Radiation measurements in the International Space Station, Columbus module, in 2020-2022 with the LIDAL detector.

The Light Ion Detector for ALTEA (LIDAL) is a new instrument designed to measure flux, energy spectra and Time of Flight of ions in a space habitat. It was installed in the International Space Station (Columbus) on January 19, 2020 and it is still operating. This paper presents the results of LIDAL measurements in the first 17 months of operation (01/2020-05/2022). Particle flux, dose rate, Time of Flight and spectra are presented and studied in the three ISS orthogonal directions and in the different geomagnetic regions (high latitude, low latitude, and South Atlantic Anomaly, SAA). The results are consistent with previous measurements. Dose rates range between 1.8 nGy/s and 2.4 nGy/s, flux between 0.21 particles/(sr cm2 s) and 0.32 particles/(sr cm2 s) as measured across time and directions during the full orbit. These data offer insights concerning the radiation measurements in the ISS and demonstrate the capabilities of LIDAL as a unique tool for the measurement of space radiation in space habitats, also providing novel information relevant to assess radiation risks for astronauts.

Respirometric techniques coupled with laboratory-scale tests for kinetic and stoichiometric characterisation of fungal and bacterial tannin-degrading biofilms.

In environmental biotechnology applications for wastewater treatment, bacterial-based bioprocesses are mostly implemented; on the contrary, the application of fungal-based bioprocesses, is still challenging under non-sterile conditions. In a previous laboratory-scale study, we showed that when specific tannins are used as the sole carbon source, fungi can play a key role in the microbial community, under non-sterile conditions and in the long term. In a previous study, an engineered ecosystem, based on fungal tannin biodegradation, was successfully tested in a laboratory-scale bioreactor under non-sterile conditions. In the present study, a kinetic and stoichiometric characterisation of the biomass developed therein was performed through the application of respirometric techniques applied to the biomass collected from the above-mentioned reactor. To this aim, a respirometric set-up was specifically adapted to obtain valuable information from tannin-degrading fungal biofilms. A mathematical model was also developed and applied to describe both the respirometric profiles and the experimental data collected from the laboratory-scale tests performed in the bioreactor. The microbial growth was described through a Monod-type kinetic equation as a first approach. Substrate inhibition, decay rate and tannin hydrolysis process were included to better describe the behaviour of immobilised biomass selected in the tannin-degrading bioreactor. The model was implemented in AQUASIM using the specific tool Biofilm Compartment to simulate the attached fungal biofilm. Biofilm features and transport parameters were either measured or assumed from the literature. Key kinetic and stoichiometric unknown parameters were successfully estimated, overcoming critical steps for scaling-up a novel fungal-based technology for tannins biodegradation.

Stability of fungal biomass continuously fed with tannic acid in a non-sterile moving-packed bed reactor.

A number of bacteria and fungi are known to degrade tannins. In this study, the efficiency of the white-rot fungus, Bjerkandera adusta MUT 2295, was evaluated for the treatment of a synthetic solution prepared with tannic acid. Tests were performed in continuously fed, bench-scale, packed-bed reactors, operated under non-sterile conditions with biomass immobilized within PolyUrethane Foam cubes (PUFs). The main parameters monitored to evaluate the process efficiency were: soluble Chemical Oxygen Demand (sCOD), Total Organic Carbon (TOC) removal, and activities. of Tannase and Lignin Peroxidase. At the end of the process, additional parameters were evaluated, including the increase of fungal dry weight and the presence of ergosterol. The reactor was operative for 210 days, with maximum sCOD and TOC removal of 81% and 73%, respectively. The reduction of sCOD and TOC were positively correlated with the detection of Tannase and Lignin Peroxidase (LiP) activities. Increases in biomass within the PUF cubes was associated with increases in ergosterol concentrations. This study proved that the fungal-based system tested was efficient for the degradation of tannic acid over a period of time, and under non-sterile conditions.

Effect of cellulose as co-substrate on old landfill leachate treatment using white-rot fungi.

