Osteoporosis, which is supposed to be a prevalent systemic skeletal disease worldwide, features low bone mass, microarchitectural deterioration of bone tissue and decreased bone strength, thus bringing about increased bone fragility as well as elevated fracture risk (Kanis et al., 2019, Stumpf et al., 2022). It was estimated that over 9.9 million Americans suffered from osteoporosis and about 43.1 million lived with low bone mass (Cosman et al., 2014, Wright et al., 2014). A case of a previous study testified that various risk factors were found to be the contributors to osteoporosis, such as smoking, alcohol abuse, coffee, glucocorticoid treatment as well as low body mass index (Kanis et al., 2019, Tanski et al., 2021). It is acknowledged that osteogenic differentiation acts as a critical factor in bone regeneration; therefore, the discussion of the regulatory mechanism of osteoblast differentiation as well as calcification is vital for the improvement of bone-related diseases (Fang et al., 2015).

Being cell surface receptors for the secretory ligand Slits, roundabout (Robo) family proteins were initially discovered to be repulsive guidance cues for axon pathfinding and migrating neurons in the course of nervous system advancement (Fan et al., 2012). In its extracellular domain, Robo2, which is a transmembrane protein, contains five Ig motifs as well as three fibronectin type III (FNIII) repeats (Dickson and Gilestro, 2006). It was reported that the mRNA expression of Robo2 was ascended in murine pre-osteoblastic MC3T3-E1 cells as well as in rat bone marrow stromal cells during differentiation (Sun et al., 2009). Besides, osteosclerosis protein, a negative regulator of bone formation, induced declined Robo2 expression in osteoblasts (Shi et al., 2018), implying that Robo2 can be involved in bone formation.

Being an 84-amino acid peptide hormone, parathyroid hormone (PTH), which is synthesized and secreted by the parathyroid glands, acts as a pivotal regulatory player in calcium and phosphorus metabolism as well as imparts promotive impacts on osteoblast differentiation (Arumugam et al., 2019). Additionally, PTH could alleviate osteoclast damage through autophagy induction (Zhu et al., 2017). In view of this, whether Robo2/autophagy could be induced by PTH was addressed in this paper.

To sum up, this study utilized bone morphogenetic protein 2 (BMP2) to induce pre-osteoblastic MC3T3-E1 cells so as to establish a cellular model of osteoporosis in vitro. In this paper, the role of Robo2 in osteoporosis as well as its hidden mechanism was resolved, intending to seek for possible methods to promote osteoblast differentiation and mineralization.