Reports of itchy skin have been increasing in recent years, with pruritus being an uncomfortable sensation on the skin. Suppressing itch is indispensable as it not only lessens the disagreeable feeling but also hinders further scratching, which is generally a sign of multiple common diseases, such as thyroid disorders and atopic dermatitis. It is widely accepted that histamine is the main cause of allergic and itching reactions [1, 7]. In medical clinics, anti-histamines are the most widely used medications to relieve itching [24]. A multitude of studies have demonstrated that natural plant extracts, such as osthole [25], quercetin [12] and red ginseng extract [13] possess the ability to suppress histamine-induced itch. This is the initial investigation to suggest that SL, a kind of biosurfactant, can have a counter-pruritic action in C57BL/6J mice against scratching behavior caused by histamine, as well as diminish skin harm caused by scratching.
Histamine is identified when it binds to histamine receptors, initiating the downstream histamine pathway and subsequently activating a number of intracellular pathways, such as the adenylate cyclase and PLC pathways [12]. The increase in intracellular calcium resulting from this phenomenon can result in the upregulated levels of of pro-inflammatory cytokines and may even induce cell death. It has been established in prior studies that an overabundance of intracellular calcium can lead to certain negative pharmacological effects associated with the receptor, the most common of which is pruritus [26, 27]. This study, especially results shown in Fig. 2, for the first time reported that SL has a dose-dependent capability to impede histamine-induced calcium influx.
Histamine action is widely recognized as being mediated by four distinct receptors, namely H1-H4 [25, 28]. Of these, histamine H1 and H4 receptors are essential components in the communication of histamine-associated itch signals between epidermal cells and peripheral sensory nerve endings [7, 11, 29]. Despite the use of antagonists of histamine H1 and H4 receptors, histamine-caused scratching behavior cannot be fully inhibited. Previous studies have demonstrated that the histamine-induced scratching behavior is nearly completely suppressed when both histamine H1 and H4 receptor antagonists are associated with the transmission process of the itch signaling to the central nervous system [25, 29]. Itching caused by specific receptor activators cannot be relieved by other receptor blockers. For example, thiopamine, a H3/H4 receptor antagonist, has been found to be ineffective in reducing scratching behavior caused by HTMT [30]. Extensive research has been conducted in recent years to elucidate the effects of H1 and H4 receptors on histamine-caused calcium influx in both in vitro and in vivo settings [9, 10]. It has been demonstrated by various researchers that in human epidermal keratinocytes, histamine receptor blockers play an inhibitory role in histamine-induced calcium ion influx [12, 19, 27]. To ascertain if the inhibitory effect is associated with histamine receptors, we conducted experiments to evaluate if SL could further inhibit histamine-induced calcium ion influx when histamine receptor blockers are present. The observations in this study suggested that SL impeded the rise in [Ca2+]i caused by histamine, and this inhibition is not solely dependent on a single histamine receptor. In addition, activation of H1 or H4 receptors by H1/H4 receptor antagonists leads to an increase in intracellular calcium, which can be suppressed by different concentrations of SL. It is conceivable that SL may not act as a direct selective agent of H1 or H4 receptors, but instead, SL plays a partial role via the interaction with H1 and H4 receptors to inhibit the signaling cascade that follows.
Following histamine binding to its corresponding receptors, the subsequent PLC, IP3, and DAG pathways are activated, resulting in the opening of specific calcium ion channels and allowing for increased calcium influx, in turn leading to the production of action potentials and the increase of neuro-sensitivity [18]. IP3 attaches to a particular IP3 receptor (IP3R) and is involved in the regulation of calcium ion movement. It has been observed that IP3 and DAG stimulate the corresponding PKC, further linked to the conveyance of calcium ions [18, 19]. We employed an RT-PCR analysis to examine the effect of SL on genes that had been stimulated by histamine. Our research showed that histamine significantly increased the mRNA levels of PLCγ1, IP3R, and PKCα, and SL reduced their expression to a certain degree.
TRPV1 has been well-documented as a virtual ion channel within the downstream signaling process of histamine H1 and H4 receptors [14, 31]. Research evidence has indicated that TRPV1 is involved in the calcium response activated by histamine and the conduction of itch signals when the H1/H4 receptor is stimulated [10, 14]. We hypothesized that SL controls histamine-induced itching by controlling the activity of TRPV1, which was validated by demonstrating that SL was able to prevent the rise in [Ca2+]i caused by capsaicin (Fig. 5). The findings of the present study demonstrate that TRPV1 may be one of the critical factors in the suppression of histamine-induced reactions by SL. It is unclear why this outcome did not increase or decrease with the amount of the dose. A high dose of SL (25 µg/mL) was not able to further suppress the influx of capsaicin. In fact, it was in line with our previous data in the Section 3.3 that the degree of SL's inhibition of the H1/H4 receptor agonist-caused [Ca2+]i elevation was not affected by the dose. As a result, additional verification is required to explain the mechanism. Molecular docking results suggest that SL has a strong affinity for the TRPV1 protein complex and could act as an inhibitor by binding to the protein.
Although our data demonstrated the antipruritic effect of SL with keratinocyte and mice models, there are several limitations that were not covered in the current study. The mechanism of itching is complex and we need to investigate more molecular pathways. Apart from histamine-dependent itch, there is also the presence of itch that is not related to histamine. Chloroquine, an anti-malaria drug, has proved to induce histamine-independent pruritus as a side effect [13]. Therefore, for a comprehensive understanding of SL's antipruritic effect, further examination on chloroquine-induced itch is required.