Recent advances in characterizing biological mechanisms underlying UV-induced wrinkles: a pivotal role of fibrobrast-derived elastase

In clinical studies, the formation of facial wrinkles has been closely linked to the loss of elastic properties of the skin. Cumulative irradiation with ultraviolet (UV) B at suberythemal doses significantly reduces the elastic properties of the skin, resulting in the formation of wrinkles. In in vitro studies, we identified a paracrine pathway between keratinocytes and fibroblasts, which leads to wrinkle formation via the up-regulation of fibroblast elastases that degrade elastic fibers. UVB irradiation stimulates the activity of fibroblast elastases in animal skin. Scanning electron microscopy revealed that cumulative UVB irradiation elicits a marked alteration in the three-dimensional structure of elastic fibers, which is closely associated with the subsequent reduction in the elastic properties of the skin, resulting in wrinkle formation. Studies using anti-wrinkle treatments suggest a close relationship between the recovery of wrinkles and an improvement in the linearity of elastic fibers. Those studies also suggest a close correlation between the recovery in the linearity of elastic fibers and the improvement in skin elasticity. In a study using ovariectomized animals, we characterized the important role of elastase in their high vulnerability to UV-induced wrinkle formation. A synthetic inhibitor specific for fibroblast elastases significantly prevents wrinkle formation without reducing the elastic properties of the skin, accompanied by minor damage in elastic fibers. Finally, we identified an effective extract of Zingiber officinale (L.) Rose from a screen of many herb extracts, which has a safe and potent inhibitory activity against fibroblast elastases. Animal studies using the L. Rose extract revealed that it has significant preventive effects against UVB-induced wrinkle formation, which occur in concert with beneficial effects on skin elasticity. A 1-year clinical study on human facial skin to determine the efficacy of the L. Rose extract demonstrated that it inhibits the UV-induced decrease in skin elasticity and prevents or improves wrinkle formation in skin around the corner of the eye without changing the water content of the stratum corneum. Our long-term studies support our hypothesis for a mechanism of wrinkle formation in which cytokine expression is activated by UV irradiation and triggers dermal fibroblasts to increase the expression of elastase. That increase in elastase results in the deterioration of the three-dimensional architecture of elastic fibers, reducing skin elasticity and finally leading to the formation of wrinkles.     

Selective inhibition of skin fibroblast elastase elicits a concentration-dependent prevention of ultraviolet B-induced wrinkle formation

We previously reported that wrinkle formation in the skin following long-term ultraviolet B irradiation is accompanied by decreases in skin elasticity and the curling of elastic fibers in the dermis. We further showed that wrinkles could be repaired by treatment with retinoic acid and that this was concomitant with the recovery of skin elasticity ascribed to the repair of damaged elastic fibers. Those studies suggested that decreasing the tortuosity of dermal elastic fibers is an important factor involved in inhibiting or repairing wrinkle formation. Therefore, it is of particular interest to determine whether the inhibition of elastase activity in vivo would prevent the damage of dermal elastic fibers and might abolish wrinkle formation associated with the loss of skin elasticity. Because the major elastase in the skin under noninflammatory conditions is skin fibroblast elastase, we used a specific inhibitor of that enzyme to assess its biologic role in wrinkle formation. The hind limb skins of Sprague-Dawley rats were irradiated with ultraviolet B at a suberythemal dose three times a week for 6 wk. During that period, 0.1-10.0 mM N-phenetylphosphonyl-leucyl-tryptophane, an inhibitor of skin fibroblast elastase, was applied topically five times a week. N-phenetylphosphonyl-leucyl-tryptophane application at concentrations of 0.1-1.0 mM abolished wrinkle formation in a concentration-dependent manner, with a peak for inhibition at 1.0 mM. This inhibition was accompanied by a continued low tortuosity of dermal elastic fibers and a maintenance of skin elasticity. Measurement of elastase activity after 6 wk of ultraviolet B irradiation demonstrated that whereas phosphoramidon-sensitive elastase activity was significantly enhanced in the ultraviolet B-exposed skin, there was no significant increase in that activity in the ultraviolet B-exposed, N-phenetylphosphonyl-leucyl-tryptophane-treated skin. These findings suggest that skin fibroblast elastase plays an essential part in the degeneration and/or tortuosity of elastic fibers induced by cumulative ultraviolet B irradiation.     

