Increased levels of circulating microparticles in primary Sj?gren's syndrome,systemic lupus erythematosus and rheumatoid arthritis and relation with disease activity

Introduction

Cell stimulation leads to the shedding of phosphatidylserine (PS)-rich microparticles (MPs). Because autoimmune diseases (AIDs) are characterized by cell activation, we investigated level of circulating MPs as a possible biomarker in primary Sjögren''s syndrome (pSS), systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA).

Methods

We measured plasma levels of total, platelet and leukocyte MPs by prothrombinase capture assay and flow cytometry in 43 patients with pSS, 20 with SLE and 24 with RA and in 44 healthy controls (HCs). Secretory phospholipase A2 (sPLA2) activity was assessed by fluorometry. Soluble CD40 ligand (sCD40L) and soluble P-selectin (sCD62P), reflecting platelet activation, were measured by ELISA.

Results

Patients with pSS showed increased plasma level of total MPs (mean ± SEM 8.49 ± 1.14 nM PS equivalent (Eq), P < 0.0001), as did patients with RA (7.23 ± 1.05 n PS Eq, P = 0.004) and SLE (7.3 ± 1.25 nM PS Eq, P = 0.0004), as compared with HCs (4.13 ± 0.2 nM PS Eq). Patients with AIDs all showed increased level of platelet MPs (P < 0.0001), but only those with pSS showed increased level of leukocyte MPs (P < 0.0001). Results by capture assay and flow cytometry were correlated. In patients with high disease activity according to extra-glandular complications (pSS), DAS28 (RA) or SLEDAI (SLE) compared with low-activity patients, the MP level was only slightly increased in comparison with those having a low disease activity. Platelet MP level was inversely correlated with anti-DNA antibody level in SLE (r = -0.65; P = 0.003) and serum β2 microglobulin level in pSS (r = -0.37; P < 0.03). The levels of total and platelet MPs were inversely correlated with sPLA2 activity (r = -0.37, P = 0.0007; r = -0.36, P = 0.002, respectively). sCD40L and sCD62P concentrations were significantly higher in pSS than in HC (P ≤ 0.006).

Conclusions

Plasma MP level is elevated in pSS, as well as in SLE and RA, and could be used as a biomarker reflecting systemic cell activation. Level of leukocyte-derived MPs is increased in pSS only. The MP level is low in case of more severe AID, probably because of high secretory phospholipase A2 (sPLA2) activity, which leads to consumption of MPs. Increase of platelet-derived MPs, sCD40L and sCD62P, highlights platelet activation in pSS.     

TGF-beta1 genotypes in cirrhosis: relationship with the occurrence of liver cancer

This study aimed to verify whether specific single nucleotide polymorphisms (SNPs) of the transforming growth factor-beta1 (TGF-beta1) may predispose to end-stage liver disease and/or hepatocellular carcinoma (HCC). One hundred eighty-eight consecutive patients transplanted for liver cirrhosis (HBV N=21, HCV N=68, alcoholic N=55 and others N=23) and a control group of 140 healthy blood donors were investigated. Four SNPs were studied by restriction fragment length assays: -800G>A, -509C>T, Leu10Pro and Arg25Pro. Patients were found to possess the -509T/ * (TT 53/188, CT 85/188, CC 50/188 vs TT 22/140, CT 61/140, CC 57/140; p<0.002) and Arg25Pro C/ * genotypes (CC 1/188, CG 31/188, GG 156/188 vs CC 0/140, CG 13/140, GG 127/140; p<0.05) more frequently than controls. Patients with cirrhosis complicated by HCC possessed more frequently the Leu10Pro T/ * genotype than patients without HCC (TT 20/54, CT 26/54, CC 8/54 vs TT 31/134, CT 69/134, CC 34/134; p<0.05). The analysis of molecular variance detected significant genotypic differentiations between controls and cirrhotics but not between cirrhotics with or without HCC. In conclusion, TGF-beta1 SNPs probably facilitate the development of liver cirrhosis, while they seem to have a limited role in predicting the occurrence of HCC.     

Identification of long chain alkylidenemalonates as novel small molecule modulators of histone acetyltransferases

Pentadecylidenemalonate 1b, a simplified analogue of anacardic acid, was identified as the first mixed activator/inhibitor of histone acetyltransferases (HATs). It potentiates PCAF HAT activity while inhibiting those of p300/CBP and recombinant CBP. The remarkable apoptotic effect together with the ability to selectively acetylate histone versus non-histone substrates appoint 1b as a lead for the development of anticancer drugs.     

Chemical Characterization and Biological Activities of Phenolic‐Rich Fraction from Cauline Leaves of Isatis tinctoria L. (Brassicaceae) Growing in Sicily,Italy

The present work focused on the evaluation of the antioxidant and cytotoxic activities of the phenolic‐rich fraction (ItJ‐EAF) obtained from cauline leaves collected in January from Isatis tinctoria L. (Brassicaceae) growing wild around Acireale (Sicily, Italy). The total phenolic, flavonoid, and condensed tannin contents of the fraction were determined spectrophotometrically, whereas the phenolic profile was assessed by HPLC‐PDA/ESI‐MS analysis. A total of 20 compounds were positively identified and twelve out of them were never previously reported in I. tinctoria leaves. The fraction exhibited good radical scavenging activity in DPPH test (IC₅₀ = 0.6657 ± 0.0024 mg/ml) and reducing power (3.87 ± 0.71 ASE/ml), whereas, it neither showed chelating activity nor was able to counteract H₂O₂ induced oxidative stress damage in Escherichia coli. The antiproliferative effect was evaluated in vitro on two human anaplastic thyroid carcinoma cell lines (CAL‐62 and 8505C) by MTT assay. At the highest tested concentration ItJ‐EAF significantly reduced (80%) the growth of CAL‐62 cells. No cytotoxicity against Artemia salina was observed. It can be concluded that I. tinctoria cauline leaves represent a source of phenolic compounds which could be potentially used as chemopreventive or adjuvant agents against cancer.     

Immunosenescence in aging: between immune cells depletion and cytokines up-regulation

Background

The immunosenescence is a relatively recent chapter, correlated with the linear extension of the average life began in the nineteenth century and still in progress. The most important feature of immunosenescence is the accumulation in the “immunological space” of memory and effector cells as a result of the stimulation caused by repeated clinical and subclinical infections and by continuous exposure to antigens (inhalant allergens, food, etc.). This state of chronic inflammation that characterizes senescence has a significant impact on survival and fragility. In fact, the condition of frail elderly occurs less frequently in situations characterized by poor contact with viral infections and parasitic diseases. Furthermore the immunosenescence is characterized by a particular “remodelling” of the immune system, induced by oxidative stress. Apoptosis plays a central role in old age, a period in which the ability of apoptosis can change. The remodelling of apoptosis, together with the Inflammaging and the up-regulation of the immune response with the consequent secretion of pro-inflammatory lymphokines represents the major determinant of the rate of aging and longevity, as well as of the most common diseases related with age and with tumors. Other changes occur in the innate immunity, the first line of defence providing rapid, but unspecific and incomplete protection, consisting mostly of monocytes, natural killer cells and dendritic cells, acting up to the establishment of a adaptive immune response, which is slower, but highly specific, which cellular substrate consists of T and B lymphocytes. The markers of “Inflammaging” in adaptive immunity in centenarians are characterized by a decrease in T cells “naive.” The reduction of CD8 virgins may be related to the risk of morbidity and death, as well as the combination of the increase of CD8+ cells and reduction of CD4+ T cells and the reduction of CD19+ B cells. The immune function of the elderly is weakened to due to the exhaustion of T cell-virgin (CD95?), which are replaced with the clonal expansion of CD28-T cells.

Conclusions

The increase of pro-inflammatory cytokines is associated with dementia, Parkinson’s disease, atherosclerosis, diabetes type 2, sarcopenia and a high risk of morbidity and mortality. A correct modulation of immune responses and apoptotic phenomena can be useful to reduce age-related degenerative diseases, as well as inflammatory and neoplastic diseases.

     

Structure, organization and characterization of the gene cluster involved in the production of microcin E492, a channel-forming bacteriocin

Microcin E492 is a low-molecular-weight, channel-forming bacteriocin produced and excreted by Klebsiella pneumoniae RYC492. A 13 kb chromosomal DNA fragment from K. pneumoniae RYC492 was sequenced, and it was demonstrated by random Tn5 mutagenesis that most of this segment, which has at least 10 cistrons, is needed for the production of active microcin and its immunity protein. Genes mceG and mceH correspond to an ABC exporter and its accessory protein, respectively, and they are closely related to the colicin V ABC export system. The microcin E492 system also requires the product of gene mceF as an additional factor for export. Despite the fact that this bacteriocin lacks post-translational modifications, genes mceC, mceI and mceJ are needed for the production of active microcin. Genes mceC and mceI are homologous to a glycosyl transferase and acyltransferase, respectively, whereas mceJ has no known homologue. Mutants in these three genes secrete an inactive form of microcin, able to form ion channels in a phospholipidic bilayer, indicating that the mutation of these microcin genes does not alter the process of membrane insertion. On the other hand, microcin isolated from mutants in genes mceC and mceJ has a lethal effect when incubated with spheroplasts of sensitive cells, indicating that the microcin defects in these mutants are likely to alter receptor recognition at the outer membrane. A model for synthesis and export is proposed as well as a novel maturation pathway that would involve conformational changes to explain the production of active microcin E492.     

Actin remodeling driven by circLIMA1: sperm cell as an intriguing cellular model

CircRNA cargo in spermatozoa (SPZ) participates in setting cell quality, in terms of morphology and motility. Cannabinoid receptor CB1 activity is correlated with a proper spermatogenesis and epididymal sperm maturation. Despite CB1 promotes endogenous skill to circularize mRNAs in SPZ, few notions are reported regarding the functional link between endocannabinoids and spermatic circRNA cargo. In CB1 knock-out male mice, we performed a complete dataset of spermatic circRNA content by microarray strategy.Differentially expressed (DE)-circRNAs, as a function of genotype, were identified. Within DE-circRNAs, we focused the attention on circLIMA1, as putative actin-cytoskeleton architecture regulator. The validation of circLIMA1 dependent-competitive endogenous RNA (ceRNA) network (ceRNET) in in vitro cell line confirmed its activity in the regulation of the cytoskeletal actin. Interestingly, a dynamic actin regulation in SPZ nuclei was found during their epididymal maturation.In this scenario, we showed for the first time an intriguing sperm nuclear actin remodeling, regulated via a ceRNET-independent pathway, consisting in the nuclear shuttling of circLIMA1-QKI interactome and downstream in Gelsolin regulation. In particular, the increased levels of circLIMA1 in CB1 knock-out SPZ, associated with an inefficient depolymerization of nuclear actin, specifically illustrate how endocannabinoids, by regulating circRNA cargo, may contribute to sperm morpho-cellular maturation.