Deletion of Tmem268 in mice suppresses anti-infectious immune responses by downregulating CD11b signaling.

Transmembrane protein 268 (TMEM268) is a novel, tumor growth-related protein first reported by our laboratory. It interacts with the integrin subunit ß4 (ITGB4) and plays a positive role in the regulation of the ITGB4/PLEC signaling pathway. Here, we investigated the effects and mechanism of TMEM268 in anti-infectious immune response in mice. Tmem268 knockout in mice aggravated cecal ligation and puncture-induced sepsis, as evidenced by higher bacterial burden in various tissues and organs, congestion, and apoptosis. Moreover, Tmem268 deficiency in mice inhibited phagocyte adhesion and migration, thus decreasing phagocyte infiltration at the site of infection and complement-dependent phagocytosis. Further findings indicated that TMEM268 interacts with CD11b and inhibits its degradation via the endosome-lysosome pathway. Our results reveal a positive regulatory role of TMEM268 in ß2 integrin-associated anti-infectious immune responses and signify the potential value of targeting the TMEM268-CD11b signaling axis for the maintenance of immune homeostasis and immunotherapy for sepsis and related immune disorders.

RNF115/BCA2 deficiency alleviated acute liver injury in mice by promoting autophagy and inhibiting inflammatory response.

The E3 ubiquitin ligase RING finger protein 115 (RNF115), also known as breast cancer-associated gene 2 (BCA2), has been linked with the growth of some cancers and immune regulation, which is negatively correlated with prognosis. Here, it is demonstrated that the RNF115 deletion can protect mice from acute liver injury (ALI) induced by the treatment of lipopolysaccharide (LPS)/D-galactosamine (D-GalN), as evidenced by decreased levels of alanine aminotransaminase, aspartate transaminase, inflammatory cytokines (e.g., tumor necrosis factor α and interleukin-6), chemokines (e.g., MCP1/CCL2) and inflammatory cell (e.g., monocytes and neutrophils) infiltration. Moreover, it was found that the autophagy activity in Rnf115-/- livers was increased, which resulted in the removal of damaged mitochondria and hepatocyte apoptosis. However, the administration of adeno-associated virus Rnf115 or autophagy inhibitor 3-MA impaired autophagy and aggravated liver injury in Rnf115-/- mice with ALI. Further experiments proved that RNF115 interacts with LC3B, downregulates LC3B protein levels and cell autophagy. Additionally, Rnf115 deletion inhibited M1 type macrophage activation via NF-κB and Jnk signaling pathways. Elimination of macrophages narrowed the difference in liver damage between Rnf115+/+ and Rnf115-/- mice, indicating that macrophages were linked in the ALI induced by LPS/D-GalN. Collectively, for the first time, we have proved that Rnf115 inactivation ameliorated LPS/D-GalN-induced ALI in mice by promoting autophagy and attenuating inflammatory responses. This study provides new evidence for the involvement of autophagy mechanisms in the protection against acute liver injury.

RIG-I Promotes Cell Viability, Colony Formation, and Glucose Metabolism and Inhibits Cell Apoptosis in Colorectal Cancer by NF-κB Signaling Pathway.

BACKGROUND: Retinoic acid-inducible gene-I (RIG-I) has crucial effects on various cancers, while RIG-I's detailed roles and mechanism in colorectal cancer (CRC) are uncovered. METHODS: qRT-PCR was used to detect the expression of RIG-I in CRC, adjacent nontumor specimens, and five cell lines. CCK-8, colony formation, and flow cytometry assays were conducted to study CRC cell viabilities. Extracellular acidification rates, lactate analysis, and ATP analysis were conducted to study the cell viabilities and glucose metabolism of CRC cells. Western blot is used to determine the proteins of NF-κBp65 in the nucleus and cytoplasm. RESULTS: This study revealed the upregulation of RIG-I in CRC tissues and cells and that high RIG-I expression was correlated with poor prognosis of CRC patients. In addition, silencing RIG-I inhibited cell viability as well as colony formation and promoted cell apoptosis in CRC cells, while RIG-I knockdown suppressed transplanted tumor growth and facilitated apoptosis in nude mice. Moreover, silencing RIG-I inhibited glucose metabolism by decreasing extracellular acidification rate, lactate production, adenosine triphosphate, and content of hypoxia-inducible factor 1α and pyruvate kinase isoform. 2.2-Deoxy-d-glucose, a glycolysis inhibitor, reduced the growth of CRC cells and promoted apoptosis in vitro and in vivo. In addition, RIG-I knockdown decreased NF-κB nuclear translocation. Besides, inhibiting NF-κB effectively eliminated RIG-I overexpression roles in cell viability and glucose metabolism in CRC cells. CONCLUSION: In summary, this study revealed that RIG-I mediated CRC cell proliferation, apoptosis, and glucose metabolism at least partly by NF-κB signaling pathway.

Does Helicobacter pylori infection affect the structure of bacteria in the gastric mucosa and fluid in patients with chronic antral gastritis?

This study aimed to evaluate the composition of the gastric microbiota in the gastric mucosa and gastric fluid of patients with chronic antral gastritis. Specifically, we sought to determine whether Helicobacter pylori (Hp) infection changes the bacterial community in the gastric mucosa or alters the microbiota in the gastric fluid. The bacterial community at another site in the stomach was also investigated. DNA was extracted from 160 samples collected from 40 patients with chronic antral gastritis (20 Hp-positive and 20 Hp-negative cases). Three tissue samples of the gastric mucosa (gastric angle, body, and antral mucosa) and one tube of gastric fluid were collected from every patient. A 16S rRNA amplification library was created, and high-throughput sequencing was performed. A profile of the community composition was obtained using bioinformatics methods, including cluster, taxonomy, and diversity analyses. Analysis of the gastric bacterial community revealed that the community compositions of the gastric mucosa and gastric fluid of patients without Hp are similar to but show differences from those of Hp-positive patients. The microbiota in Hp-positive patients exhibited reduced microbial diversity, and the gastric fluid of these patients contained a small proportion of Hp. The richness of Leptotrichia in mucosal samples was greater than that in gastric fluid samples from Hp-negative patients with chronic antral gastritis. Hp changes the growth of other microbiota in the mucosa and affects the microbiota in the gastric fluid of patients with chronic antral gastritis. In addition to Hp, the presence of other bacteria might be related to the development of chronic antral gastritis.

Stromal interaction molecule 1 promotes tumor growth in Esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is a disease with poor prognosis which urgently is in need of effective prognostic marker. To discover novel prognostic protein marker for ESCC, we applied a high-throughput monoclonal antibody microarray to compare tumor and adjacent non-tumor tissues from ESCC patients. Antibody #ESmAb270 was consistent higher expressed in tumors and it was identified via mass spectrometry to be stromal interaction molecule 1 (STIM1). STIM1 H scores in tumor tissues were significantly up-regulated in esophageal tumor tissues compared to non-tumor tissues in 105 ESCC patients. We also observed that high STIM1 expression was correlated with advanced tumor grade and poor prognosis of ESCC. In addition, attenuation of STIM1 by siRNA or chemical inhibitors significantly inhibited cell viability and migration of ESCC cells. Evidence from high-throughput monoclonal antibody microarray, IHC microarray with associated survival data and functional analysis show that STIM1 is an unfavorable prognostic biomarker in ESCC.

Impact of tumour size on metastasis and survival in patients with pancreatic neuroendocrine tumours (PNETs): A population based study.

