In recent years, as gene therapy technology has rapidly developed, there has been growing concern that it could be misused by athletes as a means of doping. However, current testing methods for gene doping have a range of limitations and require further improvement. Furthermore, significant progress has been made in the fields of blood storage, next-generation sequencing (NGS), and LabDroid (experimental robots). Against this background, this study was implemented to develop a test method for gene doping using dried blood spot (DBS), NGS, and the LabDroid ”Maholo”. As a first step, recombinant adeno-associated virus containing the human erythropoietin gene (hEPO) was injected into mice to establish a gene doping model. Subsequently, DBS was created using whole blood. Maholo was used to extract DNA from the DBS and to create DNA libraries for NGS. NGS in combination with bioinformatic analysis clearly identified DNA fragments that provided definitive evidence of gene doping in the mouse model, which were absent in the control mouse. To the best of our knowledge, this is the first attempt to use a biological model of hEPO gene doping in conjunction with Maholo, NGS, and DBS. This method should facilitate the further development of gene doping tests.

In recent years, bath additives developed to enhance the physiological benefits of bathing have become a popular part of self-care routines. However, there has been little investigation of the biological effects of these additives, especially in terms of their impact on gene expression. To bridge this research gap, this study was implemented to reveal the effects of a newly developed complex bath additive, Karada Totonou ProBath (KTPB), on gene expression, using RNA sequencing (RNA-seq). Two human skin-derived cell lines (keratinocytes and fibroblasts) were treated with KTPB at a final concentration of 100 μg/mL, and RNA extracted between 15 and 240 min after treatment was subjected to RNA-seq analysis. The results showed that KTPB induced significant changes in the expression of over 80 genes in both cell lines. In particular, the expression levels of genes encoding Early Growth Response Protein 1 and Hyaluronic Acid Synthases increased over time following the treatment. Enrichment analysis showed the upregulation of pathways associated with cell activation in both cell types, with no evidence of KTPB-related cytotoxicity. The findings suggest that KTPB activates the metabolism of skin cells through hormesis without inducing toxicity. Our results provide new insights promoting future research in this field.