The effect of anti-coagulant and anti-platelet agents on testicular arterial blood flow following DXR treatment.

<p>Blood flow was measured by the PW Doppler mode and quantified by analyzing the VTI using the appropriate VisualSonics software. (<b>a</b>) Blood flow of testicular arteries was continuously monitored by PW Doppler in mice pre-treated with LMWH (clexane; 100µg/mouse), before and following DXR (8mg/kg BW; n=8, n=number of imaged arteries) injection and analyzed 3, 6, 9, 12 and 15 minutes afterwards. Graphic illustration of testicular arterial blood flow; points represent percent of control (dashed line; mean±SEM), (*)-significantly different from DXR treatment values (P<0.05). (<b>b</b>) Blood flow of testicular arteries was continuously monitored by PW Doppler in mice pre-treated with eptifibatide (integrilin; 75µg/mouse), before and following DXR (8mg/kg BW; n=9, n=number of imaged arteries) injection and analyzed 3, 6, 9, 12 and 15 minutes afterwards. Graphic illustration of testicular arterial blood flow; points represent percent of control (dashed line; mean±SEM), (*)-significantly different from DXR treatment values (P<0.05).</p

Effect of Doxorubicin on platelet aggregation and adhesion.

<p>(A) PRP was pre-incubated for 15 minutes with increasing concentrations of DXR then aggregation was induced by ADP (5 µM) and maximal aggregation is presented as mean ± SD; statistical analysis was tested by ANOVA (p<0.001; n=9). (B) Whole blood was pre-incubated for 15 minutes with increasing concentrations of DXR then subjected to the Impact-R test and both surface coverage and average size of the aggregates are presented as mean ± SD (n=5).</p

DXR induces testicular vascular changes.

<p>Histological sections of testes from saline or DXR (5 mg/kg) treated mice were immunohistochemically stained with Hoecst 33342 (nuclei labeling; blue) and anti CD34 primary antibody (1:200) followed by alexa555 anti-rat secondary antibody (red; 1:400). Saline (A,C) and DXR (B,D) at one week (A,B) or one month (C,D) after treatment. Bar=240µm.</p

Platelet adhesion to DXR-treated aortic endothelial cells.

<p>Confluent endothelial cells were exposed to growth medium without (A) or with Doxorubicin (B; 100 µM) for 4 hr followed by exposure to whole blood for 5 minutes at 37°C under defined shear rates (750 s^-1) and then fixed and platelets were stained by immunohistochemistry kit using monoclonal antibody against the platelet integrin CD41a.</p

Schematic diagram of the experiments.

<p>Mice of SF, SFV and backup SF groups were co-adapted in housing groups of 3 mice each, SF mice were adapted to paste food diet, and mice of <i>in vivo</i> subgroups passed through preliminary tests. After transportation to and adaptation at Baikonur, SF mice were flown aboard the Bion-M 1 satellite for 30 days. After landing mice were examined and transported to Moscow, where animals of the <i>in vitro</i> subgroup were dissected, while recovery dynamics was followed in the <i>in-vivo</i> subgroup before dissection 7 days after landing. Ground control experiment replicated the principal stages of the spaceflight experiment.</p

Bodyweight (A) and relative BW change (B) after telemetry probe implantation.

<p>Differences significant at p<0.05 are marked with an asterisk. As can be considered from bodyweight data, acute recovery was over by day 5 after surgery.</p

Post-flight open field behavior parameters expressed as percent of pre-flight background values.

<p>Total (A), center (B) and periphery (C) distance moved, rearing frequency (D), center entries frequency (E), time in center (F), latency to the first center zone entry (G) and grooming duration (H). SF mice displayed reduced activity compared to any of the control groups (GC, SFV or GCV). Mice housed in habitats (SF and GC) were reluctant to explore the center of the arena. Statistical analysis was performed using Mann-Whitney test (*−p<0.05, **−p<0.01, ***− p<0.005 and ****−p<0.0001, ns – not significant or the p value is indicated).</p

Climate parameters in flight, control experiments and the animal facility.

<p>na – data not available.</p

A representative photograph of mice in the flight habitats.

<p>Note that mice occupy the floor grid before launch (upper row) and cling to the feeder (lower row) in microgravity. The same cages are shown.</p

Number of animals in experimental groups after post-flight adjustment.

<p>Number of animals in experimental groups after post-flight adjustment.</p