Novel RNA molecular bioengineering technology efficiently produces functional miRNA agents.

Genome-derived microRNAs (miRNAs or miRs) govern posttranscriptional gene regulation and play important roles in various cellular processes and disease progression. While chemo-engineered miRNA mimics or biosimilars made in vitro are widely available and used, miRNA agents produced in vivo are emerging to closely recapitulate natural miRNA species for research. Our recent work has demonstrated the success of high-yield, in vivo production of recombinant miRNAs by using human tRNA (htRNA) fused precursor miRNA (pre-miR) carriers. In this study, we aim to compare the production of bioengineered RNA (BioRNA) molecules with glycyl versus leucyl htRNA fused hsa-pre-miR-34a carriers, namely, BioRNAGly and BioRNALeu, respectively, and perform the initial functional assessment. We designed, cloned, overexpressed, and purified a total of 48 new BioRNA/miRNAs, and overall expression levels, final yields, and purities were revealed to be comparable between BioRNAGly and BioRNALeu molecules. Meanwhile, the two versions of BioRNA/miRNAs showed similar activities to inhibit non-small cell lung cancer cell viability. Interestingly, functional analyses using model BioRNA/miR-7-5p demonstrated that BioRNAGly/miR-7-5p exhibited greater efficiency to regulate a known target gene expression (EGFR) than BioRNALeu/miR-7-5p, consistent with miR-7-5p levels released in cells. Moreover, BioRNAGly/miR-7-5p showed comparable or slightly greater activities to modulate MRP1 and VDAC1 expression, compared with miRCURY LNA miR-7-5p mimic. Computational modeling illustrated overall comparable 3D structures for exemplary BioRNA/miRNAs with noticeable differences in htRNA species and payload miRNAs. These findings support the utility of hybrid htRNA/hsa-pre-miR-34a as reliable carriers for RNA molecular bioengineering, and the resultant BioRNAs serve as functional biologic RNAs for research and development.

The Growing Class of Novel RNAi Therapeutics.

The clinical use of RNA interference (RNAi) molecular mechanisms has introduced a novel, growing class of RNA therapeutics capable of treating diseases by controlling target gene expression at the posttranscriptional level. With the newly approved nedosiran (Rivfloza), there are now six RNAi-based therapeutics approved by the United States Food and Drug Administration (FDA). Interestingly, five of the six FDA-approved small interfering RNA (siRNA) therapeutics [patisiran (Onpattro), lumasiran (Oxlumo), inclisiran (Leqvio), vutrisiran (Amvuttra), and nedosiran] were revealed to act on the 3'-untranslated regions of target mRNAs, instead of coding sequences, thereby following the common mechanistic action of genome-derived microRNAs (miRNA). Furthermore, three of the FDA-approved siRNA therapeutics [patisiran, givosiran (Givlaari), and nedosiran] induce target mRNA degradation or cleavage via near-complete rather than complete base-pair complementarity. These features along with previous findings confound the currently held characteristics to distinguish siRNAs and miRNAs or biosimilars, of which all converge in the RNAi regulatory pathway action. Herein, we discuss the RNAi mechanism of action and current criteria for distinguishing between miRNAs and siRNAs while summarizing the common and unique chemistry and molecular pharmacology of the six FDA-approved siRNA therapeutics. The term "RNAi" therapeutics, as used previously, provides a coherently unified nomenclature for broader RNAi forms as well as the growing number of therapeutic siRNAs and miRNAs or biosimilars that best aligns with current pharmacological nomenclature by mechanism of action. SIGNIFICANCE STATEMENT: The common and unique chemistry and molecular pharmacology of six FDA-approved siRNA therapeutics are summarized, in which nedosiran is newly approved. We point out rather a surprisingly mechanistic action as miRNAs for five siRNA therapeutics and discuss the differences and similarities between siRNAs and miRNAs that supports using a general and unified term "RNAi" therapeutics to align with current drug nomenclature criteria in pharmacology based on mechanism of action and embraces broader forms and growing number of novel RNAi therapeutics.

Green and Efficient Extraction of Phenolic Components from Plants with Supramolecular Solvents: Experimental and Theoretical Studies.

