Effect of leukoreduction on tumor-associated cytokine accumutation in supernatant of stored packed red cells and its effect on tumor cell proliferation in vitro / 中国实验血液学杂志

<p><b>OBJECTIVE</b>This study was to investigate whether prestorage leukoreduction could decrease the accumulative concentration of tumor-associated cytokines in supernatant of stored packed red blood cells (pRBC) and to study the effect of prestorage leukoreduction on proliferation of HepG2 tumor cells by in vitro. The leukoreduced (LR) and non-leukoreduced (NLR) pRBC were equally obtained from one donation and were stored under 2 °C-6°C. The supernatants of pRBC in these two group were performed by centrifugation with 1 006×g for 10 min at day 0 and 35 d. The enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of normal T cells and secretory factor (RANTES/CCL5), as well as the accumulative concentrations of tumor-necrosis factor (TNF-α), platelet derived growth factor (PDGF), vascular endothelial growth factor (VEGF) and monocyte chemotactic protein-1 (MCP-1) in pRBC supernantant of above-mentioned two groups. After HepG2 cells was cultured with the supernatant of NLR-pRBC and LR-pRBC at the end of day 35 together for 48 hours, the methyl thiazolil tetracolium (MTT) method was used to measure the proliferation of tumor cells in vitro.</p><p><b>RESULTS</b>The accumulative concentration of 5 cytokines in supernatants of above menthioned two groups increased in different degrees along with the prolongation of storage time,that is, the accumulative concentrations of 5 cytokines at 35 d were higher than that at day 0, in which the change of VEGF accumu-lative concentration showed statistical significance, its accumulative concentration in NLR group at day 35 elevated to 549.61 ± 299.43 pg/ml, and was higher than that in LR group (95.46 ± 110.87 pg/ml) (P < 0.05). The experiment of HepG2 cell proliferation indicated that the supernatant of LR pRBC group produced less proliferation of tumor cells with OD value 0.40 (95% CI, 0.38-0.42) than that of NLR pRBC group with OD value 0.49 (95% CI, 0.43-0.55) (P < 0.05).</p><p><b>CONCLUSION</b>The prestorage leukoreduction has been confirmed to decrease the accumulative level of cytokines, particalarly decrease the accumulative level of VEGF, moreover, it may be a factor for inhibiting the proliferation of tumor cells in vitro.</p>

Effect of Storage Time on Cumulation of Platelet-related Growth Factors in the Supernatant of Leukoreduced Packed Red Cells and Tumor Cell Proliferation In Vitro / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of storage time on accumulation of platelet-related growth factors in the supernatant of leukoreduced packed red blood cells (LR-pRBC) and on tumor cell proliferation in vitro.</p><p><b>METHODS</b>LR-pRBC were quartered and stored at 2 °C-6 °C. The supernatant of pRBC was obtained by centrifugation with 1 006 × g for 10 min at day 0, 14, 21 and 35 d. The enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of platelet-derived growth factor(PDGF) and vascular endothelial growth factor (VEGF). After HepG2 cells was cultured with the supernatant of LR pRBC at day 0 and day 35 together for 48 hours, methylthiazoliltetracolium (MTT) method was used to measure the proliferation of tumor cells in vitro.</p><p><b>RESULTS</b>The concentrations of 2 cytokines were still increased with the storage time prolonging. As compared to LR-pRBC at day 0 [611.84 (95%CI, 356.45-867.23) pg/ml], the level of VEGF reached 900.16 (95% CI, 552.26-1248.07) pg/ml (P<0.05). There was a similar tendency in PDGF level with less increment in the supernatant of LR pRBC at day 35 [2.23 (95% CI, 0-5.37) ng/L vs 5.66 (95% CI, 0-12.48), P=0.073], but there was no significant statistical difference. Likewise, in vitro study of HepG2 cell proliferation showed that the LR-pRBC at day 35 promoted more proliferation of tumor cells with OD value 0.40 (95% CI, 0.38-0.42) (P<0.05).</p><p><b>CONCLUSION</b>The residual platelets in LR-pRBC were activated, disintegrated and released the dense granules and α-granules which induce the accumulation of VEGF and PDGF. It seemed that the supernatant of LR-pRBC promoted the proliferation of tumor cells in vitro.</p>

