Research on the Interaction Mechanisms between ScCO2 and Low-Rank/High-Rank Coal with the ReaxFF-MD Force Field.

CO2 geological sequestration in coal seams can be carried out to achieve the dual objectives of CO2 emission reduction and enhanced coalbed methane production, making it a highly promising carbon capture and storage technology. However, the injection of CO2 into coal reservoirs in the form of supercritical fluid (ScCO2) leads to complex physicochemical reactions with the coal seam, altering the properties of the coal reservoir and impacting the effectiveness of CO2 sequestration and methane production enhancement. In this paper, theoretical calculations based on ReaxFF-MD were conducted to study the interaction mechanism between ScCO2 and the macromolecular structures of both low-rank and high-rank coal, to address the limitations of experimental methods. The reaction of ScCO2 with low-rank coal and high-rank coal exhibited significant differences. At the swelling stage, the low-rank coal experienced a decrease in aromatic structure and aliphatic structure, and high-rank coal showed an increase in aromatic structure and a decrease in aliphatic structure, while the swelling phenomenon was more pronounced in high-rank coal. At the dissolution stage, low-rank coal was initially decomposed into two secondary molecular fragments, and then these recombined to form a new molecular structure; the aromatic structure increased and the aliphatic structure decreased. In contrast, high-rank coal showed the occurrence of stretches-breakage-movement-reconnection, a reduction in aromatic structure, and an increase in aliphatic structure. The primary reasons for these variations lie in the distinct molecular structure compositions and the properties of ScCO2, leading to different reaction pathways of the functional group and aromatic structure. The reaction pathways of functional groups and aromatic structures in coal can be summarized as follows: the breakage of the O-H bond in hydroxyl groups, the breakage of the C-OH bond in carboxyl groups, the transformation of aliphatic structures into smaller hydrocarbon compounds or the formation of long-chain alkenes, and various pathways involving the breakage, rearrangement, and recombination of aromatic structures. In low-rank coal, there is a higher abundance of oxygen-containing functional groups and aliphatic structures. The breakage of O-H and C-OH chemical bonds results in the formation of free radical ions, while some aliphatic structures detach to produce hydrocarbons. Additionally, some of these aliphatic structures combine with carbonyl groups and free radical ions to generate new aromatic structures. Conversely, in high-rank coal, a lower content of oxygen-containing functional groups and aliphatic structures, along with stronger intramolecular forces, results in fewer chemical bond breakages and makes it less conducive to the formation of new aromatic structures. These results elucidate the specific deformations of different chemical groups, offering a molecular-level understanding of the interaction between CO2 and coal.

CYTOR-NFAT1 feedback loop regulates epithelial-mesenchymal transition of retinal pigment epithelial cells.

Epithelial mesenchymal transition (EMT) occurring in retinal pigment epithelial cells (RPE) is a crucial mechanism that contributes to the development of age-related macular degeneration (AMD), a pivotal factor leading to permanent vision impairment. Long non-coding RNAs (lncRNAs) have emerged as critical regulators orchestrating EMT in RPE cells. In this study, we explored the function of the lncRNA CYTOR (cytoskeleton regulator RNA) in EMT of RPE cells and its underlying mechanisms. Through weighted correlation network analysis, we identified CYTOR as an EMT-related lncRNA associated with AMD. Experimental validation revealed that CYTOR orchestrates TGF-ß1-induced EMT, as well as proliferation and migration of ARPE-19 cells. Further investigation demonstrated the involvement of CYTOR in regulating the WNT5A/NFAT1 pathway and NFAT1 intranuclear translocation in the ARPE-19 cell EMT model. Mechanistically, CHIP, EMSA and dual luciferase reporter assays confirmed NFAT1's direct binding to CYTOR's promoter, promoting transcription. Reciprocally, CYTOR overexpression promoted NFAT1 expression, while NFAT1 overexpression increased CYTOR transcription. These findings highlight a mutual promotion between CYTOR and NFAT1, forming a positive feedback loop that triggers the EMT phenotype in ARPE-19 cells. These discoveries provide valuable insights into the molecular mechanisms of EMT and its association with AMD, offering potential avenues for targeted therapies in EMT-related conditions, including AMD.

