Search

Background: Porcine epidemic diarrhea (PED) is a contagious intestinal disease caused by porcine epidemic diarrhea virus (PEDV) characterized by vomiting, diarrhea, anorexia, and dehydration, which has caused huge economic losses around the world. However, it is very hard to find completely valid approaches to control the transmission of PEDV. At present, vaccine immunity remains the most effective method. To better control the spread of PED and evaluate the validity of different immunization strategies, 240 PED outbreak cases from 577 swine breeding farms were collected and analyzed. The objective of the present study was to analyze the epidemic regularity of PEDV and evaluate two kinds of different immunization strategies for controlling PED., Results: The results showed that the main reasons which led to the outbreak of PED were the movement of pig herds between different pig farms (41.7%) and delaying piglets from the normal production flow (15.8%). The prevalence of PEDV in the hot season (May to October) was obviously higher than that in the cold season (January to April, November to December). Results of different vaccine immunity cases showed that immunization with the highly virulent live vaccine (NH-TA2020 strain) and the commercial inactivated vaccine could significantly decrease the frequency of swine breeding farms (5.9%), the duration of PED epidemic (1.70 weeks), and the week batches of dead piglets (0.48 weeks weaned piglets), compared with immunization with commercial attenuated vaccines and inactivated vaccine of PED. Meanwhile, immunization with the highly virulent live vaccine and the commercial inactivated vaccine could bring us more cash flows of Y̶275,274 per year than immunization with commercial live attenuated vaccine and inactivated vaccine in one 3000 sow pig farm within one year., Conclusion: Therefore, immunization with highly virulent live vaccine and inactivated vaccine of PED is more effective and economical in the prevention and control of PED in the large-scale swine farming system., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Background: At present, there is no consensus on whether preoperative immunotherapy (PIT) without chemotherapy followed by surgery could benefit patients with advanced gastric cancer (AGC). Here, we report a six-case series study to describe the safety and efficacy of PIT plus gastrectomy in patients with AGC., Methods: This study involved six patients with AGC who received PIT and surgery at our center between January 2019 and July 2021. Demographic characteristics, preoperative gastroscope biopsy pathology, surgical tissue pathology, radicalness of tumor resection, surgical safety, and recovery parameters were reported., Results: Six patients, including four patients with Epstein-Barr virus (EBV)-positive gastric cancer (GC) and two patients with microsatellite instability-high (MSI-H)/expression deficiency of mismatch repair (dMMR) protein GC, were enrolled in this study. Four patients experienced immunotherapy-related adverse events (irAEs), without severe adverse events (SAEs). Five patients underwent R0 resection, and one patient underwent palliative gastrectomy due to liver and hilar lymph node metastasis. Pathologic responses from the surgical tissue were observed in all patients, including two pathological complete response (pCR). No operative complications or postoperative deaths occurred. Three patients (50%) experienced mild or moderate postoperative complications without severe postoperative complications. All six patients eventually recovered and were discharged., Conclusion: This study indicated that PIT was effective and tolerant in some patients with MSI-H/dMMR and/or EBV-positive AGC. PIT followed by gastrectomy might be an alternative treatment option for these selected patients., Competing Interests: Conflicts of interest: The authors declare that they have no conflicts of interest related to the subject matter or materials discussed in this article., (Copyright © 2023, the Chinese Medical Association.)

Introduction: Porcine circovirus type 2 (PCV2) is the primary etiological agent of porcine circovirus diseases (PCVD), which are widespread in most pig herds, causing huge economic losses in the global pig industry. Therefore, it is critical to assess the infection characteristics of PCV2 in different swine herds to develop effective strategies against PCVD., Methods: In this study, routine diagnostic and monitoring protocols were used to collect 12,714 samples from intensive farms in China, and PCV2 was tested for by qPCR to determine positivity rates and viral loads in samples from different herds and materials., Results: PCV2 was found to be prevalent throughout China, and fattening farms had higher positivity rates than breeding farms. The PCV2 positivity rates in breeding farms in Southern China were higher than those in Northern China. Growing-finishing pigs demonstrated the highest positivity rate in the tested samples, while pre-weaning piglets and adult sows had the lowest. Meanwhile, samples with viral loads exceeding 106 copies/mL in growing-finishing pigs had 27.2% positivity, compared to 1.9% and 3.3% in sows and piglets, respectively. The results of the viral loads in the serum samples followed a similar trend., Discussion: The findings reveal that PCV2 circulates in different herds from intensive farms, with positivity increasing from pre-weaning to growing-finishing herds. It is urgent to develop effective strategies to reduce PCV2 positivity in growing-finishing herds and prevent viral circulation among pigs., Competing Interests: XL, MF, LB, XT, ZH, WW, LS, GY, SL, LY, LW, YiW, YoW, and ZY were employed by Shandong New Hope Liuhe Agriculture and Animal Husbandry Technology Co., Ltd., New Hope Liuhe Co., Ltd., and Xiajin New Hope Liuhe Agriculture and Animal Husbandry Co., Ltd. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Fan, Bian, Tian, Hu, Wu, Sun, Yuan, Li, Yue, Wang, Wu, Wang, Yan, Ren and Li.)

Objective: To explore the change and feasibility of surgical techniques of laparoscopic transhiatal (TH)-lower mediastinal lymph node dissection (LMLND) for adenocarcinoma of the esophagogastric junction (AEG) according to Idea, Development, Exploration, Assessment, and Long-term follow-up (IDEAL) 2a standards., Methods: Patients diagnosed with AEG who underwent laparoscopic TH-LMLND were prospectively included from April 14, 2020, to March 26, 2021. Clinical and pathological information as well as surgical outcomes were quantitatively analyzed. Semistructured interviews with the surgeon after each operation were qualitatively analyzed., Results: Thirty-five patients were included. There were no cases of transition to open surgery, but three cases involved combination with transthoracic surgery. In qualitative analysis, 108 items under three main themes were detected: explosion, dissection, and reconstruction. Revised instruction was subsequently designed according to the change in surgical technique and the cognitive process behind it. Three patients had anastomotic leaks postoperatively, with one classified as Clavien-Dindo IIIa., Conclusions: The surgical technique of laparoscopic TH-LMLND is stable and feasible; further IDEAL 2b research is warranted., Competing Interests: Conflicts of Interest: The authors have no conflicts of interest to declare., (Copyright ©2023 Chinese Journal of Cancer Research. All rights reserved.)

