ADRB2, pain and opioids in mice and man

2016 ◽  
Vol 12 (1) ◽  
pp. 122-122
Author(s):  
O. Kambur ◽  
A. Samoshkin ◽  
M. Kaunisto ◽  
J.S. Wieskopf ◽  
J.S. Mogil ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Cellular Localization ◽  
Analgesic Efficacy ◽  
Breast Cancer Surgery ◽  
Cold Pain ◽  
Cellular Effects ◽  
Ici 118,551 ◽  
Opioid Induced Hyperalgesia

Abstract Aims We aim to characterize the effects of variation within ADRB2-gene on pain and opioid requirements in human patients. We will assess ADRB2-OPRM1-6TM heterodimer as a molecular mechanism, potentially explaining pronociceptive and antianalgetic effects, using preclinical in vitro and in vivo models. We will further assess clinical significance via its genetic proxy, rs563649 in humans Methods In humans, experimental and postoperative pain and opioid responses were assessed in 1000 breast cancer surgery patients. Association of ADRB2 (n = 40) and OPRM1 (n = 1) polymorphisms was assessed using linear regression and analysis of variances (ANOVA). In vitro methods involved immunofluorescence microscopy (IF), cellular localization and translocation of 6TM/β2AR-heterodimers and Ca2+-measurements. Behavioral in vivo characterization was performed in mice using formalin, von Frey, hot plate and cold plate tests after administration of morphine, specific OPRM1-6TM agonist IBNtxA and ADRB2-antagonist ICI118,551. Results In humans, several ADRB2 SNPs were associated with pain and opioid phenotypes. The strongest associations were seen between cold pain phenotypes and rs17108817 & rs11957757 (p < 0.0001). In vitro, coexpression with β2-Ars increased translocation of 6TM-MOR to plasma membrane and Ca2+responses after treatment with selective 6TM-agonist, IBNtxA, compared with the cells expressing OPRM1-6TM alone. In vivo, co-administration of P2AR selective antagonist ICI 118,551 increased analgesic efficacy of opioids in a synergistic manner and reduced opioid-induced hyperalgesia. Conclusions Our findings suggest that ADRB2 and genetic variation in ADRB2-gene are involved in the modulation of human pain and opioid responses. 6TM-MOR/P2-AR heterodimerization represents a molecular mechanism causing excitatory cellular effects and sufficient explanatory potential to explain pronociceptive and antianalgesic effects. Our animal findings further confirmed the concept of β2-AR and 6TM-MOR interaction in vivo. We suggest that co-administration of β-blockers with opioids might increase efficacy and safety of OPRM1 agonists.

scholarly journals Isolation and structural elucidation of dimeric epigallocatechin-3-gallate autoxidation products and their antioxidant capacity

2021 ◽  
Author(s):  
Julian Alfke ◽  
Uta Kampermann ◽  
Svetlana Kalinina ◽  
Melanie Esselen
Keyword(s):  
Antioxidant Capacity ◽  
Antioxidant Properties ◽  
Cd Spectroscopy ◽  
Trolox Equivalent Antioxidant Capacity ◽  
Published Data ◽  
Dietary Polyphenols ◽  
Cellular Effects ◽  
The Impact

AbstractDietary polyphenols like epigallocatechin-3-gallate (EGCG)—which represents the most abundant flavan-3-ol in green tea—are subject of several studies regarding their bioactivity and health-related properties. On many occasions, cell culture or in vitro experiments form the basis of published data. Although the stability of these compounds is observed to be low, many reported effects are directly related to the parent compounds whereas the impact of EGCG degradation and autoxidation products is not yet understood and merely studied. EGCG autoxidation products like its dimers theasinensin A and D, “P2” and oolongtheanin are yet to be characterized in the same extent as their parental polyphenol. However, to investigate the bioactivity of autoxidation products—which would minimize the discrepancy between in vitro and in vivo data—isolation and structure elucidation techniques are urgently needed. In this study, a new protocol to acquire the dimers theasinensin A and D as well as oolongtheanin is depicted, including a variety of spectroscopic and quadrupole time-of-flight high-resolution mass spectrometric (qTOF-HRMS) data to characterize and assign these isolates. Through nuclear magnetic resonance (NMR) spectroscopy, polarimetry, and especially circular dichroism (CD) spectroscopy after enzymatic hydrolysis the complementary atropisomeric stereochemistry of the isolated theasinensins is illuminated and elucidated. Lastly, a direct comparison between the isolated EGCG autoxidation products and the monomer itself is carried out regarding their antioxidant properties featuring Trolox equivalent antioxidant capacity (TEAC) values. These findings help to characterize these products regarding their cellular effects and—which is of special interest in the flavonoid group—their redox properties.

scholarly journals PGC7 suppresses TET3 for protecting DNA methylation

2013 ◽  
Vol 42 (5) ◽  
pp. 2893-2905 ◽  
Author(s):  
Chunjing Bian ◽  
Xiaochun Yu
Keyword(s):  
Dna Methylation ◽  
Molecular Mechanism ◽  
Biological Process ◽  
Cpg Islands ◽  
Binding Motifs ◽  
Genome Wide Analysis ◽  
Dna Motif ◽  
Genome Wide

Abstract Ten-eleven translocation (TET) family enzymes convert 5-methylcytosine to 5-hydroxylmethylcytosine. However, the molecular mechanism that regulates this biological process is not clear. Here, we show the evidence that PGC7 (also known as Dppa3 or Stella) interacts with TET2 and TET3 both in vitro and in vivo to suppress the enzymatic activity of TET2 and TET3. Moreover, lacking PGC7 induces the loss of DNA methylation at imprinting loci. Genome-wide analysis of PGC7 reveals a consensus DNA motif that is recognized by PGC7. The CpG islands surrounding the PGC7-binding motifs are hypermethylated. Taken together, our study demonstrates a molecular mechanism by which PGC7 protects DNA methylation from TET family enzyme-dependent oxidation.

Enhanced xenobiotic transporter expression in normal teleost hepatocytes: response to environmental and chemotherapeutic toxins

2001 ◽  
Vol 204 (2) ◽  
pp. 217-227
Author(s):  
J.A. Albertus ◽  
R.O. Laine
Keyword(s):  
Mammalian Cells ◽  
Cellular Localization ◽  
Fundulus Heteroclitus ◽  
Environmental Pollutants ◽  
Human Tumor ◽  
Aquatic Organisms ◽  
In Vivo Studies ◽  
Multixenobiotic Resistance

Many aquatic organisms are resistant to environmental pollutants, probably because their inherent multi-drug-resistant protein extrusion pump (pgp) can be co-opted to handle man-made pollutants. This mechanism of multixenobiotic resistance is similar to the mechanism of multidrug resistance exhibited in chemotherapy-resistant human tumor cells. In the present study, a variety of techniques were used to characterize this toxin defense system in killifish (Fundulus heteroclitus) hepatocytes. The cellular localization and activity of the putative drug efflux system were evaluated. In addition, in vitro and in vivo studies were used to examine the range of expression of this putative drug transporter in the presence of environmental and chemotherapeutic toxins. The broad range of pgp expression generally observed in transformed mammalian cells was found in normal cells of our teleost model. Our findings suggest that the expression of the pgp gene in the killifish could be an excellent indicator of toxin levels or stressors in the environment.

scholarly journals Insights into the Chemical Discovery of Remifentanil

2020 ◽  
Vol 132 (5) ◽  
pp. 1229-1234 ◽  
Author(s):  
Paul L. Feldman
Keyword(s):  
Analgesic Efficacy ◽  
American Chemical Society ◽  
Hepatic Function ◽  
Chemical Society ◽  
Opioid Agonist ◽  
Duration Of Action ◽  
Design Synthesis ◽  
Long Acting

Abstract Design, Synthesis, and Pharmacological Evaluation of Ultrashort- to Long-acting Opioid Analgetics. By Feldman PL, James MK, Brackeen MF, Bilotta JM, Schuster SV, Lahey AP, Lutz MW, Johnson MR, Leighton HJ. J Med Chem 1991; 34:2202-8. Copyright 1991 American Chemical Society. Reprinted with permission. In an effort to discover a potent ultrashort-acting µ-opioid analgetic that is capable of metabolizing to an inactive species independent of hepatic function, several classes of 4-anilidopiperidine analgetics were synthesized and evaluated. One series of compounds displayed potent µ-opioid agonist activity with a high degree of analgesic efficacy and an ultrashort to long duration of action. These analgetics, 4-(methoxycarbonyl)-4-[1-oxopropyl)phenylamino]-1-piperidinepropanoic acid alkyl esters, were evaluated in vitro in the guinea pig ileum for µ-opioid activity, in vivo in the rat tail withdrawal assay for analgesic efficacy and duration of action, and in vitro in human whole blood for their ability to be metabolized in blood. Compounds in this series were all shown to be potent µ agonists in vitro, but depending upon the alkyl ester substitution, the potency and duration of action in vivo varied substantially. The discrepancies between the in vitro and in vivo activities and variations in duration of action are probably due to different rates of ester hydrolysis by blood esterase(s). The [structure–activity relationships] with respect to analgesic activity and duration of action as a function of the various esters synthesized is discussed. It was also demonstrated that the duration of action for the ultrashort-acting analgetic, 8, does not change upon prolonged infusion or administration of multiple bolus injections.

Abstract 356: Krueppel-Like Factor 15 Regulates Wnt/β-Catenin Transcription and Controls Cardiac Progenitor Cell Fate in the Postnatal Heart

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Claudia Noack ◽  
Maria P Zafiriou ◽  
Anke Renger ◽  
Hans J Schaeffer ◽  
Martin W Bergmann ◽  
...  
Keyword(s):  
Cell Fate ◽  
Transcriptional Activation ◽  
Progenitor Cell ◽  
Cellular Localization ◽  
Target Genes ◽  
Critical Role ◽  
Isolated Cells ◽  
Cardiac Progenitor Cell

Wnt/β-catenin signaling controls adult heart remodeling partly by regulating cardiac progenitor cell (CPC) differentiation. We now identified and characterized a novel cardiac interaction of the transcription factor Krueppel-like factor 15 (KLF15) with the Wnt/β-catenin signaling on adult CPCs. In vitro mutation, reporter gene assays and co-localization studies revealed that KLF15 requires two distinct domains for nuclear localization and for repression of β-catenin-mediated transcription. KLF15 had no effect on β-catenin stability or cellular localization, but interacted with its co-factor TCF4, which is required for activation of β-catenin target gene expression. Moreover, increased TCF4 ubiquitination was induced by KLF15. In line with this finding we found KLF15 to interact with the Nemo-like kinase, which was shown to phosphorylate and target TCF4 for degradation. In vivo analyses of adult Klf15 functional knock-out (KO) vs. wild-type (WT) mice showed a cardiac β-catenin-mediated transcriptional activation and reduced TCF4 degradation along with cardiac dysfunction assessed by echocardiography (n=10). FACS analysis of the CPC enriched-population of KO vs. WT mice revealed a significant reduction of cardiogenic-committed precursors identified as Sca1+/αMHC+ (0.8±0.2% vs. 1.8±0.1%) and Tbx5+ (3.5±0.3% vs. 5.2±0.5%). In contrast, endothelial Sca1+/CD31+ cells were significantly higher in KO mice (11.3±0.4% vs. 8.6±0.4%; n≥9). In addition, Sca1+ isolated cells of Klf15 KO showed increased RNA expression of endothelial markers von Willebrand Factor, CD105, and Flk1 along with upregulation of β-catenin target genes. CPCs co-cultured on adult fibroblasts resulted in increased endothelial Flk1 cells and reduction of αMHC and Hand1 cardiogenic cells in KO vs. WT CPCs (n=9). Treating these co-cultures with Quercetin, an inhibitor of nuclear β-catenin, resulted in partial rescue of the observed phenotype. This study uncovers a critical role of KLF15 for the maintenance of cardiac tissue homeostasis. Via inhibition of β-catenin transcription, KLF15 controls cardiomyogenic cell fate similar to embryonic cardiogenesis. This knowledge may provide a tool for activation of endogenous CPCs in the postnatal heart.

scholarly journals MiR-302b as a Combinatorial Therapeutic Approach to Improve Cisplatin Chemotherapy Efficacy in Human Triple-Negative Breast Cancer

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2261 ◽  
Author(s):  
Alessandra Cataldo ◽  
Sandra Romero-Cordoba ◽  
Ilaria Plantamura ◽  
Giulia Cosentino ◽  
Alfredo Hidalgo-Miranda ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Triple Negative ◽  
Standard Of Care ◽  
Integrin Subunit ◽  
Breast Cancers ◽  
Therapeutic Tool ◽  
Response To Chemotherapy ◽  
Cisplatin Sensitivity

Introduction: Chemotherapy is still the standard of care for triple-negative breast cancers (TNBCs). Here, we investigated miR-302b as a therapeutic tool to enhance cisplatin sensitivity in vivo and unraveled the molecular mechanism. Materials and Methods: TNBC-xenografted mice were treated with miR-302b or control, alone or with cisplatin. Genome-wide transcriptome analysis and independent-validation of Integrin Subunit Alpha 6 (ITGA6) expression was assessed on mice tumor samples. Silencing of ITGA6 was performed to evaluate cisplatin response in vitro. Further, potential transcription factors of ITGA6 (E2F transcription facor 1 (E2F1), E2F transcription factor 2 (E2F2), and Yin Yang 1 (YY1)) were explored to define the miRNA molecular mechanism. The miR-302b expression was also assessed in TNBC patients treated with chemotherapy. Results: The miR–302b-cisplatin combination significantly impaired tumor growth versus the control through indirect ITGA6 downregulation. Indeed, ITGA6 was downmodulated in mice treated with miR-302b–cisplatin, and ITGA6 silencing increased drug sensitivity in TNBC cells. In silico analyses and preclinical assays pointed out the regulatory role of the E2F family and YY1 on ITGA6 expression under miR-302b–cisplatin treatment. Finally, miR-302b enrichment correlated with better overall survival in 118 TNBC patients. Conclusion: MiR-302b can be exploited as a new therapeutic tool to improve the response to chemotherapy, modulating the E2F family, YY1, and ITGA6 expression. Moreover, miR-302b could be defined as a new prognostic factor in TNBC patients.

scholarly journals Gallbladder cancer-associated fibroblasts promote vasculogenic mimicry formation and tumor growth in gallbladder cancer via upregulating the expression of NOX4, a poor prognosis factor, through IL-6-JAK-STAT3 signal pathway

2020 ◽  
Vol 39 (1) ◽  
Author(s):  
Mu-Su Pan ◽  
Hui Wang ◽  
Kamar Hasan Ansari ◽  
Xin-Ping Li ◽  
Wei Sun ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Gallbladder Cancer ◽  
Molecular Mechanism ◽  
Poor Prognosis ◽  
Vasculogenic Mimicry ◽  
Tube Formation ◽  
Signal Pathway ◽  
Cancer Associated Fibroblasts

Abstract Background Cancer-associated fibroblasts (CAFs) and vasculogenic mimicry (VM) play important roles in the occurrence and development of tumors. However, the relationship between CAFs and VM formation, especially in gallbladder cancer (GBC) has not been clarified. In this study, we investigated whether gallbladder CAFs (GCAFs) can promote VM formation and tumor growth and explored the underlying molecular mechanism. Methods A co-culture system of human GBC cells and fibroblasts or HUVECs was established. VM formation, proliferation, invasion, migration, tube formation assays, CD31-PAS double staining, optic/electron microscopy and tumor xenograft assay were used to detect VM formation and malignant phenotypes of 3-D co-culture matrices in vitro, as well as the VM formation and tumor growth of xenografts in vivo, respectively. Microarray analysis was used to analyze gene expression profile in GCAFs/NFs and VM (+)/VM (−) in vitro. QRT-PCR, western blotting, IHC and CIF were used to detected NOX4 expression in GCAFs/NFs, 3-D culture/co-culture matrices in vitro, the xenografts in vivo and human gallbladder tissue/stroma samples. The correlation between NOX4 expression and clinicopathological and prognostic factors of GBC patients was analyzed. And, the underlying molecular mechanism of GCAFs promoting VM formation and tumor growth in GBC was explored. Results GCAFs promote VM formation and tumor growth in GBC; and the finding was confirmed by facts that GCAFs induced proliferation, invasion, migration and tube formation of GBC cells in vitro, and promoted VM formation and tumor growth of xenografts in vivo. NOX4 is highly expressed in GBC and its stroma, which is the key gene for VM formation, and is correlated with tumor aggression and survival of GBC patients. The GBC patients with high NOX4 expression in tumor cells and stroma have a poor prognosis. The underlying molecular mechanism may be related to the upregulation of NOX4 expression through paracrine IL-6 mediated IL-6/JAK/STAT3 signaling pathway. Conclusions GCAFs promote VM formation and tumor growth in GBC via upregulating NOX4 expression through the activation of IL-6-JAK-STAT3 signal pathway. NOX4, as a VM-related gene in GBC, is overexpressed in GBC cells and GCAFs, which is related to aggression and unfavorable prognosis of GBC patients.

scholarly journals PTRF/Cavin1 remodels phospholipid metabolism to promote tumor proliferation and suppress immune responses in glioblastoma by stabilizing cPLA2

2020 ◽  
Author(s):  
Kaikai Yi ◽  
Qi Zhan ◽  
Qixue Wang ◽  
Yanli Tan ◽  
Chuan Fang ◽  
...  
Keyword(s):  
Lipid Metabolism ◽  
Immune Responses ◽  
Molecular Mechanism ◽  
Mouse Models ◽  
Phospholipid Composition ◽  
Tumor Infiltrating Lymphocytes ◽  
Intracranial Tumor ◽  
Tumor Proliferation

Abstract Background Metabolism remodeling is a hallmark of glioblastoma (GBM) that regulates tumor proliferation and the immune microenvironment. Previous studies have reported that increased polymerase 1 and transcript release factor (PTRF) levels are associated with a worse prognosis in glioma patients. However, the biological role and the molecular mechanism of PTRF in GBM metabolism remain unclear. Methods The relationship between PTRF and lipid metabolism in GBM was detected by non-targeted metabolomics profiling and subsequent lipidomics analysis. Western blotting, qRT-PCR, and immunoprecipitation were conducted to explore the molecular mechanism of PTRF in lipid metabolism. A sequence of in vitro and in vivo experiments (both xenograft tumor and intracranial tumor mouse models) were used to detect the tumor-specific impacts of PTRF. Results Here, we show that PTRF triggers a cytoplasmic phospholipase A2 (cPLA2)-mediated phospholipid remodeling pathway that promotes GBM tumor proliferation and suppresses tumor immune responses. Research in primary cell lines from GBM patients revealed that cells overexpressing PTRF show increased cPLA2 activity —resulting from increased protein stability —and exhibit remodeled phospholipid composition. Subsequent experiments revealed that PTRF overexpression alters the endocytosis capacity and energy metabolism of GBM cells. Finally, in GBM xenograft and intracranial tumor mouse models, we showed that inhibiting cPLA2 activity blocks tumor proliferation and prevents PTRF-induced reduction in CD8 + tumor-infiltrating lymphocytes. Conclusions The PTRF-cPLA2 lipid remodeling pathway promotes tumor proliferation and suppresses immune responses in GBM. In addition, our findings highlight multiple new therapeutic targets for GBM.

scholarly journals Autoradiographic visualization of beta-adrenergic receptors in normal and denervated skeletal muscle.

1979 ◽  
Vol 27 (10) ◽  
pp. 1308-1311 ◽  
Author(s):  
B Lavenstein ◽  
W K Engel ◽  
N B Reddy ◽  
S Carroll
Keyword(s):  
Skeletal Muscle ◽  
Blood Vessels ◽  
Adrenergic Receptors ◽  
Cellular Localization ◽  
Muscle Fibers ◽  
Beta Adrenergic Receptors ◽  
Beta Adrenergic ◽  
Adrenergic Blocker

Autoradiographic localization of beta-adrenergic receptors in rat skeletal muscle in vivo was achieved utilizing [125I]-iodohydroxybenzylpindolol, a potent beta-adrenergic blocker with high affinity and specificity for those receptors. In normal muscle the beta-adrenergic receptors were localized mainly to blood vessels, arterioles greater than venules, with much less concentration of grains over the fascicles of muscle fibers. One week after denervation there was an increase in binding both to blood vessels and muscle fibers, more so in soleus and gactrocnemius than in extensor digitorum longus. While these results parallel in vitro biochemical studies, they dictate caution when inferring cellular localization of beta-adrenergic receptors (and other molecules) solely on the basis of biochemical techniques applied to subcellular fractions of whole-organ homogenates.

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