Unilateral dermis-fat graft implantation in the pediatric orbit.

The purpose of this study was to evaluate the role of the dermis-fat graft (DFG) as an orbital implant in the pediatric age group. A retrospective study was made of a series of 16 patients who had undergone unilateral orbital implantation of a DFG. The ages of the patients at the time of surgery ranged from 2 months to 17 years, with followup ranging from 2 to 15 years. Growth of the graft was clinically apparent in the younger children. Increasing proptosis required surgical debulking of the graft in six of eight children who were 4 years old or younger at the time of DFG implantation. None of the eight children who were 9 years or older at the time of DFG implantation required surgical debulking. Indeed, five of the older patients demonstrated some degree of graft atrophy. Dermis-fat grafts placed in the orbits of young children appear to grow after implantation. This growth of the implant may help stimulate orbital growth, potentially leading to more symmetry between the involved and uninvolved sides.

Cardiac hypertrophy in rats after intravenous administration of CI-959, a novel antiinflammatory compound: morphologic features and pharmacokinetic and pharmacodynamic mechanisms.

CI-959 is an antiallergic/antiinflammatory agent currently in development. In rats, daily bolus intravenous administration of CI-959 at doses > or = 10 mg/kg was associated with development of cardiac hypertrophy. There was no morphologic or biochemical evidence of myocyte injury, and cardiac hypertrophy rapidly reversed after treatment was discontinued. Cardiac hypertrophy was not evident when CI-959 was given orally or by continuous intravenous infusion with ALZA osmotic pumps. Maximum plasma drug concentrations (Cmax) were significantly higher when CI-959 was given by bolus intravenous injection, suggesting that cardiac effects were dependent on high Cmax concentrations. When neonatal rat cardiomyocytes were exposed to CI-959 in vitro, there was no evidence of myocyte enlargement or increased protein content. Cardiac hypertrophy was prevented by pretreatment with nonselective beta- and beta 1-selective adrenoceptor blockers as well as with central sympatholytics. beta 2- and alpha-adrenoceptor blockers were ineffective in preventing cardiac hypertrophy. Bolus intravenous CI-959 administration resulted in prolonged hypotension and associated increase in plasma catecholamine levels, with apparent inhibition of reflex tachycardia. We conclude that CI-959-associated cardiac hypertrophy in rats was not a direct drug effect but instead was probably mediated by endogenous catecholaminergic stimulation of cardiac beta 1-adrenoceptors.

Increased risk of renal cell cancer among women using diuretics in the United States.

Use of prescription diuretics and incidence of renal cell cancer have increased in the United States in the past 25 years. Recent interview-based epidemiologic studies have reported an association between diuretic use and renal cell cancer risk. Our study evaluated this hypothesis using, for the first time, medical records as the source of the information on prescription diuretic use. Using medical records of women from a prepaid health plan, we identified 191 cases and 191 controls matched on age, membership duration, and membership at diagnosis. Diuretics use and history of potential confounding factors were ascertained by a standardized review of the medical records of each subject, without reference to case or control status. There was a strong and statistically significant association between renal cell cancer and prescription diuretics (odds ratio [OR] adjusted for hypertension, smoking, and obesity = 2.9, 95 percent confidence interval [CI] = 1.7-4.7). Risk tended to increase with dose, measured by number of prescriptions. Since renal cancer can induce hypertension, which is treated by diuretics, and thereby confound the association with diuretics, we examined diuretic use 10 or more years prior to diagnosis when secondary hypertension would be unlikely. The OR for prescriptions 10 or more years before diagnosis was 3.5 (CI = 1.7-7.4). Our results support earlier reports of an excess risk of renal cell cancer among users of prescription diuretics and indicate need for further study to evaluate this relationship, especially due to the extensive use of diuretics and the increasing incidence of this cancer.

Formation of chloramine derivatives of histamine: role of histamine chloramines in bronchoconstriction.

Hypochlorous acid generated by myeloperoxidase reacts with histamine to produce chloramines. At pH 7, one mole of histamine monochloramine (HisCl) was generated per mole of H2O2 provided as substrate for myeloperoxidase. At pH 5, one mole of histamine dichloramine (HisCl2) was generated per two moles of H2O2. HisCl and HisCl2 had two and four oxidizing equivalents per molecule, respectively. In vitro, 30 microM HisCl and HisCl2 induced mepyramine-sensitive guinea pig lung parenchyma contraction with 89 and 56 percent of the response of an equivalent concentration of histamine. Pretreatment of lung strips with chloramines reduced the subsequent contractile response of the tissues to methacholine. These results suggest that H-1 histamine receptors provide for targeting of histamine chloramines to pulmonary tissue which may facilitate modification of tissue responses.

A novel cyclosporine binding assay.

The binding of [3H]cyclosporine A (CsA) to BALB/c mouse spleen cells was examined with a novel centrifugation assay which rapidly removes free [3H]CsA from cell surfaces with a minimal loss of low affinity specifically bound [3H]CsA. A single class of specific and saturable CsA binding sites was found under equilibrium binding conditions. Scatchard analysis of the data resulted in a straight line with KD and Bmax values of 95 nM and 2.4 million binding sites/cell, respectively. Kinetic studies conducted with a wider range of [3H]CsA concentrations revealed two distinct binding sites, with KD's of 290 nM and 9.6 microM, respectively. [3H]CsA bound only nonspecifically to phosphatidylcholine:cholesterol liposomes. Specific [3H]CsA binding sites were found in murine WEHI-5 B-lymphoma cells, murine N1E-115 neuroblastoma cells and human A204 rhabdosarcoma cells. We conclude from these results that there are at least two nonlipid CsA binding sites in BALB/c mouse spleen cells and that CsA binding sites are present in both lymphoid and nonlymphoid tissue.

Collateral sensitivity to cross-linking agents exhibited by cultured L1210 cells resistant to oxazaphosphorines.

The sensitivity of cultured L1210 and P388 cells, sensitive (L1210/0, P388/0) and resistant (L1210/CPA, P388/CPA) to cyclophosphamide in vivo, to five oxazaphosphorine and eight nonoxazaphosphorine cross-linking agents was determined. Each of the resistant sublines was cross-resistant to all of the oxazaphosphorines tested. The P388/CPA cell line was also cross-resistant to all of the nonoxazaphosphorines but, in most cases, not nearly to the same extent. The L1210/CPA cell line was collaterally sensitive to all but one of the nonoxazaphosphorines, in which case it was equisensitive. Changes in sensitivity could not be accounted for by changes in intracellular pH values, or by changes in intracellular inorganic phosphate or acid-soluble organic phosphate concentrations. Inasmuch as the L1210/CPA cell line was specifically resistant to the oxazaphosphorines, identification of the phenotypic basis for this resistance should serve to identify a potentially important determinant with regard to the basis for the oncotoxic specificity of this group of agents.

Further studies on the conversion of 4-hydroxyoxazaphosphorines to reactive mustards and acrolein in inorganic buffers.

The rates at which the 4-hydroxyoxazaphosphorines, 4-hydroxycyclophosphamide and 4-hydroxyifosfamide, are converted to reactive mustards and acrolein in phosphate and bicarbonate buffers were determined as a function of pH, ionic strength, temperature, and buffer concentration. Conversion was first-order with respect to both the 4-hydroxyoxazaphosphorine and phosphate or carbonate serving as a catalyst. The catalytic constant for dianionic phosphate-catalyzed conversion of 4-hydroxyifosfamide to isophosphoramide mustard and acrolein at 37 degrees was 0.189 min-1; a value of 0.194 min-1 M-1 was obtained when dianionic phosphate-catalyzed conversion of 4-hydroxycyclophosphamide to phosphoramide mustard and acrolein was examined. A catalytic constant of 3.09 min-1 M-1 was obtained for carbonate-catalyzed conversion of 4-hydroxycyclophosphamide to phosphoramide mustard and acrolein. Hydroxyl ion and water also catalyzed the reaction; catalytic constants were 2710 and 0.000006 min-1 M-1, respectively. The rate at which the 4-hydroxyoxazaphosphorines were converted to reactive mustards and acrolein in phosphate buffer increased as the pH, ionic strength, and temperature increased. The energy of activation was about 20 kcal/mol. Dianionic phosphate, carbonate, hydroxyl ion, and water were apparently acting as general base catalysts of the rate-limiting step (probably the conversion of the intermediate aldehyde to the corresponding reactive mustard and acrolein) of the overall reaction, although specific base-general acid catalysis could not be ruled out. Bifunctional catalysis of the rate-limiting step did not appear to be significant and certainly was not obligatory as concluded previously by our laboratory. Assuming that the oncotoxic specificity of the oxazaphosphorines resides with the 4-hydroxyoxazaphosphorines and that their cytotoxic effect at therapeutic doses is largely mediated by the reactive mustards released within cells, these observations offer the possibility that intracellular general base catalytic activity serves as an important determinant with regard to the oncotoxic potential and specificity of the oxazaphosphorines. General base catalytic activity varies directly with pH, ionic strength, temperature, and the concentration of the base. The influence of some of these factors on the development of cyclophosphamide-induced bladder toxicity has already been demonstrated.

Influence of diuretics on urinary general base catalytic activity and cyclophosphamide-induced bladder toxicity.

The influence of diuretics on the induction of bladder toxicity by cyclophosphamide was investigated in rats. Following ip administration, about 3.5% of the cyclophosphamide was excreted as 4-hydroxycyclophosphamide. This amount was found to be compatible with the view that the urotoxic effects of cyclophosphamide are caused by the acrolein generated in the urine from 4-hydroxycyclophosphamide, the primary metabolite of cyclophosphamide. In situ, acrolein was more potent than 4-hydroperoxycyclophosphamide with regard to producing an increase in bladder weight; phosphoramide mustard was essentially without urotoxic activity. The urotoxic potency of 4-hydroperoxycyclophosphamide, but not that of acrolein, increased as the pH and/or the phosphate concentration of the infusion medium increased. This was as expected in view of the knowledge that release of acrolein from 4-hydroxycyclophosphamide or 4-hydroperoxycyclophosphamide is facilitated by the presence of general base catalysts, eg, phosphate and bicarbonate, and that the rate at which this reaction proceeds in the presence of these catalysts increases as their concentration and the pH increases. In vivo, diuretics that acidified the urine, eg, ammonium chloride and furosemide, prevented the increase in bladder weight ordinarily elicited by the dose of cyclophosphamide used in these experiments. In contrast, a diuretic, acetazolamide, that markedly increased urinary bicarbonate concentration and alkalinized the urine, did not. None of the diuretics altered the systemic metabolism and urinary excretion of cyclophosphamide nor did they alter the systemic action, as judged by spleen weight, of cyclophosphamide. These observations demonstrate that the pH of the urine and the urinary concentration of general base catalysts greatly influence the urotoxic potential of oxazaphosphorines such as cyclophosphamide. They indicate that while the use of acidifying diuretics is likely to be beneficial in minimizing oxazaphosphorine-induced bladder toxicity, the use of alkalinizing diuretics may not be helpful.

Conversion of 4-hydroperoxycyclophosphamide and 4-hydroxycyclophosphamide to phosphoramide mustard and acrolein mediated by bifunctional catalysis.

The rates at which 4-hydroperoxycyclophosphamide and 4-hydroxycyclophosphamide are converted to phosphoramide mustard and acrolein were determined as a function of buffer composition, buffer concentration, and pH. Conversion of 4-hydroperoxycyclophosphamide to 4-hydroxycyclophosphamide in 0.5 M Tris buffer, pH 7.4, 37 degrees, was first-order (k = 0.016 min-1), but subsequent conversion of 4-hydroxycyclophosphamide to phosphoramide mustard and acrolein under these conditions was negligible. Phosphoramide mustard and acrolein were readily generated from 4-hydroperoxycyclophosphamide or 4-hydroxycyclophosphamide when either of these agents was placed in phosphate buffer. Conversion of 4-hydroxycyclophosphamide to phosphoramide mustard and acrolein was first-order with respect to 4-hydroxycyclophosphamide (k = 0.126 min-1 in 0.5 M phosphate buffer, pH 8, 37 degrees) as well as first-order with respect to phosphate serving as a catalyst. The rate-determining step in the reaction was pH dependent only insofar as the hydrogen ion concentration governed the relative amounts of monobasic and dibasic phosphate present. Pseudo-first-order rate constants were 0.045 M-1 min-1 for monobasic phosphate and 0.256 M-1 min-1 for dibasic phosphate. The role of phosphate in this reaction was as that of a bifunctional catalyst. The reaction was not subject to specific or general, acid or base, catalysis. Other bifunctional catalysts such as glucose-6-phosphate and bicarbonate also catalyzed the reaction, albeit less efficiently. Aldophosphamide apparently exists only transiently; its presence could not be established by 31P nuclear magnetic resonance spectroscopy. We conclude that, in the reaction sequence 4-hydroxycyclophosphamide leads to aldophosphamide leads to phosphoramide mustard + acrolein, the conversion of 4-hydroxycyclophosphamide to aldophosphamide is rate limiting and is subject to bifunctional catalysis; this reaction can proceed efficiently only in the presence of a bifunctional catalyst. Assuming that the oncotoxic specificity of cyclophosphamide resides with 4-hydroxycyclophosphamide and that its cytotoxic effect at therapeutic doses is largely mediated by phosphoramide mustard released within cells, these observations offer the possibility that the intracellular concentration of bifunctional catalysts, whether in the form of inorganic phosphates, organic phosphates, enzymes, or other species, serve as important determinants with regard to the oncotoxic potential and specificity of cyclophosphamide. Similarly, the concentration of bifunctional catalysis in the urine as well as the pH of the urine may be important with regard to the potential of cyclophosphamide to induce, via acrolein, hemorrhagic cystitis.