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Reading Papers ARRIVE Guidelines
METHODS…continued
7. Experimental procedures
For each experiment and each experimental group, including controls, provide precise details of all
procedures performed. For example:
a. How (e.g. drug formulation and dose, site and route of administration, anaesthesia and analgesia used
[including monitoring], surgical procedure, method of euthanasia). Provide details of any specialist
equipment used, including supplier(s).
b. When (e.g. time of day).
c. Where (e.g. home cage, laboratory, water maze).
d. Why (e.g. rationale for choice of specific anaesthetic, route of administration,
drug dose used).
Reading Papers ARRIVE Guidelines
__________________________________________________
Source Mice Source Innoculum No.EAE/Total
__________________________________________________
Company X Company X 1/20
Company X Baker Lab 9/10
Baker Lab Company X 4/10
Baker Lab Baker Lab 10/10
________________________________________________
6-8 week oude, vrouwelijke ABH muizen werden subcutaan in 2 plaatsen in de rug van de flank met 0.1ml een sonicated (10min in
Bransonic Ultrasonicator, Sigma Ltd, Poole, GB) emulsie van 1mg syngeneic ruggenmerg homogenate fosfaat beschermd zouthoudend
en de volledige hulp van Freund ingespoten die 60µg kobalt-gebestralen Mycobacterium tuberculose op dag 0 en 7.
6-8 week old, female ABH mice were injected subcutaneously in 2 sites in the dorsum of the flank with 0.1ml a sonicated (10min in
Bransonic Ultrasonicator, Sigma Ltd, Poole, UK) emulsion of 1mg syngeneic spinal cord homogenate phosphate buffered saline and
Freund’s complete adjuvant containing 60µg cobalt-irradiated Mycobacterium tuberculosis on day 0 and 7.
METHODS…continued
7. Experimental procedures….continued
Al-Izki S et al. Multiple Sclerosis and Related Disorders 2012;1: 29–38
Reading Papers ARRIVE Guidelines
METHODS…continued
8. Experimental animals
a. Provide details of the animals used, including species, strain, sex,
b. Provide further relevant information such as the source of animals,
International strain nomenclature,
Genetic modification status (e.g. knock-out or transgenic) & Genotype,
Health/immune status, Drug or Test naïve, Previous procedures, etc.
PROVIDE DETAILS OF DEMOGRAPHICS!
_____________________________________
Source No. EAE/Total
_____________________________________
In-house Bred ABH mice 10/10
Commercial Bred ABH mice 5/10
_____________________________________
Reading Papers ARRIVE Guidelines
METHODS…continued
9. Housing and husbandry (This part of the Ethical review process) & Environment Enrichment
a. Housing (e.g. type of facility,e.g. specific pathogen free [SPF]; type of cage or housing; bedding
material; number of cage companions.
b. Husbandry Conditions (e.g. breeding programme, light/dark cycle, temperature, quality of water, access
to food and water, environmental enrichment).
c. Welfare-Related Assessments and Interventions that were carried out prior to, during, or after the
experiment.
Reading Papers
METHODS…continued
10. Sample size
a. Specify the total number of animals used in each experiment and the number
of animals in each experimental group.
b. Explain how the number of animals was arrived at. Provide details of any sample size calculation used.
c. Indicate the number of independent replications of each experiment, if relevant.
POWER ANALYSIS!
https://www.nc3rs.org.uk/experimental-design-assistant-eda
Experimental Design Assistant
Vesterinen et al. Mult scler; 2010, 16:1044 2010
Reading Papers
METHODS…continued
11. Allocating Animals to Experimental Groups
a. Give full details of how animals were allocated to experimental groups, including randomisation or
matching if done.
b. Describe the order in which the animals in the different experimental groups were treated and assessed.
