Neurodegenerative disease

Plasmalogens, Part 2

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This post compares Dr. Goodenowe’s clinical trial mentioned in Part 1 with other researchers’ human plasmalogen studies this decade. One of its findings was:

“Figure 1A illustrates that plasmalogen precursor DHA-AAG dose-dependently elevated both direct and indirect target species [DHA-PL, DHA-PE, and (LA + AA)-PL] and had no effect on levels of biochemically unrelated PE species index (LA + AA)-PE.

  • DHA-AAG had a greater elevating effect on its direct target, DHA-PL than its indirect targets.
  • The 1-month washout period resulted in decreased levels of both direct and indirect target species and no effect on unrelated PE species.

Figures 1A,B illustrate that DHA-AAG is converted to its direct and indirect target species in humans as predicted from animal studies on similar AAG plasmalogen precursors (Wood et al., 2011d).”

fcell-10-864842-g001A

Given this century’s background of numerous animal studies, there’s a need to know what translates to humans. Here are the three most recent human plasmalogen studies in descending order where I could access the full study:

2022

“Forty unmarried male students aged 18–22 years (20 in the plasmalogen group and 20 in the placebo group) were randomly allocated to either plasmalogen (2 mg per day) or placebo treatment of 4 weeks’ duration and ingested two capsules of 0.5 mg plasmalogen or placebo twice daily.

  • The primary efficacy outcome was the Total Mood Disturbance (TMD) T-score of POMS 2–Adult Short.
  • Secondary outcomes included the seven individual scales of POMS 2, other psychobehavioral measures (Athens Insomnia Scale and Uchida-Kraepelin test), physical performance test (shuttle run, grip muscle strength, and standing long jump), plasmalogen levels in plasma and erythrocytes, plasma levels of brain-derived neurotrophic factor (BDNF), urinary 8-hydroxy-2′-deoxyguanosine (8-OHdG), body mass index, and percent body fat.

Lipid composition of purified ether phospholipids from scallop is shown below. One capsule contained 0.48 mg of ethanolamine plasmalogen and 0.02 mg of choline plasmalogen. Plasmalogen and placebo capsules were prepared by a manufacturer (B&S Corporation, Tokyo).

fcell-10-894734-t001

There were no between-group differences in physical and laboratory measurements. It is suggested that orally administered plasmalogens alleviate negative mood states and sleep problems, and also enhance mental concentration.”

https://www.frontiersin.org/articles/10.3389/fcell.2022.894734/full “Orally Administered Plasmalogens Alleviate Negative Mood States and Enhance Mental Concentration: A Randomized, Double-Blind, Placebo-Controlled Trial”

There was no dose / response investigation, so there’s no data to corroborate that this 2 mg treatment produced these effects. It isn’t difficult to think of other factors that could influence the primary outcome of a 18-22 year-old unmarried male’s moods.

2020

“Effects of ascidian-derived plasmalogens on cognitive performance improvement were assessed in a randomized, double-blind, placebo-controlled study including Japanese adult volunteers age 45.6 ± 11.1 years with mild forgetfulness. An allocation controller who was not directly involved in the study equally, but randomly, assigned participants to either the intervention group (n＝33) or the placebo group (n＝33), based on normalized Cognitrax composite memory score (the primary outcome), sex, and age at time of screen. Participants were administered either one active capsule (200 mg medium-chain triglyceride (MCT) oil including ascidian plasmalogen oil) or placebo capsule (200 mg MCT oil) per day with water, any time during the day for 12 weeks.

Ascidian plasmalogen oil was extracted from ascidians (Halocynthia roretzi) and sold by NIHON PHARMACEUTICAL CO., LTD. Based on a previous study, 33％ of lipids contained in ascidians are phospholipids, 23％ of which are plasmalogens, and fatty acids of the sn-2 position of plasmalogens are mainly EPA, DHA, oleic acid, and arachidonic acid. The active capsule contains 1 mg plasmalogen.

Compared to the placebo group, the intervention group showed a significant increase score in composite memory (eight weeks: 3.0 ± 16.3 points, 12 weeks: 6.7 ± 17.5 points), which was defined as the sum of verbal and visual memory scores. These results indicate consumption of ascidian-derived plasmalogen maintains and enhances memory function.”

https://www.jstage.jst.go.jp/article/jos/69/12/69_ess20167/_article “The Impact of Ascidian (Halocynthia roretzi)-derived Plasmalogen on Cognitive Function in Healthy Humans: A Randomized, Double-blind, Placebo-controlled Trial”

Again no dose / response investigation, so no corroborating data. Standard deviations many times larger than a sample’s mean indicated wild variability (aka noise). Maybe intervention participants experienced memory loss (3.0 mean – 16.3 SD = -13.3; 6.7 mean – 17.5 SD = -10.8)? Yet statistics inferred a signal that allowed interpreting this treatment as producing meaningful positive changes in cognitive function.

“Ten Parkinson’s disease (PD) patients age 67.80 (7.41) years received oral administration of 1 mg/day of purified ether phospholipids derived from scallop for 24 weeks. Clinical symptoms and blood tests were checked at 0, 4, 12, 24, and 28 weeks. Blood levels of plasmalogens in patients with PD were compared with those of 39 age-matched normal controls.

B&S Corporation Co. Ltd. (Tokyo) was involved in provision of capsules containing ether phospholipids derived from scallop. Ethanolamine ether phospholipids (ePE) in plasma from PD and relative composition of ethanolamine plasmalogen (plsPE) of erythrocyte membrane in PD were significantly low as compared to those of age-matched normal controls.

Oral administration of purified ether phospholipids derived from scallop for 24 weeks increased plasma ePE and erythrocyte plsPE to almost normal levels, and concomitantly improved some clinical symptoms of patients with PD. Results indicate the efficacy of oral administration of purified ether phospholipids derived from scallop to some nonmotor symptoms of PD. Physiological mechanisms of the efficacy of purified ether phospholipid derived from scallop remained to be elucidated.”

https://www.hindawi.com/journals/pd/2020/2671070/ “Improvement of Blood Plasmalogens and Clinical Symptoms in Parkinson’s Disease by Oral Administration of Ether Phospholipids: A Preliminary Report

Again no dose / response investigation, so no corroborating data. These researchers asserted their 2017 study to be a plasmalogen gold standard, as did the other two above studies.

Here’s part of what Dr. Goodenowe said about that 2017 study in a 2019 review Plasmalogen deficiency and neuropathology in Alzheimer’s disease: Causation or coincidence?:

“They did not observe a significant elevation of plasma levels of plasmalogens in the treated group relative to the baseline. Lower dose of plasmalogens (1 mg twice daily) and the labile nature of the vinyl-ether bond might have limited absorption of the intact molecule and might have contributed to the lack of response in terms of plasmalogen levels in blood as well as the cognitive function. Reported instability of plasmalogens in acidic environments questions the stability of preformed plasmalogens in gastric juice during digestion which might reduce plasmalogen bioavailability.”

Also see Part 1’s explanation of why using age-matched controls in plasmalogen studies is ridiculous.

Continued in Part 3.

Plasmalogens, Part 1

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The person who knows the most about this subject is Dayan Goodenowe, PhD. Some recent publications include:

https://www.frontiersin.org/articles/10.3389/fcell.2022.864842/full “Targeted Plasmalogen Supplementation: Effects on Blood Plasmalogens, Oxidative Stress Biomarkers, Cognition, and Mobility in Cognitively Impaired Persons”

https://www.frontiersin.org/articles/10.3389/fcell.2022.866156/full “Brain ethanolamine phospholipids, neuropathology and cognition: A comparative post-mortem analysis of structurally specific plasmalogen and phosphatidyl species”

plasmalogens and cognition

A sample of links freely available at https://drgoodenowe.com/.

1. Presentations to professional groups. Have your mouse ready to click the pause button.

https://drgoodenowe.com/dr-goodenowe-presents-at-the-iagg2023-in-yokohama-japan/ “A rare children’s disease that may be the key to reversing neurological decline in aging”

Includes videos of a treatment’s effects on a child.

https://neomarkgroup.wistia.com/medias/0qln0wy93t “The most influential biomarkers for aging and disease”

Despite the title, a considerable number of studies were presented on prenatal, infant, and early childhood development. He misspoke a few times, so read the slides.

“Phenotype is reality. Genotype is possibility. Communications links between different fields are very poorly connected in science.

Peroxisomes are islands. They don’t have DNA like your mitochondria do. Peroxisomal transport issues are important things to understand.

All aging-related cross-sectional analyses are on the rate of decline. You’re declining from a previous well state. Age-matched controls are the most ridiculous thing to do.”

2. I’ll highlight the longest of several interviews because there was plenty of room to expand on points. Maybe the best detailed explanations came as responses to that interviewer challenging with contrasting AD, traumatic brain injury, and cholesterol paradigms. Its transcript is more accurate than a usual YouTube interpretation, but there are still mistakes such as “fossil lipid” vs. phospholipid.

https://www.betterhealthguy.com/episode186 “Plasmalogens with Dr. Dayan Goodenowe, PhD”

“Science is how do you push things to its failure, until you can’t fail it again. We’ve lost that. It’s become more hypothesis proving.

Plasmalogens levels go up for a different reason than people think. The reason why it peaks in our 40s and 50s is because we’ve been myelinating. The white matter of our brain is still increasing. It’s not because we’re making more plasmalogens. It’s because the lake, the reservoir, gets full. What you’re measuring in blood is overflow from the lake. The lower plasmalogens start trickling down in your blood, the bigger drain that’s occurring on that system.

Low plasmalogens don’t just predict dementia in the elderly population. It predicts the rate of decline of that dementia. It predicts the rate of death.

The biggest drivers of plasmalogen manufacturing and the biggest reasons why they decrease with age, or in other circumstances is two things. One, the failure to maintain a fasting state of the human body. The second one is muscle atrophy.

Amyloid has absolutely nothing to do with Alzheimer’s, or dementia. It’s just a bystander on the road watching an accident happen.

Age-related cognitive decline is clearly where plasmalogens have the greatest impact. You’re always going to have mixed pathologies in the brain.

Nutritional availability of plasmalogens is virtually non-existent. As soon as they hit the hydrochloric acid of your stomach, they’re gone. They don’t make it past the stomach, or the upper intestine.”

I came across Dr. Goodenowe’s work last month from clicking a comment on this blog that linked back to her blog. Always be curious.

Continued in Part 2.

Bridging Nrf2 and autophagy

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Three more 2023 papers that cited Precondition your defenses with broccoli sprouts, starting with a review:

“Examining crosstalk between Nrf2 antioxidant signaling and autophagy provides insights into how they are interconnected and proteins that mediate their communication. These factors are potential therapeutic targets for diseases with both autophagy dysfunction and oxidative stress.

A working model illustrates mechanisms of bridging factors (SQSTM1, TFEB, Sestrin2, TRIM16, Ca2+, and miRNAs) connecting autophagy (left) and the main antioxidant Nrf2-Keap1-ARE pathway (right) and feedback loops between these factors.

fcell-11-1232241-g003

  • A network forms that connects Nrf2, SQSTM1, TFEB, and mTOR.
  • Other non-canonical autophagy regulatory proteins like Sestrin2 and tripartite motif-containing protein 16 (TRIM16) also participate in regulation of Nrf2 and mTOR via direct or indirect interactions.
  • Ca2+ is the most widespread intracellular messenger whose role in autophagy has been studied extensively.
  • At post-transcriptional level, microRNAs have been reported to impact both the regulation of autophagy and Nrf2 antioxidant signaling.

Since these regulatory proteins seem intricately entangled, potential side effects in practical scenarios should also be taken into consideration. Further studies on understanding the complex crosstalk between autophagy and antioxidant pathways are yet to be conducted.”

https://www.frontiersin.org/articles/10.3389/fcell.2023.1232241/full “An update on the bridging factors connecting autophagy and Nrf2 antioxidant pathway”

A second review subject was improving autophagy:

“Lysosomes are crucial degradative organelles that maintain cellular homeostasis. During the pathogenesis of neurodegenerative diseases and aging, functions of lysosomes are impaired, and lysosomal degradative capacity is consequently reduced.

Transcription factor EB-mediated lysosome biogenesis enhances autolysosome-dependent degradation, which subsequently alleviates neurodegenerative diseases. Small-molecule compounds that enhance TFEB activity and lysosome biogenesis are potential therapeutic agents.”

https://journals.lww.com/nrronline/fulltext/2023/11000/enhancement_of_lysosome_biogenesis_as_a_potential.7.aspx “Enhancement of lysosome biogenesis as a potential therapeutic approach for neurodegenerative diseases”

A third review tied mitochondrial participation into these processes:

“Mitochondria play an essential role in neural function, such as supporting normal energy metabolism, regulating reactive oxygen species, buffering physiological calcium loads, and maintaining the balance of morphology, subcellular distribution, and overall health through mitochondrial dynamics. Given recent technological advances in the assessment of mitochondrial structure and functions, mitochondrial dysfunction has been regarded as the early and key pathophysiological mechanism of cognitive disorders.

Mitochondrial dysfunction caused by acute and chronic brain injury is difficult to be distinguished because they may exhibit similar structural and functional impairments. Mitochondrial physiological function and morphology are integral, so when one is damaged, the other is also involved.

We recommend that all of the above methods can be used to explore mitochondrial dysfunction in different pathological pathways of cognitive disorders. Results may be related to special pathological pathways, sensitivity of the method, experiment cost, and degree of proficiency.”

https://journals.lww.com/nrronline/fulltext/2024/04000/latest_assessment_methods_for_mitochondrial.18.aspx “Latest assessment methods for mitochondrial homeostasis in cognitive diseases”

PXL_20231003_110600182

Ergothioneine dosing

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Four 2023 papers that outlined or used different ergothioneine doses, starting with a human/rodent study:

“We found that cognitive function and hippocampal neurogenesis were lower in mice fed an ERGO-free diet than in those fed the control diet. Mice fed an ERGO-free diet were orally administered ERGO (0, 2, and 20 mg/kg) for two weeks which reversed these effects.

trkb ratio

Phosphorylated brain-derived neurotrophic factor receptor TrkB, the activated form of TrkB, was also detected in extracellular vesicles (EVs) derived from serum samples of 52 volunteers who had been orally administered ERGO-containing tablets (5 mg/day for 12 weeks). The ratio of serum EV-derived phosphorylated TrkB was significantly higher in the ERGO-treated group than in the placebo-treated group and was positively correlated with both serum ERGO concentrations and several cognitive domain scores from Cognitrax.

cognitrax

The ratio of p-TrkB to TrkB in serum EVs was proposed as a quantitative diagnostic marker of long-term ERGO-induced cognitive improvement.”

https://www.researchsquare.com/article/rs-2626422/v1 “TrkB phosphorylation in serum extracellular vesicles correlates with cognitive function enhanced by ergothioneine in humans”

Human equivalents of all rodent ergothioneine doses were higher than the 5 mg/day for 12 weeks 2020 human study, cited as Reference 21. I couldn’t access that paper, so here’s its Abstract:

Effect of ergothioneine on the cognitive function improvement in healthy volunteers and mild cognitive impairment subjects – a randomized, double-blind, parallel-group comparison study

“These results indicate that continuous intake of ergothioneine improves cognitive function in healthy subjects.”

A rodent study compared effects of a fermented product with 0.1 and 1.0 mg/g (human equivalent 6 mg (1 mg x .081) x  70 kg) ergothioneine doses:

“Our present study demonstrated for the first time the preventive effect of Rice-koji fermented extracts made by Aspergillus oryzae on anxiety, impaired recognition, and nociception using a psychophysically stressed model. Our results also demonstrated preventive effects of ergothioneine (EGT) on stress-induced anxiety- and pain-like behaviors.

Daily administration of High dose Rice-koji or 0.1 mg/kg EGT decreased anxiety- and pain-like behaviors. These findings suggest that inhibitory effects of Rice-koji on psychological stress might be mediated through the actions of EGT.”

https://www.mdpi.com/2072-6643/15/18/3989 “Preventive Roles of Rice-koji Extracts and Ergothioneine on Anxiety- and Pain-like Responses under Psychophysical Stress Conditions in Male Mice”

Here’s one of several reviews that cited a 2017 clinical trial (duplicately Reference 39 and 61 for some reason) of 5 and 25 mg ergothioneine doses:

“In this pharmacokinetic study, forty-five healthy humans received placebo, 5, or 25 mg encapsulated ergothioneine/d for 7 d and were followed up for an additional 4 weeks. Ergothioneine was rapidly absorbed and largely retained by the body, with large increases in plasma ergothioneine levels and only minimal increases (<4 %) in urinary excretion observed. While plasma levels of ergothioneine decreased when supplementation was withdrawn, levels in whole blood continued to increase in a dose–response fashion, reaching maximal levels 3 weeks after withdrawal of supplement, which were sustained at 4 weeks follow-up.

A large difference in basal concentrations of ergothioneine in whole blood was observed. Participants with the highest basal levels of ergothioneine also appeared to take up more of supplemented ergothioneine.”

https://www.cambridge.org/core/journals/british-journal-of-nutrition/article/ergothioneine-an-underrecognised-dietary-micronutrient-required-for-healthy-ageing/92CED7FF201A9FB23BEAFF0D3EAD7316 “Ergothioneine: an underrecognised dietary micronutrient required for healthy ageing?”

Wrapping up with a deep dive into seven mushroom varieties’ compounds:

“Mushrooms contain multiple essential nutrients and health-promoting bioactive compounds, including amino acid L-ergothioneine. We compared metabolomes of fresh raw white button, crimini, portabella, lion’s mane, maitake, oyster, and shiitake mushrooms using untargeted liquid chromatography mass spectrometry (LC/MS)-based metabolomics.

Results indicate significantly higher concentrations of L-ergothioneine in lion’s mane and oyster mushrooms compared to the remaining five mushroom varieties, which had concentrations ranging from 1.94 ± 0.55 to 5.26 ± 1.23 mg/100 g wet weight (mean ± SD). There was also variability in concentration of L-ergothioneine between mushroom varieties of the same farm. Different numbers denote significance (p < 0.05).

foods-12-02985-g008

Mushrooms and their bioactive extracts are considered functional foods. Mushrooms have several bioactive compounds, including polysaccharides, lectins, terpenoids, sterols, and alkaloids, among others, which may positively impact health.

Cell walls of mushrooms contain polysaccharides, including β-glucans and chitin, which positively affect health, through modulating the immune system and protecting the cardiovascular system through improvements in glucose and lipid metabolism. Effects on the cardiovascular system are also attributable to lovastatin and polyphenols, known for their lipid-lowering and antioxidant properties, respectively.

While the 1344 compounds in common among the seven mushroom varieties support some level of similarity, detection of hundreds of unique-to-mushroom-variety compounds and differences in amino acid profiles indicate that not all mushrooms are chemically comparable. Given detection of >400 unique-to-mushroom-variety compounds in lion’s mane, maitake, oyster, and shiitake mushrooms, we suggest further targeted investigations on compounds detected and potential health benefits.”

https://www.mdpi.com/2304-8158/12/16/2985 “Metabolomics Profiling of White Button, Crimini, Portabella, Lion’s Mane, Maitake, Oyster, and Shiitake Mushrooms Using Untargeted Metabolomics and Targeted Amino Acid Analysis”

I eat around 200 grams of mushrooms daily, having temporarily overridden the boredom of eating AGE-less chicken vegetable soup every day. I prep all the top package’s frozen umami bomb (283 grams) and half of the bottom’s fresh mushrooms (340 grams) into the soup:

PXL_20230921_193708552

It makes servings for three days, including one for prep day dinner. I’d guess from “concentrations ranging from 1.94 ± 0.55 to 5.26 ± 1.23 mg/100 g (mean ± SD)” that my daily mushroom ergothioneine dose is around 7 mg ((1.94 mg + 5.26 mg) / 2) = 3.6 mg per 100 grams x 2 (for 200 grams).

Continued in Part 2.

Nrf2 and senescence, Part 1

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A 2023 rodent study investigated Nrf2’s capacity to cause cell senescence:

“The KEAP1-NRF2 pathway is a stress response pathway which has been maintained by natural selection due to its ability to benefit survival of the host organism. One important distinction between this pathway and other stress response pathways such as p53, is that chronic activation of NRF2 has not been associated directly with a mechanism to promote cell death if survival of the cell becomes deleterious to the host.

Some unexplained observations suggest that NRF2 activation has additional physiological outputs which have yet to be described. For example, despite the fact that oxidative stress plays an important role in etiology of many aging-related diseases, genetic activation of Nrf2 in mice is associated with decreased lifespan.

We found that NRF2 functions to prime cells to become senescent in response to irreparable damage. In diseased states, NRF2 promotes transcriptional activation of a specific subset of the senescence-associated secretory phenotype (SASP) gene program, which we have named the NRF2-induced secretory phenotype (NISP).

full vs nisp sasp

As Nrf2 also promotes monocyte and macrophage invasion in mouse disease models of steatohepatitis, colitis, pancreatitis, and autoimmune nephritis, we would posit that it represents a central component of the Nrf2 response in damaged epithelial tissues, and that the NRF2-NISP-Immune recruitment model represents a framework through which these disease phenotypes can be understood.

This pathway represents the final stage of the oxidative stress response, as it allows cells to be safely removed if macromolecular damage caused by the original stressor is so extensive that it is beyond the repair capacity of the cell.”

https://www.sciencedirect.com/science/article/pii/S221323172300246X “A NRF2-induced secretory phenotype activates immune surveillance to remove irreparably damaged cells”

Continued in Part 2.

Ripe wild grapes

PXL_20230905_110321298.MP

Fructose and survival

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This 2023 paper provided mechanistic evidence, evolutionary theory, and testable scenarios for fructose metabolism differences from other nutrients:

“The fructose survival hypothesis proposes that obesity and metabolic disorders may have developed from over-stimulation of an evolutionary-based biologic response (survival switch) that aims to protect animals in advance of crisis. The response is characterized by hunger, thirst, foraging, weight gain, fat accumulation, insulin resistance, systemic inflammation, and increased blood pressure.

Unlike other nutrients, fructose reduces the active energy (adenosine triphosphate) in the cell, while blocking its regeneration from fat stores. This is mediated by intracellular uric acid, mitochondrial oxidative stress, inhibition of AMP kinase, and stimulation of vasopressin.

rstb20220230f04

Fructose metabolism is associated with oxidative stress, mitochondrial dysfunction, loss of cytoprotective transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), and a reduction in sirtuins that characterize the ageing process. Fructose also induces generation of advanced glycation end products much more effectively than glucose.

