A Comparative Study of Microneedling Combined with Topical 5-Fluorouracil versus Microneedling Alone in Treatment of Localized Stable Vitiligo.

CONTEXT: Vitiligo is an acquired pigmentary disorder of the skin that is characterized by circumscribed, depigmented macules, and patches. AIMS: To compare the therapeutic efficacy and safety of microneedling combined with topical 5-fluorouracil solution versus microneedling alone on stable patches of vitiligo. SETTINGS AND
DESIGN: Hospital-based interventional prospective study. METHODS AND MATERIAL: We selected 46 patients in the age group of 10-50 years having patch stability of at least 1-year duration. The patients were randomly assigned into two groups of 23 patients each. Group A was treated with microneedling followed by application of 5-fluorouracil which was available in the form of a solution (50 mg/mL) whereas Group B was treated with microneedling alone. The procedure was repeated after every 15 days up to 4 months and patient follow-up along with the grading of repigmentation was done till 6 months. The side effects were noted. STATISTICAL ANALYSIS USED: Statistical Package for Social Sciences (SPSS) software, version 25.0. The results were analyzed using the Chi-square test and t-test and P < 0.05 was considered significant.
RESULTS: At the end of 6 months, excellent response (>75% repigmentation) was reported in 35 patches (48.6%) in Group A treated with microneedling combined with 5-fluorouracil whereas only 12 patches (16.9%) showed an excellent response in Group B treated with microneedling alone.
CONCLUSIONS: Combining microneedling with 5-fluorouracil is a simple, safe, well tolerable, and effective modality than microneedling alone for treating small localized vitiligo patches. Copyright:

Introduction

Vitiligo, one of the most common cutaneous disorders of depigmentation with complex genetic, autoimmune processes, and environmental interactions continues to be a major dermatologic challenge, in spite of the availability of many therapeutic options. Despite being asymptomatic, it can have a significant impact on an individual's quality of life. They may experience significant psychosocial manifestations, including low self-esteem and depression.

It is stated that vitiligo affects 0.5–2% of the world's population[1] and has no predilection for gender and race.

The three important hypotheses proposed for its pathogenesis are autoimmune, neural, and biochemical/cytotoxic. Many studies have reported that autoimmune thyroid diseases are related to vitiligo in the pattern of hyperthyroidism or hypothyroidism.[2] The primary mode of therapy includes topical and oral immunomodulators and phototherapy. The complete therapeutic response varies according to the disease's clinical form and stability as well as the available therapeutic options.

Advancement in the understanding of the pathogenesis of vitiligo has contributed to newer therapeutic options. Surgical methods are emerging as an important solution for stable vitiligo refractory to medical therapy. These surgical modalities are based on a common principle: to transplant autologous melanocytes from a normal pigmented area to the affected depigmented skin. Various surgical techniques have been developed to date which include tissue grafts (full-thickness punch graft, split-thickness grafts, suction blister grafts) and cellular grafts (cultured melanocytes, non-cultured epidermal cellular grafts). Microneedling followed by topical application of 5-fluorouracil is a recent advancement to the treatment mode for stable vitiligo and a newer option to investigate.

Subjects and Methodology

Study design

It was a hospital-based single-blinded interventional prospective study which was conducted in the outpatient department of dermatology over 6 months from September 2019 to February 2020 after obtaining clearance from the institutional ethical committee.

Sample size

It was calculated using the sealed envelope power calculator for the binary outcome superiority trial. On the basis of previous studies,[3] it was obtained considering 48% success in the experimental group and 12% success in the control group, at a significance level of 5% and power of 80%. The sample size thus obtained was 23 patients in each group.

Study subjects

A total of 46 patients attending the outpatient department during the study period who satisfied the following inclusion criteria were enrolled in the study: age between 10 and 50 years, patch stability for at least 1-year duration, size of the patches less than 10 cm in diameter.

The patients allergic to 5-fluorouracil, patches on the mucosal membrane, having a keloidal tendency, active Koebner's phenomenon, active infection, bleeding disorder, anti-coagulant therapy, immunocompromised, age less than 10 years, size of patch more than 10 cm, were excluded from the study. The presence of prognostic factors like leukotrichia, site, and previously taken treatment were recorded. The patients were explained regarding the side effects like pain, erythema, burning, erosion, hyperpigmentation which might occur. Informed written consent was obtained from each patient.

Study intervention

The patients were randomly assigned into two groups of 23 patients in each study arm. A systematic random procedure was chosen to select the patients for both groups. Every even patient was put into Group A and the remaining patients were included in Group B. Group A with 72 patches was treated with microneedling combined with 5-fluorouracil, whereas Group B with 71 patches was treated with microneedling alone.