Expansion of G4C2 hexanucleotide repeats in the chromosome 9 ORF 72 (C9ORF72) gene is the most common genetic cause of amyotrophic lateral sclerosis (ALS) with frontotemporal dementia (C9-ALS/FTD). Dipeptide repeats generated by unconventional translation, especially the R-containing poly(GR), have been implicated in C9-ALS/FTD pathogenesis. Mutations in other genes, including TAR DNA-binding protein 43 KD (TDP-43), fused in sarcoma (FUS), and valosin-containing protein, have also been linked to ALS/FTD, and upregulation of amyloid precursor protein (APP) is observed at the early stage of ALS and FTD. Fundamental questions remain as to the relationships between these ALS/FTD genes and whether they converge on similar cellular pathways. Here, using biochemical, cell biological, and genetic analyses in Drosophila disease models, patient-derived fibroblasts, and mammalian cell culture, we show that mechanistic target of rapamycin complex 2 (mTORC2)/AKT signaling is activated by APP, TDP-43, and FUS and that mTORC2/AKT and its downstream target valosin-containing protein mediate the effect of APP, TDP-43, and FUS on the quality control of C9-ALS/FTD-associated poly(GR) translation. We also find that poly(GR) expression results in reduction of global translation and that the coexpression of APP, TDP-43, and FUS results in further reduction of global translation, presumably through the GCN2/eIF2α-integrated stress response pathway. Together, our results implicate mTORC2/AKT signaling and GCN2/eIF2α-integrated stress response as common signaling pathways underlying ALS/FTD pathogenesis., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: This study aims to verify the feasibility and efficacy of laparoscopic lower mediastinal lymphadenectomy for Siewert type II/III adenocarcinoma of esophagogastric junction (AEG)., Setting: An exploratory, observational, prospective, cohort study will be carried out under the Idea, Development, Exploration, Assessment and Long-term Follow-up (IDEAL) framework (stage 2b)., Participants: The study will recruit 1,036 patients with cases of locally advanced AEG (Siewert type II/III, clinical stage cT2-4aN0-3M0), and 518 will be assigned to either the laparoscopy group or the open group., Interventions: Patients will receive lower mediastinal lymphadenectomy along with either total or proximal gastrectomy., Primary and Secondary Outcome Measures: The primary endpoint is the number of lower mediastinal lymph nodes retrieved, and the secondary endpoints are the surgical safety and prognosis, including intraoperative and postoperative lower-mediastinal-lymphadenectomy-related morbidity and mortality, rate of rehospitalization, R0 resection rate, 3-year local recurrence rate, and 3-year overall survival., Conclusions: The study will provide data for the guidance and development of surgical treatment strategies for AEG., Trial Registration Number: The study has been registered in ClinicalTrials.gov (No. NCT04443478)., (© Chinese Journal of Cancer Research. All rights reserved.)

Microalgae-based technology is an effective and environmentally friendly method for antibiotics-contaminated wastewater treatment. To assess the tolerance and removal ability of Chlorella sorokiniana to ciprofloxacin (CIP), this study comprehensively revealed the responses of C. sorokiniana to CIP exposure and its degradation processes through physiological and transcriptomic analyses. Although the photosynthetic system was inhibited, the growth of C. sorokiniana was not negatively affected by CIP. Dissolved organic matter was analyzed and indicated that humic-like substances were released to alleviate the stress of CIP. In addition, the maximum removal of CIP was 83.3% under 20 mg L -1 CIP exposure. HPLC-MS/MS and RNA-Seq analyses suggested that CIP could be bioaccumulated and biodegraded by C. sorokiniana through the reactions of hydroxylation, demethylation, ring cleavage, oxidation, dehydrogenation, and decarboxylation with the help of intracellular oxidoreductases, especially cytochrome P450. Collectively, this research shows that C. sorokiniana have a great potential for removing CIP from wastewater., Competing Interests: The authors declare no competing interests., (© 2022 The Author(s).)

Environmental pollution caused by petroleum hydrocarbons is being paid more and more attention worldwide. Surfactants are able to improve the solubility of petroleum hydrocarbons, but their effects on petroleum hydrocarbon degradation in composting systems are still unclear. In this study, the effects on microbial community succession were investigated by adding petroleum hydrocarbons and rhamnolipids during composting of organic wastes. The results showed that the compost and the addition of rhamnolipids could effectively reduce the petroleum hydrocarbon content with an efficiency of 73.52%, compared to 53.81% for the treatment without addition. Network analyses and Structural Equation Model suggested that there were multiple potential petroleum degraders microbes that might be regulated by nitrogen. The findings in this study can also provide an implication for the treatment of petroleum hydrocarbon pollutants from oil-polluted soil, and the technology can be potentially applied on an industrial scale in practice., Competing Interests: The authors declare no competing interests., (© 2022 The Author(s).)

Stalled mRNA translation results in the production of incompletely synthesized proteins that are targeted for degradation by ribosome-associated quality control (RQC). Here we investigated the fate of defective proteins translated from stall-inducing, nonstop mRNA that escape ubiquitylation by the RQC protein LTN1. We found that nonstop protein products accumulated in nucleoli and this localization was driven by polylysine tracts produced by translation of the poly(A) tails of nonstop mRNA. Nucleolar sequestration increased the solubility of invading proteins but disrupted nucleoli, altering their dynamics, morphology, and resistance to stress in cell culture and intact flies. Our work elucidates how stalled translation may affect distal cellular processes and may inform studies on the pathology of diseases caused by failures in RQC and characterized by nucleolar stress.

Purpose: To compare the short-term surgical outcomes of laparoscopic and open total/proximal gastrectomy using transorally inserted anvil (OrVil TM )., Method: Patients diagnosed with gastric cancer and underwent total or proximal gastrectomy using OrVil TM for reconstruction were included. Clinical and pathological characteristics, as well as postoperative outcomes, were analyzed. Propensity score matching was used to balance baseline factors., Results: From April 2012 to April 2020, 199 patients at our center were included. A total of 166 underwent open total or proximal gastrectomy (OTG/OPG), and 33 underwent laparoscopic total or proximal gastrectomy (LTG/LPG). Twenty-seven patients from each group were paired with propensity score matching. The operation time was significantly shorter in the OTG/OPG group after matching. The overall complication rate and the incidence of each complication did not show significant differences between the two groups before and after matching., Conclusion: LTG/LPG and OTG/OPG using OrVil TM for the alimentary tract reconstruction are both feasible and can achieve similar short-term outcomes.

