Muscle secreted factors enhance activation of the PI3K/Akt and ß-catenin pathways in murine osteocytes.

Skeletal muscle and bone interact at the level of mechanical loading through the application of force by muscles to the skeleton and more recently focus has been placed on molecular/biochemical coupling of these two tissues. We sought to determine if muscle and muscle-derived factors were essential to the osteocyte response to loading. Botox® induced muscle paralysis was used to investigate the role of muscle contraction during in vivo tibia compression loading. 5-6 month-old female TOPGAL mice had their right hindlimb muscles surrounding the tibia injected with either BOTOX® or saline. At four days post injections when muscle paralysis peaked, the right tibia was subjected to a single session of in vivo compression loading at ∼2600 µÎµ. At 24 h post-load we observed a 2.5-fold increase in ß-catenin signaling in osteocytes in the tibias of the saline injected mice, whereas loading of tibias from Botox® injected mice failed to active ß-catenin signaling in osteocytes. This suggests that active muscle contraction produces a factor(s) that is necessary for or conditions the osteocyte's ability to respond to load. To further investigate the role of muscle derived factors, MLO-Y4 osteocyte-like cells and a luciferase based ß-catenin reporter (TOPflash-MLO-Y4) cell line we developed were treated with conditioned media (CM) from C2C12 myoblasts (MB) and myotubes (MT) and ex vivo contracted Extensor Digitorum Longus (EDL) and Soleus (Sol) muscles under static or loading conditions using fluid flow shear stress (FFSS). 10 % C2C12 myotube CM, but not myoblast or NIH3T3 fibroblast cells CM, induced a rapid activation of the Akt signaling pathway, peaking at 15 min and returning to baseline by 1-2 h under static conditions. FFSS applied to MLO-Y4 cells for 2 h in the presence of 10 % MT-CM resulted in a 6-8 fold increase in pAkt compared to a 3-4 fold increase under control or when exposed to 10 % MB-CM. A similar response was observed in the presence of 10 % EDL-CM, but not in the presence of 10 % Sol-CM. TOPflash-MLO-Y4 cells were treated with 10 ng/ml Wnt3a in the presence or absence of MT-CM. While MT-CM resulted in a 2-fold activation and Wnt3a produced a 10-fold activation, the combination of MT-CM + Wnt3a resulted in a 25-fold activation of ß-catenin signaling, implying a synergistic effect of factors in MT-CM with Wnt3a. These data provide clear evidence that specific muscles and myotubes produce factors that alter important signaling pathways involved in the response of osteocytes to mechanical load. These data strongly suggest that beyond mechanical loading there is a molecular coupling of muscle and bone.

[Acetabular revision arthroplasty with a novel cementless augment-and-modular-cage system (MRS-C) : Surgical technique and preliminary clinical results]. / Hüftpfannenwechsel mit einem neuartigen zementfreien "Augment-and-modular-cage"-Revisionssystem (MRS-C) : Chirurgische Technik und erste klinische Ergebnisse.

OBJECTIVE: Optimal reconstruction of acetabular anatomy and primarily stable bridging even of large bone defects in hip revision surgery by intraoperative cementless application of highly modular components of the MRS-TITAN Comfort (MRS-C) system according to the augment-and-modular-cage principle. INDICATIONS: Acetabular defect situations in hip revision surgery, hereditary/acquired acetabular deformities, severe secondary hip osteoarthritis. CONTRAINDICATIONS: Extended periacetabular defects of the dorsal pillar, pelvic discontinuities, persisting posttraumatic or periprosthetic infections, lack of intraoperative bony anchorage options. SURGICAL TECHNIQUE: Conventional positioning. Flexible selection of approach (posterolateral, anterolateral or transgluteal) to the hip joint, if possible using a previous approach. Implant removal, preparation and intraoperative acetabular defect evaluation. Optional impaction bone grafting for biological downsizing. Selection of modular components and augments depending on the defect. Preparation of the obturator foramen for anchorage of the caudal flange. Optional preparation of the os ilium for additional fixation of the cage with flanges. Fixation with polyaxial screws and selection of version/inclination of the cup component. Selection of the inlay (polyethylene/ceramic). POSTOPERATIVE MANAGEMENT: Partial weight-bearing with 20 kg body-weight for 6 weeks with crutches. Prophylaxis for deep vein thrombosis and heterotopic ossification. Additional adapted antibiotic therapy in cases of infection. RESULTS: A minimum 1­year follow-up of 42 patients showed significant improvements of Harris Hip Scores (p < 0.0001), adequate reconstruction of the physiologic center of rotation (x-ray controlled in the vertical plane, referenced to the contralateral side), and a clear reduction of preoperative leg length discrepancies. No significant changes in range of motion. No implant-associated complications noted. Based on the short follow-up results, the implant proves as suitable for reconstruction of even large acetabular bone defects.

