Journal: PLoS ONE

Article Title: Non-Specific Protein Modifications by a Phytochemical Induce Heat Shock Response for Self-Defense

doi: 10.1371/journal.pone.0058641

Figure Lengend Snippet: (A) Hepa1c1c7 cells were treated with ZER (100 µM) for 1 or 3 hours. HSF1 phosphorylated at Ser326 in cell lysates was detected by ELISA. As a positive control, cells were exposed to heat shock (HS; incubation at 43°C in waterbath) for 15 minutes and examined. This experiment was performed in quadruplicates. (B) Cells were treated with the vehicle, ZER (Z; 10, 25, 50 µM) or geldanamycin (G; 1 µM) for 3 hours, then total RNA was subjected to qRT-PCR to semi-quantify the expressions of HSP90α, HSP90β, HSP70, and HSP40. HPRT expressions were also measured as internal standards. This experiment was performed in triplicates. a, versus CTL by Dunnett's test ( P <0.05). (C) Cells were treated with the vehicle, ZER (50 µM), or geldanamycin (GEL; 1 µM) for 6-24 hours, then lysed for western blot analysis. (D) Cells were treated with Lipofectamine™ 2000 and a siRNA solution (control and HSF1, 75 nM) for 6 hours. The culture medium was replaced with DMEM containing 10% FBS and incubated for another 24 hours. Cell lysates were subjected to western blot analysis. (E) Cells were treated with Lipofectamine™ 2000 and a siRNA solution (control and HSF1, 75 nM) for 6 hours, then the culture medium was replaced with DMEM containing 10% FBS and incubation was performed for another 24 hours. Then, siRNA-transfected cells were treated with the vehicle, ZER (50 µM), or geldanamycin (GEL; 1 µM) for 15 hours, and total RNA was subjected to qRT-PCR to semi-quantify the expression of HSP70. HPRT expression was also determined as an internal standard. This experiment was performed in quadruplicates. The various characteristics were significantly different, as shown by Tukey-Kramer test result ( P <0.05). (F) Cells were treated with the vehicle, ZER (50 µM) or tunicamycin (TUN; 1 µM) for 6 hours, then total RNA was subjected to qRT-PCR to semi-quantify the expressions of CHOP and GRP78. HPRT expressions were also measured as internal standards. This experiment was performed in triplicates. * P <0.05 vs. DMSO by Dunnett's test.

Article Snippet: After washing with PBS twice, cells were lysed, and lysates were subjected to a pSer 326 HSF1 EIA Kit according to the manufacturer's instructions (Enzo Biochem Inc., New York, NY).

Techniques: Enzyme-linked Immunosorbent Assay, Positive Control, Incubation, Quantitative RT-PCR, Western Blot, Transfection, Expressing

Journal: PLoS ONE

Article Title: Non-Specific Protein Modifications by a Phytochemical Induce Heat Shock Response for Self-Defense

doi: 10.1371/journal.pone.0058641

Figure Lengend Snippet: Xenobiotic phytochemicals like ZER are bound to numerous cellular proteins with less selectivities, which are recognized as proteostress by HSP90. Subsequently, HSF1 was dissociated from HSP90 and activated for HSPs induction.

Article Snippet: After washing with PBS twice, cells were lysed, and lysates were subjected to a pSer 326 HSF1 EIA Kit according to the manufacturer's instructions (Enzo Biochem Inc., New York, NY).

Techniques:

Journal: Frontiers in Physiology

Article Title: Mechanisms of lymphoid depletion in bowel obstruction

doi: 10.3389/fphys.2022.1005088

Figure Lengend Snippet: Osteopontin expression in the colon (A) and plasma OPN levels (B) during bowel obstruction. Rats with sham operation and bowel obstruction (BO) for 7 days were euthanized for collections of colon tissues and blood. The muscularis externa tissues of the colonic segments 2 cm proximal (Pro) and 1 cm distal (Dis) to the site of obstruction band were taken for real-time qPCR study. Left ventricle blood was taken for preparation of plasma samples and EIA measurements of OPN. N = 5 or 6. * p ≤ 0.05 vs. Sham.

Article Snippet: The levels of corticosterone and osteopontin in plasma samples were determined in enzyme immunoassay (EIA) as described ( ; ) with corticosterone and norepinephrine EIA kits (Cayman Chemical, Ann Arbor, MI) and osteopontin EIA kit (Enzo, Farmingdale, NY), respectively according to the manufacturers’ instruction.

Techniques: Expressing

Journal: Frontiers in Physiology

Article Title: Mechanisms of lymphoid depletion in bowel obstruction

doi: 10.3389/fphys.2022.1005088

Figure Lengend Snippet: Effects of bowel obstruction on specific lymphocytes in lymphoid organs in wild-type (OPN+/+) and osteopontin knock out (OPN−/−) mice. (A) Numbers of total live lymphocytes (A) , T cells (B) , CD4 + cells (C) , CD8 + cells (D) and B cells (E) in the spleen of OPN +/+ and OPN −/− mice with sham operation (Sh) and obstruction (BO, 4 days). (B) Numbers of total live lymphocytes (A) and T cells (B) in the thymus of OPN +/+ and OPN −/− mice with sham operation (Sh) and obstruction (BO, 4 days). (C) Numbers of total live lymphocytes (A) and B cells (B) in the bone marrow of OPN +/+ and OPN −/− mice with sham operation (Sh) and obstruction (BO, 4 days). N = 6 or 7. * p ≤ 0.05 vs. sham of the group. # p ≤ 0.05 vs. sham of OPN +/+ . $ p ≤ 0.05 vs. BO of OPN +/+ .

Article Snippet: The levels of corticosterone and osteopontin in plasma samples were determined in enzyme immunoassay (EIA) as described ( ; ) with corticosterone and norepinephrine EIA kits (Cayman Chemical, Ann Arbor, MI) and osteopontin EIA kit (Enzo, Farmingdale, NY), respectively according to the manufacturers’ instruction.

Techniques: Knock-Out

Journal: Frontiers in Physiology

Article Title: Mechanisms of lymphoid depletion in bowel obstruction

doi: 10.3389/fphys.2022.1005088

Figure Lengend Snippet: Osteopontin expression in the colon (A) and plasma OPN levels (B) during bowel obstruction. Rats with sham operation and bowel obstruction (BO) for 7 days were euthanized for collections of colon tissues and blood. The muscularis externa tissues of the colonic segments 2 cm proximal (Pro) and 1 cm distal (Dis) to the site of obstruction band were taken for real-time qPCR study. Left ventricle blood was taken for preparation of plasma samples and EIA measurements of OPN. N = 5 or 6. * p ≤ 0.05 vs. Sham.

Article Snippet: The levels of corticosterone and osteopontin in plasma samples were determined in enzyme immunoassay (EIA) as described ( ; ) with corticosterone and norepinephrine EIA kits (Cayman Chemical, Ann Arbor, MI) and osteopontin EIA kit (Enzo, Farmingdale, NY), respectively according to the manufacturers’ instruction.

Techniques: Expressing