The fructose pathway is almost inevitably strongest in early disease states, for over time there is often fibrosis, inflammation, or mitochondrial loss that results in persistence of the disease process. The best time for intervention may turn out to be in early disease before conditions become less reversible.”

https://royalsocietypublishing.org/doi/10.1098/rstb.2022.0230 “The fructose survival hypothesis for obesity”

Time to exit fructose survival mode.

PXL_20230904_140453607

Acetyl-L-carnitine dosing

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Haven’t curated acetyl-L-carnitine papers recently. Here are three 2023 studies, beginning with a human case report:

“It is believed that 75% of the required amount of carnitine is taken from diet and the remaining 25% is synthesized in the body. Long-term use of a carnitine-free diet is thought to increase the risk of carnitine deficiency.

Dosage for long-term tube-fed patients with disorders of consciousness and convulsive seizures, such as in the present cases, is not specified. Instructions accompanying the medication list gastrointestinal symptoms such as nausea, vomiting, and diarrhea as side effects of L-carnitine. They indicate a maximum dosage of 3 g/day, and a maximum single dose of 1 g.

L-Carnitine is efficiently absorbed in the gastrointestinal tract when taken in small amounts, but when taken in large amounts, the transporter is saturated and bioavailability is only about 10%–20%. Although safety of oral L-carnitine administration is considered high because there is an upper limit to the amount that can be absorbed, clinicians should remain aware of side effects noted above.

To the best of our knowledge, this is the first report in which L-carnitine was administered to a patient with impaired consciousness after stroke with the result that symptoms improved. It is possible that carnitine deficiency is overlooked in some patients in rehabilitation wards, and measurement of ammonia may be useful in its detection. Because carnitine deficiency might interfere with active rehabilitation, nutritional management with attention to carnitine deficiency is important in rehabilitation wards.”

https://www.jstage.jst.go.jp/article/prm/8/0/8_20230019/_html/-char/en “Disorders of Consciousness after Subacute Stroke Might Partly be Caused by Carnitine Deficiency: Two Case Reports”

I currently take one gram of acetyl-L-carnitine three times a day.

Next is a clinical trial with amyotrophic lateral sclerosis (ALS) patients that used two different doses of acetyl-L-carnitine:

“Our findings did not confirm an effect of ALCAR 3 g/day on survival in ALS subjects at 24 months. An effect was observed in those treated with ALCAR 1.5 g/day.

In addition, we did not detect an effect on self-sufficiency at 12 months as previously seen in the pilot trial. These differences could be explained by:

  • The study design (retrospective observational study vs prospective randomized trial);
  • Selection bias (subjects from the real-world clinical practice are less selected than those included in a clinical trial); and
  • Drug compliance (subjects enrolled in a clinical trial perform several onsite evaluations in which compliance is verified by tablets accounting, while in clinical practice this is not done).

Our hypothesis is that the presence of residual confounding might explain our unexpected results. Residual confounding refers to the presence of an unmeasured or uncontrolled variable that could affect the relationship between treatment (ALCAR) and outcome.

This study provided additional information on the potential effect of ALCAR on disease progression and survival, and adds evidence to justify the use of ALCAR in ALS subjects.”

https://link.springer.com/article/10.1007/s00415-023-11844-6 “Retrospective observational study on the use of acetyl-L-carnitine in ALS”

This study’s dosing method wasn’t clear on exactly how doses were administered every day. I’ll guess that if both 1.5 and 3 grams were given all at once, they might have been roughly equivalent doses per the first paper’s cited bioavailability saturation effect.

Next is a rodent aging study:

“The aim of this study was to examine effects of long-term L-Carnitine (β-hydroxy-γ-trimethylaminobutyric acid, LC) administration on cardiomyocyte contraction and intracellular Ca2+ transients in aging rats. LC (50 mg/kg body weight/day) was dissolved in distilled water and orally administered for a period of 7 months.

LC increased cardiomyocyte cell shortening and resting sarcomere length. LC supplementation led to a reduction in resting [Ca2+]i level and an increase in the amplitude of [Ca2+]i transients, indicative of enhanced contraction. Consistent with these results, decay time of Ca2+ transients also decreased significantly in the LC-treated group.

Long-term administration of LC may help restore Ca2+ homeostasis altered during aging, and could be used as a cardioprotective medication in cases where myocyte contractility is diminished.”

https://link.springer.com/article/10.1007/s00418-023-02215-3 “L-Carnitine improves mechanical responses of cardiomyocytes and restores Ca2+ homeostasis during aging” (not freely available)

A human equivalent of this study’s daily dose is (50 mg x .162) x 70 kg = 567 mg. A human equivalent of this study’s duration using the maximum lifespan method is (7 months x 32.2) = 225.4 months. The subjects began at 11 months old (human equivalent age 29.5 years) and ended at 18 months old (human equivalent age 48.3 years).

This study illustrated how heart dysfunctions with subclinical symptoms advance with aging, and that starting to do something preventative before human equivalent age 30 may work.

PXL_20230814_101039592

How long does sulforaphane keep?

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This 2023 clinical trial with broccoli sprout powder investigated ways of improving blood plasma measurements:

“Quantifying sulforaphane (SFN) and its thiol metabolites in biological samples using liquid chromatography-tandem mass spectrometry is complicated by SFN’s electrophilic nature and the facile dissociation of SFN-thiol conjugates. We used the alkylating agent iodoacetamide (IAA) to both release SFN from protein thiols and force the dissociation of SFN metabolites.

After 15 min of incubation, approximately 90% of the SFN was recovered. After 2 hours of incubation, SFN loss to thiol conjugation was significant, with approximately 30% recovered. The percentage of SFN lost in this manner in human plasma upon protein precipitation would likely be greater than the loss we observed in fetal bovine serum (FBS), as free thiols in human plasma are approximately 6 times higher than in FBS.

images_medium_jf3c01367_0002

Subjects were 12 healthy, young (19–30 y), men (n = 4), and women (n = 8). They consumed three EnduraCell capsules with water.

Per the manufacturer, each capsule contained 700 mg of 100% whole broccoli sprout powder, including active myrosinase and 21 mg of glucoraphanin, which upon full conversion to SFN would yield ∼8 mg, equaling ∼24 mg of SFN total per three-capsule dose. We note that full conversion to SFN, even with active myrosinase in the supplement, is not expected.

SFN concentrations in plasma increased for all subjects, with the highest being 193 nM for subject 4 one hour post consumption. Asterisks for the 2 and 3 hour time points indicate a p-value of <0.05, comparing that time point to the previous time point for that participant.

images_medium_jf3c01367_0005

This thiol-blocking method increased SFN percent recovery from FBS from 32 to 94 ± 5%. Applying the method to clinical samples, SFN concentrations were on average 6 times greater than when IAA was omitted.”

https://pubs.acs.org/doi/10.1021/acs.jafc.3c01367 “Bioavailable Sulforaphane Quantitation in Plasma by LC–MS/MS Is Enhanced by Blocking Thiols”

I didn’t highlight subject bioavailability statistics or researcher generalizations. Someday, researchers will be interested and forward-thinking enough about their field to plan ahead and investigate likely occurrences such as:

  • What caused Subjects 3 and 4 to have much higher responses than Subjects 2 and 6?
  • What caused Subjects 5 and 11 to have their highest sulforaphane levels at their 3-hour points, whereas 3-hour points were most other subjects’ lowest levels?

And, of course, what was the actual sulforaphane intake by researcher measurements vs. relying on manufacturer statements? Especially when researcher lab equipment was available to measure that.

This study’s proof of concept demonstrated a 10% sulforaphane loss after 15 minutes, and a 68% loss after two hours. Applying their finding to home sprouting, I wouldn’t expect any advance preparation method of broccoli sprouts or seeds to adequately address sulforaphane degradation.

I’d suggest microwaving up to 60°C (140°F) and/or grinding and/or blending broccoli sprouts and seeds just before eating to optimize your sulforaphane intake. Also, taking the very reactive sulforaphane by itself to optimize your results.

Take Vitamin K2 to protect against aluminum toxicity

gettingwell4 4-5 stars Advanced knowledge, Acetylcholine, Cerebrum, Dopamine, Epigenetics, Hippocampus, Limbic system, Memory, Neurochemicals, Stress , , ,

This 2023 rodent study investigated relationships of MK-7 menaquinone, aluminum trichloride, and brain health:

“A variety of endogenous and exogenous agents, such as metals and environmental toxins (aluminum, mercury, etc.), can contribute to neurodegeneration, which is of multifactorial clinical occurrence.

The current study showed that Alzheimer’s Disease (AD)-like condition was induced in mice by AlCl3 treatment affecting spatial and recognition memory. Neuropathological alterations included neuroinflammation, oxidative stress, an increase in brain amyloid β levels, and loss of hippocampal neurons.

Aluminium chloride (AlCl3; 100 mg/kg for 3 weeks orally) was administered to Swiss albino mice to induce neurodegeneration and Vitamin K2 (100 mcg/kg for 3 weeks orally) was applied as treatment. This was followed by behavioral studies to determine memory changes.

Antioxidants like glutathione and SOD were low compared to the control group, while oxidative stress marker MDA was elevated. BDNF levels increased in the Vitamin K2 treated animals, suggesting its neuroprotective functions.

k2 abstract

vitamin K2 BDNF

Vitamin K2 could partially reverse AlCl3-mediated cognitive decline. It increased hippocampal acetylcholine and BDNF levels while reducing oxidative stress, neuroinflammation, and β-amyloid deposition, protecting hippocampal neurons from AlCl3-mediated damage.“

https://link.springer.com/article/10.1007/s10787-023-01290-1 “Vitamin K2 protects against aluminium chloride-mediated neurodegeneration” (not freely available)

This study’s human equivalent Vitamin K2 dose is (100 mcg x .081) x 70 kg = 567 mcg. I’ve taken 600 mcg MK-7 every day for the past two years.

Found out last week that I’ve also been inadvertently dosing myself with aluminum every day. This is the underside of my former 3-year-old drip coffee maker with its cover removed:

PXL_20230813_172709641

I’m certain its aluminum tubing that heats reservoir water started to corrode a long time ago. Currently trying out methods of making aluminum-free coffee.

Reversing biological age in rats

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This 2023 rodent study wrapped together findings of the original study curated in A rejuvenation therapy and sulforaphane, and the second follow-on study mentioned in Signaling pathways and aging. I’ll start by highlighting specifics of the later study:

“Pronounced rejuvenation effects in male rats prompted us to conduct further confirmatory experiments. A particularly important consideration is the effectiveness of E5 with regards to sex, as sex-dependent rejuvenation by some interventions have previously been reported.

To assess E5’s applicability to both male and female Sprague Dawley rats, we studied 12 males (6 treated with E5, 6 with saline) and 12 females (6 treated with E5, 6 with saline). These rats were treated every 45 days with an injection of E5 or saline. Rats were monitored for 165 days, and blood was drawn at six time points: 0, 15, 30, 60, 150 and 165 days from the first injection.

We observed highly significant improvements in TNF alpha and IL-6 levels for both males and females in the blood of E5-injected rats over that of saline controls. We also observed a substantial improvement in grip strength.

Our study shows age reversal effects in both male and female rats, but E5 is more effective in males.”

Another experimental group was started with old rats of both sexes. Using the human / rat relative clock developed in the original study, a human equivalent age to these rats at 26 months old was ((112.7 weeks / 197.6 weeks maximum rat lifespan) x 122.5 years maximum human lifespan) = 69.8 years:

“To validate our epigenetic clock results, we conducted a second set of E5 experiments with Sprague Dawley rats of both sexes. When these rats turned 26 months old, half (9 rats) received the E5 treatment while the other half (8 rats) received only the control treatment (saline injection). We analyzed methylation data from two blood draws: blood draw before treatment (baseline) and a follow up sample (15 days after the E5/saline treatment).”

Treatment measurements were affected by one female control group outlier. Panels F through J were recalculated after removing the outlier to show significant effects in both sexes:

second follow-on results

“A) Final version of the rat clock for blood. Baseline measurement (x-axis) versus follow up measurement (15 days after treatment, y-axis). Points (rats) are colored by treatment: red=treated by E5, black=treated with saline only. Rotated grey numbers underneath each bar reports the group sizes. Each bar plot reports the mean value and one standard error.

B,D,E) Difference between follow up measurement and baseline measurement (y-axis) versus treatment status in B) all rats, D) female rats only, E) male rats only. C) is analogous to B) but uses the pan tissue clock for rats.

Panels in the second row (F,G,H,I,J) are analogous to those in the first row but the analysis omitted one control rat (corresponding to the black dot in the lower right of panel A).”

https://www.biorxiv.org/content/10.1101/2023.08.06.552148v1 “Reversal of Biological Age in Multiple Rat Organs by Young Porcine Plasma Fraction”

A description of how E5 plasma fraction was made starts on page 16 of the *.pdf file. The next E5 study will be done with dogs per July 2023 updates in blog post comments:

“On E5 our entire team is working hard towards the launch of an old Beagle dogs trial this month. We want to make them really young, healthy, happy, and jumping around like 1 and 2 year olds.

Primary endpoint is safety and toxicology to test various dose strengths and frequencies. Secondary endpoints are more than 20.

As you know, we like to test exhaustively to get a sharper perspective of what’s happening. In rat studies we tested 30 biomarkers, including functional. We are especially keen to check kidney markers.

There are two clocks for dogs we are interested in to get third party confirmation of age reversal. Horvath dog clock is ready and GlycanAge dog clock is under construction.

We are requesting all organizations that support pets and aging to financially support their project of building an accurate dog clock. Not only will it help veterinary aging research like ours, but also all the dog owners that may want to know how much improvement their dog received from treatment. Dr. Matt Kaeberlain is an advisor on their project.”

36 holes in your roof

gettingwell4 4-5 stars Advanced knowledge, Cerebrum, Cortisol, Hippocampus, Immune system, Limbic system, Memory, Neurochemicals, Stress, Telomeres, Testosterone , , , , ,

An August 2023 interview with Dr. Dale Bredesen, who has reversed Alheizmer’s disease in many people, which will never be acknowledged by the corrupt paradigm:

“How much do you want me to go into things that are relatively controversial and how much do you want me to stick with kind of the more standard line?

For Alzheimer’s we noticed initially there are 36 different potential contributors. You need to patch as many as possible to have an effect.

All of these things, your estradiol level, your progesterone level, pregnenolone, free T3, TSH, Vitamin D, testosterone, these things are all critical. They all feed into the equation.

You have over a hundred trillion contacts in your brain. Will you be able to keep them? Or do you not have what it takes to keep them, and you have to downsize?

The reality is Alzheimer’s disease should be a rare disease. If everybody would get on appropriate prevention or early reversal, we could make it a rare disease.”

https://brokenscience.org/podcasts-ep-5/ “Dale Bredesen – Reversing Alzheimer’s Fate”

See A therapy to reverse cognitive decline for previous curation of Dr. Bredesen’s work.

A blood plasma aging clock, Part 2

gettingwell4 2-3 stars Required further work, Epigenetics, Hippocampus, Immune system, Limbic system, Stress, Testosterone , ,

Quite a few people recently looked at Part 1 which curated “Undulating changes in human plasma proteome across lifespan are linked to disease” in December 2019. Let’s start with a 2023 human study coauthored by Part 1’s lead researcher:

“The aim of this study is to identify a set of proteins in human plasma associated with aging by integration of data of four independent, large-scaled datasets. We identified a set of 273 plasma proteins significantly associated with aging (aging proteins, APs) across these cohorts consisting of healthy individuals and individuals with comorbidities and highlight their biological functions.

arthur and robbins cohorts

Although these presented proteins may be different compared to other presented proteomic clocks [like Part 1’s], this can be explained due to a variety of factors. Across studies there may be several technical factors, such as used anti-coagulants, and biological differences, such as different age ranges, ethnicity and corrections for BMI, which may influence the plasma proteome in the cohorts. To overcome these differences, we focused on the overlap between the different studies as they also present several of these confounding factors.

We show that individuals presenting accelerated or decelerated aging based on their plasma proteome, respectively have a more aged or younger systemic environment. These results provide novel insights in understanding the aging process and its underlying mechanisms and highlight potential modulators contributing to healthy aging.”

https://www.frontiersin.org/articles/10.3389/fragi.2023.1112109/full “Markers of aging: Unsupervised integrated analyses of the human plasma proteome”

A 2023 human study cited the above study and found:

“Our cross-sectional study of adults adherent and non-adherent to recommended lifestyle habits established strong group differences for 39 proteins primarily related to innate immunity and lipoprotein metabolism. Many of these protein differences were best explained by group contrasts in adiposity and visceral fat. The relatively small number of upregulated and downregulated proteins associated with good lifestyle habits should facilitate development of a targeted lifestyle proteomic panel that can be used in future studies to determine efficacy of various prevention and treatment strategies.”

https://www.researchsquare.com/article/rs-3097901/v1 “Adherence to Lifestyle Recommendations Linked to Innate Immunity and Lipoprotein Metabolism: A Cross-Sectional Comparison Using Untargeted Proteomics”

A 2023 human study from Google-owned Calico:

“In most cases, direction of effects between cause-specific and all-cause mortality was concordant, but all-cause mortality association was not statistically significant. Neither do we have insight into conditional causal effects of these proteins nor interaction effects between them.”

https://www.researchsquare.com/article/rs-2626017/v1 “Plasma Proteomic Determinants of Common Causes of Mortality”

“Undulating” in Part 1 described plasma proteins changing over time with peaks at ages 34, 60, and 78. Those peaks don’t provide a base for linearly extrapolating all-cause mortality.

peaks

A 2023 rodent study did a touch better with one of Part 1’s 46 proteins of a conserved aging signature that changed in the same direction with mice and humans, although it didn’t fully investigate protein expression over time.

“Interactions between CHRDL1 levels, age, and plasma lipids that might affect cardiometabolic health should be further investigated.”

https://www.mdpi.com/2073-4409/12/4/624 “Chordin-like 1, a Novel Adipokine, Markedly Promotes Adipogenesis and Lipid Accumulation”

Sulforaphane, TFEB, and ADH1

gettingwell4 4-5 stars Advanced knowledge, Epigenetics, Hippocampus, Immune system, Limbic system, Memory, Stress , , , ,

Looked for a recent follow-on study of the 2021 Precondition your defenses with broccoli sprouts, specifically:

“NFE2L2/NRF2 is a target gene of TFEB (transcription factor EB), a master regulator of autophagic and lysosomal functions, which we show here to be potently activated by sulforaphane.”

Some interesting papers cited it, but no studies continued its sulforaphane/TFEB line of inquiry. A 2022 review made a good point when citing this study for TFEB, but didn’t tie it in with sulforaphane:

“TFEB is translocated into the nucleus with a moderate increase of ROS through a Ca2+-dependent, but mTOR (mechanistic target of rapamycin kinase)-independent mechanism. Essential genes involved in lysosome biogenesis and autophagosome are activated, which are crucial for removal of damaged mitochondria.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9730074/ “Phytochemicals and modulation of exercise-induced oxidative stress: a novel overview of antioxidants”

A search of TFEB brought up a 2023 nematode study:

“We searched for effectors acting downstream of the transcription factor EB (TFEB), known as HLH-30 in C. elegans, because TFEB/HLH-30 is necessary across anti-aging interventions. Its overexpression is sufficient to extend C. elegans lifespan, and reduce biomarkers of aging in mammals including humans.

While investigating the potential role of autophagy in hlh-30 dependent longevity of the mxl-3 C. elegans mutant, we found that the current model has exceptions. Contrary to expectation, we found that autophagy is not activated in the mxl-3 mutant, and that neither autophagy nor lysosomal activity are required for the longevity phenotype observed in these mutant animals. mxl-3 longevity is hlh-30-dependent but autophagy-independent.

Instead, we found the gene encoding Alcohol DeHydrogenase ADH-1 induced in mxl-3 and other hlh-30-dependent anti-aging interventions. adh-1 is induced in an hlh-30-dependent manner in longevity models caloric restriction (eat-2), insulin insensitivity (daf-2), and mTOR inhibition (let-363 RNAi).

insulin insensitivity longevity model

We present an alcohol-dehydrogenase-mediated anti-aging response (AMAR) that is essential for C. elegans longevity driven by HLH-30 overexpression, caloric restriction, mTOR inhibition, and insulin-signaling deficiency. Overexpression of ADH-1 is sufficient to activate AMAR, which extends healthspan and lifespan by reducing levels of glycerol, an age-associated and aging-promoting alcohol.”

https://www.cell.com/current-biology/fulltext/S0960-9822(23)00128-8 “Increased alcohol dehydrogenase 1 activity promotes longevity” (not freely available) Thanks to Dr. Eyleen O’Rourke for providing a copy.

A 2022 human study found that chronic ADH1 activation occurs in liver disease:

“Activity of total ADH, ADH isoenzymes and aldehyde dehydrogenase (ALDH) was evaluated in the blood serum of patients with primary biliary cholangitis (PBC), a chronic autoimmune disease of the liver. An increase in class I ADH and total ADH activity indicates that the isoenzyme class I ADH is released by compromised liver cells and can be useful diagnostic markers of PBC.”

https://link.springer.com/article/10.1007/s00005-022-00667-4 “An Assessment of the Serum Activity of ADH and ALDH in Patients with Primary Biliary Cholangitis”

Chronically activating any of the body’s systems points to a problem. There’s has to be a balance.

A 2022 rodent study investigated ADH1 activation and MEK1/2 inhibitors for beneficial effects:

“Alcohol is mainly catabolized by class I alcohol dehydrogenase (ADH1) in liver. ADH deficiency can aggravate ethanol-induced tissue injury.

Extracellular signal-regulated kinases 1/2 (ERK1/2) is involved in alcohol metabolism. However, the relationship between ERK1/2 and ADH1 remains unclear.

Mitogen-activated protein kinases 1/2 (MEK1/2) is required to phosphorylate and activate ERK1/2. Protein expression of phosphorylated ERK1/2 in liver is inversely associated with ethanol-induced liver injury and hepatocytes apoptosis, suggesting inhibition of ERK1/2 may protect hepatocytes from ethanol-induced cytotoxicity. We hypothesize that inhibition of ERK1/2 by MEK1/2 inhibitors may protect hepatocytes from ethanol cytotoxicity by activating ADH1 expression.

Results showed MEK1/2 inhibitors significantly increased ADH1 protein expression by inducing its transcription activity. Our findings revealed inhibition of ERK1/2 can significantly increase ADH1 expression, indicating MEK1/2 inhibitors may possess potential application in alcohol-related diseases.”

https://link.springer.com/article/10.1007/s11033-022-07361-w “MEK1/2 inhibitors induce class I alcohol dehydrogenase (ADH1) expression by regulating farnesoid X receptor in hepatic cell lines and C57BL/6J mouse” (not freely available)

Chronically inhibiting any of the body’s systems also points to a problem.

