Journal: Cell Death Discovery

Article Title: Trifluoperazine causes mast cell apoptosis through a secretory granule-mediated pathway

doi: 10.1038/s41420-026-03122-x

Figure Lengend Snippet: A Human lung smooth muscle cells (HLSMCs), lung fibroblasts (HLFs), and small airway epithelial cells (HSAECs) were treated with TFP at the indicated concentrations for 24 h. Human peripheral blood eosinophils and neutrophils were treated with TFP for 2 h. Cell viability was assessed by staining the cells with Annexin V (AnnV) and DRAQ7. Viable cells, AnnV − DRAQ7 − ; apoptotic cells, AnnV + DRAQ7 − ; necrotic/late apoptotic cells, AnnV + DRAQ7 + . HSAECs, n = 6 from three independent experiments; HLSMCs, eosinophils, neutrophils, n = 4 from four independent experiments; HLFs, n = 3 from one individual experiment representative of three independent experiments (One-way ANOVA for HLSMCs, HLFs; Kruskal-Wallis for HSAECs; Friedman test for eosinophils, neutrophils). B – C Bone marrow-derived MCs (BMMCs) and peritoneal cell-derived MCs (PCMCs) treated under the same conditions as in ( A ) for 24 h. BMMCs, n = 5 from two independent experiments; PCMCs, n = 3 from one individual experiment representative of three independent experiments (One-way ANOVA). Untreated (control) cells were used for statistical comparisons to all other groups in all figures. The bar charts show mean values + SEM or median + interquartile range. * P < 0.05; ** P < 0 .01; **** P < 0.0001. D Effect of TFP on DNA degradation. MCs were preincubated with bafilomycin A1 (Baf A1) (20 nM) for 2 h followed by treatment with TFP (10 μΜ) for 2 h. DNA was extracted from MCs and fragmentation was assessed by agarose gel electrophoresis. St standard marker.

Article Snippet: Primary human lung fibroblasts (HLFs) (PCS-201-013) and primary human lung smooth muscle cells (HLSMCs)(PCS-130-010) were obtained from American Type Culture Collection (ATCC, Manassas, VA) and cultured following the manufacturer’s instructions.

Techniques: Staining, Derivative Assay, Control, Agarose Gel Electrophoresis, Marker

Journal: International Journal of Molecular Sciences

Article Title: A Simple Ionic-Gelation Method for Chitosan Nanoparticle Synthesis and Standardized Protocols for Biological Safety Assessment: Antibacterial Activity, Phytotoxicity, and Biocompatibility

doi: 10.3390/ijms27083673

Figure Lengend Snippet: Cell viability of HDF cells treated with Ch NPs at concentrations of 2.5–160 µg/mL for 24 h. Viability was determined using the MTT assay and expressed as mean ± SE (n = 3). Different letters indicate statistically significant differences ( p < 0.05).

Article Snippet: The biocompatibility of Ch NPs was evaluated using primary HDF (ATCC ® PCS-201-012TM [ ], American Type Culture Collection, Manassas, VA, USA) and HaCaT cells (Cat. No. T0020001, AddexBio, San Diego, CA, USA).

Techniques: MTT Assay

Journal: Cell metabolism

Article Title: Generation of human fatty livers using custom-engineered induced pluripotent stem cells with modifiable SIRT1 metabolism