The supramolecular solvent (SUPRAS) has garnered significant attention as an innovative, efficient, and environmentally friendly solvent for the effective extraction and separation of bioactive compounds from natural resources. However, research on the use of a SUPRAS for the extraction of phenolic compounds from plants, which are highly valued in food products due to their exceptional antioxidant properties, remains scarce. The present study developed a green, ultra-sound-assisted SUPRAS method for the simultaneous determination of three phenolic acids in Prunella vulgaris using high-performance liquid chromatography (HPLC). The experimental parameters were meticulously optimized. The efficiency and antioxidant properties of the phenolic compounds obtained using different extraction methods were also compared. Under optimal conditions, the extraction efficiency of the SUPRAS, prepared with octanoic acid reverse micelles dispersed in ethanol-water, significantly exceeded that of conventional organic solvents. Moreover, the SUPRAS method demonstrated greater antioxidant capacity. Confocal laser scanning microscopy (CLSM) images revealed the spherical droplet structure of the SUPRAS, characterized by a well-defined circular fluorescence position, which coincided with the position of the phenolic acids. The phenolic acids were encapsulated within the SUPRAS droplets, indicating their efficient extraction capacity. Furthermore, molecular dynamics simulations combined with CLSM supported the proposed method's mechanism and theoretically demonstrated the superior extraction performance of the SUPRAS. In contrast to conventional methods, the higher extraction efficiency of the SUPRAS can be attributed to the larger solvent contact surface area, the formation of more types of hydrogen bonds between the extractants and the supramolecular solvents, and stronger, more stable interaction forces. The results of the theoretical studies corroborate the experimental outcomes.

The volatile organic compounds generated from the Maillard reaction between l-ascorbic acid and l-cysteine in hot compressed water.

BACKGROUND: Hot compressed water (HCW), also known as subcritical water (SCW), refers to high-temperature compressed water in a special physical and chemical state. It is an emerging technology for natural product extraction. The volatile organic compounds (VOCs) generated from the Maillard reaction between l-ascorbic acid (ASA) and l-cysteine (Cys) have attracted significant interest in the flavor and fragrance industry. This study aimed to explore the formation mechanism of VOCs from ASA and Cys and examine the effects of reaction parameters such as temperature, time, and pH in HCW. RESULTS: The identified VOCs were predominantly thiophene derivatives, polysulfides, and pyrazine derivatives in HCW. The findings indicated that thiophene derivatives were formed under various pH conditions, with polysulfide formation favored under acidic conditions and pyrazine derivative formation preferred under weak alkaline conditions, specifically at pH 8.0. CONCLUSION: The Maillard reaction between ASA and Cys mainly produced thiophene derivatives, polysulfides, and pyrazine derivatives in HCW. The generation mechanism was significantly dependent on the surrounding pH conditions. © 2024 Society of Chemical Industry.

[Analysis of differential metabolites of spikes of Prunella vulgaris at different stages based on UPLC-MS/MS].

Prunella vulgaris, aptly named for its withering at the summer solstice, displays significant variation in quality arising from differing harvest time. However, research on the chemical composition changes of its spikes at various stages is limited, and the specific metabolites remain unclear. In order to elucidate the metabolites and metabolic pathways of the spikes of P. vulgaris, the current study deployed ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) and targeted metabolomics to characterize the compound variability in the spikes of P. vulgaris across different periods. Multivariate statistical techniques such as principal component analysis(PCA) and orthogonal partial least squares-discriminant analysis(OPLS-DA) were used to identify the differences in metabolites, and relevant metabolic pathways were analyzed. A total of 602 metabolites were identified by metabolomics, of which organic acids and their derivatives were the most abundant, followed by flavonoids. Multiple differential metabolites, including p-hydroxybenzoic acids and gallic acids were identified based on variable importance in projection(VIP)>1 and P<0.05. The results of enrichment analysis suggested that isoflavonoids biosynthesis, aminobenzoate degradation, benzoate degradation, anthocyanins biosynthesis, metabolic pathways, microbial metabolism in different environments, secondary plant metabolite biosynthesis, tryptophan metabolism, and phenylpropanoid synthesis were the main metabolic pathways. These results intend to elucidate the dynamic changes of differential metabolites of P. vulgaris and provide a theoretical basis for further study of the harvesting mechanism of spikes of P. vulgaris.

RNAi-Based Therapeutics and Novel RNA Bioengineering Technologies.

RNA interference (RNAi) provides researchers with a versatile means to modulate target gene expression. The major forms of RNAi molecules, genome-derived microRNAs (miRNAs) and exogenous small interfering RNAs (siRNAs), converge into RNA-induced silencing complexes to achieve posttranscriptional gene regulation. RNAi has proven to be an adaptable and powerful therapeutic strategy where advancements in chemistry and pharmaceutics continue to bring RNAi-based drugs into the clinic. With four siRNA medications already approved by the US Food and Drug Administration (FDA), several RNAi-based therapeutics continue to advance to clinical trials with functions that closely resemble their endogenous counterparts. Although intended to enhance stability and improve efficacy, chemical modifications may increase risk of off-target effects by altering RNA structure, folding, and biologic activity away from their natural equivalents. Novel technologies in development today seek to use intact cells to yield true biologic RNAi agents that better represent the structures, stabilities, activities, and safety profiles of natural RNA molecules. In this review, we provide an examination of the mechanisms of action of endogenous miRNAs and exogenous siRNAs, the physiologic and pharmacokinetic barriers to therapeutic RNA delivery, and a summary of the chemical modifications and delivery platforms in use. We overview the pharmacology of the four FDA-approved siRNA medications (patisiran, givosiran, lumasiran, and inclisiran) as well as five siRNAs and several miRNA-based therapeutics currently in clinical trials. Furthermore, we discuss the direct expression and stable carrier-based, in vivo production of novel biologic RNAi agents for research and development. SIGNIFICANCE STATEMENT: In our review, we summarize the major concepts of RNA interference (RNAi), molecular mechanisms, and current state and challenges of RNAi drug development. We focus our discussion on the pharmacology of US Food and Drug Administration-approved RNAi medications and those siRNAs and miRNA-based therapeutics that entered the clinical investigations. Novel approaches to producing new true biological RNAi molecules for research and development are highlighted.

Recombinant Technologies Facilitate Drug Metabolism, Pharmacokinetics, and General Biomedical Research.

The development of safe and effective medications requires a profound understanding of their pharmacokinetic (PK) and pharmacodynamic properties. PK studies have been built through investigation of enzymes and transporters that drive drug absorption, distribution, metabolism, and excretion (ADME). Like many other disciplines, the study of ADME gene products and their functions has been revolutionized through the invention and widespread adoption of recombinant DNA technologies. Recombinant DNA technologies use expression vectors such as plasmids to achieve heterologous expression of a desired transgene in a specified host organism. This has enabled the purification of recombinant ADME gene products for functional and structural characterization, allowing investigators to elucidate their roles in drug metabolism and disposition. This strategy has also been used to offer recombinant or bioengineered RNA (BioRNA) agents to investigate the posttranscriptional regulation of ADME genes. Conventional research with small noncoding RNAs such as microRNAs (miRNAs) and small interfering RNAs has been dependent on synthetic RNA analogs that are known to carry a range of chemical modifications expected to improve stability and PK properties. Indeed, a novel transfer RNA fused pre-miRNA carrier-based bioengineering platform technology has been established to offer consistent and high-yield production of unparalleled BioRNA molecules from Escherichia coli fermentation. These BioRNAs are produced and processed inside living cells to better recapitulate the properties of natural RNAs, representing superior research tools to investigate regulatory mechanisms behind ADME. SIGNIFICANCE STATEMENT: This review article summarizes recombinant DNA technologies that have been an incredible boon in the study of drug metabolism and PK, providing investigators with powerful tools to express nearly any ADME gene products for functional and structural studies. It further overviews novel recombinant RNA technologies and discusses the utilities of bioengineered RNA agents for the investigation of ADME gene regulation and general biomedical research.

Common and unique alterations of functional connectivity in major depressive disorder and bipolar disorder.

OBJECTIVE: Major depressive disorder (MDD) and bipolar disorder (BD) are considered whole-brain disorders with some common clinical and neurobiological features. It is important to investigate neural mechanisms to distinguish between the two disorders. However, few studies have explored the functional dysconnectivity between the two disorders from the whole brain level. METHODS: In this study, 117 patients with MDD, 65 patients with BD, and 116 healthy controls completed resting-state functional magnetic resonance imaging (R-fMRI) scans. Both edge-based network construction and large-scale network analyses were applied. RESULTS: Results found that both the BD and MDD groups showed decreased FC in the whole brain network. The shared aberrant network across patients involves the visual network (VN), sensorimotor network (SMN), dorsal attention network (DAN), and ventral attention network (VAN), which is related to the processing of external stimuli. The default mode network (DMN) and the limbic network (LN) abnormalities were only found in patients with MDD. Furthermore, results showed the highest decrease in edges of patients with MDD in between-network FC in SMN-VN, whereas in VAN-VN of patients with BD. CONCLUSIONS: Our findings indicated that both MDD and BD are extensive abnormal brain network diseases, mainly aberrant in those brain networks correlated to the processing of external stimuli, especially the attention network. Specific altered functional connectivity also was found in MDD and BD groups, respectively. These results may provide possible trait markers to distinguish the two disorders.

CSNK2A2 promotes hepatocellular carcinoma progression through activation of NF-κB pathway.

INTRODUCTION AND OBJECTIVES: Breast and non-small cell lung cancers harbor an upregulated CSNK2A2 oncogene that encodes the protein kinase CK2 alpha', a catalytic subunit of the highly conserved serine/threonine kinase CK2. However, its role and biological significance in hepatocellular carcinoma (HCC) remains unclear. MATERIALS AND METHODS: Western-blotting and immunohistochemistry were used to measure the expression of CSNK2A2 in HCC tumor tissues and cell lines. CCK8, Hoechst staining, transwell, tube formation assay in vitro and nude mice experiments in vivo were used to measure the effects of CSNK2A2 on HCC proliferation, apoptosis, metastasis, angiogenesis and tumor formation. RESULTS: In the study, we showed that CSNK2A2 was highly expressed in HCC comparison with matched control tissues, and was linked with lower survival of patients. Additional experiments indicated that silencing of CSNK2A2 promoted HCC cell apoptosis, while inhibited HCC cells migrating, proliferating, angiogenesis both in vitro and in vivo. These effects were also accompanied by reduced expression of NF-κB target genes, including CCND1, MMP9 and VEGF. Moreover, treatment with PDTC counteracted the promotional effects of CSNK2A2 on HCC cells. CONCLUSIONS: Overall, our results suggested that CSNK2A2 could promote HCC progression by activating the NF-κB pathway, and this could serve as a biomarker for future prognostic and therapeutic applications.

Application of CUBE-STIR MRI and high-frequency ultrasound in contralateral cervical 7 nerve transfer surgery.

OBJECTIVE: The objective of the study was to investigate the feasibility of CUBE-SITR MRI and high-frequency ultrasound for the structural imaging of the brachial plexus to exclude neoplastic brachial plexopathy or structural variation and measure the lengths of anterior and posterior divisions of the C7 nerve, providing guidelines for surgeons before contralateral cervical 7 nerve transfer. METHODS: A total of 30 patients with CNS and 20 with brachial plexus injury were enrolled in this retrospective study. All patients underwent brachial plexus CUBE-STIR MRI and high-frequency ultrasound, and the lengths of the anterior and posterior divisions of C7 nerve were measured before surgery. Precise length of anterior and posterior divisions of contralateral C7 nerve was measured during surgery. RESULTS: MRI-measured lengths of anterior and posterior divisions of C7 nerves were positively correlated with that measured during surgery (anterior division, r = 0.94, p < .01; posterior division, r = 0.92, p < .01). High agreement was found between MRI-measured and intra-surgery measured length of anterior and posterior divisions of C7 nerve by BLAD-ALTMAN analysis. Ultrasonography could feasibly image supraclavicular C7 nerve and recognize small variant branches derived from middle trunk of C7 nerve root, which could be dissected intra-operatively and confirmed by electromyography during the procedure of contralateral C7 nerve transfer. CONCLUSION: CUBE-STIR MRI had advantages for the imaging of the brachial plexus and measurement of the length of root-trunk-anterior/posterior divisions of C7 nerve. The clinical role of ultrasonography may be a simple way of evaluating general condition of C7 nerve and provide guidelines for contralateral C7 nerve transfer surgery.

