Dynamic network biomarker C1QTNF1 regulates tumor formation at the tipping point of hepatocellular carcinoma.

Identifying the precise moment before the onset of hepatocellular carcinoma (HCC) remains a significant challenge in the medical field. The existing biomarkers fall short of pinpointing the critical point preceding HCC formation. This study aimed to determine the exact tipping point for the transition from cirrhosis to HCC, identify the core Dynamic Network Biomarker (DNB), and elucidate its regulatory effects on HCC. A spontaneous HCC mouse model was established to mimic HCC formation in patients with chronic hepatitis. Using the DNB method, C1q and tumor necrosis factor (TNF) related 1 (C1QTNF1) protein was identified as the key DNB at the crucial tipping time of spontaneous HCC development. Both in vitro and in vivo studies showed that C1QTNF1 could inhibit tumor growth. Overexpression of C1QTNF1 before the tipping point effectively prevented HCC occurrence. Patients with elevated C1QTNF1 expression demonstrated improved overall survival (OS) (P = 0.03) and disease-free survival (DFS) (P = 0.03). The diagnostic value of C1QTNF1 was comparable to that of alpha-fetoprotein (AFP) (area under the curve [AUC] = 0.84; sensitivity 85%; specificity 80%). Furthermore, our research indicated that platelet-expressed C1QTNF1 is involved in cancer-associated signaling pathways. Our findings introduce a novel perspective by highlighting C1QTNF1 as the pivotal biomarker at the tipping point of primary HCC formation using DNB. We propose C1QTNF1 as a prognostic biomarker for HCC, potentially influencing tumor development through a platelet-related cancer signaling pathway.

MRI-based automated machine learning model for preoperative identification of variant histology in muscle-invasive bladder carcinoma.

OBJECTIVES: It is essential yet highly challenging to preoperatively diagnose variant histologies such as urothelial carcinoma with squamous differentiation (UC w/SD) from pure UC in patients with muscle-invasive bladder carcinoma (MIBC), as their treatment strategy varies significantly. We developed a non-invasive automated machine learning (AutoML) model to preoperatively differentiate UC w/SD from pure UC in patients with MIBC. METHODS: A total of 119 MIBC patients who underwent baseline bladder MRI were enrolled in this study, including 38 patients with UC w/SD and 81 patients with pure UC. These patients were randomly assigned to a training set or a test set (3:1). An AutoML model was built from the training set, using 13 selected radiomic features from T2-weighted imaging, semantic features (ADC values), and clinical features (tumor length, tumor stage, lymph node metastasis status), and subsequent ten-fold cross-validation was performed. A test set was used to validate the proposed model. The AUC of the ROC curve was then calculated for the model. RESULTS: This AutoML model enabled robust differentiation of UC w/SD and pure UC in patients with MIBC in both training set (ten-fold cross-validation AUC = 0.955, 95% confidence interval [CI]: 0.944-0.965) and test set (AUC = 0.932, 95% CI: 0.812-1.000). CONCLUSION: The presented AutoML model, that incorporates the radiomic, semantic, and clinical features from baseline MRI, could be useful for preoperative differentiation of UC w/SD and pure UC. CLINICAL RELEVANCE STATEMENT: This MRI-based automated machine learning (AutoML) study provides a non-invasive and low-cost preoperative prediction tool to identify the muscle-invasive bladder cancer patients with variant histology, which may serve as a useful tool for clinical decision-making. KEY POINTS: • It is important to preoperatively diagnose variant histology from urothelial carcinoma in patients with muscle-invasive bladder carcinoma (MIBC), as their treatment strategy varies significantly. • An automated machine learning (AutoML) model based on baseline bladder MRI can identify the variant histology (squamous differentiation) from urothelial carcinoma preoperatively in patients with MIBC. • The developed AutoML model is a non-invasive and low-cost preoperative prediction tool, which may be useful for clinical decision-making.

Discovery of Kinetin in inhibiting colorectal cancer progression via enhancing PSMB1-mediated RAB34 degradation.

