Journal: bioRxiv

Article Title: Magnetic torque-driven living microrobots for enhanced tumor infiltration

doi: 10.1101/2022.01.03.473989

Figure Lengend Snippet: ( A ) Representative confocal images of immunostaining for VE-cadherin (green) on HMEC-1 cells cultured on a Transwell membrane. Cell nuclei were stained using Hoechst 33342 (blue). ( B ) Evaluation of the lower and upper compartment fluorescence intensity measurements following 1 hour of passive diffusion of liposomes across HMEC-1 monolayers (n = 3; mean ± SD). ( C ) Comparison of lower compartment MTB-LP concentrations after 1 hour for unactuated controls and exposure to either DMF (12 mT) and out-of-plane RMF (20 mT and 24 Hz) (n = 3; mean ± SD; *P < 0.05, ANOVA). No gradients were applied in these experiments. ( D ) Pre- and post-experimental transendothelial electrical resistance (TEER) measurements for unactuated controls and out-of-plane RMF ( n = 3; mean ± SD). ( E ) Pre- and post-experimental Lucifer yellow (LY) rejection values for unactuated controls and out-of-plane RMF ( n = 3; mean ± SD).

Article Snippet: For semiquantitative biodistribution analysis, far-red fluorescence signals were measured using an ex vivo fluorescence imaging system (Sapphire Biomolecular Imager, Azure Biosystems).

Techniques: Immunostaining, Cell Culture, Membrane, Staining, Fluorescence, Diffusion-based Assay, Liposomes, Comparison

Journal: bioRxiv

Article Title: Magnetic torque-driven living microrobots for enhanced tumor infiltration

doi: 10.1101/2022.01.03.473989

Figure Lengend Snippet: ( A ) Representative confocal images of live MCF-7 spheroids following 1 hour exposure to RMF (20 mT and 24 Hz), thorough washing, and 24 hours incubation. Control refers to unactuated samples. Images were captured at 10 μm increments from the bottom of the spheroids and show the localization of DiO-labelled liposomes (green). Outline depicts the shape of the spheroid at its largest circumference. Scale bar = 100 μm. ( B ) Plot of mean intensity for each section at 10 μm intervals from the bottom of the spheroids along the z-axis up to a depth of 100 μm. ( C ) Representative fluorescence intensity distribution of the 80 µm section. Values were normalized to overall minimum and maximum fluorescence intensity values. Spheroid diameter was normalized along the x-axis. ( D ) Summation of mean intensity values for consecutive Z-plane images up to a depth of 100 μm ( n = 3; mean ± SD; ** P < 0.01, Student’s t-test). ( E ) Representative z-projections of MTB stained with a far-red proliferative dye in live MCF-7 spheroids following 1 hour exposure to RMF (20 mT and 24 Hz), thorough washing, and incubation for up to 120 h without actuation. Control refers to unactuated samples. Images were captured at 24 and 120 h. Outline depicts the shape of the spheroid at its largest circumference. Representative normalized fluorescence intensity distribution of actuated samples and controls at 24 and 120 h (center). Values were normalized to overall minimum and maximum fluorescence intensity values. Spheroid diameter was normalized along x-axis. Image-based quantification of fluorescence intensity values from z-projections at 24 and 120 h (right; n = 3; mean ± SD; * P < 0.05, ** P < 0.01, Student’s t-test). ( F ) Representative images of 5 μm histology sections for MCF-7 spheroids 144 h after actuation.

Article Snippet: For semiquantitative biodistribution analysis, far-red fluorescence signals were measured using an ex vivo fluorescence imaging system (Sapphire Biomolecular Imager, Azure Biosystems).

Techniques: Incubation, Liposomes, Fluorescence, Staining

Journal: bioRxiv

Article Title: Magnetic torque-driven living microrobots for enhanced tumor infiltration

doi: 10.1101/2022.01.03.473989

Figure Lengend Snippet: A) Representative z-projections from live HCT 116 spheroids following 1 hour exposure to RMF (20 mT and 24 Hz), thorough washing, and incubation for up to 120 h without actuation. MTB were stained with a far-red proliferative stain and images were captured at 120 h. Control refers to unactuated samples. Scale bar = 200 μm. Normalized fluorescence intensity distribution of actuated samples and controls at 120 h (center). Values were normalized to overall minimum and maximum fluorescence intensity values. Spheroid diameter was normalized along x-axis. Image-based quantification of fluorescence intensity values from z-projections at 120 h (right; n = 3; mean ± SD).

