Thesis (Ph. D.)--University of Wisconsin--Madison, 1982. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

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The cellular origin of prostaglandins in human tumors was investigated using cell fractionation procedures and high resolution gas chromatography. Additionally, the role of macrophages and prostaglandins on human tumor cloning in vitro was investigated. Spontaneous human tumors were prepared as single cell suspensions which were subsequently manipulated to yield macrophage-enriched, and tumor cell enriched (macrophage-depleted) subpopulations of cells. A fused silica capillary gas chromatographic analysis with electron capture detection was developed to measure derivatized prostaglandins in the supernatant of the cell subpopulation incubations. The derivative used for the analysis was the pentafluorobenzyl ester-methoxime-trimethyl-silyl ether. The assay showed a detection limit of 25 picograms of prostaglandins E₁, E₂, F₂ₐ, and I₂ (which was detected as 6-Keto-PGF₁ₐ). Analysis of cell subpopulations of seventeeen tumor samples showed that the macrophage-enriched cells were responsible for the large majority of prostaglandin production in vitro (p ≤ 0.02). It was found that the major prostaglandins were PGE₂ and PGI₂. The range of values measured for macrophage produced prostaglandin E₂ was 1.1 to 704.8 ng/ml after a 24 hour incubation of 10⁶ cells. Prostaglandin I₂ was also produced by the macrophage-enriched cell subpopulation with values ranging from 1.2 to 334.3 ng/ml. This is the first report of prostaglandin I₂ production by host macrophages infiltrating human tumors. Studies of the effect of macrophage depletion and reconstitution on the ability of tumor cells to form colonies in vitro were performed. A two layer soft agar assay was used to evaluate tumor cells clonogenicity. The results demonstrated that macrophages infiltrating human carcinoma samples function in a supportive role for tumor cell colony formation in vitro. Using a prostaglandin synthesis inhibitor, flurbiprofen, it was shown that this support was not the result of a direct effect of prostaglandins on the tumor cells. Possible roles for macrophage produced prostaglandins in cancer are discussed.

The mechanical and biochemical effects of prostaglandins E₁ and F₁α were studied on rat heart using isolated right and left atria and the Langendorff perfused whole heart preparation. Preliminary experiments were performed to establish optimal perfusion conditions for the Langendorff preparation. Hearts were perfused at different perfusate temperatures and at different filling pressures. Heart rate and coronary flow rate were monitored at all combinations of perfusate temperature and filling pressure. A constant temperature water recirculating pump setting of 38°C and a filling pressure of 40 cm of H₂O were chosen as the optimal perfusion conditions. Hearts perfused under the above conditions responded normally to bolus injections of isoproterenol. Isoproterenol produced a dose dependent increase in the contractile force of the Langendorff preparation and the cyclic AMP increasing effect of isoproterenol preceeded the positive inotropic effect in a time course study. Prostaglandin E₁ (PGE₁) did not produce any effect on heart rate or tension development in the Langendorff preparation, when infused over a dose range of 0.03 Ug to 5.0 μg/min. Infusion of prostaglandin F₁α (PGF₁α) (0.1 to 5.0 μg/min) produced an increase in tension development which was associated with a negative chronotropic effect. The positive inotropic effect of PGF₁α was secondary to the drop in rate as the positive inotropic effect was completely abolished when the hearts were paced at 6 Hz. In the rat right atrium, PGE₁ produced a dose dependent increase in the rate which developed very slowly. PGE₁ had no effect on the tension development of the.rat left atrium. PGF₁α produced a slow, dose dependent positive chronotropic effect on the right atrium and a slight but not significant effect on the force of contraction of the left atrium. Both prostaglandins were equipotent in exerting their positive chronotropic effect on the right atrium. The PD₁ value for PGE₁ was 5.54 ±0.25 and for PG₁α 5.59 ± 0.18. In the right atrium 10⁻⁴ M PGE₁ increased the rate and cyclic AMP content without changing phosphorylase a activity or cyclic GMP content. PGE₁ (10⁻⁴M) slightly but not significantly increased the left atrial cyclic AMP con-; tent and did not change the cyclic GMP content. 10⁻⁴ M PGF₁α did not affect either right or left atrial cyclic AMP or cyclic GMP content. The effect of a 1 μg/min infusion of either PGE₁ or PGF₁α on the changes of cyclic AMP and cyclic GMP contents and phosphorylase a activity with time were studied in the Langendorff preparation. A 1 μg/min infusion of PGE₁ increased the myocardial cyclic AMP levels by about 57 per cent above control at 30 sees after starting the infusion and the cyclic AMP levels were still elevated by 50 per cent over control at the end of a one minute period of infusion. PGE₁ did not change cyclic GMP content or phos-phorylase a activity at any time point. A 1 ug/min infusion of PGF₁α did not alter cyclic AMP and cyclic GMP levels or phosphorylase a activity in the rat heart within one minute. These results supported the earlier reported observation that PGE₁ selectively increased rat myocardial cyclic AMP content without altering myocardial contractile force or phosphorylase a activity. PGE₁ might be selectively increasing a pool of cyclic AMP and activating a cyclic AMP-dependent protein kinase in the cardiac cells that is not associated with contractile force or phosphorylase activation. PGF₁α did not possess this selective effect of PGE₁. Cyclic GMP is not involved in the mediation of the actions of either PGE₁, or PGF₁α, on the rat heart. / Pharmaceutical Sciences, Faculty of / Graduate

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