## 4 Discussion  
Chronic inflammation and the overproduction of ROS are precursors of metabolic diseases, such as cancer, atherosclerosis, and diabetes [31]. Controlling ROS synthesis and inflammatory processes constitutes a promising strategy to prevent metabolic diseases. Therefore, extensive work has previously focused on the development of efficient and novel antioxidants and anti-inflammatory molecules. Various natural antioxidants have been characterized, and several anti-inflammatory agents, such as aspirin, are widely used [32]. However, the development of effective anti-inflammatory agents is still necessary. Although appropriate amounts of ROS and NO play essential roles in signaling, cellular homeostasis, and cardioprotection [10, 33], ROS overexpression causes inflammation and tissue damage [34]. We assessed the inflammatory and antioxidant effects of plant extracts by measuring the efficacy of LPS on the level of NO and ROS, markers of inflammation and oxidative stress, respectively [35, 36]. Among the 100 plant extracts tested, _Sorseris hirsuta_ had the greatest effect on the inhibition of NO levels (Supplemental Table S1). _Sorseris hirsuta_ has not been studied well, and the anti-inflammatory and antioxidant effects of _Sorseris hirsuta_ are unknown. Thus, we investigated the effect of several concentrations of SHE on LPS-induced NO levels and cell viability in RAW 264.7 cells. As expected, consistent with the normal dose of SHE, the other concentrations of SHE significantly inhibited NO levels without causing cell cytotoxicity in LPS-induced RAW 264.7 cells. The production of NO and prostaglandin E2 (PGE2) by iNOS and COX-2, respectively, contribute to inflammation [37]. Although SHE did not inhibit COX-2 expression in LPS-stimulated cells, it was shown to significantly inhibit iNOS expression. The results of specifically suppressing iNOS expression suggested that SHE exhibits anti-inflammatory properties. Cranberry is well known for its inhibitory action on NO production. [PERSON] et al. suggested that cranberry polyphenol and volatile extracts induced a 46-fold decrease in NO production in LPS-activated RAW 264.7 macrophages [38]. In the present study, SHE caused 38% inhibition of NO production induced by LPS in RAW 264.7 cells. Therefore, SHE and cranberry have very similar inhibitory activities on NO production. Moreover, the inhibitory effect of SHE on the increase in _IL-1\(\beta\)_ mRNA expressed by LPS treatment in RAW 264.7 cells was significantly different from that of the group not treated with SHE. This  
Figure 6: Effects of _Sorseris hirsuta_ extract (SHE) on lipopolysaccharide-induced _interleukin-1 beta (IL-1\(\beta\))_ mRNA expression in RAW 264.7 cells. The cells were first treated with SHE 1 h, and then LPS and SHE were co-treated and left for 24 h. Subsequent procedures were carried out as described in the Materials and Methods section. # \(p\) < 0.001 and *** \(p\) < 0.001.  
is of particular interest as inflammatory cytokines can cause systemic inflammation [39] and, particularly, IL-1\(\beta\) causes a fibrosis reaction and an acute inflammatory reaction in the lungs [40, 41]. Macrophages recognize LPS through TLR4, a membrane receptor that activates an intracellular signaling cascade. Consequently, IKK phosphorylates the regulator of NF-\(\kappa\)B, signaling IxB, that undergoes proteasome degradation. NF-\(\kappa\)B then freely moves to the nucleus to stimulate transcription [42]. NF-\(\kappa\)B plays a pivotal role in the development of inflammation by activating the transcription of inflammatory genes. Therefore, NF-\(\kappa\)B signaling molecules constitute promising targets to prevent inflammation. Because iNOS and IL-1\(\beta\) are target genes of NF-\(\kappa\)B [43], we assumed that the inhibitory effect of SHE on LPS-induced _iNOS_ and _IL-1\(\beta\)_ mRNA expressions may affect the NF-\(\kappa\)B signaling pathway. Our results clearly displayed that SHE blocked the phosphorylation of IKK and p65 induced by LPS in RAW 264.7 cells.