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Heterozygous mutations in the Berardinelli-Seip congenital lipodystrophy ( BSCL2) gene have been associated with different clinical phenotypes including Silver syndrome/spastic paraplegia 17, distal hereditary motor neuropathy type V, and Charcot-Marie-Tooth disease type 2 (CMT2) with predominant hand involvement. We studied an Italian family with a CMT2 phenotype with pyramidal signs that had subclinical sensory involvement on sural nerve biopsy. Direct sequencing analysis of the BSCL2 gene in the three affected siblings revealed an S90L mutation. This report confirms the variability of clinical phenotypes associated with a BSCL2 Ser90Leu mutation and describes the first Italian family with this mutation. Muscle Nerve, 2010 [ABSTRACT FROM AUTHOR]

OBJECTIVE: Erdheim-Chester disease (ECD) is a rare form of non-Langerhans' cell histiocytosis (LCH) of unknown etiology, characterized by diffuse histiocyte infiltration of bones and soft tissue. The purpose of this study was to assess cell proliferation and expression of cytokines, chemokines, and chemokine receptors that may potentially be important in histiocyte accumulation in ECD lesions. METHODS: Biopsies were performed on 3 patients with ECD. The diagnosis of the disease was based on clinical signs including typical radiologic osteosclerosis, and on the detection of foamy CD68+,CD1a- non-Langerhans' cell histiocytes on histologic examination. The expression of the proliferation marker Ki-67 as well as of selected chemokine/chemokine receptor pairs and cytokines was analyzed by immunohistochemistry. RESULTS: In all samples, Ki-67 was undetectable in CD68+ histiocytes. Conversely, these cells expressed the chemokines CCL2 (monocyte chemotactic protein 1), CCL4/macrophage inflammatory protein 1beta (MIP-1beta), CCL5/RANTES, CCL20/MIP-3alpha, and CCL19/MIP-3beta, and their counter-receptors CCR1, CCR2, CCR3, CCR5, CCR6, and CCR7. Moreover, ECD histiocytes expressed interferon-gamma-inducible 10-kd protein (CXCL10), which is specifically induced by interferon-gamma, and interleukin-6 and RANKL, which are both implicated in bone remodeling. Finally, all cases showed a Th1-type lymphocyte infiltrate. CONCLUSION: Our data indicate that, similar to LCH, ECD lesions are characterized by a complex cytokine and chemokine network, which may orchestrate histiocyte activation and accumulation through an autocrine loop and contribute to the pathogenesis of the disease. [ABSTRACT FROM AUTHOR]

The effects of maternal active and passive smoking during pregnancy on childhood central nervous system (CNS) tumours were assessed in a population-based case-control study. The mothers of 244 children aged 0-15 years with CNS tumours and 502 control mothers were interviewed about their smoking habits. All families were resident in the region of Lombardy, Italy. Risk estimates were calculated by unconditional logistic regression, adjusted for age, sex and area of residence. Active smoking by the parents before pregnancy was not associated with increased risk of CNS tumours in the children. Active smoking by the mother during early pregnancy (approximately the first 5 weeks) was associated with a slightly increased risk of the child developing a CNS tumour (odds ratio [OR] 1.5 [95% CI 1.0, 2.3]). An increased risk of CNS tumours was found in the children of non-smoking mothers exposed regularly to tobacco smoke both in early pregnancy (OR 1.8 [95% CI 1.2, 2.6]) and in late pregnancy (OR 1.7 [95% CI 1.2, 2.5]). Although this study was retrospective, the results confirm our previous findings and suggest an association between the risk of developing CNS tumours in children and regular passive smoking by the mother during pregnancy. [ABSTRACT FROM AUTHOR]

in seven human adenocarcinomas and a non-neoplastic granulomatous disease of the lung, γδ+ infiltrating lymphocytes (TIL) could be isolated and selectively expanded in vitro upon culture in interleukin-2 (IL-2), without any additional stimuli, indicating a prior activation in vivo. In most cases γδ TIL were predominantly Vδ1+, despite a normal Vδ2:Vδ1 ratio in paired peripheral blood lymphocytes, suggesting a possible expansion of this subset in response to localized antigens/ superantigens. Moreover, in five patients it was possible to identify a Vδ1- Vδ2- TIL population which by polymerase chain reaction (PCR) analysis was shown to be heterogeneous as Vδ gene usage, inclusive of Vδ 3,4,5,6,7 and 8. Of note, these Vδ regions have not been found in peripheral blood so far. Finally, in all cases, γδ TIL displayed killing activity of the autologous turnout, which appeared to be more restricted in the case of Vδ1+ cells. Altogether, these findings suggest a preferential expansion, at the tumour site, of Vδ1+ cells and of cells expressing Vδ genes other than Vδ2. [ABSTRACT FROM AUTHOR]

The effects of short-chain fatty acids (SCFA) produced by anaerobic bacteria, namely propionic, butyric and iso-butyric on T-cell proliferation was investigated. A dose-dependent inhibition of both phytohemagglutinin-induced blastogenesis and mixed lymphocyte culture was observed in the millimolar range of SCFA concentrations. The tested SCFA displayed different levels of suppression. The degree of activity was in the following order: butyrate > propionate > isobutyrate. T-cell inhibition was partially reversed, at least for propionic and isobutyric acids, by increasing the concentration of macrophages in the assay system. Furthermore, butyric acid displayed an interesting biphasic stimulation of monocyte interleukin-1 beta production, a cytokine with a powerful bone-resorbing activity. Since millimolar concentrations of SCFA are present in gingival fluid from periodontal pockets, the observed results support the role of these by products of anaerobic metabolism in the pathogenesis of periodontal diseases. [ABSTRACT FROM AUTHOR]

Murine γ/δ T lymphocytes localize to different epithelial tissues and are phenotypically distinct from peripheral γ/δ T cell-populations in that they show limited TCR diversity, express the CD8 α/α homodimer and lack the CD8β chain. In humans, a compartmentalization of γ/δ cells sharing similar phenotypic features has been documented to date only in the case of intestinal epithelium. In the present study we show that about half of Vδ1+ (as well as Vδ1-Vδ2-) γ/δ lymphocytes, which can be selectively expanded from human lung cancers, coexpress the CD8α/α homodimer. The accumulation of intraepithelial CD8+ γ/δ+ lymphocytes might then be a more general phenomenon, possibly as a result of common mechanisms operating at those sites. [ABSTRACT FROM AUTHOR]

Human peripheral blood large granular lymphocytes (LGL)--that is, cells with intracytoplasmic azurophilic (electron-dense) granules, with a positivity for the cytochemical localization of certain acid hydrolases, and with avid surface receptors for the Fc portion of IgG--have been purified on Percoll density gradients. Approximately 30% of these cells expressed receptors for the third complement component (C3R). They were separated into C3R-positive and C3R-negative cells. C3R+ cells had a significantly greater natural killer (NK) activity against K562 target cells than C3R+ cells. This difference was unrelated to the presence in the C3R+ cells of a contaminant cell type incapable of NK activity, since cytochemical and ultrastructural analysis revealed that C3R+ and C3R- fractions contained comparable LGL numbers. Agarose cytotoxicity assays at the single-cell level demonstrated that C3R+ LGL contained a large number of cells that bound to but did not lyse the target. The remaining fully cytotoxic C3R+ LGL had, however, the same killing and recycling properties as the cells from the C3R- fraction. Electron microscopy and cytochemical studies showed that C3R+ cells had fewer electron-dense granules than C3R- cells and stained more faintly for the localization of alpha-naphtyl acetate esterase. In contrast to C3R- cells, C3R+ LGL displayed morphological features suggesting that an active process of granule formation was taking place. Taken together, the data indicate that C3R+ cells represent a discrete subset or a maturational stage of LGL. [ABSTRACT FROM AUTHOR]

Large granular lymphocytes (LGL) are nonadherent cells with cytoplasmic azurophitic granules, avid receptors for the Fc portion of lgG, and a paranuclear localization of alpha-naphthyl acid esterase or acid phosphatase. LGL constitute the bulk of TG cells (cells wish receptors for sheep erythrocytes and for IGC molecules) and null cells (non-T, non-B cells). In the present study we demonstrate that 20-33% of the circulating human LGL express receptors (or the third complement component (C3R). When TG or null cell fractions from normal individuals or non-T cells from a patient with infantile agammaglobulinaemia (which contained almost exclusively LGL) were rosetted with erythrocytes coated with antibody and complement, a variable number of C3R-bearing cells were detected. Such cells were isolated and analysed further; the great majority of them displayed she cytochemical and ultrastructural features of LGL. [ABSTRACT FROM AUTHOR]

Endogenous and surface labelling techniques were used on human lymphoid cells to characterize intracytoplasmic, membrane and secreted IgD. IgD synthesized by lymphocytes and inserted into the cell membrane displayed a single molecular form with the same mobility in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) as the previously described slow migrating serum IgD1. Plasma cells produced and secreted IgD1 and another IgD corresponding to the faster-moving serum IgD2. Conversion of one molecular form into the other was never observed, thus indicating that neither molecule is a precursor or a degradation product of the other. [ABSTRACT FROM AUTHOR]

Human IgD present in the serum of normal individuals or of patients with Hodakin's disease (having high IgD concentrations) was characterized and compared with five IgD myeloma proteins. IgD was isolated using a highly specific anti-δ insoluble immunoabsorbent from which the bound material was critical with sodium dodecyl sulphate (SDS) or urea. The latter reagent could be removed by extensive dialysis, thus making possible the renaturation of the eluted molecules. The purity ot the IgD thus isolated was confirmed by antigenic analysis. Both k and z light chain determinants were present on serum IgD. although z light chain was predominant with a ratio over the K chain of 2:1, SDS-polyacrylamide slah gel electrophoresis analysis revealed two different molecular forms of serum IgD, one (IgDI) migrating identically to monoclonal IgD, the other (lgD2) having a faster mobility. The difference between the two molecules was entirely, due to the different sizes of their constituent δ chains. Peptide mapping of the two chains (δ,1 and δ2 respectively) and of the δ chain of an IgD myeloma protein was carried out using labelled material. The three molecules displayed a high degree of homolog the δ2 chain differing by the presence of three characteristic extra peptides. The significance of these extra peptides is discussed in the light of the peptide mapping technique employed. [ABSTRACT FROM AUTHOR]

Congenital hypomyelination neuropathy (CHN) has been related with mutations of the MPZ gene that codes for P0, the major structural protein of the peripheral myelin and of EGR2/Krox20 gene that codes for a transcription factor essential for the normal development of myelinating Schwann cell. More recently, we reported the association between CHN and a Ser72Leu mutation of the peripheral myelin protein 22 (PMP22), a quantitatively minor component of the compact myelin of peripheral nerves, whose functions are still debated. Here we describe a second patient with CHN associated with a novel mutation of PMP22. The patient is the 4-year-old daughter of healthy nonconsanguineous parents and is affected with early delay of motor development. At seven years she could not walk without support; examination disclosed muscle weakness and atrophy, foot drop, pes cavus, scoliosis, areflexia, and impairment of all sensory modalities. MNCV at the median nerve was 3.6 m/s. Nerve biopsy disclosed marked loss of fibers; residual fibers were devoid of myelin or encircled by extremely thin myelin sheaths and were surrounded by basal laminae onion bulbs. Clinical follow up demonstrated no progression of the disease. Molecular analysis disclosed a novel heterozygous T-to-C transition at nucleotide 374 of PMP22, that is predicted to cause a nonconservative substitution of cysteine109 (a phylogenetically conserved residue) with arginine, in the third transmembrane domain. The duplication of PMP22 causes the common demyelinating form of Charcot-Marie-Tooth neuropathy type 1 (CMT1). Our reports indicate that missense mutations of the same protein may cause dysmyelination rather than demyelination, and prompt to investigate the function of PMP22 in myelinogenesis as well as in Schwann cell differentiation. [ABSTRACT FROM AUTHOR]

Background: Charcot-Marie-Tooth disease type 1 (CMT1) and related Dejerine-Sottas syndrome (DSS) and Hereditary Neuropathy with liability to Pressure Palsy (HNPP) have a high degree of genetic heterogeneity; mutation analysis performed by direct nucleotide sequencing is highly sensitive but time-consuming and expensive. Objective: To evaluate the sensitivity of denaturing high performance liquid chromatography (DHPLC), a recently developed technology for fast mutational analysis. Methods: Optimal conditions for analysing Cx32, P0 and PMP22 genes by DHPLC were developed on the basis of 39 mutations (Cx32 = 21; P0 = 10; PMP22 = 8) detected in the period 1997–2002. Patients: During 2003, 44 patients fitting the clinical and electrophysiological criteria of CMT1, DSS or HNPP who were negative for the 17p11.2 duplication/deletion were analysed either by nucleotide sequencing or by DHPLC. Results: In the last year we identified a total of 3 mutations in Cx32, 3 mutations in MP0 (2 novels mutations, one recessive) and 2 mutations in PMP22 (2 novels nonsense mutations). Conclusions: DHPLC was capable of detecting all mutations identified by sequencing, thus appearing as a reliable approach for the mutational analysis of CMT1. [ABSTRACT FROM AUTHOR]

