Remodeling of the transverse tubular system after myocardial infarction in rabbit correlates with local fibrosis: A potential role of biomechanics.

The transverse tubular system (t-system) of ventricular cardiomyocytes is essential for efficient excitation-contraction coupling. In cardiac diseases, such as heart failure, remodeling of the t-system contributes to reduced cardiac contractility. However, mechanisms of t-system remodeling are incompletely understood. Prior studies suggested an association with altered cardiac biomechanics and gene expression in disease. Since fibrosis may alter tissue biomechanics, we investigated the local microscopic association of t-system remodeling with fibrosis in a rabbit model of myocardial infarction (MI). Biopsies were taken from the MI border zone of 6 infarcted hearts and from 6 control hearts. Using confocal microscopy and automated image analysis, we quantified t-system integrity (ITT) and the local fraction of extracellular matrix (fECM). In control, fECM was 18 ± 0.3%. ITT was high and homogeneous (0.07 ± 0.006), and did not correlate with fECM (R2 = 0.05 ± 0.02). The MI border zone exhibited increased fECM within 3 mm from the infarct scar (30 ± 3.5%, p < 0.01 vs control), indicating fibrosis. Myocytes in the MI border zone exhibited significant t-system remodeling, with dilated, sheet-like components, resulting in low ITT (0.03 ± 0.008, p < 0.001 vs control). While both fECM and t-system remodeling decreased with infarct distance, ITT correlated better with decreasing fECM (R2 = 0.44) than with infarct distance (R2 = 0.24, p < 0.05). Our results show that t-system remodeling in the rabbit MI border zone resembles a phenotype previously described in human heart failure. T-system remodeling correlated with the amount of local fibrosis, which is known to stiffen cardiac tissue, but was not found in regions without fibrosis. Thus, locally altered tissue mechanics may contribute to t-system remodeling.

Cellular electrophysiological principles that modulate secretion from synovial fibroblasts.

Rheumatoid arthritis (RA) is a progressive disease that affects both pediatric and adult populations. The cellular basis for RA has been investigated extensively using animal models, human tissues and isolated cells in culture. However, many aspects of its aetiology and molecular mechanisms remain unknown. Some of the electrophysiological principles that regulate secretion of essential lubricants (hyaluronan and lubricin) and cytokines from synovial fibroblasts have been identified. Data sets describing the main types of ion channels that are expressed in human synovial fibroblast preparations have begun to provide important new insights into the interplay among: (i) ion fluxes, (ii) Ca2+ release from the endoplasmic reticulum, (iii) intercellular coupling, and (iv) both transient and longer duration changes in synovial fibroblast membrane potential. A combination of this information, knowledge of similar patterns of responses in cells that regulate the immune system, and the availability of adult human synovial fibroblasts are likely to provide new pathophysiological insights.

Analyzing Remodeling of Cardiac Tissue: A Comprehensive Approach Based on Confocal Microscopy and 3D Reconstructions.

Microstructural characterization of cardiac tissue and its remodeling in disease is a crucial step in many basic research projects. We present a comprehensive approach for three-dimensional characterization of cardiac tissue at the submicrometer scale. We developed a compression-free mounting method as well as labeling and imaging protocols that facilitate acquisition of three-dimensional image stacks with scanning confocal microscopy. We evaluated the approach with normal and infarcted ventricular tissue. We used the acquired image stacks for segmentation, quantitative analysis and visualization of important tissue components. In contrast to conventional mounting, compression-free mounting preserved cell shapes, capillary lumens and extracellular laminas. Furthermore, the new approach and imaging protocols resulted in high signal-to-noise ratios at depths up to 60 µm. This allowed extensive analyzes revealing major differences in volume fractions and distribution of cardiomyocytes, blood vessels, fibroblasts, myofibroblasts and extracellular space in control vs. infarct border zone. Our results show that the developed approach yields comprehensive data on microstructure of cardiac tissue and its remodeling in disease. In contrast to other approaches, it allows quantitative assessment of all major tissue components. Furthermore, we suggest that the approach will provide important data for physiological models of cardiac tissue at the submicrometer scale.

IKK-2 inhibitor TPCA-1 represses nasal epithelial inflammation in vitro.

BACKGROUND: Nasal polyposis (NP) is considered a subgroup within chronic rhinosinusitis. NP can be further subdivided into aspirin sensitive- and aspirin tolerant types (ASNP/ ATNP). Although the true etiology of NP has not been identified so far, it is agreed that NP represents an inflammatory disease of the nasal mucosa. Alterations of cellular kinase activities including that of IKK-2 might play a role in this inflammatory process. METHODS: Paraffin sections of ASNP, ATNP and controls were immunostained with Phospho-IkB-α antibody that detects the direct IKK-2 product (IkB-α. Intensity of epithelial staining was analysed semi-quantitatively by two independent observers. In cultured nasal polyp epithelial cells (NPECs) epithelial derived cytokines IL-8 and GRO α were induced by TNF-α or Staphylococcal supernatants and subsequently repressed by IKK-2 inhibitor TPCA-1. RESULTS: Significant Phospho-IkB-α staining was observed in the nasal epithelium of ASNP compared to ATNP and controls suggesting strong IKK-2 activation in patients with ASNP in vivo. In vitro, pro-inflammatory cytokines IL-8 and GRO-α in NPECs were significantly repressed by TPCA-1. CONCLUSION: IKK-2 activity is increased in the subgroup of ASNP. IL-8 and GRO-α responses were repressed by IKK-2 inhibitor TPCA-1 in vitro. IKK-2 inhibitors might represent a potential target for anti-inflammatory intervention in ASNP.

Minimum Information about a Cardiac Electrophysiology Experiment (MICEE): standardised reporting for model reproducibility, interoperability, and data sharing.

Cardiac experimental electrophysiology is in need of a well-defined Minimum Information Standard for recording, annotating, and reporting experimental data. As a step towards establishing this, we present a draft standard, called Minimum Information about a Cardiac Electrophysiology Experiment (MICEE). The ultimate goal is to develop a useful tool for cardiac electrophysiologists which facilitates and improves dissemination of the minimum information necessary for reproduction of cardiac electrophysiology research, allowing for easier comparison and utilisation of findings by others. It is hoped that this will enhance the integration of individual results into experimental, computational, and conceptual models. In its present form, this draft is intended for assessment and development by the research community. We invite the reader to join this effort, and, if deemed productive, implement the Minimum Information about a Cardiac Electrophysiology Experiment standard in their own work.

Models of cardiac tissue electrophysiology: progress, challenges and open questions.

Models of cardiac tissue electrophysiology are an important component of the Cardiac Physiome Project, which is an international effort to build biophysically based multi-scale mathematical models of the heart. Models of tissue electrophysiology can provide a bridge between electrophysiological cell models at smaller scales, and tissue mechanics, metabolism and blood flow at larger scales. This paper is a critical review of cardiac tissue electrophysiology models, focussing on the micro-structure of cardiac tissue, generic behaviours of action potential propagation, different models of cardiac tissue electrophysiology, the choice of parameter values and tissue geometry, emergent properties in tissue models, numerical techniques and computational issues. We propose a tentative list of information that could be included in published descriptions of tissue electrophysiology models, and used to support interpretation and evaluation of simulation results. We conclude with a discussion of challenges and open questions.

Staphylococcus aureus invades the epithelium in nasal polyposis and induces IL-6 in nasal epithelial cells in vitro.

BACKGROUND: Staphylococcus aureus has been associated with chronic rhinosinusitis with nasal polyps (CRSwNP) pathogenesis but its role is still controversially discussed. Here, we demonstrate S. aureus detection in the mucosa of CRSwNP. In addition, intracellular residency of S. aureus in nasal polyp epithelial cells (NPECs) and its capability to induce TH-2 cytokines were analyzed in vitro. METHODS: Staphylococcus aureus detection in CRSwNP (n = 25), CRS without polyps (CRSsNP, n = 5), and turbinate mucosa (TM, n = 10) was performed by peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) and microbial cultivation from tissue biopsies. Intracellular residency was examined by intracellular persistence assay and electron microscopy. IL-6 and IL-13 responses to S. aureus infection and supernatants were quantified by ELISA. RESULTS: Peptide nucleic acid-fluorescence in situ hybridization positive bacterial cells were significantly increased in the epithelium of CRSwNP (17/25) compared to CRSsNP (0/5) and TM (1/10). Good concordance of PNA-FISH results and S. aureus cultivation was found applying Cohen's κ for CRSwNP (κ = 0.841) and TM (κ = 1.0). Intracellular persistence assay with S. aureus strain Newman and its corresponding small-colony variant mutant strain III33 demonstrated intracellular survival and replication of S. aureus within NPECs. Both S. aureus strains significantly induced IL-6 but not IL-13 in infected NPECs and in NPECs challenged with corresponding staphylococcal supernatants. CONCLUSION: Invasion of the epithelium by S. aureus was a phenomenon seen predominantly in CRSwNP. Regardless of an intra- or extracellular localization in the epithelium, S. aureus is capable to induce IL-6 synthesis in vitro and thus may contribute to the TH-2 cytokine pattern in CRSwNP.

[Nasal surgery in patients with systemic disorders]. / Rhinochirurgie bei Systemerkrankungen.

Systemic disorders represent a heterogenous group of diseases which can primarily manifest at the nose and sinuses as limited disease or secondarily as part of the systemic involvement. Rhinologists therefore play an important role in the diagnostic but also therapeutic process. Although therapy of systemic disorders is primary systemic, additional nasal surgery may become necessary. Surgical procedures include sinus surgery for the treatment of complications of the orbit and the lacrimal duct, septorhinoplasty due to saddle nose deformity and closure of septal perforation. Since many systemic disorders represent very rare diseases, recommendations are based on the analysis of single case reports and studies with a limited number of patients. Even though data is still limited, experiences published so far have shown that autogenous cartilage or bone transplants can be used in nasal reconstruction of deformities caused by tuberculosis, leprosy, wegener's granulomatosis, sarcoidosis and relapsing polychondritis. Experiences gained from these diseases support our observation that well-established techniques of septorhinoplasty can be used in systemic disorders as well. However, reaching a state of remission is an essential condition before considering any rhinosurgery in these patients. Under these circumstances revision surgery has to be expected more frequently compared to the typical collective of patients undergoing septorhinoplasty. These observations in part may also be useful for the treatment of nasal septal perforations since implantation of cartilage- or bone grafts represents an essential step in the closure of septal perforations. Apart from the treatment of orbital complications, sinus surgery has been proven beneficial in reducing nasal symptoms and increasing quality of life in patients refractory to systemic treatment.

Anti-inflammatory effects of ciprofloxacin in S. aureus Newman induced nasal inflammation in vitro.

OBJECTIVES: Chronic rhinosinusitis (CRS) is a chronic inflammatory disease of the nasal mucosa. Recent studies suggest that S. aureus enterotoxins may play an etiologic role in the development of CRS. Apart from surgery and repeated courses of steroids, macrolide antibiotics have been reported to exert anti-inflammatory effects in CRS. Similar effects have been reported for fluoroquinolones on various cell types. Since these effects have poorly been characterized in CRS, we examined anti-inflammatory effects of ciprofloxacin on human nasal epithelial cells (HNECs). METHODS: Inflammation was induced in HNECs cultured from nasal turbinate mucosa with supernatants of S. aureus Newman for 12 hours. Subsequently, HNECs were coincubated with S. aureus Newman and ciprofloxacin (1.5 x 10-5 M), clarithromycin (10-6 M) or prednisolone (10-5 M) for another 12 hours. IL-8 synthesis was quantified after 12 and 24 hours by ELISA. RESULTS: Stimulation with S. aureus Newman supernatants was associated with an increase of IL-8 synthesis after 12 hours in all experiments. During the second 12 hours, IL-8 synthesis decreased and this effect was independent from any stimulus or inhibitor. However, coincubation of HNECs with ciprofloxacin was associated with a more extensive decrease of IL-8 synthesis. Similarly, addition of clarithromycin was associated with a reduction of IL-8 synthesis although this effect was not significant. Coincubation with prednisolone resulted in a significant reduction of IL-8 levels. CONCLUSION: Ciprofloxacin exerts anti-inflammatory effects in S. aureus Newman driven nasal inflammation. Inhibitory effects were comparable to those of prednisolone and clarithromycin.

