Achieving negative surgical margins remains a critical determinant of local recurrence and survival in head and neck cancer (HNC) surgery. Current intraoperative margin assessment techniques, including frozen section analysis, suffer from sampling errors and procedural delays. Tumor-targeted fluorescence imaging offers real-time tumor visualization but lacks standardized quantitative approaches for clinical decision-making. We developed a Tumor Probability Mapping (TPM) framework using panitumumab-IRDye800 fluorescence imaging in 16 HNC patients. Ex vivo specimens and gross tissue sections were imaged using near-infrared fluorescence systems. A total of 5,442 regions of interest (ROIs) were manually distributed across fluorescence images of gross specimen sections validated by histopathology. Signal-to-background ratios (SBR) were calculated and used to train the following predictive models: generalized linear model fit standard logistic regression (MATLAB, glmfit), standard logistic regression (R, LOG), mixed-effects logistic regression (GLMER), and Bayesian mixed-effects regression (BRMS). Model performance was evaluated using receiver operating characteristic and area under the curve (ROC-AUC) analysis, sensitivity, specificity, along with beta-calibration and model fit. All models demonstrated excellent (> 90%) discriminative ability between tumor and normal tissue. The glmfit model, selected for clinical implementation, achieved 95.8% accuracy, 90.8% sensitivity, 98.8% specificity, and an AUC of 0.989 on test data. The final TPM algorithm provides real-time probability assessment of tumor presence based on fluorescence intensity quantified by histopathology validated historical data. TPM represents a significant advancement in fluorescence-guided surgery by converting qualitative fluorescence signals into quantitative probability assessments validated against histopathology. This approach provides surgeons with standardized, real-time tumor probability information that extends beyond qualitative assessments and/or binary threshold determinations, potentially improving surgical outcomes by enhancing margin assessment and reducing local recurrence rates.
Monitoring therapeutic response is essential for improving the precision and efficacy of photodynamic therapy (PDT), yet quantitative and real-time evaluation remain challenging. Herein, we report a programmable DNA nanotube-based dual-mode biosensing platform for monitoring PDT-induced apoptosis through complementary fluorescence and electrochemical signals. In this system, methylene blue (MB) functions as a multifunctional component, simultaneously acting as a photosensitizer, near-infrared imaging probe, and electrochemical reporter. A caspase-3-responsive peptide is integrated into the DNA nanotube scaffold, enabling apoptosis-triggered fluorescence activation. Caspase-3 activation triggers peptide cleavage, resulting in fluorescence recovery and enhanced electrochemical signals, with limits of detection (LOD) of 0.786 ng/mL for the fluorescence method and 0.622 ng/mL for square-wave voltammetry (SWV). Gold nanoparticles incorporated into the DNA nanotube further catalyze endogenous H2O2 decomposition to generate oxygen, alleviating tumor hypoxia and enhancing PDT efficacy. The platform demonstrated accurate dual-mode detection capability in complex biological samples. These results demonstrate that the programmable DNA nanotube-based platform enables synchronized electrochemical and fluorescence biosensing through a shared caspase-3-responsive mechanism, thereby allowing real-time therapeutic feedback during photodynamic therapy.
Herein, we have successfully synthesized a colorimetric and ratiometric fluorescent probe C-BF2 for biogenic amines through a specific reaction between β-diketone boron difluoride and amine. C-BF2 presents rapid response, low limit of detection, excellent reversibility and significant color change. By incorporating C-BF2 into filter paper, we created a smart label capable of detecting gaseous biogenic amines through distinct absorption and fluorescence changes. Utilizing a smartphone and color-processing software, we established a correlation between food storage time and fluorescence color parameters (G/R) of the smart label. Importantly, the fluorescence color variations of label exhibited a linear quantitative relationship with total volatile basic nitrogen value of food, validating the accuracy and reliability in food monitoring. Furthermore, we verified the potential applications of C-BF2 as a fluorescent ink and fingerprint powder. This multipurpose detection platform is expected to provide new ideas for the field of biogenic amine detection, meat food safety monitoring, and utilization in fluorescent dyeing materials.