Maintaining the fidelity of nascent peptide chain (NP) synthesis is essential for proteome integrity and cellular health. Ribosome-associated quality control (RQC) serves to resolve stalled translation, during which untemplated Ala/Thr residues are added C terminally to stalled peptide, as shown during C-terminal Ala and Thr addition (CAT-tailing) in yeast. The mechanism and biological effects of CAT-tailing-like activity in metazoans remain unclear. Here we show that CAT-tailing-like modification of poly(GR), a dipeptide repeat derived from amyotrophic lateral sclerosis with frontotemporal dementia (ALS/FTD)-associated GGGGCC (G4C2) repeat expansion in C9ORF72 , contributes to disease. We find that poly(GR) can act as a mitochondria-targeting signal, causing some poly(GR) to be cotranslationally imported into mitochondria. However, poly(GR) translation on mitochondrial surface is frequently stalled, triggering RQC and CAT-tailing-like C-terminal extension (CTE). CTE promotes poly(GR) stabilization, aggregation, and toxicity. Our genetic studies in Drosophila uncovered an important role of the mitochondrial protease YME1L in clearing poly(GR), revealing mitochondria as major sites of poly(GR) metabolism. Moreover, the mitochondria-associated noncanonical Notch signaling pathway impinges on the RQC machinery to restrain poly(GR) accumulation, at least in part through the AKT/VCP axis. The conserved actions of YME1L and noncanonical Notch signaling in animal models and patient cells support their fundamental involvement in ALS/FTD., Competing Interests: The authors declare no competing interest.

Purpose: To investigate the efficacy of percutaneous transhepatic biliary drainage (PTBD) combined with iodine-125 ( 125 I) stranded seeds for the treatment of malignant bile duct obstruction (MBO)., Material and Methods: A retrospective study was performed on 58 consecutive MBO patients. Twenty patients underwent PTBD combined with 125 I stranded seeds (group A). Thirty-eight patients underwent percutaneous trans-hepatic biliary drainage (group B). Total bilirubin, direct bilirubin, and indirect bilirubin levels were compared preoperatively, 1-week, 1-month, and 3-months post-operatively. Carbohydrate antigen 19-9 (CA19-9), cancer antigen 125 (CA125), and carcino-embryonic antigen (CEA) levels were compared at preoperative and 3-month post-operative stages. The time free from biliary obstruction and survival times were compared., Results: The differences in total bilirubin, direct bilirubin, and indirect bilirubin levels between the two groups were not significant preoperatively ( p = 0.857, p = 0.719, and p = 0.870), and 1-week post-operatively ( p = 0.259, p = 0.395, and p = 0.145). However, 1-month ( p = 0.012, p = 0.005, and p = 0.049) and 3-months post-operatively ( p < 0.001, p = 0.001, and p = 0.001), group A was lower than group B. Differences in CA19-9, CA125, and CEA levels between the two groups were not significant preoperatively ( p = 0.229, p = 0.116, and p = 0.273) and 3-months post-operatively ( p = 0.159, p = 0.342, and p = 0.306). The median biliary obstruction free time was 7.0 months for group A and 5.0 months for group B ( p < 0.001). The median survival time was 9.0 months for group A and 6.0 months for group B ( p = 0.001)., Conclusions: PTBD combined with 125 I stranded seeds seem to reduce bilirubin levels and prevents biliary obstruction, promoting survival., Competing Interests: The authors report no conflict of interest., (Copyright © 2020 Termedia.)

Improving soil structure, fertility, and production is of major concern for establishing sustainable agroecosystems. Further research is needed to evaluate whether different methods of straw returning determine the variations of soil aggregation and the microbial community in aggregates in the long term. In this study, we comparatively investigated the effects of long-term fertilization regimes performed over six years, namely, non-fertilization (CK), chemical fertilization (CF), continuous straw return (CS), and continuous straw-derived biochar amendment (CB), on soil aggregation and bacterial communities in rice-wheat rotation systems. The results showed that straw/biochar application increased soil nutrient content and soil aggregate size distribution and stability at both 0-20 cm and 20-40 cm soil depths, compared with those of CF and CK; CB performed better than CS. CB increased bacterial community diversity and richness in 0-20 cm soil, and evenness in 0-40 cm soil (p < 0.05); CS had no significant effect on these aspects. Variations in the relative abundance of Actinobacteria, Chloroflexi, Bacteroidetes, Nitrospirae, Gemmatimonadetes, and Latescibacteria in specific aggregates confirmed the different effects of straw/biochar on bacterial community structure. The partial least squares discrimination analysis and permutation multivariate analysis of variance revealed that fertilization, aggregate size fractions, and soil depth affected the bacterial community, although their effects differed. This study suggests that CB may reduce chemical fertilizer usage and improve the sustainability of rice-wheat cropping systems over the long term, with a better overall outcome than CS.

It is an intractable issue to improve the low-temperature SO 2 -tolerant selective catalytic reduction (SCR) of NO x with NH 3 because deposited sulfates are difficult to decompose below 300 °C. Herein, we established a low-temperature self-prevention mechanism of mesoporous-TiO 2 @Fe 2 O 3 core-shell composites against sulfate deposition using experiments and density functional theory. The mesoporous TiO 2 -shell effectively restrained the deposition of FeSO 4 and NH 4 HSO 4 because of weak SO 2 adsorption and promoted NH 4 HSO 4 decomposition on the mesoporous-TiO 2 . The electron transfer at the Fe 2 O 3 (core)-TiO 2 (shell) interface accelerated the redox cycle, launching the "Fast SCR" reaction, which broadened the low-temperature window. Engineered from the nano- to macro-scale, we achieved one-pot self-installation of mesoporous-TiO 2 @Fe 2 O 3 composites on the self-tailored AlOOH@Al-mesh monoliths. After the thermal treatment, the mesoporous-TiO 2 @Fe 2 O 3 @Al 2 O 3 monolith catalyst delivered a broad window of 220-420 °C with NO conversion above 90% and had superior SO 2 tolerance at 260 °C. The effective heat removal of Al-mesh monolithcatalysts restrained NH 3 oxidation to NO and N 2 O while suppressing the decomposition of NH 4 NO 3 to N 2 O, and this led to much better high-temperature activity and N 2 selectivity. This work supplies a new point for the development of low-temperature SO 2 -tolerant monolithic SCR catalysts with high N 2 selectivity, which is of great significance for both academic interests and practical applications.

