recombinant igf2  (Cusabio)

Journal: iScience

Article Title: Reprogramming of fibroblasts into cancer-associated fibroblasts via IGF2-mediated autophagy promotes metastasis of lung cancer cells

doi: 10.1016/j.isci.2024.111269

Figure Lengend Snippet: Lung cancer cells induced CAFs activation via IGF2 secretion (A) mRNA levels of IGF2 in cells were detected by qPCR. (B) IGF2 concentrations in cell culture medium were detected by ELISA. (C) NFs were treated with IGF2 (50 and 100 ng/mL) for 24 h. Cell morphology was observed under a microscope. (D) NFs were treated with IGF2. α-SMA and FAP expressions were detected by western blotting. (E) NFs were treated with IGF2. Cell proliferation was examined by CCK-8 kit after 48 h. (F and G) NFs were treated with IGF2. NFs migration was detected by trans-well assay after 24 h. (H) NFs were treated with CM or CM + IGF2 neutralizing antibody. Cell morphology was observed under a microscope. (I) NFs were treated with CM or CM + IGF2 neutralizing antibody. α-SMA and FAP expressions were detected by western blotting. (J) NFs were treated with CM or CM + IGF2 neutralizing antibody, and IGF2 (100 ng/mL). Cell proliferation was examined by CCK-8 kit after 48 h. (K and L) NFs were treated with CM or CM + IGF2 neutralizing antibody. NFs migration was detected by trans-well assay after 24 h. (M) The effect of IGF2 knockdown by RNAi. (N) IGF2 expression was knocked down in lung cancer cells, and then the CM was collected and used to culture NFs. α-SMA and FAP expressions were detected by western blotting. (O and P) IGF2 expression was knocked down in lung cancer cells by RNAi, and then the CM was collected and used to culture NFs. NFs migration was detected by trans-well assay after 24 h. Ab, neutralizing antibody. Scale bar, 100 μm. Data represents the mean ± SD from three independent experiments. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 .

Article Snippet: Recombinant human IGF2 , Sino Biological , Cat#13032-HNAY.

Techniques: Activation Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Microscopy, Western Blot, CCK-8 Assay, Migration, Knockdown, Expressing

Journal: iScience

Article Title: Reprogramming of fibroblasts into cancer-associated fibroblasts via IGF2-mediated autophagy promotes metastasis of lung cancer cells

doi: 10.1016/j.isci.2024.111269

Figure Lengend Snippet: IGF2-induced autophagy mediated the effect of lung cancer cells on CAFs activation (A) NFs were treated with CM and then were stained with 0.5 μg/mL AO for 15 min. The acidic vesicular organelles (AVOs) formation was observed under a fluorescence microscope (scale bar, 20 μm). (B) NFs were treated with CM. The expression of LC-3II and ATG5 were detected by western blotting. (C) NFs were treated with IGF2 (50 or 100 mg/mL) for 24 h. The AVOs formation was examined by AO staining (scale bar, 20 μm). (D) NFs were treated with IGF2. The expression of LC-3II and ATG5 was detected by western blotting. (E) NFs were treated with IGF2 or IGF2 + 3-MA (5 mM). The AVOs formation was examined by AO staining (scale bar, 20 μm). (F) NFs were treated with IGF2 or IGF2 + 3-MA. Protein expressions were detected by western blotting. (G) NFs were treated with IGF2 or IGF2 + 3-MA. Cell proliferation was examined by CCK-8 kit after 48 h. (H and I) NFs were treated with IGF2 or IGF2+3-MA. NFs migration was detected by trans-well assay after 24 h (scale bar, 100 μm). (J) NFs were treated with IGF2, CM, CM + IGF2 Ab, and CM + 3-MA. The AVOs formation was examined by AO staining (scale bar, 20 μm). (K) NFs were treated with CM, IGF2, and CM + IGF2 Ab. Protein expressions were detected by western blotting. (L) NFs were treated with IGF2, CM, CM + IGF2 Ab, and CM + 3-MA. Cell proliferation was examined by CCK-8 kit after 48 h. (M and N) NFs were treated with IGF2, CM, CM + IGF2 Ab, and CM + 3-MA. NFs migration was detected by trans-well assay after 24 h (scale bar, 100 μm). (O and P) NFs were treated with RAPA (100 nM); α-SMA and FAP expressions were detected by western blotting (scale bar, 20 μm). (Q and R) NFs were treated with RAPA; NFs migration was detected by trans-well assay after 24 h. Ab, neutralizing antibody (scale bar, 100 μm). Data represents the mean ± SD from three independent experiments. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 .