1,3-Diphenylguanidine (DPG), an additive widely utilized in industrial production, has been reported to exhibit numerous hazardous effects. However, effective methods for its real-time detection remain limited. In this study, an F, N-codoped carbon dots@Mn2+ (F, N-CDs@Mn2+) system is successfully developed for the rapid fluorescent detection of DPG. Initially, upon the addition of Mn²⁺, a blue shift (from 489 to 475 nm) is observed in the F, N-CDs@Mn2+ system due to chelation-enhanced fluorescence. Following the subsequent introduction of DPG, a significant fluorescence quenching accompanied by a pronounced red shift (from 475 to 501 nm) occurs in the F, N-CDs@Mn2++DPG, owing to aggregation-induced quenching and photoinduced electron transfer. This sensor exhibits a broad linear range spanning from 0 to 10.55 µg/mL, along with a low limit of detection (LOD) of 23.63 ng/mL. Moreover, the F, N-CDs@Mn2+ system is combined with RGB analysis for the real-time detection of DPG, yielding satisfactory results with an LOD of 37.12 ng/mL and recoveries ranging from 97% to 114.33%. Consequently, highly sensitive and selective detection of DPG is achieved, providing a real-time visual analytical platform for safeguarding human health and enabling early warning of environmental pollution.
Lead exposure remains a major global health concern, contributing to an estimated 1.5% of annual deaths worldwide and causing widespread, irreversible neurodevelopmental and cardiovascular damage, particularly in children. A central challenge in understanding and mitigating lead toxicity is the determination of its chemical speciation in complex biological and environmental systems, where conventional Pb LIII-edge X-ray absorption spectroscopy (XAS) is fundamentally limited by severe core-hole lifetime broadening. Here we show that Pb Lα1 high-energy-resolution fluorescence-detected X-ray absorption spectroscopy (HERFD-XAS) provides a practical and experimentally accessible route to overcoming this limitation. Pb Lα1 HERFD-XAS yields an approximately 2-fold improvement in spectral resolution relative to conventional XAS, enabling enhanced sensitivity for quantitative speciation in heterogeneous and dilute systems. The method shows negligible chemical shifts (<0.06 eV) in the emission energy, simplifying implementation, distinguishes holodirected and hemidirected Pb(II) coordination environments, resolves biologically relevant Pb binding modes in zinc-finger model peptides, and identifies hemidirected Pb-substitution in hydroxyapatite, a key long-term reservoir of lead in humans. These results establish Pb Lα1 HERFD-XAS as a sensitive and experimentally accessible tool for determining Pb speciation in environmental and biological systems.
The high-performance detection of hexavalent chromium (Cr6+, specifically Cr2O72-) poses significant challenges and holds great importance for in-situ inspection. Among the developed probes, strong oxidizing acid roots, especially MnO4- and CrO42-, there is significant interference in the detection of Cr2O72-. In this study, a dual functional group fluorescent probe (SP) was constructed for the specific recognition of Cr2O72- based on redox reaction and chelation quenching fluorescence effect (CHQF). Moreover, SP showed excellent selectivity and sensitivity for Cr2O72- detection, the LOD = 7.08 × 10-8 M and a high binding constant (Ka = 4.53 × 104 M-1). The detection mechanism observation revealed that Cr2O72- was reduced to Cr3+, and Cr3+ could coordinate with SP and accompanied by an electron transfer process. Rapid and efficient detection of Cr2O72- in actual water samples and plants used smartphones as an aid. To sum up, the detection system based on SP recognized identity of Cr2O72- within different application situations.
In this work, we present a novel strategy for developing solid-state luminescence turn-on sensors by constructing indicator displacement assays (IDAs) within a covalent organic framework (COF) composed of 2,4,6-triformylphloroglucinol (Tp) and 5, 5'-diamino-2, 2'-bipyridine (Bpy) ligands. To illustrate our strategy, a robust electron-deficient COF was synthesized through the post-synthetic N-alkylation of its bipyridyl units. The cationic N-alkylated bipyridyl units served as receptors, while the fluorescent indicator, anionic 8-anilino-1-naphthalenesulfonate (ANS-), was incorporated within the framework via anion exchange. The resulting host-guest hybrid exhibited negligible fluorescence due to the efficient electron donor-receptor charge-transfer (CT) quenching of the ANS- indicators. Upon exposure to the strong electron-donating analyte triethylamine (TEA), the competitive CT interaction between the ANS- indicators and TEA with the cationic bipyridyl receptors initiates the indicator displacement process, leading to the significant fluorescence activation of ANS-. This rapid and sensitive fluorescence turn-on response in dioxane enables efficient detection of TE{Wang, 2020 #79}A with a limit of detection (LOD) of 0.95 nM. Moreover, practical applicability was demonstrated through the fabrication of test strips exhibiting visually observable solid-state fluorescence at TEA concentrations as low as 1 μM. This work presents the first demonstration of confining IDAs within COFs for solid-state luminescence turn-on sensing of TEA. It not only establishes a general approach for developing turn-on TEA sensors but also broadens the utility of COFs as versatile platforms for advanced sensing applications.
