| BackgroundChronic lymphocytic leukemia(CLL),as the most common leukemia in the western countries,accounts for-30%of adult leukemia and 25%of non-Hodgkin lymphoma(NHL).It is a neoplastic disease of advancing age,characterized by accumulation of functionally defective B-lymphocytes in the blood,bone marrow,spleen,lymphoid notes,and other organs,leading to functional failure and patient death.Among B-cell malignancies,CLL is a shining exception because it is far from uniform in presentation and clinical course.The symptoms of CLL tend to develop over time.For many patients,CLL symptoms may at first seem to be some kind of non-specific change in overall health,and then there may be an increased sense of fatigue or weakness.Some people may experience flu-like symptoms,like night sweats or enlarged lymph nodes.Many people are diagnosed with CLL because of a blood test for an unrelated condition.The relatively indolent nature of CLL enables detailed investigation of tumor cells,often in the absence of treatment,as well as observation of tumor behavior over time.This window of opportunity has facilitated the identification of prognostic factors that relate to pathogenesis.Furthermore,in CLL,there is the added advantage of availability of tumor cells from blood,although conclusions based on this compartment have to be tempered by the fact that critical proliferative events occur in tissue sites.B-cell receptor(BCR),responsible for the activation of B cells,plays a vital role in the pathogenesis and the progression of CLL.It is composed of a membrane immunoglobulin(IgM)noncovalently bonded to a heterodimer composed of CD79a(Iga)/CD79b(Igp).When IgM is ligated by a soluble or membrane-bound antigen,tyrosine residues in the cytoplasmic ITAM portion of CD79a and CD79b are phosphorylated by the Src family kinase Lyn and spleen tyrosine kinase(Syk).This phosphorylation results in recruitment of the signalo-some,which includes many kinases and adaptor proteins,including the kinases Syk,Bruton tyrosine kinase(Btk),and Lyn,the guanine exchange factor Vav proteins,and the adaptor proteins Grb2 and B-cell linker.The protein tyrosine kinases are primarily responsible for amplification of this activation.BTK,a member of the Tec family kinases,has a well-characterized role in BCR-signaling that regulates B-cell survival,activation,differentiation,and interaction with the microenvironment.It is recruited to the membrane-bound signalosome in the early stages of B-cell activation,and,following the activation by Src-family kinases,participates in the phosphorylation of phospholipase C,gamma 2(PLCy2),which leads to production of the second messengers diacylglycerol and inositol-1,4,5-triphosphate.This pathway is amplified in CLL and leads to prosurvival signals through its effects on phosphatidylinositol 3-kinase(PI3K),PLCy2,nuclear factor-?B(NF-?B).and MAPK.Although BTK is expressed in multiple hematopoietic cells,the primary defect in knockout mice is B cell-specific,suggesting a more selective B-cell function.BTK mutations in humans give rise to X-linked agammaglobulinemia,an inherited disorder that is characterized by severe B cell-specific defects including severely decreased levels of immunoglobulin production and the absence of B cells;further suggesting the importance and selectivity of BTK to B cells.While understanding of CLL biology has improved dramatically,until very recently integration of these findings to treatment interventions has been lacking.Given that BTK is critical for B-cell survival,the inhibition of BTK has recentlyemerged as a crucial strategy for treating CLL.Ibrutinib,a US Food and Drug Administration-approved,orally administered irreversible inhibitor of BTK,has shown to be highly effective in controlling relapsed/refractory,previously untreated or 17p deletion CLL.Ibrutinib is generally well tolerated but is associated with an increased risk of bleeding.Bleeding has been reported in up to 50%of ibrutinib-treated patients.Grade 3 or higher bleeding events(intracranial hemorrhage,gastrointestinal bleeding,hematuria,and post-procedural hemorrhage)have occurred in up to 6%of patients.Clinical observation showed that ibrutinib may increase the risk of hemorrhage in patients receiving antiplatelet or anticoagulant therapies.Furthermore,previous studies found that platelets from patients receiving ibrutinib treatment exhibit deficient responses to collagen,despite no defects in classical coagulation parameters(for example,prothrombin time and activated partial thromboplastin time).However,until now the underlying