[Sciences/BBB] Why the vitamin K shot in newborn matters

I have seen the topic of vitamin K (VitK) shot coming over and over in various discussion groups, with some parents weighing the need of the VitK in newborns. One of the main argument in favor for the injection of VitK in newborn is its ability to reduce the risk of cerebral bleeding (cerebral hemorrhage).
I thought a post on this topic would provide a great help in understanding the physiological role of VitK, the consequence of brain hemorrhage and conclude on the importance of the VitK shot.

1.What is the vitamin K?

Vitamin K is a fat-soluble vitamin that is mostly obtained by our gut microbiota and accessory from our food intake (in particular leafy greens and liver).

220px-Phylloquinone_structure.svg

During gestation, the fetus obtains it from the mother, as such vitamin passes through the placenta barrier. Vitamin K plays an important role through its biochemical cycle called “the Vitamin K cycle”. Vitamin K can convert glutamyl residues present in proteins into gamma-carboxylglutamyl residues as depicted in the picture below:

F1.large

Such modified glutamyl residues are present in particular set of proteins called “coagulation factors”. These coagulation factors are important pieces of what we refer as the “coagulation cascade”.
400px-coagulation_full-svg

I know this graph is complicated but what we care here is the final part of the cascade. The presence of intrinsic damage or trauma, we have the activation of several coagulation factors. Amongst those that are VitK-dependent, we have factor VII (seven), IX (nine) and X (ten). Prothrombin, upon activation by factor X  is converted into thrombin, which in turn cleaves the soluble fibrinogen into the insoluble fibrin. Fibrin acts as a mesh and forms a fibrin clot that will patch the bleeding area. This is an important physiological response when you rupture a blood vessel. The coagulation cascade will create a clot that will stop the bleeding process, saving you from a risk of loosing too much blood and entering an hypovolemic shock. One organ is particularly sensible to brain bleed, this organ is the brain.

2. Brain hemorrhage: small numbers, big damage

In this section, I will mostly discuss about brain bleeds in regards of hemorrhagic stroke but you can apply the same pathophysiology to brain bleeds induced by brain trauma. Brain bleeds are the second type of stroke. They account for about 15% of total stroke events, but account for 40% of stroke-related deaths.

subarachnoid-800x416

We have different types of brain bleeds. In stroke, we usually have a type of brain bleed called “intracerebral hemorrhage” (ICH) that happens deep inside the brain. There are other types of hemorrhage called “sub-arachnoid hemorrhage”. In that case, the brain bleeds occurs in the sub-arachnoid space, a space between the brain and the skull. This type of bleed results into an ischemic stroke (due to a lack of blood perfusion in blood vessels beyond the bleed site) and a brain swelling (resulting in the crushing of the brain tissue due to increase intracranial pressure).

During the injury heme (from damaged astrocytes, neurons and red blood cells) is released in the extracellular space. Heme is a very strong pro-oxidant molecule resulting in the formation of radical oxygen species (ROS) such as anion superoxide (O2*-) and hydrogen peroxide (H2O2), which in turn further induce oxidative stress and cellular damage.

The major type of cells that suffers of such damage at the greatest extent are neurons. Neurons are highly sensible to such injury and unlike other cell types neurons do not divide anymore (post-mitotic cells). A dead neuron is a dead neuron. There are some studies suggesting a possible regeneration of neurons in certain brain regions in rodents (mice, rats), yet the presence of an evidence pointing out at similar mechanism in humans are yet to be demonstrated. Furthermore, there is still no evidence that stem cells (including cord blood stem cells from umbilical cord) can provide a repair of such brain region following injury.

As of today, a dead neuron is a dead neuron. The ability of a damaged brain region to recover is very limited.

3. Why Vitamin K shots?

As we just have explained here, we know that VitK is essential in coagulation and we also understood the impact of brain bleed on the brain. Thus, reducing such brain bleed can be done in the short-term by the induction of the coagulation cascade.
As we mentioned, babies get their VitK from the placenta, but by the time they are born, they are already coming with a low VitK. We also mentioned that the VitK is primarily produced by the gut microbiota. It will take weeks if not months for babies to get a gut microbiota that is functional enough to produce the VitK (I speculate that such microbiota is not present until the age of 12 months when baby eat a diet similar to adults). We can speculate that food (breast milk or baby formula) should provide a source of VitK but providing a steady and standardized intake from dietary is near impossible to achieve.
Furthermore, there is no lab tests or techniques that can predict the onset of a brain bleed. Furthermore, brain bleed has a very high mortality rate and very high morbidity rate including cerebral palsy and other brain damage.
Therefore, ensuring a source of VitK right at birth is the best approach to ensure the baby has enough VitK to have a functional coagulation cascade. In case of a brain bleed, we can expect to have a rapid response of the body to ensure a emergency clotting process ongoing until the doctors can intervene and stop such bleeding to happen and clean any possible brain bleed.
This is why it is important to opt-in for a VitK shot. Once a brain tissue is damaged, there is no evidence yet that there is regeneration of such area. Neurons do not divide anymore by birth and there is no evidence yet of stem cells (including stem cells from cord blood) able to repair such damage.

 

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[Punk] Brian Deneke 30/09/1978 – 12/12/1997

Today marks the 20th anniversary of the death of Brian Deneke. His death did not made the international news much, but for those that live in Europe, his death is sharing similarities with the death of Sophie Lancaster (died on 08/11/2007). Many metalheads know about her death, enough to have Delain dedicated a song on her named “We Are The Other” in their album named similarly. Both deaths share the same motivation: murdered because they were different, because they were not fitting the mold of society, because they were ostracized by their attire and their style, because they wanted to live their lives as teenagers using music subgenres as a vehicle for their catharsis. Brian found in the punk culture a liberating moment, Sophie found in the goth culture a liberating moment. Myself found mine in the doom metal culture.
I heard the story of Brian only recently, as a stranger, from my few years living in Yellow City. Some call it Bomb City, because of the nearby Pantex assembly that constitute some vital part of the economy. But I felt in the story of Brian, the same story that many others that do not fit to the mold of the society feel: being labelled as a misfit, as an outcast, as an indesirable of the society.
Interestingly, Brian and me share roughly the same age (+/- 12 months), come from the same generation (Gen Xers) and have been in our troubled teenage years in the same time. We both are not actual to the punk movement, if we consider the movement “golden age” was in the end 70s-beginning 80s. Yet, the punk movement was alive and kicking proud in the 90s. Some classmate embraced punk as a way to rebel against the system, embracing some anarchist ideas. “Fuck the system!” was the motto. I was like Teflon to embracing punk: it did not stick long on me. I rather was seeking the melodic riffs of the other major genre that grew alongside the punk movement: the New Wave of British Heavy Metal. I was driven by the escaping and incensing tunes of Iron Maiden, swinged by the lyrics of Bruce Dickinson and the guitar melodics. Sure, I was an outcast, I was a nerd and I was the dude that sit quietly in a group of friends. But what I experienced was nothing to what Brian and many teenagers in the US experienced.
As an European, the only exposure to US teenage years was through the TV: you had the posh and falsely glamour pictures from “Beverly Hills 90210” or “Melrose Place”, the funny “Saved By The Bell” (although sometimes opened discussion to serious matters) or what appeared more realistic as “21 Jump Street” (by discussing some real issues). But the one that was really in phase with me was MTV “Daria”. She was also an outcast in her sense, nixing being part of the cheerleader team or being the girlfriend of the football star. She was also a punk at heart along with her friend Jane, without having a crush on Trent (Jane’s brother) that was playing into a rock band during the weekends. It also showed me that the meaning of belonging to a tribe in the US was really much more amplified in the US than it was in Europe.
Where I had no pretension for a music career, Brian saw himself part of the local punk scene dreaming of becoming a leader for a punk band that could make a living of his art. The events that lead to Brian’s death are unknown to me and mostly garnered from reading on different sources.
It seems all started on the IHOP facing the former Western Plaza mall (what seems to be the current strip mall located on I-40@Western corner). On Saturday December 6th, an altercation occurred between Dustin Camp (a honor student and star football player at Tascosa High School) and John King, a member of the local punk scene. For those who are not familiar, Tascosa High School is usually considered one of the most preppy public HS in Yellow City, surrounded by the posh Tascosa neighborhood (since the Colonies neighborhood claim the title of “posh neighborhood). There are contested claims that Camp tried to run on King and his group with his Cadillac, some claiming King hit Camps windshield with a baton. On Friday 12th, Camp and King (alongside their group of friends) set a showdown in front of the same IHOP at 11:00pm. During the fight, Camp retreated in his car and ran over Deneke in an apparent hit-and-run.
The trial was set on Camp with a first-degree murder. The defense attorney, Warren Clark, apparently try to divert the attention of the jury by ostracizing and trying to put the blame on the punk community. Considering the Yellow City community, putting blame on the misfits is an easy target by portraying them with some infamous cliches: “They are lawless, they worship Satan, they are punks.” These are the same kind of stuff we metalheads have to go through: our music is noise, garbage. We are Satan-worshipper, we have tomb-destroyer. We are evil incarnate.
The trial concluded with Camp found guilty as involuntary manslaughter, 10 years probation and $10’000 fine. This is a very mild sentence for someone that voluntarily (according to witnesses) run over Brian and left the crime scene. This case has possibly some signs of “affluenza” in which the social position of the person prosecuted is used to downplay the severity of the crime (“He is a good boy! He is in the Honors list! He is the football star player!”), something we have been already seen. Nevertheless, he was arrested in 2001 for underage drinking, followed by charges on false statement to police and ultimately sentenced in September 2001 to 8 years in prison for violating his probation.

More recently, a movie documentary named “Bomb City” retracing the story of Brian Deneke has received some remarkable standing ovations and awards at various film festivals. I would strongly recommend to watch it and I hope to attend the local airing.

Although the punk and metal scenes rarely mingle (although some can argue that thrash, death metal are intertwining of punk and metal), the small scene in Yellow City make us closer. We don’t have much gigs in town, so when we have some local bands performing in a bar, it is more than welcome. I try to think that what if Brian was still amongst us, maybe I would have been sitting in a bar watching him perform and enjoying his gig.

However, I will never have that chance. I really feel sorry for Brian’s parents about what happened to Brian. I wished I could give them “my sincere condolences” even 20 years after the facts. Rest in Peace Brian 😦

[Metal/Melodic Death] Arch Enemy @ Sunshine Theater (Albuquerque, NM – 12/02/2017)

It has been a bit more than three years since I discovered @ArchEnemy via their album “War Eternal” and it was basically a love at first sight. It was a very big surprise because I am not much driven into death metal bands, but what makes Arch Enemy unique is the remarkable Michael Amott guitar melodics, now seconded by Jeff Loomis (ex-Nevermore). I even got into the feud between “team Angie” versus “team Alissa” (spoiler: Alissa really gained a lot of experience since War Eternal and you can get that from “Will to Power”)
I quickly ended up getting the whole discography but one thing I could not get is to attend one of their gig. Arch Enemy tours a lot in Europe, but having them tour the US is damn hard. So when they announced they will tour and have a stop in ABQ (that cuts my ride to 4 hours instead of 5.5 hours), it was a bliss for me. Top it with a show on Saturday night and that was just having the perfect astral alignment to attend it.
The only issue I have was the line-up: Fit for an Autopsy (Deathcore, Jersey City, NJ), While She Sleeps (Metalcore, Sheffield, UK) and Trivium (Groove Metal, Orlando, FL). Nothing that talk to me much, especially when you were bottle-fed with some fine European metal bands. I find it interesting the relative cleavage in the metal styles between the Old and the New continent: the former is complex, well refined and musically challenging whereas the later often is more brutal and primal in its composition. TL:DR, none of the lineup bands got my interest, they failed to impress me so I will not talk about them and move to Arch Enemy.
What was interesting though was the number of folks that left the show right after Trivium, especially teenagers and college kids. This was just like any swimming pool in a hotel: time for kids and family ran out and time to start the adult swim.

IMG_0954

Oh boy, 75 minutes of pure joy (and few bruises and weightlifting :p) and also a chance to sneak in closer to the front row. We get into the performance accompanied by “Set Flame  To The Night” with Daniel coming behind the drums and starting them like a horde of B52s flying over with the guitar riffs of Michael and Jeff with “The World Is Yours”. You know what, just watch it:
https://youtu.be/0GkXBRXXMv4

This is Arch F**king Enemy! 75 minutes of rage, furor, mosh, marvelous performance of the band! Alissa clearly walk the walk, learnt from the mentorship of Angie. Damn it feels good to listen the band live.
The band played through their whole repertoire from “Wages of Sin” until “Will to Power”.

 

 

We started with “The World Is Yours”, followed by “Ravenous” (not much my favorite though), “Stolen Life”, “My Apocalypse”, “War Eternal”, “Bloodstained Cross” (Ohhhhh yeah! As brutal than Angie!), “As The Pages Burn”, “The Eagles Fly Alone”, “We Will Rise”, “Avalanche” followed by a solo by Jeff transitioning into “Snow Bound” (one of the best guitar solo by Michael on “Wages of Sin”) and finally concluding to “Nemesis” (the classical AE song and also a good metric to compare Alissa versus Angie).
If I have to pick a concert that was awesome to me, this is THE CONCERT that I found the most awesome. Really looking forward to see them again soon!

[Neurosciences/Aluminum] Does the latest paper from Exley show a link between ASD and aluminum?

Someone brought my attention today about the most recent Exley paper out in the press titled “Aluminium in brain tissue in autism” (the title could have been better but well….) and published in the journal “Journal of Trace Elements in Medicine and Biology“.
Let me put this straight, this is not a paper that has evidence of scientific fraud or data manipulation. There is no duplicated images, no suspicious blots. The problem I have with this paper is its deep experimental flaws and data analysis that nonetheless should not have passed through the peer-review filter.

