Unique Retinol Therapy with Antioxidant and Anti-Inflammaging Complex for Naturally Reborn Skin: The Clinical Case Series Study

. One of the methods used to prevent symptoms of skin aging are chemical peels. Among retinoids, retinol is much less described in the literature than tretinoin as a substance used in the peel procedure. Te aim of our study was to determine the efcacy of using a 4% retinol solution product containing novel TGF-β activators and antioxidants in improving the condition of the skin in patients with facial skin issues caused by aging and skin disorders. Each of 15 patients went through three peel treatment series, with a 30-day gap between each session and was examined using Multi Skin Test MC 1000 Courage+ Khazaka, evaluating hydration, elasticity, and skin pigmentation and Observ 520x. All patients reported overall improvement after the peeling procedure, and most of them reported subjective improvement in the reduction of the facial skin: tone, skin pigmentation, dehydration, dryness, structure, and sebaceous gland activity. An objective evaluation revealed signifcant clinical improvement in pigmentation in study groups. Elasticity parameters were shown to increase during treatment, with a greater efect occurring in patients with mature skin with frst signs of aging and patients with postinfammatory hyperpigmentation, atrophic acne scars, and enlarged pores. Concluding, combined peel therapy of 4% retinol serum product containing novel TGF-β activators and antioxidants showed good efcacy in reducing facial pigmentation of the skin high patients’ satisfaction with the procedure. Te study revealed that the described method is simple to use, has low cost, is with rare adverse events, and has well tolerability.


Introduction
Te skin is the largest single organ in the human body with a surface area of around 2 m 2 and a mass of 15% of the total body mass [1].Te main functions of the skin include regulating body temperature, protecting the organism from mechanical injuries and infections, and ensuring waterelectrolyte management.Despite these very important functions, it also has an important cosmetic role.It is known that young-looking skin and appearance may have a positive infuence on people's social behavior [2][3][4].According to the literature, factors such as age, skin diseases, and exposure to sunlight may change the skin condition.With age, the skin becomes thinner and progressively less hydrated, mostly due to a reduction in the quantity of natural moisturizers [5,6].
Over the years, as a result of the declining collagen synthesis in the dermis and the resilience of its already-existing collagen and elastin fbers, the skin also loses its elasticity.Tese processes defned by the clinical, histological, and physiological decrements that occur in the sun-protected skin are called in the literature as intrinsic aging [5,[7][8][9].Te second factor causing skin aging is long-term exposure to sunlight which belongs to extrinsic aging factors.It commonly manifests as pigmentation changes (hypopigmentation or hyperpigmentation), rough skin, dryness, telangiectasias, and wrinkles [9][10][11].Histologically, these are the result of alterations in dermal connective tissue, accumulation of disorganized elastin, its microfbrillar component fbrillin in the deep layers of the dermis, and acute loss of interstitial collagen [12].
One of the methods used to prevent symptoms of skin aging are chemical peels [13].Tese are divided into superfcial, medium-depth, and deep kinds, based on how deeply they penetrate the skin.Not only the substance but also its concentration, pH of the solution, and time of application have an impact on the procedure efects [14].Commonly used superfcial peels include tretinoin, salicylic acid, Jessner solution, and also glycolic and pyruvic acid.Tey target the epidermis and the epidermal-dermal interface, causing decreased corneocyte adhesion, epidermolysis, and increased dermal collagen deposition [15].Retinol (vitamin A, all-trans-retinol) is much less described in the literature than tretinoin as a substance used in the peel procedure.Retinol was proven to be efective in reducing wrinkles, loss of skin elasticity, and pigmentation; nonetheless, the evidence is weak [16][17][18][19][20].
Te aim of our study was to determine the efcacy of using a 4% retinol solution product containing novel TGFβ activators and antioxidants in improving the condition of the skin in patients with facial skin issues caused by aging and skin disorders.

