Peanut agglutinin in combination with CD19 monoclonal antibody has potential as a purging agent in myeloma.

Administration of high-dose chemotherapy to patients with myeloma, followed by rescue with autologous bone marrow transplantation (ABMT), sometimes induces complete disease remission but relapse is usual. We have attempted to reduce the risk of relapse by selective in vitro removal of myeloma cells from the autologous graft. A combination of the (gal-galNac)-binding lectin peanut agglutinin (PNA), which binds all plasma cells, and the pan-B monoclonal antibody CD19 was assessed for purging marrow of myeloma cells and their putative precursors using a magnetic bead method. Preliminary experiments performed on peripheral blood mononuclear cells spiked with fluorescent-labeled PNA+ Kirk tumor cells showed that a magnetic bead: target cell ratio of 40:1 resulted in a greater than 3-log reduction in PNA+ cells. This technique was then applied to 17 samples of myeloma bone marrow and to 18 samples of normal bone marrow spiked with PNA+ Kirk cells and CD19+ hairy cell leukemia cells. In each case all detectable plasma cells and CD19+ lymphocytes were effectively removed, and normal hemopoietic progenitor cell recovery was greater than 55%. This purging system deserves further study as a means of reducing relapse rates in myeloma patients treated by a combination of high-dose chemotherapy and ABMT.     

Cytoprotection by iloprost against paracetamol-induced toxicity in hamster isolated hepatocytes.

1 The ability of iloprost (ZK36374) to protect hamster isolated hepatocytes from the toxic effects of paracetamol and its reactive metabolite N-acetyl-p-benzoquinoneimine (NABQI) was investigated. The cytoprotection provided by iloprost was compared with that of N-acetyl-L-cysteine. 2 Treatment of hepatocytes with either NABQI (0.4 mM) or paracetamol (2 mM) alone resulted in a considerable loss of cell viability, as assessed by trypan blue exclusion or leakage of lactate dehydrogenase, accompanied by an increase in the percentage of viable cells that were blebbed. N-acetyl-L-cysteine (1.25 mM) pretreatment diminished the loss of cell viability and the percentage of blebbed cells resulting from exposure to NABQI or paracetamol, whereas iloprost (10(-16) M to 10(-10) M) pretreatment reduced only the loss of cell viability, not the percentage of viable cells exhibiting blebbing. Pretreatment with N-acetyl-L-cysteine significantly attenuated the depletion by paracetamol of glutathione and decreased the covalent binding of [14C]-paracetamol to cellular proteins, whereas iloprost was without any such effects. 3 The effects of iloprost and N-acetyl-L-cysteine were also investigated by use of a model of paracetamol toxicity in which it is possible to study the biochemical events leading to cell injury separate from the generation of toxic metabolites. Hamster hepatocytes were incubated with paracetamol (4 mM) for 90 min at 37 degrees C during which metabolism of paracetamol occurs with minimal loss of cell viability.(ABSTRACT TRUNCATED AT 250 WORDS)     

The zygomatico-temporal approach to the skull base: a critical review of 11 patients.

The zygomatico-temporal approach to the base of the skull is a relatively new but established surgical technique. The approach involves the removal of the zygomatic bone to provide access to the skull base, middle cranial fossa, parasellar region and interpeduncular cistern with minimal brain retraction. An excellent view of the bifurcation of the basilar artery and suprasellar region is provided. The outcome of 11 patients undergoing this procedure is reported with particular reference to the post-operative morbidity and the cosmetic result.     

Cultured human granulosa cells as a model for corpus luteum function: relative roles of gonadotrophin and low density lipoprotein studied under defined culture conditions.

