Quantification of protein-protein interactions in highly denatured whey and potato protein gels.

Understanding the stabilizing protein interactions in protein gels is of high importance for food- and biotechnology. Protein interactions in protein gels can help to predict hardness, deformability and other gel parameters. Currently there are two types methods used. One is to use protein interaction blocking agents and the other is to dissolve the gel in different buffer systems, which cleave the interactions. The first method alters the gelling mechanism, which is why the second method is the preferred one. However, currently published methods are often only suitable for specific gel systems as for example weakly bound protein gels. In this paper, a method is introduced, which is suitable for highly denatured whey and plant protein.•Suitable for strongly cross-linked whey protein and plant protein gels•Stronger buffer system to ensure cleavage of all protein interactions•More reproducible and simplified crushing of the gel without the introduction of uncontrolled shear stress excessively affecting the analysis of chemical bonds.

Establishment of an In Vitro System of the Human Intestinal Microbiota: Effect of Cultivation Conditions and Influence of Three Donor Stool Samples.

Fecal microbiota transplantation (FMT) is an alternative method for the treatment of gastrointestinal diseases with a high recovery rate. Disadvantages are ethical concerns, high donor requirements and the low storability of stool samples. The cultivation of an in vitro microbiota in a continuous bioreactor was established as an alternative to FMT to overcome these problems. In this study, the influence of the system parameters and donor stool characteristics was investigated. Each continuous colonic fermentation system was inoculated with feces from three different donors until a stable state was established. The influence of the fermentation conditions on the system's behavior regarding cell count, metabolic activity, short-chain fatty acid profile and microbiota composition as well as richness and diversity was assessed. Cultivation conditions were found to affect the microbial system: the number of cells and the production of short-chain fatty acids increased. The abundance of Actinobacteria and Firmicutes decreased, Bacteroidetes increased, while Proteobacteria and Verrucomicrobia remained largely unaffected. Diversity in the in vitro system decreased, but richness was unaffected. The cultivation of stool from different donors revealed that the performance of the created in vitro system was similar and comparable, but unique characteristics of the composition of the original stool remained.

Differential Effects of Angiotensin-II Compared to Phenylephrine on Arterial Stiffness and Hemodynamics: A Placebo-Controlled Study in Healthy Humans.

The α1-adrenoceptor agonist phenylephrine (PE) and Angiotensin II (Ang II) are both potent vasoconstrictors at peripheral resistance arteries. PE has pure vasoconstrictive properties. Ang II, additionally, modulates central nervous blood pressure (BP) control via sympathetic baroreflex resetting. However, it is unknown whether Ang II vs. PE mediated vasoconstriction at equipressor dose uniformly or specifically modifies arterial stiffness. We conducted a three-arm randomized placebo-controlled cross-over trial in 30 healthy volunteers (15 female) investigating the effects of Ang II compared to PE at equal systolic pressor dose on pulse wave velocity (PWV), pulse wave reflection (augmentation index normalized to heart rate 75/min, AIx) and non-invasive hemodynamics by Mobil-O-Graph™ and circulating core markers of endothelial (dys-)function. PE but not Ang II-mediated hypertension induced a strong reflex-decrease in cardiac output. Increases in PWV, AIx, total peripheral resistance and pulse pressure, in contrast, were stronger during PE compared to Ang II at equal mean aortic BP. This was accompanied by minute changes in circulating markers of endothelial function. Moreover, we observed differential hemodynamic changes after stopping either vasoactive infusion. Ang II- and PE-mediated BP increase specifically modifies arterial stiffness and hemodynamics with aftereffects lasting beyond mere vasoconstriction. This appears attributable in part to different interactions with central nervous BP control including modified baroreflex function.

Assessment of platelet biology in equine patients with systemic inflammatory response syndrome.

In addition to maintaining hemostasis, platelets have an important role in modulating innate and adaptive immune responses. A low platelet count has been found to be a negative prognostic factor for survival in humans and horses with critical illnesses, such as sepsis or systemic inflammatory response syndrome (SIRS). Decreased platelet aggregation, caused by in vivo activation, has been found in human patients with severe sepsis. In our prospective controlled study, we assessed platelet biology in blood samples from 20 equine SIRS cases and 120 healthy control horses. Platelet variables such as platelet count, large platelet count, clumps, plateletcrit, mean platelet volume, and mean platelet component concentration were analyzed by laser flow cytometry (Advia 2120) from K3EDTA blood and from citrate blood. Hirudin blood samples were analyzed by impedance aggregometry (Multiplate analyzer; Roche) for platelet aggregation, including spontaneous aggregation and aggregation by 4 different agonists: adenosine diphosphate (ADPtest), ADP + prostaglandin E1 (ADPtestHS), arachidonic acid (ASPItest), and collagen (COLtest). SIRS cases had significantly lower platelet counts in K3EDTA blood (p < 0.0001) compared to control horses. There were no significant differences in aggregation values between SIRS cases and controls. Non-surviving SIRS horses did not have statistically significant lower platelet counts or lower aggregation values for COLtest, ADPtest, or ADPtestHS compared to surviving SIRS horses, although 5 non-survivors were thrombocytopenic.

Non-invasive tools beyond lung function before and after specific inhalation challenges for diagnosing occupational asthma.

PURPOSE: Increases of fractional exhaled nitric oxide (FeNO), sputum eosinophils, and methacholine responsiveness have been described after specific inhalation challenges (SIC) with occupational allergens, but limited information is available about their comparative performance. It was the aim of the study to assess the diagnostic accuracy of these non-invasive tests before and after SIC for the diagnosis of occupational asthma (OA). METHODS: A total of 122 subjects with work-related shortness of breath were included. The 'gold standard' was defined as airway obstruction (pulmonary responders) and/or an increase of FeNO of at least 13 ppb after SIC. The results were compared with those obtained using the pulmonary responder status alone as 'gold standard'. RESULTS: If the pulmonary responder status and/or an increase of FeNO was used as 'gold standard' for SIC, 28 out of 39 positives (72%), but also 20 out of 83 negatives (24%) showed an increase of sputum eosinophils and/or bronchial hyperresponsiveness after SIC. If the pulmonary responder status alone was used as 'gold standard', an increase of FeNO with a sensitivity of 0.57 and a specificity of 0.82 showed a higher accuracy than increases of sputum eosinophils (0.52/0.75) or bronchial hyperresponsiveness (0.43/0.87). Individual case analyses suggest that a few cases of OA may be detected by increases of sputum eosinophils or bronchial hyperresponsiveness alone, but probably false-positive tests dominate. CONCLUSION: It is recommended to use both lung function and increase of FeNO as primary effect parameters of SIC. Changes of sputum eosinophils and bronchial hyperresponsiveness after SIC have a low additional diagnostic value, but may be useful in individual cases.

An increase of fractional exhaled nitric oxide after specific inhalation challenge is highly predictive of occupational asthma.

PURPOSE: An increase of fractional exhaled nitric oxide (FeNO) has been described after specific inhalation challenges (SICs) with occupational allergens, but the clinical role of FeNO measurements before and after SIC is unknown. It was the aim of this study to assess the diagnostic accuracy of FeNO measurements before and after SIC in subjects with suspected occupational asthma (OA). METHODS: One hundred forty-eight patients with suspected OA were examined by SIC with various occupational allergens. Subjects were assigned to pulmonary responders, nonresponders or doubtful by standard lung function criteria. FeNO was measured before SIC (baseline) and 24 h afterwards. Subjects with negative or doubtful SIC but increase of FeNO were evaluated individually by an overall expert rating. Effect modifiers of FeNO increases were assessed by regression analyses. RESULTS: Thirty-one patients (21%) were classified as pulmonary responders, 105 (71%) as nonresponders and 12 (8%) as doubtful. With the pulmonary responder status as gold standard an increase of FeNO ≥ 13 ppb showed a specificity of 0.9 and a sensitivity of 0.5. Seventeen subjects with negative or doubtful responder status showed such an increase of FeNO, among them 13 subjects with definitive or probable OA after expert rating. Regression analyses revealed no significant modifiers for the FeNO increase. CONCLUSION: An increase of FeNO after SIC is highly predictive of OA and should be regarded as an additional criterion for the interpretation of SIC with occupational agents.

Hardiness, depression, and emotional well-being and their association with appetite in older adults.

OBJECTIVES: To examine the associations between hardiness (defined as the ability to manage stress), depression, and emotional well-being and appetite in older adults. DESIGN: Cross-sectional. SETTING: Assisted-living facilities and senior centers in the Washington/Baltimore area. PARTICIPANTS: Two hundred ninety-two adults aged 60 and older. MEASUREMENTS: Depressive symptoms assessed using the 5-item Geriatric Depression Scale and categorized as 0 to 1 (normal, referent group) versus 2 to 5 (depressive symptoms present). Hardiness was measured using the 18-item Dispositional Resilience Scale II modified based on interviews with older adults and categorized as 67 or less (low hardiness) versus greater than 67 (normal, referent group). Appetite was measured using the Simplified Nutritional Appetite Questionnaire and categorized as 4 to 14 (poor appetite) versus 15 to 20 (normal, referent group). Emotional well-being was measured using a single question. RESULTS: Depression, hardiness, and emotional well-being were all significantly associated with appetite. In models controlling for confounders (data collection site, age, educational attainment, self-reported health, race, presence of chronic disease), fair to poor emotional well-being was most significantly associated with poor appetite (odds ratio (OR)=5.60, 95% confidence interval (CI)=2.60-12.07) and low commitment (a component of hardiness that indicates an individual's involvement in life) was also significantly associated with poor appetite (OR=1.35, 95% CI=1.13-1.61). CONCLUSION: These associations further elucidate the components of mental health that contribute to poor appetite in this population. Simple measures of self-reported mental health administered to older adults may predict poor appetite and lend themselves to potential interventions to prevent malnutrition and negative health outcomes.

Identification of radiation-induced expression changes in nonimmortalized human T cells.

In the event of a radiation accident or attack, it will be imperative to quickly assess the amount of radiation exposure to accurately triage victims for appropriate care. RNA-based radiation dosimetry assays offer the potential to rapidly screen thousands of individuals in an efficient and cost-effective manner. However, prior to the development of these assays, it will be critical to identify those genes that will be most useful to delineate different radiation doses. Using global expression profiling, we examined expression changes in nonimmortalized T cells across a wide range of doses (0.15-12 Gy). Because many radiation responses are highly dependent on time, expression changes were examined at three different times (3, 8, and 24 h). Analyses identified 61, 512 and 1310 genes with significant linear dose-dependent expression changes at 3, 8 and 24 h, respectively. Using a stepwise regression procedure, a model was developed to estimate in vitro radiation exposures using the expression of three genes (CDKN1A, PSRC1 and TNFSF4) and validated in an independent test set with 86% accuracy. These findings suggest that RNA-based expression assays for a small subset of genes can be employed to develop clinical biodosimetry assays to be used in assessments of radiation exposure and toxicity.

Correlates of the desire for improved cosmetic results after breast-conserving therapy and mastectomy in breast cancer patients.

The aim of this survey was to evaluate correlates for the patient's desire for surgical improvement of the cosmetic outcome after the primary operation for breast cancer. A cross-sectional study was carried out in a single follow-up outpatient clinic using a questionnaire. Patients were asked to assess their degree of satisfaction with the cosmetic results of their primary surgery and to state if they would like to undergo a further breast surgery to improve the appearance. Patients' characteristics were correlated with this desire. After breast-conserving surgery, 21.6% of the patients stated that they desired surgical improvement, in comparison with 29.8% of the patients who underwent mastectomy. In the latter group, the desire for improvement remained constant up to 5years after the initial operation, whereas it declined in the group of patients after breast-conserving surgery. Furthermore, a younger age and the perception that the appearance negatively influences femininity, partnership or sexual life were associated with a desire for further surgery. Breast reconstruction after mastectomy can be discussed with the patients even after a long follow-up, especially when the appearance seems to influence partnership issues.

Identification of genes with abnormal expression changes in acute myeloid leukemia.

Acute myeloid leukemia (AML) is one of the most common and deadly forms of hematopoietic malignancies. We hypothesized that microarray studies could identify previously unrecognized expression changes that occur only in AML blasts. We were particularly interested in those genes with increased expression in AML, believing that these genes may be potential therapeutic targets. To test this hypothesis, we compared gene expression profiles between normal hematopoietic cells from 38 healthy donors and leukemic blasts from 26 AML patients. Normal hematopoietic samples included CD34+ selected cells (N = 18), unselected bone marrows (N = 10), and unselected peripheral bloods (N = 10). Twenty genes displayed AML-specific expression changes that were not found in the normal hematopoietic cells. Subsequent analyses using microarray data from 285 additional AML patients confirmed expression changes for 13 of the 20 genes. Seven genes (BIK, CCNA1, FUT4, IL3RA, HOMER3, JAG1, WT1) displayed increased expression in AML, while 6 genes (ALDHA1A, PELO, PLXNC1, PRUNE, SERPINB9, TRIB2) displayed decreased expression. Quantitative RT/PCR studies for the 7 over-expressed genes were performed in an independent set of 9 normal and 21 pediatric AML samples. All 7 over-expressed genes displayed an increased expression in the AML samples compared to normals. Three of the 7 over-expressed genes (WT1, CCNA1, and IL3RA) have already been linked to leukemogenesis and/or AML prognosis, while little is known about the role of the other 4 over-expressed genes in AML. Future studies will determine their potential role in leukemogenesis and their clinical significance.

Size of FLT3 internal tandem duplication has prognostic significance in patients with acute myeloid leukemia.

FLT3 internal tandem duplications (FLT3/ITDs) in the juxtamembrane domain are found in approximately 25% of acute myeloid leukemia (AML) patients, ranging in size from 3 to hundreds of nucleotides. We examined whether the sizes of FLT3/ITDs were associated with clinical outcomes in 151 AML patients enrolled in Southwest Oncology Group studies: S9333 and S9500. FLT3/ITDs were identified in 32% of patients (median ITD size = 39 nucleotides; range, 15-153 nucleotides). The CR rates were 35%, 67%, and 52% for patients with large (>or= 40), small (< 40), and no ITDs, respectively (P = .19). Increasing ITD size was associated with decreasing OS (estimated 5-year OS: large = 13%, small = 26%, and no ITD = 21%, P = .072) and RFS (estimated 5-year RFS: large = 13%, small = 27%, and no ITD = 34%, P = .017). These studies suggest that ITD size may have prognostic significance.

Angiotensin I-converting enzyme insertion/deletion polymorphism and cardiac mortality and morbidity after coronary artery bypass graft surgery.

STUDY OBJECTIVES: This study was designed to evaluate whether the insertion (I)/deletion (D) polymorphism of the angiotensin I-converting enzyme (ACE) gene is associated with mortality and cardiac morbidity after coronary artery bypass graft surgery (CABG). METHODS AND RESULTS: The ACE I/D genotype was determined in 249 consecutive patients who underwent CABG. Follow-up information (after 2 years) was obtained in 247 patients (99.2%). The primary end point was total mortality; the secondary end point was mortality from cardiac reasons, or the need for myocardial revascularization (coronary angioplasty or recurrent CABG) during follow-up. At follow-up, total mortality was 9.7% (all patients). None of the 51 patients with the ACE II genotype, 14 of 125 patients with the ACE ID genotype (11.2%), and 10 of 71 with the ACE DD genotype (14.1%) died during follow-up (p < 0.05). The ACE DD genotype, older age, diabetes mellitus, decreased left ventricular ejection fraction, and lack of internal mammary artery graft were independently related to an increased mortality after CABG. The incidence of the secondary end point was 14.5% (all patients): ACE II, 5.8%; ACE ID, 9.4%; ACE DD, 30.3% (p < 0.05). The ACE DD genotype and the presence of a left main coronary artery stenosis >or= 50% were independent predictors for the secondary end point. CONCLUSION: The ACE DD genotype is associated with increased midterm mortality and cardiac morbidity after CABG.