Journal: Military Medical Research

Article Title: Octopus-inspired engineered bacteria with a plug-and-play surface display system achieves enhanced tumor-specific colonization and antitumor immunity

doi: 10.1016/j.mmr.2026.100030

Figure Lengend Snippet: Schematic of the construction and therapeutic mechanism of the octopus-inspired triple-engineered bacteria (OITE strain) . The antitumor attenuated Salmonella typhimurium AISI strain was triple-engineered. (1) ST/SC-RGD×4 “tentacles”-Surface modification: the ST sequence was expressed within the external third loop of OmpA in the AISI strain, resulting in the AISI-ST strain. The ST protein was coexpressed with OmpA (OmpA-ST fusion protein) localized to the bacterial outer membrane. An incubation with SC-RGD×4 protein (SC-RGD×4) led to the formation of AISI-ST/SC-RGD×4 strain (AISI-ST/SC-RGD×4) through ST-SC-mediated covalent conjugation. (2) Dynamic EPS “camouflage”-Immunoactivation engineering: the AISI-H-ST strain was created by introducing the quorum-sensing (QS) promoter pLuxI to control HtrA expression into AISI-ST strain, which specifically increased HtrA-mediated extracellular polysaccharide (EPS) production to amplify bacteria-mediated immune activation. (3) Anti-PD1nanobody (PD1nb) “secretion”-Checkpoint blockade engineering: further programming of AISI-H-ST strain enabled QS-triggered anti-PD1nb secretion, generating the AISI-HP-ST strain. The incubation of AISI-HP-ST strain with SC-RGD×4 protein finally produced the OITE strain (AISI-HP-ST/SC-RGD×4). The intravenously administered OITE strain highly selectively accumulates in tumors through RGD-αvβ3 integrin interactions, with subsequent bacterial proliferation initiating two therapeutic actions: HtrA-mediated immune activation via increasing immune cells infiltration and activation, and secreted PD1nb-based blockade of PD-1/PD-L1 immunosuppressive signaling. OITE. Octopus-inspired, triple-engineered bacterium; AISI. Attenuated Salmonella Δ htrA :: luxI - VNP20009 strains; ST. SpyTag; SC. SpyCatcherΔ; RGD. Arginine-glycine-aspartic acid; HtrA. High-temperature requirement A; PD1. Programmed cell death protein 1; H. HtrA; HP. HtrA and PD1nb; AHL. N-acyl homoserine lactone; MACS. Macrophages; Teffs. Effector T cells; Tregs. Regulatory T cells.

Article Snippet: To prepare the surfaces for the strain adhesion assays, the recombinant mouse integrin αvβ3 (TGAV & ITGB3) heterodimer protein (HY-P700761, MCE, USA) was first diluted to 10 μg/ml in 50 mmol/L carbonate buffer (pH 9.6).

Techniques: Bacteria, Modification, Sequencing, Membrane, Incubation, Conjugation Assay, Control, Expressing, Activation Assay, Produced

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Self-Triggered Apoptosis Enzyme Prodrug Therapy (STAEPT): Enhancing Targeted Therapies via Recurrent Bystander Killing Effect by Exploiting Caspase-Cleavable Linker.

doi: 10.1002/advs.201800368

Figure Lengend Snippet: Figure 2. Enhanced cellular uptake and cytotoxic activity of RGDEVD-DOX in integrin αvβ3 expressing cells. a) Representative confocal images (left) and quantitative analysis (right) of HDMEC and U-87 MG cells exposed to fluorescent-labeled RDEVD and RGDEVD peptides. b) Representative confocal images (left) and quantitative analysis (right) of scrambled control and integrin αv (ITGAV) siRNA-transfected HDMECs and U-87 MG cells exposed to fluorescent-labeled RGDEVD peptide. Green and blue indicate the fluorescent-labeled peptides and cell nuclei, respectively. Scale bar, 50 µm. c) Flow cytometry analysis of isotype control (top), scrambled control-transfected (lower left), and ITGAV siRNA-transfected (lower right) U87 MG cells incubated with fluorescent-labeled RGDEVD and stained with an antibody against integrin αvβ3. d) Representative confocal images (left) and quantitative analysis (right) of U-87 MG and HT-29 cells treated with RDEVD-DOX and RGDEVD-DOX. Red and blue indicate the intrinsic red fluorescence of doxorubicin and cell nuclei, respectively. Scale bar, 50 µm. e) Concentration-dependent cytotoxicity of RDEVD-DOX and RGDEVD-DOX on U-87 MG (left) and HT-29 (right) determined by MTT assay (n = 4). f) Doxorubicin release from RGDEVD-DOX when incubated in PBS (pH 7.4) containing (or not containing) carboxylesterase (n = 3). Data are mean ± s.d. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: The cells were incubated with Alexa Fluor 488-conjugated integrin αvβ3 antibody (1:100; R&D Systems, Minneapolis, MN; Cat. No. FAB3050G) for an hour at 4 °C, washed, and suspended in PBS containing 0.5% BSA.

Techniques: Activity Assay, Expressing, Labeling, Control, Transfection, Flow Cytometry, Incubation, Staining, Fluorescence, Concentration Assay, MTT Assay

Journal: Bioengineering

Article Title: Effects of Oral Cavity Stem Cell Sources and Serum-Free Cell Culture on Hydrogel Encapsulation of Mesenchymal Stem Cells for Bone Regeneration: An In Vitro Investigation

doi: 10.3390/bioengineering11010059

Figure Lengend Snippet: Effects of synthetic serum on the expression of adhesion molecules on hydrogel-encapsulated hBFP-hADCs in synthetic serum (SS-OS) and fetal bovine serum (FBS-OS). Quantitative real-time reverse-transcription polymerase chain reaction (qt-RT-PCR) exhibiting the expression fold change of alpha 5 beta 3 (ITG-αVβ3) and beta 1 (ITG-β1) integrins in FBS-OS relative to those of the SS-OS.

Article Snippet: Quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR) was performed to determine the expression levels of the osteoblast-associated gene Runx2, bone morphogenetic protein 2 (BMP2), and bone gamma-carboxyglutamate protein (BGLAP) as markers of the early, middle, and late stages of osteoblastic differentiation [ ], and the expression levels of the alpha v beta 3 (ITG-αVβ3) and beta 1 (ITG-β1) integrins were examined to demonstrate the expression levels of the adhesion molecules (QuantiStudio 3, Applied Biosystems, San Francisco, CA, USA).

Techniques: Expressing, Reverse Transcription, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction

Journal: The Journal of biological chemistry

Article Title: Chlorotoxin binds to both matrix metalloproteinase 2 and neuropilin 1.

doi: 10.1016/j.jbc.2023.104998

Figure Lengend Snippet: Figure 8. Binding of CTX-Cy5 and CTX-displaying phages to various proposed target proteins and human serum albumin in the cobalt-coated bead test. “His-tag isolation and pulldown” Dynabeads were coated with various His-tagged proteins at 0.16 μM for each. The tested target proteins were MMP-2, NRP1, CLC-3 chloride channel, TIMP-2, MT1-MMP (MMP14), annexin A2 (ANX2), αvβ3 integrin (INT), MMP-9, and human serum albumin (HSA). A, the coated beads were blocked with casein and stained with 1 μM CTX-Cy5. Control beads (CONT) were not incubated with any target protein but blocked with casein and stained. B, representative intensity distribution histograms of beads from an experiment of A. C, the coated beads were blocked with casein, incubated with CTX-displaying phages (5.5 × 1014 particles/ml), and stained to detect M13 phage binding. Control beads (CONT) were not incubated with any target protein but blocked with casein, and phages were also applied and stained. The bar graphs in A and C represent relative fluorescence intensities calculated from median fluorescence intensities of 10,000 events recorded by the flow cytometer and are presented as mean ± SD of three separate experiments with superimposed scatter plot. * and *** indicate statistically significant difference (p < 0.05 and p < 0.001) from control (ANOVA followed by Dunnett’s test). D, representative intensity distribution histograms of beads from the experiment of C. Note the very similar patterns of binding of fluorescent-labeled and phage-displayed CTX to various target proteins. CTX, chlorotoxin; MMP-2, matrix metalloproteinase 2; NRP1, neuropilin 1; TIMP-2, tissue inhibitor of metalloproteinase 2.

