RhoA/ROCK Signaling Pathway Mediates Shuanghuanglian Injection-Induced Pseudo-allergic Reactions

Abstract

Background: Shuanghuanglian injection (SHLI) is a famous Chinese medicine used as an intravenous preparation for the treatment of acute respiratory tract infections. In the recent years, the immediate hypersensitivity reactions induced by SHLI have attracted broad attention. However, the mechanism involved in these reactions has not yet been elucidated. The present study aims to explore the characteristics of the immediate hypersensitivity reactions induced by SHLI and deciphers the role of the RhoA/ROCK signaling pathway in these reactions.Methods: SHLI-immunized mice or naive mice were intravenously injected (i.v.) with SHLI (600 mg/kg) once, and vascular leakage in the ears was evaluated. Passive cutaneous anaphylaxis test was conducted using sera collected from SHLI-immunized mice. Naive mice were administered (i.v.) with a single dose of 150, 300, or 600 mg/kg of SHLI, and vascular leakage, histamine release, and histopathological alterations in the ears, lungs, and intestines were tested. In vitro, human umbilical vein endothelial cell (HUVEC) monolayer was incubated with SHLI (0.05, 0.1, or 0.15 mg/mL), and the changes in endothelial permeability and cytoskeleton were observed. Western blot analysis was performed and ROCK inhibitor was employed to investigate the contribution of the RhoA/ROCK signaling pathway in SHLI-induced hypersensitivity reactions, both in HUVECs and in mice.Results: Our results indicate that SHLI was able to cause immediate dose-dependent vascular leakage, edema, and exudates in the ears, lungs, and intestines, and histamine release in mice. These were pseudo-allergic reactions, as SHLI-specific IgE was not elicited during sensitization. In addition, SHLI induced reorganization of actin cytoskeleton and disrupted the endothelial barrier. The administration of SHLI directly activated the RhoA/ROCK signaling pathway both in HUVECs and in the ears, lungs, and intestines of mice. Fasudil hydrochloride, a ROCK inhibitor, ameliorated the SHLI-induced hypersensitivity reactions in both endothelial cells and mice indicating its protective effect. SHLI-induced pseudo-allergic reactions were mediated by the activation of the RhoA/ROCK signaling pathway.Conclusion: This study presents a novel mechanism of SHLI-induced immediate hypersensitivity reactions and suggests a potential therapeutic strategy to prevent the associated adverse reactions.

LED Lights Promote Growth and Flavonoid Accumulation of Anoectochilus roxburghii and Are Linked to the Enhanced Expression of Several Related Genes

“…All parts of A. roxburghii have a long tradition use as a folk medicine to treat various ailments, with indications from one country to the other. In China, the whole plant has been used in various dosage forms such as oral liquid, spray, capsules, and its mixture to treat hepatitis, hyperliposis, tumor, chronic hepatitis B, diabetes [ 1 , 2 , 3 , 4 , 5 ] and nephritis, cystitis, myasthenia gravis, rheumatoid arthritis, febrile convulsion, cough-variant, and snakebite [ 6 ]. In Taiwan, all plant parts are frequently used as a tonic and treat hypertension, lung disease, kidney deficiency [ 7 ], cancer, and other incurable diseases [ 4 ].…”

Forsythoside A and Forsythoside B Contribute to Shuanghuanglian Injection-Induced Pseudoallergic Reactions through the RhoA/ROCK Signaling Pathway

“…According to the information available in the commercial instructions, SHLI is clinically recommended for use at a dosage of 60 mg/kg, which is approximately equivalent to 600 mg/kg in mice [42]. In the previous study, we showed that treatment with SHLI at 600 mg/kg could result in endothelial dysfunction and aberrant vascular leakage in mice [16]. Therefore, the minimum vascular leakage-responsive doses used in the present study, 50 mg/kg of forsythoside A and 100 mg/kg of forsythoside B, were approximately 7 and 12 times as high as their equivalent clinical dosages (7.92 and 8.58 mg/kg), respectively.…”

“…In our previous study, we demonstrated that SHLI could induce IgE-independent but doserelated pseudoallergic reactions by evoking histamine release, activating the RhoA/ROCK signaling pathway, and triggering increased vascular leakage [16]. This effect, at least partly, results in the hypersensitivity reactions of SHLI in the clinic.…”

“…The ears of each mouse were shredded and preserved in a tube. Then, 2 mL of formamide was added for EB extraction [15,16]. The high doses of forsythoside A, forsythoside B, and forsythoside E were all designed to be given at 100 mg/kg.…”

“…Fluorescence was determined immediately using a microplate reader (Thermo Scientific Varioskan Flash, Thermo Fisher Scientific, MA, USA) with an excitation wavelength of 490 nm and an emission wavelength of 520 nm. The fluorescent apparent permeability coefficient (P app , cm/s) was calculated according to the equation described in Reference [16].…”