Effect of fertility regulating agents on motility and zona-free hamster egg penetration by spermatozoa of bonnet monkey.

Administration of STS-557 (17 alpha-cyanomethyl-17 beta-hydroxyestra 4,9(10)-dien-3-one; 12 mg/monkey daily) for 4 weeks either alone or in combination with 20 Aet-1 (testosterone-trans-4-n-butyl cyclohexyl carboxylate; code CDB 1781; 40 mg/monkey single administration) had no significant effect on motility and zona free hamster egg penetration by spermatozoa of bonnet monkey, but continuation of the treatment for 12 weeks reduced (in one monkey treated with STS-557) or abolished (one treated with STS-557 and two with STS-557 + 20 Aet-1) the motility as well as zona-free hamster egg penetration (by spermatozoa of all treated monkeys). Motility and the ability to penetrate zona-free hamster egg returned to normalcy after 10 weeks of withdrawal of treatments. Active immunization of monkeys with ovine FSH (4 weeks after booster) had no adverse effect on motility of spermatozoa but none of the zona-free hamster eggs was fertilized. The correlation between motility and the capacity to penetrate the zona-free hamster eggs by monkey spermatozoa varies with the treatment. Such correlation was apparent in monkeys treated with STS-557 but not in monkeys immunized with ovine FSH.     

Changes in Arabidopsis leaf ultrastructure, chlorophyll and carbohydrate content during spaceflight depend on ventilation

Leaf structure and function under spaceflight conditions have received little study despite their important implications for biological life support systems using plants. Previous reports described disruption of the membrane apparatus for photosynthesis and a general decrease in carbohydrate content in foliage. During a series of three short-duration experiments (Chromex-03, -04, -05) on the US space shuttle (STS-54, STS-51, STS-68), we examined Arabidopsis thaliana leaves. The plants were at the rosette stage at the time of loading onto the space shuttle, and received the same light, temperature, carbon dioxide and humidity regimes in the orbiter as in ground controls. The experiments differed according to the regime provided in the headspace around the plants: this was either sealed (on mission STS-54); sealed with high levels of carbon dioxide (on mission STS-51) or vented to the cabin air through a filtration system (on mission STS-68). Immediately post-flight, leaf materials were fixed for microscopy or frozen in liquid nitrogen for subsequent analyses of chlorophyll and foliar carbohydrates. At the ultrastructural level, no aberrations in membrane structure were observed in any of the experiments. When air-flow was provided, plastids developed large starch grains in both spaceflight and ground controls. In the experiments with sealed chambers, spaceflight plants differed from ground controls with regard to measured concentrations of carbohydrate and chlorophyll, but the addition of airflow eliminated these differences. The results point to the crucial importance of consideration of the foliage microenvironment when spaceflight effects on leaf structure and metabolism are studied.     

Predictive Validity of the Suicide Trigger Scale (STS-3) for Post-Discharge Suicide Attempt in High-Risk Psychiatric Inpatients

Background

The greatly increased risk of suicide after psychiatric hospitalization is a critical problem, yet we are unable to identify individuals who would attempt suicide upon discharge. The Suicide Trigger Scale v.3 (STS-3), was designed to measure the construct of an affective ‘suicide trigger state’ hypothesized to precede a suicide attempt (SA). This study aims to test the predictive validity of the STS-3 for post-discharge SA on a high-risk psychiatric-inpatient sample.

Methods

The STS-3, and a psychological test battery measuring suicidality, mood, impulsivity, trauma history, and attachment style were administered to 161 adult psychiatric patients hospitalized following suicidal ideation (SI) or SA. Receiver Operator Characteristic and logistic regression analyses were used to assess prediction of SA in the 6-month period following discharge from hospitalization.

Results

STS-3 scores for the patients who made post-discharge SA followed a bimodal distribution skewed to high and low scores, thus a distance from median transform was applied to the scores. The transformed score was a significant predictor of post-discharge SA (AUC 0.731), and a subset of six STS-3 scale items was identified that produced improved prediction of post-discharge SA (AUC 0.814). Scores on C-SSRS and BSS were not predictive. Patients with ultra-high (90^th percentile) STS-3 scores differed significantly from ultra-low (10^th percentile) scorers on measures of affective intensity, depression, impulsiveness, abuse history, and attachment security.

