A 3′UTR modification of the TNF-α mouse gene increases peripheral TNF-α and modulates the Alzheimer-like phenotype in 5XFAD mice
Authors: Sotiria Boulekou
Nikoleta Kalovyrna
Evi Paouri
Spiros Georgopoulos
Olympia Apokotou
Athena Boutou
Journal: Scientific Reports (2020)
Discipline: Multidisciplinary
Keywords: Tumor necrosis factor-α; Alzheimer's disease; Anti-TNF-α; 5XFAD mice; Brain inflammation
Affiliations: Laboratory of Cellular Neurobiology, Center of Basic Research, Biomedical Research Foundation, Academy of Athens, 11527, Athens, Greece., Laboratory of Cellular Neurobiology, Center of Basic Research, Biomedical Research Foundation, Academy of Athens, 11527, Athens, Greece. .

Abstract:  Tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine, involved in Alzheimer's disease pathogenesis. Anti-TNF-α therapeutic approaches currently used in autoimmune diseases have been proposed as a therapeutic strategy in AD. We have previously examined the role of TNF-α and anti-TNF-α drugs in AD, using 5XFAD mice, and we have found a significant role for peripheral TNF-α in brain inflammation. Here we investigated the role of mouse TNF-α on the AD-like phenotype of 5XFAD mice using a knock-in mouse with deletion of the 3'UTR of the endogenous TNF-α (TNFΔARE/+) that develops rheumatoid arthritis and Crohn's disease. 5XFAD/TNFΔARE/+ mice showed significantly decreased amyloid deposition. Interestingly, microglia but not astrocytes were activated in 5XFAD/ TNFΔARE/+ brains. This microglial activation was associated with increased infiltrating peripheral leukocytes and perivascular macrophages and synaptic degeneration. APP levels and APP processing enzymes involved in Aβ production remained unchanged, suggesting that the reduced amyloid burden can be attributed to the increased microglial and perivascular macrophage activation caused by TNF-α. Peripheral TNF-α levels were increased while brain TNF-α remained the same. These data provide further evidence for peripheral TNF-α as a mediator of inflammation between the periphery and the brain. 

Retracted: No

PubPeer comments:
 Despite the huge number of participants, this investigation must still be categorised as observational study, i.e. not a clinical trial. We should be very careful in interpreting efficacy from such a study, as the conduct was not subject to Good Clinical Practice or inspections from regulatory authorities. Normally, hypothesis generation and safety and tolerability are the main objectives of observational studies. But here, the authors focussed exclusively on efficacy. The authors reported u201cdeathsu201d, but in fact all data on deaths were u201crelated to COVID-19u201d; this means that death was counted only if the person had a positive SARS-CoV-2 test. Table S1 does not even clarify whether the test had to be done before (then: which time frame) or after the death (post mortem swabs allowed?). The protocol appended to the article does not specify the test PCR products; this is in contrast to the protocol of the pivotal clinical trial with the Pfizer-Biontech vaccine. Was the PCR really not standardised? Which CT values they accepted? Which genes they tested? The authors did not tell anything about death due to any cause or serious adverse events. , 
 #1 You ll note under the study design, it s listed as an observational study. It s not an interventional study and is not subject to Good Clinical Practice or inspection from  regulatory agencies . The study was approved by the Clalit Health Services IRB. Existing data was collected from the data repository on patients who had already been vaccinated as well as those who had not been vaccinated. This type of data is generally not subject to informed consent. You are incorrect when you state  hypothesis generation is the main objective of observational studies. It depends on the study design. Some designs, such as cross-sectional studies and ecological studies are  hypothesis-generating designs, while others are hypothesis-testing (cohorts and case-controls). This aim of this study was to evaluate the effectiveness of the vaccine work in real life situations. Does the vaccine appear to work equally among all age groups and number of co-morbid conditions? Does the vaccine prevent COVID-19 illness (per PCR, symptoms) Does the vaccine prevent hospitalization for COVID-19? Does the vaccine prevent death from COVID-19? Does the vaccine appear to be effective against circulating variants? Efficacy answers the question,  How well does the intervention work under trial conditions while effectiveness answers the question,  How well does the intervention work in real-life circumstances . The comment about the PCR testing is irrelevant as the same testing is applied to both the vaccinated and unvaccinated groups. The vaccinated group wasn t tested differently from the unvaccinated group in a systematic manner (this is what introduces bias-differential differences). Characterization of the number of adverse events and deaths is captured in the phase 3 trials. The sample sizes in those trials were more than adequate to capture adverse events. You ll note there were 596,618 vaccinated individuals matched to the same number of unvaccinated (you can see the covariate balance in figure S2). This population includes ~44K individuals over the age of 80. Based on probability alone, individuals will die from coincidental events (background rate). The number of events is less important than the differences between the two groups. The number of deaths in the vaccinated group was 9 compared to 32 deaths in the unvaccinated group. , 
 Thank you very much, dear Lithophane Adipel! I did not challenge anything about informed consent, but missing GCP also means no source data verification, no monitoring, no regulatory inspection of the data capturing, no independent validity checks. I also did not blame you for u201csellingu201d this investigation as randomised or interventional clinical trial. Unfortunately, many readers might misunderstand this investigation, because of the gigantic numbers and the neatly fitted control set. Certainly, misunderstandings by readers might not be your fault. u201cReal worldu201d is OK, but this term did not occur in the text. In fact the article stresses u201climitationsu201d only with respect to the pivotal clinical trial(s), but not with respect to the present investigation. Your response concerning the unspecified PCR test, that it was the same or similar in both cohorts, is acceptable. But it remains a pity concerning external validity. How far can we compare your data with the pivotal clinical trial or other investigations? You responded to u201cadverse eventsu201d, which I did not address. It is fully acceptable that you did not capture nonserious adverse events or adverse reactions in your study. In fact I addressed u201cserious adverse eventsu201d and u201cdeath due to any causeu201d. I have to confess that even capturing of serious adverse events is challenging for a study such as yours. Based on data from the pivotal trial with that vaccine you had to expect more than 5000 serious adverse events for your investigation; this is a lot and might have been too burdensome, at least for a rapid publication of the data. Nevertheless, it was a unique chance to get such large scale data. Your response provided nothing on the very important deaths due to any cause. With such a large database you have had the obligation to inform the world about the net benefit of the vaccination in terms of all cause mortality. This is what matters for potential vaccine recipients. Having mentioned this, I should also stress the findings of Dr Hervu00e9 Seligmann pertinent to your investigation: http: www.nakim.org israel-forums viewtopic.php?t=270873 u201cthere are on the first day of vaccination 172 positive COVID-19 cases among the 596618 vaccinated, vs. 359 cases, more than twice, among the controls that same day.u201d It is almost impossible that this occurred by chance. And this underlines the importance of source data verification and regulatory oversight, as addressed above. [Moderator: unsupported assertion deleted - see our FAQ] , 
 From a comment on social media: https: twitter.com doctor_i_am_the status 1383212936592691208?s=21 u201cThis study in a nationwide mass vaccination setting suggests emphasize the word  SUGGESTS. There is no measured T-cell or B-cell response. The flawed use of PCT and symptoms that are not specific to SARS-CoV-2 are not definitive. There is no real discussion statistically comparison (sic) of this nominal data. Taking control patients and then including in the experimental group prevents actual comparisons of at least this much of the study.u201d , 
 I m somewhat of a novice here, just a student and not a researcher, so forgive me if I have misinterpreted, but isn t the ARR from a study only going to be over the time course of the study? So in Dagan, Barda, Kepten, et al (2021), the they measured over 42 days. so wouldn t that mean that the ARR of 0u00b746% would be over the course of a 42 day span when the prevalence of the virus was whatever it was at that time? I ve been vaccinated, but I expect that protection to last beyond 42 days. At some point we want the government to relax all COVID related restrictions. We might expect that in a situation where no one is vaccinated, and no public health restrictions are in place, then the chances of catching SARS-COV-2 will over time, approach something close to 1. So isn t that how we should be evaluating the vaccines? Not over a time course of 42 days with public health restrictions in place, and a prevalence that reflects that, but over the full lifetime we expect this virus to be with us, and with the freedoms we expected pre-pandemic? ,