Faster sorting algorithms discovered using deep reinforcement learning.

Fundamental algorithms such as sorting or hashing are used trillions of times on any given day1. As demand for computation grows, it has become critical for these algorithms to be as performant as possible. Whereas remarkable progress has been achieved in the past2, making further improvements on the efficiency of these routines has proved challenging for both human scientists and computational approaches. Here we show how artificial intelligence can go beyond the current state of the art by discovering hitherto unknown routines. To realize this, we formulated the task of finding a better sorting routine as a single-player game. We then trained a new deep reinforcement learning agent, AlphaDev, to play this game. AlphaDev discovered small sorting algorithms from scratch that outperformed previously known human benchmarks. These algorithms have been integrated into the LLVM standard C++ sort library3. This change to this part of the sort library represents the replacement of a component with an algorithm that has been automatically discovered using reinforcement learning. We also present results in extra domains, showcasing the generality of the approach.

Insulin Determines Transforming Growth Factor ß Effects on Hepatocyte Nuclear Factor 4α Transcription in Hepatocytes.

Loss of hepatocyte nuclear factor 4α (HNF4α) expression is frequently observed in end-stage liver disease and associated with loss of vital liver functions, thus increasing mortality. Loss of HNF4α expression is mediated by inflammatory cytokines, such as transforming growth factor (TGF)-ß. However, details of how HNF4α is suppressed are largely unknown to date. Herein, TGF-ß did not directly inhibit HNF4α but contributed to its transcriptional regulation by SMAD2/3 recruiting acetyltransferase CREB-binding protein/p300 to the HNF4α promoter. The recruitment of CREB-binding protein/p300 is indispensable for CCAAT/enhancer-binding protein α (C/EBPα) binding, another essential requirement for constitutive HNF4α expression in hepatocytes. Consistent with the in vitro observation, 67 of 98 patients with hepatic HNF4α expressed both phospho-SMAD2 and C/EBPα, whereas 22 patients without HNF4α expression lacked either phospho-SMAD2 or C/EBPα. In contrast to the observed induction of HNF4α, SMAD2/3 inhibited C/EBPα transcription. Long-term TGF-ß incubation resulted in C/EBPα depletion, which abrogated HNF4α expression. Intriguingly, SMAD2/3 inhibitory binding to the C/EBPα promoter was abolished by insulin. Two-thirds of patients without C/EBPα lacked membrane glucose transporter type 2 expression in hepatocytes, indicating insulin resistance. Taken together, these data indicate that hepatic insulin sensitivity is essential for hepatic HNF4α expression in the condition of inflammation.

Enhancing cold resistance in Banana (Musa spp.) through EMS-induced mutagenesis, L-Hyp pressure selection: phenotypic alterations, biomass composition, and transcriptomic insights.

BACKGROUND: The cultivation of bananas encounters substantial obstacles, particularly due to the detrimental effects of cold stress on their growth and productivity. A potential remedy that has gained attention is the utilization of ethyl mesylate (EMS)-induced mutagenesis technology, which enables the creation of a genetically varied group of banana mutants. This complex procedure entails subjecting the mutants to further stress screening utilizing L-Hyp in order to identify those exhibiting improved resistance to cold. This study conducted a comprehensive optimization of the screening conditions for EMS mutagenesis and L-Hyp, resulting in the identification of the mutant cm784, which exhibited remarkable cold resistance. Subsequent investigations further elucidated the physiological and transcriptomic responses of cm784 to low-temperature stress. RESULTS: EMS mutagenesis had a substantial effect on banana seedlings, resulting in modifications in shoot and root traits, wherein a majority of seedlings exhibited delayed differentiation and limited elongation. Notably, mutant leaves displayed altered biomass composition, with starch content exhibiting the most pronounced variation. The application of L-Hyp pressure selection aided in the identification of cold-resistant mutants among seedling-lethal phenotypes. The mutant cm784 demonstrated enhanced cold resistance, as evidenced by improved survival rates and reduced symptoms of chilling injury. Physiological analyses demonstrated heightened activities of antioxidant enzymes and increased proline production in cm784 when subjected to cold stress. Transcriptome analysis unveiled 946 genes that were differentially expressed in cm784, with a notable enrichment in categories related to 'Carbohydrate transport and metabolism' and 'Secondary metabolites biosynthesis, transport, and catabolism'. CONCLUSION: The present findings provide insights into the molecular mechanisms that contribute to the heightened cold resistance observed in banana mutants. These mechanisms encompass enhanced carbohydrate metabolism and secondary metabolite biosynthesis, thereby emphasizing the adaptive strategies employed to mitigate the detrimental effects induced by cold stress.

