Identification of prognostic biomarkers for cholangiocarcinoma by combined analysis of molecular characteristics of clinical MVI subtypes and molecular subtypes.

Cholangiocarcinoma (CCA) is widely noted for its high degree of malignancy, rapid progression, and limited therapeutic options. This study was carried out on transcriptome data of 417 CCA samples from different anatomical locations. The effects of lipid metabolism related genes and immune related genes as CCA classifiers were compared. Key genes were derived from MVI subtypes and better molecular subtypes. Pathways such as epithelial mesenchymal transition (EMT) and cell cycle were significantly activated in MVI-positive group. CCA patients were classified into three (four) subtypes based on lipid metabolism (immune) related genes, with better prognosis observed in lipid metabolism-C1, immune-C2, and immune-C4. IPTW analysis found that the prognosis of lipid metabolism-C1 was significantly better than that of lipid metabolism-C2 + C3 before and after correction. KRT16 was finally selected as the key gene. And knockdown of KRT16 inhibited proliferation, migration and invasion of CCA cells.

The end of court-ordered desegregation and U.S. children's health: Quasi-experimental evidence.

School racial segregation significantly impacts racial disparities in U.S. children's health. Recently, school segregation has been increasing, partially due to Supreme Court decisions since 1991 that have made it easier for school districts to be released from court-ordered desegregation. We investigated the association of the end of court-ordered desegregation with child health using the 1997-2018 waves of the National Health Interview Survey (N=8,182 Black, 16,930 White children). We exploited quasi-random variation in the timing of school districts' releases from court orders to estimate effects on general health, body weight, mental health, and asthma, using difference-in-differences and event-study methods (including traditional and heterogeneity-robust estimators). Heterogeneity-robust difference-in-differences analyses show that release was associated with increased school segregation, improved mental health among Black children, and better self-reported health among White children. For heterogeneity-robust event-study analyses, school segregation increased steadily over time after release, with worse self-reported health and higher risk of asthma episodes among Black children 18+ years after release. Black children's mental health temporarily improved in the short term. In contrast, White children had improved self-reported health, mental health, and risk of asthma episodes in some years. Interventions to address the harms of school segregation are important for reducing racial health inequities.

Advances in difference-in-differences methods for policy evaluation research.

Difference-in-differences (DiD) is a powerful, quasi-experimental research design widely used in longitudinal policy evaluations with health outcomes. However, DiD designs face several challenges to ensuring reliable causal inference, such as when policy settings are more complex. Recent economics literature has revealed that DiD estimators may exhibit bias when heterogeneous treatment effects, a common consequence of staggered policy implementation, are present. To deepen understanding of these advancements in epidemiology, in this methodologic primer, we start by presenting an overview of DiD methods. We then summarize fundamental problems associated with DiD designs with heterogeneous treatment effects and provide guidance on recently proposed heterogeneity-robust DiD estimators, which are increasingly being implemented by epidemiologists. We also extend the discussion to violations of the parallel trends assumption, which has received less attention. Last, we present results from a simulation study that compares the performance of several DiD estimators under different scenarios to enhance understanding and application of these methods.

Tumor-suppressive zinc finger protein 24 (ZNF24) sensitizes colorectal cancer cells to 5-fluorouracil by inhibiting the Wnt pathway and activating the p53 signaling.

Carcinogenesis and colorectal cancer (CRC) development are associated with dysregulation of various pathways, including Wnt and p53. 5-fluorouracil (5-FU) is a common chemotherapeutic agent for CRC treatment, but its efficacy is restricted by drug resistance. Doxycycline is an orally active tetracycline antibiotic known for its antimicrobial and anticancer cell proliferation activities. This study intends to delineate the potential role of bioinformatically predicted ZNF24 in the 5-FU resistance of CRC cells. The expression of ZNF24 was measured in clinically collected CRC tissues and cells. Afterward, ectopic ZNF24 expression was induced by DOX to evaluate the viability, colony-forming ability and sphere-forming ability of CRC cells. It was found that ZNF24 was validated to be poorly expressed in CRC tissues, and ectopic expression of ZNF24 was revealed to restrict the malignant phenotypes of CRC cells. In addition, restored ZNF24 attenuated 5-FU resistance of CRC cells by inhibiting the Wnt pathway and activating p53 signaling. Furthermore, an inhibitor of Wnt production 2 (IWP-2) treatment was an alternative to ZNF24 up-regulation in sensitizing CRC cells to 5-FU treatment. In conclusion, our results indicate that ZNF24 inhibits 5-FU resistance of CRC cells by suppressing the Wnt pathway and activating p53 signaling, which offers a potential strategy for managing chemoresistance in CRC.

The maize WRKY transcription factor ZmWRKY64 confers cadmium tolerance in Arabidopsis and maize (Zea mays L.).

KEY MESSAGE: ZmWRKY64 positively regulates Arabidopsis and maize Cd stress through modulating Cd uptake, translocation, and ROS scavenging genes expression. Cadmium (Cd) is a highly toxic heavy metal with severe impacts on crops growth and development. The WRKY transcription factor is a significant regulator influencing plant stress response. Nevertheless, the function of the WRKY protein in maize Cd stress response remains unclear. Here, we identified a maize WRKY gene, ZmWRKY64, the expression of which was enhanced in maize roots and leaves under Cd stress. ZmWRKY64 was localized in the nucleus and displayed transcriptional activity in yeast. Heterologous expression of ZmWRKY64 in Arabidopsis diminished Cd accumulation in plants by negatively regulating the expression of AtIRT1, AtZIP1, AtHMA2, AtNRAMP3, and AtNRAMP4, which are involved in Cd uptake and transport, resulting in Cd stress tolerance. Knockdown of ZmWRKY64 in maize led to excessive Cd accumulation in leaf cells and in the cytosol of the root cells, resulting in a Cd hypersensitive phenotype. Further analysis confirmed that ZmWRKY64 positively regulated ZmABCC4, ZmHMA3, ZmNRAMP5, ZmPIN2, ZmABCG51, ZmABCB13/32, and ZmABCB10, which may influence Cd translocation and auxin transport, thus mitigating Cd toxicity in maize. Moreover, ZmWRKY64 could directly enhance the transcription of ZmSRG7, a reported key gene regulating reactive oxygen species homeostasis under abiotic stress. Our results indicate that ZmWRKY64 is important in maize Cd stress response. This work provides new insights into the WRKY transcription factor regulatory mechanism under a Cd-polluted environment and may lead to the genetic improvement of Cd tolerance in maize.

Natural antioxidants enhance the oxidation stability of blended oils enriched in unsaturated fatty acids.

BACKGROUND: Rancidity causes unpleasant tastes and smells, and the degradation of fatty acids and natural antioxidants, so that an oil is unfit to be consumed. Natural antioxidants, including tocopherols, polyphenols (sesamol, canolol, ferulic acid, caffeic acid, etc.), ß-carotene, squalene and phytosterols, contribute to delay the oxidation of vegetable oils. However, studies on the combination of natural antioxidants to lengthen the shelf life of unsaturated fatty acid-rich blended oil have not been reported. RESULTS: All of the composite antioxidants had the potential to significantly improve the oxidation stability of blended oil. Blended oil G with 0.05 g kg-1 ß-carotene, 0.25 g kg-1 sesamol and 0.25 g kg-1 caffeic acid showed the best anti-autooxidation. It is also effective in improving the oxidative stability of vegetable oils containing various fatty acids. The oxidation stability index of the blended oil containing the optimum composition of natural antioxidants was 2.17-fold longer than that of the control sample. After the end of accelerated oxidation, the oil's peroxide value, p-anisidine value and total oxidation value were 6.59 times, 12.26 times and 6.65 times lower than those of the control sample, respectively. CONCLUSION: (1) The combination of natural antioxidants ß-carotene (0.05 g kg-1 ), sesamol (0.25 g kg-1 ) and caffeic acid (0.25 g kg-1 ) enhances the oxidative stability of unsaturated fatty acid-rich blended oils. (2) ß-Carotene is the main antioxidant in the early stages of oxidation. (3) Sesamol and caffeic acid are the main antioxidants in the middle and late stages of oxidation. © 2023 Society of Chemical Industry.

From clinical variables to multiomics analysis: a margin morphology-based gross classification system for hepatocellular carcinoma stratification.

