Journal: iScience

Article Title: Therapy-induced normal tissue damage promotes breast cancer metastasis

doi: 10.1016/j.isci.2023.108503

Figure Lengend Snippet:

Article Snippet: Venetoclax , Selleckchem , Cat#S8048.

Techniques: Luciferase, Virus, Plasmid Preparation, Recombinant, Staining, Ab Array, Generated, Gene Expression, Software

Journal: Journal of Virology

Article Title: HIV-1 Nef activates proviral DNA transcription by recruiting Src kinase to phosphorylate host protein Nef-associated factor 1 to compromise its viral restrictive function

doi: 10.1128/jvi.00280-25

Figure Lengend Snippet: Nef recruits Src and promotes HIV-1 proviral DNA transcription by stimulating PI3K/AKT/mTOCR1 pathways. ( A, B ) Nef promotes HIV-1 replication. PHA-P-activated CD4 + T cells (10 6 cells) were infected with replication-competent HIV-1 NL4-3 (WT) or Nef-deficient (ΔNef) mutant virus (50 ng p24 gag amounts of viruses) ( A ), or with pseudotyped single-cycle infectious HIV-H131(ΔNef)/VSV-G or HIV-H132 (Nef)/VSV-G (5 ng p24 gag amounts of viruses) ( B ), for 5 days. Viral replication was detected by measuring the production of cell-associated gag mRNA. ( C ) Nef increases LTR-driven gene expression. Nef-expressing plasmid pCDH-CMV-MCS-EF1-Puro-Nef, HIV-1- tat expressing plasmid pRK-Flag/tat, and a luciferase reporter driven by the full-length LTR promoter derived from HIV-1 NL4-3 were co-transfected into HEK293T cells, and β-Gal-expressing vector was used to normalize transfection efficiency. At 24 h post-transfection, cells were harvested, and the reporter gene expressions were assessed. ( D, E ) Nef promotes the transcription of HIV-1 proviral DNA. C11 cells were transduced with lentiviruses (5 ng p24 gag amounts of viruses) containing HIV-1 Nef- or mutant-expressing plasmid or vector control for 2 days; the transcription of HIV-1 proviral DNA was measured by detecting GFP expression ( D ) or measuring the production of cell-associated gag mRNA ( E ). ( F ) The illumination of Nef motif and mutants. ( G through J ) Nef activates the PI3K/AKT/mTORC1 pathway. C11 cells were transfected with pCDH-CMV-MCS-EF1-Puro-Nef (0.2, 1, or 2 ng plasmids in “G”; 1 ng plasmids in “H” and “I"), mutant-expressing plasmid, or a vector control for 2 days. The expression and phosphorylation of PI3K p85 subunit, AKT, mTORC1, P70S6K S6, and CDK9 were detected by western blotting. One representative from at least five repeats is shown. The Image J software was used to calculate the gray intensity of western blotting strips, and the relative values were labeled below ( G through I ). ** P < 0.01 and *** P < 0.001 are considered significant differences determined by an unpaired t test.

Article Snippet: Endogenous Naf1 was detected with a mouse mAb at a dilution of 1:1,000 , and the following other antibodies were used: anti-Flag tag mouse (M20008; Abmart Inc., Shanghai, China), anti-GAPDH (clone 3B3) ( M20006 ; Abmart Inc., Shanghai, China), anti-Src Rabbit mAb (2109, Cell Signaling Technology), anti-phosphorylation-Src family (Tyr416) rabbit mAb (6943, Cell Signaling Technology), anti-PI3K kinase p85 subunit (4292; Cell Signaling Technology), anti-PI3K phosphorylation p85 Tyr458/p55 Tyr 199 (4228; Cell Signaling Technology), anti-AKT (pan) (C67E7) (4691S; Cell Signaling Technology), anti-AKT phosphorylation Thr308 (D25E6) (13038; Cell Signaling Technology), anti-mTORC1 (ab32028; Abcam), anti-mTORC1 phosphorylation S2448 (ab109268; Abcam), anti-p70 S6 kinase (9202; Cell Signaling Technology), anti-p70 S6 kinase phosphor Thr389 (9205; Cell Signaling Technology), anti-S6 ribosomal protein(5G10) (2217; Cell Signaling Technology), anti-S6 ribosomal protein phosphorylation Ser235/236 (D57.2.2E) (4858; Cell Signaling Technology), anti-myc tag (ab9106; Abcam), anti-Cyclin-dependent protein kinase 9 (anti-CDK9)(2316; Cell Signaling Technology), anti-phosphorylation-CDK9 (Thr186) (2549; Cell Signaling Technology), and anti-Cyclin T1 (81464; Cell Signaling Technology).

Techniques: Infection, Mutagenesis, Virus, Gene Expression, Expressing, Plasmid Preparation, Luciferase, Derivative Assay, Transfection, Transduction, Control, Phospho-proteomics, Western Blot, Software, Labeling

Journal: Journal of Virology

Article Title: HIV-1 Nef activates proviral DNA transcription by recruiting Src kinase to phosphorylate host protein Nef-associated factor 1 to compromise its viral restrictive function

doi: 10.1128/jvi.00280-25

Figure Lengend Snippet: Naf1 is required for Nef-induced transcription of HIV proviral DNA. ( A ) C11 cells were transduced with lentiviruses containing Naf1 shRNA or the scramble control for 48 h and further transduced with pCDH-CMV-MCS-EF1-Puro-Nef or a vector control for an additional 48 h. HIV proviral DNA transcription was measured by detecting GFP expression, and the protein expression and Nef-induced phosphorylation of PI3K p85 subunit, AKT, mTORC1, and CDK9 were detected by western blotting (B). ( C–E ) Naf1 suppresses the PI3K/AKT/mTOCR1 pathway. C11 cells were transfected with pCMV-Tag3B-myc-Naf1 (0.2, 1, or 2 ng plasmids) or a vector control ( C ), or transduced with lentiviruses containing Naf1 shRNA or the scramble control ( D ), for 48 h. Western blotting was performed to detect the protein expressions and the phosphorylation levels of p85, AKT, mTORC1, P70S6K, and CDK9 ( C, D ). ( E ) LTR-driven gene expression. HEK293T cells were transduced with lentiviruses containing Naf1 shRNA or scramble control for 48 h, then were further transfected with pCDH-CMV-MCS-EF1-Puro-Nef, pRK-Flag/tat, and a luciferase reporter driven by HIV-1 NL4-3 -LTR for 24 h, and the reporter gene expressions were assessed. ( F ) Viral replication. PHA-P-activated CD4 + T cells (10 6 cells) were transduced with lentiviruses containing Naf1 shRNA or scramble control for 2 days, then were further infected with HIV-1 NL4-3 (WT) or Nef-deficient (ΔNef) mutant virus (50 ng p24 gag amounts of viruses) for 5 days. Viral replication was detected by measuring the p24 gag in cell supernatant. One representative from four repeats is shown. The Image J software was used to calculate the gray intensity of western blotting strips, and the relative values were labeled below ( B–D ). ** P < 0.01 and *** P < 0.001 are considered significant differences determined by an unpaired t test.

