Cytocompatibility of novel extracellular matrix protein analogs of biodegradable polyester polymers derived from α-hydroxy amino acids

One of the challenges in regenerative medicine is the development of novel biodegradable materials to build scaffolds that will support multiple cell types for tissue engineering. Here we describe the preparation, characterization, and cytocompatibility of homo- and hetero-polyesters of α-hydroxy amino acid derivatives with or without lactic acid conjugation. The polymers were prepared by a direct condensation method and characterized using gel permeation chromatography, ¹H-nuclear magnetic resonance spectroscopy, Fourier transform infrared spectroscopy, differential scanning calorimetry, optical activity, and solubility. The surface charge of the polymers was evaluated using zeta potential measurements. The polymers were coated onto glass cover slips followed by characterization using nano-surface profiler, thin film reflectometry, and atomic force microscopy (AFM). Their interaction with endothelial and neuronal cells was assessed using adhesion, proliferation, and differentiation assays. Of the characterized polymers, Poly-HOVal-LA, but not Poly-(D)HOPhe, significantly augmented nerve growth factor (NGF)-induced neuronal differentiation of the PC12 pheochromcytoma cells. In contrast, Poly-HOLeu increased by 20% the adhesion of endothelial cells, but did not affect PC12 cell differentiation. NGF-induced Erk1/2 phosphorylation in PC12 cells grown on the different polymers was similar to the effect observed for cells cultured on collagen type I. While no significant association could be established between charge and the differentiative/proliferative properties of the polymers, AFM analysis indicated augmentation of NGF-induced neuronal differentiation on smooth polymer surfaces. We conclude that overall selective cytocompatibility and bioactivity might render α-hydroxy amino acid polymers useful as extracellular matrix-mimicking materials for tissue engineering.     

Human intestinal epithelial cells respond to β‐glucans via Dectin‐1 and Syk

Intestinal epithelial cells (IECs) are the first to encounter luminal antigens and may be involved in intestinal immune responses. Fungi are important components of the intestinal microflora. The potential role of fungi, and in particular their cell wall component β‐glucan, in modulating human intestinal epithelial responses is still unclear. Here we examined whether human IECs are capable of recognizing and responding to β‐glucans, and the potential mechanisms of their activation. We show that human IECs freshly isolated from surgical specimens, and the human IEC lines HT‐29 and SW480, express the β‐glucan receptor Dectin‐1. The β‐glucan‐consisting glycans curdlan and zymosan stimulated IL‐8 and CCL2 secretion by IEC lines. This was significantly inhibited by a Dectin‐1 blockade using its soluble antagonist laminarin. Spleen tyrosine kinase (Syk), a signaling mediator of Dectin‐1 activation, is expressed in human IECs. β‐glucans and Candida albicans induced Syk phosphorylation, and Syk inhibition significantly decreased β‐glucan‐induced chemokine secretion from IECs. Thus, IECs may respond to β‐glucans by the secretion of pro‐inflammatory chemokines in a Dectin‐1‐ and Syk‐dependent pathway, via receptors and a signaling pathway described to date only for myeloid cells. These findings highlight the importance of fungi–IEC interactions in intestinal inflammation.     

Critical role of the endogenous interferon ligand–receptors in type I and type II interferons response

Separate ligand–receptor paradigms are commonly used for each type of interferon (IFN). However, accumulating evidence suggests that type I and type II IFNs may not be restricted to independent pathways. Using different cell types deficient in IFNAR1, IFNAR2, IFNGR1, IFNGR2 and IFN‐γ, we evaluated the contribution of each element of the IFN system to the activity of type I and type II IFNs. We show that deficiency in IFNAR1 or IFNAR2 is associated with impairment of type II IFN activity. This impairment, presumably resulting from the disruption of the ligand–receptor complex, is obtained in all cell types tested. However, deficiency of IFNGR1, IFNGR2 or IFN‐γ was associated with an impairment of type I IFN activity in spleen cells only, correlating with the constitutive expression of type II IFN (IFN‐γ) observed on those cells. Therefore, in vitro the constitutive expression of both the receptors and the ligands of type I or type II IFN is critical for the enhancement of the IFN activity. Any IFN deficiency can totally or partially impair IFN activity, suggesting the importance of type I and type II IFN interactions. Taken together, our results suggest that type I and type II IFNs may regulate biological activities through distinct as well as common IFN receptor complexes.     