A 2022 rodent study investigated TFEB activation and MEK1/2 inhibitors for beneficial effects:

“Inhibiting MEK/ERK signaling using a clinically available MEK1/2 inhibitor induces protection of neurons through autophagic lysosomal activation mediated by transcription factor EB (TFEB) in a model of AD.”

https://www.nature.com/articles/s41380-022-01713-5 “MEK1/2 inhibition rescues neurodegeneration by TFEB-mediated activation of autophagic lysosomal function in a model of Alzheimer’s Disease”

PXL_20230720_102047833

Natural ways to modify GDF11

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Three 2023 studies to follow up mention of GDF11 in the Brain endothelial cells post. Two are selected for non-pharmaceutical interventions people can do on their own. Let’s start with a human exercise study:

“We explored the exercise-related regulation of Growth Differentiation Factor 11 (GDF11) in cerebrospinal fluid (CSF) and blood. Samples of serum, plasma, and CSF were obtained before and 60 min after acute exercise (90 min run) from twenty healthy young individuals. Additional serum and plasma samples were collected immediately after run. GDF11 protein content, body composition, physical fitness, and cognitive functions were evaluated.

Controversies regarding the role of GDF11 in aging originate mainly from the absence of a reliable, validated and widely accepted method of GDF11 detection. To support the reliability of our findings as well as to distinguish GDF11 from its close homologue GDF8, we determined GDF11 in CSF, serum, and plasma, by immunoblotting, using two different GDF11-specific antibodies, as well as GDF11/GDF8 non-specific antibody. These antibodies have been previously successfully used by others. Reliability of our findings is further supported by correlations between GDF11 in serum and plasma, as well as between GDF11 and serum GDF11/GDF8.

We report an association between levels of GDF11 and adiponectin in CSF as well as in serum after acute endurance exercise. These observations support potentially synergic effects of GDF11 and adiponectin on the brain. The experimental design we implement seems to represent a reliable model to study regulation of bioactive molecules, potential mediators of neuroprotective effects of exercise in the human brain.

We show for the first time a direct link between endurance exercise and GDF11 levels in human cerebrospinal fluid. This study provided the first albeit indirect (correlative) evidence on the putative role of GDF11 in promoting healthy aging in humans, by demonstrating a tight relationship between serum GDF11 and peak power output. We extend this observation by showing that the level of physical fitness is an important determinant of regulation of GDF11 by acute exercise.

In this work, we confirm in a bigger cohort our previous finding that blood-brain barrier permeability, as assessed by CSF/serum albumin ratio, is decreased after an acute bout of endurance exercise. We observed a modest positive correlation between CSF/serum albumin ratio and CSF/serum GDF11/GDF8 ratio, with a trend also for GDF11. However, exercise-induced changes of CSF/serum albumin ratio and that of GDF11 or GDF11/GDF8 did not correlate, indicating that there are other factors that could modulate levels of this growth factor rather than blood-brain barrier permeability.”

https://www.frontiersin.org/articles/10.3389/fendo.2023.1137048/full “Acute endurance exercise modulates growth differentiation factor 11 in cerebrospinal fluid of healthy young adults”

Next is a rodent study of intermittent fasting before and after cerebral ischemia:

“The present study focused on the cerebral angiogenesis effect of intermittent fasting (IF) on ischemic rats. Rats were fed within strict time periods for 8 h out of every 24 h, with free access to food between 0800 and 1600 h.

In the first step, we designed different time schedules (10 d, 1 month, and 3 months) of IF before middle cerebral artery occlusion (MCAO). We monitored whether IF accelerated neurobehavioral recovery and induced expression of endothelial cells after MCAO. Then we explored whether GDF11 and downstream signals mediated angiogenesis in the peri-infarct area.

journal.pone.0282338.g006

We found that 3 months (p < 0.01) and 1 month (p < 0.05) of IF conditioning, respectively, markedly increased GDF11-positive cells in the peri-infarct area 3 d after MCAO compared with ad libitum dietary regimen. There were no significant differences between the cerebral ischemia (CI) + ad libitum group and the CI + IF 10-day group.

We also assayed plasma expression pattern of GDF11 protein. Plasma level of GDF11 protein was significantly upregulated in the IF dietary groups compared with the ad libitum dietary group 3 d after MCAO, which was consistent with the brain level. However, short-term CI + IF 10-day group results were not statistically different from CI + ad libitum group.

Taken together, our results strongly indicated that pretreatment of long-term IF might promote circulation of GDF11 and cerebral GDF11 protein during the post-ischemic, recovery period. Preoperative long-term IF might be beneficial for inducing cerebral angiogenesis in acute cerebral infarction.

These findings suggested that the longer the period of IF before MCAO, the better the protective effects after surgery.”

https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0282338 “Long-term intermittent fasting improves neurological function by promoting angiogenesis after cerebral ischemia via growth differentiation factor 11 signaling activation”

Per Week 28 of Changing to a youthful phenotype with broccoli sprouts, using species maximum lifespan to estimate a human-equivalent multiplication factor that can be applied to a rat post-development time period is 122.5 years / 3.8 years = 32.2. Applying it to this study’s findings:

  • 10 rat days (322 human days) of intermittent fasting provided little protection from cerebral ischemia;
  • 1 rat month (32.2 human months) of intermittent fasting had better protection; and
  • 3 rat months (a little over 8 human years) of intermittent fasting had even stronger protection.

Is it worth the hassle of intermittently fasting every day for years to prevent a future stroke, or better recover from one, or keep other subclinical / clinical diseases from accelerating, or keep aging from accelerating? This study also cited more immediate benefits of intermittent fasting.

Might be too late for a gradual approach for people who are already diseased or close, though, like subjects in this human study:

“We aimed to explore the correlation among serum GDF11, the severity of coronary artery lesions, and the prognosis of patients with ST-segment elevation myocardial infarction (STEMI). A total of 367 patients were enrolled and divided into control (n = 172) and STEMI (n = 195) groups. Control group fulfilled the following criteria:

  1. Presented with typical chest tightness, chest pain, or other discomfort symptoms on admission;
  2. Electrocardiogram examination suggested ST-T changes;
  3. Levels of myocardial injury markers did not suggest abnormalities; and
  4. The diagnosis of unstable angina was considered clinically valid.

14 variables that were significant in univariate logistic regression analysis were included in the subsequent multivariate logistic regression analysis. Multivariate analysis indicated that smoking, diabetes, C-reactive protein, homocysteine, and lipoprotein (a) were positively correlated with STEMI occurrence, whereas serum GDF11 and the Apolipoprotein A1-to-Apolipoprotein B ratio were negatively correlated with STEMI occurrence.

Serum GDF11 was negatively correlated with severity of coronary lesions, and was also an independent prognostic indicator of major adverse cardiovascular events in patients with STEMI.”

https://link.springer.com/article/10.1007/s12265-023-10358-w “Correlation Between GDF11 Serum Levels, Severity of Coronary Artery Lesions, and the Prognosis of Patients with ST-segment Elevation Myocardial Infarction” (not freely available)

Brain endothelial cells

gettingwell4 4-5 stars Advanced knowledge, Acetylcholine, Cerebrum, Dopamine, Epigenetics, Glutamate, Hippocampus, Hypothalamus, Immune system, Limbic system, Memory, Neurochemicals, Stress , , , , ,

Six 2023 papers on the subject, starting with a rodent study:

“One of the primary discoveries of our study is that the endothelial cell (EC) transcriptome is dynamically regulated by both aging and heterochronic parabiosis. We found that ECs, when compared with other brain cell types, exhibited one of the highest fractions of aging-related genes that were rescued after heterochronic parabiosis in the old brain, and similarly, the highest fraction of aging-related genes that were disrupted after heterochronic parabiosis in the young brain. This finding supports our previous research that vasculature is strongly affected by aging and disease, and is capable of regrowth after heterochronic parabiosis or systemic GDF11 treatment.

parabiosis

We observed that a subset of ECs was classified as mitogenic. It is reasonable to speculate that the growth of these cells, which is probably prevented or suspended by the inflammatory environment of the aged brain, may be among the cell populations that respond to these interventions.