Direct and indirect costs of COPD and its comorbidities: Results from the German COSYCONET study.

BACKGROUND: Reliable up-to-date estimates regarding the economic impact of chronic obstructive pulmonary disease (COPD) are lacking. This study investigates COPD excess healthcare utilization, work absenteeism, and resulting costs within the German COPD cohort COSYCONET. METHODS: Data from 2139 COPD patients in GOLD grade 1-4 from COSYCONET were compared with 1537 lung-healthy control subjects from the population-based KORA platform. Multiple generalized linear models analyzed the association of COPD grades with healthcare utilization, work absence, and costs from a societal perspective while adjusting for sex, age, education, smoking status, body mass index (BMI), and several comorbidities. RESULTS: COPD was significantly associated with excess healthcare utilization, work absence, and premature retirement. Adjusted annual excess cost of COPD in 2012 for GOLD grade 1-4 amounted to €2595 [1770-3678], €3475 [2966-4102], €5955 [5191-6843], and €8924 [7190-10,853] for direct costs, and €8621 [4104-13,857], €9871 [7692-12,777], €16,550 [13,743-20,457], and €27,658 [22,275-35,777] for indirect costs respectively. Comorbidities contributed to the primary effect of COPD on direct costs only. An additional history of cancer or stroke had the largest effect on direct costs, but the effects were smaller than those of COPD grade 3/4. CONCLUSIONS: COPD is associated with substantially higher costs than previously reported.

Occlusive gloves and skin conditions: is there a problem? Results of a cross-sectional study in a semiconductor company.

BACKGROUND: Although there is poor scientific evidence that working with occlusive gloves is as damaging as wet work, prolonged glove occlusion is considered to be a risk factor for developing hand eczema similar to wet work. OBJECTIVES: To assess the effects of wearing occlusive gloves during the whole working day, without exposure to any additional hazardous substances, on skin condition and skin barrier function. METHODS: We investigated 323 employees of a semiconductor production company in Germany: 177 clean-room workers wearing occlusive gloves during the whole shift (exposed group) and 146 employees working in administration (control group). A standardized interview was performed, the skin condition of both hands was studied using the quantitative skin score HEROS, and transepidermal water loss (TEWL) and stratum corneum hydration were measured. RESULTS: There was no significant difference in skin condition between the two subgroups. Values for TEWL and corneometry were significantly higher in exposed participants (P < 0·05). However, the TEWL values were similar to control values if participants took off the occlusive gloves at least 30 min before the measurement. Hence, the effect of occlusion on skin barrier function seems to be transient. CONCLUSIONS: Prolonged wearing of occlusive gloves with clean hands and without exposure to additional hazardous substances does not seem to affect the skin negatively.

Time-frequency techniques in biomedical signal analysis. a tutorial review of similarities and differences.

OBJECTIVES: This review outlines the methodological fundamentals of the most frequently used non-parametric time-frequency analysis techniques in biomedicine and their main properties, as well as providing decision aids concerning their applications. METHODS: The short-term Fourier transform (STFT), the Gabor transform (GT), the S-transform (ST), the continuous Morlet wavelet transform (CMWT), and the Hilbert transform (HT) are introduced as linear transforms by using a unified concept of the time-frequency representation which is based on a standardized analytic signal. The Wigner-Ville distribution (WVD) serves as an example of the 'quadratic transforms' class. The combination of WVD and GT with the matching pursuit (MP) decomposition and that of the HT with the empirical mode decomposition (EMD) are explained; these belong to the class of signal-adaptive approaches. RESULTS: Similarities between linear transforms are demonstrated and differences with regard to the time-frequency resolution and interference (cross) terms are presented in detail. By means of simulated signals the effects of different time-frequency resolutions of the GT, CMWT, and WVD as well as the resolution-related properties of the interference (cross) terms are shown. The method-inherent drawbacks and their consequences for the application of the time-frequency techniques are demonstrated by instantaneous amplitude, frequency and phase measures and related time-frequency representations (spectrogram, scalogram, time-frequency distribution, phase-locking maps) of measured magnetoencephalographic (MEG) signals. CONCLUSIONS: The appropriate selection of a method and its parameter settings will ensure readability of the time-frequency representations and reliability of results. When the time-frequency characteristics of a signal strongly correspond with the time-frequency resolution of the analysis then a method may be considered 'optimal'. The MP-based signal-adaptive approaches are preferred as these provide an appropriate time-frequency resolution for all frequencies while simultaneously reducing interference (cross) terms.

Molecular pharmacological profile of a novel thiazolinone-based direct and selective 5-lipoxygenase inhibitor.

BACKGROUND AND PURPOSE: The potency of many 5-lipoxygenase (5-LOX) inhibitors depends on the cellular peroxide tone and the mechanism of 5-LOX enzyme activation. Therefore, new inhibitors that act regardless of the mode of enzyme activation need to be developed. Recently, we identified a novel class of thiazolinone-based compounds as potent 5-LOX inhibitors. Here, we present the molecular pharmacological profile of (Z)-5-(4-methoxybenzylidene)-2-(p-tolyl)-5H-thiazol-4-one, compound C06. EXPERIMENTAL APPROACH: Inhibition of 5-LOX product formation was determined in intact cells [polymorphonuclear leukocytes (PMNL), rat basophilic leukaemia-1, RAW264.7] and in cell-free assays [homogenates, 100, 000×g supernatant (S100), partially purified 5-LOX] applying different stimuli for 5-LOX activation. Inhibition of peroxisome proliferator-activated receptor (PPAR), cytosolic phospholipase A(2) (cPLA(2) ), 12-LOX, 15-LOX-1 and 15-LOX-2 as well as cyclooxygenase-2 (COX-2) were measured in vitro. KEY RESULTS: C06 induced non-cytotoxic, direct 5-LOX inhibition with IC(50) values about 0.66 µM (intact PMNL, PMNL homogenates) and approximately 0.3 µM (cell-free PMNL S100, partially purified 5-LOX). Action of C06 was independent of the stimulus used for 5-LOX activation and cellular redox tone and was selective for 5-LOX compared with other arachidonic acid binding proteins (PPAR, cPLA(2) , 12-LOX, 15-LOX-1, 15-LOX-2, COX-2). Experimental results suggest an allosteric binding distinct from the active site and the C2-like domain of 5-LOX. CONCLUSIONS AND IMPLICATIONS: C06 was identified as a potent selective direct 5-LOX inhibitor exhibiting a novel and unique mode of action, different from other established 5-LOX inhibitors. This thiazolinone may possess potential for intervention with inflammatory and allergic diseases and certain types of cancer.

Time-variant analysis of phase couplings and amplitude-frequency dependencies of and between frequency components of EEG burst patterns in full-term newborns.

OBJECTIVE: Burst activity of the 'trace alternant' (TA) EEG pattern in the quiet sleep of full-term newborns is investigated to explore the timing and the time-variant coupling characteristics of and between a burst's oscillatory components. The working hypothesis is that signal properties provide information about the neuronal initiation processes of the burst, and about the coupling and interrelation dynamics between cortical low-frequency oscillations and high-frequency spindles in thalamic structures which substantially contribute to the burst pattern. METHODS: For time-variant phase-locking index (PLI), phase-synchronization index (PSI), quadratic phase coupling (QPC) measures, and amplitude-frequency dependency analyses the Gabor and the Hilbert transformation, both implemented as fast Fourier transformation-based approaches, were used. Additionally, models of mutually coupled Duffing oscillators were adapted to the burst data derived from the neonates ('measured bursts'), and the corresponding 'modeled burst' simulations were analyzed in comparison to the measured bursts. RESULTS: A strong phase-locking of the high-frequency oscillations and synchronization between low- and high-frequency oscillatory activity at burst onset can be observed. The QPC courses and the amplitude of all oscillations rise slightly before or at the burst onset and reach their maximum within the following 1-3 s after onset. Additionally, correlative envelope-envelope and envelope-frequency couplings within and between the burst oscillations can be demonstrated. All theses time-variant signal properties can be simulated by the model. CONCLUSIONS: The amplitude-independent phase measures point to a phase stabilization of high-frequency oscillatory activity which occurs before the initiation of the low-frequency oscillation. This finding points to a trigger process in which the thalamus is initially involved. After burst onset the cortical low-frequency oscillation modulates the high-frequency oscillatory activities, where modulation and additional coupling effects can be explained by three mutually coupled oscillators. SIGNIFICANCE: The model-based analysis strategy offers an up-to-date methodological guideline and sets a new standard of analysis for the investigation of EEG patterns and event-related potentials.

Dynamics of directed interactions between brain regions during interburst-burst EEG patterns in quiet sleep of full-term neonates.

The study investigates time-variant directed interactions between brain regions during the interburst-burst EEG pattern (tracé alternant) characteristic of quiet sleep in healthy neonates. The transition from interburst to burst is of particular interest as the generation of the EEG characteristics at burst onset reflects timing and time-variant interplay between the cortical and the thalamo-cortical brain structures. To study the dynamics of the interactions, time-variant partial directed coherence (PDC), a measure of effective connectivity, was used which allows analysis in the time-frequency range. The main results of the grand mean PDC analysis are: (1) PDC time-frequency patterns are frequently associated with phase-locked oscillations. (2) Interhemispheric interactions are dominant between frontal, central and occipital electrodes and intrahemispheric interactions are much less substantial. (3) An interaction breakdown for the frequency ranges 1-4 Hz (Fp(1) ⇒ Fp(2)) and 0.5-3 Hz (Fp(2) ⇒ Fp(1)) exists which lasts about 2.5s and which is located at about burst onset. (4) Strong interactions in the high-frequency range 3.5-4.5 Hz between the frontal electrodes can be observed for both directions at the burst onset. It can be concluded that the evolution of strong interactions in the high-frequency range, which starts shortly before or at the burst onset from frontal regions to anteroposterior directions as well as the frontal interhemispheric interactions, are associated with the burst onset generation. Additionally, the collapsing of the interactions before burst onset and after the burst are indicative of neuronal reorganisation processes.

Inhibition of thromboxane A2-induced arrhythmias and intracellular calcium changes in cardiac myocytes by blockade of the inositol trisphosphate pathway.

We have recently reported that left atrial injections of the thromboxane A(2) (TXA(2)) mimetic, (5Z)-7-[(1R,4S,5S,6R)-6-[(1E,3S)-3-hydroxy-1-octenyl]-2 -oxabicyclo[2.2.1]hept-5-yl]-5-heptenoic acid (U46619), induced ventricular arrhythmias in the anesthetized rabbit. Data from this study led us to hypothesize that TXA(2) may be inducing direct actions on the myocardium to induce these arrhythmias. The aim of this study was to further elucidate the mechanism responsible for these arrhythmias. We report that TXA(2)R is expressed at both the gene and protein levels in atrial and ventricular samples of adult rabbits. In addition, TXA(2)R mRNA was identified in single, isolated ventricular cardiac myocytes. Furthermore, treatment of isolated cardiac myocytes with U46619 increased intracellular calcium in a dose-dependent manner and these increases were blocked by the specific TXA(2)R antagonist, 7-(3-((2-((phenylamino)carbonyl)hydrazino)methyl)-7-oxabicyclo(2.2.1)hept-2-yl)-5-heptenoic acid (SQ29548). Pretreatment of myocytes with an inhibitor of inositol trisphosphate (IP(3)) formation, gentamicin, or with an inhibitor of IP(3) receptors, 2-aminoethoxydiphenylborate (2-APB), blocked the increase in intracellular calcium. In vivo pretreatment of anesthetized rabbits with either gentamicin or 2-APB subsequently inhibited the formation of ventricular arrhythmias elicited by U46619. These data support the hypothesis that TXA(2) can induce arrhythmias via a direct action on cardiac myocytes. Furthermore, these arrhythmogenic actions were blocked by inhibitors of the IP(3) pathway. In summary, this study provides novel evidence for direct TXA(2)-induced cardiac arrhythmias and provides a rationale for IP(3) as a potential target for the treatment of TXA(2)-mediated arrhythmias.