Two-dimensional (2D) nanocomposites as lubricant additives have been widely studied, but the synthetic process of the nanocomposites is not always environmentally friendly or economical. In this study, a new 2D nanocomposite, Fe 3 O 4 /h-BN, has been prepared by physical mixing of exfoliated h-BN nanosheets and organically modified Fe 3 O 4 nanoparticles. The nanocomposite displays a unique 2D-layered structure without folds or wrinkles. The Fe 3 O 4 nanoparticles are uniformly dispersed on the h-BN nanosheet surfaces with the help of an elegant self-assembly strategy from van der Waals interactions. For the first time, Fe 3 O 4 /h-BN is studied as a lubricant additive and it exhibits excellent tribological properties. The coefficient of friction (COF) and the wear depth can be respectively reduced by 47% and 80% compared with the base oil. Based on the advantages of a simple and low-cost synthetic process and significant tribological properties, Fe 3 O 4 /h-BN offers great potential for lubrication application., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Hedgehog (Hh) transduces signals by promoting cell surface accumulation and activation of the G-protein-coupled receptor (GPCR)-family protein Smoothened (Smo) in Drosophila , but the molecular mechanism underlying the regulation of Smo trafficking remains poorly understood. Here, we identified the Cul4-DDB1 E3 ubiquitin ligase complex as being essential for Smo ubiquitylation and cell surface clearance. We found that the C-terminal intracellular domain of Smo recruits Cul4-DDB1 through the β subunit of trimeric G protein (Gβ), and that Cul4-DDB1-Gβ promotes the ubiquitylation of both Smo and its binding partner G-protein-coupled-receptor kinase 2 (Gprk2) and induces the internalization and degradation of Smo. Hh dissociates Cul4-DDB1 from Smo by recruiting the catalytic subunit of protein kinase A (PKA) to phosphorylate DDB1, which disrupts its interaction with Gβ. Inactivation of the Cul4-DDB1 complex resulted in elevated Smo cell surface expression, whereas an excessive amount of Cul4-DDB1 blocked Smo accumulation and attenuated Hh pathway activation. Taken together, our study identifies an E3 ubiquitin ligase complex targeting Smo for ubiquitylation and provides new insight into how Hh signaling regulates Smo trafficking and cell surface expression., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Objective: To compare the short-term safety and costs between laparoscopic assisted or totally laparoscopic uncut Roux-en-Y and Billroth II((BII() + Braun reconstruction after radical gastrectomy of distal gastric cancer., Methods: Clinical data from our prospective database of radical gastrectomy were systematically analyzed. The patients who underwent laparoscopic gastrectomy with uncut Roux-en-Y or BII(+ Braun reconstruction between March 1st, 2015 and June 30th, 2017 were screened out for further analysis. Both the reconstructions were completed by linear staplers. Uncut Roux-en-Y reconstruction was performed with a 45 mm no-knife linear stapler (ATS45NK) on the afferent loop below the gastrojejunostomy. Continuous variables were compared using independent samples t test or Mann-Whitney U. The frequencies of categorical variables were compared using Chi-squared or Fisher exact test., Results: Eighty-one patients were in uncut Roux-en-Y group and 58 patients were in BII(+Braun group. There were no significant differences between uncut Roux-en-Y group and BII(+Braun group in median age (56.0 years vs. 56.5 years, P=0.757), gender (male/female, 52/29 vs. 46/12, P=0.054), history of abdominal surgery (yes/no, 10/71 vs. 4/54, P=0.293), neoadjuvant chemotherapy (yes/no, 21/60 vs. 11/47, P=0.336), BMI (thin/normal/overweight/obesity, 2/49/26/3 vs. 3/39/14/2, P=0.591), NRS 2002 score (1/2/3/4, 58/15/5/3 vs. 47/5/3/3, P=0.403), pathological stage (0/I(/II(/III(, 3/41/20/17 vs. 1/28/13/16, P=0.755), median tumor diameter in long axis (2.5 cm vs. 3.0 cm, P=0.278), median tumor diameter in short axis (2.0 cm vs. 2.0 cm, P=0.126) and some other clinical and pathological characteristics. There were no significant differences between uncut Roux-en-Y group and BII(+Braun group in morbidity of postoperative complication more severe than grade I([12.3% (10/81) vs. 17.2% (10/58), P=0.417], morbidity of anastomotic complication [1.2%(1/81) vs. 0, P=1.000] or hospitalization costs [(94000±14000) yuan vs.(95000±16000) yuan, P=0.895]. The median first time to liquid diet (57.1 hours vs. 70.8 hours, P=0.017) and median postoperative hospital stay (9 days vs. 11 days, P=0.003) of the patients in uncut Roux-en-Y group were shorter than those in BII(+Braun group., Conclusion: Laparoscopic assisted or totally laparoscopic uncut Roux-en-Y reconstruction after radical gastrectomy of distal gastric cancer is safe and feasible with better recovery than BII(+Braun reconstruction.

Purpose: To compare the therapeutic effects of 125 I versus non- 125 I combined with transcatheter arterial chemoembolization (TACE) for the treatment of unresectable hepatocellular carcinoma (HCC) with obstructive jaundice., Methods: A retrospective analysis was conducted using the records of 54 consecutive patients who were initially diagnosed with HCC with obstructive jaundice between May 2009 and July 2016. Twenty-one cases (group A) were treated with percutaneous transhepatic biliary drainage (PTBD) followed by 125 I radioactive seed strip implantation through the PTBD tube. After the total serum bilirubin level was reduced to normal and the liver function recovered to Child-Pugh class A or early B, TACE was conducted. In 33 cases (group B) PTBD was performed in combination with TACE without applying the 125 I radioactive seeds. The duration of biliary patency and survival were analyzed., Results: The technical success rate in both groups was 100%. The median biliary patency time was 6.000 ± 0.315 months (95% CI 5.382-6.618 months) in group A and 4.000 ± 0.572 months (95% CI 2.879-5.121 months) in group B; the two groups were significantly different (P = 0.001). The median survival was 11.000 ± 0.864 months (95% CI 9.306-12.694 months) in group A and 9.000 ± 0.528 months (95% CI 7.965-10.035 months) in group B; the two groups were significantly different (P = 0.022)., Conclusions: The combination of 125I with TACE was more effective than TACE without the radioactive seeds for treating patients with unresectable HCC with obstructive jaundice. Future prospective trials with larger samples will be required to validate these results.

