Morphological changes induced by clofibrate in type-1 predominant soleus, type-2 predominant tensor fasciae latae, and type-1 and -2 mixed biceps femoris muscles and diaphragm in rats were investigated. Administration of the agent at 500 or 750 mg/kg/day by oral gavage for 14 or 28 days caused lesions in the soleus muscle and diaphragm, bur no changes in the tensor fasciae latae and biceps femoris muscles. In soleus muscle, vacuolation of muscle fibers was observed in all animals treated with clofibrate, and degeneration of muscle fibers and infiltration of leukocytes were noted at 750 mg/kg/day. In diaphragm, vacuolation of muscle fibers was also observed in all animals treated with clofibrate, and these lesions were located in type-1 skeletal muscles densely stained with NADH-TR. The vacuoles seen in soleus muscle and diaphragm were positive for oil red O staining. In addition, increase of lipid droplets and mitochondrial hypertrophy was seen in soleus muscle, ultrastructurally. These data suggest that sensitivity to clofibrate-induced muscle toxicity differs among muscles, with type-1 fibers being susceptible.

Morphological changes and mRNA expression levels in type-1 predominant soleus and type-2 predominant tensor fasciae latae muscles of rats treated with fenofibrate were investigated. After fenofibrate by oral gavage at 300 mg/kg/day for 28 days, degeneration/necrosis and regeneration of muscle fibers, cellular infiltration, and fibrosis were seen in soleus muscle. Additionally, expression of PDK4, CPT1-M, CPT2, and FACO mRNAs was increased. In contrast, no morphological changes or mRNA induction were apparent in tensor fasciae latae muscle. These data suggest that sensitivity to fenofibrate-induced muscle toxicity differs among muscles, with only type-1 fibers being susceptible. The up-regulation of PDK4, CPTs and FACO mRNA expression in soleus muscle indicates that the energy source is switched from glucose to fatty acids, and this might be related to the observed fenofibrate-induced muscular toxicity.

ABSTRACT. One-step RT-PCR procedure without initial RNA extraction step is tested for laser microdissected tissue sample. Unfixed cryosections of liver and kidney tissue of male SD rats were cut using laser microdissection system and directly used as templates for RT-PCR study. To check the sensitivity, 5, 25, 125, and 625 hepatocytes were cut and put in PCR-tube. After DNase treatment and cDNA synthesis with pd(N)6 random primer, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) cDNAs were amplified by 60 thermal cycles. GAPDH-specific bands were observed at as few as 25 hepatocytes. Specificity of this procedure was tested for hepatocytes, renal tubular epithelium and glomerular tissue using albumin PCR primers. Approximately 250 cells were cut and albumin cDNA was amplified as described above. Albumin specific band was observed only in hepatocytes sample. To apply this approach to quantitative PCR, various numbers of hepatocytes were cut and put in 0.2 mL PCR tube. After reverse transcription and 10 cycles of GAPDH cDNA amplification by regular thermal-cycler, PCR solution was transferred to 96-well plate designed for real-time PCR system, and further 40 cycles were performed. As a result, GAPDH cDNAs were successfully amplified with a good correlation between the number of template hepatocytes and the intensity of PCR signal. From these results, we concluded this approach would be very useful for the expression analysis of microdissected pathology samples. KEY WORDS: laser microdissection, one-step RT-PCR.

No abstract

Abstract:In a young adult (8 weeks old) Sprague-Dawley (SD) rat, a 7 mm-diameter hemispherical nodule with white spots was macroscopically identified on the visceral surface of the spleen. Microscopically, the nodule consisted of areas of enlarged white pulp. The periarteriolar lymphoid sheaths (PALS) area was expanded due to an increase in lymphocytes, but the marginal zone was relatively atrophied. Although minimal compressions and mitotic figures were sporadically observed, there were no structural alterations or atypical cells. Immunohistochemically, there were increased numbers of proliferating cells (Ki-67 positive cells) in the PALS and red pulp areas. In the PALS area, there were increased numbers of T-cells and macrophage/dendritic cells, while in the red pulp area, there were increased numbers of T-cells and B-cell/plasma cells. This lesion, which is believed to be hyperplastic and not neoplastic, was diagnosed as lymphoid hyperplasia, a condition considered to be uncommon in young adult SD rats. (J Toxicol Pathol 2008; 21: 243-247)