Ovariectomy is sufficient to accelerate spontaneous skin ageing and to stimulate ultraviolet irradiation-induced photoageing of murine skin

Background: Wrinkling and sagging of the skin during photoageing is physiologically associated with diminished elasticity, which can be attributed to increased fibroblast-derived elastase activity. This degrades the dermal elastic fibres needed to maintain the three-dimensional structure of the skin. We previously reported that ovariectomy accelerates ultraviolet (UV)B-induced wrinkle formation in rat hind limb skin by altering the three-dimensional structure of elastic fibres. OBJECTIVES: In this study, we used hairless mice to assess the effects of ovariectomy with or without chronic UVA or UVB radiation on sagging and wrinkling of skin, on the elasticity of skin, as well as on matrix metalloproteinase activities in the skin. METHODS: Ovariectomies or sham operations were performed on 6-week-old female ICR/HR hairless mice. RESULTS: Even in the ovariectomy group without UV irradiation, the skin elasticity was significantly decreased during the 3-13 weeks after ovariectomy, which was accompanied by a significant increase in elastase activity in the skin. After UVA or UVB irradiation, skin elasticity was significantly decreased to a greater extent in the ovariectomy group than in the sham operation group, and this was accompanied by a reciprocal increase in elastase activity but not in the activities of collagenases I or IV in the skin. Consistent with the decreased skin elasticity, UVA irradiation for 12 weeks elicited more marked sagging in the ovariectomy group than in the sham operation group. UVB irradiation for 12 weeks also induced more marked wrinkle formation in the ovariectomy group than in the sham operation group. CONCLUSIONS: These results suggest that ovariectomy alone is sufficient to accelerate skin ageing and to increase UV sensitivity, which results in the further deterioration of the skin and photoageing, and may account for the accelerated skin ageing seen in postmenopausal women.     

Carbon dioxide laser treatment promotes repair of the three-dimensional network of elastic fibres in rat skin

BACKGROUND: We have previously reported that ultraviolet (UV) B irradiation induces a loss of linearity in the three-dimensional structure of dermal elastic fibres, which results in the reduction of elastic properties of the skin and leads to wrinkle formation. We further reported that repair of wrinkles by all-trans retinoic acid is accompanied by recovery of the linearity of elastic fibres. Carbon dioxide (CO2) lasers are widely used for treating wrinkles in cosmetic surgery. OBJECTIVES: To perform CO2 laser treatment of wrinkles induced in rat skin by UVB irradiation and to evaluate changes in the three-dimensional structure of dermal elastic fibres during wrinkle repair. METHODS: Wrinkles were induced in the hind limb skin of Sprague-Dawley rats by UVB irradiation (130 mJ cm-2 three times weekly for 6 weeks), followed by CO2 laser treatment (11.3 J cm-2). The surface appearance of the skin was evaluated by replica observation 6 and 10 weeks after CO2 laser treatment followed by measurement of mechanical properties using a Cutometer. Subsequently, perfusion fixation and digestion with formic acid were performed and elastic fibres were observed by scanning electron microscopy (SEM). Image analysis of SEM micrographs was carried out to evaluate the linearity in the three-dimensional structure of elastic fibres. RESULTS: Six weeks after CO2 laser treatment, all parameters of skin mechanical properties in the UVB-irradiated group recovered to levels of the control non-irradiated group, accompanied by repair of wrinkles and a significant increase in linearity of the three-dimensional structure of elastic fibres. CONCLUSIONS: These findings indicate that CO2 laser treatment has a therapeutic potential to repair wrinkles to non-irradiated levels through recovery of the three-dimensional structure of elastic fibres.     