Conventional wastewater treatment technologies are ineffective for remediation of old LandFill Leachate (LFL), and innovative approaches to achieve satisfactory removal of this recalcitrant fraction are needed. This study focused on old LFL treatment with a selected fungal strain, Bjerkandera adusta MUT 2295, through batch and continuous tests, using packed-bed bioreactors under non-sterile conditions. To optimize the process performance, diverse types of co-substrates were used, including milled cellulose from beverage cups waste material. Extracellular enzyme production was assayed, in batch tests, as a function of a) cellulose concentration, b) leachate initial Chemical Oxygen Demand (COD) and Soluble COD (sCOD), and c) co-substrate type. Bioreactors were dosed with an initial start-up of glucose (Rg) or cellulose (Rc). An additional glucose dosage was provided in both reactors, leading to significant performance increases. The highest COD and sCOD removals were i) 63% and 53% in Rg and ii) 54% and 51% in Rc.

Chronic lymphocytic leukemia with 6q- shows distinct hematological features and intermediate prognosis.

Cytogenetic and fluorescence in situ hybridization studies were successfully performed in 217 chronic lymphocytic leukemia (CLL). In all, 13 patients with 6q21 deletion were identified and characterized in comparison with 92 patients with 'favourable' karyotype (normal or 13q-), 69 cases with 'intermediate risk' (1-2 anomalies) and 43 cases with 'unfavourable' karyotype (complex, 11q- or 17p-). Six out of 13 cases with 6q- showed an excess of atypical lymphocytes, a finding confirmed at the histologic level; >20% CD38+ cells were seen in 5/6 cases. IGVH mutational status revealed >98% homology to the germline sequence in 4/10 cases. When compared with the 'favourable' group, patients with 6q- showed a higher white blood cell (WBC) count, frequent splenomegaly, atypical morphology, CD38+ and short time from diagnosis to first treatment and short survival. A higher median WBC count was found in the 6q- group vs the intermediate-risk group; survival was shorter in the unfavourable group. To ascertain if the 6q- anomaly was an independent factor predicting for an inferior outcome among those patients with 'favourable' cytogenetics, we performed an analysis of prognostic factors in 105 patients (92 'favourable' plus 13 with 6q-), showing that the 6q- chromosome maintained its prognostic significance at multivariate analysis (P=0.02) along with stage (P=0.01). We conclude that CLL with 6q- is characterized by a high incidence of atypical morphology, classical immunophenotype with CD38 positivity and intermediate incidence of IGVH somatic hypermutation. Clinicobiological features and outcome show that this cytogenetic subset of CLL should be allocated in an intermediate-risk category.

Incidence and significance of cryptic chromosome aberrations detected by fluorescence in situ hybridization in acute myeloid leukemia with normal karyotype.

To better define the incidence and significance of cryptic chromosome lesions in acute myeloid leukemia (AML), fluorescence in situ hybridization (FISH) studies were performed in interphase cells and, when appropriate, in metaphase cells and in morphologically intact BM smears. Fifty-five adult de novo AML (group A) and 27 elderly AML or AML after myelodysplastic syndrome (AML-MDS) (group B) were tested using probes detecting the following anomalies: -5, -7, +8, deletions of 5q31, 7q31, 12p13/ETV6, 17p13/p53, 20q11. All the patients had a normal karyotype in more than 20 cells and tested negative for the common AML-associated fusion genes. No patient in group A was found to carry occult chromosome anomalies, whereas 8/27 patients in group B (P < 0.0001) showed 5q31 or 7q31 deletion (three cases each), a 17p13/p53deletion or trisomy 8 (one case each) in 33-60% interphase cells. Metaphase cells showed only one hybridization signal at 5q31 (three cases) and 7q31 (one case), whereas two normal signals at 7q31 and chromosome 8 centromeres were seen in two patients with 7q deletion and trisomy 8 in interphase cells. The majority of blast cells (76-94%) carried the chromosome anomaly in all cases; erythroid involvement in a minority of cells was seen in three patients. In group B, the presence of occult chromosome anomalies was associated with exposure to myelotoxic agents in the workplace (5/8 cases vs 3/19, P = 0.026) and with a lower complete remission rate (0/6 patients vs 7/12, P = 0.024). We arrived at the following conclusions: (1) cryptic chromosome deletions in the order of a few hundred kb magnitude may be found in a fraction of elderly AML or MDS-related AML and not in de novo adult AML with normal karyotype; (2) these chromosome lesions are usually represented by submicroscopic rearrangements; (3) they display a specific pattern of cell-lineage involvement arguing in favor of their role in the outgrowth of the leukemic blast cells; (4) they are associated with a history of exposure to myelotoxic agents in the workplace and, possibly, with resistance to induction treatment.

Clinical importance of interphase cytogenetics detecting occult chromosome lesions in myelodysplastic syndromes with normal karyotype.

At diagnosis, approximately half of myelodysplastic (MDS) patients presents a normal karyotype by conventional cytogenetic analysis (CCA). Fluorescent in situ hybridization (FISH) is more sensitive than CCA allowing for the detection of minor clones and of submicroscopic lesions. We have analyzed by FISH 101 MDS patients with normal karyotype for the occurrence of the abnormalities which are most frequently observed in MDS (ie -5/5q-, -7/7q-, +8, 17p-). In 18 patients, 15 to 32% of interphase cells were found to carry one FISH abnormality. Six patients presented trisomy 8, five had del(5)(q31), five del(7)(q31), one monosomy 7 and one del(17)(p13). FISH abnormalities were more frequently observed among patients with an increased percentage of bone marrow blasts (P = 0.001). FISH abnormalities were also associated with a higher rate of progression into AML (13/18 vs 12/83, P < 0.001) and were predictive for a worse prognosis (P < 0.001). Multivariate analysis indicated that FISH positivity and IPSS risk group were independent predictors for a poor survival (P = 0.0057 and 0.0123, respectively) and for leukemic transformation (P = 0.0006 and 0.035, respectively). Leukemic transformation in FISH-positive patients was associated in all cases with an expansion of the abnormal clone. Our data demonstrated that a significant proportion of MDS patients with normal karyotype presented, if analyzed by FISH, clones of cytogenetically abnormal cells which played a determinant role in the progression of the disease. The presence of FISH abnormalities identified a group of MDS patients with normal karyotype characterized by an inferior prognosis.

A novel recurrent translocation t(11;14)(p11;q32) in splenic marginal zone B cell lymphoma.

A novel recurrent translocation t(11;14)(p11;q32) was found in three patients with splenic marginal zone B cell lymphoma (MZBCL). Fluorescence in situ hybridization (FISH) studies with IgH probes revealed in all cases involvement of the IgH locus, with breakpoint downstream of the IGVH sequences. Partner genes at 11p11 were not identified. The translocation defined the stem line in two patients, who carried additional cytogenetic aberrations, including a 17p deletion, present in both cases. In one patient a 7q- chromosome was the primary cytogenetic defect, the t(11;14) having been found in four out of 11 abnormal metaphase cells at the time of transformation into high-grade MZBCL. Hematological features in all cases included splenomegaly with peripheral blood (PB) involvement by a monoclonal B cell population consisting of lymphocytes with villous projections and several blast-like cells. The immunophenotype was CD19+; CD22bright+; CD23-, CD10-, CD5-, surface Igbright+. A bone biopsy in one patient revealed an interstitial infiltration with an intrasinusoidal pattern of growth. Histological studies on spleen specimens in two patients showed an expanded marginal zone, with small lymphocytes and several blast-like cells. One patient had a therapy-demanding disease, with partial, short-term responses to cytotoxic treatment; one patient transformed into a high-grade MZBCL involving the gut, the PB and the bone marrow 2 years after diagnosis; one patient was unresponsive to cytotoxic treatment and underwent splenectomy. The t(11;14)(p11;q32) may define a subset of splenic MZBCL with a high-grade component and a relatively aggressive clinical behavior.

Secondary chromosome changes in mantle cell lymphoma: cytogenetic and fluorescence in situ hybridization studies.

To better define the incidence and nature of secondary chromosome anomalies in mantle cell lymphoma (MCL) carrying the t(11:14)/BCL1 rearrangement, cytogenetic and fluorescence in situ hybridization studies (FISH) were performed in 42 patients (39 classical histology, 3 blastoid variant), using 6q21, 9p21/p16, 13q14, 17p13/p53 and chromosome-12-specific probes. Karyotypes from 89 cases published in 5 recent series including patients diagnosed in a homogeneous fashion were reviewed. In our series, FISH confirmed the interpretation of the karyotype in all cases and disclosed cryptic chromosome deletions in a sizeable fraction of cases. One patient (2.4% of total) was found with a cryptic 9p21 deletion by FISH. Two cases (4.8%) had a 6q21 deletion at CCA and at FISH; +12 was found in three cases by CCA plus nine by FISH (28.6%); 13q14 deletion was found in six cases by CCA plus 16 by FISH (52.4%), 17p13 deletion in three cases by CCA plus 8 by FISH (26.2%). In 131 patients (42 present series plus 89 in the literature) secondary chromosome aberrations seen by conventional cytogenetic analysis in more than 5 cases included deletions/translocations (del/t) 6q15-23 [15 cases]; -13 [14 cases]; del/t 1p21-31 [12 cases]; +3q [11 cases]; del/t 17p [9 cases]; 8p translocations and del(Y) [8 cases each]; -20 [7 cases]; 13q14 deletion, del/t 11q22-23, del/t 9q, del(10)(q22q24), -20, -21, -22 and -X [6 cases each]. We arrived at the following conclusions: i) though no secondary anomaly is specific for MCL, there is a distinct profile of recurrent chromosome lesions in MCL with 1p21-31 deletions, 8p translocations, 11q22-23 anomalies having a strong association with CD5+ B-cell lymphomas of low-to-intermediate grade histology; ii) FISH enabled the detection of cryptic chromosome 12, 13q and 17p rearrangements in a sizeable fraction of cases; iii) 9p21/p16 deletions did not occur at a high incidence in this series, possibly because of the low number of cases with blastoid variant.

Multilineage involvement in the 5q- syndrome: a fluorescent in situ hybridization study on bone marrow smears.

BACKGROUND AND OBJECTIVES: A pluripotent progenitor cell was demonstrated to be involved in myelodysplastic syndromes (MDS) with normal karyotype or with numerical chromosome aberrations, but the pattern of lineage involvement by the 5q31 deletion in the 5q- syndrome is unknown. We performed this study in order to define the distribution pattern of the 5q- anomaly better in the non-lymphoid cell compartment DESIGN AND METHODS: Bone marrow (BM) smears from 8 patients with the 5q- syndrome were studied by a modification of the fluorescent in situ hybridization (FISH) technique that allowed direct visualization of cell morphology. A commercial LSI EGR1 probe (Vysis Inc.) for the 5q31 band was used simultaneously in dual-color experiments with a chromosome-5-centromeric probe (Vysis Inc.) on BM smears from 8 patients with the 5q-syndrome. As additional internal controls a chromosome-7-centromeric probe and a 7q31 probe were used. To establish the sensitivity limit of this approach 5 normal BM smears were studied. All 8 patients had the 5q- chromosome as the sole anomaly in 45% to 75% of the interphase cells. RESULTS: For each patient 20-40 erythroblasts were analyzed: they were mostly proerythroblasts and basophilic erythroblasts. In all patients a clone carrying the 5q31 deletion was detected (35-50% of the cells, median 45%). Between 20-50 granulocyte precursors were scored; the 5q31 deletion was found in 40%-50% (median 45%) in all cases. The proportion of neutrophils carrying the 5q deletion was consistently lower than the corresponding value in promyelocytes (28.7% vs 45.6%). In the 20-25 megakaryocytes analyzable in all patients, the overall incidence of 5q31 deletion was 52-68%. Equal proportions of large multilobular megakaryocytes and hypolobular megakaryocytes characteristic of the 5q- syndrome were scored: the latter cells showed the 5q31 deletion more frequently than the former cells (93.6% vs 19.3% of the cells). In 66% to 100% of the cases (median 83%) a few cells with uncondensed nuclear chromatin pattern, and two or three prominent nucleoli with cytoplasmatic hypogranulation were seen in each sample carrying the 5q31 deletion. INTERPRETATION AND CONCLUSIONS: We arrived at the following conclusions: i) the transformation in the 5q- syndrome involves an early progenitor cell retaining the ability to proceed along multiple differentiation pathways; ii) there is a preferential distribution of the 5q31 deletion within immature cells and morphologically abnormal megakaryocytes.

