• BMB Reports
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• 제40권4호
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• pp.475-485
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• 2007

Methamphetamine is an illicit drug that is often abused and can cause neuropsychiatric and neurotoxic damage. Repeated administration of psychostimulants such as methamphetamine induces a behavioral sensitization. According to a previous study, Bax was involved in neurotoxicity by methamphetamine, but the function of Bax in rewarding effect has not yet been elucidated. Therefore, we have studied the function of Bax in a rewarding effect model. In the present study, we treated chronic methamphetamine exposure in a Bax-deficient mouse model and examined behavioral change using a conditioned place preference (CPP) test. The CPP score in Bax knockout mice was decreased compared to that of wild-type mice. Therefore, we screened for Bax-related genes that are involved in rewarding effect using microarray technology. In order to confirm microarray data, we applied the RT-PCR method to observe relative changes of Bcl2, a pro-apoptotic family gene. As a result, using our experiment microarray, we selected genes that were associated with Bax in microarray data, and eventually selected the Tgfbr2 gene. Expression of the Tgfbr2 gene was decreased by methamphetamine in Bax knockout mice, and the gene was overexpressed in Bax wild-type mice. Additionally, we confirmed that Creb, FosB, and c-Fos were related to rewarding effect and Bax using immunohistochemistry.

• IMMUNE NETWORK
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• 제4권2호
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• pp.81-87
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• 2004

Background: In the thymus, developing thymocytes continually interact with thymic epithelial cell components. Self MHC restriction of mature T cells are imposed in the thymus through interaction of immature double positive thymocytes and thymic cortical epithelial cells. The site of negative selection, however, is a matter of debate. Through systemic injection of anti-TCR antibody or antigenic peptides, investigators suggested that most of the negative selection occurs in the thymic cortex. But the requirements for negative selection, i.e cellular counterparts and costimulatory molecules are more available in the medulla or cortico-medullary junction rather than in the thymic cortex. Methods: The direct and indirect pathways of thymocyte death after systemic anti-TCR antibody injection were separated through several experimental systems. B6 mice were either adrenalectomized or sham-adrenalectomized to evaluate the role of endogenous glucocorticoids from adrenal gland. Role of TNF were evaluated through using TNF receptor double knockout mice. Results: We found that without indirectly acting mediators such as $TNF-\alpha$ or corticosteroid, double positive thymocyte death were minimal by systemic injection of anti-TCR antibody in TNF receptor double knockout neonatal mice. Also by analyzing neonatal wild-type mice with adoptively transferred mature T cells, only peripheral activation of mature T cells could induce extensive double positive thymocyte death. Conclusion: Thus, systemically injected anti-TCR antibody mediated thymocyte death are mostly induced through indirect pathway.

• Biomolecules & Therapeutics
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• 제32권1호
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• pp.115-122
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• 2024

Heat shock protein (HSP) 90 is expressed in most living organisms, and several client proteins of HSP90 are necessary for cancer cell survival and growth. Previously, we found that HSP90 was cleaved by histone deacetylase (HDAC) inhibitors and proteasome inhibitors, and the cleavage of HSP90 contributes to their cytotoxicity in K562 leukemia cells. In this study, we first established mouse xenograft models with K562 cells expressing the wild-type or cleavage-resistant mutant HSP90β and found that the suppression of tumor growth by the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) was interrupted by the mutation inhibiting the HSP90 cleavage in vivo. Next, we investigated the possible function of thioredoxin interacting protein (TXNIP) in the HSP90 cleavage induced by SAHA. TXNIP is a negative regulator for thioredoxin, an antioxidant protein. SAHA transcriptionally induced the expression of TXNIP in K562 cells. HSP90 cleavage was induced by SAHA also in the thymocytes of normal mice and suppressed by an anti-oxidant and pan-caspase inhibitor. When the thymocytes from the TXNIP knockout mice and their wild-type littermate control mice were treated with SAHA, the HSP90 cleavage was detected in the thymocytes of the littermate controls but suppressed in those of the TXNIP knockout mice suggesting the requirement of TXNIP for HSP90 cleavage. We additionally found that HSP90 cleavage was induced by actinomycin D, β-mercaptoethanol, and p38 MAPK inhibitor PD169316 suggesting its prevalence. Taken together, we suggest that HSP90 cleavage occurs also in vivo and contributes to the anti-cancer activity of various drugs in a TXNIP-dependent manner.

• BMB Reports
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• 제49권12호
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• pp.651-652
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• 2016

EphA2 has been implicated in amplifying ErbB2 tumorigenic signaling. One protein that interacts with EphA2 is the Anks1a PTB adaptor. However, the precise role of Anks1a in EphA2-mediated tumorigenesis is unclear. We demonstrated that Anks1a localizes to the ER upon phosphorylation and that the Ankyrin repeats and PTB of Anks1a bind to EphA2 and Sec23, respectively. Thus, Anks1a facilitates the selective packaging of EphA2 into COPII vesicles. Additionally, Anks1a knockout mice, a phenocopy of EphA2 knockout mice, exhibited markedly reduced ErbB2-induced breast tumorigenesis. Strikingly, ErbB2 did not localize to the cell surface following Anks1a knockdown in primary mammary tumor cells over-expressing ErbB2. Importantly, EphA2 was critical for stabilizing ErbB2 through complex formation, but its interaction with Anks1a also facilitated ErbB2 loading into COPII carriers. These findings suggest a novel role for Anks1a in the molecular pathogenesis of breast tumors and possibly other human diseases.

• Interdisciplinary Bio Central
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• 제2권4호
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• pp.15.1-15.5
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• 2010

RIKEN BioResource Center (BRC) has collected, preserved, conducted quality control of, and distributed mouse resources since 2002 as the core facility of the National BioResource Project by the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan. Our mouse resources include over 5,000 strains such as humanized disease models, fluorescent reporters, and knockout mice. We have developed novel mouse strains such as tissue-specific Cre-drivers and optogenetic strains that are in high demand by the research community. We have removed all our specified pathogens from the deposited mice and used our quality control tests to examine their genetic modifications and backgrounds. RIKEN BRC is a founding member of the Federation of International Mouse Resources and the Asian Mouse Mutagenesis and Resource Association, and provides mouse resources to the one-stop International Mouse Strain Resource database. RIKEN BRC also participates in the International Gene Trap Consortium, having registered 713 gene-trap clones and their sequences in a public library, and is an advisory member of the CREATE (Coordination of resources for conditional expression of mutated mouse alleles) consortium which represents major European and international mouse database holders for the integration and dissemination of Cre-driver strains. RIKEN BRC provides training courses in the use of advanced technologies for the quality control and cryopreservation of mouse strains to promote the effective use of mouse resources worldwide.

• 한국수정란이식학회지
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• 제29권2호
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• pp.195-200
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• 2014

Endothelin 2 (EDN2) induces follicular rupture by constricting periovulatory follicles. In this study, it was investigated the mechanisms of EDN2 action on follicular rupture with respect of receptor using the conditionally granulosa cell specific EDN2 receptor type A (ETa) KO mice (gcETaKO; $ETa^{flox/-}{\cdot}Amhr2^{Cre}$). It was generated the gcETaKO mice by breeding with $ETa^{flox/-}$ mice after mono-alleic ETa knockout by $ZP3^{Cre}$ and $Amhr2^{Cre}$ mice. Fertility, ovulation and maturation rates of ovulated oocytes after super ovulation were investigated in the gcETaKO mice compared with wild-type mice ($ETa^{flox/flox}$ and $ETa^{flox/-}$) as a control group. In the gcETaKO mice, normal fertility after breeding with male mice was shown compared with wild-type mice. And, there was no significant differences in ovulation rates after super ovulation, however its maturation rates was lower than that of wild type mice. These findings show that EDN2 in follicular rupture for ovulation is related with an other ETa not in granulosa cells. Further studies are needed to investigate how EDN2 is acted in ovarian follicular rupture for ovulation.

• 생명과학회지
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• 제25권3호
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• pp.336-344
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• 2015

비타민 C는 신진대사에 연관되어 있으며 특히 항산화 기능을 가지고 있다. 본 연구에서는 생체에서 비타민 C를 합성할 수 없는 SMP 30 녹아웃 마우스에 간 절제술을 시행하여 간 재생에서 비타민 C의 역할을 관찰하였다. 간 절제술은 마우스 중간엽 및 좌엽을 제거한 부분절제술을 수행하였다. 마우스는 간 절제술 후 비타민 C를 투여한 군(KV)와 비타민 C를 투여하지 않는 군(KO)로 나누어서 비타민 C의 효과를 관찰하였다. 결과 비타민 C를 투여한 KV 마우스의 간 회복이 투여하지 않는 KO 마우스에 비해 촉진되었다. KV 마우스의 혈액에서 관찰된 간소상 지표인 아스파르타산 아미노전달효소 및 간 손상 정도가 KO 마우스에 비해 낮게 관찰되었다. KV 마우스에서는 HGF와 c-Met에 의해서 TGF-베타 수용체 신호전달계가 활성화되고 세포주기 조절인자인 cyclin D1과 PCNA의 발현이 빠르게 증가되었다. 반면 KO 마우스에서는 활성화 되지 않았다. 또한 ERK와 GSK-3β 단백질의 활성화가 관찰되었으며 세포분열 간세포들의 유의적인 증가가 관찰되었다. 그리고 KV 마우스에서는 혈중 알부민의 농도가 높은 것으로 확인되었다. 따라서 본 실험결과는 SMP 30 결핍 마우스에서 비타민 C 투여는 간 재생시스템의 활성화와 이에 따른 빠른 회복을 초래한다.

• 생명과학회지
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• 제21권8호
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• pp.1163-1167
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• 2011

과다한 음주는 알츠하이머 및 파킨슨 질병과 같은 각종 만성 퇴행성 뇌질환의 대표적인 원인 중 하나로 알려져 있다. 체내에 유입된 에탄올은 알코올 탈수소효소(alcohol dehydrogenase, ADH)에 의해 아세트알데히드로 대사된 후 다시 알데히드탈수소효소 2(aldehyde dehydrogenase 2, ALDH2)에 의해 아세트산으로 대사되어 배출된다. 에탄올의 대사과정 중에는 다량의 free radical이 생성되어 체내에서 산화적 스트레스를 유발하는 것으로 알려져 있고, 아세트알데히드는 활성산소를 생산하는 독성물질로 잘 알려져 있다. 본 연구에서는 8주간 에탄올에 노출된 Aldh2 knockout 마우스를 사용하여 ALDH2 효소 활성이 뇌 조직과 소변의 지질과산화에 미치는 영향에 대하여 살펴보았으며, 지질과산화 정도를 측정하기 위해 HPLC를 통한 TBARS 정도를 측정하였다. 연구결과, 마우스에서 만성 에탄올 섭취는 뇌 조직 TBARS 생성에 영향을 주지 않는 것으로 나타났으나, 소변 TBARS는 Aldh2 (-/-) 마우스에서 에탄올을 투여함에 따라 유의한 증가를 보였다(p<0.05). 본 연구 결과로부터 8주간 에탄올을 경구 투여한 마우스에서 ALDH2의 활성은 체내의 전반적인 활성산소 생성에는 중요하게 관여하는 것으로 보이지만 뇌조직에서의 활성산소 생성에는 영향을 주지 않는 것으로 보이며, 이는 에탄올 노출과 이에 따른 활성산소가 다양한 만성 뇌질환을 유발한다는 기존의 가설에서 ALDH2의 활성이 중요하게 관여하지 않을 가능성을 시사한다.

• Journal of Oral Medicine and Pain
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• 제40권3호
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• pp.110-114
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• 2015

Purpose: Growth and differentiation factor (GDF)-11 is a transforming growth factor-${\beta}$ family member that plays important regulatory roles in development of multiple tissues which include axial skeletal patterning, palatal closure, and tooth formation. Proteins that have been identified as GDF-11 inhibitors include GDF-associated serum protein (GASP)-1 and GASP-2. Recently, we found that mice genetically engineered to lack both Gasp1 and Gdf11 have an increased frequency of cleft palate. The goal of this study was to investigate the roles of GDF-11 and its inhibitors, GASP-1 and GASP-2, during dental and craniofacial development and growth. Methods: Mouse genetic studies were used in this study. Homozygous knockout mice for Gasp1 ($Gasp1^{-/-}$) and Gasp2 ($Gasp2^{-/-}$) were viable and fertile, but Gdf11 homozygous knockout ($Gdf11^{-/-}$) mice died within 24 hours after birth. The effect of either Gasp1 or Gasp2 deletion in $Gdf11^{-/-}$ mice during embryogenesis was evaluated in $Gasp1^{-/-}$;$Gdf11^{-/-}$ and $Gasp2^{-/-}$;$Gdf11^{-/-}$ mouse embryos at 18.5 days post-coitum (E18.5). For the analysis of adult tissues, we used $Gasp1^{-/-}$;$Gdf11^{+/-}$ and $Gasp2^{-/-}$;$Gdf11^{+/-}$ mice to evaluate the potential haploinsufficiency of Gdf11 in $Gasp1^{-/-}$ and $Gasp2^{-/-}$ mice. Results: Although Gasp2 expression decreased after E10.5, Gasp1 expression was readily detected in various ectodermal tissues at E17.5, including hair follicles, epithelium in nasal cavity, retina, and developing tooth buds. Interestingly, $Gasp1^{-/-}$;$Gdf11^{-/-}$ mice had abnormal formation of lower incisors: tooth buds for lower incisors were under-developed or missing. Although $Gdf11^{+/-}$ mice were viable and had mild transformations of the axial skeleton, no specific defects in the craniofacial development have been observed in $Gdf11^{+/-}$ mice. However, loss of Gasp1 in $Gdf11^{+/-}$ mice occasionally resulted in small and abnormally shaped auricles. Conclusions: These findings suggest that both GASP-1 and GDF-11 play important roles in dental and craniofacial development both during embryogenesis and in adult tissues.

• Journal of Microbiology and Biotechnology
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• 제26권8호
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• pp.1446-1451
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• 2016

Clostridium difficile toxin A causes acute gut inflammation in animals and humans. It is known to downregulate the tight junctions between colonic epithelial cells, allowing luminal contents to access body tissues and trigger acute immune responses. However, it is not yet known whether this loss of the barrier function is a critical factor in the progression of toxin A-induced pseudomembranous colitis. We previously showed that NADH:quinone oxidoreductase 1 (NQO1) KO (knockout) mice spontaneously display weak gut inflammation and a marked loss of colonic epithelial tight junctions. Moreover, NQO1 KO mice exhibited highly increased inflammatory responses compared with NQO1 WT (wild-type) control mice when subjected to DSS-induced experimental colitis. Here, we tested whether toxin A could also trigger more severe inflammatory responses in NQO1 KO mice compared with NQO1 WT mice. Indeed, our results show that C. difficile toxin A-mediated enteritis is significantly enhanced in NQO1 KO mice compared with NQO1 WT mice. The levels of fluid secretion, villus disruption, and epithelial cell apoptosis were also higher in toxin A-treated NQO1 KO mice compared with WT mice. The previous and present results collectively show that NQO1 is involved in the formation of tight junctions in the small intestine, and that defects in NQO1 enhance C. difficile toxin A-induced acute inflammatory responses, presumably via the loss of epithelial cell tight junctions.