Integrated analysis of the functions and prognostic values of RNA binding proteins in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC), one of the most common malignant tumors worldwide, ranks as the fifth most common cancer and has been the second most frequent cause of cancer-related death. RNA binding proteins (RBPs) are proteins that interact with different classes of RNA and are commonly detected in cells. METHODS: We used RNA sequencing data from TCGA to display dysfunctional RBPs microenvironments and provide potential useful biomarkers for HCC diagnosis and prognosis. RESULTS: 330 differently expressed RBPs (208 upregulated and 122 downregulated) were identified. KEGG were mainly enriched in RNA degradation, Influenza A, Hepatitis C, RIG-I-like receptor signaling pathway, Herpes simplex virus 1 infection and RNA transport. CBioPortal results demonstrated that these genes were altered in 50 samples out of 357 HCC patients (14%) and the amplification of BRCA1 was the largest frequent copy-number alteration. CONCLUSION: Based on the online database, we identified novel RBPs markers for the prognosis of hepatocellular carcinoma.

Host genetics in malaria: lessons from mouse studies.

Malaria remains a deadly parasitic disease caused by Plasmodium, claiming almost half a million lives every year. While parasite genetics and biology are often the major targets in many studies, it is becoming more evident that host genetics plays a crucial role in the outcome of the infection. Similarly, Plasmodium infections in mice also rely heavily on the genetic background of the mice, and often correlate with observations in human studies, due to their high genetic homology with humans. As such, murine models of malaria are a useful tool for understanding host responses during Plasmodium infections, as well as dissecting host-parasite interactions through various genetic manipulation techniques. Reverse genetic approach such as quantitative trait loci studies and random mutagenesis screens have been employed to discover novel host genes that affect malaria susceptibility in mouse models, while other targeted studies utilize mouse models to validate observation from human studies. Herein, we review the findings from the past and present studies on murine models of hepatic and erythrocytic stages of malaria and speculate on how the current mouse models benefit from the recent development in CRISPR/Cas9 gene editing technology.

Low-dose bortezomib and dexamethasone as primary therapy in elderly patients with WaldenstrÓ§m macroglobulinemia.

OBJECTIVE: This retrospective study was designed to determine the efficacy and safety of low-dose bortezomib and dexamethasone (lBD) in elderly Chinese patients with WaldenstrÓ§m macroglobulinemia (WM). METHODS: Ten patients with WM aged over 60 years received first-line treatment with lBD. RESULTS: The median age was 70 years (range, 61-77 years). The overall response rate was 80%, including 1 patient who achieved a complete response, 1 patient with very good partial response, and 6 patients with a partial response. Median time to response was 1.8 months after treatment with lBD. Six (60%) patients achieved a partial response, including 2 (20%) patients who had a more than 75% reduction in serum immunoglobulin M levels. A rapid reduction in paraprotein was observed in three patients who received plasmapheresis. After a median follow-up period of 36 months, all patients were still alive and six had no disease progression. The estimated median time to progression was 39 months (range, 15-60 months). The most common adverse events were anemia, thrombocytopenia, neuropathy, and neutropenia. Peripheral neuropathy was the most common non-hematological toxicity in six (60%) patients, but did not result in the discontinuation of bortezomib. CONCLUSIONS: Our findings show that lBD is an effective and tolerable treatment regimen for elderly patients with WM.

[Expression and Clinical Significance of LINC00475 in Multiple Myeloma].

