Factors Associated with Slow Disease Progression in Macaques Immunized with an Adenovirus-Simian Immunodeficiency Virus (SIV) Envelope Priming-gp120 Boosting Regimen and Challenged Vaginally with SIVmac251

Rhesus macaques were immunized with a combination vaccine regimen consisting of adenovirus type 5 host range mutant-simian immunodeficiency virus envelope (Ad5hr-SIVenv) recombinant priming and boosting with native SIV gp120. Upon intravaginal challenge with SIVmac251, both persistently and transiently viremic animals were observed (S. L. Buge, E. Richardson, S. Alipanah, P. Markham, S. Cheng, N. Kalyan, C. J. Miller, M. Lubeck, S. Udem, J. Eldridge, and M. Robert-Guroff, J. Virol. 71:8531-8541, 1997). Long-term follow-up of the persistently viremic immunized macaques, which displayed significantly reduced viral burdens during the first 18 weeks postchallenge compared to controls, has now shown that one of four became a slow progressor, clearing virus from plasma and remaining asymptomatic with stable CD4 counts for 134 weeks postchallenge. Reboosting of the transiently viremic macaques did not reactivate latent virus. Rechallenge with two sequential SIVmac251 intravaginal exposures again resulted in partial protection of one of two immunized macaques, manifested by viral clearance and stable CD4 counts. No single immune parameter was associated with partial protection. Development of a strong antibody response capable of neutralizing a primary SIVmac251 isolate together with SIV-specific cytotoxic T lymphocytes were implicated, while CD8(+) T-cell antiviral activity and mucosal immune responses were not associated with delayed disease progression. Our data show that even a third immunization with the same Ad5hr-SIVenv recombinant can elicit significant immune responses to the inserted gene product, suggesting that preexisting Ad antibodies may not preclude effective immunization. Further, the partial protection against a virulent, pathogenic SIV challenge observed in two of six macaques immunized with a vaccine regimen based solely on the viral envelope indicates that this vectored-vaccine approach has promise and that multicomponent vaccines based in the same system merit further investigation.     

Outer membrane permeabilization by the membrane attack complex sensitizes Gram-negative bacteria to antimicrobial proteins in serum and phagocytes

Infections with Gram-negative bacteria form an increasing risk for human health due to antibiotic resistance. Our immune system contains various antimicrobial proteins that can degrade the bacterial cell envelope. However, many of these proteins do not function on Gram-negative bacteria, because the impermeable outer membrane of these bacteria prevents such components from reaching their targets. Here we show that complement-dependent formation of Membrane Attack Complex (MAC) pores permeabilizes this barrier, allowing antimicrobial proteins to cross the outer membrane and exert their antimicrobial function. Specifically, we demonstrate that MAC-dependent outer membrane damage enables human lysozyme to degrade the cell wall of E. coli. Using flow cytometry and confocal microscopy, we show that the combination of MAC pores and lysozyme triggers effective E. coli cell wall degradation in human serum, thereby altering the bacterial cell morphology from rod-shaped to spherical. Completely assembled MAC pores are required to sensitize E. coli to the antimicrobial actions of lysozyme and other immune factors, such as Human Group IIA-secreted Phospholipase A2. Next to these effects in a serum environment, we observed that the MAC also sensitizes E. coli to more efficient degradation and killing inside human neutrophils. Altogether, this study serves as a proof of principle on how different players of the human immune system can work together to degrade the complex cell envelope of Gram-negative bacteria. This knowledge may facilitate the development of new antimicrobials that could stimulate or work synergistically with the immune system.     

Tolerance to nickel: oral nickel administration induces a high frequency of anergic T cells with persistent suppressor activity.

We adapted our mouse model of allergic contact hypersensitivity to nickel for the study of tolerance. Sensitization in this model is achieved by the administration of nickel ions with H(2)O(2); nickel ions alone are unable to prime naive T cells, but can restimulate primed ones. A 4-wk course of oral or i.p. administration of 10 mM NiCl(2) to naive mice induced tolerance, preventing the induction of hypersensitivity for at least 20 wk; long term desensitization of nickel-sensitized mice, however, required continuous NiCl(2) administration. When splenic T cells of orally tolerized donors, even after a treatment-free interval of 20 wk, were transferred to naive recipients, as with lymph node cells (LNC), they specifically prevented sensitization of the recipients. The LNC of such donors were anergic, because upon in vivo sensitization with NiCl(2) in H(2)O(2) and in vitro restimulation with NiCl(2), they failed to show the enhanced proliferation and IL-2 production as seen with LNC of mice not tolerized before sensitization. As few as 10(2) bulk T cells, consisting of both CD4(+) and CD8(+) cells, were able to specifically transfer tolerance to nickel. A hypothesis is provided to account for this extraordinarily high frequency of nickel-reactive, suppressive T cells; it takes into account that nickel ions fail to act as classical haptens, but form versatile, unstable metal-protein and metal-peptide complexes. Furthermore, a powerful amplification mechanism, such as infectious tolerance, must operate which allows but a few donor T cells to tolerize the recipient.     

Prevalence of Incidental Pancreatic Cysts on 3 Tesla Magnetic Resonance

Objectives

To ascertain the prevalence of pancreatic cysts detected incidentally on 3-Tesla magnetic resonance imaging (MRI) of the abdomen and correlate this prevalence with patient age and gender; assess the number, location, and size of these lesions, as well as features suspicious for malignancy; and determine the prevalence of incidentally detected dilatation of the main pancreatic duct (MPD).

Methods

Retrospective analysis of 2,678 reports of patients who underwent abdominal MRI between January 2012 and June 2013. Patients with a known history of pancreatic conditions or surgery were excluded, and the remaining 2,583 reports were examined for the presence of pancreatic cysts, which was then correlated with patient age and gender. We also assessed whether cysts were solitary or multiple, as well as their location within the pancreatic parenchyma, size, and features suspicious for malignancy. Finally, we calculated the prevalence of incidental MPD dilatation, defined as MPD diameter ≥ 2.5 mm.

Results

Pancreatic cysts were detected incidentally in 9.3% of patients (239/2,583). The prevalence of pancreatic cysts increased significantly with age (p<0.0001). There were no significant differences in prevalence between men and women (p=0.588). Most cysts were multiple (57.3%), distributed diffusely throughout the pancreas (41.8%), and 5 mm or larger (81.6%). In 12.1% of cases, cysts exhibited features suspicious for malignancy. Overall, 2.7% of subjects exhibited incidental MPD dilatation.

Conclusions

In this sample, the prevalence of pancreatic cysts detected incidentally on 3T MRI of the abdomen was 9.3%. Prevalence increased with age and was not associated with gender. The majority of cysts were multiple, diffusely distributed through the pancreatic parenchyma, and ≥ 5 mm in size; 12.1% were suspicious for malignancy. An estimated 2.7% of subjects had a dilated MPD.     

Educational Inequalities in Exit from Paid Employment among Dutch Workers: The Influence of Health,Lifestyle and Work

Background

Individuals with lower socioeconomic status are at increased risk of involuntary exit from paid employment. To give sound advice for primary prevention in the workforce, insight is needed into the role of mediating factors between socioeconomic status and labour force participation. Therefore, it is aimed to investigate the influence of health status, lifestyle-related factors and work characteristics on educational differences in exit from paid employment.

Methods

14,708 Dutch employees participated in a ten-year follow-up study during 1999–2008. At baseline, education, self-perceived health, lifestyle (smoking, alcohol, sports, BMI) and psychosocial (demands, control, rewards) and physical work characteristics were measured by questionnaire. Employment status was ascertained monthly based on tax records. The relation between education, health, lifestyle, work-characteristics and exit from paid employment through disability benefits, unemployment, early retirement and economic inactivity was investigated by competing risks regression analyses. The mediating effects of these factors on educational differences in exit from paid employment were tested using a stepwise approach.

Results

Lower educated workers were more likely to exit paid employment through disability benefits (SHR:1.84), unemployment (SHR:1.74), and economic inactivity (SHR:1.53) but not due to early retirement (SHR:0.92). Poor or moderate health, an unhealthy lifestyle, and unfavourable work characteristics were associated with disability benefits and unemployment, and an unhealthy lifestyle with economic inactivity. Educational differences in disability benefits were explained for 40% by health, 31% by lifestyle, and 12% by work characteristics. For economic inactivity and unemployment, up to 14% and 21% of the educational differences could be explained, particularly by lifestyle-related factors.

Conclusions

There are educational differences in exit from paid employment, which are partly mediated by health, lifestyle and work characteristics, particularly for disability benefits. Health promotion and improving working conditions seem important measures to maintain a productive workforce, particularly among workers with a low education.     

Preconceptional folic acid‐containing supplement use in the national birth defects prevention study

Background: Despite public health campaigns encouraging women to take a daily folic acid supplement, the proportion of reproductive age women, in the United States, who comply with this recommendation is less than optimal. The objective of this analysis was to identify predictors of preconceptional folic acid‐containing supplement use to define subgroups of women who may benefit from targeted folic acid campaigns. Methods: This study included 6570 mothers of live born infants from the control population of National Birth Defects Prevention Study (1997–2005). Logistic regression analyses were used to identify predictors of preconceptional folic acid supplementation. A classification and regression tree (CART) analysis was used to define subgroups of women with different patterns of preconceptional folic acid supplementation. Results: Race/ethnicity, education, age at delivery, nativity, employment, income, number of dependents, smoking, and birth control use were significantly associated with preconceptional folic acid‐containing supplement use. Based on a CART analysis, education, race/ethnicity, and age were the most distinguishing factors between women with different preconceptional supplementation patterns. Non‐white women with <4 years of a college education were the least likely to use folic acid‐containing supplements (11%). However, even in the most compliant subgroup (women with ≥4 years of college), only 60% of women supplemented with folic acid. Conclusion: These results demonstrate the need for continued efforts to increase folic acid supplementation among all reproductive aged women. However, the success of such efforts may be improved if maternal characteristics such as education, race/ethnicity, and age, are considered in the development of future interventions. Birth Defects Research (Part A) 100:472–482, 2014. © 2014 Wiley Periodicals, Inc.     

Distinct localization of the complement C5b‐9 complex on Gram‐positive bacteria

The plasma proteins of the complement system fulfil important immune defence functions, including opsonization of bacteria for phagocytosis, generation of chemo‐attractants and direct bacterial killing via the Membrane Attack Complex (MAC or C5b‐9). The MAC is comprised of C5b, C6, C7, C8, and multiple copies of C9 that generate lytic pores in cellular membranes. Gram‐positive bacteria are protected from MAC‐dependent lysis by their thick peptidoglycan layer. Paradoxically, several Gram‐positive pathogens secrete small proteins that inhibit C5b‐9 formation. In this study, we found that complement activation on Gram‐positive bacteria in serum results in specific surface deposition of C5b‐9 complexes. Immunoblotting revealed that C9 occurs in both monomeric and polymeric (SDS‐stable) forms, indicating the presence of ring‐structured C5b‐9. Surprisingly, confocal microscopy demonstrated that C5b‐9 deposition occurs at specialized regions on the bacterial cell. On Streptococcus pyogenes, C5b‐9 deposits near the division septum whereas on Bacillus subtilis the complex is located at the poles. This is in contrast to C3b deposition, which occurs randomly on the bacterial surface. Altogether, these results show a previously unrecognized interaction between the C5b‐9 complex and Gram‐positive bacteria, whichmight ultimately lead to a new model of MAC assembly and functioning.     

A novel biosensor based on Lactobacillus acidophilus for determination of phenolic compounds in milk products and wastewater

Different branches of industry need to use phenolic compounds (PCs) in their production, so determination of PCs sensitively, accurately, rapidly, and economically is very important. For the sensitive determination of PCs, some biosensors based on pure polyphenol oxidase, plant tissue and microorganisms were developed before. But there has been no study to develop a microbial phenolic compounds biosensor based on Lactobacillus species, which contain polyphenol oxidase enzyme. In this study, we used different forms of Lactobacillus species as enzyme sources of biosensor and compared biosensor performances of these forms for determination of PCs. For this purpose, we used lyophilized Lactobacillus cells (containing L. bulgaricus, L. acidophilus, Streptococcus thermophilus), pure L. acidophilus, pure L. bulgaricus, and L. acidophilus- and L. bulgaricus adapted to catechol in Lactobacilli MRS Broth. The most suitable form was determined and optimization studies of the biosensor were carried out by using this form. For preparing the bioactive layer of the biosensor, the Lactobacillus cells were immobilized in gelatin by using glutaraldehyde. In the study, we used catechol as a substrate. Phenolic compound determination is based on the assay of the differences on the respiration activity of the cells on the oxygen meter in the absence and the presence of catechol. The microbial biosensor response depends directly on catechol concentration between 0.5 and 5.0 mM with 18 min response time. In the optimization studies of the microbial biosensor the most suitable microorganism amount was found to be 10 mg, and also phosphate buffer (pH 8.0; 50 mM) and 37.5 °C were obtained as the optimum working conditions. In the characterization studies of the microbial biosensor some parameters such as substrate specificity on the biosensor response and operational and storage stability were examine. Furthermore, the determination of PC levels in synthetic wastewater, industrial wastewater, and milk products was investigated by using the developed biosensor under optimum conditions.     

Metabolomics of prolonged fasting in humans reveals new catabolic markers

Fasting is one of the simplest metabolic challenges that can be performed in humans. We here report for the first time a comprehensive analysis of the human ??fasting metabolome?? obtained from analysis of plasma and urine samples in a small cohort of healthy volunteers, using nuclear magnetic resonance (NMR), gas chromatography- and liquid chromatography-mass spectrometry (GC-MS and LC-MS). Intra- and inter-individual variation of metabolites was on measurement of four overnight fasting samples collected from each volunteer over a four week period. One additional sample per volunteer was collected following a prolonged fasting period of 36?h. Amongst a total of 377 quantified entities in plasma around 44% were shown to change significantly in concentration when volunteers extended fasting from 12 to 36?h. In addition to known markers (plasma free fatty acids, glycerol, ketone bodies) that reflect changes in the body??s fuel management under fasting conditions a wide range of ??new?? entities such as ??-aminobutyrate as well as other amino and keto acids were identified as fasting markers. Based on multiple correlations amongst the metabolites and selected hormones in plasma such as leptin or insulin-like-growth-factor-1 (IGF-1), a robust metabolic network with coherent regulation of a wide range of metabolites could be identified. The metabolomics approach described here demonstrates the plasticity of human metabolism and identifies new and robust markers of the fasting state.     

Responses of peripheral blood flow to acute hypoxia and hyperoxia as measured by optical microangiography

Oxygen availability is regarded as a critical factor to metabolically regulate systemic blood flow. There is a debate as to how peripheral blood flow (PBF) is affected and modulated during hypoxia and hyperoxia; however in vivo evaluating of functional PBF under oxygen-related physiological perturbation remains challenging. Microscopic observation, the current frequently used imaging modality for PBF characterization often involves the use of exogenous contrast agents, which would inevitably perturb the intrinsic physiologic responses of microcirculation being investigated. In this paper, optical micro-angiography (OMAG) was employed that uses intrinsic optical scattering signals backscattered from blood flows for imaging PBF in skeletal muscle challenged by the alteration of oxygen concentration. By utilizing optical reflectance signals, we demonstrated that OMAG is able to show the response of hemodynamic activities upon acute hypoxia and hyperoxia, including the modulation of macrovascular caliber, microvascular density, and flux regulation within different sized vessels within skeletal muscle in mice in vivo. Our results suggest that OMAG is a promising tool for in vivo monitoring of functional macro- or micro-vascular responses within peripheral vascular beds.