Journal:

Article Title: Insulin-like growth factor I receptor blockade enhances chemotherapy and radiation responses and inhibits tumour growth in human gastric cancer xenografts

doi: 10.1136/gut.2004.048926

Figure Lengend Snippet: Expression and roles of the insulin-like growth factor/insulin-like growth factor I receptor (IGF/IGF-Ir) axis in human gastric cancer cell lines. (A) Representative data of reverse transcription-polymerase chain reaction analyses. Both MKN45 and MKN74 cells expressed IGF-I message (396 bp) but neither MKN28 nor NUGC4 showed any expression. All cells expressed mRNAs of IGF-II (468 bp), IGF-Ir (755 bp), and insulin-like growth factor receptor 2 (IGF-2r) (430 bp). Controls were β-actin (540 bp) and GAPDH (300 bp). (B–E) After MKN45 cells were cultured to 60% confluence (1×106) in six well plates, medium was replaced with the indicated medium (complete medium (CM) or serum free medium (SFM)) for an additional 48 hours and then cell growth evaluated by trypan blue assay. Growth ratio was calculated compared with the cell number in complete medium with 10% fetal calf serum (1.76 (0.05)×106). Cell growth was suppressed by serum withdrawal (SFM, p<0.0001) but 100 ng/ml IGF-I ((B); p = 0.0230, SFM without IGF-I v SFM with IGF-I) and 100 ng/ml IGF-II ((C); p = 0.0227, SFM without IGF-II v SFM with IGF-II) partially restored growth. (D) Insulin-like growth factor binding protein 3 (IGFBP3) reduced cell growth in complete medium. (E) The growth suppressing effect of IGFBP3 was also seen in serum free medium dose dependently. (F–H) Both IGF-I (F) and IGF-II (100 ng/ml (G, H)) blocked induction of 5% ethanol (EtOH one hour) induced apoptosis in both MKN45 (F, G) and MKN74 (H). (F) p<0.0001, no stimulation v ethanol stimulation without IGF-I; p = 0.0003, ethanol without IGF-I v ethanol with 200 ng/ml IGF-I. (G) p = 0.0097, no stimulation v ethanol stimulation without IGF-II; p = 0.0179, ethanol without IGF-I v ethanol with IGF-I. (H) p = 0.0022, no stimulation v ethanol stimulation without IGF-II; p = 0.0033, ethanol without IGF-I v ethanol with IGF-I.

Article Snippet: Recombinant human IGF-I and IGF-II were purchased from R&D systems (Minneapolis, Minnesota, USA) and NH2 terminally truncated IGF-I (des(1–3) IGF-I) from GroPep (Adelaide, Australia).

Techniques: Expressing, Reverse Transcription, Polymerase Chain Reaction, Cell Culture, Binding Assay

Journal:

Article Title: Insulin-like growth factor I receptor blockade enhances chemotherapy and radiation responses and inhibits tumour growth in human gastric cancer xenografts

doi: 10.1136/gut.2004.048926

Figure Lengend Snippet: Downstream signals from insulin-like growth factor I receptor (IGF-Ir) in gastric cancer cells, MKN45 (A–F), NUGC4 (G, H), and MKN74 (I, J). (A) In MKN45, western blotting showed that 20 ng/ml insulin-like growth factor I (IGF-I) phosphorylated Akt-1, extracellular signal regulated kinase (ERK), and Bad. Both Akt and Bad phosphorylation was reduced by dominant negative forms (dns) of adenoviruses expressing IGF-Ir (Ad-IGF-Ir/dns, 30 multiplicity of infection (moi)). However, Ad-IGF-Ir/dns did not influence ERK-1/-2 phosphorylation to the same degree as Akt. pAkt, phosphorylated Akt-1; tAkt, total Akt-1; pERK, phosphorylated ERK-1/-2; tERK, total ERK-1/-2; pBad, phosphorylated Bad; tBad, total Bad. (B) Western blotting revealed that 10 ng/ml IGF-II phosphorylated Akt which was reduced by infection with adenoviruses expressing truncated IGF-Ir of 482 amino acids long (Ad-IGF-Ir/482st) (30 moi). (C) IGF-Ir/482st blocked p38 activity by p38 kinase assay. ATF2 is a substrate of p38 MAPK. (D) Insulin induces Akt phosphorylation which was not influenced by IGF-Ir/482st. (E) NH2 terminally truncated IGF-I (des(1-3)IGF-I 20 ng/ml) phosphorylated Akt to the same degree as IGF-I. Ad-IGF-Ir/482st reduced des(1-3)IGF-I inducing Akt phosphorylation. dIGF-I, des(1-3)IGF-I. (F) IGF-Ir/482st did not block Akt phosphorylation stimulated by high dose IGF-I, especially 200 ng/ml IGF-I. (G) In NUGC4, IGF-I stimulated phosphorylation of Akt and p38 mitogen activated protein kinase (MAPK) were blocked by both IGF-Ir/dns (30 moi) but phosphorylation of ERKs were not. p-p38, phosphorylated p38 MAPK; t-p38, total p38 MAPK. (H) IGF-Ir/dn blocked IGF-II induced Akt phosphorylation. (I, J) In MKN74, IGF induced Akt phosphorylation was blocked by 100 moi of Ad-IGF-Ir/482st, but phosphorylated ERK was not. LacZ, control (adenovirus expressing β-galactosidase).

Article Snippet: Recombinant human IGF-I and IGF-II were purchased from R&D systems (Minneapolis, Minnesota, USA) and NH2 terminally truncated IGF-I (des(1–3) IGF-I) from GroPep (Adelaide, Australia).

Techniques: Western Blot, Phospho-proteomics, Dominant Negative Mutation, Expressing, Infection, Activity Assay, Kinase Assay, Blocking Assay, Control

Journal: Journal of Neurotrauma

Article Title: Central Infusion of Insulin-Like Growth Factor-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function after Traumatic Brain Injury

doi: 10.1089/neu.2017.5374

Figure Lengend Snippet: Dose-dependent effects of insulin-like growth factor (IGF-1) delivered by central infusion on hippocampal neurogenesis and memory retention after controlled cortical impact (CCI). (A) Representative images of doublecortin (DCX; green) immunoreactivity in the ipsilateral hippocampus of vehicle (Veh) and IGF-1–infused CCI-injured mice at 7 days post-injury. CCI + Veh mice exhibited a robust loss of DCX immunoreactivity in the dentate gyrus granular layer. DCX immunoreactivity appeared to increase in a dose-dependent manner in CCI + IGF-1 mice. Granular layer (GL) and hilus (H). Scale bar represents 50 μm. (B) In the hippocampus ipsilateral to impact, immature neuron density increased as a function of IGF-1 infusate concentration, peaking at 3 μg/day of IGF-1, which produced a significantly higher density of DCX+ cells compared to vehicle infusion (*p < 0.05). Immature neuron density in the hippocampus contralateral to impact was not significantly changed. (C) Recognition indices (% time exploring novel object) at 3 and 7 days post-CCI. Brain-injured mice receiving vehicle or 0.3 μg/day of IGF-1 performed at chance levels, whereas CCI mice receiving 1, 3, or 10 μg/day of IGF-1 performed above chance at either 3 days, 7 days, or both time points (+p < 0.05 compared to 50%). Data are presented as mean + SEM (n = 4 vehicle-treated CCI-injured and n = 5/dose IGF-1–treated CCI-injured mice). SEM, standard error of the mean. Color image is available online at www.liebertpub.com/neu

Article Snippet: Quantification of insulin-like growth factor-1 by enzyme-linked immunosorbent assay By using recombinant hIGF-1, exogenously infused IGF-1 could be distinguished from endogenous mouse IGF-1 using an ELISA kit for hIGF-1. hIGF-1 was quantified using the highly specific Quantikine ® human IGF-1 ELISA kit (DG100; R&D Systems Inc., Minneapolis, MN) according to the manufacturer's instructions. hIGF-1 was not detected in serum or brain samples of vehicle-treated mice.

