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Topical Treatment of Hair Loss with Formononetin by Modulating Apoptosis

Introduction

Hair loss is characterized by abnormal hair cycles. Three phases, anagen (growth), catagen (regression) and telogen (resting) organize the hair cycle and so control the hair development [1]. These dermatologic processes frequently co-occur perifollicular lymphocytic infiltrate-affected anagen, induction of apoptosis-regression catagen and prolonged telogen. Most of all, dystrophic hair follicles affected by the induction of apoptosis in the catagen phase exhibit reduced size and inconsistent pigmentation, resulting in a disabled state of production of hair fiber [2]. These imbalances between the follicle growth and the induction of apoptosis result in hair loss [3].

Abstract

Formononetin is one of the main components of red clover plants and its role on hair regrowth against hair loss has not been established yet. In the present study, we assessed the potential effects of formononetin on alopecia, along with impaired hair cycles by induction of apoptosis-regression.

Depilated C57BL/6 mice were used for monitoring the hair cycles. Formononetin (1 and 100 µM) was topically treated to the dorsal skin for 14 days.

Topical formononetin treatment induced miniaturized hair follicles to recover to normal sizes. Tapering hair shaft began to grow newly, emerging from the hair follicles by formononetin. In addition, formononetin inhibited the activation of caspase-8 and decreased the procaspase-9 expression.

As a result of formononetin treatment, anti-apoptotic Bcl-2 was up-regulated, whereas pro-apoptotic Bax and p53 were down-regulated, resulting in a decrease of caspase-3 activation.

Formononetin showed the obvious inhibition of apoptosis under terminal deoxynucleotidyl transferase dUTP nick end labeling staining thereafter. Taken together, our findings demonstrate that formononetin exerted the hair regrowth effect on hair loss, in which the underlying mechanisms were associated with Fas/Fas L-induced caspase activation, thus inhibiting apoptosis.

Results

For 2 weeks, the depilated backs remained hairless in the depilated group. In contrast, mice treated with formononetin showed markedly hair regrowth. By treating formononetin, the area of depilated back skin was covered with black fur. Furthermore, FNT 1 and 100 µM group showed black skin color, while the depilated mice still contained pink or gray skin color. These changes were appeared in histomorphometry. Hair shafts affected by hair loss were broken and fragmented in the depilated group (l ” Fig. 1A). There were miniaturized hair follicles with no hair fiber (l ” Fig. 1 B). Treatment with formononetin recovered not only the lengths of hair shafts, but also the sizes of hair follicles. Wellstraight hair shafts with numerous hair fibers got through the surface of the epidermis after treatment with formononetin. In addition, the size of the dystrophic hair follicles returned to normal levels.

Topical formononetin treatment inhibited the activation of casepase-8 and procaspase-9 (l” Fig. 2A). Additionally, formononetin treatment decreased the caspase-3-positive cells and caspase-3 protein levels, while those still were remained in catagen-like depilated hair follicles (l” Fig. 3). In mice treated with formononetin, the protein expressions of Bax and p53 were decreased, while the expression of Bcl-2, but not of Bcl-xL, was increased compared to depilated mice (l” Fig. 2 B, C). These results revealed an increase of Bcl-2/Bax ratio by formononetin.

Numerous cell death pattern in catagen phase of hair cycle were seen in depilated skin region. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells were located in the epithelial cells and surrounded the dermal papilla of hair follicles (l” Fig. 4). Formononetin decreased the number of TUNEL-positive cells at the both of treated groups.

Authors : Mi Hye Kim1, You Yeon Choi1, Ji Eun Lee1, Kyuseok Kim2, Woong Mo Yang1
Affiliations : 1 College of Korean Medicine and Institute of Korean Medicine, Kyung Hee University, Seoul, Korea
2 Department of Ophthalmology, Otorhinolaryngology and Dermatology of Korean Medicine, College of Korean Medicine, Kyung Hee University, Seoul, Republic of Korea

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