Cryopreserved buffy-coat-derived platelets reconstituted in platelet additive solution: A safe and available product with sufficient haemostatic effectiveness.

BACKGROUND: Platelets (PLTs) stored at 20-24 °C have a short shelf life of only 5 days, which can result in their restricted availability. PLT cryopreservation extends the shelf life to 2 years. METHODS: We implemented a method of PLT freezing at -80 °C in 5-6% dimethyl sulfoxide. Buffy-coat-derived leucodepleted fresh PLTs blood group O (FP) were used for cryopreservation. Cryopreserved pooled leucodepleted PLTs (CPP) were thawed at 37 °C, reconstituted in PLT additive solution SSP + and compared to FP regarding PLT content, PLT concentration, pH, volume, PLT loss, anti-A/B antibody titre, total protein, plasma content, and PLT swirling. Clot properties were evaluated via rotational thromboelastometry. PLT microparticle number and surface receptor phenotype were assessed via flow cytometry. RESULTS: CPP met the required quality parameters. The mean freeze-thaw PLT loss was 22.24 %. Anti-A/B antibody titre and plasma content were significantly lower in CPP. CPP were characterised by faster clot initiation and form stable PLT clots. The number of PLT microparticles increased 25 times in CPP and there were more particles positive for the activation marker CD62 P compared to FP. CONCLUSION: Thawing and reconstitution are easy and fast processes if platelet additive solution is used. Low anti-A/B antibody titre and plasma content make possible the use of CPP of blood group O reconstituted in SSP + as universal ABO products, including clinical situations where washed PLTs are required. Clot properties evaluated via rotational thromboelastometry demonstrated that CPP retain a significant part of their activity compare to FP and are haemostatically effective.

The use of platelet-rich fibrin in the surgical treatment of medication-related osteonecrosis of the jaw: 40 patients prospective study.

OBJECTIVES: Medication-related osteonecrosis of the jaw (MRONJ) is defined as exposed bone in the maxillofacial region persisting for more than eight weeks in patients who are or were treated with antiresorptive or antiangiogenic agents and had no radiation therapy to the craniofacial region or obvious metastatic disease of the jaws. It is a recognised side effect of antiresorptive or antiangiogenic medication. To date, there is no specific gold standard treatment for MRONJ cases. The aim of this study was to evaluate the successful rate of surgical treatment with adjuvant local application of platelet rich fibrin. METHODS: 40 patients treated with necrotic bone resection and adjuvant local application of platelet-rich fibrin (PRF) were included. Treatment outcomes were evaluated after 12 months. RESULTS: The outcome of surgical treatment was successful in 34 of all 40 patients (85%), in 12 months follow-up. If we evaluate only cases where removal of all necrotic bone was possible the success rate was increased to 94%. A significant association between size of necrotic bone and treatment response was found (P=0.014, Wilcoxon rank sum test with continuity correction). CONCLUSIONS: Surgical treatment of MRONJ with adjuvant local PRF application proved to be very effective and safe, especially in early stages when all necrotic bone can be easily removed.

Dendritic Cell-Based Immunotherapy in Advanced Sarcoma and Neuroblastoma Pediatric Patients: Anti-cancer Treatment Preceding Monocyte Harvest Impairs the Immunostimulatory and Antigen-Presenting Behavior of DCs and Manufacturing Process Outcome.

