Biologically derived epicardial patch induces macrophage mediated pathophysiologic repair in chronically infarcted swine hearts.

There are nearly 65 million people with chronic heart failure (CHF) globally, with no treatment directed at the pathologic cause of the disease, the loss of functioning cardiomyocytes. We have an allogeneic cardiac patch comprised of cardiomyocytes and human fibroblasts on a bioresorbable matrix. This patch increases blood flow to the damaged heart and improves left ventricular (LV) function in an immune competent rat model of ischemic CHF. After 6 months of treatment in an immune competent Yucatan mini swine ischemic CHF model, this patch restores LV contractility without constrictive physiology, partially reversing maladaptive LV and right ventricular remodeling, increases exercise tolerance, without inducing any cardiac arrhythmias or a change in myocardial oxygen consumption. Digital spatial profiling in mice with patch placement 3 weeks after a myocardial infarction shows that the patch induces a CD45pos immune cell response that results in an infiltration of dendritic cells and macrophages with high expression of macrophages polarization to the anti-inflammatory reparative M2 phenotype. Leveraging the host native immune system allows for the potential use of immunomodulatory therapies for treatment of chronic inflammatory diseases not limited to ischemic CHF.

On demand delivery and analysis of single molecules on a programmable nanopore-optofluidic device.

Nanopore-based single nanoparticle detection has recently emerged as a vibrant research field with numerous high-impact applications. Here, we introduce a programmable optofluidic chip for nanopore-based particle analysis: feedback-controlled selective delivery of a desired number of biomolecules and integration of optical detection techniques on nanopore-selected particles. We demonstrate the feedback-controlled introduction of individual biomolecules, including 70S ribosomes, DNAs and proteins into a fluidic channel where the voltage across the nanopore is turned off after a user-defined number of single molecular insertions. Delivery rates of hundreds/min with programmable off-times of the pore are demonstrated using individual 70S ribosomes. We then use real-time analysis of the translocation signal for selective voltage gating of specific particles from a mixture, enabling selection of DNAs from a DNA-ribosome mixture. Furthermore, we report optical detection of nanopore-selected DNA molecules. These capabilities point the way towards a powerful research tool for high-throughput single-molecule analysis on a chip.

Do group dynamics affect colour morph clines during a range shift?

Species exhibiting colour polymorphism are thought to have an ecological advantage at the landscape scale, because spatial segregation of alternatively adapted ecotypes into diverse habitats can increase the species' niche breadth and thus confer greater geographic range size. However, morph frequencies are also influenced by intrapopulational processes such as frequency- or density-dependent social interactions. To identify how social feedback may affect clinal variation in morph frequencies, we investigated reciprocal interactions between morph-specific thermal tolerance, local climatic conditions and social environments, in the context of a colour-morph frequency cline associated with a recent range expansion in blue-tailed damselflies (Ischnura elegans) in Sweden. Cold tolerances of gynochromes (female-like female morph) were positively correlated with local gynochrome frequencies, suggesting a positive frequency-dependent fitness benefit. In contrast, androchrome (male-mimic female morph) cold tolerances were improved following recent exposure to cold weather, suggesting a beneficial environmental acclimation effect. Thus, according to an environment-matching hypothesis for clinal variation, androchrome frequencies should therefore increase towards the (cooler) range limit. In contrast to this prediction, gynochrome frequencies increased at the expanding range limit, consistent with a positive frequency-dependent social feedback that is beneficial when invading novel climates. Our results suggest that when phenotypes or fitnesses are affected by interactions with conspecifics, beneficial social effects on environmental tolerances may (i) facilitate range shifts, and (ii) reverse or counteract typical patterns of intraspecific interactions and environment-matching clines observed in stable populations observed over broader geographic scales.

The role of pleiotropy vs signaller-receiver gene epistasis in life history trade-offs: dissecting the genomic architecture of organismal design in social systems.

