Monoclonal antibody production to human and bovine 2′:3′-cyclic nucleotide 3′-phosphodiesterase (CNPase): high-specificity recognition in whole brain acetone powders and conservation of sequence between CNP1 and CNP2

Monoclonal antibodies against human and bovine 2′:3′-cyclic nucleotide 3′-phosphodiesterase (CNPase) were generated by fusing FOX-NY myeloma cells with spleen cells from RBF/Dn mice previously immunized with the purified brain antigens. The enzyme isolated from bovine brain was quite basic, with an isoelectric point of 9.71 and both the bovine and human enzymes consisted of a closely spaced doublet at approximately 44 and 46 kDa on SDS-PAGE. Six monoclonals were identified as strongly recognizing the enzyme on both ELISA plates and on immunoblots of whole brain protein. Four monoclonals very weakly cross-reacted with guinea pig myelin basic protein. In contrast with two previous reports, some of our monoclonal antibodies did immunostain 2 or 3 protein bands in peripheral nerve, two bands closely corresponding to those immunostained in central nervous system (CNS) myelin, the Wolfgram protein fraction and in acetone powders of whole brain. Each of the 6 monoclonals reacting strongly on immunoblots recognized the enzyme in from 2 to 5 of the species examined (human, bovine, rat, mouse and rabbit). In addition, all 6 monoclonals that immunostained the enzyme in whole brain, myelin and Wolfgram protein immunoblots recognized both CNP1 (44 kDa) and CNP2 (46 kDa). The two closely spaced protein bands observed on SDS-PAGE and previously stained on immunoblots of CNS CNPase using polyvalent rabbit anti-bovine CNPase antisera, and now different monoclonal antibodies, appear to be immunologically related and to contain highly conserved sequences.     

BIOCHEMICAL CORRECTION IN THE SYNDROME OF HYPERTENSION AND HYPERKALAEMIA BY SEVERE DIETARY SALT RESTRICTION SUGGESTS RENIN-ALDOSTERONE SUPPRESSION CRITICAL IN PATHOPHYSIOLOGY

1. Plasma potassium and chloride concentrations were raised and plasma renin activity, aldosterone, bicarbonate and arterial pH were reduced in two brothers with the syndrome of hypertension and hyperkalaemia with normal glomerular filtration rate (Gordon's syndrome), on unrestricted or moderately restricted sodium diets. 2. These abnormalities were corrected in both patients within 10 days of severe sodium restriction. 3. Pressor sensitivity to cold and angiotensin II decreased on low sodium diet, associated with a fall in blood pressure. 4. Increasing distal tubular sodium delivery by infusion of normal saline increased fractional excretion of potassium when aldosterone had been stimulated by severely restricted sodium diet, but not when aldosterone levels were low on unrestricted sodium diet. 5. These findings are consistent with excessive sodium reabsorption as the primary renal lesion in Gordon's syndrome, leading to volume expansion and suppression of renin and aldosterone. Severe dietary sodium restriction leading to volume contraction, by stimulating renin and aldosterone and promoting kaliuresis, corrects the abnormalities.