Journal: eBioMedicine

Article Title: Leukemia inhibitory factor is a therapeutic target for renal interstitial fibrosis

doi: 10.1016/j.ebiom.2022.104312

Figure Lengend Snippet: LIF expression in renal biopsies and urine from CKD patients. (a) The expression of LIF along with the decline of eGFR (n = 39). (b) The association between the log2 of LIF expression and eGFR (n = 39). (c) Differential expression of IL6 family cytokines between subjects with (n = 33) and without (n = 6) TIF. (d) The expression of LIF in different stages of Oxford-T grades of IgAN (n = 39). (e) Differential expression of IL6 family cytokines between CKD (n = 28) and healthy subjects (n = 9) from the ERCB Nephrotic Syndrome Data set. (f) The association between the log2 of LIF expression and eGFR in subjects (n = 33) from the ERCB Nephrotic Syndrome Data set. (g) Western blot analysis of LIF protein in 24 h-urine. (h) The amount of LIF protein in 24 h-urine was increased along with the stage of Oxford-T grades of IgAN (n = 50) and healthy controls (n = 8). The amount of LIF protein in 24 h-urine was measure by ELISA. (i) The amount of LIF protein in 24 h-urine negatively correlated with eGFR of IgAN patients (n = 50). (j–l) The level of uLIF at baseline predicts the risk of CKD progression to ESRD. Tertiles of uLIF-7 levels had a graded relationship with the risk of CKD progression to ESRD in total cohort (j), in subgroups of patients with eGFR <60 ml/min/1.73 m 2 (k) and patients with proteinuria >1.0 g/d (l). Data in (a), (d) and (h) were represented as median ± interquartile range (IQR) and P values were determined by one-way ANOVA (Least-Significant Difference test). The Spearman correlation analysis was used in (b), (f) and (i). The R package limma (3.46.0) was used in (c) and (e) for gene differential expression analysis between two groups. Data in (j), (k) and (l) were generated using the Kaplan–Meier method and compared using the log-rank test.

Article Snippet: The concentration of LIF in the culture medium of normal rat kidney interstitial fibroblast cell lines (NRK49F) was determined using rat-specific LIF ELISA kits (ELK5072, ELK Biotechnology, Wuhan, China).

Techniques: Expressing, Quantitative Proteomics, Western Blot, Enzyme-linked Immunosorbent Assay, Generated

Journal: eBioMedicine

Article Title: Leukemia inhibitory factor is a therapeutic target for renal interstitial fibrosis

doi: 10.1016/j.ebiom.2022.104312

Figure Lengend Snippet: Upstream regulators of LIF expression. (a–e) LIF upregulated the expression of itself in NRK49F cells through ERK-EGR1 axis. NRK49F cells were treated with LIF for 24 h. (a) Real-time PCR showed that LIF upregulated itself in NRK49F cells. Relative Lif mRNA levels were shown as fold induction over controls after normalization with gapdh, respectively. ∗ P < 0.01 versus controls (n = 3). (b and c) Western blot analyses (b) and quantitative data (c) showed that LIF induced the expression of itself and EGR1 in a dose-dependent manner. # P < 0.05, ∗ P < 0.01 versus controls (n = 4). (d and e) Western blot analyses (d) and quantitative data (e) showed that knocking down EGR1 diminished LIF-induced LIF production. # P < 0.05, ∗ P < 0.01 versus NC+LIF (n = 4). (f) Upstream regulators of Lif expression. NRK49F cells were treated with OSM (20 ng/ml) and IL11 (10 ng/ml). (g) Upstream regulators of Lif expression. NRK49F cells were treated with TGF-β1 (8 ng/ml), TNF-α (8 ng/ml) and Ang II (10 nM/ml) for 24 h. Real-time PCR measured the mRNA level of Lif . Relative Lif mRNA levels were showed as fold induction over controls after normalization with gapdh, respectively. ∗ P < 0.01 versus controls (n = 4). (h) The mRNA level of Lif were induced by TGF-β1 induced LIF expression in a dose-dependent manner in NRK49F cells. ∗ P < 0.01 versus controls (n = 4). (i and j) Western blotting Representative Western blot (i) and quantitative data (j) showed that the protein level of LIF were induced by TGF-β1 in a dose-dependent manner in NRK49F cells. # P < 0.05, ∗ P < 0.01 versus controls (n = 4). (k) NRK49F cells were treated with TGF-β1 (8 ng/ml) for 24 h, and the LIF concentration in the culture medium was determined by ELISA. ∗ P < 0.01 versus controls (n = 4). Data were expressed as means ± SD. Student's t-test in (f), (g) and (k) or one-way ANOVAs (Least-Significant Difference test or Dunnett's T3 test) in (a), (c), (e), (h) and (j) was used for determining the P values while comparing between each pair of groups, respectively.

Article Snippet: The concentration of LIF in the culture medium of normal rat kidney interstitial fibroblast cell lines (NRK49F) was determined using rat-specific LIF ELISA kits (ELK5072, ELK Biotechnology, Wuhan, China).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Concentration Assay, Enzyme-linked Immunosorbent Assay