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Aim: To better define clinically relevant non-classic radiation-induced liver disease (RILD) following stereotactic body radiotherapy (SBRT) in patients with small hepatocellular carcinoma (HCC)., Methods: We retrospectively evaluated the influence of acute liver toxicities on fatal hepatic failure in HCC patients treated with SBRT. Between April 2006 and February 2012, 194 HCC were treated with SBRT. Among them, patients followed up for more than 6 months were eligible. Laboratory results and Child-Pugh (CP) scores were obtained before treatment and at monthly follow-up visits. Toxicities were evaluated by the Common Terminology Criteria for Adverse Events version 4.0. Possible definitions of RILD were evaluated with respect to fatal hepatic failure within 12 months., Results: One hundred and eighty HCC were evaluated with a median follow-up of 28.2 months. Fatal hepatic failure within 12 months occurred in eight patients (4%). On univariate analysis, grade 3 or more elevated transaminases, CP score of 8 or more, and/or grade 3 or more decreased platelet count significantly predicted fatal hepatic failure within 12 months. Combinations of these factors (i.e. having at least one criterion) also predicted fatal hepatic failure within 12 months (16% with criteria vs 1% without criteria). Two-year overall survival rates for patients with and without RILD was 64.9% and 83.8% (P < 0.001), respectively., Conclusion: We identified three criteria that affected overall survival in HCC patients treated with SBRT. Further prospective studies are warranted to validate the safety and effect of SBRT for HCC., (© 2014 The Japan Society of Hepatology.)

Introduction: We investigated tolerable doses to organs at risk (OARs) in the mediastinum and pulmonary hilum following stereotactic body radiotherapy for centrally located lung tumors., Methods: Between 2005 and 2012, 381 patients with lung tumors were treated with stereotactic body radiotherapy of 40 to 60 Gy in five fractions. From among these patients, we extracted those who received greater than 25 Gy irradiation to OARs and analyzed dosimetric factors in relation to grade 3 to 5 toxicities., Results: In total, 398 OARs in 133 patients were analyzed, with a median follow-up of 33 (range, 3-87) months. The numbers receiving greater than 25 Gy irradiation to the aorta, vena cava, pulmonary artery, pulmonary vein, bronchus, trachea, heart, and esophagus were 72, 33, 73, 60, 55, 13, 69, and 23, respectively. The corresponding median Dmax 0.5 ml were 43.8, 32.0, 32.2, 29.1, 28.4, 28.7, 41.1, and 21.7 Gy. Of these patients, two developed grade 5 and one grade 3 hemoptysis, and two had grade 3 obstructive pneumonia. Two patients with grade 5 hemoptysis received high doses at the pulmonary artery and bronchus (59.2 and 54.4 Gy, and 61.3 and 59.6 Gy, respectively). No other grade 3 to 5 toxicities occurred., Conclusion: Therapeutic indications and dose-intensity should be carefully determined for patients with central tumors, especially when doses to the pulmonary artery and bronchus in the pulmonary hilum exceed 50 Gy. Tolerable doses for other OARs might, however, be higher than in this study, though longer follow-up is necessary to assess this possibility.

Systemic chemotherapy has enabled prolongation of survival in patients with stage IV colorectal cancer. This has subsequently increased the relative significance of local therapy for patients with oligometastases because they can be cured by removal of oligometastatic lesions. One of the most frequently reported tumor histologies for oligometastases is colorectal cancer. Resection is the standard therapy in most settings of oligometastases. Recently, studies have shown that stereotactic body radiotherapy (SBRT) may become a treatment option that provides high local control with minimal morbidity. Two-year local control rates following SBRT for hepatic and pulmonary oligometastases are almost over 80% and are even higher for patients treated with high-dose regimens. The indications of SBRT for other metastatic sites or conditions include isolated lymph nodes, spinal and adrenal metastasis, and post-surgical pelvic recurrence. Many retrospective studies have indicated that SBRT for various lesions results in good outcomes with low morbidity, both in the curative and palliative setting. However, few reports with a high level of evidence have indicated the efficacy of SBRT compared to standard therapy. Hereafter, the optimal indication of SBRT needs to be prospectively investigated to obtain convincing evidence.

The integration of new technologies has raised an interest in liver tumor radiotherapy, with literature evolving to support its efficacy. These advances, particularly stereotactic body radiation therapy (SBRT), have been critical in improving local control or potential cure in liver lesions not amenable to first-line surgical resection or radiofrequency ablation. Active investigation of SBRT, particularly for hepatocellular carcinoma (HCC), has recently started, yielding promising local control rates. In addition, data suggest a possibility that SBRT can be an alternative option for HCC unfit for other local therapies. However, information on optimal treatment indications, doses, and methods remains limited. In HCC, significant differences in patient characteristics and treatment availability exist by country. In addition, the prognosis of HCC is greatly influenced by underlying liver dysfunction and treatment itself in addition to tumor stage. Since they are closely linked to treatment approach, it is important to understand these differences in interpreting outcomes from various reports. Further studies are required to validate and maximize the efficacy of SBRT by a large, multi-institutional setting.

Background: Since 2005, we have treated hepatocellular carcinoma (HCC) with stereotactic body radiotherapy (SBRT) uniformly at two dose levels, according to baseline liver function and normal liver dose. We retrospectively examined the outcomes for these patients., Material and Methods: Between 2005 and 2012, 221 HCC patients were treated with SBRT. Eligibility criteria for SBRT included a single (either solitary or recurrent) HCC lesion; unfeasible, difficult or refusal to undergo other surgery or percutaneous ablative therapies; Child-Pugh Classification (CPC) A or B; tumors ≤ 5 cm; dose to the bowels < 25 Gy/5 fractions; curative intent. Patients followed up ≥ 6 months were eligible. The prescribed dose depended on liver function and liver dose: 40 Gy for CPC-A and 35 Gy for CPC-B, in 5 fractions, requiring a 5-Gy dose reduction if the proportion of the liver receiving ≥ 20 Gy exceeded 20%. Treatment outcomes and safety were analyzed., Results: A total of 185 patients (n = 48 in the 35-Gy group; n = 137 in the 40-Gy group) were eligible, with a median follow-up duration of 24 months (range 3-80). The three-year local control and overall survival rates were 91% and 70%, respectively. There were no significant differences in outcomes between dose levels: the three-year local control and overall survival rates in the 35-Gy and 40-Gy groups were 91% and 89% (log-rank p = 0.99) and 66% and 72% (p = 0.54), respectively. Acute toxicities ≥ grade 3 were observed in 24 (13.0%) patients, and 19 (10.3%) patients had worsening of CPC score by two points. All but three (1.6%) patients promptly recovered to grade 1-2. Grade 5 liver failure occurred in two patients in the 35-Gy group., Conclusion: SBRT for HCC was safe and provided equivalent outcomes when administered either in 35 or 40 Gy/5 fractions.