Chronic lymphocytic leukemia (CLL) is a hematological disorder with complex clinical and biological behavior. TP53 mutational status and cytogenetic assessment of the deletion of the corresponding locus (17p13.1) are considered the most relevant biomarkers associated with pharmaco-predictive response, chemo-refractoriness, and worse prognosis in CLL patients. The implementation of Next Generation Sequencing (NGS) methodologies in the clinical laboratory allows for comprehensively analyzing the TP53 gene and detecting mutations with allele frequencies ≤10%, that is, "subclonal mutations". We retrospectively studied TP53 gene mutational status by NGS in 220 samples from 171 CLL patients. TP53 mutations were found in 60/220 (27.3%) samples and 47/171 (27.5%) patients. Interestingly, subclonal mutations could be detected in 31/60 samples (51.7%) corresponding to 25 patients (25/47, 53.2%). We identified 44 distinct subclonal TP53 mutations clustered in the central DNA-binding domain of p53 protein (exons 5-8, codons 133-286). Missense mutations were predominant (>80%), whereas indels, nonsense, and splice site variants were less represented. All subclonal TP53 variants but one [p.(Pro191fs)] were already described in NCI and/or Seshat databases as "damaging" and/or "probably damaging" mutations (38/44, 86% and 6/44, 14%, respectively). Longitudinal samples were available for 37 patients. Almost half of them displayed at least one TP53 mutant subclone, which could be alone (4/16, 25%) or concomitant with other TP53 mutant clonal ones (12/16, 75%); different patterns of mutational dynamics overtimes were documented. In conclusion, utilization of NGS in our "real-life" cohort of CLL patients demonstrated an elevated frequency of subclonal TP53 mutations. This finding indicates the need for precisely identifying these mutations during disease since the clones carrying them may become predominant and be responsible for therapy failures., (© 2022 The Authors. Hematological Oncology published by John Wiley & Sons Ltd.)

Considering the emergence of antifungal resistance on Sporothrix brasiliensis, we aimed to assess new benzylidene-carbonyl compounds against feline-borne S. brasiliensis isolates. The compounds were designed as bioisosteres from previously reported benzylidene-ketones generating the p-coumaric (1), cinnamic (2), p-methoxycinnamic (3) and caffeic acid (4) analogues. The corresponding compounds were tested against feline isolates of S. brasiliensis with sensitivity (n = 4) and resistance (n = 5) to itraconazole (ITZ), following the M38-A2 protocol (CLSI, Reference method for broth dilution antifungal susceptibility testing of filamentous fungi M38-A2 Guideline, 2008). Eleven analogues showed activity against all fungal strains with minimum inhibitory concentrations (MIC) ≤1 mg/ml (1a-d, 2e, 3b, 3e, 4, 4a and 5e) and fungicidal concentrations (MFC) ≤1 mg/ml (1b, 1d, 3e and 4a), whereas 3 was the less active with both MIC and MFC values above 1 mg/ml. Compound 3e (4-methoxy-N-butylcinnamamide) was the most potent (MIC range 0.08-0.16 mg/ml; MFC range 0.32-0.64 mg/ml) from the set, suggesting a different role of the substituents in ester and amide derivatives. The designed compounds proved to be important prototypes with improved drug-likeness to achieve compounds with higher activity against ITZ-resistant S. brasiliensis., (© 2021 John Wiley & Sons A/S.)

