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Objectives: To evaluate the outcomes of robot-assisted partial nephrectomy (RAPN) in cystic renal tumors. Materials and Methods: We retrospectively analyzed patients who underwent RAPN for either c...

Alanine-serine-cysteine transporter 2 (ASCT2) has been associated with increased levels of metabolism in various malignant tumors. However, its biological significance in the proliferation of prostate cancer (PCa) cells remains under investigation. We used the cBioPortal database to assess the effect of ASCT2 expression on the oncological outcomes of 108 PCa patients. To evaluate the function of ASCT2 in castration-sensitive PCa (CSPC) and castration-resistant PCa (CRPC), LNCaP cells and the ARV7-positive PCa cell line, 22Rv1, were assessed using cell proliferation assays and Western blot analyses. The ASCT2 expression level was associated with biochemical recurrence-free survival after prostatectomy in patients with a Gleason score ≥ 7. In vitro experiments indicated that the growth of LNCaP cells after combination therapy of ASCT2 siRNA and enzalutamide treatment was significantly reduced, compared to that following treatment with enzalutamide alone or ASCT2 siRNA transfection alone (p < 0.01, 0.01, respectively). After ASCT2 inhibition by siRNA transfection, the growth of 22Rv1 cells was significantly suppressed as compared with negative control siRNA via downregulation of ARV7 both in fetal bovine serum and androgen-deprivation conditions (p < 0.01, 0.01, respectively). We demonstrated that ASCT2 inhibition significantly reduced the proliferation rates of both CSPC and CRPC cells in vitro.

To compare perioperative and long-term oncological outcomes and recurrence patterns between robot-assisted radical cystectomy with intra-corporeal urinary diversion (iRARC) and open radical cystectomy (ORC). We retrospectively analyzed 177 bladder cancer patients who received iRARC or ORC at Fujita Health University between 2008 and 2020. Our primary endpoint was long-term oncological outcomes. As a secondary endpoint, we examined perioperative outcomes, complications, and recurrence patterns. These outcome measures were compared between the propensity score (PS)-matched cohorts. PS-matched analysis resulted in 60 matched pairs from iRARC and ORC groups. The iRARC cohort was associated with significantly longer operative time (p = 0.02), lower estimated blood loss (p

The significance of BRCA alterations has been implicated in the development of metastatic castration-resistant prostate cancer (PC). On the other hand, the details of the frequency and significance of BRCA alterations in localized PC remain unknown. In this study, we investigated the frequency and clinical significance of BRCA alterations in localized PCs using an in-house next-generation sequencer (NGS) system. NGS analyses revealed that the BRCA2 CN decreased in 17 patients (13.5%) and the BRCA1 CN in six (4.8%). NGS-based CN values were shown to be highly correlated with droplet digital PCR-based CN values. Tissue-specific BRCA expression investigated using the Human Protein Atlas showed that the decreased CN of BRCA2, but not BRCA1, is responsible for the decreased BRCA activity in PC. Ten of the 22 patients with decreased BRCA2 CN were presumed to have somatic heterozygous deletion. There were no observed associations between the heterozygous deletion of BRCA2 and various clinicopathological parameters. Furthermore, three of 10 patients developed biochemical recurrence within 3 months after surgery. Multivariate analyses revealed that the initial prostate-specific antigen levels and BRCA2 CN were independent factors for biochemical recurrence. Our results suggest that a decrease in BRCA2 CN may be used as a biomarker for predicting recurrence after surgery in localized PC. Early screening for somatic alterations in BRCA2 using NGS may help to broadly predict the risk of PC progression. Citation Format: Makoto Sumitomo, Takuhisa Nukaya, Eiji Sugihara, Mayu Takeda, Sachio Nohara, Shigeki Tanishima, Masashi Takenaka, Kenji Zennami, Kiyoshi Takahara, Ryoichi Shiroki, Hideyuki Saya. Determination of BRCA2 deletion status by copy number estimation using next generation sequencing to predict prognosis of clinically localized prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6097.

Indoleamine 2,3‐dioxygenase 1 (IDO1) is a key enzyme associated with immunomodulation through its regulation of the tryptophan‐kynurenine (Kyn) pathway in advanced cancers, including metastatic renal cell carcinoma (mRCC). However, the failure of IDO1 inhibitors when used in combination with immune checkpoint inhibitors (ICIs), as observed in clinical trials, raises a number of questions. This study aimed to investigate the association of tryptophan 2,3‐dioxygenase (TDO) and IDO1 with cancer development and resistance to immunotherapy in patients with RCC. In our analysis of RCC tissue samples, tissue Kyn levels were elevated in advanced‐stage RCC and correlated well with TDO expression levels in RCC tumor cells. In patients with mRCC, TDO rather than IDO1 was expressed in RCC tumor cells, showing a strong association with Kyn expression. Furthermore, immunohistochemical staining of TDO was strongly associated with the staining intensity of forkhead box P3, as well as ICI therapy response and survival in patients with mRCC. Our study is the first to show that TDO expression in tumor tissues is associated with progression and survival, confirming its potential as a predictive biomarker of primary resistance to immunotherapy in patients with mRCC. Our findings suggest that strategies aimed at inhibiting TDO, rather than IDO1, in combination with ICI therapy may aid in the control of mRCC progression., This study aimed to investigate the association of tryptophan 2,3‐dioxygenase (TDO) and indoleamine 2,3‐dioxygenase 1 (IDO1) with cancer development and resistance to immunotherapy in renal cell carcinoma (RCC) patients. Our results suggest that TDO expression in tumor cells is associated with kynurenine accumulation, progression, and survival, confirming its potential as a predictive biomarker of primary resistance to immunotherapy in RCC patients. We believe that our study makes a significant contribution to the literature because, to the best of our knowledge, it is the first to show that the accumulation of kynurenine induced by TDO in tumor tissues is associated with progression and survival. Further, we believe that this paper will be of interest to the readership of your journal because our results strongly recommend the testing of specific TDO inhibitors or dual inhibitors of IDO1 and TDO with ICI treatment in future preclinical and clinical trials focusing on mRCC.

Objectives To assess the impact of two cycles of neoadjuvant chemotherapy (NAC) in patients who underwent nephroureterectomy for high-risk cN0M0 upper tract urothelial carcinoma (UTUC), and to evaluate the efficacy of NAC in patients with localised disease (≤cT2). Patients and methods We retrospectively analysed patients with high-risk cN0M0 UTUC who received NAC followed by surgery, compared with a matched cohort who underwent initial surgery at Fujita Health University during 2005-2019. Baseline and tumour characteristics, overall survival (OS), cancer-specific survival (CSS), and recurrence-free survival (RFS) were compared between the cohorts. Cox proportional hazards models were used to identify predictors of survival. Results There were 117 and 67 patients in the study group and the control group, respectively. Significantly higher pathological downstaging (pDS) and lower lymphovascular invasion (LVI) were observed in the study group than in the control group (48% vs 22%, P = 0.008 and 29% vs 46%, P = 0.045, respectively). The NAC group had significantly better 5-year OS (79% vs 53%, P = 0.003), 5-year CSS (84% vs 66%, P = 0.008), and 5-year RFS (80% vs 61%, P = 0.001) than the control group. The OS benefit of NAC was observed even in patients with localised (≤cT2) disease (P = 0.019). Patients with LVI showed significantly worse CSS both in pathologically locally advanced (≥pT3) and in localised (≤pT2) tumours (P = 0.048 and P = 0.018, respectively). Multivariate analysis identified LVI, NAC, and pDS as independent predictors of OS. Male sex and post-NAC LVI were identified as predictors of worse survival in patients who underwent NAC. Conclusions Two cycles of NAC improved the survival of patients with high-risk UTUC, even in patients with localised disease. Although two cycles of NAC appear to be effective in cN0M0 high-risk UTUC including localised disease, additional larger sample size multicentre prospective studies comparing short-course NAC regimens followed by surgery and surgery alone are required.

This study aimed to compare the perioperative and long-term functional outcomes between robot-assisted partial nephrectomy (RAPN) and open partial nephrectomy (OPN) in Japanese patients.We retrospectively analyzed 242 patients who underwent either RAPN or OPN between 2007 and 2017 at our hospital. Propensity score matching was carried out between the two groups at a ratio of 1:1. Perioperative outcomes and postoperative estimated glomerular filtration rates (eGFR) were compared at one and three years of follow-up.After propensity score matching, we evaluated 39 patients from each group. The ischemia duration of the RAPN group was significantly shorter than that of the OPN group (18 vs. 24,RAPN resulted in shorter ischemia durations and lower rates of EBL than did OPN; however, no differences in long-term renal function were observed between RAPN and OPN in our propensity-score matched Japanese cohort.

The incidence of secondary bladder cancer after treatment for localized prostate cancer (PCa) remains unclear. In this study, PCa cases treated with brachytherapy (BT) were evaluated to assess the incidence of a second malignancy of bladder cancer in a Japanese cohort.Overall, 969 patients treated with BT at our hospital between July 2006 and January 2019 were included in the study cohort. The incidence and predictors of secondary bladder cancer were also assessed.The incidence of secondary bladder cancer was 1.5% (n = 14). Of the seven factors (age, pretreatment PSA, Gleason score, cTNM stage, prostate volume, total activity, and combined external beam), prostate volume and total activity showed significant differences between the cohorts with and without secondary bladder cancer (P = .03 and P = .001, respectively). Upon comparison of the seven parameters for the 969 patients treated with BT, we found that only the total activity factor was affected by the incidence of secondary bladder cancer in the multivariate analysis (P = .007).The incidence of secondary bladder cancer was evaluated after BT for PCa. Total activity was associated with the incidence of secondary bladder cancer in Japanese patients who received BT.

INTRODUCTION AND OBJECTIVE:The glutamine (Gln) pathway has been implicated in tumor progression in various malignancies including renal cell carcinoma (RCC). However, the detail underlying this pat...

INTRODUCTION AND OBJECTIVE:Indoleamine 2,3-dioxygenase 1 (IDO1) is a key enzyme associated with immunomodulation through its regulation of the tryptophan (Trp)-kynurenine (Kyn) pathway in advanced ...

This study aimed to assess the risks and benefits of selective clamping in robot-assisted partial nephrectomy (RAPN). We retrospectively analyzed 372 patients who had undergone RAPN at our hospital between July 2010 and March 2021. After propensity score matching between the full and selective clamping groups, perioperative outcomes and postoperative preservation ratio of the estimated glomerular filtration rate (eGFR) were compared at 6 and 12 months of follow-up. After propensity score matching, we evaluated 47 patients from each group. While no significant differences were observed in surgical time, warm ischemia time, or incidence rates of all grades of complications between the two cohorts, the estimated blood loss (EBL) was significantly lower in the full clamping group than in the selective clamping group (30 vs. 60, p = 0.046). However, no significant intergroup differences were observed in the postoperative preservation ratio of eGFR at 6 or 12 months of follow-up (full clamping 94.0% vs. selective clamping 92.7%, p = 0.509, and full clamping 92.0% vs. selective clamping 91.6%, p = 0.476, respectively). Selective clamping resulted in higher EBL rates than did full clamping in RAPN. However, selective clamping provided no renal functional advantage over full clamping in our propensity-score-matched cohort.

To evaluate the risk factors for postoperative ileus in patients who underwent robot-assisted radical cystectomy with intracorporeal urinary diversion.We retrospectively analyzed 78 patients with bladder cancer who underwent robot-assisted radical cystectomy with intracorporeal urinary diversion at Fujita Health University between 2011 and 2021. Baseline characteristics and perioperative outcomes were compared between the cohorts with and without ileus. Logistic regression analysis was used to identify the risk factors for postoperative ileus.Out of the 78 patients included in this study, 20 (25.6%) developed postoperative ileus. The ileus cohort was associated with a significantly lower Geriatric-8 score (P = 0.003) and a higher rate of previous abdominal/pelvic surgery (P = 0.04) compared with those of the nonileus cohort. Significantly longer intestinal tract reconstruction time, hospital stay, time to mobilization, fluid intake, solid intake, flatus, and stool were observed in the ileus cohort. According to the results of the logistic regression analysis, the Geriatric-8 sum (P = 0.009), time to mobilization (P = 0.03), and time to fluid intake (P = 0.004) were independent predictors of postoperative ileus. In the model predicting postoperative ileus, the area under the receiver operating characteristic curve was 0.716, and the cutoff value of the Geriatric-8 sum was 13.Early mobilization and fluid intake and low Geriatric-8 scores were significant risk factors for postoperative ileus. Preoperative Geriatric-8 evaluation is a useful tool for predicting postoperative ileus. Comprehensive enhanced recovery after surgery, including key components, may help bowel recovery and prevent subsequent ileus.