Techniques: Concentration Assay, Produced

Journal: Journal of Neurotrauma

Article Title: Central Infusion of Insulin-Like Growth Factor-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function after Traumatic Brain Injury

doi: 10.1089/neu.2017.5374

Figure Lengend Snippet: Central infusion of 10 μg/day human insulin-like growth factor-1 (hIGF-1) over 7 days elevates brain levels of hIGF-1 and enhances Akt activation in the hippocampus after controlled cortical impact (CCI). (A) hIGF-1 was detected in the ipsilateral (ipsi) cortex (CTX), and ipsilateral and contralateral (contra) hippocampus (HP) in sham and CCI mice at 7 days post-injury (*p < 0.05, contra HP compared to ipsi CTX or HP). (B) Representative western blot images of phosphorylated Akt (pAkt) and actin, as a control protein, for vehicle (Veh) and IGF-1–treated sham and CCI mice at 7 days post-injury. (C) Infusion of IGF-1 in CCI-injured mice resulted in a significant increase in Akt phosphorylation within the hippocampus at 7 days post-injury, compared to vehicle infusion (*p < 0.005). Optical density (OD) for each pAkt band was normalized to its respective control protein actin band OD; the mean of the triplicate samples for each mouse was normalized to the mean OD value of the vehicle-treated sham group. The results are presented as mean + SEM (n = 5 sham-injured/treatment and n = 9 CCI-injured/treatment). Akt, protein kinase B; SEM, standard error of the mean.

Article Snippet: Quantification of insulin-like growth factor-1 by enzyme-linked immunosorbent assay By using recombinant hIGF-1, exogenously infused IGF-1 could be distinguished from endogenous mouse IGF-1 using an ELISA kit for hIGF-1. hIGF-1 was quantified using the highly specific Quantikine ® human IGF-1 ELISA kit (DG100; R&D Systems Inc., Minneapolis, MN) according to the manufacturer's instructions. hIGF-1 was not detected in serum or brain samples of vehicle-treated mice.

Techniques: Activation Assay, Western Blot

Journal: Journal of Neurotrauma

Article Title: Central Infusion of Insulin-Like Growth Factor-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function after Traumatic Brain Injury

doi: 10.1089/neu.2017.5374

Figure Lengend Snippet: Central infusion of 10 μg/day of insulin-like growth factor-1 (IGF-1) attenuates cognitive and motor impairment after controlled cortical impact (CCI). (A) At 3 days post-injury, novel object recognition (NOR) task performance varied across groups (ANOVA, p < 0.05), but memory retention of injured cohorts was not significantly less than sham cohorts in post-hoc testing. (B) At 7 days post-injury, CCI + Veh mice exhibited a significant reduction in the recognition index compared to either sham group (*p < 0.005). In contrast, memory function of CCI + IGF-1 mice was significantly improved when compared to CCI + Veh mice (#p < 0.05) and was not statistically different from that of sham groups. (C) CCI + Veh mice exhibited significant impairment in motor function at 1, 2, 3, and 5 days post-injury as assessed by a modified neurological severity score (NSS) (*p < 0.001, compared to sham + Veh mice). CCI + IGF-1 mice also exhibited a significant motor impairment at 1, 2, and 3 days post-injury when compared to sham + IGF-1 (*p < 0.001). However, CCI + IGF-1 mice showed improved motor function at 3 and 5 days post-injury compared to CCI + Veh mice (#p < 0.01). Data are presented as mean ± SEM (n = 9 sham-injured/treatment and n = 19 CCI-injured/treatment). ANOVA, analysis of variance; SEM, standard error of the mean; Veh, vehicle.

Article Snippet: Quantification of insulin-like growth factor-1 by enzyme-linked immunosorbent assay By using recombinant hIGF-1, exogenously infused IGF-1 could be distinguished from endogenous mouse IGF-1 using an ELISA kit for hIGF-1. hIGF-1 was quantified using the highly specific Quantikine ® human IGF-1 ELISA kit (DG100; R&D Systems Inc., Minneapolis, MN) according to the manufacturer's instructions. hIGF-1 was not detected in serum or brain samples of vehicle-treated mice.

