Inhibition of cardiac myocyte maturation by 1,25-dihydroxyvitamin D3.

The signals that regulate cardiac myocyte maturation in the neonatal heart are not completely understood. In our study we examined the effects of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] on primary cultures of ventricular myocytes isolated from neonatal rat hearts. Our data show that 1,25-(OH)2D3 inhibited an increase in both the muscle-specific form of creatine kinase and V1 myosin isoenzyme levels in myocytes induced to mature by serum withdrawal. Thus, in contrast to other cell types studied to date, in the heart, 1,25-(OH)2D3 blocks cell maturation. Treating cultures with phorbol 12-myristate 13-acetate induced a decrease in the muscle-specific isoenzyme of creatine kinase similar to the effects of 1,25-(OH)2D3. Interestingly, we found that staurosporine, a protein kinase-C inhibitor, blunts the effects of 1,25-(OH)2D3 and has the opposite effect of phorbol 12-myristate 13-acetate on cultured myocytes induced to mature by serum withdrawal. Thus, our data identify a novel role for 1,25-(OH)2D3 in the regulation of myocardial development and suggest that 1,25-(OH)2D3 may be acting through a protein kinase-dependent mechanism to maintain cardiac myocytes in an immature state.

Serum percent-free PSA does not predict extraprostatic spread of prostate cancer.

Percent-free prostate-specific antigen (proportion of free prostate-specific antigen [PSA] to total PSA) has been shown recently in studies on frozen serum samples to be more useful than total PSA alone in distinguishing prostate cancer from benign conditions of the prostate gland. The primary purpose of our study was to determine whether percent-free PSA could predict extraprostatic spread of prostate cancer. We also sought to evaluate the freeze-thaw stability of free PSA. Percent-free PSA values in fresh serum samples were compared with those in aliquots subjected to one to five freeze-thaw cycles. Percent-free PSA values in frozen serum samples from 130 men undergoing radical prostatectomy for clinically localized prostate cancer were compared across pathologic stages. Free PSA levels remained stable for up to five freeze-thaws. Great overlap was found in percent-free PSA values for men with organ-confined disease and those with extraprostatic spread. These results indicate that multiple freeze-thaw cycles do not significantly affect free PSA levels and percent-free PSA is not useful in identifying ideal candidates for radical prostatectomy.

Half-life determination of serum free prostate-specific antigen following radical retropubic prostatectomy.

OBJECTIVES: Prostate-specific antigen (PSA) continues to be the the most clinically useful tumor marker for prostate cancer. Recently, several molecular forms of PSA have been detected and characterized. These specific forms, including free PSA and PSA complexed to alpha 1-antichymotrypsin, can be measured and their proportions determined. In doing so, the sensitivity of PSA as a tumor marker can be maintained while the specificity is improved. In order to maximize the clinical utility of free PSA, the half-life and elimination kinetics of free PSA from the serum were determined. METHODS: Twenty-five patients, ages 43-74 years (mean 60 years) with biopsy proven, organ-confined adenocarcinoma of the prostate who underwent anatomic radical retropubic prostatectomy, were identified. For each patient, venous blood samples were obtained preoperatively, and at 60-minute intervals beginning 1 hour after the prostate was removed. The specimens were handled and stored in a consistent fashion. Using the AxSYM immunoassay analyzer (Abbott Diagnostics, Abbott Park, IL), the serum free PSA values were determined and plotted as a function of time for each patient. From the 25 individual elimination curves that were generated, the half-life of serum free PSA was determined. RESULTS: The mean half-life of serum free PSA was 110 minutes +/- 18.6 minutes (SD). Analysis of the individual and cumulative elimination curves indicates that the elimination of free PSA from the serum following radical prostatectomy follows a biphasic pattern. CONCLUSIONS: Unlike PSA, which has a half life of 2-3 days, the half-life of serum free PSA is 110 minutes (1.83 hours). This short half-life may have significant implications for the use of percentage of free PSA as a clinically useful tool in distinguishing patients with early, curable prostate cancer from men with benign prostatic hyperplasia (BPH) only.

Effect of vehicle on the nasal absorption of epinephrine during cardiopulmonary resuscitation.

STUDY OBJECTIVES: We have shown in previous studies that epinephrine administered intranasally is a feasible route of administration during cardiopulmonary resuscitation (CPR). To promote the absorption of epinephrine we administered phentolamine prior to epinephrine and used a bile salt as a vehicle to dissolve the epinephrine. The purpose of this study was to compare the effect of two different vehicles (bile salt vs surfactant) in promoting the absorption of nasally administered epinephrine during CPR and to determine their effects on the nasal mucosa. STUDY DESIGN: A randomized, blinded study. SETTING: A controlled laboratory environment. SUBJECTS: Eleven mongrel dogs. INTERVENTIONS: Each dog underwent 3 minutes of unassisted ventricular fibrillation (VF) followed by 7 minutes of VF with CPR. Five minutes after the start of VF, 10 dogs received intranasal phentolamine 0.25 mg/kg/nostril followed 1 minute later by intranasal epinephrine 7.5 mg/kg/nostril. The epinephrine was dissolved in a randomly assigned vehicle consisting of either taurodeoxycholic acid (group A, bile salt) or polyoxyethylene-9-lauryl ether (group B, surfactant). One animal acted as a control and received 0.9% sodium chloride nasally. MEASUREMENTS AND MAIN RESULTS: Data from eight dogs (one control) were included for analysis. Histology of the nasal cavity demonstrated severe multifocal erosion and ulceration of the respiratory epithelium for groups A and B compared with the control. The severity was similar between the two groups. In addition, no significant differences in plasma epinephrine concentrations or blood pressure responses were seen between the groups. CONCLUSION: Based on histology, polyoxyethylene-9-lauryl ether offered no advantage over taurodeoxycholic acid in its effect on the nasal mucosa. The data available for changes in epinephrine concentration and pressure also suggest no difference between the two vehicles in promoting the absorption of epinephrine during CPR in an animal model.

Endogenous norepinephrine regulates tumor necrosis factor-alpha production from macrophages in vitro.

Evidence for the extraneuronal accumulation of norepinephrine has been demonstrated to occur in macrophage (M phi), yet the physiologic role of this system remains undefined. We have assessed the response of murine peritoneal M phi to adrenergic antagonists. We have also defined a physiologic role of a M phi-associated pool of the nonspecific adrenergic agonist norepinephrine. We investigated the constitutive involvement of alpha-adrenergic and beta-adrenergic receptors in LPS-induced TNF-alpha production. CFA-elicited M phis were incubated with LPS (1 microgram/ml) in the presence or absence of adrenergic agonists and/or antagonists. Although stimulation of alpha-adrenergic receptors increased TNF production and gene expression, beta-adrenergic receptors decreased it. Interestingly, when adrenergic antagonists along with LPS alone were added to M phi, they generated the response opposite to that produced by their suitable agonist, suggesting a role for endogenous norepinephrine in M phi. Thus, although alpha 2-adrenergic antagonists attenuated TNF production, beta-adrenergic antagonists augmented TNF expression in a concentration-dependent manner. Norepinephrine and epinephrine were found in M phi as determined by HPLC and LPS stimulation induced a significant decrease in their content. M phis were also incubated with LPS or medium only, washed, and then challenged 12 h later with LPS. When given a second LPS stimulation, M phis were found to have an increased response to alpha 2-adrenergic agonists and decreased response to alpha 2-adrenergic antagonists. Therefore, M phi-associated norepinephrine appears to regulate LPS-induced TNF production in an autocrine fashion.

Interference by goat-derived anti-lymphocyte globulin in a double-antibody radioimmunoassay of digoxin.

Therapy with anti-lymphocyte and anti-thymocyte globulins, two animal-derived preparations used as immunosuppressive agents, results in high concentrations of foreign immunoglobulin circulating in plasma. We describe a case that illustrates how the presence of these immunoglobulins can interfere with double-antibody radioimmunoassay procedures.

Stability of tacrolimus (FK 506) and cyclosporin G in whole blood.

The stability of two new immunosuppressants, tacrolimus and cyclosporin G (CsG), was evaluated in whole blood following incubation of patient specimens at ambient temperature or 4 degrees C for time periods of 2 (48 h) and 7 days (168 h) after collection. No decrease in CsG concentrations was noted over a 7-day period for specimens stored at either ambient temperature or 4 degrees C. For tacrolimus, the concentrations of drug in whole blood stored at ambient temperature for 7 days had a slight downward trend. At 7 days, the ratio of tacrolimus concentrations to day zero concentrations ranged from 0.86 to 1.15, with a mean decrease of 5%. However, these changes are within the published precision characteristics of the tacrolimus immunoassay. Evaluation of a smaller group of whole blood specimens provided evidence of adequate stability of these two drugs over a 13-day period.

Natriuresis associated with elevated plasma atrial natriuretic hormone during supraventricular tachycardia.

