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The aim of this study was to apply a two-stage deep model combining multi-scale feature maps and the recurrent attention model (RAM) to assist with the pathological diagnosis of breast cancer histological subtypes by the use of whole slide images (WSIs).In this article, we propose an integrated framework combining multi-scale feature maps from Inception V3 and the recurrent attention model to classify the six histological subtypes of breast cancer. This model was trained with 194 WSIs, and on 63 validation WSIs the model achieved accuracies of 0.9030 for patch-level classification and 0.8889 for WSI-level classification. In the testing stage, a total of 65 WSIs were used to achieve an accuracy of 0.8462 without any form of data curation. The t-distributed stochastic neighbour embedding showed that features extracted by the feature network of the RAM from WSIs of the same category can cluster together after training, and the visualization of decision steps showed that the decision-making glimpses are focused on the middle tumour area of an example from test WSIs. Finally, the false classification patches indicated that the morphological similarities between tumour tissues of different subtypes or non-tumour tissues and tumour tissues in patches might contribute to misclassification.This model can imitate the diagnostic process of pathologists, pay attention to a series of local features on the pathology image, and summarize related information, in order to accurately classify breast cancer into its histological subtypes, which is important for treatment and prognosis.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide, and identification of novel targets is necessary for its diagnosis and treatment. This study aimed to investigate the biological function and clinical significance of tweety homolog 3 (TTYH3) in HCC. TTYH3 overexpression promoted cell proliferation, migration, and invasion and inhibited HCCM3 and Hep3B cell apoptosis. TTYH3 promoted tumor formation and metastasis

The demand for artificial vascular grafts in clinical applications is increasing, and it is urgent to design a tissue-engineered vascular graft with good biocompatibility and sufficient mechanical strength. In this study, three-layer small diameter artificial vascular grafts were constructed by electrospinning. Polycaprolactone (PCL) and collagen (COL) were used as the inner layer to provide good biocompatibility and cell adhesion, the middle layer was PCL to improve the mechanical properties, and gelatin (GEL) and PCL were used to construct the outer layer for further improving the mechanical properties and biocompatibility of the vascular grafts in the human body environment. The electrospun artificial vascular graft had good biocompatibility and mechanical properties. Its longitudinal maximum stress reached 2.63 ± 0.12 MPa, which exceeded the maximum stress that many natural blood vessels could withstand. The fiber diameter of the vascular grafts was related to the proportion of components that made up the vascular grafts. In the inner structure of the vascular grafts, the hydrophilicity of the vascular grafts was enhanced by the addition of COL to the PCL, and human umbilical vein endothelial cells (HUVECs) adhered more easily to the vascular grafts. In particular, the cytocompatibility and proliferation of HUVECs on the scaffold with an inner structure PCL:COL = 2:1 was superior to other ratios of vascular grafts. The vascular grafts did not cause hemolysis of red blood cells. Thus, the bionic PCL-COL@PCL@PCL-GEL composite graft is a promising material for vascular tissue engineering.

Based on the theory of planned behavior, the aim of this study was to describe the influencing factors of patient delay intentions and behaviors in benign prostatic hyperplasia (BPH) patients and to provide a reference for the development of a patient delay intention scale. This study was carried out over 4 months in 2021 in Daqing, Heilongjiang, China. The participants were 20 patients with BPH who were aged 60 to 82 years and experienced patient delay; participants were selected through a purposive sampling method. The data were collected via face-to-face semistructured interviews. Five main themes emerged from the interviews, including an insufficient understanding of symptoms, experiences of coping instead of seeking health care, negative attitudes toward care-seeking, the influence of others on decision-making for care-seeking, and obstacles to seeking health care. In conclusion, the patient delay intentions and behaviors of BPH patients are the result of a combination of many factors.

Cervical cancer is one of the most prevalent gynecological malignancies. Although the functions of long non-coding RNA (lncRNA) plasmacytoma variant translocation 1 (PVT1) and c-Myc in tumorigenesis have been acknowledged, the roles of c-Myc and lncRNA-PVT1 in the proliferation of cervical cancer are still unclear. Our study is designed to demonstrate the regulatory network involving c-Myc and lncRNA-PVT1 in cervical cancer. Quantitative real-time PCR and western blot assays were performed in our research to estimate the expression levels of RNA and proteins. CCK8 assays were applied to demonstrate the viability of HeLa and SiHa cells. Immunofluorescence assay was then used to investigate the co-localization of lncRNA-PVT1 and miR-486-3p. Binding of c-Myc to the promoter region of PVT1 was identified by ChIP-assay. Functionally, upregulation of lncRNA-PVT1 enhanced the proliferation and viability of cervical cancer cells. Mechanistically, lncRNA-PVT1 sponged miR-486-3p and released its repression of extracellular matrix protein 1. Besides, c-Myc functioned as an activator of lncRNA-PVT1 and upregulated its expression by binding to the promoter of PVT1 in cervical cancer cells. lncRNA-PVT1 was upregulated by c-Myc and thus enhanced the proliferation of cervical cancer cells by sponging miR-486-3p.

Tumor-associated macrophage (TAM) is a major component of tumor microenvironment (TME) and plays critical role in the progression of cancer metastasis. However, TAM-mediated regulation in gallbladder cancer (GBC) has not been fully characterized. Here, we found that exosomes derived from GBC cell polarized macrophage to M2 phenotype, which then facilitated the invasion and migration of GBC cells. We discovered that leptin was enriched in GBC cell-derived exosomes. Exosomal leptin levels promoted invasion and migration of GBC-SD cells. The inhibition of leptin not only attenuated M2 macrophage of polarization but also inhibited the invasive and migratory ability of GBC cell. In addition, GBC-SD cell-derived exosomal leptin induced M2 polarization of macrophage via activation of STAT3 signal pathway. Taken together, our results suggested that GBC cells secrete exosome-enclosed leptin facilitated cell invasion and migration via polarizing TAM.

We report a case of childhood coronavirus disease 2019 infection with pleural effusion complicated by possible secondary Mycoplasma pneumoniae infection. Fever and pulmonary lesions on computed tomography were the early clinical manifestations, and the patient developed nonproductive cough later. The hydrothorax in this coronavirus disease 2019 case was exudative, showing predominantly mature lymphocytes.

In an effort to develop novel Bax activators for breast cancer treatment, a series of diverse analogues have been designed and synthesized based on lead compound SMBA1 through several strategies, including introducing various alkylamino side chains to have a deeper access to S184 pocket, replacing carbon atoms with nitrogen, and reducing the nitro group of 9H-fluorene scaffold. Compounds 14 (CYD-2-11) and 49 (CYD-4-61) have been identified to exhibit significantly improved antiproliferative activity compared to SMBA1, with IC50 values of 3.22 μM and 0.07 μM against triple-negative breast cancer MDA-MB-231 and 3.81 μM and 0.06 μM against ER-positive breast cancer MCF-7 cell lines, respectively. Mechanism of action studies of compound 49 indicated that it can activate Bax protein to induce cytochrome c release and regulate apoptotic biomarkers, leading to cancer cell apoptosis. Further in vivo efficacy studies of compounds 14 and 49 in nude mice bearing MDA-MB-231 tumor xenografts demonstrated that these drug candidates can significantly suppress tumor growth, indicating their therapeutic potential for the treatment of breast cancer.

