Research Article Protease Hydrolysates Ameliorates Inflammation and Intestinal Flora Imbalance in DSS-Induced Colitis Mice
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Hindawi Journal of Food Quality Volume 2021, Article ID 5536148, 11 pages https://doi.org/10.1155/2021/5536148 Research Article Protease Hydrolysates Ameliorates Inflammation and Intestinal Flora Imbalance in DSS-Induced Colitis Mice Lixia Zhao ,1 Weijie Tian,1 Wenjuan Qu,2 Ting Li,1 Junsong Zhu,1 and Haile Ma 2 1School of Food and Biological Engineering, Jiangsu University, Zhenjiang 212013, China 2Institute of Food Physical Processing, Jiangsu University, Zhenjiang 212013, China Correspondence should be addressed to Haile Ma; [email protected] Received 27 February 2021; Revised 26 April 2021; Accepted 29 May 2021; Published 7 June 2021 Academic Editor: Shengbao Cai Copyright © 2021 Lixia Zhao et al. +is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Wheat germ and fish skin usually has not been completely utilized and sometimes may be discarded, thus causing a lot of waste. Here, we aim at exploring the therapeutic anti-inflammatory effects of protease hydrolysates of wheat germ and fish skin on the ulcerative colitis (UC) mice. In the current study, wheat germ protein hydrolysates (WGPH) and fish skin gelatin hydrolysates (FSGH) treated mice had a longer colon than the DSS-induced mice. Moreover, protease hydrolysates reversed DSS-induced gut dysbiosis. Protease hydrolysates were likely to shift the balance of the intestinal flora on inflammation. In summary, these findings suggested that protease hydrolysates might serve as a latent therapy for UC treatment. 1. Introduction composes the intestinal mucosal barrier of the human body and shields the intestines [13]. Once the intestinal flora is Ulcerative colitis (UC) is recognized as a type of chronic and disturbed, it may cause inflammation of the colon [14]. recurrent inflammatory bowel disease (IBD) [1]. Unfortu- +e incidence of inflammation is increasing year by year, nately, UC patients often experienced abdominal pain and and the drugs for treating inflammation are always ex- bloody diarrhea, and the course of the disease is quite long [2]. pensive along with side effects [15, 16]. Given the concerns Additionally, the recurrence of UC is always along with grave about the side effects of long-term use of these drugs, food- complications and disappointing prognosis [3, 4]. Moreover, based functional food as alternatives to anti-inflammatory the incidence of UC has been rising year by year, particularly in drugs has become a research hotspot [17]. A number of the developed countries [5]. However, the etiology of UC is still studies have reported that some nutrients or food compo- unclear. +us, it is emerging to find effective therapy for UC. nents exert beneficial effects on UC well beyond their +e continuous improvement in the understanding of conventional nutritional value [18]. Previous studies have UC has evidenced that heredity, environmental factors, shown that the wheat oligopeptides have a protective effect dietary habits, immune factors, and gut microbiota are on DSS-induced colitis in mice [19, 20]. Vitexin and ana- intently related to the occurrence of UC [6, 7]. Among them, pigenin-8-C-glucoside have been shown that they could the interaction between gut microbiota and the host immune possess the positive effects on human health as well as gut system is considered as the major origin of colonic in- microbiota [21]. Corn proteolysis product has been proved flammation [8, 9]. Moreover, the amount of some ill bacteria to exert a good anti-inflammatory effect on trinitrobenzene (including Bacteroides fragilis, Escherichia coli, and Heli- sulfonic acid-induced colitis in rats [22]. Fish protein hy- cobacter) is flourishing in the UC patients’ intestinal tract, drolysates have multiple biological activities, containing which is markedly higher than that in healthy ones and is antioxidative, anticancer, lipid homeostasis modulation, probably connected to the incidence of UC [10, 11]. On this antihypertensive activities, anti-inflammatory, and neuro- basis, researchers have observed the correlation between UC protective effects which make them promising nutraceutical and gut microbiota [12]. +e ordinary gut microbiota ingredients for application in food [23]. Moreover, oral 2 Journal of Food Quality Control Model WGPH(L) WGPH(H) FSGH(L) FSGH(H) (a) 10 9 8 7 ∗∗∗ 6 ∗∗ ∗ 5 ### 4 Colon length (cm) 3 2 1 0 Model Control FSGH(L) FSGH(H) WGPH(L) WGPH(H) (b) Figure 1: Protease hydrolysates could alleviate the symptoms of DSS-induced UC mice. (a) Protease hydrolysates validly improved the shorten of the colon tissue. (b) Quantitative results of colon length in different groups. ###P < 0:001 vs. control; ∗P < 0:05, ∗∗P < 0:01, and ∗∗∗P < 0:001 vs. model. bovine collagen peptides have also been proved to obsess the sulfate sodium salt (DSS) salt buy in MP Biomedicals analgesic and anti-inflammatory effects on patients with Company. arthritis [24]. +e raw wheat germ was obtained and cleaned to remove However, the specific mechanism of the effects of contaminants. N-hexane (1 : 8, w/w) was used to defat the WGPH and FSGH on colitis is still unclear. +erefore, the wheat germ flour [25, 26], and the defatted wheat germ flour DSS-induced UC mouse model was utilized to detect the (DWGF) was scattered into the 1M NaCl solution (1 : 8, w/v) intervention of foodborne peptides on UC and observe the and stirred for 30 min at ambient temperature [27, 28]. effects of peptides on the intestinal tract, which would +en, the pH was adjusted to 9.5. After stirring for 30 min, provide new idea for the treatment of UC. the suspension was centrifuged at 8,000 g for 20 min at 4°C. +en, the pH of the supernatant was adjusted to 4.0 and 2. Materials and Methods centrifuged again at 8,000 g for 20 min at 4–8°C. Finally, the precipitate was obtained [29, 30]. 2.1. Protein Isolation Preparation. Wheat gluten protein +e skin of tilapia was homogenized with distilled peptide was extracted from our laboratory (Jiangsu water (1 : 20, w/v). Alcalase (7674 U/g protein) was added Jiangda Wukesong Biotechnology Co., Ltd). Male c57 to the suspension, and the pH was adjusted to 9.0. +en, it bl/6 mice, 6–8-week-old, weight 18∼24 g, were selected was hydrolyzed for 4.92 h at 55°C. Afterwards, the re- and purchased from Jiangsu University Experimental action was terminated by heating the solution in a Animal Center, license SCXK (Su) 2018–0012. Dextran waterbath at 95°C for 20 min. +e supernatant was Journal of Food Quality 3 100x Control Model WGPH(L) 100x WGPH(H) FSGH(L) FSGH(H) (a) 200x Control Model WGPH(L) 200x WGPH(H) FSGH(L) FSGH(H) (b) Figure 2: Histological observation and evaluation. (a) Representative photos of colonic segments (×100). (b) Representative photos of colonic segments (x200). collected by centrifugation at 10,000 rpm for 15 min and were the control group, DSS group, DSS + WGPH (300 mg/ then freeze-dried [31]. kg·d), DSS + WGPH (600 mg/kg·d), DSS + FSGH (300 mg/ kg·d), and DSS + FSGH (600 mg/kg·d). We treated the mice in the control group with pure water, and 3% DSS solution was 2.2. )e Design of Experiments. Sixty male C57BL/6 mice supplied to the other five groups to induce UC disease. +e (6–8-week-old) were obtained from Jiangsu University and WGPH and FSGH were orally administrated once a day at the housed at a constant temperature and humidity room. All mice same time according to the abovementioned volume for 7 days. were divided into 6 groups randomly with 10 mice each; they +e control and DSS groups were given normal saline, and all 4 Journal of Food Quality Observed_species 1500 600 1000 400 OTUs detected 500 Observed_species 200 0 0 5 10 15 20 0 5000 10000 15000 Number of samples sequenced Sequences_per_sample SampleID s491 s474 s444 s492 s101 s446 s496 s477 s448 s499 s480 s440 s452 s481 s463 s453 s487 s470 s460 s488 s466 s472 s489 s468 (a) (b) Figure 3: Rarefaction curves species (a) and accumulation curves (b) OTUs clustered at 97% sequence identity of all the samples. the mice were endowed for cervical dislocation on the 15th day +e conditions of PCR assay were listed as follows: primary for anatomical sampling. All experiments were approved by the denaturation at 94°C for 5 min; 32 cycles of 94°C for 30 sec, Institution Animal Care and Use Committee of Jiangsu Uni- annealing at 55°C for 30 sec, and extension 72°C for 60 sec; versity (UJS-IACUC-201920960) and met the guidelines of the and the final step was an extension at 72°C for 7 min. PCR National Institutes of Health Guide for the Care and Use of amplification was carried out in a total volume of 25 µL Laboratory Animals. reaction mixture including 3 µL BSA, 12.5 µL 2xTaq Plus Master Mix, 30 ng of template DNA and ddH2O, and 1 µL of each primer. We collected and purified the PCR products via 2.3. HE Assay. +e mice’ colon tissues were cut into 5 μm QIAquick Gel Extraction Kit (QIAGEN, Germany). After thick slices. After dewaxing and hydration, the slices were that, we prepared the amplicon pools for sequencing, size, incubated in hematoxylin solution for about 3–5 min and and quantity of the amplicon library through the Agilent later hatched in ethanol hydrochloric acid for 15 sec [32]. 2100 Bioanalyzer (Agilent, USA) as well as the Library Subsequently, the sections were washed by PBS solution, and Quantification Kit for Illumina, separately [12]. then, the sections were placed in eosin solution and incu- Next, the samples were then sequenced through the bated for about 2-3 min.