Journal: Cell Reports Medicine

Article Title: Unacylated-Ghrelin Impairs Hippocampal Neurogenesis and Memory in Mice and Is Altered in Parkinson’s Dementia in Humans

doi: 10.1016/j.xcrm.2020.100120

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Article Snippet: For IGF-1 and GH, platelet-rich plasma samples were analyzed using Human IGF-1 DuoSet ELISA (cat. No. DY291, R&D Systems), and Human GH DuoSet ELISA (Cat. No. DY1067, R&D Systems), using half-volume Nunclon Microwell 96-well plates.

Techniques: Clinical Proteomics, Recombinant, Saline, RNAscope, Bicinchoninic Acid Protein Assay, Enzyme-linked Immunosorbent Assay, Software, Microscopy

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: Animal

Article Title: Effects of angiogenin on granulosa and theca cell function in cattle

doi: 10.1017/s1751731116002044

Figure Lengend Snippet: Figure 2 Effect of angiogenin on basal, LH- and IGF1-induced androstenedione (a) and progesterone (b) production by large follicle theca cells (experiment 2). Cells were cultured for 48h as described in ‘Materials and methods’ section, and then treated for an additional 48h with 0, 30 or 100ng/ml of angiogenin (ANG) and: Control (no IGF1 or LH), IGF1 (30ng/ml), LH (30 ng/ml) or LH plus IGF1. Values are means ± SEM of three separate experiments (n = 6). *Within a panel and treatment, mean differs (P < 0.05) from its respective 0 ANG mean. #Within a panel, mean differs (P < 0.05) from its respective Control mean. +Within a panel, mean differs (P < 0.05) from its respective LH alone treatment mean.

Article Snippet: Hormones and reagents The hormones and reagents used in cell culture were: ovine FSH (NIDDK-oFSH-20; activity: 175 X NIH-FSH-S1 U/mg) and ovine LH (NIDDK-oLH-26; activity: 1.0 X NIH-LH-S1 U/mg) from the National Hormone & Pituitary Program (Torrance, CA, USA), recombinant human ANG and IGF-1 (IGF1) from R&D Systems (Minneapolis, MN, USA), testosterone from Steraloids (Wilton, NH, USA), fetal calf serum (FCS) from EquiTech-Bio Inc. (Kerrville, TX, USA), and collagenase and DNase from Sigma-Aldrich Corp. (St. Louis, MO, USA).

Techniques: Cell Culture, Control

Journal: Animal

Article Title: Effects of angiogenin on granulosa and theca cell function in cattle

doi: 10.1017/s1751731116002044

Figure Lengend Snippet: Figure 1 Effect of angiogenin on basal, FSH- and IGF1-induced estradiol (a) and progesterone (b) production by small-follicle granulosa cells (experiment 1). Cells were cultured for 48 h as described in ‘Materials and methods’ section, and then treated for an additional 48 h with 0, 30 or 100 ng/ml of angiogenin (ANG) and: Control (no IGF1 or FSH), IGF1 (30 ng/ml), FSH (30 ng/ml) or FSH plus IGF1. Values are means ± SEM of three separate experiments (n = 6). *Within a panel and treatment, mean differs (P < 0.05) from its respective 0 ANG mean. #Within a panel, mean differs (P < 0.05) from its respective Control mean. +Within a panel, mean differs (P < 0.05) from its respective IGF1 alone or FSH alone treatment mean.

Article Snippet: Hormones and reagents The hormones and reagents used in cell culture were: ovine FSH (NIDDK-oFSH-20; activity: 175 X NIH-FSH-S1 U/mg) and ovine LH (NIDDK-oLH-26; activity: 1.0 X NIH-LH-S1 U/mg) from the National Hormone & Pituitary Program (Torrance, CA, USA), recombinant human ANG and IGF-1 (IGF1) from R&D Systems (Minneapolis, MN, USA), testosterone from Steraloids (Wilton, NH, USA), fetal calf serum (FCS) from EquiTech-Bio Inc. (Kerrville, TX, USA), and collagenase and DNase from Sigma-Aldrich Corp. (St. Louis, MO, USA).