In primates, corpus luteum development involves both gonadotrophin stimulation and exposure to low density lipoprotein (LDL) delivered through vascularization of the granulosa cell-derived layer. These regulatory influences were modelled in vitro using granulosa cells obtained during in-vitro fertilization (IVF) cycles controlled with gonadotrophin releasing hormone (GnRH) analogue, human menopausal gonadotrophin (HMG) and human chorionic gonadotrophin (HCG). Granulosa cells were cultured in defined medium on extracellular matrix. Without gonadotrophin or LDL in the medium, progesterone production declined progressively. With LDL alone, there was a short-lived elevation of progesterone output which subsequently declined. Culture with HCG alone resulted in a relatively unchanged rate of steroid production over 5 days despite morphological development. This contrasted with a marked and sustained increase in progesterone output over the same time when granulosa cells were cultured with combined HCG/LDL. Cultures were challenged with combined HCG/LDL on day 5. Where initial incubation included HCG, the challenge resulted in a recovery of progesterone output to values comparable to those of granulosa cells exposed to continuous HCG/LDL. Initial incubation without gonadotrophin led to a reduced response. Results suggest that LDL delivery to granulosa cells of the early corpus luteum causes a short-lived period of progesterone production. Sustained luteinization of granulosa cells and maintenance of gonadotrophin responsiveness requires continued exposure to gonadotrophin in the luteal phase.     

Freshly isolated hepatocytes as a model for studying the toxicity of paracetamol

The toxicity of paracetamol has been investigated in freshly isolated hamster hepatocytes. Two phases of toxicity have been identified. In phase 1, metabolic activation of paracetamol occurs with depletion of glutathione. In phase 2, there is progressive morphological damage, leading ultimately to cell death. This occurs even in the absence of further exposure to paracetamol. The thiol reductant, dithiothreitol, added at the start of phase 2, prevents and reverses the toxicological damage that would otherwise occur. Thus, it is most likely that paracetamol causes hepatotoxicity through oxidation of SH groups in key enzymes. N-Acetylcysteine, but not methionine, has an effect similar to that of dithiothreitol. This difference is probably due to oxidation of the enzymes involved in the conversion of methionine to cysteine, whereas N-acetylcysteine can still serve as a precursor of glutathione. The glutathione can act both by adduct formation with the metabolite of paracetamol and as a thiol reductant. Species differences in sensitivity to paracetamol toxicity were shown to be due to differences in the rate of oxidation of the drug to its toxic metabolite. Most people are relatively poor activators of paracetamol, but in few subjects the reaction proceeds quite rapidly, rendering such individuals more sensitive to the hepatotoxic effects of the drug.     

Molecular determinants of benzodiazepine receptor affinities and anticonvulsant activities

In vivo convulsant activities profiles and receptor binding studies together with the techniques of theoretical chemistry were used to characterize 15 compounds, from five different chemical families, known to bind to the BDZ receptor. The experimental goals of this study were to determine the affinity of these analogs for this receptor, the effect of gamma-aminobutyric acid on the affinity, and, in a self-consistent manner, the nature of the activity, agonist (anticonvulsant), antagonist, or inverse agonist (proconvulsant, convulsant), elicited by binding to this receptor. To these ends, in vivo studies were made to determine the proconvulsant, convulsant, and anticonvulsant activities and antagonism to anticonvulsant activities of the 15 analogs. Their receptor affinities at 25 degrees were also determined by competitive inhibition of [3H] flunitrazepam and [3H]Ro 15-1788 in the absence and presence of gamma-aminobutyric acid. The goal of the theoretical studies was to identify and calculate molecular properties that modulate these affinities and types of activities and from them to develop a model of receptor recognition and activation that could consistently explain observed behavior and predict new results. Thus, molecular orbital calculations were carried out for all analogs, using semiempirical quantum mechanical methods. In addition to the optimization of structures, a number of electronic properties, such as polarizations, partition coefficients, and proton and electron affinities were computed and examined for their ability to modulate relative affinities and modes of activation of the receptor. From these studies, a model for receptor recognition involving two anchoring hydrogen bond-acceptor sites and for activation involving interaction of the most lipophilic aromatic region of each compound with the receptor was developed, which could systematically account for the three different types of behavior, agonist, antagonist, and inverse agonist, observed for these analogs. Electronic rather than structural properties were found to be the principal modulator of both recognition and activation. A possible mechanism of agonist activation of the receptor involving electron transfer to the agonist, as well as a possible induced conformational change in the receptor, is also suggested by these results. Finally, by complementarity, some steric and electronic characteristics of the receptor binding site could be deduced.     

Technical Note: A comparison of a novel direct ophthalmoscope, the OptyseTM, to conventional direct ophthalmoscopes

Despite the current popularity of binocular indirect ophthalmoscopy, direct ophthalmoscopes are still commonly used by clinicians for fundus examination. They are considered to be expensive, however, and it has been suggested that this cost can prevent their use by healthcare professionals in developing countries. The Optyse Lens Free Ophthalmoscope is a novel direct ophthalmoscope, without a lens focus system, that allows for comparatively inexpensive manufacture and supply. We compared the clarity of view with the Optyse to that with standard direct ophthalmoscopes, over a sequential cohort of patients with a variety of refractive errors and ocular conditions. The grade of clarity of view with the Optyse Lens Free Ophthalmoscope was less than conventional ophthalmoscopes (Wilcoxon signed rank test, p < 0.0001). This grade of clarity of view was not associated with the ametropia of the ophthalmoscopic observation (Spearman r < or = 0.03, p > or = 0.28) but was with the presence of cataracts (chi2 test, p < 0.0001) with both the Optyse and the conventional ophthalmoscopes. Despite its limitations, the retinal view with Optyse was often within acceptable clinical limits suggesting that this relatively inexpensive ophthalmoscope may have a place when cost prohibits any other type of ophthalmoscope use.     

[3H]Ketanserin labels 5-HT2 receptors and α1-adrenoceptors in human and pig brain membranes

The binding characteristics of [3H]ketanserin (a reported selective radioligand for serotonin 5-HT2 receptors) and [125I]BE 2254 (which labels selectively alpha 1-adrenoceptors) were characterized in brain frontal cortex membranes of pig and man. Saturation experiments indicated that both radioligands label apparently a homogeneous class of binding sites in human and pig fontal cortex membranes. Competition experiments with [125I]BE 2254 using 17 agonists and antagonists showed monophasic and steep curves in human and pig frontal cortex membranes. The pharmacological profile of these sites is typical of alpha 1-adrenoceptors. In competition experiments with [3H]ketanserin, most of the tested compounds displayed shallow or biphasic curves. In particular, alpha 1-adrenoceptor-selective antagonists (prazosin, WB 4101, BE 2254...) displaced with nanomolar affinity about 15 and 40% of the specific [3H]ketanserin binding in human and pig frontal cortex membranes, respectively. The minor component of [3H]ketanserin binding correlated highly significantly with [125I]BE 2254 binding in both membrane preparations. The major component of [3H]ketanserin binding to pig and human frontal cortex membranes correlated significantly with [3H]ketanserin binding in rat brain cortex membranes (which is essentially to 5-HT2 receptors). The present data demonstrate that [3H]ketanserin in nanomolar concentrations binds significantly to alpha 1-adrenoceptors in human and pig frontal cortex membranes; this suggests a rather limited degree of selectivity of ketanserin for 5-HT2 receptors in pig and human tissues.     

De novo generation of histamine in sputum and the effect of antibiotics

We have performed experiments to test the hypothesis that bacteria may contribute to the presence of histamine in sputum. Sputum samples obtained from 7 patients with exacerbations of chronic bronchitis and 7 patients with cystic fibrosis were incubated at 37 degrees C for 72 hours. Serial sputum histamine estimations, performed by a recently-developed HPLC technique, showed large, progressive increases in both groups of samples. Both the pre-heating of samples at 100 degrees C prior to incubation and the addition of antibiotics to the incubates substantially reduced these increases. These findings strongly suggest that bacteria may contribute to sputum histamine in infective lung disease.