Differentiation-associated modulation of heparan sulfate structure and function in CaCo-2 colon carcinoma cells

Heparan sulfate species expressed by different cell and tissue types differ in their structural and functional properties. Limited information is available on differences in regulation of heparan sulfate biosynthesis within a single tissue or cell population under different conditions. We have approached this question by studying the effect of cell differentiation on the biosynthesis and function of heparan sulfate in human colon carcinoma cells (CaCo-2). These cells undergo spontaneous differentiation in culture when grown on semipermeable supports; the differentiated cells show phenotypic similarity to small intestine enterocytes. Metabolically labeled heparan sulfate was isolated from the apical and basolateral media from cultures of differentiated and undifferentiated cells. Compositional analysis of disaccharides, derived from the contiguous N-sulfated regions of heparan sulfate, indicated a greater proportion of 2-O- sulfated iduronic acid units and a smaller amount of 6-O-sulfated glucosamine units in differentiated than in undifferentiated cells. By contrast, the overall degree of sulfation, the chain length and the size distribution of the N-acetylated regions were similar regardless the differentiation status of the cells. The structural changes were found to affect the binding of heparan sulfate to the long isoform of platelet-derived growth factor A chain but not to fibroblast growth factor 2. These findings show that heparan sulfate structures change during cell differentiation and that heparan sulfate-growth factor interactions may be affected by such changes.      

Ultrastructure of lung elastin and collagen in mouse models of spontaneous emphysema.

The tight-skin (Tsk) and beige (bg) mutants of the C57B1/6J strain of mouse spontaneously develop air-space enlargement reminiscent of human emphysema. To determine if this enlargement is accompanied by matrix destruction, as in the human disease, we examined the elastin and collagen matrices of the lungs of both mutants. The ultrastructure of these matrix components was separately visualized by scanning electron microscopy following controlled alkali digestion, which preserves collagen, and formic acid digestion, which enables visualization of elastin. Significant elastin destruction suggestive of an elastolytic process was observed in the lungs of Tsk mice. Thickening of elastin lamellae was observed in the lungs of bg mice, suggesting that congenital matrix remodeling may underlie air-space enlargement in this strain.     

Ulysses - an application for the projection of molecular interactions across species

We developed Ulysses as a user-oriented system that uses a process called Interolog Analysis for the parallel analysis and display of protein interactions detected in various species. Ulysses was designed to perform such Interolog Analysis by the projection of model organism interaction data onto homologous human proteins, and thus serves as an accelerator for the analysis of uncharacterized human proteins. The relevance of projections was assessed and validated against published reference collections. All source code is freely available, and the Ulysses system can be accessed via a web interface .     

Fetal microchimerism in kidney biopsies of lupus nephritis patients may be associated with a beneficial effect

IntroductionMicrochimeric male fetal cells (MFCs) have been associated with systemic lupus erythematosus, and published studies have further correlated MFC with lupus nephritis (LN). In the present study, we evaluated the frequency of MFC in the renal tissue of patients with LN.MethodsTwenty-seven renal biopsies were evaluated: Fourteen were from women with clinical and laboratory findings of LN, and thirteen were from controls. Genomic DNA was extracted from kidney biopsies, and the male fetal DNA was quantified using real-time quantitative polymerase chain reactions for the detection of specific Y chromosome sequences.ResultsMFCs were detected in 9 (64%) of 14 of patients with LN, whereas no MFCs were found in the control group (P = 0.0006). No differences in pregnancy history were found between patients with LN and the control group. Significantly higher amounts of MFCs were found in patients with LN with serum creatinine ≤1.5 mg/dl. Furthermore, women with MFCs had significantly better renal function at the time of biopsy (P = 0.03). In contrast, patients with LN without MFCs presented with more severe forms of glomerulonephritis (World Health Organization class IV = 60% and class V = 40%).ConclusionsOur data indicate a high prevalence of MFCs in renal biopsy specimens from women with LN, suggesting a role for MFCs in the etiology of LN. The present report also provides some evidence that MFCs could have a beneficial effect in this disease.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0615-4) contains supplementary material, which is available to authorized users.     

USP18 lack in microglia causes destructive interferonopathy of the mouse brain

Microglia are tissue macrophages of the central nervous system (CNS) that control tissue homeostasis. Microglia dysregulation is thought to be causal for a group of neuropsychiatric, neurodegenerative and neuroinflammatory diseases, called “microgliopathies”. However, how the intracellular stimulation machinery in microglia is controlled is poorly understood. Here, we identified the ubiquitin‐specific protease (Usp) 18 in white matter microglia that essentially contributes to microglial quiescence. We further found that microglial Usp18 negatively regulates the activation of Stat1 and concomitant induction of interferon‐induced genes, thereby terminating IFN signaling. The Usp18‐mediated control was independent from its catalytic activity but instead required the interaction with Ifnar2. Additionally, the absence of Ifnar1 restored microglial activation, indicating a tonic IFN signal which needs to be negatively controlled by Usp18 under non‐diseased conditions. These results identify Usp18 as a critical negative regulator of microglia activation and demonstrate a protective role of Usp18 for microglia function by regulating the Ifnar pathway. The findings establish Usp18 as a new molecule preventing destructive microgliopathy.     

Evaluation of APOE polymorphisms and the risk for age-related macular degeneration in a Southeastern Brazilian population

This study aimed to evaluate the role of APOE polymorphisms (rs429358 and rs7412) in the risk of age-related macular degeneration in a sample of the Southeastern Brazilian population. Seven hundred and five unrelated individuals were analyzed, 334 with age-related macular degeneration (case group), and 371 without the disease (control group). In the case group, patients were further stratified according to disease phenotypes, divided into dry and wet age-related macular degeneration, and non-advanced and advanced age-related macular degeneration. APOE polymorphisms (rs429358 and rs7412) were evaluated through polymerase chain reaction and direct sequencing. In the comparison of cases vs. controls, none of the associations reached statistical significance, considering the Bonferroni-adjusted P-value, although there was a suggestive protection for the E3/E4 genotype (OR = 0.626; P-value = 0.037) and E4 carriers (OR = 0.6515; P-value = 0.047). Statistically significant protection for both the E3/E4 genotype and E4 carriers was observed in the comparisons: advanced age-related macular degeneration vs. controls (OR = 0.3665, P-value = 0.491 × 10⁻³ and OR = 0.4031, P-value = 0.814 × 10⁻³, respectively), advanced age-related macular degeneration vs. non-advanced age-related macular degeneration (OR = 0.2529, P-value = 0.659 × 10⁻⁴ and OR = 0.2692, P-value = 0.631 × 10⁻⁴, respectively). In the comparison of wet age-related macular degeneration vs. control, protection was statistically significant only for E3/E4 (OR = 0.4052, P-value = 0.001). None of the comparisons demonstrated any significant association for E2 genotypes or E2 carriers in age-related macular degeneration risk in this study. Findings suggest a protective role of the E4 haplotype in the APOE gene in the risk for advanced and wet forms of age-related macular degeneration, in a sample of the Brazilian population. To our knowledge, this is the first Brazilian study to show the association between APOE polymorphisms and age-related macular degeneration.     

Chitooligosaccharide inhibits RANKL-induced osteoclastogenesis and ligation-induced periodontitis by suppressing MAPK/ c-fos/NFATC1 signaling

Considering the high rate of osteoclast-related diseases worldwide, research targeting osteoclast formation/function is crucial. In vitro, we demonstrated that chitooligosaccharide (CS) dramatically inhibited osteoclastogenesis as well as osteoclast function dose-dependently. CS suppressed osteoclast-specific genes expression during osteoclastogenesis. Furthermore, we found that CS attenuated receptor activator of nuclear factor kappa B ligand (RANKL)-mediated mitogen-activated protein kinase (MAPK) pathway involving p38, erk1/2, and jnk, leading to the reduced expression of c-fos and nuclear factor of activated T cells c1 (NFATc1) during osteoclast differentiation. In vivo, we found CS protected rats from periodontitis-induced alveolar bone loss by micro-computerized tomography and histological analysis. Overall, CS inhibited RANKL-induced osteoclastogenesis and ligature-induced rat periodontitis model, probably by suppressing the MAPK/c-fos/NFATc1 signaling pathway. Therefore, CS may be a safe and promising treatment for osteoclast-related diseases.     

HERG1 Currents in Native K562 Leukemic Cells

The human ether-a-go-go related gene (HERG1) K⁺ channel is expressed in neoplastic cells, in which it was proposed to play a role in proliferation, differentiation and/or apoptosis. K562 cells (a chronic myeloid leukemic human cell line) express both the full-length (herg1a) and the N-terminally truncated (herg1b) isoforms of the gene, and this was confirmed with Western blots and coimmunoprecipitation experiments. Whole-cell currents were studied with a tail protocol. Seventy-eight percent of cells showed a HERG1-like current: repolarization to voltages negative to −40 mV produced a transient peak inward tail current, characteristic of HERG1 channels. Cells were exposed to a HERG-specific channel blocker, E4031. Half-maximal inhibitory concentration (IC₅₀) of the blocker was 4.69 nm. The kinetics of the HERG1 current in K562 cells resembled the rapid component of the native cardiac delayed rectifier current, known to be conducted by heterotetrameric HERG1 channels. Fast and slow deactivation time constants at −120 mV were 27.5 and 239.5 ms, respectively. Our results in K562 cells suggest the assembling of heterotetrameric channels, with some parameters being dominated by one of the isoforms and other parameters being intermediate. Hydrogen peroxide was shown to increase HERG1a K⁺ current in heterologous expression systems, which constitutes an apoptotic signal. However, we found that K562 HERG1 whole-cell currents were not activated by H₂O₂.