Monomeric, monovalent derivative of Maackia amurensis leukoagglutinin. Preparation and application to the study of cell surface glycoconjugates by flow cytometry

A stable subunit of Maackia amurensis leukoagglutinin (MMAL) was prepared by the selective reduction of disulfide bridges between the subunits followed by alkylation with 4-vinylpyridine. MMAL failed to precipitate fetuin, whereas it retained its ability to bind to the same glycoprotein coated on a plastic plate, indicating the monovalency of this derivative. This binding to immobilized fetuin was inhibited by a haptenic sugar, Neu5Ac alpha 2-3lactose, with the same inhibitory potency as against the native M. amurensis leukoagglutinin. Microscopic observation as well as flow cytometric analyses showed that Chinese hamster ovary cells were clearly stained with fluorescein isothiocyanate-labeled MMAL without any detectable agglutination. This staining was inhibited by the addition of fetuin or by the sugar chains of fetuin. Differences in the types of sialylated glycoconjugates on the cell surface of several cell lines were detected by the combined use of fluorescein isothiocyanate-labeled MMAL and the monomeric derivative of elderberry bark lectin (specific for the Neu5Ac alpha 2-6Gal/GalNAc sequence) by flow cytometry. These results demonstrate the usefulness of these monovalent derivatives of sialylated oligosaccharide-specific lectins as probes for the analysis of cell surface glycoconjugates containing sialic acid by the technique of flow cytometry.     

Modified hippocampal long-term potentiation in PKC gamma-mutant mice

Calcium-phospholipid-dependent protein kinase (PKC) has long been suggested to play an important role in modulating synaptic efficacy. We have created a strain of mice that lacks the gamma subtype of PKC to evaluate the significance of this brain-specific PKC isozyme in synaptic plasticity. Mutant mice are viable, develop normally, and have synaptic transmission that is indistinguishable from wild-type mice. Long-term potentiation (LTP), however, is greatly diminished in mutant animals, while two other forms of synaptic plasticity, long-term depression and paired-pulse facilitation, are normal. Surprisingly, when tetanus to evoke LTP was preceded by a low frequency stimulation, mutant animals displayed apparently normal LTP. We propose that PKC gamma is not part of the molecular machinery that produces LTP but is a key regulatory component.     

Effect of 5-fluorouracil and UFT on experimental liver metastasis model of colorectal cancer using mouse colon 26 cells

Effects of 5-fluorouracil (5-FU) and UFT on an experimental liver metastasis model were compared at equi-effective dosage levels against subcutaneous tumor of mouse colon 26. 5-FU at the dosage level of 40 mg/kg suppressed the subcutaneous tumor growth by 70.0% and 45.0% on day 13 and day 18, respectively, and UFT at 20 mg/kg provided almost equal suppression (63.0% and 48.0%). In the liver metastasis model, 5-FU at 40 mg/kg showed more potent prevention of the formation of metastatic foci (94.9%) than did UFT (60.4%) at 20 mg/kg. 5-FU at 40 mg/kg produced a much higher peak serum level of 5-FU than did UFT at 20 mg/kg and also showed a much higher AUC (area under the curve) level in the portal blood. These results suggest that oral administration of 5-FU might be useful in prevention of liver metastasis of colorectal cancer.     

Erythroid potentiating activity of tissue inhibitor of metalloproteinases on the differentiation of erythropoietin-responsive mouse erythroleukemia cell line, ELM-I-1-3, is closely related to its cell growth potentiating activity

The erythroid-potentiating activity (EPA) of the tissue inhibitor of metalloproteinase-1 (TIMP-1) was re-examined using ELM-I-1-3, a mouse erythroleukemia cell line, which responded well to erythropoietin. Depletion of pre-existing TIMP-1 from fetal calf serum in culture medium using monoclonal antibody suppressed erythropoietin-induced differentiation as measured by the induction of hemoglobin, commitment assay and globin mRNAs. The removal of TIMP-1 also suppressed the proliferation of ELM-I-1-3 as measured by cell number and de novo DNA synthesis. These changes were reversed by the addition of purified TIMP-1 to the culture medium. Anti-TIMP-1 antibody also blocked both hexamethylene bisacetamide (HMBA)-induced erythroid differentiation and the proliferation of both ELM-I-1-3 and Friend erythroleukemia cells. Considering previous reports analyzing the chemical induction of Friend mouse erythroleukemia cell differentiation, our results suggest that erythropoietin- or HMBA-induced erythroid differentiation might also be coupled with cell proliferation. Our 3H thymidine-uptake experiment shows that TIMP-1 removal was also effective in the inhibition of cell growth of various other cell lines in addition to erythroleukemia cell lines. These results suggest that EPA action of TIMP-1 on erythroid leukemia cell lines is closely related to its activity to promote the cell growth of various cell lines and cells including erythroleukemia cell lines.     

Serotype-specific amplification of Rickettsia tsutsugamushi DNA by nested polymerase chain reaction

Polymerase chain reaction (PCR) with nested primer pairs was used to diagnose scrub typhus and identify the Rickettsia tsutsugamushi serotype. The primer pairs used for PCR were designed on the basis of the nucleotide sequence of the gene that encodes the 56-kDa antigen. Serotype-specific primers were used in the second PCR amplification. Five serovariants, the Gilliam, Karp, Kato, Kawasaki, and Kuroki strains of R. tsutsugamushi, were identified by nested PCR. In addition, the serotype identified by PCR with DNA from blood clots was the same as that of the strain isolated from five patients with scrub typhus. These findings indicate that this method is useful for diagnosis and identification of the rickettsial serotype in infected patients.     

