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Next generation sequencing (NGS) technologies have revolutionized cancer research allowing the comprehensive study of cancer using high throughput deep sequencing methodologies. These methods detect genomic alterations, nucleotide substitutions, insertions, deletions and copy number alterations. SOLiD (Sequencing by Oligonucleotide Ligation and Detection, Life Technologies) is a promising technology generating billions of 50 bp sequencing reads. This robust technique, successfully applied in gene identification, might be helpful in detecting novel genes associated with cancer initiation and progression using formalin fixed paraffin embedded (FFPE) tissue. This study's aim was to compare the validity of whole exome sequencing of fresh-frozen vs. FFPE tumor tissue by normalization to normal prostatic FFPE tissue, obtained from the same patient. One primary fresh-frozen sample, corresponding FFPE prostate cancer sample and matched adjacent normal prostatic tissue was subjected to exome sequencing. The sequenced reads were mapped and compared. Our study was the first to show comparable exome sequencing results between FFPE and corresponding fresh-frozen cancer tissues using SOLiD sequencing. A prior study has been conducted comparing the validity of sequencing of FFPE vs. fresh frozen samples using other NGS platforms. Our validation further proves that FFPE material is a reliable source of material for whole exome sequencing.

Background: Biomarkers of the immune system are currently not used as prognostic factors in breast cancer. We analyzed the association of the B cell/plasma cell marker immunoglobulin kappa C (IGKC) and survival of untreated node-negative breast cancer patients., Material and Methods: IGKC expression was evaluated by immunostaining in a cohort of 335 node-negative breast cancer patients with a median follow-up of 152 months. The prognostic significance of IGKC for disease-free survival (DFS) and breast cancer-specific overall survival (OS) was evaluated with Kaplan-Meier survival analysis as well as univariate and multivariate Cox analysis adjusted for age at diagnosis, pT stage, histological grade, estrogen receptor (ER) status, progesterone receptor (PR) status, Ki-67 and human epidermal growth factor receptor 2 (HER-2) status., Results: 160 patients (47.7%) showed strong expression of IGKC. Univariate analysis showed that IGKC was significantly associated with DFS (P = 0.017, hazard ratio [HR] = 0.570, 95% confidence interval [CI] = 0.360-0.903) and OS (P = 0.011, HR = 0.438, 95% CI = 0.233-0.822) in the entire cohort. The significance of IGKC was especially strong in ER negative and in luminal B carcinomas. In multivariate analysis IGKC retained its significance independent of established clinical factors for DFS (P = 0.004, HR = 0.504, 95% CI = 0.315-0.804) as well as for OS (P = 0.002, HR = 0.371, 95% CI = 0.196-0.705)., Conclusion: Expression of IGKC has an independent protective impact on DFS and OS in node-negative breast cancer.

The enantiomers and the cis isomers of two previously studied 4,5-disubstituted oxazolidinones have been synthesized, and their binding to the T-box riboswitch antiterminator model RNA has been investigated in detail. Characterization of ligand affinities and binding site localization indicates that there is little stereospecific discrimination for binding antiterminator RNA alone. This binding similarity between enantiomers is likely due to surface binding, which accommodates ligand conformations that result in comparable ligand-antiterminator contacts. These results have significant implications for T-box antiterminator-targeted drug discovery and, in general, for targeting other medicinally relevant RNA that do not present deep binding pockets.

We retrospectively reviewed clinic charts of 21 children and adolescents with developmental disabilities including autism spectrum disorders (ASD) treated consecutively with aripiprazole (ARI) for irritability and severe challenging behaviors. Data extracted include age, sex, and race; level of intellectual disability (ID); Diagnostic and Statistical Manual-IV diagnoses including comorbidity, ARI dosage, and treatment duration; other psychoactive medications and Clinical Global Impressions-Improvement (CGI-I) at baseline and end point; weight; height; and side effects. Body mass index (BMI) z scores are compared with Centers for Disease Control norms. Eleven boys and 10 girls with ID and/or ASD ages 8 to 18 years (mean age 13.4 years) received ARI; mean dose was 8.4 mg/day (range 2.5 to 15); average duration was 60.6 weeks (7 to 132). Eleven of 21 patients (52%) met CGI-I response of ≤ 2. ARI was well tolerated, including together with stimulants, divalproex, or less commonly other medications. Mean BMI was 23.8 ± 5.9 at baseline and 24.2 ± 5.2 at end. Mean BMI z score increase was 0.06 ± 0.67. Four individuals (19%) manifested early intolerable weight gain. In this long-term clinical sample, ARI was effective in 52% and well tolerated. ARI was mostly weight neutral; early weight gain was intolerable in 19%. Larger long-term outcome studies are warranted in this population.

We previously described the large-scale production of RBCs from hematopoietic stem cells (HSCs) of diverse sources. Our present efforts are focused to produce RBCs thanks to an unlimited source of stem cells. Human embryonic stem (ES) cells or induced pluripotent stem cell (iPS) are the natural candidates. Even if the proof of RBCs production from these sources has been done, their amplification ability is to date not sufficient for a transfusion application. In this work, our protocol of RBC production was applied to HSC isolated from fetal liver (FL) as an intermediate source between embryonic and adult stem cells. We studied the erythroid potential of FL-derived CD34(+) cells. In this in vitro model, maturation that is enucleation reaches a lower level compared to adult sources as observed for embryonic or iP, but, interestingly, they (i) displayed a dramatic in vitro expansion (100-fold more when compared to CB CD34(+)) and (ii) 100% cloning efficiency in hematopoietic progenitor assays after 3 days of erythroid induction, as compared to 10-15% cloning efficiency for adult CD34(+) cells. This work supports the idea that FL remains a model of study and is not a candidate for ex vivo RBCS production for blood transfusion as a direct source of stem cells but could be helpful to understand and enhance proliferation abilities for primitive cells such as ES cells or iPS.

An isolated defect of respiratory chain complex I activity is a frequent biochemical abnormality in mitochondrial disorders. Despite intensive investigation in recent years, in most instances, the molecular basis underpinning complex I defects remains unknown. We report whole-exome sequencing of a single individual with severe, isolated complex I deficiency. This analysis, followed by filtering with a prioritization of mitochondrial proteins, led us to identify compound heterozygous mutations in ACAD9, which encodes a poorly understood member of the mitochondrial acyl-CoA dehydrogenase protein family. We demonstrated the pathogenic role of the ACAD9 variants by the correction of the complex I defect on expression of the wildtype ACAD9 protein in fibroblasts derived from affected individuals. ACAD9 screening of 120 additional complex I-defective index cases led us to identify two additional unrelated cases and a total of five pathogenic ACAD9 alleles.

Epstein Barr virus latent membrane protein 1 (LMP1) induces NF-κB activation through transformation effector sites (TES) 1 and 2, both of which are critical for B-lymphocyte transformation. TES2 principally activates canonical NF-κB, which we confirm is NF-κB essential modifier (NEMO)-dependent and requires an intact ubiquitin binding in A20 binding inhibitor of NF-κB and NEMO (UBAN) domain. LMP1 TES2 activated NF-κB in Jurkat cell lines harboring NEMO truncated at 372 (A45) or NEMO with an in-frame deletion of 133-224 (2C), whereas TNFα, 12-O-Tetradecanoylphorbol-13-acetate, human T-cell leukemia virus 1 Tax, and CD40 did not. In both A45 and 2C Jurkat cell lines, LMP1 TES2-mediated NF-κB activation was blocked by siRNAs to TNFα receptor-associated factor 6 and NEMO, by IκB kinase inhibitors, and by the IκBα superrepressor, indicating that the NEMO mutants function to support canonical NF-κB activation. Expression of A45 or 2C mutants in NEMO-deficient murine embryonic fibroblasts reproduced the Jurkat phenotypes: LMP1 TES2 activated NF-κB in fibroblasts lacking NEMO amino acids 133-224 or 373-419, but TNFα and Tax did not. Further analysis indicated that TES2 did not activate NF-κB in cells expressing the double deletion mutant Δ133-224/Δ372-419. These data provide further evidence of the essential role for NEMO in LMP1 TES2 NF-κB activation and highlight the importance of unique domains within NEMO for sensing distinct NF-κB stimuli.

The cultivation of erythroid cells at large scale would have to be performed in suitable bioreactors which would most likely employ some mode of agitation to ensure optimal mass and gas transfer and prevent culture inhomogeneity. The effect of low agitation at 15-20 rpm on ex vivo erythropoiesis of PB CD34+ derived cultures was investigated and found to have significant impact on erythroid development. Agitated cultures showed a reduced lag phase and increased cell expansion during the early stages of culture. Additionally, agitation accelerated erythroid differentiation as seen by the loss of early development markers, acquisition of late erythroid markers and premature cell cycle arrest, although not yielding higher fractions of terminally differentiated cells in comparison to stationary culture. However, agitation at 20 rpm led to significantly increased loss of cell viability after day 15 in culture, an effect that could be reduced by decreasing the agitation rate to 15 rpm. On the one hand these results imply that agitation may improve cell yields and reduce expensive cytokine-dependent early culture stages but on the other hand it might introduce the risk of increased cell death in large scale culture., (2010. Published by Elsevier B.V.)

The potential of peripheral blood derived CD34+ cells for ex vivo erythropoiesis was investigated in a stroma-free culture system using a novel strategy of daily passaging. By expanding PB-derived CD34+ cells up to 1.5 x 10(6)-fold this method achieved expansion factors previously only reported for CD34+ cells derived from more potent stem cell sources such as cord blood, bone marrow and mobilized peripheral blood. Analysis of cell surface markers showed differentiation of immature CD34+ cells to populations with 80% CD71-/GpA+ cells and up to 45% enucleated cells, indicating a significant amount of terminal maturation. Cell crowdedness was found to have decisive effects on in vitro erythropoiesis. Cell density per surface area rather than cell concentration per media volume determined cell expansion during exponential growth where more crowded cells showed reduced overall expansion. In late stage erythropoiesis, however, when cells no longer proliferating, increased cell density was seen to enhance cell viability. These results indicate that peripheral blood derived haematopoietic stem cells can be an alternative to cells sourced from bone marrow, cord blood or leukapheresis in terms of expansion potential. This provides distinct advantages in terms of availability for studies of conditions for scale-up and maturation, and may have particular clinical applications in the future.

Background: Early detection of breast cancer reduces breast cancer-related mortality. Breast cancer biomarkers offer a promising means of detecting this disease at the earliest and most treatable stages., Patients and Methods: The aim of this study was to generate a protein biomarker profile in tear fluid for breast cancer patients. This established biomarker profile was then used to discriminate between cancer patients and healthy controls. Potential biomarkers were screened in tear fluid from 50 women with breast cancer and 50 healthy women, matched for age. Tear fluid was drawn prior to surgery. Surface-enhanced laser desorption-ionisation time-of-flight mass spectrometry was used for protein profiling with two different active surfaces on the protein chips: a cationic exchanger (CM-10) and a reverse-phase surface (H50). The data were analyzed by multivariate statistical techniques and artificial neural networks., Results: A total of 404 peaks were found with different molecular weights at different laser intensities and a statistically significant (p<0.05) panel with 20 biomarkers was generated. Use of the biomarker panel resulted in 71.19% of the samples being correctly classified as cancer samples (42 out of 59) and 70.69% as control samples (41 out of 58), thus overall 70.94% were correctly classified. The diagnostic pattern was able to differentiate cancer patients from healthy women with a specificity and sensitivity of approximately 70% using tear fluid., Conclusion: In this study a biomarker panel in tear fluid was successfully generated to allow breast cancer patients to be discriminated from healthy women. The study suggests that the proteomic pattern of tear fluid may be useful in the diagnosis of breast cancer and for high-throughput biomarker discovery.

Background: Submicroscopic imbalances in the subtelomeric regions of the chromosomes are considered to play an important role in the aetiology of mental retardation (MR). The aim of the study was to evaluate a quantitative PCR (qPCR) protocol established by Boehm et al. (2004) in the clinical routine of subtelomeric testing., Results: 296 patients with MR and a normal karyotype (500-550 bands) were screened for subtelomeric imbalances by using qPCR combined with SYBR green detection. In total, 17 patients (5.8%) with 20 subtelomeric imbalances were identified. Six of the aberrations (2%) were classified as causative for the symptoms, because they occurred either de novo in the patients (5 cases) or the aberration were be detected in the patient and an equally affected parent (1 case). The extent of the deletions ranged from 1.8 to approximately 10 Mb, duplications were 1.8 to approximately 5 Mb in size. In 6 patients, the copy number variations (CNVs) were rated as benign polymorphisms, and the clinical relevance of these CNVs remains unclear in 5 patients (1.7%). Therefore, the overall frequency of clinically relevant imbalances ranges between 2% and 3.7% in our cohort., Conclusion: This study illustrates that the qPCR/SYBR green technique represents a rapid and versatile method for the detection of subtelomeric imbalances and the option to map the breakpoint. Thus, this technique is highly suitable for genotype/phenotype studies in patients with MR/developmental delay and/or congenital defects.

Background: New biomarkers are needed to improve the early detection of breast cancer. This study describes the use of surface-enhanced laser desorption/ionisation time-of-flight mass spectroscopy (SELDI-TOF-MS) in serum and tear fluid., Materials and Methods: Blood and tear fluid of 10 women with breast cancer and 10 healthy age-matched women were screened for potential biomarkers. Blood samples and tear fluid were drawn prior to surgery. SELDI-TOF-MS were used for protein profiling with three different active surfaces of the protein chips. The data were analyzed by multivariate statistical techniques and artificial neural networks., Results: Complex protein and peptide patterns were found on all three surfaces. We identified the main proteins in tear fluid. Statistically significant differences in the protein pattern (p<0.001) were found between breast cancer patients and healthy controls. The diagnostic pattern differentiated cancer patients from controls with a specificity and sensitivity of approximately 90% in serum and tear fluid., Conclusion: Protein chip technology facilitates the discovery of new and better biomarkers in breast cancer. It is a promising approach to analyse a large number of patients with high sensitivity and specificity. Analysing tear fluid could show some advantages.

Background: Paraoxonase 1 (PON1) is an extracellular enzyme, which in the gastrointestinal tract may act as a local detoxifier, antioxidant, immunomodulator, and/or quorum-quenching factor. There are no data on PON1 activity in Crohn's disease (CD)., Methods: PON1 phenotype and activity were determined spectrophotometrically in 52 subjects with CD, 67 with ulcerative colitis (UC), and 99 healthy individuals, and related to lipid peroxidation and disease phenotype, clinical and biochemical activity, and therapeutic strategy. Diagnostic utility of PON1 was evaluated by ROC analysis and compared with C-reactive protein (CRP)., Results: In comparison with controls (166 U), PON1 was reduced only in active CD (110 U, P < 0.0001) and UC (126 U, P < 0.0001), and correlated with disease activity (r = -0.47, P = 0.001 in CD and r = -0.50, P < 0.001 in UC). PON1 significantly correlated with erythrocyte sedimentation rate (ESR) (r = -0.36), platelets (r = -0.35), interleukin-6 (r = -0.45), hemoglobin (r = 0.29), transferrin (r = 0.46), albumin (r = 0.60) in CD, and CRP (r = -0.29), ESR (r = -0.37), platelets (r = -0.43), leukocytes (r = -0.50), interleukin-6 (r = -0.45), hemoglobin (r = 0.34), transferrin (r = 0.54), and albumin (r = 0.50) in UC. PON1 correlated positively with lipids but not with their peroxidation markers (thiobarbituric acid-reactive substances, lipid hydroperoxides, ox-LDL, and ox-LDL autoantibodies). PON1 phenotype B (protective against IBD) tended to be less frequent in IBD patients than controls, and associated with lower concentration of inflammatory indices. PON1 was a poorer indicator of CD or UC than CRP., Conclusions: PON1 was reduced in IBD, despite treatment with antioxidant 5'-aminosalicylate derivatives. PON1 reflected disease activity, inflammation severity, and anemia but not lipid peroxidation. The diagnostic power of PON1 was insufficient for its clinical application.

