The yak is a unique bovine species living in the high-altitude areas of Central Asia, with significant economic, cultural, and social value [1]. Due to intense UV radiation, short growing season of pastures, and the cold, dry climate of the plateau, yaks have evolved a set of peculiar physical structures and physiological mechanisms to resist the frigid, hypoxic, and rough feeding conditions of high-altitude environments [2].

Cryptorchidism, a common disease of the male reproductive system, is one of the leading cause of male yak infertility. It induces disturbances in the seminiferous epithelium and interstitial tissue of ectopic testes [3], causes changes in testicular capillary angiogenesis or degeneration, thickening of the endovascular cortex, and ultimately impedes material exchange [4]. Currently, research on cryptorchid infertility is focused on humans and rodents, with a lack of attention to cryptorchidism in large domestic animals, especially rare large animals such as yaks and camels [5].

As the only somatic cell type in seminiferous tubules, Sertoli cells (SCs) are the most sensitive cells to temperature and oxygen content, play a vital role in supporting and nourishing the germ cells (GCs). The tight junctions (TJs) between the SCs are the key components to form the “blood-testis-barrier” (BTB). BTB is a specialized structure consisting of TJs, gap junctions, and anchor junctions (β-catenin and N-cadherin). These junctions connect SCs to SCs. The TJs include occludins and zonula occludens (ZO-1) [6]. The function of the BTB is not only to avoid the adverse effects of anti-sperm antibodies and autoimmune reactions but also to prevent harmful substances from entering the seminiferous tubules, which provide a unique and stable environment for the development of GCs [7]. Cryptorchidism is accompanied by tissue fibrosis and calcification [8], and changes in the spermatogenic environment induce autophagy and apoptosis in GCs [9,10].

Autophagy is a highly regulated and evolutionarily conserved cellular process that maintains the stability of the intracellular environment by eliminating damaged organelles and misfolded proteins [[11], [12], [13]]. Rapamycin (mTOR) mammalian target is a vital autophagy regulator [14]. Cumulative research has revealed that autophagy is involved in endocrinological processes, acrosome biogenesis, and spermatogenesis [15]. Conditional knockout of autophagy-related genes in mice severely affects fertility, resulting in the production of malformed and/or absent acrosomes, large cytoplasmic droplets, and reduced motility [16,17], similar to those observed in cryptorchidism.

Heme oxygenase 1 (HO1), a significant rate-limiting enzyme, has emerged as a central effector of the mammalian stress response. It is also known as heat shock protein 32 (Hsp32) and is associated with anti-apoptotic functions and cellular antioxidant defense [18,19]. Furthermore, HO1 has been shown to resist or alleviate various types of cell death stress, such as anti-apoptosis, anti-necrosis, anti-necroptosis, anti-pyroptosis, and anti-ferrocytosis [20]. Unfortunately, few studies have investigated the relationship between HO1 and the male reproductive system. Some studies have shown that in SCs cultured in vitro in mice, heat stress increased HO1 expression, which alleviated apoptosis of SCs [21].

In this study, we selected SCs as in vitro models to simulate the occurrence of cryptorchidism and its effect on reproductive processes in hypoxia. The regulatory function of HO1 in autophagy and apoptosis of yak SCs under hypoxic conditions was then demonstrated by knocking out and over-expression of HO1. This study aimed to provide a theoretical basis for improving male fertility and treating male infertility in plateau organisms.