In Group A, under aseptic conditions, microneedling of vitiligo patches was done with 1.5 mm dermapen. It was followed by topical application of 5-fluorouracil solution available as ampoules in the strength of 50 mg per mL and again microneedling was done. Occlusive dressing was then done and kept for 1 day. Antibacterial agents were given for 5–7 days to prevent any infection. This procedure was repeated every 15 days up to 4 months. The patients were examined in every sitting for the appearance of pigmentation and complication if any. Serial photographs of the patches were taken before and after the treatment and follow-up was done for 6 months. The side effects were also monitored throughout the study period.

Similarly, in Group B, microneedling was done alone over the vitiligo patches. The rest of the procedure was the same.

Follow-up

The patients were followed-up for 2 months after the last session. Using serial photographs, the assessment was done at the baseline, and thereafter, every month. The grading of repigmentation was done by the subjective assessment score shown in [Table 1].

Table 1

Grading of repigmentation

Grades	Repigmentation	Response
G0	Absent	No response
G1	<25%	Poor
G2	25-50%	Good
G3	50-75%	Very good
G4	>75%	Excellent

Statistical analysis

After collecting information, data were compiled, tabulated, and analyzed using the SPSS software version 25.0. The parameters of mean, standard deviation, ratio, frequency, and percentage were used for analyzing descriptive statistics. Chi-square test and t-test were applied. P <0.05 was considered statistically significant. The statistical power of the sample was 80% and type 1 error of 0.05.

Results

In our study, 46 patients in the age group of 10–50 years were enrolled for the study and randomly assigned into two groups of 23 patients each. In Group A, 13 were females and 10 were males with a female to male sex ratio of 1.3:1. In Group B, 14 were females and 9 were males with a female to male ratio of 1.55:1. The maximum number of patients were in the age group of 20–40 years in both groups. The different morphological types of vitiligo were vitiligo vulgaris in 28 patients (60.86%), acrofacial in 12 patients (26.1%), and focal vitiligo in 6 patients (13.1%). Out of the 143 patches, 60 (41.9%) were on the trunk/abdomen, 51 (35.66%) were on the limbs, and 32 (22.37%) were distributed acrally. The family history of vitiligo was positive in two patients in Group A whereas 1 patient in Group B. The baseline characteristics of the patients are mentioned in Table 2.

Table 2

Baseline parameters of the studied groups

Clinical data	Group A	Group B	P
Age (mean±SD) years	29.4±8.66	29±8.90	0.87
Gender
Male	10 (43.47%)	9 (39.1%)
Female	13 (56.5%)	14 (60.86%)
Duration (mean±SD) years	5.04±1.49	4.82±1.41	0.61
Family history
Present	2 (8.69%)	1 (4.34%)
Absent	21 (91.30%)	22 (95.6%)
Type of vitiligo
Vitiligo vulgaris	15 (65.2%)	13 (56.52%)
Acrofacial	6 (26.1%)	5 (21.73%)
Focal	2 (8.69%)	5 (21.73%)
Leukotrichia
Present	3 (13.04%)	2 (8.69%)
Absent	20 (86.95%)	21 (91.30%)
Site of vitiligo patches
Trunk	29 (40.27%)	31 (43.66%)
Limbs	26 (36.11%)	25 (35.21%)
Acral	17 (23.6%)	15 (21.12%)

Treatment outcomes of the studied groups

A total of 72 and 71 vitiligo patches were treated in Group A and Group B, respectively. After 1 month of therapy, the initiation of repigmentation, i.e. Grade 1 pigmentation was noted in 44 patches (61.1%) in Group A whereas 29 patches (40.84%) in Group B [Table 3].

Table 3

After 1 month

Response on patches	Group A	Group B	P
Grade 1	44 (61.1%)	29 (40.84%)	0.003
Grade 0	28 (28.9%)	42 (59.1%)
	Total=72 patches	Total=71 patches

At the end of 6 months, a statistically significant difference between the two groups was observed (P = 0.0001), i.e., an excellent response (>75% repigmentation) was reported in 35 patches (48.6%) in Group A whereas only 12 patches (16.9%) showed an excellent response in Group B [Table 4].

