SARS-CoV-2 RNA stabilizes host mRNAs to elicit immunopathogenesis.

SARS-CoV-2 RNA interacts with host factors to suppress interferon responses and simultaneously induces cytokine release to drive the development of severe coronavirus disease 2019 (COVID-19). However, how SARS-CoV-2 hijacks host RNAs to elicit such imbalanced immune responses remains elusive. Here, we analyzed SARS-CoV-2 RNA in situ structures and interactions in infected cells and patient lung samples using RIC-seq. We discovered that SARS-CoV-2 RNA forms 2,095 potential duplexes with the 3' UTRs of 205 host mRNAs to increase their stability by recruiting RNA-binding protein YBX3 in A549 cells. Disrupting the SARS-CoV-2-to-host RNA duplex or knocking down YBX3 decreased host mRNA stability and reduced viral replication. Among SARS-CoV-2-stabilized host targets, NFKBIZ was crucial for promoting cytokine production and reducing interferon responses, probably contributing to cytokine storm induction. Our study uncovers the crucial roles of RNA-RNA interactions in the immunopathogenesis of RNA viruses such as SARS-CoV-2 and provides valuable host targets for drug development.

Capture RIC-seq reveals positional rules of PTBP1-associated RNA loops in splicing regulation.

RNA-binding proteins (RBPs) bind at different positions of the pre-mRNA molecules to promote or reduce the usage of a particular exon. Seeking to understand the working principle of these positional effects, we develop a capture RIC-seq (CRIC-seq) method to enrich specific RBP-associated in situ proximal RNA-RNA fragments for deep sequencing. We determine hnRNPA1-, SRSF1-, and PTBP1-associated proximal RNA-RNA contacts and regulatory mechanisms in HeLa cells. Unexpectedly, the 3D RNA map analysis shows that PTBP1-associated loops in individual introns preferentially promote cassette exon splicing by accelerating asymmetric intron removal, whereas the loops spanning across cassette exon primarily repress splicing. These "positional rules" can faithfully predict PTBP1-regulated splicing outcomes. We further demonstrate that cancer-related splicing quantitative trait loci can disrupt RNA loops by reducing PTBP1 binding on pre-mRNAs to cause aberrant splicing in tumors. Our study presents a powerful method for exploring the functions of RBP-associated RNA-RNA proximal contacts in gene regulation and disease.

Complementary Alu sequences mediate enhancer-promoter selectivity.

Enhancers determine spatiotemporal gene expression programs by engaging with long-range promoters1-4. However, it remains unknown how enhancers find their cognate promoters. We recently developed a RNA in situ conformation sequencing technology to identify enhancer-promoter connectivity using pairwise interacting enhancer RNAs and promoter-derived noncoding RNAs5,6. Here we apply this technology to generate high-confidence enhancer-promoter RNA interaction maps in six additional cell lines. Using these maps, we discover that 37.9% of the enhancer-promoter RNA interaction sites are overlapped with Alu sequences. These pairwise interacting Alu and non-Alu RNA sequences tend to be complementary and potentially form duplexes. Knockout of Alu elements compromises enhancer-promoter looping, whereas Alu insertion or CRISPR-dCasRx-mediated Alu tethering to unregulated promoter RNAs can create new loops to homologous enhancers. Mapping 535,404 noncoding risk variants back to the enhancer-promoter RNA interaction maps enabled us to construct variant-to-function maps for interpreting their molecular functions, including 15,318 deletions or insertions in 11,677 Alu elements that affect 6,497 protein-coding genes. We further demonstrate that polymorphic Alu insertion at the PTK2 enhancer can promote tumorigenesis. Our study uncovers a principle for determining enhancer-promoter pairing specificity and provides a framework to link noncoding risk variants to their molecular functions.

RIC-seq for global in situ profiling of RNA-RNA spatial interactions.