Study Design.
Te study was conducted between August 2022 and December 2022.It was approved by the Independent Bioethics Committee for Scientifc Research at the Medical University of Gdansk, Poland (NKBBN/618/ 2022).Prior to the study, each patient supplied written informed consent.In total, 15 healthy women were enrolled according to inclusion and exclusion criteria and qualifed into three groups conforming to facial skin problems as follows: (i) Group S: 5 patients with mature skin with frst signs of aging (such as fne lines and wrinkles and dry and dehydrated skin) (ii) Group P: 5 patients with superfcial and deep hyperpigmentation (such as melasma and sun spots) (iii) Group NSS: 5 patients with postinfammatory hyperpigmentation, atrophic acne scars, and enlarged pores.
Patients from each group were subjected to chemical peel sessions according to the same protocol, indicated by the producer.Each patient had three peel treatment series, with a 30-day gap between each session.Troughout the research, they were required to utilize the same daily maintenance and photoprotection cosmetic items provided by the creator.Before the treatment series and before each peel session during the treatment day, patients were examined using Multi Skin Test MC 1000 Courage + Khazaka, evaluating hydration, elasticity, and skin pigmentation; Observ 520x and dermatoscope.Te last examination with the measurement above parameters was performed 30 days after the last peel procedure.In total, the patients were examined 5 times.
Before the study, patients were asked, with the option to answer yes or no, regarding the occurrence of several subjective facial skin symptoms among others: itching, burning, roughness, redness, exfoliation, and tightness.After the peel treatment series, they subjectively assessed the achieved improvement, according to the symptoms: redness, skin tone, dehydration skin pigmentation, dryness, wrinkles, structures, sebaceous gland activity, scar, and overall improvement.Patients assessed the reduction of the abovementioned symptoms as an answer: yes; slightly; no.

Inclusion Criteria
(i) Women with I-III Fitzpatrick skin type including features such as frst signs of aging, mature skin, shallow discoloration, postinfammatory discoloration, acne scars, dilated sebaceous glands, and uneven texture.(ii) Age: 35-50 years (iii) Sex: women (iv) Good health condition (v) Signed informed consent.

Exclusion Criteria
(i) Pregnancy (ii) Breastfeeding period (iii) Open wounds, violation of the continuity of the epidermis (iv) Hypersensitivity to any of the preparation ingredients (v) Acute and chronic infectious processes, e.g., tuberculosis, active herpes simplex virus infection (vi) Use of isotretinoin in the last 6 months (vii) Epilepsy (viii) Emotional instability (ix) Facial surgery in the last 6 months (x) Tendency to keloids (xi) Occurrence of skin diseases: rosacea, acne tarda, atopic dermatitis, contact dermatitis, and seborrheic dermatitis.

Procedure of Chemical
Peeling.Te chemical peeling procedure was performed in the Department of Dermatology, according to the producer's protocol.Te area of the procedure was well ventilated, and patients' eyes were covered during the peeling procedure to avoid irritation.At frst, make-up removal of the skin was made after which the skin was treated with a degreasing agent, Retix.C PRE-PEEL.Subsequently, one ampoule of Retix.C BOOSTER serum was applied over the face surface and left until complete absorption.Next, the contents of the vial of Retix.C Retinol TGF Activator were spread with a brush over the entire surface of the face.Te solution was left on the skin for 6 hours, after which it was washed of with lukewarm water.

Dermatologic Terapy
After the procedure patients were asked to use daily skin care: Ultra Repair Moisturizer and SPF50+ cream in the morning and Retimodeling serum for oily and Anti-Aging Face Cream Retix C for dry skin in the evening.
2.5.Products.Te chemical peeling procedure was performed using two products.Firstly, an antioxidant product Retix C BOOSTER Serum was applied on the facial skin; subsequently, patients were treated with Retix C Retinol TGF Activator, containing 4% retinol and TGF activators.

Statistical Analysis.
Completed surveys were downloaded for statistical analysis.Te collected data were analyzed in Statistica 12.0 software.Te Shapiro-Wilk W test was used to check whether the quantitative variable came from a normally distributed population.Te Leven (Brown-Forsythe) test was used to test the hypothesis of equal variances.Te signifcance of diferences between more than two groups was tested by F (ANOVA) or Kruskal-Wallis test.In case of statistically signifcant differences between the groups, post hoc tests were used (Tukey's test for F, Dunn's test for Kruskal-Wallis).Te signifcance of diferences between more than two in the linked variable model was checked by repeated measures analysis of variance or Friedman's test.Chi-square tests were used for qualitative variables (using Yates correction for cell counts below 10, checking Cochran conditions, and Fisher's exact test, respectively).A p value equal to or less than 0.05 was considered statistically signifcant.

Results
A total of 15 patients (all females), with age range 37-50 (mean age: 41) completed the study.Subjective evaluation, according to the survey data collected during the last peel session, showed a good efcacy of the therapy.Most of the patients reported improvement in the reduction of the facial skin: uneven tone, skin pigmentation, dehydration, dryness, uneven structure, and sebaceous gland activity.All patients reported overall improvement after the peeling procedure.No improvement was reported in the following symptoms: redness, liquidation of pigmentations, wrinkle reduction and liquidation, as well as scar reduction and liquidation (Table 1).
An objective evaluation revealed a signifcant clinical improvement in pigmentation overall in study groups, specifcally in patients with mature skin with frst signs of aging (Table 2) (Figure 1).Elasticity parameters were shown to increase during treatment, with a greater efect occurring in patients with mature skin with frst signs of aging and patients with postinfammatory hyperpigmentation, atrophic acne scars, and enlarged pores; however, none of the diferences were statistically signifcant (Table 3) (Figure 2).Furthermore, there was no improvement noticed referring to the hydration of the skin during the therapy (Table 1) (Figure 3).Improvement of skin condition as photo documentation in groups P, S, and NSS is shown in Figures 4-6, respectively.

Adverse Events.
Tere were no serious adverse events in patients during the study.Only one patient reported mild itching during skin exfoliation, and the same patient noticed dryness of the skin after the procedure.

Discussion
Vitamin A was frst found in about 1906 and synthesized in 1947 [21].Since then, research on the use of retinol in medical indications has been underway.Its efectiveness in terms of antiaging was frst suggested by Kang S. et al., who demonstrated, similarly to retinoic acid, the administration of all-trans-retinol to normal human skin efects on epidermal thickening and increases the expression of CRABP II and CRBP mRNAs and proteins.Te scientists also noted that, in contrast to tretinoin, retinol caused very little erythema and discomfort.
Literature regarding the efcacy of retinol used as a peeling substance seems to be poor yet promising.Wojcik et al. investigated the efects of 2% retinol peel on skin lipids in the face and neck area.Te authors observed a signifcant growth of sebum on the left cheek, nose, chin, and left side of the neck.Te methods used were similar to ours.Te 2% retinol chemical peel was applied 3 times in 3 weeks of intervals in 21 women.However, there was no other parameter investigated besides sebum secretion [22].In another study by Sadick et al., 24 patients (photodamage group, n � 14 (with an acne subgroup, n � 5]; melasma group, n � 5; skin of color, n � 5) were treated with 3% retinol peel in 6 weeks of intervals and a series of 2-4 peels.Nonetheless, only the photodamage group was evaluated.Dermatologist clinical grading of fne lines, wrinkles, pore size, laxity, mottled pigmentation, lack of clarity/radiance, and overall photodamage was signifcantly improved (P < 0.05) In comparison to our study, in that research efcacy was evaluated mainly based on the dermatologist; thus, the results might be not fully objective [23].
Te study group of our research consisted of patients only with Fitzpatrick I-III skin type.Furthermore, there is currently no research investigating the efectiveness of retinol chemical peel in the dark skin population.However, the efectiveness of other chemical peel substances has been described in the literature.Te systematic review and metaanalysis of comparative trials by Dorgham et al. reported satisfactory results of glycolic acid (GA) and trichloroacetic acid peel as a melasma treatment in dark skin patients, with better efects and safety profle of GA.Undoubtedly, there is a need to perform research evaluating the efectiveness of retinol peel in the population with Fitzpatrick IV-VI skin type [24].
According to the literature, transforming growth factor, beta (TGF-beta), plays a crucial role in regulating the extracellular matrix (ECM), which becomes remodeled in the aging skin, including photoaging.TGF-beta regulates the diferentiation of the epidermal layer by inhibiting the growth of epithelial cells.On the other hand, it promotes fbroblast proliferation in connective tissue and thus has an impact on type I procollagen production by TGF-beta type II receptor activation [25][26][27].Terefore, well-balanced TGF-β Dermatologic Terapy    Dermatologic Terapy signaling is important to keep the skin in a healthy condition and support the maintenance of a young appearance.Given the cited literature data, studied chemical peel products contain novel substances which are the activators of the transforming growth factor, beta.To the best of our knowledge, it is the frst study describing the efects of chemical peel containing the transforming growth factor, beta activators.Nevertheless, as a component of topical cosmeceuticals, they have been under investigation for a couple of years.Te current literature suggests that topically applied growth factors in clinical contexts have a rejuvenation efect on the skin; notwithstanding, there is a high need for further studies confrming the thesis [28][29][30].
Structures providing structural integrity and resilience in the skin are called collagen fbrils, mainly composed of type I collagen (COL1) [31].Due to natural aging and photoaging,  Dermatologic Terapy the content of skin COL1 declines with the passage of time.Tis in turn results in the loss of elasticity, thinning, and wrinkling of the skin which contributes to an elderly appearance.Tus, restoration of collagen 1 synthesis and ultimately its content in the skin could contribute to enhancing the skin's resistance and frmness.It has been proven that retinol has a positive efect on metabolism of collagen which impacts the skin aging [17,32].Topical retinol restores Type I collagen production in the photoaged forearm skin within four weeks.Interestingly, Varani et al. postulated that, with increasing age, MMP levels are increased which can mediate a destruction of collagen [17].Retinol, on the other hand, inhibits UV induction of MMP and stimulates collagen synthesis in the skin.Hence, by MMP inhibition, topical retinol has the ability to suppress degradation of newly synthesized procollagen and as a result leads to enhanced procollagen expression.Taking into consideration the use of retinol in skin care products for daily use, confrmed in numerous studies, it can be concluded that retinol is efective in treating aging and photoaging of the skin [9].What is more, retinol has been also shown to have skin brightening properties.In 2003, Yoshimura et al. investigated the efciency and adverse efects of 10% all-trans retinol (ROL) gel for improvement of skin hyperpigmentation [33].Te authors enrolled 21 Japanese patients with hyperpigmented lesions on the face.After more than 10 weeks of a follow-up, 18 patients were analyzed.Sixteen out of eighteen analyzed subjects showed improvement of pigmentation after an average treatment period of 11.3 weeks.Pigmentation was almost totally eliminated in 6 of the treated subjects.Reported side efects included erythema and scaling.It was concluded by the authors that ROL has the ability to improve skin hyperpigmentation as effectively as tretinoin when used at high concentration.Other studies, however using retinol in combination, also confrmed the abovementioned thesis [34,35].

