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The rare occurrence of human cryptococcosis caused by Cryptococcus gattii sensu lato leads to difficulties in establishing the antifungal susceptibility profile between species of this potentially lethal pathogen, which may be crucial for treating cryptococcosis.To establish an antifungal susceptibility profile of C. gattii s.l. in Taiwan.A total of 104 environmental C. gattii s.l. strains (including multilocal sequence typing ST7, ST106, ST274, ST328, ST546, ST548 and ST630) and 21 previously collected clinical strains (including ST7, ST44, ST06, ST274, ST328 and ST329) were included in this study. We determined the minimum inhibitory concentrations (MICs) of six antifungal agents (itraconazole, fluconazole, voriconazole, posaconazole, flucytosine and amphotericin B) against environmental C. gattii s.l. strains and compared the antifungal susceptibility profiles of environmental strains with those of clinical strains.The antifungal susceptibility data demonstrated that the MICs of antifungal agents against environmental strains were comparable to those against clinical strains. Compared with strains of Cryptococcus deuterogattii, those of C. gattii sensu stricto were more susceptible to azoles and flucytosine. The differences in antifungal susceptibility between the strains of each sequence type (ST) were significant. Correlation analysis of MICs revealed cross-resistance between azoles in environmental strains of C. gattii s.l. Geographic differences in the antifungal susceptibility of C. gattii s.l. isolated from different cities in Taiwan were observed in this study.Clinical and environmental strains were indistinguishable in antifungal susceptibility. The antifungal susceptibility of C. gattii s.l. is associated with STs. Therefore, establishing an ST-oriented domestic antifungal susceptibility database may help treat C. gattii s.l.-induced cryptococcosis.

Late initiation (LI) of combination antiretroviral therapy (cART)-defined as having a CD4This multicenter retrospective cohort study enrolled 1198 patients with newly diagnosed HIV infection during 2009-2019 who were grouped by status at cART initiation: those without LI (non-LI group, 56.01%); those with LI but without late presentation (LP) of HIV (LP: a CD4 + count of 200 cells/μL at HIV presentation or AIDS events ≤ 3 months of HIV diagnosis) [LILP(-) group, 4.51%]; and those with LI and with LP of HIV [LILP(+) group, 39.48%]. Joinpoint regression was used to identify changes in LI proportion.The median CD4Given the rise in LI from 2015 in the era of treat-all and rapid cART initiation, strategic interventions to increase earlier cART initiation must be intensified in Taiwan, especially among populations with delayed access to HIV testing services.

Background The rare occurrence of cryptococcosis caused by Cryptococcus gattii sensu lato (C. gattii s.l.) leads to the difficulties in studying the molecular epidemiology of this globally emerging disease. Objectives To establish the molecular epidemiological profile of C. gattii s.l. in Taiwan, and understand the genetic relationship between locally endemic and global isolates. Methods A nationwide survey on environmental C. gattii s.l. in Taiwan was conducted from 2017 to 2019. The geographic distribution and molecular epidemiology based on multilocus sequence typing (MLST) data of the environmental isolates were compared with 18 previously collected clinical isolates. Phylogenetic analysis was performed to elucidate the genetic relationship between the global isolates and the isolates endemic to Taiwan. Results From a total of 622 environmental samples, 104 (16.7%) were positive for C. gattii s.l.. Seven sequence types were identified among the environmental isolates. The genetic population structure showed that the environmental and clinical isolates were closely linked by sequence types and geographical locations. Phylogenetic analysis revealed the association between the C. gattii s.l. isolates in Taiwan and those from South America and South Asia. The recombination test suggested that, in Taiwan, the C. gattii sensu stricto (C. gattii s.s). isolates undergo clonal reproduction and sexual recombination, whereas C. deuterogattii isolates were clonal. Conclusions The molecular epidemiology of environmental C. gattii s.l. isolates is closely linked to the clinical isolates. Phylogenetic analysis of the environmental isolates provides an insight into the mechanisms underlying reproduction and dispersal of C. gattii s.l. in Taiwan.

Carbapenemase-producing combined porin loss is one of the primary mechanisms for carbapenem resistance. Although mutations in ompK35 and ompK36 genes have often been identified in carbapenem-resistant Klebsiella pneumoniae, reports on the porin protein gene disruption by insertion sequence (IS) elements are varied. The ompK36 porin protein gene disruption by IS elements and OmpK36 production loss in six blaKPC-2-carrying K. pneumoniae isolates were detected in this study. IS903, ISEc68, and IS1 insertions were noted in the 3, 2, and 1 isolates, respectively. The six K. pneumoniae isolates showed five different pulsed-field gel electrophoresis patterns and belonged to four multilocus sequence typing types, ST4, ST11, ST15, and ST39. This study increases our understanding of the genetic background of KPC-2 carbapenemases in porin-defective clinical isolates and the contribution of OmpK36 production loss to carbapenem resistance.

Introduction. Antimicrobial resistance associated with animal hosts is easily transmitted to humans either by direct contact with resistant organisms or by transferring resistance genes into human pathogens. Gap statement. There are limited studies on antimicrobial resistance genes and genetic elements of multidrug-resistant (MDR) Escherichia coli in veterinary hospitals in Taiwan. Aim. The aim of this study was to investigate antimicrobial resistance genes in multidrug-resistant Escherichia coli from animals. Methodology. Between January 2014 and August 2015, 95 multidrug-resistant Escherichia coli isolates were obtained from pigs (n=66), avians (n=18), and other animals (n=11) in a veterinary hospital in Taiwan. Susceptibility testing to 24 antimicrobial agents of 14 antimicrobial classes was performed. Antimicrobial resistance genes, integrons, and insertion sequences were analysed by polymerase chain reaction and nucleotide sequencing. Pulsed-field gel electrophoresis (PFGE), and multi-locus sequence typing were used to explore the clonal relatedness of the study isolates. Results. Different antimicrobial resistance genes found in these isolates were associated with resistance to β-lactams, tetracycline, phenicols, sulfonamides, and aminoglycosides. Fifty-five of 95 E. coli isolates (55/95, 57.9 %) were not susceptible to extended-spectrum cephalosporins, and bla CTX-M-55 (11/55, 20.0 %) and bla CMY-2 (40/55, 72.7 %) were the most common extended-spectrum β-lactamase (ESBL) and AmpC genes, respectively. Both bla CTX-M and bla CMY-2 were present on conjugative plasmids that contained the insertion sequence ISEcp1 upstream of the bla genes. Plasmid-mediated FOX-3 β-lactamase-producing E. coli was first identified in Taiwan. Forty isolates (40/95, 42 %) with class 1 integrons showed seven resistance phenotypes. Genotyping of 95 E. coli isolates revealed 91 different XbaI pulsotypes and 52 different sequence types. PFGE analysis revealed no clonal outbreaks in our study isolates. Conclusion. This study showed a high diversity of antimicrobial resistance genes and genotypes among MDR E. coli isolated from diseased livestock in Taiwan. To our knowledge, this is the first report of plasmid-mediated ESBL in FOX-3 β-lactamase-producing E. coli isolates in Taiwan. MDR E. coli isolates from animal origins may contaminate the environment, resulting in public health concerns, indicating that MDR isolates from animals need to be continuously investigated.

Cryptococcosis is an opportunistic infection caused by the Cryptococcus species complex. An outbreak of cryptococcosis caused by Cryptococcus gattii (AFLP6/VGII) in North America has indicated the need for studies of this organism and its environmental niche. Difficulties in isolating the Cryptococcus spp. because of the overgrowth of filamentous fungi onto culture media and its low fungal population size under natural conditions limit studies of these pathogenic yeasts. We designed a selective medium that inhibits the growth of environmental filamentous fungi but does not inhibit that of Cryptococcus cells. After enrichment in acidified YPD media and inoculation onto selective media, Cryptococcus cells in brown-coloured colonies were isolated from environmental materials. This two-step method is useful for isolating environmental members of the Cryptococcus species complex, which is essential for further studies involving diversity and the microbe-environment relationship of this yeast.

