Preoperative blood markers and intra-abdominal infection after colorectal cancer resection.

BACKGROUND: Colorectal cancer (CRC) has one of the highest morbidity and mortality rates among digestive tract tumors. Intra-abdominal infection (IAI) is a common postoperative complication that affects the clinical outcomes of patients with CRC and hinders their rehabilitation process. However, the factors influencing abdominal infection after CRC surgery remain unclear; further, prediction models are rarely used to analyze preoperative laboratory indicators and postoperative complications. AIM: To explore the predictive value of preoperative blood markers for IAI after radical resection of CRC. METHODS: The data of 80 patients who underwent radical resection of CRC in the Anorectal Surgery Department of Suzhou Hospital affiliated with Anhui Medical University were analyzed. These patients were categorized into IAI (n = 15) and non-IAI groups (n = 65) based on whether IAI occurred. Influencing factors were compared; general data and laboratory indices of both groups were identified. The relationship between the indicators was assessed. Further, a nomogram prediction model was developed and evaluated; its utility and clinical applicability were assessed. RESULTS: The risk factors for IAI after radical resection of CRC were neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), systemic immune-inflammation index (SII), and carcinoembryonic antigen (CEA) levels. NLR was correlated with PLR and SII (r = 0.604, 0.925, and 0.305, respectively), while PLR was correlated with SII (r = 0.787). The nomogram prediction model demonstrated an area under the curve of 0.968 [95% confidence interval (CI): 0.948-0.988] in the training set (n = 60) and 0.926 (95%CI: 0.906-0.980) in the validation set (n = 20). The average absolute errors of the calibration curves for the training and validation sets were 0.032 and 0.048, respectively, indicating a good model fit. The decision curve analysis curves demonstrated high net income above the 5% threshold, indicating the clinical practicality of the model. CONCLUSION: The nomogram model constructed using NLR, PLR, SII, and CEA levels had good accuracy and reliability in predicting IAI after radical resection of CRC, potentially aiding clinical treatment decision-making.

Lack of evolutionary convergence in multiple primary lung cancer suggests insufficient specificity of personalized therapy.

Multiple primary lung cancer (MPLC) is an increasingly prevalent subtype of lung cancer. According to recent genomic studies, the different lesions of a single MPLC patient exhibit functional similarities that may reflect evolutionary convergence. We perform whole-exome sequencing for a unique cohort of MPLC patients with multiple samples from each lesion found. Using our own and other relevant public data, evolutionary tree reconstruction reveals that cancer driver gene mutations occurred at the early trunk, indicating evolutionary contingency rather than adaptive convergence. Additionally, tumors from the same MPLC patient are as genetically diverse as those from different patients, while within-tumor genetic heterogeneity is significantly lower. Furthermore, the aberrant molecular functions enriched in mutated genes for a sample show a strong overlap with other samples from the same tumor, but not with samples from other tumors or other patients. Overall, there is no evidence of adaptive convergence during the evolution of MPLC. Most importantly, the similar between-tumor diversity and between-patient diversity suggest that personalized therapies may not adequately account for the genetic diversity among different tumors in an MPLC patient. To fully exploit the strategic value of precision medicine, targeted therapies should be designed and delivered on a per-lesion basis.

A Novel PD-L1-Containing MSLN Targeting Vaccine for Lung Cancer Immunotherapy.

Therapeutic tumor vaccines have become an important breakthrough in the treatment of various solid tumors including lung cancer. Dendritic cells (DCs)-based tumor vaccines targeting tumor-associated antigens (TAAs) play a key role in immunotherapy and immunoprevention. However, the weak immunogenicity of TAAs and low immune response rates are a major challenge faced in the application of therapeutic tumor vaccines. Here, we tested whether targeting an attractive target Mesothelin (MSLN) and PD-L1 immune checkpoint molecule to DCs in vivo would elicit therapeutic antitumor cytotoxic T lymphocyte (CTL) response. We generated specific MSLN fragment combined with PD-L1 and GM-CSF peptide immunogen (MSLN-PDL1-GMCSF) based on the novel anti-PD-L1 vaccination strategy we recently developed for the cancer treatment and prevention. We found that DCs loaded with MSLN-PDL1-GMCSF vaccine elicited much stronger endogenous anti-PD-L1 antibody and T cell responses in immunized mice and that antigen specific CTLs had cytolytic activities against tumor cells expressing both MSLN and PD-L1. We demonstrated that vaccination with MSLN-PDL1-GMCSF potently inhibited the tumor growth of MSLN+ and PD-L1+ lung cancer cells, exhibiting a significant therapeutic anti-tumor potential. Furthermore, PD-1 blockade further improved the synergistic antitumor therapeutic efficacy of MSLN-PDL1-GMCSF vaccine in immunized mice. In summary, our data demonstrated for the first time that this PD-L1-containing MSLN therapeutic vaccine can induce persistent anti-PD-L1 antibody and CTL responses, providing an effective immunotherapeutic strategy for lung cancer immunotherapy by combining MSLN-PDL1-GMCSF vaccine and PD-1 blockade.

Single-cell transcriptomes identifies characteristic features of mouse macrophages in liver Mallory-Denk bodies formation.

Mallory-Denk bodies (MDBs) consist of intracellular aggregates of misfolded proteins in ballooned hepatocytes and serve as important markers of progression in certain liver diseases. Resident hepatic macrophage-mediated inflammation influences the development of chronic liver diseases and cancer. Here, the first systematic study of macrophages heterogeneity in mice was conducted to illustrate the pathogenesis of MDB formation using single-nucleus RNA sequencing (snRNA-seq). Furthermore, we provided transcriptional profiles of macrophages obtained from the fractionation of mouse liver tissues following chronic injury. We equally identified seven discrete macrophage subpopulations, each involved in specific cellular activated pathways such as basal metabolism, immune regulation, angiogenesis, and cell cycle regulation. Among these, a specific macrophage cluster (Cluster4), a subpopulation specifically expressing genes that regulate cell division and the cell cycle, was identified. Interestingly, we found that CCR2 was significantly induced in Cluster2, thereby inducing monocytes to migrate to macrophages to promote MDB pathogenesis. Thus, our study is the first to demonstrate the heterogeneity of macrophages associated with liver MDB formation in mice through single-cell resolution. This serves as the basis for further insights into the pathogenesis of liver MDB formation and molecular mechanisms of chronic liver disease progression.

A Novel Therapeutic Tumor Vaccine Targeting MUC1 in Combination with PD-L1 Elicits Specific Anti-Tumor Immunity in Mice.

Dendritic cells (DCs), as professional antigen-presenting cells (APCs), play a key role in the initiation and regulation of humoral and cellular immunity. DC vaccines loaded with different tumor-associated antigens (TAAs) have been widely used to study their therapeutic effects on cancer. A number of clinical trials have shown that DCs are safe as an antitumor vaccine and can activate certain anti-tumor immune responses; however, the overall clinical efficacy of DC vaccine is not satisfactory, so its efficacy needs to be enhanced. MUC1 is a TAA with great potential, and the immune checkpoint PD-L1 also has great potential for tumor treatment. Both of them are highly expressed on the surface of various tumors. In this study, we generated a novel therapeutic MUC1-Vax tumor vaccine based on the method of PD-L1-Vax vaccine we recently developed; this novel PD-L1-containing MUC1-Vax vaccine demonstrated an elevated persistent anti-PD-L1 antibody production and elicited a much stronger protective cytotoxic T lymphocyte (CTL) response in immunized mice. Furthermore, the MUC1-Vax vaccine exhibited a significant therapeutic anti-tumor effect, which significantly inhibited tumor growth by expressing a high MUC1+ and PD-L1+ level of LLC and Panc02 tumor cells, and prolonged the survival of cancer-bearing animals. Taken together, our study provides a new immunotherapy strategy for improving the cross-presentation ability of therapeutic vaccine, which may be applicable to pancreatic cancer, lung cancer and for targeting other types of solid tumors that highly express MUC1 and PD-L1.

Characterization of the complete chloroplast genome of Achnatherum pekinense (Poaceae), a widespread weed.

Achnatherum pekinense belongs to Poaceae. The complete chloroplast genome of A. pekinense was reported in this study. The chloroplast genome was 137,837 bp in size with a canonical quadripartite structure, including two inverted repeat regions (IR) of 21,635 bp for each, a large single-copy (LSC) region of 81,787 bp in length, and a small single-copy (SSC) region of 12,780 bp in length. The overall guanine-cytosine (GC) content of this chloroplast genome was 38.8%, and the corresponding values of the LSC, SSC, and IR regions were 36.9%, 33.1%, and 44.1%, respectively. A total of 113 unique genes were annotated in this chloroplast genome, including four rRNA genes, 31 tRNA genes, and 78 protein-coding genes. The phylogenetic analysis showed that A. pekinense was clustered with A. inebrians.

Distinct Mitochondria-Mediated T-Cell Apoptosis Responses in Children and Adults With Coronavirus Disease 2019.

BACKGROUND: Lymphopenia is a key feature for adult patients with coronavirus disease 2019 (COVID-19), although it is rarely observed in children. The underlying mechanism remains unclear. METHODS: Immunohistochemical and flow cytometric analyses were used to compare the apoptotic rate of T cells from COVID-19 adults and children and apoptotic responses of adult and child T cells to COVID-19 pooled plasma. Biological properties of caspases and reactive oxygen species were assessed in T cells treated by COVID-19 pooled plasma. RESULTS: Mitochondria apoptosis of peripheral T cells were identified in COVID-19 adult patient samples but not in the children. Furthermore, increased tumor necrosis factor-α and interleukin-6 in COVID-19 plasma induced mitochondria apoptosis and caused deoxyribonucleic acid damage by elevating reactive oxygen species levels of the adult T cells. However, the child T cells showed tolerance to mitochondrial apoptosis due to mitochondria autophagy. Activation of autophagy could decrease apoptotic sensitivity of the adult T cells to plasma from COVID-19 patients. CONCLUSIONS: Our results indicated that the mitochondrial apoptosis pathway was activated in T cells of COVID-19 adult patients specifically, which may shed light on the pathophysiological difference between adults and children infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 ).

Continuous expression of reprogramming factors induces and maintains mouse pluripotency without specific growth factors and signaling inhibitors.

OBJECTIVES: Derivation and maintenance of pluripotent stem cells (PSCs) generally require optimized and complex culture media, which hinders the derivation of PSCs from various species. Expression of Oct4, Sox2, Klf4, and c-Myc (OSKM) can reprogram somatic cells into induced PSCs (iPSCs), even for species possessing no optimal culture condition. Herein, we explored whether expression of OSKM could induce and maintain pluripotency without PSC-specific growth factors and signaling inhibitors. METHODS: The culture medium of Tet-On-OSKM/Oct4-GFP mouse embryonic stem cells (ESCs) was switched from N2B27 with MEK inhibitor, GSK3ß inhibitor, and leukemia inhibitory factor (LIF) (2iL) to N2B27 with doxycycline. Tet-On-OSKM mouse embryonic fibroblast (MEF) cells were reprogrammed in N2B27 with doxycycline. Cell proliferation was traced. Pluripotency was assessed by expression of ESC marker genes, teratoma, and chimera formation. RNA-Seq was conducted to analyze gene expression. RESULTS: Via continuous expression of OSKM, mouse ESCs (OSKM-ESCs) and the resulting iPSCs (OSKM-iPSCs) reprogrammed from MEF cells propagated stably, expressed pluripotency marker genes, and formed three germ layers in teratomas. Transcriptional landscapes of OSKM-iPSCs resembled those of ESCs cultured in 2iL and were more similar to those of ESCs cultured in serum/LIF. Furthermore, OSKM-iPSCs contributed to germline transmission. CONCLUSIONS: Expression of OSKM could induce and maintain mouse pluripotency without specific culturing factors. Importantly, OSKM-iPSCs could produce gene-modified animals through germline transmission, with potential applications in other species.

