Myeloid vitamin D receptor regulates Paneth cells and microbial homeostasis.

Cross talk between immune cells and the intestinal crypt is critical in maintaining intestinal homeostasis. Recent studies highlight the direct impact of vitamin D receptor (VDR) signaling on intestinal and microbial homeostasis. However, the tissue-specific role of immune VDR signaling is not fully understood. Here, we generated a myeloid-specific VDR knockout (VDRΔLyz ) mouse model and used a macrophage/enteroids coculture system to examine tissue-specific VDR signaling in intestinal homeostasis. VDRΔLyz mice exhibited small intestine elongation and impaired Paneth cell in maturation and localization. Coculture of enteroids with VDR-/- macrophages increased the delocalization of Paneth cells. VDRΔLyz mice exhibited significant changes in the microbiota taxonomic and functional files, and susceptibility to Salmonella infection. Interestingly, loss of myeloid VDR impaired Wnt secretion in macrophages, thus inhibiting crypt ß-catenin signaling and disrupting Paneth cell differentiation in the epithelium. Taken together, our data have demonstrated that myeloid cells regulate crypt differentiation and the microbiota in a VDR-dependent mechanism. Dysregulation of myeloid VDR led to high risks of colitis-associated diseases. Our study provided insight into the mechanism of immune/Paneth cell cross talk in regulating intestinal homeostasis.

Paneth Cell Alertness to Pathogens Maintained by Vitamin D Receptors.

BACKGROUND AND AIMS: Vitamin D exerts a regulatory role over mucosal immunity via the vitamin D receptor (VDR). Although Paneth cells and their products are known to regulate the commensal and pathogenic microbiota, the role that VDRs in Paneth cells play in these responses is unknown. METHODS: We identified the decreased intestinal VDR significantly correlated with reduction of an inflammatory bowel disease risk gene ATG16L1 and Paneth cell lysozymes in patients with Crohn's disease. We generated Paneth cell-specific VDR knockout (VDRΔPC) mice to investigate the molecular mechanisms. RESULTS: Lysozymes in the Paneth cells were significantly decreased in the VDRΔPC mice. Isolated VDRΔPC Paneth cells exhibited weakened inhibition of pathogenic bacterial growth and displayed reduced autophagic responses. VDRΔPC mice had significantly higher inflammation after Salmonella infections. VDRΔPC mice also showed high susceptibility to small intestinal injury induced by indomethacin, a nonsteroidal anti-inflammatory drug. Co-housing of VDRΔPC and VDRlox mice made the VDRΔPC less vulnerable to dextran sulfate sodium colitis, suggesting the transmission of protective bacterial from the VDRlox mice. Thus, a lack of VDR in Paneth cells leads to impaired antibacterial activities and consequently increased inflammatory responses. Genetically and environmentally regulated VDRs in the Paneth cells may set the threshold for the development of chronic inflammation, as observed in inflammatory bowel diseases. CONCLUSIONS: We provide new insights into the tissue-specific functions of VDRs in maintaining Paneth cell alertness to pathogens in intestinal disorders. Targeting the VDR affects multiple downstream events within Paneth cells that inhibit intestinal inflammation and establish host defense against enteropathogens.

Imbalance of autophagy and apoptosis in intestinal epithelium lacking the vitamin D receptor.

Apoptosis and autophagy are dynamic processes that determine the fate of cells. Vitamin D receptor (VDR) deficiency in the intestine leads to abnormal Paneth cells and impaired autophagy function. Here, we will elucidate the mechanisms of the intestinal epithelial VDR regulation of autophagy and apoptosis. We used in vivo VDRlox and VDR∆IEC mice and ex vivo organoids generated from small intestine and colon tissues. We found that VDR deficiency induced more apoptotic cells and significantly increased cell death in the small intestine and colon of VDR∆IEC mice. The proapoptotic protein B-cell lymphoma 2 (BCL-2) associated X protein (Bax) was enhanced, whereas autophagy related 16 like 1 (ATG16L1) and Beclin-1 were decreased in the intestines of VDRΔIEC mice. Apoptosis induced by Bax reduced autophagy by decreasing Beclin-1. Physical interactions between Beclin-1 and Bcl-2 were increased in the VDR-deficient epithelia from mice. The growth of VDR∆IEC organoids was significantly slower with fewer Paneth cells than that of VDR+/+ organoids. The expression levels of Beclin-1 and lysozyme were decreased in VDR∆IEC organoids. Bacterial endotoxin levels were high in the serum from VDR∆IEC mice and made mice susceptible to colitis. In the organoids and colitis IL-10-/- mice, vitamin D3 treatment increased VDR and ATG16L1 protein expression levels, which activated autophagic responses. In summary, intestinal epithelial VDR regulates autophagy and apoptosis through ATG16L1 and Beclin-1. Our studies provide fundamental insights into the tissue-specific function of VDR in modulating the balance between autophagy and apoptosis.-Lu, R., Zhang, Y.-G., Xia, Y., Sun, J. Imbalance of autophagy and apoptosis in intestinal epithelium lacking the vitamin D receptor.

