DNA-functionalized MOF fluorescent probes for the enzyme-free and pretreatment-free detection of MicroRNA in serum.

MicroRNA (miRNA) is a promising biomarker that plays an important role in various biomedical applications, especially in cancer diagnosis. However, the current miRNA detection technology has inherent limitations such as complex operation, expensive testing cost and excessive detection time. In this study, a dual signal amplification biosensor based on DNA-functionalized metal-organic frameworks (MOFs) fluorescent probes, MFPBiosensor, was established for the enzyme-free and pretreatment-free detection of the colon cancer (CC) marker miR-23a. DNA-functionalized MOFs NH2-MIL-53(Al) (DNA@MOFs) were synthesized as fluorescent probes with specific recognition functions. A single DNA@MOF carries a large number of fluorescent ligands 2-aminoterephthalic acid (NH2-H2BDC), which can generate strong fluorescence signals after alkaline hydrolysis. Combined with catalyzed hairpin assembly (CHA), an efficient isothermal amplification technique, the dual signal enhancement strategy reduced matrix interference and sensitized the signal response. The established MFPBiosensor successfully detected extremely low levels of miRNA in complex biological samples with acceptable sensitivity and specificity. With a single detection cost of $0.583 and a test time of 50 min, the excellent inexpensive and rapid advantage of the MFPBiosensor is highlighted. More importantly, the subtle design enables the MFPBiosensor to achieve convenient batch detection, where miRNA in serum can be directly detected without any pretreatment process or enzyme. In conclusion, MFPBiosensor is a promising biosensor with substantial potential for commercial miRNA detection and clinical diagnostic applications of CC.

Multiregion WES of metastatic pancreatic neuroendocrine tumors revealed heterogeneity in genomic alterations, immune microenvironment and evolutionary patterns.

Pancreatic neuroendocrine tumors (PanNETs), though uncommon, have a high likelihood of spreading to other body parts. Previously, the genetic diversity and evolutionary patterns in metastatic PanNETs were not well understood. To investigate this, we performed multiregion sampling whole-exome sequencing (MRS-WES) on samples from 10 patients who had not received prior treatment for metastatic PanNETs. This included 29 primary tumor samples, 31 lymph node metastases, and 15 liver metastases. We used the MSK-MET dataset for survival analysis and validation of our findings. Our research indicates that mutations in the MEN1/DAXX genes might trigger the early stages of PanNET development. We categorized the patients based on the presence (MEN1/DAXXmut, n = 7) or absence (MEN1/DAXXwild, n = 3) of these mutations. Notable differences were observed between the two groups in terms of genetic alterations and clinically relevant mutations, confirmed using the MSK-MET dataset. Notably, patients with mutations in MEN1/DAXX/ATRX genes had a significantly longer median overall survival compared to those without these mutations (median not reached vs. 43.63 months, p = 0.047). Multiplex immunohistochemistry (mIHC) analysis showed a more prominent immunosuppressive environment in metastatic tumors, especially in patients with MEN1/DAXX mutations. These findings imply that MEN1/DAXX mutations lead PanNETs through a unique evolutionary path. The disease's progression pattern indicates that PanNETs can spread early, even before clinical detection, highlighting the importance of identifying biomarkers related to metastasis to guide personalized treatment strategies.

Short-term and Long-term Outcomes of Robotic Enucleation of Tumors Located in the Pancreatic Head and Uncinate Process.