Characterization of respiratory microbial dysbiosis in hospitalized COVID-19 patients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic of Coronavirus disease 2019 (COVID-19). However, the microbial composition of the respiratory tract and other infected tissues as well as their possible pathogenic contributions to varying degrees of disease severity in COVID-19 patients remain unclear. Between 27 January and 26 February 2020, serial clinical specimens (sputum, nasal and throat swab, anal swab and feces) were collected from a cohort of hospitalized COVID-19 patients, including 8 mildly and 15 severely ill patients in Guangdong province, China. Total RNA was extracted and ultra-deep metatranscriptomic sequencing was performed in combination with laboratory diagnostic assays. We identified distinct signatures of microbial dysbiosis among severely ill COVID-19 patients on broad spectrum antimicrobial therapy. Co-detection of other human respiratory viruses (including human alphaherpesvirus 1, rhinovirus B, and human orthopneumovirus) was demonstrated in 30.8% (4/13) of the severely ill patients, but not in any of the mildly affected patients. Notably, the predominant respiratory microbial taxa of severely ill patients were Burkholderia cepacia complex (BCC), Staphylococcus epidermidis, or Mycoplasma spp. (including M. hominis and M. orale). The presence of the former two bacterial taxa was also confirmed by clinical cultures of respiratory specimens (expectorated sputum or nasal secretions) in 23.1% (3/13) of the severe cases. Finally, a time-dependent, secondary infection of B. cenocepacia with expressions of multiple virulence genes was demonstrated in one severely ill patient, which might accelerate his disease deterioration and death occurring one month after ICU admission. Our findings point to SARS-CoV-2-related microbial dysbiosis and various antibiotic-resistant respiratory microbes/pathogens in hospitalized COVID-19 patients in relation to disease severity. Detection and tracking strategies are needed to prevent the spread of antimicrobial resistance, improve the treatment regimen and clinical outcomes of hospitalized, severely ill COVID-19 patients.

New therapeutic options for persistent low-level viremia in patients with chronic hepatitis B virus infection: Increase of entecavir dosage.

Chronic hepatitis B virus (HBV) infection (CHB) is a public health concern worldwide. Current therapies utilizing nucleos(t)ide analogs (NA) have not resulted in a complete cure for CHB. Furthermore, patients on long-term NA treatment often develop low-level viremia (LLV). Persistent LLV, in addition to causing the progression of liver disease or hepatocellular carcinoma, may shed light on the current plight of NA therapy. Here, we review the literature on LLV, NA treatment, and various doses of entecavir to find a strategy for improving the efficacy of this antiviral agent. For LLV patients, three therapeutic options are available, switching to another antiviral monotherapy, interferon-α switching therapy, and continuing monotherapy. In real-world clinical practice, entecavir overdose has been used in antiviral therapy for CHB patients with NA refractory and persistent LLV, which encouraged us to conduct further in-depth literature survey on dosage and duration related entecavir studies. The studies of pharmacodynamics and pharmacokinetics show that entecavir has the maximal selected index for safety, and has great potential in inhibiting HBV replication, in all of the NAs. In the particular section of the drug approval package published by the United States Food and Drug Administration, entecavir doses 2.5-20 mg/d do not increase adverse events, and entecavir doses higher than 1.0 mg/d might improve the antiviral efficacy. The literature survey led us to two suggestions: (1) Increasing entecavir dose to 1.0 mg/d for the treatment of NA naïve patients with HBV DNA >2 × 106 IU/mL is feasible and would provide better prognosis; and (2) Further research is needed to assess the long-term toxic effects of higher entecavir doses (2.5 and 5.0 mg/d), which may prove beneficial in treating patients with prior NA treatment, partial virological response, or LLV state.

Population Bottlenecks and Intra-host Evolution During Human-to-Human Transmission of SARS-CoV-2.

The emergence of the novel human coronavirus, SARS-CoV-2, causes a global COVID-19 (coronavirus disease 2019) pandemic. Here, we have characterized and compared viral populations of SARS-CoV-2 among COVID-19 patients within and across households. Our work showed an active viral replication activity in the human respiratory tract and the co-existence of genetically distinct viruses within the same host. The inter-host comparison among viral populations further revealed a narrow transmission bottleneck between patients from the same households, suggesting a dominated role of stochastic dynamics in both inter-host and intra-host evolutions.

Altered regulation of LncRNA analysis of human alcoholic hepatitis with Mallory-Denk Bodies (MDBs) is revealed by RNA sequencing.

Mallory-Denk Bodies (MDBs) are prevalent in a variety of liver diseases including alcoholic hepatitis (AH) and are formed in mice livers by feeding DDC. Long noncoding RNAs (lncRNAs) are considered as emerging new gene regulators, which participates in many functional activities through diverse mechanisms. We previously reported the mechanisms involved in the formation of liver MDBs in mouse model and in AH livers where MDBs had formed. To investigate the regulation of mRNAs expression and the probable role of lncRNAs in AH livers with MDBs, RNA-Seq analyses was further conducted to determine the mRNA and lncRNA expression profiles of the AH livers compared with the normal livers. It showed that different lncRNAs have different information contribution degrees by principal component analysis, and the integrated analysis of lncRNA-mRNA co-expression networks were linked to endocytosis, cell cycle, p53 signaling pathways in the human. Based on the co-expression networks, we identify 36 mRNAs that could be as potential biomarkers of alcoholic liver disease (ALD) and hepatocellular carcinoma (HCC). To our knowledge, this is the first report on the regulatory network of lncRNAs associated with liver MDB formation in human, and these results might offer new insights into the molecular mechanisms of liver MDB formation and the progression of AH to HCC.