Analysis of Factors Influencing Posttransfusion Effectiveness in 26045 Cases of Platelet Transfusion / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the factors influencing platelet transfusion results so as to improve the platelet transfusion efficiency.</p><p><b>METHODS</b>According to the clinical symptoms (bleeding condition is stopped or improved)and the corrected count of increment (CCI), the patients were divided into efficient transfusion and inefficient transfusion groups. A total of 20 671 patients' clinical data and main platelet transfusion parameters in 26 045 tranfusions including platelet count of per- and post- transfusion, platelet component sorts, storage time and transfusion number were analysed.</p><p><b>RESULTS</b>The comparison of platelet transfusion efficiency in age and sex between two groups did not showed statistical difference (P > 0.05), the platelet count before transfusion between two groups showed statistical difference (t = -5.59, P < 0.001) after converting to log, a significant linear correlation did not exist between storage time of the platelet and CCI (corrected count of increment), but there was statistical difference in transfusion efficiency of patients with different diseases. The patients with hematologic diseases showed lower efficiency of platelet transfusion. According to the results of Wilcocon test detection, there was difference between different times of transfusion and transfusion efficiency, that is to say, the transfusion frequency was higher, the transfusion efficency was lower. The Fisher test indicated that the transfusion efficiency of single platelet transfusion was lower than that of transfussed platelet with other blood components (P < 0.01).</p><p><b>CONCLUSION</b>Platelet transfusion efficiency associates with many factors, including different diseases, whether being transfused with other blood components, the platelet count before transfusion, transfusion frequency, but the time of storage does not relate to the transfusion efficacy.</p>

Assessment of RBC transfusion volume and its effect on postoperative pulmonary complications in on-pump CABG patients / 中国实验血液学杂志

This study was purposed to investigate the effect of the transfused RBC amount on pulmonary complications after on-pump CABG surgery, and to explore the influencing factors on RBC transfusion volume. 292 adult patients receiving on-pump CABG surgery were divided into non-RBC transfusion group (n = 71), 1-4 U RBC transfusion group (n = 144) and >4 U RBC transfusion group (n = 77). Adjusted multivariable regression analysis was performed to examine the correlation between transfused RBC amount and the odds of pulmonary complications, and multivariable linear regression was used to analyze the influencing factors on RBC transfusion volume. The results showed that compared the three groups, there was the significant difference in postoperative pulmonary complications (1.4% vs 14.6% vs 24.7%, P < 0.001). A stronger and graded correlation was found between transfused RBC amount and pulmonary complications in on-pump CABG patients, the adjusted odds were increased to 1.251 (95% CI: 1.120-1.398, P < 0.001), and influencing factors on RBC transfusion volume were as follows: age (B:0.102; 95% CI: 0.046-0.157, P < 0.001), sex (B:1.825; 95% CI: 0.692-2.957, P = 0.002), preoperative Hct (B:-36.044; 95% CI:-47.724--25.163, P < 0.001), CPB time (B: 0.031; 95% CI:0.013-0.050, P = 0.001) and acute myocardiac infarction (B:2.769; 95% CI: 1.295-4.243, P < 0.001). It is concluded that the transfused RBC amount is related with postoperative pulmonary complications, and the influencing factors on RBC transfusion volume include preoperative Hct, age, acute myocardiac infarction, sex and CPB time.

Accumulative regularity of tumor-associated noncellular components in supernatant of stored packed red cells / 中国实验血液学杂志

This study was purposed to investigate the accumulative regularity of tumor-associated noncellular components in supernatant of stored packed red blood cells (PRBC) during storage. The supernatant of PRBC was obtained by centrifugation with 1 006×g for 10 min at day 0, 7, 14, 21, 28 and 35 d. The enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of T cells and the accumulative levels of secreted RANTES/CCL5, tumor necrosis factor-alpha (TNF-α), platelet derived growth factor (PDGF), vascular endothelial growth factor (VEGF) and monocyte chemotactic protein-1 (MCP-1). The results showed that the high concentration of RANTES/CCL5 and TNF-α was found in fresh PRBC, and their accumulative concentration did not increase along with the prolonging of storage time. The VEGF level in PRBC at day 0 of storage was 229.9 pg/ml, and increased to 749.08 pg/ml at end of day 7, then it was stable, and increased to 760.67 pg/ml at end of day 35. The PDGF level in supernatant of PRBC was 13.54 ng/L at dag 0 of storage, and increased stably during storage, then decreased at day 28, however PDGF rapidly rose to 22.13 ng/L at the end of day 35 (P < 0.05). The MCP-1 level in supernatant of PRBC was 39.98 ng/L at day 0 of storage, then slowly increased during storage time, at end of day 35 MCP-1 level increased to 49.83 ng/L. It is concluded that along with the prolongation of storage time, the growth factors in the supernatant of PRBC display the tendency of accumulative increment and RANTES/CCL5 and TNF-α show relative high level at day 0 of storage, moreover, no obvious increase of accumulation is observed along with prolonging of the storage time, suggesting no relation of concentration with storage time.