Background: The relationship between tumour size and metastasis rate is poorly recognized in patients with pancreatic neuroendocrine tumours (PNETs). The impact of tumour size on prognosis was controversial in previous investigations. Methods: PNETs cases diagnosed from 1988 to 2013 were retrieved from the Surveillance, Epidemiology, and End Results (SEER) database. Clinicopathologic features were retrospectively analyzed. Survival was calculated by the Kaplan-Meier method. Multivariable Cox regression models with hazard ratios (HRs) were constructed to analyze survival outcomes and risk factors. Cubic spline analysis was used to assess relationship between tumor size and probability of metastasis. Results: A total of 5424 patients were identified, 1226 (22.6%) with tumour size of 20mm or less. The probability of metastasis increased in a non-linear fashion with increasing tumour size. Univariate analysis showed that tumour size was significantly correlated with survival (P<0.001), no matter surgery was performed or not. However, subgroup analysis suggested this association to be linear for patients with localized and regional tumours (P<0.001), but stochastic in patients with distant stages (P=0.703). On multivariate analysis, tumour size was an indicator for metastasis (HR=1.010, 95%CI: 1.008-1.013, P<0.001) and size≤20mm was an independent prognostic factor for good survival. For tumours≤20mm, surgical treatment was associated with significantly improved survival (P<0.001). Conclusions: Tumour size affects the probability of metastasis. Its prognostic impact on survival is restricted to patients with localized and regional disease. For patients with tumour size ≤20mm, surgical treatment should be considered preferably.

Klotho-mediated targeting of CCL2 suppresses the induction of colorectal cancer progression by stromal cell senescent microenvironments.

Senescent microenvironments play an important role in tumor progression. Here, we report that doxorubicin (DOX)-pretreated or replicative senescent stromal cells (WI-38 and HUVEC) promote colorectal cancer (CRC) cell growth and invasion in vitro and in vivo. These pro-tumorigenic effects were attenuated by exogenous administration of Klotho, an anti-aging factor. We subsequently identified several senescence-associated secretory phenotype (SASP)-associated genes, including CCL2, which were significantly upregulated in both types of senescent stromal cells during replication and DNA damage-induced senescence. Importantly, we found that the secretion of CCL2 by senescent stromal cells was significantly higher than that seen in nonsenescent cells or in senescent cells pretreated with Klotho. Notably, CCL2 was found to accelerate CRC cell proliferation and invasion, while this effect could be blocked by administration of a specific CCR2 antagonist. We further show that Klotho can suppress NF-κB activation during DOX-induced senescence and thus block CCL2 transcription. Low expression of Klotho, or high expression of CCL2 in patient tumor tissues, correlated with poor overall survival of CRC patients. Collectively, our findings suggest that senescent stromal cells are linked to progression of CRC. Klotho can suppress the senescent stromal cell-associated triggering of CRC progression by inhibiting the expression of SASP factors including CCL2. The identification of key SASP factors such as CCL2 may provide potential therapeutic targets for improving CRC therapy.

Usefulness of the thread-traction method in endoscopic full-thickness resection for gastric submucosal tumor: a comparative study.

BACKGROUND: Endoscopic full-thickness resection (EFTR) has shown great prospects in treating gastric submucosal tumors (SMTs) from the muscularis propria. However, it is very difficult sometimes to ideally expose the tumor and gain adequate visualization for the dissection site. In the present study, we applied the thread-traction (TT) method to assist EFTR in treating gastric SMTs and investigated the feasibility and effectiveness of this strategy. METHODS: A total of 28 patients were involved in the study. 13 patients were treated by TT-assisted EFTR (TT group) and the others by non-assisted EFTR (NA group). Data on clinical characteristics and therapeutic outcomes were collected for analysis. RESULTS: The average tumor size was 1.6 ± 0.4 cm. En bloc resection rate was 92.9%. Histopathological evaluation indicated that 22 tumors were gastrointestinal stromal tumors (78.6%), all at low- or very low-risk, and 6 tumors were leiomyomas (21.4%). The total complication rate was 32.1%. All complications were managed intra-operatively or conservatively. Both the total procedure time and the perforation time were significantly shorter in patients of TT group than those of NA group (71.9 ± 30.5 vs. 107.5 ± 35.8 min, P = 0.010; 38.3 ± 22.0 vs. 68.6 ± 24.2 min, P = 0.002). The pain score evaluated by visual analogue system after operation was significantly lower in patients of TT group than those of NA group (4.5 ± 1.1 vs. 5.8 ± 1.4, P = 0.014). Although complication rate was lower in patients of TT group than those of NA group, the difference was not statistically significant (15.4% vs. 46.7%, P = 0.114). No residual or recurrent tumors were observed during a mean follow-up period of 17.9 ± 4.4 months. CONCLUSIONS: The TT method could effectively assist EFTR to shorten operation time and decrease the risk of complications.