OBJECTIVE: Selecting interventions for patients with solitary hepatocellular carcinoma (HCC) remains a challenge. Despite gross classification being proposed as a potential prognostic predictor, its widespread use has been restricted due to inadequate studies with sufficient patient numbers and the lack of established mechanisms. We sought to investigate the prognostic impacts on patients with HCC of different gross subtypes and assess their corresponding molecular landscapes. DESIGN: A prospective cohort of 400 patients who underwent hepatic resection for solitary HCC was reviewed and analysed and gross classification was assessed. Multiomics analyses were performed on tumours and non-tumour tissues from 49 patients to investigate the mechanisms underlying gross classification. Inverse probability of treatment weight (IPTW) was used to control for confounding factors. RESULTS: Overall 3-year survival rates varied significantly among the four gross subtypes (type I: 91%, type II: 80%, type III: 74.6%, type IV: 38.8%). Type IV was found to be independently associated with poor prognosis in both the entire cohort and the IPTW cohort. The four gross subtypes exhibited three distinct transcriptional modules. Particularly, type IV tumours exhibited increased angiogenesis and immune score as well as decreased metabolic pathways, together with highest frequency of TP53 mutations. Patients with type IV HCC may benefit from adjuvant intra-arterial therapy other than the other three subtypes. Accordingly, a modified trichotomous margin morphological gross classification was established. CONCLUSION: Different gross types of HCC showed significantly different prognosis and molecular characteristics. Gross classification may aid in development of precise individualised diagnosis and treatment strategies for HCC.

Low dissolved oxygen supply functions as a global regulator of the growth and metabolism of Aurantiochytrium sp. PKU#Mn16 in the early stages of docosahexaenoic acid fermentation.

BACKGROUND: Thraustochytrids accumulate lipids with a high content of docosahexaenoic acid (DHA). Although their growth and DHA content are significantly affected by the dissolved oxygen (DO) supply, the role of DO on the transcriptional regulation of metabolism and accumulation of intracellular metabolites remains poorly understood. Here we investigate the effects of three different DO supply conditions (10%, 30%, and 50%) on the fed-batch culture of the Aurantiochytrium PKU#Mn16 strain to mainly reveal the differential gene expressions and metabolite profiles. RESULTS: While the supply of 10% DO significantly reduced the rates of biomass and DHA production in the early stages of fermentation, it achieved the highest amounts of biomass (56.7 g/L) and DHA (6.0 g/L) on prolonged fermentation. The transcriptome analyses of the early stage (24 h) of fermentation revealed several genes involved in the central carbon, amino acid, and fatty acid metabolism, which were significantly downregulated at a 10% DO level. The comparative metabolomics results revealed the accumulation of several long-chain fatty acids, amino acids, and other metabolites, supporting the transcriptional regulation under the influence of a low oxygen supply condition. In addition, certain genes involved in antioxidative systems were downregulated under 10% DO level, suggesting a lesser generation of reactive oxygen species that lead to oxidative damage and fatty acid oxidation. CONCLUSIONS: The findings of this study suggest that despite the slow growth and metabolism in the early stage of fermentation of Aurantiochytrium sp. PKU#Mn16, a constant supply of low dissolved oxygen can yield biomass and DHA content better than that with high oxygen supply conditions. The critical information gained in this study will help to further improve DHA production through bioprocess engineering strategies.

Saturated and Polyunsaturated Fatty Acids Production by Aurantiochytrium limacinum PKU#Mn4 on Enteromorpha Hydrolysate.

Thraustochytrids are unicellular marine heterotrophic protists, which have recently shown a promising ability to produce omega-3 fatty acids from lignocellulosic hydrolysates and wastewaters. Here we studied the biorefinery potential of the dilute acid-pretreated marine macroalgae (Enteromorpha) in comparison with glucose via fermentation using a previously isolated thraustochytrid strain (Aurantiochytrium limacinum PKU#Mn4). The total reducing sugars in the Enteromorpha hydrolysate accounted for 43.93% of the dry cell weight (DCW). The strain was capable of producing the highest DCW (4.32 ± 0.09 g/L) and total fatty acids (TFA) content (0.65 ± 0.03 g/L) in the medium containing 100 g/L of hydrolysate. The maximum TFA yields of 0.164 ± 0.160 g/g DCW and 0.196 ± 0.010 g/g DCW were achieved at 80 g/L of hydrolysate and 40 g/L of glucose in the fermentation medium, respectively. Compositional analysis of TFA revealed the production of equivalent fractions (% TFA) of saturated and polyunsaturated fatty acids in hydrolysate or glucose medium. Furthermore, the strain yielded a much higher fraction (2.61-3.22%) of eicosapentaenoic acid (C20:5n-3) in the hydrolysate medium than that (0.25-0.49%) in the glucose medium. Overall, our findings suggest that Enteromorpha hydrolysate can be a potential natural substrate in the fermentative production of high-value fatty acids by thraustochytrids.

Transcriptomic analysis reveals key genes and pathways corresponding to Cd and Pb in the hyperaccumulator Arabis paniculata.

Soil and water are increasingly at risk of contamination from the toxic heavy metals lead (Pb) and cadmium (Cd). Arabis paniculata (Brassicaceae) is a hyperaccumulator of heavy metals (HMs) found widely distributed in areas impacts by mining activities. However, the mechanism by which A. paniculata tolerates HMs is still uncharacterized. For this experiment, we employed RNA sequencing (RNA-seq) in order to find Cd (0.25 mM)- and Pb (2.50 mM)-coresponsive genes A. paniculata. In total, 4490 and 1804 differentially expressed genes (DEGs) were identified in root tissue, and 955 and 2209 DEGs were identified in shoot tissue, after Cd and Pb exposure, respectively. Interestingly in root tissue, gene expression corresponded similarly to both Cd and Pd exposure, of which 27.48% were co-upregulated and 41.00% were co-downregulated. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses showed that the co-regulated genes were predominantly involved in transcription factors (TFs), cell wall biosynthesis, metal transport, plant hormone signal transduction, and antioxidant enzyme activity. Many critical Pb/Cd-induced DEGs involved in phytohormone biosynthesis and signal transduction, HM transport, and transcription factors were also identified. Especially the gene ABCC9 was co-downregulated in root tissues but co-upregulated in shoot tissues. The co-downregulation of ABCC9 in the roots prevented Cd and Pb from entering the vacuole rather than the cytoplasm for transporting HMs to shoots. While in shoots, the ABCC9 co-upregulated results in vacuolar Cd and Pb accumulation, which may explain why A. paniculata is a hyperaccumulator. These results will help to reveal the molecular and physiological processes underlying tolerance to HM exposure in the hyperaccumulator A. paniculata, and aid in future efforts to utilize this plant in phytoremediation.

Biphasic action potential and chaos in a symmetrical Chua Corsage Memristor-based circuit.

Neuromorphic computing provides unique computing and memory capabilities that could break the limitation of conventional von Neumann computing. Toward realizing neuromorphic computing, fabrication and synthetization of hardware elements and circuits to emulate biological neurons are crucial. Despite the striking progress in exploring neuron circuits, the existing circuits can only reproduce monophasic action potentials, and no studies report on circuits that could emulate biphasic action potentials, limiting the development of neuromorphic devices. Here, we present a simple third-order memristive circuit built with a classical symmetrical Chua Corsage Memristor (SCCM) to accurately emulate biological neurons and show that the circuit can reproduce monophasic action potentials, biphasic action potentials, and chaos. Applying the edge of chaos criterion, we calculate that the SCCM and the proposed circuit have the symmetrical edge of chaos domains with respect to the origin, which plays an important role in generating biphasic action potentials. Also, we draw a parameter classification map of the proposed circuit, showing the edge of chaos domain (EOCD), the locally active domain, and the locally passive domain. Near the calculated EOCD, the third-order circuit generates monophasic action potentials, biphasic action potentials, chaos, and ten types of symmetrical bi-directional neuromorphic phenomena by only tuning the input voltage, showing a resemblance to biological neurons. Finally, a physical SCCM circuit and some experimentally measured neuromorphic waveforms are exhibited. The experimental results agree with the numerical simulations, verifying that the proposed circuit is suitable as artificial neurons.

A negative capacitor emulator circuit and its application in chaotic circuits.

Negative capacitance usually exists in ferroelectric materials, and its instinct nonlinearity and negative capacitance render its potential applications. To date, the single negative capacitance device is commonly unavailable. Hence, it is necessary to build a negative capacitor emulator in hardware for further investigating its electrical characteristics and applications. Based on a simple negative capacitor mathematical model, an emulator circuit that can mimic the S-shaped voltage-charge characteristics of the negative capacitor is proposed. The proposed emulator is composed of commercial off-the-shelf components, such as operational amplifiers, resistors, and capacitors. Based on the negative capacitor, we design a new chaotic circuit that can generate the single-period, double-period, single-scroll chaos, double-scroll chaos, and so on. The theoretical calculation, simulation analysis, and hardware experimental verification manifest that this proposed emulator circuit can be operated as a negative capacitor and applied in the chaotic circuit.

