B-cell immunity in the context of T-cell tolerance after combined kidney and bone marrow transplantation in humans.

Five patients with end-stage kidney disease received combined kidney and bone marrow transplants from HLA haploidentical donors following nonmyeloablative conditioning to induce renal allograft tolerance. Immunosuppressive therapy was successfully discontinued in four patients with subsequent follow-up of 3 to more than 6 years. This allograft acceptance was accompanied by specific T-cell unresponsiveness to donor antigens. However, two of these four patients showed evidence of de novo antibodies reactive to donor antigens between 1 and 2 years posttransplant. These humoral responses were characterized by the presence of donor HLA-specific antibodies in the serum with or without the deposition of the complement molecule C4d in the graft. Immunofluorescence staining, ELISA assays and antibody profiling using protein microarrays demonstrated the co-development of auto- and alloantibodies in these two patients. These responses were preceded by elevated serum BAFF levels and coincided with B-cell reconstitution as revealed by a high frequency of transitional B cells in the periphery. To date, these B cell responses have not been associated with evidence of humoral rejection and their clinical significance is still unclear. Overall, our findings showed the development of B-cell allo- and autoimmunity in patients with T-cell tolerance to the donor graft.

Evaluation of the pharmacodynamic and pharmacokinetic interaction between calcium antagonists and digoxin.

Therapeutic uses of calcium antagonists have expanded to include not only ischemic heart disease but arrhythmias, systemic hypertension, congestive heart failure, and various pulmonary and gastrointestinal diseases. Many patients receiving a calcium antagonist concomitantly receive digoxin. Although the potential interaction between these agents has frequently been investigated, literature reports are confusing and inconsistent. We summarized the pharmacodynamics, pharmacokinetics, and mechanisms of interaction to help clinicians evaluate the potential calcium antagonist-digoxin interaction.

Pyramidal tract of the cat: axon size and morphology.

The purpose of this work was to determine the number and morphology of pyramidal tract (PT) axons in the cat, using electron microscopy, modern methods of fixation, and computer-assisted morphometric analysis. Sections taken at the level of the medullary pyramids in three animals were fixed and magnified up to 10,000 X to produce photomicrographs. Morphological data were entered into computer files for analysis by tracing axon perimeters on micrographs mounted on a digitizer tablet. The number of axons per PT averaged 415,000, of which 88% were myelinated and 12% were unmyelinated. 90% of the myelinated axons fell in the diameter range 0.5-4.5 microns. Axons larger than 9 microns diameter accounted for 1% of the total; the largest were 20-23 microns. Myelinated axon mean diameter was 1.98 microns; because of the skewed distribution, with many small axons and a few very large axons, median diameter was 1.60 micron. Size distribution was relatively uniform throughout the PT cross section, with all sizes represented in all regions. However, the more medial regions had a higher proportion of small fibers than the more lateral regions: mean medial diameter was 1.85 micron while mean lateral diameter was 2.09 microns. Myelin sheath thickness averaged 7.9% of fiber diameter for axons up to 11 microns, but was constant at 0.9 micron for larger fibers. Myelinated fibers were distorted from the circular shape in cross section, with a mean circularity index (or form factor) of 0.85, which implies that the fibers could swell about 15% without rupture of the cell membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of food on the absorption of methotrexate sodium tablets in healthy volunteers.

OBJECTIVE: To examine the effect of food on the absorption and bioavailability of low-dose orally administered methotrexate sodium tablets. METHODS: In this randomized, 2-way crossover study, a 7.5-mg dose of methotrexate (three 2.5-mg tablets) was administered to 12 healthy male volunteers after an overnight fast or within 10 minutes of consuming a high fat-content breakfast. Serum methotrexate concentrations over the next 24 hours were used to determine the area under the concentration-time curve (AUC), the maximum concentration, the time to maximum concentration (tmax), and the serum half-life for each phase. RESULTS: Food delayed the tmax by approximately 30 minutes, but the extent of absorption, as measured by the AUC, for both phases was similar. CONCLUSION: These results demonstrate that the bioavailability of low-dose orally administered methotrexate sodium tablets is not influenced by food.

Bioequivalence of two leucovorin calcium tablet formulations.

In a randomized, single-dose, two-way crossover study, 36 male volunteers received 25 mg each of two oral formulations of leucovorin calcium. Reduced serum folate concentrations were determined over the 24 hours after dosing. There were no statistically significant differences in areas under the serum concentration-time curves for total L-tetrahydrofolates, L-leucovorin (L-5-formyltetrahydrofolate), and L-5-methyltetrahydrofolate (the active metabolite of leucovorin and the predominant circulating form of reduced folate after oral administration). The peak serum concentrations and times to peak serum concentrations were also not significantly different. We conclude that the two leucovorin calcium 5 mg tablet formulations are bioequivalent.

Liquid-chromatographic determination of ciprofloxacin in serum and urine.

We describe the liquid-chromatographic determination of ciprofloxacin in patients' serum and urine. Serum samples were prepared by precipitating protein with perchloric acid. Urine samples were diluted 100-fold with mobile phase. The mobile phase, consisting of pH 3 phosphate buffer/acetonitrile/methanol (81/5/14, by vol), was pumped through a mu Bondapak C18 reversed-phase column at 1.5 mL/min. Fluorescence of the effluent was monitored, at wavelengths for excitation and emission of 270 and 440 nm, respectively. Standard curves were linearly related to concentration from 0.08 to 10 mg/L for serum, 1 to 20 mg/L for urine. The procedure was evaluated in a clinical setting to determine its usefulness in studying the pharmacokinetics of ciprofloxacin in patients with concurrent diseases and receiving multiple drug therapies.

Estimation of the steady-state volume of distribution for digoxin: a comparison of model-independent methods with a two-compartment model in healthy volunteers.

Digoxin serum concentration vs. time data have been described in the literature by a linear two-compartment model. When calculating the steady-state volume of distribution for digoxin after oral dosing, a computer fitting program is often used because of the complex first-order absorption, two-compartment model employed. Since computer programs are not always available, we computed and compared the steady-state volume of distribution/bioavailability for digoxin using both a model-independent (area) and compartmental approach. Six healthy subjects participated in the study; each received digoxin 0.2 mg in capsule form daily for ten days. The mean steady-state volume of distribution/bioavailability calculated by noncompartmental analysis was 785 L and the mean for compartmental analysis was 784 L. The small difference between methods suggests that area analysis offers a simpler alternative to computerized compartmental fitting to determine this parameter for digoxin.

Bioequivalence of verapamil hydrochloride extended-release pellet-filled capsules when opened and sprinkled on food and when swallowed intact.

A study was performed to determine whether verapamil hydrochloride administered in extended-release pellet-filled capsules is bioequivalent to the same formulation administered by sprinkling the contents of the capsules onto food. Thirty-two healthy subjects participated in the randomized, two-way crossover study. In treatment A, the subjects swallowed the contents of a verapamil hydrochloride extended-release pellet-filled capsule, 240 mg, that had been sprinkled on applesauce. In treatment B, the subjects swallowed the same type of capsule intact. Blood samples were drawn at baseline, every hour for 10 hours, and at 12, 15, 24, 30, 36, and 48 hours after each dose administration. The plasma was analyzed for verapamil and norverapamil by high-performance liquid chromatography. The following calculations were performed: AUC0-48, AUC0-infinity, Cmax, tmax, and k. Results for the two treatments were compared by analysis of variance. There were no significant differences between the AUC0-48, AUC0-infinity, Cmax, tmax, and k for the two methods of dose administration. For verapamil the differences for all variables were less than 5%, and for norverapamil the differences were less than 4% for all variables except tmax (9.5%). The 90% confidence intervals were within acceptable limits for all variables except the norverapamil tmax comparison. Sprinkling the contents of extended-release pellet-filled capsules onto food provides verapamil hydrochloride that is bioequivalent to that obtained from the intact capsules.

Role of Borrelia burgdorferi in the pathogenesis of morphea/scleroderma and lichen sclerosus et atrophicus: a PCR study of thirty-five cases.

Morphea (localized scleroderma), and lichen sclerosus et atrophicus (LSA) share common features with acrodermatitis chronica atrophicans (ACA), a known chronic form of borreliosis. These include similar histologic findings such as diffuse dermal fibrosis. These observations have led several investigators to consider the possibility of Borrelia burgdorferi (Bb) as a common etiologic factor among all of these diseases. The aim of this study is to investigate the role of Bb in the pathogenesis of morphea and LSA, by assaying for its presence in lesional skin biopsies from patients with these diseases. We utilized the nested polymerase chain reaction (PCR) technique to selectively amplify a longer segment of a Bb-specific somatic gene, on DNA from paraffin-embedded, formalin-fixed tissues. The results revealed no Bb-specific DNA sequence in 28 specimens of morphea/scleroderma and 7 of LSA with varying stages of disease. Furthermore, confirmatory Southern blot of the PCR product, resulted in similar findings. These data seriously question the role played by this spirochete in the pathogenesis of morphea and LSA, at least in the southeastern part of the USA.