Biology subjects

Gu, Y.

Publications and source records attributed to Gu, Y..

At least 217 records · Page 12Linked to original sources

bioRxiv2020

Coupling metabolic addiction with negative autoregulation to improve strain stability and pathway yield

Metabolic addiction, an organism that is metabolically addicted with a compound to maintain its growth fitness, is an underexplored area in metabolic engineering. Microbes with heavily engineered pathways or genetic circuits tend to experience metabolic burden leading to degenerated or abortive production phenotype during long-term cultivation or scale-up. A promising solution to combat metabolic instability is to tie up the end-product with an intermediary metabolite that is essential to the growth of the producing host. Here we present a simple strategy to improve both metabolic stability and pathway yield by coupling chemical addiction with negative autoregulatory genetic circuits. Naringenin and lipids compete for the same precursor with inversed pathway yield in oleaginous yeast. Negative autoregulation of the lipogenic pathways, enabled by CRISPRi and fatty acid-inducible promoters, repartitioned malonyl-CoA to favor flavonoid synthesis and increased naringenin production by 74.8%. With flavonoid-sensing hybrid promoters to control leucine synthesis, this flavonoid addiction phenotype confers a selective growth advantage to the naringenin-producing cell. The engineered yeast persisted 90.9% of naringenin titer up to 324 generations. Cells without flavonoid addiction regained growth fitness but lost 94.5% of the naringenin titer after cell passage beyond 300 generations. Metabolic addiction and negative autoregulation may be generalized as basic tools to eliminate metabolic heterogeneity, improve strain stability and pathway yield.

synthetic biology↗

bioRxiv2020

Isolating multiple formats of human monoclonal neutralizing antibodies against SARS-CoV-2 by in vitro site-directed antibody screening

Neutralizing antibody is one of the most effective interventions for acute pathogenic infection. Currently, over three million people have been identified for SARS-CoV-2 infection but SARS-CoV-2-specific vaccines and neutralizing antibodies are still lacking. SARS-CoV-2 infects host cells by interacting with angiotensin converting enzyme-2 (ACE2) via the S1 receptor-binding domain (RBD) of its surface spike glycoprotein. Therefore, blocking the interaction of SARS-CoV-2-RBD and ACE2 by antibody would cause a directly neutralizing effect against virus. In the current study, we selected the ACE2 interface of SARS-CoV-2-RBD as the targeting epitope for neutralizing antibody screening. We performed site-directed screening by phage display and finally obtained one IgG antibody (4A3) and several domain antibodies. Among them, 4A3 and three domain antibodies (4A12, 4D5, and 4A10) were identified to act as neutralizing antibodies due to their capabilities to block the interaction between SARS-CoV-2-RBD and ACE2-positive cells. The domain antibody 4A12 was predicted to have the best accessibility to all three ACE2-interfaces on the spike homotrimer. Pseudovirus and authentic SARS-CoV-2 neutralization assays showed that all four antibodies could potently protect host cells from virus infection. Overall, we isolated multiple formats of SARS-CoV-2-neutralizing antibodies via site-directed antibody screening, which could be promising candidate drugs for the prevention and treatment of COVID-19.

immunology↗

bioRxiv2020

GRP78 and Integrins Play Different Roles in Host Cell Invasion During Mucormycosis

Mucormycosis, caused by Rhizopus species, is a life-threatening fungal infection that occurs in patients immunocompromised by diabetic ketoacidosis (DKA), cytotoxic chemotherapy, immunosuppressive therapy, hematologic malignancies or severe trauma. Inhaled Rhizopus spores cause pulmonary infections in patients with hematologic malignancies, while patients with DKA are much more prone to rhinoorbital/cerebral mucormycosis. Here we show that R. delemar interacts with glucose-regulated protein 78 (GRP78) on nasal epithelial cells via its spore coat protein CotH3 to invade and damage the nasal epithelial cell. Expression of the two proteins is significantly enhanced by high glucose, iron and ketone body levels (hallmark features of DKA), potentially leading to frequently lethal rhinoorbital/cerebral mucormycosis. In contrast, R. delemar CotH7 recognizes integrin {beta}1 as a receptor on alveolar epithelial cells causing the activation of epidermal growth factor receptor (EGFR) leading to host cell invasion. Anti-integrin {beta}1 antibodies inhibit R. delemar invasion of alveolar epithelial cells and protect mice from pulmonary mucormycosis. Our results show that R. delemar interacts with different mammalian receptors depending on the host cell type. Susceptibility of patients with DKA primarily to rhinoorbital/cerebral disease can be explained by host factors typically present in DKA and known to upregulate CotH3 and nasal GRP78 thereby trapping the fungal cells within the rhino-orbital milieu, leading to subsequent invasion and damage. Our studies highlight that mucormycosis pathogenesis can potentially be overcome by the development of novel customized therapies targeting niche-specific host receptors or their respective fungal ligands. ImportanceMucormycosis caused by Rhizopus species is a fungal infection with often fatal prognosis. Inhalation of spores is the major route of entry, with nasal and alveolar epithelial cells among the first cells that encounter the fungi. In patients with hematologic malignancies or those undergoing cytotoxic chemotherapy, Rhizopus causes pulmonary infections. On the other hand, DKA patients predominantly suffer from rhinoorbital/cerebral mucormycosis. The reason for such disparity in disease types by the same fungus is not known. Here we show that, the unique susceptibility of DKA subjects to rhinoorbital/cerebral mucormycosis is likely due to specific interaction between nasal epithelial cell GRP78 and fungal CotH3, the expression of which increase in the presence of host factors present in DKA. In contrast, pulmonary mucormycosis is initiated via interaction of inhaled spores expressing CotH7 with integrin {beta}1 receptor which activates EGFR to induce fungal invasion of host cells. These results introduce plausible explanation to disparate disease manifestations in DKA versus hematologic malignancy patients and provide a foundation for development of therapeutic interventions against these lethal forms of mucormycosis.

microbiology↗

bioRxiv2020

Single-cell transcriptomic analysis identifies neocortical developmental differences between human and mouse

BackgroundThe specification and differentiation of neocortical projection neurons is a complex process under precise molecular regulation; however, little is known about the similarities and differences in cerebral cortex development between human and mouse at single-cell resolution. ResultsHere, using single-cell RNA-seq (scRNA-seq) data we explore the divergence and conservation of human and mouse cerebral cortex development using 18,446 and 7,610 neocortical cells. Systematic cross-species comparison reveals that the overall transcriptome profile in human cerebral cortex is similar to that in mouse such as cell types and their markers genes. By single-cell trajectories analysis we find human and mouse excitatory neurons have different developmental trajectories of neocortical projection neurons, ligand-receptor interactions and gene expression patterns. Further analysis reveals a refinement of neuron differentiation that occurred in human but not in mouse, suggesting that excitatory neurons in human undergo refined transcriptional states in later development stage. By contrast, for glial cells and inhibitory neurons we detected conserved developmental trajectories in human and mouse. ConclusionsTaken together, our study integrates scRNA-seq data of cerebral cortex development in human and mouse, and uncovers distinct developing models in neocortical projection neurons. The earlier activation of cognition -related genes in human may explain the differences in behavior, learning or memory abilities between the two species.

bioinformatics↗

bioRxiv2020

Engineering Yarrowia lipolytica as a chassis for de novo synthesis of aromatic-derived natural products and chemicals

AbstractsYarrowia lipolytica is a novel microbial chassis to upgrade renewable low-cost carbon feedstocks to high-value commodity chemicals and natural products. In this work, we systematically characterized and removed the rate-limiting steps of the shikimate pathway and achieved de novo synthesis of five aromatic chemicals in Y. lipolytica. We determined that eliminating amino acids formation and engineering feedback-insensitive DAHP synthases are critical steps to mitigate precursor competition and relieve the feedback regulation of shikimate pathway. Further overexpression of heterologous phosphoketolase and deletion of pyruvate kinase provided a sustained metabolic driving force that channels E4P (erythrose 4-phosphate) and PEP (phosphoenolpyruvate) precursors through the shikimate pathway. Precursor competing pathways and byproduct formation pathways were also blocked by inactivating chromosomal genes. To demonstrate the utility of our engineered chassis strain, three natural products, 2-phenylethanol (2-PE), p-coumaric acid and violacein, which were derived from phenylalanine, tyrosine and tryptophan, respectively, were chosen to test the chassis performance. We obtained 2426.22 {+/-} 48.33 mg/L of 2-PE, 593.53 {+/-} 28.75 mg/L of p-coumaric acid, 12.67 {+/-} 2.23 mg/L of resveratrol, 366.30 {+/-} 28.99 mg/L of violacein and 55.12 {+/-} 2.81 mg/L of deoxyviolacein from glucose in shake flask. The 2-PE production represents a 286-fold increase over the initial strain (8.48 {+/-} 0.50 mg/L). Specifically, we obtained the highest 2-PE, violacein and deoxyviolacein titer ever reported from the de novo shikimate pathway in yeast. These results set up a new stage of engineering Y. lipolytica as a sustainable biorefinery chassis strain for de novo synthesis of aromatic compounds with economic values.

