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Exosomes are seen as promising biomarkers for minimally invasive liquid biopsies and disease surveillance. However, the complexity of body fluids, inherent heterogeneity, and tiny size of exosomes impede their extraction, consequently restricting their clinical application. In this study, in order to efficiently isolate exosomes from clinical samples, an irregular serpentine channel microfluidic chip (ExoSIC) was designed to continuously separate exosomes from plasma based on a magnetic-nanowaxberry (MNWB). In the ExoSIC, irregular serpentine microchannels are utilized to increase fluid chaotic mixing, hence improving exosome capture efficiency. In comparison to commonly used spherical magnetic particles, the designed MNWB can not only enhance the capture efficiency of exosomes, but also possess a size-exclusion effect to improve exosome purity. Consequently, the ExoSIC exhibited a large yield (24 times higher than differential centrifugation), optimum purity (greater than precipitation and similar to differential centrifugation), and high specificity. Furthermore, the ExoSIC was utilized for plasma-based cancer diagnosis by multiplex monitoring of five exosomal biomarkers (exosomal concentration, EGFR, EpCAM, SAA1 and FV), and the AUC reached 0.791. This work provides a comprehensive framework for exosome-based cancer diagnostics in order to meet clinical requirements for exosome isolation and downstream analysis. [ABSTRACT FROM AUTHOR]

Deoxyribonucleic acid (DNA) methylation is one of the epigenetic characteristics that result in heritable and revisable phenotype changes but without sequence changes in DNA. Aberrant methylation occurring at a specific locus was reported to be associated with cancers, insulin resistance, obesity, Alzheimer's disease, Parkinson's disease, etc. Therefore, locus‐specific DNA methylation can serve as a valuable biomarker for disease diagnosis and therapy. Recently, Clustered regularly interspaced short palindromic repeats (CRISPR)‐Cas systems are applied to develop biosensors for DNA, ribonucleic acid, proteins, and small molecules detection. Because of their highly specific binding ability and signal amplification capacity, CRISPR‐Cas assisted biosensor also serve as a potential tool for locus‐specific detection of DNA methylation. In this perspective, based on the detection principle, a detailed classification and comprehensive discussion of recent works about the latest advances in locus‐specific detection of DNA methylation using CRISPR‐Cas systems are provided. Furthermore, current challenges and future perspectives of CRISPR‐based locus‐specific detection of DNA methylation are outlined. [ABSTRACT FROM AUTHOR]

Rapid recruitment of neutrophils to an inflamed site is one of the hallmarks of an effective host defense mechanism. The main pathway through which this happens is by the innate immune response. Neutrophils, which play an important part in innate immune defense, migrate into lungs through the modulation actions of chemokines to execute a variety of pro-inflammatory functions. Despite the importance of chemokines in host immunity, little has been discussed on their roles in host immunity. A holistic understanding of neutrophil recruitment, pattern recognition pathways, the roles of chemokines and the pathophysiological roles of neutrophils in host immunity may allow for new approaches in the treatment of infectious and inflammatory disease of the lung. Herein, this review aims at highlighting some of the developments in lung neutrophil-immunity by focusing on the functions and roles of CXC/CC chemokines and pattern recognition receptors in neutrophil immunity during pulmonary inflammations. The pathophysiological roles of neutrophils in COVID-19 and thromboembolism have also been summarized. We finally summarized various neutrophil biomarkers that can be utilized as prognostic molecules in pulmonary inflammations and discussed various neutrophil-targeted therapies for neutrophil-driven pulmonary inflammatory diseases. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) encapsulated in tumor-derived exosomes are becoming ideal biomarkers for the early diagnosis and prognosis of lung cancer. However, the accuracy and sensitivity are often hampered by the extraction process of exosomal miRNA using traditional methods. Herein, this study developed a fluorogenic quantitative detection method for exosomal miRNA using the fluorescence quenching properties of molybdenum disulfide (MoS2) nanosheets and the enzyme-assisted signal amplification properties of duplex-specific nuclease (DSN). First, a fluorescently-labeled nucleic acid probe was used to hybridize the target miRNA to form a DNA/RNA hybrid structure. Under the action of the DSN, the DNA single strand in the DNA/RNA hybrid strand was selectively digested into smaller oligonucleotide fragments. At the same time, the released miRNA target triggers the next reaction cycle, so as to achieve signal amplification. Then, MoS2 was used to selectively quench the fluorescence of the undigested probe leaving the fluorescent signal of the fluorescently-labeled probe fragments. The fluorometric signals for miRNA-21 had a maximum excitation/emission wavelength of 488/518 nm. Most importantly, the biosensor was then applied for the accurate quantitative detection of miRNA-21 in exosome lysates extracted from human plasma and this method was able to successfully distinguish lung cancer patients from healthy people. This biosensor provides a simple, rapid, and a highly specific quantitative method for exosomal miRNA and has promising potential to be used in the early diagnosis of lung cancer. [ABSTRACT FROM AUTHOR]

Increasing epidemiological studies suggest that there is an association between vitamin D deficiency and risk of type 2 diabetes mellitus (T2DM). Therefore, randomized clinical trials (RCTs) have been performed to observe the effect of vitamin D supplementation on preventing T2DM, decreasing fasting plasma glucose (FPG) and improving insulin resistance to confirm the association between vitamin D and T2DM. However, the results of RCTs on controlling FPG level, improving insulin resistance and preventing T2DM in non-diabetics are inconsistent. In the present study, a systematic meta-analysis considering individual variation and intervention strategy was conducted to establish an objective and definitive conclusion. The results suggested that vitamin D supplementation had no significant effect on controlling FPG level, improving insulin resistance or preventing T2DM in non-diabetics in a pooled meta-analysis of 23 articles (containing 28 RCTs). However, stratified analysis indicated that supplementation of vitamin D had differential effects on FPG control, insulin sensitivity improvement and T2DM prevention in individuals with different baseline states: FPG was decreased for those with BMI <25 (P=0.048) or 20≤ 25(OH)D <30 ng/ml (P=0.002); insulin resistance was improved for those with 25(OH)D ≥30 ng/ml (P=0.021); and risk of T2DM was lower for pre-diabetic individuals (P=0.047) or for those with 25≤ BMI <30 (P=0.032). Additionally, the effect on T2DM prevention was improved when the supplement dose was >2,000 IU/day (P=0.047) and with intervention without calcium (P=0.047). Thus, further trials should focus on individual baselines and the supplementation strategy of vitamin D in the prevention of T2DM. [ABSTRACT FROM AUTHOR]

This work aimed to develop an ultrasensitive and specific immunosorbent assay for simultaneous detection of double DNA methylation marks. Being considered the most important indicators in disease diagnosis, clinical treatment, and prognosis, 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) were chosen as the proof-of-concept targets. The described strategy consisted of Phos-tag Biotin anchoring at streptavidin-magnetic nanoparticles, specific immune recognition of anti-5mC antibody and anti-5hmC antibody and labeling of Barcode-antibody, signal amplification of immune PCR and digital PCR machine. Under optimal conditions, the digital immuno-PCR assay showed a board dynamic range from 2.7 × 10−13 mol/L to 2.7 × 10−9 mol/L and the detection limits were 61.7 fmol/L for 5mC, and of 0.111 pmol/L for 5hmC. A 16-fold and 186-fold improvement of LOD were obtained by the proposed approach for 5mC and 5hmC detection compared with real-time immune PCR. The approach also showed ideal specificity, repeatability and stability. The recovery test demonstrated that the digital immuno-PCR assay is a promising platform for the simultaneous determination of the two DNA methylation marks in human serum sample. In this work, a simple and sensitive digital immune PCR assay was developed for simultaneous determination of 5-methylcytosine and 5-hydroxymethylcytosine in human serum. This simultaneous detection method can effectively save serum sample and improve efficiency. [Display omitted] • Simultaneous detection of 5mC and 5hmC were of great importance for physiological and pathological study. • We successfully developed a novel digital immuno-PCR assay for simultaneous detection of 5mC and 5hmC in human serum. • The LOD of digital immuno-PCR signal readout for 5mC was 6.17 × 10−14 mol/L (61.7 fmol/L) and for 5hmC was 1.11 × 10−13 mol/L (0.111 pmol/L). [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) encapsulated in tumor-derived exosomes are becoming ideal biomarkers for the early diagnosis and prognosis of lung cancer. However, the accuracy and sensitivity are often hampered by the extraction process of exosomal miRNA using traditional methods. Herein, this study developed a fluorogenic quantitative detection method for exosomal miRNA using the fluorescence quenching properties of molybdenum disulfide (MoS 2 ) nanosheets and the enzyme-assisted signal amplification properties of duplex-specific nuclease (DSN). First, a fluorescently-labeled nucleic acid probe was used to hybridize the target miRNA to form a DNA/RNA hybrid structure. Under the action of the DSN, the DNA single strand in the DNA/RNA hybrid strand was selectively digested into smaller oligonucleotide fragments. At the same time, the released miRNA target triggers the next reaction cycle, so as to achieve signal amplification. Then, MoS 2 was used to selectively quench the fluorescence of the undigested probe leaving the fluorescent signal of the fluorescently-labeled probe fragments. The fluorometric signals for miRNA-21 had a maximum excitation/emission wavelength of 488/518 nm. Most importantly, the biosensor was then applied for the accurate quantitative detection of miRNA-21 in exosome lysates extracted from human plasma and this method was able to successfully distinguish lung cancer patients from healthy people. This biosensor provides a simple, rapid, and a highly specific quantitative method for exosomal miRNA and has promising potential to be used in the early diagnosis of lung cancer.