How to measure biological age?

As mentioned in Week 127, I had biological age measured earlier this month, and received five reports two days ago on Sunday. Part of the company’s process is to follow up their reports (intrinsic aging, immune aging, pace of aging, telomere length, weight loss) with a consulting session to review and interpret, which lasted an hour yesterday.

Part of our conversation revolved around comparing my measurements with other customers. These people are a different population than people usually sampled for aging and other biomarkers, because people who pay to get their biological age measured probably actionably want to improve it.

We’ll see which items I asked the consultant to pass on to the company produce responses, and which interfere with their business or they’re too busy to get back to me. I offered more than a half-dozen specifics, but held back on items I didn’t think the consultant would adequately communicate.

I didn’t argue with the consultant’s recommendations for quercetin supplementation (at 4% bioavailability?) as part of a treatment for senescence (not measured in any of the reports?). I didn’t offer to follow-up with studies demonstrating yeast cell wall β-glucan (new to the consultant) effects on immune report findings here in my 19th year of taking it every day.

I did argue with their recommendation to take DHEA-S. I changed my mind about taking it a year and a half ago ago, but left blog posts up such as Take responsibility for your one precious life – DHEA for evidence that I’m learning.

Epigenetic clocks per The epigenetic clock theory of aging generally view biological aging as “an unintended consequence of both developmental programmes and maintenance programmes, the molecular footprints of which give rise to DNAm [DNA methylation] age estimators.”

So what would be appropriate anti-aging actions for customers to take? Should customers try to emulate youthful biological markers, and supplement DHEA-S to impact serum levels of insulin-like growth factor 1?

I don’t think so. Our bodies never evolved feedback mechanisms to determine “Time to stop the growth programs, you’ve survived to reproduction age.” Older people achieving teenagers’ DHEA-S levels and activating IGF-1 pathways, pretty much guarantees further biological aging as “an unintended consequence of both developmental programmes and maintenance programmes.”


It’s too early to recommend these biological aging measurements. We’ll see where it goes.

One good thing is the company wants their customers to tell them everything about what they’re doing. I exercise at least a half hour every day, eat Avena nuda oats for breakfast and AGE-less chicken vegetable soup for dinner, and take the following:

Before breakfast
– 3-day-old microwaved broccoli / red cabbage / mustard sprouts started from 10.7 grams of seeds, with nothing else an hour before or after
– Yeast cell wall β-glucan (Glucan 300), 1500 mg, with nothing else an hour before or after
– Calcium alpha-ketoglutarate 1 g

Breakfast, lunch, and dinner
– Hyaluronic acid, Nature’s Lab, 1 serving total
– Boron, Swanson Triple Boron Complex, 9 mg total

Breakfast and dinner
Acetyl-L-carnitine, 1 g total
– Balance oil, which blends linoleic acid 1400 mg with linolenic acid 350 mg, 2 times
– Betaine anhydrous, 3 g total
– Glucosamine hydroxychloride 1.5 g total, with chondroitin sulfate 1.2 g total
Taurine, 2 g total
– 3-day-old Avena sativa oat sprouts started from 20 g seeds, 2 times

Breakfast only
– Minerals and vitamins, RDA mainly, Kirkland Signature Daily Multi
– D3 25 mcg

Lunch only
– Vitamin K2 MK-7 600 mcg

Dinner only
– D3 50 mcg
– Zinc monomethionine 30 mg with 0.3 mg copper
– Lutein 25 mg with 5 mg zeaxanthin


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Broccoli sprout compounds at different growth stages

This 2022 study investigated 12 glucosinolate compounds in 9 broccoli cultivars across seeds, 3-, 11-, and 17-day-old sprouts:

“Broccoli is rich in glucosinolates (GLs) which makes it an excellent source of these nutraceuticals. Composition and concentration of GLs vary among broccoli cultivars and throughout developmental stages of the plant.

9 aliphatic GLs and 3 indole GLs were identified from 9 broccoli cultivars. Aliphatic GLs concentrations decreased with broccoli sprouts and seedling growth for most cultivars. Indole GLs amounts increased after germination and reached the highest level in Stage B 3-day sprouts or C 11-day seedlings, and fell back to a low level in D 17-day seedlings.

Stage B was a stage that sprouts grew with no lights in medium and were about to be transplanted into pots. Stage C was a period that seedlings began to grow a main leaf, while in Stage D they were growing the second main leaf.

stages

Relatively high accumulation of glucoraphanin and glucoerucin in Chunqiujiali seeds suggests that CQJL broccoli seeds could be used for extraction of beneficial aliphatic GLs in nutraceutical industry.”

https://www.mdpi.com/2223-7747/11/12/1563/htm “Variation in Glucosinolate Accumulation among Different Sprout and Seedling Stages of Broccoli (Brassica oleracea var. italica)”


These researchers are probably early in their careers. They may have otherwise measured broccoli sprout compounds like glucosinolate-hydrosolate isothiocyanates and others, as did the cited 3-day-old broccoli sprouts have the optimal yields. As those researchers said:

“From the perspective of comparison methods, broccoli varieties, and germination processes, there is still lack of a systematic comparison of SF yields and other bioactive compounds contents between broccoli seeds and sprouts.”

Glucoraphanin is not sulforaphane highlighted one pitfall in concluding “CQJL broccoli seeds could be used for extraction of beneficial aliphatic GLs in nutraceutical industry.” Selecting broccoli varieties highlighted another, and provided an example of how human-applicable broccoli sprout compound research across different varieties could be done:

“We found a clear difference in selecting functional broccoli by considering only the GSL content or hydrolysates.

  • Even if total GSL content and individual GSL content were high, ITC content could not be produced at a high level.
  • When GSL content is high, if nitrile formation rate was also high, more nitrile than ITC would be produced.”

Preteen practicing handstands 15 minutes before sunrise

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Minds of their own

It’s the weekend, so it’s time for: Running errands? Watching sports? Other conditioned behavior?

Or maybe broadening our cognitive ability with Dr. Michael Levin’s follow-ups to his 2021 Basal cognition paper and 2020 Electroceuticals presentation with a 2022 paper and presentation starting around the 13:30 mark:

Michael Levin - Cell Intelligence in Physiological and Morphological Spaces

“A homeostatic feedback is usually thought of as a single variable such as temperature or pH. The set point has been found to be a large-scale geometry, a descriptor of a complex data structure.”


His 2022 paper Technological Approach to Mind Everywhere: An Experimentally-Grounded Framework for Understanding Diverse Bodies and Minds:

“It is proposed that the traditional problem-solving behavior we see in standard animals in 3D space is just a variant of evolutionarily more ancient capacity to solve problems in metabolic, physiological, transcriptional, and morphogenetic spaces (as one possible sequential timeline along which evolution pivoted some of the same strategies to solve problems in new spaces).

Developmental bioelectricity works alongside other modalities such as gene-regulatory networks, biomechanics, and biochemical systems. Developmental bioelectricity provides a bridge between the early problem-solving of body anatomy and the more recent complexity of behavioral sophistication via brains.

This unification of two disciplines suggests a number of hypotheses about the evolutionary path that pivoted morphogenetic control mechanisms into cognitive capacities of behavior, and sheds light on how Selves arise and expand.

While being very careful with powerful advances, it must also be kept in mind that existing balance was not achieved by optimizing happiness or any other quality commensurate with modern values. It is the result of dynamical systems properties shaped by meanderings of the evolutionary process and the harsh process of selection for survival capacity.”


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Gut microbiota, SCFAs, and hypertension

Two 2022 rodent studies from the same research group on short-chain fatty acid effects, beginning with butyrate:

“Maternal nutrition, gut microbiome composition, and metabolites derived from gut microbiota are closely related to development of hypertension in offspring. A plethora of metabolites generated from diverse tryptophan metabolic pathways show both beneficial and harmful effects.

Butyrate, one of the short-chain fatty acids (SCFAs), has shown vasodilation effects. We examined whether sodium butyrate administration in pregnancy and lactation can prevent hypertension induced by a maternal tryptophan-free diet in adult progeny, and explored protective mechanisms.

Decreased tryptophan metabolites indole-3-acetamide and indoleacetic acid observed in offspring born to dams that received the trytophan-free (TF) diet coincided with hypertension. This suggested that gut microbiota-derived tryptophan metabolites might be an offsetting mechanism, but not a cause of TF-induced hypertension. Considering that TF intervention reduced abundance of Romboutsia and Akkermansia, and many species are able to metabolize tryptophan, further studies linking abundance of bacterial species and concentrations of tryptophan metabolites are still required to identify main tryptophan metabolite producers.

