[A cryobank as an attribute of omics technologies]. / Kriobank kak atribut omiksnykh tekhnologii.

Biobanks are systematic and annotated collections of biological samples based on the system of standard operating procedures (SOP) and corresponding to the recommendations of the International Society for Biological and Environmental Repositories (ISBER). Standardization of conditions of obtaining, processing, storage of samples and providing to an end user are crucial in the activities of the biobank. The attributes of biobanks include common principles of labeling and annotation of biological samples using specialized software, an automated monitoring system of storage conditions, and registration of biosamples. Cryobanks are the biobanks maintained at the storage conditions from -196°C to -150°C that provide better cell viability and the highest preservation of biological molecules. Cryobanking is the most essential part of the infrastructure of population and personalized medicine, pharmaceuticals and biopharmacology, conservation of rare and endangered species, as well as biotechnology in general. Next Generation Biobanking, a concept especially designed for omics technologies, involves annotating biological samples on many biomarkers based on Next Generation Sequencing techniques, as well as collecting biological material from the same patient at different time points (for example, at different stages of the disease, before and after the operation, at different periods of therapy) with a detailed annotation of physiological, biochemical and clinical data. Epigenetic studies (DNA methylation, microRNA, etc.), as well as bioinformatic data analysis are of great importance in the activity of Next Generation Biobanking. Such biobanks should function based on the new ethical principles of the post-genomic era.

Coronary atherosclerosis in unheralded sudden coronary death under age 50: histo-pathologic comparison with 'healthy' subjects dying out of hospital.

AIM: sudden coronary death (SCD) in older individuals is generally associated with extensive coronary atherosclerosis, although it may be the first manifestation of ischaemic heart disease. In younger age-groups, SCD may occur in the presence of less severe disease. We sought to (1) examine the extent of coronary atherosclerosis in young victims of SCD compared with age- and sex-matched controls, (2) analyse the composition of atherosclerotic plaques in these patients, (3) identify the predominant mechanism of SCD, and (4) evaluate the possibility of detecting this mechanism on the basis of morphologic plaque features, in particular presence and amount of lipid accumulation and calcific deposits. METHODS AND RESULTS: coronary arteries were obtained at autopsy from 28 victims of SCD under age 50 with no prior clinical manifestation of ischaemic heart disease (IHD) and no myocardial scar formation and from 16 age- and sex-matched subjects dying of noncardiac causes out of hospital. Sections of all available major coronary arteries were cut in 5-mm intervals to yield a total of 1357 histologic sections, which were analysed using digitised planimetry. Victims of SCD had significantly more major coronary arteries per subject with luminal area narrowing > or = 75% than controls (on average, 2.1 vs. 0.2). Plaque area per histologic section was 5.1 +/- 2.1 mm(2) in SCD cases and 2.0 +/- 0.9 mm(2) in controls (P < 0.001). The major constituent of all plaques was fibrous tissue. Lipid core area per section was 0.49 +/- 0.59 mm(2) in SCD cases and 0.004 +/- 0.01 mm(2) in controls (P < 0.001), and calcified plaque area was 0.18 +/- 0.19 mm(2) in SCD cases and 0.02 +/- 0.05 mm(2) in controls (P < 0.001), both defining significant differences between SCD cases and controls. Arterial thrombosis, most often with underlying plaque rupture was the mechanism of SCD in > 80% of the cases. Considering histologic sections with > or = 50 and with > or = 75% area stenosis, plaque rupture was independently predicted by lipid core area. Calcific deposits were a frequent feature of plaque rupture but were only associated with it in univariate analysis. CONCLUSIONS: the extent and severity of coronary atherosclerosis in young victims of SCD as the first manifestation of IHD was substantially greater than in age-and sex-matched controls and comparable with that previously reported in SCD cases with a broader age range. Lipid core and calcified plaque areas provided for excellent separation between the two groups, which may have implications for identifying persons at increased risk for SCD by non invasive visualisation and assessment of the coronary arteries.

Time trends in the prevalence of atherosclerosis: a population-based autopsy study.

PURPOSE: Mortality from coronary heart disease is declining but little is known about trends in the prevalence of atherosclerosis. Autopsy rates in Olmsted County, Minnesota, are higher than the national average, offering an opportunity to address this matter. In this study, we determined the prevalence of anatomic coronary disease among autopsied Olmsted County residents and examined the generalizability of these findings. SUBJECTS AND METHODS: Reports of the 2,562 autopsies performed between 1979 and 1994 on Olmsted County residents > or =20 years of age were reviewed for the presence of coronary disease. RESULTS: Among autopsied decedents less than 60 years old at death and among coroner's cases, the prevalence of anatomic coronary disease declined with time (P for trend = 0.05); no trend was detected among older persons or noncoroner's cases. By logistic regression analysis, the crude odds ratio ([OR] per 5 years) for the association between time and anatomic coronary disease was 0.94 (95% confidence interval [CI]: 0.86 to 1.03; P = 0.18]. Age, sex, and antemortem diagnosis of heart disease were also strongly related to the presence of disease. After adjustment for sex and antemortem diagnosis of heart disease, the prevalence of anatomic coronary disease decreased more in younger people than in older people (age 40 years: OR 0.43 [95% CI: 0.24 to 0.80]; age 60 years: OR 0.62 [95% CI: 0.45 to 0.87]; age 80 years: OR 0.89 [95% CI: 0.64 to 1.23]). CONCLUSION: The prevalence of anatomic coronary disease at autopsy decreased between 1979 and 1994, particularly among younger people, supporting the notion that the burden of coronary disease has shifted toward the elderly. These results suggest that the decreased incidence of coronary artery disease has contributed to the recent decrease in coronary mortality, particularly among younger people.

Referral to autopsy: effect of antemortem cardiovascular disease: a population-based study in Olmsted County, Minnesota.

PURPOSE: Autopsy studies can provide insight into disease trends and their determinants, including data on the prevalence of atherosclerosis. However, such studies are subject to autopsy bias, which limits their generalizability to the source population. The impact of this bias on autopsy based estimates of time trends in heart disease prevalence is unknown. To report on the trends over time in autopsy rates in Olmsted County, MN, to examine the association between clinical diagnoses of cardiovascular diseases (CVDs) and referral to autopsy and how this association may have changed over time. METHODS: We examined the trends in autopsy rates between 1979 and 1994 in Olmsted County, and the association between antemortem characteristics including cardiovascular diagnoses and autopsy referral. RESULTS: From 1979 to 1994, a total of 9110 residents died in Olmsted County. The average annual autopsy rate was 30%. Autopsy rates declined from 36% in 1979 to 23% in 1994, corresponding to an average decline of 0.6%/year (p < 0.01). Referral to autopsy was positively associated with younger age, male sex, in-hospital place of death, antemortem diagnoses of myocardial infarction (MI) or peripheral vascular disease (PVD), and earlier calendar period. There was no evidence of an interaction between calendar period and any of these predictor variables. Antemortem diagnosis of heart failure was associated with a decrease in the odds of referral to autopsy over time as compared to persons without such diagnosis. CONCLUSIONS: In Olmsted County, autopsy rates, although declining over time, have remained on average approximately 30%. Antemortem diagnoses of MI or PVD are associated with autopsy referral but this association did not change over time. While the greater decline overtime in the use of autopsy observed among decedents with an antemortem diagnosis of congestive heart failure (CHF) deserves further studies, the present findings reduce the concern for bias of time trends in the prevalence of atherosclerosis by changes in the clinical characteristics of decedents referred to autopsy.

Calciphylaxis associated with cholangiocarcinoma treated with low-molecular-weight heparin and vitamin K.

Calciphylaxis is a rare disorder of small-vessel calcification and cutaneous infarction associated with chronic renal failure. Rare cases of calciphylaxis not associated with chronic renal failure have been reported with breast cancer, hyperparathyroidism, and alcoholic cirrhosis. To our knowledge, we report the first case of calciphylaxis without chronic renal failure associated with cholangiocarcinoma and the first attempt to treat calciphylaxis with vitamin K. A 56-year-old woman presented with necrotic leg ulceration. She was treated initially with low-molecular-weight heparin, with no effect. A coagulation work-up showed vitamin K deficiency. During vitamin K therapy, the patient had fulminant progression of the calciphylaxis. She died, and an autopsy showed metastatic cholangiocarcinoma. Thrombosis and protein C deficiency have been implicated in the pathophysiology of calciphylaxis. Functional protein C deficiency may be one of several factors contributing to the development of calciphylaxis. Vitamin K therapy was ineffective in our patient and may have been detrimental.

Effects of acute and chronic angiotensin receptor blockade on myocardial vascular blood volume and perfusion in a pig model of coronary microembolization.

Based on the reduction of ischemic cardiac events in clinical trials and experimental observations, inhibition of the effects of angiotensin II on coronary microcirculatory function may afford myocardial protection after injury. The immediate effects of intracoronary AT1 receptor blockade with irbesartan were examined in a pig model in the healthy myocardium and in acute ischemia induced by injection of 30-microm microspheres into the left anterior descending coronary artery (LAD). Electron-beam computed tomography was performed for in-vivo quantitative measurements of regional intramyocardial vascular blood volume (V(B)) and perfusion (F(M)), as well as left ventricular ejection fraction (LVEF) and muscle mass. Ratios of V(B) and F(M) in the anterior (LAD-supplied)/ inferior (control) myocardium were generated. At baseline, 0.2 mg/kg irbesartan injected into the LAD increased V(B) and F(M) ratios significantly by 27 +/- 8% and 51 +/- 13%, respectively. After anterior coronary microembolization, V(B) and F(M) ratios were 0.60 +/- 0.05 and 0.51 +/- 0.05, respectively, and were significantly increased by irbesartan (by 24 +/- 10% and by 36 +/- 11%, respectively). After 4 weeks of treatment with oral irbesartan (n = 7) or placebo (n = 7), an improved LVEF (56 +/- 4% v 44 +/- 4%, P = .046) was observed in irbesartan-treated animals, but no difference in LV end-diastolic volumes or muscle mass. Resting V(B) (0.95 +/- 0.06 v 0.76 +/- 0.06; P = .047) and F(M) (0.84 +/- 0.05 v 0.64 +/- 0.04; P = .016) ratios were significantly greater in irbesartan-treated animals. Using adenosine, there was a trend for higher V(B) and F(M) ratios in irbesartan- v placebo-treated animals. Therefore, in a pig model of acute myocardial ischemia, AT1 receptor blockade by irbesartan induced microvascular vasodilation and, ostensibly, conveyed myocardial protection. Long-term treatment with irbesartan resulted in moderate enhancements of resting V(B) and F(M) compared with placebo, suggesting a role for coronary microcirculatory effects of chronic AT1 receptor blockade in preserving LVEF.

Event-related potentials reveal topographical and temporal distinct neuronal activation patterns for spatial and object working memory.

This study examined whether working memory processes for object and spatial information are associated with different patterns of ERP activity. Subjects performed two versions of a delayed match-to-sample task in which either object forms or two-dimensional spatial configurations (S1) had to be encoded and retained in working memory for 6800 ms for comparison with a subsequent stimulus (S2). Event-related potentials (ERPs) were recorded from 29 electrode sites in the S1-S2 interval. Negative slow wave activity in the ERP varied with both, the type of memory task and the amount of materials held in working memory. When spatial information was maintained in working memory negative slow wave activity rapidly rose at recording sites overlying posterior parietal and occipital cortical areas. At these recording sites, slow wave increased in amplitude with increasing spatial memory load. For object information, load-sensitive negative slow wave activity was obtained approximately 2000 ms later than in the spatial task and it was focused to mid-frontal recording sites. Moreover, in the object memory task more pronounced negative slow wave activity was found at right inferior temporal recording sites indicating a larger involvement of the right temporal lobe in the processing of object as compared to spatial information. The results provide evidence for the notion that encoding and retention processes for object information and for spatial information can be functionally dissociated and involve differential patterns of neuronal activation rapidly shifting in time.

Event-related brain potentials during natural speech processing: effects of semantic, morphological and syntactic violations.