Intracerebral hemorrhage (ICH) has high disability rates and fatality. This study aims to investigate whether fat mass and obesity-associated protein (FTO) exacerbate ICH-induced brain injury by regulating autophagy-dependent ferroptosis and to identify potential therapeutic targets. An ICH model was established in rats via autologous blood injection. FTO expression was knocked down using adeno-associated virus-delivered short hairpin RNA (shRNA). Neurological scores, brain edema, and histopathology were assessed. Autophagy, oxidative stress, and ferroptosis markers were measured by Western blot and enzyme-linked immunosorbent assay (ELISA).Immunofluorescence was performed for FTO with neuronal nuclei (NeuN), glial fibrillary acidic protein (GFAP), ionized calcium-binding adapter molecule 1 (Iba-1), LC3, and GPX4/microtubule-associated protein 2 (MAP2). FTO expression was significantly upregulated post-ICH, correlating with neurological deterioration, cerebral edema, neuronal loss, and inflammatory infiltration. Immunofluorescence showed FTO colocalized with NeuN. FTO knockdown attenuated neurological deficits, reduced cerebral edema, and suppressed neuronal loss. FTO knockdown inhibited autophagy-related protein (ATG5)/microtubule-associated protein 1 light chain 3B (LC3B)-mediated autophagy activation, thereby mitigating iron overload, lipid peroxidation, and ferroptosis markers (decreased glutathione peroxidase 4 [GPX4], elevated acyl-CoA synthetase long-chain family member 4 [ACSL4], and cyclooxygenase-2 [COX2]). FTO knockdown also reduced LC3 fluorescence and restored GPX4/MAP2 colocalization. Rescue experiments further confirmed that ATG5 overexpression reversed the neuroprotective effects of FTO knockdown. FTO aggravates ICH-induced brain injury by promoting ATG5/LC3B-mediated autophagy and subsequent ferroptosis. Targeting FTO represents a promising therapeutic strategy to mitigate secondary brain damage post-ICH.
Terconazole is an effective antifungal drug with a broad spectrum of activity against many types of fungi. In the study, a spectrofluorimetric method was developed for the quantification of terconazole. Eosin Y, an efficient fluorescence probe, was used in the developed work. The developed method is based on forming a non-fluorescent association complex between terconazole and eosin Y, which quenches eosin Y's inherent fluorescence intensity at both excitation and emission spectra. This quenching effect is highly correlated with the concentration of terconazole, with a linear range of 0.15-1.2 µg/mL. Furthermore, Stern-Volmer analysis was carried out to study the quenching fluorescence strength of eosin Y by terconazole. Moreover, the developed method exhibits an LOD of 0.041 µg/mL and a LOQ of 0.125 µg/mL. Moreover, the recommended method was verified in accordance with ICH guidelines. In addition, with respectable recovery values, the established procedures were successfully applied to the analysis of terconazole in vaginal cream dosage forms. Furthermore, the greenness of the entire proposed project was assessed using the following assessment tools: MoGAPI, Complex MoGAPI, MoGSA, BAGI, and RAPI. The green profiles provided outstanding evidence of the greenness and environmental friendliness of the proposed approach.
Therapeutic angiogenesis represents a promising strategy for recovery following myocardial infarction (MI). Yiqi Huoxue Formula (YQHX), a well-known traditional Chinese medicinal prescription, is widely utilized in clinical practice to treat myocardial ischemia and enhance cardiac performance in patients with MI. This study aims to elucidate the regulatory role of YQHX on mitochondria-associated membranes (MAMs) via the cGMP/PKG signaling pathway in promoting post-MI angiogenesis and restoring cardiac function. An in vivo MI model was established via left anterior descending (LAD) coronary artery ligation in rats, and an in vitro model was developed using hypoxia-induced injury in human umbilical vein endothelial cells (HUVECs). The effects of YQHX on cardiac function and HUVEC behaviors (proliferation, migration, and tube formation) were evaluated. Myocardial histopathology and endothelial damage were assessed using HE/Masson staining and biochemical assays. Angiogenesis in the infarct border zone was visualized by platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) immunofluorescence. Ultrastructural changes in MAMs, the endoplasmic reticulum (ER), and mitochondria were observed via transmission electron microscopy (TEM). Intracellular reactive oxygen species (ROS) and calcium (Ca 2+) levels were detected using fluorescent probes. Transcriptomic sequencing was performed to identify key therapeutic pathways, followed by validation of the cGMP/PKG axis and MAMs-associated proteins (including cGMP, PKG, IP3R2, GRP75, FUNDC1, VDAC1, CYPD, and MCU) using ELISA, Western blotting, and immunofluorescence. Echocardiography and biochemical analysis demonstrated that high-dose YQHX demonstrated comparable efficacy to the first-line clinical drug Perindopril in preserving cardiac function and mitigating myocardial injury. Immunofluorescence revealed that while the infarct border zone in the MI group exhibited disordered and dysfunctional capillary proliferation, YQHX treatment promoted the formation of organized and functional microvessels. TEM revealed significant disruption of MAMs ultrastructure post-MI. Furthermore, the colocalization of IP3R2 and VDAC1 was markedly reduced in MI tissues and hypoxic HUVECs, an effect that was reversed by YQHX. Transcriptomic analysis identified the cGMP/PKG pathway as a pivotal mechanism. Activation of this pathway by YQHX restored MAMs structural integrity and rescued impaired cytosolic Ca2+ signaling, as confirmed by TEM, Western blotting, triple-labeling immunofluorescence, and calcium assays. Consequently, YQHX-mediated MAMs repair enhanced HUVEC proliferation, migration, and angiogenic capacity. This study demonstrates that MI/hypoxia impairs endothelial function by disrupting the structural and functional integrity of MAMs. YQHX effectively preserves MAMs architecture and restores intracellular Ca2+ signaling via the cGMP/PKG signaling axis, thereby promoting post-MI angiogenesis and improving cardiac performance. These findings identify a novel therapeutic target for MI and highlight the unique advantages of traditional Chinese medicine formulas in modulating subcellular organelle interactions. However, given the inherent variability of botanical materials, these mechanistic findings are based on a single validated batch, and future multi-batch standardizations are warranted.
The pink-to-purple apatite from the Xuebaoding tungsten‑tin‑beryllium (W-Sn-Be) deposit in Sichuan, Southwest China, is of significant gemmological and geological interest owing to its distinctive coloration and typical tabular crystal habit. In this study, six apatite samples exhibiting a range of colors were systematically analyzed using a comprehensive suite of techniques, including Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS), electron paramagnetic resonance (EPR), ultraviolet-visible spectrophotometry (UV-Vis), photoluminescence (PL), and three-dimensional fluorescence spectroscopy. Our findings indicate that the pink hues of apatite are primarily determined by Mn2+ d-d transitions, whereas purple tones arise from color centers associated with F vacancies. The Mn/Fe ratio shows a positive correlation with color saturation. Trace elements such as Nd3+, Sm3+, and Y3+ collectively contribute to modulating the color saturation. XRD Rietveld refinement shows that the sample is apatite of relatively high purity, accompanied by a small amount of muscovite. Both FTIR and UV-Vis spectra jointly reveal marked anisotropy in absorption peak intensities relative to crystal orientation, which can provide a basis for crystal orientation. Fluorescence studies show that the blue-violet emission is dominated by the 5d → 4f transitions of Ce3+, the additional contributions from Dy3+. In the 620-750 nm range, Mn2+, Sm3+, and Pr3+ synergistically induce pinkish-orange fluorescence. This study elucidates the color origins and luminescent behavior of pink-to-purple apatite from the Xuebaoding deposit.
