Table 1 Experimental design.
ARRIVE essential 10 | ||
|---|---|---|
Study design | 1 | All mice were male and stratified into three groups housed in different cages: those on a standard chow diet (CD) for 40Â weeks served as a control, while the rest of the mice were fed a high-fat diet in a longitudinal fashion. Mice in the Pre-T group were on this diet for 40Â weeks, while those in the Post-T group were on the diet for 48Â weeks and developed HCC, as shown in the development of this DIAMOND mice model26,27 |
Sample size | 2 | The total number of DIAMOND mice used was six, with two mice stratified in each group (Ctrl n = 2, Pre-T n = 2, and Post-T n = 2) in order to capture cellular changes in the liver during HCC progression. Sample size was decided based on cost of sequencing services and our ability to perform a base level of statistical analyses |
Inclusion and exclusion criteria | 3 | No animals were excluded from this study, nor was an exclusion criterion established |
Randomization | 4 | No randomization occurred, as all mice were housed in cages on the same rack, and their cage location remained the same throughout the study |
Blinding | 5 | No blinding was performed, as knowledge of which cage the mice were in was required to provide them with the correct experimental diet and water (CD and regular water compared to high-fat diet and sugar water) |
Outcome measures | 6 | Any behavioral changes in mice were monitored throughout the study. The outcome measure assessed was examination of the livers from our experimental groups of mice via snRNA-seq |
Statistical methods | 7 | DESeq2 processed data was filtered on a p-value < 0.01, followed by subsequent filtering in IPA of only molecules with a z-score > 2. Quantification of specific molecules of interest was performed in each replicate (n = 2 per group) in order to generate an average and standard error mean (SEM) for plotting. CellChat analyses only presented predicted signaling pathways deemed significant by the program (p-value < 0.05) |
Experimental animals | 8 | DIAMOND mice (129A1/SvIm and C57BL/6J cross mice) were all males in this study and started on their respective diets at two months of age for 40–48 weeks |
Experimental procedures | 9 | No experimental procedures were conducted during the time on high-fat diet. At the end of the time course mice were humanely euthanized to snap-freeze resected livers and subjected to snRNA-seq |
Results | 10 | Analyses of snRNA-seq data assessed both replicates (n = 2 per group) pooled together for visualization of data in Seurat and cell type annotation. Replicates were separated in R for statistical measures during analysis with DESeq2 and quantification of cells expressing specific molecules of interest |
