Long non-coding ribonucleic acid W5 inhibits progression and predicts favorable prognosis in hepatocellular carcinoma

BACKGROUND Accumulating evidence has revealed that several long non-coding ribonucleic acids(lncRNAs)are crucial in the progress of hepatocellular carcinoma(HCC).AIM To classify a long non-coding RNA,i.e.,lncRNA W5,and to determine the clinical significance and potential roles of lncRNA W5 in HCC.METHODS The results showed that lncRNA W5 expression was significantly downregulated in HCC cell lines and tissues.Analysis of the association between lncRNA W5 expression levels and clinicopathological features suggested that low lncRNA W5 expression was related to large tumor size(P<0.01),poor histological grade(P<0.05)and serious portal vein tumor thrombosis(P<0.05).Furthermore,Kaplan-Meier survival analysis showed that low expression of lncRNA W5 predicts poor overall survival(P=0.016).RESULTS Gain-of-loss function experiments,including cell counting kit8 assays,colony formation assays,and transwell assays,were performed in vitro to investigate thebiological roles of lncRNA W5.In vitro experiments showed that ectopic overexpression of lncRNA W5 suppressed HCC cell proliferation,migration and invasion;conversely,silencing of lncRNA W5 promoted cell proliferation,migration and invasion.In addition,acting as a tumor suppressor gene in HCC,lncRNA W5 inhibited the growth of HCC xenograft tumors in vivo.CONCLUSION These results showed that lncRNA W5 is down-regulated in HCC,and it may suppress HCC progression and predict poor clinical outcomes in patients with HCC.LncRNA W5 may serve as a potential HCC prognostic biomarker in addition to a therapeutic target.

Blood exosomal micro ribonucleic acid profiling reveals the complexity of hepatocellular carcinoma and identifies potential biomarkers for differential diagnosis

BACKGROUND Hepatocellular carcinoma(HCC)is one of the leading causes of cancer-related deaths worldwide,but there is a shortage of effective biomarkers for its diagnosis.AIM To explore blood exosomal micro ribonucleic acids(miRNAs)as potential biomarkers for HCC diagnosis.RESULTS The principal component analysis suggested that daily alcohol consumption could alter the blood exosomal miRNA profiles of hepatitis B virus positive non-HCC patients through miR-3168 and miR-223-3p.The miRNA profiles also revealed the tumor stages of HCC patients.High expression of miR-455-5p and miR-30c-5p,which significantly correlated with better overall survival in tumor tissues,could also be detected in blood exosomes.Two pairs of miRNAs(miR-584-5p/miR-106-3p and miR-628-3p/miR-941)showed a 94.1%sensitivity and 68.4%specificity to differentiate HCC patients from non-HCC patients.The specificity of the combination was substantially influenced by alcohol consumption habits.CONCLUSION This study suggested that blood exosomal miRNAs can be used as new noninvasive diagnostic tools for HCC.However,their accuracy could be affected by tumor stage and alcohol consumption habits.

S100 calcium binding protein A6 and associated long noncoding ribonucleic acids as biomarkers in the diagnosis and staging of primary biliary cholangitis