Non-toxic natural products useful in the formulation of cosmetics are of considerable interest. Recent efforts have focused on the identification of substances that inhibit tyrosinase activity or suppress formation of reactive oxygen species (ROS) in skin cells. Since tyrosinase is the rate-limiting enzyme in the synthesis of melanin, the pigment responsible for the color of human skin, tyrosinase inhibitors may have skin-whitening effects. Since ROS have been implicated in the aging of human skin, agents that suppress the production of ROS may retard such aging. In the present study, ethanol (95%) extracts of 26 plants endemic to Taiwan were examined for their tyrosinase-inhibitory or melanin-reducing activities in human epidermal melanocytes, as well as in vitro free radical scavenging activity. Among the preparations tested, extracts of Pyracantha koidzumii (M-165) were found to be the least cytotoxic and to possess the highest cellular tyrosinase inhibitory activity (IC50, 54.8 μg/mL). Extracts of Acer albopurpurascens (M-51), Hygrophila pogonocalyx (M169), Machilus japonica var. kusanoi (M-67), and Eriobotrya deflexa (M-50) exhibited the most potent free radical scavenging activity against hydroxyl, superoxide, and 2, 2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) anion radicals. The IC(subscript 50) values of M-51, M169, M-67, and M-50 were 3.1, 0.8, 6.6, and 1.8 μg/mL for hydroxyl radical; 5.3, 12.8, 12.4, and 6.1 μg/mL for superoxide radical; 2.4, 7.9, 5.7, and 2.9 μg/mL for ABTS anion radical, respectively. Nevertheless, the phenolic contents were not all correlated with these activities. These plants thus serve as potential sources of ingredients, which could be combined in cosmetic products. Further investigation of the substances responsible for the observed tyrosinase-inhibitory and free radical scavenging activities is therefore warranted.

Carbapenemase-producing combined porin loss is one of the primary mechanisms for carbapenem resistance. Although mutations in

Purpose. Carbapenem-resistant Klebsiella pneumoniae (CRKP) has emerged as a major challenge for global healthcare systems. The objectives of this study were to determine the nosocomial spread of CRKP clones and analyse the molecular characteristics of CRKP in our hospital. Methodology. Ninety-eight non-duplicated clinical CRKP isolates were collected from March 2014–June 2015. Clinical, demographic and microbiological data of patients with CRKP were reviewed. Pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing were applied to investigate the genetic relationship between the 98 isolates. Antibiotic resistance genes were identified by conventional PCR-sequencing. Results. PFGE patterns were grouped into 26 clusters. Two main PFGE clusters were identified: L (53 isolates, belonging to ST11) and N (11 isolates, belonging to ST11). The most dominant ST was ST11 (79 %, 77/98), followed by ST273 (5 %, 5/98). KPC-2 (n=82) was the predominant carbapenemase followed by OXA-48 (n=64). Fifty isolates (51 %, 50/98) harboured bla KPC-2 and bla OXA-48 simultaneously, and three of these isolates were detected with the third carbapenemase genes (bla IMP-8 or bla VIM-1). Conclusion. The clonal spread of K. pneumoniae ST11 expressing OXA-48, KPC-2 and CTX-M-14 β-lactamases was the cause of an outbreak of CRKP. To the best of our knowledge, a single strain harbouring A-, B- and D-class carbapenemase genes has not previously been identified. There is a high prevalence of plasmid-encoded KPC-2- and OXA-48-producing CRKP in our hospital; most isolates were members of ST11, which may be representative of a high-risk CRKP clone disseminating in central Taiwan.

Cortical spreading depolarization (CSD) reflects neuronal depolarization slowly propagating in the cortex and is accompanied with vascular response. Dynamic ultrafast Doppler has potential to access the hemodynamic response to CSD, specifically to the penetrating vessels in the cortex. In this study, the CSD was experimentally triggered by topical application of potassium chloride (KCl) on the surface of the brain. The vascular response caused by KCl induced CSD was successfully detected during the dynamic ultrafast Doppler. The propagation of vascular responses from the center of right cortex (stimulated side) to the medial and lateral side was observed. The perfusion change (ΔCBV) decreased over 30 % from the baseline at the beginning, and the CBV response subsequently elevated dramatically to over 30% very likely due to vasodilation. At the end of episode, CBV decreased slowly along the time. In this study, we firstly detect the vascular responses caused by KCl induced CSD with the dynamic ultrafast Doppler.

Increased Crabp2 levels have been found in various types of cancer, and are associated with poor patients’ survival. Although Crabp2 is found to be overexpressed in lung cancer, its role in metastasis of lung cancer is unclear. In this study, Crabp2 was overexpressed in high-metastatic C10F4 than low-metastatic lung cancer cells. Analysis of clinical samples revealed that high CRABP2 levels were correlated with lymph node metastases, poor overall survival, and increased recurrence. Knockdown of Crabp2 decreased migration, invasion, anoikis resistance, and in vivo metastasis. Crabp2 was co-immunoprecipitated with HuR, and overexpression of Crabp2 increased HuR levels, which promoted integrin β1/FAK/ERK signaling. Inhibition of HuR or integrin β1/FAK/ERK signaling reversed the promoting effect of Crabp2 in migration, invasion, and anoikis resistance. Knockdown of Crabp2 further inhibited the growth of cancer cells as compared with that by gemcitabine or irinotecan alone. The expression of Crabp2 in human lung tumors was correlated with stress marker CHOP. In conclusion, our findings have identified the promoting role of Crabp2 in anoikis resistance and metastasis. CRABP2 may serve as a prognostic marker and targeting CRABP2 may be exploited as a modality to reduce metastasis.

Up-regulation of hypoxia-inducible factor-1α (HIF-1α), even in normoxia, is a common feature of solid malignancies. However, the mechanisms of increased HIF-1α abundance, and its role in regulating breast cancer plasticity are not fully understood. We have previously demonstrated that dimethyl-2-ketoglutarate (DKG), a widely used cell membrane-permeable α-ketoglutarate (α-KG) analogue, transiently stabilizes HIF-1α by inhibiting prolyl hydroxylase 2. Here, we report that breast cancer tumorigenicity can be acquired through prolonged treatment with DKG. Our results indicate that, in response to prolonged DKG treatment, mitochondrial respiration becomes uncoupled, leading to the accumulation of succinate and fumarate in breast cancer cells. Further, we found that an early increase in the oxygen flux rate was accompanied by a delayed enhancement of glycolysis. Together, our results indicate that these events trigger a dynamic enrichment for cells with pluripotent/stem-like cell markers and tumorsphere-forming capacity. Moreover, DKG-mediated metabolic reprogramming results in HIF-1α induction and reductive carboxylation pathway activation. Both HIF-1α accumulation and the tumor-promoting metabolic state are required for DKG-promoted tumor repopulation capacity in vivo. Our data suggest that mitochondrial adaptation to DKG elevates the ratio of succinate or fumarate to α-KG, which in turn stabilizes HIF-1α and reprograms breast cancer cells into a stem-like state. Therefore, our results demonstrate that metabolic regulation, with succinate and/or fumarate accumulation, governs the dynamic transition of breast cancer tumorigenic states and we suggest that HIF-1α is indispensable for breast cancer tumorigenicity.

Most lung cancer patients are diagnosed late with metastasis, which is the major cause of cancer-related death and recurrent tumors that often exhibit chemoresistance. In the present study, we initially identified gap junction beta-4 protein (Gjb4) to be overexpressed in highly metastatic cancer cells selected by their enhanced binding to serum components. Overexpression or knockdown of Gjb4 increased or decreased lung metastasis of syngeneic mice, respectively. We found that Gjb4 expression was higher in lung tumors than normal tissues (p = 0.0026), and Gjb4 levels in blood buffy coat samples showed significant performance in diagnosing stage I–III (p = 0.002814) and stage IV (p

The catalyst layer (CL) of a polybenzimidazole (PBI) membrane electrode assembly (MEA) consists of Pt–C (Pt on a carbon support), PBI, and H 3 PO 4 . Two series of catalyst ink solutions each containing Pt–C, N,N′-dimethyl acetamide, and PBIs comprising four different molecular weights (MWs) ( i.e. , M w = 1.1 × 10 4 , 4.4 × 10 4 , 9.0 × 10 4 , and 17.4 × 10 4 g mol −1 ) are used to fabricate CLs. One catalyst ink solution series is mixed with LiCl, while the other solution series lacks LiCl. We demonstrate that the CL prepared using a lower MW PBI has a higher electrochemical surface area, lower charge transfer resistance, and higher fuel cell performance. The addition of LiCl enhances the dispersion of the high MW PBIs in the catalyst ink solution and acts as a foaming agent in CL, thus improving fuel cell performance. However, LiCl exerts small influence on the fuel cell performance of the MEAs fabricated using low MW PBIs.

The neurotoxin 6-hydroxydopamine (6-OHDA) has been widely used to generate an experimental model of Parkinson's disease. This model is crucial in the search for compounds that diminish 6-OHDA-induced nerve growth factor (NGF)-differentiated PC12 cell death. Nephrocizin (luteolin-7-O-β-D-glucopyranoside), a flavone glycoside, was isolated from widely distributed plants. The protective effects of pre-treatment with nephrocizin on the induced neurotoxicity in PC12 cells by 6-OHDA and its oxidative products, H₂O₂-, and p-quinone, were evaluated herein. Nephrocizin promoted cell viability, scavenged ROS-related products, increased cellular glutathione (GSH) levels, and reduced caspase-3 and -8 activities in 6-OHDA-, H₂O₂-, or p-quinone-treated PC12 cells. Furthermore, nephrocizin-conjugated metabolites in PC12 cells were identified with the boronate-affinity method and LC-MS technology, and preferential regioselectivity at the C2' and C5' positions by the nephrocizin-GSH (or NAC) adduct method was observed. These lines of evidence established that nephrocizin could form a dimer to diminish the intracellular ROS. These results demonstrate the first neuroprotective mechanism of nephrocizin against 6-OHDA-, H₂O₂- or p-quinone-induced cytotoxicity in PC12 cells via chemical and biological studies. These dietary antioxidants are potential candidates for use in intervention in neurodegenerative diseases.

Congenital insensitivity to pain with anhidrosis (hereditary sensory and autonomic neuropathy type IV) is a rare autosomal recessive disorder caused by a defect in neurotrophic tyrosine kinase receptor and nerve growth factor, as reported in previous studies. This report is of a 6-month-old male infant with typical symptoms and signs of congenital insensitivity to pain with anhidrosis. He had a homozygous insertion mutation with c.2086_2087 ins C of neurotrophic tyrosine kinase receptor type 1 (NTRK1) gene with both parents as heterozygous carriers. This mutation may have a strong relation to hereditary sensory and autonomic neuropathy type IV Taiwanese patients. This is the youngest reported patient in Taiwan and first reported with congenital insensitivity to pain with mutation of NTRK1 gene inherited from the parents. Early diagnosis may provide appropriate medical care and education for these children and their families for better prognosis.

Multiple advantages-including the short generation time, large numbers of fertilized eggs, low cost of cultivation and easy maintenance favor the use of fish as bioreactors for the production of pharmaceutical proteins. In the present study, zebrafish eggs were used as bioreactors to produce mature tilapia insulin-like growth factors (IGFs) proteins using the oocyte-specific zona pellucida (zp3) promoter. The chimeric expression plasmids, pT2-ZP-tIGFs-IRES-hrGFP, in which hrGFP was used as reporter of tilapia IGFs expression, were designed to established Tg (ZP:tIGFs:hrGFP) transgenic lines for the expression of tilapia IGF-1 and IGF-2. Recombinant tilapia IGF-1 and IGF-2 were expressed as soluble forms in cytoplasm of fertilized eggs. The content level of tilapia IGF-1 and IGF-2 were 6.5 and 5.0% of the soluble protein, respectively. Using a simple Ni-NTA affinity chromatography purification process, 0.58 and 0.49 mg of purified tilapia IGF-1 and IGF-2 were obtained, respectively, from 650 fertilized eggs. The biological activity of the purified tilapia IGF-1 and IGF-2 was confirmed via a colorimetric bioassay to monitor the growth stimulation of zebrafish embryonic cells (ZF4), tilapia ovary cells (TO-2) and human osteosarcoma epithelial cells (U2OS). These results demonstrate that the use of zebrafish eggs as bioreactors is a promising approach for the production of biological recombinant proteins.

Since the mid-1990s, novel G9 rotaviruses have been detected in many countries, suggesting that G9 is a globally important serotype. The molecular epidemiology of G9 rotaviruses in Taiwan from 2000 to 2002 was investigated in this study. G9 rotavirus first appeared in 2000 with 4 cases and constituted 33.8% and 54.8% of the rotavirus-positive samples in 2001 and 2002, respectively. These G9 strains belonged to P[8]G9, subgroup II, and long electropherotype, except one belonged to P[4]G9, subgroup II, and short electropherotype. Nucleotide sequencing and phylogenetic analysis of 52 Taiwanese G9 rotaviruses showed that the VP7 genes shared a high degree of identity to overseas G9 rotaviruses detected after 1993 and that the VP8* portions of the VP4 genes were more closely related to those of local rotaviruses of other G types. The two P[8]G9 strains with high nucleotide identities in the VP7 and the partial VP4 genes, 01TW591 of Taiwan from 2001 and 95H115 of Japan from 1995, varied in four genes, genes 2, 3, 7, and 8, which was revealed by RNA-RNA hybridization. Representative strains for different RNA patterns were also analyzed in the partial VP2 and VP3 genes; the nucleotide identities were high between Taiwanese G9 strains and local G3 or G2 strains. These results suggested that Taiwanese G9 rotaviruses possibly had evolved through reassortment between overseas G9 strains and circulating rotaviruses of other G types.

Plant phenolic compounds isolated from a 70% aqueous acetone extract of the leaves of Malus doumeri A. CHEV. var. formosana (KAWAK. & KOIDZ.) S. S. YING, a type of Taiwanese indigenous plant, were evaluated for potential application in the field of skin care. A phytochemical investigation of the active fractions resulted in the isolation of seven compounds of which the structures were identified by spectroscopic characterization. In the present study, the isolated phenolic compounds were evaluated for their free radical-scavenging activities against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and the superoxide radicals, anti-elastase, and for their anti-matrix metalloproteinase-1 (MMP-1) activity in human skin fibroblast cells. Of these compounds, 3-hydroxyphloridzin (2), 3-hydroxyphloretin (6), and quercetin (7) exhibited the strongest DPPH and superoxide radical-scavenging activities. The IC50 values of these compounds were 9.2, 7.7, and 15.4 microM, respectively, for the DPPH radical, and 25.0, 19.6, and 42.6 microM, respectively, for the superoxide radical. 3-Hydroxyphloridzin (2) and 3-hydroxyphloretin (6) also showed xanthine oxidase inhibitory activity, with IC(50) values of 52.1 and 22.4 muM, respectively. In the test for elastase inhibitory activity, phloretin (5) and 3-hydroxyphloretin (6) were the most potent compounds. Phloretin (5), 3-hydroxyphloretin (6), and quercetin (7) showed better inhibition of MMP-1 production in fibroblast cells. To the best of our knowledge, this is the first time that the active phenolic compounds from M. doumeri var. formosana have been isolated, reported, and described. The above results suggest that the extract of M. doumeri var. formosana containing phenolic compounds could be suitable naturally occurring active constituents for use in anti-aging or cosmetic products.

Progranulin (PGRN) is a secreted growth factor that has been implicated in diverse biological processes including wound healing, embryo development, morphogenesis and disease. We previously showed that PGRN is induced along with IGF-1 in tilapia liver upon GH administration. In the present study, we demonstrate that the coinduction of PGRN and IGF-1 by GH administration is a common regulation in fish also presented in zebrafish. To understand the regulatory mechanism of GH-induced PGRN expression, the zebrafish PGRN promoter was isolated and analyzed. We found that a region from -2400 to -3000 within PGRN promoter is essential for GH-induced PGRN expression. A promoter competition assay showed that HNF-3 β, HNF-1 α and C/EBP β binding motifs within the -2400/-3000 region of PGRN promoter contributed to GH-induced PGRN expression. The individual or concomitant transfection of HNF-3 β, HNF-1 α and C/EBP β into HepG2 cells showed that the three transcriptional regulators all participate in GH-induced PGRN expression with independent activities. These results demonstrated that the liverenriched transcription factors HNF-3 β, HNF-1 α and C/EBP β are involved in GH-induced PGRN expression in liver. We expect that these results will be useful in understanding the regulatory relationship between PGRN and IGF-1 in response to GH regulation.

Background Cyclic vomiting syndrome (CVS) is an episodic nausea and non-bilious vomiting disorder characterized by recurrent stereotypic symptoms with disease-free intervals. CVS in children is associated with a high prevalence of migraine, and is commonly considered a precursor to migraine. This study aimed to investigate the clinical manifestations of pediatric CVS and its prognosis, and to clarify its relationship with the risk of migraine development in children. Methods The clinical features of children diagnosed with CVS before the age of 18 years at the designated hospital were retrospectively studied over the past 30 years (1976–2006) based on the Rome III or ICHD II criteria. Clinical evaluations, including age of onset, sex, family history, symptoms and duration during attacks, frequency, trigger events, electroencephalogram, treatment and subsequent development of migraine were assessed from chart records and telephone interviews. Results Thirty-five children (17 males and 18 females) were enrolled. Their age of onset ranged from 2 to 17 years (mean, 6.8 ± 3.1 years) and frequency of attacks ranged from once to 36 times per year (mean, 8.2 ± 7.6 times). Duration of symptoms during each attack ranged from 1 to 45 days (mean, 5.9 ± 7.3 days). Of 20 children assessed for migraine development, seven subsequently developed typical migraine symptoms. There was younger onset age in the migraine-positive subgroup (5 ± 1.7 years) than in the migraine-negative subgroup (8.9 ± 3 years; p = 0.001). Co-morbid headache during CVS attack was also more evident in the migraine-positive subgroup (28.6% vs. 0%). Conclusion Results of the study show that younger onset age and headache during CVS attacks may have increased risk of migraine development. Large-scale prospective studies are warranted to further clarify the relationship between CVS and migraine.

Chronic neurodegenerative disorders are having an increasing impact on public health as human longevity increases. Parkinson's disease (PD) is a degenerative disorder of the central nervous system and is characterized by motor system disorders resulting in loss of dopamine-producing brain cells. Pueraria thomsonii Benth. (Fabaceae) is an herbal medicine that has traditionally been used as an antipyretic agent. In the present study, the active constituents, daidzein and genistein, were isolated from P. thomsonii. Both compounds exhibited neurocytoprotective effects against 6-hydroxydopamine (6-OHDA)-induced cytotoxicity in nerve growth factor (NGF)-differentiated PC12 cells. Neither daidzein nor genistein affected 6-OHDA-induced cellular reactive oxygen species (ROS) generation according to flow cytometric analysis. Rather, they inhibited caspase-8 and partially inhibited caspase-3 activation, providing a protective mechanism against 6-OHDA-induced cytotoxicity in NGF-differentiated PC12 cells. The present results imply that daidzein and genistein may be useful in the development of future strategies for the treatment of PD.

Aim of the Study There is greater consumer awareness of plant-based skin-care products. Sophora japonica L. (Fabaceae) has been used traditionally as a hemostatic agent and also has skin-care and whitening benefits. The effect of the isolated active compounds of Sophora japonica L. (Fabaceae) that inhibits tyrosinase activity in human epidermal melanocytes (HEMn) was examined. Materials and methods We used the mushroom tyrosinase inhibitory assay to isolate active constituents from the extracts. The structures of these constituents were characterized by physical and spectroscopic analyses. Cellular tyrosinase kinetics were analyzed and showed by Lineweaver–Burk plot. Results A new compound, N -feruloyl- N ′- cis -feruloyl-putrescine ( 8 ), together with four flavonoids and three putrescine derivatives were obtained after assay-guided isolation of S. japonica . In HEMn, compound 8 was minimally cytotoxic (cell viability >90% at 100 μM) and the IC 50 value for suppression of cellular tyrosinase activity was estimated as 85.0 μM. Zymography analysis demonstrated the compound's concentration-dependent effects and the kinetic analysis indicated the compound's mixed-inhibitory action. Conclusions We concluded that the new compound 8 is the most potent component of S. japonica yet discovered. Its pigment inhibition activity may be exploitable cosmetically.

Based on nucleotide sequence and phylogenetic analysis of the partial VP6 genes, group A rotaviruses can be mainly differentiated into two genogroups. In this study, a method employing reverse transcription-PCR (RT-PCR) and degenerate primers was established to assign the VP6 genogroup. VP6 genogroup I and genogroup II could be determined according to the sizes of the amplicons: 380 and 780 bp, respectively. The VP6 genogroup of human reference strains of G1 to G4 and G9 types and RotaTeq vaccine strains could be properly assigned by RT-PCR. Eighty rotavirus-positive fecal samples were subjected to enzyme-linked immunosorbent assay (ELISA), RT-PCR, and sequencing of the partial VP6 gene for subgroup and genogroup determination. The results correlated well among these three methods, except for seven samples whose subgroups could not be determined by ELISA. VP6 genogroups of another 150 rotavirus strains recovered between 1981 and 2005 were determined by RT-PCR and sequencing, and the same results were obtained by these two methods. Furthermore, an additional 524 rotavirus-positive fecal samples were tested by RT-PCR, and the VP6 genogroups could be easily determined. The RT-PCR assay developed here provided a reliable and convenient method for assigning the VP6 genogroups of human rotaviruses with a wide range of genetic variation.

Tyrosinase is a copper-containing monooxygenase that catalyzes melanin synthesis in skin melanocytes. Herein, 13 compounds from the Formosan apple (Malus doumeri var. formosana), an indigenous Taiwanese plant, were isolated and identified. The active constituents were identified as 3-hydroxyphloretin (7) and catechol (9); they exhibited potent hydroxyl radical-scavenging (IC(50) values, 0.6 and 1.1 microM) and cellular tyrosinase-reducing activities (IC(50) values, 32 and 22 microM) in human epidermal melanocytes. In addition, we evaluated the level of several tyrosinase-related proteins by Western blot analysis. In contrast to 3-hydroxyphloretin (7), which showed no effect on the level of these proteins, catechol (9) reduced their activity and the expression of the respective genes, as determined by quantitative real-time PCR. In a kinetic analysis of mushroom tyrosinase, 3-hydroxyphloretin (7) was a competitive inhibitor. These two constituents exhibited metal-coordinating interactions with copper ions in a virtual model of molecular docking with human tyrosinase. Thus, 3-hydroxyphloretin (7) and catechol (9) were the most active constituents from the Formosan apple; they exhibited anti-oxidant and tyrosinase reducing activities, suggesting their possible use as cosmetic agents.

