Publications by authors named "Umberto Basile"

39 Publications

Salivary Biomarkers in COVID-19 Patients: Towards a Wide-Scale Test for Monitoring Disease Activity.

J Pers Med 2021 May 8;11(5). Epub 2021 May 8.

Dipartimento di Scienze di Laboratorio e Infettivologiche, Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Università Cattolica del Sacro Cuore, 00168 Rome, Italy.

The ongoing outbreak of coronavirus disease 2019 (COVID-19), which impairs the functionality of several organs, represents a major threat to human health. One of the hardest challenges in the fight against COVID-19 is the development of wide-scale, effective, and rapid laboratory tests to control disease severity, progression, and possible sudden worsening. Monitoring patients in real-time is highly demanded in this pandemic era when physicians need reliable and quantitative tools to prioritize patients' access to intensive care departments. In this regard, salivary biomarkers are extremely promising, as they allow for the fast and non-invasive collection of specimens and can be repeated multiple times.

Methods: We compare salivary levels of immunoglobulin A subclasses (IgA1 and IgA2) and free light chains (kFLC and λFLC) in a cohort of 29 SARS-CoV-2 patients and 21 healthy subjects.

Results: We found that each biomarker differs significantly between the two groups, with -values ranging from 10 to 10. A Receiving Operator Curve analysis shows that λFLC level is the best-suited candidate to discriminate the two groups (AUC = 0.96), with an accuracy of 0.94 (0.87-1.00 95% CI), a precision of 0.91 (0.81-1.00 95% CI), a sensitivity of 1.00 (0.96-1.00 95% CI), and a specificity of 0.86 (0.70-1.00 95% CI).

Conclusion: These results suggest λFLC as an ideal indicator of patient conditions. This hypothesis is strengthened by the consideration that the λFLC half-life (approximately 6 h) is significantly shorter than the IgA one (21 days), thus confirming the potential of λFLC for effectively monitoring patients' fluctuation in real-time.
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http://dx.doi.org/10.3390/jpm11050385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8151878PMC
May 2021

Serum and urine free light chains measurements in patients with systemic sclerosis: novel biomarkers for disease activity.

Clin Exp Immunol 2021 May 1. Epub 2021 May 1.

Area Diagnostica di Laboratorio, Università Cattolica del Sacro Cuore, Fondazione Policlinico Universitario 'A. Gemelli', IRCCS, Rome, Italy.

Circulating free light chains (FLCs), considered biomarkers of B cell activity, are frequently elevated in patients affected by systemic inflammatory autoimmune diseases. As the systemic sclerosis (SSc) clinical course can be variable, this study is aimed at evaluating FLCs levels in affected individuals as biomarkers of disease activity. We assessed FLC levels in serum and urine of 72 SSc patients and 30 healthy controls (HC). Results were analyzed in comparison with overall clinical and laboratory findings, disease activity index (DAI) and disease severity scale (DSS). SSc patients displayed increased levels of κ and λ FLC in serum significantly higher than HC (p = 0.0001) alongside the mean values of free κ/λ ratio and κ + λ sum (p = 0.0001). SSc patients showed increased free κ in urine with a κ/λ higher than HC (p = 0.0001). SSc patients with increased κ + λ in serum showed that erythro-sedimentation rate (p = 0.034), C-reactive protein (p = 0.003), DAI (p = 0.024) and DSS (p = 0.015) were higher if compared to SSc patients with normal levels of FLC. A positive linear correlation was found between serum levels of free κ and DAI (r = 0.29, p = 0.014). In addition, SSc patients with increased free κ in urine had higher DAI (p = 0.048) than SSc patients with normal κ levels. Our results strengthen the role of serum FLC as useful biomarker in clinical practice to early diagnosis and monitor disease activity, showing for the first time that also urine FLC levels correlated with disease activity in SSc patients.
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http://dx.doi.org/10.1111/cei.13611DOI Listing
May 2021

Hematological and Genetic Markers in the Rational Approach to Patients With HCV Sustained Virological Response With or Without Persisting Cryoglobulinemic Vasculitis.

Hepatology 2021 Mar 15. Epub 2021 Mar 15.

MaSVE Interdepartmental Hepatology Center, Department of Experimental and clinical Medicine, University of Florence, Center for Research and Innovation CRIA-MaSVE, AOU Careggi, Florence, Italy.

Background And Aims: Direct-acting antivirals (DAAs) usually lead to improvement/remission of cryoglobulinemic vasculitis (CV), although symptoms may persist/recur after a sustained virological response (SVR). We evaluated hematological and genetic markers in patients with HCV-SVR vasculitis with and without persisting/recurring symptoms to early predict the CV outcome.

Approach And Results: Ninety-eight patients with HCV-CV were prospectively enrolled after a DAA-induced SVR: Group A: 52 with complete clinical response; Group B: 46 with symptom maintenance/recurrence. Monoclonal B-cell lymphocytosis, t(14;18) translocation, and abnormal free light chains κ/λ ratios were detected by flow cytometry or nested-PCR or nephelometry in 4% Group A versus 17% Group B (P = 0.04) patients, 17% Group A versus 40% Group B patients (P = 0.02), and 17% Group A versus 47% Group B (P = 0.003) patients, respectively. At least 1 out of 3 clonality markers was altered/positive in 29% of Group A versus 70% of Group B patients (P < 0.0001). When available, pretherapy samples were also tested for t(14;18) translocation (detected in 12/37 [32%] Group A and 21/38 [55%] Group B) and κ/λ ratios (abnormal in 5/35 [14%] Group A and 20/38 [53%] Group B) (P = 0.0006), whereas at least one clonality marker was detected/altered in 16/37 (43%) Group A and 30/38 (79%) Group B (P = 0.002). CV-associated single-nucleotide polymorphisms were tested by real-time PCR. Among them, notch4 rs2071286 T minor allele and TT genotype showed a higher frequency in Group B versus Group A (46% vs. 29%, P = 0.01, and 17% vs. 2%, P = 0.006, respectively).

Conclusions: Hematological or genetic analyses could be used to foresee the CV clinical response after DAA therapy and could be valuable to assess a rational flowchart to manage CV during follow-up.
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http://dx.doi.org/10.1002/hep.31804DOI Listing
March 2021

Solving the mystery of HBV-related mixed cryoglobulinemia: potential biomarkers of disease progression.

Rheumatology (Oxford) 2021 02 16. Epub 2021 Feb 16.

Area Diagnostica di Laboratorio, Fondazione Policlinico Universitario "A. Gemelli", I.R.C.C.S, Rome, Italy.

Objectives: The biomarkers of an immunological dysregulation due to a chronic Hepatitis B virus (HBV) infection are indeed understudied. If untreated, this condition may evolve into liver impairment also co-occurred by extrahepatic involvements. Here, we aim to identify a new panel of biomarkers including IgG subclasses, Rheumatoid Factor (RF), and Free Light Chains (FLC) that may result useful and reliable for clinical evaluation of HBV-related cryoglobulinemia.

Methods: We retrospectively analyzed clinical data from 44 HBV positive patients. Patients were stratified on the presence/absence of mixed cryoglobulinemia in two group: 22 with cryoglobulins (CGs) and 22 without CGs. Samples from 20 healthy blood donors (HD) were used as negative controls. Serum samples were tested for IgG subclasses, RF (IgM, IgG, and IgA type), and FLC.

Results: We detected a strikingly different serum IgG subclasses distribution between HD and HBV positive patients together with different RF isotypes; FLC were significantly increased in HBV-patients if compared with HD while no significant difference was shown between HBV with/without MC.

Conclusion: The immune-inflammatory response triggered by HBV may be monitored by a peculiar profile of biomarkers. Our results open new perspective in precision medicine era; in these challenging times, they could be also employed to monitor the clinical course of COVID-19 patients, at high risk of HBV reactivation due to liver impairment and/or immunosuppressive therapies.
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http://dx.doi.org/10.1093/rheumatology/keab157DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7928620PMC
February 2021

Immune dyscrasia in adult growth hormone deficiency: Evaluation of hemolytic complement activity (CH50) and IgG subclasses.

Biomed Pharmacother 2020 Nov 19;131:110757. Epub 2020 Sep 19.

Dipartimento di Medicina e Chirurgia Traslazionale, Università Cattolica del Sacro Cuore, Rome, Italy; Fondazione Policlinico Universitario A. Gemelli IRCCS, Rome, Italy. Electronic address:

CH50 is a screening assay for the activation of the classical complement pathway, the immunoglobulins-mediated one, activated in several inflammatory diseases. Adult growth hormone deficiency (aGHD) is recognized as a chronic inflammatory condition, although poorly evaluated under the profile of inflammatory biomarkers. The aim of this case-control observational study is to analyze CH50 and immunoglobulins G (IgG) subclasses production in aGHD, comparing this condition to healthy controls. 38 subjects were included and divided as follows: aGHD (n = 18, 6 females and 12 males); healthy controls (n = 20, 10 females and 10 males). GHD was diagnosed with dynamic test using Growth Hormone-Releasing Hormone (GHRH 50 μg i.v. + arginine 0,5 g/Kg), with a peak GH response < 9 μg/L when BMI was <30 kg/m or < 4 μg/L when BMI was >30 kg/m. The two groups were evaluated for hormonal and metabolic parameters, CH50 and IgG subtypes. IgG1 and IgG2 were significantly higher in controls than in aGHD, while IgG3 and IgG4 showed a trend to higher levels in controls, although not significant. Furthermore, CH50 levels were significantly higher in aGHD. These data substantiate the hypothesis of a dyscrasia in IgG subclasses production in aGHD. As IgG levels decrease, CH50 levels do not.
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http://dx.doi.org/10.1016/j.biopha.2020.110757DOI Listing
November 2020

Cryoglobulins: putative effectors of adaptive immune response.

Clin Exp Rheumatol 2021 Mar-Apr;39 Suppl 129(2):171-179. Epub 2020 Oct 29.

Department of Translational and Precision Medicine, Sapienza University of Rome, Italy.

Cryoglobulinaemia consists of circulating monoclonal and/or polyclonal immunoglobulins with rheumatoid factor (RF) activity that precipitate at temperatures <37°C. Cryoglobulinaemic syndrome, characterised by clinical signs of systemic vasculitis, is associated with chronic infection of hepatitis C virus (HCV) and might evolve in B-cell malignancies. In about one third of all HCV infection cases, serum autoantibodies are commonly found. This is probably due directly to the transformation of infected B cells but, also, indirectly, to the viral chronic stimulation of a pool of autoreactive B cells. The pattern of IgG subclasses seems to contribute to the worsening progression of HCV infection into lymphoproliferative and/or autoimmune diseases. Many evidences showed that B cells circulating in patients with HCV-associated mixed cryoglobulinaemia (MC) are profoundly abnormal; moreover, in most of cases, normal B cells are replaced by expanded clonal B cells characterized by the low expression of CD21. After viral eradication, these cells persist in circulation and their occurrence does not correlate with serum cryoglobulins nor with vasculitis response or relapse. It is probably due to the persistence of monoclonal B cells producing RF, that in course of MC can be reactivated by circulating immune complexes, highly produced during infections or tumours. Here, we aimed to review current literature focusing the pathogenesis of MC referring to specificity and immunochemical characteristics of the immunoglobulins involved in cryoprecipitation.
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May 2021

COVID-19 and hepatic involvement: The liver as a main actor of the pandemic novel.

Scand J Immunol 2021 Mar 26;93(3):e12977. Epub 2020 Sep 26.

Dipartimento di Scienze di Laboratorio e Infettivologiche, Fondazione Policlinico Universitario "A. Gemelli" I.R.C.C.S, Rome, Italy.

