Rapamycin

Manufactured by Targetmol

Sourced in United States

Rapamycin is a macrolide compound isolated from the bacterium Streptomyces hygroscopicus. It is a potent inhibitor of the mammalian target of rapamycin (mTOR) signaling pathway, which is involved in the regulation of cell growth, proliferation, and survival.

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Lab products found in correlation

Protein a g plus agarose immunoprecipitation reagent, by Santa Cruz Biotechnology (1 mentions) Anti glutathione peroxidase 4, by Abcam (1 mentions) Anti calnexin, by Proteintech (1 mentions) Anti gabarap, by Abcepta (1 mentions) Anti sqstm1 p62, by Abcam (1 mentions) Anti tom20, by Proteintech (1 mentions) Fer 1, by Targetmol (1 mentions) Pd146176, by Merck Group (1 mentions) Zileuton, by Selleck Chemicals (1 mentions) Lipofectamine ltx reagent with plus reagent, by Thermo Fisher Scientific (1 mentions) Liperfluo, by Dojindo Laboratories (1 mentions) Anti atg4b, by Proteintech (1 mentions) Ml355, by Selleck Chemicals (1 mentions) Lipoxygenase inhibitor screening assay kit, by Cayman Chemical (1 mentions) Anti flag, by Merck Group (1 mentions) Bafilomycin a1, by Merck Group (1 mentions) Normal rabbit igg, by Santa Cruz Biotechnology (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions) Anti gfp, by Abcam (1 mentions) Goat anti rabbit igg hrp, by Fudebio (1 mentions)

8 protocols using rapamycin

Ferroptosis and Autophagy Assessment

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Anti-15Lipoxygenase 1 (Abcam, ab244205), anti-Glutathione Peroxidase 4 (GPX4) (Abcam, ab125066), anti-Calnexin (Proteintech, 10427-2-AP), anti-ATG4B (CST, 13507S), anti-ATG4B (Proteintech, 15131-1-AP), anti-LC3B (CST, 2775), anti-GABARAP (Abgent, AP1821a), anti-SQSTM1/p62 (Abcam, ab56416), anti-GFP (Abcam, ab290), anti-FLAG (Sigma, F1804), anti-Tom20 (Proteintech, 11802-1-AP), anti-GAPDH (Fudebio-tech, FD0063), anti-β-actin (Fudebio-tech, FD0060), normal rabbit IgG (Santa Cruz, sc-2027), goat anti-mouse IgG-HRP (Fudebio-tech, FDM007), goat anti-rabbit IgG-HRP (Fudebio-tech, FDR007), 1S, 3R-RSL3 (TargetMol, T3646), Fer-1 (Targetmol, T6500), PD146176 (Sigma, P4620), ML355 (Selleck, S6557), Zileuton (Selleck, S1443), Rapamycin (TargetMol, T1537), Bafilomycin A1 (BaF1, Sigma, B1793) Lipofectamine™ LTX Reagent with PLUS™ Reagent (Thermo Fisher Scientific, 15338100), Protein A/G Plus-Agarose Immunoprecipitation Reagent (Santa Cruz, sc-2003), DAPI (Beyotime, C1002), LiperFluo (DOJINDO, L248), Lipoxygenase Inhibitor Screening Assay Kit (Cayman, 760700). PC (1,2-dioleoyl-sn-glycero-3-phosphocholine, Avanti, 850375), PE (1-stearoyl-2-eicasotetraenoyl-sn-glycero-3-phosphatidylethanolamine, Avanti, 850804), PE-OOH (1-stearoyl-2-15-hydroperoxy-eicasotetraenoyl-sn-glycero-3-phosphatidylethanolamine, Cayman, Cay25856-100).

Li W., Luo L.X., Zhou Q.Q., Gong H.B., Fu Y.Y., Yan C.Y., Li E., Sun J., Luo Z., Ding Z.J., Zhang Q.Y., Mu H.L., Cao Y.F., Ouyang S.H., Kurihara H., Li Y.F., Sun W.Y., Li M, & He R.R. (2022). Phospholipid peroxidation inhibits autophagy via stimulating the delipidation of oxidized LC3-PE. Redox Biology, 55, 102421.

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SARS-CoV-2 RNA Characterization Reagents

Cited 2 times

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Anti-Flag M2 (F3165, 1:5000) was from Sigma-Aldrich; anti-N (40143-R019, 1:5000) was from Sino-Biological; anti-N (ARG66782, 1:1000) was from Arigo Biolaboratories; anti-β-Actin (20536-1-AP, 1:2000) was from Proteintech Group. Anti-human GAPDH (1:5000) was prepared in our laboratory and generated by immunizing rabbits with human GAPDH protein. Naproxen^{42 (link)} (T0855), Nucleozin^{43 (link)} (T7330), (-)-Gallocatechin gallate^{44 (link)} (T3807), Sapanisertib^{9 (link)} (T1838), Rapamycin^{9 (link)} (T1537), and Silmitasertib^{9 (link)} (T2259) were from TargetMol. AB-423⁴⁵ (HY-112142) was from MedChemExpress. BAY41-4109 Racemic^{46 (link)} (S0285) was from Selleck. TMCB^{9 (link)} (B7464) was from APExBIO. (-)-Epigallocatechin gallate^{47 (link)} (E4143) and Doxycycline hyclate (D9891) were from Sigma-Aldrich. Poly(I:C) (tlrl-pic) and 5′ppp-dsRNA (tlrl-3prna) were from InvivoGen. Full-length SARS-CoV-2 3′ UTR and its complementary RNA were in vitro transcribed and labeled with HyperScribe T7 High Yield Cyanine 5 (Cy5) RNA Labeling Kit (K1062, APExBIO), and the annealed dsRNA (3′ UTR) was from the two transcribed RNAs. Cyanine 3 (Cy3)-labeled 55-nt vRNA (segment of 5′ UTR), Cy5-labeled 10-nt to 60-nt vRNA (segment of Nsp1) and 6-carboxy-fluorescein (FAM)-labeled ssDNA were generated by Tsingke Biological Technology. Sequence information is provided in Supplementary Data 3.

Zhao M., Yu Y., Sun L.M., Xing J.Q., Li T., Zhu Y., Wang M., Yu Y., Xue W., Xia T., Cai H., Han Q.Y., Yin X., Li W.H., Li A.L., Cui J., Yuan Z., Zhang R., Zhou T., Zhang X.M, & Li T. (2021). GCG inhibits SARS-CoV-2 replication by disrupting the liquid phase condensation of its nucleocapsid protein. Nature Communications, 12, 2114.

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Matrine and Fluoxetine Neuroprotective Effects

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Matrine (purity >98%) and fluoxetine were bought from Sigma and dissolved in 1% DMSO in 0.9% saline. LY294002 was purchased from Tocris, rapamycin was purchased from Target Mol, and they were dissolved in 1% DMSO in ACSF. The doses of Matrine (15, 30, 60, and 120 mg/kg), fluoxetine (20 mg/kg), LY294002 (10 nmol/site), and rapamycin (0.2 nmol/site) were chosen based on previous reports (Jiang et al., 2015 (link); Gong et al., 2016 (link); Ludka et al., 2016 (link); Jiang et al., 2017 (link)). Matrine and fluoxetine were i.p. injected in a volume of 10 mL/kg. LY294002 and rapamycin were intracerebroventricularly (i.c.v.) infused.

Wu Z., You Z., Chen P., Chen C., Chen F., Shen J, & Xu H. (2018). Matrine Exerts Antidepressant-Like Effects on Mice: Role of the Hippocampal PI3K/Akt/mTOR Signaling. International Journal of Neuropsychopharmacology, 21(8), 764-776.

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Antibodies for Flow Cytometry Analysis

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The antibodies used for flow cytometric analyses are listed in Supplementary Table S1. Purified Fc Block (anti-mouse FcRγII/III monoclonal antibody) and isotype control antibodies were obtained from eBioscience. Rapamycin and STAT3 inhibitor were purchased from TargetMol (China). Carboxyfluorescein diacetate succinimidyl ester (CFSE) was purchased from Invitrogen (United States). L-NG-Monomethylarginine, Acetate Salt (L-NMMA) and N(omega)-hydroxy-nor-L-arginine (nor-NOHA) were purchased from MCE (United States). Recombinant murine IL-2 was obtained from PeproTech (United States).

Li X., Li Y., Yu Q., Xu L., Fu S., Wei C., Wang L., Luo Y., Shi J., Qian P., Huang H, & Lin Y. (2021). mTOR Signaling Regulates the Development and Therapeutic Efficacy of PMN-MDSCs in Acute GVHD. Frontiers in Cell and Developmental Biology, 9, 741911.

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Modulation of Autophagy and mTORC1 in ZNP Studies

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3-Benzyl-5-(2-nitrophenoxymethyl)-γ-butyrolactone derivative (3BDO; TargetMol, USA) is an inhibitor of autophagy and an activator of mTORC1 [19] [20] [21] . Rapamycin (RAPA; TargetMol, USA) is an activator of autophagy and an inhibitor of mTORC1 [22] [23] [24] . These reagents were diluted in DMSO. In vivo, phosphate buffer solution-diluted 3BDO was applied to the skin surface at 6 h before ZNP application once daily. In vitro, cells were pretreated with 3BDO and RAPA for 3 h before ZNP treatment. An equal volume of DMSO was added as the negative control.

Lai X., Wang M., Zhang Z., Chen S., Tan X., Liu W., Liang H., Li L, & Shao L. (2024). ZNPs reduce epidermal mechanical strain resistance by promoting desmosomal cadherin endocytosis via mTORC1-TFEB-BLOC1S3 axis. Journal of nanobiotechnology, 22(1).

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Regulation of VSMC Autophagy by UA

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Primary VSMCs (HA-VSMC line) and culture medium 231 were purchased from Cascade Biologics (Portland, OR, USA). Cells were cultured with different concentrations of UA for 48 h. Rapamycin (TargetMol, Boston, MA, USA) and MHY1485 were used to promote and inhibit autophagy, respectively. To knock out KLF4 in HA-VSMCs, the cells were transfected with an anti-KLF4 siRNA using Lipofectamine 2000 (Invitrogen, Waltham, MA, USA), according to the manufacturer's instructions.

Lu Y., Zhang H., Han M., Wang P, & Meng L. (2024). Impairment of Autophagy Mediates the Uric-Acid-Induced Phenotypic Transformation of Vascular Smooth Muscle Cells. Pharmacology, 109(1).

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Immune Cell Isolation and Culture

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The following reagents and drugs were used in this study: RPMI-1640 medium (Hyclone, SH30809.01B); collagenase IV (Gibco, 17104-019), PBS (Solebao, P1020); bovine serum albumin (BSA, Hyclone, SH30574.03); 1% Triton x-100 (Senbega, SBJ-1141); double antibody (Beijing Xia Si Biotechnology Co., Ltd., SV30010); 0.25% Trypsin (Gibco, 15050065); rapamycin (Targetmol, T1537); hemin (Sigma, 51280-1 G); γ-IFN (Sigma, I4777); FBS (Gibco, 16000-044).

Yuan S., Liang X., He W., Liang M., Jin J, & He Q. (2021). ATF4-dependent heme-oxygenase-1 attenuates diabetic nephropathy by inducing autophagy and inhibiting apoptosis in podocyte. Renal Failure, 43(1), 968-979.

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Generating Engineered T Cells for Therapy

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Peripheral blood mononuclear cells (PBMC) were separated by density gradient centrifugation, and T cells were separated using a CD3 T-cell Separation Kit (Miltenyi Biotec). On day 0, T cells were resuspended in X-VIVO-15 Medium (Lonza) supplemented with 5% Human AB Serum (GEMINI), 100 U/mL penicillin, 100 μg/mL streptomycin (Solarbio), and 2 mmol/L GlutaMAX (Gibco) at a concentration of 5 × 10⁵ cells/mL, and anti-CD3/CD28 Dynabeads (Thermo Fisher Scientific) were added at a 1: 1 ratio; at the same time, IL2 was added at a final concentration of 100 U/mL and then supplemented every 2 days. After 24 hours, concentrated lentivirus (MOI = 50) and polybrene (Sigma) with a final concentration of 8 μg/mL were added to activated T cells, and were centrifuged at 800 × g for 1 hour at 32°C. The medium was changed after 6–8 hours and rapamycin (Sigma) was added at a final concentration of 20 nmol/L, everolimus (TargetMol, 20 nmol/L), temsirolimus (TargetMol, 20 nmol/L), JR-AB2–011 (TargetMol, 1μmol/L), KU0063794 (TargetMol, 20 nmol/L); rapamycin was supplemented every 2 days. The cell culture was monitored daily and the medium was added to maintain a cell concentration of 0.5–1 × 10⁶ cells/mL. On day 4, the Dynabeads were removed. The CAR-T cells were harvested after day 6 for analysis or in vivo experiments.

Nian Z., Zheng X., Dou Y., Du X., Zhou L., Fu B., Sun R., Tian Z, & Wei H. (2021). Rapamycin Pretreatment Rescues the Bone Marrow AML Cell Elimination Capacity of CAR-T Cells. Clinical Cancer Research, 27(21), 6026-6038.

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