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Platelets are key players in cardiovascular disease and platelet aggregation represents a central pharmacological target, particularly in secondary prevention. However, inhibition of adenosine diphosphate and thromboxane signaling has low efficacy in preventing venous thromboembolism, necessitating the inhibition of the plasmatic coagulation cascade in this disease. Anticoagulation carries a significantly higher risk of bleeding complications, highlighting the need of alternative therapeutic approaches. We hypothesized that procoagulant activation (PA) of platelets promotes venous thrombus formation and that targeting PA could alleviate venous thrombosis. Here, we found elevated levels of procoagulant platelets in the circulation of patients with deep vein thrombosis (DVT) and pulmonary embolism, and in mice developing DVT following inferior vena cava stenosis. Further, we detected procoagulant activation of recruited platelets within murine and human venous thrombi. Mice with platelet-specific deficiency in central pathways of procoagulant activation - cyclophilin D and transmembrane protein 16F - were more resistant towards low flow-induced venous thrombosis. Finally, we found that a clinically approved carbonic anhydrase inhibitor, methazolamide, reduced platelet procoagulant activity and alleviated murine thrombus formation without affecting trauma-associated hemostasis. These findings identify an essential role of platelet procoagulant function in venous thrombosis and delineate novel pharmacological strategies targeting platelets in preventing venous thromboembolism., (Copyright © 2024 American Society of Hematology.)

Background: Transcatheter aortic valve replacement (TAVR) is an effective and safe therapy for severe aortic stenosis. Rapid or fast pacing is required for implantation, which can be performed via a pre-existing cardiac implantable electric device (CIED). However, safety data on CIEDs for pacing in TAVR are missing., Objectives: The aim of this study was to elucidate procedural safety and feasibility of internal pacing with a CIED in TAVR., Methods: Patients undergoing TAVR with a CIED were included in this analysis. Baseline characteristics, procedural details, and complications according to Valve Academic Research Consortium 3 (VARC-3) criteria after TAVR were compared between both groups., Results: A total of 486 patients were included. Pacing was performed using a CIED in 150 patients and a transient pacemaker in 336 patients. No differences in technical success according to VARC-3 criteria or procedure duration occurred between the groups. The usage of transient pacers for pacing was associated with a significantly higher bleeding rate (bleeding type ≥2 according to VARC-3-criteria; 2.0% vs 13.1%; P < 0.01). Furthermore, impairment of the CIED appeared in 2.3% of patients after TAVR only in the group in which pacing was performed by a transient pacer, leading to surgical revision of the CIED in 1.3% of all patients when transient pacemakers were used., Conclusions: Internal pacing using a CIED is safe and feasible without differences of procedural time and technical success and might reduce bleeding rates. Furthermore, pacing using a CIED circumvents the risk of lead dislocation. Our data provide an urgent call for the use of a CIED for pacing during a TAVR procedure in general., Competing Interests: Funding Support and Author Disclosures Drs Steffen, Deseive, and Lackermair have received speaker honoraria from AstraZeneca. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Social determinants of health (SDOH) significantly affect individuals' health outcomes, yet universal electronic SDOH screening is not standard in primary care. Our study explores the implementation of an electronic SDOH screening in the electronic health record (EHR) and follow-up intervention among primary care pediatric patients within an academic clinic., Methods: Beginning in August of 2022, patients and their families determined to have at least one SDOH need qualified for an in-clinic referral to a coordinated care team member. We assessed the overall efficacy and feasibility of the implementation., Results: Over the 4-month pilot, 1473 of 2064 (71.4%) eligible patients were screened, with 472 (32%) patients screening positive on at least one SDOH domain. Of the 472 screened positive, 48 (10.2%) declined a referral. Two hundred and forty-seven of the 424 (58.3%) received a referral to a care coordination team member., Conclusions: This study demonstrates the feasibility of a universal electronic SDOH screening tool within the EHR within an urban, academic-based clinic., Competing Interests: Declaration of Competing Interest None., (Copyright © 2024 Academic Pediatric Association. Published by Elsevier Inc. All rights reserved.)

Background: Produce prescription programs are gaining traction in the U.S.; however, data on the impact of such approaches in pediatric populations are limited. The purpose of our clinic-based comparative effectiveness randomized controlled trial (CE RCT) is to evaluate the preliminary effectiveness of two produce prescription strategies (at-home delivery and grocery store vouchers) implemented by the Brighter Bites non-profit organization in improving obesity-related health outcomes and dietary behaviors among low-income 5-12-year-olds in Houston, Texas. This paper presents the study design, intervention components, and the study measures., Methods: Participants (n = 150) are being recruited from two pediatric clinics in Houston, Texas. Child eligibility criteria are aged 5-12 years, Medicaid recipients, body-mass index (BMI) percentile ≥85 and living within 10 miles of a Brighter Bites distribution site. Following consent and baseline measures, children are randomized into one of three arms: (1) Bi-weekly $25 vouchers redeemable for produce at stores (n = 50), (2) Bi-weekly produce delivery to participants' homes through DoorDash (n = 50), and (3) wait-list usual care controls (n = 50). Intervention participants also receive Brighter Bites nutrition education materials. Main child outcome measures are BMI z-scores, blood pressure, hemoglobin A1c, liver panels, and lipid panels. Other outcomes including household food insecurity, child diet quality, and home nutrition environment will be collected through parent surveys. Outcome measures are collected at baseline and post-intervention. Process evaluation will measure program dosage, reach, acceptability, and feasibility., Conclusions: Our paper presents the design and next steps to ensure the successful implementation of a produce prescription program in a pediatric clinic setting., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Dr. Shreela Sharma serves on the board of directors of Brighter Bites 501c3 non-profit organization. This is an unpaid board position. No other authors report any conflicts of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Long COVID is a public health emergency affecting millions of people worldwide, characterized by heterogeneous symptoms across multiple organ systems. Here, we discuss the current evidence linking thromboinflammation to postacute sequelae of COVID-19. Studies have found persistence of vascular damage with increased circulating markers of endothelial dysfunction, coagulation abnormalities with heightened thrombin generation capacity, and abnormalities in platelet counts in postacute sequelae of COVID-19. Neutrophil phenotype resembles acute COVID-19 with an increase in activation and Neutrophil Extracellular Trap formation. These insights are potentially linked by elevated platelet-neutrophil aggregate formation. This hypercoagulable state in turn can lead to microvascular thrombosis, evidenced by microclots and elevated D-dimer in the circulation as well as perfusion abnormalities in the lungs and brains of patients with long COVID. Also, COVID-19 survivors experience an increased rate of arterial and venous thrombotic events. We discuss 3 important, potentially intertwined hypotheses that might contribute to thromboinflammation in long COVID: lasting structural changes, most prominently endothelial damage, caused during initial infection; a persistent viral reservoir; and immunopathology driven by a misguided immune system. Finally, we outline the necessity for large, well-characterized clinical cohorts and mechanistic studies to clarify the contribution of thromboinflammation to long COVID., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2023 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are based on a range of novel platforms, with adenovirus-based approaches (like ChAdOx1 nCov-19) being one of them. Recently, a novel complication of SARS-CoV-2-targeted adenovirus vaccines has emerged: immune thrombocytopenia, either isolated, or accompanied by thrombosis (then termed VITT). This complication is characterized by low platelet counts, and in the case of VITT, also by platelet-activating platelet factor 4 antibodies reminiscent of heparin-induced thrombocytopenia, leading to a prothrombotic state with clot formation at unusual anatomic sites. Here, we detected antiplatelet antibodies targeting platelet glycoprotein receptors in 30% of patients with proven VITT (n = 27) and 42% of patients with isolated thrombocytopenia after ChAdOx1 nCov-19 vaccination (n = 26), indicating broad antiplatelet autoimmunity in these clinical entities. We use in vitro and in vivo models to characterize possible mechanisms of these platelet-targeted autoimmune responses leading to thrombocytopenia. We show that IV but not intramuscular injection of ChAdOx1 nCov-19 triggers platelet-adenovirus aggregate formation and platelet activation in mice. After IV injection, these aggregates are phagocytosed by macrophages in the spleen, and platelet remnants are found in the marginal zone and follicles. This is followed by a pronounced B-cell response with the emergence of circulating antibodies binding to platelets. Our work contributes to the understanding of platelet-associated complications after ChAdOx1 nCov-19 administration and highlights accidental IV injection as a potential mechanism of platelet-targeted autoimmunity. Hence, preventing IV injection when administering adenovirus-based vaccines could be a potential measure against platelet-associated pathologies after vaccination., (© 2022 by The American Society of Hematology.)