Article Snippet: Recombinant human IGF2 , Sino Biological , Cat#13032-HNAY.

Techniques: Activation Assay, Staining, Fluorescence, Microscopy, Expressing, Western Blot, CCK-8 Assay, Migration

Journal: iScience

Article Title: Reprogramming of fibroblasts into cancer-associated fibroblasts via IGF2-mediated autophagy promotes metastasis of lung cancer cells

doi: 10.1016/j.isci.2024.111269

Figure Lengend Snippet:

Article Snippet: Recombinant human IGF2 , Sino Biological , Cat#13032-HNAY.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: Reprogramming of fibroblasts into cancer-associated fibroblasts via IGF2-mediated autophagy promotes metastasis of lung cancer cells

doi: 10.1016/j.isci.2024.111269

Figure Lengend Snippet: PCR primer sequences

Article Snippet: Recombinant human IGF2 , Sino Biological , Cat#13032-HNAY.

Techniques:

Journal: The FASEB Journal

Article Title: Insulin‐Like Growth Factor 2 Reverses Synaptic and Cognitive Deficits in Fetal Growth Restriction Mice

doi: 10.1096/fj.202402934R

Figure Lengend Snippet: FGR induces hippocampal Igf2 downregulation by DNA hypomethylation, and neuronal Igf2 deletion impairs synaptic plasticity. (A) qPCR analysis of Igf family mRNA levels in the hippocampi from 8‐week‐old NS and FGR mice. n = 6 mice in each group. (B, C) WB representative picture (B) and analysis (C) of IGF2 protein levels in the hippocampi at Postnatal Day 1, 1, 4, 8 weeks from NS and FGR mice. n = 3 mice in each group. (D) Bisulfite methylation sequencing of ICR in the hippocampi from 8‐week‐old NS and FGR mice. n = 9 mice in each group. (E, F) qPCR analysis of H19 (E) and Dnmt1 (F) mRNA levels in the hippocampi of 8‐week‐old NS and FGR mice. n = 6 mice in each group. (G) WB analysis of IGF2 protein level in the hippocampi of Ctrl and IKO mice. n = 4 mice in each group. (H) LTP measurements were performed in the CA1 region of acute slices from Ctrl and IKO mice. n = 3 in each group. (I) Representative Golgi staining images of the hippocampal DG neurons from Ctrl and IKO mice. Scale bar, 100 μm. (J and K) Quantification of the dendritic branches (J) and total dendritic length (K) of the hippocampal DG neurons. n = 6 mice in each group, 4–5 neurons in each mouse. (L) Representative Golgi staining images of hippocampal DG neuronal dendritic spines. Scale bar, 5 μm. (M) Quantification of the dendritic spine density. n = 6 mice in each group, 4–5 spine pictures in each mouse. The data are shown as mean ± SEM. Two‐way ANOVA with Sidak's multiple comparisons test (H) or unpaired Student's t ‐test (A, C–F, H, J, K, M). ** p < 0.01 and *** p < 0.001. See also Figures and .

Article Snippet: Igf2 recombination protein (CF61, 50 ng/mL, Novoprotein, China) was administered to the medium on Day 3, 9, or 12.

Techniques: Methylation, Sequencing, Staining

Journal: The FASEB Journal

Article Title: Insulin‐Like Growth Factor 2 Reverses Synaptic and Cognitive Deficits in Fetal Growth Restriction Mice

