Comparison of glycoconjugates at the surface of developing type II pneumocytes and Clara cells

Summary The apical surface coat of type II pneumocytes and Clara cells in pre- and post-natal rat lung was examined with lectin histochemical methods. Lectins fromHelix pomatia (HPA), peanut (PNA) andMaclura pomifera (MPA) were conjugated with horseradish peroxidase and used to stain paraffin sections of fixed lung with or without certain pre-treatments. HPA and MPA were observed to react with almost all type II pneumocytes at postnatal day 1. Type II pneumocytes that stained with a sialidase—PNA sequence increased from a few positive cells at postnatal day 5 to many in the adult. It has been reported that the surface coat of type II pneumocytes closely resembles that of Clara cells in its staining with histochemical methods employing cationic dyes or lectins including MPA and PNA. However, staining with HPA, especially after periodic acid oxidation, revealed many type II pneumocytes with strong reactivity but showed only a few Clara cells that were faintly positive. HPA also stained alveolar macrophages. The HPA affinity of macrophages, however, was labile to oxidation with periodic acid or galactose oxidase unlike that of type II pneumocytes. This difference suggests that HPA recognizes more than one type of sugar structure.To whom all correspondence and reprint requests should be addressed.     

Can we determine the biological availability of sediment-bound trace elements?

It is clear from available data that the susceptibility of biological communities to trace element contamination differs among aquatic environments. One important reason is that the bioavailability of metals in sediments appears to be altered by variations in sediment geochemistry. However, methods for explaining or predicting the effect of sediment geochemistry upon metal bioavailability are poorly developed. Experimental studies demonstrate that ingestion of sediments and uptake from solution may both be important pathways of metal bioaccumulation in deposit/detritus feeding species. Relative importance between the two is geochemistry dependent. Geochemical characteristics of sediments also affect metal concentrations in the tissues of organisms collected from nature, but the specific mechanisms by which these characteristics influence metal bioavailability have not been rigorously demonstrated. Several prerequisites are necessary to better understand the processes that control metal bioavailability from sediments. 1) improved computational or analytical methods for analyzing distribution of metals among components of the sediments; 2) improved computational methods for assessing the influences of metal form in sediments on sediment-water metal exchange; and 3) a better understanding of the processes controlling bioaccumulation of metals from solution and food by metazoan species directly exposed to the sediments. Such capabilities would allow mechanistic explanations essential to the development of practical tools sought for determining sediment quality criteria for metals.     

Association of catalytic and regulatory subunits of cyclic AMP-dependent protein kinase requires a negatively charged side group at a conserved threonine.

In Saccharomyces cerevisiae, as in higher eucaryotes, cyclic AMP (cAMP)-dependent protein kinase is a tetramer composed of two catalytic (C) subunits and two regulatory (R) subunits. In the absence of cAMP, the phosphotransferase activity of the C subunit is inhibited by the tight association with R. Mutation of Thr-241 to Ala in the C1 subunit of S. cerevisiae reduces the affinity of this subunit for the R subunit approximately 30-fold and results in a monomeric cAMP-independent C subunit. The analogous residue in the mammalian C subunit is known to be phosphorylated. Peptide maps of in vivo 32P-labeled wild-type C1 and mutant C1(Ala241) suggest that Thr-241 is phosphorylated in yeast cells. Substituting Thr-241 with either aspartate or glutamate partially restored affinity for the R subunit. Uncharged and positively charged residues substituted at this site resulted in C subunits that failed to associate with the R subunit. Replacement with the phosphorylatable residue serine resulted in a C subunit with wild-type affinity for the R subunit. Analysis of this protein revealed that it appears to be phosphorylated on Ser-241 in vivo. These data suggest that the interaction between R and C involves a negatively charged phosphothreonine at position 241 of yeast C1, which can be mimicked by either aspartate, glutamate, or phosphoserine.     