Deletion of sorting nexin 27 suppresses proliferation in highly aggressive breast cancer MDA-MB-231 cells in vitro and in vivo.

BACKGROUND: Sorting Nexin 27 (SNX27) belongs to a family of sortin nexins and possesses a unique binding domain at the C-terminus which mediates protein-protein interaction in intracellular trafficking, membrane remodeling, organelle motility, and tight junctions. However, its role in cancer development, especially in vivo, remains largely unknown. METHODS: We have generated a stable SNX27 knockdown clone in a highly aggressive breast cancer cell line MDA-MB-231 using an inducible lentiviral shRNA system. Cell migration and proliferation of SNX27 knockdown (KD) cells were compared with wild-type (WT) cells by MTT and wound healing assay, respectively. The differences in colony formation between SNX27-KD and WT cells were detected by soft agar culture and matrigel 3D culture. Furthermore, tumor growth was examined in a xenograft nude mouse model using SNX27-KD and WT MDA-MB-231 cells. The critical EMT (epithelial-mesenchymal transition) regulators were examined in vitro and in vivo. RESULTS: The wound healing assay showed that SNX27 knockdown significantly decreased cell motility and proliferation. Colony formation in soft agar showed that the SNX27 knockdown cells formed significantly fewer and smaller colonies than the parental MDA-MB-231 cells. Western blots and immunostaining showed that knockdown of SNX27 led to increased expression of E-cadherin and ß-catenin proteins, which facilitate adhesion formation and reverse EMT. EMT is a cellular program that allows polarized, immotile epithelial cells to convert to motile mesenchymal cells, promoting carcinoma invasion. The expression levels of Vimentin, the transcription factor of EMT, and tight junction protein Claudin-5, were significantly diminished in the SNX27 knockdown cells. The expression of PCNA, the cell proliferation marker, was increased in SNX27-KD cells transfected with E-cadherin siRNA. In a xenograft nude mouse model, we found that knockdown of SNX27 significantly inhibited tumor growth. The tumors from mice with SNX27-KD cells showed less proliferation compared to tumors from mice injected with wildtype cells. The increase in E-cadherin and ß-catenin and decrease in Vimentin and Claudin-5 were observed in tumors of mice injected with SNX27-KD cells. CONCLUSIONS: Our data have demonstrated that SNX27 plays a crucial role in tumor growth in vitro and in vivo.

Salmonella enteritidis Effector AvrA Stabilizes Intestinal Tight Junctions via the JNK Pathway.

Salmonella pathogenesis studies to date have focused on Salmonella typhimurium, and the pathogenesis of a second major serotype, Salmonella enteritidis, is poorly understood. Salmonella spp. possess effector proteins that display biochemical activities and modulate host functions. Here, we generated a deletion mutant of the effector AvrA, S.E-AvrA-, and a plasmid-mediated complementary strain, S.E-AvrA-/pAvrA+ (S.E-AvrA+), in S. Enteritidis. Using in vitro and in vivo infection models, we showed that AvrA stabilizes epithelial tight junction (TJ) proteins, such as ZO-1, in human intestinal epithelial cells. Transepithelial electrical resistance was significantly higher in cells infected with S.E-AvrA+ than in cells infected with S.E-AvrA- Inhibition of the JNK pathway suppresses the disassembly of TJ proteins; we found that enteritidis AvrA inhibited JNK activity in cells infected with wild type or S.E-AvrA+ strains. Therefore, Enteritidis AvrA-induced ZO-1 stability is achieved via suppression of the JNK pathway. Furthermore, the S.E-AvrA- strain led to enhanced bacterial invasion, both in vitro and in vivo Taken together, our data reveal a novel role for AvrA in S. Enteritidis: Enteritidis AvrA stabilizes intestinal TJs and attenuates bacterial invasion. The manipulation of JNK activity and TJs in microbial-epithelial interactions may be a novel therapeutic approach for the treatment of infectious diseases.