Molecular cytogenetic characterization of marginal zone B-cell lymphoma: correlation with clinicopathologic findings in 14 cases.

BACKGROUND AND OBJECTIVES: To improve the definition of the incidence and significance of chromosome lesions occurring in marginal zone B-cell lymphoma (MZBCL). DESIGN AND METHODS: Fourteen cases of MZBCL diagnosed according to the REAL classification were studied by conventional chromosome analysis (CCA) and by interphase fluorescence in situ hybridization (FISH) using the following probes: 3q27/BCL6, 6q21, 7q31, 9p21/p16, 11q22/ATM, 13q14, 17p13, centromeres of #3, #7, #12. Pertinent clinical data were collected. RESULTS: Primary disease presentation consisted of histologically documented splenic MZBCL in 9 cases, nodal MZBCL in 3 cases and extra-nodal MZBCL in 2 cases. Four cases showed evolution into a high-grade lymphoma, due to the presence of a predominant large cell or blast cell component. Clonal karyotype anomalies were detected by CCA in 12 cases, 6 of which had a complex karyotype, including all 4 cases with high-grade histology. Interphase FISH confirmed cytogenetic data and revealed several cryptic chromosomal lesions. Overall, total/partial +12 was found in five cases; 13q14 and 17p13 deletion were found in four cases each; +3, 7q31 deletion and a BCL6 split signal were found in three cases; deletions at 6q21 and 11q22.3 in two cases each; +7 and a 9p21 deletion were found in one case each. INTERPRETATION AND CONCLUSIONS: i) Besides +3 and 7q-, 13q14 deletion, total/partial +12, BCL6 rearrangement, and deletions at 6q21, 11q22-23, and 17p13.3 are relatively frequent events in MZBCL; ii) unlike in mantle cell lymphoma, 9p21 deletion occurred infrequently in MZBCL; iii) a switch into high grade histology is usually associated with complex chromosome defects, including 6q-, 11q-, +12, and 17p.

BCL-1 rearrangements and p53 mutations in atypical chronic lymphocytic leukemia with t(11;14)(q13;q32).

BACKGROUND AND OBJECTIVES: The translocation t(11;14) (q13;q32), typically described in mantle cell lymphomas (MCL), has also been found in some cases of non-MCL lymphoproliferative disorders, such as splenic lymphoma with villous lymphocytes (SLVL), multiple myeloma (MM), prolymphocytic leukemia (PLL), typical and atypical chronic lymphocytic leukemia (CLL and aCLL). In order to define better the genetic features of aCLL with t(11;14), which could represent a distinct disease subset, we looked for genetic lesions in the BCL-1 locus and in BCL-2, BCL-6, c-myc and p53 genes. DESIGN AND METHODS: We investigated a panel of B-lymphoproliferative disorders with translocation t(11;14)(q13;q32) including nine aCLL, six MCL and one MM. Southern and Northern blot analysis was used to investigate DNA structure and RNA expression; SSCP and direct sequencing were used to detect and characterize p53 point mutations; cytofluorimetric analysis was used to quantify p53 protein. RESULTS: Alterations of BCL-2, BCL-6 and c-myc were not detected. Conversely, BCL-1 rearrangements were present in 4 out of 7 aCLL and in 2 out of 4 MCL. A high incidence of p53 gene alterations was found, almost equivalent in aCLL and MCL. INTERPRETATION AND CONCLUSIONS: Our results indicate that the occurrence of BCL-1 locus lesions in aCLL selected for t(11;14) is as high as in MCL. Interestingly, rearrangements in the mTC1 (minor translocation cluster 1) were only found in aCLL. Therefore, the two B-cell chronic lymphoproliferative disorders share similar molecular rearrangements and the t(11;14) identifies a subset of B-CLL sharing molecular features with MCL and characterized by aggressive clinical evolution.

Acquired chromosome 11q deletion involving the ataxia teleangiectasia locus in B-cell non-Hodgkin's lymphoma: correlation with clinicobiologic features.

PURPOSE: To study the clinicobiologic significance of acquired 11q deletions involving the ataxia teleangiectasia locus (ATM+/-) in B-cell non-Hodgkin's lymphomas (NHL). PATIENTS AND METHODS: Fifty-three indolent lymphomas and 82 aggressive lymphomas were studied by conventional cytogenetic analysis and by fluorescence in situ hybridization using an 11q22-23 probe recognizing ATM sequences. Pertinent clinical data were collected. RESULTS: A hemizygous ATM deletion was seen in 44% to 88% of the interphase cells in 15 cases (11.1%); four patients had an indolent lymphoma (follicular center cell lymphoma), and 11 patients had an aggressive lymphoma (five mantle-cell lymphomas [MCLs] and six diffuse large-cell lymphomas). Dual-color hybridization studies showed ATM deletion to be possibly a secondary aberration in three patients with MCL. Ten out of 15 ATM+/- patients had a complex karyotype, 11 out of 15 had more than 90% abnormal metaphases (AA karyotype status), and +12, 13q14 deletion, and 17p13 deletion were seen in seven, four, and five cases, respectively. Patients with ATM+/- more frequently had a complex karyotype (P =.01) and the AA karyotype (P =.04) compared with patients without ATM+/-. With the exception of a poor performance status (P =.001), no correlation was found between ATM+/-, initial clinical variables, and complete remission rate; whereas a highly significant association was found with shorter survival (P <.0001). This cytogenetic lesion maintained its prognostic importance in multivariate analysis (P =.0004), along with performance status (P =.0006), serum lactate dehydrogenase level (P =.03), splenomegaly (P =.01), and histologic grade (P =.03). When analyzing indolent lymphomas and aggressive lymphomas separately, ATM+/- maintained its prognostic importance as an independent variable in both histologic groups (P =.0001 and P =.016, respectively). CONCLUSION: Though possibly not representing a primary genetic lesion in the majority of cases, the acquired ATM+/- status has clinicobiologic importance in NHL, possibly representing a major cytogenetic determinant of outcome.

Cytogenetic and molecular cytogenetic characterization of 6 new cases of idiopathic hypereosinophilic syndrome.

BACKGROUND AND OBJECTIVE: Idiopathic hypereosinophilic syndrome (HES) is defined as a peripheral blood eosiniphilia greater than 1, 500 cells/microL for longer than 6 months, absence of other apparent etiologies for eosinophilia and signs and symptoms of organ involvement. HES may be a reactive condition or a chronic myeloproliferative disorder but scanty information is available concerning its cytogenetic profile. DESIGN AND METHODS: Six patients with HES were studied by cytogenetic analysis. To increase the sensitivity of cytogenetic analysis, interphase FISH studies were performed to detect some cryptic chromosomal lesions involving the regions known to be frequently involved in myeloproliferative disorders (i.e. BCR/ABL, 5q31, 7q31.1, 11q23, 13q14, 17p13). Clinical parameters were recorded in all patients. RESULTS: A 3q deletion was detected in one patient; two unrelated clones with +14 and +11 were present in another patient who had a cryptic 5q31 deletion as disclosed by FISH; both patients had a mild clinical course. The 5q31 deletion was shown to involve the eosinophilic lineage and not the lymphoid cells. No chromosome abnormalities were found by karyotyping or interphase FISH in the remaining 4 cases. In two of these cases the clinical course was aggressive, with progressive leukocytosis and marked splenomegaly in one patient, central nervous system and cardiac involvement as well as bone marrow failure in the other. INTERPRETATION AND CONCLUSIONS: The 3q deletion, +11 and +14, and a cryptic 5q31 deletion involving the cells of the eosinophilic lineage are three novel chromosome abnormalities occurring in HES. We did not find a correlation between evolving or aggressive disease and the presence of chromosome anomalies. Our data confirm that HES is a clinically and biologically heterogeneous condition and suggest that more cases need to be studied to identify clinically significant chromosome changes in this rare condition. Some patients may benefit from treatment with interferon.