OBJECTIVE: To assess short-term and long-term outcomes following robotic enucleation (REn) of tumors in the proximal pancreas. BACKGROUND: Despite the advantages of preserving function via pancreatic enucleation, controversies persist, since this can be associated with severe complications, such as clinically relevant postoperative pancreatic fistula, especially when performed near the main pancreatic duct. The safety and efficacy of REn in this context remain largely unknown. METHODS: A retrospective analysis was performed of all patients who underwent REn for benign and low-grade malignant neoplasms in the pancreatic head and uncinate process between January 2005 and December 2021. Clinicopathologic, perioperative, and long-term outcomes were compared with a similar open enucleation (OEn) group. RESULTS: Of 146 patients, 92 underwent REn with a zero conversion-to-open rate. REn was superior to OEn in terms of shorter operative time (90.0 minutes vs 120.0 minutes, P<0.001), decreased blood loss (20.0 mL vs 100.0 min, P=0.001), and lower clinically relevant postoperative pancreatic fistula rate (43.5% vs 61.1%, P=0.040). Bile leakage rate, major morbidity, 90-day mortality, and length of hospital stay were comparable between groups. No post-REn grade C POPF or grade IV/V complication was identified. Subgroup analyses for uncinate process tumors and proximity to the main pancreatic duct did not demonstrate inferior postoperative outcomes. In a median follow-up period of 50 months, REn outcomes were comparable to OEn regarding recurrence rate and pancreatic endocrine or exocrine function. CONCLUSIONS: REn for pancreatic head and uncinate process tumors improved clinically relevant outcomes without increased major complications compared to OEn, while demonstrating comparable long-term oncological and functional outcomes.

Berberine protects mice against type 2 diabetes by promoting PPARγ-FGF21-GLUT2-regulated insulin sensitivity and glucose/lipid homeostasis.

Type 2 diabetes (T2D) is a chronic, burdensome disease that is characterized by disordered insulin sensitivity and disturbed glucose/lipid homeostasis. Berberine (BBR) has multiple therapeutic actions on T2D, including regulation of glucose and lipid metabolism, improvement of insulin sensitivity and energy expenditure. Recently, the function of BBR on fibroblast growth factor 21 (FGF21) has been identified. However, if BBR ameliorates T2D through FGF21, the underlying mechanisms remain unknown. Herein, we used T2D wild type (WT) and FGF21 global knockout (FKO) mice [mouse T2D model: established by high-fat diet (HFD) feeding plus streptozotocin (STZ) injection], and hepatocyte-specific peroxisome proliferator activated receptor γ (PPARγ) deficient (PPARγHepKO) mice, and cultured human liver carcinoma cells line, HepG2 cells, to characterize the role of BBR in glucose/lipid metabolism and insulin sensitivity. We found that BBR activated FGF21 expression by up-regulating PPARγ expression at the cellular level. Meanwhile, BBR ameliorated glucosamine hydrochloride (Glcn)-induced insulin resistance and increased glucose transporter 2 (GLUT2) expression in a PPARγ/FGF21-dependent manner. In T2D mice, BBR up-regulated the expression of PPARγ, FGF21 and GLUT2 in the liver, and GLUT2 in the pancreas. BBR also reversed T2D-induced insulin resistance, liver lipid accumulation, and damage in liver and pancreas. However, FGF21 deficiency diminished these effects of BBR on diabetic mice. Altogether, our study demonstrates that the therapeutic effects of BBR on T2D were partly accomplished by activating PPARγ-FGF21-GLUT2 signaling pathway. The discovery of this new pathway provides a deeper understanding of the mechanism of BBR for T2D treatment.

High-resolution genome mapping and functional dissection of chlorogenic acid production in Lonicera maackii.

Amur honeysuckle (Lonicera maackii) is a widely used medicinal plant of the Caprifoliaceae family that produces chlorogenic acid. Research on this plant mainly focuses on its ornamental value and medicinal compounds, but a reference genome sequence and molecular resources for accelerated breeding are currently lacking. Herein, nanopore sequencing and high-throughput chromosome conformation capture (Hi-C) allowed a chromosome-level genome assembly of L. maackii (2n = 18). A global view of the gene regulatory network involved in the biosynthesis of chlorogenic acid and the dynamics of fruit coloration in L. maackii was established through metabolite profiling and transcriptome analyses. Moreover, we identified the genes encoding hydroxycinnamoyl-CoA quinate transferase (LmHQT) and hydroxycinnamoyl-CoA shikimic/quinate transferase (LmHCT), which localized to the cytosol and nucleus. Heterologous overexpression of these genes in Nicotiana benthamiana leaves resulted in elevated chlorogenic acid contents. Importantly, HPLC analyses revealed that LmHCT and LmHQTs recombinant proteins modulate the accumulation of chlorogenic acid (CGA) using quinic acid and caffeoyl CoA as substrates, highlighting the importance of LmHQT and LmHCT in CGA biosynthesis. These results confirmed that LmHQTs and LmHCT catalyze the biosynthesis of CGA in vitro. The genomic data presented in this study will offer a valuable resource for the elucidation of CGA biosynthesis and facilitating selective molecular breeding.

Identification of a novel immune checkpoint molecule V-set immunoglobulin domain-containing 4 that leads to impaired immunity infiltration in pancreatic ductal adenocarcinoma.

BACKGROUND: Checkpoint-based immunotherapy has failed to elicit responses in the majority of patients with pancreatic cancer. In our study, we aimed to identify the role of a novel immune checkpoint molecule V-set Ig domain-containing 4 (VSIG4) in pancreatic ductal adenocarcinoma (PDAC). METHODS: Online datasets and tissue microarray (TMA) were utilized to analyze the expression level of VSIG4 and its correlation with clinical parameters in PDAC. CCK8, transwell assay and wound healing assay were applied to explore the function of VSIG4 in vitro. Subcutaneous, orthotopic xenograft and liver metastasis model was established to explore the function of VSIG4 in vivo. TMA analysis and chemotaxis assay were conducted to uncover the effect of VSIG4 on immune infiltration. Histone acetyltransferase (HAT) inhibitors and si-RNA were applied to investigate factors that regulate the expression of VSIG4. RESULTS: Both mRNA and protein levels of VSIG4 were higher in PDAC than normal pancreas in TCGA, GEO, HPA datasets and our TMA. VSIG4 showed positive correlations with tumor size, T classification and liver metastasis. Patients with higher VSIG4 expression were related to poorer prognosis. VSIG4 knockdown impaired the proliferation and migration ability of pancreatic cancer cells both in vitro and in vivo. Bioinformatics study showed positive correlation between VSIG4 and infiltration of neutrophil and tumor-associated macrophages (TAMs) in PDAC, and it inhibited the secretion of cytokines. According to our TMA panel, high expression of VSIG4 was correlated with fewer infiltration of CD8+ T cells. Chemotaxis assay also showed knockdown of VSIG4 increased the recruitment of total T cells and CD8+ T cells. HAT inhibitors and knockdown of STAT1 led to decreased expression of VSIG4. CONCLUSIONS: Our data indicate that VSIG4 contributes to cell proliferation, migration and resistance to immune attack, thus identified as a promising target for PDAC treatment with good prognostic value.

CircRNA circ_POSTN promotes the malignancy of glioma by regulating the miR-433-3p/SPARC axis.

Glioma is a common tumor in the brain. CircRNA hsa_circ_0030018, also termed as hsa_circPOSTN_001 (circ_POSTN), is reported to exert a promoting influence on the development of glioma. Our study intends to deeply explore its regulation mechanism of circ_POSTN. Expression of circ_POSTN, microRNA-433-3p (miR-433-3p) and Secreted protein acidic and rich in cysteine (SPARC) was detected by qRT-PCR or western blot assay. The function of circ_POSTN was analyzed by loss-of-function experiments. The targeting relationship between miR-433-3p and circ_POSTN or SPARC was predicted by bioinformatics analysis and validated by dual-luciferase reporter assay. Xenograft modeling was employed to validate the function of circ_POSTN in glioma in vivo. circ_POSTN and SPARC were upregulated while miR-433-3p was downregulated in glioma tissues and cells. Both circ_POSTN and SPARC knockdown inhibited clonogenicity, migration, and promoted apoptosis of glioma cells. Circ_POSTN sponged miR-433-3p to regulate SPARC expression. Gain of SPARC largely attenuated circ_POSTN knockdown or miR-433-3p overexpression-mediated effects on glioma cell clonogenicity, migration, and apoptosis. Furthermore, silencing of circ_POSTN decreased xenograft tumor growth in vivo. Inhibition of circ_POSTN repressed glioma development, at least in part, via regulating the miR-433-3p/SPARC axis, providing evidence for circ_POSTN as a potential therapeutic target for glioma.