Techniques: Modification

Journal: Journal of Neurotrauma

Article Title: Central Infusion of Insulin-Like Growth Factor-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function after Traumatic Brain Injury

doi: 10.1089/neu.2017.5374

Figure Lengend Snippet: Central infusion of 10 μg/day of insulin-like growth factor-1 (IGF-1) increases immature neuron density in the injured hippocampus after controlled cortical impact (CCI). (A) Representative images of doublecortin (DCX; green) immunoreactivity in the ipsilateral hippocampus of vehicle (Veh) and IGF-1–infused sham- and CCI-injured mice at 7 days post-injury. Scale bar represents 50 μm. Granular layer (GL) and Hilus (H). (B) Brain injury resulted in a significant decrease in DCX+ cell density in vehicle-treated mice (*p < 0.005, compared to sham + Veh), but not in CCI mice infused with IGF-1. DCX+ cell density was significantly greater in IGF-1–treated brain-injured mice than in vehicle-treated counterparts at 7 days after CCI injury (#p < 0.005, compared to CCI + Veh mice). Immature neuron counts obtained from the ipsilateral granular layer were normalized to the volume of the ipsilateral granular layer to calculate cellular density (1000/mm3). Data are expressed as mean + SEM (n = 4 sham-injured/treatment, n = 10 CCI + Veh, and n = 9 CCI + IGF-1). SEM, standard error of the mean. Color image is available online at www.liebertpub.com/neu

Article Snippet: Quantification of insulin-like growth factor-1 by enzyme-linked immunosorbent assay By using recombinant hIGF-1, exogenously infused IGF-1 could be distinguished from endogenous mouse IGF-1 using an ELISA kit for hIGF-1. hIGF-1 was quantified using the highly specific Quantikine ® human IGF-1 ELISA kit (DG100; R&D Systems Inc., Minneapolis, MN) according to the manufacturer's instructions. hIGF-1 was not detected in serum or brain samples of vehicle-treated mice.

Techniques:

Journal: Journal of Neurotrauma

Article Title: Central Infusion of Insulin-Like Growth Factor-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function after Traumatic Brain Injury

doi: 10.1089/neu.2017.5374

Figure Lengend Snippet: Gross histopathology and regional water content at 7 days after controlled cortical impact (CCI). (A–E) Representative images of cresyl violet staining of the cortex (CTX) and hippocampus (HP) ipsilateral (ipsi) to impact at 7 days post-injury from vehicle (Veh) and IGF-1–treated mice subjected to sham or CCI injury. Sham-injured mice infused with either (A) vehicle or (B) IGF-1 exhibited brain swelling at the craniotomy site, which was not present at the time of surgery. (C) Injured mice treated with vehicle exhibited cortical cavitation and hippocampal cell loss consistent with CCI. (D) The majority of IGF-1–infused brain-injured mice (7 of 10) exhibited characteristic histopathology with minimal or no brain swelling. (E) However, a subset of IGF-1–infused CCI-injured mice (3 of 10) showed distortion of the hippocampus at 7 days post-injury, raising concerns that central infusion of IGF-1 could exacerbate cerebral edema (n = 4 sham-injured/treatment and n = 10 CCI-injured/treatment). Scale bar represents 500 μm. (F) Water content was quantified in mice subjected to CCI injury without cannulation or infusion (CCI), CCI injury with central infusion of vehicle (Veh CCI), or CCI injury with central infusion of 10 μg/day of IGF-1 (IGF-1 CCI; n = 6/group). Comparison by one-way ANOVA revealed infusion of IGF-1 did not result in significant regional cerebral edema; however, a small subset of mice (2 of 6) had markedly increased water content in the ipsilateral (ipsi) hippocampus. Contralateral (contra). Individual data points are superposed with bars representing mean + SEM. ANOVA, analysis of variance; SEM, standard error of the mean.

Article Snippet: Quantification of insulin-like growth factor-1 by enzyme-linked immunosorbent assay By using recombinant hIGF-1, exogenously infused IGF-1 could be distinguished from endogenous mouse IGF-1 using an ELISA kit for hIGF-1. hIGF-1 was quantified using the highly specific Quantikine ® human IGF-1 ELISA kit (DG100; R&D Systems Inc., Minneapolis, MN) according to the manufacturer's instructions. hIGF-1 was not detected in serum or brain samples of vehicle-treated mice.

