Data of multiple different high-fat diets

Background and Objectives: The spectrum of non-alcoholic fatty liver disease (NAFLD), known as non-alcoholic steatohepatitis (NASH), can lead to advanced liver disease. It is known that a variety of diets play a significant role in the development of NAFLD/NASH. The goal of this study is to determine the most appropriate composition of diet to induce NASH in an animal model. Methods: This research used Rattus norvegicus strain Wistar (n=27), which were divided into four groups and given each diet for 12 weeks: normal diet (ND, n=7), high-fat diet (HFD, n=6), Western diet (WD, n=7), high-fat-high-fructose diet (HFHFD, n=7). Subjects were documented for body weight. Blood samples were taken for biochemical analysis: low-density lipoprotein (LDL), triglyceride, alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), hepatic lipase, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and lipopolysaccharide (LPS). Feces were taken for short-chain fatty acid (SCFA) analysis. Liver histology was analyzed using NAS (NAFLD activity score). The comparison test was carried out using the one-way ANOVA or Kruskal–Wallis test. Results: The highest mean of body weight was in the WD group (346.14 g). Liver enzymes, LDL, triglyceride, propionic acid, and acetic acid in each group were not significantly different. TNF-α, IL-6, and hepatic lipase were significant (p = 0.000; p = 0.000; p = 0.004) and the highest was in the HFD group. Butyrate level was significant (p = 0.021) and the least was in the HFHFD group (4.77 mMol/g). Only WD and HFHFD had an NAS ≥ 5 (14% and 14%). The highest percentage of borderline NAS was found in WD (57%). Conclusions: The HFD group induced significant liver inflammation but did not produce NASH histologically, whereas the WD and HFHFD groups had the potential to develop NASH because the diets affected metabolic and inflammatory parameters as well as liver histology.Software that is required to open the data files in format .sav is SPSS.The Ethical Committee of the Faculty of Medicine, Universitas Brawijaya reviewed and approved all procedures (No. 66/EC/KEPK/02/2021). Twenty seven male Wistar rats were obtained from the Universitas Gadjah Mada. Rat inclusion criteria included: male rat with shiny white fur, healthy, active, and have normal behavior; about 8–12 weeks old; the average body weight was 150–180 grams. Exclusion criteria included: the appearance of dull fur, loss and baldness; less or inactive activity; rats that during the study did not want to eat; weight loss >10% after adaptation period; Disabled, sick and/or dead rat. In minimizing pain and discomfort to experimental animals, this research uses the refinement principle for ensuring the welfare of experimental animals until the end of the study. We provided food and drink regularly every day with a certain type of diet according to the type of treatment. Cage maintenance, cage cleaning, and husk replacement were carried out every day with attention to light, temperature, and humidity. We monitored and evaluated rats condition every day and placed them individually in each cage. Before being treated, the rats were acclimatized for 2 weeks, given a standard diet, and placed inside cages at the Pharmacology Laboratory, Faculty of Medicine, University Brawijaya. The Wistar rats were randomly assigned using table of random numbers, then categorized into four groups: normal diet (ND) (67% carbohydrate, 21% protein, 7% fat, 5% fiber); HFD (67.1% carbohydrate, 16.5% fat, 16.4% protein), WD (52% carbohydrate, 16.1% protein, 31.7% fat), and HFHFD (41.5% carbohydrate, 10.3% fat, 10.2% protein, 38% fructose). All diets were given for 12 weeks. All four groups were euthanized with ketamine–xylazine intravenously to relieve pain on the same day before surgery was performed. The entire liver was taken out and weighed. For further analysis, the livers were either collected and stored at -20°C or fixed in 10% paraformaldehyde. Rat serum was used to analyze biochemical parameters in the Clinical Pathology Laboratory, Universitas Brawijaya, Indonesia. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) were analyzed chemically using colorimetric analysis (ADVIA 2400 Clinical Chemistry System (Siemens, Germany). Serum hepatic lipase, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and lipopolysaccharide (LPS) were analyzed with the sandwich enzyme-linked immunosorbent assay (ELISA) method. A total of 0.5 gram fecal samples from the colon were used, and a dilution ratio of 1:4 between samples and sterilized water was established. We subsequently vortexed the samples for 1 min and centrifuged the homogenate at 10000 x g for 10 min at 4°C. Filtration of SCFA-containing supernatant was completed by using cellulose acetate membrane (GyroDisc, Orange Scientific, CA, US) and it was stored at -80°C until further analysis was performed. At the time of analysis, 20 mg of the faecal sample supernatant poured into a 1.5 ml microtube and then added 200uL of distilled water homogenized using an iron spatula. 200 µL of sample, 200 µL of a mixture of N-butanol, tetrahydrofuran and acetonitrile at a ratio of 50:30:20 40 µl 0.1 M HCl, 20 mg of citric acid and 40 mg of sodium chloride were added to 1.5 mL microtubules. The microtubules were then shaken using a vortex for 1 minute. Sample centrifuged at 14870 x g at room temperature for 10 minutes. Supernatant transferred using a micropipette to the chromatography vial for later Gas Chromatography analysis was carried out. The results of the analysis are entered into the software EZchrome and GC solution. SCFA analysis was performed at Food Technology and Agricultural Products Laboratory, Universitas Gadjah Mada, Indonesia. The liver was sliced, fixed with 10% buffered formalin, embedded in paraffin, and stained with hematoxylin–eosin (HE) stain at a thickness of 5 μm (11). The sample preparation was performed at the Anatomical Pathology Laboratory of Universitas Brawijaya, Indonesia. Liver histopathology was used to find the NAFLD Activity Score (NAS). Three parameters (steatosis score 0–3; lobules inflammation score 0–3; ballooning score 0–2) were used to know NAFLD staging. Scores of 0–2 are defined non NASH, scores of 3–4 are defined as borderline, while scores ≥ 5 are considered diagnostic of NASH. Data were presented as the mean ± standard deviation and were analyzed with SPSS 25.0 (RRID:SCR_002865) for Windows. A one way ANOVA was carried out when the data were normally distributed and then continued with the Tukey Honest Significant Difference (HSD) post hoc test if the data were significant. The Kruskal–Wallis test was used when the data distribution was not normal. If the results were significant then the Mann Whitney test was performed. When p