Synthesis of 1H-isoindolin-1-ones via a simple photodecarboxylative addition of carboxylates to phthalimides and evaluation of their antibiotic activity.

A variety of 3-hydroxy-isoindolin-1-one derivatives were synthesized using the photodecarboxylative addition of carboxylates to phthalimide derivatives in aqueous media. Subsequent acid-catalyzed dehydration furnished 3-(alkyl and aryl)methyleneisoindolin-1-ones with variable E-diastereoselectivity in good to excellent overall yields. Noteworthy, the parent 3-phenylmethyleneisoindolin-1-one underwent isomerization and oxidative decomposition when exposed to light and air. Selected 3-hydroxy-isoindolin-1-one and 3-(alkyl and aryl)methyleneisoindolin-1-one derivatives showed moderate antibacterial activity that justifies future elaboration and study of these important bioactive scaffolds.

Sex-Specific Relationship Between Parathyroid Hormone and Platelet Indices in Phenotypes of Heart Failure-Results From the MyoVasc Study.

Background: Heart failure (HF) is a multifactorial syndrome with pathophysiological complexities still not fully understood. Higher mean platelet volume (MPV), a potential marker of platelet activation, and high concentrations of parathyroid hormone (PTH) have been implicated in the pathogenesis of HF. Aim: This study aims to investigate sex-specifically the association between PTH concentrations and platelet indices in phenotypes of HF. Methods and Results: PTH and platelet indices (MPV and platelet count) were available in 1,896 participants from the MyoVasc study in Mainz, Germany. Multivariable linear regression models, adjusted for age, sex, season, vitamin D status, cardiovascular risk factors, comorbidities, estimated glomerular filtration rate, and medication, were used to assess the associations between platelet indices and PTH. The results showed distinct sex-specific associations between PTH and platelet indices. A positive association between PTH and MPV was found in females with symptomatic HF with reduced ejection fraction (HFrEF) only [ß = 0.60 (0.19; 1.00)]. Platelet count was inversely associated with PTH in male HFrEF individuals [ß = -7.6 (-15; -0.30)] and in both males and females with HF with preserved ejection fraction (HFpEF). Conclusion: This study reports differential, sex-specific relationships between PTH and platelet indices in HF individuals independent of vitamin D status and clinical profile. Particularly in phenotypes of symptomatic HF, distinct associations were observed, suggesting a sex-specific mechanism involved in the interaction between PTH and platelets.

In Vitro Entero-Capillary Barrier Exhibits Altered Inflammatory and Exosomal Communication Pattern after Exposure to Silica Nanoparticles.

The intestinal microvasculature (iMV) plays multiple pathogenic roles during chronic inflammatory bowel disease (IBD). The iMV acts as a second line of defense and is, among other factors, crucial for the innate immunity in the gut. It is also the therapeutic location in IBD targeting aggravated leukocyte adhesion processes involving ICAM-1 and E-selectin. Specific targeting is stressed via nanoparticulate drug vehicles. Evaluating the iMV in enterocyte barrier models in vitro could shed light on inflammation and barrier-integrity processes during IBD. Therefore, we generated a barrier model by combining the enterocyte cell line Caco-2 with the microvascular endothelial cell line ISO-HAS-1 on opposite sides of a transwell filter-membrane under culture conditions which mimicked the physiological and inflamed conditions of IBD. The IBD model achieved a significant barrier-disruption, demonstrated via transepithelial-electrical resistance (TER), permeability-coefficient (Papp) and increase of sICAM sE-selectin and IL-8. In addition, the impact of a prospective model drug-vehicle (silica nanoparticles, aSNP) on ongoing inflammation was examined. A decrease of sICAM/sE-selectin was observed after aSNP-exposure to the inflamed endothelium. These findings correlated with a decreased secretion of ICAM/E-selectin bearing exosomes/microvesicles, as evaluated via ELISA. Our findings indicate that aSNP treatment of the inflamed endothelium during IBD may hamper exosomal/microvesicular systemic communication.

Mean Platelet Volume and Arterial Stiffness - Clinical Relationship and Common Genetic Variability.

Vessel wall stiffening is an important clinical parameter, but it is unknown whether platelets, key elements in the pathogenesis of arterial thrombosis, are associated with arterial stiffness. The present studies sought to determine whether mean platelet volume (MPV), a potential marker of platelet activation, is linked to vascular elasticity as assessed by the augmentation index (AIx), in 15,010 individuals from the population-based Gutenberg Health Study. Multivariable analysis showed that MPV in both males (ß 0.776; 95thCI [0.250;1.16]; p = 0.0024) and females (ß 0.881[0.328;1.43]; p = 0.0018) is strongly associated with AIx. Individuals with MPV and AIx above the sex-specific medians had worse survival. Association analysis between MPV-related genetic variants and arterial stiffness identified four genetic variants in males and one in females related with AIx. Cox regression analysis for mortality identified one of these joint genetic variants close to ring finger protein 145 gene (RNF145, rs10076782) linked with increased mortality (hazard ratio 2.02; 95thCI [1.35;3.02]; p = 0.00061). Thus, these population-based data demonstrate a close relation between platelet volume as a potential marker of platelet activation and arterial stiffness in both sexes. Further research is warranted to further elucidate the mechanisms underlying larger platelets' role in arterial stiffening including the role of shared common genetics.