Objective: Diagnostic laparoscopy is recommended for the pretherapeutic staging of gastric cancer to detect any unexpected or unconfirmed intra-abdominal metastasis. The aim of this study was to evaluate the role and indications of diagnostic laparoscopy in the detection of intra-abdominal metastasis., Methods: Standard diagnostic laparoscopy with peritoneal cytology examination was performed prospectively on patients who were clinically diagnosed with primary local advanced gastric cancer (cT≥2M0). We calculated the rate of intra-abdominal metastases identified by diagnostic laparoscopy, and examined the relationship between peritoneal dissemination (P) and cytology results (CY). Split-sample method was applied to find clinical risk factors for intra-abdominal metastasis. Multivariate logistic regression analysis and receiver-operator characteristic (ROC) analysis were performed in training set to find out risk factors of intra-abdominal metastasis, and then validate it in testing set., Results: Out of 249 cM0 patients, 51 (20.5%) patients with intra-abdominal metastasis were identified by diagnostic laparoscopy, including 20 (8.0%) P1CY1, 17 (6.8%) P0CY1 and 14 (5.6%) P1CY0 patients. In the training set, multivariate logistic regression analysis and ROC analysis showed that the depth of tumor invasion on computer tomography (CT) scan ≥21 mm and tumor-occupied ≥2 portions of stomach are predictive factors of metastasis. In the testing set, when diagnostic laparoscopy was performed on patients who had one or two of these risk factors, the sensitivity and positive predictive value for detecting intra-abdominal metastasis were 90.0% and 32.1%, respectively., Conclusions: According to our results, depth of tumor invasion and tumor-occupied portions of stomach are predictive factors of intra-abdominal metastasis.

Background/aims: Calycosin, a phytoestrogenic compound, has recently emerged as a promising antitumor drug. It has been shown that calycosin suppresses growth and induces apoptosis of breast cancer cells. However, the effect of calycosin on migration and invasion of breast cancer cells and the underlying molecular mechanisms have not been elucidated., Methods: Human breast cancer cells MCF-7 and T47D were treated with, or without, different doses (0, 6.25, 12.5, 25, 50, 100 or 150 μM) of calycosin, and the viability of different groups was determined by MTT assay. Next, the inhibitory effect of higher doses (50, 100 or 150 μM) of calycosin on migration and invasion of the two cell lines was determined by wound healing and transwell assay. The relative expression levels of forkhead box P3 (Foxp3), vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) in MCF-7 and T47D cells were determined by quantitative RT-PCR and Western blot., Results: Treatment with lower doses (6.25 or 12.5 μM) promoted proliferation of breast cancer cells, but with higher doses significantly reduced the viability of MCF-7 and T47D cells. Furthermore, higher doses of calycosin were found to inhibit migration and invasion of the two cell lines in a dose-dependent manner. Additionally, treatment with a higher dose of calycosin significantly reduced the expression levels of Foxp3, followed by down-regulation of VEGF and MMP-9 in both MCF-7 and T47D breast cancer cells., Conclusion: Treatment with a higher dose of calycosin tends to reduce migration and invasion capacity of human breast cancer cells, by targeting Foxp3-mediated VEGF and MMP-9 expression., (© 2017 The Author(s). Published by S. Karger AG, Basel.)

Hedgehog (Hh) signaling plays a central role in development and diseases. Hh activates its signal transducer and GPCR-family protein Smoothened (Smo) by inducing Smo phosphorylation, but whether Smo is activated through other post-translational modifications remains unexplored. Here we show that sumoylation acts in parallel with phosphorylation to promote Smo cell-surface expression and Hh signaling. We find that Hh stimulates Smo sumoylation by dissociating it from a desumoylation enzyme Ulp1. Sumoylation of Smo in turn recruits a deubiquitinase UBPY/USP8 to antagonize Smo ubiquitination and degradation, leading to its cell-surface accumulation and elevated Hh pathway activity. We also provide evidence that Shh stimulates sumoylation of mammalian Smo (mSmo) and that sumoylation promotes ciliary localization of mSmo and Shh pathway activity. Our findings reveal a conserved mechanism whereby the SUMO pathway promotes Hh signaling by regulating Smo subcellular localization and shed light on how sumoylation regulates membrane protein trafficking., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: To compare the safety and efficacy of laparoscopic distal gastrectomy (LDG) versus open distal gastrectomy (ODG) in treating locally advanced distal gastric cancer after neoadjuvant chemotherapy (NACT)., Methods: Forty-four patients with locally advanced distal gastric cancer were enrolled. The patients received neoadjuvant chemotherapy before undergoing surgery. Twenty patients were allocated into LDG after NACT group and 24 patients into ODG after NACT group. Radicalness of oncological resection, surgical safety and recovery were measured and compared., Results: All operations were successfully performed without severe postoperative complications. There were no significant differences in blood loss, mean operation time, complications, distal and proximal resection margin, and number of retrieved lymph nodes between LDG and ODG groups, but LDG group had shorter length of incision and the first aerofluxus time., Conclusion: Laparoscopic distal gastrectomy after NACT has comparable results with open distal gastrectomy in safety and efficacy in the short term.

Purpose: Angiogenesis is a promising therapeutic target to inhibit tumor growth. This review summarizes data from clinical trials of anti-angiogenic agents in gastric cancer., Design: A systematic search of PubMed, Embase and conference databases is performed to identify clinical trials with specific anti-angiogenic agents in gastric cancer treatment, Results: The risk of disease progression (37-52%) and death (19-22%) with ramucirumab as second-line treatment decreases in phase III trials in advanced gastric cancer. No significant improvement in overall survival (OS) with the addition of bevacizumab to chemotherapy is shown. Bevacizumab or ramucirumab combined with traditional chemotherapy is associated with higher adverse event rate compared to chemotherapy alone. Except for apatinib, phase II trials of other tyrosine kinase inhibitors (TKIs) may improve overall response rate, but there are no significant improvements in OS and progression-free survival (PFS) when combined with chemotherapy., Conclusion: Phase III trials in advanced gastric cancer have demonstrated improved outcome with ramucirumab as second-line treatment. Most of the other studies on anti-angiogenic agents in gastric cancer have reported improvement in response rate but not in OS compared to chemotherapy alone. Future research is expected in optimizing the anti-angiogenic therapy combined with traditional treatment., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Several studies have evaluated the efficacy of neoadjuvant treatment using oxaliplatin and fluoropyrimidines in advanced gastric cancer (GC). However, preoperative biomarkers predictive of clinical outcome remain lacking. We examined polymorphisms in the MTHFR, DPYD, UMPS, ABCB1, ABCC2, GSTP1, ERCC1, and XRCC1 genes to evaluate their usefulness as pharmacogenetic markers in a cohort of 103 GC patients treated with preoperative chemotherapy. DNA was extracted from peripheral blood cells, and the genotypes were analyzed using a SNaPShotTM assay, polymerase chain reaction amplification, and sequencing. The ABCC2-24C > T (rs717620) genotype was associated with pathologic response to neoadjuvant chemotherapy. Patients with the TT and TC genotypes responded to neoadjuvant chemotherapy 3.80 times more often than those with the CC genotype (95% CI: 1.27-11.32). Patients with the CC genotype also had poorer outcomes than those with other genotypes. Thus, ABCC2-24C > T polymorphism may help to predict the response to preoperative chemotherapy in GC patients.

