Sb-doped FeOCl nanozyme-based biosensor for highly sensitive colorimetric detection of glutathione.

Nanozymes have been emerging as substitutes for natural enzymes to construct biosensors towards biomolecular detection. However, the detection of glutathione (GSH) by nanozyme-based biosensors still remains a great challenge for research on catalytic activity enhancement and the detection mechanism. In this work, Sb-doped iron oxychloride (Sb-FeOCl) with a well-defined nanorod-like structure is prepared by high-temperature calcination. Sb-FeOCl nanorods have high peroxidase-like activity, which can catalyze the decomposition of H2O2 into ·OH and then oxidize 3,3',5,5'-tetramethylbenzidine (TMB). In view of these intriguing observations, a reliable colorimetric method with a simple mixing and detection strategy is developed for the detection of GSH. The linear range of GSH detection is 1-36 µM. The detection limit of GSH reaches a low level of 0.495 µM (3σ/slope). The GSH sensing system also exhibits excellent specificity and anti-interference. Taking advantage of the advantages of the Sb-FeOCl nanorod-based biosensor, it can be used to quantitatively detect GSH levels in human serum. It can be anticipated that the Sb-FeOCl nanorods have broad prospects in the field of enzymatic biochemical reactions.

Establishment and Performance Evaluation of Multiplex PCR-Dipstick DNA Chromatography for Mycoplasma pneumoniae and Chlamydia pneumoniae Rapid Detection.

Methods: Nasopharyngeal swab samples of 300 children with an acute respiratory tract infection were detected by a multiplex PCR-dipstick chromatography assay, and the results were compared with the DNA sequencing and serum IgM antibody assay. Results: A multiplex PCR-dipstick DNA assay can specifically detect Mycoplasma pneumoniae and Chlamydia pneumoniae and shows a good specificity, with a minimum detection limit of 10 CFU/mL, respectively. Using DNA sequencing results as the gold standard, the sensitivity, specificity, positive predictive value, and negative predictive value of the multiplex PCR-dipstick DNA chromatography assay for the diagnosis of Mycoplasma pneumoniae were 96.61%, 100%, 100%, and 99.18% respectively, and those of Chlamydia pneumoniae were 95.24%, 100%, 100%, and 99.64% respectively. There was no statistical significance MP and CP diagnosis by the multiplex PCR-dipstick DNA assay and DNA sequencing (MP: P = 0.5; CP: P = 1.0), and the two assays had very high statistical consistency (MP: kappa = 0.979; CP: kappa = 0.974). The positive rate of the multiplex PCR-dipstick chromatography assay was significantly higher than that of the serum IgM antibody assay, with MP (17.7% vs. 13.3%), CP (5.7% vs. 3.3%), and mixed infection of MP and CP (1.3% vs. 0.67%). Conclusions: A multiplex PCR-dipstick chromatography assay was successfully established for the joint detection of Mycoplasma pneumoniae and Chlamydia pneumoniae within 2 hours. It is simple, fast, sensitive, accurate, cost-effective with good diagnostic performance, which can be used for small laboratories and point-of-care diagnosis.

Highly Sensitive Colorimetric Detection of Glutathione in Human Serum Based on Iron-Copper Metal-Organic Frameworks.

Emerging metal-organic framework (MOF)-based mimic enzymes have been exploited to design a colorimetric sensor for the detection of biomolecules. However, it is challenging to figure out the glutathione (GSH) detection method and the corresponding sensing mechanism using an MOF-based colorimetric sensor. In this work, a novel iron-copper MOF with high activity is synthesized by a wet-chemical method. A GSH colorimetric sensor based on the peroxidase-like properties of the iron-copper MOF is developed. Hydrogen peroxide is converted to hydroxyl radicals by the peroxidase-like properties of the iron-copper MOF mimic enzyme, which can catalyze the colorless 3,3',5,5'-tetramethylbenzidine (TMB) to blue oxidized TMB (ox-TMB). The kinetic constant of the MOF mimic enzyme (0.02 mM for H2O2) is superior to horseradish peroxidase (HRP). The GSH content can be quantified by proposing a sensor based on the colorimetric method and color turn-off mechanism. The turn-off mechanism of GSH analysis includes two aspects. On the one hand, the blue ox-TMB can be deoxidized to colorless TMB by GSH. On the other hand, hydroxyl radicals (•OH) can be consumed by GSH. The linear range and limit of detection are 2-20 and 0.439 µM, respectively. At the same time, GSH detection also shows good specificity and anti-interference characteristics. Therefore, MOF-based colorimetric sensors have been used to qualitatively and quantitatively measure GSH contents in human serum. The mechanism and application of the iron-copper MOF pave a way for the development of mimic enzymes with polymetallic active sites in the field of colorimetric sensing.

Mechanism and Application of Surface-Charged Ferrite Nanozyme-Based Biosensor toward Colorimetric Detection of l-Cysteine.

Peroxidase-like nanozymes with robust catalytic capacity and detection specificity have been proposed as substitutes to natural peroxidases in biochemical sensing. However, the catalytic activity enhancement, detection mechanism, and application of nanozyme-based biosensors toward l-cysteine (l-Cys) detection still remain significant challenges. In this work, a doped ferrite nanozyme with well-defined structure and surface charges is fabricated by a two-step method of continuous flow coprecipitation and high-temperature annealing. The resulted ferrite nanozyme possesses an average size of 54.5 nm and a zeta-potential of 6.45 mV. A high-performance biosensor is manufactured based on the peroxidase-like catalytic feature of the doped ferrite. The ferrite nanozyme can oxidize the 3,3',5,5'-tetramethylbenzidine (TMB) with the assistance of H2O2 because of the instinctive capacity to decompose H2O2 into ·OH. The Michaelis-Menten constants (0.0911 mM for TMB, 0.140 mM for H2O2) of the ferrite nanozyme are significantly smaller than those of horseradish peroxidase. A reliable colorimetric method is established to selectively analyze l-Cys via a facile mixing-and-detecting methodology. The detection limit and linear range are 0.119 µM and 0.2-20 µM, respectively. Taking the merits of the ferrite nanozyme-based biosensors, the l-Cys level in the human serum can be qualitatively detected. It can be anticipated that the surface-charged ferrite nanozyme shows great application prospects in the fields of bioanalytical chemistry and point-of-care testing.

Application of the amplification-free SERS-based CRISPR/Cas12a platform in the identification of SARS-CoV-2 from clinical samples.

The control of contagious or refractory diseases requires early, rapid diagnostic assays that are simple, fast, and easy-to-use. Here, easy-to-implement CRISPR/Cas12a-based diagnostic platform through Raman transducer generated by Raman enhancement effect, term as SERS-CRISPR (S-CRISPR), are described. The S-CRISPR uses high-activity noble metallic nanoscopic materials to increase the sensitivity in the detection of nucleic acids, without amplification. This amplification-free platform, which can be performed within 30-40 min of incubation time, is then used for detection of SARS-CoV-2 derived nucleic acids in RNA extracts obtained from nasopharyngeal swab specimens (n = 112). Compared with the quantitative reverse transcription polymerase chain reaction (RT-qPCR), the sensitivity and specificity of S-CRISPR reaches 87.50% and 100%, respectively. In general, the S-CRISPR can rapidly identify the RNA of SARS-CoV-2 RNA without amplification and is a potential strategy for nucleic acid point of care test (POCT).

Construction of an 11-microRNA-based signature and a prognostic nomogram to predict the overall survival of head and neck squamous cell carcinoma patients.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is a fatal malignancy owing to the lack of effective tools to predict overall survival (OS). MicroRNAs (miRNAs) play an important role in HNSCC occurrence, development, invasion and metastasis, significantly affecting the OS of patients. Thus, the construction of miRNA-based risk signatures and nomograms is desirable to predict the OS of patients with HNSCC. Accordingly, in the present study, miRNA sequencing data of 71 HNSCC and 13 normal samples downloaded from The Cancer Genome Atlas (TCGA) were screened to identify differentially expressed miRNAs (DEMs) between HNSCC patients and normal controls. Based on the exclusion criteria, the clinical information and miRNA sequencing data of 67 HNSCC samples were selected and used to establish a miRNA-based signature and a prognostic nomogram. Forty-three HNSCC samples were assigned to an internal validation cohort for verifying the credibility and accuracy of the primary cohort. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to explore the functions of 11 miRNA target genes. RESULTS: In total, 11 DEMs were successfully identified. An 11-miRNA risk signature and a prognostic nomogram were constructed based on the expression levels of these 11 DEMs and clinical information. The signature and nomogram were further validated by calculating the C-index, area under the curve (AUC) in receiver-operating characteristic curve analysis, and calibration curves, which revealed their promising performance. The results of the internal validation cohort shown the reliable predictive accuracy both of the miRNA-based signature and the prognostic nomogram. GO and KEGG analyses revealed that a mass of signal pathways participated in HNSCC proliferation and metastasis. CONCLUSION: Overall, we constructed an 11-miRNA-based signature and a prognostic nomogram with excellent accuracy for predicting the OS of patients with HNSCC.

Prognostic value of S1PR1 and its correlation with immune infiltrates in breast and lung cancers.

BACKGROUND: Sphingosine-1-phosphate receptor (S1PR1) is involved in vascular development, a key process in tumorigenesis. This study aimed to evaluate its roles in tumor development and prognosis. METHODS: S1PR1 expression levels were analyzed using TIMER and Oncomine database, and the prognostic significance of S1PR1 was assessed using PrognoScan and Kaplan-Meier plotter databases. The relationship between S1PR1 and tumor-infiltrated immune cells was analyzed using TIMER. RESULTS: S1PR1 expression was remarkably lower in breast and lung cancer tissues than in the corresponding normal tissues. Lower expression was related to poor overall survival and disease-free survival in breast invasive carcinoma (BRCA), lung adenocarcinoma (LUAD), and lung squamous cell carcinoma (LUSC). A functional network analysis confirmed the function of S1PR1 in regulating vasculogenesis. In addition, S1PR1 levels were significantly negative with regard to the tumor purity of BRCA (r = - 0.508, P = 1.76e-66), LUAD (r = - 0.353, P = 6.05e-16), and LUSC (r = - 0.402, P = - 5.20e-20). Furthermore, S1PR1 levels were significantly positive with regard to infiltrating CD8+ (r = 0.38, P = 5.91e-35) and CD4+ T cells (r = 0.335, P = 1.03e-26), macrophages (r = 0.219, P = 3.67e-12), neutrophils (r = 0.168, P = 2.03e-7), and dendritic cells (DCs) (r = 0.208, P = 9.14e-11) in BRCA; S1PR1 levels were significantly positive with regard to CD8+ T cells (r = 0.308, P = 3.61e-12), macrophages (r = 0.376, P = 1.01e-17), neutrophils (r = 0.246, P = 4.15e-8), and DCs (r = 0.207, P = 4.16e-6) in LUAD; and positive with regard to B cells (r = 0.356, P = 1.57e-15), CD8+ (r = 0.459, P = 3.83e-26) and CD4+ T cells (r = 0.338, P = 3.98e-14), macrophages (r = 0.566, P = 2.61e-45), neutrophils (r = 0.453, P = 1.79e-25), and DCs (r = 0.56, P = 2.12e-40) in LUSC. CONCLUSIONS: S1PR1 levels are positively correlated with multiple immune markers in breast and lung cancer. These observed correlations between S1PR1 and the prognosis and immune cell infiltration provide a foundation for further research on its immunomodulatory role in cancer.

Development and production of nanobodies specifically against green fluorescence protein.

Variable domains of heavy chains of camelid heavy-chain antibodies (VHHs) are known as nanobodies. Nanobodies are approximately 15 kDa in size with high affinity to their antigens. They can be easily manipulated and produced in microorganisms. In this study, an alpaca was immunized with purified green fluorescence protein (GFP) and a VHH library from lymphocytes of the immunized alpaca was constructed with a capacity of 6.7 × 107. The library was biopanned against GFP by phage display technique and four unique DNA sequences coding for anti-GFP nanobodies were identified by enzyme-linked immunosorbent assay, named a12, e6, d5, and b9. The four DNA sequences were then cloned into pADL-10b-6×His or pBAD24-Flag-6×His for expression in bacteria. Purified A12, E6, D5, and B9 were demonstrated to bind GFP specifically both in vitro by enzyme-linked immunosorbent assay and native-PAGE analysis and in vivo by immunofluorescence and immunoprecipitation. Taken together, our results demonstrate that anti-GFP nanobodies are successfully selected from the immune library, are produced in bacteria, and are available for basic research.Key Points• Four different GFP binders were successfully obtained from an immune VHH library.• The four GFP binders were successfully purified from bacteria. • Purified GFP binders can bind GFP both in vitro and in vivo and are ready for use in basic research.

Detection of Eight Respiratory Bacterial Pathogens Based on Multiplex Real-Time PCR with Fluorescence Melting Curve Analysis.

Background and Objective. Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae, Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Pseudomonas aeruginosa, and Mycobacterium tuberculosis are primary respiratory bacterial pathogens contributing to morbidity and mortality in developing countries. This study evaluated the diagnostic performance of multiplex real-time PCR with fluorescence melting curve analysis (MCA) assay, which was used to detect eight respiratory bacterial pathogens simultaneously. METHODS: A total of 157 sputum specimens were examined by multiplex real-time with fluorescence MCA, and the results were compared with the conventional culture method. RESULTS: Multiplex real-time PCR with fluorescence MCA specifically detected and differentiated eight respiratory bacterial pathogens by different melting curve peaks for each amplification product within 2 hours and exhibited high repeatability. The limit of detection ranged from 64 to 102 CFU/mL in the multiplex PCR system. Multiplex real-time PCR with fluorescence MCA showed a sensitivity greater than 80% and a 100% specificity for each pathogen. The kappa correlation of eight bacteria ranged from 0.89 to 1.00, and the coefficient of variation ranged from 0.05% to 0.80%. CONCLUSIONS: Multiplex real-time PCR with fluorescence MCA assay is a sensitive, specific, high-throughput, and cost-effective method to detect multiple bacterial pathogens simultaneously.

Potential circulating biomarkers of circulating chemokines CCL5, MIP-1ß and HA as for early detection of cirrhosis related to chronic HBV (hepatitis B virus) infection.

BACKGROUND: Due to no clinical symptoms in the compensated stage of cirrhosis, it is usually diagnosed when decompensated complications occur. In this study, the noninvasive circulating biomarkers for early detection to compensated stage of cirrhosis in patients with chronic HBV (hepatitis B virus) infection was explored. METHODS: According to the Guideline of Prevention and Treatment of Chronic Hepatitis B (2015 Update), 78 patients with CHB (chronic hepatitis B) were divided into mild group, moderate-to-advanced group, while 73 patients with HBV-related cirrhosis were divided into compensated group and decompensated group. Nineteen cytokines and chemokines, four serum liver fibrosis markers were measured using chemiluminescence. The expression of CCL5 in liver tissue was determined with immunohistochemistry. RESULTS: The CCL5 expression level in serum increased in CHB patients with aggravated liver injury and significantly decreased in cirrhosis patients with advanced liver fibrosis. ROC analysis revealed that the serum levels of CCL5, HA and MIP-1ß were effective in distinguishing patients with cirrhosis from patients with CHB, especially for CCL5. Increasing serum level of CCL5 in CHB patients was severely associated with disease progression. CONCLUSIONS: The serum levels of CCL5, HA and MIP-1ß maybe used to distinguish cirrhosis from CHB patients, moreover, CCL5 was the most reliable marker. The increasing serum levels of CCL5 were significantly related to disease progression in CHB patients.

miR-9 is an essential oncogenic microRNA specifically overexpressed in mixed lineage leukemia-rearranged leukemia.

MicroRNAs (miRNAs), small noncoding RNAs that regulate target gene mRNAs, are known to contribute to pathogenesis of cancers. Acute myeloid leukemia (AML) is a group of heterogeneous hematopoietic malignancies with various chromosomal and/or molecular abnormalities. AML with chromosomal translocations involving the mixed lineage leukemia (MLL) gene are usually associated with poor survival. In the present study, through a large-scale, genomewide miRNA expression assay, we show that microRNA-9 (miR-9) is the most specifically up-regulated miRNA in MLL-rearranged AML compared with both normal control and non-MLL-rearranged AML. We demonstrate that miR-9 is a direct target of MLL fusion proteins and can be significantly up-regulated in expression by the latter in human and mouse hematopoietic stem/progenitor cells. Depletion of endogenous miR-9 expression by an appropriate antagomiR can significantly inhibit cell growth/viability and promote apoptosis in human MLL-rearranged AML cells, and the opposite is true when expression of miR-9 is forced. Blocking endogenous miR-9 function by anti-miRNA sponge can significantly inhibit, whereas forced expression of miR-9 can significantly promote, MLL fusion-induced immortalization/transformation of normal mouse bone marrow progenitor cells in vitro. Furthermore, forced expression of miR-9 can significantly promote MLL fusion-mediated leukemogenesis in vivo. In addition, a group of putative target genes of miR-9 exhibited a significant inverse correlation of expression with miR-9 in a series of leukemia sample sets, suggesting that they are potential targets of miR-9 in MLL-rearranged AML. Collectively, our data demonstrate that miR-9 is a critical oncomiR in MLL-rearranged AML and can serve as a potential therapeutic target to treat this dismal disease.

Aggregated silver nanoparticles based surface-enhanced Raman scattering enzyme-linked immunosorbent assay for ultrasensitive detection of protein biomarkers and small molecules.

Lowering the detection limit is critical to the design of bioassays required for medical diagnostics, environmental monitoring, and food safety regulations. The current sensitivity of standard color-based analyte detection limits the further use of enzyme-linked immunosorbent assays (ELISAs) in research and clinical diagnoses. Here, we demonstrate a novel method that uses the Raman signal as the signal-generating system of an ELISA and combines surface-enhanced Raman scattering (SERS) with silver nanoparticles aggregation for ultrasensitive analyte detection. The enzyme label of the ELISA controls the dissolution of Raman reporter-labeled silver nanoparticles through hydrogen peroxide and generates a strong Raman signal when the analyte is present. Using this assay, prostate-specific antigen (PSA) and the adrenal stimulant ractopamine (Rac) were detected in whole serum and urine at the ultralow concentrations of 10(-9) and 10(-6) ng/mL, respectively. The methodology proposed here could potentially be applied to other molecules detection as well as PSA and Rac.

MicroRNA-196b promotes cell proliferation and suppress cell differentiation in vitro.

MicroRNA-196b (miR-196b) is frequently amplified and aberrantly overexpressed in acute leukemias. To investigate the role of miR-196b in acute leukemias, it has been observed that forced expression of this miRNA increases proliferation and inhibits apoptosis in human cell lines. More importantly, we show that this miRNA can significantly increase the colony-forming capacity of mouse normal bone marrow progenitor cells alone, as well as partially blocking the cells from differentiation. Taken together, our studies suggest that miRNA-196b may play an essential role in the development of MLL-associated leukemias through inhibiting cell differentiation and apoptosis, while promoting cell proliferation.

Two-layer cascaded catalytic hairpin assemblies based on locked nucleic acids for one-step and highly sensitive ctDNA detection.

Enzyme-free signal amplification of catalytic hairpin assembly (CHA) has enabled sensitive detection of circulating tumor DNA (ctDNA) in early clinical diagnosis. Conventional CHA strategies are restrained by the limited amplification efficiency of the single-stage system, and signal leakage from "breathing" influence and nuclease degradation. Here, we introduced two-layer cascaded locked nucleic acid (LNA)-assisted CHA circuits with the intelligent incorporation of LNA in the hairpins and reporter for the highly sensitive one-step detection of scarce ctDNA. The target-triggered upstream CHA reaction continuously generates hybrid products to catalyze the downstream CHA reaction for transducing the primary sensing event, and the released target and the produced hybrid product trigger the next catalytic reaction round at the same time and finally cascade to amplify the target ctDNA fluorescence output signal. Meanwhile, the stronger binding affinity of the LNA-DNA duplex endows the two-layer LNA-assisted CHA system with thermodynamic stability and nuclease resistance, and thus our designed system exhibits an excellent detection performance for target ctDNA in the range from 2 pM to 5 nM with a low detection limit of 0.6 pM. Significantly, the two-layer LNA-assisted CHA circuits have been successfully implemented for the feasible analysis of clinical samples. This two-layer cascaded LNA-assisted CHA strategy provides a promising high sensitivity tool for one-step detection of scarce ctDNA from complex clinical samples and would facilitate the reconfiguration of DNA circuit-based DNA nanotechnology for the precise analysis of other biomarkers in clinical research fields.

Mathematical Modeling of Initial Exothermic Behavior and Thixotropic Properties in Nanoclay-Enhanced Cementitious Materials.

In the realm of cementitious materials, integrating nanoclay shows promise in enhancing properties relevant to additive manufacturing. This paper presents a novel mathematical model that combines simple empirical dissolution/nucleation Avrami-like kinetics with a thixotropic kinetics equation. To analyze the initial exothermic peak, two sets of the calculation parameter function are built to describe the exothermic rate as a function of time, following an exponential pattern. This allows for the prediction of the changes in cumulative heat and heat rate during hydration, considering different concentrations of nanoclay. In the rheological aspect, the relationship between shear stress, shear rate, and time is modeled as a combination of exponential dependencies. This enables the prediction of the variations in shear stress with one variable while holding the other constant (either time or shear rate). By integrating these aspects, this model effectively describes both the first exothermal peak and the rheological behavior during cement hydration with the inclusion of nanoclay. Validated against experimental results, these models demonstrate good accuracy (overall below 3% error), reliability, and applicability. The findings offer valuable insights into the thermal and rheological aspects of concrete printing, enabling informed design decisions for both scientific and industrial applications.

Aberrant overexpression and function of the miR-17-92 cluster in MLL-rearranged acute leukemia.

MicroRNA (miRNA)-17-92 cluster (miR-17-92), containing seven individual miRNAs, is frequently amplified and overexpressed in lymphomas and various solid tumors. We have found that it is also frequently amplified and the miRNAs are aberrantly overexpressed in mixed lineage leukemia (MLL)-rearranged acute leukemias. Furthermore, we show that MLL fusions exhibit a much stronger direct binding to the locus of this miRNA cluster than does wild-type MLL; these changes are associated with elevated levels of histone H3 acetylation and H3K4 trimethylation and an up-regulation of these miRNAs. We further observe that forced expression of this miRNA cluster increases proliferation and inhibits apoptosis of human cells. More importantly, we show that this miRNA cluster can significantly increase colony-forming capacity of normal mouse bone marrow progenitor cells alone and, particularly, in cooperation with MLL fusions. Finally, through combinatorial analysis of miRNA and mRNA arrays of mouse bone marrow progenitor cells transfected with this miRNA cluster and/or MLL fusion gene, we identified 363 potential miR-17-92 target genes that exhibited a significant inverse correlation of expression with the miRNAs. Remarkably, these potential target genes are significantly enriched (P < 0.01; >2-fold) in cell differentiation, hematopoiesis, cell cycle, and apoptosis. Taken together, our studies suggest that overexpression of miR-17-92 cluster in MLL-rearranged leukemias is likely attributed to both DNA copy number amplification and direct up-regulation by MLL fusions, and that the miRNAs in this cluster may play an essential role in the development of MLL-associated leukemias through inhibiting cell differentiation and apoptosis, while promoting cell proliferation, by regulating relevant target genes.

A rapid and highly sensitive ctDNA detection platform based on locked nucleic acid-assisted catalytic hairpin assembly circuits.

As a promising biomarker of liquid biopsy, circulating tumor DNA (ctDNA) plays a paramount role in the early diagnosis of noninvasive cancer. The isothermal catalytic hairpin assembly (CHA) strategy has great potential for in vitro detection of ctDNA in low abundance. However, a traditional CHA strategy for ctDNA detection at the earlier stages of cancer remains extremely challenging, as annoying signal leakage from the 'breathing' phenomenon and nuclease degradation occur. Herein, we report a locked nucleic acid (LNA)-incorporated CHA circuit for the rapid and sensitive detection of target ctDNA. The target ctDNA intelligently catalyzed LNA-modified hairpins H1 and H2via a range of toehold-mediated strand displacement processes, leading to the continuous generation of an H1-H2 hybrid for the amplified fluorescence signal. In comparison to conventional CHA circuits, the stronger binding affinity of LNA-DNA bases greatly inhibited the breathing effect, which endowed it with greater thermodynamic stability and resistance to nuclease degradation in the LNA-assisted CHA system, thus achieving a high signal gain. The developed CHA circuit demonstrated excellent performance during target ctDNA detection, with a linear range from 10 pM to 5 nM, and its target detection limit was reached at 3.3 pM. Moreover, this LNA-assisted CHA system was successfully applied to the analysis of target ctDNA in clinical serum samples of breast cancer patients. This updated CHA system provides a general and robust platform for the sensitive detection of biomarkers of interest, thus facilitating the accurate identification and diagnosis of cancers.

An injectable, in situ forming and NIR-responsive hydrogel persistently reshaping tumor microenvironment for efficient melanoma therapy.

BACKGROUND: Melanoma is a highly aggressive form of skin cancer with increasing incidence and mortality rates. Chemotherapy, the primary treatment for melanoma, is limited by hypoxia-induced drug resistance and suppressed immune response at the tumor site. Modulating the tumor microenvironment (TME) to alleviate hypoxia and enhance immune response has shown promise in improving chemotherapy outcomes. METHODS: In this study, a novel injectable and in situ forming hydrogel named MD@SA was developed using manganese dioxide (MnO2) nanosheets pre-loaded with the chemotherapy drug doxorubicin (DOX) and mixed with sodium alginate (SA). The sustainable drug delivery, oxygen generation ability, and photothermal property of MD@SA hydrogel were characterized. The therapeutic efficacy of hydrogel was studied in B16F10 in vitro and B16F10 tumor-bearing mice in vivo. The immune effects on macrophages were analyzed by flow cytometry, real-time quantitative reverse transcription PCR, and immunofluorescence analyses. RESULTS: The MD@SA hydrogel catalyzed the tumoral hydrogen peroxide (H2O2) into oxygen, reducing the hypoxic TME, down-regulating hypoxia-inducible factor-1 alpha (HIF-1α) and drug efflux pump P-glycoprotein (P-gp). The improved TME conditions enhanced the uptake of DOX by melanoma cells, enhancing its efficacy and facilitating the release of tumor antigens. Upon NIR irradiation, the photothermal effect of the hydrogel induced tumor apoptosis to expose more tumor antigens, thus re-educating the M2 type macrophage into the M1 phenotype. Consequently, the MD@SA hydrogel proposes an ability to constantly reverse the hypoxic and immune-inhibited TME, which eventually restrains cancer proliferation. CONCLUSION: The injectable and in situ forming MD@SA hydrogel represents a promising strategy for reshaping the TME in melanoma treatment. By elevating oxygen levels and activating the immune response, this hydrogel offers a synergistic approach for TME regulation nanomedicine.

Towards Robust Person Re-Identification by Defending Against Universal Attackers.

Recent studies show that deep person re-identification (re-ID) models are vulnerable to adversarial examples, so it is critical to improving the robustness of re-ID models against attacks. To achieve this goal, we explore the strengths and weaknesses of existing re-ID models, i.e., designing learning-based attacks and training robust models by defending against the learned attacks. The contributions of this paper are three-fold: First, we build a holistic attack-defense framework to study the relationship between the attack and defense for person re-ID. Second, we introduce a combinatorial adversarial attack that is adaptive to unseen domains and unseen model types. It consists of distortions in pixel and color space (i.e., mimicking camera shifts). Third, we propose a novel virtual-guided meta-learning algorithm for our attack-defense system. We leverage a virtual dataset to conduct experiments under our meta-learning framework, which can explore the cross-domain constraints for enhancing the generalization of the attack and the robustness of the re-ID model. Comprehensive experiments on three large-scale re-ID benchmarks demonstrate that: 1) Our combinatorial attack is effective and highly universal in cross-model and cross-dataset scenarios; 2) Our meta-learning algorithm can be readily applied to different attack and defense approaches, which can reach consistent improvement; 3) The defense model trained on the learning-to-learn framework is robust to recent SOTA attacks that are not even used during training.

Rapid Evaluation of Lung Adenocarcinoma Progression by Detecting Plasma Extracellular Vesicles with Lateral Flow Immunoassays.

Extracellular vesicles (EVs) have been widely used in liquid biopsy to diagnose and monitor cancers. However, since samples containing EVs are usually body fluids with complex components, the cumbersome separation steps for EVs during detection limit the clinical application and promotion of EV detection methods. In this study, a dyad lateral flow immunoassay (LFIA) strip for EV detection, containing CD9-CD81 and EpCAM-CD81, was developed to detect universal EVs and tumor-derived EVs, respectively. The dyad LFIA strip can directly detect trace plasma samples and effectively distinguish the cancerous sample from healthy plasma. The limit of detection for detecting universal EVs was 2.4 × 105 mL-1. The whole immunoassay can be performed in 15 min and only consumes 0.2 µL of plasma for one test. To improve the suitability of a dyad LFIA strip in complex scenarios, a smartphone-based photographic method was developed, which provided a consistency of 96.07% to a specialized fluorescence LFIA strip analyzer. In further clinical testing, EV-LFIA discriminated lung cancer patient groups (n = 25) from healthy controls (n = 22) with 100% sensitivity and 94.74% specificity at the best cutoff. The detection of EpCAM-CD81 tumor EVs (TEVs) in lung cancer plasma revealed the differences in TEVs in individuals, which reflected the different treatment effects. TEV-LFIA results were compared with CT scan findings (n = 30). The vast majority of patients with increased TEV-LFIA detection intensity had lung masses that enlarged or remained unchanged in size, which reported no response to treatment. In other words, patients who reported no response (n = 22) had a high TEV level compared with patients who reported a response to treatment (n = 8). Taken together, the developed dyad LFIA strip provides a simple and rapid platform to characterize EVs to monitor lung cancer therapy outcomes.