Effect of mirasol pathogen reduction technology system on immunomodulatory molecules of apheresis platelets.

Pathogen inactivation for platelets by riboflavin system (MIRASOL) efficiently reduces transfusion related pathogen transmission. However little is known about its impact on platelets' immunomodulatory biochemical profile. We aimed was to assess the effects of MIRASOL treatment on platelet quality parameters and immunomodulatory molecules CD62P, RANTES, and CD40L in Single Donor Platelets (SDPs) resuspended in plasma (SDP-P) or T-PAS and additive solution (SDP-A). Twenty nine SDPs (15 SDP-P and 14 SDP-A) were included in the study. Samples were collected before, after MIRASOL treatment and just before transfusion. P-selectin (CD62P), RANTES, and CD40L were tested by ELISA. Platelet products quality assays were also performed. Platelet count/unit decreased after Mirasol treatment by 13 %. The pH of all units decreased over the 5-day storage period but remained above expected limits and the swirling test was positive throughout storage. P-selectin levels were not different between the three different time points in both SDPs-P and SDPs-A while RANTES levels were found to differ statistically significantly at the three different time points in all units and in the SPD-A subgroup. CD40L levels in all SDP products increased slightly during storage but this was not statistically significant. CD62P, RANTES, and CD40L in all time points were elevated in SDPs-A compared to SDPs-P but not at a statistically significant level. In conclusion MIRASOL treatment apart from RANTES increase does not seem to substantially affect platelets associated other cytokines and immunomodulatory molecules namely P-selectin and sCD40L which are implicated in immune transfusion reactions.

HEMOSTASIS DISORDERS IN CORONARY ARTERY DISEASE: A PROSPECTIVE COMPARATIVE STUDY OF 130 PATIENTS.

The laboratory detection of factors that participate in coagulation mechanisms in patients with coronary heart disease may lead to important findings regarding the contribution of endothelial function to atherosclerotic lesions of coronary arteries. The main purpose of this study was to investigate the role of high-sensitivity C-reactive protein (hs-CRP), von Willebrand factor (vWF) activity, thrombomodulin (TM), ADAMTS13 activity and myeloperoxidase (MPO) in patients undergoing coronary angiography due to non-ST-elevation myocardial infarction (NSTEMI), unstable angina (UA) and stable angina pectoris with positive stress testing-induced myocardial ischemia (controls). Furthermore, the measured biomarkers were examined among patients with classical cardiovascular risk factors. 50 NSTEMI patients, 50 UA patients and 30 controls referred to coronary angiography were included in the study. The blood samples were collected before the catheter procedure. MPO, TM and ADAMTS13 activity were measured by enzyme-linked immunosorbent assay (ELISA), while vWF activity was calculated with INNOVANCE vWF Ac. When the laboratory results were compared between the three study groups, hs-CRP was found to be higher in NSTEMI patients compared to UA patients (p=0.0015) and controls (p<0.0001). ADAMTS13 activity was higher in NSTEMI (p=0.0035) and UA patients (p=0.0102) compared to controls and TM was lower in NSTEMI patients compared to UA patients (p=0.0307) and controls (p=0.0002). Moreover, MPO was higher in UA patients compared to the control group (p=0.0227). Finally, each of the aforementioned biomarkers was compared in the presence of the following cardiovascular risk factors: smoking, diabetes mellitus, arterial hypertension, dyslipidemia, chronic kidney disease (CKD) and peripheral artery disease (PAD). The results of this study add more data to the current medical literature concerning the role of coagulation disorders, endothelial damage and immunothrombosis in patients with coronary artery disease and their correlation with traditional risk factors for cardiovascular disease.

Real-world data from a single Greek centre on the use of secukinumab in plaque psoriasis: effectiveness, safety, drug survival, and identification of patients that sustain optimal response.