Plant phenolic compounds isolated from a 70% aqueous acetone extract of the leaves of Malus doumeri A. CHEV. var. formosana (KAWAK.KOIDZ.) S. S. YING, a type of Taiwanese indigenous plant, were evaluated for potential application in the field of skin care. A phytochemical investigation of the active fractions resulted in the isolation of seven compounds of which the structures were identified by spectroscopic characterization. In the present study, the isolated phenolic compounds were evaluated for their free radical-scavenging activities against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and the superoxide radicals, anti-elastase, and for their anti-matrix metalloproteinase-1 (MMP-1) activity in human skin fibroblast cells. Of these compounds, 3-hydroxyphloridzin (2), 3-hydroxyphloretin (6), and quercetin (7) exhibited the strongest DPPH and superoxide radical-scavenging activities. The IC50 values of these compounds were 9.2, 7.7, and 15.4 microM, respectively, for the DPPH radical, and 25.0, 19.6, and 42.6 microM, respectively, for the superoxide radical. 3-Hydroxyphloridzin (2) and 3-hydroxyphloretin (6) also showed xanthine oxidase inhibitory activity, with IC(50) values of 52.1 and 22.4 muM, respectively. In the test for elastase inhibitory activity, phloretin (5) and 3-hydroxyphloretin (6) were the most potent compounds. Phloretin (5), 3-hydroxyphloretin (6), and quercetin (7) showed better inhibition of MMP-1 production in fibroblast cells. To the best of our knowledge, this is the first time that the active phenolic compounds from M. doumeri var. formosana have been isolated, reported, and described. The above results suggest that the extract of M. doumeri var. formosana containing phenolic compounds could be suitable naturally occurring active constituents for use in anti-aging or cosmetic products.

Spatholobus suberectus Dunn (Leguminosae) is a traditional Chinese herbal medicine used to treat rheumatism, anemia, menoxenia, and other disorders. The extent to which this herbal medicine is useful to skin cells, however, has not been evaluated. Constituents of the 95% ethanol extracts of the dried vine stems of S. suberectus were therefore isolated and examined for their skin-whitening capacity. A bio-guided phytochemical investigation, involving use of the mushroom tyrosinase inhibitory system, of active fractions of the extracts resulted in the isolation of 12 constituents. The structures of these constituents, which were characterized by various spectroscopic techniques, consisted of one flavone, three isoflavones, five flavanones, two flavanonols, and one chalcone. Of these constituents 3',4',7-trihydroxyflavone (1), eriodictyol (3), plathymenin (5), dihydroquercetin (6), butin (7), neoisoliquiritigenin (8), dihydrokaempferol (9), liquiritigenin (10), and 6-methoxyeriodictyol (12) represented compounds isolated for the first time from S. suberectus. These constituents were evaluated their ability to inhibit cellular tyrosinase activity and for their melanin inhibitory activity in human epidermal melanocytes (HEMn). Butin (7) was the most efficacious of these constituents and exhibited concentration-dependent effects. Western blot analysis revealed that expression of tyrosinase and tyrosinase-related proteins 1 and 2 (TRP1 and TRP2) was decreased in butin (7)-treated HEMn cells. Additionally, quantitative real-time PCR (qRT-PCR) analysis disclosed that expression of mRNAs for tyrosinase, TRP1 and TRP2 was suppressed by butin (7). It is concluded that butin (7) is the most active of the components of S. suberectus in inhibiting pigmentation and that this inhibition is exerted through inhibition of transcription of the genes encoding tyrosinase, TRP1 and TRP2.

Corrigendum to ‘‘Constituents from the Formosan apple reduce tyrosinase activity in human epidermal melanocytes’’ [Phytochemistry 68 (2007) 1189–1199] Yi-Pei Lin , Feng-Lin Hsu , Chien-Shu Chen , Ji-Wang Chern , Mei-Hsien Lee b,* a Graduate Institute of Pharmacognosy, College of Pharmacy, Taipei Medical University, 250 Wu Hsing Street, Taipei 110, Taiwan b School of Pharmacy, College of Medicine, National Taiwan University, Taipei 100, Taiwan