CEBPB promotes gastrointestinal motility dysfunction after severe acute pancreatitis via the MALAT1/CIRBP/ERK axis.

Severe acute pancreatitis (SAP) is a kind of reversible inflammatory process of the exocrine pancreas with gastrointestinal motility dysfunction involved. Studies have highlighted the role of long noncoding RNA metastasis associated lung adenocarcinoma transcript 1 (MALAT1) in AP. However, the mechanism underlying its role in the gastrointestinal motility dysfunction remains undefined. Hence, we explored the regulatory role of MALAT1 in gastrointestinal motility dysfunction following SAP. Then, the expression of CCAAT/enhancer-binding protein beta (CEBPB), MALAT1 and cold-inducible RNA binding protein (CIRBP) was detected in plasma of SAP patients and pancreatic and intestinal tissues of SAP mouse models with their correlation analyzed also. Additionally, the effect of MALAT1 on the pancreatic and intestinal injury, expression of inflammatory factors and the ERK pathway-related genes as well as gastrointestinal motility dysfunction was assessed using ectopic expression and depletion experiments. CEBPB, MALAT1 and CIRBP were highly expressed in plasma of SAP patients and pancreatic and intestinal tissues of SAP mice. Further analysis showed that knockdown of MALAT1 could alleviate pancreatic and intestinal injury, reduce inflammation, and prevent gastrointestinal motility dysfunction in SAP mice. The transcription factor CEBPB could bind to the promoter region of MALAT1, thus activating the transcription of MALAT1. MALAT1 interacted with CIRBP and inhibited the degradation of CIRBP, leading to activated extracellular signal-regulated kinase (ERK) pathway and the resultant gastrointestinal motility dysfunction. In conclusion, CEBPB exhibits a promoting activity towards gastrointestinal motility dysfunction in SAP by pumping up MALAT1 expression and activating the CIRBP-dependent ERK pathway.

Construction and application of a multifunctional CHO cell platform utilizing Cre/lox and Dre/rox site-specific recombination systems.

During the development of traditional Chinese hamster ovary (CHO) cell lines, target genes randomly integrate into the genome upon entering the nucleus, resulting in unpredictable productivity of cell clones. The characterization and screening of high-yielding cell lines is a time-consuming and expensive process. Site-specific integration is recognized as an effective approach for overcoming random integration and improving production stability. We have designed a multifunctional expression cassette, called CDbox, which can be manipulated by the site-specific recombination systems Cre/lox and Dre/rox. The CDbox expression cassette was inserted at the Hipp11(H11) locus hotspot in the CHO-K1 genome using CRISPR/Cas9 technology, and a compliant CHO-CDbox cell platform was screened and obtained. The CHO-CDbox cell platform was transformed into a pool of EGFP-expressing cells using Cre/lox recombinase-mediated cassette exchange (RMCE) in only 2 weeks, and this expression remained stable for at least 75 generations without the need for drug stress. Subsequently, we used the Dre/rox system to directly eliminate the EGFP gene. In addition, two practical applications of the CHO-CDbox cell platform were presented. The first was the quick construction of the Pembrolizumab antibody stable expression strain, while the second was a protocol for the integration of surface-displayed and secreted antibodies on CHO cells. The previous research on site-specific integration of CHO cells has always focused on the single functionality of insertion of target genes. This newly developed CHO cell platform is expected to offer expanded applicability for protein production and gene function studies.

The Evaluation of Clinical Status of Endoscopic Retrograde Cholangiography for the Placement of Metal and Plastic Stents in Cholangiocarcinoma Therapy.