BACKGROUND Primary biliary cholangitis(PBC)is a chronic and slowly progressing cholestatic disease,which causes damage to the small intrahepatic bile duct by immunoregulation,and may lead to cholestasis,liver fibrosis,cirrhosis and,eventually,liver failure.AIM To explore the potential diagnosis and staging value of plasma S100 calcium binding protein A6(S100A6)messenger ribonucleic acid(mRNA),LINC00312,LINC00472,and LINC01257 in primary biliary cholangitis.METHODS A total of 145 PBC patients and 110 healthy controls(HCs)were enrolled.Among them,80 PBC patients and 60 HCs were used as the training set,and 65 PBC patients and 50 HCs were used as the validation set.The relative expression levels of plasma S100A6 mRNA,long noncoding ribonucleic acids LINC00312,LINC00472 and LINC01257 were analyzed using quantitative reverse transcription-polymerase chain reaction.The bile duct ligation(BDL)mouse model was used to simulate PBC.Then double immunofluorescence was conducted to verify the overexpression of S100A6 protein in intrahepatic bile duct cells of BDL mice.Human intrahepatic biliary epithelial cells were treated with glycochenodeoxycholate to simulate the cholestatic environment of intrahepatic biliary epithelial cells in PBC.RESULTS The expression of S100A6 protein in intrahepatic bile duct cells was up-regulated in the BDL mouse model compared with sham mice.The relative expression levels of plasma S100A6 mRNA,log10 LINC00472 and LINC01257 were upregulated while LINC00312 was down-regulated in plasma of PBC patients compared with HCs(3.01±1.04 vs 2.09±0.87,P<0.0001;2.46±1.03 vs 1.77±0.84,P<0.0001;3.49±1.64 vs 2.37±0.96,P<0.0001;1.70±0.33 vs 2.07±0.53,P<0.0001,respectively).The relative expression levels of S100A6 mRNA,LINC00472 and LINC01257 were up-regulated and LINC00312 was down-regulated in human intrahepatic biliary epithelial cells treated with glycochenodeoxycholate compared with control(2.97±0.43 vs 1.09±0.08,P=0.0018;2.70±0.26 vs 1.10±0.10,P=0.0006;2.23±0.21 vs 1.10±0.10,P=0.0011;1.20±0.04 vs 3.03±0.15,P<0.0001,respectively).The mean expression of S100A6 in the advanced stage(III and IV)of PBC was up-regulated compared to that in HCs and the early stage(II)(3.38±0.71 vs 2.09±0.87,P<0.0001;3.38±0.71 vs 2.57±1.21,P=0.0003,respectively);and in the early stage(II),it was higher than that in HCs(2.57±1.21 vs 2.09±0.87,P=0.03).The mean expression of LINC00312 in the advanced stage was lower than that in the early stage and HCs(1.39±0.29 vs 1.56±0.33,P=0.01;1.39±0.29 vs 2.07±0.53,P<0.0001,respectively);in addition,the mean expression of LINC00312 in the early stage was lower than that in HCs(1.56±0.33 vs 2.07±0.53,P<0.0001).The mean expression of log10 LINC00472 in the advanced stage was higher than those in the early stage and HCs(2.99±0.87 vs 1.81±0.83,P<0.0001;2.99±0.87 vs 1.77±0.84,P<0.0001,respectively).The mean expression of LINC01257 in both the early stage and advanced stage were up-regulated compared with HCs(3.88±1.55 vs 2.37±0.96,P<0.0001;3.57±1.79 vs 2.37±0.96,P<0.0001,respectively).The areas under the curves(AUC)for S100A6,LINC00312,log10 LINC00472 and LINC01257 in PBC diagnosis were 0.759,0.7292,0.6942 and 0.7158,respectively.Furthermore,the AUC for these four genes in PBC staging were 0.666,0.661,0.839 and 0.5549,respectively.The expression levels of S100A6 mRNA,log10 LINC00472,and LINC01257 in plasma of PBC patients were decreased(2.35±1.02 vs 3.06±1.04,P=0.0018;1.99±0.83 vs 2.33±0.96,P=0.036;2.84±0.92 vs 3.69±1.54,P=0.0006),and the expression level of LINC00312 was increased(1.95±0.35 vs 1.73±0.32,P=0.0007)after treatment compared with before treatment using the paired t-test.Relative expression of S100A6 mRNA was positively correlated with log10 LINC00472(r=0.683,P<0.0001);serum level of collagen type IV was positively correlated with the relative expression of log10 LINC00472(r=0.482,P<0.0001);relative expression of S100A6 mRNA was positively correlated with the serum level of collagen type IV(r=0.732,P<0.0001).The AUC for the four biomarkers obtained in the validation set were close to the training set.CONCLUSION These four genes may potentially act as novel biomarkers for the diagnosis of PBC.Moreover,LINC00472 acts as a potential biomarker for staging in PBC.