In the natural history of SARS-CoV-2 infection, liver injury is frequent but quite mild and it is defined as any liver damage occurring during disease progression and treatment of infection in patients with or without pre-existing liver diseases. The underlying mechanisms for hepatic injury in patients with COVID-19 are still unclear but the liver damage in SARS-CoV-2 infection seems to be directly caused by virus-induced cytopathic effects. In this review, we will summarize all data of updated literature, regarding the relationship between SARS-CoV-2 infection, acute response and liver involvement. An overview will be given on liver injury, liver transplant and the possible consequences of COVID-19 in patients with pre-existing liver diseases.
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http://dx.doi.org/10.1111/sji.12977DOI Listing
March 2021

Immunological Role of IgG Subclasses.

Immunol Invest 2021 May 11;50(4):427-444. Epub 2020 Jun 11.

Area Diagnostica di Laboratorio, Fondazione Policlinico Universitario "A. Gemelli", Rome, Italy.

The loss of tolerance to self-antigens is the unequivocal "red line" of autoimmunity: both development of autoreactive T and B cells and production of polyclonal autoantibodies represent seminal keys to the pathogenesis of protean autoimmune diseases. Most of these autoantibodies are immunoglobulins G (IgG), functionally distinguished in four subclasses named IgG1, IgG2, IgG3, and IgG4, due to structural differences in the hinge and heavy chain constant regions. Different studies analyzed serum levels of IgG subclasses in the course of different disorders, showing that they might have a pathogenic role by regulating interactions among immunoglobulins, Fc-gamma receptors, and complement. To date, the mechanisms promoting different IgG subclasses distribution during the natural history of most autoimmune diseases remain somewhat unclear. Evidence from the medical literature shows that the serum IgG profile is peculiar for many autoimmune diseases, suggesting that different subclasses could be specific for the underlying driving autoantigens. A better knowledge of IgG subsets may probably help to elucidate their pathological task, but also to define their relevance for diagnostic purposes, patients' personalized management, and prognosis assessment.
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http://dx.doi.org/10.1080/08820139.2020.1775643DOI Listing
May 2021

Long-Lasting Rituximab-Induced Reduction of Specific-But Not Total-IgG4 in MuSK-Positive Myasthenia Gravis.

Front Immunol 2020 5;11:613. Epub 2020 May 5.

Istituto di Neurologia, Università Cattolica del Sacro Cuore, Rome, Italy.

The use of rituximab (RTX), an anti-CD20 monoclonal antibody (Ab), in refractory myasthenia gravis (MG) is associated with a better response in patients with Abs to the muscle-specific tyrosine kinase (MuSK) than in other MG subgroups. Anti-MuSK Abs are mostly IgG4 with proven pathogenicity and positive correlation with clinical severity. The rapid and sustained response to RTX may be related to MuSK Ab production by short-lived Ab-secreting cells derived from specific CD20 B cells. Here, we investigated the long-term effects of RTX in nine refractory MuSK-MG patients with a follow-up ranging from 17 months to 13 years. In patients' sera, we titrated MuSK-specific IgG (MuSK-IgG) and MuSK-IgG4, along with total IgG and IgG4 levels. Optimal response to RTX was defined as the achievement and maintenance of the status of minimal manifestations (MM)-or-better together with ≥ 50% steroid reduction, withdrawal of immunosuppressants, and no need for plasma-exchange or intravenous immunoglobulin. After a course of RTX, eight patients improved, with optimal response in six, while only one patient did not respond. At baseline, MuSK-IgG and MuSK-IgG4 serum titers were positive in all patients, ranging from 2.15 to 49.5 nmol/L and from 0.33 to 46.2 nmol/L, respectively. MuSK Abs mostly consisted of IgG4 (range 63.80-98.86%). RTX administration was followed by a marked reduction of MuSK Abs at 2-7 months and at 12-30 months ( < 0.02 for MuSK-IgG and < 0.01 for MuSK-IgG4). In patients with a longer follow-up, MuSK Ab titers remained suppressed, paralleling clinical response. In the patient who achieved long-term complete remission, MuSK-IgG4 was no longer detectable within 2 years, while MuSK-IgG remained positive at very low titers up to 10 years after RTX. In the patient who did not respond, MuSK-IgG and MuSK-IgG4 remained unchanged. In this patient series, total IgG and IgG4 transiently decreased ( < 0.05) at 2-7 months after RTX. The different trends of reduction between MuSK-IgG4 and total IgG4 after RTX support the view that short-lived Ab-secreting cells are the main producers of MuSK Abs. The ratio between short-lived Ab-secreting cells and long-lived plasma cells may influence the response to RTX, and B-cell severe depletion may reduce self-maintaining autoimmune reactivity.
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http://dx.doi.org/10.3389/fimmu.2020.00613DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7214629PMC
March 2021

Biomarkers of minimal residual disease in rituximab-treated patients with mixed cryoglobulinemia.

Biotechnol Appl Biochem 2021 Apr 17;68(2):319-329. Epub 2020 May 17.

Department of Translational and Precision Medicine, Sapienza University of Rome, Rome, Italy.

Hepatitis C virus (HCV) represents the major risk factor for mixed cryoglobulinemia (MC), a small-vessel vasculitis that may evolve into an overt B-cell non-Hodgkin's lymphoma. Here, we aimed to identify a biomarker signature for the early diagnosis of minimal residual disease (MRD). We assessed free light chains (FLCs), IgM k,and IgM λ heavy/light chain (HLC) pairs, and vascular endothelial growth factor (VEGF) in sera from 34 patients with MC vasculitis (32 HCV- and 2 HBV-related), treated with low-dose rituximab (RTX). FLCs and IgM HLCs were measured by turbidimetric assay; VEGF by an enzyme-linked immunosorbent assay. After RTX, the positive (complete + partial) clinical and laboratory responses were of 85.29% and 50%, respectively; in contrast, the mean levels of FLCs, IgM HLCs, and VEGF were substantially unaffected in most patients and still above the normal range. In those achieving a reduction of FLCs and IgM k and λ chains values within the range of normality, we found that post-treatment free λ chains and IgM k values correlated with clinical and laboratory response. Our results suggest that high levels of FLCs, IgM HLCs, and VEGF could represent the signature of "dormant" B cell clones' activity that could be very useful to identify MRD indicative of possible relapse or worsening outcome.
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http://dx.doi.org/10.1002/bab.1929DOI Listing
April 2021

The prognostic impact of monoclonal immune globulin and free light chain secretion in diffuse large B cell lymphoma (DLBCL).

Leuk Lymphoma 2020 05 31;61(5):1133-1139. Epub 2019 Dec 31.

Area di Ematologia, Fondazione Policlinico Universitario A. Gemelli, IRCCS, Rome, Italy.

We analyzed the prognostic impact of levels of free light chains (FLC) in 106 patients with DLBCL, selecting 61 patients with a monoclonal (M) protein in serum, and 45 patients without a M protein as an IPI-matched control group. Patients with a M protein had higher levels of FLC, but these were not of prognostic significance in this group. The presence of a M protein nullified associations of κ-FLC with several laboratory parameters indicating immune system activation observed in patients without a M protein. Patients without M protein and κ-FLC >50 mg/L had a significant inferior event-free survival ( = .004). The presence of M protein of an IgM type at diagnosis was a negative outcome predictor ( = .008), while a non-IgM M protein did not significantly impact on prognosis. In conclusion, the prognostic performance of the FLC assay is altered by the presence of a M protein.
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http://dx.doi.org/10.1080/10428194.2019.1706731DOI Listing
May 2020

Sentinel biomarkers in HCV positive patients with mixed cryoglobulinemia.

J Immunol Methods 2020 01 25;476:112687. Epub 2019 Oct 25.

Dipartimento di Medicina Interna e Gastroenterologia, Fondazione Policlinico Universitario "A. Gemelli" - I.R.C.C.S, Università Cattolica del Sacro Cuore, Rome, Italy.

Background: Infections, autoimmunity and cancer play a role as determinants of etiology in Hepatitis C virus (HCV) related mixed cryoglobulinemia (MC). Several factors of risk have been suggested as markers of pathogenesis and progression of HCV-related MC into B cell Non-Hodgkin's Lymphoma (B-NHL). Here, we evaluated IgG subclass distribution, free light chains (FLCs) and vascular endothelial growth factor (VEGF) as a new combination of biomarkers.

Methods: We measured IgG1-4 subclasses, FLCs and VEGF levels in sera 53 from HCV-related MC, in comparison with 40 sera from HCV negative patients with rheumatoid arthritis (RA) and 30 from healthy blood donors (HBD).

Results: IgG3 levels were significantly higher in HCV-MC patients with a decrement of IgG2 and IgG4; FLC levels significantly increased in both MC and RA patients' groups; serological VEGF was higher in HCV-MC patients than in HBD in correlation with k and λ levels.

Conclusion: Our results suggest that a specific IgG subclasses pattern together with raised levels of FLCs and VEGF could represent the biomarker "signature" of an inflammation multistage of acquired immune system.
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http://dx.doi.org/10.1016/j.jim.2019.112687DOI Listing
January 2020

The Laboratory Role in anti-TNF Biological Therapy Era.

Immunol Invest 2020 Apr 12;49(3):317-332. Epub 2019 Jul 12.

Istituto di Patologia Generale - Fondazione Policlinico Universitario Agostino Gemelli- IRCCS, Università Cattolica del Sacro Cuore, Rome, Italy.

Along years, the advent of biological therapy widely modified treatment of rheumatic diseases and other disorders. However, many agents may elicit in anti-drug antibodies (ADAbs) upon consecutive infusions, with a loss of response. For the right strategy of a personalized medicine, the therapeutic monitoring of TNF-α inhibitors and ADAbs represents an important effort in diagnostic-therapeutic pathway, to improve overall patient management and favoring an appropriate clinical approach. A raising number of diagnostic tests have been designed to elucidate the efficacy and/or safety of a specific drug or class of drugs for a targeted patient's group. Our paper reviewed the current understanding of the immunogenicity of biological drugs employed in the treatment of inflammatory diseases underlying the laboratory role.
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http://dx.doi.org/10.1080/08820139.2019.1637434DOI Listing
April 2020

Aggressive Light Chain Myeloma Originating a Double Peak on Serum Electrophoresis: What's Underneath?

Acta Haematol 2020 3;143(1):91-92. Epub 2019 Jul 3.

Istituto di Ematologia, Università Cattolica del Sacro Cuore, Rome, Italy.

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http://dx.doi.org/10.1159/000500873DOI Listing
April 2020

Free light chains and autoimmunity.

Autoimmun Rev 2019 May 4;18(5):484-492. Epub 2019 Mar 4.

Area Diagnostica di Laboratorio, Fondazione Policlinico Universitario A. Gemelli IRCCS, Rome, Italy.

The study of free light chains (FLCs) has grown as area of enormous interest for many clinicians with the aim of disclosing the exact biological role and potential use of FLCs in the clinical routine. Moreover, the attention given to immunological functions of FLCs has sparked a new light into their pathogenic contribution in different chronic autoimmune-based inflammatory diseases. The release of intracellular antigens following cell death or ineffective clearance of apoptotic debris, modification of self-antigens, and molecular mimicry may trigger the production of immunoglobulins after activation and polyclonal expansion of B cells, by which FLCs are released. The discovery of polyclonal FLCs as potential biomarkers started with the observation of their increased concentrations in a variety of biological fluids related to patients with autoimmune diseases. This review deals with the use of polyclonal FLCs for identifying severity and monitoring outcome after treatment in some autoimmune diseases, namely systemic lupus erythematosus, myasthenia gravis, systemic sclerosis, rheumatoid arthritis and Sjögren's syndrome, as supported by the fact that levels of FLCs correlate with both B cell activation markers and other specific markers of disease activity. In a near future, following the evidence shown, FLCs might probably work as early prognostic markers of severity and also as indicators of response to treatment or early assessment of relapse in selected autoimmune diseases.
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http://dx.doi.org/10.1016/j.autrev.2019.03.003DOI Listing
May 2019

Plasmatic free light chains in polycystic ovary syndrome.

