Pien Tze Huang Alleviates 5-fluorouracil-induced Intestinal Toxicity

Objective:To evaluate the effect of Pien Tze Huang (PZH) on alleviating 5-fluorouracil-induced intestinal mucositis in vivo, and to explore the underlying mechanisms. The aim of this study was to provide theoretical basis for the clinical application of the drug.Methods:After detached through trypsinization. CT-26 cells were resuspended in serum-free RPMI-1640.2×106 cells mixed with matrigel (1:1) were injected subcutaneously into the right flank region of each mouse. When the tumor size reached 300 to 350 mm3, the mice were randomly divided into 4 groups (n=8):control group, PZH group,5-FU group, and 5-FU+PZH group.250 mg/kg of PZH was intragastrically given, daily for 4 days; and 5-FU (150 mg/kg) was administered intraperitoneally only on the first day of treatment. Body weight was measured and diarrhea score was calculated every day. On 5 days, mice were sacrificed by cervical dislocation. The excised jejunum tissues fixed in 4% paraformaldehyde were embedded in paraffin and sectioned at 4 μm thickness. And then haematoxylin and eosin (H&E) staining was used to observe the morphology of jejunum, and the histopath-ology assessment was performed by using a modification of the histopathological scores described by Macpherson & Pfeiffer. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was used to detect cell apoptosis. Immunohistochemical (IHC) staining was performed to determine the expression of Bcl-2 and Bax.Result:Compared with control or PZH alone group,5-FU challenge significantly reduced body weight and caused serious diarrhea in CT-26 xenograft mice, and caused severe histological damages in jejunum. Although PZH did not neutralize body weight loss caused by 5-FU, it significantly alleviated 5-FU-induced diarrhea and intestinal damage in experimental animals. TUNEL assay indicated that 5-FU challenge significantly increased the percentage of TUNEL-positive intestinal crypt cells in xenograft mice, as compared with mice in either control or PZH alone group, which however was reduced by PZH treatment. In addition, data from IHC staining showed that 5-FU significantly reduced anti-apoptotic Bcl-2 protein level in intestinal crypt cells, whereas the level of pro-apoptotic Bax protein was significantly increased after 5-FU challenge. However,5-FU-induced alteration of Bcl-2/Bax expression was remarkably reversed by PZH treatment. No significant difference between control group and PZH alone group.Conclusion:(1) PZH can significantly alleviate 5-FU-induced diarrhea and intestinal mucositis in CT-26 tumor-bearing mice, suggesting that PZH had a protective effect on 5-FU-induced intestinal damage. (2) PZH can significantly inhibit 5-FU-induced apoptosis of intestinal crypt cell through suppression the pro-apoptotic Bax/Bcl-2 ratio, which may prompting PZH can ameliorate 5-FU-induced intestinal damages.