Journal: JCI Insight

Article Title: Nononcogenic restoration of the intestinal barrier by E . coli –delivered human EGF

doi: 10.1172/jci.insight.125166

Figure Lengend Snippet: (A) EGFR expression levels in the colon or ileum of patients with Crohn’s disease (CD) or ulcerative colitis (UC). Three data sets (Sleiman’s, Gene Expression Omnibus [GEO] ID GSE10616, n = 58; Vemeire’s, GEO ID GSE75214, n = 194; and Haberman’s GEO ID GSE57945, n = 322) were compared. Haberman’s samples were only from the ileum from patients with UC or ileocolonic CD (iCD). The asterisks in box-and-whisker plots (min to max) represent significant differences between 2 groups (*P < 0.05, **P < 0.01, and ***P < 0.001 using 2-tailed, unpaired Student’s t test). The box plot depicts the minimum and maximum values (whiskers), the upper and lower quartiles, and the median. The length of the box represents the interquartile range. (B) The scheme for the bacterial secretion of LARD3-linked EGF protein through prtDEF. Genes for human mature EGF polypeptide linked to 1 fragment of LARD3 and prtDEF in pKOV vector were encoded in the bacterial chromosome by a crossing-over recombination at the OmpC region. Based on SWISS-MODEL (https://swissmodel.expasy.org/), expected binding modes in the bacterial membrane are depicted, based on SWISS-MODEL. (C) Colonization activity of E. coli OP50 and EGF-EcN in C. elegans intestine in the presence of mucoadherent enteropathogenic E. coli (EPEC). Gut colonization was assessed in wild-type (N2) C. elegans after 24 hours of preincubation with mCherry-labeled nonpathogenic E. coli (OP50 or EGF-EcN) in the presence of GFP-labeled EPEC (original magnification, ×100). The pictures are representative of 10 independent observations. (D) Stool samples from EGF-EcN–treated mice were collected at indicated times after gavage. Colonized bacteria levels were estimated based on PCR with EcN-specific primers for Muta5/6. (E) GFP-labeled EGF-EcN (1 × 109 CFU/200 μL) in the gut after gavage. The isolated intestines were observed on the 4th and 29th days after gavage. The colonizing bacteria were visualized in the intestine via an in vivo imaging system.

Article Snippet: To assess bacterial colonization in the gut, recombinant bacteria transformed with pGFP-UV plasmid were applied to the mice via gavage, and the green fluorescence in the isolated intestine was detected by using a fluorescence in vivo imaging system (FM03-B, NeoScience).

Techniques: Expressing, Gene Expression, Whisker Assay, Plasmid Preparation, Binding Assay, Membrane, Activity Assay, Labeling, Bacteria, Isolation, In Vivo Imaging

Journal: JCI Insight

Article Title: Nononcogenic restoration of the intestinal barrier by E . coli –delivered human EGF

doi: 10.1172/jci.insight.125166

Figure Lengend Snippet: Eight-week-old female C57BL/6 mice (n = 8–15) were treated twice with 3% DSS after oral gavages with 1 × 109 EcN or EGF-EcN. Mice were then injected (intraperitoneally) with EGFR inhibitor (1 mg AG1478/mouse, Selleckchem) twice at 2-day intervals before and after the last day of DSS exposure. (A) Schematic outline of EGFR inhibition in the EcN or EGF-EcN treatment and DSS-induced colitis model. (B) Expression of p-EGFR in mucosa (green with the white arrow) was quantified (left box-and-whisker plot, min to max, based on fluorescence microscopy observations, shown at right) (*P < 0.05; **P < 0.01; ***P < 0.001; ns using 2-tailed, unpaired Student’s t test). (C) Representative hematoxylin and eosin (H&E) staining of the intestinal lesions as demonstrated by microscopy (original magnification, ×200). Scale bar: 100 μm. (D) Colons were isolated for analysis of goblet cells and mucin production and were stained with Alcian blue (original magnification, ×400. Scale bar: 100 μm). Each histogram represents events at an increasing Alcian blue level. A quantitative comparison is shown in the right graph (*P < 0.05; **P < 0.01; ***P < 0.001; ns using 2-tailed, unpaired Student’s t test). (E) Gram staining (original magnification, ×400. Scale bar: 100 μm). The asterisks represent significant differences between 2 groups (the right graph) (**P < 0.01; ***P < 0.001 using 2-tailed, unpaired Student’s t test).

