Journal: Scientific Reports

Article Title: Efficient and highly reproducible production of red blood cell-derived extracellular vesicle mimetics for the loading and delivery of RNA molecules

doi: 10.1038/s41598-024-65623-y

Figure Lengend Snippet: Production of miR-210-loaded RBCEVs and biological effect. ( A ) Absolute miR-210 concentrations have been calculated by a standard curve set up with synthetic RNA standards at 20–0.0002 nM. miR-210 concentration into loaded RBCEVs is compared to UL RBCEVs and to the starting concentration obtained into the mother RBCs (UL and L) before vesiculation. High loading efficiency can be appreciated. The amount of miR-210 found in UL RBCs and RBCEVs is due to the endogenous miRNA. Data are mean and SEM (n = 4). In ( B ), miR-210 relative quantification into HUVEC is reported. Relative quantification has been performed using U6 snRNA as a reference gene and UL sample as a control. miR-210 concentration found in HUVEC treated with L RBCEVs are compared to cells treated with UL RBCEVs and to cells transfected at different miRNA concentrations. Data are mean and SEM, n = 4 (Unpaired t-test; *two-tailed p-values < 0.05). In ( C ), the evaluation of the effect of miR210-loaded RBCEVs at the mRNA level is shown. Relative quantification of PTB1B mRNA has been performed using ATCB as reference gene and UL sample as control. PTP1B mRNA found in HUVEC treated with L RBCEVs is compared to cells treated with UL RBCEVs and to cells transfected at different miRNA concentrations. Data are mean and SEM, n = 4 (Unpaired t-test; *two-tailed p-values < 0.05). ( D ) Western blot of PTP1B in protein extracts from HUVEC treated with RBCEVs UL or L and transfected with different amounts of miR-210. Lanes 1–5: RBCEVs UL, RBCEVs L, miR-210 1 nM, 10 nM, and 50 nM plus Transit-2X. This figure has been cropped in order to report the most relevant results and the original blot is available in . In ( E ), the evaluation of the effect of miR210-loaded RBCEVs at the protein level. Quantification of PTP1B band normalized to total proteins. Data are the mean and SEM, n = 4 (Unpaired t-test; p-values * < 0.05, *** < 0.001, **** < 0.0001). ( F ) Glycolytic and mitochondrial ATP production rates obtained in both miR210-transfected and RBCEVs-treated HUVEC (Unpaired t-test; p-values *** < 0.001).

Article Snippet: For the PTP1B mRNA expression assay, the reaction was done with 1 μl of diluted cDNA, 10 μl of the same master mix as small RNAs, and 1 μl of the specific TaqMan® Gene Expression Assay (Hs00942477_m1, Catalog #4331182, ThermoFisher Scientific).

Techniques: Concentration Assay, Quantitative Proteomics, Control, Transfection, Two Tailed Test, Western Blot

Journal: Nature Communications

Article Title: Metal-organic polyhedra maintain the self-renewal of embryonic stem cells

doi: 10.1038/s41467-025-63811-6

Figure Lengend Snippet: A The photo of MOP-1 aqueous solution (left). Monodisperse structure of MOP-1 in H 2 O (right). Color codes: V, green; O, red; C, gray; N, blue; Cl, bright blue. The large pink sphere represents the free space inside the molecular cage. For clarity, H atoms were omitted. Schematic (right panel) was created with Diamond software. B Experimental and simulated PXRD patterns of MOP-1. C TEM image of MOP-1 (left). Scale bar, 20 nm. The particle size of MOP-1 in H 2 O (right). Three experiments were repeated independently with similar results. D The schematic diagram showed that the SHP-2 mediated STAT3 inactivation. Schematic diagram was created with Microsoft Office PowerPoint. E , F An in-depth mechanism investigation of mESC pluripotency control by MOP-1. Binding model from a global view of a complex composed of SHP-2 and MOP-1 illustrated by electrostatic surface potential ( E ). Binding modes are illustrated by ribbon diagrams of a complex composed of SHP-2 and MOP-1 (the left panel), a complex composed of SHP-2 with MOP (the middle panel) and a complex composed of SHP-2 with ZrMOP (the right panel). The top panel is the global view of the catalytic PTP structure of SHP-2, the bottom panel is the focused view of binding modes illustrated by the Ribbon diagrams ( F ). G Binding kinetics of MOP-1 (top panel) and MOP-2 (bottom panel) with SHP-2 were measured by the SPR assay. H The ICP-MS analysis of the binding quantity between MOPs and SHP-2 (mean ± s.e.m, n = 6). I The inhibition efficiency of SHP-2, JAK2, JAK1, SHP-1, PTP1B, Cyt c , ACP and lipase by MOP-1 at a concentration of 2 μM using enzyme assay (mean ± s.e.m, n = 3). Data in ( H ) and ( I ) are analyzed by one-way ANOVA. **** P < 0.0001, the binding between MOP-1 and SHP-2 vs. the binding between MOP-2 and SHP-2, relative activity of SHP-2 vs. relative activity of JAK2, JAK1, PTP1B, Cyt c , ACP and lipase. *** P < 0.001, relative activity of SHP-2 vs. relative activity of SHP-1.