Transcriptome-wide 1-methyladenosine functional profiling of messenger RNA and long non-coding RNA in bladder cancer.

Introduction: Post-transcriptional RNA modifications are crucial regulators of tumor development and progression. In many biological processes, N1-methyladenosine (m1A) plays a key role. However, little is known about the links between chemical modifications of messenger RNAs (mRNAs) and long noncoding RNAs (lncRNAs) and their function in bladder cancer (BLCA). Methods: Methylated RNA immunoprecipitation sequencing and RNA sequencing were performed to profile mRNA and lncRNA m1A methylation and expression in BLCA cells, with or without stable knockdown of the m1A methyltransferase tRNA methyltransferase 61A (TRMT61A). Results: The analysis of differentially methylated gene sites identified 16,941 peaks, 6,698 mRNAs, and 10,243 lncRNAs in the two groups. Gene ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analyses of the differentially methylated and expressed transcripts showed that m1A-regulated transcripts were mainly related to protein binding and signaling pathways in cancer. In addition, the differentially genes were identified that were also differentially m1A-modified and identified 14 mRNAs and 19 lncRNAs. Next, these mRNAs and lncRNAs were used to construct a lncRNA-microRNA-mRNA competing endogenous RNA network, which included 118 miRNAs, 15 lncRNAs, and 8 mRNAs. Finally, the m1A-modified transcripts, SCN2B and ENST00000536140, which are highly expressed in BLCA tissues, were associated with decreased overall patient survival. Discussion: This study revealed substantially different amounts and distributions of m1A in BLCA after TRMT61A knockdown and predicted cellular functions in which m1A may be involved, providing evidence that implicates m1A mRNA and lncRNA epitranscriptomic regulation in BLCA tumorigenesis and progression.

Lateral root primordium: Formation, influencing factors and regulation.

Roots are the primary determinants of water and nutrient uptake by plants. The structure of roots is largely determined by the repeated formation of new lateral roots (LR). A new lateral root primordium (LRP) is formed between the beginning and appearance of LR, which defines the organization and function of LR. Therefore, proper LRP morphogenesis is a crucial process for lateral root formation. The development of LRP is regulated by multiple factors, including hormone and environmental signals. Roots integrate signals and regulate growth and development. At the molecular level, many genes regulate the growth and development of root organs to ensure stable development plans, while also being influenced by various environmental factors. To gain a better understanding of the LRP formation and its influencing factors, this study summarizes previous research. The cell cycle involved in LRP formation, as well as the roles of ROS, auxin, other auxin-related plant hormones, and genetic regulation, are discussed in detail. Additionally, the effects of gravity, mechanical stress, and cell death on LRP formation are explored. Throughout the text unanswered or poorly understood questions are identified to guide future research in this area.

Postnatal treatment and evolution patterns of giant fetal hepatic hemangioma: a case series of 29 patients.

OBJECTIVES: To explore the clinical characteristics, postnatal treatment and prognosis of giant fetal hepatic hemangioma (GFHH). METHOD: Retrospective analysis was performed on children with giant fetal hepatic hemangioma (maximum tumor diameter > 40 mm) diagnosed by prenatal ultrasound and MRI from December 2016 to December 2020. These patients were observed and treated at the Children's Hospital of Fudan University after birth. The clinical data were collected to analyze the clinical characteristics, treatment, and prognosis of GFHH using independent sample t tests or Fisher's exact tests. RESULTS: Twenty-nine patients who were detected by routine ultrasound in the second and third trimester of pregnancy with giant fetal hepatic hemangiomas were included. The first prenatal ultrasound diagnosis of gestational age was 34.0 ± 4.3 weeks, ranging from 22 to 39 weeks. Of the patients, 28 had focal GFHHs and 1 had multifocal GFHHs. Surgery was performed, and the diagnosis was confirmed histopathologically in two patients. There were 8 cases with echocardiography-based evidence of pulmonary hypertension, 11 cases had a cardiothoracic ratio > 0.6, and 4 cases had hepatic arteriovenous fistula (AVF). The median follow-up time was 37 months (range: 14-70 months). During the follow-up, 12 patients received medical treatment with propranolol as the first-line therapy. The treatment group had a higher ratio of cardiothoracic ratio > 0.6 (P = 0.022) and lower albumin levels (P = 0.018). Four (14.8%) lesions showed postnatal growth before involuting. Complete response was observed in 13 (13/29) patients, and partial response was observed in 16 (16/29) patients. CONCLUSIONS: Fetal giant hepatic hemangioma is mainly localized, and its clinical outcome conforms to RICH (rapidly involuting) and PICH (partially involuting), but some fetal giant hepatic hemangiomas will continue to grow after birth and then gradually decrease. For uncomplicated giant fetal hepatic hemangioma, postnatal follow-up is the main concern, while those with complications require aggressive medical treatment. Propranolol may have no effect on the volume change of GFHH.

Application of Machine Learning and Deep EfficientNets in Distinguishing Neonatal Adrenal Hematomas From Neuroblastoma in Enhanced Computed Tomography Images.

Background: The aim of the study was to employ a combination of radiomic indicators based on computed tomography (CT) imaging and machine learning (ML), along with deep learning (DL), to differentiate between adrenal hematoma and adrenal neuroblastoma in neonates. Methods: A total of 76 neonates were included in this retrospective study (40 with neuroblastomas and 36 with adrenal hematomas) who underwent CT and divided into a training group (n = 38) and a testing group (n = 38). The regions of interest (ROIs) were segmented by two radiologists to extract radiomics features using Pyradiomics package. ML classifications were done using support vector machine (SVM), AdaBoost, Extra Trees, gradient boosting, multi-layer perceptron (MLP), and random forest (RF). EfficientNets was employed and classified, based on radiometrics. The area under curve (AUC) of the receiver operating characteristic (ROC) was calculated to assess the performance of each model. Results: Among all features, the least absolute shrinkage and selection operator (LASSO) logistic regression selected nine features. These radiomics features were used to construct radiomics model. In the training cohort, the AUCs of SVM, MLP and Extra Trees models were 0.967, 0.969 and 1.000, respectively. The corresponding AUCs of the test cohort were 0.985, 0.971 and 0.958, respectively. In the classification task, the AUC of the DL framework was 0.987. Conclusion: ML decision classifiers and DL framework constructed from CT-based radiomics features offered a non-invasive method to differentiate neonatal adrenal hematoma from neuroblastoma and performed better than the clinical experts.

Indole-Fused Benzoxepine Synthesis via Visible-Light-Driven Aerobic Dehydrogenative [5 + 2] Annulation.

A photocatalyzed oxidative dehydrogenative annulation between 2-(1H-indol-2-yl)phenols and alkenylphenols is presented. Various indole-fused benzoxepines can be obtained at room temperature using atom-efficient strategies. This method not only avoids the use of stoichiometric amounts of oxidants but also exhibits excellent atom economy by generating H2O as the only theoretical byproduct.

Immunogenicity and immunoprotection of the functional TL-HN fragment derived from tetanus toxin.

Tetanus toxin (TeNT) is a protein toxin produced by Clostridium tetani bacteria, which causes hyperreflexia and rhabdomyolysis by spastic paralysis. Like botulinum neurotoxin, TeNT comprises a heavy chain (HC) and a light chain (LC) linked via an interchain disulfide bond, which include the following three functional domains: a receptor-binding domain (Hc), a translocation domain (HN), and a catalytic domain (LC). Herein, we produced and characterized three functional domains of TeNT and three types of TeNT-derived L-HN fragments (TL-HN, TL-GS-HN and TL-2A-HN), which contained L and HN domains but lacked the Hc domain. The immunological effects of these different functional domains or fragments of TeNT were explored in an animal model. Our investigations showed the TL-HN functional fragment provided the best immunoprotection among all the TeNT functional domains. The TL-HN fragment, as a protective antigen, induced the highest levels of neutralizing antibodies, indicating that it might contain some crucial epitopes. Further experiments revealed that the protective effect of TL-HN was superior to that of the THc, TL, or THN fragments, either individually or in combination. Therefore, the TL-HN fragment exerts an important function in immune protection against tetanus toxin, providing a good basis for the development of TeNT vaccines or antibodies, and could serve as a promising subunit vaccine to replace THc or tetanus toxoid (TT).

Seasonal variations of macrozoobenthic community trophic structure in the waters adjacent to the seaweed beds of Dalian Island in the North Yellow Sea.

To investigate the effects of algal detritus export on the trophic structure of macrozoobenthic community in the adjacent benthic habitat during the bloom and decline of macroalgae, we collected macrozoobenthos from the adjacent sea area of Dalian Island in the North Yellow Sea in May (the algal bloom period) and August (the algal decay period) of 2020. We quantifyied the seasonal changes in the trophic structure of macrozoobenthic community by using carbon and nitrogen stable isotope techniques. Results showed that δ13C and δ15N values of macrozoo-benthos in May ranged from -23.14‰ to -14.24‰, 6.21‰ to 12.90‰, respectively, and -22.36‰ to -14.13‰, 5.33‰ to 12.00‰, respectively in August. Results of PERMANOVA analysis showed that δ13C values of macrozoobenthos differed significantly between the two months, while δ15N values were not significantly different. Based on the Euclidean distance, the macrozoobenthic communities in both months could be classified into five trophic functional groups. The trophic levels of macrozoobenthos ranged from 2.00 (Nitidotellina minuta) to 3.97 (Glycera onomichiensis) in May and from 2.00 (N. minuta) to 3.96 (G. onomichiensis) in August. The δ13C range, δ15N range, mean centroid distance, total area and corrected standard ellipse areas which represented community trophic structure indices in August were higher than those in May. Our results indicated that the trophic diversity level and trophic niche width of the macrozoobenthic community in the adjacent sea area of the seaweed bed were higher in the algal decline season.

Deciphering pathological behavior of pediatric medullary thyroid cancer from single-cell perspective.

Background: Pediatric medullary thyroid cancer (MTC) is one of the rare pediatric endocrine neoplasms. Derived from C cells of thyroid glands, MTC is more aggressive and more prompt to metastasis than other types of pediatric thyroid cancer. The mechanism remains unclear. Methods: We performed single-cell transcriptome sequencing on the samples of the primary tumor and metastases lymph nodes from one patient diagnosed with MTC, and it is the first single-cell transcriptome sequencing data of pediatric MTC. In addition, whole exome sequencing was performed and peripheral blood was regarded as a normal reference. All cells that passed quality control were merged and analyzed in R to discover the association between tumor cells and their microenvironment as well as tumor pathogenesis. Results: We first described the landscape of the single-cell atlas of MTC and studied the interaction between the tumor cell and its microenvironment. C cells, identified as tumor cells, and T cells, as the dominant participant in the tumor microenvironment, were particularly discussed in their development and interactions. In addition, the WES signature of tumor cells and their microenvironment were also described. Actively immune interactions were found, indicating B cells, T cells and myeloid cells were all actively participating in immune reaction in MTC. T cells, as the major components of the tumor microenvironment, proliferated in MTC and could be divided into clusters that expressed proliferation, immune effectiveness, and naive markers separately.

Three-Dimensional Pore Structure Characterization of Bituminous Coal and Its Relationship with Adsorption Capacity.

Bituminous coal reservoirs exhibit pronounced heterogeneity, which significantly impedes the production capacity of coalbed methane. Therefore, obtaining a thorough comprehension of the pore characteristics of bituminous coal reservoirs is essential for understanding the dynamic interaction between gas and coal, as well as ensuring the safety and efficiency of coal mine production. In this study, we conducted a comprehensive analysis of the pore structure and surface roughness of six bituminous coal samples (1.19% < Ro,max < 2.55%) using various atomic force microscopy (AFM) techniques. Firstly, we compared the microscopic morphology obtained through low-pressure nitrogen gas adsorption (LP-N2-GA) and AFM. It was observed that LP-N2-GA provides a comprehensive depiction of various pore structures, whereas AFM only allows the observation of V-shaped and wedge-shaped pores. Subsequently, the pore structure analysis of the coal samples was performed using Threshold and Chen's algorithms at ×200 and ×4000 magnifications. Our findings indicate that Chen's algorithm enables the observation of a greater number of pores compared to the Threshold algorithm. Moreover, the porosity obtained through the 3D algorithm is more accurate and closely aligns with the results from LP-N2-GA analysis. Regarding the effect of magnification, it was found that ×4000 magnification yielded a higher number of pores compared to ×200 magnification. The roughness values (Rq and Ra) obtained at ×200 magnification were 5-14 times greater than those at ×4000 magnification. Interestingly, despite the differences in magnification, the difference in porosity between ×200 and ×4000 was not significant. Furthermore, when comparing the results with the HP-CH4-GA experiment, it was observed that an increase in Ra and Rq values positively influenced gas adsorption, while an increase in Rsk and Rku values had an unfavorable effect on gas adsorption. This suggests that surface roughness plays a crucial role in gas adsorption behavior. Overall, the findings highlight the significant influence of different methods on the evaluation of pore structure. The 3D algorithm and ×4000 magnification provide a more accurate description of the pore structure. Additionally, the variation in 3D surface roughness was found to be related to coal rank and had a notable effect on gas adsorption.

Divergent structures and functions of the Cupin proteins in plants.

Cupin superfamily proteins have extensive functions. Their members are not only involved in the development of plants but also responded to various stresses. Whereas, the research on the Cupin members has not attracted enough attention. In this article, we summarized the research progress on these family genes in recent years and explored their evolution, structural characteristics, and biological functions. The significance of members of the Cupin family in the development of plant cell walls, roots, leaves, flowers, fruits, and seeds and their role in stress response are highlighted. Simultaneously, the prospective application of Cupin protein in crop enhancement was introduced. Some members can enhance plant growth, development, and resistance to adversity, thereby increasing crop yield. It will be as a foundation for future effective crop research and breeding.

Characteristics and associations of microbial community in the eye, anterior nares, and oropharynx of healthy adults: eye-nose microbiota transmission.

PURPOSE: We compared the microbial communities of the eye, anterior nares (ANs) and oropharynx (OP) of healthy adults to provide a basic understanding of the microbial associations among the three sites. METHODS: The name of the registry of our prospective study was "Study on the diversity of microbial flora in the eye, nose and oropharynx of healthy people". The trial number is ChiCTR2300067724 (https://www.chictr.org.cn/index.aspx). Swabs were collected from the eye, ANs and OP of 48 healthy adult participants for 16S rRNA gene amplicon sequencing. The bacterial community profiles and their functional associations were compared among the three sites. RESULTS: At a phylum level, the basic bacterial compositions in the eye and ANs were generally similar, and the predominant phyla were Actinobacteria, Proteobacteria and Firmicutes. In contrast, the OP microbiota was characterized by an increased abundance of Bacteroidetes. At a genus level, Corynebacterium, Cutibacterium and Staphylococcus were the most abundant in the eye and ANs. Prevotella-7, Alloprevatella, Haemophilus and Streptococcus were more abundant in the OP. Correlation analysis of the eye and ANs microbiota suggested that Cutibacterium and Micrococcus may migrate from the eye to nose (P < 0.05). CONCLUSIONS: The bacterial flora composition and function predictions of eye and ANs were similar, but differed from those of the OP in healthy adults. The OP bacterial flora distribution was markedly different, showing characteristics similar to that of the digestive tract flora. Thus, the eye and ANs microorganisms may be related in healthy individuals. Cutibacterium and Micrococcus may migrate from the eye to the nose.

Whole-genome sequencing of Paracoccus species isolated from the healthy human eye and description of Paracoccus shanxieyensis sp. nov.

