Quantitative analysis and visualization of literature on acupuncture and related TCM therapies for the treatment of colorectal cancer based on CiteSpace.

Objective: We analyzed the literature describing the results of treatment of colorectal cancer (CRC) using acupuncture in the past three decades from the Web of Science (WoS) and Chinese databases (including CNKI, WANGFANG and VIP), and summarized the current development of CRC treatment as well as future research directions through the presentation of maps and visualization analysis. Methods: We searched the WoS and Chinese databases. Relevant articles were exported, and the data were organized using Excel software and was visualized and analyzed using CiteSpace software. Results: A total of 355 articles from the WoS and 95 articles from Chinese databases were selected for inclusion in the analysis. The articles in WoS were sourced from 174 journals, 1274 institutions, and 66 countries, and covered 299 keywords. The articles in the Chinese databases were sourced from 43 journals, 111 institutions, and 3 countries, and included 126 keywords. The article with the most citations in the WoS was cited 128 times and in the Chinese databases, the article with the most citations was cited 120 times. Acupuncture, CRC, rectal cancer, apoptosis, warm acupuncture, traditional Chinese medicine (TCM) and gastrointestinal function were mentioned most frequently in the WoS. CRC, electroacupuncture, gastrointestinal function, rectal cancer, acupuncture and moxibustion, acupuncture, and colon cancer were mentioned most frequently in the Chinese databases. Conclusion: Both the WoS and Chinese databases showed a gradual increase in the number of articles related to acupuncture treatment for CRC, indicating a growing interest in this area. Acupuncture treatments are diverse, including warm acupuncture, auricular acupuncture, acupuncture injection, and electroacupuncture. They are often used in combination with drugs to treat symptoms such as depression, nausea and vomiting, pain, diarrhea, and urinary and fecal incontinence, which are commonly associated with CRC.

Knockdown of Long Noncoding RNA 01124 Inhibits the Malignant Behaviors of Colon Cancer Cells via miR-654-5p/HAX-1.

Background: Previous studies have shown that long noncoding RNAs (lncRNAs) play a key role in cancer, including colon cancer (CC). However, the exact role of long noncoding RNA 01124 (LINC01124) in CC and its mechanisms of action remain unknown. In this study, we investigated the functional effects and the possible mechanism of LINC01124 in CC. Methods: We first determined the expression of LINC01124 in CC tissues (The Cancer Genome Atlas (TCGA) database) and cell lines (quantitative real-time polymerase chain reaction (qRT-PCR)). Functional analysis via Cell Counting Kit-8 (CCK-8), colony formation, cell cycle, wound healing and Transwell assays were performed, and a mechanistic experiment was performed with the western blotting. The function of LINC01124 was also determined in vivo using nude BALB/c mice. Results: The results showed that LINC01124 was upregulated in CC tissues and cell lines. Functional studies showed that knockdown of LINC01124 significantly suppressed the proliferation, migration, and invasion of colon cancer cells in vitro and in vivo. Subsequent mechanistic experiments indicated that LINC01124 acted as a sponge to suppress microRNA 654-5p, which targeted HAX-1. Downregulation of LINC01124 decreased the expression of HAX-1, and overexpression of the miR-654-5p inhibitor attenuated the sh-LINC01124-induced inhibition of CC cell proliferation, migration, and invasion. Conclusion: Collectively, this study revealed that the knockdown of LINC01124 inhibited the malignant behaviors of CC via the miR-654-5p/HAX-1 axis, suggesting that LINC01124 might be a therapeutic target for CC treatment.

Dermcidin Enhances the Migration, Invasion, and Metastasis of Hepatocellular Carcinoma Cells In Vitro and In Vivo.

