**5.1 Background**

It was reported in the late 60's that a single bolus injection of high-dose (250 mg/ kg body weight) induced rampant renal hypertrophy and cell proliferation [33]. A large increase in the number of dividing cells was reported within 18 hours, with the maximum division of cells within the renal medulla occurring at 24-hours post FAN whereas the cortex reached its peak at 26 hours. This appears to be due to renal tubular epithelial damage that appear to be mediated by both the deposition of folic acid crystals and subsequent tubular obstruction as well as direct nephrotoxicity due to high-dose FAN [34]. The histological evidence of tubular injury is accompanied by reduced renal function as measured by circulating creatinine and blood urea nitrogen levels. Both increase in proportion to the dose of FAN, with the 250 mg/kg dose resulting in severe acute kidney injury with significant increase (greater than double) in circulating markers and histological evidence of acute tubular necrosis at 48 hours post injection. If the animals survive, the FAN model is also commonly used to study renal fibrosis (key finding in chronic kidney disease) that develops 7–14 days after injection [14, 35] and continues to progress weeks thereafter and can be exacerbated with repeated administration of FA [36, 37].