Comparison of synchronous dual wavelength diode laser versus conventional endo-knives for esophageal endoscopic submucosal dissection: an animal study.

BACKGROUND: Endoscopic submucosal dissection (ESD) is widely used for en bloc resection in early gastrointestinal cancer. However, it is technically complex with long procedure time and high adverse event rates with using conventional knives. The aim of this animal study was to verify the feasibility of ESD using a novel 980/1470 nm dual diode laser (DDL-ESD) in comparison with conventional knives (C-ESD) in esophagus. METHODS: This was an in vivo animal study using eight porcine models. Four were allocated in DDL-ESD group and four were in C-ESD group. Every model underwent two ESDs to remove half circumferential esophageal mucosa. Each model's esophagus was harvested during postmortem at 24 h after ESD. Each specimen underwent gross inspection and histopathological examination was carried out. Procedure time, completeness of en bloc resection, adverse events (bleeding and perforation) and histological injury to muscularis propria were assessed. RESULTS: A total of 16 ESD procedures were performed with 100% en bloc resection rate. The procedure speed in DDL-ESD group was significantly faster as compared to C-ESD group (0.27 cm2/min vs. 0.21 cm2/min, p = 0.001). The number of intraoperative bleeding points and the use of haemostatic forceps were significantly lesser in DDL-ESD group (4 ± 2 vs. 8 ± 3, p = 0.016; 1 ± 1 vs. 3 ± 2, p = 0.029). Histological assessment showed that injury to muscularis propria in DDL-ESD was milder than C-ESD. There was no perforation observed in both groups. CONCLUSIONS: DDL-ESD technique appears to be safer and faster than C-ESD with less bleeding and injury to deep tissues.

Deployment of carboxymethyl cellulose sheets to prevent esophageal stricture after full circumferential endoscopic submucosal dissection: A porcine model.

BACKGROUND AND AIM: Esophageal stricture is a serious adverse event secondary to extensive endoscopic submucosal dissection (ESD). The present study aimed to investigate the efficacy of carboxymethyl cellulose (CMC) sheets for the prevention of esophageal stricture after full circumferential ESD in an animal model. METHODS: Fourteen porcine models were randomized into a control group (n = 7) and a CMC group (n = 7). Five-centimeter-long circumferential esophageal ESD was carried out at a distance of 40 to 45 cm from the incisors in all models. In the CMC group, CMC sheets were placed over the mucosal defect completely after ESD, whereas the control group underwent routine ESD only. Endoscopic examination was conducted after the first and second week post-ESD. Esophageal specimens were harvested during post-mortem and were evaluated for macroscopic and histological appearance. Blood serum levels of four pro-inflammatory or profibrotic cytokines were measured quantitatively. RESULTS: The CMC group had better food tolerability during the second week post-ESD. The CMC group showed a significantly lower esophageal mucosal stricture rate compared to the control group. Histological assessments showed less fibrosis in the submucosal layer, milder damage to the muscularis propria, and enhanced re-epithelization in the CMC group. Serum transforming growth factor beta 1 levels were significantly lower in the CMC group post-ESD. CONCLUSION: Deployment of CMC sheets on the mucosal defect appears to be a promising method for preventing esophageal strictures after extensive ESD.

Zhen-wu-tang protects against podocyte injury in rats with IgA nephropathy via PPARγ/NF-κB pathway.

Zhen-wu-tang (ZWT) has been widely applied in chronic kidney diseases. However, the mechanism of ZWT remains unclear. Peroxisome proliferator-activated receptors-γ (PPARγ) is known as a protective factor for podocyte and kidney function. This study is aimed to investigate the protective effects of ZWT on IgA nephropathy (IgAN) in rats against podocyte injury and the underlying mechanism related to PPARγ. IgAN model rats were induced by administering bovine serum albumin, lipopolysaccharide, and carbon tetrachloride. ZWT at two doses and GW9662 (PPARγ antagonist) was administered once daily for 4 weeks respectively. Cultured podocyte induced by LPS were used to evaluate the podocyte-protective effect and related mechanism of ZWT in vitro. Results showed that ZWT observably reduced proteinuria and hematuria excretion, as well as the levels of blood urea nitrogen, serum creatinine, serum uric acid, low-density lipoprotein cholesterol, total cholesterol and triglycerides, but increased the contents of high-density lipoprotein cholesterol, ameliorating renal function and hyperlipidemia state in IgAN rats. Besides, both ZWT administration groups alleviated kidney pathological lesion, macrophage infiltration, IgA and C3 deposition in glomeruli. To further demonstrate the protective effects of ZWT, we found that podocyte damage was markedly ameliorated with ZWT treatments in IgAN rats and LPS-induced podocyte injury model by suppressing the expressions of desmin, reducing podocyte apoptosis and augmenting nephrin and podocin levels. Moreover, ZWT inhibited the phosphorylation of NF-κB and IκBα, simultaneously upregulated PPARγ. However, GW9662 made no difference in all the above effects compared to the model group, and was reversed by ZWT in vitro study. In conclusion, these results demonstrated that ZWT ameliorated IgAN-induced podocyte injury via upregulation PPARγ and the underlying mechanism might involve the inhibition of NF-κB pathway.

Cyclovirobuxinum D alleviates cardiac hypertrophy in hyperthyroid rats by preventing apoptosis of cardiac cells and inhibiting the p38 mitogen-activated protein kinase signaling pathway.

OBJECTIVE: To investigate the underlying mechanisms of cyclovirobuxinum D (Cvb-D) on alleviating cardiac hypertrophy in rats. METHODS: Sprague-Dawley rats were randomly divided into 5 groups: control group; levothyroxine-induced cardiac hypertrophy group (model); levothyroxine-induced cardiac hypertrophy + Cvb-D group (Cvb-D); levothyroxine-induced cardiac hypertrophy + captopril group (captopril); levothyroxine-induced cardiac hypertrophy + SB203580 group (SB203580), n=10 for each group. Rats were daily administered the respective drugs continuously for14 days by gastric gavage. A rat model of cardiac hypertrophy was established by intraperitoneal injection of levothyroxine to investigate whether Cvb-D protects against cardiac hypertrophy by inhibiting the p38 mitogen-activated protein kinase (MAPK) signaling pathway and preventing apoptosis of cardiac cells. RESULTS: Treatment with Cvb-D significantly deceased left ventricle hypertrophy, improved the histopathology, hemodynamic conditions, and cardiac function in rats with cardiac hypertrophy. Compared with the normal control group, in rats with cardiac hypertrophy, expression of bax in the heart and phospho-p38 MAPK protein levels were significantly up-regulated (P<0.01 or 0.05), whereas the bcl-2 protein level was down-regulated (P<0.01). In contrast, Cvb-D treatment reversed the changes in bax and phospho-p38 MAPK protein levels but increased the bcl-2 protein level (P<0.01 or 0.05), and these effects were similar to those of captopril and SB203580 (a specific p38MAPK inhibitor) treatment. Furthermore, both Cvb-D, captopril and SB203580 reduced mRNA expression of p38α, p38ß, c-fos, and c-jun mRNA, and Cvb-D had a stronger effect (P<0.01). CONCLUSION: These results demonstrate that Cvb-D protects against cardiac hypertrophy, which is possibly mediated by prevention of cardiac cell apoptosis and inhibition of the p38MAPK signaling pathway.