Co-infection with both hepatitis B virus (HBV) and hepatitis D virus (HDV) can aggravate the severity of the end-stage liver disease and accelerate its progression. However, no combined diagnostic method for HDV and HBV nucleic acids exists. In this study, we have developed a highly sensitive and specific dual detection method for HDV RNA and HBV DNA using the CRISPR-Cas system. We established a dual detection method combining CRISPR-Cas12a with recombinase polymerase amplification (RAA) for HBV, and CRISPR-Cas13a with RT-RAA for HDV. Validation was performed using specimens from 70 co-infected patients. RAA primers and crRNAs were designed and optimized to establish a dual fluorescence detection method (DF) and lateral flow strip-based dual detection (DL) within the same CRISPR-Cas13a/Cas12a system for HDV RNA and HBV DNA. The system demonstrated 100% specificity, and both DF and DL methods exhibited a sensitivity of 10 copies/μL for synthetic positive plasmids and samples. Peak fluorescence detection was achieved with T7 RNA polymerase, while the best detection efficiency was at ssRNA: ssDNA ratio of 1:1.5. In the validation of plasma samples from 70 co-infected clinical patients, the positive concordance rates for RT-RAA-CRISPR-Cas13a/Cas12a DF and DL were 85.7% (60/70) and 82.9% (58/70), respectively. We developed a CRISPR-Cas13a/Cas12a-based dual assay for sensitive, specific, and accurate detection of HDV RNA and HBV DNA, offering an effective tool for the early detection, treatment, and monitoring of HDV and HBV infections.
Alzheimer's disease (AD), the most prevalent form of dementia, is marked by progressive memory impairment, cognitive decline, reduced acetylcholine level, oxidative stress, amyloid-β (Aβ) aggregation, and disturbances in metal homeostasis. Our earlier work on caffeic acid-based multifunctional inhibitors prompted us to explore structure-activity relationships (SARs) to improve BChE cholinesterase inhibition while maintaining or enhancing the multifunctional activity of the previously identified lead molecule 12d. For this purpose, the glycinamide linker was rationally replaced with benzylpiperazine and glycinamide-piperazine, yielding six new series of derivatives. Comprehensive SAR analysis identified 7b as the most promising molecule, displaying markedly enhanced potency compared to caffeic acid and our earlier analog EJMC-12d. 7b exhibited approximate 3.6-fold improvement in BChE inhibition compared to EJMC-12d. While AChE inhibition of 7b was found to be comparable to EJMC-12d. The enzyme kinetic studies showed a mixed-type inhibitory mechanism for 7b at both AChE and BChE. Molecular docking studies also confirmed that 7b binds to the CAS and PAS sites of AChE. The DPPH assay on synthesized derivatives revealed strong antioxidant properties of 7b (IC₅₀ = 4.89 ± 0.72 μM) compared to EJMC-12d (IC₅₀ = 6.32 ± 0.15 μM).7b also showed efficient metal-chelating properties. 7b exhibited potent inhibitory activity against Aβ1-42 aggregation, as confirmed by the ThT fluorescence assay and validated by fluorescence microscopy. The PAMPA-BBB permeability assessment showed that 7b (Pe = 3.96 ± 0.38) can cross the blood-brain barrier. Cell viability studies with 7b in SH-SY5Y cells demonstrated cytocompatibility at higher doses. The neuroprotection studies against H₂O₂ and ROS generation confirmed that 7b exhibits strong antioxidant activity and a neuroprotective effect against oxidative stress. 7b was found to be safe in the acute toxicity studies at a higher dose of 500 mg/kg. In the in vivo Aβ-induced stereotaxic mouse model, 7b significantly restored spatial working memory. The overall findings highlighted 7b as a promising, efficacious, multifunctional candidate with significant therapeutic potential for AD. Despite this, there is scope for structural modifications in 7b to further improve the multifunctional properties. Overall, this scaffold demonstrates strong potential and can be further explored as a promising therapeutic candidate for AD management.
Understanding the rapid adjustments of plants to high-light exposure remains challenging, as multiple excitation and de-excitation pathways are simultaneously activated. In this study, we examined carotenoid pigment conversions at the second-scale in three tree species in parallel with high temporal resolution (<1 s) in vivo fluorescence and absorption spectroscopy. Our results reveal that both β-branch (violaxanthin, antheraxanthin, zeaxanthin) and α-branch (lutein, lutein epoxide) xanthophylls exhibit remarkably fast and oscillating pool dynamics within the first 20 seconds of illumination, reaching even maximal values in that timeframe. Prompt (0-20 s) conversion of the lutein is observed at the expense of both lutein epoxide and α-carotene in certain species, while accumulation of antheraxanthin and zeaxanthin is seen both prompt (0-20 s) and slower (>30 s). Interestingly, mirror trends between whole α- and β-branch carotenoids seem to indicate balancing trends, involving dynamic precursor shifts between α- and β-carotenes. Further, we observe that quick xanthophyll changes match the kinetic trends of fitted Gaussian-modeled absorbance peaks (approx. at 520, 535, 560 nm) within the early seconds. These quick changes in photon absorption are followed by slower-triggered non-photochemical de-excitation through a particular xanthophyll, seen from the dominant 535-nm peak, and likely attributed to antheraxanthin or zeaxanthin. The quick xanthophylls conversion redistributing the excessive excitation energy while quenching fluorescence (EET phase) is shown as one of the first responses to excessive light, before regulated energy dissipation as heat is initiated. These observations invite to interpret the non-steady state conditions and their parametrization more carefully, considering different photoprotective strategies across species.
