Yu-Mi Yang

Yonsei University, Sŏul, Seoul, South Korea

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Publications (13)38.1 Total impact

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    ABSTRACT: Objective: In reaching the airways inhaled allergens pass through and contact with the oral mucosa. Although they are often responsible for initiating asthmatic attacks, it is unknown whether airborne allergens can also trigger chronic inflammation of gingival epithelial cells leading to chronic periodontitis. In this study, we investigated the inflammatory responses of human gingival epithelial cells (HGECs) to airborne allergens, particularly German cockroach extract (GCE) with a focus on calcium signaling. Design: HGECs isolated from healthy donors were stimulated with GCE. Intracellular Ca(2+) concentration ([Ca(2+)]i) was measured with Fura-2-acetoxymethyl ester (Fura-2/AM) staining. Expression of inflammatory cytokines interleukin (IL)-8, IL-1β, IL-6, and NOD-like receptor family, pyridine domain-containing (NLRP) 3 was analyzed using reverse transcription-polymerase chain reaction (RT-PCR). Results: GCE promoted increase in the [Ca(2+)]i in a dose-dependent manner. Depletion of endoplasmic reticulum (ER) Ca(2+) by the ER Ca(2+) ATPase inhibitor thapsigargin (Tg) but not the depletion of extracellular Ca(2+) abolished the GCE-induced increase in [Ca(2+)]i. Treatment of phospholipase C (PLC) inhibitor (U73122) or 1,4,5-trisinositolphosphate (IP3) receptor inhibitor (2-APB) also prevented GCE-induced increase in [Ca(2+)]i. Protease activated receptor (PAR)-2 activation mainly mediated the GCE-induced increase in [Ca(2+)]i and enhanced the expression of IL-8, NLRP3, IL-1β, and IL-6 in HGECs. Conclusions: GCE activates PAR-2, which can induce PLC/IP3-dependent Ca(2+) signaling pathway, ultimately triggering inflammation via the production of pro-inflammatory cytokines such as IL-1β, IL-6, IL-8, and NLRP 3 in HGECs.
    No preview · Article · Nov 2015 · Archives of oral biology
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    ABSTRACT: Purpose: We evaluated whether DA-6034 is involved in mucin secretion via P2Y receptor activation and/or intracellular Ca2+ concentration ([Ca2+]i) change. Also, we investigated the effect of P2Y receptor inhibitors or Ca2+ chelators on the DA-6034-induced mucin secretion and [Ca2+]i increases. Methods: Effects of DA-6034 on mucin expression in primary, cultured, conjunctival epithelial cells was studied using RT-PCR, Western blot analysis, and periodic acid-schiff (PAS) staining. To evaluate thin film layer thickness generated by mucin and fluid secretion, cells were incubated in DA-6034 with/without P2Y antagonists or extracellular/intracellular Ca2+ chelators, and were imaged with confocal microscope using Texas Red-dextran dye. In addition, DA-6034-induced Ca2+-dependent Cl- channels opening was evaluated using perforated patch clamp. Fluo-4/AM was used to measure changes in [Ca2+]i induced by DA-6034 in Ca2+-free or Ca2+-containing buffered condition, as well as P2Y antagonists. Results: DA-6034 induced the expression of mucin genes, production of mucin protein, and increase of number of mucin-secreting cells. P2Y antagonists inhibited DA-6034-induced mucin and fluid secretion, which was also affected by extracellular/intracellular Ca2+ chelators. DA-6034 stimulated Cl- channel opening and [Ca2+]i elevation. Further, [Ca2+]i increases induced by DA-6034 were lacking in either P2Y antagonists or Ca2+-free buffered condition, and diminished when endoplasmic reticulum Ca2+ was depleted by cyclopiazonic acid in Ca2+-free buffered condition. Conclusions: This study demonstrated that DA-6034 has a potential to induce mucin secretion via Ca2+-dependent pathways through P2Y receptors in multilayer, cultured, human conjunctival epithelial cells.
    No preview · Article · Sep 2014 · Investigative Ophthalmology & Visual Science
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    ABSTRACT: Homer proteins are scaffold molecules with a domain structure consisting of an N-terminal Ena/VASP homology 1 protein-binding domain and a C-terminal leucine zipper/coiled-coil domain. The Ena/VASP homology 1 domain recognizes proline-rich motifs and binds multiple Ca2+-signaling proteins, including G protein-coupled receptors, inositol 1,4,5-triphosphate receptors, ryanodine receptors, and transient receptor potential channels. However, their role in Ca2+ signaling in nonexcitable cells is not well understood. In this study, we investigated the role of Homer2 on Ca2+ signaling in parotid gland acinar cells using Homer2-deficient (Homer2−/−) mice. Homer2 is localized at the apical pole in acinar cells. Deletion of Homer2 did not affect inositol 1,4,5-triphosphate receptor localization or channel activity and did not affect the expression and activity of sarco/endoplasmic reticulum Ca2+-ATPase pumps. In contrast, Homer2 deletion markedly increased expression of plasma membrane Ca2+-ATPase (PMCA) pumps, in particular PMCA4, at the apical pole. Accordingly, Homer2 deficiency increased Ca2+ extrusion by acinar cells. These findings were supported by co-immunoprecipitation of Homer2 and PMCA in wild-type parotid cells and transfected human embryonic kidney 293 (HEK293) cells. We identified a Homer-binding PPXXF-like motif in the N terminus of PMCA that is required for interaction with Homer2. Mutation of the PPXXF-like motif did not affect the interaction of PMCA with Homer1 but inhibited its interaction with Homer2 and increased Ca2+ clearance by PMCA. These findings reveal an important regulation of PMCA by Homer2 that has a central role on PMCA-mediated Ca2+ signaling in parotid acinar cells.
    Preview · Article · Jul 2014 · Journal of Biological Chemistry
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    ABSTRACT: DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.
    Full-text · Article · Apr 2014 · Korean Journal of Physiology and Pharmacology
  • Yu-Mi Yang · Sung Jun Lee · Dong Min Shin

    No preview · Article · Dec 2013
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    ABSTRACT: Receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis is accompanied by intracellular Ca(2+) mobilization in a form of oscillations, which plays essential roles by activating sequentially Ca(2+)/calmodulin-dependent protein kinase, calcineurin and NFATc1, necessary in the osteoclast differentiation. However, it is not known whether Ca(2+) mobilization which is evoked in RANKL-independent way induces to differentiate into osteoclasts. In present study, we investigated Ca(2+) mobilization induced by aluminum fluoride (AlF4 (-)), a G-protein activator, with or without RANKL and the effects of AlF4 (-) on the osteoclastogenesis in primary cultured mouse bone marrow-derived macrophages (BMMs). We show here that AlF4 (-) induces intracellular Ca(2+) concentration ([Ca(2+)]i) oscillations, which is dependent on extracellular Ca(2+) influx. Notably, co-stimulation of AlF4 (-) with RANKL resulted in enhanced NFATc1 expression and formation of tartrate-resistant acid phosphatase (TRAP) positive multinucleated cells. Additionally, we confirmed that mitogen-activated protein kinase (MAPK) is also activated by AlF4 (-). Taken together, these results demonstrate that G-protein would be a novel modulator responsible for [Ca(2+)]i oscillations and MAPK activation which lead to enhancement of RANKL-mediated osteoclastogenesis.
    Full-text · Article · Oct 2013 · Korean Journal of Physiology and Pharmacology
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    ABSTRACT: The transient receptor potential melastatin type 7 (TRPM7) channel is a widely expressed non-selective cation channel with fusion to the C-terminal alpha kinase domain and regarded as a key regulator of whole body Mg(2+) homeostasis in mammals. However, the roles of TRPM7 during osteoclastogenesis in RAW264.7 cells and bone marrow-derived monocyte/macrophage precursor cells (BMMs) are not clear. In the present study, we investigate the roles of TRPM7 in osteoclastogenesis using methods of small interfering RNA (siRNA), RT-PCR, patch-clamp, and calcium imaging. RANKL (receptor activator of NF-κB ligand) stimulation did not affect the TRPM7 expression and TRPM7-mediated current was activated in HEK293, RAW264.7, and BMM cells by the regulation of Mg(2+). Knock-down of TRPM7 by siTRPM7 reduced intracellular Ca(2+) concentration ([Ca(2+)](i)) increases by 0 mM [Mg(2+)](e) in HEK293 cells and inhibited the generation of RANKL-induced Ca(2+) oscillations in RAW264.7 cells. Finally, knock-down of TRPM7 suppressed RANKL-mediated osteoclastogenesis such as activation and translocation of NFATc1, formation of multinucleated cells, and the bone resorptive activity, sequentially. These results suggest that TRPM7 plays an essential role in the RANKL-induced [Ca(2+)](i) oscillations that triggers the late stages of osteoclastogenesis.
    Preview · Article · Feb 2013 · Korean Journal of Physiology and Pharmacology
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    ABSTRACT: Protease-activated receptor 2 (PAR2), a G protein-coupled receptor expressed in airway epithelia and smooth muscle, plays an important role in airway inflammation. In this study, we demonstrated that activation of PAR2 induces mucus secretion from the human airway gland and examined the underlying mechanism using the porcine and murine airway glands. The mucosa with underlying submucosal glands were dissected from the cartilage of tissues, pinned with the mucosal side up at the gas/bath solution interface of a physiological chamber, and covered with oil so that secretions from individual glands could be visualized as spherical bubbles in the oil. Secretion rates were determined by optical monitoring of the bubble diameter. The Ca(2+)-sensitive dye Fura2-AM was used to determine intracellular Ca(2+) concentration ([Ca(2+)](i)) by means of spectrofluorometry. Stimulation of human tracheal mucosa with PAR2-activating peptide (PAR2-AP) elevated intracellular Ca(2+) and induced glandular secretion equal to approximately 30% of the carbachol response in the human airway. Porcine gland tissue was more sensitive to PAR2-AP, and this response was dependent on Ca(2+) and anion secretion. When the mouse trachea were exposed to PAR2-AP, large amounts of secretion were observed in both wild type and ΔF508 cystic fibrosis transmembrane conductance regulator mutant mice but there is no secretion from PAR-2 knock out mice. In conclusion, PAR2-AP is an agonist for mucus secretion from the airway gland that is Ca(2+)-dependent and cystic fibrosis transmembrane conductance regulator-independent.
    Full-text · Article · Aug 2012 · PLoS ONE
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    ABSTRACT: Human and pig airway submucosal glands secrete mucus in response to substance P (SubP), but in pig tracheal glands the response to SubP is >10-fold greater than in humans and shares features with cholinergically produced secretion. CFTR-deficient pigs provide a model for human cystic fibrosis (CF), and in newborn CF pigs the response of tracheal glands to SubP is significantly reduced (Joo et al. J Clin Invest 120: 3161-3166, 2010). To further define features of SubP-mediated gland secretion, we optically measured secretion rates from individual adult porcine glands in isolated tracheal tissues in response to mucosal capsaicin and serosal SubP. Mucosal capsaicin (EC(50) = 19 μM) stimulated low rates of secretion that were partially inhibited by tetrodotoxin and by inhibitors for muscarinic, VIP, and SubP receptors, suggesting reflex stimulation of secretion by multiple transmitters. Secretion in response to mucosal capsaicin was inhibited by CFTR(inh)-172, but not by niflumic acid. Serosal SubP (EC(50) = 230 nM) stimulated 10-fold more secretion than mucosal capsaicin, with a V(max) similar to that of carbachol. Secretion rates peaked within 5 min and then declined to a lower sustained rate. SubP-stimulated secretion was inhibited 75% by bumetanide, 53% by removal of HCO(3)(-), and 85% by bumetanide + removal of HCO(3)(-); it was not inhibited by atropine but was inhibited by niflumic acid, clotrimazole, BAPTA-AM, nominally Ca(2+)-free bath solution, and the adenylate cyclase inhibitor MDL-12330A. Ratiometric measurements of fura 2 fluorescence in dissociated gland cells showed that SubP and carbachol increased intracellular Ca(2+) concentration by similar amounts. SubP produced rapid volume loss by serous and mucous cells, expansion of gland lumina, mucus flow, and exocytosis but little or no contraction of myoepithelial cells. These and prior results suggest that SubP stimulates pig gland secretion via CFTR- and Ca(2+)-activated Cl(-) channels.
    Full-text · Article · Dec 2010 · AJP Lung Cellular and Molecular Physiology
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    ABSTRACT: Adequate fluid secretion from airway mucosa is essential for maintaining mucociliary clearance, and fluid hypersecretion is a prominent feature of inflammatory airway diseases such as allergic rhinitis. House dust mite extract (HDM) has been reported to activate protease-activated receptors (PARs), which play various roles in airway epithelia. However, the role of HDM in regulating ion transporters and fluid secretion has not been investigated. We examined the effect of HDM on ion transport in human primary nasal epithelial cells. The Ca(2+)-sensitive dye Fura2-AM was used to determine intracellular Ca(2+) concentration ([Ca(2+)](i)) by means of spectrofluorometry in human normal nasal epithelial cells (NHNE). Short-circuit current (Isc) was measured using Ussing chambers. Fluid secretion from porcine airway mucosa was observed by optical measurement. HDM extract (10 microg/Ml) effectively cleaved the PAR-2 peptide and induced an increase of [Ca(2+)](i) that was abolished by desensitization with trypsin, but not with thrombin. Apical application of HDM-induced Isc sensitive to both a cystic fibrosis transmembrane conductance regulator (CFTR) inhibitor and a Ca(2+)-activated Cl(-) channel (CaCC) inhibitor. HDM extract also stimulated fluid secretion from porcine airway mucosa. HDM extract activated PAR-2 and apical Cl(-) secretion via CaCC and CFTR, and HDM-induced fluid secretion in porcine airway mucosa. Our results suggest a role for PAR-2 in mucociliary clearance and fluid hypersecretion of airway mucosa in response to air-borne allergens such as HDM.
    No preview · Article · Apr 2010 · Journal of Cellular Biochemistry
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    ABSTRACT: RANKL (receptor activator of NF-κB ligand) induces osteoclastogenesis by activating multiple signaling pathways in osteoclast precursor cells, chief among which is induction of long lasting oscillations in the intracellular concentration of Ca2+ ([Ca2+]i). The [Ca2+]i oscillations activate calcineurin, which activates the transcription factor NFATc1. The pathway by which RANKL induces [Ca2+]i oscillations and osteoclastogenesis is poorly understood. Here we report the discovery of a novel pathway induced by RANKL to cause a long lasting increase in reactive oxygen species (ROS) and [Ca2+]i oscillations that is essential for differentiation of bone marrow-derived monocytes into osteoclasts. The pathway includes RANKL-mediated stimulation of Rac1 to generate ROS, which stimulate phospholipase Cγ1 to evoke [Ca2+]i oscillations by stimulating Ca2+ release from the inositol 1,4,5-trisphosphate pool and STIM1-regulated Ca2+ influx. Induction and activation of the pathway is observed only after 24-h stimulation with RANKL and lasts for at least 3 days. The physiological role of the pathway is demonstrated in mice with deletion of the Peroxiredoxin II gene and results in a mark increase is ROS and, consequently, a decrease in bone density. Moreover, bone marrow-derived monocytes in PrxII−/− primary culture show increased ROS and spontaneous [Ca2+]i oscillations. These findings identify the primary RANKL-stimulated pathway to trigger the late stages of osteoclastogenesis and regulate bone resorption.
    Preview · Article · Mar 2010 · Journal of Biological Chemistry
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    ABSTRACT: A mutation of Atp2a2 gene encoding the sarco/endoplasmic reticulum Ca(2+)-ATPase 2 (SERCA2) causes Darier's disease in human and null mutation in one copy of Atp2a2 leads to a high incidence of squamous cell tumor in a mouse model. In SERCA2 heterozygote (SERCA2(+/-)) mice keratinocytes, mechanisms involved in partial depletion of SERCA2 gene and its related tumor induction have not been studied. In this study, we investigated Ca(2+) signaling and differential gene expression in primary cultured keratinocytes from SERCA2(+/-) mice. SERCA2(+/-) keratinocytes showed reduced initial increases in intracellular concentration of calcium in response to ATP, a G-protein coupled receptor agonist, and higher store-operated Ca(2+) entry with the treatment of thapsigargin, an inhibitor of SERCA, compared to wild type kerationcytes. Protein expressions of plasma membrane Ca(2+) ATPases, NFATc1, phosphorylated ERK, JNK, and phospholipase gamma1 were increased in SERCA2(+/-) keratinocytes. Using the gene fishing system, we first found in SERCA2(+/-) keratinocytes that gene level of tumor-associated calcium signal transducer 1, crystalline alphaB, procollagen XVIII alpha1, and nuclear factor I-B were increased. Expression of involucrin, a marker of keratinocyte differentiation, was decreased in SERCA2(+/-) keratinocytes. These results suggest that the alterations of Ca(2+) signaling by SERCA2 haploinsufficiency alternate the gene expression of tumor induction and differentiation in keratinocytes.
    No preview · Article · Oct 2009 · Progress in Biophysics and Molecular Biology
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    ABSTRACT: RANKL is essential for the terminal differentiation of monocytes/macrophages into osteoclasts. RANKL induces long-lasting oscillations in the intracellular concentration of Ca(2+) ([Ca(2+)](i)) only after 24 h of stimulation. These Ca(2+) oscillations play a switch-on role in NFATc1 expression and osteoclast differentiation. Which Ca(2+) transporting pathway is induced by RANKL to evoke the Ca(2+) oscillations and its specific role in RANKL-mediated osteoclast differentiation is not known. This study examined the effect of a partial loss of sarco/endoplasmic reticulum Ca(2+) ATPase type 2 (SERCA2) on osteoclast differentiation in SERCA2 heterozygote mice (SERCA2(+/-)). The BMD in the tibias of SERCA2(+/-) mice increased >1.5-fold compared with wildtype mice (WT). RANKL-induced [Ca(2+)](i) oscillations were generated 48 h after RANKL treatment in the WT mice but not in the SERCA2(+/-) bone marrow-derived macrophages (BMMs). Forty-eight hours after RANKL treatment, there was a lower level of NFATc1 protein expression and markedly reduced translocation of NFATc1 into the nucleus during osteoclastogenesis of the SERCA2(+/-) BMMs. In addition, RANKL treatment of SERCA2(+/-) BMMs incompletely induced formation of multinucleated cells, leading to reduced bone resorption activity. These results suggest that RANKL-mediated induction of SERCA2 plays a critical role in the RANKL-induced [Ca(2+)](i) oscillations that are essential for osteoclastogenesis.
    Preview · Article · Jun 2009 · Journal of bone and mineral research: the official journal of the American Society for Bone and Mineral Research