Studies of the diphtheria toxin receptor on chinese hamster cells

Concanavalin A, wheat germ agglutinin and the ovalbumin glycopeptide are all inhibitors of the cytotoxic effect of diphtheria toxin on Chinese hamster cells. Ovalbumin glycopeptide loses its inhibitory property after treatment with β-N-acetylglucosaminidase. This demonstrates the importance of the glycopeptide structure for the mechanism of inhibition. The glycopeptide may be a toxin cell-surface receptor analogue. Diphtheria toxin-resistant mutants were isolated in order to search for cells that might have an altered toxin receptor. One mutant was 10-to 15-fold more resistant to diphtheria toxin than wild-type cells when protein synthesis was measured as a function of toxin concentration. However, when protein synthesis was measured as a function of time at a high toxin concentration, the time before onset of inhibition was identical in the mutant and wild-type cells. We present evidence indicating that the resistance of this mutant can be accounted for by a decreased affinity of toxin for a cell-surface receptor.     

Theoretical analysis of the effects of two pH regulation patterns on the temperature sensitivities of biological systems in nonhomeothermic animals.

When protonation reactions comprise a part of biochemical reaction schemes in vivo, the temperature sensitivities of chemical equilibria and of enzymatic rates are modulated according to the variation of pH with T_B (body temperature). Two patterns of pH regulation have been established, each pattern controlling the ionization states of different titratable groups as T_B changes: $\text{dpH}\text{dT}{}_{\mathrm{<mtext>B</mtext>}}\text{? 0}$ pH unit/° C for the blood of heterothermic mammals (constant charge for phosphate groups, e.g.), and $\text{dpH}\text{dT}{}_{\mathrm{<mtext>B</mtext>}}$ ? ?0.017 pH unit/° C for intracellular and blood compartments of vertebrate and invertebrate poikilotherms (constant charge for histidine imidazole and -SH groups). Calculations demonstrate the feasibility of the following. (i) A large effect of T_B on a metabolic branch point is possible when the competition of two pathways for common substrate is governed by two titratable groups with large differences in ΔH_i^o values (ionization enthalpy), e.g., phosphate and alpha amino groups. (ii) The effect of temperature on the amount of free energy released from equilibrium chemical reactions (as might occur for reactions not controlled by rate-limiting enzymes) would depend strongly on the value of ΔH_i⁰, and on which of the two pH regulation patterns was present. (iii) If free energy released from ionization reactions were coupled to activation processes of enzyme catalysis, so as to effectively decrease free energy barriers, the temperature sensitivities of reaction rates would be smaller in pH environments in which pH varied inversely with T_B than in environments of constant pH, regardless of ΔH_i⁰. (iv) Different mechanisms for the modulation of metabolism as a function of T_B might have evolved for animals with different values of $\text{dpH}\text{dT}{}_{\mathrm{B}}$.     

Amaurosis fugax and carotid artery disease: indications for angiography.

A prospective study of 80 patients presenting with amaurosis fugax was performed in an attempt to relate clinical features to angiographic findings in the internal carotid artery. Carotid bruit, transient cerebral ischaemic attacks, hypertension, and claudication were associated with a high prevalence of angiographic abnormality. Every patient who showed all these features had an operable lesion, as did 88% of those who had three features. In patients over 50 years of age carotid stenosis and atheromatous ulceration were occasionally found in the absence of bruit and transient cerebral ischaemia, but only one patient aged under 50 had an operable lesion and no associated features. Clinical features were therefore valuable in predicting the outcome of angiography, but it seems prudent to restrict angiography to patients aged over 50, who are most likely to benefit from surgery on the carotid artery.     

Prospectively randomised trial of proximal gastric vagotomy either with or without pyloroplasty in treatment of uncomplicated duodenal ulcer.

A consecutive series of 100 men with uncomplicated duodenal ulcer was randomly divided into two groups: one group of 52 underwent proximal gastric vagotomy (PGV), the other group (48) underwent PGV with pyloroplasty (PGVP). Preoperative peak acid output (PAOP) was measured in all patients. Those with a higher preoperative PAOP were significantly more likely to develop recurrent ulceration. Three patients developed recurrent ulceration after PGV and seven after PGVP. Dumping was both more common and more severe after PGVP than PGV. An overall satisfactory result was achieved in 92% after PGV and 81% after PGVP. We conclude that combining pyloroplasty with PGV has no appreciable advantages.     

Inhibition of lymphoma cell proliferation by supernatant from fibrosarcoma cultures: Preliminary evidence that the inhibitory material is prostaglandin E

Supernatants obtained from mouse fibrosarcoma cultures 48 hr after the addition of fresh medium contained dialyzable material which inhibited the proliferation of syngeneic lymphoma cells , as measured by ³H-thymidine incorporation. Three lines of evidence indicate that the supernatant inhibitory material is probably prostaglandin (PG) E. First, the supernatant and dialysate of the supernatant contained a substance with the same characteristics as PGE₁ or PGE₂ as detected by thin layer chromatography. Second, PGE₂-treatment of lymphoma cells mimicked the inhibition of proliferation observed with supernatant inhibitory substance. Third, indomethacin treatment of fibrosarcoma cultures reduced the amount of supernatant inhibitory substance present.     

