Journal:

Article Title: Relationships between Colony Morphotypes and Oil Tolerance in Rhodococcus rhodochrous

doi:

Figure Lengend Snippet: Growth of rhodococci on seawater-based medium containing the AF of Arabian light crude oil a

Article Snippet: The results supported and generalized our notion that S-2 EPS protects rough Rhodococcus strains from the toxicity of n -hexadecane. table ft1 table-wrap mode="anchored" t5 TABLE 3 caption a7 Strain % of surviving cells Without EPS With EPS R. coprophilus ATCC 29080 0.1 (±0.2) 1.8 (±0.78) R. erythropolis IFO 15567 0.9 (±1.2) 75.3 (±81.6) R. erythropolis JCM 3201 0.1 (±0.1) 48.6 (±25.4) R. globerulus IFO 14531 0.01 (±0.01) 8.6 (±0.1) R. opacus ATCC 51881 1.1 (±1.0) 18.2 (±1.1) R. rhodochrous ATCC 13808 0 (±0) 72.6 (±6.7) R. zopfii ATCC 51349 0.5 (±0.5) 46.7 (±23.6) Open in a separate window a The EPS concentration used in this test was 100 μg/ml.

Techniques:

Journal:

Article Title: Relationships between Colony Morphotypes and Oil Tolerance in Rhodococcus rhodochrous

doi:

Figure Lengend Snippet: Effect of S-2 EPS on the survival of resting cells of Rhodococcus strains treated with 10% (vol/vol) n -hexadecane a

Article Snippet: The results supported and generalized our notion that S-2 EPS protects rough Rhodococcus strains from the toxicity of n -hexadecane. table ft1 table-wrap mode="anchored" t5 TABLE 3 caption a7 Strain % of surviving cells Without EPS With EPS R. coprophilus ATCC 29080 0.1 (±0.2) 1.8 (±0.78) R. erythropolis IFO 15567 0.9 (±1.2) 75.3 (±81.6) R. erythropolis JCM 3201 0.1 (±0.1) 48.6 (±25.4) R. globerulus IFO 14531 0.01 (±0.01) 8.6 (±0.1) R. opacus ATCC 51881 1.1 (±1.0) 18.2 (±1.1) R. rhodochrous ATCC 13808 0 (±0) 72.6 (±6.7) R. zopfii ATCC 51349 0.5 (±0.5) 46.7 (±23.6) Open in a separate window a The EPS concentration used in this test was 100 μg/ml.

Techniques:

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: ( A ) Schematic representation of the injury model used in this study. Mice underwent left (L) renal pedicle clamping (IR) for 31 minutes, and nephrectomy of the right kidney (R UNX) was performed 8 days later. Mice were euthanized at the time points indicated. d–1 indicates preinjured mice. ( B ) Levels of phosphorylated and total DDR1 were analyzed by Western blot in kidney cortices isolated from uninjured (d–1) or injured WT mice at the time points indicated. ( C ) DDR1 and β-tubulin bands were quantified by densitometry, and values are expressed as DDR1/β-tubulin ratio and represent the mean ± SD of n ≥ 3 mice/group. Statistical analysis: 1-way ANOVA followed by Dunnett’s multiple-comparison test versus uninjured mice. * P < 0.05, *** P < 0.001, **** P < 0.0001. ( D ) Kidney sections from uninjured (d–1) or injured Ddr1 tm1a/tm1a mice (d–1 n = 3, d3 n = 4, d7 n = 3, d10 n = 1, d13 n = 1, d28 n = 3) euthanized at the time points indicated were stained for β-gal (blue staining) and lotus tetragonolobus lectin (LTL, a marker of proximal tubules, red staining) as described in the Methods. Note the increased expression of β-gal staining in injured proximal tubules. Scale bar: 25 μm.

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Western Blot, Isolation, Comparison, Staining, Marker, Expressing

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: ( A ) Havcr1 (KIM-1) mRNA levels in WT and Ddr1-KO mice uninjured (d–1) or 3 days after IR were analyzed by quantitative PCR and normalized to Gapdh mRNA. Circles represent individual kidneys, and the bars show mean ± SD. Uninjured WT and Ddr1-KO n = 4, injured WT n = 8, injured Ddr1-KO n = 6. ( B ) Periodic acid–Schiff (PAS) staining of kidneys from WT and Ddr1-KO mice uninjured (d–1) and d3 after IR. Scale bar: 50 μm. ( C ) Acute injury was evaluated in uninjured (d–1) (WT and Ddr1-KO n = 3) and d3 injured mice (WT n = 15, Ddr1-KO n = 9). Circles represent individual kidneys, and bars show mean ± SD. ( D ). Blood urea nitrogen (BUN) was measured prior to injury (d–1) and at d9 and d28 after injury. Circles represent individual mice, and bars show mean ± SD. ( E ) PAS staining of kidneys from WT and Ddr1-KO mice uninjured (d–1) and 28 days after injury. Scale bar: 50 μm. ( F ) Tubular injury scores were evaluated in uninjured (d–1) (WT and Ddr1-KO n = 5) and d28 injured (WT n = 10, Ddr1-KO n = 6) mice. Circles represent single mice, and bars are mean ± SD. Statistical analysis: 1-way ANOVA followed by Tukey’s multiple-comparison test for A , C , and F and 2-way repeated measures ANOVA followed by Sidak’s multiple-comparison test for D .

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Real-time Polymerase Chain Reaction, Staining, Comparison

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: ( A ) Images of kidney sections from WT and Ddr1-KO mice uninjured (d–1) or 28 days after IR stained with anti–α-SMA antibody. Scale bar: 50 μm. ( B ) The percentage of α-SMA–positive area per microscopic field was evaluated at d28 using ImageJ. Circles represent individual kidneys (WT and Ddr1-KO n = 4), and bars are mean ± SD. ( C ) Picrosirius red staining of kidney sections from WT and Ddr1-KO mice uninjured (d–1) or 28 days after IRI. Scale bar: 50 μm. ( D ) The percentage of Picrosirius red–positive area was evaluated using ImageJ. Circles represent individual kidneys (d–1 WT and Ddr1-KO n = 4, d28 WT and Ddr1-KO n = 5), and bars are mean ± SD. ( E ) Kidney lysates from uninjured WT (d–1) and WT and Ddr1-KO 28 days after IR were analyzed for the level of collagen I by Western blot analysis. ( F ) Collagen I and β-tubulin bands were quantified by densitometry analysis, and collagen I is expressed as collagen I/β-tubulin ratio. Circles represent individual kidneys (d–1 WT n = 4, d28 WT and Ddr1-KO n = 5), and bars are mean ± SD. Statistical analysis: 2-tailed t test for B and 1-way ANOVA followed by Tukey’s multiple-comparison test for D and F .

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Staining, Western Blot, Comparison

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: ( A ) MCP-1 levels were measured by ELISA in conditioned medium of RPTECs treated with vehicle or collagen I (CI, 50 μg/mL) with or without the DDR1 inhibitor Cmp-1 (3 μM). Each circle represents 1 experiment performed in triplicates. Data represent mean ± SD of 5 experiments and are expressed as fold-change relative to vehicle-treated cells assigned a value of 1. ( B ) Ccl2 (MCP-1) mRNA levels were measured in kidneys of uninjured (d–1) or 3 days injured WT and Ddr1-KO mice by real-time quantitative PCR and normalized to Gapdh mRNA. Circles represent individual kidneys (d–1 WT and Ddr1-KO n = 3, d3 WT n = 9, d3 Ddr1-KO n = 6), and bars are mean ± SD. I Images of kidney sections from uninjured (d–1) or 3 days injured WT and Ddr1-KO mice stained with anti-F4/80 antibody. Scale bar: 50 μm. ( D ) The number of F4/80 positive cells per microscopic field was evaluated and expressed as F4/80-positive cells/microscopic field. Circles represent individual kidneys (d–1 WT and Ddr1-KO n = 3, d3 WT and Ddr1-KO n = 7), and bars are mean ± SD. Statistical analysis: 1-way ANOVA followed by Dunnett’s multiple comparison versus CI-treated cells for A and 1-way ANOVA followed by Tukey’s multiple-comparison test for B and D .

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Staining, Comparison

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: ( A ) Vector- or DDR1-HEK cells were treated with collagen I (50 μg/mL) or vehicle, then lysed and immunoprecipitated with anti-Flag antibody, and analyzed for levels of total and phosphorylated DDR1 or BCR. ( B ) DDR1-HEK or DDR1-K655A-HEK cells were treated with collagen I (50 μg/mL), and the cell lysates were analyzed for levels of total and phosphorylated DDR1 or BCR. ( C ) RPTECs were treated with collagen I (50 μg/mL) ± Cmp-1 as indicated and then analyzed as described in B . The black vertical line separates 2 gels that were run and developed at the same time. ( D ) pBCR and BCR bands in C were quantified by densitometry, and the pBCR level is expressed as pBCR/BCR ratio. Dara shown are mean ± SD of 1 experiment performed in triplicate ( n = 3 experiments). Statistical analysis: 1-way ANOVA followed by Dunnett’s multiple-comparison test versus untreated cells for pBCR and 2-way ANOVA followed by Sidak’s multiple-comparison test for Cmp-1–treated versus untreated cells. ( E ) Primary RPTECs isolated from WT (WT-mRPTECs) and Ddr1-KO ( Ddr1-KO -mRPTECs) mice were treated with vehicle or collagen I (50 μg/mL) for 30 minutes and then analyzed as described above. ( F ) pBCR and BCR bands were quantified using the software provided by Odyssey CLx imaging system. Circles represent cells isolated from a single mouse and values represent mean ± SD n = 5 mice for each group. Statistical analysis: 1-way ANOVA followed by Tukey’s multiple-comparison test. ( G ) Kidney cortices from uninjured (d–1), d1, and d3 injured WT and Ddr1-KO mice were analyzed by Western blot for the levels of pBCR and BCR. ( H ) pBCR and BCR bands at d3 were quantified by densitometry, and pBCR is expressed as pBCR/BCR ratio. Circles represent individual kidneys, values represent mean ± SD, WT n = 10, Ddr1-KO n = 8. Statistical analysis: 2-tailed t test.

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Plasmid Preparation, Immunoprecipitation, Comparison, Isolation, Software, Imaging, Western Blot

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: ( A ) TGF-β was measured by ELISA in conditioned medium of RPTECs treated ± collagen I (CI) ± Cmp-1 (3 μM). Circles represent a single experiment performed in triplicate. Values are mean ± SD of 3 experiments and represent fold-changes versus vehicle-treated cells assigned as 1. Statistical analysis: 1-way ANOVA followed by Dunnett’s multiple-comparison test versus CI-treated group. ( B ) RPTECs were treated ± CI ± Cmp-1 (3 μM) and then analyzed by Western blot for phosphorylated (Tyr705) and total STAT3. The black vertical line separates 2 gels that were run and developed at the same time. ( C ) pSTAT3 and STAT3 bands were quantified by densitometry and pSTAT3 is expressed as pSTAT3/STAT3 ratio. Circles and values are as in A . Statistical analysis: 1-way ANOVA followed by Tukey’s multiple-comparison test. ( D and E ) Kidney cortices from uninjured (d–1), d3, and d28 injured WT and Ddr1-KO mice were analyzed by Western blot for levels of pSTAT3 and STAT3. The black vertical line separates 2 different gels. pSTAT3/STAT3 ratio was calculated as described in C . Circles represent an individual kidney (d–1 WT n = 4, d3 WT and Ddr1-KO n = 12, d28 WT and Ddr1-KO n = 5). Values are mean ± SD and represent fold-change versus d–1 assigned as 1. Statistical analysis was performed as in C . ( F and G ) Kidney sections from uninjured (d–1), d3, and d28 injured WT and Ddr1-KO mice were stained with anti-pSTAT3 antibody and LTL. Scale bar: 20 μm. The number of pSTAT3-positive and total number of proximal tubule cells was evaluated and expressed as described in the Methods. Circles represent a single kidney. Values represent mean ± SD (d–1 WT n = 3, d3 WT n = 5, d3 Ddr1-KO n = 3, d28 WT n = 6, d28 Ddr1-KO n = 5). Statistical analysis was performed as in C .

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Enzyme-linked Immunosorbent Assay, Comparison, Western Blot, Staining

Journal: JCI Insight

Article Title: DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

doi: 10.1172/jci.insight.150887

Figure Lengend Snippet: Upregulation of DDR1 in injured RPTECs leads to a) phosphorylation of BCR, thus enabling β-catenin to regulate the production of proinflammatory MCP-1; and b) phosphorylation of STAT3 that induces the expression of the profibrotic TGF-β.

Article Snippet: For immunoblotting, membranes were incubated with the following primary antibodies from Cell Signaling Technology: DDR1 (catalog 5583), pY513-DDR1 (catalog 14531), pY792-DDR1 (catalog 11994), BCR (catalog 3902), pY177-BCR (catalog 3901), Stat3 (catalog 4904), pY705-Stat3 (catalog 9145), β-tubulin (catalog 15115), β-actin (catalog 4970), NF-κB-p65 (catalog 8242), pS536–NF-κB–p65 (catalog 3033), GAPDH (catalog 2118), N-cadherin (catalog 13116), and PARP (catalog 9532).

Techniques: Phospho-proteomics, Expressing

Journal: Scientific Reports

Article Title: Impaired retinoic acid signaling in cerebral cavernous malformations

doi: 10.1038/s41598-023-31905-0

Figure Lengend Snippet: Expression of retinoic acid pathway components is misregulated in in vitro and in vivo models of CCM. (a ) Retinoic acid synthesis and degradation pathway. Synthesis of retinoic acid from Vitamin A (retinol) is dependent on availability of NADPH. In this study, all-trans RA and the Cyp26 inhibitor Talarozole were used to modulate RA levels. ( b ) Comparative expression levels of RA pathway genes in ccm2 mutant zebrafish hearts, pan-endothelial Ccm2 knock-out mouse veins, and siRNA CCM2-depleted HUVECs. Fold changes in expression levels are depicted in circle sizes (grouped by fold changes between 0 and 5, see legend). Downregulated transcripts are represented with blue circles whereas upregulated genes are shown in red circles. Transcripts not represented in the datasets do not contain a circle. Gene names are represented as human orthologs except for genes for which only co-orthologs are known (marked with asterisks). ( c ) Whole-mount in situ hybridization of raldh2 performed on 72 hpf old zebrafish and magnifications of the heart region (see box). Stained valve leaflets are marked by an arrow.