All-trans retinoic acid promotes the repair of tortuosity of elastic fibres in rat skin

To elucidate the repair effects of all-trans retinoic acid (t-RA) on ultraviolet (UV)-induced tortuosity of elastic fibres in rat skin, the hind limbs of Sprague-Dawley rats were irradiated at a suberythemal dose of UV (three times/week for 6 weeks) and 0.025% t-RA in ethanol was applied topically five times/week for 6 weeks. The three-dimensional arrangement of elastic fibres with special reference to their linearity was quantified by image analysis using a scanning electron microscope following a combination of intravascular resin injection and selective digestion using formic acid. When t-RA was topically applied for 6 weeks on wrinkles formed by 6 weeks' exposure of the skin of rat hind limbs to UV radiation, the wrinkles disappeared to control levels, concomitant with an improvement in skin elasticity. The linearity of the elastic fibres was significantly (P < 0.01) increased compared with age-matched UV-irradiated controls. These findings suggest that the degeneration and deposition of elastic fibres accompanied by an increase in their linearity is involved in the process of wrinkle repair by topical application of t-RA.     

The formation of wrinkles caused by transition of keratin intermediate filaments after repetitive UVB exposure

It has been reported that the formation of wrinkles involves changes in the elastic properties of the dermis due to the denaturation of elastic fibers. Several studies have shown that the hydration condition of the stratum corneum is also important in wrinkle formation. It is, however, still unclear how the stratum corneum contributes to wrinkle formation. Here we investigated the relationship between the formation of wrinkles and changes in the physical properties and condition of the skin after repetitive ultraviolet B (UVB) irradiation of hairless mice (HR/ICR). Repetitive UVB irradiation caused wrinkles on the dorsal skin of the mice. The elasticity (E) of the stratum corneum of UVB-irradiated mice was significantly lower than that of age-matched control (unirradiated) mice. UVB exposure also caused a deterioration of the fibrous ultrastructure of keratin intermediate filaments (KIFs) in the skin. We conclude that the deterioration of KIFs in the stratum corneum caused by repetitive UVB irradiation decreases the elastic properties of the stratum corneum, resulting in the formation of wrinkles.     

Loss of elastic fibers causes skin wrinkles in sun-damaged human skin

BACKGROUND: Although wrinkling is the most obvious sign of aged skin, the detailed pathomechanism of wrinkle development has not been elucidated. OBJECTIVES: In this study, we investigated the role of elastic fibers in the formation of skin wrinkles. METHODS: Loss of elastic fibers was measured quantitatively in the facial skins of subjects representing seven decades, and then compared with wrinkle severities. We also investigated whether topical retinoic acid treatment to photoaged human skin can restore destroyed elastic fiber, and the correlation between wrinkle improvement with increase in elastic fibers in RA-treated facial skin. RESULTS: We found a significant correlation between decreases in the length, width, number and total area of oxytalan fibers and wrinkle severity. Furthermore, we found that topical application of retinoic acid (0.025%) to chronically photodamaged skin regenerated and restored elastic fibers, and that there was a significant positive correlation between the amount of newly regenerated elastic fiber and the wrinkle improvement caused by retinoic acid. CONCLUSIONS: Our results provide an objective insight into the role of elastic fibers in skin wrinkle formation by providing a quantitative correlation between changes in oxytalan fibers and the severity of skin wrinkling.     

Mechanics of wrinkle formation: micromechanical analysis of skin deformation during wrinkle formation in ultraviolet‐irradiated mice

