シュワン細胞に着目したタキサン系抗がん薬誘発末梢神経障害の機序解明およびドラッグ・リポジショニングによる新規治療薬の探索

京都大学新制・課程博士博士(薬学)甲第23147号薬博第847号新制||薬||242(附属図書館)京都大学大学院薬学研究科薬学専攻(主査)教授 中山 和久, 教授 土居 雅夫, 准教授 中川 貴之学位規則第4条第1項該当Doctor of Pharmaceutical SciencesKyoto UniversityDFA

Diversifying selection and functional analysis of interleukin-4 suggests antagonism-driven evolution at receptor-binding interfaces

<p>Abstract</p> <p>Background</p> <p>Interleukin-4 (IL4) is a secreted immunoregulatory cytokine critically involved in host protection from parasitic helminths <abbrgrp><abbr bid="B1">1</abbr></abbrgrp>. Reasoning that helminths may have evolved mechanisms to antagonize IL4 to maximize their dispersal, we explored mammalian IL4 evolution.</p> <p>Results</p> <p>This analysis revealed evidence of diversifying selection at 15 residues, clustered in epitopes responsible for IL4 binding to its Type I and Type II receptors. Such a striking signature of selective pressure suggested either recurrent episodes of pathogen antagonism or ligand/receptor co-evolution. To test the latter possibility, we performed detailed functional analysis of IL4 allotypes expressed by <it>Mus musculus musculus </it>and <it>Mus musculus castaneus</it>, which happen to differ at 5 residues (including three at positively selected sites) in and adjacent to the site 1 epitope that binds the IL4Rα subunit shared by the Type I and Type II IL4 receptors. We show that this intra-species variation affects the ability of IL4 neither to bind IL4 receptor alpha (IL4Rα) nor to signal biological responses through its Type I receptor.</p> <p>Conclusions</p> <p>Our results -- reminiscent of clustered positively selected sites revealing functionally important residues at host-virus interaction interfaces -- are consistent with IL4 having evolved to avoid recurrent pathogen antagonism, while maintaining the capacity to bind and signal through its cognate receptor. This work exposes what may be a general feature of evolutionary conflicts fought by pathogen antagonists at host protein-protein interaction interfaces involved in immune signaling: the emergence of receptor-binding ligand epitopes capable of buffering amino acid variation.</p

Effect of riboflavin deficiency on development of the cerebral cortex in Slc52a3 knockout mice

Riboflavin transporter 3 (RFVT3), encoded by the SLC52A3 gene, is important for riboflavin homeostasis in the small intestine, kidney, and placenta. Our previous study demonstrated that Slc52a3 knockout (Slc52a3−/−) mice exhibited neonatal lethality and metabolic disorder due to riboflavin deficiency. Here, we investigated the influence of Slc52a3 gene disruption on brain development using Slc52a3−/− embryos. Slc52a3−/− mice at postnatal day 0 showed hypoplasia of the brain and reduced thickness of cortical layers. At embryonic day 13.5, the formation of Tuj1+ neurons and Tbr2+ intermediate neural progenitors was significantly decreased; no significant difference was observed in the total number and proliferative rate of Pax6+ radial glia. Importantly, the hypoplastic phenotype was rescued upon riboflavin supplementation. Thus, it can be concluded that RFVT3 contributes to riboflavin homeostasis in embryos and that riboflavin itself is required during embryonic development of the cerebral cortex in mice

Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

Impairment of peripheral neurons by anti-cancer agents, including taxanes and platinum derivatives, has been considered to be a major cause of chemotherapy-induced peripheral neuropathy (CIPN), however, the precise underlying mechanisms are not fully understood. Here, we examined the direct effects of anti-cancer agents on Schwann cells. Exposure of primary cultured rat Schwann cells to paclitaxel (0.01 μM), cisplatin (1 μM), or oxaliplatin (3 μM) for 48 h induced cytotoxicity and reduced myelin basic protein expression at concentrations lower than those required to induce neurotoxicity in cultured rat dorsal root ganglion (DRG) neurons. Similarly, these anti-cancer drugs disrupted myelin formation in Schwann cell/DRG neuron co-cultures without affecting nerve axons. Cisplatin and oxaliplatin, but not paclitaxel, caused mitochondrial dysfunction in cultured Schwann cells. By contrast, paclitaxel led to dedifferentiation of Schwann cells into an immature state, characterized by increased expression of p75 and galectin-3. Consistent with in vitro findings, repeated injection of paclitaxel increased expression of p75 and galectin-3 in Schwann cells within the mouse sciatic nerve. These results suggest that taxanes and platinum derivatives impair Schwan cells by inducing dedifferentiation and mitochondrial dysfunction, respectively, which may be important in the development of CIPN in conjunction with their direct impairment in peripheral neurons

Sex-inducing effects toward planarians widely present among parasitic flatworms

Summary Various parasitic flatworms infect vertebrates for sexual reproduction, often causing devastating diseases in their hosts. Consequently, flatworms are of great socioeconomic and biomedical importance. Although the cessation of parasitic flatworm sexual reproduction is a major target of anti-parasitic drug design, little is known regarding bioactive compounds controlling flatworm sexual maturation. Using the planarian Dugesia ryukyuensis, we observed that sex-inducing substances found in planarians are also widespread in parasitic flatworms, such as monogeneans and flukes (but not in tapeworms). Reverse-phase HPLC analysis revealed the sex-inducing substance(s) eluting around the tryptophan retention time in the fluke Calicophoron calicophorum, consistent with previous studies on the planarian Bipalium nobile, suggesting that the substance(s) is likely conserved among flatworms. Moreover, six of the 18 ovary-inducing substances identified via transcriptome and metabolome analyses are involved in purine metabolism. Our findings provide a basis for understanding and modifying the life cycles of various parasitic flatworms.journal articl

Polymorphisms and Body Mass Index Across Life Course

Background: Obesity is a reported risk factor for various health problems. Genome-wide association studies (GWASs) have identified numerous independent loci associated with body mass index (BMI). However, most of these have been focused on Europeans, and little evidence is available on the genetic effects across the life course of other ethnicities. Methods: We conducted a cross-sectional study to examine the associations of 282 GWAS-identified single nucleotide polymorphisms with three BMI-related traits, current BMI, BMI at 20 years old (BMI at 20), and change in BMI (BMI change), among 11,586 Japanese individuals enrolled in the Japan Multi-Institutional Collaborative Cohort study. Associations were examined using multivariable linear regression models. Results: We found a significant association (P < 0.05/282 = 1.77 × 10−4) between BMI and 11 polymorphisms in or near FTO, BDNF, TMEM18, HS6ST3, and BORCS7. The trend was similar between current BMI and BMI change, but differed from that of the BMI at 20. Among the significant variants, those on FTO were associated with all BMI traits, whereas those on TMEM18 and HS6SR3 were only associated with BMI at 20. The association of FTO loci with BMI remained, even after additional adjustment for dietary energy intake. Conclusions: Previously reported BMI-associated loci discovered in Europeans were also identified in the Japanese population. Additionally, our results suggest that the effects of each loci on BMI may vary across the life course and that this variation may be caused by the differential effects of individual genes on BMI via different pathways

Improvement of peripheral vascular impairment by a phosphodiesterase type 5 inhibitor tadalafil prevents oxaliplatin-induced peripheral neuropathy in mice

Oxaliplatin, a platinum-based chemotherapeutic drug, frequently induces peripheral neuropathy. Accumulating evidences suggest a possible relationship between peripheral vascular impairment and peripheral neuropathy. In this study, we investigated the effects of vasodilators on cumulative peripheral neuropathy induced by repeated injections of oxaliplatin (10 mg/kg) once a week for 8 weeks in mice. Single injections of vasodilators, including a phosphodiesterase type 5 inhibitor tadalafil acutely alleviated oxaliplatin-induced cold hypersensitivity, while tadalafil had no effect on the mechanical hypersensitivity. By contrast, long-term administration of tadalafil (0.1% in chow diets) during the oxaliplatin injection period reduced the oxaliplatin-induced decreases in skin temperature and blood flow without affecting platinum concentrations in blood, sciatic nerves, and dorsal root ganglion. The long-term administration significantly suppressed cold, mechanical, and electrical current hypersensitivities as well as thermal hypoesthesia. Furthermore, it prevented the decreases in sensory nerve conductance velocity and the number of endoneurial microvessels, and axon degeneration in the sciatic nerves. In vitro studies confirmed that tadalafil does not interfere with the cytotoxicity of oxaliplatin against human cancer cell lines. Altogether, these results suggest that improvement of peripheral vascular impairment by tadalafil could alleviate and prevent oxaliplatin-induced peripheral neuropathy

細菌性スーパー抗原黄色ブドウ球菌腸管毒素A(SEA)に応答するいくつかのマウスT細胞集団はSEA刺激に対する応答性が異なる

我々はスーパー抗原,黄色ブドウ球菌腸管毒素A(SEA)2回投与の影響をC57BL/6,β^+ミクログロブリンノックアウトおよびCD4ノックアウトマウスを用いて検討した.1回目のSEA投与では,SEA応答性T細胞4画分(Vβ3^+D4^+,Vβ3^+D8^+,Vβ11^+CD4^+,Vβ11^+CD8^+)は共に投与2日目をピークに増幅し,その後対照のレベルまで減少する.しかし,1回目の投与後,2日目にSEAを再投与するとSEA応答性T細胞画分はそれぞれ異なった反応を示した.増幅したVβ3^+CD4^+細胞はさらに投与後2日目まで増幅した.増幅したVβ3^+CD8^+およびVβ11^+CD4^+T細胞は再投与後2日目まで増幅した状態を維持した.一方,増幅したVβ11^+CD8^+T細胞は再投与後,すぐに減少した.SEA2回目投与後,3時間目の各細胞集団のIL-2レセプターα鎖の発現量はVβ3^+CD4^+およびVβ11^+CD4^+T細胞で90%以上,Vβ3^+CD8^+T細胞では約80%,Vβ11^+CD8^+T細胞では約70%であり,これらの細胞集団の大多数はSEA認識能力を保持していた.以上の結果より,Vβ11^+CD8^+T細胞のアナジー感受注は著しく高く,Vβ3^+CD4^+T細胞は低く,Vβ3^+CD8^+およびVβ11^+CD4^+T細胞はその中間であろうと推測した.We examined the immunologic behaviors of a superantigen Staphylococcal enterotoxin A (SEA)-reactive four distinct T cell populations (Vβ3^+CD4^+, Vβ3^+CD8^+, Vβ11^+CD4^+ and Vβ11^+CD8^+ T cells) in mice injected with SEA twice in a 2-day interval. The four T cell populations increased equally at 2 days after the first injection and thereafter decreased equally to the control level. However, these four T cell populations expanded by the first SEA injection exhibited different behaviors upon the second SEA injection, depending on the T cell receptor Vβ elements expressed and the T cell subsets. Vβ3^+CD4^+ T cells expanded exhibited further expansion, and Vβ11^+CD8^+ T cells expanded decreased rapidly. Vβ3^+CD8^+ and Vβ11^+CD4^+ T cells expanded were sustained in similar levels for 2 days after the second injection. Peak of serum IL-2 activity was seen in a higher level and at earlier hours after the second SEA injection than the first injection. Levels of IL-2 receptor a chain expression were more than 90% in Vβ3^+CD4^+ T cells and Vβ11^+CD4^+ T cells, about 80% in Vβ3^+CD8^+ T cells, and about 70% in Vβ11^+CD8^+ T cells, suggesting that large parts of these T cell populations can recognized SEA. These results suggest that Vβ3^+CD4^+ T cells are low and Vβ11^+CD8^+ T cells are high in the susceptibility to anergic induction with SEA. Vβ3^+CD8^+ and Vβ11^+CD4^+ T cells may be sustained at intermediate level