Generation and characterization of mature hepatocyte organoids for liver metabolic studies.

Hepatocyte organoids (HOs) generated in vitro are powerful tools for liver regeneration. However, previously reported HOs have mostly been fetal in nature with low expression levels of metabolic genes characteristic of adult liver functions, hampering their application in studies of metabolic regulation and therapeutic testing for liver disorders. Here, we report development of novel culture conditions that combine optimized levels of triiodothyronine (T3) with the removal of growth factors to enable successful generation of mature hepatocyte organoids (MHOs) of both mouse and human origin with metabolic functions characteristic of adult livers. We show that the MHOs can be used to study various metabolic functions including bile and urea production, zonal metabolic gene expression, and metabolic alterations in both alcoholic liver disease and non-alcoholic fatty liver disease, as well as hepatocyte proliferation, injury and cell fate changes. Notably, MHOs derived from human fetal hepatocytes also show improved hepatitis B virus infection. Therefore, these MHOs provide a powerful in vitro model for studies of human liver physiology and diseases. The human MHOs are potentially also a robust research tool for therapeutic development.

Roflumilast inhibits tumor growth and migration in STK11/LKB1 deficient pancreatic cancer.

Pancreatic cancer is a malignant tumor of the digestive system. It is highly aggressive, easily metastasizes, and extremely difficult to treat. This study aimed to analyze the genes that might regulate pancreatic cancer migration to provide an essential basis for the prognostic assessment of pancreatic cancer and individualized treatment. A CRISPR knockout library directed against 915 murine genes was transfected into TB 32047 cell line to screen which gene loss promoted cell migration. Next-generation sequencing and PinAPL.py- analysis was performed to identify candidate genes. We then assessed the effect of serine/threonine kinase 11 (STK11) knockout on pancreatic cancer by wound-healing assay, chick agnosia (CAM) assay, and orthotopic mouse pancreatic cancer model. We performed RNA sequence and Western blotting for mechanistic studies to identify and verify the pathways. After accelerated Transwell migration screening, STK11 was identified as one of the top candidate genes. Further experiments showed that targeted knockout of STK11 promoted the cell migration and increased liver metastasis in mice. Mechanistic analyses revealed that STK11 knockout influences blood vessel morphogenesis and is closely associated with the enhanced expression of phosphodiesterases (PDEs), especially PDE4D, PDE4B, and PDE10A. PDE4 inhibitor Roflumilast inhibited STK11-KO cell migration and tumor size, further demonstrating that PDEs are essential for STK11-deficient cell migration. Our findings support the adoption of therapeutic strategies, including Roflumilast, for patients with STK11-mutated pancreatic cancer in order to improve treatment efficacy and ultimately prolong survival.

The prevalence and factors associated with posttraumatic growth after 3-years outbreak of COVID-19 among resident physicians in China: a cross-sectional study.

Introduction: The Coronavirus disease 2019 (COVID-19) pandemic is a global traumatic event that has profoundly struck individuals' mental health. However, this might potentially promote positive transformation such as posttraumatic growth (PTG). Studies have indicated that the COVID-19 pandemic negatively affected the well-being of resident physicians, but little is known about PTG among this vulnerable population in China. Therefore, this study investigated the prevalence and associated factors of PTG among Chinese resident physicians after 3-years outbreak of COVID-19. Methods: An online survey was conducted from 9 March to 20 March in 2023. PTG was assessed using the 10-item Posttraumatic Growth Inventory-Short Form (PTGI-SF). Scores ≥30 implied moderate-to-high PTG. We also collected possible associated factors for PTG, including socio-demographic and psychological variables. Data was analyzed by applying descriptive statistics, univariable and multivariable logistic regression models. Results: In total, 2267 Chinese resident physicians provided validated data. 38.7% of them reported moderate-to-high PTG. In the multivariable logistic regression models, age (odds ratio, OR = 1.039; 95% confidence interval, 95%CI = 1.008-1.070), female (OR = 1.383, 95%CI = 1.151-1.662), satisfied or neutral with annual income (OR = 2.078, 95%CI = 1.524-2.832; OR = 1.416, 95%CI = 1.157-1.732), sufficient support at work (OR = 1.432, 95%CI = 1.171-1.751) and resilience (OR = 1.171, 95%CI = 1.096-1.252) were significantly positively associated with moderate-to-high PTG. On the contrary, burnout (OR = 0.653, 95%CI = 0.525-0.812), depression symptoms (OR = 0.700, 95%CI = 0.552-0.889), and stress (OR = 0.757, 95%CI = 0.604-0.949) were significantly negatively associated with moderate-to-high PTG. Discussion: Overall, resident physicians in China experienced relatively high prevalence of PTG that could be associated with several psychosocial factors. Findings may provide evidence to develop interventions for resident physicians to systematically and constructively process traumatic events related to the pandemic and foster their PTG.