Article Snippet: For semiquantitative biodistribution analysis, far-red fluorescence signals were measured using an ex vivo fluorescence imaging system (Sapphire Biomolecular Imager, Azure Biosystems).

Techniques: Incubation, Staining, Fluorescence

Journal: bioRxiv

Article Title: Magnetic torque-driven living microrobots for enhanced tumor infiltration

doi: 10.1101/2022.01.03.473989

Figure Lengend Snippet: ( A ) BALB/c nude mice bearing subcutaneous MCF-7 tumors in one hind flank were intravenously administered with 1 × 10 9 MTB stained with a far-red proliferative dye. The mice were anaesthetized for 1 h in the absence of exposure to magnetic actuation (control) or were placed on a magnetic field generator with tumors positioned in the workspace (RMF). Mice were returned to the cage for 24 h, after which the tumor and major organs were harvested for further analysis. ( B ) Plots of expected field magnitude and gradients produced in the workspace of the magnetic field generator for an applied field of 20 mT in x. ( C ) Representative ex vivo fluorescence intensity images of major organs and tumors 24 h after injection of far-red stained MTB. (D) Quantitative biodistribution from harvested organs and tumors ( n = 3; mean ± SD; ** P < 0.01, Student’s t-test). (E) C mag values for homogenized tumors placed in liquid culture for 8 days ( n = 3 control, n = 4 RMF; mean ± SD). (F) Representative images of 10 μm histology sections which were sectioned at depth of approximately 1000 μm in the tumour. Cell nuclei were stained using Hoechst 33342 (blue). (G) Mean intensity values with increasing distance from the periphery of the tumor sections (n = 3; mean ± SD). (H) Representative transverse tumor sections. Cell nuclei were stained using Hoechst 33342 (blue). (I) Plot of mean intensity for each traverse tumor section at 200 μm intervals. Values were normalized to overall minimum and maximum fluorescence intensity values. ( J ) Summation of mean intensity values for consecutive traverse tumor sections ( n = 3; mean ± SD).

Article Snippet: For semiquantitative biodistribution analysis, far-red fluorescence signals were measured using an ex vivo fluorescence imaging system (Sapphire Biomolecular Imager, Azure Biosystems).

Techniques: Staining, Produced, Ex Vivo, Fluorescence, Injection

Journal: bioRxiv

Article Title: Magnetic torque-driven living microrobots for enhanced tumor infiltration

doi: 10.1101/2022.01.03.473989

Figure Lengend Snippet: ( A ) Schematic of MCF-7 spheroid with illustration of z-axis planes. Images were captured at 10 μm intervals from the bottom of the spheroids up to a depth of 100 μm. ( B ) Z-projection of Hoechst image stack with overlaid regions of interest (ROIs). To quantify the average fluorescence of DiO liposomes for each spheroid, ROIs were defined using the Hoechst image stack by tracing a contour around the spheroid at 0 and 100 µm. Intermediate ROIs were defined using interpolation, with approximately 3 µm radial increments. These ROIs were then applied to the corresponding DiO liposome image stack. Scale bar = 200 μm. ( C ) Representative images of sections from DiO liposome image stacks showing an interpolated ROI (yellow line). These fluorescence values were normalized to the average fluorescence of the surrounding media (ROI = gray rectangle). Scale bar = 200 μm.

Article Snippet: For semiquantitative biodistribution analysis, far-red fluorescence signals were measured using an ex vivo fluorescence imaging system (Sapphire Biomolecular Imager, Azure Biosystems).

