Carbonylation of Runx2 at K176 by 4-Hydroxynonenal Accelerates Vascular Calcification.

BACKGROUND: Vascular calcification, which is characterized by calcium deposition in arterial walls and the osteochondrogenic differentiation of vascular smooth muscle cells, is an actively regulated process that involves complex mechanisms. Vascular calcification is associated with increased cardiovascular adverse events. The role of 4-hydroxynonenal (4-HNE), which is the most abundant stable product of lipid peroxidation, in vascular calcification has been poorly investigated. METHODS: Serum was collected from patients with chronic kidney disease and controls, and the levels of 4-HNE and 8-iso-prostaglandin F2α were measured. Sections of coronary atherosclerotic plaques from donors were immunostained to analyze calcium deposition and 4-HNE. A total of 658 patients with coronary artery disease who received coronary computed tomography angiography were recruited to analyze the relationship between coronary calcification and the rs671 mutation in aldehyde dehydrogenase 2 (ALDH2). ALDH2 knockout (ALDH2-/-) mice, smooth muscle cell-specific ALDH2 knockout mice, ALDH2 transgenic mice, and their controls were used to establish vascular calcification models. Primary mouse aortic smooth muscle cells and human aortic smooth muscle cells were exposed to medium containing ß-glycerophosphate and CaCl2 to investigate cell calcification and the underlying molecular mechanisms. RESULTS: Elevated 4-HNE levels were observed in the serum of patients with chronic kidney disease and model mice and were detected in calcified artery sections by immunostaining. ALDH2 knockout or smooth muscle cell-specific ALDH2 knockout accelerated the development of vascular calcification in model mice, whereas overexpression or activation prevented mouse vascular calcification and the osteochondrogenic differentiation of vascular smooth muscle cells. In patients with coronary artery disease, patients with ALDH2 rs671 gene mutation developed more severe coronary calcification. 4-HNE promoted calcification of both mouse aortic smooth muscle cells and human aortic smooth muscle cells and their osteochondrogenic differentiation in vitro. 4-HNE increased the level of Runx2 (runt-related transcription factor-2), and the effect of 4-HNE on promoting vascular smooth muscle cell calcification was ablated when Runx2 was knocked down. Mutation of Runx2 at lysine 176 reduced its carbonylation and eliminated the 4-HNE-induced upregulation of Runx2. CONCLUSIONS: Our results suggest that 4-HNE increases Runx2 stabilization by directly carbonylating its K176 site and promotes vascular calcification. ALDH2 might be a potential target for the treatment of vascular calcification.

VEGF Mediates Tumor Growth and Metastasis by Affecting the Expression of E-Cadherin and N-Cadherin Promoting Epithelial to Mesenchymal Transition in Gastric Cancer.

Background: Gastric cancer (GC) is the fifth leading cancer in the world, and there is a high mortality rate in China. Exploring the relationship between the prognosis of GC and the expression of related genes is helpful to further understand the common characteristics of the occurrence and development of GC and provide a new method for the identification of early GC, so as to provide the best therapeutic targets. Methods: Vascular endothelial growth factor (VEGF) and markers of epithelial-mesenchymal transition (EMT) were investigated immunohistochemically using tumor samples obtained from 196 GC tissues and adjacent tumor tissues. The correlation of the expression level with histopathologic features and survival was investigated. Results: Here, we show that VEGF and EMT markers expression were significantly correlated with depth of tumor invasion and GC stage (P < .05), degree of differentiation and lymph node metastasis (P < .001). We found that the rate of VEGF positivity in GC tissues was 52.05%, which was significantly higher than that in adjacent cancer tissues (16.84%). In GC, the association between VEGF and E-cadherin was negative (r = -0.188, P < .05), whereas VEGF and N-cadherin were positively correlated (r = 0.214, P < .05). Furthermore, the Kaplan-Meier analysis and a Cox regression model were used to analyze the effect of VEGF and EMT marker expression on the survival of the patients. We found that the overall survival of GC patients was correlated with VEGF (P < .001), N-cadherin (P < .001), E-cadherin (P = .002) expression, and some histopathologic features. Conclusions: Vascular endothelial growth factor and EMT markers exist side by side and play a part together in the development of GC, which provides new ideas for evaluating the prognosis of GC and researching targeted drugs.

PLXDC1 Can Be a Biomarker for Poor Prognosis and Immune Evasion in Gastric Cancer.

Background: Research has revealed that Plexin domain containing 1 (PLXDC1) is correlated with the prognosis of a variety of tumors, but its role in the tumor microenvironment (TME) of gastric cancer has not been reported. Methods: In this study, we analyzed PLXDC1 expression in gastric cancer using the Oncomine and the Cancer Genome Atlas (TCGA) databases and immunohistochemical staining experiments, and performed prognostic assessment with data from the TCGA and Kaplan-Meier Plotter databases. The immunomodulatory role of PLXDC1 in the gastric cancer TME was analyzed by signaling pathway enrichment, immune cell correlation analysis, immunomodulator risk model construction and immunohistochemical staining experiments of immune cells. Results: The results indicated that PLXDC1 was overexpressed in gastric cancer and that its overexpression was associated with poor prognosis. Multivariate Cox analysis revealed that PLXDC1 could be an independent biomarker of the risk of gastric cancer. Signaling pathway enrichment revealed that high PLXDC1 expression was involved in signaling pathways related to immune activation and stromal activation, and Tumor Immune Dysfunction and Exclusion (TIDE) assessment indicated that high PLXDC1 expression was associated with a significantly higher risk of immune evasion than low PLXDC1 expression. A Cox risk model based on PLXDC1-associated immunomodulators also presented poor prognosis, and immune evasion was significantly higher in the high-risk group than in the low-risk group. In addition, immunohistochemical staining of CD8/CD3/CD4+ T cells in the high and low PLXDC1 expression groups also observed immune cell distribution characteristics of immune evasion. Conclusion: This study analyzed PLXDC1 from multiple biological perspectives and revealed that PLXDC1 can be a biomarker for poor prognosis and immune evasion in gastric cancer.

Severe immunochemotherapy-induced toxicities in a patient with dyskeratosis congenita and literature review.

OBJECTIVES: Dyskeratosis congenita (DC) is a rare inherited disease characterized by the triad of reticulate hyperpigmentation, nail dystrophy and oral leukoplakia. DC patients are considered vulnerable to external pressure, such as immunochemotherapy. There are very few cases reporting severe therapy-induced toxicities in patients with DC. METHODS: A 27-year-old woman was admitted to our hospital with a 4-month history of pancytopenia and a 7-day history of dyspnea with coughing. She was diagnosed with non-Hodgkin's lymphoma 5 months ago. She received immunochemotherapy due to non-Hodgkin's lymphoma but experienced recurrent fever, oral ulcer, pancytopenia, dyspnea and other symptoms during immunochemotherapy. On admission, she experienced an aggravation of respiratory symptoms, recurrent infections and acute heart failure. RESULTS: Laboratory examination confirmed pancytopenia, and chest computed tomography showed interstitial lung disease (ILD). Genetic analysis results confirmed the presence of DC and a TINF2 gene mutation. With continuous supportive and anti-infection treatment, her condition finally stabilized. She was discharged from the hospital after nearly 2 months. DISCUSSION: We reviewed similar cases and found common features that could be useful. However, the reported cases are very limited. More cases and studies are needed. CONCLUSION: These cases indicate that DC patients seem more vulnerable to therapy toxicities; thus, physicians should be careful when treating these patients with chemotherapy drugs or radiation therapy. Reduced-intensity therapy may be considered.

