ALBERT

Beschreibung: Abstract 3435 Introduction Packed red blood cells in blood bank undergo a series of changes and this so-called “storage lesions” increases with time. It is believed that longer storage is associated with adverse transfusion-related complications, but the reason for this is not clear. Many bioactive substances are generated during blood storage and one or more of them may be responsible for adverse events. Among them, red cell microparticles (RMP) are a leading candidate for adverse effects, but conditions influencing their release are not well identified. Elucidation of these conditions is an important step toward minimizing storage lesions. Methods (I) MP generation: Non-leukoreduced and leukoreduced PC of known blood types (A+, B+, AB+, O+) were obtained from the blood bank within 2–4 days of collection, then stored at 4°C. Time of receipt was defined as day 0. At days 0, 10, 20, 30, and 40, forty mL samples were centrifuged at 1000×g for 20 min to remove cells. The supernatants were then assayed for subtypes of MP by flow cytometry comprising (a) RMP assessed by CD235a, (b) leukocyte MP (LMP) by CD45, (c) platelet MP (PMP) by CD41, and (d) generic MP by Ulex Europaeus (Ulex) or annexin V (AnV); MP-mediated procoagulant and proinflammatory activities were determined by TEG and CD11b expression, respectively. (II) Storage under anaerobic conditions: Anaerobic test units were processed by an OCDD (Oxygen Carbon dioxide Depletion Device) to deplete O2 and CO2, then stored anaerobically in an Anaerobic Storage Bag (ASB) [Transfusion 2011:51S, SP89]. After 42 days, MP were assayed as above. Results (a) Time of storage: In non-leukoreduced RBC, multiple MP types (PMP, LMP, RMP) were detected and seen to increase with time, but at distinctive rates. RMP increased little until day 10 when they rose exponentially; PMP counts rose steadily from day 0 and peaked at day 20; LMP showed little change until day 20 when they began to increase, then rose sharply after day 30. Levels of PMP (days 0 to 20) and RMP (days 20 to 40) correlated with increasing MP-mediated procoagulant and inflammatory markers. (b) Leukoreduction: Pre-storage leukoreduction decreased RMP generation by 20–40% and completely suppressed PMP and LMP generation. Leukoreduction decreased total MP-mediated procoagulant and inflammatory markers by 40–60%. This suggests that residual leukocytes and/or platelets potentiate RMP generation. (c) Residual platelet concentrations: We added increasing numbers of platelets to leukoreduced RBC, and then evaluated RMP generation during storage. The levels of RMP released were proportional to platelet counts in the storage bags. (d) Residual oxygen: We observed that storage of leukoreduced RBC under anaerobic conditions resulted in 40 – 60% reductions in RMP and annexin V+ MP generation measured at 42 days. Conclusions Multiple MP species (RMP, PMP, LMP) are released during storage of non-leukoreduced PC and increased with time. Procoagulant and proinflammatory activities increased in parallel. Leukoreduction eliminates LMP and PMP generation and reduces RMP generation by 40–50%. This was accompanied by reduction of procoagulant and proinflammatory activities by 60%. We have identified residual platelets, leukocytes, and oxygen levels as important factors governing MP release in stored blood. Reduction or elimination of factors influencing MP generation such as residual platelets, leukocytes and oxygen levels would improve future blood storage condition. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: Small cell volume or diameter (3–7μm) appears to be one of the characteristics that may be used to identify early stem cells. Hematopoietic stem cells may be characterized by the expression of cell surface markers such as CD34, CD117 and CD133. In addition, stem cells such as “SP cells” can be recognized by the efflux of dyes such as Hoechst 33342 and presence of aldehyde dehydrogenase (ALDH). In prior studies we used the Beckman Coulter® Cell Lab Quanta(™) SC MPL analyzer to determine the electronic cell volume, diameter and marker expression (CD34, CD90, CD117 and CD133) of cells from peripheral blood apheresis (HPC, Apheresis) samples (Shariatmadar et al. Cytometry B, 74B:182–188, 2008; Sharma et al. Cytometry A, 73A:160–167, 2008). In the present study, we used this analyzer to determine cell volume, diameter and marker expression of ALDHpositive stem cells from HPC, Apheresis samples of 44 patients with hematologic malignancies mobilized with granulocyte colony-stimulating factor. The mean electronic volume of the ALDHpositive cells was 286μm3 (SD±27) corresponding to a diameter of 8.2μm. The mean percentage of ALDHpositive cells analyzed was 0.51% (SD± 0.004). CD34 expression was seen in 0.13% (SD±0.001) of the ALDHpositive cells. The mean electronic volume of the ALDH and CD34 positive