Highly structured RNA molecules usually interact with each other, and associate with various RNA-binding proteins, to regulate critical biological processes. However, RNA structures and interactions in intact cells remain largely unknown. Here, by coupling proximity ligation mediated by RNA-binding proteins with deep sequencing, we report an RNA in situ conformation sequencing (RIC-seq) technology for the global profiling of intra- and intermolecular RNA-RNA interactions. This technique not only recapitulates known RNA secondary structures and tertiary interactions, but also facilitates the generation of three-dimensional (3D) interaction maps of RNA in human cells. Using these maps, we identify noncoding RNA targets globally, and discern RNA topological domains and trans-interacting hubs. We reveal that the functional connectivity of enhancers and promoters can be assigned using their pairwise-interacting RNAs. Furthermore, we show that CCAT1-5L-a super-enhancer hub RNA-interacts with the RNA-binding protein hnRNPK, as well as RNA derived from the MYC promoter and enhancer, to boost MYC transcription by modulating chromatin looping. Our study demonstrates the power and applicability of RIC-seq in discovering the 3D structures, interactions and regulatory roles of RNA.

Locus Coeruleus Integrity Is Linked to Response Inhibition Deficits in Parkinson's Disease and Progressive Supranuclear Palsy.

Parkinson's disease (PD) and progressive supranuclear palsy (PSP) both impair response inhibition, exacerbating impulsivity. Inhibitory control deficits vary across individuals and are linked with worse prognosis, and lack improvement on dopaminergic therapy. Motor and cognitive control are associated with noradrenergic innervation of the cortex, arising from the locus coeruleus (LC) noradrenergic system. Here we test the hypothesis that structural variation of the LC explains response inhibition deficits in PSP and PD. Twenty-four people with idiopathic PD, 14 with PSP-Richardson's syndrome, and 24 age- and sex-matched controls undertook a stop-signal task and ultrahigh field 7T magnetization-transfer-weighted imaging of the LC. Parameters of "race models" of go- versus stop-decisions were estimated using hierarchical Bayesian methods to quantify the cognitive processes of response inhibition. We tested the multivariate relationship between LC integrity and model parameters using partial least squares. Both disorders impaired response inhibition at the group level. PSP caused a distinct pattern of abnormalities in inhibitory control with a paradoxically reduced threshold for go responses, but longer nondecision times, and more lapses of attention. The variation in response inhibition correlated with the variability of LC integrity across participants in both clinical groups. Structural imaging of the LC, coupled with behavioral modeling in parkinsonian disorders, confirms that LC integrity is associated with response inhibition and LC degeneration contributes to neurobehavioral changes. The noradrenergic system is therefore a promising target to treat impulsivity in these conditions. The optimization of noradrenergic treatment is likely to benefit from stratification according to LC integrity.SIGNIFICANCE STATEMENT Response inhibition deficits contribute to clinical symptoms and poor outcomes in people with Parkinson's disease and progressive supranuclear palsy. We used cognitive modeling of performance of a response inhibition task to identify disease-specific mechanisms of abnormal inhibitory control. Response inhibition in both patient groups was associated with the integrity of the noradrenergic locus coeruleus, which we measured in vivo using ultra-high field MRI. We propose that the imaging biomarker of locus coeruleus integrity provides a trans-diagnostic tool to explain individual differences in response inhibition ability beyond the classic nosological borders and diagnostic criteria. Our data suggest a potential new stratified treatment approach for Parkinson's disease and progressive supranuclear palsy.

TMPRSS13 promotes the cell entry of swine acute diarrhea syndrome coronavirus.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) has caused severe intestinal diseases in pigs. It originates from bat coronaviruses HKU2 and has a potential risk of cross-species transmission, raising concerns about its zoonotic potential. Viral entry-related host factors are critical determinants of susceptibility to cells, tissues, or species, and remain to be elucidated for SADS-CoV. Type II transmembrane serine proteases (TTSPs) family is involved in many coronavirus infections and has trypsin-like catalytic activity. Here we examine all 18 members of the TTSPs family through CRISPR-based activation of endogenous protein expression in cells, and find that, in addition to TMPRSS2 and TMPRSS4, TMPRSS13 significantly facilitates SADS-CoV infection. This is confirmed by ectopic expression of TMPRSS13, and specific to trypsin-dependent SADS-CoV. Infection with pseudovirus bearing SADS-CoV spike protein indicates that TMPRSS13 acts at the entry step and is sensitive to serine protease inhibitor Camostat. Moreover, both human and pig TMPRSS13 are able to enhance the cell-cell membrane fusion and cleavage of spike protein. Overall, we demonstrate that TMPRSS13 is another host serine protease promoting the membrane-fusion entry of SADS-CoV, which may expand its host tropism by using diverse TTSPs.

