Development of a technique for introduction of an expressed complementary deoxyribonucleic acid into parathyroid cells by direct injection

Kazuhiro Shiizaki, Ikuji Hatamura, Masafumi Fukagawa, Eiko Nakazawa, Fumie Saji, Yuko Watanabe, Tadao Akizawa, Eiji Kusano

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

PTH is a major mediator of bone and mineral metabolism. However, physiological and pathological investigations of parathyroid cells (PTCs) have been limited because of the lack of available cell lines and because the organ is too small for detailed studies. Here, we describe a novel method for adenovirus-mediated cDNA transfer into PTCs, and we show the accuracy of the method in a rat model of uremia-induced secondary hyperparathyroidism. Rats underwent a 5/6-nephrectomy and were fed with a high-phosphate diet for 8 wk. The parathyroid glands were surgically exposed and adenoviruses containing LacZ or Ca-sensing receptor (CaSR) were directly injected into the glands under a zoom-stereo microscope. The parathyroid glands were analyzed for infection of adeno-virus and immunohistochemically for expression of CaSR. The functional activity of exogenous CaSR in PTCs after this treatment was investigated based on changes of the calcium and PTH curve. A virus concentration of more than 109 plaque-forming units/ml was required for adequate infection of PTCs within 7 d after treatment. Marked increase of CaSR-positive PTCs by 2.39±0.72 times relative to control treatment, and significant colocalization of CaSR overexpression and virus labeling, were observed in glands after gene introduction. The calcium and PTH curve was shifted to the left from the basal position (set point, 1.10 ± 0.09 to 0.76 ± 0.12 mM; P < 0.0001), indicating successful introduction of a functionally active cDNA into the PTCs. This technique may facilitate an elucidation of biological effects through targeting and identification of specific features of PTCs, which may provide the basis for new clinical approaches.

Original languageEnglish (US)
Pages (from-to)4031-4038
Number of pages8
JournalEndocrinology
Volume151
Issue number8
DOIs
StatePublished - Aug 2010

Fingerprint

Calcium-Sensing Receptors
Injections
DNA
Parathyroid Glands
Adenoviridae
Complementary DNA
Viruses
Calcium
Secondary Hyperparathyroidism
Uremia
Virus Diseases
Nephrectomy
Minerals
Therapeutics
Phosphates
Diet
Bone and Bones
Cell Line
Infection
Genes

ASJC Scopus subject areas

  • Endocrinology

Cite this

Shiizaki, K., Hatamura, I., Fukagawa, M., Nakazawa, E., Saji, F., Watanabe, Y., ... Kusano, E. (2010). Development of a technique for introduction of an expressed complementary deoxyribonucleic acid into parathyroid cells by direct injection. Endocrinology, 151(8), 4031-4038. https://doi.org/10.1210/en.2010-0012

Development of a technique for introduction of an expressed complementary deoxyribonucleic acid into parathyroid cells by direct injection. / Shiizaki, Kazuhiro; Hatamura, Ikuji; Fukagawa, Masafumi; Nakazawa, Eiko; Saji, Fumie; Watanabe, Yuko; Akizawa, Tadao; Kusano, Eiji.

In: Endocrinology, Vol. 151, No. 8, 08.2010, p. 4031-4038.

Research output: Contribution to journalArticle

Shiizaki, K, Hatamura, I, Fukagawa, M, Nakazawa, E, Saji, F, Watanabe, Y, Akizawa, T & Kusano, E 2010, 'Development of a technique for introduction of an expressed complementary deoxyribonucleic acid into parathyroid cells by direct injection', Endocrinology, vol. 151, no. 8, pp. 4031-4038. https://doi.org/10.1210/en.2010-0012
Shiizaki, Kazuhiro ; Hatamura, Ikuji ; Fukagawa, Masafumi ; Nakazawa, Eiko ; Saji, Fumie ; Watanabe, Yuko ; Akizawa, Tadao ; Kusano, Eiji. / Development of a technique for introduction of an expressed complementary deoxyribonucleic acid into parathyroid cells by direct injection. In: Endocrinology. 2010 ; Vol. 151, No. 8. pp. 4031-4038.
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