Multimodal evaluation of the effects of low-intensity ultrasound on cerebral blood flow after traumatic brain injury in mice.

Traumatic brain injury (TBI) is one of the leading causes of death and disability worldwide, and destruction of the cerebrovascular system is a major factor in the cascade of secondary injuries caused by TBI. Laser speckle imaging (LSCI)has high sensitivity in detecting cerebral blood flow. LSCI can visually show that transcranial focused ultrasound stimulation (tFUS) treatment stimulates angiogenesis and increases blood flow. To study the effect of tFUS on promoting angiogenesis in Controlled Cortical impact (CCI) model. tFUS was administered daily for 10 min and for 14 consecutive days after TBI. Cerebral blood flow was measured by LSCI at 1, 3, 7 and 14 days after trauma. Functional outcomes were assessed using LSCI and neurological severity score (NSS). After the last test, Nissl staining and vascular endothelial growth factor (VEGF) were used to assess neuropathology. TBI can cause the destruction of cerebrovascular system. Blood flow was significantly increased in TBI treated with tFUS. LSCI, behavioral and histological findings suggest that tFUS treatment can promote angiogenesis after TBI.

Discovery of a NADPH oxidase inhibitor, (E)-3-cyclohexyl-5-(4-((2-hydroxyethyl)(methyl)amino)benzylidene)-1-methyl-2-thioxoimidazolidin-4-oneone, as a novel therapeutic for Parkinson's disease.

Several lines of evidence indicated that generation of NADPH oxidase (Nox)-mediated reactive oxygen species are associated with neuronal inflammation, leading to Parkinson's disease (PD). Novel benzylidene-1-methyl-2-thioxoimidazolidin-one derivatives as Nox inhibitors were designed and synthesized in order to increase blood-brain barrier (BBB) permeability to target Nox in brain cells. In lucigenin chemiluminescence assay, eight compounds showed excellent inhibition activity against NADPH oxidases and parallel artificial membrane permeability assay (PAMPA) identified compound 11 with high passive permeability. To validate the effect of compound 11 on neuronal inflammation, we tested the regulatory activity of compound 11 in lipopolysaccharide (LPS)-induced production of pro-inflammatory cytokines in BV-2 microglial cells and LPS-mediated microglial migration. Treatment of BV2 cells with compound 11 resulted in suppressed production of pro-inflammatory cytokines and migration activity of BV2 cells in response to LPS. To evaluate the therapeutic efficacy of compound 11 in PD animal model, compound 11 was applied to MPTP-induced PD mouse model. Oral administration of compound 11 (30 mg/kg/daily, 4 weeks) into the mice resulted in suppression of dopaminergic neuronal death in substantia nigra (SN) and in striatum as well as inhibition of microglial migration into SN. These results implicate compound 11 as a novel therapeutic agent for the treatment of PD.

Circular RNA circ_0004277 Inhibits Acute Myeloid Leukemia Progression Through MicroRNA-134-5p / Single stranded DNA binding protein 2.

Circular RNAs (circRNAs) are crucial non-coding RNAs in the process of tumorigenesis. Nevertheless, the biological function of circ_0004277 in acute myeloid leukemia (AML) is blurred. Microarray data of circRNAs were utilized to evaluate circRNAs' differential expression in AML. Quantitative real-time polymerase chain reaction (qRT-PCR) was executed to determine circ_0004277 and microRNA-134-5p (miR-134-5p) expression levels. The growth, migration and invasion of AML cells were tested by the cell counting kit-8 and Transwell experiment. Dual-luciferase reporter gene experiment, RNA immunoprecipitation (RIP) experiment and RNA pull-down experiment were executed to determine the targeting relationship between circ_0004277 and miR-134-5p. Western blot assay was used to detect single stranded DNA binding protein 2 (SSBP2) expression. We observed that circ_0004277 was down-regulated in AML, while miR-134-5p was up-regulated. Functionally, circ_0004277 overexpression or inhibition of miR-134-5p remarkably suppressed AML cell viability, migration and invasion. Furthermore, miR-134-5p served as a direct downstream target of circ_0004277 and SSBP2 was identified as a target of miR-134-5p. Compensation experiments showed that miR-134-5p mimics abolished the biological function of circ_0004277 on malignant phenotypes of AML cells. Collectively, circ_0004277 impedes AML development by adsorbing miR-134-5p and up-regulating SSBP2.

