We studied the effects of recombinant human macrophage colony-stimulating factor (M-CSF) on the leukemic blast progenitors from 10 acute myeloblastic leukemia patients. Recombinant human (rh)M-CSF stimulated leukemic blast progenitors in methylcellulose in four patients, but the colonies by rhM-CSF were smaller in size and number than those by rh-granulocyte-CSF or human bladder carcinoma cell line 5637 conditioned medium. rhM-CSF did not increase the number of clonogenic cells in long-term suspension culture. The blast colony formation in methylcellulose and the exponential growth of clonogenic cells in long-term suspension culture are considered to reflect the terminal divisions and the self-renewal of blast progenitors, respectively. The results show that M-CSF stimulates terminal divisions weakly but does not stimulate self-renewal of leukemic blast progenitors. M-CSF did not induce differentiation of blasts either in methylcellulose or in suspension culture.

Cell Differentiation↗

PubMed1988

[Associative visual agnosia--a case report].

A 61 year-old right-handed man, a president of a company fell downstairs and was found unconscious on January 1, 1985. He was admitted to a hospital and he had been unresponsive for 10 days. As he recovered from the consciousness disturbance, he complained of difficulties in discrimination of common objects and in recognition of familiar faces. The patient was admitted to our hospital for the closer examination on July 29, 1985. There was no abnormality in the general physical examination on admission. Neurological examinations revealed no significant deficits except visual field disturbance. In the results of WAIS, the verbal IQ was 108, but the performance IQ was unmeasurable because of a presence of object agnosia. His spontaneous speech was fluent without paraphasia, perseveration or dysprosody. His auditory comprehension of spoken language and repetition were spared. Neuropsychological examinations revealed an impairment in the recent verbal memory, alexia, agraphia, object agnosia, color naming difficulty, prosopagnosia and visuospatial constructional disability. The patient exhibited three major neuropsychological characteristics. Firstly, the recognition of the objects which were presented by the photograph or line drawing was more severely impaired than the real objects. Secondly, the object naming difficulty was more pronounced in the presentation at an unusual angle than in the free presentation. Thirdly, the patient made poor performance in reproduction of the models by photographic presentation as compared with the presentation of the real models in the three-dimensional block construction test. The CT scan performed on January 4, 1985 demonstrated subcortical hematomas with surrounding edema in the temporo-occipital regions of both hemispheres.(ABSTRACT TRUNCATED AT 250 WORDS)

Agnosia↗

PubMed1988

[Clinical significance of serum RNase activity in patients with eosinophilia].

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Adult↗

PubMed1987

Stimulation by GTP-binding proteins (Gi, Go) of partially purified phospholipase C activity from human platelet membranes.

A phospholipase C exhibiting preferential hydrolytic activity for polyphosphoinositides was partially purified from the deoxycholate extract of human platelet membranes by Q-Sepharose and Heparin-Sepharose column chromatographies. The activity of this purified phospholipase C free of the GTP gamma S-binding activity was stimulated at a similar level by addition of purified rat brain Gi or Go. These results suggest that GTP-binding proteins may interact directly with a solubilized membrane phospholipase C to stimulate its activity.

Animals↗

PubMed1987

Redistribution of protein kinase C in pancreatic acinar cells stimulated with caerulein or carbachol.

We examined phospholipid/calcium-dependent protein kinase (protein kinase C) activity and amylase secretion in isolated pancreatic acinar cells, when exposed to caerulein or carbachol. Upon stimulation with 10(-10) M caerulein or 10(-6) M carbachol cytosolic protein kinase C activity was increased in accordance with amylase secretion. Effect of carbachol on increase in membrane-associated protein kinase C activity was maximal at 10(-6) M where the rate of amylase secretion was highest. On the other hand, caerulein showed the maximal secretion of amylase at 10(-9) M, but the activity of the protein kinase C associated with membranes increased progressively with increasing concentration of caerulein. These results indicate different profiles of redistribution of protein kinase C upon stimulation of pancreatic acinar cells with carbachol or caerulein, and they were discussed in terms of amylase secretion.

Amylases↗

PubMed1987

Rat testosterone 7 alpha-hydroxylase. Isolation, sequence, and expression of cDNA and its developmental regulation and induction by 3-methylcholanthrene.

