Publications by authors named "Jianming Pei"

88 Publications

Activation of κ-opioid receptor inhibits inflammatory response induced by sodium palmitate in human umbilical vein endothelial cells.

Cytokine 2021 Jul 28;146:155659. Epub 2021 Jul 28.

Department of Physiology and Pathophysiology, National Key Discipline of Cell Biology, Air Force Medical University, No. 169 West Changle Road, Xi'an 710032, Shaanxi Province, People's Republic of China; School of Life Sciences, Northwest University, No.1 North Taibai Road, Xi'an 710069, Shaanxi Province, People's Republic of China. Electronic address:

Objectives: The current study aims to investigate the effect of κ-opioid receptor (κ-OR) activation on sodium palmitate (SP)-induced human umbilical vein endothelial cells (HUVECs) inflammatory response and elucidate the underlying mechanisms.

Methods: A hyperlipidemic cell model was established and treated with κ-OR agonist (U50,488H), and antagonist (norbinaltorphimine, nor-BNI), or inhibitors targeting PI3K, Akt or eNOS (LY294002, MK2206-2HCl or L-NAME, respectively). Furthermore, the expression levels of NLRP3, caspase-1, p-Akt, Akt, p-eNOS, and total eNOS were evaluated. Additionally, the production of reactive oxygen species, and levels of inflammatory factors, such as TNF-α, IL-1β, IL-6, IL-1 and adhesion molecules, such as ICAM-1, VCAM-1, P-selectin, and E-selectin were determined. The adherence rates of the neutrophils and monocytes were assessed as well.

Results: The SP-induced hyperlipidemic cell model demonstrated increased expression of NLRP3 and caspase-1 proteins (P < 0.05) and elevated ROS levels (P < 0.01), and decreased phosphorylated-Akt and phosphorylated-eNOS expression (P < 0.05). In addition, SP significantly increased TNF-α, IL-1β, IL-6, ICAM-1, VCAM-1, P-selectin, and E-selectin levels (P < 0.01), decreased IL-10 levels (P < 0.01), and increased the adhesion rates of monocytes and neutrophils (P < 0.01). The SP-induced inflammatory response in HUVECs was ameliorated by κ-OR agonist, U50,488H. However, the protective effect of U50,488H was abolished by κ-OR antagonist, nor-BNI, and inhibitors of PI3K, Akt and eNOS.

Conclusion: Our findings suggest that κ-OR activation inhibits SP-induced inflammation by activating the PI3K/Akt/eNOS signaling pathway.
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http://dx.doi.org/10.1016/j.cyto.2021.155659DOI Listing
July 2021

SMARCA2-NR4A3 is a novel fusion gene of extraskeletal myxoid chondrosarcoma identified by RNA next-generation sequencing.

Genes Chromosomes Cancer 2021 Jun 14. Epub 2021 Jun 14.

Department of Pathology, Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

Extraskeletal myxoid chondrosarcoma (EMC) is a rare sarcoma of uncertain differentiation, characterized by recurrent chromosomal translocation involving NR4A3 (9q22.33) in more than 90% of cases. Five fusion partners for NR4A3 have been described including: EWSR1 (22q12.2), TAF15 (17q12), FUS (16p11.2), TCF12 (15q21), and TFG (3q12.2). This report describes a patient with an EMC at the dorsum of the right foot. The tumor showed a cord-like and reticular pattern in a background of myxoid matrix. The tumor cells demonstrated an epithelioid morphology with prominent nucleoli. The tumor cells were positive for synaptophysin, GFAP, with focal positivity for CD117, S100, Cam5.2, and NSE, and negative for AE1/3, desmin, and SMA. An RNA next-generation sequencing test showed a SMARCA2-NR4A3 gene fusion which has not been previously reported. The exon 3 of SMARCA2 was fused to exon 3 of NR4A3. This fusion was confirmed by NR4A3 break-apart FISH, although both SMARCA2 (9p24.3) and NR4A3 (9q22.33) are located on chromosome 9. The tumor cells showed retained expression of INI1 and SMARCA2 by immunohistochemistry.
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http://dx.doi.org/10.1002/gcc.22976DOI Listing
June 2021

κ-opioid receptor stimulation alleviates rat vascular smooth muscle cell calcification via PFKFB3-lactate signaling.

Aging (Albany NY) 2021 05 20;13(10):14355-14371. Epub 2021 May 20.

Department of Physiology and Pathophysiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an 710032, Shaanxi Province, China.

In the present study, the effects and mechanism of action of U50,488H (a selective κ-opioid receptor agonist) on calcification of rat vascular smooth muscle cells (VSMCs) induced by β-glycerophosphate (β-GP) were investigated. VSMCs were isolated and cultured in traditional FBS-based media. A calcification model was established in VSMCs under hyperphosphatemia and intracellular calcium contents. Alkaline phosphatase (ALP), lactate dehydrogenase (LDH), and lactate were detected in cell culture supernatants before and after treatment. Alizarin red staining was used to detect the degree of calcification of VSMCs. Expression levels of key molecules of osteogenic markers, fructose-2,6-biphosphatase 3 (PFKFB3), and proline hydroxylase 2 (PHD2), were determined using western blotting. Further, vascular calcification was induced by vitamin plus nicotine in rats and isolated thoracic aortas, calcium concentration was assessed in rat aortic rings . We demonstrated that U50,488H inhibited VSMC calcification in a concentration-dependent manner. Moreover, U50,488H significantly inhibited osteogenic differentiation and ALP activity in VSMCs pretreated with β-GP. Further studies confirmed that PFKFB3 expression, LDH level, and lactate content significantly increased during calcification of VSMCs; U50,488H reversed these changes. PHD2 expression showed the opposite trend compared to PFKFB3 expression. nor-BNI or 3-PO abolished U50,488H protective effects. Besides, U50,488H inhibited VSMC calcification in rat aortic rings . Collectively, our experiments show that κ-opioid receptor activation inhibits VSMC calcification by reducing PFKFB3 expression and lactate content, providing a potential drug target and strategy for the clinical treatment of vascular calcification.
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http://dx.doi.org/10.18632/aging.203050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8202865PMC
May 2021

Punicalagin Protects Against Diabetic Cardiomyopathy by Promoting Opa1-Mediated Mitochondrial Fusion Regulating PTP1B-Stat3 Pathway.

Antioxid Redox Signal 2021 Jun 10. Epub 2021 Jun 10.

Department of Geriatrics Cardiology, The Second Affiliated Hospital, School of Medicine, Xi'an Jiaotong University, Xi'an, China.

