Bone loss is a known complication of severe burn injury. It is, in part, due to increased endogenous glucocorticoids that contribute to the reduction in bone formation and osteoblast differentiation, hypercalciuria secondary to hypoparathyroidism, and vitamin D deficiency. In this study we attempted to prevent post-burn bone loss by acute intravenous administration of the bisphosphonate pamidronate. We enrolled 43 children, with burns of > 40% total body surface area, in a randomized, double-blind, placebo-controlled study, administering the study drug within 10 days of burn injury and again 1 week later. Dual energy X-ray absorptiometry was performed prior to drug therapy, at hospital discharge and at 6 months post-burn. Urine specimens were obtained at baseline and discharge for determination of calcium and free deoxypyridinoline. Blood was obtained along with the urine specimens for measurement of intact parathyroid hormone (iPTH) and ionized calcium (Ca) levels. Following doxycycline labeling, intra-operative iliac crest bone biopsies were obtained, and bone histomorphometry was determined. At time of discharge there were no differences in total body bone mineral content (BMC), but lumbar spine BMC was significantly higher in the pamidronate group (P < 0.005). By 6 months post-burn the differences in lumbar spine BMC persisted, but, now, total body BMC was significantly higher in the pamidronate group (P < 0.05). Bone histomorphometry and levels of urine Ca and free deoxypyridinoline failed to show significant increases in bone formation or decreases in bone resorption. Pamidronate did not exacerbate the hypocalcemia in burn patients. In summary, acute intravenous pamidronate administration following burns may help to preserve bone mass, perhaps by inhibiting the glucocorticoid-induced apoptosis of osteoblasts and osteocytes.

Natural polyamines, i.e. putrescine, spermidine and spermine, are excellent promoters of triplex DNA. Using melting temperature (Tm) measurements and CD spectroscopy, we found that structural alterations on spermidine backbone, including methylation, or acetylation at the N1-, N4- and/or N8-positions had a profound influence on the stability and conformation of poly(dA).2poly(dT) triplex. The conformation of the polynucleotide complex underwent sequential changes from B-DNA to triplex DNA as the concentration of spermidine increased from 0 to 50 microM in a buffer containing 10 mM sodium cacodylate and 1 mM EDTA (pH 7.2). At 60 microM spermidine, the CD spectrum of triplex DNA was comparable with that of psi-DNA, with a strong positive band centred around 260 nm. A negative band was also found at 295 nm. At higher concentrations of spermidine, however, the intensity of the positive band progressively decreased and the peak intensity was found at a 1:0.3 molar ratio of DNA phosphate:spermidine. Temperature-dependent CD analysis showed that the psi-DNA structure melted to single-stranded DNA at temperatures above the Tm determined from the absorbance versus temperature profile. Comparable effects were exerted on the conformation of triplex DNA by Co(NH3)6(3+), an inorganic trivalent cation. Substitution of the N4-hydrogen of spermidine by a cyclohexyl ring or the fusion of the N4-nitrogen in a cyclic ring system, as in piperidine, enhanced the ability of spermidine analogues to stabilize triplex and psi-DNA forms over a wider concentration range compared with spermidine. These data demonstrate a differential effect of trivalent cations in stabilizing triplex DNA and provoking unusual conformations such as psi-DNA. Synthetic homologues of spermidine that stabilize triplex DNA over a wider range of concentrations than that stabilized by spermidine itself might have potential therapeutic applications in the development of an anti-gene strategy against several diseases, including cancer and AIDS.

