Neutron activation of holmium poly(L-lactic acid) microspheres for hepatic arterial radio-embolization: a validation study.

Poly(L-lactic acid) microspheres loaded with holmium-166 acetylacetonate (166Ho-PLLA-MS) are a novel microdevice for intra-arterial radio-embolization in patients with unresectable liver malignancies. The neutron activation in a nuclear reactor, in particular the gamma heating, damages the 166Ho-PLLA-MS. The degree of damage is dependent on the irradiation characteristics and irradiation time in a particular reactor facility. The aim of this study was to standardize and objectively validate the activation procedure in a particular reactor. The methods included light- and scanning electron microscopy (SEM), particle size analysis, differential scanning calorimetry, viscometry, thermal neutron flux measurements and energy deposition calculations. Seven hours-neutron irradiation results in sufficient specific activity of the 166Ho-PLLA-MS while structural integrity is preserved. Neutron flux measurements and energy deposition calculations are required in the screening of other nuclear reactors. For the evaluation of microsphere quality, light microscopy, SEM and particle size analysis are appropriate techniques.

Production of GMP-grade radioactive holmium loaded poly(L-lactic acid) microspheres for clinical application.

Radioactive holmium-166 loaded poly(L-lactic acid) microspheres are promising systems for the treatment of liver malignancies. The microspheres are loaded with holmium acetylacetonate (HoAcAc) and prepared by a solvent evaporation method. After preparation, the microspheres (Ho-PLLA-MS) are activated by neutron irradiation in a nuclear reactor. In this paper, the aspects of the production of a (relatively) large-scale GMP batch (4 g, suitable for treatment of 5-10 patients) of Ho-PLLA-MS are described. The critical steps of the Ho-PLLA-MS production process (sieving procedure, temperature control during evaporation and raw materials) were considered and the pharmaceutical quality of the microspheres was evaluated. The pharmaceutical characteristics (residual solvents, possible bacterial contaminations and endotoxins) of the produced Ho-PLLA-MS batches were in compliance with the requirements of the European Pharmacopoeia. Moreover, neutron irradiated Ho-PLLA-MS retained their morphological integrity and the holmium remained stably associated with the microspheres; it was observed that after 270h (10 times the half-life of Ho-166) only 0.3+/-0.1% of the loading was released from the microspheres in an aqueous solution. In conclusion, Ho-PLLA-MS which are produced as described in this paper, can be clinically applied, with respect to their pharmaceutical quality.

Albumin restores lysophosphatidylcholine-induced inhibition of vasodilation in rat aorta.

BACKGROUND: Impairment of vasodilation by oxidized low-density lipoprotein has been attributed to lysophosphatidylcholine (LPC). Albumin avidly binds LPC. Therefore, hypoalbuminemia may directly impair vasodilation and thus contribute to increased risk of atherosclerosis in nephrotic syndrome. The addition of albumin reduces LPC in erythrocytes and endothelial cells. We hypothesized that the addition of albumin will salvage vasodilation in aortic rings previously exposed to LPC. LPC increases superoxide production and disturbs L-arginine availability. Therefore, we also decreased superoxide with a superoxide dismutase mimic, MnCl(2), and supplemented L-arginine in an attempt to restore vasodilation. METHODS: Rat aorta rings, which had been incubated with various concentrations of LPC and human serum albumin (HSA), were mounted in organ chambers. Relaxation was studied with acetylcholine (0.01 to 100 micromol/L) after precontraction with phenylephrine (CON, 0.3 micromol/L; LPC, 0.03 micromol/L). In some studies MnCl(2) or L-arginine was added to the organ chamber. RESULTS: LPC had time- and dose-dependent inhibitory effects on acetylcholine-mediated vasodilation, but no effect on nitroprusside-mediated vasodilation. Preincubation with albumin (50 or 6 g/L) could protect vasodilation against very high levels of LPC. After preincubation with LPC, the addition of albumin to the incubation salvaged vasodilation. Albumin was more effective after short LPC incubation. MnCl(2) had no specific effect on the LPC-mediated disturbance in vasodilation. L-arginine completely salvaged vasodilation at low concentrations of LPC. However, even high concentrations of L-arginine (1 mmol/L) could not improve vasodilation at LPC levels at which vasodilation was restored by albumin. CONCLUSIONS: LPC affects several pathways that inhibit vasodilation, all of which are salvaged by addition of albumin.

Placental superoxide is increased in pre-eclampsia.

One of the current hypotheses on the pathophysiology of pre-eclampsia (PE) states that the placenta secretes one or more cytotoxic factors resulting in maternal endothelial dysfunction. Among the candidate factors are the products of increased oxidative stress. Although there is circumstantial evidence of such an increase, direct evidence is still lacking. Electron paramagnetic spin trap resonance (EPR), the most direct method to detect free radicals in tissues, was used to measure superoxide levels in placentae from normal pregnancies (n=13) and pregnancies complicated by PE (n=10). The superoxide level was significantly increased in the placental tissue of pre-eclamptic women. Moreover, upon inhibition of Cu-Zn superoxide dismutase (SOD) activity the relative increase of the superoxide levels was significantly smaller in the placentae from the PE patients, implying decreased basal Cu-Zn SOD activity. These findings lend direct support to the hypothesis that oxidative stress in placental tissue is increased in PE.

Donor-specific antibodies after transplantation by flow cytometry: relative change in fluorescence ratio most sensitive risk factor for graft survival.

BACKGROUND: There is no consensus on the role of donor-directed antibodies after renal transplantation detected by complement-dependent cytotoxicity (CDC) or by flow cytometry (FC). METHODS: Therefore, antibody formation was studied by FC and correlated with clinical course in a group of patients who received transplants between 1983 and 1993. All had a negative current CDC crossmatch and were treated with cyclosporine. Current and posttransplant sera from 143 donor-recipient combinations were studied retrospectively. Antibodies were considered present in FC if the fluorescence ratio between serum and negative control was > 2.65. RESULTS: Of 143 patients, 17 (11.9%) were found to be positive in the posttransplant FC crossmatch and 126 (88.1%) were negative. Of the positive patients, 3 were already positive in the current FC crossmatch, whereas 14 demonstrated a positive posttransplant FC crossmatch after a negative current FC crossmatch. It was noteworthy that, from 16 patients with a positive current FC crossmatch, 13 turned negative in the posttransplant crossmatch. In 113 recipients (79%), both pre- and posttransplant FC crossmatches were negative. The development of a positive FC crossmatch after transplantation was a significant risk factor for graft survival in Cox regression analysis (P = 0.01). The results were also studied as relative change in fluorescence ratio (RCFR). RCFR was determined by classifying the recipients in quartiles according to their change in flow cytometric value from current to posttransplant serum. Quartiles were defined as follows: quartile 1, decrease > 10%; quartile 2, decrease 0-10%; quartile 3, increase > 0-30%; and quartile 4, increase > 30%. RCFR proved to be the only significant risk factor for graft survival (odds ratio for quartile 4 vs. quartile 1, 3.27; P < 0.02). More rejections were shown for increasing quartile numbers (P < 0.001). CONCLUSIONS: Classification of patients by RCFR detected more patients with unfavorable clinical outcome (25% vs. 11%) than by FC crossmatch.

Effects of low-dose angiotensin II infusion on loop segment reabsorption: a free-flow micropuncture study in rats.

1. Little direct information is available on the actions of angiotensin II beyond the proximal tubule. We therefore studied the effect of a mildly vasoconstrictive dose of angiotensin II on tubular handling of water, sodium (Na+) and lithium (Li+) in rats by means of free-flow micropuncture at the late proximal tubule and the early distal tubule. 2. Endogenous angiotensin II was suppressed by pretreatment with enalapril. Compared with a control group, angiotensin II increased mean arterial pressure by 15 mmHg. Glomerular filtration rate decreased from 1.32 +/- 0.05 to 1.10 +/- 0.05 ml/min, Na+ excretion from 0.43 +/- 0.09 to 0.13 +/- 0.03 mumol/min, fractional delivery of water at the late proximal tubule from 50.1 +/- 1.7 to 42.9 +/- 3.2%, fractional delivery of Na+ at the late proximal tubule from 46.5 +/- 1.3 to 39.1 +/- 3.5% and fractional delivery of water at the early distal tubule from 26.4 +/- 1.4 to 21.9 +/- 1.0% (P < 0.05 for each variable). Fractional delivery of Na+ at the early distal tubule did not change significantly. 3. Similar experiments were performed during partial aortic constriction to exclude the effects of increased perfusion pressure. The data obtained were similar, except that in this group the fractional delivery of Na+ at the early distal tubule decreased from 8.6 +/- 0.7 to 6.8 +/- 0.9% (P < 0.05). 4. The relation between late proximal tubule Na+ delivery and Na+ reabsorption between late proximal and early distal tubule was not disturbed by angiotensin II. For water, however, this relation tended to shift to a higher reabsorption rate.(ABSTRACT TRUNCATED AT 250 WORDS)

Prostaglandin synthesis inhibition stimulates lithium reabsorption in Henle's loop in rats.

The effect of prostaglandin (PG) synthesis inhibition on tubular lithium (Li) handling in the rat was studied by micropuncture at the late proximal (LPT) and early distal (EDT) tubules. Animals received no treatment (N = 7), meclofenamate (MECLO; 5 mg/kg i.p. and 5 mg/kg i.v.; N = 7) or indomethacin (INDO, 1 mg/kg i.v.; N = 6). Whole kidney fractional Li excretion fell from 26.2 +/- 1.5% in control rats to 12.8 +/- 1.3% and 14.6 +/- 1.3% in MECLO and INDO treated rats. Fractional delivery of Li to the LPT was not affected by PG synthesis inhibition. All of the rise in tubular Li reabsorption took place between the LPT and EDT; at the EDT fractional Li delivery fell from 32.3 +/- 2.0% in the control group to 19.1 +/- 1.6% and 20.0 +/- 1.4% in the rats given MECLO or INDO. Water reabsorption between the LPT and EDT also increased. The tubular fluid/plasma Li concentration ratio ([T/P]Li) at the LPT was approximately 1.15 in all groups of rats. At the EDT, this ratio was approximately 1.06 in control rats, but only approximately 0.82 (P < 0.01) in rats subjected to PG synthesis inhibition. The finding that the [T/P]Li fell from values exceeding unity at the LPT to values below unity at the EDT during PG synthesis inhibition indicates that reabsorption of Li between these sites was stimulated to a greater extent than that of water. This dissociation strongly suggests that at least part of the increased Li reabsorption between the LPT and EDT took place in the water-impermeable thick ascending limb.(ABSTRACT TRUNCATED AT 250 WORDS)