Objective: Cholangiocarcinoma is a common malignant tumor that occurs in the bile duct system, which can be treated by using the endoscopic retrograde cholangiography (ERCP). This study was aimed at exploring the therapeutic effect of ERCP with metal stent and plastic stent for cholangiocarcinoma. Methods: The clinical data of 71 patients with cholangiocarcinoma treated by ERCP in our hospital from June 2020 to October 2021 were retrospectively analyzed. According to different stent types, the patients were divided into plastic stent group (n = 43) and metal stent group (n = 28). Patients in the plastic stent group and metal stent group were received with plastic stent and metal stent, respectively. The indexes of liver function (serum alkaline phosphatase (ALT), direct bilirubin (DBIL), glutamic oxaloacetic transaminase (AST), alkaline phosphatase (ALP), and total bilirubin (TBIL)), postoperative complications, success rate of stent implantation, and survival time of patients in the two groups were determined. Logistic multivariate regression analysis was used to analyze the prognostic factors of postoperative cholangiocarcinoma. Results: The liver function indexes of the two groups were significantly improved after treatment with the stent, in which the ameliorative effect in the metal stent group was better than that in the plastic stent group (P < 0.05). The incidence of postoperative complications in the plastic stent group and the metal stent group was 53.49% and 14.29%, respectively, and the success rate of stent placement was 60.47% and 96.43%, respectively. The incidence of complications in the metal stent group was lower than that in the plastic stent group, and the success rate of stent placement was higher than that in the plastic stent group (P < 0.05). The median survival time of patients in the plastic stent group and the metal stent group was 8.15 and 11.83 months, respectively. The survival time of patients in the metal stent group was longer than that of the plastic stent group. The median survival time of patients with types I, II, III, and IV was 12.73, 11.54, 10.57, and 9.36 months, respectively. The survival time of patients with stage I was significantly higher than that of patients with types II, III, and IV. There was an inverse relationship between the disease type and the survival time of patients. Logistic multivariate regression analysis showed that tumor diameter ≥ 5 cm, portal vein invasion, lymph node metastasis, and classification of hilar cholangiocarcinoma were the risk factors (P < 0.05) and metal stent type was the protective factor (P < 0.05). Conclusion: In the clinical treatment of patients with cholangiocarcinoma, the placement of metal stent and plastic stent under ERCP plays an important role. The placement of the metal stent under ERCP has a higher success rate and better prognosis and can prolong the survival time of patients to a greater extent, but the price of the metal stent is relatively expensive. For patients with an expected survival period of more than 4-6 months, the metal stent should be considered; otherwise, the plastic stent can be used to maintain cost-effectiveness. Therefore, it is necessary to comprehensively analyze the patient's economic affordability, expected survival time, stent drainage time, and personal needs and then select an appropriate treatment method.

A nanobody-based molecular toolkit for ubiquitin-proteasome system explores the main role of survivin subcellular localization.

Targeted protein degradation is a powerful tool for determining the function of specific proteins nowadays. Survivin is the smallest member of the inhibitor of the apoptosis protein (IAP) family. It exists in the cytoplasm and nucleus of cells, but the exact function of survivin in different subcellular locations retained unclear updates due to the lack of effective and simple technical means. In this study, we created a novel nanoantibody-based molecular toolkit, namely, the ubiquitin-proteasome system (Nb4A-Fc-T2A-TRIM21), that can target to degrade survivin localized in cytoplasmic and cell nuclear by ubiquitinating, and by which to verify the potential roles of survivin subcellular localization. Also, the results showed that the cytoplasmic survivin mainly plays an anti-apoptotic function by directly or indirectly inhibiting the caspase pathway, and the nuclear survivin mainly promotes cell proliferation and participates in the regulation of the cell cycle. In addition, the Nb4A-Fc-T2A-TRIM21 system can degrade the endogenous survivin protein in a large amount by the ubiquitin-proteasome pathway, and the system can provide theoretical support for ubiquitination degradation targeting other endogenous proteins.

Cannabidiol Effectively Promoted Cell Death in Bladder Cancer and the Improved Intravesical Adhesion Drugs Delivery Strategy Could Be Better Used for Treatment.

Cannabidiol (CBD), a primary bioactive phytocannabinoid extracted from hemp, is reported to possess potent anti-tumorigenic activity in multiple cancers. However, the effects of CBD on bladder cancer (BC) and the underlying molecular mechanisms are rarely reported. Here, several experiments proved that CBD promoted BC cells (T24, 5637, and UM-UC-3) death. For example, T24 cells were treated with 12 µM CBD for 48 h, flow cytometry analysis demonstrated that early and late apoptotic cells were accounted for by 49.91%, indicating CBD enhanced cell apoptosis ability. To deeper explore molecular mechanisms, the CBD-treated T24 cell transcriptome libraries were established. KEGG analysis implied that the significantly changed genes were enriched in the PI3K/Akt pathway. qRT-PCR and Western blot assays verified that CBD regulated BC cells growth and migration and induced apoptosis by inactivating the PI3K/Akt pathway. Meanwhile, the developed chitosan to wrap CBD-loaded PLGA nanoparticles can significantly enhance the adhesion of the material to the mouse bladder wall, and the binding efficiency of mucin to chitosan-PLGA nanoparticles reached 97.04% ± 1.90%. In summary, this work demonstrates that CBD may become a novel reliable anticancer drug and the developed intravesical adhesion system is expected to turn into a potential means of BC chemotherapy drug delivery.

LncRNA MAFG-AS1 Upregulates Polo-Like Kinase-1 by Sponging miR-505 to Promote Gastric Adenocarcinoma Cell Proliferation.

