Our objective has been to establish a pro-angiogenic role for exosomes in endometriosis and to determine whether a differential expression profile of cellular and exosomal microRNAs (miRNAs) exists in endometriosis. We performed an in vitro study of human primary endometrial stromal cells (ESCs) and human umbilical vein endothelial cells (HUVECs). We isolated and characterized exosomes from ESCs from five endometriosis patients and five phase-matched controls. Exosomes were characterized by transmission electron microscopy and NanoSight technology. MiRNA was assessed by deep sequencing and reverse transcription with quantitative polymerase chain reaction. Exosome uptake studies were achieved by means of confocal microscopy. The pro-angiogenic experiments were executed by treating HUVECs with ESC-derived exosomes. We observed differential profiles of exosomal miRNA expression between exosomes derived from endometriosis lesion cells and diseased eutopic stromal cells compared with exosomes derived from control ESCs. We also demonstrated autocrine cellular uptake of exosomes and paracrine functional angiogenic effects of exosomes on HUVECs. The results of this study support the hypothesis that exosomes derived from ESCs play autocrine/paracrine roles in the development of endometriosis, potentially modulating angiogenesis. The broader clinical implications are that Sampson’s theory of retrograde menstruation possibly encompasses the finding that exosomes work as intercellular communication modulators in endometriosis.
Despite improvements in treatment strategies, colorectal cancer (CRC) still has high mortality rates. Most CRCs develop from adenopolyps via the adenoma-carcinoma sequence. A mechanism for inhibition of this sequence in individuals with a high risk of developing CRC is urgently needed. Differential studies of mitochondrial (mt) gene expressions in the progressive stages of CRC with villous architecture are warranted to reveal early risk assessments and new targets for chemoprevention of the disease. In the present study, reverse transcription-quantitative PCR (RT-qPCR) was used to determine the relative amount of the transcripts of six mt genes [MT-RNR1, MT-ND1, MT-COI, MT-ATP6, MT-ND6, and MT-CYB (region 648–15887)] which are involved in the normal metabolism of mitochondria. A total of 42 pairs of tissue samples obtained from colorectal adenopolyps, adenocarcinomas, and their corresponding adjacent normal tissues were examined. Additionally, electron transport chain (ETC), complexes I (NADH: ubiquinone oxidoreductase) and III (CoQH2-cytochrome C reductase), and carbonyl protein group contents were analyzed. Results indicate that there were differential expressions of the six mt genes and elevated carbonyl protein contents among the colorectal adenopolyps compared to their paired adjacent normal tissues (p < 0.05). The levels of complexes I and III were higher in tumor tissues relative to adjacent normal tissues. Noticeably, the expression of MT-COI was overexpressed in late colorectal carcinomas among all studied transcripts. Our data suggest that increased expressions in certain mt genes and elevated levels of ROS may potentially play a critical role in the colorectal tumors evolving from adenopolyps to malignant lesions.
BackgroundA majority of primary ovarian neoplasms arise from cell surface epithelium of the ovaries. Although old age and a positive family history are associated risk factors, the etiology of the epithelial ovarian tumors is not completely understood. Additionally, knowledge of factors involved in the histogenesis of the various subtypes of this tumor as well as those factors that promote progression to advanced stages of ovarian malignancy are largely unknown. Current evidence suggests that mitochondrial alterations involved in cellular signaling pathways may be associated with tumorigenesis.MethodsIn this study, we determined the presence of polymorphisms and other sequence variants of mitochondrial DNA (mtDNA) in 102 epithelial ovarian tumors including 10 matched normal tissues that paired with some of the tumors. High-resolution restriction endonucleases and PCR-based sequencing were used to assess the mtDNA variants spanning 3.3 kb fragment that comprised the D-Loop and 12S rRNA-tRNAphe, tRNAval, tRNAser, tRNAasp, tRNAlys, ATPase 6, ATPase 8, cytochrome oxidase I and II genes.ResultsThree hundred and fifty-two (352) mtDNA sequence variants were identified, of which 238 of 352 (68%) have not been previously reported. There were relatively high frequencies of three mutations in the 12S rRNA gene at np 772, 773, and 780 in stage IIIC endometrioid tumors, two of which are novel (773delT and 780delC), and occurred with a frequency of 100% (7/7). Furthermore, two mutations were observed in serous tumors only at np 1657 in stage IV (10/10), and at np 8221delA in benign cystadenomas (3/3) and borderline tumors (4/4). A high frequency, 81% (13/16) of TC insertion at np 310 was found only in early stages of serous subtype (benign cystadenomas, 3/3; borderline tumors, 4/4; stage I tumors, 2/5 and matched normal tissues 4/4).ConclusionOur findings indicate that certain mtDNA mutations can reliably distinguish the different histologic subtypes of epithelial ovarian tumors. In addition, these data raise the possibility that certain mtDNA mutations may be useful biomarkers for predicting tumor aggressiveness and may play a potential role in tumorigenesis.
Serous ovarian cancer (SOC) is usually diagnosed at late stage and stage-adjusted five year survival rate is low. Mortality is relatively heavy on African-Americans/Black (AA) affected with SOC compared to their Caucasian counterparts, though the cause for the disparity remains unclear. DNA damage induced by oxidative stress has been linked to ovarian cancer, but the role of oxidative stress in distinguishing differences in aggressive SOC tumors among patients is yet to be determined. This study aims to determine the levels of reactive oxygen species (ROS), malondialdehyde (MDA), reactive carbonyl groups and antioxidants in primary SOC normal, precancerous (cystadenoma, borderline) and invasive (III/IV) tissue samples obtained from AA and Caucasian subgroups. Additionally, the study seeks to investigate significant changes in the level of ROS between AA and Caucasian SOC samples. A fluorogenic probe, dichlorodihydrofluorescein (DCFH-DiOxyQ), was used to scavenge reactive oxygen species in SOC normal, precancerous and malignant stages III/IV tissue samples. Malondialdehyde (MDA), a lipid peroxidation marker, and reactive carbonyl groups were measured as indicators of oxidative injury. Moreover, antioxidant status was assessed by estimating glutathione peroxidase 3 (GPX3) enzyme levels. Results indicate ROS concentration was approximately 96% higher in the malignant tissues in comparative to the normal non-diseased controls. In addition, ROS concentration among AA women was approximately 9% higher than Caucasian women. MDA levels increased exponentially from non-disease control and precancerous tissues relative to malignant tissues. Furthermore, malignant serous ovarian samples showed significantly higher reactive carbonyl content compared to the non-disease controls (p=0.009), while GPX3 levels decreased considerably in serous cystadenoma and malignant tissue samples, and non-diseased control compared to borderline disease. The results suggest accumulation of ROS and MDA levels may be a causative factor for SOC. Elevated levels of MDA and reactive carbonyl proteins could override the GPX3 enzyme capacity therefore, initiating serous ovarian neoplasm.
Serous subtype of ovarian cancer is considered to originate from fallopian epithelium mucosa that has been exposed to physiological changes resulting from ovulation. Ovulation influences an increased in inflammation of epithelial ovarian cells as results of constant exposure of cells to ROS. The imbalance between ROS and antioxidant capacities, as well as a disruption of redox signaling, causes a wide range of damage to DNA, proteins, and lipids. This study applied spectrophotometric, dinitrophenylhydrazone (DNPH) assay, two-dimensional gel electrophoresis, and Western blot analyses to assess the levels of oxidatively modified proteins in 100 primary serous epithelial ovarian carcinoma and normal/surrounding tissues. These samples were obtained from 56 Caucasian and 44 African-American patients within the age range of 61 ± 10 years. Analyses showed that the levels of reactive protein carbonyl groups increased as stages progressed to malignancy. Additionally, the levels of protein carbonyls in serous ovarian carcinoma among African Americans are 40% (P < 0.05) higher relative to Caucasian at similar advanced stages. Results suggest that oxidative stress is involved in the modification of carbonyl protein groups, leading to increased aggressiveness of epithelial ovarian tumors and may contribute to the disease's invasiveness among African Americans.
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