Different Detection Strategies of Pediocin-Like Produced by Pediococcus pentosaceus.

In this study, Pediococcus pentosaceus C-2-1 and C23221 contained genes encoding penocin and pediocin PA-1, mined by antiSMASH. The penocin structural gene pedA from Pediococcus pentosaceus C-2-1 was successfully expressed in Escherichia coli BL21. The presence of a 6.5 kDa recombinant penocin was confirmed by Tricine-SDS-PAGE, and the specific activity increased by 1.54-fold. The bacteriocins produced by Pediococcus pentosaceus C23221 were purified using acetic ether extraction, Sepharose Fast Flow, Sephadex G-25 gel chromatography, and reversed-phase high-performance liquid chromatography (RP-HPLC); the amino acid sequence of this bacteriocin was identical to pediocin PA-1 by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), which confirmed the expression of pediocin PA-1 gene; and the specific activity increased by 24.39-fold. The heterologous expression and purification of bacteriocins have proved the expression of pediocin-like produced by Pediococcus pentosaceus. This provides a theoretical basis for the subsequent development and application of pediocin-like.

Intervention mechanism of marine-based chito-oligosaccharide on acute liver injury induced by AFB1 in rats.

Aflatoxin B1 (AFB1) is extremely hepatotoxic, a causative agent of liver cancer, and can cause symptoms of acute or chronic liver damage. Chito-oligosaccharides (COS), obtained from the degradation of chitosan derived from shrimp and crab shells, is a natural antioxidant substance and its antitumor properties have been widely studied, but less research has been done on the prevention of AFB1-induced acute liver injury. In this study, rats were acutely exposed to 1 mg/kg BW AFB1 and simultaneously gavaged with different doses of COS for 8 days. The results showed that COS attenuated the hepatic histopathological changes and reduced serum biochemical indices (ALT, AST, ALP, and TBIL) in rats. It significantly inhibited MDA content and promoted SOD and GSH-Px activity production. Moreover, it also improved hepatocyte apoptosis. Furthermore, AFB1-vs-HCOS differential genes were enriched with 622 GO entries, and 380 were Biological Processes, 170 were Molecular Functions, 72 were Cellular Components. Differentially expressed genes (DEGs) analyzed by KEGG enrichment were more enriched in pathways, such as metabolism, PPAR signaling pathway, and peroxisome. Q-PCR technique verified that Lama5, Egr1, Cyp2b1, and Gadd45g in DEGs were associated with oxidative stress damage and apoptosis. In conclusion, COS intervention reduces the effect of AFB1 on hepatic genes and thus reduces the changes in hepatic gene function.

Antibacterial Ingredients and Modes of the Methanol-Phase Extract from the Fruit of Amomum villosum Lour.

Epidemics of infectious diseases threaten human health and society stability. Pharmacophagous plants are rich in bioactive compounds that constitute a safe drug library for antimicrobial agents. In this study, we have deciphered for the first time antibacterial ingredients and modes of the methanol-phase extract (MPE) from the fruit of Amomum villosum Lour. The results have revealed that the antibacterial rate of the MPE was 63.64%, targeting 22 species of common pathogenic bacteria. The MPE was further purified by high performance liquid chromatography (Prep-HPLC), and three different constituents (Fractions 1-3) were obtained. Of these, the Fraction 2 treatment significantly increased the cell membrane fluidity and permeability, reduced the cell surface hydrophobicity, and damaged the integrity of the cell structure, leading to the leakage of cellular macromolecules of Gram-positive and Gram-negative pathogens (p < 0.05). Eighty-nine compounds in Fraction 2 were identified by ultra HPLC-mass spectrometry (UHPLC-MS) analysis, among which 4-hydroxyphenylacetylglutamic acid accounted for the highest 30.89%, followed by lubiprostone (11.86%), miltirone (10.68%), and oleic acid (10.58%). Comparative transcriptomics analysis revealed significantly altered metabolic pathways in the representative pathogens treated by Fraction 2 (p < 0.05), indicating multiple antibacterial modes. Overall, this study first demonstrates the antibacterial activity of the MPE from the fruit of A. villosum Lour., and should be useful for its application in the medicinal and food preservative industries against common pathogens.

Development of a UPLC-MS/MS-based method for simultaneous determination of advanced glycation end products and heterocyclic amines in stewed meat products.

A novel analytical strategy was proposed to simultaneously quantify two advanced glycation end products (AGEs) including Nε-(Carboxymethyl)lysine (CML), Nε-(Carboxyethyl)lysine (CEL) and eight heterocyclic amines (HAs) including IQ, MeIQ, MeIQx, 4,8-DiMeIQx, 7,8-DiMeIQx, PhIP, Harman, and Norharman. The procedure was based on a two-step extraction, solid phase extraction (SPE) purification followed by ultra performance liquid chromatography tandem mass spectrometry. The established method showed a good linearity (R2 ≥ 0.9950), rapid processing time (8 min per sample), satisfactory recoveries (matrix spiked recoveries range from 72.2% to 119.6%) and precision (intra-day and inter-day RSDs were <19.3%). The limit of quantification (LOQ) and limit of detection (LOD) resulted to be between 0.05-15 ng/g and 0.2-50 ng/g, respectively. The validated technique was further applied to determine HAs and AGEs in eight stewed meat product samples consumed in Shanghai, with the amount of HAs and AGEs ranging from 2.851 to 18.289 ng/g and 118.158-543.493 ng/g, respectively.

Resistant starch intake facilitates weight loss in humans by reshaping the gut microbiota.

Emerging evidence suggests that modulation of gut microbiota by dietary fibre may offer solutions for metabolic disorders. In a randomized placebo-controlled crossover design trial (ChiCTR-TTRCC-13003333) in 37 participants with overweight or obesity, we test whether resistant starch (RS) as a dietary supplement influences obesity-related outcomes. Here, we show that RS supplementation for 8 weeks can help to achieve weight loss (mean -2.8 kg) and improve insulin resistance in individuals with excess body weight. The benefits of RS are associated with changes in gut microbiota composition. Supplementation with Bifidobacterium adolescentis, a species that is markedly associated with the alleviation of obesity in the study participants, protects male mice from diet-induced obesity. Mechanistically, the RS-induced changes in the gut microbiota alter the bile acid profile, reduce inflammation by restoring the intestinal barrier and inhibit lipid absorption. We demonstrate that RS can facilitate weight loss at least partially through B. adolescentis and that the gut microbiota is essential for the action of RS.

Effect of chemical modification of κ-carrageenan on its inhibitory effect against heterocyclic amine (HAs) formation in roasted tilapia fish patties.

Sulfated κ-carrageenan (S-KC), carboxymethylated κ-carrageenan (C-KC), acetylated κ-carrageenan (A-KC) and phosphorylated κ-carrageenan (P-KC) were synthesized and tested for their inhibitory effect on heterocyclic amine (HAs) formation in roasted tilapia fish patties. Fish patties with 1 % of each hydrocolloid prepared by 90 % of fish and 10 % of an aqueous hydrocolloid dispersion were determined for HAs-levels after roasting. P-KC showed the strongest inhibitory effect against total HAs formation (20.95 %). Moreover, P-KC increased the content of creatinine and glucose but decreased the content of free amino acids in fish patties, indicating that P-KC may compete with creatinine and glucose to react with amino acids to suppress HAs generation. In addition, P-KC plus naringenin had a stronger inhibitory effect against HAs formation than P-KC or naringenin alone. P-KC at 1 % (w/w) and P-KC (0.5 %, w/w) plus naringenin (0.5 %, w/w) showed no significant effects on the color and textural properties compared to the control group (100 % fish), and had less impact on food quality than 1 % (w/w) KC. Therefore, our results suggest that chemical modification could enhance the inhibitory effect of some hydrocolloids on HAs formation, and an appropriate combination of hydrocolloids and flavonoids contributes to the attenuation of dietary exposure to genotoxic HAs.

Resistant starch decreases intrahepatic triglycerides in patients with NAFLD via gut microbiome alterations.

