scholarly journals Platelet storage at 22 degrees C: role of gas transport across plastic containers in maintenance of viability

Blood ◽  
1975 ◽  
Vol 46 (2) ◽  
pp. 209-218 ◽  
Author(s):  
S Murphy ◽  
FH Gardner
Keyword(s):  
Platelet Count ◽  
Lactate Concentration ◽  
Platelet Concentrates ◽  
Pasteur Effect ◽  
Platelet Storage ◽  
Low Levels ◽  
Microscopic Morphology ◽  
Plastic Containers

Containers constructed of polyvinylchloride (PVC) are used for the storage of platelet concentrates (PC) for transfusion, At 22 degrees C, pH often falls to such low levels (pH is less that 6.0) that viability is lost. Far lesser degrees of pH fall are observed in bags constructed of polyethylene (PE). In this study, pH, PO2, PCO2, platelet count, lactate concentration, microscopic morphology, and viability after 51- chromium labeling were evaluated during storage at 22 degrees C under a variety of circumstances. The results indicate that (1) pH falls because of the generation of lactic acid by platelet glycolysis and, under some circumstances, the retention of CO2. (2) Rate of pH fall is, therefore, roughly proportional to the platelet count. (3) PE is more permeable to gases, thereby allowing CO2 escape from and easier O2 entry into the stored PC; the higher O2 tensions suppress glycolysis by the Pasteur effect. (4) Adequate agitation and container size are critical if the beneficial effect of PE is to be obtained. (5) In general, platelets stored in PE containers have excellent viability in vivo although CO2 escape can result in elevations in pH which are deleterious. (6) Storage in a 10% CO2 atmosphere prevents these deletrrious pH elevations without otherwise impairing platelet viability; (7) Results similar to those achieved with PE can be achieved with PVC if this material is made thinner to allow easier penetration of gases.

scholarly journals Platelet storage at 22 degrees C: role of gas transport across plastic containers in maintenance of viability

Blood ◽  
1975 ◽  
Vol 46 (2) ◽  
pp. 209-218 ◽  
Author(s):  
S Murphy ◽  
FH Gardner
Keyword(s):  
Platelet Count ◽  
Lactate Concentration ◽  
Platelet Concentrates ◽  
Pasteur Effect ◽  
Platelet Storage ◽  
Low Levels ◽  
Microscopic Morphology ◽  
Plastic Containers

Abstract Containers constructed of polyvinylchloride (PVC) are used for the storage of platelet concentrates (PC) for transfusion, At 22 degrees C, pH often falls to such low levels (pH is less that 6.0) that viability is lost. Far lesser degrees of pH fall are observed in bags constructed of polyethylene (PE). In this study, pH, PO2, PCO2, platelet count, lactate concentration, microscopic morphology, and viability after 51- chromium labeling were evaluated during storage at 22 degrees C under a variety of circumstances. The results indicate that (1) pH falls because of the generation of lactic acid by platelet glycolysis and, under some circumstances, the retention of CO2. (2) Rate of pH fall is, therefore, roughly proportional to the platelet count. (3) PE is more permeable to gases, thereby allowing CO2 escape from and easier O2 entry into the stored PC; the higher O2 tensions suppress glycolysis by the Pasteur effect. (4) Adequate agitation and container size are critical if the beneficial effect of PE is to be obtained. (5) In general, platelets stored in PE containers have excellent viability in vivo although CO2 escape can result in elevations in pH which are deleterious. (6) Storage in a 10% CO2 atmosphere prevents these deletrrious pH elevations without otherwise impairing platelet viability; (7) Results similar to those achieved with PE can be achieved with PVC if this material is made thinner to allow easier penetration of gases.

Impaired CD163-mediated hemoglobin-scavenging and severe toxic symptoms in patients treated with gemtuzumab ozogamicin

Blood ◽  
2008 ◽  
Vol 112 (4) ◽  
pp. 1510-1514 ◽  
Author(s):  
Maciej Bogdan Maniecki ◽  
Henrik Hasle ◽  
Lennart Friis-Hansen ◽  
Birgitte Lausen ◽  
Ove Juul Nielsen ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
High Plasma ◽  
Gemtuzumab Ozogamicin ◽  
Intravascular Hemolysis ◽  
Soluble Cd163 ◽  
Plasma Hemoglobin ◽  
Monocytes And Macrophages ◽  
Low Levels

Abstract We describe a novel syndrome of severe toxic symptoms during intravascular hemolysis due to impaired hemoglobin scavenging in 2 children with acute myeloid leukemia undergoing CD33-directed therapy with the immunotoxin gemtuzumab ozogamicin (GO). A simultaneous high plasma hemoglobin, haptoglobin, and low bilirubin after septicemia-induced intravascular hemolysis indicated abrogated clearance of haptoglobin-hemoglobin complexes. This was further supported by low levels of plasma soluble CD163 and a concordant low number of CD163-expressing monocytes. We show that CD163 positive monocytes and macrophages from liver, spleen, and bone marrow coexpress CD33, thus suggesting that the GO-induced cellular cytotoxicity of CD33 positive cells eradicates a significant part of the CD163 positive monocytes and macrophages. The risk of severe toxic symptoms from plasma hemoglobin should be considered after CD33-targeted chemotherapy when the disease is complicated by a pathologic intravascular hemolysis. Furthermore, the cases provide further circumstantial evidence of a key role of (CD163-expressing) monocytes/macrophages in plasma hemoglobin clearance in vivo.

Leucocyte-Reduced Buffy Coat Platelet Concentrates (BCPC) Treated with INTERCEPT™ Stored up to 7 Days: Hemostatic Function in a Whole Blood Flow System.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 956-956
Author(s):  
Miguel Lozano ◽  
Ana Galan ◽  
Roberto Mazzara ◽  
Laurence Corash ◽  
Gines Escolar
Keyword(s):  
Blood Flow ◽  
Platelet Count ◽  
Whole Blood ◽  
Flow System ◽  
Ex Vivo ◽  
Buffy Coat ◽  
Platelet Concentrates ◽  
Platelet Storage ◽  
Pre Treatment ◽  
Additive Solution

Abstract Background: The risk of bacterial growth has limited the shelf life of platelet concentrates (PC) to 5 days. Modern platelet storage containers facilitate storage for up to 7 days, if bacterial contamination is prevented. INTERCEPT (Baxter, La Chatre, France; Cerus, Concord, CA) photochemical treatment (PCT) for pathogen reduction based on amotosalen (150μM) and UVA illumination (3 J/cm2) inactivates high titers of bacteria in PC (Transfusion2004; 44: 1496–1504). Adhesion and aggregation of platelets to injured vascular surfaces are critical aspects of platelet hemostatic function. In this study, the adhesion and aggregation of leucocyte-reduced buffy coat derived PC (BCPC), treated with INTERCEPT and stored up to 7 days, were measured on injured vascular surfaces using an ex-vivo blood flow system. Methods: BCPC were prepared from 450 mL-whole blood donations with the top and bottom method (Optipress II, Baxter). Five BCPC, of the same ABO group, were pooled with additive solution (Intersol™) the day following collection, after viral screening testing was completed. Following centrifugation and leukocyte depletion, two BCPC pools of the same ABO group were mixed and divided. One pooled BCPC was treated with INTERCEPT (I-BCPC) and the other was prepared by conventional methods (C-BCPC); and both were stored in 1.3 liter PL2410 plastic containers (Baxter R4R7012) at 22 ± 2°C with continuous agitation for 7 days. Samples for hemostatic function testing were taken immediately after preparation before splitting for treatment and after 5 and 7 days of storage. Platelet counts were performed in K3EDTA in a Coulter MD II counter (Coulter, Miami, FL). Samples of I-BCPC and C-BCPC were added to citrate anticoagulated blood, previously depleted of platelets and leukocytes by filtration, and adjusted to a final platelet count of 150x109/L. Enzymatically denuded vascular segments were exposed to circulating whole blood, reconstituted with I-BCPC and C-BCPC, in Baumgartner chambers at a shear rate of 800 s−1 for 10 minutes. The proportion (%) of the vascular surface area covered by platelets after perfusion was measured for each type of BCPC (N = 9) at each storage time point. Platelets and groups of platelets were classified as adhesive when platelet masses were less than 5 μm in height and as thrombi when height exceeded 5 μm. Data were analyzed with the SPSS 12.0.1 statistical package with significance at p < 0.05, and expressed at the mean ± SEM Results(Table). Conclusion: The platelet count of I- BCPC decreased by 12.3% including PCT processing losses and 7 days of storage. However, I- BCPC platelet adhesive and aggregatory capacities under flow conditions were similar to C- BCPC, and were well conserved for up to 7 days of storage. Hemostatic Function of Stored I-BCPC and C-BCPC Parameter I-BCPC C-BCPC p Day 1(Pre Treatment) Platelet Count (109/L) 945±40 945±40 Platelet Coverage (%) 26.0±3.7 26.0±4.2 Adhesion(%) 24.0±3.7 24.0±3.7 Thrombus(%) 1.9±0.6 1.9±0.6 Day 5 Storage Platelet Count (109/L 844±41 902±44 0.004 Platelet Coverage (%) 20.9±2.2 20.6±1.6 0.9 Adhesion(%) 19.9±2.1 19.3±1.4 0.8 Thrombus(%) 0.9±0.3 1.2±0.4 0.5 Day 7 Storage Platelet Count (109/L) 829±32 923±48 0.008 Platelet Coverage (%) 27.1±2.9 21.2±2.8 0.06 Adhesion(%) 26.0±2.7 20.4±2.7 0.06 Thrombus(%) 1.2±0.3 0.7±0.2 0.16

