rapamycin signaling pathway  (Musashi Engineering Inc)

Journal: Molecular Therapy. Nucleic Acids

Article Title: Potential effects of endogenous RNA/DNA hybrids on CRISPR-Cas9-mediated homology-directed repair

doi: 10.1016/j.omtn.2026.102880

Figure Lengend Snippet: Modulation of HDR efficiency by R-loop resolution, cell-cycle perturbation, and locus context (A) ddPCR quantification of HDR events in HEPA1-6/SpCas9 cells transiently overexpressing human RNase H1 and transduced with 10,000 vg/cell of scAAVDJ-sgMid Alb + 10,000 vg/cell ssAAVDJ-donor Mid Alb, or 10,000 vg/cell of scAAVDJ-sg3′ Alb + 10,000 vg/cell ssAAVDJ-donor 3′ Alb. (B) Schematic representation of cell cycle profiles in HEPA1-6 cells and murine hepatocytes. (C) Flow cytometry analysis of HEPA1-6/SpCas9 cells treated for 24 h with rapamycin (1 μM, 5 μM) or palbociclib (PD; 5 μM, 10 μM). DMSO-treated cells served as controls. (D) DRIP-qPCR analysis of HEPA1-6/SpCas9 cells after 24 h treatment with 5 μM rapamycin; DMSO-treated cells served as controls. (E) ddPCR detection of HDR events in HEPA1-6/SpCas9 cells pre-treated with 5 μM rapamycin for 24 h, followed by 72 h transduction with 10,000 vg/cell of scAAVDJ-sgMid Alb + 10,000 vg/cell ssAAVDJ-donor Mid Alb, or scAAVDJ-sg3′ Alb + ssAAVDJ-donor 3′ Alb. (F) IGV visualization of the Alb and Actb loci showing gRNA (sgActb) and donor (donor Actb) design targeting the 5′ region of Actb . (G) ddPCR detection of HDR events in HEPA1-6/SpCas9 cells transduced for 72 h with 10,000 vg/cell of scAAVDJ-sgActb + 10,000 vg/cell ssAAVDJ-donor Actb. Scrambled gRNA with donor served as negative control. (H) ddPCR detection of HDR events in HEPA1-6/SpCas9 cells pre-treated with 5 μM rapamycin for 24 h, followed by 72 h transduction with scAAVDJ-sgActb + ssAAVDJ-donor Actb. (I) ddPCR detection of HDR events in HEPA1-6/SpCas9 cells transiently overexpressing human RNase H1 and transduced with scAAVDJ-sgActb + ssAAVDJ-donor Actb. (J) DRIP-qPCR analysis of CD4 + T cells from a healthy donor, pre- and post-activation with 10 μg/mL PHA, 50 IU/mL IL-2, 5 ng/mL IL-7, and 5 ng/mL IL-15. DRIP, immunoprecipitated samples treated with S9.6 antibody which are enriched in R loops; RNAseH + , samples treated with RNAseH1+S9.9 antibody which are depleted of R-loops. (K) Table summarizing the effects of R-loops levels on indels, HDR, and AAV integration. N/A, not affected; ↑ increase; ↓ = decrease. Statistical analysis: (A and E) multiple t test; (C) two-way ANOVA with Dunnett’s post hoc test; (G–I) Student’s t test. p < 0.05, ∗ p < 0.01, ∗∗ p < 0.001, ∗∗∗ p < 0.0001. Error bars represent mean ± SD.

Article Snippet: HEPA1-6/SpCas9 cells were treated with rapamycin (Selleck, S1039) or palbociclib (Selleck, S1116) for 24 h, fixed in 70% ethanol, stained with propidium iodide, and analyzed by flow cytometry (NovoCyte Penteon, Agilent).

Techniques: Transduction, Flow Cytometry, Negative Control, Activation Assay, Immunoprecipitation

Journal: Neurobiology of Stress

Article Title: USP11 drives stress-induced synaptic structural deficits and depression-like behaviors through GSK3β/mTOR signaling

doi: 10.1016/j.ynstr.2026.100791

Figure Lengend Snippet: USP11 knockout alleviates stress-induced depressive-like behaviors and associated with mTOR Signaling (A) Western blot analysis of USP11 (110 kDa), p-mTOR (Ser2448, 289 kDa), total mTOR (289 kDa), p-GSK3β (Ser9, 47 kDa), total GSK3β (47 kDa), PSD95 (95 kDa), and Tubulin (55 kDa) in mouse mPFC from wild-type (WT) and USP11 knockout (USP11 −/− ) male mice (n = 6, Tubulin as loading control). (B–E) Quantification of baseline protein band intensity in wild-type control (WT-CON) and USP11 knockout control (KO-CON) groups: (B) USP11 (relative to Tubulin, p < 0.0001), (C) p-GSK3β (relative to total GSK3β, p = 0.0072), (D) p-mTOR (relative to total mTOR, p = 0.0028), (E) PSD95 (relative to Tubulin, p = 0.0159). n = 6/group. (F–I) Behavioral results for four groups: WT-CON, KO-CON, WT-CUMS, and KO-CUMS (OFT, distance [cm], F [3, 28] = 8.234, p = 0.0004; OFT, velocity [cm/s], F [3, 28] = 8.233, p = 0.0004; FST, F [3, 28] = 8.721, p = 0.0003; TST, F [3, 29] = 5.378, p = 0.0046). n = 8/group. (J) Western blot analysis of USP11 (110 kDa), p-mTOR (Ser2448, 289 kDa), total mTOR (289 kDa), SYN (synaptophysin, 77 kDa), and Tubulin (55 kDa) in mPFC from all four groups (n = 3). (K-M) Quantification of (K) USP11 (relative to Tubulin, F (3, 8) = 139.5, p < 0.0001), (L) p-mTOR (relative to total mTOR, F (3, 8) = 8.298, p = 0.0077), (M) SYN (relative to Tubulin, F (3, 8) = 8.811, p = 0.0065). n = 3/group. (N) Schematic overview of the experimental design, including a 7-day acclimation period, a 28-day chronic unpredictable mild stress (CUMS) procedure, the rapamycin dosing regimen (3 mg/kg, i.p., three times per week; from day 14 of CUMS until 24 h before tissue collection), and the behavioral test battery in male USP11 −/− mice. (O-R) Behavioral results for three groups in USP11 −/− mice: CON + Veh, CUMS + Veh and CUMS + Rapa. (SPT, F (2, 18) = 7.019, p = 0.0056; OFT, center time [s], F [2, 18] = 8.788, p = 0.0022; OFT, velocity [cm/s], F [2, 18] = 0.09090, p = 0.9135; TST, F [2, 18] = 7.797, p = 0.0036). n = 7/group.) (T) Quantification of p-mTOR (relative to total mTOR, F (2, 6) = 38.49, p = 0.0004) Data are shown as mean ± SEM. Determined by t -test (baseline comparisons) or one-way ANOVA (multiple groups) unless otherwise indicated. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. (S) Representative immunoblots of p-mTOR (Ser2448, 289 kDa), total mTOR (289 kDa) in USP11 −/− mice under the indicated conditions. (n = 3, Tubulin as loading control).

Article Snippet: Rapamycin (MedChemExpress, Cat. No. 53123-88-9) was formulated in a mixed vehicle containing 10% DMSO, 40% PEG300, 5% Tween-80, and 45% sterile 0.9% saline and prepared fresh before injection( ).

Techniques: Knock-Out, Western Blot, Control, Battery

Journal: JAAD Case Reports

Article Title: Treatment of extramammary Paget’s disease with sirolimus: A case series

doi: 10.1016/j.jdcr.2025.10.021

Figure Lengend Snippet: Clinical images of patient 1 at baseline (A) , 8 wks on sirolimus twice daily (B) , and 16 wks on sirolimus twice daily (C) .

Article Snippet: While the pathogenesis of EMPD is not completely understood, the Musashi-1- mammalian target of rapamycin signaling pathway is thought to play a role in disease development.

Techniques:

Journal: JAAD Case Reports

Article Title: Treatment of extramammary Paget’s disease with sirolimus: A case series

doi: 10.1016/j.jdcr.2025.10.021

Figure Lengend Snippet: Clinical images of patient 3 at baseline (A) , 4 wks on sirolimus twice daily (B) , and 8 wks on sirolimus twice daily (C) .

Article Snippet: While the pathogenesis of EMPD is not completely understood, the Musashi-1- mammalian target of rapamycin signaling pathway is thought to play a role in disease development.

Techniques: