Development of a Rabbit Iliac Arterial Stenosis Model Using a Controlled Cholesterol Diet and Pullover Balloon Injury.

Purpose: This study aimed to develop a rabbit iliac stenosis model and evaluate the effects of different mechanical injury techniques on the degree of arterial stenosis. Materials and Methods: Eighteen rabbits were divided into three groups: cholesterol-fed with pullover balloon injury (group A; n = 6), cholesterol-fed with localized balloon dilatation (group B; n = 6), and chow-diet with pullover balloon injury (group C; n = 6). After baseline angiography, the left iliac arteries of all rabbits were injured with a 3 × 10 mm noncompliant balloon using either a wide pullover technique (groups A and C) or a localized balloon dilatation technique (group B). A nine-week follow-up angiography was performed, and the angiographic late lumen loss and percentage of stenosis were compared. Results: Group A exhibited the most severe late lumen loss (A vs. B, 0.67 ± 0.13 vs. 0.04 ± 0.13 mm, p < 0.0001; A vs. C, 0.67 ± 0.13 vs. 0.26 ± 0.29 mm, p < 0.05; stenosis percentage 32.02% ± 6.54%). In contrast, group B showed a minimal percentage of stenosis (1.75% ± 6.55%). Conclusion: Pullover-balloon injury can lead to significant iliac artery stenosis in rabbits with controlled hypercholesterolemia. This model may be useful for elucidating the pathogenesis of atherosclerosis and for evaluating the efficacy of novel therapeutic interventions.

56 Fe ion exposure increases the incidence of lung and brain tumors at a similar rate in male and female mice.

The main deterrent to long-term space travel is the risk of Radiation Exposure Induced Death (REID). The National Aeronautics and Space Administration (NASA) has adopted Permissible Exposure Levels (PELs) to limit the probability of REID to 3% for the risk of death due to radiation-induced carcinogenesis. The most significant contributor to current REID estimates for astronauts is the risk of lung cancer. Recently updated lung cancer estimates from Japan's atomic bomb survivors showed that the excess relative risk of lung cancer by age 70 is roughly four-fold higher in females compared to males. However, whether sex differences may impact the risk of lung cancer due to exposure to high charge and energy (HZE) radiation is not well studied. Thus, to evaluate the impact of sex differences on the risk of solid cancer development post-HZE radiation exposure, we irradiated Rb fl/fl ; Trp53 fl/+ male and female mice infected with Adeno-Cre with various doses of 320 kVp X-rays or 600 MeV/n 56 Fe ions and monitored them for any radiation-induced malignancies. We observed that lung adenomas/carcinomas and esthesioneuroblastomas (ENBs) were the most common primary malignancies in X-ray and 56 Fe ion-exposed mice, respectively. In addition, 1 Gy 56 Fe ion exposure compared to X-rays led to a significantly higher incidence of lung adenomas/carcinomas (p=0.02) and ENBs (p<0.0001). However, we did not find a significantly higher incidence of any solid malignancies in female mice as compared to male mice, regardless of radiation quality. Furthermore, gene expression analysis of ENBs suggested a distinct gene expression pattern with similar hallmark pathways altered, such as MYC targets and MTORC1 signaling, in X-ray and 56 Fe ion-induced ENBs. Thus, our data revealed that 56 Fe ion exposure significantly accelerated the development of lung adenomas/carcinomas and ENBs compared to X-rays, but the rate of solid malignancies was similar between male and female mice, regardless of radiation quality.

Covered stent placement for gastroduodenal artery stump hemorrhage after pancreaticoduodenectomy: long-term patency and risk factor of stent failure.

OBJECTIVE: To evaluate the long-term outcomes of covered stent placement in patients with gastroduodenal artery (GDA) stump hemorrhage after pancreaticoduodenectomy (PD) and to identify risk factors of stent failure. METHODS AND MATERIALS: Covered stent was placed in total of 21 patients for GDA stump hemorrhage after PD from September 2012 to March 2021. Technical and clinical success, complications, and stent patency were retrospectively evaluated. Nine relevant variables were analyzed to determine risk factors for stent failure. RESULTS: In 20 of 21 patients (95.2%), the GDA stump was completely excluded with covered stent placement. Immediate hemostasis was achieved in the 20 patients and rebleeding from jejunal artery occurred in one patient which was successfully embolized one day after the stent placement. There was no procedure-related complication or early mortality (<30 days). During follow-up period (median 655.5 days), stent thrombosis was found on CT in 10 patients (50.0%, 10/20) without any laboratory or CT abnormalities. One thrombosed stent migrated into the jejunum 20 months after placement. The six-month, one-year, and two-year stent patency were 81.9%, 52.9%, and 37.8%, respectively (median 620 days). The recurrence of primary malignancy was associated with stent failure (HR 5.70; 95% CI 1.18-27.76, p = 0.03). CONCLUSIONS: Covered stent placement is an effective and safe management of postoperative GDA stump hemorrhage. Stent failure occurred frequently (50%) but did not cause liver ischemia. Stent failure was associated with recurrence of primary malignancy. ADVANCES IN KNOWLEDGE: 1. Covered stent placement is an effective and safe management of postoperative GDA stump hemorrhage.2. Stent failure occurred frequently (50%) but did not cause liver ischemia.3. Stent failure was associated with recurrence of primary malignancy.

Noninvasive assessment of foot perfusion in cholesterol-fed rabbits using dynamic volume perfusion CT with an upslope method.

To evaluate the feasibility of dynamic foot volume CT with the upslope method and to demonstrate macrovascular reactivity and microvascular perfusion during cuff-induced reactive hyperemia state in cholesterol-fed rabbits. 30 New Zealand male rabbits were divided into 2 groups: dietary hypercholesterolemia (n = 10) and normal diet control (n = 20). To measure for macrovascular reactivity, perfusion parameters of the left posterior tibial artery was measured at baseline and at reactive hyperemia state. For the evaluation of microvascular perfusion, color-coded perfusion map of the plantar dermis was generated for perfusion CT scan by an in-house developed dedicated analysis software based on upslope method. Dermal perfusion values were measured and analyzed before and after cuff-induced reactive hyperemia. Foot dynamic volume CT with the upslope method demonstrated significant impairment of both macrovascular reactivity and microvascular perfusion in cholesterol-fed rabbits without significant macrovascular lesions during cuff-induced reactive hyperemia (CRH) state. Arterial time-to-peak of cholesterol-fed rabbits failed to show acceleration while chow-fed rabbits showed significant decrease in time. Microvascular perfusion calculated by perfusion value (P < 0.01) and perfusion ratio (P = .014) showed decreased microvascular perfusion in cholesterol-fed rabbits compared to chow-fed rabbits during CRH state. Post-CT pathologic examination revealed decreased endothelial cell density in cholesterol-fed rabbits (P < 0.001). Foot perfusion CT using upslope method provides perfusion parameters for large arteries and a perfusion map of the foot during cuff-induced reactive hyperemia in cholesterol-fed rabbits. It may be a useful tool to assess microvascular reactivity in patients with peripheral artery disease but no apparent macrovascular lesions.

Appendiceal Visualization on 2-mSv CT vs. Conventional-Dose CT in Adolescents and Young Adults with Suspected Appendicitis: An Analysis of Large Pragmatic Randomized Trial Data.

OBJECTIVE: We compared appendiceal visualization on 2-mSv CT vs. conventional-dose CT (median 7 mSv) in adolescents and young adults and analyzed the undesirable clinical and diagnostic outcomes that followed appendiceal nonvisualization. MATERIALS AND METHODS: A total of 3074 patients aged 15-44 years (mean ± standard deviation, 28 ± 9 years; 1672 female) from 20 hospitals were randomized to the 2-mSv CT or conventional-dose CT group (1535 vs. 1539) from December 2013 through August 2016. A total of 161 radiologists from 20 institutions prospectively rated appendiceal visualization (grade 0, not identified; grade 1, unsure or partly visualized; and grade 2, clearly and entirely visualized) and the presence of appendicitis in these patients. The final diagnosis was based on CT imaging and surgical, pathologic, and clinical findings. We analyzed undesirable clinical or diagnostic outcomes, such as negative appendectomy, perforated appendicitis, more extensive than simple appendectomy, delay in patient management, or incorrect CT diagnosis, which followed appendiceal nonvisualization (defined as grade 0 or 1) and compared the outcomes between the two groups. RESULTS: In the 2-mSv CT and conventional-dose CT groups, appendiceal visualization was rated as grade 0 in 41 (2.7%) and 18 (1.2%) patients, respectively; grade 1 in 181 (11.8%) and 81 (5.3%) patients, respectively; and grade 2 in 1304 (85.0%) and 1421 (92.3%) patients, respectively (p < 0.001). Overall, undesirable outcomes were rare in both groups. Compared to the conventional-dose CT group, the 2-mSv CT group had slightly higher rates of perforated appendicitis (1.1% [17] vs. 0.5% [7], p = 0.06) and false-negative diagnoses (0.4% [6] vs. 0.0% [0], p = 0.01) following appendiceal nonvisualization. Otherwise, these two groups were comparable. CONCLUSION: The use of 2-mSv CT instead of conventional-dose CT impairs appendiceal visualization in more patients. However, appendiceal nonvisualization on 2-mSv CT rarely leads to undesirable clinical or diagnostic outcomes.

Can Patient Triaging with Clinical Scoring Systems Reduce CT Use in Adolescents and Young Adults Suspected of Having Appendicitis?

Background There are ongoing efforts to reduce CT radiation exposure for the diagnosis of appendicitis. Recent guidelines recommend using clinical scoring systems to triage patients who need imaging examinations. Purpose To determine whether patient triaging with scoring systems can reduce CT use without a loss of diagnostic accuracy in adolescents and young adults suspected of having appendicitis. Materials and Methods This retrospective study used data from a previous multicenter randomized controlled trial conducted between December 2013 and August 2016. Five scoring systems (adult appendicitis, appendicitis inflammatory response, modified Alvarado, Broek, and Christian scores) were used to categorize patients into low-, intermediate-, or high-probability groups. CT use was simulated for only the intermediate-probability group. The primary outcomes were CT reduction rate, sensitivity, and specificity. The CT reduction rate was defined as the proportion of patients in low- and high-probability groups who would not have to undergo CT among all patients. Sensitivity and specificity were calculated in the overall diagnostic pathway using each scoring system and subsequent CT. As a secondary analysis, to maintain the diagnostic accuracy to a level of when CT was used for all patients with suspected appendicitis, new cutoff values for probability group stratification targeting 97.6% sensitivity and 94.9% specificity were applied for each of the scoring systems. Results A total of 2888 patients (mean age ± standard deviation, 28 years ± 9; 1580 women and 1308 men) with suspected appendicitis were evaluated, of whom 1088 had and 1800 did not have appendicitis. The CT reduction rates of the five scoring systems ranged from 55.6% (1606 of 2888 patients) to 71.1% (2053 of 2888), but at the cost of sensitivity (range, 48.7% [530 of 1088] to 81.2% [883 of 1088]) and specificity (range, 79.0% [1422 of 1800] to 97.8% [1761 of 1800]). Targeting 97.6% sensitivity and 94.9% specificity, the CT reduction rates of all five scoring systems were 0% (0 of 2888). Conclusion Using clinical scoring systems in triaging patients for selective CT use led to a considerable loss of diagnostic accuracy. © RSNA, 2021 Online supplemental material is available for this article. See also the editorial by Mellnick in this issue.

Final diagnosis and patient disposition following equivocal results on 2-mSv CT vs. conventional-dose CT in adolescents and young adults with suspected appendicitis: a post hoc analysis of large pragmatic randomized trial data.

OBJECTIVE: To compare 2-mSv CT and conventional-dose CT (CDCT, typically 7-8 mSv) regarding final diagnosis and patient disposition following equivocal CT results in adolescents and young adults with suspected appendicitis. METHODS: In total, 3074 patients of 15-44 years (28 ± 9 years, 1672 women) from 20 hospitals were randomized to undergo contrast-enhanced 2-mSv CT (n = 1535) or CDCT (n = 1539) from December 2013 through August 2016. One hundred sixty-one radiologists prospectively rated the likelihood of appendicitis in a Likert scale (i.e., grades 1-5). The final diagnosis was based on CT image, surgical, pathologic, and clinical findings. Post hoc analysis was performed for final diagnosis, surgical procedure, and delay in patient management following equivocal results (i.e., grade 3). RESULTS: The 2-mSv CT and CDCT groups were comparable for final diagnosis following equivocal results, including confirmed appendicitis (1.2% [18 patients] vs. 1.2% [19], p > 0.99), negative appendectomy (0.1% [2] vs. 0.3% [4], p = 0.53), and perforated appendicitis (0.1% [1] vs. 0.2% [3], p = 0.53). More patients were confirmed as not having appendicitis following equivocal results in the CDCT group than in the 2-mSv CT group (2.2% [34] vs. 1.0% [16], p = 0.016). The two groups were comparable for the need of appendectomy (1.4% [22] vs. 1.5% [23], p > 0.99), need of additional imaging tests (0.7% [11] vs. 1.1% [17], p = 0.35), and delay in patient management following equivocal results. CONCLUSION: 2-mSv CT is comparable to CDCT regarding final diagnosis and patient disposition following equivocal CT results. KEY POINTS: • Our results strengthen evidence justifying the use of low-dose CT instead of conventional-dose CT (CDCT) in adolescents and young adults with suspected appendicitis. • The 2-mSv CT and CDCT groups were comparable for final diagnosis following equivocal CT results, including confirmed appendicitis (1.2% vs. 1.2%, p > 0.99), negative appendectomy (0.1% vs. 0.3%, p = 0.53), and perforated appendicitis (0.1% vs. 0.2%, p = 0.53). • The two groups were comparable for the need for appendectomy (1.4% vs. 1.5%, p > 0.99), need for additional imaging tests (0.7% vs. 1.1%, p = 0.35), and delay in patient management, following equivocal CT results.

An extra copy of p53 suppresses development of spontaneous Kras-driven but not radiation-induced cancer.

The tumor suppressor p53 blocks tumor progression in multiple tumor types. Radiation-induced cancer following exposure to radiation therapy or space travel may also be regulated by p53 because p53 has been proposed to respond to DNA damage to suppress tumorigenesis. Here, we investigate the role of p53 in lung carcinogenesis and lymphomagenesis in LA-1 KrasG12D mice with wild-type p53 or an extra copy of p53 (super p53) exposed to fractionated total body irradiation with low linear energy transfer (low-LET) X-rays or high-LET iron ions and compared tumor formation in these mice with unirradiated controls. We found that an additional copy of p53 suppressed both Kras-driven lung tumor and lymphoma development in the absence of radiation. However, an additional copy of p53 did not affect lymphoma development following low- or high-LET radiation exposure and was unable to suppress radiation-induced expansion of thymocytes with mutated Kras. Moreover, radiation exposure increased lung tumor size in super p53 but not wild-type p53 mice. These results demonstrate that although p53 suppresses the development of spontaneous tumors expressing KrasG12D, in the context of exposure to ionizing radiation, an extra copy of p53 does not protect against radiation-induced lymphoma and may promote KrasG12D mutant lung cancer.

HIF-1 Alpha Regulates the Response of Primary Sarcomas to Radiation Therapy through a Cell Autonomous Mechanism.

Hypoxia is a major cause of radiation resistance, which may predispose to local recurrence after radiation therapy. While hypoxia increases tumor cell survival after radiation exposure because there is less oxygen to oxidize damaged DNA, it remains unclear whether signaling pathways triggered by hypoxia contribute to radiation resistance. For example, intratumoral hypoxia can increase hypoxia inducible factor 1 alpha (HIF-1α), which may regulate pathways that contribute to radiation sensitization or radiation resistance. To clarify the role of HIF-1α in regulating tumor response to radiation, we generated a novel genetically engineered mouse model of soft tissue sarcoma with an intact or deleted HIF-1α. Deletion of HIF-1α sensitized primary sarcomas to radiation exposure in vivo. Moreover, cell lines derived from primary sarcomas lacking HIF-1α, or in which HIF-1α was knocked down, had decreased clonogenic survival in vitro, demonstrating that HIF-1α can promote radiation resistance in a cell autonomous manner. In HIF-1α-intact and -deleted sarcoma cells, radiation-induced reactive oxygen species, DNA damage repair and activation of autophagy were similar. However, sarcoma cells lacking HIF-1α had impaired mitochondrial biogenesis and metabolic response after irradiation, which might contribute to radiation resistance. These results show that HIF-1α promotes radiation resistance in a cell autonomous manner.

Tumor cells, but not endothelial cells, mediate eradication of primary sarcomas by stereotactic body radiation therapy.

Cancer clinics currently use high-dose stereotactic body radiation therapy as a curative treatment for several kinds of cancers. However, the contribution of vascular endothelial cells to tumor response to radiation remains controversial. Using dual recombinase technology, we generated primary sarcomas in mice with targeted genetic mutations specifically in tumor cells or endothelial cells. We selectively mutated the proapoptotic gene Bax or the DNA damage response gene Atm to genetically manipulate the radiosensitivity of endothelial cells in primary soft tissue sarcomas. Bax deletion from endothelial cells did not affect radiation-induced cell death in tumor endothelial cells or sarcoma response to radiation therapy. Although Atm deletion increased endothelial cell death after radiation therapy, deletion of Atm from endothelial cells failed to enhance sarcoma eradication. In contrast, deletion of Atm from tumor cells increased sarcoma eradication by radiation therapy. These results demonstrate that tumor cells, rather than endothelial cells, are critical targets that regulate sarcoma eradication by radiation therapy. Treatment with BEZ235, a small-molecule protein kinase inhibitor, radiosensitized primary sarcomas more than the heart. These results suggest that inhibiting ATM kinase during radiation therapy is a viable strategy for radiosensitization of some tumors.

Automated measurement of microcirculatory blood flow velocity in pulmonary metastases of rats.

Because the lung is a major target organ of metastatic disease, animal models to study the physiology of pulmonary metastases are of great importance. However, very few methods exist to date to investigate lung metastases in a dynamic fashion at the microcirculatory level, due to the difficulty to access the lung with a microscope. Here, an intravital microscopy method is presented to functionally image and quantify the microcirculation of superficial pulmonary metastases in rats, using a closed-chest pulmonary window and automated analysis of blood flow velocity and direction. The utility of this method is demonstrated to measure increases in blood flow velocity in response to pharmacological intervention, and to image the well-known tortuous vasculature of solid tumors. This is the first demonstration of intravital microscopy on pulmonary metastases in a closed-chest model. Because of its minimized invasiveness, as well as due to its relative ease and practicality, this technology has the potential to experience widespread use in laboratories that specialize on pulmonary tumor research.

Atm deletion with dual recombinase technology preferentially radiosensitizes tumor endothelium.

Cells isolated from patients with ataxia telangiectasia are exquisitely sensitive to ionizing radiation. Kinase inhibitors of ATM, the gene mutated in ataxia telangiectasia, can sensitize tumor cells to radiation therapy, but concern that inhibiting ATM in normal tissues will also increase normal tissue toxicity from radiation has limited their clinical application. Endothelial cell damage can contribute to the development of long-term side effects after radiation therapy, but the role of endothelial cell death in tumor response to radiation therapy remains controversial. Here, we developed dual recombinase technology using both FlpO and Cre recombinases to generate primary sarcomas in mice with endothelial cell-specific deletion of Atm to determine whether loss of Atm in endothelial cells sensitizes tumors and normal tissues to radiation. Although deletion of Atm in proliferating tumor endothelial cells enhanced the response of sarcomas to radiation, Atm deletion in quiescent endothelial cells of the heart did not sensitize mice to radiation-induced myocardial necrosis. Blocking cell cycle progression reversed the effect of Atm loss on tumor endothelial cell radiosensitivity. These results indicate that endothelial cells must progress through the cell cycle in order to be radiosensitized by Atm deletion.

Assessing cardiac injury in mice with dual energy-microCT, 4D-microCT, and microSPECT imaging after partial heart irradiation.

PURPOSE: To develop a mouse model of cardiac injury after partial heart irradiation (PHI) and to test whether dual energy (DE)-microCT and 4-dimensional (4D)-microCT can be used to assess cardiac injury after PHI to complement myocardial perfusion imaging using micro-single photon emission computed tomography (SPECT). METHODS AND MATERIALS: To study cardiac injury from tangent field irradiation in mice, we used a small-field biological irradiator to deliver a single dose of 12 Gy x-rays to approximately one-third of the left ventricle (LV) of Tie2Cre; p53(FL/+) and Tie2Cre; p53(FL/-) mice, where 1 or both alleles of p53 are deleted in endothelial cells. Four and 8 weeks after irradiation, mice were injected with gold and iodinated nanoparticle-based contrast agents, and imaged with DE-microCT and 4D-microCT to evaluate myocardial vascular permeability and cardiac function, respectively. Additionally, the same mice were imaged with microSPECT to assess myocardial perfusion. RESULTS: After PHI with tangent fields, DE-microCT scans showed a time-dependent increase in accumulation of gold nanoparticles (AuNp) in the myocardium of Tie2Cre; p53(FL/-) mice. In Tie2Cre; p53(FL/-) mice, extravasation of AuNp was observed within the irradiated LV, whereas in the myocardium of Tie2Cre; p53(FL/+) mice, AuNp were restricted to blood vessels. In addition, data from DE-microCT and microSPECT showed a linear correlation (R(2) = 0.97) between the fraction of the LV that accumulated AuNp and the fraction of LV with a perfusion defect. Furthermore, 4D-microCT scans demonstrated that PHI caused a markedly decreased ejection fraction, and higher end-diastolic and end-systolic volumes, to develop in Tie2Cre; p53(FL/-) mice, which were associated with compensatory cardiac hypertrophy of the heart that was not irradiated. CONCLUSIONS: Our results show that DE-microCT and 4D-microCT with nanoparticle-based contrast agents are novel imaging approaches complementary to microSPECT for noninvasive assessment of the change in myocardial vascular permeability and cardiac function of mice in whom myocardial injury develops after PHI.

PD-0332991, a CDK4/6 inhibitor, significantly prolongs survival in a genetically engineered mouse model of brainstem glioma.

Diffuse intrinsic pontine glioma (DIPG) is an incurable tumor that arises in the brainstem of children. To date there is not a single approved drug to effectively treat these tumors and thus novel therapies are desperately needed. Recent studies suggest that a significant fraction of these tumors contain alterations in cell cycle regulatory genes including amplification of the D-type cyclins and CDK4/6, and less commonly, loss of Ink4a-ARF leading to aberrant cell proliferation. In this study, we evaluated the therapeutic approach of targeting the cyclin-CDK-Retinoblastoma (Rb) pathway in a genetically engineered PDGF-B-driven brainstem glioma (BSG) mouse model. We found that PD-0332991 (PD), a CDK4/6 inhibitor, induces cell-cycle arrest in our PDGF-B; Ink4a-ARF deficient model both in vitro and in vivo. By contrast, the PDGF-B; p53 deficient model was mostly resistant to treatment with PD. We noted that a 7-day treatment course with PD significantly prolonged survival by 12% in the PDGF-B; Ink4a-ARF deficient BSG model. Furthermore, a single dose of 10 Gy radiation therapy (RT) followed by 7 days of treatment with PD increased the survival by 19% in comparison to RT alone. These findings provide the rationale for evaluating PD in children with Ink4a-ARF deficient gliomas.

Oncogene-dependent control of miRNA biogenesis and metastatic progression in a model of undifferentiated pleomorphic sarcoma.

Undifferentiated pleomorphic sarcoma (UPS) is one of the most common soft tissue malignancies. Patients with large, high-grade sarcomas often develop fatal lung metastases. Understanding the mechanisms underlying sarcoma metastasis is needed to improve treatment of these patients. Micro-RNAs (miRNAs) are a class of small RNAs that post-transcriptionally regulate gene expression. Global alterations in miRNAs are frequently observed in a number of disease states including cancer. The signalling pathways that regulate miRNA biogenesis are beginning to emerge. To test the relevance of specific oncogenic mutations in miRNA biogenesis in sarcoma, we used primary soft tissue sarcomas expressing either Braf(V600E) or Kras(G12D). We found that Braf(V600E) mutant tumours, which have increased MAPK signalling, have higher levels of mature miRNAs and enhanced miRNA processing. To investigate the relevance of oncogene-dependent alterations in miRNA biogenesis, we introduced conditional mutations in Dicer and showed that Dicer haploinsufficiency promotes the development of distant metastases in an oncogene-dependent manner. These results demonstrate that a specific oncogenic mutation can cooperate with mutation in Dicer to promote tumour progression in vivo.