Article Snippet: For treatment, either 0.1 µM Retinoic Acid (#R2625, Sigma) or 1–10 µM Talarozole (14531-10, Hycultec GmbH) was added and controls were supplemented with the same volume of DMSO.

Techniques: Expressing, In Vitro, In Vivo, Mutagenesis, Knock-Out, In Situ Hybridization, Staining

Journal: Scientific Reports

Article Title: Impaired retinoic acid signaling in cerebral cavernous malformations

doi: 10.1038/s41598-023-31905-0

Figure Lengend Snippet: Curative effects upon Talarozole treatment of krit1 mutant zebrafish and retinoic acid treatment of siCCM2 depleted HUVECs. (a ) Treatment scheme for zebrafish from 18 to 48 hpf using Talarozole. Endocardial cell numbers (a readout for CCM-associated cardiac phenotype in zebrafish) were analyzed after treatment in two independent experiments. ( b ) Representative images of zebrafish heart morphologies visualized by the transgene kdrl:EGFP at 48 hpf after treatment. Upper: heart morphology of controls treated with DMSO. Lower: hearts treated with Talarozole. Scale bars: 100 µm. ( c ) Endocardial cell counts at 48 hpf of zebrafish treated with several doses of Talarozole or DMSO (*P < 0.05; ****P < 0.0001 vs. control). Each data point represents one heart. ( d ) Treatment regimen of HUVECS. SiRNA transfection was done for 24 h, followed by treatment with RA 24 h later. Cell morphologies were analyzed after 48 h of incubation and antibody staining. ( e ) Immunohistochemistry of HUVECS for beta-catenin, F-actin, and DAPI. First column: control cells (siCT) with wild-type CCM2 expression. Second column: untreated siCCM2 -depleted HUVECs. Third column: representative image showing siCCM2 HUVECs treated with 100 nM RA. Scale bars: 10 µm.

Article Snippet: For treatment, either 0.1 µM Retinoic Acid (#R2625, Sigma) or 1–10 µM Talarozole (14531-10, Hycultec GmbH) was added and controls were supplemented with the same volume of DMSO.

Techniques: Mutagenesis, Transfection, Incubation, Staining, Immunohistochemistry, Expressing

Journal: Scientific Reports

Article Title: Impaired retinoic acid signaling in cerebral cavernous malformations

doi: 10.1038/s41598-023-31905-0

Figure Lengend Snippet: Assessment of KLF2/4 mRNA expression in treated animal models of CCM. (a ) Expression levels of treated BEC iCcm2 mouse cerebella after curative RA treatment (~ 20 mg/kg/day for 21 days) as mean fold changes determined by qRT-PCR. (ns = P > 0.05; **P < 0.01;***P < 0.001;****P < 0.0001 vs. control). Each data point represents one sample. Error bars show mean with SD. ( b ) Fold changes of KLF2/4 mRNA expression in siCCM2 -transfected HUVECs treated with RA for 48 h compared to untreated cells (*P < 0.05; **P < 0.01). Each data point represents one sample. ( c ) Whole-mount in situ hybridization of klf2a in 48 hpf zebrafish embryos treated with DMSO or Talarozole.

Article Snippet: For treatment, either 0.1 µM Retinoic Acid (#R2625, Sigma) or 1–10 µM Talarozole (14531-10, Hycultec GmbH) was added and controls were supplemented with the same volume of DMSO.

Techniques: Expressing, Quantitative RT-PCR, Transfection, In Situ Hybridization

Journal: PLoS ONE

Article Title: Opposite Effects of Methanandamide on Lipopolysaccharide-Induced Prostaglandin E2 and F2α Synthesis in Uterine Explants from Pregnant Mice

doi: 10.1371/journal.pone.0039532

Figure Lengend Snippet: Uterine explants were incubated without/with m-AEA (100 nM) for 24 h. A) PGE 2 production assessed by RIA; B) PGEM quantification assessed by PGE metabolite EIA kit. Student t Test. a, p<0.001 vs control; b, p<0.01 vs control. n = 5. Both metabolites were assessed in the same sample. PGEM, Prostaglandin E 2 metabolite.

Article Snippet: PGE 2 metabolite was assayed in culture supernatants with the PGE metabolite EIA kit (catalog No. 14531; Cayman Chemical Co) according to the manufacturer’s recommendations.

Techniques: Incubation, Control