Hsa_circ_0007321 regulates Zika virus replication through miR-492/NFKBID/NF-κB signaling pathway.

IMPORTANCE: Over the past decade, increasing evidence has shown that circular RNAs (circRNAs) play important regulatory roles in viral infection and host antiviral responses. However, reports on the role of circRNAs in Zika virus (ZIKV) infection are limited. In this study, we identified 45 differentially expressed circRNAs in ZIKV-infected A549 cells by RNA sequencing. We clarified that a downregulated circRNA, hsa_circ_0007321, regulates ZIKV replication through targeting of miR-492 and the downstream gene NFKBID. NFKBID is a negative regulator of nuclear factor-κB (NF-κB), and we found that inhibition of the NF-κB pathway promotes ZIKV replication. Therefore, this finding that hsa_circ_0007321 exerts its regulatory role on ZIKV replication through the miR-492/NFKBID/NF-κB signaling pathway has implications for the development of strategies to suppress ZIKV and possibly other viral infections.

Simple, sensitive, and visual detection of 12 respiratory pathogens with one-pot-RPA-CRISPR/Cas12a assay.

Respiratory infections pose a serious threat to global public health, underscoring the urgent need for rapid, accurate, and large-scale diagnostic tools. In recent years, the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated) system, combined with isothermal amplification methods, has seen widespread application in nucleic acid testing (NAT). However, achieving a single-tube reaction system containing all necessary components is challenging due to the competitive effects between recombinase polymerase amplification (RPA) and CRISPR/Cas reagents. Furthermore, to enable precision medicine, distinguishing between bacterial and viral infections is essential. Here, we have developed a novel NAT method, termed one-pot-RPA-CRISPR/Cas12a, which combines RPA with CRISPR molecular diagnostic technology, enabling simultaneous detection of 12 common respiratory pathogens, including six bacteria and six viruses. RPA and CRISPR/Cas12a reactions are separated by paraffin, providing an independent platform for RPA reactions to generate sufficient target products before being mixed with the CRISPR/Cas12a system. Results can be visually observed under LED blue light. The sensitivity of the one-pot-RPA-CRISPR/Cas12a method is 2.5 × 100 copies/µL plasmids, with no cross-reaction with other bacteria or viruses. Additionally, the clinical utility was evaluated by testing clinical isolates of bacteria and virus throat swab samples, demonstrating favorable performance. Thus, our one-pot-RPA-CRISPR/Cas12a method shows immense potential for accurate and large-scale detection of 12 common respiratory pathogens in point-of-care testing.

Autophagy-related protein 5 (ATG5) interacts with bone marrow stromal cell antigen 2 (BST2) to stimulate HBV replication through antagonizing the antiviral activity of BST2.

Hepatitis B virus (HBV) infection is a major global health burden with 820 000 deaths per year. In our previous study, we found that the knockdown of autophagy-related protein 5 (ATG5) significantly upregulated the interferon-stimulated genes (ISGs) expression to exert the anti-HCV effect. However, the regulation of ATG5 on HBV replication and its underlying mechanism remains unclear. In this study, we screened the altered expression of type I interferon (IFN-I) pathway genes using RT² Profiler™ PCR array following ATG5 knock-down and we found the bone marrow stromal cell antigen 2 (BST2) expression was significantly increased. We then verified the upregulation of BST2 by ATG5 knockdown using RT-qPCR and found that the knockdown of ATG5 activated the Janus kinase/signal transducer and activator of transcription (JAK-STAT) signaling pathway. ATG5 knockdown or BST2 overexpression decreased Hepatitis B core Antigen (HBcAg) protein, HBV DNA levels in cells and supernatants of HepAD38 and HBV-infected NTCP-HepG2. Knockdown of BST2 abrogated the anti-HBV effect of ATG5 knockdown. Furthermore, we found that ATG5 interacted with BST2, and further formed a ternary complex together with HBV-X (HBx). In conclusion, our finding indicates that ATG5 promotes HBV replication through decreasing BST2 expression and interacting with it directly to antagonize its antiviral function.

O-GlcNAc of STING mediates antiviral innate immunity.

BACKGROUND: O-GlcNAcylation modification affects multiple physiological and pathophysiolocal functions of cells. Altered O-GlcNAcylation was reported to participate in antivirus response. Stimulator of interferon genes (STING) is an adaptor mediating DNA virus-induced innate immune response. Whether STING is able to be modified by O-GlcNAcylation and how O-GlcNAcylation affects STING-mediated anti-DNA virus response remain unknown. METHODS: Metabolomics analysis was used for detecting metabolic alterations in HSV-1 infection cells. Succinylated wheat germ agglutinin (sWGA), co-immunoprecipitation, and pull-down assay were employed for determining O-GlcNAcylation. Mutagenesis PCR was applied for the generation of STING mutants. WT and Sting1-/- C57BL/6 mice (KOCMP-72512-Sting1-B6NVA) were infected with HSV-1 and treated with O-GlcNAcylation inhibitor for validating the role of STING O-GlcNAcylation in antiviral response. RESULTS: STING was functionally activated by O-GlcNAcylation in host cells challenged with HSV-1. We demonstrated that this signaling event was initiated by virus infection-enhanced hexosamine biosynthesis pathway (HBP). HSV-1 (or viral DNA mimics) promotes glucose metabolism of host cells with a marked increase in HBP, which provides donor glucosamine for O-GlcNAcylation. STING was O-GlcNAcylated on threonine 229, which led to lysine 63-linked ubiquitination of STING and activation of antiviral immune responses. Mutation of STING T229 to alanine abrogated STING activation and reduced HSV-1 stimulated production of interferon (IFN). Application of 6-diazo-5-oxonorleucine (DON), an agent that blocks the production of UDP-GlcNAc and inhibits O-GlcNAcylation, markedly attenuated the removal of HSV-1 in wild type C57BL/6 mice, leading to an increased viral retention, elevated infiltration of inflammatory cells, and worsened tissue damages to those displayed in STING gene knockout mice. Together, our data suggest that STING is O-GlcNAcylated in HSV-1, which is crucial for an effective antiviral innate immune response. CONCLUSION: HSV-1 infection activates the generation of UDP-Glc-NAc by upregulating the HBP metabolism. Elevated UDP-Glc-NAc promotes the O-GlcNAcylation of STING, which mediates the anti-viral function of STING. Targeting O-GlcNAcylation of STING could be a useful strategy for antiviral innate immunity.

New horizons for the role of RNA N6-methyladenosine modification in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common malignancy, presenting a formidable challenge to the medical community owing to its intricate pathogenic mechanisms. Although current prevention, surveillance, early detection, diagnosis, and treatment have achieved some success in preventing HCC and controlling overall disease mortality, the imperative to explore novel treatment modalities for HCC remains increasingly urgent. Epigenetic modification has emerged as pivotal factors in the etiology of cancer. Among these, RNA N6-methyladenosine (m6A) modification stands out as one of the most prevalent, abundant, and evolutionarily conserved post-transcriptional alterations in eukaryotes. The literature underscores that the dynamic and reversible nature of m6A modifications orchestrates the intricate regulation of gene expression, thereby exerting a profound influence on cell destinies. Increasing evidence has substantiated conspicuous fluctuations in m6A modification levels throughout the progression of HCC. The deliberate modulation of m6A modification levels through molecular biology and pharmacological interventions has been demonstrated to exert a discernible impact on the pathogenesis of HCC. In this review, we elucidate the multifaceted biological functions of m6A modifications in HCC, and concurrently advancing novel therapeutic strategies for the management of this malignancy.

Cascaded signal amplification strategy for ultra-specific, ultra-sensitive, and visual detection of Shigella flexneri.

Pathogen infections including Shigella flexneri have posed a significant threat to human health for numerous years. Although culturing and qPCR were the gold standards for pathogen detection, time-consuming and instrument-dependent restrict their application in rapid diagnosis and economically less-developed regions. Thus, it is urgently needed to develop rapid, simple, sensitive, accurate, and low-cost detection methods for pathogen detection. In this study, an immunomagnetic beads-recombinase polymerase amplification-CRISPR/Cas12a (IMB-RPA-CRISPR/Cas12a) method was built based on a cascaded signal amplification strategy for ultra-specific, ultra-sensitive, and visual detection of S. flexneri in the laboratory. Firstly, S. flexneri was specifically captured and enriched by IMB (Shigella antibody-coated magnetic beads), and the genomic DNA was released and used as the template in the RPA reaction. Then, the RPA products were mixed with the pre-loaded CRISPR/Cas12a for fluorescence visualization. The results were observed by naked eyes under LED blue light, with a sensitivity of 5 CFU/mL in a time of 70 min. With no specialized equipment or complicated technical requirements, the IMB-RPA-CRISPR/Cas12a diagnostic method can be used for visual, rapid, and simple detection of S. flexneri and can be easily adapted to monitoring other pathogens.

Integrated Metabolomic-Transcriptomic Analyses of Flavonoid Accumulation in Citrus Fruit under Exogenous Melatonin Treatment.

The flavonoids in citrus fruits are crucial physiological regulators and natural bioactive products of high pharmaceutical value. Melatonin is a pleiotropic hormone that can regulate plant morphogenesis and stress resistance and alter the accumulation of flavonoids in these processes. However, the direct effect of melatonin on citrus flavonoids remains unclear. In this study, nontargeted metabolomics and transcriptomics were utilized to reveal how exogenous melatonin affects flavonoid biosynthesis in "Bingtangcheng" citrus fruits. The melatonin treatment at 0.1 mmol L-1 significantly increased the contents of seven polymethoxylated flavones (PMFs) and up-regulated a series of flavonoid pathway genes, including 4CL (4-coumaroyl CoA ligase), FNS (flavone synthase), and FHs (flavonoid hydroxylases). Meanwhile, CHS (chalcone synthase) was down-regulated, causing a decrease in the content of most flavonoid glycosides. Pearson correlation analysis obtained 21 transcription factors co-expressed with differentially accumulated flavonoids, among which the AP2/EREBP members were the most numerous. Additionally, circadian rhythm and photosynthesis pathways were enriched in the DEG (differentially expressed gene) analysis, suggesting that melatonin might also mediate changes in the flavonoid biosynthesis pathway by affecting the fruit's circadian rhythm. These results provide valuable information for further exploration of the molecular mechanisms through which melatonin regulates citrus fruit metabolism.

FOXA2 prevents hyperbilirubinaemia in acute liver failure by maintaining apical MRP2 expression.

OBJECTIVE: Multidrug resistance protein 2 (MRP2) is a bottleneck in bilirubin excretion. Its loss is sufficient to induce hyperbilirubinaemia, a prevailing characteristic of acute liver failure (ALF) that is closely associated with clinical outcome. This study scrutinises the transcriptional regulation of MRP2 under different pathophysiological conditions. DESIGN: Hepatic MRP2, farnesoid X receptor (FXR) and Forkhead box A2 (FOXA2) expression and clinicopathologic associations were examined by immunohistochemistry in 14 patients with cirrhosis and 22 patients with ALF. MRP2 regulatory mechanisms were investigated in primary hepatocytes, Fxr -/- mice and lipopolysaccharide (LPS)-treated mice. RESULTS: Physiologically, homeostatic MRP2 transcription is mediated by the nuclear receptor FXR/retinoid X receptor complex. Fxr-/- mice lack apical MRP2 expression and rapidly progress into hyperbilirubinaemia. In patients with ALF, hepatic FXR expression is undetectable, however, patients without infection maintain apical MRP2 expression and do not suffer from hyperbilirubinaemia. These patients express FOXA2 in hepatocytes. FOXA2 upregulates MRP2 transcription through binding to its promoter. Physiologically, nuclear FOXA2 translocation is inhibited by insulin. In ALF, high levels of glucagon and tumour necrosis factor α induce FOXA2 expression and nuclear translocation in hepatocytes. Impressively, ALF patients with sepsis express low levels of FOXA2, lose MRP2 expression and develop severe hyperbilirubinaemia. In this case, LPS inhibits FXR expression, induces FOXA2 nuclear exclusion and thus abrogates the compensatory MRP2 upregulation. In both Fxr -/- and LPS-treated mice, ectopic FOXA2 expression restored apical MRP2 expression and normalised serum bilirubin levels. CONCLUSION: FOXA2 replaces FXR to maintain MRP2 expression in ALF without sepsis. Ectopic FOXA2 expression to maintain MRP2 represents a potential strategy to prevent hyperbilirubinaemia in septic ALF.

ETNPPL modulates hyperinsulinemia-induced insulin resistance through the SIK1/ROS-mediated inactivation of the PI3K/AKT signaling pathway in hepatocytes.

Hyperinsulinemia is a critical risk factor for the pathogenesis of insulin resistance (IR) in metabolic tissues, including the liver. Ethanolamine phosphate phospholyase (ETNPPL), a newly discovered metabolic enzyme that converts phosphoethanolamine (PEA) to ammonia, inorganic phosphate, and acetaldehyde, is abundantly expressed in liver tissue. Whether it plays a role in the regulation of hyperinsulinemia-induced IR in hepatocytes remains elusive. Here, we established an in vitro hyperinsulinemia-induced IR model in the HepG2 human liver cancer cell line and primary mouse hepatocyte via a high dose of insulin treatment. Next, we overexpressed ETNPPL by using lentivirus-mediated ectopic to investigate the effects of ETNPPL per se on IR without insulin stimulation. To explore the underlying mechanism of ETNPPL mediating hyperinsulinemia-induced IR in HepG2, we performed genome-wide transcriptional analysis using RNA sequencing (RNA-seq) to identify the downstream target gene of ETNPPL. The results showed that ETNPPL expression levels in both mRNA and protein were significantly upregulated in hyperinsulinemia-induced IR in HepG2 and primary mouse hepatocytes. Upon silencing ETNPPL, hyperinsulinemia-induced IR was ameliorated. Under normal conditions without IR in hepatocytes, overexpressing ETNPPL promotes IR, reactive oxygen species (ROS) generation, and AKT inactivation. Transcriptome analysis revealed that salt-inducible kinase 1 (SIK1) is markedly downregulated in the ETNPPL knockdown HepG2 cells. Moreover, disrupting SIK1 prevents ETNPPL-induced ROS accumulation, damage to the PI3K/AKT pathway and IR. Our study reveals that ETNPPL mediates hyperinsulinemia-induced IR through the SIK1/ROS-mediated inactivation of the PI3K/AKT signaling pathway in hepatocyte cells. Targeting ETNPPL may present a potential strategy for hyperinsulinemia-associated metabolic disorders such as type 2 diabetes.