Techniques: Fluorescence, Liposomes

Journal: Materials Today Bio

Article Title: Robust drug bioavailability and safety for rheumatoid arthritis therapy using D-amino acids-based supramolecular hydrogels

doi: 10.1016/j.mtbio.2022.100296

Figure Lengend Snippet: Design and characterization of the hydrogels. (a) The chemical structures of Nap-GFFY, Nap-G D F D F D Y, MTX-GFFY, and MTX-G D F D F D Y. (b) TEM images of the hydrogels. Inserted images: hydrogels formed by 10.317 ​mM (1 ​wt%) of different peptides in PBS. Scale bar ​= ​100 ​nm. (c) Cirular dichrorism (CD) revealed the chirality of the hydrogles. (d) Gelation behavior of MTX-G D F D F D Y hydrogels before and after injection. (e) HPLC analysis of the hydrogels exposed to proteinase K. (f) Compound remained rate calculated from (e). (g) Cumulative MTX release from MTX-GFFY and MTX-G D F D F D Y hygrogels in PBS within 24 ​h ( n ​= ​3). (h) Fluoresence imaging of the mice after the intra-articular injection of Cy5.5-GFFY or Cy5.5-G D F D F D Y hydrogel at day 0, 1, 5, 7, and 10.

Article Snippet: At day 0, 1, 5, 7, and 10, mice were imaged using an in vivo fluoresence imaging system (FX PRO, BRUKER, Germany) with an excitation wavelength of 630 nm and an emission wavelength of 700 nm.

Techniques: Injection, 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: (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: Bioactive Materials

Article Title: Modular assembled biomimetic nanobubbles for synergistic therapy of ischemic stroke via cascade modulation thrombo-inflammatory network

doi: 10.1016/j.bioactmat.2025.06.054

Figure Lengend Snippet: In vitro and in vivo biological effects and targeting characterization of PFNBs. (A) Schematic diagram of the endothelial barrier penetration ability evaluation of PFNBs, and (B) quantitative analysis results (n = 3). (C) Schematic diagram of the enhanced uptake effect of PFNBs by microglia with/without inflammatory activation. (D) confocal microscopy images, and (E) quantitative analysis results (n = 3). Scale bar, 10 μm. (F) Comparison of the PNBs and PFNBs uptaken by microglia with/without inflammatory activation at 4 h (n = 3). (G) Near-infrared fluorescence images of PFNBs targeting AIS lesions, and (H) quantitative analysis results (n = 3). (I) Near-infrared fluorescence images of PFNBs distribution in brains and major organs, as well as (J) quantitative analysis results (n = 3). Error bars: mean ± standard deviation. ns indicates non-significant (p > 0.05). ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001 (two-tailed Student's t -test).

Article Snippet: The plasma pharmacokinetics of DiR-labeled PNBs and PFNBs were assessed by monitoring the fluorescence signal using a near-infrared fluorescence in vivo imaging system (IVIS Spectrum Imaging System, Caliper Life Sciences, USA).

Techniques: In Vitro, In Vivo, Activation Assay, Confocal Microscopy, Comparison, Fluorescence, Standard Deviation, Two Tailed Test

Journal: Bioactive Materials

Article Title: Modular assembled biomimetic nanobubbles for synergistic therapy of ischemic stroke via cascade modulation thrombo-inflammatory network

doi: 10.1016/j.bioactmat.2025.06.054

Figure Lengend Snippet: Evaluation of the immunomodulatory effects of PFNBs in vitro and in vivo . (A) Confocal fluorescence microscopy images of iNOS and CD206 immunofluorescence staining in inflammatory activated microglia after being incubated with Saline, free FTY720, PNBs, and PFNBs. Scale bar, 50 μm. (B) Quantitative analysis of iNOS (M1) and CD206 (M2) fluorescence intensity based on confocal images (n = 3). (C) WB bands of iNOS, CD206, p-STAT3 and t-STAT3 proteins. (D) Quantification of iNOS/GAPDH, CD206/GAPDH and p-STAT3/t-STAT3 levels,/Saline represents the change of experimental group relative to Saline group (n = 3). (E) Immunofluorescence staining and quantitative analysis of M1 (F) and M2 (G) phenotypes of microglia in the ischemic lesion (n = 5). Scale bar: 50 μm. (H) WB bands of iNOS, CD206, p-STAT3, and t-STAT3 proteins in lesion brain tissues of AIS mice. (I) Quantification of iNOS/GAPDH, CD206/GAPDH and p-STAT3/t-STAT3 levels in lesion brain tissues of AIS mice. Saline represents the change of experimental group relative to Saline group (n = 3). Error bars: mean ± standard deviation. ns indicates non-significant (p > 0.05). ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001 (two-tailed Student's t -test).