Non-alcoholic fatty liver disease (NAFLD) is a hepatic manifestation of metabolic dysfunction for which effective interventions are lacking. To investigate the effects of resistant starch (RS) as a microbiota-directed dietary supplement for NAFLD treatment, we coupled a 4-month randomized placebo-controlled clinical trial in individuals with NAFLD (ChiCTR-IOR-15007519) with metagenomics and metabolomics analysis. Relative to the control (n = 97), the RS intervention (n = 99) resulted in a 9.08% absolute reduction of intrahepatic triglyceride content (IHTC), which was 5.89% after adjusting for weight loss. Serum branched-chain amino acids (BCAAs) and gut microbial species, in particular Bacteroides stercoris, significantly correlated with IHTC and liver enzymes and were reduced by RS. Multi-omics integrative analyses revealed the interplay among gut microbiota changes, BCAA availability, and hepatic steatosis, with causality supported by fecal microbiota transplantation and monocolonization in mice. Thus, RS dietary supplementation might be a strategy for managing NAFLD by altering gut microbiota composition and functionality.

Effects of Stigmasterol on 3-Chloropropane-1,2-diol Fatty Acid Esters and Aldehydes Formation in Heated Soybean Oil.

In this study, we investigated the inhibitory effects of three soybean isoflavones and two soybean phytosterols on the formation of 3-chloropropane-1,2-diol fatty acid esters (3-MCPDE) and aldehydes in heated soybean oil model. 0.4 mM of genistin, genistein, daidzein, stigmasterol, and ß-sitosterol significantly reduced 3-MCPDE formation by 25.7, 51.4, 21.4, 61.6, and 55.7%, and total aldehydes formation by 42.03, 43.94, 28.36, 54.74, and 39.23%, respectively. Further study showed that stigmasterol reduced the content of glycidyl esters (GEs) and glycidol, two key intermediates of 3-MCPDE, and prevented fatty acids degradation in the oils. Moreover, the effects of continuous frying time on the content of stigmasterol and the migration of stigmasterol were evaluated in the fried dough sticks model system. The content of stigmasterol in soybean oil was found to be significantly decreased with prolonged heating time. The concentrations of stigmasterol in fried dough sticks and the migration rates of stigmasterol from soybean oil to fried dough sticks decreased with repeated frying sessions. In addition, stigmasterol undergoes oxidative changes during heat treatment, and the oxidation products including 5,6α-epoxystigmasterol, 5,6ß-epoxystigmasterol, 7α-hydroxystigmasterol, 7ß-hydroxystigmasterol, stigmasterlol-3ß,5α,6ß-triol, and 7-ketostigmasterol were identified in the frying oils but not in the fried dough sticks. Overall, stigmasterol could be added to soybean oil to reduce 3-MCPDE and aldehydes formation, and reacting with GEs/glycidol and protection of lipid acids from oxidation may be the mechanism of action of stigmasterol.

Multi-omics reveal key enzymes involved in the formation of phenylpropanoid glucosides in Artemisia annua.

Although mainly known for producing artemisinin, Artemisia annua is enriched in phenylpropanoid glucosides (PGs) with significant bioactivities. However, the biosynthesis of A. annua PGs is insufficiently investigated. Different A. annua ecotypes from distinct growing environments accumulate varying amounts of metabolites, including artemisinin and PGs such as scopolin. UDP-glucose:phenylpropanoid glucosyltransferases (UGTs) transfers glucose from UDP-glucose in PG biosynthesis. Here, we found that the low-artemisinin ecotype GS produces a higher amount of scopolin, compared to the high-artemisinin ecotype HN. By combining transcriptome and proteome analyses, we selected 28 candidate AaUGTs from 177 annotated AaUGTs. Using AlphaFold structural prediction and molecular docking, we determined the binding affinities of 16 AaUGTs. Seven of the AaUGTs enzymatically glycosylated phenylpropanoids. AaUGT25 converted scopoletin to scopolin and esculetin to esculin. The lack of accumulation of esculin in the leaf and the high catalytic efficiency of AaUGT25 on esculetin suggest that esculetin is methylated to scopoletin, the precursor of scopolin. We also discovered that AaOMT1, a previously uncharacterized O-methyltransferase, converts esculetin to scopoletin, suggesting an alternative route for producing scopoletin, which contributes to the high-level accumulation of scopolin in A. annua leaves. AaUGT1 and AaUGT25 responded to induction of stress-related phytohormones, implying the involvement of PGs in stress responses.

Comparative genomics reveals key molecular targets for mutant Pediococcus pentosaceus C23221 producing pediocin.

Listeria monocytogenes is a common microorganism that causes food spoilage. Pediocins are some biologically active peptides or proteins encoded by ribosomes, which have a strong antimicrobial activity against L. monocytogenes. In this study, the antimicrobial activity of previously isolated P. pentosaceus C-2-1 was enhanced by ultraviolet (UV) mutagenesis. A positive mutant strain P. pentosaceus C23221 was obtained after 8 rounds of UV irradiation with increased antimicrobial activity of 1448 IU/mL, which was 8.47 folds higher than that of wild-type C-2-1. The genome of strain C23221 and wild-type C-2-1 was compared with identify the key genes for higher activity. The genome of the mutant strain C23221 consists of a chromosome of 1,742,268 bp, with 2052 protein-coding genes, 4 rRNA operons, and 47 tRNA genes, which is 79,769 bp less than the original strain. Compared with strain C-2-1, a total of 19 deduced proteins involved in 47 genes are unique to C23221 analyzed by GO database; the specific ped gene related to bacteriocin biosynthesis were detected using antiSMASH in mutant C23221, indicating mutant C23221 produced a new bacteriocin under mutagenesis conditions. This study provides genetic basis for further constituting a rational strategy to genetically engineer wild-type C-2-1 into an overproducer.

Assessment of lipid composition and eicosapentaenoic acid/docosahexaenoic acid bioavailability in fish oil obtained through different enrichment methods.

In this study, we analyzed the eicosapentaenoic acid/docosahexaenoic acid (EPA/DHA) lipid composition of fish oil obtained through enzymatic treatment, fractional distillation and silica gel column purification, and further assessed EPA/DHA bioavailability. Lipid subclass composition information was obtained through ultra-performance liquid chromatography-electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS), and bioavailability tests were performed using the Caco-2 cell monolayer model. Results showed that enzymatic treatment improved the incorporation of EPA/DHA as diacylglycerol (DG) while silica gel column chromatography enriched the content of EPA/DHA as phosphatidylglycerol (PG) (12.58%) and phosphatidylethanolamine (PE) (4.99%). Furthermore, increasing the purity of EPA/DHA could improve its bioavailability and after 24 incubation, binding forms of triglyceride (TG) was superior to ethyl ester (EE) (p < 0.05) at the same purity level. Those findings are helpful to provide research basis for exploring the bioactivity of fish oil.

Insight into mechanism of quality changes in tilapia fillets during salting from physicochemical and microstructural perspectives.

The effects and mechanisms of salting on quality properties of tilapia fillets were investigated in the present study. Salting under high NaCl concentrations (12 % and 15 %) resulted in low water content and decreased yields, due to the salting-out effects and low pH. Water in fillets increased in the later stage of salting in 3 % and 6 % NaCl solutions (p < 0.05). The released proteins accumulated with increasing time (p < 0.05). The TBARS value increased from 0.01 to 0.20 mg/kg after 10 h in 15 % NaCl solution (p < 0.05). The quality changes were mainly correlated to the shrinking or swelling of myofibers, extracellular spaces, and existential state of muscle proteins. In consideration of fish quality and increasing call for low sodium intake, it was recommended to prepare fillets below 9 % NaCl with short times. The finding provided instructions to obtain target quality properties from tilapia by controlling salting conditions.

Effect of acrolein on the formation of harman and norharman in chemical models and roast beef patties.

Harman and norharman were the most abundant ß-carboline-type heterocyclic amines (HCAs) detected in various foodstuffs. Unsaturated fatty acids in foods may undergo rapid oxidative deterioration during transportation, storage and heat treatment, forming reactive carbonyl species (RCS). This work studied the effects of acrolein, a highly reactive RCS, on the formation of harman and norharman in the tryptophan model system. Results showed that 0.005, 0.01, 0.015, 0.02, 0.05, 0.1 and 0.2 mmol of acrolein led to harman production increased by 528 %, 752 %, 981 %, 1172 %, 1375 %, 1288 % and 768 % respectively, and led to norharman formation increased by 116 %, 129 %, 152 %, 169 %, and 197 %, 185 % and 157 %, respectively. Furthermore, acrolein addition reduced the residue of tryptophan (up to 63.19 %), but increased the level of the intermediates including formaldehyde (up to 352 %), acetaldehyde (up to 491 %), (1S,3S)-1-Methyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid (MTCA, up to 1936 %), and 1,2,3,4-tetrahydro-ß-carboline-3-carboxylicacid (THCA, up to 2142 %) in the tryptophan model system. Acrolein might react with tryptophan, harman and norharman to eliminate them directly. These data suggested that acrolein may contribute to harman and norharman formation through participating in the above complex chemical reactions. In addition, the content of harman and norharman produced in roast beef patties made of minced beef oxidized for 2, 4, 6, 8, and 10 days increased by 118 %, 188 %, 267 %, 137 %, and 48 %, respectively, and led to norharman formation increased by 140 %, 132 %, 90 %, 86 %, and 74 %, respectively compared with those made of fresh minced beef, which further illustrated that lipid oxidation products potentially contributed to harman and norharman formation.

