Structural Artifacts and Advantages of Cytocentrifugation of Cells as Viewed by Scanning Electron Microscopy

Cytocentrifugation of cell suspensions onto glass slides is a widely used procedure in contemporary cytology. We employed here scanning electron microscopy (SEM) to investigate putative morphological changes induced in cells submitted to cytocentrifugation. The fine structure of murine pleural exudate cells (macrophages mainly) processed by spinning was compared with that of similar cells treated without centrifugation (poly-L-lysine attachment of the cells to glass slides at 1 g). Cells of cytocentrifuged preparations showed a significant increase in diameter and smoothening of the cell surface as compared with the morphology of non-centrifuged cells. Cytocentrifugation also induced the formation of thin elongations coming out of the cellular outlines. The centrifugation-induced flattening of the pleural macrophages improved the detection of large intracellular inclusions (containing tungsten particles): these bodies were readily identified by secondary-electron imaging mode of SEM in cytospinned cells whereas their detection in non-centrifuged spherical cells required the use of the backscattered-electron imaging mode of SEM. We conclude that the cytocentrifugation methodology, on one hand, requires caution on the interpretation of the microanatomy of the cells and, on the other hand, the procedure may be an adequate method to improve the identification of large intracellular inclusions by routine (secondary-electron imaging mode) SEM

Effects of low-frequency noise on cardiac collagen and cardiomyocyte ultrastructure: an immunohistochemical and electron microscopy study

"Introduction: Low-frequency noise (LFN) leads to the development of tissue fibrosis. We previously reported the development of myocardial and perivascular fibrosis and a reduction of cardiac connexin43 in rats, but data is lacking concerning the affected type of collagen as well as the ultrastructural myocardial modifications. Objectives: The aim of this study was to quantify cardiac collagens I and III and to evaluate myocardial ultrastructural changes in Wistar rats exposed to LFN. Methods: Two groups of rats were considered: A LFN-exposed group with 8 rats continuously submitted to LFN during 3 months and a control group with 8 rats. The hearts were sectioned and the mid-ventricular fragment was selected. After immunohistochemical evaluation, quantification of the collagens and muscle were performed using the image J software in the left ventricle, interventricular septum and right ventricle and the collagen I/muscle and collagen III/muscle ratios were calculated. Transmission electron microscopy (TEM) was used to analyze mid-ventricular samples taken from each group. Results: The collagen I/muscle and collagen III/muscle ratios increased in totum respectively 80% (p<0.001) and 57.4% (p<0.05) in LFN-exposed rats. TEM showed interstitial collagen deposits and changes in mitochondria and intercalated discs of the cardiomyocytes in LFN-exposed animals. Conclusions: LFN increases collagen I and III in the extracellular matrix and induces ultrastructural alterations in the cardiomyocytes. These new morphological data open new and promising paths for further experimental and clinical research regarding the cardiac effects of low-frequency noise.

Arrest in ciliated cell expansion on the bronchial lining of adult rats caused by chronic exposure to industrial noise

Workers chronically exposed to high-intensity/low-frequency noise at textile plants show increased frequency of respiratory infections. This phenomenon prompted the herein investigation on the cytology of the bronchial epithelium of Wistar rats submitted to textile noise. Workplace noise from a cotton-mill room of a textile factory was recorded and reproduced in a sound-insulated animal room. The Wistar rats were submitted to a weekly schedule of noise treatment that was similar to that of the textile workers (8 h/day, 5 days/week). Scanning electron microscopy (SEM) was used to compare the fine morphology of the inner surface of the bronchi in noise-exposed and control rats. SEM quantitative cytology revealed that exposure to noise for 5-7 months caused inhibition in the natural expansion of the area occupied by ciliated cells on the bronchial epithelium as adult rats grow older. This difference between noise-exposed and age-matched control rats was statistically significant (P<0.05) and documents that the cytology of the rat bronchial epithelium is mildly altered by noise exposure. The decrease in the area of bronchial cilia may impair the mucociliar clearance of the respiratory airways and, thus, increase vulnerability to respiratory infection