In Vivo Evaluation of Extended Stored Platelet Concentrates.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 946-946 ◽  
Author(s):  
Sherrill J. Slichter ◽  
Doug Bolgiano ◽  
Jill Corson ◽  
Mary Kay Jones ◽  
Todd Christoffel ◽  
...  
Keyword(s):  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Storage Conditions ◽  
Mean Value ◽  
Platelet Concentrates ◽  
In Vivo Evaluation ◽  
Platelet Storage ◽  
Volunteer Donor ◽  
Simple Ratio

Abstract Background: With the introduction of bacterial testing, extended storage of platelets is now possible as long as platelet viability is maintained. Although apheresis platelets have recently been licensed for seven days of storage, platelet concentrates are still only licensed for five days. We performed studies to determine the post-storage viability of platelets stored as concentrates. As yet, there are no established criteria for platelet viability that must be met at the end of storage. However, two different criteria for evaluating post-storage platelet viability have been suggested: compare the stored platelet results with each donor’s fresh platelet results; or establish a fixed platelet standard with an expected mean value along with the lower limit of the 95% confidence interval (lower 95% CI) of the mean. Methods: Forty-four normal volunteers donated a unit of whole blood, the whole blood was centrifuged to give platelet-rich plasma (PRP), and the PRP was then centrifuged to produce a platelet concentrate. The platelet concentrates were re-suspended and stored in either 100% plasma or in 20% plasma and 80% Plasmalyte (a platelet storage solution) for five to eight days in Terumo Teruflex bags. At the end of storage, a sample of blood was obtained from each volunteer to prepare fresh platelets to compare the results of a fresh platelet transfusion with the transfusion of the same volunteer’s stored platelets. The fresh and stored platelets were alternately radiolabeled with either 51Cr or 111In, re-injected into their volunteer donor, and serial blood samples were drawn from the volunteer after transfusion of the fresh and stored platelets to determine platelet recoveries and survivals. Results: Results are shown in Table 1. Conclusions: After seven days of storage in plasma, the platelets could qualify for standards of: 74% and 53% as a simple ratio of fresh to stored platelet recoveries and survivals, respectively; according to Dumont’s method of analysis (Transfusion, in press), achievable stored platelet recoveries would be 63% of fresh and survivals would be 40% of fresh; and as a fixed standard, platelet recoveries averaged 45% and survivals 4.4 days with lower 95% CI’s of 35% and 3.3 days, respectively. After eight days of storage in plasma, platelet recoveries and survivals gave unacceptable results. Instead of increasing or at least maintaining post-storage platelet viability, Plasmalyte markedly reduced platelet viability compared to plasma stored platelets at each storage interval. In Vivo Radiolabeled Autologous Platelet Recoveries and Survivals STORAGE CONDITIONS PLATELET RECOVERIES (%) PLATELET SURVIVALS (Days) Solution Time (Days) N Fresh Stored Fresh Stored Data are given as the average ±1 S.D. Plasma 5 10 62±12 55±11 7.7±1.5 6.1±1.0 Plasma 6 10 61±7 46±12 8.2±1.5 5.1±1.6 Plasmalyte 6 3 47±30 29±16 5.1±3.4 1.8±0.2 Plasma 7 10 61±11 45±14 8.3±1.0 4.4±1.5 Plasmalyte 7 5 62±17 30±12 7.4±0.6 2.2±1.1 Plasma 8 3 67±12 31±8 8.3±0.6 2.9±0.8 Plasmalyte 8 3 70±3 21±8 8.2±0.8 0.7±0.1

scholarly journals Platelet storage at 22 degrees C: effect of type of agitation on morphology, viability, and function in vitro

