ZNF281 enhances cardiac reprogramming by modulating cardiac and inflammatory gene expression.

Direct reprogramming of fibroblasts to cardiomyocytes represents a potential means of restoring cardiac function following myocardial injury. AKT1 in the presence of four cardiogenic transcription factors, GATA4, HAND2, MEF2C, and TBX5 (AGHMT), efficiently induces the cardiac gene program in mouse embryonic fibroblasts but not adult fibroblasts. To identify additional regulators of adult cardiac reprogramming, we performed an unbiased screen of transcription factors and cytokines for those that might enhance or suppress the cardiogenic activity of AGHMT in adult mouse fibroblasts. Among a collection of inducers and repressors of cardiac reprogramming, we discovered that the zinc finger transcription factor 281 (ZNF281) potently stimulates cardiac reprogramming by genome-wide association with GATA4 on cardiac enhancers. Concomitantly, ZNF281 suppresses expression of genes associated with inflammatory signaling, suggesting the antagonistic convergence of cardiac and inflammatory transcriptional programs. Consistent with an inhibitory influence of inflammatory pathways on cardiac reprogramming, blockade of these pathways with anti-inflammatory drugs or components of the nucleosome remodeling deacetylase (NuRD) complex, which associate with ZNF281, stimulates cardiac gene expression. We conclude that ZNF281 acts at a nexus of cardiac and inflammatory gene programs, which exert opposing influences on fibroblast to cardiac reprogramming.

Improved Cardiac Function in Postischemic Rats Using an Optimized Cardiac Reprogramming Cocktail Delivered in a Single Novel Adeno-Associated Virus.

BACKGROUND: Cardiac reprogramming is a technique to directly convert nonmyocytes into myocardial cells using genes or small molecules. This intervention provides functional benefit to the rodent heart when delivered at the time of myocardial infarction or activated transgenically up to 4 weeks after myocardial infarction. Yet, several hurdles have prevented the advancement of cardiac reprogramming for clinical use. METHODS: Through a combination of screening and rational design, we identified a cardiac reprogramming cocktail that can be encoded in a single adeno-associated virus. We also created a novel adeno-associated virus capsid that can transduce cardiac fibroblasts more efficiently than available parental serotypes by mutating posttranslationally modified capsid residues. Because a constitutive promoter was needed to drive high expression of these cell fate-altering reprogramming factors, we included binding sites to a cardiomyocyte-restricted microRNA within the 3' untranslated region of the expression cassette that limits expression to nonmyocytes. After optimizing this expression cassette to reprogram human cardiac fibroblasts into induced cardiomyocyte-like cells in vitro, we also tested the ability of this capsid/cassette combination to confer functional benefit in acute mouse myocardial infarction and chronic rat myocardial infarction models. RESULTS: We demonstrated sustained, dose-dependent improvement in cardiac function when treating a rat model 2 weeks after myocardial infarction, showing that cardiac reprogramming, when delivered in a single, clinically relevant adeno-associated virus vector, can support functional improvement in the postremodeled heart. This benefit was not observed with GFP (green fluorescent protein) or a hepatocyte reprogramming cocktail and was achieved even in the presence of immunosuppression, supporting myocyte formation as the underlying mechanism. CONCLUSIONS: Collectively, these results advance the application of cardiac reprogramming gene therapy as a viable therapeutic approach to treat chronic heart failure resulting from ischemic injury.

Ultrasmall 2D Sn-Doped MAPbBr3 Nanoplatelets Enable Bright Pure-Blue Emission.

Synthesis of perovskites that exhibit pure-blue emission with high photoluminescence quantum yield (PLQY) in both nanocrystal solutions and nanocrystal-only films presents a significant challenge. In this work, a room-temperature method is developed to synthesize ultrasmall, monodispersed, Sn-doped methylammonium lead bromide (MAPb1- xSnxBr3) perovskite nanoplatelets (NPLs) in which the strong quantum confinement effect endows pure blue emission (460 nm) and a high quantum yield (87%). Post-treatment using n-hexylammonium bromide (HABr) repaired surface defects and thus substantially increased the stability and PLQY (80%) of the NPL films. Concurrently, high-precision patterned films (200-µm linewidth) are successfully fabricated by using cost-effective spray-coating technology. This research provides a novel perspective for the preparation of high PLQY, highly stable, and easily processable perovskite nanomaterials.