Modified Whipf's Polychrome: A Connective Tissue Stain with Special Application for Demonstrating Leishmania

A polychrome stain procedure was developed to demonstrate amastigotes of the protozoan parasite Leishmania braziliensis as well as cytoplasmic and other tissue components in cutaneous lesions of infected animals. The procedure is as follows: stain nuclei for 10 minutes with an iron hematoxylin containing 0.5% hematoxylin and 0.75% ferric ammonium sulfate dissolved in 1:1 0.6 N H₂SO₄:95% ethanol; rinse 4 minutes in distilled water. Cytoplasmic staining is achieved by exposing tissues for 10 minutes to a solution containing 0.25% Biebrich scarlet, 0.45% orange G, 0.5% phosphomolybdic acid and 0.5% phosphotungstic acid in 1% aqueous acetic acid. These first two solutions are modified from Whipf's polychrome stain. Sections are differentiated for 10 seconds in 50% ethanol, rinsed in water, stained 3 minutes in 0.1% aniline blue WS in saturated aqueous picric acid, rinsed in water and differentiated for 1 minute in absolute ethanol containing 0.05% acetic acid. Mordanting overnight in 6% picric acid in 95% ethanol produced optimal results.

This procedure was applied to sectioned material from experimental animals with various protozoa. Trypanosoma cruzi, Besnoitia Jellisoni, Toxoplasma gondii and especially Leishmania braziliensis were well demonstrated. Combining cytoplasmic dyes and phosphomolybdic-phosphotungstic acids into one solution afforded differential staining of tissues by Biebrich scarlet and orange G; connective tissues were stained by this solution. Substantially improved definition of connective tissues resulted after counterstaining. This procedure differs from the Massou sequence in which connective tissues are first stained by cytoplasmic dyes along with other tissues and then destained prior to specific counter-staining. in comparing dyes structurally related to Biebrich scarlet, it was found that Crocein scarlet MOO, but not Poncenu S, was an acceptable substitute. Sirius supra blue GL and Sirius red FSBA were not useful as replacements for aniline blue WS in this procedure.     

Comparison of in vivo and in vitro ribosomal RNA synthesis in nucleolar mutants ofXenopus laevis

Summary Cells of embryos carrying a lethal nucleolar mutation have been maintained in vitro for extended periods of time. Normally these mutants live only 9 to 12 days after fertilization but their cells in culture will survive for more than 3 months. The extent of ribosomal RNA (rRNA) synthesis was determined in primary cultures prepared from normal embryos and nucleolar mutants having different numbers of ribosomal RNA genes. We found that the accumulation of radioactivity into rRNA for normal and mutant embryos was similar in vivo and in vitro. In primary cultures of normal embryos which have two nucleoli per cell and mutant embryos which have only one nucleolus per cell, the incorporation of radio-activity into rRNA was similar even though the normal cells have twice as many rRNA genes. Thus the mechanism which regulates dosage compensation of the rRNA genes operates both in vivo and in vitro. This work was supported by Grant GB38651 from the National Science Foundation.     

The spatial frequency sensitivity of bipolar cells

Retinal bipolar cells constitute the output stage of the outer layer of the retina. There are several constraints on the ability of the bipolar cell array to respond to the different spatial frequency components of the visual image, including (i) electrical coupling in the dendritic tree receiving receptor input; (iii) the "lateral inhibition" mediated by horizontal cells. Using simple mathematical models, we derive analytical expressions for the spatial frequency response of the bipolar cell array for the case in which horizontal cells are presynaptic to bipolar cells (feedforward model) and also for the case in which horizontal cells are presynaptic to receptors (feedback model). The results illustrate the importance of the three factors mentioned in determining the bipolar cells' properties. The optimal spatial frequency for stimulating the bipolar cell array, and the range of spatial frequencies transmitted onward to the inner plexiform layer, are thus related to the anatomical and electrical properties of the cells in the outer plexiform layer.     

Duplication of the segment q12.2→qter of chromosome 22 due to paternal inversion 22(p13q12.2)

Summary A 1730-g male infant, born at 37 weeks gestation, had multiple congenital anomalies, consisting of microcephaly, hypertelorism, bilateral cleft lip and palate, micrognathia, lowset ears, and cryptorchidism. Chromosome analysis showed a recombinant 22 derived from the paternal inversion (22) (p13q12.2). The proband's karyotype is 46,XY,rec(22),dup q,inv(22)(p13q12.2)pat, which has a duplication of q12.2qter. An identical recombinant has been reported in a female infant in Mexico whose mother was a carrier of the inversion. Similar congenital anomalies present in these two patients demonstrate the phenotype of duplication of the distal long arm 22. This report also documents the occurrence of an identical inversion in two apparently unrelated Mexican families.