Biochemical systematics of Japanese fireflies of the subfamily Luciolinae and their flash communication systems

Japanese fireflies of the subfamily Luciolinae are biochemically analyzed using 13 allozymes, and the phylogenetic relationships obtained from this analysis are compared with their flash communication systems. As a result, the Japanese Luciolinae can be divided into three groups.Hotaria parvula andH. tsushimana together withLuciola yayeyamana andL. kuroiwae from the first group, and they use the same communication system.L. lateralis, Curtos okinawana, andC. costipennis make up the second group, and their communication systems are also the same.L. cruciata makes up the last one, and its communication system is different from the other fireflies of Luciolinae. Therefore, their taxonomical arrangement and communication systems are not congruent. However, the genetic similarity deduced by allozymic analysis of the members of the Japanese Luciolinae is highly consistent with their flash communication systems.     

Inhibition of gap junctional intercellular communication in heptachlor- and heptachlor epoxide-treated normal human breast epithelial cells

Based on the concern of organochlorides in the environment and in human tissue, this study was designed to determine whether various noncytotoxic levels of heptachlor and heptachlor epoxide could inhibit, reversibly, gap junctional intercellular communication in human breast epithelial cells (HBEC). Cytotoxicity and gap junctional intercellular communication (GJIC) were evaluated by lactate dehydrogenase assay and fluorescence redistribution after photobleaching analysis, respectively. Both heptachlor and heptachlor epoxide were noncytotoxic up to 10 μg/ml. At this concentration, heptachlor and heptachlor epoxide inhibited GJIC of normal human breast epithelial cells after 1 h treatment. Within a 24 h treatment with heptachlor and heptachlor epoxide at 10 μg/ml, recovery of GJIC had not returned. GJIC completely recovered after a 12 h treatment of 1 μg/ml heptachlor epoxide, but it did not recover after a 24 h treatment of 1 μg/ml heptachlor. RT-PCR and Western blots were analyzed to determine whether the heptachlor or heptachlor epoxide might have altered the steady-state levels of gap junction mRNA and/or connexin protein levels or phosphorylation state. No significant difference in the level of connexin 43 (Cx43) message between control and heptachlor-treated cells was observed. Western blot analyses showed hypophosphorylation patterns in cells treated with 10 μg/ml heptachlor and heptachlor epoxide for 1 h with no recovery within 24 h. Immunostaining of Cx43 protein in normal HBEC indicated that heptachlor and heptachlor epoxide caused a loss of Cx43 from the cell membranes at noncytotoxic dose levels. Taken together, these results suggest that heptachlor and heptachlor epoxide can alter GJIC at the post-translational level, and that, under the conditions of exceeding a threshold concentration in the breast tissue containing ‘initiated’ cells for a long time and not being counteracted by anti-tumor-promoting chemicals, they could act as breast tumor promoters.     

Development of human monoclonal antibodies: A review

This article describes the current status in the development of human monoclonal antibodies. Over the last ten years a lot of information about the human immune system has emerged. Combining these with the many new (bio-)technologies it is plausible that the long awaited breakthrough of this technology is close. This paper focuses on the classical cell-biological methods of achieving stable, antibody-producing human cell lines via cell fusion methods or virus derived transformations of human B-lymphocytes, as well as genetic engineering methods e.g. DNA libraries or phage display technology. The available in vitro immunization methods are critically reviewed and their impact on this topic is discussed. Therapeutic applications for cancer treatment or passive immunization against infectious diseases with antibodies derived by both ways are also reviewed.     

The gap junctional intercellular communication is no prerequisite for the stabilization of xenobiotic metabolizing enzyme activities in primary rat liver parenchymal cells in vitro

Summary In primary monocultures of adult rat liver parenchymal cells (PC), the activities of the xenobiotic metabolizing enzymes microsomal epoxide hydrolase (mEH_b), soluble epoxide hydrolase (sEH), glutathione S-transferases (GST), and phenolsulfotransferase (ST) were reduced after 7 d to values below 33% of the initial activities. Furthermore, the gap junctional intercellular communication (GJIC), measured after microinjection by dye transfer, decreased from 90% on Day 1 to undetectable values after 5 d in monoculture. Co-culture of PC with nonparenchymal rat liver epithelial cells (NEC) increased (98% on Day 1) and stabilized (82% on Day 7) the homotypic GJIC of PC. Additionally, most of the measured xenobiotic metabolizing enzyme activities were well stabilized over 1 wk in co-culture. Because GJIC is one of several mechanisms playing an important role in cell differentiation, the importance of GJIC for the stabilization of xenobiotic metabolizing enzymes in PC was investigated. PC in monoculture were, therefore, treated with 2% dimethyl sulfoxide (DMSO), a differentiation promoting factor, and 1,1,1-trichloro-2,2,-bis (p-chlorophenyl) ethane (DDT) (10 μg/ml), a liver tumor promotor and inhibitor of GJIC, was given to co-cultures of PC with NEC. DMSO significantly stabilized (68% on Day 7), while DDT significantly inhibited (8% on Day 7) homotypic GJIC of PC in the respective culture systems. In contrast, the activities of mEH_b, sEH, GST, and ST were not affected in the presence of DMSO or DDT. These results lead to the assumption that the differentiation parameters measured in this study (i.e., homotypic GJIC and the activities of xenobiotic metabolizing enzymes) are independently regulated in adult rat liver PC.     

Multiple peptide synthesis

The synthesis of large numbers of peptides can be very labor intensive and, if a conventional peptide synthesizer is used, only small numbers of peptides can be produced within a reasonable time. The techniques described below can make large numbers of different peptides simultaneously with varying degrees of mechanization, ranging from the wholly manual methods, to those involving complete mechanization of the whole synthesis process. Most of the multiple synthesis methods are primarily intended for small scale production ranging from microgram amounts up to a few tens of milligrams. All of the systems are economical in use of solvents and reagents, enabling cost-effective synthesis. The techniques described can also be used to prepare peptide libraries, containing several millions of peptide sequences, to enable the rapid screening of all possible permutations of amino acids within short peptides. However, it is considered that multiple synthesis methods are not particularly suited where extreme high purity or very long peptides are required.     

The biotechnology and applications of antibody engineering

The exquisite specificity of monoclonal antibodies (MAb) has long provided the potential for creating new reagents for the in vivo delivery of therapeutic drugs or toxins to defined cellular target sites or improved methods of diagnosis. However, many difficulties associated with their production, affinity, specificity, and use in vivo have largely confined their application to research or in vitro diagnostics. This situation is beginning to change with the recent developments in the applied molecular techniques that allow the engineering of the genes that encode antibodies rather than the manipulation of the intact antibodies themselves. Techniques, such as the polymerase chain reaction, have provided essential methods with which to generate and modify the genetic constituents of antibodies, allow their conjugation to toxins or drugs, provide ways of humanizing murine antibodies, and allow discrete modular antigen binding components to be produced. More recent developments of in vitro expression systems and powerful phage surface display technologies will without doubt play a major role in future antibody engineering and in the successful development of new diagnostic and therapeutic antibody-based reagents.     

Advanced sub social behaviour in the scorpionHeterometrus fulvipes Brunner (Arachnida)

Scorpions arc generally non-social, solitary animals that interact with conspecifics at birth, courtship or predation only. The present study reports the presence of advanced sub social behaviour inHeterometrus fulvipes Brunner and evaluates the importance of its burrowing as a cause for such social behaviour.Heterometrus fulvipes constructed deep angular burrows at the base of plants. Burrows provided (i) protection against predation, (ii) increased availability of food and (iii) ideal microclimate for year round activity of the scorpions. No cannibalism was observed in laboratory maintained colonies. The risk of predation and the difficult by immatures to dig tunnels during dry soil conditions may have forced the mother and offspring to live together in the burrow for longer durations. The cohabitation of relative offsprings transforms the burrow into a nest. The members of a colony exhibits division of labour for nest expansion and in foraging. The mother communicates with the immatures through “Buzz” sound and may provide premasticated food. There is food sharing also among colony members. All these behaviours indicate the presence of advanced sub social behaviour inHeterometrus fulvipes.     

Modulation of gap junction-mediated intercellular communication in embryonic chick mesenchyme during tissue remodeling in vitro

Gap junction-mediated intercellular communication was analyzed in a model system in which tissue necrosis and remodeling could be modulated. This in vitro system, previously used for analysis of epithelial-mesenchymal tissue interaction, was modified to permit analysis of the presence and extent of intercellular communition by monitoring intercellular transfer of the micro-injected fluorescent dye, Lucifer Yellow. Light and transmission electronmicroscopy were employed to correlate the presence and degree of gap junctional communication (coupling) with tissue morphology. Digital image analysis was used to determine cell density and mitotic indices within the outgrowths of explants. Our results indicated that cell communication in outgrowths adjacent to necrotic foci within an explant was minimal or absent. Cell-coupling in outgrowths adjacent to a compartment of viable mesenchyme was significantly higher-equivalent to unseparated control cultures. A time-course study demonstrated correlation of increased levels of cell-coupling in outgrowths with the level of tissue remodeling within an explant. Our conclusions from these studies are that embryonic mesenchymal cell populations may be selectively uncoupled as a result of alterations in the microenvironment produced by a proximate impaired cell population. It is proposed that endogenous factors in the microenvironment (wound signals), emanating from impaired cell populations, regulate gap junction-mediated intercellular communication in adjacent viable tissue. Normal, unimpaired populations of cells surrounding an area of injury are thereby isolated from the effects of a potentially toxic environment. This could serve as a protective function in development and may represent, in a more general sense, part of the repertoire of events associated with tissue repair and remodeling.     

Oscillations in the electrical resistance of Nitellopsis obtusa nodes

Oscillatory changes of the electrical resistance across the nodal complex of Nitellopsis obtusa (Desv. in Lois) J. Gr. were observed in experiments performed for 40–150 min with the use of external electrodes and microelectrodes. Three main patterns of node resistance oscillations were similar to those found for membrane potential and resistance. The presented findings indicate an oscillatory behaviour of the plasmodesmata system at the node, which may be connected with e.g. pulsatile variations in the number of open plasmodesmata.     

A 70-amino acid zinc-binding polypeptide fragment from the regulatory chain of aspartate transcarbamoylase causes marked changes in the kinetic mechanism of the catalytic trimer.

Interaction between a 70-amino acid and zinc-binding polypeptide from the regulatory chain and the catalytic (C) trimer of aspartate transcarbamoylase (ATCase) leads to dramatic changes in enzyme activity and affinity for active site ligands. The hypothesis that the complex between a C trimer and 3 polypeptide fragments (zinc domain) is an analog of R state ATCase has been examined by steady-state kinetics, heavy-atom isotope effects, and isotope trapping experiments. Inhibition by the bisubstrate ligand, N-(phosphonacetyl)-L-aspartate (PALA), or the substrate analog, succinate, at varying concentrations of substrates, aspartate, or carbamoyl phosphate indicated a compulsory ordered kinetic mechanism with carbamoyl phosphate binding prior to aspartate. In contrast, inhibition studies on C trimer were consistent with a preferred order mechanism. Similarly, 13C kinetic isotope effects in carbamoyl phosphate at infinite aspartate indicated a partially random kinetic mechanism for C trimer, whereas results for the complex of C trimer and zinc domain were consistent with a compulsory ordered mechanism of substrate binding. The dependence of isotope effect on aspartate concentration observed for the Zn domain-C trimer complex was similar to that obtained earlier for intact ATCase. Isotope trapping experiments showed that the compulsory ordered mechanism for the complex was attributable to increased "stickiness" of carbamoyl phosphate to the Zn domain-C trimer complex as compared to C trimer alone. The rate of dissociation of carbamoyl phosphate from the Zn domain-C trimer complex was about 10(-2) that from C trimer.(ABSTRACT TRUNCATED AT 250 WORDS)