Background Management of recurrent hepatocellular cancer (HCC) after liver resection is challenging, with unsatisfactory long-term patient outcomes. Liver re-resection, in theory, is a good treatment option. We therefore studied prognosis and risk factors of patients who undergo re-hepatectomy. Methods We retrospectively analyzed 103 patients who underwent re-hepatectomy. Results The re-resection postoperative complication rate was 31.1% (32/103). Patients with gross vascular invasion (GVI), cirrhosis, or hepatitis B (HBV) infections not treated with antiviral therapy had higher morbidity than patients without these diseases (per chi-square tests). In bivariate regression analysis, cirrhosis (odds ratio [OR]: 10.308, P = 0.031) and HBV not treated with antiviral therapy (OR: 3.982, P = 0.011) were associated with immediate postoperative morbidity. Median overall survival (OS) after re-resection was 65.0 months (range: 2.1–119.3 months); cumulative OS rates were 1-year: 92.1%, 2-year: 78.2%, and 5-year: 54.4%. Independent risk factors for worse survival were serum AFP level > 20 ng/mL at first resection, portal hypertension (PH) and GVI at recurrence. In the non-PH group, microvascular invasion (micro-VI), GVI and pTNM III–IV disease were associated with poor prognosis; patients with pTNM I–II disease had significantly less micro-VI and GVI than did patients with advanced disease. Conclusion Repeat hepatectomy has favorable long-term outcomes. Cirrhosis and HBV not treated with antiviral therapy were associated with immediate postoperative morbidity. Serum AFP > 20 ng/mL at first resection, PH, and GVI at recurrence are independent prognostic factors. For patients without PH, TNM staging can predict prognosis.

Intrahepatic cholangiocarcinoma (ICC) can invade and metastasize. EIF5A2 is involved in the invasive metastatic process of several digestive malignancies. However, its role in ICC is yet to be elucidated.Immunohistochemistry (IHC) and Western blot (WB) were used to detect the level of EIF5A2 in the tumor specimens of ICC patients and evaluate the correlation between its expression and clinicopathological characteristics. The significance of EIF5A2 in the prognosis of ICC patients was further evaluated by Kaplan-Meier and Cox regression analysis. In addition, CCK-8, EdU, Transwell invasion, and scratch assays were utilized to detect tumor cell proliferation, invasion, and metastasis. Furthermore, the role of EIF5A2 in ICC cells was evaluated after modification of EIF5A2 expression.The level of EIF5A2 protein was significantly higher in ICC than in adjacent tissues. This high expression in the tumor samples was significantly associated with malignant phenotypes, such as lymph node metastasis (LNM), microvascular or bile duct invasion, and poor differentiation. ICC patients with high expression of EIF5A2 had short overall survival and a high cumulative recurrence rate. The multifactorial analysis showed that EIF5A2 is an independent prognostic marker. Furthermore, high levels of EIF5A2 may activate the PI3K/AKT/mTOR signaling pathway and upregulate Cyclin D1, Cyclin D3, MMP2, and MMP9 to promote ICC cell proliferation, migration, and invasion.The current study found that EIF5A2 promotes ICC progression and is a prognostic biomarker and candidate therapeutic target for ICC patients.

In our previous research, we developed a 32-gene risk index model that may be utilized as a robust prognostic method for predicting prostate cancer (PCa) recurrence after surgery. Among the 32 genes, the Fifth Ewing Variant (FEV mRNA expression was analyzed and correlated to clinical outcomes in PCa patients who underwent prostatectomy at the Massachusetts General Hospital. Specimens from tissue microarray (TMA) including 102 prostate cancer patients were analysis for the expression of FEV. Meanwhile, FEV expression profiles were also assessed in PCa cell lines and in BPH-1 prostate epithelial cells using western blotting and quantitative reverse transcription-PCR (qRT-PCR). Furthermore, we transfected LNCaP and PC-3 cells with either an empty vector or full-length FEV gene and performed in vitro cell functional assays. The part FEV plays in tumor xenograft growth was also assessed in vivo.Of the 191 patients included in this study base on the DASL dataset, 77 (40.3%) and 24 (13.6%), respectively, developed prostate-specific antigen (PSA) relapse and metastasis postradical prostatectomy. Significant FEV downregulation was observed in PCa patients showing PSA failure and metastasis. The protein expression of FEV was significantly negatively correlated with the Gleason score and pathological stage in prostate cancer tissues. Similarly, FEV expression significantly decreased in all PCa cell lines relative to BPH-1 (allThe results of this study suggest an association between downregulated FEV expression and PSA relapse in PCa patients. In addition, FEV may act as a tumor suppressor in PCa.

Background Breast cancer is the most common cause of cancer-related death among women, and prognosis is especially poor for patients with triple-negative breast cancer (TNBC); therefore, there is an urgent need for new effective therapies. Recent studies have demonstrated that the uracil auxotroph Toxoplasma gondii vaccine displays anti-tumor effects. Here, we examined the immunotherapy effects of an attenuated uracil auxotroph strain of T. gondii against 4T1 murine breast cancer. Methods We constructed a uracil auxotroph T. gondii RH strain via orotidine 5′-monophosphate decarboxylase gene deletion (RH-Δompdc) with CRISPR/Cas9 technology. The strain’s virulence in the T. gondii-infected mice was determined in vitro and in vivo by parasite replication assay, plaque assay, parasite burden detection in mice peritoneal fluids and survival analysis. The immunomodulation ability of the strain was evaluated by cytokine detection. Its anti-tumor effect was evaluated after its in situ inoculation into 4T1 tumors in a mouse model; the tumor volume was measured, and the 4T1 lung metastasis was detected by hematoxylin and eosin and Ki67 antibody staining, and the cytokine levels were measured by an enzyme-linked immunosorbent assay. Results The RH-Δompdc strain proliferated normally when supplemented with uracil, but it was unable to propagate without the addition of uracil and in vivo, which suggested that it was avirulent to the hosts. This mutant showed vaccine characteristics that could induce intense immune responses both in vitro and in vivo by significantly boosting the expression of inflammatory cytokines. Inoculation of RH-Δompdc in situ into the 4T1 tumor inhibited tumor growth, reduced lung metastasis, promoted the survival of the tumor-bearing mice and increased the secretion of Th1 cytokines, including interleukin-12 (IL-12) and interferon-γ (INF-δ), in both the serum and tumor microenvironment (TME). Conclusion Inoculation of the uracil auxotroph RH-Δompdc directly into the 4T1 tumor stimulated anti-infection and anti-tumor immunity in mice, and resulted in inhibition of tumor growth and metastasis, promotion of the survival of the tumor-bearing mice and increased secretion of IL-12 and IFN-γ in both the serum and TME. Our findings suggest that the immunomodulation caused by RH-Δompdc could be a potential anti-tumor strategy. Graphical abstract

Min Xu,1,2,* Chong Yao,3,* Wei Zhang,1 Shen Gao,1 Hao Zou,1 Jing Gao1,4 1School of Pharmacy, Second Military Medical University, Shanghai, 200433, People’s Republic of China; 2Department of Pharmacy, Changzheng Hospital, Shanghai, 200003, People’s Republic of China; 3Clinical Pharmacy Center, Department of Pharmacy, Chinese PLA General Hospital, Beijing, 100850, People’s Republic of China; 4State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing, 100850, People’s Republic of China*These authors contributed equally to this workCorrespondence: Jing Gao; Hao ZouSchool of Pharmacy, Second Military Medical University, No. 325 Guohe Road, Yangpu District, Shanghai, People’s Republic of ChinaTel +86-10-6687-4665Email gjsmmu@126.com; haozou@smmu.edu.cnPurpose: Nanocarriers, with a high drug loading content and good safety, to achieve desirable therapeutic effect are always the goals for industry and research.Methods and Results: In the present study, we developed a docetaxel loaded poly-2-oxazoline polymer micellar system which employed poly-2-butyl-2 oxazoline and poly-2-methyl-2 oxazoline as the hydrophobic chain and hydrophilic chain, respectively. This micellar system achieves a high load up to 25% against the docetaxel, and further demonstrates an IC50 as low as 40% of the commercialized docetaxel injection in vitro and a double maximum tolerated dose in MCF-7 cells in vivo.Conclusion: The high drug loading content, superior safety, and considerable anti-cancer activity make this newly developed docetaxel loaded poly(2-oxazoline) micelle go further in future clinical research.Keywords: poly(2-oxazoline)s, micelle, docetaxel, high loading, anti-cancer activity

Background: The present study aimed to investigate the prognostic value of the neutrophil-to-lymphocyte ratio (NLR) in distal cholangiocarcinoma (DCC) following radical surgery. Methods: The clinicopathological data of 59 patients with DCC were retrospectively reviewed. Patients were treated by radical surgery and diagnosed by postoperative pathology at the Second Affiliated Hospital of Kunming Medical University (Yunnan, China), between July 2015 and December 2017. The optimal cut-off value for the NLR was determined by generating receiver operating characteristic (ROC) curves. Kaplan–Meier survival analysis and Cox proportional hazards models were used to determine the risk factors and independent risk factors influencing the prognosis of patients with DCC. Results: According to the ROC curve, the optimal cut-off value for the NLR was 2.933. The results of Kaplan–Meier survival analysis and the Cox proportional hazards model showed that carbohydrate antigen 125, NLR, perineural, vascular and fat invasion, regional lymph node metastasis, and the American Joint Committee on Cancer stage were risk factors for DCC; the only independent risk factor to affect the prognosis of DCC patients was the NLR. Conclusions: The preoperative NLR plays an important guiding role in evaluating the prognosis of patients with DCC, and an increase in the NLR is associated with poor patient prognosis.