Kinetics of viral load and antibody response in relation to COVID-19 severity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent for coronavirus 2019 (COVID-19) pneumonia. Little is known about the kinetics, tissue distribution, cross-reactivity, and neutralization antibody response in patients with COVID-19. Two groups of patients with RT-PCR-confirmed COVID-19 were enrolled in this study: 12 severely ill patients in intensive care units who needed mechanical ventilation and 11 mildly ill patients in isolation wards. Serial clinical samples were collected for laboratory detection. Results showed that most of the severely ill patients had viral shedding in a variety of tissues for 20-40 days after onset of disease (8/12, 66.7%), while the majority of mildly ill patients had viral shedding restricted to the respiratory tract and had no detectable virus RNA 10 days after onset (9/11, 81.8%). Mildly ill patients showed significantly lower IgM response compared with that of the severe group. IgG responses were detected in most patients in both the severe and mild groups at 9 days after onset, and remained at a high level throughout the study. Antibodies cross-reactive to SARS-CoV and SARS-CoV-2 were detected in patients with COVID-19 but not in patients with MERS. High levels of neutralizing antibodies were induced after about 10 days after onset in both severely and mildly ill patients which were higher in the severe group. SARS-CoV-2 pseudotype neutralization test and focus reduction neutralization test with authentic virus showed consistent results. Sera from patients with COVID-19 inhibited SARS-CoV-2 entry. Sera from convalescent patients with SARS or Middle East respiratory syndrome (MERS) did not. Anti-SARS-CoV-2 S and N IgG levels exhibited a moderate correlation with neutralization titers in patients' plasma. This study improves our understanding of immune response in humans after SARS-CoV-2 infection.

Postoperative intensity-modulated radiation therapy reduces local recurrence and improves overall survival in III-N2 non-small-cell lung cancer: A single-center, retrospective study.

PURPOSE: To determine the postoperative effects of radiotherapy (PORT) on the local recurrence-free survival (LRFS) and overall survival (OS) of stage III-N2 non-small-cell lung cancer (NSCLC). MATERIALS AND METHODS: 183 patients with resected stage III-pN2 NSCLC from Hunan Cancer Hospital between 2013 and 2016 were divided into two groups for postoperative chemotherapy (POCT) (n = 105) or combination chemotherapy and radiotherapy (POCRT) (n = 78). The LRFS and OS were compared and the factors affecting local recurrence were illustrated in these two groups. The sites of failure based on the lobe of the primary tumor in two groups were described. RESULTS: PORT leads to a strikingly lower risk for local recurrence and brought superior OS benefit. For different pN2 Subclassification, Patients with multiple-station pN2 ± pN1 disease had the worst LRFS (11 months) and single-station pN2 + multiple station pN1 disease had a relatively short LRFS (24 months) in group POCT. Short LRFS is correlated with multiple-station pN2, older age (Y > 55), patients with a high positive LN ratio > 1/3 and a poor tumor histological differentiation degree. In group POCT, the most frequent failure site occurs at the ipsilateral hilum (21.0%), the bronchial stump (20.0%), followed by LNs4R (19.0%), LNs4L (18.1%), LNs7 (15.2%), most of left-sided tumors more frequently involved the contralateral mediastinum, whereas the ipsilateral recurrences dominated for right-sided tumors, especially for LNs4R. In group POCRT, the highest failure site was the bronchial stump (11.5%), followed by LNs4L (8.97%), LNs1 (7.69%), the ipsilateral hilum (6.41%) and LNs4R (6.41%). CONCLUSION: PORT remarkably reduced local recurrence and improved OS in stage III-pN2 NSCLC, especially in the multiple-station pN2 group.

Preliminary evidence from a multicenter prospective observational study of the safety and efficacy of chloroquine for the treatment of COVID-19.

