Pleiotropic role of GAS6 in cardioprotection against ischemia-reperfusion injury.

INTRODUCTION: Myocardial ischemia-reperfusion (IR) injury is a common medical issue contributing to the onset and progression of ischemic heart diseases (IHD). Growth arrest-specific gene 6 (GAS6), a vitamin K-dependent secretory protein, promotes cell proliferation and inhibits inflammation and apoptosis through binding with Tyro3, Axl, and Mertk (TAM) receptors. OBJECTIVES: Our study aimed to examine the effect of GAS6 pathways activation as a potential new treatment in myocardial IR injury. METHODS: Gain- and loss-of-function experiments were utilized to determine the roles of GAS6 in the pathological processes of myocardial IR injury. RESULTS: Our results revealed down-regulated levels of GAS6, Axl, and SIRT1 in murine hearts subjected to IR injury, and cardiomyocytes challenged with hypoxia reoxygenation (HR) injury. GAS6 overexpression significantly improved cardiac dysfunction in mice subjected to myocardial IR injury, accompanied by reconciled mitochondrial dysfunction, oxidative stress, and apoptosis. In vitro experiments also observed a protective effect of GAS6 in cardiomyocytes. SIRT1 was found to function as a downstream regulator for GAS6/Axl signaling axis. Through screening a natural product library, a polyphenol natural compound catechin was identified to exhibit a protective effect by turning on GAS6/Axl-SIRT1 cascade. CONCLUSIONS: Together, our findings indicate that GAS6 emerges as a potential novel target in the management of myocardial IR injury and other related anomalies.

The regulatory role of PI3K in ageing-related diseases.

Ageing is a physiological/pathological process accompanied by the progressive damage of cell function, triggering various ageing-related disorders. Phosphatidylinositol 3-kinase (PI3K), which serves as one of the central regulators of ageing, is closely associated with cellular characteristics or molecular features, such as genome instability, telomere erosion, epigenetic alterations, and mitochondrial dysfunction. In this review, the PI3K signalling pathway was firstly thoroughly explained. The link between ageing pathogenesis and the PI3K signalling pathway was then summarized. Finally, the key regulatory roles of PI3K in ageing-related illnesses were investigated and stressed. In summary, we revealed that drug development and clinical application targeting PI3K is one of the focal points for delaying ageing and treating ageing-related diseases in the future.

Activation of NR1H3 signaling pathways by psoralidin attenuates septic myocardial injury.

Sepsis can cause various organ dysfunction, which heart failure may be associated with significant mortality. Recently, natural plant extracts have gradually attracted people's attention in the clinical treatment of cardiovascular diseases. Psoralidin (PSO) is one of the main bioactive compounds from the seeds of Psoralea corylifolia L and exhibits remarkable protective effects in diseases, including cancer, osteoporosis, and depression. Recently, NR1H3 is one of the emerging nuclear receptors targets for the various drugs. This study first reported the porotective role of PSO in septic myocardial injury, which was mainly attributed to the NR1H3-dependent manner. NR1H3 knockout mice subjected to cecal ligation and puncture (CLP) were used to investigate the involvement of NR1H3 in PSO protection. Our results showed that PSO prominently improved cardiac function, attenuated inflammation, inhibited oxidative stress, improved mitochondrial function, regulated ERS, suppressed apoptosis, and particularly increased NR1H3 and p-AMPK levels. However, NR1H3 knockout reversed the positive role of PSO in septic mice. Furthermore, activation of NR1H3 by T0901317 also increased the activity of AMPK and ACC in the HL-1 cardiomyocytes, indicating the regulatory relationship between NR1H3 and AMPK signaling. Together, this study demonstrated the beneficial effect of PSO in septic myocardial injury through activation of NR1H3/AMPK pathway.