Traditional life history theory ignores trade-offs due to social interactions, yet social systems expand the set of possible trade-offs affecting a species evolution--by introducing asymmetric interactions between the sexes, age classes and invasion of alternative strategies. We outline principles for understanding gene epistasis due to signaller-receiver dynamics, gene interactions between individuals, and impacts on life history trade-offs. Signaller-receiver epistases create trade-offs among multiple correlated traits that affect fitness, and generate multiple fitness optima conditional on frequency of alternative strategies. In such cases, fitness epistasis generated by selection can maintain linkage disequilibrium, even among physically unlinked loci. In reviewing genetic methods for studying life history trade-offs, we conclude that current artificial selection or gene manipulation experiments focus on pleiotropy. Multi-trait selection experiments, multi-gene engineering methods or multiple endocrine manipulations can test for epistasis and circumvent these limitations. In nature, gene mapping in field pedigrees is required to study social gene epistases and associated trade-offs. Moreover, analyses of correlational selection and frequency-dependent selection are necessary to study epistatic social system trade-offs, which can be achieved with group-structured versions of Price's (1970) equation.

Corticosterone manipulation reveals differences in hierarchical organization of multidimensional reproductive trade-offs in r-strategist and K-strategist females.

Life history trade-offs are often hierarchical with decisions at one level affecting lower level trade-offs. We investigated trade-off structure in female side-blotched lizards (Uta stansburiana), which exhibit two evolved strategies: yellow-throated females are K-strategists and orange-throated are r-strategists. Corticosterone treatment was predicted to differentially organize these females' reproductive decisions. Corticosterone-treated yellow females suppressed reproduction but survived well, and augmented egg mass without decreasing clutch size. Conversely, corticosterone enhanced mortality and reproductive rates in orange females, and increased egg mass only after lengthy exposure. Corticosterone did not affect post-laying condition, suggesting that corticosterone increased egg mass through enhanced energy acquisition (income breeding). Corticosterone enhanced survival of lightweight females, but decreased survival of heavy females, introducing a foraging vs. predation trade-off. We conclude that rather than being a direct, functional relationship, observed trade-offs between offspring size and number represent evolved differences in hierarchical organization of multidimensional trade-offs, particularly in response to stress.

Suppression of lung inflammation in rats by prevention of NF-kappaB activation in the liver.

Activation of NF-kappaB and production of NF-kappaB-dependent chemokines are thought to be involved in the pathogenesis of neutrophilic lung inflammation. Calpain-1 inhibitor (CI-1) blocks activation of NF-kappaB by preventing proteolysis of the inhibitory protein IkappaB-alpha by the ubiquitin/proteasome pathway. We hypothesized that inhibition of proteasome function with CI-1 would block NF-kappaB activation in vivo after intraperitoneal (i.p.) treatment with bacterial lipopolysaccharide (LPS), and that NF-kappaB inhibition would be associated with suppression of chemokine gene expression and attenuation of neutrophilic alveolitis. We treated rats with a single i.p. injection of CI-1 (10 mg/kg) two hours prior to i.p. LPS (7 mg/kg). Treatment with Cl-1 prevented degradation of IkappaB-alpha and activation of NF-kappaB in the liver in response to LPS; however, Cl-1 treatment had no detected effect on NF-kappaB activation in lung tissue. CI-1 treatment prior to LPS resulted in 40% lower MIP-2 concentration in lung lavage fluid compared to rats treated with vehicle prior to LPS (502 +/- 112 pg/ml vs. 859 +/-144 pg/ml, P < 0.05). In addition, CI-1 treatment substantially inhibited LPS-induced neutrophilic alveolitis (2.7+ /- 1.2 x 10(5) vs. 43.7 +/- 12.2 x 10(5) lung lavage neutrophils, P < 0.01). These data indicate that NF-kappaB inhibition in the liver can alter lung inflammation induced by systemic LPS treatment and suggest that a liver-lung interaction contributes to the inflammatory response of the lung.

Multiorgan nuclear factor kappa B activation in a transgenic mouse model of systemic inflammation.

