stc2  (R&D Systems)

Journal: Life Science Alliance

Article Title: Identification and characterization of a membrane receptor that binds to human STC1

doi: 10.26508/lsa.202201497

Figure Lengend Snippet: (A) hSTC1, (B) hSTC2, and (C) hCREG binding to human IGF2R.

Article Snippet: To perform kinetic affinity assays, hSTC1-His, human His-tagged STC2 (hSTC2-His, 9405-SO-050; R&D Systems), and human His-tagged cellular repressor of E1A-stimulated gene (hCREG-His, 2380-CR-025/CF; R&D Systems) were prepared at 10 μg/ml in a running buffer, followed by a twofold serial dilution to give 10, 5, 2.5, 1.25, 0.625, 0.313, 0.156, 0.078, 0.039, and 0.019 μg/ml solutions.

Techniques: Binding Assay

Journal: Cell death discovery

Article Title: Stanniocalcin 2 drives malignant transformation of human glioblastoma cells by targeting SNAI2 and Matrix Metalloproteinases.

doi: 10.1038/s41420-022-01090-6

Figure Lengend Snippet: Fig. 1 Aggressive GBM expresses higher level of STC2. a, b Representative tissue microarray data of STC2 in clinical specimens (a, GL2082; b, GL208, Biomax). The scale bar represents 100 µm. STC2 staining score from 0-4 were analyzed in different tissues. N, normal tissues; A, astrocytoma; GBM, glioblastoma. One-way ANOVA, *p < 0.05, **p < 0.01, ***p < 0.001, n.s., no significance. c The levels of STC2 mRNA expression in low grade glioma (LGG) versus glioblastoma (GBM) tissues obtained from TCGA Research Network were analyzed. Unpaired t- test, **p < 0.01. d The mRNA expression of STC2 in various glioblastoma cell lines were analyzed. After normalizing with 18 S rRNA in each sample, the relative mRNA levels were calculated using A172 = 1. Means ± SD; n = 3 biological replicates. Oneway ANOVA, **p < 0.01, ***p < 0.001, n.s., no significance. e, f Representative images of Western blotting against STC2 and β-ACTIN in whole lysates of glioblastoma cell lines (e) and concentrated culture media (Conditioned media CM) (f).

Article Snippet: Human recombinant STC2 (Cat#9405-SO) was purchased from R&D Systems.

Techniques: Microarray, Staining, Expressing, Western Blot

Journal: Cell death discovery

Article Title: Stanniocalcin 2 drives malignant transformation of human glioblastoma cells by targeting SNAI2 and Matrix Metalloproteinases.

doi: 10.1038/s41420-022-01090-6

Figure Lengend Snippet: Fig. 2 Overexpression of STC2 results in invasive phenotypes of GBM cell lines. a, b STC2 mRNA (a) and protein (b) expression was validated after modulation of STC2. LN18 cell was transfected with pRS-shSTC2 to knockdown STC2 whereas A172 cell was transfected with Myc-DDK- tagged-STC2 to overexpress STC2. Means ± SD; n = 3 biological replicates; Student’s two tailed t-test, ***p < 0.001. c Cells were seeded to 6-well plates at a density of 1000 cells per well and colony formation was determined after 10 days. Absorbance at 590 nm was measured after dissolving with 10% acetic acid. Means ± SD; n = 3 biological replicates; Student’s two tailed t-test, ***p < 0.001. d Cell growth rates after STC2 modulation were determined by MTT assay. Means ± SD; n = 12 biological replicates. Student’s two tailed t-test, *p < 0.05, ***p < 0.001. e In vitro invasion and motility were determined onto Matrigel-coated or non-coated Transwell chambers for 48 h. Scale bar = 100 µm. Means ± SD; n = 5 biological replicates; Student’s two tailed t-test, *p < 0.05, ***p < 0.001. f In vitro migration was determined for 24 h after STC2 modulation. Scale bar = 100 µm.

Article Snippet: Human recombinant STC2 (Cat#9405-SO) was purchased from R&D Systems.

Techniques: Over Expression, Expressing, Transfection, Knockdown, Two Tailed Test, MTT Assay, In Vitro, Migration

Journal: Cell death discovery

Article Title: Stanniocalcin 2 drives malignant transformation of human glioblastoma cells by targeting SNAI2 and Matrix Metalloproteinases.

doi: 10.1038/s41420-022-01090-6

Figure Lengend Snippet: Fig. 3 Secreted STC2 induces invasive phenotypes of neighboring GBM cells. a Cell growth rates after treatment of recombinant STC2 (100 ng/mL) or STC2-containing CM were determined using MTT assay. Means ± SD; n = 12 biological replicates. Student’s two tailed t-test, *p < 0.05, **p < 0.01, ***p < 0.001. Significance was analyzed against shSTC2 in LN18, Con in A172. b, c) Colony formation (b) and in vitro invasion (c) were determined after treatment of recombinant STC2 (100 ng/mL) or STC2-containing CM. Absorbance at 590 nm was measured after dissolving with 10% acetic acid. Means ± SD; n = 3 biological replicates; One-way ANOVA, **p < 0.01, ***p < 0.001. d In vitro migration was determined after treatment of recombinant STC2 (100 ng/mL) or STC2-containing CM. Scale bar = 100 µm. e Migrating cells were visualized using fluorescent phalloidin (green) and DAPI (blue) staining. Scale bar = 50 µm.

