Positive Association Between Fluoroquinolone Exposure and Tendon Disorders: A Nationwide Population-Based Cohort Study in Taiwan.

Introduction: Fluoroquinolone exposure is reportedly associated with a higher risk of tendon disorders, tendonitis, or tendon rupture. However, studies in East Asian populations have not confirmed these risks in patients with comorbidities or concomitant medication use. This cohort study was designed to investigate the associations among fluoroquinolone exposure, comorbidities, medication use, and tendon disorders in Taiwan. Materials and Methods: This population-based, nationwide, observational, cohort study used data from the National Health Insurance Research database in Taiwan, a nationwide claims database that covers more than 99% of the Taiwanese population. The study period was from January 2000 to December 2015, and the median follow-up time was 11.05 ± 10.91 years. Patients who were exposed to fluoroquinolones for more than three consecutive days were enrolled, and patients without fluoroquinolone exposure who were matched by age, sex, and index year were enrolled as controls. The associations of comorbidities and concomitant medication use with tendon disorder occurrence were analyzed using Cox regression models. Results: The incidence of tendon disorders were 6.61 and 3.34 per 105 person-years in patients with and without fluoroquinolone exposure, respectively (adjusted hazard ratio, 1.423; 95% confidence interval [1.02,1.87]; p = 0.021). Sensitivity analyses yielded similar results. Patients under 18 and over 60 years with fluoroquinolone exposure; those with chronic kidney disease, diabetes, rheumatologic disease, cardiac disease, lipid disorder, or obesity; and those who concomitantly used statins, aromatase inhibitors, or glucocorticoids, had a significantly higher risk of tendon disorders. Conclusion: The long-term risk of tendon disorders was higher in patients with fluoroquinolone exposure than in those without fluoroquinolone exposure. Clinicians should assess the benefits and risks of fluoroquinolone use in patients at high risk of tendon disorders who require fluoroquinolone administration.

New insights into precocious puberty and ADHD: a nationwide cohort study.

BACKGROUND: Attention deficit-hyperactivity disorder (ADHD) is one of the most common neurodevelopmental disorders in children; however, studies delineating the association between ADHD and central precocious puberty are limited. This study aimed to understand whether children with ADHD are at a higher risk of central precocious puberty. METHODS: This population-based retrospective cohort study was conducted using the National Health Insurance Research Database of Taiwan to investigate the association between ADHD and the incidence of central precocious puberty between 2000-2015. We identified ADHD individuals treated with methylphenidate, atomoxetine or not. The control cohort consisted of individuals without ADHD. The outcome measure was central precocious puberty diagnosis. RESULTS: Among 290,148 children (mean age: 5.83 years), central precocious puberty incidence was 4.24 and 1.95 per 105 person-years in the ADHD and control groups, respectively. Children with ADHD treated with medication had a higher risk than those without ADHD. However, medication use did not affect the incidence of central precocious puberty among children with ADHD. CONCLUSION: This study showed an association between ADHD and a higher risk of central precocious puberty. Early referral of children with ADHD to a pediatric endocrinologist for evaluation may facilitate correct diagnoses and early interventions. IMPACT: ADHD is associated with a higher risk of central precocious puberty. This study provides relevant findings, as it is the first nationwide, population-based cohort study to investigate the association between ADHD and the risk of central precocious puberty with a 15-year follow-up. Early referral of children with ADHD to a pediatric endocrinologist for the evaluation of suspected precocious puberty could facilitate correct diagnosis. Early intervention treatment with gonadotropin-releasing hormone agonist might improve final height in children with central precocious puberty.

Factors Influencing the Reputation of Assistive Technology Resources Center: An Example from Yunlin County, Taiwan.

By the end of 2020, a total of 34 multifunctional Assistive Technology Resources Centers (ATR centers) were set up in 22 counties and cities in Taiwan. This study examines the perceptions of the users and their caregivers of the government-established ATR centers in Taiwan and examines the impact on the reputation of the public institution. The research framework and hypotheses were developed by examining the factors of "service convenience", "center-related factors", "justice", and "perceived value" and using "perceived value" as a mediating variable. The data were collected through a questionnaire survey, and the structural equation model was used to test the model and verify the hypotheses. Research data was collected in various townships in Yunlin County A total of 320 questionnaires were collected. Of these respondents, 22% were aged 51-60. All the research hypotheses were positively and significantly verified. Of these, justice was the most important factor affecting the value of the ATR center's services compared to convenience and center-related factors. Of convenience, service value and justice, service value was the most important factor affecting the perceived reputation of the public institution. According to the findings of this study, it is beyond expectation that the convenience of ATR is not the main factor influencing the service value, but rather the perceived justice is the most important factor. Therefore, ATR should be prioritized from the perspective of the service recipient, especially the perceived justice of the service, in order to best enhance the value of the service and improve the reputation of the public institution.

Implantation of monocusp valve prolongs the duration of chest tube drainage in children with tetralogy of fallot after corrective surgery.

BACKGROUND: Right ventricular outflow tract obstruction relief is one of the major procedures during the total correction of tetralogy of Fallot (TOF). Pulmonary insufficiency (PI) is usually inevitable after a transannular incision with a patch repair is performed. Therefore, some surgeons advocate to place a monocusp valve within the transannular patch (TAP) in order to decrease the severity of the PI. However, the monocusp valve seemed not be very effective in some patients who underwent the complete TOF repair. METHODS: Patients who had the classic form of TOF between January 2009 and January 2017 and underwent the corrective surgery with a TAP by the same cardiovascular surgeon were identified for further analysis. Clinical information including demographics at operation, perioperative data, and postoperative outcome were collected retrospectively and compared between the group with and without a monocusp valve. RESULTS: A total of 24 TOF cases were included in the final analysis, and 16 (66.7%) patients received a monocusp valve placement. The patients' characteristics before and during the surgery were similar between the two groups. The median duration of chest tube drainage after the total correction in the monocusp group was longer than those without the valve (p = 0.04). There was no difference in the immediate postoperative data, including the inflammation/infection status, the duration of mechanical ventilation, and the length of ICU and hospital stay. CONCLUSION: Implantation of a monocusp valve during the total TOF correction using a TAP did not bring benefit to improve the immediate postoperative outcomes, especially the duration of the pleural drainage. Further study with a prospective design and a larger number of cases is needed.

Baicalein ameliorates ulcerative colitis by improving intestinal epithelial barrier via AhR/IL-22 pathway in ILC3s.

Ulcerative colitis (UC) is a chronic inflammatory disease of the gastrointestinal tract, which is closely related to gut barrier dysfunction. Emerging evidence shows that interleukin-22 (IL-22) derived from group 3 innate lymphoid cells (ILC3s) confers benefits on intestinal barrier, and IL-22 expression is controlled by aryl hydrocarbon receptor (AhR). Previous studies show that baicalein protects the colon from inflammatory damage. In this study we elucidated the molecular mechanisms underlying the protective effect of baicalein on intestinal barrier function in colitis mice. Mice were administered baicalein (10, 20, 40 mg·kg-1·d-1, i.g.) for 10 days; the mice freely drank 3% dextran sulfate sodium (DSS) on D1-D7 to induce colitis. We showed that baicalein administration simultaneously ameliorated gut inflammation, decreased intestinal permeability, restored tight junctions of colons possibly via promoting AhR/IL-22 pathway. Co-administration of AhR antagonist CH223191 (10 mg/kg, i.p.) partially blocked the therapeutic effects of baicalein in colitis mice, whereas AhR agonist FICZ (1 µg, i.p.) ameliorated symptoms and gut barrier function in colitis mice. In a murine lymphocyte line MNK-3, baicalein (5-20 µM) dose-dependently increased the expression of AhR downstream target protein CYP1A1, and enhanced IL-22 production through facilitating AhR nuclear translocation, these effects were greatly diminished in shAhR-MNK3 cells, suggesting that baicalein induced IL-22 production in AhR-dependent manner. To further clarify that, we constructed an in vitro system consisting of MNK-3 and Caco-2 cells, in which MNK-3 cell supernatant treated with baicalein could decrease FITC-dextran permeability and promoted the expression of tight junction proteins ZO-1 and occluding in Caco-2 cells. In conclusion, this study demonstrates that baicalein ameliorates colitis by improving intestinal epithelial barrier via AhR/IL-22 pathway in ILC3s, thus providing a potential therapy for UC.

Association Between Tetralogy of Fallot and Psychiatric Disorders: A Nationwide Cohort Study.

OBJECTIVE: The tetralogy of Fallot (TOF) has been reported to be associated with some neurodevelopmental impairment and psychiatric disorders. Nevertheless, a nationwide study to clarify the risk between TOF and comorbid psychiatric disorders is lacking. Using a nationwide database in Taiwan, this study aimed to explore the role of TOF in various psychiatric disorders and analyze whether there are patient-related risk factors. METHODS: A total of 16,824 enrolled patients, including 4,206 study subjects who were diagnosed with TOF and 12,618 controls with TOF matched (1:3) for sex, age, hospital visits, and index year, were randomly selected from the Taiwanese National Health Insurance Research Database (NHIRD) between 2000 and 2015. Patients' diagnoses in the NHIRD were encoded using International Classification of Diseases, 9th Revision, Clinical Modification codes. RESULTS: Of patients with TOF, 256 (6.09%) developed psychiatric disorders compared to 394 (3.12%) in the control group. After adjusting for covariates, the adjusted hazard ratio of psychiatric disorders for patients with TOF was 3.192 (95% CI, 2.683-3.798; P < .001). After exclusion of psychiatric diagnoses within the first 5 years, TOF was associated with an increased risk of anxiety (P < .001), depression (P < .001), bipolar disorder (P < .001), and sleep disorders (P = .005). CONCLUSIONS: This study revealed that TOF patients have a nearly 3-fold higher risk of psychiatric disorders, including anxiety, depressive, bipolar, and sleep disorders, than the general population. Therefore, continued mental health screening and surveillance are warranted in TOF patients.

A Novel Multiprotein Bridging Factor 1-Like Protein Induces Cyst Wall Protein Gene Expression and Cyst Differentiation in Giardia lamblia.

The capacity to synthesize a protective cyst wall is critical for infectivity of Giardia lamblia. It is of interest to know the mechanism of coordinated synthesis of three cyst wall proteins (CWPs) during encystation, a differentiation process. Multiprotein bridging factor 1 (MBF1) gene family is a group of transcription coactivators that bridge various transcription factors. They are involved in cell growth and differentiation in yeast and animals, or in stress response in fungi and plants. We asked whether Giardia has MBF1-like genes and whether their products influence gene expression. BLAST searches of the Giardia genome database identified one gene encoding a putative MBF1 protein with a helix-turn-helix domain. We found that it can specifically bind to the AT-rich initiator promoters of the encystation-induced cwp1-3 and myb2 genes. MBF1 localized to cell nuclei and cytoplasm with higher expression during encystation. In addition, overexpression of MBF1 induced cwp1-3 and myb2 gene expression and cyst generation. Mutation of the helixes in the helix-turn-helix domain reduced cwp1-3 and myb2 gene expression and cyst generation. Chromatin immunoprecipitation assays confirmed the binding of MBF1 to the promoters with its binding sites in vivo. We also found that MBF1 can interact with E2F1, Pax2, WRKY, and Myb2 transcription factors that coordinately up-regulate the cwp genes during encystation. Using a CRISPR/Cas9 system for targeted disruption of mbf1 gene, we found a downregulation of cwp1-3 and myb2 genes and decrease of cyst generation. Our results suggest that MBF1 is functionally conserved and positively regulates Giardia cyst differentiation.

Baicalin prevents LPS-induced activation of TLR4/NF-κB p65 pathway and inflammation in mice via inhibiting the expression of CD14.

Previous studies have shown that baicalin, an active ingredient of the Chinese traditional medicine Huangqin, attenuates LPS-induced inflammation by inhibiting the activation of TLR4/NF-κBp65 pathway, but how it affects this pathway is unknown. It has been shown that CD14 binds directly to LPS and plays an important role in sensitizing the cells to minute quantities of LPS via chaperoning LPS molecules to the TLR4/MD-2 signaling complex. In the present study we investigated the role of CD14 in the anti-inflammatory effects of baicalin in vitro and in vivo. Exposure to LPS (1 µg/mL) induced inflammatory responses in RAW264.7 cells, evidenced by marked increases in the expression of MHC II molecules and the secretion of NO and IL-6, and by activation of MyD88/NF-κB p65 signaling pathway, as well as the expression of CD14 and TLR4. These changes were dose-dependently attenuated by pretreatment baicalin (12.5-50 µM), but not by baicalin post-treatment. In RAW264.7 cells without LPS stimulation, baicalin dose-dependently inhibit the protein and mRNA expression of CD14, but not TLR4. In RAW264.7 cells with CD14 knockdown, baicalin pretreatment did not prevent inflammatory responses and activation of MyD88/NF-κB p65 pathway induced by high concentrations (1000 µg/mL) of LPS. Furthermore, baicalin pretreatment also inhibited the expression of CD14 and activation of MyD88/NF-κB p65 pathway in LPS-induced hepatocyte-derived HepG2 cells and intestinal epithelial-derived HT-29 cells. In mice with intraperitoneal injection of LPS and in DSS-induced UC mice, oral administration of baicalin exerted protective effects by inhibition of CD14 expression and inflammation. Taken together, we demonstrate that baicalin pretreatment prevents LPS-induced inflammation in RAW264.7 cells in CD14-dependent manner. This study supports the therapeutic use of baicalin in preventing the progression of LPS-induced inflammatory diseases.

Social Image Impacting Attitudes of Middle-Aged and Elderly People toward the Usage of Walking Aids: An Empirical Investigation in Taiwan.

The elderly need the assistance of walking aids due to deterioration of their physical functions. However, they are often less willing to use these aids because of their worries about how others may think of them. Not using professional walking aids often makes elderly people fall easily when walking. This study explores the behavioral intention factors of middle-aged people (45-64 years old) and elderly people (65 years and older) that affect the use of walking aids. Based on the Theory of Reasoned Action (TRA), subjective norms, attitude toward usage, behavior intention, safety, and usefulness were combined with social image to establish the research framework. This study used questionnaire surveys both in paper form assisted by volunteers and in online electronic form. A total of 457 questionnaires were collected. Data analysis was carried out in three stages: descriptive analysis, measurement model verification, and structural equation model analysis. The results showed that social image had a significant impact on the attitude toward using walking aids. Factors such as attitude toward usage, subjective norms, and safety of walking aids also had a significant positive impact on behavioral intention. Finally, through the research results, some suggestions are proposed for stakeholders to improve the elderly's concerns about the social image of using walking aids.

Diagnosis and Surgical Outcomes of Patients with Anomalous Left Coronary Artery from the Pulmonary Artery: A Single Taiwanese Medical Center Experience.

BACKGROUND: Anomalous left coronary artery from the pulmonary artery (ALCAPA), a very rare congenital cardiac anomaly, is associated with a high mortality rate among infants who are not diagnosed or treated in a timely manner. Surgical intervention with the reconstruction for a two-coronary-system circulation is the main treatment; however, there have been very few reported cases from Taiwan. In this study, we aim to describe the clinical manifestations, diagnostic methods, surgery types, and surgical outcomes in patients with ALCAPA from a single Taiwanese medical center. METHODS: We retrospectively reviewed patients diagnosed with ALCAPA who underwent surgery at our institution between January 2001 and October 2018. Clinical presentations, noninvasive and invasive study results, surgical methods, and postoperative follow-up results were assessed from medical records. Moreover, literature on this particular cardiovascular anomaly was reviewed. RESULTS: The study included 6 patients (5 children and 1 adult). The diagnosis was confirmed using cardiac catheterization and coronary angiography in four patients and only echocardiography in two patients. All patients underwent surgical correction and survived. Four patients showed improvements in left ventricular function and mitral regurgitation (MR). CONCLUSION: Early diagnosis and timely surgical intervention could avoid mortality regardless of the method of operation. ALCAPA can be definitively diagnosed using noninvasive echocardiography. Both left ventricular systolic function and mitral insufficiency could improve after the surgical intervention in pediatric patients. Repair or replacement of the mitral valve could be reserved for persistent MR complicated with congestive heart failure, particularly in patients who received the initial operation beyond infancy.

Oroxindin inhibits macrophage NLRP3 inflammasome activation in DSS-induced ulcerative colitis in mice via suppressing TXNIP-dependent NF-κB pathway.

Oroxindin is a flavonoid isolated from the traditional Chinese medicine Huang-Qin, which has shown various pharmacological activities including anti-inflammatory, antitumor, antioxidant, etc. Thus far, the effect of oroxindin on colonic inflammation and the underlying mechanism remain unknown. In this study, we investigated the tissue distribution of oroxindin and its therapeutic effects on ulcerative colitis (UC) as well as the underlying mechanisms. UC model was established in mice by administrating dextran sulfate sodium (DSS) in drinking water for 7 d. We first showed that oroxindin was largely absorbed by the colon as an active ingredient after normal mice received Huang-Qin-Tang, a traditional Chinese medicine decoction. UC mice were then treated with oroxindin (12.5, 25, 50 mg ·kg-1 ·d-1, i.g.) for 10 d. We found that oroxindin treatment greatly suppressed massive macrophages infiltration and attenuated pathological changes in colonic tissue. Furthermore, oroxindin treatment significantly inhibited the generation of IL-1ß and IL-18 in the colon via inhibiting the nucleotide-binding oligomerization domain-like receptor 3 (NLRP3) inflammasome formation and activation. In cultured macrophages, LPS induced NLRP3 inflammasome formation and caspase-1 activation, which were suppressed by oroxindin (12.5-50 µM). In LPS-treated macrophages, oroxindin dose-dependently restored the expression of TXNIP protein, leading to suppressing TXNIP-dependent NF-κB activation. In conclusion, these results demonstrate that oroxindin could be absorbed by the colon and attenuate inflammatory responses via inhibiting NLRP3 inflammasome formation and activation, which is related to the inhibitory effect on TXNIP-dependent NF-κB-signaling pathway. Hence, oroxindin has the potential of becoming an effective drug for treating UC.

EXIA2: web server of accurate and rapid protein catalytic residue prediction.

We propose a method (EXIA2) of catalytic residue prediction based on protein structure without needing homology information. The method is based on the special side chain orientation of catalytic residues. We found that the side chain of catalytic residues usually points to the center of the catalytic site. The special orientation is usually observed in catalytic residues but not in noncatalytic residues, which usually have random side chain orientation. The method is shown to be the most accurate catalytic residue prediction method currently when combined with PSI-Blast sequence conservation. It performs better than other competing methods on several benchmark datasets that include over 1,200 enzyme structures. The areas under the ROC curve (AUC) on these benchmark datasets are in the range from 0.934 to 0.968.

CELLO2GO: a web server for protein subCELlular LOcalization prediction with functional gene ontology annotation.

CELLO2GO (http://cello.life.nctu.edu.tw/cello2go/) is a publicly available, web-based system for screening various properties of a targeted protein and its subcellular localization. Herein, we describe how this platform is used to obtain a brief or detailed gene ontology (GO)-type categories, including subcellular localization(s), for the queried proteins by combining the CELLO localization-predicting and BLAST homology-searching approaches. Given a query protein sequence, CELLO2GO uses BLAST to search for homologous sequences that are GO annotated in an in-house database derived from the UniProt KnowledgeBase database. At the same time, CELLO attempts predict at least one subcellular localization on the basis of the species in which the protein is found. When homologs for the query sequence have been identified, the number of terms found for each of their GO categories, i.e., cellular compartment, molecular function, and biological process, are summed and presented as pie charts representing possible functional annotations for the queried protein. Although the experimental subcellular localization of a protein may not be known, and thus not annotated, CELLO can confidentially suggest a subcellular localization. CELLO2GO should be a useful tool for research involving complex subcellular systems because it combines CELLO and BLAST into one platform and its output is easily manipulated such that the user-specific questions may be readily addressed.

Reevaluating reference ranges of oxygen saturation for healthy full-term neonates using pulse oximetry.

BACKGROUND: We compared our clinical experience with currently available reference oxygen saturation level (SpO(2)) values from the American Academy of Pediatrics/American Heart Association (AAP/AHA) neonatal resuscitation program guidelines. METHODS: We enrolled 145 healthy full-term neonates; infants showing respiratory distress and those with serious congenital anomalies were excluded. SpO(2) values at every 1 minute until 10 minutes after birth were measured and recorded. Infants were classified into the cesarean section (CS) and normal spontaneous delivery (NSD) groups for evaluating differences. The 10(th) percentiles of SpO(2) at each minute were used as the lower limits of normal oxygen saturation, and these were compared with the lowest target values recommended in the AAP/AHA guidelines. RESULTS: Overall, 130 vigorous full-term neonates (median gestational age: 38 5/7 weeks; body weight at birth: 2405-3960 g) were analyzed. The median SpO(2) were 67% and 89% at the 1(st) and 4(th) minute, respectively. On average, SpO(2) values reached >90% at the 5(th) minute. No statistical differences were noted in the SpO(2) values between the CS and NSD groups after 5 minutes; however, a trend of higher SpO(2) was observed in the NSD group. We noted a gradually increasing trend for SpO(2) values over time, similar to that noted in the AAP/AHA guidelines. However, SpO(2) values at the 10(th) percentiles of each minute within the first 5 minutes in our study were equal to or significantly lower than those in the AAP/AHA guidelines; moreover, at the 10(th) minute, SpO(2) values at the 10(th) percentiles were significantly higher than those in the guidelines. CONCLUSION: The delivery modes did not affect the SpO(2) values of full-term healthy neonates. Discrepancies in SpO(2) changes in full-term neonates not requiring resuscitation between this study and the AAP/AHA guidelines were significant. SpO(2) ranges for each time point within the first 10 minutes after birth should therefore be reevaluated locally.

On the structural context and identification of enzyme catalytic residues.

Enzymes play important roles in most of the biological processes. Although only a small fraction of residues are directly involved in catalytic reactions, these catalytic residues are the most crucial parts in enzymes. The study of the fundamental and unique features of catalytic residues benefits the understanding of enzyme functions and catalytic mechanisms. In this work, we analyze the structural context of catalytic residues based on theoretical and experimental structure flexibility. The results show that catalytic residues have distinct structural features and context. Their neighboring residues, whether sequence or structure neighbors within specific range, are usually structurally more rigid than those of noncatalytic residues. The structural context feature is combined with support vector machine to identify catalytic residues from enzyme structure. The prediction results are better or comparable to those of recent structure-based prediction methods.

Accurate prediction of protein catalytic residues by side chain orientation and residue contact density.

Prediction of protein catalytic residues provides useful information for the studies of protein functions. Most of the existing methods combine both structure and sequence information but heavily rely on sequence conservation from multiple sequence alignments. The contribution of structure information is usually less than that of sequence conservation in existing methods. We found a novel structure feature, residue side chain orientation, which is the first structure-based feature that achieves prediction results comparable to that of evolutionary sequence conservation. We developed a structure-based method, Enzyme Catalytic residue SIde-chain Arrangement (EXIA), which is based on residue side chain orientations and backbone flexibility of protein structure. The prediction that uses EXIA outperforms existing structure-based features. The prediction quality of combing EXIA and sequence conservation exceeds that of the state-of-the-art prediction methods. EXIA is designed to predict catalytic residues from single protein structure without needing sequence or structure alignments. It provides invaluable information when there is no sufficient or reliable homology information for target protein. We found that catalytic residues have very special side chain orientation and designed the EXIA method based on the newly discovered feature. It was also found that EXIA performs well for a dataset of enzymes without any bounded ligand in their crystallographic structures.