A P-450, designated P-450a, with high testosterone 7 alpha-hydroxylase activity was purified from rat liver microsomes. Specific polyclonal antibody against this P-450 was used to screen a lambda gt11 expression cDNA library and a 1687-base pair cDNA was isolated and sequenced. The deduced protein had 492 amino acids, a calculated Mr of 56,016, and it shared 51 and 45% amino acid similarities to P-450e and P-450f, respectively. Regions of similarity were distributed in distinct areas of high and low similarity along the P-450a primary sequence. P-450a cDNA was introduced into yeast cells using the expression vector pAAH5, and the resultant yeast microsomes contained both a protein of identical electrophoretic mobility to that of rat P-450a and testosterone 7 alpha-hydroxylase activity. These results confirm enzyme reconstitution data and antibody inhibition data that P-450a possesses testosterone 7 alpha-hydroxylase activity. The antibody and cDNA probes were used to examine the mechanism of regulation of P-450a by inducers and during development. P-450a and its mRNA were present at low level in newborn rats and increased to maximal level at 1 week of age in both males and females. At age 12 weeks, however, the P-450a level decreased in males but remained elevated in females. Concomitant with the decrease in P-450a in adult males was an increase in level of another immunologically related P-450. In adult male rats, P-450a was induced almost 5-fold by administration of 3-methylcholanthrene and this induction was the result of an increase in its mRNA. These results establish testosterone 7 alpha-hydroxylase as a member of the P-450e gene family that is developmentally regulated, sex-dependent, and markedly inducible by 3-methylcholanthrene.

Amino Acid Sequence↗

PubMed1987

Redistribution of phospholipid/Ca2+-dependent protein kinase in mast cells activated by various agonists.

Three types of agonists; receptor-mediated concanavalin A), direct (phorbol ester), and membrane-perturbing (compound 48/80), elicit histamine secretion from rat peritoneal mast cells. We tested whether activation of the mast cells by these agents is accompanied by subcellular redistribution of protein kinase C. Phorbol ester treatment predictably caused a profound decrease of phospholipid/Ca2+-dependent histone kinase activity in the cytosol and a concomitant increase of [3H]PMA-binding capacity in the membrane fraction, in a time- and concentration-dependent manner. Similar, but less marked effects were observed with stimulations by concanavalin A and compound 48/80. When mast cells labeled with [32P] and then stimulated with the agents, phosphorylation of a 50,000-Dalton protein was enhanced in the membrane fraction. These results suggest that protein kinase C may play a role in mast cell activation through phosphorylation of the membrane protein.

Animals↗

PubMed1987

A quantitative study of physiological cerebral atrophy with aging. A statistical analysis of the normal range.

A new method of discriminating pathological cerebral atrophy from physiological atrophy during aging is reported. The authors advocate a pixel counting method using a minicomputer for the quantitative measurement of cerebral atrophy. Five hundred cases were studied with this quantitative method and the normal range of the physiological atrophy was determined statistically. In order to estimate the degree of cerebral atrophy easily, the conventional linear measurement methods were compared with the pixel counting method using multivariant analysis, and a simple formula for the calculation of the degree of cerebral atrophy is proposed. Using this formula and the normal range, pathological cerebral atrophy is easily detectable.

Adolescent↗

PubMed1987

Effects of retinoic acid on receptors for epidermal growth factor in mouse palatal mesenchymal cells in vitro.

Retinoic acid (RA), one of the inducers of cleft palate in vivo, specifically increased the 125I-epidermal growth factor (EGF) binding capacity of embryonic palatal mesenchymal cells; the increase was completely inhibited by actinomycin D, suggesting that transcription is required for this change. When the increase in EGF receptors was related to the stimulation of DNA synthesis by EGF in the presence of various concentrations of RA, the maximum stimulation of [3H]-thymidine by EGF was at 10(-9) M RA, but the maximum increase in the number of EGF receptors was at 10(-6) M RA. Thus the increase in the number of EGF receptors cannot be related to EGF-stimulated DNA synthesis.

Animals↗

PubMed1987

Immunocytochemical demonstration of simultaneous synthesis of types I, III and V collagen and fibronectin in mouse embryonic palatal mesenchymal cells in vitro.

An immunocytochemical study was made on the palatal mesenchymal cells obtained from mouse embryos during palatal development with special reference to synthesis of types of collagen and fibronectin in vitro. Most cells showed positive staining with antibodies against the four proteins examined. The staining for type I collagen was most intense among the four proteins and was distributed perinuclearly. The staining for type III collagen was quite similar as that for type I collagen but less intense, whereas that for type V collagen was weak and its staining pattern was different from those for types I and III collagen in that the surface of the plasma membrane, in addition to the perinuclear cytoplasm, showed weak staining for type V collagen. Antibodies to fibronectin showed perinuclear and extracellular fibrous staining. These data suggest that palatal mesenchymal cells synthesize types I, III, and V collagen and fibronectin simultaneously.

Animals↗

PubMed1987

pH-dependent function, purification, and intracellular location of a major collagen-binding glycoprotein.