Article Snippet: The plasma pharmacokinetics of DiR-labeled PNBs and PFNBs were assessed by monitoring the fluorescence signal using a near-infrared fluorescence in vivo imaging system (IVIS Spectrum Imaging System, Caliper Life Sciences, USA).

Techniques: In Vitro, In Vivo, Fluorescence, Microscopy, Immunofluorescence, Staining, Incubation, Saline, Standard Deviation, Two Tailed Test

Journal: Bioactive Materials

Article Title: Modular assembled biomimetic nanobubbles for synergistic therapy of ischemic stroke via cascade modulation thrombo-inflammatory network

doi: 10.1016/j.bioactmat.2025.06.054

Figure Lengend Snippet: Evaluation of the protective effects on BBB permeability and vascular integrity of PFNBs in vitro and in vivo . (A) Schematic time line protocol of in vitro BBB model establishment and subsequent operations. (B) TEER value monitoring of endothelial cell monolayers after OGD/R treatment with addition of Saline, free FTY720, PNBs, and PFNBs (n = 3). (C) Proteomic analysis of the species and expression abundance of growth factors carried by platelet membranes in PLTs, PMVs, PNBs, and PFNBs (n = 3). (D) TEER value monitoring of endothelial cell monolayers seeded with inflammatory activated microglia in the lower chamber after OGD/R treatment with addition of Saline, free FTY720, PNBs, and PFNBs (n = 3). Confocal fluorescence microscopy images of CD34 and ZO-1 (E) or occluding (F) double-labeled immunofluorescence staining in the lesion area 24 h after AIS modeling and administration. Scale bar, 20 μm. Quantitative analysis of the ratio of ZO-1/CD34 positive area (G) or occludin/CD34 positive area (H) (n = 5). Photos of AIS mice brains in each group after Evans blue injection (I) and quantitative analysis of Evans blue leakage (J) (n = 3). (K) Quantitative analysis of brain edema in each group of AIS mice (n = 3). Error bars: mean ± standard deviation. ns indicates non-significant (p > 0.05). ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001 (two-tailed Student's t -test).

Article Snippet: The plasma pharmacokinetics of DiR-labeled PNBs and PFNBs were assessed by monitoring the fluorescence signal using a near-infrared fluorescence in vivo imaging system (IVIS Spectrum Imaging System, Caliper Life Sciences, USA).

Techniques: Permeability, In Vitro, In Vivo, Saline, Expressing, Fluorescence, Microscopy, Labeling, Immunofluorescence, Staining, Injection, Standard Deviation, Two Tailed Test

Journal: Gut Microbes

Article Title: Extracellular vesicles of Fusobacterium nucleatum compromise intestinal barrier through targeting RIPK1-mediated cell death pathway

doi: 10.1080/19490976.2021.1902718

Figure Lengend Snippet: FnEVs increase gut barrier leakage in experimental colitis models . (a) Representative images and ex vivo imaging with the intestine, liver, heart, spleen and kidney of mice. (b) Relative fluorescence intensity of translocated EGFP-labeled E.coli in every tissues. (c) Representative images of immunohistochemical stainings of ZO-1, claudin-1 and occludin in the colon on day 3 after colitis induction. Scale bar = 50 um. (d) The relative mRNA level of mucin1, mucin2, ZO-1 and claudin-1 was detected in colon samples on day 3 after colitis induction. *p < .05, **p < .01, ***p < .001. All data were presented as the means ± SD (n = 6 mice per group)

Article Snippet: After 24 h, the mice were sacrificed to harvest intestinal and extra-intestinal organs (gut, liver, heart, spleen, and kidney) to study with the Bruker in vivo fluorescence imaging system.

Techniques: Ex Vivo, Imaging, Fluorescence, Labeling, Immunohistochemical staining