Journal: Frontiers in Neuroscience

Article Title: Mito-metformin protects against mitochondrial dysfunction and dopaminergic neuronal degeneration by activating upstream PKD1 signaling in cell culture and MitoPark animal models of Parkinson’s disease

doi: 10.3389/fnins.2024.1356703

Figure Lengend Snippet: Activation of PKD1 by Mito-Met in N27 cells. (A) N27 dopaminergic neuronal cells were treated with metformin (100 and 1,000 μM) for 3 h. (B) N27 cells were treated with Mito-Met (100 and 300 nM) for 3 h. Cell lysates were prepared and subjected to Western blot analysis. Representative immunoblots of PKD1 S744/748 phosphorylation are shown. (C) N27 cells were treated with 100 and 300 nM Mito-Met for 3 and 6 h. Cell lysates were prepared and subjected to Western blot analysis. Representative immunoblots of total PKD1, PKD1 S744/748, and S916 phosphorylation are shown. (D) The graph represents the densitometric analysis of phospho-PKD1 S744/748 levels in (C) . Results are the mean ± SEM of at least three independent experiments ( * p ≤ 0.05; *** p < 0.001).

Article Snippet: The PKD1 inhibitor CID755673 was purchased from Tocris Bioscience (Ellisville, MO).

Techniques: Activation Assay, Western Blot, Phospho-proteomics

Journal: Frontiers in Neuroscience

Article Title: Mito-metformin protects against mitochondrial dysfunction and dopaminergic neuronal degeneration by activating upstream PKD1 signaling in cell culture and MitoPark animal models of Parkinson’s disease

doi: 10.3389/fnins.2024.1356703

Figure Lengend Snippet: Activation of Akt and AMPK by Mito-Met in N27 cells. (A) N27 cells were treated with 100 and 300 nM Mito-Met for 3 and 6 h. Cell lysates were prepared and phospho-Akt (S473) and phospho-AMPKα (Thr172) levels were determined by Western blot analysis. (B) N27 cells were pretreated with 50 μM PKD1 inhibitor CID755673 for 1 h and then cotreated with 100 nM Mito-Met for 3 h. Cell lysates were prepared and subjected to Western blot analyses of phospho-PKD1 (S916), phospho-Akt (S473), and phospho-AMPKα (Thr172).

Article Snippet: The PKD1 inhibitor CID755673 was purchased from Tocris Bioscience (Ellisville, MO).

Techniques: Activation Assay, Western Blot

Journal: Antibiotics

Article Title: Novel Antibacterial Agents SAAP-148 and Halicin Combat Gram-Negative Bacteria Colonizing Catheters

doi: 10.3390/antibiotics12121743

Figure Lengend Snippet: Bactericidal efficacies of SAAP-148 peptide and halicin against antibiotic-resistant Gram-negative strains isolated from catheters.

Article Snippet: Halicin (SU 3327; Mw 261.3) was purchased from Tocris Bioscience (Bristol, UK).

Techniques: Isolation

Journal: Antibiotics

Article Title: Novel Antibacterial Agents SAAP-148 and Halicin Combat Gram-Negative Bacteria Colonizing Catheters

doi: 10.3390/antibiotics12121743

Figure Lengend Snippet: Reduction in bacterial counts within biofilms upon exposure to SAAP148 and halicin. In short, 24 h Gram-negative biofilms on silicone elastomer discs were exposed for 4 h and 24 h to increasing concentrations of SAAP-148 ( A ) and halicin ( B ); then, the biofilms were sonicated, and the number of surviving bacteria was assessed microbiologically. The results are expressed as the mean of the number of viable bacteria in log 10 CFU/disc. Four experiments were undertaken in triplicate. The data obtained after 4 h and 24 h exposures are represented by open black and open grey symbols, respectively. Mann–Whitney U test: * p < 0.05 indicates significantly different from control (0 ⸸ , no agent). DMSO control (0 ⸸⸸ ): control for halicin, i.e., the concentration of DMSO in the highest concentration of halicin that was tested. ns: not significantly different. Kruskal–Wallis test indicated that SAAP-148 as well as halicin at 4 h and 24 h were significantly effective against all the strains.

Article Snippet: Halicin (SU 3327; Mw 261.3) was purchased from Tocris Bioscience (Bristol, UK).

Techniques: Sonication, Bacteria, MANN-WHITNEY, Control, Concentration Assay

Journal: Antibiotics

Article Title: Novel Antibacterial Agents SAAP-148 and Halicin Combat Gram-Negative Bacteria Colonizing Catheters

doi: 10.3390/antibiotics12121743

Figure Lengend Snippet: Effects of SAAP-148 and halicin on persisters derived from antibiotic-exposed mature biofilms. In short, bacterial biofilms were cultured for seven days, washed, exposed for 3 days to antibiotics, washed, and then exposed for 4 h to increasing concentrations of the peptide or halicin. Thereafter, the biofilms were sonicated to obtain a suspension of persisters, enabling the microbiological detection of the bacterial counts. The results are expressed as the mean of the number of viable bacteria in log 10 CFU/mL. Three independent experiments in duplicate were undertaken. To enrich for persisters, E. coli EC2 and K. pneumoniae KP1 and KP2 were exposed to 50 × MBC ciprofloxacin; A. baumannii AB1, 50 × MBC gentamicin. The Kruskal–Wallis test indicated that SAAP-148 and halicin were significantly effective against all the persisters at 4 h. 0 ⸸ : number of bacteria before treatment with antibiotic. 0 ⸸⸸ : number of bacteria after 3 days of exposure to the antibiotic. ** p < 0.01 (Mann–Whitney U test) indicated significant differences from controls exposed to the antibiotic (0 ⸸⸸ ) but not SAAP-148/halicin.

Article Snippet: Halicin (SU 3327; Mw 261.3) was purchased from Tocris Bioscience (Bristol, UK).

Techniques: Derivative Assay, Cell Culture, Sonication, Suspension, Bacteria, MANN-WHITNEY

Journal: Antibiotics

Article Title: Novel Antibacterial Agents SAAP-148 and Halicin Combat Gram-Negative Bacteria Colonizing Catheters

doi: 10.3390/antibiotics12121743

Figure Lengend Snippet: Pre-incubation for 2 h of UV-inactivated GNB strains with SAAP-148, but not halicin, reduced the ability of GNBs to induce IL-12-p40 production by whole human blood leukocytes. In short, UV-killed bacteria were pre-incubated with increasing concentrations of SAAP-148 or halicin (from 1 to 1000 nM) for 2 h and then mixed with 5-fold diluted whole blood. After 20 h of incubation at 37 °C, the cells were spun down, and the levels of IL-12p40 in the supernatants were assessed using ELISA. The results are expressed as the percentage of IL-12p40 compared with that of the control (0 ⸸ ), i.e., UV-inactivated bacteria exposed to PBS instead of the peptide. 0 ⸸⸸ : UV-inactivated bacteria exposed to the highest concentration of DMSO instead of the halicin concentration that was tested. Values are the means of three independent experiments, each performed in duplicate. The Kruskal–Wallis test indicated that SAAP-148 significantly reduced the abilities of all the bacteria (EC2 and AB1: p < 0.0001; KP1: p = 0.0004 and KP2: p = 0.0005) to induce IL-12p40 production. ** p < 0.01 compared to the control (no peptide) using Mann–Whitney U test.

Article Snippet: Halicin (SU 3327; Mw 261.3) was purchased from Tocris Bioscience (Bristol, UK).

Techniques: Incubation, Bacteria, Enzyme-linked Immunosorbent Assay, Control, Concentration Assay, MANN-WHITNEY

Journal: Antibiotics

Article Title: Novel Antibacterial Agents SAAP-148 and Halicin Combat Gram-Negative Bacteria Colonizing Catheters

doi: 10.3390/antibiotics12121743

Figure Lengend Snippet: Hemolytic activities of SAAP-148 in PBS ( A ) and 50% plasma ( B ) and of halicin in PBS ( C ) and 50% plasma ( D ). Values are means of three independent experiments, each performed in triplicate.

Article Snippet: Halicin (SU 3327; Mw 261.3) was purchased from Tocris Bioscience (Bristol, UK).

Techniques:

Journal: Antibiotics

Article Title: Novel Antibacterial Agents SAAP-148 and Halicin Combat Gram-Negative Bacteria Colonizing Catheters

doi: 10.3390/antibiotics12121743

Figure Lengend Snippet: Synergistic effects of SAAP-148 and halicin alone/in combination against GNB preformed biofilms and fractional biofilm eradication concentration indexes (ΣFBEC) for combinations of SAAP-148 and halicin.

Article Snippet: Halicin (SU 3327; Mw 261.3) was purchased from Tocris Bioscience (Bristol, UK).

Techniques: Concentration Assay

Journal: Cellular signalling

Article Title: The role of PKC and PKD in CXCL12 and CXCL13 directed malignant melanoma and acute monocytic leukemic cancer cell migration.

doi: 10.1016/j.cellsig.2023.110966

Figure Lengend Snippet: Fig. 3. Effect of PKC and PKD inhibitors upon CXCL12 and CXCL13 stimulated THP-1 cell migration. THP-1 cells were treated with 30 μM GF109203X, 10 μM ZIP, 100 nM staurosporine, 30 μM CID2011756 or 30 μM CID755673 and stimulated with either A) 5 nM CXCL12 or B) 5 nM CXCL13 for 4 h. Data are means ± SEM, N = 4 analysed by two-tailed, unpaired t-tests, comparing PKC/ PKD inhibitor treatment to 5 nM CXCL12/CXCL13. Ns not significant and ****p < 0.0001.

Article Snippet: GF109203X I. Hamshaw et al. Cellular Signalling 113 (2024) 110966 (stock solution of 23.5 mM in DMSO), CID2011756 (stock solution of 50 mM in DMSO), CID755673 (stock solution of 50 mM in DMSO) and lovastatin (stock solution 7 mM in DMSO) were purchased from Tocris (Abingdon, UK).

Techniques: Migration, Two Tailed Test

Journal: Cellular signalling

Article Title: The role of PKC and PKD in CXCL12 and CXCL13 directed malignant melanoma and acute monocytic leukemic cancer cell migration.

doi: 10.1016/j.cellsig.2023.110966

Figure Lengend Snippet: Fig. 4. PKC inhibitors decrease CXCL12 stimulated SK-MEL-28 cell shape changes. SK-MEL-28 cells were treated with 30 μM GF109203X, 100 nM staurosporine, 10 μM ZIP, 30 μM CID2011756 or 30 μM CID755673 ± 10 nM CXCL12 or 10 nM CXCL13 for 24 h. Actin cytoskeleton visualised using Phalloidin-iFluor 488 reagent (green) with nuclei indicated by DAPI staining (blue). Negative control visualised using DAPI staining only. Data shows representative cells from 4 independent experiments with similar findings. Acquired with Leica imaging suite, 40× objective (22× overall magnification). Scale bar is μm. Cell area and circularity calculated in Fig. 5. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: GF109203X I. Hamshaw et al. Cellular Signalling 113 (2024) 110966 (stock solution of 23.5 mM in DMSO), CID2011756 (stock solution of 50 mM in DMSO), CID755673 (stock solution of 50 mM in DMSO) and lovastatin (stock solution 7 mM in DMSO) were purchased from Tocris (Abingdon, UK).

Techniques: Staining, Negative Control, Imaging

Journal: Cellular signalling

Article Title: The role of PKC and PKD in CXCL12 and CXCL13 directed malignant melanoma and acute monocytic leukemic cancer cell migration.

doi: 10.1016/j.cellsig.2023.110966

Figure Lengend Snippet: Fig. 5. PKC inhibitors decrease CXCL12 stimulated SK-MEL-28 cell surface area. SK-MEL-28 cells were treated with 30 μM GF109203X, 100 nM staurosporine, 10 μM ZIP, 30 μM CID2011756 or 30 μM CID755673 ± 10 nM CXCL12 or 10 nM CXCL13 for 24 h as depicted in Fig. 4. Cell area and circularity were measuring using ImageJ analysis software. 10 cells were measured per condition. Data are mean ± SEM, N = 4 analysed by two-tailed, unpaired t-tests. Ns not significant, *p < 0.05, **p < 0.01 and ***p < 0.001.

Article Snippet: GF109203X I. Hamshaw et al. Cellular Signalling 113 (2024) 110966 (stock solution of 23.5 mM in DMSO), CID2011756 (stock solution of 50 mM in DMSO), CID755673 (stock solution of 50 mM in DMSO) and lovastatin (stock solution 7 mM in DMSO) were purchased from Tocris (Abingdon, UK).

Techniques: Software, Two Tailed Test