Limitations
Te main limitations of the study were the small sample size and carrying out of the research in a single center, which was caused by the preliminary character of the research.However, the number of study participants remains equal to another comparable research.[36] Another limitation was the assessment of the efects no later than 3 weeks after the last peeling treatment and lack of the control group; therefore, there is a need to evaluate longer periods in further studies.

Conclusions
Combined peel therapy of 4% retinol serum product containing novel TGFβ activators and antioxidants showed good efcacy in reducing facial pigmentation of the skin.Despite the fact that there were no statistically signifcant results in improvement of skin hydration and elasticity, subjectively all patients showed high satisfaction with the procedure.Te study revealed that the described method is simple to use, has low cost, is with rare adverse events, and has well tolerability.Te next step will be a randomized control trial with an expanded study group to investigate the efcacy of therapy on discussed facial skin parameters.

Figure 1 :
Figure1: Skin pigmentation reduction during the therapy, with a statistically signifcant diference in group S. P: patients with Superfcial and deep hyperpigmentation, S: patients with mature skin with frst signs of aging, NSS: patients with postinfammatory hyperpigmentation, atrophic acne scars, and enlarged pores.

Figure 2 :Figure 3 :
Figure 2: Skin elasticity changes during the therapy.P: patients with Superfcial and deep hyperpigmentation, S: patients with mature skin with frst signs of aging, NSS: patients with postinfammatory hyperpigmentation, atrophic acne scars, and enlarged pores.

Figure 4 :
Figure 4: Photo of the patient from group P (patients with superfcial and deep hyperpigmentation) before (left site) and after (right site) the peel treatment.

Figure 5 :
Figure 5: Photo of the patient from group S (patients with mature skin with frst signs of aging) before (left site) and after (right site) the peel treatment.

Table 1 :
Patients' subjective assessment of facial skin symptom changes.

Table 1 :
Continued.: patients with superfcial and deep hyperpigmentation, S: patients with mature skin with frst signs of aging, NSS: patients with postinfammatory hyperpigmentation, atrophic acne scars, and enlarged pores. P

Table 2 :
Skin pigmentation reduction during the therapy, with a statistically signifcant diference in group S.