Colorectal cancer (CRC) is one of the most prevalent malignancies worldwide. Understanding the underlying mechanism driving CRC progression and identifying potential therapeutic drug targets are of utmost urgency. We previously utilized LC-MS-based proteomic profiling to identify proteins associated with postoperative progression in stage II/III CRC. Here, we revealed that proteasome subunit beta type-1 (PSMB1) is an independent predictor for postoperative progression in stage II/III CRC. Mechanistically, PSMB1 binds directly to onco-protein RAB34 and promotes its proteasome-dependent degradation, potentially leading to the inactivation of the MEK/ERK signaling pathway and inhibition of CRC progression. To further identify potential anticancer drugs, we screened a library of 2509 FDA-approved drugs using computer-aided drug design (CADD) and identified Kinetin as a potentiating agent for PSMB1. Functional assays confirmed that Kinetin enhanced the interaction between PSMB1 and RAB34, hence facilitated the degradation of RAB34 protein and decreased the MEK/ERK phosphorylation. Kinetin suppresses CRC progression in patient-derived xenograft (PDX) and liver metastasis models. Conclusively, our study identifies PSMB1 as a potential biomarker and therapeutic target for CRC, and Kinetin as an anticancer drug by enhancing proteasome-dependent onco-protein degradation.

TCMBank: bridges between the largest herbal medicines, chemical ingredients, target proteins, and associated diseases with intelligence text mining.

Traditional Chinese Medicine (TCM) has long been viewed as a precious source of modern drug discovery. AI-assisted drug discovery (AIDD) has been investigated extensively. However, there are still two challenges in applying AIDD to guide TCM drug discovery: the lack of a large amount of standardized TCM-related information and AIDD is prone to pathological failures in out-of-domain data. We have released TCM Database@Taiwan in 2011, and it has been widely disseminated and used. Now, we developed TCMBank, the largest systematic free TCM database, which is an extension of TCM Database@Taiwan. TCMBank contains 9192 herbs, 61 966 ingredients (unduplicated), 15 179 targets, 32 529 diseases, and their pairwise relationships. By integrating multiple data sources, TCMBank provides 3D structure information of ingredients and provides a standard list and detailed information on herbs, ingredients, targets and diseases. TCMBank has an intelligent document identification module that continuously adds TCM-related information retrieved from the literature in PubChem. In addition, driven by TCMBank big data, we developed an ensemble learning-based drug discovery protocol for identifying potential leads and drug repurposing. We take colorectal cancer and Alzheimer's disease as examples to demonstrate how to accelerate drug discovery by artificial intelligence. Using TCMBank, researchers can view literature-driven relationship mapping between herbs/ingredients and genes/diseases, allowing the understanding of molecular action mechanisms for ingredients and identification of new potentially effective treatments. TCMBank is available at https://TCMBank.CN/.

Aquaporin 9 is involved in CRC metastasis through DVL2-dependent Wnt/ß-catenin signaling activation.

Background: Aquaporin 9 (AQP9) is permeable to water or other small molecules, and plays an important role in various cancers. We previously found that AQP9 was related to the efficacy of chemotherapy in patients with colorectal cancer (CRC). This study aimed to identify the role and regulatory mechanism of AQP9 in CRC metastasis. Methods: The clinical significance of AQP9 was analysed by using bioinformatics and tissue microarray. Transcriptome sequencing, Dual-Luciferase Reporter Assay, Biacore, and co-immunoprecipitation were employed to demonstrate the regulatory mechanism of AQP9 in CRC. The relationship between AQP9 and CRC metastasis was verified in vitro and in vivo by using real-time cell analysis assay, high content screening, and liver metastasis models of nude mice. Results: We found that AQP9 was highly expressed in metastatic CRC. AQP9 overexpression reduced cell roundness and enhanced cell motility in CRC. We further showed that AQP9 interacted with Dishevelled 2 (DVL2) via the C-terminal SVIM motif, resulting in DVL2 stabilization and the Wnt/ß-catenin pathway activation. Additionally, we identified the E3 ligase neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) as a modulator regulating the ubiquitination and degradation of AQP9. Conclusions: Collectively, our study revealed the important role of AQP9 in regulating DVL2 stabilization and Wnt/ß-catenin signaling to promote CRC metastasis. Targeting the NEDD4L-AQP9-DVL2 axis might have therapeutic usefulness in metastatic CRC treatment.

