Journal: Transplant international : official journal of the European Society for Organ Transplantation

Article Title: Intracellular sirolimus concentration is reduced by tacrolimus in human pancreatic islets in vitro.

doi: 10.1111/tri.12617

Figure Lengend Snippet: Figure 1 Intracellular concentrations of TAC in human islets. Human islets were cultured with TAC (10 or 30 lg/l), SRL (10 or 30 lg/l), or the combi- nation thereof for 24–48 h before the intracellular concentration of TAC was measured in islet lysate and normalized to total protein as detailed in methods. Data are presented as the mean SD, n = 6 for each group. TAC, tacrolimus; SRL, sirolimus; *P < 0.04; ** P < 0.007; *** P < 0.0006.

Article Snippet: The islets were exposed to 10 and 30 lg/l of tacrolimus (Santa Cruz Biotechnology, Dallas, TX, USA) or sirolimus (Toronto Research Chemicals, Toronto, Ontario, Canada) or the combination thereof for 24 h and 48 h at 37 °C (5% CO2).

Techniques: Cell Culture, Concentration Assay

Journal: Transplant international : official journal of the European Society for Organ Transplantation

Article Title: Intracellular sirolimus concentration is reduced by tacrolimus in human pancreatic islets in vitro.

doi: 10.1111/tri.12617

Figure Lengend Snippet: Figure 2 Intracellular concentrations of SRL in human islets. Human islets were cultured with TAC (10 or 30 lg/l), SRL (10 or 30 lg/l), or the combi- nation thereof for 24–48 h before the intracellular concentration of SRL was measured in islet lysate and normalized to total protein as detailed in methods. Data are presented as the mean SD, n = 6 for each group. TAC: tacrolimus; SRL: sirolimus; *** P < 0.001; **** P < 0.0001.

Article Snippet: The islets were exposed to 10 and 30 lg/l of tacrolimus (Santa Cruz Biotechnology, Dallas, TX, USA) or sirolimus (Toronto Research Chemicals, Toronto, Ontario, Canada) or the combination thereof for 24 h and 48 h at 37 °C (5% CO2).

Techniques: Cell Culture, Concentration Assay

Journal: Transplant international : official journal of the European Society for Organ Transplantation

Article Title: Intracellular sirolimus concentration is reduced by tacrolimus in human pancreatic islets in vitro.

doi: 10.1111/tri.12617

Figure Lengend Snippet: Figure 3 Effect of CsA on intracellular concentration of SRL in human islets. Human islets were cultured with the combination of SRL (30 lg/l) and CsA (5 lg/ml), or the drug alone for 24 h before the intracellular concentration of SRL (a) or CsA (b) was measured in islet lysate and nor- malized to total protein as detailed in methods. Data are calculated as percentages of control and are presented as mean SD, n = 6 for each group. CsA, cyclosporine A; SRL, sirolimus.

Article Snippet: The islets were exposed to 10 and 30 lg/l of tacrolimus (Santa Cruz Biotechnology, Dallas, TX, USA) or sirolimus (Toronto Research Chemicals, Toronto, Ontario, Canada) or the combination thereof for 24 h and 48 h at 37 °C (5% CO2).

Techniques: Concentration Assay, Cell Culture, Control

Journal: Transplant international : official journal of the European Society for Organ Transplantation

Article Title: Intracellular sirolimus concentration is reduced by tacrolimus in human pancreatic islets in vitro.

doi: 10.1111/tri.12617

Figure Lengend Snippet: Figure 4 The effect of SRL, TAC, or CsA on phosphorylation of p70S6k in islets. Human islets were cultured with TAC (30 lg/l), SRL (30 lg/l), or the combination thereof for 24 h before the presence of p- p70s6k was assessed by the cell-signaling Bio-Plex assay in human islet cell lysate and normalized to total protein (a). In a parallel experiment, human islets were cultured with SRL (30 lg/l), CsA (5 lg/ml), or the combination thereof for 24 h before p-p70S6k was detected in the lysate and normalized to total protein. Data are calculated as ratio to control and are presented as mean SD, n = 3–6 for each group. TAC, tacrolimus; SRL, sirolimus; CsA, cyclosporine A, ** P < 0.01, **** P < 0.0001.

Article Snippet: The islets were exposed to 10 and 30 lg/l of tacrolimus (Santa Cruz Biotechnology, Dallas, TX, USA) or sirolimus (Toronto Research Chemicals, Toronto, Ontario, Canada) or the combination thereof for 24 h and 48 h at 37 °C (5% CO2).

Techniques: Phospho-proteomics, Cell Culture, Plex Assay, Control

Journal: Transplant international : official journal of the European Society for Organ Transplantation

Article Title: Intracellular sirolimus concentration is reduced by tacrolimus in human pancreatic islets in vitro.

doi: 10.1111/tri.12617

Figure Lengend Snippet: Figure 5 Oxygen consumption rates (OCR) in human islets after treatment of TAC, SIR, or SIR+TAC. Human islets were cultured with TAC (30 lg/l), SIR (30 lg/l), or a combination thereof for 24 h before the glucose-stimulated OCR was measured as indicated in methods. OCR is expressed as percentage of baseline and is presented as the mean SD, n = 6 for each group. TAC, tacrolimus; SRL, sirolimus, ** P < 0.01.

Article Snippet: The islets were exposed to 10 and 30 lg/l of tacrolimus (Santa Cruz Biotechnology, Dallas, TX, USA) or sirolimus (Toronto Research Chemicals, Toronto, Ontario, Canada) or the combination thereof for 24 h and 48 h at 37 °C (5% CO2).

Techniques: Cell Culture

Journal: Transplant international : official journal of the European Society for Organ Transplantation

Article Title: Intracellular sirolimus concentration is reduced by tacrolimus in human pancreatic islets in vitro.

doi: 10.1111/tri.12617

Figure Lengend Snippet: Figure 6 Expression of ABCB1 (Pgp), OATP1B1, and CYP3A4 in human islets. Human islets were cultured for 24 h before the expression of the drug transporter (ABCB1(Pgp) and OATP1B1), and the metabolic enzyme CYP3A4 was evaluated. RNA was prepared and subjected to qPCR as detailed in methods. The reference gene index is calculated by the mean of ALAS1, B2M, and RPL13A expression and used to normalize the expression of target genes in isolated hepatocytes relative to the mRNA level of ABCB1(Pgp), OATP1B1, and CYP3A4 in human islets (a). OATB1 mRNA expression in human islets was normalized to reference gene index after exposure to TAC (30 lg/l), SRL (30 lg/l), or a combination thereof for 24 h (b). Represen- tative immunofluorescence image of dispersed human islets stained for insulin (green), ABCB1(Pgp) (red) and nuclear staining with DAPI (blue) (c), or glucagon (green), ABCB1(Pgp) (red) and nuclear staining wit DAPI (blue) (d). Data are presented as mean SD, n = 4–5 for each group. TAC, tacroli- mus; SRL, sirolimus; *P < 0.05; ** P < 0.01.

Article Snippet: The islets were exposed to 10 and 30 lg/l of tacrolimus (Santa Cruz Biotechnology, Dallas, TX, USA) or sirolimus (Toronto Research Chemicals, Toronto, Ontario, Canada) or the combination thereof for 24 h and 48 h at 37 °C (5% CO2).

Techniques: Expressing, Cell Culture, Isolation, Staining