12. Experimental outcomes
Clearly define the primary and secondary experimental outcomes assessed
In Human Trials failure to change primary outcome measure is a FAILURE!](https://image.slidesharecdn.com/bakeresnihandoutsreadingpapers-190717105407/75/Baker-esni-handouts-reading-papers-7-2048.jpg)
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- 1. 17/07/2019 1 Professor David Baker (david.baker@qmul.ac.uk) ESNIAmsterdam 2019 Reading Papers • Paper Construction • Reporting & Understanding Basic Science & Clinical Papers www.ms-res.org • Understanding multiple sclerosis models Reading Papers CLINICAL TRIAL PAPERS RESULTS: EFFICACY & SAFETY LITTLE INTERPRETATION SCIENCE PAPERS RESULTS: RESULTS EVOLVE (OVER) INTERPRETATION MECHANISM OF ACTION RESEARCH PAPERS Often written by Professional Writers Reading Papers Questions to think about: • RELEVANCE • RELIABILITY • Are the methods appropriate to the study aims? • Is the Analysis appropriate? • Are the Data & Pictures convincing? • VALIDITY - How convincing are the authors arguments for their results in relations to others? DON’T BELIEVE EVERYTHING YOU READ READ READ, READ, READ Reading Papers KNOWLEDGE IS THE KEY TO GAINING KNOWLEDGE Allows You to Understand the Paper Allows You to Interpret the Results Allows you to Understand the Discussion Allows you to Place the Work in Context of its Relevance Allows you to see the flaws that may affect interpretation Allows you to Judge the Value of the Work Allows you to Exploit the Information from the work Allows you to learn from your and others mistakes GIVES YOU THE CONFIDENCE IN ALL OF THE ABOVE READ READ, READ, READ Reading Papers NON PEER-REVIEWED Patents Meeting Abstracts Internet/Web pages There is a lot of Rubbish Out there! Media Sensationalisation PEER-REVIEWED Reviews Lots of References, Brings Threads together, Sorts conflict Are citations broad or focused? Dogma/Expertise/Conflicts of Interest? Case Reports Clinical Trials Quality Class of Evidence Original Articles Quality & Supporting Evidence PERCIEVED Serious Science/Pop Science QUALITY OF JOURNAL TYPE OF RESEARCH PAPER Reading Papers CONSORT GUIDELINES (Human Trials): Consolidated Standards of Reporting Trials (www.consort-statement.org) ARRIVE GUIDELINES (Animals Studies) Animal Research: Reporting of In Vivo Experiments (Kilkenny, C. et al (2009), PLoSOne, 4 (11)) •Checklists of Reporting Items •Improve Transparency and Replication •Ensure critical appraisal and interpretation NEUROIMMUNOLOGICAL ARRIVE Baker D, Amor S. Publication guidelines for refereeing and reporting on animal use in experimental autoimmune encephalomyelitis. J Neuroimmunol. 2012; 242:78. Animal Studies Sample Size Calculations: <5% Randomization <20% Blinding <30% RESEARCH PAPER-STRUCTURE
- 2. 17/07/2019 2 Reading Papers DEMOGRAPHICS: ComparingLike for Like (Sex, Age, History) SAMPLE SIZE Power to Detect Change Too Small- Miss Important Events Too Large-Extra Cost/Risk to People in Study BIAS REDUCTION: •RANDOMIZATION: Methods to Achieve this. Was it achieved? •BLINDING (Single Blind/Double Blind) METHODS OF Is the correct analysis used? ASSESSMENT OUTCOMES: Are they fit for purpose? Are the Outcomes Responsive? Control Response (Placebo Effect) COMPLIANCE Drugs don’t work if people do not take them SIDE-EFFECTS Tolerability * * * IMPORTANT ASPECTS OF HUMAN STUDIES Reading Papers 1. TITLE & ABSTRACT 2. BACKGROUND & OBJECTIVES 3. TRIAL DESIGN 4. PARTICIPANTS (Eligibility) 5. TREATMENT 6. OUTCOME MEASURES 7. SAMPLE SIZE CALCULATION 8. RANDOMIZATION METHOD 9. ALLOCATION OF RANDOMISATION 10. IMPELEMENTATION OF RANDOMISATION 11. BLINDING 12. STATISTICS 13. PARTICIPANT FLOW RESULTS 14. RESULTS-RECRUITMENT 15. RESULTS-BASELINE DEMOGRAPHICS 16. RESULTS-NUMBERS- HOWMANY PEOPLEIN EACH GROUP 17. RESULTS-OUTCOMES 18. RESULTS ADDITIONAL ANALYSIS 19. RESULTS-SIDE EFFECTS 20. DISCUSSION-LIMITATIONS 21. DISCUSSION- GENERALISABILITY (VALIDITY) 22. DISCUSSION- INTERPRETATION 23. REGISTRATION (www.clinicaltrials.gov) 24. SOURCE OFPROTOCOL 25. FUNDING Checklists of Reporting Items Improve Transparency and Replication Ensure critical appraisal and interpretation http://www.consort-statement.org/ DESIGN OF TRIAL Reading Papers DESIGN OF TRIAL Assessment for Eligibility Randomization Allocated to Intervention Received Intervention Follow Up Analysed Did Not Receive Lost to Follow Up Not Analysed Allocated to Intervention Received Intervention Follow Up Analysed Did Not Receive Lost to Follow Up Not Analysed Excluded TEST CONTROL Reading Papers IMPORTANT ASPECTS OF HUMAN STUDIES BIAS