findings contribute to the development of advanced nanozyme-based biosensors, addressing specificity regulation challenges and expanding their practical application in biosensing and disease diagnostics.

Real-life observation of wildfire-smoke impaired COVID-19 vaccine immunity.

Wildfires are increasingly common with wildfire smoke affecting millions globally, yet its impact on immune responses is poorly understood. Natural Killer (NK) cells play a role in mediating air pollutant effects and regulating vaccine immunity. This real-world study, conducted on participants in the Pfizer BNT162b2 COVID-19 vaccine trial, studied the effects of wildfire smoke exposure on long-term vaccine effects. We collected blood samples from 52 healthy, non-smoking participants (ages 26-83) before and 1 month after placebo or vaccine injections during heavy wildfire smoke events in Sacramento. The study included 28 vaccinated (Group 1) and 24 placebo-injected (Group 2) individuals, the latter vaccinated several months later, outside wildfire season. Blood samples from both Group 1 and 2 were also investigated 6 months after the second dose of vaccine. We analyzed intracellular cytokines, B and NK cell markers by flow cytometry, and serum immunoglobulin levels against common coronaviruses using multiplex assays. A robust S-RBD-specific IgG response observed 1 month post booster, declined variably 6 months later. Wildfire smoke acutely increased IL-13 expression by CD56bright NK cells. IL-13+CD56bright NK cells at the time of vaccination negatively correlated with anti-S-RBD IgG (r=-0.41, p<0.05) one month later. Total IgG levels on the other hand, positively correlated with the air quality index (AQI) measured during vaccination (r=0.96, p<0.01). Similarly to age (but not sex, BMI or race/ethnicity), the two-week AQI averages during vaccination showed a significant negative correlation with anti-S-RBD IgG levels 6 months later (r=-0.41, p<0.05). Wildfire smoke may lead to inappropriate immunoglobulin production and diminished vaccine immunity. Our novel findings highlight a previously unrecognized pathway involving NK-cell derived IL-13 and non-specific B-cell activation and underscore the significance of environmental exposures in shaping immunity.

Characteristics of the proinflammatory response of monocytes/macrophages in patients with metabolic syndrome, exemplified by the production of IL-6 and MCP-1

Metabolic syndrome (MS) is one of the most common socially significant diseases. Around 1.9 billion people suffer from this disease, which places an enormous burden on healthcare systems around the world. This is particularly true in connection with concomitant diseases. With the progression of MS, disorders in the function of the immune system occur in the body, including those associated with mitochondrial dysfunction, leading to the development of chronic inflammation. In particular, there is an increase in the number of circulating monocytes actively recruited to inflamed adipose tissue, where there is non-specific proinflammatory activation of innate immune cells, which adopt an M1-like phenotype and become less sensitive to anti-inflammatory stimuli. This ultimately leads to a decrease in the functional activity of monocytes/macrophages and their immunoplasticity. The subject of the study was the venous blood of patients and the CD14+ monocytes/macrophages obtained from it by immunomagnetic separation. In our work, we focused on the search for significant relationships between markers of chronic inflammation and the formation of immune tolerance of monocytes/macrophages in patients with metabolic syndrome. Biochemical parameters, basal and LPS-stimulated production of proinflammatory cytokines (IL-6 and MCP-1) were investigated by culture of monocytes/macrophages in patients with metabolic syndrome. The evaluation of biochemical parameters in blood samples from MS patients and healthy donors revealed that the levels of ALAT, AST, GGT, alkaline phosphatase, uric acid, C-reactive protein, glucose and insulin were significantly higher in MS patients than in healthy donors. The levels of P-amylase and high-density lipoprotein were significantly lower than in the control group. Within the experimental model, repeated stimulation showed a decrease in cytokine production in response to LPS compared to the first stimulation on day 7. It was also found that the response to the primary stimulus was higher in cells from patients with a body mass index (BMI) 40 kg/m2, which could indirectly indicate the presence of a phenotype associated with chronic inflammation and consequently with reduced plasticity of the monocyte/macrophage immune response.