Direct comparison of the specificity of gene silencing using antisense oligonucleotides and RNAi

RNAi (RNA interference) and ASO (antisense oligonucleotide) technologies are the most commonly used approaches for silencing gene expression. However, the specificity of such powerful tools is an important factor to correctly interpret the biological consequences of gene silencing. In the present study, we examined the effects of acute loss of Ser/Thr kinase PDK1 (3-phosphoinositide-dependent kinase 1) expression using ASO and RNAi, and compared, for the first time, these two techniques using Affymetrix microarrays. We show that both ASO- and siRNA (small interfering RNA)-mediated knock-down of PDK1 expression strongly inhibited cell proliferation, although by different mechanisms, thereby questioning the specificity of these reagents. Using microarray analysis, we characterized the specificity of the ASO- and siRNA-mediated gene silencing of PDK1 by examining expression profiles 48 and 72 h following oligonucleotide transfection. At 48 h, a PDK1-dependent pattern of gene alterations was detectable, despite a large number of non-specific changes due to transfection of control nucleic acids. These non-specific alterations became more apparent at the 72 h time point, and obscured any PDK1-specific pattern. This study underscores the importance of defining appropriate control ASOs and siRNAs, using multiple oligonucleotides for each target and preferably short time points following transfection to avoid misinterpretation of the phenotype observed.

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Efficacy of compressed sodium chloride (CSC) against E. coli and Candida auris in minutes and methods improvement for testing

Scientific Reports ◽

10.1038/s41598-020-79212-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Lan N. Truong ◽

Brayden D. Whitlock

Keyword(s):

Sodium Chloride ◽

Improved Method ◽

Candida Auris ◽

Antimicrobial Surfaces ◽

E Coli ◽

Route Of Transmission ◽

The World ◽

Short Time ◽

First Time ◽

Time Point

AbstractControlling infections has become one of the biggest problems in the world, whether measured in lives lost or money spent. This is worsening as pathogens continue becoming resistant to therapeutics. Antimicrobial surfaces are one strategy being investigated in an attempt to decrease the spread of infections through the most common route of transmission: surfaces, including hands. Regulators have chosen two hours as the time point at which efficacy should be measured. The objectives of this study were to characterize the new antimicrobial surface compressed sodium chloride (CSC) so that its action may be understood at timepoints more relevant to real-time infection control, under two minutes; to develop a sensitive method to test efficacy at short time points; and to investigate antifungal properties for the first time. E. coli and Candida auris are added to surfaces, and the surfaces are monitored by contact plate, or by washing into collection vats. An improved method of testing antimicrobial efficacy is reported. Antimicrobial CSC achieves at least 99.9% reduction of E. coli in the first two minutes of contact, and at least 99% reduction of C. auris in one minute.

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Faculty Opinions recommendation of Molecular-timetable methods for detection of body time and rhythm disorders from single-time-point genome-wide expression profiles.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1021199.242550 ◽

2004 ◽

Author(s):

Victor Reus

Keyword(s):

Expression Profiles ◽

Single Time ◽

Genome Wide ◽

Genome Wide Expression ◽

Single Time Point ◽

Time Point

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Faculty Opinions recommendation of Direct comparison of the specificity of gene silencing using antisense oligonucleotides and RNAi.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1030460.359446 ◽

2006 ◽

Author(s):

Brent Cochran

Keyword(s):

Gene Silencing ◽

Antisense Oligonucleotides

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Faculty Opinions recommendation of Efficient gene silencing by delivery of locked nucleic acid antisense oligonucleotides, unassisted by transfection reagents.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1166571.627578 ◽

2009 ◽

Author(s):

Fritz Eckstein

Keyword(s):

Nucleic Acid ◽

Gene Silencing ◽

Antisense Oligonucleotides ◽

Locked Nucleic Acid ◽

Efficient Gene

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Advances in the Design of (Nano)Formulations for Delivery of Antisense Oligonucleotides and Small Interfering RNA: Focus on the Central Nervous System

Molecular Pharmaceutics ◽

10.1021/acs.molpharmaceut.0c01238 ◽

2021 ◽

Author(s):

Monique C. P. Mendonça ◽

Ayse Kont ◽

Maria Rodriguez Aburto ◽

John F. Cryan ◽

Caitriona M. O’Driscoll

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Antisense Oligonucleotides ◽

Small Interfering Rna ◽

The Central Nervous System ◽

Interfering Rna

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Bactericidal activity of three different antiseptic ophthalmic preparations as surgical prophylaxis

Graefe s Archive for Clinical and Experimental Ophthalmology ◽

10.1007/s00417-021-05361-3 ◽

2021 ◽

Author(s):

Daniele Tognetto ◽

Marco R. Pastore ◽

Gian Marco Guerin ◽

Giuliana Decorti ◽

Martina Franzin ◽

...