Effective therapies are urgently needed for the SARS-CoV-2 pandemic. Chloroquine has been proved to have antiviral effect against coronavirus in vitro. In this study, we aimed to assess the efficacy and safety of chloroquine with different doses in COVID-19. In this multicenter prospective observational study, we enrolled patients older than 18 years old with confirmed SARS-CoV-2 infection excluding critical cases from 12 hospitals in Guangdong and Hubei Provinces. Eligible patients received chloroquine phosphate 500 mg, orally, once (half dose) or twice (full dose) daily. Patients treated with non-chloroquine therapy were included as historical controls. The primary endpoint is the time to undetectable viral RNA. Secondary outcomes include the proportion of patients with undetectable viral RNA by day 10 and 14, hospitalization time, duration of fever, and adverse events. A total of 197 patients completed chloroquine treatment, and 176 patients were included as historical controls. The median time to achieve an undetectable viral RNA was shorter in chloroquine than in non-chloroquine (absolute difference in medians -6.0 days; 95% CI -6.0 to -4.0). The duration of fever is shorter in chloroquine (geometric mean ratio 0.6; 95% CI 0.5 to 0.8). No serious adverse events were observed in the chloroquine group. Patients treated with half dose experienced lower rate of adverse events than with full dose. Although randomized trials are needed for further evaluation, this study provides evidence for safety and efficacy of chloroquine in COVID-19 and suggests that chloroquine can be a cost-effective therapy for combating the COVID-19 pandemic.

Dihydroartemisinin increases apoptosis of colon cancer cells through targeting Janus kinase 2/signal transducer and activator of transcription 3 signaling.

As a derivative of artemisinin, dihydroartemisinin is effective in the treatment of malaria. Dihydroartemisinin has been identified to possess inhibitory effects in numerous types of animal model with tumors, indicating that it has an antineoplastic effect. The aim of the present study was to analyze the potential anticancer effects of dihydroartemisinin, particularly its effect on apoptosis of colon cancer cells. In the present study, it was identified that dihydroartemisinin inhibited cell viability, promoted cell apoptosis, increased B-cell lymphoma-2-associated X-protein expression, increased caspase-3/9 activities, decreased poly(ADP-ribose) polymerase levels, decreased phosphorylation of extracellular-signal-regulated kinase, and increased phosphorylation of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase in colon cancer cells. Conversely, the phosphorylation of Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) was suppressed by dihydroartemisinin in colon cancer cells. These results demonstrate that the potential anticancer effects of dihydroartemisinin may increase apoptosis of colon cancer cells through targeting JAK2/STAT3 signaling.

Association of mast cell infiltration with gastric cancer progression.

The present study aimed to determine the expression of mast cells, C-C motif chemokine ligand 2 (CCL-2) and C-C motif chemokine receptor 2 (CCR2) in gastric cancer tumor tissue; and the association of mast cells with the proliferation, migration, invasion and apoptosis of gastric cancer cells. In addition, whether the stem cell factor (SCF)/c-Kit pathway was associated with the secretion of CCL-2 by gastric cancer cells was explored. Flow cytometry analysis and immunohistochemistry were used to observe the relative number of mast cells, and reverse transcription-quantitative polymerase chain reaction and western blot analysis were utilized to determine the expression of CCL-2 and CCR2 mRNA and protein. Following the co-culture of the mast cell line HMC-1 and the gastric cancer cell line BGC-823, a Transwell assay was used to validate the effect of mast cells on the migration and invasion of gastric cancer cells. Furthermore, Cell Counting kit-8 and dual acridine orange/ethidium bromide fluorescent staining assays were performed to determine the proliferation and apoptosis of gastric cancer cells, following co-culture with mast cells. The expression of SCF and c-Kit were also determined with a western blot analysis. A specific phosphoinositide 3-kinase (PI3K) inhibitor, wortmannin, was used to test the effect of PI3K inhibition on the secretion of CCL-2 in gastric cancer. The results demonstrated that the proportion of infiltrating mast cells, and the mRNA/protein expression of CCL-2 and CCR2, were significantly increased in tumor tissue relative to adjacent tissues. In addition, the migration and invasion of gastric cancer cells were significantly increased when mast cells were used as an attractant. When co-cultured with mast cells, the viability of gastric cancer cells was significantly increased and H2O2-induced apoptosis was inhibited. In gastric cancer tissue samples, the expression of SCF, c-Kit and phosphorylated (p)-Akt protein were significantly increased compared with normal adjacent tissues. It was hypothesized that SCF/c-Kit signaling pathway was activated by PI3K-Akt, resulting in an increase in the expression of CCL-2 mRNA and protein. Furthermore, it was demonstrated that CCL-2 mRNA and protein expression was significantly inhibited by treatment with the PI3K inhibitor wortmannin. Additionally, wortmannin intervention significantly inhibited gastric cancer cell migration and invasion. Therefore, the results of the present study demonstrated that mast cells may promote gastric cancer cell proliferation, migration and invasion, and inhibit apoptosis. In addition, the activation of the SCF/c-Kit signaling pathway was identified to promote the expression of CCL-2, which is associated with the development and metastasis of gastric cancer.