Background/purpose: The mechanical aspects of wrinkle formation were studied in the dorsal skin of hairless mice. Methods: Wrinkles were induced by irradiating with ultraviolet (UV) B for 10 weeks, while observing skin deformation during wrinkle formation. Changes in skin dimensions were also observed during the specimen excision process. Wrinkle depth and interval were measured before and after removal of the cutaneous muscle layer. Local deformation of wrinkled skin during uniaxial stretch was also measured. Changes in curvature of skin specimens upon muscle layer removal were then observed to determine the force balance in skin layers. Results: The skin showed spontaneous contraction in response to UV irradiation. Wrinkled skin showed a marked decrease in the wrinkle depth and a slight increase in wrinkle interval following muscle layer removal, a peculiar mechanical response that cannot be explained by homogeneous deformation of the skin. This response was due to compressive deformations of dermal tissue caused by the muscle layer and concentrated at valleys of the wrinkles. Curvature measurements indicated that the muscle layer compressed the dermal tissue predominantly in the craniocaudal direction. Morphological observations showed that the wrinkles coincided with rows of pores and sulci cutis, where the structural stiffness of the horny layer was relatively low. The horny layer showed significant thickening. Conclusion: Taken together, we propose the following hypothetical mechanisms of wrinkle formation during UV irradiation: spontaneous contraction of the dermis while maintaining or increasing the epidermal area induces buckling of the epidermis into the dermis at mechanically weak lines, namely, the rows of pores and sulci cutis, and buckling may be amplified by the axial compression of the dermis by the muscle layer.     

Epidermal changes caused by chronic low-dose UV irradiation induce wrinkle formation in hairless mouse

To investigate the effects of chronic low-dose UV irradiation on the skin, hairless mice were irradiated with a 1/3 minimal erythemal dose (MED) of UV. We examined the relationship between visible changes and skin damage in the dermis and epidermis. Hairless mice were irradiated with UVB (20 mJ/cm2) and UVA (14 J/cm2) three times a week for 10 weeks, followed by a 24-week non-irradiation period. Visible fine wrinkling was present after 4 weeks of irradiation, and the wrinkling progressively worsened throughout the period of irradiation. The wrinkles remained after irradiation was discontinued. In dermal components, no significant histological changes in the collagen fibers and elastic fibers were found, and the amount of hydroxyproline was also not changed. Thus, in the epidermis, there was a significant increase in the number of stratum corneum layers and the terminal-differentiation marker, filaggrin, positive cells. The intensity of staining for the differentiation marker, keratin 1, was reduced. These changes were accompanied by wrinkle formation, and remained after discontinuance of irradiation. These findings suggested that chronic low-dose UV irradiation induces structural and quantitative changes in the epidermis as a result of keratinization impairment, and that this damage in the epidermis is an important causative factor in wrinkle formation.     

Accumulation of elafin in actinic elastosis of sun-damaged skin: elafin binds to elastin and prevents elastolytic degradation

Elafin has a primary structure with two functional domains; a transglutaminase substrate domain at the N-terminus and a protease inhibitor domain at the C-terminus. Elafin expression has so far been reported only for epithelial tissues. Accumulation of elafin was immunohistochemically detected in the actinic elastosis of sun-damaged skin. Exposure of normal skin to UVA induced elafin expression that colocalized with elastic fibers. Incubation of synthetic transglutaminase substrate domain of elafin and elastin molecules in the presence of tissue transglutaminase in vitro resulted in the formation of a higher molecular complex on SDS-PAGE. Elafin expression was not detected in normal cultured skin fibroblasts, but was induced by UVA irradiation at both messenger RNA and protein levels. When radiolabeled insoluble elastin was incubated with recombinant full-length elafin and tissue transglutaminase, insoluble elastin became more resistant to neutrophil elastase digestion. These results indicate that (1) dermal fibroblasts potentially express elafin on UV irradiation, (2) UV-mediated elafin interacts with elastin, and (3) the elafin-elastin complex protects elastic fibers from elastolytic degradation, leading to the accumulation of elastic fibers in the actinic elastosis of sun-damaged skin. The transglutaminase substrate moiety of elafin plays an important role in anchoring elafin at its proper sites of action during UV-induced aging processes.     