Background and Aims: Stereotactic ablative body radiotherapy (SABR) is a relatively new treatment for hepatocellular carcinoma (HCC). The outcomes of SABR for previously untreated solitary HCC unfit for ablation and surgical resection were evaluated., Methods: Untreated solitary HCC patients treated with SABR were retrospectively studied. Between 2005 and 2012, 221 HCC patients underwent SABR. Among them, patients with untreated solitary HCC, treated with only SABR or SABR preceded by transarterial chemoembolization, were eligible. Based on baseline liver function and liver volume receiving ≥ 20 Gy, 35-40 Gy in five fractions was prescribed to the planning target volume surface., Results: Sixty-three patients were eligible, with a median follow-up duration of 31.1 (range 12.0-88.1) months. No patients were lost to follow-up. Twenty patients were treated with only SABR. In 43 patients treated with SABR preceded by transarterial chemoembolization, accumulation of lipiodol in the tumor remained complete in five, a partial defect in 38 on pre-SABR computed tomography. The 1-, 2-, and 3-year local control rates were 100%, 95%, and 92%, respectively; the intrahepatic recurrence-free rates were 76%, 55%, and 36%, respectively; and the overall survival rates were 100%, 87%, and 73%, respectively. Grade 3 laboratory toxicities in the acute, subacute, and chronic phases were observed in 10, 9, and 13 patients, respectively, and ascites occurred in one patient., Conclusions: Local control and overall survival after SABR for untreated solitary HCC were excellent despite the candidates being unfit for resection and ablation. SABR is safe and might be an alternative to resection and ablation., (© 2013 Journal of Gastroenterology and Hepatology Foundation and Wiley Publishing Asia Pty Ltd.)

Purpose: Focal liver reaction (FLR) appears on radiographic images after stereotactic ablative body radiation therapy (SABR) in patients with hepatocellular carcinoma (HCC) and chronic liver disease. We investigated the threshold dose (TD) of FLR and possible factors affecting the TD on gadoxetate acid (Gd-EOB-DTPA)-enhanced magnetic resonance imaging (MRI)., Methods and Materials: In 50 patients who were treated with SABR for small HCC and followed up by MRI for >6 months, FLR, seen as a hypointense area, was evaluated on the hepatobiliary phase of Gd-EOB-DTPA-enhanced MRI. The follow-up MRI with the largest extent of FLR was fused to the planning computed tomography (CT) image, and patients with good image fusion concordance were eligible. After delineating the border of the FLR manually, a dose-volume histogram was used to identify the TD for the FLR. Clinical and volumetric factors were analyzed for correlation with the TD., Results: A total of 45 patients were eligible for analysis with a median image fusion concordance of 84.9% (range, 71.6-95.4%). The median duration between SABR and subsequent hepatobiliary phase MRI with the largest extent of FLR was 3 months (range, 1-6 months). The median TD for FLR was 28.0 Gy (range, 22.3-36.4 Gy). On univariate analysis, pre-treatment Child-Pugh (CP) score and platelet count were significantly correlated with the TD. On multiple linear regression analysis, CP score was the only parameter that predicted TD. Median TDs were 30.5 Gy (range, 26.2.3-36.4 Gy) and 25.2 Gy (range, 22.3-27.5 Gy) for patients with CP-A and CP-B disease, respectively., Conclusion: The TD was significantly correlated with baseline liver function. We propose 30 Gy for CP-A disease and 25 Gy for CP-B disease in 5 fractions as TDs for FLR after SABR for patients with HCC and chronic liver disease. Use of these TDs will help to predict potential loss of liver tissue after SABR., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Purpose: To assess optimal relative prescribed dose values in stereotactic ablative body radiation therapy (SABR) using dynamic conformal multiple arc therapy (DCMAT) for liver tumors., Methods and Materials: We generated SABR plans for 8 typical liver tumors that received SABR with 50 Gy in 5 fractions. The prescribed dose had previously been defined as 80% of the maximal dose ("80% isodose plan"). Alternatively, 20%-90% isodose plans were created to compare dosimetric factors., Results: The mean liver volume values (%) that received >20 Gy (V20) and the mean liver dose were both the lowest with a 70% isodose plan and were the second lowest with a 60% isodose plan. The V20 dose was 5.19% lower (11.14%) with a 70% isodose plan and 4.51% lower (11.22%) with a 60% isodose plan compared with the value with an 80% isodose plan (11.75%). Mean planning target volume (PTV) dose increased as the % isodose decreased. The mean PTV dose was 10% higher (62.4 Gy) with a 70% isodose plan and 21% higher (68.9 Gy) with a 60% isodose plan compared with the value with an 80% isodose plan (56.8 Gy)., Conclusions: During SABR treatment planning using DCMAT for liver tumors, target doses increased as the percentage isodose value decreased, which could result in better outcomes. In contrast, a 70% isodose plan had the lowest normal liver dose and a 60% isodose plan had the second lowest. An optimal percentage isodose level might be adjusted depending on tumor radiation sensitivity and liver function reserve. Further investigation is warranted to determine whether these dosimetric advantages result in improved outcomes., (Copyright © 2014 American Society for Radiation Oncology. Published by Elsevier Inc. All rights reserved.)

Introduction: The maximum standardized uptake value (SUVmax) on F-fluorodeoxyglucose positron emission tomography is a predictor for overall survival (OS) in non-small-cell lung cancer (NSCLC) after resection. We investigated the association between SUVmax and outcomes in NSCLC after stereotactic body radiotherapy., Methods: Between 2005 and 2012, 283 patients with early NSCLC (T1a-2N0M0) were treated with stereotactic body radiotherapy; the total doses were 40 to 60 Gy in five fractions. Patients who underwent staging F-fluorodeoxyglucose positron emission tomography scans by a single scanner and were followed up for more than or who died within 6 months were eligible. The optimal threshold SUVmax was calculated for each outcome. Outcomes were analyzed using the Kaplan-Meier method and log-rank test. Prognostic significance was assessed by univariate and multivariate analyses., Results: One hundred fifty-two patients were eligible. Median follow-up was 25.3 (range, 1.3-77.4) months. Local, regional, and distant recurrences, cancer-specific deaths, and deaths from other reasons occurred in 14, 11, 27, 21, and 31 patients, respectively. The optimal threshold SUVmax for local, regional, and distant recurrences, and disease-free survival (DFS), cancer-specific survival, and OS were 2.47 to 3.64. Outcomes of patients with SUVmax lower than each threshold were significantly better than those with higher SUVmax (all p<0.005): 3-year DFS rates were 93.0% versus 58.3% (p<0.001) and 3-year OS rates were 86.5% versus 42.2% (p<0.001), respectively. By multivariate analysis, higher SUVmax was a significantly worse predictor for DFS (p<0.01) and OS (p=0.04)., Conclusions: SUVmax was a predictor for DFS and OS. A high SUVmax may be considered for intensive treatment to improve outcomes.

