Lastly, the collected μ Opioid Receptor/MOR Formulation intestinal SIRT2 Storage & Stability tissue samples had been used for transcription analysis. 2.4. Morphological Section Analysis of Rabbit Intestine We chosen seven rabbits with diarrhea and 3 healthy rabbits for intestinal morphological section evaluation. The intestinal tissue specimens have been fixed with four paraformaldehyde, trimmed, dehydrated, embedded in paraffin, sectioned, dewaxed with xylene, stained with hematoxylin and eosin (HE), dehydrated with alcohol, and sealed with resin. The histological changes from the whole tissue sections were observed below a microscope (HE, bar = one hundred , one hundred, and regular areas and obvious lesion places were recorded with a microscope imaging method. two.5. RNA Extraction, cDNA Library Construction, and Sequencing The duodenum, jejunum, ileum, cecum, colon, and rectum of the DIA and CON have been selected for RNA-seq evaluation (n = three). Total RNA was extracted from intestinal tissues of rabbits with diarrhea and healthy handle rabbits by regular extraction process. The effective RNA was screened by Agilent 2100 biological analyzer (Agilent RNA 6000 nano). The screening criteria had been RNA 7.0 Rin value and 28s/18S 1.8. The purified doublestranded cDNA was repaired at the end, an A tail was added, as well as the sequencing connector was connected. The 37020 bp cDNA was screened, amplified by PCR, and purified once again, and also the library was ultimately obtained. The NEBNextUltraTM RNA Library Prep Kit for Illumina(San Diego, CA, USA) [23] was utilised to construct the library. Then Illumina sequencing was performed. two.six. RNA-Seq Data Analyses Image data measured by high-throughput sequencer have been converted into sequence information (reads) by CASAVA base recognition and clean reads have been obtained. Q20, q30, and GC contents of clean reads were calculated (S 1). The transcripts were reconstructed and annotated employing StringTie (1.three.3b) [24]. Additionally, principal element evaluation (PCA) was performed on all samples. DESeq2 [25] application (1.20.0) was applied for differential expression analysis among the two comparative combinations. p-value 0.05, FDR 0.01, and genes with |log2 (foldchange)| 1 had been regarded to become significantly differentially expressed genes. To be able to predict the primary biological and molecular functions of those DEGs, we used gene ontology (GO) classification and functional description. GO includes three elements: molecular function, cellular elements, and biological processes (http://geneontology.org/, accessed on 15 June 2020). Rabbit genes had been annotated by KEGG (Kyoto Encyclopedia of Genes and Genomes) plus the differential gene sets were enriched and analyzed by KEGG pathway. Significance levels for all GO and KEGG terms were corrected by controlling for the false discovery price (FDR) of several pairing comparisons. Statistical software program R (R version four.0.5) and python (Python version 3.9.0) have been utilised for statistical analysis. three. Results 3.1. Rabbit Intestinal Tissue Section The HE-stained intestinal tissue samples (Figure 1) showed that the intestinal samples of rabbits with diarrhea had been mostly manifested in different degrees of necrosis. In serious situations, the entire layer of intestinal wall showed necrosis, plus the quantity of lymphocytes in most intestinal lymph follicles was drastically decreased. In far more significant situations, intestinal coagulative necrosis was the primary outcome, followed by mucosal epithelial necrosis and falling off to type erosion, plus the clinical manifestation was diarrhea. Conversely, the intestinal3.1. Ra