Experimental and computational studies of strain-conduction velocity relationships in cardiac tissue.

Velocity of electrical conduction in cardiac tissue is a function of mechanical strain. Although strain-modulated velocity is a well established finding in experimental cardiology, its underlying mechanisms are not well understood. In this work, we summarized potential factors contributing to strain-velocity relationships and reviewed related experimental and computational studies. We presented results from our experimental studies on rabbit papillary muscle, which supported a biphasic relationship of strain and velocity under uni-axial straining conditions. In the low strain range, the strain-velocity relationship was positive. Conduction velocity peaked with 0.59 m/s at 100% strain corresponding to maximal force development. In the high strain range, the relationship was negative. Conduction was reversibly blocked at 118+/-1.8% strain. Reversible block occurred also in the presence of streptomycin. Furthermore, our studies revealed a moderate hysteresis of conduction velocity, which was reduced by streptomycin. We reconstructed several features of the strain-velocity relationship in a computational study with a myocyte strand. The modeling included strain-modulation of intracellular conductivity and stretch-activated cation non-selective ion channels. The computational study supported our hypotheses, that the positive strain-velocity relationship at low strain is caused by strain-modulation of intracellular conductivity and the negative relationship at high strain results from activity of stretch-activated channels. Conduction block was not reconstructed in our computational studies. We concluded this work by sketching a hypothesis for strain-modulation of conduction and conduction block in papillary muscle. We suggest that this hypothesis can also explain uni-axially measured strain-conduction velocity relationships in other types of cardiac tissue, but apparently necessitates adjustments to reconstruct pressure or volume related changes of velocity in atria and ventricles.

Proinflammatory impact of Staphylococcus epidermidis on the nasal epithelium quantified by IL-8 and GRO-alpha responses in primary human nasal epithelial cells.

BACKGROUND: Bacterial etiology of chronic rhinosinusitis (CRS) still remains controversial. Whereas Staphylococcus aureus enterotoxins have been detected in CRS, the impact of Staphylococcus epidermidis, a major commensal inhabitant of the nose, has not been studied. Among others, serine and cysteine proteases have been identified as factors of virulence in S. epidermidis. METHODS: S. epidermidis was examined in tissue biopsies of 30 CRS patients (16 with nasal polyposis) using standard procedures. Primary human nasal epithelial cells from inferior nasal turbinates (HNECs), from nasal polyps (NPECs) and A549 airway epithelial cells were stimulated with S. epidermidis supernatants DSM20044 or ATCC35984 and the IL-8 and GRO-alpha response was quantified by ELISA. Protease-triggered chemokine responses and involvement of NF-kappaB were investigated by addition of protease or NF-kappaB inhibitors. Activation of NF-kappaB was demonstrated by quantitative DNA binding assay. RESULTS: S. epidermidis was the most frequently isolated bacteria in the majority of CRS patients. HNECs and NPECs revealed no different IL-6 and IL-8 synthesis following stimulation with DSM20044 or ATCC35984. Stimulation of HNECs and A549 cells with S. epidermidis supernatants resulted in increased IL-8 and GRO-alpha expression which could be suppressed by the serine protease inhibitor AEBSF and the NF-kappaB inhibitor BAY 11 but not by the cysteine protease inhibitor E64. Results obtained for A549 cells were similar to HNECs. CONCLUSION: S. epidermidis was present in the majority of CRS specimens. Proinflammatory impact of S. epidermidis supernatants on nasal epithelial cells was demonstrated by serine protease-triggered and NF-kappaB-dependent chemokine responses.

PAR-2 activation regulates IL-8 and GRO-alpha synthesis by NF-kappaB, but not RANTES, IL-6, eotaxin or TARC expression in nasal epithelium.

BACKGROUND: The effects of protease-activated receptor-2 (PAR-2) stimulation on inflammation mechanisms of chronic rhinosinusitis (CRS) are still unknown. METHODS: PAR-2 receptor expression was investigated by immunohistochemistry and Taqman mRNA analysis in the mucosa of different rhinosinusitis entities. In primary nasal epithelial cell cultures, the function of PAR-2 and its ability to produce CXC, CC chemokines, and IL-6 were measured by calcium mobilization and stimulation tests. Inhibition tests were performed using cortisone, serine protease inhibitors, cysteine protease inhibitors, Pertussis toxin (PTX) and nuclear transcription factor (NF-kappaB) inhibition (BAY 11-7085). Signal transduction pathways were analysed by electromobility shift assays (EMSA) and NF-kappaB binding studies. RESULTS: The expression of PAR-2 was found to be increased in CRS specimens. The activation of PAR by trypsin or PAR-2-specific activating peptide (AP) caused an increase in cytosolic calcium, as well as the release of the CXC chemokines IL-8 and growth-related oncogene (GRO)-alpha, but not the release of CC chemokines or IL-6. AP-induced CXC chemokine was sensitive to PTX and activation of NF-kappaB was inhibited by BAY11-7085. Furthermore, a serine protease inhibitor significantly inhibited chemokine synthesis stimulated by trypsin and culture supernatants of staphylococci, whereas steroids and cysteine protease inhibitors had little effect. CONCLUSION: PAR-2 plays a role in serine protease-mediated regulation - staphylococcal and non-staphylococcal origin - of IL-8 and GRO-alpha in nasal epithelial cells, but not in the regulation of CC chemokines. PAR-2 may therefore be involved in the pathophysiology of CRS and NP at different sites of activation, namely (i) proteases, (ii) the PAR-2 receptor itself or (iii) the application of novel agents that block NF-kappaB/IkappaB-alpha signalling.

Primary role of growth-related oncogene-alpha and granulocyte chemotactic protein-2 as neutrophil chemoattractants in chronic rhinosinusitis.

BACKGROUND: The aetiology of chronic rhinosinusitis (CRS) remains unclear. The purpose of this study was to investigate neutrophil-attracting chemokine patterns in CRS without nasal polyposis. METHODS: The biological activity of the chemokines was identified using a two-step high-performance liquid chromatography (HPLC) technique combined with a bioassay in extracts from 55 CRS patients, and in the turbinate mucosa (TM) of patients (N=51) undergoing septumplasty. The biologic activity of each chemokine was assessed using blocking antibodies to chemokines. Immunolocalization of detected neutrophil chemokines was performed by quantitative evaluation of immunohistochemistry. Besides, PCR analysis was performed to quantify neutrophil chemokine mRNA. RESULTS: In CRS, the chemokines primarily detected by two-step HPLC were growth-related oncogene-alpha (GRO-alpha) and the granulocyte chemotactic protein-2 (GCP-2). Blocking of GCP-2 and GRO-alphad each resulted in chemotaxis inhibition rates of 43.3% and 35.9%, respectively, whereas anti-IL-8 and anti-ENA-78 had no effect. Both GCP-2 and GRO-alphad were generally synthesized by the surface epithelium and mucosal glands while GRO-alpha in particular was synthesized by endothelial cells, as shown by immunohistochemistry. The concentrations of the chemokines IL-8 and epithelial cell-derived neutrophil attractant-78 (ENA-78) were low in CRS and TM. CONCLUSION: It appears that both GRO-alpha and GCP-2 contribute to neutrophil chemotaxis in CRS, whereas IL-8 and ENA-78 appear to be of secondary importance for the chemotaxis of neutrophils in this condition. The expression of chemokines in mucosal gland cells is the main phenomenon involved in constitutive neutrophil chemotaxis in the TM.

Induction of CXC chemokines in A549 airway epithelial cells by trypsin and staphylococcal proteases - a possible route for neutrophilic inflammation in chronic rhinosinusitis.

While various microorganisms have been recovered from patients with chronic rhinosinusitis, the inflammatory impact of virulence factors, in particular proteases from Staphylococcus aureus and coagulase negative staphylococci on the nasal epithelium, has not yet been investigated. Expression of CXC chemokines was determined in the epithelium of patients with chronic rhinosinusitis by immunohistochemistry. In a cell culture system of A549 respiratory epithelial cells, chemokine levels were quantified by enzyme-linked immunosorbent assay (ELISA) after stimulation with supernatants originating from three different staphylococcal strains or with trypsin, representing a serine protease. Inhibition experiments were performed with prednisolone, with the serine protease inhibitor 4-(2-aminoethyl)-benzenesulphonylfluoride (AEBSF) and with the nuclear transcription factor (NF)-kappaBeta inhibitor (2E)-3-[[4-(1,1-dimethylethyl)phenyl]sulphonyl]-2-propenenitrite (BAY) 11-7085. Electromobility shift assays (EMSA) were used to demonstrate NF-kappaB-dependent protein synthesis. CXC chemokines interleukin (IL)-8, growth-related oncogene alpha (GRO-alpha) and granulocyte chemotactic protein-2 (GCP-2) were expressed in the patients' epithelium whereas epithelial cell-derived neutrophil attractant 78 (ENA-78) was rarely detected. In A549 cells, chemokines IL-8, ENA-78 and GRO-alpha but not GCP-2 were induced by trypsin and almost equal levels were induced by staphylococcal supernatants. IL-8, GRO-alpha and ENA-78 synthesis was suppressed almost completely by AEBSF and BAY 11-7085, whereas prednisolone reduced chemokine levels differentially dependent on the supernatant added. CXC chemokines were detectable in the epithelium of patients with chronic rhinosinusitis. Staphylococcal serine proteases induced CXC chemokines in A549 cells, probably by the activation of proteases activated receptors, and thus might potentially be involved in neutrophilic inflammation in chronic sinusitis.

[Malignant fibrous histiocytoma in the parotid gland. Case series and literature review]. / Maligne fibröse Histiozytome in der Glandula parotis. Fallserie und Literaturübersicht.

BACKGROUND: Malignant fibrous histiocytoma (MFH), a soft tissue sarcoma that is predominantly localized in the extremities and retroperitoneum, rarely occurs in the head and neck. This study presents the clinicopathological features of three patients with MFH of the parotid gland treated at the ENT department of the University Hospital of Muenster (Germany) between 1991 and 2002. PATIENTS: The clinical course of all three patients was defined by a rapidly growing mass in the parotid area. Surgical therapy was the first treatment of choice. In two patients, radical parotidectomy was performed, whereas one patient underwent partial parotidectomy. Selective neck dissection was performed in one case. In two cases, post-surgical treatment involved radiation and/or chemotherapy. Two patients died as a consequence of local recurrence within the first year after diagnosis, whereas one patient is alive and free of disease after a follow up of 14 months. CONCLUSION: Our own experiences, taken together with those reported in the literature, suggest that clear surgical margins are probably the most important factor for avoiding a recurrence and to improve disease free survival.

A software framework for solving bioelectrical field problems based on finite elements.

Computational modeling and simulation can provide important insights into the electrical and electrophysiological properties of cells, tissues, and organs. Commonly, the modeling is based on Maxwell's and Poisson's equations for electromagnetic and electric fields, respectively, and numerical techniques are applied for field calculation such as the finite element and finite differences methods. Focus of this work are finite element methods, which are based on an element-wise discretization of the spatial domain. These methods can be classified on the element's geometry, e.g. triangles, tetrahedrons and hexahedrons, and the underlying interpolation functions, e.g. polynomials of various order. Aim of this work is to describe finite element-based approaches and their application to extend the problem-solving environment SCIRun/BioPSE. Finite elements of various types were integrated and methods for interpolation and integration were implemented. General methods for creation of finite element system matrices and boundary conditions were incorporated. The extension provides flexible means for geometric modeling, physical simulation, and visualization with particular application in solving bioelectric field problems.