Article Snippet: The binding kinetics and affinity of MOP-1 or MOP-2 to SHP-2 (MCE, HY-P700618), SHP-1 (MCE, HY- P71141 ), JAK1 (MCE, HY-P700583), JAK2 (MCE, HY-P701102) and PTP1B (MCE, HY- P73685 ) were analyzed by SPR (Biacore 8 K, Cytiva).

Techniques: Software, Control, Binding Assay, SPR Assay, Inhibition, Concentration Assay, Enzymatic Assay, Activity Assay

Journal: Cells

Article Title: PTPN1 Deficiency Modulates BMPR2 Signaling and Induces Endothelial Dysfunction in Pulmonary Arterial Hypertension

doi: 10.3390/cells12020316

Figure Lengend Snippet: PTPN1 is a novel regulator of BMPR2 signaling pathway. A siRNA-mediated HTS ( n = 22,124) was performed to identify possible BMPR2 signaling modifiers in mouse myoblastoma BRE-ID1-LUC incorporated reporter cells. After 48 h knockdown of the genes, cells were treated with BMP4 to activate the signaling for two hours and then measured for ID1-linked luciferase levels. A colorimetric trypan blue cell viability was also performed. ( A ) Changes in ID1-linked luciferin expression ( n = 22,124) ( X -axis) versus cell viability ( Y -axis) were plotted. Red dots denote the pre-selected protein tyrosine kinases (PTPs) selected from all major PTPs. ( B ) Changes in ID1-linked luciferin expression of the selected PTPs (% changes from NTi). ( C ) Selected PTPs tested with individual siRNA (reconstructed from the secondary screening data n = 96 from ). Data represented as mean ± SEM ( n = 2–3). ( D – J ) PTPN1, BMPR2 and ID1 expression were measured by qRT-PCR in PAECs silenced to either PTPN1 or BMPR2 for 48 h. Data represented as mean ± SEM ( n = 3), student t-test. * p < 0.05, ** p < 0.01, **** p < 0.0001.

Article Snippet: Total RNA was then converted to cDNA using a high-capacity cDNA reverse transcription kit (Cat # 4368813, Applied Biosystems TM , Foster City, CA, USA) and gene expression was assessed by TaqMan qRT-PCR with targeted TaqMan assay probes, human PTPN1 (Hs00942477), GAPDH (Hs01786624_g1), 18S (Hs99999901_s1), BMPR2 (Hs00176148_m1), ID1 (Hs03676575_s1), rat PTPN1 (Rn01423685_m1), rat Gapdh (Rn01775763_g1), and TaqManTM 2× Universal PCR Master Mix (Cat # 4304437).