We studied the effects of natural and synthetic polyamines on the conformation of an oligodeoxyribonucleotide (ODN1) harboring the estrogen response element (ERE) by circular dichroism (CD) spectroscopy and polyacrylamide gel electrophoresis. Putrescine and spermidine had no marked effect on the CD spectrum of ODN1. In contrast, spermine provoked and stabilized two characteristic changes in the CD spectrum. The first change was indicated by an increase in the intensity of the CD band at 280 nm at 0.5 mM spermine in Tris-HCl buffer containing 50 mM NaCl. This change appears to be related to changes in base tilt and conformational alterations similar to A-DNA. At 1-2 mM spermine, the CD spectrum was characterized by a loss of positive bands at 220 and 270 nm. This change might have contributions from polyamine-induced condensation/aggregation of DNA. Spectral measurements were also conducted in Tris-HCl buffer containing 150 mM NaCl to minimize contributions from condensation and aggregation of ODN1. Under these conditions, CD spectral changes were retained by (ODN1), although the magnitude of the change was diminished. In contrast, a control oligdeoxyribonucleotide (ODN2) having similar base composition did not show any significant change in the CD spectrum in the presence of 150 mM NaCl and 2 mM spermine. The changes in the CD spectrum of ODN1 were highly sensitive to polyamine structure, as evidenced by experiments using spermine analogs with altered number of -CH2- groups separating the amino and imino groups. Electrophoretic mobility shift analysis further showed ODN1 stabilization by spermine and its analogs. These data demonstrate the ability of an ODN containing ERE to undergo conformational transitions in the presence of polyamines and suggest a possible mechanism for polyamine-mediated alterations in the interaction of estrogen receptor with ERE.

A study was conducted to examine tolerability and pharmacodynamics of single doses of yohimbine in healthy volunteers using measures of mood, heart rate, blood pressure, and serum catecholamine levels. Participants were given single oral doses of yohimbine hydrochloride as high as 21.6 mg. Plasma concentrations of yohimbine, epinephrine, norepinephrine, and MHPG (3-methoxy-4-hydroxyphenylethylene-glycol) were quantified by means of high-performance liquid chromatography with electrochemical detection. Mood was assessed by visual analogue scale (VAS), the Profile of Mood States, and the Spielberger State Anxiety Index. Yohimbine was well tolerated and rapidly absorbed and eliminated. Dose-related increases in area under the concentration-time curve (AUC) were observed. Administration of yohimbine in the presence of a high fat meal diminished both the rate and extent of drug absorption. Significant intersubject variability in the pharmacokinetic parameters of yohimbine was observed, with some individuals exhibiting greatly increased oral bioavailability of yohimbine. Increases in blood pressure, respiratory rate, plasma catecholamine levels, and total VAS score were observed in participants with elevated AUC values. The AUC of yohimbine had the largest effect on total VAS score. The results indicate that higher doses of yohimbine are both well tolerated and produce dose-related increases in AUC, which are associated with more pronounced autonomic effects. Increases in respiratory rate and plasma MHPG appear to be the most reliable pharmacodynamic measures for single oral doses of yohimbine. Individual differences in the pharmacokinetics of yohimbine are important in determining pharmacodynamic effects and should be considered in evaluations of its clinical effectiveness.

We sought to determine the safety, pharmacodynamic response, and single- and multiple-dose pharmacokinetic profile of yohimbine hydrochloride. Thirty-two healthy volunteers received 6 days of yohimbine, 5.4 mg 3 times daily (t.i.d.), 10.8 mg t.i.d., 16.2 mg t.i.d., or 21.6 mg twice daily (b.i.d.), with determination of plasma catecholamine levels and mood/anxiety-inventory scores. The pharmacokinetic profile of yohimbine was determined after the first and last dose. Yohimbine exhibited one-compartment elimination in most subjects, with dose-dependent increases in maximal concentration (Cmax) and area under the curve (AUC) but no evidence of drug accumulation. At least two subjects in each cohort exhibited two-compartment elimination of yohimbine, with nonsignificant increases in day 7 AUC, Cmax, and terminal elimination half-life (t1/2beta). Plasma catecholamine levels increased significantly in relation to both average yohimbine AUC and Cmax, but there were no significant effects on heart rate, blood pressure, or anxiety/mood-inventory scores. The single- and multiple-dose pharmacokinetic profile of yohimbine exhibits a substantial degree of interpatient and intrapatient variability, possibly resulting from variability in first-pass and hepatic metabolism. There is a significant correlation between plasma norepinephrine levels and yohimbine AUC or Cmax. Further multiple-dose studies are warranted definitively to address the relation between yohimbine AUC or Cmax and pharmacologic effect.