Enkephalinergic innervation of GABAergic neurons in the dorsal raphe nucleus of the rat

The preembedding double immunoreaction method was used to study interrelations of enkephalinergic and GABAergic neuronal elements in the dorsal raphe nucleus of the Wistar albino rat. The enkephalin-like neuronal elements were immunoreacted by the peroxidase-antiperoxidase method and silver-gold intensified, which showed strongly and was specific. The GABA-like immunoreactive neurons were immunoreacted by the peroxidase-antiperoxidase method only. GABA-like neural somata were postsynaptic to both the enkephalin-like immunoreactive and the non-immunoreactive axon terminals. The enkephalin-like immunoreactive axon terminals were also found to synapse GABA-like immunoreactive dendrites. The GABA-like immunoreactive neuronal elements were also found to receive synapses from other non-immunoreactive as well as GABA-like immunoreactive axon terminals. Almost all of the synapses appeared to be asymmetrical. Possible functional activity of interactions among the enkephalinergic, GABAergic, and serotonergic neuronal elements in the dorsal raphe nucleus are discussed.     

F-actin bundling activity of Tetrahymena elongation factor 1 alpha is regulated by Ca2+/calmodulin

Translation elongation factor 1 alpha (EF-1 alpha) catalyzes the GTP-dependent binding of amino-acyl-tRNA to the ribosome. Previously, Tetrahymena 14-nm filament-associated protein was identified as EF-1 alpha [Kurasawa et al. (1992) Exp. Cell Res. 203, 251-258]. This and several other studies suggest that EF-1 alpha functions not only in translation but also in regulation of some part of the cytoskeleton. Tetrahymena EF-1 alpha bound to F-actin and induced bundling of F-actin. We investigated the effects of GTP/GDP and Ca2+/calmodulin on F-actin bundling activity of EF-1alpha. The presence of GTP, GDP, or guanylyl-imidodiphosphate (GMP-PNP) slightly decreased the amount of EF-1 alpha which bound to F-actin, but each had virtually no effect on the F-actin bundling activity. The formation of F-actin bundles by EF-1 alpha was Ca(2+)-insensitive. In the absence of Ca2+, calmodulin did not bind to EF-1 alpha and F-actin. On the other hand, in the presence of Ca2+, calmodulin directly bound to EF-1 alpha but did not have any serious influence on EF-1 alpha/F-actin binding. Under the conditions, electron microscopy demonstrated that Ca2+/calmodulin completely inhibited the F-actin bundling by EF-1 alpha. These results indicate that CA2+/calmodulin regulates the F-actin bundling activity of EF-1 alpha without inhibition of the binding between Ef-1 alpha and F-actin.     

Caspase 1-independent IL-1beta release and inflammation induced by the apoptosis inducer Fas ligand

Anchietia salutaris tea is traditionally used in Brazil to treat allergies, suggesting it contains compounds with antagonistic activity on the allergic mediators. We have evaluated extracts and semi-purified fractions of Anchietia salutaris as a source of compounds having this type of antagonism on the contraction induced in guinea-pig lung parenchymal strips and on platelet aggregation and shape change. After 10 min pre-incubation dichloromethane extracts containing 30 or 100 microg mL(-1) inhibited the contraction induced by prostaglandin D2 (PGD2) in guinea-pig lung parenchymal strips with dose ratios (DR) of 0.76+/-0.14 and 0.93+/-0.19, respectively; the amount of inhibition depended both on the concentration and on the time of pre-incubation (DR after 30 min pre-incubation was 1.21+/-0.51). The dichloromethane extract and its semi-purified fractions also inhibited the contractions induced by U46619, a more potent, stable, synthetic agonist of thromboxane A2 (TxA2) prostanoid (TP) receptors, the receptors acted upon by PGD2 to produce lung contractions. The dichloromethane extract did not inhibit the lung parenchymal contractions induced by histamine, leukotriene D4 (LTD4) or platelet-activating factor (PAF). Platelet aggregation induced by U46619, adenosine 5'-diphosphate (ADP) or PAF was not inhibited by the dichloromethane extract. Indeed, the extract potentiated platelet aggregation induced by low concentrations of these agonists and also potentiated the shape change induced by U46619. These results imply that the dichloromethane extract of Anchietia salutaris and its semi-purified fractions contain an active principle that competitively inhibits TxA2 TP receptors, the stimulation of which causes lung parenchymal contraction. The inhibition seems to be selective for this receptor subtype, because the extract fails to inhibit platelet aggregation or shape change. This provides additional support of earlier reports suggesting the occurrence of TP receptor subtypes.     

R&D activities on wireless systems in China

Various wireless systems and the research preceding their practical use in China are described. The topics discussed include research establishments, research and development projects underway, challenges inherent to digital radio, and expansion opportunities afforded by satellite communications     

An efficient technique for detecting catastrophic convolutional codes

Catastrophic convolutional codes (CC) cause an infinite number of decoded data bit errors when decoding a finite number of code symbols. A CC displays a catastrophic error propagation if the generating polynomials have a common factor. An efficient algorithm for polynomial factorization in GF(2^m) is used for detecting catastrophic CC for any rate n/m and constraint length k. A general formula is derived to calculate the number of catastrophic codes in any (m, n, k) CC.