BACKGROUND: Few studies have investigated the long-term outcomes of secukinumab in real-life psoriasis treatment where diverse patient profiles require a personalized approach. OBJECTIVES: To determine long-term performance of secukinumab in moderate-to-severe plaque psoriasis, and identify potential clinical factors predictive of sustained optimal response under real-world conditions. METHODS: In this 78-week, single-centre, retrospective study, effectiveness, safety and drug survival of secukinumab were evaluated. Effectiveness data are reported as observed. Co-primary endpoints were absolute Psoriasis Area and Severity Index (PASI) ≤3 at week 4, 16, 52, 78, and clinical predictors of PASI ≤3 and PASI100 responses at week 52 and 78. RESULTS: A total of 85 patients (75.3% male; mean age 48.6 years) were included. Absolute PASI ≤3 was achieved in 73% and 83% of patients at week 52 and 78, respectively. PASI 75/90/100 responses at week 52 (71.6%, 50.8%, and 40.3%, respectively) were sustained at week 78 (73.6%, 64.2%, and 45.3%, respectively). Median absolute PASI remained low at week 52/78 (0.9/0.6, respectively), while mean absolute PGA also sustained low (0-1) values after 16-78 weeks. Investigator's Global Assessment 0/1 response rate was maintained by week 52/78 (72/83%, respectively). The drug survival rate of secukinumab at week 78 was 79.1%. Treatment was discontinued in 17.9% of patients after an average of 41.7 weeks, mainly due to loss of effectiveness (10.4%). A total of 27% experienced adverse events, without critical safety concerns. Based on multivariate analysis, advanced body mass index (BMI) and presence of ≥3 comorbidities decreased the chance of achieving PASI ≤3 at week 78 [OR (95% CI) 0.78 (0.64-0.97); P = 0.024, and OR (95% CI) 0.045 (0.002-0.83); P = 0.037, respectively]. CONCLUSIONS: Secukinumab showed consistently high effectiveness in this real-life cohort, with an acceptable safety profile. Over time, persistence of PASI ≤3 response appears to be lower in patients with high BMI or multiple comorbidities.

Correlation of single nucleotide polymorphisms in the promoter region of the ANXA5 (annexin A5) gene with recurrent miscarriages in women of Greek origin.

Background: Recent findings show that a number of single nucleotide polymorphisms (SNPs) within the promoter region of the annexin A5-gene (ANXA5) reduce the expression of the reporter gene and so they display a significant association with recurrent pregnancy loss (RPL).Objective: The objective of the present study aimed to address the contribution of ANXA5 M2 haplotype consisting of four minor alleles: (SNP1: (-)467G > A, SNP2: (-)448A > C, SNP3: (-)422T > C, and SNP4: (-)373G > A) in the occurrence of recurrent pregnancy losses in the Greek population, and the role of further two minor alleles: SNP5: (-)302 T > G and SNP6: (-)1C > T as independent risk factors for RPL.Methods: A 752-bp genomic region of ANXA5 promoter was amplified by PCR using specific primers. Genotypic analysis by Sanger sequencing was performed for these six SNPs (minor alleles) in the promoter region of ANXA5 gene, in 100 (100) Greek women with recurrent miscarriages (median =3) and 70 (70) fertile controls. Statistical analysis was done using the SAS 9.3 for Windows (SAS Institute Inc, NC, USA) and SPSS packages for Windows (C.DiMaggio 2013, SAS Institute 2014).Results: This case-control study revealed that there is no significantly increased risk of RPL among the M2/ANXA5 haplotype carriers in the Greek population, as there were no statistical differences between the patients with recurrent pregnancy losses and the fertile controls (11.5% in RPL cases versus 9.29% in controls, p-value: .6364). There was no difference in SNP5 and SNP6 minor carriership between the two groups. In particular, carriers of SNP5 and SNP6 had an increased risk for RPL state with odds ratio: 1.2472 and 1.3846 respectively, however without statistically significant importance.Conclusion: The M2/ANXA5 haplotype does not differ between RPL patients and controls in the Greek population. Also, it is the first time that SNP5 and SNP6 minor alleles were evaluated extensively in women of European origin with recurrent pregnancy losses (RPL), and they do not seem to be independent risk factors in the occurrence of RPL in the Greek population. Though, this has to be confirmed in further and larger clinical trials with women of European origin.

The role of prasugrel in the management of acute coronary syndromes: a systematic review.