Boosting Reactive Oxygen Species Formation Over Pd and VOδ Co-Modified TiO2 for Methane Oxidation into Valuable Oxygenates.

Direct photocatalytic methane oxidation into value-added products provides a promising strategy for methane utilization. However, the inefficient generation of reactive oxygen species (ROS) partly limits the activation of CH4. Herein, it is reported that Pd and VOδ co-modified TiO2 enables direct and selective methane oxidation into liquid oxygenates in the presence of O2 and H2. Due to the extra ROS production from the in situ formed H2O2, a highly improved yield rate of 5014 µmol g-1 h-1 for liquid oxygenates with a selectivity of 89.3% is achieved over the optimized Pd0.5V0.2-TiO2 catalyst at ambient temperature, which is much better than those (2682 µmol g-1 h-1, 77.8%) without H2. Detailed investigations also demonstrate the synergistic effect between Pd and VOδ species for enhancing the charge carrier separation and transfer, as well as improving the catalytic activity for O2 reduction and H2O2 production.

The causal relationship between COVID-19 and seventeen common digestive diseases: a two-sample, multivariable Mendelian randomization study.

OBJECTIVES: In clinical practice, digestive symptoms such as nausea, vomiting are frequently observed in COVID-19 patients. However, the causal relationship between COVID-19 and digestive diseases remains unclear. METHODS: We extracted single nucleotide polymorphisms associated with the severity of COVID-19 from summary data of genome-wide association studies. Summary statistics of common digestive diseases were primarily obtained from the UK Biobank study and the FinnGen study. Two-sample Mendelian randomization analyses were then conducted using the inverse variance-weighted (IVW), Mendelian randomization-Egger regression (MR Egger), weighted median estimation, weighted mode, and simple mode methods. IVW served as the primary analysis method, and Multivariable Mendelian randomization analysis was employed to explore the mediating effect of body mass index (BMI) and type 2 diabetes. RESULTS: MR analysis showed that a causal association between SARS-CoV-2 infection (OR = 1.09, 95% CI 1.01-1.18, P = 0.03), severe COVID-19 (OR = 1.02, 95% CI 1.00-1.04, P = 0.02), and COVID-19 hospitalization (OR = 1.04, 95% CI 1.01-1.06, P = 0.01) with gastroesophageal reflux disease (GERD). Mediation analysis indicated that body mass index (BMI) served as the primary mediating variable in the causal relationship between SARS-CoV-2 infection and GERD, with BMI mediating 36% (95% CI 20-53%) of the effect. CONCLUSIONS: We found a causal relationship between SARS-CoV-2 infection and gastroesophageal reflux disease. Furthermore, we found that the causal relationship between SARS-CoV-2 infection and GERD is mainly mediated by BMI.

Postoperative seizure outcomes and antiseizure medication utilization based on histopathological diagnosis: A retrospective cohort study.

OBJECTIVE: Analyze the association between histopathology, seizure outcomes, and drug load of antiseizure medications (ASMs) 5-8 years after epilepsy surgery to inform preoperative decision-making and consultation. METHODS: In this retrospective, non-interventional, single-center study, patients who visited the epilepsy clinic at West China Hospital, Sichuan University from Jan 1, 2015 to Dec 31, 2020 were assessed. Patients with postoperative histopathology after epilepsy resection were included and categorized into 13 etiological groups. The primary outcomes were achieving Engel class 1 at 1, 2, 3, 5, and 8 years postoperative. Secondary outcomes included the use of ASMs and comparison of postoperative seizure outcomes between adults and children. Univariate and multivariable analyses were conducted to explore the association between clinical characteristics such as histopathology and seizure outcomes. RESULTS: A total of 315 patients were include. Patients with embryonic dysplastic neuroepithelial tumor (DNT) achieved the best seizure outcomes (84.6 % Engel class 1). DNT (odds ratio, OR=0.103, 95 %CI=0.012-0.899), cavernous hemangiomas (OR=0.140, 95 %CI=0.024-0.819) and meningioma (OR=0.137, 95 %CI=0.021-0.910) were independently associated with a higher probability of seizure-free outcome. The results of epileptic seizures in adult and pediatric groups with different pathologies were significantly different, and the preoperative and postoperative ASM dosages were also different among adult patients with various etiologies. Additionally, multivariate analysis showed that early age at onset (adjusted hazard ratio (HR) = 1.754, 95 % CI=1.049-2.934, P=0.032), late surgical age (HR=0.569, 95 %CI=0.339-0.954, P=0.032), and longer duration from seizure onset to surgery (HR=1.735, 95 % CI=1.028-2.928, P=0.039) were independent predictors of unfavorable outcomes in epileptic seizures. CONCLUSIONS: we demonstrated that the seizure outcomes of focal epilepsy have high pathological specificity, with histopathological diagnosis serving as a crucial and independent determinant of seizure outcome. Surgical assessment should be contemplated for all patients with presumed refractory focal epilepsy, irrespective of their age.

