Localization of cathepsin D in rat liver. Immunocytochemical study using post-embedding immunoenzyme and protein A-gold techniques

Light and electron microscopic localization of cathepsin D in rat liver was investigated by post-embedding immunoenzyme and protein A-gold techniques. By light microscopy, cytoplasmic granules of parenchymal cells and Kupffer cells were stained for cathepsin D. Weak staining was also noted in sinusoidal endothelial cells. In the parenchymal cells many of positive granules located around bile canaliculi. In the Kupffer cells and the endothelial cells, diffuse staining was noted in the cytoplasm in addition to granular staining. By electron microscopy, gold particles representing the antigenic sites for cathepsin D were seen in typical secondary lysosomes and some multivesicular bodies of the parenchymal cells and Kupffer cells. The lysosomes of the endothelial cells and fat-storing cells were weakly labeled. Quantitative analysis of the labeling density in the lysosomes of these three types of cells demonstrated that the lysosomes of parenchymal cells and Kupffer cells are main containers of cathepsin D in rat liver. The results suggest that cathepsin D functions in the intracellular digestive system of parenchymal cells and Kupffer cells but not so much in that of the endothelial cells.     

Alterations in cerebrospinal fluid uridine,hypoxanthine, and xanthine in head-injured patients

1. Examination of the cerebrospinal fluid (CSF) of head-injured patients reveals that the concentration of intraventricular xanthine is elevated and that of uridine is decreased relative to those of adult lumbar CSF. 2. No correlations were observed between CSF lactate and CSF hypoxanthine, xanthine, or uridine, suggesting that changes in purine metabolites and the pyrimidine nucleoside do not index similar cellular events as does lactic acid production. 3. Ventricular CSF from hydrocephalic infants had uridine and hypoxanthine concentrations not significantly different from those of normal adult lumbar CSF, but xanthine was significantly elevated. 4. Since uridine has anticonvulsant properties and is a crucial substrate for cerebral metabolism, it may be useful to evaluate this pyrimidine for use in the management of patients with head injury.     

Microflora of the upper respiratory tract in young children under normal conditions and in respiratory tract diseases. Microbial count of the upper respiratory tract in healthy children and in children with pneumonia

The results of the inoculation of material taken from the anterior section of the nasal cavity and from the pharyngeal mucosa of 50 healthy young children and 298 acute pneumonia patients were analyzed. 23 microbial species were isolated. In the samples taken from the anterior section of the nasal cavity, monocultures were detected in 86 samples and 54 variants of associations including 2-4 species, in 139 samples. In the samples taken from the pharynx, monocultures were detected in 59 samples and 180 variants of associations including 2-6 species, in 282 samples. Differences in the contamination of the nasal cavity and the pharynx in healthy children and in pneumonia patients were revealed. These differences were manifested in the structure of the microflora (monocultures, associations, their composition), the assortment of microbial species and their concentration. In young children with pneumonia the microflora of the upper respiratory tract was found to reflect the severity of acute pneumonia and the intensity of the pathological process in the lungs (uncomplicated, pyodestructive pneumonia, pyodestructive pneumonia with fatal termination, acute purulent pleurisy).     

Transcriptional control, translation and function of the products of the five open reading frames of the Escherichia coli nir operon

Five open reading frames designated nirB, nirD, nirE, nirC and cysG have been identified from the DNA sequence of the Escherichia coli nir operon. Complementation experiments established that the NirB, NirD and CysG polypeptides are essential and sufficient for NADH-dependent nitrite reductase activity (EC 1.6.6.4). A series of plasmids has been constructed in which each of the open reading frames has been fused in-phase with the beta-galactosidase gene, lacZ. Rates of beta-galactosidase synthesis during growth in different media revealed that nirB, -D, -E and -C are transcribed from the FNR-dependent promoter, p-nirB, located just upstream of the nirB gene: expression is co-ordinately repressed by oxygen and induced during anaerobic growth. Although the nirB, -D and -C open reading frames are translated into protein, no translation of nirE mRNA was detected. The cysG gene product is expressed from both p-nirB and a second, FNR-independent promoter, p-cysG, located within the nirC gene. No NADH-dependent nitrite reductase activity was detected in extracts from bacteria lacking either NirB or NirD, but a mixture of the two was as active as an extract from wild-type bacteria. Reconstitution of enzyme activity in vitro required stoichiometric quantities of NirB and NirD and was rapid and independent of the temperature during mixing. NirD remained associated with NirB during the initial stages of purification of the active enzyme, suggesting that NirD is a second structural subunit of the enzyme.     

