Survival after total-body irradiation. I. Effects of partial small bowel shielding

The small intestine of the rat was shielded during total-body irradiation (TBI) to evaluate the effects of radiation dose and length of intestine shielded on survival. Sprague-Dawley rats were anesthetized in groups of 10. Using aseptic surgical procedures 80, 40, 20, or 10 cm, or none of the proximal or distal small intestine were temporarily exteriorized and shielded during irradiation with photons from an 18 MeV linear accelerator. Less than 17% of the dose was delivered to the shielded intestines. In unshielded animals deaths occurred from Days 4 to 6 with 13, 15, or 17 Gy and from Days 8 to 30 with 9, 11, and 12 Gy. However, in all animals exposed to 15 Gy with all or part of the small intestine shielded, survival was increased to between 5 and 9 days. Shielding of the distal small intestine was more effective in prolonging survival than shielding of the proximal small intestine. The previously identified target of radiation damage in the small intestine is the crypt stem cell. In this study, the analysis of histological specimens of shielded and irradiated small intestine suggested that humoral factors also influence intestinal histology and survival after irradiation. These humoral factors are thought to originate from the irradiated body tissues, the shielded proximal intestine, and the shielded distal intestine. Further studies are required to identify these factors and to determine their mode of action and their therapeutic potential after radiation damage to the small intestine.     

Intermediates in membrane fusion and bilayer/nonbilayer phase transitions imaged by time-resolved cryo-transmission electron microscopy.

Bilayer-to-nonbilayer phase transitions in phospholipids occur by means of poorly characterized intermediates. Many have proposed that membrane fusion can also occur by formation of these intermediates. Structures for such intermediates were proposed in a recent theory of these transition mechanisms. Using time-resolved cryo-transmission electron Microscopy (TRC-TEM), we have directly visualized the evolution of inverted phase micro-structure in liposomal aggregates. We have identified one of the proposed intermediates, termed an interlamellar attachment (ILA), which has the structure and dimensions predicted by the theory. We show that ILAs are likely to be the structure corresponding to "lipidic particles" observed by freeze-fracture electron microscopy. ILAs appear to assemble the inverted cubic (III) phase by formation of an ILA lattice, as previously proposed. ILAs are also observed to mediate membrane fusion in the same systems, on the same time scale, and under nearly the same conditions in which membrane fusion was observed by fluorescence methods in earlier studies. These earlier studies indicated a linkage between a membrane fusion mechanism and III phase formation. Our micrographs suggest that the same intermediate structure mediates both of those processes.     

Stoichiometry of estramustine phosphate binding to MAP2 measured by the disassembly of chick brain MAP2:tubulin microtubules

The concentration of estramustine phosphate required to inhibit the assembly or to induce the disassembly of chick brain MAP2:tubulin microtubules is markedly dependent upon the microtubule protein concentration. Analysis of this relationship shows that estramustine phosphate and tubulin compete for common MAP2 sites, that MAP2 can bind 5-6 moles.mole-1 estramustine phosphate, and that the Kd of these sites is congruent to 20 microM estramustine phosphate. It is proposed that two molecules of estramustine phosphate interact with each of the three tubulin-binding sites of MAP2 and inhibit the MAP2:tubulin interaction by neutralising two highly conserved basic residues.     

Missense mutation in the lacI gene of Escherichia coli. Inferences on the structure of the repressor protein

The lac repressor has been studied extensively but a precise three-dimensional structure remains unknown. Studies using mutational data can complement other information and provide insight into protein structure. We have been using the lacI gene-repressor protein system to study the mutational specificity of spontaneous and induced mutation. The sequencing of over 6000 lacI- mutations has revealed 193 missense mutations generating 189 amino acid replacements at 102 different sites within the lac repressor. Replacement sites are not distributed evenly throughout the protein, but are clustered in defined regions. Almost 40% of all sites and over one-half of all substitutions found occur within the amino-terminal 59 amino acid residues, which constitute the DNA-binding domain. The core domain (residues 60 to 360) is less sensitive to amino acid replacement. Here, substitution is found in regions involved in subunit aggregation and at sites surrounding residues that are implicated in sugar-binding. The distribution and nature of missense mutational sites directs attention to particular amino acid residues and residue stretches.     