RNA Drugs and RNA Targets for Small Molecules: Principles, Progress, and Challenges.

RNA-based therapies, including RNA molecules as drugs and RNA-targeted small molecules, offer unique opportunities to expand the range of therapeutic targets. Various forms of RNAs may be used to selectively act on proteins, transcripts, and genes that cannot be targeted by conventional small molecules or proteins. Although development of RNA drugs faces unparalleled challenges, many strategies have been developed to improve RNA metabolic stability and intracellular delivery. A number of RNA drugs have been approved for medical use, including aptamers (e.g., pegaptanib) that mechanistically act on protein target and small interfering RNAs (e.g., patisiran and givosiran) and antisense oligonucleotides (e.g., inotersen and golodirsen) that directly interfere with RNA targets. Furthermore, guide RNAs are essential components of novel gene editing modalities, and mRNA therapeutics are under development for protein replacement therapy or vaccination, including those against unprecedented severe acute respiratory syndrome coronavirus pandemic. Moreover, functional RNAs or RNA motifs are highly structured to form binding pockets or clefts that are accessible by small molecules. Many natural, semisynthetic, or synthetic antibiotics (e.g., aminoglycosides, tetracyclines, macrolides, oxazolidinones, and phenicols) can directly bind to ribosomal RNAs to achieve the inhibition of bacterial infections. Therefore, there is growing interest in developing RNA-targeted small-molecule drugs amenable to oral administration, and some (e.g., risdiplam and branaplam) have entered clinical trials. Here, we review the pharmacology of novel RNA drugs and RNA-targeted small-molecule medications, with a focus on recent progresses and strategies. Challenges in the development of novel druggable RNA entities and identification of viable RNA targets and selective small-molecule binders are discussed. SIGNIFICANCE STATEMENT: With the understanding of RNA functions and critical roles in diseases, as well as the development of RNA-related technologies, there is growing interest in developing novel RNA-based therapeutics. This comprehensive review presents pharmacology of both RNA drugs and RNA-targeted small-molecule medications, focusing on novel mechanisms of action, the most recent progress, and existing challenges.

Acorus tatarinowii Schott: A Review of Its Botany, Traditional Uses, Phytochemistry, and Pharmacology.

Acorus tatarinowii Schott (A. tatarinowii) is a natural medicinal plant. It plays an indispensable role in the treatment of diseases by the empirical medicine system and has achieved remarkable curative effects. A. tatarinowii is often used to treat various diseases, such as depression, epilepsy, fever, dizziness, heartache, stomachache, etc. More than 160 compounds of different structural types have been identified in A. tatarinowii, including phenylpropanoids, terpenoids, lignans, flavonoids, alkaloids, amides, and organic acids. These bioactive ingredients make A. tatarinowii remarkable for its pharmacological effects, including antidepressant, antiepileptic, anticonvulsant, antianxiety, neuroprotective, antifatigue, and antifungal effects, improving Alzheimer's disease, and so on. It is noteworthy that A. tatarinowii has been widely used in the treatment of brain diseases and nervous system diseases and has achieved satisfactory therapeutic effects. This review focused on the research publications of A. tatarinowii and aimed to summarize the advances in the botany, traditional uses, phytochemistry, and pharmacology, which will provide a reference for further studies and applications of A. tatarinowii.

Nuclear receptor RORγ inverse agonists/antagonists display tissue- and gene-context selectivity through distinct activities in altering chromatin accessibility and master regulator SREBP2 occupancy.

The nuclear receptor RORγ is a major driver of autoimmune diseases and certain types of cancer due to its aberrant function in T helper 17 (Th17) cell differentiation and tumor cholesterol metabolism, respectively. Compound screening using the classic receptor-coactivator interaction perturbation scheme led to identification of many small-molecule modulators of RORγ(t). We report here that inverse agonists/antagonists of RORγ such as VTP-43742 derivative VTP-23 and TAK828F, which can potently inhibit the inflammatory gene program in Th17 cells, unexpectedly lack high potency in inhibiting the growth of TNBC tumor cells. In contrast, antagonists such as XY018 and GSK805 that strongly suppress tumor cell growth and survival display only modest activities in reducing Th17-related cytokine expression. Unexpectedly, we found that VTP-23 significantly induces the cholesterol biosynthesis program in TNBC cells. Our further mechanistic analyses revealed that VTP-23 enhances the local chromatin accessibility, H3K27ac mark and the cholesterol master regulator SREBP2 recruitment at the RORγ binding sites, whereas XY018 exerts the opposite activities. Yet, they display similar inhibitory effects on circadian rhythm program. Similar distinctions and contrasting activities between TAK828F and SR2211 in their effects on local chromatin structure at Il17 genes were also observed. Together, our study shows for the first-time that structurally distinct RORγ antagonists possess different or even contrasting activities in tissue/cell-specific manner. Our findings also highlight that the activities at natural chromatin are key determinants of RORγ modulators' tissue selectivity.

Mixed-Linker Strategy for the Construction of Metal-Organic Framework Combined with Dyes toward Alcohol Detection.

Herein, a N-rich metal-organic framework (MOF) with four kinds of cages, Zn4(ade)2(TCA)2(H2O) (NENU-1000, Hade = adenine, H3TCA = 4,4',4″-tricarboxytriphenylamine, NENU = Northeast Normal University), was prepared by the mixed-ligand strategy. Cationic dyes can be selectively absorbed by NENU-1000 at proper concentrations, but not neutral and anionic dyes, which perhaps can be assigned to the N-rich neutral framework of NENU-1000. When NENU-1000 was introduced to a relatively lower concentration of cationic dye solutions (e.g., rhodamine B or basic red 2), the colors of these systems faded quickly. Furthermore, the faded solutions can be used for the detection of methanol and other small alcohol molecules with either the naked eye or common UV-vis spectra. The effect of the length of carbon chain, the position of the -OH group, and the number of the hydroxyl group of the alcohols was explored for the color development rate. In addition, the performance of NENU-1000 in iodine sorption and release was also studied.

A Novel Integrated Pharmacokinetic-Pharmacodynamic Model to Evaluate Combination Therapy and Determine In Vivo Synergism.

Understanding pharmacokinetic (PK)-pharmacodynamic (PD) relationships is essential in translational research. Existing PK-PD models for combination therapy lack consideration of quantitative contributions from individual drugs, whereas interaction factor is always assigned arbitrarily to one drug and overstretched for the determination of in vivo pharmacologic synergism. Herein, we report a novel generic PK-PD model for combination therapy by considering apparent contributions from individual drugs coadministered. Doxorubicin (Dox) and sorafenib (Sor) were used as model drugs whose PK data were obtained in mice and fit to two-compartment model. Xenograft tumor growth was biphasic in mice, and PD responses were described by three-compartment transit models. This PK-PD model revealed that Sor (contribution factor = 1.62) had much greater influence on overall tumor-growth inhibition than coadministered Dox (contribution factor = 0.644), which explains the mysterious clinical findings on remarkable benefits for patients with cancer when adding Sor to Dox treatment, whereas there were none when adding Dox to Sor therapy. Furthermore, the combination index method was integrated into this predictive PK-PD model for critical determination of in vivo pharmacologic synergism that cannot be correctly defined by the interaction factor in conventional models. In addition, this new PK-PD model was able to identify optimal dosage combination (e.g., doubling experimental Sor dose and reducing Dox dose by 50%) toward much greater degree of tumor-growth inhibition (>90%), which was consistent with stronger synergy (combination index = 0.298). These findings demonstrated the utilities of this new PK-PD model and reiterated the use of valid method for the assessment of in vivo synergism. SIGNIFICANCE STATEMENT: A novel pharmacokinetic (PK)-pharmacodynamic (PD) model was developed for the assessment of combination treatment by considering contributions from individual drugs, and combination index method was incorporated to critically define in vivo synergism. A greater contribution from sorafenib to tumor-growth inhibition than that of coadministered doxorubicin was identified, offering explanation for previously inexplicable clinical observations. This PK-PD model and strategy shall have broad applications to translational research on identifying optimal dosage combinations with stronger synergy toward improved therapeutic outcomes.

Classical swine fever virus infection suppresses claudin-1 expression to facilitate its replication in PK-15 cells.

