Journal: eLife

Article Title: Renal interstitial cells promote nephron regeneration by secreting prostaglandin E2

doi: 10.7554/eLife.81438

Figure Lengend Snippet: ( A–C ) lhx1a mRNA levels were assessed by RT-PCR at 7 dpi. β-actin was used as a sample control. ( D–K ) lhx1a whole-mount in situ hybridization (WISH) showing the trunk kidney region at 7 dpi. XAV939 ( F ) could reduce the number of lhx1a + cell aggregates and dmPGE2 could rescue the influence of XAV939 treatment ( G ); ICRT 14 ( H ) or ICG-001 ( J ) could reduce the number of lhx1a + cell aggregates and dmPGE2 could not rescue the influence of ICRT 14 ( I ) or ICG-001 ( K ) treatment. n = 5–7 in each condition. ( L ) lhx1a + cell aggregates of whole kidney were calculated using ImageJ. Data were analyzed by ANOVA, ***p<0.001; ns, no significant difference. ( M–V ) Immunofluorescence staining of β-catenin in Tg(lhx1a:DsRed ) zebrafish kidneys at 5 dpi. ( M, N ) Zebrafish injected DMSO as a control group, and the amount of β-catenin could be detected in lhx1a + cell aggregates during renal progenitor cell (RPC) aggregation ( M ) or proliferation ( N ). ( O ) β-catenin level in lhx1a + cell aggregates of cox2a -/- was significantly less than the control group, and injection of dmPGE2 ( P, Q ) could rescue the influence of Cox2a deficiency. ( R ) β-catenin level in lhx1a + cell aggregates of wnt4a -/- was significantly less than the control group, and injection of dmPGE2 ( S, T ) could rescue the influence of Wnt4a deficiency. Injection of PKI ( U ) could reduce β-catenin level in lhx1a + cell aggregates, while injection of dmPGE2 ( V ) could not rescue the influence of PKI treatment. n = 3–6 in ( M–V ). Scale bar in ( M–V ), 50 μm. ( W ) Bar chart depicting β-catenin levels following acute kidney injury (AKI) ( M–V ). Fluorescent intensities per unit area were measured at the lhx1a + RPC aggregates using ImageJ. β-catenin levels of lhx1a + RPCs during RPC aggregation normalized as 1. Ag, aggregation; Pr, proliferation. n = 3–6 in each condition. Data were analyzed by ANOVA, ***p<0.001; ns, no significant difference. Figure 6—source data 1. Original gel files of . Figure 6—source data 2. Numerical data for , , , and .

Article Snippet: During the nephron regeneration stage, the Indo (I7378-5G, Sigma; 400 μM, 10 μL per fish), NS-398 (N194-5MG, Sigma; 140 μM, 10 μL per fish), TG4-155 (S6793, Selleck; 400 μM, 10 μL per fish), dmPGE2 (D0160, Sigma; 600 μM, 10 μL per fish), GW627368X (T1978, TOPSCIENCE; 200 μM, 10 μL per fish), XAV939 (S1180, Selleck; 200 μM, 10 μL per fish), ICRT 14 (HY-16665, MCE; 10 μM, 10 μL per fish), ICG-001 (HY-14428, MCE; 100 μM, 10 μL per fish), PKI (HY-P1290A, MCE; 20 μM, 10 μL per fish), and H89 (HY-15979, MCE; 200 μM, 10 μL per fish) were intraperitoneally injected into zebrafish at 2, 4, and 6 dpi.

Techniques: Reverse Transcription Polymerase Chain Reaction, Control, In Situ Hybridization, Immunofluorescence, Staining, Injection

Journal: eLife

Article Title: Renal interstitial cells promote nephron regeneration by secreting prostaglandin E2