BK channel modulation by positively charged peptides and auxiliary γ subunits mediated by the Ca2+-bowl site.

The large-conductance, Ca2+-, and voltage-activated K+ (BK) channel consists of the pore-forming α (BKα) subunit and regulatory ß and γ subunits. The γ1-3 subunits facilitate BK channel activation by shifting the voltage-dependence of channel activation toward the hyperpolarization direction by about 50-150 mV in the absence of Ca2+. We previously found that the intracellular C-terminal positively charged regions of the γ subunits play important roles in BK channel modulation. In this study, we found that the intracellular C-terminal region of BKα is indispensable in BK channel modulation by the γ1 subunit. Notably, synthetic peptide mimics of the γ1-3 subunits' C-terminal positively charged regions caused 30-50 mV shifts in BKα channel voltage-gating toward the hyperpolarization direction. The cationic cell-penetrating HIV-1 Tat peptide exerted a similar BK channel-activating effect. The BK channel-activating effects of the synthetic peptides were reduced in the presence of Ca2+ and markedly ablated by both charge neutralization of the Ca2+-bowl site and high ionic strength, suggesting the involvement of electrostatic interactions. The efficacy of the γ subunits in BK channel modulation was reduced by charge neutralization of the Ca2+-bowl site. However, BK channel modulation by the γ1 subunit was little affected by high ionic strength and the positively charged peptide remained effective in BK channel modulation in the presence of the γ1 subunit. These findings identify positively charged peptides as BK channel modulators and reveal a role for the Ca2+-bowl site in BK channel modulation by positively charged peptides and the C-terminal positively charged regions of auxiliary γ subunits.

Functional classification of prostate cancer­associated miRNAs through CRISPR/Cas9­mediated gene knockout.

The aim of the present study was to use the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR­associated (Cas) 9­mediated gene knockout technology for the rapid classification of the differential function of micro (mi)RNAs screened using miRNA expression profiling by microarray. The rational design of single guide RNAs for the CRISPR/Cas9 system was verified to function in human LNCaP cells with rapid and efficient target gene editing. miRNA (miR)­205, miR­221, miR­222, miR­30c, miR­224, miR­455­3p, miR­23b and miR­505 were downregulated in patients with prostate cancer (PCa) and were experimentally validated to function as tumor suppressors in prostate cancer cells, affecting tumor proliferation, invasion and aerobic glycolysis. In addition, the data of the present study suggested that miR­663a and mfiR­1225­5p were upregulated in prostate cancer tissues and cell proliferation of miR­663a and miR­1225­5p knockout PCa cells was significantly lower compared with miR­NC cells. Furthermore, knockout of miR­1225­5p and miR­663a significantly decreased the lactate production in LNCaP cells in vitro. In conclusion, the present study offered a simple and efficient method for rapidly classifying miRNA function by applying CRISPR/Cas9 in LNCaP cells. The present study suggested, for the first time to the best of the authors' knowledge, that the aberrant expression of miR­663a and miR­1225­5p may be involved with the progression of prostate cancer, implying their potential as candidate markers for this type of cancer. However, the precise role of miR­663a and miR­1225­5p in accelerating the development of prostate cancer and promoting tumor progression remains to be elucidated.

Transient Cell-to-Cell Signaling Before Mitosis in Cultures of Human Bone Marrow-Derived Mesenchymal Stem/Stromal Cells.

Some types of cells, if not all, that undergo signal exchanges in culture need to contact other cells for various reasons, such as cell-to-cell contact for growth inhibition. However, signal exchanges by cell-to-cell contact before proliferation have never been reported. Using time-lapse recording, we discovered the emergence of several astonishing cell-to-cell contact modes in bone marrow-derived mesenchymal stem/stromal cells (MSCs) before the cells divided. When the cells contacted with another, a huge temporary synapse-like structure formed for molecule exchanges; a cell-tissue particle was taken in by a recipient cell; two cell membranes formed infusion-like structure for a short time; and even a 20-µm long and 5-µm wide cell tail was grafted to another cell. A total of 87% of cells underwent cell-to-cell contact before dividing. After epidermal growth factor-green fluorescent protein (EGF-GFP) vectors were transfected into MSCs and the cells were cocultured with unmanipulated MSCs, the unmanipulated MSCs took in EGF-GFP particles from EGF-GFP expressed MSCs, immediately increased in mitogen genes, and then divided. These results suggest that cells which may lack signal molecules may need to obtain these molecules from other cells through various types of cell-to-cell contact, as mentioned above. Our study provided valuable information to better understand the behaviors of cell-to-cell contact and communication before mitosis.

[Laparoscopic percutaneous extraperitoneal closure does not affect vas deferens orientation or testicular volume and perfusion].

OBJECTIVE: To investigate the influence of single-port laparoscopic percutaneous extraperitoneal closure (LPEC) on the orientation of the vas deferens and the volume and perfusion of the testis in pediatric patients undergoing inguinal hernia repair. METHODS: A total of 92 consecutively enrolled boys diagnosed with unilateral inguinal hernia underwent single-port LPEC between June 2013 and June 2014. The orientation of the vas deferens and the testicular volume and perfusion of the patients were ultrasonographically assessed preoperatively and at 1 and 6 months after surgery. RESULTS: All the surgical procedures were performed successfully without conversion or serious perioperative complications. Ultrasonography showed no angulation or distortion of the vas deferens on the surgical side during a six-month follow-up period. Similarly, no obvious changes were observed in the testicular volume or perfusion. CONCLUSIONS: Single-port LPEC is safe and effective in the treatment of pediatric inguinal hernia and does not affect the orientation of the vas deferens or testicular volume and perfusion.

BCL9, a coactivator for Wnt/ß-catenin transcription, is targeted by miR-30c and is associated with prostate cancer progression.

B-cell lymphoma 9 (BCL9), a component of aberrantly activated Wnt signaling, is an important contributing factor to tumor progression. Our previous data indicated that downregulation of the tumor suppressor microRNA-30c (miR-30c) was a frequent pathogenetic event in prostate cancer (PCa). However, a functional link between miR-30c and BCL9/Wnt signaling, and their clinical and pathological significance in PCa, have not been well established. The present study demonstrated that miR-30c serves as a key negative regulator targeting BCL9 transcription in PCa cells. Ectopic expression of miR-30c was associated with reduced expression of Wnt pathway downstream targets, including c-Myc, cluster of differentiation 44 and sex determining region Y-box 9 in DU145 human PCa cells. Examination of clinical prostate specimens revealed higher levels of BCL9 expression in PCa compared with that in benign prostate tissues. After substantiating this finding by patient sample analysis, BCL9 expression or activity was observed to be closely correlated with PCa biochemical recurrence (BCR) and disease progression, whereas it was inversely associated with miR-30c. Furthermore, overexpression of BCL9 in PCa acted cooperatively with miR-30c low expression to predict earlier BCR in PCa. These findings indicate that inhibition of BCL9/Wnt signaling by miR-30c is important in the progression of PCa. Furthermore, the combined analysis of miR-30c and BCL9 may be valuable tool for prediction of BCR in PCa patients following radical prostatectomy.

Protein regulator of cytokinesis 1 overexpression predicts biochemical recurrence in men with prostate cancer.

BACKGROUND: Protein regulator of cytokinesis 1 (PRC1) has been reported to be implicated into the completion of cytokinesis and is dys-regulated in a cancer-specific manner. However, it roles in human prostate cancer (PCa) remain unclear. In the current study, we aimed to investigate the expression pattern of PRC1 and its clinical significance in this malignancy. MATERIALS AND METHODS: PRC1 protein expression in human PCa and non-cancerous prostate tissues was detected by immunohistochemistry, which was validated by microarray-based Taylor data at mRNA level. Then, the associations of PRC1 expression with clinicopathological features and clinical outcome of PCa patients were statistically analyzed. RESULTS: PRC1 expression in PCa tissues, at both mRNA and protein levels, were significantly higher than those in non-cancerous prostate tissues. In addition, the PCa patients with PRC1 overexpression more frequently had high Gleason score, advanced pathological stage, positive metastasis, short overall survival time and positive PSA failure than those with low Gleason score, early pathological stage, negative metastasis, long overall survival time and negative PSA failure (all P<0.05). Moreover, PRC1 expression was identified as an unfavorable prognostic factor of biochemical recurrence-free survival in PCa patients (P<0.001). CONCLUSION: These findings suggest that the aberrant expression of PRC1 may predict biochemical recurrence in men with PCa highlighting its potential as a prognostic marker of this malignancy.

Molecular cloning, phylogenetic analysis, and expression profiling of endoplasmic reticulum molecular chaperone BiP genes from bread wheat (Triticum aestivum L.).

BACKGROUND: The endoplasmic reticulum chaperone binding protein (BiP) is an important functional protein, which is involved in protein synthesis, folding assembly, and secretion. In order to study the role of BiP in the process of wheat seed development, we cloned three BiP homologous cDNA sequences in bread wheat (Triticum aestivum), completed by rapid amplification of cDNA ends (RACE), and examined the expression of wheat BiP in wheat tissues, particularly the relationship between BiP expression and the subunit types of HMW-GS using near-isogenic lines (NILs) of HMW-GS silencing, and under abiotic stress. RESULTS: Sequence analysis demonstrated that all BiPs contained three highly conserved domains present in plants, animals, and microorganisms, indicating their evolutionary conservation among different biological species. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) revealed that TaBiP (Triticum aestivum BiP) expression was not organ-specific, but was predominantly localized to seed endosperm. Furthermore, immunolocalization confirmed that TaBiP was primarily located within the protein bodies (PBs) in wheat endosperm. Three TaBiP genes exhibited significantly down-regulated expression following high molecular weight-glutenin subunit (HMW-GS) silencing. Drought stress induced significantly up-regulated expression of TaBiPs in wheat roots, leaves, and developing grains. CONCLUSIONS: The high conservation of BiP sequences suggests that BiP plays the same role, or has common mechanisms, in the folding and assembly of nascent polypeptides and protein synthesis across species. The expression of TaBiPs in different wheat tissue and under abiotic stress indicated that TaBiP is most abundant in tissues with high secretory activity and with high proportions of cells undergoing division, and that the expression level of BiP is associated with the subunit types of HMW-GS and synthesis. The expression of TaBiPs is developmentally regulated during seed development and early seedling growth, and under various abiotic stresses.

Molecular characterisation and evolution of HMW glutenin subunit genes in Brachypodium distachyon L.

Brachypodium distachyon, a small wild grass within the Pooideae family, is a new model organism for exploring the functional genomics of cereal crops. It was shown to have close relationships to wheat, barley and rice. Here, we describe the molecular characterisation and evolutionary relationships of high molecular weight glutenin subunits (HMW-GS) genes from B. distachyon. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), high performance capillary electrophoresis (HPCE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses demonstrated that there was no HMW-GS expression in the Brachypodium grains due to the silencing of their encoding genes. Through allele-specific polymerase chain reaction (AS-PCR) amplification and cloning, a total of 13 HMW-GS encoding genes from diploid, tetraploid and hexaploid Brachypodium species were obtained, and all of them had typical structural features of y-type HMW-GS genes from common wheat and related species, particularly more similar to the 1Dy12 gene. However, the presence of an in-frame premature stop codon (TAG) at position 1521 in the coding region resulted in the conversion of all the genes to pseudogenes. Further, quantitative real-time PCR (qRT-PCR) analysis revealed that HMW-GS genes in B. distachyon displayed a similar trend, but with a low transcriptional expression profile during grain development due to the occurrence of the stop codon. Phylogenetic analysis showed that the highly conserved Glu-1-2 loci were presented in B. distachyon, which displayed close phylogenetic evolutionary relationships with Triticum and related species.