Thoracolumbar supernumerary ribs (TSRs) are classified as less severe skeletal anomalies in rat developmental toxicity studies, although their incidence is relatively high in rodent studies. To investigate the characteristics of the critical window for chemically-induced TSR, in this study, rats were administered 5-fluorocytocine (5-FC) or sodium salicylate (SAL) at one of three time periods on gestational day (GD) 9, early morning (7:00 AM), midday (12:00 PM to 1:00 PM), or late afternoon (4:00 PM or 7:00 PM). The incidence of TSR and other anomalies were assessed in GD20 fetuses. A single treatment with both chemicals on GD9-induced TSR, with the incidence highest when administered at 7:00 AM, decreasing gradually when administered later. This trajectory was clearer in rats treated with 5-FC than with SAL. The critical period of TSR induction is shorter in rats administered 5-FC than SAL. The characteristics of the critical window may cause variability in the incidence of TSR observed in developmental toxicity studies. K E Y W O R D S 5-fluocytocine, critical period, rats, sodium salicylate, thoracolumbar supernumerary rib 1 | INTRODUCTION Several types of fetal anomalies encountered in developmental toxicity studies can occur spontaneously, and are not because of chemical exposure. Understanding the toxicological characteristics of these anomalies is important to determine whether these anomalies indicate a chemical-induced developmental toxicity or a spontaneous finding.Thoracolumbar supernumerary ribs (TSRs) have been classified as less severe skeletal anomalies, with a relatively high spontaneous incidence in rat fetuses. 1-4 Current historical control data in Japan have shown that the spontaneous incidence of TSR is 0.09% to 15.1% in Sprague-Dawley (SD) rats, and 17.8% to 58.1% in Wistar Hannover rats. 2 TSR can also be induced in rodent fetuses by maternal stress, 5 maternal hypoglycemia, 6 and exposure to chemicals and herbs during pregnancy. 7-11 If the incidence of TSR is dependent on dose of a chemical, the tested item should be regarded as a risk factor for TSR induction.The rates of chemically-induced TSR vary considerably, perhaps because of differences in fertility times of dams. Alternatively, this variability may also be because of the existence of a relatively narrow critical window. This study therefore assessed TSR induced in rat fetuses by a single treatment on gestational day (GD) nine with two teratogenic chemicals, 5-fluorocytocine (5-FC) and sodium salicylate (SAL). 9,12,13 To determine characteristic of critical window for TSR, these chemicals were administered at three different times on GD9. | MATERIALS AND METHODS | AnimalPregnant SLC:SD rats were obtained from SLC Japan, Inc. (Shizuoka, Japan), and housed individually in an environmentally controlled animal room. Lights were on from 7:00 AM to 7:00 PM, and rat chow (CE-2, CLEA Japan Inc., Tokyo, Japan) and drinking water were provided ad libitum. The day on which sperm were observed, was designated as GD0. On GD9, 5-FC or SAL was ad...

-5-Fluorocytosine (5-FC) is an antimycotic and teratogenic compound. Oral administration of 5-FC to pregnant rats on gestation days (GD) 9 and 13 was shown to induce thoracolumbar supernumerary ribs (TSR, 14th rib) and abnormal digits, respectively, in fetuses. This study investigated the effects of 5-FC on homeobox genes, which control the anterior-posterior-axis. 5-FC (75 mg/kg) was administered orally on GD9 and GD13, and tissues collected from cranial and caudal regions of TSR sites were analyzed. Following 5-FC administration on GD9, the levels of expression of Hoxa10, which determine the position of the thoracic and lumbar vertebrae, were decreased at GD13. Analysis of hindlimbs 6 hours after administration on GD13 showed decreases in expression of Hoxa11, Hoxd12, and Hoxd13, the Hox genes responsible for limb formation from the proximal to distal, and from the anterior to posterior directions. The present findings showed that altered expression of Hox genes contributes to 5-FC teratogenicity.