CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas.

Chaperonins, which contain t-complex polypeptide 1 (CCT), are critical for correct protein folding to generate stable and functional protein conformations, which are important for cell growth and survival. However, little is known about the expression and prognostic significance of CCT8 (subunit 8 of the CCT complex chaperonin) in lung cancer. In this study, we demonstrated that CCT8 expression is frequently increased in human lung cancer. Survival analysis indicated that CCT8 expression is closely correlated with inferior overall survival in lung adenocarcinoma (LUAD), but not in lung squamous carcinoma (LUSC). Subsequently, ectopic expression of CCT8 facilitated cell migration and tumor metastasis, and vice versa. Mechanistically, CCT8 interacted and activated ATK. Inhibition of AKT suppressed CCT8-induced cell migration and tumor metastasis. Our findings support CCT8 as a biomarker for LUAD prognosis and as a target for LUAD therapy.

The Effect of miR-505-5p on Inhibition of Serum Uromodulin Ameliorates Myocardial Inflammation and Apoptosis Induced by Ischemia-Reperfusion.

Background: It has been found that miR-505-5p is closely related to cardiovascular metabolic risk factors. Nonetheless, there is little research analyzing miR-505-5p for its role as well as molecular mechanism in myocardial injury caused by ischemia-reperfusion (I/R). Methods: This work utilized quantitative reverse transcriptase PCR (qRT-PCR) for detecting miR-505-5p and serum uromodulin (sUmod) levels. sUmod, interleukin-1beta (IL-1ß), IL-6, IL-10, caspase7, caspase9, tumor necrosis factor-alpha (TNF-α), Bax, and Bcl-xL expression was detected by western blot. Bioinformatics database was used for target prediction and miR-505-5's target was determined by luciferase reporter gene assay. Results: Relative to sham group, sUmod was highly expressed within myocardial I/R injury (MIRI), whereas sUmod silencing significantly decreased the heart weight/body weight ratio, reduced serum myocardial enzymes expression, ameliorated I/R-mediated myocardial apoptosis, and inflammation. TargetScan bioinformatics database and luciferase reporter genes confirmed that sUmod was miR-505-5p's direct target gene, besides, miR-505-5p overexpression significantly improved the myocardial injury score, increased IL-10, decreased TNF-α, IL-1ß, IL-6 expression, decreased caspase7, caspase9, Bax expression, and increased Bcl-xL expression. More importantly, overexpression of sUmod abolished miR-505-5p overexpression's role in I/R-mediated myocardial apoptosis and inflammation. Conclusion: miR-505-5p can improve I/R-mediated myocardial apoptosis and inflammation by targeting sUmod. In this study, miR-505-5p is related to MIRI pathogenesis, which provides the new possible targeted therapy in patients with MIRI.

FAT10 Combined with Miltefosine Inhibits Mitochondrial Apoptosis and Energy Metabolism in Hypoxia-Induced H9C2 Cells by Regulating the PI3K/AKT Signaling Pathway.

Hypoxia-induced cardiomyocyte apoptosis is the main contributor to heart diseases. Human leukocyte antigen F-associated transcript 10 (FAT10), the small ubiquitin-like protein family subtype involved in apoptosis, is expressed in the heart and exhibits cardioprotective functions. This study explored the impact of FAT10 on hypoxia-induced cardiomyocyte apoptosis and the involved mechanisms. The cardiomyocyte cell line H9C2 was cultivated in hypoxia-inducing conditions (94% N2, 5% CO2, and 1% O2) and the expression of FAT10 in hypoxia-stimulated H9C2 cells was identified. For this, FAT10 overexpression/interference vectors were exposed to transfection into H9C2 cells with/without the PI3K/AKT inhibitor, miltefosine. The results indicated that hypoxia exposure decreased the FAT10 expression, suppressed H9C2 cell growth, disrupted mitochondrial metabolism, and promoted H9C2 cell apoptosis and oxidative stress. However, these impacts were reversed by the FAT10 overexpression. In addition, the inhibition of PI3K/AKT in FAT10-overexpressing cells suppressed cell proliferation, impaired mitochondrial metabolism, and promoted apoptosis and oxidative stress response. The findings demonstrated that FAT10 inhibited mitochondrial apoptosis and energy metabolism in hypoxia-stimulated H9C2 cells through the PI3K/AKT pathway. This finding can be utilized for developing therapeutic targets for treating heart disorders associated with hypoxia-induced apoptosis.

A novel 125I seeds fixed on gastric tube for treatment of inoperable esophageal carcinoma: A case report.

ABSTRACT: To explore the treatment benefit of 125I seeds fixed on a gastric tube in the early inoperable esophageal carcinoma (EC).Three senile patients with early inoperable EC who were treated with brachytherapy between October 2017 and February 2019 were included in this study. 125I seeds were fixed on the gastric tube, which was then inserted on the surface of the EC. One patient suffered from severe pulmonary insufficiency; 1 patient underwent colon cancer surgery one week before treatment and suffered from liver dysfunction and esophageal varices; 1 patient suffered from venous embolism of lower extremities and pulmonary artery embolism.All three patients were successfully operated and completed treatment. During the operation, no displacement and shedding of 125I seed gastric tube occurred. After surgery, the discomfort while swallowing and pain after eating were significantly improved. Moreover, dysphagia was relieved in patient 3. In addition, no complications, such as perforation or bleeding, occurred. Local lesions were effectively controlled.Gastric tube with 125I seeds provides a new treatment protocol for inoperable EC and malignant obstruction of esophageal carcinoma.

Combined Administration of Metformin and Atorvastatin Attenuates Diabetic Cardiomyopathy by Inhibiting Inflammation, Apoptosis, and Oxidative Stress in Type 2 Diabetic Mice.

Diabetic cardiomyopathy (DCM), a common complication of diabetes mellitus, may eventually leads to irreversible heart failure. Metformin is the cornerstone of diabetes therapy, especially for type 2 diabetes. Statins are widely used to reduce the risk of cardiovascular diseases. In this study, we aimed to investigate whether the combined administration of metformin and atorvastatin could achieve superior protective effects on DCM and to elucidate its molecular mechanism. Here, db/db mice (9-10 weeks old) were randomly divided into four groups, including sterile water group (DM), metformin group (MET, 200 mg/kg/day), atorvastatin group (AVS, 10 mg/kg/day), and combination therapy group (MET + AVS). Mice were treated with different drugs via gavage once per day for 3 months. After 3 months of treatment, the pathological changes (inflammation, fibrosis, hypertrophy, and oxidative stress makers) were detected by histopathological techniques, as well as Western blotting. The H9C2 cardiomyocytes were treated with palmitate (PAL) to mimic diabetic condition. The cells were divided into control group, PAL treatment group, MET + PAL treatment group, AVS + PAL treatment group, and MET + AVS + PAL treatment group. The effects of MET and AVS on the cell viability and inflammation of H9C2 cells subjected to PAL condition were evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay, immunofluorescence staining, and Western blotting. Both MET and AVS prevented diabetes-induced fibrosis, hypertrophy, and inflammation. The combination therapy showed superior effects in protecting myocardial tissue against diabetes-induced injury. Mechanistically, the combination therapy significantly inhibited oxidative stress and the expression levels of inflammation-related proteins, e.g., NLRP3, caspase-1, interleukin-1ß (IL-1ß), Toll-like receptor 4 (TLR4), and P-p65/p65, in both cardiac tissues and H9C2 cells. TUNEL assay showed that the combination therapy significantly attenuated the apoptosis of cardiomyocytes; decreased the expression level of pro-apoptotic-related proteins, such as cleaved caspase-3 and BAX; and enhanced the expression level of anti-apoptotic protein (Bcl-2). Furthermore, the combination therapy remarkably upregulated the expression levels of 5'-AMP-activated protein kinase (AMPK) and SIRT1. Our findings indicated that the anti-inflammation and anti-apoptosis effects of the combination therapy may be related to activation of AMPK/SIRT1 signaling pathway.

LIMD1 Increases the Sensitivity of Lung Adenocarcinoma Cells to Cisplatin via the GADD45α/p38 MAPK Signaling Pathway.

Objective: To investigate the effect of LIM domain-containing protein 1 (LIMD1) on the sensitivity of lung adenocarcinoma cells to cisplatin and explore the mechanism. Methods: A549 and H1299 cells were transfected with lentivirus to establish LIMD1-overexpressing cell lines and their respective controls. The protein expression of DNA damage-inducible 45 alpha (GADD45α) and p38 mitogen-activated protein kinase (MAPK) was detected by Western blot. The survival of A549-vec, A549-LIMD1, H1299-vec, and H1299-LIMD1 cells after cisplatin treatment was observed by CCK-8, and the viability was calculated accordingly. Then, SB203580 was used to inhibit the activity of the p38 MAPK signaling pathway, after which the survival of A549-vec, A549-LIMD1, H1299-vec, and H1299-LIMD1 cells in response to cisplatin was observed again by CCK-8, and the viability was calculated accordingly. Results: When LIMD1 was overexpressed in A549 and H1299 cells, the levels of GADD45α and p-p38 MAPK were increased, but total p38 MAPK expression showed no significant change. After adding 30 µM cisplatin, the optical density (OD) values of A549-LIMD1 and H1299-LIMD1 cells were significantly lower than those of their respective controls at 24, 48, and 72 h. The viability of A549-LIMD1 and H1299-LIMD1 cells was significantly lower than that of their respective controls at all the times tested (p < 0.05). The Western blot results showed that the expression of apoptotic proteins cleaved caspase 3 and cleaved PARP in cisplatin-treated A549-LIDM1 and H1299-LIMD1 cells was significantly higher than that in their respective control cells. Flow cytometry showed that the apoptosis rates of A549-LIMD1 and H1299-LIMD1 cells were significantly higher than those of their respective controls (p < 0.05). SB203580 significantly inhibited the activation of the p38 MAPK signaling pathway in lung adenocarcinoma cells; however, neither the OD values nor the viability of A549-LIMD1 cells and H1299-LIMD1 cells showed no significant difference from those of their controls at 24, 48, and 72 h after cisplatin and SB203580 treatment (p > 0.05 for both). Western blot analysis showed that after SB203580 was added, the expression of cleaved caspase 3 and cleaved PARP in A549-LIMD1 and H1299-LIMD1 cells presented no significant difference compared with that in their respective controls. Conclusion: LIMD1 increases the sensitivity of lung adenocarcinoma cells to cisplatin by activating the GADD45α/p38 MAPK signaling pathway.

Placenta-specific protein 8 promotes the proliferation of lung adenocarcinoma PC-9 cells and their tolerance to an epidermal growth factor receptor tyrosine kinase inhibitor by activating the ERK signaling pathway.

Placenta-specific protein 8 (PLAC8) is a conserved protein with a molecular weight of 12.5 kDa. The specific function of this protein has not been fully elucidated, however, PLAC8 has been found to play an important tumor regulatory role in certain types of cancer, including colon, pancreatic and liver cancer. PLAC8 also participates in the regulation of the cell cycle, autophagy, epithelial-mesenchymal transition and other cellular functions, indicating its potential as a molecular target worth further investigation. The present study investigated the effect of PLAC8 on the proliferation of lung adenocarcinoma PC-9 cells and their sensitivity to gefitinib, an epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI). It was found that the inhibition of PLAC8 expression in PC-9 cells resulted in significantly decreased proliferation, whereas overexpression of PLAC8 significantly increased the proliferation (P<0.05) of PC-9 cells. Furthermore, inhibition of PLAC8 expression resulted in decreased activity of the ERK signaling pathway, while PLAC8 overexpression increased activity of this pathway. Inhibition of the ERK signaling pathway with U0126 reversed the effects induced by inhibiting or overexpressing PLAC8 on cell proliferation. In addition, overexpression of PLAC8 significantly decreased the sensitivity of PC-9 cells to gefitinib, and this effect was reversed by U0126. Overall, these results suggest that PLAC8 is involved in the regulation of proliferation of lung adenocarcinoma PC-9 cells and impacts their sensitivity to an EGFR-TKI. Thus, PLAC8 is a potential novel target in lung adenocarcinoma for future studies.

The long non-coding RNA LINC01606 contributes to the metastasis and invasion of human gastric cancer and is associated with Wnt/ß-catenin signaling.

The dysregulation of long non-coding RNA (lncRNA) has increasingly been linked to human gastric cancer (GC). However, the LINC01606 expression level and clinical values, and its role in the molecular mechanism underlying GC remain largely unknown. In our research, we found that LINC010606 was elevated aberrantly and correlated with metastasis and invasion in GC patients. Moreover, we found that LINC01606 expression level was associated with Wnt/ß-catenin signaling. In addition, subsequent functional experiments showed that JW74, a specific Wnt/ß-catenin signaling inhibitor, inhibited the transcription of LINC01606 and suppressed migration and invasion in GC cell lines. We also revealed that LINC01606 might be associated with miR-423-5p to regulate the level to which the Wnt/ß-catenin signaling pathway is activated. In summary, the findings of this study, based on competing endogenous RNA (ceRNA) theory, combine new data on the interaction between miR-423-5p and Wnt3a and introduce LINC01606 as a new focus for research, thus providing new insight into possible molecular-level approaches to preventing the migration and invasion of GC.

Downregulation of microRNA­199a­5p protects cardiomyocytes in cyanotic congenital heart disease by attenuating endoplasmic reticulum stress.

Chronic hypoxia is a key pathological change in patients with cyanotic congenital heart defect (CCHD). It has been demonstrated that enhanced myocardial unfolded protein response (UPR) increases the capacity to buffer endoplasmic reticulum (ER) stress and to avoid subsequent apoptosis caused by the hypoxia that underlies CCHD. The present study was performed to determine the regulatory role of microRNAs (miRNAs) in this cytoprotective UPR process. The results revealed that miR­199a­5p was markedly downregulated in the cardiac tissue of patients with CCHD and in human myocardial cells cultured in hypoxic conditions. The two major UPR modulators, 78 kDa glucose­regulated protein (GRP78) and activating transcription factor 6 (ATF6), were potential target genes of miR­199a­5p in CCHD myocardial specimens. In addition, the miR­199a­5p mimic and inhibitor were evidently able to change GRP78 and ATF6 gene expression and ER stress­associated apoptosis in hypoxia­treated cardiomyocytes. The interaction between miR­199a­5p and the ATF6 and GRP78 3'­UTR binding sites in myocardial cells was also confirmed by luciferase assay. Thus, it is concluded that myocardial downregulation of miR­199a­5p favors the UPR against hypoxia­induced ER stress in CCHD, which contributes to myocardial protection.

A novel approach for characterizing variations in serum peptides in rheumatic heart disease.

BACKGROUND & OBJECTIVES: Acute rheumatic fever and rheumatic heart disease (RHD) are important public health problems in developing countries. In this study, peptidomic analyses on RHD patients and healthy individuals were performed to characterize variations in serum peptide levels using label-free quantitation approaches. METHODS: Blood samples were obtained from 160 healthy controls and 160 RHD patients. Of the 448 identified peptides, 272 were analyzed by two label-free mass spectrometry methods, the spectral count and spectral index. RESULTS: There were 38 proteins and 95 peptides with significant (adjusted P<0.001) differences in the abundance of peptides between healthy controls and RHD patients, including multiple peptides derived from histone H2B, villin-like protein, complement C4-B and motile sperm domain containing protein-2. The levels of 10 peptides were upregulated, and 85 peptides were downregulated in patients compared to controls. In addition, in patients, the levels of four proteins were upregulated and 34 were downregulated compared to controls. INTERPRETATION & CONCLUSIONS: This study shows that detection of significant changes in serum peptides reflects the difference between RHD patients and healthy controls. This label-free method may be helpful for clinicians to treat RHD patients during the perioperative period.

Upregulated ATF6 contributes to chronic intermittent hypoxia-afforded protection against myocardial ischemia/reperfusion injury.

In the present study, we investigated the role of activating transcription factor 6 (ATF6) in the mechanism by which chronic intermittent hypoxia (CIH) increases tolerance to myocardial ischemia/reperfusion (I/R). Experiments were conducted using a rat model of I/R injury in vivo and isolated Langendorff-perfused rat hearts ex vivo. The role of Akt in this process was also investigated in vitro using rat myoblast H9c2 cells. Cell viability was measured using a cell counting kit-8 assay. Lactate dehydrogenase (LDH) and creatine kinase cardiac isoenzyme activity were also measured as markers of cellular damage. ATF6, Akt and phosphorylated (p)-Akt expression was analyzed by western blot analysis. RNA interference (RNAi) was used to suppress ATF6 expression. We noted that ATF6 expression in the ventricular myocardium was significantly increased in rats exposed to CIH. Furthermore, we noted that CIH preserved cardiac function after I/R in vivo and improved post-ischemic recovery of myocardial performance in isolated rat hearts. ATF6 and p-Akt expression was upregulated in cultured H9c2 cells exposed to chronic mild hypoxia compared with those cultured under normoxic conditions. Chronic mild hypoxia attenuated subsequent simulated I/R injury in H9c2 cells (48 h), as evidenced by increased cell viability and decreased LDH activity. By contrast, decreased cell viability and increased LDH activity were observed in siRNA-ATF6-transfected H9c2 cells, with a concomitant reduction in p-Akt levels. These results indicated that ATF6 upregulation is involved in the mechanism by which CIH attenuates myocardial I/R injury, possibly through upregulation of p-Akt, which is a key regulator of cardiomyocyte survival.

MicroRNA-146b inhibition augments hypoxia-induced cardiomyocyte apoptosis.

MicroRNAs (miRs) regulate a number of physiological and pathological processes, including myocardial chronic hypoxia. Previous studies revealed that the expression of miR-146b is increased in vitro and in vivo following the induction of hypoxia. In the present study, the role of miR­146b in hypoxic cardiomyocytes, and the mechanisms underlying its activity, were investigated. The expression of miR­146b was measured in tissue samples from patients with congenital heart disease by reverse transcription­quantitative polymerase chain reaction. The rat H9c2 cardiomyocyte cell line was transfected with an miR­146b inhibitor or the experimental controls, and the cells were maintained under hypoxic conditions for 72 h. The expression of miR­146b increased following the induction of hypoxia. Transfection with the miR­146b inhibitor enhanced the release of lactate dehydrogenase and increased hypoxia­induced apoptosis, as determined by terminal deoxynucleotidyl transferase dUTP nick­end labeling, Hoechst 33258 staining, JC­1 assay (measuring mitochondrial membrane permeability) and annexin V/propidium iodide analysis. A decreased expression of Bcl­2 was observed, whereas the expression levels of cleaved­caspase 3 and Bax were increased. Western blot analysis and a dual luciferase reporter assay confirmed that ribonuclease L is a direct target of miR­146b. Furthermore, inhibition of miR-146b increased the activation of nuclear factor-κB and signal transducer and activator of transcription 3. In conclusion, the inhibition of miR­146b may increase hypoxia-induced cardiomyocyte apoptosis.

Serum diamine oxidase as a hemorrhagic shock biomarker in a rabbit model.

BACKGROUND: In prolonged hemorrhagic shock, reductions in intestinal mucosal blood perfusion lead to mucosal barrier damage and systemic inflammation. Gastrointestinal failure in critically ill patients has a poor prognosis, so early assessment of mucosal barrier injury in shock patients is clinically relevant. Unfortunately, there is no serum marker that can accurately assess intestinal ischemia-reperfusion injury. OBJECTIVE: The aim of this study was to assess if serum diamine oxidase levels can reflect intestinal mucosal injury subsequent to prolonged hemorrhagic shock. METHODS: Thirty New Zealand white rabbits were divided into three groups: a control group, a medium blood pressure (BP) group (exsanguinated to a shock BP of 50 to 41 mm Hg), and a low BP group (exsanguinated to a shock blood pressure of 40 to 31 mm Hg), in which the shock BP was sustained for 180 min prior to fluid resuscitation. RESULTS: The severity of hemorrhagic shock in the low BP group was significantly greater than that of the medium BP group according to the post-resuscitation BP, serum tumor necrosis factor (TNF)-α, and arterial lactate. Intestinal damage was significantly more severe in the low BP group according to Chiu's scoring, claudin-1, intercellular adhesion molecule (ICAM)-1, and myeloperoxidase expression. Serum diamine oxidase was significantly increased in the low BP group compared to the medium BP and control groups and was negatively correlated with shock BP. CONCLUSION: Serum diamine oxidase can be used as a serological marker in evaluating intestinal injury and shows promise as an indicator of hemorrhagic shock severity.

Utilization of extracorporeal membrane oxygenation alleviates intestinal ischemia-reperfusion injury in prolonged hemorrhagic shock animal model.

Intestinal ischemia-reperfusion injury is one of the main factors leading to multiple organ failure after resuscitation of prolonged hemorrhagic shock; however, the current conventional fluid resuscitation still cannot effectively reduce intestinal injury caused by prolonged hemorrhagic shock. To investigate the effect of ECMO resuscitation on alleviating intestinal ischemia-reperfusion injury in a prolonged hemorrhagic shock rabbit model. Thirty New Zealand white rabbits were randomly divided into three groups: control group, conventional fluid resuscitation group, and ECMO resuscitation group. The prolonged hemorrhagic shock model was established by keeping the arterial blood pressure from 31 to 40 mmHg for 3 h through the femoral artery bleeding, and performing the resuscitation for 2 h by conventional fluid resuscitation and ECMO resuscitation, respectively. Chiu's score of intestinal injury, serum lactate and TNF-α levels, intestinal mucosamyeloperoxidase (MPO) activity, intercellular adhesion molecule (ICAM-1), and Claudin-1expression were detected. The mean arterial blood pressure in Group 2 was significantly higher after resuscitation than in Group 1, but serum lactate and inflammatory cytokines TNF-α level were significantly lower. And Chiu's score of intestinal injury and myeloperoxidase (MPO) activity level and ICAM-1 expression were significantly lower in the ECMO resuscitation group, in which the Claudin-1 levels were significantly increased. ECMO resuscitation for the prolonged hemorrhagic shock improves tissue perfusion and reduces the systemic inflammation, and thus alleviates intestinal damage caused by prolonged hemorrhagic shock.

[Clinical significance of cellular immune function in patients with viral myocarditis].

AIM: To probe into the clinical significance of cellular immune function in patients with viral myocarditis. METHODS: Choose our hospital in January 2008-December 2009 admitted during the 75 patients of viral myocarditis as the experimental group, 67 healthy patients served as control groups, respectively, and hospital admission within 24 h after the venous blood, before and after treatment T cell subsets, natural killer cells (NK) activity and tumor necrosis factor (TNF) were detected and analyzed. RESULTS: Compared with admission, 24 h after admission test in peripheral blood T lymphocyte subsets CD3, CD4, CD8 and CD4/CD8 were improved and CD3, CD4 and CD4/CD8 were statistically significant differences (P<0.05), but still slightly lower in the control group, while the difference was not statistically significant. At the same time, compared with admission, 24 h after admission of natural killer cells in the experimental group (NK) and tumor necrosis factor were improved, the differences were statistically significant (P<0.05), but still slightly lower than the control group, but the difference was not statistically significant. CONCLUSION: More in-depth study of viral myocarditis in patients with immune injury mechanism will help to better guide the treatment, and has important clinical significance.