Browsing by Author "Chen, Xingjuan"
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Item Capsaicin and TRPV1 Channels in the Cardiovascular System: The Role of Inflammation(MDPI, 2021-12) Munjuluri, Sreepadaarchana; Wilkerson, Dru A.; Sooch, Gagandeep; Chen, Xingjuan; White, Fletcher A.; Obukhov, Alexander G.; Pharmacology and Toxicology, School of MedicineCapsaicin is a potent agonist of the Transient Receptor Potential Vanilloid type 1 (TRPV1) channel and is a common component found in the fruits of the genus Capsicum plants, which have been known to humanity and consumed in food for approximately 7000–9000 years. The fruits of Capsicum plants, such as chili pepper, have been long recognized for their high nutritional value. Additionally, capsaicin itself has been proposed to exhibit vasodilatory, antimicrobial, anti-cancer, and antinociceptive properties. However, a growing body of evidence reveals a vasoconstrictory potential of capsaicin acting via the vascular TRPV1 channel and suggests that unnecessary high consumption of capsaicin may cause severe consequences, including vasospasm and myocardial infarction in people with underlying inflammatory conditions. This review focuses on vascular TRPV1 channels that are endogenously expressed in both vascular smooth muscle and endothelial cells and emphasizes the role of inflammation in sensitizing the TRPV1 channel to capsaicin activation. Tilting the balance between the beneficial vasodilatory action of capsaicin and its unwanted vasoconstrictive effects may precipitate adverse outcomes such as vasospasm and myocardial infarction, especially in the presence of proinflammatory mediators.Item Catechol estrogens stimulate insulin secretion in pancreatic β-cells via activation of the transient receptor potential A1 (TRPA1) channel(American Society for Biochemistry and Molecular Biology, 2019-02-22) Ma, Wenzhen; Chen, Xingjuan; Cerne, Rok; Syed, Samreen K.; Ficorilli, James V.; Cabrera, Over; Obukhov, Alexander G.; Efanov, Alexander M.; Cellular and Integrative Physiology, School of MedicineEstrogen hormones play an important role in controlling glucose homeostasis and pancreatic β-cell function. Despite the significance of estrogen hormones for regulation of glucose metabolism, little is known about the roles of endogenous estrogen metabolites in modulating pancreatic β-cell function. In this study, we evaluated the effects of major natural estrogen metabolites, catechol estrogens, on insulin secretion in pancreatic β-cells. We show that catechol estrogens, hydroxylated at positions C2 and C4 of the steroid A ring, rapidly potentiated glucose-induced insulin secretion via a nongenomic mechanism. 2-Hydroxyestrone, the most abundant endogenous estrogen metabolite, was more efficacious in stimulating insulin secretion than any other tested catechol estrogens. In insulin-secreting cells, catechol estrogens produced rapid activation of calcium influx and elevation in cytosolic free calcium. Catechol estrogens also generated sustained elevations in cytosolic free calcium and evoked inward ion current in HEK293 cells expressing the transient receptor potential A1 (TRPA1) cation channel. Calcium influx and insulin secretion stimulated by estrogen metabolites were dependent on the TRPA1 activity and inhibited with the channel-specific pharmacological antagonists or the siRNA. Our results suggest the role of estrogen metabolism in a direct regulation of TRPA1 activity with potential implications for metabolic diseases.Item Ex Vivo Method for Assessing the Mouse Reproductive Tract Spontaneous Motility and a MATLAB-based Uterus Motion Tracking Algorithm for Data Analysis(Journal of Visualized Experiments, 2019-09-01) Liang, Kaley L.; Bursova, Julia O.; Lam, Frank; Chen, Xingjuan; Obukhov, Alexander G.; Cellular and Integrative Physiology, School of MedicineDysmenorrhea, or painful cramping, is the most common symptom associated with menses in females and its severity can hinder women's everyday lives. Here, we present an easy and inexpensive method that would be instrumental for testing new drugs decreasing uterine contractility. This method utilizes the unique ability of the entire mouse reproductive tract to exhibit spontaneous motility when maintained ex vivo in a Petri dish containing oxygenated Krebs buffer. This spontaneous motility resembles the wave-like myometrial activity of the human uterus, referred to as endometrial waves. To demonstrate the effectiveness of the method, we employed a well-known uterine relaxant drug, epinephrine. We demonstrate that the spontaneous motility of the entire mouse reproductive tract can be quickly and reversibly inhibited by 1 µM epinephrine in this Petri dish model. Documenting the changes of uterine motility can be easily done using an ordinary smart phone or a sophisticated digital camera. We developed a MATLAB-based algorithm allowing motion tracking to quantify spontaneous uterine motility changes by measuring the rate of uterine horn movements. A major advantage of this ex vivo approach is that the reproductive tract remains intact throughout the entire experiment, preserving all intrinsic intrauterine cellular interactions. The major limitation of this approach is that up to 10-20% of uteri may exhibit no spontaneous motility. Thus far, this is the first quantitative ex vivo method for assessing spontaneous uterine motility in a Petri dish model.Item Furanocoumarins are a novel class of modulators for the transient receptor potential vanilloid type 1 (TRPV1) channel(ASBMB, 2014-04-04) Chen, Xingjuan; Sun, Weiyang; Ginaris, Nicholas G.; Riley, Ashley M.; Cummins, Theodore R.; Fehrenbacher, Jill C.; Obukhov, Alexander G.; Department of Cellular & Integrative Physiology, IU School of MedicineFuranocoumarin imperatorin is the major active component of Angelica dahurica root extracts, widely used in traditional medicine to treat headache, toothache, and orbital eye pain. In this study, we investigated the mechanisms that may underlie the pain-relieving effects of the compound. We found that imperatorin significantly inhibited formalin- and capsaicin-induced nocifensive responses but did not alter baseline thermal withdrawal thresholds in the rat. We established that imperatorin is a weak agonist of TRPV1, a channel implicated in detecting several noxious stimuli, exhibiting a 50% effective concentration (EC50) of 12.6 ± 3.2 μM. A specific TRPV1 antagonist, JNJ-17203212 (0.5 μM), potently inhibited imperatorin-induced TRPV1 activation. Site-directed mutagenesis studies revealed that imperatorin most likely acted via a site adjacent to or overlapping with the TRPV1 capsaicin-binding site. TRPV1 recovery from desensitization was delayed in the presence of imperatorin. Conversely, imperatorin sensitized TRPV1 to acid activation but did not affect the current amplitude and/or the activation-inactivation properties of Na(v)1.7, a channel important for transmission of nociceptive information. Thus, our data indicate that furanocoumarins represent a novel group of TRPV1 modulators that may become important lead compounds in the drug discovery process aimed at developing new treatments for pain management.Item HIV-Nef Protein Transfer to Endothelial Cells Requires Rac1 Activation and Leads to Endothelial Dysfunction Implications for Statin Treatment in HIV Patients(American Heart Association, 2019-08-27) Chelvanambi, Sarvesh; Gupta, Samir K.; Chen, Xingjuan; Ellis, Bradley W.; Maier, Bernhard F.; Colbert, Tyler M.; Kuriakose, Jithin; Zorlutuna, Pinar; Jolicoeur, Paul; Obukhov, Alexander G.; Clauss, Matthias; Medicine, School of MedicineRationale Even in antiretroviral therapy (ART) treated patients, HIV continues to play a pathogenic role in cardiovascular diseases. A possible cofactor may be persistence of the early HIV response gene Nef, which we have demonstrated recently to persist in the lungs of HIV+ patients on ART. Previously, we have reported that HIV strains with Nef, but not Nef-deleted HIV strains, cause endothelial proinflammatory activation and apoptosis. Objective To characterize mechanisms through which HIV-Nef leads to the development of cardiovascular diseases using ex vivo tissue culture approaches as well as interventional experiments in transgenic murine models. Methods and Results EV (extracellular vesicles) derived from both peripheral blood mononuclear cells (PBMC) and plasma from HIV+ patient blood samples induced human coronary artery endothelial cells dysfunction. Plasma derived EV from ART+ patients that were HIV-Nef+ induced significantly greater endothelial apoptosis compared to HIV-Nef- plasma EV. Both HIV-Nef expressing T cells and HIV-Nef-induced EV increased transfer of cytosol and Nef protein to endothelial monolayers in a Rac1-dependent manner, consequently leading to endothelial adhesion protein upregulation and apoptosis. HIV-Nef induced Rac1 activation also led to dsDNA breaks in endothelial colony forming cells (ECFC), thereby resulting in ECFC premature senescence and eNOS downregulation. These Rac1 dependent activities were characterized by NOX2-mediated ROS production. Statin treatment equally inhibited Rac1 inhibition in preventing or reversing all HIV-Nef-induction abnormalities assessed. This was likely due to the ability of statins to block Rac1 prenylation as geranylgeranyl transferase inhibitors were effective in inhibiting HIV-Nef-induced ROS formation. Finally, transgenic expression of HIV-Nef in endothelial cells in a murine model impaired endothelium-mediated aortic ring dilation, which was then reversed by 3-week treatment with 5mg/kg atorvastatin. Conclusion These studies establish a mechanism by which HIV-Nef persistence despite ART could contribute to ongoing HIV related vascular dysfunction which may then be ameliorated by statin treatment.Item In vitro and in vivo inhibition of the host TRPC4 channel attenuates Zika virus infection(EMBO Press, 2024) Chen, Xingjuan; Yan, Yunzheng; Liu, Zhiqiang; Yang, Shaokang; Li, Wei; Wang, Zhuang; Wang, Mengyuan; Guo, Juan; Li, Zhenyang; Zhu, Weiyan; Yang, Jingjing; Yin, Jiye; Dai, Qingsong; Li, Yuexiang; Wang, Cui; Zhao, Lei; Yang, Xiaotong; Guo, Xiaojia; Leng, Ling; Xu, Jiaxi; Obukhov, Alexander G.; Cao, Ruiyuan; Zhong, Wu; Anatomy, Cell Biology and Physiology, School of MedicineZika virus (ZIKV) infection may lead to severe neurological consequences, including seizures, and early infancy death. However, the involved mechanisms are still largely unknown. TRPC channels play an important role in regulating nervous system excitability and are implicated in seizure development. We investigated whether TRPCs might be involved in the pathogenesis of ZIKV infection. We found that ZIKV infection increases TRPC4 expression in host cells via the interaction between the ZIKV-NS3 protein and CaMKII, enhancing TRPC4-mediated calcium influx. Pharmacological inhibition of CaMKII decreased both pCREB and TRPC4 protein levels, whereas the suppression of either TRPC4 or CaMKII improved the survival rate of ZIKV-infected cells and reduced viral protein production, likely by impeding the replication phase of the viral life cycle. TRPC4 or CaMKII inhibitors also reduced seizures and increased the survival of ZIKV-infected neonatal mice and blocked the spread of ZIKV in brain organoids derived from human-induced pluripotent stem cells. These findings suggest that targeting CaMKII or TRPC4 may offer a promising approach for developing novel anti-ZIKV therapies, capable of preventing ZIKV-associated seizures and death.Item Long-Term Diabetic Microenvironment Augments the Decay Rate of Capsaicin-Induced Currents in Mouse Dorsal Root Ganglion Neurons(MDPI, 2019-02-21) Chen, Xingjuan; Duan, Yaqian; Riley, Ashley M.; Welch, Megan A.; White, Fletcher A.; Grant, Maria B.; Obukhov, Alexander G.; Cellular and Integrative Physiology, School of MedicineIndividuals with end-stage diabetic peripheral neuropathy present with decreased pain sensation. Transient receptor potential vanilloid type 1 (TRPV1) is implicated in pain signaling and resides on sensory dorsal root ganglion (DRG) neurons. We investigated the expression and functional activity of TRPV1 in DRG neurons of the Ins2+/Akita mouse at 9 months of diabetes using immunohistochemistry, live single cell calcium imaging, and whole-cell patch-clamp electrophysiology. 2',7'-Dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescence assay was used to determine the level of Reactive Oxygen Species (ROS) in DRGs. Although TRPV1 expressing neuron percentage was increased in Ins2+/Akita DRGs at 9 months of diabetes compared to control, capsaicin-induced Ca2+ influx was smaller in isolated Ins2+/Akita DRG neurons, indicating impaired TRPV1 function. Consistently, capsaicin-induced Ca2+ influx was decreased in control DRG neurons cultured in the presence of 25 mM glucose for seven days versus those cultured with 5.5 mM glucose. The high glucose environment increased cytoplasmic ROS accumulation in cultured DRG neurons. Patch-clamp recordings revealed that capsaicin-activated currents decayed faster in isolated Ins2+/Akita DRG neurons as compared to those in control neurons. We propose that in poorly controlled diabetes, the accelerated rate of capsaicin-sensitive TRPV1 current decay in DRG neurons decreases overall TRPV1 activity and contributes to peripheral neuropathy.Item Long-term spironolactone treatment reduces coronary TRPC expression, vasoconstriction, and atherosclerosis in metabolic syndrome pigs(Springer, 2017) Li, Wennan; Chen, Xingjuan; Riley, Ashley M.; Hiett, S. Christopher; Temm, Constance J.; Beli, Eleni; Long, Xin; Chakraborty, Saikat; Alloosh, Mouhamad; White, Fletcher A.; Grant, Maria B.; Sturek, Michael; Obukhov, Alexander G.; Ophthalmology, School of MedicineCoronary transient receptor potential canonical (TRPC) channel expression is elevated in metabolic syndrome (MetS). However, differential contribution of TRPCs to coronary pathology in MetS is not fully elucidated. We investigated the roles of TRPC1 and TRPC6 isoforms in coronary arteries of MetS pigs and determined whether long-term treatment with a mineralocorticoid receptor inhibitor, spironolactone, attenuates coronary TRPC expression and associated dysfunctions. MetS coronary arteries exhibited significant atherosclerosis, endothelial dysfunction, and increased histamine-induced contractions. Immunohistochemical studies revealed that TRPC6 immunostaining was significantly greater in the medial layer of MetS pig coronary arteries compared to that in Lean pigs, whereas little TRPC6 immunostaining was found in atheromas. Conversely, TRPC1 immunostaining was weak in the medial layer but strong in MetS atheromas, where it was predominantly localized to macrophages. Spironolactone treatment significantly decreased coronary TRPC expression and dysfunctions in MetS pigs. In vivo targeted delivery of the dominant-negative (DN)-TRPC6 cDNA to the coronary wall reduced histamine-induced calcium transients in the MetS coronary artery medial layer, implying a role for TRPC6 in mediating calcium influx in MetS coronary smooth muscles. Monocyte adhesion was increased in Lean pig coronary arteries cultured in the presence of aldosterone; and spironolactone antagonized this effect, suggesting that coronary mineralocorticoid receptor activation may regulate macrophage infiltration. TRPC1 expression in atheroma macrophages was associated with advanced atherosclerosis, whereas medial TRPC6 upregulation correlated with increased histamine-induced calcium transients and coronary contractility. We propose that long-term spironolactone treatment may be a therapeutic strategy to decrease TRPC expression and coronary pathology associated with MetS.Item Long-Term Treatment with Gadopentetic Acid or Gadodiamide Increases TRPC5 Expression and Decreases Adriamycin Nuclear Accumulation in Breast Cancer Cells(MDPI, 2023-05-03) Zhang, Weiheng; Wang, Mengyuan; Lv, Weizhen; White, Fletcher A.; Chen, Xingjuan; Obukhov, Alexander G.; Anesthesia, School of MedicineGadopentetic acid and gadodiamide are paramagnetic gadolinium-based contrast agents (GBCAs) that are routinely used for dynamic contrast-enhanced magnetic resonance imaging (MRI) to monitor disease progression in cancer patients. However, growing evidence indicates that repeated administration of GBCAs may lead to gadolinium (III) cation accumulation in the cortical bone tissue, skin, basal ganglia, and cerebellum, potentially leading to a subsequent slow long-term discharge of Gd3+. Gd3+ is a known activator of the TRPC5 channel that is implicated in breast cancer's resistance to chemotherapy. Herein, we found that gadopentetic acid (Gd-DTPA, 1 mM) potentiated the inward and outward currents through TRPC5 channels, which were exogenously expressed in HEK293 cells. Gd-DTPA (1 mM) also activated the Gd3+-sensitive R593A mutant of TRPC5, which exhibits a reduced sensitivity to GPCR-Gq/11-PLC dependent gating. Conversely, Gd-DTPA had no effect on TRPC5-E543Q, a Gd3+ insensitive TRPC5 mutant. Long-term treatment (28 days) of human breast cancer cells (MCF-7 and SK-BR-3) and adriamycin-resistant MCF-7 cells (MCF-7/ADM) with Gd-DTPA (1 mM) or gadodiamide (GDD, 1 mM) did not affect the IC50 values of ADM. However, treatment with Gd-DTPA or GDD significantly increased TRPC5 expression and decreased the accumulation of ADM in the nuclei of MCF-7 and SK-BR-3 cells, promoting the survival of these two breast cancer cells in the presence of ADM. The antagonist of TRPC5, AC1903 (1 μM), increased ADM nuclear accumulation induced by Gd-DTPA-treatment. These data indicate that prolonged GBCA treatment may lead to increased breast cancer cell survival owing to the upregulation of TRPC5 expression and the increased ADM resistance. We propose that while focusing on providing medical care of the best personalized quality in the clinic, excessive administration of GBCAs should be avoided in patients with metastatic breast cancer to reduce the risk of promoting breast cancer cell drug resistance.Item Mechanisms underlying capsaicin effects in canine coronary artery: implications for coronary spasm(Oxford University Press, 2014-09-01) Hiett, S. Christopher; Owen, Meredith K.; Li, Wennan; Chen, Xingjuan; Riley, Ashley; Noblet, Jillian; Flores, Sarah; Sturek, Michael; Tune, Johnathan D.; Obukhov, Alexander G.; Department of Cellular & Integrative Physiology, IU School of MedicineAIMS: The TRPV1, transient receptor potential vanilloid type 1, agonist capsaicin is considered to be beneficial for cardiovascular health because it dilates coronary arteries through an endothelial-dependent mechanism and may slow atheroma progression. However, recent reports indicate that high doses of capsaicin may constrict coronary arterioles and even provoke myocardial infarction. Thus far, the mechanisms by which TRPV1 activation modulates coronary vascular tone remain poorly understood. This investigation examined whether there is a synergistic interplay between locally acting vasoconstrictive pro-inflammatory hormones (autacoids) and capsaicin effects in the coronary circulation. METHODS AND RESULTS: Experiments were performed in canine conduit coronary artery rings and isolated smooth muscle cells (CASMCs). Isometric tension measurements revealed that 1-10 μM capsaicin alone did not affect resting tension of coronary artery rings. In contrast, in endothelium-intact rings pre-contracted with a Gq/11-coupled FP/TP (prostaglandin F/thromboxane) receptor agonist, prostaglandin F2α (PGF2α; 10 μM), capsaicin first induced transient dilation that was followed by sustained contraction. In endothelium-denuded rings pre-contracted with PGF2α or thromboxane analogue U46619 (1 μM, a TP receptor agonist), capsaicin induced only sustained contraction. Blockers of the TP receptor or TRPV1 significantly inhibited capsaicin effects, but these were still observed in the presence of 50 μM nifedipine and 70 mM KCl. Capsaicin also potentiated 20 mM KCl-induced contractions. Fluorescence imaging experiments in CASMCs revealed that the Gq/11-phospholipase C (PLC)-protein kinase C (PKC) and Ca(2+)-PLC-PKC pathways are likely involved in sensitizing CASMC TRPV1 channels. CONCLUSION: Capsaicin alone does not cause contractions in conduit canine coronary artery; however, pre-treatment with pro-inflammatory prostaglandin-thromboxane agonists may unmask capsaicin's vasoconstrictive potential.