KjG The Korean Journal of Gastroenterology Korean J Gastroenterol 1598-9992 2233-6869 Korean Society of Gastroenterology 10.4166/kjg.2018.71.3.132 kjg-71-132 Original Article Combined Extracts of Artemisia and Green Tea, Mitigated Alcoholic Gastritis Via Enhanced Heat-shock Protein 27 KimYong Seok 1 JeongMigyeong 2 HanYoung Min 2 ParkJong-Min 2 KwonSang Oh 3 HongSeong Pyo 4 HahmKi Baik 2 4 Department of Biochemistry and Molecular Biology, Hanyang University College of Medicine, Seoul, Korea CHA Cancer Preventive Research Center, CHA Bio Complex, College of Medicine, CHA University, Pangyo, Korea S&D Research Center, S&D Foods, Cheongwon, Korea Digestive Disease Center, CHA University Bundang Medical Center, College of Medicine, CHA University, Seongnam, Korea Correspondence to: Ki Baik Hahm, Digestive Disease Center, Cha University Bundang Medical Center, 59 Yatap-ro, Bundang-gu, Seongnam 13496, Korea. Tel: +82-31-881-7250, Fax: +82-31-881-7185, E-mail: hahmkb@cha.ac.kr 10 2018 04 10 2018 713 132 142 31102017 13122017 Copyright © 2018 Korean Ophthalmological Society 2018 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background/Aims

Several lines of evidence from epidemiologic and laboratory studies have shown that the consumption of Artemisia or green tea extracts (MPGT) is inversely associated with the risk of alcohol-induced damage and other chronic diseases. Supported by previous studies showing that the combined extract of Artemisia and green tea, MPGT, exerted significantly either anti-oxidative or anti-inflammatory actions against Helicobacter pyloriassociated gastric diseases, it was hypothesized that MPGT can offer protection against alcoholic gastritis.

Methods

Ethanol was administered to induce gastric damage in Wistar rats, which had been pretreated with various doses of MPGT, to measure the rescuing action of a MPGT pretreatment against ethanol-induced gastric damage. In addition, the molecular mechanisms for the preventive effects were examined.

Results

The MPGT pretreatment (100, 300, and 500 mg/kg) alleviated the ethanol-induced gastric damage, which was evidenced by the significant decrease in calcium-dependent phospholipase A2, MAPKs, and NF- B levels compared to ethanol alone. Furthermore, the MPGT pretreatment preserved 15-prostaglandin dehydrogenase, whereas cyclooxygenase-2 was decreased significantly. All of these biochemical changes led to the significant alleviation of alcoholassociated gastric mucosal damage. Ethanol significantly increased the TUNEL positivity in the stomach, but MPGT decreased the apoptotic index significantly, which was associated with significantly lower pathological scores of ethanol-induced mucosal ulcerations. The significant protective changes observed alcoholic gastritis with MPGT were related to the increased expression of cytoprotective genes, such as heat-shock protein (HSP)27, HSP60, and PDGF.

Conclusions

The efficient anti-inflammatory, anti-apoptotic, and regenerative actions of MPGT make it a potential nutrient phytoceut-ical to rescue the stomach from alcoholic gastritis.

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MPGT pretreatment alleviated ethanol-induced gastric damage. (A) Experimental scheme. Five animal groups were set: Group 2 as the control and Groups 3, 4, and 5 pretreated with 100 mg/kg, 300 mg/kg, and 500 mg/kg MPGT. The rats were sacrificed after 1 hour of 100% ethanol. Representative pictures of the gross appearance of the rat stomach after alcohol ingestion in the absence or presence of 100, 300, and 500 mg/kg MPGT pretreatment. (B) Gross and microscopic photograph of ethanol-induced gastric mucosal lesions according to the group, ×40 magnification. (C, D) The scoring criteria are written in Methods. The mean score for the gross and pathologic lesion scores. The mean levels of the gross lesion index and mean scores of pathological evaluation are presented as the mean±SD from 8 rats/group. MPGT, extracts of artemisia and green tea; SD, standard deviation.

Status of ethanol-induced gastric inflammation. (A) Mean score for inflammation according to group. (B) Changes in IL-8, phosphate ERK, phosphate JNK, and phosphate p38 mRNA and their protein expression. (C) Immunohistochemistry staining with the NF-κ B p65 anti-body, ×100 magnification, and the mean positivity according to the group. ERK, extracellular signal-regulated kinases; JNK, c-Jun N-terminal kinases; mRNA, messenger RNA; SD, standard deviation.

Status of ethanol-induced mucosal and submucosal damage. (A) Score of mucosal and submucosal damage. Pathology image representing the mucosal and submucosal condition from Groups 2 and 5 ×100 magnification. (B) Western blot for cPLA2 and 15-PGDH (C) RT-PCR and Western blot for COX-2 according group (left) (D) Immunohistochemical staining with COX-2 antibody (middle), ×100 magnification, and mean COX-2 positivity. cPLA2, calcium-dependent phospholipase A2; COX-2, cyclooxygenase-2; ERK, extracellular signal-regulated kinases; JNK, c-Jun N-terminal kinases, SD, standard deviation.

Status of an ethanol-induced gastric ulcer. (A) Mean score for ethanol-induced gastric ulcers. The arrow shows the significant denudation and exfoliation of gastric mucosa. (B) TUNEL staining for apoptotic cells. All images were obtained from the non-ulcerated mucosa of each group and significantly increased TUNEL positive cells were noted in Group 2 even in the non-ulcerated gastric mucosa. (C) Mean TUNEL positive (apoptotic index) according to the group. (D) Western blot for BAX expression. TUNEL, terminal deoxynucleotidyl transferase mediated dUTP nick-end labeling; SD, standard deviation.

Western blot for HSP27 and HSP. (A) Changes in HSP27 and HSP70 according to group. (B) Changes in Hsp27 and Hsp70 mRNA by RT-PCT. (C) Changes in Pdgf mRNA by RT-PCR. HSP, heat-shock protein; mRNA, messenger RNA; SD, standard deviation.

Determination of the Score in H. pylori– Induced Inflammation and Ulcers

  Scores
Inflammation  
   1/3 portion of mucosa 1
   2/3 portion of mucosa 2
   3/3 portion of mucosa 3
Grade  
   0 μ m 0
   0–1,000 μ m 1
   1,001–2,000 μ m 2
   2,001–3,000 μ m 3
   3,001 μ m 4
Ulcers, erosion length×depth (in mucosa)  
   Gland 0.5
   1/3 portion of mucosa 1
   2/3 portion of mucosa 2
   3/3 portion of mucosa 3
Ulcers size  
   Ulcer or erosion up to 1 mm n×2
   Ulcer or erosion larger than 1 mm n×3

H. pylori, Helicobacter pylori.