Neuroprotective effect of Caulerpa lentillifera extracts against lipopolysaccharide-induced neuroinflammation in raw 264.7 macrophages and SK-N-SH neuroblastoma cells

Current existing therapies against Alzheimer’s disease (AD) relieve only the clinical symptoms but fail to halt the neurodegenerative disease progression. This raises the need for alternative neuroprotective agents that can prevent AD. Caulerpa lentillifera J. Agardh appears to be useful against the neuroinflammation-driven AD given its antioxidant and anti-inflammation properties. To date, no study has reported C. lentillifera’s neuroprotective potential against neuroinflammation. Hence, this study screened four C. lentillifera extracts for their anti-oxidative, acetylcholinesterase (AChE) inhibitory and anti-inflammatory potentials using the high-performance thin layer chromatography (HPTLC). To this end, freeze-dried C. lentillifera was extracted using sequential liquid-liquid partitioning with methanol, hexane, chloroform and ethyl acetate. HPTLC analyses found all four C. lentillifera extracts to exhibit varying degrees of free radical scavenging activities. Amongst the extracts, the C. lentillifera chloroform extract (CLCE) exhibited the highest free radical scavenging activity (6.135 μg/ 1 mg GAE). However, none of the extracts exhibited detectable AChE inhibitory activity. All C. lentillifera extracts exhibited varying degrees of COX-1 inhibitory activities, with the C. lentillifera ethyl acetate extract (CLEAE) demonstrated the highest COX-1 inhibitory activity (9,613.62 μg/ 1 mg SAE). Qualitative test for phenols indicated the presence of phenolic compounds in all C. lentillifera extracts. The HPTLC-based FeCl₃ assay confirmed and found CLCE to contain the highest total phenolic content (40.79 μg/ 1 mg GAE). NMR phytochemical profiling revealed that the C. lentillifera methanolic extract (CLME) contained sugar, sterols and amino acids; the C. lentillifera hexane extract (CLHE) contained fatty acids, sterols and olefinic compounds, CLCE contained palmitic acids, polysaccharides and amino acids; CLEAE contained polysaccharides, terpenoids and phenolic compounds. The present study went on to examine the neuroprotective potentials of the four C. lentillifera extracts and unravel the underlying mechanisms through oxidative stress, inflammatory and apoptotic markers in LPS-induced RAW 264.7 macrophages and SK-N-SH neuroblastoma cells. With regards to LPS-induced RAW 264.7 macrophages, all four C. lentillifera extracts significantly reduced ROS production, with CLCE and CLHE being the two most potent extracts that significantly reduced ROS production (p < 0.05) by 79.8% and 63.8%, respectively when compared to controlLPS. All four C. lentillifera extracts also significantly reduced NO production, with CLCE being the most potent extract that significantly reduced NO production (p < 0.05) by 80.1% when compared to controlLPS. Nevertheless, only CLCE resulted in significant reduction of Caspase-3 activity (p < 0.05; -29.6%) when compared to controlLPS. Whilst only CLCE and CLHE significantly upregulated NOS2 and NRF2 expression, only CLCE significantly upregulated NOS3 expression. With regards to LPS-induced SK-N-SH neuroblastoma cells, all four C. lentillifera extracts significantly reduced ROS production, with CLHE emerged as the most potent extract that significantly reduced ROS production (p < 0.05) by 72.7% when compared to controlLPS. All four C. lentillifera extracts also significantly reduced Caspase activity, with CLHE and CLCE emerged as the two most potent extracts that significantly reduced Caspase-3 activity (p < 0.05) by 48.9% and 45.3%, respectively. However, none of the extracts resulted in significant changes to NOS2, NOS3 and NRF2 expressions. The present findings implied the potentials of the C. lentillifera extracts (especially CLCE and CLHE) as natural neuroprotective agents against the neuroinflammation-driven AD.