Mixed invasive ductal lobular carcinoma is clinically and pathologically more similar to invasive lobular than ductal carcinoma.

BACKGROUND: Mixed invasive ductal lobular carcinoma (mDLC) remains a poorly understood subtype of breast cancer composed of coexisting ductal and lobular components. METHODS: We sought to describe clinicopathologic characteristics and determine whether mDLC is clinically more similar to invasive ductal carcinoma (IDC) or invasive lobular carcinoma (ILC), using data from patients seen at the University of Pittsburgh Medical Center. RESULTS: We observed a higher concordance in clinicopathologic characteristics between mDLC and ILC, compared to IDC. There is a trend for higher rates of successful breast-conserving surgery after neoadjuvant chemotherapy in patients with mDLC compared to patients with ILC, in which it is known to be lower than in those with IDC. Metastatic patterns of mDLC demonstrate a propensity to develop in sites characteristic of both IDC and ILC. A meta-analysis evaluating mDLC showed shared features with both ILC and IDC with significantly more ER-positive and fewer high grades in mDLC compared to IDC, although mDLCs were significantly smaller and included fewer late-stage tumours compared to ILC. CONCLUSIONS: These findings support clinicopathologic characteristics of mDLC driven by individual ductal vs lobular components and given the dominance of lobular pathology, mDLC features are often more similar to ILC than IDC. This study exemplifies the complexity of mixed disease.

A sparse negative binomial mixture model for clustering RNA-seq count data.

Clustering with variable selection is a challenging yet critical task for modern small-n-large-p data. Existing methods based on sparse Gaussian mixture models or sparse $K$-means provide solutions to continuous data. With the prevalence of RNA-seq technology and lack of count data modeling for clustering, the current practice is to normalize count expression data into continuous measures and apply existing models with a Gaussian assumption. In this article, we develop a negative binomial mixture model with lasso or fused lasso gene regularization to cluster samples (small $n$) with high-dimensional gene features (large $p$). A modified EM algorithm and Bayesian information criterion are used for inference and determining tuning parameters. The method is compared with existing methods using extensive simulations and two real transcriptomic applications in rat brain and breast cancer studies. The result shows the superior performance of the proposed count data model in clustering accuracy, feature selection, and biological interpretation in pathways.

A hierarchical regulatory network ensures stable albumin transcription under various pathophysiological conditions.

BACKGROUND AND AIMS: It remains unknown how patients with liver failure maintain essential albumin levels. Here, we delineate a hierarchical transcription regulatory network that ensures albumin expression under different disease conditions. APPROACH AND RESULTS: We examined albumin levels in liver tissues and serum in 157 patients, including 84 with HCC, 38 decompensated cirrhosis, and 35 acute liver failure. Even in patients with liver failure, the average serum albumin concentrations were 30.55 g/L. In healthy subjects and patients with chronic liver diseases, albumin was expressed in hepatocytes. In patients with massive hepatocyte loss, albumin was expressed in liver progenitor cells (LPCs). The albumin gene (ALB) core promoter possesses a TATA box and nucleosome-free area, which allows constitutive RNA polymerase II binding and transcription initiation. Chromatin immunoprecipitation assays revealed that hepatocyte nuclear factor 4 alpha (HNF4α), CCAAT/enhancer-binding protein alpha (C/EBPα), and forkhead box A2 (FOXA2) bound to the ALB enhancer. Knockdown of either of these factors reduced albumin expression in hepatocytes. FOXA2 acts as a pioneer factor to support HNF4α and C/EBPα. In hepatocytes lacking HNF4α and C/EBPα expression, FOXA2 synergized with retinoic acid receptor (RAR) to maintain albumin transcription. RAR nuclear translocation was induced by retinoic acids released by activated HSCs. In patients with massive hepatocyte loss, LPCs expressed HNF4α and FOXA2. RNA sequencing and quantitative PCR analyses revealed that lack of HNF4α and C/EBPα in hepatocytes increased hedgehog ligand biosynthesis. Hedgehog up-regulates FOXA2 expression through glioblastoma family zinc finger 2 binding to the FOXA2 promoter in both hepatocytes and LPCs. CONCLUSIONS: A hierarchical regulatory network formed by master and pioneer transcription factors ensures essential albumin expression in various pathophysiological conditions.