Nine-long non-coding ribonucleic acid signature can improve the survival prediction of colorectal cancer

BACKGROUND Investigating molecular biomarkers that accurately predict prognosis is of considerable clinical significance.Accumulating evidence suggests that long noncoding ribonucleic acids(lncRNAs)are frequently aberrantly expressed in colorectal cancer(CRC).AIM To elucidate the prognostic function of multiple lncRNAs serving as biomarkers in CRC.METHODS We performed lncRNA expression profiling using the lncRNA mining approach in large CRC cohorts from The Cancer Genome Atlas(TCGA)database.Receiver operating characteristic analysis was performed to identify the optimal cutoff point at which patients could be classified into the high-risk or low-risk groups.Based on the Cox coefficient of the individual lncRNAs,we identified a ninelncRNA signature that was associated with the survival of CRC patients in the training set(n=175).The prognostic value of this nine-lncRNA signature was validated in the testing set(n=174)and TCGA set(n=349).The prognostic models,consisting of these nine CRC-specific lncRNAs,performed well for risk stratification in the testing set and TCGA set.Time-dependent receiver operating characteristic analysis indicated that this predictive model had good performance.RESULTS Multivariate Cox regression and stratification analysis demonstrated that this nine-lncRNA signature was independent of other clinical features in predicting overall survival.Functional enrichment analysis of Kyoto Encyclopedia of Genes and Genomes pathways and Gene Ontology terms further indicated that these nine prognostic lncRNAs were closely associated with carcinogenesis-associated pathways and biological functions in CRC.CONCLUSION A nine-lncRNA expression signature was identified and validated that could improve the prognosis prediction of CRC,thereby providing potential prognostic biomarkers and efficient therapeutic targets for patients with CRC.

Association between homeobox protein transcript antisense intergenic ribonucleic acid genetic polymorphisms and cholangiocarcinoma

BACKGROUND Cholangiocarcinoma(CCA)represents a rare but highly aggressive malignancy that is often challenging to diagnose,especially in early stages.The role of existing tumor biomarkers for CCA diagnosis,remains controversial due to their low sensitivity and specificity.Increasing evidence has implicated long non-coding ribonucleic acid polymorphisms with cancer susceptibility in a variety of tumor types.The association between long non-coding ribonucleic acid homeobox protein transcript antisense intergenic ribonucleic acid(HOTAIR)polymorphisms and CCA risk has not been reported yet.AIM To investigate the influence of HOTAIR variants on the risk of CCA development.METHODS We conducted a case-control study in which three HOTAIR single nucleotide polymorphisms(rs920778,rs4759314 and rs7958904)were genotyped in a Greek cohort.Our study population included 122 CCA patients(80 males and 42 females)and 165 healthy controls.The polymorphisms under investigation were examined in peripheral blood samples.RESULTS HOTAIR rs4759314 AG and GG genotypes were associated with a significantly increased CCA risk[P=0.004,odds ratio:3.13;95%confidence interval:1.65-5.91 and P=0.005,odds ratio:12.31;95%confidence interval:1.48-101.87,respectively].However,no significant associations of HOTAIR rs920778,and rs7958904 were detected.Similarly,we found no significant associations between rs4759314 AA genotype and CCA susceptibility.CONCLUSION HOTAIR rs4759314 AG and GG genotypes may be implicated with CCA development and may serve as a potential diagnostic biomarker.

Potential role of micro ribonucleic acids in screening for anal cancer in human papilloma virus and human immunodeficiency virus related malignancies

Despite advances in antiretroviral treatment(ART),human immunodeficiency virus(HIV)continues to be a major global public health issue owing to the increased mortality rates related to the prevalent oncogenic viruses among people living with HIV(PLWH).Human papillomavirus(HPV)is the most common sexually transmitted viral disease in both men and women worldwide.High-risk or oncogenic HPV types are associated with the development of HPV-related malignancies,including cervical,penile,and anal cancer,in addition to oral cancers.The incidence of anal squamous cell cancers is increasing among PLWH,necessitating the need for reliable screening methods in this population at risk.In fact,the currently used screening methods,including the Pap smear,are invasive and are neither sensitive nor specific.Investigators are interested in circulatory and tissue micro ribonucleic acids(miRNAs),as these small non-coding RNAs are ideal biomarkers for early detection and prognosis of cancer.Multiple miRNAs are deregulated during HIV and HPV infection and their deregulation contributes to the pathogenesis of disease.Here,we will review the molecular basis of HIV and HPV co-infections and focus on the pathogenesis and epidemiology of anal cancer in PLWH.The limitations of screening for anal cancer and the need for a reliable screening program that involves specific miRNAs with diagnostic and therapeutic values is also discussed.