This study aims to explore the efficacy of punicalagin (PG) on diabetic cardiomyopathy (DCM), with a specific focus on the mechanisms underlying the effects of PG on mitochondrial fusion/fission dynamics. Cardiac structural and functional abnormalities were ameliorated in diabetic rats receiving PG administration as evidenced by increased ejection fraction, and attenuated myocardial fibrosis and hypertrophy. PG enhanced mitochondrial function and inhibited mitochondria-derived oxidative stress by promoting Opa1-mediated mitochondrial fusion. The benefits of PG could be abrogated by knockdown of Opa1 and . Inhibitor screening and chromatin immunoprecipitation analysis showed that Stat3 directly regulated the transcriptional expression of Opa1 by binding to its promoter and was responsible for PG-induced Opa1-mediated mitochondrial fusion. Moreover, pharmmapper screening and molecular docking studies revealed that PG embedded into the activity pocket of PTP1B and inhibited the activity of PTP1B. Overexpression of PTP1B blocked the promoting effect of PG on Stat3 phosphorylation and Opa1-mediated mitochondrial fusion, whereas knockdown of PTP1B mimicked the benefits of PG in high-glucose-treated cardiomyocytes. Our study is the first to identify PG as a novel mitochondrial fusion promoter against hyperglycemia-induced mitochondrial oxidative injury and cardiomyopathy by upregulating Opa1 regulating PTP1B-Stat3 pathway. PG protects against DCM by promoting Opa1-mediated mitochondrial fusion, a process in which PG interacts with PTP1B and inhibits its activity, which in turn increases Stat3 phosphorylation and then enhances the transcriptional expression of Opa1. These results suggest that PG might be a promising new therapeutic approach against diabetic cardiac complication.
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http://dx.doi.org/10.1089/ars.2020.8248DOI Listing
June 2021

Clinical Application of Chromosome Microarray Analysis in the Diagnosis of Lipomatous Tumors.

Appl Immunohistochem Mol Morphol 2021 Mar 17. Epub 2021 Mar 17.

Department of Pathology, Fox Chase Cancer Center, Philadelphia, PA.

Well-differentiated liposarcoma/atypical lipomatous tumor (WDLS/ALT) and dedifferentiated liposarcoma (DDLS) have characteristic supernumerary ring and giant marker chromosomes involving the chromosomal region 12q13-15 which contains MDM2 (12q15), CDK4 (12q14.1), HMGA2 (12q14.3), YEATS4 (12q15), CPM (12q15), and FRS2 (12q15). Detecting MDM2 amplification by fluorescence in situ hybridization (FISH) is considered to be the gold standard for the diagnosis of WDLS/ALT and DDLS. In this study, formalin fixed paraffin embedded clinical specimens (16 liposarcomas and 19 benign lipomatous tumors) were used to detect MDM2 amplification and other chromosomal alterations in WDLS/ALT and DDLS by single nucleotide polymorphism-based chromosome microarray (CMA). All 16 liposarcomas showed MDM2 amplification with a MDM2/cep12 ratio from 2.4 to 8.4 by CMA. Ten (62.5%) of these cases had CDK4/cep12 ratio ≥2.0. All the cases without CDK4 amplification were from the thigh. The MDM2/cep12 ratio of all the benign lipomatous tumors (19/19) was within the normal limits. Twenty-one of the 35 benign lipomatous tumors and liposarcomas were also tested for MDM2 amplification by FISH. All the FISH results were consistent with the CMA results (100%). Along with MDM2 amplification, all 16 liposarcomas (100%) also showed amplification of YEATS4, CPM and FRS2. Only 11 of 16 (69%) cases showed HMGA2 amplification. In conclusion, this study demonstrated that CMA on routine formalin fixed paraffin embedded tissue is a sensitive and specific clinical test for detection of MDM2 gene amplification. Moreover, CMA allows simultaneous detection of genomic changes of interest including CDK4 and others, which provides enriched information for diagnosing lipomatous tumors.
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http://dx.doi.org/10.1097/PAI.0000000000000923DOI Listing
March 2021

BOC-PLAG1, a new fusion gene of pleomorphic adenoma: Identified in a fine-needle aspirate by RNA next-generation sequencing.

Diagn Cytopathol 2021 Jun 12;49(6):790-792. Epub 2021 Mar 12.

Department of Pathology, Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

Pleomorphic adenoma (PA) is the most common benign salivary gland tumor. Fine-needle aspiration (FNA) of PA exhibits variable combinations of bland ductal epithelial cells, myoepithelial cells, and characteristic magenta fibrillary stroma on Diff-Quik/Romanowsky stain. However, a cellular PA with scant chondromyxoid stroma can be a diagnostic challenge on FNA. Around 70% of PAs have a translocation involving PLAG1 or HMGA2. The presence of either PLAG1 or HMGA2 fusion gene can be used to diagnose PA since they have not been reported in other salivary gland tumors except for carcinoma ex PA. In this case report, we describe a case of cellular PA initially diagnosed on FNA as a "low grade salivary gland neoplasm, favor PA." RNA next-generation sequencing performed on the cell block showed a BOC-PLAG1 fusion gene. The presence of PLAG1 fusion gene in conjunction with cytomorphology supported a diagnosis of PA. The mass was surgically removed and proved to be a cellular PA with scattered foci of chondromyxoid and collagenous stroma. To our knowledge, this is the first reported PA bearing BOC-PLAG1. RNA next-generation sequencing performed on cytology specimens can be helpful in achieving a more specific diagnosis of salivary gland tumors.
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http://dx.doi.org/10.1002/dc.24714DOI Listing
June 2021

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition).

Autophagy 2021 Jan 8;17(1):1-382. Epub 2021 Feb 8.

University of Crete, School of Medicine, Laboratory of Clinical Microbiology and Microbial Pathogenesis, Voutes, Heraklion, Crete, Greece; Foundation for Research and Technology, Institute of Molecular Biology and Biotechnology (IMBB), Heraklion, Crete, Greece.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
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http://dx.doi.org/10.1080/15548627.2020.1797280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7996087PMC
January 2021

Genetic ablation of fas-activated serine/threonine kinase ameliorates alcoholic liver disease through modulating HuR-SIRT1 mRNA complex stability.

Free Radic Biol Med 2021 04 18;166:201-211. Epub 2021 Feb 18.

Department of Physiology and Pathophysiology, National Key Discipline of Cell Biology, Basic Medicine School, China. Electronic address:

Chronic alcoholism often causes liver injuries characterized by hepatic steatosis, inflammation as well as oxidative stress and finally leads to advanced cirrhosis and liver cancer. Fas-activated serine/threonine kinase (FASTK) and its homologs are gradually known as multifunctional proteins involved in various biological processes; however, the role of FASTK and its family members in alcoholic liver disease (ALD) is still unexplored. Here we found that, among FASTK family members, the expression of FASTK was specifically induced both in livers of mice received chronic ethanol ingestion and in ethanol-stimulated hepatocytes. Animal studies showed that genetic deletion of FASTK attenuated chronic ethanol ingestion-induced liver damage, steatosis, and inflammation. Moreover, FASTK deficiency was associated with improved oxidative/anti-oxidative system homeostasis and reduced reactive oxygen species (ROS) generation in livers upon chronic ethanol stimulation. Importantly, FASTK ablation preserved hepatic sirtuin-1 (SIRT1) expression/activity upon chronic ethanol ingestion and SIRT1 silencing via adenovirus-mediated small interfering RNA transfer diminished FASTK deletion-elicited beneficial effects on alcohol-associated hepatic steatosis, inflammation, and oxidative stress. Mechanistically, ethanol increased the phosphorylation of human antigen R (HuR, a RNA binding protein that stabilizes SIRT1 mRNA) and triggered the dissociation of HuR-SIRT1 mRNA complex, in turn promoting SIRT1 mRNA decay. Genetic deletion of FASTK diminished ethanol-induced HuR phosphorylation and HuR-SIRT1 mRNA complex dissociation, thereby enhancing SIRT1 mRNA stability. Collectively, these findings for the first time highlight a critical role of FASTK in the pathogenesis of ALD and implicate HuR-SIRT1 mRNA complex involves in this process.
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http://dx.doi.org/10.1016/j.freeradbiomed.2021.02.002DOI Listing
April 2021

Ibrutinib and venetoclax target distinct subpopulations of CLL cells: implication for residual disease eradication.