doi: 10.1096/fj.202402934R

Figure Lengend Snippet: Increasing Igf2 rescues dendritic, synaptic, and memory impairments in cultured FGR hippocampal neurons and FGR mice. (A, B) Representative fluorescence images (A) and Sholl analysis of dendritic morphology (B) of cultured mouse hippocampal neurons from NS and FGR mice added with IGF2 protein or solvent control (Ctrl). Scale bar, 100 μm. n = 3 mice in each group, 8–10 neurons in each experiment. (C–E) Representative fluorescence images of synaptic sites (C), quantification of the percentage of intact synaptic contacts (D), and the total spine density (E) in primary hippocampal neurons of four groups. Scale bar, 20 μm. n = 3 in each group, 7–10 spine pictures in each experiment. (F) Representative Golgi staining images of the hippocampal DG neurons from NS and FGR mice injected with AAV‐Igf2 virus or its control. Scale bar, 50 μm. (G and H) Quantification of the dendritic branches (G) and total dendritic length (H) of the hippocampal DG neurons in four groups. n = 6 mice in each group, 4–7 neurons in each mouse. (I and J) Representative Golgi staining of dendritic spines (I) and dendritic spine density (J) in four groups. Scale bar, 20 μm. n = 9 mice in each group, 9–10 spine pictures in each mouse. (K–N) MWM performance in four groups. (NS Ctrl: n = 12, NS Igf2: n = 11, FGR Ctrl: n = 12, FGR Igf2: n = 10). (K) Escape latency. (L) Time spent in each quadrant. The number of platform crossings was recorded 24 h (M) and 3 weeks (N) after the final training session. (O and P) Discrimination index (O) and object preferences (P) of the NOR test. (NS Ctrl: n = 13, NS Igf2: n = 11, FGR Ctrl: n = 13, FGR Igf2: n = 13). (Q and R) Freezing behavior in the 24 h (Q) and 7 days (R) fear recall test of the CFC test. n = 10 mice in each group. The data are shown as mean ± SEM. Two‐way ANOVA with Sidak's multiple comparisons test (B, K, L and P), or one‐way ANOVA with Tukey's multiple comparisons test (D, E, G, H, J, M–O, Q, R). * p < 0.05, ** p < 0.01, and *** p < 0.001. See also Figure .

Article Snippet: Igf2 recombination protein (CF61, 50 ng/mL, Novoprotein, China) was administered to the medium on Day 3, 9, or 12.

Techniques: Cell Culture, Fluorescence, Solvent, Control, Staining, Injection, Virus

Journal: The FASEB Journal

Article Title: Insulin‐Like Growth Factor 2 Reverses Synaptic and Cognitive Deficits in Fetal Growth Restriction Mice

doi: 10.1096/fj.202402934R

Figure Lengend Snippet: Running exercise elevates hippocampal Igf2 expression and ameliorates cognitive defects in FGR mice. (A) qPCR analysis of Igf2 and Igf1 expression levels in the hippocampus of NS and FGR mice at 24 h after sedentary (Sed) or exercised (Run) treatment. n = 3 in each group. (B and C) qPCR analysis of Igf2, Igf1(B) and Dnmt1 (C) expression levels in the hippocampus of NS and FGR mice at 2 months after exercise training. n = 6 in each group. (D) Bisulfate methylation sequencing analysis of CTS1 in mouse hippocampi at 2 months after exercise training. n = 9 mice in each group. (E) Representative Golgi staining images of the hippocampal DG neurons from NS and FGR mice after sedentary (Sed) or exercised (Run) treatment. Scale bar, 50 μm. (F and G) Quantification of the dendritic branches (F) and total dendritic length (G) of the hippocampal DG neurons in four groups. n = 6 mice in each group, 4–6 neurons in each mouse. (H and I) Representative Golgi staining dendritic spines (H) and dendritic spine density (I) in four groups. Scale bar, 20 μm. n = 9 mice in each group, 9–11 spine pictures in each mouse. (J–M) MWM performance in four groups. (NS Sed: n = 12, NS Run: n = 15, FGR Sed: n = 14, FGR Run: n = 12). (J) Escape latency. (K) Time spent in each quadrant. The number of platform crossings was recorded 24 h (L) and 3 weeks (M) after the final training session. (N and O) Discrimination index (N) and object preferences (O) of the NOR test. (NS Sed: n = 11, NS Run: n = 12, FGR Sed: n = 12, FGR Run: n = 12). (P and Q) Freezing behavior in the 24 h (P) and 7 days (Q) fear recall test of the CFC test (NS Sed: n = 14, NS Run: n = 11, FGR Sed: n = 13, FGR Run: n = 10). The data are shown as mean ± SEM. One‐way ANOVA with Tukey's multiple comparisons test (A–D, F, G, I, L–N, P, Q), or two‐way ANOVA with Sidak's multiple comparisons test (J, K, O) * p < 0.05, ** p < 0.01, and *** p < 0.001. See also Figure .