The Impact of Additional Para-aortic Dissection During Pancreaticoduodenectomy for Resectable Pancreatic Cancer.

BACKGROUND: The short-term outcome and long-term survival of pancreaticoduodenectomy with additional para-aortic dissection (PAD) for patients with resectable pancreatic cancer remain obscure. PATIENTS AND METHODS: Consecutive patients who underwent radical pancreaticoduodenectomy for resectable pancreatic cancer in a single high-volume center during a 7-year period were included retrospectively. Both short- and long-term effects of PAD were compared between the PAD group and the no PAD group. Then, the PAD group was divided into the non-metastatic para-aortic lymph node (PALN-) group and the metastatic PALN (PALN+) group to further analyze the prognosis of PALN+. RESULTS: Of the 909 included patients, 280 (30.8%) underwent PAD during pancreaticoduodenectomy. The PAD group had a higher rate of intra-abdominal infection compared with the no PAD group (28.6% vs. 20.7%, P = 0.009) but no differences were found in the incidence of other complications. The overall survival (OS) and recurrence-free survival (RFS) were also comparable between the two groups. Subgroup analysis showed that patients with PALN+ had a worse OS than patients in the PALN- group (median of 14 vs. 20 months, P = 0.048). Multivariate Cox regression analysis further revealed that PALN+ was an independent adverse predictor of OS (hazard ratio: 1.70, P = 0.007). CONCLUSIONS: This study suggests that the addition of PAD during pancreaticoduodenectomy does not improve the prognosis of patients with resectable pancreatic cancer and may lead to an increased risk of infection. However, the accurate preoperative assessment and appropriate treatment strategy for patients with PALN+ need further investigation due to the poor prognosis.

Differential gene expression and potential regulatory network of fatty acid biosynthesis during fruit and leaf development in yellowhorn (Xanthoceras sorbifolium), an oil-producing tree with significant deployment values.

Xanthoceras sorbifolium (yellowhorn) is a woody oil plant with super stress resistance and excellent oil characteristics. The yellowhorn oil can be used as biofuel and edible oil with high nutritional and medicinal value. However, genetic studies on yellowhorn are just in the beginning, and fundamental biological questions regarding its very long-chain fatty acid (VLCFA) biosynthesis pathway remain largely unknown. In this study, we reconstructed the VLCFA biosynthesis pathway and annotated 137 genes encoding relevant enzymes. We identified four oleosin genes that package triacylglycerols (TAGs) and are specifically expressed in fruits, likely playing key roles in yellowhorn oil production. Especially, by examining time-ordered gene co-expression network (TO-GCN) constructed from fruit and leaf developments, we identified key enzymatic genes and potential regulatory transcription factors involved in VLCFA synthesis. In fruits, we further inferred a hierarchical regulatory network with MYB-related (XS03G0296800) and B3 (XS02G0057600) transcription factors as top-tier regulators, providing clues into factors controlling carbon flux into fatty acids. Our results offer new insights into key genes and transcriptional regulators governing fatty acid production in yellowhorn, laying the foundation for efforts to optimize oil content and fatty acid composition. Moreover, the gene expression patterns and putative regulatory relationships identified here will inform metabolic engineering and molecular breeding approaches tailored to meet biofuel and bioproduct demands.

Potential allopolyploid origin of Ericales revealed with gene-tree reconciliation.

Few incidents of ancient allopolyploidization (polyploidization by hybridization or merging diverged genomes) were previously revealed, although there is significant evidence for the accumulation of whole genome duplications (WGD) in plants. Here, we focused on Ericales, one of the largest and most diverse angiosperm orders with significant ornamental and economic value. Through integrating 24 high-quality whole genome data selected from ~ 200 Superasterids genomes/species and an algorithm of topology-based gene-tree reconciliation, we explored the evolutionary history of in Ericales with ancient complex. We unraveled the allopolyploid origin of Ericales and detected extensive lineage-specific gene loss following the polyploidization. Our study provided a new hypothesis regarding the origin of Ericales and revealed an instructive perspective of gene loss as a pervasive source of genetic variation and adaptive phenotypic diversity in Ericales.

