Browsing by Author "Cummings, Oscar W."
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Item Automated assessment of steatosis in murine fatty liver(PLOS, 2018-05-10) Sethunath, Deepak; Morusu, Siripriya; Tuceryan, Mihran; Cummings, Oscar W.; Zhang, Hao; Yin, Xiao-Ming; Vanderbeck, Scott; Chalasani, Naga; Gawrieh, Samer; Computer and Information Science, School of ScienceAlthough mice are commonly used to study different aspects of fatty liver disease, currently there are no validated fully automated methods to assess steatosis in mice. Accurate detection of macro- and microsteatosis in murine models of fatty liver disease is important in studying disease pathogenesis and detecting potential hepatotoxic signature during drug development. Further, precise quantification of macrosteatosis is essential for quantifying effects of therapies. Here, we develop and validate the performance of automated classifiers built using image processing and machine learning methods for detection of macro- and microsteatosis in murine fatty liver disease and study the correlation of automated quantification of macrosteatosis with expert pathologist’s semi-quantitative grades. The analysis is performed on digital images of 27 Hematoxylin & Eosin stained murine liver biopsy samples. An expert liver pathologist scored the amount of macrosteatosis and also annotated macro- and microsteatosis lesions on the biopsy images using a web-application. Using these annotations, supervised machine learning and image processing techniques, we created classifiers to detect macro- and microsteatosis. For macrosteatosis prediction, the model’s precision, sensitivity and area under the receiver operator characteristic (AUROC) were 94.2%, 95%, 99.1% respectively. When correlated with pathologist’s semi-quantitative grade of steatosis, the model fits with a coefficient of determination value of 0.905. For microsteatosis prediction, the model has precision, sensitivity and AUROC of 79.2%, 77%, 78.1% respectively. Validation by the expert pathologist of classifier’s predictions made on unseen images of biopsy samples showed 100% and 63% accuracy for macro- and microsteatosis, respectively. This novel work demonstrates that fully automated assessment of steatosis is feasible in murine liver biopsies images. Our classifier has excellent sensitivity and accuracy for detection of macrosteatosis in murine fatty liver disease.Item CD4 T cells but not Th17 cells are Required for Mouse Lung Transplant Obliterative Bronchiolitis(Wiley, 2015-07) Wu, Qiang; Gupta, Pawan Kumar; Suzuki, Hidemi; Wagner, Sarah R.; Zhang, Chen; Cummings, Oscar W.; Fan, Lin; Kaplan, Mark H.; Wilkes, David S.; Shilling, Rebecca A.; Department of Pediatrics, Indiana University School of MedicineLung transplant survival is limited by obliterative bronchiolitis (OB), but the mechanisms of OB development are unknown. Previous studies in a mouse model of orthotopic lung transplantation suggested a requirement for IL-17. We have used this orthotopic mouse model to investigate the source of IL-17A and the requirement for T cells producing IL-17A. The major sources of IL-17A were CD4+ T cells and γδ T cells. Depletion of CD4+ T cells led to a significantly decreased frequency and number of IL-17A+ lymphocytes and was sufficient to prevent acute rejection and OB. However, mice with STAT3-deficient T cells, which are unable to differentiate into Th17 cells, rejected lung allografts and developed OB similar to control mice. The frequency of IL-17A+ cells was not decreased in mice with STAT3-deficient T cells due mainly to the presence of IL-17A+ γδ T cells. Deficiency of γδ T cells also did not affect the development of airway fibrosis. Our data suggest that CD4+ T cells are required for OB development and expansion of IL-17A responses in the lung, while Th17 and γδ T cells are not absolutely required and may compensate for each other.Item Contribution of the anaphylatoxin receptors, C3aR and C5aR, to the pathogenesis of pulmonary fibrosis(Federation of American Societies for Experimental Biology, 2016-06) Gu, Hongmei; Fisher, Amanda J.; Mickler, Elizabeth A.; Duerson, Frank, III; Cummings, Oscar W.; Peters-Golden, Marc; Twigg, Homer L., III; Woodruff, Trent M.; Wilkes, David S.; Vittal, Ragini; Medicine, School of MedicineComplement activation, an integral arm of innate immunity, may be