Techniques: Cell Culture, Control

Journal: Animal

Article Title: Effects of angiogenin on granulosa and theca cell function in cattle

doi: 10.1017/s1751731116002044

Figure Lengend Snippet: Figure 3 Effect of angiogenin on proliferation of large follicle granulosa cells induced by 10% FCS (a) (experiment 5) and 3H-thymidine incorporation into large follicle granulosa and theca cells (b) treated with IGF1 (experiment 6). (a) Cells were cultured for 48 h as described in ‘Materials and methods’ section, and then treated for an additional 48 h with 0 or 300 ng/ml of angiogenin in the presence of 10% FCS; cells were enumerated via Coulter counting. (b) Cells were cultured for 48 h as described in ‘Materials and methods’ section, serum-starved for 24 h, and then treated for an additional 40h with 3H-thymidine and IGF1 (30 ng/ml) and either 0 or 300 ng/ml of angiogenin. *Within a panel, asterisk (*) indicates mean differs (P < 0.05) from its respective control (0 ANG) mean. Values are means ± SEM of three separate experiments (n = 6).

Article Snippet: Hormones and reagents The hormones and reagents used in cell culture were: ovine FSH (NIDDK-oFSH-20; activity: 175 X NIH-FSH-S1 U/mg) and ovine LH (NIDDK-oLH-26; activity: 1.0 X NIH-LH-S1 U/mg) from the National Hormone & Pituitary Program (Torrance, CA, USA), recombinant human ANG and IGF-1 (IGF1) from R&D Systems (Minneapolis, MN, USA), testosterone from Steraloids (Wilton, NH, USA), fetal calf serum (FCS) from EquiTech-Bio Inc. (Kerrville, TX, USA), and collagenase and DNase from Sigma-Aldrich Corp. (St. Louis, MO, USA).

Techniques: Cell Culture, Control

Journal: Animal

Article Title: Effects of angiogenin on granulosa and theca cell function in cattle

doi: 10.1017/s1751731116002044

Figure Lengend Snippet: Figure 4 Effect of angiogenin (ANG) on steroidogenic enzyme gene expression in granulosa and theca cells (experiment 7). (a) Effect of effect of ANG (300ng/ml) on CYP19A1 (aromatase) and CYP11A1 (side-chain cleavage enzyme) mRNA abundance in small follicle granulosa cells. (b) Effect of ANG (300 ng/ml) on CYP17A1 and CYP11A1 mRNA abundance in large follicle theca cells. Granulosa cells from small follicles and theca cells from large follicles were cultured for 48h in the presence of 10% FCS, and then treated in serum-free medium with ANG for 24 h. Granulosa cells were also concomitantly treated with 30ng/ml of IGF1 and 30ng/ml of FSH, and theca cells were concomitantly treated with 30ng/ml of IGF1 and 30ng/ml of LH. Values are means of three separate experiments (±SEM) and normalized to constitutively expressed 18S ribosomal RNA. *Mean differs (P <0.05) from its respective control (0 ng/ml of ANG).

Article Snippet: Hormones and reagents The hormones and reagents used in cell culture were: ovine FSH (NIDDK-oFSH-20; activity: 175 X NIH-FSH-S1 U/mg) and ovine LH (NIDDK-oLH-26; activity: 1.0 X NIH-LH-S1 U/mg) from the National Hormone & Pituitary Program (Torrance, CA, USA), recombinant human ANG and IGF-1 (IGF1) from R&D Systems (Minneapolis, MN, USA), testosterone from Steraloids (Wilton, NH, USA), fetal calf serum (FCS) from EquiTech-Bio Inc. (Kerrville, TX, USA), and collagenase and DNase from Sigma-Aldrich Corp. (St. Louis, MO, USA).

Techniques: Gene Expression, Cell Culture, Control