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Anti-VEGF therapy has been used as the anti-angiogenic agent in cancer treatment. However, the treatment often couples with severe complications. Complementary natural products that offer similar anti-angiogenic potency with less side effects are sought as alternatives. Study showed that Malaysian Tualang Honey (MTH) can inhibit inflammation-induced vascular hyperpermeability in vitro and in vivo, but the effect on the angiogenesis process remains unclear. Thus, this study aims to determine the anti-angiogenic effects of MTH. Effects of MTH ranging from 0.3% to 0.9% on VEGF-induced human umbilical vein endothelial cells (HUVEC) angiogenesis was determined by proliferation, migration and tube-formation assays. Matrix metalloproteinase-2 (MMP-2) secretion from HUVEC and VEGF production from MCF7 cancer cells in response to MTH were also quantified by using ELISA kits. Suramin, an angio-suppressive agent was used as positive control. MTH significantly suppressed HUVEC proliferation (from 155% proliferation rate to 54%); migration (~ 50% inhibition rate) and tube-formation (69% reduction) induced by VEGF. These findings were explained by the significant reduction (p < 0.05) of VEGF-induced MMP-2 secretion in HUVEC with 39% suppression exhibited by 0.9% of MTH. MTH also significantly (p < 0.05) reduced VEGF secretion (19% reduction compared to control) in MCF7 breast cancer cells. Our findings suggest that the anti-angiogenic effects of MTH mainly targets endothelial cell through inhibition cell proliferation, migration and tube formation capacity, via suppression of MMP-2 secretion by the endothelial cells. However, MTH has less prominent effect on suppressing VEGF secretion by the MCF7 cancer cells.

Background: Clinacanthus nutans (Burm. f.) Lindau. has traditionally been using in South East Asia countries to manage cancer. However, scientific evidence is generally lacking to support this traditional claim. This study aims to investigate the in vitro, ex-vivo and in vivo effects of C. nutans extracts on angiogenesis. Methods: C. nutans leaves was extracted with 50–100% ethanol or deionised water at 1% (w/v). Human umbilical veins endothelial cell (HUVEC) proliferation was examined using MTT assay. The in vitro anti-angiogenic effects of C. nutans were assessed using wound scratch, tube formation and transwell migration assays. The VEGF levels secreted by human oral squamous cell carcinoma (HSC-4) cell and HUVEC permeability were also measured. Besides, the rat aortic ring and chick embryo chorioallantoic membrane (CAM) assays, representing ex vivo and in vivo models, respectively, were performed. Results: The MTT assay revealed that water extract of C. nutans leaves exhibited the highest activity, compared to the ethanol extracts. Therefore, the water extract was chosen for subsequent experiments. C. nutans leaf extract significantly suppressed endothelial cell proliferation and migration in both absence and presence of VEGF. However, the water extract failed to suppress HUVEC transmigration, differentiation and permeability. C. nutans water extract also did not suppress HSC-4 cell-induced VEGF production. Importantly, C. nutans water extract significantly abolished the sprouting of vessels in aortic rings as well as in chick embryo CAM. Conclusion: In conclusion, these findings reveal potential anti-angiogenic effects of C. nutans, providing new evidence for its potential application as an anti-angiogenic agent.

Inflammation is the immediate response to tissue damage or harmful stimuli. Though inconvenient, its role is significant and important as the protective and physiological response of our body. It directly sets the stage for tissue repair particularly increasing endothelial permeability which then contributes to the healing process. However, in some cases inflammation may progress out of control causing various inflammatory diseases. Neurogenic inflammation is a sub-set of inflammation and is characterized by an increase in neuronal chemical mediators such as Substance P (SP). In this study, we investigated the involvement of SP in enhancing endothelial permeability on HUVECs monolayer. Neurogenic inflammation was induced through the administration of SP (1 nM to 100 nM) on HUVECs monolayer inserts, and incubated with varying short (10, 20 and 30 minutes) and longer (6, 12 and 24 hours) time-points. FITC-Dextran were finally added to cell culture inserts for 5 minutes to let the fluorescence molecule pass through the gaps. Endothelial permeability is directly proportional with extravasation of FITC-Dextran, determined by fluorescence intensity reading. Based on our data, there were no significant differences between control group (non-treated cell) and the different concentrations of SP at different time-points. Our current findings suggest that SP was unable to increase the endothelial permeability on HUVECs monolayer inflammatory experimental model. Experiments that use this model to mimic vascular inflammation in laboratory settings may require further elucidation in the future.