Anti-psoriatic effects of indigo naturalis on the proliferation and differentiation of keratinocytes with indirubin as the active component

Department of Traditional Chinese Medicine, Center for Traditional Chinese Medicine, Chang Gung Memorial Hospital, Taoyuan, Taiwan.
Journal of dermatological science (Impact Factor: 3.42). 04/2009; 54(3):168-74. DOI: 10.1016/j.jdermsci.2009.02.007
Source: PubMed


Indigo naturalis has shown efficacy in treating psoriasis in our previous clinical studies.
To investigate the potential effect of indigo naturalis on regulating keratinocyte proliferation and differentiation.
Skin samples from six patients were analyzed for proliferating cell nuclear antigen (PCNA) and involucrin expression by immunohistochemical staining. In addition, indigo naturalis extracts from 10 to 500 microg/ml were added to cultured keratinocytes and cell viability determined. Real-time RT-PCR, Western blotting analysis and indirect immunofluorescent labeling were used to investigate the messenger (m)RNA and protein expressions of PCNA and involucrin. Finally, high-performance liquid chromatography (HPLC) was used to identify major components of indigo naturalis and their in vitro effects compared.
Immunohistochemical results demonstrated decreased PCNA and increased involucrin in psoriatic lesions after indigo naturalis treatment. Cultured keratinocytes decreased after indigo naturalis treatment, while G(0)/G(1) arrest was observed to dose-dependently increase. Staining revealed decreased PCNA-stained nuclei and increased cytosolic involucrin in treated keratinocytes. Decreased PCNA and increased involucrin at both the mRNA and protein levels were confirmed. Both major components, indirubin and indigo, could cause G(0)/G(1) phase arrest; however, only indirubin modulated the expressions of PCNA and involucrin similar to indigo naturalis.
Together, these findings indicate that the anti-psoriatic effects of indigo naturalis are mediated, at least in part, by modulating the proliferation and differentiation of keratinocytes, with indirubin as the major active component.

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    • "Thus, the search for less toxic efficacious compounds remains a worldwide challenge (McKellar et al., 2007). Indigo naturalis (Apiaceae), an herbal medicine, has been used to treat skin disorders including eczema, aphtha, furuncles, and psoriasis (Lin et al., 2009). Indirubin, which is isolated from Indigo naturalis, is a purple 3,2-bisindole and a stable isomer of indigo. "
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    ABSTRACT: Ethnopharmacological relevance: Indirubin, isolated from Indigo naturalis (Apiaceae) is a purple 3,2- bisindole and a stable isomer of indigo. Although it is known to have anti-inflammatory activities, its mechanism of action has not been elucidated. Materials and methods: Seven-week-old female BALB/c mice were sensitized with 1-chloro-2,4-dinitrobenzene (DNCB) to induce skin inflammation. Hematoxylin and eosin staining was performed to assess epidermal and dermal hyperplasia, which were determined by measuring the thicknesses of the epidermis and dermis, respectively. We also evaluated serum immunoglobulin E (IgE) levels and cytokines production, such as tumor necrosis factor (TNF)-alpha, interleukin (IL)-4, 6 and Interferon (IFN)-gamma. In addition, we investigated nuclear factor (NF)-κB, IκB-α and mitogen-activated protein (MAP) kinase activities for verifying the molecular mechanism of inflammation. Results: Indirubin treatment suppressed skin inflammation in DNCB-exposed mice. The skin lesions were significantly thinner in the Indirubin-treated group than in untreated controls, and the hyperkeratosis disappeared. Indirubin reduced the total serum IgE level and cytokines production. In addition, it normalized NF-κB, IκB-α and MAP kinase expression. Conclusions: Indirubin might be a useful treatment for allergic contact dermatitis via regulating the co-expression of T helper (Th) 1 and 2 cell-mediated immune responses.
    Journal of ethnopharmacology 11/2012; 145(1). DOI:10.1016/j.jep.2012.10.055 · 3.00 Impact Factor
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    • "Indirubin, as demonstrated by chromatographic fingerprint of indigo naturalis in previous study [8], was the major component of indigo naturalis that exerted the anti-proliferating effect on keratinocytes. To determine the effect of indirubin on the gene expression of CDC25B, keratinocytes were treated with 0, 1, 5, 10, and 20 mM indirubin for 24 h and then analyzed by RT/real-time PCR and western blotting. "
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    ABSTRACT: Topical indigo naturalis ointment is clinically proved to be an effective therapy for plaque-type psoriasis. Indirubin, as the active component of indigo naturalis, inhibits cell proliferation of epidermal keratinocytes. However, the detailed underlying mechanism is not fully understood. To further investigate the anti-proliferating effects of indigo naturalis and indirubin on epidermal keratinocytes. The decreased expression of CDC25B in indigo naturalis- or indirubin-treated epidermal keratinocytes, as revealed by cDNA microarray analysis, was studied. The CDC25B expression was examined under different serum concentrations and compared between primary and immortalized keratinocytes. The activation of EGFR and the effect of EGF on the cell proliferation and CDC25B expression were also investigated in epidermal keratinocytes. RT/real-time PCR and western blot method were used to analyze the CDC25B expression at the mRNA and protein levels, respectively. Indigo naturalis and indirubin were confirmed to down-regulate CDC25B expression significantly at both the mRNA and protein levels. The growth-dependent expression of CDC25B was demonstrated by the increased expression in serum-stimulated and immortalized keratinocytes. The activation of EGF receptor, known to be highly expressed in psoriatic lesions, was inhibited by indigo naturalis or indirubin. The cell proliferation and CDC25B expression of epidermal keratinocytes were induced by EGF alone and confirmed to be inhibited by indigo naturalis or indirubin. Except being a common therapeutic target in various cancers, CDC25B also plays an important role in the hyper-proliferation of epidermal keratinocytes which can be suppressed by anti-psoriatic drug indigo naturalis and its component, indirubin.
    Journal of dermatological science 06/2012; 67(2):140-6. DOI:10.1016/j.jdermsci.2012.05.008 · 3.42 Impact Factor
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    • "Indigo naturalis is a dark-blue powder from the leaves of indigo-bearing plants. It inhibits proliferation, promotes differentiation of epidermal keratinocytes, inhibits neutrophil proinflammatory responses and suppresses TNF- induced vascular cell adhesion molecule 1 expression in endothelial cells thereby exerting an antipsoriatic effect (Lin et al, 2009). Six patients with psoriasis who had been treated for 4 years with indigo naturalis ointment or systemic Chinese herbs and whose skin lesions had responded well whereas the nails remained unchanged were treated twice daily with 1 to 2 drops of indigo naturalis oil extract onto the nail plate, fold and hyponychium. "

    Psoriasis, 02/2012; , ISBN: 978-953-307-878-6
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