Article Snippet: Igf2 recombination protein (CF61, 50 ng/mL, Novoprotein, China) was administered to the medium on Day 3, 9, or 12.

Techniques: Expressing, Methylation, Sequencing, Staining

Journal: The FASEB Journal

Article Title: Insulin‐Like Growth Factor 2 Reverses Synaptic and Cognitive Deficits in Fetal Growth Restriction Mice

doi: 10.1096/fj.202402934R

Figure Lengend Snippet: Running exercise rescues impaired cognition in FGR mice via hippocampal Igf2. (A) Schematics of AAV9 construct knockdown of Igf2 expression and flow diagram of mouse experiments. (B, C) WB representative picture (B) and analysis (C) of IGF2 protein level in the hippocampal DGs of mice injected with AAV‐shIgf2 virus and its control. n = 3 in each group. (D) Representative Golgi staining images of the hippocampal DG neurons from FGR mice injected with control virus after sedentary treatment (FGR‐Ctrl‐Sed) or exercised treatment (FGR‐Ctrl‐Run), or injected with AAV‐shIgf2 virus after sedentary treatment (FGR‐shIgf2‐Sed) or exercised treatment (FGR‐shIgf2‐Run). Scale bar, 100 μm. (E and F) Quantification of the dendritic branches (E) and total dendritic length (F) of the hippocampal DG neurons in four groups. n = 6 mice in each group, 4–5 neurons in each mouse. (G and H) Representative Golgi staining dendritic spines (G) and dendritic spine density (H) in four groups. Scale bar, 5 μm. n = 6 mice in each group, 5 spine pictures in each mouse. (I–L) MWM performance in four groups. n = 8 mice in each group. (I) Escape latency. (J) Time spent in each quadrant. The number of platform crossings was recorded at 24 h (K) and 3 weeks (L) after the final training session. (M and N) Discrimination index (M) and object preferences (N) of the NOR test. n = 8 mice in each group. (O and P) Freezing behavior in the 24 h (O) and 7 days (P) fear recall test of the CFC test. n = 8 mice in each group. The data are shown as mean ± SEM. One‐way ANOVA with Tukey's multiple comparations test (E, F, H, K–M, O and P), two‐way ANOVA with Sidak's multiple comparisons test (I, J and N) or unpaired Student's t ‐test (C). * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: Igf2 recombination protein (CF61, 50 ng/mL, Novoprotein, China) was administered to the medium on Day 3, 9, or 12.

Techniques: Construct, Knockdown, Expressing, Injection, Virus, Control, Staining

Journal: The Journal of Reproduction and Development

Article Title: The imprinted Igf2-Igf2r axis is critical for exosome biogenesis during the early development of bovine placenta

doi: 10.1262/jrd.2024-081

Figure Lengend Snippet: Histological analysis of cotyledon villi of bovine placenta at 2, 3, and 4 months of gestation. (A) The cotyledon villi at 2, 3, and 4 months of pregnancy were collected for HE staining; the black arrows indicate mononuclear trophoblasts (MNCs) and binuclear trophoblasts (BNCs), Scale bars = 100 μm. Plasma was collected from pregnant dairy cows at 60, 90, and 120 days gestation, and the secretion of IGF2 (B) and IGF2R (C) was detected using ELISA kits according to the manufacturer’s recommendations. Data represent the mean ± standard deviations from six independent experiments. Bars with different letters indicate significant differences (* P < 0.05).

Article Snippet: After incubation with deionized water at room temperature for 10 min, after rinsing with PBS buffer, TSG101 (1:200 dilution, bs-1365R, Bioss, Beijing, China), Rab11 (1:200 dilution, 15903-1-AP, Proteintech, ​ Chicago, IL, USA), CD63 (1:200 dilution; bs-1523R; Bioss), CD9 (1:200 dilution; bs-2486R; Bioss), IGF2R (1:200 dilution; 14364S; Cell Signaling Technology, Danvers, MA, USA) and IGF2 (1:200 dilution; 12220-1-AP; Proteintech) were added, incubated at room temperature for 60 min, rinsed with PBS buffer, and then incubated with enzyme-labeled sheep anti-mouse IgG polyclonal antibody (1:200 dilution; HS201; TransGen Biotech, Beijing, China) at room temperature for 30 min.