Objective: The purpose of this study is to evaluate the CT appearances of tumor responses following hypofractionated stereotactic ablative body radiotherapy for small hypervascular hepatocellular carcinomas (HCCs) and to assess the relationship between tumor responses and local control., Materials and Methods: Among 277 HCC tumors treated with stereotactic ablative body radiotherapy (35 or 40 Gy per five fractions), we selected enhanced lesions on arterial phase CT performed before stereotactic ablative body radiotherapy. Radiographic findings after stereotactic ablative body radiotherapy were evaluated during a 2-year follow-up period with the modified Response Evaluation Criteria in Solid Tumors. Local control and survival rates were calculated with the Kaplan-Meier method., Results: Forty-two tumors with a median size of 2.1 cm (range, 1.0-3.8 cm) were selected with a median follow-up of 23.3 months (range, 9-56 months). Local recurrence was observed in two tumors after achieving a complete response (CR). The 2-year local control rate was 97%, and the overall survival rate was 81%. CR increased from 10 (24%) to 28 (67%) to 30 (71%) tumors at 3, 6, and 12 months after stereotactic ablative body radiotherapy. Overall CR at maximum follow-up was 39 tumors (93%), yet three enhanced tumors persisted for more than 2 years. The median time to achieve CR was 5.9 months (range, 1.2-34.2 months)., Conclusion: The CR rate in hypervascular HCCs after hypofractionated stereotactic ablative body radiotherapy increased during the 2-year follow-up period. Cautious and continuous observation until tumor regrowth is considered relevant to evaluate a true effect of this treatment. Further studies for the optimal evaluation of treatment outcome after stereotactic ablative body radiotherapy are warranted.

Background: For isolated postsurgical local recurrences (IPSLR) of lung cancer, salvage resection is often unfeasible due to a high risk of morbidity and death. Stereotactic ablative body radiotherapy (SABR) provides excellent therapeutic effects, with mild toxicities, for patients with medically inoperable lung cancer. However, the outcomes of SABR for IPSLR have not been reported., Methods: Patients with IPSLR who were treated with SABR between 2005 and 2012 were retrospectively identified. The prescribed doses were 40 to 60 Gy per 5 to 10 fractions. Treatment outcomes and toxicities were evaluated., Results: We identified 23 patients with IPSLR, including 21 with bronchial stump or staple line recurrences and 2 with chest wall recurrences. During follow-up, IPSLR occurred at a median of 36.7 months (range, 5.0 to 190 months) after resection. All patients were N0 M0, and the T stages at recurrence were T1a, T1b, T2a, and T4 in 6, 5, 3, and 9 patients, respectively. The initial pathologic diagnoses were adenocarcinoma in 17 patients and squamous cell carcinoma in 6. At a median follow-up duration of 17.0 months (range, 6.0 to 89.6 months) after SABR, there were 2 local recurrences. Local control and overall survival rates at 1 and 2 years were 94.7% and 86.8% and 84.0% and 76.4%, respectively. Grade 3 to 5 radiation pneumonitis occurred in 1 patient each. Grade 3 temporary but repeated obstructive pneumonia occurred in 2 patients., Conclusions: SABR for IPSLR achieved high local control with limited toxicities. SABR may lead to a potential cure and should be considered as a salvage treatment option for IPSLR., (Copyright © 2013 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Background: The target volume for postoperative breast irradiation is the remaining breast tissue, and the axillary region is not an intentional target volume., Patients and Methods: Between 2001 and 2009, eligible women with pT1-2cN0/pN0(sn) breast cancer underwent breast-conserving therapy without axillary dissection. Treatment outcomes between 2 radiotherapy planning groups, high tangent fields with 2-dimensional (2-D) simulation-based planning and 3-dimensional (3-D) computed tomography-based planning with a field-in-field technique, were compared. The correlating factors for axillary failure were also calculated., Results: In total, 678 patients were eligible. As of May 2009, the median follow-up times for the 2-D (n = 346) and 3-D (n = 332) groups were 94 and 52 months, respectively. Patient characteristics were balanced, except for a younger population in the 2-D group and more lymphovascular invasion in the 3-D group. On multivariate analysis, 2-D planning was the only risk factor for axillary failure. In the 2-D and 3-D groups, the 5-year cumulative incidences of axillary failure were 8 (3.1%) and 1 (0.3%) (log-rank p = 0.009), respectively. The respective 5-year overall survival rates were 97.4 and 98.4% (p = 0.4)., Conclusion: High tangent irradiation with 3-D planning improved axillary control compared to that with 2-D planning, suggesting that optimizing axillary dose distribution may impact outcomes.

We performed a detailed analysis of hysterectomy specimens of uterine cervical cancer to determine the appropriate length of uterine body to include within the clinical target volume. Between 2008 and 2011, 54 patients with uterine cervical carcinoma underwent hysterectomy. Those with quality pre-operative magnetic resonance imaging (MRI) data were included for analysis. Tumor sizes measured by MRI and microscopy were compared with regard to brachytherapy-oriented parameters. Detailed descriptive analysis focusing on the extent of tumor involvement was also performed. A total of 31 specimens were analyzed. The median maximal tumor length measured by MRI was slightly shorter than microscopic length (19 vs. 24 mm, respectively), while the maximal radius was almost identical. No tumors with a maximal size <2 cm by MRI (n = 6) extended to the uterine body ≥ 1/3. The majority of maximal tumor length underestimation on MRI was within 1 cm. Precise tumor delineation can be made by MRI. For patients with tumors <2 cm on MRI, treating the entire uterine body length may not be necessary. A 1-cm margin around an MRI-based gross tumor seems to be adequate to cover the actual tumor involvement.

