Comparison of a two-step Tempus600 hub solution single-tube vs. container-based, one-step pneumatic transport system.

OBJECTIVES: Efficient and timely transportation of clinical samples is pivotal to ensure accurate diagnoses and effective patient care. During the transportation process, preservation of sample integrity is crucial to avoid pre-analytical aberrations on laboratory results. Here, we present a comparative analysis between a two-step Tempus600 hub solution single-tube and a one-step, container-based pneumatic transport system (PTS) from Airco, for the in-house transportation of blood samples. METHODS: Ten blood samples from healthy volunteers were split in 10 mL collection tubes filled at full or half capacity for transportation with the two PTS (about 250 m). To compare the impact of transportation, markers of hemolysis such as lactate dehydrogenase (LDH), potassium (K+), and the hemolysis index (HI), were determined. Additionally, differences in HI in routine samples and repeated transportation was investigated. To assess and compare the mechanistic impact profiles, we recorded the acceleration profiles of the two PTS using a shock data logger. RESULTS: Transportation using the Tempus600 hub solution resulted in 49 and 46 % higher HI with samples filled to total or half capacity, respectively. Routine samples transported with the Tempus600 hub solution showed a higher median HI by 23 and 33 %. Additionally, shock logger analysis showed an elevated amount of shocks (6.5 fold) and shock intensities (1.8 fold). CONCLUSIONS: The Tempus600 hub solution caused an increased number of unreportable LDH or K+ results based on the hemolysis index. However, it was only statistically significant for LDH (p<0.01 and p<0.08) - while the comparisons for K+ were not statistically significant (p<0.28 and p<0.56).

Mapping the dynamic high-density lipoprotein synapse.

BACKGROUND AND AIMS: Heterogeneous high-density lipoprotein (HDL) particles, which can contain hundreds of proteins, affect human health and disease through dynamic molecular interactions with cell surface proteins. How HDL mediates its long-range signaling functions and interactions with various cell types is largely unknown. Due to the complexity of HDL, we hypothesize that multiple receptors engage with HDL particles resulting in condition-dependent receptor-HDL interaction clusters at the cell surface. METHODS: Here we used the mass spectrometry-based and light-controlled proximity labeling strategy LUX-MS in a discovery-driven manner to decode HDL-receptor interactions. RESULTS: Surfaceome nanoscale organization analysis of hepatocytes and endothelial cells using LUX-MS revealed that the previously known HDL-binding protein scavenger receptor B1 (SCRB1) is embedded in a cell surface protein community, which we term HDL synapse. Modulating the endothelial HDL synapse, composed of 60 proteins, by silencing individual members, showed that the HDL synapse can be assembled in the absence of SCRB1 and that the members are interlinked. The aminopeptidase N (AMPN) (also known as CD13) was identified as an HDL synapse member that directly influences HDL uptake into the primary human aortic endothelial cells (HAECs). CONCLUSIONS: Our data indicate that preformed cell surface residing protein complexes modulate HDL function and suggest new theragnostic opportunities.

Decoding Functional High-Density Lipoprotein Particle Surfaceome Interactions.

High-density lipoprotein (HDL) is a mixture of complex particles mediating reverse cholesterol transport (RCT) and several cytoprotective activities. Despite its relevance for human health, many aspects of HDL-mediated lipid trafficking and cellular signaling remain elusive at the molecular level. During HDL's journey throughout the body, its functions are mediated through interactions with cell surface receptors on different cell types. To characterize and better understand the functional interplay between HDL particles and tissue, we analyzed the surfaceome-residing receptor neighborhoods with which HDL potentially interacts. We applied a combination of chemoproteomic technologies including automated cell surface capturing (auto-CSC) and HATRIC-based ligand-receptor capturing (HATRIC-LRC) on four different cellular model systems mimicking tissues relevant for RCT. The surfaceome analysis of EA.hy926, HEPG2, foam cells, and human aortic endothelial cells (HAECs) revealed the main currently known HDL receptor scavenger receptor B1 (SCRB1), as well as 155 shared cell surface receptors representing potential HDL interaction candidates. Since vascular endothelial growth factor A (VEGF-A) was recently found as a regulatory factor of transendothelial transport of HDL, we next analyzed the VEGF-modulated surfaceome of HAEC using the auto-CSC technology. VEGF-A treatment led to the remodeling of the surfaceome of HAEC cells, including the previously reported higher surfaceome abundance of SCRB1. In total, 165 additional receptors were found on HAEC upon VEGF-A treatment representing SCRB1 co-regulated receptors potentially involved in HDL function. Using the HATRIC-LRC technology on human endothelial cells, we specifically aimed for the identification of other bona fide (co-)receptors of HDL beyond SCRB1. HATRIC-LRC enabled, next to SCRB1, the identification of the receptor tyrosine-protein kinase Mer (MERTK). Through RNA interference, we revealed its contribution to endothelial HDL binding and uptake. Furthermore, subsequent proximity ligation assays (PLAs) demonstrated the spatial vicinity of MERTK and SCRB1 on the endothelial cell surface. The data shown provide direct evidence for a complex and dynamic HDL receptome and that receptor nanoscale organization may influence binding and uptake of HDL.

Rapid and sensitive multiplex detection of COVID-19 antigens and antibody using electrochemical immunosensor-/aptasensor-enabled biochips.

We report protein- and aptamer-based electrochemical biochips for low-cost, one-step, sensitive and accurate multiplex detection of SARS-CoV-2 spike (S) and nucleocapsid (N) proteins, and IgG antibody in unprocessed clinical samples, allowing citizens to achieve rapid diagnosis at home or in community settings.

Reproducible Determination of High-Density Lipoprotein Proteotypes.

High-density lipoprotein (HDL) is a heterogeneous mixture of blood-circulating multimolecular particles containing many different proteins, lipids, and RNAs. Recent advancements in mass spectrometry-based proteotype analysis show promise for the analysis of proteoforms across large patient cohorts. In order to create the required spectral libraries enabling these data-independent acquisition (DIA) strategies, HDL was isolated from the plasma of more than 300 patients with a multiplicity of physiological HDL states. HDL proteome spectral libraries consisting of 296 protein groups and more than 786 peptidoforms were established, and the performance of the DIA strategy was benchmarked for the detection of HDL proteotype differences between healthy individuals and a cohort of patients suffering from diabetes mellitus type 2 and/or coronary heart disease. Bioinformatic interrogation of the data using the generated spectral libraries showed that the DIA approach enabled robust HDL proteotype determination. HDL peptidoform analysis enabled by using spectral libraries allowed for the identification of post-translational modifications, such as in APOA1, which could affect HDL functionality. From a technical point of view, data analysis further shows that protein and peptide quantities are currently more discriminative between different HDL proteotypes than peptidoforms without further enrichment. Together, DIA-based HDL proteotyping enables the robust digitization of HDL proteotypes as a basis for the analysis of larger clinical cohorts.

Continuous rhPTH (1-34) treatment in chronic hypoparathyroidism.

SUMMARY: Standard treatment of hypoparathyroidism consists of supplementation of calcium and vitamin D analogues, which does not fully restore calcium homeostasis. In some patients, hypoparathyroidism is refractory to standard treatment with persistent low serum calcium levels and associated clinical complications. Here, we report on three patients (58-year-old male, 52-year-old female, and 48-year-old female) suffering from severe treatment-refractory postsurgical hypoparathyroidism. Two patients had persistent hypocalcemia despite oral treatment with up to 4 µg calcitriol and up to 4 g calcium per day necessitating additional i.v. administration of calcium gluconate 2-3 times per week, whereas the third patient presented with high frequencies of hypocalcemic and treatment-associated hypercalcemic episodes. S.c. administration of rhPTH (1-34) twice daily (40 µg/day) or rhPTH (1-84) (100 µg/day) only temporarily increased serum calcium levels but did not lead to long-term stabilization. In all three cases, treatment with rhPTH (1-34) as continuous s.c. infusion via insulin pump was initiated. Normalization of serum calcium and serum phosphate levels was observed within 1 week at daily 1-34 parathyroid hormone doses of 15 µg to 29.4 µg. Oral vitamin D and calcium treatment could be stopped or reduced and regular i.v. calcium administration was no more necessary. Ongoing efficacy of this treatment has been documented for up to 7 years so far. Therefore, we conclude that hypoparathyroidism that is refractory to both conventional treatment and s.c. parathyroid hormone (single or twice daily) may be successfully treated with continuous parathyroid hormone administration via insulin pump. LEARNING POINTS: Standard treatment of hypoparathyroidism still consists of administration of calcium and active vitamin D. Very few patients with hypoparathyroidism also do not respond sufficiently to standard treatment or administration of s.c. parathyroid hormone once or twice daily. In those cases, continuous s.c. administration of parathyroid hormone via insulin pump may represent a successful treatment alternative.

'Break the Chains 2015' community-based HIV prevention campaign for men who have sex with men in Switzerland: non-randomised evaluation and cost analysis.

OBJECTIVES: To study the implementation, effects and costs of Break the Chains, a community-based HIV prevention campaign for men who have sex with men (MSM) in Switzerland, from March to May 2015, which aimed to reduce early HIV transmission by promoting the campaign message to adopt short-term risk reduction followed by HIV testing. DESIGN: Non-randomised evaluation and cost analysis. SETTING: Gay venues in 11 of 26 cantons in Switzerland and national online media campaign. PARTICIPANTS: MSM in online surveys (precampaign n=834, postcampaign n=688) or attending HIV testing centres (n=885); campaign managers (n=9); and campaign staff (n=38) or further intermediaries (n=80) in an online survey. PRIMARY AND SECONDARY OUTCOME MEASURES: The primary outcome measure was the proportion of MSM at risk of HIV acquisition or transmission who adhered to the campaign message. Secondary outcomes were postcampaign test uptake, knowledge about HIV primary infection and sense of belonging to the gay community. RESULTS: Campaign staff estimated that they contacted 17 145 MSM in 11 cantons. Among 688 respondents to the postcampaign survey, 311 (45.2%) were categorised as MSM at risk. Of 402/688 (58.5%) MSM who had heard about Break the Chains 2015, MSM categorised as being at risk were less likely to report adherence to the campaign message than MSM not at risk (adjusted OR 0.24; 95% CI 0.14 to 0.42). Twenty per cent of MSM with a defined risk of HIV acquisition or transmission who adopted risk reduction declared having done so because of the campaign. Costs for one MSM at risk to adhere to the campaign message were estimated at USD purchasing power parity 36-55. The number of HIV tests in the month after the campaign was twice the monthly average. CONCLUSION: Break the Chains increased HIV testing, implying that community-based campaigns are useful HIV prevention strategies for MSM. Additional interventions are needed to reach MSM at the highest risk of infection more effectively.

Prednisolone is associated with a worse bone mineral density in primary adrenal insufficiency.

CONTEXT: Patients with primary adrenal insufficiency (PAI) or congenital adrenal hyperplasia (CAH) receive life-long glucocorticoid (GC) therapy. Daily GC doses are often above the physiological cortisol production rate and can cause long-term morbidities such as osteoporosis. No prospective trial has investigated the long-term effect of different GC therapies on bone mineral density (BMD) in those patients. OBJECTIVES: To determine if patients on hydrocortisone (HC) or prednisolone show changes in BMD after follow-up of 5.5 years. To investigate if BMD is altered after switching from immediate- to modified-release HC. DESIGN AND PATIENTS: Prospective, observational, longitudinal study with evaluation of BMD by DXA at visit1, after 2.2 ± 0.4 (visit2) and after 5.5 ± 0.8 years (visit3) included 36 PAI and 8 CAH patients. Thirteen patients received prednisolone (age 52.5 ± 14.8 years; 8 women) and 31 patients received immediate-release HC (age 48.9 ± 15.8 years; 22 women). Twelve patients on immediate-release switched to modified-release HC at visit2. RESULTS: Prednisolone showed significantly lower Z-scores compared to HC at femoral neck (-0.85 ± 0.80 vs -0.25 ± 1.16, P < 0.05), trochanter (-0.96 ± 0.62 vs 0.51 ± 1.07, P < 0.05) and total hip (-0.78 ± 0.55 vs 0.36 ± 1.04, P < 0.05), but not at lumbar spine, throughout the study. Prednisolone dose decreased by 8% over study time, but no significant effect was seen on BMD. BMD did not change significantly after switching from immediate- to modified-release HC. CONCLUSIONS: The use of prednisolone as hormone replacement therapy results in significantly lower BMD compared to HC. Patients on low-dose HC replacement therapy showed unchanged Z-scores within the normal reference range during the study period.

BclI polymorphism of the glucocorticoid receptor and adrenal crisis in primary adrenal insufficiency.

CONTEXT: Patients with primary adrenal insufficiency (PAI) or congenital adrenal hyperplasia (CAH) are at a high risk of adrenal crisis (AC). Glucocorticoid sensitivity is at least partially genetically determined by polymorphisms of the glucocorticoid receptor (GR). OBJECTIVES: To determine if a number of intercurrent illnesses and AC are associated with the GR gene polymorphism BclI in patients with PAI and CAH. DESIGN AND PATIENTS: This prospective, longitudinal study over 37.7 ± 10.1 months included 47 PAI and 25 CAH patients. During the study period, intercurrent illness episodes and AC were documented. RESULTS: The study period covered 223 patient years in which 21 AC occurred (9.4 AC/100 pat years). There were no significant differences between BclI polymorphisms (CC (n = 29), CG (n = 34) and GG (n = 9)) regarding BMI, hydrocortisone equivalent daily dose and blood pressure. We did not find a difference in the number of intercurrent illnesses/patient year among BclI polymorphisms (CC (1.5 ± 1.4/pat year), CG (1.2 ± 1.2/pat year) and GG (1.6 ± 2.2/pat year)). The occurrence of AC was not significantly different among the homozygous (GG) genotype (32.5 AC/100 pat years), the CC genotype (6.7 AC/100 pat years) and the CG genotype (4.9 AC/100 pat years). Concomitant hypothyroidism was the highest in the GG genotype group (5/9), compared to others (CC (11/29) and CG (11/34)). CONCLUSIONS: Although sample sizes were relatively small and results should be interpreted with caution, this study suggests that the GR gene polymorphism BclI may not be associated with the frequencies of intercurrent illnesses and AC.

Immediate versus modified release hydrocortisone in mitotane-treated patients with adrenocortical cancer.

OBJECTIVE: Mitotane induces hepatic CYP3A4 activity, resulting in accelerated cortisol inactivation, and also increases cortisol binding globulin (CBG). Therefore, higher hydrocortisone doses are required in patients with adrenocortical cancer (ACC) on mitotane treatment. Modified release hydrocortisone has not been used in mitotane-treated ACC patients yet. AIM: Case series to compare serum cortisol, calculated free serum cortisol and ACTH levels in ACC patients on mitotane treatment with immediate and modified release hydrocortisone. DESIGN: Pharmacokinetics of immediate and modified release hydrocortisone, each administered at a dose of 40-20-0 mg, in nine patients with ACC and adjuvant mitotane treatment. For comparison, ten patients with secondary adrenal insufficiency (SAI) on three different hydrocortisone regimens and ten healthy males were included. METHODS: Serum cortisol and plasma ACTH were measured by chemiluminescent enzyme immunoassay, and CBG by RIA, followed by calculation of free cortisol. RESULTS: Calculated free serum cortisol levels after 40 mg immediate release hydrocortisone in ACC patients (46 ± 14 nmol/l) were similar to those after 10 mg immediate release hydrocortisone intake in men with SAI (64 ± 16 nmol/l) or to the physiological morning free cortisol levels in healthy subjects (31 ± 5 nmol/l). Compared to immediate release hydrocortisone, free cortisol levels after 40 mg modified release hydrocortisone in ACC patients were significantly lower (12 ± 3 nmol/l; P = 0·03) resulting in a generally lower AUC (98 ± 21 vs 149 ± 37 nmol h/l; P = 0·02). CONCLUSIONS: 40-20-0 mg immediate release, but not modified release hydrocortisone, resulted in sufficient glucocorticoid coverage in patients with ACC receiving mitotane treatment. The use of equivalent doses of modified release hydrocortisone preparation should be avoided in patients on mitotane treatment.

Variable RBE in proton therapy: comparison of different model predictions and their influence on clinical-like scenarios.

BACKGROUND: In proton radiation therapy a constant relative biological effectiveness (RBE) of 1.1 is usually assumed. However, biological experiments have evidenced RBE dependencies on dose level, proton linear energy transfer (LET) and tissue type. This work compares the predictions of three of the main radio-biological models proposed in the literature by Carabe-Fernandez, Wedenberg, Scholz and coworkers. METHODS: Using the chosen models, a spread-out Bragg peak (SOBP) as well as two exemplary clinical cases (single field and two fields) for cranial proton irradiation, all delivered with state-of-the-art pencil-beam scanning, have been analyzed in terms of absorbed dose, dose-averaged LET (LET D ), RBE-weighted dose (D RBE) and biological range shift distributions. RESULTS: In the systematic comparison of RBE predictions by the three models we could show different levels of agreement depending on (α/ß) x and LET values. The SOBP study emphasizes the variation of LET D and RBE not only as a function of depth but also of lateral distance from the central beam axis. Application to clinical-like scenario shows consistent discrepancies from the values obtained for a constant RBE of 1.1, when using a variable RBE scheme for proton irradiation in tissues with low (α/ß) x , regardless of the model. Biological range shifts of 0.6- 2.4 mm (for high (α/ß) x ) and 3.0 - 5.4 mm (for low (α/ß) x ) were found from the fall-off analysis of individual profiles of RBE-weighted fraction dose along the beam penetration depth. CONCLUSIONS: Although more experimental evidence is needed to validate the accuracy of the investigated models and their input parameters, their consistent trend suggests that their main RBE dependencies (dose, LET and (α/ß) x ) should be included in treatment planning systems. In particular, our results suggest that simpler models based on the linear-quadratic formalism and LETD might already be sufficient to reproduce important RBE dependencies for re-evaluation of plans optimized with the current RBE = 1.1 approximation. This approach would be a first step forward to consider RBE variations in proton therapy, thus enabling a more robust choice of biological dose delivery. The latter could in turn impact clinical outcome, especially in terms of reduced toxicities for tumors adjacent to organs at risk.

Reduction in daily hydrocortisone dose improves bone health in primary adrenal insufficiency.

OBJECTIVE: Individuals with primary adrenal insufficiency (PAI) or congenital adrenal hyperplasia (CAH) receive life-long glucocorticoid (GC) replacement therapy. Current daily GC doses are still higher than the reported adrenal cortisol production rate. This GC excess could result in long-term morbidities such as osteoporosis. No prospective trials have investigated the long-term effect of GC dose changes in PAI and CAH patients. METHODS: This is a prospective and longitudinal study including 57 subjects with PAI (42 women) and 33 with CAH (21 women). Bone mineral density (BMD) was measured by dual energy X-ray absorptiometry at baseline and after 2 years. Subjects were divided into three groups (similar baseline characteristics) depending on changes in daily hydrocortisone equivalent dose (group 1: unchanged 25.2±8.2  mg (mean±S.D., n=50); group 2: increased 18.7±10.3 to 25.9±12.0  mg (n=13); group 3: decreased 30.8±8.5 to 21.4±7.2  mg (n=27)). RESULTS: Subjects in group 1 showed normal lumbar and femoral Z-scores which were unchanged over time. Group 2 subjects showed a significant decrease in femoral neck Z-scores over time (-0.15±1.1 to -0.37±1.0 (P<0.05)), whereas group 3 subjects showed a significant increase in lumbar spine and hip Z-scores (L1-L4: -0.93±1.2 to -0.65±1.5 (P<0.05); total hip: -0.40±1.0 to -0.28±1.0 (P<0.05)). No changes in BMI over time were seen within any group. Reduction in GC dose did not increase the risk of adrenal crisis. CONCLUSION: This study demonstrates for the first time that cautious reduction in hydrocortisone equivalent doses leads to increases in BMD, whereas dose increments reduced BMD. These data emphasize the need for the lowest possible GC replacement dose in AI patients to maintain health and avoid long-term adverse effects.

Clinical implementation and range evaluation of in vivo PET dosimetry for particle irradiation in patients with primary glioma.

PURPOSE: The physical and biological properties of ion-beams offer various advantages in comparison to conventional radiotherapy, though uncertainties concerning quality assurance are still left. Due to the inverted depth dose profile, range accuracy is of paramount importance. We investigated the range deviations between planning simulation and post-fractional PET/CT measurement from particle therapy in primary glioblastoma. METHODS AND MATERIALS: 20 patients with glioblastoma undergoing particle therapy at our institution were selected. 10 received a proton-boost, 10 a carbon-ion-boost in addition to standard treatment. After two fractions, we performed a PET/CT-scan of the brain. We compared the resulting range deviation based on the Most-likely-shift method between the two measurements, and the measurements with corresponding expectations, calculated with the Monte-Carlo code FLUKA. RESULTS: A patient's two measurements deviated by 0.7mm (±0.7mm). Overall comparison between measurements and simulation resulted in a mean range deviation of 3.3mm (±2.2mm) with significant lower deviations in the (12)C-arm. CONCLUSION: The used planning concepts display the actual dose distributions adequately. The carbon ion group's results are below the used PTV safety margins (3mm). Further adjustments to the simulation are required for proton irradiations. Some anatomical situations require particular attention to ensure highest accuracy and safety.

How funding structures for HIV/AIDS research shape outputs and utilization: a Swiss case study.

BACKGROUND: Research policy in the field of HIV has changed substantially in recent decades in Switzerland. Until 2004, social science research on HIV/AIDS was funded by specialized funding agencies. After 2004, funding of such research was "normalized" and integrated into the Swiss National Science Foundation as the main funding agency for scientific research in Switzerland. This paper offers a longitudinal analysis of the relationship between the changing nature of funding structures on the one hand and the production and communication of policy-relevant scientific knowledge in the field of HIV on the other hand. METHODS: The analysis relies on an inventory of all social sciences research projects on HIV in Switzerland that were funded between 1987 and 2010, including topics covered and disciplines involved, as well as financial data. In addition, in-depth interviews were conducted with 18 stakeholders. RESULTS: The analysis highlights that the pre-2004 funding policy ensured good coverage of important social science research themes. Specific incentives and explicit promotion of social science research related to HIV gave rise to a multidisciplinary, integrative and health-oriented approach. The abolition of a specific funding policy in 2004 was paralleled by a drastic reduction in the number of social science research projects submitted for funding, and a decline of public money dedicated to such research. Although the public administration in charge of HIV policy still acknowledges the relevance of findings from social sciences for the development of prevention, treatment and care, HIV-related social science research does not flourish under current funding conditions. CONCLUSIONS: The Swiss experience sheds light on the difficulties of sustaining social science research and multidisciplinary approaches related to HIV without specialized funding agencies. Future funding policy might not necessarily require specialized agencies, but should better take into account research dynamics and motivations in the field of social sciences.

Speech-in-noise intelligibility does not correlate with efferent olivocochlear reflex in humans with normal hearing.

CONCLUSION: According to the presented data, speech-in-noise intelligibility (SI) does not correlate with olivocochlear efferent activity - as measured by contralateral suppression (CS) of distortion product otoacoustic emissions (DPOAE) in humans with normal auditory threshold. OBJECTIVES: Literature data indicate a possible role of the medial olivocochlear efferents in speech intelligibility, especially in background noise. The objective of this study was to investigate this relationship. MATERIALS AND METHODS: SI was evaluated in three independent sessions by determining the ratio speech level/noise level, at which 50% of the words are understood (i.e. speech reception threshold, SRT). Efferent activity was inferred measuring CS of DPOAE, using two different paradigms with extensive variation of stimulus parameters and duplicate measurements. RESULTS: For optimum measurement of CS, the study was restricted to subjects (n =49) with valid DPOAE down to primary tone levels L1=47/L2 =20 dB SPL. Average SRT was -6.66 dB (-4.50 to -7.65 dB, SD 0.63 dB). CS increased with decreasing primary tone levels, with mean absolute CS values in the range of 0.6-6 dB SPL. Test-retest repeatability was good. Statistical evaluation revealed no significant relationship between SI and CS of DPOAE.