OBJECTIVE: Dual antiplatelet therapy (DAPT) is the treatment of choice in the medical management of patients with acute coronary syndrome (ACS). The combination of aspirin and a P2Y12 inhibitor in patients who receive a coronary stent reduces the rate of stent thrombosis and the rates of major adverse cardiovascular events. However, patients with acute coronary syndrome remain at risk of recurrent cardiovascular events despite the advance of medical therapy. The limitations of clopidogrel with variable antiplatelet effects and delayed onset of action are well established and lead to the development of newer P2Y12 inhibitors. Prasugrel is a selective adenosine diphosphate (ADP) receptor antagonist indicated for use in patients with ACS. Prasugrel provides greater inhibition of platelet aggregation than clopidogrel and has a rapid onset of action. We have conducted a systematic review to retrieve current evidence regarding the role of prasugrel in the management of ACS. Evidence comparing prasugrel, clopidogrel, and ticagrelor remain scant. MATERIALS AND METHODS: A complete literature survey was performed using PubMed database search to gather available information regarding management of acute coronary syndromes and prasugrel. An explorative comparison of the safety and efficacy of prasugrel, clopidogrel, and ticagrelor was also conducted. RESULTS: Prasugrel and ticagrelor are more efficacious than clopidogrel in reducing the occurrence of non-fatal myocardial infarction, stroke, or cardiovascular (CV) death but they have also an increased risk of major bleeding in comparison to clopidogrel. CONCLUSIONS: Prasugrel and ticagrelor are today the recommended first-line agents in patients with ACS. The estimation of which drug is superior over the other cannot be reliably established from the current trials.

Alloimmunisation during pregnancy in Greece: need for nationwide HDFN prevention programme.

AIM/OBJECTIVES: To access the incidence and specificity of maternal red blood cells alloimmunisation and its relevant clinical impact in Greece. BACKGROUND: The rate of alloimmunisation in pregnant women in Greece is unknown. MATERIALS/METHODS: We performed a 4-year study in two tertiary hospitals in Greece. Demographics, transfusion and obstetric history were analysed. Maternal alloimmunisation was detected with indirect anti-globulin test. RESULTS: We investigated 4368 pregnant women. Of which 3292 (75·37%) were Greek and 1076 (24·63%) were migrants. In 39 alloimmunised women, 41 alloantibodies were detected (0·89%). The incidence of alloimmunisation was 0·66% (22/3292) in Greeks and 1·76% (17/1076) in migrants (P = 0·01). Anti-D was the most frequent alloantibody (0·18%). Anti-D was more frequent in migrants; 5·76% compared to 0·56% in Greek RhD negative women (P = 0·002). Other antibody specificities in declining frequency rank were anti-K, anti-E, anti-Lea, anti-M, anti-c, anti-Ce, anti-Jka, anti-Jkb and anti-C. Primiparae vs para >2 and past history of blood transfusion were significantly associated with alloimmunisation during pregnancy (P = 0·0088, P < 0·0001, respectively). CONCLUSIONS: Our results depict differences in the delivery of health care between migrants and Greek women, as well as the heterogeneity in practices for the prevention of haemolytic disease of foetus and newborn in Greece and highlight the need for the implementation of nationwide guidelines.

Effect of packed red blood cells transfusion on plasma fibronectin during liver resection.

Our study aimed at evaluating the effect of blood transfusion - allogeneic or autologous - on plasma levels of fibronectin during liver resections. Thirty-five patients scheduled for liver resection were randomly allocated to receive autologous (group autologous blood transfusion (ABT), n= 19) or allogeneic (homologous) (homologous blood transfusion (HBT), n= 16) packed red blood cell to maintain serum haemoglobin concentration above 9 g. Serum levels of fibronectin were measured before induction of anaesthesia, at the end of operation and at first, third and sixth postoperative day. Perioperative morbidity and survival rate were also recorded. Serum fibronectin levels were significantly higher (P < 0.05) in the autologous group than in the allogeneic, at the first (134 +/- 49 microg mL(-1) vs. 89 +/- 31 microg mL(-1)) and third (178 +/- 51 microg mL(-1) vs. 96 +/- 41 microg mL(-1)) postoperative day. No differences in survival and complication rate between the two groups were observed. Concentrations of serum fibronectin seem to be adversely affected by allogeneic blood transfusion during liver resection surgery, although this does not seem to affect patients' morbidity and mortality.

Predictive factors for survival in myeloma patients who undergo autologous stem cell transplantation: a single-centre experience in 211 patients.

High-dose therapy with autologous stem cell therapy (ASCT) has become the treatment of choice for eligible patients with myeloma. We analysed retrospectively the prognostic influence of pre-transplant characteristics and transplant modalities on response and survival in 211 myeloma patients who were transplanted in our centre between 1994 and 2004. All patients received peripheral blood stem cell support after conditioning with melphalan alone (183 patients), or melphalan and total blood irradiation (28 patients). We evaluated the influence of age, type of multiple myeloma, status prior and post ASCT, previous treatment regimens, time of ASCT from diagnosis, year of autograft, dose of re-infused CD34(+) cells, plasma cell infiltration and beta2-microglobulin at diagnosis on overall survival (OS) and event-free survival (EFS) to define patients with better prognosis. Median OS and EFS from transplantation were 50.9 and 20.1 months, respectively. Median OS from diagnosis was 68.8 months. Transplant-related mortality was 1.4%. Lower beta2-microglobulin levels, achievement of complete remission (CR) post transplant and lower plasma cell infiltration at diagnosis and transplant correlated with longer EFS and OS, whereas CR at transplant and low international prognostic index at transplant correlated with better EFS. Higher CD34(+) cell dose correlated with improved OS. We conclude that ASCT is safe and effective and the outcome is independent of age, time from diagnosis, previous treatment and conditioning regimen.

The combination of intermediate doses of thalidomide with dexamethasone is an effective treatment for patients with refractory/relapsed multiple myeloma and normalizes abnormal bone remodeling, through the reduction of sRANKL/osteoprotegerin ratio.

The aim of this study was the evaluation of the effect of intermediate doses of thalidomide with dexamethasone (Thal/Dex) on disease course and bone disease in patients with refractory/relapsed myeloma who were under zoledronic acid therapy. We studied 35 patients, who received thalidomide at a dose of 200 mg/daily. We measured, pre-, 3 and 6 months post-treatment soluble receptor activator of nuclear factor-kappaB ligand (sRANKL), osteoprotegerin (OPG), osteopontin (OPN), markers of bone resorption and formation. Before treatment, patients had increased levels of sRANKL/OPG ratio, bone resorption markers and OPN, while they had suppressed bone formation. The pretreatment sRANKL/OPG ratio correlated with the extent of bone disease. Thal/Dex administration resulted in a significant reduction of sRANKL/OPG ratio, and bone resorption. Bone formation, OPG and OPN did not show any alteration. Changes of sRANKL/OPG ratio correlated with changes of bone resorption markers. Thal/Dex was given for a median time of 10 months and the median follow-up period was 22 months. The response rate was 65.7%. The median survival was 19.5 months. beta2-microglobulin, type of response and International Staging System predicted for survival. These results suggest that the combination of intermediate dose of Thal/Dex is effective in patients with refractory/relapsed myeloma and improves abnormal bone remodeling through the reduction of sRANKL/OPG ratio.

Colorectal adenomas with a low frequency of ki-ras mutations: a sample from a population with a low mortality rate from colorectal cancer.

OBJECTIVE: To detect and analyse ki-ras point mutations in colorectal adenomas and carcinomas in patients from Greece, where mortality rate from colorectal cancer is lower than that in other European countries and the USA. PATIENTS AND METHODS: As a method we used PCR-RFLP technique, to examine ki-ras codons 12 and 13 mutations in 450 paraffin embedded tissues, consisting of 141 normal mucosas, 165 adenomas and 144 sporadic colorectal cancers taken from 141 patients. RESULTS: Frequency of ki-ras mutations was 3.2% (3/93) in small adenomas, 5.2% (3/57) in greater than 1 cm adenomas, 20% (3/15) in tissues where adenoma and cancer coexisted and 39.5% (57/144) in colorectal cancer. CONCLUSION: These data showed that the frequency of ki-ras mutations was low in all sizes of polyps and high in carcinomas in a sample of a population, characterized by a low mortality rate from colorectal cancer. Ki-ras mutations were not an early event of colon carcinogenesis in these patients and the low frequency of ki-ras mutations in adenomas may probably have some relevance to the low mortality rate. Our findings could be explained, not by the dietary habits of this population, but by the presence of other hitherto unknown environmental factors.

A comparative study of invasive and oscillometric methods of arterial blood pressure measurement in the anesthetized rabbit.

INTRODUCTION: The aim of this project was to evaluate the reliability and accuracy of direct, using the central ear artery (CEA), and oscillometric, using limb-cuffs, methods of arterial blood pressure (AP) measurement in the anesthetized rabbit. METHODS: New Zealand rabbits were anesthetized using a xylazine-ketamine-isoflurane protocol. Using the abdominal aorta (ABA) as direct "gold standard" for AP measurements, ABA pressure readings, via femoral artery catheterization, were compared with those made simultaneously from the ascending aorta after median sternotomy. Thereafter, direct CEA as well as forelimb-(FL) and hindlimb-(HL) cuff oscillometric readings were compared with those made simultaneously from ABA. RESULTS: The blood pressure in the ABA correlated with that from ascending aorta. Furthermore, CEA correlated with the ABA readings. Nevertheless, at high pressures, their divergence from "true" pressure tended to increase. Oscillometric readings at the FL site correlated well with "true" pressure while those at the HL site did not. Their divergence tended to increase at high pressures when using the FL site, while it varied when using the HL site. The accuracy of measurements was moderate for the FL site while poor for the HL site. DISCUSSION: Our results suggest that the CEA can be readily used with high reliability and accuracy for direct AP measurements in the anesthetized rabbit. On the other hand, the FL-cuff oscillometric method should only be used for the evaluation of AP at low and normal pressure ranges.

Autologous stem cell transplantation normalizes abnormal bone remodeling and sRANKL/osteoprotegerin ratio in patients with multiple myeloma.

The osteoprotegerin (OPG)/receptor activator of NF-kappa B ligand (RANKL) system has a major role in the pathogenesis of bone disease in myeloma (MM). The effect of autologous stem cell transplantation (ASCT) on bone turnover in MM was evaluated in 51 patients (35M/16F). Markers of bone resorption (NTX, TRACP-5b), bone formation (bone-alkaline phosphatase (bALP), osteocalcin), OPG and sRANKL were measured pre- and every month post-ASCT. The median follow-up period was 12 months. Four patients were transplanted in CR, 44 were transplanted in PR and three patients had progressive/resistant disease. All patients received bisphosphonates both pre- and post-ASCT. At baseline the majority of patients had increased NTX, TRACP-5b levels, and sRANKL/OPG ratio, while markers of bone formation were strongly suppressed. ASCT produced a significant reduction of sRANKL/OPG ratio, with a concomitant decrease of NTX, and TRACP-5b levels, starting the second month post-ASCT. Bone formation markers, osteocalcin and bALP, started to increase after the 9th and 11th month post-ASCT, respectively, while the increase of OPG preceded this. These results provide biochemical evidence that ASCT normalizes the abnormal bone resorption in MM patients possibly through the decrease of RANKL/OPG ratio, while bone formation requires a longer period to return to normal.

An unusual case of CD4+ CD7+ CD56+ acute leukemia with overlapping features of type 2 dendritic cell (DC2) and myeloid/NK cell precursor acute leukemia.

Type 2 dendritic cell (DC2) acute leukemia has been recently described. We report here an unusual case of a 17-yr-old adolescent with overlapping features of DC2 and myeloid/NK cell precursor acute leukemia as defined by Suzuki et al. The patient presented with lymphadenopathy and hepatosplenomegaly without extranodal manifestations in skin or elsewhere. The morphologic, cytochemical and immunophenotypic features were compatible with those described in DC2 acute leukemia, with co-expression of CD4, CD56 and CD123 antigens. The novel markers BDCA-4 and BDCA-2 considered specific for DC2s were co-expressed. However, bright CD7 positivity along with a dim expression of CD33 (57%) and CD117 (27%) were also noted. Additionally, there was bright expression of NG2 monoclonal antibody 7.1, a frequent finding in myeloid/NK cell precursor acute leukemia. The interpretation of the immunophenotypic profile leads to the hypothesis on the existence of borderline cases between DC2 and myeloid/NK cell precursor acute leukemia. Still, other hypotheses can not be overlooked, such as the possibility for a kind of variant monoblastic leukemia or of another rare entity of acute unclassified leukemia.

Genetic heterogeneity underlies juvenile hemochromatosis phenotype: analysis of three families of northern Greek origin.

Hereditary hemochromatosis is a genetically heterogeneous disease. Common HFE mutations (C282Y and H63D) are related to the majority of hereditary hemochromatosis cases in populations of Northern European ancestry (HFE1). Juvenile hemochromatosis (JH) is a more severe iron overload disorder, usually presenting at the second decade of life. The gene responsible for JH lies on a genetic locus at chromosome 1q. We have performed a genetic linkage study in three families of Northern Greek origin with typical clinical features of JH. In two families results were in accordance with linkage to chromosome 1q. In one family linkage of the disease to the genetic loci at 1q21, 7q22, and 6p22 was excluded. We suggest that more than one gene may underlie the JH phenotype. This genetic type of hemochromatosis may be designated 1q unlinked juvenile hemochromatosis. Family studies are necessary to establish the genetic diagnosis of JH.