Alcohol Injury Damages Intestinal Stem Cells.

BACKGROUND: Alcohol consumption is associated with intestinal injury including intestinal leakiness and the risk of developing progressive gastrointestinal cancer. Alcoholics have disruption of intestinal barrier dysfunction that persists weeks after stopping alcohol intake, and this occurs in spite of the fact that intestinal epithelial cells turn over every 3 to 5 days. The renewal and functional regulation of the intestinal epithelium largely relies on intestinal stem cells (ISCs). Chronic inflammation and tissue damage in the intestine can injure stem cells including accumulation of mutations that may result in ISC dysfunction and transformation. ISCs are a key element in intestinal function and pathology; however, very little is known about the effects of alcohol on ISCs. We hypothesize that dysregulation of ISCs is one mechanism by which alcohol induces long-lasting intestinal damage. METHODS: In Vivo: Small intestinal samples from alcohol- and control-fed mice were assessed for ISC markers (Lgr5 and Bmi1) and the changes of the ß-catenin signaling using immunofluorescent microscopy, Western blotting, and RT-PCR. Ex Vivo: Organoids were generated from small intestine tissue and subsequently exposed to alcohol and analyzed for ISC markers, ß-catenin signaling. RESULTS: Chronic alcohol consumption significantly decreased the expression of stem cell markers, Bmi1 in the small intestine of the alcohol-fed mice and also resulted in dysregulation of the ß-catenin signaling-an essential regulator of its target gene Lgr5 and ISC function. Exposure of small intestine-derived organoids to 0.2% alcohol significantly reduced the growth of the organoids, including budding, and total surface area of the organoid cultures. Alcohol also significantly decreased the expression of Lgr5, p-ß-catenin (ser552), and Bmi1 in the organoid model. CONCLUSIONS: Both chronic alcohol feeding and acute exposure of alcohol resulted in ISC dysregulation which might be one mechanism for alcohol-induced long-lasting intestinal damage.

Prevalence and Clinical Characteristics of Spontaneous Splenorenal Shunt in Liver Cirrhosis: A Retrospective Observational Study Based on Contrast-Enhanced Computed Tomography (CT) and Magnetic Resonance Imaging (MRI) Scans.

BACKGROUND This is a retrospective observational study evaluating the prevalence and clinical characteristics of spontaneous splenorenal shunt in liver cirrhosis. MATERIAL AND METHODS We included a total of 105 cirrhotic patients who were admitted to our hospital between June 2012 and December 2013 and underwent contrast-enhanced CT and/or MRI scans at admissions. Spontaneous splenorenal shunt was identified. Clinical and laboratory data were compared between cirrhotic patients with and without spontaneous splenorenal shunt. RESULTS The prevalence of spontaneous splenorenal shunt was 10.5% (11/105). The prevalence of hepatic encephalopathy was higher in patients with spontaneous splenorenal shunt than in those without spontaneous splenorenal shunt (18.2% vs. 4.3%, p=0.062), but the difference between them was not statistically significant. The prevalence of acute upper-gastrointestinal bleeding was lower in patients with spontaneous splenorenal shunt than in those without spontaneous splenorenal shunt (0% vs. 18.1%, p=0.205), but the difference between them was not statistically significant. Patients with spontaneous splenorenal shunt had significantly higher Child-Pugh scores (9.50±1.65 vs. 7.43±2.02, p=0.002) and MELD scores (11.26±7.29 vs. 5.67±6.83, p=0.017) than those without spontaneous splenorenal shunt. In-hospital mortality was similar between them (0% vs. 4.3%, p=1.000). CONCLUSIONS Spontaneous splenorenal shunt might be associated with worse liver function in liver cirrhosis, but not with in-hospital mortality.

Splenectomy Causes 10-Fold Increased Risk of Portal Venous System Thrombosis in Liver Cirrhosis Patients.

BACKGROUND Portal venous system thrombosis (PVST) is a life-threatening complication of liver cirrhosis. We conducted a retrospective study to comprehensively analyze the prevalence and risk factors of PVST in liver cirrhosis. MATERIAL AND METHODS All cirrhotic patients without malignancy admitted between June 2012 and December 2013 were eligible if they underwent contrast-enhanced CT or MRI scans. Independent predictors of PVST in liver cirrhosis were calculated in multivariate analyses. Subgroup analyses were performed according to the severity of PVST (any PVST, main portal vein [MPV] thrombosis >50%, and clinically significant PVST) and splenectomy. Odds ratios (ORs) and 95% confidence intervals (CIs) were reported. RESULTS Overall, 113 cirrhotic patients were enrolled. The prevalence of PVST was 16.8% (19/113). Splenectomy (any PVST: OR=11.494, 95%CI=2.152-61.395; MPV thrombosis >50%: OR=29.987, 95%CI=3.247-276.949; clinically significant PVST: OR=40.415, 95%CI=3.895-419.295) and higher hemoglobin (any PVST: OR=0.974, 95%CI=0.953-0.996; MPV thrombosis >50%: OR=0.936, 95%CI=0.895-0.980; clinically significant PVST: OR=0.935, 95%CI=0.891-0.982) were the independent predictors of PVST. The prevalence of PVST was 13.3% (14/105) after excluding splenectomy. Higher hemoglobin was the only independent predictor of MPV thrombosis >50% (OR=0.952, 95%CI=0.909-0.997). No independent predictors of any PVST or clinically significant PVST were identified in multivariate analyses. Additionally, PVST patients who underwent splenectomy had a significantly higher proportion of clinically significant PVST but lower MELD score than those who did not undergo splenectomy. In all analyses, the in-hospital mortality was not significantly different between cirrhotic patient with and without PVST. CONCLUSIONS Splenectomy may increase by at least 10-fold the risk of PVST in liver cirrhosis independent of severity of liver dysfunction.

Serum Liver Fibrosis Markers in the Prognosis of Liver Cirrhosis: A Prospective Observational Study.

BACKGROUND The prognostic role of serum liver fibrosis markers in cirrhotic patients remains unclear. We performed a prospective observational study to evaluate the effect of amino-terminal pro-peptide of type III pro-collagen (PIIINP), collagen IV (CIV), laminin (LN), and hyaluronic acid (HA) on the prognosis of liver cirrhosis. MATERIAL AND METHODS All patients who were diagnosed with liver cirrhosis and admitted to our department were prospectively enrolled. PIIINP, CIV, LN, and HA levels were tested. RESULTS Overall, 108 cirrhotic patients were included. Correlation analysis demonstrated that CIV (coefficient r: 0.658, p<0.001; coefficient r: 0.368, p<0.001), LN (coefficient r: 0.450, p<0.001; coefficient r: 0.343, p<0.001), and HA (coefficient r: 0.325, p=0.001; coefficient r: 0.282, p=0.004) levels, but not PIIINP level (coefficient r: 0.081, p=0.414; coefficient r: 0.090, p=0.363), significantly correlated with Child-Pugh and MELD scores. Logistic regression analysis demonstrated that HA (odds ratio=1.00003, 95% confidence interval [CI]=1.000004-1.000056, p=0.022) was significantly associated with the 6-month mortality. Receiver operating characteristics analysis demonstrated that the area under the curve (AUC) of HA for predicting the 6-month mortality was 0.612 (95%CI=0.508-0.709, p=0.1531). CONCLUSIONS CIV, LN, and HA levels were significantly associated with the severity of liver dysfunction, but might be inappropriate for the prognostic assessment of liver cirrhosis.

Intestinal epithelial vitamin D receptor deletion leads to defective autophagy in colitis.

OBJECTIVE: Vitamin D and the vitamin D receptor (VDR) appear to be important immunological regulators of inflammatory bowel diseases (IBD). Defective autophagy has also been implicated in IBD, where interestingly, polymorphisms of genes such as ATG16L1 have been associated with increased risk. Although vitamin D, the microbiome and autophagy are all involved in pathogenesis of IBD, it remains unclear whether these processes are related or function independently. DESIGN: We investigated the effects and mechanisms of intestinal epithelial VDR in healthy and inflamed states using cell culture models, a conditional VDR knockout mouse model (VDR(ΔIEC)), colitis models and human samples. RESULTS: Absence of intestinal epithelial VDR affects microbial assemblage and increases susceptibility to dextran sulfate sodium-induced colitis. Intestinal epithelial VDR downregulates expressions of ATG16L1 and lysozyme, and impairs antimicrobial function of Paneth cells. Gain and loss-of-function assays showed that VDR levels regulate ATG16L1 and lysozyme at the transcriptional and translational levels. Moreover, low levels of intestinal epithelial VDR correlated with reduced ATG16L1 and representation by intestinal Bacteroides in patients with IBD. Administration of the butyrate (a fermentation product of gut microbes) increases intestinal VDR expression and suppresses inflammation in a colitis model. CONCLUSIONS: Our study demonstrates fundamental relationship between VDR, autophagy and gut microbial assemblage that is essential for maintaining intestinal homeostasis, but also in contributing to the pathophysiology of IBD. These insights can be leveraged to define therapeutic targets for restoring VDR expression and function.