Synchronized time lens based temporal magnifier assisted by the phase lock loop.

We propose a synchronized time lens based temporal magnifier for the characterization of an ultrafast pulse, which is assisted by the phase lock loop (PLL) to actively lock the repetition rates of the pump laser and signal laser. A feedback control system combining a proportional-integral (PI) circuit and a piezoelectric transducer (PZT) inside the signal laser cavity is used to synchronize the repetition rates between the pump and signal lights. Benefitting from the PLL technique, the temporal position of the signal pulse remains as the numerical aperture of the time lens system, and therefore it has a high short-time stability for pulse measurement. This synchronized time lens based temporal magnifier can record single-shot pulses within continuous round trips. By engineering the dispersion of the pump, signal, and idler lights, we demonstrate a 200× magnification of the signal pulse. Our technique offers a simple synchronized method in a time lens system for ultrafast temporal characterization, which provides new insights into the observation of the fiber laser dynamics.

Induction of DNMT1-dependent demethylation of SHP-1 by the natural flavonoid compound Baicalein overcame Imatinib-resistance in CML CD34+ cells.

BACKGROUND: The most significant cause of treatment failure in chronic myeloid leukemia (CML) is a persistent population of minimal residual cells. Emerging evidences showed that methylation of SHP-1 contributed to Imatinib (IM) resistance. Baicalein was reported to have an effect on reversal of chemotherapeutic agents resistance. However, the molecular mechanism of Baicalein on JAK2/STAT5 signaling inhibition against drug resistance in bone marrow (BM) microenvironment that had not been clearly revealed. METHODS: We co-cultured hBMSCs and CML CD34+ cells as a model of SFM-DR. Further researches were performed to clarify the reverse mechanisms of Baicalein on SFM-DR model and engraftment model. The apoptosis, cytotoxicity, proliferation, GM-CSF secretion, JAK2/STAT5 activity, the expression of SHP-1 and DNMT1 were analyzed. To validate the role of SHP-1 on the reversal effect of Baicalein, the SHP-1 gene was over-expressed by pCMV6-entry shp-1 and silenced by SHP-1 shRNA, respectively. Meanwhile, the DNMT1 inhibitor decitabine was used. The methylation extent of SHP-1 was evaluated using MSP and BSP. The molecular docking was replenished to further explore the binding possibility of Baicalein and DNMT1. RESULTS: BCR/ABL-independent activation of JAK2/STAT5 signaling was involved in IM resistance in CML CD34+ subpopulation. Baicalein significantly reversed BM microenvironment-induced IM resistance not through reducing GM-CSF secretion, but interfering DNMT1 expression and activity. Baicalein induced DNMT1-mediated demethylation of the SHP-1 promoter region, and subsequently activated SHP-1 re-expression, which resulted in an inhibition of JAK2/STAT5 signaling in resistant CML CD34+ cells. Molecular docking model indicated that DNMT1 and Baicalein had binding pockets in 3D structures, which further supported Baicalein might be a small-molecule inhibitor targeting DNMT1. CONCLUSIONS: The mechanism of Baicalein on improving the sensitivity of CD34+ cells to IM might be correlated with SHP-1 demethylation by inhibition of DNMT1 expression. These findings suggested that Baicalein could be a promising candidate by targeting DNMT1 to eradicate minimal residual disease in CML patients. Video Abstract.

The real-time detection of acupuncture-induced extracellular ATP mobilization in acupoints and exploration of its role in acupuncture analgesia.

