*3.8. Effect of CYP on MAPK Pathway in Oxidative Stressed IEC-6 Cells*

MAPK is associated with oxidative stress, and the excessive activation of the MAPK pathway can lead to oxidative damage in cells [8]. Meanwhile, ROS overproduction could trigger the MAPK pathway when oxidative stress occurs in cells [45]. As observed in Figure 5, the ratios of p-JNK to JNK, p-ERK to ERK and p-P38 to P38 were significantly increased after H2O2 treatment compared to the control group. It suggested that oxidative damage to cells might be related to the MAPK pathway.

The JNK family is a key molecule in cellular signaling in response to various stressors induced by stressors and is involved in cellular responses to radiation, osmotic stress, temperature changes, and other stressors. After stimulation by H2O2, JNK protein was phosphorylated, while the level of intracellular JNK phosphorylation was significantly decreased after pretreatment with CYP.

ERK is a protein involved in the regulation of cell proliferation and differentiation. Upon oxidative stimulation, ERK phosphorylation in cells significantly increased. This phenomenon was effectively inhibited by CYP at 400 and 800 mg/mL.

P38 mediates inflammation and apoptosis. In the model group, the phosphorylation level of P38 was significantly higher than that of the control group, and the phosphorylation level of the polysaccharide group decreased relative to that of the model group. Unlike the JNK and ERK proteins, the phosphorylation level of the polysaccharide group did not decrease to that of the control group, which may be due to the fact that as an important mediator of apoptosis, the cells underwent apoptosis after oxidative damage stimulated by H2O2.

The ratios of all three proteins and their phosphorylated proteins were significantly decreased after pretreatment with CYP, which indicated that CYP might inhibit the activation of the MAPK pathway and thus protect IEC-6 cells.

#### **4. Conclusions**

In the present study, we isolated a polysaccharide and evaluated its protective effect against oxidative damage in cells. A water-soluble acidic polysaccharide (molecular weight = 20.89 kDa) was successfully extracted from Chinese yam. Analysis of monosaccharide composition showed that CYP primarily comprised galactose (28.57%), glucose (11.28%), and galacturonic acid (37.59%). CYP was able to increase SOD activity, inhibit MDA production, and reduce intracellular ROS production. Moreover, CYP exhibited better antioxidant activity, and it was able to alleviate the reduction of cell viability induced by oxidative stress due to H2O2 by inhibiting the activation of the MAPK pathway. It also provided a theoretical basis for the development of functional foods and clinical therapeutics against the damage caused by oxidative stress.

**Author Contributions:** Data curation, Z.L.; Formal analysis, Z.L.; Methodology, M.S.; Supervision, M.S.; Validation, W.Z.; Writing—original draft, Z.L.; Writing—review and editing, W.X., J.X., Q.Y., Y.C. and M.S. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research work was financially supported by the National Natural Science Foundation of China (81960708).

**Data Availability Statement:** Not applicable.

**Conflicts of Interest:** The authors declare no conflict of interest.
