In basic terms, colitis is an inflammation of the large intestine's inner lining. Numerous unpleasant symptoms are brought on by the colon getting irritated by this inflammation.The underlying cause of all the issues is inflammation and swelling of the colon. This inflammation can occur for several reasons, such as autoimmune disorders, infections, and inadequate blood supply to the colon. The symptoms of colitis include cramps, diarrhea (often with blood or mucus), abdominal pain, and even weight loss [1]. Millions of people worldwide are thought to suffer from colitis, yet there is no accurate statistic for this condition. The number of people estimated to have colitis worldwide in 2019 ranges from 3.3 million to 5.5 million. Due to variations in diagnosis and underdiagnosis, it is challenging to determine an accurate number [2]. In the human lower intestinal tract, one kind of naturally occurring short-chain fatty acid is sodium butyrate (SCFA). It gets generated from gut bacteria, microbiota,and fermenting dietary fiber. Some foods, including cheese, butter, and breast milk, also contain sodium butyrate. Among the many possible health advantages of sodium butyrate is that Colonocytes mostly obtain their energy from sodium butyrate. It maintains the colon's health and functionality [3]. Recent researchhas shown that the health of gut microbiota is directly correlated with varied disorders inside the gut and surrounding systems such as the lungs [4, 5], endometriosis [6] and too many others.

Handling of the experimental animals

We used fully grown male rats that were 8 - 9 weeks old and weighed between 200 - 210 g. Housing was typically equipped with controlled temperature, humidity percentage, and light time. Before the rats were acclimated to the study, they underwent two weeks of conditioning. The animals were given free access to commercial pellets and water for the duration of the trial.

Study design

Sodium butyrate (SB) (200 mg/kg BW) was given to rats [7]. Rats received 1% DSS in tap water [8] . Both drugs were given every day for nine days. Ten animals each were assigned to one of four groups: the first one was the control group; the second one received 200 mg/kg of sodium butyrate (SB) daily; the third one received 1% of DSS in tap water; and the fourth one received both DSS and sodium butyrate in tap water.

Sample gathering and Laboratory examinations

Rats were given intramuscular injections of a combination of ketamine 60 mg/kg BW and xylazine 40 mg/kg BW to put them unconscious after the research [9]. To examine some biochemical parameters blood was taken from the eye [10].

Malondialdehyde and glutathione were quantified following the manufacturer's instructions using commercially available ELISA kits (Elabscience). After blood was drawn, colon samples were taken, preserved in buffered formalin, embedded in paraffin, and hematoxylin-eosin-stained for micro-investigation. To verify the expression of the PPAR-γ gene, the spleen was removed [11].

Statistical analysis

One-Way ANOVA in the SPSS was used to detect significant differences between study groups at P<0.05 [12].

Sodium butyrate administration minimizes clinical indicators in rats with DSS-induced colitis

Figure 1-A is a line graph representing the loss/ or gain of weight percentile. Conventionally group C, the control group shows a meager elevation, enlisted as normal/expected, when looking at group SB, which also shows a minor increase. Latterly group D shows an immense drop in weight, by DSS. Importantly onward to group SB+D who show an impediment to weight loss, maintaining similar weight percentile as the day’s progress. 

Figure 1-B represents the colon length after 9 days of administration of sodium butyrate, anddextran sodium sulfate. The colon size may be variable due to extreme conditions, and they read as follows, group C the control group having an average size colon of 16.6 were knowingly normal. Group SB comes slightly below the C group which is insignificant in vivo. However, when looking at group D there is a massive decrease in length due to the high chroniculcerative colitis. When looking at group SB+D there is a slight differentiation and scoring just below the SB group. 

Figure (1): Shown the effect of sodium butyrate, DSS, and their combination in A- weight gain /loss %, B- colon length, and C-Macroscopic Score in adult male rats. Values are expressed as mean ±SE. n=10/each group, * p<0.05

Impact of sodium butyrate treatment on the redox system in rats with DSS-trigged colitis

Figure 2-A is a bar graph representing MDA (malondialdehyde) is essentially a marker used in samples to display studies related to free radicals and, hence substantially suitable for our experiment. In the consecutive groups C and SB, there is a minuscule portion of lipid peroxidation, scoring low as there is no combative substance included, however, when approaching group D there was a massive escalation of MDA levels, predominantly due to the colitis formed from DSS introduction. When looking at group SB+D there is a perceptible increment of MDA levels, however not as significant as the D group alone, on the strength of the attenuative SB. Figure 2-B is a bar graph representing GSH (glutathione) very enlightening antioxidant capable of preventing damage to tissue from peroxides, reactive oxygen species, and of course free radicals. As enlisted, group C shows normal results, however a dramatic uprise in the SB group due to the helpful effect of sodium butyric acid, accordingly,protecting the tissues. Group D has DSS hence showing a momentous shrinkage in presence, exposing the tissues to further oxidative damage. The SB+D group shows an advantageous increase in glutathione, reasonably arduous to maintain and protect the surrounding tissue. 