Soil moisture and pH differentially drive arbuscular mycorrhizal fungal composition in the riparian zone along an alpine river of Nam Co watershed.

The riparian zone is an important ecological corridor connecting the upstream and downstream rivers. Its highly complex biological and physical environments significantly affect the biogeographical pattern of species and various ecosystem functions. However, in alpine riparian ecosystems, the distribution patterns and drivers of arbuscular mycorrhizal (AM) fungi, a group of functionally important root-associated microorganisms, remain poorly understood. In this study, we investigated the AM fungal diversity and community composition in near-bank (wetland) and far-bank (alpine meadows) soils along the Niaqu River in the Nam Co watershed, and assessed the relative importance of abiotic and biotic filtering in shaping these distributions. Overall, 184 OTUs were identified in the riparian ecosystem, predominantly belonging to the genus Glomus, especially in the downstream soils, and Claroideoglomus in near-bank soils. AM fungal colonization, spore density, and α diversity showed an overall increasing trend along the river, while the extraradical hyphae declined dramatically from the middle of the river. AM fungal communities significantly varied between the wetland and alpine meadows in the riparian zone, mainly driven by the geographic distance, soil water content, soil pH, and plant communities. Specifically, soil pH was the principal predictor of AM fungal community in near-bank wetland soils, while soil water content had a most substantial direct effect in alpine meadows. These findings indicate that abiotic factors are the most important divers in shaping AM fungal communities at the watershed scale, which could be helpful in alpine riparian biodiversity conservation and management.

Integrated metabolomics and transcriptomic analysis of the flavonoid regulatory networks in Sorghum bicolor seeds.

BACKGROUND: The objective of this study was to reveal the flavonoid biosynthesis pathway in white (Z6), red (Z27) and black (HC4) seeds of the sweet sorghum (Sorghum bicolor) using metabolomics and transcriptomics, to identify different flavonoid metabolites, and to analyze the differentially expressed genes involved in flavonoid biosynthesis. RESULTS: We analyzed the metabolomics and transcriptomics data of sweet sorghum seeds. Six hundred and fifty-one metabolites including 171 flavonoids were identified in three samples. Integrated analysis of transcriptomics and metabolomics showed that 8 chalcone synthase genes (gene19114, gene19115, gene19116, gene19117, gene19118, gene19120, gene19122 and gene19123) involved in flavonoid biosynthesis, were identified and play central role in change of color. Six flavanone including homoeriodictyol, naringin, prunin, naringenin, hesperetin and pinocembrin were main reason for the color difference. CONCLUSIONS: Our results provide valuable information on the flavonoid metabolites and the candidate genes involved in the flavonoid biosynthesis pathway in sweet sorghum seeds.

Hydrochemical characteristics and formation mechanisms of groundwater in west Zoucheng City, Shandong Province, China.

Groundwater is an important water source for domestic, industrial, and agricultural use in the western part of Zoucheng, China. Understanding its hydrochemical characteristics and formation mechanisms is important for the sustainable development and utilization of groundwater. In this study, 36 water samples were collected during the wet and dry seasons, respectively, and the hydrochemical components such as K+, Na+, Ca2+, Mg2+, Cl-, SO42-, HCO3-, NO3-, F-, TH, and TDS were analyzed. A graphical method, correlation analysis, and principal component analysis were applied to explore the hydrochemical characteristics and evolution mechanisms of groundwater in the study area. The results show that the orders of the anion and cation concentrations of karst groundwater and pore groundwater are Ca2+ > Na+ > Mg2+ > K+ and HCO3- > SO42- > Cl- > NO3- > F-, respectively. On the whole, the karst groundwater quality is better than the pore groundwater quality, which in turn is better than the surface water quality. In addition, water quality in the dry season is better than water quality in the wet season for all the three water sources. The hydrochemical types of groundwater are complex and changeable. Compared with dry seasons, HCO3 and SO4 type water increase during the wet seasons, while the Cl type and Mg type water decrease. Na type is significantly more prevalent in pore groundwater than in karst groundwater. The chemical formations of karst groundwater and pore groundwater in the dry and wet seasons are mainly affected by water-rock interactions and human activities.

PLGA microspheres carrying miR-20a-5p improved intestinal epithelial barrier function in patients with Crohn's disease through STAT3-mediated inhibition of Th17 differentiation.

BACKGROUND: Recent studies have shown that microRNAs (miRNAs) are aberrantly expressed in patients with Crohn's disease (CD). This suggests that the aberrant expression of miRNAs may contribute to the development of CD. Currently, the specific miRNAs involved in CD development have not been clearly identified. Therefore, we aimed to identify CD-associated miRNAs and explore their functions. METHODS: miRNA microarray analysis was performed to screen for differentially expressed miRNAs in colon tissues from normal controls (NC) and CD patients. The identified miRNAs were validated using quantitative real-time PCR (qPCR). The therapeutic roles of miR-20a-5p mimics via the delivery of poly(lactic-co-glycolic acid) microspheres (PLGA MSs) were further investigated in IL-10-/- mice with spontaneous chronic colitis that were used as a model of CD. The target genes of miR-20a-5p and the associated signaling pathways were identified through bioinformatic analysis and experimental verification of the interactions between the targets predicted by the algorithms and dysregulated mRNAs. RESULTS: The analysis showed that miR-20a-5p was the most significantly downregulated miRNA in patients with CD. Treatment with PLGA MSs carrying miR-20a-5p significantly ameliorated the colitis, decreased mucosal inflammation, and improved epithelial barrier function. Bioinformatic analysis and experimental studies showed that miR-20a-5p inhibition enhanced Th17 differentiation and improved intestinal epithelial barrier function by targeting STAT3. CONCLUSIONS: Downregulation of miR-20a-5p improved the intestinal epithelial barrier function and prevented CD development through the STAT3/IL-17 signaling pathway. Therefore, the delivery of miR-20a-5p by PLGA MSs may serve as a potential therapeutic strategy for CD treatment.

Yap-Sox9 signaling determines hepatocyte plasticity and lineage-specific hepatocarcinogenesis.

BACKGROUND & AIMS: Primary liver tumors comprise distinct subtypes. A subset of intrahepatic cholangiocarcinoma (iCCA) can arise from cell fate reprogramming of mature hepatocytes in mouse models. However, the underpinning of cell fate plasticity during hepatocarcinogenesis is still poorly understood, hampering therapeutic development for primary liver cancer. As YAP activation induces liver tumor formation and cell fate plasticity, we investigated the role of Sox9, a transcription factor downstream of Yap activation that is expressed in biliary epithelial cells (BECs), in Yap-induced cell fate plasticity during hepatocarcinogenesis. METHODS: To evaluate the function of Sox9 in YAP-induced hepatocarcinogenesis in vivo, we used several genetic mouse models of inducible hepatocyte-specific YAP activation with simultaneous Sox9 removal. Cell fate reprogramming was determined by lineage tracing and immunohistochemistry. The molecular mechanism underlying Yap and Sox9 function in hepatocyte plasticity was investigated by transcription and transcriptomic analyses of mouse and human liver tumors. RESULTS: Sox9, a marker of liver progenitor cells (LPCs) and BECs, is differentially required in YAP-induced stepwise hepatocyte programming. While Sox9 has a limited role in hepatocyte dedifferentiation to LPCs, it is required for BEC differentiation from LPCs. YAP activation in Sox9-deficient hepatocytes resulted in more aggressive HCC with enhanced Yap activity at the expense of iCCA-like tumors. Furthermore, we showed that 20% of primary human liver tumors were associated with a YAP activation signature, and tumor plasticity is highly correlated with YAP activation and SOX9 expression. CONCLUSION: Our data demonstrated that Yap-Sox9 signaling determines hepatocyte plasticity and tumor heterogeneity in hepatocarcinogenesis in both mouse and human liver tumors. We identified Sox9 as a critical transcription factor required for Yap-induced hepatocyte cell fate reprogramming during hepatocarcinogenesis. LAY SUMMARY: Sox9, a marker of liver progenitor cells and bile duct lining cells, is a downstream target of YAP protein activation. Herein, we found that YAP activation in hepatocytes leads to a transition from mature hepatocytes to liver progenitor cells and then to bile duct lining cells. Sox9 is required in the second step during mouse hepatocarcinogenesis. We also found that human YAP and SOX9 may play similar roles in liver cancers.

