Inflammatory Cell Interactions Research Articles

Ebselen Reduces Inflammation and Microvascular Perfusion Failure After Blunt Skeletal Muscle Injury of the Rat

Soft tissue trauma induces an local inflammatory response and yields a microvascular perfusion failure due to trauma-induced oxidative stress. Using high-resolution multifluorescence microscopy, we herein report on the efficiency of treatment with the oxygen radical scavenger ebselen to improve compromised perfusion of traumatized muscle tissue and to minimize secondary tissue damage. By using a pneumatically driven computer-controlled impact device, closed soft tissue trauma of the left hind limb was induced in pentobarbital-anesthetized rats that received either ebselen (30 mg/kg body weight, intraperitoneally) or equal volumes of the vehicle dimethyl sulfoxide (DMSO). In an additional series of animals, ebselen or DMSO were applied without soft tissue trauma. Ebselen restored microcirculatory impairment within the injured muscle, as given by values of nutritive perfusion (763 +/- 44 cm/cm2), nicotinamide adenine dinucleotide levels (56 +/- 3 aU) and inflammatory cell interaction (leukocytes: 226 +/- 31 mm(-2)) at 24 hours after trauma, being not different to those found in noninjured muscle tissue of controls. In contrast, skeletal muscle in DMSO-treated animals revealed persistent perfusion failure (564 +/- 32 cm/cm2) with tissue hypoxia (nicotinamide adenine dinucleotide 75 +/- 11 aU) and enhanced endothelial interaction of leukocytes (383 +/- 18 mm(-2)) at 24 hours after trauma. Treatment of skeletal muscle soft tissue trauma with the glutathione peroxidase mimic ebselen is highly effective in restoration of disturbed microcirculation. Moreover, reduced inflammatory cell response helps to prevent leukocyte-dependent secondary tissue injury.

Biocompatibility of porous polyethylene implants tissue‐engineered by extracellular matrix and VEGF

Rapid ingrowth of blood vessels and low inflammatory response are considered major prerequisites for successful implantation of biomaterials in reconstructive surgery. Aim of the present study was to evaluate whether tissue-engineered porous polyethylene (PPE) implants providing extracellular matrix components (ECM) and vascular endothelial growth factor (VEGF) in vivo improve microvascular ingrowth and mechanical integration with regard to initial inflammatory responses. PPE implants (3 x 3 x 0.1 mm(3), pore size approximately 100-200 microm) were tissue-engineered by incorporation of ECM components (GFR-Matrigel) adding recombinant murine VEGF (1 microg/mL) and grafted into dorsal skinfold chamber preparations of C57BL/6 mice. Control animals received uncoated implants or implants coated with ECM components alone (n = 6 per group). Using in vivo fluorescence microscopy angiogenic activity and inflammatory leukocyte-endothelial cell interactions were analyzed for 2weeks. Finally, mechanical integration was quantified by measurement of dynamic desintegration strengths at the host-implant border. Functional vessel density, red blood cell velocity, and vessel diameters increased continuously in all groups indicating that rapid microvascular integration of PPE occurred even without incorporation of ECM or VEGF. However, a transient initial inflammatory response with increased leukocyte-endothelial cell adherence on day 7 in uncoated control implants was efficiently reduced by incorporation of ECM and VEGF. Measurement of dynamic breaking strengths revealed no significant differences between the groups although there was a tendency to improved mechanical integration in tissue-engineered implants. Therefore, novel tissue- engineered constructs of PPE implants providing ECM and VEGF in high local concentrations can increase biocompatibility especially under unfavorable conditions for implantation.

Initiation and Progression of Axonopathy in Experimental Autoimmune Encephalomyelitis

Axonal loss is the principal cause of chronic disability in multiple sclerosis and experimental autoimmune encephalomyelitis (EAE). In C57BL/6 mice with EAE induced by immunization with myelin oligodendrocyte glycoprotein peptide 35-55, the first evidences of axonal damage in spinal cord were in acute subpial and perivascular foci of infiltrating neutrophils and lymphocytes and included intra-axonal accumulations of the endovesicular Toll-like receptor TLR8, and the inflammasome protein NAcht leucine-rich repeat protein 1 (NALP1). Later in the course of this illness, focal inflammatory infiltrates disappeared from the spinal cord, but there was persistent activation of spinal cord innate immunity and progressive, bilaterally symmetric loss of small-diameter corticospinal tract axons. These results support the hypothesis that both contact-dependent and paracrine interactions of systemic inflammatory cells with axons and an innate immune-mediated neurodegenerative process contribute to axonal loss in this multiple sclerosis model.

Atherosclerosis

Atherosclerosis is a very complex pathology. Over the past two decades we have come to appreciate that a major component of this pathological process is chronic inflammation. This inflammatory state is not only characterized by infiltration of lipid laden macrophages, the hallmark cell of this disease, into the vascular wall, but also is characterized by the infiltration of a host of other inflammatory cells. The complex interaction of inflammatory cells with the normal residents of the vascular wall (ie, endothelial and smooth muscle cells) directs the progression of the disease. Inflammatory cells secrete a variety of chemokines and cytokines which have a profound effect on the development and progression of the plaque. Moreover, many of these cytokines can mediate both pro- and antiatherogenic processes depending on the cellular milieu. The tumor necrosis factor (TNF) and TNF receptor (TNFR) superfamilies encompass numerous cytokines and receptors suggested to play a pivotal role in atherogenesis. Although many members of these families such as TNF-α and CD40 ligand and their putative receptors are considered to be proatherogenic,1–4 studies have also demonstrated an antiatherogenic role for others, including the p55 TNFR.5 TNF-like weak inducer of apoptosis (TWEAK) and its putative receptor, fibroblast growth factor-inducible 14 (Fn14), are also members of the TNF and TNFR superfamilies, respectively. Both TWEAK and Fn14 have been detected in human atherosclerotic plaques, suggesting that these molecules may also play a role in the atherogenic process. Clinical …

Functional profiling in stage IV NSCLC: A phase II trial of individualized therapy

e19079 Background: The introduction of new classes of agents has improved survival in NSCLC yet the accurate selection of candidates for these often costly & toxic drugs remains a challenge. Genomic platforms have identified several prognostic & predictive factors e.g. ERCC1, RRM1/2, EGFr mutations but the comparatively small number of known cellular analytes has limited their broad application. Methods: To address the complexity and redundancy of cell-death signaling pathways, we applied Ex Vivo Analysis of Programmed Cell Death (EVA/PCD) (Nagourney, R. Curr Treat Options in Oncol. 2006) to select therapies for chemo- naïve pts with metastatic NSCLC. This functional platform uses morphologic and metabolic endpoints to gauge cellular response to drugs & signal transduction inhibitors in human tumor microspheroids isolated from surgical biopsies. Modified Z-scores & synergy analyses (median-effect) were used to identify the ex vivo best regimen (EVBR) for each pt. from FDA-approved NSCLC drugs. All pts signed informed consent. Results: 22/25 (88%) pts are evaluable with 3/25 (12%) excluded, 1 for CVA & 2 inadequate PFS at time of treatment. There were CR = 2/22 (9%); PR = 10/22 (45%); SD = 9/22 (41%) & PD = 1/22 (5%) for an ORR = 12/22 (54%) & CBR = 21/22 (95%). 1st line EVBR were CDDP/Gemcitabine 10/25 (40%); CDDP/Taxane 7/25(28%); Erlotinib 5/25(20%); CDDP/Vinorelbine 1/25 (4%); CDDP/Irinotecan1/25(4%) & Ifex/Gemcitabine1/25(4%). Six of 22 (27%) stage IV pts were converted to surgical and/or radiation candidates by therapy. Accrual continues with current TTP ranges from 1.2–35+ months and OS from 1.7 to 46+ months. Conclusions: EVA/PCD ORR in Stage IV NSCLC of 54%, many durable, compares very favorably with reports in this population. By examining drug-induced cell death events in native-state microspheroids, this platform has the unique capacity to capture stromal, vascular and inflammatory cell interactions with tumor cells, now known to be crucial for clinical response prediction. EVA/PCD analysis in NSCLC warrants further evaluation. Supported by the Memorial Medical Center Foundation of Long Beach. [Table: see text]

Functional profiling in stage IV colorectal cancer: A phase II trial of individualized therapy

e15124 Background: The introduction of new classes of agents has improved survival in colorectal cancer (CRC) yet the accurate selection of candidates for these often costly & toxic drugs remains a challenge. Genomic platforms have identified several prognostic & predictive factors e.g. ERCC1, TS, kRas mutations but the comparatively small number of known cellular analytes has limited their broad application. Methods: To address the complexity and redundancy of cell-death signaling pathways, we applied Ex Vivo Analysis of Programmed Cell Death (EVA/PCD) (Nagourney, R. Curr Treat Options in Oncol. 2006) to select therapies for chemo-naïve pts with metastatic CRC. This functional platform uses morphologic and metabolic endpoints to gauge cellular response to drugs & signal transduction inhibitors in human tumor microspheroids isolated from surgical biopsies. Modified Z-scores & synergy analyses (median-effect) were used to identify the ex vivo best regimen (EVBR) for each pt. from FDA-approved CRC drugs. All pts signed informed consent. Results: 12/13 (92%) pts are evaluable. Of the 12 there was 1/12 (8%) CR; 7/12 (58%) PR; 3/12 (25%) SD & 1/12 (8%) PD for an ORR = 8/12 (66%) & CBR = 11/12 (92%). 1st line EVBR were FOLFOX 3/12 (25%); FOLFIRI 2/12 (16%); FOLFOX/Erbitux 2/12(16%); FOLFOX/Avastin 1/12 (8%); FOLFOX/Avastin/Erbitux 1/12(8%); Irinotecan/Avastin/Erbitux 2/12(16%); IROX/Avastin 1/12(8%). Three of 12 inoperable pts (25%) converted to surgical candidates. There was 1 treatment-related death. Accrual continues with current TTP ranges from 1.2–17.3 months and OS from 2.5 to 30 months. Conclusions: EVA/PCD ORR in Stage IV CRC of 66%, many durable, compares favorably with reports in this population. By examining drug-induced cell death events in native-state microspheroids, this platform has the unique capacity to capture stromal, vascular and inflammatory cell interactions with tumor cells, now known to be crucial for clinical response prediction. EVA/PCD analysis warrants further evaluation in CRC. Supported by the Memorial Medical Center Foundation of Long Beach. [Table: see text]

Involvement of Müller glial cells in epiretinal membrane formation

Proliferative retinopathies are considered to represent maladapted retinal wound repair processes driven by growth factor- and cytokine-induced overstimulation of proliferation, migration, extracellular matrix production and contraction of retinal cells. The formation of neovascular membranes represents an attempt to reoxygenize non-perfused retinal areas. Müller glial cells play a crucial role in the pathogenesis of proliferative retinopathies. This review summarizes the present knowledge regarding the role of Müller cells in periretinal membrane formation, especially in the early steps of epiretinal membrane formation, which involve an interaction of inflammatory and glial cells, and gives a survey of the factors which are suggested to be implicated in the induction of Müller cell gliosis and proliferation. Alterations in the membrane conductance of Müller cells suggest that Müller cells may alter their phenotype into progenitor-like cells in the course of proliferative retinopathies; transdifferentiated Müller cells may have great impact for the development of new cell-based therapies.

Reduced responses of macrophages on nanometer surface features of altered alumina crystalline phases

Extensive prolonged interactions of inflammatory cells (such as macrophages) at the host–implant interface may lead to implant failure. While previous studies have shown increased in vitro and in vivo bone cell adhesion, proliferation and mineralization on nanophase compared to currently implanted ceramics, few studies have been conducted to elucidate inflammatory cell responses on such nanophase ceramics. Controlling surface feature size and corresponding surface roughness on implants may clearly alter immune cell responses, which would be an extremely important consideration for the use of nanostructured materials as improved biomaterials. In this study, reduced macrophage density was observed on alumina (Al 2O 3) compacts with greater nanometer surface roughness accompanied by changes in crystallinity for up to 24 h in culture. Since alumina is a commonly used ceramic in orthopedic applications, this in vitro study continues to support the use of nanophase ceramics as improved orthopedic implants by demonstrating reduced macrophage responses.

