In vitro activation of leukocyte glycogen synthetase

The activity of leukocyte glycogen synthetase in a freshly prepared homogenate is almost completely in the $\text{b}$ form. Incubation of the homogenate at 30°C caused a time dependent increase in the activity measured in the absence of G-6-P ($\text{b}$ to $\text{a}$ conversion). The K_a for G-6-P decreased from 0.7 to 0.01 mM. Freezing of the homogenate resulted in a complete loss of the capacity for activation. These results demonstrate that glycogen synthetase from leukocytes of normal human subjects can be converted $\text{in vitro}$ to a form, which is almost independent of G-6-P for activity.     

Leukotriene A: stereochemistry and enzymatic conversion to leukotriene B

Leukotriene A was assigned the structure 5(S)-$\text{trans}$-5,6-oxido-7,9-$\text{trans}\text{-11,14-}\text{cis}$-eicosatetraenoic acid by the enzymatic conversion of a synthetic product of known stereochemistry into the naturally occurring isomer of 5(S),12(R)-dihydroxy-6,8,10,14-eicosatetraenoic acid in human polymorphonuclear leukocytes.     

Comparison of circulating and peritoneal leukocyte cationic proteins in release of histamine from rat mast cells

Lysosomal cationic proteins which release histamine from rat peritoneal mast cells were prepared from circulating as well as peritoneal leukocytes of rabbits. The release of histamine by cationic proteins and by compound $\text{48}\text{80}$ was compared as a function of temperature, pH and concentration. Cationic protein-mediated histamine release appears to be a non-cytotoxic energy requiring process similar to compound $\text{48}\text{80}\text{-}\text{mediated release}$. It was inhibited by iodoacetate, n-ethylmaleimide, 2,4-dinitrophenol, malonate, oxamate, glutamate and slightly inhibited by 2-deoxyglucose. Pharmacologic inhibition of release by isoproterenol, aminophylline, dibutyryl cyclic AMP and prednisone was also demonstrated.     

Chemotactic migration of neutrophils under agarose.

A simple test for neutrophil chemotaxis is described. Wells were cut in soft agarose gel and filled with human peripheral blood leukocytes, chemotactin and control substances. Neutrophils consistently migrated under agarose towards the well with chemotactin, but not towards wells with control substances. Chemotaxis was quantitated as the mean distance travelled by 10 cells farthest from the well of origin, at specified time-intervals after filling the wells. Approximately $\text{1}\text{3}$ distance was covered in 2 hours, $\text{3}\text{4}$ in 4 hours and 90 per cent in 6 hours. The migrating cells examined after fixation and staining were found to be predominantly neutrophils with occasional eosinophils and monocytes.     

Enzymatic formation of 14,15-leukotriene A and C(14)-sulfur-linked peptides

Human leukocytes converted [³H]-(S)-15-HPETE into [³H]-14,15-LTA. Rat basophilic leukemia cells transformed 14,15-LTA into two bioactive C(14)-S-linked peptides, which have been characterized as 15(S)-hydroxy-14(R)-S-glutathionyl-5,8$\text{Z}$,10,12$\text{E}$-icosatetraenoic acid and 15-(S)-hydroxy-14(R)-S-cysteinylglycyl-5,8$\text{Z}$,10,12$\text{E}$-icosatetraenoic acid by comparison with synthetic specimens.     

Phylogeny of immune responsiveness in invertebrates

Existence of essential immunocompetence at different levels of complexity has recently been demostrated among diverse phylums of invertebrates. $\text{Immunorecognition}$ or capacity for non-self recognition of allogeneic tissue followed by incompatibility reactions is already found in Coelenterates as shown in colonial hydroids, gorgonians, and hard corals. Antagonistic reactions reveal convincing specificity, with compatible fusion preceding manifestations of incompatibility, but no memory component has been demonstrated. Allogeneic contact incompatibilities found in protochordates as well as mixed lymphocyte culture reactions between allogeneic cells of various vertebrates may tentatively be assigned to this “sipmle” immunorecognition category. $\text{Primordial Cell-Mediated Immunity}$ (PCMI) represents a higher level of immunoevolution revealed by specific allograft reactions with at least short-term memory as found in Annelids and Echinoderms. PCMI is inherent in populations of leukocytes and definitely has an immune memory component. $\text{Integrated Cell-Mediated and Humoral Antibody Immunity}$ may be regarded as the highest level of immunoevolution characteristic of all vertebrates from primitive fishes to mammals. Vertebrate-type immunoglobulin antibodies have yet to be demonstrated in any invertebrate species, although cell-surface receptors akin to antibodies are apparent on leukocytes of advanced invertebrates. The less elaborate immunoresponsiveness evident among invertebrates awaits definitive characterization.     

Leukotriene A4: Enzymatic conversion to leukotriene C4

An unstable epoxide, leukotriene A₄ (5(S)-$\text{trans}$-5,6-oxido-7,9-$\text{trans}$-11,14-$\text{cis}$-eicosatetraenoic acid), was earlier proposed to be an intermediate in the conversion of arachidonic acid into the slow reacting substance (SRS), leukotriene C₄. In the present work synthetic leukotriene A₄ was incubated with human leukocytes or murine mastocytoma cells. A lipoxygenase inhibitor, BW755C, was added in order to prevent leukotriene formation from endogenous substrate. Leukotriene C₄ and 11-$\text{trans}$-leukotriene C₄ were the main products with SRS activity. It was not established whether the 11-$\text{trans}$-compound was formed by isomerization at the leukotriene A₄ or C₄ stage.     

Tuftsin: A hormone-like tetrapeptide with antimicrobial and antitumor activities

A specific fraction of immunoglobulin G binds to polymorphonuclear neutrophils and stimulates their phagocytic activity. This phagocytosis-stimulating activity resides solely in a small peptide termed tuftsin, of the sequence Thr-Lys-Pro-Arg, which has been isolated from the leukophilic immunoglobulin G fraction. The physiological significance of tuftsin has been demonstrated in splenectomized patients and patients with a congenital tuftsin abnormality, in whom the low levels of tuftsin in sera (measurable by radioimmunoassay) coincides with a high incidence of infection. Tuftsin has also been shown to enhance bactericidal activity in addition to phagocytosis. Its biological activities appear to be mediated via specific tuftsin receptors which have been found on macrophages, monocytes and granulocytes. In addition, tuftsin possesses chemotactic, migration-enhancing and mitogenic properties for leukocytes and has recently been shown to enhance their anti-tumor activity $\text{in}$$\text{vitro}$ as well as $\text{in}$$\text{vivo}$. Other known activities of tuftsin include effects on the activity of the hexose monophosphate shunt, on the concentrations of intracellular cyclic nucleotides and on the efflux of Ca²⁺ in leukocytes. Tuftsin has been chemically synthesized in various laboratories using different procedures and also is available commercially. The above features of tuftsin plus the expected low toxicity of this peptide make tuftsin a very attractive agent for immunotherapy against infection and cancer. However, a great deal of caution needs to be exercised when using tuftsin due to inhibitory contaminants found in certain commercial preparations.     

Characterization of two crystal forms of neutrophil cationic protein NP2, a naturally occurring broad-spectrum antimicrobial agent from leukocytes

A hexagonal crystal form (P6₃22, $\text{a = b = 34.0}\text{A}\text{?}\text{, c = 113.5}\text{A}\text{?}$) and a monoclinic form (P2₁, $\text{a = 37.1}\text{A}\text{?}\text{, b = 32.2}\text{A}\text{?}\text{, c = 32.4}\text{A}\text{?}\text{, β = 110 °}$) of neutrophil cationic protein NP2, isolated from rabbit leukocytes, have been characterized. The monoclinic form, containing two promoters (M_r = 3844) per asymmetric unit, diffracts to at least 1.8 Å and is suitable for high-resolution structural studies.     

Transfer factor and cellular reactivity to varicella-zoster antigen in childhood leukemia

The present studies were designed to evaluate possible efficacy of specific human transfer factor (TF) in preventing or attenuating varicella-zoster (VZ) infection in children with acute lymphocytic leukemia (ALL). TF was prepared following leukapheresis of adult donors who were convalescing from chicken pox. Recipients were VZ seronegative children with ALL, 12 in remission and 3 in relapse; a single injection of TF was given equivalent to 10⁸ lymphocytes per 7 kg body wt. The following VZ-specific parameters were measured: lymphocyte blastogenesis, cytotoxicity and leukocyte inhibitory factor (LIF) production, and indirect fluorescent and CF antibody titers. No patients in relapse converted immune response while $\text{10}\text{12}$ in remission developed positive reactivity in at least one assay of cell-mediated immunity (CMI); $\text{3}\text{12}$ were positive in all three parameters of CMI and $\text{8}\text{12}$ in two assays. Cytotoxicity was the most consistently positive test following TF administration. No patients developed VZ antibody. Seventeen months follow-up has demonstrated persistent cellular immunity.     

A role of cathepsin B1 in polymorphonuclear leukocytes chemotaxis.

Cathepsin B_I¹ was purified from rat liver lysosomal fraction by ammonium sulfate fractionation, followed by chromatography on Sephadex G-200 and DEAE-Sephadex. Formation of chemotactic factor for guinea pig polymorphonuclear (PMN) leukocytes was demonstrated $\text{in vitro}$ when guinea pig serum was incubated with cathepsin B_I. This factor formation was dependent on SH-reagent dithiothreitol (DTT) and was maximal at pH 6.0. ZnSO₄, an inhibitor of cathepsin B_I, inhibited the chemotactic factor formation likewise.     

Mathematical analysis of a capillary migration assay for cellular immune sensitivity: simple theory for dose-response.

A simple model is used to investigate parametric dependences of in vitro leukocyte assays for cellular immune sensitivity. The model is first tested by computing areas of cell migration occurring when migration inhibition factors (MIF) are absent from the culture medium. The theory subsequently is modified to include MIF production, and analytic expressions for the migration inhibition index $\text{I}$ are derived. These then are evaluated in order to study consequences of varying such factors as the ratios of constituent leukocytes or kinetic parameters pertaining to the ability of the culture medium to sustain migration.     

The separation of phage promoter from bacterial lac promoter for beta-galactosidase expression in transducing phage lambda plac5.

The replication defective transducing phage λp$\text{lac}\text{5}\text{O}\text{29}\text{P}$3 carries a portion of the $\text{E.}\text{coli}\text{lac}$ operon in the $\text{b}$2 region of the lambda phage. This $\text{lac}$ operon segment contains the $\text{lac}$ promoter, the $\text{lac}$ operator, and the β-galactosidase $\text{z}$ gene, but does not contain the $\text{lac}$ repressor $\text{i}$ gene. The $\text{z}$ gene can be expressed from both the inserted $\text{lac}$ promoter and the phage promoter. When $\text{E.}\text{coli}$ strain 594 ($\text{z}$^?, $\text{i}$⁺) or JC6256 (Δ$\text{lac}$) is infected by λp$\text{lac}\text{5}\text{O}\text{29}\text{P}$3 in the absence of additional cyclic AMP, β-galactosidase synthesis is shown to be expressed from the phage promoter. When 594 (λ⁺) or JC6256 (λ⁺) is infected by λp$\text{lac}\text{5}\text{O}\text{29}\text{P}$3 in the presence of additional cyclic AMP and IPTG, β-galactosidase synthesis is shown to be expressed from the inserted $\text{lac}$ promoter.The ability to separate the phage promoter from the inserted $\text{lac}$ promoter for β-galactosidase expression will simplify the interpretation whenever λp$\text{lac}$5 is used.     

The biosynthesis of phosphorylated tyrosine hydroxylase by organ cultures of rat adrenal medulla and superior cervical ganglia: a correction.

Determination of the complete amino acid sequence of the rubredoxin isolated from the sulfate reducing bacterium $\text{Desulfovibrio}$$\text{gigas}$ showed that the molecule consists of a single polypeptide chain of 52 residues. The sequence of the first 42 residues was determined using an automatic Protein Sequencer. Peptides derived from tryptic hydrolysis and from specific cleavage at tryptophan residue were used to construct the total sequence. Compared with the sequence of $\text{Desulfovibrio}$$\text{vulgaris}$ rubredoxin, 37 positions are identical, and with the sequences of $\text{Clostridium}$$\text{pasteurianum}$, $\text{Peptostreptococcus}$$\text{elsdenii}$, $\text{Micrococcus}$$\text{aerogenes}$ and $\text{D.}$$\text{vulgaris}$ rubredoxins, 20 matching residues occur. A crystallographic study of the $\text{D.}$$\text{gigas}$ rubredoxin is in progress.     

Two sex pheromone components of the tobacco budworm moth, Heliothis virescens

Two compounds were isolated from female $\text{Heliothis}$$\text{virescens}$ (Lepidoptera: Noctuidae) extracts and identified as $\text{cis}$-9-tetradecenal and $\text{cis}$-ll-hexadecenal. Together they elicit intense male $\text{H}$. $\text{virescnes}$ response in laboratory tests and have attracted males in the field. Although $\text{cis}$-ll-hexadecenal is an $\text{H}$. $\text{zea}$ sex pheromone, no evidence was obtained for $\text{cis}$-9-tetradecenal in $\text{H}$. $\text{zea}$.     

Seasonal effects on female reproductive functions in the bovine (Indian breeds)

Reproductive function is mediated by season in the Indian breeds of cattle ($\text{Bos}$$\text{indicus}$). The reproductive endocrinology of $\text{Bos}$$\text{indicus}$ cattle differs from that of $\text{Bos}$$\text{taurus}$ breeds; the estrus is shorter and less intense and occurs late in relation to an estrogen stimulus. Moreover, the $\text{Bos}$$\text{indicus}$ female has a smaller preovulatory surge of luteinizing hormone (LH), which occurs earlier relative to the onset of estrus, and she ovulates sooner after the onset of estrus. The corpus luteum is smaller and contains less progesterone, and the serum progesterone concentration is lower in $\text{Bos}$$\text{indicus}$ females. Furthermore, they have fewer preovulatory LH surges than $\text{Bos}$$\text{taurus}$ females and their luteal cells are less responsive to LH in vitro during the winter. Their fertility is lower during the late fall and winter months. For $\text{Bos}$$\text{indicus}$ cattle, recovery of transferable embryos and survival of embryos in the recipient are at their maximum from July through October.     

Hepatic microsomal N-oxidation and N-demethylation of N,N-dimethylaniline in red-winged blackbird compared with rat and other birds.

Hepatic microsomes prepared from red-winged blackbirds ($\text{Agelaius phoeniceus}$) and albino rats were incubated with $\text{N}$,$\text{N}$-dimethylaniline (DMA)_in complete incubation mixtures at pH 7.9 and 37°C for 10 min. Formaldehyde and $\text{N}$,$\text{N}$-dimethylaniline-$\text{N}$-oxide produced from DMA were measured. Redwings were found to have significantly lower $\text{N}$-demethylation activities than rats, and redwings had only marginal or no $\text{N}$-oxidation activities. Hepatic microsomes from redwings did not further metabolize the $\text{N}$-oxide. The $\text{N}$-oxidation and $\text{N}$-demethylation activities of brown-headed cowbirds ($\text{Molothrus ater}$), common grackles ($\text{Quiscalus quiscula}$), and starlings ($\text{Sturnus vulgaris}$) were similar to those of redwings.