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1.  Intakes of Lutein, Zeaxanthin, and Other Carotenoids and Age-Related Macular Degeneration During 2 Decades of Prospective Follow-up 
JAMA ophthalmology  2015;133(12):1415-1424.
Importance
Despite strong biological plausibility, evidence from epidemiologic studies and clinical trials on the relations between intakes of lutein and zeaxanthin and age-related macular degeneration (AMD) has been inconsistent. The roles of other carotenoids are less thoroughly investigated.
Objective
To investigate the associations between intakes of carotenoids and AMD.
Design, Setting, and Participants
Prospective cohort study, with cohorts from the Nurses' Health Study and the Health Professionals Follow-up Study in the United States. A total of 63 443 women and 38 603 men were followed up, from 1984 until May 31, 2010, in the Nurses' Health Study and from 1986 until January 31, 2010, in the Health Professionals Follow-up Study. All participants were aged 50 years or older and were free of diagnosed AMD, diabetes mellitus, cardiovascular disease, and cancer at baseline.
Main outcomes and Measures
Predicted plasma carotenoid scores were computed directly from food intake, assessed by repeated food frequency questionnaires at baseline and follow-up, using validated regression models to account for bioavailability and reporting validity of different foods, and associations between predicted plasma carotenoid scores and AMD were determined.
Results
We confirmed 1361 incident intermediate and 1118 advanced AMD cases (primarily neovascular AMD) with a visual acuity of 20/30 or worse by medical record review. Comparing extreme quintiles of predicted plasma lutein/zeaxanthin score, we found a risk reduction for advanced AMD of about 40% in both women and men (pooled relative risk comparing extreme quintiles = 0.59; 95% CI, 0.48-0.73; P for trend < .001). Predicted plasma carotenoid scores for other carotenoids, including β-cryptoxanthin, α-carotene, and β-carotene, were associated with a 25% to 35% lower risk of advanced AMD when comparing extreme quintiles. The relative risk comparing extreme quintiles for the predicted plasma total carotenoid index was 0.65 (95% CI, 0.53-0.80; P for trend < .001). We did not identify any associations of carotenoids, either as predicted plasma score or calculated intake, with intermediate AMD.
Conclusions and Relevance
Higher intake of bioavailable lutein/zeaxanthin is associated with a long-term reduced risk of advanced AMD. Given that some other carotenoids are also associated with a lower risk, a public health strategy aimed at increasing dietary consumption of a wide variety of fruits and vegetables rich in carotenoids may reduce the incidence of advanced AMD.
doi:10.1001/jamaophthalmol.2015.3590
PMCID: PMC5119484  PMID: 26447482
2.  Multi-omic data integration enables discovery of hidden biological regularities 
Nature Communications  2016;7:13091.
Rapid growth in size and complexity of biological data sets has led to the ‘Big Data to Knowledge' challenge. We develop advanced data integration methods for multi-level analysis of genomic, transcriptomic, ribosomal profiling, proteomic and fluxomic data. First, we show that pairwise integration of primary omics data reveals regularities that tie cellular processes together in Escherichia coli: the number of protein molecules made per mRNA transcript and the number of ribosomes required per translated protein molecule. Second, we show that genome-scale models, based on genomic and bibliomic data, enable quantitative synchronization of disparate data types. Integrating omics data with models enabled the discovery of two novel regularities: condition invariant in vivo turnover rates of enzymes and the correlation of protein structural motifs and translational pausing. These regularities can be formally represented in a computable format allowing for coherent interpretation and prediction of fitness and selection that underlies cellular physiology.
Translating omics data sets into biological insight is one of the great challenges of our time. Here, the authors make headway by synchronising pairs of omics data types via invariants across conditions and by integrating datasets into a genome-scale model of E. coli metabolism and gene expression.
doi:10.1038/ncomms13091
PMCID: PMC5095171  PMID: 27782110
3.  Standardizing data exchange for clinical research protocols and case report forms: An assessment of the suitability of the Clinical Data Interchange Standards Consortium (CDISC) Operational Data Model (ODM) 
Efficient communication of a clinical study protocol and case report forms during all stages of a human clinical study is important for many stakeholders. An electronic and structured study representation format that can be used throughout the whole study life-span can improve such communication and potentially lower total study costs. The most relevant standard for representing clinical study data, applicable to unregulated as well as regulated studies, is the Operational Data Model (ODM) in development since 1999 by the Clinical Data Interchange Standards Consortium (CDISC). ODM's initial objective was exchange of case report forms data but it is increasingly utilized in other contexts. An ODM extension called Study Design Model, introduced in 2011, provides additional protocol representation elements.
Using a case study approach, we evaluated ODM's ability to capture all necessary protocol elements during a complete clinical study lifecycle in the Intramural Research Program of the National Institutes of Health. ODM offers the advantage of a single format for institutions that deal with hundreds or thousands of concurrent clinical studies and maintain a data warehouse for these studies. For each study stage, we present a list of gaps in the ODM standard and identify necessary vendor or institutional extensions that can compensate for such gaps. The current version of ODM (1.3.2) has only partial support for study protocol and study registration data mainly because it is outside the original development goal. ODM provides comprehensive support for representation of case report forms (in both the design stage and with patient level data). Inclusion of requirements of observational, non-regulated or investigator-initiated studies (outside Food and Drug Administration (FDA) regulation) can further improve future revisions of the standard.
doi:10.1016/j.jbi.2015.06.023
PMCID: PMC4714951  PMID: 26188274
Clinical research informatics; Clinical protocol; Standards; Clinical trial; electronic Case Report Forms
4.  Selective expression of Parkinson's disease-related Leucine-rich repeat kinase 2 G2019S missense mutation in midbrain dopaminergic neurons impairs dopamine release and dopaminergic gene expression 
Human Molecular Genetics  2015;24(18):5299-5312.
Preferential dysfunction/degeneration of midbrain substantia nigra pars compacta (SNpc) dopaminergic (DA) neurons contributes to the main movement symptoms manifested in Parkinson's disease (PD). Although the Leucine-rich repeat kinase 2 (LRRK2) G2019S missense mutation (LRRK2 G2019S) is the most common causative genetic factor linked to PD, the effects of LRRK2 G2019S on the function and survival of SNpc DA neurons are poorly understood. Using a binary gene expression system, we generated transgenic mice expressing either wild-type human LRRK2 (WT mice) or the LRRK2 G2019S mutation (G2019S mice) selectively in the midbrain DA neurons. Here we show that overexpression of LRRK2 G2019S did not induce overt motor abnormalities or substantial SNpc DA neuron loss. However, the LRRK2 G2019S mutation impaired dopamine homeostasis and release in aged mice. This reduction in dopamine content/release coincided with the degeneration of DA axon terminals and decreased expression of DA neuron-enriched genes tyrosine hydroxylase (TH), vesicular monoamine transporter 2, dopamine transporter and aldehyde dehydrogenase 1. These factors are responsible for dopamine synthesis, transport and degradation, and their expression is regulated by transcription factor paired-like homeodomain 3 (PITX3). Levels of Pitx3 mRNA and protein were similarly decreased in the SNpc DA neurons of aged G2019S mice. Together, these findings suggest that PITX3-dependent transcription regulation could be one of the many potential mechanisms by which LRRK2 G2019S acts in SNpc DA neurons, resulting in downregulation of its downstream target genes critical for dopamine homeostasis and release.
doi:10.1093/hmg/ddv249
PMCID: PMC4550828  PMID: 26123485
5.  The Hygiene Hypothesis and Its Inconvenient Truths about Helminth Infections 
PLoS Neglected Tropical Diseases  2016;10(9):e0004944.
Author Summary
Current iterations of the hygiene hypothesis suggest an adaptive role for helminth parasites in shaping the proper maturation of the immune system. However, aspects of this hypothesis are based on assumptions that may not fully account for realities about human helminth infections. Such realities include evidence of causal associations between helminth infections and asthma or inflammatory bowel disease as well as the fact that helminth infections remain widespread in the United States, especially among populations at greatest risk for inflammatory and autoimmune diseases.
doi:10.1371/journal.pntd.0004944
PMCID: PMC5025185  PMID: 27632204
6.  Proteomic Identification of Immunodiagnostic Antigens for Trypanosoma vivax Infections in Cattle and Generation of a Proof-of-Concept Lateral Flow Test Diagnostic Device 
PLoS Neglected Tropical Diseases  2016;10(9):e0004977.
Trypanosoma vivax is one of the causative agents of Animal African Trypanosomosis in cattle, which is endemic in sub-Saharan Africa and transmitted primarily by the bite of the tsetse fly vector. The parasite can also be mechanically transmitted, and this has allowed its spread to South America. Diagnostics are limited for this parasite and in farm settings diagnosis is mainly symptom-based. We set out to identify, using a proteomic approach, candidate diagnostic antigens to develop into an easy to use pen-side lateral flow test device. Two related members the invariant surface glycoprotein family, TvY486_0045500 and TvY486_0019690, were selected. Segments of these antigens, lacking N-terminal signal peptides and C-terminal transmembrane domains, were expressed in E. coli. Both were developed into ELISA tests and one of them, TvY486_0045500, was developed into a lateral flow test prototype. The tests were all evaluated blind with 113 randomised serum samples, taken from 37 calves before and after infection with T. vivax or T. congolense. The TvY486_0045500 and TvY486_0019690 ELISA tests gave identical sensitivity and specificity values for T. vivax infection of 94.5% (95% CI, 86.5% to 98.5%) and 88.0% (95% CI, 75.7% to 95.5%), respectively, and the TvY486_0045500 lateral flow test prototype a sensitivity and specificity of 92.0% (95% CI, 83.4% to 97.0%) and 89.8% (95% CI, 77.8% to 96.6%), respectively. These data suggest that recombinant TvY486_0045500 shows promise for the development of a pen-side lateral flow test for the diagnosis of T. vivax animal African trypanosomosis.
Author Summary
African Animal Trypanosomosis presents a significant problem for agricultural development in sub-Saharan Africa and leads to large economic losses. One of the main parasites responsible is Trypanosoma vivax. Current diagnostic methods are either symptom-based or too costly and technologically demanding for use in endemic regions. Here, we identified T. vivax proteins selectively recognized by infected cattle sera and developed two related proteins into ELISA tests and one of these into a lateral flow test prototype. All three tests performed well when tested against randomised calf sera, suggesting good potential for the development of a pen-side T. vivax animal African trypanosomosis diagnostic device for use in endemic regions.
doi:10.1371/journal.pntd.0004977
PMCID: PMC5015970  PMID: 27606593
7.  CrossLink: a novel method for cross-condition classification of cancer subtypes 
BMC Genomics  2016;17(Suppl 7):549.
Background
We considered the prediction of cancer classes (e.g. subtypes) using patient gene expression profiles that contain both systematic and condition-specific biases when compared with the training reference dataset. The conventional normalization-based approaches cannot guarantee that the gene signatures in the reference and prediction datasets always have the same distribution for all different conditions as the class-specific gene signatures change with the condition. Therefore, the trained classifier would work well under one condition but not under another.
Methods
To address the problem of current normalization approaches, we propose a novel algorithm called CrossLink (CL). CL recognizes that there is no universal, condition-independent normalization mapping of signatures. In contrast, it exploits the fact that the signature is unique to its associated class under any condition and thus employs an unsupervised clustering algorithm to discover this unique signature.
Results
We assessed the performance of CL for cross-condition predictions of PAM50 subtypes of breast cancer by using a simulated dataset modeled after TCGA BRCA tumor samples with a cross-validation scheme, and datasets with known and unknown PAM50 classification. CL achieved prediction accuracy >73 %, highest among other methods we evaluated. We also applied the algorithm to a set of breast cancer tumors derived from Arabic population to assign a PAM50 classification to each tumor based on their gene expression profiles.
Conclusions
A novel algorithm CrossLink for cross-condition prediction of cancer classes was proposed. In all test datasets, CL showed robust and consistent improvement in prediction performance over other state-of-the-art normalization and classification algorithms.
doi:10.1186/s12864-016-2903-z
PMCID: PMC5001207  PMID: 27556419
8.  Predictors of successful non-operative management of grade III & IV blunt pancreatic trauma 
Introduction
Although surgery is the preferred treatment for grade III&IV pancreatic trauma, there is a growing movement for non-operative management. in blunt pancreatic trauma. Very few studies compare operative versus non-operative management in adult patients.
Methods
Retrospective analysis of a prospectively maintained database was performed from 2004 to 2013 in the department of gastrointestinal surgery, NIMS, Hyderabad. Comparative analysis was performed between patients who failed versus those who were successfully managed with non-operative management.
Results
34 patients had grade III/IV trauma out of which 8 were operated early with the remaining 26 initially under a NOM strategy, 10 of them could be successfully managed without any operation. Post-traumatic pancreatitis, Necrotizing pancreatitis, Ileus, contusion on CT, surrounding organ injuries are independently associated with failure of NOM on a univariate analysis. On multivariate logistic regression presence of necrosis& associated organ injury are factors that predict failure of NOM independently. Development of a pseudocyst is the only significant factor that is associated with a success of NOM.
Conclusions
Non-operative measures should be attempted in a select group of grade III&IV blunt pancreatic trauma. In hemodynamically stable patients with a controlled leak walled off as a pseudocyst without associated organ injuries and pancreatic necrosis, NOM has a higher success rate.
Highlights
•Non-operative measures should be attempted in a select group of high grade (grade III/IV) pancreatic trauma.•Controlled leak walled off as a pseudocyst, absent necrosis&organ injuries predict high success rate for NOM.•Dedicated nutritional, gastrointestinal and interventional radiological support are the key components of care.
doi:10.1016/j.amsu.2016.08.003
PMCID: PMC4995476  PMID: 27594995
ISGPF: International study group of pancreatic fistula; NOM: non-operative management; AAST: American association for the surgery of trauma
9.  Structure-Based Design of Head-Only Fusion Glycoprotein Immunogens for Respiratory Syncytial Virus 
PLoS ONE  2016;11(7):e0159709.
Respiratory syncytial virus (RSV) is a significant cause of severe respiratory illness worldwide, particularly in infants, young children, and the elderly. Although no licensed vaccine is currently available, an engineered version of the metastable RSV fusion (F) surface glycoprotein—stabilized in the pre-fusion (pre-F) conformation by “DS-Cav1” mutations—elicits high titer RSV-neutralizing responses. Moreover, pre-F-specific antibodies, often against the neutralization-sensitive antigenic site Ø in the membrane-distal head region of trimeric F glycoprotein, comprise a substantial portion of the human response to natural RSV infection. To focus the vaccine-elicited response to antigenic site Ø, we designed a series of RSV F immunogens that comprised the membrane-distal head of the F glycoprotein in its pre-F conformation. These “head-only” immunogens formed monomers, dimers, and trimers. Antigenic analysis revealed that a majority of the 70 engineered head-only immunogens displayed reactivity to site Ø-targeting antibodies, which was similar to that of the parent RSV F DS-Cav1 trimers, often with increased thermostability. We evaluated four of these head-only immunogens in detail, probing their recognition by antibodies, their physical stability, structure, and immunogenicity. When tested in naïve mice, a head-only trimer, half the size of the parent RSV F trimer, induced RSV titers, which were statistically comparable to those induced by DS-Cav1. When used to boost DS-Cav1-primed mice, two head-only RSV F immunogens, a dimer and a trimer, boosted RSV-neutralizing titers to levels that were comparable to those boosted by DS-Cav1, although with higher site Ø-directed responses. Our results provide proof-of-concept for the ability of the smaller head-only RSV F immunogens to focus the vaccine-elicited response to antigenic site Ø. Decent primary immunogenicity, enhanced physical stability, potential ease of manufacture, and potent immunogenicity upon boosting suggest these head-only RSV F immunogens, engineered to retain the pre-fusion conformation, may have advantages as candidate RSV vaccines.
doi:10.1371/journal.pone.0159709
PMCID: PMC4963090  PMID: 27463224
10.  Evaluating the Anticancer Potential of Ethanolic Gall Extract of Terminalia chebula (Gaertn.) Retz. (Combretaceae) 
Pharmacognosy Research  2016;8(3):209-212.
Plants have been an important source for discovery of anticancer compounds. With the current decline in the number of new molecular entities from the pharmaceutical industry, novel anticancer agents are being sought from traditional medicines; therefore the anticancer efficacy of many plants that are used in traditional medicine is yet to be verified. The objective of the study was to evaluate the cytotoxic potential of ethanolic leaf gall extract of Terminalia chebula are evaluated against buffalo rat liver 3A, MCF-7 (Human mammary gland adenocarcinoma) and A-549 (Human lung cancer) cell lines. The cytotoxic effect of the ethanolic extract was evaluated by MTT assay. The extract was potent and effective in inducing cytotoxic effects in all the cell lines with an IC50 value of 305.18 ± 1.7 μg/mL, 643.13 ± 4.2 μg/mL, and 208.16 ± 3.7 μ/mL, respectively. The extract was more effective against A549 cell lines when compared to others. The presences of phenolics, triterpenoids, and flavonoids were identified in the extract. The extract showed total phenolic and flavonoid content of 478 ± 2.2 mg of gallic acid equivalent/g d.w and 538 ± 1.4 mg of quercetinequivalent/g d.w, respectively. This higher content of total phenolics and flavonoids found in the ethanolic extract was directly associated to higher cytotoxicity activity.
Conclusion:
The ethanolic leaf gall extract of T. chebula showed effective cytotoxic activities; which might be attributed to the phenolics/flavonoids present in higher concentration. Future work will be interesting to know the chemical composition of the extract and also better understand the mechanism of action of the constituents present in the extract to develop it as drug for therapeutic application.
SUMMARY
The present investigation establishes the anticancer activities of T. chebula leaf gall extracts on BRL3A, MCF-7, and A-549 cells. Presumably, these activities could be attributed in part to the phenolics/flavanoids features of the extract that has been demonstrated to act as cytotoxic agents. The experimental evidence obtained in the laboratory model could provide a rationale for the traditional use of plant as a source of easily available effective anticancer agents to the people, particularly in developing countries.
doi:10.4103/0974-8490.182919
PMCID: PMC4908852  PMID: 27365992
Alternative medicine; drugs; indian medicinal plants; inhibition; traditional medicine
11.  Temple-Baraitser Syndrome and Zimmermann-Laband Syndrome: one clinical entity? 
BMC Medical Genetics  2016;17:42.
Background
KCNH1 encodes a voltage-gated potassium channel that is predominantly expressed in the central nervous system. Mutations in this gene were recently found to be responsible for Temple-Baraitser Syndrome (TMBTS) and Zimmermann-Laband syndrome (ZLS).
Methods
Here, we report a new case of TMBTS diagnosed in a Lebanese child. Whole genome sequencing was carried out on DNA samples of the proband and his parents to identify mutations associated with this disease. Sanger sequencing was performed to confirm the presence of detected variants.
Results
Whole genome sequencing revealed three missense mutations in TMBTS patient: c.1042G > A in KCNH1, c.2131 T > C in STK36, and c.726C > A in ZNF517. According to all predictors, mutation in KCNH1 is damaging de novo mutation that results in substitution of Glycine by Arginine, i.e., p.(Gly348Arg). This mutation was already reported in a patient with ZLS that could affect the connecting loop between helices S4-S5 of KCNH1 with a gain of function effect.
Conclusions
Our findings demonstrate that KCNH1 mutations cause TMBTS and expand the mutational spectrum of KCNH1 in TMBTS. In addition, all cases of TMBTS were reviewed and compared to ZLS. We suggest that the two syndromes are a continuum and that the variability in the phenotypes is the result of the involvement of genetic modifiers.
doi:10.1186/s12881-016-0304-4
PMCID: PMC4901505  PMID: 27282200
Temple-Baraitser syndrome; Whole genome sequencing; KCNH1; Zimmermann-Laband syndrome
12.  Immigrant Latino Neighborhoods and Mortality among Infants Born to Mexican-Origin Latina Women 
Maternal and child health journal  2015;19(6):1354-1363.
Objectives
To compare the association between neighborhood Latino immigrant concentration and infant mortality by maternal nativity among singleton births to Mexican-origin women in Los Angeles County.
Methods
Information about births, infant deaths, and infant and maternal characteristics were obtained from geocoded Los Angeles County vital statistics records (2002–2005). Linked data on neighborhood characteristics (census tracts) were obtained from the 2000 Census. Logistic regression models were used to predict infant mortality while accounting for spatial clustering by census tract.
Results
Two-thirds of births to Mexican-origin mothers were to foreign-born women. Foreign-born mothers were older, had less education, and were more likely to have delivery costs paid by Medicaid than US-born mothers. Infants born to foreign-born women had a lower infant mortality rates than infants born to US-born women (3.8/1000 live births vs. 4.6, p=.002)). Among infants of foreign-born mothers, the odds of infant mortality increased with increasing immigrant concentration (OR: 1.29; 95%CI: 1.01–1.66). There was a similar pattern of association between immigrant concentration and mortality for infants of US-born mothers (OR: 1.29; 95% CI: 0.99–1.67).
Conclusions
In Los Angeles County, the odds of infant mortality among foreign-born Mexican-origin Latina were higher in higher-density immigrant neighborhoods, with a similar trend among US-born mothers. Thus, living in immigrant enclaves likely does not help to explain the lower than expected infant mortality rate among infants born to Latina women. Instead, higher neighborhood Latino immigrant concentration may indicate a neighborhood with characteristics that negatively impact maternal and infant health for Latinos.
doi:10.1007/s10995-014-1640-7
PMCID: PMC4447583  PMID: 25430802
13.  Clinical Appraisal of Fosfomycin in the Era of Antimicrobial Resistance 
Antimicrobial Agents and Chemotherapy  2015;59(12):7355-7361.
Fosfomycin is recommended as one of the first-line agents for treatment of urinary tract infections (UTIs) in the latest guidelines endorsed by the Infectious Diseases Society of America (IDSA) and the European Society for Clinical Microbiology and Infectious Diseases (ESCMID). We evaluated the use of fosfomycin among inpatients at a tertiary care hospital between 2009 and 2013. UTI cases were defined using physician diagnosis and the National Healthcare Safety Network (NHSN) surveillance definitions. The number of patients treated with fosfomycin increased from none in 2009 to 391 in 2013. Among 537 patients who received fosfomycin for any indication during this period, UTI was the most common indication (74%), followed by asymptomatic bacteriuria (10%). All except 19 patients received a single dose of fosfomycin. Escherichia coli was the most common organism involved (52%). For 119 patients with UTIs, after exclusion of those with negative urine culture results, negative urinalysis results, receipt of additional agents, or indeterminate clinical outcomes, the clinical success rate at 48 h was 74.8%. Of 89 patients who met the criteria for NHSN-defined UTIs, 89.9% had successful outcomes. Recurrent infections occurred in 4.3% of cases, and mild adverse events were observed in 2.0%. All 100 randomly selected extended-spectrum β-lactamase (ESBL)-producing E. coli clinical isolates from this period were susceptible to fosfomycin. In conclusion, the use of fosfomycin has increased substantially since implementation of the updated guidelines at this hospital. Fosfomycin was used mainly for the treatment of physician-diagnosed UTIs, and the clinical outcomes were generally favorable. Fosfomycin maintained activity against E. coli despite the increased use of the agent.
doi:10.1128/AAC.01071-15
PMCID: PMC4649162  PMID: 26369978
14.  Sublingual injection of microparticles containing glycolipid ligands for NKT cells and subunit vaccines induces antibody responses in oral cavity 
Carbohydrate research  2014;405:87-92.
Natural Killer T (NKT) cells are a unique type of innate immune cells which exert paradoxical roles in animal models through producing either Th1 or Th2 cytokines and activating dendritic cells. Alpha-galactosylceramide (αGalCer), a synthetic antigen for NKT cells, was found to be safe and immune stimulatory in cancer and hepatitis patients. We recently developed microparticle-formulated αGalCer, which is selectively presented by dendritic cells and macrophages, but not B cells, and thus can avoid the anergy of NKT cells. In this study, we have examined the immunogenicity of microparticles containing αGalCer and protein vaccine components through sublingual injection in mice. The results showed that sublingual injection of microparticles containing αGalCer and ovalbumin triggered IgG responses in serum (titer >1:100,000), which persisted for more than 3 months. Microparticles containing ovalbumin alone also induced comparable level of IgG responses. However, immunoglobulin subclass analysis showed that sublingually injected microparticles containing αGalCer and ovalbumin induced 20 fold higher Th1 biased antibody (IgG2c) than microparticles containing OVA alone (1:20,000 as compared to 1:1000 titer). Sublingual injection of microparticles containing αGalCer and ovalbumin induced secretion of both IgG (titer >1:1000) and IgA (titer =1:80) in saliva secretion, while microparticles containing ovalbumin alone only induced secretion of IgG in saliva. Our results suggest that sublingual injection of microparticles and their subsequent trafficking to draining lymph nodes may induce adaptive immune responses in mucosal compartments. Ongoing studies are focused on the mechanism of antigen presentation and lymphocyte biology in the oral cavity, as well as the toxicity and efficacy of these candidate microparticles for future applications.
doi:10.1016/j.carres.2014.11.007
PMCID: PMC4874192  PMID: 25555750
Microparticles; alpha-galactosylceramide; Natural Killer T cells; Vaccines; Mucosal Immunity; Antibody Class Switch
15.  Infectious SIV resides in adipose tissue and induces metabolic defects in chronically infected rhesus macaques 
Retrovirology  2016;13:30.
Background
HIV reservoirs pose major challenges to viral eradication. The main cellular reservoirs include CD4 T cells and macrophages, whereas anatomic reservoirs are thought to be primarily lymphoid tissues. Adipose tissue represents a potentially important non-lymphoid location for HIV replication and persistence because the stromal-vascular-fraction (AT-SVF) contains activated innate and adaptive immune cells that increase in number during infections, obesity, and chronic inflammation.
Results
Adipose tissue from two groups of SHIV-SF162p3-infected (~4 weeks acute infection) or SIVmac251-infected (~38 weeks chronic infection) rhesus macaques (N = 8 for each group) were studied for immune cell content, viral infectiousness, and metabolic health. The AT-SVF cells from SHIV-infected monkeys contained abundant memory CD4 and CD8 T cells, with fewer NKT cells and macrophages, and no B cells. Proviral DNA (Gag and Env) was readily detectable by nested PCR in AT-SVF cells from multiple adipose depots (subcutaneous and visceral) of acutely infected monkeys, but mostly from visceral fat. More importantly, viral outgrowth assays using input CD4 T cells derived from AT-SVF cells or peripheral blood of chronically infected monkeys resulted in robust replication of infectious virus from both AT-SVF and peripheral blood CD4 T cells. Chronically infected monkeys also experienced adipocyte dysfunction (suppression of major adipogenic genes) and systemic dyslipidemia (decreased serum total cholesterol and free fatty acids, and increased triglycerides), similar to metabolic abnormalities of HIV patients.
Conclusions
Adipose tissues of SIV-infected rhesus macaques become major compartments for infected immune cells, which in turn induce defects in adipose tissue metabolism.
Electronic supplementary material
The online version of this article (doi:10.1186/s12977-016-0260-2) contains supplementary material, which is available to authorized users.
doi:10.1186/s12977-016-0260-2
PMCID: PMC4847269  PMID: 27117277
Adipose tissue; CD4 T cells; HIV latency; HIV reservoirs; Rhesus macaques; SIV reservoirs
16.  Reversal of oxidative stress-induced apoptosis in T and B lymphocytes by Coenzyme Q10 (CoQ10) 
Coenzyme Q10, (CoQ10) an electron transporter and an antioxidant, protects a variety of cell types against oxidative stress and apoptosis. However, protective effect of CoQ10 on oxidative stress-induced apoptosis in lymphocytes has not been studied in detail. In this study, we investigated the effect of CoQ10 on oxidative stress-induced apoptosis in lymphocytes. An exposure of peripheral blood lymphocytes to oxidative stressors, rotenone or hydrogen peroxide, lead to apoptosis. Pre-treatment of lymphocytes with CoQ10 resulted in a significantly reduced level of oxidative stress-induced apoptosis, which was associated with decreased reactive oxygen species production, an inhibition of mitochondrial membrane depolarization, and inhibition of activation of caspase-9 and caspase-3. Furthermore, CoQ10 inhibited oxidative stress induced apoptosis in both CD4+ T, and CD8+ T, and CD19+ B cells. Our findings suggest that CoQ10 may provide new therapeutic strategies for preventing oxidative stress-induced cell death and dysfunction in lymphocytes and lymphocyte subsets.
PMCID: PMC4858601  PMID: 27168954
Coenzyme Q10; oxidative stress; apoptosis; lymphocytes
17.  Using the Panel Study of Income Dynamics To Analyze Housing Decisions, Dynamics, and Effects 
Cityscape (Washington, D.C.)  2016;18(1):185-199.
The Panel Study of Income Dynamics (PSID) is the world’s longest running household panel survey. It started in 1968 and has followed the same families—and their descendants—for nearly 50 years. PSID was conducted annually from 1968 through 1997 and has been conducted biennially since 1997. As of 2015, 39 waves of data have been collected. In 2015, interviews were completed with more than 9,000 households and information was collected on about 25,000 household members. PSID has achieved high wave-to-wave response rates throughout most of its history. Since the beginning of the study, detailed information has been collected on family composition, income, assets and debt, public program participation, and housing. At the beginning of the recent housing crisis, PSID began collecting information about mortgage distress and foreclosure activity. PSID currently includes several major supplemental studies. The Child Development Supplement and the Transition into Adulthood Supplement collect detailed information about behavior and outcomes among children and young adults in PSID families, such as educational achievement, health, time use, family formation, and housing-related decisions among young adults. PSID data are publicly available free of charge to researchers; some data available only under contract to qualified researchers allow linkage with various administrative databases and include information such as census tract and block of residence that can be used to describe neighborhood characteristics. PSID data have been widely used to study topics of major interest to Cityscape readers, including housing decisionmaking, housing expenditures and financing, residential mobility and migration, and the effects of neighborhood characteristics on a variety of measures of child and family well-being. This article provides an overview of PSID and its housing- and neighborhood-related measures. We briefly describe studies using PSID on housing-related topics. Finally, we point readers to resources needed to begin working with PSID data.
PMCID: PMC4839387  PMID: 27110321
18.  Cohort Profile: The Panel Study of Income Dynamics' Child Development Supplement and Transition into Adulthood Study 
The Child Development Supplement (CDS) was started in 1997 to collect information on children and caregivers in families in the USA that participated in the Panel Study of Income Dynamics (PSID), an ongoing national longitudinal household survey that began in 1968. CDS was launched with the goal of creating a comprehensive, nationally representative, prospective database of young children and their families for studying the dynamic process of children’s health and development. The same children and their caregivers were interviewed in up to three waves approximately every 5 years (1997, 2002–03, and 2007–08), with a child-based response rate of 90% in the most recent wave. Upon reaching age 18 years and finishing or leaving high school, the children in the CDS cohort shifted to a six-wave follow-up study launched in 2005 called the PSID Transition into Adulthood (TA) study. The TA data have been collected biennially through 2013, with a final wave planned for 2015. Once these young adults form their own economically independent households, they join the PSID. The main categories of data emphasize the major developmental tasks of childhood and young adulthood, including influences on successful development in the domains of family, schools and neighbourhoods. The majority of data and documentation are freely and publicly available through the PSID Online Data Center.
doi:10.1093/ije/dyu076
PMCID: PMC4553706  PMID: 24706732
19.  Intranasal Vaccination Affords Localization and Persistence of Antigen-Specific CD8+ T Lymphocytes in the Female Reproductive Tract 
Vaccines  2016;4(1):7.
Immunization strategies generating large numbers of antigen-specific T cells in the female reproductive tract (FRT) can provide barrier protection against sexually-transmitted pathogens, such as the human immunodeficiency virus (HIV) and human papillomaviruses (HPV). The kinetics and mechanisms of regulation of vaccine-induced adaptive T cell-mediated immune responses in FRT are less well defined. We present here evidence for intranasal delivery of the model antigen ovalbumin (OVA) along with alpha-galactosylceramide adjuvant as a protein vaccine to induce significantly higher levels of antigen-specific effector and memory CD8+ T cells in the FRT, relative to other systemic and mucosal tissues. Antibody blocking of the CXCR3 receptor significantly reduced antigen-specific CD8+ T cells subsequent to intranasal delivery of the protein vaccine suggesting an important role for the CXCR3 chemokine-receptor signaling for T cell trafficking. Further, intranasal vaccination with an adenoviral vector expressing OVA or HIV-1 envelope was as effective as intramuscular vaccination for generating OVA- or ENV-specific immunity in the FRT. These results support the application of the needle-free intranasal route as a practical approach to delivering protein as well as DNA/virus vector-based vaccines for efficient induction of effector and memory T cell immunity in the FRT.
doi:10.3390/vaccines4010007
PMCID: PMC4810059  PMID: 26999228
Female reproductive tract; CD8+ T cells; intranasal immunization
20.  Adaptive dosing of anticancer drugs in neonates: facilitating evidence-based dosing regimens 
Purpose
Selection of the most appropriate chemotherapy dosing regimens for neonates treated within the first weeks of life represents a significant clinical dilemma. Due to a lack of information relating to the clinical pharmacology of anticancer drugs in these challenging patients, current dosing guidelines are based on limited scientific rationale. In the current study, we investigate the utilisation of therapeutic drug monitoring approaches in neonates with localised hepatoblastoma, Wilms’ tumour and stage 4S neuroblastoma, being treated with widely used anticancer drugs.
Methods
Plasma concentrations of cisplatin, vincristine, etoposide and carboplatin were quantified in two neonates being treated within the first 3 weeks of life and in a 32-week preterm infant treated at a gestational age of 40 weeks. Therapeutic drug monitoring was carried out where appropriate, based on the pharmacokinetic data obtained in conjunction with clinical response and toxicity.
Results
Treatment of a child aged 2 weeks with a recommended cisplatin dose reduction for weight to 1.8 mg/kg resulted in achievement of unbound cisplatin plasma concentrations of 0.01–0.08 µg/mL, markedly lower than exposures previously reported in infants and older children. A dose increase to 2.7 mg/kg was implemented, leading to the achievement of levels more in-line with those previously reported. This increased dose level was well tolerated over six courses of treatment, resulting in a good response to cisplatin monotherapy and the patient remains in remission at 3.5 years. In contrast, a 50 % vincristine dose reduction for weight in a 3-week-old neonate resulted in plasma concentrations comparable to levels observed in older children, leading to successful treatment and continued remission at 2 years. In a third patient, etoposide and carboplatin clearance values normalised to body weight were comparable to those reported in older children, resulting in comparatively lower exposures following reduced dosing.
Conclusions
The current report provides unique data on the pharmacokinetics of several widely used anticancer drugs in neonates treated within the first few weeks of life. The provision of these data acts as a useful reference point to support future dosing decisions to be made by clinicians in the treatment of these challenging patients.
doi:10.1007/s00280-016-2975-0
PMCID: PMC4819938  PMID: 26875154
Neonates; Chemotherapy; Hepatoblastoma; Neuroblastoma; Wilms’ tumour; Cisplatin; Vincristine; Carboplatin; Etoposide; Therapeutic drug monitoring
21.  Crystal structure, conformational fixation, and entry-related interactions of mature ligand-free HIV-1 Env 
As the sole viral antigen on the HIV-1-virion surface, trimeric Env is a focus of vaccine efforts. Here we present the structure of the ligand-free HIV-1-Env trimer, fix its conformation, and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies, but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C-433C (DS) variant, specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer bound by a single CD4 without the typical antigenic hallmarks of CD4 induction. Antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like particle and soluble formats providing a new generation of vaccine antigens.
doi:10.1038/nsmb.3051
PMCID: PMC4706170  PMID: 26098315
22.  Deciphering the Dynamics of Non-Covalent Interactions Affecting Thermal Stability of a Protein: Molecular Dynamics Study on Point Mutant of Thermus thermophilus Isopropylmalate Dehydrogenase 
PLoS ONE  2015;10(12):e0144294.
Thermus thermophilius isopropylmalate dehydrogenase catalyzes oxidative decarboxylation and dehydrogenation of isopropylmalate. Substitution of leucine to alanine at position 172 enhances the thermal stability among the known point mutants. Exploring the dynamic properties of non-covalent interactions such as saltbridges, hydrogen bonds and hydrophobic interactions to explain thermal stability of a protein is interesting in its own right. In this study dynamic changes in the non-covalent interactions are studied to decipher the deterministic features of thermal stability of a protein considering a case study of a point mutant in Thermus thermophilus isopropylmalate dehydrogenase. A total of four molecular dynamic simulations of 0.2 μs were carried out on wild type and mutant’s functional dimers at 300 K and 337 K. Higher thermal stability of the mutant as compared to wild type is revealed by root mean square deviation, root mean square fluctuations and Cα-Cα distance with an increase in temperature from 300 K to 337 K. Most of the regions of wild type fluctuate higher than the corresponding regions of mutant with an increase in temperature. Cα-Cα distance analysis suggests that long distance networks are significantly affected in wild type as compared to the mutant. Short lived contacts are higher in wild type, while long lived contacts are lost at 337 K. The mutant forms less hydrogen bonds with water as compared to wild type at 337 K. In contrast to wild type, the mutant shows significant increase in unique saltbridges, hydrogen bonds and hydrophobic contacts at 337 K. The current study indicates that there is a strong inter-dependence of thermal stability on the way in which non-covalent interactions reorganize, and it is rewarding to explore this connection in single mutant studies.
doi:10.1371/journal.pone.0144294
PMCID: PMC4689552  PMID: 26657745
23.  Prefusion F–specific antibodies determine the magnitude of RSV neutralizing activity in human sera 
Science translational medicine  2015;7(309):309ra162.