Figure (2): Shown the effect of sodium butyrate, DSS, and their combination in A- MDA level B- GSH level in adult male rats. Values are expressed as mean ±SE. n=10/each group, * p<0.05

Gene expression in colitis brought on by DSS is altered by sodium butyrate treatment

Figure 3 is a bar graph representing the PPAR-γ (Peroxisome proliferator-activated receptor gamma) triggering, which is crucial in homeostasis and metabolic function and activities. 14 “Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor superfamily that regulate the expression of genes related to lipid and glucose metabolism and inflammation.”. Knowing this we can conclude that PPAR-GAMMA is an essential part of the normal activities of rats and important in normal life functions. When looking at Group C, there is an invariable average score recorded, however, when SB group results were established, a vast increase was recorded. Group D shows a poor score. Group SB+D shows a comparable result with group C in number, with the awareness that the rat was administered DSS we can summarize with vindication.

Figure (3): Shown the effect of sodium butyrate, DSS, and their combination in PPAR-gamma expression in spleen in adult male rats. Values are expressed as mean ±SE. n=10/each group, * p<0.05

Sodium butyrate improves histopathological examination in rats with DSS-induced colitis

Figure 4 represents the pathological examination in rat colon panel A: section of colon mucosal fold (control) showed normal mucosal epithelium (Red arrow), normal tubular glands (Black arrows), and muscularis (Asterisk).Panel B: section of colon (SB) showed normal appearance and size of the fold with normal (Black arrows), the epithelium (Red arrow) & muscularis (Asterisks). Panel C: section of colon (D) showed severe necrotic colitis characterized by necrosis of tubular glands (Red arrows) with severe infiltration of mononuclear leukocytes (Red arrows), epithelial sloughing (Black arrows) & necrotic debris (Blue arrow). Panel D: section of colon (SB+D) showed mild thickening of folds associated with moderate aggregation lymphocytes (Arrow), and mild atrophy of tubular glands.

Figure (4): Shown the effect of sodium butyrate, DSS, and their combination in the colon section (A) There was no visible damage to the colon's cellular structure in the control rats. (B)Rat colonocytes show no pathogenic changes and are normal when given sodium butyrate orally. (C) giving medication to rats of DSS with severe necrotic colitis in the tubular glands and severe infiltration of leukocytes, (D) When rats were administered sodium butyrate orally and DSS in tap water, they had mild thickening of the mucosal folds. H and E, 100×

In the majority of inflammatory bowel disease (IBD) studies, weight loss may be a suitable single outcome measure of DSS colitis with colon shortening. The animal may become less interested in eating if they have symptoms like nausea and stomach pain. Their body may find it more difficult to absorb nutrients from meals if they have diarrhea.Extreme dehydration, a damaged colon, reduced bone density, and inflammation of the skin, joints, and eyes are further effects of ulcerative colitis. These issues can arise over time. Additionally, it may raise your risk of colon cancer and blood clots. Numerous investigations have demonstrated that the disruption of tight junction and mucus layer proteins is linked to intestinal epithelial barrier dysfunction in the pathophysiology of DSS-induced colitis [13]. Propionate, acetate, and butyrate are the SCFAs that are most prevalent. All have been thoroughly investigated in mouse models for potential effects on appetite, elevation in weight, and colon health [14].

Along side with the findings of Yao et al. and Zhao et al. suggest that DSS exacerbates oxidative stress and inhibits the action of antioxidants in acute colitis models. Specifically, colon MPO and MDA levels rose while SOD and GSH activity declined in the colitis group relative to controls [15, 16]. Alike, DSS significantly decreased the activities of endogenous antioxidant enzymes, including SOD, GSH-Px, catalase, and GSH, in the colon tissue of colitis mice [17]. Lipid peroxidation (malondialdehyde) and increased disease activity index (DAI) were the two main causes of the histological alterations brought on by DSS delivery that led to acute colitis [18] A previous study demonstrated that Mice exposed to DSS showed decreased body weight, elevated myeloperoxidase, and malonaldehyde levels, proinflammatory cytokine mRNA levels (TNF-α, IL- 1β, IL-6, and IL-17), and decreased activities of enzymatic antioxidants, Catalase, Glutathione reductase, and Glutathione peroxidase. In a comparable manner, sulfasalazine greatly reduces the DSS-induced rise in MDA levels, bringing them back to levels that are almost under control, according to a different investigation. Reactive oxygen species were reduced and AGE-induced oxidative stress was mitigated through sodium butyrate [19], enhanced superoxide dismutase material, elevated malondialdehyde, and cytokines associated with inflammation interleukin-1β and tumor necrosis factor-α [20]

According to studies, butyric acid can activate PPAR-γ, which raises GLUT4 expression and enhances cells' capacity to absorb glucose and decrease inflammation [21]. A key player in the pathophysiology of ulcerative colitis and colitis-associated malignancies is PPAR-γ. A mouse model of AOM/DSS-induced colitis-associated neoplasia demonstrated that 5‑ASA greatly reduces the tumor burden and colitis and increases the expression of PPAR-γ in the intestinal tract [22]. Peroxisome proliferator-activated receptors γ (PPARγ/NR1C3) are trans-activated by butyrate, which also modulates the PPARγ target gene Angptl4 in gut colonic cells [23]. Neutrophils, platelets, and macrophages can change their production of pro- to anti-inflammatory mediators in response to PPARγ. via changing macrophage trafficking, boosting efferocytosis and phagocytosis, encouraging alternative M2 macrophage activation, suppressing platelet-leukocyte interactions, and promoting neutrophils death [24]. Butyrate prevented the development of colitis brought on by the adoptive transfer of CD4+CD45R T cells in Rag1−/− mice and stimulated the differentiation of Treg cells both in vitro and in vivo. Also, histone H3 acetylation in the conserved non-coding sequence regions and promoter of the Foxp3 locus was enhanced when naive T cells were treated with butyrate under Treg-cell-polarizing conditions the processes by which interactions between microbes and hosts maintain immunological homeostasis in the gut [25]. Oral butyrate considerably enhanced trophism and decreased leukocyte infiltration (neutrophils and eosinophils) in the colon mucosa in ulcerative colitis mice. Additionally, it increased the cecal lymph nodes' inflammatory profile, which included activated B, T, and macrophage cells. The DSS therapy increased interleukin-10 levels, reduced duodenal transforming growth factor-β, and improved dendritic and memory T cells in Peyer's patches, despite the fact that the mucosa histology in the small intestine was equivalent between the groups. Butyrate supplementation was able to undo these modifications [26]. Complete relativity revealed that the DSS group's colon tissue had lost its crypt architecture and had extensive inflammatory lesions throughout the mucosa. while butyrate was able to preserve the structure of the colonic mucosa, improving the epithelial surface, crypt structure, and mucosal integrity [27]. Giving mice TNBS-induced animals and LPS-induced macrophagetreatment with sodium butyrate significantly reduced their inflammatory response [28]. It had suggested that activation of the GPR109A receptor has a protective impact against IBD. Dietary fiber is thought to have a number of positive benefits on intestinal homeostasis maintenance. This is because intestinal microbiota ferments fibers, producing short-chain fatty acids like butyrate, propionate, and acetate [29].

This study concluded that oral sodium butyrate can be used to decreasing the severity of colitis, and upregulated PPAR-gamma gene expression. These findings suggest that sodium butyrate may mitigate the colitis triggered by DSS in rats by boosting the redox system's homeostasis.

Acknowledgment

The University of Baghdad's College of Veterinary Medicine is acknowledged by the authors for helping with animal breeding along with offering experimental animals.

Authors’ Declaration

No Conflicts of Interest

Ethics Clearance

The University of Baghdad's College of Veterinary Medicine's local ethics committee accepted the project and granted permission to carry out this scientific inquiry (file No. 2083/P. G.).

Authors’ contribution

Every experiment was designed by Amira Mohammed. All of the tests were carried out by Fatimah Elias, who also gathered the data and drafted the report. Amira Mohammed helped with data analysis to complete the manuscript in preparation for journal submission. The final draft of the work was reviewed by all authors, who gave their approval for publication.

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