Intratumor Heterogeneity in Interstitial Fluid Pressure in Cervical and Pancreatic Carcinoma Xenografts.

Preclinical studies have suggested that interstitial fluid pressure (IFP) is uniformly elevated in the central region of tumors, whereas clinical studies have revealed that IFP may vary among different measurement sites in the tumor center. IFP measurements are technically difficult, and it has been claimed that the intratumor heterogeneity in IFP reported for human tumors is due to technical problems. The main purpose of this study was to determine conclusively whether IFP may be heterogeneously elevated in the central tumor region, and if so, to reveal possible mechanisms and possible consequences. Tumors of two xenograft models were included in the study: HL-16 cervical carcinoma and Panc-1 pancreatic carcinoma. IFP was measured with Millar SPC 320 catheters in two positions in each tumor and related to tumor histology or the metastatic status of the host mouse. Some tumors of both models showed significant intratumor heterogeneity in IFP, and this heterogeneity was associated with a compartmentalized histological appearance (i.e., the tissue was divided into compartments separated by thick connective tissue bands) in HL-16 tumors and with a dense collagen-I-rich extracellular matrix in Panc-1 tumors, suggesting that these connective tissue structures prevented efficient interstitial convection. Furthermore, some tumors of both models developed lymph node metastases, and of the two IFP values measured in each tumor, only the higher value was significantly higher in metastatic than in non-metastatic tumors, suggesting that metastatic propensity was determined by the tumor region having the highest IFP.

DCE-MRI and Quantitative Histology Reveal Enhanced Vessel Maturation but Impaired Perfusion and Increased Hypoxia in Bevacizumab-Treated Cervical Carcinoma.

PURPOSE: This study had a dual purpose: to investigate (1) whether bevacizumab can change the microvasculature and oxygenation of cervical carcinomas and (2) whether any changes can be detected with dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). METHODS AND MATERIALS: Two patient-derived xenograft models of cervical cancer (BK-12 and HL-16) were included in the study. Immunostained histologic preparations from untreated and bevacizumab-treated tumors were analyzed with respect to microvascular density, vessel pericyte coverage, and tumor hypoxia using CD31, α-SMA, and pimonidazole as markers, respectively. DCE-MRI was performed at 7.05 T, and parametric images of Ktrans and ve were derived from the data using the Tofts pharmacokinetic model. RESULTS: The tumors of both models showed decreased microvascular density, increased vessel pericyte coverage, and increased vessel maturation after bevacizumab treatment. Bevacizumab-treated tumors were more hypoxic and had lower Ktrans values than untreated tumors in the BK-12 model, whereas bevacizumab-treated and untreated HL-16 tumors had similar hypoxic fractions and similar Ktrans values. Significant correlations were found between median Ktrans and hypoxic fraction, and the data for untreated and bevacizumab-treated tumors were well fitted by the same curve in both tumor models. CONCLUSIONS: Bevacizumab-treated tumors show less abnormal microvessels than untreated tumors do, but because of treatment-induced vessel pruning, the overall function of the microvasculature might be impaired after bevacizumab treatment, resulting in increased tumor hypoxia. DCE-MRI has great potential for monitoring bevacizumab-induced changes in tumor hypoxia in cervical carcinoma.

Increasing aggressiveness of patient-derived xenograft models of cervix carcinoma during serial transplantation.

Four patient-derived xenograft (PDX) models (BK-12, ED-15, HL-16, LA-19) of carcinoma of the uterine cervix have been developed in our laboratory, and their stability during serial transplantation in vivo was investigated in this study. Two frozen cell stocks were established, one from xenografted tumors in passage 2 (early generation) and the other from xenografted tumors transplanted serially in mice for approximately two years (late generation), and the biology of late generation tumors was compared with that of early generation tumors. Late generation tumors showed higher incidence of lymph node metastases than early generation tumors in three models (ED-15, HL-16, LA-19), and the increased metastatic propensity was associated with increased tumor growth rate, increased microvascular density, and increased expression of angiogenesis-related and cancer stem cell-related genes. Furthermore, late generation tumors showed decreased fraction of pimonidazole-positive tissue (i.e., decreased fraction of hypoxic tissue) in two models (HL-16, LA-19) and decreased fraction of collagen-I-positive tissue (i.e., less extensive extracellular matrix) in two models (ED-15, HL-16). This study showed that serially transplanted PDXs may not necessarily mirror the donor patients' diseases, and consequently, proper use of serially transplanted PDX models in translational cancer research requires careful molecular monitoring of the models.