Research Progress on the Integrated Detection Technology for Forensic Deoxyribonucleic Acid Genetic Markers and Ribonucleic Acid Molecular Markers

Deoxyribonucleic acid(DNA)genetic markers and ribonucleic acid(RNA)molecular markers have been widely used in forensic practices including individual identification,parentage testing,body fluid identification,determination of the age of stains,and molecular pathological diagnosis.Variant information of biological evidence and their interrelation could be revealed by the integrated detection of DNA/RNA markers.The integrated detection workflow aims to simplify working procedures,reduce time consuming and save valuable samples collected from crime scenes.Next-generation sequencing(NGS)may be an effective method for integrated DNA/RNA detection.In this review,DNA/RNA co-extraction strategies,simultaneous detection methods based on capillary electrophoresis were summarized.Research on NGS-based integrated detection methods of DNA and RNA markers was reviewed to provide a reference for forensic medicine researches and applications.

Circulating cell-free nucleic acids as prognostic and therapy predictive tools for metastatic castrate-resistant prostate cancer

Metastatic castrate-resistant prostate cancer remains a disease hard to cure,and for this reason predictive tools to monitor disease progression and therapy response are an urgent need.In this respect,liquid biopsy on circulating cell-free nucleic acids represents an interesting strategy based on robust data.The low invasiveness and the possibility to target circulating cell-free tumor deoxyribonucleic acid underline the high specificity,sensitivity and clinical usability of the technique.Moreover,it has been observed that the cell-free tumor deoxyribonucleic acid of metastatic castrate-resistant prostate cancer patients can be representative of the tumor heterogeneity.Cell-free tumor deoxyribonucleic acids express the same behaviors as mutations:Variation in gene copy number or the methylation rate of the tumor tissue.Recently,circulating cell-free ribonucleic acid molecules have emerged as interesting markers to stratify the disease.Due to high-throughput technologies,liquid biopsy on circulating cell-free nucleic acids will soon be utilized in the clinical management of metastatic castrate-resistant prostate cancer patients.

Sequential extraction of RNA,DNA and protein from cultured cells of the same group

BACKGROUND Efficient extraction of nucleic acids and proteins(ENAP)from cells is a prerequisite for precise annotation of gene function,and has become laboratory routine for revealing the mysteries of life.However,cell samples are often from different culture dishes,resulting in inevitable experimental errors and sometimes poor repeatability.AIM To explore a method to improve the efficiency of ENAP,minimizing errors in ENAP processes,enhancing the reliability and repeatability of subsequent experimental results.METHODS A protocol for the sequential isolation of RNA,DNA,and proteins from the same cultured HepG2 cells using RNAzol reagent is presented here.The first step involves culturing HepG2 cells to the exponential phase,followed by the sequential isolation of RNA,DNA,and proteins from the same cultured cells in the second step.The yield of nucleic acids and proteins is detected in the third step,and their purity and integrity are verified in the last step.RESULTS The procedure takes as few as 3-4 d from the start to quality verification and is highly efficient.In contrast to the existing kits and reagents,which are primarily based on independent isolation,this RNAzol reagent-based method is characterized by the sequential isolation of RNA,DNA,and proteins from the same cells,and therefore saves time,and has low cost and high efficiency.CONCLUSION The RNA,DNA,and proteins isolated using this method can be used for reverse transcription-polymerase chain reaction,polymerase chain reaction,and western blotting,respectively.

Nanotechnology-based gene therapy as a credible tool in the treatment of Alzheimer’s disease

Toxic aggregated amyloid-βaccumulation is a key pathogenic event in Alzheimer’s disease.Treatment approaches have focused on the suppression,deferral,or dispersion of amyloid-βfibers and plaques.Gene therapy has evolved as a potential therapeutic option for treating Alzheimer’s disease,owing to its rapid advancement over the recent decade.Small interfering ribonucleic acid has recently garnered considerable attention in gene therapy owing to its ability to down-regulate genes with high sequence specificity and an almost limitless number of therapeutic targets,including those that were once considered undruggable.However,lackluster cellular uptake and the destabilization of small interfering ribonucleic acid in its biological environment restrict its therapeutic application,necessitating the development of a vector that can safeguard the genetic material from early destruction within the bloodstream while effectively delivering therapeutic genes across the bloodbrain barrier.Nanotechnology has emerged as a possible solution,and several delivery systems utilizing nanoparticles have been shown to bypass key challenges regarding small interfering ribonucleic acid delivery.By reducing the enzymatic breakdown of genetic components,nanomaterials as gene carriers have considerably enhanced the efficiency of gene therapy.Liposomes,polymeric nanoparticles,magnetic nanoparticles,dendrimers,and micelles are examples of nanocarriers that have been designed,and each has its own set of features.Furthermore,recent advances in the specific delivery of neurotrophic compounds via gene therapy have provided promising results in relation to augmenting cognitive abilities.In this paper,we highlight the use of different nanocarriers in targeted gene delivery and small interfering ribonucleic acid-mediated gene silencing as a potential platform for treating Alzheimer’s disease.

