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-->   Miho Matsuda Last modified date：2024.05.10 Associate Professor / Division of Oral Biological Sciences Department of Dental Science Faculty of Dental Science 1 Research Interests 2 Current and Past Project 3 Academic Activities 3.1 Papers 3.2 Reports 3.3 Presentations 4 Membership in Academic Society 5 Awards 6 Educational Activities 7 Other Educational Activities 8 Professional and Outreach Activ... Graduate School Department of Dental Science Graduate School of Dental Science E-Mail　*Since the e-mail address is not displayed in Internet Explorer, please use another web browser:Google Chrome, safari. Homepage https://kyushu-u.elsevierpure.com/en/persons/miho-matsuda　Reseacher Profiling Tool　Kyushu University Pure http://www.dent.kyushu-u.ac.jp/sosiki/a04/index.html Phone 092-642-6321 Fax 092-642-6322 Academic Degree PhD Country of degree conferring institution (Overseas) No Field of Specialization molecular biology biochemistry cell biology Total Priod of education and research career in the foreign country 03years00months Research Research Interests Study on intestinal immunity system and an novel molecular mechanism of bioregulationkeyword : GALT, PRIP, lymphoid cell2021.04. Study on novel molecular mechanisms in the regulation of bone metabolismkeyword : bone metabolism, signal transduction, PRIP, hormone, secretion2008.04. Function of a novel inositol 1,4,5-trisphosphate binding protein, PRIP, in the regulation of reproductive systemkeyword : hormone, secretion, reproductive system, gonadotropin2006.05. Current and Past Project Study on the involvement of a novel signaling molecule, PRIP in the regulation of bone metabolism. Study on the involvement of a novel signaling molecule, PRIP in the regulation of bone metabolism. Study on Ca2+ dependent regulation of gene expressions. Study on brain specific regulation of PRIP-1 gene expression. Academic Activities Reports  Show All Reports >> Papers  Show All Papers >> 1. Miho Matsuda, Masato Hirata, Phospholipase C-related but catalytically inactive proteins regulate ovarian follicle development, ASBMB/ Journal of Biological Chemistry, 292, 20, 8369-8380, 2017.04, Phospholipase C-related but catalytically inactive proteinsPRIP-1 and -2 are inositol-1,4,5-trisphosphate binding proteinsthat are encoded by independent genes. Ablation of the PRIPgenes in mice impairs female fertility, which is manifestedby fewer pregnancies, a decreased number of pups, and thedecreased and increased secretion of gonadal steroids andgonadotropins, respectively. We investigated the involvementof the PRIPs in fertility, focusing on the ovaries of Prip-1 and -2double-knock-out (DKO) mice. Multiple cystic follicles wereobserved in DKO ovaries, and a superovulation assay showed amarkedly decreased number of ovulated oocytes. Cumulusoocytecomplexes showed normal expansion, and artificialgonadotropin stimulation regulated the ovulation-related genesin a normal fashion, suggesting that the ovulation itself wasprobably normal.Ahistological analysis showed atresia in fewerfollicles of the DKO ovaries, particularly in the secondary folliclestages. The expression of luteinizing hormone receptor(LHR) was aberrantly higher in developing follicles, and thephosphorylation of extracellular signal-regulated protein kinase,a downstream target of LH-LHR signaling, was higher in DKOgranulosa cells. This suggests that the up-regulation of LH-LHRsignaling is the cause of impaired follicle development. Theserum estradiol level was lower but estradiol production wasunchanged in theDKOovaries. These results suggest that PRIPsare positively involved in the development of follicles via theirregulation of LH-LHR signaling and estradiol secretion. FemaleDKO mice had higher serum levels of insulin, testosterone, anduncarboxylated osteocalcin, which together with reduced fertility,are reminiscent of polycystic ovary syndrome in humans.. 2. Ayako Murakami, Miho Matsuda, Masato Hirata, Phospholipase C-related, but catalytically inactive protein (PRIP) up-regulates osteoclast differentiation via calcium-calcineurin-NFATc1 signaling, Journal of Biological Chemistry, 292, 7994-8006, 2017.03, Phospholipase C-related, but catalytically inactive protein(PRIP) was previously identified as a novel inositol 1,4,5-trisphosphate-binding protein with a domain organization similarto that of phospholipase C- but lacking phospholipase activity.We recently showed that PRIP gene knock-out (KO) in miceincreases bone formation and concomitantly decreases boneresorption, resulting in increased bone mineral density and trabecularbonevolume.However, the role ofPRIPin osteoclastogenesishas not yet been fully elucidated. Here, we investigated the effectsof PRIP on bone remodeling by investigating dynamic tooth movementin mice fitted with orthodontic devices. Morphological analysisindicated that the extent of tooth movement was smaller inthe PRIP-KO mice than in wild-type mice. Histological analysisrevealed fewer osteoclasts on the bone-resorption side in maxillarybones of PRIP-KO mice, and osteoclast formation assays and flowcytometry indicated lower osteoclast differentiation in bone marrowcells isolated from these mice. The expression of genes implicatedinboneresorptionwaslower in differentiatedPRIP-KOcells,and genes involved in osteoclast differentiation, such as the transcriptionfactor NFATc1, exhibited lower expression in immaturePRIP-KO cells initiated by M-CSF. Moreover, calcineurin expressionand activity were also lower in the PRIP-KO cells. ThePRIP-KO cells also displayed fewer M-CSF-induced changes inintracellular Ca2 and exhibited reduced nuclear localization ofNFATc1. Up-regulation of intracellularCa2restored osteoclastogenesisof the PRIP-KO cells. These results indicate that PRIP deficiencyimpairs osteoclast differentiation, particularly at the earlystages, and that PRIP stimulates osteoclast differentiation throughcalcium-calcineurin-NFATc1 signaling via regulating intracellularCa2.. 3. Matsuda, M, Tsutsumi, K., Kanematsu, T., Fukami, K., Terada, Y., Takenawa, T., Nakayama, K.I., Hirata, M., Involvement of phospholipase C-related inactive protein in the mouse reproductive system through the regulation of gonadotropin levels. , Biology of reproduction, 81, 681, 2009.07. Presentations  Show All Presentations >> 1. Bone is an important tissue contributing to the regulation of homeostasis in the body and bone remodeling is stringently regulated by a balance between bone resorption by osteoclasts and bone formation by osteoblasts.Phospholipase C (PLC)-related, but catalytically inactive protein (PRIP) was first identified as an inositol 1,4,5-trisphosphate binding protein, which is conserved among different species, but its biological function remains elusive. We therefore generated PRIP gene-deficient (KO) mice to gain insights into the biological function. Base on the expression pattern of PRIP, we initially focused on its function in the inhibitory signaling in the central nervous system and in the regulation of protein phosphorylation in signal transduction. However, we unexpectedly found an increased bone mass in KO mice and therefore have been studying the involvement of PRIP in bone metabolism. Our results indicate that PRIP positively regulates osteoclast differentiation through calcium-calcineurin-NFAT signaling.We thus revealed the involvement of PRIP in bone metabolism through reverse genetics, however we never imagined of this possibility when we generated the KO mice. Reverse genetics is one of the best approaches to identify diverse function of a gene/protein. Furthermore, it all started when we noticed a thing in KO mice.. 2. Miho Matsuda, Ayako Murakami, Masato Hirata, Involvement of phospholipase C-related but catalytically inactive protein in the early stage of osteoclast differentiation, cell biology 2016 ASCB Annual Meeting, 2016.12, Phospholipase C-related but catalytically inactive protein (PRIP) was first identified as an inositol 1,4,5-trisphosphate binding protein. We generated PRIP gene-deficient (PRIP-KO) mice and reported that PRIP-KO mice exhibited the upregulation of the bone mineral density and trabecular bone volume, due to the enhancement of bone formation while decreased bone resorption. In this study, we investigated the possible mechanisms by which PRIP regulates bone resorption. Orthodontic tooth movement was performed using wild type (WT) and PRIP-KO mice equipped with orthodontic devices and analyzed by micro-computed tomography and TRAP staining. In the mutant mice, tooth movement was reduced compared with wild type (WT) mice, and osteoclasts were observed at the lesser amount on pressure side in KO maxillary bones. In vitro osteoclast formation assay and flow cytometric analysis were also performed and revealed the decreased differentiation for osteoclast in bone marrow cells isolated from KO mice. The expression of genes implicated in osteoclast differentiation and bone resorption was decreased in KO mice. Immunoblot analysis using cultured bone marrow cells indicated the lower expression of calcineurin in KO cells, and the phosphatase activity by calcineurin was decreased in KO cells. The expression of nuclear factor of activated T cells, cytoplasmic 1 (Nfatc1), which is an essential factor for osteoclast differentiation and the activity is regulated by calcineurin, was also decreased in KO cells in the early stage of osteoclast differentiation. These results suggest that PRIP positively regulates osteoclast differentiation, especially in the early stage, probably through calcium-calcineurin-NFAT signaling. . 3. Phospholipase C-related catalytically inactive protein (comprising PRIP-1 and -2) was first identified as a novel inositol 1,4,5-trisphosphate binding protein, but the biological functions have remained elusive. We therefore generated PRIP-1 and -2 double knockout (DKO) mice and found their reduced fertility: DKO mice exhibited reduced fertility in the female such as decreased number of pups, longer estrous days, increased secretion of gonadotropins, etc. This time we examined the ability of the ovaries in response to gonadotropins: the total number of the ovulated oocytes were remarkably decreased in DKO female mice, indicating that PRIP plays an important role in follicle maturation and/or ovulation, but not fertilization or implantation. Microarray and quantitative real-time PCR analyses revealed the increased expressions of molecules involved in ovulation including Has2, Ptgs2, PTX3, Tnfaip6 and Areg in both genotype. We examined the ability of COC expansion in both genotypes but little difference was observed. On the other hand, histological analysis revealed more immature follicles and fewer mature follicles or corpus lutea in DKO ovaries. Immunological analysis showed the expression of luteinizing hormone receptors (LHR) at earlier stages of follicle maturation in DKO ovary, and elevated level of ERK phosphorylation in DKO granulosa cells. These results suggest that the appearance of LHR at immature follicle stages leads to suppress follicle maturation by excessive LH signaling in DKO ovary, indicating that PRIP may be involved in positive regulation of ovarian follicle maturation through LH signaling. . 4. Phospholipase C-related catalytically inactive protein (comprising PRIP-1 and -2) was first identified as a novel inositol 1,4,5-trisphosphate binding protein, but the biological functions have remained elusive. We therefore generated PRIP-1 and –2 double knockout (DKO) mice to gain insight into the biological function. DKO mice exhibited reduced fertility in the female such as decreased number of pups, longer estrous days, increased secretion of gonadotropins, etc. We examined the ability of the ovaries in response to gonadotropins: the total number of the ovulated oocytes were remarkably decreased in DKO female mice, indicating that PRIP plays an important role in follicle maturation and/or ovulation, but not fertilization or implantation. Microarray and quantitative real-time PCR analyses revealed the increased expressions of molecules involved in ovulation including Has2, Ptgs2, PTX3, Tnfaip6 and Areg. We examined the ability of COC expansion in both genotypes but little difference was observed. On the other hand, histological analysis of ovaries from both genotypes revealed more immature follicles and fewer mature follicles or corpus lutea in DKO ovaries. Immunofluorescence analysis showed the expression of luteinizing hormone receptors (LHR) at earlier stages of follicle maturation in DKO ovary and more increase of antral follicles. These results are summarized as below: the increased LH secretion from pituitary gland and the appearance of LHR at immature follicle stages lead excessive LH signaling to suppress follicle maturation in DKO females, indicating that PRIP may be involved in positive regulation of ovarian follicle maturation through LH signaling. . 5. Miho Matsuda, Masato Hirata, Involvement of a novel signaling molecule, PRIP in the regulation of reproduction system, The 22th IUBMB, 37st FEBS Congress, 2012.09, PRIP (PLC-Related, but catalytically Inactive Protein) は、Ins(1,4,5)P3 結合性の分子として見いだされた新規分子であり、種を越えて広く存在するタンパク質である。２つのサブタイプが存在し、PRIP-1は脳に多く発現しているのに対し、PRIP-2は比較的ユビキタスな発現パターンを示す。我々は、PRIP-1及びPRIP-2のダブルノックアウトマウスを作製し、本変異マウスにおいて総出産仔数の減少、血中性腺刺激ホルモン量の増加などの表現型を見いだし、性周期制御にPRIPが強く関与することを明らかにした。排卵数を調べるため、PMSG（妊馬血清性ゴナドトロピン）、HCG（人絨毛生性腺刺激ホルモン）を用いた排卵誘発を行ったところ、変異型で激減していた。このことから、出産仔数の減少は、排卵後の授精や着床における異常によるものではなく、卵成熟を含めた排卵に至るまでの過程における異常によるものであることが示唆された。そこで、まず排卵前後における遺伝子発現の変化を検討するために、卵巣RNAを用いたマイクロアレイ解析を行った。その結果、PMSG投与後48時間でHCGを投与した卵巣において、排卵前の卵胞で発現するHas2 (hyaluronan synthase 2)、Ptgs2 (cyclooxygenase 2)、PTX3 (pentraxin 3)、Tnfαip6 (TNFα-induced protein 6)などの発現量が、変異型で増加していた。また、同時期の卵巣の組織学的解析より、変異型において卵母細胞が残ったまま黄体化したような所見があり、これらのことから排卵時の制御におけるPRIPの関与が示唆された。そこで現在、野生型及び変異型マウスにおいて、排卵直前に起こる急激な変化（卵丘細胞-卵母細胞複合体のexpansionやヒアルロン酸の発現量及び局在等）について解析を進めている。. 6. Phospholipase C-related but catalytically inactive protein (comprising PRIP-1 and -2) was first identified as a novel D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] binding protein, but the biological functions have remained elusive. We therefore generated PRIP-1 and –2 double knockout (DKO) mice to gain insight into the biological function. DKO mice showed dysfunction of the reproductive system in the female. We examined the ability of the ovaries in response to gonadotropins: the total number of the ovulated oocytes were significantly decreased in mutant female mice, compared to that seen with wild-type, which was also confirmed by histological analysis of the ovaries after ovulation. These results suggested that PRIP plays an important role in not fertilization or implantation but follicle maturation and/or ovulation. Microarray analysis indicated the increased expressions of ovulation related molecules such as Has2, Ptgs2, PTX3, Tnfαip6 and Ereg. Historogical analysis of ovaries, just before ovulation, showed some abnormal follicles in the DKO. We also examined the ability of COC expansion in both genotypes and the significantly difference was not observed. On the other hand, TUNEL assay was performed to detect apoptotic follicles related to follicle maturation. There were some differences between in WT and DKO ovaries, depending on the follicle stages. Further analyses are underway to reveal the role of PRIP on follicle maturation and ovulation. . 7. Phospholipase C-related but catalytically inactive protein (comprising PRIP-1 and -2) was first identified as a novel D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] binding protein, but the biological functions have remained elusive. We therefore generated PRIP-1 and –2 double knockout (DKO) mice to gain insight into the biological function. DKO mice apparently grew normally and became fertile; however, during animal maintenance, we noticed that mutant couples exhibited decreased litter events and litter size, indicating dysfunction of the reproductive system. Cross-mating experiments indicated that the cause appeared to be on the female side. The observation of the estrous cycle in mice by histological analysis of vaginal smears showed that the estrous days were apparently increased in DKO mice. Levels of serum LH and FSH were measured for 5-6 consecutive days, and were significantly higher in the mutant, which was also confirmed by examining the secretion of LH from the explant culture of anterior pituitary glands of wild-type and DKO mice. We also examined the ability of the ovaries in response to gonadotropins: the total number of the ovulated oocytes were significantly decreased in mutant female mice, compared to that seen with wild-type, which was also confirmed by histological analysis of the ovaries after ovulation. These results suggest that PRIP plays an important role in female reproductive system, especially in gonadotropin secretion, ovarian follicle maturation and ovulation.. 8. Phospholipase C-related but catalytically inactive protein (comprising PRIP-1 and -2) was first identified as a novel D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] binding protein, but the biological functions have remained elusive. We therefore generated PRIP-1 and –2 double knockout (DKO) mice to gain insight into the biological function. DKO mice apparently grew normally and became fertile; however, during animal maintenance, we noticed that mutant couples exhibited decreased litter events and litter size, indicating dysfunction of the reproductive system. Cross-mating experiments indicated that the cause appeared to be on the female side. The observation of the estrous cycle in mice by histological analysis of vaginal smears showed that the estrous days were apparently increased in DKO mice. Levels of serum LH and FSH were measured for 5-6 consecutive days, and were significantly higher in the mutant, which was also confirmed by examining the secretion of LH from the explant culture of anterior pituitary glands of wild-type and DKO mice. We also examined the ability of the ovaries in response to gonadotropins: the total number of the ovulated oocytes were significantly decreased in mutant female mice, compared to that seen with wild-type, which was also confirmed by histological analysis of the ovaries after ovulation. The analysis also showed that the numbers of follicles originally were not significantly different between WT and DKO ovaries. These results suggest that PRIP plays an important role in female reproductive system, especially in gonadotropin secretion and ovarian follicle maturation.. 9. PRIP (phospholipase C related, but catalytically inactive protein) was identified as a novel inositol 1,4,5-trisphosphate binding protein, whose domain organization is similar to that of phospholipase C-_1 but is catalytically inactive, comprizing two isoforms, PRIP-1 and PRIP-2. To get insight into the biological functions, we generated the double knockout (DKO) mice of both PRIP-1 and PRIPﾐ2, followed by the phenotypic analyses. DKO mice grew normally and became fertile. However DKO female exhibited the decreased litter occasion and litter size, indicating the dysfunction of female reproductive system. Then we examined the estrus cycle by cytological analysis of daily vaginal smears, resulting in the irregular cycle in DKO mice. We further noticed that the mutant mice had apparently smaller sized uterus by gross anatomical observation. Basal levels of serum leuteinizing hormone and follicle stimulating hormone were significantly higher in the mutant, the event of which was also confirmed by examining the secretion from the tissue culture of anterior pituitary glands. These results suggest that PRIP is involved in maternal reproduction, through the gonadotropine secretion.. 10. PRIP (phospholipase C related, but catalytically inactive protein) was identified as a novel inositol 1,4,5-trisphosphate binding protein, whose domain organization is similar to that of phospholipase C-_1 but is catalytically inactive, comprizing two isoforms, PRIP-1 and PRIP-2. To get insight into the biological functions, we generated the double knockout (DKO) mice of both PRIP-1 and PRIPﾐ2, followed by the phenotypic analyses. DKO mice grew normally and became fertile. However DKO female exhibited the decreased litter occasion and litter size, indicating the dysfunction of female reproductive system. Then we examined the estrus cycle by cytological analysis of daily vaginal smears, resulting in the irregular cycle in DKO mice. We further noticed that the mutant mice had apparently smaller sized uterus by gross anatomical observation. Basal levels of serum leuteinizing hormone and follicle stimulating hormone were significantly higher in the mutant, the event of which was also confirmed by examining the secretion from the tissue culture of anterior pituitary glands. These results suggest that PRIP is involved in maternal reproduction, through the gonadotropine secretion.. Membership in Academic Society Japanese Society for Immunology The Molecular Biology Society of Japan The Japanese biochemical Society Awards Nuclear factor-κB (NF-κB) is a transcription factor that regulates the expression of a wide variety of genes involved in inflammatory and immune responses. Recently, as a new transcriptional regulatory mechanism of NF-κB, phosphorylation of serine at 536 (534 in mice; S534) of the p65 subunit by inhibitor of κB (IκB) kinase  (IKK) has been reported. This discovery may lead to a new periodontitis treatment to resist bone resorption by host inflammatory and immune cell influences. To unveil the relation of S534 phosphorylation and periodontitis, we generated S534A knock-in (KI) mouse, in which S534 of p65 was substituted with alanine. There was no significant difference in growth rate and weight gain between wild-type (WT) and S534A KI mice. We next prepared mouse embryonic fibroblasts (MEF) from WT and S534A KI mice and stimulated the cells with tumor necrosis factor alpha (TNFa). TNFa induced phosphorylation of S354 within 30 minutes in WT MEFs, but not in S534A MEFs. TNFa-induced degradation of IκBa and phosphorylation of p38 did not change between WT and S534A MEFs. The expression level of interleukin-1beta (IL-1beta) mRNA, which is a target gene of NF-κB, after TNFa stimulation was higher in S534A MEFs than that of WT MEFs. Thus, we compared alveolar bone resorption induced by ligature in WT with S534A mice. The maxillary right second molar of each mouse was ligated using 6-0 silk string for 7 days. The degree of alveolar bone resorption was evaluated by micro-computed tomography (mCT) scanning. Ligature-induced alveolar bone resorption was enhanced in S534A mice compared to WT mice. Taken together, it could be concluded that the phosphorylation of S534 of p65 is a new regulator of bone resorption in periodontitis. The ovary is the main regulator of female reproduction and also the responsible organ for female whole life. It produces reproductive hormones and organizes homeostasis of female body and mentality with complicated endocrine system. For studying female anti-aging, it’s necessary to reveal the molecular mechanism of ovarian function. PRIP (Phospholipase C-related but catalytically inactive protein) was identified as an inositol-1,4,5-trisphosphate binding protein, comprising type1 and 2, whose gene ablation impaired female fertility manifested by decreased number of pups, longer estrous days and increased secretion of gonadotropins. Herein, we investigated the involvement of PRIP in fertility through ovary. Superovulation assay revealed the remarkable decrease of the ovulated oocyte in ovaries of PRIP knockout (KO) mice. Histological analysis of the ovaries revealed more immature follicles and fewer corpus lutea in KO ovaries. The promoted expression of luteinizing hormone receptors (LHR) at early stages of follicle development was observed in KO ovaries. The phosphorylation of extracellular signal-regulated protein kinase (ERK), a downstream target of LH signaling, was enhanced in KO granulosa cells. Serum level of estradiol was lower in DKO mice without affecting the production. These results suggest that PRIP is involved in follicle development at the early stages through the regulation of LH signaling and estradiol secretion, and also in the regulation of ovarian aging. Educational Educational Activities *Biochemical practical course*Research exposure *Lectures of molecular biology*Early exposure Other Educational Activities 2019.07. 2015.09. 2016.09. Social Professional and Outreach Activities This committee is authorized by Ministry of Health, Labour and Welfare, Japan.. Unauthorized reprint of the contents of this database is prohibited. Copyright © 2006, Kyushu University. All rights reserved.