Integrins as therapeutic targets in the organ-specific metastasis of human malignant melanoma.

Integrins are a large family of adhesion molecules that mediate cell-cell and cell-extracellular matrix interactions. Among the 24 integrin isoforms, many have been found to be associated with tumor angiogenesis, tumor cell migration and proliferation, and metastasis. Integrins, especially αvß3, αvß5 and α5ß1, participate in mediating tumor angiogenesis by interacting with the vascular endothelial growth factor and angiopoietin-Tie signaling pathways. Melanoma patients have a poor prognosis when the primary tumor has generated distant metastases, and the melanoma metastatic site is an independent predictor of the survival of these patients. Different integrins on the melanoma cell surface preferentially direct circulating melanoma cells to different organs and promote the development of metastases at specific organ sites. For instance, melanoma cells expressing integrin ß3 tend to metastasize to the lungs, whereas those expressing integrin ß1 preferentially generate lymph node metastases. Moreover, tumor cell-derived exosomes which contain different integrins may prepare a pre-metastatic niche in specific organs and promote organ-specific metastases. Because of the important role that integrins play in tumor angiogenesis and metastasis, they have become promising targets for the treatment of advanced cancer. In this paper, we review the integrin isoforms responsible for angiogenesis and organ-specific metastasis in malignant melanoma and the inhibitors that have been considered for the future treatment of metastatic disease.

Metastatic pathway and the microvascular and physicochemical microenvironments of human melanoma xenografts.

BACKGROUND: Malignant melanoma of the skin can metastasize through blood vessels and lymphatics. The primary tumor develops a vascular microenvironment characterized by abnormal blood vessels and lymphatics and a physicochemical microenvironment characterized by low oxygen tension, regions with hypoxic tissue, and high interstitial fluid pressure (IFP). This study aimed at identifying relationships between the metastatic route of melanomas and characteristic features of the microvascular and physicochemical microenvironments of the primary tumor. METHODS: Two patient-derived xenograft (PDX) models (E-13, N-15) and four cell line-derived xenografts (CDX) models (C-10, D-12, R-18, T-22) of human melanoma were included in the study. Tumors were transplanted to an orthotopic site in BALB/c-nu/nu mice, and when the tumors had grown to a volume of 500-600 mm3, the IFP of the primary tumor was measured and the hypoxia marker pimonidazole was administered before the host mouse was euthanized. The primary tumor, lungs, and six pairs of lymph nodes were evaluated by examining hematoxylin/eosin-stained and immunostained histological preparations. The expression of angiogenesis-related genes was assessed by quantitative PCR. RESULTS: C-10, D-12, and E-13 tumors disseminated primarily by the hematogenous route and developed pulmonary metastases. These tumors showed high angiogenic activity and high expression of the F3 gene as well as ANGPT2 and TIE1, genes encoding proteins of the angiopoietin-tie system. N-15, R-18, and T-22 tumors disseminated mainly by the lymphogenous route and developed metastases in draining lymph nodes. These tumors had highly elevated IFP and showed high expression of NRP2, a gene encoding neuropilin-2. CONCLUSION: The primary metastatic route of orthotopic human melanoma xenografts and the development of lung and lymph node metastases are influenced significantly by the microvascular and physicochemical microenvironments of the primary tumor.

MtDNA depletion influences the transition of CD44 subtypes in human prostate cancer DU145 cells.