Sodium butyrate treatment during pregnancy and lactation offset effects of maternal tryptophan-deficiency-induced offspring hypertension, mainly related to shaping gut microbiome, mediating SCFA receptor GPR41 and GPE109A, and restoring the renin–angiotensin system. A better understanding of mechanisms behind tryptophan metabolism implicated in programming of hypertension is critical for developing gut microbiota-targeted therapies to halt hypertension.”

https://www.sciencedirect.com/science/article/abs/pii/S0955286322001619 “Sodium butyrate modulates blood pressure and gut microbiota in maternal tryptophan-free diet-induced hypertension rat offspring” (not freely available) Thanks to Dr. You-Lin Tain for providing a copy.


A second study was on propionate effects:

“Early-life disturbance of gut microbiota has an impact on adult disease in later life. Propionate, one of predominant SCFAs, has been shown to have antihypertensive property.

We examined whether perinatal propionate supplementation can prevent offspring hypertension induced by maternal chronic kidney disease (CKD). CKD is closely linked to adverse maternal and fetal outcomes, and is reported to affect at least 3%-4% women of childbearing age.

Male offspring were divided into four groups: control, CKD, control+propionate (CP), and CKD+propionate (CKDP).

nutrients-14-03435-g001

Perinatal propionate supplementation:

  • Prevented offspring hypertension;
  • Shaped gut microbiota with increases in species richness and evenness;
  • Increased plasma propionate level; and
  • Upregulated renal GPR41 expression.

Results reveal the feasibility of manipulating gut microbiota by altering their metabolites with early-life use of propionate to prevent offspring hypertension in later life.”

https://www.mdpi.com/2072-6643/14/16/3435/htm “Perinatal Propionate Supplementation Protects Adult Male Offspring from Maternal Chronic Kidney Disease-Induced Hypertension”


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An inflammation clock

Here are six 2022 papers that either cited the second study of Variable aging measurements, or provided further evidence for its findings. Let’s start with a citing study:

“This study aimed to investigate expression patterns and prognostic values of the inflammatory aging clock (iAge) in glioblastoma (GBM), and its relations with stem cells. Similar to epigenetic clocks and transcriptomic clocks, iAge could track multifaceted aging phenotypes and have clinical significance in translation medicine.

iAge was positively correlated with chronological age, and highly associated with immune cells and inflammatory activities. iAge could serve as a prognostic biomarker for overall survival, and could precisely predict GBM stem cells stemness.

We identified the physiological importance and function of iAge in GBM, and provided novel insights into how iAge is a critical event for development of GBM.”

https://www.frontiersin.org/articles/10.3389/fgene.2022.925469/full “Inflammatory aging clock: A cancer clock to characterize the patients’ subtypes and predict the overall survival in glioblastoma”


Beginning with a human osteoporosis study, five papers investigated cytokine CXCL9, which the iAge study found to be “clearly actionable as shown by our experiments in CXCL9 where we can reverse aging phenotypes.”

“We assessed whether levels of CXCL9 and CXCL10 were elevated in human serum samples of older adults who had incident hip fractures. Our findings revealed higher serum levels of CXCL9 in pre-fracture blood samples of men with subsequent hip fractures, compared with their non-fracture controls. There was no such difference in CXCL9 serum levels between cases and controls in women.

Serum CXCL9 improved the prediction of osteoporotic hip fracture in men. The association between CXCL10 and hip fracture risk was not statistically significant in either sex.

While our epidemiologic findings are supported by experimental data providing the mechanistic pathway for CXCL9 in regulating osteoclast recruitment, further studies are needed to confirm validity of our findings and determine their generalizability to other study populations. Underlying biological mechanisms that limit our findings to men but not women require further investigation.”

https://asbmr.onlinelibrary.wiley.com/doi/10.1002/jbmr.4646 “CXCL9 Predicts the Risk of Osteoporotic Hip Fracture in a Prospective Cohort of Chinese Men—A Matched Case–Control Study”


Two immune-mediated skin diseases, with a vitiligo review:

“Current findings emphasize the critical role of immune cells and their mediators in the immunopathogenesis of vitiligo. IFN-γ [interferon gamma] is the primary cytokine mediator that activates the JAK/STAT pathway, causing keratinocytes to produce the key chemokines CXCL9 and CXCL10.

Interactions between immune and non-immune cells finally result in apoptosis of melanocytes. Additional investigations of these pathways may provide an opportunity for finding possible therapeutic targets, as there are currently no targeted biological drugs available for treatment of vitiligo.”

https://www.mdpi.com/2227-9059/10/7/1639/htm “Current Concepts of Vitiligo Immunopathogenesis”

and a study of psoriasis:

“CXCL9 is an important chemokine involved in T cell recruitment, and is up-regulated in plasma of patients with psoriasis. Increased CXCL9 expression can aggravate the progression of psoriasis.

cxcl9 expression

IL-1β and CXCL9 were up-regulated and CLDN8 was down-regulated in psoriasis with statistically significant differences. Identification of potential key molecular markers and signaling pathways provides potential research directions for further understanding molecular mechanisms of psoriasis.”

https://www.wjgnet.com/2307-8960/full/v10/i18/5965.htm “Identification of potential key molecules and signaling pathways for psoriasis based on weighted gene co-expression network analysis”


Two lung-related studies, first, an editorial for a human lung transplant study that isn’t freely available:

“CXCL9 and CXCL10 are chemokines that bind to the shared receptor CXCR3, potentiating T cells, mononuclear cells, and natural killer (NK) cells. Previous studies demonstrated that presence of these chemokines in bronchoalveolar lavage samples preceded development of chronic lung allograft dysfunction (CLAD).

Acute rejection and acute lung injury are known risk factors to the development of CLAD, yet this study found that increased risk was dependent on the presence of CXCL9/CXCL10 plasma elevation. Early identification of patients at risk, possibly during the active inflammatory phase, rather than once abnormal wound healing pathways dominate resulting in irreversible injury, provides an attractive opportunity for intervention.”

https://onlinelibrary.wiley.com/doi/10.1111/ajt.17135 “CXCL9 and CXCL10 plasma levels: Potential keys to unlocking CLAD risk”

and a study of smoking effects:

“We collected blood samples from 78 healthy male volunteers aged 18–60, including non-smokers (n = 30), current smokers (n = 30), and ex-smokers (n = 18). Expression levels of CXCL9/MIG [monokine induced by IFN-γ] and sIL-6R significantly increased after smoking, and continued to increase after quitting smoking.

cxcl9 smoking

Changes in related cytokines after smoking cessation are mainly restorative, while some cytokines further strengthen the trend of smoking-related changes.”

https://www.mdpi.com/1420-3049/27/12/3715/htm “Effects of Smoking on Inflammatory-Related Cytokine Levels in Human Serum”


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Natural sulforaphane effects

This 2022 rodent cell study used the natural form of sulforaphane to replicate experiments performed with mixtures of its natural and unnatural forms:

“Natural sulforaphane (SFN) exists as a single enantiomer with a RS absolute configuration. Most studies focusing on its biological activities, in particular its anti-inflammatory and antioxidant activities, have been conducted using its racemic (rac) form. rac-SFN has shown these effects in several in vitro and in vivo models.

(R)-sulforaphane

These findings demonstrate that (R)-SFN was able to:

  • Modulate inflammatory response and oxidative stress induced by LPS stimulation in murine peritoneal macrophages;
  • Reduce pro-inflammatory enzyme expression (iNOS, COX-2 and mPGES-1) and cytokine production (IL-1β, IL-6, IL-17, IL-18 and TNF-α);
  • Inhibit MAPK, JAK2/STAT-3, and canonical and non-canonical inflammasome signaling pathways;
  • Reduce NO and ROS levels and up-regulate the Nrf-2/HO-1 axis; and
  • Modulate epigenetic changes through histone methylation (H3K9me3) and deacetylation (H3K18ac).

(R)-SFN could be a new epinutraceutical compound useful for management of several immunoinflammatory diseases.”

https://www.mdpi.com/1424-8247/15/8/966/htm “Immunomodulatory Effects of (R)-Sulforaphane on LPS-Activated Murine Immune Cells: Molecular Signaling Pathways and Epigenetic Changes in Histone Markers”


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Epigenetic effects of plasma concentrate

“We use data from a safety study (n = 18, mean age 74) to investigate whether human umbilical cord plasma concentrate (hereinafter Plasma Concentrate) injected weekly (1 ml intramuscular) into elderly human subjects over a 10-week period affects different biomarkers, including epigenetic age measures, standard clinical biomarkers of organ dysfunction, mitochondrial DNA copy number (mtDNA-CN), and leukocyte telomere length.

More than 20 clinical biomarkers were significantly and beneficially altered. Telomere length and mtDNA-CN were not significantly affected by treatment.

An increase in entropy means that the methylome becomes noisier. We found that entropy was significantly decreased after treatment. Decreased entropy may implicate rejuvenation of the epigenetic landscape after plasma concentrate treatments.

changes in methylation entropy

Treatment reduced DNA methylation-based GrimAge by an average of 0.82 years, suggesting a reduction in morbidity and mortality risk. By contrast, no significant results could be observed for epigenetic clocks that estimate chronological age.

Our study lends credence to the notion that there are youth-promoting factors in the secretome of umbilical cord plasma. This conclusion has also been reached by other researchers that have provided treatment with stem cells, which do not work by plasma dilution but primarily by providing humoral factors and changing the microenvironment of cells and tissues. While there may be youth-promoting microvesicles or humoral factors that are at work, we do not want to rule out the possibility that it is ‘young and undamaged’ albumin that leads to the improvements noted, especially in light of recent evidence for such a mechanism.

This first human epigenetic clock study of plasma concentrate treatments revealed age-reversal effects according to a well-established DNA methylation-based estimator of morbidity and mortality risk. Future placebo-controlled replication studies are warranted with a larger number of participants over a longer study period, which our laboratory has undertaken to pursue.”

https://onlinelibrary.wiley.com/doi/10.1111/acel.13696 “Umbilical cord plasma concentrate has beneficial effects on DNA methylation GrimAge and human clinical biomarkers”


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Garlic vs. broccoli

This 2022 human study compared effects of two supplements:

“We test the hypothesis that consuming glucoraphanin (from broccoli) or alliin (from garlic) results in the accumulation of sulforaphane and alliin and their associated metabolites in the human prostate gland in a randomised, double-blinded, 2 × 2-factorial, dietary supplement, four-week intervention study.

The predominant sulphur-containing metabolite in garlic is alliin, which is odourless and non-volatile. When the plant tissue is damaged, alliinase enzymes rapidly convert alliin to allysulfenates that condense to form allicin and other thiosulfinates, predominantly γ-glutamyl S-allyl-L cysteine (γ-SAC) and S-allyl-L cysteine (SAC).

The BroccoMax/GRN supplements (530 mg) contained 97.7 ± 6.70 µmol glucoraphanin. The Kwai/alliin supplements (715 mg) contained four garlic-derived metabolites: alliin (35.2 ± 0.52 µmol), γ-SAC (19.3 ± 1.91 µmol), SAC (1.8 ± 0.16 µmol), and allicin (21.4 ± 2.10 µmol).

Mean excretion of sulforaphane and its metabolites as a percentage of ingested glucoraphanin [aka bioavailability] was 56.21% (range 21–91%, SD ± 18.66).

sulforaphane bioavailability

Alliin was detected within the prostate of every participant. Estimation of dietary intake of alliaceous vegetables is challenging due to their widespread presence in processed foods, and it is likely that intake is often underestimated.

We provide evidence that sulforaphane can be detected in human prostate tissue following regular consumption of glucoraphanin supplements. In contrast, alliin and associated metabolites were not more abundant in prostates of men receiving the alliin garlic-derived supplement. It is conceivable that alliin does accumulate in human prostate tissue, but its turnover is much slower than that of sulforaphane so that a longer allium-free diet is required prior to an intervention to assess its accumulation.

Accumulation of sulforaphane and presence of alliin in prostate tissue, as demonstrated in this study, may result in local effects on healthy and cancerous cells through a variety of mechanisms. This may explain the reduced risk of prostate cancer incidence and progression following consumption of cruciferous and alliaceous vegetables.”

https://www.mdpi.com/2072-6643/14/16/3263/htm “Accumulation of Sulforaphane and Alliin in Human Prostate Tissue”


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Epigenetic clocks so far in 2022

2022’s busiest researcher took time out this month to update progress on epigenetic clocks. If I curated every study he’s contributed to, it would require at least three blog posts a week. I’ll link to a few he’s posted in August 2022 that are more appreciated in the researcher community.

“In my lab, we are looking for clocks that apply to multiple species at the same time, for example, universal pan-mammalian clocks. It’s all about enhancing translation.

If you have an intervention that rejuvenates a mouse, a rat, a dog, and a cat according to the same clock, then chances are high that it will also work in humans. Naked Mole-Rat Hyaluronan Synthase 2 Promotes Longevity and Enhances Healthspan in Mice

Several groups, including mine, are working on single cell methylation clocks. Researchers are building clocks that respond to lifestyle interventions, such as exercise.

Moving away from methylation, it would be nice to build similar clocks for other ‘omics’ data. Many researchers build clocks on the basis of other omics data, such as for chromatin, proteomics, and gene expression.

There are different platforms, but they all attempt to measure the same thing: biological age. LINE-1 RNA causes heterochromatin erosion and is a target for amelioration of senescent phenotypes in progeroid syndromes

Epigenetic clocks are ‘life course clocks.’ I don’t know any other biomarkers of aging that applies to fetal tissues as well, because most other biomarkers measure organ dysfunction. Epigenetic profiling and incidence of disrupted development point to gastrulation as aging ground zero in Xenopus laevis

There’s this company called Intervene Immune, founded by Greg Fahy, and they are using GrimAge and other epigenetic clocks in clinical trials. They are doing a Phase II clinical trial. By the way, I’m one of the participants.

I could name several other groups who are using epigenetic clocks in clinical trials. It would be interesting if more people would measure epigenetic age in clinical trials in humans, at least as a secondary outcome, because there’s always an opportunity to make a discovery.

If you compare GrimAge to other biomarkers, such as cholesterol or glucose levels, you will see similar noise levels there. Epigenetic clocks are remarkably robust compared to what else is used in the clinic. I would say that the issue with technical noise in epigenetic clocks has been solved.

I’m really glad that different companies and researchers pursue different avenues, since it diversifies our risk. If one of these approaches works, it will change the world.”

https://www.lifespan.io/news/steve-horvath-on-the-present-and-future-of-epigenetic-clocks/ “Steve Horvath on the Present and Future of Epigenetic Clocks”


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Non-patentable boron benefits

To follow up Is boron important to health? I’ll highlight a 2022 review of boron intake:

“Boron is essential for activity of several metabolic enzymes, hormones, and micronutrients. It is important for growth and maintenance of bone, reduction in inflammatory biomarkers, and increasing levels of antioxidant enzymes.

The average person’s daily diet contains 1.5 to 3 milligrams of boron. Boron intakes of 1–3 mg/day have been shown to improve bone and brain health in adults when compared to intakes of 0.25–0.50 mg/day.

One week of 10 mg/d boron supplementation resulted in a 20% reduction in inflammatory biomarkers TNF-α, as well as significant reductions (nearly 50%) in plasma concentrations of hs-CRP and IL-6. Calcium fructoborate, a naturally occurring, plant-based boron-carbohydrate complex, had beneficial effects on osteoarthritis (OA) symptoms. A double-blind study in middle-aged patients with primary OA found that all groups except the placebo group saw a reduction in inflammatory biomarkers after 15 days of food supplementation with calcium fructoborate.

Dietary boron intake significantly improves brain function and cognitive functioning in humans. Electroencephalograms showed that boron pharmacological intervention after boron deficiency improved functioning in older men and women, such as less drowsiness and mental alertness, better psychomotor skills (for example, motor speed and dexterity), and better cognitive processing (e.g., attention and short-term memory). Boron compounds can help with both impaired recognition and spatial memory problems.

We discussed the role of boron-based diet in memory, boron and microbiome relation, boron as anti-inflammatory agents, and boron in neurodegenerative diseases. Boron reagents will play a significant role to improve dysbiosis.”

https://www.mdpi.com/1420-3049/27/11/3402/htm “The Role of Microbiome in Brain Development and Neurodegenerative Diseases”


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Findings, or fun with numbers?

This 2022 rodent study investigated bone mass phenotypes and sulforaphane:

“Mouse strains can have divergent basal bone mass, yet this phenotype is seldom reflected in the design of studies seeking to identify new modulators of bone resorption by osteoclasts. Sulforaphane exerts inhibitory effects on in vitro osteoclastogenesis in cells from C57BL/6 mice. We explore whether a divergent basal bone mass in different mouse strains is linked both to in vitro osteoclastogenic potential and to SFX-01 sensitivity.

osteoclasts in three mouse strains

Powerful antioxidants are an alternative to achieve beneficial bone effects and avoidance of osteoporotic bone loss. Sulforaphane (SFN) is a natural antioxidant found at high levels (as glucoraphanin) in cruciferous vegetables. SFN activates the NRF2 pathway and has anti-inflammatory effects, protecting against oxidative stress in many cell types.

These findings suggest that BM cells derived from animals with a high in vivo bone mass are less sensitive to M-CSF and RANKL in vitro leading to lower osteoclastogenesis. They also support the hypothesis that similar sensitivity extends to inhibitory effects of SFX-01 on osteoclast formation/function.

It is important to stress that osteoclasts generated in these strains may simply undergo multinucleation in a manner related to their underpinning genetics, and that by coincidence alone this is matched to their bone mass.”

https://onlinelibrary.wiley.com/doi/10.1002/cbf.3734 “High bone mass in mice can be linked to lower osteoclast formation, resorptive capacity, and restricted in vitro sensitivity to inhibition by stable sulforaphane”


I curated this study primarily for its honesty. I’ll link this post to future posts of studies where researchers lack similar honesty.

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Broccoli sprouts and your brain

A 2022 review of Nrf2 signaling hilariously avoided mentioning sulforaphane, although of ~4,000 sulforaphane published articles, two were cited. I’ll curate it anyway to highlight referenced brain effects.

“A good stability of NRF2 activity is crucial to maintain redox balance and therefore brain homeostasis. In this review, we have gathered recent data about the contribution of the NRF2 pathway in the healthy brain as well as during metabolic diseases, ageing, and ageing-related neurodegenerative diseases.

A functional NRF2 system is important to regulate both neuroinflammation, i.e., activation of microglia and astrocytes, and oxidative stress in the brain. NRF2 and NF-κB transcription factors regulate cellular responses to inflammation and oxidative stress in order to maintain brain homeostasis. Both pathways have been described to inhibit each other.

Nrf2 brain aging

Future challenges will be to establish novel therapies to:

  • Increase NRF2 activation in specific cell types and/or brain regions; and
  • Modulate NRF2 pathway in senescent cells.

Modulation of NRF2 signalling pathway by using specific food products [like unmentioned broccoli sprouts] and phytochemicals [like unmentioned sulforaphane], dietary supplements [like unmentioned Vitamin D3], drugs, and epigenetic modifiers, alone or in combination, will help to limit inflammatory diseases, ageing process, and subsequently ageing-related diseases.”

https://www.mdpi.com/2076-3921/11/8/1426/htm “Normal and Pathological NRF2 Signalling in the Central Nervous System”


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Eat broccoli sprouts for your offspring

This 2022 rodent study investigated effects of glucoraphanin supplementation during pregnancy and lactation:

“We investigated whether dietary intake of sulforaphane glucosinolate (SGS [properly termed glucoraphanin]) during pregnancy and lactation influenced composition of gut microbiota in offspring:

  • Dietary intake of SGS during pregnancy and lactation caused significant changes in diversity of gut microbiota in 3-week-old offspring (SGS-3W) and 10-week-old offspring (SGS-10W).
  • Plasma levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in SGS-10W after injection of lipopolysaccharide were significantly lower than those of CON-10W group.
  • There were sex differences of gut microbiota composition in both SGS-3W and SGS-10W offspring.

glucoraphanin during pregnancy and lactation

This study has some limitations:

  1. We did not investigate mechanisms of how dietary intake of SGS during pregnancy and lactation modulated gut microbial communities in offspring.
  2. We found several signaling pathways in beneficial effects of SGS food pellet, and further study of the role of maternal intake of SGS food in these pathways is needed.
  3. We did not investigate mechanisms of relationships between maternal intake of SGS and long-term anti-inflammatory action in adult offspring, and further detailed study including epigenetic modification is needed.

These data suggest that dietary intake of SGS during pregnancy and lactation might produce long-lasting beneficial effects in adult offspring through persistent modulation of gut microbiota. It is likely that modulation of gut microbiota by maternal nutrition may confer resilience versus vulnerability to stress-related psychiatric disorders in offspring.”

https://www.sciencedirect.com/science/article/pii/S0955286322001681 “Long-lasting beneficial effects of maternal intake of sulforaphane glucosinolate on gut microbiota in adult offspring”


This study published results of a mother’s glucoraphanin intake where offspring never ate glucoraphanin, with beneficial effects at both 3 weeks (~prepubescent human) and 10 weeks (~young human adult). Maybe future studies will continue this paradigm on to a second or third generation to see whether there are also transgenerational epigenetic effects.

This study’s methods extracted glucoraphanin from 1-day-old broccoli sprouts into a powder containing 135 mg (0.31 mmol) glucoraphanin per gram. Each 1 kg of of treatment chow included pellets containing (2.3 mmol / 0.31 mmol) x 135 mg = 1 gram of broccoli sprout powder, 0.1% of food intake.

Per Drying broccoli sprouts, dried 3-day-old broccoli sprouts contain 10% moisture, and fresh 3-day-old broccoli sprouts contain 82.6% moisture. A gram of 1-day-old broccoli sprout powder may be an approximate equivalent of (.826 / .1) = 8 grams fresh 3-day-old broccoli sprouts for a mouse / kg of daily food intake. A human equivalent dose is (.826 / .1) x .081 x 70 kg = 47 grams of fresh 3-day-old broccoli sprouts / kg of daily food intake.

That’s about how much 3-day-old, microwaved, glucoraphanin-containing broccoli and red cabbage sprouts I eat every day, starting from 7.2 grams of seeds. I sprout another 3.5 grams of yellow mustard seeds into the mixture for taste.


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Week 120 of Changing to a youthful phenotype with sprouts

It was time for an annual physical last Wednesday. My focus was to see whether reducing sulforaphane intake per Week 87 had the desired effect on thyroid measurements.

That and other adjustments did! Readings of TSH 2.91 (0.45 – 4.50 uIU/mL), free T4 1.22 (0.82 − 1.77 ng/dL), and free T3 2.4 (2.0 – 4.4 pg/mL) were all in-range. 🙂

thyroid


I won’t repeat the Week 63 workbook calculations done after last year’s annual physical. To me, that’s another form of magical thinking.

Every one of those reference ranges, and optimal ranges built from all-cause mortality statistics, requires a suffix “of people who didn’t positively change their healthspan and lifespan.” What value is there in optimizing (pick a measurement) against those outcomes? Why compare my efforts, or results, or any other aspect of my life, to people who didn’t actionably care about their one precious life?

I’m not deflecting with poor measurements:

  • 3 of the 5 values in last year’s optimal ranges got better, and the other 2 stayed the same; and
  • 2 of the 4 values that weren’t in last year’s optimal ranges came into those ranges, and the other 2 got better but stayed outside an optimal range.

We each have a lot at stake. Bad things like diseases of old age happen on their own. If we want good things to happen, we have to make them happen.

Consider this from The impact of transgenerational epigenetic inheritance and early life experiences:

“Every disease is connected to the immune system.”

Are people making good choices every day for their immune systems?

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Eat broccoli sprouts for metabolic syndrome

This 2022 rodent study investigated sulforaphane’s effects on insulin resistance:

“Insulin resistance is one of the defining clinical traits of metabolic syndrome, which represents a constellation of metabolic disorders, mainly comprising obesity, type 2 diabetes mellitus, atherogenic dyslipidemia, and hypertension. This study aimed to investigate therapeutic effects and potential mechanisms of sulforaphane (SFN) on high-fat diet (HFD)-induced insulin resistance. Control chow diet was 10 kcal% fat, and HFD was 60 kcal% fat.

  • SFN was found to effectively reduce body weight, fasting blood glucose, and hyperlipidemia, and improve liver function in HFD-fed mice.
  • SFN led to increased expression of antioxidant genes downstream of Nrf2, and decreased accumulation of lipid peroxides MDA and 4-HNE.
  • SFN significantly reduced glutathione peroxidase 4 (GPx4) inactivation-mediated oxidative stress by activating the AMPK and Nrf2 signaling pathways.

Data suggested that GPx4 could be a key target through which SFN might activate Nrf2/ARE signaling pathway to decrease the extent of insulin resistance induced in HFD-fed mice. Taken together, SFN ameliorated HFD-induced insulin resistance by activating the AMPK-Nrf2-GPx4 pathway, providing new insights into SFN as a therapeutic compound for alleviation of insulin resistance.”

https://www.sciencedirect.com/science/article/pii/S075333222200662X “Sulforaphane alleviates high fat diet-induced insulin resistance via AMPK/Nrf2/GPx4 axis”


This study’s sulforaphane dose was the same as Eat broccoli sprouts for your heart and Broccoli sprouts activate the AMPK pathway at 0.5 mg / kg. It was administered five times a week for 8 weeks to a subgroup of HFD-fed mice starting after 8 weeks of HFD.

A human equivalent oral dose to these three studies’ subcutaneous doses would be a low (0.5 mg x .081) x 70 kg = ~3 mg sulforaphane. Per Eat broccoli sprouts for your gut, my sulforaphane intake is six times that every day.

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