OBJECTIVE: To investigate the relative expression level and clinical significance of LINC00475 in serum of patients with multiple myeloma (MM). METHODS: The expression of LINC00475 in serum of 108 MM patients and five MM cell lines including RPMI 8226, NCI-H929, U266, OPM2 and CAG were detected by real-time fluorescence quantitative PCR. The diagnostic value of LINC00475 in MM was evaluated by receiver operating characteristic (ROC) curve analysis. The correlation of LINC00475 with patients' characteristics was analyzed. RESULTS: Compared with control groups, the expression of LINC00475 was up-regulated in serum of MM patients and MM cell lines (all P < 0.05). ROC curve analysis showed that the optimal cut-off value of LINC00475 was 262.4, the area under curve (AUC) was 0.924(95%CI : 0.884-0.964), and sensitivity and specificity was 83.3% and 91.7%, respectively, which indicated that LINC00475 had good evaluation value in MM patients. Compared with low-LINC00475 expression group, patients in high-LINC00475 expression group had higher levels of ß2microglobulin (ß2-MG) and Cystatin C (Cys-C) but lower albumin (ALB) (all P < 0.05). Compared with MM patients with International Staging System (ISS) stage I, the expression level of LINC00475 was significantly higher in patients with stage II and III (both P < 0.05). CONCLUSION: LINC00475 is helpful to distinguish MM patients from healthy adults, which is correlated with the prognostic indicators such as ß2-MG, ALB, and ISS stage.

Effective Invasiveness Recognition of Imbalanced Data by Semi-Automated Segmentations of Lung Nodules.

Over the past few decades, recognition of early lung cancers was researched for effective treatments. In early lung cancers, the invasiveness is an important factor for expected survival rates. Hence, how to effectively identify the invasiveness by computed tomography (CT) images became a hot topic in the field of biomedical science. Although a number of previous works were shown to be effective on this topic, there remain some problems unsettled still. First, it needs a large amount of marked data for a better prediction, but the manual cost is high. Second, the accuracy is always limited in imbalance data. To alleviate these problems, in this paper, we propose an effective CT invasiveness recognizer by semi-automated segmentation. In terms of semi-automated segmentation, it is easy for doctors to mark the nodules. Just based on one clicked pixel, a nodule object in a CT image can be marked by fusing two proposed segmentation methods, including thresholding-based morphology and deep learning-based mask region-based convolutional neural network (Mask-RCNN). For thresholding-based morphology, an initial segmentation is derived by adaptive pixel connections. Then, a mathematical morphology is performed to achieve a better segmentation. For deep learning-based mask-RCNN, the anchor is fixed by the clicked pixel to reduce the computational complexity. To incorporate advantages of both, the segmentation is switched between these two sub-methods. After segmenting the nodules, a boosting ensemble classification model with feature selection is executed to identify the invasiveness by equalized down-sampling. The extensive experimental results on a real dataset reveal that the proposed segmentation method performs better than the traditional segmentation ones, which can reach an average dice improvement of 392.3%. Additionally, the proposed ensemble classification model infers better performances than the compared method, which can reach an area under curve (AUC) improvement of 5.3% and a specificity improvement of 14.3%. Moreover, in comparison with the models with imbalance data, the improvements of AUC and specificity can reach 10.4% and 33.3%, respectively.

[Analysis of the efficacy and compliance of conventional immunotherapy and rush immunotherapy in patients with allergic rhinitis].

Objective:To analyze the efficacy and compliance of conventional immunotherapy(CIT)and rush immunotherapy(RIT)in patients with allergic rhinitis.Method:This trial was a prospective study involved 404 patients with persistent AR who were allergic to house dust mite.328 patients were assigned to the conventional immunotherapy reaching the maintenance dose within 14 weeks,and 76 patients were assigned to the rush immunotherapy reaching the maintenance dose within 1 week.The visual analog scale(VAS)score and the patients' compliance were recorded during treatment and follow-up.Result:After CIT and RIT,the VAS score were significantly reduced in each group,but the decrement of VAS score of RIT group was more evident than that of CIT in half ayear(P<0.05).After 5 years follow-up,the VAS score of two groups was also significantly reduced.The rate of treatment continuation of CIT group in 1 year,2 years and 3 years were 18.5%,39.0% and 57.3%,higher than RIT group(11.8%,26.3%,42.1%),respectively.Conclusion:Both CIT and RIT were beneficial for allergic rhinitis patients,and the clinical efficacy lasts for at least 5 years.But RIT has the superiority of faster onset and better compliance.

[Assessment of the horizontal semicircular canal function after cochlear implantation by video head impulse test and caloric test].