Although proteostasis in brain ECs has not been thoroughly investigated, they are apparently long-lived cells and, like neurons, might therefore accumulate protein aggregates with age, potentially compromising their function. ECs become senescent with age, but parabiosis may reverse that phenotype as well.

These findings underline the strong susceptibility and malleability of ECs, which are directly exposed to secreted factors in both brain parenchyma and blood, to adapt to changes in their microenvironment. ECs, despite comprising <5% of the total number of brain cells, are a promising and accessible target for treatment of aging and its associated diseases.”

https://www.nature.com/articles/s43587-023-00373-6 “Heterochronic parabiosis reprograms the mouse brain transcriptome by shifting aging signatures in multiple cell types”

A review elaborated on endothelial cell senescence:

“ECs form highly dynamic and differentiated monolayers arranged in a vascular network. Within brain tissue, the ECs of arteries, capillaries, and veins present different molecular characteristics. The main functions of ECs as a major cellular component of the blood-brain barrier (BBB) are to express cell membrane transport proteins, produce inflammatory mediators, deliver nutrients to brain tissue, and prevent drugs and toxins from entering the central nervous system.

ECs are the first echelons of cells affected at the onset of senescence due to their special structural position in the vascular network. Senescent ECs produce reactive oxygen species (ROS), which directly inhibit smooth muscle potassium channels and cause vasoconstriction.

The vascular endothelium is in a constant process of damage and repair, and once damage occurs, ECs replenish themselves to remove damaged cells. EC senescence makes the endothelium less capable of self-repair. With the decline in endothelial function, excess accumulated senescent cells express senescence-associated secretory phenotypes (SASPs), which result in senescence of adjacent cells, and eventually degeneration of vascular function.”

https://www.aginganddisease.org/EN/10.14336/AD.2023.0226-1 “Endothelial Senescence in Neurological Diseases”

A human study investigated above-mentioned differences in brain endothelial cells:

“We performed single nucleus RNAseq on tissue from 32 Alzheimer’s Disease (AD) and non-AD donors each with five cortical regions: entorhinal cortex, inferior temporal gyrus, prefrontal cortex, visual association cortex, and primary visual cortex. Analysis of 51,586 endothelial cells revealed unique gene expression patterns across the five regions in non-AD donors.

Visual cortex areas, which are affected late in AD progression and experience less neurodegeneration, expressed more genes related to vasculogenesis and angiogenesis. Highly vulnerable areas such as the entorhinal cortex expressed more oxidative stress-related genes in normal aged brain, suggesting endothelial dysfunction in this region even in the absence of severe AD pathology.

The present work shows that senescence-related gene signatures are increased across several brain regions, and confirms these changes in endothelial cells in the absence of other vascular cell types. While endothelial cells are not typically associated with protein aggregation, upregulated protein folding pathways suggest that proteostatic stress is a key pathway in this cell type.”

https://www.biorxiv.org/content/10.1101/2023.02.16.528825v1.full “Endothelial Cells are Heterogeneous in Different Brain Regions and are Dramatically Altered in Alzheimer’s Disease”

A human cell study abstract on above-mentioned blood-brain barrier endothelial cells:

“The BBB is a semi-permeable and protective barrier of the brain, primarily composed of endothelial cells interconnected by tight junction proteins, that regulates movement of ions and molecules between blood and neural matter. In pathological conditions such as traumatic brain injury (TBI), disruption of the BBB contributes to leakage of solutes and fluids into brain parenchyma, resulting in onset of cerebral edema and elevation of intracranial pressure.

The objective of this study was to determine upstream regulators of NLRP3 signaling and BBB hyperpermeability, particularly to determine if extracellular adenosine triphosphate (ATP) via P2X7R, a purinergic receptor, promotes NLRP3 inflammasome activation. Extracellular ATP is a major contributor of secondary injuries following TBI.

Our results suggest that extracellular ATP promotes NLRP3 inflammasome activation. Subsequent caspase-1 and MMP-9-mediated tight junction disorganization are major pathways that lead to BBB dysfunction and hyperpermeability following conditions such as TBI.”

https://journals.physiology.org/doi/abs/10.1152/physiol.2023.38.S1.5732827 “Regulation of Blood-Brain Barrier Endothelial Cell Hyperpermeability by NLRP3 Inflammasome Inhibition”

A human study further investigated effects of traumatic brain injury on brain endothelial cells:

“We previously demonstrated that extracellular vesicles (EVs) released from injured brains led to endothelial barrier disruption and vascular leakage. Here, we enriched plasma EVs from TBI patients (TEVs), detected high mobility group box 1 (HMGB1) exposure to 50.33 ± 10.17% of TEVs, and found the number of HMGB1+TEVs correlated with injury severity. We then investigated for the first time the impact of TEVs on endothelial function using adoptive transfer models.

HMGB1 is secreted by activated cells or passively released by necrotic or injured cells. After post-translational modifications, it interacts with receptors such as toll-like receptors (TLRs; e.g., TLRs 2, 4, and 9) and the receptor for advanced glycation end products (RAGE) to trigger multiple signaling pathways and mediate inflammatory and immune responses. Extracellular HMGB1 promotes endothelial dysfunction, leukocyte activation and recruitment, as well as thrombosis.

These results suggest that circulating EVs isolated from patients with TBI alone are sufficient to induce endothelial dysfunction. They contribute to secondary brain injury that are dependent on immunologically active HMGB1 exposed on their surface. This finding provided new insight for development of potential therapeutic targets and diagnostic biomarkers for TBI.”

https://www.sciencedirect.com/science/article/pii/S1043661823001470 “Circulating extracellular vesicles from patients with traumatic brain injury induce cerebrovascular endothelial dysfunction”

To wrap up, eat mushrooms to protect your brain endothelial cells!

“Natural compound ergothioneine (ET), which is synthesised by certain fungi and bacteria, has considerable cytoprotective potential. We previously demonstrated anti-inflammatory effects of ET on 7-ketocholesterol (7KC)-induced endothelial injury in human blood-brain barrier endothelial cells (hCMEC/D3). 7KC is an oxidised form of cholesterol present in atheromatous plaques and sera of patients with hypercholesterolaemia and diabetes mellitus. The aim of this study was to elucidate the protective effect of ET on 7KC-induced mitochondrial damage.

Protective effects of ET were diminished when endothelial cells were coincubated with verapamil hydrochloride (VHCL), a nonspecific inhibitor of the ET transporter OCTN1 (SLC22A4). This outcome demonstrates that ET-mediated protection against 7KC-induced mitochondrial damage occurred intracellularly and not through direct interaction with 7KC.

OCTN1 mRNA expression itself was significantly increased in endothelial cells after 7KC treatment, consistent with the notion that stress and injury may increase ET uptake. Our results indicate that ET can protect against 7KC-induced mitochondrial injury in brain endothelial cells.”

https://www.mdpi.com/1422-0067/24/6/5498 “Protective Effect of Ergothioneine against 7-Ketocholesterol-Induced Mitochondrial Damage in hCMEC/D3 Human Brain Endothelial Cells”