mechanism of ibrutinib affecting platelet function is still not well understood.As we know,platelets are the most important blood cells to prevent bleeding after vascular injury.They are highly reactive circulating blood cells derived from megakaryocytes.Like red blood cells,platelets lack nuclei and consequently are unable to replace damaged proteins encoded in the nuclear genome.However,unlike red blood cells,platelets contain actively metabolizing mitochondria.For many years,platelet mitochondria were viewed only as the `power-house',providing ATP to meet a significant portion of energy demands of quiescent cells.However,accumulating evidence suggests that mitochondria regulate platelet activation beyond simply providing ATP.Treatment of platelets with exogenous hydrogen peroxide has been shown to lead to activation,and intracellular ROS generation has been implicated as an integral second messenger in collagen-induced activation of platelets.Whereas the exact downstream targets of the ROS are not completely clear,scavenging of intracellular hydrogen peroxide has been shown to activate phospholipase C and arachidonic acid metabolism leading to the inhibition of calcium mobilization,ATP release from granules and platelet aggregation.Although multiple source of ROS exist within the platelet,the mitochondrial electron transport chain can generate significant concentration of superoxide,which are rapidly converted into hydrogen peroxide by superoxide dismutase.In CLL patients,cytogenetic alterations such as chromosomal aberrations are rather common.Some of the cytogenetic changes are associated with poor prognosis and aggressive disease progression.Of note,chromosome 17p deletion(17p-)is a prominent cytogenic alteration in CLL associated with resistance to chemotherapy and poor clinical outcomes.As the tumor suppressor p53 gene is located in human chromosome 17p,it is suspected that the loss of p53function in CLL cells with 17p-may be responsible for the poor prognosis of this subgroup of CLL patients.Interestingly,recent study suggests a very high concordance(over 70%)in 17p deletion and mutations in the remaining p53 allele.Furthermore,p53 dysfunction may also arise via alternative mechanisms such as functional inactivation,which may explain certain CLL cases with poor prognosis but without apparent structural changes in p53 gene such as 17p deletion or mutations.Thus,it seems clear that the loss of p53 function has a profound effect on CLL response to drug treatment and the disease development.However,the underlying mechanisms remain to be elucidated.MicroRNAs(miRNAs)are a family of small,non-coding RNAs that regulate the expression of target messenger RNAs at the post-transcriptional level.Previous studies have suggested that miRNAs are extensively involved in the proliferation and differentiation of hematopoietic cells.Aberrant expression of certain miRNAs has been observed in CLL.Associations between miRNAs and chromosomal abnormalities suggest that miRNAs may be involved in the pathogenesis of CLL with 17p deletion.In this study,we first investigated the biological effect of ibrutinib on collagen-induced platelet aggregation.Next,to further investigate the underlying mechanism,we performed a comparison of the effects of short-term and long-term ibrutinib treatment on mitochondrial respiratory chain using clinical platelet samples and platelets from megakaryocytes.We identified that ibrutinib suppresses mitochondrial respiration,leading to defects in collagen-induced platelet aggregation,potentially through inhibiting the activation of the Syk pathway.Moreover,we generated a mouse colony with TCL1 transgenic and p53 deletion(TCL1-Tg:p53-/-)genotype by crossing the TCL1-Tg mice with p53-/-mice.The TCL1-Tg:p53-/-mice develop leukemia that seems to resemble human aggressive CLL disease.The leukemia cells from TCL1-Tg:p53-/-mice exhibited higher proliferation and survival capacity than the leukemia cells from the TCL1 transgenic mice.Further analysis of microRNA expression profile showed significant decrease in miR-15a and miR-16-1 expression,suggesting miRNAs may be involved in the development of 17p deletion CLL.Materials and methodsCell lines,reagents,antibiotics and antibodies.MEG-01 cells were cultured in RPMI-1640(GIBCO)supplemented with 10%fetal bovine serum(GIBCO).Cells were maintained at 37 C in a humidified atmosphere of 5%C02.Antibodies specific to OXPHOS and ?