Journal: RSC Advances

Article Title: Nanodiamond-based layer-by-layer nanohybrids mediate targeted delivery of miR-34a for triple negative breast cancer therapy

doi: 10.1039/c8ra00907d

Figure Lengend Snippet: Schematic illustration and characterization of nanohybrids. (A) Schematic illustration of the construction of FA/PS/miR-34a/PS@NDs through self-assembly approach. (B) Surface zeta potential of nanohybrids which was expressed as means ± S.D ( n = 3). (C) The size distribution of nanohybrids, which was expressed as means ± S.D ( n = 3). (D) TEM imaging. (a) PS/NDs, (b) miR-34a/PS@NDs, (c) PS/miR-34a/PS@NDs, (d) FA/PS/miR-34a/PS@NDs. (e) Magnified TEM image of the marked part in (d). (f) The corresponding FFT pattern of FA/PS/miR-34a/PS@NDs. The scale bars represent 5 nm.

Article Snippet: MicroRNA-34a (miR-34a) mimics, Cy3-labeled miR-34a (Cy3-miR-34a) mimics and scramble microRNA (NC miRNA) mimics were obtained from RiboBio Co. Ltd. RMPI 1640 medium, PBS, fetal bovine serum (FBS), trypsin/EDTA solution and penicillin/streptomycin were purchased from Gibco BRL.

Techniques: Imaging

Journal: RSC Advances

Article Title: Nanodiamond-based layer-by-layer nanohybrids mediate targeted delivery of miR-34a for triple negative breast cancer therapy

doi: 10.1039/c8ra00907d

Figure Lengend Snippet: Serum stability and cell uptake of nanohybrids. (A) The serum stability of nanohybrid-encapsulated miR-34a was evaluated by gel electrophoresis. Free miR-34a was served as control. (B) Cell uptake of Cy3-labeled miR-34a analyzed by flow cytometry. Data were presented as means ± S.D ( n = 3), ** p < 0.01. (C) Cellular uptake of Cy3-labeled miR-34a detected by confocal microscopy.

Article Snippet: MicroRNA-34a (miR-34a) mimics, Cy3-labeled miR-34a (Cy3-miR-34a) mimics and scramble microRNA (NC miRNA) mimics were obtained from RiboBio Co. Ltd. RMPI 1640 medium, PBS, fetal bovine serum (FBS), trypsin/EDTA solution and penicillin/streptomycin were purchased from Gibco BRL.

Techniques: Nucleic Acid Electrophoresis, Labeling, Flow Cytometry, Confocal Microscopy

Journal: RSC Advances

Article Title: Nanodiamond-based layer-by-layer nanohybrids mediate targeted delivery of miR-34a for triple negative breast cancer therapy

doi: 10.1039/c8ra00907d

Figure Lengend Snippet: (A) Intracellular delivery of miR-34a and analysis of Fra-1 expression. (A) miR-34a level was detected by RT-PCR. (B) Fra-1 expression was quantitatively measured by RT-PCR. Data were presented as means ± S.D ( n = 3), ** p < 0.01. (C) Fra-1 expression was analyzed by western blot.

Article Snippet: MicroRNA-34a (miR-34a) mimics, Cy3-labeled miR-34a (Cy3-miR-34a) mimics and scramble microRNA (NC miRNA) mimics were obtained from RiboBio Co. Ltd. RMPI 1640 medium, PBS, fetal bovine serum (FBS), trypsin/EDTA solution and penicillin/streptomycin were purchased from Gibco BRL.

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

Journal: RSC Advances

Article Title: Nanodiamond-based layer-by-layer nanohybrids mediate targeted delivery of miR-34a for triple negative breast cancer therapy

doi: 10.1039/c8ra00907d

Figure Lengend Snippet: In vivo antitumor activity. (A) Tumor volumes were measured at shown time-points and demonstrated as average of duplicate measurements with standard error bars ( n = 5) for each group, ** p < 0.01. (B) The tumors volumes were calculated at time of sacrifice and presented as mean ± SD, ** p < 0.01. (C) miR-34a level was quantitatively measured by RT-PCR and shown as mean ± SD, ** p < 0.01. (D) Expression of Fra-1 was quantitatively analyzed by RT-PCR and demonstrated as mean ± SD, ** p < 0.01. (E) Expression of Fra-1 was detected by western blot. (F) Representative data of H&E staining of tumor tissues. Scale bar: 50 μm.