Vitamin D receptor pathway is required for probiotic protection in colitis.

Low expression of vitamin D receptor (VDR) and dysfunction of vitamin D/VDR signaling are reported in patients with inflammatory bowel disease (IBD); therefore, restoration of VDR function to control inflammation in IBD is desirable. Probiotics have been used in the treatment of IBD. However, the role of probiotics in the modulation of VDR signaling to effectively reduce inflammation is unknown. We identified a novel role of probiotics in activating VDR activity, thus inhibiting inflammation, using cell models and VDR knockout mice. We found that the probiotics Lactobacillus rhamnosus strain GG (LGG) and Lactobacillus plantarum (LP) increased VDR protein expression in both mouse and human intestinal epithelial cells. Using the VDR luciferase reporter vector, we detected increased transcriptional activity of VDR after probiotic treatment. Probiotics increased the expression of the VDR target genes, such as antimicrobial peptide cathelicidin, at the transcriptional level. Furthermore, the role of probiotics in regulating VDR signaling was tested in vivo using a Salmonella-colitis model in VDR knockout mice. Probiotic treatment conferred physiological and histologic protection from Salmonella-induced colitis in VDR(+/+) mice, whereas probiotics had no effects in the VDR(-/-) mice. Probiotic treatment also enhanced numbers of Paneth cells, which secrete AMPs for host defense. These data indicate that the VDR pathway is required for probiotic protection in colitis. Understanding how probiotics enhance VDR signaling and inhibit inflammation will allow probiotics to be used effectively, resulting in innovative approaches to the prevention and treatment of chronic inflammation.

Diagnostic Accuracy of APRI, AAR, FIB-4, FI, and King Scores for Diagnosis of Esophageal Varices in Liver Cirrhosis: A Retrospective Study.

BACKGROUND Aspartate aminotransferase-to-platelet ratio index (APRI), aspartate aminotransferase-to-alanine aminotransferase ratio (AAR), FIB-4, fibrosis index (FI), and King scores might be alternatives to the use of upper gastrointestinal endoscopy for the diagnosis of esophageal varices (EVs) in liver cirrhosis. This study aimed to evaluate their diagnostic accuracy in predicting the presence and severity of EVs in liver cirrhosis. MATERIAL AND METHODS All patients who were consecutively admitted to our hospital and underwent upper gastrointestinal endoscopy between January 2012 and June 2014 were eligible for this retrospective study. Areas under curve (AUCs) were calculated. Subgroup analyses were performed according to the history of upper gastrointestinal bleeding (UGIB) and splenectomy. RESULTS A total of 650 patients with liver cirrhosis were included, and 81.4% of them had moderate-severe EVs. In the overall analysis, the AUCs of these non-invasive scores for predicting moderate-severe EVs and presence of any EVs were 0.506-0.6 and 0.539-0.612, respectively. In the subgroup analysis of patients without UGIB, their AUCs for predicting moderate-severe varices and presence of any EVs were 0.601-0.664 and 0.596-0.662, respectively. In the subgroup analysis of patients without UGIB or splenectomy, their AUCs for predicting moderate-severe varices and presence of any EVs were 0.627-0.69 and 0.607-0.692, respectively. CONCLUSIONS APRI, AAR, FIB-4, FI, and King scores had modest diagnostic accuracy of EVs in liver cirrhosis. They might not be able to replace the utility of upper gastrointestinal endoscopy for the diagnosis of EVs in liver cirrhosis.

Overexpression of forkhead box C1 promotes tumor metastasis and indicates poor prognosis in hepatocellular carcinoma.

