Lactic acid sting test and capsaicin test differentially induce facial erythematous reaction in subjects with sensitive skin

Sensitivity skin (SS) is a common skin disorders, which have a various of clinical manifestation. Facial erythema is common objective symptom of SS. However, the reasons for the occurrence of erythema in sensitive skin are not fully understood.


| INTRODUC TI ON
Sensitive skin (SS), a common skin condition, is defined as occurrence of unpleased sensations, including tingling, burning, pain, and pruritus, upon stimulation which normally do not occur in normal individuals.The skin can appear normal or be accompanied by erythema. 1The prevalence of SS is 49.6% in Chinese women. 2 Over 70% of subjects with SS have erythema, while individuals with pruritus and tingling account for 61% and 39%, respectively. 3SS negatively impacts quality of patients' lives and imposes a heavy economic burden to both patients and society. 46][7][8] These cause a low sensitivity threshold in sensitive skin.Previous study had shown that lactic acid sting test (LAST) and capsaicin test (CAT) were commonly used in clinical to measure the sensory reactivity of sensitive skin. 9r previous study found that sensitive skin subjects have significant differences in reactions to LAST and CAT.LAST scores positively correlated with transepidermal water loss (TEWL) rates and could be used to identify subjects with sensitive skin characterized mainly by stinging and itching. 10Capsaicin is a natural agonist of the transient receptor potential vanilloid-1 (TRPV1). 11Activation of TRPV1 can lead to an increase in activities of adenylate cyclase and protein kinase A (PKA), enhancement of the productions of 3′,5′-cyclic adenosine monophosphate (cAMP) and nitric oxide (NO), and induction of vasodilation and neurogenic inflammation. 12,13Therefore, CAT was associated with burning sensation in sensitive skin. 14However, whether facial erythema in sensitive skin is associated with activation of TRPV1 or impaired barrier function of the stratum corneum is not clear.
In this study, we evaluated the facial erythema response of sensitive skin subjects after stimulation with lactic acid and capsaicin to analyze the possible causes of facial erythema.

| Participants
SS subjects were identified by a questionnaire (Table 1).Subjects with acne, rosacea, or glucocorticoid dependent dermatitis and other facial skin diseases were excluded from the study.A total of 197 subjects, aged 19-36 years old with a mean age of 25.25 years old, were enrolled in the study (Table 2).A written informed consent was obtained from all participants.This study was approved by the Human Research Ethics Committee (2020-L-57), and carried from November 2020 to November 2021.

| Assessment of facial erythema by physician
After washing their faces with clean water, subjects were instructed to rest in a temperature-and humidity-controlled room (temperature, 20-25°C; relative humidity, 50%-60%) for at least 30 min before assessment of erythema score, and applications of LAST and CAT.Severity of erythema was scored by physician according to a 0-to 3-scale score (0 = none, 3 = very strong). 15

| LAST
Lactic acid (purity >98%, Sigma, USA) was diluted in distilled water to make a working concentration of 10%.Following placement of filter paper soaked with 50 μL of 10% lactic acid on the folds of the nasolabial triangle, participants self-evaluated the intensity of sting sensation at 2.5 and 5 min, according to a scoring system (0 = no stimulation, 1 = mild stimulation, 2 = moderate stimulation, 3 = severe stimulation) was used.A cumulative score of ≥3 points at 2.5 and 5 min was considered as LAST positive.Otherwise, it was considered as negative. 16

| CAT
An ethanol solution containing 10% capsaicin (Macklin Co, Shanghai, China) was diluted a 0.001% solution.Capsaicin solution (50 μL) was applied to the right cheek and nasolabial fold.Within 30 s after capsaicin application, subjects reported the intensity of sensation, using a 1-5 rating scale (1 = barely perceived; 2 = slight sensation; 3 = obvious sensation; 4 = severe sensation; 5 = pain, extremely severe sensation).The CAT was positive if this sensation lasted longer than 30 s with a scores of ≥3.Otherwise, it was negative. 17

| Subject grouping
Subjects with SS were divided into four groups according to the results of LAST and CAT: LAST positive (LAST + ), both LAST and CAT positive (L + C + ), both LAST and CAT negative (L − C − ), and CAT positive (CAT + ).

| Noninvasive measurements of skin physiological parameters
Biophysical parameters of each subject were measured at the test area before and after LAST and CAT.Erythema index (EI) TA B L E 1 Sensitive skin questionnaire.
1 Cannot your face tolerate cold, hot weather, or environment? 2 Cannot your face/neck skin withstand rapid changes in temperature?
was measured using a Mexameter (MX 18; Courage and Khazaka Electronic GmbH, Köln, Germany), while the value of a* was measured using LAB (Chromameterâ-2600d, Minolta, Japan), and the tissue viability imaging (TIVI) was taken using a skin tissue viability imaging instrument (Wheels-Bridge AB, Linköping, Sweden).The values of EI and a* parameters represent the severity of skin erythema.The higher the values are, the stronger the erythema is. 18VI value represents red blood cell concentration in the cutaneous blood vessels, reflecting the function of microcirculation. 19

| Comparison of erythema scores among the five groups
Proportions of subjects without erythema (erythema score = 0 point) did no differ significantly among groups except in subjects with L + C + , which was significantly lower than the other groups (p < 0.05-0.001).In contrast, a higher portion of subjects with L + C + exhibited erythema score of 3 points in comparison to other groups although overall erythema score did not differ significantly among the five groups (Table 3).Moreover LAST + group did not differ significantly compared to the CAT + group, L − C − group and NC, and there were also no significant difference in CAT + group compared with NC and L − C − group (p > 0.05).The results indicated that subjects with LAST + , CAT + groups, L − C − group and NC cannot be distinguish by naked eye.