-actin were from Abcam Inc.'Antibodies specific to p-Syk,Syk and p-Tyr were from Cell Signaling Technology Inc.Platelet isolation from MEG-01 cellsPlatelet-like particles were isolated from differentiated MEG-01 cells by sequential centrifugation.Briefly,cells were pelletted at 150 x g for 10 min and the resulting supernatant was centrifuged again at 500 x g for 15 min to remove all contaminant cells.Platelets were further centrifuged at 1000 x g for 15 min and resuspended in Tyrode's buffer(100 mM Hepes,1.3 M NaCl,29 mM KC1,120 mM NaHC03,pH 7.4),containing 1/10(v/v)ACD(112 mM glucose,130 mM citric acid,152 mM sodium citrate)and 2 mM glucose.Platelet isolation from clinical patientsBlood was drawn from healthy donors that had given informed consent and using procedures approved by the University of Reading Research Ethics Committee and collected into 50 ml syringes containing 4%(w/v)sodium citrate and acid citrate dextrose(ACD,2.5%sodium citrate,2%D-glucose and 1.5%citric acid).PRP was prepared by centrifugation of whole blood at 100g for 20 mins.Washed platelets were prepared by centrifugation of PRP at 800g for 20 mins,the platelet pellet wasresuspended in Tyrode's buffer(134 mM NaCl,12 mM NaHC03,2.9 mM KC1,0.34 mM Na2HP04,1 mM MgC12,10 mM Hepes,pH 7.4)containing 5 mM glucose and 3 mg/ml BSA.Flow cytometryImmunophenotyping of cells was analyzed by flow cytometry in a FACSCanto instrument(Beckton Dickinson).Briefly,MEG-01 cells or platelet-like particles were fixed in 4%paraformaldehyde and stained with appropriate cluster of differentiation(CD)antibodies.The antibodies used were:fluorescein isothiocyanate(FITC)-conjugated mouse anti-human CD49b and PE-conjugated mouse anti-human CD41a(Beckton Dickinson).For each analysis,10 thousand events were acquired and analyzed using the Flowjo software(TreeStar).Flow cytometric analysis of cellular ROSCellular ROS levels were quantified by flow cytometry analysis.Cells or platelets were transferred to a 24-well plate and incubated with 1 ?M CM-H2DCF-DA(Invitrogen,Carlsbad,CA)in regular culture medium for 60 min at 37? in the dark.Stained cells were rinsed twice with PBS and analyzed immediately by flow cytometry using a FACSCalibur equipped with CellQuest Pro software.Platelet aggregation in a microplate readerPlatelets in plasma or buffer were aggregated in a microplate reader instrument type EL340(Bio-Tek Instruments).The instrument could maintain room temperature during the experiment period and continuously shake(forth and back)the microplate between readings(20 see).For the aggregation process temperature 370C was set and readings were performed with a 595 nm filter and were recorded in a computer memory for further evaluation.Aggregation process was carried out in 100 u1 volume in a 96 well flat-bottom microplate(COSTAR).Collagen was added to PRP or platelet suspension,using a multichannel pipette(COSTAR),just before start of the reading.The sample surfaces in a microplate have to be free from any air-bubbles in order to get well reproducible and smooth aggregation curves.Controls for 100%transmission(PPP or buffer)and zero level(PRP or platelet suspension)were observed during the whole period of measurements.The last control allowed us to observe a spontaneous aggregation of platelets under the experimental conditions.The collected data were finally converted to aggregation curves and evaluated using our own computer program.Oxygen consumption rates02 consumption was determined using Seahorse XF24 extracellular Flux analyzer as previously described by Qian and Van Houten 24.Briefly,cells were seeded in the 24-well XF24 cell culture plate in the respective culture media.For MEG-01 cells,10000 cells were seeded in unbuffered RPMI-1640 and plates were incubated at 37?for 30 minutes for pH and temperature stabilization.Three mitochondrial inhibitors;oligomycin(1 mM),FCCP(500 mM)and rotenone(2 mM)were sequentially injected after measurements,respectively.After all the measurements were completed,the collected data were finally converted to oxygen consumption curves.RNA extraction and reverse transcriptionTotal RNA was prepared by using the RNeasy Mini Kit(Qiagen)following the manufacturer's protocol.For formalin-fixed paraffin-embedded(FFPE)samples,total RNAwas extractedfrom five totwenty 10 ?m-thick sectionsusing miRNeasy FFPE Kit(Qiagen)according to the manufacturer's instructions.The quantity and quality of the total RNA were evaluatedwith the NanoDrop spectrophotometer(Thermo Fisher ScientificIncorporated,USA)according to the manufacturer's instructions.One ?g of total RNA from each sample was reverse-transcribed(RT)to cDNA with PrimeScript reverse transcriptase reagent kit(TaKaRa,China)according to the manufacturer's instructions.Real-time Quantitative RT-PCR of mRNA and miRNAQuantitative real-time PCR was carried out in ABI Prism 7500 Sequence Detection System(Applied Biosystems,Foster City,CA)using SYBR Premix Ex Taq(TaKaRa)according to the manufacturer's instructions.The miRNA sequence-specific RT-PCR for miR-15a/16-1 and endogenous control U6 was performed according to Hairpin-itTM miRNAs,qPCR quantitation kit and U6 snRNA real-time PCR normalization kit(GenePharma).All quantitative PCR reactions,including no-template controls,were performed in triplicate.Expression levels ofeach mRNA or miRNA were evaluated using comparative threshold cycle(Ct)method as normalized to that of GAPDH or U6(2??CT).Western blotting and co-immunoprecipitationCells were lysed and sonicated in a solution containing 0.5%sodium deoxycholate(w/v),0.2%SDS(w/v),1%Triton X-100(v/v),5 mM EDTA,10 mg/ml leupeptin,10mg/ml aprotinin and 1mM phenylmethyl sulfonyl-uoride supplemented with 1:1000 dilution of protease inhibitor cocktail.The homogenates were then centrifuged at 40C for 10min to remove cell debris.Supernatants were harvested and concentrations were determined by the DC protein assay(Bio-Rad Laboratories,Hercules,CA).Equivalent amounts of protein were electrophoresed on an 8%polyacrylamide gel and then transferred to a PVDF membrane.For Western blotting,the membrane was blocked with 5%milk(w/v)in TBS containing 0.1%Tween 20(TBS/T)at room temperature for 2h and incubated with primary antibodies including OXPHOS,p-Syk,Syk,p-Tyr and?-actin overnight at 40?,washed three times with TBST,followed by incubation with appropriate secondary antibodies at room temperature for 1h.The immune complexes were detected by an enhanced chemiluminescence(ECL)system(Amersham,Arlington Heights,IL,USA).miRNA expression arrayTotal RNA was isolated from spleen cells from TCL1-Tg and TCL1-Tg:p53_/_mice or from human CLL cells with or without 17p deletion,using a microRNA isolation kit(Ambion,Grand Island,NY,USA).For analysis of microRNA expression profiles,5 mg total RNA isolated from three mice or three CLL patient samples per group was analyzed using a commercial microRNA array(LC Sciences,Houston,TX,USA).To independently validate the individual microRNA expression pattern from the microRNA array result,total RNA was converted to cDNA synthesis using Taqman MicroRNA Reverse Transcription kit and Taqman RT primers(Applied Biosystems,LLC,Foster city,CA,USA).MicroRNA-specific real-time PCRs were performed using Taqman Universal PCR Master Mix and Taqman small assays according to the manufacturer's recommended protocol.Statistical analysisData are represented as mean± SD.Possible differences between groups were analyzed using Independent-samples T test or one-way ANOVA..Survival curves were obtained by Kaplain-Meier analysis.A P value<0.05 was considered significant.All statistical analyses were carried out with SPSS 17.0 software(SPSS Inc.,Chicago,IL,USA).ResultsIbrutinib has no effect on platelet count and MEG-01 cell proliferationTo understand the effect of ibrutinib on platelet count,we collected 9 CLL patients who had received ibrutinib treatment.Comparison of pre-and post-treatment platelet information showed that only 1 of 9 patients had sharp decrease in platelet count,while for other patients platelet counts changed slightly.To test the effect of ibrutinib on platelet production in vitro,we first treated MEG-01 cells with or without 0.5 uM ibrutinib for 1,3,or 5 days and then isalated the platelets from MEG-01 cells.Given that platelets express CD41a and CD49b.flow cytometry was performed to detect the expression of these specific platelet surface antigens,confirming the platelet production from MEG-01 cells.Next,the platelets were counted by Beckman coulter.Results showed that no significant difference between ctrl group and treatment group.In summary,ibrutinib has no obvious effect no matter on clinical platelet samples or MEG-01 derived platelets.We next determine whether ibrutinib could influence MEG-01 cell proliferation.The MTT assay was performed to measure cell proliferation after incubation with ibrutinib for 1,2,3 or 4 days.The viability of MEG-01 cells incubated with ibrutinib hardly changed compared with that of ctrl.These data suggested no effect of ibrutinib on MEG-01 cell proliferation.Ibrutinib inhibits collagen-induced platelet aggregationPlatelet rich plasma(PRP)or platelets released from MEG-01 were preincubated with or without ibrutinib for 30min or 1h.Compared to untreated controls,collagen-mediated aggregation was significantly