Erythropoietin Protects Critically Perfused Flap Tissue

The objective of this study was to analyze whether erythropoietin (EPO) protects from necrosis of critically perfused musculocutaneous tissue and the mechanisms by which this protection is achieved. EPO is the regulator of erythropoiesis and is used to treat patients with anemia of different causes. Recent studies suggest that EPO has also other tissue-protective effects, irrespective of its erythropoietic properties. C57BL/6-mice were treated with 3 doses of EPO at 500 IU/kg intraperitoneally. EPO was given either before (preconditioning, n = 7), before and after (overlapping treatment, n = 7), or after (treatment, n = 7) surgery. Animals receiving only saline served as controls (CON). Acute persistent ischemia was induced by elevating a randomly perfused flap in the back of the animals. This critically perfused tissue demonstrates an initial microvascular failure of approximately 40%, resulting in approximately 50% tissue necrosis if kept untreated. Repetitive fluorescence microscopy was performed over 10 days, assessing angiogenesis, functional capillary density, inflammatory leukocyte-endothelial cell interaction, apoptotic cell death, and tissue necrosis. Additional molecular tissue analyses included the determination of inducible nitric oxide synthase, erythropoietin receptor (EPO-R), and vascular endothelial growth factor (VEGF). EPO preconditioning did not affect hematocrit and EPO-R expression, but increased inducible nitric oxide synthase in the critically perfused tissue. This correlated with a significant arteriolar dilation, which resulted in a maintained functional capillary density (CON: 0 +/- 0 cm/cm(2); preconditioning: 37 +/- 21 cm/cm(2); overlapping treatment: 72 +/- 26 cm/cm(2); P < 0.05). EPO pretreatment further significantly reduced microvascular leukocyte adhesion and apoptotic cell death. Moreover, EPO pretreatment induced an early VEGF upregulation, which resulted in new capillary network formation (CON: 0 +/- 0 cm/cm(2); preconditioning: 40 +/- 3 cm/cm(2); overlapping treatment: 33 +/- 3 cm/cm(2); P < 0.05). Accordingly, EPO pretreatment significantly reduced tissue necrosis (CON: 48% +/- 2%; preconditioning: 26% +/- 3%; overlapping treatment: 20% +/- 3%; P < 0.05). Of interest, EPO treatment was only able to alleviate ischemia-induced inflammation but could not improve microvascular perfusion and tissue survival. EPO pretreatment improves survival of critically perfused tissue by nitric oxide -mediated arteriolar dilation, protection of capillary perfusion, and VEGF-initiated new blood vessel formation.

Targeted Liposomal Paclitaxel Raises Tumor Vessel Leakiness

Problem Paclitaxel encapsulated in cationic liposomes (EndoTAG-1) is a vascular targeting formulation for the treatment of solid tumors. It triggers intratumoral microthrombosis causing significant inhibition of tumor perfusion and tumor growth associated with endothelial cell apoptosis. Here, we quantified the effects of repeated EndoTAG-1 therapy on tumor microvascular leakiness with respect to leukocyte-endothelial cell interactions, the targeting property of cationic liposomes and the therapeutic combination with conventional cisplatin chemotherapy. Methods Using dorsal skinfold chamber preparations in Syrian Golden hamsters in vivo fluorescence microscopy experiments were performed after repeated EndoTAG-1 treatment of A-Mel-3 tumors. Controls received glucose, paclitaxel alone or cationic liposomes devoid of paclitaxel. Extravasation of rhodamine-labelled albumin was measured to calculate microvessel permeability and intratumoral leukocyte-endothelial cell interactions were quantified. Subcutaneous tumor growth was evaluated after combination therapy followed by histological analysis. Results Microvascular permeability was significantly increased only after treatment with EndoTAG-1, while intratumoral leukocyte-endothelial cell interactions were not affected by any treatment. In separate skinfold chamber experiments fluorescently-labeled cationic liposomes kept their targeting property for tumor endothelial cells after repeated EndoTAG-1 treatment and no signs of extravasation were observed. Subcutaneous A-Mel-3 tumor growth was significantly inhibited by the combination of cisplatin and EndoTAG-1. Conclusion These data show that vascular targeting with EndoTAG-1 increases tumor microvessel leakiness probably due to vascular damage. This mechanism is not mediated by inflammatory leukocyte-endothelial cell interactions. Manipulating the blood-tumor barrier by repeated tumor microvessel targeting using EndoTAG-1 can effectively be combined with tumor cell-directed conventional cisplatin chemotherapy. Significance Antivascular therapy using liposomal paclitaxel (EndoTAG-1) is currently undergoing Phase II clinical studies in human cancer patients. Referring to the present experimental data showing increased permeability of intratumoral microvessels combination therapy with conventional therapy appears to be a very promising strategy. Support This study was supported by grants of Munich Biotech AG, Neuried, Germany and by grants of the Novartis Foundation for Therapeutic Research, Nürnberg, Germany.

Paclitaxel Encapsulated in Cationic Liposomes Increases Tumor Microvessel Leakiness and Improves Therapeutic Efficacy in Combination with Cisplatin

Paclitaxel encapsulated in cationic liposomes (EndoTAG-1) is a vascular targeting formulation for the treatment of solid tumors. It triggers intratumoral microthrombosis, causing significant inhibition of tumor perfusion and tumor growth associated with endothelial cell apoptosis. Here, we quantified the effects of repeated EndoTAG-1 therapy on tumor microvascular leakiness with respect to leukocyte-endothelial cell interactions, the targeting property of cationic liposomes, and the therapeutic combination with conventional cisplatin chemotherapy. Using dorsal skinfold chamber preparations in Syrian Golden hamsters, in vivo fluorescence microscopy experiments were done after repeated EndoTAG-1 treatment of A-Mel-3 tumors. Controls received glucose, paclitaxel alone, or cationic liposomes devoid of paclitaxel. Extravasation of rhodamine-labeled albumin was measured to calculate microvessel permeability, and intratumoral leukocyte-endothelial cell interactions were quantified. Subcutaneous tumor growth was evaluated after combination therapy followed by histologic analysis. Microvascular permeability was significantly increased only after treatment with EndoTAG-1, whereas intratumoral leukocyte-endothelial cell interactions were not affected by any treatment. In separate skinfold chamber experiments, fluorescently labeled cationic liposomes kept their targeting property for tumor endothelial cells after repeated EndoTAG-1 treatment and no signs of extravasation were observed. Subcutaneous A-Mel-3 tumor growth was significantly inhibited by the combination of cisplatin and EndoTAG-1. These data show that vascular targeting with EndoTAG-1 increases tumor microvessel leakiness probably due to vascular damage. This mechanism is not mediated by inflammatory leukocyte-endothelial cell interactions. Manipulating the blood-tumor barrier by repeated tumor microvessel targeting using EndoTAG-1 can effectively be combined with tumor cell-directed conventional cisplatin chemotherapy.

Intercellular adhesion molecule-1 (ICAM-1) expression and soluble ICAM-1 (sICAM-1) production by cytokine-activated human aortic endothelial cells: a possible role for ICAM-1 and sICAM-1 in atherosclerotic aortic aneurysms.

The interactions of inflammatory cells, cytokines, and cell adhesion molecules (CAM) may be important in the pathogenesis of vascular diseases such as abdominal aortic aneurysms (AAA), in which inflammation plays a role. The aim of this study was to investigate the pathogenic role of ICAM-1, a molecule involved in leucocyte-endothelial interactions, in vascular inflammation. ELISA of human explant culture supernatants revealed a four-fold increase in sICAM-1 production by AAA (n = 9) versus normal (n = 8) aortic explants. Human aortic endothelial cell (hAEC) culture was used for further studies as an in vitro model for aortic inflammatory conditions. Tumour necrosis factor-alpha (TNF-alpha) or IL-1 beta treatment of hAEC resulted in an up to 1.8-fold significant increase in sICAM-1 production compared with resting cells. In addition, the expression of ICAM-1 on cytokine-stimulated versus resting hAEC was measured by radioimmunoassay. TNF-alpha significantly induced ICAM-1 expression on these cells. These results suggest that different forms of ICAM-1, present on or released by the activated aortic endothelium, may be involved in leucocyte adhesion to and migration into the vessel wall.

CD13 is a novel mediator of monocytic/endothelial cell adhesion

During inflammation, cell surface adhesion molecules guide the adhesion and migration of circulating leukocytes across the endothelial cells lining the blood vessels to access the site of injury. The transmembrane molecule CD13 is expressed on monocytes and endothelial cells and has been shown to mediate homotypic cell adhesion, which may imply a role for CD13 in inflammatory monocyte trafficking. Here, we show that ligation and clustering of CD13 by mAb or viral ligands potently induce myeloid cell/endothelial adhesion in a signal transduction‐dependent manner involving monocytic cytoskeletal rearrangement and filopodia formation. Treatment with soluble recombinant (r)CD13 blocks this CD13‐dependent adhesion, and CD13 molecules from monocytic and endothelial cells are present in the same immunocomplex, suggesting a direct participation of CD13 in the adhesive interaction. This concept is strengthened by the fact that activated monocytic cells adhere to immobilized recombinant CD13. Furthermore, treatment with anti‐CD13 antibodies in a murine model of peritonitis results in a decrease in leukocyte infiltration into the peritoneum, suggesting a potential role for CD13 in leukocyte trafficking in vivo. Therefore, this work supports a new direction for CD13 biology, where these cell surface molecules act as true molecular interfaces that induce and participate in critical inflammatory cell interactions.