Gastric cancer is a commonly diagnosed, often fatal malignancy and requires novel anticancer therapies and preventative approaches. This study described the involvement of MAFG-AS1, a lncRNA with important functions in cancer biology, in gastric adenocarcinoma (GA). Thirty-six male and forty-two female GA patients with an average age of 51.9 ± 5.7 years in the range of 35 to 68 years were enrolled. Paired gastric cancer (GC) and non-tumor tissues were collected from each patient. MAFG-AS1 expression was determined. RNA interaction prediction, dual luciferase reporter assay, RT-qPCR assay, Western blot, and CCK-8 assay were conducted. The results indicated that MAFG-AS1 was highly expressed in GA and closely correlated with poor survival. MAFG-AS1 interacted with miR-505, but MAFG-AS1 and miR-505 overexpression showed no significant effects on each other's expression. In addition, MAFG-AS1 increased the expression of PLK1, a miR-505 target. MAFG-AS1 and PLK1 overexpression increased GC cell proliferation rate. MiR-505 overexpression reduced the effects of MAFG-AS1 and PLK1 overexpression on cell proliferation. Therefore, MAFG-AS1 might upregulate PLK1 by sponging miR-505 to promote GA cell proliferation.

Discovering and Characterizing of Survivin Dominant Negative Mutants With Stronger Pro-apoptotic Activity on Cancer Cells and CSCs.

Survivin as a member of the inhibitor of apoptosis proteins (IAPs) family is undetectable in normal cells, but highly expressed in cancer cells and cancer stem cells (CSCs) which makes it an attractive target in cancer therapy. Survivin dominant negative mutants have been reported as competitive inhibitors of endogenous survivin protein in cancer cells. However, there is a lack of systematic comparative studies on which mutants have stronger effect on promoting apoptosis in cancer cells, which will hinder the development of novel anti-cancer drugs. Here, based on the previous study of survivin and its analysis of the relationship between structure and function, we designed and constructed a series of different amino acid mutants from survivin (TmSm34, TmSm48, TmSm84, TmSm34/48, TmSm34/84, and TmSm34/48/84) fused cell-permeable peptide TATm at the N-terminus, and a dominant negative mutant TmSm34/84 with stronger pro-apoptotic activity was selected and evaluated systematically in vitro. The double-site mutant of survivin (TmSm34/84) showed more robust pro-apoptotic activity against A549 cells than others, and could reverse the resistance of A549 CSCs to adriamycin (ADM) (reversal index up to 7.01) by decreasing the expression levels of survivin, P-gp, and Bcl-2 while increasing cleaved caspase-3 in CSCs. This study indicated the selected survivin dominant negative mutant TmSm34/84 is promising to be an excellent candidate for recombinant anti-cancer protein by promoting apoptosis of cancer cells and their stem cells and sensitizing chemotherapeutic drugs.

Description of Ornithinibacillus gellani sp. nov., a halophilic bacterium isolated from lake sediment, and emended description of the genus Ornithinibacillus.

A Gram-stain-positive, strictly aerobic, motile and rod-shaped bacterium, designated strain LJ137T, was isolated from the sediment of Taihu Lake in China. A polyphasic approach was used to investigate its taxonomic position. Strain LJ137T grew optimally at pH 7.5, at 37 °C and with 2.5 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain LJ137T was most closely related to the genera Ornithinibacillus and Oceanobacillus. The closest phylogenetic neighbours were Ornithinibacillus halophilus KCTC 13822T, Ornithinibacillus salinisoli LCB256T and Oceanobacillus limi KCTC 13823T, with 95.2, 96.5 and 95.6 % 16S rRNA gene sequence similarity, respectively. The peptidoglycan amino acid type was A4α (l-Lys-d-Asp). The major respiratory quinone was menaquinone-7 (MK-7). The polar lipids of strain LJ137T contained diphosphatidylglycerol, phosphatidylglycerol, three unidentified phospholipids, two aminophospholipids and one unidentified lipid. The G+C content of the genomic DNA was 40.4 mol%. The dominant cellular fatty acids were anteiso-C15 : 0, anteiso-C17 : 0 and iso-C15 : 0. Based on the phenotypic, chemotaxonomic, phylogenetic and genome sequence characteristics of this strain, a novel species, Ornithinibacillus gellani sp. nov., is proposed. The type strain is LJ137T (=CGMCC 1.13678T=NBRC 113552T). An emended description of the genus Ornithinibacillus is presented.

Brevundimonas lutea sp. nov., isolated from lake sediment.