Techniques: Histopathology, Staining, Comparison

Journal: Journal of Neurotrauma

Article Title: Central Infusion of Insulin-Like Growth Factor-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function after Traumatic Brain Injury

doi: 10.1089/neu.2017.5374

Figure Lengend Snippet: Delayed infusion of 10 μg/day of insulin-like growth factor-1 (IGF-1) increased hippocampal immature neuron density 1 week after controlled cortical impact (CCI). (A) Compared to CCI + Veh mice, CCI + IGF-1 mice showed increased doublecortin (DCX; green) immunoreactivity. Scale bar represents 100 μm. (B) Infusion of IGF-1 post-CCI resulted in significantly increased immature neuron density at 7 days post-injury, compared to vehicle treatment (#p < 0.05). Data are presented as mean + SEM (n = 3 CCI + Veh and n = 5 CCI + IGF-1). SEM, standard error of the mean. Color image is available online at www.liebertpub.com/neu

Article Snippet: Quantification of insulin-like growth factor-1 by enzyme-linked immunosorbent assay By using recombinant hIGF-1, exogenously infused IGF-1 could be distinguished from endogenous mouse IGF-1 using an ELISA kit for hIGF-1. hIGF-1 was quantified using the highly specific Quantikine ® human IGF-1 ELISA kit (DG100; R&D Systems Inc., Minneapolis, MN) according to the manufacturer's instructions. hIGF-1 was not detected in serum or brain samples of vehicle-treated mice.

Techniques:

Journal: iScience

Article Title: Contrasting consequences of podocyte insulin-like growth factor 1 receptor inhibition

doi: 10.1016/j.isci.2024.109749

Figure Lengend Snippet: In vitro differential suppression of podocyte IGF1R activity reveals that partial inhibition is beneficial but near total loss is highly detrimental (A) Representative Western blot shows >90% reduction of IGF1R protein in NC-IGF1RKD cells but no reduction of IR protein expression. Bar graphs show densitometry expressed as the mean fold change +/− SEM, t-test, ∗∗∗∗ p < 0.0001, n = 18 independent experiments. (B) Phosphorylation of AKT and p44/42MAPK in response to acute IGF1 stimulation at 10 ng and 100 ng/mL for 10 min was significantly reduced in NC-IGF1RKD podocytes. Data are expressed as the mean ± SEM, one-way ANOVA with Tukey’s multiple comparison test, ∗∗ p < 0.005, n = 3 independent experiments. (C) Western blot shows that IGF1R expression is reduced by ∼70% in wild-type podocytes exposed to 100 nM picropodophyllin for 24 h. Data are expressed as the mean ± SEM, t-test, ∗ p < 0.05, n = 3 independent experiments. No significant change in IR expression was observed. (D) Western blot shows the phosphorylation of AKT and p44/42MAPK in response to acute IGF1 stimulation at 10 ng and 100 ng/mL for 10 min in podocytes exposed to 100 nM picropodophyllin for 24 h. Data expressed as the mean ± SEM, ∗ p < 0.05, n = 3 independent experiments. (E) ∼50% of NC-IGF1RKD cells survive 7 days after gene excision. Treatment of wild-type podocytes with 100 nM picropodophyllin for 24 h has no effect on cell survival. Data are expressed as the mean ± SEM, t-test, ∗∗ p < 0.005, n = 3–4 independent experiments. See also Figures S5 and .

Article Snippet: Recombinant Mouse IGF1 protein , Novus , Cat# NBP2-35081.

Techniques: In Vitro, Activity Assay, Inhibition, Western Blot, Expressing, Comparison

Journal: iScience

Article Title: Contrasting consequences of podocyte insulin-like growth factor 1 receptor inhibition

doi: 10.1016/j.isci.2024.109749

Figure Lengend Snippet:

Article Snippet: Recombinant Mouse IGF1 protein , Novus , Cat# NBP2-35081.

Techniques: Recombinant, Protease Inhibitor, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Staining, RNAscope, Western Blot, Cell Culture, Software, Plasmid Preparation