Background and Aim of the Study: The aim of this meta-analysis was to compare short- and long-term outcomes of patients undergoing mitral annuloplasty (MA) with or without papillary muscle surgery (PMS) for the treatment of ischemic mitral regurgitation (IMR)., Methods: A systematic review and meta-analysis in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement were performed., Results: Nine studies met the inclusion criteria. This meta-analysis identified 478 patients: 228 patients underwent MA alone and 250 patients underwent concomitant PMS. Early mortality was similar between two groups (odds ratio [OR] 1.14, 95% confidence interval [CI], 0.51-2.53; P = .75). PMS was associated at follow-up with a higher freedom from cardiac-related events (P = .050); moreover, although both surgical techniques had a positive impact on ventricular remodeling, the PMS group showed a significant higher reduction of left ventricle end-diastolic diameter (OR, 4.89, 95% CI, 2.77-7.01; P < .001) and left ventricle end-systolic diameter values (OR, 4.11, 95% CI, 1.98-6.24; P < .001). Finally, PMS compared with MA alone was associated with a significant reduction of recurrent mitral regurgitation at follow-up (OR, 3.25, 95% CI, 1.60-6.59; P = .001)., Conclusions: This meta-analysis demonstrated superiority in terms of ventricular remodeling of a combined approach encompassing PMS and MA over MA alone in IMR. Moreover, the association of subvalvular surgery with restrictive MA decreases the incidence of mitral regurgitation recurrence and cardiac-related events at follow-up., (© 2020 Wiley Periodicals, Inc.)

Background: Aortic stenosis is the most common valvular disease and has a dismal prognosis without surgical treatment. The aim of this meta-analysis was to quantitatively assess the comparative effectiveness of the Perceval (LivaNova) valve versus conventional aortic bioprostheses., Methods and Results: A total of 6 comparative studies were identified, including 639 and 760 patients who underwent, respectively, aortic valve replacement with the Perceval sutureless valve (P group) and with a conventional bioprosthesis (C group). Aortic cross-clamping and cardiopulmonary bypass duration were significantly lower in the P group. No difference in postoperative mortality was shown for the P and C groups (2.8% versus 2.7%, respectively; odds ratio [OR]: 0.99 [95% confidence interval (CI), 0.52-1.88]; P =0.98). Incidence of postoperative renal failure was lower in the P group compared with the C group (2.7% versus 5.5%; OR: 0.45 [95% CI, 0.25-0.80]; P =0.007). Incidence of stroke (2.3% versus 1.7%; OR: 1.34 [95% CI, 0.56-3.21]; P =0.51) and paravalvular leak (3.1% versus 1.6%; OR: 2.52 [95% CI, 0.60-1.06]; P =0.21) was similar, whereas P group patients received fewer blood transfusions than C group patients (1.16±1.2 versus 2.13±2.2; mean difference: 0.99 [95% CI, -1.22 to -0.75]; P =0.001). The incidence of pacemaker implantation was higher in the P than the C group (7.9% versus 3.1%; OR: 2.45 [95% CI, 1.44-4.17]; P =0.001), whereas hemodynamic Perceval performance was better (transvalvular gradient 23.42±1.73 versus 22.8±1.86; mean difference: 0.90 [95% CI, 0.62-1.18]; P =0.001), even during follow-up (10.98±5.7 versus 13.06±6.2; mean difference: -2.08 [95% CI, -3.96 to -0.21]; P =0.030). We found no difference in 1-year mortality., Conclusions: The Perceval bioprosthesis improves the postoperative course compared with conventional bioprostheses and is an option for high-risk patients., (© 2018 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.)

The chronic lymphocytic leukemia International Prognostic Index (CLL-IPI) combines 5 parameters (age, clinical stage, TP53 status [normal vs. del(17p) and/or TP53 mutation], IGHV mutational status, serum β2-microglobulin) to predict survival and time-to-first-treatment (TTFT) in CLL patients. We performed an observational study in 337 prospectively collected, Binet stage A patients to validate the ability of the CLL-IPI to predict TTFT in an independent cohort of early stage CLL patients. The CLL-IPI score stratified Binet stage A patients into three subgroups with different outcome. Since the CLL-IPI was originally developed to predict survival, we next investigated the optimal cut-off score to predict TTFT in Binet stage A patients. Recursive partitioning analysis identified three subsets with scores of 0 (n = 139), 1 (n = 90), and ≥ 2(n = 108). The probability of remaining free from therapy 5 years after diagnosis was 85%, 67% and 46% in these three categories (P < 0.0001.; C-statistic:c = 0.72; 95% CI:0.58-0.81). This optimized CLL-IPI scoring for TTFT was subsequently validated in an independent cohort of Binet A patients from the Mayo Clinic (n = 525). The ability of either original or optimized CLL-IPI to predict TTFT was equivalent to other prognostic models specifically designed for this endpoint (2011 MDACC score and O-CLL1 score). Although originally developed to predict suvival, the CLL-IPI is useful for predicting TTFT in early stage CLL patients. Am. J. Hematol. 91:1090-1095, 2016. © 2016 Wiley Periodicals, Inc., Competing Interests: Conflict-of-interest disclosure: The authors have no conflict of interest to disclose., (© 2016 Wiley Periodicals, Inc.)

A risk score based on three biological features (CD38, ZAP-70, and IGHV mutational status) was previously developed to predict progression-free survival (PFS) in untreated Binet A CLL patients. Here we perform a score validation analysis in a prospective and independent cohort of patients. Biological markers (CD38, ZAP-70, and IGHV mutational status) and gene expression profiles (GEP) of leukemic cells from CLL patients included in a prospective multicenter observational study (O-CLL1-GISL protocol, clinicaltrial.gov ID:NCT00917549) were used to assess the value and reproducibility of this score. To date, 468 Binet A patients were classified as low- (0 positive marker), intermediate- (1 positive marker), or high-risk (2 or 3 positive markers) using the progression risk score. The 3-year PFS probability was 91.7%, 82.9%, and 57.4% for low-, intermediate-, and high-risk (P < 0.0001) cases, respectively. These values were similar to those found in the original cohort. At Cox multivariate analysis, Rai stage, absolute lymphocyte count, progression risk score, and β-2 microglobulin maintained an independent prognostic impact on PFS. This score remained a predictor of progression when analysis was limited to 371 Rai 0 cases (P < 0.0001). Finally, the cells from the different CLL risk groups showed differences in their gene expression patterns. These results confirm the ability of this progression risk score to predict PFS among Binet A patients. The utility of the score was also extended by demonstrating that it retains prognostic value when applied exclusively to Rai 0 patients. Specific transcriptional patterns were significantly associated with risk groups., (© 2014 Wiley Periodicals, Inc.)

NOTCH1 mutations have recently emerged as new genetic lesions significantly correlated with survival in chronic lymphocytic leukaemia (CLL). We performed deep next generation sequencing of the NOTCH1 mutation hotspot in 384 cases at diagnosis, including 100 monoclonal B cell lymphocytosis (MBL) and 284 Binet stage A CLL cases, enrolled in the Gruppo Italiano Studio Linfomi O-CLL1 multicentre trial. The NOTCH1 c.7541&#95;7542delCT dinucleotide deletion was detected and confirmed by an extremely sensitive polymerase chain reaction-based approach in 11% of MBL and 13·4% of CLL patients. Remarkably, the NOTCH1 mutation was often observed at low clonal level, mainly in MBL patients. Sequential analyses in a fraction of cases showed that the NOTCH1 mutation generally does not occur during the disease course and that the mutational load in positive cases tends to be stable over time. NOTCH1-mutated cases, even at low clonal level, displayed a significant reduction in median progression-free survival, although NOTCH1 mutation lost its prognostic impact in a multivariate analysis including 11q and/or 17p deletion, IGHV mutational status, and MBL or CLL status. Our data highlight the importance of using highly sensitive methods to measure NOTCH1 mutations, in order to improve prognostic stratification and obtain useful information for potential therapeutic approaches., (© 2014 John Wiley & Sons Ltd.)

Total body computed tomography (TB-CT) scan is not mandatory in the diagnostic/staging algorithm of chronic lymphocytic leukemia (CLL). The aim of this study was to determine the value and prognostic significance of TB-CT scan in early stage CLL patients. Baseline TB-CT scan was performed in 240 Binet stage A CLL patients (179 Rai low- and 61 Rai intermediate-risk) included in a prospective multicenter observational study (clinicaltrial.gov ID:NCT00917549). The cohort included 69 clinical monoclonal B lymphocytosis (cMBLs). Patients were restaged considering only radiological data. Following TB-CT scans, 20% of cases reclassified as radiologic Binet (r-Binet) stage B. r-Binet B patients showed a higher incidence of unfavorable cytogenetic abnormalities (P = 0.027), as well as a shorter PFS (P = 0.001). At multivariate analysis, r-Binet stage [HR = 2.48; P = 0.004] and IGHV mutational status [HR = 3.01; P = 0.002] retained an independent predictive value for PFS. Among 179 Rai low-risk cases, 100 were redefined as r-Rai intermediate-risk based upon TB-CT scan data, showing a higher rate of cases with higher ZAP-70 (P = 0.033) and CD38 expression (P = 0.029) and β2-microglobulin levels (P < 0.0001), as well as a shorter PFS than those with r-Rai low-risk (P = 0.008). r-Rai stage [HR = 2.78; P = 0.046] and IGHV mutational status [HR = 4.25; P = 0.009] retained a significant predictive value for PFS at multivariate analysis. Forty-two percent of cMBL patients were reclassified as r-small lymphocytic lymphomas (r-SLLs) by TB-CT scan. TB-CT scan appears to provide relevant information in early stage CLL related to the potential and the timing of patients to progress towards the more advanced disease stages., (Copyright © 2013 Wiley Periodicals, Inc.)