Purpose: To retrospectively investigate treatment outcomes of stereotactic ablative body radiation therapy (SABR) for octogenarians with non-small cell lung cancer (NSCLC)., Methods and Materials: Between 2005 and 2012, 109 patients aged ≥80 years with T1-2N0M0 NSCLC were treated with SABR: 47 patients had histology-unproven lung cancer; 62 patients had pathologically proven NSCLC. The prescribed doses were either 50 Gy/5 fractions for peripheral tumors or 40 Gy/5 fractions for centrally located tumors. The treatment outcomes, toxicities, and the correlating factors for overall survival (OS) were evaluated., Results: The median follow-up duration after SABR was 24.2 (range, 3.0-64.6) months. Only limited toxicities were observed, except for 1 grade 5 radiation pneumonitis. The 3-year local, regional, and distant metastasis-free survival rates were 82.3%, 90.1%, and 76.8%, respectively. The OS and lung cancer-specific survival rates were 53.7% and 70.8%, respectively. Multivariate analysis revealed that medically inoperable, low body mass index, high T stage, and high C-reactive protein were the predictors for short OS. The OS for the operable octogenarians was significantly better than that for inoperable (P<.01)., Conclusions: Stereotactic ablative body radiation therapy for octogenarians was feasible, with excellent OS. Multivariate analysis revealed that operability was one of the predictors for OS. For medically operable octogenarians with early-stage NSCLC, SABR should be prospectively compared with resection., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Introduction: Several single institution phase I and phase II trials of stereotactic ablative body radiotherapy (SABR) for liver tumors have reported promising results and high local control rates of over 90%. However, there are wide variations in dose and fractionation due to different prescription policies and treatment methods across SABR series that have been published to date.This study aims to assess and minimize inter-institutional variations in treatment planning using SABR for hepatocellular carcinoma (HCC) in preparation for a prospective multi-institutional study., Methods: Four institutions (A-D) participated in this study. Each institution was provided with data from four cases, including planning and diagnostic CT images and clinical information, and asked to implement three plans (a practice plan and protocol plans 1 and 2). Practice plans were established based on the current treatment protocols at each institution. In protocol plan 1, each institution was instructed to prescribe 40 Gy in five fractions within 95% of the planning target volume (PTV). After protocol plan 1 was evaluated, we made protocol plan 2, The additional regulation to protocol plan 1 was that 40 Gy in five fractions was prescribed to a 70% isodose line of the global maximum dose within the PTV. Planning methods and dose volume histograms (DVHs) including the median PTV D50 (Dm50) and the median normal liver volume that received 20 Gy or higher (Vm20) were compared., Results: In the practice plan, Dm50 was 48.4 Gy (range, 43.6-51.2 Gy). Vm20 was 15.9% (range, 12.2-18.9%). In protocol plan 1, the Dm50 at institution A was higher (51.2 Gy) than the other institutions (42.0-42.2 Gy) due to differences in dose specifications. In protocol plan 2, variations in DVHs were reduced. The Dm50 was 51.9 Gy (range, 51.0-53.1 Gy), and the Vm20 was 12.3% (range, 10.4-13.2%). The homogeneity index was nearly equivalent at all institutions., Conclusions: There were notable inter-institutional differences in practice planning using SABR to treat HCC. The range of PTV and normal liver DVH values was reduced when the dose was prescribed to an isodose line within the PTV. In multi-institutional studies, detailed dose specifications based on collaboration are necessary.

