title: Long-term response of cultured rat parathyroid glands to calcium and calcitriol: the effect of cryopreservation.
abstract: OBJECTIVE
  This study aimed to investigate the longest period in which parathyroid glands cultured in vitro maintained their viability and functionality in order to study the response of the glands to factors that exert their main action in the long term (1-7 days).
  
  
  METHODS
  Rat parathyroid glands from 104 Wistar rats were used. Cell viability was measured by flow cytometry for 7 days. Parathyroid tissue functionality was determined by parathyroid hormone (PTH) secretion in basal conditions and in the response of the glands to calcium and calcitriol. Calcium sensing receptor (CaR) synthesis was determined measuring protein levels by immunohistochemistry. Parathyroid glands were cryopreserved to study them in the same way as fresh tissue.
  
  
  RESULTS
  Intact parathyroid glands maintained their cell viability >80% until the 6th day in culture, while the functional capacity was limited to 4 days: PTH release was stable for 4 days, whilst from the 5th day onwards, PTH secretion reduced to undetectable levels. Parathyroid glands responded accurately when calcium was reduced in the culture medium; a mean increase >50% in PTH secretion was observed. No differences were observed in CaR levels before and after the culture period. PTH synthesis and secretion inhibition was observed when the parathyroid glands were cultured with calcitriol; this inhibition achieved 90% after 4 days in culture. Cryopreserved parathyroid glands maintained their viability, but partially lost their functionality, as they were unable to respond to calcium.
  
  
  CONCLUSIONS
  Intact parathyroid glands cultured in vitro maintained their functionality and their capacity to respond to their effectors for longer periods than in previously developed studies. It seems that part of this capacity is lost after cryopreservation. Nevertheless, this long-term culture model can be useful to study the response of the parathyroid glands.
authors.affiliation: Bone and Mineral Research Unit, Hospital Universitario Central de Asturias, Instituto Reina Sofia de Investigacion, Oviedo, Spain.
authors.forename: Daniel
authors.initials: D
authors.lastname: Alvarez-Hernandez
authors.forename: Ignacio
authors.initials: I
authors.lastname: Gonzalez-Suarez
authors.forename: Manuel
authors.initials: M
authors.lastname: Naves
authors.forename: Natalia
authors.initials: N
authors.lastname: Carrillo-Lopez
authors.forename: Teresa
authors.initials: T
authors.lastname: Fdez-Coto
authors.forename: Jose L
authors.initials: JL
authors.lastname: Fernandez-Martin
authors.forename: Jorge B
authors.initials: JB
authors.lastname: Cannata-Andia
mesh_terms: D000818:Animals; D002117:Calcitriol / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology*; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D015925:Cryopreservation; D008297:Male; D010280:Parathyroid Glands / Q000187:drug effects* / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors; D046509:Tissue Culture Techniques / Q000379:methods*; D014022:Tissue Survival / Q000187:drug effects*
chemical_list: D002120:Calcium Channel Agonists; D002117:Calcitriol; D002118:Calcium