The effects of 4-aminopyridine on focal nerve conduction block

In order to test whether 4-aminopyridine (4-AP), a potassium channel blocker, may be of therapeutic value in demyelinating neuropathies, a focal tibial nerve conduction block with demyelination was produced in adult rats by an intraneural microinjection of potassium tellurite. Onset and recovery of the lesion were monitored by evoked compound muscle action potentials (CMAP) activated from the proximal and distal nerve one day before and 1, 4, 7, 14, 21 and 28 days after the injection. Intraperitoneal 4-AP (2 mg/kg) or buffered saline were injected prior to the potassium tellurite and 6 days per week for 28 days. The data show that 4-AP is tolerated, it does not prevent conduction block, and only has a modest effect on increasing its recovery from day 4 to 7 (91 % increase in CMAP ratio compared with control of 35%). Recovery is similar by day 28 in 4-AP treated or untreated animals. These results suggest that 4-AP will have limited use in the therapy of subacute demyelinating neuropathies.     

Comparison of DNA adducts and sister chromatid exchange in lung cancer cases and controls

In a molecular epidemiological study of lung cancer cases (n = 81) and noncancer controls (n = 67), polycyclic aromatic hydrocarbon (PAH)-DNA adducts were evaluated in peripheral blood leukocytes from all subjects and in a smaller number of lung tissue specimens collected prior to or at surgery. Sister chromatid exchanges (SCE) in lymphocytes were also studied in a subset of cases and controls. Questionnaire, medical record, or tumor registry data provided a family history of cancer, as well as information on cigarette smoking, dietary and occupational exposure to PAHs, and other factors related to SCEs. In both cases and controls PAH-DNA adducts in leukocytes measured by an enzyme-linked immunosorbent assay were not significantly related to age, sex, ethnicity, amount of cigarette smoking, passive smoking, dietary charcoal, or caffeine consumption. Nor did family history of cancer or histological type of cancer significantly affect adduct levels. However, when subjects were stratified by smoking status (current, former, and nonsmoker), lung cancer cases who were current smokers had significantly higher levels of covalent adducts than current smoker controls. A seasonal variation was observed in PAH-DNA binding, with a peak in adduct levels during July-October. This peak corresponds to that seen in a prior study of aryl hydrocarbon hydroxylase inducibility by other investigators. The finding of significant levels of PAH-DNA adducts in former smokers and non-smokers supports an earlier observation that this marker is not smoking specific but reflects a pervasive and variable "background" exposure to PAH. These results are consistent with a genetically determined enhancement of PAH-DNA adduct formation in leukocytes of lung cancer cases which is evident in current smokers. The results in lung tissue are limited by the small number of samples. Adduct levels were not significantly increased in lung tissue of smokers compared with nonsmokers. An inverse linear correlation was seen between adduct values in lung tissue and age of the donors. SCEs were significantly related to pack years of smoking. However, there was no difference in the frequency of SCE between cases and controls; nor were SCE and DNA adducts significantly correlated in this small sample.     

Role of mast cells in experimental anti-glomerular basement membrane glomerulonephritis

Recently, divergent reports on the role of mast cells (MC) in different glomerular diseases have brought our attention to their role in an accelerated model of anti-glomerular basement membrane (GBM) glomerulonephritis (GN). Genetically MC-deficient Kit(W)/Kit(W-v) mice, MC-reconstituted Kit(W)/Kit(W-v) mice and Kit+/+ control mice were subjected to anti-GBM GN. Kit(+/+) mice developed moderate proteinuria and glomerular damage following the induction of anti-GBM nephritis. In contrast, proteinuria and glomerular damage were dramatically increased in MC-deficient Kit(W)/Kit(W-v) mice. MC-reconstituted Kit(W)/Kit(W-v) mice showed proteinuria and glomerular damage comparable to Kit+/+ mice. A significant increase in infiltrating T cells and macrophages was detected in MC-deficient Kit(W)/Kit(W-v) mice as compared to Kit+/+ control mice and MC-reconstituted Kit(W)/Kit(W-v) mice. Accordingly, we observed an increase of TGF-beta1 mRNA in kidneys from Kit(W)/Kit(W-v) mice. Interestingly, we did not detect MC in the kidney using either Giemsa staining or RT-real-time PCR, but MC were found in the regional lymph nodes. Finally, mortality of Kit(W)/Kit(W-v) mice was significantly increased after the induction of anti-GBM GN due to uremia. Our report provides the first direct evidence that MC are protective in anti-GBM GN, possibly by modulating the influx of effector T cells and macrophages to inflammatory sites in the kidney.     