doi: 10.1016/j.cmet.2019.06.017

Figure Lengend Snippet: (A) Immunohistochemical staining micrographs of SIRT1 show cytoplasmic and nuclear decrease of SIRT1 expression in NASH human livers (n=4) compared to Normal human livers (n=3). Western blot analysis and quantification of SIRT1 normalized to β-Actin in Normal (n=3) and NASH (n=3) human livers (P=0.400, Mann-Whitney test). Quantitative gene expression analysis of SIRT1 expression normalized to actb in Normal (n=4) and NASH (n=4) human livers (P=0.200, Mann-Whitney test). (B) Quantitative gene expression analysis of puromycin selection cassette gene normalized to actb on human fibroblasts transduced with lentiviral vector for –iRFP (hFib-iRFP) or –iKD-SIRT1 (hFibiKD-SIRT1) and non transduced human fibroblasts (hFib) as control (*P=0.0338, *P=0.0130, Kruskal-Wallis test and Dunnett’s multiple comparisons). Quantitative gene expression analysis SIRT1 normalized to actb in hFib-iRFP and hFib-iKD-SIRT1 in the presence or absence of doxycycline (*P=0.0422, Kruskal-Wallis test and Dunnett’s multiple comparisons). Western blot analysis and quantification of SIRT1 normalized to GAPDH on hFib-iRFP (n=4) and hFib-iKD-SIRT1 (n=4) with and without doxycycline treatment (*P=0.0395, Kruskal-Wallis test and Dunnett’s multiple comparisons). (C) Immunofluorescence micrographs of SIRT1 in hFF-iKD-SIRT1 with and without doxycycline treatment, human fetal hepatocytes and human adult hepatocytes were used as controls. Light Red fluorescence and bright light micrographs were used to analyze hFib-iRFP with and without doxycycline treatment. hFib-iRFP (n=4) hFib-iKD-SIRT1 (n=4), human Fetal hepatocytes (n=4), Adult primary hepatocytes untreated (n=3). (D) Light Red fluorescence micrographs of hiPS-iRFP colony after doxycycline exposure for 48h. DAPI was used as counterstaining. Quantitative gene expression analysis of SIRT1 expression normalized to actb show knockdown of SIRT1 in hiPS-iKD-SIRT1-#17 but not in hiPS-IRFP-#3 after exposure to docycycline (*P = 0.0360, *P=0.0140, Kruskal-Wallis test and Dunnett’s multiple comparisons). Western blot analysis and quantification of SIRT1 normalized to GAPDH in hiPS-IRFP-#3 (n=4) and hiPS-iKDSIRT1-# 17 (n=4) with and without doxycycline exposure for 48h (*P=0.0222, Kruskal-Wallis test and Dunnett’s multiple comparisons). (E) Immunofluorescence micrographs of pluripotency markers Nanog, Oct4, TRA-1–60 and SSEA-4 in hiPS-IRFP-#3 and hiPS-iKD-SIRT1-#17. Quantitative gene expression analysis of pluripotency markers c-myc, Lin28 and Oct3/4 normalized to actb shows that hiPS-IRFP and hiPS-iKD-SIRT1 express bona fide pluripotency markers comprable to human Embryonic Stem (hES) cells. hiPS-IRFP#3 with (n=4) and without (n=4) doxycycline, hiPS-iKDSIRT1# 17 with (n=4), and without DOX (n=4), human embryonic stem cells (n=3) were included as controls. hiPS-IRFP-#3 and hiPS-iKD-SIRT1-#17 both carry a normal female karyotype by G-banding analysis.

Article Snippet: At day 10, the liver tissues were fixed in 4% paraformaldehyde for 12 h and 70% ethanol overnight at 4C, and then embedded in paraffin. (See ) Liver organoids formation. iHeps (representing 62.5%), Human Umbilical Vein Endothelial Cells (Lonza, Walkersville, MD) (representing 12.5%), human mesenchymal stromal cells (hMSCs) (ATCC, Manassas, VA) (representing 6.25%) and human fibroblast (representing 6.25%) and Clonal Human Macrophage Primary Cell derived from Human Peripheral Blood (Celprogen, Torrance CA) (representing 12.5%) were plated at a density of 2 × 10 4 cells per cm 2 on low attachment Petri dishes on differentiation medium.

Techniques: Immunohistochemical staining, Staining, Expressing, Western Blot, MANN-WHITNEY, Gene Expression, Selection, Transduction, Plasmid Preparation, Control, Immunofluorescence, Fluorescence, Knockdown

Journal: Cell metabolism

Article Title: Generation of human fatty livers using custom-engineered induced pluripotent stem cells with modifiable SIRT1 metabolism

doi: 10.1016/j.cmet.2019.06.017

Figure Lengend Snippet: (A) Photograph of the organ perfusion and culture organ system constituted by the organ culture chamber, perfusion pump, cell infusion pump and bubble trap. Photograph of recellularized liver matrix with iHeps, human microvascular endothelial cells, mesenchymal cells and fibroblast. Also shown are haematoxylin and eosin staining of engineered human liver tissue–iRFP and –iKD-SIRT1 in the presence of doxycycline. Human normal and fatty livers were used as controls. Asterisks (*) indicate large vacuoles of triglyceride fat with compression and displacement of the nuclei to the periphery of affected hepatocytes consistent with macrovesicular steatosis. (B) Quantitative gene expression analysis of pro-inflammatory marker CD80 and anti-inflammatory marker CD163 (*P=0.0286, Mann-Whitney test) (n=4) in co-cultured iHeps-iKD-SIRT1 or iHeps-iRFP with human primary macrophages in the presence or absence of doxycycline and free fatty acids (FFA). (C) Histological analysis of engineered human fatty liver tissue –iKD-SIRT1 (doxycycline and free fatty acids treated) in the presence or absence of human primary macrophages (n=5). Oil Red O staining show macrovesicular steatosis. Also shown are immunohistochemistry analysis of CD68, NFκB p65, MCP-1 and IL-6 showing increased parenchymal inflammation with addition of human macrophages and the absence of SIRT1 as demonstrated by immunohistochemistry quantification (*P= 0.0116, **P=0.0059, Kruskal-Wallis test and Dunnett’s multiple comparisons). Human fatty livers (n=3) were included as controls. (D) Quantitative gene expression analysis of FGF21 and Selenoprotein-P (From left to right: FGF21, *P=0.0178; Selenoprotein-P, *P=0.0318, *P=0.0213, Kruskal-Wallis test and Dunnett’s multiple comparisons). Hematoxylin and eosin (H&E) staining of engineered human fatty liver tissue –iKD-SIRT1 (free fatty acids treated) in the presence (n=7) or absence of Doxycycline (n=6) in comparison to human normal livers and human NASH livers (n=5). Immunohistochemistry analysis and quantification of FGF21 and Selenoprotein-P in human fatty liver tissues –iKD-SIRT1 −/+ doxycycline compared to human normal and NASH livers. Immunohistochemistry analysis of zonation markers Glutamine synthetase and E-cadherin in human fatty liver tissue –iKD-SIRT1 −/+ doxycycline compared to human normal and NASH liver. Immunohistochemistry analysis and quantification of Ki-67 exhibited a significant higher percentage of positive hepatocytes in human fatty liver tissue –iKD-SIRT1 −/+ doxycycline and human NASH liver compared to human normal liver. (From left to right: *P= 0.0410, *P=0.0247, Kruskal-Wallis test and Dunnett’s multiple comparisons). (E) Human NASH livers and human iPS-derived fatty liver tissues-iKD-SIRT1 with and without doxycycline were scored by Brunt scoring for histologic nonalcoholic fatty liver disease.

Article Snippet: At day 10, the liver tissues were fixed in 4% paraformaldehyde for 12 h and 70% ethanol overnight at 4C, and then embedded in paraffin. (See ) Liver organoids formation. iHeps (representing 62.5%), Human Umbilical Vein Endothelial Cells (Lonza, Walkersville, MD) (representing 12.5%), human mesenchymal stromal cells (hMSCs) (ATCC, Manassas, VA) (representing 6.25%) and human fibroblast (representing 6.25%) and Clonal Human Macrophage Primary Cell derived from Human Peripheral Blood (Celprogen, Torrance CA) (representing 12.5%) were plated at a density of 2 × 10 4 cells per cm 2 on low attachment Petri dishes on differentiation medium.

Techniques: Organ Culture, Staining, Gene Expression, Marker, MANN-WHITNEY, Cell Culture, Immunohistochemistry, Comparison, Derivative Assay