Respiratory syncytial virus (RSV) is estimated to claim more lives among infants <1 year old than any other single pathogen, except malaria, and poses a substantial global health burden. Viral entry is mediated by a type I fusion glycoprotein (F) that transitions from a metastable prefusion (pre-F) to a stable postfusion (post-F) trimer. A highly neutralization-sensitive epitope, antigenic site Ø, is found only on pre-F. We determined what fraction of neutralizing (NT) activity in human sera is dependent on antibodies specific for antigenic site Ø or other antigenic sites on F in healthy subjects from ages 7 to 93 years. Adsorption of individual sera with stabilized pre-F protein removed >90% of NT activity and depleted binding antibodies to both F conformations. In contrast, adsorption with post-F removed ~30% of NT activity, and binding antibodies to pre-F were retained. These findings were consistent across all age groups. Protein competition neutralization assays with pre-F mutants in which sites Ø or II were altered to knock out binding of antibodies to the corresponding sites showed that these sites accounted for ~35 and <10% of NT activity, respectively. Binding competition assays with monoclonal antibodies (mAbs) indicated that the amount of site Ø–specific antibodies correlated with NT activity, whereas the magnitude of binding competed by site II mAbs did not correlate with neutralization. Our results indicate that RSV NT activity in human sera is primarily derived from pre-F–specific antibodies, and therefore, inducing or boosting NT activity by vaccination will be facilitated by using pre-F antigens that preserve site Ø.
doi:10.1126/scitranslmed.aac4241
PMCID: PMC4672383  PMID: 26468324
24.  Natural Killer T Cell and TLR9 Agonists as Mucosal Adjuvants for Sublingual Vaccination with Clade C HIV-1 Envelope Protein 
Vaccine  2014;32(51):6934-6940.
The vast majority of HIV-1 infections occur at mucosa during sexual contact. It may therefore be advantageous to provide mucosal barrier protection against this entry by mucosal vaccination. While a number of mucosal routes of vaccination are possible, many like enteric oral vaccines or intranasal vaccines have significant impediments that limit vaccine efficacy or pose safety risks. In contrast, immunogens applied to the sublingual region of the mouth could provide a simple route for mucosal vaccination. While sublingual immunization is appealing, this site does not always drive strong immune responses, particularly when using protein antigens. To address this issue, we have tested the ability of two mucosal adjuvants: alpha-galactosylceramide (αGalCer) that is a potent stimulator of natural killer T cells and CpG-oligodeoxynucleotide (CpG-ODN) a TLR9 agonist for their ability to amplify immune responses against clade C gp140 HIV-1 envelope protein antigen. Immunization with envelope protein alone resulted in a weak T cell and antibody responses. In contrast, CD4+ and CD8+ T cells responses in systemic and mucosal tissues were significantly higher in mice immunized with gp140 in the presence of either αGalCer or CpG-ODN and these responses were further augmented when the two adjuvants were used together. While both the adjuvants effectively increased gp140-specific serum IgG and vaginal IgA antibody levels, combining both significantly improved these responses. Memory T cell responses 60 days after immunization revealed αGalCer to be more potent than CpG-ODN and the combination of the αGalCer and CpG-ODN adjuvants was more effective than either alone. Serum and vaginal washes collected 60 days after immunization with gp140 with both αGalCer and CpG-ODN adjuvants had significant neutralization activity against Tier 1 and Tier 2 SHIVs. These data support the utility of the sublingual route for mucosal vaccination particularly in combination with αGalCer and CpG-ODN adjuvants.
doi:10.1016/j.vaccine.2014.10.051
PMCID: PMC4254499  PMID: 25444819
25.  No apparent transmission of transgenic α–synuclein into nigrostriatal dopaminergic neurons in multiple mouse models 
Background
α–synuclein (α–syn) is the main component of intracytoplasmic inclusions deposited in the brains of patients with Parkinson’s disease (PD) and certain other neurodegenerative disorders. Recent studies have explored the ability of α–syn to propagate between or across neighboring neurons and supposedly “infect” them with a prion–like mechanism. However, much of this research has used stereotaxic injections of heterologous α–syn fibrils to induce the spreading of inclusions in the rodent brains. Whether α–syn is able to transmit from the host cells to their neighboring cells in vivo is unclear.
Methods
Using immunestaining, we examined the potential propagation of α–syn into nigrostriatal dopaminergic (DA) neurons in three lines of transgenic mice that overexpress human wild–type α–syn (hα–syn) in different neuron populations.
Results
After testing for three different routes by which hα–syn propagation might occur, we were unable to find any evidence that hα–syn behaved like a prion and could be transmitted overtime into the DA neurons initially lack of hα–syn expression.
Conclusions
In transgenic mice hα–syn does not have the ability to propagate at pathologically significant levels between or across neurons. It must be noted that these observations do not disprove the studies that show its prion–like qualities, but rather that propagation is not detectable in transgenic models that do not use any injections of heterologous proteins or viral vectors to induce a spreading state.
doi:10.1186/s40035-015-0046-9
PMCID: PMC4668690  PMID: 26635953
Parkinson’s disease; α-synuclein; Propagation; Dopaminergic neurons; Transgenic mice

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