Lawrence Levine

Differentiated Rat Glial Cell Strain in Tissue Culture

Rat glial tumors, induced by injections of N-nitrosomethylurea, were plated and propagated in culture. Among a few cell strains obtained, one clone contains S-100 protein, which is unique to brain in vertebrates. Stationary-phase cultures contain approximately ten times more S-100 protein per cell than exponentially growing cells. When injected into newborn rats, cells producing S-100 grew as a glial tumor, which contained S-100 protein.

Antibodies to Bradykinin and Angiotensin: A Use of Carbodiimides in Immunology

Antibodies to bradykinin and angiotensin have been produced in rabbits by the use of conjugates containing albumin and the hapten, covalently bound. The use of water-soluble carbodiimide reagents provided an easy and rapid method of synthesis of the antigenic conjugates.

Accumulation of Cyclooxygenase Products of Arachidonic Acid Metabolism in Gerbil Brain During Reperfusion After Bilateral Common Carotid Artery Occlusion

: Several of the cyclooxygenase products of arachidonic acid were measured in the cerebral hemispheres of gerbils subjected to transient interruption of the cerebral circulation. The levels of PGD2, PGF2α, PGE2, TXB2, 13,14‐H2‐15‐keto‐PGE2, and the stable nonenzymic product of prostacyclin, 6‐keto‐PGF1α, were not altered at the end of a 5‐min period of ischemia. However, the onset of reperfusion was accompanied by a rapid accumulation of these products. Levels were highest during the initial period of reperfusion, then decreased to approach control levels after 120 min. PGD2, PGF2α, and PGE2 were the predominant metabolites detected. This postischemic accumulation of arachidonic acid metabolites could be blocked by prior administration of inhibitors of cyclooxygenase activity.

Captopril-induced Changes in Prostaglandin Production: RELATIONSHIP TO VASCULAR RESPONSES IN NORMAL MAN

Captopril is a potent hypotensive agent whose efficacy has hitherto been attributed to its ability to alter either angiotensin II formation or kinin degradation. Our purpose was to examine captoprils acute effect on prostaglandin production, because changes in neither the renin-angiotensin nor the kallikrein-kinin systems appear adequate to account for the fall in arterial pressure. The plasma levels of angiotensin II, kinins, and prostaglandins were determined in response to increasing doses (5, 12.5, and 25 mg) of captopril and these responses were compared with the change in arterial pressure observed in nine supine normal male subjects studied on both a high (200 meq) and low (10 meq) sodium intake.Captopril significantly (P < 0.01) increased the levels of the 13,14-dihydro-15-keto metabolite of prostaglandin E(2) (PGE(2)-M), a potent vasodilator, with similar responses being observed on both a high and a low sodium intake. No significant changes in the plasma levels of 6-keto-prostaglandin F (1)alpha, or thromboxane B(2), the stable products of prostacyclin and thromboxane A(2), respectively, occurred. The depressor response to captopril correlated with the change in PGE(2)-M (r = 0.52, t = 5.44, P < 0.0001). On the other hand, although significant (P < 0.02) decrements in angiotensin II and increments in plasma kinins accompanied the hypotensive response in sodium-restricted subjects, in sodium-loaded subjects where the renin-angiotensin system was suppressed, no change in angiotensin II, and only a modest change in kinins was noted, even though significant (P < 0.01) decrements in diastolic blood pressure occurred (-10+/-2 mm Hg).Thus, changes in depressor prostaglandin production can better account for the hypotensive response to captopril, thereby extending to yet another vasoactive system an influence by this class of drugs and providing a new approach to dissecting the abnormality in the control of vascular tone in patients with hypertension.

Prostaglandin-stimulated bone resorption by rheumatoid synovia. A possible mechanism for bone destruction in rheumatoid arthritis.

Synovial tissue from patients with rheumatoid arthritis was maintained in organ culture for 3-14 days. Conditioned media from these synovial cultures contained bone resorption-stimulating activity, measured in vitro by using calcium release from mouse calvaria as the assay system. The synovial cultures also produce prostaglandin E2 (PGE2) as measured by serologic methods. The production of both the bone resorption-stimulating activity and PGE2 was inhibited by more than 90% by treatment of the synovial cultures with indomethacin (5 mug/ml). In contrast, treatment of the synovial cultures with colchicine (0.1 mug/ml) caused a marked and parallel increase in the concentration of both bone resorption-stimulating activity and PGE2 in the conditioned media. The bone resorption-stimulating activity was quantitatively extracted into diethyl ether. Within the limits of experimental error, all of the bone resorption-stimulating activity in medium was accounted for by its content of PGE2, itself a potent osteolytic factor. We conclude that the bone resorption-stimulating activity produced by rheumatoid synovia in culture is PGE2.

Epidermal growth factor stimulates prostaglandin production and bone resorption in cultured mouse calvaria.

Murine epidermal growth factor (EGF) stimulated the production of prostaglandin E2 (PGE2) and bone resorption in neonatal mouse calvaria in organ culture. The effect of EGF on bone resorption occurred at low concentrations of the polypeptide (half-max stimulation = 0.4 ng/ml, 6.6 × 10−11 M). All concentrations of EGF which stimulated resorption also stimulated the production of PGE2 by bone; concentrations of EGF which did not stimulate resorption did not enhance PGE2 production. EGF-induced formation of PGE2 and bone resorption were inhibited completely by indomethacin (200 ng/ml) and hydrocortisone (3 × 10−6 M). Indomethacin did not inhibit the bone resorption-stimulating activity of exogenous PGE2. The time courses of action of EGF, parathyroid hormone and exogenous PGE2 on bone resorption were similar. Brief exposure (15 or 60 min) to EGF (10 ng/ml) did not cause bone resorption or an increase in PGE2 accumulation in a subsequent 48-h incubation in the absence of EGF. High concentrations (30 to 100 ng/ml) of bovine fibroblast growth factor (FGF) also stimulated the production of PGE2 and bone resorption. We conclude that concentrations of EGF equal to or less than those present in mouse plasma stimulate the resorption of mouse bone in organ culture by a mechanism that involves the enhanced local production of PGE2.

Transient cerebral ischemia and brain prostaglandins.

Abstract Prostaglandin levels were measured in gerbil brain during cerebral ischemia for up to 2 hours, and during reperfusion after various periods of transient ischemia. The levels of PGF2α and its metabolite (13,14-dihydro-15-keto-PGF2α) were low and did not change during ischemia. However, PGF2α, 13,14-dihydro-15-keto-PGF2α, PGE2, and thromboxane B2 increased during reperfusion after brief episodes of ischemia. Indomethacin or aspirin inhibited this increase. Animals pretreated with indomethacin recovered more rapidly and were more active during reperfusion than those without treatment.

Contribution of prostaglandins to the antihypertensive action of captopril in essential hypertension.

SUMMARY To determine whether prostaglandins contribute to the depressor response to the converting enzyme inhibitor, captopril, we measured the plasma prostaglandln levels by radioimmunoassay before and after captopril administration, and then examined the effect of prostaglandin synthetase inhibition on captoprils antihypertensive effect. When a single oral captopril dose (25-100 mg) was given to 31 sodium-restricted patients with essential hypertension, the levels of the stable transformation product of prostacyclin remained immeasurable and that of thromboxane A, did not change, while the metabolite of PGE, (PGE-M) increased by 53% (34 ± 4 pg/ml pre-captopril, 52 ± 5 pg/ral after; p < 0.001). As expected, blood pressure (BP) and angiotensin II (AH) levels fell, and kinin levels rose (all changesp < 0.001). We then blocked prostaglandin synthesis in 18 of these subjects for 24 hours with either indomethacin (n = 10) or aspirin (n = 8) before repeating the captopril dose, to assess the importance of these PGE-M increments. The PGE-M responses to captopril were effectively blocked in nine of 10 subjects receiving indomethacin and four of eight receiving aspirin. In these 13 patients, the depressor response to captopril was significantly blunted (− 20 ± 3 mm Hg pre-synthetase inhibition vs − 13 ± 2 mm Hg post; p < 0.05). When these agents did not block the PGE-M response to captopril, the BP response was also unchanged (-15 ± 4 mm Hg pre, −18 ± 5 mm Hg post). Neither indomethacin nor aspirin changed the AH or kinin responses to captopril. We conclude that the prostaglandins may be important mediators of captoprils antihypertensive effect in the sodium-restricted state.

APPEARANCE OF A BRAIN SPECIFIC ANTIGEN (THE S‐100 PROTEIN) DURING HUMAN FOETAL DEVELOPMENT

—The concentration of a protein specific to brain, the S‐100 protein, was measured in various regions of the human foetal brain at gestational ages ranging from 10 weeks until term. The relative increase in concentrations of the S‐100 protein during development of the human foetal brain proceeded in a caudal‐rostral fashion. This observation is emphasized by the delayed appearance of the S‐100 protein in the frontal cerebral cortex until the 30th week of gestation.

Hypercalcemia and tumor-prostaglandins: The VX2 carcinoma model in the rabbit☆☆☆

The VX2 carcinoma produces profound hypercalcemia (17-22 mg/100 ml) in the rabbit about 3-4 wk after transplantation. A bone resorption-stimulation factor (assayed in vitro with mouse calvaria in culture) has been extracted with diethyl ether from the tumor tissue and from the medium of a clonal strain of VX2 cells grown in culture. Serologic methods reveal that the tumors contain 294 plus or minus 51 ng/g fresh weight (mean plus or minus SE, 25 tumors) of prostaglandin E2 (PGE2), a potent bone resorption-stimulating agent. VX2 cells in culture produce 0.5-3.0 mug PGE2 per mg cell protein per 24 hr. The production of bone resorption-stimulating activity and PGE2 by VX2 cells in culture were both inhibited by indomethacin (100 ng/ml). Tumors from normocalcemic, indomethacin-treated rabbits (10-40 mg/rabbit/24 hr) contained little or no bone resorption-stimulating activity nor PGE2. Tumor-bearing rabbits receiving indomethacin continuously did not develop hypercalcemia, however, following cessation of indomethacin administration, hypercalcemia developed rapidly and was again reversed by reinstitution of indomethacin feeding. In untreated, hypercalcemic, tumor-bearing rabbits, initiation of indomethacin treatment was followed by a rapid return of the plasma calcium to the normal range. Systemic venous plasma from hypercalcemic tumor-bearing plasma contained higher concentrations of PGE2 than plasma from normocalcemic control rabbits. Venous drainage of the tumor contained even higher plasma PGE2 concentrations than systemic venous plasma in hypercalcemic animals; plasma PGE2 concentrations locally and in systemic plasma were unmeasurable (less than 70 pg/ml) in normocalcemic, indomethacin-treated, tumor-bearing rabbits. We conclude that PGE2 is a bone resorption-stimulating factor produced by VX2 tumor cells, and that secretion of PGE2 by the tumor in vivo may well be responsible for the hypercalcemia observed in tumor-bearing rabbits.