Unusual wrinkle formation after temporary skin fixation followed by UVB irradiation in hairless mouse skin

Abstract Generally, many wrinkles form on the human face, and temporary wrinkles eventually become permanent. We evaluated the effects of temporary skin fixation on wrinkle formation after UVB irradiation using the back skin of hairless mice. In the group treated with UVB irradiation immediately after production using cyanoacrylate resin of an artificial groove parallel to the midline, wrinkles formed parallel to the midline, an uncommon direction for wrinkle formation in this mouse model. These wrinkles did not disappear even when the skin was stretched. No such changes were observed in the group in which only the temporary groove alone was produced without UVB irradiation. In 3-D surface parameter analysis, all roughness parameters in the group treated with UVB irradiation immediately after production of an artificial groove were significantly increased relative to the age-matched control group. In contrast, no differences were observed between the group in which only the temporary groove alone was produced without UVB irradiation and age-matched controls. The results of this study suggest that both a temporary groove in the skin and UVB irradiation are necessary for wrinkle formation in the back skin of hairless mice.     

Loss of skin elasticity precedes to rapid increase of wrinkle levels

BACKGROUND: Decreased skin elasticity is considered as a factor that promotes wrinkle formation. However, the relationship between decreased skin elasticity and the formation of wrinkles is not fully understood. OBJECTIVE: The purpose of this study is to characterize the relationship between skin elasticity and wrinkle formation using quantitative methods. METHODS: Skin elasticity at the corner of the eye was measured using a Cutometer SEM575. Wrinkle levels at the corner of the eye were determined from three-dimensional analysis of surface replicas. Ninety healthy female volunteers living in Tokyo, Japan (aged 18-76 years) were examined in this study. RESULTS: In each scatter plot examined, women with lower U(r)/U(f) values or with higher U(v)/U(e) values showed a tendency for increased R(a) and R(max) values. However, this study determined widely scattered values for both types of parameters. These relationships were then reanalyzed in relation to age. Results were compared within a young (YNG) group of 27 women (aged 18-29), a middle-aged (MED) group of 26 women (aged 30-48) and an elderly (OLD) group of 37 women (aged 50-76). For elasticity parameters, U(r)/U(f) and U(v)/U(e) values, significant differences were found between the YNG and MED groups and also between the MED and OLD groups. For roughness parameters, a significant difference was found only between the MED and OLD groups. These results suggest that changes in R(a) or R(max) values lag about 20 years behind changes in skin elasticity values. CONCLUSIONS: Despite a strong relationship, elasticity parameters are unlikely to be an independent predictor of wrinkle levels. The amount of UV exposure to skin with decreased elasticity seems to be the variable that determines wrinkle levels. This finding represents another complexity in the mechanism of wrinkle formation.     

Fibrillin-rich microfibrils are reduced in photoaged skin. Distribution at the dermal-epidermal junction

Chronic sun exposure results in photoaged skin with deep coarse wrinkles and loss of elasticity. We have examined the distribution and abundance of fibrillin-rich microfibrils, key structural components of the elastic fiber network, in photoaged and photoprotected skin. Punch biopsies taken from photoaged forearm and from photoprotected hip and upper inner arm of 16 subjects with a clinical range of photoaging were examined for fibrillin-1 and fibrillin-2 expression and microfibril distribution. In situ hybridization revealed decreased fibrillin-1 mRNA but unchanged fibrillin-2 mRNA levels in severely photoaged forearm biopsies relative to photoprotected dermal sites. An immunohistochemical approach demonstrated that microfibrils at the dermal-epidermal junction were significantly reduced in moderate to severely photoaged forearm skin. Confocal microscopy revealed that the papillary dermal microfibrillar network was truncated and depleted in photoaged skin. These studies highlight that the fibrillin-rich microfibrillar network associated with the upper dermis undergoes extensive remodeling following solar irradiation. These changes may contribute to the clinical features of photoaging, such as wrinkle formation and loss of elasticity.     