Purpose: To evaluate biliary toxicity after stereotactic body radiation therapy (SBRT) for liver tumors., Methods and Materials: Among 297 consecutive patients with liver tumors treated with SBRT of 35 to 50 Gy in 5 fractions, patients who were irradiated with >20 Gy to the central biliary system (CBS), including the gallbladder, and had follow-up times >6 months were retrospectively analyzed. Toxicity profiles, such as clinical symptoms and laboratory and radiologic data especially for obstructive jaundice and biliary infection, were investigated in relation to the dose volume and length relationship for each biliary organ., Results: Fifty patients with 55 tumors were irradiated with >20 Gy to the CBS. The median follow-up period was 18.2 months (range, 6.0-80.5 months). In the dose length analysis, 39, 34, 14, and 2 patients were irradiated with >20 Gy, >30 Gy, >40 Gy, and >50 Gy, respectively, to >1 cm of the biliary tract. Seven patients were irradiated with >20 Gy to >20% of the gallbladder. Only 2 patients experienced asymptomatic bile duct stenosis. One patient, metachronously treated twice with SBRT for tumors adjacent to each other, had a transient increase in hepatic and biliary enzymes 12 months after the second treatment. The high-dose area >80 Gy corresponded to the biliary stenosis region. The other patient experienced biliary stenosis 5 months after SBRT and had no laboratory changes. The biliary tract irradiated with >20 Gy was 7 mm and did not correspond to the bile duct stenosis region. No obstructive jaundice or biliary infection was found in any patient., Conclusions: SBRT for liver tumors adjacent to the CBS was feasible with minimal biliary toxicity. Only 1 patient had exceptional radiation-induced bile duct stenosis. For liver tumors adjacent to the CBS without other effective treatment options, SBRT at a dose of 40 Gy in 5 fractions is a safe treatment with regard to biliary toxicity., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Purpose: Stereotactic body radiotherapy (SBRT) is the standard care for medically inoperable early non-small-cell lung cancer (NSCLC). However, it can be difficult to differentiate local recurrence from non-recurrence radiation-induced lung opacity. We retrospectively assessed (18)F-FDG PET/CT to detect local recurrence after SBRT for NSCLC., Methods: Between 2005 and 2011, 273 NSCLCs in 257 patients were treated with SBRT. Prescribed doses were 50Gy and 40Gy per 5 fractions for peripheral and central lesions, respectively. Tri-monthly follow-up CT scans were acquired. (18)F-FDG PET/CT scans were scheduled for screening at one year after SBRT or when recurrence was highly suspected. The dual-time-point maximum standardized uptake values (SUVmaxs) and their retention indexes (RIs) were obtained., Results: A total of 214 (18)F-FDG PET/CT scans were obtained for 164 localized NSCLC tumors in 154 patients. The median follow-up period was 24.9 months (range: 6.3-72.1). Among these, 21 scans of 17 tumors were diagnosed as local recurrence. The median SUVmaxs on early and late images of recurrence and their RI were 5.0 (range: 3.2-10.7), 6.3 (range: 4.2-13.4), and 0.20 (range; 0-0.41), respectively. These were significantly higher than the respective values of non-recurrence images of 1.8 (range: 0.5-4.6), 1.7 (range: 0.5-6.1), and 0.00 (range: -0.37-0.41) (all p<0.05). For SUVmaxs on early and late images, optimal thresholds were identified as 3.2 and 4.2. Using each threshold, the sensitivity and specificity were 100% and 96-98%, respectively. CT findings were classified into ground-glass opacity (N=9), scar or fibrotic change (N=96), consolidation with air-bronchogram (N=34), consolidation only (N=22), and nodule (N=17); the respective numbers of recurrence were 0, 0, 1, 3, and 17., Conclusion: SUVmaxs of (18)F-FDG PET/CT could detect local recurrence after SBRT for localized NSCLC. In contrast, CT scan results had a limited ability to diagnose local recurrence., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Purpose: We analyzed whether axillary nodal irradiation could control clinically node-negative disease, including those patients with a positive sentinel lymph node biopsy (SLNB), most of whom received regional nodal irradiation. We also evaluated toxicity profiles that resulted from nodal irradiation., Patients and Methods: From 1988 to 2011, 2107 patients with cT1-T2N0M0 breast cancer underwent breast conservation therapy in the absence of axillary dissection: nx group (n = 1548), without any axillary surgery; the sn(-) group (n = 518), with a negative SLNB; and sn(+) group (n = 104), with a positive SLNB., Results: The median follow-up times were 88, 56, and 55 months for the nx, sn(-), and sn(+) groups, respectively. The nx group had more risk factors than did the other 2 groups in terms of age, grade, or T stage. Ninety-eight percent of the sn(-)group received only tangent irradiation, and 100% and 83% of the sn(+) and nx group, respectively, received additional regional nodal irradiation. The 5-year cumulative incidences of axillary failure and regional nodal failure were 34, 3, and 0 (2.7%, 0.7%, and 0%; P = .02, log-rank test) and 57, 4, and 0 (4.4, 1%, and 0; P = .04), respectively. Overall survival rates in 5 years were 96.4%, 98.9%, and 97.6% (P = .03), respectively. Symptomatic but transient radiation pneumonitis developed in 31, 16, and 6 (2.0%, 3.1%, and 5.7%). Mild arm edema was observed in 1, 4, and 0 (0.06%, 0.8%, and 0%) in the nx, sn(-), sn(+) groups, respectively., Conclusions: Treatment without axillary dissection showed excellent outcomes with negligible toxicity for patients with clinically node negative, including those with a positive SLNB. Regional nodal irradiation after a positive SLNB is a reasonable alternative to axillary dissection., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Purpose: To investigate threshold dose (TD) of focal liver reaction (FLR) following stereotactic body radiotherapy (SBRT) for patients with hepatocellular carcinoma (HCC) and liver cirrhosis., Materials and Methods: In consecutive 50 patients receiving SBRT for small HCC, 38 patients receiving SBRT and follow up >6 months, FLR on follow-up CT had been previously studied. Patients with good concordance between FLR and highly irradiated area were eligible. Dose volume histogram (DVH) was used to identify TDs for FLR. Clinical factors were analyzed for correlation with TDs., Results: Of 24 eligible patients, 23 had Child-Pugh score A and 1 scored B. Presence of FLR peaked at a median of 6 (range; 3-12) months. The median and 95% confidential intervals of TDs of pre-contrast and portal-venous phase CT were 32.4 Gy (30.3-35.4) and 34.4 Gy (31.9-36.0), respectively. Each median coefficient representing the concordance was 74.9% (range; 55.8-98.0%) and 80.5% (range; 70.8-92.4%), respectively. No clinical factors significantly correlated with the TDs., Conclusion: We proposed 30 Gy/5 fractions as TD of FLRs following SBRT for patients with HCC and liver cirrhosis. This TD will enable us to predict injured liver volume and to avoid complication beforehand from toxicity. Further pathological and clinical studies, in addition to more practical and precise data of DVH, are needed to clarify the significance of FLRs., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Introduction: In non-surgical candidates with solitary pulmonary nodules (SPNs) and no histological confirmation, optimal management remains uncertain., Methods: Between February 2005 and February 2011 we treated 298 lung cancers with stereotactic body radiotherapy (SBRT), including SPNs clinically diagnosed as lung cancer (CDLC). Among them, we extracted patients treated with a total dose of 40-50 Gy per 5 fractions and followed up more than 6 months. Patients who had a history of previously treated lung cancer, or were diagnosed pathologically, or suspected as having small-cell lung cancer or large cell neuroendcrine cancer were excluded from this study. The remaining patients were divided into two groups; CDLC and non-small-cell lung cancer (NSCLC) patients and their outcomes were assessed and compared., Results: Fifty-eight CDLC and 115 NSCLC patients were included in this study. The proportions of female and inoperable cases were significantly higher in the CDLC group. Other characteristics, including T stage and standard uptake value, were well balanced. Median follow-up durations were 20.2 (range, 6.0-58.8) and 21.2 (range, 6-63.7) months, respectively. The 3-year local control, regional-free, metastasis-free, progression-free, cause-specific survival, and overall survival rates were 80% and 87% (p = 0.73), 88% and 91% (p = 0.72), 70% and 74% (p = 0.57), 64% and 67% (p = 0.45), 74% and 71% (p = 0.17), 54% and 57% (p = 0.48), respectively., Conclusion: These results indicate that the treatment outcome of CDLC group was almost identical to that of NSCLC and that few benign lesions seemed to be included. We advocate that SBRT can be legitimately applied to CDLC, provided that they are carefully diagnosed by integrating various clinical findings., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Background and Purpose: The purpose of this study was to evaluate the radiotherapy (RT) quality assurance (QA) for JCOG 0303., Methods and Materials: JCOG 0303 was a multi-center phase II/III trial that compared two types of chemotherapy administered concomitantly with RT for locally advanced esophageal cancer. RT requirements included a total dose of 60 Gy in 30 fractions and CTV with a 2-cm margin cranio-caudally to the primary tumor. The QA assessment was given as per protocol (PP), deviation acceptable (DA), violation unacceptable (VU), and incomplete/not evaluable following predefined criteria for quality parameters., Results: A total of 142 cases were accrued. After excluding 36 incomplete/not evaluable, 106 (75%) were fully evaluable for RT quality review. Of these 106, there were 4 VU (4%) and overall RT compliance (PP + DA) was 96%. Comparing the incidence of VU based on the numbers enrolled by institution, the highest quarter of enrollment (≥7 cases) had no VU, while all VU (4; 11%) were from institutions enrolling <7 patients., Conclusions: The results of the RTQA assessment for JCOG 0303 were sufficient to provide reliable results. Additional improvements will be needed for institutions with low accrual rates.