Keyword(s):

Antimicrobial Activity ◽

Povidone Iodine ◽

Bacterial Load ◽

Viable Cells ◽

Polyethylene Glycol 1000 ◽

Oil Concentration ◽

Short Time ◽

First Time ◽

Antiseptic Solutions

Abstract Purpose In the era of antibiotic resistance, there is an increased interest in antiseptic solutions that might represent a reliable option for ocular surface disinfection. The objective of this study is to compare for the first time three different antiseptic ophthalmic preparations to assess their in vitro antimicrobial activity. Methods The antiseptic activity of three commercial ophthalmic solutions, IODIM (povidone-iodine 0.6% in hyaluronic acid vehicle—Medivis, Catania, Italy), OZODROP (nanoemulsion with ozonated oil—concentration not specified—FBVision, Ophthalmic Pharmaceuticals, Rome, Italy), and DROPSEPT (chlorhexidine 0.02% and vitamin E 0.5% Tocopherol Polyethylene Glycol 1000 Succinate—TPGS, Sooft Italia, Montegiorgio, Italy), was tested in vitro on six reference strains by time-killing assays. Viable cells were evaluated after 1, 15, 30 min; 2, 6, and 24 h exposure by seeding 100 µl of the suspension (or appropriate dilutions) on LB agar or Sabouraud-dextrose agar. All plates were incubated at 37 °C for 24 h and evaluated by manually counting the colonies. Results IODIM solution showed a very rapid microbicidal activity: the number of viable cells for all the tested strains was under the detection limit (less than 10 CFU/ml) already after 1 min exposure, and this result was maintained at every incubation time. The rapid antimicrobial activity of povidone-iodine was not replicated when testing the other two antiseptics. Conclusions The study reports the great efficacy in reducing bacterial load in a very short time of povidone-iodine 0.6% compared with other antiseptic preparations.

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Dynamics of the seroprevalence of SARS-CoV-2 antibodies among healthcare workers at a COVID-19 referral hospital in Milan, Italy

Occupational and Environmental Medicine ◽

10.1136/oemed-2020-107060 ◽

2021 ◽

pp. oemed-2020-107060

Author(s):

Laura Milazzo ◽

Alessia Lai ◽

Laura Pezzati ◽

Letizia Oreni ◽

Annalisa Bergna ◽

...

Keyword(s):

Healthcare Workers ◽

Referral Hospital ◽

Control Measures ◽

Referral Centre ◽

Single Centre ◽

Factors Associated ◽

Infection Control Measures ◽

Professional Exposure ◽

First Time ◽

Time Point

ObjectivesHealthcare workers (HCWs) are at high risk of developing SARS-CoV-2 infection. The aim of this single-centre prospective study was to evaluate the trend of SARS-CoV-2 seroprevalence in HCWs working at the primary referral centre for infectious diseases and bioemergencies (eg, COVID-19) in Northern Italy and investigate the factors associated with seroconversion.MethodsSix hundred and seventy-nine HCW volunteers were tested for anti-SARS-CoV-2 antibodies three times between 4 March and 27 May 2020 and completed a questionnaire covering COVID-19 exposure, symptoms and personal protective equipment (PPE) training and confidence at each time.ResultsSARS-CoV-2 seroprevalence rose from 3/679 to 26/608 (adjusted prevalence: 0.5%, 95% CI 0.1 to 1.7% and 5.4%, 95% CI 3.6 to 7.9, respectively) between the first two time points and then stabilised, in line with the curve of the COVID-19 epidemic in Milan. From the first time point, 61.6% of the HCWs had received training in the use of PPE and 17 (61.5%) of those who proved to be seropositive reported symptoms compatible with SARS-CoV-2 infection. Contacts with ill relatives or friends and self-reported symptoms were independently associated with an increased likelihood of seroconversion (p<0.0001 for both), whereas there was no significant association with professional exposure.ConclusionThe seroprevalence of SARS-CoV-2 among the HCWs at our COVID-19 referral hospital was low at the time of the peak of the epidemic. The seroconversions were mainly attributable to extrahospital contacts, probably because the hospital readily adopted effective infection control measures. The relatively high number of asymptomatic seropositive HCWs highlights the need to promptly identify and isolate potentially infectious HCWs.

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Time Dependent Gene Expression Changes in the Liver of Mice Treated with Benzene

Biomarker Insights ◽

10.4137/bmi.s590 ◽

2008 ◽

Vol 3 ◽

pp. BMI.S590 ◽

Cited By ~ 3

Author(s):

Han-Jin Park ◽

Jung Hwa Oh ◽

Seokjoo Yoon ◽

S.V.S. Rana

Keyword(s):

Gene Expression ◽

Expression Profiles ◽

Gene Expression Profiles ◽

General Purpose ◽

Time Dependent ◽

Expression Data ◽

Benzene Exposure ◽

Microarray Analyses ◽

First Time ◽

Affymetrix Gene Chip

Benzene is used as a general purpose solvent. Benzene metabolism starts from phenol and ends with p-benzoquinone and o-benzoquinone. Liver injury inducted by benzene still remains a toxicologic problem. Tumor related genes and immune responsive genes have been studied in patients suffering from benzene exposure. However, gene expression profiles and pathways related to its hepatotoxicity are not known. This study reports the results obtained in the liver of BALB/C mice (SLC, Inc., Japan) administered 0.05 ml/100 g body weight of 2% benzene for six days. Serum, ALT, AST and ALP were determined using automated analyzer (Fuji., Japan). Histopathological observations were made to support gene expression data. c-DNA microarray analyses were performed using Affymetrix Gene-chip system. After six days of benzene exposure, twenty five genes were down regulated whereas nineteen genes were up-regulated. These gene expression changes were found to be related to pathways of biotransformation, detoxification, apoptosis, oxidative stress and cell cycle. It has been shown for the first time that genes corresponding to circadian rhythms are affected by benzene. Results suggest that gene expression profile might serve as potential biomarkers of hepatotoxicity during benzene exposure.

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