Design, synthesis and fungicidal activity of novel sclerotiorin derivatives.

Sclerotiorin, a chlorine-containing azaphilone-type natural product, was first isolated from Penicillium sclerotiorum and has been reported to exhibit weak fungicidal activity. Optimization of the substituents at the 3- and 5-positions of the sclerotiorin framework was investigated with the aim of discovering novel fungicides with improved activity. The design of sclerotiorin analogues involved replacing the diene side chain with a phenyl group or an aromatic- or heteroaromatic-containing aliphatic side chain. The designed compounds were synthesized by cycloisomerization and subsequent oxidation of suitable 2-alkynylbenzaldehydes, in which a variety of substituents were introduced using a Sonogashira coupling reaction. The structures of these newly prepared compounds were confirmed by 1H and 13C NMR spectroscopy, HRMS and single-crystal X-ray analysis. The antifungal activity of the synthesized compounds was evaluated against seven phytopathogenic species. Compounds 3, 9g and 9h were found to have a broad spectrum of fungicidal activity, and these structurally simpler products can be recognized as lead compounds for further optimization.

Synthesis and antifungal activity of novel sclerotiorin analogues.

Sclerotiorin 1, first isolated from Penicillium sclerotiorum, has weak antifungal activity and belongs to the azaphilone-type family of natural products. Several series of sclerotiorin analogues were designed and synthesized with the aim of discovering novel fungicides with improved activity. The syntheses involved two key steps, cycloisomerization and then oxidation, and used a simple and efficient Sonogashira cross-coupling reaction to construct the required functionalized precursor. With sclerotiorin as a control, the activities of the newly synthesized analogues were evaluated against seven fungal pathogens, and several promising candidates (compounds 3a1, 3d2, 3e2, 3f2 and 3k2) with greater activity and simpler structures than sclerotiorin were discovered. In addition, preliminary structure-activity relationships were studied, which revealed that not only the chlorine or bromine substituent at the 5-position of the nucleus but also the phenyl group at the 3-position and the substituent pattern on it contributed crucially to the observed antifungal activity. Analogues with a methyl substituent at the 1-position have reduced levels of activity, while those with a free hydroxyl group in place of acetoxy at the quaternary center of the bicyclic ring system retain activity.

A novel E2F-like protein involved in transcriptional activation of cyst wall protein genes in Giardia lamblia.

Giardia lamblia differentiates into resistant walled cysts for survival outside the host and transmission. During encystation, synthesis of cyst wall proteins is coordinately induced. The E2F family of transcription factors in higher eukaryotes is involved in cell cycle progression and cell differentiation. We asked whether Giardia has E2F-like genes and whether they influence gene expression during Giardia encystation. Blast searches of the Giardia genome database identified one gene (e2f1) encoding a putative E2F protein with two putative DNA-binding domains. We found that the e2f1 gene expression levels increased significantly during encystation. Epitope-tagged E2F1 was found to localize to nuclei. Recombinant E2F1 specifically bound to the thymidine kinase and cwp1-3 gene promoters. E2F1 contains several key residues for DNA binding, and mutation analysis revealed that its binding sequence is similar to those of the known E2F family proteins. The E2F1-binding sequences were positive cis-acting elements of the thymidine kinase and cwp1 promoters. We also found that E2F1 transactivated the thymidine kinase and cwp1 promoters through its binding sequences in vivo. Interestingly, E2F1 overexpression resulted in a significant increase of the levels of CWP1 protein, cwp1-3 gene mRNA, and cyst formation. We also found E2F1 can interact with Myb2, a transcription factor that coordinate up-regulates the cwp1-3 genes during encystation. Our results suggest that E2F family has been conserved during evolution and that E2F1 is an important transcription factor in regulation of the Giardia cwp genes, which are key to Giardia differentiation into cysts.