Article Snippet: MicroRNA-34a (miR-34a) mimics, Cy3-labeled miR-34a (Cy3-miR-34a) mimics and scramble microRNA (NC miRNA) mimics were obtained from RiboBio Co. Ltd. RMPI 1640 medium, PBS, fetal bovine serum (FBS), trypsin/EDTA solution and penicillin/streptomycin were purchased from Gibco BRL.

Techniques: In Vivo, Activity Assay, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, Staining

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: Physicochemical characterization of dPG-NH2-miR-34a polyplex. A. Schematic structure of dPG-NH2-miRNA polyplex. B. Electrophoresis Mobility Shift Assay with dPG-NH2-miR-34a at N/P ratios of 0, 0.4, 0.9, 2.5, 4.5 and 9. C-D. U-87 MG glioblastoma cells were treated with dPG-NH2-miR-34a or dPGNH2-NC-miR at N/P ratios of 2.2, 4.5 and 9 at dPG-NH2 concentrations of 100, 200 and 400 nM, respectively. C. Activity of dPG-NH2-miR polyplexes at different N/P ratios was monitored by a dual luciferase assay. D. Cells viability was assessed by XTT proliferation assay. E. Zeta potential. F. Size distribution. G. SEM image. H. Table summarizing zeta potential, hydrodynamic diameter and average diameter of dPG-NH2-miR-34a polyplex (N/P 9).

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques: Electrophoresis, Mobility Shift, Activity Assay, Luciferase, Proliferation Assay, Zeta Potential Analyzer

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: dPG-NH 2 –miR-34a polyplex-treated human GBM cells express higher levels of miR-34a and lower levels of its target genes. (A-E) A172, U-87 MG and T98 human GBM cells were treated with dPG-NH 2 –miR-34a, dPG-NH 2 –NC-miR or left untreated. Twenty-four hours later, RNA was isolated and qPCR was performed to evaluate miR-34a, C-MET, CDK6, Notch1 and Bcl-2 expression levels. *** P value ≤ 0.01, ** P value ≤ 0.05 related to untreated control and to NC-miR.

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques: Isolation, Expressing, Control

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: dPG-NH 2 –miR-34a polyplex inhibits GBM miR-34a activity, cell cycle progression and cell survival. (A) Activity of a dPG-NH 2 –miR-34a polyplex monitored by a dual luciferase assay in U-87 MG, U251 and U373 GBM cells. (B-D) GBM cells were treated with dPG-NH 2 –miR-34a or dPG-NH 2 –NC-miR (100 nM miR). (B) U-87 MG cells were analyzed by flow cytometry 72 h following treatment with polyplex. (C-D) Four days later, cell proliferation was assessed by Coulter Counter. *** P value ≤ 0.01, ** P value ≤ 0.05 related to untreated control and/or NC-miR. (C) U-87 MG, A172 and T98G human GBM cell lines. (D) Human patient-derived GBM cells.

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques: Activity Assay, Luciferase, Flow Cytometry, Control, Derivative Assay

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: dPG-NH 2 –miR-34a polyplex inhibits migration of GBM cells toward serum and migration of endothelial cells toward GBM cells. Representative images and quantification of migration experiments. (A) U-87 MG. *** P value ≤ 0.01, ** P value ≤ 0.05 related to control and to NC-miR. (B) A172 cells. (C) Human umbilical vein endothelial cells (HUVEC) toward conditioned media (C.M.) from GBM cells treated with dPG-NH 2 –miR-34/NC-miR polyplex.

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques: Migration, Control

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: miR-34a and dPG-NH 2 –miR-34a polyplex physiological characterization. (A) Naked miR-34a and dPG-NH 2 –miR-34a were incubated at 37 °C in mouse plasma, with or without heparin, and analyzed by gel electrophoresis. (B) Tumor necrosis factor-alpha (TNF-α) and interleukin 6 (IL-6) cytokines secreted from human PBMCs following treatment with dPG-NH 2 , miR-34a or dPG-NH 2 –miR-34a (concentration in nM). Lipopolysaccharide (LPS) was used as positive control, PBS as negative control.

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques: Incubation, Clinical Proteomics, Nucleic Acid Electrophoresis, Concentration Assay, Positive Control, Negative Control

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: Systemically-administered dPG-NH 2 –miR-34a polyplex accumulates in glioblastoma tumors. (A-B) Cy5-dPG-NH 2 (blue) complexed with miR-34a was injected intravenously (i.v.) to mice bearing subcutaneous (A) and intracranial mCherry-labeled (B) U-87 MG tumors. Accumulation of Cy5-dPG-NH 2 –miR-34a in the tumor was followed via intravital non-invasive imaging (Maestro CRI™), quantified and plotted in a graph ( in vivo ). Mice were euthanized, and tumors and organs were resected and imaged. Cy5-polyplex distribution in tumor and organs are plotted in a graph ( ex vivo ). Percent colocalization represents the % of Cy5 fluorescence in mCherry-labeled (U-87 MG tumor) area. (C) Half-life of polyplex in plasma. Cy5-dPG-NH 2 –miR-34a was i.v. injected to non-tumor-bearing mice. Polyplex levels were monitored in blood samples taken from mice at several time points (n = 3). Fluorescence intensity ( λ Ex = 630 nm, λ Em = 670 nm) was measured using SpectraMax M5 plate reader and is plotted in a graph ± stdev.

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques: Injection, Labeling, Imaging, In Vivo, Ex Vivo, Fluorescence, Clinical Proteomics

Journal: Nanomedicine

Article Title: Restoring the oncosuppressor activity of microRNA-34a in glioblastoma using a polyglycerol-based polyplex

doi: 10.1016/j.nano.2016.05.016

Figure Lengend Snippet: dPG-NH 2 –miR-34a polyplex inhibits U-87 MG tumor growth. (A) Tumor growth. (B) Kaplan–Meier analysis following 3 consecutive treatments (marked by arrows in A ) with dPG-NH 2 –miR-34a (miR-34a), dPG-NH 2 –NC-miR (NC-miR), or PBS. Data in tumor volume graph represent mean ± s.e.m. For miR-34a treated toward NC-miR treated mice, P < 0.05 on days 20 to 30, P < 0.01 on days 32 to 62.

Article Snippet: microRNA-34a (miR-34a) (50 pmol) (Biospring, Germany) was mixed with FITC-labeled dPG-NH 2 (100 pmol) in serum-free medium, incubated for 20 min at RT, and then added to U-87 MG cells previously seeded on cover glasses.

Techniques:

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: An illustration of the study design, in vitro & in vivo tests used for evaluation of immune response after tsEV-miR-34a-mimic therapy

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: In Vitro, In Vivo

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: tsEV characterization and its loading with miR-34a. (a) tsEV average size measured by DLS (130 ± 10.99 nm). (b) Transmission electron microscopy (TEM) image of isolated tsEVs pointed by red arrows. (c) Field emission scanning electron microscopy image (FESEM) of isolated tsEVs. (d) Relative expression of miR-34a in 4T1 cells after treatment with tsEV-miR-34a-mimic, compared to tsEV, tsEV-miR-34a-inhibitor and non-treated cells (one way ANOVA, means ± S.E.M, * P < 0.05, ***P < 0.001 )

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: Transmission Assay, Electron Microscopy, Isolation, Expressing

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: CD4T and CD8T frequency and their ratio in tumor, IDLN, and spleen of different treatments. a. The frequency of CD4T, and CD8 T cells in the tsEV-miR-34a-mimic group compared to tsEV, NC-tsEV, tsEV-miR-34a-inhibitor, and the control group by Two-way ANOVA showed a pattern of increasing in CD8T cells in tumor and IDLN but not spleen. b. CD4 T/CD8T ratio analysis by one-way ANOVA in tsEV-miR-34a-mimic group compared to tsEV, NC-tsEV, tsEV-miR-34a-inhibitor and control group revealed a decline in tsEV-miR-34a mimic therapy ( n = 3, means ± S.E.M *P < 0.05 , **P < 0.01 , ***P < 0.001 ,, ns, non significant)

