Ubiquitin-specific peptidase 15 regulates the TFAP4/PCGF1 axis facilitating liver metastasis of colorectal cancer and cell stemness.

The tumor recurrence and metastasis of colorectal cancer (CRC) are responsible for most of CRC-linked mortalities. It is an urgent need to deeply investigate the pathogenesis of CRC metastasis and look for novel targets for its treatment. The current study aimed to investigate the effects of ubiquitin-specific peptidase 15 (USP-15) on the CRC progression. In vivo, a mouse model of liver metastasis of CRC tumor was established to investigate the role of USP-15. In vitro, the migrated and invasive abilities of CRC cells were assessed by transwell assay. Cell stemness was evaluated by using sphere formation assay. The underlying mechanism was further explored by employing the co-immunoprecipitation, dual luciferase reporter assay, oligonucleotide pull-down assay, and chromatin immunoprecipitation assay. The results showed that USP-15 was upregulated in CRC patients with liver metastasis and high metastatic potential cell lines of CRC. Loss of USP-15 repressed the epithelial-to-mesenchymal transition (EMT), migration, invasion, and stemness properties of CRC cells in vitro. Downregulation of USP-15 reduced the liver metastasis of mice in vivo. USP-15 upregulation obtained the contrary effects. Subsequently, USP-15 deubiquitinated transcription factor AP-4 (TFAP4) and enhanced its protein stability. TFAP4 could transcriptionally activated polycomb group ring finger 1 (PCGF1). The pro-cancer effects of USP-15 were rescue by the knockdown of TFAP4 or PCGF1. In conclusions: USP-15 facilitated the liver metastasis by the enhancement of cell stemness and EMT in CRC, which was at least partly mediated by the deubiquitination of TFAP4 upon the upregulation of PCGF1.

Tumor budding and the prognosis of patients with metastatic colorectal cancer: a meta-analysis.

PURPOSE: Tumor budding has been suggested to be associated with poor survival of patients with colorectal cancer (CRC). However, it is unclear whether the association remains in patients with metastatic CRC (mCRC). The aim of the systematic review and meta-analysis was to investigate the potential predictive role of tumor budding for the prognosis of patients with mCRC. METHODS: PubMed, Embase, Cochrane Library, and Web of Science were searched for relevant observational studies comparing the survival outcomes between mCRC patients with high versus low tumor budding. Data collection, literature searching, and statistical analysis were conducted independently by two authors. Using a heterogeneity-incorporating random-effects model, the results were pooled. RESULTS: In this meta-analysis, 1503 patients from nine retrospective cohort studies were included. Pooled results showed that compared to those with low tumor budding, mCRC patients with high tumor budding were associated with a poor progression-free survival (hazard ratio (HR), 1.65; 95% confidence interval (CI), 1.31 to 2.07, p < 0.001; I2 = 30%) and overall survival (HR, 1.60; 95% CI, 1.33 to 1.93; p < 0.001; I2 = 0%). Influencing analysis by excluding one study at a time showed consistent results (p all < 0.05). Subgroup analyses showed consistent results in tumor budding evaluated from the primary cancer and metastases, in studies with a high tumor budding defined as ≥ 10 or 15 and ≥ 5 buds/HPF and in studies analyzed with univariate and multivariate regression models (p for subgroup difference all > 0.05). CONCLUSION: A high-degree tumor budding may be associated with poor prognosis in patients with mCRC.

Karyopherin Subunit Alpha 1 Enhances the Malignant Behaviors of Colon Cancer Cells via Promoting Nuclear Factor-κB p65 Nuclear Translocation.