Objective:To analyze the functional change of horizontal semicircular canals after cochlear implantation.Method:Eighteen patients were enrolled in this study.Their vestibular function was evaluated by using the caloric test and video head impulse test before and one week,one month after CI surgery,respectively.The unilateral weakness(UW),slow phase velocity(SPV)in caloric test and gain in video head impulse test(vHIT-G)were observed.Caloric test was abnormal when UW>25% or SPV mean<6°/s,while vHIT was abnormal when vHIT-G<0.8.Result:The SPV of the implanted ear were[(10.36±8.01)°/s;(14.77±14.24)°/s]pre-operatively,[(6.45±7.52)°/s;(5.14±4.67)°/s]1 week post-operatively and[(6.05±3.86)°/s;(6.27±4.17)°/s]1 month post-operatively.Statistically significant difference(P<0.05)was found between pre-and post-operative period.The vHIT-G of the implanted ear were(0.73±0.33)pre-operatively,(0.65±0.32)1 week post-operatively and(0.71±0.36)1 month post-operatively.There was no statistically significant difference of vHIT-G between preand post-operative period(P(pre-operative/1 week post-operative)=0.084,P(pre-operative/1 month post-operative)=0.679).Four patients presented with vertigo and one of them manifested slight unsteadiness post-operatively.All symptoms resolved within 7 days.These symptoms had no correlate with age,gender,implantedear and results of vestibular test.Conclusion:Cochlear implantation can affect the horizontal semicircular canal function,and the video head impulse test and caloric test should be used in a complementary fashion.

Ankyrin-1 Gene Exhibits Allelic Heterogeneity in Conferring Protection Against Malaria.

Allelic heterogeneity is a common phenomenon where a gene exhibits a different phenotype depending on the nature of its genetic mutations. In the context of genes affecting malaria susceptibility, it allowed us to explore and understand the intricate host-parasite interactions during malaria infections. In this study, we described a gene encoding erythrocytic ankyrin-1 (Ank-1) which exhibits allelic-dependent heterogeneous phenotypes during malaria infections. We conducted an ENU mutagenesis screen on mice and identified two Ank-1 mutations, one resulting in an amino acid substitution (MRI95845), and the other a truncated Ank-1 protein (MRI96570). Both mutations caused hereditary spherocytosis-like phenotypes and confer differing protection against Plasmodium chabaudi infections. Upon further examination, the Ank-1(MRI96570) mutation was found to inhibit intraerythrocytic parasite maturation, whereas Ank-1(MRI95845) caused increased bystander erythrocyte clearance during infection. This is the first description of allelic heterogeneity in ankyrin-1 from the direct comparison between two Ank-1 mutations. Despite the lack of direct evidence from population studies, this data further supported the protective roles of ankyrin-1 mutations in conferring malaria protection. This study also emphasized the importance of such phenomena in achieving a better understanding of host-parasite interactions, which could be the basis of future studies.

Erythrocyte ß spectrin can be genetically targeted to protect mice from malaria.

The malaria parasite hijacks host erythrocytes to shield itself from the immune system and proliferate. Red blood cell abnormalities can provide protection from malaria by impeding parasite invasion and growth within the cell or by compromising the ability of parasites to avoid host clearance. Here, we describe 2 N-ethyl-N-nitrosourea-induced mouse lines, SptbMRI26194 and SptbMRI53426 , containing single-point mutations in the erythrocyte membrane skeleton gene, ß spectrin (Sptb), which exhibit microcytosis but retain a relatively normal ratio of erythrocyte surface area to volume and are highly resistant to rodent malaria. We propose the major factor responsible for malaria protection is the specific clearance of mutant erythrocytes, although an enhanced clearance of uninfected mutant erythrocytes was also observed (ie, the bystander effect). Using an in vivo erythrocyte tracking assay, we established that this phenomenon occurs irrespective of host environment, precluding the involvement of nonerythrocytic cells in the resistance mechanism. Furthermore, we recapitulated this phenotype by disrupting the interaction between ankyrin-1 and ß spectrin in vivo using CRISPR/Cas9 genome editing technology, thereby genetically validating a potential antimalarial target. This study sheds new light on the role of ß spectrin during Plasmodium infection and highlights how changes in the erythrocyte cytoskeleton can substantially influence malaria susceptibility with minimal adverse consequences for the host.

A novel ENU-induced ankyrin-1 mutation impairs parasite invasion and increases erythrocyte clearance during malaria infection in mice.

Genetic defects in various red blood cell (RBC) cytoskeletal proteins have been long associated with changes in susceptibility towards malaria infection. In particular, while ankyrin (Ank-1) mutations account for approximately 50% of hereditary spherocytosis (HS) cases, an association with malaria is not well-established, and conflicting evidence has been reported. We describe a novel N-ethyl-N-nitrosourea (ENU)-induced ankyrin mutation MRI61689 that gives rise to two different ankyrin transcripts: one with an introduced splice acceptor site resulting a frameshift, the other with a skipped exon. Ank-1(MRI61689/+) mice exhibit an HS-like phenotype including reduction in mean corpuscular volume (MCV), increased osmotic fragility and reduced RBC deformability. They were also found to be resistant to rodent malaria Plasmodium chabaudi infection. Parasites in Ank-1(MRI61689/+) erythrocytes grew normally, but red cells showed resistance to merozoite invasion. Uninfected Ank-1(MRI61689/+) erythrocytes were also more likely to be cleared from circulation during infection; the "bystander effect". This increased clearance is a novel resistance mechanism which was not observed in previous ankyrin mouse models. We propose that this bystander effect is due to reduced deformability of Ank-1(MRI61689/+) erythrocytes. This paper highlights the complex roles ankyrin plays in mediating malaria resistance.

Treatment of erythrocytes with the 2-cys peroxiredoxin inhibitor, Conoidin A, prevents the growth of Plasmodium falciparum and enhances parasite sensitivity to chloroquine.

The human erythrocyte contains an abundance of the thiol-dependant peroxidase Peroxiredoxin-2 (Prx2), which protects the cell from the pro-oxidant environment it encounters during its 120 days of life in the blood stream. In malarial infections, the Plasmodium parasite invades red cells and imports Prx2 during intraerythrocytic development, presumably to supplement in its own degradation of peroxides generated during cell metabolism, especially hemoglobin (Hb) digestion. Here we demonstrate that an irreversible Prx2 inhibitor, Conoidin A (2,3-bis(bromomethyl)-1,4-dioxide-quinoxaline; BBMQ), has potent cytocidal activity against cultured P. falciparum. Parasite growth was also inhibited in red cells that were treated with BBMQ and then washed prior to parasite infection. These cells remained susceptible to merozoite invasion, but failed to support normal intraerythrocytic development. In addition the potency of chloroquine (CQ), an antimalarial drug that prevents the detoxification of Hb-derived heme, was significantly enhanced in the presence of BBMQ. CQ IC50 values decreased an order of magnitude when parasites were either co-incubated with BBMQ, or introduced into BBMQ-pretreated cells; these effects were equivalent for both drug-resistant and drug-sensitive parasite lines. Together these results indicate that treatment of red cells with BBMQ renders them incapable of supporting parasite growth and increases parasite sensitivity to CQ. We also propose that molecules such as BBMQ that target host cell proteins may constitute a novel host-directed therapeutic approach for treating malaria.

[Sorting of side population cells from multiple myeloma cell lines and analysis of their biological characteristics].

This study was aimed to sort the side population (SP) cells from human multiple myeloma cell lines, then detect the biological characteristics of those SP cells. After Hoechst33342 staining, intracellular Hoechst33342 fluorescence staining differences of myeloma cell lines observed by the fluorescence microscopy. The fluorescence-activated cell sorting (FACS) technology was used to isolate SP cells and main population (MP) cells; proliferative capacity in vitro was determined by cell growth curve; the cell colony forming ability was compared by colony forming test. The CD138 expression was detected by flow cytometry. The expression of ABCG2 mRNA was detected by reverse transcription PCR; CCK-8 assay and colony forming test were used to evaluate the effect of bortezomib on the cell proliferation, vitality and colony forming ability of the two populations. The results showed that the myeloma cell lines had a small proportion of SP cells, especially, RPMI 8226 cells accounted for the highest proportion of SP cells (7.10 ± 2.69)%, which have also been confirmed under the fluorescence microscope; the proliferative activity and cell colony forming ability of SP cells were significantly higher than those of MP cells (P < 0.05). The expression levels of CD138 in SP and MP cells were not significantly different (P > 0.05). RT-PCR results showed that SP cells expressed the drug-resistance gene ABCG2, but MP cells hardly express these genes. The inhibition rate of bortezomib on SP cells was significantly lower than that on MP cells (P < 0.05), however, the difference was not significant (P > 0.05) at bortezomib 40 nmol/L. Bortezomib could reduce colony formation in the both two cell populations, but more severe reduction appeared in the MP cells. It is concluded that the myeloma cell line contain a small amount of SP cells with the cancer stem cell characteristics.