The upregulation of stromal antigen 3 expression suppresses the phenotypic hallmarks of hepatocellular carcinoma through the Smad3-CDK4/CDK6-cyclin D1 and CXCR4/RhoA pathways.

BACKGROUND: The stromal antigen 3 (STAG3) gene encodes an adhesion complex subunit that can regulate sister chromatid cohesion during cell division. Chromosome instability caused by STAG3 gene mutation may potentially promote tumor progression, but the effect of STAG3 on hepatocellular carcinoma (HCC) and the related molecular mechanism are not reported in the literature. The mechanism of the occurrence and development of HCC is not adequately understood. Therefore, the biological role of STAG3 in HCC remains to be studied, and whether STAG3 might be a sensitive therapeutic target in HCC remains to be determined. METHODS: The expression and clinical significance of STAG3 in HCC tissues and cell lines were determined by RT-qPCR and immunohistochemistry analyses. The biological functions of STAG3 in HCC were determined through in vitro and in vivo cell function tests. The molecular mechanism of STAG3 in HCC cells was then investigated by western blot assay. RESULTS: The mRNA expression of STAG3 was lower in most HCC cells than in normal cells. Subsequently, an immunohistochemical analysis of STAG3 was performed with 126 samples, and lower STAG3 expression was associated with worse overall survival in HCC patients. Moreover, cytofunctional tests revealed that the lentivirus-mediated overexpression of STAG3 in HCC cells inhibited cell proliferation, migration, and invasion; promoted apoptosis; induced G1/S phase arrest in vitro; and inhibited tumor growth in vivo. Furthermore, studies of the molecular mechanism suggested that the overexpression of STAG3 increased Smad3 expression and decreased CDK4, CDK6, cyclin D1, CXCR4 and RhoA expression. CONCLUSION: STAG3 exhibits anticancer effects against HCC, and these effects involve the Smad3-CDK4/CDK6-cyclin D1 and CXCR4/RhoA pathways. STAG3 is a tumor-suppressor gene that may serve as a potential target for molecular therapy, which provides a new idea for the treatment of HCC.

ANKFN1 plays both protumorigenic and metastatic roles in hepatocellular carcinoma.

Ankyrin repeat and fibronectin type III domain containing 1 (ANKFN1) is reported to be involved in human height and developmental abnormalities, but the expression profile and molecular function of ANKFN1 in hepatocellular carcinoma (HCC) remain unknown. This study aimed to evaluate the clinical significance and biological function of ANKFN1 in HCC and investigate whether ANKFN1 can be used for differential diagnosis in HCC. Here, we showed that ANKFN1 was upregulated in 126 tumor tissues compared with adjacent nontumorous tissues in HCC patients. The upregulation of ANKFN1 in HCC was associated with cirrhosis, alpha-fetoprotein (AFP) levels and poor prognosis. Moreover, silencing ANKFN1 expression suppressed HCC cell proliferation, migration, invasion, and metastasis in vitro and subcutaneous tumorigenesis in vivo. However, ANKFN1 overexpression promoted HCC proliferation and metastasis in an orthotopic liver transplantation model and attenuated the above biological effects in HCC cells. ANKFN1 significantly affected HCC cell proliferation by inducing G1/S transition and cell apoptosis. Mechanistically, we demonstrated that ANKFN1 promoted cell proliferation, migration, and invasion via activation of the cyclin D1/Cdk4/Cdk6 pathway by stimulating the MEK1/2-ERK1/2 pathway. Moreover, ANKFN1-induced cell proliferation, migration, and invasion were partially reversed by ERK1/2 inhibitors. Taken together, our results indicate that ANKFN1 promotes HCC cell proliferation and metastasis by activating the MEK1/2-ERK1/2 signaling pathway. Our work also suggests that ANKFN1 is a potential therapeutic target for HCC.

Peri-operative Takotsubo syndrome after non-cardiac surgery: a retrospective nested case-control study.

