Published online Nov 21, 2020. doi: 10.3748/wjg.v26.i43.6782
Peer-review started: May 7, 2020
First decision: May 15, 2020
Revised: May 28, 2020
Accepted: October 1, 2020
Article in press: October 1, 2020
Published online: November 21, 2020
Intestinal inflammatory disorders are associated with the infiltration of immune cells and the proinflammatory release of cytokines that play a critical role in the onset and progression of colitis-associated cancer (CAC). Recent studies suggested that the intestinal microbiota has an essential role in carcinogenesis. Probiotic supplementation is an alternative means of favorably modulating the intestinal microbiota. Currently, it has become increasingly evident that intestinal microbiota plays a crucial role in the pathogenesis of inflammatory bowel diseases (IBD) and colorectal cancer. Moreover, increasing evidence suggests that probiotics prevent inflammation and carcinogenesis and several bacteria strains have been used for the prevention and treatment of the infectious colitis, IBD. Thus, probiotic modulation of intestinal microbiota has emerged as a potential chemo-preventive agent.
Although supplementation with probiotics have been reported to prevent CAC, little is known about the administration of strains of Lactobacillus bulgaricus (L. bulgaricus), as well as their impact on neoplastic changes in the intestinal mucosa. Our study may contribute to address the gaps in the literature of how this probiotic, dose and supplementation time used for this experimental model impact on colitis, serum cytokines and neoplastic development.
The purpose of this study is to investigate the effect of the probiotic L. bulgaricus during the development of an experimental model of CAC. Overall, this study intents to strengthen data from preclinical studies, encouraging clinical trials to investigate their role in preventing colitis and CAC in humans.
We used an experimental model of CAC. For mice treatment, 1 × 109 CFU were diluted in 200 μL of PBS and orally given to each mouse, 3 times a week during all experimental period. Prior to tumor induction, C57BL/6 mice were randomly distributed in 2 groups (n = 10) and treated with PBS (control group) or L. bulgaricus (Lb group) by gavage (0.2 mL/mouse) for one week. For CAC induction, mice were intraperitoneally (i.p.) injected with a single dose (10 mg/kg in 300 μL solution) of azoxymethene (Sigma-Aldrich), followed by 3 cycles of one week of 2.5% dextran sulfate sodium (DSS) diluted in drinking water intercalated for 2 wk of normal water. Mice were euthanized 12th week after CAC induction. Intestinal inflammation in vivo, or disease score, was determined by scoring clinical signs. The severity of intestinal inflammation was assessed by measuring the length of the entire large intestine. Also, the dimensions of the colorectal tumors were measured with pachymeter and the volumes were calculated by the formula: (width)2 × length/2. For histological analysis, distal colon parts were fixed in 4% p-formaldehyde in phosphate-buffered formalin and unblocked in paraffin. Tissue sections (4.0 μm) were prepared from the paraffin-embedded tissue blocks, stained with hematoxylin and eosin and evaluated in a blinded fashion by an experienced pathologist. Cytokines levels were determined from colon and/or tumor samples by ELISA. Statistical analyses were performed using GraphPad Prism version 6.0. A 2-tailed P value < 0.05 was considered to be statistically significant.
We have shown that L. bulgaricus treatment inhibited the total tumor volume and mean size of tumors. Although we did not observe differences in body weight loss between control and L. bulgaricus-treated, we found differences in clinical signals in L. bulgaricus-treated mice, which showed a lower clinical score on the 13th and 15th days after tumor initiation. In addition to the attenuation of intestinal inflammation score, we observed that the treatment with L. bulgaricus reduced the DSS-induced shortening of the colon. In segments of the large intestine that did not present tumors (inflamed colon) we also observed a reduction of at least 2-fold in the levels of the cytokines TNF-α, IL-1β, IL-23 and IL-17 in L. bulgaricus-treated mice in comparison to controls. In contrast, increased concentrations of IFN-γ were also observed in Lb group. Regarding the cytokines measured in tumor tissues, we observed a pattern similar to that found in the inflamed colon with a negative regulation of proinflammatory cytokines in mice treated with the probiotic and an increase in IFN-γ levels in this group. Overall, these findings highlight the protective effect of L. bulgaricus in the regulation of gut inflammation and preventing CAC development. Thus, further clinical trials are needed to confirm these preclinical insights.
We found an anti-inflammatory role and consequent antitumor effect of L. bulgaricus on CAC that may be used as a promising tool for the prevention and treatment of CAC. In summary, L. bulgaricus treatment during colitis-associated colorectal carcinogenesis model may be responsible for anti-inflammatory and antitumor role by lowering proinflammatory cytokine expression.
The present study has shown that L. bulgaricus inhibited CAC via a negative regulation of intestinal inflammation. Hence, has demonstrates promising evidence on L. bulgaricus probiotic has a preventive potential in CAC development. Therefore, clinical trials are needed to confirm this hypothesis and increase the therapeutic arsenal against CAC.