Table 4

After 6 months

Response on patches	Group A	Group B	P
Grade 4	35 (48.6%)	12 (16.9%)	0.0001
Grade 3	27 (37.5%)	20 (28.1%)
Grade 2	08 (11.11)	31 (43.7%)
Grade 1	02 (2.8%)	08 (11.26%)
	Total=72 patches	Total=71 patches

The side effects such as pain, erythema, itching were almost similar in both the groups as shown in Table 5. There was no statistically significant difference in the occurrence of the side effects between the two groups. Ulceration was seen in four patients (17.39%) in Group A whereas two patients (8.69%) in Group B. Hyperpigmentation was seen in 11 patients (47.82%) in Group A and 6 (26.1%) patients in Group B.

Table 5

Side effects

	Group A (total=23 patients)	Group B (total=23 patients)
Pain	18 (78.26%)	20 (86.9%)
Erythema	15 (65.21%)	17 (73.9%)
Itching	10 (43.47%)	09 (39.13%)
Ulceration	04 (17.39%)	02 (8.69%)
Hyperpigmentation	11 (47.82%)	06 (26.1%)

In our study, the excellent response was seen in the patches over the trunk [Figure 1a and b] followed by the limbs [Figure 2a and b]. None of the acral patches showed an excellent response in any of the groups [Figure 5].

Figure 1

(a) At baseline, (b) Grade 4 (>75% repigmentation) at 6 months treated with microneedling combined with 5-fluorouracil

Figure 2

(a) At baseline, (b) Grade 4 response in a vitiligo patch over the limb at 6 months treated with microneedling combined with 5-fluorouracil

Figure 5

Showing excellent response with respect to location of patch

Discussion

In 1983, Tsuji and Hamada[4] first introduced the application of 5-fluorouracil after therapeutic wounding. Various techniques were used for therapeutic wounding in different studies like derma- abrasion,[5] laser ablation, cryosurgery, and needling.[6]

Various theories have been given which explain the mechanism of action of 5-fluorouracil with needling. As a result of wounding, an inflammatory reaction takes place due to which various inflammatory mediators like leukotrienes release[7] that lead to stimulation of the melanocytes. Also, there occurs local edema that increases the intercellular spaces of the basal layer of the epidermis.[78] Therefore, during re-epithelization, active melanocytes migrate toward the achromic areas.

Our study constituted patients in the age group of 10–50 years. The females outnumbered the males in our study (56.52% in Group A and 60.86% in Group B). Similar studies conducted by George et al.[9] noted female predominance. A clinico-epidemiological study done by Gandhi et al.[10] also reported a higher number of females, reflecting the treatment-seeking behavior in the female patients due to social stigma.

We observed that most of our patients were suffering from Vitiligo vulgaris followed by acrofacial and focal vitiligo. A study conducted by Shashikiran et al.[6] also observed Vitiligo vulgaris as the common morphological type (58% patients).

In our study, Grade 1 repigmentation started at the end of the 4th week, i.e., 44 patches (61.1%) in Group A and 29 patches (40.84%) in Group B which are seen as brown, perifollicular macular pigmentation at the hairy areas which are gradually seen as coalescing during further sessions. In non-hairy areas, pigmentation started from the periphery indicating the stimulation of peripheral epidermal melanocytes during re-epithelization. Garg et al.[11] in their study noted the initiation of pigmentation within 2 months in 50% of the patches.

At the end of 6 months, an excellent response was noted in 35 patches (48.6%) in Group A which is statistically significant in comparison to Group B where only 12 patches (16.9%) showed an excellent response. Similar results were obtained in the study done by Shashikiran et al.,[6] where 49% of the patches showed >75% repigmentation. Ghiya et al.[12] showed an excellent response in 60% of the children treated with combined 5-fluorouracil and dermaroller.

In our study, the lesions over the limbs and trunk had the best response whereas the acrofacial lesions [Figure 3a and b], lesions associated with leukotrichia [Figure 4a and b], showed poor response to therapy.

Figure 3

(a) At baseline, (b) Grade 3 response (50–75% repigmentation) on the patch over the face at 6 months treated with microneedling alone

Figure 4

(a) At baseline. (b)Poor response to treatment over vitiligo patch associated with leukotrichia treated with combined microneedling and 5-fluorouracil

The side effects recorded in our study were minimal pain and burning sensation during the procedure. Also, the lesion developed mild erythema which used to subside in 1 or 2 days in comparison to the other study by Sethi et al.[5] where therapeutic wounding was done by dermabraider causing intense pain and erythema. Also, there was serous discharge and pseudomembrane formation seen as side effects in their study. In our study, hyperpigmentation was noted in 12 patients in Group A whereas it was noted in 7 patients in Group B which persisted for 3 months.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.