## 3 Results
### Effect on mRNA Expression of Pro-Inflammatory Cytokines and Antioxidant Genes  
Consistent with its secretion level, _Tnf-\(\alpha\)_ mRNA expression was significantly reduced by high doses of CHA in the sustained delivery model (Figure 4). Neither a low dose nor  
Figure 2: Effect of different delivery models on iNOS and COX-2 expression. (**a**) Cells were treated with a serial concentration of CHA with regular instantaneous delivery. Protein expression of iNOS was determined. (**b**) Low doses and high doses of CHA were delivered to a culture medium using a pipette or syringe pump. Representative images of iNOS and COX-2 protein expression are shown. (**c**) Protein expression levels were quantified. (**d**) mRNA levels of _iNos_ and _Cox-2_ were measured. Bars without a common letter indicate significant differences (\(p<0.05\)). CHA: chlorogenic acid; LPS: lipopolysaccharideacharide; iNOS: inducible nitric oxide synthase; COX: cyclooxygenase; L: low dose administered via pipette; LS: low dose administered via syringe pump; H: high dose administered via pipette; HS: high dose administered via syringe pump.  
a high dose of CHA administered via instant delivery showed significant alterations in mRNA expression.  
Although sustained delivery of a high dose of CHA suppressed the elevation in _Il-1b_ mRNA, unlike the secretion level of IL-1\(\beta\), sustained delivery of a low dose and instantaneous delivery of a high dose of CHA did not exert an inhibitory effect on _Il-1\(\beta\)  
Figure 4: Effect of different delivery models on mRNA expression of pro-inflammatory cytokines and antioxidant genes. Cells were treated with LPS (100 ng/mL) and CHA for 6 h. (**a**) mRNA levels of _Tnf-a_, _Il-1\(\beta\)_, _Il-10_, and _Map-1_ were determined 6 h after treatment. (**b**) mRNA levels of _Hmax-1_, _Sod_, Cat, and Gpx-1 were determined 6 h after treatment. Bars without a common letter indicate significant differences (_p_ < 0.05). CHA: chlorogenic acid; LPS: lipopolysaccharideacharide; TNF: tumor necrosis factor; IL: interleukin; Hmox: Hem oxygenase; Sod: superoxide dismutase; Mcp: monocyte chemoattractant protein; Cat: catalase; Gpx: glutathione peroxidase; L: low dose administered via pipette; LS: low dose administered via syringe pump; H: high dose administered via pipette; HS: fmapk.  
Figure 3: Effect of different delivery models on the secretion of TNF-\(\alpha\) and IL-1\(\beta\). Cells were treated with LPS (100 ng/mL) and CHA for 24 h. (**a**) TNF-\(\alpha\) concentration in the conditioned medium and (**b**) IL-1\(\beta\) concentration in the conditioned medium were determined. Bars without a common letter indicate significant differences (_p_ < 0.05). CHA: chlorogenic acid; LPS: lipopolysaccharideacharide; TNF: tumor necrosis factor; IL-interleukin; L: low dose administered via pipette; LS: low dose administered via syringe pump; H: high dose administered via pipette; HS: high dose administered via syringe pump.  
mRNA expression. This suggests that CHA modulates the secretion of IL-1\(\beta\) through both transcriptional and post-transcriptional regulation. In addition, mRNA levels of the other two cytokines, ll-10 and monocyte chemoattractant protein-1 (Mcp-1), were not altered by either LPS or CHA.  
LPS can produce intracellular reactive oxygen species and induce oxidative stress in RAW 264.7 cells [22]. LPS suppressed the mRNA expression of the antioxidant genes glutathione peroxidase 1 (_Gpx1_) and catalase (_Cat_). Sustained delivery of low doses of CHA showed a significant elevation in _Gpx1_ mRNA levels, and instantaneous delivery of high doses of CHA showed a significant elevation in _Cat_ mRNA expression. However, no differences were observed between different CHA treatments. Heme oxygenase 1 (_Hmox1_) mRNA expression was elevated by LPS exposure and further augmented by CHA treatment. While high doses of CHA showed a higher ability to induce the expression of _Hmox1_ than low doses, sustained delivery enhanced _Hmox1_ expression at low doses but not at high doses.