School racial segregation and long-term cardiovascular health among Black adults in the US: A quasi-experimental study.

BACKGROUND: Cardiovascular disease (CVD) disproportionately affects Black adults in the United States. This is increasingly acknowledged to be due to inequitable distribution of health-promoting resources. One potential contributor is inequities in educational opportunities, although it is unclear what aspects of education are most salient. School racial segregation may affect cardiovascular health by increasing stress, constraining socioeconomic opportunities, and altering health behaviors. We investigated the association between school segregation and Black adults' CVD risk. METHODS AND FINDINGS: We leveraged a natural experiment created by quasi-random (i.e., arbitrary) timing of local court decisions since 1991 that released school districts from court-ordered desegregation. We used the Panel Study of Income Dynamics (PSID) (1991 to 2017), linked with district-level school segregation measures and desegregation court order status. The sample included 1,053 Black participants who ever resided in school districts that were under a court desegregation order in 1991. The exposure was mean school segregation during observed schooling years. Outcomes included several adult CVD risk factors and outcomes. We fitted standard ordinary least squares (OLS) multivariable linear regression models, then conducted instrumental variables (IV) analysis, using the proportion of schooling years spent in districts that had been released from court-ordered desegregation as an instrument. We adjusted for individual- and district-level preexposure confounders, birth year, and state fixed effects. In standard linear models, school segregation was associated with a lower probability of good self-rated health (-0.05 percentage points per SD of the segregation index; 95% CI: -0.08, -0.03; p < 0.001) and a higher probability of binge drinking (0.04 percentage points; 95% CI: 0.002, 0.07; p = 0.04) and heart disease (0.01 percentage points; 95% CI: 0.002, 0.15; p = 0.007). IV analyses also found that school segregation was associated with a lower probability of good self-rated health (-0.09 percentage points; 95% CI: -0.17, -0.02, p = 0.02) and a higher probability of binge drinking (0.17 percentage points; 95% CI: 0.04, 0.30, p = 0.008). For IV estimates, only binge drinking was robust to adjustments for multiple hypothesis testing. Limitations included self-reported outcomes and potential residual confounding and exposure misclassification. CONCLUSIONS: School segregation exposure in childhood may have longstanding impacts on Black adults' cardiovascular health. Future research should replicate these analyses in larger samples and explore potential mechanisms. Given the recent rise in school segregation, this study has implications for policies and programs to address racial inequities in CVD.

Vertical community patterns of Labyrinthulomycetes protists reveal their potential importance in the oceanic biological pump.

The biological pump plays a vital role in exporting organic particles into the deep ocean for long-term carbon sequestration. However, much remains unknown about some of its key microbial players. In this study, Labyrinthulomycetes protists (LP) were used to understand the significance of heterotrophic microeukaryotes in the transport of particulate organic matter from the surface to the dark ocean. Unlike the sharp vertical decrease of prokaryotic biomass, the LP biomass only slightly decreased with depth and eventually exceeded prokaryotic biomass in the bathypelagic layer. Sequencing identified high diversity of the LP communities with a dominance of Aplanochytrium at all depths. Notably, ASVs that were observed in the surface layer comprised ~20% of ASVs and ~60% of sequences in each of the deeper (including bathypelagic) layers, suggesting potential vertical export of the LP populations to the deep ocean. Further analyses of the vertical patterns of the 50 most abundant ASVs revealed niche partitioning of LP phylotypes in the pelagic ocean, including those that could decompose organic detritus and/or facilitate the formation of fast-sinking particles. Overall, this study presents several lines of evidence that the LP can be an important component of the biological pump through their multiple ecotypes in the pelagic ocean.

De novo production of versatile oxidized kaurene diterpenes in Escherichia coli.