Proteus syndrome review: molecular,clinical, and pathologic features

Proteus syndrome is caused by an activating AKT1 mutation (c.49G>A, p.Glu17Lys). Many variable features are possible in this mosaic disorder, including: (i) disproportionate, asymmetric, and distorting overgrowth; (ii) bone abnormalities different from those observed in other disorders; (iii) a characteristic cerebriform connective tissue nevus made up of highly collagenized connective tissue; (iv) epidermal nevi in early life, consisting of acanthosis and hyperkeratosis; (v) vascular malformations of the capillary, venous, or lymphatic types; (vi) dysregulated adipose tissue including lipomas, lipohypoplasia, fatty overgrowth, and localized fat deposits; (vii) other unusual features, including bullous lung alterations; specific neoplasms; a facial phenotype associated with intellectual disability and/or seizures, and/or brain malformations; and (viii) deep vein thrombosis, resulting in premature death. Concluding remarks address diagnostic criteria, natural history, management, psychosocial issues, and differential diagnosis.     

Temporal Kinetics and Quantitative Analysis of Cryptococcus neoformans Nonlytic Exocytosis

Cryptococcus neoformans is a facultative intracellular pathogen and the causative agent of cryptococcosis, a disease that is often fatal to those with compromised immune systems. C. neoformans has the capacity to escape phagocytic cells through a process known as nonlytic exocytosis whereby the cryptococcal cell is released from the macrophage into the extracellular environment, leaving both the host and pathogen alive. Little is known about the mechanism behind nonlytic exocytosis, but there is evidence that both the fungal and host cells contribute to the process. In this study, we used time-lapse movies of C. neoformans-infected macrophages to delineate the kinetics and quantitative aspects of nonlytic exocytosis. We analyzed approximately 800 macrophages containing intracellular C. neoformans and identified 163 nonlytic exocytosis events that were further characterized into three subcategories: type I (complete emptying of macrophage), type II (partial emptying of macrophage), and type III (cell-to-cell transfer). The majority of type I and II events occurred after several hours of intracellular residence, whereas type III events occurred significantly (P < 0.001) earlier in the course of macrophage infection. Our results show that nonlytic exocytosis is a morphologically and temporally diverse process that occurs relatively rapidly in the course of macrophage infection.     

Epstein–Barr virus antibodies mark systemic lupus erythematosus and scleroderma patients negative for anti‐DNA

Systemic lupus erythematosus (SLE) is an autoimmune disease that can attack many different body organs; the triggering event is unknown. SLE has been associated with more than 100 different autoantibody reactivities – anti‐dsDNA is prominent. Nevertheless, autoantibodies to dsDNA occur in only two‐thirds of SLE patients. We previously reported the use of an antigen microarray to characterize SLE serology. We now report the results of an expanded study of serology in SLE patients and scleroderma (SSc) patients compared with healthy controls. The analysis validated and extended previous findings: two‐thirds of SLE patients reacted to a large spectrum of self‐molecules that overlapped with their reactivity to dsDNA; moreover, some SLE patients manifested a deficiency of natural IgM autoantibodies. Most significant was the finding that many SLE patients who were negative for autoantibodies to dsDNA manifested abnormal antibody responses to Epstein–Barr virus (EBV): these subjects made IgG antibodies to EBV antigens to which healthy subjects did not respond or they failed to make antibodies to EBV antigens to which healthy subjects did respond. This observation suggests that SLE may be associated with a defective immune response to EBV. The SSc patients shared many of these serological abnormalities with SLE patients, but differed from them in increased IgG autoantibodies to topoisomerase and centromere B; 84% of SLE patients and 58% of SSc patients could be detected by their abnormal antibodies to EBV. Hence an aberrant immune response to a ubiquitous viral infection such as EBV might set the stage for an autoimmune disease.