Given the healthcare costs associated with obesity (especially in childhood), governments have tried several fiscal and policy interventions such as lowering tax and giving rebates to encourage parents to choose healthier food for their family. The efficacy of such fiscal policies is currently being debated. Here we address this issue by investigating how behavioral and brain-based responses in parents with low socioeconomic status change when rebates and lower taxes are offered on healthy food items. We performed behavioral and brain-based experiments, with the latter employing electroencephalography (EEG) acquired from parents while they shop in a simulated shopping market as well as follow up functional magnetic resonance imaging (fMRI) in the more restricted scanner environment. Behavioral data show that lower tax and rebate on healthy foods increase their purchase significantly compared to baseline. Rebate has a higher effect than lower tax treatment. From the EEG and fMRI experiments, we first show that healthy/unhealthy foods elicit least/maximal reward response in the brain, respectively. Further, by offering lower tax or rebate on healthy food items, the reward signal for such items in the brain is significantly enhanced. Second, we demonstrate that rebate is more effective than lower tax in encouraging consumers to purchase healthy food items, driven in part, by higher reward-related response in the brain for rebate. Third, fiscal interventions decreased the amount of frontal cognitive control required to buy healthy foods despite their lower calorific value as compared to unhealthy foods. Finally, we propose that it is possible to titrate the amount of tax reductions and rebates on healthy food items so that they consistently become more preferable than unhealthy foods.

Toxoplasma gondii are obligate intracellular protoza, and due to their small genome and limited encoded proteins, they have to exploit host factors for entry, replication, and dissemination. Such host factors can be defined as host dependency factors (HDFs). Though HDFs are inessential for cell viability, they are critical for pathogen infection, and potential ideal targets for therapeutic intervention. However, information about these HDFs required by T. gondii infection is highly deficient. In this study, the genes of human foreskin fibroblast (HFF) cells were comprehensively edited using the lentiviral CRISPR-Cas9-sgRNA library, and then the lentivirus-treated cells were infected with T. gondii at multiplication of infection 1 (MOI = 1) for 10 days to identify HDFs essential for T. gondii infection. The survival cells were harvested and sent for sgRNA sequencing. The sgRNA sequence matched genes or miRNAs were potential HDFs. Some cells in the lentivirus-treated group could survive longer than those in the untreated control group after T. gondii infection. From a pool of 19,050 human genes and 1,864 human pri-miRNAs, 1,193 potential HDFs were identified, including 1,183 genes and 10 pri-miRNAs (corresponding with 17 mature miRNAs). Among them, seven genes and five mature miRNAs were validated with siRNAs, miRNA inhibitors, and mimics, respectively. Bioinformatics analysis revealed that, among the 1,183 genes, 53 potential HDFs were associated with regulation of host actin cytoskeleton and 23 potential HDFs coded immune negative regulators. This result indicated that actin dynamics were indispensable for T. gondii infection, and some host immune negative regulators may be involved in disarming host defenses. Our findings contribute to the current limited knowledge about host factors required by T. gondii infection and provide us with new targets for medication therapy and vaccine exploitation.

To mitigate climate change, the governments of various countries have formulated and implemented corresponding low-carbon emission reduction policies. Meanwhile, consumers’ awareness of the necessity of environmental protection is gradually improving, and more consumers pay attention to the environmental attributes of products, which all encourages enterprises to have great power to implement low carbon technology. As rational decision makers, members tend to show the characteristics of risk aversion. How to meet the needs of consumers and reduce their own risks has become a key point of low-carbon supply chain management. Considering carbon quota policy, in this paper, the optimal pricing decision-making process of a supply chain system is discussed under risk-neutral and risk-avoidance decision-making scenarios by game theory, and a cost-sharing contract is used to coordinate the decision-making process of a supply chain system. By analyzing the influence of the risk aversion coefficient on the optimal strategies of participants, we find that when the manufacturer has the risk aversion characteristic, the risk aversion coefficient will further reduce the carbon emission rate, the wholesale price of the product and the manufacturer’s profit but increase the product order quantity and the retailer’s profit. In addition, if consumers have a high preference for low-carbon products, the manufacturer’s risk-aversion coefficient will lead to a lower selling price than in the centralized decision-making situation, and the profit of the supply chain system will also be further reduced. When the cost-sharing contract is adopted for coordination, the Pareto improvement of supply chain members’ profits can be achieved when the parameters of the cost-sharing contract are appropriate, regardless of the manufacturer’s risk-neutral decision or risk-aversion decision.

Dazhong Chen,1–3,* Xiaoli Pan,4,* Fangyuan Xie,5,* Ying Lu,2 Hao Zou,2 Chuan Yin,6 Yu Zhang,7 JieGao1,2 1Department of Gastrointestinal Surgery, The Second Affiliated Hospital of Wenzhou Medical University, Wenzhou 325027, China; 2Department of Pharmaceutical Sciences, Second Military Medical University, Shanghai 200433, China; 3Department of Planning, Kunming General Hospital of Chengdu Military Command, Yunnan 650032, China; 4Division of Gastroenterology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China; 5Department of Pharmacy, Shanghai Eastern Hepatobiliary Surgery Hospital, Shanghai 200438, China; 6Department of Gastroenterology, Changzheng Hospital, Second Military Medical University, Shanghai 200003, China; 7Department of Oncology, Xiangyang Central Hospital, Affiliated Hospital of Hubei University of Arts and Science, Hubei 441000, China *These authors contributed equally to this work Purpose: Liver cancer is the third leading cause of cancer-related deaths worldwide. Liver cancer stem cells (LCSCs) are a subpopulation of cancer cells that are responsible for the initiation, progression, drug resistance, recurrence, and metastasis of liver cancer. Recent studies have suggested that the eradication of both LCSCs and liver cancer cells is necessary because the conversion of cancer stem cells (CSCs) to cancer cells occasionally occurs. As ATP-binding cassette (ABC) transporters are overexpressed in both CSCs and cancer cells, combined therapies using ABC transporter inhibitors and chemotherapy drugs could show superior therapeutic efficacy in liver cancer. In this study, we developed poly(lactide-co-glycolide)/d-alpha-tocopherol polyethylene glycol 1000 succinate nanoparticles to accomplish the simultaneous delivery of an optimized ratio of doxorubicin (DOX) and elacridar (ELC) to target both LCSCs and liver cancer cells.Methods: Median-effect analysis was used for screening of DOX and ELC for synergy in liver cancer cells (HepG2 cells) and LCSCs (HepG2 tumor sphere [HepG2-TS]). Then, nanoparticles loaded with DOX and ELC at the optimized ratio (NDEs) were prepared by nanoprecipitation method. The cytotoxicity and colony and tumor sphere formation ability of nanoparticles were investigated in vitro, and the tissue distribution and antitumor activity of nanoparticles were evaluated in vivo.Results: We demonstrated that a DOX/ELC molar ratio of 1:1 was synergistic in HepG2 cells and HepG2-TS. NDEs were shown to exhibit significantly increased cytotoxic effects against both HepG2 and HepG2-TS compared with DOX-loaded nanoparticles (NDs) or ELC-loaded nanoparticles (NEs) in vitro. In vivo studies demonstrated that the nanoparticles exhibited better tumor targeting, with NDE showing the strongest antitumor activity with lower systemic toxicity.Conclusion: These results suggested that NDE represented a promising combination therapy against liver cancer by targeting both liver cancer cells and CSCs. Keywords: combined therapy, cancer stem cells, liver cancer, doxorubicin, elacridar, nanoparticles&nbsp