Article Snippet: To assess bacterial colonization in the gut, recombinant bacteria transformed with pGFP-UV plasmid were applied to the mice via gavage, and the green fluorescence in the isolated intestine was detected by using a fluorescence in vivo imaging system (FM03-B, NeoScience).

Techniques: Injection, Inhibition, Expressing, Whisker Assay, Fluorescence, Microscopy, Staining, Isolation, Comparison

Journal: International Journal of Molecular Sciences

Article Title: Chemokine Receptors CCR6 and PD1 Blocking scFv E27 Enhances Anti-EGFR CAR-T Therapeutic Efficacy in a Preclinical Model of Human Non-Small Cell Lung Carcinoma

doi: 10.3390/ijms24065424

Figure Lengend Snippet: Anti-tumor function of EGFR-T cells enhanced in E27 and CCR6 in vivo. ( A ) Schematic diagram of the mouse animal model. ( B ) The monitoring chart of fluorescence values of tumor development of mice in each group at the time of D39. ( C ) Statistics on fluorescence values of mice. The highest fluorescence value was recorded at each time point. The error line represents SEM, n = 5. ( D ) Dissected tumors and their sizes. ( E ) Statistical results of tumor weight among different groups; SEM was the error line, and the statistical method was the unpaired t test, p < 0.05 (*).

Article Snippet: The injection was performed by anesthetizing the mice with 300 mg/kg of tribromoethanol injected into the abdominal cavity, followed by a tail vein injection of 150 mg/kg body weight of in vivo fluorescence imaging substrate (Promega, Madison, WI, USA).

Techniques: In Vivo, Animal Model, Fluorescence

Journal: PLOS Pathogens

Article Title: Mucosal immunization with the lung Lactobacillus -derived amphiphilic exopolysaccharide adjuvanted recombinant vaccine improved protection against P . aeruginosa infection

doi: 10.1371/journal.ppat.1012696

Figure Lengend Snippet: (A) Representative TEM images of EPS301 and EPS301@rPcrV particles. Scale bars, 100 nm. (B) Size distributions of EPS301 and EPS301@rPcrV. DLS results showed the size of self-assembled EPS301 about 24 ± 6 nm, EPS301@rPcrV about 43.10 ± 5 nm. (C) The Zeta potentials of EPS301, rPcrV and EPS301@rPcrV. Zeta potentials results showed EPS301 about -38.47 ± 0.53, rPcrV about -20.83 ± 0.19, EPS301@rPcrV about -17.05 ± 0.24. (D) Representative in vivo fluorescence images of mice at the indicated time points after intranasal administration of free rPcrV or EPS301@ rPcrV. rPcrV in both groups labeled with Cy7. (E) Relative fluorescence intensity of rPcrV or EPS301@rPcrV in mouse nasal cavity and lung. (F) Representative ex vivo fluorescence images of major tissues (nasal cavity, lung, heart, liver, spleen and kidney) at 9 h, 48 h and 96 h after intranasal administration of free rPcrV or EPS301@ rPcrV. rPcrV in both groups labeled with Cy7. Significant differences were calculated with One- or Two-way ANOVA followed by Tukey’s multiple comparisons test. ns, not significant, *p < 0.05, **p < 0.01, ***p < 0.001. Data are presented as means ± SEM.

Article Snippet: Mice were anesthetized by isoflurane and imaged by a fluorescence vivo imaging system (VISQUE, Smart-LF, Kibero) at different time points (5mins, 3 h, 6 h, 9 h, 12 h, 24 h, 48 h, 72 h and 96 h) after inoculation.

Techniques: In Vivo, Fluorescence, Labeling, Ex Vivo