FSHW: Pepsin hydrolyzes the peptide part extracted from sturgeon muscle through MAPK and NF-κB pathway to LPS stimulated RAW264.

Original title: FSHW: Pepsin-hydrolyzed peptides from sturgeon muscle exert anti-inflammatory effects on LPS-stimulated RAW264.

Sturgeon muscle protein produces anti-inflammatory peptides

Sturgeon peptides have strong anti-inflammatory and antioxidant activities

Sturgeon peptide exerts anti-inflammatory effects in RAW264.

Inflammation is the body's defense response to various inflammatory factors, and is the most common pathological process in many diseases.

At present, many studies have confirmed that oxidative damage is the key to accelerating the development of chronic inflammation.

Sturgeon is an ancient and endangered commercial aquaculture species with high economic and ecological value.

Five kinds of proteases (Alcalase, papain, Neutrase, pepsin, trypsin) were used to hydrolyze sturgeon muscle protein.

A.

Figure 1 The effect of sturgeon muscle enzymatic hydrolysate on cell viability

Purification process of anti-inflammatory peptides

Next, the PeH was purified by ultrafiltration and Sephadex G-15 gel filtration.

A.

Figure 2 The effect of PeH ultrafiltration components on the anti-inflammatory activity of RAW264.

A.
Chromatogram of <3 kDa peptide separated by Sephadex G-15; B.
NO production; C.
IL-1β level.

Figure 3 Purification of anti-inflammatory peptides from PeH

A.
NO production; B.
IL-1β level; C.
IL-6 level; D.
TNF-α level.

Figure 4 The effect of F2 component on the production of pro-inflammatory cytokines in RAW264.
7 cells stimulated by LPS

F2's anti-inflammatory mechanism

As we all know, JAK-STAT, MAPK and NF-κB pathways play a key role in the inflammatory response.
LPS alone (2 μg/mL) can significantly increase the phosphorylation of ERK1/2, JNK, and p38 (4.
66, 5.
9, and 8.
0 times, respectively).
After pretreatment with F2, the phosphorylation levels of ERK1/2, JNK and p38 were significantly reduced by 25.
5%, 29.
7% and 46.
29% (P<0.
01) (Figure 5A).
In addition, compared with LPS alone, pretreatment with 0.
5 mg/mL F2 can significantly inhibit the phosphorylation and degradation of IκBα in the cytoplasm (inhibition rate is 37.
11%) (P<0.
01).
At the same time, LPS increased the expression of p65 in the nucleus by 2.
29 times, which was significantly reduced after F2 pretreatment (Figure 5B).
The results show that the anti-inflammatory effect of F2 is at least partially attributable to the inhibition and inhibition of IκBα phosphorylation and p65 nuclear translocation by the MAPK signaling pathway.

A.
Western blot analysis of p-ERK, ERK, p-JNK, JNK, p-p38 and p38 levels; B.
Western blot analysis of NF-κB p65, IκBα in the cytoplasm and nuclear extracts of RAW264.
7 cells Content.

Figure 5 The effect of F2 component on MAPKs and NF-κB protein stimulated by LPS in RAW264.
7 cells

Conclusion

The results of this study indicate that sturgeon peptides exert anti-inflammatory effects by down-regulating the MAPK and NF-κB pathways of RAW264.
7 macrophages stimulated by LPS.
As a rich anti-inflammatory food ingredient, sturgeon muscle hydrolysate has broad application prospects.
However, the specific active peptide sequence in the hydrolysate still needs further exploration.
At the same time, it is necessary to further study and evaluate whether this inhibitory property will also affect the secretion of other inflammatory factors and chemokines, and further in vivo studies on sturgeon peptides are needed to clarify the significance of the current research.

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