Introduction
The treatment of acne in post-adolescent females often fails1,2, although there are a number of treatments with a relative high degree of effectiveness. Some patients suffer from severe and repetitive acne, and others have widespread occurrences of acne such as on the chest and back. Inflammation, scars, and post-inflammatory hyperpigmentation impair their appearance. The treatment of acne is based on two separate concepts: 1) anti-symptomatic treatments such as the use of chemical peeling, lasers, and antibiotics3 and 2) inhibitory treatments such as the use of oral retinoids and anti-androgen therapies (retinoids also have symptomatic effects). Systemic treatments are especially needed for treating cases with severe, repetitive, or the widespread type of acne throughout the body.
Hormonal anti-androgenic treatments could be most effective for post-adolescent females to inhibit sebum production and acne formation, though they can induce side effects such as menstrual irregularity. Cyproterone acetate, a synthetic progestin, is the first steroidal anti-androgen agent widely used in the treatment of advanced prostatic carcinoma and severe acne. However, it was reported that long-term use of cyproterone acetate can induce severe hepatocellular dysfunction and potential carcinogenesis4,5, and the sale of the agent was discontinued in Japan in1999. It is still widely used in single-use form or in combination with ethinyl estradiol, a contraceptive (e.g. Diane-35), for severe acne in many western countries. Flutamide is a non-steroidal anti-androgenic agent, recognized as a competitive agonist of the androgen receptor and a beneficial compound for the treatment of prostate cancer, benign prostatic hyperplasia, and hirsutism. Despite its proven validity, hepatotoxicity due to this agent has been reported and the use of this agent is restricted6. In addition, flutamide inducing hepatotoxicity is reported to be higher in Asians than in Caucasians7. For this reason, in Japan, monthly blood tests for checking liver function are required during the use of flutamide. Spironolactone is well known as a diuretic and has been used for over 20 years as a representative antagonist of androgen in the treatment of acne and hirsutism8,9. There have been several randomized studies evaluating the efficacy of these agents in the treatment of hirsutism10-13. Serious adverse drug-related experiences and the discontinuation of therapy have been uncommon in spironolactone therapy14, although gynecomastia15 and menstrual irregularities as side effects are frequently seen accompanying high-dose use of spironolactone. Other adverse side effects, such as lethargy, fatigue, dizziness, or headache, were reported8,9,16,17.
As is well known, hormonal actions, serum hormone levels, and side effects of hormonal agents frequently differ among races or populations. As far as the Oriental population is concerned, spironolactone appears to be safest for treating acne among the three agents discussed above with anti-androgenic action. The present study was performed in order to assess the therapeutic and side effects of oral spironolactone in Japanese patients.
Patients and Methods
Spironolactone was administered orally to 139 Japanese patients (ages 15 to 46 years, average 26.0 ± 6.8; 116 female and 23 male) suffering from acne at Ritz Medical Clinic. Informed consent approved by IRB was obtained from each patient or legal guardian. The uses of other therapies for acne were not permitted for the duration of the study. A 20-week course of therapy was designed; the initial dose was 200 mg/day for the first 8 weeks. After that, the doses were progressively decreased by 50 mg every 4 weeks to 150 mg/day, 100 mg/day, and 50 mg/day (Fig.1).
The effectiveness of this treatment was evaluated from the clinical results of 64 patients (all female) who completed the 20-week treatment. A digital camera was used to take clinical photographs before, during, and after treatment. The therapeutic results were graded according to the Consensus Conference on Acne Classification18. Evaluations of improvement were undertaken before and after treatment by two experienced plastic surgeons who did not perform this treatment. The investigators assessed each patient from his or her photographs using a 3-point scale (2 = excellent improvement, 1 = good improvement, and 0 = poor improvement).
In order to find related adverse events, we tested the blood of the patients and collected before-and-after data from 25 patients. Alanine transferase (ALT), aspartate transferase (AST), blood urea nitrogen (BUN), creatinine (Cr), Na+, K+, Cl-, luteinizing hormone (LH), dehydroepiandrostenedione sulfate (DHEAS), sex hormone binding globulin (SHBG), total testosterone (T), and free T in serum were measured, although not all of the data were measured from all patients due to the shortage of blood collected from some patients.

Statistical Analysis
Data were reported as mean ± standard error. The nonparametric Mann-Whitney U-test was used to compare clinical and hormonal data from before and after treatment

Results
All 64 patients evaluated exhibited clinical improvement (Table 1). Thirty-four patients (53.1%) showed fewer inflammatory spots (grade 2; Figs. 2, 3 and 4). Thirty patients (46.9%) showed a decrease in new acne papules (grade 1).
On the subject of side effects and complications, about 80% of 116 female patients suffered from menstrual irregularities. Five patients had no menstrual bleeding during the first 3 months of treatment, and four had severe menstrual irregularities. Both of these were therefore given an intramuscular injection of estrogen and progesterone to induce menstrual bleeding.
Among the 23 male patients, 3 (13%) showed gynecomastia after 4 to 8 weeks of treatment (all 3 patients were between 19 and 21 years old). Therefore, the administration of spironolactone was discontinued to all male patients.
Other adverse side effects that had previously been reported such as lethargy, fatigue, dizziness, or headache were not observed in this trial. Changes in urinary frequency were occasionally observed, however these did not seem to reduce the patients’ quality of life. Other side effects occurred in only a few patients: drug eruptions were seen in three of 139 patients and edema in the lower extremities was also seen in three.
We analyzed the changes in serum hormones, laboratory data, and blood levels of electrolytes resulting from oral spironolactone treatment (Fig. 5). Regarding testosterone (T) values, only a small number of patients showed a high total T or free T level before treatment, and a number of patients showed a low serum level of total T or free T. There were no particular trends in total T or free T before and after treatment, but some patients with high total T or free T before treatment tended to normalize their value after the treatment. Similarly, patients with low total T or free T showed a tendency to elevate their hormonal levels. The values of LH, DHEAS, and SHBG of almost all patients were within normal limits.
Indices of liver functions (ALT and AST) and those of kidney functions (BUN and Cre) were not affected by oral spironolactone. Unexpectedly, Na+ and K+ were also not significantly affected by this urinary agent. Only Cl- showed a significant decrease (p<0.05) compared with the pretreatment value, however almost all values were within normal limits.