Classical swine fever (CSF) is one of the most epidemic viral diseases in swine industry. The causative pathogen is CSF virus (CSFV), a small enveloped RNA virus of Flaviviridae family. Claudin-1 was reported to be involved in the infections of a number of viruses, including many from Flaviviridae family, but no studies have investigated the role of porcine claudin-1 during CSFV infection in PK-15 cells. In this study, on the one hand, we demonstrated that CSFV infection reduced the claudin-1 expression at both mRNA and protein levels; on the other hand, CSFV infection was enhanced after claudin-1 knockdown, but inhibited by claudin-1 overexpression in a dose-dependent manner. Furthermore, negative correlation was demonstrated between the claudin-1 expression and CSFV titer. In conclusion, claudin-1 might be a barrier for CSFV infection in PK-15 cells, while CSFV bypasses the barrier through lysosome mediated degradation of claudin-1, which could be repressed by bafilomycin A1. Although the elaborate mechanisms how claudin-1 plays its roles in CSFV infection require further investigations, this study may advance our understanding of the molecular host-pathogen interaction mechanisms underlying CSFV infection and suggests enhancement of porcine claudin-1 as a potential preventive or therapeutic strategy for CSF control.

Role of miR-211 in a PC12 cell model of Alzheimer's disease via regulation of neurogenin 2.

NEW FINDINGS: What is the central question of this study? What is the mechanism of miR-211 in an Alzheimer's disease cell model? What is the main finding and its importance? miR-211 was upregulated in an Alzheimer's disease cell model. It targeted neurogenin 2, reduced the activation of the phosphoinositide 3-kinase-Akt signalling pathway, inhibited the proliferation of the Alzheimer's disease cell model and promoted apoptosis. ABSTRACT: MicroRNAs (miRs) are aberrantly expressed in Alzheimer's disease (AD) patients. This study was intended to investigate the effect of miR-211 on an AD cell model and the involvement of neurogenin 2 (Ngn2). The appropriate dose and time for the effect of Aß1-42 on PC12 cells were determined to establish an AD cell model. An effect of miR-211 expression on cell viability, proliferation and apoptosis was detected after cell transfection. Online prediction and a dual luciferase reporter gene assay were utilized to confirm the binding sequence of miR-211 and Ngn2. qRT-PCR and western blot analysis were applied to measure Ngn2 expression. A gain and loss of function assay of miR-211 and Ngn2 was performed, and activation of the phosphoinositide 3-kinase (PI3K)-Akt signaling pathway was detected. The AD cell model was induced by Aß1-42 treatment. miR-211 expression was significantly enhanced after miR-211 transfection, leading to suppressed proliferation and promotion of apoptosis in Aß1-42 -treated PC12 cells. In addition, miR-211 could downregulate Ngn2 mRNA and protein expression, while overexpression of Ngn2 could reverse the effects of miR-211 on Aß1-42 -treated PC12 cells and significantly enhance the phosphorylated Akt and PI3K protein levels. miR-211 could inhibit growth of PC12 cells by suppressing Ngn2 expression and inactivating the PI3K-Akt signalling pathway.

Application and Development of Noncontact Detection Method of α-Particles Based on Radioluminescence.

The detection of α particles is of great significance in military and civil nuclear facility management. At present, the contact method is mainly used to detect α particles, but its shortcomings limit the broad application of this method. In recent years, preliminary research on noncontact α-particle detection methods has been carried out. In this paper, the theory of noncontact α-particles detection methods is introduced and studied. We also review the direct detection and imaging methods of α particles based on the different wavelengths of fluorescence photons, and analyze the application and development of this method, providing an important reference for researchers to carry out related work.

MicroRNA-1291-5p Sensitizes Pancreatic Carcinoma Cells to Arginine Deprivation and Chemotherapy through the Regulation of Arginolysis and Glycolysis.

