[Stroke/Vaccines] Varicella-Associated Stroke (Vora et al., J Pediatrics 2018)

727899-fig1

You may have heard about the case report of a 11-month baby suffering from an ischemic stroke injury. After assessing patient history, the patient has experienced (along with his siblings) a chickenpox infection (Varicella-Zoster virus) 2-3 months before the stroke event, such infection was classified as mild. The suspicion of VZV was confirmed by the presence of anti-VZV IgG and IgM in patient serum, but also by the presence of detectable viral particles (DNA) in the CSF of the patient 2 days after admission (~900 copies/mL, over the  500 copies/mL minimal quantitative limit: http://www.questdiagnostics.com/testcenter/BUOrderInfo.action?tc=18270&labCode=AMD).
The patient was put on antiviral regimen and showed signs of improvement. However, the patient shown signs of arteriopathy,  suggesting this patient is at risk for subsequent stroke events.
For me, it is an interesting learning experience as I never have heard about the occurrence of stroke events following chickenpox. It is certainly breaking the image of a “harmless childhood” disease that maybe associated prior the introduction of the chickenpox vaccine.
Indeed it appears that such complication is not unheard of and you can find several case reports in the literature about pediatric ischemic stroke associated with a previous viral infection. Indeed, contracting the varicella-zoster virus increases the risk of developing an ischemic stroke by 4-fold from 0-6 months post-infection (https://www.ncbi.nlm.nih.gov/pubmed/24092802 and https://www.ncbi.nlm.nih.gov/pubmed/27138380). In the other hand, vaccination against VZV showed no signs of increased risk of stroke  in vaccinated population versus not-vaccinated (https://www.ncbi.nlm.nih.gov/pubmed/29655630).
Considering the limited if not-existent therapeutic arsenal for the treatment of ischemic stroke injury, prevention appears as the better approach at this point. In summary, don’t underestimate the chickenpox and vaccinate your kids.

 

 

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[Movies/SciFi] 2001 – A Space Odyssey (50th anniversary)

EDIT: This blog post was in my drawer since April 3rd. For some reasons, it got never published. Time to give it justice and publish it.
Today we celebrated the 50th anniversary of 2001: A Space Odyssey. As until today, this movie is still considered one of the best science-fiction movie ever made until now, setting the cornerstone for a whole generation of filmmakers. Unconsciously, almost everyone ended up associating the mythical piece of Richard Strauss “Also Sprach Zarathustra” with the moon landing and the first men to walk of mankind on the moon.
I still remember watching it the first time on the TV, as part of an “homework” given by my 4th grade teacher (including a note to the parents to allow me to stay late).
It is fairly impressive the details and the realism in the decor used in this movie that still holds until now. It is also impressive that as today we are still speculating about the meaning and the philosophy behind the movies. Considering Kubrick, it was expected.
Lets just quickly discuss the plot. We start with what appears as the dawn of humanity, with some hominids that are ape-like. I assume that they are our common ancestor somewhere in the African Rif Valley. Nothing seems to disturb much these hominids, until the appearance of a black monolith. First, it creates fear amongst these hominids. Until one breaks such fear and leave it to curiosity, approaching and touching the black monolith. This creates a spark into this hominid, as he wakes up and grab a bone, learning to use it as a tool (for hunting, as we see an animal clubbed to death) and a weapon (as it club to death the alpha male of a rival creed), with a photography blended with the mythical “Also Sprach Zarathustra”.

I interpret this is a metaphor of humankind, as we as mankind were able to transition from belief (and fear-based)-based society to a secular (and science-driven) one.  As a cognitive animal, we learnt to develop skills beyond any other animals yet still driven by our reptilian brain (technology as a good or a bad usage).
This transition us into space, hearing the “Blue Danube Waltz” by Strauss.

The genius of Kubrick here was to brilliantly integrate classical music as a movie soundtrack, especially in sci-fi movies. This is I think inspired Georges Lucas to seek collaboration with John Williams for “Star Wars”.
It is funny to see the Pan-Am logo on the shuttlecraft heading to the space station. The photography design of the space station is simply awesome by its brightness (the white iPod feel) and the simplicity (I also assume it served as inspiration for Steve Jobs). The plot brings us to the discovery of an artifact from unknown origin on the Moon, rapidly followed by the loss of communication with Clavius Base, the outpost at the center of the discovery. Again, the artifact turns out to be the same black monolith structure (or a similar one). Its discovery leads us to hear a very-high and unpleasant pitch sound.
We are thrown 18 months later in a spacecraft “Discovery One” heading to Jupiter. Such spacecraft holds a supercomputer with Artificial Intelligence named HAL 9000 (some attributes its acronym to IBM), a powerful computer so powerful that it is considering itself as “fool-proof and incapable of making mistakes”. I don’t want to spoil much more but the story becomes more interesting and convoluted as we go through the plot.
Kubrick showed some realistic and valid points in his movie, as we are not talking about “Warp Drive” as in Trek. No aliens to be seen, but only humans. Space is only a dark, cold vacuum environment. There are technologies that appears completely Sci-Fi back then (something like an iPad, a LCD screen, the use of visioconference, AI computers….) that discarded the fiction out of the science.
This movie really influenced a lot of the sci-fi and pop-culture but what comes in my mind is Lenny Kravitz “Believe” that is very very influenced by the movie for the movie clip.
https://youtu.be/spPWnh3JvZ0

[Sciences/BBB/Drug Delivery] Red blood cell-hitchhiking boosts delivery of nanocarriers to chosen organs by orders of magnitude (Brenner et al., Nat Comm 2018)

I know, I know I have been fairly quiet. I have to tell that between attending a scientific meeting, teaching a summer course, taking care of two grants proposals and finally handling three manuscripts. However, sometimes I also like to share some papers published in the field that are interesting or bringing novel ideas or concepts. This one has been suggested by one of the author and this is his quick summary:
We bind nanoparticles to the surface of red blood cells. These nanoparticles are going to be released from the red cells once they pass the first capillary bed. Therefore we can concentrate the nanoparticles in the target tissue. In addition, these nanoparticles potentially can carry one or multiple selected drug(s). So if we inject intravenously the particles are going to be release in the lungs but if we inject through the brain arteries the nanoparticles will be release into the brain vasculature.
It is a very nice of work here published in Nature Communication (I consider this is one of the top OA journal to get published in) and you can download the full-text here.
You can also appreciate this paper likely went into at least one round of review, with a time period of about 7 months between it got accepted for review and accepted for publications.

In this paper, the authors have been working on trying to develop alternative drug delivery carrier, in that case use red blood cells as “piggyback” cells to enhance drug delivery. They tried different formulations on isolated RBCs and identified some suitable for carrying antibodies or proteins.  They called these piggybacking RBCs as RBC-Hitchiking (RH) (I wonder if it is some Easter egg towards the “Hitchiking Guide to Galaxy”).

Upon identification of the right nanoparticle materials, the authors investigated the distribution and the delivery of the conjugates in different organs, as liver and lungs. What is interesting is the amount of injected dose recovered is much higher in the RH than the free-circulating one, in particular in lungs, whether they are bare-naked or bound with a protein. These nano-carriers can be delivered to endothelial cells.

But the interesting snack-bite from this paper is the intra-arterial injection in the carotid artery, in which there was a significant increase in nano carriers delivery in the brain.  Nano-carriers alone show a %ID lesser than 1% (that is about what you expect from delivering antibodies from an IV route towards the brain)  to over 10%.   The delivery was also maintaining the ipsilateral injection site, which is good considering you are likely to treat one brain hemisphere.

Now, time for me to be picky and kind of wish we had information on the PK profile, especially if this approach increased the stability of the nanocarriers. It would be also interesting to see how this technique fare in a experimental disease model (for instance a xenograft brain tumor and see if you can deliver targeted chemotherapy in mice).

Nevertheless it is a good paper that take us out from the classical nanoparticles formulation and try here an innovative and novel approach in drug delivery.

