Implementation of ALARA radiation protection on the ISS through polyethylene shielding augmentation of the Service Module Crew Quarters.

With 5-7 month long duration missions at 51.6 degrees inclination in Low Earth Orbit, the ionizing radiation levels to which International Space Station (ISS) crewmembers are exposed will be the highest planned occupational exposures in the world. Even with the expectation that regulatory dose limits will not be exceeded during a single tour of duty aboard the ISS, the "as low as reasonably achievable" (ALARA) precept requires that radiological risks be minimized when possible through a dose optimization process. Judicious placement of efficient shielding materials in locations where crewmembers sleep, rest, or work is an important means for implementing ALARA for spaceflight. Polyethylene (CnHn) is a relatively inexpensive, stable, and, with a low atomic number, an effective shielding material that has been certified for use aboard the ISS. Several designs for placement of slabs or walls of polyethylene have been evaluated for radiation exposure reduction in the Crew Quarters (CQ) of the Zvezda (Star) Service Module. Optimization of shield designs relies on accurate characterization of the expected primary and secondary particle environment and modeling of the predicted radiobiological responses of critical organs and tissues. Results of the studies shown herein indicate that 20% or more reduction in equivalent dose to the CQ occupant is achievable. These results suggest that shielding design and risk analysis are necessary measures for reducing long-term radiological risks to ISS inhabitants and for meeting legal ALARA requirements. Verification of shield concepts requires results from specific designs to be compared with onboard dosimetry.

Co-localization of cell surface receptors at high spatial resolution by single-particle fluorescence imaging.

Dual-wavelength single-particle fluorescence imaging has been used to quantify the co-localization of receptors and/or ligands on cells by widefield microscopy. Methods for correction of chromatic aberration and identification of submicroscopic artefacts are presented, with data for the lipopolysaccharide/CD14 and MHC class II/CD74 systems.

Measurements of associations of cell-surface receptors by single-particle fluorescence imaging.

SPFI (single-particle fluorescence imaging) uses the high sensitivity of fluorescence to visualize individual molecules that have been selectively labelled with small fluorescent particles. The images of particles are diffraction-limited spots that are analysed by fitting with a two-dimensional Gaussian function. The spot intensities depend on whether they arise from one or more particles; this provides the basis for determining self-association of cell-surface receptors. We have used this approach to determine dimerization of MHC class II molecules and its disruption by interface peptides. We have also exploited the positional information obtained from SPFI to detect co-localization of cell-surface molecules. This involves labelling two different molecules with different coloured fluorophores and determining their positions separately by dual wavelength imaging. The images are analysed to quantify the overlap of the particle images and hence determine the extent of co-localization of the labelled molecules. The technique provides quantification of the extent of co-localization and can detect whether co-localized molecules occur singly or in clusters. We have obtained preliminary data for co-localization of lipopolysaccharide and CD14 on intact cells. We also show that HLA-DR (human leukocyte antigen-DR) and CD74 are partially co-localized and that interaction between these molecules involves the peptide-binding groove of HLA-DR.

Neutropenia is a risk factor for gram-negative bacillus bacteremia in human immunodeficiency virus-infected patients: results of a nested case-control study.

A previous cohort study demonstrated a relation between neutropenia and bacteremia due to gram-negative bacilli among patients infected with human immunodeficiency virus (HIV). To explore further the relation between neutropenia and bacteremia due to Escherichia coli, Klebsiella pneumoniae, or Pseudomonas aeruginosa among HIV-infected patients, controlling for confounding factors, the authors conducted a nested case-control study with matching and risk-set sampling of controls. The cohort included 1,645 HIV-infected patients followed at the University of California, San Diego, Medical Center in San Diego, California, between 1991 and 1995. Absolute neutrophil count (ANC) was summarized as mean ANC during the 7-day interval preceding the index date of bacteremia. Covariates were ascertained by medical record review. The matching ratio was 6:1 (controls:cases). Odds ratios were estimated using conditional logistic regression. Forty-four incident cases of bacteremia were identified. After adjustment for covariates, the estimated odds ratio for the effect of neutropenia (ANC=500 vs. >500/microl) during the 7 days preceding the index date was 8.1 (95% confidence interval confidence interval 1.5-43.1). The rate of bacteremia due to E. coli, K. pneumoniae, or P. aeruginosa is increased eightfold if the average current-week ANC is less than or equal to 500/microl compared with more than 500/microl.

Estimation of the effect of neutropenia on rates of clinical bacteraemia in HIV-infected patients.

A retrospective cohort study was conducted to quantitate the relationship between neutropenia and rates of clinical bacteraemia among adults with HIV infection receiving medical care at one institution between 1991-5. The primary exposure, absolute neutrophil count (ANC), was summarized as mean ANC within a given week, using a five-level stratification (reference > 1000/microl). ANC stratum-specific rates of bacteraemia were calculated, by organism type. Linear trend tests were performed to assess dose-response relationship between neutropenia and rates of bacteraemia. The cohort included 1645 patients contributing 26,785 patients-weeks and 191 episodes of bacteraemia. The unadjusted effect of neutropenia was most evident for bacteraemia due to E. coli (RR 3.4), Klebsiella pneumoniae (RR 16.7), and P. aeruginosa (RR 10.4). For bacteraemia due to any of these three organisms (47 episodes), with reference ANC > 1000/microl, relative rates were: 751-1000/microl, 1.12; 501-750/microl, 2.11; 251-500/microl, 13.58; 0-250/microl, 21.89.