cells was 270μm3 (SD± 33) corresponding to a diameter of 8.0μm. CD133, CD117 and CD90 expression was seen in 0.28% (SD± 0.01), 0.17 (SD± 0.05) and 0.04% (SD± 0.03) of the ALDHpositive cells. The mean electronic volume of the ALDHpositive cells with CD133 expression was 275μm3 (SD±29) corresponding to a diameter of 8.1μm while that of the ALDHpositive cells with CD117 and CD90 expression was 284μm3 (SD± 32) and 265μm3 (SD± 35), corresponding to a diameter of 8.15 and 7.9μm. Stem Cell Marker Electronic Volume (μm3) Diameter (μm) CD90 (Thy-1) 300μm3 (SD± 38.5) 8.3μm (95% CI=8.1–8.4) CD117 (c-Kit) 349μm 3 (SD± 47.4) 8.7μm (95% CI=8.5–8.8) CD133 (Sca-1) 322μm3 (SD± 44.5) 8.5μm (95 % CI= 8.3–8.6) ALDH+ 286μm3 (SD± 27.0) 8.2μm (95 % CI= 8.1–8.3) ALDH+CD34+ 270μm3 (SD± 33.0) 8.0μm (95 % CI= 7.8–8.2) ALDH+ CD90+ 265μm3 (SD± 35.0) 7.9μm (95 % CI= 7.8–8.1) ALDH +CD117+ 284μm 3 (SD± 32.0) 8.15μm (95 % CI= 7.9–8.2) ALDH +CD133+ 275μm3 (SD± 29.0) 8.1μm (95 % CI= 8.0–8.3) The use of electronic cell volume in conjunction with side scatter might provide a useful method for characterization of stem/progenitor cell populations with specific marker expression. In our study, hematopoietic stem cells expressing ALDH and bearing early stem cell markers were relatively larger than those previously reported. Future studies will aim to characterize the long term repopulating capability of this fraction of stem cells in in-vivo models.

Beschreibung: BACKGROUND. Endothelial damage is a prominent feature of TTP, evidenced by elevated EMP levels during acute episodes of TTP. However, endothelial repair in TTP remains to be investigated. ECFC are key players in endothelial regeneration, and their assay in blood has been employed to monitor endothelial remodeling. We investigated circulating ECFC in TTP and their relationship with EMP. METHODS. Six patients with acquired TTP were studied. All were treated with therapeutic plasma exchange (TPE). All but one underwent complete remission (CR). The one who failed to achieve CR with TPE suffered multiple strokes and developed chronic renal failure. We measured the following in acute phase and remission, as well as pre- and post-TPE: EMP defined by CD31+/CD42b- (EMP31) or CD62E+ (EMP62) by flow cytometry; ECFC in blood by the method of Ingram et al [Blood 104:2752, 2004]; ADAMTS13 activity in blood as well as in supernatants of ECFC-derived EC by the FRETS-vWF73 fluorogenic method. In addition, we investigated effects of microparticles (MP) from TTP plasma on normal ECFC. In separate experiments, EC cultured from TTP patients were assayed for surface expression of CD106 and CD54, and EMP released in response to TNF-α. RESULTS. In the acute phase, EMP were elevated: EMP31 (1.89 ± 0.65 × 106 counts/mL vs. 0.28 ±0.1 for controls, p=0.001) and EMP62 (5.7 ±1.3 × 106 counts/mL vs. 0.40 ±0.12 for controls, p

Beschreibung: Purpose: To report our experience with large volume leukapheresis (LVL) for allogeneic hematopoietic progenitor cell (HPC) transplantation, evaluating its safety and efficacy in 69 consecutive healthy donors. Methods: Donor medical records from 1995–2004 were reviewed. These consisted of 38 males, 31 females, age range 16–65 (median: 37). All were mobilized with G-CSF, based on a dose of 7.5 mcg/kg/day (median: 600 mcg, range 300–960 mcg) for four days prior to outpatient LVL (Cobe spectra). Vascular access was femoral vein in 59 and antecubital veins in 10 donors. Pre- and post-LVL determinations included: WBC count, hemoglobin, hematocrit, platelet count, serum potassium, and calcium (Ca++). Oral KCL and oral/intravenous Ca++ were given as indicated by laboratory values and symptoms. Catheter-related complications and blood product requirements associated with collection were assessed. Final CD34/kg recipient body weight (rec. BW) and recipient engraftment data was evaluated. Results: LVL was well tolerated by all donors undergoing a total of 74 procedures. Data/LVL Procedure Mean Range Procedures per patient 1 1–2 Blood volume processed (L) 24.2 15.0–34.4 Time (minutes) 273 180–386 Flow rate (ml/min) 92 65–100 ACD–A used (ml) 1008 636–1449 Heparin used (units) 3000 Not applicable CD34/kg rec. BW 8.2 1.36–33.9 Adverse reactions were minor and occurred in 40% of the donors. Local parasthesias accounted for majority (86%) of the adverse events. Bone pain secondary to G-CSF accounted for the remainder of the cases and was well controlled with analgesics. Oral electrolyte supplements were administered in 61% of the donors. Eleven (15.9%) required intravenous Ca++ during collection. There were no catheter-related complications and no requirement for blood products during or after collection. Target values of 2 x 106 CD34/kg rec. BW was obtained in 68 cases (98.6%). Engraftment was achieved in 92% of the cases (6 died prior to engraftment). There were no early or late graft failures. Median time to ANC recovery was 10 days (range 8–25 days). Self-sustained platelet count was achieved in a median of 13 days. Conclusion: LVL was a safe and efficacious method of allogeneic HPC collection. The use of a lower G-CSF dose for mobilization is more economical and well tolerated. This dose allows for adequate collection of CD34 positive cells. These cells support the sustained recovery from high-dose chemotherapy for allogeneic transplantation.