Quantitative genetics of seed size variation inPhlox

Summary Because seed size is often associated with survival and reproduction in plant populations, genetic variation for seed size may be reduced or eliminated by natural selection. To test this hypothesis we assessed genetic sources of variation in seed size in a population ofPhlox drummondii to determine whether genetic differences among seeds influence the size they attain. A diallel cross among 12 plants from a population at Bastrop, Texas, USA allowed us to partition variance in the mass of seeds among several genetic and parental effects. We found no evidence of additive genetic variance or dominance genetic variance for seed mass in the contribution of plants to their offspring. Extranuclear maternal effects accounted for 56% of the variance in seed mass. A small interaction was observed between seed genotype and maternal plant. Results of this study support theory that predicts little genetic variation for traits associated with fitness.     

New antifungal agents for the systemic mycoses

The azoles are the prominent broad spectrum oral antifungal agents in use or under clinical investigation for the systemic mycoses. This class of antifungal agents is represented by the marketed drug ketoconazole (Nizoral) and the experimental triazoles furthest along in clinical trials in the United States, itraconazole and fluconazole. Ketoconazole use is limited by its side effect profile and activity spectrum. Itraconazole appears to be better tolerated and less toxic to liver function, does not cause adrenal suppression and is more active against Aspergillus and Sporothrix schenckii. Fluconazole appears to be a highly promising agent due its highly favorable pharmacokinetic profile; it is water soluble, is well tolerated, is not metabolized to inactive constituents, it has a long half-life and, unlike the other azoles, high cerebrospinal fluid levels are readily attained for consideration in meningeal mycoses. It remains to be determined what place these new triazoles have in managing immunosuppressed patients including those with acquired immune deficiency syndrome known as AIDS. Other experimental antifungal agents, including ambruticin, amphotericin B methyl ester and saramycetin are also described. Sales figures are presented of drugs marketed in the United States for the systemic mycoses and reflect the growing problem of fungal diseases in the population.Presented as part of the Everett S. Beneke Symposium in Mycology, May 27, 1988.     

Creatine kinase activity of urinary bladder and skeletal muscle from control and streptozotocin-diabetic rats

The urinary bladder depends on intracellular ATP for the support of a number of essential intracellular processes including contraction. The concentration of ATP is maintained constant primarily via the rapid transfer of a phosphate from creatine phosphate (CP) to ADP catalyzed by the enzyme creatine kinase (CK). Since muscular pathologies associated with diabetes are in part related to intracellular alterations in metabolism, we have characterized the CK activity in both skeletal muscle and urinary bladder from control and streptozotocin-diabetic rats.The following is a summary of the results: 1) Bladder tissue from control rats showed linear kinetics with a V_max = 390 nmoles/mg protein/min, and a K_m = 275 µM. 2) Urinary bladder tissue isolated from diabetic rats displayed biphasic kinetics with V_max = 65 and 324 nmoles/mg protein/min, and K_m's = 10 µM and 190 µM respectively. 3) Skeletal muscle isolated from control rats showed linear kinetics with an approximate V_max of 800 nmoles/mg protein/min and a K_m of 280 µM CP. 4) Homogenates of skeletal muscle from diabetic rats showed complex kinetics not separable into distict component forms. 5) The K_m for ADP for both skeletal muscle and bladder was approximately 10 µM.These studies demonstrate that whereas bladders isolated from both control and diabetic rats possess a low-affinity isomer(s) of CK with similar maximum enzymatic activity, there is a high affinity isomer present within the urinary bladder muscle of diabetic rats that is not present in bladder tissue isolated from control rats. Skeletal muscle isolated from both diabetic and control rats exhibited a maximal activity 2 to 3 times higher than that of the bladder.     

Analysis of class II (hydrolytic) and class I (beta-lyase) apurinic/apyrimidinic endonucleases with a synthetic DNA substrate.