The aberrant aggregation of human superoxide dismutase 1 (hSOD1) into β-sheet-rich amyloid fibrils is a crucial process in the pathogenesis of amyotrophic lateral sclerosis (ALS), enhancing motor neuron degeneration and disease progression. The P66R mutation in SOD1 destabilizes local structure and promotes β-sheet-driven fibrillation, which makes it a suitable model for exploring approaches for reducing pathogenic aggregation. Here, we evaluate silymarin, a polyphenolic compound with known antioxidant and neuroprotective properties, for its potential to inhibit P66R-hSOD1 aggregation. ThT fluorescence and transmission electron microscopy analyses demonstrate a significant decrease in amyloid fibril formation in the presence of silymarin; in addition, FTIR spectroscopy confirms the suppression of β-sheet formation. Fluorescence quenching and ANS binding assays indicate a moderate-affinity binding between silymarin and the mutant protein, along with a reduction in surface hydrophobicity. Hemolysis assays confirm its protective effect against membrane damage induced by aggregates, while molecular docking and dynamic simulations indicate that silymarin stabilizes aggregation-prone areas with hydrogen bonding and hydrophobic interactions, thereby promoting compact conformations and reducing solvent-exposed surfaces. The findings identified silymarin as an effective anti-amyloidogenic agent that reduces β-sheet accumulation and fibril formation while also decreasing cytotoxicity, highlighting its potential as a therapeutic candidate for ALS.
Pyrromethene 597 (PM597), a widely utilized commercial laser dye, is known to exhibit a comparatively lower fluorescence quantum yield than its structural analogs such as PM546 and PM560. To elucidate the underlying mechanism for this lower emission efficiency in PM597, we conducted a comprehensive investigation of its excited-state dynamics using femtosecond fluorescence upconversion and transient absorption spectroscopy. Our studies definitively reveal ultrafast decay pathways for the excited state of PM597 immediately following light absorption. Crucially, we were able to monitor, in real-time, the formation of a new excited-state species arising from an excited-state ring-puckering process. Our findings demonstrate that this ring-puckering process is highly sensitive to the viscosity of the medium, occurring with a time constant of ≈1.3 ps in low-viscosity solvents. These experimental results are in excellent agreement with previously proposed theoretical calculations. Furthermore, we have also elucidated the significant role played by substituents on the BODIPY ring in modulating this excited-state ring-puckering process.
This investigation aims to assess surface soil contamination by heavy metals in the new city of Ali Mendjeli (Constantine, Algeria). The methodological approach is based on energy-dispersive X-ray fluorescence (EDXRF), associated with multivariate statistical tools and pollution indices (enrichment factor (EF), geo-accumulation index ( I geo ), and contamination factor (CF)), in order to characterize the contamination levels and identify potential sources of pollution. X-ray fluorescence spectrometry analysis revealed the presence of trace metallic elements. These mainly consist of the following elements: chromium, zinc, nickel, and lead, with average concentrations exceeding the limit values set by the AFNOR U44-04 regulations. Moreover, no copper contamination was observed. The EF values indicate moderate pollution for Ni, Cr, Pb, and Zn, and low pollution for Cu. Meanwhile, the positive values of I geo varied between 0 and 2 for Cr, Ni, Zn, and Pb indicate low to moderate contamination, confirming the consistency between the pollution indices. On the other hand, the CF values for Cr, Zn, Ni, and Pb, ranging between 1 and 6, indicate moderate to high contamination, mainly of anthropogenic origin. Principal component analysis (PCA), coupled with hierarchical clustering (HC), indicates that the main sources of heavy metal contamination come from agricultural activities, metallurgical industries, gravel quarries, waste incineration, and road traffic.
Endometriosis progression is driven by oxidative stress and excessive angiogenesis within an inflammatory microenvironment. To overcome these challenges, we designed ROS/pH dual-responsive Alpelisib-loaded nanoparticles (Alp@TAT-AT7-NPs) functionalized with an anti-NRP1 peptide for targeted therapy. The nanoparticles exhibited superior stability, responsive drug release, and selective internalization by NRP1-overexpressing endothelial cells. In vitro results showed efficient inhibition of NRP1 and downstream PI3K/AKT signaling, along with decreased reactive oxygen species (ROS) and enhanced antioxidant enzyme activities. In an endometriosis rat model, treatment with Alp@TAT-AT7-NPs significantly reduced ectopic lesion burden and angiogenic markers (VEGF, CD34), while suppressing systemic inflammation and oxidative injury indicators such as IL-6, TNF-α, and MDA. Fluorescence imaging confirmed preferential accumulation of nanoparticles in CD31⁺ vascular regions. Mechanistic studies demonstrated that modulation of the Sema3A-NRP1-PI3K/AKT signaling axis restored redox homeostasis and inhibited pathological angiogenesis. These findings identify Alp@TAT-AT7-NPs as a synergistic nanoplatform that integrates microenvironment responsiveness with NRP1-targeted intervention, providing a promising therapeutic strategy for endometriosis.
Glycosylation is a ubiquitous post-translational modification that can reshape protein stability and protein-protein recognition, yet its mechanistic impact on cytokine-receptor interactions remains incompletely understood. Here, the influence of glycosylation of tumor necrosis factor alpha (TNFα) on tumor necrosis factor receptor I (TNFR1) and II (TNFR2) binding was investigated. For this purpose, biophysical measurements with molecular modeling and molecular dynamics (MD) simulations were conducted. The thermal stability of glycosylated and non-glycosylated TNFα, glycosylated receptor ectodomains, and their complexes was assessed by nano differential scanning fluorescence (nanoDSF). Glycosylated TNFα exhibited higher thermal stability and formed receptor complexes with increased apparent thermal stability compared with the non-glycosylated preparation. Binding affinity was quantified using microscale thermophoresis (MST), revealing that glycosylated TNFα binds TNFR2 more tightly than non-glycosylated TNFα corresponding to a four-fold affinity increase. For TNFR1, binding was detected for glycosylated TNFα, whereas no detectable binding of the non-glycosylated preparation was observed under the MST conditions used. Complementary MD simulations of TNFα/TNFR1 and TNFα/TNFR2 complexes, indicated that TNFα glycosylation are associated with increased complex stability, with effects dependent on glycan structure and receptor type. However, linear interaction energy analysis showed consistently favorable contributions of glycans only to complex stability, with stronger stabilization for longer glycans and a more pronounced effect for TNFR1 than for TNFR2. Together, our results are consistent with an ensemble-based qualitative model in which glycosylated TNFα forms more stable receptor complexes and may display enhanced receptor binding affinity through glycan-associated stabilization effects.
A major obstacle in oxycodone addiction treatment is relapse during abstinence. In rats, oxycodone seeking intensifies during abstinence, and orbitofrontal cortex (OFC) is critical for this incubation. Here we studied the role of submedius thalamus (Sub) and its reciprocal connections with OFC in this incubation in male rats. First, using the pharmacological inactivation approach, we examined the causal role of Sub in incubation of oxycodone craving. Next, combining fluorescence-conjugated cholera toxin subunit b (a retrograde tracer) with Fos (a neuronal activity marker), we assessed whether the activation of Sub and OFC connections was associated with incubated oxycodone seeking on abstinence day 15. Using an anatomical disconnection procedure, we then examined the causal role of interactions between Sub and OFC in incubated oxycodone seeking. We also tested the effect of unilateral Sub inactivation on incubated oxycodone seeking, and contralateral disconnection of Sub and OFC on non-incubated oxycodone seeking on abstinence day 1. We found that bilateral Sub inactivation decreased oxycodone seeking on abstinence day 15 but not on day 1, and incubated oxycodone seeking was associated with activation of both Sub→OFC and OFC→Sub projections. Further, both contralateral and ipsilateral disconnection of Sub and OFC decreased incubated oxycodone seeking. Unilateral Sub inactivation had no effect on the total responding during oxycodone seeking but impacted within-session responses. Lastly, contralateral disconnection of Sub and OFC had no effect on non-incubated oxycodone seeking. Together, these results demonstrated a novel role of Sub and its interactions with OFC in incubation of oxycodone craving.