Orbital blow out fracture is a common disease in emergency department and a delay or failure in diagnosis can lead to permanent visual changes. This study aims to evaluate the ability of an automatic orbital blowout fractures detection system based on computed tomography (CT) data.Orbital CT scans of adult orbital blowout fractures patients and normal cases were obtained from Shanghai Ninth People's Hospital between January and March 2017. The region of fractures was annotated using 3D Slicer. The Inception V3 convolutional neural networks were constructed utilizing the Python programming language with PyTorch as the framework to extract high dimension features from each slice in a CT scan. These extracted features are processed through a XGBoost model to make the final differentiation of fracture cases and nonfracture ones. Accuracy, receiver operating characteristics, and area under the curve were evaluated.This study used 94 CT scans diagnosed with orbital blowout fractures and 94 healthy control cases. The automatic detection system showed accuracy of 92% in single-image classification and 87% in patient level detection. The area under the receiver operating characteristic curve was 0.9574.Using a deep learning-based automatic detection system of orbital blowout fracture can accurately detect and classify orbital blowout fractures from CT scans. The convolutional neural networks model combined with an accurate annotation system could achieve good performance in a small dataset. Further studies with large and multicenter data are required to refine this technology for possible clinical applications.

Background/Aims: Doublecortin-like kinase 1 (DCLK1) is emerging as a tumor-specific stem cell marker in pancreatic cancer (PC). MicroRNA-195 (miR-195) plays an important role in many types of tumors. However, the roles of DCLK1 in cancer and miRNAs that directly regulate DCLK1 have not been elucidated. The goal of this study is to assess the effects of miR-195 on inhibiting DCLK1 and to clarify the regulating mechanism of miR-195-DCLK1 in PC cells. Methods: The expression of DCLK1 protein and miR-195 in PC tissues and adjacent healthy pancreatic tissues was detected by Western blot and quantitative reverse transcription polymerase chain reaction (qRT-PCR), respectively and the correlation between overall survival of PC patients and expression of DCLK1 was measured by Kaplan-Meier analysis. Bioinformatics tools were used to identify the target gene of miR-195. Effects of miR-195 and DCLK1 on proliferation and cell cycle of PC cells were analyzed by MTT, colony formation assays and flow cytometry. Transwell and wound-healing experiments were employed to examine the cellular migration and invasion. A xenograft mouse model was also used to test the effects of miR-195 on tumor growth and metastasis in vivo. Results: The expression level of DCLK1 and miR-195 shows an inverse correlation in PC tissues and cell lines. A higher DCLK1 level is associated with higher TNM (tumor, node, and metastasis) stage, higher rate of lymph node metastasis, and poor survival. Luciferase reporter assay shows that miR-195 directly targets DCLK1. Overexpression of miR-195 inhibits proliferation, migration and invasion of PC cells, whereas downregulation of miR-195 has an opposite role. These actions were similar to the effects of knockdown and overexpression of DCLK1, respectively. Conclusions: These data suggest that miR-195 has tumor suppressor roles in PC by targeting DCLK1. MiR-195-DCLK1 pathway may provide insight into PC progression and represent a novel, promising diagnostic and therapeutic target for PC.

Recent reports show that B7-H4 is highly expressed in a variety of tumor cells, functions as a negative regulator of T cells and then promotes tumor progression. However, its expression and role in intrahepatic cholangiocarcinoma (ICC) remain unclear. In present study, B7-H4 expression in ICC and peritumoral tissues was determined at the level of mRNA and protein, and its bioactivity in ICC cells was studied after modification of B7-H4 expression. Then, the mechanism related to tumor progression induced by B7-H4 expression in ICC cells was explored. Finally, clinical significance of B7-H4 expression in ICC patients was further analyzed. The results showed that B7-H4 expression in ICC was much higher than that in peritumoral tissues at the level of both mRNA and protein. The high level of B7-H4 in ICC cells induced epithelial-to-mesenchymal transitions and promoted invasion and metastasis of tumor cells through activation of ERK1/2 signaling. The elevated B7-H4 expression was associated with the downregulated Bax, upregulated Bcl-2 expression, and activation of caspase-3. Clinically, high B7-H4 expression in tumor samples was significantly related to malignant phenotype, such as lymph node metastasis, high tumor stage, and poor differentiation. ICC patients with high expression of B7-H4 had shorter overall survival (OS) and disease-free survival. Moreover, the B7-H4 expression was an independent prognostic factor for predicting OS and tumor recurrence of ICC patients after operation. In conclusion, high expression of B7-H4 promotes tumor progression of ICC and may be a novel therapeutic target for ICC patients.

Aim: This article aims to explain the necrosis mechanisms of cancer cells specifically induced by gold nanorods (GNRs). Methods: The intracellular route and location of GNRs, the interaction between GNRs and lysosome, lysosome damage, cathepsin B release, necrosis complex formation, receptor-interacting protein 1 and TNF-α expression were systematically investigated. Results: The GNRs with serum corona were internalized quickly by cancer cells and finally taken up by lysosomes. The GNRs damaged the lysosomal membrane, resulting in the leakage of cathepsin B, which promoted the activation of receptor-interacting protein 1 and necrosomes formation. Necrotic cells and their debris or ill cellular contents were engulfed by macrophages resulting in high-level release of TNF-α, which further confirmed necrosis. Conclusion: GNRs can specifically trigger lysosome-dependent necrosis in cancer cells.

YY1 is a DNA-binding transcription factor and reported to be involved in cancer progression. Histone deacetylase inhibitor (HDACi) could inhibit proliferation and promote apoptosis of Hepatocellular carcinoma (HCC) cells. However, it is unclear about the roles of YY1 in the sensitivity of HCC cells to HDACi. In this study, firstly, we identified two drug-response profiles to HDACi in HCC cell lines, while our results showed that HDAC1 expression was positively correlated with YY1 in HCC cell lines and primary tumor tissues. Secondly, YY1 decreased the sensitivity of HCC cells to HDACi in vitro and in vivo. Furthermore, we found that YY1 promoted HDAC1 expression by binding to its promoter, while HDAC1 in turn up-regulated the expression of YY1. In conclusion, our results showed that YY1 could reduce the sensitivity of HCC cells to HDACi and might be a potential therapeutic target in HCC.

Aim: The application of cationic liposomes (CLs) as nonviral vectors is hampered by their cellular toxicity. Thus we aim to investigate the mechanisms underlying the cellular toxicity of CLs. Materials & methods: The effect of CLs on the autophagic flux, autophagosome-lysosome fusion, lysosome membrane permeabilization and cell necrosis of liver cells was investigated. Results & conclusion: Our results reveal a novel mechanism of CL-induced cell necrosis involving the induction of lysosome membrane permeabilization and late-stage autophagic flux inhibition that resulted in cytoplasmic release of cathepsin B, mitochondrial dysfunction and reactive oxygen species production, which are the key mediators of cell necrosis. Our study is important for revealing the cellular toxicity of CLs and designing safer gene delivery systems.

