Alogliptin: a novel approach against cyclophosphamide-induced hepatic injury via modulating SIRT1/FoxO1 pathway

Abstract

Cyclophosphamide (CP) is one of the most potent alkylating agents and is widely used in the treatment of numerous neoplastic conditions, autoimmune diseases and following organ transplantation. Due to its ability to induce oxidative stress and subsequent apoptosis, CP is affiliated with many adverse effects with special emphasis on the highly prevalent hepatotoxicity. Dipeptidyl peptidase 4 (DDP-IV) inhibitors are being rediscovered for new biological effects due to their ability to target multiple pathways, among which is the phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt) axis. This could offer protection to multiple organs against reactive oxygen species (ROS) through modulating sirtuin 1 (SIRT1) expression and, in turn, inactivation of forkhead box transcription factor of the O class 1 (FoxO1), thus inhibiting apoptosis. Accordingly, the current study aimed to investigate the potential therapeutic benefit of alogliptin (Alo), a DPP-IV inhibitor, against CP-induced hepatotoxicity through enhancing PI3K/Akt/SIRT1 pathway. Forty male Wistar rats were randomly divided into four groups. The CP-treated group received a single dose of CP (200 mg/kg; i.p.). The Alo-treated group received Alo (20 mg/kg; p.o.) for 7 days with single CP injection on Day 2. Alo successfully reduced hepatic injury as witnessed through decreased liver function enzymes, increased phospho (p)-PI3K, p-Akt, superoxide dismutase (SOD) levels, SIRT1 expression, p-FoxO1 and anti-apoptotic B-cell lymphoma 2 (Bcl-2). This resulted in decreased apoptosis, as witnessed through decreased caspase-3 levels and improved histopathological picture. In conclusion, the current study succeeded to elaborate, for the first time, the promising impact of Alo in ameliorating chemotherapy-induced liver injury.

Keywords: FoxO1; SIRT1; alogliptin; cyclophosphamide; hepatotoxicity.

Graphical Abstract

Figure 1

Effect of alogliptin (Alo) on serum (A) alanine transaminase (ALT), and (B) aspartate transaminase (AST) in cyclophosphamide (CP)-induced liver toxicity in rats. Data are presented as the mean ± SD (n = 8 per group; one-way ANOVA followed by Tukey’s multiple comparison test; ^***P < 0.001, vs. the control group; ^###P < 0.001, vs. the CP-treated group).

Figure 2

(A) Representative western blot bands of hepatic p-phosphoinositide 3–kinase (p-PI3K) (Tyr607), p-protein kinase B (p-Akt) (Thr450) and p-forkhead box transcription factor of the O class 1 (p-FoxO1) (Ser249) protein levels. (B) Quantitation of hepatic p-PI3K, p-Akt, and p-FoxO1 protein levels in control, alogliptin (Alo) (20 mg/kg/day; p.o.), cyclophosphamide (CP) (200 mg/kg single dose; i.p.) and CP + Alo-treated groups. Data are presented as the mean ± SD (n = 8 per group; one-way ANOVA followed by Tukey’s multiple comparison test; ^***P < 0.001, vs. the control group; ^###P < 0.001, vs. the CP-treated group).

Figure 3

Effect of alogliptin (Alo) on gene expression of (A) sirtuin 1(SIRT1), and levels of (B) superoxide dismutase (SOD) and (C) B-cell lymphoma 2 (Bcl-2), and (D) caspase-3 activity in cyclophosphamide (CP)-induced liver toxicity in rats. Data are presented as the mean ± SD (n = 8 per group; one-way ANOVA followed by Tukey’s multiple comparison test; ^***P < 0.001, vs. the control group; ^###P < 0.001, vs. the CP-treated group).

Figure 4

Photomicrographs of liver tissue sections. (a, b): Control and alogliptin (Alo)-treated groups show normal hepatocellular architecture portrayed through normal hepatic cords (HC) arrangement around the central vein (CV), separated by blood sinusoids (S), polyhedral eosinophilic granular hepatocytes with central rounded vesicular one or two nuclei (white arrows). The portal triad (P) at the corner of hepatic lobule shows a branch of hepatic artery, portal vein and bile duct (black arrow). (c): Cyclophosphamide (CP)-treated group displays disturbed hepatic lobular architecture. Congestion of both central (CV) and portal veins is observed. Marked mononuclear leukocyte infiltration (L) at the portal tract area (black star) and within the congested blood sinusoids (S) is noticed. Widespread vacuolated-ballooned hepatocytes in the form of microvacuolization of cytoplasm with centrally placed pyknotic nuclei (black arrows) are detected. Hepatocytes at portal tract area displayed apoptotic changes in form of dark eosinophilic cytoplasm and small deeply stained pyknotic nuclei (white arrowhead). (d): CP + Alo-treated group shows a potentially alleviated hepatocellular architecture compared to control group and minimal changes in the form of few vacuolated hepatocytes (white arrowhead) (H & E stain, X200).