Journal: PLoS ONE

Article Title: Adaptability and Persistence of the Emerging Pathogen Bordetella petrii

doi: 10.1371/journal.pone.0065102

Figure Lengend Snippet: (A) The DiversiLab Non fermentor typing kit was used for rep-PCR typing of B. petrii using DNA from clinical and reference strains. Amplicons were detected with the Agilent 2100 bioanalyzer (Agilent Technologies, Palo Alto, CA) and data analyzed with the DiversiLab software (version 3.3). Results generated include a dendrogram (left) and virtual gel images (right). (B) Genomic DNA was digested with the restriction endonuclease XbaI and separated by PFGE with a CHEF Mapper system. Asterisks indicate band differences among the patient strains. Ladder: Lambda DNA Ladder 48.5 KB–1 MB kb plugs (Lonza). (C) Growth curves were performed on LB broth at 37°C and growth assessed by colony-forming units (CFU) performed with serially diluted aliquots plated on SBA plates. Graph shows mean and SEM from three experiments. B. petrii 1–5: strains of B. petrii serially isolated from our patient; BAA-461: type strain of B. petrii (ATCC BAA-461); 13363: first described clinical strain of B. petrii (NCTC 13363).

Article Snippet: PCR products from our strains and B. petrii NCTC 13363 were sequenced by ACGT Inc. (Wheeling, IL).

Techniques: Software, Generated, Lambda DNA Preparation, Isolation

Journal: PLoS ONE

Article Title: Adaptability and Persistence of the Emerging Pathogen Bordetella petrii

doi: 10.1371/journal.pone.0065102

Figure Lengend Snippet: (A) Five ml overnight cultures from each bacterial strain ( B. petrii 1, B. petrii 3, NCTC 13363 and ATCC BAA-461) were spun 10 min at 8000g to harvest bacterial cells and LPS isolated by using a LPS extraction kit. Extracted LPS samples were separated on a 12% SDS-PAGE and stained using Pro-Q Emerald 300 Lipopolysaccharide gel stain kit. LPS concentrations were measured spectrophotometrically at 205 nm. Control E coli LPS was obtained commercially. I, II and III indicate LPS bands I, II and III respectively, described in the text (B) Immunoblots of our patient’s serum against LPS were performed as described in Fig. 4A. (C) Purified LPS (200 ng/ml) was added to human peripheral blood mononuclear cells (PBMCs) from normal volunteers, and supernatants were collected for cytokine measurements after 20 hours. p <0.001 for NCTC 13363 vs B. petrii 1, B. petrii 3 or BAA-461. Graph shows mean and SEM from three experiments.

Article Snippet: PCR products from our strains and B. petrii NCTC 13363 were sequenced by ACGT Inc. (Wheeling, IL).

Techniques: Isolation, SDS Page, Staining, Western Blot, Purification

Journal: Particle and Fibre Toxicology

Article Title: Chronic exposure to polystyrene microplastics induced male reproductive toxicity and decreased testosterone levels via the LH-mediated LHR/cAMP/PKA/StAR pathway

doi: 10.1186/s12989-022-00453-2

Figure Lengend Snippet: PS-MPs treatment downregulated the expression of steroidogenic enzymes and StAR in testes. Mice were given drinking water comprising PS-MPs of various sizes for 180 sustained days. A , B The expression of 3β-HSD, 17β-HSD, P450scc, P450c17, and StAR in testes was measured by western blotting. The expression levels were quantified with ImageJ (n = 3). Data are expressed as means ± SD. * P < 0.05, ** P < 0.01 vs. control. # P < 0.05, ## P < 0.01 vs. 100 μg/L group. C , D The expression of StAR in mouse testicular tissues of the control group and 1000 μg/L group was tested by immunofluorescence staining. Testicular tissues were stained with StAR (red) and DAPI (blue), arrows: positive expression (scale bar = 50 µm). The expression levels of StAR were quantified with ImageJ (n = 3). Data are expressed as means ± SD. ** P < 0.01 vs. control. # P < 0.05, *** P < 0.001 vs. control

Article Snippet: Transferred blots were incubated with rabbit anti-CYP11A (P450scc) (Proteintech Group, Rosemont, IL, USA), rabbit anti-CYP17A (P450c17) (Proteintech Group, Rosemont, IL, USA), rabbit anti-StAR (Cell Signaling Technology, USA), mouse anti-HSD3β (Proteintech Group, Rosemont, IL, USA), rabbit anti-HSD17β2 (Proteintech Group, Rosemont, IL, USA), rabbit anti-LHR (Proteintech Group, Rosemont, IL, USA), and mouse anti-GAPDH (Proteintech Group, Rosemont, IL, USA) overnight at 4° C. Specific information on the antibodies was shown in Table .

Techniques: Expressing, Western Blot, Immunofluorescence, Staining

Journal: Particle and Fibre Toxicology

Article Title: Chronic exposure to polystyrene microplastics induced male reproductive toxicity and decreased testosterone levels via the LH-mediated LHR/cAMP/PKA/StAR pathway

doi: 10.1186/s12989-022-00453-2

Figure Lengend Snippet: The expression of steroidogenic enzymes and StAR decreased in Leydig cells after PS-MPs treatment. Primary Leydig cells were exposed to 0.5 μm PS-MPs for 24 h at various concentrations as indicated. A The content of testosterone in supernatant was determined by ELISA assays (n = 3). B The mRNA expression levels of 3β-HSD, 17β-HSD, P450scc, P450c17, and StAR in Leydig cells after treatment with PS-MPs were determined by qRT-PCR (n = 3). C , D The expression of 3β-HSD, 17β-HSD, P450scc, and P450c17 in cells was measured by western blotting. The expression levels were quantified with ImageJ (n = 3). Data are expressed as means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 vs. control

Article Snippet: Transferred blots were incubated with rabbit anti-CYP11A (P450scc) (Proteintech Group, Rosemont, IL, USA), rabbit anti-CYP17A (P450c17) (Proteintech Group, Rosemont, IL, USA), rabbit anti-StAR (Cell Signaling Technology, USA), mouse anti-HSD3β (Proteintech Group, Rosemont, IL, USA), rabbit anti-HSD17β2 (Proteintech Group, Rosemont, IL, USA), rabbit anti-LHR (Proteintech Group, Rosemont, IL, USA), and mouse anti-GAPDH (Proteintech Group, Rosemont, IL, USA) overnight at 4° C. Specific information on the antibodies was shown in Table .

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Western Blot

Journal: Particle and Fibre Toxicology

Article Title: Chronic exposure to polystyrene microplastics induced male reproductive toxicity and decreased testosterone levels via the LH-mediated LHR/cAMP/PKA/StAR pathway

doi: 10.1186/s12989-022-00453-2

Figure Lengend Snippet: Overexpression of LHR alleviated the reduction in StAR, steroidogenic enzymes and testosterone levels induced by PS-MPs treatment in primary Leydig cells. A Primary cells were infected with LHR or empty vector with lentivirus for 72 h. The mRNA levels of LHR were tested by qRT-PCR (n = 3; *** P < 0.001 vs. LV-NC group). B Primary cells were infected with LHR or empty vector with lentivirus for 72 h. Then, the cells were exposed to 0.5 μm PS-MPs at a concentration of 0.2 mg/mL for 24 h. The testosterone content in the supernatant was examined by ELISA (n = 3; *** P < 0.001 vs. LV-NC group; ## P < 0.01 vs. LV-NC + PS-MPs group). C , D Primary cells were infected with LHR or empty vector with lentivirus for 72 h. Then, the cells were treated with 0.5 μm PS-MPs with a concentration at 0.2 mg/mL for 24 h. The expression of 3β-HSD, 17β-HSD, P450scc, P450c17, StAR, and LHR in cells was measured by western blotting. The expression levels were quantified with ImageJ (n = 3; * P < 0.05, ** P < 0.01 vs. LV-NC group; # P < 0.05, ## P < 0.01, ### P < 0.01 vs. LV-NC + PS-MPs group)

Article Snippet: Transferred blots were incubated with rabbit anti-CYP11A (P450scc) (Proteintech Group, Rosemont, IL, USA), rabbit anti-CYP17A (P450c17) (Proteintech Group, Rosemont, IL, USA), rabbit anti-StAR (Cell Signaling Technology, USA), mouse anti-HSD3β (Proteintech Group, Rosemont, IL, USA), rabbit anti-HSD17β2 (Proteintech Group, Rosemont, IL, USA), rabbit anti-LHR (Proteintech Group, Rosemont, IL, USA), and mouse anti-GAPDH (Proteintech Group, Rosemont, IL, USA) overnight at 4° C. Specific information on the antibodies was shown in Table .

Techniques: Over Expression, Infection, Plasmid Preparation, Quantitative RT-PCR, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing, Western Blot

Journal: Particle and Fibre Toxicology

Article Title: Chronic exposure to polystyrene microplastics induced male reproductive toxicity and decreased testosterone levels via the LH-mediated LHR/cAMP/PKA/StAR pathway

doi: 10.1186/s12989-022-00453-2

Figure Lengend Snippet: Specifications of primary antibodies

Article Snippet: Transferred blots were incubated with rabbit anti-CYP11A (P450scc) (Proteintech Group, Rosemont, IL, USA), rabbit anti-CYP17A (P450c17) (Proteintech Group, Rosemont, IL, USA), rabbit anti-StAR (Cell Signaling Technology, USA), mouse anti-HSD3β (Proteintech Group, Rosemont, IL, USA), rabbit anti-HSD17β2 (Proteintech Group, Rosemont, IL, USA), rabbit anti-LHR (Proteintech Group, Rosemont, IL, USA), and mouse anti-GAPDH (Proteintech Group, Rosemont, IL, USA) overnight at 4° C. Specific information on the antibodies was shown in Table .

Techniques: