forming what is defined as cornified cell envelope. Cornified envelope consists mostly of loricin (85%), small proline rich (SPR) protein, involucrin, keratins, desmoplakin, and filaggrin [25]. Keratins are crosslinked to cornified envelope at the final stage in the lipid lamellae, which has the vital mechanical and water permeability barrier functions for skin homeostasis. Since keratins are the main cytoskeletal component of keratinized epithelium, the differences in the types of keratins found in the keratinized vs. non-keratinized epithelium also suggest significant functional differences (Figure 1.4). In the keratinized epithelium layers, keratins K5 and K14 are expressed in the basal layer, while K19 is expressed in the non-keratinized epithelium [27]. In the differentiating suprabasal layers, the keratinized epithelium expresses keratins K1 and K10, while K4 and K13 are expressed mostly in non-keratinized epithelium. In K1 and K10 deficient mice, the missing keratins in the cornified envelope maturation process induced significant inflammation, hyperkeratosis, and skin fragility, thus suggesting the functional need for these keratins in skin homeostasis [28, 29].
Using a double immunostaining technique with Nile Red and anti-Involucrin, Hirao and coworkers were able to shed light into the maturation level of cornified envelopes at various locations on the body, including the lips [30, 31]. This method utilized stratum corneum obtained by tape stripping, followed by double labeling with Nile Red and anti-Involucrin. The immature cornified envelope was double labeled with the hydrophobic Nile Red dye and anti-Involucrin, while the mature cornified envelope was labeled with Nile Red only. In this work, the maturation process of cornified envelope was illustrated by sequential tape stripping, where they illustrated both the presence of immature cornified envelope on the 25th tape strip and the transition towards more mature cornified envelope in the first tape strip [30]. They went on studying different locations of the body to demonstrate potential regional differences in the cornified envelopes. Immature cornified envelopes were discovered at the forehead and lips, but not in the trunk and extremities [30]. Hirao et al. hypothesized that the cornified envelope maturity is correlated with the barrier function, where locations (such as lips) with more immature cornified envelope also have higher trans-epidermal water loss [30].
Figure 1.4 Keratin assembly in the different layers of keratinized vs. non-keratinized oral epidermis [27].
The difference in functional properties between the lips and the cheeks was illustrated by Kobayashi and Tagami where they measured transepidermal water loss (TEWL), conductance, size of corneocytes, blood flow and surface skin temperature in over three hundred volunteers [32]. They reported significantly higher TEWL (33 vs. 11 g/m2/h), lower conductance (61 vs. 163 µS), larger and more parakeratotic corneocytes (824 vs. 776 µm2), higher blood flow (16 vs. 7 mL/min*g) and higher surface temperature (33 vs. 31 °C) on lips vs. on cheeks. The reported values not only verified the relationship of the poor barrier function of the lips with the physiological makeup of the lip tissue, but it also illustrated the need for lip moisturizing products, especially in dry and cold environments.
The ceramide profile can be used to shed light on the relationship of barrier function and surface properties for both the skin and lips. Ceramides, which consist of 30-40% of stratum corneum lipids by weight, have an integral role in barrier homeostasis [33, 34]. To date, there are over 300 ceramide species that have been identified on the human stratum corneum, and they can be loosely classified into 12 groups based on their hydroxylation patterns in the sphingoid bases and fatty acid moieties [34–37]. There are 4 main classes of sphingoid bases: Dihydrosphingosine (dS), sphingosine (S), 6-hydroxy sphingosine (H) and phytosphingosine (P) [33, 34]. In terms of the fatty acid moieties, there are 3 general classes: non-hydroxy fatty acid (N), α-hydroxy fatty acid (A) and esterified ω-hydroxy fatty acid (EO) [33]. These fatty acid moieties and sphingoid bases combine to make up the different ceramide classifications such as CER [NP], CER [EOS] and CER [NS]. The different ceramides containing different sphingoid bases and fatty acid moieties are generalized in Table 1.1.
Table 1.1 Structure and nomenclature of human stratum corneum ceramides [33].
| Fatty acid Sphingoid |
Non-hydroxy fatty acid [N] |
α-hydroxy fatty acid [A] |
Esterified ω-hydroxy fatty acid [EO] |
|
Dihydrosphingosine [DS] |
CER [NDS] | CER [ADS] | CER [EODS] |
|
Sphingosine [S] |
CER [NS] | CER [AS] | CER [EOS] |
|
6-Hydroxysphingosine [H] |
CER [NH] | CER [AH] | CER [EOH] |
|
Phytosphingosine [P] |
CER [NP] | CER [AP] | CER [EOP] |
There are 4 main classes of sphingoid bases: Dihydrosphingosine (DS), sphingosine (S), 6-hydroxy sphingosine (H) and phytosphingosine (P) [33, 34]. In terms of the fatty acid moieties, there are 3 general classes: non-hydroxy fatty acid (N), α-hydroxy fatty acid (A) and esterified ω-hydroxy fatty acid (EO) [33]. These fatty acid moieties and sphingoid bases combine to make up the different ceramide classifications such as CER [NP], CER [EOS] and CER [NS].
To illustrate the body regional difference and seasonal impact on the ceramide profile, Ishikawa et al. conducted a one-year study on ten Japanese subjects with normal skin. In this study, their research team measured TEWL and capacitance, as well as taking stratum corneum samples to quantify the different ceramide groups from 10 different locations of the body [33]. Their findings suggested that lips, palm and fingers have least amounts of measurable total ceramides, while forehead, lower legs, upper arms, and buttocks have the most [33]. Lips, along with the palm, fingers, and cheeks showed lower ratio of CER[NP], CER[EOS], CER[EOH] and CER[EOP] and a higher ratio of CER[NS] and CER[AS] [33]. Furthermore, amongst all body locations, lips were found to have the lowest carbon numbers in CER[NS] and the highest level in C34-CER[NS] [33]. These findings are very interesting, as ceramides with a shorter free fatty acid chain length and an increase in C34 ceramide and unsaturated fatty acids have been identified in the stratum corneum of atopic dermatitis patients [38, 39]. Similarly to the findings by Kobayashi et al., Ishikawa et al. also found lips as one of the anatomic regions with the highest TEWL values while having relatively