REDUCTION RANDOMIZATION Process by which people are allocated to an intervention after assessment eligibility • It eliminates selection bias • It eliminates confounding issues by adjusting co-variants (gets same demographics in test and control groups) • Facilitates Blinding of investigators, participants, and assessors BLINDING/MASKING Individuals withheld the knowledge of the treatments OPEN LABEL No Blinding SINGLE BLIND pwMS unaware of their Treatment DOUBLE BLIND Neuro & pwMS unaware of their treatment Reading Papers DATA ANALYSIS BIOLOGICALLY SIGNIFICANT A difference between groups that has a real impact on biology STATISICALY SIGNIFICANT Mathematical Measure of a difference between groups. Does this occur at a level greater than may be expected by chance. PROBABILITY VALUES Likelihood of obtaining a statistical result by chance, assuming there is no Difference in the results (null hypothesis) being investigated P=0.05 is 1 in twenty chance of event occurring P=0.01 is 1 in one hundred chance of event occurring by chance P=0.001 is 1 in one thousand chance of event occurring by change Less Likely due to chance Reading Papers DATA ANALYSIS A (50%) B (25%) (4%) D (21%) C PIE CHARTS These show the proportions of response, seldom used in trials, but may occur in Marketing material
- 3. 17/07/2019 3 Reading Papers DATAANALYSIS BAR CHARTS These show the magnitude of the response/outcome TreatmentA TreatmentB Count Bar Chart 0 Measurement (Units) Frequency(%) Histogram 10 20 30 40 50 Reading Papers DATA ANALYSIS LINE GRAPHS These show the analysis over a period of time (serial analysis) Reading Papers Error Bars & Variability No Difference DifferentHelp Statistics Needed Overlap of Error Bars DATA ANALYSIS Reading Papers KAPLAN-MEIER SURVIVAL CURVE The curve represents the proportion of the study population that are free of the event (Progression, Time to relapse etc ) as successive times. i.e Time to an event Commonly used to represent overall efficacy. The larger the difference in the curves the larger the difference DATA ANALYSIS Reading Papers DATA ANALYSIS HAZARD RATIO The Hazard Ratio is the Relative Risk of the Event (e.g. Disease progression) happening in one arm of a trial compared to another arm over the time of the trial Difference between groups that has a real impact on biology Hazard Ratio = 1 means there is no difference Hazard Ratio = 2 means there is twice/double the risk Hazard Ratio = 0.5 means there is half the risk. Reduced risk by 50% This methodology uses all available information, including people who didn’t complete the study Reading Papers DATA ANALYSIS ABSOLUTE RISK REDUCTION (ARR) The Difference between the number of events (e.g. pwMS who progressed) in the test verses the control group. e.g. Placebo progressed = 15% Test Drug = 10% progressed ARR of the Test drug = 15%-10% = 5% RELATIVE RISK REDUCTION (RRR). The RRR looks at the same difference Between groups given different treatments but expresses the change in risk As a proportion, rather than an absolute difference e.g. RRR of Progression by Test drug = 15-10/divided by progression of control 15-10/15 = 5/15 = 33% INTENTION TO TREAT (ITT) ANALYSIS ITT analysis of the results of a trial is based on the initial treatment assignment and not on the treatment eventually received. ITT analysis is intended to avoid various misleading artefacts that can arise such as loss of subjects, that may break random assignment to the groups. Medical investigators often have difficulties in accepting ITT analysis because of clinical trial issues like missing data
- 4. 17/07/2019 4 Reading Papers DATAANALYSIS CONFIDENCE INTERVAL (CI) The CI Percentage indicates the certainty that a result will fall into a range of results noted by lower and upper limits HR 0.55 (95% CI 0.40-0.77) The risk of converting to clinically definite MS is reduced by 45% with estimates ranging from 60% reduction to 23% reduction and 95% certainty that the result would lie between 23% and 60% risk Reduction The narrower the range the more precise the result. The CI are used as an indication how a study would be reflected in the general population Reading Papers DATA ANALYSIS NUMBER NEEDED TO TREAT (NNT) The NNT is a statistic that tells one the actual number of pwMS that would need to be treated with a given therapy for a one patient to get a particular endpoint benefit. Determines whether Large number or small number of pwMS benefit. The Lower the NTT the more effective the treatment. ODDS RATIO. A measure of treatment effectiveness. The OR is the odds of an event happening in the treatment group expressed as a proportion of the odds of the event happening in the control group. OR close to 1 the smaller the difference in the treatment effect If greater (or lesser) than 1, then the treatment is greater (or less) than the control treatment. Reading Papers FOREST PLOT Following meta-analysis of a number of different trials some showing benefit some not showing for treatment or for analysis of subgroups within a trial that may respond differently. A Forest plot is a way to compare different groups of data. Size of square is proportional to the importance of that data set and the horizontal lines are the confidence intervals DATA ANALYSIS Reading Papers DATA ANALYSIS Regression to the mean is the phenomenon that if a variable is extreme on its first measurement, it will tend to be closer to the average on its second measurement—and if it is extreme on its second measurement, it will tend to have been closer to the average on its first. In trials people are more likely to volunteer for trials if they are not doing as well and natural history of disease will mean the flair resolves and they get better Reading Papers SCATTER GRAPH (Relationships) Individual data points are plotted showing two variables (e.g. height verses weight) to show relationships Random Positive relationship Negative relationship UnrelatedUnrelated Subset DATA ANALYSIS Reading Papers DATA ANALYSIS REGRESSION (Relationships) Spearman's rank correlation coefficient is a non-parametric measure of stastical dependence between two variables. It assesses how well the relationship between two variables can be described. R= +1 perfect Correlation or R = −1 perfect inverse correlation. R-squared is a statistical measure of how close the data are to the fitted linear regression line. It is the percentage of the response variable variation that is explained by a linear model. Or: R- squared = Explained variation / Total variation (It is between 0 and 100% or 0 to 1:) 90% explained40% explained
- 5. 17/07/2019 5 Reading Papers DATAANALYSIS BOX & WHISKER PLOT Rather than show all the distribution of individual data. The data is presented as a box with the whisker indicating deviation. Reading Papers Grade 1 Asymptomatic(no) or mild symptoms; for this grade of AE, treatment is not indicated. Grade 2 Moderate; for this grade of AE minimal, local or non-invasive intervention may be indicated, for example, this might involve treating the symptom (e.g. nausea and vomiting caused by a treatment managed/prevented by giving the patient drugs to stop sickness,rather than stopping the drug). Grade 3 Severe but not immediately life-threatening; for such AEs hospitalization and investigation/ management are often indicated. Grade 4 Usually life-threatening consequences; for these AEs urgent intervention is indicated. Grade 5 Death related to AE. IMPORTANT ASPECTS OF HUMAN STUDIES Reading Papers • Improve Reporting of Research Using Animals. • Guide Authors as to the Essential Information to Include in a Manuscript, • Flexible Enough to Accommodate Reporting a Wide-Range of Research Areas and Experimental Protocols. • Promote Reproducible, Transparent, Accurate, Comprehensive, Concise, Logically-Ordered & Well-Written Manuscripts. • Improve the Communication of the Research Findings to the Broader Scientific Community. Animals in Research: Reporting In Vivo Experiments Call for Transparency Reading Papers Call for Transparency 1 1 O C T O B E R 2 0 1 2 | VO L 4 9 0 | N AT U R E | 1 8 7 Reading Papers Call for Transparency bioRxiv preprint first posted online Jul. 15, 2019; doi: http://dx.doi.org/10.1101/703181. The Ten Essentials Plus the Recommended Set Reading Papers Essential Ten Percie de Sert et al. 2019
- 6. 17/07/2019 6 Reading Papers TITLE Provide asaccurate and concise a description of the content of the article as possible. ABSTRACT Provide an accurate summary of the background, research objectives, including details of the species or strain of animal used, key methods, principal findings and conclusions of the study. IF YOU DON’T DO THIS….. NO ONE WITH READ YOUR PAPER! ARRIVE Guidelines Reading Papers ARRIVE Guidelines INTRODUCTION 3. Background (a). Include sufficient scientific background to understand the motivation and context for the study, and explain the experimental approach and rationale. (Include relevant references to previous work) (b). Explain how and why the animal species and model being used can address the scientific objectives and, where appropriate, the study’s relevance to human biology. 4. Objectives Clearly describe the primary and any secondary objectives of the study, or specific hypotheses being tested. “A clear statement of the objective of the study, or the main hypothesis being tested, was described in the introduction by 95% of the 271 publications (Kilkenny et al. 2009)” Reading Papers ARRIVE Guidelines METHODS 5. Ethical statement Indicate the nature of the ethical review permissions, relevant licenses and national or institutional guidelines for the care and use of animals, UK Protecting Research Animals through Law since 1876. Animals (Scientific Procedures) Act 1986. European Union. Directive 2010/63/EU. (Enshrined the Concept of 3Rs and Ethical Review Process) . Animals Welfare Act 1966 USA 2000 Law for the Humane Treatment and Management of Animals in Japan Reading Papers ARRIVE Guidelines METHODS 5. Ethical statement Indicate the nature of the ethical review permissions, relevant licenses and national or institutional guidelines for the care and use of animals, UK Protecting Research Animals through Law since 1876. Animals (Scientific Procedures) Act 1986. European Union. Directive 2010/63/EU. (Enshrined the Concept of 3Rs and Ethical Review Process) . Animals Welfare Act 1966 USA 2000 Law for the Humane Treatment and Management of Animals in Japan Reading Papers ARRIVE Guidelines METHODS…continued 6. Study design For each experiment, give brief details of the study design including: a. The number of experimental and control groups. b. Any steps taken to minimise the effects of subjective bias when allocating animals to treatment (e.g. randomisation procedure) and when assessing results (e.g. if done, describe who was blinded and when). c. The experimental unit (e.g. a single animal, group, or cage of animals). EVIDENCE OF BLINDING ~ 14% Randomly Selected papers (Kilkenny et al.2009). ~ 29% Experimental Stroke papers ~ 16% Experimental MS papers (Vesterinen et al. 2010) EVIDENCE OF RANDOMISATION ~ 12% Randomly Selected papers (Kilkenny et al.2009). ~ 36% Experimental Stroke papers ~ 9% Experimental MS papers (Vesterinen et al. 2010) Blinding Experiments Reading Papers ARRIVE Guidelines Blinded Assessment of Outcome 0 10 20 30 40 50 No Yes %ImprovementinNeurologicalScore P<0.002 BLINDING Random Allocation to Group %ImprovementinNeurologicalScore 0 10 20 30 40 50 No Yes P<0.002 RANDOMIZATION METHODS…continued 6. Study design Vesterinen et al. Mult scler; 2010, 16:1044 2010
- 7. 17/07/2019 7 Reading Papers ARRIVEGuidelines METHODS…continued 7. Experimental procedures For each experiment and each experimental group, including controls, provide precise details of all procedures performed. For example: a. How (e.g. drug formulation and dose, site and route of administration, anaesthesia and analgesia used [including monitoring], surgical procedure, method of euthanasia). Provide details of any specialist equipment used, including supplier(s). b. When (e.g. time of day). c. Where (e.g. home cage, laboratory, water maze). d. Why (e.g. rationale for choice of specific anaesthetic, route of administration, drug dose used). Reading Papers ARRIVE Guidelines __________________________________________________ Source Mice Source Innoculum No.EAE/Total __________________________________________________ Company X Company X 1/20 Company X Baker Lab 9/10 Baker Lab Company X 4/10 Baker Lab Baker Lab 10/10 ________________________________________________ 6-8 week oude, vrouwelijke ABH muizen werden subcutaan in 2 plaatsen in de rug van de flank met 0.1ml een sonicated (10min in Bransonic Ultrasonicator, Sigma Ltd, Poole, GB) emulsie van 1mg syngeneic ruggenmerg homogenate fosfaat beschermd zouthoudend en de volledige hulp van Freund ingespoten die 60µg kobalt-gebestralen Mycobacterium tuberculose op dag 0 en 7. 6-8 week old, female ABH mice were injected subcutaneously in 2 sites in the dorsum of the flank with 0.1ml a sonicated (10min in Bransonic Ultrasonicator, Sigma Ltd, Poole, UK) emulsion of 1mg syngeneic spinal cord homogenate phosphate buffered saline and Freund’s complete adjuvant containing 60µg cobalt-irradiated Mycobacterium tuberculosis on day 0 and 7. METHODS…continued 7. Experimental procedures….continued Al-Izki S et al. Multiple Sclerosis and Related Disorders 2012;1: 29–38 Reading Papers ARRIVE Guidelines METHODS…continued 8. Experimental animals a. Provide details of the animals used, including species, strain, sex, b. Provide further relevant information such as the source of animals, International strain nomenclature, Genetic modification status (e.g. knock-out or transgenic) & Genotype, Health/immune status, Drug or Test naïve, Previous procedures, etc. PROVIDE DETAILS OF DEMOGRAPHICS! _____________________________________ Source No. EAE/Total _____________________________________ In-house Bred ABH mice 10/10 Commercial Bred ABH mice 5/10 _____________________________________ Reading Papers ARRIVE Guidelines METHODS…continued 9. Housing and husbandry (This part of the Ethical review process) & Environment Enrichment a. Housing (e.g. type of facility,e.g. specific pathogen free [SPF]; type of cage or housing; bedding material; number of cage companions. b. Husbandry Conditions (e.g. breeding programme, light/dark cycle, temperature, quality of water, access to food and water, environmental enrichment). c. Welfare-Related Assessments and Interventions that were carried out prior to, during, or after the experiment. Reading Papers METHODS…continued 10. Sample size a. Specify the total number of animals used in each experiment and the number of animals in each experimental group. b. Explain how the number of animals was arrived at. Provide details of any sample size calculation used. c. Indicate the number of independent replications of each experiment, if relevant. POWER ANALYSIS! https://www.nc3rs.org.uk/experimental-design-assistant-eda Experimental Design Assistant Vesterinen et al. Mult scler; 2010, 16:1044 2010 Reading Papers METHODS…continued 11. Allocating Animals to Experimental Groups a. Give full details of how animals were allocated to experimental groups, including randomisation or matching if done. b. Describe the order in which the animals in the different experimental groups were treated and assessed. 12. Experimental outcomes Clearly define the primary and secondary experimental outcomes assessed In Human Trials failure to change primary outcome measure is a FAILURE!
- 8. 17/07/2019 8 Reading Papers Time Post-Induction (Days) 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 MeanDiseaseScoreSEM 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 Drug Not-Interesting? Vehicle Limp tail Hind Limb Paralysis Partial Paralysis Impaired Righting Reflex Drug Treatment METHODS…continued 11. Allocating Animals to Experimental Groups Reading Papers Time Post- Induction (Days) 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 MeanDiseaseScoreSEM 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 Drug Not-Interesting? Vehicle Limp tail Hind Limb Paralysis Partial Paralysis Impaired Righting Reflex Drug Treatment METHODS…continued 11. Allocating Animals to Experimental Groups Reading Papers METHODS…continued 13. Statistical methods a. Provide details of the statistical methods used for each analysis. b. Specify the unit of analysis for each dataset (e.g. single animal, group of animals, single neuron). c. Describe any methods used to assess whether the data met the assumption of the statistical approach. Reading Papers DATA HANDLING Mean = Average Standard Deviation = Approximation of Variability 1.96SD contains 95% of sample Standard Error (Square Root of SD) place were mean will occur in 95% of studies Median = Middle Number Range = Top – Bottom result Quartile= 0, 25, 50, 75,100% Percentile = 1-100th division Confidence Interval 95% CI = 95% that mean will be with in limits ANALYSIS Proportions: Chi-Square Two Groups: t test, Mann Whitney U, Wilcoxon Rank Test Many Groups: ANOVA, kursall and Wallace Is data Normally Distributed are the Variances Equal If not use non-parametic stats Reading Papers Baker D, et al. PLoS Biol. 2014; 12:e1001756. Analysis of Non-Parametric Clinical Score Data with Parametric Statistics 55% (CI 46.7%-62.3%)39% (CI 32.5%-46.8%) 13% (CI 8.7%-18.58%) CNS–infiltration to Clinical Score is Not Linear Gupta S et al. PLoS One. 2013 Oct 4;8(10):e76330. Reading Papers Figure legend. Clinical scores of two independent EAE experiments at d23 post disease induction. Individual scores as well as the mean score of two independent experiments are shown. Control: n=10, vehicle: n=13, xxxxx-345: n=11. Control versus vehicle: P=0.620, control versus xxxxx-345: P=0.017, vehicle versus xxxxx-345 P=0.029. * indicate P values <0.05 and ** indicate P values <0.005 based on a non-paired Student’s t test. Error bars are s.e.m The assumptions of a t test (a) Data is normally distributed. You test this and it passes the test p=0.152, (b) data groups have equal variances Test for that and it fails P<0.05. (c) Data is Parametric. Fail. It is non-parametric. t test is not a valid test Mann Whitney test.P=0.082.........Ooooops.
- 9. 17/07/2019 9 Reading Papers RESULTS 14. Baselinedata. For each experimental group, report relevant characteristics and health status of animals (e.g. weight, microbiological status, and drug- or test-naïve) prior to treatment or testing (this information can often be tabulated). 15. Numbers Analysed Report the number of animals in each group included in each analysis. Report absolute numbers (e.g. 10/20, not 50%). b. If any animals or data were not included in the analysis, explain why. 16. Outcomes and Estimation Report the results for each analysis carried out, with a measure of precision (e.g. standard error or confidence interval). Reading Papers Time Post- Induction (Days) 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 MeanDiseaseScoreSEM 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 Drug Not-Interesting? Vehicle RESULTS Measure of Precision Limp tail Hind Limb Paralysis Partial Paralysis Impaired Righting Reflex Reading Papers Time Post- Induction (Days) 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 MeanDiseaseScoreSEM 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 Drug Not-Interesting? Vehicle RESULTS Measure of Precision Limp tail Hind Limb Paralysis Partial Paralysis Impaired Righting Reflex How many Animals Got Disease? Severity Changed? Skewing due to Non-susceptibility? Death/Endpoint? Reading Papers RESULTS…continued 16 (old 17) Animal Care & Monitoring Describe methods to reduce suffering. Increase the Dose Until it Does Something! _________________________________ Treatment No.EAE/Total _________________________________ Saline Amount 1 * 12/12 Saline Amount 2 11/12 Saline Amount 3 4/8 _________________________________ * = RDA ARE THE DOSES IN THE TEST IN VITRO OR ANIMALS RELEVANT TO THE HUMAN CONDITION? Reading Papers DISCUSSION 18. Interpretation/scientific implications a. Interpret the results, taking into account the study objectives and hypotheses, current theory and other relevant studies in the literature. b. Comment on the study limitations including any potential sources of bias, any limitations of the animal model, and the imprecision associated with the result. c. Describe any implications of your experimental methods or findings for the replacement, refinement or reduction (the 3Rs) of the use of animals in research. Reading Papers DISCUSSION….Continued 18. (Previous 19) Generalisability/ translation Comment on whether, and how, the findings of this study are likely to translate to other species or systems, including any relevance to human biology. A NEW POTENTIAL CURE COMES EVER WEEK?…….. WHILST JOURNALISTS MAY OVER-INTERPRET YOUR FINDINGS, REMEMBER, PEOPLE WITH SERIOUS DISEASES ARE DESPERATE FOR THAT CURE…… BE REALISTIC & DON’T CREATE TOO MANY FALSE HOPES! Public Engagement
- 10. 17/07/2019 10 Reading Papers 19. PROTOCOLREGISTRATION Was a protocol done before the work was done and where is it registered? Human Trials are Registered at Clinical Trials.gov (USA), EudraCT (EU) Perhaps may help limitation of repetition of animals work Public Disclosure stops the capacity to file a patent Reading Papers NEW 20-DATA ACCESS STATEMENT Consider deposition of Raw Data as a Supplementary file • Help with review • Allow further analysis and allow others to understand the data • Provide data for sample size calculations Levy Barazany et al Exp Neurol 2014: 255:63-70 Asychronous relapsing disease Baker et al. Ann clin trans neurol 2019. doi: 10.1002/acn3.792 Myelin oligodendrocyte glycoprotein induced disease in Non Obese Diabetic Mice Asychronous relapsing disease Reading Papers NEW 21-CONFLICT OF INTEREST STATEMENT Real and Perceived Conflicts of Interest Funding (Previous 20) Todays Research Can Be Tomorrows Drug June 2010 KNOWLEDGE IS THE KEY TO GAINING KNOWLEDGE Allows You to Understand the Paper Allows You to Interpret the Results Allows you to Place the Work in Context of its Relevance Allows you to see the flaws that may affect interpretation Allows you to Judge the Value of the Work Allows you to Exploit the Information from the work Allows you to learn from yours and others mistakes GIVES YOU THE CONFIDENCE IN ALL OF THE ABOVE Reading Papers Thank you for Listening