synthetic biology↗

bioRxiv2020

KMT2D Links TGF-β Signalling to Non-Canonical Activin Pathway and Regulates Pancreatic Cancer Cell Plasticity

Although KMT2D, also known as MLL2, is known to play an essential role in development, differentiation, and tumor suppression, its role in pancreatic cancer development is not well understood. Here, we discovered a novel signaling axis mediated by KMT2D, which links TGF-{beta} to the activin A pathway. We found that TGF-{beta} upregulates a microRNA, miR-147b, which in turn leads to post-transcriptional silencing of KMT2D. Loss of KMT2D induces the expression and secretion of activin A, which activates a non-canonical p38 MAPK-mediated pathway to modulate cancer cell plasticity, promote a mesenchymal phenotype, and enhance tumor invasion and metastasis in mice. We observed a decreased KMT2D expression in human primary and metastatic pancreatic cancer. Furthermore, inhibition or knockdown of activin A reversed the pro-tumoral role of KMT2D. These findings reveal a tumor-suppressive role of KMT2D and identify miR-147b and activin A as novel therapeutic targets in pancreatic cancer.

cancer biology↗

bioRxiv2020

Renewable Fatty Acid Ester Production in Clostridium

Production of renewable chemicals through biological routes is considered as an urgent solution for fossil energy crisis. However, endproduct toxicity inhibits microbial performance and is a key bottleneck for biochemical production. To address this challenge, here we report an example of biosynthesis of high-value and easy-recoverable derivatives to alleviate endproduct toxicity and enhance bioproduction efficiency. By leveraging the natural pathways in solventogenic clostridia for co-producing acyl-CoAs, acids and alcohols as precursors, through rational screening for host strains and enzymes, systematic metabolic engineering-- including rational organization of ester-synthesizing enzymes inside of the cell, and elimination of putative prophages, we developed strains that can produce 20.3 g/L butyl acetate and 1.6 g/L butyl butyrate respectively, which were both the unprecedented levels in microbial hosts. Techno-economic analysis indicated a production cost of $986 per metric tonne for butyl acetate production from corn stover comparing to the market price of $1,200-1,400 per metric tonne of butyl acetate, suggesting the economic competitiveness of our developed bioprocess. Our principles of selecting the most appropriate host for specific bioproduction and engineering microbial chassis to produce high-value and easy-separable endproducts are highly applicable to other bioprocesses, and could lead to breakthroughs in biofuel/biochemical production and general bioeconomy.

bioengineering↗

bioRxiv2020

Accurate Diagnosis of Colorectal Cancer Based on Histopathology Images Using Artificial Intelligence

BackgroundAccurate and robust pathological image analysis for colorectal cancer (CRC) diagnosis is time-consuming and knowledge-intensive, but is essential for CRC patients treatment. The current heavy workload of pathologists in clinics/hospitals may easily lead to unconscious misdiagnosis of CRC based on their daily image analyses. MethodsBased on a state-of-the-art transfer-learned deep convolutional neural network in artificial intelligence (AI), we proposed a novel patch aggregation strategy for clinic CRC prediction/diagnosis using weakly labeled pathological whole slide image (WSI) patches. This approach was trained and validated using an unprecedented and enormously large number of 170,099 patches, >14,680 WSIs, from >9,631 subjects that covered diverse and representative clinical cases from multi-independent-sources across China, U.S., and Germany. ResultsOur innovative AI tool was consistently nearly perfectly agreed with (average Kappa-statistic 0.896) and even often better than most of the experienced expert pathologists when tested in diagnosing CRC WSIs from multi-centers. The average area under the receiver operating characteristics curve (AUC) of AI was greater than that of the pathologists (0.981 vs 0.970) and achieved the best performance among the application of other AI methods to CRC diagnosis. Our AI-generated heatmap highlights the image regions of cancer tissue/cells. ConclusionsThis first-ever generalizable AI system can handle large amounts of WSIs consistently and robustly without potential bias due to fatigue commonly experienced by clinical pathologists. Hence, it will drastically alleviate the heavy clinical burden of daily pathology diagnosis, and improve the treatment for CRC patients. This tool is generalizable to other cancer diagnosis based on image recognition.

pathology↗

bioRxiv2020

Oxytocin differentially modulates amygdala responses during top-down and bottom-up aversive anticipation

The ability to successfully regulate negative emotions such as fear and anxiety is vital for mental health. The neuropeptide oxytocin (OXT) acts as important modulator of emotion regulation, as reflected by reduced amygdala responses but increased amygdala-prefrontal cortex (PFC) functional connectivity in response to threatening stimuli. The present randomized, between-subject, placebo (PLC)-controlled pharmacological study combined intranasal administration of OXT with functional MRI during an explicit (cognitive) emotion regulation (i.e. distancing reappraisal) paradigm in 65 healthy male participants to investigate the modulatory effects of OXT on both bottom-up and top-down emotion regulation. OXT attenuated the activation in posterior insular cortex and amygdala during anticipation of top-down regulation of predictable threat stimuli in participants with high trait anxiety, providing evidence to support the anxiolytic action of OXT. In contrast, OXT enhanced amygdala activity during bottom-up anticipation of an unpredictable threat stimulus in participants with low trait anxiety. OXT may thus facilitate top-down goal-directed attention by attenuating amygdala activity in high anxiety individuals, while promote bottom-up attention/vigilance to unexpected threat by enhancing anticipatory amygdala activity in low anxiety individuals. The opposite effects of OXT on anticipatory amygdala activation in high versus low anxiety individuals may suggest a baseline anxiety level dependent mechanism via which OXT promotes optimal levels of amygdala activation during the anticipation of an imminent threat. OXT may thus have the potential to promote an adaptive balance between bottom-up and top-down attention systems depending on individual levels of pre-treatment trait anxiety levels.

neuroscience↗

bioRxiv2020

Preserving Vascular Integrity Protects Mice Against Multidrug-Resistant Gram-Negative Bacterial Infection

The rise in multidrug resistant (MDR) organisms portends a serious global threat to the healthcare system with nearly untreatable infectious diseases, including pneumonia and its often fatal sequelae, acute respiratory distress syndrome (ARDS) and sepsis. Gram-negative bacteria (GNB) including Acinetobacter baumannii, Pseudomonas aeruginosa, and carbapenemase-producing Klebsiella pneumoniae (CPKP), are among the World Health Organization and National Institutes of Healths high priority MDR pathogens for targeted development of new therapies. Here we show that stabilizing the hosts vasculature by genetic deletion or pharmacological inhibition of the small GTPase ADP-ribosylation factor 6 (ARF6) increases survival rates of mice infected with A. baumannii, P. aeruginosa, CPKP pneumonia. We show that pharmacological inhibition of ARF6-GTP phenocopies endothelial-specific Arf6 disruption in enhancing survival of mice with A. baumannii pneumonia, suggesting that inhibition is on target. Finally, we show that the mechanism of protection elicited by these small molecule inhibitors is by restoration of vascular integrity disrupted by GNB lipopolysaccharide (LPS) activation of TLR4/MyD88/ARNO/ARF6 pathway. By targeting the hosts vasculature with small molecule inhibitors of ARF6 activation, we circumvent microbial drug resistance and provide a potential alternative/adjunctive treatment for emerging and re-emerging pathogens.

microbiology↗

bioRxiv2020

Developing an endogenous quorum-sensing based CRISPRi circuit for autonomous and tunable dynamic regulation of multiple targets in industrial Streptomyces

Dynamic regulation has emerged as an effective strategy to improve product titers by balancing metabolic networks, which can be implemented by coupling gene expression to pathway-independent regulatory elements, such as quorum-sensing (QS) systems. However, these QS-based circuits are often created on heterologous systems and must be carefully tuned through tedious testing and optimization process to make them work well, which hampers their application in industrial microbes including streptomycetes. In this study, we design a pathway-independent QS circuit by directly integrating an endogenous QS system with CRISPRi (named EQCi) in the industrial rapamycin-producing strain Streptomyces rapamycinicus. EQCi has the advantages of both the QS system and CRISPRi, which enables tunable, fully autonomous and dynamic regulation of multiple targets simultaneously. To demonstrate its effectiveness, we downregulate three key nodes in essential pathways separately to divert metabolic flux toward rapamycin biosynthesis. In each case, significant increases in rapamycin titers are achieved. We further apply EQCi to simultaneously control these three key nodes with proper repression strength by changing sgRNA targeting positions. The final rapamycin titer reaches to 1836{+/-}191 mg/L, which is the highest reported titer. Notably, compared to traditional static engineering strategy, which result in growth arrest and suboptimal rapamycin titers, EQCi regulation substantially promote rapamycin titers without affecting cell growth, which indicates that it can achieve the trade-off between essential pathways and product synthesis. Collectively, this study provides a simple and effective strategy for optimizing product titers and may have the potential to apply to other industrial microorganisms.

bioengineering↗

bioRxiv2020

Derivation and characterization of chimera-competent eXtra-Embryonic eNdoderm 1 (XEN) cells from pig blastocysts

In this article, we report for the first time the derivation and characterization of extra-embryonic endoderm (XEN) cells from primitive endoderm (PrE) of porcine (p) embryos. The pXEN cells can be reliably and reproducibly generated from parthenote, in vitro and in vivo derived embryos. The pXEN cells retained all the hallmarks of PrE including expression of canonical PrE and XEN cell markers (GATA4, GATA6, SOX17, SALL4, FOXA2, and HNF4A). Transcriptome analysis further confirmed their XEN cell origin. The pXEN cells when introduced into blastocyst stage embryo contributed to wide-spread chimerism including visceral yolk sac, chorion, as well as embryonic gut and liver primordium in the fetus. The pXEN cells were shown to be an efficient nuclear donor for generating cloned offspring. Taken together, pXEN cells fulfil a longstanding need for a stable, chimera-competent, and nuclear transfer-compatible porcine embryonic cells with applications for agriculture and medicine. Significance StatementWe report for the first time, the derivation and characterization of extraembryonic endoderm (XEN) stem cells from porcine (p) embryos. The pXEN cells can be reliably and reproducibly derived from primitive endoderm precursors. When injected into blastocyst-stage embryos, the pXEN cells have contributed to wide-spread chimerism including visceral yolk sac, chorion of the extraembryonic membranes, as well as definitive endoderm of the fetus, primarily the embryonic gut and liver primordium. Additionally, these XEN cells have proven to be an efficient nuclear donor for generating cloned offspring. These newly discovered stem cells provide a novel model for studying lineage segregation, as well as a source for interspecies chimeras for generating endodermal organs, and for genome editing in livestock.

developmental biology↗

bioRxiv2019

Light-induced polyp retraction and tissue rupture in the photosensitive, reef-building coral Acropora muricata

Coral reefs are in alarming decline due to climate emergency, pollution and other man-made disturbances. The numerous ecosystem services derived from coral reefs are underpinned by the growth and physical complexity of reef-forming corals. Our knowledge of their fundamental biology is limited by available technology. We need a better understanding of larval settlement and development, skeletogenesis, interactions with pathogens and symbionts, and how this biology interacts with environmental factors such as light exposure, temperature, and ocean acidification. We here focus on a fast-growing key coloniser, Acropora muricata. To enable dynamic imaging of the photosensitive organism at different scales, we developed light-sheet illumination for fluorescence microscopy of small coral colonies. Our approach reveals live polyps in previously unseen detail. An imaging range for Acropora muricata with no measurable photodamage is defined based upon polyp expansion, coral tissue reaction, and photobleaching. We quantify polyp retraction as a photosensitive behavioural response and show sparse zooxanthellar expulsion and coral tissue rupture at higher intensities of blue light. The simple and flexible technique enables non-invasive continuous dynamic imaging of highly photosensitive organisms with sizes between 1 mm3 and 5 cm3, for eight hours, at high temporal resolution, on a scale from multiple polyps down to cellular resolution. This live imaging tool opens a new window into the dynamics of reef-building corals.