The present study investigated different aspects of auditory language comprehension. The sentences which were presented as connected speech were either correct or incorrect including a semantic error (selectional restriction), a morphological error (verb inflection), or a syntactic error (phrase structure). After each sentence, a probe word was presented auditorily, and subjects had to decide whether this word was part of the preceding sentence or not. Event-related brain potentials (ERPs) were recorded from 7 scalp electrodes. The ERPs evoked by incorrect sentences differed significantly from the correct ones as a function of error type. Semantic anomalies evoked a 'classical' N400 pattern. Morphological errors elicited a pronounced negativity between 300 and 600 ms followed by a late positivity. Syntactic errors, in contrast, evoked an early negativity peaking around 180 ms followed by a negativity around 400 ms. The early negativity was only significant over the left anterior electrode. The present data demonstrate that linguistic errors of different categories evoke different ERP patterns. They indicate that with using connected speech as input, different aspects of language comprehension processes cannot only be described with respect to their temporal structure, but eventually also with respect to possible brain systems subserving these processes.

Minimally invasive saphenous vein harvesting: endothelial integrity and early clinical results.

BACKGROUND: Endoscopic harvesting of vein grafts may reduce wound complications but the effect on the venous endothelium is unknown. Endothelium-derived vasoactive substances may be important in vein graft disease prevention. We investigated the impact of endoscopic harvesting on the release of these factors and proceeded to evaluate the clinical applicability. METHODS AND RESULTS: Porcine veins were harvested in either an endoscopic or open fashion. Superfusion bioassay from endoscopic veins had a similar basal secretion as control veins (6.5% +/- 1.5% versus 3.2% +/- 2.2%, respectively; n = 5, p = 0.39). Calcium ionophore A23187 stimulation was similar in both groups (24.6% +/- 5.1% versus 27.3% +/- 9.6%; n = 5, p = 0.68). Light and electron microscopy documented a normal endothelial monolayer in both groups with no endothelial cell or connective tissue loss. Encouraged by these results, 38 patients have subsequently undergone this procedure at our institution. Total operative time for harvesting 35 to 45 cm of saphenous vein was 62.3 +/- 5.3 minutes (range, 35 to 120 minutes). The procurement time in the most recent five patients was 41.6 +/- 3.3 minutes. Patients had little incisional pain, but did have mild ecchymosis. CONCLUSIONS: Endothelial release of vasoactive substances after endoscopic harvesting is similar to that after the traditional, extended incision technique, and microscopy confirmed similar histology. These laboratory findings support the satisfactory early clinical experience with endoscopic harvesting of saphenous veins.

Endothelial regulation of vascular contraction in radial and internal mammary arteries.

BACKGROUND: The extent to which the endothelium regulates radial artery (RA) contractions is unknown. The goals of this study were to characterize endothelium-dependent relaxations in the RA, compare these responses with those in the internal mammary artery (IMA), and, subsequently, manipulate nitric oxide production in the RA with adenovirus-mediated gene transfer. METHODS: Segments of RA and IMA from 43 patients were studied initially in organ chambers. Endothelial function was evaluated and gene transfer, was examined. RESULTS: After precontraction to 80% maximum tension with prostaglandin F2alpha, acetylcholine produced lesser relaxations in the RA (21.5%+/-5.8%) than in the IMA (66.7%+/-10.6%); human thrombin and adenosine 5'-diphosphate yielded similar results. Reduced relaxations in the RA (16.8%+/-4.2%) compared with those in the IMA (71.6%+/-11.9%) were noted with calcium ionophore. Superfusion bioassay demonstrated a similar baseline release in both arteries but a reduced stimulated production of vasoactive substances in the RA, results confirmed by cyclic guanosine monophosphate level determination. The RA produced less 6-keto-prostaglandin F1alpha than the IMA. Light microscopy demonstrated an intact endothelium in both arteries. Adenovirus-mediated gene transfer of nitric oxide synthase augmented relaxations of the RA to acetylcholine. CONCLUSIONS: Reduced production of endothelium-derived relaxing factors suggests diminished endothelial regulation of vascular smooth muscle in the RA compared with the IMA. This finding may explain, in part, the predisposition to vasoconstriction in RA grafts.

Gamma responses and ERPs in a visual classification task.

OBJECTIVE: We examined event-related potentials (ERPs) and gamma range EEG activity in a visual classification task to assess which variables affect these responses. METHODS: Ten subjects silently counted the occurrence of rare Kanizsa squares (targets) among Kanizsa triangles and non-Kanizsa figures (standards). By applying a time-frequency analysis to the data and selectively calculating topographical maps of certain frequencies. RESULTS: We were able to find 3 different types of gamma responses to Kanizsa figures: an early phase-locked gamma response at 40 Hz in the N100 time range, late phase-locked gamma activity (200-300 ms) at 40 Hz and a continuous phase-locked gamma response at 80 Hz due to the monitor refresh frequency. The two 40 Hz responses were significantly higher for Kanizsa figures than for non-Kanizsa figures and within the Kanizsa figures were higher for the target figure than for the non-target. CONCLUSION: The phase-locking of these two responses, previously found also as non-phase-locked activity, could be synchronized due to the monitor flicker frequency. Also, our findings suggest that the gamma responses are not solely associated with the binding of stimulus features, but reflect some processes related to target processing.

An immunological and genetic study of patients with gestational diabetes mellitus.

The aim of the study was to evaluate the frequency of islet cell (ICA) and insulin (IAA) antibodies and of HLA antigen typing in a group of subjects diagnosed with gestational diabetes mellitus (GDM) in a screening-diagnostic program during pregnancy. ICA, complement-fixing (CF) ICA and other autoantibodies, absolute number and percentage of lymphocyte subpopulations, and HLA antigens were evaluated in 68 women with GDM and compared with those of matched controls. ICA were found in 2 (2.9%) and IAA in 1 (1.5%). Both ICA-positive women had CF-ICA; one of them was receiving insulin therapy. while the other was on a special diet. No correlations were found between ICA and IAA, nor between IAA and insulin treatment. As far as lymphocyte subsets were concerned, we found a significant increase in the absolute number of total and activated (CD3+HLA-DR+) T lymphocytes and a significant increase in the absolute number and percentage of suppressor/cytotoxic T lymphocytes (CD8) and NK lymphocytes (CD57) in GDM patients compared with normal pregnant controls. Concerning frequency for HLA A, B, C, DR antigens in the GDM population, only Cw7 was found to be significantly increased and A10 significantly decreased in comparison with controls. Our study suggests that GDM is a heterogeneous disorder in which few patients present with the immunologic and genetic markers of type 1 diabetes.

Comparison of CO2 laser, electrocautery, and scalpel incisions on acute-phase reactants in rat skin.

Lasers and electrocautery devices have been applied as an alternative to the scalpel because of better hemostasis and lymphatic sealing. However, previous studies have demonstrated conflicting data regarding the effects of these modalities on the inflammatory response, the first reaction by tissue during wound healing. The purpose of this study is to quantitate inflammatory responses in rat skin following laser, electrocautery, and scalpel injury by measuring T-kininogen (T-KGN), a major acute-phase protein in the rat and its endogenous substrate, cathepsin B, an important inflammatory mediator. Full-thickness wounds (6 cm) were created on the dorsum of Sprague Dawley rats by using a laser, electrocautery, or scalpel. Tissue samples were harvested at 1 hour to 21 days after injury. T-KGN levels were radioimmunoassayed; cathepsin B activity was assayed by using a synthetic substrate Z-Arg-Arg-MCA. Data were analyzed by analysis of variance. T-KGN levels peaked at 3 days for all modalities, although the laser group was statistically (P < or = 0.01) higher at 1, 3, and 7 days after injury. In contrast, cathepsin B activity was significantly (P < or = 0.01) lower at 3 days in the laser group. CO2 laser ablation incites a greater inflammatory response than electrocautery or scalpel injuries. High levels of T-KGN may provide protection from proteolytic damage associated with cathepsins.

Processing of surgical instruments for minimally invasive surgery.

Processing of surgical instruments for Minimally Invasive Surgery (MIS). General information on the development of Minimally Invasive Surgery and MIS instruments is given. The "MIS" scientific study group was formed to discuss and find solutions for design and hygienic problems in MIS. The article describes various groups of MIS instruments, i.e. disposable instruments, instruments with a cleaning channel, and instruments which can be dismantled. Aspects of how the instruments can be processed effectively will be discussed in detail, including an example of a survey carried out in a German hospital with a special monitoring method. Requirements are described for MIS instrumentation with regard to design, material, and surface conditions. Requirements for hygienic processing methods are listed. The article also describes the necessary control and confirmation of cleanliness, which proves to be a very particular problem. Conclusions on the present status of technology and the possibilities for future development with regard to the processing of MIS-instrumentation are given.

Strategic thinking and stress.

In this study we investigate the behaviour and the performance of experimental subjects, who had to solve a series of tasks of 'strategic thinking' under a stress and a non-stress condition. No significant differences between the conditions could be found with respect to performance and improvement of performance within the series of tasks, but the experimental subjects exhibited under the different conditions different forms of behaviour. Unexpectedly the Ss working under the stress-condition exhibited a more 'analytical' behaviour and increased their workload, instead of decreasing it. Additionally they exhibited a tendency for measures with salient effects and proved to be better in finding the right points of main effort. The results are discussed with respect to a general theory of action regulation.

Metabolic support: how to form a service.

Organizing a metabolic and nutritional support service requires consideration of issues such as departmental affiliation, staffing, and budgeting.

Preoperative renal tumor embolization. A useful procedure?

The medical reports of 113 patients operated upon for renal tumors were retrospectively reviewed to assess the value of preoperative embolization. Coils or ethanol were used preoperatively in 55 patients while 58 patients underwent nephrectomy without embolization. The tumors were staged according to the TNM and the Robson staging classification. T1 and T2 tumors as well as T3 and T4 tumors were grouped together for statistical evaluation. There was no significant difference in intraoperative blood loss, operation time and survival rate between patients undergoing preoperative embolization and those who had only nephrectomy. Duration of hospital stay was always longer in the preoperatively embolized group. Preoperative embolization of renal tumors neither improved the patients' outcome nor reduced the surgical risk. Robson's classification, stage III/IV, did not show any significant differences for embolized patients or for those nephrectomized without embolization.

Principles of the molecular construction of multienzyme templates for peptide biosynthesis in integrated reaction sequences.

The amino acid sequences of the genes coding for four multienzyme peptide synthetases, operating by the thiotemplate mechanism are compared, to show underlying principles in the biosynthetic mechanism. Alignment with other carboxylic acid activating enzymes shows the sequences. LAY(V/I)I(Y/F)TSGT(T/S)GxPKGV and GELx(L/I)GGxG(V/I) to be involved in MgATP2-binding and adenylate formation, and two other sequences, one containing the element FxLGG(H/D)S(I/L) to be involved in covalent binding of the amino acid. As a general rule, 1000 amino acid building blocks are responsible for the incorporation of one amino acid into the nascent peptide.

The adenylation domain of tyrocidine synthetase 1--structural and functional role of the interdomain linker region and the (S/T)GT(T/S)GXPKG core sequence.

Sequence analysis of peptide synthetases revealed extensive structure similarity with firefly luciferase, whose crystal structure has recently become available, providing evidence for the localization of the active site at the interface between two subdomains separated by a distorted linker region [Conti, E., Franks, N. P. & Brick, P. (1996) Structure 4, 287-298]. The functional importance of two flexible loops, corresponding to the linker region of firefly luciferase and the highly conserved (S/T)GT(T/S)GXPKG core sequence, has been studied in view of the proposed conformational changes by the use of mutant analysis, limited proteolysis and chemical modification of tyrocidine synthetase 1. Substitution of the highly conserved Arg416, residing in the loop separating the subdomains of the adenylation domain, resulted in profound loss of activity. Limited proteolysis of the mutant suggested significant structural changes as manifested by lack of protection to degradation in the presence of substrates, revealing a probable disturbance of the induced-fit mechanism regulating the transformation from an open to a closed conformation. Mutants, obtained by replacement of the conserved Lys186 from the (S/T)GT(T/S)GXPKG core sequence, displayed only minor differences in substrate-binding affinity despite significant reduction of catalytic efficiency. Residue Lys186 appears to play an important role in either stabilization of the bound substrate through charge-charge-interactions, and/or fixing of the loop for maintainance of the active-site conformation.