Bioinformatics method identifies potential biomarkers of dilated cardiomyopathy in a human induced pluripotent stem cell-derived cardiomyocyte model.

Dilated cardiomyopathy (DCM) is the most common type of cardiomyopathy that account for the majority of heart failure cases. The present study aimed to reveal the underlying molecular mechanisms of DCM and provide potential biomarkers for detection of this condition. The public dataset of GSE35108 was downloaded, and 4 normal induced pluripotent stem cell (iPSC)-derived cardiomyocytes (N samples) and 4 DCM iPSC-derived cardiomyocytes (DCM samples) were utilized. Raw data were preprocessed, followed by identification of differentially expressed genes (DEGs) between N and DCM samples. Crucial functions and pathway enrichment analysis of DEGs were investigated, and protein-protein interaction (PPI) network analysis was conducted. Furthermore, a module network was extracted from the PPI network, followed by enrichment analysis. A set of 363 DEGs were identified, including 253 upregulated and 110 downregulated genes. Several biological processes (BPs), such as blood vessel development and vasculature development (FLT1 and MMP2), cell adhesion (CDH1, ITGB6, COL6A3, COL6A1 and LAMC2) and extracellular matrix (ECM)-receptor interaction pathway (CDH1, ITGB6, COL6A3, COL6A1 and LAMC2), were significantly enriched by these DEGs. Among them, MMP2, CDH1 and FLT1 were hub nodes in the PPI network, while COL6A3, COL6A1, LAMC2 and ITGB6 were highlighted in module 3 network. In addition, PENK and APLNR were two crucial nodes in module 2, which were linked to each other. In conclusion, several potential biomarkers for DCM were identified, such as MMP2, FLT1, CDH1, ITGB6, COL6A3, COL6A1, LAMC2, PENK and APLNR. These genes may serve significant roles in DCM via involvement of various BPs, such as blood vessel and vasculature development and cell adhesion, and the ECM-receptor interaction pathway.

MicroRNA-145 Inhibits Cell Migration and Invasion and Regulates Epithelial-Mesenchymal Transition (EMT) by Targeting Connective Tissue Growth Factor (CTGF) in Esophageal Squamous Cell Carcinoma.

BACKGROUND This study investigated the mechanism of miR-145 in targeting connective tissue growth factor (CTGF), which affects the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of ESCC cells. MATERIAL AND METHODS A total of 50 ESCC tissues and their corresponding normal adjacent esophageal tissue samples were collected. Then, miR-145 expression in both ESCC clinical specimens and cell lines was detected using quantitative real-time PCR. CTGF protein was detected using immunohistochemistry. Dual luciferase reporter gene assay was employed to assess the effect of miR-145 on the 3'UTR luciferase activity of CTGF. Eca109 cells were transfected with miR-145 mimics and CTGF siRNA, respectively, and changes in cellular proliferation, migration, and invasion were detected via MTT assay, wound-healing assay, and Transwell assay, respectively. Western blotting assay was used to detect the expression of marker genes related to EMT. RESULTS MiR-145 was significantly down-regulated in ESCC tissues and cell lines compared with normal tissues and cell lines (P<0.05). We found significantly more positively expressed CTGF protein in ESCC tissues was than in normal adjacent esophageal tissues (P<0.01). Dual luciferase reporter gene assay showed that miR-145 can specifically bind with the 3'UTR of CTGF and significantly inhibit the luciferase activity by 55% (P<0.01). Up-regulation of miR-145 or down-regulation of CTGF can suppress the proliferation, migration, invasion, and EMT process of ESCC cells. CONCLUSIONS MiR-145 was significantly down-regulated in ESCC tissues and cell lines, while the protein expression of CTGF exhibited the opposite trend. MiR-145 inhibited the proliferation, migration, invasiveness, and the EMT process of ESCC cells through targeted regulation of CTGF expression.