 

[Metal/Symphonic] Adrana – Pertubatio (10th Anniversary)

Few days ago marked the release of Adrana’s first album “Perturbatio” (according to Metal-Archives.com, it was released on June 8th 2008). What can I tell about from one of my favorite French metal band? That it was an interesting first album, that it can sounds sometimes goofy as any first albums but also also has some valuable gold nuggets, just by listening to the soprano voice of Anne is enough to bring it up into the ranking. And also because it has a lots of tracks sung in French, so thats another perk to add.
It is a 14 track album, totaling about 50 mins of songs. We get into it with an intro sequence “Enter Prusias” that sounds like a medieval swordfight (for me, it sounded like more a cutelry fight) which you can guess is a ramp to “Prusias”. With “Prusias”, you get insta-slapped by the voice of Anne, singing with her high vocal ranges really making the tone of Adrana over a “Symphonic metal” band and more into its own “Opera metal” subgenre.  Blending soprano opera feminine voices with metal riffs is my guilty pleasure and Adrana in my guilry indulgence. The second is the pace and musical arrangment, raw and not much relying on the synth keybaords. “Saneday” brings on with a piano solo and progressively introduce metal elements into the composition. “The Moonchildren” is one of the track that I found it a bit goofy, as Anne French accent betrayed her in the introducting narrative and it is quickly followed by “Mortelle Fourberie Infantine”, brining the fast-paced rhythm with Anne’s voice. One of my favoirte. It is followed by “The Nymph’s Corpse”, another of my favorite. But the “piece of resistance” is coming with the following track named “Gabrielus”. Oh boy, this is a battlecry song, that feel like jumping a ride on a white armored horse and raise your sword in the sky calling for the ultimate assault on the legion of orcs, all sang in French and with this might power metal tunes (the 14th track, is an acoustic version of “Gabrielus”). If I have to have someone introduced someone to Adrana, I would definitely use this track.
“Secret Gathering” is an another track that has some goofy elements, especially at the beginning. “l’Eveil de Marklus”, “The Lords of The Tapestry”, “Oprane” and “Ingalrian” are nice tracks but nothing transcendental for me. You can skip them without missing much in my opinion. But the one you should not skip is “Mortualia” that is almost an acapella/acoustic version of Anne with a baritone. Very beautiful if you are into opera voice.
In conclusion, “Pertubatio” is an album like any first album with some goofiness but also some potentials, both from Anne and from musical elements. Indeed, the band further blacksmithed their core element and perfected it with their following up album “The Ancient Realms”.

 

[Sciences/Junk Sciences/Vaccines] HPV vaccine safety – a tale of two studies

“It was the best of times, it was the worst of times” Charles Dickens – A Tale Of Two Cities

If you are following a bit the current news about immunization and vaccines, you likely heard about two studies in regards of the effect of HPV vaccines on population safety, in particular in terms of risk of developing complications or chronic conditions.
Interestingly two studies (to be honest one study and one comment) weighing the pros and cons of HPV public immunization were published within weeks.
One of them was claiming that HPV vaccines increased the risk of cervical cancer in the Swedish population, the other the incidence of autoimmune diseases in the Canadian population, in particular in the population of Ontario province.
One of them was published in a society journal with an reasonably impact factor (IF~8 and Scimago score of 1.7), the other in a journal with inexistent impact factor (Scimago score of 0.2) showing behavior similar to “predatory journals”.
One of them was published “in a peer-reviewed general medical journal that publishes original clinical research, commentaries, analyses, and reviews of clinical topics, health news, clinical practice updates and thought-provoking editorials.”; the other “is a multi-disciplinary academic journal providing a platform for publication of original material and discussion on all aspects of healthcare ethics and the humanities, relevant to and/or from the perspective of India and other developing countries.
One of them was a full-length study with several authors, the other one a rapid communication labelled as “comment” and purposely falsified the author’s name and affiliation (hiding behind an outlook email address), claiming the fear of retaliation by the opposing group.
One of them was a controlled population study, investigating two groups (vaccinated versus unvaccinated) with a sample size of 100’000+ each; the other one tossed epidemiological data together without further stratification and cherry-picked the information.
One of them was concluding the safety of HPV vaccine and absence of increased risk of autoimmunity, the other questioned the safety of HPV vaccine straight from the abstract as “I discuss the possibility that HPV vaccination could play a role in the increase in the incidence of cervical cancer by causing instead of preventing cervical cancer disease in women previously exposed to HPV. A time relationship exists between the start of vaccination and the increase in the incidence of cervical cancer. The HPV vaccines were approved in 2006 and 2007, respectively and most young girls started to be vaccinated during 2012–2013.
After a media firestorm and the strange support of the editorial board towards the author (and still consider the data as legitimate), it got finally retracted. The corresponding author has also been informed he will have four other retractions upcoming.
Guess which one was the flawed study published with a falsified author information and in a journal with a scope outside the content of the article published?
You can these publications here and here.

[Vaccines/Junk Sciences] “Murine hypothalamic destruction with vascular cell apoptosis subsequent to combined administration of human papilloma virus vaccine and pertussis toxin” (Aratani et al., Sci Rep 2016 – RETRACTED) Lesson from a paper that went completely fritz on the scientific method

You maybe heard about the recent retraction notice of a study published in Scientific Reports that raised concern about the safety of Gardasil(R) (HPV vaccine) and got retracted this week. Another anti-vaccine paper that bit the dust. I would have say, I am not surprised at all. Anti-vaccine studies have this very annoying habit of either being a proven fraud (remember the latest Shaw paper?), botching the experimental protocol with omission of proper controls (thats the Exley paper I have reviewed) or conveniently sweeping under the rug some data that are not fitting the narrative (that goes for one recent paper published by Gherardi).
But this one is interesting at several levels, because there is a bit of blood-brain barrier in  it, and also adds to the list of papers retraction in Scientific Reports and recent threats by scientists in the editorial board to resign from their positions due to ambiguous and unjustified decision on a flawed paper (Disclaimer: I have a study authored in this journal and I have peer-reviewed for them a couple of times).

To be honest, I only heard about this paper during the last weekend and took some times to read the paper. I will be honest, I don’t see any scientific fraud in the sense of data manipulation. What concerns me is how such a botched study could even pass through peer-review process? Considering I self-impose a quality of standard in my manuscripts and still get challenged by peer-reviewers, seeing such junk studies getting a free pass is a bit vexing. I agree that open-access may not have the same stringency in terms of peer-review filter but considering Scientific Reports as part of Nature Publishing Group, you expect a rigor found for any Nature-related journals applied in this journal too.

But lets go through the paper, it is retracted but you can still access it here.

What is the wrong with this paper?

1. The experimental design in terms of groups

The first problem arises from Figure 1.

Aratani_Fig1

We have six groups: vehicle (PBS), pertussis toxin (PTX), Gardasil(R) (MSD, HPV vaccine 4-strains) (G), G+PTX, EAE and EAE+PTX. Let’s breakdown first these oddities.  Why the author has included a PTX group, even more adding PTX with Gardasil? There is not much explanation in the text explaining the rationale (also the writing style is odd, very odd. I completely understand that the first author is not a Native English speaker. As an ESL myself, I completely understand that). Also, I am not aware about a higher risk on contracting pertussis upon vaccines.
The second aspect is the use of the EAE mouse model. EAE stands for experimental autoimmune encephalitis. It is the “gold standard” for a mouse model of multiple sclerosis (MS). The idea behind is to inject a brain protein (myelin basic protein or MBP), which will trigger an immune reaction (as the brain is a immune-privileged organ) and result in symptoms similar to MS. I would understand to compare the incidence of Gardasil(R) on MS patients by comparing EAE mice versus non-EAE mice but that is never the case (they even administer PTX to a sub-group).
So here we already start with a wrong experimental design: it just make no sense. A more rationale approach would have been the following:
Vehicle (PBS), Gardasil (G), EAE, EAE+G. That would have saved two groups and precious lives sacrificed in another useless study.