Infrasound exposure promotes development of atrial fibrosis in rats

INTRODUCTION: Recent data has shown a significant association between noise exposure and atrial fibrillation (AF) in a large cohort [1] but the pathophysiology remains unclear. The acoustic spectrum of industrial environments is particularly rich in high-intensity infrasound (IFS), which we have previously found to induce coronary perivascular fibrosis in rat hearts [2–4]. The role of atrial fibrosis in AF is well documented and remains the cornerstone of atrial pathology in patients with this arrhythmia [5]. The aim of this study was to evaluate and measure the atrial interstitial fibrosis in rats exposed to high-intensity IFS. MATERIAL AND METHODS: Twelve Wistar rats exposed to high-intensity IFS (110 dB, <20Hz) during a period of 6 weeks and 12 age-matched controls were studied. All the handling and care of the experimental animals was performed by authorised researchers and was done in accordance with the EU Commission on Animal Protection for Experimental and Scientific Purposes (2010/63/EU). Hearts were transversely sectioned and the atrial fragment was selected for analysis. Chromotrope-aniline blue staining was used for histological observation and the images were obtained with an optical microscope using 400× magnifications. For each atrium, three optical fields containing more prominent fibrotic development in the absence of any arterial vessel were selected. The measurement of fibrosis was performed using Image J software. Mann–Whitney test was used to compare the groups. RESULTS: The mean values of atrial interstitial fibrosis were 8.96 ± 4.08 and 4.91 ± 1.46, respectively, in IFS-exposed rats and controls. IFS-exposed rats exhibited a significant increase in atrial interstitial fibrosis (p = .005). DISCUSSION AND CONCLUSION: High-intensity IFS induces atrial interstitial fibrosis in rats. This finding reinforces the need for further experimental and clinical studies concerning the effects of IFS on the heart

Efeito da exposição prolongada a ruído ocupacional na função respiratória de trabalhadores da indústria têxtil Effects of long term exposure to occupational noise on textile industry workers' lung function

O presente estudo procurou detectar eventuais alterações da função respiratória em 28 operárias de uma fábrica de fiação, submetidas a uma exposição prolongada ao ruído (> 10 anos) com amplitude elevada e baixa frequência, susceptível de causar doença vibro-acústica (DVA). Foram realizados testes da função respiratória, incluindo espirometria, oscilometria de impulso e estudo da Capacidade de difusão alvéolo-capilar pelo CO. Os resultados foram comparados com os de outra população de igual número de mulheres, do mesmo grupo etário, consideradas normais no aspecto respiratório, com cargos administrativos, não sujeitas a agressão acústica detectável. Realizou-se um estudo estatístico dos resultados obtidos comparando as duas populações, tendo sido calculado para cada um dos parâmetros, escolhidos a partir do estudo funcional respiratório, os valores médios, o desvio-padrão e o grau de significância (pVibroacoustic disease is a pathology caused by long occupational exposure to large pressure amplitude and low frequency noise. It is a systemic disease, with evolvement of respiratory structures. The exposure workers to this noise of textile industry may favour alterations in lung function. We studied 28 women working more than ten years in cotton-mill rooms to evaluate their lung function, including Spirometry, forced oscillation technique (I.O.S.) and Diffusion capacity. These results were compared with those of 30 women of similar ages not exposed to similar noise and not presenting respiratory disease. Statistical significance (p <0.05) was found with FEV25, R5 and D Rs5-Rs20. There was a resistance frequency dependence in 36% of the population exposed to noise, not statistically confirmed. Neither restriction nor changes in diffusing capacity where detected. Conclusion: The analysis of global alterations of lung function parameters suggests small airways aggression by noise. However we cannot definitively exclude the influence of cotton dust inhalation in itself which effects could be increased by the loss of ciliated cells and impairment of airways clearance caused by noise

Immunosuppression abrogates resistance of young rabbits to Rabbit Haemorrhagic Disease (RHD)

International audienceRabbit Haemorrhagic Disease (RHD) is caused by a calicivirus (RHDV) that kills 90% of infected adult European rabbits within 3 days. Remarkably, young rabbits are resistant to RHD. We induced immunosuppression in young rabbits by treatment with methylprednisolone acetate (MPA) and challenged the animals with RHDV by intramuscular injection. All of these young rabbits died within 3 days of infection due to fulminant hepatitis, presenting a large number of RHDV-positive dead or apoptotic hepatocytes, and a significant seric increase in cytokines, features that are similar to those of naïve adult rabbits infected by RHDV. We conclude that MPA-induced immunosuppression abrogates the resistance of young rabbits to RHD, indicating that there are differences in the innate immune system between young and adult rabbits that contribute to their distinct resistance/susceptibility to RHDV infection