Techniques: Knockdown, Luciferase, Expressing, Quantitative RT-PCR

Journal: Cells

Article Title: PTPN1 Deficiency Modulates BMPR2 Signaling and Induces Endothelial Dysfunction in Pulmonary Arterial Hypertension

doi: 10.3390/cells12020316

Figure Lengend Snippet: PTPN1 deficiency induced endothelial dysfunction in PAECs. ( A ) siRNA-mediated knockdown of PTPN1 showed decreased PAEC viability, as assessed by MTT assay and hemocytometer countings. ( B ) PTPN1 silencing induced apoptosis, as evidenced by increased caspase 3/7 levels in PAECs. ( C ) PTPN1 silencing decreased ability of PAEC tube formation in a Matrigel tube formation assay. Angiogenesis was quantified in the images using ImageJ software. Total tube lengths were presented in µm. The number of nodes were counted in the analyzed area. Scale bar = 1000 µm. Data represented as mean ± SEM ( n = 3–6), student t-test. * p < 0.05, *** p < 0.001.

Article Snippet: Total RNA was then converted to cDNA using a high-capacity cDNA reverse transcription kit (Cat # 4368813, Applied Biosystems TM , Foster City, CA, USA) and gene expression was assessed by TaqMan qRT-PCR with targeted TaqMan assay probes, human PTPN1 (Hs00942477), GAPDH (Hs01786624_g1), 18S (Hs99999901_s1), BMPR2 (Hs00176148_m1), ID1 (Hs03676575_s1), rat PTPN1 (Rn01423685_m1), rat Gapdh (Rn01775763_g1), and TaqManTM 2× Universal PCR Master Mix (Cat # 4304437).

Techniques: Knockdown, MTT Assay, Tube Formation Assay, Software

Journal: Cells

Article Title: PTPN1 Deficiency Modulates BMPR2 Signaling and Induces Endothelial Dysfunction in Pulmonary Arterial Hypertension

doi: 10.3390/cells12020316

Figure Lengend Snippet: PTPN1 is downregulated in healthy human PAECs exposed to hypoxia and in the lung sugen5416/hypoxia-induced PH rats. ( A ) 150,000 PAECs were seeded onto 6-well plates and exposed to 72 h of hypoxia. After that, PTPN1 expression was measured by qRT-PCR. Induction of hypoxia was verified by measuring VEGF expression by qRT-PCR . ( B , C ) PTPN1 mRNA and protein expression was also measured in the lung of sugen5416/hypoxia rate models by qRT-PCR and western blotting (please see the PH model description and phenotypes data in ). Data are represented as mean ± standard error mean, n = 3–5; student t -test was performed to compare data between two samples. **** p < 0.0001. ns, not significant.

Article Snippet: Total RNA was then converted to cDNA using a high-capacity cDNA reverse transcription kit (Cat # 4368813, Applied Biosystems TM , Foster City, CA, USA) and gene expression was assessed by TaqMan qRT-PCR with targeted TaqMan assay probes, human PTPN1 (Hs00942477), GAPDH (Hs01786624_g1), 18S (Hs99999901_s1), BMPR2 (Hs00176148_m1), ID1 (Hs03676575_s1), rat PTPN1 (Rn01423685_m1), rat Gapdh (Rn01775763_g1), and TaqManTM 2× Universal PCR Master Mix (Cat # 4304437).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Cells

Article Title: PTPN1 Deficiency Modulates BMPR2 Signaling and Induces Endothelial Dysfunction in Pulmonary Arterial Hypertension

doi: 10.3390/cells12020316

Figure Lengend Snippet: PTPN1 is downregulated in the blood but not in the PAECs of PAH patients. We analyzed RNA-seq data of PTPN1 expression in the whole blood ((n = 72 healthy and n = 359 PAH), for subject characteristics, please see ) ( A ) and in PAECs (n = 9/group, (GSE0126262, )) of PAH patients ( B ). TPM values for PTPN1 expression is shown in the graphs. PTPN1 was correlated with expression of BMPR2, SMAD5, and SMAD9 in healthy and PAH PAECs (GSE0126262) ( C – E ).

Article Snippet: Total RNA was then converted to cDNA using a high-capacity cDNA reverse transcription kit (Cat # 4368813, Applied Biosystems TM , Foster City, CA, USA) and gene expression was assessed by TaqMan qRT-PCR with targeted TaqMan assay probes, human PTPN1 (Hs00942477), GAPDH (Hs01786624_g1), 18S (Hs99999901_s1), BMPR2 (Hs00176148_m1), ID1 (Hs03676575_s1), rat PTPN1 (Rn01423685_m1), rat Gapdh (Rn01775763_g1), and TaqManTM 2× Universal PCR Master Mix (Cat # 4304437).