Recent studies have described chromosome 2p gain as a recurrent lesion in chronic lymphocytic leukemia (CLL). We investigated the 2p gain and its relationship with common prognostic biomarkers in a prospective series of 69 clinical monoclonal B-cell lymphocytosis (cMBL) and 218 early stage (Binet A) CLL patients. The 2p gain was detected by FISH in 17 patients (6%, 16 CLL, and 1 cMBL) and further characterized by single nucleotide polymorphism-array. Overall, unfavorable cytogenetic deletions, i.e., del(11)(q23) and del(17)(p13) (P = 0.002), were significantly more frequent in 2p gain cases, as well as unmutated status of IGHV (P < 1 × 10(-4) ) and CD38 (P < 1 × 10(-4) ) and ZAP-70 positive expression (P = 0.003). Furthermore, 2p gain patients had significantly higher utilization of stereotyped B-cell receptors compared with 2p negative patients (P = 0.009), and the incidence of stereotyped subset #1 in 2p gain patients was significantly higher than that found in the remaining CLLs (P = 0.031). Transcriptional profiling analysis identified several genes significantly upregulated in 2p gain CLLs, most of which mapped to 2p. Among these, NCOA1 and ROCK2 are known for their involvement in tumor progression in several human cancers, whereas among those located in different chromosomes, CAV1 at 7q31.1 has been recently identified to play a critical role in CLL progression. Thus, 2p gain can be present since the early stages of the disease, particularly in those cases characterized by other poor prognosis markers. The finding of genes upregulated in the cells with 2p gain provides new insights to define the pathogenic role of this lesion., (Copyright © 2012 Wiley Periodicals, Inc.)

CD26 (dipeptidyl peptidase IV, DPP IV) is widely expressed by T and natural killer (NK) cells, epithelial and endothelial cells of different tissues, and it is strongly upregulated in activated B-cells; moreover it plays a regulatory role in the neoplastic transformation and progression of various types of tumours. CD26 expression was evaluated by means of flow cytometry in various peripheral B-cell lymphoid tumours: 12 follicular and 12 mantle cell lymphomas, 20 multiple myelomas (MMs), 12 hairy cell leukaemias (HCLs), 112 chronic lymphocytic leukaemias (CLLs), 20 CD5(negative) B-cell chronic lymphoproliferative diseases (CD5(neg) B-CLPDs) and 12 diffuse large cell lymphomas (DLCLs). CD26 expression was absent or barely detectable in follicular and mantle cell lymphomas, high in MMs and HCLs, and variable in CLLs, in CD5(neg) B-CLPDs and in DLCLs. CD26 significantly correlated with CD49d and CD38 expressions (p < 0.0001) in B-CLLs, and there was a significant correlation between CD26 and ZAP-70 expressions or IgVH mutational status (p < 0.0001). After a median follow-up of 36 months, 65 B-CLL patients were treated; taking 10% as the best CD26 cut-off value, Kaplan-Meier curves revealed a significantly shorter time to treatment in the CD26-positive cases (p < 0.0001). Overall, our data indicate that CD26 expression may identify subsets of B-CLL patients with an unfavourable clinical outcome in terms of therapeutic need, thus suggesting its potential role as a marker (together with CD38 and CD49d) in a future routine cytofluorimetric panel to be validated for the prognostic stratification of B-CLLs.

IGHV mutational status and ZAP-70 or CD38 expression correlate with clinical course in B-cell chronic lymphocytic leukaemia (CLL). The three markers may be discordant in the single case and there is no consensus on their combined use in clinical practise. This multicenter study investigated this issue. Two-hundred and sixty-two Binet stage A patients were studied for the three markers. Sixty patients were profiled with HG-U133A gene expression chips. Disease progression was determined by time from diagnosis to treatment (TTT). The probability of being treatment-free at 3 years was significantly shorter in patients with unmutated IGHV genes (IGHVunmut 66% vs. 93%, chi square of log-rank = 30, P < 0.0001), ZAP-70 positive (ZAP-70pos 73% vs. 96%, chi square of log-rank = 8.2, P = 0.004) or CD38-positive cells (CD38pos 68% vs. 91%, chi square of log-rank = 21, P < 0.0001). Cox multivariate regression analysis showed that the three markers had an independent predictive value for TTT of similar power. A prognostic system based on presence of none (low-risk), one (intermediate-risk) or two or three (high-risk) markers was generated. Based on such criteria, 56%, 23% and 21% of cases were clustered in low (HR = 1), intermediate [HR = 2.8, 95% confidence interval (CI) 2.4-5.8] and high-risk group (HR = 8.0, 95% CI 3.9-16.2). Specific transcriptional patterns were significantly associated with risk groups.

The use of proteasome inhibitors have been a major advance in the treatment of multiple myeloma (MM), but their mechanisms of action remain largely unclear. A better understanding of the cellular events downstream of proteasome inhibition is essential to improve the response and identify new combination therapies for MM and other malignancies. This study analysed the relationships between redox homeostasis and bortezomib treatment in MM cells. Our data showed that decreasing intracellular glutathione through buthionine sulfoximine treatment strongly enhances bortezomib toxicity, whilst antioxidants protect MM cells from bortezomib-mediated cell death. Bortezomib treatment decreases intracellular glutathione both in MM cell lines and in malignant plasma cells obtained from MM patients. Glutamate-cysteine ligase (GCLM) and haem-oxygenase-1 (HMOX1), two genes involved in the Nrf-2-mediated antioxidant response, as well as two eIF2alpha-downstream transcription factors, activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP), are upregulated, indicating that redox-related adaptive responses are initiated in bortezomib-treated MM cells. These findings demonstrate tight links between sensitivity to proteasome inhibition and redox homeostasis in MM cells and have potential implications for treatment.

This study reports the characterization of a spontaneous lymphoblastoid cell line (LCL) raised from the peripheral blood of a patient with Kostmann's congenital neutropenia. The LCL was composed of EBV-infected polyclonal B cells and displayed surface markers and pattern of growth in vitro typical of normal LCLs. The supernatant of the LCL contained a colony inhibiting activity (CIA) that decreased the cloning efficiency of normal committed haemopoietic progenitors and was identified as immunoreactive transforming growth factor beta 1 (TGF-beta 1) by neutralization experiments with a specific antiserum. Control studies with a panel of LCLs spontaneously derived from the peripheral blood of patients seropositive for Epstein-Barr virus (EBV) infections showed that 5/30 LCLs produced a CIA. This CIA was not identifiable as TGF-beta 1 but rather was due to the combined effects of tumour necrosis factor alpha (TNF alpha), tumour necrosis factor beta (TNF beta) and interferon alpha (IFN alpha), that were present in the LCL supernatants. The hypothesis that the B cells latently infected by EBV in vivo and possibly expanded as a consequence of the infection may have contributed to the inhibition of the patient granulopoiesis by releasing TGF-beta 1 will be discussed.

Clones of dermal fibroblasts from the skin of 4 normal subjects and 5 patients with progressive systemic sclerosis (PSS; scleroderma) were established, and their synthetic and proliferative characteristics were compared. A limiting-dilution assay was used to determine frequencies of cloning in the microcultures of dermal fibroblasts plated. The clones derived from single cells were expanded in vitro and examined (in passages C-H) for growth and synthesis of glycosaminoglycan (GAG) and collagenase-sensitive protein (CSP). The clonogenicity of PSS fibroblasts was not significantly different from that of normal fibroblasts. Normal fibroblast clones were characterized by low levels of GAG and CSP synthesis, and there was a correlation between the GAG and CSP phenotypes. In contrast, clones of PSS fibroblasts were often, but not always, high producers of GAG and CSP, but there was no correlation between the levels of GAG and CSP synthesis. It appears that scleroderma skin is composed of fibroblast clones that are unable to regulate the synthesis of connective tissue components and often synthesize large amounts of connective tissue macromolecules.