BACKGROUND/AIMS: Aberrant nuclear factor-κB p65 (NF-κB p65) nuclear import commonly occurs in multiple cancers, including colon cancer. According to BioGRID, we noted that Karyopherin subunit alpha 1 (KPNA1), an important molecular transporter between the nucleus and the cytoplasm, may interact with NF-κB p65. KPNA1 itself is highly expressed in colon adenocarcinoma samples (N = 286) based on The Cancer Genome Atlas (TCGA) database. We aimed to explore the role of KPNA1 in colonic carcinogenesis and to determine whether NF-κB p65 nuclear translocation was involved. METHODS: KPNA1 expressions at mRNA and protein levels were analyzed in colon cancer tissues. The regulatory effect of KPNA1 on malignant biological properties was detected in SW480 and HCT116 colon cancer cells. Coimmunoprecipitation and immunofluorescence were performed to verify the relationship between KPNA1 and NF-κB p65. KPNA1 ubiquitination was also preliminarily investigated. RESULTS: KPNA1 was firstly confirmed as a significantly upregulated gene in our collected clinical colon cancer samples (N = 35). KPNA1 depletion inhibited cell proliferation, induced cell cycle arrest, and diminished migratory and invasive capacity of SW480 and HCT116 cells. Colon cancer cells overexpressing KPNA1 acquired more aggressive behaviors. KPNA1 acted as a transporter to induce the nuclear accumulation of NF-κB p65, thereby activating NF-κB signaling pathway in colon cancer cells. Furthermore, HECT, C2, and WW Domain-Containing E3 Ubiquitin (HECW2) interacted with KPNA1 to induce its ubiquitination. KPNA1 labeled with polyubiquitins was degraded through ubiquitin-proteasome system. CONCLUSION: The present study uncovers a role of KPNA1-NF-κB p65 axis in promoting colonic carcinogenesis.

MORC4 plays a tumor-promoting role in colorectal cancer via regulating PCGF1/CDKN1A axis in vitro and in vivo.

MORC family CW-type zinc finger 4 (MORC4) possessing nuclear matrix binding domains has been observed to be involved in multiple cancer development. By digging three gene expression omnibus (GEO) gene microarrays (GSE110223, GSE110224 and GSE24514), we found that MORC4 was overexpressed in colorectal cancer (CRC) samples (log2 Fold change >1, p < 0.05). We aimed to investigate the role of MORC4 in CRC malignant behaviors, with an emphasis on polycomb group ring finger 1 (PCGF1)/cyclin-dependent kinase inhibitor 1A (CDKN1A) axis. Firstly, we confirmed MORC4 as an upregulated gene in 60 pairs of frozen CRC and adjacent normal samples. MORC4 overexpression increased proliferation and metastasis, and decreased apoptosis in SW480 and HT29 cells, which was diminished by the knockdown of PCGF1, a transcriptional repressor of CDKN1A (a potent cyclin-dependent kinase inhibitor). MORC4 was further identified as a novel molecule that interacted with PCGF1 via coimmunoprecipitation. MORC4 itself did not substantially suppress CDKN1A transcriptional activity, but it augmented PCGF1's effect on CDKN1A. Additionally, MORC4 acted as the substrate of HECT, C2, and WW domain-containing E3 ubiquitin protein ligase 2 (HECW2) and was degraded through ubiquitin-proteasome system. Collectively, our work suggested that MORC4 accelerated CRC progression via governing PCGF1/CDKN1A signaling.

Adeno-Associated Virus-Mediated Knockdown of Agmatinase Attenuates Inflammation and Tumorigenesis in a Mouse Model of Colitis-Associated Colorectal Cancer.

Agmatinase (AGMAT) is an enzyme that hydrolyzes agmatine to putrescine and urea. In this study, we explored the functions of AGMAT in colorectal cancer (CRC). By performing gain-of-function and loss-of-function experiments, we investigated the roles of AGMAT in proliferation, cell cycle progression, and apoptosis of CRC cells. We also established a colitis-associated colorectal cancer model by challenging mice with azoxymethane (AOM) and dextran sodium sulfate (DSS), and we subsequently silenced AGMAT expression in mice by adeno-associated virus 9 (AAV9)-mediated delivery of short hairpin RNA (shRNA). In vitro experiments showed that overexpression of AGMAT accelerated the proliferation and inhibited the apoptosis of CRC cells, and AGMAT knockdown exhibited the opposite effects. Interestingly, the oncogenic transcription factor, c-Myc, could bind to the AGMAT promoter and transcriptionally increase AGMAT expression in CRC cells. Additionally, c-Myc and AGMAT were upregulated in the colon of AOM/DSS-treated mice, and AGMAT silencing significantly mitigated colitis in AOM/DSS-treated mice, as evidenced by the increased colon length, attenuated crypt damage, and reduced levels of inflammatory indicators (myeloperoxidase, interleukin-6, tumor necrosis factor-α, inducible nitric oxide synthase, and phosphorylated p65) in colon tissues. Notably, AGMAT silencing decreased both the number and size of tumors, reduced expression of proliferating cell nuclear antigen, and inhibited phosphorylation of protein kinase B (AKT) and extracellular signal-regulated kinase in the colon of AOM/DSS-treated mice. Overall, we determined that AGMAT facilitates tumor progression in CRC. Our findings will be helpful in the search for potential therapeutic targets for CRC.

