Xian Luo-Owen

Spaceflight effects on T lymphocyte distribution, function and gene expression

The immune system is highly sensitive to stressors present during spaceflight. The major emphasis of this study was on the T lymphocytes in C57BL/6NTac mice after return from a 13-day space shuttle mission (STS-118). Spleens and thymuses from flight animals (FLT) and ground controls similarly housed in animal enclosure modules (AEM) were evaluated within 3-6 h after landing. Phytohemagglutinin-induced splenocyte DNA synthesis was significantly reduced in FLT mice when based on both counts per minute and stimulation indexes (P < 0.05). Flow cytometry showed that CD3(+) T and CD19(+) B cell counts were low in spleens from the FLT group, whereas the number of NK1.1(+) natural killer (NK) cells was increased (P < 0.01 for all three populations vs. AEM). The numerical changes resulted in a low percentage of T cells and high percentage of NK cells in FLT animals (P < 0.05). After activation of spleen cells with anti-CD3 monoclonal antibody, interleukin-2 (IL-2) was decreased, but IL-10, interferon-gamma, and macrophage inflammatory protein-1alpha were increased in FLT mice (P < 0.05). Analysis of cancer-related genes in the thymus showed that the expression of 30 of 84 genes was significantly affected by flight (P < 0.05). Genes that differed from AEM controls by at least 1.5-fold were Birc5, Figf, Grb2, and Tert (upregulated) and Fos, Ifnb1, Itgb3, Mmp9, Myc, Pdgfb, S100a4, Thbs, and Tnf (downregulated). Collectively, the data show that T cell distribution, function, and gene expression are significantly modified shortly after return from the spaceflight environment.

Effects of spaceflight on innate immune function and antioxidant gene expression

Spaceflight conditions have a significant impact on a number of physiological functions due to psychological stress, radiation, and reduced gravity. To explore the effect of the flight environment on immunity, C57BL/6NTac mice were flown on a 13-day space shuttle mission (STS-118). In response to flight, animals had a reduction in liver, spleen, and thymus masses compared with ground (GRD) controls (P < 0.005). Splenic lymphocyte, monocyte/macrophage, and granulocyte counts were significantly reduced in the flight (FLT) mice (P < 0.05). Although spontaneous blastogenesis of splenocytes in FLT mice was increased, response to lipopolysaccharide (LPS), a B-cell mitogen derived from Escherichia coli, was decreased compared with GRD mice (P < 0.05). Secretion of IL-6 and IL-10, but not TNF-alpha, by LPS-stimulated splenocytes was increased in FLT mice (P < 0.05). Finally, many of the genes responsible for scavenging reactive oxygen species were upregulated after flight. These data indicate that exposure to the spaceflight environment can increase anti-inflammatory mechanisms and change the ex vivo response to LPS, a bacterial product associated with septic shock and a prominent Th1 response.

Low-Dose Photons Modify Liver Response to Simulated Solar Particle Event Protons

Abstract Gridley, D. S., Coutrakon, G. B., Rizvi, A., Bayeta, E. J. M., Luo-Owen, X., Makinde, A. Y., Baqai, F., Koss, P., Slater, J. M. and Pecaut, M. J. Low-Dose Photons Modify Liver Response to Simulated Solar Particle Event Protons. Radiat. Res. 169, 280–287 (2008). The health consequences of exposure to low-dose radiation combined with a solar particle event during space travel remain unresolved. The goal of this study was to determine whether protracted radiation exposure alters gene expression and oxidative burst capacity in the liver, an organ vital in many biological processes. C57BL/6 mice were whole-body irradiated with 2 Gy simulated solar particle event (SPE) protons over 36 h, both with and without pre-exposure to low-dose/low-dose-rate photons (57Co, 0.049 Gy total at 0.024 cGy/h). Livers were excised immediately after irradiation (day 0) or on day 21 thereafter for analysis of 84 oxidative stress-related genes using RT-PCR; genes up or down-regulated by more than twofold were noted. On day 0, genes with increased expression were: photons, none; simulated SPE, Id1; photons + simulated SPE, Bax, Id1, Snrp70. Down-regulated genes at this same time were: photons, Igfbp1; simulated SPE, Arnt2, Igfbp1, Il6, Lct, Mybl2, Ptx3. By day 21, a much greater effect was noted than on day 0. Exposure to photons + simulated SPE up-regulated completely different genes than those up-regulated after either photons or the simulated SPE alone (photons, Cstb; simulated SPE, Dctn2, Khsrp, Man2b1, Snrp70; photons + simulated SPE, Casp1, Col1a1, Hspcb, Il6st, Rpl28, Spnb2). There were many down-regulated genes in all irradiated groups on day 21 (photons, 13; simulated SPE, 16; photons + simulated SPE, 16), with very little overlap among groups. Oxygen radical production by liver phagocytes was significantly enhanced by photons on day 21. The results demonstrate that whole-body irradiation with low-dose-rate photons, as well as time after exposure, had a great impact on liver response to a simulated solar particle event.

