Vascular control by infrarenal aortic cross-clamping in placenta accreta spectrum disorders: description of technique.

We describe a novel surgical technique in 31 women with histopathologically confirmed placenta accreta spectrum (PAS) disorders managed by a multidisciplinary team using a prophylactic infrarenal abdominal aortic cross-clamping technique during caesarean hysterectomy. We conclude that this new surgical procedure is a relatively safe technique to potentially control operative blood loss. Our work may stimulate others to develop protocols assessing this innovative technique to improve the surgical outcome of PAS disorders.

Decreased expression of H3K27me3 in human ovarian carcinomas correlates with more aggressive tumor behavior and poor patient survival.

It has been confirmed that trimethylation of lysine 27 on histone H3 (H3K27me3) plays an important role in epigenetic process of tumorigenesis. However, the status of H3K27me3 in ovarian cancer and its impact on patients' clinicopathologic characteristics and prognosis are unclear. In the present study, the immunohistochemistry (IHC) was utilized to detect protein expression of H3K27me3 in 12 normal ovaries, 26 ovarian cystadenomas, 31 borderline ovarian tumors and 168 ovarian carcinomas by tissue microarray. The association between H3K27me3 expression with clinicopathologic features and patient prognosis were also evaluated using various statistical models. The expression of H3K27me3 was decreased in 2 of 12 (16.7%) cases of the normal ovaries, 8 of 26 (30.8%) cases of cystadenomas, 12 of 31 (38.7%) cases of borderline ovarian tumors, and 93 of 168 (55.4%) cases of primary ovarian carcinomas, respectively (P<0.05). Further correlation analysis suggested that decreased expression of H3K27me3 in ovarian carcinomas was significantly correlated with more advanced pM and FIGO stages (P<0.05). In addition, a significant association between decreased expression of H3K27me3 and shortened patient survival (mean 66 months versus 101 months, p=0.019) was demonstrated by univariate survival analysis of the ovarian carcinoma cohorts. Importantly, H3K27me3 expression provided a significant independent prognostic factor in multivariate analysis (p=0.028). These findings confirmed that decreased expression of H3K27me3 in primary ovarian cancer might be correlated with the acquisition of an invasive and/or aggressive phenotype of tumor, and might serve as an independent biomarker for poor prognosis in patients with ovarian carcinoma.

Oncogenic miR-20a and miR-106a enhance the invasiveness of human glioma stem cells by directly targeting TIMP-2.

Emerging evidence has shown that cancer stem cells (CSCs) are the cellular determinants to promote cancer invasion and metastasis. However, the mechanism underlying CSC invasion remains unknown. MicroRNAs are evolutionally conserved small noncoding RNAs that are critical for the regulation of gene expression, and their expressions are often dysregulated in cancers. In the present study, we demonstrated that two functionally related microRNAs, miR-20a and -106a (miR-20a/106a), were capable of enhancing the invasiveness of CD133(+) glioma stem cells (GSCs) isolated from both glioblastoma cell line U87 and primary human glioma specimens. We found that the level of miR-20a/106a in GSCs was significantly higher than that in the committed CD133(-) glioma cells, and correlated with the invasive capability of GSCs. By bioinformatic analysis, we identified tissue inhibitor of metalloproteinases-2 (TIMP-2) as one of the miR-20a/106a-targeted genes. TIMP-2 level correlated inversely with miR-20/106 expression. Directly targeting by miR-20a/106a on 3'-untranslation region (3'-UTR) of TIMP-2 mRNA was confirmed by 3'-UTR dual-luciferase reporter assay. Knockdown of miR-20a/106a in GSCs increased endogenous TIMP-2 protein abundance, thereby inhibiting GSC invasion. We also found that Nordy, a synthetic lipoxygenase inhibitor, inhibited GSC invasiveness by elevating the expression of TIMP-2 via downregulation of miR-20a/106a. Our results indicate that miR-20a/106a has a key role in GSC invasion and may serve as targets for treatment of glioblastoma.

MiR-29c mediates epithelial-to-mesenchymal transition in human colorectal carcinoma metastasis via PTP4A and GNA13 regulation of ß-catenin signaling.

BACKGROUND: Distant metastasis is the major cause of cancer-related death, and epithelial-to-mesenchymal transition (EMT) has a critical role in this process. Accumulating evidence indicates that EMT can be regulated by microRNAs (miRNAs). miR-29c has been implicated as a tumor suppressor in several human cancers. However, the role of miR-29c in the progression of colorectal cancer (CRC) metastasis remains largely unknown. PATIENTS AND METHODS: The expression of miR-29c was examined by qRT-PCR in a cohort of primary CRC (PC) and distant liver metastasis (LM) tissues. A series of in vivo and in vitro assays were carried out in order to elucidate the functions of miR-29c and the molecular mechanisms underlying the pathogenesis of metastatic CRC. RESULTS: miR-29c was markedly downregulated in PCs with distant metastasis and determined to be an independent predictor of shortened patient survival. But LM tissues showed higher levels of miR-29c than that in PC tissues. In CRC cells, miR-29c dramatically suppressed cell migration and invasion abilities in vitro and cancer metastasis in vivo. In addition, miR-29c inhibited EMT and negatively regulated Wnt/ß-catenin signaling pathway. Guanine nucleotide binding protein alpha13 (GNA13) and protein tyrosine phosphatase type IVA (PTP4A) were identified as direct targets of miR-29c, which acted through ERK/GSK3ß/ß-catenin and AKT/GSK3ß/ß-catenin pathways, respectively, to regulate EMT. Furthermore, significant associations between miR-29c, its target genes (GNA13 and PTP4A) and EMT markers were validated in both PC and LM tissues. CONCLUSION: Our findings highlight the important role of miR-29c in regulating CRC EMT via GSK-3ß/ß-catenin signaling by targeting GNA13 and PTP4A and provide new insights into the metastatic basis of CRC.

HGFK1 inhibits bone metastasis in breast cancer through the TAK1/p38 MAPK signaling pathway.

Breast cancer metastasis to bone represents a devastating complication of advanced breast cancer, frequently resulting in significant increases in morbidity and mortality. An understanding of the mechanisms that govern breast cancer metastasis at the molecular level should lead to more effective therapies. Recently, the kringle 1 domain of human hepatocyte growth factor (HGFK1) was identified as a candidate metastasis suppressor gene. Here, we investigated whether HGFK1 is a key regulator of breast cancer bone metastasis. Of the 193 human breast carcinoma tissue samples examined, HGFK1 expression was relative higher in 82 (42.4%) by western blot and in 84 (43.5%) by quantitative real-time PCR. The higher expression of HGFK1 was significantly associated with a better prognostic value (P<0.001) and inversely correlated with bone metastasis (P=0.003). The efficacy of adeno-associated virus carrying HGFK1 (AAV-HGFK1) in osteolytic bone metastasis was then evaluated using an in vivo bone metastasis model. AAV-HGFK1 significantly inhibited osteolytic bone metastasis and prolonged the survival of mice in this model (P<0.01). In vitro, HGFK1 expression resulted in significant anti-invasion effects, enhanced the phosphorylation of TAK1 (transforming growth factor-ß-activated kinase 1), p38 MAPK (mitogen-activated protein kinase) and MAPKAPK2 (MAPK-activated protein kinase 2) and decreased the expression of receptor activator of nuclear factor-κB (RANK), which was abrogated by the p38 MAPK inhibitor SB203580. This study shows for the first time that HGFK1 significantly inhibits the metastasis of breast cancer to bone by activating the TAK1/p38 MAPK signaling pathway and inhibiting RANK expression. Thus, AAV-HGFK1 treatment represents a potential therapy for bone metastasis in breast cancer.

Epigenetic inactivation of paired box gene 5, a novel tumor suppressor gene, through direct upregulation of p53 is associated with prognosis in gastric cancer patients.

Using genome-wide methylation screening, we identified that paired box gene 5 (PAX5) is involved in human cancer development. However, the function of PAX5 in gastric cancer (GC) development is largely unclear. We analyzed its epigenetic inactivation, biological functions and clinical application in GC. PAX5 was silenced in seven out of eight GC cell lines. A significant downregulation was also detected in paired gastric tumors compared with adjacent non-cancerous tissues. The downregulation of PAX5 was closely linked to the promoter hypermethylation status and could be restored with demethylation treatment. Ectopic expression of PAX5 in silenced GC cell lines (AGS and BGC823) inhibited colony formation and cell viability, arrested cell cycle, induced apoptosis, suppressed cell migration and invasion and repressed tumorigenicity in nude mice. Consistent with the induction of apoptosis by PAX5 in vitro, terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) staining showed significantly enhanced apoptotic cells in PAX5-expressed tumors compared with the vector control tumors. On the other hand, knockdown of PAX5 by PAX5-short hairpin RNA increased the cell viability and proliferation. The anti-tumorigenic function of PAX5 was revealed to be mediated by upregulating downstream targets of tumor protein 53 (p53), p21, BCL2-associated X protein, metastasis suppressor 1 and tissue inhibitors of metalloproteinase 1, and downregulating BCL2, cyclin D1, mesenchymal-epithelial transition factor (MET) and matrix metalloproteinase 1. Immunoprecipitation assay demonstrated that PAX5 directly bound to the promoters of p53 and MET. Moreover, PAX5 hypermethylation was detected in 77% (144 of 187) of primary GCs compared with 10.5% (2/19) of normal gastric tissues (P<0.0001). GC patients with PAX5 methylation had a significant poor survival compared with the unmethylated cases as demonstrated by Cox regression model and log-rank test. In conclusion, PAX5 is a novel functional tumor suppressor in gastric carcinogenesis. Detection of methylated PAX5 can be utilized as an independent prognostic factor in GC.

Anthracyclines disrupt telomere maintenance by telomerase through inducing PinX1 ubiquitination and degradation.

Telomere maintenance is essential for cancer growth. Induction of telomere dysfunction, for example, by inhibition of telomeric proteins or telomerase, has been shown to strongly enhance cancer cells' sensitivity to chemotherapies. However, it is not clear whether modulations of telomere maintenance constitute cancer cellular responses to chemotherapies. Furthermore, the manner in which anti-cancer drugs affect telomere function remains unknown. In this study, we show that anthracyclines, a class of anti-cancer drugs widely used in clinical cancer treatments, have an active role in triggering telomere dysfunction specifically in telomerase-positive cancer cells. Anthracyclines interrupt telomere maintenance by telomerase through the downregulation of PinX1, a protein factor responsible for targeting telomerase onto telomeres, thereby inhibiting telomerase association with telomeres. We further demonstrate that anthracyclines downregulate PinX1 by inducing this protein degradation through the ubiquitin-proteasome-dependent pathway. Our data not only reveal a novel action for anthracyclines as telomerase functional inhibitors but also provide a clue for the development of novel anti-cancer drugs based on telomerase/telomere targeting, which is actively investigated by many current studies.

EZH2 supports nasopharyngeal carcinoma cell aggressiveness by forming a co-repressor complex with HDAC1/HDAC2 and Snail to inhibit E-cadherin.

The enhancer of zeste homolog 2 (EZH2) is upregulated and has an oncogenic role in several types of human cancer. However, the abnormalities of EZH2 and its underlying mechanisms in the pathogenesis of nasopharyngeal carcinoma (NPC) remain unknown. In this study, we found that high expression of EZH2 in NPC was associated closely with an aggressive and/or poor prognostic phenotype (P<0.05). In NPC cell lines, knockdown of EZH2 by short hairpin RNA was sufficient to inhibit cell invasiveness/metastasis both in vitro and in vivo, whereas ectopic overexpression of EZH2 supported NPC cell invasive capacity with a decreased expression of E-cadherin. In addition, ablation of endogenous Snail in NPC cells virtually totally prevented the repressive activity of EZH2 to E-cadherin, indicating that Snail might be a predominant mediator of EZH2 to suppress E-cadherin. Furthermore, co-immunoprecipitation (IP), chromatin IP and luciferase reporter assays demonstrated that in NPC cells, (1) EZH2 interacted with HDAC1/HDAC2 and Snail to form a repressive complex; (2) these components interact in a linear fashion, not in a triangular fashion, that is, HDAC1 or HDAC2 bridge the interaction between EZH2 and Snail; and (3) the EZH2/HDAC1/2/Snail complex could closely bind to the E-cadherin promoter by Snail, but not YY1, to repress E-cadherin. The data provided in this report suggest a critical role of EZH2 in the control of cell invasion and/or metastasis by forming a co-repressor complex with HDAC1/HDAC2/Snail to repress E-cadherin, an activity that might be responsible, at least in part, for the development and/or progression of human NPCs.

Localization of 5-HT1A and 5-HT2A positive cells in the brainstems of control age-matched and Alzheimer individuals.

Serotonin receptor 1A and 2A positive cells in postmortem brainstems were demonstrated via immunohistochemistry in eight control age-matched elderly individuals and eight Alzheimer patients. The 5-HT1A positive cells were found in substantia nigra, pontile nucleus, and vagal as well as dorsal raphe nucleus, while 5-HT2A receptor positive cells were found in motor, sensory and spinal trigeminal nuclei, pontile nucleus, substantia nigra, and nucleus solitarius. A comparison in density of positive cells per unit area was made between control age-matched and Alzheimer individuals. Statistically significant differences (p ≤ 0.01) in density were observed in 5-HT1A cells in pontile, dorsal raphe, and vagal nuclei between control age-matched and Alzheimer, and in 5-HT2A positive cells in the sensory trigeminal nucleus, between control and Alzheimer. This de novo study indicated the presence of 5-HT1A and 5-HT2A receptor positive cells in the above nuclei of human brainstem and revealed differences in density between control age-matched and Alzheimer, indicating possible functional derangements in Alzheimer patients in these areas. In addition, colocalization studies indicated that 5-HT1A receptors were in cholinergic cells and gamma-aminobutyric acid positive fibers were linked to 5-HT2A receptor positive cells. It is hoped that understanding these two important 5-HT receptors and their localization might lead to advances in future therapeutic development.

Low expression of Mel-18 predicts poor prognosis in patients with breast cancer.

BACKGROUND: Our previous study suggested that melanoma nuclear protein 18 (Mel-18) acted as a tumor suppressor in human breast cancer. This study was designed to investigate the clinical and prognostic significance of Mel-18 in breast cancer patients. PATIENTS AND METHODS: Mel-18 was detected by immunohistochemistry in paraffin-embedded tissues from 287 breast cancer patients, of which 287 were from primary cancer sites, 63 from matched adjacent noncancerous sites, and 35 from metastatic lymph nodes. Differences in Mel-18 expression and clinical characteristics were compared by χ² test. Prognostic outcomes correlated with Mel-18 were examined using Kaplan-Meier analysis and Cox proportional hazards model. RESULTS: The decreased Mel-18 expression is incremental depending upon the magnitude of cancer progression (P < 0.001). Mel-18 was conversely correlated with the pathological classifications (P < 0.001 for T, N, and M classifications, respectively), clinical staging (P < 0.001), and progesterone receptor (P = 0.030). Furthermore, patients with higher level of Mel-18 showed prolonged overall survivals (P < 0.001). The diminished Mel-18 expression may be a risk factor for the patients' survival (P < 0.001). CONCLUSIONS: Lower Mel-18 expression is correlated with advanced clinicopathologic classifications and a poor overall survival in breast cancer patients. These findings suggest that Mel-18 may serve as a useful marker in prognostic evaluation for patients.

Overexpression of AIB1 negatively affects survival of surgically resected non-small-cell lung cancer patients.

BACKGROUND: The amplified in breast cancer 1 (AIB1) gene has been considered to play an oncogenic role in human cancers, but its clinical/prognostic significance in non-small-cell lung cancer (NSCLC) is still unclear. PATIENTS AND METHODS: The methods of immunohistochemistry and FISH were utilized to examine protein expression and amplification of AIB1 in 230 informative surgically resected NSCLCs and in 30 samples of normal lung tissues. RESULTS: Overexpression and amplification of AIB1 were found in 48.3% and 8.2% of NSCLCs, respectively. AIB1 overexpression was associated with AIB1 gene amplification and cell proliferation but not related to estrogen receptor (ER)-alpha, ER-beta, progesterone receptor or androgen receptor status. A positive correlation between AIB1 overexpression and an ascending pathologic node stage in lung adenocarcinoma (ADC) was observed (P = 0.043). Univariate survival analysis demonstrated a significant association of AIB1 overexpression with shortened patient survival, especially for those with stage III disease (P < 0.001). Importantly, AIB1 expression was evaluated as the most significant predictor for survival in multivariate analysis (hazards ratio = 2.069, P < 0.001). CONCLUSION: Overexpression of AIB1 might provide a selective advantage for lymph node metastasis of lung ADC and serve as a useful biomarker for poor prognosis for NSCLC patients.

Development of recombinant adeno-associated virus and adenovirus cocktail system for efficient hTERTC27 polypeptide-mediated cancer gene therapy.

The low in vivo transduction efficiency of recombinant adeno-associated virus (rAAV) and the undesirably strong immunogenicity of adenovirus (rAdv) have limited their clinical utilization in cancer gene therapy. We have previously demonstrated that intratumoral injection of rAAV expressing a C-terminal polypeptide of human telomerase reverse transcriptase (rAAV-hTERTC27) effectively inhibits the growth of glioblastoma xenografts in nude mice. To further improve its efficacy, we combined rAAV-hTERTC27 with rAdv and investigated the efficiency of the cocktail vectors in vivo. At a nontherapeutic dose (1 x 10(8) plaque-forming units (PFUs)), rAdv-null and rAdv-hTERTC27 were equipotent in enhancing the therapeutic efficacy of rAAV-hTERTC27 (1.5 x 10(11) v.g.), and complete tumor regression was achieved in 25% of the treated animals. Importantly, the combination of rAAV-hTERTC27 and a therapeutic dose (2.5 x 10(9) PFU) of rAdv-hTERTC27 significantly augmented the therapeutic effects and led to a 38% complete tumor regression rate. In vivo optical imaging also showed that rAAV-luc/rAdv-luc cocktail vectors could synergistically enhance the early transient and latent sustained expression of luciferase, as compared to rAdv-luc and rAAV-luc alone. These findings suggest that the combination of rAAV-hTERTC27 and a therapeutic dose of rAdv-hTERTC27 is potentially a promising treatment for glioblastoma, and the rAAV/rAdv cocktail vector system warrants further development for cancer gene therapy.

Screening of active ingredients of herbal medicine for interaction with CYP450 3A4.

A two-step algorithm is adopted in the screening of herbal species which possess significant inhibitory effects on cytochrome P450 3A4 (CYP450 3A4). The algorithm comprises an initial stage of high throughput screening with Herbochip for the identification of herbal fractions that exhibit interactions with CYP450 3A4. Fifty commonly used TCM species were screened with seven showing a positive signal reflecting interaction. In the inhibition assays that followed, six of the seven species gave a signal. Sophora flavescens stood out as it gave the highest number of wells with a response, the highest maximum index was 0.96, and the median index was 0.55. The selection of TCM species with inhibitory effects on CYP450 carries the potential role of its use to boost the effects of known therapeutic agents, a mechanism that has been exploited in the design of regimens for the treatment of HIV infection.