Cancer cells are dysregulated and addicted to continuous supply and metabolism of nutritional glucose and amino acids (e.g., arginine) to drive the synthesis of critical macromolecules for uncontrolled growth. Recent studies have revealed that genome-derived microRNA (miRNA or miR)-1291-5p (miR-1291-5p or miR-1291) may modulate the expression of argininosuccinate synthase (ASS1) and glucose transporter protein type 1 (GLUT1). We also developed a novel approach to produce recombinant miR-1291 agents for research, which are distinguished from conventional chemo-engineered miRNA mimics. Herein, we firstly demonstrated that bioengineered miR-1291 agent was selectively processed to high levels of target miR-1291-5p in human pancreatic cancer (PC) cells. After the suppression of ASS1 protein levels, miR-1291 perturbed arginine homeostasis and preferably sensitized ASS1-abundant L3.3 cells to arginine deprivation therapy. In addition, miR-1291 treatment reduced the protein levels of GLUT1 in both AsPC-1 and PANC-1 cells, leading to a lower glucose uptake (deceased > 40%) and glycolysis capacity (reduced approximately 50%). As a result, miR-1291 largely improved cisplatin efficacy in the inhibition of PC cell viability. Our results demonstrated that miR-1291 was effective to sensitize PC cells to arginine deprivation treatment and chemotherapy through targeting ASS1- and GLUT1-mediated arginolysis and glycolysis, respectively, which may provide insights into understanding miRNA signaling underlying cancer cell metabolism and development of new strategies for the treatment of lethal PC. SIGNIFICANCE STATEMENT: Many anticancer drugs in clinical use and under investigation exert pharmacological effects or improve efficacy of coadministered medications by targeting cancer cell metabolism. Using new recombinant miR-1291 agent, we revealed that miR-1291 acts as a metabolism modulator in pancreatic carcinoma cells through the regulation of argininosuccinate synthase- and glucose transporter protein type 1-mediated arginolysis and glycolysis. Consequently, miR-1291 effectively enhanced the efficacy of arginine deprivation (pegylated arginine deiminase) and chemotherapy (cisplatin), offering new insights into development of rational combination therapies.

Pharmacokinetic and Pharmacodynamic Factors Contribute to Synergism between Let-7c-5p and 5-Fluorouracil in Inhibiting Hepatocellular Carcinoma Cell Viability.

Pharmacological interventions for hepatocellular carcinoma (HCC) are hindered by complex factors, and rational combination therapy may be developed to improve therapeutic outcomes. Very recently, we have identified a bioengineered microRNA let-7c-5p (or let-7c) agent as an effective inhibitor against HCC in vitro and in vivo. In this study, we sought to identify small-molecule drugs that may synergistically act with let-7c against HCC. Interestingly, we found that let-7c exhibited a strong synergism with 5-fluorouracil (5-FU) in the inhibition of HCC cell viability as manifested by average combination indices of 0.3 and 0.5 in Hep3B and Huh7 cells, respectively. By contrast, coadministration of let-7c with doxorubicin or sorafenib inhibited HCC cell viability with, rather surprisingly, no or minimal synergy. Further studies showed that protein levels of multidrug resistance-associated protein (MRP) ATP-binding cassette subfamily C member 5 (MRP5/ABCC5), a 5-FU efflux transporter, were reduced around 50% by let-7c in HCC cells. This led to a greater degree of intracellular accumulation of 5-FU in Huh7 cells as well as the second messenger cyclic adenosine monophosphate, an endogenous substrate of MRP5. Since 5-FU is an irreversible inhibitor of thymidylate synthetase (TS), we investigated the interactions of let-7c with 5-FU at pharmacodynamic level. Interestingly, our data revealed that let-7c significantly reduced TS protein levels in Huh7 cells, which was associated with the suppression of upstream transcriptional factors as well as other regulatory factors. Collectively, these results indicate that let-7c interacts with 5-FU at both pharmacokinetic and pharmacodynamic levels, and these findings shall offer insight into molecular mechanisms of synergistic drug combinations. SIGNIFICANCE STATEMENT: Combination therapy is a common strategy that generally involves pharmacodynamic interactions. After identifying a strong synergism between let-7c-5p and 5-fluorouracil (5-FU) against hepatocellular carcinoma cell viability, we reveal the involvement of both pharmacokinetic and pharmacodynamic mechanisms. In particular, let-7c enhances 5-FU exposure (via suppressing ABCC5/MRP5 expression) and cotargets thymidylate synthase with 5-FU (let-7c reduces protein expression, whereas 5-FU irreversibly inactivates enzyme). These findings provide insight into developing rational combination therapies based on pharmacological mechanisms.