Irinotecan-induced mucositis manifesting as diarrhoea corresponds with an amended intestinal flora and mucin profile

Abstract

Chemotherapy-induced diarrhoea is a major oncological problem, caused by the cytotoxic effects of cancer chemotherapy. Irinotecan is linked with severe mucositis and diarrhoea, the mechanisms of which remain poorly understood. Bacterial beta-glucuronidase is thought to be involved in the metabolism of irinotecan, implicating the intestinal flora. Intestinal mucins may also be implicated in the development of chemotherapy-induced diarrhoea. Rats were treated with 200 mg/kg of irinotecan and killed at 96, 120 and 144 h. The rats were monitored for diarrhoea. Pathology and immunohistochemical staining was performed. The samples were cultured and faecal DNA was analysed using real-time polymerase chain reaction. Severe diarrhoea was observed from 72 to 96 h. A decrease in body mass was also observed after treatment. Significant changes in goblet cell numbers (both complete and cavitated cells) were observed in the small and large intestines. Changes in MUC gene expression were observed in the small intestine only. Modifications were observed to the intestinal flora profile, especially Escherichia coli, and an increase in the expression of beta-glucuronidase was detected. In conclusion, irinotecan-induced diarrhoea may be caused by an increase in some beta-glucuronidase-producing bacteria, especially E. coli, exacerbating the toxicity of active metabolites. Accelerated mucous secretion and mucin release may also contribute to the delayed onset of diarrhoea.

Figure 1

Clinical signs of mucositis. (a) Incidence of diarrhoea. An increased incidence of diarrhoea is evident from 72 to 96 h after irinotecan treatment. (b) Bodyweight. Mean bodyweight of treated rats is lower than that of untreated rats, with greatest difference between groups at 72–96 h after irinotecan treatment.

Figure 2

Serum electrolyte analysis. Sodium and bicarbonate levels were decreased in treated rats 96 h after treatment. Potassium levels were higher in the treated rats at all time points, and anion gap was raised 96 h after treatment. Osmolality levels were lower in treated rats at all time points investigated.

Figure 3

Histopathological changes seen in the jejunum and colon following treatment with irinotecan. The jejunum showed increased mitotic activity at 96 h, and occasional apoptotic bodies from 96 to 144 h. The colon showed considerable damage from 96 to 120 h, with patchy crypt degeneration, dilated crypts with attenuated epithelium and a few desquamated epithelial polymorphonuclear cells in the lumina. There was also evidence of increased hyperplasia and mitotic figures.

Figure 4

Alcian Blue-PAS staining and goblet cell counts in the (a) jejunum and (b) colon. Counts expressed as mean ± SD. The total number of goblet cells increased over time in the jejunum after irinotecan treatment. The total number of goblet cells in the colon decreased at 96 h.

Figure 5

MUC gene immunostaining in the jejunum. (a) MUC 1 expression in the crypts. MUC 1 expression did not alter significantly in the jejunum. (b) MUC 2 expression in the crypts. The expression of MUC 2 did not alter significantly in the jejunum or colon at any time point investigated. (c) MUC 4 expression in the villi. MUC 4 expression was significantly decreased at 96 h in the jejunum villi. (d) KLF 4 expression in the villi. KLF 4-expressing cells were significantly reduced in the jejunum villi at 96 and 144 h.

Figure 6

Qualitative microbiology results for the jejunum and colon. The majority of changes were seen from 96 to 120 h after treatment. In the jejunum, there were increases in Escherichia spp., Clostridium spp. and Staphylococcus spp. Changes were also seen in the colon with increases in Escherichia spp., Clostridium spp., Enterococcus spp., Serratia spp. and Staphylococcus spp.

Figure 7

Quantitative real-time polymerase chain reaction results showing an increase in Escherichia coli and decrease in Bifidobacterium spp. after treatment. β-glucuronidase expression in the colon is also increased after treatment.