The LINC00623/NAT10 signaling axis promotes pancreatic cancer progression by remodeling ac4C modification of mRNA.

BACKGROUND: Although a substantial increase in the survival of patients with other cancers has been observed in recent decades, pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest diseases. No effective screening approach exists. METHODS: Differential exosomal long noncoding RNAs (lncRNAs) isolated from the serum of patients with PDAC and healthy individuals were profiled to screen for potential markers in liquid biopsies. The functions of LINC00623 in PDAC cell proliferation, migration and invasion were confirmed through in vivo and in vitro assays. RNA pulldown, RNA immunoprecipitation (RIP) and coimmunoprecipitation (Co-IP) assays and rescue experiments were performed to explore the molecular mechanisms of the LINC00623/NAT10 signaling axis in PDAC progression. RESULTS: A novel lncRNA, LINC00623, was identified, and its diagnostic value was confirmed, as it could discriminate patients with PDAC from patients with benign pancreatic neoplasms and healthy individuals. Moreover, LINC00623 was shown to promote the tumorigenicity and migratory capacity of PDAC cells in vitro and in vivo. Mechanistically, LINC00623 bound to N-acetyltransferase 10 (NAT10) and blocked its ubiquitination-dependent degradation by recruiting the deubiquitinase USP39. As a key regulator of N4-acetylcytidine (ac4C) modification of mRNA, NAT10 was demonstrated to maintain the stability of oncogenic mRNAs and promote their translation efficiency through ac4C modification. CONCLUSIONS: Our data revealed the role of LINC00623/NAT10 signaling axis in PDAC progression, showing that it is a potential biomarker and therapeutic target for PDAC.

Astaxanthin Alleviates Ochratoxin A-Induced Cecum Injury and Inflammation in Mice by Regulating the Diversity of Cecal Microbiota and TLR4/MyD88/NF-κB Signaling Pathway.

Ochratoxin A (OTA) is a common environmental pollutant found in a variety of foods and grains, and excessive OTA consumption causes serious global health effects on animals and humans. Astaxanthin (AST) is a natural carotenoid that has anti-inflammatory, antiapoptotic, immunomodulatory, antitumor, antidiabetes, and other biological activities. The present study is aimed at investigating the effects of AST on OTA-induced cecum injury and its mechanism of action. Eighty C57 mice were randomly divided into four groups, including the control group, OTA group (5 mg/kg body weight), AST group (100 mg/kg body weight), and AST intervention group (100 mg/kg body weight AST+5 mg/kg body weight OTA). It was found that AST decreased the endotoxin content, effectively prevented the shortening of mouse cecum villi, and increased the expression levels of tight junction (TJ) proteins, consisting of occludin, claudin-1, and zonula occludens-1 (ZO-1). AST increased the number of goblet cells, the contents of mucin-2 (MUC2), and defensins (Defa5 and ß-pD2) significantly, while the expression of mucin-1 (MUC1) decreased significantly. The 16S rRNA sequencing showed that AST affected the richness and diversity of cecum flora, decreased the proportion of lactobacillus, and also decreased the contents of short-chain fatty acids (SCFAs) (acetate and butyrate). In addition, AST significantly decreased the expression of TLR4, MyD88, and p-p65, while increasing the expression of p65. Meanwhile, the expression of inflammatory factors including TNF-α and INF-γ decreased, while the expression of IL-10 increased. In conclusion, AST reduced OTA-induced cecum injury by regulating the cecum barrier function and TLR4/MyD88/NF-κB signaling pathway.

Does Pre-operative Biliary Drainage Influence Long-Term Survival in Patients With Obstructive Jaundice With Resectable Pancreatic Head Cancer?