Purification of smooth-muscle myosin free of calmodulin and myosin light-chain kinase. Susceptibility to oxidation.

Smooth-muscle myosin purified as described by Persechini & Hartshorne [(1983) Biochemistry 22, 470-476] contains trace amounts of calmodulin and myosin light-chain kinase, which can be removed by Ca2+-dependent hydrophobic-interaction chromatography followed by calmodulin-Sepharose affinity chromatography. The resultant column-purified myosin exhibits properties similar to those of the non-purified myosin, e.g. actin activation of the Mg2+-ATPase requires Ca2+/calmodulin-dependent phosphorylation of the two 20 kDa light chains. However, unlike the non-purified myosin, the column-purified myosin undergoes a time-dependent transition to a form which no longer requires phosphorylation for actin activation of the myosin Mg2+-ATPase. This transition is identified as a time-dependent change in conformation of the column-purified myosin from a 10 S to 6 S form and is caused by slow oxidation of the column-purified myosin, since it could be prevented by storage under N2 and reversed by 5 mM-dithiothreitol.     

Capillary growth and diffusion distances in muscle

1. Tissue capillarity in muscle was modelled as square-ordered arrays with capillary-to-fiber ratios (C/F) from 0.5 to 'infinity'. 2. C/F up to two had marked effects on diffusion distances, but C/F above had only slight effects on average distances and almost no effect on maximal distances. 3. Capillary growth during normal maturation results in C/F around two. Thus, capillary growth in adult muscle may not be an adaptive mechanism for reducing diffusion distances.     

Tunicamycin-resistant mutants of Bacillus amyloliquefaciens are deficient in amylase, protease and penicillinase synthesis and have altered sensitivity to antibiotics and autolysis

Mutants of Bacillus amyloliquefaciens resistant to at least 10 micrograms/ml of tunicamycin were isolated and shown to be pleiotropic. The mutants were more resistant to streptomycin, chloramphenicol, kanamycin and neomycin than was the parent strain but less resistant to penicillin G and tetracycline. They were more autolytic, presumably due to an altered cell wall. The mutants produced reduced levels of amylase, penicillinase and both metal and serine protease besides having an enhanced sporulation frequency and being more motile.     

Estimate of mean tissue O2 consumption at onset of exercise in males

A mathematical model has been developed that permitted the calculation of the flow-weighted mean tissue O2 consumption (VO2T) at the onset of a step increase in work rate. From breath-by-breath measurements of alveolar O2 consumption (VO2A) and cardiac output (Q) by impedance cardiography and assumptions about the site of depletion of O2 stores, the rate of change in O2 stores (VO2s) was determined. The sum of VO2A + VO2s = VO2T. Six very fit males performed six repetitions of each of two step increases in work rate. STlo was a transition from rest to 100-W cycling; SThi was a transition from 100- to 200-W cycling. For each work rate transition, the responses of VO2A and Q were averaged over the six repetitions of each subject and the model was solved to yield VO2T. The responses of VO2A, VO2T, and Q after the increase in work rate were fit with a monoexponential function. This function included a time constant and time delay, the sum of which gave the mean response time (MRT). In the STlo test, the MRT of VO2A (24.9 +/- 1.1 s, mean +/- SE) was longer than that of VO2T (15.3 +/- 1.3 s) and of Q (16.5 +/- 6.5 s) (P less than 0.05). The MRT of VO2T and Q did not differ significantly. Also for SThi, the MRT of VO2A (34.4 +/- 3.3 s) was significantly longer than that of VO2T (30.0 +/- 3.4 s) (P less than 0.05). The MRT of VO2T and Q (30.3 +/- 5.5 s) were not significantly different at this work rate either.(ABSTRACT TRUNCATED AT 250 WORDS)