Further Assessment of Rosette Inhibition Test in Clinical Organ Transplantation

The rosette inhibition test was used in the clinical management of organ allografts to estimate the amount of immunosuppressive drugs necessary to prevent rejection. In patients surviving more than three months renal function appeared to be better than in a similar group of patients managed without the test. It is suggested that this was due to a reduction in the number of clinical or subclinical rejection episodes. On the other hand, the test indicates that in many cases the level of immunosuppression should be much higher, and if this advice is followed the patients become increasingly exposed to the risk of infection. In other words, those patients with good renal function remained well, whereas those who might otherwise have rejected their kidney and survived had in fact died of sepsis.     

Studies of Raman Spectra of Water Solutions of Adenosine Tri-, Di-, and Monophosphate and Some Related Compounds

Using a He-Ne CW laser source together with a digital photon counting system, we have obtained well resolved Raman spectra for adenosine mono-, di-, and triphosphate (AMP, ADP, ATP) in aqueous solution. Spectra of these compounds were studied as a function of pH from pH = 0.5 to 13.5 and between 550 and 1700 cm(-1). It was found possible to distinguish spectroscopically between the three phosphates over the pH range studied. A qualitative analysis of vibrational modes responsible for various spectral lines is given. Lines at about 960 and 1100 cm(-1) were found to be good indications of the degree of ionization of the terminal phosphate group.     

Plasma concentrations of pituitary hormones in 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated male rats

Experiments were conducted to test the hypothesis that acute TCDD toxicity is associated with pituitary hypofunction. Sexually mature male Sprague-Dawley rats were given graded doses of TCDD (0-100 micrograms/kg) and evaluated 7 days later. Despite pronounced hypophagia and body weight loss, plasma concentrations of growth hormone (GH), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) were not significantly affected by any dose of TCDD. Only prolactin (PRL) concentrations were reduced, while, as previously reported, thyroid-stimulating hormone concentrations were elevated. Also, plasma LH, PRL, and adrenocorticotropic hormone (ACTH) concentrations were not significantly affected 1, 2, 3, 4, 5, or 7 days after a single dose of TCDD (50 micrograms/kg). We conclude that (1) pituitary hypofunction is not a major cause of the initial stages of acute TCDD toxicity, (2) growth retardation in TCDD-treated rats is not the result of a deficiency of GH, (3) alterations in plasma corticosterone concentrations are due to altered responsiveness of the adrenal to ACTH stimulation rather than to changes in plasma ACTH concentrations, and (4) that impaired spermatogenesis is not associated with a decrease in plasma FSH concentrations. In addition, the lack of a consistent effect on plasma PRL concentrations suggests that alterations in plasma PRL concentrations do not play a critical role in the toxicity of TCDD. Finally, because TCDD treatment causes a serious androgenic deficiency without increasing the rates at which androgens are catabolized or excreted, the fact that plasma LH concentrations were unaffected indicates that TCDD treatment must reduce the responsiveness of the testis to LH stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Monoclonal antibody against a melanosomal protein in melanotic and amelanotic human melanoma cells

BALB/c mice were immunized with tyrosinase, partially purified in two stages from a human melanoma cell line. A hybridoma was obtained which produced monoclonal antibody (MoAb 1C11) reactive with 8/10 melanoma cell lines and 10/10 primary cultures of human melanocytes, neval cells, and melanomas. Immunoreactivity correlated to a certain extent with tyrosinase activity but not with melanin content. No crossreactivity was obtained with neuroblastoma, medulloblastoma, fibroblasts, keratinocytes, lymphoid cells, or murine melanomas. Purification of the antigen directly from cell lysates with a MoAb 1C11 CNBr-Sepharose affinity column gave a green-brown protein of 56 kDa with no detectable tyrosinase activity. This protein was therefore different from 60 kDa active tyrosinase, identified by enzyme activity and Western blotting with a MoAb derived previously (MoAb 5C12). Unlike 5C12, 1C11 reactivity was not destroyed by pretreatment of the antigen with periodate. Immunogold labelling showed that the 1C11-reactive antigen was associated with melanosomes, and there was close correlation between 5C12 and 1C11 reactivity in resistance to trypsin and in staining various melanocytic cell populations. MoAb 1C11 may therefore recognise a polypeptide epitope in a molecule closely linked to melanin biosynthesis.     

Critical micelle concentration and hemolytic activity--a correlation suggested by the marine sterol, halistanol trisulfate.

The marine natural product, halistanol trisulfate, has a relatively low critical micelle concentration of 0.001% m/v (14.5 microM) and strong hemolytic potency with an EC50 of 0.00046% m/v (6.67 microM). As expected of a detergent, it inhibits the growth of gram-positive but not gram-negative bacteria. The hemolytic activity of halistanol trisulfate and other detergents has been shown to correlate with critical micelle concentration. This correlation may have important implications in the mechanism of membranolytic bioactivity.     

Linkage of the acetylcholine transporter-vesamicol receptor to proteoglycan in synaptic vesicles.

The relationship of the acetylcholine transporter-vesamicol receptor (AcChT-VR) to proteoglycan in Torpedo electric organ synaptic vesicles was investigated. The cholate-solubilized VR was immunoprecipitated by a monoclonal antibody directed against the SV1 epitope located in the glycosaminoglycan portion of the proteoglycan. AcChT that was photoaffinity-labeled with a tritiated high-affinity analogue of AcCh [cyclohexylmethyl cis-N-(4-azidophenacyl)-N-methylisonipecotate] and then denatured in sodium dodecyl sulfate also immunoprecipitated. The labeled AcChT exhibited a M(r) range of 100,000-200,000. Proteoglycan did not engage in detectable nonspecific reversible aggregation that might mask the presence of another subunit during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In vesicles permeabilized with cholate, the enzymes keratanase and testicular hyaluronidase inactivated binding of vesamicol and destroyed the SV1 epitope without detectable proteolysis. Other glycosaminoglycan-degrading enzymes were without effect. The results demonstrate that the AcChT-VR and proteoglycan are very strongly linked and that glycosaminoglycan-like polysaccharide controls the conformation of the VR. The unexpected linkage to proteoglycan suggests that AcChT-VR in intact terminals might communicate with extracellular matrix and participate in stabilization and operation of the synapse.     

Anonymous nuclear DNA markers in the American oyster and their implications for the heterozygote deficiency phenomenon in marine bivalves

A puzzling population-genetic phenomenon widely reported in allozyme surveys of marine bivalves is the occurrence of heterozygote deficits relative to Hardy-Weinberg expectations. Possible explanations for this pattern are categorized with respect to whether the effects should be confined to protein-level assays or are genomically pervasive and expected to be registered in both protein- and DNA-level assays. Anonymous nuclear DNA markers from the American oyster were employed to reexamine the phenomenon. In assays based on the polymerase chain reaction (PCR), two DNA-level processes were encountered that can lead to artifactual genotypic scorings: (a) differential amplification of alleles at a target locus and (b) amplification from multiple paralogous loci. We describe symptoms of these complications and prescribe methods that should generally help to ameliorate them. When artifactual scorings at two anonymous DNA loci in the American oyster were corrected, Hardy-Weinberg deviations registered in preliminary population assays decreased to nonsignificant values. Implications of these findings for the heterozygote-deficit phenomenon in marine bivalves, and for the general development and use of PCR-based assays, are discussed.