Figure 2 – Stratum corneum structure and
organization: A) “Bricks and mortar” Two- model (adapted
from (Prausnitz et al., 2012)); B) Stratum corneumstructure: cornified envelope (blue);
corneocyte
lipid envelope (CLE, black); corneodesmosome (brown). C)Lamellar membrane structure with intercellular lipids. Insert, electron
micrograph: Murine skin was fixed in Karnovsky’s fixative, and
post-fixed with 1 % aqueous osmium tetroxide, containing 1.5 %
potassium ferrocyanide (adapted from Uchida (Uchida and Park, 2016)).
The interior of the corneocytes is filled with keratin filaments
embedded in a matrix mainly consisted of filaggrin (filament aggregating
histidine-rich protein) and its breakdown products. These amino acids
together with certain ions, such as chloride, sodium, lactate and urate
form the natural moisturizing
factors (Ovaere et al., 2009). These substances are endogenous
humectants that keep the hydration of the SC on an adequate
level. It is important for skin elasticity and permits regularly
function for hydrolytic enzymes of desquamation (Fowler, 2012). Reduced
levels of filaggrin and properly natural moisturizing factors are caused
by mutations of the filaggrin gene connected with atopic dermatitis,
ichthyosis vulgaris, psoriatic skin, ichthyosis and general xerosis (dry
skin) (Uchida and Park, 2016, Fowler, 2012).
The cornified envelope (CE) that
encloses the Corneocytes is formed by structural proteins crosslinked by
transglutaminases (Prausnitz et al., 2012). CE has a stable stiff
property to withstand mechanical barrier stress (Figure 2B) and is
surrounded by a layer of corneocyte lipid envelope (CLE) (Uchida and
Park, 2016). This CLE is a support to form lamellar membrane structures
and controls exit of hydrophilic agents from corneocytes, but the roles
of CLE in the SC remain totally understood (Uchida and Park,
2016).
The extracellular space in the SC is filled by lipid-enriched
layers (Uchida and Park, 2016) which are composed by ceramides
(45–50%), cholesterol (25%), cholesterol sulphate (5%) and free
fatty acids (FFAs) (10–15%) (Ng and Lau, 2015, Uchida and Park, 2016,
Madison, 2003). The lipids form lamellar membranes that stretch out in a
horizontal direction parallel to the corneocytes (Figure 2C). In
addition, corneodesmosomes are found in the lamellar membrane structure.
They are modified desmosomes including corneodesmosin, a structural
protein that determines corneocyte adhesion (Ovaere et al., 2009).
Corneodesmosomes consist of desmoglein-1 and desmocollin-1 connected to
other corneocytes, intensify cohesion and regulate desquamation by their
pH-dependent degradation (Uchida and Park, 2016).
Ceramides are mainly responsible for the skin renewal process (Mizutani
et al., 2009) and play a critical role for the lamellar organization of
this layer barrier (Prausnitz et al., 2012). At least 14 different
classes of ceramides have been isolated from the human epidermis
(Meckfessel and Brandt, 2014, Jurek et al., 2019, Sofia A. Costa Lima.,
2018, Mojumdar et al., 2016). Each ceramide molecule contains a polar
head group from sphingoid nature (sphingosine, phytosphingosine,
6-hydroxysphingosine or dihydrosphingosine) and two hydrocarbon chains
derived from a fatty acid, a sphingoid or fatty acid ester moiety.
Cholesterol is the second most prevalent lipid by weight in theSC (Zbytovská et al., 2008, Pouillot et al., 2008, Prausnitz et
al., 2012). The increment of cholesterol concentration in the lipid
bilayers correlates with the diminution of the membrane thickness and
density, and at the same time with an extension of the membrane surface
area. Cholesterol fluidizes the SC lipid bilayers at skin
temperature (Ng and Lau, 2015, Zbytovská et al., 2008).
The content of FFAs is 10 – 15%
of the SC lipids and they consist mainly of very-long-chain
species with ≥18 carbon atom and are mostly saturated, like ceramides.
In addition to lipid species, cholesterol sulphate is typically at 2–5
% weight ratio. Its potential functions are to help in the formation of
the lipid lamellae and stabilization of the SC by inhibiting
enzymatic degradation of corneodesmosomes (Prausnitz et al., 2012, Ng
and Lau, 2015).
The majority of lipids is synthetized by keratinocytes in stratum
granulosum and packed in lamellar bodies. These organelles deliver
their lipid content in the extracellular space by fusion with the plasma
membrane of keratinocytes of the stratum granulosum (Pouillot et
al., 2008). Any changes in the concentrations of these main lipid
components can damage barrier integrity that mediates normal barrier
function.
These lipid structures prevent the excessive loss of water from the
organism and block the entry of most topically administered drugs that
have high molecular weight and low lipophilicity. Thereby, this barrier
represents a great challenge for drug delivery across the skin, intended
either for local effects or for systemic therapy (Groen et al., 2008).
Concerning the functions of SC , it is established that this layer
is responsible for the barrier and immune functions, namely preventing
excessive water loss, maintaining body temperature, preventing the entry
of xenotoxic chemicals and allergens as well as the invasion of
microbes. Its functions also comprise the protection of the epidermis
from oxidative stress and from mechanical stress.
The epidermal permeability barrier prevents intake and uptake of
compounds. The SC barrier functions may be partially connected to
its low hydration of 15%–20% and its very high density (1.4
g/cm3 in the dry state) (Walters and Roberts, 2002).
The body temperatures maintenance and water balance are regulated by
blocking surplus evaporation of water from nucleated layers. One of the
basic properties of this barrier is the prevention of penetration of
allergens, microbial pathogens and xenotoxic chemicals (Uchida and Park,
2016). As a general rule, a compound with
molecular weight larger than 500
Dalton cannot pass through the SC (Bos and Meinardi, 2000).
The epidermis has a pH gradient: the extracellular pH stays neutral
between the stratum granulosum and SC , where it becomes
more acidic to ca. 4.5 at the outer SC layer. Acidification
enables antimicrobial activity and regulates, by enzymatic activity
performed by proteases, the formation and desquamation of the epidermal
barrier. The use of alkaline or neutral soaps lead to the increase of
the SC pH, which conducts to untimely cleavage of
corneodesmosomes, decline in SC cohesion and following impairment
of the epidermal barrier. The pH changes are reported to matter in the
pathogenesis of skin diseases such as atopic dermatitis, acne vulgaris
and Candida albicans infections (Ovaere et al., 2009). For
example, with pH below 5.5, the growth of Pseudomonasacne , Staphylococcus epidermidis and a problematic
microbial pathogen, Staphylococcus aureus , are inhibited (Uchida
and Park, 2016). The permanent bacterial flora in the skin presents a
comprehensive ecosystem. Staphylococcus and Micrococcusstrains and Diphtheroid bacilli are the main part of
nonpathogenic microflora. They consume the sebum like basic nutrient and
confine skin colonization by potentially pathogenic organisms (Pouillot
et al., 2008).
SC plays an important role in the innate immunity, which is
related with the presence of antimicrobial peptides such as
cathelicidin, dermcidin, RNase7, S100A7/psoriasin and defensins (Uchida
and Park, 2016). They have revealed potent antimicrobial activities
against a wide spectrum of microbes, including gram-negative and
gram-positive bacteria, fungi and some viruses.
Cathelicidin antimicrobial
peptide (CAMP) is inducible with infection, injury or inflammatory
response and stimulates the production of a signal lipid,
sphingosine-1-phosphate (S1P) under stress conditions and activate
vitamin D receptor (VDR). Also, defensins are classified in three
subfamilies, α-, β-, and θ-defensin. They are inducible peptides in
epidermis in response to microbial infection, inflammation. and
differentiation (Uchida and Park, 2016). Adaptive immunity in theSC is associated with the availability of urocanic acid. Thetrans isomer of urocanic acid is generated from histidine
(principally from NMF) by histidase but can be converted to thecis isomer through exposure to
ultraviolet (UV) radiation.Cis -urocanic acid binds to the serotonin
[5-hydroxytryptamine (5-HT)]
receptor to eliminate immune function (Egawa et al., 2010). Urocanic
acid is an epidermal major chromophore, which works as a powerful
endogenic UV absorbent. Most lower UV wavelengths (UVB = 280–315 nm)
are absorbed in the epidermis, but longer wavelengths (UVA = 315–340
nm) get to the dermis. The bulk amounts of proteins, lipids, and
nucleotides has individually low potent chromophores, but they can form
the UV barrier together (Uchida and Park, 2016). The SC is
constantly exposed to oxidants, including UV light, chemical oxidants
and air pollutants. α- and γ-Tocopherol, ascorbic acid and glutathione
are the chief hydrophobic antioxidants of SC , providing the
lipid bilayers stability and safeguarding from lipid peroxidation
(Pouillot et al., 2008).
Dermis
The dermis is the bigger layer of the skin, with a thickness of
approximately 1–2 mm and provides important physical properties, namely
flexibility, elasticity and tensile strength. It is an integrated system
of fibrous and connective tissue, composed by collagenous and elastin
fibbers, accommodating epidermally derived appendages (hair follicles,
nails, sebaceous glands and sweat glands) and sensory nerve endings,
lymphatic vessels and blood capillaries. Thus, metabolic exchanges
between the epidermis and the blood systems may occur as well as
clearance of cell metabolic products and penetrated foreign agents. The
dermis contains resident cells (e.g., fibroblasts and mast cells) and
cells from the immune system, including macrophages and dermal dendritic
cells.
The thermal barrier, energy depositary and protection from physical
stroke are mainly connected with adipose tissue associated with collagen
fibbers, found in the lower reticular dermis layer. The water content
reaches 70%, favouring hydrophilic drug uptake. Below the reticular
dermis layer, the fibrous connective tissue transitions to the adipose
tissue of the hypodermis. This is mainly constituted of adipocytes
interconnected by collagen fibbers, forming a thermal barrier able to
store energy and protect from physical shock (Sofia A. Costa Lima.,
2018, Forster et al., 2009).
Hypodermis
The subcutaneous layer or hypodermis is the innermost layer of the skin
and consists of fat cells. However, this layer can be absent in some
thin skin, for example on the eyelid. The hypodermis is between the skin
and the subjacent tissues of the body, such as muscles and bones. Larger
lymphatic and blood vessels are standing in this layer. Consequently,
the major functions of the hypodermis are insulation, mechanical
integrity and support and conductance of the vascular and neural signals
of the skin (Alkilani et al., 2015, Ng and Lau, 2015).
Drug skin penetration
routes
The skin is an attractive site for delivery of drugs and cosmetics. But
normal skin is a serious barrier to drug absorption, which is why
pharmacologists and cosmetologists became interested in the development
of new drugs, formulations and ways of delivery.
Drugs can be administrated through the skin providing a local action
(topical administration) or a systemic effect, reaching the bloodstream.
The types of administration of drugs through the skin can be organized
in many ways, depending of the criteria selected. A possible
classification can be performed considering the invasive and
non-invasive nature of the different routes. Regarding invasive routes
of administration, they can be categorized as the routes in which the
drugs can enter through the skin by needle injections (subcutaneous,
intramuscular or intravenous routes) and those that consist in the
implantation of a device. In the subcutaneous route, the needle is
inserted into fatty tissue thus reaching the bloodstream. This type of
administration is usually used for the administration of many protein
drugs (eg. insulin) as they are destroyed in the digestive tract.
Intramuscular route is considered preferably over subcutaneous route for
the administration of larger volumes of drugs. In the intravenous route,
the drug is administrated directly into a vein and is used to give a
drug in a rapid and in a well-controlled manner the drug is delivered
immediately to the bloodstream. Drugs can also be administrated by
implantation of a device which is inserted under the skin. This type of
administration is probably the less commonly used and is usually
considered for the delivery of the drugs by controlled release along the
time for longer periods .
Considering the non-invasive methods, there are four possible routes of
drug penetration across the skin: intercellular, intrafollicular,
transcellular and polar (Pouillot et al., 2008), as depicted in Figure
3A. Sometimes, the diffusion through the skin appendages (e.g., fair
follicles, sebaceous glands and sweat glands) is classified as
appendageal route (Prausnitz et al., 2004, Ng and Lau, 2015).
Intercellular and transcellular ways are considered transepidermal
pathways.