decreased by ibrutinib.Platelets released from MEG-01 had a 34%reduction in response to collagen after ibrutinib treatment,and PRP had a 37%reduction induced by ibrutinib incubation.Short-term ibrutinib incubation increases ROS level in both platelets and MEG-01 cellsROS,as the second intracellular signal,exerts a vital role in the process of collagen-induced platelet activation.The deficiencies in platelet aggregation induced by ibrutinib promoted us to further investigate whether ROS was involved in this side effect.We first treated either platelets or MEG-01 cells with ibrutinib and then analyzed the variation of ROS level by flow cytometry.Results showed that ibrutinib could significantly increase ROS level in both platelets and MEG-01 cells.Short-term ibrutinib treatment dampens oxidative phosphorylation in MEG-01 cellsMitochondrial electron transport chain is the major origin of ROS,thus abnormal ROS level may be caused by the deficiency in mitochondrial electron transport chain.To test this hypothesis,we determine 02 consumption rate(OCR)using the Seahorse XF24 extracellular flux analyzer.We have used a pharmacological profiling approach,by combining the use of three mitochondrial inhibitors(rotenone,FCCP and oligomycin)and the Seahorse instrument.The OCR in response to the chemical compounds were analyzed in MEG-01 cells with or without ibrutinib treatment.The basal OCR(measure of OXPHOS)is the amount oxygen consumption that is linked to ATP synthesis in the mitochondria and represents the mean basal levels of oxygen consumption minus the mean of the two values following oligomycin treatment.Oligomycin inhibits ATP synthase by binding to the oligomycin sensitivity-conferring protein(OSCP)at the F0 subunit of the ATP synthase.This binding blocks the proton conductance resulting in loss of electron transfer and 02 consumption.Addition of oligomycin resulted in decreased levels of OCR.MEG-01 cells without ibrutinib treatment showed higher basal 02 consumption rates when compared to that with treatment.FCCP is a mitochondrial uncoupler that dissipates the proton gradient,and therefore uncouples electron transport and mitochondrial respiration from ATP synthesis.FCCP treatment has been shown to increase 02 consumption to its maximum in various cell types.In fact FCCP resulted in an increase of OCR levels in both groups,with the more pronounced response being observed in the untreated group.The difference between FCCP OCR and basal OCR is a good measure of respiratory capacity(also designed of reserve capacity)of these cells.Indeed we observed that the respiratory capacity of untreated MEG-01 cells is higher than that observed for ibrutinib-treated cells.Rotenone is a mitochondrial inhibitor that prevents electron transfer from complex ? to ubiquinone by blocking the ubiquinone binding site.Treatment with rotenone decreased OCR levels in both groups although untreated MEG-01 showed a more pronounced response to the treatment.Overall these data suggest that ibrutinib-treated cells exhibited a lower mitochondrial activity rate,leading to a dramatic reduction in OXPHOS.Rotenone and antimycin mimic the effect of ibrutinib on plateletsRotenone is a mitochondrial complex ? inhibitor and antimycin is a mitochondrial complex ? inhibitor.Since ibrutinib could dampen the activity of mitochondrial electron transport chain,we further test whether rotenone and antimycin could induce similar effects as ibrutinib.Flow cytometry was performed to detect the ROS level in platelets,after rotenone or antimycin treatment,and results showed that both treatments could enhance ROS production,which is similar as ibrutinib.Furthermore,similar results also could be observed in platelet aggregation assay.Both rotenone and antimycin could impair platelet aggregation.Taken together,these results further indicated that ibrutinib may impair platelet aggregation though inhibition of mitochondrial complex activity.Ibrutinib has no effect on OXPHOS complex gene and protein expressionTo further detect whether the inhibition of mitochondrial complex activity comes from the reduction of mitochondrial complex expression,we analyzed OXPHOS expression at both mRNA and protein levels after short-term ibrutinib treatment.Results showed that short-term ibrutinib incubation had no effect on OXPHOS complex gene and protein expression.Long-term ibrutinib treatment suppress ROS production and OXPHOS expressionGiven that platelets from clinical patients could survive in vitro for only 4-5 hours,we treat clinical platelet samples with ibrutinib for a short time.But it is not the case in clinical practice.Patients take ibrutinib daily,and platelets are affected by ibrutinib for a long time.So we further analyze the effect of long-term ibrutinib treatment.ROS level analysis reflected that MEG-01 cells had an enhanced ROS production when treated with ibrutinib for only 1 day.However,ROS level decreased slowly with time,and finally was lower than untreated cells after ibrutinib treatment for 7 days.OXPHOS expression analysis after long-term ibrutinib treatment also is different from that after short-term treatment.We incubated MEG-01 cells and platelets with or without ibrutinib for 3 days and then analyze OXPHOS expression.Western blotting showed that OXPHOS expression obviously decreased at both gene and protein level,suggesting long-term ibrutinib treatment could inhibit OXPHOS expression,whch may account for the reduction of ROS production.OXPHOS expression in CLL patients that received ibrutinib treatmentWe collect the pre-and post-treatment blood samples from 9 CLL patients.Platelets were isolated from blood to detect the change of OXPHOS expression.Results showed that 6 of 9 patients had decreased OXPHOS expression after ibrutinib treatment for 28 days.TCL1-Tg:p53-/-mice develop aggressive CLL with early disease onset and short lifespanP53 is one of the most frequently mutated genes in cancers.Currently there is no CLL mouse model with loss of p53 for investigating the pathological process of this aggressive CLL.To create such an animal model,we used the well characterized TCL1 transgenic CLL mice to cross breed with p53-/-mice to generate progenies that harbor TCL1-Tg:p53-/-genotype.The TCL1-Tg:p53-/-mice developed CLL disease with early disease onset at the age of?3 months,and most of the mice developed severe splenomegaly by 4-5 months,while the spleens of TCL1-Tg mice with wt p53 or p53-/-mice appeared relatively normal in size.The significant difference in the weights of the spleens from 4-month-old TCLl-Tg mice and 4-month-old TCL1-Tg:p53-/-mice is shown.Furthermore,blood smear revealed significant expansion of the white blood cells(WBC)in TCLI-Tg:p53-/-mice compared with TCLl-Tg mice.Most of the TCL1-Tg:p53-/-mice died at the age of 3-5 months,while most of the TCL1-Tg mice survived more than 12 months.This was consistent with the clinical observations that CLL patients with 17p deletion have significantly shorter overall survival compared with the CLL patients without 17p deletion.MiRNA expression arrayFor analysis of microRNA expression profiles,total RNA isolated from mice was analyzed using a commercial microRNA array.Among the>300 miRNAs detected by the microarray analysis,miR-15a and miR-16-1 consistently showed a marked decrease in the spenocytes of Tg:p53-/-mice compared to that of the TCL1-Tg mice.The decrease of miR-15a and miR-16-1 in cells from TCL1-Tg:p53-/-mice was further confirmed by real-time RT-PCR using the leukemia cells isolated from the spleens and PCs of TCLl-Tg and TCL1-Tg:p53-/-mice.Importantly,the decrease in miR-15a/16 expression was further confirmed in primary CLL cells isolated from five patients with 17p deletion.These data together suggest that suppression of miR15a/miR16-1 expression may be an important mechanism for the development of CLL with 17p deletion.Conclusion1.ibrutinib had no effect on platelet count but could inhibit collagen-inducedplatelet aggregation.2.Short-time ibrutinib treatment suppresses the activity of mitochondrial respiratorychain and impedes ROS production.3.Long-term ibrutinib treatment impairs ROS production through inhibiting OXPHOS expression.4.The inhibition of mitochondrial respiratory chain activity could dampen the activation of syk signaling,which is required for collagen-induced platelet aggregation.5.TCL1-Tg:p53-/-mice develop aggressive CLL with early disease onset and short lifespan.6.MiR-15a and miR-16-1 are decreased by 17p deletion in CLL.Taken together,our data showed in the current study confirm the effect of ibrutinib on collagen-induced platelet aggregation.Moreover,we provide the first line of evidence suggesting that Ibrutinib inhibits platelet aggregation through inhibition of mitochondrial respiratory chain function and suppression of Syk activation.In addition,we created TCL1-Tg:p53-/-mouse mode to investigate the pathological process of 17p deletion CLL.miRNA array analysis showed miR15a/miR16-1 were significantly decreased,suggesting that they may be involved in the mechanism for the development of CLL with 17p deletion. |
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