  1. Before we dive into the paper, lets put the paper into context
    Lets just put the paper in the context. It was received on October 26th (Thursday). Came back in its revised form on November 21st on Tuesday and accepted for publication on November 23rd (Thanksgivings for the US, but since the editor-in-chief (EIC) is in Europe no Thanksgiving here). Let that sink it a bit: in a bit more than three weeks, it got send to review, came back from review and got revised in 26 days. In my standard of reviewing for journals and publishing my papers, thats some faster-than-light peer-reviews. I usually wait 4-5 weeks by the time I submit mine and get the editor reply to my submission with the infamous reviewers comments. Does a fast-reviewed manuscript means a bad manuscript? Not necessarily, but it can mean that maybe the peer-reviewed was not optimal, rushed or even worse just botched. Based on the quality of the data presented, I am leaning towards a botched review. Thats quite disappointing because the journal holds a decent impact factor (~3 for 5-year impact factor) and you expect an okay review.
    Then comes another problem. Exley published this paper (as well as few others) in the journal…..in which he holds a seat in the editorial board. Nobody can exclude the possible conflict of interest. Consider that: if you were an EIC, would you provide the same rigor and objective decision on a paper submitted by a colleague sitting in your editorial board than a paper submitted by Doe and colleagues?
    Not forbidden, but if you can avoid it, avoid it. Transparency is key and publishing in diversified journals (unless it is society-official journals) is an indicator of an healthy research.
    Finally, the last thing to keep in mind before I deconstruct the paper is the funding source. According to the acknowledgment section “The research is supported by a grant from the Children’s Medical Safety Research Institute (CMSRI), a not-for-profit research foundation based in Washington DC, USA.”  Behind the fancy name is just another anti-vaccine foundation that will play “the vaccine safety” card to peddle their pseudosciences. So we can claim that Exley is a shill for CMSRI, since he received monetary support for his research. Does that mean the research is completely bogus? No, but it means it will require further scrutiny, especially when the claim of the study goes against the consensus in the field (aluminum in vaccines is safe).
    Same goes if a study funded by Big Tobacco claimed the absence of correlation between lung cancer and smoking or if Big Sugar claimed the absence of correlation between type 2 diabetes mellitus and consumption of sweetened beverages.
  2. So what is wrong with this paper?
    For those who wants to read the paper with me, you can download it here (I assume it is open-access, so you should not have an issue with the paywall). Exley has a publication record on aluminum, especially when it comes to its possible ecotoxicity and the impact of aluminum on certain biological processes.
    The introduction is damn short, half a page of a double-spaced document but set the tone, this study will investigate the relationship between autism and aluminum in the brain.
    Samples are obtained from the Oxford Brain Bank, but felt short to indicate the source of the tissue (like a catalog number) and how this source of materials was complying with the institutional review boards (IRB). Basically, for any research involving human subjects or human tissues, you have to comply with the IRB that such specimens are used for a certain and defined use and foremost been anonymized.
    We have 5 patients that were diagnosed as on the autism spectrum and immediately we can pinpoint an important issue: there are no controls and that’s one of the big and unforgiving flaw of this paper.
    The authors then used two techniques to localize and quantify the Al in different cortical regions (and sometimes hippocampal regions). They have used three technical replicates (random sampling from the same cortical lobe) for measuring the Al content using an atomic absorption spectrometry and used lumogallion (aka4-chloro-3-(2,4-dihydroxyphenylazo)-2-hydroxybenzene-1-sulphonic acid, a fluorescent dye initially described to localize Al in plant roots). This dye have an excitation/emission spectra close from FITC/Alexa Fluor 488. It has been also used for live cell imaging , in particular to study how macrophages process Al present in vaccines adjuvants (http://www.sciencedirect.com/science/article/pii/S0022175915001222).
    Considering the equipment mentioned in the method, the microscope used provides the right excitation bandwidth filter and provide a long pass emission filter for anything over 510nm.Then things get weird, in the result sections, the authors mention the following:”We examined serial brain sections from 10 individuals (3 females and 7 males) who died with a diagnosis of ASD and recorded the presence of aluminium in these tissues (Table S1).“Where is the number coming from? Why don’t we have the same numbers in the Materials and methods?The other problem is the over interpretation of the data. To be brief, the lumogallion will show some punctuated pictures. The authors show some brightfield pictures overlapping to show the tissue structure but does not really help the reader. A DAPI stain (to stain cell nuclei) as counterstain would have been much more informative, it would helped to distinguish background noise from possible Al inclusion. Again, keep in mind we have no controls. The other issues with immunostaining is the high risk to cherry pick the data. You will be naturally inclined to show the presence of a positive risk but this cannot be used for quantitation. Thus, the use of the second method is welcomed as a complementary technique.
    For those not familiar with fluorescence, there is an important notion to keep in mind when analyzing the data: ensuring you keep the same exposure time, the same brightness or contrast and foremost have a negative control to set your exposure time. You can see a sketch explaining here on one of my fluorescence staining (based on my data, I concluded the expression was weak if not negative).

    Slide2

    The background subtraction is also a bit weird. I acknowledge the assessment of autofluorescence is a good control, but you expect to see a low staining. But foremost, you cannot overlap two distinct slices, as proximal as it can be. For instance, in Figure 1, you see some lumogallion staining and below the fluorescence  from the “control” using the adjacent slice. The lumogallion also seems to have a very high background.
    Picture1
    It seems lipid-rich environment increase dramatically the fluorescence of lumogallion (if you look at the spectra, the dissolution of the dye in Triton-X100 solution (b, a detergent) dramatically increase the excitation and emission spectra compared to water (a)).
    What I found troubling is this sentence in the results section: “We examined serial brain sections from 10 individuals (3 females and 7 males) who died with a diagnosis of ASD and recorded the presence of aluminium in these tissues (Table S1). Excitation of the complex of aluminium and lumogallion emits characteristic orange fluorescence that appears increasingly bright yellow at higher fluorescence intensities. Aluminium, identified as lumogallion-reactive deposits, was recorded in at least one tissue in all 10 individuals. Autofluorescence of immediately adjacent serial sections confirmed“.
    If you are a bit a fluorescence microscopy savvy, you know that the “emission color” we see in the objective is never caught by the CCD camera. These camera have in the most majority a B&W output for the simple reason that they have a much higher sensitivity than color cameras. You can always re-create colors in the micrograph pictures using various “lookup tables” (LUTs) that will give a pseudo color based on the level of grays. This is very useful when you samples different excitation/emission channels (for instance, samples stained with DAPI and two antibodies, one conjugated with Alexa Fluor 488 and the other with Alexa Fluor 546 or further down).
    The problem inherent with fluorescence is you can make thing fluoresce or end up with a false-positive signal if you increase the light beam (usually never happens because it is set) or if you increase the exposure time of your camera (this is the most common issue). As you increase exposure, you increase the risk to capture non-specific signal like autofluorescence signals.
    The other problem here is how to explain this sudden shift from orange to yellow?  This seems more like a subjective observation than something caught on camera.  That can be due to different things. You can have some bleed-through of the dye that is normally emitting in a certain wavelength but if it is strong enough can appears in neighboring emission channels. This thing rarely happens with a good fluorescence microscope that have defined filter cubes that allows the diffusion of certain emission wavelengths (for instance, my microscope have a DAPI, Alexa Fluor 488 and Alexa Fluor 555 cubes that only let the respective emission wavelengths  with 20nm-margin error to cross through the objective and reach the camera and binocular).
    Usually, we have to deal with bleed-through when you use flow cytometry and usually is solved using fluorescent dyes latex beads and by following a protocol called “compensation” (this has the result of removing any noise and keeping only the signals).
    We cannot also exclude that such fluorescence is just an autofluorescence from lipofuscine inclusion bodies. Lipofuscin is a lipid-based compound naturally produced by our cells. It has an important concentration in the central nervous system, however it is normally cleared out by cells. Failure in the clearance of lipofuscin is associated with different diseases called “lipofucsinosis” such as Batten’s disease. Even the author admit the possible presence of lipofuscin inclusions “Intracellular aluminium was identified in likely neurones and glia-like cells and often in the vicinity of or colocalised with lipofuscin (Fig. 5).” Lipofuscin is also capable of autofluorescence, although it is more in the wavelengths matching DAPI. Lipofuscin has an excitation/emission peaks at 360 and 435nm respectively but has been reported to also show fluorescence at 510nm when excited at 488nm (https://www.sciencedirect.com/topics/neuroscience/lipofuscin).
    Compared to the lumogallion excitation/emission spectra (507/567), we cannot exclude the presence of a phenomenon called “FRET” (Fosterman Resonance Energy Transfer) in which the excitation of lipofuscin (as the microscope excitation bandwidth is 470-495nm) provide enough energy to the photons emitted by the lipofucsin to excite nearby lumogallion dyes. Because the microscope setting used in this paper has no restricted bandwidth (it let pass any photons harboring a wavelength of 510nm and more), it may explain this orange-to-yellow transition noted by the author. The presence of a DAPI nuclear stain would greatly helped to identify this region as grey matter (rich in cells) or white matter (rich in lipid-rich myelin sheets). Thus, we can legitimately questions the nature of these as it these punctae labelled as “Al inclusion” are simply lipid inclusion or some artificial noise due to the tissue processing. This is where controls come as critical, it can help you sort the signal from the noise.

     

    The second big issue with this paper is the over-interpretation of what the experimenter see. The experimenter wants to see Al inclusion in monocytes? So be it: “Aluminium-loaded mononuclear white blood cells, probably lymphocytes, were identified in the meninges and possibly in the process of entering brain tissue from the lymphatic system“. Or maybe these are astrocytes, or neurons, or microglial cells, or blood vessels….or whatever the author wants to believe in: “Aluminium could be clearly seen inside cells as either discrete punctate deposits or as bright yellow fluorescence. Aluminium was located in inflammatory cells associated with the vasculature (Fig. 2). In one case what looks like an aluminium-loaded lymphocyte or monocyte was noted within a blood vessel lumen surrounded by red blood cells while another probable lymphocyte showing intense yellow fluorescence was noted in the adventitia (Fig. 2b). Glial cells including microglia-like cells that showed positive aluminium fluorescence were often observed in brain tissue in the vicinity of aluminium-stained extracellular deposits (Figs. 3&4). Discrete deposits of aluminium approximately 1m in diameter were clearly visible in both round and amoeboid glial cell bodies (e.g. Fig. 3b). Intracellular aluminium was identified in likely neurones and glia-like cells and often in the vicinity of or colocalised with lipofuscin (Fig. 5). Aluminium-selective fluorescence microscopy was successful in identifying aluminium in extracellular and intracellular locations in neurones and non-neuronal cells and across all brain tissues studied (Figs.1-5). The method only identifies aluminium as evidenced by large areas of brain tissue without any characteristic aluminium-positive fluorescence (Fig. S1).
    This is the second big mistake of this paper. If the author wants to make the claim he proposed here, then he has the obligation to show a counterstain using selective markers for neurons (e.g. MAP2, bIII-tubulin, NeuN….), astrocytes (e.g. GFAP), microglial cells (CD11b), leukocytes (CD3), macrophages (CD45), blood vessels (e.g. PECAM-1, claudin-5). This could have been easily performed (using a secondary antibody conjugated with Alexa Fluor 555 or better Alexa Fluor 647)  and would have give support to this claim.
    If the author can identify cells by the naked eye, he is either equipped with  Superman X-ray eyes or he is just imagining things.

    The discussion quickly gets into an anti-vaxxer diatribe and throws the minimal amount of scientific data under the bus.
    For example, the author throws this sentence as is: “We recorded some of the highest values for brain aluminium content ever measured in healthy or diseased tissues in these male ASD donors including values of 17.10, 18.57 and 22.11 g/g dry wt. (Table 1).” Firstly, where does it get this data? You cannot sum technical replicates, you have to average them (even with considering the huge variability between technical replicates). Secondly, how can the author make a claim like this without providing values from controls (well there are no controls) or from the literature. It is like “we have recorded the highest amount of leukocytes in ASD patients blood samples with values of 11.3, 12.0 and 11.5 x10e3 cells/mm3.” I cannot make an interpretation or conclusion without knowing the reference from the normal population (normal range 4.5-11x 10e3 cells/mm3) or from control groups. The average Al level was 2.38-4.79 microg/g tissues in male ASD and 1.15 in the female ASD patient. Such levels were very similar to those reported in samples from patients suffering from familial form of Alzheimer’s disease.
    Slide3

    The data is interesting but we are lacking additional female samples to make a claim as he did: “All 4 male donors had significantly higher concentrations of brain aluminium than the single female donor.” He lacks the proper conditions to run the statistics (you need same number of patients in male and female to make such claims) and even the important inter-individual variability makes it unlikely that he could achieve the statistical significance. This is a statement that would put a graduate student in shame for overconfidence in the data.
    Then goes the tirade “What discriminates these data from other analyses of brain aluminium in other diseases is the age of the ASD donors. Why, for example would a 15 year old boy have such a high content of aluminium in their brain tissues? There are no comparative data in the scientific literature, the closest being similarly high data for a 42 year old male with familial Alzheimer’s disease (fAD) [19].” (another Exley paper published…..in the same journal). We are dealing with the same issues (lack of controls, huge variability in the technical replicates…..).
    Now if you plot the average patient Al levels agains the age, regardless of the condition, you end up with an homogenous cloud. Now, two things have to be noted here: seems there is no impact of Al levels based on the disease (only age seems to matter between ASD and AD) and there is no correlation between increase in brain Al and age, at least in the very small sample size.
    Data 2

    No pun intended, but the data scatter looks vaguely like the United States map. Again, it shows the need of data from asymptomatic patients to estimate the burden of Al in the brain.
    Since we have not access to Al content in the brain, we have to see some values in the literature. A study by Andrasi and colleagues (https://content.iospress.com/articles/journal-of-alzheimers-disease/jad00432) provide some Al levels in control samples. According to their study, the average Al content in control samples were between 1.4 to 2.5µg/g dry tissue. We are indeed not far from the value reported by this study, especially when you consider the important standard deviation in these samples.

    Maybe it is also to consider the other study by Exler on Al level in brain samples from patients associated with familial form of Alzheimers disease (fAD) and familial dementia. In that study, all reported with Alzheimers (some with early onset, some with late onset based on age), the Al values reported were ranging from 0.34microg/g tissue (male) to 6.55microg/g (female, presenting a mutation in the PSEN1 gene, a known gene in FAD). So are we just measuring noise and try to extrapolate data from noise? Thats some bold statement that should have been smashed already by a decent reviewer in the field of neurosciences.
    But seeing these two papers went through in a apparent free ride is not looking good for the journal integrity.

  3. Conclusive Remarks
    To make a claim is one thing, to back it up with robust data is another thing. I think Exley jumped the shark a while ago and started to aluminum as the big bad wolf in every little things. But a wolf can be tamed, kept out from showing danger to the community and somehow co-exist. But for Exley, like Shaw, like Gherardi, aluminum is the devil incarnate. God forbid it has been used for 70 years and showed barely more than simple coincidence in its association with some disease, aluminum is their dead horse that worth being beaten again and again. If your funding sponsor will give you money for showing a link between aluminum and autism, lets give them what they want. Ethically it is insane, but when you need to keep your lab and your faculty position afloat, sometimes making the pact with the devil and throwing the scientific integrity and the philosophism that is given to you  following your thesis defense can be tempting. Sometimes, it feels that anti-vaccines researchers are like Faust and succumbed to the offer made by Mephistopheles offer. But this come with a price and a hefty price to pay: the loss of your integrity as a scientist.
    So my question is what is coming next to patients on the spectrum: does this study will be used to support the anti-vaccine agenda (another reason to yell “Aluminum is a chemikillz” in parenting groups?) and breakdown the herd immunity? Bogus remedies by bleach enemas and drops (the infamous CD/MMS)? or give a support to chelation therapy? gluten-free/casein-free diet? Or like Exley once claimed have these people drink ad nauseam silicon-rich water like Fiji water or Volvic water with the magic claims that the silicon with drain your brain from the Al contained inside it?
    This kind of deeply-flawed studies, lacking proper controls and driven by an ideology over the facts are dangerous because they prey on the meek and enrich modern snake oil sellers.

 

[Sciences/BBB] About the Thanksgiving tryptophan comatose and the BBB

Happy Thanksgiving everyone, I hope you are enjoying your family gathering. I know many of you are dreading to meet the family and extended family to discuss about controversial topics and differences in opinion.
But the other big menace coming in, that is particularly feared by the Black Friday shoppers: “The Thanksgiving turkey comatose” myth. This myth is perpetuating the idea that the Thanksgiving feast will induce a lethargic state attributed to the tryptophan present in turkey. Lets use this time to talk about tryptophan, turkey and of course the BBB in all that.

1. What is tryptophan?

Tryptophan is one of the 22 amino acids forming the building bricks of each of our proteins. It belongs to one of the few amino acids that our body cannot produce and therefore has to get it from our food supply.
In addition to its role in proteins, tryptophan is also an interesting molecule for the central nervous system, because it serves as a precursor for serotonin (a neurotransmitter also known as 5-hydroxytryptamine) and melatonin (commonly known as the “clock hormone”). You can see the similarities in structure of these molecules below:
Picture1

Tryptophan is particularly enriched in meat. According to the USDA, turkey meat contains the highest level of tryptophan from all foods, followed by white eggs, soybean and seaweeds. This partly support the claim of turkey being rich in tryptophan.