Techniques: Staining, Enzyme-linked Immunosorbent Assay

Journal: The Journal of Reproduction and Development

Article Title: The imprinted Igf2-Igf2r axis is critical for exosome biogenesis during the early development of bovine placenta

doi: 10.1262/jrd.2024-081

Figure Lengend Snippet: Localization and expression of IGF2, IGF2R, and exosomal marker proteins. (A) Immunohistochemical assays were utilized to detect the localization of IGF2, IGF2R, and exosomal marker proteins CD9, CD63, TSG101, and Rab11 in the cotyledon villi of bovine placenta at 2, 3, and 4 months of gestation. Scale bar = 50 μm. (B) Western blot analysis was utilized to detect the expressions of IGF2, IGF2R, and exosomal marker proteins CD63, CD9, TSG101, and Rab11 in the cotyledon villi of bovine placenta at 2, 3, and 4 months of gestation. (C) Band intensity measurement of the IGF2, IGF2R, and exosomal marker proteins CD63, CD9, TSG101, and Rab11 to β-actin were determined by densitometry. The data represent the mean ± standard deviations from three independent experiments. Bars with different letters indicate statistically significant differences (* P < 0.05).

Article Snippet: After incubation with deionized water at room temperature for 10 min, after rinsing with PBS buffer, TSG101 (1:200 dilution, bs-1365R, Bioss, Beijing, China), Rab11 (1:200 dilution, 15903-1-AP, Proteintech, ​ Chicago, IL, USA), CD63 (1:200 dilution; bs-1523R; Bioss), CD9 (1:200 dilution; bs-2486R; Bioss), IGF2R (1:200 dilution; 14364S; Cell Signaling Technology, Danvers, MA, USA) and IGF2 (1:200 dilution; 12220-1-AP; Proteintech) were added, incubated at room temperature for 60 min, rinsed with PBS buffer, and then incubated with enzyme-labeled sheep anti-mouse IgG polyclonal antibody (1:200 dilution; HS201; TransGen Biotech, Beijing, China) at room temperature for 30 min.

Techniques: Expressing, Marker, Immunohistochemical staining, Western Blot

Journal: The Journal of Reproduction and Development

Article Title: The imprinted Igf2-Igf2r axis is critical for exosome biogenesis during the early development of bovine placenta

doi: 10.1262/jrd.2024-081

Figure Lengend Snippet: Knockdown of Igf2r gene in BTCs represses the expression of exosome biogenesis-related proteins in BTCs and BTCs-derived exosomes. (A) The expressions of IGF2, IGF2R, CD63, CD9, TSG101, and Rab11 proteins in Igf2r knockdown BTCs were detected using western blot analysis. (B) Quantification of band intensities was determined using densitometric analysis. The data represent the mean ± standard deviations from three independent experiments. Bars with different letters indicate statistically significant differences (* P < 0.05). (C) The expressions of CD63, CD9, TSG101, and Rab11 proteins in Igf2r knockdown BTCs-derived exosomes were identified with western blot analysis. (D) Quantification of band intensities was determined using densitometric analysis. The data represent the mean ± standard deviations from three independent experiments. Bars with different letters indicate statistically significant differences (* P < 0.05).

Article Snippet: After incubation with deionized water at room temperature for 10 min, after rinsing with PBS buffer, TSG101 (1:200 dilution, bs-1365R, Bioss, Beijing, China), Rab11 (1:200 dilution, 15903-1-AP, Proteintech, ​ Chicago, IL, USA), CD63 (1:200 dilution; bs-1523R; Bioss), CD9 (1:200 dilution; bs-2486R; Bioss), IGF2R (1:200 dilution; 14364S; Cell Signaling Technology, Danvers, MA, USA) and IGF2 (1:200 dilution; 12220-1-AP; Proteintech) were added, incubated at room temperature for 60 min, rinsed with PBS buffer, and then incubated with enzyme-labeled sheep anti-mouse IgG polyclonal antibody (1:200 dilution; HS201; TransGen Biotech, Beijing, China) at room temperature for 30 min.