Hedgehog (Hh) signaling controls embryonic development and adult tissue homeostasis through the G protein coupled receptor (GPCR)-family protein Smoothened (Smo). Upon stimulation, Smo accumulates on the cell surface in Drosophila or primary cilia in vertebrates, which is thought to be essential for its activation and function, but the underlying mechanisms remain poorly understood. Here we show that Hh stimulates the binding of Smo to a plasma membrane-associated kinase Gilgamesh (Gish)/CK1γ and that Gish fine-tunes Hh pathway activity by phosphorylating a Ser/Thr cluster (CL-II) in the juxtamembrane region of Smo carboxyl-terminal intracellular tail (C-tail). We find that CL-II phosphorylation is promoted by protein kinase A (PKA)-mediated phosphorylation of Smo C-tail and depends on cell surface localization of both Gish and Smo. Consistent with CL-II being critical for high-threshold Hh target gene expression, its phosphorylation appears to require higher levels of Hh or longer exposure to the same level of Hh than PKA-site phosphorylation on Smo. Furthermore, we find that vertebrate CK1γ is localized at the primary cilium to promote Smo phosphorylation and Sonic hedgehog (Shh) pathway activation. Our study reveals a conserved mechanism whereby Hh induces a change in Smo subcellular localization to promote its association with and activation by a plasma membrane localized kinase, and provides new insight into how Hh morphogen progressively activates Smo.

The NF-κB pathway plays important roles in immune responses. Although its regulation has been extensively studied, here, we report an unknown feedforward mechanism for the regulation of this pathway by Toll-like receptor (TLR) ligands in macrophages. During bacterial infections, TLR ligands upregulate the expression of the 11S proteasome subunit PSME3 via NF-κB-mediated transcription in macrophages. PSME3, in turn, enhances the transcriptional activity of NF-κB by directly binding to and destabilizing KLF2, a negative regulator of NF-κB transcriptional activity. Consistent with this positive role of PSME3 in NF-κB regulation and importance of the NF-κB pathway in host defense against bacterial infections, the lack of PSME3 in hematopoietic cells renders the hosts more susceptible to bacterial infections, accompanied by increased bacterial burdens in host tissues. Thus, this study identifies a substrate for PSME3 and elucidates a proteolysis-dependent, but ubiquitin-independent, mechanism for NF-κB regulation that is important for host defense and innate immunity., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: The relationship between pathological factors and lymph node metastasis of pathological stage early gastric cancer has been extensively investigated. By contrast, the relationship between preoperative factors and lymph node metastasis of clinical stage early gastric cancer has not been investigated. The present study was to investigate discrepancies between preoperative and postoperative values., Methods: From January 2011 to December 2013, 1042 patients with clinical stage early gastric cancer who underwent gastrectomy with lymphadenectomy were enrolled. Preoperative and postoperative values were collected for subsequent analysis. Receiver operating characteristics curves were computed using independent predictive factors., Results: Several discrepancies were observed between preoperative and postoperative values, including existence of ulcer, gross type, and histology (all McNemar p-values were <0.001). Multivariate analyses identified the following independent predictive factors for lymph node metastasis: postoperative values including age (p = 0.002), tumor size (p < 0.001), and tumor depth (p < 0.001); preoperative values including age (p = 0.017), existence of ulcer (p = 0.037), tumor size (p = 0.009), and prediction of the presence of lymph node metastasis in computed tomography scans (p = 0.002). These postoperative and preoperative independent predictive factors produced areas under the receiver operating characteristics curves of 0.824 and 0.660, respectively., Conclusions: Surgeons need to be aware of limitations in preoperative predictions of the presence of lymph node metastasis for clinical stage early gastric cancer.

The Hippo (Hpo) tumor suppressor pathway is an evolutionarily conserved signaling pathway that controls tissue growth and organ size in species ranging from Drosophila to human, and its malfunction has been implicated in many types of human cancer. In this study, we conducted a kinome screen and identified Happyhour (Hppy)/MAP4K3 as a novel player in the Hpo pathway. Our biochemical study showed that Hppy binds and phosphorylates Wts. Our genetic experiments suggest that Hppy acts in parallel and partial redundantly with Misshapen (Msn)/MAP4K4 to regulate Yki nuclear localization and Hpo target gene expression in Drosophila wing imaginal discs. Furthermore, we showed that cytoskeleton stress restricts Yki nuclear localization through Hppy and Msn when Hpo activity is compromised, thus providing an explanation for the Wts-dependent but Hpo-independent regulation of Yki in certain contexts. Our study has unraveled an additional layer of complexity in the Hpo signaling pathway and laid down a foundation for exploring how different upstream regulators feed into the core Hpo pathway.

Background: Although our previous randomized controlled trial showed that there was no difference in postoperative complications after gastric cancer surgery between patients with and without a prophylactic drains (PDs), PDs are commonly used by most surgeons and at most institutions. However, these results have not yet been validated elsewhere. The purpose of this study was to analyze the incidence, characteristics, and risk factors for a postoperative percutaneous catheter drainage (PCD) procedure after gastric cancer surgery when PDs were not used., Methods: We reviewed data from 1989 patients who underwent gastrectomy with lymphadenectomy for gastric cancer with curative intent from January 2012 to December 2013., Results: The incidence of PCD in the abdomen was 1.8 % (22/1249) and 9.1 % (67/740) in patients with and without PD, respectively. In the without-PD group, age [odds ratio (OR) 1.032; p = 0.013], male gender (OR for female 0.38; p = 0.005), open surgery (OR for minimally invasive surgery 0.16; p = 0.013), and longer operative time (OR 1.01; p < 0.001) were independent risk factors for postoperative PCD in the abdomen. In the without-PD group, no microbes were detected in the peritoneal fluid obtained by PCD in 72.1 % (44/61) of patients who underwent PCD, and the most commonly identified organisms were Escherichia coli and Candida albicans., Conclusion: Not using a PD increased the risk of PCD postoperatively, but no microbes in peritoneal fluid were detected in the most patients. Selective use of PD in patients during gastric cancer surgery may be possible using our risk factor analysis.

