Shed Your Skin. A multiple vector strategy to reverse skin ageing

The photo is a colored scanning electron micrograph of the epidermis, the outermost layer of the skin you see, at lower magnification, when you look in the mirror. It consists of dead cells, so anti-ageing cosmeceuticals do not work here. They work in the deeper tissues, and they are becoming more effective. But we are still only scratching at the surface.

As the science of ageing has developed it has become apparent that the phenomenology of ageing is driven by multiple sub-routines. The ageing of the skin is a good example of this.

The characteristic macroscopic changes are driven by micro-structural changes in the extra-cellular matrix (ECM), in conjunction with micro-circulatory, mitochondrial and immunological decline, epigenetic shift, the related accumulation of senescent cells and time-related changes in the dermal and epidermal microbiota. Chronic inflammatory stress plays an important role in many of these drivers.

The overall ageing process is too complex to be reversed by any one magic bullet, but each one of the above sub-routines can be individually accessed and modified using pharmaco-nutritional tools.

Starting with the very obvious inter-individual differences in apparent skin ageing, and the ability of protective multi-component regimes such as the Mediterranean diet to slow ageing (1) and reduce cutaneous inflammatory stress (2), a combinatorial approach designed to counter all known ageing sub-routines would likely generate supra-additive benefits. These should show up as improved skin thickness, tensile strength, smoothness, viscoelasticity and overall appearance.

We can be more specific.

There are at least 8 major sub-routines involved in ageing of the skin. In no particular order, these include:

1. Degradation of the Extra-Cellular Matrix (ECM). This is due to reduced synthesis and increased breakdown, which both fall under the broad heading of ECM economics.
2. Senescence: the accumulation of dysfunctional and pro-inflammatory cells in the dermis
3. Epigenetic ageing: gradual shifts in methylation and acetylation patterns in DNA and histones respectively.
4. Cellular energetic decline: mitochondrial damage and dysfunction, leading to reduced energy (ATP) generation and increased ROS formation
5. Immunological decline
6. Microbiology shift: the development of an ‘old’ and pro-ageing skin microbiota
7. Declining dermal capillary blood flow
8. Falling levels of skin hydration, leading to dryness and loss of viscoelasticity

These sub-routines and the pharmaco-nutritional tools that counter each of them are discussed below.

1. ECM economics

Physical characteristics of skin such as thickness, tensile strength, elasticity and hydration capacity, and hence its overall appearance, are largely determined by the quantity and quality of the extracellular matrix (ECM).  One of the main mechanisms of age-related dermal atrophy is a reduction in the amount of ECM, particularly collagen in the dermis (1). This is largely due to decreased collagen production and increased degradation of collagen (3-6).

Other ECM components including elastin, glycosaminglycans (GAGs), and proteoglycans (PGs), also change during aging, contributing to a deterioration in fibroblast function as these cells transform from elongated and attached to the ECM, to round and unattached, and therefore functionally uncoupled from tensile forces within the ECM. This process, which leads to increased matrix metalloprotease (MMP) release and further losses of beta-signalling and reduced ECM formation (7, 8), is exacerbated by oxidative and inflammatory stressors such as UV, smoking and diabetes (9-11). The increase in MMP expression is accompanied by a reduction in endogenous protease inhibitors (12-14), leading to accelerated loss of ECM.

Significant elements in this complex are reversible. Various polyphenols inhibit MMP’s directly (15), reduce their expression via epigenetic effects (16), and up-regulate tissue matrix metalloprotease inhibitors such as TIMP-1 (17). This trio of effects favours ECM regeneration. Aged (rounded) fibroblasts regain their elongated appearance and function when they come into contact with intact ECM (18), indicating that the ECM microenvironment is a major factor in the functional aging of fibroblasts.

This last finding is particularly interesting, as the traditional model describes a loss of functional fibroblasts and an increase in senescent fibroblasts with age (ie 19). It was thought that this would inevitably lead to further skin ageing, due to the senescent cells pro-inflammatory (SASP) profile, but now it seems that this feed-forward cycle can be at least partially reversed (18). This could be usefully combined with senolytics (see below).

If ECM can be even partially restored – and there is good evidence that it can (20) – enabling fibroblast reattachment with the restoration of function, then further ECM regeneration would ensue providing that the local environment is shielded against chronic inflammatory, glycative, carbonylative and carbamylative stresses.

Compounds that target ECM turnover

1. 1-3, 1-4 beta glucans. These activate fibroblasts, macrophages and Langerhans cells via inter alia Dectin-1 and TLR-4 receptors (21, 22), leading to enhanced ECM formation (14) and enhanced dermal immune functions.
2. Copper-binding peptides. These exhibit an array of anti-ageing properties including improved wound contraction and epithelization, increased production of growth factors and activity of antioxidant enzymes, enhanced ECM formation and reduced signs of skin ageing (23, 24).
3. Anti-inflammatory and anti-oxidant agents such as polyphenols. These compounds act via multiple mechanisms including the up-regulation of NRF2 and the down-regulation of inflammatory signalling, MMP expression and activity (25). The amphiphilic polyphenols oleuropein and phlorotannin (26, 27) achieve dermal penetration and are effective not only at reducing inflammatory and MMP activity (15, 28, 29), but also at increasing TIMP-1 activity (18).
4. Retinoids. These compounds, which exert multiple effects on skin cells (30), are well known to promote keratinocyte proliferation and reduce MMP activity (ie 31).
5. Tricin. This O-methylated flavone derived from rice bran stabilises lysosomal membranes leading to further reductions in MMP and protease secretion, and enhanced ECM stability (32). In my experience, this is a potent stand-alone anti-inflammatory agent with multiple clinical applications.

Mast cells in the skin are centrally involved in neuroimmune inflammatory responses activated by stress (33). These cells are stabilized by the polyphenol quercetin (34).

2. Senescent cells and senolytics

The accumulation of senescent cells in the skin is associated with age-related tissue dysfunction via mechanisms which include the depletion of stem and progenitor cells, and an increase in local inflammatory tone via the Senescence-Associated Inflammatory Phenotype (SASP) (35). These mechanisms contribute to loss of dermal ECM and an increased cancer risk via the degraded immune function (ie 36). Numbers of senescent cells increase after exposure to UV, a known cause of accelerated skin ageing (37). The selective removal of senescent cells has been shown to lead to improved tissue, organ and organism function (38).

Compounds that act as selective senolytics

Various polyphenols including Procyanidin C1 have been demonstrated to exert senolytic activity (39). This compound restores young functional and micro-structural aspects of skin in pre-clinical models (40).

3. Epigenetic ageing

While the running down of epigenetic clocks and the accumulation of senescent cells appear to be distinct processes, they run in parallel and are thought to be inter-connected (41). The epigenetic clock developed by Horvath et al (42) appears to effectively measure the ageing of inter alia skin cells, and shows characteristic changes in methylation and acetylation of DNA and histone sites. Critically, these changes are slowed by better nutrition (43), which is associated with more successful ageing (44). Improved nutritional profiles include raised intakes of methyl groups, butyrate and other dietary constituents that contribute to higher methylation and acetylation status.

Progressive changes in methyl groups occur over time, involving both hypo- and hyper-methylation. In some DNA sites hyper-methylation develops but globally there is a loss of methyl groups (45), and in skin, the ageing accelerant UV causes hypomethylation at multiple sites (46, 47). At the whole-body level, hypomethylation at individual age-associated CpG sites is associated with reduced life expectancy (48).

Given that dietary shift has created a wide-spread decline in methyl status as measured by plasma homocysteine (HyC) (ie 49, 50), and that this is also associated with reduced life expectancy (51), strategies designed to enhance methylation status generally and in skin cells would be expected to respectively confer global and local anti-ageing effects.

With ageing there is also a global loss of acetyl groups, the status of which are significantly determined by histone deacetylase (HDAC). This enzyme removes acetyl groups on histones, allowing chromatin structure compaction and thence gene silencing. HDAC is efficiently inhibited by butyrate (52), the only significant dietary source of which is prebiotic fiber via fermentation by saccharolytic gram-positive bacteria in the large bowel.

Post-transitional dietary shift has reduced prebiotic fiber intakes by up to 2 orders of magnitude (53-56), contributing to reduced butyrate status, increased HDAC activity and reduced chromatin acetylation. Lower fiber intake (and butyrate status) is associated with a very significantly increased rate of ageing and risk of death (57), via multiple mechanisms. As butyrate also has significant anti-inflammatory properties (58), some of which are mediated via HDAC inhibition (59), enhanced acetylation status in the skin via upregulated butyrate synthesis would be expected to confer anti-ageing effects.

NB Optimizing acetylation status in the dermis would likely require intakes of prebiotic fiber more akin to mid-Victorian levels, ie 35-45 g/day.

As epigenetic regulatory mechanisms collaborate to ensure proper epidermal homeostasis, a program designed to slow or partially reverse the epigenetic clock would be expected to result in improved skin tone and texture.

Compounds that act to enhance methyl / acetylation status

Methyl enhancement: The atypical amino acid betaine is an efficient source of methyl groups with proven epigenetic impact (60), and has ancillary anti-inflammatory activity (61).

Acetyl enhancement: The HDAC inhibitor butyrate can be delivered directly to the skin, but is somewhat malodorous and would require masking.

The copper peptides are active epigenetic co-factors, and exert many of their anti-ageing effects via epigenetic mechanisms which involve a shift to anabolic dominance (62, 63); as do the retinoids (64).

4. Cellular energetics: mitochondrial ageing

Mitochondrial dysfunction and oxidative stress are components of ageing in all tissues, including skin. Within the basal layer constant cell division requires high respiration rates and consequent mitochondrial reactive oxygen species (ROS) generation. This causes progressive cell damage, and is a significant contributor to skin ageing (65).

Mitochondrial damage accumulates with age and UV exposure in skin cells (66), and adversely affects skin structure and function via ROS signalling (67).

Compounds that maintain / enhance mitochondrial function

The sulfur-containing atypical amino acid ergothioneinine scavenges reactive oxygen and nitrogen species via its sulfhydryl group (68). It accumulates in the mitochondria (69) and has been shown to protect mitochondrial DNA from oxidative damage due to hydrogen peroxide (70) or UV exposure (71). Ergothioneine also exhibits potentially chemo-protective actions, especially when combined with TLR-agonists such as the 1-3, and 1-6 beta glucans (72). The resulting changes in IL-6 and IL-12p40 (68) are generally anti-inflammatory, and ergothioneine has recently been designated a mitochondrial cytoprotectant (69, 70).

Melatonin and CoEnzyme Q10 have broadly similar mitoprotective effects (73), as does glytein, the orally bioavailable precursor for glutatione (74). Betaine has an ancillary mitochondrial supporting role (60).

5. Immunological decline

Ageing brings an increased incidence of skin infections and cancers. The increased risk of infection reflects reduced physico-chemical barrier functions of aged skin and immunological decline, which impacts cancer risk also. There is a loss of Langerhans cells, a parallel decline in adaptive immune competence (75) and reduced production of anti-microbial polypeptides (76). The resulting changes in the host / microbiota nexus contribute to chronic inflammation (‘inflammageing’) (77), which accelerates skin ageing by degrading the ECM.

Compounds that maintain / enhance immune function

The best-known immunomodulators are the 1-3, 1-6 beta glucans derived from baker’s yeast, natural compounds which provide increased immuno-surveillance and immune-modulatory effects (78). These exert anti-inflammatory effects (79) and act on immune (80) and non-immune (81) skin cells to accelerate wound healing (82), a complex sequence of actions that includes enhanced ECM formation (79). This has been shown to increase tensile strength in scar tissue (83) and elasticity in intact skin (84); both proxies for increased ECM quality and quantity.

Nb. 1-3, 1-6 beta-glucan is a PAMP or pathogen-associated molecular pattern. Other PAMPs and DAMPs (danger- or damage-associated molecular patterns) are less well researched, but likely to exert similar effects.

6. Skin microbiology

The ageing process is characterised by subtle but significant and fairly consistent changes in the microbiome (85) and in the skin microbiota (86). Evidence that dysbiosis predisposes to immune (87) and general senescence (88), that different skin bacteria exert a wide range of epigenetic effects on host tissue (89) and that the interplay between host anti-microbial factors and various microbiotal species plays an important role in determining skin health and appearance (90), makes a case for modifying aspects of the ageing process in the skin via microbiotal enhancement.

Metagenomic analyses show an abundance of Streptococcus pneumoniae, infantis and thermophilus on the skin of younger individuals and older persons with younger (more elastic) skin (91). Increasing the numbers of streptococcus improves skin structure and barrier function by upregulating collagen and lipid synthesis in aged cells, with resulting improvements in skin elasticity and hydration (89, 90).

Compounds that restore the epidermal host/microbiota interface:

The above effects of streptococcal species on the skin are mediated largely or wholly via the bacterial spermidine (91). A polyamine with established wound-healing (growth-promoting) effects when applied systemically or topically on the skin (92), spermidine can be characterised as an epimutagen, in the sense that it induces a favourable and consistent epigenetic shift linked to improved repair. This is similar to the case made for actives cited in Sections 1 through 4.

7. Dermal microvascular blood flow

By age 70, blood flow to the skin has fallen by 40% below levels occurring at age 20 (93), due inter alia to age-associated deficits in vascular control systems (94-96). These changes, together with decreased capillary density and organization (97), contribute to impaired skin perfusion and diffusive transport capacity of nutrients and waste products. This is not an end-state; anti-ageing strategies including smoking cessation and physical exercise have been shown to regenerate the dermal micro-vascular system and improve skin perfusion in elderly subjects (98, 99).

Compounds that enhance dermal microvascular blood flow:

Many polyphenols have the ability to maintain / enhance microvascular function (100-103). A selected group of polyphenols with known vascular effects, including rutin, quercetin, oleuropein and phlorotannin, would be expected to gain this particular physiological advantage.

8. Skin moisturising / hydration

As skin ages, the progressive loss of ECM contributes to increasing dryness with loss of elasticity and increased wrinkling (104). In tandem with ECM regeneration, improved hydration improves dermal vioscoelasticity and reduces wrinkle count and depth (104).

Compounds that enhance dermal/epidermal hydration:

Betaine is a highly effective hydrating agent (105, 106), as is hyaluronic acid (107). The structural and cosmetic effects of these compounds are relatively short-lasting, but can be used to generate early results in more sustained anti-ageing campaigns.

SUMMARY

Until very recently, the above multi-component approach was entirely theoretical. Private funding was made available for a small pilot study in which a version of this approach was combined with the use of near-IR laser irradiation, over a period of 4 weeks. While it is unusual to present results informally, I have been given permission by the funders to present a brief overview of the interim findings while a more formal study is put in place.

The investigators recorded up-regulation of the majority of genes involved in ECM formation, and down-regulation of most genes that play a role in ECM breakdown. Genes that coded for anti-inflammatory effects were down-regulated and genes involved in enhanced barrier functions were up-regulated, along with SIRT1 and NRF2. Mitochondrial numbers in fibroblasts in the treated areas, and telomere length, both increased very significantly.

These are obviously not scientifically satisfactory data, but I present them here in outline for consideration as they seem to outline the possible. Further and more extensive work will determine whether these results are also repeatable, and whether we really can reverse the arrows and the slings of time.

A paper presenting the intervention and the epigenetic changes in considerably more detail is pending in the Aesthetic Surgery Journal.

Next week: Children of the Corn. Be very afraid.

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