Statins prevent endothelial cell activation induced by antiphospholipid (anti-beta2-glycoprotein I) antibodies: effect on the proadhesive and proinflammatory phenotype.

OBJECTIVE: To investigate the ability of statins, the inhibitors of the hydroxymethylglutaryl-coenzyme A reductase enzyme, to affect endothelial cell activation induced by anti-beta2-glycoprotein I (anti-beta2GPI) antibodies in vitro. METHODS: Human umbilical vein endothelial cell (HUVEC) activation was evaluated as U937 monocyte adhesion, E-selectin, and intercellular adhesion molecule I (ICAM-1) expression by cell enzyme-linked immunosorbent assay and as interleukin-6 (IL-6) messenger RNA (mRNA) expression by RNA protection assay. E-selectin-specific nuclear factor kappaB (NF-kappaB) DNA-binding activity was evaluated by the gel-shift assay. HUVECs were activated by polyclonal affinity-purified IgG, human monoclonal IgM anti-beta2GPI antibodies, human recombinant IL-1beta, tumor necrosis factor alpha, or lipopolysaccharide (LPS). RESULTS: Fluvastatin reduced, in a concentration-dependent manner (1-10 microM), the adhesion of U937 to HUVECs and the expression of E-selectin and ICAM-1 induced by anti-beta2GPI antibodies as well as by cytokines or LPS. Another lipophilic statin, simvastatin, displayed similar effects but to a lesser extent than fluvastatin. The inhibition of E-selectin expression exerted by fluvastatin was related to the impairment of NF-kappaB binding to DNA. Moreover, the drug attenuated the expression of IL-6 mRNA in HUVEC exposed to anti-beta2GPI antibodies or cytokines. Incubation of HUVECs with mevalonate (100 microM), concomitantly with fluvastatin, greatly prevented the inhibitory effect of statin. CONCLUSION: Endothelial activation mediated by anti-beta2GPI antibody can be inhibited by statins. Because of the suggested role of endothelial cell activation in the pathogenesis of antiphospholipid syndrome (APS), our data provide, for the first time, a rationale for using statins as an additional therapeutic tool in APS.     

Rituximab Therapy in Lupus Nephritis: Current Clinical Evidence

The complexity of the therapeutic approach in lupus nephritis (LN) is increased by the large number of patients who do not respond to first-line therapies and by relapses after initial clinical remission. The emergence of biological agents has increased the therapeutic armamentarium available in these complex situations, but their use is limited by the lack of licensing. We analysed current evidence on the therapeutic use of rituximab in adult LN patients by systematic analysis of seven observational studies published since 2005 (four in 2009), which included 106 LN patients treated with rituximab. A complete or partial therapeutic response was achieved in 73 (69%) patients. The response according to the type of LN was stated in 79 cases: 8 (80%) patients with type III LN had a favourable, 26 (67%) of those with type IV, 4 (57%) of those with type V and 18 (78%) of those with mixed membranous-proliferative LN. The main factors associated with no response were younger age, black race and lack of CD19⁺ cell depletion. The lowest rates of complete response were observed in patients with type V LN, especially those with associated proliferative lesions. Although it is not yet possible to make definite recommendations, the global analysis of these cases supports the off-label use of rituximab in severe, refractory LN cases.     

Fine specificity of autoantibodies to calreticulin: epitope mapping and characterization

Extracellular calreticulin (CRT) as well as anti‐CRT antibodies have been reported in patients with various autoimmune disorders and CRT has been implicated in ‘epitope spreading’ to other autoantigens such as the Ro/SS‐A complex. In addition, antibodies against parasite forms of the endoplasmic reticulum chaperone, CRT, have been found in patients suffering from onchocerciasis and schistosomiasis. In this study, we screened sera for anti‐CRT antibodies from patients with active and inactive systemic lupus ertythematosus (SLE) and primary or secondary Sjögren’s syndrome. Approximately 40% of all SLE patients were positive for anti‐CRT antibodies. The antigenic regions of CRT were determined using full length CRT and fragments of CRT prepared in yeast and Escherichia coli, respectively. Synthetic 15mer peptides corresponding to the major autoantigenic region of CRT (amino acids 1–289), each one overlapping by 12 amino acids, were used to map the B cell epitopes on the CRT protein recognized by autoimmune sera. Major antigenic epitopes were found to be associated with the N‐terminal half of the protein in 69% of the SLE sera from active disease patients, while the C‐domain was not antigenic. Major epitopes were found to be reactive with antibodies in sera from SLE patients with both active and inactive disease, spanning different regions of the N and P‐domains. Sera from both healthy and disease controls and primary Sjögren’s syndrome patients were non‐reactive to these sequences. Limited proteolysis of CRT with two major leucocyte serine proteases, elastase and cathepsin G, demonstrated that an N‐terminal region of CRT is resistant to digestion. Interestingly, some of the epitopes with the highest reactivity belong to the fragments of the protein which bind to C1q and inhibit complement activation. Whether C1q association with CRT is a pathological or protective interaction between these two proteins is currently under investigation.     

Neonatal Liver Disease Associated with Placental Transfer of Anti-mitochondrial Antibodies

Background: Anti-mitochondrial antibody is the diagnostic hallmark of primary biliary cirrhosis. Its role in the aetiology of primary biliary cirrhosis is controversial. Methods: Two cases of neonatal hepatitis seropositive for anti-mitochondrial antibody are described. Anti-mitochondrial antibody Ig isotype and epitopic specificity were investigated by immunofluorescence and enzyme immunoassays. Results: In both infants anti-mitochondrial antibody was of the G class, mainly G1 and G3 subclasses, and reacted with two synthetic peptides reproducing major M2 epitopic regions: inner lipoyl domain pyruvate dehydrogenase complex (PDC)-E2_162-176 and PDC-E3 binding protein (PDC-E3BP)_86-100. One infant also reacted with outer lipoyl domain PDC-E2_35-49, and 2-oxoglutarate dehydrogenase complex (OGDC)-E2_99-113. An identical pattern of reactivity was present in their mothers, indicating the maternal origin of the antibodies. Anti-mitochondrial antibody disappeared in the infants with the disappearance of the liver pathology. Conclusions: The simultaneous disappearance of hepatitis and anti-mitochondrial antibody in the infants suggests a possible causal link between the two.     

Antiphospholipid antibodies in homozygous sickle cell disease.

Serum samples from 108 unselected Jamaican patients with homozygous sickle cell disease and 116 control subjects with normal haemoglobin were screened for the presence of antiphospholipid antibodies. Slightly increased levels of IgG antiphospholipid antibodies were found in nine patients with sickle cell disease and in none of the control subjects. Serial control samples confirmed the increased levels of antiphospholipid antibodies. A comparison of the haematological and clinical features of patients with positive and negative antiphospholipid sickle cell disease did not highlight any differences between the groups.     

A technique of using gated-CT images to determine internal target volume (ITV) for fractionated stereotactic lung radiotherapy.

BACKGROUND AND PURPOSE: To develop and evaluate a technique and procedure of using gated-CT images in combination with PET image to determine the internal target volume (ITV), which could reduce the planning target volume (PTV) with adequate target coverage. PATIENTS AND METHODS: A skin marker-based gating system connected to a regular single slice CT scanner was used for this study. A motion phantom with adjustable motion amplitude was used to evaluate the CT gating system. Specifically, objects of various sizes/shapes, considered as virtual tumors, were placed on the phantom to evaluate the number of phases of gated images required to determine the ITV while taking into account tumor size, shape and motion. A procedure of using gated-CT and PET images to define ITV for patients was developed and was tested in patients enrolled in an IRB approved protocol. RESULTS: The CT gating system was capable of removing motion artifacts for target motion as large as 3-cm when it was gated at optimal phases. A phantom study showed that two gated-CT scans at the end of expiration and the end of inspiration would be sufficient to determine the ITV for tumor motion less than 1-cm, and another mid-phase scan would be required for tumors with 2-cm motion, especially for small tumors. For patients, the ITV encompassing visible tumors in all sets of gated-CT and regular spiral CT images seemed to be consistent with the target volume determined from PET images. PTV expanded from the ITV with a setup uncertainty margin had less volume than PTVs from spiral CT images with a 10-mm generalized margin or an individualized margin determined at fluoroscopy. CONCLUSIONS: A technique of determining the ITV using gated-CT images was developed and was clinically implemented successfully for fractionated stereotactic lung radiotherapy.