A novel glioblastoma cancer gene therapy using AAV-mediated long-term expression of human TERT C-terminal polypeptide.

Glioblastoma multiforme is the most aggressive form of human brain tumor, which has no effective cure. Previously, we have demonstrated that overexpression of the C-terminal fragment of the human telomerase reverse transcriptase (hTERTC27) inhibits the growth and tumorigenicity of human cervical cancer HeLa cells. In this study, the therapeutic effect and molecular mechanisms of hTERTC27-mediated cancer gene therapy were further explored in vivo in established human glioblastoma xenografts in nude mice. We showed that intratumoral injection of adeno-associated virus carrying hTERTC27 (rAAV-hTERTC27) is highly effective in reducing the growth of the subcutaneously transplanted glioblastoma tumors. Histological analyses showed that rAAV-hTERTC27 treatment leads to profound necrosis, apoptosis, infiltration of polymorphonuclear neutrophils and reduced microvessel density in the tumor samples. To study the molecular mechanism of rAAV-hTERTC27-mediated antitumor effects, we analyzed the global gene expression profiles of the rAAV-hTERTC27-treated tumor tissues and cell line as compared with that of the control rAAV-green fluorescent protein-treated samples by DNA microarray. Our results suggest that hTERTC27 exerts its effect through complex mechanisms, which involve genes regulating apoptosis, cell adhesion, cell cycle, immune responses, metabolism, signal transduction, transport, transcription and telomere maintenance.

Behavioral and biochemical studies of total furocoumarins from seeds of Psoralea corylifolia in the chronic mild stress model of depression in mice.

Depression is related to alterations of the monoamine oxidase (MAO), hypothalamic-pituitary-adrenal (HPA) axis, and oxidative systems, and some antidepressants achieve their therapeutic effects through alteration of following biochemical markers of depression: MAO-A and MAO-B activities, cortisol levels, superoxide dismutase (SOD) activity and malondialdehyde (MDA) levels. The seeds of Psoralea corylifolia, otherwise known as Buguzhi, have long been used for treatments of various symptoms associated with aging in China. Furocoumarins are the most widespread secondary metabolites in this species. The present study was designed to evaluate the potential antidepressant-like activity of total furocoumarins of P. corylifolia (TFPC) in the chronic mild stress (CMS) model of depression. Mice subjected to CMS exhibited a reduction in sucrose intake. Conversely, brain MAO-A and MAO-B activities, plasma cortisol levels, and liver SOD activity and MDA levels were increased following CMS exposures. The time-course for reversal of CMS-induced deficits in sucrose consumption by TFPC was dose-dependent. Thus, the statistically significant effect of the higher dose of TFPC (50 mg/kg body wt.) was observed after 3 days of treatment, while 6 days of treatment were required in the group receiving a lower dose (30 mg/kg body wt.) of TFPC. TFPC reversed these biochemical changes. These results suggest that TFPC may possess potent and rapid antidepressant properties that are mediated via MAO, the HPA axis and oxidative systems and these antidepressant actions could make TFPC a potentially valuable drug for the treatment of depression in the elderly.

Serum proteomic patterns for gastric lesions as revealed by SELDI mass spectrometry.

SELDI mass spectrometry was used to investigate protein expression in sera of patients with gastric cancer and gastritis compared to normal volunteers. Differences in peak morphology and intensity were observed in regions of 5910 Da, 5084 Da, 6640 Da and 8691 Da. Patients with gastric cancer exhibited an up-regulation of the 5910 Da peak and a down-regulation of the 8691 Da peak compared to the healthy volunteers; there was also some bi-partitioning and tri-partitioning at the 5084 Da peak. When comparing the sera of these cancer patients with those of gastritis, the former had an up-regulation of the 5910 Da peak and a down-regulation of the 6640 Da peak. This is the first report showing that SELDI sera analysis may be useful in the screening of gastric lesions.