Jacobs, Elizabeth R.

Medhora, Meetha

Molthen, Robert C.

Moulder, John E.

Schmirler, G.

Wu, Q.

English

epublications@Marquette

Marquette Universitye-Publications@MarquetteBiomedical Engineering Faculty Research andPublications Biomedical Engineering, Department of8-1-2007Pulmonary Microvascular Injury Resulting from aSingle Exposure to Low-Dose Thoracic RadiationQ. WuG. SchmirlerJohn E. MoulderMedical College of WisconsinElizabeth R. JacobsMedical College of WisconsinMeetha MedhoraMedical College of WisconsinSee next page for additional authorsPublished version. Published as part of the proceedings of the conference, Proceedings of the 8thWorld Congress for Microcirculation, Milwaukee, Wisconsin, 2007: 233-238. Publisher Link. © 2007Medimond Editore, International Proceedings Division. Used with permission.Robert Molthen was affiliated with the Medical College of Wisconsin at the time of publication.AuthorsQ. Wu, G. Schmirler, John E. Moulder, Elizabeth R. Jacobs, Meetha Medhora, and Robert C. MolthenThis conference proceeding is available at e-Publications@Marquette: https://epublications.marquette.edu/bioengin_fac/243Pulmonary Microvascular Injury Resulting from a Single Exposure to Low-Dose Thoracic Radiation Q. WlI, G. Schmirler, J.E. Moulder, E.R. Jacobs, M . Medhora and R. Molthen Medical College of Wisconsin (l nd Deparrmelll of Vetera ns Affairs, Zablocki VAMC. Milwaukee. USA Summary We describe a rat model of pos t-irrad iation (p.i.) pulmonary vascular in-jury resultant from low-dose radiation exposure. The model was developed to investi gate radiation exposure injuries expected in casualties resulting from a rad iological attack. Rats were challenged wi th a single dose of 5 or lOGy of radi ation or a sham irradiation procedure. Animals were studied at time points from 3-days to 12-months. We measured various phys iological parameters and used a novel technique of X-ray microfocal angiographic imaging to investigate pulmonary arterial morphology. With our analysis, exposure to 5 Gy had no measurable effect on pulmonary arterial structure/function, whereas lOGy resulted in a number of significant phys io logical abnormalities includ-ing inc reased vascular res istance, a decrease in to tal lung ACE activity, and vessel dropout. Introduction We hypothes ize that pu lmonary II1Jury will mani fes t in survivo rs of radio-logical attacks after they have been treated for more acute injury to the bone marrow, gastro intestinal tract and kidney. In our model of low-dose injury, we use radi ation limited to the thorax to reduce the confounding hemopoietic, gastrointestinal and renal effects in o rder to isolate lung pathobiology. The long-term goals of thi s work, in response to the cited need "', are to find Uea tments and/or mitigators that will improve lung health and functi on'- In order to accomplish our goa ls, we have worked on methods to qu antify ©2()()7 by MEDIMOND s.r.I . HS15C0267 233 234 8th World Congress for Microcirculation pneumopathy caused by radia t ion exposure level s likely to be present in casualties associated with a radiologica l dispersal device, or lower exposures compared to existing radiation therapy models' -6 . Although the pathogenesis of radiation -induced pneumopathy remains controversial, some groups have focused on radiation-induced endothelial cell death or dysfunction ' since changes in angiotensin converting enzyme, prostaglandins, thromboxane, endothelin, plasminogen activator, and platelet activating factor have been found in models of lung irradiation. Others be-lieve type II pneumocytes are the critical target and that surfactant released by the damaged type II cells triggers the radiation injury response'- A more likely scenario is that the radiation response includes some contribution from both mechanisms·. Our studies examined iso lated perfused lung preparations to measure metabolic , hemodynamic, and structural changes that result at different p.i. time points. At the 5 and lOGy dosages used in our present models, there appears to be an early response, which includes phased changes in and recoverable damage to the vasculature, and a late response resulting in interstitial lung disease and fibrosis. Materials and Methods All the studies were done under approval of the Medical College of Wis-consin and Zablocki V A Medical Center IACUC review boards and in com-pliance with the National Research Counci l's Guide for the Care and Use of Laboratory Animals. Unanesthetized female W AG/RJ MCW rats (110-170 gm, N= 144) were placed in a plexiglass holding jig a~d exposed to 0, 5 , or 10 Gy of radiation limited to the thorax by lead shielding. The X-ray field was from posterior to anterior at a dose rate of -2 Gy/minute. Exposed ani -mal s and their age-matched controls were studied at time points of 3-days, 2-weeks, and I , 2, 5, and 12-months(m) after irradiation or similar sham pro-cedure . Lung preparation, imaging methods and analysis follow those previ-ously described '0. Briefly, animals were anesthetized with sodium pentobar-bital (40mg/kg), a midline sternotomy performed, the animal was heparinized and blood sampled via right ventricle, the u'achea and pulmonary artery were cunnulated , and the heart and lungs removed . The left and right heart were weighed to assess hypertrophy. The lungs were hung by the cannulae, ven-tilated, rinsed of blood, and perfused with a 5% BSA physiological saline so lution . Angiotensin converting enzy me activity" and pulmonary vascular resistance were measured in the isolated perfused lungs. Then the perfusate in the arteries was replaced with perfiuorooctyl bromide for X-ray contrast and high-resolution angiograms of the lungs were acquired and used to analyze morphological changes in the vessels. Lastly, the lungs were placed in a drying oven for several days then weighed and normalized to body wei ght to access changes in total lung composition. All statistical comparisons were performed using at-test (Pd"0.02) in SigmaStat 3.0 (SPSS , Chicago, IL). Milwaukee, Wisconsill, USA , August 15-19, 2007 235 Results There was a dose dependent response to irradiation, with the 5Gy groups showing little measurable changes (no significant differences) whereas the lOGy groups had significant differences, in a number of parameters, that peak around 2-ms p.i., compared to their respective age matched controls. Hematocrit measured from blood sampled in the animals a t time of sacrifice was not significantly different in any of the groups (data not presented). Total lung ACE activity was sign ifi cantly reduced in the lungs of lOGy rats beginning I-m p.i. , see Figure I . A qualitative ana lysis of ang iograms of the lungs indicated that micro-vascular density was reduced beginning at I-m p.i. This rarefaction or vessel dropout appears to peak at 2-m p.i. and then return to near control status by 12-m p.i. , see Figure 2. The dry lung weight, which was normalized to body weight, was significantly increased in lOGy rats begin-ning at I-m p.i. , see Figure 3. Conclusion Thoracic irradiation of 5 and lOGy had a dose dependent but delayed impact on the normal structure and function of the pulmonary vasculature and related cardiovascular parameters, most of which reached significance l-to-2-m p.i. in the lOGy group. Irradiation with 5Gy had no significant effect, in contrast lOGy irradiation resu lted in decreased ACE activity after I-m, and ,. c & 12 E 0.. 10 « rJl !. e ~ ~ 6 " .. w u « 0> c: 2 " ...J 0'- '-,-LJ L I.,- L I.,- I.,- L t.,.-U .. U .. U .. U .. U .. U .. " ~ -" ~ Ii' Cl Ii' Cl E Cl E ~ 0 ~ 0 0 0 0 0 ... ~ ~ ~ ~ N ~ ,;, ~ ..:. -i' ... ~ ~ ~ ~ '" , N N '" N Figure I . Total lung ACE activity presel11ed as mean sUI/ace area product (MSAP) measured in lungs of control and irradiated rat at 3-d. 2-wk, I-m, 2-m, 5-m. and 12-m. 236 8th World Congress for Microcirculation increased normalized dry lung weight after I-m p.i .. There was a decrease in micro-vascular « 200"m) density after I-m p.i, which peaked at the 2-m p.i ., after which some of the injuries began to resolve . Although our experiments did not directly test for measures of fibrosis such as tota l lung compliance, 10 Figure 2. Hig h-magnification X-ray angiograms ofexarnple lungs/rom OGy (control) and 10 Gy irradiated ra ts at 3-d, 2-wk. I -m, 2-m, 5-m. and 12-m. In travascular pressure was sel to 12 mmHg a nd qinvay pressure 6 mmHg. Scale on boltom-center image indicates 100,.lm. Milwaukee, Wisconsin, USA, Augusl J 5- J 9, 2007 OOQ1$ 1: 00014 '" '0; ~ 000'2 '" § 00010 ...J ~ 00008 'C .. 00006 .~ ;;; E 00004 ~ 0 Z 00002 00000 ,., () is If -237 ,., () g Ii on Figure 3. Normalizetl dry /ullg weigh! measured il1 OCy (co fllrol) and /0 Gy irradiated rats at J-d, 2-wk. 1-111, 2-m, 5-111. alld 12 -m. increased lung weight and s ig nificantl y decreased body weight at the 12-m p .i. time point is likely re n ec ti ve of this later stage proce::ss. Supported by Department of Veterans Affairs and N IH U 19AI067734. References I. STONE, H. B., et aI. , Model s for evaluating agents intended for the proph ylaxis, mitigation and treatment of radiation injuries. Report of an NCI Workshop, De cember 3-4, 2003, Radiat Res, 6, 71 1-28, 2004. 2. AUG USTINE, A. D., et aI., Animal mode ls for radiatio n injury, prolecti on and therapy, Radiat Res, I, 100-9, 2005. 3. MOULDER, J . E. , Post-irradiation approaches to treatmcnl of radi ali on injuri es in the context of radiological terrorism and radiation acc idents: a review, 1m J Radiut Bioi, I, 3- 10, 2004. 4. MOLTENI, A., et aI., Control o f radiation-induced pneumopathy and lung librosis by angi otensin-convening enzy me inhibitors and an angiotensin II type I receptor blocker, lnt J Radiat Bioi , 4, 523-32, 2000. 5. KWOCK, L. , et aI. , The e ffects of ionizing radiation on the pulmonary vasc ul ature of intact rats and isolated pulmonary endothelium, Radial Res, 2, 276-91. t 987. 6. PETERSON_ L. M., et aI., Vascular response 10 radiation injury in Ihe ral lung, Radiat Res, 2 , 139-48, 1992. 238 8th World COllgress for Microcirculatioll 7. RUBIN, P. and CASARETI, G. W. , Clinical radiation pathology as applied to curative radiotherapy, Cancer. 4, 767·78, 1968. 8. VAN DEN BRANK, H. A. , Radiation effects of the pulmonary systemBaltimore, USA, William and Wilkins, 197 1. 9. WARD, W. F., MOLTENl, A. , TSAO, c., TAYLOR, J ., Endothelial orien ted strategies of intervention Boca Raton, FL, USA, CRe Press, 1998. 10. MOL THEN, R. c., et aI. , Quantitative models of the rat pulmonary arterial tree morphometry applied to hypoxia· induced arterial remodeling, J Appl Physiol, 6, 2372·84; discussion 2354, 2004. I I. HOLMQU IST, B., et a I. , A continuous spectrophotometric assay for angiotensin converting enzyme, Anal Biochem, 2, 540·8, 1979 . 

Pulmonary Microvascular Injury Resulting from a Single Exposure to Low-Dose Thoracic Radiation

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Pulmonary Microvascular Injury Resulting from a Single Exposure to Low-Dose Thoracic Radiation

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