Antral Inflammatory Cells, Gastric Emptying, and Electrogastrography in Pediatric Functional Dyspepsia

The aims of the current study were to determine the activation states of antral eosinophils and mast cells and to evaluate the interactions of antral inflammatory cells with gastric emptying and electrogastrography (EGG) in 30 pediatric patients with functional dyspepsia. Eosinophil degranulation was moderate in 42% and extensive in 54% of patients. Mast cell degranulation was > 50% in 81% of patients. Elevated mast cell density was associated with slower one hour gastric emptying and increased preprandial dysrhythmia. Mast cell density correlated with the preprandial percentage tachygastria. CD3+ cell density correlated with the preprandial percentage tachygastria also, but only in patients with increased eosinophil density. In conclusion, antral eosinophils and mast cells are significantly activated in pediatric functional dyspepsia. Mast cell density is associated with delayed gastric emptying and preprandial dysrhythmia, suggesting that there may be an interaction between antral inflammation and gastric electromechanical dysfunction in the pathophysiology of pediatric functional dyspepsia.

Role of thrombin in interleukin-5 expression from basophils

Interleukin-5 (IL-5) plays a key role in the pathogenesis of bronchial asthma. Thrombin is a procoagulant factor that has been also reported to participate in the inflammatory response by stimulating the secretion of cytokines. Interaction of inflammatory cells with airway epithelial cells may also promote the secretion of cytokines. However, the role of thrombin and cell-to-cell interaction in pathogenesis of allergic inflammation is unclear. In this study, we evaluated the role of thrombin and cell-to-cell interaction in the secretion of IL-5 from basophils. The human basophil cell line KU-812 was used in the assays. Thrombin and co-culture with alveolar epithelial cells significantly stimulated the secretion of IL-5 from KU-812 cells as compared to controls. Secretion of IL-5 was synergistically stimulated when KU-812 cells were incubated in the presence of both thrombin and alveolar epithelial cells. Co-culture of KU-812 cells with epithelial cells significantly increased the expression of tissue factor, an activator of coagulation activation, in a cell dose-dependent manner. Secretion of IL-5 from KU-812 basophils co-cultured with epithelial cells was significantly inhibited by LY294002, an inhibitor of phosphatidylinositol 3-kinase. These results suggest that thrombin and cell interaction with lung epithelial cells may augment the inflammatory response in allergic diseases by stimulating the secretion of IL-5 from basophils.

Vascularization and biocompatibility of scaffolds consisting of different calcium phosphate compounds

Scaffolds for tissue engineering of bone should mimic bone matrix and promote vascular ingrowth. Whether synthetic hydroxyapatite and acellular dentin, both materials composed from calcium phosphate, fulfill these material properties has not been studied yet. Therefore, we herein studied in vivo the host angiogenic and inflammatory response to these biomaterials. Porous scaffolds of hydroxyapatite and isogeneic acellular dentin were implanted into the dorsal skinfold chamber of balb/c mice. Additional animals received perforated implants of isogeneic calvarial bone displaying pores similar in size and structure to those of both scaffolds. Chambers of animals without implants served as controls. Angiogenesis and neovascularization as well as inflammatory leukocyte-endothelial cell interaction and microvascular leakage were analyzed over a 14-day time period using intravital fluorescence microscopy. Implantation of both hydroxyapatite and dentin scaffolds showed a slight increase in leukocyte recruitment compared with controls. This was associated with an elevation of microvascular permeability, which was comparable to that observed in response to isogeneic bone. In addition, hydroxyapatite as well as dentin scaffolds induced a marked angiogenic response, which resulted in complete vascularization of the implants until day 14. Of interest, in hydroxyapatite scaffolds, the newly formed capillaries were not as densely meshed as in dentin scaffolds, in which the functional capillary density was comparable to that measured in bone implants. Hydroxyapatite and, in particular, dentin scaffolds promote vascularization and exhibit a biocompatibility comparable to that of isogeneic bone. This may guarantee the rapid incorporation of these materials into the host tissue.

Aortic valve stenosis: an active atheroinflammatory process

To summarize the current understanding of the pathobiology of aortic valve stenosis and portray the major advances in this field. Stenotic aortic valves are characterized by atherosclerosis-like lesions, consisting of activated inflammatory cells, including T lymphocytes, macrophages, and mast cells, and of lipid deposits, calcific nodules, and bone tissue. Active mediators of calcification and cells with osteoblast-like activity are present in diseased valves. Extracellular matrix remodeling, including collagen synthesis and elastin degradation by matrix metalloproteinases and cathepsins, contributes to leaflet stiffening. In experimental animals, hypercholesterolemia induces calcification and bone formation in aortic valves, which can be inhibited by statin treatment. The potential of statins to retard progression of aortic valve stenosis has also been recognized in clinical studies; however, further prospective trials are needed. Angiotensin II-forming enzymes are upregulated in stenotic valves. Angiotensin II may participate in profibrotic progression of aortic valve stenosis and may serve as a possible therapeutic target. Recent findings regarding the interaction of inflammatory cells, lipids, mediators of calcification, and renin-angiotensin system in stenotic valves support the current opinion of aortic valve stenosis being an actively regulated disease, potentially amenable to targeted molecular therapy. Evidence from prospective clinical studies is eagerly awaited.

Physiological 3D tissue model of the airway wall and mucosa

This protocol describes the setup, maintenance and characteristics of a tissue-engineered model of the human bronchial mucosa that can be used for basic physiology and pathophysiology studies. The model includes a well-differentiated epithelium with functional cilia, mucus secretion and subepithelial fibroblasts within type I collagen. The tissue is created within porous polymeric wells to prevent gel contraction and allow culture at the air-liquid interface. It requires at least 2 wk to be established and can be maintained thereafter for over 4 wk, with tissue differentiation moving towards a more physiologically relevant phenotype with increasing time in culture. Over time, the extracellular matrix also remodels, depositing proteins such as types III and IV collagen and fibronectin. Because it recapitulates many key anatomical and functional features of the airway wall, this model is well suited for a wide range of studies, including those on airway remodeling, transepithelial transport and inflammatory cell interactions with the mucosa. The entire protocol takes 4-6 wk, including cell expansion, depending on the extent of ciliogenesis desired.

In vitro Evaluation of Macrophage Adhesion and Proliferation on Alumina

ABSTRACTExtensive interactions of inflammatory cells (such as macrophages) with biomaterials at the host-implant interface are often blamed for failure of implanted biomedical devices [1]. While previous studies have shown increased in vitro and in vivo bone cell (osteoblast) responses on nanophase ceramics [2], few (if any) studies have been conducted to elucidate inflammatory cell responses on such novel materials. In this study, we reported that macrophage adhesion and proliferation on nanophase (97.7 nm grain size) alumina (Al2O3) was significantly less than conventional (187.4 nm grain size) alumina, respectively, after 4, 12, 24 h. The present study provides evidence of the ability of nanophase alumina to down-regulate macrophage adhesion and proliferation, which is imperative for the future consideration of nanophase materials for orthopedic and dental applications.

Human Nasal Polyp Microenvironment Maintained in Viable and Functional States as Xenografts in SCID Mice

We undertook to maintain human nasal polyp tissue in a viable and functional state in SCID (severe combined immunodeficiency) mice. Small, nondisrupted pieces of human nasal polyp tissues were subcutaneously implanted into SCID mice depleted of natural killer cells. The resulting xenografts were examined histologically, and the sera were evaluated for the presence of human protein. The original histologic architecture of the polyp was maintained in the xenografts. The tissues, including pseudostratified columnar epithelial-lined polyps and subepithelial stroma, remained viable, and goblet cells continued to produce mucin for up to 26 weeks after engraftment. Human inflammatory leukocytes, including CD3+ T cells, CD20+ B cells, CD138+ plasma cells, and CD68+ monocytes and/or macrophages, were present. Identification of human immunoglobulin and human interferon-gamma in the sera of xenograft-bearing mice indicated that the B cells or plasma cells and T cells within the xenografts remained functional for 2 weeks after engraftment. The ability to engraft and maintain nasal polyps provides an in vivo human/mouse chimeric model with which to investigate the role of inflammatory leukocytes and stromal cells in the maintenance and progression of polyposis and to determine how exogenous cytokines may alter the interaction of inflammatory cells, stromal cells, and epithelial cells in the polyp.