A Comparative Transcriptional Landscape of Two Castor Cultivars Obtained by Single-Molecule Sequencing Comparative Analysis.

Background and Objectives: Castor (Ricinus communis L.) is an important non-edible oilseed crop. Lm-type female strains and normal amphiprotic strains are important castor cultivars, and are mainly different in their inflorescence structures and leaf shapes. To better understand the mechanisms underlying these differences at the molecular level, we performed a comparative transcriptional analysis. Materials and Methods: Full-length transcriptome sequencing and short-read RNA sequencing were employed. Results: A total of 76,068 and 44,223 non-redundant transcripts were obtained from high-quality transcripts of Lm-type female strains and normal amphiprotic strains, respectively. In Lm-type female strains and normal amphiprotic strains, 51,613 and 20,152 alternative splicing events were found, respectively. There were 13,239 transcription factors identified from the full-length transcriptomes. Comparative analysis showed a great variety of gene expression of common and unique transcription factors between the two cultivars. Meanwhile, a functional analysis of the isoforms was conducted. The full-length sequences were used as a reference genome, and a short-read RNA sequencing analysis was performed to conduct differential gene analysis. Furthermore, the function of DEGs were performed to annotation analysis. Conclusion: The results revealed considerable differences and expression diversity between the two cultivars, well beyond what was reported in previous studies and likely reflecting the differences in architecture between these two cultivars.

A self-amplifying loop of YAP and SHH drives formation and expansion of heterotopic ossification.

Heterotopic ossification (HO) occurs as a common complication after injury or in genetic disorders. The mechanisms underlying HO remain incompletely understood, and there are no approved prophylactic or secondary treatments available. Here, we identify a self-amplifying, self-propagating loop of Yes-associated protein (YAP)-Sonic hedgehog (SHH) as a core molecular mechanism underlying diverse forms of HO. In mouse models of progressive osseous heteroplasia (POH), a disease caused by null mutations in GNAS, we found that Gnas-/- mesenchymal cells secreted SHH, which induced osteoblast differentiation of the surrounding wild-type cells. We further showed that loss of Gnas led to activation of YAP transcription activity, which directly drove Shh expression. Secreted SHH further induced YAP activation, Shh expression, and osteoblast differentiation in surrounding wild-type cells. This self-propagating positive feedback loop was both necessary and sufficient for HO expansion and could act independently of Gnas in fibrodysplasia ossificans progressiva (FOP), another genetic HO, and nonhereditary HO mouse models. Genetic or pharmacological inhibition of YAP or SHH abolished HO in POH and FOP and acquired HO mouse models without affecting normal bone homeostasis, providing a previously unrecognized therapeutic rationale to prevent, reduce, and shrink HO.

Expression of Twist, Slug and Snail in esophageal squamous cell carcinoma and their prognostic significance.

Esophageal cancer is one of the most common types of malignancy worldwide. At present, surgical resection is the main treatment for esophageal cancer, but recurrence and distant metastasis are the main causes of mortality. The transcription factors Twist, Slug and Snail regulate epithelial-mesenchymal transition and thereby participate in tumor invasion and metastasis. The aim of the present study was to investigate the expression of Twist, Slug and Snail in esophageal squamous cell carcinoma (ESCC) and their prognostic significance. The expression of Twist, Slug and Snail in 229 paraffin-embedded ESCC and matched normal mucosal tissues was detected by immunohistochemistry. The expression differences of Twist, Slug and Snail in the ESCC and normal tissues were compared by χ2 test, and the associations between the three proteins and the clinicopathological parameters of ESCC were analyzed. The expression levels of Twist, Slug and Snail in 29 fresh frozen ESCC and matched normal mucosal tissues were detected by reverse transcription-quantitative PCR. The correlations among Twist, Slug and Snail in ESCC were examined by Pearson's correlation analyses. In addition, single factor and multivariate Cox regression analyses were used to analyze the influence of Twist, Slug and Snail on the prognosis of ESCC. Twist was found to be highly expressed in ESCC. The difference of Slug expression in ESCC was associated with differentiation degree, TNM stage and vascular invasion, but no significant association was observed between Snail expression and any clinicopathological parameters. In ESCC, there were significant differences in protein expression between Twist and Snail, and Slug and Snail. The mRNA expression level of Twist in ESCC was significantly higher than that in normal esophageal mucosa. However, the mRNA expression of Slug in normal esophageal mucosa was higher than that in ESCC, and the mRNA expression levels of Twist and Snail were positively correlated in ESCC. Kaplan-Meier analysis of 229 patients with ESCC revealed that Snail influenced the overall survival, as did the co-expression of Twist and Snail. Nerve invasion was also identified as an independent factor affecting the progression-free survival of ESCC. The results indicate that Twist is highly expressed, Slug may be a tumor suppressor, and Snail is an independent prognostic factor in ESCC. Twist and Snail are positively correlated, and the simultaneous inhibition of Twist and Snail protein expression may be beneficial for prolonging the overall survival of patients with ESCC.

The Expression and Prognostic Value of FGF2, FGFR3, and FGFBP1 in Esophageal Squamous Cell Carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma was treated by operation and chemoradiotherapy. However, the prognosis of most patients is poor after treatment, and most studies have shown that FGF2 and its receptor (FGFR) are involved in the development of various malignant tumors. FGF2 plays an important role in tumor progression and malignancy. In this study, the immunohistochemistry of FGF2, FGFR3, and FGFBP1 was used to further verify the expression of the three proteins in 172 patients with esophageal squamous cell carcinoma (ESCC) who had not received preoperative chemoradiotherapy and its effect on the prognosis of ESCC. METHODS: (1) χ 2 test was used to analyze the relationship between proteins and clinicopathological parameters. Survival analysis was used to investigate the effect of three proteins on prognosis. (2) Paired sample t-test was used to analyze the mRNA expression of the three proteins in fresh ESCC tissues and adjacent normal tissues. RESULTS: FGF2 was correlated with tumor size (p = 0.026), gender (p = 0.047), and lymph metastasis (p = 0.007) in ESCC tissues. The high expression of FGFR3 was associated with tumor differentiation (p = 0.043 and p < 0.05), lymph node metastasis (p = 0.078 and p < 0.1), and race (p = 0.033 and p < 0.05). The high expression of FGFBP1 was significantly associated with the degree of tumor differentiation (p = 0.012), age (p = 0.045), and lymph node metastasis (p = 0.032) of ESCC patients. The expression of FGF2, FGFR3, and FGFBP1-mRNA in ESCC tissues was significantly higher than that in adjacent tissues (p < 0.001, p < 0.001, and p = 0.001). Patients with high expression of FGF2, FGFBP1, and FGFR3 had poor prognosis. There was a weak positive correlation between FGF2 and FGFBP1, as well as FGFR. CONCLUSION: The FGF2-FGFR3 axis may promote the progression of esophageal squamous cell carcinoma. The FGF2-FGFR3 axis may be a new direction of targeted therapy for esophageal squamous cell carcinoma. FGF2 and FGFR3 may be used as prognostic markers of esophageal squamous cell carcinoma.

Metabolite Profiling of Sorghum Seeds of Different Colors from Different Sweet Sorghum Cultivars Using a Widely Targeted Metabolomics Approach.

Sweet sorghum (Sorghum bicolor) is one of the most important cereal crops in the world with colorful seeds. To study the diversity and cultivar-specificity of phytochemicals in sweet sorghum seeds, widely targeted metabolomics was used to analyze the metabolic profiles of the white, red, and purple seeds from three sweet sorghum cultivars Z6, Z27, and HC4. We identified 651 metabolites that were divided into 24 categories, including fatty acids, glycerolipids, flavonoids, benzoic acid derivatives, anthocyanins, and nucleotides and its derivatives. Among them, 217 metabolites were selected as significantly differential metabolites which could be related to the seed color by clustering analysis, principal component analysis (PCA), and orthogonal signal correction and partial least squares-discriminant analysis (OPLS-DA). A significant difference was shown between the red seed and purple seed samples, Z27 and HC4, in which 106 were downregulated and 111 were upregulated. The result indicated that 240 metabolites were significantly different, which could be related to the purple color with 58 metabolites downregulated and 182 metabolites upregulated. And 199 metabolites might be involved in the red phenotype with 54 downregulated and 135 upregulated. There were 45 metabolites that were common to all three cultivars, while cyanidin O-malonyl-malonyl hexoside, cyanidin O-acetylhexoside, and cyanidin 3-O-glucosyl-malonylglucoside were significantly upregulated red seeds, which could be the basis for the variety of seed colors. Generally, our findings provide a comprehensive comparison of the metabolites between the three phenotypes of S. bicolor and an interpretation of phenotypic differences from the point of metabolomics.

Piezo1/2 mediate mechanotransduction essential for bone formation through concerted activation of NFAT-YAP1-ß-catenin.

Mechanical forces are fundamental regulators of cell behaviors. However, molecular regulation of mechanotransduction remain poorly understood. Here, we identified the mechanosensitive channels Piezo1 and Piezo2 as key force sensors required for bone development and osteoblast differentiation. Loss of Piezo1, or more severely Piezo1/2, in mesenchymal or osteoblast progenitor cells, led to multiple spontaneous bone fractures in newborn mice due to inhibition of osteoblast differentiation and increased bone resorption. In addition, loss of Piezo1/2 rendered resistant to further bone loss caused by unloading in both bone development and homeostasis. Mechanistically, Piezo1/2 relayed fluid shear stress and extracellular matrix stiffness signals to activate Ca2+ influx to stimulate Calcineurin, which promotes concerted activation of NFATc1, YAP1 and ß-catenin transcription factors by inducing their dephosphorylation as well as NFAT/YAP1/ß-catenin complex formation. Yap1 and ß-catenin activities were reduced in the Piezo1 and Piezo1/2 mutant bones and such defects were partially rescued by enhanced ß-catenin activities.

The expression and prognostic value of SOX2, ß-catenin and survivin in esophageal squamous cell carcinoma.

Aim: We mainly explored the effect of SOX2, ß-catenin and survivin on prognosis in esophageal squamous cell carcinoma. Materials & methods: Immunohistochemistry was used to examine the expression of SOX2, ß-catenin and survivin. χ2 test was used to analyze the relationship between proteins and clinicopathological parameters. Survival analysis was used to investigate the effect of three proteins on prognosis. Results: SOX2 was related to lymph node metastasis (p = 0.004) and vascular invasion (p = 0.041). ß-catenin was associated with depth of invasion (p = 0.014), lymph node metastasis (p = 0.032) and postoperative chemoradiotherapy (p < 0.001). Survivin was related to gender (p = 0.022) and nerve invasion (p = 0.014). There was a positive correlation between SOX2 and ß-catenin. Patients with SOX2 and ß-catenin overexpression had poor prognosis. Survivin-positive patients who received postoperative chemoradiotherapy had a short time. Conclusion: SOX2, ß-catenin and survivin can be used as prognostic markers of esophageal squamous cell carcinoma.

The Silencing of SFRP2 Expression in ESCC Is Due to Methylation of the Gene Promoter.

OBJECTIVE: Our aim of the study was to investigate the expression level and methylation status of the secreted frizzled-related protein 2 in esophageal squamous cell carcinoma and to evaluate the clinical utility of the marker. MATERIAL AND METHODS: We first used Immunohistochemistry (ICH) to explore the expression level of secreted frizzled-related protein 2 protein in esophageal squamous cell carcinoma tissues and adjacent normal tissues and then used methylation-specific polymerase chain reaction and bisulfite sequencing polymerase chain reaction to detect methylation status of secreted frizzled-related protein 2. RESULTS: Secreted frizzled-related protein 2 expression was notably reduced in patients with esophageal squamous cell carcinoma, whereas methylation of secreted frizzled-related protein 2 was increased in the majority of esophageal squamous cell carcinoma specimens. CONCLUSION: Sum up, we have demonstrated the abnormal DNA hypermethylation, causing reduced or absent gene expression. Methylation testing of secreted frizzled-related protein 2 using epigenetic marker may be a significative screening method for patients with esophageal squamous cell carcinoma.

Dietary Nondigestible Polysaccharides Ameliorate Colitis by Improving Gut Microbiota and CD4+ Differentiation, as Well as Facilitating M2 Macrophage Polarization.

BACKGROUND: The aim of this study was to investigate the therapeutic mechanism of a specific multifiber mix diet (MF) designed to match the fiber content of a healthy diet in interleukin-10 knockout (IL-10-/- ) mice with spontaneous chronic colitis displaying similar characteristics to those of human Crohn's disease (CD). METHODS: Sixteen-week-old IL-10-/- mice were used for the experiments with MF diet for 4 weeks. Severity of colitis, the composition of the fecal microbiota, expression of Th1/Th17 cells, myeloperoxidase (MPO) concentrations, and inflammatory cytokines and chemokines (tumor necrosis factor-α [TNF-α], IL-6, macrophage inflammatory protein [MIP]-2, monocyte chemoattractant protein-1 [MCP-1], and MIP-1α), as well as arginase 1 (Arg1) and signal transducers and activators of transcription 6 (STAT6) proteins, were measured at the end of the experiment. In addition, the corresponding metabolites (short-chain fatty acids) of MF on CD4+ CD25+ Foxp3+ regulatory T cells (Tregs) were also detected in vivo and in vitro. RESULTS: MF treatment significantly ameliorated colitis associated with decreased lamina propria frequency of Th1/Th17 cells, MPO concentrations, and inflammatory cytokines and chemokines (TNF-α, IL-6, MIP-2, MCP-1, and MIP-1α). An increase in gut microbial diversity was observed after MF treatment compared with IL-10-/- mice, including a significant increase in bacteria belonging to the Firmicutes phylum and a significant decrease in bacteria belonging to the Proteobacteria phylum. Moreover, MF treatment increased the differentiation of CD4+ CD25+ Foxp3+ Tregs mainly by microbial metabolites butyrate. In addition, Arg1 and STAT6 proteins were also significantly increased after MF treatment. CONCLUSIONS: These results shed light on the contribution of MF treatment to the CD mouse model and suggest that MF has potential as a therapeutic strategy for enhancing efficacy in inducing remission in patients with active CD.

Gαs signaling controls intramembranous ossification during cranial bone development by regulating both Hedgehog and Wnt/ß-catenin signaling.

How osteoblast cells are induced is a central question for understanding skeletal formation. Abnormal osteoblast differentiation leads to a broad range of devastating craniofacial diseases. Here we have investigated intramembranous ossification during cranial bone development in mouse models of skeletal genetic diseases that exhibit craniofacial bone defects. The GNAS gene encodes Gαs that transduces GPCR signaling. GNAS activation or loss-of-function mutations in humans cause fibrous dysplasia (FD) or progressive osseous heteroplasia (POH) that shows craniofacial hyperostosis or craniosynostosis, respectively. We find here that, while Hh ligand-dependent Hh signaling is essential for endochondral ossification, it is dispensable for intramembranous ossification, where Gαs regulates Hh signaling in a ligand-independent manner. We further show that Gαs controls intramembranous ossification by regulating both Hh and Wnt/ß-catenin signaling. In addition, Gαs activation in the developing cranial bone leads to reduced ossification but increased cartilage presence due to reduced cartilage dissolution, not cell fate switch. Small molecule inhibitors of Hh and Wnt signaling can effectively ameliorate cranial bone phenotypes in mice caused by loss or gain of Gnas function mutations, respectively. Our work shows that studies of genetic diseases provide invaluable insights in both pathological bone defects and normal bone development, understanding both leads to better diagnosis and therapeutic treatment of bone diseases.

Laparoscopic Surgery Within an Enhanced Recovery after Surgery (ERAS) Protocol Reduced Postoperative Ileus by Increasing Postoperative Treg Levels in Patients with Right-Side Colon Carcinoma.

BACKGROUND The aim of this study was to determine the effects of laparoscopic surgery within an ERAS program on outcomes and immunological function in patients with a carcinoma in the right colon. MATERIAL AND METHODS Patient data were acquired from a prospectively maintained database, and 176 patients diagnosed with right colon carcinoma with surgery were selected from the database. These patients were divided into a laparoscopic group (Lap group, n=86) and an open operation group (Open group, n=90). All patients received treatment according to a standardized ERAS protocol. We collected data on CRP levels, CD4+/CD8+ ratios, and Treg values in peripheral blood, baseline and surgical characteristics, postoperative complications, and postoperative ileus (POI). RESULTS Circulating CD4+/CD8+ ratios and Treg values were decreased and CRP levels were increased in both groups after the operation. However, the values in the Lap group patients recovered much more quickly than those of patients in the Open group (P<0.05). Patients undergoing laparoscopic surgery had significantly less preoperative bleeding (P<0.01), reduced ratio of overall POI (mainly early ileus), and shorter postoperative hospital stay (P=0.03). Multivariate logistic regression analysis showed that POD1 Treg value was an independent predicator for postoperative ileus in patients with right colon carcinoma resection. CONCLUSIONS In patients with a carcinoma in the right colon, laparoscopic surgery within an ERAS protocol leads to better immunity preservation after surgery, and POD1 Treg value may be an independent predicator for postoperative ileus, which could, at least in part, explain the shorter hospital stay after surgery.

Prognostic value of SOX2, Cyclin D1, P53, and ki-67 in patients with esophageal squamous cell carcinoma.

In this study, we evaluated SOX2, Cyclin D1, p53, and ki-67 expression immunohistochemically in 117 samples of surgically resected esophageal squamous cell carcinoma (ESCC) and matched normal tumor adjacent tissues and correlated the expression with clinicopathological finding and patient survival. Lymph node metastasis was observed in 36.8% of patients, and organ metastasis was observed in 17.9%. We detected high expression of SOX2, Cyclin D1, p53, and ki-67 in 46.1%, 70.1%, 54.7%, and 32.5% of ESCC tissues, respectively. SOX2 is localized in the tumor cell nuclei, and its expression was significantly associated with N stage (p=0.034) and differentiation (p=0.003) and ki-67 expression (p=0.001), whereas increased Cyclin D1 expression was correlated with high p53 (p=0.015). With regard to survival, we found that ESCC patients with high SOX2 expression had significantly better survival time than those with low SOX2 expression (p=0.021). A multivariate Cox analysis revealed that therapy and high p53 expression and venous invasion were independent predictors of unfavorable prognosis in overall survival (p=0.039, p=0.004, and p=0.023, respectively). Furthermore, higher T stage, clinical stage (pTNM), venous invasion, and high p53 expression were independent predictors of a worse progression-free survival. Notably, co-overexpression of p53 and Cyclin D1 was significantly correlated with poor overall survival and progression-free survival (p=0.029 and p=0.0227, respectively). Therefore, SOX2 might be considered as a potential prognostic indicator and a potential target for therapeutic targets in ESCC. p53 staining and combined p53 and Cyclin D1 expression had significantly unfavorable prognostic value for patients with ESCC. These findings provide more insight into ESCC; thus, further investigations into molecular mechanisms of drug resistance are essential.