AIMS: Takotsubo syndrome (TTS) is an acute reversible cardiac dysfunction that may occur during the peri-operative period and among patients with serious illness. We aimed to evaluate the clinical characteristics, peri-operative management, and prognosis of peri-operative TTS (pTTS) and explore the factors associated with pTTS. METHODS: We conducted a retrospective nested case-control study using the database of patients who underwent in-hospital non-cardiac surgeries between January 2017 and December 2020 in Peking University Third hospital. Cases were adult patients diagnosed TTS at discharge who were matched with four controls based on operative types. Multivariable conditional logistic regression was used to identified the factors associated with pTTS. The area under the curve (AUC) was used to evaluate the diagnostic efficacy. RESULTS: Among the 128 536 patients underwent non-cardiac surgery, 20 patients with pTTS and 80 patients without were enrolled in this study. The incidence of pTTS was about 0.016% in our centre. The median age of patients with pTTS was 52.5 (38.25, 76.25) years, although 90% of them were female. Fifty per cent (9 cases) of female patients were pre-menopausal. Caesarean section has the highest proportion of pTTS (30% of the pTTS cases) with the incidence of caesarean section-related pTTS of 0.06% in our centre. A high prevalence of non-apical ballooning pattern of regional wall motion abnormality (seven cases, 35%) and a high mortality (two cases, 10%) were observed. Left ventricular ejection fraction (LVEF) of patients with pTTS was significantly decreased (41.7 ± 8.8%). In the acute phase, supportive treatments aiming to reduce life-threatening complications were main treatment strategies. After receiving systematic treatment, significant improvements were observed in LVEF (63.1 ± 13.5%), with median recovery time of LVEF of 7.48 days. Leucocyte count [odds ratio (OR): 4.59; 95% confidence interval (CI): 1.10-19.15], haemoglobin (HGB) (OR: 10.52; 95% CI: 1.04-106.36), and the revised cardiac risk index (RCRI) score (OR: 6.30; 95% CI: 1.05-37.88) were the factors significantly associated with pTTS. The RCRI score performed poorly in the prediction of pTTS (AUC: 0.630; 95% CI: 0.525-0.735). After adding leucocyte count and HGB into the RCRI score, the AUC was significantly improved (AUC: 0.768; 95% CI: 0.671-0.865; P = 0.001). CONCLUSIONS: Patients with pTTS have some differences compared with common TTS, including higher proportion of pre-menopausal female, higher prevalence during caesarean section, higher prevalence of non-apical ballooning pattern of regional wall motion abnormality, and higher mortality. The RCRI score performed poorly in the evaluation of pTTS. Adding HGB and leucocyte count into the RCRI score could significantly improve its predictive performance.

Malignant Peripheral Nerve Sheath Tumor of the Cervix.

   The occurrence of primary malignant peripheral neurilemmoma in the female cervix without neurofibromatosis type 1 is extremely rare. We, herein, report a case of a 46-year woman with malignant peripheral nerve sheath tumor (MPNST) involving the cervix. The patient was admitted to the hospital because of irregular vaginal bleeding. Pelvic CT suggested the presence of cervical mass, and the patient accepted radical hysterectomy. Following the surgery, MPNST was diagnosed by a combination of histology and immunohistochemistry. One year later, the patient developed recurrence and pulmonary metastasis and received sequentially epiubicin/ifosfamide, dacarbazine, and etoposide/cisplatin multi-line chemotherapy. The prognosis of MPNST is poor, which is related to its invasiveness, high recurrence and metastasis rates. The patient was treated with surgery, multi-line chemotherapy and targeted therapy, but the overall survival (OS) was 44 months. It is the first reported case of cervical MPNST receiving combined treatment. Key Words: Malignant peripheral nerve sheath tumor, Cervix, Pulmonary metastasis, Hysterectomy, Chemotherapy.

Development and Validation of Predictive Model-HASBLAD Score-For Major Adverse Cardiovascular Events During Perioperative Period of Non-cardiac Surgery: A Single Center Experience in China.