Oncogenic BRAF, which drives cell transformation and proliferation, has been detected in approximately 50% of human malignant melanomas and 5% to 15% of colorectal cancers. Despite the remarkable clinical activities achieved by vemurafenib and dabrafenib in treating BRAF(V600E) metastatic melanoma, their clinical efficacy in BRAF(V600E) colorectal cancer is far less impressive. Prior studies suggested that feedback activation of EGFR and MAPK signaling upon BRAF inhibition might contribute to the relative unresponsiveness of colorectal cancer to the first-generation BRAF inhibitors. Here, we report characterization of a dual RAF kinase/EGFR inhibitor, BGB-283, which is currently under clinical investigation. In vitro, BGB-283 potently inhibits BRAF(V600E)-activated ERK phosphorylation and cell proliferation. It demonstrates selective cytotoxicity and preferentially inhibits proliferation of cancer cells harboring BRAF(V600E) and EGFR mutation/amplification. In BRAF(V600E) colorectal cancer cell lines, BGB-283 effectively inhibits the reactivation of EGFR and EGFR-mediated cell proliferation. In vivo, BGB-283 treatment leads to dose-dependent tumor growth inhibition accompanied by partial and complete tumor regressions in both cell line-derived and primary human colorectal tumor xenografts bearing BRAF(V600E) mutation. These findings support BGB-283 as a potent antitumor drug candidate with clinical potential for treating colorectal cancer harboring BRAF(V600E) mutation., (©2015 American Association for Cancer Research.)

Background: Systemic chemotherapy is the key treatment for advanced gastric cancer. The benefit of adjuvant surgery following preoperative chemotherapy in gastric cancer with liver metastasis has not been well established., Methods: Forty-nine gastric cancer patients diagnosed with synchronous liver metastasis initially treated with chemotherapy were categorized into the following two groups: surgery group: 25 patients who underwent gastrectomy and subsequently received postoperative chemotherapy and control group: 24 patients who received chemotherapy alone., Results: The median overall survival of patients in the surgery group and control group was 20.5 and 9.1 months, respectively, (P = 0.006). The median progression-free survival in the surgery group was 10.9 months, with statistical significance when compared with 5.0 months in the control group (P = 0.001). Multivariate analysis demonstrated that response to chemotherapy was the only independent factor in predicting prognosis. The survival of patients who achieved partial response (PR) was prolonged if they received adjuvant surgery (P = 0.024). No significant difference in the survival of patients underwent combined hepatic resection when compared with patients performed gastrectomy only., Conclusions: For gastric cancer with synchronous liver metastasis, adjuvant gastrectomy followed by chemotherapy might be beneficial for survival comparing with chemotherapy alone, especially in patients response to initial preoperative chemotherapy.

Xenotransplantation of human cancers into immunodeficient mice is a very useful approach for studying human tumor biology. However, the occasional occurrence of lymphomagenesis in some mice can spoil the model and must be investigated in detail. We found that a high percentage (32.5%, 26/80) of cancer patient-derived xenografts (PDXs) resembled lymphoma in NOD/SCID mice. Of the 26 xenografts, 23 were human-derived expressing human CD45 (hCD45+) and proved to be of the B-cell subtype (CD3-/CD20+), and they were all positive for Epstein-Barr virus (EBV). The remaining 3 xenografts proved to be mouse-derived for both hCD45- and negative amplification of a human gene. The most interesting finding is that gastric cancer had much higher rates (24/126, 19.0%) of lymphoma formation in the PDX model than did colorectal cancer (1/43, 2.3%). Statistical analysis revealed that cancer type and inflammation in the parent tumor are significantly associated with lymphomagenesis. Further validation discovered lymphomagenesis by inoculating only gastritis mucosa. Therefore, our findings suggest that it is necessary to take precautions when directly xenografting cancer tissues with remarkable baseline inflammation, such as gastric cancer into immunodeficient NOD/SCID strains. Further, the established xenograft models should be validated by both leukocyte markers and human gene signatures.

Many adult tissues are maintained by resident stem cells that elevate their proliferation in response to injury. The regulatory mechanisms underlying regenerative proliferation are still poorly understood. Here we show that injury induces Hedgehog (Hh) signaling in enteroblasts (EBs) to promote intestinal stem cell (ISC) proliferation in Drosophila melanogaster adult midgut. Elevated Hh signaling by patched (ptc) mutations drove ISC proliferation noncell autonomously. Inhibition of Hh signaling in the ISC lineage compromised injury-induced ISC proliferation but had little if any effect on homeostatic proliferation. Hh signaling acted in EBs to regulate the production of Upd2, which activated the JAK-STAT pathway to promote ISC proliferation. Furthermore, we show that Hh signaling is stimulated by DSS through the JNK pathway and that inhibition of Hh signaling in EBs prevented DSS-stimulated ISC proliferation. Hence, our study uncovers a JNK-Hh-JAK-STAT signaling axis in the regulation of regenerative stem cell proliferation., (© 2015 Tian et al.)

Hedgehog (Hh) signaling governs many developmental processes by regulating the balance between the repressor (Ci(R)/Gli(R)) and activator (Ci(A)/Gli(A)) forms of Cubitus interruptus (Ci)/glioma-associated oncogene homolog (Gli) transcription factors. Although much is known about how Ci(R)/Gli(R) is controlled, the regulation of Ci(A)/Gli(A) remains poorly understood. Here we demonstrate that Casein kinase 1 (CK1) sustains Hh signaling downstream of Costal2 and Suppressor of fused (Sufu) by protecting Ci(A) from premature degradation. We show that Hh stimulates Ci phosphorylation by CK1 at multiple Ser/Thr-rich degrons to inhibit its recognition by the Hh-induced MATH and BTB domain containing protein (HIB), a substrate receptor for the Cullin 3 family of E3 ubiquitin ligases. In Hh-receiving cells, reduction of CK1 activity accelerated HIB-mediated degradation of Ci(A), leading to premature loss of pathway activity. We also provide evidence that Gli(A) is regulated by CK1 in a similar fashion and that CK1 acts downstream of Sufu to promote Sonic hedgehog signaling. Taken together, our study not only reveals an unanticipated and conserved mechanism by which phosphorylation of Ci/Gli positively regulates Hh signaling but also provides the first evidence, to our knowledge, that substrate recognition by the Cullin 3 family of E3 ubiquitin ligases is negatively regulated by a kinase.

Similar to the mammalian intestine, the Drosophila adult midgut has resident stem cells that support growth and regeneration. How the niche regulates intestinal stem cell activity in both mammals and flies is not well understood. Here, we show that the conserved germinal center protein kinase Misshapen restricts intestinal stem cell division by repressing the expression of the JAK-STAT pathway ligand Upd3 in differentiating enteroblasts. Misshapen, a distant relative to the prototypic Warts activating kinase Hippo, interacts with and activates Warts to negatively regulate the activity of Yorkie and the expression of Upd3. The mammalian Misshapen homolog MAP4K4 similarly interacts with LATS (Warts homolog) and promotes inhibition of YAP (Yorkie homolog). Together, this work reveals that the Misshapen-Warts-Yorkie pathway acts in enteroblasts to control niche signaling to intestinal stem cells. These findings also provide a model in which to study requirements for MAP4K4-related kinases in MST1/2-independent regulation of LATS and YAP., (Copyright © 2014 Elsevier Inc. All rights reserved.)

A preclinical trial identified 4 of 20 (20%) gastric cancer (GC) patient-derived xenografts responded to cetuximab. Genome-wide profiling and additional investigations revealed that high EGFR mRNA expression and immunohistochemistry score (3+) are associated with tumor growth inhibition. Furthermore, EGFR amplification were observed in 2/4 (50%) responders with average copy number 5.8 and >15 respectively. Our data suggest that a GC subtype with EGFR amplification and overexpression benefit from cetuximab treatment.

Objective: To study the effect of pretreatment with apoptotic donor spleen cells on spleen lymphocyte function of recipient rats undergoing islet transplantation to explore new approaches to prolong islet graft survival., Methods: Apoptotic spleen cells from donor rats were obtained by exposure to γ-ray irradiation from (60)Co. Diabetic SD rat models were randomly divided into 4 groups to receive tail vein injections with saline (group A), normal cells (group B), apoptotic donor cells (group C), or necrotic donor cells (group D). One week later, orthotopic transplantation of islets under the renal capsule was performed. Before and at 1 and 2 weeks after islet transplantation, the recipient rats were examined for proliferative activity of spleen lymphocytes with CFSE cell staining and for IL-2 and IL-10 expressions in the cells using ELISA., Results: Pretreatment with donor apoptotic cells significantly suppressed the proliferative activity of recipient spleen lymphocytes before and at 1 and 2 weeks after islet transplantation as compared with the other three groups (P<0.05). The level of IL-2 was significantly decreased while IL-10 increased in apoptotic donor cell pretreatment group compared with those in the other 3 groups at each time point of observation., Conclusion: The effect of pretreatment with apoptotic donor cells on recipient spleen lymphocytes suggest an important role of apoptotic donor spleen cells in immune tolerance of grafts.

Objective: To investigate the expression status of human carcinoma antigen (HCA) in human cholangiocellular carcinomas, and to determine the relationship between HCA and clinical features., Methods: Tissues from 60 intrahepatic cholangiocellular carcinoma (ICC) patients, and normal liver tissues from 20 hepatic hemangioma patients selected randomly were assayed for the expression of HCA by immunohistochemistry, and Western blots. Areas of poorly differentiated (n=20), moderately-well differentiated (n=30), highly differentiated tumors (n=10) from different cases were evaluated. Results were recorded as positive (≥5% of cells staining and staining intensity 2+ or 3+) or negative (<5% of cells staining and staining intensity<2+) and analyzed using the χ2 test., Results: BCE075 and BDD048 antibodies showed similar staining patterns. The positive immunostaining of BCE075 was mainly localized in the cytoplasm and cell secretions. The staining was positive in 15% of poorly differentiated ICC, 72% of moderately-well differentiated, 100% of highly differentiated tumors. But, staining was not detected in adjacent normal tissue. The differences in HCA expression among these tissues were statistically significant. Also, we found expression of HCA to be closely associated with the degree of differentiation of ICC and tumor cell morphology. There was a correlation between expression of HCA and serum CA19-9., Conclusion: The data suggest that HCA is a potential marker for the diagnosis of cholangiocellular carcinoma., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The proproliferative transcription factor KLF5 plays an important role in promoting cell proliferation and tumorigenesis. KLF5 is a short-lived protein that can be rapidly degraded through the ubiquitin-proteasome pathway in cancer cells. However, the mechanisms regulating protein stability remain poorly understood. In this study, the tumor suppressor Fbw7, a component of the SCF complex (SCF(Fbw7)) E3 ubiquitin ligase, specifically promoted the degradation and ubiquitination of KLF5 but had little effect on the stability of KLF4. Fbw7 interacted with KLF5 in a CDC4 phosphodegron (CPD)-dependent manner. Three CPDs were found in the KLF5 protein. Simultaneous mutation of these CPDs significantly abolished Fbw7-mediated ubiquitination and degradation. Furthermore, Fbw7 deficiency dramatically delayed KLF5 turnover and led to the accumulation of KLF5 protein in cancer cells. Glycogen synthase kinase-3beta could phosphorylate and promote Fbw7-mediated KLF5 degradation. More importantly, Fbw7 negatively regulated the biological activity of KLF5 in gene regulation and cell proliferation. Taken together, these data indicate that Fbw7 is a key negative regulator controlling KLF5-mediated cell proliferation and suggest an additional mechanism linking the loss of Fbw7 function to tumorigenesis.

We report the cloning of a novel beta-glucosidase-like gene by function-based screening of a metagenomic library from uncultured soil microorganisms. The gene was named bgllC and has an open reading frame of 1,443 base pairs. It encodes a 481 amino acid polypeptide with a predicted molecular mass of about 57.8 kDa. The deduced amino acid sequence did not show any homology with known beta-glucosidases. The putative beta-glucosidase gene was subcloned into the pETBlue-2 vector and overexpressed in E. coli Tuner (DE3) pLacI; the recombinant protein was purified to homogeneity. Functional characterization with a high performance liquid chromatography method demonstrated that the recombinant BgllC protein hydrolyzed D-glucosyl-beta-(l-4)-D-glucose to glucose. The maximum activity for BgllC protein occurred at pH 8.0 and 42 degrees C using p-nitrophenyl-beta-D-glucoside as the substrate. A CaCl(2) concentration of 1 mM was required for optimal activity. The putative beta-glucosidase had an apparent K(m) value of 0.19 mM, a V(max) value of 4.75 U/mg and a k (cat) value of 316.7/min under the optimal reaction conditions. The biochemical characterization of BgllC has enlarged our understanding of the novel enzymes that can be isolated from the soil metagenome.