Comprehensive molecular analysis to predict the efficacy ofchemotherapy containing bevacizumab in patients with metastaticcolorectal cancer

Background:Although bevacizumab is an important treatment for metastatic colorectal cancer(CRC),not allpatients with CRC benefit from it;in unselected patient populations,only modest survival benefits have been reported.Methods:We evaluated clinical outcomes in 110 patients using comprehensive molecular characterization to identifybiomarkers for a response to bevacizumab-containing treatment.The molecular analysis comprised whole-exomesequencing,ribonucleic acid sequencing,and a methylation array on patient tissues.Results:Genomic and molecularcharacterization was successfully conducted in 103 patients.Six of 103 CRC samples were hypermutated,and none ofthe non-hypermutant tumors were microsatellite unstable.Among those 103 patients,89 had adenocarcinoma(ADC),15 were diagnosed with mucinous ADC,and six had signet-ring cell carcinoma(SRCC).Consensus molecular subtype(CMS)2 was unique to ADC.Of the four SRCCs,two were CMS1,one was CMS4,and the other was CMS3.APCmutation status was a significantly enriched factor in responders to bevacizumab treatment.Fibroblast growth factorreceptor(FGFR)1/2 signaling was upregulated in non-responders,whereas cell cycle,transfer ribonucleic acidprocessing,nucleotide excision repair,and oxidative phosphorylation pathways were enriched in responders.Inaddition,IGF1 was differentially expressed in non-responders(log2 fold change=−1.43,p=4.11×10^(−5),falsediscovery rate=0.098),and FLT1 was highly methylated in non-responders(p=7.55×10^(−3)).When the molecularpathways were reanalyzed separately according to the backbone chemotherapy(FOLFOX vs.FOLFIRI),thesignificance of the molecular pathways varied according to the backbone chemotherapy.Conclusions:This studysought a subset of CRC patients with a distinct clinical response to chemotherapy containing bevacizumab.Ourresults need to be validated in a large group of homogenous patient cohort and examined according to the differentchemotherapy backbones to create personalized therapeutic opportunities in CRC.

Molecular pathways of liver regeneration:A comprehensive review

The liver is a unique parenchymal organ with a regenerative capacity allowing it to restore up to 70%of its volume.Although knowledge of this phenomenon dates back to Greek mythology(the story of Prometheus),many aspects of liver regeneration are still not understood.A variety of different factors,including inflammatory cytokines,growth factors,and bile acids,promote liver regeneration and control the final size of the organ during typical regeneration,which is performed by mature hepatocytes,and during alternative regeneration,which is performed by recently identified resident stem cells called“hepatic progenitor cells”.Hepatic progenitor cells drive liver regeneration when hepatocytes are unable to restore the liver mass,such as in cases of chronic injury or excessive acute injury.In liver maintenance,the body mass ratio is essential for homeostasis because the liver has numerous functions;therefore,a greater understanding of this process will lead to better control of liver injuries,improved transplantation of small grafts and the discovery of new methods for the treatment of liver diseases.The current review sheds light on the key molecular pathways and cells involved in typical and progenitor-dependent liver mass regeneration after various acute or chronic injuries.Subsequent studies and a better understanding of liver regeneration will lead to the development of new therapeutic methods for liver diseases.

