PMP22 overexpression causes dysmyelination in mice.

Charcot-Marie-Tooth (CMT) disease is the most frequent hereditary peripheral neuropathy in humans. Its prevalence is about one in 2500. A subform, CMT1A, is transmitted as an autosomal dominant trait. An estimated 75% of patients are affected. This disorder has been shown to be associated with the duplication of a 1.5 Mb region of the short arm of chromosome 17, in which the PMP22 gene has been mapped. We have constructed a murine model of CMT1A by inserting into the murine genome a human YAC containing peripheral myelin protein 22 (PMP22) and its flanking controlling elements. We describe the behaviour of the C22 line (seven copies of YAC, 2.1 times PMP22 overexpression) during the myelination process. Electron microscopy, morphometry, electrophysiology, nerve conduction and expression of specific markers (e.g. Krox20) in normal and pathological Schwann cells demonstrated that PMP22 overexpression leads to a defect in the myelination of axons. The largest axons are the most affected. Only a few demyelination/remyelination processes were observed. Moreover, PMP22 overexpression probably enhances collagen synthesis by fibroblasts, before myelination, demonstrating that structures other than Schwann cells are affected by PMP22 overexpression. Classically, CMT1A was thought to be induced by a demyelination process following a phase of normal myelination, yet our data suggest that dysmyelination should be considered as a major factor for the disease.

Fish oil supplementation prevents diabetes-induced nerve conduction velocity and neuroanatomical changes in rats.

Diabetic neuropathy has been associated with a decrease in nerve conduction velocity, Na,K-ATPase activity and characteristic histological damage of the sciatic nerve. The aim of this study was to evaluate the potential effect of a dietary supplementation with fish oil [(n-3) fatty acids] on the sciatic nerve of diabetic rats. Diabetes was induced by intravenous streptozotocin injection. Diabetic animals (n = 20) were fed a nonpurified diet supplemented with either olive oil (DO) or fish oil (DM), and control animals (n = 10) were fed a nonpurified diet supplemented with olive oil at a daily dose of 0.5 g/kg by gavage for 8 wk. Nerves were characterized by their conduction velocity, morphometric analysis and membrane Na, K-ATPase activity. Nerve conduction velocity, as well as Na,K-ATPase activity, was improved by fish oil treatment. A correlation was found between these two variables (R = 0.999, P < 0.05). Moreover, a preventive effect of fish oil was observed on nerve histological damage [endoneurial edema, axonal degeneration (by 10-15%) with demyelination]. Moreover, the normal bimodal distribution of the internal diameter of myelinated fibers was absent in the DO group and was restored in the DM group. These data suggest that fish oil therapy may be effective in the prevention of diabetic neuropathy.

Digitization of microcalcifications in breast radiographs. Correlation with pathologic data.

A series of 104 nonpalpable breast lesions detected by mammograms and containing microcalcifications were studied. Intraoperative radiographs of intact and sliced specimens were assessed, followed by microscopic diagnosis on frozen sections of areas containing microcalcifications. Microcalcifications detected on mammograms and radiographs of the specimens were digitized and evaluated in accordance with morphometric parameters, including the mean surface, shape factor, bend energy, envelope surface and total surface, total number and concentration of microcalcifications. Benign disorders, atypical hyperplasia and carcinomas accounted for 47.2%, 4.8% and 48% of the tissue lesions, respectively, but the disorders were most often heterogeneous and mixed. Most, but not all, parameters significantly correlated with the three types of radiographs, although radiographs of the sliced specimens provided images of the best quality. Only two parameters, mean size and bend energy, were significantly different (P = .008, .0036) in benign and malignant lesions. It is concluded that image analysis of digitized microcalcifications in radiographs may provide quantitative data helpful in mammogram interpretation.

Morphological polarization of human polymorphonuclear leucocytes in response to three different chemoattractants: an effector response independent of calcium rise and tyrosine kinases.

Chemoattractants such as interleukin-8, C5a and N-formylmethionyl-leucyl-phenylalanine induce a cytosolic calcium rise involved in triggering the secretory functions of human polymorphonuclear leucocytes. We studied the possible role of calcium rise in membrane ruffling, actin polymerization, filamentous actin distribution, and morphological polarization, which are all events contributing to chemotaxis. Membrane ruffling was assessed by right-angle light-scatter changes, the cellular content of polymerized actin by fluorescence of bodipy phallacidin, the intracellular distribution of filamentous actin by fluorescence microscopy and image digitization, and morphological polarization by scanning electron microscopy. Pretreatment of polymorphonuclear leucocytes with 50 microM BAPTA/AM, an intracellular calcium chelator, lowered the basal level in cell calcium and inhibited the transient calcium rise stimulated by 2 nM interleukin-8, 2 nM C5a, and 10 nM N-formylmethionyl-leucyl-phenylalanine. However, BAPTA pretreatment of polymorphonuclear leucocytes did not modify membrane ruffling, actin polymerization, filamentous actin distribution, and morphological polarization stimulated by chemoattractants. Downstream effectors may be protein tyrosine kinases. However, the tyrosine kinase inhibitor tyrphostin did not affect the cytoskeletal characteristics elicited by chemoattractants. Taken together, our results suggest that the transductional pathway leading to cytoskeleton organization and morphological polarization of polymorphonuclear leucocytes is different from that leading to secretion.

Effects of castration and testosterone on Fel dI production by sebaceous glands of male cats: II--Morphometric assessment.

A morphometric study of cat sebaceous glands was performed to evaluate the effects of castration and testosterone treatments. Skin biopsies were taken in six cats before castration, after castration and after the testosterone injections administered after castration (total number of biopsies: 18). Ninety 8 microns thick sections of each biopsy were assessed for image analysis processing (SAMBA 2005, ALCATEL TITN). The variations in glands and cells size were evaluated on digitized microscopic images by morphometric parameters included in the SAMBA software package. An original software was developed for the analysis of the spacial gland structure. The best morphometric parameters were selected in a first step of the study, and included the nuclear surface (NS), the cell surface (CS) and the nuclear/cellular surface ratio (N/C). These three parameters were then compared in each group of samples for the six cats. It was shown that after castration the N/C (21%) significantly increased compared with prior to castration (12.6%). This 59.8% increase was mainly due to cell cytoplasm shrinking reflecting a decrease of the cell activity. The testosterone administered after castration produced a reverse effect with a N/C ratio back to normal (11.4%) and a significant cell cytoplasm and gland enlargement as shown by the three dimension constructions. This morphometric data correlated with the measurement of sebum and Fel dI productions. The negative effects of castration and the positive effects of testosterone on the sebaceous cells and glands volume favour the hypothesis that cat sebaceous cells are subject to hormonal control this is also likely to apply to the Fel dI production.

X-raying of sliced surgical specimens during surgery: an improvement of the histological diagnosis of impalpable breast lesions with microcalfications.

Surgical specimens, from 78 patients operated for impalpable breast lesions containing microcalcifications detected on mammographs (breast cancer screening program) were systematically examined by a pathologist during the surgical intervention. The per-operative procedures encompassed (i) X-raying of the unfixed specimens to ascertain that the latter did enclose the screened lesion, (ii) slicing and subsequent X-raying of the specimens slices in order to locate the microcalcifications, (iii) histological evaluation of areas containing the microcalcifications on frozen sections. Thirty-two of the lesions were histologically benign, 55% were malignant and 13% borderline. Forty of the carcinomas were in situ and 60% were invasive. The per-operative histological diagnosis was correct in 65% of the case, erroneous in 10% and uncertain in 25%. Malignancy was never overscored. In 65% of the carcinomas diagnosed during the surgical intervention, the in sano margins excision and axillary lymph node removal could be performed (one stage surgical treatment). These results suggest that X-raying of sliced specimens and histological evaluation during the surgical intervention ascertains that lesions screened by mammographs are effectively and completely removed, and that they are precisely and extensively histologically examined after being identified and located.