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BACKGROUND: The wide diffusion of multicomponent collection in donor apheresis has led to the yielding of different components, such as plasma-reduced platelet-pheresis at high PLT concentration. We investigated whether this collection modality could induce more PLT activation compared to standard plateletpheresis. STUDY DESIGN AND METHODS: Forty-one plateletpheresis collections (20 Trima and 21 Spectra LRS Turbo v.7.0, COBE) were evaluated. Donor, procedure, and product data were recorded. ADP, collagen, and U46619 (a thromboxane-A2 analog)-induced PLT aggregation was investigated in basal (donor) and final (plateletpheresis unit) samples. The expression of PLT activation marker P-selectin (CD62P) was studied using flow cytometry in basal and final samples. In all cases, P-selectin was investigated in final samples after stimulation with ADP to assess for a possible further release of the antigen. Four additional plateletpheresis procedures were performed in donors from Group A, using the traditional, nonplasma-reduced program. RESULTS: Plateletpheresis obtained by means of the Trima device showed a lower response to in-vitro induced PLT aggregation and a higher percentage of P-selectin-expressing PLT when compared to products obtained using the Spectra device. Moreover, P-selectin release after ADP stimulation was reduced in plateletpheresis units obtained using the Trima device. These differences disappeared when a nonplasma-reduced collection program was used. In-vivo evaluation did not detect any difference between platelepheresis obtained by means of the two cell separators. CONCLUSIONS: Plateletpheresis units obtained by means of multicomponent collection show a higher degree of PLT activation compared to traditional plateletpheresis procedures when high-concentration plasma-reduced products are collected. Randomized clinical studies are needed to assess the real impact of these findings in terms of in-vivo efficacy of plasma-reduced plateletpheresis units.

We investigated the possible involvement of vascular damage in the pathogenesis of Alzheimer's disease (AD), by assessment of plasma levels of tissue factor pathway inhibitor (TFPI), a serine protease inhibitor induced by endothelial injury, and homocysteine (Hcy), a known risk factor for cerebrovascular disorders, folate levels were also measured. 110 probable AD, 38 mild cognitive impairment, 31 patients affected by idiopathic Parkinson's disease (without dementia) and 100 healthy controls, who displayed no vascular disorders were enrolled. TFPI and Hcy were significantly higher in AD patients with respect to other groups. The levels of TFPI and Hcy were positively correlated in hyperhomocysteinemic AD and mild cognitive impairment subjects, and were negatively correlated with folate levels. Our findings suggest that an impairment of endothelial function associated with high Hcy levels may occur in AD patients, despite the absence of manifest cerebrovascular lesions. Therefore, TFPI may represent a candidate marker of endothelial damage in AD and might be used for the identification and monitoring of patients that would benefit from folate supplementation treatment.

Circulating endothelial cells (CECs) have been studied in cardiovascular disorders and as a marker of angiogenetic activity; clinical applications are limited by a lack of consensus on their phenotypic identification and quantification. We determined CECs in essential thrombocythemia (ET) patients, to investigate their possible pathogenetic role. We considered CECs as CD146⁺/CD45⁻ nucleated cells, detected in peripheral blood from 21 healthy controls and 39 ET patients, performing a combination of pre-enrichment of CD146⁺ circulating cells and multiparametric flow cytometry measurement (FCM). Levels of CECs in ET patients were higher with respect to controls (median 2844 CECs/mL vs. 121.3 CECs/mL, P < 0.0001). Apparently hydroxyurea treatment did not influence the levels of CECs. As another established marker of endothelial activation, we also assessed soluble E-selectin (sE-selectin) levels in 31 of the ET patients and compared with 39 healthy volunteers: median sE-selectin level in ET patients was 35.3 ng/mL, higher with respect to controls (24.48 ng/mL), P = 0.0369. Our data suggest that endothelium in ET is activated, reflecting a significant role of angiogenesis in this disorder and suggesting an important endothelial contribution in the hypercoagulable state of ET patients.

Transfusion-related acute lung injury (TRALI) is the leading cause of transfusion-associated mortality. Antibodies against human leucocyte antigens (HLA) and human neutrophil antigens (HNA) are often detected in the implicated donors. We investigated the incidence and aetiology of TRALI in Lombardy. Moreover, we determined the rate of HLA and HNA alloimmunisation and the HNA genotype in a cohort of local blood donors.During a 2-year observational study in eight blood transfusion services, suspected TRALI cases were collected and characterised by means of HLA and HNA antibody screening of implicated donors, donor/recipient cross-matching and HLA/HNA molecular typing. In addition, 406 Italian donors were evaluated for alloimmunisation and in 102 of them HNA gene frequencies were determined.Eleven cases were referred to the central laboratory, of whom three were diagnosed as having TRALI, seven as having possible TRALI and one as having transfusion-associated circulatory overload. Seven TRALI cases were immune-mediated whereas in three we did not find either alloantibodies in implicated donors or a positive reaction in the cross-match. The most frequently implicated blood component was red blood cells (in 5 males and in 1 female), whereas four cases of TRALI were associated with transfusion of fresh-frozen plasma (in 3 females and in 1 male). The frequency of reported TRALI/possible TRALI cases was 1:82,000 for red blood cells and 1:22,500 for fresh-frozen plasma. No cases were observed for platelets. Overall, the frequency of HLA or HNA alloimmunisation in blood donors was 29% for females and 7% for males. The latter could be related, at least in part, to natural antibodies. HNA gene frequencies showed that HNA-1b is more frequent than HNA-1a in our sample of donors.The recently adopted national policy to prevent TRALI, i.e. using only plasma donated by males, would have had a positive impact in our setting.

The course of early stage CLL patients may be very different with patients who never need therapy and other who need therapy at different times. The presence or absence of somatic mutations in the immunoglobulin heavy-chain gene of the CLL cell (mutational status) was recently identified as one of the most predictive factors for the time from diagnosis to initial treatment. CD-38 and ZAP-70 were also shown to have a similar prognostic value and they were sometimes considerate as a “surrogate” markers of mutational state. This study was aimed to evaluate the prognostic impact of different prognostic factors on time from diagnosis to initial treatment in this subgroup of CLL patients. We studied 127 patients (71 M; 56 F); median age was 64,3 (range 42 – 84). All they had a stage A CLL at diagnosis. When Rai stage was considered, 83 had stage 0, 32 stage I and 12 patients stage II. Median follow-up was 65 months (range 6 – 232 months). Absolute lymphocyte count was: median 19 cells × 10^9/l (range 2,5 – 99). Median hemoglobin was 13,82 g/dL (range 11,3 g/dL – 17,5 g/dL). Median platelet count was 209 × 109/L (range 102 × 109/L e 533 × 109/L.). Patients were treated when symptomatic or progressive disease developed, according to National Cancer Institute Working Group criteria. The following cut-off values were used: CD38 >20%; ZAP-70 > 20%; VH unmutated: >98% homology with germ-line sequence. 26% of patients were CD38 positive, 28% were ZAP-20 positive; 24% patients were VH unmutated. 39/127 patients included in the study had received one or more chemotherapy regimens. First line therapy was chlorambucil in 28 patients, fludarabine in six, different therapies (cyclophosphamyde, prednisone, rituximab) in 5. Median interval between diagnosis and initial therapy was 30 months (range 1–96). 22 % of Rai 0 pts, 44 % of Rai 1 pts 50% of Rai 2 pts were treated. 65% of ZAP-70 positive were treated vs 18% of ZAP-70 negative. 55% of unmutated were treated vs 18% of mutated pts. 42% of CD-38 positive were treated vs 26% of CD-38 negative. When the interval from diagnosis to therapy was analyzed with a Kaplan-Meier survival curve (see figures), mutational status and ZAP-70 positivity were the most significant prognostic factors. Median treatment-free time was 35 months for ZAP-70 negative patients vs. 190 months for ZAP-70 positive patients (p= 0,001). Median treatment-free time was 40 months for unmutated vs. 190 months for mutated patients (p= 0,002). When CD-38 was analyzed, a borderline significant value was obtained (p=.0.08) with median treatment-free interval of 70 months in CD-38 positive patients vs. 98 months in CD-38 negative patients. Absolute lymphocyte count (with a cut-off at 15 cells × 10^9/l) also showed a significant impact on the need of treatment. Time from diagnosis to initial treatment and IgVH omology Time from diagnosis to initial treatment and IgVH omology Time from diagnosis to initial therapy and ZAP 70 Time from diagnosis to initial therapy and ZAP 70