doi: 10.7554/eLife.81438

Figure Lengend Snippet: ( A, B ) lhx1a mRNA levels were evaluated by RT-PCR at 7 dpi. β-actin was used as a sample control. ( C–H ) lhx1a whole-mount in situ hybridization (WISH) showing the trunk kidney region at 7 dpi. PKI ( E ) or H89 ( G ) treatment reduced the number of lhx1a + cell aggregates, while injection of dmPGE2 could not rescue the influence of PKI ( F ) or H89 ( H ) treatment. ( I ) lhx1a + cell aggregates of whole kidney were calculated using ImageJ. n = 5–7 in each condition. Data were analyzed by ANOVA, ***p<0.001; ns, no significant difference. ( J–N ) Immunofluorescence staining of p-S9-GSK3β (arrowheads) in Tg(lhx1a:DsRed ) zebrafish kidneys at 5 dpi. ( J, K ) Zebrafish injected with DMSO as a control group, and the amount of p-S9-GSK3β could be detected in lhx1a + cell aggregates cytoplasm during RPC aggregation ( J ) or proliferation ( K ). ( L ) p-S9-GSK3β in lhx1a + cell aggregates of cox2a -/- was hardly detectable, and injection of dmPGE2 ( M, N ) could rescue the influence of Cox2a deficiency. Injection of PKI ( O ) could reduce p-S9-GSK3β level in lhx1a + cell aggregates, while injection of dmPGE2 ( P ) could not rescue the influence of PKI treatment. ( Q–W ) Immunofluorescence staining of p-S675-β-catenin in Tg(lhx1a:DsRed ) zebrafish kidneys at 5 dpi. ( Q, R ) Injection of DMSO as a control group and amounts of p-S675-β-catenin could be detected in lhx1a + cell aggregates during RPC aggregation ( Q ) or proliferation ( R ). ( S ) p-S675-β-catenin level in lhx1a + cell aggregates of cox2a -/- was hardly detectable, and injection of dmPGE2 ( T, U ) could rescue the influence of Cox2a deficiency. Injection of PKI ( V ) could reduce p-S675-β-catenin level in lhx1a + cell aggregates, while injection of dmPGE2 ( W ) could not rescue the influence of PKI treatment. Scale bar, 50 μm. ( X, Y ) Bar chart depicting p-S9-GSK3β ( X ) and p-S675-β-catenin ( Y ) levels following acute kidney injury (AKI) ( J–W ). Fluorescent intensities per unit area were measured at the lhx1a + RPC aggregates using ImageJ. p-S9-GSK3β or p-S675-β-catenin levels of lhx1a + RPCs during RPC aggregation normalized as 1. Ag, aggregation; Pr, proliferation. n = 3–6 in each condition. Data were analyzed by ANOVA, ***p<0.001; ns, no significant difference. Figure 7—source data 1. Original gel files of . Figure 7—source data 2. Numerical data for .

Article Snippet: During the nephron regeneration stage, the Indo (I7378-5G, Sigma; 400 μM, 10 μL per fish), NS-398 (N194-5MG, Sigma; 140 μM, 10 μL per fish), TG4-155 (S6793, Selleck; 400 μM, 10 μL per fish), dmPGE2 (D0160, Sigma; 600 μM, 10 μL per fish), GW627368X (T1978, TOPSCIENCE; 200 μM, 10 μL per fish), XAV939 (S1180, Selleck; 200 μM, 10 μL per fish), ICRT 14 (HY-16665, MCE; 10 μM, 10 μL per fish), ICG-001 (HY-14428, MCE; 100 μM, 10 μL per fish), PKI (HY-P1290A, MCE; 20 μM, 10 μL per fish), and H89 (HY-15979, MCE; 200 μM, 10 μL per fish) were intraperitoneally injected into zebrafish at 2, 4, and 6 dpi.

Techniques: Reverse Transcription Polymerase Chain Reaction, Control, In Situ Hybridization, Injection, Immunofluorescence, Staining

Journal: Frontiers in Microbiology

Article Title: Cyclic AMP Pathway Activation and Extracellular Zinc Induce Rapid Intracellular Zinc Mobilization in Candida albicans

doi: 10.3389/fmicb.2018.00502

Figure Lengend Snippet: Inhibition of cAMP signaling inhibits glucose-induced zinc mobilization. (A) Adenylate cyclase (AC) inhibitor MDL12330 (MDL, 417 μM) or PKA inhibitor myr-PKI (25 μM) was added for 20 min (stimuli) before addition of 0.25% glucose. (B) MDL12330 (MDL, 417 μM) and myr-PKI (25 μM) was added for 20 min (stimuli) before 20 μM zinc was added for 20 min (Zn 2+ ). Finally 100 μM EDTA was added for 20 min. Open triangle (Δ) indicate p < 0.01 in one-way ANOVA test for both myr-PKI and MDL compared to control at the specified time point, while closed triangle ( ) indicate p ≤ 0.01 in one-way ANOVA test for only myr-PKI n = 3–6.

Article Snippet: The following compounds were used: 25 μl of dithiothreitol (DTT, D0632, Sigma), theophylline (T1633, Sigma), caffeine (C8960, Sigma), dibutyryl cAMP (D0260, Sigma), MDL12330 (sc-3537, Santa Cruz,) or myristoylated PKI (14–22) (myr-PKI, sc-471154, Santa Cruz,).

Techniques: Inhibition

Journal: Frontiers in Microbiology

Article Title: Cyclic AMP Pathway Activation and Extracellular Zinc Induce Rapid Intracellular Zinc Mobilization in Candida albicans

doi: 10.3389/fmicb.2018.00502

Figure Lengend Snippet: Proposed mechanism for intracellular zinc mobilization. Glucose induces cAMP-PKA-dependent intracellular zinc release and can be inhibited by the AC inhibitor MDL12330 or the PKA inhibitor myr-PKI. The cAMP analog, dibutyryl cAMP and PDE inhibitors, also induces intracellular zinc release. Extracellular Zn 2+ induces ER zinc efflux through a cAMP-PKA independent mechanism.

Article Snippet: The following compounds were used: 25 μl of dithiothreitol (DTT, D0632, Sigma), theophylline (T1633, Sigma), caffeine (C8960, Sigma), dibutyryl cAMP (D0260, Sigma), MDL12330 (sc-3537, Santa Cruz,) or myristoylated PKI (14–22) (myr-PKI, sc-471154, Santa Cruz,).

Techniques: