Detection of heterozygous large deletions in the antithrombin gene using multiplex polymerase chain reaction and denatured high performance liquid chromatography

The present study reports a method for the easy, rapid and cost effective detection of heterozygous large deletions. As a model gene all exons of the antithrombin gene were amplified in a one tube multiplex polymerase chain reaction (PCR) and the products separated according to their size by reverse-phase ion-pair high performance liquid chromatography. A significant reduction in the height of a peak in the probandOs sample compared to in the control indicates the presence of a large deletion of the corresponding allele. Using this approach we identified heterozygous deletions in four patients: the deletions affected exons 1 and 2, exon 7 and the whole antithrombin gene.     

Development of the S-303 Pathogen Inactivation Technology for Red Blood Cell Concentrates

Pathogen inactivation systems are in use in many European countries as routine procedures. However, a pathogen inactivation system for erythrocytes is currently not available. Although significant improvements have been made to decrease the incidence of transfusion-transmitted infections, risks remain for infectious disease agents specific to red blood cell concentrates, such as parasitic infections resulting in babesiosis and malaria. The pathogen inactivation system for erythrocytes utilizes S-303 and glutathione for the treatment of red blood cell concentrates. Preclinical studies to assess the pathogen inactivation efficacy and toxicology as well as preliminary clinical studies have been completed. Preclinical studies have shown log reduction for leukocytes, several viruses and bacteria in excess of 4 to 6 logs. Preclinical toxicology studies were conducted to enable the initiation of two phase III clinical studies in the USA for support of acute and chronic anemia. A second-generation system was developed after observation of an unexpected immune response in two chronic anemia patients. Preclinical pathogen inactivation studies, serological evaluations and a clinical study to evaluate survival of S-303-treated erythrocytes have been completed to support advanced development of the S-303 pathogen inactivation system. A functional system for the inactivation of red blood cell concentrates has been completed and is reaching clinical application.     

HLA-DR/DQ interaction in patients with erosive rheumatoid arthritis presenting articular and extraarticular disease manifestations.

In the present study we have analysed the effect of HLA-DRB1 and -DQB1 alleles on disease progression and genetic predisposition among 201 RA patients. We clearly confirm the association of RA with HLA class II alleles sharing the (Q)R/KRAA amino acid (AA) cassette in the third hypervariable region (HVR3) of the DR beta-chain. The HVR3 (Q)R/KRAA motif was significantly overrepresented among RA patients (79% vs. 40%, P < 0.001), with one third of the patients homozygous (28% vs. 6.7%, P < 10(-9)) and the number of rheumatoid factor positive (RF+) patients was significantly increased among HVR3 (Q)R/KRAA homozygous in comparison to HVR3 (Q)R/KRAA negative individuals. Erosive disease defined by the Larsen Score and personal disability determined using the Health Assessment Questionnaire (HAQ) was significantly increased among patients positive for the HVR3 motif with the worst outcome among HVR3 (Q)R/KRAA homozygous patients. In contrast, there was no association of the shared HVR3 AA cassette and disease severity in the majority of patients presenting systemic (extraarticular) disease. Homozygosity for the shared HVR3 motif was only marginally increased among patients presenting 'severe' extraarticular disease in comparison to patients with articular disease (33% vs. 43%, P = ns). Similarly, patients with nodular disease were not more often homozygous for the HVR3 (Q)R/KRAA motif. Furthermore, we observed no HLA-DR independent association of DQB1 alleles among HVR3 (Q)R/KRAA positive patients and controls. Our analysis supports the predominant role of HLA-DR for genetic susceptibility to RA. In the clinical setting, however, HLA-DR typing may be limited to assess the individual risk of patients for disease progression.     

Transplantation of human acute myeloid leukemia (AML) cells in immunodeficient mice reveals altered cell surface phenotypes and expression of human endothelial markers

To better characterize acute myeloid leukemia (AML) development in non-obese diabetic (NOD)/severe combined immunodeficiency (SCID) mice, we transplanted samples from patients with AML or KG-1 and EOL-1 cell lines. We found 9/12 primary AML samples and both cell lines to engraft within 2-8 weeks, with 5-80% human cells in bone marrow. Compared with freshly isolated AML cells, percentages of human CD33+, CD38+, CD31+ CD13+ or CD15+ subpopulations increased after transplantation, whereas percentages of CD34+ cells decreased. Engrafted mice frequently showed expression of human endothelial cell markers. Thus, transplantation of human AML into NOD/SCID mice reveals expression of hematopoietic and endothelial differentiation markers.     

Distance learning on the Internet: web-based archived curriculum

Web-based education through archived educational modules offers a significant opportunity to provide didactic education. By archiving lectures and teaching materials, it reduces the educators' time of preparation, especially when many students will need to take the same curriculum over a long period of time. The site can package educational material in multiple formats including audio, video, and readable text, allowing the student to tailor the educational experience to his/her learning preferences. This can be a stand-alone program, or integrated into a program combining distance and in-person education. Assessment through on-line tests can also be conducted, but these must be considered open-book assessments where collaboration cannot be prevented. As such, this vehicle can be utilized effectively for continuing education programs in health care, where open book is permitted and credits are generally awarded on the honor system. However, tests for certificate courses should only be given with a proctor in attendance. In this instance, on-line tests can be used as pre-tests for the student, while being structured to enhance further learning.     

The antioxidant conundrum in cancer

The health-related effects of interactions between reactive oxygen species (ROS) and dietary antioxidants and the consequences of dietary antioxidant supplementation on human health are by no means clear. Although ROS, normal byproducts of aerobic metabolism, are essential for various defense mechanisms in most cells, they can also cause oxidative damage to DNA, proteins, and lipids, resulting in enhanced disease risk. Dietary antioxidants (e.g., vitamin E, vitamin C, beta-carotene, and selenium), as well as endogenous antioxidant mechanisms, can help maintain an appropriate balance between the desirable and undesirable cellular effects of ROS. However, any health-related effects of interactions between dietary antioxidants and ROS likely depend on the health status of an individual and may also be influenced by genetic susceptibilities. Clinical studies of antioxidant supplementation and changes in either oxidative status, disease risk, or disease outcome have been carried out in healthy individuals, populations at risk for certain diseases, and patients undergoing disease therapy. The use of antioxidants during cancer therapy is currently a topic of heated debate because of an overall lack of clear research findings. Some data suggest antioxidants can ameliorate toxic side effects of therapy without affecting treatment efficacy, whereas other data suggest antioxidants interfere with radiotherapy or chemotherapy. Overall, examination of the evidence related to potential interactions between ROS and dietary antioxidants and effects on human health indicates that consuming dietary antioxidant supplements has pros and cons for any population and raises numerous questions, issues, and challenges that make this topic a fertile field for future research. Overall, current knowledge makes it premature to generalize and make specific recommendations about antioxidant usage for those at high risk for cancer or undergoing treatment.     

Anti-fetal immune response mechanisms may be involved in the pathogenesis of placental abruption

Placental abruption is an unpredictable severe complication in pregnancy. In order to investigate the possibility that the activation of the fetal nonadaptive immune system may be involved in the pathogenesis of this disease, IL-6 release from cord blood monocytes was examined by intracellular cytokine staining and flow cytometric analysis. Our results demonstrate that preterm placental abruption (n = 15) in contrast to uncontrollable preterm labor (n = 33) is associated with significantly (P < 0.001) increased release of IL-6 from the fetal monocytes. The same holds true for rhesus disease (n = 9, P < 0.001) that is characterized by a maternal production of antibodies against the rhesus-D antigen expressed by the fetal erythrocytes. This suggests that during rhesus disease, IL-6 release of monocytes is induced by antibody-mediated cross-linking of these cells to the erythrocytes in the fetal circulation. Hence, this assumption favors the idea that also in case of placental abruption, an increased maternal antibody production against paternal antigens leads to an elevated IL-6 release by the fetal monocytes. To elucidate this potential mechanism, the presence of anti-HLA-antibodies was assessed in the maternal circulation of patients with placental abruption (n = 17) and patients with uncontrollable preterm labor (n = 29). The percentage of women producing anti-paternal HLA-antibodies was significantly (P < 0.01) increased in the group of women with preterm placental abruption (47%) in comparison to women with uncontrollable preterm labor (14%). Therefore, our results suggest that an increased humoral immune response of the mother against the fetus may be decisively involved in the pathogenesis of placental abruption.