Background Kidney transplant recipients are at increased risk of developing cancer in comparison with the general population. To effectively manage post-transplantation malignancies, it is essential to proactively monitor patients. A long-term intensive screening program was associated with a reduced incidence of cancer after transplantation. This study evaluated the usefulness of the gene expression profiling of peripheral blood samples obtained from kidney transplant patients and adopted a screening test for detecting cancer of the digestive system (gastric, colon, pancreas, and biliary tract). Study Design and Method Nineteen patients were included in this study and a total of 53 gene expression screening tests were performed. The gene expression profiles of blood-delivered total RNA and whole genome human gene expression profiles were obtained. We investigated the expression levels of 2665 genes associated with digestive cancers and counted the number of genes in which expression was altered. A hierarchical clustering analysis was also performed. The final prediction of the cancer possibility was determined according to an algorithm. Results The number of genes in which expression was altered was significantly increased in the kidney transplant recipients in comparison with the general population (1091 ± 63 vs 823 ± 94; P = .0024). The number of genes with altered expression decreased after the induction of mechanistic target of rapamycin (mTOR) inhibitor (1484 ± 227 vs 883 ± 154; P = .0439). No cases of possible digestive cancer were detected in this study period. Conclusion The gene expression profiling of peripheral blood samples may be a useful and noninvasive diagnostic tool that allows for the early detection of cancer of the digestive system.

To evaluate the prognostic value of the warm ischemic time and the validity of the Kidney Donor Profile Index/Kidney Donor Risk Index for predicting the survival of donors after cardiac death grafts.We retrospectively assessed 315 kidneys retrieved from donors after cardiac death at Fujita Health University Hospital, Toyoake, Aichi, Japan. The Kidney Donor Profile Index/Kidney Donor Risk Index was calculated and the grafts were enrolled.The median follow-up period was 11.9 years. The Kidney Donor Profile Index had a markedly asymmetric distribution (median 94%), and the Kidney Donor Risk Index had high index rates (0.79-2.94, median 1.70). The overall 5-, 10- and 15-year graft survival rates were 67.5%, 52.1% and 38.9%, respectively. The Kidney Donor Profile Index correlated with graft survival. The 5-, 10- and 15-year graft survival rates for the Kidney Donor Profile Index1.2 were 87.7%, 73.5% and 59.2%; those for the Kidney Donor Risk Index2.0 were 55.0%, 34.7% and 22.1%, respectively. A Cox multivariate analysis identified the Kidney Donor Risk Index (hazard ratio 2.06, 95% confidence interval 1.48-2.86, P 0.0001) and warm ischemic time (hazard ratio 1.21, 95% confidence interval 1.09-1.34, P = 0.0010) as independent risk factors for graft loss. The addition of warm ischemic time30 min had a significant effect, as measured by the C-index (0.708-0.731, P = 0.032), improving the net reclassification improvement score (0.256, P = 0.0039) and integrated discrimination improvement score (0.042, P = 0.0022).The Kidney Donor Profile Index/Kidney Donor Risk Index is a good prognostic tool for determining the outcomes of donors after cardiac death grafts. However, the warm ischemic time should also be included in the allocation system for donors after cardiac death grafts.

Introduction Three-dimensional (3-D) printing systems allow for the creation of surgical models mimicking real tissue. We developed a kidney graft and pelvic cavity replica as a patient-specific 3-D model using a 3-D printing system with simultaneous jetting of different materials and subsequently evaluated the usefulness of surgical simulation and navigation of living kidney transplantation. Methods After generating a stereolithographic file of the organ surface based on multidetector computed tomographic data, we created a 3-D organ model using an inkjet 3-D printer and manufactured a pelvic cavity replica using patient-specific data. Results The patients' individual 3-D printed models were used to plan and guide the surgical procedures for laparoscopic donor nephrectomy and recipient transplantation surgery. The 3-D organ replicas obtained using transparent materials allowed for the creation of models that showed the visceral organs, blood vessels, and other details, thereby overcoming the limitations of conventional image-guided navigation. Our pelvic replicas can be made according to each patient's specific anatomical data, thus representing personalized surgical procedures. This level of detail of the anatomy enables the surgeons and trainees to virtually treat various pelvic conditions before they perform the surgical procedure. The use of these replicas may also reduce the length of the operation and provide better anatomical reference tools for tailor-made simulation and navigation of kidney transplantation surgery, consequently helping to improve training for the operating room staff, students, and trainees. Conclusions We believe that our sophisticated personalized donor graft and pelvic replications obtained using a 3-D printing system are advantageous for kidney transplantation surgery.

BACKGROUND The goal of this study was to define the role of T-cell sub-sets in the pathogenesis of autoimmunity-induced obliterative airway disease by passive transfer of CD8+ or CD4+ T cells. METHODS Antibodies to major histocompatibility complex (MHC) class I were administered intrabronchially into C57BL/6 animals. Lungs were analyzed by histopathology and immunohistochemistry. The CD8+ and CD4+ T-cell sub-sets were purified from the lung-infiltrating cells and intrabronchially transferred. Frequency of cells secreting interleukin-17, interferon-γ, or interleukin-10 to self-antigens was enumerated by enzyme-linked immunospot assay. Myeloperoxidase and antibodies to self-antigens were determined by enzyme-linked immunosorbent assay. Cytokine and growth factor expression was determined by quantitative reverse-transcription polymerase chain reaction. RESULTS Passive transfer of lung-infiltrating CD8 T cells isolated after anti-MHC class I administration, along with sub-optimal dose, induced significantly higher cellular infiltration (89.3% ± 7.9% vs 62.8% ± 10.1%, p p p CONCLUSION Ligation of MHC molecules by its specific antibodies induced early injury with neutrophils, macrophages, and CD8 T cells, which leads to exposure of cryptic self-antigens and their presentation by the infiltrating CD4+ T cells and B cells, leading to the development of immune responses to self-antigens and culminating in obliterative airway disease.

Background The IL-17 axis is implicated in pathogenesis of chronic rejection after human lung transplantation. Using a murine model of obliterative airway disease (OAD), we recently demonstrated that Abs to MHC class I antigens can induce immune responses to self-antigens that contributes to immunopathogenesis of chronic rejection. Using a murine model of OAD, we determined the role of IL-17 family members in induction of autoimmunity leading to OAD after ligation of MHC class I. Methods Anti-MHC class I or control antibodies (Abs) were administered intrabronchially to wild-type (WT) and IL-17a knock out (IL-17A-/-) C57BL/6. Results By day 30, anti-MHC I administered endobronchially in IL-17A-/- mice demonstrated significant reduction in cellular infiltration, a 36.8% reduction in CD4 T cells, 62.7% in CD11b macrophages, 37.5% in degree of fibrosis, 1.94 fold and 2.17 fold decrease in anti-KAT and anti-Col-V, respectively, when compared with wild-type mice. Analysis of lung infiltrating cells in anti-MHC I WT revealed increase in IL-17A (KAT:92+21,Col-V:103+19spm) and IL-17F (KAT:5.03%,Col-V:2.75%) secreting CD4+ T cells. However, administration of anti-MHC I in IL-17A-/- demonstrated increase only in IL-17F for KAT (13.70%) and Col-V (7.08%). Anti-IL-17(A-F) mAb administration after anti-MHC I abrogated OAD in both WT and IL-17A-/-. Conclusion Our findings indicate that IL-17A and IL-17F secreted by CD4+Th17 cells specific to lung self-antigens are critical mediators of autoimmunity leading to the pathogenesis of OAD.

Using a murine model, we demonstrated that endobronchial administration of antibodies (Abs) to major histocompatibility complex (MHC) class I results in cellular infiltration, epithelial metaplasia, fibrosis and obstruction of the small airways (obliterative airway disease [OAD]) mediated predominantly by Th17 responses to self-antigens. This resembles bronchiolitis obliterans syndrome developed following human lung transplantation. Since B cells play a crucial role in induction of autoimmune responses, we defined the role of B cells and its antigen presenting properties in induction of OAD in this study. Anti-MHC class I was administered endobronchially in B(-/-) and wild-type mice. In contrast to wild type, B(-/-) animals did not demonstrate cellular infiltration, epithelial metaplasia and obstruction of airways following anti-MHC. Frequency of K-α1 tubulin and CollagenV-specific IL-17 cells was significantly decreased in B(-/-) mice. As expected, Abs against self-antigens and germinal center formation were not developed in B(-/-) mice. Thus, we conclude that B cells and its antigen presenting capacity play an important role in induction of immune responses to self-antigens and immunopathogenesis of OAD following the administration of anti-MHC. Therefore, strategies to block B-cell and its antigen presenting functions should be considered for preventing the development of chronic rejection.

Alloimmune-induced immune responses to self-antigens are involved in the development of chronic lung allograft rejection. Angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin II receptor blockers (ARBs) have been shown to modulate autoimmune diseases. This study investigated the effect of modulation of the renin angiotensin aldosterone system (RAAS) a murine model of obliterative airways disease (OAD).Major histocompatibility complex (MHC) class I antibodies were administered intrabronchially to C57Bl/6 mice on Days 1, 2, 3, and 6, and weekly thereafter. ACEI/ARB (10 mg/kg/day) were administered in water 5 days before antibody administration. Antibodies were analyzed by enzyme-linked immunosorbent assay, cytokines by Luminex, Th-frequency by enzyme-linked immunosorbent spot, and transcription factors by Western blotting and real-time polymerase chain reaction.Significant decreases (50%-70%) in airway lesions and fibrous deposition were noted in lungs at Day 30 in the animals administered ACEI and ARB vs controls. Antibody concentrations to self-antigens also decreased from 14 ± 21 to 62 ± 18 μg/ml for collagen V and from 263 ± 43 to 84 ± 28 μg/ml for K-α1 tubulin. Th-precursor frequency and cytokine analysis showed increased interleukin (IL)-10 (3-fold increase) and decreased levels of IL-6 (3.4-fold) and IL-17 (4-fold decrease; p0.05) in ACEI and ARB groups. There was also messenger RNA level downregulation of tumor necrosis factor-α (8.6-fold) and p38/mitogen-activated protein (MAP)kinase (3.1-fold) in the treatment groups.Our results demonstrate that modulation of RAAS leads to downregulation of IL-17 through tumor necrosis factor-α-dependant IL-6 through p38/MAPKinase pathway and thus abrogation of anti-MHC-induced OAD.

Corrosion tests of 10 kinds of stainless steels have been conducted for 5 years under three conditions of marine environments, which are atmospheric corrosion, intermittent seawater spray and intermittent seawater immersion. Rating number increased with PRE(Cr+3.3Mo+16N;mass%) value in all test conditions and the stainless steels of which the value were over 37 showed excellent corrosion resistance under marine environments. Maximum pit depth decreased with increasing PRE value in all test conditions. Rating number settled after a couple of months from the start and then the number almost unchanged. On the contrary maximum pit depth grew with increasing test duration. It is suggested that the progress of corrosion mainly depened on the pit growth.

The molecular mechanisms leading to the development of chronic lung allograft dysfunction following de novo development of antibodies to mismatched donor MHC remain undefined. We demonstrated that intrabronchial administration of antibodies to MHC class I resulted in induction of both innate and adaptive cellular immune responses characterized by a predominance of Th17 specific to lung associated self antigens Kα1-tubulin and Collagen-V leading to the development of obliterative airway lesions (OAD), correlate of chronic rejection following human lung transplantation. To determine the role of regulatory T cells (Treg) in the pathogenesis of OAD, we administered anti-MHC class I to mice, in which Treg were depleted by conditional ablation of FoxP3+cells. Under this condition, we observed a three-fold increase in pulmonary cellular infiltration, luminal occlusion and fibrous deposition when compared anti-MHC class I Ab administered mice maintaining FoxP3. OAD lesions were accompanied with enhanced accumulation of neutrophils along with self-antigen specific Th17 and humoral responses. However, IL-17-blockade or adoptive transfer of Treg abrogated OAD. We conclude that Treg exerts a suppressive effect on anti-MHC induced IL-8 mediated neutrophil infiltration and innate immune responses that leads to inhibition of Th17 immune responses to lung associated self antigens which is critical for development of OAD.

Previous studies have shown that intrabronchial administration of antibodies (Abs) to MHC class I resulted in development of obliterative airway disease (OAD), a correlate of chronic human lung allograft rejection. Since development of Abs specific to mismatched donor HLA class II have also been associated with chronic human lung allograft rejection, we analyzed the role of Abs to MHC class II in inducing OAD. Administration of MHC class II Abs (M5/114) to C57BL/6 mice induced the classical features of OAD even though MHC class II expression is absent de novo on murine lung epithelial and endothelial cells. The induction of OAD was accompanied by enhanced cellular and humoral immune responses to self-antigens (Collagen V and K- α1Tubulin). Further, lung-infiltrating macrophages demonstrated a switch in their phenotype predominance from MΦ1 (F4/80(+)CD11c(+)) to MΦ2 (F4/80(+)CD206(+)) following administration of Abs and prior to development of OAD. Passive administration of macrophages harvested from animals with OAD but not from naïve animals induced OAD lesions. We conclude that MHC class II Abs induces a phenotype switch of lung infiltrating macrophages from MΦ1 (F4/80(+)CD11c(+)) to MΦ2 (F4/80(+)CD206(+)) resulting in the breakdown of self-tolerance along with an increase in autoimmune Th17 response leading to OAD.

Aim Our previous results demonstrated that 200 μg complement fixing antibodies (Abs) to MHC class I (cfAbs) or non-complement fixing Abs (ncAbs) result in obliterative airway disease (OAD) in mouse model of intrabronchial anti-MHC administration. In human lung transplantation, recipients may develop both complement fixing and non-complement fixing DSA that may augment chronic rejection. The aim of this study was to test if low levels of cfAbs and ncAbs can synergize leading to OAD. Methods Low dose anti-MHC class I ncAbs mouse IgG1 (100 μg) or cfAbs IgG2a (25 μg) were given alone or in combination intrabronchially in F1 C57Bl/6 × B.10 mice on day 0,1,2,6 and weekly after. Isotype IgG were given in control. Lungs analyzed by histopathology. Serum tested for Abs to Kalpha1Tubulin (K α 1T) and collagenV (ColV) by ELISA. IFNγ, IL17 and IL10 cells specific to Kα1T and ColV enumerated by ELISPOT. Cytokines and growth factors in lung determined by RTPCR. Results Low dose of ncAbs (100 μg) or cfAbs (25 μg) alone did not lead to OAD including isotype control. Combination of ncAbs and cfAbs at same low dose resulted in OAD with cellular infiltration, fibrosis and luminal occlusion of vessels and bronchioles. Mice given both ncAbs and cfAbs developed serum Abs to Kα1T (667 ± 63 μg/mL, p = 0.065) and ColV (323 ± 41, p = 0.0034) compared to ncAbs alone, cfAbs alone or isotype mice. This was associated with increased IFNγ (p = 0.002), IL17 (p = 0.004) specific to Kα1T and ColV and suppression of IL10 compared to other groups. Further, there was upregulation of TGFβ, MCP-1, IL-6, IL-1β and VEGF. Conclusions High dose ncAbs and cfAbs to MHC lead to OAD, but low dose of ncAbs or cfAbs given alone did not induce OAD. However, low levels of ncAbs and cfAbs in combination, can synergize and potentiate each other resulting in activation of inflammatory and fibrotic cascades leading to OAD. Hence, in human lung transplant recipients low titers of de novo DSA which are cfAbs and ncAbs may synergize augmenting chronic lung allograft rejection.

Antibodies to MHC Induce Autoimmunity Leading to Obliterative Airway Disease: Definition of Mechanisms V. Tiriveedhi, M. Takenaka, V. Subramanian, A. Gelman, G.A. Patterson, T. Mohanakumar. Surgery, Washington University School of Medicine, St. Louis, MO; Pathology & Immunology, Washington University School of Medicine, St. Louis, MO; Cardiothoracic Surgery, Washington University School of Medicine, St. Louis, MO.

Recent evidence strongly suggest that alloimmunity induced autoimmune process plays an important role in the development of chronic rejection following human lung transplantation. We have previously shown that intrabronchial administration of MHC Class I antibodies (Abs) to mice causes obliterative airway disease (OAD, which is considered as a model for chronic rejection following human lung transplantation. Our goal is to study the impact of FoxP3+ regulatory T-cells (Tregs) in this established murine model of OAD. FoxP3DTR transgenic Bl/6 mice (kindly provided by Dr. Rudensky), following diphtheria toxin (DT) administration the Treg frequency at 48hrs decreased by 98%. Intrabronchial administration of H2Kb Abs into DT treated mice demonstrated a 3 fold increase in airway lesions and fibrous deposition over non-Treg depleted animals. Antibody concentrations to self antigens increased from 154 ± 32 to 265 ± 48 µg/mL for Collagen-V and 243 ± 43 to 394 ± 48 µg/mL for K- α1Tubulin. ELISpot analysis for Th-precursor frequency showed enhanced levels of IFN-γ (2.3 fold), IL-17 (3.3 fold) and decreased levels of IL-10 (4.1 fold) in Treg depleted cohort. RT-PCR analysis on lung tissue demonstrated upregulation of chemokines and pro-fibrotic growth factors in Treg depleted cohort. We conclude that depletion of T reg increases the severity of OAD as well as development of autoimmune responses to self-antigens leading to pathogenesis of chronic rejection.