kidney tubular epithelial hk2 cells  (ATCC)

Journal: Redox Biology

Article Title: MAOB promotes ROS-mediated DNA damage, triggering a cyclic MAOB-HNF1A-53BP1-p53 axis that suppresses the malignancy of clear cell renal cell carcinoma

doi: 10.1016/j.redox.2025.103945

Figure Lengend Snippet: The enzymatic activity of MAOB is pivotal in suppressing growth of ccRCC cells via inducing ROS production. A MAOB protein levels in renal proximal tubule epithelial cells (HK2) and ccRCC cell lines were estimated by Western blotting, with GAPDH as an internal loading control. B MAOB was overexpressed in Caki-1 cells and knocked down in 786-O and HK2 cells, as determined by Western blotting. C–F MAOB expression was negatively correlated with ccRCC cell viability (Caki-1, n = 6; 786-O, n = 6; HK2, n = 4) ( C ), proliferation ( n = 3) ( D ), colony formation (Caki-1, n = 7; 786-O, n = 7; HK2, n = 4) ( E ), and sphere formation ( F ). Data in (C–E) are presented as the mean ± standard deviation (SD). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. G Viability of Caki-1/Neo and Caki-1/MAOB cells after treatment with or without 10 μM of various MAOB inhibitors (selegiline, pargyline, or rasagiline) for 48 h ( n = 4). H Viability of Caki-1 cells after transduction with wild-type MAOB, MAOB/Y435W, or a control vector ( n = 6). I, J Oxidative stress were measured using a DCFDA fluorescent probe and flow cytometry in Caki-1/Neo and Caki-1/MAOB cells treated with or without 10 μM selegiline ( n = 3) ( I ) or 5 mM of the antioxidant reagents NAC and GSH ( n = 3) ( J ). K Intracellular H 2 O 2 levels were quantified using the PO-1 fluorescent probe followed by flow cytometric analysis in Caki-1/EV and Caki-1/MAOB cells, with or without treatment with 10 μM selegiline or 5 mM NAC ( n = 3). Exogenous treatment of 200 μM H 2 O 2 was used as a positive control. L Colony-forming ability was evaluated in Caki-1/Neo and Caki-1/MAOB cells treated with or without NAC and GSH. ∗∗ p < 0.01, ∗∗∗ p < 0.001 compared to the vector control group ( n = 3). # p < 0.05, ## p < 0.01, ### p < 0.001 compared to the MAOB-overexpressing group. For comparisons between two groups, a t -test was performed. For comparisons involving more than two groups, ANOVA followed by Tukey's post hoc test was used.

Article Snippet: The human 786-O, A498, and Caki-1 ccRCC cell lines, as well as kidney tubular epithelial HK2 cells, were sourced from American Type Culture Collection (Manassas, VA, USA).

Techniques: Activity Assay, Western Blot, Control, Expressing, Standard Deviation, Transduction, Plasmid Preparation, Flow Cytometry, Positive Control

Journal: Biological research

Article Title: Alpha-kinase1 promotes tubular injury and interstitial inflammation in diabetic nephropathy by canonical pyroptosis pathway.

doi: 10.1186/s40659-023-00416-7

Figure Lengend Snippet: Fig. 4 A, A1 The protein levels of ALPK1 in HK-2 cells treated with 5.5, 10, 20 or 30 mM d-glucose for 24 h were determined by western blot. B, B1 The protein levels of ALPK1 in HK-2 cells treated with 30 mM d-glucose for different periods of time (0, 12, 24 or 48 h) were determined by western blot. Experiments were performed in triplicate. Data were presented as mean ± SD (ns no significance, **P < 0.01, ****P < 0.0001)

Article Snippet: Human kidney proximal tubular epithelial cells (HK2) (ATCC, USA) were cultured in DMEM/F12 (Gibco) medium supplemented with 10% fetal bovine serum and antibiotics in a humidified 5% CO2 incubator at 37 °C.

Techniques: Western Blot

Journal: Biological research

Article Title: Alpha-kinase1 promotes tubular injury and interstitial inflammation in diabetic nephropathy by canonical pyroptosis pathway.

doi: 10.1186/s40659-023-00416-7

Figure Lengend Snippet: Fig. 5 HK-2 cells were treated with ALPK1 siRNA or non-target siRNA for 6 h prior to 24 h-HG (30 mM) treatment. A Densitometry analysis shows knocking down ALPK1 using siRNA transfection downregulated ALPK1 (A1), caspase-1 (A2), caspase-1 P20 (A3), GSDMD (A4), GSDMD-NT (A5) and α-SMA (A6) which increased under high glucose environment. B, B1 Flow cytometry analysis shows high glucose (5.63%) increased proportion of pyroptotic HK-2 cells compared with the control (3.41%) and the proportion was reversed after knockdown of ALPK1 (3.98%). Quadrant 2 presented ratio of pyroptosis cells. Experiments were performed in triplicate. Data were presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (compared with LG group). #P < 0.05, ##P < 0.01, ###P < 0.001, ####P < 0.0001 (compared with HG group)

Article Snippet: Human kidney proximal tubular epithelial cells (HK2) (ATCC, USA) were cultured in DMEM/F12 (Gibco) medium supplemented with 10% fetal bovine serum and antibiotics in a humidified 5% CO2 incubator at 37 °C.

Techniques: Transfection, Flow Cytometry, Control, Knockdown

Journal: Biological research

Article Title: Alpha-kinase1 promotes tubular injury and interstitial inflammation in diabetic nephropathy by canonical pyroptosis pathway.

doi: 10.1186/s40659-023-00416-7

Figure Lengend Snippet: Fig. 6 HK-2 cells were treated with ALPK1 siRNA or non-target siRNA for 6 h prior to 24 h-HG (30 mM) treatment. A Representative images of immunofluorescence staining for NF-κB P65 (green) with nuclei marked by DAPI (blue). ALPK1 knockdown inhibited the nuclear translocation of NF-κB P65 (arrow shows) (scale bar, 50 μm; magnification ×400). B, B1 The protein level of phospho-NF-κB P65 in four groups were determined by western blot. C, C1 Western blot determined cellular distribution of P65 in HK-2 cells. Experiments were performed in triplicate. Data were presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (compared with LG group). #P < 0.05, ##P < 0.01, ###P < 0.001, ####P < 0.0001 (compared with HG group)

Article Snippet: Human kidney proximal tubular epithelial cells (HK2) (ATCC, USA) were cultured in DMEM/F12 (Gibco) medium supplemented with 10% fetal bovine serum and antibiotics in a humidified 5% CO2 incubator at 37 °C.

Techniques: Immunofluorescence, Staining, Knockdown, Translocation Assay, Western Blot

Journal: Biological research

Article Title: Alpha-kinase1 promotes tubular injury and interstitial inflammation in diabetic nephropathy by canonical pyroptosis pathway.

doi: 10.1186/s40659-023-00416-7

Figure Lengend Snippet: Fig. 7 HK-2 cells were treated with ALPK1 siRNA or non-target siRNA for 6 h prior to 24 h-HG (30 mM) treatment. A Representative images of immunofluorescence staining for GSDMD (green) with nuclei marked by DAPI (blue) show knockdown of ALPK1 inhibited translocation of GSDMD to the cell membrane (arrow shows) induced by high glucose (scale bar, 50 μm; magnification ×400). B The protein levels of IL-1β and IL-18 in HG-treated HK-2 cells were determined by western blot. C IL-18 level in supernatant of HK-2 cells was measured by ELISA. Experiments were performed in triplicate. Data were presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (compared with LG group). #P < 0.05, ##P < 0.01, ###P < 0.001, ####P < 0.0001 (compared with HG group)

Article Snippet: Human kidney proximal tubular epithelial cells (HK2) (ATCC, USA) were cultured in DMEM/F12 (Gibco) medium supplemented with 10% fetal bovine serum and antibiotics in a humidified 5% CO2 incubator at 37 °C.