Elevated plasma levels of atrial natriuretic hormone (ANH) have been found in patients during paroxysmal supraventricular tachycardia (SVT) and other clinical syndromes. However, physiologic effects of this endogenous ANH have not been demonstrated. To determine whether the rise in ANH during SVT is associated with either a natriuresis or kaliuresis, urine sodium and potassium levels were measured in five patients at baseline and during SVT simulated by rapid atrioventricular pacing. Plasma ANH levels increased from 149 +/- 35 pmol/L at baseline to 187 +/- 31 pmol/L (p = 0.007) during SVT. Plasma vasopressin and renin levels were unchanged. Urine sodium levels increased 49% from 1.54 +/- 0.66 mEq/hr at baseline to 2.29 +/- 0.89 mEq/hr (p = 0.044) during SVT, and urine potassium levels increased 22% from 4.14 +/- 0.10 mEq/hr to 5.04 +/- 1.25 mEq/hr (p = 0.018). Urine sodium and potassium levels returned to baseline values 1 hour after pacing. Thus elevated plasma levels of ANH during SVT are associated with both a natriuresis and kaliuresis, which may represent physiologic effects of the endogenously secreted hormone.

Effect of dose on the nasal absorption of epinephrine during cardiopulmonary resuscitation.

Delay in epinephrine administration during cardiopulmonary resuscitation (CPR) due to technical difficulties in obtaining an access site may be detrimental. To avoid this potential delay, we have previously shown that intranasal administration of phentolamine and epinephrine is a rapidly obtainable and feasible route of administration during CPR. A randomized blinded dose ranging study was performed in a controlled laboratory environment. Thirty mongrel dogs were randomized to one of the following dosage regimens: phentolamine, 0.25 or 2.5 mg/kg/nostril; epinephrine, 0.075, 0.75, or 7.5 mg/kg/nostril. Phentolamine was administered intranasally 1 minute before the intranasal administration of epinephrine to improve absorption. Each dog underwent 3 minutes of ventricular fibrillation followed by 7 minutes of closed chest CPR. Epinephrine was administered was administered at 3 minutes of CPR. Data from 26 dogs were included for analysis. Treatment B (0.25 and 7.5 mg/kg/nostril of phentolamine and epinephrine, respectively) produced the greatest elevation in coronary perfusion pressure (17 +/- 11 vs. 4 +/- 3 mm Hg for the next highest group, P < .003) and in epinephrine plasma concentrations (1,403 +/- 1,400 vs 290 +/- 182 ng/mL for the next highest group, P > .05). In addition, treatment B had the highest resuscitation rate, 100% (5/5) versus 0% to 50% for the other groups (P < .05). These data show that there is a dose response effect, with 0.25 and 7.5 mg/kg/nostril of phentolamine and epinephrine being the optimal dose studied. In addition, when administered in appropriate doses, intranasal epinephrine reaches the systemic circulation and increases coronary perfusion pressure and successful resuscitation during CPR in this canine model.

The effects of a blood-salvaging device on blood containing a hemoglobin-based oxygen carrier, HBOC-201.

OBJECTIVES: Hemoglobin-based oxygen carriers will be used concurrently with intraoperative blood salvage. The effects of salvage and processing on blood containing one such solution (HBOC-201; Biopure Corp, Boston, MA) were studied. DESIGN: Prospective, randomized. SETTING: Laboratory. INTERVENTIONS: Sixteen blood units from healthy volunteers had either HBOC (1,500 mg/dL; n = 10) or normal saline (equivalent volume; n = 6) added. All units were salvaged and processed using a blood salvage device. Samples were analyzed for the concentration and molecular weight distribution of plasma hemoglobin and red cell morphology presalvage (pre) and following processing and washing (post 1). Five of the HBOC units underwent a second 1,000 mL wash (post 2). MEASUREMENTS AND MAIN RESULTS: Processing and washing decreased the concentration of plasma hemoglobin (mg/dL) in HBOC units (1311 +/- 265 pre to 27.8 +/- 19.6 post 1 to 6.5 +/- 2.19 post 2), but did not change the plasma hemoglobin concentration in saline units (2.05 +/- 1.27 pre v 3.18 +/- 0.79 post 1). Total plasma hemoglobin in HBOC units (6.56 +/- 2.19) was significantly greater than in saline units (3.18 +/- 0.79), even after the second wash (post 2). The concentration of unstable hemoglobin in the plasma phase was not different between groups. Red cell morphology was altered by the salvage process but was not different between groups. CONCLUSIONS: Salvage and processing of blood containing HBOC yield concentrated red cells that are indistinguishable from those obtained from blood without HBOC. Residual HBOC remains but is unchanged from the HBOC initially administered.