Abscopal effect of low-LET γ-radiation mediated through Rel protein signal transduction in a mouse model of nontargeted radiation response.

Ascertaining the ionizing radiation (IR)-induced bystander response and its preceding molecular regulation would increase our understanding of the mechanism of acute and delayed radiobiological effects. Recent evidence clearly prompted that radiation-induced nuclear factor kappa B (NF-κB) would play a key role in bystander responses in nontargeted cells. Accordingly, we investigated the orchestration of NF-κB signaling after IR in a nontargeted distant organ. Heart tissues from C57/BL6 mice either mock irradiated or exposed (limited to lower abdomen 1 cm diameter) to single-dose IR (SDR: 2 or 10 Gy) or fractionated IR (FIR, 2 Gy per day for 5 days) were examined for onset of abscopal NF-κB signal transduction, translated activity, downstream functional signaling and associated DNA damage. Radiation significantly induced NF-κB DNA binding activity in nontargeted heart. Transcriptional profiling showed that 51, 46 and 26 of 88 genes were significantly upregulated after 2 Gy, 10 Gy and FIR. Of these genes, 22 showed dose- and fractionation-independent upregulation. Immunohistochemistry revealed a robust increase in p65 and cMyc expression in distant heart after SDR and FIR. Immunoblotting revealed increased phosphorylation of p38 after 2 Gy and extracellular signal-regulated kinases 1/2 after 10 Gy in nontargeted heart. In addition, IR exposure significantly enhanced DNA fragmentation in nontargeted heart. Together, these data clearly indicated an induced abscopal response in distant organ after clinically relevant IR doses. More importantly, the results imply that orchestration of NF-κB signal transduction in nontargeted tissues may serve as an effector and could play a key role in induced abscopal responses.

Curcumin-altered p53-response genes regulate radiosensitivity in p53-mutant Ewing's sarcoma cells.

AIM: Curcumin has been demonstrated to have antitumor effects including radiosensitization by modulating many molecular targets including p53. Herein, we investigated the radiosensitizing effect of curcumin in p53 mutant Ewing's sarcoma (ES) cells. MATERIALS AND METHODS: Cells exposed to radiation with or without curcumin were examined for transcriptional and translational levels of p53 downstream targets and its influence in regulated apoptosis, DNA fragmentation, cell survival and clonal expansion. RESULTS: Curcumin significantly caused radiation induced expression of p21 and Bax, and reduced BclXl, Mcl1 with only marginal Bcl2 modulation. As a positive control to the study, both transcriptional and translational levels of p53 remained unchanged after radiation with/without curcumin. Conversely, curcumin caused radiation-induced apoptosis and DNA fragmentation. Consistently, curcumin enhanced radiation-induced cytotoxicity and clonal expansion. CONCLUSION: These results suggest that curcumin potentially radiosensitizes p53-mutant ES cells by regulating IR-modulated p53-response genes. However, the curcumin-associated p53-independent regulation of downstream targets remains to be explored.

Effects of isoflurane, pentobarbital, and urethane on apoptosis and apoptotic signal transduction in rat kidney.

BACKGROUND: Renal cell apoptosis contributes significantly to the pathogenesis of acute renal failure. Anesthetic agents have been shown to modulate apoptotic signal transduction in various tissues. We examined the effects of 6 h of different general anesthetic techniques on renal cell apoptosis in rat kidneys. METHODS: Twenty-one male Sprague-Dawley rats were randomly allocated into four groups: (i) control, non-anesthetized rats (n= 3) and rats anesthetized with (ii) inhaled isoflurane (n= 6), (iii) intraperitoneal pentobarbital (n= 6), and (iv) intraperitoneal urethane (n= 6). Animals were sacrificed 6 h after the induction of anesthesia. RESULTS: Apoptosis was assessed by terminal deoxynucleotidyl transferase-fluorescein end-labeling analysis. RNA was extracted from the left kidney to probe cDNA microarrays. Gene expression was measured as a percentage of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and subsequently confirmed using reverse transcriptase-polymerase chain reaction (RT-PCR). Compared with the control (no anesthesia), urethane significantly (P < 0.001) induced apoptosis in both the renal cortex and medulla. Isoflurane significantly (P < 0.001) inhibited apoptosis in the medulla. Microarray analysis revealed that urethane up-regulated more (74) genes than pentobarbital (16) and isoflurane (10). Isoflurane down-regulated more genes (85) than pentobarbital (74) and urethane (12). These anesthetic-induced modulations were significant (P < 0.05) for 60 isoflurane-, 30 pentobarbital- and 4 urethane-modulated genes. CONCLUSION: Our results suggest that general anesthetic drugs have an effect on renal cell apoptosis and apoptotic signal transduction, and thus may potentially affect the risk of subsequent acute renal failure.