Article Snippet: The used target proteins were the following (with National Center for Biotechnology Information accession number [segment amino acid sequence]; His-tag position: C- or N-terminal; supplier; and catalog number listed in parentheses): MMP2 (NP_004521.1 [amino acids: 110–660]; N-; ProSpec, catalog no.: ENZ 769), NRP1 (NP_001019799.2 [amino acids: 2–644]; C-; Sino Biological; catalog no.: 10011-H08H); MMP-9 (NP_004985.2 [amino acids: 20–701]; C-; ProSpec; catalog no.: ENZ 1091), TIMP-2 (NP_003246.1 [amino acids: 27–220]; N-; ProSpec; catalog no.: ENZ-646), MMP 14 (NP_004986 [amino acids: 24–524]; C-; ThermoFisher; catalog no.: RP77533), ANX2 (NP_001002857.1 [amino acids: 1–339]; N-; ProSpec; catalog no.: PRO-777), αvβ3 integrin (AAA52589.1 & NP_002196.4 heterodimer; both with C-terminal His tag; Native Antigen Company; catalog no.: REC31719-100), CLC-3 (NP_001820.2 [amino acids: 1–818]; C-; Creative Biomart; custom made by recombinant expression in E. coli), and human serum albumin (NP_000468.1 [amino acids: 25–609]; C-; Abcam; catalog no.: ab217817).

Techniques: Binding Assay, Isolation, Staining, Control, Incubation, Cytometry, Labeling

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Expression of FN and αvβ3 in osteosarcoma and corresponding osteochondroma

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques: Expressing

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Correlation between FN and αvβ3 expression in osteosarcoma

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques: Expressing

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Association between individual FN and αvβ3 expression and clinicopathological characteristics in osteosarcoma

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques: Expressing

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Association between co-expression of FN and αvβ3 and clinicopathological characteristics in osteosarcoma

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques:

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Univariate analysis of FN and αvβ3 expression and osteosarcoma patient survival based on the log-rank test

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques: Expressing

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Kaplan-Meier analyses of disease-free survival (DFS) and overall survival (OS) time by FN and αvβ3 expression. a , b Significant differences in DFS ( P < 0.001) and OS ( P < 0.001) time were observed between high FN expression (FN + ) and low/negative FN expression (FN − ) groups. c , d Significant differences in DFS ( P < 0.001) and OS ( P < 0.001) time were observed between high αvβ3 expression (αvβ3 + ) and low/negative αvβ3 expression (αvβ3 − ) groups. e , f Significant differences in DFS ( P < 0.001) and OS ( P < 0.001) time were demonstrated between FN + /αvβ3 + and “others” groups. g A significant difference in DFS ( P < 0.001) time was demonstrated between FN + /αvβ3 + and FN − /αvβ3 − groups. h Significant differences in OS time were demonstrated between FN + /αvβ3 + and FN − /αvβ3 − groups ( P < 0.001) and between FN + /αvβ3 + and “others” groups ( P = 0.005). *, “others” included the single high-expression and double low/negative-expression groups (FN + /αvβ3 − plus FN − /αvβ3 + plus FN − /αvβ3 − ); †, “others” included the single high-expression groups (FN + /αvβ3 − plus FN − /αvβ3 + )

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques: Expressing

Journal: World Journal of Surgical Oncology

Article Title: Clinicopathological and prognostic values of fibronectin and integrin αvβ3 expression in primary osteosarcoma

doi: 10.1186/s12957-019-1566-z

Figure Lengend Snippet: Co-expression of FN and αvβ3 and osteosarcoma patient survival based on multivariate analysis

Article Snippet: The sections were incubated overnight at 4 °C with antibodies against FN (mouse monoclonal 2755-8; 1:50; Santa Cruz, USA) and αvβ3 (rabbit polyclonal orb10927; 1:50; Biorbyt, UK).

Techniques: Comparison