Binding of Monoclonal Antibody (4F2) to its Cell Surface Antigen on Dispersed Adenomatous Parathyroid Cells Raises Cytosolic Calcium and Inhibits Parathyroid Hormone Secretion*

Abstract

In the course of characterizing monoclonal antibodies (MAbs) recognizing cell surface antigens on dispersed human parathyroid cells (dPTCs), we identified one MAb (4F2) that bound avidly to parathyroid cells and had marked effects on parathyroid function. The binding of MAb 4F2 to human adenomatous dPTCs resulted in a marked [53.8 +/- 7.9% (+/- SEM)] reduction in low calcium (Ca)-stimulated PTH secretion to levels equivalent to those in cell suppressed by high extracellular Ca (1.5 mM). Typically, these functional effects were optimal at antibody dilutions of 1:10(4) to 1:10(5). Cell viability was confirmed at the conclusion of each experiment by trypan blue exclusion (greater than 90-95%) and cell surface immunofluorescence. Parallel studies using the Ca-sensitive dye Quin-2 showed that inhibition of PTH secretion in 4F2-treated cells was associated with a concomitant increase in cytosolic Ca (Cai) of 188% in 0.5 mM Ca; these values also approached Cai levels in control cells incubated in high Ca. Mab controls, P3 X 63, which do not bind to dPTCs, and Mab LC7-2, which recognizes a different epitope of the same antigen as 4F2 on dPTCs, did not alter PTH secretion or Cai. Immunoprecipitation of 125I-labeled parathyroid cell extracts with MAb 4F2 demonstrated proteins with mol wt of approximately 145, 85, and 45 under nonreducing conditions and 85 and 45 kilodaltons after reduction with 5% mercaptoethanol. These studies suggest that 1) Mab-4F2 binding to its cell surface antigen inhibits PTH secretion by human adenomatous parathyroid cells in vitro; 2) the alterations in secretory function could be related to by an attendant increase in Cai; 3) the 4F2 antigen on dPTCs is a heterodimeric protein of (approximately) 85K and 45K; and 4) the 4F2 antigen may be an important component of the Ca-sensing and/or signal-transducing mechanism in this cell.

Enhanced tumorigenicity caused by truncation of the extracellular domain of GP125/CD98 heavy chain

“…The involvement of CD98 in cellular Na + -dependent calcium uptake has been reported in cardiac and skeletal muscle (Michalak et al, 1986), and in the parathyroid gland (Posillico et al, 1987). In this context, CD98 has been identi®ed as a major surface receptor for a mammalian galactoside-binding protein, Galectin-3 (Dong and Hughes, 1996), which binds to CD98 and induces an increase in intracellular calcium in human Jurkat T leukemia cells.…”

High extracellular Ca2+ and Mg2+ stimulate accumulation of inositol phosphates in bovine parathyroid cells

“…That is, Ca 2+ produces an initial 'spike' in cytosolic Ca 2÷, which arises from intracellular Ca 2÷ stores, followed by a sustained increase in cytosolic Ca 2÷ which derives from uptake of extracellular Ca 2÷ [4,6]. Finally a variety of factors binding to the parathyroid cell surface, such as lectins [11], naturally occurring autoantibodies [12], and monoclonal antibodies [13] modulate PTH release [11][12][13] and the cytosolic calcium concentration [13], consistent with a role for cellsurface moeities in regulating parathyroid function.…”

β1 integrins show specific association with CD98 protein in low density membranes

“…The CD98 molecule is also widely expressed on rapidly growing non-hematopoietic cells, where it may modulate oncogenic transformation [10,11], metal ion transport [12,13], cell fusion [14,15], and amino acid transport [16][17][18][19]. CD98 is also expressed on normal proliferating tissue such as the basal layer of squamous epithelia [20] and on cells having secretion or transport functions [21].…”