Gynecol Endocrinol 2019 Aug 5;35(8):710-713. Epub 2019 Mar 5.

e Area di Medicina di Laboratorio , Fondazione Policlinico Universitario A. Gemelli IRCCS, Università Cattolica del Sacro Cuore , Rome , Italy.

Polycystic ovary syndrome (PCOS), as systemic disease, is accompanied by different indexes of inflammation. Free light chains of immunoglobulins (FLCs), produced by plasmacells, are released in slight excess for the immune requests, with still poorly defined physiological role but surely they represent a marker of inflammation. In order to evaluate their levels and correlate them with hyperandrogenism, we have studied a group of PCOS patients, age range 18-37 yrs, mean ± SEM body mass index (BMI) 24.1 ± 0.9 kg/m), compared with age- and BMI-matched controls, with assay of and FLCs, by turbidimetric method, and their ratio in blood plasma. PCOs exhibited higher levels vs. controls: (mean ± SEM : 10.0 ± 0.85 mg/L vs. 8.41 ± 0.45 mg/L; : 12.45 ± 0.72 mg/L vs. 6.41 ± 0.34 mg/L; /: 1.31 ± 0.07 vs. 0.78 ± 0.04). A significant direct correlation was observed between -FLCs and testosterone levels, no correlation was indeed found with HOMA-IR index. These data confirm high levels of FLCs in PCOS, suggesting systemic inflammatory state and a possible role in the pathophysiology of such complex syndrome.
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http://dx.doi.org/10.1080/09513590.2019.1581759DOI Listing
August 2019

Serological profile of asymptomatic HCV positive patients with low level of cryoglobulins.

Biofactors 2019 May 18;45(3):318-325. Epub 2018 Dec 18.

Dipartimento di Medicina Interna e Gastroenterologia, Fondazione Policlinico Universitario Agostino Gemelli, Università Cattolica del Sacro Cuore, Rome, Italy.

Clinical spectrum of hepatitis C virus (HCV)-related cryoglobulinemia varies from an asymptomatic presentation to severe vasculitis and lymphoma. A recent study in HCV-negative patients suggests that low cryoglobulins (CGs) levels are responsible for severe renal and neurological complications. The aim of this study was to identify a panel of serological biomarkers associated with low levels of CGs in HCV-positive patients. We studied a population of 79 untreated patients with chronic HCV infection: 13 naïve patients without CGs; 28 patients with asymptomatic mixed cryoglobulinemia (MC) and low levels of CGs (16/28 with polyclonal type III and 12/28 with microheterogeneous type III CGs); 38 patients with symptomatic MC and high levels of type II CGs. Serum samples were collected and examined for rheumatoid factor (RF) IgG and IgM, free light chains (FLCs) and C3 and C4 complement components. We found that RF-IgG and IgM, free k chains and k+λ were increased while C4 component was reduced, both in symptomatic and asymptomatic patients. Our results suggest that, even in absence of MC symptoms, the low levels of CGs may represent a trigger of activation for immune system in course of HCV infection. The identification of a correlated biomarkers panel could improve the clinical management of these patients and pave the way for target treatment strategies. © 2018 BioFactors, 45(3):318-325, 2019.
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http://dx.doi.org/10.1002/biof.1485DOI Listing
May 2019

Plasmatic free light chains as inflammatory marker in insulin resistance: comparison of metabolic syndrome with adult growth hormone deficiency.

Biofactors 2018 Sep 3;44(5):480-484. Epub 2018 Sep 3.

Operative Unit of Endocrinology, Fondazione Policlinico Universitario A. Gemelli IRCCS - Università Cattolica del Sacro Cuore, Rome, Italy.

Biological functions of immunoglobulin-free light chains (FLCs), other than in chronic inflammatory diseases, are still poorly defined; the field of insulin resistance (IR) has not been investigated, despite the strict relationships with oxidative stress (OS) and inflammation. Therefore, we evaluated FLCs levels and their relationships with metabolic parameters in adult growth hormone deficiency (GHD) and metabolic syndrome (MetS), both characterized by IR. One hundred subjects were enrolled: group A, patients with GHD [n =31, 24-69 years, mean ± SEM body mass index (BMI) 26.8 ± 1.5 kg/m ]; group B, patients with MetS (n = 29, 21-70 years, BMI 31.9 ± 1.3); group C, controls (N = 40, 21-62 years, BMI 21.6 ± 1.1). Groups were matched by age range and, for patients, by BMI. Morning blood sample was collected for metabolic parameters and FLCs, assessed by turbidimetric assay. GHD patients show levels of FLCs significantly higher than MetS and controls (mean ± SEM κ 37.21 ± 6.97, 15.27 ± 0.86, 12.34 ± 0.85 mg/l; λ 19.44 ± 2.61, 11.78 ± 0.72 and 11.67 ± 0.77 mg/l; κ/λ ratio 1.77 ± 0.13, 1.38 ± 0.09; and 1.10 ± 0.06, respectively); only κ were higher in MetS versus controls. Therefore, the ratio showed progressive declining values in GHD versus MetS versus controls. Our data show increased FLCs levels in GHD and MetS, with the highest values in the former. Both conditions show OS, but with different molecular patterns. FLCs may contribute to chronic inflammation, leading to OS, and cardiovascular complications of GHD. Prognostic and therapeutic implications require further investigation. © 2018 BioFactors, 44(5):480-484, 2018.
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http://dx.doi.org/10.1002/biof.1444DOI Listing
September 2018

Serological Immunoglobulin-Free Light Chain Profile in Myasthenia Gravis Patients.