Techniques: Immunofluorescence, Staining, Knockdown, Translocation Assay, Membrane, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Renal Failure

Article Title: Isoliquiritigenin attenuates septic acute kidney injury by regulating ferritinophagy-mediated ferroptosis

doi: 10.1080/0886022X.2021.2003208

Figure Lengend Snippet: Measurement of MDA and nitric oxide after ISL treatment upon LPS induction. LPS induce AKI mice models were developed by intraperitoneal ( i.p. ) LPS injection. A total of 30 mice were randomly divided into six groups ( n = 5): control, ISL, Fer, LPS, LPS plus ISL, and LPS plus Fer. An intraperitoneal injection of LPS (10 mg/kg) was made to induce septic AKI. ISL was administered via gavage at 50 mg/kg 30 min before LPS injection. HK2 cells were treated with 50 μM or 100 μM ISL for 5 h, before septic AKI was induced using 2 μg/mL LPS. Cells were collected 24 h after LPS inducing. And the cell experiments were repeated at three times. (A) MDA measurement of mice kidney tissue homogenate. (B) MDA measurement of HK2 cell homogenate. (C) Nitric Oxide Assay of murine serum. (D) Nitric Oxide Assay of HK2 cell supernatant. ‘*’ means compared with the LPS group and p < 0.05. ‘**’ means compared with the LPS group and p < 0.01. ‘***’ means compared with the LPS group and p < 0.001. ‘#’ means compared with the control group and p < 0.05. ‘##’ means compared with the control group and p < 0.01. ‘###’ means compared with the control group and p < 0.001.

Article Snippet: Human kidney epithelial tubular cell line HK2 (ATCC® CRL-2190TM) was grown in DME/F-12 medium (SH30023.01, HyClone, Logan, UT), supplemented with 10% FBS (Gibco, Life Technologies, Lofer, Austria), 100 units/mL penicillin, and 100 units/mL streptomycin (1705694, HyClone, Logan, UT), at 37 °C in a 5% carbon dioxide humidified incubator.

Techniques: Injection, Control, Nitric Oxide Assay

Journal: Renal Failure

Article Title: Isoliquiritigenin attenuates septic acute kidney injury by regulating ferritinophagy-mediated ferroptosis

doi: 10.1080/0886022X.2021.2003208

Figure Lengend Snippet: ISL inhibited Fe 2+ and lipid peroxidation accumulation in LPS-stimulated cells. HK2 cells were treated with 50 μM or 100 μM ISL for 5 h, before septic AKI was induced using 2 μg/mL LPS. Cells were collected 24 h after LPS induction. FerroOrange was used as fluorescent probe to measure the level of Fe 2+ in HK2 cells. Liperfluo was employed to detect the level of lipid peroxidation in HK2 cells. Mito-Ferrogreen could further detect Fe 2+ ions in the mitochondria of HK2 cells. Magnification: ×60 oil.

Article Snippet: Human kidney epithelial tubular cell line HK2 (ATCC® CRL-2190TM) was grown in DME/F-12 medium (SH30023.01, HyClone, Logan, UT), supplemented with 10% FBS (Gibco, Life Technologies, Lofer, Austria), 100 units/mL penicillin, and 100 units/mL streptomycin (1705694, HyClone, Logan, UT), at 37 °C in a 5% carbon dioxide humidified incubator.

Techniques:

Journal: Renal Failure

Article Title: Isoliquiritigenin attenuates septic acute kidney injury by regulating ferritinophagy-mediated ferroptosis

doi: 10.1080/0886022X.2021.2003208

Figure Lengend Snippet: ISL inhibited the expression of HMGB1 and increased the expression of GPX4 both in vivo and in vitro following LPS stimulation. (A) Western blot assay about HMGB1 and GPX4 in murine kidney tissues. (B) Western blot assay about HMGB1 and GPX4 in HK2 cells.

Article Snippet: Human kidney epithelial tubular cell line HK2 (ATCC® CRL-2190TM) was grown in DME/F-12 medium (SH30023.01, HyClone, Logan, UT), supplemented with 10% FBS (Gibco, Life Technologies, Lofer, Austria), 100 units/mL penicillin, and 100 units/mL streptomycin (1705694, HyClone, Logan, UT), at 37 °C in a 5% carbon dioxide humidified incubator.

Techniques: Expressing, In Vivo, In Vitro, Western Blot

Journal: Renal Failure

Article Title: Isoliquiritigenin attenuates septic acute kidney injury by regulating ferritinophagy-mediated ferroptosis

doi: 10.1080/0886022X.2021.2003208

Figure Lengend Snippet: ISL reduced the expression of NOCA4 both in vivo and in vitro upon LPS induction (A) Western blot assay about NCOA4 in murine kidney tissues. (B) Western blot assay about NCOA4 in HK2 cells. (C) Immunohistochemical staining with NCOA4 in mice kidney tissues. Magnification for immunohistochemical staining: ×400.

