Inhibition of intestinal epithelial apoptosis improves survival in a murine model of radiation combined injury

World conditions place large populations at risk from ionizing radiation (IR) from detonation of dirty bombs or nuclear devices. In a subgroup of patients, ionizing radiation exposure would be followed by a secondary infection. The effects of radiation combined injury are potentially more lethal than either insult in isolation. The purpose of this study was to determine mechanisms of mortality and possible therapeutic targets in radiation combined injury. Mice were exposed to IR with 2.5 Gray (Gy) followed four days later by intratracheal methicillin-resistant Staphylococcus aureus (MRSA). While either IR or MRSA alone yielded 100% survival, animals with radiation combined injury had 53% survival (p = 0.01). Compared to IR or MRSA alone, mice with radiation combined injury had increased gut apoptosis, local and systemic bacterial burden, decreased splenic CD4 T cells, CD8 T cells, B cells, NK cells, and dendritic cells, and increased BAL and systemic IL-6 and G-CSF. In contrast, radiation combined injury did not alter lymphocyte apoptosis, pulmonary injury, or intestinal proliferation compared to IR or MRSA alone. In light of the synergistic increase in gut apoptosis following radiation combined injury, transgenic mice that overexpress Bcl-2 in their intestine and wild type mice were subjected to IR followed by MRSA. Bcl-2 mice had decreased gut apoptosis and improved survival compared to WT mice (92% vs. 42%; p<0.01). These data demonstrate that radiation combined injury results in significantly higher mortality than could be predicted based upon either IR or MRSA infection alone, and that preventing gut apoptosis may be a potential therapeutic target

Pheochromocytoma presenting with arterial and intracardiac thrombus in a 47-year-old woman: a case report

<p>Abstract</p> <p>Introduction</p> <p>Pheochromocytoma is a rare cause of hypertension but it could have severe consequences if not recognized and treated appropriately. The association of pheochromocytoma and thrombosis is even rarer but significantly increases management complexity, morbidity and mortality. To the best of our knowledge, this is the first report of a patient with pheochromocytoma presenting with left axillary arterial and intracardiac thrombus.</p> <p>Case presentation</p> <p>A 47-year-old Caucasian woman with a past medical history of hypertension presented for medical attention with left arm numbness. Doppler ultrasound showed an obstructing thrombus in her left axillary artery. She had symptom resolution after stent placement in her left axillary artery. A subsequent echocardiogram demonstrated a large intracardiac mass and abdominal computed tomography revealed a 7 cm mass between her spleen and left kidney. Labile blood pressure was noted during admission and she had very high levels of plasma and 24-hour urine catecholamines and metanephrines tests. A (123)I- metaiodobenzylguanidine scan showed intense uptake in the left abdominal mass. After adequate alpha blockage with phenoxybenzamine, laparoscopic tumor resection was performed without complications. She had normal metanephrines and complete symptom resolution afterwards. The intracardiac mass also disappeared with anticoagulation. All other endocrine laboratory abnormalities returned to normal after surgery.</p> <p>Conclusion</p> <p>Arterial and ventricular thrombosis occurring in patients with pheochromocytoma is rare. A multi-disciplinary approach is necessary in caring for this type of patient. Catecholamines likely contributed to the development of thrombosis in our patient. Early recognition of pheochromocytoma is the key to improving outcome.</p

Radiation combined injury causes increased systemic levels of IL-6, IL-10 and G-CSF.

<p>While neither IR nor MRSA caused alterations in systemic cytokines (except for an increase in G-CSF following MRSA), IL-6 (p<0.005), IL-10 (p<0.01) and G-CSF (p<0.05) were significantly increased in IR/MRSA mice compared to NR/sham, IR/sham, NR/MRSA mice (n = 10−14/group).</p

Radiation combined injury increases mortality compared to IR or MRSA alone.

<p>Mice that received either 2.5% survival. In contrast, when IR was followed four days later by MRSA, survival was only 53% (n = 8−15/group; p = 0.01). Note that day 0 in this survival curve represents when MRSA or sham pneumonia was induced, and IR was given four days earlier.</p

Increased gut apoptosis with radiation combined injury is associated with changes in the mitochondrial pathway.

<p>Real time PCR was performed on mediators in both the mitochondrial pathway and receptor-mediated pathway of apoptosis. Radiation combined injury caused an increase in mRNA levels of Bcl-2 (A), Bcl-x_L (B), and Bax (C, p<0.05 IR/MRSA compared to all other groups). In contrast, no alterations were noted in Bid (D), Fas (E), FADD (F) or TRADD levels (G, n = 7–8/group for all mediators).</p

Gut Bcl-2 overexpression prevents intestinal epithelial apoptosis and improves survival following radiation combined injury.

<p>Gut apoptosis was lower in <i>Fabpl</i>-Bcl-2 mice than WT mice following IR/MRSA by both H&E (A) and active caspase-3 (B) staining (p<0.05 for H&E, p<0.005 for active caspase-3, n = 12–15). Preventing gut apoptosis led to improved outcome, as survival was 92% in <i>Fabpl</i>-Bcl-2 mice subjected to radiation combined injury compared to 42% in WT mice given the same insult (p<0.01, n = 12–13/group). Note that day 0 in this survival curve represents when MRSA pneumonia was induced, and IR was given four days earlier.</p

Radiation combined injury causes increased BAL IL-6 and G-CSF.

<p>MRSA pneumonia caused an increase in BAL IL-6, G-CSF, TNFα, IL-1β and IL-10 compared to NR/sham mice (p<0.001 for IL-6 and IL-1β, p<0.0001 for G-CSF, TNFα, and IL-10). A further increase in IL-6 and G-CSF (p<0.001) was seen in IR/MRSA mice compared to NR/MRSA animals (n = 12−17/group for al comparisons).</p

Radiation combined injury worsens the decrease in splenic immune populations seen following IR.

<p>Isolated splenocytes were stained for CD4 T cells (A), CD8 T cells (B), B cells (C), NK cells (D), dendritic cells (E), and macrophages (F) to determine differential splenic cell counts (n = 9−12 IR/sham, NR/MRSA, IR/sham, n = 5 NR/sham). Mice subjected to IR alone had lower cell counts than NR/sham mice for all populations measured except macrophages (p<0.01 for all). Cell counts were further diminished in animals subjected to radiation combined injury (p<0.001 for CD4, p<0.0001 for CD8, B cells, NK cells, p<0.01 for dendritic cells comparing IR/MRSA to IR/sham).</p

Radiation combined injury increases local and systemic infection following MRSA pneumonia.

<p>Bacterial colony counts were measured in BAL fluid (A) and blood (B) from mice subjected to MRSA pneumonia alone or radiation combined injury (n = 9−10/group). Animals subjected to IR/MRSA had higher levels of MRSA recovered from BAL fluid (p<0.05) and blood (p<0.001) 24 hours after induction of pneumonia.</p