Exophiala dermatitidis

Exophiala is a genus comprising several species of opportunistic black yeasts,which belongs to Ascomycotina.It is a rare cause of fungal infections.However,infections are often chronic and recalcitrant,and while the number of cases is steadily increasing in both immunocompromised and immunocompetent people,detailed knowledge remains scarce regarding infection mechanisms,virulence factors,specific predisposing factors,risk factors,and host response.The most common manifestations of Exophiala infection are skin infections,and the most frequent type of deep infection is pulmonary infection due to inhalation.The invasive disease ranges from cutaneous or subcutaneous infection to systemic dissemination to internal organs.The final identification of the causative organism should be achieved through a combination of several methods,including the newly introduced diagnostic analysis,matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry,together with sequencing of the ribosomal ribonucleic acid internal transcribed spacer region of the fungi,and histological and culture findings.Regarding treatment,because anti-infective agents and natural compounds exhibited poor antibiofilm activity,few treatments have ultimately been found to be effective for specific antifungal therapy,so the optimal antifungal therapy and duration of therapy for these infections remain unknown.Therefore,most forms of disease caused by Exophiala dermatitidis require aggressive combination therapies:Both surgical intervention and aggressive antifungal therapy with novel compounds and azoles are necessary for effective treatment.

Probing Lanthanide Ions Binding on tRNA^(Phe) by ~1H NMR

The structure of phenylalanine transfer ribonucleic acid(tRNA Phe ) in solution was explored by 1 H NMR spectroscopy to evaluate the effect of lanthanide ion on the structural and conformational change. It was found that La 3+ ions possess specific effects on the imino proton region of the 1 H NMR spectra for yeast tRNA Phe . The dependence of the imino proton spectra of yeast tRNA Phe as a function of La 3+ concentration was examined, and the results suggest that the tertiary base pair G 15 ·C 48 , which is located in the terminal in the augmented dihydrouridine helix(D helix), was markedly affected by La 3+ (shifted to downfield by as much as 0 35). Base pair U 8·A 14 in yeast tRNA Phe , which are stacked on G 15 ·C 48 , was also affected by added La 3+ when 1～2 Mg 2+ were also present. Another imino proton that may be affected by La 3+ in yeast tRNA Phe is that of the tertiary base pair G 19 ·C 56 . The assignment of this resonance in yeast tRNA Phe is tentative since it is located in the region of highly overlapping resonances beween 12 6 and 12 2. This base pair helps to anchor the D loop to the TΨC loop. The binding of La 3+ caused conformational change of tRNA, which is responsible for shifts to upfield or downfield in 1 H NMR spectra.

Research Progress of microRNA in Regulating Gynecological Tumours

The incidence of gynaecological tumours,the main factor threatening women’s health and safety,is increasing with the daily heavier pressure of women’s life and work;Due to its serious threat to women,the disease has grasped increasingly more vigilance and attention.The discovery and research of micro ribonucleic acid(microRNA),a general term for a class of small-molecule RNA,showed that there was a close relationship between microRNA and the occurrence and development of gynaecological tumours.In this paper,based on the recent researches on the relationship between microRNA and gynaecological tumours,a comprehensive analysis was made to do a favour for scientific research and clinical treatment of gynaecological tumours.

Principles of amino-acid–ribonucleotide interaction revealed by binding affinities between homogeneous oligopeptides and singlestranded RNA molecules

We have determined the binding strengths between ribonucleotides of adenine(A),guanine(G),uracil(U),and cytosine(C)in homogeneous single-stranded ribonucleic acids(ssRNAs)and homo-decapeptides consisting of 20 common amino acids.We use a bead-based fluorescence assay for these measurements in which decapeptides are immobilized on the bead surface and ssRNAs are in solutions.The results provide a molecular basis for analyzing selectivity,specificity,and polymorphisms of amino-acid–ribonucleotide interactions.Comparative analyses of the distribution of the binding energies reveal unique binding strength patterns assignable to each pair of amino acid and ribonucleotide originating from the chemical structures.Pronounced favorable(such as Arg–G)and unfavorable(such as Met–U)binding interactions can be identified in selected groups of amino acid and ribonucleotide pairs that could provide basis to elucidate energetics of amino-acid–ribonucleotide interactions.Such interaction selectivity,specificity,and polymorphism manifest the contributions from RNA backbone,RNA bases,as well as main chain and side chain of the amino acids.Such characteristics in peptide–RNA interactions might be helpful for understanding the mechanism of protein–RNA specific recognition and the design of RNA nano-delivery systems based on peptides and their derivatives.