Blood group B and decreased risk of coronary artery disease in hypertensive elderly / Grupo sanguíneo B y disminución del riesgo de enfermedad arterial coronaria en ancianos hipertensos

Abstract Although ABO blood groups have been associated with cardiovascular disease, little is known about whether ABO blood groups contribute to the risk of the presence and severity of coronary artery disease (CAD) in elderly individuals with hypertension. This study was aimed to explore this association. A total of 793 hypertensive patients aged ≥60 years out of 2095 patients who underwent primary coronary angiography were retrospectively included. They were divided into CAD and non-CAD groups. Demographic and clinical characteristics, ABO blood groups and other biochemical parameters were compared. Further evaluation was performed to determine the impact of ABO blood groups on CAD severity using the Gensini score and the number of significantly diseased vessels. A logistic regression model was constructed to identify the association of ABO blood groups with CAD. There was a substantial difference in the distribution of ABO blood groups in elderly and hypertensive adults with and without CAD (p=0.022). Hypertensive patients with CAD had a significantly lower proportion of the blood group B than those without CAD (p=0.008). Compared to those with non-Blood group B, hypertensive elderly with a blood group B tended to have significantly lower concentrations of TC, LDL -C and Apo B, and a lower number of significantly stenosed vessels. The blood group B was found to be an independent protective factor for CAD in elderly with hypertension. The blood group B is significantly associated with a decreased risk of CAD and is inversely correlated with the severity of coronary stenosis in the elderly with hypertension.

Resumen Aunque los tipos de sangre ABO están asociados con enfermedades cardiovasculares, se sabe poco sobre si los tipos de sangre ABO estás relacionados con la presencia y gravedad de la enfermedad arterial coronaria (CAD) en pacientes de edad avanzada con hipertensión. El objetivo de este estudio fue explorar esta relación. Se incluyeron retrospectivamente un total de 793 pacientes hipertensos de ≥60 años tomados de un grupo de 2095 pacientes sometidos a angiografía coronaria primaria. Se dividieron en el grupo de cardiopatía coronaria (CAD) y el grupo sin cardiopatía coronaria (no-CAD). Se compararon las características demográficas y clínicas, el grupo sanguíneo ABO y otros parámetros bioquímicos. El efecto del grupo sanguíneo ABO sobre la gravedad de la CAD se evaluó con la puntuación Gensini y el número de vasos sanguíneos patológicos significativos. Se construyó un modelo de regresión logística para determinar la relación entre el grupo sanguíneo ABO y la CAD. Hubo una diferencia significativa en la distribución de los grupos sanguíneos ABO entre los ancianos con y sin cardiopatía coronaria y los adultos con hipertensión (P=0,022). La proporción del grupo sanguíneo B en pacientes hipertensos con cardiopatía coronaria fue significativamente menor que en pacientes sin cardiopatía coronaria (P=0,008). En comparación con los grupos sanguíneos no B, las concentraciones de TC, LDL - C y Apolipoproteína B en los ancianos hipertensos del Grupo B fueron significativamente menores, y el número de estenosis vascular fue significativamente menor. El Grupo B es un factor protector independiente de la CAD en pacientes de edad avanzada con hipertensión. El Grupo B se correlacionó significativamente con la reducción del riesgo de cardiopatía coronaria y negativamente con la gravedad de la estenosis coronaria en pacientes de edad avanzada con hipertensión.

CTRP3 alleviates myocardial ischemia/reperfusion injury in mice through activating LAMP1/JIP2/JNK pathway.