Protection Effect of Zhen-Wu-Tang on Adriamycin-Induced Nephrotic Syndrome via Inhibiting Oxidative Lesions and Inflammation Damage.

Zhen-wu-tang (ZWT), a well-known formula in China, is widely used to treat chronic kidney diseases. However, very little information on ZWT's mechanism of action is currently available. In this study, we investigated the possible protective role and underlying mechanism of ZWT on nephrotic syndrome (NS) induced by Adriamycin (intravenous injection, 6.0 mg/kg) in rats using biochemical and histopathological approaches. ZWT decreased urine protein excretion and the serum levels of total cholesterol, triglycerides, blood urea nitrogen, and creatinine significantly in diseased rats. A decrease in plasma levels of total protein and albumin was also recorded in nephropathic rats. Pathological results show an improved pathological state and recovering glomerular structure in ZWT treatment groups. ZWT decreased renal IL-8 level but increased renal IL-4 level. In addition, rats subjected to ZWT exhibited less IgG deposition in glomerulus compared with model group. RT-PCR results showed that ZWT decreased the mRNA expression of NF- κ B p65 and increased the mRNA expression of I κ B. Furthermore, ZWT reduced the level of MDA and increased SOD activity. These results demonstrated that ZWT ameliorated Adriamycin-induced NS in rats possibly by inhibiting Adriamycin-induced inflammation damage, enhancing body's antioxidant capacity, thereby protecting glomerulus from injury.

Qi-Dan Fang ameliorates adriamycin-induced nephrotic syndrome rat model by enhancing renal function and inhibiting podocyte injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Nephrotic syndrome (NS) is a clinical syndrome with a variety of causes, mainly characterized by heavy proteinuria. Podocyte injury plays a key role in proteinuria, one of the principal means for the control of NS is to prevent podocyte injury. Qi-Dan Fang consists of two of the most extensively applied herbal remedies among Traditional Chinese Medicine (TCM) (Radix Astragali Mongolici and Radix Salviae Miltiorrhizae, with a weight ratio of 5:1) which are specifically used for the treatment of various kidney diseases. In previous studies, we found that Qi-Dan Fang provides improvement to patients with adriamycin-induced nephrotic syndrome by alleviating proteinuria and serum lipid. The aim of this study is to study the efficiency of Qi-Dan Fang on NS model rat with renal dysfunction and podocyte injury, something which has not been carried out yet. MATERIALS AND METHODS: The rats were divided into Normal, Model, Jin Gui Shen Qi Pill (4.12 g/kg), Qi-Dan Fang (3.09, 6.17 and 12.34 g/kg/d) groups, they were each given a single tail intravenous injection of Adriamycin (6.0 mg/kg) except for the Normal group and were orally administered dosages of Qi-Dian Fang and Jin Gui Shen Qi pills once daily for 7 weeks. Following the treatment, the content of cystation C (CysC), blood urea nitrogen (BUN), serum creatinine (Scr) were measured with an autobiochemical analyser. The pathomorphological changes to the glomeruli, the mRNA expressions of nephrin, podocin, CD2AP genes and p53, bax, bcl-2 proteins expressions were also carried out to probe the effects of Qi-Dan Fang. RESULTS: (1) Qi-Dan Fang treatment raised the level of CysC in blood serum while lowering the content of BUN and Scr in the adriamycin-induced nephrotic syndrome rat model; (2) Long-term administration of Qi-Dan Fang was able to ameliorate pathomorphological change of glomeruli and repair the organization structure of Glomerulus; (3) Qi-Dan Fang could increase the mRNA expression of nephrin, podocin and CD2AP genes, down-regulate the expression of p53, bax proteins, while increased bcl-2 protein to protect the podocyte and restore Glomerular selective filtration function. CONCLUSIONS: Results of our present studies reveal that Qi-Dan Fang is able to enhance renal function, inhibit podocyte injury to provide improvements to the Adriamycin-induced nephrotic syndrome.