This study investigates the effects of lysine-induced acute renal failure. Female dogs received a lysine hydrochloride (lysine) of 4500 mg/kg/day (3.75 ml/kg/hr) for 3 consecutive days. The dogs were observed for clinical signs. Body weights were recorded, food consumption and water consumption calculated, and urinalysis and blood biochemistry were performed daily. Plasma samples for amino acid determinations were obtained from all dogs, which were necropsied on Day 3. Histopathological examinations were done on all test animals. Compound-related findings include the following. Blood biochemistry results showed increases in ammonia, blood urea nitrogen, blood urea nitrogen/creatinine ratio, and creatinine. Urinary changes consisted of increases in urine volume, total protein, albumin, γ-glutamyl transpeptidase, and N-acetyl-β-D-glucosaminidase. In addition, macroscopic findings consisted of pale, congested capsule; microscopic findings consisted of hypertrophy of proximal convoluted tubule (mainly S1 segment), and degeneration/desquamation of urinary tubule (mainly S3 segment with hyaline casts) in the kidney. From these findings, it can be concluded that lysine is nephrotoxic in dogs. Nephrotoxicity of lysine may relate to direct tubular toxicity and to tubular obstruction.
Airway epithelium is exposed to inhaled exogenous sources. Injury of the alveolar epithelium by cigarette smoking is presumed to be an important process in the pathogenesis of smoking-related pulmonary diseases. Current mechanistic assays that measure the toxicity of cigarette smoke focus on carcinogenesis. However, there is a need to design assays relevant to other disease processes. Oxidative stress is implicated in the pathogenesis of many respiratory diseases including chronic obstructive pulmonary disease. Therefore, we evaluated whether in vitro studies of cigarette smoking are appropriate to examine HO-1 mRNA expression. The human lung epithelial cell line A549 was exposed to the particulate fraction of cigarette smoke (Cigarette Smoke Condensate; CSC) and examined for the induction of HO-1 mRNA. HO-1 gene expression by CSC is increased dose-dependently. In comparison of the induction of HO-1 mRNA by CSC prepared from flue-cured or Burley tobacco, CSC from flue-cured tobacco seems to tend to induce an mRNA of HO-1 higher than CSC from Burley tobacco. The adaptation of HO-1 mRNA expression assay as a biologically relevant indicator of cigarette smoke-induced stress may be exemplified in this study whereby CSC derived from cigarette smoke positively correlated with an increase in HO-1 expression and the difference of the type of tobacco can be detected.
We performed a flow cytometric (FCM) analysis of the maturity of reticulocytes using peripheral blood obtained from rats administered 5-fluorouracil (5-FU) at 10, 50 and 100 mg/kg or acethylphenyl hydrazine (APHZ) at 1 and 3 mg/kg to clarify whether the FCM method is useful for assessing toxicity. In the 5-FU-administered rats, a decrease and recovery of the immature reticulocyte fraction (Cell Maturity Index, CMI; Retic Distribution Index, RDI) was observed more rapidly (several days prior to changes in the reticulocyte ratio), and sensitively regarding dose-dependency (clear changes were observed at 10 mg/kg, whereas the reticulocyte ratio was only slightly affected). In addition, there was good agreement between the microscopic results obtained by counting Heilmyer's reticulocyte maturation groups, especially for type I and II, and CMI/RDI assessed by the FCM method after the administration of 50 and 100 mg/kg of 5-FU, the dose at which clear changes were obtained with the microscopic method. In the APHZ-administered rats, a dose-dependent increase in CMI/RDI coinciding with the enhancement of reticulocyte production was observed. The results suggested that the automated FCM method could be a useful and valuable tool to assess and predict impairments of erythropoiesis, especially for CMI and RDI, and could help in the diagnosis of hematological disorders in experimental animals.
To investigate the toxicity of pierisin-1, a cytotoxic protein present in the cabbage butterfly, Pieris rapae, pierisin-1 was administered via intraperitoneally in mice and rats and the effects examined. Common findings in these experiments were hypoactivity with a gradual decrease in body weight due to decreased food intake, relative polycythemia with low serum albumin concentration and atrophy of the thymus, spleen, seminal vesicles and adipose tissue. Characteristic findings were diarrhea, fusion and atrophy of the villi and dilatation of the crypts in the small intestine at 6-100 μg/kg in BALB/c mice as well as elevation of LDH activity and creatinine value, hemolysis and renal and hepatic injuries at 1,000 and 10,000 μg/kg in BALB/c mice. In the case of ICR mice, severer renal injury was observed. On the other hand, in Fischer 344/Du rats, sudden stop of food intake, elevation of both AST and ALT activities, interlobar adhesion of the right hepatic lobe, capsular thickening, septal fibrosis and single cell necroses of subcapsular hepatocytes in the liver and basophilic tubules in the kidneys were observed. Oral administration of pierisin-1 at a dose of 10,000 μg/kg in BALB/c mice did not exert any obvious effects. Thus, existence of species and strain differences in toxicity of pierisin-1 to animals was demonstrated.
This study was conducted to evaluate the feasibility of total parenteral nutrition (TPN) in rats using continuous intravenous infusion (tail cuff method) via the posterior vena cava. A catheter was inserted into the posterior vena cava from the femoral vein of 10 females (Experiment 1) and 16 females (Experiment 2). The depth of the inserted catheter from the femoral vein was set at 4.5 cm for Experiment 1 and was set at 6 cm for Experiment 2. The test animals were divided into two groups in each experiment: a 5% D-Mannitol (MAN) group and a TPN group. In Experiment 1, TPN rats showed macroscopic lesions (edema in peritoneum, increased collateral vasculature, induration in perivenous tissue, and thrombus) at the tip of the catheter. The diameter of the posterior vena cava (2.86 ± 0.16 mm, mean ± S.D.) was significantly greater than that of the anterior vena cava (2.45 ± 0.22 mm) in 10 rats of Experiment 1. In Experiment 2, TPN rats showed no abnormalities at necropsy. Our findings suggest that TPN administered via the posterior vena cava in Sprague-Dawley rats requires the catheter to be inserted to a depth of 6 cm from the femoral vein. We hypothesize that this is because it is inserted to the level of the renal vein branch where the diameter of the posterior vena cava may be greatest.
The unfolded protein response (UPR) events triggered by the accumulation of unfolded protein in endoplasmic reticulum (ER) activate the three UPR signaling pathways mediated by IRE1, ATF6 and PERK. Spliced XBP1 mRNA induced by activated IRE1 is translated to the protein, a potent transcription factor that induces BiP expression. XBP1 is also induced by activated ATF6. It is thus thought to be an important marker reflecting both IRE1 and ATF6 signaling in response to ER stress. For quantitative measurement of XBP1 gene expression, it is important to distinguish between the spliced and non-spliced form of XBP1 mRNA. We have developed a new method to detect the spliced XBP1 mRNA by means of real-time PCR and we compared the result with measurements of the expression of the ER stress inducible gene BiP. A good correlation was found between spliced XBP1 expression and BiP expression. Thus, our method may be useful for simple and quantitative evaluation of ER stress.
During the characterization of hemorrhagic factor in venom of Rhabdophis tigrinus tigrinus, so-called Yamakagashi in Japan, one of the Colubridae family, a novel metalloproteinase with molecular weight of 38 kDa in the Duvernoy's gland of Yamakagashi was identified by gelatin zymography and by monitoring its proteolytic activity using a fluorescence peptide substrate, MOCAc-PLGLA2pr(Dnp)AR-NH 2, which was developed for measuring the well-known matrix metalloproteinase (MMP) activity. After purification by gel filtration HPLC and/or column switch HPLC system consisting of an affinity column, which was immobilized with a synthetic BS-10 peptide (MQKPRCGVPD) originating from propeptide domain of MMP-7 and a reversed-phase column, the N-terminal amino acid sequence of the 38 kDa metalloproteinase was identified as FNTFPGDLK which shared a high homology to Xenopus MMP-9. The 38 kDa metalloproteinase required Zn2 + and Ca2+ ions for its proteolytic activity. In addition, the proteolytic activity was almost completely inhibited by BS-10, a MMP inhibitor, but not by the serine proteinase inhibitors, cysteine proteinase inhibitors and aspartic proteinase inhibitors.Together these results demonstrated that the 38 kDa proteinase is a novel snake verom metalloproteinase (SVMP) containing HExGHxxGxxH motif which possesses high affinity to the BS-10 peptide, into its molecule, and the enzymatic properties are closed to that of MMPs. Based on the results obtained in the present study, we concluded that the 38 kDa metalloproteinase is a novel metalloproteinase whose activity may be regulated by the cysteine switch mechanism, and could be classified as one of the matrix metalloproteinases rather than snake venom metalloproteinases.
This study's aim is to investigate the relationship between stage and degree of restricted feeding during the gestation period and occurrence of abortion, premature birth or fetal damage in rabbits. The study was composed of 5 groups of pregnant Kbl:NZW rabbits that consisted of 8 animals each. These groups were subjected to restricted feeding in the following ways: (A) control group, free access to food, (B) 60 g per day from gestational days (GD) 6 to 18 (middle period), (C) 20 g per day from GD 6 to 18, (D) 20 g per day from GD 19 to 28 (post-middle period), and (E) 20 g per day from GD 6 to 28 (middle and post-middle periods). Even though all dams in Groups A, B and C went to full term, abortion or premature birth occurred to 2/8 and 8/8 dams in Groups D and E, respectively. Fetal lethality increased in Group C, which was subjected to restricted feeding at 20 g/head/day in the middle period. Slight inhibition of fetal growth was recorded only in Group D, which was subjected to restricted feeding in the post-middle period. Restricted feeding at 20 g/head/day in the middle period induced no abortion or premature birth, but increased fetal lethality that in the middle and post-middle periods resulted in abortion or premature birth of all dams, and that in the post-middle period resulted in abortion or premature birth at low incidence and slightly inhibited fetal growth. These results demonstrated that the post-middle period is vulnerable to effects of reduced food consumption in pregnant rabbits.