Pertussis toxin and target eukaryotic cells: binding, entry, and activation.

Pertussis toxin, a protein virulence factor produced by Bordetella pertussis, is composed of an A protomer and a B oligomer. The A protomer consists of a single polypeptide, termed the S1 subunit, which disrupts transmembrane signaling by ADP-ribosylating eukaryotic G-proteins. The B oligomer, containing five polypeptides, binds to cell receptors (most likely containing carbohydrate) and delivers the S1 subunit. Current knowledge suggests that expression of ADP-ribosyltransferase activity in target eukaryotic cells arises after 1) nucleotides and membrane lipids allosterically promote the release of the S1 subunit; and 2) the single disulfide bond in the S1 subunit is reduced by reductants such as glutathione. This model suggests conditions for the proper use of the toxin as an experimental reagent.     

trans rescue of a mutant poliovirus RNA polymerase function.

A series of three-nucleotide insertions was engineered into the P2 and P3 coding regions of the T7 expression plasmid pT7(tau)-PV1, which encodes a full-length copy of poliovirus type 1 (Mahoney) cDNA. When RNA derived in vitro from these mutated templates was used to transfect HeLa cells, viable virus mutants were recovered. One mutant, Sel-3D-18, which contained a single amino acid insertion in the 3Dpol coding region, was temperature sensitive for growth at 39 degrees C and showed defects in both RNA synthesis and P1 protein processing at the nonpermissive temperature. The RNA replication defect in Se1-3D-18 was identified at the level of RNA chain elongation. A highly specific and sensitive method was developed for analyzing the ability of mutant RNA templates to replicate in the presence or absence of helper functions provided in trans. This approach was used to demonstrate that RNA synthesis in Se1-3D-18 can be rescued by helper functions provided in trans.     

Effect of microspore stage and media on anther culture of peanut (Arachis hypogaea L.)

This study was designed to study the effects of stage of microspore development and culture medium on androgenic response in peanut (Arachis hypogaea L.). Anthers of various developmental stages were cultured for 7 days, then fixed and observed cytologically. Three sets of media, involving different basal media, growth regulators, sucrose levels and glutamine concentrations, were tested. In all experiments, the stage of development of the microspores at the time of culture was highly significant. The early uninucleate microspores stage was identified as producing the highest anther response rating. The effect of media was nonsignificant in all experiments. However, the stepwise modification of the media through the course of the study resulted in an almost 8 x increase in anther response rating. Numerically, the best media tested was N₆ basal medium with 1 mg 1^-1 NAA, 0.1 mg 1^-1 BA, 5.5% sucrose, and 3.5 g 1^-1 glutamine. While no haploids were obtained, four-nucleate cells were observed, indicating the potential in peanuts for an androgenic reponse.     

A comparative assessment of purification techniques for mesophyll protoplasts of Brassica napus L.

The effects of three different general purification protocols have been assessed quantitatively using mesophyll protoplasts of Brassica napus. Within the initial sample two distinct sub-populations were determined. The methods used influenced the ratio of the vacuolated to chloroplastic type protoplast sub-populations. Overall recovery rates of the initial sample varied according to the method used from 38% to 27%, but the relative recovery of the sub-populations varied considerably with a purified ratio of between 1.0:0.78 to 1.0:7.0. Size distribution profiles of the initial and purified populations are also presented.     

A survey of hospital toilet facilities.

OBJECTIVE--To assess the quality of toilet facilities available for disabled people in a large provincial teaching hospital. DESIGN--Survey of toilet facilities for patients on the wards and in the outpatient department. SETTING--Teaching hospital in Leeds. RESULTS--Although the quality of toilet facilities varied, none met the standards recommended by the British Standards Institution. The worst facilities were found on a ward accommodating elderly patients, where the toilets were unsuitable for use by disabled people and bedside commodes had to be used instead. CONCLUSION--Toilet provision within a major hospital failed to meet standards required for disabled people. Admission to hospital may therefore result in loss of independence and dignity. If hospitals are to be centres of excellence, greater consideration must be given to the requirements of disabled people in the design of new wards, and current inadequate facilities should be upgraded.