Objectives: Whether pre-operative biliary drainage (PBD) affects long-term survival of patients with obstructive jaundice with pancreatic ductal adenocarcinoma (PDAC) who underwent pancreaticoduodenectomy is still controversial. Most of the previous research did not include the important total serum bilirubin (TB) level before intervention as well as before surgery. The aim of this study is to evaluate the impact of PBD on long-term survival after considering the TB level. Methods: Data were collected retrospectively from patients with obstructive jaundice who underwent resection of pancreatic head cancer in a high-volume center. X-Tile software and Kaplan-Meier survival analysis were applied to determine the optimal cut-off levels for TB and age based on the minimal probability (P)-value and the largest χ2-value. Multivariate Cox regression analyses were performed after univariate analysis to assess independent prognostic factors for overall survival (OS) and disease-free survival (DFS). Results: Of 426 patients with obstructive jaundice who underwent pancreaticoduodenectomy for resectable pancreatic head cancer during a 7 year period, 242 (56.8%) received PBD and 184 (43.2%) underwent surgery directly. The OS of patients who received PBD was significantly worse than that of patients who did not receive PBD by univariate analysis (median of 16.6 vs. 22.2 months, P = 0.048). After including liver function parameters in the multivariate Cox regression, we found that the use of PBD was not associated with OS or DFS, while TB before intervention >150 µmol/L was an independent adverse prognostic factor for both OS [hazard ratio (HR), 1.42; 95% CI, 1.05-1.91] and DFS (HR, 1.38; 95% CI, 1.08-1.77). Conclusions: In patients with obstructive jaundice with resectable pancreatic head cancer, undergoing PBD before pancreaticoduodenectomy did not impair or benefit survival rates compared with surgery alone. However, TB before intervention >150 µmol/L predicted an unfavorable prognosis, irrespective of the PBD procedure.

Preoperative biliary drainage of severely obstructive jaundiced patients decreases overall postoperative complications after pancreaticoduodenectomy: A retrospective and propensity score-matched analysis.

OBJECTIVES: The influence of preoperative biliary drainage (PBD) for obstructive jaundiced patients before pancreaticoduodenectomy is debated in the past decades. The aim of this study is to assess the impact of preoperative biliary drainage on intraoperative and postoperative outcomes in patients with severely obstructive jaundice. METHODS: Data were collected retrospectively from severely obstructive jaundiced patients with serum total bilirubin level exceeding 250 µmol/L and undergoing pancreaticoduodenectomy from January 2012 to December 2017. The univariate and multivariate analyses were performed to assess independent risk factors for overall postoperative complications. A propensity score-matched (PSM) analysis was performed to adjust baseline characteristics between PBD and direct surgery (DS) groups. After PSM, intraoperative data and postoperative complications were compared between the two groups. RESULTS: A total of 200 patients were included. The rate of overall postoperative complication occurred in 119 (59.5%) patients, with prealbumin <150 mg/L (OR = 3.03; 95%CI = [1.63-5.62]; p < 0.001), ASA (American Society of Anesthesiology score) classification II-III (OR = 2.27; 95%CI = [1.21-4.27]; p = 0.011), and direct surgery (OR = 3.88; 95%CI = [1.67-8.99]; p = 0.002) identified as independent risk factors in multivariate analysis. After PSM, there was similar operative time and intraoperative transfusion between PBD and DS group. However, DS group had a higher incidence of overall postoperative complication (p = 0.005), grades B and C of post-pancreatectomy hemorrhage (PPH) (p = 0.032), and grades B and C of postoperative pancreatic fistula (POPF) (p = 0.045) compared to PBD group. CONCLUSIONS: In this retrospective study, in order to reduce overall postoperative complications, PBD should be performed routinely for those patients with serum total bilirubin level exceeding 250 µmol/L and undergoing pancreaticoduodenectomy.

NF-κB signaling and integrin-ß1 inhibition attenuates osteosarcoma metastasis via increased cell apoptosis.