Currently, the genus Paracoccus comprises 76 recognized species. Members of Paracoccus are mostly isolated from environmental, animal, and plant sources. This report describes and proposes a novel species of Paracoccus isolated from clinical specimens of the human ocular surface. We isolated two aerobic, Gram-stain-negative, non-spore-forming, coccoid or short rod-shaped, and non-motile strains (designated DK398T and DK608) from conjunctival sac swabs of two healthy volunteers. The results showed that the strains grew best under the conditions of 28°C, pH 7.0, and 1.0 % (w/v) NaCl. Sequence analysis based on the 16S rRNA gene showed that strains DK398T and DK608 were members of Paracoccus, most similar to Paracoccus laeviglucosivorans 43PT (98.54 and 98.62 %), Paracoccus litorisediminis GHD-05T (98.34 and 98.41 %), and Paracoccus limmosus NB88T (98.21 and 98.29 %). Phenotypic analysis showed that DK398T and DK608 were positive for catalase and oxidase, negative for producing N-acetyl-ß-glucosaminic acid, arginine dihydrolase, and ß-glucuronidase but positive for leucine arylamidase. The predominant isoprenoid quinone was Q-10, and the major polar lipids included phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, and an unidentified glycolipid. The major fatty acids (>10%) were summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c) and C16 : 0. The meso-diaminopimelic acid was found in the cell wall peptidoglycan of DK398T. The major cell wall sugars were ribose and galactose. Based on the results of phylogenetic analyses, low (<83.22 %) average nucleotide identity, digital DNA-DNA hybridization (<26.0%), chemotaxonomic analysis, and physiological properties, strain DK398T represents a novel species of the genus Paracoccus, for which the name Paracoccus shanxieyensis sp. nov. is proposed. The type strain is DK398T (=CGMCC 1.17227T=JCM 33719T).

Emergence of an ancient and pathogenic mammarenavirus.

Emerging zoonoses of wildlife origin caused by previously unknown agents are one of the most important challenges for human health. The Qinghai-Tibet Plateau represents a unique ecological niche with diverse wildlife that harbours several human pathogens and numerous previously uncharacterized pathogens. In this study, we identified and characterized a novel arenavirus (namely, plateau pika virus, PPV) from plateau pikas (Ochotona curzoniae) on the Qinghai-Tibet Plateau by virome analysis. Isolated PPV strains could replicate in several mammalian cells. We further investigated PPV pathogenesis using animal models. PPV administered via an intraventricular route caused trembling and sudden death in IFNαßR-/- mice, and pathological inflammatory lesions in brain tissue were observed. According to a retrospective serological survey in the geographical region where PPV was isolated, PPV-specific IgG antibodies were detected in 8 (2.4%) of 335 outpatients with available sera. Phylogenetic analyses revealed that this virus was clearly separated from previously reported New and Old World mammarenaviruses. Under the co-speciation framework, the estimated divergence time of PPV was 77-88 million years ago (MYA), earlier than that of OW and NW mammarenaviruses (26-34 MYA).

Insights into the environmental factors shaping lateral root development.

Roots are important organs of plants. Plants rely on roots for water, nutrients, and organic salts. In the whole root system, lateral roots (LRs) account for a large proportion and are critical to the development of the plant. Many environmental factors affect LR development. Therefore, a systematic understanding of these factors can provide a theoretical basis for creating optimal growth conditions for plants. In this paper, the factors affecting LR development are systematically and comprehensively summarized, and the molecular mechanism and regulatory network of LR development are described. Changes in the external environment not only lead to hormone homeostasis in plants but also affect the composition and activity of rhizosphere microbial communities, which in turn affect plants' nitrogen and phosphorus uptake and growth dynamics. LR development is influenced by hormone levels and external environment. In particular, auxin and abscisic acid coordinate with each other to maintain normal LR development. Of course, changes in the external environment are also important for root development, and they affect the intrinsic hormone levels of plants by affecting the accumulation and transport of hormones. For example, nitrogen, phosphorus, reactive oxygen species, nitric oxide, water, drought, light, and rhizosphere microorganisms affect LR development and plant tolerance in a variety of ways, including regulating hormone levels. This review summarizes the factors affecting LR development and the regulatory network and points out the direction for future research.

Expression dynamics of periodic transcripts during cancer cell cycle progression and their correlation with anticancer drug sensitivity.

