L -Theanine alleviates heat stress via modulating the transcriptional regulation of Hsph1 by Hsf1

Abstract Strategies altering dietary nutrient composition, such as the addition of appropriate essential amino acids or phytochemicals, are effective in addressing the adverse effects of heat stress (HS) under rising temperatures. The new food ingredient L-theanine (LTA), a natural non-protein amino acid found in tea, showed a promising effect in alleviating HS. However, systematic studies on the mechanisms by which LTA mitigates HS are scarce. Here, we evaluated the mechanisms by which LTA alleviates HS both in vivo and in vitro. The results showed that LTA suppressed HS-induced decreases in cell viability, antioxidant enzyme activity, mitochondrial membrane potential and ATP content. It reduced reactive oxygen species levels and apoptosis in heat-stressed BRL-3A cells. In addition, LTA affected the expression of heat shock proteins, ribosomal proteins and apoptosis-relate genes during HS and HS recovery (HSR) process. Similar results were observed in heat-stressed rats. Mechanistically, the alleviating effect of LTA on HS may be mediated by heat shock factor 1 (Hsf1). Specifically, at the initiation of HS, LTA promoted the binding of Hsf1 to the promoter of heat shock protein family H member 1 (Hsph1), which facilitated Hsph1 transcription and accelerated the response to HS. Following the end of HS treatment, LTA repressed the expression of Hsph1, thus restoring intracellular homeostasis. Taken together, these results suggest that LTA effectively regulates the transcription of Hsph1 via Hsf1, thereby mitigating HS-induced hepatic injury. This provides scientific evidence supporting the potential of LTA, a promising anti-HS phytochemical that deserves further development.