2. How does the tryptophan enters the central nervous system?

Like you expect, the blood-brain barrier is impermeable to any charged molecule. This is the case of many amino acids circulating in the blood (pH=7.4). Thus amino acids can enter the brain only by using special “revolving door” called solute carriers (SLCs). Tryptophan is transported by a particular amino acid transporter called large amino acid transporter 1 (LAT1). LAT1 is a particular transporter because it is formed by two subunits named SLC3A2 (also named CD98) and SLC7A5.
LAT1 is not specific to tryptophan, it also allows the transport of other aromatic amino acids like phenylalanine and tyrosine, but also chained amino acids such as leucine or arginine.

The impact of dysfunction in LAT1 remains poorly understood, however a study by Mykkaenen and colleagues noted several point mutations in SLC7A7 with a rare disease named lysinuric protein intolerance, a rare autosomal disease primarily described in patients from Finnish and Japanese origin marked by the impaired transport and elimination of basic amino acids following a protein-rich diet.

3. What is the function of tryptophan in the brain?

As I have previously mentioned, tryptophan is the precursor of two major neuromediators: serotonin and melatonin.
Serotonin is produced by a certain type of neurons named “serotoninergic neurons”. Like other neurons expressing a particular neurotransmitter other than glutamate or gamma-aminobutyrate (GABA), these neurons are restricted to a certain localization usually referred as “nucleus” (kernel, core). These neurons can project their axons all through the brain via a process called projections, allowing these neurons to interact with far-fetched neurons localized in a remote location.

In the case of serotoninergic (5-HT) neurons, these neurons are located in a structure called “raphe nucleus” and project to areas in which such neurotransmitter interact with 5-HT receptors. Through the interactions with the receptor, serotonin plays an important role in the modulation of several behavior including appetite, emotional (depression, anxiety), cognitive (schizophrenia) motor and autonomous (for instance emesis, the scientific term of “puking“).

In addition to the biological effects on the brain, the serotonin system is also linked to the circadian rhythm system (what we can call the “biological clock”) as depicted in the picture below:

sadserotoninfigure

We are diurnal animals as our main activity occurs during daylight and concludes with our sleep cycle during the dark period. In opposite, some animals like rodents are nychthemeral animals (active during dark phase and sleeping during daylight).

The light/dark cycle phase is determined by our eyes and retina. Such retina will transmit the presence of light to a particular nucleus named “suprachiasmatic nucleus” (SCN) . This nucleus is consisted by cells and nuclei functioning as oscillators. You can think about a pendulum in perpetual movement or a ticking clock. When darkness settles, the retina start to slowdown the information coming to the SCN.
In turn, the SCN becomes less active and relieve the blockade of the activity of the pineal gland. The pineal gland in turn start to secrete melatonin (aka the sleep hormone) that act as a “negative feedback loop” further shutting down the SCN and stimulate the production of serotonin via the raphe nucleus. All these events ultimately giving us the feeling of being sleepy and the process of sleeping.

4. So why we claim the “turkey comatose” is real?

As you can see in this myth, we are facing a post-hoc ergo fallacy. “I feel sleepy after Thanksgiving dinner. I ate large amount of turkey meat at Thanksgiving dinner. Turkey contains tryptophan and sleep is controlled by melatonin (a tryptophan derivative). Thus the tryptophan contained in the turkey meat is responsible of the food comatose”.

As you have seen, this does not make sense as the sleep/wake cycle is driven by the light exposure. This is also explaining partly why some people feel more tired and less motivated during winter times.

One explanation we can discuss is the particular food intake we all face during Thanksgiving that exceed our usual amount of food. We rarely experience such a feast and copious meal during the year. The table is furnished with so different plates, rich in proteins and carbohydrates.
This create a spike in food intake and food digestion that will likely create a urge of blood flow towards the gastrointestinal tract. This physiological phenomenon is named “postpandrial torpor”, making you feel sleepy and tired after a large meal, even if the meal was completely turkey-free.

So in conclusion, if you want to avoid “the turkey comatose”, don’t blame it on the turkey. Blame it on your eyes having a bigger appetite that your stomach can sustain. Keep it in moderation and now you know about the tryptophan transport at the BBB.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

[Metal/Gothic] Evanescence – Synthesis (85%)

