Ultrastructural studies of human basophils and mast cells.

Ultrastructural studies of human mast cells (HMCs) and basophils (HBs) are reviewed. Sources of HMCs include biopsies of tissue sites and in situ study of excised diseased organs; isolated, partially purified samples from excised organs; and growth-factor-stimulated mast cells that develop de novo in cultures of cord blood cells. Sources of HBs for study include partially purified peripheral blood basophils, basophils in tissue biopsies, and specific growth factor-stimulated basophils arising de novo from cord blood cells. The ultrastructural studies reviewed deal with identity, secretion, vesicles, recovery, and synthesis issues related to the biology of these similar cells.     

Ultrastructural cytochemical and autoradiographic demonstration of nonspecific esterase(s) in guinea pig basophils

We used ultrastructural autoradiographic and cytochemical methods to localize esterase activities in unstimulated guinea pig basophils and in basophils undergoing degranulation or recovery from degranulation. We used tritium-labeled diisopropylfluorophosphate (DFP) as a probe for serine enzymes and localized this probe by ultrastructural autoradiography to cytoplasmic granules of immature or mature unstimulated basophils, as well as to granules released by degranulating basophils. Ultrastructural cytochemistry using alpha naphthyl acetate (ANA) as substrate localized nonspecific esterase activity to extruded granules, either within the interiors of degranulation sacs or within granules completely separated from degranulating basophils. Extruded granules retained their esterase activity for as long as 24 hr after antigen-induced degranulation. The plasma membranes of unstimulated or degranulating basophils, as well as of basophils recovering from degranulation, displayed prominent cell surface ANA esterase ectoenzyme activity. Lipid bodies, organelles present in the cytoplasm of both control and recovering basophils, were also alpha naphthyl acetate esterase (ANAE)-positive. Thus, cytochemical and autoradiographic techniques localized esterase and/or [3H]-DFP-binding activities to cytoplasmic granules, lipid bodies, and cell surface of basophils, and these enzyme activities persisted during both degranulation and recovery from degranulation.     

Modified Box-Cox transform for modulating the dynamic range of flow cytometry data

We describe an algorithm, Vout = Integer ([2(12)-1/2(12 lambda)-1] V lambda in-1) + 1; lambda greater than 0 based upon Box-Cox transformations as an alternative to nonlinear electronic amplifiers to expand or compress high- or low-amplitude flow cytometer-derived signals. If the indexing parameter lambda less than 1, input channels in the high-amplitude input range are compressed in the output range as occurs when an electronic logarithmic amplifier is used. However, if lambda greater than 1, input channels in the low-amplitude input range are compressed in the output range as occurs when an electronic power amplifier is used. Our modified Box-Cox transform can be implemented either during data collection or off-line for the transformation of previously collected raw data. The transform is the equivalent of an infinite class of nonlinear amplifiers. As the transform is implemented in software, it does not suffer from many of the disadvantages of nonlinear electronic amplifiers.     

Trypanosoma cruzi: histochemical characterization of parasitized skeletal muscle fibers

C57B1/6 mice were infected with Brasil strain Trypanosoma cruzi trypomastigotes. The leg muscles of the mice were serial-sectioned with a cryostat, and individual fibers were classified histochemically as type I or type II on the basis of succinic dehydrogenase or adenosine triphosphatase activity. Although markedly more type II fibers were present in the leg muscles, the percentage of infected type I fibers was nearly five-fold higher than type II. This is the first demonstration of a preferential in vivo distribution of T. cruzi in muscle fibers based upon muscle type.     

Trypanosoma cruzi: Histochemical Characterization of Parasitized Skeletal Muscle Fibers1

C57B1/6 mice were infected with Brasil strain Trypanosoma cruzi trypomastigotes. The leg muscles of the mice were serial-sectioned with a cryostat, and individual fibers were classified histochemically as type I or type II on the basis of succinic dehydrogenase or adenosine triphosphatase activity. Although markedly more type II fibers were present in the leg muscles, the percentage of infected type I fibers was nearly five-fold higher than type II. This is the first demonstration of a preferential in vivo distribution of T. cruzi in muscle fibers based upon muscle type.     

The centrifugal horizontal cells in the lobula plate of the blowfly, Phaenicia sericata

Intracellular recordings combined with iontophoretic injection of Procion Yellow M4RAN were used to study the anatomy and physiology of the centrifugal horizontal cells (CH-cells) in the lobula plate of the blowfly, Phaenicia sericata.Anatomy: The CH-cells comprise a set of two homolateral, giant visual interneurones (DCH, VCH) at the rostral surface of each lobula plate. Their extensive arborizations in the lobula plate possess bulbous swellings (boutons terminaux). The arborization of one cell (DCH) covers the dorsal, and the arborization of the other cell (VCH) the ventral half of the lobula plate. Their axons run jointly with those of the horizontal cells through the chiasma internum and the optic peduncle. Their protocerebral arborization possesses spines; they form a dense network together with the axonal arborization of the horizontal cells, a second type of giant homolateral cell most sensitive to horizontal motion. The protocerebral arborization of the CH-cells gives rise to a cell body fibre which traverses the protocerebrum dorsally to the oesophageal canal. The cell body lies on the contralateral side laterally and slightly dorsally to the oesophageal canal in the frontal cell body layer.Physiology: The CH-cells respond with graded potentials to rotatory movements of their surround. Cells in the right lobula plate respond with excitation (excitatory postsynaptic potentials, membrane depolarization) to clockwise motion (contralateral regressive, ipsilateral progressive), and with inhibition (inhibitory postsynaptic potentials, membrane hyperpolarization) to counterclockwise motion in either or both receptive fields; CH-cells respond to motion presented to the ipsilateral and/or contralateral eye. Cells of the left lobula plate respond correspondingly to the reverse directions of motion. Vertical pattern motion and stationary patterns are ineffective.The heterolateral H1-neurone elicits excitatory postsynaptic potentials in the DCH-cell; these postsynaptic potentials are tightly correlated 1:1 to the preceding H1-action potentíal. The delay between the peak of the action potential and the beginning of the DCH-postsynaptic potential is 1.15 msec, agreeing very well with the value reported previously for the blowfly, Calliphora (Hausen, 1976a). The synaptic input and output connections of the CH-cells are discussed.     

Chimeric gene expression using maize intron in cultured cells of breadwheat

High voltage electrical pulses were used to introduce the CAT reporter gene into cultured protoplasts of breadwheat,Triticum aestivum. Four DNA constructs harboring the CAT gene and the 35S or mannipine synthase promoter were tested for levels of CAT activity 40–45 hr after electroporation of protoplasts. One construct, containing a maize intron sequence between 35S and CAT sequences, conferred 30 to 185 fold greater CAT activity over the other three constructs. Data from these experiments suggest that a maize intron or sequences with similar effects may be required in DNA constructs for efficient heterologous gene expression in cultured cells of breadwheat.Abbreviations CAT Chloramphenicol acetyl transferase - NPT II neomycin phosphotransferase - 35S the 35S promoter of Cauliflower Mosaic Virus - PEG Polyethylene glycol - MES 2-[N-morpholino] ethanesulfonic acid     

Genome origins of Triticum cylindricum,Triticum triunciale,and Triticum ventricosum (Poaceae) inferred from variation in restriction patterns of repeated nucleotide sequences: a methodological study

Three methods of phylogenetic inferences on polyploid plants employing variation in restriction sites in repeated nucleotide sequences were compared. Allotetraploid Triticum species of well-established origin were used as a model. Methods based on determination of the proportion of restriction fragments shared between a polyploid and its diploid relatives generated biased results because of uneven numbers of restriction fragments among diploid species and presence of common bands in phylogenetically related diploid species. A method employing restriction fragments unique to a diploid species (marker bands) was not affected by either factor and generated results consistent with cytogenetic inferences. It is shown that the latter method can be used to investigate the origin of a polyploid species even when one of its progenitors is extinct or when the polyploid and its diploid progenitors have diverged.     

Characterization of the Early Stages of Genetic Salt-Stress Responses in Salt-Tolerant Lophopyrum elongatum,Salt-Sensitive Wheat,and Their Amphiploid

Eleven unique cDNA clones corresponding to genes showing enhanced mRNA accumulation in the early stages of salt stress (early salt stress induced, ESI) were previously isolated. The accumulation of these mRNAs in Lophopyrum elongatum (Host) A. Love, salt-sensitive wheat (Triticum aestivum L.), and their amphiploid is compared. The accumulation of ESI mRNAs was much greater in the L. elongatum roots than in the shoots. Additionally, mRNA accumulation in the roots of the three genotypes showed a biphasic response. The first phase occurred within a few hours after the onset of stress and had a large osmotic shock component, as indicated by induction of the accumulation of these mRNAs by a nonsaline osmoticum. The ion-specific component, however, also played a role. External Ca2+ reduced this response. The second phase was characterized by either constantly elevated mRNA levels or gradually increasing mRNA levels. The same biphasic response was elicited by exogenous abscisic acid (ABA). The response of all mRNAs to ABA closely approximated the response to 250 mM NaCl treatment in all three genotypes. The differences among the three genotypes in response to NaCl and ABA treatments were largely confined to the first phase of the response, in which mRNA levels were highest in L. elongatum and lowest in wheat. The levels of ESI mRNAs in the amphiploid closely approximated levels calculated on the basis of the doses of wheat and L. elongatum genomes in the amphiploid, which indicated an additive contribution of the genomes to early salt stress response in the amphiploid. The inducer of the ESI mRNA accumulation in response to NaCl and other osmotica is produced in the stressed roots and shows only minor, if any, translocation. A putative candidate for this inducer is root ABA.