BACKGROUND: The cell cycle is at the center of cellular activities and is orchestrated by complex regulatory mechanisms, among which transcriptional regulation is one of the most important components. Alternative splicing dramatically expands the regulatory network by producing transcript isoforms of genes to exquisitely control the cell cycle. However, the patterns of transcript isoform expression in the cell cycle are unclear. Therapies targeting cell cycle checkpoints are commonly used as anticancer therapies, but none of them have been designed or evaluated at the alternative splicing transcript level. The utility of these transcripts as markers of cell cycle-related drug sensitivity is still unknown, and studies on the expression patterns of cell cycle-targeting drug-related transcripts are also rare. METHODS: To explore alternative splicing patterns during cell cycle progression, we performed sequential transcriptomic assays following cell cycle synchronization in colon cancer HCT116 and breast cancer MDA-MB-231 cell lines, using flow cytometry and reference cell cycle transcripts to confirm the cell cycle phases of samples, and we developed a new algorithm to describe the periodic patterns of transcripts fluctuating during the cell cycle. Genomics of Drug Sensitivity in Cancer (GDSC) drug sensitivity datasets and Cancer Cell Line Encyclopedia (CCLE) transcript datasets were used to assess the correlation of genes and their transcript isoforms with drug sensitivity. We identified transcripts associated with typical drugs targeting cell cycle by determining correlation coefficients. Cytotoxicity assays were used to confirm the effect of ENST00000257904 against cyclin dependent kinase 4/6 (CDK4/6) inhibitors. Finally, alternative splicing transcripts associated with mitotic (M) phase arrest were analyzed using an RNA synthesis inhibition assay and transcriptome analysis. RESULTS: We established high-resolution transcriptome datasets of synchronized cell cycle samples from colon cancer HCT116 and breast cancer MDA-MB-231 cells. The results of the cell cycle assessment showed that 43,326, 41,578 and 29,244 transcripts were found to be periodically expressed in HeLa, HCT116 and MDA-MB-231 cells, respectively, among which 1280 transcripts showed this expression pattern in all three cancer cell lines. Drug sensitivity assessments showed that a large number of these transcripts displayed a higher correlation with drug sensitivity than their corresponding genes. Cell cycle-related drug screening showed that the level of the CDK4 transcript ENST00000547281 was more significantly associated with the resistance of cells to CDK4/6 inhibitors than the level of the CDK4 reference transcript ENST00000257904. The transcriptional inhibition assay following M phase arrest further confirmed the M-phase-specific expression of the splicing transcripts. Combined with the cell cycle-related drug screening, the results also showed that a set of periodic transcripts, for example, ENST00000314392 (a dolichyl-phosphate mannosyltransferase polypeptide 2 isoform transcript), was more associated with drug sensitivity than the levels of their corresponding gene transcripts. CONCLUSIONS: In summary, we identified a panel of cell cycle-related periodic transcripts and found that the levels of transcripts of drug target genes showed different values for predicting drug sensitivity, providing novel insights into alternative splicing-related drug development and evaluation.

Application of High-Throughput Sequencing Technology in Identifying the Pathogens in Endophthalmitis.

Infectious endophthalmitis is an important cause of vision loss worldwide. It is an inflammatory reaction caused by bacteria, fungi, and other micro-organisms and often occurs as a complication of intraocular surgery, especially following cataract surgery or intravitreal injection. The focus of the prevention and treatment of infectious endophthalmitis is the early detection of microbial flora, such as fungi or bacteria. Current identification methods for bacteria include Gram staining-based, culture-based, and polymerase chain reaction (PCR)-based methods. The matrix-assisted laser desorption/ionization time-of-flight mass spectrometry technology is now the standard identification method of bacteria and fungi after their isolation in culture. The remarkable sensitivity of PCR technology for the direct detection of micro-organisms in clinical samples makes it particularly useful in culture-positive and culture-negative endophthalmitis. Furthermore, PCR increases the rate of microorganism detection in intraocular samples by 20% and can provide a microbiology diagnosis in approximately 44.7-100% of the culture-negative cases. This review aims to introduce the development of different methods for the detection and identification of micro-organisms causing endophthalmitis through a literature review; introduce the research status of the first, second, and third-generation sequencing technologies in infectious endophthalmitis; and understand the research status of endophthalmitis microbial flora. For slow-growing and rare micro-organisms, high-throughput sequencing (HTS) offers advantages over conventional methods and provides a basis for the identification of pathogens in endophthalmitis cases with negative culture. It is a reliable platform for the identification of pathogenic bacteria of infectious endophthalmitis in the future and provides a reference for the clinical diagnosis and treatment of infectious endophthalmitis. The application of HTS technology may also be transformative for clinical microbiology and represents an exciting future direction for the epidemiology of ocular infections.