Toxoplasma gondii is an intracellular parasite that infects humans and other warm-blooded animals. Exosomes are endocytic-derived vesicles released by cells, representing an important mode of intercellular communication. In exosomes, specific molecules of proteins, lipids, and mRNAs or miRNAs have been detected, some of which are capable of transferring biologically active molecules to recipient cells. Dendritic cells (DCs) are the only antigen-presenting cells (APCs) that activate the initial immune response. In this study, high-throughput sequencing was used to analyze the exosomal miRNA profile of DC2.4 cells infected with Toxoplasma gondii for 28 h, compared with those of uninfected DC2.4 cells. Differential exosomal miRNAs (DEmiRs) from these two cell groups were analyzed. Through high-throughput sequencing, 3434 DEmiRs were obtained, and 12 stably enriched DEmiRNAs were verified by Reverse Transcription-quantitative Polymerase Chain Reaction (RT-qPCR) and selected for further analysis. The target genes of these 12 miRNAs were predicted with online analysis software and subjected to bioinformatics analyses including protein&ndash, protein interaction (PPI) network analysis, key driver analysis (KDA), gene ontology (GO) enrichment, and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis. These DEmiRs were found to be associated with a variety of biological processes and signaling pathways involved in host ubiquitin system, innate immunity, biosynthesis, and transferase activity and could be potential biomarkers for T. gondii infection.

Irisin is a circulating myokine induced by exercise, which is a cleaved version of fibronectin type III domain containing protein 5 (FNDC5). It can promote the browning of white fat tissue, increase energy consumption, and decrease weight. Irisin plays an important role in the regulation of various diseases, such as diabetes and coronary heart disease. Different types of exercise have different effects on irisin level in blood circulation, and moderate exercise can reduce cardiovascular symptoms. In this paper, the cardiovascular protective effect of irisin and its research progress in the field of exercise are reviewed, hoping to provide a new target for the prevention and treatment of cardiovascular diseases.

Nanotechnology has been materialized as a proficient technology for the development of anticancer nanoparticles all the way through an environment-friendly approach. Conventionally, nanoparticles have been assembled by dissimilar methods, but regrettably rely on the negative impact on the natural environment. Amalgamation of nanoparticles by means of plant extract is alternate conservative methods. In the present study, we equipped gold nanoparticles (AuNPs) from Strychni semen; displayed as a less toxic and environment-friendly. Integration of AuNPs was famed by UV-absorbance which displays peak values. Moreover, high-resolution transmission electron microscopy (HR-TEM), energy dispersive X-ray analysis (EDX) and atomic force microscopy (AFM) substantiate the shape of the AuNPs in the combined materials. FTIR results exhibit the active molecules positioned in the flat surface of the AuNPs. Similarly, the anticancer effectiveness of AuNPs is considered in KMCH-1 cells. Also, AuNPs successfully aggravate cytotoxicity and apoptosis by conjugating apoptotic gene expressions in KMCH-1 cells. Eventually, our results confirm the synthesis of AuNPs from Strychni Semen shows anticancer effects with environment-friendly manner.

Emerging evidence has shown that leptin, an adipocyte-derived cytokine that is closely associated with obesity, play a significant role in carcinogenesis and tumorigenesis. However, its impact on gallbladder cancer (GBC) remains unclear. In this study, we firstly found that leptin and its functional receptor OB-Rb were significantly co-expressed in human GBC tissues and cell lines, the content of which were higher than those in normal human gallbladder tissues. Treatment with leptin promoted the proliferation, migration and invasion of GBC cells, which were attenuated by OB-Rb shRNA. Blocking in the G2/M period of cell cycle, increasing of MMP3 and MMP9, increasing of VEGF-C/D, activation of SOCS3/JAK2/p-STAT3 pathway was demonstrated after treatment with leptin. All of these positive responses were attenuated by OB-Rb receptor shRNA. Taken together, our findings suggest that leptin promoted the proliferation, migration and invasion of GBC cells by increasing OB-Rb expression through the SOCS3/JAK2/p-STAT3 signal pathway. Targeting the leptin/OB-Rb axis could be an attractive therapeutic strategy for treatment of GBC.

The long noncoding RNA plasmacytoma variant translocation 1 gene (LncRNA PVT1) has an important role in tumor occurrence and development, yet the role and underlying molecular mechanisms of this RNA in cervical cancer have not yet been elucidated. In the present study, three cervical cancer cell lines (HeLa, Ca Ski and SiHa) were used to verify how LncRNA PVT1 mediates cervical cancer development, and the H8 cell line was used as a control. A LncRNA PVT1 overexpression vector or small interfering RNAs targeting LncRNA PVT1 were transfected into cervical cancer cells to generate LncRNA PVT1 overexpression and silencing in these cells. LncRNA PVT1 overexpression accelerated the growth of cervical cancer cells by advancing the cell cycle and inhibiting cellular apoptosis; increases in Cyclin D1 (CCND1) mRNA and activated Bcl‑2 protein expression levels also supported this finding. Furthermore, NF‑κB activation and expression was increased by LncRNA PVT1 overexpression. In addition, NF‑κB activation or inhibition induced changes in cell viability, accompanied by changes in CCND1 and Bcl‑2 expression. Increases or decreases in microRNA‑16 (miR‑16) expression (using miR mimics and inhibitors) also corresponded to changes in LncRNA PVT1 expression, in vitro. miR‑16 mimics and inhibitor had opposite effects to those of NF‑κB activity, and miR‑16 was demonstrated to directly interact with the NF‑κB gene as measured using the dual‑luciferase assay. In summary, LncRNA PVT1 inhibits the effect of miR‑16, promoting the cell cycle and inhibiting cellular apoptosis of cervical cancer cells, potentially via the NF‑κB pathway. The data from the present study will contribute to the current knowledge surrounding the theoretical basis of cervical cancer and provide a new perspective for the treatment of cervical cancer.

The incidence of intrahepatic cholangiocarcinoma (ICC) is progressively increasing worldwide, and its prognosis remains poor. Accumulating evidence has demonstrated that tumor necrosis factor receptor-associated factor 4 (TRAF4), an adaptor protein, is involved in the carcinogenesis and progression of several tumor types. However, the function of TRAF4 in predicting prognosis, and mediating migration and invasion of ICC remains to be elucidated. In the present study, immunohistochemistry, western blotting and reverse transcription-quantitative polymerase chain reaction assays were used to determine that the expression of TRAF4 at the mRNA and protein levels in ICC tissues was significantly higher compared with that in non‑tumor tissues. The overexpression of TRAF4 was positively correlated with poor differentiation, regional lymphatic metastasis, and high tumor‑node-metastasis staging. Inhibiting the expression of TRAF4 using small interfering RNA decreased the migration and invasion of ICC cells in vitro. In addition, the AKT inhibitor perifosine eliminated the effect of TRAF4 on the invasion and migration of ICC cells in vitro. Clinically, the overexpression of TRAF4 was correlated with shorter overall survival rate and elevated recurrence rate in patients with ICC. Furthermore, patients with ICC with a high expression of TRAF4 and lymphatic metastasis were closely associated with a poorer prognosis compared with the other groups. Multivariate analysis indicated that the overexpression of TRAF4 was an independent prognostic indicator for patients with ICC. It was identified that a high level of TRAF4 facilitated the invasiveness of ICC cells via the activation of AKT signaling. The overexpression of TRAF4 may be a prognostic biomarker and candidate therapeutic target for patients with ICC.