Pneumatosis Intestinalis Predictive Evaluation Study: A multicenter epidemiologic study of the American Association for the Surgery of Trauma

BACKGROUND Our group has previously published a retrospective review defining variables predictive of transmural bowel ischemia in the setting of pneumatosis intestinalis (PI). We hypothesize this prospective study will confirm the findings of the retrospective review, enhancing legitimacy to the predictive factors for pathologic PI previously highlighted. METHODS Data were collected using the Research Electronic Data Capture. Forward logistic regression was utilized to identify independent predictors for pathologic PI. Statistical significance was defined as p ⩽ 0.05. RESULTS During the 3-year study period, 127 patients with PI were identified. Of these, 79 had benign disease, and 49 pathologic PI defined by the presence of transmural ischemia during surgical exploration or autopsy. Laboratory values such as elevated international normalized ratio (INR), decreased hemoglobin, and a lactate value of greater than 2.0 mmol/L were predictive of pathologic PI, as well as clinical factors including adynamic ileus, peritoneal signs on physical examination, sepsis, and hypotension. The location was also a significant factor, as patients with small bowel PI had a higher incidence of transmural ischemia than colonic PI. On multiple logistic regression, lactate value of greater than 2.0 mmol/L (odds ratio, 5.1, 1.3–19.5; p = 0.018), elevated INR (odds ratio, 3.2, 1.1–9.6; p = 0.031), peritonitis (15.0, 2.9–78; p = 0.001), and decreased hemoglobin (0.70, 0.50–0.97, 0.031) remained significant predictors of transmural ischemia (area under the curve, 0.90; 0.83–0.97). A lactate value of 2.0 mmol/L or greater and peritonitis are common factors between the retrospective review and this prospective study. CONCLUSIONS We recommend surgical exploration to be strongly considered for those PI patients presenting also with a lactate greater than 2 mmol/L and/or peritonitis. We suggest strong suspicion for necrosis in those patient with PI and small bowel involvement, ascites on computed tomography scan, adynamic ileus, anemia, and a high INR. LEVEL OF EVIDENCE Prognostic study, level II; therapeutic study, level II.

Low-dose Photon and Simulated Solar Particle Event Proton Effects on Foxp3+ T Regulatory Cells and other Leukocytes

Radiation is a major factor in the spaceflight environment that has carcinogenic potential. Astronauts on missions are continuously exposed to low-dose/low-dose-rate (LDR) radiation and may receive relatively high doses during a solar particle event (SPE) that consists primarily of protons. However, there are very few reports in which LDR photons were combined with protons. In this study, C57BL/6 mice were exposed to 1.7 Gy simulated SPE (sSPE) protons over 36 h, both with and without pre-exposure to 0.01 Gray (Gy) LDR γ-rays at 0.018 cGy/h. Apoptosis in skin samples was determined by immunohistochemistry immediately post-irradiation (day 0). Spleen mass relative to body mass, white blood cells (WBC), major leukocyte populations, lymphocyte subsets (T, Th, Tc, B, NK), and CD4+ CD25+ Foxp3+ T regulatory (Treg) cells were analyzed on days 4 and 21. Apoptosis in skin samples was evident in all irradiated groups; the LDR+sSPE mice had the greatest expression of activated caspase-3. On day 4 post-irradiation, the sSPE and LDR+sSPE groups had significantly lower WBC counts in blood and spleen compared to non-irradiated controls (p < 0.05 vs. 0 Gy). CD4+ CD25+ Foxp3+ Treg cell numbers in spleen were decreased at day 4, but proportions were increased in the sSPE and LDR+sSPE groups (p < 0.05 vs. 0 Gy). By day 21, lymphocyte counts were still low in blood from the LDR+sSPE mice, especially due to reductions in B, NK, and CD8+ T cytotoxic cells. The data demonstrate, for the first time, that pre-exposure to LDR photons did not protect against the adverse effects of radiation mimicking a large solar storm. The increased proportion of immunosuppressive CD4+ CD25+ Foxp3+ Treg and persistent reduction in circulating lymphocytes may adversely impact immune defenses that include removal of sub-lethally damaged cells with carcinogenic potential, at least for a period of time post-irradiation.