Background and Aims: Hepatocellular carcinoma (HCC) is a common primary liver neoplasm with high mortality. Dermcidin (DCD), an antimicrobial peptide, has been reported to participate in oncogenesis. This study assessed the effects and underlying molecular events of DCD overexpression and knockdown on the regulation of HCC progression in vitro and in vivo. Methods: The serum DCD level was detected using enzyme-linked immunosorbent assay. DCD overexpression, knockdown, and Ras-related C3 botulinum toxin substrate 1 (Rac1) rescue were performed in SK-HEP-1 cells using plasmids. Immunofluorescence staining, quantitative PCR, and Western blotting were used to detect the expression of different genes and proteins. Differences in HCC cell migration and invasion were detected by Transwell migration and invasion assays. A nude mouse HCC cell orthotopic model was employed to verify the in vitro data. Results: The level of serum DCD was higher in patients with HCC and in SK-HEP-1 cells. DCD overexpression caused upregulation of DCD, fibronectin, Rac1, and cell division control protein 42 homologue (Cdc42) mRNA and proteins as well as actin-related protein 2/3 (Arp2/3) protein (but reduced Arp2/3 mRNA levels) and activated Rac1 and Cdc42. Phenotypically, DCD overexpression induced HCC cell migration and invasion in vitro, whereas knockout of DCD expression had the opposite effects. A Rac1 rescue experiment in DCD-knockdown HCC cells increased HCC cell migration and invasion and increased the levels of active Rac1/total Rac1, Wiskott-Aldrich syndrome family protein (WASP), Arp2/3, and fibronectin. DCD overexpression induced HCC cell metastasis to the abdomen and liver in vivo. Conclusions: DCD promotes HCC cell migration, invasion, and metastasis through upregulation of noncatalytic region of tyrosine kinase adaptor protein 1 (Nck1), Rac1, Cdc42, WASP, and Arp2/3, which induce actin cytoskeletal remodeling and fibronectin-mediated cell adhesion in HCC cells.

CRMP5 regulates cell proliferation and development of colorectal cancer via MAPK-dependent signaling.

Colorectal cancer (CRC) is one of the most common malignancies worldwide. Collapsin response mediator protein 5 (CRMP5) belongs to a family of five cytosolic proteins that serve a major role in neural development. CRMP5 has been identified as a biomarker of numerous cancer types, including lung cancer and glioblastoma. However, the role of CRMP5 in CRC remains unclear. In the present study, CRMP5 was characterized as a novel biomarker of poor survival in CRC. CRMP5-overexpression in CRC cells promoted cell proliferation and migration while CPMP5-knockdown decreased cell growth and migration. A novel mechanism was uncovered, by which CRMP5 regulates MAPK signaling to drive CRC cell proliferation and development. Furthermore, CRMP5-overexpression induced chemotherapy resistance and tumor recurrence in CRC. Taken together, these results demonstrated the important role of CRMP5 in the development and proliferation of CRC cells and suggested that CRMP5 may be a novel therapeutic target for CRC.

lncRNA LINC00963 downregulation regulates colorectal cancer tumorigenesis and progression via the miR­10b/FGF13 axis.

Long non-coding RNAs (lncRNAs) serve a key role in different types of cancer, including colorectal cancer (CRC). The exact roles and mechanisms underlying lncRNA00963 [long intergenic non­protein coding RNA 963 (LINC00963)] in CRC are not completely understood. The present study aimed to identify the effects and mechanisms underlying LINC00963 in CRC. Firstly, the LINC00963 expression was detected using reverse transcription­quantitative PCR and the results demonstrated that LINC00963 expression levels were significantly increased in CRC tissues and cell lines compared with healthy tissues and HpoEpiC cells, respectively. Online database analysis indicated that high levels of LINC00963 were associated with low survival rates. The results of functional experiments, such as CCK­8 assay, colony formation assay, wound healing assay and Transwell invasion assay, indicated that LINC00963 knockdown significantly inhibited CRC cell proliferation, colony formation, migration and invasion compared with the small interfering RNA (si)­negative control (NC) group. Furthermore, the luciferase reporter indicated that LINC00963 competitively regulated microRNA (miR)­10b by targeting fibroblast growth factor 13 (FGF13). Compared with si­NC, LINC00963 knockdown decreased the expression levels of FGF13, vimentin and N­cadherin, and increased the expression of E­cadherin as detecting by western blotting. miR­10b inhibitors partly attenuated si­LINC00963­induced inhibition of CRC cell proliferation, migration and invasion. Collectively, the results of the present study suggested a potential role of the LINC00963/miR-10b/FGF13 axis in the tumorigenesis and progression of CRC, indicating a novel lncRNA-based diagnostic or therapeutic target for CRC.

Long non­coding RNA LINC00312 regulates breast cancer progression through the miR­9/CDH1 axis.

Long non­coding RNAs (lncRNAs) are important mediators of the initiation and progression of tumors, including breast cancer (BC). The exact role of long intergenic non­coding RNA 00312 (LINC00312) in BC and its mechanism of action have not been reported to date. In the present study, LINC00312 was found to be downregulated in human BC tissues and cell lines by RT­qPCR. The findings of a functional study indicated that overexpression of LINC00312 suppressed the proliferation, colony forming ability, migration and invasiveness of BC cell lines. Mechanistically, LINC00312 was found to induce suppression of cell migration and invasion by directly binding to miR­9. Overexpression of LINC00312 increased the expression of cadherin 1 (CDH1), a direct target of miR­9, and decreased the expression of vimentin (VIM), a major cytoskeletal component of mesenchymal cells as determined by western blot analysis. miR­9 partly abrogated the upregulation of CDH1 and downregulation of VIM induced by LINC00312. Taken together, the results of the present study indicate a role for the LINC00312/miR­9/CDH1 axis in the progression of BC, and suggest a novel lncRNA­based diagnostic biomarker or therapeutic target for BC.

Corilagin Inhibits Esophageal Squamous Cell Carcinoma by Inducing DNA Damage and Down-Regulation of RNF8.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC), the most prevalent histologic subtype of esophageal cancer, is an aggressive malignancy with poor prognosis and a high incidence in the East. Corilagin, an active component present in Phyllanthus niruri L., has been shown to suppress tumor growth in various cancers. However, the effects of corilagin on ESCC and the mechanisms for its tumor suppressive function remain unknown. METHODS: Cell proliferation was measured by Cell Counting Kit-8 assay and colony formation assays. Annexin V/PI double-staining was performed to assess cell apoptosis. Immunofluorescence staining and western blotting were used to evaluate the protein expression. A xenograft mice model was used to assess the in vivo antitumor effects of corilagin alone or in combination with cisplatin. RESULTS: We for the first time showed that corilagin was effectively able to inhibit ESCC cell proliferation and induce cell apoptosis. Additionally, our results validated its antitumor effects in vivo using a xenograft mouse model. Mechanistically, we found that corilagin caused significant DNA damage in ESCC cells. We found that corilagin could significantly attenuate the expression of the E3 ubiquitin ligase RING finger protein 8 (RNF8) through ubiquitin-proteasome pathway, leading to the inability of DNA damage repair response and eventually causing cell apoptosis. Furthermore, we also showed that corilagin substantially enhanced the antitumor effects of chemotherapy drug cisplatin both in vitro and in vivo. CONCLUSION: Our results not only provided novel and previously unrecognized evidences for corilagin-induced tumor suppression through inducing DNA damage and targeting RNF8 in ESCC, but also highlighted that corilagin might serve as an adjunctive treatment to conventional chemotherapeutic drugs in ESCC patients.

The Role of Dermcidin in the Diagnosis and Staging of Hepatocellular Carcinoma.

INTRODUCTION: Hepatocellular carcinoma (HCC) is a major contributor to cancer-related deaths due to its often late stage diagnosis. Our previous study showed that dermcidin (DCD) may have the potential to be used as a serum biomarker for HCC for more timely diagnoses. MATERIALS AND METHODS: In this study, we measured serum DCD and alpha-fetoprotein (AFP) levels in 87 HCC patients; 33 liver cirrhosis (LC); and 44 normal controls (NC), evaluated the relationship between DCD levels and clinicopathological parameters. RESULTS: Serum DCD levels in HCC patients (27.03 ng/mL) were significantly higher than in LC patients (24.78 ng/mL, p < 0.05), and NC subjects (18.98 ng/mL, p < 0.001). The optimum cutoff values were 25.75 ng/mL for DCD and 9.86 ng/mL for AFP. DCD had a greater area under the receiver operating characteristic curve (AUC) for differentiating HCC from the controls than AFP (AUC = 0.769 vs. 0.729, respectively). Importantly, our cohort revealed that serum DCD levels were positively correlated with metastasis in HCC patients versus HCC patients without metastatic disease (32.31 vs. 23.95, p < 0.001). Western blot results showed that DCD expression was significantly upregulated in seven tumor tissues compared with the noncancerous adjacent tissues. Immunohistochemistry performed in four paired samples confirmed the upregulation of DCD expression in the tumor tissues. CONCLUSIONS: Our results showed that serum DCD levels were significantly increased in HCC patients and in cancerous tissue. DCD could potentially be used as a biomarker for the diagnosis of HCC.

Evaluation of two high-abundance protein depletion kits and optimization of downstream isoelectric focusing.

Disease biomarkers for diagnostic and prognostic purposes are most likely within an extremely low concentration range and are thus masked by the presence of high­abundance proteins. Therefore, removing high­abundance proteins is the main challenge for identifying disease biomarkers. In addition, the solution obtained from high­abundance protein depletion kits contains a rich array of compounds, which interfere with isoelectric focusing (IEF). In the present study, the effect of two commercial kits was evaluated and the downstream IEF protocol was optimized. High­resolution results could be obtained according to the following conditions: The ProteoPrep Blue Albumin and IgG Depletion kit depleted albumin and IgG; immobilized pH gradient strips (typically 18 cm) were rehydrated with sample buffer containing 250 µg serum proteins at 30 v for 6 h, 60 v for 6 h, 200 v for 2 h, 500 v for 2 h, 1,000 v for 2 h, 5,000 v for 2 h, 10,000 v for 2 h and then focusing at 10,000 v up to 110 k vhs. In addition, the protein spots identified by matrix­assisted laser desorption ionization time­of­flight mass spectrometry demonstrated that all proteins had a low abundance. The present study not only provides a definite and effective method for removing high­abundance proteins, but also provides a proper protocol (protocol C) for downstream IEF. The present study includes a comprehensive investigation of serum proteomics, which paves the way for serum protein research.

A novel interaction between the SH2 domain of signaling adaptor protein Nck-1 and the upstream regulator of the Rho family GTPase Rac1 engulfment and cell motility 1 (ELMO1) promotes Rac1 activation and cell motility.

Nck family proteins function as adaptors to couple tyrosine phosphorylation signals to actin cytoskeleton reorganization. Several lines of evidence indicate that Nck family proteins involve in regulating the activity of Rho family GTPases. In the present study, we characterized a novel interaction between Nck-1 with engulfment and cell motility 1 (ELMO1). GST pull-down and co-immunoprecipitation assay demonstrated that the Nck-1-ELMO1 interaction is mediated by the SH2 domain of Nck-1 and the phosphotyrosine residues at position 18, 216, 395, and 511 of ELMO1. A R308K mutant of Nck-1 (in which the SH2 domain was inactive), or a 4YF mutant of ELMO1 lacking these four phosphotyrosine residues, diminished Nck-1-ELMO1 interaction. Conversely, tyrosine phosphatase inhibitor treatment and overexpression of Src family kinase Hck significantly enhanced Nck-1-ELMO1 interaction. Moreover, wild type Nck-1, but not R308K mutant, significantly augmented the interaction between ELMO1 and constitutively active RhoG (RhoG(V12A)), thus promoted Rac1 activation and cell motility. Taken together, the present study characterized a novel Nck-1-ELMO1 interaction and defined a new role for Nck-1 in regulating Rac1 activity.