Article Snippet: Human recombinant STC2 (Cat#9405-SO) was purchased from R&D Systems.

Techniques: Recombinant, MTT Assay, Two Tailed Test, In Vitro, Migration, Staining

Journal: Cell death discovery

Article Title: Stanniocalcin 2 drives malignant transformation of human glioblastoma cells by targeting SNAI2 and Matrix Metalloproteinases.

doi: 10.1038/s41420-022-01090-6

Figure Lengend Snippet: Fig. 4 STC2 targets SNAI2 and MMPs. a SNAI2, MMP-2, and MMP-9 mRNA expressions were validated after modulation of STC2 in LN18 or A172 cell lines. Means ± SD; n = 3 biological replicates; Student’s two tailed t-test, *p < 0.05, **p < 0.01, ***p < 0.001. b SNAI2 protein expression was validated after modulation of STC2. c Proteolytic activities of MMP-2 and MMP-9 were determined by gelatin zymography after STC2 modulation. d The correlation of SNAI2 expression with STC2 (left), the correlation of MMP-2 expression with SNAI2 (middle), and the correlation of MMP-9 expression with SNAI2 (right) were determined in the TCGA data. e SNAI2, MMP-2, and MMP-9 mRNA expressions were measured after treatment of recombinant STC2 (50, 100 ng/mL) or STC2-containing CM (50%). Means ± SD; n = 3; One-way ANOVA, *p < 0.05, **p < 0.01, ***p < 0.001, n.s., no significance. f SNAI2 protein expression was validated after treatment of recombinant STC2 (50, 100, 200 ng/mL) or STC2- containing CM (50, 100%). g In vitro migration was determined for 6 h after recombinant STC2 (100 ng/mL) treatment. Migrated cells were visualised by staining with fluorescent phalloidin and DAPI. Scale bar = 50 µm. h Proteolytic activities of MMP-2 and MMP-9 were determined by gelatin zymography after recombinant STC2 (50, 100, 200 ng/mL) or STC2-containing CM (50, 100%).

Article Snippet: Human recombinant STC2 (Cat#9405-SO) was purchased from R&D Systems.

Techniques: Two Tailed Test, Expressing, Zymography, Recombinant, In Vitro, Migration, Staining

Journal: Cell death discovery

Article Title: Stanniocalcin 2 drives malignant transformation of human glioblastoma cells by targeting SNAI2 and Matrix Metalloproteinases.

doi: 10.1038/s41420-022-01090-6

Figure Lengend Snippet: Fig. 5 Secreted STC2 regulates SNAI2 and MMPs through p38 MAPK pathway. a, b LN18 and A172-STC2 cells were treated with small molecule signaling pathway inhibitors (LGK974, 500 nM; LY3214996, 50 nM; Rapamycin, 10 nM; LY294002 1 µM; and SB202190, 10 µM) for 48 h and STC2 expression was detected by real-time PCR (a) and Western blot analysis (b). Means ± SD; n = 3; On-way ANOVA, ***ī< 0.001. c Wild- type A172 cells were co-treated with small molecule signaling pathway inhibitors and recombinant STC2 (100 ng/mL) for 48 h and SNAI2, MMP- 2, and MMP-9 mRNA expressions were determined by real-time PCR. The relative mRNA levels were calculated against vehicle control of each inhibitors. Small molecule inhibitors were added 30 min prior to recombinant STC2 treatment. Means ± SD; n = 3; Student’s two tailed t-test, *p < 0.05, **p < 0.01, ***p < 0.001. d A172 cells were treated with recombinant STC2 (100 ng/mL) or STC2-containing CM (50%) with or without p38 MAPK inhibitor SB202190 for 48 h. SNAI2 protein expression along with phosphorylated p38 (p-p38) and total p38 (t-p38) proteins were determined by Western blot analysis.

Article Snippet: Human recombinant STC2 (Cat#9405-SO) was purchased from R&D Systems.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Recombinant, Control, Two Tailed Test

Journal: Nature Communications

Article Title: Electrical modulation of transplanted stem cells improves functional recovery in a rodent model of stroke

doi: 10.1038/s41467-022-29017-w

Figure Lengend Snippet: a Heatmap of transcriptome changes in NPCs from electrical stimulation (NPC – unstimulated, NPC Stim – electrically stimulated; blue-downregulated, orange-upregulated). b Volcano plot demonstrating changes in genes with electrical stimulation (blue-downregulated, orange-upregulated). c Top Gene set enrichment pathways with STC2 as the leading edge. d qRT-PCR analysis indicated that STC2 in NPCs was upregulated by electrical stimulation. e ELISA study indicated that the level of STC2 protein after electrical stimulation (NPC Stim ) is much higher than that in non-stimulated NPCs (NPC). d , e Analyzed using a one-way ANOVA, followed by Tukey’s HSD post-hoc test with ** P < 0.01, **** P < 0.0001, data shown as mean ± SEM, n = 4.

Article Snippet: About 24 h prior to implantation, recombinant human STC2 protein (4 ng/mL of 1X PBS, Novus Biologicals) was loaded on to the osmotic pump (Azlet mini-osmotic pump model 2001, Braintree Scientific) and incubated at 37 C as per the manufacturer protocol.

Techniques: Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: Nature Communications

Article Title: Electrical modulation of transplanted stem cells improves functional recovery in a rodent model of stroke

doi: 10.1038/s41467-022-29017-w

Figure Lengend Snippet: a Vibrissae-forepaw (WP) and total neurological score (NS). There is a statistically significant difference in NPC Stim , STC2 UP , and Scramble KD+Stim groups versus all other groups starting at week 3 and onward for WP Scores and week 5 and onward for NS scores. STC2 KD and STC2 KD+Stim showed no significant behavioral recovery at any timepoint in either WP Scores or NS Scores. b Representative images of fluorescently labeled cells. Green indicates BrdU positive cells, whereas Blue indicates cell nucleus. Scale bar indicates 200 µm. c BrdU positive cells in certain areas of interest. Data are shown as mean ± SEM, n = 8 images/4 rats. d PAX6/BrdU or Nestin/BrdU positive cells in certain areas of interest. Data are shown as mean ± SEM, n = 16 images/4 rats. e Representative images of fluorescently labeled cells. Green indicates Nestin positive cells whereas Red indicates PAX6 positive cells. Blue indicates BrdU positive cells. Scale bar indicates 200 µm. a Analyzed using a Kruskal–Wallis test followed by post-hoc pairwise Mann–Whitney U test with Benjamini–Hochberg correction to control the false discovery rate at the 0.05 level. For both, * P < 0.01, ** P < 0.001, data shown as mean ± SEM, n = 10 per group. c , d Analyzed using a one-way ANOVA, followed by Tukey’s HSD post-hoc test with * P < 0.05, ** P < 0.01, and **** P < 0.0001.

Article Snippet: About 24 h prior to implantation, recombinant human STC2 protein (4 ng/mL of 1X PBS, Novus Biologicals) was loaded on to the osmotic pump (Azlet mini-osmotic pump model 2001, Braintree Scientific) and incubated at 37 C as per the manufacturer protocol.

Techniques: Labeling, MANN-WHITNEY, Control

Journal: Nature Communications

Article Title: Electrical modulation of transplanted stem cells improves functional recovery in a rodent model of stroke

doi: 10.1038/s41467-022-29017-w

Figure Lengend Snippet: a Schematic of the brain slice view (left) and top view (right) in the skull for STC2 protein delivery via mini-osmotic pump. The cannula of the osmotic pump is implanted at 0.8 mm posterior (Y) and 1.5 mm contralateral (X) to the bregma at a depth of 3.5 mm at 1-week after stroke. b Behavioral testing. Vibrissae-forepaw (WP) behavioral testing (left) and overall neurological score (right). Based on WP, STC2 group exhibited statistically significantly greater recovery beginning at 4 weeks post-stroke compared to other groups. c Endogenous neuroblasts. Representative images of DCX + /BrdU+ cells (left) and the cell count of number co-positive cells (right) at peri-infarct region. b Analyzed using a log-transform two-way repeated measures ANOVA which revealed a statistically significant interaction between the effects of treatment group and WP Score ( F [12, 162] = 7.058, P < 0.0001); followed by Dunnett’s multiple comparisons test. Neurological scores (NS) were analyzed using a Kruskal–Wallis test ( P = 0.018), followed by Dunn’s multiple comparisons test. For both, * P < 0.05, ** P < 0.01, data shown as mean ± SEM, n = 10 per group. c Analyzed using one-way ANOVA, followed by Tukey’s HSD post-hoc test with * P < 0.05. Data shown as mean ± SEM, n = 20 images/4 rats.

Article Snippet: About 24 h prior to implantation, recombinant human STC2 protein (4 ng/mL of 1X PBS, Novus Biologicals) was loaded on to the osmotic pump (Azlet mini-osmotic pump model 2001, Braintree Scientific) and incubated at 37 C as per the manufacturer protocol.

Techniques: Slice Preparation, Cell Counting