| Comparison baseline values of EI, a*, TIVI among the five groups
The EI and a* value of CAT + group are higher than that of LAST + group, but there is no significant difference (p > 0.05).However, CAT + and L + C + groups had higher erythema values compared with LAST + group, L − C − group, and NC through non-invasive skin detect instruments.

| Comparison of ΔEI, Δa*, and ΔTIVI after LAST and CAT
Following LAST, the changes in either EI or a* or TIVI did not differ significantly among five groups (

| DISCUSS ION
Studies have shown that the occurrence of SS was related to a variety of mechanisms.Epidermal dysfunction is one of pathogenesis of SS, and leading to a decrease in the protective function of sensory nerve endings. 20Moreover, non-neuronal transient receptor potential channels play an important role in SS, which can be activated by capsaicin. 21,22Therefore, Yokota et al. divided people with sensitive skin into three types, type I characterizes by impaired skin barrier function, type II characterizes by skin inflammation, type III is subhealth skin with nerve hypersensitivity. 23Since different individuals respond differently to different reagents, both lactic acid and capsaicin are commonly used to diagnose sensitive skin with specific symptoms and signs. 14,24r previous study showed that LAST scores negatively correlated with EI. 10 In this study, we also find that EI and a* value were significantly higher in subjects with L + C + and CAT + , and TIVI value significantly higher in subjects with L + C + .However, there was no significant difference in EI, a*, and TIVI values in LAST + group compared with normal group.Moreover, we also found that the erythema in L + C + and CAT + group were higher than that in LAST + group, although the difference was not significant.These results indicate that sensitive skin subjects with CAT + are more likely to appear erythema reactions.Then, we further compared the changes of EI, a* and TIVI values before and after lactic acid and capsaicin stimulation.Our research showed that TIVI value significantly changed after CAT in subjects with either CAT + or L + C + group, and there were no significant changes after lactate stimulation.Therefore, the CAT is more suitable for evaluating the severity of facial erythema in SS.
Our previous study also showed that LAST scores positively correlated with TEWL, and SS with stinging, itching, tightness and scaling had impaired epidermal barrier. 10LAST, the most effective method to evaluate SS, can stimulate the production of inflammatory mediators by Langerhans cells in the skin, resulting in the development of symptoms. 6Previous studies have shown that capsaicin can activate TRPV1 channel in sensory nerves, producing neurogenic inflammation, such as substance P (SP) and calcitonin gene-related peptide (CGRP), leading to vasodilation and delaying epidermal barrier recovery. 25sensitive skin individuals had higher expression of TRPV1 Mrna. 26Jerome Srour et al found that inhibition of TRPV1 channel in skin can improve facial erythema. 27ere results suggested that TRPV1 plays an important role in the development of facial erythema, our research further indicated this conclusion.
TRPV1, as a non-selective cation channel can be activated by capsaicin to release the neuropeptides from sensory nerve endings.
Moreover neuropeptides act on skin cell including microvascular cells and mast cells, leading to degranulation, vasodilation and extravasation of leukocytes. 259][30][31] TIVI values represent the concentration of red blood cells in skin microvessels, which reflect the instantaneous change of erythema. 32When assessing erythematous reactions following application of different chemical agents, we found subjects with CAT + and L + C + groups had higher TIVI values after capsaicin stimulation.
This indicates that facial erythema is also associated with vascular hyperreactivity.This result also indicates that the generation of facial erythema is related to vasodilation.
In summary, erythema in subjects with SS is a complex symptom, it may associate with the activate of TRPV1 channel, which induce neurogenic inflammation and vascular hyperreactivity.When treating SS, in addition to repairing the epidermal barrier, it is also important to reduce neurohyperresponsiveness and vascular relaxation, which are related to the generation of facial erythema in SS.
Due to few volunteers are willing to proceed skin biopsies clinically, we were unable to detect the expression of TRPV1 of SS subjects with CAT + in this study.If the condition allows, Further research on the expression of TRPV1 from facial erythema is warranted.

Table 5 )
. In contrast, stimulation with capsaicin markedly increased TIVI in subjects with either CAT + or L + C + group in comparison to NC.CAT + group was significantly higher in Δa* than NC (P < 0.05).Moreover, remarkable increases in EI were observed in subjects with L + C + than those with L − C − (P < 0.05).But the LAST + group had no significantly increase compared with CAT + group and NC after capsaicin stimulation.The results indicate that subjects with either CAT + or L + C + are more likely to occur erythema response after capsaicin stimulation, and are more sensitive to CAT.
3.5 | Correlation CAT scores with EI, a* and TIVI values Since subjects with either CAT + or L + C + displayed higher EI, a*, and TIVI values, we analyzed next the correlation of CAT scores with EI, a* and TIVI values.As shown in Figure 1, CAT scores correlated positively with EI, a*, and TIVI values.The results indicate that CAT is associated with facial erythema.