• 한국산학기술학회논문지
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• 제12권7호
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• pp.3096-3102
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• 2011

바닐로이드 수용체 TRPV1(transient receptor potential vanilloid 1)은 캡사이신, pH, 열 등의 통증 유발물질에 의해 활성화되는 비특이적 양이온 채널로서 통증발현에 핵심적인 막 단백질이다. TRPV1의 막 수송에 관한 연구가 미미한 가운데 FIP3(family of Rab11 interacting protein 3)가 TRPV1 채널과 결합하여 막수송에 관여한다고 보고되었다. FIP3는 Rab11과 결합하는 단백질인데 최근 Rab11 단백질이 여러 채널 단백질의 막수송에 직접적으로 또는 간접적으로 중요하다고 보고되었다. 그러므로 본 연구에서는 Rab11이 TRPV1의 막 수송에서의 역할을 알아보기 위하여 세포 생물학적, 생화학적으로 알아보았다. 공촛점 현미경을 통하여 확인한 결과 Rab11은 실제로 세포내에서 TRPV1과 동일한 위치에서 발현되어 있음을 확인하였다. 하지만 생화학적인 방법인 GST-pulldown을 실시하였을 때 TRPV1과 Rab11간에는 서로 직접적인 결합은 하지 않는 것으로 나타났다. 비록 직접적인 결합은 하지 않지만 Rab11이 TRPV1의 막 수송에 관여한다고 가정하고 Rab11의 TRPV1의 막수송에서의 역할을 더 자세히 알아보기 위하여 세포내 Rab11a의 발현을 siRNA를 사용하여 Rab11a의 발현을 50%수준으로 저해한 후 TRPV1의 세포막으로의 이동을 알아본 결과 Rab11 발현 저해 시 세포막에 이동된 TRPV1이 현저히 감소함을 확인할 수 있었다. 이 결과로부터 Rab11이 아마도 FIP3을 포함하는 방법으로 TRPV1의 막 수송에 영향을 주는 것으로 결론지을 수 있다.

• International Journal of Oral Biology
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• 제46권3호
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• pp.119-126
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• 2021

Activation of transient receptor potential vanilloid 1 (TRPV1), a calcium permeable channel expressed in primary sensory neurons, induces the release of glutamate from their central and peripheral afferents during normal acute and pathological pain. However, little information is available regarding the glutamate release mechanism associated with TRPV1 activation in primary sensory neurons. To address this issue, we investigated the expression of vesicular glutamate transporter (VGLUT) in TRPV1-immunopositive (+) neurons in the rat trigeminal ganglion (TG) under normal and complete Freund's adjuvant (CFA)-induced inflammatory pain conditions using behavioral testing as well as double immunofluorescence staining with antisera against TRPV1 and VGLUT1 or VGLUT2. TRPV1 was primarily expressed in small and medium-sized TG neurons. TRPV1+ neurons constituted approximately 27% of all TG neurons. Among all TRPV1+ neurons, the proportion of TRPV1+ neurons coexpressing VGLUT1 (VGLUT1+/TRPV1+ neurons) and VGLUT2 (VGLUT2+/TRPV1+ neurons) was 0.4% ± 0.2% and 22.4% ± 2.8%, respectively. The proportion of TRPV1+ and VGLUT2+ neurons was higher in the CFA group than in the control group (TRPV1+ neurons: 31.5% ± 2.5% vs. 26.5% ± 1.2%, VGLUT2+ neurons: 31.8% ± 1.1% vs. 24.6% ± 1.5%, p < 0.05), whereas the proportion of VGLUT1+, VGLUT1+/TRPV1+, and VGLUT2+/TRPV1+ neurons did not differ significantly between the CFA and control groups. These findings together suggest that VGLUT2, a major isoform of VGLUTs, is involved in TRPV1 activation-associated glutamate release during normal acute and inflammatory pain.

• 한국산학기술학회논문지
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• 제12권1호
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• pp.312-317
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• 2011

캡사이신 채널로 알려진 바닐로이드 수용체 TRPV1 (캡사이신채널, Transient Receptor Potential Vanilloid 1)은 통증발현에서 중요한 역할을 하는 것으로 알려져 있다. 하지만 TRPV1의 활성조절에 관여하는 단백질에 대하여는 알려진 바가 많지 않다. 최근 rat TRPV1과 직접적으로 결합하는 단백질을 탐색하여 mouse Rab11-FIP3 (rab11-family interaction protein 3)가 rat TRPV1과 직접적으로 결합한다는 것이 보고되었다. Rab11은 여러 가지의 세포내 이동에 관여하는 것으로 보고되었다. 그러므로 Rab11-FIP3과의 결합을 통해 TRPV1의 세포막으로의 이동에 관여할 것으로 추측할 수 있다. 본 연구에서는 전에 보고된 연구가 mouse와 rat 이라는 다른 종의 단백질끼리의 결합이기 때문에 같은 종에서의 상호작용을 확인하고 Rab11-FIP3의 TRPV1의 세포막으로의 이동에서의 역할을 알아보고자 현재까지 동정되지 않은 rat의 Rab11-FIP3의 유전자를 GenBank 서열을 바탕으로 rat 뇌의 RNA 로부터 cDNA 를 클로닝하여 유전자를 분리하고 TRPV1 과의 관계를 세포생물학적으로 알아보았다. 연구결과 rat의 Rab11-FIP3는 489개의 아미노산 서열을 가지고 있으며 human과는 80%, mouse와는 90% 이상 아미노산 서열의 상동성을 보였다. 조직별 분포는 심장, 뇌, 간, 콩팥, 정소에서 발현되고 있는 것을 northern blot assay와 western blot assay 로 확인하였다. rat 의 뇌조직에서 TRPV1 과 Rab11-FIP3 단백질이 결합하여 colocalize 하는 것을 면역화학방법으로 확인하였다. 이 결합은 같은 family 의 TRPV2 와는 결합하지 않는 특이적 결합이므로 Rab11-FIP3 가 TRPV1 과 상호작용하여 세포막으로의 이동에 관여할 것이라는 것을 시사한다.

• 한국동물생명공학회지
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• 제34권4호
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• pp.311-317
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• 2019

Reproductive potential decreases with age. A decrease in male fertility is due to a combination of morphological and molecular alterations in the testes. Transient receptor potential vanilloid receptor-1 (TRPV1) is associated with aging and lifespan, and its activation causes apoptotic cell death in various cell types. However, the effect of TRPV1 on testicular apoptosis in aged mice has not yet been reported. TRPV1 knockout (KO) mice had a longer lifespan than that of wild-type (WT) mice. Lifespan was increased by 11.8% in male TRPV1 KO mice compared to that in WT mice. TRPV1 KO males lived approximately 100 days longer than WT males on average, and the maximum lifespan was markedly extended in TRPV1 KO mice compared with that in WT mice. The TRPV1 expression levels were highly increased in the testes of older mice. TRPV1 was expressed in the entire testes region of the old mice. In addition, old TRPV1 KO mice had lower testicular apoptosis than that of WT mice. Our results show that TRPV1 induces testicular apoptosis and suggest that TRPV1 may be associated with testicular aging.

• Yonsei Medical Journal
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• 제59권9호
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• pp.1131-1137
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• 2018

Purpose: Previous reports have shown that hyperglycemia-induced inhibition of transient receptor potential vanilloid sub type 4 (TRPV4), a transient receptor potential ion channel, affects the severity of hearing impairment (HI). In this study, we explored the role of TRPV4 in HI using HEI-OC1 cells exposed to high glucose (HG). Materials and Methods: HEI-OC1 cells were cultured in a HG environment (25 mM D-glucose) for 48 hours, and qRT-PCR and Western blotting were used to analyze the expression of TRPV4 at the mRNA and protein level. TRPV4 agonist (GSK1016790A) or antagonist (HC-067047) in cultured HEI-OC1 cells was used to obtain abnormal TRPV4 expression. Functional TRPV4 activity was assessed in cultured HEI-OC1 cells using the MTT assay and a cell death detection ELISA. Results: TRPV4 agonists exerted protective effects against HG-induced HI, as evidenced by increased MTT levels and inhibition of apoptosis in HEI-OC1 cells. TRPV4 overexpression significantly increased protein levels of phosphorylated p38 mitogen-activated protein kinase (p-p38 MAPK), while TRPV4 antagonists had the opposite effect. Our results indicated that TRPV4 is a hyperglycemia-related factor that can inhibit cell proliferation and promote cell apoptosis by activating the MAPK signaling pathway in HEI-OC1 cells. Conclusion: Our results show that the overexpression of TRPV4 can attenuate cell death in HEI-OC1 cells exposed to HG.

• Applied Microscopy
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• 제42권1호
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• pp.27-33
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• 2012

Transient receptor potential ankyrin 1 (TRPA1)은 $17^{\circ}C$보다 낮은 유해한 온도 및 자극적인 화합물에 의해 활성화되며 통각을 조절한다. 그러나 TRPA1에 의한 통각정보가 어떻게 처리되는지에 대한 정보는 많이 알려져 있지 않다. 본 연구에서는 흰쥐의 삼차신경절에서 TRPA1을 발현하는 신경세포의 특성을 규명하기 위해서, 면역형광기법을 사용하여 TRPA1을 발현하는 신경세포에서 다른 통각수용기들에서 발현되며, 특징적인 기능을 수행하는 수용기인 transient receptor potential vanilloid 1 (TRPV1)와 $P2X_3$와의 발현양상을 조사하였다. TRPA1을 발현하는 신경세포에서 열감각수용기이며, 통각표지자인 TRPV1과의 공존을 조사해 본 결과, TRPA1 면역양성 신경세포 중에서 58.8% (328/558)가 TRPV1을 동시에 발현하였으며, 41.2% (230/558)가 TRPA1만 발현하고 TRPV1을 발현하지 않았다. TRPA1을 발현하는 신경세포 중 TRPV1을 동시에 발현하는 신경세포는 대부분 작거나 중간크기였다. 또한 TRPA1과 조직의 손상, 그리고 염증 시 분비되는 ATP와 결합하는 $P2X_3$와의 공존을 조사해 본 결과, TRPA1 면역양성 신경세포 중에서 26.1% (310/1186)의 신경세포에서 $P2X_3$을 동시에 발현하였으며, 73.9% (876/1186)의 신경세포에서 TRPA1만 발현하였다. TRPA1을 발현하는 신경세포 중 $P2X_3$을 동시에 발현하는 신경세포는 대부분 작거나 중간크기였다. 이러한 결과는 TRPA1을 발현하는 신경세포가 TRPV1 또는 $P2X_3$를 동시에 발현함으로써 동일한 신경세포가 구강안면영역에서의 냉통각 및 열통각을 조절할 뿐 아니라, 냉통각 및 염증성동통을 동시에 전달하는 등 하나의 신경세포가 여러 가지 통각의 전달에 관여하는 것을 시사한다.

• Animal cells and systems
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• 제15권3호
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• pp.181-187
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• 2011

Transient receptor potential vanilloid type 1 (TRPV1) receptor plays an important role as a molecular detector of noxious signals in primary sensory neurons. Activity of TRPV1 can be modulated by the change in the environment such as redox state and extracellular cations. In the present study, we investigated the effect of the mercury chloride ($HgCl_2$) on the activity of TRPV1 in rat dorsal root ganglia (DRG) neurons using whole-cell patch clamp technique. Extracellular $HgCl_2$ reversibly reduced the magnitudes of capsaicin-activated currents ($I_{cap}$) in DRG neurons in a dose-dependent manner. The blocking effect of $HgCl_2$ was prevented by pretreatment with the reducing agent dithiothreitol (DTT). Inhibition of $I_{cap}$ by $HgCl_2$ was abolished by point mutation of individual cysteine residues located on the extracellular surface of TRPV1. These results suggest that three extracellular cysteines of TRPV1, Cys616, Cys634 and Cys621, are responsible for the oxidative modulation of $I_{cap}$ by $HgCl_2$.

• Molecules and Cells
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• 제37권3호
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• pp.257-263
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• 2014

A mammalian cell renovates itself by autophagy, a process through which cellular components are recycled to produce energy and maintain homeostasis. Recently, the abundance of gap junction proteins was shown to be regulated by autophagy during starvation conditions, suggesting that transmembrane proteins are also regulated by autophagy. Transient receptor potential vanilloid type 1 (TRPV1), an ion channel localized to the plasma membrane and endoplasmic reticulum (ER), is a sensory transducer that is activated by a wide variety of exogenous and endogenous physical and chemical stimuli. Intriguingly, the abundance of cellular TRPV1 can change dynamically under pathological conditions. However, the mechanisms by which the protein levels of TRPV1 are regulated have not yet been explored. Therefore, we investigated the mechanisms of TRPV1 recycling using HeLa cells constitutively expressing TRPV1. Endogenous TRPV1 was degraded in starvation conditions; this degradation was blocked by chloroquine (CLQ), 3MA, or downregulation of Atg7. Interestingly, a glucocorticoid (cortisol) was capable of inducing autophagy in HeLa cells. Cortisol increased cellular conversion of LC3-I to LC-3II, leading autophagy and resulting in TRPV1 degradation, which was similarly inhibited by treatment with CLQ, 3MA, or downregulation of Atg7. Furthermore, cortisol treatment induced the colocalization of GFP-LC3 with endogenous TRPV1. Cumulatively, these observations provide evidence that degradation of TRPV1 is mediated by autophagy, and that this pathway can be enhanced by cortisol.

• 한국동물생명공학회지
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• 제36권3호
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• pp.145-153
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• 2021

The role of transient receptor potential vanilloid receptor-1 (TRPV1) has been primarily investigated in pain sensory neurons. Relatively, little research has been performed in testicular cells. TRPV1 is abundantly expressed in Leydig cells of young adult mice. This study was conducted to determine the role of the TRPV1 channel in Leydig cells. TRPV1 modulators and testosterone were treated to the mouse Leydig cell line TM3 cells for 24 h. Capsaicin, a TRPV1 activator, dose-dependently induced cell death, whereas capsazepine, a TRPV1 inhibitor, inhibited capsaicin-induced cell death. Testosterone treatment reduced capsaicin-induced cell death. High concentrations of testosterone decreased TRPV1 mRNA and protein expression levels. However, TRPV1 modulators did not affect testosterone production. These results showed that capsaicin induced cell death of Leydig cells and that testosterone reduced capsaicin-induced cell death. Our findings suggest that testosterone may regulate the survival of Leydig cells in young adult mice by decreasing the expression level of TRPV1.

• The Korean Journal of Pain
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• 제34권3호
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• pp.262-270
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• 2021

Background: Transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel implicated in pain sensation in response to heat, protons, and capsaicin (CAPS). It is well established that TRPV1 is involved in mechanical allodynia. This study investigates the effect of Ononis spinosa (Fabaceae) in CAPS-induced mechanical allodynia and its mechanism of action. Methods: Mechanical allodynia was induced by the intraplantar (ipl) injection of 40 ㎍ CAPS into the left hind paw of male Wistar rats. Animals received an ipl injection of 100 ㎍ O. spinosa methanolic leaf extract or 2.5% diclofenac sodium 20 minutes before CAPS injection. Paw withdrawal threshold (PWT) was measured using von Frey filament 30, 90, and 150 minutes after CAPS injection. A molecular docking tool, AutoDock 4.2, was used to study the binding energies and intermolecular interactions between O. spinosa constituents and TRPV1 receptor. Results: The ipsilateral ipl injection of O. spinosa before CAPS injection increased PWT in rats at all time points. O. spinosa decreased mechanical allodynia by 5.35-fold compared to a 3.59-fold decrease produced by diclofenac sodium. The ipsilateral pretreatment with TRPV1 antagonist (300 ㎍ 4-[3-Chloro-2-pyridinyl]-N-[4-[1,1-dimethylethyl] phenyl]-1-piperazinecarboxamide [BCTC]) as well as the β2-adrenoreceptor antagonist (150 ㎍ butoxamine) attenuated the action of O. spinosa. Depending on molecular docking results, the activity of the extract could be attributed to the bindings of campesterol, stigmasterol, and ononin compounds to TRPV1. Conclusions: O. spinosa alleviated CAPS-induced mechanical allodynia through 2 mechanisms: the direct modulation of TRPV1 and the involvement of β2 adrenoreceptor signaling.