Purification of cardiac myocytes from human heart biopsies for gene expression analysis.

The collection of gene expression data from human heart biopsies is important for understanding the cellular mechanisms of arrhythmias and diseases such as cardiac hypertrophy and heart failure. Many clinical and basic research laboratories conduct gene expression analysis using RNA from whole cardiac biopsies. This allows for the analysis of global changes in gene expression in areas of the heart, while eliminating the need for more complex and technically difficult single-cell isolation procedures (such as flow cytometry, laser capture microdissection, etc.) that require expensive equipment and specialized training. The abundance of fibroblasts and other cell types in whole biopsies, however, can complicate gene expression analysis and the interpretation of results. Therefore, we have designed a technique to quickly and easily purify cardiac myocytes from whole cardiac biopsies for RNA extraction. Human heart tissue samples were collected, and our purification method was compared with the standard nonpurification method. Cell imaging using acridine orange staining of the purified sample demonstrated that >98% of total RNA was contained within identifiable cardiac myocytes. Real-time RT-PCR was performed comparing nonpurified and purified samples for the expression of troponin T (myocyte marker), vimentin (fibroblast marker), and alpha-smooth muscle actin (smooth muscle marker). Troponin T expression was significantly increased, and vimentin and alpha-smooth muscle actin were significantly decreased in the purified sample (n = 8; P < 0.05). Extracted RNA was analyzed during each step of the purification, and no significant degradation occurred. These results demonstrate that this isolation method yields a more purified cardiac myocyte RNA sample suitable for downstream applications, such as real-time RT-PCR, and allows for more accurate gene expression changes in cardiac myocytes from heart biopsies.

Acute and long-term proteome changes induced by oxidative stress in the developing brain.

The developing mammalian brain experiences a period of rapid growth during which various otherwise innocuous environmental factors cause widespread apoptotic neuronal death. To gain insight into developmental events influenced by a premature exposure to high oxygen levels and identify proteins engaged in neurodegenerative and reparative processes, we analyzed mouse brain proteome changes at P7, P14 and P35 caused by an exposure to hyperoxia at P6. Changes detected in the brain proteome suggested that hyperoxia leads to oxidative stress and apoptotic neuronal death. These changes were consistent with results of histological and biochemical evaluation of the brains, which revealed widespread apoptotic neuronal death and increased levels of protein carbonyls. Furthermore, we detected changes in proteins involved in synaptic function, cell proliferation and formation of neuronal connections, suggesting interference of oxidative stress with these developmental events. These effects are age-dependent, as they did not occur in mice subjected to hyperoxia in adolescence.

Oxidative stress related DNA adducts in the liver of female rats fed with sunflower-, rapeseed-, olive- or coconut oil supplemented diets.

Both animal and epidemiological studies support an effect of fatty acid composition in the diet on cancer development, in particular on colon cancer. We investigated the modulating effect of supplementation of the diet of female F344 rats with sunflower-, rapeseed-, olive-, or coconut oil on the formation of the promutagenic, exocyclic DNA adducts in the liver, an organ where major metabolism of fatty acids takes place. 1,N(6)-ethenodeoxyadenosine (etheno-dA), 3,N(4)-ethenodeoxycytidine (etheno-dC) and 1,N(2)-propandodeoxyguanosine from 4-hydroxy-2-nonenal (HNE-dGp) were determined as markers for DNA-damage derived from lipid peroxidation products and markers for oxidative stress. 8-Oxo-deoxyguanosine (8-Oxo-dG) was also measured as direct oxidative stress marker. The body weight of the rats was not influenced by the four diets containing the different vegetable oils during the 4-week feeding period. Highest adduct levels of etheno-dC (430 +/- 181 adducts/10(9) parent bases), HNE-dGp (617 +/- 96 adducts/10(9) parent bases) and 8-Oxo-dG (37,400 +/- 12,200 adducts/10(9) parent bases) were seen in rats on sunflower oil diet (highest linoleic acid content). Highest adducts levels of etheno-dA (133 +/- 113 adducts/10(9) parent bases) were found in coconut oil diet (lowest content of linoleic acid). Weakly positive correlations between linoleic acid content in the four diet groups were only observed for levels of HNE-dGp and 8-Oxo-dG. Neither the diet based on olive oil (which contains mainly oleic acid) nor the diet based on rapeseed oil (containing alpha-linolenic acid) exerted any significant protective effect against oxidative DNA damage. Our results indicate that a high linoleic acid diet may contribute to oxidative stress in the liver of female rats leading to a marginal increase in oxidative DNA-damage.