[Molecular mechanism of locus coeruleus involved in acupuncture anti-myocardial ischemia based on transcriptome sequencing].

OBJECTIVE: To explore the molecular mechanism of locus coeruleus（LC） involved in electroacupuncture (EA) anti myocardial ischemia. METHODS: Twenty-four SD rats were randomly divided into sham-operation, model, EA and EA +lesion groups, with 6 rats in each group. The acute myocardial ischemia (AMI) model was established by ligation of the left anterior descending branch of coronary artery. EA (2 Hz/15 Hz, 1 mA) was applied to bilateral "Shenmen" (HT7) -"Tongli" (HT5) and the middle-point between HT7 and HT5 for 30 min, once daily for 3 days. For rats of the EA +lesion group, the virus (300 nL) was injected into bilateral LC before EA treatment. Serum aspartate aminotransferase (AST) was detected by ELISA. The gene expression profiles of rat heart were detected by transcriptome sequencing, the differentially expressed genes were screened, and Gene Ontology (GO) functional classification and Kyoto Encyclopedia of genes and genomes (KEGG) metabolic pathway enrichment analysis were performed. RESULTS: Compared with the sham-operation group, serum AST content was significantly increased in the model group (P<0.01). Following the intervention, serum AST was significantly reduced in the EA group (P<0.01), while the serum AST in the EA + lesion group was significantly higher compared with the EA group (P<0.05). Differential expression analysis showed that 1 138 differentially expressed genes were screened out between the model group and the sham-operation group, 1 330 differentially expressed genes between model and EA group, and 804 differentially expressed genes between EA and EA + lesion group. Among them, 218 differential genes were involved in the regulation of EA anti-myocardial ischemia in LC. GO functional classification analysis showed that these differentially expressed genes mainly involved in cell processes, metabolic processes and biological regulation in biological processes. KEGG pathway analysis showed that these differentially expressed genes were enriched in sulfur relay system, thiamine metabolism, glutathione metabolism, C5 branch dicarboxylic acid metabolism, cell adhesion molecules and Th1 and Th2 cell differentiation. CONCLUSION: EA intervention has a positive effect in anti-myocardial ischemia, which may be related to the sulfur relay system, thiamine metabolism, glutathione metabolism, C5 branch dicarboxylic acid metabolism, cell adhesion molecules and Th1 and Th2 cell differentiation involved in LC.

[Analysis of "Tianyou (TE 16) and five regions" in the human body].

Focusing on the original text record in Huangdi Neijing (Inner Canon of Yellow Emperor), the relevant theories of "Tianyou (TE 16) and five regions" are explored, e.g. acupoint names, meridians and acupoint features, and the clinical application of these acupoints has been analyzed. It is discovered that "Tianyou (TE 16) and five regions" are mainly used in treatment of the disorders in the nervous system, five sensory organs and motor system. Besides, in terms of the relevant theories, "Tianyou (TE 16) and five regions" has been compared with "root and knot" and "twelve divergent meridians". It is found that "Tianyou (TE 16) and five regions" communicates the externally-internally related meridians and is applicable in treatment of the disorders with both exterior and interior involved. It is the essential acupoint composition of the human body.

[Clinical analysis for fixation failure of 13 patients with intertrochanteric fractures].