ortexAfter several years of hiatus, Amy Lee and Evanescence broke their silence by releasing “Synthesis”, an album providing a re-orchestration of their previous songs. Amy dropped the guitars and the drums to let place to other instruments. Amy is already gifted by a beautiful voice and a talent to play piano. When Evanescence mixed her voice with her piano melodics, this is where my auditory and limbic system explode into a serendipity.
Evanescence played a sort of gateway for me to explore the symphonic metal genre (this is how I got introduced to Nightwish and Within Temptation) but never went fully symphonic. How does the album stands without metal riffs and drums?
It stands good, some songs are impressive by their intimacy and their simpleness, some are kind of a disappointment. Let’s go through together the 16-tracks spanning a bit more than 63 minutes.
We start with “Overture”, an instrumental opening track that introduce us to the piano and violin tones that are the trademark of the album. The first track is the cover of “Never Go Back” from their third album “Evanescence”. Listening to the piano opening sequence is just a thrill on the skin, giving me some goosebumps. This is an example of a successful cover by its ability to remove the heaviness of the original and still sounds so wonderful to the ears. The third track, “Hi-Lo”, was one track that was initially planned to be in “Evanescence” but was not retained. As is it is, it is such a beautiful and intimate song that makes you wonder how it would have sounded in its metal version.
In the opposite, “My Heart is Broken” (the 4th track) sounds much better in its original cover because it already provided such a wonderful blend of the metal, piano and Amy’s voice. The 5th and 6th tracks were sounding as good as the original cover are “Lacrymosa” (The Open Door), “The End of The Dream” (Evanescence).
“Unraveling”, the 7th track posing as an instrumental and melancholic piano solo interlude is sounding so beautiful by its own.
“Imaginary”, the 8th track, is one of the biggest disappointment. The original one, already present in their EP “Origin” was simply perfect as it was, a perfect blend of Amy musical and vocal performance with the heaviness. It was one of my favorite of “Fallen”. This cover just axed it and fail to give justice. What the heck Amy on that one?
“Secret Door” is as good as the original one. Both were instrumental in their conception, very light and sober so they are very comparable. Some of the finest compositions of the band and kind of make you wish they pursued into the direction of Within Temptation or Nightwish. “Lithium”, another of my favorite from “Fallen”, also received the good treatment even amplifying the beauty of the original version.
“Lost in Paradise”, the 12th track, is another failed attempt and liked the original more than the cover in this one. “Your Star”, contrasts and shows how a cover can give back some justice to a track often went unnoticed in the original album (The Open Door).
Track 14, “My Immortal” is the litmus test. I have listened to two versions of the track back in the days: the radio edit version that was heavy and one bootleg downloaded from P2P (Kazaa or eDonkey, I don’t remember) that had the instrumental segment purely on the piano. The cover one revives the bootleg ones, embellishing it and making it even better. Damn, I wished this track and “Bring Me to Life” were not appended to the cinematic turd called “Daredevil”.
“The In-Between” is a piano solo new song written for the album. Oh my! Such a delight of listening Amy playing the piano on this one and emphasize the grave notes giving this melancholic feel to the song. Nobody plays the piano in the metal scene than Amy.
“Imperfection”, the last track is the last “meh” on that album. Not a big fan of it, sounds like a mishmash of styles that makes in my opinion an undigestible thing to listen.Be a judge by yourself you can listen to it below.
https://youtu.be/7_XoKJP1XK4

Now I am really mad and disappointed i missed on the latest Evanescence tour (due to scheduling conflict) because after 15 years and the loss of Ben after “Fallen”, Evanescence still stand strong of its feet and really should explore some facets of the symphonic metal. A recent meet and greet between Amy and Simone Simons (Epica) could open up some collaboration, especially with some input from Mark.

 

 

[Metal/Gothic] Beyond Forgiveness – The Great Wall (80%)

US metal bands playing Gothic Metal or Symphonic Metal are fairly rare birds, in contrast to Europe in which I found the market on the verge to precipitate. Beyond Forgiveness (Colorado Springs, CO) is one of these band that attracted me attention with their EP “The Ferryman’s Shore” and released a couple of weeks ago their first album “The Great Wall”.
How does it fare for a first album? It fares very well, navigating into the current of gothic, symphonic and folk metal keeping upmost of the musical composition “hand-crafted” (in others not over-relying on the keyboards and the studio processing). The result is very pleasant, maybe lacking some ambitious songs that hook you on, but a very nice escaping album that result me in calling the band some metal “troubadours” telling tales, fables and fiends straight out of the middle ages.
This is an album spanning over 10 tracks spanning over 58 minutes. The good point is all the stuff inside are all original materials, no rearrangement of existing materials from their previous EP and singles.
We get into it by the “End of Times”, their first track. It is setting the tones: some good classic metal tones, from the early days of gothic and symphonic metal. Personally, I like that especially considering that a lot of new comers playing the “symphonic” card abuse and overuse the keyboards and post-processing. We quickly transition into the second track and third tracks “The Great Wall” and “Sanctuary”. Good stuff so far. “Imprisoned” fare into more the symphonic metal than the previous track.
“Interlude”, an instrumental melodic ballad track gives the rest of the wandering bard, very nice interlude allowing to appreciate the guitar plays from Richard and Greg. This interlude track also announce a transition in the album, towards a more symphonic type of tone. Whereas “Moment of Truth” sounds more like the first half of the album, “Never Before” has this kind of old-style Nightwish style into it that is not bad. “Dream Before I sleep” is what I call an acoustic track with the voice of Talia accompanied mainly by a piano solo. Good. “I will fight Till The End” brings back the heaviness, just for this track as the last track “Every Breath” brings it more a folk feeling to the music.
Overall the album is quite good but I failed to find something to hook me on. Usually I have one or two “must-listen” song in an album but not this time. The album is very homogenous, maybe a bit too much. Something is still missing to make it a great, I don’t know something bold.
Nevertheless, it is a good starter, give a try and if you like your metal seasoned into some old school gothic and symphonic, give these guys a try, their album is available on Bandcamp and iTunes.