Our earlier study revealed that long-term ethidium bromide application causes mitochondrial DNA depletion in human prostate cancer DU145 cell line (DU145MtDP), and this DU145MtDP subline appears to have expanded CD44Bright cell population than its parental wild type DU145 cells (DU145WT). Increasing evidence suggests that CD44Bright cells are highly cancer stem cell like, but it is not clear about their dynamic transition between CD44Dim and CD44Bright phenotypes in prostate cancer cells, and how it is affected by mitochondrial DNA depletion. To address these questions, four cell subpopulations were isolated from both DU145WT and DU145MtDP cell lines based on their CD44 expression level and mitochondrial membrane potential. The cell motility and colony formation capability of the fluorescence activated cell sorting-sorted cell subpopulations were further examined. It was discovered in the DU145WT cells that CD44Dim cells could transit into both CD44Dim and CD44Bright phenotypes and that CD44Bright cells were prone to sustain their CD44Bright phenotype as renewal. However, such transition principle was altered in the DU145MtDP cells, in which CD44Bright cells showed similar capability to sustain a CD44Bright phenotype, while the transition of CD44Dim cells to CD44Bright were suppressed. It is concluded that mitochondrial DNA depletion in the human prostate cancer DU145 cells influences their renewal and CD44 subphenotype transition. Such alterations may be the driving force for the enrichment of CD44Bright DU145 cells after the mitochondrial DNA depletion, although the molecular mechanisms remain unclear.

Metabolic reprogramming is associated with flavopiridol resistance in prostate cancer DU145 cells.

Flavopiridol (FP) is a pan-cyclin dependent kinase inhibitor, which shows strong efficacy in inducing cancer cell apoptosis. Although FP is potent against most cancer cells in vitro, unfortunately it proved less efficacious in clinical trials in various aggressive cancers. To date, the molecular mechanisms of the FP resistance are mostly unknown. Here, we report that a small fraction human prostate cancer DU145 cells can survive long-term FP treatment and emerge as FP-resistant cells (DU145FP). These DU145FP cells show accumulated mitochondrial lesions with stronger glycolytic features, and they proliferate in slow-cycling and behave highly migratory with strong anti-apoptotic potential. In addition, the cells are less sensitive to cisplatin and docetaxel-induced apoptotic pressure, and over-express multiple stem cell associated biomarkers. Our studies collectively uncover for the first time that FP-resistant prostate cancer cells show metabolic remodeling, and the metabolic plasticity might be required for the FP resistance-associated cancer cell stemness up-regulation.

Lymph node metastasis and the physicochemical micro-environment of pancreatic ductal adenocarcinoma xenografts.

Pancreatic ductal adenocarcinoma (PDAC) patients develop lymph node metastases early and have a particularly poor prognosis. The poor prognosis has been shown to be associated with the physicochemical microenvironment of the tumor tissue, which is characterized by desmoplasia, abnormal microvasculature, extensive hypoxia, and highly elevated interstitial fluid pressure (IFP). In this study, we searched for associations between lymph node metastasis and features of the physicochemical microenvironment in an attempt to identify mechanisms leading to metastatic dissemination and growth. BxPC-3 and Capan-2 PDAC xenografts were used as preclinical models of human PDAC. In both models, lymph node metastasis was associated with high IFP rather than high fraction of hypoxic tissue or high microvascular density. Seven angiogenesis-related genes associated with high IFP-associated lymph node metastasis were detected by quantitative PCR in each of the models, and these genes were all up-regulated in high IFP/highly metastatic tumors. Three genes were mutual for the BxPC-3 and Capan-2 models: transforming growth factor beta, angiogenin, and insulin-like growth factor 1. Further comprehensive studies are needed to determine whether there is a causal relationship between the up-regulation of these genes and high IFP and/or high propensity for lymph node metastasis in PDAC.

Cancer stem cells (CSCs), cervical CSCs and targeted therapies.

Accumulating evidence has shown that cancer stem cells (CSCs) have a tumour-initiating capacity and play crucial roles in tumour metastasis, relapse and chemo/radio-resistance. As tumour propagation initiators, CSCs are considered to be promising targets for obtaining a better therapeutic outcome. Cervical carcinoma is the most common gynaecological malignancy and has a high cancer mortality rate among females. As a result, the investigation of cervical cancer stem cells (CCSCs) is of great value. However, the numbers of cancer cells and corresponding CSCs in malignancy are dynamically balanced, and CSCs may reside in the CSC niche, about which little is known to date. Therefore, due to their complicated molecular phenotypes and biological behaviours, it remains challenging to obtain "purified" CSCs and continuously culture CSCs for further in vitro studies without the cells losing their stem properties. At present, CSC-related markers and functional assays are used to purify, identify and therapeutically target CSCs both in vitro and in vivo. Nevertheless, CSC-related markers are not universal to all tumour types, although some markers may be valid in multiple tumour types. Additionally, functional identifications based on CSC-specific properties are usually limited in in vivo studies. Furthermore, an optimal method for identifying potential CCSCs in CCSC studies has not been previously published, and these techniques are currently of great importance. This article updates our knowledge on CSCs and CCSCs, reviews potential stem cell markers and functional assays for identifying CCSCs, and describes the potential of targeting CCSCs in the treatment of cervical carcinoma.

Decreased expression of pyruvate dehydrogenase A1 predicts an unfavorable prognosis in ovarian carcinoma.

Pyruvate dehydrogenase A1 (PDHA1) serves as a gate-keeper enzyme link between glycolysis and the mitochondrial citric acid cycle. The inhibition of PDHA1 in cancer cells can result in an increased Warburg effect and a more aggressive phenotype in cancer cells. This study was conducted to investigate the expression of PDHA1 in ovarian cancer and the correlation between PDHA1 expression and the prognosis of patients. The PDHA1 protein expression in 3 ovarian cancer cell lines (OVCAR-3, SKOV-3 and ES-2) and 248 surgically removed ovarian carcinoma samples was immunocytochemically examined. Statistical analyses were performed to evaluate the correlations between PDHA1 expression and the clinicopathological characteristics of the patients as well as the predictive value of PDHA1. The results showed the presence of variable expression of PDHA1 in the three ovarian cancer cell lines. Of the 248 ovarian cancer tissue specimens, 45 cases (18.1%) were negative in tumor cells for PDHA1, 162 cases (65.3%) displayed a low expression level, and 41 cases (16.5%) had a relatively high PDHA1 staining. The expression of PDHA1 was associated with the histological subtype (P=0.004) and FIGO stage (P=0.002). The median OS time in the PDHA1 negative group, low expression group and high expression group were 0.939 years, 1.443 years and 9.900 years, respectively. The median PFS time in the above three groups were 0.287 years, 0.586 years and 9.900 years, respectively. Furthermore, the high expression of PDHA1 in ovarian carcinoma cells was significantly associated with better OS and PFS by statistical analyses. Multivariate analyses showed that PDHA1 expression was also an independent prognostic factor for higher OS in ovarian cancer patients (HR=0.705, 95% CI 0.541-0.918, P=0.01). Our study indicated that the decreased expression of PDHA1 might be an independent prognostic factor in unfavorable outcomes.

Functional intratumoral lymphatics in patient-derived xenograft models of squamous cell carcinoma of the uterine cervix: implications for lymph node metastasis.

Studies of cell line-derived human tumor xenografts have suggested that the lymphatics seen in immunohistochemical preparations from non-peripheral regions of tumors are nonfunctional. In this investigation, lymphangiogenesis, hemangiogenesis, and lymph node metastasis were studied in patient-derived xenograft (PDX) models of carcinoma of the uterine cervix. Lymph vessel density (LVD) and blood vessel density (BVD) were measured in immunohistochemical preparations. The expression of angiogenesis-related genes was investigated by quantitative PCR. Lymphatic functionality was assessed with the ferritin assay, and tumor interstitial fluid pressure (IFP) was measured with a Millar catheter. The PDX models mirrored the angiogenesis and aggressiveness of the donor patients' tumors, and two highly aggressive models developed functional lymphatics within the tumor mass. Tumors with functional intratumoral lymphatics showed low IFP, high LVD, high BVD, high expression of a large number of angiogenesis-related genes, and high incidence of lymph node metastases. LVD correlated with BVD, and lymph node metastasis was associated with high LVD and high BVD. Nine angiogenesis-related genes associated with the development of functional intratumoral lymhatics were identified. High expression of these genes, high LVD, and high BVD may be important biomarkers for poor outcome in cervix carcinoma.

MicroRNA-155 is a potential molecular marker of natural killer/T-cell lymphoma.

Natural killer/T-cell lymphoma (NKTCL) is characterized by its highly aggressive nature and rapid progression. MicroRNAs (miRNAs) play key roles in the development of NKTCL. We utilized next-generation Solexa high-throughput sequencing to compare miRNA expression in the SNK-6 and YTS NKTCL cell lines with expression in normal NK cells. We found that 195 miRNAs were upregulated in the SNK-6 cells and 286 miRNAs were upregulated in the YTS cells. Based on those results, we selected six miRNAs, including miRNA-155, and confirmed their expression using real-time polymerase chain reaction. Expression of miRNA-155 was higher in SNK-6 and YKS cells than in normal NK cells. We next determined the levels of miRNA-155 in the serum of healthy individuals and NKTCL patients, and correlated its expression with clinical parameters and inflammatory factors detected using enzyme-linked immunoabsorbent assays. Levels of miRNA-155 were higher in NKTCL patients' serum than in serum from healthy individuals. miRNA-155 expression was higher in patients with stable or progressive disease (SD+PD) than in those with partial or complete remission (PR+CR). While further studies are needed to clarify the underlying molecular mechanisms, it appears miRNA-155 may be a molecular marker of NKTCL.

Expression of zinc finger E-box-binding homeobox factor 1 in epithelial ovarian cancer: A clinicopathological analysis of 238 patients.

A growing body of evidence indicates that aberrant activation of epithelial-to-mesenchymal transition (EMT) plays a key role in tumor cell invasion and metastasis. Zinc finger E-box-binding homeobox factor 1 (ZEB1), as a crucial mediator of EMT, contributes to the malignant progression of various epithelial tumors. To determine whether ZEB1 is involved in the progression of ovarian cancer, we immunohistochemically evaluated the expression of ZEB1 in 238 cases of epithelial ovarian cancer (EOC) and analyzed its associations with clinicopathological parameters. Positive expression of ZEB1 was observed in 32.8% (78/238) of EOCs and it was found to be significantly associated with advanced tumor stage (P=0.001). The survival analysis indicated that the expression of ZEB1 was associated with a poor 5-year progression-free survival (PFS) (P=0.021). A similar tendency was also observed between the expression of ZEB1 and 5-year overall survival, although it did not reach statistical significance (P=0.118). Moreover, the multivariate analysis demonstrated that ZEB1 expression was an independent risk factor for 5-year PFS in ovarian cancer. Taken together, our data provide evidence that ZEB1 may play a crucial role in promoting aggressive ovarian carcinoma progression. Therefore, ZEB1 may serve as an effectively predictive marker and a potential target for therapeutic intervention in EOC.

Patient-derived xenograft models of squamous cell carcinoma of the uterine cervix.

Patient-derived xenograft (PDX) models of cancer are considered to reflect the biology and treatment response of human tumors to a larger extent than xenograft models initiated from established cell lines. The characterization of a panel of four novel PDX models of cervical carcinoma of the uterine cervix is described in this communication. The outcome of treatment differed substantially among the donor patients, and the PDX models were found to mirror the histology, aggressiveness, and metastatic propensity of the donor patients' tumors. Two of the models (BK-12 and LA-19) were highly metastatic, one model (ED-15) was poorly metastatic, and one model (HL-16) was non-metastatic. The primary tumors of the two highly metastatic models showed high density of intratumoral lymphatics, whereas the other two models did not develop intratumoral lymphatics. The potential of the models to metastasize to lymph nodes was associated with high expression of both angiogenesis-related genes and cancer stem cell-related genes. The models may be highly valuable for studying mechanisms linking lymph node metastasis to lymphangiogenesis, hemangiogenesis, and the presence of cancer stem cells.

Nuclear, cytoplasmic, and stromal expression of ZEB1 in squamous and small cell carcinoma of the esophagus.

Zinc finger E-box-binding homeobox 1 (ZEB1) is a transcriptional factor known to repress E-cadherin promoter and thus induce EMT. Expression of ZEB-1 has in numerous cancers been associated with aggressive disease and poor clinical outcome. Our aim was to investigate the expression of ZEB1 in esophageal squamous- and small-cell carcinomas. Immunohistochemical staining was performed on tissue sections obtained from 151 patients with esophageal squamous cell carcinoma (ESCC) and 25 patients with primary small-cell carcinoma of the esophagus (PSCCE). Semi-quantitative analysis, and thus statistical analysis, has been accomplished on the samples. Immunohistochemistry revealed ZEB1 expression in the cytoplasm (64.9% of cases), in nuclei (11.3% of cases) and in tumor stroma (80.1% of cases) of ESCC. In PSCCE only nuclear staining (88.0% of cases) was observed. Weak cytoplasmic expression of ZEB1 in ESCC was associated with longer survival. Immunohistochemical evaluation of ZEB1 cytoplasmic expression in ESCC may have clinical prognostic value, but further studies are needed to fully understand the function as well as potential clinical and therapeutic implications of ZEB1 expression in cancers.

Decreased Expression of PDHE1α Predicts Worse Clinical Outcome in Esophageal Squamous Cell Carcinoma.

BACKGROUND: Aerobic glycolysis is an important hallmark of cancer cells. Deficiency of pyruvate dehydrogenase component α subunit (PDHE1α) leads to mitochondrial dysfunction and promotes glycolysis. Therefore, studies on the expression of PDHE1α in cancer are warranted. PATIENTS AND METHODS: The PDHE1α protein was immunohistochemically investigated and analyzed in 157 samples of surgically dissected esophageal squamous cell carcinoma (ESCC) and 21 'normal' esophageal epithelia tissues. RESULTS: PDHE1α protein expression was lower in ESCC. Absence of PDHE1α protein expression in tumor cells was found in 77.1% of cases. Negativity for PDHE1α protein in tumor cells was correlated with poor tumor differentiation (p<0.05) and shorter overall survival (p<0.05). CONCLUSION: Lack of PDHE1α protein expression in ESCC is associated with poor prognosis. Our findings also verify the existence of aerobic glycolysis switch in ESCC, and strongly advocate novel therapeutic strategies in reversing the Warburg effect in tumors.

The expression of aldehyde dehydrogenase 1 (ALDH1) in ovarian carcinomas and its clinicopathological associations: a retrospective study.

BACKGROUND: Aldehyde dehydrogenase 1 (ALDH1) is widely used as a specific cancer stem cell marker in a variety of cancers, and may become a promising target for cancer therapy. However, the role of its expression in tumor cells and the microenvironment in different cancers is still controversial. METHODS: To clarify the clinicopathological effect of ALDH1 expression in ovarian carcinoma, a series of 248 cases of paraffin-embedded formalin fixed ovarian carcinoma tissues with long term follow-up information were studied by immunohistochemistry. RESULTS: The immunostaining of ALDH1was variably detected in both tumor cells and the stromal cells, although the staining in tumor cells was not as strong as that in stromal cells. Statistical analyses showed that high ALDH1 expression in tumor cells was significantly associated with histological subtypes, early FIGO stage, well differentiation grade and better survival probability (p < 0.05). The expression of ALDH1 in the stromal cells had no clinicopathological associations in the present study (p > 0.05). CONCLUSIOMS: High expression of cancer stem cell marker ALDH1 in ovarian carcinoma cells may thus portend a favorable prognosis, but its expression in tumor microenvironment may have no role in tumor behavior of ovarian carcinomas. More studies are warranted to find out the mechanisms for this.

Mitochondrial DNA Deficiency in Ovarian Cancer Cells and Cancer Stem Cell-like Properties.

BACKGROUND: A low quantity of mitochondrial DNA (mtDNA) is a risk factor in a variety of tumor types. However, it is unclear how mtDNA reduction influences tumor behavior. MATERIAL AND METHODS: mtDNA-deficient ovarian cancer cells were established by ethidium bromide (EtBr) treatment with additive combination of pyruvate and uridine. RESULTS: The mtDNA-deficient cells had a low growth and colony-forming efficiency compared to the control cells. RNA sequencing revealed down-regulation of mitochondrion-related genes and up-regulation of genes related to cell proliferation and anti-apoptosis. The expression of genes involved in cancer metastasis, proliferation, angiogenesis, drug resistance and cancer cell stemness were also up-regulated. Intriguingly, cancer stem cell markers CD90 and CD117 were both up-regulated by EtBr dose-dependently in both cell lines. CONCLUSION: MtDNA deficiency may induce ovarian cancer stem cell-like properties through different ways in vitro, therefore contributing to different tumor behaviors.

Renalase protects the cardiomyocytes of Sprague-Dawley rats against ischemia and reperfusion injury by reducing myocardial cell necrosis and apoptosis.

BACKGROUND/AIMS: Renalase, a novel flavoprotein expressed in the kidney and heart, reduces renal tubular necrosis and apoptosis, which suggests that it might protect against necrosis and/or apoptosis in myocardial ischemia reperfusion injury (MIRI). The present study thus explored the effects of renalase on Sprague-Dawley (SD) rats subjected to MIRI. METHODS: We used Lentivirus-mediated RNA interference (RNAi) to inhibit the renalase gene expression in the heart tissue via pericardial cavity injection. The MIRI animal modal was established by blocking the left anterior descending artery for 45mins followed by 4h of reperfusion. Real-time PCR and western blotting were used to detect renalase expression in the heart tissue. Double staining and TUNEL were used to detect the necrosis and apoptosis in the myocardial cells, respectively. RESULTS: All rats subjected to MIRI exhibited lower levels of renalase in the heart tissue than did the sham-operated group (P<0.05, n=6). The (RNAi) group rats exhibited lower renalase levels than did the controls and also exhibited more serious necrosis (7.12±0.56% vs. 3.32±0.93%, P<0.05, n=6) and apoptosis (151.8±8.2% vs. 66.8±6.5%, P<0.05, n=6); however, pretreatment with the recombinant renalase significantly reduced myocardial cell necrosis (1.51±0.12% vs. 4.13±0.02%, P<0.05, n=6) and apoptosis (21.3±5.0% vs. 52.6±10.4%, P<0.05, n=6) relative to the control rats. CONCLUSIONS: Exogenous recombinant renalase protein reduced myocardial cell necrosis and apoptosis. Recombinant renalase protein might be a new cardiovascular drug for ischemia/IR injury.

[A case of repeated unconsciousness caused by excrescence locking bicuspid aortic valve].

We received a patient who repeated unconsciousness due to excrescence locking bicuspid aortic valve. He experienced unconsciousness and treatments with anti infection, surgical operation, valve replacement and recovery. It was a rare case, which made us realize that the heart color Doppler ultrasound should be regularly performed in patients with aortic valve abnormalities. Once patients were found to have infective endocarditis combined with the valve vegetations, they should be formally treated as soon as possible.