UNLABELLED: Recurrence and metastasis remain the most common causes of lethal outcomes in hepatocellular carcinoma (HCC) after curative resection. Thus, it is critical to discover the mechanisms underlying HCC metastasis. Forkhead box C1 (FoxC1), a member of the Fox family of transcription factors, induces epithelial-mesenchymal transition (EMT) and promotes epithelial cell migration. However, the role of FoxC1 in the progression of HCC remains unknown. Here, we report that FoxC1 plays a critical role in HCC metastasis. FoxC1 expression was markedly higher in HCC tissues than in adjacent noncancerous tissues. HCC patients with positive FoxC1 expression had shorter overall survival times and higher recurrence rates than those with negative FoxC1 expression. FoxC1 expression was an independent, significant risk factor for recurrence and survival after curative resection. FoxC1 overexpression induced changes characteristic of EMT and an increase in HCC cell invasion and lung metastasis. However, FoxC1 knockdown inhibited these processes. FoxC1 transactivated Snai1 expression by directly binding to the Snai1 promoter, thereby leading to the inhibition of E-cadherin transcription. Knockdown of Snai1 expression significantly attenuated FoxC1-enhanced invasion and lung metastasis. FoxC1 expression was positively correlated with Snai1 expression, but inversely correlated with E-cadherin expression in human HCC tissues. Additionally, a complementary DNA microarray, serial deletion, site-directed mutagenesis, and a chromatin immunoprecipitation assay confirmed that neural precursor cell expressed, developmentally down-regulated 9 (NEDD9), which promotes the metastasis of HCC cells, is a direct transcriptional target of FoxC1 and is involved in FoxC1-mediated HCC invasion and metastasis. CONCLUSIONS: FoxC1 may promote HCC metastasis through the induction of EMT and the up-regulation of NEDD9 expression. Thus, FoxC1 may be a candidate prognostic biomarker and a target for new therapies.

Upregulation of IL-23 expression in patients with chronic hepatitis B is mediated by the HBx/ERK/NF-κB pathway.

IL-23 is a newly discovered proinflammatory cytokine that contributes to the maintenance and expansion of Th17 cells. IL-23 has recently been identified as playing a critical role in a number of chronic inflammatory diseases. However, the regulatory mechanism of IL-23 in chronic hepatitis B (CHB) remains largely unknown. The aims of this study were to detect the expression of IL-23 in CHB patients and to explore the molecular mechanism of hepatitis B virus (HBV)-induced IL-23 expression. Serum levels and hepatic expression of IL-23 were significantly upregulated in CHB patients. A positive correlation was found between IL-23 expression and the histological activity index score, HBV DNA load, and serum alanine aminotransferase and aspartate aminotransferase levels. HBx protein increased IL-23 expression in a dose-dependent manner. It also aided in the nuclear translocation of NF-κB, which directly bound to the promoters of IL-23 subunits p19 and p40 to facilitate their transcription. NF-κB inhibitors blocked the effect of HBx on IL-23 induction, and NF-κB subunits p65 and p50 increased the augmented IL-23 expression. Inhibition of ERK1/2 activation and transfection with ERK dominant-negative plasmid significantly blocked the HBx-induced IL-23 expression. Furthermore, PI3K and Ras-MEK-MAPK inhibitors significantly decreased the ERK1/2 activation and IL-23 expression. Thus, we report a new molecular mechanism for HBV-induced IL-23 expression, which involves the activation of the ERK/NF-κB pathway by HBx, leading to the transactivation of the IL-23 p19 and p40 promoters.

DBC1 is over-expressed and associated with poor prognosis in colorectal cancer.

BACKGROUND: Deleted in breast cancer 1 (DBC1) was initially cloned from a region homozygously deleted in breast cancers, but its role in colorectal cancer remains unknown. The present study aims to examine the expression level of DBC1 and assess its prognostic value in human colorectal cancer. METHODS: Immunohistochemical staining was performed to detect the expression level of DBC1 in a series of 186 colorectal cancer patients. Immunohistochemical staining results were analyzed and compared statistically with various clinicopathological characters and overall survival. RESULTS: Compared with the corresponding non-tumor tissues, a higher expression level of DBC1 was detected in colorectal cancer (P < 0.01). Tissue microarray analysis revealed that DBC1 expression is significantly associated with tumor histological grade, TNM stage and metastatic status (P < 0.01). Importantly, Kaplan-Meier analysis showed that DBC1 expression is associated with shorter overall survival (P < 0.01). Univariate Cox regression suggested that DBC1 expression, poorly differentiation status and the presence of lymph node metastasis predict shorter overall survival in colorectal cancer (P < 0.05). Multivariate Cox regression analysis indicated that DBC1 acts as an independent prognostic factor in colorectal cancer (P < 0.01). CONCLUSIONS: These results suggest that DBC1 is over-expressed in colorectal cancer and that it might serve as a predictor for selecting patients at high risk of poor prognosis.