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: Control

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: Treg frequency and its ratio in proportion to CD8T cells in different tissues after treatment. (a) Bar plots of comparisons of evaluated Treg frequencies after different therapies. The population of Treg fell in the tumor, IDLN, and spleen by tsEV-miR-34a-mimic therapy. (b) Treg/CD8T ratios in different tissues of tested subjects. The highest decline was seen in the Treg/CD8T ratio of tsEV-miR-34a-mimic therapy which reflected a major CD8T rise and Treg drop by this treatment toward other therapies. The data was shown as means ± SEM, ( n = 3,*P < 0.05, **P < 0.01, ***P< 0.001 , ns, non significant) and analyzed by one-way ANOVA

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques:

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: Relative expression of tumor-promoting and immune-inhibitory targets of miR-34a in tumor tissue. In total a sharp drop was observed in tsEV-miR-34a-mimic group toward other treated groups in all evaluated genes except for MMP2. The comparison was made between control, NC-tsEV, tsEV, tsEV-miR-34a-mimic/inhibitor by one-way ANOVA and represented as means ± SEM ( n = 3,*P < 0.05, **P < 0.01, ***P < 0.001 , ns, non significant)

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: Expressing, Comparison, Control

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: T cells master transcription factors relative expression in tumor tissue. Differences in transcription factors of T cells main lineages in the tumor after tsEV-miR-34a-mimic therapy with domination of Th1 response exhibited as mean ± SEM and analyzed by one-way ANOVA ( n = 3, *P < 0.05, **P < 0.01 ***P < 0.001 ,ns, non significant)

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: Expressing

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: Killing capacity and stimulating index of IDLN and spleen isolated lymphocytes.( a ) Comparison of percentages of 4T1 apoptosis in tsEV-miR-34a-mimic treated mice across other treatments by IDLN lymphocytes with one-way ANOVA test. ( b ) 4T1 cells apoptosis percentage after exposure to splenocytes analyzed by one way ANOVA. c. Rate of proliferation in Lymphocytes of different treated groups toward control after stimulation with 4T1 cell lysate analyzed by two-way ANOVA test. ( d ). Rate of proliferation in splenocytes of different treated groups toward control after stimulation with 4T1 cell lysate. (Two-way ANOVA test) The data was represented as mean ± SEM ( n = 3, *P < 0.05 , **P < 0.01, ***P< 0.001 , ns, non significant)

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: Isolation, Comparison, Control

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: Histopathological findings. Metastasis in IDLN ( a ), lung ( b ), and liver( c ) of different therapies. In tsEV-miR-34a-mimic treated mice all evaluated tissues were metastasis-free compared to PBS received group. ( d ) necrosis of tumor tissue which was bordered by dotted lines and black stars. The highest rate of necrosis was detected in the tsEV-miR-34a-mimc group compared to other therapeutic agents. ( e ) Tumor infiltrating lymphocytes after different treatments were shown by red arrows (x400). An increase in the accumulation of stromal TIL was seen in the tsEV-miR-34a-mimic-treated group. (Scale bar x100: 1600 μm, Scale bar x400: 400 μm, A: PBS, B: NC-tsEV, C: tsEV, D: tsEV-miR-34a-mimic, E: tsEV-miR-34a-inhibitor)

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques:

Journal: Cancer Cell International

Article Title: Tumor-derived small extracellular vesicles loaded with functionally active miR-34a mimic can modify the anti-tumor response in 4T1 breast cancer animal model

doi: 10.1186/s12935-025-03994-6

Figure Lengend Snippet: The anti-tumor effects of tsEV-miR-34a-mimic in 4T1 animal model. (a) Tumor volume was decreased significantly in the tsEV-miR-34a-mimic treated group compared to PBS receiving group (b) In all treated groups tumor weight was reduced drastically toward control. The maximum decline was caused by the tsEV-miR-34a-mimic group. c. tsEV-miR-34a-mimic treatment induced an increase in survival days of treated mice against PBS administered group. The data was represented as mean ± SEM and analyzed by Kaplan-Meier method ( n = 5, *P < 0.05, **P < 0.01, ***P < 0.001, ns, non significant)

Article Snippet: miR-34a-mimic and inhibitor (Bioneer, Korea) were separately loaded in tsEVs based on the modified CaCl 2 method [ ].

Techniques: Animal Model, Control