Isolation, antibacterial characterization, and alternating tangential flow-based preparation of viable cells of Lacticaseibacillus paracasei XLK 401: Potential application in milk preservation.

It is desirable to obtain high levels of viable Lacticaseibacillus paracasei, a widely used food probiotic whose antibacterial activity and potential application in milk remain largely uninvestigated. Here, we isolated and purified the L. paracasei strain XLK 401 from food-grade blueberry ferments and found that it exhibited strong antibacterial activity against both gram-positive and gram-negative foodborne pathogens, including Staphylococcus aureus, Salmonella paratyphi B, Escherichia coli O157, and Shigella flexneri. Then, we applied alternating tangential flow (ATF) technology to produce viable L. paracasei XLK 401 cells and its cell-free supernatant (CFS). Compared with the conventional fed-batch method, 22 h of ATF-based processing markedly increased the number of viable cells of L. paracasei XLK 401 to 12.14 ± 0.13 log cfu/mL. Additionally, the CFS exhibited good thermal stability and pH tolerance, inhibiting biofilm formation in the abovementioned foodborne pathogens. According to liquid chromatography-mass spectrometry analysis, organic acids were the main antibacterial components of XLK 401 CFS, accounting for its inhibition activity. Moreover, the CFS of L. paracasei XLK 401 effectively inhibited the growth of multidrug-resistant gram-positive Staph. aureus and gram-negative E. coli O157 pathogens in milk, and caused a reduction in the pathogenic cell counts by 6 to 7 log cfu/mL compared with untreated control, thus considerably maintaining the safety of milk samples. For the first time to our knowledge, ATF-based technology was employed to obtain viable L. paracasei on a large scale, and its CFS could serve as a broad-spectrum biopreservative for potential application against foodborne pathogens in milk products.

The Enhanced Performance of NiCuOOH/NiCu(OH)2 Electrode Using Pre-Conversion Treatment for the Electrochemical Oxidation of Ammonia.

Electrochemical oxidation of ammonia is an attractive process for wastewater treatment, hydrogen production, and ammonia fuel cells. However, the sluggish kinetics of the anode reaction has limited its applications, leading to a high demand for novel electrocatalysts. Herein, the electrode with the in situ growth of NiCu(OH)2 was partially transformed into the NiCuOOH phase by a pre-treatment using highly oxidative solutions. As revealed by SEM, XPS, and electrochemical analysis, such a strategy maintained the 3D structure, while inducing more active sites before the in situ generation of oxyhydroxide sites during the electrochemical reaction. The optimized NiCuOOH-1 sample exhibited the current density of 6.06 mA cm-2 at 0.5 V, which is 1.67 times higher than that of NiCu(OH)2 (3.63 mA cm-2). Moreover, the sample with a higher crystalline degree of the NiCuOOH phase exhibited lower performance, demonstrating the importance of a moderate treatment condition. In addition, the NiCuOOH-1 sample presented low selectivity (<20%) towards NO2- and stable activity during the long-term operation. The findings of this study would provide valuable insights into the development of transition metal electrocatalysts for ammonia oxidation.

Sorting of small RNAs into Arabidopsis argonaute complexes is directed by the 5' terminal nucleotide.

Argonaute (AGO) proteins recruit small RNAs to form the core of RNAi effector complexes. Arabidopsis encodes ten AGO proteins and a large network of small RNAs. How these small RNAs are sorted into specific AGO complexes remains largely unknown. We have cataloged small RNAs resident in four AGO complexes. We found that AGO2 and AGO4 preferentially recruit small RNAs with a 5' terminal adenosine, whereas AGO1 harbors microRNAs (miRNAs) that favor a 5' terminal uridine. AGO5 predominantly binds small RNAs that initiate with cytosine. Changing the 5' terminal nucleotide of an miRNA predictably redirected it into a different AGO complex and alters its biological activity. These results reveal a role for small RNA sequences in assorting among AGO complexes. This suggests that specialization of AGO complexes might involve remodeling the 5' end-binding pocket to accept certain small RNA sequences, perhaps explaining the evolutionary drive for miRNAs to initiate with uridine.

Dynamic alterations of striatal-related functional networks in juvenile absence epilepsy.

PURPOSE: To explore the features of dynamic functional connectivity (dFC) variability of striatal-cortical/subcortical networks in juvenile absence epilepsy (JAE). METHODS: We collected resting-state functional magnetic imaging data from 18 JAE patients and 28 healthy controls. The striatum was divided into six pairs of regions: the inferior-ventral striatum (VSi), superior-ventral striatum (VSs), dorsal-caudal putamen, dorsal-rostral putamen, dorsal-caudate (DC) and ventral-rostral putamen. We assessed the dFC variability of each subdivision in the whole brain using the sliding-window method, and correlated altered circuit with clinical variables in JAE patients. RESULTS: We found altered dFC variability of striatal-cortical/subcortical networks in patients with JAE. The VSs exhibited decreased dFC variability with subcortical regions, and dFC variability between VSs and thalamus was negatively correlated with epilepsy duration. For the striatal-cortical networks, the dFC variability was decreased in VSi-affective network but increased in DC-executive network. The altered dynamics of striatal-cortical networks involved crucial nodes of the default mode network (DMN). CONCLUSION: JAE patients exhibit excessive stability in the striatal-subcortical networks. For striatal-cortical networks in JAE, the striatal-affective circuit was more stable, while the striatal-executive circuit was more variable. Furthermore, crucial nodes of DMN were changed in striatal-cortical networks in JAE.

Association of ß-arrestin1 and p53-Mdm2 signaling in the development of missed abortion.

BACKGROUND: Missed abortion is a peculiar form of spontaneous abortion before 20 weeks' gestation. The definite etiology and pathogenesis are not fully understood. Recent studies have demonstrated that p53/Mdm2-mediated ubiquitination of the IGF-1R may be closely related to G-protein-coupled receptor kinases (GRK)/ß-arrestin1 system. Our previous studies have confirmed that the elevated expression of p53 and Mdm2 may be responsible for apoptosis during missed abortion. However, there was no information surrounding ß-arrestin1 in missed abortion. METHODS: The mRNA levels of ß-arrestin1 in villous samples of 30 missed abortion patients and 31 healthy controls were determined by real-time quantitative polymerase chain reaction (PCR). Immunohistochemistry was used to explore the expression and location of ß-arrestin1, p53, Mdm2, VEGF and HIF-lα in trophoblasts. Transwell assays were performed to examine the influences of ß-arrestin1 expression on cell invasion. Furthermore, we tested the effect of ß-arrestin1 on the expression of p53, Mdm2, ERK, AKT and NF-κB. RESULTS: The expression of ß-arrestin1 in the villous samples of missed abortion group was dramatically lower than control group by quantitative real-time-PCR and immunohistochemistry. Furthermore, the patients with missed abortion showed significantly higher levels of p53, Mdm2, HIF-lα and lower level of VEGF than healthy controls by immunohistochemistry. Functional studies showed that suppression of ß-arrestin1 in HTR-8 cells inhibited cell invasion. The protein expressions of ERK and AKT in HTR-8 cells were significantly downregulated by reducing the expression of ß-arrestin1, while the expressions of p53, Mdm2, NF-κB were enhanced. Overexpression of ß-arrestin1 exhibited the adverse effect. CONCLUSION: Our data indicated that ß-arrestin1 play an important role in maintaining the maternal-fetal tolerance, the decreased expression of ß-arrestin1 in the villous samples may be related with the development of missed abortion.

[Effects of Bupleurum chinense on the diversity of intestinal flora in depressed mice].

Bupleuri Radix, serving as the sovereign medicinal in many antidepressant compound preparations, has been proved effective in treating depression in mice, but its effect on the intestinal flora remains unclear. The present study aimed to investigate the effects of Bupleurum chinense(one of the original materials of Bupleuri Radix) on the behaviors and the diversity of intestinal flora of depressed mice. A depression mouse model was induced by repeated social defeat stress. Specifically, C57 BL/6 J male mice were exposed to the attack from the CD-1 mice. Then, C57 BL/6 J male mice were divided into a depression group and a B. chinense group, with normal saline and B. chinense administered(ig) respectively. Sucrose preference test and tail suspension test were conducted during and after the experiment respectively, to analyze the effects of B. chinense on the behaviors of the depressed mice. The feces were collected after the experiment. The V3-V4 16 S rDNA regions of intestinal flora of mice in each group were sequenced by Ion S5 TMXL for the analysis of the number of operational taxonomic units(OTUs), richness, alpha and beta diversity indexes, and differential phyla and genera. The results indicated that B. chinense could decrease depressive-like behaviors of mice, increase sucrose preference, and shorten the time of immobility in tail suspension test. After B. chinense intervention, the relative abundance of Firmicutes was significantly decreased, while that of Bacteroidetes was increased at the phylum level. At the genus level, the relative abundance of Lactobacillus and Lachnoclostridium decreased(P<0.05), while that of Bacteroides, Alistopes, etc. was elevated(P<0.05). The findings demonstrate that B. chinense can regulate the intestinal flora and improve the depressive-like behaviors of mice with depression.

Blockade to pathological remodeling of infarcted heart tissue using a porcupine antagonist.

The secreted Wnt signaling molecules are essential to the coordination of cell-fate decision making in multicellular organisms. In adult animals, the secreted Wnt proteins are critical for tissue regeneration and frequently contribute to cancer. Small molecules that disable the Wnt acyltransferase Porcupine (Porcn) are candidate anticancer agents in clinical testing. Here we have systematically assessed the effects of the Porcn inhibitor (WNT-974) on the regeneration of several tissue types to identify potentially unwanted chemical effects that could limit the therapeutic utility of such agents. An unanticipated observation from these studies is proregenerative responses in heart muscle induced by systemic chemical suppression of Wnt signaling. Using in vitro cultures of several cell types found in the heart, we delineate the Wnt signaling apparatus supporting an antiregenerative transcriptional program that includes a subunit of the nonfibrillar collagen VI. Similar to observations seen in animals exposed to WNT-974, deletion of the collagen VI subunit, COL6A1, has been shown to decrease aberrant remodeling and fibrosis in infarcted heart tissue. We demonstrate that WNT-974 can improve the recovery of heart function after left anterior descending coronary artery ligation by mitigating adverse remodeling of infarcted tissue. Injured heart tissue exposed to WNT-974 exhibits decreased scarring and reduced Col6 production. Our findings support the development of Porcn inhibitors as antifibrotic agents that could be exploited to promote heart repair following injury.

Fluorimetric Detection of Candida albicans Using Cornstalk N-Carbon Quantum Dots Modified with Amphotericin B.

Due to the advanced fluorescence property of N-carbon quantum dots (N-CQDs), a new method to detect pathogenic fungi by newly synthesized cornstalk N-CQDs modified with water-soluble amphotericin B (N-CQDs@AmpB) was developed. Specifically, N-CQDs with blue fluorescence were initially synthesized according to a previous report and modified with amphotericin B on their surfaces. Subsequently, the as-prepared N-CQDs@AmpB was used to detect Candida albicans, exhibiting a linear range of 2.60 × 105 to 1.99 × 108 cfu/mL and a detection limit of 1124 cfu/mL. Compared with other common methods, the method largely shortened the detection time and enabled the process to be performed with minimal interference from complex samples such as beef sausage. The high cost of water-soluble amphotericin B may hamper the large-scale application of the new detection method using N-CQDs@AmpB. Thus, alcohol-soluble amphotericin B was used in subsequent experiments, confirming its potential to broaden avenues for the detection of fungi.

Long Noncoding RNA NEAT1 Promotes Myocardiocyte Apoptosis and Suppresses Proliferation Through Regulation of miR-129-5p.

Recent studies have revealed the important role of long noncoding RNAs (lncRNAs) in heart development and pathogenesis. This study was aimed to investigate the role of NEAT1 in hypoxia-induced cardiac injury and explore its possible molecular mechanism. Real-time PCR (RT-PCR) was used to determine the relative RNA expression of NEAT1 and its potential target microRNA, miR-129-5p, in the plasma of patients with acute myocardial infarction, heart failure, and angina, as well as in H2O2-treated H9c2 cells. The role of NEAT1 overexpression or inhibition in H9c2 cell migration and proliferation was assessed by transwell assay and Edu staining, respectively. Collagen deposition and apoptosis were evaluated by Western blot detection of collagen and apoptotic proteins, including Capase3, Bax, and Bcl2. We showed that H2O2 treatment significantly decreased H9c2 cell migration and proliferation while increasing H9c2 cell apoptosis. Inhibition of NEAT1 attenuated the cell apoptosis and alleviated proliferation inhibition induced by hypoxia. Bioinformatics analysis showed that miR-129-5p was the direct target of NEAT1, which was confirmed by luciferase assay. NEAT1 upregulation aggravated apoptosis by downregulating miR-129-5p. In conclusion, we uncovered a novel NEAT1-miR-129 axis and its implication in H2O2-induced heart failure.

DNA methylation mediated by a microRNA pathway.

In plants, the known microRNAs (miRNAs) are produced as approximately 21 nucleotide (nt) duplexes from their precursors by Dicer-like 1 (DCL1). They are incorporated into Argonaute 1 (AGO1) protein to regulate target gene expression primarily through mRNA cleavage. We report here the discovery of a class of miRNAs in the model monocot rice (Oryza sativa). These are 24 nt in length and require another member of the Dicer family, DCL3, for their biogenesis. The 24 nt long miRNAs (lmiRNAs) are loaded into AGO4 clade proteins according to hierarchical rules, depending on the upstream biogenesis machinery and the 5'-terminal nucleotide. We demonstrated that lmiRNAs direct DNA methylation at loci from which they are produced as well as in trans at their target genes and play roles in gene regulation. Considered together, our findings define a miRNA pathway that mediates DNA methylation.

Akt1/protein kinase B enhances transcriptional reprogramming of fibroblasts to functional cardiomyocytes.

Conversion of fibroblasts to functional cardiomyocytes represents a potential approach for restoring cardiac function after myocardial injury, but the technique thus far has been slow and inefficient. To improve the efficiency of reprogramming fibroblasts to cardiac-like myocytes (iCMs) by cardiac transcription factors [Gata4, Hand2, Mef2c, and Tbx5 (GHMT)], we screened 192 protein kinases and discovered that Akt/protein kinase B dramatically accelerates and amplifies this process in three different types of fibroblasts (mouse embryo, adult cardiac, and tail tip). Approximately 50% of reprogrammed mouse embryo fibroblasts displayed spontaneous beating after 3 wk of induction by Akt plus GHMT. Furthermore, addition of Akt1 to GHMT evoked a more mature cardiac phenotype for iCMs, as seen by enhanced polynucleation, cellular hypertrophy, gene expression, and metabolic reprogramming. Insulin-like growth factor 1 (IGF1) and phosphoinositol 3-kinase (PI3K) acted upstream of Akt whereas the mitochondrial target of rapamycin complex 1 (mTORC1) and forkhead box o3 (Foxo3a) acted downstream of Akt to influence fibroblast-to-cardiomyocyte reprogramming. These findings provide insights into the molecular basis of cardiac reprogramming and represent an important step toward further application of this technique.

Investigation of the ischioanal fossa: Application to abscess spread.

The location of perianal abscesses and the course of the fistula follow certain patterns, especially in the relationship between external and internal openings. However, it is still not clear how the contents of the ischioanal fossa, especially the fibrous network of fat tissue, affect the route for such diseases. Ten male adult cadavers were selected for the study. Seven horizontal transverse section planes from 1 cm above the pubic symphysis to the inferior border of the lesser trochanter of the femur were recorded after P45 sheet plastination. We observed characteristics of fiber distribution in the ischioanal fossa and its relationship with surrounding structures in every plane. There was a dense strip-type fiber connecting with junction fascia between the obturator internus and gluteus maximus muscles. Close to the levator ani, obturator internus, and gluteus maximus, the fibers were very dense and continuous with the fascia on the surfaces of these three muscles. The function of the fibrous network was considered to be not only the support of fat tissue in the fossa but also cushioning during physiological actions such as defecation. We hope that these morphological results could help to elucidate the passage of fistulae and the locations susceptible to perianal abscesses. Clin. Anat. 30:1029-1033, 2017. © 2017 Wiley Periodicals, Inc.

Exploration of potential drug targets for Glaucoma by plasma proteome screening.

BACKGROUND: Glaucoma is the leading cause of irreversible blindness. However, the current available treatment methods are still unsatisfactory. Therefore, the exploration of new drug targets for the treatment of glaucoma is of paramount importance. METHODS: We conducted two-sample Mendelian randomization (MR) using plasma protein quantitative trait loci (pQTL) data from two datasets (n = 734, n = 4907) and their instrumental variables to investigate the causal relationship between plasma proteins and glaucoma. The analysis was validated by replacing the exposure and outcome cohorts. Additionally, we utilized protein-protein interaction networks to assess the associations between these potential drug targets and existing drug targets. RESULTS: Through two-sample Mendelian randomization analysis, we identified causal relationships between Glaucoma and the following proteins: AZU1, OBP2B, ENPP5, INPP5B, KREMEN1, LYPLAL1, and PTPRJ. External validation confirmed the protective effect of LYPLAL1 on Glaucoma, while ENPP5, KREMEN1, and PTPRJ increased the risk of Glaucoma. Reverse MR and Steiger filtering did not indicate any reverse causal associations of the aforementioned proteins with Glaucoma. CONCLUSION: Our study demonstrates a causal impact of ENPP5, KREMEN1, PTPRJ, and LYPLAL1 on the risk of Glaucoma. These findings suggest that these four proteins may serve as promising drug targets for Glaucoma treatment. SIGNIFICANCE: Currently, the pharmacological treatment of glaucoma primarily focuses on lowering intraocular pressure, which has its limitations. Targeted therapy is a personalized treatment approach that aims to inhibit or block the development and progression of diseases such as cancer and inflammation by selectively acting on specific biomolecules or signaling pathways. Our research employs a two-sample Mendelian randomization (MR) method, integrating a large amount of GWAS and pQTL data to perform MR analysis. This has enabled us to explore several plasma proteins as potential drug targets for glaucoma, providing direction and a research foundation for future investigations into glaucoma drug targets.

Efficient and markerless gene integration with SlugCas9-HF in Kluyveromyces marxianus.

The nonconventional yeast Kluyveromyces marxianus has potential for industrial production, but the lack of advanced synthetic biology tools for precise engineering hinders its rapid development. Here, we introduce a CRISPR-Cas9-mediated multilocus integration method for assembling multiple exogenous genes. Using SlugCas9-HF, a high-fidelity Cas9 nuclease, we enhance gene editing precision. Specific genomic loci predisposed to efficient integration and expression of heterologous genes are identified and combined with a set of paired CRISPR-Cas9 expression plasmids and donor plasmids to establish a CRISPR-based biosynthesis toolkit. This toolkit enables genome integration of large gene modules over 12 kb and achieves simultaneous quadruple-locus integration in a single step with 20% efficiency. As a proof-of-concept, we apply the toolkit to screen for gene combinations that promote heme production, revealing the importance of HEM4Km and HEM12Sc. This CRISPR-based toolkit simplifies the reconstruction of complex pathways in K. marxianus, broadening its application in synthetic biology.