Background: Major adverse cardiovascular events (MACEs) represent a significant reason of morbidity and mortality in non-cardiac surgery during perioperative period. The prevention of perioperative MACEs has always been one of the hotspots in the research field. Current existing models have not been validated in Chinese population, and have become increasingly unable to adapt to current clinical needs. Objectives: To establish and validate several simple bedside tools for predicting MACEs during perioperative period of non-cardiac surgery in Chinese hospitalized patients. Design: We used a nested case-control study to establish our prediction models. A nomogram along with a risk score were developed using logistic regression analysis. An internal cohort was used to evaluate the performance of discrimination and calibration of these predictive models including the revised cardiac risk index (RCRI) score recommended by current guidelines. Setting: Peking University Third Hospital between January 2010 and December 2020. Patients: Two hundred and fifty three patients with MACEs and 1,012 patients without were included in the training set from January 2010 to December 2019 while 38,897 patients were included in the validation set from January 2020 and December 2020, of whom 112 patients had MACEs. Main Outcome Measures: The MACEs included the composite outcomes of cardiac death, non-fatal myocardial infarction, non-fatal congestive cardiac failure or hemodynamically significant ventricular arrhythmia, and Takotsubo cardiomyopathy. Results: Seven predictors, including Hemoglobin, CARDIAC diseases, Aspartate aminotransferase (AST), high Blood pressure, Leukocyte count, general Anesthesia, and Diabetes mellitus (HASBLAD), were selected in the final model. The nomogram and HASBLAD score all achieved satisfactory prediction performance in the training set (C statistic, 0.781 vs. 0.768) and the validation set (C statistic, 0.865 vs. 0.843). Good calibration was observed for the probability of MACEs in the training set and the validation set. The two predictive models both had excellent discrimination that performed better than RCRI in the validation set (C statistic, 0.660, P < 0.05 vs. nomogram and HASBLAD score). Conclusion: The nomogram and HASBLAD score could be useful bedside tools for predicting perioperative MACEs of non-cardiac surgery in Chinese hospitalized patients.

CHSY3 can be a Poor Prognostic Biomarker and Mediates Immune Evasion in Stomach Adenocarcinoma.

Background: Chondroitin sulphate synthase 3 (CHSY3) is an important enzyme that regulates glycosylation, but it has not been reported in tumours. This study explored for the first time the oncological features of CHSY3 in stomach adenocarcinoma (STAD). Methods: We analysed CHSY3 expression in STAD through the Cancer Genome Atlas (TCGA) database and verified our findings by immunohistochemical staining and Western blot experiments. The prognostic value of CHSY3 in STAD was analysed through the biological aspects of CHSY3 in STAD, such as communal clinical follow-up survival data, methylation sites, tumour immune microenvironment (TIME) and immune cell surface checkpoints. Finally, the immune-evasion potential of CHSY3 in STAD was assessed on the Tumor Immune Dysfunction and Exclusion (TIDE) website and immunohistochemical staining experiment. Results: CHSY3 overexpression in STAD was associated with a poor prognosis based on immunohistochemical staining and Western blot experiments. Multivariate Cox analysis suggested that CHSY3 could be an independent prognostic risk factor. Pathway enrichment and TIME analysis demonstrated that CHSY3 up-regulated mesenchymal activation and immune activation signals in STAD, while TIDE assessment revealed that the risk of immune evasion was significantly higher in the high CHSY3 expression group than in the low CHSY3 expression group. Risk model scores based on CHSY3-associated immune cell surface checkpoints also presented poor prognosis, and immune evasion was significantly higher in the high-risk group than in the low-risk group. Conclusions: This study analysed CHSY3 from multiple biological perspectives and revealed that CHSY3 can be a biomarker of poor prognosis and mediates the TIME immune-evasion status in STAD.

miR-107 regulates the effect of MCM7 on the proliferation and apoptosis of colorectal cancer via the PAK2 pathway.