The oxidized kaurene (Ox-Kau) compounds are the core structures of many important diterpenoids with biological activities and economical values. However, easy access to diverse Ox-Kau products is still limited by low natural abundance, and large-scale manufacture remain challenging due to lack of proper heterologous production. To achieve an abundant source alternative to natural extracts, we here report a highly effective Escherichia coli-based platform for the de novo production of multiple Ox-Kau molecules from simple carbon source. Pathway optimization in prokaryotic cells through modification of transmembrane CYP450 oxidases, cytochrome b5 co-expression and AlphaFold-based protein engineering improved a 50-fold yield of steviol (1.07 g L-1), a key intermediate in the kaurenoid biosynthesis. Combinatorial biosynthetic strategy further led to a series of oxidized derivatives (20-600 mg L-1) with rich oxygenated functional groups on C3, C7, C16 and C19 previously hard to be introduced. Our engineered strains not only laid a foundation for realizing the industrial fermentation of gram-scale ent-kaurene diterpenoids, but also provided a reliable platform for characterization and utilization of kaurene-modifying oxidases, which may generate naturally rare or unnatural ent-kaurenoids with potential bioactivity.

Development and validation of a preoperative CT-based radiomic nomogram to predict pathology invasiveness in patients with a solitary pulmonary nodule: a machine learning approach, multicenter, diagnostic study.

OBJECTIVES: To develop and validate a preoperative CT-based nomogram combined with radiomic and clinical-radiological signatures to distinguish preinvasive lesions from pulmonary invasive lesions. METHODS: This was a retrospective, diagnostic study conducted from August 1, 2018, to May 1, 2020, at three centers. Patients with a solitary pulmonary nodule were enrolled in the GDPH center and were divided into two groups (7:3) randomly: development (n = 149) and internal validation (n = 54). The SYSMH center and the ZSLC Center formed an external validation cohort of 170 patients. The least absolute shrinkage and selection operator (LASSO) algorithm and logistic regression analysis were used to feature signatures and transform them into models. RESULTS: The study comprised 373 individuals from three independent centers (female: 225/373, 60.3%; median [IQR] age, 57.0 [48.0-65.0] years). The AUCs for the combined radiomic signature selected from the nodular area and the perinodular area were 0.93, 0.91, and 0.90 in the three cohorts. The nomogram combining the clinical and combined radiomic signatures could accurately predict interstitial invasion in patients with a solitary pulmonary nodule (AUC, 0.94, 0.90, 0.92) in the three cohorts, respectively. The radiomic nomogram outperformed any clinical or radiomic signature in terms of clinical predictive abilities, according to a decision curve analysis and the Akaike information criteria. CONCLUSIONS: This study demonstrated that a nomogram constructed by identified clinical-radiological signatures and combined radiomic signatures has the potential to precisely predict pathology invasiveness. KEY POINTS: • The radiomic signature from the perinodular area has the potential to predict pathology invasiveness of the solitary pulmonary nodule. • The new radiomic nomogram was useful in clinical decision-making associated with personalized surgical intervention and therapeutic regimen selection in patients with early-stage non-small-cell lung cancer.

The long-term effect of the Earned Income Tax Credit on women's physical and mental health.

Using a novel method, and data from the National Longitudinal Survey of Youth 1979 (NLSY79), we estimate the cumulative, long-term, causal effect of Earned Income Tax Credit (EITC) eligibility on women's physical and mental health at age 50. We find that an increase in lifetime eligible EITC benefits is associated with long-term improvements in physical health, such as reduced occurrence of activity-limiting health problems and reduced reported diagnoses of mild and severe diseases. We explore intermediate health behaviors and outcomes, and find that an increase in lifetime eligible EITC benefits increases the number of hours worked and access to employer-sponsored health insurance, and decreases body mass index in the short-term. We find no significant effects of the EITC on mental health at age 50. Finally, we find that White women benefit disproportionately from the EITC in terms of mobility-related health issues, while Black and Hispanic women benefit in terms of lung-related illnesses like asthma, as well as cancer and stroke.

Characteristics, goal-oriented treatments and survival of pulmonary arterial hypertension in China: Insights from a national multicentre prospective registry.

BACKGROUND AND OBJECTIVE: Nationally representative reports on the characteristics and long-term survival of pulmonary arterial hypertension (PAH) from developing countries are scarce. The applicability of the current main risk stratifications and the longitudinal changes in goal-oriented treatments have yet to be elucidated in real-world settings. Therefore, we aimed to provide insights into the characteristics, goal-oriented treatments and survival of PAH in China and to explore the applicability of the main risk stratifications in our independent cohort. METHODS: PAH patients were consecutively enrolled from a national prospective multicentre registry. Data on baseline, follow-up re-evaluation and therapeutic changes were collected. RESULTS: A total of 2031 patients were enrolled, with congenital heart disease (CHD)-PAH (45.2%) being the most common aetiology. The mean age was 35 ± 12 years, and 76.2% were females. At baseline, approximately 20% of the patients with intermediate or high risk received combination treatment. At follow-up, approximately half of the re-evaluated patients did not achieve low-risk profiles, and even among patients who received combination therapy at baseline, 4% of them still worsened. The rate of combination therapy increased significantly from 6.7% before 2015 to 35.5% thereafter. The main risk assessment tools demonstrated good performance for predicting survival both at baseline and at follow-up. CONCLUSION: Chinese PAH patients show both similar and distinct features compared to other countries. Current main risk stratifications can significantly discriminate patients at different risk levels. There were still many patients not achieving low-risk profiles at follow-up, indicating more aggressive treatment should be implemented to optimize the goal-oriented treatment strategy.

Nitrogen Starvation Enhances the Production of Saturated and Unsaturated Fatty Acids in Aurantiochytrium sp. PKU#SW8 by Regulating Key Biosynthetic Genes.

Nitrogen deprivation is known to improve lipid accumulation in microalgae and thraustochytrids. However, the patterns of fatty acid production and the molecular mechanisms underlying the accumulation of unsaturated and saturated fatty acids (SFAs) under nitrogen starvation remain largely unknown for thraustochytrids. In this study, batch culture experiments under nitrogen replete and nitrogen starvation conditions were performed, and the changes in the transcriptome of Aurantiochytrium sp. PKU#SW8 strain between these conditions were investigated. Our results showed improved yields of total fatty acids (TFAs), total unsaturated fatty acids, and total SFAs under nitrogen starvation, which suggested that nitrogen starvation favors the accumulation of both unsaturated and saturated fatty acids in PKU#SW8. However, nitrogen starvation resulted in a more than 2.36-fold increase of SFAs whereas a 1.7-fold increase of unsaturated fatty acids was observed, indicating a disproportionate increase in these groups of fatty acids. The fabD and enoyl-CoA hydratase genes were significantly upregulated under nitrogen starvation, supporting the observed increase in the yield of TFAs from 2.63 ± 0.22 g/L to 3.64 ± 0.16 g/L. Furthermore, the pfaB gene involved in the polyketide synthase (PKS) pathway was significantly upregulated under nitrogen starvation. This suggested that the increased expression of the pfaB gene under nitrogen starvation may be one of the explanations for the increased yield of docosahexaenoic acid by 1.58-fold. Overall, our study advances the current understanding of the molecular mechanisms that underlie the response of thraustochytrids to nitrogen deprivation and their fatty acid biosynthesis.

Culturable Diversity of Thraustochytrids from Coastal Waters of Qingdao and Their Fatty Acids.

Thraustochytrids have gained significant attention in recent years because of their considerable ecological and biotechnological importance. Yet, the influence of seasons and habitats on their culturable diversity and lipid profile remains poorly described. In this study, a total of 58 thraustochytrid strains were isolated from the coastal waters of Qingdao, China. These strains were phylogenetically close to five thraustochytrid genera, namely Botryochytrium, Oblongichytrium, Schizochytrium, Thraustochytrium, and Sicyoidochytrium. Most of the isolated strains were classified into the genera Thraustochytrium and Oblongichytrium. Further diversity analysis revealed that samples collected from nutrient-rich habitats and during summer/fall yielded significantly higher culturable diversity of thraustochytrids than those from low-nutrient habitats and winter/spring. Moreover, sampling habitats and seasons significantly impacted the fatty acid profiles of the strains. Particularly, the Oblongichytrium sp. OC931 strain produced a significant amount (153.99 mg/L) of eicosapentaenoic acid (EPA), accounting for 9.12% of the total fatty acids, which was significantly higher than that of the previously reported Aurantiochytrium strains. Overall, the results of this study fill the gap in our current understanding of the culturable diversity of thraustochytrids in the coastal waters and the impact of the sampling habitats and seasons on their capacity for lipid accumulation.