Partial characterization of the C-terminal non-collagenous domain (NC1) of collagen type X.

Collagen type X is composed of three identical alpha 1(X) chains of 59 kDa, each containing a triple-helical region of 45 kDa flanked by a short N-terminal sequence and a larger non-collagenous C-terminal (NC1) domain of approx. 15 kDa. Collagen type X molecules can associate via their C-termini to form a regular hexagonal lattice in vitro, which in vivo may provide a modified extracellular matrix for the events of endochondral ossification. The NC1 domain of chick collagen type X was isolated and purified from a highly purified bacterial collagenase digest of hypertrophic chondrocyte medium proteins. The structure and aggregation properties of the NC1 domain of collagen X were investigated, independently of the triple helix. A trimer, a dimer and a monomer of the individual alpha-chain NC1 polypeptides were identified from a bacterial collagenase digest of cartilage collagens using [14C]tyrosine labelling, N-chlorosuccinimide peptide mapping and N-terminal sequencing. The trimer (50 kDa) remained intact in Laemmli sample buffer unless boiled, upon which it dissociated into the dimer (38 kDa) and the monomer (20 kDa). The dimer persisted even after prolonged periods of heating or reduction with beta-mercaptoethanol, and in preparations obtained from chondrocyte cultures treated with beta-aminoproprionitrile, indicating the presence of non-reducible, non-lysine-derived, covalent cross-links. Hexamers of the individual C-termini were observed in rotary-shadowed preparations of purified NC1 domain, reflecting the ability of collagen type X to self-assemble via its C-termini under appropriate conditions.

Expression and distribution of cartilage matrix macromolecules in avian tibial dyschondroplasia.

Tibial dyschondroplasia (TD) is a disorder of endochondral ossification characterized by the presence of an avascular, non-mineralized cartilage lesion extending from the growth plate into the metaphysis. Cells within the TD growth plate fail to differentiate to full hypertrophy, and instead appear to maintain a 'pre-hypertrophic' or 'transitional' status. The synthesis and distribution of aggrecan, biglycan, decorin, and collagen types II and X in the growth plates of normal and tibial dyschondroplastic chickens have been investigated using in situ hybridization and immunolocalization. Marked reductions in the amount of aggrecan and biglycan core protein mRNAs were observed in the tibial dyschondroplastic lesion by in situ hybridization. Reduction in mRNA production seemed to be specific to the extracellular matrix components since total mRNA expression showed no significant difference between normal and dyschondroplastic cartilage. In addition, expression of collagen type II and decorin did not differ significantly between normal and TD cartilage. Distribution of aggrecan biglycan, decorin, type II and X collagens were examined using immunohistochemistry. Normal hypertrophic cartilage showed a strong matrix labelling for aggrecan and biglycan. Type X collagen in the normal hypertrophic cartilage showed strong pericellular and matrix distribution, whereas in TD cartilage labelling for aggrecan, biglycan and collagen X was located intracellularly with a very low level of signal in the matrix. In contrast, collagen type II was found to be present throughout the extracellular matrix of both the normal growth plate and the TD lesion, suggesting that the differences observed in aggrecan, biglycan and type X collagen distribution are specific to these proteins and not a general disturbance of matrix macromolecular metabolism. The reduced deposition of these macromolecules may have implications in normal and pathological bone development.

Electroanesthesia (EA) studies: current applications to human volunteers to produce general and local anesthesia.

Electrical currents found to be of sufficient intensity to produce EA in animals were applied to human subjects several hundred times, to determine whether, and how, clinical general anesthesia, and local anesthesia in various parts of the body, could be obtained. General anesthesia was not produced in any subject in any test, the obstacle in every instance being pain. Local analgesia of the arm was obtained in one subject, but in all other subjects muscle spasm and vibration pain prevented application of enough current to produce analgesia or anesthesia in the arm. Anesthesia of the hand was produced several times in all subjects, with a total loss of pain sensation.

A serum enzyme test to detect pulmonary capillary injury.

Lung converting enzyme (LCE) was extracted from normal human lung obtained at autopsy. The specificity of the enzyme preparation was confirmed in vitro by incubation with Angiotensin I (AI) and paper chromatographic identification of reaction products. A method was developed for assay of the enzyme preparation in human serum using tritiated Angiotensin I (H-3A-I) as substrate. Enzyme activity is quantitated by scintillation counting the radioactive end product tritiated dipeptide histadylleucine (H-3His-leu). Serum from the authors and pooled serum from the hospital laboratory caused less than 3.5% conversion of H-3A-I to H-3His-leu. Serum from two patients with severe lung damage caused a maximum 20% conversion of H-3A-I to H-3His-leu. Percent conversion in these patients correlated with clinical and laboratory evidence of lung dysfunction.