Blood ◽  
1978 ◽  
Vol 52 (2) ◽  
pp. 425-435 ◽  
Author(s):  
S Holme ◽  
K Vaidja ◽  
S Murphy
Keyword(s):  
Platelet Count ◽  
Shape Change ◽  
Normal Platelet ◽  
Platelet Storage ◽  
Platelet Morphology ◽  
And Function ◽  
Change Response ◽  
Citrate Phosphate

Abstract Recovery in vivo after 51Cr labeling, platelet morphology, and platelet aggregation were studied with platelet concentrates (PC) stored for transfusion under carefully controlled conditions. PC were prepared to a final volume of 50 ml from whole blood anticoagulated with citrate- phosphate-dextrose (CPD). The platelet count was kept between 0.8 and 1.6 X 10(12) platelets/liter. The PC were stored in bags constructed of polyvinylchloride (PVC) or polyethylene (PE) at 22 degrees C for 72 hr. The bags were placed on a horizontal shaker or a ferris wheel for agitation during storage. No significant changes in pH or platelet count were observed during storage. PC stored on the wheel showed moderate loss of viability and a marked deterioration of platelet morphology and aggregation compared to the shaker. PC stored on the shaker in bags made of PE showed better aggregation with ADP and thrombin but had the same viability and morphology as PC in bags constructed of PVC. Maintenance of normal platelet morphology as determined by phase-contrast microscopy, extent of shape change response, and the size distribution according to the Coulter Counter correlated with recovery in vivo.

scholarly journals Platelet storage at 22 degrees C: effect of type of agitation on morphology, viability, and function in vitro

Blood ◽  
1978 ◽  
Vol 52 (2) ◽  
pp. 425-435 ◽  
Author(s):  
S Holme ◽  
K Vaidja ◽  
S Murphy
Keyword(s):  
Platelet Count ◽  
Shape Change ◽  
Normal Platelet ◽  
Platelet Storage ◽  
Platelet Morphology ◽  
And Function ◽  
Change Response ◽  
Citrate Phosphate

Recovery in vivo after 51Cr labeling, platelet morphology, and platelet aggregation were studied with platelet concentrates (PC) stored for transfusion under carefully controlled conditions. PC were prepared to a final volume of 50 ml from whole blood anticoagulated with citrate- phosphate-dextrose (CPD). The platelet count was kept between 0.8 and 1.6 X 10(12) platelets/liter. The PC were stored in bags constructed of polyvinylchloride (PVC) or polyethylene (PE) at 22 degrees C for 72 hr. The bags were placed on a horizontal shaker or a ferris wheel for agitation during storage. No significant changes in pH or platelet count were observed during storage. PC stored on the wheel showed moderate loss of viability and a marked deterioration of platelet morphology and aggregation compared to the shaker. PC stored on the shaker in bags made of PE showed better aggregation with ADP and thrombin but had the same viability and morphology as PC in bags constructed of PVC. Maintenance of normal platelet morphology as determined by phase-contrast microscopy, extent of shape change response, and the size distribution according to the Coulter Counter correlated with recovery in vivo.

scholarly journals The Potential Role of Thrombopoietin and Interleukin-6 in the Thrombocytosis Effect of Carica papaya Leaves

2020 ◽  
Vol 11 (SPL4) ◽  
pp. 1928-1934
Author(s):  
Laila M Abdelrahim ◽  
Zetty NM Zain ◽  
Siti NS Abdul Jalil ◽  
Zahidah A Seman ◽  
Fadlul AF Mansur ◽  
...  
Keyword(s):  
Platelet Count ◽  
Carica Papaya ◽  
Potential Role ◽  
Scientific Evidence ◽  
Specific Treatment ◽  
Clinical Feature ◽  
Developing Regions ◽  
Methanol Extracts

Dengue fever is endemic in tropical urban developing regions worldwide. Thrombocytopenia is an important clinical feature which may result in bleeding. However, there is no specific treatment for Dengue-induced thrombocytopenia. Carica papaya leaves (CPL) is a popular remedy in South East Asia to treat Dengue-induced thrombocytopenia. Development of CPL into pharmaceutical therapeutic agents is not forthcoming due to lack of rigorous scientific evidence and unknown mechanism of action. This study investigated the role of thrombopoietin (TPO) and interleukin (IL-6) in the thrombocytosis effect of CPL in vivo. These experiments were conducted using busulfan-induced thrombocytopenic rats. Treatment of aqueous and methanol extracts of CPL at 600mg/kg were administered orally for 7 consecutive days and serum platelet count was determined intermittently until day 15. At the end of experiments, serum Thrombopoietin (TPO) and IL-6 levels were determined by ELISA. Both aqueous and methanol extracts of CPL significantly increased platelet count compared to the control groups (x2 (2) = 25.373, P = 0.00). Investigations into the mechanism of thrombocytosis showed that TPO and IL-6 levels were increased compared to controls but was not statistically significant (H (3) = 5.339 P = 0.149) (H (3) = 4.412 P = 0.220) respectively. This study is the first to document the thrombocytosis effect of both aqueous and methanol CPL extracts in a rodent model system. Our findings showed that aqueous extract of CPL demonstrated an increase of TPO and IL-6 levels. We suggested that the possible mechanism could be linked with the upregulation of major thrombopoietic cytokines such as TPO and IL-6.

scholarly journals Resistance to antibiotics in Gram-negative bacteria isolated from broiler carcasses

2002 ◽  
Vol 54 (1) ◽  
pp. 1-7
Author(s):  
M.A.S. Moreira ◽  
C.A. Moraes
Keyword(s):  
Antibiotic Resistance ◽  
Resistant Bacteria ◽  
Processing Plant ◽  
Growth Promoter ◽  
Gram Negative Bacteria ◽  
Beta Lactams ◽  
Gram Negative ◽  
Low Levels

One hundred and ninety-seven isolates of Gram-negative bacteria, comprising 10 genera, were isolated from poultry carcasses at a processing plant in order to investigate resistance to low levels of antibiotics. The samples were taken just after evisceration and before inspection. Most of the isolates were of Samonella and Escherichia. Other genera present were Enterobacter, Serratia, Klebsiella, Kluyvera, Erwinia, Citrobacter, Pseudomonas and Aeromonas. Distinct profiles of antibiotic resistance were detected. Resistance to more than two antibiotics predominated and spanned several classes of antibiotics. Salmonellae and escherichiae were mainly resistant to the aminoglycosides, followed by tetracycline, nitrofuran, sulpha, macrolide, chloramphenicol, quinolones and beta-lactams. Most isolates were sensitive to 30mug/ml olaquindox, the growth promoter in use at the time of sampling. However, many were resistant to a level of 10mug/ml and 13mug/ml olaquindox, levels present in the gut due to the dilution in the feed. The results suggest a possible role of low level administration of antibiotics to broilers in selecting multi-resistant bacteria in vivo.

scholarly journals The Use of Lymphocyte Function–Associated Antigen (Lfa)-1–Deficient Mice to Determine the Role of Lfa-1, Mac-1, and α4 Integrin in the Inflammatory Response of Neutrophils

2001 ◽  
Vol 194 (2) ◽  
pp. 219-226 ◽  
Author(s):  
Robert B. Henderson ◽  
Lina H.K. Lim ◽  
Philippe A. Tessier ◽  
Felicity N.E. Gavins ◽  
Meg Mathies ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Neutrophil Recruitment ◽  
Lymphocyte Function ◽  
Adhesion Receptor ◽  
Low Levels ◽  
First Time ◽  
Deficient Mice ◽  
Β2 Integrins

After injury or infection, neutrophils rapidly migrate from the circulation into tissues by means of an orderly progression of adhesion receptor engagements. Neutrophils have been previously considered to use selectins exclusively to roll on vessels before an adhesion step mediated by the β2 integrins, lymphocyte function–associated antigen (LFA)-1, and Mac-1. Here we use LFA-1−/− mice, function blocking monoclonal antibodies, and intravital microscopy to investigate the roles of LFA-1, Mac-1, and α4 integrins in neutrophil recruitment in vivo. For the first time, we show that LFA-1 makes a contribution to neutrophil rolling by stabilizing the transient attachment or tethering phase of rolling. In contrast, Mac-1 does not appear to be important for either rolling or firm adhesion, but instead contributes to emigration from the vessel. Blocking Mac-1 in the presence of LFA-1 significantly reduces emigration, suggesting cooperation between these two integrins. Low levels of α4β1 integrin can be detected on neutrophils from LFA-1+/+ and −/− mice. These cells make use of α4β1 during the rolling phase, particularly in the absence of LFA-1. Thus LFA-1 and α4β1, together with the selectins, are involved in the rolling phase of neutrophil recruitment, and, in turn, affect the later stages of the transmigration event.

Genome-Wide Chromatin Immunopreciptiation and Gene Expression Analysis Indicates That MMSET Is a Transcriptional Repressor in Vivo

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 627-627
Author(s):  
Dong-Joon Min ◽  
Julia Meyer ◽  
Eva Martinez-Garcia ◽  
Josh Lauring ◽  
Jonathan D. Licht
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Line ◽  
Signaling Pathway ◽  
Target Genes ◽  
Transcriptional Repressor ◽  
Low Levels

Abstract Multiple myeloma (MM) is a malignancy of plasma cells characterized by frequent chromosomal translocations of the immunoglobulin heavy chain (IgH) locus. The multiple myeloma SET domain (MMSET) gene is a recurrent chromosomal partner in the t(4;14) translocation, and MMSET levels are elevated in these patients relative to other myeloma cases and normal cells. Previously, we showed that MMSET is a histone methyltransferase with specific activity for lysine 20 on histone H4 and acts as a transcriptional repressor when tethered to a model target gene. To reveal the function of MMSET in t(4;14) MM in vivo, we identified MMSET target genes in the KMS11 t(4;14) MM cell line. Chromatin from these cells was subjected to immunoprecipitation with a polyclonal anti-MMSET antibody in biological replicate, amplified by ligation-mediated PCR and hybridized to NimbleGen 2.7kB promoter arrays, which represent 24,659 human promoters. Data analysis using the MaxFour algorithm ranked putative binding sites based upon intensities of 4 consecutive probes. The top 2000 promoters identified from each experiment were combined to yield a list of 1,412 putative MMSET target genes. This list was analyzed using the DAVID program (david.abcc.ncifcrf.gov/). Genes bound by MMSET includes those implicated in antigen processing and presentation (p<8.7×10-4), cell cycle (p<2.2×10-3), the p53 signaling pathway (p<0.03) apoptosis (p<1.6×10-6) and DNA repair (p<3.3×10-4) Among genes bound by MMSET were XBP1, IRF2, and BCL6, all important transcription factors regulating B cell development. Real-time quantitative PCR validated MMSET binding in 6/6 promoters tested so far. To investigate the role of MMSET in transcriptional regulation, we profiled gene expression in KMS11 cells using Illumina arrays to determine the expression of MMSET bound genes. Nearly 50% of genes bound by MMSET had very low levels of expression (≤100, Range on arrays 10–18,000) while only 13% of genes bound by MMSET were expressed at high levels (>1000). This supports the notion that MMSET represses target genes in vivo. Functional annotation of genes bound by MMSET and expressed at low levels showed over-representation of genes implicated in toll-like receptor signaling pathway (p<2.8×10-3), cytokine-signaling (p<3.3×10-3) and JAK2/STAT signaling (p<0.08), transmembrane receptor function (p<4×10-8), and apoptosis (p<0.01), while those bound yet expressed at high levels were implicated in oxidative phosphorylation (p<3.9×10-4) and protein synthesis (p<4.1×10-6). The effects of MMSET on gene expression were further investigated using KMS11KO cells in which the rearranged MMSET allele was ablated by homologous recombination. RNA from KMS11 and KMS11KO cells was profiled by Illumina arrays and genes showing a significant change in gene expression were determined by significance analysis of microarray (SAM) testing with 1% of false discovery rate. Among the 720 genes bound by MMSET and expressed at a level of >100 in the wild-type KMS11 cells, 35 genes were up-regulated and 20 genes were down-regulated (>1.5 fold) in the KMS11KO cell line. Among the 692 genes bound by MMSET and expressed at a level of ≤100, 9 genes were up-regulated in KMS11KO cells. The up-regulated genes (presumably bound and repressed by MMSET) were categorized in cytokine receptor (p<0.02) and JAK2/STAT signaling pathway (p<0.05), nucleosome assembly (p<6×10-4), apoptosis (p<0.01), and cell differentiation (p<0.05). Collectively these data suggest that MMSET may interfere with signal transduction, chromatin modulation and apoptosis pathways involved in the terminal differentiation of the plasma cell. Intriguingly MMSET also bound and was associated with repression of RB1 and RBL2 suggesting a role of MMSET in cell cycle control. Chromatin immunoprecipitation analysis of a MMSET bound gene (ARHGAP25) revealed that MMSET binding was correlated with increased tri-methylation of H4K20, a repression-associated chromatin mark. MMSET binding of this promoter was decreased but still detectable in the KMS11KO cells. Collectively these data suggest that MMSET binds and represses many target genes in vivo. However MMSET could still bind to genes expressed at a high level and MMSET ablation was associated with activation of some MMSET target genes, suggesting that its role in gene regulation may be complex and potentially gene-specific.

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