Blood Cancer J 2021 02 18;11(2):39. Epub 2021 Feb 18.

Fox Chase Cancer Center, Philadelphia, PA, USA.

Ibrutinib inhibits Bruton tyrosine kinase while venetoclax is a specific inhibitor of the anti-apoptotic protein BCL2. Both drugs are highly effective as monotherapy against chronic lymphocytic leukemia (CLL), and clinical trials using the combination therapy have produced remarkable results in terms of rate of complete remission and frequency of undetectable minimal residual disease. However, the laboratory rationale behind the success of the drug combination is still lacking. A better understanding of how these two drugs synergize would eventually help develop other rational combination strategies. Using an ex vivo model that promotes CLL proliferation, we show that modeled ibrutinib proliferative responses, but not viability responses, correlate well with patients' actual clinical responses. Importantly, we demonstrate for the first time that ibrutinib and venetoclax act on distinct CLL subpopulations that have different proliferative capacities. While the dividing subpopulation of CLL responds to ibrutinib, the resting subpopulation preferentially responds to venetoclax. The combination of these targeted therapies effectively reduced both the resting and dividing subpopulations in most cases. Our laboratory findings help explain several clinical observations and contribute to the understanding of tumor dynamics. Additionally, our proliferation model may be used to identify novel drug combinations with the potential of eradicating residual disease.
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http://dx.doi.org/10.1038/s41408-021-00429-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7893066PMC
February 2021

Molecular Profiling of Exceptional Responders to Cancer Therapy.

Oncologist 2021 03 28;26(3):186-195. Epub 2020 Nov 28.

Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

Background: The vast majority of metastatic cancers cannot be cured. Palliative treatment may relieve disease symptoms by stopping or slowing cancer growth and may prolong patients' lives, but almost all patients will inevitably develop disease progression after initial response. However, for reasons that are not fully understood, a very few patients will have extraordinary durable responses to standard anticancer treatments.

Materials And Methods: We analyzed exceptional responders treated at Fox Chase Cancer Center between September 2009 and November 2017. An exceptional response was defined as a complete response lasting more than 1 year or a partial response or stable disease for more than 2 years. Tumor samples were analyzed using an Ambry Genetics test kit with a 142-gene panel. Messenger RNA expression was evaluated using NanoString's nCounter PanCancer Pathways Panel and Immune Profiling Panel and compared with matched controls for gender, age, and cancer type.

Results: Twenty-six exceptional responders with metastatic bladder, kidney, breast, lung, ovarian, uterine, and colon cancers were enrolled. Mutations were identified in 45 genes. The most common mutation was an EPHA5 nonsynonymous mutation detected in 87.5% of patients. Mutations in DNA damage repair pathway genes were also frequent, suggesting increased genome instability. We also found varying expression of 73 genes in the Pathways panel and 85 genes in the Immune Profiling panel, many of them responsible for improvement in tumor recognition and antitumor immune response.

Conclusions: The genomic instability detected in our exceptional responders, plus treatment with DNA damage compounds combined with favorable anticancer immunity, may have contributed to exceptional responses to standard anticancer therapies in the patients studied.

Implications For Practice: With recent advances in the treatment of cancer, there is increased emphasis on the importance of identifying molecular markers to predict treatment outcomes, thereby allowing precision oncology. In this study, it was hypothesized that there is a "specific biologic signature" in the biology of the cancer in long-term survivors that allows sensitivity to systemic therapy and durability of response. Results showed that DNA damage repair pathway alterations, combined with favorable anticancer immunity, may have contributed to exceptional responses. It is very likely that an in-depth examination of outlier responses will become a standard component of drug development in the future.
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http://dx.doi.org/10.1002/onco.13600DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7930427PMC
March 2021

Accelerated FASTK mRNA degradation induced by oxidative stress is responsible for the destroyed myocardial mitochondrial gene expression and respiratory function in alcoholic cardiomyopathy.

Redox Biol 2021 01 1;38:101778. Epub 2020 Nov 1.

Department of Physiology and Pathophysiology, Basic Medicine School, National Key Discipline of Cell Biology, Air Force Medical University, China. Electronic address:

Chronic alcoholism disrupts mitochondrial function and often results in alcoholic cardiomyopathy (ACM). Fas-activated serine/threonine kinase (FASTK) is newly recognized as a key post-transcriptional regulator of mitochondrial gene expression. However, the modulatory role of FASTK in cardiovascular pathophysiology remains totally unknown. In experimental ACM models, cardiac FASTK expression markedly declined. Ethanol directly suppressed FASTK expression at post-transcriptional level through NADPH oxidase-derived reactive oxygen species (ROS). Ethanol destabilized FASTK mRNA 3'-untranslated region (3'-UTR) and accelerated its decay, which was blocked by the clearance of ROS. Regnase-1 (Reg1), a ribonuclease regulating mRNA stability, was induced by ROS in ethanol-stimulated cardiomyocytes. Reg1 directly bound to FASTK mRNA 3'-UTR and promoted its degradation, whereas silencing of Reg1 reversed ethanol-induced FASTK downregulation. Compared to wild type control, alcohol-related myocardial morphological (hypertrophy, fibrosis and cardiomyocyte apoptosis) and functional (reduced ejection fraction and compromised cardiomyocyte contraction) anomalies were worsened in FASTK deficient mice. Mechanistically, FASTK ablation repressed NADH dehydrogenase subunit 6 (MTND6, a mitochondrial gene encoding a subunit of complex I) mRNA production and reduced complex I-supported respiration. Importantly, cardiomyocyte-specific upregulation of FASTK through intra-cardiac AAV9-cTNT injection mitigated myocardial mitochondrial dysfunction and restrained ACM progression. In vitro study showed that overexpression of FASTK ameliorated ethanol-induced MTND6 mRNA downregulation, complex I inactivation, and cardiomyocyte death, whereas these beneficial effects were counteracted by rotenone, a complex I inhibitor. Collectively, ROS-accelerated FASTK mRNA degradation via Reg1 underlies chronic ethanol ingestion-associated mitochondrial dysfunction and cardiomyopathy. Restoration of FASTK expression through genetic approaches might be a promising therapeutic strategy for ACM.
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http://dx.doi.org/10.1016/j.redox.2020.101778DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7677712PMC
January 2021

Immortalization of human primary prostate epithelial cells via CRISPR inactivation of the CDKN2A locus and expression of telomerase.

Prostate Cancer Prostatic Dis 2021 03 1;24(1):233-243. Epub 2020 Sep 1.

Fels Institute for Cancer Research and Molecular Biology, Philadelphia, PA, USA.

Background: Immortalization of primary prostate epithelial cells (PrEC) with just hTERT expression is particularly inefficient in the absence of DNA tumor viral proteins or p16 knockdown.

Materials And Methods: Here, we describe the establishment of immortalized normal prostate epithelial cell line models using CRISPR technology to inactivate the CDKN2A locus concomitantly with ectopic expression of an hTERT transgene.

Results: Using this approach, we have obtained immortal cell clones that exhibit fundamental characteristics of normal cells, including diploid genomes, near normal karyotypes, normal p53 and pRB cell responses, the ability to form non-invasive spheroids, and a non-transformed phenotype. Based on marker expression, these clones are of basal cell origin.

Conclusions: Use of this approach resulted in the immortalization of independent clones of PrEC that retained normal characteristics, were stable, and non-transformed. Thus, this approach could be used for the immortalization of normal primary prostate cells. This technique could also be useful for establishing cell lines from prostate tumor tissues of different tumor grades and/or from patients of diverse ethnicities to generate cell line models that facilitate the study of the molecular basis of disease disparity.
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http://dx.doi.org/10.1038/s41391-020-00274-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7917161PMC
March 2021

Genetic ablation of Fas-activated serine/threonine kinase ameliorates obesity-related hepatic glucose and lipid metabolic disorders via sirtuin-1 signaling.

Biochem Biophys Res Commun 2020 09 30;529(4):1066-1072. Epub 2020 Jul 30.

Department of Geriatrics, Xijing Hospital, Air Force Medical University, Xi'an, Shaanxi, China. Electronic address:

Obesity has become a worldwide pandemic and is associated with various metabolic diseases such as type 2 diabetes mellitus and non-alcoholic fatty liver disease. Fas-activated serine/threonine kinase (Fastk) is a multifunctional protein localized in the mitochondrion; however, the role of Fastk in obesity-related metabolic disorders remains unexplored. Here we found that Fastk expression was specifically induced in livers of high fat (HF) diet-fed mice and in saturated fatty acid (such as palmitate)-loaded hepatocytes. Genetic ablation of Fastk ameliorated HF diet-induced insulin resistance, glucose intolerance, and hepatic steatosis. Further studies confirmed that Fastk knockout suppressed hepatic gluconeogenesis and lipogenesis in HF diet-stressed livers and in palmitate-loaded hepatocytes. Mechanistically, Fastk ablation significantly preserved sirtuin-1 (SIRT1) expression and activity in livers of HF diet-fed mice and in palmitate-loaded hepatocytes. Inhibition of SIRT1 activity by EX-527 (a specific inhibitor of SIRT1) totally abolished the suppressive effects of Fastk knockout on gluconeogenesis and lipogenesis in cultured hepatocytes. In conclusion, these data for the first time demonstrate that Fastk critically controls hepatic gluconeogenesis and lipogenesis mainly through modulating SIRT1 signaling. Intervening Fastk expression or activity might be a promising therapeutic strategy for the treatment of obesity-associated metabolic diseases.
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http://dx.doi.org/10.1016/j.bbrc.2020.06.049DOI Listing
September 2020

Overall tumor genomic instability: an important predictor of recurrence-free survival in patients with localized clear cell renal cell carcinoma.

Cancer Biol Ther 2020 05 1;21(5):424-431. Epub 2020 Mar 1.

Genomics Facility, Fox Chase Cancer Center, Philadelphia, PA, USA.

Measurement of a tumor's overall genomic instability has gathered recent interest over the identification of specific genomic imbalances, as it may provide a more robust measure of tumor aggressiveness. Here we demonstrate the association of tumor genomic instability in the prediction of disease recurrence in patients with clinically localized clear cell renal cell carcinoma (ccRCC). Genomic copy number analysis was performed using SNP-based microarrays on tumors from 103 ccRCC patients. The number of copy number alterations (CNAs) for each tumor was calculated, and a genomic imbalance threshold (GIT) associated with high stage and high-grade disease was determined. Cox proportional hazards regression analyzes were performed to assess the effect of GIT on recurrence-free survival adjusting for known confounders. In the cohort, copy number losses in chromosome arms 3p, 14q, 6q, 9p, and 1p and gains of 5q and 7p/q were common. CNA burden significantly increased with increasing stage ( < .001) and grade ( < .001). The median CNA burden associated with patients presenting with advanced stage (IV) and high-grade (III/IV) tumors was ≥9, defining the GIT. On regression analysis, GIT was a superior predictor of recurrence (Hazard Ratio 4.44 [CI 1.36-14.48], = .01) independent of stage, with similar results adjusting for grade. These findings were confirmed using an alternative measure of genomic instability, weighted Genomic Integrity Index. Our data support a key role for genomic instability in ccRCC progression. More importantly, we have identified a GIT (≥ 9 CNAs) that is a superior and independent predictor of disease recurrence in high-risk ccRCC patients.
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http://dx.doi.org/10.1080/15384047.2020.1721251DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7515487PMC
May 2020

Genetic Variants Detected Using Cell-Free DNA from Blood and Tumor Samples in Patients with Inflammatory Breast Cancer.

Int J Mol Sci 2020 Feb 14;21(4). Epub 2020 Feb 14.

Fox Chase Cancer Center, Philadelphia, PA 19111, USA.

We studied genomic alterations in 19 inflammatory breast cancer (IBC) patients with advanced disease using samples of tissue and paired blood serum or plasma (cell-free DNA, cfDNA) by targeted next generation sequencing (NGS). At diagnosis, the disease was triple negative (TN) in eleven patients (57.8%), ER+ Her2- IBC in six patients (31.6%), ER+ Her2+ IBC in one patient (5.3%), and ER- Her2+ IBC in one other patient (5.3%). Pathogenic or likely pathogenic variants were frequently detected in (47.3%), (26.3%), (26.3%), (10.5%), (10.5%), (10.5%) and (10.5%); other affected genes included , , , , , , , , , , and In 15 of the 19 patients in which tissue and paired blood were collected at the same time point, 80% of the variants detected in tissue were also detected in the paired cfDNA. Higher concordance between tissue and cfDNA was found for variants with higher allele fraction in tissue (AF ≥ 5%). Furthermore, 86% of the variants detected in cfDNA were also detected in paired tissue. Our study suggests that the genetic profile measured in blood cfDNA is complementary to that of tumor tissue in IBC patients.
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http://dx.doi.org/10.3390/ijms21041290DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7072950PMC
February 2020

The effect of activated κ-opioid receptor (κ-OR) on the role of calcium sensing receptor (CaSR) in preventing hypoxic pulmonary hypertension development.

Biomed Pharmacother 2020 May 14;125:109931. Epub 2020 Feb 14.

Department of Physiology and Pathophysiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an, Shaanxi Province, 710032, China. Electronic address:

κ-opioid receptor (κ-OR) plays a key role in preventing hypoxic pulmonary hypertension (HPH) development after activated by exogenous agonist U50,488H. Calcium sensing receptor (CaSR) activation induces HPH by promoting vasoconstriction and vascular remodeling. The activated κ-OR is reported to inhibit the expression of CaSR in pulmonary artery smooth muscle cells (PASMCs). Thus, in this study, we aimed to explore the effect of activated κ-OR on the role of CaSR in preventing HPH development. An HPH rat model was constructed using Sprague-Dawley rats. Changes in mean pulmonary arterial pressure (mPAP) and right ventricular pressure (RVP) mediated by κ-OR agonist U50,488H and CaSR inhibitor NPS2143 were observed. The effects of CaSR agonist spermine and inhibitor NPS2143 on pulmonary artery tension were tested. The expression and localization of κ-OR and CaSR were measured in isolated PASMCs. A cell-counting kit-8 assay was performed to evaluate the effect of spermine in PASMC proliferation. Expression of proliferating cell nuclear antigen (PCNA), Erk, and p-Erk was evaluated by western blot analysis. Results showed that κ-OR and CaSR were co-expressed and colocalized in PASMCs under normoxic and hypoxic conditions. Interactions between κ-OR and CaSR were also observed. Spermine improved vasoconstriction in the pulmonary artery in HPH rats, which was abolished by U50,488H. RVP and mPAP were significantly increased in HPH rats under CaSR stimulation, but were significantly reduced when the rats were pretreated with U50,488H and NPS2143 (P < 0.01). Spermine treatment significantly promoted PASMC proliferation, which was significantly inhibited by U50,488H, p38 inhibitor SB203580, JNK inhibitor SP600125, Erk inhibitor SCH772984, and MEK inhibitor U0126, especially Erk inhibitor (P < 0.01). Spermine significantly increased PCNA and P-Erk expression in hypoxic conditions, which was inhibited by U50,488H and NPS2143. κ-OR stimulation prevented HPH development via the CaSR/MAPK signaling pathway.
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http://dx.doi.org/10.1016/j.biopha.2020.109931DOI Listing
May 2020

P53/PANK1/miR-107 signalling pathway spans the gap between metabolic reprogramming and insulin resistance induced by high-fat diet.

J Cell Mol Med 2020 03 12;24(6):3611-3624. Epub 2020 Feb 12.

Department of Physiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an, China.

High-fat diet (HFD) leads to obesity, type II diabetes mellitus (T2DM) and increases the coincidence of cardiovascular diseases and cancer. Insulin resistance (IR) is considered as the 'common soil' of those diseases. Furthermore, people on HFD showed restrained glycolysis and enhanced fatty acid oxidation, which is the so-called metabolic reprogramming. However, the relationship between metabolic reprogramming and IR induced by HFD is still unclear. Here, we demonstrate that PANK1 and miR-107 were up-regulated in the liver tissue of mice on HFD for 16 weeks and involved in metabolic reprogramming induced by palmitate acid (PA) incubation. Importantly, miR-107 within an intron of PANK1 gene facilitated IR by targeting caveolin-1 in AML12 cells upon PA incubation. Moreover, we identify that HFD enhanced P53 expression, and activation of P53 with nutlin-3a induced PANK1 and miR-107 expression simultaneously in transcriptional level, leading to metabolic reprogramming and IR, respectively. Consistently, inhibition of P53 with pifithrin-α hydrobromide ameliorated PA-induced metabolic reprogramming and IR. Thus, our results revealing a new mechanism by which P53 regulate metabolism. In addition, the results distinguished the different roles of PANK1 and its intron miR-107 in metabolic regulation, which will provide more accurate intervention targets for the treatment of metabolic diseases.
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http://dx.doi.org/10.1111/jcmm.15053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7131928PMC
March 2020

Application of Chromosome Microarray Analysis for the Differential Diagnosis of Low-grade Renal Cell Carcinoma With Clear Cell and Papillary Features.

Appl Immunohistochem Mol Morphol 2020 02;28(2):123-129

Department of Pathology.

Clear cell renal cell carcinoma (ccRCC) and papillary renal cell carcinoma (pRCC) are the 2 most common RCCs. However, some RCCs can have both clear cell and papillary features, including clear cell papillary RCC (ccpRCC). They can be a diagnostic challenge in daily practice. Accurate diagnosis of these tumors is important for both patient prognosis and appropriate treatment. Fourteen RCCs with papillary architecture, clear cytoplasm and low Fuhrman grade were analyzed by SNP-based chromosome microarray (CMA). Seven cases had pathologic features of ccpRCC, and all had normal genomic profiles except one that had copy neutral loss of heterozygosity (cnLOH) of chromosome 3 and loss of one copy of the X chromosome. The remaining 7 cases also had papillae and clear cytoplasm. Two of these cases showed losses of chromosome 3 which are typically found in ccRCC. One had a gain of chromosome 7, which is commonly seen in pRCC. The remaining 4 had no alterations of chromosome 3 or 7. However, 3 of these 4 had monosomy 8, which are consistent with RCC with monosomy 8. The remaining case had no copy number alterations. This study shows that low-grade RCC with papillae and clear cell phenotype represents a heterogeneous group, including ccpRCC, ccRCC, pRCC, and RCC with monosomy 8. CMA analysis can be useful for the differential diagnosis of these neoplasms.
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http://dx.doi.org/10.1097/PAI.0000000000000704DOI Listing
February 2020

κ-opioid receptor activation promotes mitochondrial fusion and enhances myocardial resistance to ischemia and reperfusion injury via STAT3-OPA1 pathway.

Eur J Pharmacol 2020 May 4;874:172987. Epub 2020 Feb 4.

Department of Physiology and Pathophysiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an, Shaanxi, 710032, China. Electronic address:

Mitochondrial dynamics, determining mitochondrial morphology, quality and abundance, have recently been implicated in myocardial ischemia and reperfusion (MI/R) injury. The roles of κ-opioid receptor activation in cardioprotection have been confirmed in our previous studies, while the underlying mechanism associated with mitochondrial dynamics remains unclear. This study aims to investigate the effect of κ-opioid receptor activation on the pathogenesis of MI/R and its underlying mechanisms. MI/R mouse model and hypoxia-reoxygenation cardiomyocyte model were established in this study. Mitochondrial dynamics were analyzed with transmission electron microscopy in vivo and confocal microscopy in vitro. STAT3 phosphorylation and OPA1 expression were detected by Western blotting. We show here that κ-opioid receptor activation with its selective receptor agonist U50,488H promoted mitochondrial fusion and enhanced myocardial resistance to MI/R injury, while these protective effects were blockaded by nor-BNI, a selective κ-opioid receptor antagonist. In addition, κ-opioid receptor activation increased STAT3 phosphorylation and OPA1 expression, which were blockaded by nor-BNI. Furthermore, inhibition of STAT3 phosphorylation by stattic, a specific STAT3 inhibitor, repressed the effects of κ-opioid receptor activation on promoting OPA1 expression and mitochondrial fusion, as well as inhibiting cell apoptosis and oxidative stress both in vivo and in vitro during MI/R injury. Overall, our data for the first time provide evidence that κ-opioid receptor activation promotes mitochondrial fusion and enhances myocardial resistance to MI/R injury via STAT3-OPA1 pathway. Targeting the pathway regulated by κ-opioid receptor activation may be a potential therapeutic strategy for MI/R injury.
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http://dx.doi.org/10.1016/j.ejphar.2020.172987DOI Listing
May 2020

Monosomy of Chromosome 9 Is Associated With Higher Grade, Advanced Stage, and Adverse Outcome in Clear-cell Renal Cell Carcinoma.

Clin Genitourin Cancer 2020 02 26;18(1):56-61. Epub 2019 Sep 26.

Department of Pathology, Fox Chase Cancer Center, Temple Health System, Philadelphia, PA.

Background: Clear-cell renal cell carcinoma (ccRCC) is one of the most common malignancies in humans and is usually associated with poor outcomes. Cancers are considered to be genetic diseases. Therefore, a better understanding of genetic alterations that are related to disease progression or poor prognosis can help to more precisely identify high-risk patients and treat them more effectively. The aim of this study was to examine the frequency of whole chromosome 9 loss (monosomy of chromosome 9) and its prognostic value in patients with ccRCC.

Materials And Methods: Single nucleotide polymorphism-based chromosome microarray (CMA) analysis was performed on 103 resected specimens from patients with ccRCC who had undergone partial or radical nephrectomy between January 2002 and March 2017 at Fox Chase Cancer Center. Monosomy 9 was correlated with clinicopathologic parameters and recurrence-free survival.

Results: Chromosome 9 loss was detected in 31 (30%) of 103 tumors. Tumors with chromosome 9 loss had higher histologic grade (3 and 4; P < .001) and pathologic stage (P < .001). In 59 patients with non-metastatic ccRCC, chromosome 9 loss was also associated with higher recurrence rate and shorter recurrence-free survival (RFS) (12-month RFS, 77.8%; 95% confidence interval, 36.5%-93.9% for chromosome 9 loss vs. 95.7%; 95% confidence interval, 84.0%-98.9% for no loss; P = .002).

Conclusions: Chromosome 9 loss was found in 30% of patients with ccRCC and correlated with higher grade, advanced stage, and shorter RFS in patients with Stage I to III ccRCC.
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http://dx.doi.org/10.1016/j.clgc.2019.09.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7781234PMC
February 2020

Clinical application of RNA sequencing in sarcoma diagnosis: An institutional experience.

Medicine (Baltimore) 2019 Jun;98(25):e16031

Department of Pathology.

Accurate diagnoses of sarcoma are sometimes challenging on conventional histomorphology and immunophenotype. Many specific genetic aberrations including chromosomal translocations have been identified in various sarcomas, which can be detected by fluorescence in situ hybridization and polymerase chain reaction analysis. Next-generation sequencing-based RNA sequencing can screen multiple sarcoma-specific chromosome translocations/fusion genes in 1 test, which is especially useful for sarcoma without obvious differentiation. In this report, we utilized RNA sequencing on formalin-fixed paraffin-embedded (FFPE) specimens to investigate the possibility of diagnosing sarcomas by identifying disease-specific fusion genes. Targeted RNA sequencing was performed on 6 sarcoma cases. The expected genetic alterations (clear cell sarcoma/EWSR1-ATF1, Ewing sarcoma/EWSR1-FLI1, myxoid liposarcoma/DDIT3-FUS) in four cases were detected and confirmed by secondary tests. Interestingly, three SS18 fusion genes (SS18-SSX2B, SS18-SSX2, and SS18-SSX4) were identified in a synovial sarcoma case. A rare fusion gene (EWSR1-PATZ1) was identified in a morphologically challenging case; which enabled us to establish the diagnosis of low grade glioneural tumor. In conclusion, RNA sequencing on FFPE specimen is a reliable method in establishing the diagnosis of sarcoma in daily practice.
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http://dx.doi.org/10.1097/MD.0000000000016031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6636967PMC
June 2019

Circulating tumor cell and cell-free RNA capture and expression analysis identify platelet-associated genes in metastatic lung cancer.

BMC Cancer 2019 Jun 19;19(1):603. Epub 2019 Jun 19.

Protocol Support Laboratory, Fox Chase Cancer Center, Philadelphia, PA, 19111, USA.

Background: Circulating tumor cells (CTC) and plasma cell-free RNA (cfRNA) can serve as biomarkers for prognosis and treatment response in lung cancer. One barrier to the selected or routine use of CTCs and plasma cfRNA in precision oncology is the limited quantity of both, and CTCs are only seen in metastatic disease. As capture of CTCs and plasma cfRNA presents an opportunity to monitor and assess malignancies without invasive procedures, we compared two methods for CTC capture and identification, and profiled mRNA from CTCs and plasma cfRNA to identify potential tumor-associated biomarkers.

Methods: Peripheral blood was collected from ten patients with small cell lung cancer (SCLC), ten patients with non-small cell lung cancer (NSCLC) and four healthy volunteers. Two methods were used for CTC capture: the standard epithelial cell adhesion molecule (EpCam) CellSearch kit (unicapture) and EpCAM plus HER2, EGFR and MUC-1 specific combined ferrofluid capture (quadcapture). For the quadcapture, anti-cytokeratin 7 (CK7) was additionally used to assist in CTC identification. NanoString analysis was performed on plasma cfRNA and on mRNA from combined ferrofluid isolated CTCs. Expression data was analyzed using STRING and Reactome.

Results: Unicapture detected CTCs in 40% of NSCLC and 60% of SCLC; whereas, quadcapture/CK7 identified CTCs in 20% of NSCLC and 80% of SCLC. Bioinformatic analysis of NanoString data identified high expression of a platelet factor 4 (PF4)-related group of transcripts.

Conclusions: Quadcapture ferrofluid reagent did not significantly improve CTC capture efficacy. NanoString analysis based on CTC and plasma cfRNA data highlighted an intriguing PF-4-centric network in patients with metastatic lung cancer.
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http://dx.doi.org/10.1186/s12885-019-5795-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6582501PMC
June 2019

Detecting MYB and MYBL1 fusion genes in tracheobronchial adenoid cystic carcinoma by targeted RNA-sequencing.

Mod Pathol 2019 10 25;32(10):1416-1420. Epub 2019 Apr 25.

Department of Pathology, Fox Chase Cancer Center, Philadelphia, USA.

Primary tracheobronchial adenoid cystic carcinoma is rare, accounting for less than 1% of all lung tumors. Many adenoid cystic carcinomas have been reported to have a specific chromosome translocation t(6;9)/MYB-NFIB. More recently, t(8;9)/MYBL1-NFIB gene fusion was reported in salivary gland adenoid cystic carcinomas which lacked a t(6;9)/MYB-NFIB. Two prior studies showed t(6;9)/MYB-NFIB in tracheobronchial adenoid cystic carcinoma; however, only rare cases of MYBL1 rearrangement have been reported in this carcinoma. In this study, we used targeted RNA sequencing to investigate fusion genes in tracheobronchial adenoid cystic carcinoma at our institution. Fusions of either MYB or MYBL1 genes were detected in 7 of 7 carcinomas. Three cases had MYB-NFIB, and 3 had MYBL1-NFIB. The remaining case showed a rare MYBL1-RAD51B fusion. These findings suggest that rearrangement involving MYB or MYBL1 is a hallmark of tracheobronchial adenoid cystic carcinoma.
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http://dx.doi.org/10.1038/s41379-019-0277-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6763356PMC
October 2019

κ-Opioid receptor stimulation reduces palmitate-induced apoptosis via Akt/eNOS signaling pathway.

Lipids Health Dis 2019 Feb 14;18(1):52. Epub 2019 Feb 14.

Department of Physiology and Pathophysiology, National Key Discipline of Cell Biology, Fourth Military Medical University, No. 169 West Changle Road, Xi'an, 710032, Shaanxi Province, China.

Background: This study was designed to test the hypothesis that κ-opioid receptor (κ-OR) stimulation reduces palmitate-induced HUVECs apoptosis and to investigate its mechanisms.

Methods: HUVECs were subjected to sodium palmitate, apoptosis and cell viability were determined, HUVECs were treated with specific inhibitors to PI3K, Akt, eNOS and siRNAs targeting κ-OR and Akt. Groups were divided as follows: the control group, the sodium palmitate group, the sodium palmitate+U50,488H (a selective κ-OR agonist) group and the sodium palmitate+U50,488H + nor-BNI (a selective κ-OR antagonist) group.

Results: Treatment with sodium palmitate significantly reduced cell viability and increased apoptosis rate which were significantly alleviated by pretreatment with U50,488H, the effect of U50,488H was abolished by nor-BNI. Phosphorylation of Akt and eNOS, as well as NO production were attenuated and accompanied by an increased expression of caspase 3 when HUVECs were subjected to sodium palmitate, and all these changes were restored by pretreatment with U50,488H, the effects of U50,488H were abolished by nor-BNI, and specific inhibitors to PI3K, Akt, eNOS, respectively. SiRNAs targeting κ-OR or Akt abolished the effects of U50,488H on phosphorylation of Akt and eNOS as well as the expressions of caspase 3, Bax and Bcl-2. SiRNAs targeting Akt elicited no effect on the expression of κ-OR.

Conclusion: This study provides the evidence for the first time that κ-OR stimulation possesses anti-palmitate-induced apoptosis effect, which is mediated by PI3K/Akt/eNOS signaling pathway.
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http://dx.doi.org/10.1186/s12944-019-0989-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6376663PMC
February 2019

NEAT1-TFE3 and KAT6A-TFE3 renal cell carcinomas, new members of MiT family translocation renal cell carcinoma.

Mod Pathol 2019 05 8;32(5):710-716. Epub 2019 Jan 8.

Department of Pathology, Fox Chase Cancer Center, Philadelphia, PA, 19111, USA.

Microphthalmia-associated transcription factor (MiT) family translocation renal cell carcinoma harbors variable gene fusions involving either TFE3 or TFEB genes. Multiple 5' fusion partners for TFE3 have been reported, including ASPSCR1, CLTC, DVL2, LUC7L3, KHSRP, PRCC, PARP14, NONO, SFPQ1, MED15, and RBM10. Each of these fusion genes activates TFE3 transcription which can be detected by immunostaining. Using targeted RNA-sequencing, TFE3 fusion gene partners were identified in 5 cases of TFE3 immunohistochemistry positive translocation renal cell carcinoma. Three cases demonstrated known fusions: ASPSCR1-TFE3, MED15-TFE3 and RBM10-TFE3. However, two cases showed unreported NEAT1-TFE3 and KAT6A-TFE3 fusion transcripts. The NEAT1-TFE3 RCC arose in a 59-year-old male; which demonstrated overlapping morphological features seen in NEAT2(MALAT1)-TFEB t(6;11) renal cell carcinoma, including biphasic alveolar/nested tumor cells with eosinophilic cytoplasm. The KAT6A-TFE3 renal cell carcinoma demonstrated typical morphological features of TFE3/Xp11 renal cell carcinoma including papillae, eosinophilic cytoplasm with focal clearing and abundant psammoma bodies. KAT6A gene fusion was reported in some cases of acute myeloid leukemia, which has not been previously reported in solid tumors. This report highlights the genetic complexity of TFE3 translocation renal cell carcinoma; and RNA-sequencing is a powerful approach for elucidating the underlying genetic alterations.
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http://dx.doi.org/10.1038/s41379-018-0191-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6486435PMC
May 2019

Mitochondrial Dysfunction and Apoptosis Are Attenuated on κ-Opioid Receptor Activation Through AMPK/GSK-3β Pathway After Myocardial Ischemia and Reperfusion.

J Cardiovasc Pharmacol 2019 02;73(2):70-81

Department of Physiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an, China.

Previous studies have shown that κ-opioid receptor activation possesses cardioprotection against myocardial ischemia and reperfusion (MI/R) injury. The current study was designed to investigate whether mitochondrial dysfunction after MI/R is regulated by the κ-opioid receptor and to further explore the underlying mechanisms involved. MI/R rat model was established in vivo, and a hypoxia and reoxygenation cardiomyocytes model was used in vitro. Mitochondrial morphology and function as well as myocardial apoptosis were determined. Our data indicated that treatment with U50,488H (a selective κ-opioid receptor agonist) not only reduced apoptosis but also significantly improved mitochondrial morphology and function. These effects were blocked by nor-binaltorphimine (nor-BNI, a selective κ-opioid receptor antagonist), Compound C (an AMPK inhibitor), and AR-A014418 (a GSK3β inhibitor). Moreover, in cardiomyocytes, treatment with U50,488H significantly increased the expression in phosphorylation of AMPK and the phosphorylation of GSK3β. Treatment of cardiomyocytes with AMPKα siRNA decreased the phosphorylation of AMPK and GSK3β. Moreover, AMPK activation resulted in the phosphorylation of GSK3β. Our findings suggested that U50,488H exerted cardioprotective effects by improving mitochondrial morphology and function against MI/R injury through activation of the κ-opioid receptor-mediated AMPK/GSK3β pathway.
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http://dx.doi.org/10.1097/FJC.0000000000000635DOI Listing
February 2019

Sfrp1 attenuates TAC-induced cardiac dysfunction by inhibiting Wnt signaling pathway- mediated myocardial apoptosis in mice.

Lipids Health Dis 2018 Aug 28;17(1):202. Epub 2018 Aug 28.

Department of Physiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an, Shaanxi Province, China.

Background: Hemodynamic overload causes cardiac hypertrophy leading to heart failure. Wnt signaling pathway was reported activated in heart failure. Secreted frizzled related protein 1 (Sfrp1) is a suppressor of Wnt signaling activation. The aim of the present study was to investigate the protective effect of Sfrp1 on hemodynamic overload- induced cardiac dysfunction.

Methods: A mice transverse aortic constriction (TAC)- induced heart failure model was established. A recombinant adeno-associated virus 9 (AAV9) vector was used to deliver Sfrp1 gene into myocardium. Fluorescence and immunohistochemistry staining was used to evaluate the effectiveness of viral vector delivery. Invasive hemodynamic examination was used to evaluate cardiac systolic and diastolic functions. Myocardium apoptosis was detected by TUNEL assay. The expression levels of Sfrp1, β-catenin, caspase3, Bax, Bcl-2 and c-Myc were measured by Western blotting.

Results: Increased mean arterial pressure and impaired cardiac function confirmed the establishment of TAC model. Sfrp1 protein expression was effectively increased in myocardium of mice treated with AAV9-Sfrp1 viral vector. The viral vector administration improved both systolic and diastolic cardiac functions by reducing myocardial apoptosis in TAC mice. The expression levels of β-catenin, caspase3 and Bax were significantly reduced while the expression levels of Bcl-2 and c-Myc were dramatically increased in myocardium by the viral vector treatment in TAC mice.

Conclusions: AAV9 viral vector delivered sfrp1 expression gene into myocardium, which attenuated TAC-induced cardiac dysfunction by inhibiting Wnt signaling pathway activation- mediated apoptosis.
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http://dx.doi.org/10.1186/s12944-018-0832-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6114876PMC
August 2018

κ-opioid receptor activation protects against myocardial ischemia-reperfusion injury via AMPK/Akt/eNOS signaling activation.

Eur J Pharmacol 2018 Aug 30;833:100-108. Epub 2018 May 30.

Department of Physiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an 710032, Shaanxi Province, China. Electronic address:

This study aims to investigate the effect of κ-opioid receptor activation on myocardial ischemia and reperfusion(I/R) injury and elucidate the underlying mechanisms. Myocardial I/R rat model and simulated I/R cardiomyocytes model were established. In vivo study showed that U50,488 H improved cardiac function, reduced myocardial infarct size and serum cTnT significantly. The effect of U50,488 H was abolished by nor-BNI(a κ-opioid receptor antagonist), Compound C(an AMPK inhibitor), Akt inhibitor and L-NAME(an eNOS inhibitor). AICAR, an AMPK activator, mimicked the effect of U50,488 H. U50,488 H up-regulated p-AMPK, p-Akt, and p-eNOS, which were abolished by nor-BNI. AICAR increased p-Akt and p-eNOS, which was abolished by Compound C. In vitro study showed that U50,488 H increased p-AMPK, p-Akt, and p-eNOS via κ-OR activation. The effect of U50,488 H on p-AMPK was abolished by compound C, but not Akt inhibitor and L-NAME. The effect of U50,488 H on p-Akt was abolished by compound C and Akt inhibitor, but not L-NAME. AICAR increased p-Akt and p-eNOS, which was abolished by Akt inhibitor, but not L-NAME. U50,488 H and AICAR also increased the viability of cardiomyocytes subjected to simulated I/R, the effects of U50,488 H and AICAR were blocked by nor-BNI, Compound C, Akt inhibitor, and L-NAME, respectively. In conclusion, κ-OR activation confers cardioprotection via AMPK/Akt/eNOS signaling.
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http://dx.doi.org/10.1016/j.ejphar.2018.05.043DOI Listing
August 2018

Quaternary ammonium salt of U50,488H elicits protective effects against hypoxic pulmonary hypertension.

Eur J Pharmacol 2018 Aug 18;832:129-137. Epub 2018 May 18.

Departemnt of Emergency, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi Province, China; Department of Physiology, National Key Discipline of Cell Biology, Fourth Military Medical University, Xi'an, Shaanxi Province, China. Electronic address:

The present study aimed to investigate the role of quaternary ammonium salt of U50,488H (Q-U50,488H) in hypoxic pulmonary hypertension (HPH) and underlying mechanisms involved. A HPH animal model was established in rats under hypoxia and the mean pulmonary arterial pressure (mPAP) and right ventricular pressure (RVP) were measured. Relaxation of the pulmonary artery in response to Q-U50,488H was determined. In addition, expression and activity of endothelial nitric oxide (NO) synthase (eNOS) and inducible NO synthase (iNOS) with NO content, Akt expression, total antioxidant capacity (T-AOC), and gp91phox were evaluated. Cell viability was determined by the cell counting kit-8 (CCK-8) assay. We demonstrated that both the molecular weight and solubility of Q-U50,488H were higher than that of U50,488H. Q-U50,488H reduced mPAP and RVP and prevented the development of HPH. Moreover, Q-U50,488H relaxed the pulmonary arteries from both normal and HPH rats in a time-dependent manner. Under hypoxic conditions, Q-U50,488H significantly increased Akt phosphorylation, eNOS phosphorylation, NO content in serum, and T-AOC in pulmonary arteries of HPH rats. In addition, the activity of eNOS was elevated, but the activity of iNOS was reduced when Q-U50,488H was given under hypoxia. Q-U50,488H significantly counteracted the increase of gp91phox expression in pulmonary arteries under hypoxia. In addition, in vitro studies suggested that Q-U50,488H inhibited pulmonary artery smooth muscle cells (PASMCs) proliferation under hypoxic conditions and that the effects of Q-U50,488H were blocked by nor-binaltorphimine (nor-BNI). Thus, our results provided evidence that Q-U50,488H plays a protective role against HPH via κ-opioid receptor stimulation.
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http://dx.doi.org/10.1016/j.ejphar.2018.05.025DOI Listing
August 2018

Melatonin prevents Drp1-mediated mitochondrial fission in diabetic hearts through SIRT1-PGC1α pathway.

J Pineal Res 2018 Sep 14;65(2):e12491. Epub 2018 Apr 14.

Department of Physiology, National Key Discipline of Cell Biology, School of Basic Medicine, Fourth Military Medical University, Xi'an, China.

Myocardial contractile dysfunction is associated with an increase in mitochondrial fission in patients with diabetes. However, whether mitochondrial fission directly promotes diabetes-induced cardiac dysfunction is still unknown. Melatonin exerts a substantial influence on the regulation of mitochondrial fission/fusion. This study investigated whether melatonin protects against diabetes-induced cardiac dysfunction via regulation of mitochondrial fission/fusion and explored its underlying mechanisms. Here, we show that melatonin prevented diabetes-induced cardiac dysfunction by inhibiting dynamin-related protein 1 (Drp1)-mediated mitochondrial fission. Melatonin treatment decreased Drp1 expression, inhibited mitochondrial fragmentation, suppressed oxidative stress, reduced cardiomyocyte apoptosis, improved mitochondrial function and cardiac function in streptozotocin (STZ)-induced diabetic mice, but not in SIRT1 diabetic mice. In high glucose-exposed H9c2 cells, melatonin treatment increased the expression of SIRT1 and PGC-1α and inhibited Drp1-mediated mitochondrial fission and mitochondria-derived superoxide production. In contrast, SIRT1 or PGC-1α siRNA knockdown blunted the inhibitory effects of melatonin on Drp1 expression and mitochondrial fission. These data indicated that melatonin exerted its cardioprotective effects by reducing Drp1-mediated mitochondrial fission in a SIRT1/PGC-1α-dependent manner. Moreover, chromatin immunoprecipitation analysis revealed that PGC-1α directly regulated the expression of Drp1 by binding to its promoter. Inhibition of mitochondrial fission with Drp1 inhibitor mdivi-1 suppressed oxidative stress, alleviated mitochondrial dysfunction and cardiac dysfunction in diabetic mice. These findings show that melatonin attenuates the development of diabetes-induced cardiac dysfunction by preventing mitochondrial fission through SIRT1-PGC1α pathway, which negatively regulates the expression of Drp1 directly. Inhibition of mitochondrial fission may be a potential target for delaying cardiac complications in patients with diabetes.
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http://dx.doi.org/10.1111/jpi.12491DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6099285PMC
September 2018
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