Intensification of ultraviolet-induced dermal damage by infrared radiation

Summary Abnormal dermal deposition of elastic fibers is the earliest and most striking effect of prolonged sun exposure (solar elastosis). The hyperplastic fibers are usually ascribed to ultraviolet (UV) rays. Nonetheless, other portions of the solar spectrum may play contributing roles. Heat, for example, enhances experimental UV tumorigenesis. Heat induces erythema ab igne in which the structural alterations resemble those of actinically damaged skin, including the development of premaligant and malignant lesions. In regions of high insolation, infrared radiation (IR) is a constant companion of UV. To assess the role of IR in actinic damage to the dermis, albino guinea pigs were irradiated for 45 weeks with UV-B and UV-A, with and without IR. Control animals received IR only or no irradiation at all. Unirradiated dermis contains small amounts of elastic fibers in the upper dermis with greater depositions around follicles and sebaceous glands. After irradiation with UV, the fibers became more numerous, thicker, and more twisted; IR alone produced many fine, feathery fibers. The addition of IR to UV resulted in dense matlike elastic fiber depositions that exceeded what was observed with either irradiation alone. In combination or alone UV and IR radiation produced a large increase in ground substance, a finding also seen in actinically damaged human skin. Infrared radiation, in the physiologic range, though pleasant is not innocuous.     

Skin ageing: clinical and histopathologic study of permanent and reducible wrinkles

Wrinkles are modifications of the skin associated with cutaneous ageing and develop preferentially on sun-exposed skin. The aim of the study was to analyse the clinicopathological features of wrinkles, among the different types of skin relief modifications. Despite its importance in dermato-cosmetology and skin ageing, few studies have been specifically devoted to wrinkles. In the present study, we analyzed the histological features of the pre-auricular wrinkle compared to retro-auricular skin, obtained from sixteen patients undergoing facial surgery; skin samples were immediately processed for routine histology and histochemical staining. Four types of skin depressions could be defined according to their depth: folds, permanent wrinkles, reducible wrinkles and skin micro-relief. Two different types of pre-auricular wrinkles were observed: (i) permanent wrinkles which were conserved after sampling and, (ii) reducible wrinkles which required in vivo staining to be visible at histology. Histological analysis of the epidermis and dermis of the skin forming the pre-auricular wrinkle revealed a normal skin morphology, identical to that of the skin immediately adjacent to the wrinkle. This was particularly striking for the reducible wrinkles which could not be individualized in the absence of in vivo staining. Both types of wrinkles comprised skin modifications observed in sun-exposed skin, however, in the upper dermis, permanent wrinkles showed a more pronounced accumulation of basophilic fibers, i.e. actinic elastosis, than reducible wrinkles did. These data suggest that the development of wrinkles could be secondary to actinic elastosis and to the disappearance of microfibrils and collagen fibers at the dermal-epidermal junction.     

A novel treatment option for photoaged skin

Background DNA damage as a result of ultraviolet (UV) exposure plays an important role in the progression of cutaneous aging. Both folic acid and creatine have been linked to the process of DNA protection and repair. Aims This study aims to investigate the effects of a commercially available folic acid– and creatine‐containing formulation to fight the clinical signs of premature skin aging. Patients/methods Both in vitro and in vivo home‐in‐use studies using a folic acid– and creatine‐containing formulation were performed aiming to elucidate the efficacy in terms of improvement of skin regeneration, protection from UV‐induced DNA damage (Comet assay), reduction of wrinkle volume, and skin visco‐elasticity. Furthermore, clinical evaluation and photography were carried out to determine the improvement of clinically graded parameters after treatment. Results Cultured full‐thickness epidermal skin models supplemented with folic acid and creatine after epithelial perturbation showed an accelerated skin regeneration compared to untreated control models. Similarly, application of a folic acid– and creatine‐containing formulation significantly improved epidermal turnover in vivo as evidenced by smaller corneocytes derived from the treated sites relative to the vehicle‐treated sides. In addition, topical in vivo application of this formulation significantly protected from UV‐induced DNA lesions, increased skin firmness, and reduced wrinkle volume compared to untreated control areas. Expert grading confirmed a significant decrease of fine and coarse wrinkles in the periocular region as well as overall wrinkles, tactile roughness, and laxity. Conclusions Taken together, these results show that the combination of folic acid and creatine significantly accelerates epidermal skin regeneration in vitro and in vivo. Together with the finding of improved biomechanical skin properties, we conclude that the described topical formulation provides an effective treatment option for (photo)‐aged skin.     