Apigenin and its octoic acid diester attenuated glycidol-induced autophagic-dependent apoptosis via inhibiting the ERK/JNK/p38 signaling pathways in human umbilical vein endothelial cells (HUVECs).

Glycidol is a well-known food contaminant mainly formed in refined edible oils and various thermally processed foods. Here, we studied the toxicity effects and related mechanism of glycidol on Human umbilical vein endothelial cells (HUVECs). Glycidol was found to induce Gap period 2 (G2)/Mitosis (M) phase cell cycle arrest, apoptosis, and autophagy in HUVECs. Inhibition of autophagy by 3-methyladenine (3-MA) attenuated glycidol-induced cell death, suggesting that glycidol-induced apoptosis was autophagy-dependent. Moreover, glycidol treatment induced phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal protein kinase (JNK), and p38. Inhibition of ERK, JNK, and p38 phosphorylation by the inhibitors U0126, SP600125, and SB203580 attenuated glycidol-induced autophagy and prevented glycidol-mediated reduction in cell viability, demonstrating that glycidol inhibited HUVECs growth by inducing autophagic-dependent apoptosis through activation of the ERK, JNK and p38 signaling pathways. In addition, apigenin (API) and its octoic acid diester apigenin-7 (API-C8), 4'-O-dioctanoate were found to significantly attenuate glycidol-induced cell growth inhibition by inhibiting the above signaling pathways. Collectively, glycidol induces autophagic-dependent apoptosis via activating the ERK/JNK/p38 signaling pathways in HUVECs and API-C8 could attenuate the toxicity effects.

Effects of quercetin on emissions of aldehydes from heated docosahexaenoic acid (DHA)-fortified soybean oil.

Home cooking has been considered as an indoor pollution problem since cooking oil fumes contain various toxic chemicals such as aldehydes. Fortifying edible oils with docosahexaenoic acid (DHA) has been applied to enhance the nutritional value of oils. This study designed a frying simulation system and examined the effect of oil type, DHA fortification, heating time, and addition of natural antioxidant on the emissions of aldehydes from heated oils. Results showed that linseed oil had the highest total aldehyde emissions, followed by soybean oil, peanut oil, and palm oil. Fortifying soybean oil with DHA increased the toxic aldehydes emitted. Quercetin, a flavonoid, significantly reduced aldehydes emitted from DHA-fortified soybean oil (by up to 39.80%) to levels similar to those of normal soybean oil. Further analysis showed that DHA-fortified soybean oil with quercetin had a significantly higher DHA and unsaturated fatty acids (UFAs) content than the control oil at each heating time point. The result indicated that quercetin inhibited emissions of aldehydes, at least in part, by protecting UFAs from oxidation. Collectively, quercetin could be used as a natural additive in DHA-fortified and normal cooking oils to reduce aldehyde emissions, indoor air pollution, and preserve functional DHA and other UFAs.

Quality improvement of tilapia fillets by light salting during repeated freezing-thawing: Contribution of structural rearrangement and molecular interactions.

The present study evaluated the effects and underlying mechanisms of light salting on quality properties of tilapia fillets during repeated freezing-thawing. Light salting was found to improve water-holding capacity and decelerated texture softening in tilapia fillets during repeated freezing-thawing. Instead of tissue distortion and heterogeneous aggregates in control groups, light salting promoted myofibril disassembly and formation of an ordered protein network with the solubilized myofibrillar proteins. The myofibrils presented an overall amorphous appearance with the loss of M-lines, removing the restraints to myofibril swelling and solubilization from A-binds in salted groups during repeated freezing-thawing. The structural rearrangement caused by light salting facilitated the enlargement of water-holding space, transformation of tissue water, and tissue recoverability, improving water-holding capacity and texture properties of tilapia fillets during freezing-thawing. The finding provided novel insight into the improvement of quality properties of tilapia fillets by light salting when subjected to drastic temperature fluctuations.

Apigenin and apigenin-7, 4'-O-dioctanoate protect against acrolein-aggravated inflammation via inhibiting the activation of NLRP3 inflammasome and HMGB1/MYD88/NF-κB signaling pathway in Human umbilical vein endothelial cells (HUVEC).

Exposure to acrolein, one environmental and dietary pollutant, has been shown to cause inflammation. Here, we reported for the first time that acrolein aggravated lipopolysaccharide (LPS)-induced inflammation in Human umbilical vein endothelial cells (HUVEC) as evidenced by the further increased mRNA expression of three pro-inflammatory cytokines, including interleukin 1ß (IL-1ß), interleukin 6 (IL-6), and tumor necrosis factor-alpha (TNF-α). Acrolein also further increased the generation of reactive oxygen species (ROS) and decreased the activity of glutathione peroxidase (GSH-Px) in LPS-pretreated HUVEC. Moreover, acrolein treatment further increased the nucleotide oligomerization domain-like receptor protein 3 (NLRP3) and apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) expression, caspase-1 cleavage, and downstream matures interleukin 18 (IL-18) and IL-1ß level in LPS-pretreated HUVEC. Acrolein treatment also further increased the expressions of high-mobility group box 1 (HMGB1), toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), and phospho-NF-κB P65 (P-P65) in the LPS pre-treated HUVEC. Thus, acrolein aggravated LPS-induced HUVEC inflammation through induction of oxidative stress, and activation of NLRP3 inflammasome and HMGB1/MYD88/NF-κB signaling pathway. In addition, apigenin and apigenin-7, 4'-O-dioctanoate attenuated acrolein-aggravated inflammation by targeting the above signaling pathways. Our findings could help to develop potential therapeutic strategies against acrolein-enhanced inflammation.

Effect of acrolein, a lipid oxidation product, on the formation of the heterocyclic aromatic amine 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in model systems and roasted tilapia fish patties.

The effect of acrolein on the formation of the 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) was investigated in a chemical model. Acrolein was found to increase PhIP formation at each tested addition level. 0-0.2 mmol of acrolein increased PhIP formation dose-dependently, while high levels of acrolein (>0.2 mmol) did not further increase PhIP formation. Mechanistic study showed that acrolein addition decreased the residue of phenylalanine and creatinine, but increased the content of some key intermediates. Further analysis indicated that acrolein can react with phenylalanine, creatinine, and PhIP to form adducts. These results suggested that acrolein was able to contribute to PhIP formation as a consequence of its comprehensive ability to facilitate Strecker degradation of phenylalanine and react with phenylalanine, creatinine, and PhIP. In addition, oxidation of the tilapia fish increased the PhIP formation in the roasted fish patties, further supporting the potential contribution role of lipid oxidation products to the formation of PhIP.

FIP-fve Stimulates Cell Proliferation and Enhances IL-2 Release by Activating MAP2K3/p38α (MAPK14) Signaling Pathway in Jurkat E6-1 Cells.

FIP-fve, a fungal fruiting body protein from Flammulina velutipes, has potential immunomodulatory properties. Here, we investigated the immunomodulation mechanism of FIP-fve in Jurkat E6-1 cells by conducting a cell viability assay and IL-2 release assay. Kinase inhibitors experiment and proteomics analysis were also involved in the mechanism study. It was found that FIP-fve stimulated cell proliferation and enhanced IL-2 secretion in a dose-dependent manner in Jurkat E6-1 cells. Unbiased high-throughput proteomics analysis showed that 4 T cell immune activation markers, including ZAP-70, CD69, CD82, and KIF23, were upregulated in response to FIP-fve treatment. Further pathway analysis indicated that MAP2K3/p38 pathway-related proteins, including MAP2K, p38, ELK, AATF, FOS, and JUN-B, were unregulated. In addition, losmapimod (p38 inhibitor) and gossypetin (MAP2K3 inhibitor) inhibited FIP-fve enhanced cell proliferation and IL-2 release in Jurkat E6-1 cells. Our results demonstrate that FIP-fve stimulates cell proliferation and enhances IL-2 secretion through MAP2K3/p38α activation.