Evaluating pulmonary toxicity of Shuang–Huang–Lian in vitro and in vivo
ETHNOPHARMAOCOLOGICAL RELEVANCE: Shuang-Huang-Lian (SHL) is a traditional Chinese formula and has been used for the treatment of respiratory tract infections by inhalation. However, the pulmonary toxicity via inhalation is largely uninvestigated. To evaluate the pulmonary toxicity of SHL following in vivo intratracheal spray to rats and in vitro exposures to A549 and Calu-3 cells. Calu-3 and A549 cells were exposed to SHL, chlorogenic acid, baicalin and forsythin solutions and in vitro cytotoxicity was evaluated using an MTT assay, whilst rats were subjected to intratracheal administration of SHL solutions and in vivo toxicity was indicated by assaying the LDH activity and total protein content in bronchoalveolar lavage fluid (BALF) and observing the histopathologic changes of the lungs. Secretion of inflammatory mediators, including IL-6, IL-8 and TNF-α, in cell culture media and BALF was quantified by ELISA. The MTT cell viability data revealed the presence of minor toxicity to Calu-3 or A549 cells following exposure to SHL and its major ingredients for 24h or 48 h. However, the cell cultural media showed no sign of inflammatory responses. The in vivo results showed that exposures to SHL at doses of up to 50mg/kg did not significantly increase the total protein content, the LDH activity and the concentrations of IL-6, IL-8 and TNF-α in BALF. However, although intratracheal sprayed SHL at doses of up to 6 mg/kg for histopathologic study and up to 25mg/kg for cell counts showed no sign of adverse effects, inhaled SHL at elevated doses appeared to induce alveolar fusion in the lung and significant increases in the cell number of monocytes and granulocytes in the BALF. The results demonstrated that the pulmonary safety of inhaled SHL was dependent on the administered dose. Inhalation therapy of SHL may be safely used when the inhaled dose was properly controlled.