Our and in vitro studies had confirmed that mechanosensitive ATP release and accumulation in acupoints was elicited by acupuncture (AP), which might be a pivotal step for triggering AP analgesia. But to date, the dynamics of extracellular ATP (eATP) in the interstitial space during AP process was poorly known, mainly due to the low temporal resolution of the current detection approach. This study attempted to capture rapid eATP signals in vivo in the process of needling, and further explored the role of this eATP mobilization in initiating AP analgesic effect. Ipsilateral 20-min needling was applied on Zusanli acupoint (ST36) of complete Freund's adjuvant (CFA)-induced ankle arthritis rats. Pain thresholds were assessed in injured-side hindpaws. eATP in the interstitial space was microdialyzed and real-time quantified by luciferin-luciferase assay at 1-min interval with the aid of the microfluid chip. We revealed in behavioral tests that modulation of eATP levels in ST36 influenced AP analgesic effect on ankle arthritis. A transient eATP accumulation was induced by needling that started to mobilize at 4 min, climbed to the peak of 11.21 nM within 3.25 min and gradually recovered. Such AP-induced eATP mobilization was significantly impacted by ankle inflammation, needling depth, needle manipulation, and the presence of local ecto-nucleotidases. This work reveals that needling elicits a transient eATP mobilization in acupoints, which contributes to initiating AP analgesia. This study will help us better understand the peripheral mechanism of AP analgesia and guide clinicians to optimize the needle manipulations to improve AP efficacy.

COVID-19-associated liver injury: Adding fuel to the flame.

COVID-19 is mainly characterized by respiratory disorders and progresses to multiple organ involvement in severe cases. With expansion of COVID-19 and SARS-CoV-2 research, correlative liver injury has been revealed. It is speculated that COVID-19 patients exhibited abnormal liver function, as previously observed in the SARS and MERS pandemics. Furthermore, patients with underlying diseases such as chronic liver disease are more susceptible to SARS-CoV-2 and indicate a poor prognosis accompanied by respiratory symptoms, systemic inflammation, or metabolic diseases. Therefore, COVID-19 has the potential to impair liver function, while individuals with preexisting liver disease suffer from much worse infected conditions. COVID-19 related liver injury may be owing to direct cytopathic effect, immune dysfunction, gut-liver axis interaction, and inappropriate medication use. However, discussions on these issues are infancy. Expanding research have revealed that angiotensin converting enzyme 2 (ACE2) expression mediated the combination of virus and target cells, iron metabolism participated in the virus life cycle and the fate of target cells, and amino acid metabolism regulated immune response in the host cells, which are all closely related to liver health. Further exploration holds great significance in elucidating the pathogenesis, facilitating drug development, and advancing clinical treatment of COVID-19-related liver injury. This article provides a review of the clinical and laboratory hepatic characteristics in COVID-19 patients, describes the etiology and impact of liver injury, and discusses potential pathophysiological mechanisms.

Production of Tetramethylpyrazine from Cane Molasses by Bacillus sp. TTMP20.

2,3,5,6-Tetramethylpyrazine (TTMP) is an active ingredient of Ligusticum wallichii Franch. It can be used in medicine and food fields. In this study, Bacillus sp. TTMP20 was applied to produce TTMP using cane molasses as a carbon source. After pretreatment with phosphoric acid, 170 mL/L treated molasses, combined with 10 g/L yeast powder, 30 g/L tryptone and 30 g/L (NH4)2HPO4 were used for fermentation. After 36 h, TTMP output reached the highest value of 208.8 mg/L. The yield of TTMP using phosphoric acid-treated molasses as carbon source was 145.59% higher than control. Under the sulfuric acid treatment process of molasses (150 g), the maximum yield of TTMP was 895.13 mg/L, which was 183.18% higher than that of untreated molasses (316.1 mg/L). This study demonstrated that molasses is a high-quality and inexpensive carbon source for the manufacture of TTMP, laying the groundwork for the future industrial production of TTMP.