Regulation Of Macrophage Cellular Response By Clinacanthus Nutans Extracts In J774.2 Macrophage Cell Line
Loading...
Date
2019-07
Authors
Noor, Nur Mazidah Haji
Journal Title
Journal ISSN
Volume Title
Publisher
Abstract
Clinacanthus nutans, which is a medicinal plant from Acanthaceae family, is well-known as 'Belalai gajah' and Sabah Snake Grass in Malaysia. C. nutans extracts have been reported to exhibit anti-inflammatory, analgesic, anti-oxidant, anti-cancer, anti-viral and anti-bacterial properties. However, the effects of C. nutans extract towards the immune system and its immunomodulatory capabilities have not been well explored until today. Macrophage activation, which is necessary to elicit an immune response, is lacking in certain macrophage related immunodeficiency cases whereas macrophage activation occurs constitutively in certain inflammatory diseases. Thus, the immunomodulatory roles of C. nutans extracts in regulating macrophage cellular response in J774.2 mouse macrophages were investigated in this study as it can potentially provide benefits to patients with macrophage related inflammatory diseases as well as to patients with severe macrophage related immunodeficiencies. The macrophage cell line was characterised through morphological observation under inverted phase contrast microscope and expression of a few novel macrophage markers (CD11b, F4/80, CD80 and CD86) using flow cytometric analysis (FACS). The cytotoxicity effect of C. nutans leaves ethanol (EtOH), ethanol-aqueous (EtOH-AQ) and aqueous (AQ) extracts on J774.2 macrophages was determined using PrestoBlue assay. The secretion of a mixture of pro- and anti-inflammatory cytokines by the macrophages after 48 hours of extracts incubation was assessed using multiplexed cytokine bead-based assay. The phagocytic activity of 48-hours extracts-treated J774.2 macrophages was assessed using Green pHrodo-conjugated E. coli Bioparticles by both fluorescence microscopy and flow cytometry approaches. Finally, the M1 (CD80 and CD86) and M2 (CD71 and CD206) activation markers expression on the macrophages after 48 hours of extract incubation was assessed using flow cytometry technique. The results showed that J774.2 macrophages have high expression of CD11b, F4/80 and CD86 and all three extracts tested did not exhibit any significant cytotoxic effect towards J774.2 macrophages within the extract concentration range tested. All three extracts displayed their anti-inflammatory properties by reducing LPS-induced inflammatory cytokines secretion without disintegrating the phagocytic ability of the macrophages. EtOH-AQ extract has the highest potential in downregulating LPS-induced inflammatory cytokines production and CD86 expression level as well as stimulating the phagocytic function of J774.2 macrophages. However, the effect of the extracts on J774.2 macrophage polarisation could not be determined with these limited findings. Further studies should be carried out to further explore and confirm the role of C. nutans extracts in modulating macrophage activation and function.
Description
Keywords
Cellular immunity