The species-level microbiota of healthy eyes revealed by the integration of metataxonomics with culturomics and genome analysis.

Objectives: To characterize the healthy ocular surface microbiota at the species level, including cultured and uncultured taxa. Methods: We integrated the metataxonomic method with culturomics and genome sequencing analysis of selected isolated strains to better illustrate the taxonomic structure of the ocular surface microbiota. The metataxonomics used the full-length 16S rRNA gene sequences and the operational phylogenetic unit strategy, which can precisely identify the cultured and uncultured or potentially new taxa to species level based on the phylogenetic tree constructed. Results: We detected 1,731 operational phylogenetic units (OPUs) in 196 healthy eyes from 128 people, affiliated to 796 cultured species, 784 potentially new species, and 151 potentially new higher taxa. The microbiota for each eye had 49.17 ± 35.66 OPUs. Of the 796 cultured species, 170 (21.36%) had previously caused clinical infections. Based on where they were initially isolated, the ocular surface microbiota mainly came from human body sites (34.55%), the environment (36.93%), plants (9.05%), animals (4.90%), and others; 428 strains were isolated from 20 eyes, affiliated to 42 species, and had come from the environment (33.33%) and the skin (16.67%). Of these, 47.62% had previously caused clinical infections. Genome analysis of 73 isolators revealed that 68.5% of them carried antibiotic resistance genes. The most frequently isolated genera, namely Staphylococcus, Streptococcus, and Moraxella, had an average of 5.30, four, and three resistance genes per strain, respectively. Discussion: The study found that the ocular surface microbiota mainly came from the environment, plants, animals, food, and human body sites such as the skin, oral cavity, upper respiratory tract, etc. No core member of ocular surface microbiota was detected at the species level. The human eyes were invaded and colonized by bacteria from the exposed environment, some of which were capable of causing infections in humans and carried antibiotic resistance genes. Preventive measures should be developed to protect our eyes from danger.

Comparison of the ocular surface microbiota between thyroid-associated ophthalmopathy patients and healthy subjects.

Purpose: Thyroid-associated ophthalmopathy (TAO) is a chronic autoimmune disease. In this study, high-throughput sequencing was used to investigate the diversity and composition of the ocular microbiota in patients with TAO. Methods: Patients with TAO did not receive treatment for the disease and did not have exposed keratitis. Patients with TAO (TAO group) and healthy individuals (control group) were compared. All samples were swabbed at the conjunctival vault of the lower eyelid. The V3 to V4 region of the 16S rDNA was amplified using polymerase chain reaction and sequenced on the Illumina HiSeq 2500 Sequencing Platform. Statistical analysis was performed to analyze the differences between the groups and the correlation between ocular surface microbiota and the disease. The ocular surface microbiota of patients and healthy individuals were cultured. Results: The ocular surface microbiota structure of TAO patients changed significantly. The average relative abundance of Bacillus and Brevundimonas increased significantly in the TAO group. Corynebacterium had a significantly decreased relative abundance (P<0.05). Paracoccus, Haemophilus, Lactobacillus, and Bifidobacterium were positively correlated with the severity of clinical manifestations or disease activity (P<0.05). Bacillus cereus and other opportunistic pathogens were obtained by culture from TAO patients. Conclusions: This study found that the composition of ocular microbiota in patients with TAO was significantly different from that in healthy individuals. The ocular surface opportunistic pathogens, such as Bacillus, Brevundimonas, Paracoccus, and Haemophilus in TAO patients, increase the potential risk of ocular surface infection. The findings of this study provide a new avenue of research into the mechanism of ocular surface in TAO patients.