Anabolic steroids are chemically synthetic derivatives of the male sex hormone testosterone. They are used in medicine for their ability to support muscle growth and healing and by athletes for esthetic purposes and to increase sports performance, but another major use is in fattening animals to increase meat production. The more people there are on Earth, the greater the need for meat production and anabolic steroids accelerate the growth of animals and, most importantly, increase the amount of muscle mass. Anabolic steroids also have proven side effects that affect all organs and tissues, such as liver and kidney parenchymal damage, heart muscle degeneration, organ growth, coagulation disorders, and increased risk of muscle and tendon rupture. Anabolic steroids also have a number of harmful effects on the developing brain, such as brain atrophy and changes in gene expression with consequent changes in the neural circuits involved in cognitive functions. Behavioral changes such as aggression, irritability, anxiety and depression are related to changes in the brain. In terms of long-term toxicity, the greatest impact is on the reproductive system, i.e., testicular shrinkage and infertility. Therefore, their abuse can be considered a public health problem. In many countries around the world, such as the United States, Canada, China, Argentina, Australia, and other large meat producers, the use of steroids is permitted but in all countries of the European Union there is a strict ban on the use of anabolic steroids in fattening animals. Meat from a lot of countries must be carefully inspected and monitored for steroids before export to Europe. Gas or liquid chromatography methods in combination with mass spectrometry detectors and immunochemical methods are most often used for the analysis of these substances. These methods have been considered the most modern for decades, but can be completely ineffective if they face new synthetic steroid derivatives and want to meet meat safety requirements. The problem of last years is the application of “cocktails” of anabolic substances with very low concentrations, which are difficult to detect and are difficult to quantify using conventional detection methods. This is the reason why scientists are trying to find new methods of detection, mainly based on changes in the structure of tissues and cells and their metabolism. This review gathered this knowledge into a coherent form and its findings could help in finding such a combination of changes in tissues that would form a typical picture for evidence of anabolic misuse.

Anabolic steroid hormones (AASs) are used in most countries of the world to accelerate the growth of animals, increase the volume of their muscles and thereby increase meat production. However, there is a strict ban on the use of AASs in the fattening of all animals in all countries of the European Union, and there must therefore be effective methods of detection and control of these substances. Methods based on chromatography and mass spectrometry may no longer be completely effective when faced with new synthetic steroids of unknown chemical structures and low concentrations. Therefore, there is an effort to develop new methods of AAS detection, based primarily on the monitoring of biological changes at the level of gene expression or changes in metabolism or structure at the cellular level. More detailed knowledge of the mechanisms of action of AASs on tissues is essential for these methods, and histological changes are one of them. In this study, we report histological changes in muscle structure after AAS application, specifically in the size of muscle fibers, the amount of endomysium and the number of nuclei and satellite cells in muscle fibers. A pig model was also intentionally used for the study, as no such study has been carried out on this species, and at the same time, pork is one of the most consumed meats across Europe. The results of histology and fluorescent antibody labeling showed that AASs increased the diameter and surface area of muscle fibers and also significantly increased the number of satellite cells on the fiber surface. The evident correlations between the number of satellite cells, all nuclei and the diameters of muscle fibers between some experimental groups provide evidence that the selected histological parameters could be additional detection mechanisms for screening a large number of samples and indicate the possibility of the presence of AASs in pork meat in the future.