Retrospective study of modified SMILE chemotherapy for advanced-stage, relapsed, or refractory extranodal natural killer (NK)/T cell lymphoma, nasal type.

Extranodal natural killer/T cell lymphomas, nasal type (ENKLs), which are a group of non-Hodgkin lymphomas with poor prognoses, are much more common in China than in Western countries. Here, we retrospectively assessed the impact of two treatment regimens on clinical response and survival among 42 ENKL patients. All patients were diagnosed with stage IV, relapsed, or refractory ENKL. Twenty patients received modified SMILE (consisting of L-asparaginase, methotrexate, ifosphamide, etoposide, and dexamethasone) chemotherapy, and 22 control patients received CHOP (consisting of cyclophosphamide, doxorubicin, vincristine, and prednisone) treatment. Higher complete response (CR) and overall response rates (ORR) (CR 45.0 vs. 13%, ORR 70 vs. 36%) were observed among the patients treated with the modified SMILE regimen (Fisher's exact = 0.040, Pearson χ(2) P = 0.030). Similarly, a higher ORR rate was observed among Epstein-Barr virus-positive patients (ORR 50.0 vs. 18.0%, Fisher's exact = 0.049). The treatment group was also significantly associated with longer overall survival (OS) and progression-free survival (PFS) (Log-rank, P = 0.0341, P = 0.0142, respectively), but OS did not seem to be longer. Treatment-related toxicity was monitored in all patients throughout the protocol. There were no significant differences in the incidence of hematological and non-hematological toxicities between the two groups (P < 0.05), with the exception of peripheral neuropathy (treatment = 0 control = 5, Fisher's exact = 0.049).

[Effects of down-regulated TRAF6 gene expression on the proliferation and apoptosis in multiple myeloma cells].

OBJECTIVE: To investigate the down-regulated TRAF6 gene expression and its effects on proliferation and apoptosis in multiple myeloma (MM) cells. METHODS: Detection of TRAF6 expression were conducted by RT-PCR and Western blot in MM cell lines of KM3, U266, RPMI8226 and primary cells from patients. RPMI8226 cell lines were transfected with siRNA of TRAF6. The efficiency of transfection was identified by using of fluorescence microscope, RT-PCR, and Western blot. The levels of proliferation were analyzed by CCK-8 method under the different concentrations of siRNA. Apoptosis rate were detected with Hoechst33258/PI double staining by flow cytometry. Apoptosis related proteins Bcl-2, BAX, and NF-κB signal pathway were observed before and after siRNA transfection by Western blot. RESULTS: The levels of TRAF6 mRNA and protein in MM cell lines, especially in primary myeloma cells, were significantly higher than those in controls. After transfected with 50 nmol/L siRNA in RPMI8226 cells, the relative level of TRAF6 mRNA (0.49±0.24) was significantly lower than that in non-transfected group (1.87±0.23) and idling group (1.74±0.35). The proliferation rate of siRNA transfected cells decreased with dose dependence (P<0.01). The apoptosis rates increased from 11.20% (before transfection) to 51.82% (after transfection), accompanied by down-regulated Bcl-2 protein, NF-κB signal pathway (p-p65 and p52), and up-regulated BAX protein. CONCLUSION: TRAF6 expression was high in myeloma cells. TRAF6 siRNA could inhibit proliferation of myeloma cells and induce apoptosis mediated by NF-κB classical and alternative pathway in myeloma cells.

Platelet factor 4 and Duffy antigen required for platelet killing of Plasmodium falciparum.

Platelets restrict the growth of intraerythrocytic malaria parasites by binding to parasitized cells and killing the parasite within. Here, we show that the platelet molecule platelet factor 4 (PF4 or CXCL4) and the erythrocyte Duffy-antigen receptor (Fy) are necessary for platelet-mediated killing of Plasmodium falciparum parasites. PF4 is released by platelets on contact with parasitized red cells, and the protein directly kills intraerythrocytic parasites. This function for PF4 is critically dependent on Fy, which binds PF4. Genetic disruption of Fy expression inhibits binding of PF4 to parasitized cells and concomitantly prevents parasite killing by both human platelets and recombinant human PF4. The protective function afforded by platelets during a malarial infection may therefore be compromised in Duffy-negative individuals, who do not express Fy.

[The Effects of IFN-γ on AKT activated 32D Cells and its Mechanisms].

OBJECTIVE: To investigate the effects of activated AKT on murine myeloid precursor cells (32D cells), and the effects of IFN-γ on 32D cells and its mechanisms. METHODS: Plasmid transduction was used to enhance the expression of AKT on 32D cells. After the transfected cells treated with IFN-γ for 24 hours, proliferation rate was tested by WST-1, apoptosis by flow cytometry, expression of phosphorylated Erk1/2, Stat3 and phosphorylated Stat3 was determined by Western blot. RESULTS: (1) IFN-γ at low concentration (100 U/ml) enhanced the growth and proliferation of 32D cells, while at high concentration (1000 U/ml) suppressed them. (2) Compared with control groups, low concentration IFN-γ increased (1124 ± 13) Stat3 phosphorylation in 32D-cell, while it high concentration IFN-γ decreased (601 ± 13). 32D cells transfected with activated Akt grew rapidly (0.287 ± 0.010) and had a low apoptotic rate [(9.57 ± 0.17)% (P < 0.05)]. (3) The expression of p-Erk1/2 in transfected 32D-cell was significantly reduced (P < 0.05). (4) Apoptosis rate of IFN-γ treated group was significantly decreased in transfected 32D cells (P < 0.05). CONCLUSIONS: IFN-γ has dual effects on 32D cells, namely, at low concentration enhanced the growth and proliferation of 32D cells, while at high concentration suppressed them. Its mechanisims is possibly through Stat3 pathway. Activated Akt can significantly promote the growth and proliferation of 32D cell and significantly inhibit apoptosis and IFN-γ can regulate cell proliferation and apoptosis through AKT. AKT activation can inhibit the Erk signal pathway, which may be affected by inhibition the modificaton of Raf1.

[Effect of zoledronic acid on the growth and CD138 expression of myeloma cell line KM3].

OBJECTIVE: To study the inhibitory effect of zoledronic acid (ZA) on the growth and CD138 expression of myeloma cell line KM3. METHODS: KM3 cells were treated with different concentrations of ZA The growth of KM3 cells was measured by trypan blue dye exclusion, and the changes of apoptosis rate, cell cycle and expression of CD138 induced by ZA by flow cytometry. RESULTS: Within the concentration of 10(-5)-10(-3) mol/L, ZA obviously inhibited the growth of KM3 cells in a dose dependent manner. IBN at 10(-5)-10(-4) moL/L increased Annexin V positive rate, blocked cells at the S/G2 boundary, reduced the expression of CD138 and its fluorescence intensity. CONCLUSION: ZA can inhibit the growth of KM3 cells in a dose-dependent manner and inhibited CD138 expression. The mechanism is probably related to induction cell cycle accumulation in S phase and apoptosis.

[Antiangiogenic activity of thalidomide in vitro mediated by cytochrome CYP2C19].

This study was aimed to investigate the effect of metabolic system in human hepatic cell microsome on antiangiogenic in vitro activity of thalidomide used in treating multiple myeloma and to explore the role of cytochrome CYP2C19. Human umbilical cord vein endothelial cells (hUCVECs) were treated with thalidomide alone or thalidomide co-incubated with human hepatic cell microsome. Cell proliferation ability was assessed by MTT assay, cell cycle analysis and detection of apoptosis were carried out by flow cytometry (FCM), migration activity of hUCVECs was determined by modified Boyden chamber and differentiation of hUCVECs was assayed by tube formation test. The results showed that thalidomide alone had no obvious direct effect on hUCVEC viability or apoptosis, mild effect on cell migration and no effect on tube formation. However, when co-incubated with human hepatic cell microsome, thalidomide significantly inhibited the hUCVECs viability. At 100 microg/ml, thalidomide co-incubated with human hepatic cell microsome, the proliferation ability of hUCVECs decreased by (11.7 +/- 3.9)%, apoptosis cells increased by 27.2%, the cell migration was down-regulated significantly, and the tube formation was obviously inhibited. When omeprazole, a specific inhibitor of cytochrome CYP2C19, was added into the co-incubation mixture, the effects of thalidomide on cell proliferation ability, apoptosis, migration and tube formation decreased. It is concluded that effect of human hepatic cell microsome is required for thalidomide's antiangiogenic activity in vitro and cytochrome CYP2C19 may be involved in the antiangiogenic effect of thalidomide.

[Effect of cytochrome CYP2C19 on the anti-myeloma activity of thalidomide in vitro].

OBJECTIVE: To investigate the effect of human liver microsome on anti-myeloma activity of thalidomide (TH) in vitro and identify the role of cytochrome CYP2C19 in it. METHODS: Human multiple myeloma (MM) cell lines U266, NCI-H929, RPMI 8226, LP-1 and CZ-1 were treated with TH or TH pre-incubated with human liver microsome. Cell viability was detected by MTT assay, and cell cycle and apoptosis by flow cytometry (FCM). RESULTS: TH treatment had no direct effect on cell viability at concentrations of 10 microg/ml, 50 microg/ml and 100 microg/ml, the viabilities of the 5 MM cell lines were 96.2% - 103.7%, 96.3% - 103.7% and 97.9% - 106.5% respectively, being no significant difference from that of control (P > 0.05). However, when preincubated with human liver microsome, TH significantly inhibited the cell viability with a dose-dependent manner. At concentrations of 10 microg/ml, 50 microg/ml and 100 microg/ml, TH pre-incubated with human liver microsome led to 12.2% - 22.9%, 25.9% - 36.4% and 34.9% - 46.3% decreases of cell viability, respectively (P < 0.05). TH at concentration of 100 microg/ml pre-incubated with human liver microsome caused an increase of 18.5% - 32.5% in apoptosis cells. When omeprazole, a specific inhibitor of cytochrome CYP2C19, was added in the incubation system, the inhibition of cell viability by TH was weakened. At concentrations of 5 micromol/L and 10 micromol/L, omeprazole reversed the cell viability by 7.5% - 21.9% and 19.1% - 38.3%, respectively (P < 0.05). CONCLUSION: Treatment of TH with human liver microsome is essential for its anti-myeloma activity in vitro, and cytochrome CYP2C19 is involved with this metabolism process.

[Study on negative regulatory factors in bone marrow mononuclear cells of myelodysplastic syndromes].

To explore the difference of negative regulatory factors among T lymphocyte subsets in bone marrow (BM) of myelodysplastic syndromes (MDS) and their relations to apoptotic gene Fas, different lymphocyte subsets in BM were categorized by monoclonal antibodies with 3 color fluorescence using flow cytometry, and the intracellular cytokines such as tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma) were determined following marrow cells culture. Then Fas mRNA of bone marrow mononuclear cells (BMMNC) were examined by RT-PCR. The results showed that TNF-alpha, IFN-gamma levels in BM of MDS both increased, the former produced by cells CD4+CD45RO+, CD8+CD45RO+, the latter by cells CD4+CD45RO+, CD8+CD45RO+, CD8+CD45RA+, in which the cells CD8+CD45RO+ were dominant. Fas mRNA expression had relationship with IFN-gamma produced by T cells but not with TNF-alpha. It is concluded that in hematopoietic microenvironment of MDS, not only the T lymphocyte subsets are in disorder, but also negative regulatory factors secreted by T lymphocyte increase. T lymphocytes play an important role in producing IFN-gamma in patients with MDS.