Four novel non-random chromosome rearrangements in B-cell chronic lymphocytic leukaemia: 6p24-25 and 12p12-13 translocations, 4q21 anomalies and monosomy 21.

Nine patients with previously unreported chromosome changes were identified among 209 B-cell chronic lymphocytic leukaemia (CLL) cases: three patients had a translocation involving 6p24-25; three had a 12p12-13 translocation; two had 4q21 involvement (one with coexisting 6p anomaly); and two had monosomy 21. Interphase fluorescence in situ hybridization (FISH) detected some cryptic aberrations (+12, 6q-, 17p-, 11q-) in those patients with 6p translocations, whereas only a cytogenetically undetected 13q14 deletion was found in the remaining cases. Atypical morphology was noted in six cases, including both cases with monosomy 21, two cases with 6p and 4q21 anomaly and one case with 12p involvement. Four of these cases also had more than one phenotype deviation with respect to the classical CLL phenotype. Disease progression after 21-51 months (median 41) was noted in two cases with 6p and 4q21 involvement and in one case with 12p anomaly and monosomy 21. We arrived at the following conclusions: (i) 6p24-25 and, possibly, 4q21 lesions represent non-random events in CLL, occurring in association with other well-known unbalanced rearrangements; (ii) 12p rearrangements and monosomy 21 may possibly represent early chromosome defects that are not associated with the classical DNA gains and losses known to be present in the majority of CLL; and (iii) atypical morphology and immunophenotype as well as disease progression were frequently observed in these cases

Assessment of distribution of CD34 epitope classes in fresh and cryopreserved peripheral blood progenitor cells and acute myeloid leukemic blasts.

BACKGROUND AND OBJECTIVE: So far several reports have described changes in the expression of surface antigens in progenitor cells and blasts following cryopreservation. However, there are no data on the effects of cryopreservation on the expression of the three CD34 epitope classes, and on their relationship with the clonogenic capacity of PBPC collected by leukapheresis. DESIGN AND METHODS: In order to analyze the effects of freezing/thawing procedures (Eth 80C storage for 3 months) and use of dimethylsulfoxide (DMSO) on the immunophenotype profile and colony production of peripheral blood progenitor cells (PBPC) in apheresis products derived from 20 patients with stage 0-III non-Hodgkin's lymphoma (nHL), a flow cytometry study was undertaken using different CD34 monoclonal antibodies (MoAbs) capable of recognizing the 3 epitope classes of CD34 molecule (class III: HPCA-2/FITC, HPCA-2/PE, 581/FITC, 581/PE; class II: Q-Bend 10/PE; class I: ICH3/PE, BI3C5-PE, Immu-133-PE). CD34 epitope expression was also analyzed in thawed CD34+ blasts obtained from 14 patients with acute myeloid leukemia (AML), who were analyzed using a larger number (#17) of CD34 epitope class I, II, and III reactive MoAbs. RESULTS: Under our experimental conditions it was found that class III and class II CD34 epitopes (differentially resistant to enzymatic cleavage with neuraminidase, chymopapain and glycoprotease) are better preserved than class I epitope Eth sensitive to degradation Eth after cell exposure to cryoprotectant DMSO and the freezing- thawing procedures. Results further showed a concomitant decrease in class I CD34+ counts and in BFU-E colony production. A significant increase in CD34 antigen expression levels (i.e. antibody binding capacity, ABC) by cryopreserved cells stained with CD34 epitope class III, and class II reactive MoAbs was also documented, while no changes after cryopreservation were noted using class I-reactive MoAbs. The slight increase in the percentage of CD34+ cells detected after frozen storage was correlated to a concomitant decrease in the number of more mature myeloid cells (CD15+, CD13+, CD33+). Compared to pre-cryopreservation values, a slight reduction in class I CD34 epitope expression was also found in thawed CD34+ AML blasts. INTERPRETATION AND CONCLUSIONS: As far as the reduction of class I CD34 epitope is concerned, it may be hypothesized that the freezing procedure, use of DMSO, and/or lysis methodology may either damage a CD34 subset, or induce distinct alterations of the CD34 glycoprotein, possibly determining a reduction in their immunoreactivity with some CD34 MoAbs. In conclusion, this study has shown that exposure to the cryoprotectant DMSO and the freezing/thawing procedures modifies the distribution of CD34 epitopes as well as the clonogenic capacity of PBPCs from nHL patients, and CD34+ blasts from AML. These findings need to considered when selecting CD34 MoAbs for enumeration and positive selection of stem/progenitor cells for research and clinical purposes.

13q14 deletion in non-Hodgkin's lymphoma: correlation with clinicopathologic features.

BACKGROUND AND OBJECTIVE: 13q14 deletion frequently occurs as a single anomaly in chronic lymphocytic leukemia (CLL) with favorable prognosis. This study was performed to assess the distribution of 13q14 deletion in non-Hodgkin's lymphoma (NHL) and to analyze its correlation with salient clinicopathologic features. DESIGN AND METHODS: One hundred and twenty-five NHL were analyzed by cytogenetics and by interphase fluorescence in situ hybridization (FISH), using a 13q14 cosmid probe recognizing DNA sequences between the Rb gene and the D13S25 marker. Clinical records all patients were surveyed. RESULTS: A 13q14 rearrangement was present in the stemline in 10 patients; 15 additional cases were shown by FISH to carry 13q14 deletion in 55-90% of the interphase cells, giving a 20% overall incidence for this anomaly. Six of 44 patients had a low-grade NHL, 14/28 had mantle cell lymphoma (MCL), 5/42 had a high grade NHL (p<0.0001). There was not correlation between 13q, karyotype status and complexity. A statistically significant association was found between 13q-, presence of splenomegaly and PB involvement, lower probability of attaining complete remission (CR) and shorter survival. These findings were not simply a function of the association of 13q- with MCL. In multivariate analysis, a complex karyotype had prognostic importance (p=0.0078), along with age (p=0.01), histology (p=0.001), LDH (p=0.03), PS (p=0.001), sex (p=0.03) and splenomegaly (p=0.02). INTERPRETATION AND CONCLUSIONS: 13q14 deletion represented an early chromosome change and showed a preferential association with MCL, though it was found in virtually all principal histologic subtypes, irrespective of clinical stage, karyotype status and complexity. Patients with 13q14 deletions had a low CR rate, suggesting that genes relevant to lymphomagenesis are located in this chromosome segment that warrants molecular cytogenetic investigation.

Cytogenetic profile of lymphoma of follicle mantle lineage: correlation with clinicobiologic features.

Conventional chromosome analysis (CCA) and interphase fluorescence in situ hybridization (FISH) was performed in 42 patients with mantle-cell lymphoma (MCL), with BCL1 rearrangement. The t(11;14)(q13;q32) or 11q abnormalities were detected by CCA in 34 cases, 20 of which had additional aberrations. A normal karyotype was observed in 8 cases. Probes detecting the chromosome aberrations that were observed in at least 3 cases by CCA, ie, +12, 13q14 deletion, and 17p deletion, were used for interphase FISH analysis. FISH detected total or partial +12, 13q14 deletion and 17p- in 28.5%, 52.4%, and 26% of the cases, respectively. The presence of these anomalies was not a function of karyotype complexity. Based on the results of CCA/FISH, three groups of increasing karyotype complexity were recognized: group 1, including 11 patients without detectable aberrations in addition to BCL1 rearrangement; group 2, including 14 patients with 1 to 2 additional anomalies; and group 3, including 17 patients with three or more additional anomalies. Clinical parameters associated with shorter survival were male sex (P =.006) and primary lymph-node involvement compared with primary bone marrow involvement (P =.015). Trisomy 12 was the only single cytogenetic parameter predictive of a poor prognosis (P =.006) and the best prognostic indicator was the derived measure of karyotype complexity (P <.0001), which maintained statistical significance in multivariate analysis (P<.0001). We arrived at the following conclusions: 13q14 deletion occurs at a high incidence in MCL; 17p deletion and total/partial +12 are relatively frequent events in MCL, the latter aberration being associated with a shorter survival; and the degree of karyotype complexity has a strong impact on prognosis in this neoplasia.