Transcriptional responses of rat skeletal muscle following hypoxia-reoxygenation and near ischaemia-reperfusion.

AIM: The effect of ischaemia/reperfusion or hypoxia/reoxygenation on gene expression has not been extensively studied. We hypothesized that in skeletal muscle, tissue hypoxia of similar magnitude but induced by different mechanisms would lead to different transcriptional responses. METHODS: Muscle gene transcription was assessed using microarray analysis and reverse transcriptase polymerase chain reaction in 18 rats exposed to regional hind limb near ischaemia/reperfusion (n = 6), hypoxia/reoxygenation (n = 6) or sham operation (n = 6). Hypoxic burden was measured by the area under the PtO(2)-time curve. RESULTS: PtO(2) was reduced in both the near ischaemia/reperfusion and hypoxia/reoxygenation groups. Although the hypoxic burden was similar, the genomic response was different for each condition. Near ischaemia/reperfusion had a greater effect on gene expression than hypoxia/reoxygenation. Using stringent criteria for changes in gene expression (i.e. more than or equal to twofold change vs. control) unique patterns of gene expression could be identified suggesting individualized transcriptional responses to each of these injuries. Several genes, including insulin-like growth factor 1 (IGF-1) and cyclin-dependent kinase inhibitor (p27(Kip1)) were induced by both injury types and these may have potential clinical application as markers of tissue damage. In contrast, no single gene was downregulated by both injury conditions. CONCLUSIONS: The mechanism of skeletal muscle hypoxia has a profound effect on its subsequent transcriptional response. We identified several potential candidates as markers of skeletal muscle ischaemic damage.

Post-translational modification of I-kappa B alpha activates NF-kappa B in human monocytes exposed to 56Fe ions.

The objective of this study was to investigate whether heavy ion (56Fe) radiation exposure activates one of the key transcriptional regulators, nuclear factor-kappa B (NF-kappa B), in normal human monocytes (Mono Mac 6 cells: MM6). The study revealed that the exposure of MM6 cells to 56Fe ions resulted in increased NF-kappa B DNA-binding activity. The activation was both dose- and time-dependent, with a maximum response at the 2 h time point after a 0.7 Gy dose. Cells pre-incubated with inhibitors of the phosphorylation and proteasome signaling pathway, completely blocked heavy ion-induced activation of NF-kappa B. These results clearly indicate that 56Fe ions can induce NF-kappa B DNA-binding activity in normal human monocytes, that the activation is rapid and persistent, and that the heavy ion-induced activation of NF-kappa B is mediated through phosphorylation of I-kappa B alpha and the subsequent proteasome-dependent degradation pathway. Since activation of NF-kappa B by extracellular stimuli is implicated in inflammation, infection and cancer induction, as well as in protection of cells against insult, it will be important in subsequent studies to elucidate whether heavy ion-induced NF-kappa B activation is involved in downstream gene expression.

Cutaneous SCC: metastases with a predilection for lungs 6 years after excision of the primary tumour.

An 80-year-old woman developed an ulcerated, moderately differentiated squamous cell carcinoma (SCC) on the lower leg. Despite local excision and radiotherapy, the patient presented 6 years later with multiple lung metastases which were histologically indistinguishable from the original skin tumour. There was no evidence of metastases to lymph nodes or other viscera.