Isoforms of alkaline phosphatase from mouse internal organs after bilateral adrenalectomy

Abstract

The activity of alkaline phosphatase of female CBA and BALB/c mice is studied after bilateral adrenalectomy. Interstrain differences in enzyme activity are revealed in some organs of the control and experimental animals. The expression of new isoforms of alkaline phosphatase in hypocorticoidism is demonstrated.Key Words: alkaline phosphatase; isoforms; regulation; adrenalectomy Alkaline phosphomonoesterases (EC 3.1.3.1) have been extensively studied in experimental and clinical medicine, since these enzymes play an important role in adaptive and pathological responses [7,10]. The fact that alkaline phosphatase (AP) activity is high in actively absorbing or secreting tissues indicates how important this enzyme is for the development and function of living systems. By dephosphorylating NADP and NADPH, alkaline phosphatase affects numerous oxidation-reduction processes in the cell [3]; dephosphorylation of histones and glycogen synthetase testifies to the important role of AP in the regulation of glucose metabolism and the expression of the mammalian genome [2,12].According to data in the literature data [6,9], adrenal hormones are involved in the expression of AP. In view of the considerable variations of steroid hormone levels during ontogenesis and the role of these hormones in adaptive, stress, and pathological mechanisms, it is important to study the activity of various isoforms of alkaline phosphomonoesterase under conditions of decreased concentrations of corticosteroids in the body after experimental adrenalectomy. MATERIALS AND METHODSExperiments were performed on pubertal female CBA and BALB/c mice weighing 18-20 g. A deficiency of glucocorticoid hormones was achieved by bilateral adrenalectomy performed under light ether anesthesia [4]. The "cleanness" of adrenalectomy was checked visually during the surgery and by measuring the plasma concentration of 11-hydroxycorticosteroids [5]. Animals with sham adrenalectomy (without removal of the adrenals) were taken for comparison. The activity and spectrum of AP from various organs and tissues were studied seven days after the surgery, when the corticosteroid levels were minimal. Homogenates of the liver, kidneys, lungs, and bone (caudal vertebrae) were prepared in 0.05 M Tris-HCI buffer (pH 7.2) containing Triton X-100. The AP activity was assayed in the supernatants obtained by centrifugation of the homogenates at 6000 rpm for 20 min, the assay [13] being based on measurements of the intensity of fluorescence of c~-naphthol formed after the hydrolysis of c~-naphthylphosphate at pH 9.6. The AP activity was expressed in #g o~-naphthol per mg wet weight of tissue during 15 min.Isoforms of AP were studied by vertical slab electrophoresis at room temperature in 2251~x130x2 mm gel as described elsewhere [8].