2. The experimental design in terms of statistics and power of analysis

Another important issue is the blatant dismissal of consideration of biostatistics and the power of analysis in the experimental design. For those that are not familiar with scientific research, you have to ensure you have a statistical meaning to your data to ensure the effect observed is real and not due to simple coincidence. This statement is especially true when working with vertebrates animals. Any animal experiment has to be approved by the institutional IACUC that ensure you have a clear idea of what are the purpose of your experiments, how you will ensure a humane treatment to animals and also have an optimal number of animals to achieve a statistical significance.
An important aspect for in vivo (animal) studies is to achieve at least a sample size of 8 or more animals per group (n=8). From Figure 1a, we have already a violation of this as only the G and the G+PTX have enough animals (n=14 and 21 respectively). All other groups are below the n=8 threshold. In addition, you have very different number of animals per groups (control has n=6, EAE has n=5) making the statistical power weak and also restricting the use of common statistical methods such as the use of ANOVA (ANOVA recommends that all groups have an equal number of samples).

3. The experimental procedure and treatment

This is where the firestorm came in: the experimental data. So lets bring this to the table: “Groups of 11 week-old female C57BL/6 mice were intramuscularly administrated 100 μ l of Gardasil or phosphate-buffered saline (PBS) for a total of five times. Ptx was intraperitoneally administrated 2 and 24 hours after immunization. The Gardasil vaccine or Ptx were administrated at 2- weeks or 4-week intervals“.
A key element in a paper is the methods section and this one utterly failed. Based on this information I have absolutely no idea why they injected five times (Gardasil immunization is maximum 3 times), why they injected the PTX right after the immunization (2 and 24 hours, suggesting a double-induction) and when they injected the Gardasil and PTX (how do they separate the 2-weeks versus the 4-weeks? When did they started?).
Also the use of 11 week-old female is not reflective of a human case scenario. If we approximate 1 human year to about 3.6 mice-days, you would expect to use young mice (males and females, to have a gender-balanced study) that are about 6-weeks old (~43.2 days). That would be about 12 years, the age of puberty.
Here we are basically injecting HPV to adult females, which is known to not provide an additional benefit as such population may have already been exposed to HPVs.
The next problem is the injection dose. It is 100microL, thats the equivalent of 0.1mL. A single dose of HPV is 0.5mL. Lets ignore the scale law and assume a mouse is an equivalent to a human. A 50th percentile weight at age 12 is about 40kgs. Lets assume a 6-weeks old mice to be about 20g.
If we assume 0.1mL injection to a 20g mouse, then the human dose-equivalent would be about 200 dose-equivalent injected at once! This is a serious issue because there is absolutely no chance that such things to happen in a lifetime (at grand maximum you may have 3-4 HPV injections, spaced in time). Also, if we assume the age scale, we are expecting to have mice receive their two doses within 48 to 96 hours, not 2 to 4-weeks.
I am not even entering the rationale to inject PTX right after immunization, which is utterly no-sense and just scramble defining the effect of HPV versus the effect of PTX. Another disastrous example of how this paper was flawed from the beginning.

4. Failure to report weight and clinical score

If we want to follow an EAE protocol, it is important to show the evolution of animal weight over a period of times (up to 15-21 days) as well as a clinical score. The clinical score is a well-described protocol in which features found in EAE mice are score from mild (tail flaccid) to severe (inability to move hindlimb or complete immobility).
These two graphs are almost present in any EAE paper outside in the literature.
I assume this is what Figure 1b and 1c wanted to show but very poorly. Indeed Figure 1c does not really show up anything. We dont know when these data have been taken, we have no idea about the onset time of symptoms and furthermore we have no indication of a statistical differences. This is already a waste of data.

5. The constant cherry-picking of the data and incomplete picture

The methods used are honestly laughable: some hematoxylin-eosin staining (a common histological stain that does not tell much unless you have massive brain damage or the growth brain tumor),  Kluver-Barrera staining (for myelin staining), TUNEL stringing (for apoptosis), a behavioral test relegated in Supplementary Figure S1 (in which the author thinks that a P-value of 0.1 has a statistical meaning).

Aratani_Fig2

Aratani_FigS1
Where is the Evans blue extravasation staining to show a disrupted BBB? Where are the GFAP staining to show astrocytes activation? Where are the CD11b and F4/80 staining to show microglial cells activation and macrophages infiltration? Nowhere to be seen. We have to comptent ourselves with some miserable histological staining in Figure 2 and 3. Also no-one of the data about EAE is never shown past Figure 1. Figure 4 is even more laughable as the author only shows the staining of the G+PTX, giving the middle finger to the reader to how such staining looks like in vehicle, or G.

Aratani_Fig3

How can the author be confident that it was the Gardasil treatment, not the PTX treatment (despite being mentioned as a BBB disrupting toxin) being the sole contributor of all this?

6. Conclusions

Another anti-vaccine study, another case of botched science resulting in a junk paper, the sacrifice of animals over a useless experiment. That should not have been passing through the peer-review filter at all because of its deficiencies, yet was able to go through. If I was the reviewer behind it, I would have been ashamed to have this paper not outright rejected for major flaws in the study. Should we assume that the author recommended some complacent reviewers  to this paper? Or should we question the integrity of the editorial board in accepting papers for publication that fail to address some scientific integrity? Again, anti-vaccine studies shows that they cannot challenge vaccine safety and can only make fool of themselves by producing junk studies like this time.

The first and senior authors of this paper produced a paper that is so bad, they should feel ashamed to even had published at first.

 

[Metal/Gothic] Theatre of Tragedy – Aegis (20th Anniversary Release)

Today marks the 20th anniversary of the release of the second album by the Norwegian gothic metal band Theatre Of Tragedy. Listening to it reminds me how great the 90s was for the gothic metal scene and one of the most prolific period in which we have seen most of the metal divas entering the scene and in their trails new genre of metal in Europe and in particular from Scandinavian countries (Norway, Sweden, Finland).
Indeed, that period was the golden age for the second wave coming from Norway. This time a much different style and approach than the first wave made notoriously famous by the church burning and the intestine hatred between some black metal bands (You cannot go without mentioning the name of Varg Vikenes).
This second wave brought one of the most refined sounds to be heard in gothic metal by two Norwegian bands (Tristania and Theatre of Tragedy) supported by some of the most talented feminine voice in metal.
One of them is Liv Kristine, certainly one of the most hypnotizing voice in the “female-fronted metal bands” (I hate that word but I use it applied to the context of that period).
In this second album, named “Aegis”, Theatre of Tragedy learnt from their first album and perfected their sound and Liv her vocal performance. The result? A masterpiece of Gothic Metal. The album is a 10-track album spanning for about an hour.
We get aspired into it straight by the first title “Cassandra” wrapped by the mysterious atmosphere and the vocals of Liv. It is rapidly followed by “Lorelei”, that is more fast-paced than “Cassandra” but in perfect manner. “Angelique”, takes more into a melodic ballad that is reminiscent of a Lacuna Coil. A very relaxing track to listen. “Aoede”, the fourth track, is original and maybe very disorienting because at first we don’t know if they sing in Norwegian on this track until you read the lyrics and assume they wrote in Old English. Thats give the extra amount of mystery and stimulation. The fifth track, “Siren”, is certainly my favorite. By its composition, by its lyrics and by the voice of Liv Kristin. Definitely Liv is the Siren in this song, as we can only succumb to her voice and drown in the waves of melancholy and sadness. I would give any day a shoot to a “Theatre of Tragedy” to a “Leaves Eyes”. This is the kind of track that makes this album a must-have in your collection.
The rest of the album is also very good with tracks such as “Samantha”, “Venus”, “Poppaea”, giving an escape and some well refined gothic metal songs. “Bacchante” is probably my least favorite and its rightfully eclipsed by “Virago”, my second favorite of the album.
If you want to judge the quality of the album, submit it to the test of time and give it a listening session 20 years after the release. While in the 90s, some claimed the metal scene was deadbeat by grunge and alternative, it was only an appearance. It just needed to be searched, to be initiated by a friend or by a savvy record store to realize that underneath the visible, a whole new genre of metal was growing and morphing giving us some of the best sounds of the 90s. And Theatre of Tragedy was one of them.