Low Prognostic Nutritional Index Is Common and Associated with Poor Outcomes following Curative-Intent Resection for Gastro-Entero-Pancreatic Neuroendocrine Tumors.

INTRODUCTION: To investigate the impact of prognostic nutritional index (PNI) on short- and long-term outcomes of patients who underwent curative-intent resection for gastro-entero-pancreatic neuroendocrine tumors (GEP-NETs). METHODS: Patients with GET-NETs who underwent curative-intent resection were identified from a multi-center database. The prognostic impact of clinicopathological factors including PNI on post-operative outcomes were evaluated. A novel nomogram was developed and externally validated. RESULTS: A total of 2,099 patients with GEP-NETs were included in the training cohort; 255 patients were in the external validation cohort. Median PNI (n = 973) was 47.4 (IQR 43.1-52.4). At the time of presentation, 1,299 (61.9%) patients presented with some type of clinical symptom. Low-PNI (≤42.2) was associated with gastrointestinal symptoms, as well as nodal metastasis and distant metastasis (all p < 0.05). Patients with a low PNI had a higher incidence of severe (≥Clavien-Dindo grade IIIa: low PNI 24.9% vs. high PNI 15.4%, p = 0.001) and multiple (≥3 types of complications: low PNI 14.5% vs. high PNI 9.2%, p = 0.024) complications, as well as a worse overall survival (OS)(5-year OS, low PNI 73.7% vs. high PNI 88.5%, p < 0.001), and RFS (5-year RFS, low PNI 68.5% vs. high PNI 79.8%, p = 0.008) versus patients with high PNI (>42.2). A nomogram based on PNI, tumor grade and metastatic disease demonstrated excellent discrimination and calibration to predict OS in both the training (C-index 0.748) and two external validation (C-index 0.827, 0.745) cohorts. CONCLUSIONS: Low PNI was common and associated with worse short- and long-term outcomes among patients with GEP-NETs.

Molecular mechanisms of HCG18 in the sorafenib resistance of hepatocellular carcinoma.

Sorafenib has been approved for advance hepatocellular carcinoma (HCC), however, drug resistance often occurred. Therefore, it is of great significance to clarify the underlying mechanisms of sorafenib resistance and to find out the effective strategies to overcome sorafenib resistance. The expression of HCG18 was detected by qPCR, MTT, colony formation, flow cytometry and TUNEL assay were used to explore the function of HCG18 on sorafenib resistance in HCC. RNA pull-down, RNA immunoprecipitation, immunofluorescence labeling, luciferase reporter assay, western blot and qPCR were used to investigate the mechanism of HCG18 regulating sorafenib resistance in HCC. Our results showed that HCG18 was significantly increased in HCC, which resulted in shorter 5-year survival for patients with HCC. Sorafenib can induce the expression of HCG18, suggesting HCG18 might be involved in sorafenib resistance in HCC. Further analysis showed that knockdown of HCG18 can reduce viability and increase apoptosis of HCC cells. Mechanistically, HCG18 can bind to USP15, further regulated the protein stability of p65, TAB2 and TAB3, and nuclear location of p65, which finally modulated the NF-κB signaling. Our findings showed that HCG18 played an important role in sorafenib resistance in HCC. And knockdown of HCG18 can promote the sensitivity of HCC cells to sorafenib, inferring that targeting HCG18 might be an effective strategy to overcome sorafenib resistance in HCC.

New sexually transmitted HIV infections from 2016 to 2050 in Guangdong Province, China: a study based on a dynamic compartmental model.

BACKGROUND: In Guangdong Province, China, there is lack of information on the HIV epidemic among high-risk groups and the general population, particularly in relation to sexual transmission, which is a predominant route. The new HIV infections each year is also uncertain owing to HIV transmission from men who have sex with men (MSM) to women, as a substantial proportion of MSM also have female sexual partnerships to comply with social demands in China. METHODS: A deterministic compartmental model was developed to predict new HIV infections in four risk groups, including heterosexual men and women and low- and high-risk MSM, in Guangdong Province from 2016 to 2050, considering HIV transmission from MSM to women. The new HIV infections and its 95% credible interval (CrI) were predicted. An adaptive sequential Monte Carlo method for approximate Bayesian computation (ABC-SMC) was used to estimate the unknown parameter, a mixing index. We calibrated our results based on new HIV diagnoses and proportions of late diagnoses. The Morris and Sobol methods were applied in the sensitivity analysis. RESULTS: New HIV infections increased during and 2 years after the COVID-19 pandemic, then declined until 2050. New infections rose from 8,828 [95% credible interval (CrI): 6,435-10,451] in 2016 to 9,652 (95% CrI: 7,027-11,434) in 2019, peaking at 11,152 (95% CrI: 8,337-13,062) in 2024 before declining to 7,084 (95% CrI: 5,165-8,385) in 2035 and 4,849 (95% CrI: 3,524-5,747) in 2050. Women accounted for approximately 25.0% of new HIV infections, MSM accounted for 40.0% (approximately 55.0% of men), and high-risk MSM accounted for approximately 25.0% of the total. The ABC-SMC mixing index was 0.504 (95% CrI: 0.239-0.894). CONCLUSIONS: Given that new HIV infections and the proportion of women were relatively high in our calibrated model, to some extent, the HIV epidemic in Guangdong Province remains serious, and services for HIV prevention and control are urgently needed to return to the levels before the COVID-19 epidemic, especially in promoting condom-based safe sex and increasing awareness of HIV prevention to general population.

YOLOv8-RMDA: Lightweight YOLOv8 Network for Early Detection of Small Target Diseases in Tea.

In order to efficiently identify early tea diseases, an improved YOLOv8 lesion detection method is proposed to address the challenges posed by the complex background of tea diseases, difficulty in detecting small lesions, and low recognition rate of similar phenotypic symptoms. This method focuses on detecting tea leaf blight, tea white spot, tea sooty leaf disease, and tea ring spot as the research objects. This paper presents an enhancement to the YOLOv8 network framework by introducing the Receptive Field Concentration-Based Attention Module (RFCBAM) into the backbone network to replace C2f, thereby improving feature extraction capabilities. Additionally, a mixed pooling module (Mixed Pooling SPPF, MixSPPF) is proposed to enhance information blending between features at different levels. In the neck network, the RepGFPN module replaces the C2f module to further enhance feature extraction. The Dynamic Head module is embedded in the detection head part, applying multiple attention mechanisms to improve multi-scale spatial location and multi-task perception capabilities. The inner-IoU loss function is used to replace the original CIoU, improving learning ability for small lesion samples. Furthermore, the AKConv block replaces the traditional convolution Conv block to allow for the arbitrary sampling of targets of various sizes, reducing model parameters and enhancing disease detection. the experimental results using a self-built dataset demonstrate that the enhanced YOLOv8-RMDA exhibits superior detection capabilities in detecting small target disease areas, achieving an average accuracy of 93.04% in identifying early tea lesions. When compared to Faster R-CNN, MobileNetV2, and SSD, the average precision rates of YOLOv5, YOLOv7, and YOLOv8 have shown improvements of 20.41%, 17.92%, 12.18%, 12.18%, 10.85%, 7.32%, and 5.97%, respectively. Additionally, the recall rate (R) has increased by 15.25% compared to the lowest-performing Faster R-CNN model and by 8.15% compared to the top-performing YOLOv8 model. With an FPS of 132, YOLOv8-RMDA meets the requirements for real-time detection, enabling the swift and accurate identification of early tea diseases. This advancement presents a valuable approach for enhancing the ecological tea industry in Yunnan, ensuring its healthy development.

Assessing Mailuoning injection in wound healing and thrombophlebitis management: A rat model study.

Thrombophlebitis is the inflammatory condition characterized by obstruction of one or more vessels, commonly in the legs, due to the formation of blood clots. It has been reported that traditional Chinese medicine, including Mailuoning injection, is advantageous for treating inflammatory and blood disorders. This research assessed the therapeutic efficacy of Mailuoning injection in the treatment of thrombophlebitis in rodents, as well as investigated its impact on fibrinolysis, inflammation, and coagulation. An experimental setup for thrombophlebitis was established in rodents via modified ligation technique. Five groups comprised the animals: sham operation group, model group, and three Mailuoning treatment groups (low, medium, and high dosages). The pain response, edema, coagulation parameters (PT, APTT, TT, FIB), serum inflammatory markers (IL-6, TNF-α, CRP), and expression levels of endothelial markers (ICAM-1, VCAM-1, NF-κB) were evaluated. Blood flow and vascular function were further assessed by measuring hemorheological parameters and the concentrations of TXB2, ET, and 6-k-PGF1α. In contrast to the sham group, model group demonstrated statistically significant increases in endothelial expression levels, coagulation latencies, and inflammatory markers (p < 0.05). The administration of mailing, specifically at high and medium dosages, resulted in a substantial reduction in inflammatory markers, enhancement of coagulation parameters, suppression of ICAM-1 and VCAM-1 expression, and restoration of hemorheological measurements to baseline (p < 0.05). Significantly higher concentrations of 6-k-PGF1α and lower levels of TXB2 and ET were observed in high-dose group, suggesting that pro- and anti-thrombotic factors were restored to equilibrium. Utilization of Mailuoning injection in rat model of thrombophlebitis exhibited significant therapeutic impact. This effect was manifested through pain alleviation, diminished inflammation, enhanced blood viscosity and facilitation of fibrinolysis. The study indicated that Mailuoning injection may serve as a viable therapeutic option for thrombophlebitis, potentially aiding in the improvement of wound healing by virtue of its anti-inflammatory and blood flow-enhancing characteristics.

Prefrontal Cortical Connectivity Mediates Locus Coeruleus Noradrenergic Regulation of Inhibitory Control in Older Adults.

Response inhibition is a core executive function enabling adaptive behavior in dynamic environments. Human and animal models indicate that inhibitory control and control networks are modulated by noradrenaline, arising from the locus coeruleus. The integrity (i.e., cellular density) of the locus coeruleus noradrenergic system can be estimated from magnetization transfer (MT)-sensitive magnetic resonance imaging (MRI), in view of neuromelanin present in noradrenergic neurons of older adults. Noradrenergic psychopharmacological studies indicate noradrenergic modulation of prefrontal and frontostriatal stopping-circuits in association with behavioral change. Here, we test the noradrenergic hypothesis of inhibitory control, in healthy adults. We predicted that locus coeruleus integrity is associated with age-adjusted variance in response inhibition, mediated by changes in connectivity between frontal inhibitory control regions. In a preregistered analysis, we used MT MRI images from N = 63 healthy humans aged above 50 years (of either sex) who performed a Stop-Signal Task (SST), with atlas-based measurement of locus coeruleus contrast. We confirm that better response inhibition is correlated with locus coeruleus integrity and stronger connectivity between presupplementary motor area (preSMA) and right inferior frontal gyrus (rIFG), but not volumes of the prefrontal cortical regions. We confirmed a significant role of prefrontal connectivity in mediating the effect of individual differences in the locus coeruleus on behavior, where this effect was moderated by age, over and above adjustment for the mean effects of age. Our results support the hypothesis that in normal populations, as in clinical settings, the locus coeruleus noradrenergic system regulates inhibitory control.SIGNIFICANCE STATEMENT We show that the integrity of the locus coeruleus, the principal source of cortical noradrenaline, is related to the efficiency of response inhibition in healthy older adults. This effect is in part mediated by its effect on functional connectivity in a prefrontal cortical stopping-network. The behavioral effect, and its mediation by connectivity, are moderated by age. This supports the psychopharmacological and genetic evidence for the noradrenergic regulation of behavioral control, in a population-based normative cohort. Noradrenergic treatment strategies may be effective to improve behavioral control in impulsive clinical populations, but age, and locus coeruleus integrity, are likely to be important stratification factors.

Elevated FAM134B expression induces radiation-sensitive in hepatocellular carcinoma.

BACKGROUND: Previous studies have shown that Family with sequence similarity 134 member B (FAM134B) was involved in the occurrence and development of malignancy, however, the function and molecular mechanism of FAM134B in Hepatocellular Carcinoma (HCC) radiotherapy resistance remain unclear. Therefore, it may clinical effective to clarify the molecular mechanism and identify novel biomarker to overcome radiotherapy resistance in HCC. METHODS: The protein and mRNA expression of FAM134B were determined using Real-time PCR and Western blot, respectively. IHC assay was performed to investigate the association between FAM134B expression and the clinicopathological characteristics of 132 HCC patients. Functional assays, such as in situ model, colon formation, FACS, and Tunel assay were used to determine the oncogenic role of FAM134B in human HCC progression. Furthermore, western blotting and luciferase assay were used to determine the mechanism of FAM134B promotes radiation-sensitive in HCC cells. RESULTS: We noted that FAM134B was downregulated in HCC, which was correlated with the radiation resistance in patients with HCC. Overexpression of FAM134B contribute to radiation sensitive in HCC; however, inhibition of FAM134B confers HCC cell lines to radiation resistance both in vitro and in vivo. Moreover, we found that FAM134B interacts with FMS related receptor tyrosine kinase 3 (FLT3) and downregulation of FAM134B activated JAK/Stat3 signaling pathway. Importantly, pharmacological inhibition of JAK/Stat3 signaling pathway significantly counteracted downregulation of FAM134B-induced radiation resistance and enhanced radiation therapeutic efficacy in HCC. CONCLUSIONS: Our findings suggest that FAM134B may be a potential therapeutic biomarker for the treatment of HCC patients with radiotherapy tolerance.

Noradrenergic deficits contribute to apathy in Parkinson's disease through the precision of expected outcomes.

Apathy is a debilitating feature of many neuropsychiatric diseases, that is typically described as a reduction of goal-directed behaviour. Despite its prevalence and prognostic importance, the mechanisms underlying apathy remain controversial. Degeneration of the locus coeruleus-noradrenaline system is known to contribute to motivational deficits, including apathy. In healthy people, noradrenaline has been implicated in signalling the uncertainty of expectations about the environment. We proposed that noradrenergic deficits contribute to apathy by modulating the relative weighting of prior beliefs about action outcomes. We tested this hypothesis in the clinical context of Parkinson's disease, given its associations with apathy and noradrenergic dysfunction. Participants with mild-to-moderate Parkinson's disease (N = 17) completed a randomised double-blind, placebo-controlled, crossover study with 40 mg of the noradrenaline reuptake inhibitor atomoxetine. Prior weighting was inferred from psychophysical analysis of performance in an effort-based visuomotor task, and was confirmed as negatively correlated with apathy. Locus coeruleus integrity was assessed in vivo using magnetisation transfer imaging at ultra-high field 7T. The effect of atomoxetine depended on locus coeruleus integrity: participants with a more degenerate locus coeruleus showed a greater increase in prior weighting on atomoxetine versus placebo. The results indicate a contribution of the noradrenergic system to apathy and potential benefit from noradrenergic treatment of people with Parkinson's disease, subject to stratification according to locus coeruleus integrity. More broadly, these results reconcile emerging predictive processing accounts of the role of noradrenaline in goal-directed behaviour with the clinical symptom of apathy and its potential pharmacological treatment.

Potential candidate genes and pathways related to cytoplasmic male sterility in Dianthus spiculifolius as revealed by transcriptome analysis.

KEY MESSAGE: We introduced the candidate gene DsHSP70 into Arabidopsis thaliana, resulting in male gametophyte sterility and abnormal degeneration of sepals and petals. Cytoplasmic male sterility (CMS) is a useful tool for hybrid production. However, the regulatory mechanism of CMS in Dianthus spiculifolius remains unclear. In this study, we investigated whether male-sterile line of D. spiculifolius has a malformed tapetum and fails to produce normal fertile pollen. RNA sequencing technology was used to compare the gene expression patterns of the D. spiculifolius male-sterile line and its male fertility maintainer line during anther development. A total of 12,365 differentially expressed genes (DEGs) were identified, among which 1765 were commonly expressed in the S1, S2 and S3 stages. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses indicated that these DEGs were mainly involved in oxidation-reduction processes, signal transduction and programmed cell death. Additionally, weighted correlation network analysis (WGCNA) showed that three modules may be related to male sterility. A putative regulatory pathway for the male sterility traits was constructed based on the reproductive development network. After introducing the candidate DsHSP70 gene into Arabidopsis thaliana, we found that overexpressing plants showed anther abortion and shorter filaments, and accompanied by abnormal degeneration of sepals and petals. In summary, our results identified potential candidate genes and pathways related to CMS in D. spiculifolius, providing new insights for further research on the mechanism of male sterility.

CircSOD2 Contributes to Tumor Progression, Immune Evasion and Anti-PD-1 Resistance in Hepatocellular Carcinoma by Targeting miR-497-5p/ANXA11 Axis.

Circular RNAs (circRNAs) can function as functional molecules in hepatocellular carcinoma (HCC). Herein, circRNA superoxide dismutase 2 (circSOD2) was researched in HCC progression and immune system. The real-time polymerase chain reaction (qRT-PCR) was used for quantification of circSOD2, microRNA-497-5p (miR-497-5p) and Annexin A11 (ANXA11). Cell assays were performed by 3-(4, 5-dimethylthiazol-2-y1)-2, 5-diphenyl tetrazolium bromide (MTT) and colony formation assays for proliferation, flow cytometry for apoptosis and cell cycle, wound healing assay for migration and transwell assay for migration/invasion. ANXA11 and metastatic protein levels were measured by western blot. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were performed to analyze target binding. CD8+ T cell immunity was assessed by Immunohistochemistry (IHC) assay, and the effect of circSOD2 on programmed cell death 1 (PD-1) immune checkpoint inhibitors (anti-PD-1) therapy was evaluated by mice xenograft assay. CircSOD2 was upregulated in HCC tissues and cells. Knockdown of circSOD2 resulted in HCC cell growth inhibition, apoptosis promotion, cell cycle arrest and metastasis suppression. Mechanically, circSOD2 promoted HCC development by acting as a miR-497-5p sponge and miR-497-5p played a tumor-inhibitory role in HCC cells by targeting ANXA11. Moreover, circSOD2 induced upregulation of ANXA11 expression by interacting with miR-497-5p. Also, the promoting effects of circSOD2 on immune evasion and anti-PD-1 resistance were related to miR-497-5p/ANXA11 axis. This study elucidated the pivotal function of circSOD2 in HCC progression and immunosuppression by mediating miR-497-6p/ANXA11 axis. CircSOD2/miR-497-5p/ANXA11 axis was a novel view of circRNA research in HCC.

LIM Homeobox 2 Increases Adhesion-Regulating Molecule 1 Transcription to Facilitate the Pathological Progression of Oxidized Low-Density Lipoprotein-Stimulated Atherosclerotic Cell Models.