J Immunol Res 2018 25;2018:9646209. Epub 2018 Mar 25.

Dipartimento di Medicina di Laboratorio, Fondazione Policlinico Universitario Agostino Gemelli, Università Cattolica del Sacro Cuore, Rome, Italy.

Background: Serological levels of free immunoglobulin light chains (FLCs), produced in excess of heavy chains during synthesis of immunoglobulins by plasma cells, can be considered a direct marker of B cell activity in different systemic inflammatory-autoimmune conditions and may represent a useful predictor of rituximab (RTX) therapeutic efficacy, as reported for rheumatoid arthritis and systemic lupus erythematosus. Myasthenia gravis (MG) is an autoimmune disease of the neuromuscular junction with antibodies (abs) targeting the acetylcholine receptor (AChR) or the muscle-specific tyrosine kinase (MuSK), inducing muscle weakness and excessive fatigability. As MG course may be remarkably variable, we evaluated the possible use of FLCs as biomarkers of disease activity.

Subjects And Methods: We assessed FLC levels in 34 sera from 17 AChR-MG and from 13 MuSK-MG patients, in comparison with 20 sera from patients with systemic autoimmune rheumatic diseases and 18 from healthy blood donors, along with titers of specific auto-abs and IgG subclass distribution.

Results: We found a statistically significant increase in free chains in both AChR- and MuSK-MG patients, while free chain levels were increased only in AChR-MG. We also observed a significant reduction of both free and chains in 1/4 MuSK-MG patients along with specific abs titer, two months after RTX treatment.

Conclusions: From our data, FLCs appear to be a sensitive marker of B cell activation in MG. Further investigations are necessary to exploit their potential as reliable biomarkers of disease activity.
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http://dx.doi.org/10.1155/2018/9646209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5889870PMC
September 2018

Different biochemical patterns in type II and type III mixed cryoglobulinemia in HCV positive patients.

Dig Liver Dis 2018 Sep 4;50(9):938-943. Epub 2018 Apr 4.

Institute of Internal Medicine, Fondazione Policlinico Universitario Agostino Gemelli - Università Cattolica del Sacro Cuore, Rome, Italy.

Background: Reversible cryoprecipitability of proteins is observed as a concomitant feature of immune complex formation. Mixed cryoglobulinemia (MC) is systemic vasculitis, associated with mixed IgM and IgG cryoglobulins (CGs) showing rheumatoid factor (RF) activity. It is frequently associated with hepatitis C virus (HCV). This study investigates the presence of IgG RF and anti-nuclear antibodies (ANA) in cryoprecipitates of patients with type III and type II MC, to understand the biochemical patterns associated with different types of MC to a greater degree.

Methods: Sera from 70 HCV untreated patients with type III or type II MC were tested by immunofixation for IgG3 and through ELISA for IgG RF. Cryoprecipitates were analysed for ANA by indirect immunofluorescence to identify specific patterns.

Results: After stratification according to MC type, the ANA patterns between type II and type III MC were statistically different. IgG3 levels and IgG-RF positivity were significantly higher in type III cryoprecipitate. We observed a higher positivity of IgG3 and a significant difference between the liver fibrosis stage, ANA and IgG-RF in the cryoprecipitate.

Conclusion: Results show a combination of biochemical markers and autoantibodies associated to mixed cryoglobulinemia; these findings could be further investigated in order to ascertain their usefulness in assessing the risk for the development of mixed cryoglobulinemia.
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http://dx.doi.org/10.1016/j.dld.2018.03.028DOI Listing
September 2018

Free light chains of immunoglobulins in patients with systemic sclerosis: correlations with lung involvement and inflammatory milieu.

J Clin Pathol 2018 Jul 31;71(7):620-625. Epub 2018 Jan 31.

Institute of Rheumatology and Affine Sciences, Department of Rheumatology, Fondazione Policlinico Universitario Agostino Gemelli, School of Medicine, Catholic University of the Sacred Heart, Rome, Italy.

Aim: Humoral immunity and B cells are thought to play an important role in the pathophysiology of the systemic sclerosis (SSc). The production of free light chains (FLC) of immunoglobulins is abnormally high in several pathological autoimmune conditions and reflects B cell activation. Furthermore, FLCs demonstrated different biological activities including their capability to modulate the immune system, proteolytic activity and complement cascade activation. The aims of this study are to determine the FLC levels in patients with SSc compared with healthy controls (HC) and to study their possible association with organ involvement and disease characteristics.

Methods: Sixty-five patients with SSc and 20 HC were studied. Clinical and immunological inflammatory characteristics were assessed for all the patients with SSc. κ-FLC and λ-FLC, interleukin 6 (IL-6) and B cell activating factor levels were measured.

Results: The mean serum κ-FLC levels and FLC ratio were significantly higher in patients with SSc compared with HC, while the serum λ-FLC levels were comparable.The levels of FLC were comparable in patients with diffuse skin disease and limited skin involvement, while κ-FLC levels were increased in patients with restrictive lung (forced vital capacity (FVC) <80%) disease (26.4±7.4 mg/L) when compared with patients with FVC ≥80% (19.6±7.3 mg/L, P=0.009). In patients with SSc, the levels of serum κ-FLC level directly correlated with the IL-6 levels (R=0.3, P=0.001) and disease activity (R=0.4, P=0.003).

Conclusions: FLC levels are elevated in SSc and high levels are associated with lung involvement and with a higher degree of inflammation, supporting a possible role of B cell activation in the pathophysiology of the disease.
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http://dx.doi.org/10.1136/jclinpath-2017-204656DOI Listing
July 2018

Vitamin D deficiency and supplementation in patients with aggressive B-cell lymphomas treated with immunochemotherapy.

Cancer Med 2018 01 22;7(1):270-281. Epub 2017 Dec 22.

Institute of Hematology, Università Cattolica del Sacro Cuore, Rome, Italy.