Conclusion

STS-3 transformed scores at admission to the psychiatric hospital predict suicide attempts following discharge among the high-risk group of suicidal inpatients. Patients with high transformed scores appear to comprise two clinically distinct groups; an impulsive, affectively intense, fearfully attached group with high raw STS-3 scores and a low-impulsivity, low affect and low trauma-reporting group with low raw STS-3 scores. These groups may correspond to low-plan and planned suicide attempts, respectively, but this remains to be established by future research.     

NASDA aquatic animal experiment facilities for Space Shuttle.

National Space Development Agency of Japan (NASDA) has been developed aquatic animal experiment facilities for space experiments using NASA Space Shuttle. Vestibular Function Experiment Unit (VFEU) has been firstly designed and developed for Spacelab-J mission (STS-47), and 8 days space experiment with carp has been performed. Following, the VFEU, Aquatic Animal Experiment Unit (AAEU) has been developed to accommodate small aquatic animals second International Microgravity Laboratory mission (IML-2, STS-65). Four kinds of space experiments with goldfish, medaka, newt, and newt eggs have been performed for 15 days mission duration. Then, VFEU has been improved to accommodate marine fish under low temperature condition for Neurolab (STS-90) and STS-95 missions. 17 days (STS-90) and 9 days (STS-95) experiments with oyster toadfish have been performed by using the VFEU. This report summarizes the outline of these aquatic animal experiment facilities.     

The Fluid Processing Apparatus: from flight hardware to electron micrographs

Sweet clover seeds were grown in the Fluid Processing Apparatus (FPA) flown on STS-54 (January 1993) and STS-60 (February 1994). After germination and seedling growth for 40 hours, seedlings were fixed in space. Electron microscopy was used to examine the seedling ultrastructure to verify that fixation occurred and preserved the samples. Micrographs revealed well-preserved cell structure and the presence of calcium precipitates indicating complete fixation. FPA is a useful piece of hardware for preservation/fixation during spaceflight.     

Emergency Room Validation of the Revised Suicide Trigger Scale (STS-3): A Measure of a Hypothesized Suicide Trigger State

Background

The Suicide Trigger Scale (STS) was designed to measure the construct of an affective ‘suicide trigger state.’ This study aims to extend the inpatient setting validation study of the original Suicide Trigger Scale version 2 to the revised Suicide Trigger Scale version 3 (STS-3) in an acute psychiatric emergency room setting.

Methods

The 42-item STS-3 and a brief psychological test battery were administered to 183 adult psychiatric patients with suicidal ideation or attempt in the psychiatric emergency room, and re-administered to subjects at 1 year follow up. Factor analysis, linear and logistic regressions were used to examine construct structure, divergent and convergent validity, and construct validity, respectively.

Results

The STS-3 demonstrated strong internal consistency (Cronbach’s alpha 0.94). Factor analysis yielded a three-factor solution, which explained 43.4% of the variance. Principal axis factor analysis was used to identify three reliable subscales: Frantic Hopelessness, Ruminative Flooding, and Near-Psychotic Somatization (Cronbach’s alphas 0.90, 0.80, and 0.76, respectively). Significant positive associations were observed between Frantic Hopelessness and BSI depression and anxiety subscales, between Ruminative Flooding and BSI anxiety and paranoia subscales, and Near Psychotic Somatization and BSI somatization subscales. Suicidal subjects with suicide attempt history had mean scores 7 points higher than those without history of suicide attempts. Frantic hopelessness was a significant predictor of current suicide attempt when only attempts requiring at least some medical attention were considered.

Conclusion

The STS-3 measures a distinct clinical entity, provisionally termed the ‘suicide trigger state.’ Scores on the STS-3 or select subscales appear to relate to degree of suicidality in terms of severity of ideation, history of attempt, and presence of substantive current attempts. Further study is required to confirm the factor structure and better understand the nature of these relations.     

Reproduction during spaceflight by plants in the family Brassicaceae

Researchers report on studies of reproduction in Arabidopsis thaliana in space during during the Chromex-03 on STS-54, Chromex-04 on STS-51, and Chromex-05 on STS-68 missions. The obstacles to seed formation were related to carbon dioxide levels. Other experiments examined in flight pollination and seed production in Brassica rapa during parabolic flight, a 4-1/2 month stay on Mir, and on STS-87. During the Mir experiment, Brassica seeds were harvested from seeds sown in flight. The second generation seeds grew to produce new seeds that contained more starch and less protein and lipid when compared to ground control seeds.     

STS-95 Space Experiments (plants and cell biology).

We report the outline of Space Experiments conducted on Space Shuttle (STS-95) launched in autumn of 1998. In this STS-95 mission, Japanese astronaut Dr. Chiaki Mukai achieved her 2nd space flight and conducted a part of 82 space experiments including Japanese experiments. US astronaut Senator John Glenn also achieved his second space flight, 36 years after his first space flight. Senator Glenn was a leader of the original (the first) 7 US astronauts and very famous in US because he succeeded US first orbital space flight around the earth. NASDA had started the project of space experiment using STS-95 at the summer of 1997, therefore we had only one year for the all preparation Yamashita, et al. Biological Sciences in Space, Vol.12 No.3(1998). Scientific results will be reported by investigators, therefore we report here how we had been developing the space experiment plan, on board operation procedure and ground operations including ground control experiments about four plant experiments and one cell biology experiment.     

Crystallization of biological macromolecules from flash frozen samples on the Russian Space Station Mir

One hundred eighty-three flash frozen, liquid-liquid diffusion and batch method protein and virus crystallization samples were launched aboard the Space Shuttle Discovery on June 27 (STS-71) and transferred to the Russian Space Station Mir on July 1, 1995. They were returned to earth November 20, 1995 (STS-74). Subsequent examination showed that of the 19 types of proteins and viruses investigated, 17 were crystallized during the period on Mir. The experiment demonstrates the utility of this very simple and inexpensive approach for the crystallization of biological macromolecules in space over extended time periods. The distribution of crystals among the three types of containers used indicated small samples yielded results equal or better than larger samples and that long diffusion path lengths were clearly better. Distribution of crystals within the container tubes showed a striking gradient of quality and size that indicated long, narrow tubes yield superior crystals, as predicted from other work based on crystallization in capillaries.     

Pollen and ovule development in Arabidopsis thaliana under spaceflight conditions

The development of pollen and ovules in Arabidopsis thaliana on the space shuttle 'Endeavour' (STS-54) was investigated. Plants were grown on nutrient agar for 14 days prior to loading into closed plant growth chambers that received light and temperature control inside the Plant Growth Unit flight hardware on the shuttle middeck. After 6 days in spaceflight the plants were retrieved and immediately dissected and processed for light and electron microscope observation. Reproductive development aborted at an early stage. Pistils were collapsed and ovules inside were seen to he empty. No viable pollen was observed from STS-54 plants; young microspores were deformed and empty. At a late stage, the cytoplasm of the pollen contracted and became disorganized, but the pollen wall developed and the exine appeared normal. The tapetum in the flight flowers degenerated at early stages. Ovules from STS-54 flight plants stopped growing and the integuments and nucellus collapsed and degenerated. The megasporocytes appeared abnormal and rarely underwent meiosis. Apparently they enlarged, or occasionally produced a dyad or tetrad, to assume the form of a female gametophyte with the single nucleus located in an egglike cell that lacks a cell wall. Synergids, polar nuclei, and antipodals were not observed. The results demonstrate the types of lesions occurring in plant reproductive material under spaceflight conditions.     

Genetic heterogeneity of steroid sulfatase deficiency revealed with cDNA for human steroid sulfatase

Three cDNA clones with inserts of 1.2-1.6 kb that reacted both with antibodies and oligonucleotides specific for steroid sulfatase were isolated from a human placental library in lambda gt11. The 5'-end of one of the inserts, STS-3, was sequenced and colinearity with the amino acid sequence of 3 peptides of steroid sulfatase encompassing 64 amino acids was demonstrated. STS-3 hybridized with 2.5, 4.6 and 6.3 kb species in poly(A)+RNA and with 2.5, 4 and 9 kb fragments of EcoRI digested human DNA. The frequency of the EcoRI fragments in DNA from females was approximately twice that in DNA from males. DNA from two patients with steroid sulfatase deficiency and X-linked ichthyosis did not hybridize with STS-3. DNA from a third patient showed a normal hybridization pattern. It is concluded that steroid sulfatase deficiency is a genetically heterogenous disorder.     

TULA-family proteins: Jacks of many trades and then some

UBASH3/STS/TULA is a novel two-member family, which exerts several key regulatory effects in multiple cell types. UBASH3B/STS-1/TULA-2 is a highly active protein tyrosine phosphatase; its major target appears to be a specific regulatory site of protein tyrosine kinases of the Syk family, dephosphorylation of which inhibits Syk and Zap-70 kinases and suppresses receptor signaling mediated by these kinases. UBASH3A/STS-2/TULA exhibits substantial homology to UBASH3B/STS-1/TULA-2, but possesses only a small fraction of phosphatase activity of UBASH3B/STS-1/TULA-2, and thus, its regulatory effect may be based also on the phosphatase-independent mechanisms. Critical physiologic effects of these proteins have been demonstrated in T lymphocytes, platelets, stem cells, and other important cell types. These proteins have also been shown to play a key role in such pathologic conditions as autoimmunity, cancer, and thrombosis. The review focuses on the recent studies of this important family of cellular regulators.     

Clinorotation affects soybean seedling morphology

Although spaceflight does not appear to significantly affect seed germination, it can influence subsequent plant growth. On STS-3 and SL-2, decreased growth (measured as plant length, fresh weight and dry weight) was noted for pine, oat and mung bean. In the CHROMEX-01 and -02 experiments with Haplopappus and in the CHROMEX-03 experiment with Arabidopsis, enhanced root growth was noted in the space-grown plants. In the CHROMEX-04 experiment with wheat, both leaf fresh weight and leaf area were diminished in the space-grown plants but there was no difference in total plant height (CS Brown, HG Levine, and AD Krikorian, unpublished data). These data suggest that microgravity impacts growth by whole plant partitioning of assimilates. The objective of the present study was to determine the influence of clinorotation on the growth and morphology of soybean seedlings grown in the BRIC (Biological Research In Canister) flight hardware. This experiment provided baseline data for a spaceflight experiment (BRIC-03) flown on STS-63 (Feb. 3-11, 1995).     

Once phosphorylated, tyrosines in carboxyl terminus of protein-tyrosine kinase Syk interact with signaling proteins, including TULA-2, a negative regulator of mast cell degranulation

Activation of the high affinity IgE-binding receptor (FcεRI) results in the tyrosine phosphorylation of two conserved tyrosines located close to the COOH terminus of the protein-tyrosine kinase Syk. Synthetic peptides representing the last 10 amino acids of the tail of Syk with these two tyrosines either nonphosphorylated or phosphorylated were used to precipitate proteins from mast cell lysates. Proteins specifically precipitated by the phosphorylated peptide were identified by mass spectrometry. These included the adaptor proteins SLP-76, Nck-1, Grb2, and Grb2-related adaptor downstream of Shc (GADS) and the protein phosphatases SHIP-1 and TULA-2 (also known as UBASH3B or STS-1). The presence of these in the precipitates was further confirmed by immunoblotting. Using the peptides as probes in far Western blots showed direct binding of the phosphorylated peptide to Nck-1 and SHIP-1. Immunoprecipitations suggested that there were complexes of these proteins associated with Syk especially after receptor activation; in these complexes are Nck, SHIP-1, SLP-76, Grb2, and TULA-2 (UBASH3B or STS-1). The decreased expression of TULA-2 by treatment of mast cells with siRNA increased the FcεRI-induced tyrosine phosphorylation of the activation loop tyrosines of Syk and the phosphorylation of phospholipase C-γ2. There was parallel enhancement of the receptor-induced degranulation and activation of nuclear factor for T cells or nuclear factor κB, indicating that TULA-2, like SHIP-1, functions as a negative regulator of FcεRI signaling in mast cells. Therefore, once phosphorylated, the terminal tyrosines of Syk bind complexes of proteins that are positive and negative regulators of signaling in mast cells.     