Endothelial-mesenchymal transition (EndMT) and endothelial cell apoptosis have been documented to have a role in atherosclerosis (AS) progression. To deepen knowledge in this aspect, our study investigated the effect of LIM homeobox 2 (LHX2) and adhesion-regulating molecule 1 (ADRM1) on EndMT and endothelial cell apoptosis in the oxidized low-density lipoprotein (ox-LDL) -stimulated AS cell model.Ox-LDL was utilized to treat human umbilical vein endothelial cells (HUVECs) for constructing an AS model in vitro, followed by measurement of LHX2 and ADRM1 expressions. Afterward, gain- and loss-of-function assays were performed in HUVECs, followed by detection of cell viability, invasion, migration, and apoptosis and the expression of inflammatory factors [tumor necrosis factor (TNF) -α, interleukin (IL) -1ß, and IL-6], EndMT-related proteins [CD31, vascular epithelium (VE) -cadherin, vimentin, α-smooth muscle actin (SMA), Snai1, Snai2, and Twist1], and the apoptotic protein cleaved caspase-3. Interactions between LHX2 and ADRM1 were analyzed with dual-luciferase reporter gene and chromatin immunoprecipitation assays.High levels of LHX2 and ADRM1 were observed in ox-LDL-induced HUVECs. In ox-LDL-treated HUVECs, LHX2, or ADRM1 knockdown promoted CD31 and VE-cadherin levels, viability, invasion, and migration and reduced apoptosis and the expressions of TNF-α, IL-1ß, IL-6, vimentin, α-SMA, Snai1, Snai2, Twist1, and cleaved caspase-3. Mechanistically, LHX2 bound to the ADRM1 promoter to promote ADRM1 transcription. Overexpression of ADRM1 annulled the aforementioned effects of LHX2 knockdown on ox-LDL-induced HUVECs.LHX2 facilitates the pathological progression of ox-LDL-stimulated AS cell models by increasing ADRM1 transcription.

Generation of low-cadmium rice germplasms via knockout of OsLCD using CRISPR/Cas9.

The OsLCD gene, which has been implicated in cadmium (Cd) accumulation in rice, might be a useful target for CRISPR/Cas9 editing. However, the effects of OsLCD gene editing on Cd accumulation, plant growth, and yield traits remain unknown. Here, we used CRISPR/Cas9 to generate oslcd single mutants from indica and japonica rice cultivars. We also generated osnramp5 single mutants and oslcd osnramp5 double mutants in the indica background. When grown in Cd-contaminated paddy soils, all oslcd single mutants accumulated less Cd than the wild types (WTs). Consistent with this, oslcd single mutants grown in Cd-contaminated hydroponic culture accumulated significantly less Cd in the shoots as compared to WTs. This decrease in accumulation probably resulted from the reduction of Cd translocation under Cd stress. Oxidative damage also decreased, and plant growth increased in all oslcd single mutant seedlings as compared to WTs in the presence of Cd. Plant growth and most yield traits, as well essential element concentrations in rice seedling shoots, brown rice, and rice straw, were similar between oslcd single mutants and WTs. In the presence of Cd, Cd concentrations in the brown rice and shoots of oslcd osnramp5 double mutants were significantly decreased compared with WTs as well as osnramp single mutants. Our results suggested that OsLCD knockout may reduce Cd accumulation alone or in combination with other knockout mutations in a variety of rice genotypes; unlike OsNramp5 mutations, OsLCD knockout did not reduce essential element contents. Therefore, OsLCD knockout might be used to generate low-Cd rice germplasms.

Enantioselective Synthesis of Quaternary Oxindoles: Desymmetrizing Staudinger-Aza-Wittig Reaction Enabled by a Bespoke HypPhos Oxide Catalyst.

This paper describes a catalytic asymmetric Staudinger-aza-Wittig reaction of (o-azidoaryl)malonates, allowing access to chiral quaternary oxindoles through phosphine oxide catalysis. We designed a novel HypPhos oxide catalyst to enable the desymmetrizing Staudinger-aza-Wittig reaction through the PIII/PV═O redox cycle in the presence of a silane reductant and an IrI-based Lewis acid. The reaction occurs under mild conditions, with good functional group tolerance, a wide substrate scope, and excellent enantioselectivity. Density functional theory revealed that the enantioselectivity in the desymmetrizing reaction arose from the cooperative effects of the IrI species and the HypPhos catalyst. The utility of this methodology is demonstrated by the (formal) syntheses of seven alkaloid targets: (-)-gliocladin C, (-)-coerulescine, (-)-horsfiline, (+)-deoxyeseroline, (+)-esermethole, (+)-physostigmine, and (+)-physovenine.