Skin rejuvenating effects of chemical peeling: a study in photoaged hairless mice

Background Chemical peeling is a dermatologic treatment for skin aging. However, the mechanism by which the chemical peel achieves its results is not clear. We investigated the effects of chemical peeling and the mechanism of wrinkle reduction in photoaged hairless mice skin. Methods After inducing photoaged skin in hairless mice by repetitive ultraviolet‐B irradiation applied over 14 weeks, we applied trichloroacetic acid (TCA) 30%, TCA 50%, and phenol on areas of the same size on the backs of the mice. Punch biopsies were obtained 7, 14, 28, and 60 days after the procedure for histologic and immunohistochemical analyses. Results Histologic examination showed an increase in dermal thickness, collagen fibers, and elastic fibers in the dermis of intervention groups compared with control groups. These increases were maintained significantly for 60 days. Conclusions This study demonstrates that chemical peeling reduces wrinkles and regenerates skin by increasing dermal thickness and the amount of collagen and elastic fibers in photoaged skin.     

The severity of wrinkling at the forehead is related to the degree of ptosis of the upper eyelid

Background: Most studies on wrinkle formation have focused on changes in the dermal condition that promote the fixation of transiently formed wrinkles. Little is known about the age‐dependent changes in transient wrinkle formation in response to altered facial expression or the mechanism leading to fixed wrinkle formation. Objective: To clarify the mechanism of wrinkle formation at the forehead, we investigated the factors that influence the severity of transient wrinkling and the relationship of transient with fixed wrinkles, using a newly established method to evaluate transient wrinkle formation. Methods: Transient wrinkles were generated by requesting subjects to gaze in an upward direction. Foreheads of the subjects with or without an upward gaze at a fixed angle were photographed and the severity of wrinkles at the forehead was graded from 0 to 5 in 50 healthy Japanese female volunteers in their 20s, 40s, or 60s. Skin elasticity was measured using a Cutometer^®. Frontalis muscle activity and ptosis of the upper eyelid were estimated by measuring movement of the eyebrow during upward gazing and the position of the upper eyelid of the open eye, respectively. Results: Wrinkles formed transiently at the forehead by upward gazing were highly reproducible in each subject. Their severity increased with aging and was highly correlated to that of fixed wrinkles (R=0.81, P<0.001). Therefore, this method appears to be suitable for studying the mechanism of transient wrinkle formation and the relationship between transient and fixed wrinkles at the forehead. The severity of transient wrinkles was correlated with elevation of the eyebrow during upward gazing (R=0.69, P<0.001), but not with dermal elasticity. This suggests that transient wrinkles are induced by increased frontalis muscle activity during upward gazing. Frontalis muscle activation was negatively correlated with upper eyelid position (R=−0.37, P<0.05), which descended with aging, meaning ptosis of the upper eyelid, and negatively correlated with the severity of transient wrinkles induced by upward gazing (R=−0.43, P<0.05). Furthermore, the upper eyelid position was also negatively correlated with the severity of fixed wrinkles (R=−0.44, P<0.05). Conclusion: These results suggest that ptosis of the upper eyelid is associated with increased activation of the frontalis muscle during upward gazing and increased severity of transient and fixed wrinkling at the forehead.     

Light and scanning electron microscopic studies on wrinkles in aged persons' skin

Wrinkles in six aged persons (67–82 years of age) have been investigated by light microscopy (LM) and scanning electron microscopy (SEM). There are two types of wrinkles. One is a deep wrinkle which develops on the sun-exposed skin and does not disappear on stretching (permanent wrinkle). The LM and SEM showed less elastotic change in the upper dermis in the area of wrinkle than in that of the surroundings. The other type is a shallow wrinkle which develops on sun-protected skin and disappears on stretching (temporary wrinkle). The LM and SEM showed the decrease or loss of the elastic fibres in the papillary dermis as seen in ageing skin.