Article Snippet: Endogenous Naf1 was detected with a mouse mAb at a dilution of 1:1,000 , and the following other antibodies were used: anti-Flag tag mouse (M20008; Abmart Inc., Shanghai, China), anti-GAPDH (clone 3B3) ( M20006 ; Abmart Inc., Shanghai, China), anti-Src Rabbit mAb (2109, Cell Signaling Technology), anti-phosphorylation-Src family (Tyr416) rabbit mAb (6943, Cell Signaling Technology), anti-PI3K kinase p85 subunit (4292; Cell Signaling Technology), anti-PI3K phosphorylation p85 Tyr458/p55 Tyr 199 (4228; Cell Signaling Technology), anti-AKT (pan) (C67E7) (4691S; Cell Signaling Technology), anti-AKT phosphorylation Thr308 (D25E6) (13038; Cell Signaling Technology), anti-mTORC1 (ab32028; Abcam), anti-mTORC1 phosphorylation S2448 (ab109268; Abcam), anti-p70 S6 kinase (9202; Cell Signaling Technology), anti-p70 S6 kinase phosphor Thr389 (9205; Cell Signaling Technology), anti-S6 ribosomal protein(5G10) (2217; Cell Signaling Technology), anti-S6 ribosomal protein phosphorylation Ser235/236 (D57.2.2E) (4858; Cell Signaling Technology), anti-myc tag (ab9106; Abcam), anti-Cyclin-dependent protein kinase 9 (anti-CDK9)(2316; Cell Signaling Technology), anti-phosphorylation-CDK9 (Thr186) (2549; Cell Signaling Technology), and anti-Cyclin T1 (81464; Cell Signaling Technology).

Techniques: Transduction, shRNA, Control, Plasmid Preparation, Expressing, Phospho-proteomics, Western Blot, Transfection, Gene Expression, Luciferase, Infection, Mutagenesis, Virus, Software, Labeling

Journal: Journal of Virology

Article Title: HIV-1 Nef activates proviral DNA transcription by recruiting Src kinase to phosphorylate host protein Nef-associated factor 1 to compromise its viral restrictive function

doi: 10.1128/jvi.00280-25

Figure Lengend Snippet: Naf1-Y552 phosphorylation activates PI3K/AKT/mTORC1 signaling and compromises Naf1's inhibitor roles in HIV proviral DNA transcription. HEK293T cells were co-transfected with pCMV-Tag3B-myc-Naf1 (or Naf1 mutant plasmids) and p85 plasmid. Immunoprecipitations were performed with anti-myc antibody, and p85 was immunoblotted with a specific antibody ( A ), and western blotting was performed to detect the expression and phosphorylation levels of Naf1, p85 subunit, AKT, mTORC1, and CDK9 ( B ). ( C ) Naf1-Y552D induces transcription of HIV proviral DNA. C11 cells were transduced with lentiviruses containing WT Naf1-expressing plasmid pCDH-CMV-MCS-EF1-Puro/Naf1, Y552D, or Y552F mutant-expressing plasmid. The transcription of HIV proviral DNA was measured either by detecting GFP expression or quantifying the production of cell-associated gag mRNA. The results from five repeats ( n = 5) are summarized. Mean fluorescence intensity (MFI) was calculated. ( D ) The association of Cyclin T1 with HIV-1 5`-LTR in C11 or ACH2 cells was determined by a cross-linked ChIP assay. ( E ) Nef compromises Naf1's inhibitory roles of HIV-1 replication. Jurkat CD4 + T cells were transfected with pCMV-Tag3B-myc-Naf1 or vector and then infected with HIV-1 NL4-3 or ΔNef mutant virus for 48 h. Viral replication was detected by measuring the production of cellular gag mRNA. ( F ) Nef compromises Naf1's inhibitory roles of LTR activation. HEK293T cells were transfected with pCMV-Tag3B-myc-Naf1, pCDH-CMV-MCS-EF1-Puro-Nef, pRK-Flag/tat, and a luciferase reporter driven by HIV-1 NL4-3 -LTR, for 24 h, and the reporter gene expressions were assessed. One representative from four repeats is shown. * P < 0.05, ** P < 0.01, and *** P < 0.001 denote the significant difference determined by an unpaired t test.

Article Snippet: Endogenous Naf1 was detected with a mouse mAb at a dilution of 1:1,000 , and the following other antibodies were used: anti-Flag tag mouse (M20008; Abmart Inc., Shanghai, China), anti-GAPDH (clone 3B3) ( M20006 ; Abmart Inc., Shanghai, China), anti-Src Rabbit mAb (2109, Cell Signaling Technology), anti-phosphorylation-Src family (Tyr416) rabbit mAb (6943, Cell Signaling Technology), anti-PI3K kinase p85 subunit (4292; Cell Signaling Technology), anti-PI3K phosphorylation p85 Tyr458/p55 Tyr 199 (4228; Cell Signaling Technology), anti-AKT (pan) (C67E7) (4691S; Cell Signaling Technology), anti-AKT phosphorylation Thr308 (D25E6) (13038; Cell Signaling Technology), anti-mTORC1 (ab32028; Abcam), anti-mTORC1 phosphorylation S2448 (ab109268; Abcam), anti-p70 S6 kinase (9202; Cell Signaling Technology), anti-p70 S6 kinase phosphor Thr389 (9205; Cell Signaling Technology), anti-S6 ribosomal protein(5G10) (2217; Cell Signaling Technology), anti-S6 ribosomal protein phosphorylation Ser235/236 (D57.2.2E) (4858; Cell Signaling Technology), anti-myc tag (ab9106; Abcam), anti-Cyclin-dependent protein kinase 9 (anti-CDK9)(2316; Cell Signaling Technology), anti-phosphorylation-CDK9 (Thr186) (2549; Cell Signaling Technology), and anti-Cyclin T1 (81464; Cell Signaling Technology).

Techniques: Phospho-proteomics, Transfection, Mutagenesis, Plasmid Preparation, Western Blot, Expressing, Transduction, Fluorescence, Infection, Virus, Activation Assay, Luciferase

Journal: Journal of the Anus, Rectum and Colon

Article Title: Evaluation of the Amounts of Sennosides A and B in Rhubarb-containing Kampo Medicines to Create a Ranking of Kampo Medicines for Appropriate Selection of Laxatives

doi: 10.23922/jarc.2020-102

Figure Lengend Snippet: Comparison of the Estimated and Analysed Strength Scores of Tsumura’s Medicinal Rhubarb-containing Formulas Based on Their Daiokanzoto.

Article Snippet: 1.99 , Tsumura-134 , Keishikashakuyakudaioto.

Techniques: Comparison

Journal: Surgical Endoscopy

Article Title: Evaluation of a novel home-based laparoscopic and core surgical skills programme (Monash Online Surgical Training)

doi: 10.1007/s00464-023-10669-8

Figure Lengend Snippet: The eoSim SurgTrac Core laparoscopic simulation bench trainer (eoSurgical Ltd, Scotland, UK) used with an iPad Tablet (Apple Inc, Cupertino, California, USA)

Article Snippet: Fig. 2 Example of the feedback template for the assessment task and metrics on SurgTrac, version 1.9.9, eoSurgical, Scotland UK

Techniques:

Journal: Surgical Endoscopy

Article Title: Evaluation of a novel home-based laparoscopic and core surgical skills programme (Monash Online Surgical Training)

doi: 10.1007/s00464-023-10669-8

Figure Lengend Snippet: Example of the feedback template for the assessment task and metrics on SurgTrac, version 1.9.9, eoSurgical, Scotland UK

Article Snippet: Fig. 2 Example of the feedback template for the assessment task and metrics on SurgTrac, version 1.9.9, eoSurgical, Scotland UK

Techniques:

Journal: Surgical Endoscopy

Article Title: Evaluation of a novel home-based laparoscopic and core surgical skills programme (Monash Online Surgical Training)

doi: 10.1007/s00464-023-10669-8

Figure Lengend Snippet: Major themes obtained from the thematic analysis of participants written feedback, with illustrative direct participant comment

Article Snippet: Fig. 2 Example of the feedback template for the assessment task and metrics on SurgTrac, version 1.9.9, eoSurgical, Scotland UK

Techniques: