Detection of male 2+0 and 1+0 carriers for spinal muscular atrophy by digital PCR.

Spinal muscular atrophy (SMA) is an autosomal recessive disease with a high carrier frequency. While current screening methods can identify 1+0 carriers, detecting 2+0 genotypes remains challenging, highlighting the need for additional research. Herein, we applied Digital Polymerase Chain Reaction (dPCR) to develop a novel approach for the detection of male carriers (DMC), especially for those with a 2+0 genotype. The clinical utility of DMC was evaluated in 39 semen samples. Multiple ligation-dependent probe amplification (MLPA) and pedigree analysis were performed on genomic DNA from 111 males and their family members. DMC identified 1+1, 2+1, and 1+0 genotypes in 21, 1, and 8 subjects. Importantly, seven men were identified as 2+0 carriers, while two men were excluded from the 2+0 carrier status. The results of DMC were consistent with those of MLPA and pedigree analysis. DMC provides an inexpensive and accurate method for determining the 2+0 and 1+0 genotypes.

Phosphorus bioavailability and the diversity of microbial community in sediment in response to modified calcium peroxide ceramsite capping.

Bioavailability of phosphorus (P) has close relationship with the microbial community in sediments and plays an important role in the sedimentary P release. However, little is known about the relationship between P bioavailability and microbial community under capping conditions. A 62-day experiment was conducted by capping with modified calcium peroxide ceramsite (MCPC) at different addition dosages (from 1:1 to 1:4, the ratio of mobile P in sediment to MCPC). P disappearance and release were expressed by the dissolved inorganic P (DIP) in overlying water. The results show that the proportion of disappeared P in released P was reduced sharply from 44% (Control) to 2% (1:4) under the capping with MCPC. Under the capping, the concentrations of DIP and Fe(Ⅱ) in pore water decreased markedly compared with the control, the removal up to 71.6% and 59.3% (mean, P < 0.05), respectively. The bioavailable P (BAP) presented the obvious decline under the capping with MCPC, which indicates the inhibition on the algae growth. The algae available P (AAP), water soluble P (WSP) and readily desorbable P (RDP) were reduced obviously at the 1:2 ratio compared with the other ratios. On the contrary, the biggest increase of Olsen-P in sediment was at the 1:2 ratio. This may be the explanation why the disappearance of DIP in the pore water was found. The result of microbial community structures in sediment shows that the relative abundance of Proteobacteria and Nitrospirae increased under the MCPC capping. It is analyzed that the microbial diversity is related negatively with the BAP in the sediments under the capping with MCPC, suggesting that microbial diversity is the key to control the BAP.

Effects of microwave on extracellular vesicles and microRNA in milk.

This study demonstrates the effects of microwaves on the microRNA (miRNA) content of milk and milk extracellular vesicles (EV). We determined the miRNA concentration in milk subjected to different treatments using real-time PCR and a spectrophotometer. The miRNA expression and total RNA content of the microwaved milk samples were lower when compared with untreated milk. We measured the microstructure and the size distribution by scanning electron microscopy and dynamic light scattering to verify the loss of miRNA in microwaved milk due to damage to the EV. The results revealed that 2 different-sized EV were present and had an average size of 147.50 and 22.14 nm, respectively. Furthermore, acridine orange staining showed that the total RNA content in microwaved milk EV was lower than that in cow milk. These results suggest that EV may confer the protection and the stability of the miRNA in milk.

Unlocking the Design Paradigm of In-Plane Heterojunction with Built-in Bifunctional Anion Vacancy for Unexpectedly Fast Sodium Storage.

Transition metal chalcogenide (TMD) electrodes in sodium-ion batteries exhibit intrinsic shortcomings such as sluggish reaction kinetics, unstable conversion thermodynamics, and substantial volumetric strain effects, which lead to electrochemical failure. This report unlocks a design paradigm of VSe2- x /C in-plane heterojunction with built-in anion vacancy, achieved through an in situ functionalization and self-limited growth approach. Theoretical and experimental investigations reveal the bifunctional role of the Se vacancy in enhancing the ion diffusion kinetics and the structural thermodynamics of Nax VSe2 active phases. Moreover, this in-plane heterostructure facilitates complete face contact between the two components and tight interfacial conductive contact between the conversion phases, resulting in enhanced reaction reversibility. The VSe2- x /C heterojunction electrode exhibits remarkable sodium-ion storage performance, retaining specific capacities of 448.7 and 424.9 mAh g-1 after 1000 cycles at current densities of 5 and 10 A g-1 , respectively. Moreover, it exhibits a high specific capacity of 353.1 mAh g-1 even under the demanding condition of 100 A g-1 , surpassing most previous achievements. The proposed strategy can be extended to other V5 S8- x and V2 O5- x -based heterojunctions, marking a conceptual breakthrough in advanced electrode design for constructing high-performance sodium-ion batteries.

A Chamber-Based Digital PCR Based on a Microfluidic Chip for the Absolute Quantification and Analysis of KRAS Mutation.

The Kirsten rat sarcoma virus gene (KRAS) is the most common tumor in human cancer, and KRAS plays an important role in the growth of tumor cells. Normal KRAS inhibits tumor cell growth. When mutated, it will continuously stimulate cell growth, resulting in tumor development. There are currently few drugs that target the KRAS gene. Here, we developed a microfluidic chip. The chip design uses parallel fluid channels combined with cylindrical chamber arrays to generate 20,000 cylindrical microchambers. The microfluidic chip designed by us can be used for the microsegmentation of KRAS gene samples. The thermal cycling required for the PCR stage is performed on a flat-panel instrument and detected using a four-color fluorescence system. "Glass-PDMS-glass" sandwich structure effectively reduces reagent volatilization; in addition, a valve is installed at the sample inlet and outlet on the upper layer of the chip to facilitate automatic control. The liquid separation performance of the chip was verified by an automated platform. Finally, using the constructed KRAS gene mutation detection system, it is verified that the chip has good application potential for digital polymerase chain reaction (dPCR). The experimental results show that the chip has a stable performance and can achieve a dynamic detection range of four orders of magnitude and a gene mutation detection of 0.2%. In addition, the four-color fluorescence detection system developed based on the chip can distinguish three different KRAS gene mutation types simultaneously on a single chip.

Detection and identification of oil spill species based on polarization information.

Aiming at the problem of poor oil identification accuracy in existing oil spill detection technologies, the polarization degree model of oil spill on rough sea surface under different azimuths and zenith angles was established based on Fresnel theory. The analytical expressions of visible light polarization degree in calm and fluctuating water surface were derived respectively, and the polarization degree model of oil spill in reflection space was constructed. The effectiveness of the method and its influence on the polarization distribution of oil spill were analyzed by simulation. A portable turntable was designed to test the polarization characteristics of the experiment, and the visible light polarization detection experiment was carried out. The visible light polarization images of five typical oil spills at different observation azimuth and zenth angles were obtained. The differences in the polarization degrees of different oil species were analyzed, and the correctness of the theoretical model was proved by experiments. The polarization detection experiment of visible light pBRDF was completed, which more intuitively showed the variation law of the polarization characteristics of light reflected by different oil spills in different spatial positions. Using polarization information to distinguish oil species is a useful supplement to the traditional oil spill detection method and has important significance to improve the marine pollution control ability.

Non-Invasive Measurement Using Deep Learning Algorithm Based on Multi-Source Features Fusion to Predict PD-L1 Expression and Survival in NSCLC.

Background: Programmed death-ligand 1 (PD-L1) assessment of lung cancer in immunohistochemical assays was only approved diagnostic biomarker for immunotherapy. But the tumor proportion score (TPS) of PD-L1 was challenging owing to invasive sampling and intertumoral heterogeneity. There was a strong demand for the development of an artificial intelligence (AI) system to measure PD-L1 expression signature (ES) non-invasively. Methods: We developed an AI system using deep learning (DL), radiomics and combination models based on computed tomography (CT) images of 1,135 non-small cell lung cancer (NSCLC) patients with PD-L1 status. The deep learning feature was obtained through a 3D ResNet as the feature map extractor and the specialized classifier was constructed for the prediction and evaluation tasks. Then, a Cox proportional-hazards model combined with clinical factors and PD-L1 ES was utilized to evaluate prognosis in survival cohort. Results: The combination model achieved a robust high-performance with area under the receiver operating characteristic curves (AUCs) of 0.950 (95% CI, 0.938-0.960), 0.934 (95% CI, 0.906-0.964), and 0.946 (95% CI, 0.933-0.958), for predicting PD-L1ES <1%, 1-49%, and ≥50% in validation cohort, respectively. Additionally, when combination model was trained on multi-source features the performance of overall survival evaluation (C-index: 0.89) could be superior compared to these of the clinical model alone (C-index: 0.86). Conclusion: A non-invasive measurement using deep learning was proposed to access PD-L1 expression and survival outcomes of NSCLC. This study also indicated that deep learning model combined with clinical characteristics improved prediction capabilities, which would assist physicians in making rapid decision on clinical treatment options.

Biological Functions and Cross-Kingdom Host Gene Regulation of Small RNAs in Lactobacillus plantarum-Derived Extracellular Vesicles.

Extracellular vesicle-mediated transfer of microRNAs is a novel mode of cell-to-cell genetic transmission. Extracellular vesicles produced by microbes have been shown to contain significant quantities of physiologically active molecules such as proteins, lipids, and RNA, which could be transported to host cells and play a key role in both inter-kingdom signaling and physiological responses. In this study, we identified sRNAs by sequencing small RNAs (sRNAs) from Lactobacillus plantarum-derived extracellular vesicles (LDEVs) and detected the expression levels of vesicular sRNAs using quantitative reverse transcription-polymerase chain reaction (RT-PCR), which demonstrated the presence of microRNA-sized RNAs (msRNAs) within these vesicles. We chose sRNA71, a highly expressed msRNA, for further investigation, predicted its potential target genes for the human genome, and indicated that it could be translocated into mammalian cells. The biological functions of this sRNA71 were subsequently explored through cellular proteomics, western blot, and luciferase reporter assay. According to the findings, transfection with synthetic sRNA71 mimics substantially reduced Tp53 expression in HEK293T cells and suppressed the gene expression through binding to the 3' UTR of Tp53 mRNA. In conclusion, it is hypothesized that microbial-derived extracellular vesicles serve as carriers of functional molecules such as sRNAs, which play an essential role in regulating microbial-host communication.

Application of Cell-Free Protein Synthesis System for the Biosynthesis of l-Theanine.

l-Theanine, as an active component of the leaves of the tea plant, possesses many health benefits and broad applications. Chemical synthesis of l-theanine is possible; however, this method generates chiral compounds and needs further isolation of the pure l-isoform. Heterologous biosynthesis is an alternative strategy, but one main limitation is the toxicity of the substrate ethylamine on microbial host cells. In this study, we introduced a cell-free protein synthesis (CFPS) system for l-theanine production. The CFPS expressed l-theanine synthetase 2 from Camellia sinensis (CsTS2) could produce l-theanine at a concentration of 11.31 µM after 32 h of the synthesis reaction. In addition, three isozymes from microorganisms were expressed in CFPS for l-theanine biosynthesis. The γ-glutamylcysteine synthetase from Escherichia coli could produce l-theanine at the highest concentration of 302.96 µM after 24 h of reaction. Furthermore, CFPS was used to validate a hypothetical two-step l-theanine biosynthetic pathway consisting of the l-alanine decarboxylase from C. sinensis (CsAD) and multiple l-theanine synthases. Among them, the combination of CsAD and the l-glutamine synthetase from Pseudomonas taetrolens (PtGS) could synthesize l-theanine at the highest concentration of 13.42 µM. Then, we constructed an engineered E. coli strain overexpressed CsAD and PtGS to further confirm the l-theanine biosynthesis ability in living cells. This engineered E. coli strain could convert l-alanine and l-glutamate in the medium to l-theanine at a concentration of 3.82 mM after 72 h of fermentation. Taken together, these results demonstrated that the CFPS system can be used to produce the l-theanine through the two-step l-theanine biosynthesis pathway, indicating the potential application of CFPS for the biosynthesis of other active compounds.

Artificial intelligence-assisted decision making for prognosis and drug efficacy prediction in lung cancer patients: a narrative review.

OBJECTIVE: In this review, we aim to present frontier studies in patients with lung cancer as it related to artificial intelligence (AI)-assisted decision-making and summarize the latest advances, challenges and future trend in this field. BACKGROUND: Despite increasing survival rate in cancer patients over the last decades, lung cancer remains one of the leading causes of death worldwide. The early diagnosis, accurate evaluation and individualized treatment are vital approaches to improve the survival rate of patients with lung cancer. Thus, decision making based on these approaches requires accuracy and efficiency beyond manpower. Recent advances in AI and precision medicine have provided a fertile environment for the development of AI-based models. These models have the potential to assist radiologists and oncologists in detecting lung cancer, predicting prognosis and developing personalized treatment plans for better outcomes of the patients. METHODS: We searched literature from 2000 through July 31th, 2021 in Medline/PubMed, the Web of Science, the Cochrane Library, ACM Digital Library, INSPEC and EMBASE. Key words such as "artificial intelligence", "AI", "deep learning", "lung cancer", "NSCLC", "SCLC" were combined to identify related literatures. These literatures were then selected by two independent authors. Articles chosen by only one author will be examined by another author to determine whether this article was relative and valuable. The selected literatures were read by all authors and discussed to draw reliable conclusions. CONCLUSIONS: AI, especially for those based on deep learning and radiomics, is capable of assisting clinical decision making from many aspects, for its quantitatively interpretation of patients' information and its potential to deal with the dynamics, individual differences and heterogeneity of lung cancer. Hopefully, remaining problems such as insufficient data and poor interpretability may be solved to put AI-based models into clinical practice.

The effect of microenvironment in the sediment on phosphorus immobilization under capping with ACPM and Phoslock®.

Currently, in situ capping is a typical popular geoengineering method for eutrophication control. It is crucial to better understand the effect of microenvironment change due to capping, such as amended calcium peroxide material (ACPM) and Phoslock®, on phosphorus (P) adsorption and immobilization under the addition of external P. The microenvironment in sediment was presented by the concentration of O2, NH4+, and Fe2+ and microbial activity. The P removal and immobilization were also analyzed. The results show that the stronger oxidation in the microenvironment under the capping with ACPM was due to the higher reduction of NH4+ and Fe2+ and the higher increase of microbial activity, compared to Phoslock®. Although, under the capping of ACPM, less amount of external P was removed and there was a faster release of sedimentary P, compared to Phoslock®, ACPM improved the transformation of P from mobile P fractions to inert P fractions. In addition, sedimentary P under the capping of ACPM presents less release than that under the capping of Phoslock® during the anaerobic incubation. However, the settlement of suspended solids decreased the function of capping. All these results indicated that the mechanism of P removal and immobilization was different under the capping of ACPM and Phoslock®.

[Effect of Calcium Peroxide Composite Tablets on Water Remediation and Phosphorus Control in Sediment].

Two types of calcium peroxide composite tablets (CPCTs) were prepared, and the inhibition effect on the release of endogenous phosphorus and the influence on the overlying water by mixed-dosing were investigated. The CPCTs were made of calcium peroxide (CaO2), calcined water purification sludge, and hydroxypropyl methylcellulose (HPMC), which were directly pressed into composite tablets, among which Tablet-B (T-B) contained ferrous sulfate (FeSO4) and Tablet-A (T-A) did not. Both the tablets agreed well with the Langmuir and Freundlich isothermal models; the theoretical maximum adsorption capacities of T-A and T-B on phosphorus were 110.908 mg·g-1 and 106.390 mg·g-1, respectively. Compared with the control group, the pH of overlying water was increased, the concentration of Chl-a was decreased by 42.75% and 60.82%, and the DO was increased by 53.73% and 63.30% in group A and B, respectively. The DIP of the overlying water decreased significantly by 54.93% and 25.11% in group A and B, respectively. For the interstitial water in sediment, the DIP in layer Ⅰ (0-2 cm) decreased significantly by 74.81% and 65.66% in group A and B, and the DIP in layer Ⅱ (2-4 cm) decreased significantly by 46.23% in group B, but not obviously in group A. The DIP in layer Ⅲ (4-6 cm) remained unchanged in group A and B. For phosphorus fractions in sediment, the proportion of NH4Cl-P in TP was significantly increased (layer Ⅰ: 16.87% and 13.11%; layer Ⅱ: 12.99% and 11.02%, in group A and B, respectively), and the proportion of Al-P in TP was significantly decreased (layer Ⅰ: 7.58% and 13.91%; layerⅡ: 9.86% and 7.28%, in group A and B, respectively). The other phosphorus fractions did not change significantly. Both T-A and T-B dosing can improve the microbial activity of the surface sediment, though T-A can improve the microbial activity more significantly.

Characterization of three different types of extracellular vesicles and their impact on bacterial growth.

Recently body fluids have been found to contain a class of nanoparticles released from cells, referred to as extracellular vesicles; exosomes are a type of small-diameter extracellular vesicle. We selected three types of sample: milk-derived exosomes, adipose-derived stem cell exosomes, and nanoparticles extracted from coconut water, to investigate their morphology, particle size distribution, protein content, and microRNA expression levels. Among the vesicles investigated, coconut nanoparticles had the greatest size distribution, and the protein content of coconut nanoparticles differed from that of mammalian exosomes. Using fluorescence microscopy, we determined that DiI-labeled extracellular vesicles could be absorbed by bacteria. Prominently, milk-derived exosomes could promote the growth of Escherichia coli K-12 MG1655 and Lactobacillus plantarum WCFS1. The studied extracellular vesicles could alter bacterial gene expression. Overall, this study identified differences in exogenous extracellular vesicles from different sources and revealed their supportive effects on microbial growth to make better utilization of microbial resources.

Isolation of Exosome-Like Nanoparticles and Analysis of MicroRNAs Derived from Coconut Water Based on Small RNA High-Throughput Sequencing.

In this study, the presence of microRNAs in coconut water was identified by real-time polymerase chain reaction (PCR) based on the results of high-throughput small RNA sequencing. In addition, the differences in microRNA content between immature and mature coconut water were compared. A total of 47 known microRNAs belonging to 25 families and 14 new microRNAs were identified in coconut endosperm. Through analysis using a target gene prediction software, potential microRNA target genes were identified in the human genome. Real-time PCR showed that the level of most microRNAs was higher in mature coconut water than in immature coconut water. Then, exosome-like nanoparticles were isolated from coconut water. After ultracentrifugation, some particle structures were seen in coconut water samples using 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate fluorescence staining. Subsequent scanning electron microscopy observation and dynamic light scattering analysis also revealed some exosome-like nanoparticles in coconut water, and the mean diameters of the particles detected by the two methods were 13.16 and 59.72 nm, respectively. In conclusion, there are extracellular microRNAs in coconut water, and their levels are higher in mature coconut water than in immature coconut water. Some exosome-like nanoparticles were isolated from coconut water, and the diameter of these particles was smaller than that of animal-derived exosomes.

Fermentation Results in Quantitative Changes in Milk-Derived Exosomes and Different Effects on Cell Growth and Survival.

The discovery of microRNAs encapsulated in milk-derived exosomes has revealed stability under extreme conditions reflecting the protection of membranes. We attempted to determine the variations in nanoparticles derived from milk after fermentation, and provide evidence to determine the effects of these exosomes on cells with potential bioactivity. Using scanning electron microscopy and dynamic light scattering, we compared the morphology and particle size distribution of exosomes from yogurt fermented with three different combinations of strains with those from raw milk. The protein content of the exosome was significantly reduced in fermented milk. The cycle threshold showed that the expression of miR-29b and miR-21 was relatively high in raw milk, indicating a loss of microRNA after fermentation. Milk-derived exosomes could promote cell growth and activate the mitogen-activated protein kinase pathway. These findings demonstrated biological functions in milk exosomes and provided new insight into the nutrient composition of dairy products.