Microchromosome maintenance protein 7 (MCM7), a DNA replication permitting factor, plays an essential role in initiating DNA replication. MCM7 is reported to be involved in tumor formation and progression, whereas the expression profile and molecular function of MCM7 in colorectal cancer (CRC) remain unknown. In this study, we aimed to evaluate the clinical significance and biological function of MCM7 in CRC and investigated whether MCM7 can be used for a differential diagnosis in CRC and whether it may serve as a more sensitive proliferation marker for CRC evaluation. Moreover, immunohistochemical analysis of MCM7 was performed in a total of 89 specimens, and high MCM7 expression levels were associated with worse overall survival (OS) in CRC patients. Furthermore, the cell functional test suggested that lentivirus-mediated silencing of MCM7 with shRNA in CRC cells significantly inhibited cellular proliferation and promoted apoptosis in vitro and inhibited tumor growth in vivo. Additionally, mechanistic studies further demonstrated that P21-activated protein kinase 2 (PAK2) was regulated by MCM7 via microarray analysis and cell functional recovery tests, and miR-107 played a role in regulating expression MCM7 via miRNA microarray analysis and 3'UTR reporter assays. Taken together, our results suggest that the miR-107/MCM7/PAK2 pathway may participate in cancer progression and that MCM7 may serve as a prognostic biomarker in CRC.

Identification of an antitumor effect of demethylzeylasteral on human gastric cancer cells.

Gastric cancer is a common malignancy in China, with the second highest mortality rate worldwide. Advanced gastric cancer usually exhibits a poor prognosis with a low 5-year survival rate. Therefore, developing novel drugs for the treatment of this cancer will be beneficial for patients. Demethylzeylasteral, an extract of tripterygium wilfordii, has shown positive anticancer activities. However, the possible antitumor effect of demethylzeylasteral on gastric cancer cells and its underlying molecular mechanism remain to be determined. In the present study, the Cell Counting Kit-8 and colony formation assays revealed that demethylzeylasteral impeded the proliferation of human gastric cancer cells in a dose-dependent manner. Furthermore, the Transwell assay identified an inhibitory effect of demethylzeylasteral on the migration of MKN-45 cells, while flow cytometry found that treatment with demethylzeylasteral induced apoptosis and decreased the mitochondrial membrane potential in the cancer cells. Further investigation revealed that demethylzeylasteral downregulated the phosphorylation of ERK1/2, AKT, and GSK-3ß in MKN-45 cells. Notably, decreased expression of Bcl-2 and increased expression of Bax, cleaved caspase-3, cleaved caspase-9 and cleaved PARP were detected in the cancer cells treated with demethylzeylasteral. The present study demonstrated that demethylzeylasteral exhibits therapeutic potential for gastric cancer.

Effect of LY294002 on adriamycin-induced epithelial-mesenchymal transition in human breast carcinoma cells.

OBJECTIVE: To study the effect of LY294002 on the adriamycin- induced epithelial-mesenchymal transition in human breast carcinoma cells. METHODS: Human breast carcinoma cells MCF-7 was cultured in vitro and then exposed to adriamycin with or without LY294002. The protein expression levels of Akt, phosphorylated-Akt (p-Akt), Snail, and E-cadherin was detected by Western blot analysis. The mRNA expressions of Snail and E-cadherin were determined by RT-PCR. RESULTS: Adriamycin significantly increased the protein expression of Snail and depressed the protein expression of E-cadherin (P<0.05). The pre-treatment with LY294002 significantly reversed the changes of activities and levels of the above proteins (P<0.05). CONCLUSION: LY294002 could reverse the adriamycin-induced epithelial-mesenchymal transition in human breast carcinoma cells by regulating the expressions of Snail and E-cadherin through suppressing PI3K/Akt signaling pathway.