the critical link to the pathogenesis of idiopathic pulmonary fibrosis (IPF). Whereas we have previously reported elevated anaphylatoxins-complement component 3a (C3a) and complement component 5a (C5a)-in IPF, which interact with TGF-β and augment epithelial injury in vitro, their role in IPF pathogenesis remains unclear. The objective of the current study is to determine the mechanistic role of the binding of C3a/C5a to their respective receptors (C3aR and C5aR) in the progression of lung fibrosis. In normal primary human fetal lung fibroblasts, C3a and C5a induces mesenchymal activation, matrix synthesis, and the expression of their respective receptors. We investigated the role of C3aR and C5aR in lung fibrosis by using bleomycin-injured mice with fibrotic lungs, elevated local C3a and C5a, and overexpression of their receptors via pharmacologic and RNA interference interventions. Histopathologic examination revealed an arrest in disease progression and attenuated lung collagen deposition (Masson's trichrome, hydroxyproline, collagen type I α 1 chain, and collagen type I α 2 chain). Pharmacologic or RNA interference-specific interventions suppressed complement activation (C3a and C5a) and soluble terminal complement complex formation (C5b-9) locally and active TGF-β1 systemically. C3aR/C5aR antagonists suppressed local mRNA expressions of tgfb2, tgfbr1/2, ltbp1/2, serpine1, tsp1, bmp1/4, pdgfbb, igf1, but restored the proteoglycan, dcn Clinically, compared with pathologically normal human subjects, patients with IPF presented local induction of C5aR, local and systemic induction of soluble C5b-9, and amplified expression of C3aR/C5aR in lesions. The blockade of C3aR and C5aR arrested the progression of fibrosis by attenuating local complement activation and TGF-β/bone morphologic protein signaling as well as restoring decorin, which suggests a promising therapeutic strategy for patients with IPF.-Gu, H., Fisher, A. J., Mickler, E. A., Duerson, F., III, Cummings, O. W., Peters-Golden, M., Twigg, H. L., III, Woodruff, T. M., Wilkes, D. S., Vittal, R. Contribution of the anaphylatoxin receptors, C3aR and C5aR, to the pathogenesis of pulmonary fibrosis.Item Crosstalk between TGF-β1 and complement activation augments epithelial injury in pulmonary fibrosis(Federation of American Societies for Experimental Biology, 2014-10) Gu, Hongmei; Mickler, Elizabeth A.; Cummings, Oscar W.; Sandusky, George E.; Weber, Daniel J.; Gracon, Adam; Woodruff, Trent; Wilkes, David S.; Vittal, Ragini; Department of Microbiology and Immunology, IU School of MedicineThe epithelial complement inhibitory proteins (CIPs) cluster of differentiation 46 and 55 (CD46 and CD55) regulate circulating immune complex-mediated complement activation in idiopathic pulmonary fibrosis (IPF). Our previous studies demonstrated that IL-17A mediates epithelial injury via transforming growth factor β1 (TGF-β1) and down-regulates CIPs. In the current study, we examined the mechanistic role of TGF-β1 in complement activation-mediated airway epithelial injury in IPF pathogenesis. We observed lower epithelial CIP expression in IPF lungs compared to normal lungs, associated with elevated levels of complement component 3a and 5a (C3a and C5a), locally and systemically. In normal primary human small airway epithelial cells (SAECs) treated with TGF-β1 (10 ng/ml), C3a, or C5a (100 nM), we observed loss of CIPs and increased poly(ADP-ribose) polymerase (PARP) activation [also observed with RNA interference (RNAi) of CD46/CD55]. TGF-β1-mediated loss of CIPs and Snail induction [SNAI1; a transcriptional repressor of E-cadherin (E-CAD)] was blocked by inhibiting mitogen-activated protein kinase (p38MAPK; SB203580) and RNAi silencing of SNAI1. C3a- and C5a-mediated loss of CIPs was also blocked by p38MAPK inhibition. While C3a upregulated TGFb transcripts, both C3a and C5a down-regulated SMAD7 (negative regulator of TGF-β), and whereas TGF-β1 induced C3a/C5a receptor (C3aR/C5aR) expression, pharmacologic C3aR/C5aR inhibition protected against C3a-/C5a-mediated loss of CIPs. Taken together, our results suggest that epithelial injury in IPF can be collectively amplified as a result of TGF-β1-induced loss of CIPs leading to complement activation that down-regulates CIPs and induces TGF-β1 expressionItem Findings in the Distal Colorectum are not associated with Proximal Advanced Serrated Lesions(Elsevier, 2015-02) Kahi, Charles J.; Vemulapalli, Krishna C.; Snover, Dale C.; Jawad, Khaled H. Abdel; Cummings, Oscar W.; Rex, Douglas K.; Department of Medicine, IU School of MedicineBackground & Aims Serrated lesions are an important contributor to colorectal cancer (CRC), notably in the proximal colon. Findings in the distal colorectum are markers of advanced proximal adenomatous neoplasia. However, it is not known whether they affect the odds of advanced proximal serrated lesions. Methods We performed a retrospective cross-sectional study of data from 1910 patients (59.3 ± 8.0 years, 53.8% female) who underwent an average-risk screening colonoscopy from August 2005 through April 2012 at Indiana University Hospital and an associated ambulatory surgery center. Colonoscopies were performed by an endoscopist with high rates of detection of adenomas and serrated polyps. Tissue samples of all serrated polyps (hyperplastic, sessile serrated adenoma/polyp [SSA/P], or traditional serrated adenoma) proximal to the sigmoid colon and serrated polyps >5 mm in the rectum or sigmoid colon were reviewed by a gastrointestinal pathologist and reclassified on the basis of World Health Organization criteria. Advanced serrated lesion (ASL) was defined as SSA/P with cytologic dysplasia, SSA/P ≥10 mm, or traditional serrated adenoma. Advanced conventional adenomatous neoplasia (ACN) was defined as tubular adenoma ≥10 mm, villous histology, high-grade dysplasia, or cancer. The prevalence of proximal ASL and ACN was calculated on the basis of distal colorectal findings. Multivariable logistic regression analysis was performed to determine the age-adjusted and sex-adjusted odds of advanced proximal adenomatous and serrated lesions. Secondary analyses were performed to examine the effect of variable ASL definitions. Results Fifty-two patients (2.7%) had proximal ASL, and 99 (5.2%) had proximal ACN. Of the 52 patients with proximal ASL, 27 (52%) had no distal polyps. Of the 99 patients with proximal ACN, 40 (40%) had no distal polyps. Age and type of distal adenomas were significantly associated with proximal ACN. There were no significant associations between distal polyp type and proximal ASL. In secondary analyses, distal SSA/Ps (P = .008) but not distal hyperplastic polyps or conventional adenomas were associated with any proximal SSA/P. Conclusions The findings at flexible sigmoidoscopy that traditionally serve as indications for colonoscopy (conventional adenomas) are likely to be ineffective for detection of proximal ASL. This finding, plus the observation that most patients with proximal ASL have no distal polyps, favors screening colonoscopy over sigmoidoscopy, especially in the elderly. The observation that non-advanced distal SSA/Ps are associated with any proximal SSA/P warrants further study.Item Gene Expression Profiling of Early- and Late-Relapse Nonseminomatous Germ Cell Tumor and Primitive Neuroectodermal Tumor of the Testis(AACR Publications, 2004-04-01) Sugimura, Jun; Foster, Richard S.; Cummings, Oscar W.; Kort, Eric J.; Takahashi, Masayuki; Lavery, Todd T.; Furge, Kyle A.; Einhorn, Lawrence H.; Teh, Bin Tean; Urology, School of MedicinePurpose: To better understand the molecular mechanisms that underlay the development and progression of nonseminomatous germ cell tumor of testis (NSGCTT) as well as malignant transformation of teratoma and primitive neuroectodermal tumor (PNET). Experimental Design: We studied the gene expression profiles of 17 retroperitoneal NSGCTTs (10 yolk sac tumors, 3 embryonal carcinomas, 4 teratomas) and 2 PNETs obtained from patients with two clinical outcomes. Tissue samples were obtained from the Indiana University. One group of NSGCTT and PNET patients developed metastases within 2 years (early-relapse) of initial successful treatment, and the other group developed metastases after 2 years (late-relapse). Gene expression in these groups of patients was quantified using cDNA microarrays and real-time relative quantitative PCR. Results: We demonstrate that the gene expression profiles of these tumors correlate with histological type. In addition, we identify type-specific genes that may serve as novel diagnostic markers. We also identify a gene set that can distinguish between early-relapse and late-relapse yolk sac tumors. The expression differences of these genes may underlie the differences in clinical outcome and drug response of these tumors. Conclusion: This is the first study that used gene expression profiling to examine the molecular characteristics of the NSGCTTs and drug response in early- and late-relapse tumors. These results suggest that two molecularly distinct forms of NSGCTTs exist and that the integration of expression profile data with clinical parameters could enhance the diagnosis and prognosis of NSGCTTs. More importantly, the identified genes provide insight into the molecular mechanisms of aggressive NSGCTTs and suggest intervention strategies.Item Hepatic Autophagy Deficiency Compromises FXR Functionality and Causes Cholestatic Injury(AASLD, 2019) Khambu, Bilon; Li, Tiangang; Yan, Shengmin; Yu, Changshun; Chen, Xiaoyun; Goheen, Michael; Li, Yong; Lin, Jingmei; Cummings, Oscar W.; Lee, Youngmin A.; Friedman, Scott; Dong, Zheng; Feng, Gen-Sheng; Wu, Shangwei; Yin, Xiao-Ming; Pathology and Laboratory Medicine, School of MedicineAutophagy is important for hepatic homeostasis, nutrient regeneration and organelle quality control. We investigated the mechanisms by which liver injury occurred in the absence of autophagy function. We found that mice deficient in autophagy due to the lack of Atg7 or Atg5, key autophagy‐related genes, manifested intracellular cholestasis with increased levels of serum bile acids, a higher ratio of TMCA/TCA in the bile, increased hepatic bile acid load, abnormal bile canaliculi and altered expression of hepatic transporters. In determining the underlying mechanism, we found that autophagy sustained and promoted the basal and upregulated expression of Fxr in the fed and starved conditions, respectively. Consequently, expression of Fxr and its downstream genes, particularly Bsep, and the binding of FXR to the promoter regions of these genes, were suppressed in autophagy‐deficient livers. In addition, co‐deletion of Nrf2 in autophagy deficiency status reversed the FXR suppression. Furthermore, the cholestatic injury of autophagy‐deficient livers was reversed by enhancement of FXR activity or expression, or by Nrf2 deletion.Item Histologic Abnormalities in Children with Nonalcoholic Fatty Liver Disease and Normal or Mildly Elevated Alanine Aminotransferase Levels.(Elsevier, 2014-04) Molleston, Jean P.; Schwimmer, Jeffrey B.; Yates, Katherine P.; Murray, Karen F.; Cummings, Oscar W.; Lavine, Joel E.; Brunt, Elizabeth M.; Scheimann, Ann O.; Unalp-Arida, Aynur; Department of Pediatrics, IU School of MedicineObjectives: To investigate the histological spectrum of nonalcoholic fatty liver disease (NAFLD) in children with normal, mildly elevated (26–50 U/L boys, 23–44 U/L girls), or elevated (> 50 boys, > 44 girls) serum alanine aminotransferase (ALT) levels. Study design: The Nonalcoholic Steatohepatitis Clinical Research Network (NASH CRN) enrolls children 5–18 years with NAFLD. We analyzed baseline clinical and histological data from 91 children with suspected NAFLD and normal or mildly elevated ALT and liver biopsy within 180 days of ALT, and compared them with 392 children with elevated ALT. Results: Of 91 children, 17 (19%) had normal and 74 (81%) had mildly elevated ALT levels. Overall, 45% of biopsies had ≥ 33% steatosis, lobular inflammation grade was ≥ 2 in 22%, 81% had portal inflammation, 29% had ballooned hepatocytes, 35% had “suspicious/borderline” steatohepatitis, and 8% had definite NASH, 34% had NAFLD activity score (NAS) ≥ 4. Overall, 46% had fibrosis (38% mild/moderate and 8% bridging/cirrhosis). Marked steatosis (50% vs 24%) and fibrosis (54% vs 12%) were significantly more common in mildly elevated vs normal, with no difference in ballooning, inflammation, or NAS ≥ 4. Fibrosis stage 3/4 was seen in none of the children with normal ALT, and in 9% of the mildly elevated and 15% of the elevated. Conclusions: Liver biopsies of children with NAFLD with normal or mildly elevated ALT levels show significant histologic abnormalities, including advanced fibrosis in children with mildly elevated ALT. ALT thus may underestimate liver injury in NAFLD. Appropriate ALT cut-off levels can help identify children at risk for more severe disease.Item Histologic Findings of Advanced Fibrosis and Cirrhosis in Patients With Nonalcoholic Fatty Liver Disease Who Have Normal Aminotransferase Levels(Wolters Kluwer, 2019-10-01) Gawrieh, Samer; Wilson, Laura A.; Cummings, Oscar W.; Clark, Jeanne M.; Loomba, Rohit; Hameed, Bilal; Abdelmalek, Manal F.; Dasarathy, Srinivasan; Neuschwander-Tetri, Brent A.; Kowdley, Kris; Kleiner, David; Doo, Edward; Tonascia, James; Sanyal, Arun; Chalasani, Naga; Network and the NASH Clinical Research; Medicine, School of MedicineBackground and aims: Patients with nonalcoholic fatty liver disease (NAFLD) and normal aminotransferase levels may have advanced liver histology. We conducted a study to characterize the prevalence of and factors associated with advanced liver histology in patients with histologically characterized NAFLD and normal aminotransferase levels. Methods: We evaluated 534 adults with biopsy-proven NAFLD and ALT and AST < 40 U/L within 3 months of their liver biopsy. Histological phenotypes of primary interest were NASH with stage 2-3 fibrosis (NASH F2-3) and cirrhosis. Using multiple logistic regression models with Akaike’s Information Criteria (AIC), we identified variables associated with these histological phenotypes. We developed and internally validated their clinical prediction models. Results: The prevalence of NASH F2-F3 and cirrhosis were 19% and 7%, respectively. The best multiple regression AIC model for NASH F2-3 consisted of type 2 diabetes, White race, lower LDL, lower platelet count, higher AST/ALT ratio, higher serum triglycerides, and hypertension. The best AIC model for cirrhosis consisted of lower platelet count, lower AST/ALT ratio, higher BMI, and female sex. The area under the receiver operator curves of the prediction models were 0.70 (95% CI: 0.65-0.76) for detecting NASH-F2-3 and 0.85 (95% CI: 0.77-0.92) for detecting cirrhosis. When models were fixed at maximum Youden’s index, their positive and negative predictive values were 35% and 88% for NASH F2-F3 and 30% and 98% for cirrhosis, respectively. Conclusion: Clinically significant histological phenotypes are observed in patients with NAFLD and normal aminotransferase levels. Our models can assist the clinicians in excluding advanced liver histology in NAFLD patients with normal aminotransferase levels.Item Lung Transplantation for Bronchopulmonary Dysplasia in Adults: A Clinical and Pathological Study of Three Cases(Wolters Kluwer, 2020-04) Liu, Natalia; Cummings, Oscar W.; Lagstein, Amir; Hage, Chadi A.; Chan, Kevin M.; Zhang, Chen; Pathology and Laboratory Medicine, School of MedicineBronchopulmonary dysplasia (BPD) is usually seen in premature infants who require mechanical ventilation and oxygen therapy for acute respiratory distress. Although most patients wean from oxygen therapy by the ages of 2 to 3, rehospitalization for respiratory problems is common in these patients in adulthood. There have been few studies that document the long-term outcomes of BPD survivors and information about the pulmonary function and radiographic findings of adult BPD are limited. Data on pathologic features of adult BPD are scarce. Three adult patients who underwent recent lung transplantation for BPD from 2 institutions were identified. Clinical data including clinical presentation, chest radiographic images, pulmonary function tests, cardiac catheterization, and echocardiography were retrieved from the electronic medical records. Hematoxylin and eosin and selective elastic stained sections of the explant lungs were examined. CD31 immunohistochemical stain is performed on representative sections. All 3 cases had similar clinical and radiologic features including the history of prematurity and long-term mechanical ventilation after birth, hyperexpanded lungs with air trapping and mosaic attenuation on chest computed tomographic scan, severe obstructive changes on pulmonary function test, and pulmonary hypertension. Pathologic examination showed common features including enlarged and simplified alveoli, peribronchial, subpleural, and interlobular septal fibrosis, narrowing/obliteration of the small airways by elastosis and muscular hypertrophy, thickening of venous walls by fibromuscular hyperplasia, and bronchitis/bronchiolitis. Cholesterol granulomas were seen in 2 cases. The common pathologic findings in the lungs explain the clinical and radiologic findings. Future studies are warranted to further characterize the clinical and pathologic features of adult BPD to develop optimal management strategies for these patients.