Vitamin D deficiency has been reported to be a negative prognostic factor in elderly patients with aggressive B-cell lymphomas. In vitro data suggest that vitamin D supplementation may enhance rituximab-mediated cytotoxicity. We prospectively assessed 25-hydroxyvitamin D [25(OH)D] levels at diagnosis in a cohort of 155 patients with aggressive B-cell lymphomas of whom 128 had diffuse large B-cell lymphoma (DLBCL) not otherwise specified. 25(OH)D levels were deficient (<20 ng/mL) in 105 (67%), insufficient (20-29 ng/mL) in 32 (21%), and normal (≥30 ng/mL) in 18 (12%) patients with a seasonal variation. Patient characteristics associated with lower 25(OH)D levels were poor performance status, overweight, B-symptoms, elevated LDH, lower albumin and hemoglobin levels. As a result of a change in practice pattern, 116 patients received vitamin D3 (cholecalciferol) supplementation that included a loading phase with daily replacement and subsequent maintenance phase with a weekly dose of 25,000 IU until end of treatment. This resulted in a significant increase in 25(OH)D levels, with normalization in 56% of patients. We analyzed the impact of 25(OH)D levels on event-free survival in patients treated with Rituximab-CHOP. 25(OH)D levels below 20 ng/mL at diagnosis and IPI were independently associated with inferior EFS. Moreover, patients with normalized 25(OH)D levels following supplementation showed better EFS than patients with persistently deficient/insufficient 25(OH)D levels. Our study provides the first evidence that achievement of normal 25(OH)D levels after vitamin D3 supplementation is associated with improved outcome in patients with DLBCL and deficient/insufficient 25(OH)D levels when receiving rituximab-based treatment.
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http://dx.doi.org/10.1002/cam4.1166DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5773978PMC
January 2018

Free light chains: Eclectic multipurpose biomarker.

J Immunol Methods 2017 12 18;451:11-19. Epub 2017 Sep 18.

Center for Systemic Manifestations of Hepatitis Viruses (MaSVE), Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.

The production of antibodies is accompanied by a slight excess of synthesis of κ and λ immunoglobulin light chains; small amounts of them are released in the peripheral blood and can also be found in various body fluids, such as synovial fluid, cerebrospinal fluid, urine and saliva. They are rapidly filtered by the glomerulus and >99% are reabsorbed from the cells of the proximal convoluted tubule, making them present in the urine in only trace amounts. The production of an excess of protein without a reason or a specific function in a biological system is rare. Free light chains, considered for years a waste product of Ig synthesis, are currently known to be very active molecules, able to bind antigens as well as whole immunoglobulin and helping to develop specific antibody affinity. The ability of free light chains to activate mast cells and then become an active part of the pathogenic mechanisms of chronic inflammatory diseases has increased interest in their clinical use, both as an attractive therapeutic target or as a biochemical marker of disease evolution or remission. This is an overview of relevant scientific interest that immunoglobulin light chains κ and λ have attracted over the years, a report on the progress in knowledge about their structure and function, with a special focus on their biological meaning and potential clinical utility in different diseases.
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http://dx.doi.org/10.1016/j.jim.2017.09.005DOI Listing
December 2017

Enhanced liver fibrosis test as a reliable tool for assessing fibrosis in nonalcoholic fatty liver disease in a clinical setting.

Int J Biol Markers 2017 Oct 31;32(4):e397-e402. Epub 2017 Oct 31.

Department of Internal Medicine and Gastroenterology, Gemelli University Hospital, Catholic University of Sacred Heart, Rome - Italy.

Background: Liver fibrosis is the main determinant and predictor of the clinical course of nonalcoholic fatty liver disease (NAFLD). To date, a liver biopsy is still considered the gold standard for staging fibrosis. The aim of this study was to investigate the diagnostic accuracy of the commercial enhanced liver fibrosis (ELF) test manufacturer's cutoff value (≥9.8) in identifying severe fibrosis for adult patients with histologically confirmed NAFLD.

Methods: We tested the ELF test in a clinical practice, prospective cohort of 82 consecutive patients who consecutively underwent percutaneous liver biopsy.

Results: All stages of liver fibrosis were represented in our cohort, and severe fibrosis was present in 15 of 82 patients (18.3%). The stage of fibrosis was significantly associated with ELF score (Spearman's rho = 0.483, p<0.001). The commercial ELF test manufacturer's cutoff identified severe fibrosis with good sensitivity (86.7%; 95% confidence interval [95% CI], 0.69-1.04) and high specificity (92.5%; 95% CI, 0.86-0.99), with a positive predictive value of 72% and negative predictive value of 97%.

Conclusions: Our data could support the use of the ELF test in clinical practice.
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http://dx.doi.org/10.5301/ijbm.5000292DOI Listing
October 2017

IgG3 subclass: A possible trigger of mixed cryoglobulin cascade in hepatitis C virus chronic infection.

Dig Liver Dis 2017 Nov 24;49(11):1233-1239. Epub 2017 Jun 24.

Institute of Internal Medicine, Catholic University of the Sacred Heart, Largo A. Gemelli, 00168 Rome, Italy.

HCV is a hepatotropic and lymphotropic virus and is the most frequent cause of "benign" mono-oligoclonal B-lymphocyte proliferation, observed in mixed cryoglobulinemia (MC). The study aims to investigate the presence, prevalence and characteristics of the subclasses of cryoglobulins in HCV-patients to look for a relationship with MC. Fifty HCV-infected patients with cryoglobulins were enrolled. IgG subclasses were characterized in cryoprecipitate, and serum IgG and IgM Rheumatoid Factor (RF) were determined. Patients were stratified into two subgroups according to the presence of IgG3 subclass. Differences were observed in supernatant IgM, IgG3-positive and IgG3-negative patients with a higher IgM concentration in the IgG3-negative cohort (p=0.03). Higher IgM-RF was detected in the IgG3-negative group (p=0.01). IgG3-positive group showed higher IgG-RF compared to the IgG3-negative group (p<0.0001). IgG3-negative/monoclonal-IgM patients had higher cryocrit compared to IgG3-negative/polyclonal-IgM patients (p<0.01). C4 levels were higher in the polyclonal-IgM group compared to monoclonal-IgM group (p<0.01). We speculate that cryoglobulins are part of a progressive clonal selection process in which, B-cells are stimulated to produce oligoclonal IgG3 with RF activity. The persistence of the antigenic stimulus elicits the production of polyclonal IgM-RF and subsequently the formation of oligoclonal IgG/polyclonal IgM containing cryoglobulins. In the last stage, a monoclonal IgM-RF clone is formed which may coexist with a monoclonal IgG3-RF clone.
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http://dx.doi.org/10.1016/j.dld.2017.06.003DOI Listing
November 2017

Overview of immune abnormalities in lysosomal storage disorders.

Immunol Lett 2017 08 4;188:79-85. Epub 2017 Jul 4.

Centro Italiano Macula, Rome, Italy.

The critical relevance of the lysosomal compartment for normal cellular function can be proved by numbering the clinical phenotypes that arise in lysosomal storage disorders (LSDs), a group of around 70 different monogenic autosomal or X-linked syndromes, caused by specific lysosomal enzyme deficiencies: all LSDs are characterized by progressive accumulation of heterogeneous biologic materials in the lysosomes of various parts of the body such as viscera, skeleton, skin, heart, and central nervous system. At least a fraction of LSDs has been associated with mixed abnormalities involving the immune system, while some patients with LSDs may result more prone to autoimmune phenomena. A large production of proinflammatory cytokines has been observed in Gaucher and Fabry diseases, and wide different autoantibody production has been also reported in both. Many immune-mediated reactions are crucial to the pathogenesis of different inflammatory signs in mucopolysaccharidoses, and subverted heparan sulphate catabolism might dysregulate cellular homeostasis in the brain of these patients. Furthermore, an inappropriate activation of microglia is implicated in the neurodegenerative foci of Niemann-Pick disease, in which abnormal signalling pathways are activated by impaired sphingolipid metabolism. In addition, not the simple impaired catabolism of gangliosides per se, but also the production of anti-ganglioside autoantibodies contributes to the neurological disease of gangliosidoses. Even if the exact relationship between the modification of lysosomal activities and modulation of the immune system remains obscure, there is emerging evidence of different impaired immunity responses in a variety of LSDs: in this review we investigate and summarize the immune abnormalities and/or clinical data about immune system irregularities which have been described in a subset of LSDs.
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http://dx.doi.org/10.1016/j.imlet.2017.07.004DOI Listing
August 2017

Serum free light chain quantitative assays: Dilemma of a biomarker.

J Clin Lab Anal 2018 Feb 26;32(2). Epub 2017 Apr 26.

Department of Laboratory Medicine, Catholic University of the Sacred Heart, Rome, Italy.

Background: Serum free light chains detection assays are consistently meeting greater interest for the diagnosis and monitoring of monoclonal gammopathies and plasma cell dyscrasias. Nowadays, there are neither standardized methods nor reference material for the determination of free light chains; for this reason, it is important to compare two different assays used in clinical laboratory.

Methods: We evaluated 300 serum samples from patients with B-cell disorders and compared the analytical performances of both assay. Each test was assayed on both testing platforms (Siemens Dade Behring BN II Nephelometer and SPAPLUS by The Binding Site). κ/λ ratios were determined and compared. Results were analyzed by Passing-Bablok and Bland-Altman plots to evaluate comparability of the two techniques and to determine bias.

Results: The reproducibility of both assays is acceptable, reaching minimum and desirable analytical goals derived from biological variability. However, values are not interchangeable between systems. This study shows that the two systems do not allow results to be transferred from one method to the other even if they display good agreement.

Conclusion: Our study highlights the importance of elaborating an international standard for free light chains quantification in order to offer homogeneous results as well as guarantee harmonization of values among laboratories. Moreover, the assays should be validated in specific patient groups to determine that they are clinically fit for purpose.
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http://dx.doi.org/10.1002/jcla.22243DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6817218PMC
February 2018

Cryoglobulin Test and Cryoglobulinemia Hepatitis C-Virus Related.

Mediterr J Hematol Infect Dis 2017 1;9(1):e2017007. Epub 2017 Jan 1.

Department of Laboratory Medicine, Catholic University of the Sacred Heart, Rome, Italy.

Cryoglobulins are immunoglobulins that precipitate in serum at temperatures below 37°C and resolubilize upon warming. The clinical syndrome of cryoglobulinemia usually includes purpura, weakness, and arthralgia, but the underlying disease may also contribute other symptoms. Blood samples for cryoglobulin are collected, transported, clotted and spun at 37°C, before the precipitate is allowed to form when serum is stored at 4°C in a Wintrobe tube for at least seven days. The most critical and confounding factor affecting the cryoglobulin test is when the preanalytical phase is not fully completed at 37°C. The easiest way to quantify cryoglobulins is the cryocrit estimate. However, this approach has low accuracy and sensitivity. Furthermore, the precipitate should be resolubilized by warming to confirm that it is truly formed of cryoglobulins. The characterization of cryoglobulins requires the precipitate is several times washed, before performing immunofixation, a technique by which cryoglobulins can be classified depending on the characteristics of the detected immunoglobulins. These features imply a pathogenic role of these molecules which are consequently associated with a wide range of symptoms and manifestations. According to the Brouet classification, Cryoglobulins are grouped into three types by the immunochemical properties of immunoglobulins in the cryoprecipitate. The aim of this paper is to review the major aspects of cryoglobulinemia and the laboratory techniques used to detect and characterize cryoglobulins, taking into consideration the presence and consequences of cryoglobulinemia in Hepatitis C Virus (HCV) infection.
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http://dx.doi.org/10.4084/MJHID.2017.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5224812PMC
January 2017