developmental biology↗

bioRxiv2019

Patient stratification of clear cell renal cell carcinoma using the global transcription factor activity landscape derived from RNA-seq data

Clear cell renal cell carcinoma represents the most common type of kidney cancer. Precision medicine approach to ccRCC requires an accurate stratification of patients that can predict prognosis and guide therapeutic decision. Transcription factors are implicated in the initiation and progression of human carcinogenesis. However, no comprehensive analysis of transcription factor activity has been proposed so far to realize patient stratification. Here we propose a novel approach to determine the subtypes of ccRCC patients based on global transcription factor activity landscape. Using the TCGA cohort dataset, we identified different subtypes that have distinct upregulated biomarkers and altered biological pathways. More important, this subtype information can be used to predict the overall survival of ccRCC patients. Our results suggest that transcription factor activity can be harnessed to perform patient stratification.

systems biology↗

bioRxiv2019

DNB-Based On-Chip Motif Finding (DocMF): a High-Throughput Method to Profile Different Types of Protein-DNA Interactions

Here we report a highly sensitive DNB-based on-chip Motif Finding (DocMF) system that utilizes high throughput next-generation-sequencing (NGS) chips to profile protein binding or cleaving activity. Using DocMF, we successfully identified a variety of endonuclease recognition sites and the protospacer-adjacent-motif (PAM) sequences of different CRISPR systems. Our DocMF platform can simultaneously screen both 5 and 3 PAM regions with high coverage using the same NGS library/chip. For the well-studied SpCas9, our DocMF platform identified a small proportion of noncanonical 5-NAG-3 ([~]5%) and 5-NGA-3 ([~]1.6%), in addition to its common PAMs, 5-NGG-3 ([~]89.9%). We also used the DocMF to assay two uncharacterized Cas endonucleases, VeCas9 and BvCpf1. VeCas9 PAMs were not detected by the conventional PAM depletion method. However, DocMF discovered that both VeCas9 and BvCpf1 required broader and more complicated PAM sequences for target recognition. VeCas9 preferred the R-rich motifs, whereas BvCpf1 used the T-rich PAMs. Moreover, after slightly changing the experimental protocol, we observed that dCas9, a DNA-binding protein lacking endonuclease activity, preferably binded to the previously reported PAMs 5-NGG-3. In summary, our studies demonstrate that DocMF is the first tool with the capacity to exhaustively assay both the binding and the cutting properties of different DNA-binding proteins.

molecular biology↗

bioRxiv2019

Selective targeting of an oncogenic KRAS mutant allele by CRISPR/Cas9 induces efficient tumor regression

BackgroundKRAS is one of the most frequently mutated oncogenes in human cancers, but its activating mutations have remained undruggable due to its picomolar affinity for GTP/GDP and its smooth protein structure resulting in the absence of known allosteric regulatory sites. ResultsWith the goal of treating mutated KRAS-driven cancers, two CRISPR systems, CRISPR-SpCas9 genome-editing system and transcription-regulating system dCas9-KRAB, were developed to directly deplete KRAS mutant allele or to repress its transcription in cancer cells, respectively, through guide RNA specifically targeting the mutant but not wild-type allele. The effect of in vitro proliferation and cell cycle on cancer cells as well as in vivo tumor growth was examined after delivery of Cas9 system. SpCas9 and dCas9-KRAB systems with sgRNA targeting the mutant allele both blocked the expression of mutant KRAS gene, leading to an inhibition of cancer cell proliferation. Local adenoviral injections using SpCas9 and dCas9-KRAB systems both suppressed tumor growth in vivo. The gene-depletion system (SpCas9) performed more effectively than the transcription-suppressing system (dCas9-KRAB) on tumor inhibition. Application of both Cas9 systems to wild-type KRAS tumor cells did not affect cell proliferation in vitro and in vivo. Furthermore, through bioinformatic analysis of 31555 SNP mutations of the top 20 cancer driver genes, we showed that our mutant-specific editing strategy could be extended to a list of oncogenic mutations with high editing potentials, and this pipeline can be applied to analyze the distribution of PAM sequence in the genome to survey the best targets for other editing purpose. ConclusionsWe successfully developed both gene-depletion and transcription-suppressing systems to specifically target an oncogenic mutant allele of KRAS which led to significant tumor regression. It provides a promising strategy for the treatment of tumors with driver gene mutations.