Probiotics and microbial metabolites maintain barrier and neuromuscular functions and clean protein aggregation to delay disease progression in TDP43 mutation mice.

Amyotrophic lateral sclerosis (ALS) is a neuromuscular disease. The ALS mice expressing human mutant of transactive response DNA binding protein of 43 kDa (hmTDP43) showed intestinal dysfunction before neuromuscular symptoms. We hypothesize that restoring the intestinal and microbial homeostasis with a bacterial metabolite or probiotics delays the ALS disease onset. We investigate the pathophysiological changes in the intestine and neurons, intestinal and blood-brain barriers, and inflammation during the ALS progression. We then cultured enteric glial cells (EGCs) isolated from TDP43 mice for mechanistic studies. TDP43 mice had significantly decreased intestinal mobility, increased permeability, and weakened muscle, compared with the age-matched wild-type mice. We observed increased hmTDP43 and Glial fibrillary acidic protein (GFAP), and decreased expression of α-smooth muscle actin (α-SMA), tight junction proteins (ZO-1 and Claudin-5) in the colon, spinal cord, and brain in TDP43 mice. TDP43 mice had reduced Butyryl-coenzyme A CoA transferase, decreased butyrate-producing bacteria Butyrivibrio fibrisolvens, and increased Bacteroides fragilis, compared to the WT mice. Serum inflammation cytokines (IL-6, IL-17, and IFN-γ) and LPS were elevated in TDP43 mice. EGCs from TDP43 mice showed aggregation of hmTDP43 associated with increased GFAP and ionized calcium-binding adaptor molecule (IBA1, a microglia marker). TDP43 mice treated with butyrate or probiotic VSL#3 had significantly increased rotarod time, increased intestinal mobility and decreased permeability, compared to the untreated group. Butyrate or probiotics treatment decreased the expression of GFAP, TDP43, and increased α-SMA, ZO-1, and Claudin-5 in the colon, spinal cord, and brain. Also, butyrate or probiotics treatment enhanced the Butyryl-coenzyme A CoA transferase, Butyrivibrio fibrisolvens, and reduced inflammatory cytokines in TDP43 mice. The TDP43 EGCs treated with butyrate or probiotics showed reduced GFAP, IBA1, and TDP43 aggregation. Restoring the intestinal and microbial homeostasis by beneficial bacteria and metabolites provide a potential therapeutic strategy to treat ALS.

Case Report: First attempt by off-label use of tenecteplase to treat acute extensive portal venous system thrombosis.

Acute extensive portal venous system thrombosis (PVST) can cause lethal complications. Herein, we have for the first time reported the use of anticoagulation combined with systemic thrombolysis by tenecteplase in a male patient with a diagnosis of acute extensive PVST but without liver cirrhosis. After thrombolytic therapy, abdominal pain obviously alleviated. However, urinary bleeding developed, which was reversible by stopping thrombolytic drugs. Finally, this case developed cavernous transformation of the portal vein without portal venous recanalization. In future, the efficacy and safety of tenecteplase should be explored in acute extensive PVST cases.

Intestinal Epithelial Axin1 Deficiency Protects Against Colitis via Altered Gut Microbiota.

Intestinal homeostasis is maintained by specialized host cells and the gut microbiota. Wnt/ß-catenin signaling is essential for gastrointestinal development and homeostasis, and its dysregulation has been implicated in inflammation and colorectal cancer. Axin1 negatively regulates activated Wnt/ß-catenin signaling, but little is known regarding its role in regulating host-microbial interactions in health and disease. Here, we aim to demonstrate that intestinal Axin1 determines gut homeostasis and host response to inflammation. Axin1 expression was analyzed in human inflammatory bowel disease datasets. To explore the effects and mechanism of intestinal Axin1 in regulating intestinal homeostasis and colitis, we generated new mouse models with Axin1 conditional knockout in intestinal epithelial cell (IEC; Axin1 ΔIEC) and Paneth cell (PC; Axin1 ΔPC) to compare with control (Axin1 LoxP; LoxP: locus of X-over, P1) mice. We found increased Axin1 expression in the colonic epithelium of human inflammatory bowel disease (IBD). Axin1 ΔIEC mice exhibited altered goblet cell spatial distribution, PC morphology, reduced lysozyme expression, and enriched Akkermansia muciniphila (A. muciniphila). The absence of intestinal epithelial and PC Axin1 decreased susceptibility to dextran sulfate sodium (DSS)-induced colitis in vivo. Axin1 ΔIEC and Axin1 ΔPC mice became more susceptible to DSS-colitis after cohousing with control mice. Treatment with A. muciniphila reduced DSS-colitis severity. Antibiotic treatment did not change the IEC proliferation in the Axin1 Loxp mice. However, the intestinal proliferative cells in Axin1 ΔIEC mice with antibiotic treatment were reduced compared with those in Axin1 ΔIEC mice without treatment. These data suggest non-colitogenic effects driven by the gut microbiome. In conclusion, we found that the loss of intestinal Axin1 protects against colitis, likely driven by epithelial Axin1 and Axin1-associated A. muciniphila. Our study demonstrates a novel role of Axin1 in mediating intestinal homeostasis and the microbiota. Further mechanistic studies using specific Axin1 mutations elucidating how Axin1 modulates the microbiome and host inflammatory response will provide new therapeutic strategies for human IBD.