We have developed simple and sensitive assays that distinguish the main classes of apurinic/apyrimidinic (AP) endonucleases: Class I enzymes that cleave on the 3' side of AP sites by beta-elimination, and Class II enzymes that cleave by hydrolysis on the 5' side. The distinction of the two types depends on the use of a synthetic DNA polymer that contains AP sites with 5'-[32P]phosphate residues. Using this approach, we now show directly that Escherichia coli endonuclease IV and human AP endonuclease are Class II enzymes, as inferred previously on the basis of indirect assays. The assay method does not exhibit significant interference by nonspecific nucleases or primary amines, which allows the ready determination of different AP endonuclease activities in crude cell extracts. In this way, we show that virtually all of the Class II AP endonuclease activity in E. coli can be accounted for by two enzymes: exonuclease III and endonuclease IV. In the yeast Saccharomyces cerevisiae, the Class II AP endonuclease activity is totally dependent on a single enzyme, the Apn1 protein, but there are probably multiple Class I enzymes. The versatility and ease of our approach should be useful for characterizing this important class of DNA repair enzymes in diverse systems.     

Training for general practice: a national survey.

OBJECTIVES--(a) To compare current vocational training in general practice with that ascertained by a survey in 1980; (b) to compare the training of trainees in formal training schemes with that of trainees arranging their own hospital and general practice posts. DESIGN--National questionnaire survey of United Kingdom and armed services trainees who were in a training practice on 1 April 1989. Questionnaires were distributed by course organisers. SETTING--Research project set up after an ad hoc meeting of trainees at the 1988 national trainee conference. SUBJECTS--2132 Of the 2281 trainees (93%) known to be in a training practice on 1 April 1989. RESULTS--1657 Trainees returned the questionnaires, representing 73% of all trainees known to be in a training practice on 1 April 1989. Between 1980 and 1989 there were significant improvements in the trainee year, and there was also evidence of improvements in general practice study release courses. There was no evidence of improvement in other aspects of training. General practice trainees spent an average of three years in junior hospital posts, which provided very little opportunity for study related to general practice. Training received during tenure of hospital posts differed significantly between trainees in formal schemes and those arranging their own hospital posts. During the trainee year training was almost the same for those in formal schemes and those arranging their own posts. Regions varied significantly in virtually all aspects of general practice training. CONCLUSIONS--The trainee year could be improved further by enforcing the guidelines of the Joint Committee on Postgraduate Training for General Practice. The poor training in junior hospital posts reflected the low priority that training is generally given during tenure of these posts. A higher proportion of general practice trainees should be attached to vocational training schemes. More hospital trainees could attend general practice study release courses if these were designed specifically with the needs of hospital doctors in mind.     

In vitro stimulation and inhibition of steroid hormone release from postovulatory follicles of the tilapia,Oreochromis mossambicus

Summary Postovulatory follicles of the tilapia, Oreochromis mossambicus, were incubated with graded doses of salmon gonadotropin to identify the steroid hormones released by this tissue. In addition, the effects of either cytochalasin B or colchicine on steroid hormone release were studied. After the incubation, the tissue was examined by electron microscopy. Postovulatory follicles released testosterone and estradiol-17B in a dose-dependent manner with gonadotropin. There was no detectable release of progesterone or 17a-OH-progesterone. When stimulated with high doses of gonadotropin, the steroidogenic cells showed an increase in smooth endoplasmic reticulum, Golgi complexes, and lipid droplets. Also, microfilaments became arranged in orderly bundles and were found close to the numerous secretory vesicles and lipid droplets. Upon incubation with gonadotropin and either colchicine or cytochalasin B, the cells still appeared steroidogenic, but the filaments were not organized nor associated with vesicles or lipid droplets. Release of steroid hormone decreased significantly. Also in these tissues, vesicles were no longer numerous in the apical region of the granulosa cells, but were located primarily near smooth endoplasmic reticulum and Golgi complexes. This suggests that disruption of the cytoskeleton results in reduced steroid hormone synthesis or release.