Establishment of Conditionally Immortalized Rat Retinal Pericyte Cell Lines (TR-rPCT) and Their Application in a Co-culture System Using Retinal Capillary Endothelial Cell Line (TR-iBRB2)

Kondo, Takeru; Hosoya, Ken; Hori, Satoko; Tomi, Masatoshi; Ohtsuki, Sumio; Takanaga, Hitomi; Nakashima, Emi; Iizasa, Hisashi; Asashima, Tomoko; Ueda, Masatsugu; Obinata, Masuo; Terasaki, Tetsuya

doi:10.1247/csf.28.145

Cited by 41 publications

(42 citation statements)

References 36 publications

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“…36,41) RT-PCR analysis indicated the expression of mRNA of VGLUT1 in TR-rPCT1 cells, but not VGLUT2 and VGLUT3 (Fig. 1A), implying the presence of the vesicular transport system for L-Glu via VGLUT1 in retinal pericytes.…”

Section: Discussionmentioning

confidence: 96%

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<i>In Vitro</i> Study of L-Glutamate and L-Glutamine Transport in Retinal Pericytes: Involvement of Excitatory Amino Acid Transporter 1 and Alanine–Serine–Cysteine Transporter 2

Akanuma

Zakoji

Kubo

et al. 2015

Biological & Pharmaceutical Bulletin

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L-Glutamate (L-Glu) is known to be a relaxant of pericytes and to induce changes in microcirculatory hemodynamics. Since the concentration of L-Glu which induces the dilation of retinal capillaries is reported to be high compared with the estimated concentration in the retinal interstitial fluid, it is hypothesized that some systems involving concentrative L-Glu release are present in retinal pericytes. The purpose of this study was to investigate the existence of L-Glu-storing systems, which contribute to autocrine L-Glu release, in retinal pericytes using conditionally immortalized rat retinal pericytes (TR-rPCT1 Key words excitatory amino acid transporter; L-glutamate; retinal pericyte; alanine-serine-cysteine transporter; L-glutamine Retinal pericytes reside within the basal lamina of retinal capillaries and communicate with the retinal capillary endothelial cells which form the blood-retinal barrier (BRB). 1,2)It is known that the contraction and relaxation of pericytes which are induced by physiological factors leads to a change in the diameter of the capillaries. [3][4][5] Because it has been reported that the diameter of capillaries in the central nervous system (CNS) affects the blood flow rate in the capillaries and, thus, the exchange of some compounds between the circulating blood and retina, 6) it is very important to identify the regulatory mechanisms governing the contraction and relaxation of retinal pericytes.L-Glutamate (L-Glu) is known as a relaxant of pericytes and Peppiatt et al. have reported that L-Glu treatment induces dilation of the retinal capillaries. 7) Regarding the mechanism for the L-Glu-induced dilation of retinal capillaries, it has been proposed that L-Glu induces the production of nitric oxide (NO), a relaxant of the cells, and inhibits Ca 2+ influx in retinal pericytes, thereby relaxing the pericytes. 7,8) An in vitro analysis has indicated that this L-Glu-induced NO production occurs in the presence of L-Glu at a concentration of more than 1 mM. 8) In addition, the dilation of retinal capillaries accompanied with pericytes was induced by treatment with 500 µM L-Glu.7) The compounds between the retinal interstitial fluid (ISF) and vitreous humor are considered to be freely exchangeable, and the concentration of L-Glu in the vitreous humor has been reported to be 444 µM in rats.9) Because the L-Glu level in the retinal ISF seems to be low compared with the concentration which induces dilation of the retinal capillaries and relaxation in the pericytes, it is possible that some mechanisms for concentrative L-Glu release are present in retinal pericytes and/or the cells surrounded by the pericytes. In neuronal cells, the packing of L-Glu into synaptic vesicles up to concentrations as high as 100 mM and exocytotic release of L-Glu in the vesicles into the synaptic cleft have been reported. [10][11][12] It has also been reported that the uptake of L-Glu from the cytoplasm of neurons into synaptic vesicles involves the vesicular L-Glu transporters (VGLUTs) which belong to the so...

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Section: Discussionmentioning

confidence: 96%

“…The expression of vesicular L-Glu transporters and L-Glu-synthesizing enzyme was elucidated by a reverse transcription-polymerase chain reaction (RT-PCR) in conditionally immortalized rat retinal pericytes (TR-rPCT1 cells). 36,37) Moreover, the transport characteristics of L-Glu and L-Gln was evaluated by uptake studies using TR-rPCT1 cells.…”

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confidence: 99%

<i>In Vitro</i> Study of L-Glutamate and L-Glutamine Transport in Retinal Pericytes: Involvement of Excitatory Amino Acid Transporter 1 and Alanine–Serine–Cysteine Transporter 2

Akanuma

Zakoji

Kubo

et al. 2015

Biological & Pharmaceutical Bulletin

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“…Choroid plexus epithelial cell line (TR-CSFB) (72) transthyretin, Na + , K + -ATPase The blood-brain barrier (BBB) Brain capillary endothelial cell lines (TR-BBB) (70) von Willebrand factor, acetylated LDLuptake Tight-junction strand proteins (claudine-5, occludin), drug efflux and influx transporters Retinal capillary endothelial cell lines (TR-iBRB) (73) von Willebrand factor, VEGF receptor-2, GLUT1 The blood-retinal barrier (BRB) Retinal pericyte cell lines (TR-rPCTs) (71,74) typical pericyte markers Retinal Müller cell lines (TR-MUL). (75) The blood-placental barrier (BPB) Syncytiotrophoblast cell lines (TR-TBTs) (76) A syncytium-like morphology, carrier-mediated transport The cells from brain Neuronal cell line, N14.5.…”

Section: The Blood-cerebrospinal Fluid (Bcsfb) Barriermentioning

confidence: 99%

“…Retinal pericytes surround retinal capillary endothelial cells have been proposed to play a role in regulation of endothelial cell proliferation. The established rat retinal pericyte cell lines (TR-rPCTs) (74) expressed typical pericyte markers and morphologies as similar to the brain pericyte cell line (TR-PCT). Co-culture studies of TR-rPCT1 and TR-iBRB2 cells suggested that physical contact between pericytes and retinal endothelial cells is important for the growth of retinal endothelial cells.…”

Section: Male Genital Cell Linesmentioning

confidence: 99%

The immortalized cell lines with differentiation potentials: Their establishment and possible application

Obinata

2007

Cancer Science

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“…4 By using tsSV40Tg mice, immortalized PC lines have been established only from certain restricted capillary-rich tissues, such as the retina and capillaries isolated from brain tissue. 21,22 However, PCs from capillaries in other peripheral tissues, especially in the pathological setting, have not been established.…”

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confidence: 99%

Immortalized multipotent pericytes derived from the vasa vasorum in the injured vasculature. A cellular tool for studies of vascular remodeling and regeneration

Kabara

Kawabe

Matsuki

et al. 2014

Laboratory Investigation

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Adventitial microvessels, vasa vasorum in the vessel walls, have an active role in the vascular remodeling, although its mechanisms are still unclear. It has been reported that microvascular pericytes (PCs) possess mesenchymal plasticity. Therefore, microvessels would serve as a systemic reservoir of stem cells and contribute to the tissues remodeling. However, most aspects of the biology of multipotent PCs (mPCs), in particular of pathological microvessels are still obscure because of the lack of appropriate methods to detect and isolate these cells. In order to examine the characteristics of mPCs, we established immortalized cells residing in adventitial capillary growing at the injured vascular walls. We recently developed in vivo angiogenesis to observe adventitial microvessels using collagen-coated tube (CCT), which also can be used as an adventitial microvessel-rich tissue. By using the CCT, CD146-or NG2-positive cells were isolated from the adventitial microvessels in the injured arteries of mice harboring a temperature-sensitive SV40 T-antigen gene. Several capillary-derived endothelial cells (cECs) and PCs (cPCs) cell lines were established. cECs and cPCs maintain a number of key endothelial and PC features. Co-incubation of cPCs with cECs formed capillary-like structure in Matrigel. Three out of six cPC lines, termed capillary mPCs demonstrated both mesenchymal stem cell-and neuronal stem cell-like phenotypes, differentiating effectively into adipocytes, osteoblasts, as well as schwann cells. mPCs differentiated to ECs and PCs, and formed capillary-like structure on their own. Transplanted DsRed-expressing mPCs were resident in the capillary and muscle fibers and promoted angiogenesis and myogenesis in damaged skeletal muscle. Adventitial mPCs possess transdifferentiation potential with unique phenotypes, including the reconstitution of capillary-like structures. Their phenotype would contribute to the pathological angiogenesis associated with vascular remodeling. These cell lines also provide a reproducible cellular tool for high-throughput studies on angiogenesis, vascular remodeling, and regeneration as well.

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Establishment of Conditionally Immortalized Rat Retinal Pericyte Cell Lines (TR-rPCT) and Their Application in a Co-culture System Using Retinal Capillary Endothelial Cell Line (TR-iBRB2)

Cited by 41 publications

References 36 publications

<i>In Vitro</i> Study of L-Glutamate and L-Glutamine Transport in Retinal Pericytes: Involvement of Excitatory Amino Acid Transporter 1 and Alanine–Serine–Cysteine Transporter 2

<i>In Vitro</i> Study of L-Glutamate and L-Glutamine Transport in Retinal Pericytes: Involvement of Excitatory Amino Acid Transporter 1 and Alanine–Serine–Cysteine Transporter 2

The immortalized cell lines with differentiation potentials: Their establishment and possible application

Immortalized multipotent pericytes derived from the vasa vasorum in the injured vasculature. A cellular tool for studies of vascular remodeling and regeneration

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