Gravitropic moss cells default to spiral growth on the clinostat and in microgravity during spaceflight

In addition to shoots and roots, the gravity (g)-vector orients the growth of specialized cells such as the apical cell of dark-grown moss protonemata. Each apical cell of the moss Ceratodon purpureus senses the g-vector and adjusts polar growth accordingly producing entire cultures of upright protonemata (negative gravitropism). The effect of withdrawing a constant gravity stimulus on moss growth was studied on two NASA Space Shuttle (STS) missions as well as during clinostat rotation on earth. Cultures grown in microgravity (spaceflight) on the STS-87 mission exhibited two successive phases of non-random growth and patterning, a radial outgrowth followed by the formation of net clockwise spiral growth. Also, cultures pre-aligned by unilateral light developed clockwise hooks during the subsequent dark period. The second spaceflight experiment flew on STS-107 which disintegrated during its descent on 1 February 2003. However, most of the moss experimental hardware was recovered on the ground, and most cultures, which had been chemically fixed during spaceflight, were retrieved. Almost all intact STS-107 cultures displayed strong spiral growth. Non-random culture growth including clockwise spiral growth was also observed after clinostat rotation. Together these data demonstrate the existence of default non-random growth patterns that develop at a population level in microgravity, a response that must normally be overridden and masked by a constant g-vector on earth.     

Development of the brine shrimp Artemia is accelerated during spaceflight

Developmentally arrested brine shrimp cysts have been reactivated during orbital spaceflight on two different Space Shuttle missions (STS-50 and STS-54), and their subsequent development has been compared with that of simultaneously reactivated ground controls. Flight and control brine shrimp do not significantly differ with respect to hatching rates or larval morphology at the scanning and transmission EM levels. A small percentage of the flight larvae had defective nauplier eye development, but the observation was not statistically significant. However, in three different experiments on two different flights, involving a total of 232 larvae that developed in space, a highly significant difference in degree of flight to control development was found. By as early as 2.25 days after reactivation of development, spaceflight brine shrimp were accelerated, by a full instar, over ground control brine shrimp. Although developing more rapidly, flight shrimp grew as long as control shrimp at each developmental instar or stage.     

TULA proteins regulate activity of the protein tyrosine kinase Syk

TULA belongs to a two-member family: TULA (STS-2) is a lymphoid protein, whereas STS-1/TULA-2 is expressed ubiquitously. TULA proteins were implicated in the regulation of signaling mediated by protein tyrosine kinases (PTKs). The initial experiments did not fully reveal the molecular mechanism of these effects, but suggested that both TULA proteins act in a similar fashion. It was shown recently that STS-1/TULA-2 dephosphorylates PTKs. In this study, we analyzed the effects of TULA proteins on Syk, a PTK playing an important role in lymphoid signaling. First, we have shown that TULA-2 decreases tyrosine phosphorylation of Syk in vivo and in vitro and that the intact phosphatase domain of TULA-2 is essential for this effect. We have also shown that TULA-2 exhibits a certain degree of substrate specificity. Our results also indicate that inactivated TULA-2 increases tyrosine phosphorylation of Syk in cells co-transfected to overexpress these proteins, thus acting as a dominant-negative form that suppresses dephosphorylation of Syk caused by endogenous TULA-2. Furthermore, we have demonstrated that phosphatase activity of TULA is negligible as compared to that of TULA-2 and that this finding correlates with an increase in Syk tyrosine phosphorylation in cells overexpressing TULA. This result is consistent with the dominant-negative effect of inactivated TULA-2, arguing that TULA acts in this system as a negative regulator of TULA-2-dependent dephosphorylation. To summarize, our findings indicate that TULA proteins may exert opposite effects on PTK-mediated signaling and suggest that a regulatory mechanism based on this feature may exist.     

Brine shrimp development in space: ground-based data to shuttle flight results

The brine shrimp, Artemia salina, has been used as a model system to assess microgravity effects on developing organisms. Following fertilization and early development, the egg can arrest in early gastrula as a dehydrated cyst stage that is stable to harsh environments over long time periods. When salt water is added, the cysts can reactivate, with embryonic development and egg hatching occurring in about 24 h. A series of larval molts or instars, over about a 2 week period, results in the adult crustacean. We have assessed these developmental events in a closed syringe system, a bioprocessing module, in ground-based studies, and have conducted preliminary in-orbit experiments aboard the Space Shuttle Atlantis during the flights of STS-37 and STS-43. Although the in-flight data are limited, spectacular degrees of development have been achieved.     

Cleavage of Bax is mediated by caspase-dependent or -independent calpain activation in dopaminergic neuronal cells: protective role of Bcl-2.

Two cysteine protease families, caspase and calpain, are known to participate in cell death. We investigated whether a stress-specific protease activation pathway exists, and to what extent Bcl-2 plays a role in preventing drug-induced protease activity and cell death in a dopaminergic neuronal cell line, MN9D. Staurosporine (STS) induced caspase-dependent apoptosis while a dopaminergic neurotoxin, MPP(+) largely induced caspase-independent necrotic cell death as determined by morphological and biochemical criteria including cytochrome c release and fluorogenic caspase cleavage assay. At the late stage of both STS- and MPP(+)-induced cell death, Bax was cleaved into an 18-kDa fragment. This 18-kDa fragment appeared only in the mitochondria-enriched heavy membrane fraction of STS-treated cells, whereas it was detected exclusively in the cytosolic fraction of MPP(+)-treated cells. This proteolytic cleavage of Bax appeared to be mediated by calpain as determined by incubation with [(35)S]methionine-labelled Bax. Thus, cotreatment of cells with calpain inhibitor blocked both MPP(+)- and STS-induced Bax cleavage. Intriguingly, overexpression of baculovirus-derived inhibiting protein of caspase, p35 or cotreatment of cells with caspase inhibitor blocked STS- but not MPP(+)-induced Bax cleavage. This appears to indicate that calpain activation may be either dependent or independent of caspase activation within the same cells. However, cotreatment with calpain inhibitor rescued cells from MPP(+)-induced but not from STS-induced neuronal cell death. In these paradigms of dopaminergic cell death, overexpression of Bcl-2 prevented both STS- and MPP(+)-induced cell death and its associated cleavage of Bax. Thus, our results suggest that Bcl-2 may play a protective role by primarily blocking drug-induced caspase or calpain activity in dopaminergic neuronal cells.     

A comparison of depth dependence of dose and linear energy transfer spectra in aluminum and polyethylene

A set of four tissue-equivalent proportional counters (TEPCs), with their detector heads at the centers of 0 (bare), 3, 7 and 9-inch-diameter aluminum spheres, were flown on Shuttle flight STS-89. Five such detectors at the centers of polyethylene spheres were flown 1 year earlier on STS-81. The results of dose-depth dependence for the two materials convincingly show the merits of using material rich in hydrogen to decrease the radiation exposure to the crew. A comparison of the calculated galactic cosmic radiation (GCR) absorbed dose and dose-equivalent rates using the radiation transport code HZETRN with nuclear fragmentation model NUCFRG2 and the measured GCR absorbed dose rates and dose-equivalent rates shows that they agree within root mean square (rms) error of 12.5 and 8.2%, respectively. However, there are significant depth-dependent differences in the linear energy transfer (LET) spectra. A comparison for trapped protons using the proton transport code BRYNTRN and the AP-8 MIN trapped-proton model shows a systematic bias, with the model underpredicting dose and dose-equivalent rates. These results show the need for improvements in the radiation transport and/or fragmentation models.