Revealing the origin of postsclerotherapy hyperpigmentation: Identification of melanin and hemosiderin as causative pigments in a histopathological study

Introduction

Postsclerotherapy hyperpigmentation (PSH) is the most common side effect of sclerotherapy, with a prevalence that varies widely across studies, depending on factors such as the type of treated vein or the type and concentration of sclerosant used, occurring in up to 70% of treated patients. ¹ For sclerotherapy, sclerosing agents such as polidocanol, tetradecyl sulfate, hypertonic glucose, saline, or glycerin-containing agents are injected into the target veins using visual or ultrasound guidance. ¹ Inside the veins, these agents induce endothelial damage, leading to the closure of the treated vein segment. The degree of endothelial effect increases with the concentration of the sclerosing agent used.^1,2 If polidocanol is injected as a foam, it also exerts a vasospastic effect. ² This endothelial damage can render the vessel permeable or cause it to rupture, resulting in the extravasation of erythrocytes, which are subsequently broken down into hemosiderin in the dermis.^1,3

Limited literature exists on the origin and treatment of PSH. It has been hypothesized that PSH results from postinflammatory hyperpigmentation (PIH), characterized by increased melanin production and accumulation in the superficial dermis after inflammation and hemosiderin deposition in the dermis following erythrocyte extravasation.^3,4 To the best of our knowledge, only two histopathological studies have investigated this condition. In 1987, Goldman et al. analyzed histopathological samples from six patients with PSH, finding hemosiderin deposits but no increase in melanin after 6–12 months. ³ Similar results were reported in 10 samples from six patients in a study by Zhang et al. ⁵

Although hyperpigmentation is primarily a cosmetic issue, it is often stigmatized, leading to patient dissatisfaction and a desire for lightening treatments. Spontaneous resolution of PSH is common, but PSH persists for more than 6 months in approximately 7.5% of cases. ⁶ Currently, no treatment guidelines are available for persistent hyperpigmentation. Short-pulsed lasers and various topical lightening products have been suggested.^4,7–9 Small studies, case reports, and case series have indicated a favorable response of PSH to quality-switched (QS) ruby laser, picosecond alexandrite laser, and intense pulsed light in combination with radiofrequency, whereas the pulsed dye laser has shown a poor response.^7,9–11 Based on the previous histopathological studies, which identified hemosiderin but found no increase in melanin in PSH, topical bleaching creams would likely be ineffective. This is because these creams exclusively target melanin synthesis, without affecting hemosiderin. Furthermore, their effect is limited to the skin’s surface and does not extend into the deeper dermis. To develop an effective treatment algorithm for PSH, a thorough understanding of the origins and location of pigmentation is essential. This histopathological study examined the origins of in early pigmentation within 3 months after sclerotherapy versus pigmentation persisting longer than 3 months after sclerotherapy in PSH.

Materials and methods

Nineteen patients with PSH following treatment with polidocanol for telangiectasias, reticular varicose veins, and lateral branches who presented at the Department of Dermatology of the University Hospital of Bern, Switzerland, between February 2023 and June 2024 were included in the study. Written informed consent was obtained from all patients. This study was approved by the local ethics committee (Cantonal Ethical Committee of Bern, Switzerland; project number 2023-00909).

Demographic data (age and sex), clinical parameters (skin type, type of treated vein, duration and localization of pigmentation), and polidocanol concentrations were collected. In addition, 20 skin biopsies (3–4 mm punch biopsy) from these 19 patients were performed by a dermatologist and analyzed by a dermatopathologist without knowledge about duration of hyperpigmentation. In one patient 2 biopsies were taken, one <3 months after sclerotherapy and one > 3 months after sclerotherapy. For histopathological evaluation, hematoxylin-eosin, Fontana-Masson, and Prussian blue staining combined together with CD68 (PGM1 clone) immunostaining, were used. The location was determined based on penetration depth according to grading I–V: I (junction zone), II (papillary dermis), III (papillary-reticular dermis), IV (reticular dermis), and V (subcutis). The quantity of hemosiderin, as well as epidermal and dermal melanin, was quantified semi-quantitatively as none (−), scant (+/−), mild (+), prominent (++), or abundant (+++). The presence and type of inflammatory infiltrate (lymphocytes, macrophages, melanocytes, eosinophilic granulocytes) were evaluated using the same grading scale used for pigment. Additionally, the presence of affected vessels (damaged or thrombosed) was assessed. Furthermore, in Prussian blue/ CD68 staining, the evaluation focused on whether hemosiderin was phagocytosed by macrophages.

Results

Type, localization, and quantity of pigment

Hemosiderin deposits were detected in all 20 samples, located from the reticular dermis to the subcutis and ranged in quantity from scant to abundant. In patients with DoP ≤3 months and DoP >3 months respectively, hemosiderin was found mild in the papillary-reticular dermis (1 and 1 patient), in the reticular dermis (1 and 2 patients) and the subcutis (2 and 0 patients), prominent in the reticular dermis (1 and 4 patients) and the subcutis (1 and 2 patients) and abundant in the reticular dermis (1 and 0 patients) and the subcutis (3 and 1 patients).

Increased epidermal melanin was found in one biopsy from a patient with a DoP of ≤3 months, and borderline elevation was observed in two patients with a DoP of >3 months, all with skin type II (Figure 2(a)–(d)). Dermal melanin was categorized as mild in three samples with DoP ≤3 months and scant in 4 samples with DoP >3 months (6 patients had skin type II and one had skin type III). In three of the seven patients with dermal melanin, epidermal pigmentation was slightly decreased. (Figure 3(a)–(d)) In all CD68 stains, hemosiderin was found to be primarily phagocytosed by macrophages, independent of the duration of pigmentation (Figure 4).OPEN IN VIEWER

Figure 2.

Histopathological findings of patient 1: (a) Prussian blue staining magnification ×20; iron/ hemosiderin deposit in the reticular dermis and subcutis, (b) detail with magnification ×200, (c) Masson-Fontana stain magnification ×20; melanin in the papillary dermis and normal melanin in the epidermis, (d) detail with magnification ×200.

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Figure 3.

Histopathological findings of patient 2: (a) Prussian blue staining magnification ×20; iron/ hemosiderin deposit in the reticular dermis and subcutis, (b) detail with magnification ×200, (c) Masson-Fontana stain magnification ×20; melanin in the papillary dermis and decrease of melanin in the epidermis, (d) detail with magnification ×200.

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Figure 4.

Combination of Prussian blue staining and CD68 immunostaining, demonstrating iron/hemosiderin inside the macrophages.

Discussion

In this study we demonstrate that besides hemosiderin, epidermal and dermal PIH is detected in some PSH biopsies. Nevertheless, hemosiderin remains the main causative pigment in PSH, as demonstrated by our results, which show its presence in all biopsies, consistent with previous findings.^3,5 Hemosiderin was primarily located in the reticular dermis and subcutis, and in only two biopsies, the deeper parts of the papillary and reticular dermis were involved. This is explainable by the origin of hemosiderin, which results from the destruction of erythrocytes following the extravasation of ruptured vessels after sclerotherapy. ³ Goldmann et al. described that PSH following the treatment of larger telangiectasias is usually transient due to its deeper location in the dermis and greater access to macrophages. ³ Consistent with this observation, our study found a trend of greater subcutis involvement in PSH cases with a DoP ≤3 months, suggesting faster pigment elimination in the subcutis. However, in our study, the type of vein treated did not influence the duration of PSH. In all biopsies, hemosiderin was exclusively phagocytosed by macrophages. This aligns with the healing of bruises, where erythrophagocytosis occurs within 3 days, and hemosiderin is found in wounds from day 8, decreasing from day 19, with very low levels after 1 month.^12,13 Regarding the fact that the first biopsy was performed >1 month after treatment, we can hypothesize two possibilities for pigment persistence. First, there is ongoing reflux in the same areaor a lack of ability of macrophages to eliminate the pigment, similar to that observed in tattoos and in iron stains after paravasal iron supplementation. ¹⁴ The absence of affected vessels in older PSH cases and the localized nature of the pigmentation without spreading support the idea that there is no continuous source of bleeding. Additionally, if there were an ongoing source of bleeding, we would expect to find erythrocytes in the histological analysis. In our analysis in 7/10 of the older PSH no hemorrhage or even erythrocytes could been found.