Myocardial ischemia reperfusion (I/R) injury is an important complication of myocardial infarction reperfusion therapy, and no effective treatment has been identified. Based on preexisting evidence, C1q/tumor necrosis factor-related protein 3 (CTRP3) has been reported to be closely associated with myocardial dysfunction. In this study, we found that CTRP3 was downregulated in acute coronary syndrome (ACS) patients and myocardial I/R mice. Silence of CTRP3 aggravated cardiac systolic function due to I/R of mice, while CTRP3 overexpression ameliorated cardiac function. Moreover, overexpression of CTRP3 improved I/R inhibitory effects on the levels of creatinine phosphokinase (CPK), lactate dehydrogenase (LDH) and cardiac troponin-I (cTn-I), myocardial infarction area, the intensity of the 3-nitrotyrosine (3-NT), apoptosis and protein levels of LAMP1, JNK-Interacting Protein-2 (JIP-2) and JNK, while these effects could be exacerbated by downregulation of CTRP3. Co-IP experiments could identify physical interactions between CTRP3 and lysosomal-associated membrane protein 1 (LAMP1) and Numb and JIP2. LAMP1 silence aggravated the inhibition effects of I/R on JIP2 and JNK protein expression, CPK, LDH and cTn-I levels and caspase-3 activity, while overexpression of LAMP1 recovered these inhibition effects of I/R. JNK inhibitor (SP600125) could reverse the inhibitory effects of CTRP3 overexpression on CPK, LDH, cTn-I, myocardial infarction, strong positive staining for 3-NT and apoptosis. These findings demonstrated that CTRP3 protected against injury caused by myocardial I/R through activating LAMP1/JIP2/JNK pathway to attenuate myocardial injury, improve left ventricular function, decrease myocardial infarction, and reduce myocardial apoptosis.

CTRP3 alleviates cardiac ischemia/reperfusion injury via LAMP1/JIP2/JNK signaling pathway.

BACKGROUND: C1q/tumor necrosis factor-related protein 3 (CTRP3) has been reported to be a crucial regulator in myocardial infarction. Nevertheless, the potential molecular mechanism of CTRP3 in ischemia/reperfusion (I/R) injury remains largely unclear. METHODS: The cell model of myocardial I/R injury was established by oxygen-glucose deprivation/reoxygenation (OGD/R) of rat cardiomyocyte H9C2. Expression of CTRP3 and lysosomal-associated membrane protein 1 (LAMP1) was detected in H9C2 cells treated with oxygen-glucose deprivation/reoxygenation (OGD/R). H9C2 cells were transfected with overexpression plasmids of CTRP3 (pcDNA-CTRP3) and LAMP1 (pcDNA-LAMP1), or CTRP3 small interfering RNA (si-CTRP3) or/and pcDNA-LAMP1, and cell proliferation, apoptosis and oxidative stress were testified. Co-IP assay was performed to validate the relationship among CTRP3, LAMP1 and JIP2. The role of CTRP3 and LAMP1 in JIP2/JNK pathway was evaluated with Western blot assay. Furthermore, in vivo myocardial I/R injury model was constructed to investigate the effect of CTRP3. RESULTS: Overexpression of CTRP3 and LAMP1 both significantly promoted cell proliferation, inhibited apoptosis and the production of reactive oxygen species (ROS), malondialdehyde (MAD) and cardiac troponin (cTn-I), while silencing CTRP3 exerted the opposite effects, and LAMP1 overexpression reversed the effect of silencing CTRP3 on the aspects above. CTRP3 interacted with LAMP1, and both CTRP3 and LAMP1 bound with JIP2. SP600125 (JNK inhibitor) could restore the effects of CTRP3 or LAMP1 overexpression on the expression of JIP2 and phosphorylated-JNK (p-JNK), proliferation and apoptosis. Moreover, overexpression of CTRP3 improved cardiac I/R injury in vivo. CONCLUSION: CTRP3 alleviates cardiac I/R injury by elevating LAMP1 and activating JIP2/JNK signaling pathway, which may serve as a potential therapeutic target for I/R injury.

Inhibition of EMMPRIN by microRNA-124 suppresses the growth, invasion and tumorigenicity of gliomas.

MicroRNAs (miR) are a group of non-coding, small RNAs, 18-20 nucleotides in length, that are frequently involved in the development of a variety of different types of cancer, including glioma, which is a type of severe tumor in the brain. Previous studies reported that miR-124 levels were downregulated in glioma specimens; however, the potential role of miR-124 in glioma currently remains unclear. The present study performed experiments, including dual-luciferase reporter assay (DLRA), MTT assay, transwell assay and flow cytometry, with the aim of elucidating the molecular mechanism of miR-124 in glioma. The results indicated that miR-124 expression was decreased in glioma tissues, accompanied by the increased expression of extracellular matrix metalloproteinase inducer (EMMPRIN). The expression of EMMPRIN was inhibited by miR-124 transfection. The DLRA results revealed that EMMPRIN directly targets miR-124. Furthermore, upon overexpression of miR-124 in the U87 cells, cell proliferation was significantly inhibited, apoptosis was increased, and cell migration and invasion were decreased. Furthermore, tumor growth was blocked by miR-124 in mice. Based on these results, the present study concluded that miR-124 is critical for amelioration of glioma by targeting EMMPRIN, thereby acting as a tumor suppressor. Thus, miR-124/EMMPRIN constitutes a plausible basis for the treatment of glioma.

SNHG9, delivered by adipocyte-derived exosomes, alleviates inflammation and apoptosis of endothelial cells through suppressing TRADD expression.

Exosomes are membrane-derived vesicles and play a critical role in cell signaling by transferring RNAs and proteins to target cells through fusion with the cell membrane. Long non-coding RNA-small nucleolar RNA host gene 9 (lncRNA-SNHG9) was proven to be an important element in lncRNA-mRNA interaction networks during adipocyte differentiation, suggesting its potential involvement in the development of obesity, an important risk factor of cardiovascular and cerebrovascular endothelial dysfunction. However, the role of lncRNA-SNHG9 within the exosome in endothelial dysfunction of obese patients is largely unknown. In this study, we proved that adipocytes-derived exosomal SNHG9 were downregulated in obese persons and further decreased in obese individuals with endothelial dysfunction. Functional experimentations demonstrated that adipocytes-derived exosomal SNHG9 alleviated inflammation and apoptosis in endothelial cells. Bioinformatic analysis revealed that there was a potential interaction between SNHG9 and the TNF receptor type 1-associated death domain protein (TRADD) mRNA. Then, RNA-binding protein immunoprecipitation assay based on Ago2 antibody and ribonuclease protection assay demonstrated that exosomal SNHG9 directly bound to a specific region in TRADD mRNA sequence and formed an RNA dimeric inducible silencing complex. Moreover, knockdown of TRADD markedly inhibited inflammation and apoptosis in human umbilical vein endothelial cells (HUVECs), whereas overexpression of TRADD dramatically neutralized the protective effect of exosomal SNHG9 on epithelial dysfunction. Therefore, SNHG9 could prevent endothelial dysfunction in obese patients by suppressing inflammation and apoptosis, indicating that SNHG9 may be a potential therapeutic target for obese patients with endothelial dysfunction.

Biological Response to Recombinant Human Bone Morphogenetic Protein-2 on Bone-Implant Osseointegration in Ovariectomized Experimental Design.

OBJECTIVE: The aim of this study is to investigate the effects of recombinant human bone morphogenetic protein-2 (rhBMP-2) on bone-implant osteointegration in osteoporotic rats. MATERIALS AND METHODS: Thirty-six female Wistar rats were randomly divided into 3 groups: sham-operation (SHAM), ovariectomized (OVX), and ovariectomized with rhBMP-2 (OVX + rhBMP-2). The bone density of right tibia was observed with x-ray and the serum alkaline phosphatase (ALP) activity was measured preovariectomy and postovariectomy using an ALP-kit. In OVX + rhBMP-2 group, rhBMP-2 was embedded in the peri-implant area, while SHAM and OVX groups did not contain rhBMP-2. Four and eight weeks after implantation, the rats were killed and the right tibia with implants was taken by x-ray. Histologic changes were investigated by hematoxylin and eosin staining, scanning electron microscope (SEM), and energy dispersive spectrometer examinations. RESULTS: The serum ALP level was significantly higher in ovariectomized rats compared with that before ovariectomy (P < 0.05), while no difference was found in SHAM rats. At 12 weeks after ovariectomy, radiographic and histologic findings showed significant osteoporotic changes in proximal tibial metaphyses of OVX rats, including reduced cortical bone density and enlargement of bone marrow cavity compared with SHAM ones. The results of implantation verified new bone formation around implants in OVX + rhBMP-2 and SHAM groups, indicating favorable bone healing and osseointegration. No bone resorption was found in OVX + rhBMP-2 group, while some soft tissue was observed in bone-implant interface in SHAM group. In OVX group, there was no effective bone-implant osseointegration and mature bone formed around implants, and some implants were even lost due to chronic inflammation. The percentage of calcium and phosphorous atoms was significantly higher and the percentage of sulfur element was significantly lower in peri-implant area in OVX + rhBMP-2 and SHAM groups than that in OVX group. CONCLUSION: rhBMP-2 could enhance the osseous healing and restore bone-implant osseointegration in osteoporotic rats.

miRNA143 Induces K562 Cell Apoptosis Through Downregulating BCR-ABL.

BACKGROUND Leukemia seriously threats human health and life. MicroRNA regulates cell growth, proliferation, apoptosis, and cell cycle. Whether microRNA could be treated as a target for leukemia is still unclear and the mechanism by which microRNA143 regulates K562 cells needs further investigation. MATERIAL AND METHODS miRNA143 and its scramble miRNA were synthesized and transfected to K562 cells. MTT assay was used to detect K562 cell proliferation. Flow cytometry and a caspase-3 activity detection kit were used to test K562 cell apoptosis. Western blot analysis was performed to determine breakpoint cluster region-Abelson (BCR-ABL) expression. BCR-ABL overexpression and siRNA were used to change BCR-ABL level, and cell apoptosis was detected again after lipofection transfection. RESULTS miRNA143 transfection inhibited K562 cell growth and induced its apoptosis. miRNA143 transfection decreased BCR-ABL expression. BCR-ABL overexpression suppressed miRNA143-induced K562 cell apoptosis, while its reduction enhanced miRNA143-induced apoptosis. CONCLUSIONS miRNA143 induced K562 cell apoptosis through downregulating BCR-ABL. miRNA143 might be a target for a new leukemia therapy.

Increased expression of Six1 correlates with progression and prognosis of prostate cancer.

Sineoculis homeobox homolog 1 (Six1), normally a developmentally restricted transcriptional regulator, is frequently dysregulated in mutiple cancers. Increasing evidences show that overexpression of Six1 plays a key role in tumorigenesis. However, the Six1 expression status and its relationship with the clinicopathological characteristics in prostate cancer were unclear. In this study, the mRNA and protein levels of Six1 in prostate cancer tissues and normal prostate tissues were evaluated. The clinicopathological significance of Six1 was investigated by immunohistochemistry (IHC) on a prostate cancer tissue microarray. The cut-off score for high expression of Six1 was determined by the receiver-operating characteristic (ROC) analysis. The correlation between Six1 protein expression and clinicopathological characteristics of prostate cancer was analyzed by Chi-square test. Increased expression of Six1 protein was observed in the majority of prostate cancer, compared with their paired adjacent normal prostate tissues. When Six1 high expression percentage was determined to be above 55 % (area under ROC curve = 0.881, P = 0.000), high expression of Six1 was observed in 55.6 % (80/144) of prostate cancer tissues and low expression of Six1 was observed in all normal prostate tissues by IHC. Increased expression of Six1 in patients was correlated with high histological grade (χ2 = 58.651, P = 0.00), advanced clinical stage (χ2 = 57.330, P = 0.000), high Gleason score (χ2 = 63.480, P = 0.000), high primary tumor grade (χ2 = 57.330, P = 0.000) and positive regional lymph node metastasis (χ2 = 19.294, P = 0.000). Furthermore, univariate and multivariate survival analysis suggested that Six1 was an independent prognostic indicator for overall survival (P < 0.05). This study suggests that Six1 could be served as an additional biomarker in identifying prostate cancer patients at risk of tumor progression, might potentially be used for predicting survival outcome of patients with prostate cancer.