Complex RNA-DNA hybrid nanoshapes from iterative mix-andmatch screening

Hybrid nucleic acid nanostructures partition architectural and functional roles between ribonucleic acid(RNA)joints and deoxyribonucleic acid(DNA)connectors.Nanoshapes self-assemble from nucleic acid modules through synergistic stabilization of marginally stable base pairing interactions within circularly closed polygons.Herein,we report the development of hybrid nanoshapes that include multiple different RNA modules such as internal loop and three-way junction(3WJ)motifs.An iterative mix-and-match screening approach was used to identify suitable DNA connectors that furnished stable nanoshapes for combinations of different RNA modules.The resulting complex multicomponent RNA-DNA hybrid nanoshapes were characterized by atomic force microscopy(AFM)imaging.Our research provides proof of concept for modular design,assembly and screening of RNA-DNA hybrid nanoshapes as building blocks for complex extended nucleic acid materials with features at the sub-10 nm scale.

Effect of lanthanum ions on tRNA^(Phe) structure: Imino proton NMR spectroscopy

The effect of lanthanum ions on the structural and conformational change of yeast tRNA Phe was studied by 1H NMR. The results suggest that the tertiary base pair (G-15)(C-48), which was located in the terminal in the augmented dihydrouridine helix (D-helix), was markedly affected by adding La 3+ and shifted 0.33 downfield. Based pair (U-8)(A-14), which is associated with a tertiary interaction, links the base of the acceptor stem to the D-stem and anchors the elbow of the L structure, shifted 0.20 upfield. Another imino proton that may be affected by La 3+ in tRNA Phe is the tertiary base pair (G-19)(C-56). The assignment of this resonance is tentative since it is located in the region of highly overlapping resonances between 12.6 and 12.2. This base pair helps to anchor the D-loop to the TΨC loop.

Electrically controlled mRNA delivery using a polypyrrolegraphene oxide hybrid film to promote osteogenic differentiation of human mesenchymal stem cells

Direct messenger ribonucleic acid(mRNA)delivery to target cells or tissues has revolutionized the field of biotechnology.However,the applicability of regenerative medicine is limited by the technical difficulties of various mRNA-loaded nanocarriers.Herein,we report a new conductive hybrid film that could guide osteogenic differentiation of human adipose-derived mesenchymal stem cells(hADMSCs)via electrically controlled mRNA delivery.To find optimal electrical conductivity and mRNAloading capacity,the polypyrrole-graphene oxide(PPy-GO)hybrid film was electropolymerized on indium tin oxide substrates.We found that the fluorescein sodium salt,a molecule partially mimicking the physical and chemical properties of mRNAs,can be effectively absorbed and released by electrical stimulation(ES).The hADMSCs cultivated on the PPy-GO hybrid film loaded with pre-osteogenic mRNAs showed the highest osteogenic differentiation under electrical stimulation.This platform can load various types of RNAs thus highly promising as a new nucleic acid delivery tool for the development of stem cell-based therapeutics.

3D RNA nanocage for encapsulation and shielding of hydrophobic biomolecules to improve the in vivo biodistribution

Ribonucleic acid(RNA)nanotechnology platforms have the potential of harboring therapeutics for in vivo delivery in disease treatment.However,the nonspecific interaction between the harbored hydrophobic drugs and cells or other components before reaching the diseased site has been an obstacle in drug delivery.Here we report an encapsulation strategy to prevent such nonspecific hydrophobic interactions in vitro and in vivo based on a self-assembled three-dimensional(3D)RNA nanocage.By placing an RNA three-way junction(3WJ)in the cavity of the nanocage,the conjugated hydrophobic molecules were specifically positioned within the nanocage,preventing their exposure to the biological environment.The assembly of the nanocages was characterized by native polyacrylamide gel electrophoresis(PAGE),atomic force microscopy(AFM),and cryogenic electron microscopy(cryo-EM)imaging.The stealth effect of the nanocage for hydrophobic molecules in vitro was evaluated by gel electrophoresis,flow cytometry,and confocal microscopy.The in vivo sheathing effect of the nanocage for hydrophobic molecules was assessed by biodistribution profiling in mice.The RNA nanocages with hydrophobic biomolecules underwent faster clearance in liver and spleen in comparison to their counterparts.Therefore,this encapsulation strategy holds promise for in vivo delivery of hydrophobic drugs for disease treatment.