Techniques: Knockdown, Expressing, Derivative Assay, Western Blot

Journal: The Journal of Reproduction and Development

Article Title: The imprinted Igf2-Igf2r axis is critical for exosome biogenesis during the early development of bovine placenta

doi: 10.1262/jrd.2024-081

Figure Lengend Snippet: Repression of IGF2 protein expression inhibits the expression of exosomal marker proteins in BTCs and BTC-derived exosomes. (A) The expressions of IGF2, IGF2R, CD63, CD9, TSG101, and Rab11 proteins in BTCs treated with chromeceptin for 24 h were detected with western blot analysis. (B) Quantification of band intensities was determined using densitometric analysis. The data represent the mean ± standard deviations from three independent experiments. Bars with different letters indicate statistically significant differences (* P < 0.05). (C) The expressions of CD63, CD9, TSG101, and Rab11 proteins in exosomes derived from BTCs treated with chromeceptin were detected with western blot analysis. (D) Quantification of band intensities was determined using densitometric analysis. The data represent the mean ± standard deviations from three independent experiments. Bars with different letters indicate statistically significant differences (* P < 0.05).

Article Snippet: After incubation with deionized water at room temperature for 10 min, after rinsing with PBS buffer, TSG101 (1:200 dilution, bs-1365R, Bioss, Beijing, China), Rab11 (1:200 dilution, 15903-1-AP, Proteintech, ​ Chicago, IL, USA), CD63 (1:200 dilution; bs-1523R; Bioss), CD9 (1:200 dilution; bs-2486R; Bioss), IGF2R (1:200 dilution; 14364S; Cell Signaling Technology, Danvers, MA, USA) and IGF2 (1:200 dilution; 12220-1-AP; Proteintech) were added, incubated at room temperature for 60 min, rinsed with PBS buffer, and then incubated with enzyme-labeled sheep anti-mouse IgG polyclonal antibody (1:200 dilution; HS201; TransGen Biotech, Beijing, China) at room temperature for 30 min.

Techniques: Expressing, Marker, Derivative Assay, Western Blot

Journal: Aging Cell

Article Title: Premature cognitive decline in a mouse model of tuberous sclerosis

doi: 10.1111/acel.14318

Figure Lengend Snippet: Hippocampal administration of recombinant IGF2 normalizes both memory consolidation and relative cfos expression in aged Tsc2 +/− mice. (a) Schematic overview of 7d NORT including IGF2 administration on day 2 within 2 h after second training phase. (b), 7d NORT approach of aged Tsc2 mutant mice 7 days after IGF2 injection showed a significant enhancement of memory consolidation measured as discrimination index compared to the PBS‐injected mutant mice (two tailed t test: P (baseline) = 5,89e‐9, n (WT) = 32, n ( Tsc2+/− ) = 34); p (PBS) = 0.0066, n (WT) = 13, n ( Tsc2 +/− ) = 11; p (IGF2) = 0.4433, n (WT) = 18, n ( Tsc2 +/− ) = 17. (c) Relative mRNA expression levels of cfos in the hippocampus of aged Tsc2 mutants after IGF2 administration compared to the control group. No significant difference in Tsc2 +/− mice compared to wildtype controls after IGF2 treatment (two‐tailed t test: p (PBS) = 0.0004, n (WT) = 4, n ( Tsc2 +/− ) = 7); p (IGF2) = 0.2329, n (WT) = 6, n ( Tsc2 +/− ) = 8. Values were normalized against Gapdh and are presented as mean ± SEM, * p < 0.05, *** p < 0.001. Quantification was performed using Excel.

Article Snippet: A single dose of recombinant mouse IGF2 protein (R&D Systems, #792‐MG) was stereotactically injected into the hippocampus of 8–10 months old Tsc2 +/− mice and wildtype littermates after the training phase 2 (day 2) of the 7 days NORT (Figure ).

Techniques: Recombinant, Expressing, Mutagenesis, Injection, Two Tailed Test, Control