We utilized a line of transgenic mice expressing Photinus luciferase complementary DNA (cDNA) under the control of a nuclear factor kappa B (NF-kappaB)-dependent promoter (from the 5' human immunodeficiency virus-1 [HIV-1] long terminal repeat) to examine the role of NF-kappaB activation in the pathogenesis of systemic inflammation induced by bacterial endotoxin (lipopolysaccharide [LPS]). After intraperitoneal injection of E. coli LPS, these mice displayed a time- and dose-dependent, organ-specific pattern of luciferase expression, showing that NF-kappaB-dependent gene transcription is transiently activated in multiple organs by systemic LPS administration. Luciferase expression in liver could be specifically blocked by intravenous administration of replication-deficient adenoviral vectors expressing a dominant inhibitor of NF-kappaB (IkappaB-alphaDN), confirming that luciferase gene expression is a surrogate marker for NF-kappaB activation in this line of mice. After treatment with intraperitoneal LPS, the mice were found to have increased lung tissue messenger RNA (mRNA) expression of a variety of cytokines that are thought to be NF-kappaB-dependent, as well as elevated serum concentrations of presumed NF-kappaB-dependent cytokines. In lung tissue homogenates, a close correlation was identified between luciferase activity and KC levels. These studies show that systemic treatment with LPS orchestrates a multiorgan NF-kappaB-dependent response that likely regulates the pathobiology of systemic inflammation.

The location of protein S8 and surrounding elements of 16S rRNA in the 70S ribosome from combined use of directed hydroxyl radical probing and X-ray crystallography.

Ribosomal protein S8, which is essential for the assembly of the central domain of 16S rRNA, is one of the most thoroughly studied RNA-binding proteins. To map its surrounding RNA in the ribosome, we carried out directed hydroxyl radical probing of 16S rRNA using Fe(II) tethered to nine different positions on the surface of protein S8 in 70S ribosomes. Hydroxyl radical-induced cleavage was observed near the classical S8-binding site in the 620 stem, and flanking the other S8-footprinted regions of the central domain at the three-helix junction near position 650 and the 825 and 860 stems. In addition, cleavage near the 5' terminus of 16S rRNA, in the 300 region of its 5' domain, and in the 1070 region of its 3'-major domain provide information about the proximity to S8 of RNA elements not directly involved in its binding. These data, along with previous footprinting and crosslinking results, allowed positioning of protein S8 and its surrounding RNA elements in a 7.8-A map of the Thermus thermophilus 70S ribosome. The resulting model is in close agreement with the extensive body of data from previous studies using protein-protein and protein-RNA crosslinking, chemical and enzymatic footprinting, and genetics.

The April 8, 1998 tornado: assessment of the trauma system response and the resulting injuries.

BACKGROUND: On April 8, 1998, an F5 tornado touched down in two counties of Alabama producing a wide path of destruction. The presence of a regional trauma system in this area presents an opportunity to evaluate the effectiveness of the system in responding to the victims of this natural disaster. METHODS: Emergency room logs and the regional trauma system database were searched for all patients treated for injuries sustained from the tornado, and medical records were reviewed for demographic information, mode of transportation to hospital, injuries, treatment, and outcome. Fatalities were identified by means of the coroner's office. RESULTS: A total of 224 patients were evaluated at nine area hospitals, of whom 63 (28%) required admission. There were 32 deaths: 30 persons were dead at the scene, and 2 patients subsequently died at Level I trauma centers. Among patients with nonfatal injuries, 39% were managed at Level I facilities, 46% at Level III facilities, and 15% at nontrauma facilities. Forty patients (55%) seen at Level I facilities required admission compared with 15 patients (17%) at Level III facilities and 8 patients (29%) at nontrauma facilities; Level I facilities also had the highest Injury Severity Score. Of patients requiring admission, 83% were transported by emergency medical services; these patients also had the highest Injury Severity Score. CONCLUSION: The regional trauma system facilitated appropriate and efficient triage to system hospitals, routing the most severely injured patients to the Level I centers without overwhelming them with the more numerous, less severely injured patients.

Differential NF-kappaB activation after intratracheal endotoxin.

We examined the relationship between nuclear factor (NF)-kappaB DNA binding activity, cytokine gene expression, and neutrophilic alveolitis in rats after intratracheal (IT) instillation of endotoxin [lipopolysaccharide (LPS)]. NF-kappaB activation in lung tissue mirrored neutrophilic alveolitis after IT LPS instillation, with NF-kappaB activation and neutrophilic influx beginning 2 h after IT LPS doses of 0.01 mg/kg or greater. In lung lavage fluid cells, however, transient NF-kappaB activation was present in alveolar macrophages by 15 min after IT LPS instillation, followed by a second peak of NF-kappaB activation corresponding to the onset on neutrophilic alveolitis. For cytokines thought to be NF-kappaB dependent, two different patterns of mRNA expression were found. Interleukin (IL)-1alpha, IL-1beta, and tumor necrosis factor-alpha showed increased mRNA by 30 min after IT LPS instillation, but IL-6- and cytokine-induced neutrophil chemoattractant mRNAs were not substantially increased until 2 h after IT LPS instillation. Therefore, IT LPS causes differential NF-kappaB activation in air space cells and lung tissue, which likely determines production of key cytokines and directs the evolution of neutrophilic alveolitis.

Chemotactic gradients predict neutrophilic alveolitis in endotoxin-treated rats.

We hypothesized that the intensity of neutrophilic alveolitis is related to establishing a gradient of neutrophil attractant chemokines across the alveolar-capillary barrier. In these experiments, a positive chemokine gradient toward the alveoli was induced by intratracheal instillation of endotoxin in rats (IT LPS). Alteration of the chemotactic gradient was induced by combining IT LPS (0.1 mg/kg) with an intraperitoneal injection of endotoxin (IP LPS, 6.0 mg/kg). Bronchoalveolar lavage (BAL) and peripheral blood cell counts and differentials, and lavage and serum CXC chemokines were measured 4 h after LPS treatment. Compared with IT LPS treatment alone, IP + IT LPS resulted in a 30-fold reduction in neutrophil (PMN) count in BAL and a decreased percentage of PMNs in lavage (from 82 to 24%, p < 0.01). Total lung myeloperoxidase activity, a reflection of total PMN burden, was increased in all three treatment groups compared with the control group, but differences were not apparent between treatment groups. For the rat CXC chemokines MIP-2 and CINC, high concentrations were detected in BAL from both IT and IP + IT LPS groups; however, significantly higher concentrations were found in the sera of rats treated with IP + IT LPS compared with IT LPS alone. The calculated chemokine BAL-serum gradients were significantly higher for both MIP-2 and CINC in the IT LPS group than in the IT + IP LPS or IP LPS group, and correlated with neutrophil influx into the alveolar spaces 4 h after LPS treatment. In addition, the BAL-serum MIP-2 gradient was found to be increased 24 h after IP LPS, which is the time point of peak neutrophilic alveolitis. In summary, these data show that local chemokine gradients predict the intensity of neutrophilic alveolitis after treatment with endotoxin. Interventions to limit neutrophilic alveolitis could either be targeted to block local lung chemokine production or, paradoxically, to increase systemic production of chemokines.

Characterization of functionally active subribosomal particles from Thermus aquaticus.

Peptidyl transferase activity of Thermus aquaticus ribosomes is resistant to the removal of a significant number of ribosomal proteins by protease digestion, SDS, and phenol extraction. To define the upper limit for the number of macromolecular components required for peptidyl transferase, particles obtained by extraction of T. aquaticus large ribosomal subunits were isolated and their RNA and protein composition was characterized. Active subribosomal particles contained both 23S and 5S rRNA associated with notable amounts of eight ribosomal proteins. N-terminal sequencing of the proteins identified them as L2, L3, L13, L15, L17, L18, L21, and L22. Ribosomal protein L4, which previously was thought to be essential for the reconstitution of particles active in peptide bond formation, was not found. These findings, together with the results of previous reconstitution experiments, reduce the number of possible essential macromolecular components of the peptidyl transferase center to 23S rRNA and ribosomal proteins L2 and L3. Complete removal of ribosomal proteins from T. aquaticus rRNA resulted in loss of tertiary folding of the particles and inactivation of peptidyl transferase. The accessibility of proteins in active subribosomal particles to proteinase hydrolysis was increased significantly after RNase treatment. These results and the observation that 50S ribosomal subunits exhibited much higher resistance to SDS extraction than 30S subunits are compatible with a proposed structural organization of the 50S subunit involving an RNA "cage" surrounding a core of a subset of ribosomal proteins.

Nuclear factor kappa B: a pivotal role in the systemic inflammatory response syndrome and new target for therapy.

NF-kappaB is an important transcription factor complex that appears to play a fundamental role in regulating acute inflammation through activation of the cytokine cascade and production of other pro-inflammatory mediators. There is increasing evidence that NF-kappaB is important in the pathobiology of disease states such as SIRS, MODS and ARDS; therefore, therapeutic interventions aimed at limiting NF-kappaB activation and down-regulating production of inflammatory mediators could prove to be beneficial in decreasing host-derived tissue injury and organ dysfunction. Specific interventions that hold promise for suppressing NF-kappaB activation include the use of antioxidants, inhibition of NIK and the IKK signalsome, treatment with proteasome inhibitors, induction of endotoxin tolerance and, possibly the use of corticosteroids in selected patients.

Reafference and attractors in the olfactory system during odor recognition.

Olfactory bulb activity has been postulated to be chaotic, as measured in the EEG, and to be subject to an attractor with many "wings" enabling classification of different learned odor classes. Two parallel questions are thus addressed by the work presented here: (1) what is the evidence for attractors in the olfactory system, which can mediate learned odor classes? and (2) how does the olfactory system enter a specific attractor or attractor wing associated with the learned odor during the classification process? Both of these questions address the wider notion of endogenous activity preparing the system for an expected stimulus, which is at the basis of the reafference principle. By viewing the brain as a distributed complex dynamical system with global attractors, these questions can be answered together. Rats were implanted with bipolar macroelectrodes in the Olfactory Bulb (OB), Prepyriform Cortex (PPC), Entorhinal Cortex (EC), and Dentate Gyrus (DG), and then trained in an operant paradigm to press a bar for a reward in the presence of one odor and to receive no reward in the presence of another odor. Local Field Potentials (LFP) were recorded simultaneously from the structures during the operant task. We present evidence for three endogenous events: (1) preafference, which is manifested both by the EC entering an attractor and a mid-range signal (15-30 Hz) which appears to be passed from the EC to the OB just before the OB enters an attractor; (2) afference, where the OB enters an attractor during the odor recognition period of the experiment and the LFP recordings indicate that the OB drives the other structures in all frequency bands, especially the high gamma band (65-100 Hz) associated with the OB burst frequency; and (3) reafference or post-afference, which is accompanied by a lower frequency gamma band signal (40-60 Hz) originating in the PPC and passed to both the OB and the EC just before the onset of the motor response to the odor. We use a new method, NECTAR (Nonparametric Exact Contingency Table Association Routine), related to mutual information, to verify what is seen with coherence and phase estimates, the apparent driving of each structure at different times in the odor trials, and to display evidence for non-periodic attractors governing both individual physiological structures and the system of structures. This is the first evidence of an endogenous, limbic event associated with sensory perceptual tuning in a mammal. These results are also the first experimental confirmation that the attractors governing olfactory activity involve multiple sites in the olfactory/limbic system and implement the process of attention.

The immune system in patients with renal failure. Part 1: Review of immune function.

Patients with renal failure often succumb to infection. Alterations in the immune system of these patients, whether inherent to the disease process or precipitated by therapeutic regimens, undermine the promotion of wellness. Part 1 of this article provides a comprehensive understanding of normal immune system function. Part 2 discusses common immune abnormalities related to renal failure and its treatment. In addition, there is an overview of assessment techniques that can assist nephrology nurses in identifying alterations in a patient's immunologic status and direct care to prevent complications from altered immune function.

Part 2: Effects of renal failure and its treatment on the immune system and assessment of immune system function.

Patients with renal failure often succumb to infection. Alterations in the immune system of these patients, whether inherent to the disease process or precipitated by therapeutic regimens, undermine the promotion of wellness. This article provides a comprehensive understanding of common immune abnormalities, and assessment techniques that can assist nephrology nurses in identifying problems in a patient's immunologic status and direct care to prevent infections, to treat or prevent organ rejection, and to monitor for signs and symptoms of malignancies.