Gene Expression Profile Analysis of the Molecular Mechanism of HOXD10 Regulation of Epithelial Ovarian Cancer Cells.

Epithelial ovarian cancer (EOC) is the most common type of ovarian cancer. Although studies have reported that downregulation of HOXD10 expression may contribute to the migration and invasion abilities in EOC, much about its regulation remains to be fully elucidated. The present study aimed to identify different gene expression profiles associated with HOXD10 overexpression in EOC cells. The present study confirmed that HOXD10 overexpression effectively inhibited the proliferation and motility of the TOV21G and TOV112D cells. Further, we overexpress HOXD10 in TOV112D cells, the different gene expression (DEGs) profiles induce by HOXD10 was analyze by the Human OneArray microarray. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG), ingenuity pathway analysis (IPA) was used to perform the pathway enrichment analysis for the DEGs. Integrated bioinformatics analysis showed that the DEGs were enriched for terms related to oxidative phosphorylation and mitochondrial function pathways. Dysfunction oxidative phosphorylation metabolic pathway occurs frequently in many tumors. We validated the expression of NDUFA7, UQCRB and CCL2 using qPCR, involving in metabolism-related pathway, were significantly changed by HOXD10 overexpression in EOC. The detailed regulatory mechanism that links HOXD10 and the oxidative phosphorylation genes is not yet fully understood, our findings provide novel insight into HOXD10-mediated pathways and their effects on cancer metabolism, carcinogenesis, and the progression of EOC. Thus, the data suggest that strategies to interfere with metabolism-related pathways associated with cancer drug resistance could be considered for the treatment of ovarian tumors.

Spatial and temporal relationship between native mammals and free-roaming dogs in a protected area surrounded by a metropolis.

With rapid urbanization worldwide, anthropogenic impacts such as human settlements and invasive carnivores (dogs Canis familiaris, cats Felis catus) are altering spatial distributions and temporal activity patterns of native species. In this study, we focused on spatiotemporal responses of native mammals to anthropogenic impacts in a protected area surrounded by a large metropolis (i.e. Yangmingshan National Park inside Taipei-Keelung metropolis in northern Taiwan). We collected site use data of 11 mammal species (i.e. dogs, cats, nine native species) between 2012 and 2017 with a camera system comprising 121 camera sites. We quantified anthropogenic disturbances as distance to human settlements and activity levels of free-roaming dogs and cats. Species richness and occurrences of the native mammals increased with increasing distances to human settlements and decreasing activity level of dogs, with the latter having a stronger effect than the former. Diel activity overlap between native mammals and dogs was lower during April-July season, coinciding with the breeding season for several native mammals. In contrast, activity level of cats showed no relationships with species richness, occurrences or diel activities of the native mammals. This study demonstrated negative impacts of human settlements and free-roaming dogs on native mammal communities for protected areas in urban environments, and highlights dog activity as a major anthropogenic threat to wildlife.

Demography and welfare status of free-roaming dogs in Yangmingshan National Park, Taiwan.

Free-roaming dogs (Canis familiaris) cause threats to native wildlife and public health and raise concerns for their welfare. Understanding the demography of free-roaming dog populations is essential for developing an effective management plan. An evaluation of their welfare status would be beneficial to earn public support for the management plan. In this study, we estimated the population size, survivorship, and health of a free-roaming dog population in Yangmingshan National Park (YMSNP), Taiwan, during 2016-2018. YMSNP is a rural area with human settlements but also a protected area of conservation concern. We identified 191, 176, 216 individuals at our sampling sites in 2016, 2017, and 2018, respectively. Using a photographic capture-recapture method and extrapolation, we estimated that there were 786-979 dogs in the park during this 3-year period. The annual apparent survival rate of identified dogs was 16.7% for 2016-2017 and 23.9% for 2017-2018. The dogs had a high rate of lameness and dermatosis of 5.1-8.8% and 14.2-18.1%, respectively. Thirty-five blood samples showed that 34.3% of the dogs were anemic, 37.1% showed abnormal white blood cell counts, and 68.6% exhibited abnormal platelet counts. These results suggested that the dogs were at high density with low survivorship and in poor health, and new individuals entered the population continuously. Interventions to manage this dog population and to improve their welfare must be carried out. Our study provides an example for monitoring and managing a free-roaming dog population in a rural, conservation area in Southeast Asia.

Neuroprotective and Antineuroinflammatory Effects of Hydroxyl-Functionalized Stilbenes and 2-Arylbenzo[b]furans.

The drugs currently used to treat Alzheimer's disease (AD) are limited in the benefits they confer, and no medication has been clearly proven to cure or delay the progression of AD. Most candidate AD drugs are meant to reduce the production, aggregation, and toxicity of amyloid ß (Aß) or to promote Aß clearance. Herein, we demonstrate the efficient synthesis of hydroxyl-functionalized stilbene and 2-arylbenzo[b]furan derivatives and report on the neuroprotective and anti-inflammatory effects of these phenolic compounds in vitro and in an animal model. Structure-activity relationships revealed that the presence of an acrylate group on 2-arylbenzo[b]furan confers neuroprotective and anti-inflammatory effects. Furthermore, compounds 11 and 37 in this study showed particular potential for development as disease-modifying anti-Alzheimer's drugs, based on their neuroprotective effects on neuron cells, their antineuroinflammatory effects on glial cells, and the ability to ameliorate nesting behavior in APP/PS1 mice. These results indicate that 2-arylbenzo[b]furans could be candidate compounds for the treatment of AD.

Observation of Surface Coverage-Dependent Surface-Enhanced Raman Scattering and the Kinetic Behavior of Methylene Blue Adsorbed on Silver Oxide Nanocrystals.

We report the study of surface-enhanced Raman scattering (SERS) and photocatalytic reactions of methylene blue (MB) adsorbed on cubic silver oxide (c-Ag2O) nanocrystals. The SERS activities of MB observed on c-Ag2O are surface coverage-dependent. According to the SERS selection rules, the strong intensities of the C-S-C and C-N-C bending modes imply that the MB molecules are adsorbed on the c-Ag2O surface with a near-perpendicular position. On the other hand, if these modes are weak, MB assumes a near-parallel position to the c-Ag2O surface. Under the "perpendicular" condition, the photocatalytic reaction rate of MB observed on c-Ag2O surfaces is first order at a high surface coverage, whereas under the "parallel" condition, the observed rate is zero order at a low surface coverage. The SERS intensity trends and the kinetic behavior of MB on c-Ag2O are correlated.

Differential Effects of Roads and Traffic on Space Use and Movements of Native Forest-Dependent and Introduced Edge-Tolerant Species.

Anthropogenic infrastructure such as roads and non-native species are major causes of species endangerment. Understanding animal behavioral responses to roads and traffic provides insight into causes and mechanisms of effects of linear development on wildlife and aids effective mitigation and conservation. We investigated effects of roads and traffic on space use and movements of two forest-dwelling species: endemic, forest-dependent Mount Graham red squirrels (Tamiasciurus hudsonicus grahamensis) and introduced, edge-tolerant Abert's squirrels (Sciurus aberti). To assess the effects of roads on space use and movement patterns, we compared the probability that a squirrel home range included roads and random lines in forests, and assessed effects of traffic intensity on rate of road crossing and movement patterns. Red squirrels avoided areas adjacent to roads and rarely crossed roads. In contrast, Abert's squirrels were more likely to include roads in their home ranges compared to random lines in forests. Both red squirrels and Abert's squirrels increased speed when crossing roads, compared to before and after road crossings. Increased hourly traffic volume reduced the rate of road crossings by both species. Behavioral responses of red squirrels to roads and traffic resemble responses to elevated predation risk, including reduced speed near roads and increased tortuosity of movement paths with increased traffic volume. In contrast, Abert's squirrels appeared little affected by roads and traffic with tortuosity of movement paths reduced as distance to roads decreased. We found that species with similar body size category (<1 kg) but different habitat preference and foraging strategy responded to roads differently and demonstrated that behavior and ecology are important when considering effects of roads on wildlife. Our results indicate that roads restricted movements and space use of a native forest-dependent species while creating habitat preferred by an introduced, edge-tolerant species.

Characterization of glycine substitution mutations within the putative NAD+-binding site of Bacillus licheniformis aldehyde dehydrogenase.

The NAD(+)-requiring enzymes of the aldehyde dehydrogenase (ALDH) family contain a glycine motif, GX1- 2GXXG, which is reminiscent of the fingerprint region of the Rossman fold, a conserved structural motif of the classical nicotinamide nucleotide-binding proteins. In this research, the role of three glycine residues situated within the putative NAD(+)-binding motif (211-GPGSSAG) together with Gly233 and Gly238 of Bacillus licheniformis ALDH (BlALDH) were probed by site-directed mutatgenesis. Fifteen mutant BlALDHs were obtained by substitution of the indicated glycine residues with alanine, glutamate and arginine. Except for the Ala replacement at positions 211, 213, 217 and 238, the remaining mutant enzymes lost the dehydrogenase activity completely. Tryptophan fluorescence and far-UV circular dichroism spectra allowed us to discriminate BlALDH and the inactive mutant enzymes, and unfolding analyses further revealed that they had a different sensitivity towards temperature- and guanidine hydrochloride (GdnHCl)-induced denaturation. BlALDH and the functional variants had a comparable T(m) value, but the value was reduced by more than 5.1°C in the rest of mutant enzymes. Acrylamide quenching analysis showed that the inactive mutant enzymes had a dynamic quenching constant greater than that of BlALDH. Native BlALDH started to unfold beyond ~0.21 M GdnHCl and reached an unfolded intermediate, [GdnHCl](0.5, N-U), at 0.92 M equivalent to free energy change (ΔG(N-U)(H2O)) of 12.34 kcal/mol for the N → U process, whereas the denaturation midpoints for mutant enzymes were 0.45-1.61 M equivalent to ΔG(N-U)(H2O) of 0.31-4.35 kcal/mol. Taken together, these results strongly suggest that the explored glycines are indeed important for the catalytic activity and structural stability of BlALDH.

Contribution of conserved Glu255 and Cys289 residues to catalytic activity of recombinant aldehyde dehydrogenase from Bacillus licheniformis.

Based on the sequence homology, we have modeled the three-dimensional structure of Bacillus licheniformis aldehyde dehydrogenase (BlALDH) and identified two different residues, Glu255 and Cys289, that might be responsible for the catalytic function of the enzyme. The role of these residues was further investigated by site-directed mutagenesis and biophysical analysis. The expressed parental and mutant proteins were purified by nickel-chelate chromatography, and their molecular masses were determined to be approximately 53 kDa by SDS-PAGE. As compared with the parental BlALDH, a dramatic decrease or even complete loss of the dehydrogenase activity was observed for the mutant enzymes. Structural analysis showed that the intrinsic fluorescence and circular dichroism spectra of the mutant proteins were similar to the parental enzyme, but most of the variants exhibited a different sensitivity towards thermal- and guanidine hydrochloride-induced denaturation. These observations indicate that residues Glu255 and Cys289 play an important role in the dehydrogenase activity of BlALDH, and the rigidity of the enzyme has been changed as a consequence of the mutations.

Biophysical studies of an NAD(P)(+)-dependent aldehyde dehydrogenase from Bacillus licheniformis.

Aldehyde dehydrogenase (ALDH) catalyzes the conversion of aldehydes to the corresponding acids by means of an NAD(P)(+)-dependent virtually irreversible reaction. In this investigation, the biophysical properties of a recombinant Bacillus licheniformis ALDH (BlALDH) were characterized in detail by analytical ultracentrifuge (AUC) and various spectroscopic techniques. The oligomeric state of BlALDH in solution was determined to be tetrameric by AUC. Far-UV circular dichroism analysis revealed that the secondary structures of BlALDH were not altered in the presence of acetone and ethanol, whereas SDS had a detrimental effect on the folding of the enzyme. Thermal unfolding of this enzyme was found to be highly irreversible. The native enzyme started to unfold beyond ~0.2 M guanidine hydrochloride (GdnHCl) and reached an unfolded intermediate, [GdnHCl](05, N-U), at 0.93 M. BlALDH was active at concentrations of urea below 2 M, but it experienced an irreversible unfolding under 8 M denaturant. Taken together, this study provides a foundation for the future structural investigation of BlALDH, a typical member of ALDH superfamily enzymes.

Risk factors for delayed diagnosis of scabies in hospitalized patients from long-term care facilities.

BACKGROUND: Delayed diagnosis of scabies can cause an institutional outbreak, which causes considerably economic burden to control. This study was to find the risk factors for delayed diagnosis of scabies in hospitalized patients from long-term care facilities. METHODS: We conducted a retrospective analysis of the hospitalized patients from long-term care facilities, diagnosed to have scabies between January 2006 and December 2008. A stepwise logistic regression analysis was performed to determine the risk factors for delayed diagnosis of scabies. RESULTS: A total of 706 episodes with scabies were identified retrospectively in 399 hospitalized patients from long-term care facilities. Of these, 44 episodes were considered as delayed diagnosis of scabies. These patients were more associated with chronic usage of steroid (73% vs. 10%, P < 0.001) and had longer duration of hospitalization than the others (30 vs. 13 days, P < 0.001). After logistic regression, steroid therapy was the risk factor of delayed diagnosis of scabies (odds ratio: 23.493). CONCLUSIONS: In the patients from long-term care facilities, clinical physicians should pay more attention to those with chronic usage of steroid to avoid delayed diagnosis of scabies. KEYWORDS: Scabies; Delayed diagnosis; Risk factor; Long-term care facility.

Biophysical characterization of Bacillus licheniformis and Escherichia coli γ-glutamyltranspeptidases: A comparative analysis.

The oligomeric states of Bacillus licheniformis and Escherichia coli γ-glutamyltranspeptidases (BlGGT and EcGGT) in solution have been investigated by analytical ultracentrifugation. The results showed that BlGGT has a sedimentation coefficient of 5.12S, which can be transformed into an experimental molecular mass of approximately 62,680Da. The monomeric conformation is conserved in EcGGT. SDS-PAGE analysis and cross-linking studies further proved that the autocatalytically processed BlGGT and EcGGT form a heterodimeric association. Unfolding analyses using circular dichroism and tryptophan emission fluorescence revealed that these two proteins had a different sensitivity towards temperature- and guanidine hydrochloride (GdnHCl)-induced denaturation. BlGGT and EcGGT had a T(m) value of 59.5 and 49.2°C, respectively, and thermal unfolding of both proteins was found to be highly irreversible. Chemical unfolding of BlGGT was independent to the pH value ranging from 5 to 10, whereas the pH environment was found to significantly influence the GdnHCl-induced denaturation of EcGGT. Both enzymes did not reactivate from the completely unfolded states, accessible at 6M GdnHCl. BlGGT was active in the presence of 4M NaCl, whereas the activity of EcGGT was significantly decreased at the high-salt condition. Taken together, these findings suggest that the biophysical properties of the homologous GGTs from two mesophilic sources are quite different.

Biophysical characterization of a recombinant α-amylase from thermophilic Bacillus sp. strain TS-23.

Environmental variables can significantly influence the folding and stability of a protein molecule. In the present study, the biophysical properties of a truncated Bacillus sp. TS-23 α-amylase (BACΔNC) were characterized in detail by glutaraldehyde cross-linking, analytical ultracentrifugation, and various spectroscopic techniques. With cross-linking experiment and analytical ultracentrifuge, we demonstrated that the oligomeric state of BACΔNC in solution is monomeric. Far-UV circular dichroism analysis revealed that the secondary structures of BACΔNC were significantly altered in the presence of various metal ions and SDS, whereas acetone and ethanol had no detrimental effect on folding of the enzyme. BACΔNC was inactive and unstable at extreme pH conditions. Thermal unfolding of the enzyme was found to be highly irreversible. The native enzyme started to unfold beyond ~0.2 M guanidine hydrochloride (GdnHCl) and reached an unfolded intermediate, [GdnHCl](0.5, N-U), at 1.14 M. BACΔNC was active at the concentrations of urea below 6 M, but it experienced an irreversible unfolding by >8 M denaturant. Taken together, this work lays a foundation for the future structural studies with Bacillus sp. TS-23 α-amylase, a typical member of glycoside hydrolases family 13.

Persistence of transplanted oral mucosal epithelial cells in human cornea.

PURPOSE: To determine the expression of differentiation and progenitor cell markers in corneal tissues that previously underwent autologous cultivated oral mucosal epithelial transplantation (COMET). METHODS: Four eyes from three alkaline-injured patients and one thermally injured patient underwent COMET to promote re-epithelialization or corneal reconstruction. Between 10 and 22 months (mean, 14.2 +/- 5.5 months [SD]) after COMET, the corneal tissues were obtained after penetrating keratoplasty (n = 1) or autologous limbal transplantation (n = 3). Immunoconfocal microscopy for keratin (K)3, -12, -4, -13, and -8; connexin (Cx)43; MUC5AC; laminin-5; pan-p63; ABCG2; and p75 was performed in those specimens as well as in the oral mucosa and cultivated oral mucosal epithelial cells (OMECs). RESULTS: All four specimens were unanimously positive for K3, -4, and -13 but negative for K8 and MUC5AC, suggesting that the keratinocytes were oral mucosa-derived. However, peripheral K12 staining was positive only in patient 2, suggesting a mixed oral and corneal epithelium in that case. Cx43 staining in the basal epithelium was negative in patients 1, 2, and 3, but was positive in patient 4. Small, compact keratinocytes in the basal epithelium preferentially expressed pan-p63, ABCG2, and p75. Although the staining of pan-p63 and ABCG2 tended to be more than one layer, signal for p75 was consistently localized only to the basal layer. CONCLUSIONS: The study demonstrated the persistence of transplanted OMECs in human corneas. In addition, small, compact cells in the basal epithelium preferentially expressed the keratinocyte stem/progenitor cell markers, which may be indicative of the engraftment of the progenitor cells after transplantation.