Comparison of proton and electron radiation effects on biological responses in liver, spleen and blood.

Purpose: To determine whether differences exist between proton and electron radiations on biological responses after total-body exposure. Materials and methods: ICR mice (n = 45) were irradiated to 2 Gray (Gy) using fully modulated 70 MeV protons (0.5 Gy/min) and 21 MeV electrons (3 Gy/min). At 36 h post-irradiation liver gene expression, white blood cell (WBC), natural killer (NK) cell and other analyses were performed. Results: Oxidative stress-related gene expression patterns were strikingly different for irradiated groups compared to 0 Gy (P < 0.05). Proton radiation up-regulated 15 genes (Ctsb, Dnm2, Gpx5, Il19, Il22, Kif9, Lpo, Nox4, Park7, Prdx4, Prdx6, Rag2, Sod3, Srxn1, Xpa) and down-regulated 2 genes (Apoe, Prdx1). After electron irradiation, 20 genes were up-regulated (Aass, Ctsb, Dnm2, Gpx1, Gpx4, Gpx5, Gpx6, Gstk1, Il22, Kif9, Lpo, Nox4, Park7, Prdx3, Prdx4, Prdx5, Rag2, Sod1, Txnrd3, Xpa) and 1 was down-regulated (Mpp4). Of the modified genes, only 11 were common to both forms of radiation. Comparison between the two irradiated groups showed that electrons significantly up-regulated three genes (Gstk1, Prdx3, Scd1). Numbers of WBC and major leukocyte types were low in the irradiated groups (P < 0.001 vs. 0 Gy). Hemoglobin and platelet counts were low in the electron-irradiated group (P < 0.05 vs. 0 Gy). However, spleens from electron-irradiated mice had higher WBC and lymphocyte counts, as well as enhanced NK cell cytotoxicity, compared to animals exposed to protons (P < 0.05). There were no differences between the two irradiated groups in body mass, organ masses, and other assessed parameters, although some differences were noted compared to 0 Gy. Conclusion: Collectively, the data demonstrate that at least some biological effects induced by electrons may not be directly extrapolated to protons.

Low-dose total-body γ irradiation modulates immune response to acute proton radiation.

Health risks due to exposure to low-dose/low-dose-rate radiation alone or when combined with acute irradiation are not yet clearly defined. This study quantified the effects of protracted exposure to low-dose/low-dose-rate γ rays with and without acute exposure to protons on the response of immune and other cell populations. C57BL/6 mice were irradiated with 57Co (0.05 Gy at 0.025 cGy/h); subsets were subsequently exposed to high-dose/high-dose-rate proton radiation (250 MeV; 2 or 3 Gy at 0.5 Gy/min). Analyses were performed at 4 and 17 days postexposure. Spleen and thymus masses relative to body mass were decreased on day 4 after proton irradiation with or without pre-exposure to γ rays; by day 17, however, the decrease was attenuated by the priming dose. Proton dose-dependent decreases, either with or without pre-exposure to γ rays, occurred in white blood cell, lymphocyte and granulocyte counts in blood but not in spleen. A similar pattern was found for lymphocyte subpopulations, including CD3+ T, CD19+ B, CD4+ T, CD8+ T and NK1.1+ natural killer (NK) cells. Spontaneous DNA synthesis by leukocytes after proton irradiation was high in blood on day 4 and high in spleen on day 17; priming with γ radiation attenuated the effect of 3 Gy in both body compartments. Some differences were also noted among groups in erythrocyte and thrombocyte characteristics. Analysis of splenocytes activated with anti-CD3/anti-CD28 antibodies showed changes in T-helper 1 (Th1) and Th2 cytokines. Overall, the data demonstrate that pre-exposure of an intact mammal to low-dose/low-dose-rate γ rays can attenuate the response to acute exposure to proton radiation with respect to at least some cell populations.

Analysis of a Metalloporphyrin Antioxidant Mimetic (MnTE-2-PyP) as a Radiomitigator: Prostate Tumor and Immune Status

