Arnica montana L. extract containing 6-O-methacryloylhelenalin and 6-O-isobutyrylhelenalin accelerates growth and differentiation of human subcutaneous preadipocytes and leads volumizing of skin.

OBJECTIVE: An important factor in the aging of the face is a reduction in the volume of adipose tissue. This reduction in adipose tissue contributes to decreased skin elasticity, which is also part of the aging process. Overall, these lead to wrinkle formation. Fat injection is a common means of addressing this issue and is used to reduce the effects of aging on the face and to increase the fullness of the lips and breasts. However, fat injection is an invasive surgical procedure. This study aimed to discover novel cosmetic ingredients that increase the volume of subcutaneous (pre)adipocytes to create the appearance of more youthful skin. METHODS: We focused on the number of subcutaneous preadipocytes and the accumulation of lipid droplets. To discover natural ingredients that increase both of these, extracts of 380 natural products were prepared and screened for their effects on both growth and differentiation (i.e., lipid droplet accumulation) of human subcutaneous preadipocytes. One extract was found to have the desired effects, and this was further studied to determine the active compounds. We then evaluated its efficacy in a human clinical study. RESULTS: We found that Arnica montana L. flower extract (AFE) accelerates both the growth and the differentiation of human subcutaneous preadipocytes. AFE was found to significantly increase the volume of adipocyte spheroids. The active compounds 6-O-methacryloylhelenalin and 6-O-isobutyrylhelenalin were found to be responsible for the effects of AFE on preadipocytes. In a human clinical study, gels containing 1% AFE successfully enhanced the volume of the lips and face with reduction of wrinkles with no adverse reactions. CONCLUSION: This is the first report to demonstrate that AFE and the included compounds, 6-O-methacryloylhelenalin and 6-O-isobutyrylhelenalin, act on preadipocytes. AFE would be ideal for use in products that plump the face to reduce wrinkles and create a more youthful appearance.

OBJECTIF: Un facteur important du vieillissement du visage réside dans la réduction du volume du tissu adipeux. Cette réduction du tissu adipeux contribue à une diminution de l'élasticité de la peau qui fait également partie du processus de vieillissement. Globalement, ces facteurs induisent la formation des rides. L'injection de graisse est un moyen courant pour remédier à ce problème et sert à réduire les effets du vieillissement sur le visage et à augmenter la plénitude des lèvres et des seins. Cependant, l'injection de graisse est une intervention chirurgicale invasive. Cette étude visait à découvrir des ingrédients cosmétiques innovants qui augmentent le volume des (pré)adipocytes sous-cutanés pour créer l'apparence d'une peau plus jeune. MÉTHODES: Nous avons mis l'accent sur le nombre de préadipocytes sous-cutanés et sur l'accumulation de gouttelettes lipidiques. Pour découvrir des ingrédients naturels qui augmentent ces deux facteurs, des extraits de 380 produits naturels ont été préparés et analysés en vue de la détermination de leurs effets sur la croissance et la différenciation (c'est-à-dire l'accumulation de gouttelettes lipidiques) des préadipocytes humains sous-cutanés. Un extrait s'est avéré avoir les effets escomptés et il a fait l'objet d'études approfondies visant à déterminer les composés actifs. Nous avons ensuite évalué son efficacité dans une étude clinique chez l'homme. RÉSULTATS: Nous avons découvert que l'extrait de fleur de l'Arnica montana L. (AFE) accélère à la fois la croissance et la différenciation des préadipocytes humains sous-cutanés. L'AFE s'est avéré augmenter considérablement le volume des sphéroïdes des adipocytes. Les composés actifs 6-Ométhacryloyl-hélénaline et 6-O-isobutyryl-hélénaline se sont avérés responsables des effets de l'AFE sur les préadipocytes. Dans une étude clinique chez l'homme, des gels contenant 1 % d'AFE ont permis d'améliorer le volume des lèvres et du visage avec une réduction des rides sans effets indésirables. CONCLUSION: Il s'agit du premier rapport démontrant que l'AFE et les composés inclus, 6-O-méthacryloyl-hélénaline et 6-O-isobutyryl-hélénaline, agissent sur les préadipocytes. L'AFE serait idéal pour les produits qui repulpent le visage afin de réduire les rides et de donner un aspect rajeuni.

Juniper berry extract containing Anthricin and Yatein suppresses lipofuscin accumulation in human epidermal keratinocytes through proteasome activation, increases brightness and decreases spots in human skin.

OBJECTIVE: Skin brightness and spot have a significant impact on youthful and beautiful appearance. One important factor influencing skin brightness is the amount of internal reflected light from the skin. Observers recognize the total surface-reflected light and internal reflected light as skin brightness. The more internal reflected light from the skin, the more attractive and brighter the skin appears. This study aims to identify a new natural cosmetic ingredient that increases the skin's internal reflected light, decreases spot and provides a youthful and beautiful skin appearance. METHODS: Lipofuscin in epidermal keratinocytes, the aggregating complex of denatured proteins and peroxidized lipids, is one factor that decreases skin brightness and causes of spot. Aggregates block light transmission, and peroxidized lipids lead to skin yellowness, dullness and age spot. Lipofuscin is known to accumulate intracellularly with ageing. Rapid removal of intracellular denatured proteins prevents lipofuscin formation and accumulation in cells. We focused a proteasome system that efficiently removes intracellular denatured proteins. To identify natural ingredients that increase proteasome activity, we screened 380 extracts derived from natural products. The extract with the desired activity was fractionated and purified to identify active compounds that lead to proteasome activation. Finally, the efficacy of the proteasome-activating extract was evaluated in a human clinical study. RESULTS: We discovered that Juniperus communis fruits (Juniper berry) extract (JBE) increases proteasome activity and suppresses lipofuscin accumulation in human epidermal keratinocytes. We found Anthricin and Yatein, which belong to the lignan family, to be major active compounds responsible for the proteasome-activating effect of JBE. In a human clinical study, an emulsion containing 1% JBE was applied to half of the face twice daily for 4 weeks, resulting in increased internal reflected light, brightness improvement (L-value) and reduction in yellowness (b-value) and spot in the cheek area. CONCLUSION: This is the first report demonstrating that JBE containing Anthricin and Yatein decreases lipofuscin accumulation in human epidermal keratinocytes through proteasome activation, increases brightness and decreases surface spots in human skin. JBE would be an ideal natural cosmetic ingredient for creating a more youthful and beautiful skin appearance with greater brightness and less spot.

OBJECTIF: La luminosité et les taches de peau ont un impact significatif sur la jeunesse et la beauté de l'apparence. L'un des facteurs importants influençant la luminosité de la peau est la quantité de lumière interne réfléchie par la peau. Pour les observateurs, la luminosité de la peau correspond à la somme de la lumière réfléchie par la surface et de la lumière réfléchie par l'intérieur de la peau. Plus la quantité de lumière interne réfléchie par la peau est importante, plus la peau semble attrayante et lumineuse. Cette étude vise à identifier un nouvel ingrédient cosmétique naturel qui augmente la lumière interne réfléchie par la peau, diminue les taches et donne à la peau une apparence jeune et belle. MÉTHODES: La lipofuscine dans les kératinocytes de l'épiderme, le complexe agrégé de protéines dénaturées et de lipides peroxydés, est un facteur qui diminue l'éclat de la peau et qui est à l'origine des taches. Les agrégats bloquent la transmission de la lumière et les lipides peroxydés entraînent une coloration jaune de la peau, un aspect terne et des taches de vieillesse. On sait que la lipofuscine s'accumule au niveau intracellulaire avec le vieillissement. L'élimination rapide des protéines dénaturées intracellulaires empêche la formation et l'accumulation de lipofuscine dans les cellules. Nous avons mis l'accent sur un système de protéasome qui élimine efficacement les protéines dénaturées intracellulaires. Pour identifier les ingrédients naturels qui augmentent l'activité du protéasome, nous avons passé au crible 380 extraits dérivés de produits naturels. L'extrait présentant l'activité souhaitée a été fractionné et purifié afin d'identifier les composés actifs qui conduisent à l'activation du protéasome. Enfin, l'efficacité de l'extrait activant le protéasome a été évaluée dans une étude clinique humaine. RÉSULTATS: Nous avons découvert que l'extrait de Juniperus communis fruits (baie de genièvre) augmente l'activité du protéasome et supprime l'accumulation de lipofuscine dans les kératinocytes épidermiques humains. Nous avons découvert que l'anthricine et la yateine, qui appartiennent à la famille des lignanes, sont les principaux composés actifs responsables de l'effet activateur du protéasome de l'extrait de baies de genévrier. Dans une étude clinique humaine, une émulsion contenant 1 % de JBE a été appliquée sur la moitié du visage deux fois par jour pendant 4 semaines, ce qui a entraîné une augmentation de la lumière interne réfléchie, une amélioration de la luminosité (valeur L) et une réduction de la jaunisse (valeur b) et des taches dans la zone des joues. CONCLUSION: Il s'agit du premier rapport démontrant que l'EBJ contenant de l'anthricine et de la yateine diminue l'accumulation de lipofuscine dans les kératinocytes épidermiques humains par l'activation du protéasome, augmente la luminosité et diminue les taches superficielles de la peau humaine. Le JBE serait un ingrédient cosmétique naturel idéal pour créer une peau plus jeune et plus belle, plus lumineuse et moins tachetée.

Effects of watercress extract fraction on R-spondin 1-mediated growth of human hair.

OBJECTIVE: Hair loss and greying affect men and women of all ages, often causing psychosocial difficulties. Dickkopf-1 (DKK1), a major hair loss factor secreted from dermal papilla (DP) cells in response to the secretion of dihydrotestosterone (DHT), has been reported to induce and accelerate androgenetic alopecia (AGA). In addition, DKK1 acts as a potent suppressor of melanogenesis and is closely related to hair colour. R-spondin 1 (RSPO1) is a secretory agonist of Wnt signalling known to antagonize the effects of DKK1, including DKK1-mediated hair follicle suppression. In this study, we investigated the effect of watercress extract (WCE) on the secretion of RSPO1 and DKK1 from DP cells as well as its anti-hair loss effect in human hair follicles and patients. METHODS: The in vitro secretion of RSPO1 and DKK1 was measured by ELISA. Human hair follicles were collected from the scalp of a female donor and used for ex vivo organ culture to investigate the effects of WCE on human hair loss. Finally, a 6-month human clinical trial was conducted to examine the effect of WCE-containing lotion on hair growth in a male panel. RESULTS: WCE significantly upregulated RSPO1 secretion and suppressed DKK1 secretion in a dose-dependent manner, even in the presence of DHT. WCE-treated hair follicles elongated 1.6-fold compared with the control, and the level of RSPO1 production in DP as well as RSPO1 bound to the outer root sheath (ORS) increased. In the clinical trial, the hair lotion containing 2% WCE increased hair thickness and density to improve against hair loss symptoms. CONCLUSION: WCE exhibited a strong anti-androgenic effect through its ability to suppress DKK1 secretion and antagonize DKK1 via RSPO1. These findings highlighted the potential use of WCE for the treatment of hair loss.

OBJECTIF: La perte de cheveux et le grisonnement touchent des hommes et des femmes de tous âges, ce qui entraîne souvent des difficultés psychosociales. Selon des rapports, Dickkopf-1 (DKK1), un facteur de perte de cheveux majeur sécrété par les cellules de la papille dermique (PD) en réponse à la sécrétion de dihydrotestostérone (DHT), induit et accélère l'alopécie androgénétique (AAG). En outre, DKK1 agit comme un puissant suppresseur de la mélanogenèse et est étroitement lié à la couleur des cheveux. La protéine R-spondin 1 (RSPO1) est un agoniste sécrétoire de la voie de signalisation Wnt connue pour antagoniser les effets de DKK1, notamment la suppression des follicules pileux médiée par DKK1. Dans cette étude, nous avons étudié l'effet de l'extrait de cresson sur la sécrétion de RSPO1 et de DKK1 à partir des cellules de la PD, ainsi que son effet anti-perte de cheveux sur les follicules pileux humains et chez les patients. MÉTHODES: La sécrétion in vitro de RSPO1 et de DKK1 a été mesurée à l'aide de la méthode ELISA. Des follicules pileux humains ont été prélevés sur le cuir chevelu d'une femme et utilisés pour une culture d'organes ex vivo afin d'étudier les effets de l'extrait de cresson sur la perte de cheveux humains. Enfin, un essai clinique de 6 mois chez l'être humain a été mené pour examiner l'effet d'une lotion contenant de l'extrait de cresson sur la croissance des cheveux au sein d'un panel d'hommes. RÉSULTATS: L'extrait de cresson a significativement régulé à la hausse la sécrétion de RSPO1 et a supprimé la sécrétion de DKK1 de manière dose-dépendante, même en présence de DHT. Les follicules pileux traités avec de l'extrait de cresson ont été multipliés par 1,6 par rapport au groupe témoin, et le niveau de production de RSPO1 dans la PD ainsi que le taux de RSPO1 lié à la gaine externe de la racine ont augmenté. Dans l'essai clinique, la lotion pour cheveux contenant 2 % d'extrait de cresson a augmenté l'épaisseur et la densité des cheveux, améliorant ainsi les symptômes de perte de cheveux. CONCLUSION: La capacité de l'extrait de cresson à supprimer la sécrétion de DKK1 et à antagoniser DKK1 via la protéine RSPO1 lui a conféré un effet anti-androgénique puissant. Ces résultats ont mis en évidence le potentiel de l'extrait de cresson pour le traitement de la perte de cheveux.

Extracellular Environment-Controlled Angiogenesis, and Potential Application for Peripheral Nerve Regeneration.

Endothelial cells acquire different phenotypes to establish functional vascular networks. Vascular endothelial growth factor (VEGF) signaling induces endothelial proliferation, migration, and survival to regulate vascular development, which leads to the construction of a vascular plexuses with a regular morphology. The spatiotemporal localization of angiogenic factors and the extracellular matrix play fundamental roles in ensuring the proper regulation of angiogenesis. This review article highlights how and what kinds of extracellular environmental molecules regulate angiogenesis. Close interactions between the vascular and neural systems involve shared molecular mechanisms to coordinate developmental and regenerative processes. This review article focuses on current knowledge about the roles of angiogenesis in peripheral nerve regeneration and the latest therapeutic strategies for the treatment of peripheral nerve injury.

Exogenous Application of Proteoglycan to the Cell Surface Microenvironment Facilitates to Chondrogenic Differentiation and Maintenance.

Osteoarthritis (OA), a disease that greatly impacts quality of life, has increasing worldwide prevalence as the population ages. However, its pathogenic mechanisms have not been fully elucidated and current therapeutic treatment strategies are inadequate. In recent years, abnormal endochondral ossification in articular cartilage has received attention as a pathophysiological mechanism in OA. Cartilage is composed of abundant extracellular matrix components, which are involved in tissue maintenance and regeneration, but how these factors affect endochondral ossification is not clear. Here, we show that the application of aggrecan-type proteoglycan from salmon nasal cartilage (sPG) exhibited marked proliferative capacity through receptor tyrosine kinases in chondroprogenitor cells, and also exhibited differentiation and three-dimensional structure formation via phosphorylation of Insulin-like Growth Factor-1 Receptor and Growth Differentiation Factor 5 expression. Furthermore, sPG inhibited calcification via expression of Runx2 and Col10 (factors related to induction of calcification), while increasing Mgp, a mineralization inhibitory factor. As a result of analyzing the localization of sPG applied to the cells, it was localized on the surface of the cell membrane. In this study, we found that sPG, as a biomaterial, could regulate cell proliferation, differentiation and calcification inhibition by acting on the cell surface microenvironment. Therefore, sPG may be the foundation for a novel therapeutic approach for cartilage maintenance and for improved symptoms in OA.

Cynaropicrin from Cynara scolymus L. suppresses photoaging of skin by inhibiting the transcription activity of nuclear factor-kappa B.

Aging of skin is characterized by skin wrinkling, laxity, and pigmentation induced by several environmental stress factors. Histological changes during the photoaging of skin include hyperproliferation of keratinocytes and melanocytes causing skin wrinkles and pigmentation. Nuclear factor kappa B (NF-κB) is one of the representative transcription factors active in conjunction with inflammation. NF-κB is activated by stimulation such as ultraviolet rays and inflammatory cytokines and induces the expression of various genes such as those of basic fibroblast growth factor (bFGF) and matrix metalloprotease-1 (MMP-1). We screened several plant extracts for their possible inhibitory effect on the transcriptional activity of NF-κB. One of them, an extract from Cynara scolymus L., showed a greatest effect on the suppression of NF-κB transactivation. As a result, we found that cynaropicrin, which is a sesquiterpene lactone, inhibited the NF-κB-mediated transactivation of bFGF and MMP-1. Furthermore, it was confirmed that in an in vivo mouse model cynaropicrin prevented skin photoaging processes leading to the hyperproliferation of keratinocytes and melanocytes. These findings taken together indicate that cynaropicrin is an effective antiphotoaging agent that acts by inhibiting NF-κB-mediated transactivation.

Generation of KS-133 as a Novel Bicyclic Peptide with a Potent and Selective VIPR2 Antagonist Activity that Counteracts Cognitive Decline in a Mouse Model of Psychiatric Disorders.

Worldwide, more than 20 million people suffer from schizophrenia, but effective and definitive new therapeutic drugs/treatments have not been established. Vasoactive intestinal peptide receptor 2 (VIPR2) might be an attractive drug target for the treatment of schizophrenia because both preclinical and clinical studies have demonstrated a strong link between high expression/overactivation of VIPR2 and schizophrenia. Nevertheless, VIPR2-targeting drugs are not yet available. VIPR2 is a class-B G protein-coupled receptor that possesses high structural homology to its subtypes, vasoactive intestinal peptide receptor 1 (VIPR1) and pituitary adenylate cyclase-activating polypeptide type-1 receptor (PAC1). These biological and structural properties have made it difficult to discover small molecule drugs against VIPR2. In 2018, cyclic peptide VIpep-3, a VIPR2-selective antagonist, was reported. The aim of this study was to generate a VIpep-3 derivative for in vivo experiments. After amino acid substitution and structure optimization, we successfully generated KS-133 with 1) a VIPR2-selective and potent antagonistic activity, 2) at least 24 h of stability in plasma, and 3) in vivo pharmacological efficacies in a mouse model of psychiatric disorders through early postnatal activation of VIPR2. To the best of our knowledge, this is the first report of a VIPR2-selective antagonistic peptide that counteracts cognitive decline, a central feature of schizophrenia. KS-133 may contribute to studies and development of novel schizophrenia therapeutic drugs that target VIPR2.

Ellagitannins from Rosa roxburghii suppress poly(I:C)-induced IL-8 production in human keratinocytes.

The anti-inflammatory effects of a 50% aqueous extract of Rosa roxburghii fruit (RRFE) and two ellagitannins (strictinin and casuarictin) isolated from the RRFE were evaluated in a cell model of skin inflammation induced by self-RNA released from epidermal cells damaged by UV ray (UVR) irradiation. The RRFE inhibited interleukin-8 (IL-8) mRNA expression in normal human epidermal keratinocytes (NHEKs) stimulated with polyinosinic:polycytidylic acid (poly(I:C)), a ligand of toll-like receptor-3 (TLR-3). The plant-derived anti-inflammatory agents, dipotassium glycyrrhizinate (GK2) and allantoin, had no influence on the IL-8 expression. The purified compounds, strictinin and casuarictin, inhibited the IL-8 mRNA expression and IL-8 release induced in NHEKs by poly(I:C). These ellagitannins were thus found to be responsible for the biological activity exhibited by the RRFE. This study demonstrates that RRFE and isolated RRFE compounds show promise as ingredients for products formulated to improve skin disorders induced by UVR irradiation.

Generation of KS-58 as the first K-Ras(G12D)-inhibitory peptide presenting anti-cancer activity in vivo.

Ras mutations (e.g., occur in K-Ras, N-Ras, and H-Ras) are one of the most desirable and promising drug targets in chemotherapy treatments for cancer. However, there are still no approved drugs directly targeting mutated Ras. In 2017, an artificial cyclic peptide, KRpep-2d, was discovered as the first selective inhibitor of K-Ras(G12D), the most frequent K-Ras mutation. Here, we report the generation of KS-58, a KRpep-2d derivative that is identified as a bicyclic peptide and possess unnatural amino acid structures. Our in vitro data and molecular dynamics simulations suggest that KS-58 enters cells and blocks intracellular Ras-effector protein interactions. KS-58 selectively binds to K-Ras(G12D) and suppresses the in vitro proliferation of the human lung cancer cell line A427 and the human pancreatic cancer cell line PANC-1, both of which express K-Ras(G12D). Moreover, KS-58 exhibits anti-cancer activity when given as an intravenous injection to mice with subcutaneous or orthotropic PANC-1 cell xenografts. The anti-cancer activity is further improved by combination with gemcitabine. To the best of our knowledge, this is the first report of K-Ras(G12D)-selective inhibitory peptide presenting in vivo anti-cancer activity. KS-58 is an attractive lead molecule for the development of novel cancer drugs that target K-Ras(G12D).

Improvement of Learning and Memory in Senescence-Accelerated Mice by S-Allylcysteine in Mature Garlic Extract.

S-allylcysteine (SAC), a major thioallyl compound contained in mature garlic extract (MGE), is known to be a neuroactive compound. This study was designed to investigate the effects of SAC on primary cultured hippocampal neurons and cognitively impaired senescence-accelerated mice prone 10 (SAMP10). Treatment of these neurons with MGE or SAC significantly increased the total neurite length and number of dendrites. SAMP10 mice fed MGE or SAC showed a significant improvement in memory dysfunction in pharmacological behavioral analyses. The decrease of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, N-methyl-d-aspartate (NMDA) receptor, and phosphorylated α-calcium/calmodulin-dependent protein kinase II (CaMKII) in the hippocampal tissue of SAMP10 mice fed MGE or SAC was significantly suppressed, especially in the MGE-fed group. These findings suggest that SAC positively contributes to learning and memory formation, having a beneficial effect on brain function. In addition, multiple components (aside from SAC) contained in MGE could be useful for improving cognitive function by acting as neurotrophic factors.

Cynaropicrin is dual regulator for both degradation factors and synthesis factors in the cartilage metabolism.

AIMS: The molecular mechanism of osteoarthritis (OA) has never been understood clearly, but it has been suggested that imbalance of degradation and synthesis in cartilage contribute to the underlying mechanisms of OA. In this study, we investigated the effectiveness in the cartilage metabolism of the artichoke extract that includes the compound cynaropicrin. MAIN METHODS: We evaluated the efficacy of the artichoke extract or cynaropicrin in the cartilage metabolism factors and NF-κB signaling activity stimulated by inflammatory cytokine in chondrogenic cell lines, OUMS-27 and SW1353, using qRT-PCR, immunofluorescence and immunoblotting. KEY FINDINGS: We initially found that an artichoke extract and cynaropicrin both inhibited the increase of cartilage degradation factor MMP13 and further decreased the synthesis factor aggrecan induced by TNF-α in OUMS-27. In addition, cynaropicrin suppressed the enhancement of master regulator HIF-2α on cartilage degradation and further reduced the master regulator Sox9 on cartilage synthesis induced by TNF-α. We observed that cynaropicrin suppresses NF-κB signaling, which controls HIF-2α and Sox9. Since, HIF-2α is induced by p65 (RelA), we evaluated the effect of cynaropicrin and observed that it suppressed the nuclear transport of p65 (RelA) by inhibiting phosphorylation of IκBα. Moreover, cynaropicrin not only suppressed TNF-α stimulation, it had a similar effect on IL-1ß stimulation. No significant cytotoxicity with cynaropicrin was observed. SIGNIFICANCE: These finding suggest that cynaropicrin is an effective substance that can improve the balance of cartilage metabolism, by altering the equilibrium of cartilage degradation and synthesis induced by multiple mediators know to contribute to OA.

Indoleamine 2,3-dioxygenase 1 is upregulated in activated microglia in mice cerebellum during acute viral encephalitis.

Indoleamine 2,3-dioxygenase1 (IDO1) is the rate-limiting enzyme in the kynurenine pathway that converts l-tryptophan to l-kynurenine. Encephalomyocarditis virus (EMCV) can cause acute myocarditis in various animals including mice. Previously, IDO1 has been reported to have an important immunomodulatory function in immune-related diseases. However, the pathophysiological roles of IDO1 following acute viral infection of central nervous system are not fully understood. We observed that acute EMCV infection leads to a highly reproducible neuronal degeneration in mouse cerebellum. The goal of this study is to determine tissue/cell-specific and time-dependent expressions of IDO1 during acute EMCV infection in mouse cerebellum. IDO1 was up-regulated in microglia, which was recognized to be activated morphologically and positive for ionized calcium-binding adapter molecule 1 (Iba-1), a protein expressed in microglia, within EMCV-induced cerebellar lesions showing neuronal degeneration although the very weak expression of IDO1 is detected only in cytoplasm of Purkinje cells. No GFAP immunostaining was observed in EMCV-induced cerebellar lesions although many reactive astrocytes surrounding the lesions showed strongly positive immunostaining for GFAP 10 days after the viral inoculation. Thus, IDO1 expression may affect EMCV-induced neuronal degeneration in cerebellum.