Background: Paclitaxel (PTX) could inhibit the growth of fibroblasts, which occurs in proliferative cholangitis and leads to biliary stricture. However, its use has been limited due to poor bioavailability and local administration for short time. This study designed and synthesized a new PTX-conjugated chitosan film (N-succinyl-hydroxyethyl chitosan containing PTX [PTX-SHEC]) and evaluated its safety and efficiency using in vivo and in vitro experiments. Methods: The SHEC conjugated with PTX was confirmed by nuclear magnetic resonance (NMR) and Fourier-transform infrared spectroscopy (FT-IR) measurements. Drug releases in vitro and in vivo were determined using high-performance liquid chromatography. Cell viability in vitro was measured using 3-（4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide. Rabbit biliary stricture model was constructed. All rabbits randomly divided into five groups (n = 8 in each group): the sham-operated rabbits were used as control (Group A), Groups B received laparotomies and suture, Group C received laparotomies and covered SHEC suture without the PTX coating, Group D received laparotomies and covered PTX-SHEC suture, and Group E received laparotomies and 1000 μmol/L PTX administration. Liver function tests and residual dosage of PTX from each group were measured by enzyme-linked immunosorbent assay. Histological data and α-smooth muscle actin (SMA) immunohistochemical staining of common bile duct were examined. Results: NMR and FT-IR indicated that PTX was successfully introduced, based on the appearance of signals at 7.41–7.99 ppm, 1.50 ppm, and 1.03 ppm, due to the presence of aromatic protons, methylene protons, and methyl protons of PTX, respectively. No bile leak was observed. The PTX-conjugated film could slowly release PTX for 4 weeks (8.89 ± 0.03 μg at day 30). The in vitro cell viability test revealed significantly different levels of toxicity between films with and without PTX (111.7 ± 4.0% vs. 68.1 ± 6.0%, P> 0.001), whereas no statistically significant difference was observed among the three sets of PTX-contained films (67.7 ± 5.4%, 67.2 ± 3.4%, and 59.1 ± 6.0%, P< 0.05). Histological examinations revealed that after 28 days of implantment, Groups D and E (but not Group C) had less granulation tissue and glandular hyperplasia in the site of biliary duct injury than Group B. The pattern was more obvious in Group D than Group E. Less α-SMA-positive cells were found in tissue from Groups D and E. Comparing with Group E, the liver function was improved significantly in Group D, including total bilirubin (2.69 ± 1.03 μmol/L vs. 0.81 ± 0.54 μmol/L, P= 0.014), alanine aminotransferase (87.13 ± 17.51 U/L vs. 42.12 ± 15.76 U/L, P= 0.012), and alkaline phosphatase (60.61 ± 12.31 U/L vs. 40.59 ± 8.78 U/L, P< 0.001). Conclusions: PTX-SHEC film effectively inhibites the myofibroblast proliferation and extracellular matrix over-deposition during the healing process of biliary reconstruction. This original film might offer a new way for reducing the occurrence of the benign biliary stricture. Key words: Biliary Stricture; Chitosan; Paclitaxel; Proliferative Cholangitis; Slow-Releasing

Breast cancer stem cells (BCSCs) are implicated in the initiation and progression of breast cancer and are responsible for metastasis and recurrence. In this study, we attempted to simultaneously target differentiated breast cancer cells (DBCCs) and BCSCs by using a combination of docetaxel (DTX)- and sulforaphane (SFN)-loaded poly(D, L-lactide-coglycolide)/hyaluronic acid (PLGA-b-HA)-based nanoparticles. BCSCs were identified as having an ESA+CD44+CD24– phenotype, which exhibited docetaxel resistance. Drug-loaded nanoparticles exhibited enhanced cytotoxicity towards both DBCCs and BCSCs compared with free drugs. SFN-loaded nanoparticles were more effective in inhibiting BCSCs than free SFN in vitro by down-regulating β-catenin expression. In vivo analysis of anti-tumor activity showed that the combination therapy with DTX- and SFN-loaded nanoparticles had the strongest antitumor efficacy. In vivo analysis of anti-BCSCs activity showed that the self-renewal ability of BCSCs was strongly inhibited in DTX- and SFN-loaded nanoparticle-treated groups. In conclusion, the combination of SFN- and DTX-loaded PLGA-b-HA nanoparticles shows therapeutic potential in the treatment of breast cancer by simultaneously targeting DBCCs and BCSCs.

We present a strategy to discover recombinant monoclonal antibodies (mAbs) to specific cancers and demonstrate this approach using basal subtype breast cancers. A phage antibody library was depleted of antibodies to common cell surface molecules by incubation with luminal breast cancer cell lines, and then selected on a single basal-like breast cancer cell line (MDA-MB-231) for binding associated receptor-mediated endocytosis. Additional profiling against two luminal and four basal-like cell lines revealed 61 unique basal-specific mAbs from a pool of 1440 phage antibodies. The unique mAbs were further screened on nine basal and seven luminal cell lines to identify those with the greatest affinity, specificity, and internalizing capability for basal-like breast cancer cells. Among the internalizing basal-specific mAbs were those recognizing four transmembrane receptors (EphA2, CD44, CD73 and EGFR), identified by immunoprecipitation–mass spectrometry and yeast-displayed antigen screening. Basal-like breast cancer expression of these four receptors was confirmed using a bioinformatic approach, and expression microarray data on 683 intrinsically subtyped primary breast tumors. This overall approach, which sequentially employs phage display antibody library selection, antigen identification and bioinformatic confirmation of antigen expression by cancer subtypes, offers efficient production of high-affinity mAbs with diagnostic and therapeutic utility against specific cancer subtypes.

Swertia mussotii (Gentianaceae) is a traditional Chinese medicinal plant grown in the Qinghai-Tibet Plateau. Three fractions from S. mussotii extract, named SWF50, SWF 70 and SWF100, were screened for in vitro anti-proliferative activity on two gastric cancer cell lines, MGC-803 and BGC-823 cells using MTT assay. Our results demonstrated that SMF70 showed an anti-proliferative effect in MGC-803 cells and SMF100 showed an anti-proliferative effect in BGC-823 cells in vitro. Moreover, both two fractions induced apoptosis via depolymerization of cytoskeletal filaments, increased cytoplasmic levels of ROS and Ca2+ and disrupted mitochondrial transmembrane potential. In addition, flow cytometry analysis indicated that both two fractions could induce cell apoptosis and arrest the cell cycle at S phase. Our results indicate that SMF70 induces apoptosis of MGC-803 cells and SMF100 induces apoptosis of BGC-823 cells via a mitochondrial-dependent pathway. Meanwhile, we also investigated antitumor effect of SMF70 in vivo, and exhibited effective tumor growth inhibition. Our findings demonstrate that S. mussotii extracts could be a potential new alternative therapeutic agent gastric cancer.

Aim: Salinomycin (SAL)-loaded PEG-ceramide nanomicelles (SCM) were prepared to target both liver cancer cells and cancer stem cells. Materials & methods: The synergistic ratio of SAL/PEG-ceramide was evaluated to prepare SCM, and the antitumor activity of SCM was examined both in vitro and in vivo. Results: SAL/PEG-ceramide molar ratio of 1:4 was chosen as the synergistic ratio, and SCM showed superior cytotoxic effect and increased apoptosis-inducing activity in both liver cancer cells and cancer stem cells. In vivo, SCM showed the best tumor inhibitory effect with a safety profile. Conclusion: Thus, PEG-ceramide nanomicelles could serve as an effective and safe therapeutic drug carrier to deliver SAL into liver cancer, opening up the avenue of using PEG-ceramide as therapeutic drug carriers.

Background The Barcelona Clinic Liver Cancer (BCLC) staging system is widely used to classify hepatocellular carcinoma (HCC). This study is to investigate the prognostic factors for patients with BCLC stage A HCC after R0 hepatectomy. Methods A total of 592 patients with BCLC stage A HCC following R0 hepatectomy from 1997-2012 were enrolled in this study. Kaplan-Meier analysis and Cox regression were used to analyze the risk factors associated with recurrence. Receiver operating characteristic curves were used to establish a new scoring system to evaluate the independent risk factors for recurrence. Furthermore, subgroup analyses were performed to evaluate surgical margins on tumor recurrence between the anatomic and nonanatomic resection group. Results Independent risk factors for BCLC stage A HCC recurrence were preoperative alanine transaminase >40U/L, liver cirrhosis, surgical margin 5cm. Based on these 5 risk factors, we established a new scoring system in this study, named "HCC recurrence scoring system." Patients with a high score (≥3 points, 1 point for each factor) composed the high recurrence risk group. Moreover, the subgroup analyses demonstrated that different surgical margins had no significant effect on tumor recurrence in the anatomic resection group ( P = 0.408), while it had a significant effect in the nonanatomic resection group ( P = 0.000). Conclusions For patients with BCLC stage A with scores ≥3 points, close postoperative follow-up and positive measures to prevent recurrence are particularly important. Anatomic resection is preferred for patients with BCLC stage A. Adequate surgical margins are necessary for patients with poor liver function.

Scaffolds that can achieve cell adhesion and controlled release of protein drugs are very promising in bone tissue engineering. Due to their biocompatibility and injectablity, poly(lactide-co-glycolide acid) (PLGA) porous microspheres (PLGA-pMS) present potential scaffolds in bone tissue engineering. However, their application is hampered by the burst release of protein drugs and hydrophobicity that leads to poor cell adhesion. To overcome these drawbacks, we developed novel PLGA-pMS by incorporating bovine serum albumin (BSA) loaded chitosan microspheres (CS-MS) in Gly-Arg-Gly-Asp-Ser-Pro-Cys (GRGDSPC) modified PLGA-pMS (CS-MS/PLGA-pMS). GRGDSPC was used to enhance the hydrophilicity and cell affinity of the porous microspheres. Results showed that PLGA-pMS had a size of 446.77±19.46μm, with an average surface pore size of 21.56±3.02μm, whereas CS-MS had a size of 15.98±0.96μm and 16.35±0.38μm (5% and 10% TPP-prepared CS-MS, respectively). A scanning electron microscope (SEM) and a confocal laser scanning microscope (CLSM) revealed that CS-MS were partly embedded in the PLGA matrices and the integrity of CS-MS was retained. Thermogravimetry analyzer (TGA) also demonstrated that CS-MS were incorporated into PLGA-pMS. The CS-MS/PLGA-pMS had a size of 454.02±16.09μm, with a BSA encapsulation efficiency of 53.19±1.67% and 62.16±3.44% (5% and 10% TPP-prepared CS-MS, respectively). CS-MS/PLGA-pMS exhibited a sustained FITC-BSA release for 28 days. Modification of GRGDSPC significantly improved adhesion of MG-63 cells on the porous microspheres. In conclusion, CS-MS/PLGA-pMS may act as potential bifunctional scaffolds for bone tissue engineering.

Breast cancer stem cells (BCSCs), which can fully recapitulate the tumor origin and are often resistant to chemotherapy and radiotherapy, are currently considered as a major obstacle for breast cancer treatment. To achieve the goal of both targeting BCSCs and bulk breast cancer cells, we developed 8-hydroxyquinoline-loaded hyaluronan modified mesoporous silica nanoparticles (MSN)-supported lipid bilayers (HA-MSS) and docetaxel-loaded MSS. The results showed that the size of all the nanoparticles was smaller than 200 nm. BCSCs were enriched from MCF-7 cells by a sphere formation method and identified with the CD44(+)/CD24(-) phenotype. Quantitative and qualitative analysis demonstrated that HA promotes the uptake of HA-MSS in CD44-overexpressing MCF-7 mammospheres, revealing the mechanism of receptor-mediated endocytosis. DTX or DTX-loaded MSS showed much enhanced cytotoxicity against MCF-7 cells compared with MCF-7 mammospheres, whereas 8-HQ or 8-HQ-loaded HA-MSS showed much enhanced cytotoxicity against MCF-7 mammospheres compared with MCF-7 cells. In the MCF-7 xenografts in mice, the combination therapy with DTX-loaded MSS plus 8-HQ-loaded HA-MSS produced the strongest antitumor efficacy, with little systemic toxicity (reflecting by loss of body weight) in mice. Thus, this combination therapy may provide a potential strategy to improve the therapy of breast cancer by eradication of breast cancer cells together with BCSCs.

Aims The prognosis of patients with intrahepatic cholangiocarcinoma (ICC) remains poor in terms of overall survival (OS) and recurrence rate. Mortalin, a stress chaperone, has been reported to be involved in carcinogenesis and metastasis. However, its role in ICC has not been defined. Methods Mortalin expression in tumour samples from patients with ICC was examined by Western blot and immunohistochemistry, and correlation between its expression and clinicopathological features was assessed. In addition, invasion, migration proliferation and apoptosis, and the expression of epithelial-mesenchymal transition (EMT)-related markers in ICC cells were assessed after mortalin depletion. Finally, the prognostic significance of mortalin in patients with ICC was further evaluated by Kaplan–Meier and Cox regression analysis. Results We provide evidence that expression of mortalin in human ICC tissues is higher than that in matched peritumoural tissues. The interference of mortalin expression inhibited the proliferation and invasion of ICC cells in vitro. Mechanistically, inhibition of mortalin expression in ICC cells upregulated E-cadherin expression and decreased vimentin and snail expression. Clinically, a high level of mortalin in ICC samples was associated with loss of E-cadherin, and increased expression of vimentin and snail. Patients with ICC and high mortalin expression had a shorter OS and a higher recurrence rate. Multivariate analysis revealed that mortalin overexpression was an independent prognostic indicator for patients with ICC. Conclusions Mortalin may promote cell proliferation and invasion via induction of EMT of ICC cells. A high level of mortalin may be used as a prognostic biomarker and therapeutic target for patients with ICC.

Aim: To develop salinomycin-loaded nanoliposomes (SLN), doxorubicin-loaded nanoliposomes (DLN) and nanoliposomes codelivering salinomycin and doxorubicin (SDLN) to target both liver cancer cells and cancer stem cells. Materials & methods: The characterization and antitumor activity of SLN, DLN and SDLN were evaluated. Results & conclusion: The doxorubicin/salinomycin sodium mole ratio of 1:1 had the best synergistic combination index value, and was chosen as the drug ratio in SDLN. SDLN could maintain the drug ratio between 1:1 and 3:1 in 12 h in vivo. SDLN and SLN + DLN showed the best tumor inhibitory rate, and could significantly decrease the percentage of liver cancer stem cells in vivo. SDLN and SLN + DLN may serve as an effective approach to treat liver cancer.

We previously reported the development of PE38KDEL-loaded anti-HER2 poly(lactic-co-glycolic acid) (PLGA) nanoparticles that bind and internalize in HER2-overexpressing breast cancer cells, enabling potent anti-tumor activity. To overcome the problems associated with the short half-lives of this drug delivery system, we have constructed PE38KDEL-loaded anti-HER2 PEGylated liposomes (PE-HER-liposomes). PE-HER-liposomes were constructed with Fab′ of recombinant humanized anti-HER2 monoclonal antibody (anti-HER2 Fab′) covalently linked to PEGylated liposomes containing PE38KDEL (PE-liposomes). We attached anti-HER2 Fab′ to the terminus of PEG (polyethylene glycol) on PEGylated liposomes. Incorporation of pyridylthiopropionoylamino-PEG-distearoylphosphatidylethanolamine (PDP-PEG-DSPE) into PEGylated liposomes followed by mild thiolysis of the PDP groups resulted in the formation of reactive thiol groups at the periphery of the liposomes. Efficient attachment of maleimide-derivatized anti-HER2 Fab′ took place under mild conditions. The characterization of PE-HER-liposomes, such as particle size, was evaluated by dynamic light-scattering detector. The Micro BCA method was used to determine the encapsulation efficiency of PE38KDEL and the quantity of conjugated Fab′. Flow cytometry and confocal microscopy showed that PE-HER-liposomes possessed receptor-specific binding and internalization for HER2-overexpressing SK-BR3 cells. Remarkably, PE-HER-liposomes were more cytotoxic than non-targeted PE-liposomes in HER2-overexpressing breast cancer cells. In conclusion, PE-HER-liposomes could serve as a promising therapeutic candidate for the treatment of HER2-overexpressing breast cancers.

SummaryHuman serum potently induces hyphal development of the polymorphic fungal pathogen Candida albicans, a phenotype that contributes critically to infections. The fungal adenylyl cyclase Cyr1p is a key component of the cAMP/PKA-signaling pathway that controls diverse infection-related traits, including hyphal morphogenesis. However, identity of the serum hyphal inducer(s) and its fungal sensor remain unknown. Our initial analyses of active serum fractions revealed signs of bacterial peptidoglycan (PGN)-like molecules. Here, we show that several purified and synthetic muramyl dipeptides (MDPs), subunits of PGN, can strongly promote C. albicans hyphal growth. Analogous to PGN recognition by the mammalian sensors Nod1 and Nod2 through their leucine-rich-repeat (LRR) domain, we show that MDPs activate Cyr1p by directly binding to its LRR domain. Given the abundance of PGN in the intestine, a natural habitat and invasion site for C. albicans, our findings have important implications for the mechanisms of infection by this pathogen.

The objective of this work was to develop and evaluate a floating-pulsatile drug delivery system intended for chronopharmacotherapy. Floating-pulsatile concept was applied to increase the gastric residence of the dosage form having lag phase followed by a burst release. To overcome limitations of various approaches for imparting buoyancy, we generated the system which consisted of three different parts, a core tablet, containing the active ingredient, an erodible outer shell and a top cover buoyant layer. The dry coated tablet consists in a drug-containing core, coated by a hydrophilic erodible polymer which is responsible for a lag phase in the onset of pulsatile release. The buoyant layer, prepared with Methocel K4M, Carbopol 934P and sodium bicarbonate, provides buoyancy to increase the retention of the oral dosage form in the stomach. The effect of the hydrophilic erodible polymer characteristics on the lag time and drug release was investigated. Developed formulations were evaluated for their buoyancy, dissolution and pharmacokinetic, as well gamma-scintigraphically. The results showed that a certain lag time before the drug released generally due to the erosion of the dry coated layer. Floating time was controlled by the quantity and composition of the buoyant layer. Both pharmacokinetic and gamma-scintigraphic data point out the capability of the system of prolonged residence of the tablets in the stomach and releasing drugs after a programmed lag time.

Aims: To develop novel iRGD (internalizing Arg-Gly-Asp peptide)-conjugated DSPE-PEG2000 nanomicelles (M-SAL-iRGD) for delivery of salinomycin to both liver cancer cells and cancer stem cells (CSCs). Materials & methods: The characterization, antitumor activity and mechanism of action of M-SAL-iRGD were evaluated. Results & conclusion: M-SAL-iRGD possessed a small size of around 10 nm, and drug encapsulation efficacy higher than 90%. M-SAL-iRGD showed significantly increased cytotoxic effect toward both nontargeted M-SAL (salinomycin-loaded DSPE-PEG2000 nanomicelles) and salinomycin in both liver cancer cells and CSCs. The tissue distribution and antitumor assays in mice bearing liver cancer xenograft confirmed the superior penetration tumor efficacy and antitumor activity of M-SAL-iRGD. M-SAL-iRGD represent a potential effective nanomedicine against liver cancer.

Hepatocellular carcinoma (HCC) is a malignant tumor and threatens human life worldwide, whereas the etiology and pathogenesis of HCC have not been fully determined. In the past few years, many microRNAs (miRNAs) have been proved to have important roles in tumorigenesis of HCC. In this study, we found that miR-23b was significantly upregulated in tumor tissues of HCC patients. Functional tests showed that miR-23b could promote HCC cell proliferation and metastasis in vitro and in vivo. Then, mechanistic investigations suggested that ST7L was a direct target of miR-23b and involved in the promotion effects of miR-23b on HCC tumorigenesis and metastasis. Furthermore, our study indicated that ST7L could interact with the carboxyl terminal region of AKT and suppress AKT/GSK3β/β-catenin pathway in HCC cells. In conclusion, our study revealed important roles of miR-23b and ST7L in progression of HCC.

In this paper, a method for dose calculation with pencil beam kernels constructed by point kernel superposition was proposed to accelerate the dose calculation during intensity optimization iteration. With this method, the direct aperture optimization method can be integrated in the planning system based on point kernel convolution/superposition model. The dose calculation time was also reduced during the iteration. From the result of the phantom and clinical patient data test, it was concluded that this method could be used for the intensity optimization of iteration dose calculation as the satisfied precision due to the optimization result coherence obtained. By implementing the method in the planning system product based on point kernel convolution/superposition model, a lot of additional research and development works for the pencil beam dose calculation model as well as the product maintenance cost can be avoided.

Herpes simplex virus (HSV) and varicella zoster virus (VZV) are related members of the Herpesviridae family and are well-documented human pathogens causing a spectrum of diseases, from mucocutaneous disease to infections of the central nervous system. This study was carried out to evaluate and validate the performance of a multiplex real-time polymerase chain reaction (PCR) assay in detecting and differentiating HSV1, HSV2, and VZV from clinical samples. Consensus PCR primers for HSV were designed from the UL30 component of the DNA polymerase gene of HSV, with 2 separate hydrolysis probes designed to differentiate HSV1 and HSV2. Separate primers and a probe were also designed against the DNA polymerase gene of VZV. A total of 104 clinical samples were available for testing by real-time PCR, conventional PCR, and viral culture. The sensitivity and specificity of the real-time assay was calculated by comparing the multiplex PCR result with that of a combined standard of virus culture and conventional PCR. The sensitivity of the real-time assay was 100%, with specificity ranging from 98% to 100% depending on the target gene. Both PCR methods detected more positive samples for HSV or VZV, compared with conventional virus culture. This multiplex PCR assay provides accurate and rapid diagnostic capabilities for the diagnosis and differentiation of HSV1, HSV2, and VZV infections, with the presence of an internal control to monitor for inhibition of the PCR reaction.

Gallbladder cancer (GBC) is one leading cause of cancer-related death worldwide. WW domain-containing oxidoreductase (WWOX) is a tumor suppressor gene which can suppress proliferation of a variety of tumors. However, little was known about the relationships between WWOX and gallbladder cancer. In the current study, we intended to investigate the tumor suppressive role of WWOX in gallbladder malignant cells both in vitro and in vivo, and explore the potential mechanism of tumor toxic function of WWOX. Our results have shown that WWOX triggerred apoptosis in GBC cells and increased the expression of P73 and PUMA in cytoplasm. We also have found that Bax has been upregulated after overexpression of WWOX, whereas, Bcl-2 was downregulated by WWOX. To further validate the results in vivo, we evaluated the tumor suppressive role of WWOX in mouse model of gallbladder cancer. The results have shown that the proliferation of the tumor was inhibited after delivery of WWOX, and the expressions of P73 and PUMA were upregulated in target tissues. The mice models administrated with WWOX have shown better prognosis than mice in negative control groups. The results from our study indicated that WWOX could be used as a therapeutic agent in the gene therapy of gallbladder cancer.

Aberrant expression and activation of EGF receptor (EGFR) has been implicated in the development and progression of many human cancers. As such, targeted therapeutic inhibition of EGFR, for example by antibodies, is a promising anticancer strategy. The overall efficacy of antibody therapies results from the complex interplay between affinity, valence, tumor penetration and retention, and signaling inhibition. To gain better insight into this relationship, we studied a panel of EGFR single-chain Fv (scFv) antibodies that recognize an identical epitope on EGFR but bind with intrinsic monovalent affinities varying by 280-fold. The scFv were converted to Fab and IgG formats, and investigated for their ability to bind EGFR, compete with EGF binding, and inhibit EGF-mediated downstream signaling and proliferation. We observed that the apparent EGFR-binding affinity for bivalent IgG plateaus at intermediate values of intrinsic affinity of the cognate Fab, leading to a biphasic curve describing the ratio of IgG to Fab affinity. Mathematical modeling of antibody–receptor binding indicated that the biphasic effect results from nonequilibrium assay limitations. This was confirmed by further observation that the potency of EGF competition for antibody binding to EGFR improved with both intrinsic affinity and antibody valence. Similarly, both higher intrinsic affinity and bivalent binding improved the potency of antibodies in blocking cellular signaling and proliferation. Overall, our work indicates that higher intrinsic affinity combined with bivalent binding can achieve avidity that leads to greater in vitro potency of antibodies, which may translate into greater therapeutic efficacy. Mol Cancer Ther; 11(7); 1467–76. ©2012 AACR.