Despite efforts to develop novel treatment strategies, refractory and relapsing sarcoma, and high-risk neuroblastoma continue to have poor prognoses and limited overall survival. Monocyte-derived dendritic cell (DC)-based anti-cancer immunotherapy represents a promising treatment modality in these neoplasias. A DC-based anti-cancer vaccine was evaluated for safety in an academic phase-I/II clinical trial for children, adolescents, and young adults with progressive, recurrent, or primarily metastatic high-risk tumors, mainly sarcomas and neuroblastomas. The DC vaccine was loaded with self-tumor antigens obtained from patient tumor tissue. DC vaccine quality was assessed in terms of DC yield, viability, immunophenotype, production of IL-12 and IL-10, and stimulation of allogenic donor T-cells and autologous T-cells in allo-MLR and auto-MLR, respectively. Here, we show that the outcome of the manufacture of DC-based vaccine is highly variable in terms of both DC yield and DC immunostimulatory properties. In 30% of cases, manufacturing resulted in a product that failed to meet medicinal product specifications and therefore was not released for administration to a patient. Focusing on the isolation of monocytes and the pharmacotherapy preceding monocyte harvest, we show that isolation of monocytes by elutriation is not superior to adherence on plastic in terms of DC yield, viability, or immunostimulatory capacity. Trial patients having undergone monocyte-interfering pharmacotherapy prior to monocyte harvest was associated with an impaired DC-based immunotherapy product outcome. Certain combinations of anti-cancer treatment resulted in a similar pattern of inadequate DC parameters, namely, a combination of temozolomide with irinotecan was associated with DCs showing poor maturation and decreased immunostimulatory features, and a combination of pazopanib, topotecan, and MTD-based cyclophosphamide was associated with poor monocyte differentiation and decreased DC immunostimulatory parameters. Searching for a surrogate marker predicting an adverse outcome of DC manufacture in the peripheral blood complete blood count prior to monocyte harvest, we observed an association between an increased number of immature granulocytes in peripheral blood and decreased potency of the DC-based product as quantified by allo-MLR. We conclude that the DC-manufacturing yield and the immunostimulatory quality of anti-cancer DC-based vaccines generated from the monocytes of patients were not influenced by the monocyte isolation modality but were detrimentally affected by the specific combination of anti-cancer agents used prior to monocyte harvest.

Comparison of titer results obtained using immediate spin one-dilution techniques to a reference method.

BACKGROUND: Many transfusion services determine the titer of potentially incompatible plasma-containing products by performing a one-dilution titer at their selected titer threshold. This study compared the results of immediate spin (IS) one-dilution titers determined by three methods with a reference standard method. METHODS: Plasma-containing products from group A and O donors were titered using the participant's routine IS one-dilution titer method. No time or temperature incubations were performed, and antihuman globulin reagent was not used. The samples were then tested using a reference method, which was a saline tube test with a 1-hour room temperature incubation; antihuman globulin was not used in the reference method. The results of the one-dilution titer were then compared to that obtained in the reference method. RESULTS: Nine centers participated in this study. There were 698 antibodies from 374 units tested by the manual IS tube one-dilution titer method; sensitivity was 0.88 (95% confidence interval [CI], 0.83-0.92), and specificity was 1.00 (95% CI, 0.98-1.00). There were 412 antibodies from 206 units tested by the manual and automated IS buffered gel card one-dilution titer method; sensitivity was 0.95 (95% CI, 0.91-0.98), and specificity was 0.87 (95% CI, 0.81-0.91). There were 98 antibodies from 49 units tested by an automated microplate IS one-dilution titer method; sensitivity was 0.76 (95% CI, 0.71-0.93), and specificity was 0.96 (95% CI, 0.92-0.99). All three methods had an accuracy rate of 90% or greater. CONCLUSION: The manual and automated one-dilution titer methods are suitable for screening plasma-containing units, although more evaluation of the automated microplate method might be required.

Numerical defects in CD8+CD28- T-suppressor lymphocyte population in patients with type 1 diabetes mellitus and multiple sclerosis.

Type 1 diabetes mellitus (T1D) and multiple sclerosis (MS) are organ-specific autoimmune diseases leading to an attack of auto-aggressive lymphocytes against the pancreatic beta-cells and central nervous system, respectively. Using four-colour flow cytometry, T-lymphocyte populations having an important function in autoimmune processes were analyzed. T-regulatory cells (Treg) CD4(+)CD25(+)CD127(low), T-suppressor cells (Ts) CD8(+)CD28(-), activated helper CD4(+)CD25(+)CD127(+) and cytotoxic CD8(+)CD25(+) T-cells and also naive CD4(+)CD45RA(+) and memory T-cells CD4(+)CD45RO(+) were compared in the group of patients with T1D (n=30), MS (n=31) and in the group of healthy controls (n=29). Significant differences in Ts cells, activated helper and cytotoxic cells and also memory T-cells were recognized in the group of T1D patients compared to healthy controls. Ts population was significantly lowered in MS patients as well. However, no significant differences were noticed in Treg population. The observed data demonstrate significant differences among patients with T1D and MS in comparison to healthy individuals.