Osteosarcoma is a common primary bone malignancy, and distant metastasis limited the cure estimate during last decades. Detailed investigation of osteosarcoma metastasis is valuable for improving therapeutic strategy. Our study indicated increased integrin-ß1 expression and NF-kB signaling activation in metastatic osteosarcoma tissues. Gain-of-function assays showed that integrin-ß1 knockdown significantly inhibited osteosarcoma growth and metastasis, whereas exogenous reintroducing of integrin-ß1 restored cell proliferation and metastasis in vitro and in vivo. NF-κB signaling directly modulated integrin-ß1 expression, which is an effective target for the treatment of osteosarcoma. Mechanically, integrin-ß1 blockage with AIIB2 antibody increased osteosarcoma cell apoptosis. Immunohistochemistry staining of integrin-ß1 revealed that elevated integrin-ß1 expression was correlated with poor prognosis of osteosarcoma patients and acted as an independent detrimental factor for osteosarcoma. Our data showed that integrin-ß1 and NF-κB signaling are promising therapeutic targets to improve the clinical outcome of osteosarcoma patients. The examination of integrin-ß1 expression will also identify patients with high risk of disease progression.

PIK3CA mutations confer resistance to first-line chemotherapy in colorectal cancer.

Chemotherapy represents an important treatment option for colorectal cancer (CRC), but only half of the patients benefit from these regimens. We explored the potential predicting value and mechanism of PIK3CA mutation in CRC chemotherapy. CRC specimens from 440 patients were retrospectively collected and examined with a fluorescence PCR-based method. The correlation of first-line chemotherapy response and PIK3CA mutation was evaluated according to follow-up and medical records. The underlying mechanism of PIK3CA mutation in chemotherapy resistance was assessed with CRC tumors and primary cells. The mutation frequency of the PIK3CA gene in CRC patients was 9.55%, which was correlated with late TNM staging and lower histological grade. The CRC patients with PIK3A mutation showed worse response to first-line chemotherapy than those without PIK3CA mutation. PIK3A mutation tumor cells showed poor sensitivity to first-line chemotherapy in vitro and in vivo. PIK3CA mutation induced PI3K/Akt signaling activation to increase LGR5+ CRC stem cells survival and proliferation, from which lead to chemotherapy resistance. Furthermore, PIK3CA mutation/LGR5+ expression was an independent detrimental factor for CRC patients. Our findings indicated that PIK3CA mutation induced PI3K/Akt activation contributed to CRC stem cells survival and proliferation, from which cells further resistance to chemotherapy. PIK3CA mutation/LGR5+ expression was a potential biomarker for monitoring chemotherapy resistance in CRC.

Protective effects of tiotropium alone or combined with budesonide against cadmium inhalation induced acute neutrophilic pulmonary inflammation in rats.

As a potent bronchodilator, the anti-inflammatory effects of tiotropium and its interaction with budesonide against cadmium-induced acute pulmonary inflammation were investigated. Compared to values obtained in rats exposed to cadmium, cytological analysis indicated a significant decrease of total cell and neutrophil counts and protein concentration in bronchoalveolar lavage fluid (BALF) in rats pretreated with tiotropium (70µg/15ml or 350µg/15ml). Zymographic tests showed a decrease of MMP-2 activity in BALF in rats pretreated only with high concentration of tiotropium. Histological examination revealed a significant decrease of the severity and extent of inflammatory lung injuries in rats pretreated with both tested concentrations of tiotropium. Though tiotropium (70µg/15ml) or budesonide (250µg/15ml) could not reduce cadmium-induced bronchial hyper-responsiveness, their combination significantly decreased bronchial contractile response to methacholine. These two drugs separately decreased the neutrophil number and protein concentration in BALF but no significant interaction was observed when both drugs were combined. Although no inhibitory effects on MMP-2 and MMP-9 was observed in rats pretreated with budesonide alone, the combination with the ineffective dose of tiotropium induced a significant reduction on these parameters. The inhibitory effect of tiotropium on lung injuries was not influenced by budesonide which alone induced a limited action on the severity and extent of inflammatory sites. Our findings show that tiotropium exerts anti-inflammatory effects on cadmium-induced acute neutrophilic pulmonary inflammation. The combination of tiotropium with budesonide inhibits cadmium-induced inflammatory injuries with a synergistic interaction on MMP-2 and MMP-9 activity and airway hyper-responsiveness.