Functional characterization of mouse lymphocyte subpopulations identified by their natural binding of bacteria. II. Identification of subpopulations of LY-1 + 2-3-, LY-1-2+3+ and LY-1+2+3+ cells and the localization of specific cytotoxic cells in a subset of LY-1-2+3+ cells .

Three T-cell subpopulations (T1, T2 and T3) can be identified by their binding of various bacteria (Mayer, Chen, Dray & Teodorescu, 1978). In this work we determined how the three subpopulations identified by their Ly-1, -2 and -3 alloantigens were distributed among the T1, T2 and T3 subpopulations. We found that the T1 subpopulation contained most of the Ly-1+2+3+ cells, that the T2 subpopulation contained some Ly-1+2-3- and some Ly-1-2+3+ cells and that the T3 subpopulation contained the remainder of the Ly-1+2+3+, Ly-1+2-3- and Ly-1-2+3+ cells. Thus the subpopulations identified by their bacterial adherence properties subdivided the three subpopulations identified by their Ly-1, -2 and -3 alloantigens. We also investigated whether the specific cytotoxic T lymphocytes were contained in the T1, T2 and/or T3 cells. We found that essentially all of the cytotoxic T lymphocytes were contained in the T3 subpopulation. Since the T3 cells contained a subpopulation of Ly-1-2+3+ cells the data indicated that essentially all of the cytotoxic T lymphocytes were contained in a subpopulation of Ly-1-2+3+ cells.     

Human osteoblast response to pulsed laser deposited calcium phosphate coatings

Octacalcium phosphate (OCP) and Mn(2+)-doped carbonate hydroxyapatite (Mn-CHA) thin films were deposited on pure, highly polished and chemically etched Ti substrates with pulsed laser deposition. The coatings exhibit different composition, crystallinity and morphology that might affect their osteoconductivity. Human osteoblasts were cultured on the surfaces of OCP and Mn-CHA thin films, and the cell attachment, proliferation and differentiation were evaluated up to 21 days. The cells showed a normal morphology and a very good rate of proliferation and viability in every experimental time. Alkaline phosphatase activity was always higher than the control and Ti groups. From days 7 to 21 collagen type I production was higher in comparison with control and Ti groups. The level of transforming growth factor beta 1 (TGF-beta1) was lower at 3 and 7 days, but reached the highest values during following experimental times (14 and 21 days). Our data demonstrate that both calcium phosphate coatings favour osteoblasts proliferation, activation of their metabolism and differentiation.     

Effect of tissue fixatives on telomere length determination by quantitative PCR

Telomere length is a well established marker of cellular senescence and thus biological age. Quantitative PCR allows the determination even from very low amounts of tissue by using telomere specific and single copy gene primers. Comparing a directly processed tissue sample to a 4% formaldehyde fixed one showed a significantly reduced efficiency of PCR reactions (mainly in single copy gene experiments) in a storage time-dependent manner resulting in an artificial increase in reported relative telomere length. This effect was not seen when the tissue was stored in RNA later solution. In summary, telomere length determination from formaldehyde fixed material by quantitative PCR is not a reliable method. Unfortunately therefore, many easily accessible tissue samples from pathology laboratories are unsuitable for this technique.     

A new local multiscale Fourier analysis for medical imaging

The Stockwell transform (ST), recently developed for geophysics, combines features of the Fourier, Gabor and wavelet transforms; it reveals frequency variation over time or space. This valuable information is obtained by Fourier analysis of a small segment of a signal at a time. Localization of the Fourier spectrum is achieved by filtering the signal with frequency-dependent Gaussian scaling windows. This multi-scale time-frequency analysis provides information about which frequencies occur and more importantly when they occur. Furthermore, the Stockwell domain can be directly inferred from the Fourier domain and vice versa. These features make the ST a potentially effective tool to visualize, analyze, and process medical imaging data. The ST has proven useful in noise reduction and tissue texture analysis. Herein, we focus on the theory and effectiveness of the ST for medical imaging. Its effectiveness and comparison with other linear time-frequency transforms, such as the Gabor and wavelet transforms, are discussed and demonstrated using functional magnetic resonance imaging data.     

Focus assay for varicella-zoster virus in human embryo cells stained with immunoperoxidase method

Rapid titration of varicella-zoster virus (VZV) in human embryonic fibroblasts (HEF) based on staining of virus-infected cells by indirect immunoperoxidase technique (IPA) is described. Cell monolayers were grown in wells of plastic plates (two different diameters). Foci of virus-infected cells as revealed by IPA could be counted either 48 hr post-infection, if cell-associated virus (VZV infected cells) was used as inoculum, or 72 hr p. i. if cell-free virus was used. A linear relationship was observed between virus dilution and number of foci. The first virus was detected 12 hr p. i., the highest titre at 36 hr, when cytopathic effect (CPE) involved about 50% of the monolayer.