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1.
Eat Weight Disord ; 24(4): 705-714, 2019 Aug.
Article in English | MEDLINE | ID: mdl-31165446

ABSTRACT

PURPOSE: A growing body of literature has established that food and alcohol disturbance (FAD: decreasing one's caloric intake in preparation for alcohol consumption) is a specific health risk that endangers health and wellbeing. Recent research on trends in FAD has revealed ethno-racial disparities. A sociological analysis is helpful to center race and examine the role of ethnic identity in reproducing health disparities. The current study is guided by theories of socialization into ideal body types by race. METHODS: Study uses data from a cross-sectional survey conducted among college students. The sample includes White and Black American college students, ages 18-25, and uses ordinal logistic regression to test for the impact of race and ethnic identity on engagement in FAD using the Compensatory Eating and Behaviors in Response to Alcohol Consumption Scale (CEBRACS). RESULTS: FAD prevalence was lower among Black Americans than among White Americans in the sample. Results from ordered logistic regression models indicate that stronger ethnic ties reduce likelihood of FAD among Black Americans but have the opposite effect among White Americans. This modification effect provides evidence that ethnic identity belonging protects against FAD for Black Americans but acts as a risk factor for FAD among White Americans. CONCLUSIONS: Findings shed light on the documented racial disparities in FAD and weight control behavior more broadly. Ethnic identity modifies the relationship between race and FAD in our sample. LEVEL OF EVIDENCE: Level V, cross-sectional descriptive study.


Subject(s)
Alcohol Drinking/psychology , Feeding Behavior/psychology , Feeding and Eating Disorders/epidemiology , Adolescent , Adult , Black or African American , Caloric Restriction , Cross-Sectional Studies , Feeding and Eating Disorders/psychology , Female , Humans , Male , Prevalence , White People , Young Adult
2.
RSC Adv ; 8(62): 35768-35775, 2018 Oct 15.
Article in English | MEDLINE | ID: mdl-35547932

ABSTRACT

Legionaminic acid is a member of the nonulosonic acids, which are a class of sugars considered to be a virulence factor within a wide variety of pathogenic bacteria. We have developed a synthetic pathway towards C-7 analogues of legionaminic acid starting from Neu5Ac, resulting in the complete synthesis of both legionaminic acid, and its C-7 epimer, from a common precurser. Our approach involves the late-stage introduction of the requisite C-7 nitrogen functionality, thus making our strategy amenable to the introduction of a range of different amide groups at C-7 of legionaminic acid.

3.
PLoS One ; 10(11): e0139899, 2015.
Article in English | MEDLINE | ID: mdl-26580561

ABSTRACT

In portions of South Asia, vectors and patients co-infected with dengue (DENV) and chikungunya (CHIKV) are on the rise, with the potential for this occurrence in other regions of the world, for example the United States. Therefore, we engineered an antiviral approach that suppresses the replication of both arboviruses in mosquito cells using a single antiviral group I intron. We devised unique configurations of internal, external, and guide sequences that permit homologous recognition and splicing with conserved target sequences in the genomes of both viruses using a single trans-splicing Group I intron, and examined their effectiveness to suppress infections of DENV and CHIKV in mosquito cells when coupled with a proapoptotic 3' exon, ΔN Bax. RT-PCR demonstrated the utility of these introns in trans-splicing the ΔN Bax sequence downstream of either the DENV or CHIKV target site in transformed Aedes albopictus C6/36 cells, independent of the order in which the virus specific targeting sequences were inserted into the construct. This trans-splicing reaction forms DENV or CHIKV ΔN Bax RNA fusions that led to apoptotic cell death as evidenced by annexin V staining, caspase, and DNA fragmentation assays. TCID50-IFA analyses demonstrate effective suppression of DENV and CHIKV infections by our anti-arbovirus group I intron approach. This represents the first report of a dual-acting Group I intron, and demonstrates that we can target DENV and CHIKV RNAs in a sequence specific manner with a single, uniquely configured CHIKV/DENV dual targeting group I intron, leading to replication suppression of both arboviruses, and thus providing a promising single antiviral for the transgenic suppression of multiple arboviruses.


Subject(s)
Aedes/virology , Chikungunya virus/genetics , Dengue Virus/genetics , Introns , Trans-Splicing , Viral Proteins/genetics , bcl-2-Associated X Protein/genetics , Aedes/cytology , Animals , Annexin A5/metabolism , Apoptosis/genetics , Caspases/genetics , Caspases/metabolism , Cell Line , Chikungunya virus/metabolism , DNA Fragmentation , Dengue Virus/metabolism , Epithelial Cells/metabolism , Epithelial Cells/pathology , Epithelial Cells/virology , Exons , Female , Humans , Protein Structure, Tertiary , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Tetrahymena thermophila/chemistry , Tetrahymena thermophila/genetics , Transformation, Genetic , Viral Proteins/metabolism , bcl-2-Associated X Protein/metabolism
4.
Home Health Care Serv Q ; 34(2): 101-12, 2015.
Article in English | MEDLINE | ID: mdl-25894489

ABSTRACT

Adult day services (ADS) are the leading provider of community-based care for persons with dementia and their caregivers. While the provision of caregiver respite is well-documented, little is known about the provision of other forms of dementia caregiver support. Logistic regression analyses of ADS providers (N = 297) in the MetLife Study indicated that the number of hours of social work support was a significant predictor of case management services, while nonprofit status was a significant predictor of caregiver education and support groups. These findings have implications for practice and policy related to this growing provider of dementia services.


Subject(s)
Aged , Caregivers/psychology , Day Care, Medical/psychology , Dementia/complications , Stress, Psychological/psychology , Aged, 80 and over , Day Care, Medical/statistics & numerical data , Dementia/psychology , Dementia/therapy , Humans , Regression Analysis , Stress, Psychological/complications , Surveys and Questionnaires
5.
Virology ; 481: 223-34, 2015 Jul.
Article in English | MEDLINE | ID: mdl-25840398

ABSTRACT

The HCV-IRES sequence is vital for both protein translation and genome replication and serves as a potential target for anti-HCV therapy. We constructed a series of anti-HCV group I introns (αHCV-GrpIs) to attack conserved target sites within the HCV IRES. These αHCV-GrpIs were designed to mediate a trans-splicing reaction that replaces the viral RNA genome downstream of the 5' splice site with a 3' exon that encodes an apoptosis-inducing gene. Pro-active forms of the apoptosis inducing genes BID, Caspase 3, Caspase 8, or tBax were modified by incorporation of the HCV NS5A/5B cleavage sequence in place of their respective endogenous cleavage sites to ensure that only HCV infected cells would undergo apoptosis following splicing and expression. Huh7.5 cells transfected with each intron were challenged at MOI 0.1 with HCV-Jc1FLAG2 which expresses a Gaussia Luciferase (GLuc) marker. Virus-containing supernatants were then assayed for GLuc expression as a measure of viral replication inhibition. Cellular extracts were analyzed for the presence of correct splice products by RT-PCR and DNA sequencing. We also measured levels of Caspase 3 activity as a means of quantifying apoptotic cell death. Each of these αHCV-GrpI introns was able to correctly splice their 3' apoptotic exons onto the virus RNA genome at the targeted Uracil, and resulted in greater than 80% suppression of the GLuc marker. A more pronounced suppression effect was observed with TCID50 virus titrations, which demonstrated that these αHCV-GrpIs were able to suppress viral replication by more than 2 logs, or greater than 99%. Robust activation of the apoptotic factor within the challenged cells was evidenced by a significant increase of Caspase 3 activity upon viral infection compared to non-challenged cells. This novel genetic intervention tool may prove beneficial in certain HCV subjects.


Subject(s)
Hepacivirus/genetics , Hepatitis C/physiopathology , Hepatitis C/virology , Introns , RNA, Viral/genetics , Trans-Splicing , Base Sequence , Cell Death , Cell Line, Tumor , Gene Expression Regulation, Viral , Gene Targeting , Hepacivirus/chemistry , Hepacivirus/physiology , Humans , Nucleic Acid Conformation , RNA, Viral/chemistry , RNA, Viral/metabolism
6.
Virol J ; 11: 111, 2014 Jun 13.
Article in English | MEDLINE | ID: mdl-24927852

ABSTRACT

INTRODUCTION: Approximately 100 million confirmed infections and 20,000 deaths are caused by Dengue virus (DENV) outbreaks annually. Global warming and rapid dispersal have resulted in DENV epidemics in formally non-endemic regions. Currently no consistently effective preventive measures for DENV exist, prompting development of transgenic and paratransgenic vector control approaches. Production of transgenic mosquitoes refractory for virus infection and/or transmission is contingent upon defining antiviral genes that have low probability for allowing escape mutations, and are equally effective against multiple serotypes. Previously we demonstrated the effectiveness of an anti-viral group I intron targeting U143 of the DENV genome in mediating trans-splicing and expression of a marker gene with the capsid coding domain. In this report we examine the effectiveness of coupling expression of ΔN Bax to trans-splicing U143 intron activity as a means of suppressing DENV infection of mosquito cells. RESULTS: Targeting the conserved DENV circularization sequence (CS) by U143 intron trans-splicing activity appends a 3' exon RNA encoding ΔN Bax to the capsid coding region of the genomic RNA, resulting in a chimeric protein that induces premature cell death upon infection. TCID50-IFA analyses demonstrate an enhancement of DENV suppression for all DENV serotypes tested over the identical group I intron coupled with the non-apoptotic inducing firefly luciferase as the 3' exon. These cumulative results confirm the increased effectiveness of this αDENV-U143-ΔN Bax group I intron as a sequence specific antiviral that should be useful for suppression of DENV in transgenic mosquitoes. Annexin V staining, caspase 3 assays, and DNA ladder observations confirm DCA-ΔN Bax fusion protein expression induces apoptotic cell death. CONCLUSION: This report confirms the relative effectiveness of an anti-DENV group I intron coupled to an apoptosis-inducing ΔN Bax 3' exon that trans-splices conserved sequences of the 5' CS region of all DENV serotypes and induces apoptotic cell death upon infection. Our results confirm coupling the targeted ribozyme capabilities of the group I intron with the generation of an apoptosis-inducing transcript increases the effectiveness of infection suppression, improving the prospects of this unique approach as a means of inducing transgenic refractoriness in mosquitoes for all serotypes of this important disease.


Subject(s)
Apoptosis/genetics , Dengue Virus/genetics , Gene Expression , Introns , Protein Interaction Domains and Motifs/genetics , bcl-2-Associated X Protein/genetics , Animals , Cell Line , Culicidae , Dengue/virology , Dengue Virus/classification , Exons , Gene Order , Genetic Vectors , Promoter Regions, Genetic , Serogroup , Trans-Splicing , Virus Replication/genetics , bcl-2-Associated X Protein/chemistry
7.
Methods Mol Biol ; 1103: 57-66, 2014.
Article in English | MEDLINE | ID: mdl-24318886

ABSTRACT

In vitro cleavage assays are routinely conducted to properly assess the catalytic activity of hammerhead ribozymes (HHR) against target RNA molecules like dengue virus RNA. These experiments are performed for initial assessment of HHR catalysis in a cell-free system and have been simplified by the substitution of agarose gel electrophoresis for SDS-PAGE. Substituting mobility assays enables the analysis of ribozymes in a more rapid fashion without radioisotopes. Here we describe the in vitro transcription of an HHR and corresponding target from T7-promoted plasmids into RNA molecules leading to the analysis of HHR activity against the RNA target by in vitro cleavage assays.


Subject(s)
Dengue Virus/genetics , RNA, Catalytic/genetics , RNA, Viral/genetics , Catalysis , Dengue Virus/growth & development , Electrophoresis, Agar Gel , Humans , Kinetics , Molecular Biology/methods
8.
J Biol Chem ; 288(50): 35726-35, 2013 Dec 13.
Article in English | MEDLINE | ID: mdl-24178297

ABSTRACT

Paramyxovirus fusion (F) proteins promote membrane fusion between the viral envelope and host cell membranes, a critical early step in viral infection. Although mutational analyses have indicated that transmembrane (TM) domain residues can affect folding or function of viral fusion proteins, direct analysis of TM-TM interactions has proved challenging. To directly assess TM interactions, the oligomeric state of purified chimeric proteins containing the Staphylococcal nuclease (SN) protein linked to the TM segments from three paramyxovirus F proteins was analyzed by sedimentation equilibrium analysis in detergent and buffer conditions that allowed density matching. A monomer-trimer equilibrium best fit was found for all three SN-TM constructs tested, and similar fits were obtained with peptides corresponding to just the TM region of two different paramyxovirus F proteins. These findings demonstrate for the first time that class I viral fusion protein TM domains can self-associate as trimeric complexes in the absence of the rest of the protein. Glycine residues have been implicated in TM helix interactions, so the effect of mutations at Hendra F Gly-508 was assessed in the context of the whole F protein. Mutations G508I or G508L resulted in decreased cell surface expression of the fusogenic form, consistent with decreased stability of the prefusion form of the protein. Sedimentation equilibrium analysis of TM domains containing these mutations gave higher relative association constants, suggesting altered TM-TM interactions. Overall, these results suggest that trimeric TM interactions are important driving forces for protein folding, stability and membrane fusion promotion.


Subject(s)
Cell Membrane/metabolism , Protein Folding , Protein Multimerization , Viral Fusion Proteins/chemistry , Viral Fusion Proteins/metabolism , Amino Acid Sequence , Animals , Cell Membrane/virology , Chlorocebus aethiops , Membrane Fusion , Molecular Sequence Data , Protein Stability , Protein Structure, Quaternary , Protein Structure, Tertiary , Vero Cells
9.
Virol J ; 10: 201, 2013 Jun 28.
Article in English | MEDLINE | ID: mdl-23809208

ABSTRACT

BACKGROUND: Recent epidemics of dengue viruses (DENV) coupled with new outbreaks on the horizon have renewed the demand for novel detection methods that have the ability to identify this viral pathogen prior to the manifestation of symptoms. The ability to detect DENV in a timely manner is essential for rapid recovery from disease symptoms. A modified lab-derived 10-23 DNAzyme tethered to gold nanoparticles provides a powerful tool for the detection of viruses, such as DENV. RESULTS: We examined the effectiveness of coupling DNAzyme (DDZ) activation to the salt-induced aggregation of gold nanoparticles (AuNP) to detect dengue virus (DENV) progeny in mosquito cells. A DNAzyme was designed to recognize the 5' cyclization sequence (5' CS) that is conserved among all DENV, and conjugated to AuNPs. DDZ-AuNP has demonstrated the ability to detect the genomic RNA of our model dengue strain, DENV-2 NGC, isolated from infected Aedes albopictus C6/36 cells. These targeting events lead to the rapid aggregation of AuNPs, resulting in a red to clear color transition of the reaction mixes, and thus positive detection of the DENV RNA genome. The inclusion of SDS in the reaction mixture permitted the detection of DENV directly from cell culture supernatants without additional sample processing. Specificity assays demonstrated detection is DENV-specific, while sensitivity assays confirm detection at levels of 1 × 10(1) TCID50 units. These results demonstrate DDZ-AuNP effectively detects DENV genomes in a sequence specific manner and at concentrations that are practical for field use. CONCLUSIONS: We have developed an effective detection assay using DNAzyme catalysis coupled with AuNP aggregation for the detection of DENV genomes in a sequence specific manner. Full development of our novel DDZ-AuNP detection method will provide a practical, rapid, and low cost alternative for the detection of DENV in mosquito cells and tissues, and possibly infected patient serum, in a matter of minutes with little to no specialized training required.


Subject(s)
DNA, Catalytic , Dengue Virus/isolation & purification , Dengue/virology , Gold , Nanoparticles , RNA, Viral/isolation & purification , Virology/methods , Aedes , Animals , Cell Line , Dengue/diagnosis , Dengue Virus/genetics , Humans , RNA, Viral/genetics , Sensitivity and Specificity
10.
J Virol ; 86(6): 3014-26, 2012 Mar.
Article in English | MEDLINE | ID: mdl-22238299

ABSTRACT

Hendra virus is a highly pathogenic paramyxovirus classified as a biosafety level four agent. The fusion (F) protein of Hendra virus is critical for promoting viral entry and cell-to-cell fusion. To be fusogenically active, Hendra virus F must undergo endocytic recycling and cleavage by the endosomal/lysosomal protease cathepsin L, but the route of Hendra virus F following internalization and the recycling signals involved are poorly understood. We examined the intracellular distribution of Hendra virus F following endocytosis and showed that it is primarily present in Rab5- and Rab4-positive endosomal compartments, suggesting that cathepsin L cleavage occurs in early endosomes. Hendra virus F transmembrane domain (TMD) residues S490 and Y498 were found to be important for correct Hendra virus F recycling, with the hydroxyl group of S490 and the aromatic ring of Y498 important for this process. In addition, changes in association of isolated Hendra virus F TMDs correlated with alterations to Hendra virus F recycling, suggesting that appropriate TMD interactions play an important role in endocytic trafficking.


Subject(s)
Endocytosis , Hendra Virus/metabolism , Henipavirus Infections/physiopathology , Henipavirus Infections/virology , Viral Fusion Proteins/chemistry , Viral Fusion Proteins/metabolism , Amino Acid Motifs , Amino Acid Sequence , Animals , Cell Line , Endosomes/metabolism , Hendra Virus/chemistry , Hendra Virus/genetics , Humans , Molecular Sequence Data , Protein Structure, Tertiary , Protein Transport , Sequence Alignment , Viral Fusion Proteins/genetics
11.
BMC Mol Biol ; 11: 84, 2010 Nov 15.
Article in English | MEDLINE | ID: mdl-21078188

ABSTRACT

BACKGROUND: Dengue viruses (DENV) are one of the most important viral diseases in the world with approximately 100 million infections and 200,000 deaths each year. The current lack of an approved tetravalent vaccine and ineffective insecticide control measures warrant a search for alternatives to effectively combat DENV. The trans-splicing variant of the Tetrahymena thermophila group I intron catalytic RNA, or ribozyme, is a powerful tool for post-transcriptional RNA modification. The nature of the ribozyme and the predictability with which it can be directed makes it a powerful tool for modifying RNA in nearly any cell type without the need for genome-altering gene therapy techniques or dependence on native cofactors. RESULTS: Several anti-DENV Group I trans-splicing introns (αDENV-GrpIs) were designed and tested for their ability to target DENV-2 NGC genomes in situ. We have successfully targeted two different uracil bases on the positive sense genomic strand within the highly conserved 5'-3' cyclization sequence (CS) region common to all serotypes of DENV with our αDENV-GrpIs. Our ribozymes have demonstrated ability to specifically trans-splice a new RNA sequence downstream of the targeted site in vitro and in transfected insect cells as analyzed by firefly luciferase and RT-PCR assays. The effectiveness of these αDENV-GrpIs to target infecting DENV genomes is also validated in transfected or transformed Aedes mosquito cell lines upon infection with unattenuated DENV-2 NGC. CONCLUSIONS: Analysis shows that our αDENV-GrpIs have the ability to effectively trans-splice the DENV genome in situ. Notably, these results show that the αDENV-GrpI 9v1, designed to be active against all forms of Dengue virus, effectively targeted the DENV-2 NGC genome in a sequence specific manner. These novel αDENV-GrpI introns provide a striking alternative to other RNA based approaches for the transgenic suppression of DENV in transformed mosquito cells and tissues.


Subject(s)
Dengue Virus/genetics , Genome, Viral , Introns , RNA, Catalytic/genetics , RNA, Viral/genetics , Trans-Splicing , Animals , Base Sequence , Cell Line , Culicidae/virology , Dengue/genetics , Gene Expression , Humans , Molecular Sequence Data
12.
J Gen Virol ; 89(Pt 12): 3150-3155, 2008 Dec.
Article in English | MEDLINE | ID: mdl-19008405

ABSTRACT

Comparisons of the relative activities of 11 intergenic region (IGR) internal ribosome entry site (IRES) elements of insect dicistrovirus with 5' IRES elements of the hepatitis C and encephalomyocarditis viruses were performed in insect and mammalian cells. Dual luciferase assays were performed to determine the most effective dicistrovirus IGR IRES in the lepidopteran cell lines Sf9 (Spodoptera frugiperda) and BmN (Bombyx mori), and the dipteran cell lines S2 (Drosophila melanogaster) and ATC-10 (Aedes aegypti). Evaluation of dual luciferase expression from DNA plasmids and in vitro-transcribed RNA revealed apparent splicing with certain IRES elements. Though IRES activity depended upon the cell line examined, the black queen cell and Drosophila C dicistrovirus intergenic IRES elements were most effective for coupled gene expression in the diverse insect cell lines examined.


Subject(s)
DNA, Intergenic , Enhancer Elements, Genetic , Insect Viruses/pathogenicity , RNA Viruses/pathogenicity , RNA, Messenger/metabolism , Aedes/virology , Animals , Bombyx/virology , Cell Line , Drosophila melanogaster/virology , Humans , Insect Viruses/genetics , Insect Viruses/metabolism , Luciferases/genetics , Luciferases/metabolism , Plasmids/genetics , RNA Splicing , RNA Viruses/genetics , RNA Viruses/metabolism , RNA, Messenger/genetics , RNA, Viral/genetics , RNA, Viral/metabolism , Ribosomes/genetics , Ribosomes/metabolism , Spodoptera/virology
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