We evaluated associations of interstitial changes with radiation pneumonitis (RP) for patients treated with thoracic radiotherapy. Between 2005 and 2009, patients who received thoracic radiotherapy of 40 Gy or more for lung cancer or thymic tumors and were followed-up for more than 6 months were eligible for this study. Possible risk factors for RP included the presence of interstitial changes on computed tomography before radiotherapy, and elevated C-reactive protein (CRP) and lactate dehydrogenase (LDH) levels; these were compared with the incidences of severe RP. A total of 106 patients were included. The incidences of RP were 4 (4%), 0 (0%), and 5 (5%) for grades 3, 4, and 5, respectively. For those with interstitial changes, the incidence of RP ≥ grade 3 was significantly increased from 3% (2/79) to 26% (7/27) (p < 0.001). CRP and LDH levels were also associated with increased RP, as were pulmonary emphysema and performance status ≥ 2. Among 91 patients with RP ≥ grade 1, RP grade ≥ 3 occurred significantly earlier than grades 1 and 2. In conclusion, pulmonary interstitial changes, LDH and CRP levels, pulmonary emphysema, and performance status ≥ 2 were significantly associated with RP ≥ grade 3. RP grade ≥ 3 occurred significantly earlier than grades 1 and 2. The early appearance of interstitial changes requires careful management due to the possibility of severe RP.

Background: The maximum standardized uptake value (SUVmax) of FDG-PET may predict local recurrence for localized non-small-cell lung cancer (NSCLC) after stereotactic body radiotherapy (SBRT)., Methods: Among 195 localized NSCLCs that were treated with total doses of either 40Gy or 50Gy in 5 SBRT fractions, we reviewed those patients who underwent pre-treatment FDG-PET using a single scanner for staging. Local control rates (LCRs) were obtained by the Kaplan-Meier method and a log-rank test. Prognostic significance was assessed by univariate and multivariate analyses., Results: A total of 95 patients with 97 lesions were eligible. Median follow-up was 16.0months (range: 6.0-46.3months). Local recurrences occurred in 9 lesions. By multivariate analysis, only the SUVmax of a primary tumor was a significant predictor (p=0.002). Two years LCRs for lower SUVmax (<6.0; n=78) and higher SUVmax (⩾6; n=19) were 93% and 42%, respectively. In subgroups with T1b and T2, LCRs were significantly better for lower SUVmax than for higher SUVmax (p<0.0005 and p<0.01). In both subgroups that received 40Gy and 50Gy, LCRs were also significantly better for lower SUVmax than for higher SUVmax (p<0.001 and p<0.01)., Conclusions: SUVmax was the strongest predictor for local recurrence. A high SUVmax may be considered for dose escalation to improve local control. Additional follow-up is needed to determine if SUVmax is correlated with regional recurrence, distant metastasis, and survival., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Purpose: To investigate factors associated with Grade >or=3 radiation pneumonitis (RP) in patients with lung tumors treated with stereotactic body radiotherapy (SBRT)., Methods and Materials: We retrospectively analyzed 128 patients with 133 lung tumors treated with SBRT. RP was graded according to the Common Terminology Criteria for Adverse Events version 3.0. Univariate analyses were used to identify predictive factors for RP., Results: The median follow-up period after SBRT was 12 months (range, 5-45 months). Incidences of Grades 0, 1, 2, and 3 RP were 27%, 52%, 16%, and 5%, respectively. No patients suffered Grade 4 or 5 RP. For all patients with Grade 2 or 3, symptoms occurred either simultaneously with or subsequent to graphical appearances. The latent period was the only significant factor associated with Grade >or=3 RP (p < 0.01). A latent period of 1 or 2 months indicated a 40% (6/15) risk for Grade 3. However, the risk for Grade 3 was 1.2% (1/82) 3 months after SBRT. No pretreatment clinical or dosimetric factors were significantly associated with Grade >or=3 RP. However, 4 of 7 patients with Grade 3 RP had severe pulmonary comorbidities., Conclusion: Only the latency period was a significant factor in the development of RP. No pretreatment clinical or dosimetric factors were significantly associated with Grade >or=3 RP. Patients, especially those with severe pulmonary comorbidities, should be carefully observed for the graphical appearance of RP within a few months during the follow-up period after SBRT., ((c) 2010 Elsevier Inc. All rights reserved.)

Cold acclimation requires substantial alteration in membrane property. In contrast to well-documented fatty acid unsaturation during cold acclimation, changes in phospholipid biosynthesis during cold acclimation are less understood. Here, we isolated and characterized two aminoalcoholphosphotransferase (AAPT) cDNAs, TaAAPT1 and TaAAPT2, from wheat. AAPTs utilize diacylglycerols and CDP-choline/ethanolamine as substrates and catalyze the final step of the CDP-choline/ethanolamine pathway for phosphatidylcholine (PC)/phosphatidylethanolamine (PE) synthesis, respectively. Functionality of TaAAPT1 and TaAAPT2 was demonstrated by heterologous expression in a yeast cpt1Delta ept1Delta double mutant that lacks both AAPT activities. Detailed characterization of AAPT activities from the transformed mutant cells indicated that TaAAPT1 is an ECPT-type enzyme with higher ethanolamine phosphotransferase (EPT) activity than choline phosphotransferase (CPT) activity, while TaAAPT2 is a CEPT-type with the opposite substrate preference. Transient expression of GFP-fused TaAAPT1 and TaAAPT2 proteins in wheat and onion cells indicated they are localized to both the endoplasmic reticulum and Golgi apparatus, suggesting that the final synthesis of PE and PC via the CDP-choline/ethanolamine pathway occurs in these organella. Quantitative PCR analyses revealed that TaAAPT1 expression is strongly induced by cold, while TaAAPT2 was constitutively expressed at lower levels. Measurement of phospholipid content in wheat leaves indicated that PE is more prominently increased in response to cold than PC and accordingly PE/PC ratio increased from 0.385 to 0.530 during 14 days of cold acclimation. Together, these data suggested that an increase in the PE/PC ratio during cold acclimation is regulated at the final step of the biosynthetic pathway.

Purpose: We have used dynamic conformal multiple arc therapy (DCMAT) for stereotactic body radiotherapy (SBRT) since 2001. We investigated the consistency of DCMAT for SBRT using dose-volume histogram analysis., Methods and Materials: A total of 50 patients with peripheral lung tumors underwent SBRT. The median tumor diameter was 2.4 cm (range, 0.9-5.9). Treatment planning was performed using a superposition algorithm. The prescribed 50 Gy dose was divided into five fractions. The prescribed dose was defined as 80% of the maximal dose in the planning target volume (PTV), and the leaf margins were modified to ensure the PTV was included in the 80% isodose surface. The dose-volume histogram analysis was used to assess the PTV and normal lung volume., Results: The median dose covering 95% of the PTV was 50.27 Gy (range, 46.14-52.67), essentially consistent with the prescribed dose. The median homogeneity and conformity index was 1.41 (range, 1.31-1.53) and 1.73 (range, 1.41-2.21), respectively. The median volume of lung receiving > or =20 Gy (V(20)) was 4.2% (range, 1.4-10.2%). A linear correlation was found between the tumor diameter and V(20), and an even stronger correlation was found between the PTV/(normal lung volume) and V(20). The estimated V(20) was 7.1% (range, 3.9-10.4%) for a 5-cm-diameter tumor, assumed to be the maximal size limitation for SBRT., Conclusion: SBRT with DCMAT achieved high conformity and delivered adequate doses within the PTV. The median dose covering 95% of the PTV was consistent with the prescribed dose. V(20) can be estimated using the tumor diameter and normal lung volume. DCMAT was thus both a feasible and a reproducible method of SBRT delivery.

Purpose: To retrospectively analyze the clinical outcomes of stereotactic body radiotherapy (SBRT) for patients with Stages 1A and 1B non-small-cell lung cancer., Methods and Materials: We reviewed the records of patients with non-small-cell lung cancer treated with curative intent between Dec 2001 and May 2007. All patients had histopathologically or cytologically confirmed disease, increased levels of tumor markers, and/or positive findings on fluorodeoxyglucose positron emission tomography. Staging studies identified their disease as Stage 1A or 1B. Performance status was 2 or less according to World Health Organization guidelines in all cases. The prescribed dose of 50 Gy total in five fractions, calculated by using a superposition algorithm, was defined for the periphery of the planning target volume., Results: One hundred twenty-one patients underwent SBRT during the study period, and 63 were eligible for this analysis. Thirty-eight patients had Stage 1A (T1N0M0) and 25 had Stage 1B (T2N0M0). Forty-nine patients were not appropriate candidates for surgery because of chronic pulmonary disease. Median follow-up of these 49 patients was 31 months (range, 10-72 months). The 3-year local control, disease-free, and overall survival rates in patients with Stages 1A and 1B were 93% and 96% (p = 0.86), 76% and 77% (p = 0.83), and 90% and 63% (p = 0.09), respectively. No acute toxicity was observed. Grade 2 or higher radiation pneumonitis was experienced by 3 patients, and 1 of them had fatal bacterial pneumonia., Conclusions: The SBRT at 50 Gy total in five fractions to the periphery of the planning target volume calculated by using a superposition algorithm is feasible. High local control rates were achieved for both T2 and T1 tumors.

We have developed a new target plate for matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS). This target plate enables direct electric transfer of analytes from the 1-dimensional gel electrophoresis (1-DE) gel to the target plate in one step. Incorporated with a one-step direct transfer technique, this novel 1-DE/MALDI-MS (1-DE/MS) system eliminates staining, extracting, loading, and many other time-consuming intermediate processes, thereby greatly reducing analysis time while providing high throughput proteome analysis. Furthermore, in peptidome analysis, during the 1-DE step this system separates or removes the high molecular weight plasma proteins in blood and the various low molecular weight substances in tissue extracts, which interfere with mass spectrometry. This system can therefore be used for peptide profiling of any biological sample without special pretreatment. In view of these advantages, the 1-DE/MS system will greatly improve the usefulness of current peptidomic modalities in the discovery and validation of biomarker molecules in various body fluids and tissue extracts, permitting early detection, diagnosis, and treatment of diseases.

Purpose: In our institution a CT scanner was installed in the same room as the linear accelerator. In stereotactic body radiotherapy (SBRT) we confirmed the isocenter position by serial thin-slice and long-scan-time CT images before every treatment as well as in planning. In planning we constructed digitally reconstructed radiography (DRR) of both the anterior and lateral views. At the first treatment we also checked the isocenter with linacgraphy. Then we compared the isocenter positions obtained from the DRR and linacgraphy., Materials and Methods: Between Feb. 2005 and Oct. 2006, we treated 75 lung and liver tumors with SBRT in this way. Based on bony structures, we measured the differences between in-isocenter positions for SI, LR, and AP directions between DRR and linacgraphy., Results: The median (min-max) of the differences in-isocenter positions for SI, LR, and AP directions between DRR and linacgraphy were 0.0 mm (0-6.0), 0.0 mm (0-10.0), and 0.0 mm (0-10.0), respectively, as well as 3.2 mm (0-12.3) for 3-dimensional distance. In 28 tumors (37%) the differences exceeded 5 mm in three-dimensional distance. The frequency of differences exceeding 5 mm in upper lung lesions tended to be more than that in liver lesions, and that in left pulmonary lesions was significantly more than that in right ones., Conclusion: This result suggests that the relative position of the target volume to the bony structure differ in planning and in every treatment. It was recommended to verify isocenter accuracy in institutions where isocenter position is checked only by orthogonal linacgraphy in SBRT.

Purpose: We previously reported that most of axillary regions could be irradiated by the modified tangential irradiation technique (MTIT). The purpose of this study was to determine whether the three-dimensional conformal radiotherapy (3D-CRT) with a field-in-field technique improves dosimetry for the breast and axillary nodes., Methods and Materials: Fifty patients with left-sided breast cancer were enrolled. With MTIT, we planned the radiation field to be wider in the cranial direction than the standard tangential fields to include the axillary regions. With 3D-CRT, a field-in-field technique was used to spare the heart and contralateral breast to the extent possible by applying the multileaf collimator manually. Dose-volume histograms were compared for the breast, axillary region, heart, lung, and other normal tissues., Results: There were no significant differences in the percent volume of the breast receiving >90% of the prescribed dose (V90) between MTIT and 3D-CRT. The mean V90 of the level I to III axillary regions were increased from 93.7%, 48.2%, and 41.3% with MTIT to 97.6%, 85.8%, and 82.8% with 3D-CRT. 3D-CRT significantly reduced the volume of the heart receiving >30 Gy (mean, 7.6 vs. 15.9 mL), the percent volume of the bilateral lung receiving >20 Gy (7.4% vs. 8.9%), and the volume of other normal tissues receiving >107% of the prescribed dose (0.1 vs. 2.9 mL)., Conclusion: The use of 3D-CRT with a field-in-field technique improves axillary node coverage, while decreasing doses to the heart, lungs, and the other normal tissues, compared with MTIT.