A major collagen-binding heat shock protein of molecular mass 47,000 D was found to bind to collagen by a pH-dependent interaction; binding was abolished at pH 6.3. Native 47-kD protein could therefore be purified from chick embryo homogenates in milligram quantities by gelatin-affinity chromatography and gentle acidic elution. Rat monoclonal and rabbit polyclonal antibodies were generated against the purified 47-kD protein. Immunofluorescence microscopy of cultured chick embryo fibroblasts with these antibodies revealed bright, granular perinuclear staining as well as a weaker reticular network structure towards the cell periphery, suggesting that this protein was located in the endoplasmic reticulum. No immunofluorescence staining was detected on the cell surface. Double-staining experiments with these antibodies and fluorescently labeled wheat-germ agglutinin suggested that the 47-kD protein was absent from the Golgi apparatus. Localization of the 47-kD protein in the endoplasmic reticulum but not in the Golgi complex was confirmed by immunoelectron microscopy. In vivo localization studies using immunohistochemistry of cryostat sections of chick liver revealed that the 47-kD protein was present in fibrocytes, Kupffer cells, and smooth muscle cells. It was absent from hepatocytes and the epithelia of bile ducts or sinusoidal endothelium. This major transformation- and heat shock-regulated glycoprotein is thus localized intracellularly, is expressed in only certain cells, and functions in a pH-regulated manner. These findings suggest that this glycoprotein is not likely to be a general cell-surface collagen receptor, but may instead play roles in intracellular protein processing or translocation.

Animals↗

PubMed1987

Debrisoquine 4-hydroxylase: characterization of a new P450 gene subfamily, regulation, chromosomal mapping, and molecular analysis of the DA rat polymorphism.

Debrisoquine 4-hydroxylase (P450db1) was purified from rat liver microsomes. Polyclonal antibody was produced and, in conjunction with immunoblots, was used to identify and purify a second immunorelated P450 (P450db2) that does not have debrisoquine hydroxylating activity. The cDNA clones to db1 and db2 were isolated from a lambda gt11 expression library, sequenced, and found to share 78% nucleotide and 73% deduced amino acid similarities. These similarities are evenly dispersed along the sequence except for a region of 190 nucleotides with 99% similarity near the carboxyl terminus of the protein-coding region; this similarity is probably the remnant of a gene conversion event. Both proteins share between 38% and 43% amino acid similarity with P450a, P450b, P450e, P450f, P450PB1, and P450j; these data indicate that P450db1 and P450db2 are members of a separate subfamily within the P450II gene family. Southern blot analysis and preliminary genomic cloning suggest that at least four genes exist in the subfamily, although the present evidence suggests that only db1 and db2 are expressed in rat liver. With the use of 19 mouse X hamster somatic cell hybrids, the db1 and db2 genes were localized to mouse chromosome 15 (P450-2D locus). A polymorphism has been described for debrisoquine metabolism in the DA rat strain, adult females having markedly decreased debrisoquine 4-hydroxylase activity. Our immunoblot analysis and mRNA analysis suggest that debrisoquine 4-hydroxylase deficiency in the female DA rat is not due to a decrease in db1 protein or mRNA. The db1 and db2 proteins are differentially regulated: during development db2 is present at birth while db1 is absent, and db1 increases by 1 week of age; in addition, db1 is slightly induced by phenobarbital, 3-methyl-cholanthrene, and dexamethasone whereas db2 is marginally increased by these latter two agents. These results demonstrate that debrisoquine 4-hydroxylase is a member of a new constitutively expressed P450II sub-family containing two or more genes in the rat and establish that the debrisoquine polymorphism in the DA rat is probably due to a structurally altered db1 protein.

Amino Acid Sequence↗

PubMed1987

Oligomeric structure of recombinant human and murine immune interferons by means of sedimentation equilibrium.

The oligomeric structure of two recombinant human immune interferon species, one with Cys-Tyr-Cys-Gln [( Cys-Tyr-Cys]rHu-IFN-gamma) and the other with Met-Gln-Asp-Pro (rHu-IFN-gamma) as the amino-terminal residues, and a recombinant murine immune interferon [( Cys-Tyr-Cys]rMu-IFN-gamma) were examined by means of sedimentation equilibrium. All three IFN-gamma s existed as dimers under nondenaturing conditions. Both [Cys-Tyr-Cys]rHu-IFN-gamma and [Cys-Tyr-Cys]rMu-IFN-gamma formed higher oligomers, tetramers, and octamers, respectively, under oxidative conditions, whereas rHu-IFN-gamma remained as dimers.

Animals↗

PubMed1987

In vitro synthesis of influenza viral RNA: characterization of an isolated nuclear system that supports transcription of influenza viral RNA.

An in vitro system for the synthesis of influenza viral RNA was developed using isolated nuclei prepared from influenza virus-infected HeLa cells. In this system, two species of positive-sense RNA, i.e., mRNA and cRNA (complementary RNA to vRNA), were found to be synthesized when analyzed by RNA-RNA hybridization using a minus-strand RNA probe and high resolution gel electrophoresis. The in vitro RNA synthesis required Mg2+, GTP, CTP, UTP, a high concentration of ATP, and an ATP regenerating system. Neither actinomycin D nor alpha-amanitin, potent inhibitors for cellular DNA-dependent RNA polymerases, inhibited the RNA synthesis. Addition of ApG or capped RNA, well-known primers for virion-associated RNA polymerase, markedly enhanced the extent of RNA synthesis. The maximum activity was observed with nuclei isolated from cells at 5 h after infection. This system is useful for the purification and characterization of factors involved in the transcription of these two species positive-sense RNA.

Adenosine Triphosphate↗