Background:
In wound models, bee venom (apitoxin) has both therapeutic and toxic effects through biologically active compounds such as melittin, apamin, and phospholipase A2. At high doses, the toxic effects of apitoxin become predominant. Melittin-induced membrane damage leads to fibroblast and keratinocyte death, while the excessive inflammatory response caused by phospholipase A2 results in edema, cellular damage, and necrotic areas in the wound region. C. speciosa (Flush tree), a plant rich in flavonoids (quercetin, rutin, kaempferol derivatives), phenolic acids (gallic acid, chlorogenic acid), triterpenes, and sterols, exhibit significant anti-inflammatory properties through these phytochemical constituents. This study aimed to evaluate the treatment of bee venom-induced apoptosis with C. speciosa.

Methods:
An in vitro wound model was established in the L929 cell line using a 200 µL pipette tip, followed by the application of bee venom at a toxic dose (5 µg/mL) for 15 minutes. Cells were then treated with different concentrations of C. speciosa (1 µg/mL, 2 µg/mL, and 3 µg/mL) for 48 hours. At the end of 48 hours, the experiment was terminated, and cell viability was measured using the MTT assay. TAS, TOS, LDHA, and Klotho analyses were performed to determine oxidative stress and antioxidant capacity. In addition, Annexin-V-PI and DAPI immunofluorescence staining were used to detect apoptotic features. The data obtained were statistically analyzed.

Results:
Examination of the data revealed that cell viability decreased by 42.08% in the bee venom group (P<0.05), which was correlated with an increase in LDHA levels (P<0.05). However, C. speciosa preserved cell viability in a dose-dependent manner, and viability increased with higher concentrations. The best result was obtained in the C. speciosa 3 µg/mL treatment group, where viability increased approximately two-fold compared with the bee venom group (P<0.01). At the same time, oxidative damage induced by toxicity decreased in the C. speciosa group. Klotho levels increased approximately 2.6-fold (P<0.01). Accordingly, antioxidant enzyme activity increased, DNA damage decreased, and apoptosis was suppressed. Immunofluorescence results showed a reduction in Annexin (FITC)/PI antibody levels, indicating the suppression of apoptosis.

Conclusion:
These results demonstrate that C. speciosa has anti-apoptotic and anti-inflammatory effects against bee venom toxicity.