Hypofractionated stereotactic body radiotherapy (SBRT) for pulmonary lesions provides a high local control rate, allows completely painless ambulatory treatment, and is not associated with adverse reactions in most cases. Here we report a 70-year-old lung cancer patient with slight focal pulmonary honeycombing in whom subclinical idiopathic pulmonary fibrosis was exacerbated by SBRT. This experience has important implications for the development of selection criteria prior to SBRT for pulmonary lesions. For SBRT candidates with lung tumors, attention must be paid to the presence of co-morbid interstitial pneumonia even if findings are minimal. Such patients must be informed of potential risks, and careful decision-making must take place when SBRT is being considered.

Purpose: To retrospectively analyze opacity changes near primary lung cancer tumors irradiated by using hypofractionated stereotactic radiotherapy (HSRT) to determine the presence or absence of tumor recurrence., Methods and Materials: After review-board approval for a retrospective study, we examined data from 50 patients treated with curative intent for proven or highly suspected localized peripheral-lung cancer and followed up for at least 12 months. All patients had received 50 Gy in five fractions (80% isodose) and were followed up monthly with chest X-ray until clinical and X-ray findings stabilized. Follow-up computed tomography scans were performed 1 and 3 months after HSRT and thereafter at 3-month intervals during the first 2 years., Results: Median follow-up was 30.4 months (range, 12.0-73.8 months). Abnormal opacities that were suspicious for recurrent tumor appeared in 20 patients at a median of 20.7 months (range, 5.9-61.4 months). Only 3 patients were finally found to have recurrence; 14 were recurrence free but were suspected to have fibrosis, and findings for the other 3 patients were considered equivocal because of a short follow-up period (

Aim: To investigate the efficacy and toxicity of hypofractionated stereotactic radiotherapy for the treatment of patients presenting with hepatocellular carcinoma (HCC) in a single institutional setting., Methods: Sixteen patients who presented with solitary HCC, including two patients with a tumor thrombus of the portal veins, were treated with stereotactic radiotherapy with or without transarterial chemoembolization. The criteria for stereotactic radiotherapy were existence of technical difficulties for other ablation therapies, inoperable disease or refusal to undergo surgery, tumor staged as Grade A or B according to the Child-Pugh classification, and solitary tumor distant from the gastrointestinal tract and kidney with a tumor volume <100 cm(3). In 14 of 16 patients, a total dose of 35- 50 Gy was delivered in 5-7 fractions over 5-9 days., Results: At the end of a mean follow-up of 612 days (median 611 days; range 244-994 days), all patients were alive. Eight of 16 patients had complete responses and seven others were judged as stable with lipiodol accumulation. In one patient, local recurrence developed after 489 days. Intrahepatic recurrences developed outside the treated volume in six patients and no extrahepatic metastases developed during follow-up. No serious treatment-related toxic manifestations developed., Conclusions: Stereotactic radiotherapy for HCC with or without transarterial chemoembolization is feasible therapy and provides good local control with a short treatment period. Stereotactic radiotherapy may be of clinical benefit in patients who are inoperable or for whom there are difficulties in other ablation therapies.

The authors critically reviewed previous articles concerning the significance of breast irradiation following breast-conserving surgery in terms of the following subject items: indications for breast-conserving therapy, the significance and complications of breast irradiation, the timing of the start of breast irradiation, the significance of boost irradiation, the potential improvement of survival with systemic therapy plus breast irradiation, the significance of axillary dissection, indications and the significance of regional nodal irradiation, accelerated hypofractionated radiotherapy, omission of breast irradiation in low-risk patients, and future directions. In addition, our previously reported results of breast irradiation following breast-conserving surgery at the Keio University Hospital are outlined. Our newly developed tangential irradiation technique directed to the axilla and a recently introduced three-dimensional simulation technique for radiotherapy treatment planning are also presented.

Studies of carrot embryogenesis have suggested that abscisic acid (ABA) is involved in somatic embryogenesis. A relationship between endogenous ABA and the induction of somatic embryogenesis was demonstrated using stress-induced system of somatic embryos. The embryonic-specific genes C-ABI3 and embryogenic cell proteins (ECPs) were expressed during stress treatment prior to the formation of somatic embryos. The stress-induction system for embryogenesis was clearly distinguished by two phases: the acquisition of embryogenic competence and the formation of a somatic embryo. Somatic embryo formation was inhibited by the application of fluridone (especially at 10(-4) M), a potent inhibitor of ABA biosynthesis, during stress treatment. The inhibitory effect of fluridone was nullified by the simultaneous application of fluridone and ABA. The level of endogenous ABA increased transiently during stress. However, somatic embryogenesis was not significantly induced by the application of only ABA to the endogenous level, in the absence of stress. These results suggest that the induction of somatic embryogenesis, in particular the acquisition of embryogenic competence, is caused not only by the presence of ABA but also by physiological responses that are directly controlled by stresses.

Purpose: To determine the cDNA sequences and analyze the expression of porcine optineurin and myocilin in trabecular meshwork cells (TMCs) and astrocytes from the optic nerve head under normal and experimental conditions., Methods: Both porcine optineurin and myocilin were cloned to determine the cDNA sequences. Porcine TMCs and astrocytes were isolated and treated with dexamethasone (500 nM) for 2 weeks, incubated under hypoxic conditions (7% O(2)) for 72 hours, or exposed to 33 mm Hg hydrostatic pressure for 72 hours. A 10% mechanical stretch for 24 hours was also performed on TMCs. The expression level of the optineurin and myocilin transcripts was analyzed by real-time quantitative PCR., Results: The sequences of porcine optineurin and myocilin cDNA were determined, and the expression of both genes was confirmed in both TMCs and astrocytes. Amino acid sequences of porcine optineurin and myocilin were homologous to those of humans by 84% and 82%, respectively, and shared protein motifs and modification sites. The expression of myocilin mRNA by TMCs and astrocytes was increased by 8.0- and 5.5-fold, respectively, after exposure to dexamethasone. In contrast, the expression of optineurin was suppressed to 68% in TMCs and 48% in astrocytes after exposure to dexamethasone. A significant reduction of myocilin expression was observed after 72 hours of incubation under hypoxic conditions in both types of cells, whereas optineurin was not affected. Hydrostatic pressure for 72 hours and mechanical stretching for 24 hours had minimal affects on gene expression of both optineurin and myocilin., Conclusions: The high homology of porcine optineurin and myocilin to the comparable human genes indicates that pigs can be used to study changes in gene expression in hypertensive eyes. The alterations in expression of myocilin but not of optineurin under stress suggest that different mechanisms in the phenotype of glaucoma associated with the two genes are involved in development of glaucoma.