A novel Gram-negative, aerobic, rod-shaped bacterium, designated NS26T, was isolated from a sediment sample collected from Taihu Lake in China. Colonies were orange, circular, smooth and neat-edged on Reasoner's 2A agar. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain NS26T belonged to the genus Brevundimonas and had the closest relationship with Brevundimonas halotolerans DSM 24448T (96.9 %). It grew at 20-37 °C (optimum, 28 °C), pH 5.5-10.5 (pH 7.0) and without NaCl. The major isoprenoid quinone was Q-10. The dominant cellular fatty acids were C18 : 1ω7c, C16 : 0 and C18 : 1ω7c 11-methyl. The polar lipid profile comprised 1,2-diacyl-3-O-(6-phosphatidyl-α-d-glucopyranosyl) glycerol, 1,2-di-O-acyl-3-O-α-d-glycopyranuronosyl glycerol, sulfoquinovosyl diacylglycerol, 1,2-di-O-acyl-3-O-[d-glycopyranosyl-(1→4)-α-d-glucopyranuronosyl] glycerol and phosphatidylglycerol. The G+C content of genomic DNA was 68.4 mol%. The average nucleotide identity value between strain NS26T and B.halotolerans DSM 24448T was 75.6 %. Based on the polyphasic taxonomic study, strain NS26T is suggested to be a novel species, for which the name Brevundimonas lutea sp. nov. is proposed. The type strain is NS26T (=CGMCC 1.13680T=NBRC 113554T).

Rufibacter sediminis sp. nov., isolated from freshwater lake sediment.

A novel Gram-stain-negative, aerobic and rod-shaped bacterial strain, designated H-1T, was isolated from the interfacial sediment of Taihu Lake in China and characterized by using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the organism was most closely related to Rufibacter immobilis MCC P1T and Rufibacter tibetensis 1351T, with sequence similarities of 98.0 and 97.6 %, respectively. DNA-DNA relatedness between strain H-1T and R. immobilis MCC P1T and R. tibetensis 1351T was 48.8 and 36.6 %, respectively. The major (>5 %) cellular fatty acids were summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c), C16 : 1ω5c, iso-C15 : 0, summed feature 4 (iso-C17 : 1 I and/or anteiso-C17 : 1 B) and anteiso-C15 : 0. The polar lipids comprised phosphatidylethanolamine, three unidentified aminophospholipids, an unidentified phospholipid and three unidentified lipids. The major respiratory quinone was menaquinone 7 (MK-7). Genomic DNA G+C content was 49.0 mol%. Based on its physiological, biochemical and chemotaxonomic characteristics, the strain represents a novel species of the genus Rufibacter, for which the name Rufibacter sediminis sp. nov. (type strain H-1T=CGMCC 1.16289T=NBRC 113030T) is proposed.

Model Selection for Non-Negative Tensor Factorization with Minimum Description Length.

Non-negative tensor factorization (NTF) is a widely used multi-way analysis approach that factorizes a high-order non-negative data tensor into several non-negative factor matrices. In NTF, the non-negative rank has to be predetermined to specify the model and it greatly influences the factorized matrices. However, its value is conventionally determined by specialists' insights or trial and error. This paper proposes a novel rank selection criterion for NTF on the basis of the minimum description length (MDL) principle. Our methodology is unique in that (1) we apply the MDL principle on tensor slices to overcome a problem caused by the imbalance between the number of elements in a data tensor and that in factor matrices, and (2) we employ the normalized maximum likelihood (NML) code-length for histogram densities. We employ synthetic and real data to empirically demonstrate that our method outperforms other criteria in terms of accuracies for estimating true ranks and for completing missing values. We further show that our method can produce ranks suitable for knowledge discovery.

A novel genus of the class Actinobacteria, Longivirga aurantiaca gen. nov., sp. nov., isolated from lake sediment.

A novel actinobacterial strain, designated X5T, was isolated from the sediment of Taihu Lake in China and was subjected to a polyphasic taxonomic characterization. The strain formed orange-red colonies comprising aerobic, Gram-stain-negative, rod-shaped cells on R2A agar. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the organism was closely related to the genus Sporichthya and consistently formed a distinct clade along with the members of this genus. The closest phylogenetic neighbour was Sporichthya polymorpha NBRC 12702T with 93.7 % 16S rRNA gene sequence similarity. The major fatty acids (>10 %) were iso-C16 : 0 (18.7 %), C18 : 1ω9c (18.6 %) and C17 : 1ω8c (14.0 %). The genomic DNA G+C content was 74.4 mol%. The organism contained menaquinone MK-8(H2), MK-9(H4) and an unidentified menaquinone. Polar lipids were composed of phosphatidylglycerol, an unidentified lipid, two unidentified phospholipids and two unidentified aminolipids. The whole-cell sugars contained ribose, xylose, mannose, glucose and galactose. The cell-wall peptidoglycan contained ll-diaminopimelic acid. Based on the physiological, biochemical and chemotaxonomic data, the organism is proposed to represent a novel genus and species, for which the name Longivirga aurantiaca gen. nov., sp. nov. is proposed. The type strain is X5T (=CGMCC 4.7317T=NBRC 112237T).