Effects of perioperative blood transfusion on the severity of postoperative infection / 中国实验血液学杂志

This study was purposed to explore whether the blood transfusion of surgical patients can increase the severity of postoperative infection by a retrospective analysis of patients with postoperative infection in Chinese PLA General Hospital. By using a software "clinical transfusion database" developed by our department, 150 infected surgical cases were retrieved and divided into deep infection group and superficial infection group according to the infected location. These two groups were compared in term of the patient's age, duration of hospitalization, red blood cell transfusion volume, none-red cell transfusion volume, transfusion frequency and average transfusion volume. The results showed that red blood cell transfusion volume or none-red cells transfusion volume of patients with superficial infection was 4.50 (0 - 59) U or 2.95 (0 - 119.6) U, and that of deep infection was 9.00 (0 - 153) U and 8.05 (0 - 136.6) U, the differences was significant (P < 0.05). Between two groups, the transfusion frequency showed the most significant difference, median in the patients with superficial infection was about 2 (1 - 31) times, less than the deep infection group about 4 (1 - 49) times (P < 0.001). There was no significant difference between two groups in the average transfusion volume. It is concluded that perioperative blood transfusion volume and frequency of surgical patients seems to display a positive correlation with the degree of postoperative infection.

Serological characteristics and transfusion efficacy evaluation in 61 cases of autoimmune hemolytic anemia / 中国实验血液学杂志

This study was aimed to analyze the serological characteristics, efficacy and safety of incompatible RBC transfusion in patients with autoimmune hemolytic anemia (AIHA). The patients with idiopathic or secondary AIHA were analyzed retrospectively, then the serological characteristics and the incidence of adverse transfusion reactions were investigated, and the efficacy and safety of incompatible RBC transfusion were evaluated according to the different autoantibody type and infused different RBC components. The results showed that out of 61 cases of AIHA, 21 cases were idiopathic, and 40 cases were secondary. 8 cases (13.1%) had IgM cold autoantibody, 50 cases (82.0%) had IgG warm autoantibody, and 3 cases (4.9%) had IgM and IgG autoantibodies simultaneously. There were 18 cases (29.5%) combined with alloantibodies. After the exclusion of alloantibodies interference, 113 incompatible RBC transfusions were performed for 36 patients with AIHA, total efficiency rate, total partial efficiency rate and total inefficiency rate were 56.6%, 15.1% and 28.3%, respectively. Incompatible RBC transfusions were divided into non-washed RBC group and washed RBC group. The efficiency rate, partial efficiency rate and inefficiency rate in non-washed RBC group were 57.6%, 13.0% and 29.4%, respectively. The efficiency rate, partial efficiency rate and inefficiency rate in washed RBC group were 53.6%, 21.4% and 25.0%, respectively. There was no significant difference of transfusion efficacy (P > 0.05) in two groups. Incompatible RBC transfusions were also divided into IgM cold autoantibody group and IgG warm autoantibody group. The efficiency rate, partial efficiency rate and inefficiency rate in IgM cold autoantibody group were 46.2%, 30.8% and 29.4%, respectively. The efficiency rate, partial efficiency rate and inefficiency rate in IgG warm autoantibody group were 56.7%, 13.4% and 29.9%, respectively. There was no significant difference of transfusion efficacy (P > 0.05 ) in two groups. Hemolytic transfusion reaction was not observed in all incompatible RBC transfusions. It is concluded that the same ABO type of non-washed RBC transfusion and O type washed RBC transfusion are all relatively safe for the AIHA patients with severe anemia after the exclusion of alloantibodies interference. There is no significant difference of transfusion efficacy in two groups. The same ABO type of non-washed RBC transfusion is more convenient and efficient than washed RBC transfusion, and excessive use of type O RBCs can also be avoided.

Inside quality control for whole blood preservation performed at blood transfusion compatibility testing laboratory / 中国实验血液学杂志

This study was aimed to establish the technique for preparation and storage of internal quality control pro-ducts by using existing blood sample resources of blood transfusion compatibility testing laboratory. 24 healthy blood donors with group A and RhD-positive were randomly selected, and 4 ml venous blood from these donors were collected, respectively. Based on the use of anticoagulant type, whether to add red blood cell preservation solution and the samples stored at room temperature for 1 or 2 hours daily, 24 specimens were randomly divided into 8 groups by using factorial design methodology. All samples in tube with cap were stored at 4 degrees C, and placed at room temperature for 1 or 2 hours daily. ABO, RhD blood group (recorded on the agglutination strength of the forward and reverse typing), IgM anti-B antibody titer, and free hemoglobin concentration in the supernatant for all samples were detected at 0, 7, 14, 21, 28, 35 days of products preservation. The results indicated that the red blood cell damage from the group used anticoagulants ACD-B and added the MAP red blood cell preservation solution and placed at room temperature 1 hour daily (recorded as A2B2C1 group) was kept minimal, and FHb concentration and FHb increments at each time point were the lowest (p < 0.01), the FHb concentration on 35th day was only (24.5 +/- 84.5) mg/L. There was no significant change of A antigen, D antigen and IgM anti-B antibody response activity and stability in A2B2C1 group during storage for 35 days (p > 0.05). In conclusion, blood transfusion compatibility testing laboratory can use A2B2C1 program established by this study to prepare relatively stable modified whole blood internal quality control products in the existing conditions, which can be effectively preserved and meet the requirements of internal quality control for blood transfusion compatibility testing.

The effect of leukocyte depletion by filtration on the quality of apheresis platelets / 中国实验血液学杂志

This study was aimed to investigate the effect of leukocyte depletion by filtration on the quality of apheresis platelets. 20 units of donor apheresis platelets were randomly selected and were preserved with agitation at 20 - 24 degrees C for 24 - 96 hours, then were filtered on polyester flatbed filters. The platelet concentration, mean platelet volume (MPV), volume of apheresis platelets, leukocyte count, pH value, lactate dehydrogenase (LDH) concentration, K(+) concentration and CD62p expression level on surface of platelet membrane, were detected before and after filtration, as well as the rate of leukocyte depletion and platelet loss were calculated. The results showed that the leukocyte count after filtration was remarkably lower than that before filtration (p < 0.001), and the rate of leukocyte depletion was 99.97%. Platelet loss was approximately 8%, and obviously lower than that of the national standard (p < 0.001). MPV, pH value, K(+) and LDH concentration were not significantly different before and after filtration. Compared with platelets before filtration, CD62p expression level after filtration slightly decreased (p > 0.05). CD62p expression on surface of platelet membrane in perfusion fluid obtained from filter plate was obviously higher than that before filtration (p < 0.05). MA of platelet after filtration slightly decreased (p > 0.05). It is concluded that leukocyte and partial activated platelets can be removed efficiently by using polyester flatbed filters, and platelet loss is very low. Filtration does not adversely affect coagulation activity of the platelets in vitro. Apheresis platelets after filtration can fulfil quality requirements to prevent infection of cytomegalovirus and HLA alloimmunization.

Establishment of genotyping method for fetal ABO group from pregnant maternal peripheral blood / 中国实验血液学杂志

This study was aimed to establish a genotyping method to detect ABO group gene of fetus from peripheral blood of pregnant women for prenatal diagnosis of hemolytic disease of newborn (HDN) resulting from ABO blood group incompatibility. 4 pairs of primers were designed according to ABO blood group gene DNA and mRNA sequences. 20 plasma DNA samples from healthy donors were extracted and amplified to explore the best conditions for plasma DNA extraction and PCR amplification. The O group plasma DNA was mixed with A group or B group plasmas by the ratios of 1:1, 2:1, 4:1, 8:1, 10:1, 20:1, 40:1, 100:1 to simulate the status of mixed ABO gene from pregnant maternal blood and to establish the mixed blood group ABO genotyping technology. The pregnant maternal blood samples with more than 30 weeks of gestation were selected for detecting the fetal ABO blood group genotype. The blood samples should be taken as possible as after birth for identification of ABO blood group and evaluation of sensitivity and accuracy of fetal ABO blood group genotyping technology through peripheral blood of pregnant women. The results indicated that the minimal amount of template DNA from single blood plasma for accuracy identification was at least about 0.625 ng, the DNA amount extracted from 500 microl of plasma could meet the requirement for PCR amplification. When the proportion of O group plasma DNA in mixed plasma DNA was <or=10, the non-O group gene could be accurately detected. Among 14 peripheral blood samples from O-group pregnant women, the non-O group gene was amplified in 9 samples; the non-O group gene was not amplified in 5 samples. The identification of peripheral blood ABO group for 5 newborns using serologic method showed that the A group 3 cases, B group 2 cases, O group 1 case, which consisted with genotyping results with consistent rate 100%. It is concluded that in middle and late pregnancy the fetal ABO group gene can be detected accurately by means of established fetal ABO group gene extraction and typing technology, that provides some guidances for the prenatal diagnosis and prevention of HDN.

Application of thrombelastography in evaluation of platelet function during storage / 中国实验血液学杂志

This study was aimed to explore changes of platelet function in vitro during storage by thrombelastography (TEG). 12 units plateletpheresis were randomly selected and stored at 20 to 24 degrees C with agitation. Thrombelastography variable parameters R, K values and maximal amplitude (MA) were measured on 1, 2, 3, 4, 5 days of platelet storage. Platelet concentration, mean platelet volume (MPV), hypotonic shock response (HSR), CD62p expression and CD62p reexpression on platelet surface were detected at the same time. Changes of platelet function in virto were systematically evaluated by above-mentioned indexes. The results showed that MPV augmented slightly with prolongation of preserved time (p > 0.05), and CD62p expression on platelet surface increased remarkably (p < 0.01), while CD62p reexpression decreased gradually (p < 0.01). There were no significant differences in HSR level of platelets during storage (p > 0.05). R value increased with prolongation of preserved time (p < 0.01). There were no obvious changes on K value and alpha Angle during storage (p > 0.05). There were no obvious changes in MA from 1 to 4 days, and MA decreased slightly on day 5 (p < 0.05). It is concluded that there was no significant change in MA and HSR which reflects comprehensive coagulation of platelets during storage. Platelets on the end of storage have excellent function of hemostasis; Thrombelastography parameter MA value can be used as a valuable indicator for evaluation of platelet function in vitro during storage.

Qualitative analysis of batch preparing cryopreserved fresh platelet rich plasma / 中国实验血液学杂志

To evaluate the efficiency and effectiveness of batch preparing cryopreserved fresh platelet-rich plasma (cryo-FPRP) derived from the volunteer donors, platelet count (Plt), mean platelet volume (MPV), platelet distribution width (PDW), plasma pH, plasma lactic acid concentration, and lactic dehydrogenase (LDH) concentration, germiculture, CD62p positive rate, PAC-1 positive rate, and the fluorescence intensity of platelet GPIb-IX-V were detected in ACD whole blood, fresh platelet-rich plasma (FPRP), FPRP with 5% dimethyl sulphoxide DMSO (DMSO-FPRP), and thawed cryopreserved FPRP (cryo-FPRP); the procoagulant activity of FPRP and cryo-FPR was determinated. The results showed that (1) 70 percentage of platelet were separated from the whole blood into FPRP, and the counts of residual erythrocyte and leucocyte were below 1 x 10(9), and below 1 x 10(7) per unit respectively. (2) The plasma pH, lactic acid concentration and PAC-1 positive rate retained a stable level during the preparing, storing and thawing process. (3) Plasma LDH concentration, platelet CD62p positive rate and GPIb-IX-V concentration in platelet surface were enhanced significantly after being frozen and thawing. (4) The plasma clotting time induced by cryo-FPRP were significantly shorter than that induced by FPRP. It is concluded that: (1) The batch platelet preparing process can efficiently obtain platelet from whole blood donated by volunteer, and the process didn't activate the platelet. (2) Cryopreservation can prevent lactic acid accumulation, pH reduce and activation of GPIIb/IIIa. (3) The membrane of partial platelets are affected by freezing and thawing. (4) The density of GPIb-IX-V complexes in platelet surface and its procoagulant activity are enhanced significantly after the FPRP freezing and thawing process.