Discussion
A key in treating acne, which almost always recurs, is to perform both a preventive treatment as well as a curative one. The results showed that oral spironolactone is very effective for treating acne in Japanese females without any serious side effects other than menstrual irregularities.
There were no significant abnormalities in the pretreatment data of serum TST and free TST in Japanese patients suffering from severe acne. However, in this trial, all female patients exhibited clinical improvement in their acne after at least 4 months (usually 2 months was sufficient) administration of oral spironolactone without any other combined treatments. Thus, the results strongly suggest that androgen signals affect acne in almost all patients and factors other than serum hormonal levels such as the expression level of androgen receptors in the sebaceous glands have a critical influence on sebum production and subsequent new acne formation. The hypothesis may explain why some acne patients do not respond to oral contraceptive treatments. Oral contraceptives can reduce gonadotropins such as LH and thereafter reduce T production and serum level of T, but cannot block androgen signals at the level of the androgen receptors. Thus, it is suggested that oral spironolactone is advantageous for treating acne compared with oral contraceptives. Gradually decreasing spironolactone dosage can contribute to preventing a recurrence of acne formation after the treatment. Recurrences of acne occasionally occurred several months after cessation of the administration of oral spironolactone.
Some patients who also suffered from seborrheic dermatitis on their faces showed improvement after the administration of oral spironolactone. This probably results from the inhibitory action of spironolactone for sebum production by blocking androgen signals at the receptor level. Indeed, almost all patients reported reduced sebum discharge as soon as 2 weeks after starting the oral spironolactone.
The mechanism of menstrual irregularities caused by oral spironolactone is not clearly understood. After the start of oral spironolactone administration, the serum levels of T or dehydrotestosterone (DHT) could be acutely elevated because androgen receptors are competitively blocked by spironolactone. The excessive T could then be transformed into estradiol, which can induce endometrial hyperplasia and menstrual irregularities. Gynecomastia can be induced in men for the same reason15. As the 3 male patients with gynecomastia in this study were between 19 and 22 years old, it is suggested that this age range is most susceptible to suffer from conversion from T to estradiol because of the high serum level of T during this period. Oral treatment with isotretinoin can be one important alternative for male patients with severe acne19,20.
Most patients in our trial were satisfied with the clinical results in spite of the menstrual irregularities because they had been suffering from repetitive acne for a long time and had lived a stressful life caused by disfigurement. For 9 of 64 (14%) patients, a combined injection of estrogen and progesterone was performed for an effective induction of normal menstruation, which was usually observed within 10 to 14 days after the injection. Combined administration of contraceptives can be useful for preventing possible pregnancy during the treatment.
Other side effects such as itching and allergic eruptions occurred in only a few patients. There were no abnormal blood levels of electrolytes and laboratory data such as ALT, AST, BUN, Cr, Na+, and K+ after administration of the series of oral spironolactone. Only Cl- showed a significant but minor decrease.
In summary, this study demonstrated that oral spironolactone is quite effective for any type of acne in Asian females and that this therapy is a good option, especially for repetitive, severe, resistant, or widespread types of acne in spite of its side effects such as menstrual irregularities. Other additional therapies might be helpful but not necessary for inhibiting new outbreaks of acne. Male patients require an alternative treatment such as oral isotretinoin. Oral spironolactone has been used as a diuretic for a long time and its safety has already been established for Asian people, although two other hormonal agonists of acne therapy, cyproterone acetate and flutamide, showed liver dysfunction in Asians. Menstrual disturbance is common as an adverse effect, but a compensatory hormonal injection is available for inducing menstrual regularity.

References
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Legends
Figure 1. 　Protocol of oral spironolactone administration.

Figure 2. 27-year-old female with acne vulgaris from the jawline to the neck who did not improve by repeated AHA peeling and oral antibiotics. (a) Before treatment. (b) After 20 weeks of oral spironolactone (grade2, fewer inflammatory spots).

Figure 3. 31-year-old female with acne vulgaris and seborrheic dermatitis on the whole face. (a) Before treatment. (b) After 4 months of oral spironolactone (grade 2). Sebum discharge was markedly reduced and seborrheic dermatitis and acne were both improved.

Figure 4. 28-year-old female with acne vulgaris and severe sebum discharge. (a) Before treatment. (b) After 4 months of oral spironolactone (grade 2).

Figure 5. Data of blood tests before and after administration of oral spironolactone: (a) Total T, (b) Free T, (c) LH, (d) DHEAS, (e) SHBG, (f) ALT, (g) AST, (h) BUN), (i) Cre, (j) Na+, (k) K+, and (l) Cl-. Increased or decreased data by more than 10 % of initial values were shown as blue or red lines, respectively. Free testosterone showed significant increases (p<0.05) and Cl- showed significant decreases (p<0.05) compared with pretreatment values, however almost all values were within normal limits. LH, DHEAS, SHBG, ALT, AST, BUN, Cre, Na+, and K+ did not show any significant differences.

Table 1. 3-point scale results. 2 = excellent improvement, 1 = good improvement, and 0 = poor improvement. The excellent feature showed fewer inflammatory spots.