OBJECTIVE: To analysis the reasons of fixation failure for intertrochanteric fractures, so as to select correct operation indications and fixation methods. METHODS: Retrospective analysis the clinical data of 13 patients with failed internal fixation of intertrochanteric fractures from September 1997 to September 2008, and the failure reasons were summarized. There were 7 males and 6 females,ranging in age from 58 to 93 years,averaged 71 years. Two patients were treated with intramedullary fixation, 4 patients with anatomical proximal femoral plate, 3 patients with DHS fixation, 2 patients with hollow compression screws, and 2 patients with external fixation. According to Evans types: 1 patient was type II, 7 patients were type III, and 5 patients were type IV. RESULTS: Eight patients with unstable fractures and malreduction had no grafted bone. Six patients had bad position of neck screws in the femur neck. Postoperative collodiaphyseal angle: 3 patients were under 90 degree, 7 patients 90 to 110 degree, and 3 patients 110 to 130 degree. Five patients had internal fixed screw exited, 6 patients had neck screws cutting to superior lateral, 3 patients had early weight bearing, and 10 patients were osteoporosis occurred after operation from 6 weeks to 11 months, averaged 4.5 months. CONCLUSION: The fixation failure of intertrochanteric fractures was concerned with fractures types, reduction, fixation methods, osteoporosis and the time of weight bearing.

[Studies on HPLC-fingerprint of Rhizoma Fagopyri Dibotoryis].

OBJECTIVE: To establish an HPLC fingerprint of Rhizoma fagopyri dibotoryis. METHOD: The HPLC-electrochemical detection assay was used to establish the fingerprint of Rhizoma fagopyri dibotoryis. The sample was performed on a column of Diamonsil C18 (4.6 mm x 250 mm, 5 microm) which was eluted with methanol- 0.1 mol x L(-1) phosphate buffer (pH 2.5), the flow rate was 1.0 mL x min(-1), the column temperature was 35 degrees C, the reference electrode was ISAAC (in-situ silver/silver chloride), the work electrode was glassy carbon, the counter electrode was Pt platmun. RESULT: The HPLC fingerprint profiles of 6 Rhizoma fagopyri dibotoryis contains 6 common chromatographic peaks, and gallic acid, protocatechuic acid, protocatechuic aldehyde and ( - ) -epicatechin were tested the samples. The contents of protocatechuic acid and protocatechuic aldehyde were from 0.004% to 0.05% and from 0.003% to 0.015%, respectively. CONCLUSION: The method can be used to control the quality of Rhizoma fagopyri dibotoryis.

[Identification of jizhi syrup using HPLC fingerprint].

OBJECTIVE: To establish a HPLC fingerprint for quantitative analysis of the active constituents of jizhi syrup. METHOD: HPLC analysis was performed on a Zorbax Sb C18 column (4.6 mm x 250 mm, 5 microm), the mobile phase in gradient elution was composed of (A) 1% acetic acid solution (including 0.2% triethylamine) and (B) acetonitrile, at a flow rate of 1.0 mL x min(-1). The column temperature was 30 degrees C and the wavelength was 280 nm. RESULT: 21 common peaks were tested in 10 batches of samples. Compared with the standard fingerprint, the similarity of each sample was greater than 0.99. At the same time, protocatechuic acid, protocatechu aldehyde, chlorogenic acid, caffeic acid, naringin and neohesperidin were found in all samples. CONCLUSION: The established method can be used for the quality control of jizhi syrup.

Treatment of meniscal injuries of knee joints by arthroscopy / 中国基层医药

Objective To assess the effectiveness of treatment of meniscal injuries of knee joints by arthroscopy.Methods 33 patients 35 joints were followed up and the parts,types and treatment under arthroscopy were analysed.Results 33 patients were followed up from six months to six years,the mean preoperative Lysholm score was 60.5 points,and the mean postoperative one was 86.7 points.Conclusion The advantage of treating meniscal injuries by arthroscopy was the result of correct examination and little wound of arthroscopy operation,and arthroscopic repair or partial menisectomy could effectively restore the function of the injured knee.