Detection of tyrosine­phosphorylated proteins in hepatocellular carcinoma tissues using a combination of GST­Nck1­SH2 pull­down and two­dimensional electrophoresis.

Tyrosine-phosphorylated proteins govern a host of cell functions, such as growth, division, adhesion and motility. We previously identified a group of Nck Src homology 2 (SH2) domain­binding proteins by combining the GST-Nck1-SH2 pull-down method with two-dimensional electrophoresis (2­DE) in hepatocellular carcinoma (HCC) tissues. In the present study, different methods and conditions for key procedures of GST-Nck1-SH2 pull-down and 2­DE were investigated and optimized. High-resolution results were obtained using the following conditions: a total amount of 100 µl GST-Nck1-SH2 fusion proteins/10 mg liver proteins to execute the pull-down procedure; 7 M urea and 2 M thiourea as lysis buffer; ultrafiltration depletion of interferential materials. Moreover, we performed a negative control experiment using GST-4T3 during the pull-down procedure, and further demonstrated that the proteins obtained using the aforementioned method interacted with Nck in a tyrosine phosphorylation-dependent manner. The optimized method offers a rapid, efficient alternative for the high­quantity screening of tyrosine-phosphorylated protein expression and solubility, which in turn facilitates future studies on tyrosine-phosphorylated proteins.

Identification of Dermcidin as a novel binding protein of Nck1 and characterization of its role in promoting cell migration.

A distinct feature of hepatocellular carcinoma (HCC) is the tendency of tumor cells to disperse throughout the liver. Nck family adaptor proteins function to couple tyrosine phosphorylation signals to regulate actin cytoskeletal reorganization that leads to cell motility. In order to explore the role of Nck in HCC development, we performed GST pull-down assay using the SH2 domain of Nck1 as bait. The resulting precipitates were separated by 2-DE. Mass spectrometry analysis revealed a group of Nck1 SH2 domain-binding proteins that were differentially expressed in HCC. One of these proteins, dermcidin (DCD), and its interaction with Nck1, was further validated in vitro. GST pull-down assay revealed that Nck1 SH2 domain binds to the phosphotyrosine residue at position 20 (Y20) of the DCD. Pervandate treatment significantly enhanced the interaction between DCD and Nck1. Moreover, we demonstrated that forced expression of DCD could activate Rac1 and Cdc42 and promoted cell migration. Taken together, these data suggest a role of DCD in tumor metastasis.

Ectopic expression of Ligand-of-Numb protein X promoted TGF-beta induced epithelial to mesenchymal transition of proximal tubular epithelial cells.

Ligand-of-Numb protein X (LNX) was initially characterized as a RING finger type E3 ubiquitin ligase that targeted the intrinsic cell fate determinant Numb for ubiquitination dependent degradation. However, the physiological function of LNX remains largely unknown. In the present study, we demonstrate that ectopic expression of LNX in human proximal tubular epithelial cells (HK-2 cells) significantly enhanced TGF-beta1 induced epithelial to mesenchymal transition (EMT). The EMT-promoting effect of LNX manifested as strong inhibition of E-cadherin expression, enhanced expression of vimentin, fibronectin or PAI-1, and increased cell migration. This function of LNX was shown to be independent of its ligase activity because ectopic expression of a mutant form of LNX (C48ALNX) that lacks E3 ligase activity had the similar effect as the wild-type LNX. Overexpression of E-cadherin could inhibit LNX augmented EMT. This study suggests a potential role for LNX in promoting EMT in human proximal tubular epithelial cells.