Purpose: Epithelial cell adhesion molecule (Ep-CAM) recently received increased attention not only as a prognostic factor in breast cancer but also as a potential target for immunotherapy. We examined Ep-CAM expression in 402 consecutive node-negative breast cancer patients with long-term follow-up not treated in the adjuvant setting., Experimental Design: Ep-CAM expression was evaluated by immunostaining. Its prognostic effect was estimated relative to overexpression/amplification of HER-2, histologic grade, tumor size, age, and hormone receptor expression., Results: Ep-CAM status was positive in 106 (26.4%) patients. In multivariate analysis, Ep-CAM status was associated with disease-free survival independent of age, pT stage, histologic grade, estrogen receptor (ER), progesterone receptor (PR), as well as HER2 status (P = 0.028; hazard ratio, 1.60; 95% confidence interval, 1.05-2.44). Recently, so-called triple-negative (HER-2, ER, and PR) breast cancer has received increased attention. We noticed a similar association of Ep-CAM with disease-free survival in the triple-negative group as for the entire cohort., Conclusion: In this study of untreated breast cancer patients, Ep-CAM overexpression was associated with poor survival in the entire cohort and in the subgroup of triple-negative breast cancer. This suggests that Ep-CAM may be a well-suited target for specific therapies particularly in HER-2-, ER-, and PR-negative tumors.

Objectives: To evaluate the formation of advanced oxidation protein products (AOPPs) in juvenile overweight/obesity and obesity-related disorders and to investigate the effect of weight reduction on AOPPs., Design and Methods: AOPPs were determined in 114 overweight/obese children and adolescents without/with insulin resistance and metabolic syndrome and compared with 53 lean controls. Measurements were repeated following weight reduction program (diet/exercise, bran-enriched diet/exercise, and diet/exercise plus metformin)., Results: Overweight/obese subjects had higher AOPPs than lean controls, more elevated in patients with co-occurring metabolic syndrome. AOPPs positively correlated with central obesity, triglycerides, lipid peroxidation and insulin, and negatively with glucose to insulin ratio. AOPPs decreased following obesity intervention and DeltaAOPPs correlated with DeltaBMI%. AOPPs reduction was more pronounced in subjects on bran-enriched diet. Baseline AOPPs were a better predictor of clinically significant weight reduction than BMI%., Conclusions: Juvenile overweight/obesity was associated with AOPPs accumulation, more pronounced in metabolic syndrome. Body mass reduction decreased oxidative stress, with bran-enriched diet being more effective than diet/exercise alone.

Objective: To evaluate the status and diagnostic utility of antioxidant paraoxonase 1 (PON1) in gastroesophageal cancers., Design and Methods: PON1's arylesterase/paraoxonase activities and phenotype were determined in 82 cancers and 57 controls, and related to clinicopathological features, anemia and cachexia coexistence, cholesterol levels, liver function panel, and inflammatory and angiogenic indices., Results: PON1's activities were decreased in cancer. Arylesterase activity correlated with cancer T and N stages, being an N1-independent predictor. Both activities correlated with transferrin, albumin, CRP and inflammation-based Glasgow Prognostic Score, and arylesterase activity also with interleukin-6 and midkine. Phenotype A (Q192 homozygotes) was associated with lower transferrin and higher TNF-alpha concentrations. PON1's arylesterase activity reflected anemia severity, being correlated with hemoglobin, hematocrit, and iron. PON1 arylesterase activity negatively correlated with alkaline phosphatase and gamma-glutamyl transferase, but not with bilirubin, aminotransferases, HDL or LDL cholesterol. PON1 moderately indicated cancer presence and regional metastasis., Conclusions: PON1 activity decreases in gastroesophageal cancers and corresponds to inflammation severity and cancer-related anemia. PON1 decrease indicates lymph node metastasis, but its moderate predictive power does not recommend PON1 determination alone for clinical application.

Background: Advanced oxidation protein products (AOPPs) are new protein markers of oxidative stress with pro-inflammatory properties, accumulated in many pathological conditions. The issue of their enhanced formation in IBD has not been addressed yet., Methods: The concentration of relative AOPPs (rAOPP; concentration of AOPPs divided by albumin level) were measured in 68 subjects with ulcerative colitis (UC), 50 subjects with Crohn's disease (CD) and 45 healthy volunteers, and related to disease phenotype, clinical and biochemical activity, and therapeutic strategy. Diagnostic utility of rAOPP was evaluated by ROC analysis., Results: In comparison with controls (1.367 micromol/g), rAOPP were increased in inactive (1.778 micromol/g, P = 0.053) and active (1.895 micromol/g, P = 0.013) UC and in active (1.847 micromol/g, P = 0.003) CD. In CD, but not UC, rAOPP correlated with disease activity (r = 0.42, P = 0.013). Significant correlations with the inflammatory/malnutrition indices-erythrocyte sedimentation rate (ESR) (r = 0.53), leukocytes (r = 0.33), platelets (r = 0.38), IL-6 (r = 0.36), and transferrin (r = -0.35) were demonstrated in CD. In UC, rAOPP correlated only with ESR (r = 0.35) and IL-6 (r = 0.30). Instead, associations with antioxidant dismutase (r = 0.29) and catalase (r = 0.22) were observed. The diagnostic power of rAOPP in discriminating diseased from non-diseased subjects was less than that of C-reactive protein (CRP). Simultaneous determination of rAOPP and CRP did not significantly improve the power of single CRP determination., Conclusions: IBD was associated with enhanced formation of AOPP, which differed between C and UC with respect to the relationship between rAOPP and disease activity, inflammatory and antioxidant response. These differences may reflect divergent ways that oxidative stress develops in CD and UC. The diagnostic power of rAOPP was insufficient for its clinical application.

Objectives: To validate the diagnostic utility of oxidative stress markers in the evaluation of young type 1 diabetics, as suggested elsewhere., Design: Advanced oxidation protein products (AOPP), thiobarbituric acid-reactive substances (TBARS) and total antioxidant status (TAS) were measured in sera from diabetics, their siblings and controls, with diagnostic potential evaluated by ROC analysis, and related to diabetes clinical parameters., Results: In diabetics AOPP and TBARS were elevated, TAS decreased. Similar alterations were observed for AOPP and TAS in their siblings. AOPP and TAS were good indicators of diabetes. AOPP and TBARS correlated with HbA1C (independent predictor), but were poor markers of non-adequate glycemic control. The cardiovascular disease risk factors were independent predictors of TBARS concentrations., Conclusions: AOPP accumulation and TAS reduction seem to precede diabetes and might be considered as susceptibility indicators in relatives, but not as diabetes markers in general population (no diabetes specificity has been shown). Application in monitoring of metabolic control is not validated.

Background: Neurofibromatosis type 1 (NF1) is a pheochromocytoma-associated syndrome. Because of the low prevalence of pheochromocytoma in NF1, we ascertained subjects by pheochromocytoma that also had NF1 in the hope of describing the germline NF1 mutational spectra of NF1-related pheochromocytoma., Materials and Methods: An international registry for NF1-pheochromocytomas was established. Mutation scanning was performed using denaturing HPLC for intragenic variation and quantitative PCR for large deletions. Loss-of-heterozygosity analysis using markers in and around NF1 was performed., Results: There were 37 eligible subjects (ages 14-70 yr). Of 21 patients with corresponding tumor available, 67% showed somatic loss of the nonmutated allele at the NF1 locus vs. 0 of 12 sporadic tumors (P = 0.0002). Overall, 86% of the 37 patients had exonic or splice site mutations, 14% large deletions or duplications; 79% of the mutations are novel. The cysteine-serine rich domain (CSR) was affected in 35% but the RAS GTPase activating protein domain (RGD) in only 13%. There did not appear to be an association between any clinical features, particularly pheochromocytoma presentation and severity, and NF1 mutation genotype., Conclusions: The germline NF1 mutational spectra comprise intragenic mutations and deletions in individuals with pheochromocytoma and NF1. NF1 mutations tended to cluster in the CSR over the RAS-GAP domain, suggesting that CSR plays a more prominent role in individuals with NF1-pheochromocytoma than in NF1 individuals without this tumor. Loss-of-heterozygosity of NF1 markers in NF1-related pheochromocytoma was significantly more frequent than in sporadic pheochromocytoma, providing further molecular evidence that pheochromocytoma is a true component of NF1.

Objective: Aggressive chemotherapy and radiotherapy or radical oncological surgery in young women with cancer has greatly enhanced these patients' life expectancy, but these treatments often cause infertility or premature ovarian failure due to a massive destruction of the ovarian reserve. The objective of this review is to discuss the effect of the various cancer treatments on fertility and present the various fertility sparing operations and fertility preservation strategies., Method: An extensive survey of the most up-to-date literature was performed., Results: This review discusses the impact of current cancer treatment on fertility potential and the various surgical and assisted-reproduction innovations available today for the most common cancers in young women. Although the ability to retain reproductive potential is becoming a major quality-of-life factor in an increasing number of young female cancer survivors, they are still being poorly counseled with regard to the negative impact of the treatment on their fertility and on their options for fertility preservation., Conclusion: As the emerging discipline of fertility preservation is steadily attracting increasing interest, developments in the near future promise to be very exciting. However, in everyday routine work, better interdisciplinary cooperation between gynecological and pediatric oncologists, surgeons, immunologists, and endocrinologists is necessary so that individualized options for fertility preservation can be offered in advance of surgical procedures or cancer treatments.

Objective: To describe the prenatal phenotype of the 11q deletion syndrome (Jacobsen syndrome) and present the molecular characterization of the deletion in the case presented., Case: Ultrasound at 18 and 20 weeks of gestation, on a 34-year-old woman who presented for amniocentesis, revealed slow movements, oligohydramnios and dilatation of the cerebral ventricles in the fetus. Maternal and paternal ages were 34 and 38 years, respectively., Results: Prenatal karyotyping of cultured amniotic fluid cells revealed an 11q terminal deletion, 46,XX,del(11)(q23) (Jacobsen syndrome). Real-time quantitative PCR analysis was used to identify and map the breakpoint physically to a 45-kb region located 14.5 Mb from the 11q telomere. Polymorphic DNA marker analysis showed that DNA sequences on the paternally derived chromosome are deleted. At autopsy, facial dysmorphism without major malformations was recorded. Examination of the internal organs disclosed the following abnormalities: a Meckels' diverticulum of 4-mm length, adhesion between the gall bladder and the transverse colon, and bilaterally bilobed lungs without further situs anomalies., Conclusion: Our case demonstrates significant phenotypic variability of Jacobsen syndrome at midtrimester pregnancy; the syndrome may be manifested at this stage only by mild to moderate ventriculomegaly of the brain., (2006 John Wiley & Sons, Ltd.)

Iron balance is regulated by the rate of erythropoiesis and the size of the iron stores. Anemia that accompanies infection, inflammation, and cancer (anemia of chronic disease) features normal or increased iron stores, although patients may have functional iron deficiency, namely, an imbalance between iron requirements of the erythroid marrow and the actual supply. The proportion of hypochromic red cells and the hemoglobin content of reticulocytes are direct indicators of functional iron deficiency. Biochemical markers, especially the soluble transferrin receptor/log ferritin ratio (ferritin index), are useful indicators of the iron supply to erythropoiesis. The relationship between functional iron deficiency (reticulocyte hemoglobin content) and iron supply to erythropoiesis (ferritin index) can be described in a diagnostic plot. In normoproliferative and hypoproliferative erythropoiesis, the plot allows the differentiation of classic iron deficiency from anemia of chronic disease and the combined state of functional iron deficiency with anemia of chronic disease. The therapeutic implications of the plot are to differentiate patients into those who should be administered iron supplements, epoetin, or a combination of epoetin and iron. In patients receiving epoetin therapy, the plot is an important tool for monitoring erythropoietic activity, functional iron deficiency, and adequate iron stores for new red cell production. Enhanced erythropoiesis is reflected quantitatively by the ferritin index vector. A transgression of the 1.5 (3.2) cut-off value for the ferritin index indicates that extra doses of iron need to be administered to increase the body's iron stores. A lack of increase or a reticulocyte hemoglobin content below 28 picograms indicates functional iron deficiency. The diagnostic plot is a model for differentiating iron-deficient states and predicting those patients who will respond to epoetin therapy.

Background: The purpose of this study was to assess whether, even within a normal FSH range (< or =10 mU/ml), age-specific FSH levels are predictive of ovarian reserve., Methods: Between January 1998 and December 2001, 535 women, undergoing controlled ovarian stimulation with 225 IU of recombinant (rec) FSH and 75 IU of recLH, were included in this retrospective cohort study. Criteria for enrolment were: age 25-40 years, basal FSH (b-FSH) < or =10 mU/ml and basal LH < or =12 mU/ml. Patients were assigned to three age groups (group I: 25-29 years; group II: 30-35 years; and group III: 36-40 years). Each age group was divided into quartiles according to b-FSH levels, comparing the lowest and highest b-FSH quartiles for basal hormonal patterns and outcome-related parameters., Results: At ages 25-35 years, women in the lowest FSH quartiles demonstrated significantly increased numbers of oocytes at retrieval (group I: low b-FSH quartile 8.4 +/- 3.7 versus high b-FSH quartile 6.4 +/- 2.7, P < 0.02; group II: 7.5 +/- 4.0 versus 6.3 +/- 3.0, P < 0.047), whereas no difference with regard to oocyte yield was observed in patients above age 35 (group III: low b-FSH quartile 5.5 +/- 3.1 versus high b-FSH quartile 5.6 +/- 3.5). No statistical correlation was found between FSH quartiles and clinical pregnancy rates or miscarriage., Conclusions: In young women, age-specific high b-FSH levels, even within normal ranges, are associated with significantly reduced numbers of oocytes retrieved. B-FSH concentrations should, therefore, be interpreted in an age-specific manner to allow for appropriate patient counselling in IVF.

Purpose: Minimal invasive direct coronary artery bypass grafting (MIDCAB) or off-pump coronary artery bypass grafting (OPCAB) on the beating heart with full or mini-sternotomy are becoming more common in coronary bypass surgery of the left anterior descending (LAD). In the decision, which surgical approach (MIDCAB, OPCAB or conventional surgery with cardiopulmonary bypass) will be best used, knowledge of the anatomical field is of major importance. The aim of the study was to evaluate retrospective ECG-gated 4-row multidetector CT (MDCT) in patients planned for MIDCAB as additional imaging to coronary angiography., Material and Methods: The study included 25 consecutive patients. MSCT was performed as unenhanced (collimation 4 x 2.5 mm) and contrast-enhanced examination (140-170 ml, 300 mg Iodine/ml, collimation 4 x 1 mm). The evaluation included presence of LAD calcifications, distance of LAD and left internal mammarian artery (LIMA), course of LAD and LIMA, the presence or absence of bridging through myocardium or epicardial fat and the presence of pleural fibrosis. The MDCT results were correlated with intra-operative findings., Results: All MDCTs could be assessed with reference to the demands. In 20/25 operations, MDCT had direct influence as to the selection of the surgical approach (11 MIDCAB, 7 OPCAB with mini-sternotomy and 5 with full sternotomy, 2 conventional surgeries). The distance of LAD and LIMA varied from 0.9 to 4.5 cm in MDCT. As to calcifications, 3/25 correlated patients had calcifications and 10 patients had no calcifications in the middle LAD. Seven patients had intraoperative fibrosis of the vessel wall without calcification of the middle LAD, which could not be detected with MDCT. Another 5 patients had single calcified plaques in the middle LAD, 4 of these had a fibrosis of the vessel and 1 had a normal vessel at surgery. In these cases, the anastomosis was done between the calcified plaques. No myocardial bridging was detected by MDCT and at surgery. Bridging of epicardial fat was shown by MDCT and at surgery in 9/25 patients and was excluded in 15 patients. In 1 patient, the LAD seemed to run superficially in MDCT, but was covered with fat as seen during surgery. The course of the LIMA was inconspicuous in all cases, no pleural fibrosis was found., Conclusions: The 4-row MDCT has proven to be adequate in addition to coronary angiography for preoperative evaluation in patients scheduled for MIDCAB and provides the surgeon with relevant information for the selection of the operative approach.

Severe postmenopausal virilization is a rare event in clinical practice. To evaluate ovarian or adrenal hyperandrogenism endocrine tests and imaging are useful diagnostic tools. We report a case of a postmenopausal woman with hirsutism and androgenetic alopecia. A malignant cause for the present disorder could be excluded by imaging. Selective venous sampling was administered with increased testosterone level of the right adrenal vein. Right adrenalectomy and right oophorctomy was recommended.

The aim was to develop a German questionnaire for self-assessment of shoulder function equivalent to the Constant-Score (CS). To evaluate the retest reliability, the CS questionnaire was completed twice within 1 week by 47 patients prior to shoulder surgery. For validation the CS was assessed by the physician after the second self-test. The medium selectivity of the CS questionnaire at hospital admission was 0.47, the medium item difficulty 0.40, the test-retest reliability 0.675 ( p=0.000), the internal consistency of the questionnaire 0.80, and of the physician's CS evaluation 0.85. Construct, content, and discriminative validity of the questionnaire could be demonstrated. A high correlation of the patient-based questionnaire with the physician-assessed CS was found ( p=0.82). The statistical analyses demonstrated that the CS questionnaire is a reliable and valid instrument to evaluate the CS and can therefore be used for follow-up studies.

Telomeric chromosome rearrangements may cause mental retardation, congenital anomalies, miscarriages, and hematological malignancies. Automated detection of subtle deletions and duplications involving telomeres is essential for high-throughput screening procedures, but impractical when conventional cytogenetic methods are used. Novel real-time PCR quantitative genotyping of subtelomeric amplicons using SYBR-green dye allows high-resolution screening of single copy number gains and losses by their relative quantification against a diploid genome. To assess the applicability of the technique in the screening and diagnosis of subtelomeric imbalances, we describe here a blinded study in which DNA from 20 negative controls and 20 patients with known unbalanced cytogenetic abnormalities involving at least one or more telomeres were analyzed using a novel human subtelomere-specific primer set, producing altogether 86 amplicons, in the SYBR-green I-based real-time quantitative PCR screening approach. Screening of the DNA samples from 20 unrelated controls for copy number polymorphism do not detect any polymorphism in the set of amplicons, but single-copy-number gains and losses were accurately detected by quantitative PCR in all patients, except the copy number alterations of the subtelomeric p-arms of the acrocentric chromosomes in two cases. Furthermore, a detailed mapping of the deletion/translocation breakpoint was demonstrated in two cases by novel real-time PCR "primer-jumping." Because of the simplicity and flexibility of the SYBR-green I-based real-time detection, the primer-set can easily be extended, either to perform further detailed molecular characterization of breakpoints or to include amplicons for the detection and/or analysis of syndromes that are associated with genomic copy number alterations, e.g., deletion/duplication-syndromes and malignant cancers., (Copyright 2004 Wiley-Liss, Inc.)

Purpose: To demonstrate a quantitative approach to measuring fluid volumes with standard single shot RARE sequences., Materials and Methods: In phantom experiments, magnetic resonance hydrometry (MRH), in combination with various calibration phantoms (5 mL up to 500 mL) as internal standards, was used to quantify fluid volumes. In total, 16 volume phantoms were investigated with six different calibration phantoms. In addition, noise correction was implemented to correct the quantification results and to avoid the influence of random noise in the image., Results: All MR measurements show significant correlations of up to r = 0.99 (P <.05) with the real applied volume in the investigated phantoms. However, measurements of large volumes were more precise with large calibration phantoms. Noise reduction did not change the correlation between measured and real applied volumes, but did reduce the error of the measured volumes. Calibrated magnetic resonance hydrometry (cMRH) is able to quantify volumes of fluid fast and noninvasively. The volumes of the used calibration phantoms have to be at least in the order of magnitude of the volumes that are to be measured., Conclusion: In vitro, cMRH using a single-shot rapid acquisition with refocused echoes (ssRARE) sequence and calibration phantoms is a fast and accurate method of quantifying steady amounts of fluid., (Copyright 2003 Wiley-Liss, Inc.)

Background: Ventricle-to-coronary artery bypass (VCAB) is an experimental revascularization procedure that provides predominantly systolic instead of diastolic blood flow to a coronary artery., Methods and Results: In a pig model, a stent-based procedure (VSTENT) was developed to create a VCAB. After thoracotomy, a covered VSTENT was implanted between the left ventricle and the left anterior descending coronary artery (LAD). Distal LAD flow, regional myocardial function, and intracoronary pressures were determined at different degrees of LAD stenosis and during complete LAD occlusion. During 3 hours of LAD occlusion, VSTENT preserved net forward flow at 70+/-6% and regional myocardial function at 71+/-8% of baseline. Preservation of net flow was influenced by the positioning of the VSTENT, with higher preservation also under conditions of increased oxygen demand if a "valve-like mechanism" was present during diastole. At a hemodynamically relevant level of LAD stenosis (>70%), systolic inflow was predominant after VSTENT implantation. Changes in mean diastolic intracoronary pressure that resulted from different degrees of LAD stenosis were linearly correlated to net flow after VSTENT implantation (r=0.88; P<0.001)., Conclusions: VSTENT for ventricle-to-coronary artery bypass was feasible and preserved 70+/-6% of baseline flow during complete LAD occlusion. The degree of preservation was dependent on the position of the VSTENT creating a valve-like mechanism during diastole. Residual diastolic blood flow through a high-grade LAD stenosis influenced net flow favorably, because diastolic backflow decreased with increasing mean diastolic intracoronary pressure.

Objectives: To evaluate magnetic resonance hydrometry for the calculation of bladder volume. The reference standard to assess bladder volume is urethral catheterization, which may be linked with the risk of trauma and infection. Hence, ultrasonography is the preferred diagnostic method. However, ultrasonography is investigator dependent and inaccurate in the hands of an inexperienced operator., Methods: Investigations were performed in a 1.0-Tesla clinical scanner with a manufacturer-provided single-shot turbo spin-echo sequence. We examined 30 healthy volunteers (21 males and 9 females) with a mean age of 26.4 years. To quantify the volume of fluid in a magnetic resonance image, a histogram algorithm was used and a calibration phantom applied. Prevoid and postvoid images were acquired. The bladder volume was calculated as the difference between the prevoid and postvoid image fluid volumes. The magnetic resonance-calculated data were compared with the actually voided volumes., Results: The measured voided bladder volume was 400 +/- 33 mL, whereas magnetic resonance hydrometry yielded 390 +/- 31 mL. The difference between both measurements was not statistically significant. The 95% confidence interval for the difference of both measurements ranged from -22.6 to 2.4 mL. The regression had an r2 of 0.97., Conclusions: The feasibility of magnetic resonance hydrometry to quantify the bladder volume noninvasively was demonstrated.

Aims: To show the ability of magnetic resonance hydrometry (MRH) to quantify the pancreatic secretion after secretin stimulation in order to distinguish between physiological excretion and reduced output in chronic pancreatitis., Methods: MRH images were acquired in a 1.0-T-clinical scanner using a body-array coil and a heavily T2-weighted standard single-shot TSE sequence. Thirty-one patients (14 male/17 female) who routinely underwent ERCP for suspected choledocholithiasis (n = 22), recurring abdominal pain (n = 1), icterus (n = 6 and suspected pancreatitis (n = 2) were included. During the investigation 1 CU/kg BW secretin were administered intravenously. Secreted volume of fluid, start of secretion, achievement of a plateau of secretion and a combined score of these parameters (MRH score) were assessed and evaluated. Sensitivity and specificity were calculated for these parameters., Results: 27 patients had no pancreatic pathology, and four suffered from chronic pancreatitis. Patients without pancreatic disorders produced a mean pancreatic fluid volume of 183 plus minus 86 mL, whereas patients with chronic pancreatitis secreted 61 +/- 39 mL. Secretion started after a mean time of 95 +/- 94 seconds (no pancreatic impairment) and 62 +/- 13 seconds (chronic pancreatitis). The MRH score achieved a high accuracy in the detection of chronic pancreatitis., Conclusions: Our study demonstrated the feasibility of measuring pancreatic output by MRH after stimulation with secretin. Moreover, a distinction between normal secretion and patients with chronic pancreatitis is possible.

It was the aim of this study to compare in vitro closure time (PFA-100), reflecting platelet-related primary hemostasis, to more platelet-specific tests like whole blood electrical aggregometry and platelet surface antigen expression in healthy volunteers. In vitro closure time was measured using a PFA-100. Platelet surface antigen expression (CD63, CD62-P, CD42b, CD36, CD31) was determined in accordance with the consensus protocol for flow-cytometric characterisation of platelet function. Platelet aggregometry was performed using a whole blood electrical aggregometer (ADP and arachidonic acid as agonists). Analysis of the obtained data revealed only a few significant correlations between the different platelet function tests used. This finding can be explained by the various aspects of platelet function being focused by these tests in different extents. Whenever platelet function is analysed, the investigator should be aware of the specific and limited evidence of the method used. For screening purposes, it may be useful to introduce a platelet function index, referring to basal platelet activity, platelet adhesion and platelet aggregation at low and high shear stress forces.

This prospective randomized study investigated the influence of normotensive and hypotensive general anesthesia on platelet aggregability, intraoperative blood loss and parameters of plasmatic coagulation during extensive orthognathic surgery. A total of 30 patients were randomly allocated for either normotensive anesthesia maintained by continuous infusion of propofol and remifentanil (NORMO, n=10) or hypotensive anesthesia, whereby hypotension was induced by increasing the infusion rate of remifentanil (HYPO-R, n=10) or by administration of nitroglycerin (HYPO-N, n=10). Whole blood platelet aggregability was significantly reduced during hypotension compared to normotensive anesthesia (P<.01, HYPO-N and HYPO-R vs. NORMO). Mean arterial blood pressure during hypotension correlated well with adenosinediphosphate- (R=.712, P<.001) and collagen-induced platelet aggregability (R=.685, P<.001). Within hypotensive study groups, postoperative fibrinogen levels were significantly different, whereas intraoperative platelet aggregability, postoperative platelet count, prothrombin time, activated partial thromboplastin time and antithrombin levels were not different. Normotensive anesthesia, however, caused significant decreases in platelet count (-29%), prothrombin time (-24%), fibrinogen (-41%) and antithrombin (-28%) and a significant prolongation in activated partial thromboplastin time (+21%) and thrombin time (+18%). There was a trend to reduced intraoperative blood loss in hypotensive study groups; however, differences were not significant. In conclusion, induced hypotension--independent of substances used for induction of hypotension--reduces intraoperative platelet aggregability, subsequently protecting the coagulation system against subclinical consumption coagulopathy. Induced hypotension-caused platelet dysfunction does not lead to an increased intraoperative blood loss, but quite on the contrary shows a trend to reduced intraoperative blood loss, possibly by preventing platelet-induced subclinical consumption coagulopathy.

The numbers of those affected by the HIV virus worldwide are staggering. Emerging research demonstrates a number of lower cost medical approaches that would significantly reduce the rates of transmission of the virus. Nursing plays a critical role in quality-of-life issues at time of diagnosis. Supportive caregiving by compassionate use of vocabulary and knowledgeable resourcing for patients is part of nursing practice in HIV/AIDS care.

Objective: The purpose of the study was to evaluate the best surgical approach in off-pump single vessel revascularization of the left anterior descending coronary artery (LAD)., Methods: In 256 patients a single left internal mammary artery (LIMA) to LAD bypass was performed with beating heart techniques through a left anterior minithoracotomy (minimally invasive direct coronary artery bypass (MIDCAB), n=129) or a full sternotomy (off-pump coronary artery bypass (OPCAB), n=127)., Results: In the OPCAB group, significantly more severe comorbidities (P=0.001) and redo-operations were noted (P<0.001). Conversion to sternotomy or CPB was necessary in five MIDCAB patients and one OPCAB patient. No cerebrovascular accident was seen in both groups. There was no hospital death in MIDCAB- and two deaths in OPCAB procedures (P=ns). There was a significant reduction in time of surgery (P=0.028) and coronary occlusion (P=0.009) in the OPCAB group. No differences in postoperative ventilation time, ICU stay and length of hospital stay were recorded between groups. Wound infections occurred in six MIDCAB patients (4.7%) and one OPCAB patient (0.8%). Early postoperative reoperation due to graft failure was necessary in three patients after MIDCAB and two patients after OPCAB (P=ns). Confirmed by angiography, the early graft patency rate was 96 and 98%, respectively (P=ns)., Conclusions: Both beating heart techniques showed good results with low hospital mortality, low early complications and comparable angiographic results. Nevertheless, MIDCAB is a challenging technique as demonstrated by the longer times of surgery and coronary occlusion with a tendency towards a higher risk of conversion and wound infection. Thus, this technique should only be performed in selected patients with favourable coronary anatomy. Through a sternotomy approach, single vessel revascularization can be performed safely off-pump even in high-risk patients.

Severe combined immunodeficient (SCID) mice injected i.v. with the human T-ALL cell line CCRF CEM (SCID-CEM mice) develop within 50 days life-threatening multi-organ growth of leukaemia cells. The development of leukaemia in SCID-CEM mice treated with three 10 microg i.v. doses of the anti-CD7 immunotoxin (IT) HB2-SAPORIN or the anti-CD38 IT OKT10-SAPORIN was significantly delayed compared with PBS sham-treated animals but 90% of animals treated with either IT eventually developed disseminated leukaemia cell growth. In contrast treatment of SCID-CEM mice with a combination of both ITs led not only to a significantly greater delay in time to leukaemia development but also in the numbers of animals remaining leukaemia free (60%). The native HB2 and OKT10 antibodies (both murine IgG1antibodies) exerted significant, though relatively weak therapeutic effects, probably mediated through an antibody-dependent cellular cytotoxicity (ADCC) mechanism. Moreover, there was no in vivo additivity of therapeutic effect when both antibodies were used in combination. Apparent, however, was that the combination of HB2-SAPORIN IT with OKT10 antibody led to an intermediate therapeutic effect that was significantly greater than that obtained when either was used alone but significantly less than that obtained when the two IT combination was utilized. This was similarly the case for the combination of OKT10-SAPORIN IT with HB2 antibody though the effect was less pronounced in this instance. This result suggests that the therapeutic effect of IT + antibody treatment results from an additivity between antibody-mediated delivery of saporin combined with a SCID mouse NK cell-mediated ADCC attack on the target cell directed through target cell bound antibody Fc engagement with FcgammaRIII on the NK cell surface. The combination of both ITs however gave the best therapeutic outcome in SCID-CEM mice probably as the result of (i) delivery of greater amounts of saporin to target CEM cells positive for both CD7 and CD38, (ii) delivery of an effective dose of saporin to CEM cells downregulated or negative for one of the target antigens and (iii) through ADCC mechanisms that interact additively with IT action. We have previously proposed that combination IT therapy would be one means of overcoming the problem of heterogeneity of antigen expression within a global tumour cell population and these additional findings support this and provide a further strengthening of the rationale for employing cocktails of ITs for the treatment of human malignancies., (Copyright 2001 Cancer Research Campaign.)

Purpose: To evaluate magnetic resonance (MR) hydrometry, a method of quantifying fluid amounts by using MR imaging, for assessing the exocrine function of the pancreas after stimulation with secretin., Materials and Methods: Images were obtained with a single-shot turbo spin-echo sequence by using a 1.0-T magnet with a quadrature body coil. Image postprocessing and evaluation were performed at an external workstation by using a specially designed histogram algorithm that translates the MR signal intensity of duodenal filling into an actual amount of duodenal fluid. This algorithm was tested in vitro and in vivo. Finally, MR hydrometry results in five patients were correlated with those of the secretin-cerulein test., Results: The phantom measurements showed a high correlation (r = 0.99) between the actual amount of fluid in the imaging volume and the calculated results. In vivo, the ability of MR hydrometry to enable exact quantification of fluid amounts was demonstrated. In correlating the signal intensity of duodenal filling with the exact amount of additional fluid in the duodenum in volunteers, a coefficient of 0.043 gray tones per pixel per milliliter was calculated. The correlation (r) between secretin-stimulated duodenal fluid output estimated by using tube aspiration and that estimated by using MR hydrometry was 0.946 (P <.05)., Conclusion: MR hydrometry is a promising noninvasive method of assessing fluid output as a measure of exocrine pancreatic function.

The aim of the study was to perform endoscopic coronary artery bypass grafting on the beating heart using a surgical robotic system. In the study, the surgical system ZEUS was used in combination with 3D visualization for endoscopic coronary artery bypass grafting in 25 patients. In a total of 10 cases, the coronary artery anastomosis was done on the beating heart using endoscopic stabilizers without cardiopulmonary bypass. In all cases, total OR time ranged from 4.0 to 8.0 hours (median 5.5 h); the times for endoscopic coronary artery anastomoses ranged from 14 to 50 minutes (median 25 minutes) with no difference between arrested-heart or beating-heart procedures. All patients had an uneventful angiographic control result. An endoscopic coronary artery anastomosis is possible on the arrested as well as on the beating heart.

One virulence determinant of Staphylococcus aureus is Beta-toxin, a 37 Kd magnesium-dependent sphingomyelinase C. This toxin lyses erythrocytes (RBCs) containing sphingomyelin in the outer lipid layer of their plasma membrane. Although membranes of both human polymorphonuclear leukocytes (PMNs) and lymphocytes (MNLs) contain small amounts of sphingomyelin, the effect of Beta-toxin on these cells remains controversial. The purpose of this study was to investigate the hemolytic activity of this toxin on RBCs of various species and determine the leukotoxic nature on several types of human leukocytes. One nanogram of Beta-toxin lysed 115,000 sheep erythrocytes (sRBCs) and 82,000 human erythrocytes (hRBCs) in a 'hot-cold' assay and caused cytotoxicity to 325 PMNs and MNLs. Both hemolytic and leukotoxic activity were found to be magnesium-dependent. RBC susceptibility to Beta-toxin correlated with the reported sphingomyelin content of each species. Scanning electron microscopy (SEM) demonstrated that 'hot-cold' incubation with Beta-toxin in the presence of magnesium caused significant morphological changes in the surface structure of both RBCs and PMNs. The changes included the formation of pits and membrane invaginations in the RBCs. The PMNs lost their ruffled membrane appearance and showed overall membrane disintegration. This study demonstrated that the viability of sphingomyelin-containing PMNs and MNLs was significantly decreased by the addition of Beta-toxin, indicating that this toxin does, in fact, have a leukotoxic nature. Leukocytes did not have significant membrane invaginations unlike toxin-treated RBCs; therefore, it is possible that leukotoxicity does not result from membrane lysis.

Preeclampsia is still a leading cause of maternal and fetal morbidity and mortality. There is evidence for the involvement of platelets. Therefore, we investigated the suitability of corrected whole blood impedance aggregometry as an early predictor of preeclampsia in 71 consecutive, high-risk pregnancies. According to the occurrence of preeclampsia, defined postpartum by an independent investigator, and the stage of pregnancy (early and late, cutoff: 25 weeks of gestation), four study groups were defined. Platelet aggregation data were corrected for the influence of hematocrit and platelet count by a special purpose software package. Women developing preeclampsia showed significantly higher platelet aggregation response compared to controls in early and late pregnancy. In early pregnancy, all women developing preeclampsia had aggregation responses to collagen higher than the highest responses among the controls. Hence, this test had a 100% positive predictive value of subsequent preeclampsia. Despite being significantly increased, platelet aggregability was of minor predictive value in late pregnancy. We conclude that preeclampsia is accompanied by exaggerated platelet aggregability, particularly perceptible early in the course of pregnancy. We propose collagen-induced whole blood platelet aggregation with correction for the influence of hematocrit and platelet count for early detection of preeclampsia.

Background: In order to minimize surgical trauma, video-assisted mitral valve operation has been started using the Port-Access technique with the addition of a three-dimensional visualization system (Vista Cardiothoracic Systems Inc, Westborough, MA) and a voice-controlled camera-holding robotic arm (Aesop; Computer Motion Inc, Goleta, CA)., Methods: Port-Access mitral valve replacement or repair (PAMVR) was undertaken using an endovascular cardiopulmonary bypass (CPB) system. Fifty patients underwent Port-Access mitral valve replacement or repair. A three-dimensional thoracoscope was inserted allowing complete three-dimensional projection of the mitral valve (Vista). In the last 20 patients, the camera was attached to a robotic arm (Aesop), which allowed stabilization and voice-activated movement of the camera. Mitral valve repair was performed in 26 patients, and the valve was replaced in 24 patients with a mechanical valve prosthesis., Results: Median time of operation was 4.2 hours, aortic cross-clamp time 83 minutes, CPB time 125 minutes, intensive care unit stay 1.5 days and hospitalization 9.0 days. Three months follow-up was complete in 40 patients, with 34 patients (85%) in New York Heart Association class I and 6 patients in class II. Mortality was 0% and rate of reoperation was 2%, with a follow-up time up to 1.5 years postoperatively., Conclusions: Using three-dimensional video and robotic assistance, it was possible to minimize the length of skin incision, but at the same time to optimally visualize the whole mitral valve apparatus in order to perform true Port-Access mitral valve operation, including various repair techniques.

A histoanatomic study of the rotator cuff interval was done in 13 cadaveric specimens to investigate the relation of its ligamentous structures to the long head of the biceps tendon, with a special focus on revealing a stabilizing function. After macroscopic evaluation, the lateral half of the rotator cuff interval capsule was cut into three sections: medial, middle, and lateral. These sections were embedded in methacrylate, and then serial sections were made and stained for polarized light microscopy. The superior glenohumeral ligament was seen to form a fold having the macroscopic appearance of a U-shaped anterior suspension sling for the long head of the biceps tendon. Microscopic evaluation revealed an important role of the fasciculus obliquus in the roof of this sling. Fibers of the supraspinatus tendon join the posterosuperior part of the sling. The subscapularis tendon is not involved in this suspensory mechanism. As a result of these observations, we determined that the superior glenohumeral ligament and the fasciculus obliquus are the most important ligamentous reinforcements of a stabilizing sling for the long head of the biceps tendon in the rotator cuff interval. Their histologic appearance indicates they function to protect the long head of the biceps against anterior shearing stress. A lesion of this sling might lead to anterior instability of the biceps tendon.

Objective: Several different techniques exist within the field of minimally invasive coronary artery surgery. In this study the impact of all these techniques on the total costs and economics has been evaluated., Methods: Since May 1997, 121 minimally invasive direct coronary artery bypass (MIDCAB) procedures, 125 off-pump coronary artery bypass (OPCAB), ten Port-Access coronary artery bypass (PA-CABG) and 10 endoscopic coronary artery bypass grafting (ENDO-CABG) procedures were performed at our institution. A relative cost analysis of the different procedures was carried out in addition to a thorough evaluation done in five patients of each group dividing the costs into staff-related costs, material-related costs and general hospital costs. The costs were set in relation to regular CABG procedures., Results: Specific less invasive coronary artery surgical techniques, such as the MIDCAB or OPCAB technique already are able to reduce the total costs when compared to regular CABG procedures. Within the Port-Access group as well as the ENDO-CABG group, increased material- and general costs are present when compared to regular CABG leading to increased total hospital costs for PA-CABG and Endo-CABG in Germany., Conclusion: At present, MIDCAB and OPCAB procedures are able to reduce total hospital charges, when compared to regular CABG procedures. Increased costs for Port-Access, as well as Endo-CABG surgeries may be compensated in the future by decreased costs due to a shorter phase of rehabilitation and faster return to regular professional activities.

Background: To achieve an endoscopic coronary bypass anastomoses we performed a study with endoscopic robotic instrumentation and camera guidance using three-dimensional (3-D) visualization., Methods: The surgical robotic system ZEUS (Computer Motion Inc, Goleta, CA) consists of three interactive robotic arms and a control unit allowing the surgeon to move the instrument arms in a scaled down mode. The third arm (AESOP, Computer Motion Inc, Goleta, CA) positions the endoscope via voice control. The study had three phases. Phase I: In a phantom model, end-to-side anastomoses between vein grafts and the left anterior descending coronary artery (LAD) of 109 pig hearts were performed. Phase II: In 6 dogs (FBI, 20-25 kg) the left internal mammary artery (LIMA) was harvested endoscopically. During Port-Access (Heartport Inc, Redwood City, CA) cardiopulmonary bypass (CPB), LIMA and LAD were then anastomosed endoscopically with the help of telemetric ZEUS instruments (Computer Motion Inc). Phase III: A total of seven patients were operated on with help of the ZEUS system (Computer Motion Inc). After endoscopic LIMA harvesting and CPB using the Port-Access (Heartport Inc) system, the bypass graft (LIMA to LAD) was anastomosed endoscopically through three thoracic ports in 2 patients. Another 3 patients were operated on off-pump with regional stabilization and 2 patients with sternotomy and routine CPB., Results: The practice with the phantom model and the subsequent animal experiments allowed the surgeons to gain sufficient experience for the clinical setting. In the clinical cases, times for anastomoses ranged from 20 to 42 minutes. Median internal mammary artery flow rate was 74 mL per minute (range 36-110 mL per minute). One patient in the off-pump group was converted to CPB and routine anastomosis. All patients had an uneventful angiographic control and postoperative course., Conclusions: Using telemetic technology, a completely endoscopic anastomosis of LIMA to LAD is possible on the arrested heart, as well as on the beating heart.

Sepsis and related syndromes account for a high morbidity and mortality caused by the development of multiorgan failure. Pathogenesis of sepsis is complex, involving humoral as well as cellular factors. Since the role of platelets is still undefined in this concern, we investigated CD63, CD62P, CD36, and CD31 expression on platelets of patients in septic shock (n = 18) using a flow cytometric assay in whole blood. Samples were drawn within 24 hours of onset. We found thrombocytopenia accompanied by a significantly higher expression of CD63, CD62P, and CD31 and a significant downregulation of CD36 in comparison to healthy volunteers (n = 18). Changes in CD63 and CD62P expression indicates platelet activation. Because CD62P, CD36, and CD31 mediate interaction of platelets with leukocytes, subendothelial matrix and probably endothelial cells as well as platelet adhesion/aggregation, our findings suggest an involvement of platelets in leukocyte/endothelial cell interaction in septic shock. We suspect that thrombocytopenia is not due to bone marrow depression, but rather is due to consumption of highly activated platelets in the microcirculation. We feel that our observations may offer a rationale for potentially beneficial effects of antiplatelet therapy in sepsis; however, further studies have to evaluate its beneficial impact as well as its potential risk for bleeding complications.

Objective: With the aim of performing a completely endoscopic coronary bypass anastomosis, we have undertaken an experimental and clinical study using robotic instrumentation and voice-controlled camera guidance., Methods: The ZEUS Robotic Surgical System (Computer Motion Inc, Goleta, Calif) consists of three interactive robotic arms and a control unit, allowing the surgeon to move the instrument arms in a scaled down mode. The third arm (AESOP, Computer Motion) positions the endoscope via voice control., Phase I: In a phantom model, vascular grafts were anastomosed to the left anterior descending coronary artery (LAD) of 50 pig hearts with either 2- or 3-dimensional visualization., Phase Ii: In 6 dogs (FBI 20-25 kg) the left internal thoracic artery (LITA) was harvested endoscopically. Then the animals were placed on an endovascular cardiopulmonary bypass system (Port-Access, Heartport, Inc, Redwood City, Calif). Anastomosis of the LITA to the LAD was performed endoscopically with the telemetric ZEUS instruments. Flow rates through the LITA were measured by Doppler analysis., Phase Iii: Two patients were operated on with the ZEUS system. After endoscopic harvesting of the LITA and cardiopulmonary bypass with the Port-Access system, the bypass graft (LITA-LAD) was anastomosed endoscopically with the ZEUS system through three thoracic ports., Results: In the dry laboratory, the time range required for the robotically assisted coronary anastomosis was 35 to 60 minutes with 2-dimensional visualization and 16 to 32 minutes with 3-dimensional visualization. In the animal experiments, the median time for endoscopic harvesting of the LITA was 86 minutes (range 56-120 minutes) and for the anastomosis, 42 minutes (range 35-105 minutes); flow rates through the LITA ranged between 22 and 45 mL/min. In the clinical cases, preparation times for the LITA were 83 and 110 minutes, respectively, and anastomosis times, 42 and 40 minutes, respectively. Doppler flow rates measured 125 and 85 mL/min, respectively. Both patients had an uneventful follow-up angiogram and postoperative course., Conclusions: With sophisticated robotic technology, a completely endoscopic anastomosis of the LITA to the LAD is possible, allowing technically precise operations within acceptable time limits.