There are several alternative theories that counter the vitamin D–folate hypothesis. The skin mutagenesis and skin barrier hypotheses are two prominent theories proposed as explanations for the evolution of darker skin pigmentation [23
], and the energy conservation hypothesis is an alternative theory put forth to explain subsequent depigmentation [23
]. It is important to clarify that these are not alternative hypotheses to the vitamin D–folate hypothesis per se, as there is significant overlap between these theories and the actions of vitamin D and folate in the skin. The evidence for these theories afford explanatory power; however, it remains the case that these hypotheses relate to thousands of years of environmentally driven adaptive pressures, and such paradigms are difficult to test within contemporary populations. The manifestation of several theories for the evolution of skin colour is unsurprising. Skin colour is such an extensive polygenic adaptation to our environment that the explanation for its evolution is likely a complex picture that integrates these prominent theories and current unknowns.
3.1. The Skin Mutagenesis Hypothesis
The skin mutagenesis theory proposes that skin pigmentation arose as a mechanism to protect against the development of skin cancers [23
]. This hypothesis is based on the observations that more highly pigmented individuals are at lower risks of developing skin cancers because of the ability of skin pigmentation to combat UVR impact [33
]. It stands to reason that the pigment-facilitated protection of the vitamin D and folate status is involved in this hypothesis, as vitamin D and folate exert several photoprotective actions that would combat against skin malignancies [34
]. For example, vitamin D reduces UVR-induced DNA damage and cell death via an influence on multiple cell-cycle regulators (e.g., proto-oncogenes and tumour suppressors) and levels of reactive oxygen species [34
]. The actions of folate in DNA synthesis and repair pathways are important mechanisms to repair UVR-induced DNA damage and maintain genomic integrity [40
]. This hypothesis has several limitations that affect plausibility. Notably, there is much skepticism, given that the most fatal forms of skin cancer peak after reproductive age, and it is therefore difficult to argue that a low occurrence of skin cancers in individuals of reproductive age alone would have had an impact on natural selection [42
]. However, it is proposed that a selective advantage of protecting older (post-reproductive) adults against skin malignancies can be seen when considering the importance of older generations for offspring survival in hunter–gatherer communities [44
3.2. The Skin Barrier Hypothesis
The skin barrier hypothesis proposes that highly pigmented skin arose as a barrier that protected against multiple environmental stresses [23
]. UVR exposure causes deleterious changes in skin morphology, which reduces the ability of the skin to act as a defense barrier. Such damage includes disruption to skin permeability and a subsequent increase in transepidermal water loss [46
]. The skin barrier hypothesis is based on evidence that darker-pigmented skin types possess an enhanced barrier function compared to lighter skin types, mainly attributed to the role of melanin in scattering UVR [23
]. Compared to lighter skin types, darkly pigmented skin is shown to possess more robust permeability and greater structural integrity, barrier recovery, and skin surface acidity [23
]. In areas of high UVR and extreme humidity, darker skin pigmentation would have protected against the disruption of skin permeability via UVR and subsequent excessive water losses, and the acidity of the skin would have acted as a defense mechanism against microbial invasions. However, the feasibility of this theory is potentially limited, given that considerable discrepancies exist between studies examining ethnic differences in skin morphology.
This hypothesis is proposed as a discrete theory to the vitamin D–folate hypothesis. However, vitamin D and folate exert an array of functions that regulate the skin as a barrier against environmental stresses, having roles not only in the development of skin structures, but also in defense mechanisms that protect against UVR, heat, and microbial stresses.
The role of vitamin D in the skin is an area that has received significant research. The skin is a hub of vitamin D activity, as skin keratinocytes are unique in being the only cells in the body capable of both producing and metabolising vitamin D; being our primary site of vitamin D3
synthesis and also possessing all enzymes needed to metabolise inert vitamin D to its active form, calcitriol [48
]. Vitamin D, in turn, regulates several pathways involved in maintaining skin integrity.
Vitamin D regulates many processes involved in the development of the stratum corneum, the outmost layer of our skin. This is a highly permeable layer that expresses multiple elements of our adaptive and innate immune system, operating as a barrier against extensive water loss and microbial invasion [13
]. Vitamin D promotes the differentiation of keratinocytes into cells of the stratum corneum (corneocytes) via modulating calcium levels and the expression of protein components of the skin [13
]. Vitamin D also regulates the permeability of this layer via involvement of VDR in the synthesis of long-chain glycosylceramides, which form part of the lipid-enriched membranes around corneocytes [49
]. In addition, this nutrient plays several roles in immune responses in the skin. As examples, vitamin D may increase the expression of a key antimicrobial protein, cathelicidin, and the secretion of cytokines from T-cells: two modes of antimicrobial defense expressed on the skin surface [50
]. Folate may also play a part in skin immune responses, although this role is not well understood. Folate deficiency is associated with a decline in cell-mediated immunity, driven by a reduction in T-cell proliferation [51
]. The folate status is also linked to the expression of multiple proteins involved in immune function, inflammation, and coagulation in human blood [52
]. Notably, a high folate status correlates with increases in the expression of proteins involved in the activation and regulation of the complement system, an important non-specific skin defense mechanism [52
Folate may have a role in melanogenesis by regulating the production and stabilisation of tetrahydrobiopterin [53
]. Tetrahydrobiopterin is a required cofactor for tyrosine hydroxylase, which converts tyrosine into dopa in the production of melanin pigments [57
]. It could be suggested that folate and melanin compounds are synergistic; melanin, on the one hand, protects folate from UVR-related degradation, which in turn supports the influence of folate in melanogenesis. Interestingly, tetrahydrobiopterin also acts as a cofactor in the synthesis of nitric oxide, which has its primary function as a vasodilator in blood vessels [54
]. Vasodilation is the body’s primary response to heat stress, with increased blood flow allowing body heat to be lost via the skin through convection [58
]. From an evolutionary perspective, our ability to maintain vasodilation/vasoconstriction mechanisms would have been important in surviving varying UVR environments. As these mechanisms may been seen as relatively short-term responses to temperature changes, they are likely to be of greater importance in temperate UVR environments rather than environments of high UVR. This is supported by nitric oxide dependent vasodilation shown to be reduced in darkly skinned populations [59
]. This suggests that vasodilation processes offer no advantage in extreme UVR environments but may be important in temperate UVR environments, where seasonal and daily temperature fluctuations are seen. Vitamin D is also suggested to influence vasodilation by its influence on nitric oxide synthase [60
] and vasoconstriction by influences on the renin-angiotensin system [61
3.3. The Metabolic Conservation Hypothesis
The metabolic conservation theory proposes that the depigmentation of ancestral humans can be explained by a need to draw resources away from melanin production and towards other metabolic processes [23
]. Melanogenesis involves several production steps and feedback/crosstalk mechanisms that are dependent on energy input [62
]. In migrating to lower-UVR environments, it is likely that we lost the pressure to produce melanin to counter UVR-related stresses. The metabolic conservation theory suggests that the evolution of intermediate European and Asian skin tones allowed for the shunting of resources away from melanin production, to be used instead to combat against stressors associated with colder climates. However, this theory still supports the likeliness of the extreme dilution of pigmentation amongst northern Europeans being a mechanism to facilitate vitamin D production in low-UVR environments [23
The conservation theory is therefore not entirely isolated from the vitamin D–folate hypothesis. It is not a stretch to suggest that the protection of the vitamin D and folate status via pigmentation acted as the predominant pressure for the evolution of contrasting skin types at the equator and near the poles, as the likelihood of a deficiency in these nutrients would be highest in these environments. The occurrence of intermediate skin types that display facultative pigmentation in central European and Asian populations would have allowed for adequate vitamin D production. However, the primary “driver” may have been a need to restrict melanin production and channel these resources into responding to increased energy needs associated with colder climates. However, this is not to say the importance of vitamin D in intermediate UVR environments would have been obsolete. Notably, functions of vitamin D and folate in vasodilation/vasoconstriction outlined above, as well as roles in adipocyte biology, may have been important in maintaining energy and temperature homeostasis in increasingly colder climates.
Energy stored in adipose tissue can be utilised to maintain cellular functions in cases of increased energy needs or to fuel adaptive thermogenesis in response to cold stress [63
]. When energy is scarce, energy needs can be generated in white adipose tissue via an increase in fatty acid β-oxidation and the subsequent shunting of fatty acids into the electron transport chain to generate adenosine triphosphate (ATP). A similar mechanism occurs in brown adipose tissue, which has a principal role in regulating adaptive thermogenesis. In brown adipose tissue, β-oxidation results in the generation of energy in the form of heat [63
The roles of vitamin D in regulating fatty acid β-oxidation, energy metabolism, and the formation of brown adipose tissue are indicated by studies employing VDR-null mice models [63
]. These actions are proposed to involve the role of vitamin D in regulating the expression of related genes, with the vitamin D status being associated with the expression of genes such as PGC1α
, and AMPK
involved in mitochondrial biogenesis and thermoregulation [65
]. The roles of vitamin D in adipose tissue may have been important in increasingly colder climates. Even in temperate climates, these mechanisms would have been needed to respond to daily and seasonal variations in temperature and subsequent changes in energy needs [58
]. Notably, this theory is supported by evident ethnic differences in cold responses, with darkly skinned subjects being more susceptible to cold injury compared to lighter-skinned individuals [66
]. It could therefore be suggested that the occurrence of depigmentation in areas of lower UVR was a necessary measure to not only preserve energy but also allow for more efficient responses to colder regimes.