Myeloid ecotropic viral integration site-1 inhibition promotes apoptosis, suppresses proliferation of acute myeloid leukemia cells, accentuates the effects of anticancer drugs.

To investigate the effects of myeloid ecotropic viral integration site-1 (MEIS1) on the proliferation and apoptosis of acute myeloid leukemia (AML) cells and the anticancer effects of the drug, we screened Kasumi-6, KG-1, and Kasumi-1 cells using quantitative reverse transcription polymerase chain reaction. Kasumi-6 and Kasumi-1 cells were subjected to human antigen R (HuR)-mediated interference (IV). Hexokinase 2 (HK2) expression and phosphorylation of protein kinase B (p-AKT) and mammalian target of rapamycin (p-mTOR) were observed with Western blotting. Cell proliferation was assessed using Cell Counting Kit-8, apoptosis was examined using Hoechst 33,258 staining, and glucose uptake was detected with a colorimetric biochemical assay kit. We found that, among the three cell lines tested, MEIS1 expression was highest in Kasumi-1 cells, which were therefore selected for subsequent experiments. Kasumi-1 cells receiving IV showed significantly decreased proliferation (p < 0.05) and increased apoptosis compared to the control group. Compared with the controls, IV significantly increased the expression of HK2, p-AKT, p-mTOR, multidrug resistance-associated protein 1 and P-glycoprotein (P < 0.05), but decreased glucose uptake. Treatment with adriamycin, daunorubicin and imatinib resulted in a progressive increase in inhibition of cell proliferation, with the IV group showing the highest inhibition rate among the three groups (P < 0.05). Thus, inhibition of MEIS1 activity promoted apoptosis, inhibited the proliferation of Kasumi-1 and Kasumi-6 cells, and increaseed the anticancer effect of the drugs, suggesting that inhibition of MEIS1 may be a potential strategy for the treatment of AML.

Mechanism of exosomal miR-155 derived from bone marrow mesenchymal stem cells on stemness maintenance and drug resistance in myeloma cells.

OBJECTIVE: This study was to explore the effect of exosomal miR-155 derived from bone marrow mesenchymal stem cells (BMSCs) on stemness maintenance and drug resistance in MPC-11 multiple myeloma cells. METHODS: MPC-11 cells were transfected with mimics or inhibitors of miR-155. miR-155 expression was detected by qRT-PCR, cell condition was observed, and the expression of stemness maintenance markers OCT-4 and Nanog was observed by immunofluorescence. The expression of proteins associated with the Hedgehog signaling pathway and drug resistance was evaluated by western blot. To investigate whether exosomes affect cell behavior by horizontal delivery of miR-155, MPC-11 cells were co-cultured with exosomes isolated from BMSCs that were transfected with mimics or inhibitors of miR-155. Cell proliferation and the expression of proteins related to stemness maintenance protein and drug resistance were examined. RESULTS: In function assays, after miR-155-mimics transfection, the expression levels of proteins related to stemness maintenance marker, Hedgehog signaling, and drug resistance were increased in MPC-11 cells. BMSC-derived exosomes carrying miR-155 inhibited apoptosis, promoted cell division, and upregulated the expression of protein associated with stemness maintenance, Hedgehog signaling, and drug resistance. CONCLUSION: Therefore, our findings indicate that exosomal delivery of miR-155 exerted the same effect as transfection did on the stemness maintenance and drug resistance of multiple myeloma cells.

Effect of Low Intensity Transcranial Ultrasound (LITUS) on Post-traumatic Brain Edema in Rats: Evaluation by Isotropic 3-Dimensional T2 and Multi-TE T2 Weighted MRI.

Background: Brain edema is one of the important factors affecting the prognosis of traumatic brain injury (TBI). Low-intensity transcranial ultrasound (LITUS) has significant anti-cerebral edema effect. T2-weighted image-based volume and T2 value measurements can sensitively reflect tissue edema. Purpose: To evaluate the effect and possible mechanisms of LITUS on brain edema by iso-voxel 3-dimensional T2WI (iso-3D T2WI) and multi-TE T2WI. Methods: Forty-five rats were randomly divided into sham control, TBI and TBI + LITUS groups (n = 15, respectively). Iso-voxel 3-dimensional T2WI and multi-TE T2WI sequences at 3.0T to obtain T2 value and edema volume of the injury cortex. T2 values were obtained on days 1, 7, 14, 21, 28, 35, and 42 after TBI and brain edema volume were obtained on days 7 and 14. Results: The T2 values of the damaged cortex in the TBI group showed a slow decreasing trend after a significant increase. For TBI+LITUS group, T2 values decreased with continuous LITUS treatment. At day 28, the T2 values were not significantly longer than the control group (adjusted P = 0.0535), but were significantly shorter than the TBI group at day 42 (adjusted P = 0.0003). The edema volume at day 7 and 14 in the LITUS group was significantly lower than the TBI group (P = 0.0004 and P < 0.0001, respectively). AQP-4 and ß-APP protein staining showed a strong positive reaction near the CCI point, TBI+LITUS group showed a medium positive reaction, and the sham control group showed a weak positive reaction. Conclusion: The therapeutic effect of LITUS on post-traumatic brain edema was confirmed through T2 value and edema volume, and the mechanism may be related to inhibiting the expression of AQP-4 and promoting the removal of ß-APP.

Neuroprotective Effect of Low-Intensity Transcranial Ultrasound Stimulation in Moderate Traumatic Brain Injury Rats.

Traumatic brain injury (TBI) is a kind of severe brain injury characterized with a high incidence rate and a high disability rate. Low-intensity transcranial ultrasound stimulation (LITUS) is a promising neuroprotective method for improving the functional prognosis of TBI. The fractional anisotropy (FA) value and mean diffusivity (MD) value can be sensitive to abnormal brain structure and function and can thus be used to evaluate the effect of LITUS on TBI. Our purpose was to evaluate the therapeutic effect of LITUS in a moderate TBI rat model with FA and MD values. For our method, we used 45 male Sprague Dawley rats (15 sham normal, 15 TBI, and 15 LITUS treatment rats). We used single-shot spin echo echo-planar imaging sequences at 3.0T to obtain the DTI parameters. Parameters of FA and MD on the treated side of the injury cortex were measured to evaluate the therapeutic effect of LITUS in a TBI rat model. For FA and MD values, groups were compared by using a two-way analysis of variance for repeated measures, and this was followed by Tukey's post hoc test. Differences were considered significant at P < 0.05. The results were that the FA value in the LITUS treatment group at 1 day after TBI was significantly higher than that in the control group (adjusted P = 0.0422) and significantly lower than that in the TBI group at 14, 21, and 35 days after TBI (adjusted P = 0.0015, 0.0064, and 0.0173, respectively). At the end of the scan time point, the differences between the two groups were not significant (adjusted P = 0.3242). The MD values in the LITUS treatment group were significantly higher in the early stage than that in the TBI group (adjusted P = 0.0167) and significantly lower at the following time points than in the TBI group. In conclusion, daily treatment with LITUS for 10 min effectively improved the brain damage in the Controlled Cortical Impact (CCI)-caused TBI model. FA and MD values can serve as evaluation indicators for the neuro-protective effect of LITUS.

Evaluating the Therapeutic Effect of Low-Intensity Transcranial Ultrasound on Traumatic Brain Injury With Diffusion Kurtosis Imaging.

BACKGROUND: Low-intensity transcranial ultrasound (LITUS) has a therapeutic effect on traumatic brain injury (TBI). Diffusion kurtosis imaging (DKI) might be able to evaluate the effect changes of injured brain microstructure. PURPOSE: To evaluate the therapeutic effect of LITUS in a moderate TBI rat model with DKI parameters. STUDY TYPE: Prospective case-control animal study. ANIMAL MODEL: Forty-five rats were randomly divided into sham control, TBI, and LITUS treatment groups (n = 15). FIELD STRENGTH/SEQUENCE: Single-shot spin echo echo-planar imaging and fast T2 WI sequences at 3.0T. ASSESSMENT: DKI parameters were obtained on days 1, 7, 14, 21, 28, 35, and 42 after TBI. STATISTICAL TESTS: For the mean kurtosis (MK), axial kurtosis (Ka), and radial kurtosis (Kr) values, groups were compared using a two-way analysis of variance (ANOVA). RESULTS: LITUS inhibited TBI and caused MK values to increase significantly during the early stage (LITUS vs. TBI, day 7, adjusted P < 0.0001) and decrease during the late stage (LITUS vs. TBI, day 42, adjusted P = 0.0156) in the damaged cortex. In the thalamus, the MK value of the TBI group began to rise on day 7, with no change observed in the LITUS group. TBI increases Ka value during the early stage in the cortex and decreases during the late stage in the cortex and thalamus. LITUS inhibited these Ka changes (LITUS vs. TBI, day 7, adjusted P = 0.0014; LITUS vs. TBI, day 42, adjusted P = 0.0026 and 0.0478, respectively, for cortex and thalamus). The Kr value increased slightly during the early stage in the cortex (TBI vs. Sham, day 1, adjusted P = 0.0016). DATA CONCLUSION: The DKI parameter, particularly the MK value, evaluates primary cortical injury as well as the secondary brain injury that could not be detected by conventional T2 WI. LEVEL OF EVIDENCE: 1 Technical Efficacy Stage: 4 J. Magn. Reson. Imaging 2020;52:520-531.

[Expression and Clinical Significance of Leukemia Stem Cell Antigen in Acute Myeloid Leukemia].

OBJECTIVE: To analyze the expression characteristics of leukemia stem cell (LSC) antigen in acute myeloid leukemia (AML) and to explore the correation of LSC-specific antigens with the subtypes, cytogenetics and clinical efficacy of AML. METHODS: A total of 61 newly diagnosed patients with AML (except M3) hospltalized in Department of Hematology of our hopital were selected from January 2013 to March 2016. The immun phenotypes and expression of Tim-3, CD96 and CD123 on leucamia cells were detected by direct immunofluorescenct flow cytometry. 61 patients were divided into positive expression and megative expression groups according to expression of Tim-3, CD96 and CD123; the correlation of LSC antigen expression level with high WBC count, chromosome and therapeutic efficacy was analyzed. RESULTS: Among 61 newly diagnosed patients with AML (except M3), the expression rate of Tim-3, CD96 and CD123 was 52.45%, 44.26% and 55.73% respectively. The expression rates of Tim-3, CD96 and CD123 between the AML subtypes and total patients was not stetistically different (P>0.05). The high WBC count occurred more easily in AML (except MS) patients with positive expression of Tim-3, CD96 and CD123, but compared with AML patients with negative espression, the difference was not statstically significant (P>0.05). The proportion of chromosone karyotype with poor prognosis detected in patients with positive expression of Tim-3 and CD96 was higher than that in patients with negative expreesion (P<0.05); while the preoprtion of chromosome karyotype with poor prognosis detected in patients with positive and negative expression of CD123 was not significantly different (P>0.05). After 2 courses of chemotherapy, the complete remission (CR) rate in patients with positive expression of Tim-3, CD96 and CD123 was significantly lower than that in patients with negative expression of Tim-3, CD96 and CD123 (P<0.05), the comparison of OS time in patients with positive and negative expression of Tim-3 and CD96 showed the statistical difference (P>0.05), while the difference of OS time in patients with positive and negative expression of CD123 was not significant (P>0.05). CONCLUSION: The expression levels of Tim-3, CD96 and CD123 in newly diagnosed AML (except M3) sybtype patients are not significantly different form those in total patients. The high WBC count ocours more easily in patients with positive expression of Tim-3, CD96 and CD123. After 2 course of chemotherapy, the CR rate in patients with positive expression of Tim-3, CD96 and CD123 was significantly lower than that in patients with negative expression. The proportion of chromsome karyotype with poor prognosis detected in patients with positive expression of Tim-3 and CD96 is high, moreover, OS time in patients with positive expression of Tim-3 and CD96 is shorter than that in patients with negative expression.