Sex-specific differences in genetic and nongenetic determinants of mean platelet volume: results from the Gutenberg Health Study.

Mean platelet volume (MPV), a measure of platelet size, is a potential biological marker of platelet function. To date, a comprehensive analysis including known genetic and nongenetic factors that determine MPV is still lacking. MPV has been evaluated in 15 010 individuals from the population-based Gutenberg Health Study. Genetic information was available for 4175 individuals. Our results showed that age (ß, 0.0346; 95% confidence interval [CI], 0.0255 to 0.0436), cardiovascular risk factors (CVRFs) such as smoking (ß, 0.178; 95% CI, 0.128 to 0.229), hypertension (ß, 0.05; 95% CI, 0.00289 to .0981), and high glucose level (ß, 0.00179; 95% CI, 0.0006 to 0.00299) were linked with higher MPV in males only. Intake of oral contraceptives (ß, 0.150; 95% CI, 0.0649 to 0.236) and menstruation (ß, 0.123; 95% CI, 0.0231 to 0.224) were strongly associated with higher MPV in females. Seven single nucleotide polymorphisms (SNPs) for females and 4 SNPs for males were associated with higher MPV. The full model, including age, CVRFs, laboratory parameters, medications, and genetic variation, explained 20.4% of the MPV variance in females and 18.6% in males. The curves of cumulative mortality, stratified for sex, showed worse survival for males only with MPV > 9.96 fL vs MPV ≤ 9.96 fL (P < .0001). This study provides evidence for heterogeneity in the profile of determinants for MPV between sexes. The observed interactions between genetic variability, CVRFs, and MPV and its association with the development of cardiovascular disease or thrombotic risk need to be further investigated.

Distribution, genetic and cardiovascular determinants of FVIII:c - Data from the population-based Gutenberg Health Study.

BACKGROUND: Elevated levels of FVIII: c are associated with risk for both venous and arterial thromboembolism. However, no population-based study on the sex-specific distribution and reference ranges of plasma FVIII: c and its cardiovascular determinants is available. FVIII: c was analyzed in a randomly selected sample of 2533 males and 2440 females from the Gutenberg Health Study in Germany. Multivariable regression analyses for FVIII: c were performed under adjustment for genetic determinants, cardiovascular risk factors and cardiovascular disease. RESULTS AND CONCLUSIONS: Females (126.6% (95% CI: 125.2/128)) showed higher FVIII: c levels than males (121.2% (119.8/122.7)). FVIII: c levels increased with age in both sexes (ß per decade: 5.67% (4.22/7.13) male, 6.15% (4.72/7.57) female; p<0.001). Sex-specific reference limits and categories indicating the grade of deviation from the reference were calculated, and nomograms for FVIII: c were created. FVIII: c was approximately 25% higher in individuals with non-O blood type. Adjusted for sex and age, ABO-blood group accounted for 18.3% of FVIII: c variation. In multivariable analysis, FVIII: c was notably positively associated with diabetes mellitus, obesity, hypertension and dyslipidemia and negatively with current smoking. In a fully adjusted multivariable model, the strongest associations observed were of elevated FVIII: c with diabetes and peripheral artery disease in both sexes and with obesity in males. Effects of SNPs in the vWF, STAB2 and SCARA5 gene were stronger in females than in males. The use of nomograms for valuation of FVIII: c might be useful to identify high-risk cohorts for thromboembolism. Additionally, the prospective evaluation of FVIII: c as a risk predictor becomes feasible.

Vascular endothelial cadherin expression in lung specimens of patients with sepsis-induced acute respiratory distress syndrome and endothelial cell cultures.

AIMS: Vascular endothelial (VE) cadherin is a cell adhesion molecule localized at endothelial cell (EC) junctions. As a major component of endothelial adherens junctions, its main function is the maintenance and regulation of EC integrity. In the acute respiratory distress syndrome (ARDS), increased vascular permeability is a major mechanism in pulmonary edema and lung dysfunction. In this study, VE-cadherin expression was investigated in ARDS lungs and control tissue as well as in an ARDS cell culture model. METHODS: Lung specimens of patients with ARDS due to Gram-negative sepsis (n = 20; control lung tissue: n = 41) and cell cultures of human pulmonary microvascular ECs and human umbilical vein ECs stimulated with LPS, TNF-α and IFN-γ were stained with a VE-cadherin antibody. Staining intensity was semiquantitatively evaluated by conventional light and immunofluorescence microscopy. RESULTS: VE-cadherin expression was statistically significantly reduced in the endothelium of all vessel types in ARDS lungs compared to control tissue. Cell cultures showing disrupted cellular borders confirmed these results. CONCLUSION: Reduced expression of VE-cadherin has to be considered as a major mechanism of increased vessel permeability in ARDS. The previously described vessel-type-specific expression pattern of VE-cadherin in the human lung is not influenced by ARDS.

Hyperbranched PEG by random copolymerization of ethylene oxide and glycidol.

The synthesis of hyperbranched poly(ethylene glycol) (hbPEG) in one step was realized by random copolymerization of ethylene oxide and glycidol, leading to a biocompatible, amorphous material with multiple hydroxyl functionalities. A series of copolymers with moderate polydispersity ($\overline {M} _{{\rm w}} /\overline {M} _{{\rm n}} $ < 1.8) was obtained with varying glycidol content (3-40 mol-%) and molecular weights up to 49 800 g mol(-1) . The randomly branched structure of the copolymers was confirmed by (1) H and (13) C NMR spectroscopy and thermal analysis (differential scanning calorimetry). MTS assay demonstrated low cell toxicity of the hyperbranched PEG, comparable to the highly established linear PEG.

Acute morphological and toxicological effects in a human bronchial coculture model after sulfur mustard exposure.

Sulfur mustard (SM) is a strong alkylating agent. Inhalation of SM causes acute lung injury accompanied by severe disruption of the airway barrier. In our study, we tested the acute effects after mustard exposure in an in vitro coculture bronchial model of the proximal barrier. To achieve this, we seeded normal human bronchial epithelial explant-outgrowth cells (HBEC) together with lung fibroblasts as a bilayer on filter plates and exposed the bronchial model after 31 days of differentiation to various concentrations of SM (30, 100, 300, and 500 microM). The HBEC formed confluent layers, expressing functional tight junctions as measured by transepithelial electrical resistance (TER). Mucus production and cilia formation reappeared in the coculture model. TER was measured after 2 and 24 h following treatment. Depending on the different concentrations, TER decreased in the first 2 h up to 55% of the control at the highest concentration. After 24 h, TER seemed to recover because at concentrations up to 300 microM values were equal to the control. SM induced a widening of intercellular spaces and a loss in cell-matrix adhesion. Mucus production increased with the result that cilia ceased to beat. Changes in the proinflammatory cytokines interleukin (IL)-6 and IL-8 were also observed. Apoptotic markers such as cytochrome c, p53, Fas-associated protein with death domain, and procaspase-3 were significantly induced at concentrations of less than 100 microM. In summary, SM induces morphological and biochemical changes that reflect pathological effects of SM injury in vivo. It is hoped to use this coculture model to understand further the pathogenesis of SM-induced barrier injury and to search for novel approaches in SM therapy.

Gold nanoparticles induce cytotoxicity in the alveolar type-II cell lines A549 and NCIH441.

BACKGROUND: During the last years engineered nanoparticles (NPs) have been extensively used in different technologies and consequently many questions have arisen about the risk and the impact on human health following exposure to nanoparticles. Nevertheless, at present knowledge about the cytotoxicity induced by NPs is still largely incomplete. In this context, we have investigated the cytotoxicity induced by gold nanoparticles (AuNPs), which differed in size and purification grade (presence or absence of sodium citrate residues on the particle surface) in vitro, in the human alveolar type-II (ATII)-like cell lines A549 and NCIH441. RESULTS: We found that the presence of sodium citrate residues on AuNPs impaired the viability of the ATII-like cell lines A549 and NCIH441. Interestingly, the presence of an excess of sodium citrate on the surface of NPs not only reduced the in vitro viability of the cell lines A549 and NCIH441, as shown by MTT assay, but also affected cellular proliferation and increased the release of lactate dehydrogenase (LDH), as demonstrated by Ki-67 and LDH-release assays respectively. Furthermore, we investigated the internalization of AuNPs by transmission electron microscopy (TEM) and we observed that particles were internalized by active endocytosis in the cell lines A549 and NCIH441 within 3 hr. In addition, gold particles accumulated in membrane-bound vesicles and were not found freely dispersed in the cytoplasm. CONCLUSION: Our data suggest that the presence of contaminants, such as sodium citrate, on the surface of gold nanoparticles might play a pivotal role in inducing cytotoxicity in vitro, but does not influence the uptake of the particles in human ATII-like cell lines.

Barrier functions and paracellular integrity in human cell culture models of the proximal respiratory unit.

Airway epithelial cells provide a barrier to the translocation of inhaled materials. Tight (TJ) and adherens junctions (AJ) play a key role in maintaining barrier functions, and are responsible for the selective transport of various substances through the paracellular pathway. In this study we compared a bronchial cell line (16HBE14o-) and primary bronchial cells (HBEC), both cocultivated with the fibroblast cell line Wi-38, with respect to their structural differentiation and their reaction to cytokine stimulation. HBEC formed a pseudostratified epithelial layer and expressed TJ and AJ proteins after 2 weeks in coculture. Mucus-producing and ciliated cells were found within 24 days. Additionally, a beating activity of the ciliated HBEC (14-19Hz) could be detected. 16HBE14o- in coculture showed a multilayered growth without differentiation to a pseudostratified airway epithelium. Simultaneous exposure to TNF-alpha- and IFN-gamma-induced significant changes in barrier function and paracellular permeability in the cocultures of HBEC/Wi-38 but not in the 16HBE14o-/Wi-38. In summary, HBEC in coculture mimic the structure of native polarized bronchial epithelium showing basal, mucus-producing and ciliated cells. Our system provides an opportunity to examine the factors that influence barrier and mucociliary function of bronchial epithelium within a time frame of 3 weeks up to 3 months in an in vivo-like differentiated model.

Visions for regenerative medicine: interface between scientific fact and science fiction.

This article gives a brief overview of the authors' views on the future development of tissue engineering with respect to the challenges both to the materials and life sciences. Emphasis will be placed on the advantages of three-dimensional bioresorbable polymers in combination with relevant molecular cues and the application of autologous stem or progenitor cells. There is a requirement for much more diversity in the synthesis of so-called "intelligent" materials, which respond to external stimuli, as well as the development of novel drug and gene delivery systems. In addition, much more basic research is necessary in developmental biology and the application of modern cell and molecular biology to biomaterial questions.

Expression of the endothelial markers PECAM-1, vWf, and CD34 in vivo and in vitro.

EC culture models are essential to study pathological alterations of endothelial cells (ECs) in pulmonary vascular diseases under standardized conditions. Nevertheless, little is known about the spectrum of alterations of vessel-specific endothelial phenotypes in monolayer cultures. For the comparative study of endothelial markers in vivo and in vitro we investigated immunohistochemically the expression of PECAM-1, vWf, and CD34 by pulmonary ECs in vivo and in stimulated/unstimulated human umbilical vein endothelial cells (HU-VEC) and human pulmonary microvascular endothelial cells (HPMEC). In vivo, vessel type-specific expression patterns were found for vWf and CD34, while PECAM-1 was homogeneously and strongly expressed. While all HUVEC showed a marked vWf staining, about two-thirds of HPMEC exhibited a strong and the rest a moderate vWf staining. In both in vitro models all ECs were clearly PECAM-1-positive. However, only about 20% of the HUVEC and HPMEC were CD34-positive. Our results demonstrate the reduced expression of vessel type-specific endothelial phenotypes by endothelial monolayer cultures, stressing the need to improve culture conditions as well as develop cocultures and three-dimensional culture models. Moreover, the need for endothelial markers specific for single microvascular type ECs becomes obvious in order to establish cultures consisting of only one microvascular ECs subpopulation.

Comparative study of adhesion molecule expression in cultured human macro- and microvascular endothelial cells.

Culture systems as models for disease are only valid as long as they are comparable to in vivo conditions. The phenotype of cultured endothelial cells (ECs) has only been sporadically compared to the corresponding phenotype in vivo. Thus, we compared by immunolocalization the endothelial expression of ICAM-1, VCAM, and E-selectin in vivo in stimulated/unstimulated human umbilical vein endothelial cells (HUVEC) as a model for macrovascular ECs and stimulated/unstimulated HPMEC (human pulmonary microvessel endothelial cells) as a model for pulmonary microvascular ECs with that in human lungs in vivo (normal and ARDS). Proinflammatory stimuli in vitro were used to stimulate conditions relevant for ARDS. ICAM-1 expression in stimulated HUVEC/HPMEC correlated well with in vivo expression (macro- and microvessels). For E-selectin, the staining pattern in macro/microvessels correlated moderately with unstimulated and well with stimulated HUVEC/HPMEC. For VCAM a good correlation was found for stimulated/unstimulated HUVEC and unstimulated HPMEC. The expression patterns in stimulated HUVEC corresponded well for all three molecules with those in vivo. Thus, the expression patterns in vitro are only partially transferable to in vivo conditions. The study suggests that E-selectin- and VCAM-coated beads could potentially serve in the isolation process of arteriolar and venular ECs.