Biochemical characterisation of para-aminophenol-induced nephrotoxic lesions in the F344 rat

The acute biochemical effects of the nephrotoxin p-aminophenol (PAP) were studied in detail using a combination of conventional bioanalytical and ¹H-NMR spectroscopic methods. Dosing PAP (25–100 mg/kg) to male F344 rats resulted in a dose-related proximal nephropathy with consequent elevations in urinary enzymes, glucose, and urine total protein as shown by conventional methodology. ¹H-NMR spectroscopy at 400 MHz of urine from PAP-treated rats also revealed a characteristic glycosuria, with concomitant amino aciduria. The increased excretion of these compounds indicates functional defects in the proximal tubule and reduced solute reabsorption efficiency. In addition, ¹H-NMR urinalysis and conventional enzymatic analysis showed a dose-related lactic aciduria. Other changes detected by ¹H-NMR included a dose-related reduction in the excretion of citrate (confirmed by a conventional biochemical method) and an increase in the excretion of acetate. The degree of abnormalities shown by ¹H-NMR urinalysis agreed well with histopathological observations and conventional biochemical indices of nephrotoxicity. ¹H-NMR urinalysis therefore serves to highlight changes in the excretion of low MW urine components not routinely studied by conventional biochemical analysis.Abbreviations ALP alkaline phosphatase - APAP paracetamol - BUN blood urea nitrogen - GFR glomerular filtration rate - GOT glutamate oxaloacetate transaminase - LAP leucine aminopeptidase - LDH lactate dehydrogenase - MW molecular weight - NAG N-acetyl--D-glucosaminidase - PAP p-aminophenol - ppm parts per million - TMAO trimethylamine N-oxide - UFR urine flow rate     

Unaltered thyroid function in mice responding to a highly immunogenic thyrotropin receptor: implications for the establishment of a mouse model for Graves' disease.

Grave's disease (GD) is a common disorder characterized by the presence of autoantibodies to the thyrotropin receptor. In the past, the exceedingly low expression of the thyrotropin receptor on thyrocytes has not allowed its purification in quantities sufficient to investigate the establishment of an animal model for this disease. In this study, we have purified the 398-amino acid, extracellular region of the human thyrotropin receptor (TSH-R.E) from insect cells using recombinant baculovirus, and explored its immunopathogenic properties in H-2b,d,q,k,s strains of mice. The receptor preparation was highly immunogenic since it elicited strong specific proliferative T cell responses as well as IgG responses in all strains tested. In addition, hyperimmunization with TSH-R.E induced (i) serum antibodies that blocked the binding of 125I-TSH to its receptor, a common feature of GD autoantibodies; and (ii) IgG that reacted with a synthetic peptide (residues 32-54) from the N-terminus of the receptor, a region implicated in the binding of thyroid stimulating antibodies. In SJL animals only, a weak antibody response to two other thyroid antigens, thyroglobulin and thyroid peroxidase, was also observed. The presence of these antibodies, however, was not accompanied by a detectable alteration in thyroid function as assessed by the measurement of serum TSH, T4 and iodine levels. Also mononuclear infiltration of the thyroid gland or morphological changes compatible with an activation state of thyrocytes were not apparent in TSH-R-challenged mice. In contrast, mice treated with the anti-oxidant aminotriazole showed a dramatic increase in serum TSH levels and an activated follicular epithelium. These data demonstrate that a highly immunogenic TSH-R.E in mice does not necessarily provide a proper stimulus for the induction of a hyper- or hypothyroid status as defined by hormonal or histological criteria. Main reasons for the inability to induce receptor-specific antibodies that affect thyroid function such as those generated in GD are likely to be the inappropriate folding of the recombinant extracellular domain of the receptor, or the xenogeneic nature of the autoantigen.     

Progress in the autosomal segmental aneusomy syndromes (SASs): single or multi-locus disorders?

Based on cytogenetic observations, several syndromes have been previously identified as microdeletion-based disorders. In this review, recent progress is presented regarding whether one or multiple genes can be implicated in the pathogenesis of these segmentally aneusomic syndromes. The syndromes discussed include Angelman, Alagille, Williams, Langer-Giedeon, Prader-Willi, Smith-Magenis, Miller-Dieker, and DiGeorge/velocardiofacial or the 22q11 deletion syndromes. For Angelman and Alagille syndromes, single genes have been identified, whereas for Williams and Langer-Giedion syndromes, more than one gene can be implicated. Although there has been significant progress in dissecting the molecular basis for the other disorders, the ultimate answer regarding one versus several genes remains to be determined.      

Skin ligaments: regional distribution and variation in morphology

Skin ligaments (SL) (L. retinacula cutis) are present extensively in the face, hands, feet, and in breast tissue, but have seldom been reported elsewhere in the body. The traditional histological view of the subcutaneous region is that it comprises a matrix of loose connective tissue devoid of fibrous specializations. The purpose of this study was to determine the structure and distribution of skin ligaments. Eight embalmed cadavers (3 males, 5 females, 69-90 years of age) were used in this study. Tissue was prepared using the E12 plastination technique. Macroscopic and microscopic examination demonstrated the widespread presence in the limbs and most of the rest of the body of fibrous strands linking the base of the dermis and the superficial fibers of the underlying deep fascia. The morphology and distribution of these skin ligaments were similar in the individuals examined. Variations in the structure of the skin ligaments depended on the presence of underlying muscle, neurovascular bundles, intermuscular septa and adipose tissue. We conclude that skin ligaments are complex fibrous structures that are present over most of the body. They form an extensive peripheral network in the subcutaneous fat. These 'ligaments' seem to provide an anchorage of skin to deep fascia that is flexible and yet resistant to mechanical loading from multi-directional forces. The use of the E12 plastination technique coupled with fluorescent confocal microscopy has been of benefit in visualizing and delineating SLs from other soft tissue structures in three planes.     

Expression of luminal and basal cytokeratins in human breast carcinoma

We have examined basal and luminal cell cytokeratin expression in 1944 cases of invasive breast carcinoma, using tissue microarray (TMA) technology, to determine the frequency of expression of each cytokeratin subtype, their relationships and prognostic relevance, if any. Expression was determined by immunocytochemistry staining using antibodies to the luminal cytokeratins (CKs) 7/8, 18 and 19 and the basal markers CK 5/6 and CK 14. Additionally, assessment of alpha-smooth muscle actin (SMA) and oestrogen receptor status (ER) was performed. The vast majority of the cases showed positivity for CK 7/8, 18 and 19 indicating a differentiated glandular phenotype, a finding associated with good prognosis, ER positivity and older patient age. In contrast, basal marker expression was significantly related to poor prognosis, ER negativity and younger patient age. Multivariate analysis showed that CK 5/6 was an independent indicator for relapse free interval. We were able to subgroup the cases into four distinct phenotype categories (pure luminal, mixed luminal/basal, pure basal and null), which had significant differences in relation to the biological features and the clinical course of the disease. Tumours classified as expressing a basal phenotype (the combined luminal plus basal and the pure basal) were in a poor prognostic subgroup, typically ER negative in most cases. These findings provide further evidence that breast cancer has distinct differentiation subclasses that have both biological and clinical relevance.