Conditional Vitamin D Receptor Deletion Induces Fungal and Archaeal Dysbiosis and Altered Metabolites.

A vitamin D receptor (VDR) deficiency leads to the dysbiosis of intestinal bacteria and is associated with various diseases, including cancer, infections, and inflammatory bowel disease. However, the impact of a VDR deficiency on fungi and archaea is unknown. We conditionally deleted the VDR in Paneth cells (VDRΔPC), intestinal epithelial cells (VDRΔIEC), or myeloid cells (VDRΔLyz) in mice and collected feces for shotgun metagenomic sequencing and untargeted metabolomics. We found that fungi were significantly altered in each knockout (KO) group compared to the VDRLoxp control. The VDRΔLyz mice had the most altered fungi species (three depleted and seven enriched), followed by the VDRΔPC mice (six depleted and two enriched), and the VDRΔIEC mice (one depleted and one enriched). The methanogen Methanofollis liminatans was enriched in the VDRΔPC and VDRΔLyz mice and two further archaeal species (Thermococcus piezophilus and Sulfolobus acidocaldarius) were enriched in the VDRΔLyz mice compared to the Loxp group. Significant correlations existed among altered fungi, archaea, bacteria, and viruses in the KO mice. Functional metagenomics showed changes in several biologic functions, including decreased sulfate reduction and increased biosynthesis of cobalamin (vitamin B12) in VDRΔLyz mice relative to VDRLoxp mice. Fecal metabolites were analyzed to examine the involvement of sulfate reduction and other pathways. In conclusion, a VDR deficiency caused the formation of altered fungi and archaea in a tissue- and sex-dependent manner. These results provide a foundation about the impact of a host factor (e.g., VDR deficiency) on fungi and archaea. It opens the door for further studies to determine how mycobiome and cross-kingdom interactions in the microbiome community and metabolites contribute to the risk of certain diseases.

Role of scalp nerve block in improving the quality of rehabilitation in patients after meningioma resection: A randomized controlled clinical trial.

BACKGROUND: In this study, we analyzed whether scalp nerve block with ropivacaine can improve the quality of rehabilitation in patients after meningioma resection. METHODS: We included 150 patients who were undergoing craniotomy in our hospital and categorized them into 2 groups - observation group (patients received an additional regional scalp nerve block anesthesia) and control group (patients underwent intravenous general anesthesia for surgery), using the random number table method approach (75 patients in each group). The main indicator of the study was the Karnofsky Performance Scale scores of patients at 3 days postoperatively, and the secondary indicator was the anesthesia satisfaction scores of patients after awakening from anesthesia. The application value of different anesthesia modes was studied and compared in the 2 groups. RESULTS: Patients in the observation group showed better anesthesia effects than those in the control group, with significantly higher Karnofsky Performance Scale scores at 3 days postoperatively (75.02 vs 66.43, P < .05) and anesthesia satisfaction scores. Compared with patients in the control group, patients in the observation group had lower pain degrees at different times after the surgery, markedly lower dose of propofol and remifentanil for anesthesia, and lower incidence of adverse reactions and postoperative complications. In addition, the satisfaction score of the patients and their families for the treatment was higher and the results of all the indicators were better in the observation group than in the control group, with statistically significant differences (P < .05). CONCLUSION: Scalp nerve block with ropivacaine significantly improves the quality of short-term postoperative rehabilitation in patients undergoing elective craniotomy for meningioma resection. This is presumably related to the improvements in intraoperative hemodynamics, relief from postoperative pain, and reduction in postoperative nausea and vomiting.