Case report: Breaking CNS immuno-privilege: TNFα-inhibitor triggers aseptic meningitis in a patient with rheumatoid arthritis.

Blood-brain barrier dysfunction might be driven by peripheral inflammation. TNFα inhibitors (TNF-αi) are occasionally associated with a wide spectrum of neurological immuno-mediated disorders. However, patients with systemic autoimmune disorders, including rheumatoid arthritis (RA), might be prone to develop further organ-specific, including central nervous system (CNS), autoimmunity. Here we report the case of a patient, affected by RA and treated with etanercept, who suddenly developed focal neurological symptoms. Cerebrospinal fluid, magnetic resonance imaging (MRI), and positron emission tomography (PET)/MRI findings are reported and support the diagnosis of TNF-αi -associated aseptic meningitis.

Relapsing tumefactive demyelination lesions: A unique, distinct inflammatory brain pathology.

We report the case of a patient suffering from biopsy-proven relapsing tumefactive demyelinating lesions (TDLs) of the central nervous system who had five relapses in 16 years. No signs/symptoms suggestive of alternative pathologies emerged during the follow-up. A limited benefit was observed with intravenous (IV) high-dose steroids, while both plasma exchange and IV immunoglobulin G (IgG) administration were ineffective. A long-lasting (9 years) but transient clinical stabilization was obtained with cyclophosphamide. Our case supports the view that recurrent TDL is a relapsing brain inflammation not belonging to multiple sclerosis (MS) or myelin oligodendrocyte glycoprotein (MOG)-/AQP4-associated disorders. TDL concept and clinical features should be revised.

Combination of a solid phase extraction and a two-dimensional LC-UV method for the analysis of vitamin D3 and its isomers in olive oil.

The current HPLC methods for the quantification of vitamin D3 (VitD3) and its two isomers previtamin D3 (PreVitD3) and trans-vitamin D3 (trans-VitD3) in olive oil preparations present some limitations mainly due to peak overlapping of the oily matrix components with the compounds of interest. The use of two-dimensional liquid chromatography (2D-LC) with different retention mechanism can reach higher resolving power thus allowing the analysis of complex samples. The present paper proposes a new alternative method including a solid phase extraction sample preparation step and a two-dimensional liquid chromatographic analysis using routine instrumentation, fitting the needs of quality assurance and quality control laboratories of pharmaceutical companies. The extraction protocol was demonstrated to provide a clean-up of the sample and a quantitative recovery of the species of interest. The 2D method proved its suitability in the isolation of vitamins from oil components in the first dimension and the separation and quantification of the analytes in the second dimension thanks to the orthogonal selectivities of phenyl and porous graphitic carbon (PGC) stationary phases. The method was validated following ICH guidelines and possesses an adequate sensitivity to quantify the impurity trans-VitD3 in pharmaceuticals considering the limits imposed by regulatory agencies. The applicability of the phenyl x PGC 2D-LC-UV method to quality control of medicinal products based on VitD3 in olive oil was confirmed by the successful quantification of vitamins in olive oil formulations.

Systemic Neoplastic Cryoglobulinemic Vasculitis Mimics Large Vessel Occlusion: A Case Report.

INTRODUCTION: We describe a systemic neoplastic cryoglobulinemic vasculitis presenting as a large vessel occlusion (LVO) syndrome. We focus on a rare presentation of a rare condition. CASE REPORT: A 68-year-old man was admitted to the Stroke Unit of Padova with a right middle cerebral artery syndrome. A cerebrovascular event was suspected and protocol for revascularization treatment was performed. Neuroimaging provided no evidence for infarcted tissue or medium-large vascular occlusion but hypothesized a vasculitic involvement of the small vessels of the right hemisphere. Further diagnostics demonstrated a microangiopathic involvement of the heart, kidneys, and lungs. Blood tests showed circulating cryoglobulins and further hematological investigation identified a chronic lymphatic leukemia-like lymphoproliferative disorder. High-dose steroid therapy improved the patient's clinical status and no neurological symptoms remained at discharge. CONCLUSION: We discuss the clinical-radiologic presentation of a small vessel vasculitis that mimics an LVO stroke. This case focuses on the relevance of concomitant multiorgan manifestations in the hyper-acute evaluation of LVO stroke, suggesting the clinical neurologist should consider alternative etiologies as these could provide important clinical implications.

Multi-approach LC-MS methods for the characterization of species-specific attributes of monoclonal antibodies from plants.

In this work, an analytical platform based on the use of chromatography and mass spectrometry (MS), has been applied to the characterization of Rituximab (RTX) obtained from two plant expression systems (rice and tobacco) in comparison to the mammalian cell-derived reference monoclonal antibody (mAb). Different chromatographic approaches, hyphenated to high resolution MS (HRMS), were applied to RTX structural investigation both at middle- and peptide level. In particular, cation exchange chromatography (CEX), size exclusion chromatography (SEC), reversed phase (RPLC) and hydrophilic interaction liquid chromatographic (HILIC) methods were developed and applied on intact mAbs, IdeS-, and trypsin digests in order to address critical attributes such as primary structure, glycan composition, species-related heterogeneity, glycosylation degree, charge variants, aggregation tendency and enzymatic stability. All the collected data highlight the features and criticalities of each production approach. Production in rice results in a heterogeneous but stable product over time, suggesting the absence of proteases in seeds; while tobacco expression system leads to more homogeneous glycosylation, but protein stability seems to be a critical issue probably due to the presence of proteases. This analytical strategy represents a robust support to scientists in the selection and optimization of the best plant expression system to produce recombinant humanized mAbs.

Discovery of a Novel Inhibitor of Human Purine Nucleoside Phosphorylase by a Simple Hydrophilic Interaction Liquid Chromatography Enzymatic Assay.

Human purine nucleoside phosphorylase (HsPNP) belongs to the purine salvage pathway of nucleic acids. Genetic deficiency of this enzyme triggers apoptosis of activated T-cells due to the accumulation of deoxyguanosine triphosphate (dGTP). Therefore, potential chemotherapeutic applications of human PNP inhibitors include the treatment of T-cell leukemia, autoimmune diseases and transplant tissue rejection. In this report, we present the discovery of novel HsPNP inhibitors by coupling experimental and computational tools. A simple, inexpensive, direct and non-radioactive enzymatic assay coupled to hydrophilic interaction liquid chromatography and UV detection (LC-UV using HILIC as elution mode) was developed for screening HsPNP inhibitors. Enzymatic activity was assessed by monitoring the phosphorolysis of inosine (Ino) to hypoxanthine (Hpx) by LC-UV. A small library of 6- and 8-substituted nucleosides was synthesized and screened. The inhibition potency of the most promising compound, 8-aminoinosine (4), was quantified through Ki and IC50 determinations. The effect of HsPNP inhibition was also evaluated in vitro through the study of cytotoxicity on human T-cell leukemia cells (CCRF-CEM). Docking studies were also carried out for the most potent compound, allowing further insights into the inhibitor interaction at the HsPNP active site. This study provides both new tools and a new lead for developing novel HsPNP inhibitors.

Ultrasound findings in urogenital schistosomiasis: a pictorial essay.

Urogenital schistosomiasis is a parasitic disease caused by S. haematobium which is endemic in tropical and sub-tropical areas but is increasingly diagnosed in temperate non-endemic countries due to migration and international travels. Early identification and treatment of the disease are fundamental to avoid associated severe sequelae such as bladder carcinoma, hydronephrosis leading to kidney failure and reproductive complications. Radiologic imaging, especially through ultrasound examination, has a fundamental role in the assessment of organ damage and follow-up after treatment. Imaging findings of urinary tract schistosomiasis are observed mainly in the ureters and bladder. The kidneys usually appear normal until a late stage of the disease.

Costs Associated with Surgically Treated Cases of Abdominal Cystic Echinococcosis: A Single Center's Experience from 2008 to 2014, Pavia, Italy.

Cystic echinococcosis (CE) is a globally distributed zoonosis caused by the Echinococcus granulosus sensu lato species complex. Four approaches are available for treatment of abdominal CE: surgery, percutaneous aspiration, chemotherapy with albendazole, and watch-and-wait. Allocation of patients to these different treatment options mainly depends on the stage of the cystic lesion. However, as available guidelines are not widely followed, surgery is often applied even without the correct indication outside referral centers. This is not only a disadvantage for the patient, but also a waste of money. In this study, we evaluated the cost of the surgical approach for abdominal CE by analyzing hospitalization costs for 14 patients admitted to the General Surgery Ward at the "San Matteo" Hospital Foundation in Pavia, Italy, from 2008 through 2014. We found that the total cost of a single hospitalization, including hospital stay, surgical intervention, personnel, drugs, and administrative costs ranged from €5,874 to 23,077 (median €11,033) per patient. Our findings confirm that surgery can be an expensive option. Therefore, surgical intervention should be limited to cyst types that do not benefit from nonsurgical therapies and appropriate case management can best be accomplished by using a cyst stage-specific approach.

A case of relapsing-remitting tumour-like inflammation of the central nervous system.

The case of a 37-year-old woman suffering from a relapsing-remitting tumefactive inflammatory disease of the central nervous system (CNS) is described. The patient had four severe relapses over eight years, and was treated with steroids, immunosuppression and plasma-exchange with modest benefit. No magnetic resonance imaging or cerebrospinal spinal fluid findings suggestive of multiple sclerosis emerged during the eight-year follow-up. 'Relapsing-remitting tumefactive inflammation' seems to have the features of a distinct inflammatory CNS disease.

Medical treatment versus "Watch and Wait" in the clinical management of CE3b echinococcal cysts of the liver.

BACKGROUND: Available treatments for uncomplicated hepatic cystic echinococcosis (CE) include surgery, medical therapy with albendazole (ABZ), percutaneous interventions and the watch-and-wait (WW) approach. Current guidelines indicate that patients with hepatic CE should be assigned to each option based on cyst stage and size, and patient characteristics. However, treatment indications for transitional CE3b cysts are still uncertain. These cysts are the least responsive to non-surgical treatment and often present as indolent, asymptomatic lesions that may not warrant surgery unless complicated. Evidence supporting indications for treatment of this stage is lacking. In the attempt to fill this gap before the implementation of randomized clinical trials, we compared the clinical behavior of single hepatic CE3b cysts in 60 patients followed at the WHO Collaborating Centre for Cystic Echinococcosis of the University of Pavia. METHODS: We analyzed retrospectively data of 60 patients with hepatic CE3b cysts seen at our clinic over 27 years, who either received ABZ or were monitored with WW. Univariate and multivariate analysis were performed to investigate the effect on outcome (inactivation or relapse) of variables such as age, sex, origin, treatment, cyst size and presence of other echinococcal hepatic cysts using a multiple failure Cox proportional hazard model. RESULTS: ABZ treatment was positively associated with inactivation (p < 0.001), but this was not permanent, and no association was found between therapeutic approach and relapse (p = 0.091). No difference was found in the rate of complications between groups. CONCLUSIONS: In conclusion, our study shows that ABZ treatment induces temporary inactivation of CE3b cysts, while during WW cysts remain stable over time. As the rate of adverse events during periods of ABZ treatment and WW did not differ significantly in the follow-up period considered in this study (median 43 months, IQR 10.7-141.5), expectant management might represent a valuable option for asymptomatic CE3b cysts when strict indication for surgery is absent and patients comply with regular long-term follow-up.

Cystic echinococcosis of the liver: A primer for hepatologists.

Cystic echinococcosis (CE) is a complex, chronic and neglected disease with a worldwide distribution. The liver is the most frequent location of parasitic cysts. In humans, its clinical spectrum ranges from asymptomatic infection to severe, potentially fatal disease. Four approaches exist in the clinical management of CE: surgery, percutaneous techniques and drug treatment for active cysts, and the "watch and wait" approach for inactive cysts. Allocation of patients to these treatments should be based on cyst stage, size and location, available clinical expertise, and comorbidities. However, clinical decision algorithms, efficacy, relapse rates, and costs have never been properly evaluated. This paper reviews recent advances in classification and diagnosis and the currently available evidence for clinical decision-making in cystic echinococcosis of the liver.

Intranasal immunization in mice with non-ionic surfactants vesicles containing HSV immunogens: a preliminary study as possible vaccine against genital herpes.

The purpose of this study was to investigate the potential of intranasal immunization with non-ionic surfactant vesicles (NISV) containing either the secretory recombinant form of glycoprotein B (gBs) of herpes simplex virus type 1 or a related polylysine reach peptides (DTK) for induction of protective immunity against genital herpes infection in mice. NISV were prepared by lipid film hydration method. The mean diameter of vesicles was around 390 nm for DTK-containing NISV (DTK-NISV) and 320 nm for gB1s-containing NISV (gB1s-NISV). The encapsulation efficiency of the molecules was comprised between 57% and 70%. After 7-14 day NISV maintained stable dimensions and a drug encapsulation higher than 48%. We showed that intranasal immunization with gB1s-NISV induces gB-specific IgG antibody and lymphoproliferative responses, whereas vaccination with DTK-NISV was not able to generate a gB-specific immune response. Our results indicate that vaccination of BALB/c mice with gB1s-NISV induced Th1 responses, as characterized by increased titre of IG2a in plasma and IFN-production in CD4+ splenic cells. Intranasal immunization with gB1s-NISV could elicit 90% (almost complete) protection against a heterologous lethal vaginal challenge with herpes simplex virus type 2. These data may have implications for the development of a mucosal vaccine against genital herpes.

Basal ganglia and frontal/parietal cortical atrophy is associated with fatigue in relapsing-remitting multiple sclerosis.

BACKGROUND: Fatigue is one of the most frequent symptoms suffered by patients affected by multiple sclerosis. The patho-physiological basis of multiple sclerosis-related fatigue remains to be elucidated. OBJECTIVE: Our aim was to investigate whether a particular pattern of deep and/or cortical grey matter atrophy is associated with fatigue in patients with multiple sclerosis. METHODS: A total of 152 patients with relapsing-remitting multiple sclerosis were evaluated with the Expanded Disability Status Scale, the Fatigue Severity Status Scale (FSS), the Modified Fatigue Impact Scale and the Beck Depression Inventory. The thalamic and basal ganglia volume and the regional cortical thickness were analysed by means of FreeSurfer. RESULTS: Based on Fatigue Severity Status Scale score, patients were divided into fatigued (FSS ≥ 4, 71 patients, 46.6%) and non-fatigued (FSS < 4, 81 patients, 53.4%). A significant atrophy of striatum, thalamus, superior frontal gyrus and inferior parietal gyrus was observed in fatigued patients compared with non-fatigued patients. The cognitive domain of Modified Fatigue Impact Scale significantly correlated with the volume of the striatum and with the cortical thickness of the posterior parietal cortex and middle frontal gyrus (R = 0.51-0.61), while the physical domain of Modified Fatigue Impact Scale significantly correlated with striatum volume and superior frontal gyrus cortical thickness (R = 0.50-0.54). CONCLUSIONS: The regional analysis of deep and cortical grey matter atrophy suggests an association between the neurodegenerative process taking place in the striatum-thalamus-frontal cortex pathway and the development of fatigue in relapsing-remitting multiple sclerosis. The inclusion of the posterior parietal cortex as one of the best predictors of the Modified Fatigue Impact Scale cognitive domain suggests the major role of the posterior attentional system in determining cognitive fatigue in relapsing-remitting multiple sclerosis.

No evidence of JC virus reactivation in natalizumab treated multiple sclerosis patients: an 18 month follow-up study.

BACKGROUND AND AIM: Natalizumab, used as therapy for multiple sclerosis (MS), has been associated with progressive multifocal leucoencephalopathy (PML), a potentially fatal disease caused by JC virus (JCV), which is not predictable by specific markers. This study evaluated whether JCV reactivation occurred in the urine and/or plasma in 42 MS patients treated with natalizumab over 18 months, and followed by a thorough monitoring programme. METHODS: 42 patients (F/M: 24/18, mean age 34.4±8.9 years) were followed-up by: urine and plasma JCV-DNA PCR assay, immune cell subsets analysis, clinical and MRI evaluation, quality of life, fatigue and mood assessment. RESULTS: JCV data. At baseline, 11/42 (26%) patients had JCV viruria, persistent at serial controls. One patient acquired viruria at month 1 and one patient at month 12. No patient had JCV viraemia at baseline; three patients acquired viraemic (one at month 6, one at month 13 (both transiently) and one at month 12 (persistently viraemic)). The prevalence of JCV in both urine and plasma did not change significantly from baseline to months 12 and 18. No patient had clinical or MRI evidence of PML. Immunological data. Circulating B cells showed greater expansion (300% increase in absolute number) since the first infusion. NK cell count doubled with no change in percentage while T cell count increased with a reduced percentage, reflecting a clear redistribution in the lymphocyte compartment. CD4+ and CD8+ T cells increased proportionally, with no change in their percentage. Clinical data. 27 patients (64%) were disease free after 1 year. A marked improvement in quality of life was reported by 72% of patients. CONCLUSIONS: No evidence of subclinical JCV reactivation was found in our natalizumab treated MS patients up to 18 months of therapy, notwithstanding the marked increase in circulating B cells observed. Moreover, the efficacy of natalizumab, its tolerability and the positive impact on quality of life were confirmed in this study.

Magnetic resonance evidence of cerebellar cortical pathology in multiple sclerosis.

BACKGROUND: Although neuropathological observations suggest that cerebellar cortex is a major site of demyelination in multiple sclerosis (MS), only a few MRI studies on cerebellar cortical pathology in MS are available. OBJECTIVE: To analyse cerebellar cortical volume (CCV) and leucocortical lesions (CL) in MS, and their impact on clinical disability. METHODS: The authors studied 125 patients divided into 38 Clinical Isolated Syndrome (CIS), 35 relapsing remitting multiple sclerosis (RRMS), 27 secondary progressive multiple sclerosis (SPMS) and 25 primary progressive multiple sclerosis, and 32 normal controls (NC). CCV and cerebellar CL number and volume were evaluated by means of Freesurfer software and Double Inversion Recovery, respectively. RESULTS: Compared with NC (mean 113.2 + or - 2.6 cm(3)), the CCV was significantly reduced in CIS (105.4 + or - 2.2 cm(3), p=0.018), RRMS (104.0 + or - 2.0 cm(3), p=0.012), SPMS (98.8 + or - 2.0 cm(3), p<0.001) and PPMS (100.6 + or - 2.2 cm(3), p<0.001), even after age, gender and mean cortical volume correction. CL were observed in all patient groups and were an independent predictor of CCV and cerebellar dysfunction. DISCUSSION: The authors confirm that the cerebellar cortex is severely and early affected by MS pathology. The monitoring of cerebellar cortical atrophy and CL may help to understand the mechanism underlying disability progression in MS.