Calcium overload and in vitro apoptosis of the C6 glioma cells mediated by sonodynamic therapy (hematoporphyrin monomethyl ether and ultrasound).

UNLABELLED: The objective of this study was to investigate the role of intracellular calcium overload in the in vitro apoptosis of C6 glioma cells mediated by low level ultrasound and hematoporphyrin monomethyl ether (HMME) therapy. The frequency of ultrasound was optimized by the cell viability assay using 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The apoptotic rate, reactive oxygen species (ROS) and decreased mitochondrial membrane potential (MMP) were determined by flow cytometry. Morphological changes were observed by the transmission electron microscope. Concentrations of intracellular Ca2+, [Ca2+]i were detected by a confocal microscopic laser scanning, and the release of cytochrome-c (cyt-c) was measured by western blotting. RESULTS: The SDT-mediated apoptotic effect involved an overload of [Ca2+]i derived from the intra- and extracellular sources during the early progression of apoptotosis. The process was associated with an increased ROS production, a decreased MMP, and a release of cyt-c. In conclusion,the combined use of low level ultrasound and HMME improved the apoptotic rate of C6 glioma cells mediated by ultrasound alone. The [Ca2+]i overload involving activation of mitochondrial signaling played a pivotal role in the SDT-induced apoptosis.

Low expression of miR-196b enhances the expression of BCR-ABL1 and HOXA9 oncogenes in chronic myeloid leukemogenesis.

MicroRNAs (miRNAs) can function as tumor suppressors or oncogene promoters during tumor development. In this study, low levels of expression of miR-196b were detected in patients with chronic myeloid leukemia. Bisulfite genomic sequencing PCR and methylation-specific PCR were used to examine the methylation status of the CpG islands in the miR-196b promoter in K562 cells, patients with leukemia and healthy individuals. The CpG islands showed more methylation in patients with chronic myeloid leukemia compared with healthy individuals (P<0.05), which indicated that low expression of miR-196b may be associated with an increase in the methylation of CpG islands. The dual-luciferase reporter assay system demonstrated that BCR-ABL1 and HOXA9 are the target genes of miR-196b, which was consistent with predictions from bioinformatics software analyses. Further examination of cell function indicated that miR-196b acts to reduce BCR-ABL1 and HOXA9 protein levels, decrease cell proliferation rate and retard the cell cycle. A low level of expression of miR-196b can cause up-regulation of BCR-ABL1 and HOXA9 expression, which leads to the development of chronic myeloid leukemia. MiR-196b may represent an effective target for chronic myeloid leukemia therapy.

The association between polymorphisms in the MDR1 gene and risk of cancer: a systematic review and pooled analysis of 52 case-control studies.

BACKGROUND: The multidrug resistance (MDR) 1 gene encodes a 170-kDa membrane transporter called P-glycoprotein, which plays an important role in protecting cells against lipophilic xenobiotics by the way of an ATP-dependent cellular efflux mechanism. Three polymorphisms of MDR1, 3435C > T located in exon 26, 1236C > T in exon 12 and 2677G > T/A in exon 21 were the most extensively studied and were identified functionally important and ethnically diverse mapping to the gene region. Considering the potential influence of altering MDR1 activity, it is plausible that MDR1 polymorphisms might play a role in the development of cancer. Although the effects of MDR1 polymorphisms on susceptibility to human cancer have been investigated in many studies, the results still remain conflicting. METHODS: To resolve these conflicts, we performed a quantitative synthesis of the association between these three polymorphisms and cancer risk, including 52 studies (15789 cases and 20274 controls) for 3435C > T polymorphism, 10 studies (2101 cases and 2842 controls) for 1236C > T polymorphism and 18 studies (3585 cases and 4351 controls) for 2677G > T/A polymorphism. RESULTS: The stratified analyses for 3435C > T polymorphism, individuals with T-allele in 3435C > T had significantly higher ALL risks (TT versus CC: OR =1.286, 95% CI =1.123-1.474); significantly elevated risks were observed among Caucasian populations (TT versus CC: OR =1.276, 95% CI =1.112-1.464). When restricting the analysis to the source of controls, we found that HB (hospital-based) genetic models had higher risks (TT versus CC: OR =1.307, 95% CI =1.046-1.632), as well as in PB (population-based) genetic models (TT versus CC: OR =1.294, 95% CI =1.079-1.55).The T/A-allele frequency of 2677G > T/A polymorphism was associated with higher risk of cancer (TT + TA + AA vs. GG: OR =1.348, 95% CI =1.031-1.762), significantly elevated risks were observed among Asian populations (TT + TA + AA vs. GG: OR =1.642, 95% CI =1.340-2.012), and elevated risks could be associated with PB models (TT + TA + AA vs. GG: OR =1.641, 95% CI =1.018-2.646). CONCLUSIONS: Our meta-analysis suggested that 3435C > T polymorphism and 2677G > T/A polymorphism were associated with cancer risk when all studies were pooled together, while 1236C > T polymorphism not.

Decreased microRNA-30a levels are associated with enhanced ABL1 and BCR-ABL1 expression in chronic myeloid leukemia.

Chronic myeloid leukemia (CML) is associated with overexpression of BCR-ABL1, a nonreceptor tyrosine kinase critical for malignant transformation. We investigated whether non-coding microRNAs (miRNAs) targeting BCR-ABL1 mRNA contribute to the pathogenesis of CML. Indeed, miR-30a targeted BCR-ABL1 and was underexpressed in bone marrow from CML patients. In K562 leukemia cells, overexpression of miR-30a reduced ABL1 and BCR-ABL1 protein expression, decreased proliferation, and arrested cell cycle progression between G1 and S. These findings strongly suggest that miR-30a acts as a tumor suppressor by downregulating ABL1 and BCR-ABL1 expression. Upregulation of miR-30a in hematopoietic cells may have therapeutic efficacy against CML.

[Analysis and verification of the interaction network of differentially expressed genes in invasive bladder cancer].

OBJECTIVE: To study the data of gene expression microarray by protein interaction network analysis, establish an interaction network of differentially expressed genes in invasive bladder cancer and verify the central nodes of the network. METHODS: A total of 152 differentially expressed genes in invasive bladder cancer detected by gene expression microarray were inputted into STRING database online for analysis and establishment of the interaction network. The interaction data were imported into Cytoscape 2.6.2 software for screening the central nodes of the network. KEGG database was exploited for pathway analysis and functional study of the central node genes. Real-time RT-PCR was used for verification, and the genes with maximal differential expressions were screened for exploring the molecular mechanism of carcinogenesis of invasive bladder cancer. RESULTS: The protein products of 103 differentially expressed genes in bladder cancer had interactions, forming a complicated interaction network. Twenty-six nodes involved in several signal pathways were confirmed by Cytoscape as the central nodes of the network, among which UBE2C, VEGF, TGFBR2, and CAV1 nodes were verified by real-time RT-PCR as the genes with maximal differential expressions between the bladder cancer and normal tissues, and the 2(-delta delta Ct) of these genes were 9.45, 4.17, 0.13 and 0.18 (GAPHD as the internal control), respectively. CONCLUSION: The interaction network of the differentially expressed genes, especially the central nodes of this network, can provide clues to the carcinogenesis, early diagnosis and molecular targeted therapy of invasive bladder cancer.

[Effect of vascular endothelial growth factor gene transfer on proliferation and metabolism of human bone marrow stromal cells in vitro].

OBJECTIVE: To investigate the effect of vascular endothelial growth factor 165 (VEGF165) gene transfer on the proliferation and metabolism of human bone marrow stromal cells (hBMSCs) in vitro. METHODS: hBMSCs were divided into 3 groups and subjected to adenovirus mediated VEGF165 gene transfection, transfection with empty adenoviral vector, or left untreated (control). MTT assay and flow cytometry were performed to analyze the proliferation of the cells after corresponding treatments. The third passage of hBMSCs (2x10(4)/ml), after corresponding transfection procedures, were cultured in conditional medium and tested for ALP content 2, 4 and 6 days after the transfection. Also at 3, 5 and 7 days after the transfection, the cells were examined for osteocalcin (C) and laminin (LN) contents. RESULTS: The number of cells in each group increased with the culture time without obvious differences in the optical density. No significant differences were noted between the 3 groups in the percentage of G1 phase cells or in the proliferation index (PrI) (P>0.05), but compared with the nontransfected and the empty vector-transfected cells, the cells with VEGF165 gene transfection had significantly higher ALP, OC and LN contents (P<0.05). CONCLUSION: VEGf165 gene transfer does not obviously affect the proliferation of cultured hBMSCs, but can increase the cellular secretion of AIP, C and LN, suggesting that VEGF165 promotes the differentiation of hBMSCs into osteoblasts in vitro.

[Screening differentially expressed genes in denucleated K562 cells with restriction display technique].

OBJECTIVE: To screen differentially expressed genes in cytochalasin B (CB)-induced denucleated K562 cells by restriction display (RD) technique. METHODS: The total RNA was isolated and purified from K562 cells before and after CB (10 mug/ml) treatment. The mRNA from both treated and untreated K562 cells were reversely transcribed into cDNA, and the differentially expressed genes were separated using RD technique combined with polyacrylamide gel electrophoresis and sliver staining, followed by cloning, sequencing and homology analysis against GenBank database of these genes. RESULTS: Seven differentially expressed genes were identified in CB-treated cells including aquaporin 1 (AQP1) gene, which was verified to be up-regulated after CB treatment by RT-PCR. CONCLUSION: AQP1 gene might be in close association with the regulation of denucleation processes and CB-induced proliferation inhibition of K562 cells.

[c-myc gene silencing in K562 cells with RNA interference].

OBJECTIVE: To study the inhibitory effect of small interfering RNA (siRNA) targeting c-myc gene in K562 cells. METHODS: siRNAs targeting the site 1357 of c-myc mRNA was designed and synthesized. In vitro cultured K562 cells were transfected with lipofectamine 2000 and the inhibitory effect was detected by reverse transcriptase (RT)-PCR, cell count, MTT assay and fluorescence-activated cell sorting. RESULTS: Compared with the negative and blank control group, the transfection group showed marked decrease in the c-myc expression and the K562 cells exhibited increased apoptosis rate. CONCLUSION: RNA interference can effectively inhibit c-myc expression and induce apoptosis in K562 cells.

[Construction and preliminary identification of subtracted cDNA library of leukemia cell line K562].

OBJECTIVE: Subtractive hybridization technology is a common method to screen and clone differentially expressed genes. This study was to construct subtracted cDNA library of leukemia cell line K562, and screen for differentially expressed genes. METHODS: cDNA fragments of K562 cells (tester), prepared by restriction display (RD), were subtracted with the Sau3A I-digested cDNA fragments of normal lymphocytes (driver). The subtracted cDNA fragments were re-amplified, and cloned into pMD18-T vectors. Positive clones were selected by blue-white screening. The inserts in plasmid were amplified by polymerase chain reaction (PCR), and some of which were sequenced. RESULTS: The subtracted library contained 360 positive clones with cDNA fragments distributed mainly from 200 to 800 bp. The 50 randomly sequenced clones were derived from 42 known genes. CONCLUSION: Specific subtracted cDNA library of K562 cells was successfully constructed with reliable quality, and may be used to further screen and clone differentially expressed genes of K562 cells.