PIH, one of the most common acquired hypermelanoses, is more prevalent in individuals with darker skin tones and is typically triggered by UV radiation. ¹⁵ The pathogenesis of PIH is complex. Multiple cell types, cytokines, growth factors, and cellular interactions have been suggested to be involved in the pathogenesis. ¹⁶ Inflammatory mediators such as leukotrienes (LTC4 and LTD4), prostaglandins E2 and D2, and thromboxane could promote melanogenesis by increasing tyrosinase-related proteins. Furthermore, interleukins 1 and 6, as well as TNF-α and reactive oxygen species, have been suggested to have melanocyte-stimulating properties.^16,17 In histopathology, PIH can demonstrate increased epidermal melanin, dermal melanin, or melanophages, which may be accompanied by a perivascular lymphocytic infiltrate and a decrease in epidermal melanin. ¹⁶ In our findings, melanin was detected in the papillary dermis of seven patients, three of whom exhibited a typical decrease in epidermal melanin. Furthermore, three patients presented with epidermal PIH characterized by increased epidermal melanin. These findings contradict the assumptions of previous histological studies that hemosiderin is the sole causative agent of PSH, highlighting their implications for treatment options. Both types of PIH were observed in biopsies, regardless of whether the DoP was ≤3 or >3 months. Regarding the reason why PIH has not been detected in previous studies, one possibility is that our cohort exclusively used polidocanol foam at concentrations of 0.5%–3%, whereas Goldmann’s study included polidocanol in only three patients at a concentration of 0.25%–0.75%. The type of sclerosing agent influences the risk of irritation and PSH; for polidocanol, both risks increase with concentration and when foam is used compared to liquid.^1,2,18 Furthermore, the sample sizes in previous studies were smaller than ours, which may have led to missed cases of PIH. In addition to the classical mechanism of PIH induced by inflammation after sclerotherapy, iron deposits in tissues can also stimulate melanogenesis. Several studies have described the upregulation of melanogenesis by iron.^19–22 Furthermore, iron has been suspected to play a role in the development of dermal melanocytes in stasis dermatitis. ²⁰ As PIH can cause PSH in some patients, physicians should consider risk factors for PIH, such as darker skin type and sun exposure, before treatment. The 2014 European Guidelines for sclerotherapy recommend UV protection in the first 2 weeks after sclerotherapy. ²³ However, there is limited evidence that sun exposure and Fitzpatrick skin type influence the risk of PSH. ²⁴

Short-pulse lasers, such as picosecond and QS lasers, are the first-line treatments for PSH. These lasers are widely used for treating endogenous pigment lesions as well as for removing exogenous pigments, such as tattoos and iron deposits after parenteral supplementation. ¹⁴ The target pigment absorbs light and is destroyed through photoacoustic and photothermal effects. Smaller particles are then phagocytosed by macrophages and transported via the lymphatic system. ¹⁴ Therefore, the wavelength should be selected based on the maximum absorption spectrum of the target pigment. Hemosiderin has absorption spectra at 280 nm, 320–350 nm, and 400 nm. ²⁵ Therefore, lasers with shorter wavelengths would be the first choice. However, as demonstrated in our study, hemosiderin is primarily located in the deep reticular dermis and sometimes even in the subcutis, making it less accessible to short wavelengths with limited penetration depth. Additionally, shorter wavelengths, such as those used in QS ruby lasers, carry a higher risk of PIH after treatment compared to longer wavelengths, such as the QS neodymium-doped yttrium aluminum garnet (Nd:YAG) 1064 nm laser. ²⁶ Given that PIH contributes to pigmentation in some patients, topical depigmenting agents used for treating epidermal PIH ²⁶ could be considered a more cost-effective treatment option, which may be effective in a small group of patients with PSH. While short-pulse lasers can effectively treat iron deposits and PIH, particularly dermal PIH, they remain the gold standard treatment of PSH. In these cases, the use of longer wavelengths, mainly 1064 nm, should be considered as the first choice, as they have a deep penetration depth, an effect on iron deposits as well as on PIH, and a smaller risk of inducing further PIH compared with short pulsed lasers with shorter wavelengths.

The limitations of our study include the predominance of patients with skin types II or III and the fact that cofactors such as sun exposure or the use of topical agents after treatment were not examined. In addition, the histopathologic examination was performed by an experienced dermatopathologist; however, no objective pigment quantification using image analysis algorithms was conducted on the stained samples, nor were they compared with skin type–matched healthy controls. Further studies with larger sample sizes should be conducted, taking these factors into account.

Conclusion

This study confirms that while hemosiderin remains the primary pigment responsible for post-sclerotherapy hyperpigmentation, epidermal as well as dermal postinflammatory hyperpigmentation can also contribute to pigmentation in some cases. Our findings challenge previous assumptions that hemosiderin is the sole causative agent in PSH and suggest that the type and treatment- and patient-specific factors such as skin type and sun exposure, may influence pigment development. Future research with larger cohorts and objective pigment quantification is needed to better understand the pathogenesis of postsclerotherapy hyperpigmentation.