genetics↗

bioRxiv2019

Diagnostic value of circulating tumor DNA as an effective biomarker in cervical cancer: a meta-analysis

The applications of liquid biopsy have attracted much attention in biomedical research in recent years. Circulating cell-free DNA (cfDNA) in the serum may serve as a unique tumor marker in various types of cancer. Circulating tumor DNA (ctDNA) is a type of serum cfDNA found in patients with cancer and contains abundant information regarding tumor characteristics, highlighting its potential diagnostic value in the clinical setting. However, the diagnostic value of cfDNA as a biomarker in cervical cancer remains unclear. Here, we performed a meta-analysis to evaluate the applications of ctDNA as a biomarker in cervical cancer. A systematic literature search was performed using PubMed, Embase, and WANFANG MED ONLINE databases up to March 18, 2019. All literature was analyzed using Meta Disc 1.4 and STATA 14.0 software. Diagnostic measures of accuracy of ctDNA in cervical cancer were pooled and investigated. Fifteen studies comprising 1109 patients with cervical cancer met our inclusion criteria and were subjected to analysis. The pooled sensitivity and specificity were 0.52 (95% confidence interval [CI], 0.33-0.71) and 0.97 (95% CI, 0.91-0.99), respectively. The pooled positive likelihood ratio and negative likelihood ratio were 16.0 (95% CI, 5.5-46.4) and 0.50 (95% CI, 0.33-0.75), respectively. The diagnostic odds ratio was 32 (95% CI, 10-108), and the area under the summary receiver operating characteristic curve was 0.92 (95% CI, 0.90- 0.94). There was no significant publication bias observed. In the included studies, ctDNA showed clear diagnostic value for diagnosing and monitoring cervical cancer. Our meta-analysis suggested that detection of human papilloma virus ctDNA in patients with cervical cancer could be used as a noninvasive early dynamic biomarker of tumors, with high specificity and moderate sensitivity. Further large-scale prospective studies are required to validate the factors that may influence the accuracy of cervical cancer diagnosis and monitoring.

bioinformatics↗

bioRxiv2019

Variations of gut microbiome profile under different storage conditions and preservation periods: A multi-dimensional evaluation

Gut dysbiosis contributes to the development of various human diseases. There are thousands of publications per year for investigating the role of gut microbiota in development of various diseases. However, emerging evidence has indicated data inconsistency between different studies frequently, but gained very little attention by scientists. There are many factors that can cause data variation and inconsistency during the process of microbiota study, in particular, sample storage conditions and subsequent sequencing process. Here, we systemically evaluated the impacts of six fecal sample storage conditions (including -80 {degrees}C, -80 {degrees}C with 70% ethanol (ET_-80 {degrees}C), 4{degrees}C with 70% ethanol (ET_4{degrees}C), and three commercial storage reagents including OMNIgene*GUT OMR-200 (GT), MGIEasy (MGIE), and Longsee (LS)), storage periods (1, 2 weeks or 6 months), and sequencing platform on gut microbiome profile using 16S rRNA gene sequencing. Our results suggested that -80{degrees}C is acceptable for fecal sample storage, and the addition of 70% ethanol offers some benefits. Meanwhile, we found that samples in ET_4 {degrees}Cand GT reagents are comparable, both introduced multi-dimensional variations. The use of MGIE resulted in the least alteration, while the greatest changes were observed in samples stored in LS reagents during the whole experiment. Finally, we also confirmed that variations caused by storage condition were larger than that of storage time and sequencing platform.\n\nIMPORTANCEIn the current study, we performed a multi-dimensional evaluation on the variations introduced by types of storage conditions, preservation period and sequencing platform on the basis of data acquired from 16S rRNA gene sequencing. The efficacy of preservation methods was comprehensively evaluated by DNA yield and quality, and {beta} diversity, relative abundance of the dominant bacteria and functional bacteria associated with SCFAs producing and BAs metabolism. Our results confirmed that variations introduced by storage condition were larger than that of storage periods and sequencing platform. Collectively, our study provided a comprehensive view to the impacts of storage conditions, storage times, and sequencing platform on gut microbial profile.

microbiology↗