Article Snippet: Human kidney epithelial tubular cell line HK2 (ATCC® CRL-2190TM) was grown in DME/F-12 medium (SH30023.01, HyClone, Logan, UT), supplemented with 10% FBS (Gibco, Life Technologies, Lofer, Austria), 100 units/mL penicillin, and 100 units/mL streptomycin (1705694, HyClone, Logan, UT), at 37 °C in a 5% carbon dioxide humidified incubator.

Techniques: Expressing, In Vivo, In Vitro, Western Blot, Immunohistochemical staining, Staining

Journal: Central-European Journal of Immunology

Article Title: Resveratrol alleviates sepsis-induced acute kidney injury by deactivating the lncRNA MALAT1/MiR-205 axis

doi: 10.5114/ceji.2021.109195

Figure Lengend Snippet: Primers for genes

Article Snippet: Human kidney proximal tubular epithelial cell 2 (HK2) cell line, purchased from American Type Culture Collection (ATCC, USA), was inoculated in minimum Eagle’s medium (MEM), which contained 10% fetal bovine serum (FBS), 1% penicillin-streptomycin and 1% non-essential amino acids (NEAA), under an atmosphere of 37°C and 5% CO 2 .

Techniques:

Journal: Central-European Journal of Immunology

Article Title: Resveratrol alleviates sepsis-induced acute kidney injury by deactivating the lncRNA MALAT1/MiR-205 axis

doi: 10.5114/ceji.2021.109195

Figure Lengend Snippet: Resveratrol reduced proliferation and inflammatory response of HK2 cells by regulating lncRNA MALAT1 and miR-205. A ) MALAT1 expression was determined in HK2 cells of PBS group, LPS group and LPS + Res group. *** p < 0.001. B ) MALAT1 expression in HK2 cells was altered after transfection of si-MALAT1 and pcDNA3.1- MALAT1. *** p < 0.001. C ) Viability of HK2 cell line was determined after treatments of PBS, LPS, LPS + Res, LPS + si-MALAT1 and LPS + Res + pcDNA3.1-MALAT1. *** p < 0.001 in comparison to PBS group; ### p < 0.001 in comparison to LPS group; @@@ p < 0.001 in comparison to LPS + Res group. D-F ) TNF-α (D), IL-1β (E) and IL-6 (F) levels were compared among PBS group, LPS group, LPS + Res group, LPS + si-MALAT1 group and LPS + Res + pcDNA3.1-MALAT1 group. *** p < 0.001 in comparison to PBS group; ### p < 0.001 in comparison to LPS group; @@@ p < 0.001 in comparison to LPS + Res group. G ) MiR-205 expression in HK2 cells was measured after transfection of si-MALAT1 and pcDNA3.1-MALAT1. *** p < 0.001. H) MiR-205 expression in HK2 cells was compared among PBS group, LPS group, LPS + Res group, LPS + si-MALAT1 group and LPS + Res + pcDNA3.1-MALAT1 group. *** p < 0.001 in comparison to PBS group; ### p < 0.001 in comparison to LPS group; @@@ p < 0.001 in comparison to LPS + Res group

Article Snippet: Human kidney proximal tubular epithelial cell 2 (HK2) cell line, purchased from American Type Culture Collection (ATCC, USA), was inoculated in minimum Eagle’s medium (MEM), which contained 10% fetal bovine serum (FBS), 1% penicillin-streptomycin and 1% non-essential amino acids (NEAA), under an atmosphere of 37°C and 5% CO 2 .

Techniques: Expressing, Transfection, Comparison

Journal: Journal of the Renin-Angiotensin-Aldosterone System: JRAAS

Article Title: Aldosterone Induces the Proliferation of Renal Tubular Epithelial Cells In Vivo but Not In Vitro

doi: 10.1155/2021/9943848

Figure Lengend Snippet: Effects of aldosterone on the protein expression of PCNA according to western blotting. (a) HK2 cells were treated with aldosterone (0, 10 −9 , 10 −8 , 10 −7 , and 10 −6 mol/L) for 3, 6, 12, 24, 36, or 48 h. Total cellular protein was extracted, and 30 μ g of protein was loaded for western blotting of total PCNA and β -actin. (b) mDCT cells. Data are the mean ± SD ( n = 3). HK2: human kidney proximal tubular epithelial; mDCT: mouse distal convoluted tubule epithelial; PCNA: proliferating cell nuclear antigen.

Article Snippet: Human kidney proximal tubular epithelial (HK2) cells were obtained from Procell Life Science & Technology (Wuhan, China).

Techniques: Expressing, Western Blot

Journal: Journal of the Renin-Angiotensin-Aldosterone System: JRAAS

Article Title: Aldosterone Induces the Proliferation of Renal Tubular Epithelial Cells In Vivo but Not In Vitro

doi: 10.1155/2021/9943848

Figure Lengend Snippet: Dose and time effect of aldosterone on proliferation of mDCT cells and HK2 cells. Aldosterone-stimulated cell growth was measured using the MTT assay. The assay was carried out on mDCT cells and HK2 cells. Cells were divided into untreated (0 mol/L) or treated with different concentrations of aldosterone for 3, 6, 12, 24, 36, or 48 h. The absorbance measurement at 570 nm was recorded. The value represents the mean value of five separate experiments. Each average contains six auxiliary holes, and the results are plotted in a bar chart, mean ± SD ( n = 6). mDCT: mouse distal convoluted tubule epithelial; HK2: human kidney proximal tubular epithelial.

Article Snippet: Human kidney proximal tubular epithelial (HK2) cells were obtained from Procell Life Science & Technology (Wuhan, China).

Techniques: MTT Assay

Journal: Journal of the Renin-Angiotensin-Aldosterone System: JRAAS

Article Title: Aldosterone Induces the Proliferation of Renal Tubular Epithelial Cells In Vivo but Not In Vitro

doi: 10.1155/2021/9943848

Figure Lengend Snippet: Effect of aldosterone on the cycle in HK2 cells (a) and mDCT cells (b). Effects of aldosterone (0, 10 −9 , 10 −8 , 10 −7 , and 10 −6 mol/L) on the number and DNA content of HK2 cells for 3 (a1), 6 (a2), 12 (a3), 24 (a4), 36 (a5), or 48 (a6) hours. Effects of aldosterone on the number and DNA content of mDCT cells treated with aldosterone (0, 10 −9 , 10 −8 , 10 −7 , and 10 −6 mol/L) for 3 (b1), 6 (b2), 12 (b3), 24 (b4), 36 (b5), or 48 (b6) hours. Percentage of cells in the G2/M phase cells is presented as a bar graph; each bar represents the mean ± SD ( n = 3). (c) Proliferation cycle of HK2 cells. (d) mDCT cells. mDCT: mouse distal convoluted tubule epithelial; HK2: human kidney proximal tubular epithelial.

Article Snippet: Human kidney proximal tubular epithelial (HK2) cells were obtained from Procell Life Science & Technology (Wuhan, China).

Techniques:

Journal: Journal of the Renin-Angiotensin-Aldosterone System: JRAAS

Article Title: Aldosterone Induces the Proliferation of Renal Tubular Epithelial Cells In Vivo but Not In Vitro

doi: 10.1155/2021/9943848

Figure Lengend Snippet: Growth of HK2 cells and mDCT cells observed by live imaging. Cells (1 × 10 5 cells/mL) were plated into 96-well plates and stimulated by aldosterone (0, 10 −9 , 10 −8 , 10 −7 , and 10 −6 mol/L) for 48 h. The time interval for recording images was 1 h. After 12 h, the cells began to adhere to the wall. We observed cell growth for 3, 6, 12, 24, 36, and 48 h. (a) HK2 cells. (b) mDCT cells. (A) Aldosterone concentration of 0 mol/L. (B) Aldosterone concentration of 10 −9 mol/L. (C) Aldosterone concentration of 10 −8 mol/L. (D) Aldosterone concentration of 10 −7 mol/L. (E) Aldosterone concentration of 10 −6 mol/L. 1, 2, 3, 4, 5, and 6 show cell growth at 3, 6, 12, 24, 36, and 48 h, respectively. We selected three replicate wells randomly, and the number of cells and the degree of fusion were calculated. Results are plotted as a bar chart, and values are the mean ± SD. (A1, B1) represent changes in the cell number, and (A2, B2) represent changes in cell fusion, compared with that in the group not given aldosterone.

Article Snippet: Human kidney proximal tubular epithelial (HK2) cells were obtained from Procell Life Science & Technology (Wuhan, China).

Techniques: Imaging, Concentration Assay