Role of serotonin in thromboxane A2-induced coronary chemoreflex.

We reported previously that the thromboxane A(2) (TxA(2)) mimetic U-46619 stimulates cardiac vagal afferent nerves, eliciting a reflex decrease in heart rate (HR) and arterial blood pressure (ABP). The present experiments were designed to test the hypothesis that TxA(2) evokes these changes via the release of serotonin [5-hydroxytryptamine (5-HT)] and activation of the 5-HT(3) receptor. Injections of the 5-HT(3) antagonist tropisetron (1 mg of 3-tropanyl-indole-3-carboxylate or ICS-205-930) attenuated the decreases in HR and ABP induced by left atrial injections of U-46619 (20 microg). Tropisetron administration also eliminated the U-46619-induced increase in impulse frequency in a majority of cardiac, vagal afferent units tested. Measurement of serum 5-HT levels revealed an elevation in serum 5-HT levels after U-46619 injection in those rabbits that displayed a significant HR change following injection of U-46619. These results indicate that although other factors may also contribute to these reflex responses, the release of 5-HT and stimulation of the 5-HT(3) receptor plays a significant role in coronary reflexes induced by TxA(2).

Unaltered brain levels of 1,N2-propanodeoxyguanosine adducts of trans-4-hydroxy-2-nonenal in Alzheimer's disease.

In recent years, an important role for the pathogenesis of Alzheimer's disease (AD) has been ascribed to oxidative stress. Trans-4-hydroxy-2-nonenal, a product of lipid peroxidation, forms stable adducts with a variety of nucleophilic substituents such as thiols or amino moieties. Here, we report the quantification of 1,N2-propanodeoxyguanosine adducts of trans-4-hydroxy-2-nonenal (HNE-dGp) using the specific and very sensitive method of 32P-postlabeling of deoxyguanosine adducts derived from nuclear DNA in neuron rich areas of the hippocampus, the parietal cortex, and the cerebellum of postmortem brains from patients with AD and age matched controls. Adduct levels were highest in the hippocampus, followed by the cerebellum and parietal cortex irrespective of the disease. Neither age, postmortem delay time, gender, nor the extent of neurofibrillary deposits affected tissue adduct levels in the brain areas examined. Although distinctively present in the human brain, the level of HNE-dGp adducts appears not to be useful as a biomarker for AD.

Detection of 1,N2-propanodeoxyguanosine adducts of trans-4-hydroxy-2-nonenal after gavage of trans-4-hydroxy-2-nonenal or induction of lipid peroxidation with carbon tetrachloride in F344 rats.

The 1,N2-propanodeoxyguanosine adducts of trans-4-hydroxy-2-nonenal (HNE-dGp-adducts) were quantitated in tissues of rats treated with trans-4-hydroxy-2-nonenal (HNE) or carbon tetrachloride, respectively, using a 32P-postlabeling method. The method development was based on chemically synthesized HNE-1,N2-propanodeoxyguanosine adduct standard, which was characterized by NMR and mass spectra. The adducts were enriched by Nuclease P1. They were subsequently reacted with gamma-32P-ATP to give the respective 3'-5'-bisphosphates, which were two-directionally separated on PEI-cellulose-TLC and quantitated by autoradiography. The labeling efficiency for the adduct standard was 27%, and the recovery of spiked amounts of adduct standard in the enzymatical procedure was about 80%. Internal standard was used to eliminate methodological variations. The determination of the limit of quantitation in DNA from rat tissues by spiking of HNE-dGp-adduct standard revealed a sensitivity of about 20 HNE-dGp-adducts/10(9) normal nucleotides. Background levels of HNE-dGp-adducts in tissues of rats including liver, kidney, lung, colon and forestomach were found in the range of 18-158 adducts/10(9) nucleotides with relatively high adduct levels in the liver and low adduct levels in kidney, lung and colon. These background levels were statistically significantly increased by the factor of 2 in liver, lung, colon and forestomach after induction of lipid peroxidation by carbon tetrachloride. The finding that background HNE-dGp-adduct levels may be in context with different metabolic activities of the tissues and the increase of HNE-dGp-adduct levels after application of carbon tetrachloride indicate that HNE-dGp-adducts are an endogenous lesion and that they are probably formed from radical initiated lipid peroxidation.

Meperidine: second-line agent with first-line prescribing practices.

Results from this survey need to be viewed within the context of the small number of responses (20%), which may not fully represent the knowledge base of all house officers. Recommendations established by the Agency for Health Care Policy and Research (AHCPR) define standards of care for acute and chronic pain management and address many of the problems seen nationally with meperidine and its metabolite, normeperidine. Data from this survey have assisted us in providing educational programs which are in line with these guidelines for house officers.

Development of a (32)P-postlabeling method for the detection of 1,N(2)-propanodeoxyguanosine adducts of trans-4-hydroxy-2-nonenal in vivo.

A (32)P-postlabeling method was developed for the sensitive detection of 1,N(2)-propanodeoxyguanosine adducts of the lipid peroxidation product trans-4-hydroxy-2-nonenal in vivo. The method development was based on the chemically synthesized HNE-1, N(2)-propanodeoxyguanosine adduct standard, which was characterized by NMR and mass spectra. The adducts were enriched by nuclease P1. They were subsequently reacted with [gamma-(32)P]ATP to give the respective 3'-5'-bisphosphates, which were two-directionally separated on PEI-cellulose TLC and quantitated by autoradiography. The medium labeling efficiency for the mixture of the two pairs of diastereomers was 27%, and the recovery of spiked amounts of adduct standard in the enzymatical procedure was about 80%. The method is applicable for the separation and quantitation of HNE-dGp-propano adducts in vivo. It was applied to DNA from colon and brain tissue of untreated Fischer 344 rats and humans. The determination of the limit of quantitation in DNA from rat colon by spiking of adduct standard revealed a sensitivity of <21 adducts/10(9) nucleotides. The analytical quantitation of 4-HNE-dGp-propano adducts resulted in adduct-levels per 10(9) normal nucleotides +/- the standard deviation of 223.32 +/- 79.84 in rat colon tissue, 90.37 +/- 11.94 in rat brain tissue, 378.44 +/- 52.42 in human colon tissue, and 185.15 +/- 6.48 in human brain tissue. The results clearly demonstrate the applicability of this method for the sensitive detection of endogenously formed 1,N(2)-propanodeoxyguanosine adducts of trans-4-hydroxy-2-nonenal, a specific marker for the lipid peroxidation process.

Cardiac decompensation during an ischemic event weakens the predictive power of myocardial injury markers.

Our objection was to find determinants of long-term outcome in routine data collected for differential diagnosis of suspected acute myocardial infarction. Study population consisted of 263 discharged patients who were initially hospitalized for differential diagnosis of suspected acute myocardial infarction between October 1992 and January 1993. Follow-up time for all cause and cardiac mortality was 5 years. The variables studied as predictors of outcome were computerized ECG, peak creatine kinase isoenzyme MB, peak troponin I, radiographic evidence of pulmonary congestion (cardiac decompensation), treatment for hyperlipidemia, hypertension or diabetes, smoking, previous myocardial infarction, age and gender. Total mortality was 32% at 5 years, of which 77% (64/83) was of cardiac origin. Pulmonary congestion in chest X-ray was the most powerful predictor of outcome (RR=3.3, 95% CI=2.0-5.2, P<0.001). In multivariate analysis congestion (RR=3.3, CI=2.0-5.2) was the only independent predictor of 5-year total mortality in addition to age (RR=1.06, CI=1.04-1.08). These two variables together with previous myocardial infarction (RR=1.9, CI=1.2-3.1) and hyperlipidemia (RR=2. 0, CI=1.1-3.5) were independent predictors of cardiac mortality. Radiographic evidence of cardiac decompensation during hospitalization is a strong and independent predictor of long-term outcome in unselected patients with suspected AMI. The predictive power of cardiac markers is confined to patients without pulmonary congestion.