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J Microsc Ultrastruct. 2019 Oct-Dec;7(4):147-152. doi: 10.4103/JMAU.JMAU_1_19. Epub 2019 Nov 18.

Effects of Combined Nicotine and Caffeine on the Rat Skeletal Muscles: A Histological and Immunohistochemical Study.

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Department of Anatomy, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia.
Department of Histology and Cell Biology, Faculty of Medicine, Ain Shams University, Cairo, Egypt.



Nicotine and caffeine are pharmacologically active substances that consumed widely in the whole world. Most of the nicotine users also consume caffeine. Smokers tend to drink more coffee than nonsmokers. It is important to characterize these substances with regard to their effects on the histological and immunohistological structure.


The objective of the study is to assess the impact of combined administration of nicotine and caffeine on histological structure of the skeletal muscle tissue in the adult male Wistar rats.

Materials and Methods:

Twenty adult male Wistar rats with an average weight of 200-250 g were randomly divided into four equal groups: control, nicotine, caffeine, and combined (nicotine + caffeine). The diaphragm muscle was processed and stained with hematoxylin and eosin (H and E) stain, histochemically by periodic acid-Schiff (PAS) and immunohistochemically by anti-CD68 antibodies.


After injected nicotine, thick basement membrane with apparent increase in the positive CD68 macrophages inbetween the diaphragm muscle fibers. After injected caffeine, there was an apparent accumulation of mononuclear cells around some fibers with decrease in the PAS positive fibers. Combined injected (nicotine + caffeine) group, some fibers exhibited deep acidophilic cytoplasm with flat peripheral nuclei and apparent increase of the CD68 positive cells. There was an increase in PAS positive material around fibers appearing as a thick basement membrane.


The present study proved that caffeine and nicotine either taken alone or in combination have many negative impacts on the active type of skeletal muscles like diaphragm leading to degenerative changes that may affect their function.


CD68; Caffeine; histology; nicotine; periodic acid–Schiff; skeletal muscles

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