Dapsone protects brain microvascular integrity from high-fat diet induced LDL oxidation.

Atherosclerosis was considered to induce many vascular-related complications, such as acute myocardial infarction and stroke. Abnormal lipid metabolism and its peroxidation inducing blood-brain barrier (BBB) leakage were associated with the pre-clinical stage of stroke. Dapsone (DDS), an anti-inflammation and anti-oxidation drug, has been found to have protective effects on vascular. However, whether DDS has a protective role on brain microvessels during lipid oxidation had yet to be elucidated. We investigated brain microvascular integrity in a high-fat diet (HFD) mouse model. We designed this study to explore whether DDS had protective effects on brain microvessels under lipid oxidation and tried to explain the underlying mechanism. In our live optical study, we found that DDS significantly attenuated brain microvascular leakage through reducing serum oxidized low-density lipoprotein (oxLDL) in HFD mice (p < 0.001), and DDS significantly inhibited LDL oxidation in vitro (p < 0.001). Our study showed that DDS protected tight junction proteins: ZO-1 (p < 0.001), occludin (p < 0.01), claudin-5 (p < 0.05) of microvascular endothelial cells in vivo and in vitro. DDS reversed LAMP1 aggregation in cytoplasm, and decreased the destruction of tight junction protein: ZO-1 in vitro. We first revealed that DDS had a protective role on cerebral microvessels through preventing tight junction ZO-1 from abnormal degradation by autophagy and reducing lysosome accumulation. Our findings suggested the significance of DDS in protecting brain microvessels under lipid metabolic disorders, which revealed a novel potential therapeutic strategy in brain microvascular-related diseases.

Suppression of HSP27 increases the anti­tumor effects of quercetin in human leukemia U937 cells.

Quercetin, a natural flavonoid, inhibits the growth of leukemia cells and induces apoptosis. Heat shock protein 27 (HSP27) has been reported to promote the development of leukemia by protecting tumor cells from apoptosis through various mechanisms. The present study investigated the effects of small hairpin (sh)RNA-mediated HSP27 knockdown on the anti­cancer effects of quercetin in U937 human leukemia cells. Cells were transfected with recombinant lentiviral vector pCMV­G­NR­U6­shHSP27 (shHSP27), which expressed shRNA specifically targeting the HSP27 gene, alone or in combination with quercetin. The results showed that shHSP27 and quercetin synergistically inhibited U937 cell proliferation and induced apoptosis by decreasing the Bcl2-to-Bax ratio. Furthermore, this combined treatment significantly suppressed the infiltration of tumor cells and the expression of angiogenesis­associated proteins HIF1α and VEGF. Compared with shHSP27 or quercetin alone, shHSP27 plus quercetin markedly decreased the protein expression of cyclinD1 and thus blocked the cell cycle at G1 phase. The Notch/AKT/mTOR signaling pathway is important in tumor aggressiveness; quercetin plus shHSP27 significantly decreased Notch 1 expression and the phosphorylation levels of the downstream signaling proteins AKT and mTOR. The inhibitory effects of quercetin plus shHSP27 on this pathway may thus have been responsible for the cell cycle arrest, inhibition of proliferations and infiltration as well as enhancement of apoptosis. Therefore, these findings collectively suggested that suppression of HSP27 expression amplified the anti­cancer effects of quercetin in U937 human leukemia cells, and that quercetin in combination with shHSP27 represents a promising therapeutic strategy for human leukemia.

Autophagy is involved in oral rAAV/Aß vaccine-induced Aß clearance in APP/PS1 transgenic mice.

The imbalance between ß-amyloid (Aß) generation and clearance plays a fundamental role in the pathogenesis of Alzheimer's disease (AD). The sporadic form of AD is characterized by an overall impairment in Aß clearance. Immunotherapy targeting Aß clearance is believed to be a promising approach and is under active clinical investigation. Autophagy is a conserved pathway for degrading abnormal protein aggregates and is crucial for Aß clearance. We previously reported that oral vaccination with a recombinant AAV/Aß vaccine increased the clearance of Aß from the brain and improved cognitive ability in AD animal models, while the underlying mechanisms were not well understood. In this study, we first demonstrated that oral vaccination with rAAV/Aß decreased the p62 level and up-regulated the LC3B-II/LC3B-I ratio in APP/PS1 mouse brain, suggesting enhanced autophagy. Further, inhibition of the Akt/mTOR pathway may account for autophagy enhancement. We also found increased anti-Aß antibodies in the sera of APP/PS1 mice with oral vaccination, accompanied by elevation of complement factors C1q and C3 levels in the brain. Our results indicate that autophagy is closely involved in oral vaccination-induced Aß clearance, and modulating the autophagy pathway may be an important strategy for AD prevention and intervention.

Phospholipid transfer protein (PLTP) deficiency accelerates memory dysfunction through altering amyloid precursor protein (APP) processing in a mouse model of Alzheimer's disease.

Phospholipid transfer protein (PLTP) is a widely expressed lipid transfer protein participating in the transport of cholesterol and other lipids in the plasma and peripheral tissues. Recently, elevated amyloid ß (Aß) in young and aged PLTP-deficient brains had been reported. However, the role of PLTP in amyloid precursor protein (APP) processing and Alzheimer's disease (AD) pathology remains elusive. Here we first found that deficiency of PLTP accelerated memory dysfunction in APP/PS1ΔE9 AD model mice at the age of 3 months. Further characterization showed that PLTP deficiency increased soluble Aß peptides, and intracellular accumulation of Aß was illustrated, which might be due to disrupted APP turnover and the enhanced amyloidogenic pathway. Besides, reduced brain-derived neurotrophic factor (BDNF) was found in PLTP-deficient APP/PS1ΔE9 mice, and the BDNF level was negatively correlated with Aß42 content, instead of Aß40 content. In addition, autophagic dysfunction was found in the PLTP-deficient APP/PS1ΔE9 mice. Our data presented a novel model to link phospholipid metabolism to APP processing and also suggested that PLTP played an important role in Aß metabolism and would be useful to further elucidate functions of PLTP in AD susceptibility.

Knockdown of WISP1 inhibit proliferation and induce apoptosis in ALL Jurkat cells.

WISP1, a Wnt-induced secreted protein, has been found to have anticancer activity. ALL is a leading cause of death. Here we investigate the WISP1 effects on ALL Jurkat cells. Cell viability was assessed by CCK-8. Cell cycle and apoptosis were detected by flow cytometry. Mitochondrial membrane potential (MMP) was monitored using TMRM. Generation of reactive oxygen species (ROS) was quantified using DCFH-DA. Western blot was used to detect the expression of cell proliferation and apoptosis related genes. The results showed that knockdown of WISP1 significantly inhibited proliferation of Jurkat cells. Parallelly, cell cycle distribution was increased at G1 phase and apoptotic rate was induced after WISP1 knockdown. Furthermore, knockdown of WISP1 induced apoptosis of Jurkat cells was also associated with loss of MMP and generation of ROS. Western blot results showed that the protein expression p-AKT, PCNA, CDK1, P-ERK, CDK2, VEGF, VEGFR2 and Bcl2 were decreased, while the expression of Bax was up-regulated. In conclusion, WISP1 plays an important role in proliferation and apoptosis of Jurkat cells in mitochondria dependent pathway, the specific mechanisms need further study.

Surgical stress induced depressive and anxiety like behavior are improved by dapsone via modulating NADPH oxidase level.

Surgical stress induced depression and anxiety like behavior are common complications among aged individuals suffering from surgery. Recent studies proposed that accumulation of oxidative stress is involved in the etiology of stress induced depression and anxiety. Dapsone possesses antioxidant properties, however, whether dapsone is effective in modulating surgical stress induced brain oxidative damage remains uncertain. The present study aimed to investigate the effect of dapsone on surgical stress induced depressive and anxiety like behavior, and brain oxidative stress in a well-established surgical stress model. Depressive and anxiety like behavior accompanied by elevated brain oxidative stress were observed in aged mice underwent abdominal surgery. Pretreatment with 5 mg/kg dapsone significantly improved the behavioral disorder and ameliorated brain oxidative stress in this model. Further investigation, revealed that surgical stress increased brain NADPH oxidase level, while pretreatment with dapsone abrogated the elevation of NADPH oxidase triggered by surgical stress. These findings suggest that dapsone is effective in improving surgical stress induced brain oxidative damage via down-regulating NADPH oxidase level in aged mice.

Heparin-induced thrombocytopenia in Chinese patients: 240 patients study in one center.

BACKGROUND: The study was conducted in order to analyze the incidence of heparin-induced thrombocytopenia (HIT) and heparin-induced thrombocytopenia and thrombosis syndrome (HITTS) in 240 patients from the Department of Vascular Surgery in China, to evaluate the current laboratory detection technique, and to explore the feasibility for the technique to be developed in China. METHODS: 240 patients receiving unfractionated heparin (UFH) were studied in one center. Before and after the UFH treatment, platelet count, HIT-antibody ELISA test, and heparin-induced platelet aggregation (HIPA) were tested. RESULTS: Among 240 patients, HIT was diagnosed in 36 cases. The incidence was 15%. HITTS occurred in 6 cases (2.5%). The median time of thrombocytopenia was the 8th day. Platelet counts recovered to normal level in 3 - 6 days after heparin withdrawal CONCLUSIONS: The incidence of HIT is higher; however, the incidence of HITTS is lower. Monitoring platelet count, which is simple and easy to do and available in hospitals at different levels (township hospital included), is a reliable method to diagnose HIT early. The HIT-antibody ELISA test could be introduced in specific hospitals to prescreen HIT. In view of the higher sensitivity and specificity of HIPA, it can be used in larger diagnostic centers in all the big cities of China.

Chemotherapy induces enhanced procoagulant activity through phosphatidylserine exposure in acute lymphoblastic leukemia.

INTRODUCTION: Thromboembolism is a serious complication in patients with acute lymphoblastic leukemia (ALL). Coagulation disorders can be induced and worsened by cytotoxic drugs; however, the mechanisms are largely unknown. Our study aims to investigate the effects of daunorubicin (DNR) and L-asparaginase (L-ASP) on phosphatidylserine (PS) exposure and the procoagulant activity (PCA) of Jurkat/ALL cells. The anticoagulant properties of lactadherin were also explored. MATERIALS AND METHODS: Jurkat cells and cells from 10 newly diagnosed patients with ALL were treated with DNR or L-ASP. Flow cytometry and confocal microscopy were used to quantify and locate PS exposure, respectively. PCA was evaluated using coagulation assays and purified coagulation complex assays. Lactadherin, a glycoprotein of the milk fat globule membrane with stereospecific binding to phosphatidyl-L-serine, was used as a probe for the detection of exposed PS. RESULTS: Untreated Jurkat/ALL cells exhibited higher PS exposure and greater PCA than mononuclear cells (MNCs). The PCA of cells treated with DNR or L-ASP was markedly increased. Flow cytometry and confocal microscopy indicated that the increased PCA occurred in parallel with PS exposure. The blocking of PS with lactadherin prolonged the coagulation time and inhibited approximately 85-90% of the activities of procoagulant enzyme complexes in Jurkat/ALL cells. CONCLUSIONS: Our results indicate that DNR and L-ASP increased the PCA of Jurkat/ALL cells through PS exposure and played a critical role in inducing thrombosis in ALL patients. Lactadherin is an ideal probe for PS detection at an early stage and a potential anticoagulant to improve the hypercoagulability of ALL patients.

HIF-1α inhibitors: synthesis and biological evaluation of novel moracin O and P analogues.

The natural products moracins O and P exhibited potent in vitro inhibitory activity against hypoxia-inducible factor (HIF-1), which is a key mediator during adaptation of cancer cells to tumour hypoxia. Systematic variations of the structures of benzofuran type moracins were made and structure-activity relationship analysis showed the importance of the 2-arylbenzofuran ring and the (R)-configuration of the core scaffold. Further evaluation of the representative compound 5 showed its inhibitory effect on HIF-1α protein accumulation and target gene expression under hypoxia.

LW6, a novel HIF-1 inhibitor, promotes proteasomal degradation of HIF-1alpha via upregulation of VHL in a colon cancer cell line.

Hypoxia-inducible factor HIF-1 is responsible for radiation resistance and poor prognosis in cancer therapy. As part of our drug discovery program, a novel HIF inhibitor, LW6, was identified as a small compound that inhibits the accumulation of HIF-1alpha. We found that LW6 decreased HIF-1alpha protein expression without affecting HIF-1beta expression. MG132, a proteasome inhibitor, protected HIF-1alpha from LW6-induced proteasomal degradation, indicating that LW6 affects the stability of the HIF-1alpha protein. We found that LW6 promoted the degradation of wild type HIF-1alpha, but not of a DM-HIF-1alpha with modifications of P402A and P564A, at hydroxylation sites in the oxygen-dependent degradation domain (ODDD). LW6 did not affect the activity of prolyl hydroxylase (PHD), but induced the expression of von Hippel-Lindau (VHL), which interacts with prolyl-hydroxylated HIF-1alpha for proteasomal degradation. In the presence of LW6, knockdown of VHL did not abolish HIF-1alpha protein accumulation, indicating that LW6 degraded HIF-1alpha via regulation of VHL expression. In mice carrying xenografts of human colon cancer HCT116 cells, LW6 demonstrated strong anti-tumor efficacy in vivo and caused a decrease in HIF-1alpha expression in frozen-tissue immunohistochemical staining. These data suggest that LW6 may be valuable in the development of a HIF-1alpha inhibitor for cancer treatment.

Epidermal growth factor receptor regulates MT1-MMP and MMP-2 synthesis in SiHa cells via both PI3-K/AKT and MAPK/ERK pathways.

Matrix metalloproteinase 2 (MMP-2) and membrane type 1 matrix metalloproteinase (MT1-MMP) have been identified as important participants in tumor invasion, metastasis, and angiogenesis. Membrane type 1 matrix metalloproteinase has also been recognized as a major activator of MMP-2. The purpose of this study was to investigate epidermal growth factor (EGF) mediating signal pathways in the regulation of MMP-2 and MT1-MMP in SiHa cells, a cervical cancer cell line. We showed here that EGF induced the expression of MT1-MMP and inhibited the expression of MMP-2 at both the mRNA and protein levels. Membrane type 1 matrix metalloproteinase induction was blocked by mitogen-activated protein kinase or extracellular signal-regulated kinase inhibitors PD98059 and U0126 but not by phosphatidylinositol-3 kinase (PI3-K) inhibitors LY294002 and wortmannin. Interestingly, the mitogen-activated protein kinase or extracellular signal-regulated kinase inhibitors PD98059 and U0126 actually increased MMP-2 mRNA and protein synthesis, whereas the PI3-K inhibitors LY294002 and wortmannin further suppressed the expression of MMP-2. Our results suggest that EGF receptor up-regulated the expression of MT1-MMP and down-regulated the synthesis of MMP-2 through the mitogen-activated protein kinase/extracellular signal-regulated kinase pathway while concomitantly transmitting a mild positive regulatory signal to the expression of MMP-2 via the PI3-K/AKT pathway in SiHa cells. Furthermore, we found that EGF elevated the activity of MMP-2 in culture media.