Beschreibung: Abstract 342 Background. Packed red cells (PC) stored in blood bank undergo a series of changes, so-called “storage lesion”, which increase with time. In view of some but not all recent studies, it is widely believed that transfusion with younger blood carries less risk of adverse reactions than older blood. However, there is no agreement on the “safe” age of blood, nor is it clearly understood why older blood may carry increased risks. The purpose of this study was to identify microparticle-related factors in stored PC at different time intervals that might pose risk of adverse effects. We investigated profiles of cell-derived microparticles (MP), particularly RBC-derived MP (RMP), in stored PC and assessed their procoagulant and inflammatory property. Methods. Twelve bags of fresh packed red cells (PC) of known blood types (A+, B+, AB+, O+) were obtained from blood bank (2-4 days since drawing). All were non-leuko depleted and were stored at 4°C. Time of receipt was considered day 0. At intervals of 0, 10, 20, and 30 days, 40 mL was withdrawn and centrifuged at 1000xg for 20 min to remove cells. The supernatants were then assayed for (1) quantity of different species of cell-derived MP by flow cyteometry comprising (a) RMP defined by CD235b; (b) LMP by CD45; (c) PMP by CD41; (d) EMP by CD144; (e) generic MP by Ulex Europaeus (Ulex) or Annexin V (AnV), (2) procoagulant activity by MP-mediated thrombin generation assay (TGA); (3) MP-mediated proinflammatory activity by CD 11b expression in neutrophils following incubation with RMP. Results. (1) MP Profiles. The time-course of generation of the MP subtypes varied considerably. For RMP, there was little increase before day 10, but then rose rapidly with time, to 180% at 20 days, and to 450% at 30 days. Small amounts of MP derived from leukocytes (LMP), platelets (PMP), and endothelia (EMP) were present in all bags at day 0, generally

Beschreibung: BACKGROUND: Blood transfusion (Tx) carries greater risks of adverse events (AEs) than previously appreciated. These adverse effects include higher incidence of post-surgical infections, longer hospital stay, higher mortality, more frequent serious adverse events (SAE’s), and generally poorer surgical outcomes. Accordingly, ameliorating these adverse effects constitutes an urgent challenge to medical science. Factors responsible for Tx-related adverse events (AE’s) are not well understood. Many potentially toxic substances are released during blood storage, and many of them have been implicated or postulated as culprits. Washing of packed RBC remove these products and may ameliorate transfusion-related AE’s. Benefits of washed RBC are well established for pediatric surgical patients, chiefly by preventing hyperkalemia, but use of washed RBC in adult surgical patients has not heretofore been systematically investigated. We here report results of a prospective randomized study directly comparing surgical outcomes, in terms of mortality and AE’s, between groups of adult CABG patients transfused with either washed or unwashed (conventional) RBC. METHODS: A prospective randomized study of 148 patients undergoing coronary artery bypass graft (CABG) was conducted. Fifty-eight patients were randomized to receive unwashed (conventional) RBC (UW group) and 41 to washed RBC (W group). The remaining 49 did not require Tx. The main in-hospital outcomes recorded included mortality, serious adverse events (SAE’s), non-serious adverse events (AE’s), and SOFA scores pre- and post-surgery. A telephone interview was conducted at day 30 post-discharge, and mortality at one-year was also assessed. The statistical techniques used for the comparison of the UW and W RBC groups included: independent sample t-tests for variables with normal or approximately normal distribution; Mann-Whitney tests for variables with skewed distributions and for ordinal variables; chi-squared tests or Fisher’s exact tests for discrete variables; and logistic regression model for assessing different factors as predictors of the occurrence of each kind of event. RESULTS: Between the 2 groups, demographic, clinical, and comorbidity data were similar and there was no statistically significant difference in number of serious AE’s (SAE’s). However, 4 of 6 patients died from SAE’s in the UW group but all 7of 7 with SAE in the W group survived. The in-hospital mortality was greater in the UW group (4 vs. 0, p = 0.149) but 1-year post-op mortality was significantly higher in UW group (7 vs. 0, p=0.036). Frequency of less serious AE’s was higher in UW group in every category. Negative binomial regression analyses showed that, after adjusting for comorbidities, UW-group are likely to experience 64% more AEs (p= 0.027). The 30-day follow-up showed similar trends of higher AE’s in UW-group, but only CNS-related AE’s were significant (30 vs. 5, p

Beschreibung: Abstract 3373 Introduction Several studies have indicated that transfusion with older blood carries more risk of adverse reactions than transfusion with younger blood, but this remains controversial. It is not clear why older blood may carry increased risks, or what the “safe age” of stored blood is. It is known that multiple bioactive substances are generated from blood during storage, and one or more of these substances may be involved in transfusion-related complications. Among them, MP are a recognized marker of the storage lesion, and their involvement in transfusion-related complications has been postulated. However, questions such as MP species, quantity, biological activity, and factors affecting their release are not well elucidated. The purpose of this study was to quantify MP species and their activity in stored RBC as a function of storage time, and to evaluate the impact of leukoreduction and residual platelets on MP release. Methods (I) MP generation and functional activity Thirty-four bags of packed RBC (16 non-leukoreduced, 18 leukoreduced) of known blood types (A+, B+, AB+, O+) were obtained from the blood bank within 2–4 days of drawing, and then stored at 4°C. Time of receipt was defined as day 0. At days 0, 10, 20, 30, and 40, 40 mL samples were centrifuged at 1000xg for 20 min to remove cells. The supernatants were then assayed for: (1) subtypes of MP by flow cytometry comprising (a) red cell MP (RMP) assessed by CD235a, (b) leukocyte MP (LMP) by CD45, (c) platelet MP (PMP) by CD41, (d) endothelial MP (EMP) by CD144, and (e) generic MP by Ulex Europaeus (Ulex) or Annexin V (AnV); (2) MP-mediated thrombin generation assay (TGA); (3) MP-mediated inflammatory activity by CD 11b expression in neutrophils following incubation with MP. (II) Reconstitution of increasing platelet counts in leukoreduced RBC. To investigate the effect of residual platelets on RMP generation, we mixed a constant amount of RBC with increasing amounts of type-matched platelets (0 to 250,000/μL f.c.) in standard storage bags and measured time-dependent MP release. Results (A) Time-course of MP generation (i) Non-leukoreduced. RMP, PMP and LMP all increased with time, but with different patterns. RMP increased little to day 10 but then rose exponentially, and by day 40 they were 4–6 fold higher than at day 0. PMP counts rose steadily from day 0 and peaked at day 20, being 2–3 fold higher than at day 0. LMP showed no significant change until day 20 when they started to increase, and then increased sharply after day 30, and by day 40 were 1.5–2 fold higher than at day 0. Levels of PMP (days 0 to 20) and RMP (days 20 to 40) correlated with increasing MP-mediated procoagulant and inflammatory markers. (ii) Leukoreduced. Pre-storage leukoreduction decreased RMP generation by 20–40%, completely suppressed PMP and LMP generation, and reduced total MP-mediated procoagulant and inflammatory markers by 40–60%. CBC showed that leukoreduction not only removed 〉99% WBC but also reduced residual platelets by 〉95% (from 90 ±30 ×103/μL to 3.5 ±1.3 ×103/μL). This suggests that residual leukocytes and platelets potentiate RMP generation. (B) Effects of residual platelets on RMP generation. To further study the effects of platelets on RMP generation, we mixed known counts of platelets with leukoreduced RBC, and then evaluated RMP generation over time. We found that RMP levels released were proportional to the platelet counts, as were the procoagulant and inflammatory markers. These results show that platelets in stored RBC play a key role in RMP generation. Conclusion Multiple MP types (PMP, LMP, RMP) are released during storage, and their levels increase over time but their patterns of change are different. Procoagulant and inflammatory markers increase in parallel with PMP and RMP. Our data support the hypothesis that age of stored blood could be important in transfusion-related complications, via MP production. Leukoreduction sharply reduces MP generation and procoagulant and inflammatory markers, suggesting that known benefits of leukoreduction may be attributable to reduced MP production. The finding that residual platelets in stored RBC can potentiate RMP generation suggests that minimizing platelets in non-leukoreduced packed cells could reduce the risk of transfusion-related complications. (Supported by NIH grant 1R01HL098031). Disclosures: No relevant conflicts of interest to declare.

Beschreibung: Introduction Blood transfusion is associated with serious adverse events (SAE) such as poor surgical outcome, higher surgical mortality. The cost of transfusion is escalating due to increasing demand and limited blood supply. There is a worldwide growing interest in improving blood utilization methods and storage. Identification of reliable predictors of surgical bleeding is of upmost importance in order to reduce the number of unnecessary transfusions, save limited blood resources, and lower the related costs. Cell derived microparticles (MP) are small membrane vesicles of 0.1 µm in size, released during cell activation and apoptosis. They play an important role in hemostasis and thrombosis. Methods Data presented herein are from an NIH funded prospective randomized study to investigate role of cell derived MP in surgical complications in CABG. Of a total of 122 patients, 81 received transfusion during and/or after surgery (Tx group) whereas the remaining 41 did not (NoTx group). In addition to routine laboratory tests, special assays were performed pre-surgery including concentrations of MP from platelets (PMP), red cells (RMP), endothelia (EMP), and leukocytes (LMP). The two groups were compared with respect to pre-surgical variables in order to assess potential predictors of the need for surgery. Results There were no significant differences between the two groups with respect to most of the pre-surgical demographic, clinical, and lab variables, except that Tx group was associated with higher incidence of female gender, type O blood group, and heparin use. On the other hand, the NoTx group was associated with higher pre-surgical levels of RMP and PMP. Using stepwise logistic regression analyses, four factors were identified as the most significant predictors of the need for transfusion. As depicted by an the areas under the curve (AUC) of the ROC curve, the 95% Confidence Interval of the AUC, and their corresponding p-value for each factor, were as follows. For the model combining all four variables, the AUC and its 95% CI were 0.86 [0.78 - 0.94]. In addition, analysis of post-surgery data revealed better outcomes for the NoTx group in terms of complications such as hours on mechanical ventilation (mean ± SD: 9.2 ± 7.0 vs 15.5 ± 17.1, p = 0.005), number of SAE other than death (0 vs 9 (11.1%), p =0.028), and number of in-hospital deaths (0 vs 5 (6.1%), p= 0.167). Discussion This is the first report to show that pre-surgical levels of circulating MP are highly significant predictors of need for transfusion in CABG. This finding is consistent with other evidence that MP play a major role in hemostasis. Implementation of pre-surgery MP assay will greatly imporove assessment of risk of bleeding in surgery, and requirements for transfusion. This could reduce the number of transfusions, with corresponding improvement in patient outcomes, as well as reduced costs and strains on limited blood supplies. Furthermore, it may be feasible to manipulate pre-surgical MP levels in patients to minimize bleeding and Tx requirements. This could be done by increasing rate of MP generation or retarding rate of clearance, or both. Alternatively, MP could be produced in vitro (autologous or not) for infusion during surgery to minimize blood loss. Such approach is in progress [Jy, et al Thromb Haemost, in press]. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: INTRODUCTION: Therapeutic plasma exchange (TPE) is a widely accepted therapy for TTP. A leading hypothesis for its benefit is that TPE replenishes ADAMTS13 with fresh frozen plasma (FFP) and removes inhibitory antibodies. We have reported that endothelial microparticles (EMP) positive for ULvWF are elevated in acute TTP, decline with repeated TPE, and can inhibit ADAMTS13. Presence of platelet microparticles (PMP) in FFP has been described. This led us to the hypothesis that the benefit of TPE is partly due to its effect on microparticles (MP). We investigated the effect of TPE on MP and ADAMTS13 in TTP. METHODS: Nine patients were studied longitudinally during an episode of TTP. Blood samples were obtained upon admission and daily pre- and post-TPE. Additionally, the infused and removed plasma were studied. EMP defined by CD31+/CD42b− (EMP31), CD62E+ (EMP62) and CD62E+/vWF+ (EMPVWF); PMP were defined by CD31+/CD42b+ (PMP42) and CD41+ (PMP41), all measured in platelet-poor plasma by flow cytometry; concentrations are in millions / mL. ADAMTS13 activity was assayed by a FRETS-vWF73 method of Kokame et al [Br J Haematol2005; 129:93]. vWF multimers were analyzed following Raines et al [Thromb Res1990; 60:201]. RESULTS: At admission, TTP patients exhibited very low ADAMTS13 levels, 0–10% of controls (ctl), and elevated EMP31 (2.5 ±0.7 vs. 0.35 ±0.09 ctl) and EMP62 (6.3 ±1.9 vs. 0.25 ±0.07 ctl), both p

Beschreibung: INTRODUCTION: Therapeutic plasma exchange with infusion of FFP (TPE/FFP) is a standard therapy for TTP. However TPE requires insertion of central lines and is associated with increased morbidity such as line-related infections and sepsis. Danazol has two relevant properties: immune modulation and enzyme induction. The former was exploited in autoimmune diseases such as ITP and the latter in disorders of enzyme deficiencies such as hereditary angioneurotic edema. TTP is an autoimmune disorder with deficiency of the enzyme, ADAMTS13. We investigated danazol therapy in patients with TTP and measured ADAMTS13 activities prior to and post danazol. METHODS: We studied 7 patients (pts) with diagnosis of TTP, and 10 non-TTP pts (4 ITP, 2 thrombocytopenias of other cause, 2 autoimmune hemolytic anemia, 2 lympho- or myeloproliferative disorders). Both TTP and non-TTP patients were treated with danazol. All 7 TTP patients were female with mean age of 42. Four of the TTP patients (Group A) failed or had complications with TPE/FFP, requiring over 2 wks of hospitalization prior to starting danazol. The new regimen includes (1) administration of danazol at 200 mg 2–4 times daily, (2) removal of central lines and discontinuation of TPE when platelet counts stabilized, (3) infusion of FFP through peripheral vein. While monitoring platelet counts and LDH, FFP infusion was gradually tapered and stopped and danazol was continued for an additional 8–20 wks. In the remaining pts with TTP (Group B), danazol was started on admission along with TPE in two pts and FFP alone in one. ADAMTS13 was assayed by the FRETS-vWF73 method [Brit J Haem2005; 129:93] before and after 2–5 wks of danazol. Parameters evaluated were clinical and laboratory improvement, complications, requirement for TPE and FFP, and levels of ADAMTS13 activity. RESULTS: All 4 pts in Group A, who failed or had complications with TPE/FFP, responded well to the regimen, requiring fewer TPE and FFP infusions. The mean number of TPE was reduced by 88% post-danazol and mean number of FFP reduced by 79%. Their hospital stays were shorter. The 3 pts (Group B) who received danazol on admission also responded well. Patient B#1, with 20 yr history of TTP and numerous recurrences, required 1–2 weeks of hospitalizations with the new regimen, compared to 3–8 wks in her previous admissions. Patient B#3 who had FFP infusion alone had an excellent response but danazol had to be stopped because of abnormal liver function; she then relapsed necessitating TPE/FFP. ADAMTS13 activities were measured in 5 TTP pts: mean activity rose from 25.2% pre-danazol to 80.2% post (p=0.007). In the 10 non-TTP pts, the mean activity increased from 61.1% to 75.0% (p=0.03) pre-/post-danazol. CONCLUSION. Danazol with FFP infusion was effective in all TTP pts. Danazol significantly reduced requirement of TPE and infusion of FFP and increased ADAMTS13 activity in both TTP and non-TTP patients. Use of danazol with FFP infusion through peripheral veins avoided complications of central lines required for TPE and reduced hospital stays. We suggest that early addition of danazol and early switching from TPE/FFP to the new regimen would benefit patients with TTP. However, a larger-scale randomized prospective study is needed to rigorously evaluate the efficacy and indications for this regimen.