Due to radiation-induced immune depression and development of pathologies such as cancer, there is increasing urgency to identify radiomitigators that are effective when administered after radiation exposure. The main goal of this study was to determine the radiomitigation capacity of MnTE-2-PyP[Mn(III) tetrakis (N-ethylpyridinium-2-yl) porphyrin], a superoxide dismutase (SOD) mimetic, and evaluate leukocyte parameters in spleen and blood. C57BL/6 mice were total-body exposed to 2 Gy γ-rays (Co-60), i.e., well below a lethal dose, followed by subcutaneous implantation of 5 × 105 RM-9 prostate tumor cells and initiation of MnTE-2-PyP treatment (day 0); interval between each procedure was 1–2 h. The drug was administered daily (12 times). Tumor progression was monitored and immunological analyses were performed on a subset per group on day 12. Animals treated with MnTE-2-PyP alone had significantly slower tumor growth compared to mice that did not receive the drug (P < 0.05), while radiation alone had no effect. Treatment of tumor-bearing mice with MnTE-2-PyP alone significantly increased spleen mass relative to body mass; the numbers of splenic white blood cells (WBC) and lymphocytes (B and T), as well as circulating WBC, granulocytes, and platelets, were high compared to one of more of the other groups (P < 0.05). The results show that MnTE-2-PyP slowed RM-9 tumor progression and up-regulated immune parameters, but mitigation of the effects of 2 Gy total-body irradiation were minimal.

Biological Effects of Passive Versus Active Scanning Proton Beams on Human Lung Epithelial Cells

The goal was to characterize differences in cell response after exposure to active beam scanning (ABS) protons compared to a passive delivery system. Human lung epithelial (HLE) cells were evaluated at various locations along the proton depth dose profile. The dose delivered at the Bragg peak position was essentially identical (∼4 Gy) with the two techniques, but depth dose data showed that ABS resulted in lower doses at entry and more rapid drop-off after the peak. Average dose rates for the passive and ABS beams were 1.1 Gy/min and 5.1 Gy/min, respectively; instantaneous dose rates were 19.2 Gy/min and 2,300 Gy/min (to a 0.5 × 0.5 mm2 voxel). Analysis of DNA synthesis was based on 3H-TdR incorporation. Quantitative real-time polymerase chain reaction (RT-PCR) was done to determine expression of genes related to p53 signaling and DNA damage; a total of 152 genes were assessed. Spectral karyotyping and analyses of the Golgi apparatus and cytokines produced by the HLE cells were also performed. At or near the Bragg peak position, ABS protons resulted in a greater decrease in DNA synthesis compared to passively delivered protons. Genes with >2-fold change (P < 0.05 vs. 0 Gy) after passive proton irradiation at one or more locations within the Bragg curve were BTG2, CDKN1A, IFNB1 and SIAH1. In contrast, many more genes had >2-fold difference with ABS protons: BRCA1, BRCA2, CDC25A, CDC25C, CCNB2, CDK1, DMC1, DNMT1, E2F1, EXO1, FEN1, GADD45A, GTSE1, IL-6, JUN, KRAS, MDM4, PRC1, PTTG1, RAD51, RPA1, TNF, WT1, XRCC2, XRCC3 and XRCC6BP1. Spectral karyotyping revealed numerous differences in chromosomal abnormalities between the two delivery systems, especially at or near the Bragg peak. Percentage of cells staining for the Golgi apparatus was low after exposure to passive and active proton beams. Studies such as this are needed to ensure patient safety and make modifications in ABS delivery, if necessary.

Space-relevant radiation modifies cytokine profiles, signaling proteins and Foxp3+ T cells

Abstract Purpose: The major goal was to evaluate effects of various radiation regimens on leukocyte populations relatively long-term after whole-body irradiation. Materials and methods: C57BL/6 mice were exposed to-low-dose/low-dose rate (LDR) 57Co γ-rays (0.01 Gy, 0.03 cGy/h), with and without acute 2 Gy proton (1 Gy/min) or γ-ray (0.9 Gy/min) irradiation; analyses were done on days 21 and 56 post-exposure. Results: Numerous radiation-induced changes were noted at one or both time points. Among the most striking differences (P < 0.05) were: (i) High percentage of CD4+CD25+Foxp3+ T cells in spleens from the Proton vs. LDR, Gamma and LDR + Proton groups (day 56); (ii) high interleukin-2 (IL-2) in spleen supernatants from the LDR and LDR + Proton groups vs. 0 Gy (day 56), whereas IL-10 was high in the LDR + Gamma group vs. 0 Gy (day 56); (iii) difference in transforming growth factor-β1 (TGF-β1) in spleen supernatants from Proton and LDR + Proton groups vs. Gamma and LDR + Gamma groups (both days); (iv) low TGF-β1 in blood from LDR + Proton vs. LDR + Gamma group (day 21); and (v) high level of activated cJun N-terminal kinase (JNK) in CD4+ T cells from LDR + Proton vs. LDR + Gamma group (day 21). Conclusions: The findings demonstrate that at least some immune responses to acute 2 Gy radiation were dependent on radiation quality time of assessment, and pre-exposure to LDR γ-rays.