In Vivo Engraftment and Functional Efficacy of a 3D-Bioprinted Human Parathyroid Equivalent

<i>Background and Objectives</i>: Hypocalcemia due to hypoparathyroidism (HypoPTH) is the most common complication following thyroid surgery, typically resulting from iatrogenic removal, tissue damage, or compromised vascularization of the parathyroid glands. Patients with persistent HypoPTH are at risk for long-term complications such as osteoporosis, cardiac dysfunction, and renal impairment. Lifelong regulation of calcium levels is therefore essential to prevent morbidity and mortality associated with these complications. In this study, we aimed to evaluate the functional engraftment efficacy of 3D bioprinted human parathyroid tissue constructs in a xenograft model in vivo. <i>Materials and Methods</i>: Primary cells obtained from freshly excised human parathyroid tissue specimens were isolated and 3D bioprinted using alginate-based bioink. The bioprinted tissue constructs were implanted into CD1 athymic mice. Histopathological evaluation of the grafted constructs was performed at different time points. In addition, surface calcium-sensing receptor (CaSR) expression was assessed by immunofluorescence as an indicator of functional parathyroid tissue engraftment. <i>Results</i>: The presence of CaSR on parathyroid cells within the 3D-printed scaffolds confirmed the persistence of functional parathyroid cells following implantation. In tissue samples obtained during the first, second, and third weeks after implantation, CaSR positivity was consistently observed in the parathyroid cells. However, at the three-month follow-up, the pores within the scaffolds were found to be filled with calcified material and replaced by fibrotic tissue. At this stage, the absence of parathyroid hormone (PTH) expression indicated a loss of functional activity in the grafted biomaterial. <i>Conclusions</i>: Human primary parathyroid cells were successfully isolated, and a functional, hormone-active parathyroid tissue substitute was developed ex vivo using 3D-bioprinted hydrogel scaffolds combined with autologous cells. Although short-term functional engraftment was achieved, long-term graft viability and hormonal activity were limited due to scaffold degradation and fibrosis. These findings indicate the necessity for further improvement in scaffold biocompatibility to enhance the therapeutic potential of 3D-bioprinted parathyroid tissue constructs for in vivo applications.