Production of calf anterior pituitary progenitor cell line for conditional regulation of gonadotroph differentiation

Pituitary stem/progenitor cells are essential for regulating the physiological functions mediated by anterior pituitary hormones. Studying the molecular mechanisms governing their differentiation of these cells is crucial for improving the cattle productivity, an agriculturally and industrially important animal. However, elucidating such mechanisms remain a challenge, as maintaining pituitary stem/progenitor cells in vitro is difficult. Here, we attempted to establish immortalized cells for continuous utilization of cattle pituitary stem/progenitor cells. The pituitary stem/progenitor cell marker Sox2 was constitutively expressed in the cattle pituitary gland regardless of age. Calf anterior pituitary cells expressing multiple stem/progenitor cell markers (Sox2, Pitx1, Prrx1, and/or Prop1) were immortalized by introducing temperature-sensitive simian virus 40 large T antigen. Hormonal gene expression was not detected in any cell line. Among these SOX2-positive calf anterior pituitary cell lines, a pituitary-specific marker PROP1-positive cell proliferated when the immortalization was switched on, and differentiated into gonadotroph positive for luteinizing hormone when it was off, a finding supported by the increased expression of transcription factors (Pitx1 and Egr1) that upregulate Lhb expression. However, in this cell line, no changes were observed in the differentiation lineage even when a Notch signaling inhibitor was used. Thus, we established a unique calf anterior pituitary progenitor cell line to study the mechanisms of gonadotroph differentiation.