Impairment of vascular integrity is a hallmark of inflammatory diseases. We recently reported that single immune-responsive platelets migrate and reposition themselves to sites of vascular injury to prevent bleeding. However, it remains unclear how single platelets preserve vascular integrity once encountering endothelial breaches. Here we demonstrate by intravital microscopy combined with genetic mouse models that procoagulant activation (PA) of single platelets and subsequent recruitment of the coagulation cascade are crucial for the prevention of inflammatory bleeding. Using a novel lactadherin-based compound, we detect phosphatidylserine (PS)-positive procoagulant platelets in the inflamed vasculature. We identify exposed collagen as the central trigger arresting platelets and initiating subsequent PA in a CypD- and TMEM16F-dependent manner both in vivo and in vitro. Platelet PA promotes binding of the prothrombinase complex to the platelet membrane, greatly enhancing thrombin activity and resulting in fibrin formation. PA of migrating platelets is initiated by costimulation via integrin αIIbβ3 (GPIIBIIIA)/Gα13-mediated outside-in signaling and glycoprotein VI signaling, leading to an above-threshold intracellular calcium release. This effectively targets the coagulation cascade to breaches of vascular integrity identified by patrolling platelets. Platelet-specific genetic loss of either CypD or TMEM16F as well as combined blockade of platelet GPIIBIIIA and glycoprotein VI reduce platelet PA in vivo and aggravate pulmonary inflammatory hemorrhage. Our findings illustrate a novel role of procoagulant platelets in the prevention of inflammatory bleeding and provide evidence that PA of patrolling platelet sentinels effectively targets and confines activation of coagulation to breaches of vascular integrity., (© 2022 by The American Society of Hematology.)

The prevention and treatment of arterial thrombosis continue to be clinically challenging, and understanding the relevant molecular mechanisms in detail may facilitate the quest to identify novel targets and therapeutic approaches that improve protection from ischemic and bleeding events. The chemokine CXCL12 augments collagen-induced platelet aggregation by activating its receptor CXCR4. Here we show that inhibition of CXCR4 attenuates platelet aggregation induced by collagen or human plaque homogenate under static and arterial flow conditions by antagonizing the action of platelet-secreted CXCL12. We further show that platelet-specific CXCL12 deficiency in mice limits arterial thrombosis by affecting thrombus growth and stability without increasing tail bleeding time. Accordingly, neointimal lesion formation after carotid artery injury was attenuated in these mice. Mechanistically, CXCL12 activated via CXCR4 a signaling cascade involving Bruton's tyrosine kinase (Btk) that led to integrin αIIbβ3 activation, platelet aggregation, and granule release. The heterodimeric interaction between CXCL12 and CCL5 can inhibit CXCL12-mediated effects as mimicked by CCL5-derived peptides such as [VREY]4. An improved variant of this peptide, i[VREY]4, binds to CXCL12 in a complex with CXCR4 on the surface of activated platelets, thereby inhibiting Btk activation and preventing platelet CXCL12-dependent arterial thrombosis. In contrast to standard antiplatelet therapies such as aspirin or P2Y12 inhibition, i[VREY]4 reduced CXCL12-induced platelet aggregation and yet did not prolong in vitro bleeding time. We provide evidence that platelet-derived CXCL12 is involved in arterial thrombosis and can be specifically targeted by peptides that harbor potential therapeutic value against atherothrombosis., (© 2022 by The American Society of Hematology.)

Objectives: Invasive listeriosis is a severe foodborne infection caused by Listeria(L.)monocytogenes. The aim of this investigation was to verify and describe a molecular cluster of listeriosis patients and identify factors leading to this outbreak., Methods: Whole genome sequencing and core genome multilocus sequence typing were used for subtyping L. monocytogenes isolates from listeriosis cases and food samples in Germany. Patient interviews and investigational tracing of foodstuffs offered in health-care facilities (HCF), where some of the cases occurred, were conducted., Results: We identified a German-wide listeriosis outbreak with 39 genetically related cases occurring between 2014 and 2019. Three patients died as a result of listeriosis. After identification of HCF in different regions of Germany for at least 13 cases as places of exposure, investigational tracing of food supplies in six prioritized HCF revealed meat products from one company (X) as a commonality. Subsequently the outbreak strain was analysed in six isolates from ready-to-eat meat products and one isolate from the production environment of company X. No further Sigma1 cases were detected after recall of the meat products from the market and closure of company X (as of August 2020)., Conclusions: Interdisciplinary efforts including whole genome sequencing, epidemiological investigations in patients and investigational tracing of foods were essential to identify the source of infections, and thereby prevent further illnesses and deaths. This outbreak underlines the vulnerability of hospitalized patients for foodborne diseases, such as listeriosis. Food producers and HCF should minimize the risk of microbiological hazards when producing, selecting and preparing food for patients., (Copyright © 2020. Published by Elsevier Ltd.)

Several large-scale human biomonitoring projects have been conducted in Canada, including the Canadian Health Measures Survey (CHMS) and the First Nations Biomonitoring Initiative (FNBI). However, neither of these studies included participants living in the Yukon. To address this data gap, a human biomonitoring project was implemented in Old Crow, a fly-in Gwich'in community in the northern Yukon. The results of this project provide baseline levels of contaminant and nutrient biomarkers from Old Crow in 2019. Samples of hair, blood, and/or urine were collected from approximately 44% of community residents (77 of 175 adults). These samples were analyzed for contaminants (including heavy metals and persistent organic pollutants (POPs)), and nutrients (including trace elements and omega-3 fatty acids). Levels of these analytes were compared to health-based guidance values, when available, and results from other human biomonitoring projects in Canada. Levels of lead (GM 0.64 μg/g creatinine in urine/24 μg/L blood), cadmium (GM 0.32 μg/g creatinine in urine/0.85 μg/L blood), and mercury (GM < LOD in urine/0.76 μg/L blood/0.31 μg/g hair) were below select health-based guidance values for more than 95% of participants. However, compared to the general Canadian population, elevated levels of some contaminants, including lead (approximately 2× higher), cobalt (approximately 1.5× higher), manganese (approximately 1.3× higher), and hexachlorobenzene (approximately 1.5× higher) were observed. In contrast, levels of other POPs, including insecticides such as dichlorodiphenyltrichloroethane (DDT), its metabolite, dichlorodiphenyldichloroethylene (DDE), and polychlorinated biphenyls (PCBs) were similar to, or lower than, those reported in the general Canadian population. This study can be used along with future biomonitoring programs to evaluate the effectiveness of international initiatives designed to reduce the contaminant burden in the Arctic, including the Stockholm Convention and the Minamata Convention. Regionally, this project complements environmental monitoring being conducted in the region, informing local and regional traditional food consumption advisories., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Objective: Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can lead to severe pneumonia, but also thrombotic complications and non-pulmonary organ failure. Recent studies suggest intravascular neutrophil activation and subsequent immune cell-triggered immunothrombosis as a central pathomechanism linking the heterogenous clinical picture of coronavirus disease 2019 (COVID-19). We sought to study whether immunothrombosis is a pathognomonic factor in COVID-19 or a general feature of (viral) pneumonia, as well as to better understand its upstream regulation., Approach and Results: By comparing histopathological specimens of SARS-CoV-2 with influenza-affected lungs, we show that vascular neutrophil recruitment, NETosis, and subsequent immunothrombosis are typical features of severe COVID-19, but less prominent in influenza pneumonia. Activated neutrophils were typically found in physical association with monocytes. To explore this further, we combined clinical data of COVID-19 cases with comprehensive immune cell phenotyping and bronchoalveolar lavage fluid scRNA-seq data. We show that a HLADR low CD9 low monocyte population expands in severe COVID-19, which releases neutrophil chemokines in the lungs, and might in turn explain neutrophil expansion and pulmonary recruitment in the late stages of severe COVID-19., Conclusions: Our data underline an innate immune cell axis causing vascular inflammation and immunothrombosis in severe SARS-CoV-2 infection., (© 2020 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals LLC on behalf of International Society on Thrombosis and Haemostasis.)

Alveolar echinococcosis (AE) is a rare zoonotic disease caused by the larval stage of Echinococcus multilocularis. Despite its low world-wide prevalence, this disease shows differences in the regional distribution of cases. In the present cohort study, we analyse the distribution of AE according to environmental and geographical factors in Germany. We identified the place of residence of 591 cases of AE from the national database for AE, and georeferenced these localities in the Universal Transverse Mercator coordinate system. Data on elevation, air temperature, precipitation height and land cover were mapped out and correlated with the distribution of cases of disease during the period 1992-2018. Moran's I statistic was used for spatial autocorrelation. Differences in frequency distribution between elevation, air temperature, precipitation height and landscape feature classes were analysed with the Kruskal-Wallis test. With the multiple linear regression analysis, we determined the influences and interactions of geographical and climatic factors on the number of AE cases. The results showed a heterogeneous distribution of AE cases with a higher concentration in southern Germany than in the rest of Germany (I = 0.225517, Z = 35.8182 and p < 0.001). There was a statistically significant difference in frequency distribution between precipitation height, air temperature, elevation and landscape feature classes and AE cases in Germany (p < 0.0001). In regions with higher elevations (505-672 m), moderate average air temperatures (6.0-7.9°C) and higher precipitation rates (701-1000 mm) most AE cases were recorded. It seems, that regions with higher precipitation rates, higher elevations and moderate average air temperatures have a higher infection burden and infection conditions. It is therefore extremely important to generate greater awareness of the disease in these regions, with the aim of recognising potential cases of AE as early as possible and introducing the appropriate therapeutic measures., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Intercellular bridges are an essential structural feature found in both germline and somatic cells throughout the animal kingdom. Because of their large size, the germline intercellular bridges, or ring canals, in the developing fruit fly egg chamber are an excellent model to study the formation, stabilization, and growth of these structures. Within the egg chamber, the germline ring canals connect 15 supporting nurse cells to the developing oocyte, facilitating the transfer of materials required for successful oogenesis. The ring canals are derived from a stalled actomyosin contractile ring; once formed, additional actin and actin-binding proteins are recruited to the ring to support the 20-fold growth that accompanies oogenesis. These behaviors provide a unique model system to study the actin regulators that control incomplete cytokinesis, intercellular bridge formation, and growth. By temporally controlling their expression in the germline, we have demonstrated that the Arp2/3 complex and the formin, Diaphanous (Dia), coordinately regulate ring canal size and growth throughout oogenesis. Dia is required for successful incomplete cytokinesis and the initial stabilization of the germline ring canals. Once ring canals have formed, the Arp2/3 complex and Dia cooperate to determine ring canal size and maintain stability. Our data suggest that nurse cells must maintain a precise balance between the activity of these two nucleators during oogenesis., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Objective: We describe epidemiological trends of human brucellosis in Germany over a 13 year period based on national surveillance data., Methods: We analyzed demographic, clinical, laboratory and exposure information of symptomatic laboratory-confirmed brucellosis cases notified 2006-18. Using official population data, we calculated incidences and risk ratios (RR)., Results: From 2006 to 2018, 408 brucellosis cases were notified in Germany (mean annual incidence: 0.38/1,000,000 population), of which 75% were travel-associated. Yearly notifications peaked in 2014 (n = 47) and remained elevated compared to 2006-2013 (mean: n = 25). Asylum seekers (AS) arriving in Germany accounted for 9/44 (2015) and 15/36 (2016) cases, respectively. RR AS/non-AS 2015-2016: 28, 95% CI: 17-45. Unpasteurized milk products were most frequently notified as source of infection. Imported food and occupational exposure played a role in autochthonous cases., Conclusions: The incidence of human brucellosis has markedly increased in recent years. Most of the observed rise in notifications can be explained by infections in AS. Exposure still predominantly occurs abroad. Risk factors for autochthonous infections need to be investigated further, though imported dairy products seem to play a role. Physicians should consider brucellosis as differential diagnosis in AS and people with travel to endemic regions with compatible symptoms., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Clinical observations implicate a role of eosinophils in cardiovascular diseases because markers of eosinophil activation are elevated in atherosclerosis and thrombosis. However, their contribution to atherosclerotic plaque formation and arterial thrombosis remains unclear. In these settings, we investigated how eosinophils are recruited and activated through an interplay with platelets. Here, we provide evidence for a central importance of eosinophil-platelet interactions in atherosclerosis and thrombosis. We show that eosinophils support atherosclerotic plaque formation involving enhanced von Willebrand factor exposure on endothelial cells and augmented platelet adhesion. During arterial thrombosis, eosinophils are quickly recruited in an integrin-dependent manner and engage in interactions with platelets leading to eosinophil activation as we show by intravital calcium imaging. These direct interactions induce the formation of eosinophil extracellular traps (EETs), which are present in human thrombi and constitute a substantial part of extracellular traps in murine thrombi. EETs are decorated with the granule protein major basic protein, which causes platelet activation by eosinophils. Consequently, targeting of EETs diminished thrombus formation in vivo, which identifies this approach as a novel antithrombotic concept. Finally, in our clinical analysis of coronary artery thrombi, we identified female patients with stent thrombosis as the population that might derive the greatest benefit from an eosinophil-inhibiting strategy. In summary, eosinophils contribute to atherosclerotic plaque formation and thrombosis through an interplay with platelets, resulting in mutual activation. Therefore, eosinophils are a promising new target in the prevention and therapy of atherosclerosis and thrombosis., (© 2019 by The American Society of Hematology.)

Background: Pasireotide (SOM230; Novartis Inc, Basel, Switzerland) is a multitargeted somatostatin receptor analogue likely to treat the neuroendocrine, and docetaxel resistant components within metastatic castrate-resistant prostate cancer (mCRPC). This phase I trial tested the combination of pasireotide, docetaxel, and prednisone in pretreated mCRPC., Patients and Methods: Chemotherapy naive mCRPC patients received docetaxel 75 mg/m 2 intravenously every 21 days and pasireotide intramuscularly every 28 days at escalating dose levels of 40, 60, and 80 mg. Maximum tolerated dose and recommended phase II dose (RP2D) were assessed., Results: Eighteen patients were enrolled with a median age of 65 (range, 49-75) years, and pretherapy prostate-specific antigen of 259.9 ng/mL. The dose-limiting toxicities were Grade 4 hyperglycemia unresponsive to therapy and Grade 4 neutropenia lasting for > 7 days in 1 patient each occurring at the 80-mg dose level of pasireotide. The RP2D was determined at 60 mg every 28 days. Four patients at the 60 mg dose had Grade 3 or 4 hyperglycemia, which responded adequately to therapy. Median time to progression and survival were 7.2 and 18.3 months, respectively. Three of 6 patients with circulating tumor cells ≥5 converted to circulating tumor cells < 5 post therapy. The insulin like growth factor-1 levels revealed a median 51% decrease after therapy. The neuron-specific enolase and chromogranin did not show any marked change., Conclusion: The addition of pasireotide to docetaxel and prednisone is clinically feasible at a dose level of 60 mg every 28 days. The combination showed potential for clinical efficacy but needs to be compared with the standard docetaxel and prednisone regimen., (Copyright © 2018 Elsevier Inc. All rights reserved.)

As classically described, Eagle syndrome is an entity where patients develop pain or neurologic manifestations arising from an elongated styloid process and/or an ossified stylohyoid ligament irritating or compressing adjacent cranial nerves or the carotid arteries. Over the past few years, there have been reports of actual injury to the internal carotid artery with dissection, occlusion, and strokes. We present 3 cases identified after blunt trauma: 1 due to carotid compression and 2 due to actual injury to the internal carotid artery. Eagle syndrome should be a consideration in any patient with a carotid injury due to blunt trauma or suffering a syncopal episode which led to blunt trauma. Carotid stenting is an effective treatment modality for injury to the carotid artery when anticoagulation is contraindicated. Styloidectomy is performed for symptoms due to carotid artery compression or if there is concern for future carotid injury from the styloid process., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Deep venous thrombosis (DVT) is one of the most common cardiovascular diseases, but its pathophysiology remains incompletely understood. Although sterile inflammation has recently been shown to boost coagulation during DVT, the underlying molecular mechanisms are not fully resolved, which could potentially identify new anti-inflammatory approaches to prophylaxis and therapy of DVT. Using a mouse model of venous thrombosis induced by flow reduction in the vena cava inferior, we identified blood-derived high-mobility group box 1 protein (HMGB1), a prototypical mediator of sterile inflammation, to be a master regulator of the prothrombotic cascade involving platelets and myeloid leukocytes fostering occlusive DVT formation. Transfer of platelets into Hmgb1 -/- chimeras showed that this cell type is the major source of HMGB1, exposing reduced HMGB1 on their surface upon activation thereby enhancing the recruitment of monocytes. Activated leukocytes in turn support oxidation of HMGB1 unleashing its prothrombotic activity and promoting platelet aggregation. This potentiates the amount of HMGB1 and further nurtures the accumulation and activation of monocytes through receptor for advanced glycation end products (RAGE) and Toll-like receptor 2, leading to local delivery of monocyte-derived tissue factor and cytokines. Moreover, disulfide HMGB1 facilitates formation of prothrombotic neutrophil extracellular traps (NETs) mediated by RAGE, exposing additional HMGB1 on their extracellular DNA strands. Eventually, a vicious circle of coagulation and inflammation is set in motion leading to obstructive DVT formation. Therefore, platelet-derived disulfide HMGB1 is a central mediator of the sterile inflammatory process in venous thrombosis and could be an attractive target for an anti-inflammatory approach for DVT prophylaxis., (© 2016 by The American Society of Hematology.)

Purpose: Malnutrition is common in residential care environments, primarily due to poor intake. Micronutrient deficiency, although poorly investigated to date, is also reported to be high. Improving the nutrient density of consumed foods is a potential mechanism to promote increased nutrient intake. A scoping review was conducted to: (1) explore the evidence on micronutrient food fortification strategies, (2) identify candidate nutrients and food vehicles for successful food fortification, and (3) identify gaps for future research., Methods: The scoping review framework of Arksey and O'Malley was used. A comprehensive search strategy of 4 electronic databases (MEDLINE, EMBASE, CINAHL, and Web of Science) was completed. Two reviewers were involved in screening and data extraction for all selected articles., Results: A total of 4394 relevant articles were identified for screening, and application of inclusion/exclusion criteria resulted in 6 food fortification studies (8 citations; 1 study had 3 citations). Overall, vitamin D (n = 5 studies) and calcium (n = 4 studies) were the most common micronutrients fortified; milk products, margarine, bread, and pureed foods were fortification vehicles. Most studies fortified below the RDA recommendation and did not include clinical outcomes. Samples were small and intervention periods were short (3-6 months)., Conclusions: Fortification is a viable strategy for improving the nutrient density of foods consumed in residential care. Although disparate, this literature suggests the potential for further undertaking of fortification to prevent micronutrient deficiencies among residents and future research should consider multinutrient preparations and clinical outcomes., (Copyright © 2016 AMDA – The Society for Post-Acute and Long-Term Care Medicine. Published by Elsevier Inc. All rights reserved.)

Background and Aims: Recent gene-environment interaction studies suggest that diet may influence an individual's genetic predisposition to cardiovascular risk. We evaluated whether omega-3 fatty acid intake may influence the risk for acute coronary syndrome (ACS) conferred by genetic polymorphisms among patients with early onset ACS., Methods and Results: Our population consisted of 705 patients of white European descent enrolled in GENESIS-PRAXY, a multicenter cohort study of patients aged 18-55 years and hospitalized with ACS. We used a case-only design to investigate interactions between the omega-3 index (a validated biomarker of omega-3 fatty acid intake) and 30 single nucleotide polymorphisms (SNPs) robustly associated with ACS. We used logistic regression to assess the interaction between each SNP and the omega-3 index. Interaction was also assessed between the omega-3 index and a genetic risk score generated from the 30 SNPs. All models were adjusted for age and sex. An interaction for increased ACS risk was found between carriers of the chromosome 9p21 variant rs4977574 and low omega-3 index (OR 1.57, 95% CI 1.07-2.32, p = 0.02), but this was not significant after correction for multiple testing. Similar results were obtained in the adjusted model (OR 1.55, 95% CI 1.05-2.29, p = 0.03). We did not observe any interaction between the genetic risk score or any of the other SNPs and the omega-3 index., Conclusion: Our results suggest that omega-3 fatty acid intake may modify the genetic risk conferred by chromosome 9p21 variation in the development of early onset ACS and requires independent replication., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Objectives: This study aimed to assess the effect of curing conditions--exposure time, mode, energy density, and exposure distance--on the mechanical properties of high-viscosity bulk-fill resin-based composites (RBCs) measured at simulated clinical relevant filling depths., Methods: Three high-viscosity bulk-fill RBCs were investigated by assessing the variation in micromechanical properties in 200μm steps (Vickers hardness [HV] and indentation modulus [E]) within simulated 6-mm deep fillings (n=5) polymerized under 16 different curing conditions. The exposure duration was 5, 20, and 40s in the standard power mode; 3, 4, and 8s in the high power mode; and 3 and 6s in the plasma mode; the exposure distance was 0 and 7mm. Energy density ranged from 2.63 to 47.03J/cm(2). Measurements were performed after 24h of storage in distilled water at 37°C. The depth of cure (DOC) was calculated as the 80% hardness drop-off., Results: The results were compared using one- and multiple-way ANOVAs and Tukey's HSD post hoc test (α=0.05). The effect of the parameter material was significant and strong on all measured properties (p<0.05, partial eta-squared ηP(2)=0.492 for E, 0.562 for HV, and 0.087 for DOC). Energy density exerted the strongest influence on the measured properties in all materials, whereas the influence of the exposure distance was strong on DOC, low on E and not significant on HV. The high-viscosity bulk-fill RBCs respond heterogeneously to variations in curing conditions., Conclusions: A lower energy density limit was identified, allowing for a 4mm material bulk placement (5.88J/cm(2) for EvoCeram Bulk Fill, 7.0J/cm(2) for x-tra fil, and 23.51J/cm(2) for SonicFill). This limit increased to 47.03J/cm(2) for a 5mm bulk placement, as claimed for SonicFill. To maintain mechanical properties in depth, however, an energy density of at least 23.51J/cm(2) is recommended for EvoCeram Bulk Fill and x-tra fil and 47.03J/cm(2) for SonicFill, respectively. This energy density should be achieved at moderate irradiance and increased curing time., Clinical Significance: An exposure time of 20s at moderate irradiance is recommended for all materials for a 4 mm bulk placement., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Lyme borreliosis (LB) is the most frequent vector-borne disease in Germany. For more than 10 years, data from mandatory notifications have been available from 6 federal states in the eastern part of Germany. A common case definition was applied. Clinical manifestations of erythema migrans, neuroborreliosis (radiculoneuritis, cranial neuritis, meningitis), and Lyme arthritis were notifiable. From 2009 to 2012, altogether 18,894 cases were notified. The overall incidence varied between 34.9 cases/100,000 inhabitants in 2009 and 19.54 cases/100,000 persons in 2012. LB in eastern Germany showed a pronounced seasonality with a peak in August. Decreasing as well as increasing trends were observed in different federal states. Females predominated among all cases (55.3%). The age distribution was bimodal with incidence peaks in children 5-9 years old (32.4 cases/100,000 persons in 2011) and in adults aged 60-69 years (56.7 cases/100,000 persons in 2011). Erythema migrans affected 95.4% of the patients and acute neuroborreliosis 3.3%. Among the latter, the most common manifestation was radiculoneuritis (n=316). Neuritis cranialis was more common in children than in adults (p<0.01). The same was true for meningitis (p<0.01). Altogether 2.0% of the LB cases developed Lyme arthritis. LB has a significant disease burden in the study area. Different levels of under-ascertainment in the surveillance system could explain parts of the differences in the incidence. Furthermore, there may be discrepancies in disease awareness among patients and physicians. Changes in time and differences among geographical regions could result from variations in risk factors related to human behaviour (e.g., outdoor activity). Additionally, vector-related risk factors may have varied (e.g., landscape, climate). Public health strategies with a particular focus on the high-incidence age groups should promote daily checks for ticks and prompt removal of ticks after exposure to avoid infection. Physicians should be able to recognize LB patients with early manifestations and promptly treat those appropriately., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

In vitro studies suggest that leukocytes locomote in an ameboid fashion independently of pericellular proteolysis. Whether this motility pattern applies for leukocyte migration in inflamed tissue is still unknown. In vivo microscopy on the inflamed mouse cremaster muscle revealed that blockade of serine proteases or of matrix metalloproteinases (MMPs) significantly reduces intravascular accumulation and transmigration of neutrophils. Using a novel in vivo chemotaxis assay, perivenular microinjection of inflammatory mediators induced directional interstitial migration of neutrophils. Blockade of actin polymerization, but not of actomyosin contraction abolished neutrophil interstitial locomotion. Multiphoton laser scanning in vivo microscopy showed that the density of the interstitial collagen network increases in inflamed tissue, thereby providing physical guidance to infiltrating neutrophils. Although neutrophils locomote through the interstitium without pericellular collagen degradation, inhibition of MMPs, but not of serine proteases, diminished their polarization and interstitial locomotion. In this context, blockade of MMPs was found to modulate expression of adhesion/signaling molecules on neutrophils. Collectively, our data indicate that serine proteases are critical for neutrophil extravasation, whereas these enzymes are dispensable for neutrophil extravascular locomotion. By contrast, neutrophil interstitial migration strictly relies on actin polymerization and does not require the pericellular degradation of collagen fibers but is modulated by MMPs.

Objective: Genome-wide association studies identified a risk haplotype on chromosome 9p21.3 to be associated with coronary artery disease (CAD) and myocardial infarction (MI). Since this region does not contain a clear candidate gene with known pathophysiology, we performed a haplotype-specific expression study in human macrophages during pro-inflammatory stimulation to investigate the locus-dependent expression patterns in a model of atherosclerosis, the underlying cause for CAD and MI., Methods: Blood samples were taken from 40 male stable MI patients either homozygous for 9p21.3 risk (n = 20) or non-risk haplotype (n = 20) as well as from 28 healthy male individuals (n = 14 for each haplotype). Monocytes were isolated by density gradient centrifugation followed by differentiation into macrophages via M-CSF. Macrophages were either incubated with a pro-inflammatory IFNγ-LPS cocktail or kept untreated as controls. After 24 h, RNA was isolated and applied to Affymetrix Human Exon 1.0 ST Arrays., Results: Macrophages from MI patients and controls stratified for 9p21.3 haplotypes, exhibited marked differences in gene expression. Most pronounced differences were found in inflammatory mediators, like the chemokines CCL8 and CCL2 and the lectines CLEC4E and CLEC5A. Differences in expression changes could be seen most obviously during inflammatory stimulation for both, the interleukins IL12B and IL1B, and members of metallothionein gene family., Conclusion: These findings show that gene expression is different in 9p21.3 haplotype-stratified macrophages. While these effects are relatively small in our in vitro model of atherosclerosis, these biological effects may contribute to a long term effect in risk haplotype carriers increasing susceptibility to CAD and MI., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

In parallel with a renewed interest in nuclear power and its possible environmental impacts, a new environmental radiation protection system calls for environmental indicators of radiological stress. However, because environmental stressors seldom occur alone, this study investigated the combined effects of an ecological stressor (larval density) and an anthropogenic stressor (ionizing radiation) on amphibians. Scaphiopus holbrookii tadpoles reared at different larval densities were exposed to four low irradiation dose rates (0.13, 2.4, 21, and 222 mGy d(-1)) from (137)Cs during the sensitive period prior to and throughout metamorphosis. Body size at metamorphosis and development rate served as fitness correlates related to population dynamics. Results showed that increased larval density decreased body size but did not affect development rate. Low dose rate radiation had no impact on either endpoint., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: A single nucleotide polymorphism (SNP) rs599839 located at chromosome 1p13.3 has previously been associated with risk of coronary artery disease (CAD) and with serum levels of low-density lipoprotein cholesterol (LDL-C). A functional link explaining the association of SNP rs599839 with LDL-C levels and CAD risk has not yet been elucidated., Methods: We analyzed the association of rs599839 with LDL-C in 6605 individuals across a wide age spectrum and with CAD in four case-control studies comprising 4287 cases and 7572 controls. Genome-wide expression array data was used to assess the association of SNP rs599839 with gene expression at chromosome 1p13. Finally, we overexpressed sortilin in transfected cells to study LDL-uptake in vitro., Results: Each copy of the G-allele of rs599839 associated with a decrease of serum LDL-C by 0.14 mmol/L (90% confidence interval (CI) 0.09-0.17 mmol/L, p=2.6 x 10(-11)). Moreover, each copy of the G-allele associated with a 9% decrease of CAD risk (90% CI 4-14%) in the presently studied four case-control samples and with a 13% decrease (90% CI 10-17%, p=2.18 x 10(-9)) in a pooled meta-analysis including recent genome-wide association studies on CAD. The same allele was associated with higher mRNA-expression levels of the multiligand receptor sortilin (log transformed mRNA AA vs. GG=8.31 vs. 8.55; p=0.01). Overexpression of SORT1 cDNA resulted in a significant increase in LDL-particle uptake (+23%, p=0.01)., Conclusions: Rs599839 associates with decreased LDL-C and a lower risk of CAD. Effects appear to be mediated by increased sortilin expression and subsequently enhanced LDL-uptake into cells., (Copyright (c) 2009 Elsevier Ireland Ltd. All rights reserved.)

The HDL and LDL subclass profile is an emerging cardiovascular risk factor. Yet, the biological and genetic mechanisms controlling the lipoprotein subclass distribution are unclear. Therefore, we aimed 1) to determine the heritability of the entire spectrum of LDL and HDL subclass features and 2) to identify gene loci influencing the lipoprotein subfraction pattern. Using NMR spectroscopy, we analyzed the lipoprotein subclass distribution in 1,275 coronary artery disease patients derived from the Regensburg Myocardial Infarction Family Study. We calculated heritabilities, performed a microsatellite genome scan, and calculated linkage. HDL and LDL subclass profiles showed heritabilities ranging from 23% to 67% (all P < 10(-3)) of traits using univariate calculation. After multivariate adjustment, we found heritabilities of 27-48% (all P < 0.05) for HDL and 21-44% for LDL traits. The linkage analysis revealed a significant logarithm of the odds (LOD) score (3.3) for HDL particle concentration on chromosome 18 and a highly suggestive signal for HDL particle size on chromosome 12 (2.9). After multivariate adjustment, we found a significant maximum LOD score of 3.7 for HDL size. Our study is the first to analyze heritability and linkage for the entire spectrum of LDL and HDL subclass features. Our findings may lead to the identification of genes controlling the lipoprotein subclass distribution.

A novel human cytochrome P450, CYP2W1, was cloned and expressed heterologously. No or very low CYP2W1 mRNA levels were detected in fetal and adult human tissues, expression was however seen in 54% of human tumor samples investigated (n=37), in particular colon and adrenal tumors. Western blotting also revealed high expression of CYP2W1 in some human colon tumors. In rat tissues, CYP2W1 mRNA was expressed preferentially in fetal but also in adult colon. The CYP2W1 gene was shown to encompass one functional CpG island in the exon 1-intron 1 region which was methylated in cell lines lacking CYP2W1 expression, but unmethylated in cells expressing CYP2W1. Re-expression of CYP2W1 was seen following demethylation by 5-Aza-2'-deoxycytidine. Transfection of HEK293 cells with CYP2W1 caused the formation of a properly folded enzyme, which was catalytically active with arachidonic acid as a substrate. It is concluded that CYP2W1 represents a tumor-specific P450 isoform with potential importance as a drug target in cancer therapy.

Recombinant CYP4F8 and CYP4F12 metabolize prostaglandin H2 (PGH2) analogs by omega2- and omega3-hydroxylation and arachidonic acid (20:4n-6) by omega3-hydroxylation. CYP4F8 was found to catalyze epoxidation of docosahexaenoic acid (22:6n-3) and docosapentaenoic acid (22:5n-3) and omega3-hydroxylation of 22:5n-6. CYP4F12 oxidized 22:6n-3 and 22:5n-3 in the same way, but 22:5n-6 was a poor substrate. The products were identified by liquid chromatography-mass spectrometry. The missense mutation 374A>T of CYP4F8 (Tyr125Phe in substrate recognition site-1 (SRS-1)) occurs in low frequency. This variant oxidized two PGH2 analogs, U-51605 and U-44069, in analogy with CYP4F8, but 20:4n-6 and 22:5n-6 were not oxidized. CYP4F enzymes with omega-hydroxylase activity contain a heme-binding Glu residue, whereas CYP4F8 (and CYP4F12) with omega2- and omega 3-hydroxylase activities has a Gly residue in this position of SRS-4. The mutant CYP4F8 Gly328Glu oxidized U-51605 and U-44069 as recombinant CYP4F8, but the hydroxylation of arachidonic acid was shifted from C-18 to C-19. Single amino acid substitutions in SRS-1 and SRS-4 of CYP4F8 may thus influence oxygenation of certain substrates. We conclude that CYP4F8 and CYP4F12 catalyze epoxidation of 22:6n-3 and 22:5n-3, and CYP4F8 omega3-hydroxylation of 22:5n-6.

Arachidonic acid is oxidized by cytochromes P450 2C (CYP2C) to epoxyeicosatrienoic acids (EETs), possessing vasoactive properties, with 11,12-EET as the endothelium derived hyperpolarization factor. Genetic variants of CYP2C enzymes have altered drug metabolizing capacity. Our primary aim was to determine whether EET biosynthesis differed in human liver microsomes with known CYP2C genotypes. Human liver microsomes (n = 25) of different CYP2C genotypes or yeast-expressed CYP2C enzymes were used. Analysis of metabolites was performed by liquid chromatography/mass spectrometry. Samples genotyped as CYP2C8*3/*3/CYP2C9*2/*2 exhibited a 34% (p < 0.05) decreased EET biosynthesis, compared to other CYP2C8/CYP2C9 haplotypes. Inhibition experiments suggested CYP2C8 and CYP2C9 to be the predominant catalysts of EETs. We found no differences between the three recombinantly expressed CYP2C9 variants, but CYP2C8.1 had lower K(m) than these isoforms. In conclusion, there are genetic differences in the CYP2C-dependent oxidation of arachidonic acid to vasoactive metabolites, of which the relevance to cardiovascular pathophysiology is still unclear.

Our aim was to determine the tissue distribution of CYP4F8, which occurs in human seminal vesicles and catalyzes 19-hydroxylation of prostaglandin H(1) and H(2) in vitro (J. Bylund, M. Hidestrard, M. Ingelman-Sundberg, E.H. Oliw, J. Biol. Chem. 275 (2000) 21844-21849). Polyclonal antibodies were raised in rabbits against RVEPLG, the C-terminal end of CYP4F8, and purified by affinity chromatography. Screening of 50 human tissues for CYP4F8 immunoreactivity revealed protein expression, inter alia, in seminal vesicles, epidermis, hair follicles, sweat glands, corneal epithelium, proximal renal tubules, and epithelial linings of the gut and urinary tract. The CYP4F8 transcripts were detected by reverse transcription polymerase chain reaction and by Northern blot analysis. There was a prominent induction of CYP4F8 immunoreactivity and mRNA in psoriasis in comparison with unaffected epidermis of the same patients. The cDNA of CYP4F8 from plucked scalp hair roots was identical with the genital cDNA sequence. We conclude that CYP4F8 is present in epithelial linings and up regulated in epidermis of psoriatic lesions.

We identified a novel erythropoietin (Epo)-induced protein (CIP29) in lysates of human UT-7/Epo leukemia cells using two-dimensional gel analysis and cloned its full-length cDNA. CIP29 contains 210 amino acids with a predicted MW of 24 kDa, and has a N-terminal SAP DNA-binding motif. CIP29 expression was higher in cancer and fetal tissues than in normal adult tissues. CIP29 mRNA expression is cytokine regulated in hematopoietic cells, being up-regulated by Epo in UT7/Epo cells, and by thrombopoietin (Tpo), FLT3 ligand (FL) and stem cell factor (SCF) in primary human CD34(+) cells. Up-regulation of CIP29 in UT7/Epo cells by Epo was associated with cell cycle progression but not with antiapoptosis. Epo withdrawal reduced CIP29 expression concomitant with cell cycle arrest. Overexpression of CIP29-GFP in HEK293 cells enhances cell cycle progression. CIP29 appears to be a new cytokine regulated protein involved in normal and cancer cell proliferation.

Solid-organ transplant recipients are at increased risk of various infectious diseases, some of which are vaccine preventable mmunisations are among the most efficient interventions available. Solid-organ tranplant recipients would greatly benefit from effective immunisations, provided the recommendations are based on a careful risk-benefit analysis in which the effectiveness of the vaccine is weighed against possible adverse reactions, including graft rejection. In this review, we summarise the data from studies on relevant immunisations in solid-organ transplant recipients. The major issues are the immunogenicity and safety of immunisations, the factors associated with poor immune response, and recommendations for immunisation schemes.

The Alcohol and Drug Addiction Treatment and Support Act (ADATSA) created a treatment program for indigent clients in Washington State. This research assesses the relationship between the level of treatment services received and subsequent employment outcomes. Clients who completed their plan of treatment earned more than those who did not, controlling for other factors. Those clients who received vocational services, in addition to completing treatment, earned more than those who completed treatment only. While on average wages were low, this study does show that clients once deemed 'unemployable' can become productive.

Background: n-3 Fatty acid supplementation lowered serum triacylglycerol concentrations in studies in which most of the subjects were male. The effects of n-3 fatty acid supplementation in postmenopausal women receiving and not receiving hormone replacement therapy (HRT) have received little attention., Objective: We sought to determine the effects of a fish-oil-derived n-3 fatty acid concentrate on serum lipid and lipoprotein risk factors for cardiovascular disease in postmenopausal women receiving and not receiving HRT, with an emphasis on serum triacylglycerol concentrations and the ratio of triacylglycerol to HDL cholesterol., Design: Postmenopausal women (n = 36) were grouped according to exogenous hormone use and were randomly allocated to receive 8 capsules/d of either placebo oil (control) or n-3 fatty acid-enriched oil (supplement). The supplement provided 2.4 g eicosapentaenoic acid (EPA) plus 1.6 g docosahexaenoic acid (DHA) daily. Serum lipids and the fatty acid composition of serum phospholipids were determined on days 0 and 28., Results: Supplementation with n-3 fatty acids was associated with 26% lower serum triacylglycerol concentrations (P < 0.0001), a 28% lower overall ratio of serum triacylglycerol to HDL cholesterol (P < 0.01), and markedly greater EPA and DHA concentrations in serum phospholipids (P < 0.05)., Conclusions: These results show that supplementation with a fish-oil-derived concentrate can favorably influence selected cardiovascular disease risk factors, particularly by achieving marked reductions in serum triacylglycerol concentrations and triacylglycerol:HDL cholesterol in postmenopausal women receiving and not receiving HRT. This approach could potentially reduce the risk of coronary heart disease by 27% in postmenopausal women.

Gene targeting has proven to be extremely powerful in various fields of biological research. Through this technique, knockout mice lacking a particular gene, and thus a particular protein, can be generated. One limitation to this technique is the fact that mice develop without the protein of interest and therefore, developmental compensations may have taken place, contributing to an observed phenotype. Inducible strategies, those which allow the timing of expression of a gene to be regulated, are currently being developed and should prove useful when applied to gene targeting technology. To begin to apply such new technologies to the field of gene targeting, we first created and tested several reporter constructions using the tetracycline inducible system. Here we describe the creation of several beta-galactosidase reporter constructions and the results of in vitro testing in Cos-7 cells. We then discuss future knockout strategies based upon our observations.

We have developed a new procedure utilizing microhomologous recombination in yeast to generate targeting constructs for producing targeted mutations in mice. This procedure is rapid and efficient, and should be directly applicable to all mammalian genes. Moreover, only minimal information about the locus being targeted is required. The feasibility of this approach was demonstrated by producing another allele of the mouse Tg737 polycystic kidney gene., (Copyright 1998 Elsevier Science B.V. All rights reserved.)

Background/aims: The hepatitis C virus genotypes have been shown to be differently distributed between distinct geographical areas and to be associated with different clinical presentations. In the present study we investigated the distribution of HCV genotypes in 379 German patients with chronic hepatitis C in relation to age, sex, route of infection, liver histology and viremia., Methods: Typing of HCV was done using restriction fragment length polymorphism analysis as well as a DNA enzyme immunoassay. HCV RNA concentrations were determined by quantitative polymerase chain reaction. Liver biopsies were performed in 187 patients and the histological activity was graded by the Knodell score., Results: Seventy percent were infected with genotype 1 (20% subtype 1a, 80% subtype 1b), 4% with genotype 2 and 26% with genotype 3 (all subtype 3a). Genotype 3a and 1a infection was significantly associated with intravenous drug abuse. In contrast, genotype 1 predominated in patients with post-transfusion hepatitis and infection of unknown origin. A changing relative prevalence of HCV genotypes in relation to age was also observed. Patients with genotype 3 infection showed significantly lower HCV RNA levels and a lower mean histological activity score as compared to patients with genotype 1 and genotype 2. However, using multivariate analysis, only age and mode of transmission but not histological activitiy score were shown to be independent variables., Conclusions: Our study confirms previous reports from other countries that HCV variants can be classified into a relatively small number of discrete genotypes, and that the subtype 1b clearly predominates. However, we found evidence that there is a changing relative prevalence of HCV genotypes in relation to age, and that the mode of transmission is reflected in the predominance of certain genotypes.

Background: A flavivirus designated hepatitis G virus (HGV) has been isolated from the serum of patients with non-A-E hepatitis. Hitherto, the presence of HGV RNA in serum has been detected with the reverse transcription-polymerase chain reaction (RT-PCR) amplification method. We have now developed an immunoassay for antibodies against an HGV protein., Methods: Recombinant HGV envelope protein E2 was used as antigen in an ELISA. 80 blood donors, 99 intravenous-drug users, and 11 patients with acute post-transfusion hepatitis were tested for antibodies to E2. The HGV-RNA status was assessed by RT-PCR., Findings: Anti-E2 seroprevalence was 9% among the blood donors and 41% among the drug users; HGV-RNA prevalence was 2.5% and 38%, respectively. Whereas anti-E2 prevalence increased with the duration of drug use, HGV-RNA prevalence declined in parallel. In each group, the presence of anti-E2 and HGV RNA was almost mutually exclusive: none of the blood donors and only 4% of the drug users were positive for both markers at the same time. Of the 11 post-transfusion patients--who were all HGV-RNA positive and anti-E2 negative at the onset of disease--four developed antibodies to E2 during the following year, and two of the four subsequently became HGV-RNA negative., Interpretation: We conclude that a humoral immune response to E2 is associated with loss of detectable HGV viraemia. Thus, E2-specific antibodies might serve as a useful marker for diagnosing recovery from HGV infections. The immunoassay we describe should facilitate investigation of suspected infections and may be helpful in the elucidation of the clinical significance of HGV.

The major advance during the past year was the identification of ligands for two of the previously known position-specific integrins in Drosophila. At the same time, two new Drosophila integrin subunits (one alpha and one beta) were discovered, and significant progress was made on developmental genetic analyses of integrin functions, shedding light on the roles of integrins in Drosophila development.