Techniques: RNA Sequencing, Expressing

Journal: Cells

Article Title: PTPN1 Deficiency Modulates BMPR2 Signaling and Induces Endothelial Dysfunction in Pulmonary Arterial Hypertension

doi: 10.3390/cells12020316

Figure Lengend Snippet: Correlation of PTPN1 expression with SMAD1 and ID1 in PAECs collected from healthy and PAH patients (GSE0126262). PTPN1 expression was not significantly correlated with SMAD1 ( A ) and ID1 ( B ) in the PAECs of PAH patients and healthy controls.

Article Snippet: Total RNA was then converted to cDNA using a high-capacity cDNA reverse transcription kit (Cat # 4368813, Applied Biosystems TM , Foster City, CA, USA) and gene expression was assessed by TaqMan qRT-PCR with targeted TaqMan assay probes, human PTPN1 (Hs00942477), GAPDH (Hs01786624_g1), 18S (Hs99999901_s1), BMPR2 (Hs00176148_m1), ID1 (Hs03676575_s1), rat PTPN1 (Rn01423685_m1), rat Gapdh (Rn01775763_g1), and TaqManTM 2× Universal PCR Master Mix (Cat # 4304437).

Techniques: Expressing

Journal: Toxicology and applied pharmacology

Article Title: PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

doi: 10.1016/j.taap.2015.12.021

Figure Lengend Snippet: Fig. 2. Up-regulation of PTP1B in the fibrotic livers from CCl4-treated mice and down-regulation in the spontaneous recovery livers, n = 12 per group. a. The level of PTP1B and α-SMA were analyzed by immune-histochemistry in control, model and recovery livers. Representative views from each group are presented (×100). b. The expression of PTP1B, Col1α1 and α-SMA mRNA was assessed by Real-time PCR. The results are expressed as relative expression against negative control expression. The graph represents mRNA expression as mean ± SD from three independent experiments.⁎⁎p b 0.01 vs control; #p b 0.05, ##p b 0.01 vs model. c. The expression of PTP1B, Col1α1 and α-SMA protein was assessed by Western blot. The results are expressed as relative expression against negative control expression. Representative blots of three independent experiments are shown. ⁎⁎p b 0.01 vs control; #p b 0.05, ##p b 0.01 vs model. d. Double immunofluorescence staining was performed to determine the colocalization of PTP1B (green) and α-SMA (red) in CCl4-induced mouse liver fibrosis models. Representative views from each group are presented (×100).

Article Snippet: Rabbit polyclonal anti-aSMA (Proteintech, USA) was diluted at 1:500, Rabbit polyclonal anti -PTP1B and Col1α1 (Boster, China) were diluted 1:200; mouse monoclonal anti-β-actin was diluted 1:200.

Techniques: Control, Expressing, Real-time Polymerase Chain Reaction, Negative Control, Western Blot, Staining

Journal: Toxicology and applied pharmacology

Article Title: PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

doi: 10.1016/j.taap.2015.12.021

Figure Lengend Snippet: Fig. 3. Increased expression of PTP1B during HSC activation induced by TGF-β1. a. HSC-T6 cells were stimulated with different concentrations of TGF-β1 (0, 5, 10 ng/ml) for 24 h. The mRNA level of PTP1B, Col1a1 and α-SMA were examined by Real-time PCR; the protein level of PTP1B, Col1a1 and α-SMA were assessed by Western blot. The results are expressed as relative expression against negative control expression. Data represent the mean ± SD from three independent experiments. ⁎p b 0.05; ⁎⁎p b 0.01 vs 0 ng/ml group. b. HSC-T6 cells were stimulated with TGF-β1 (10 ng/ml) for 0,12,24 and 48 h. The mRNA level of PTP1B, Col1a1 and α-SMA were examined by Real-time PCR; the protein level of PTP1B, Col1a1 and α-SMA were assessed by Western blot. The results are expressed as relative expression against negative control expression. Data represent the mean ± SD from three independent experiments. ⁎p b 0.05; ⁎⁎p b 0.01 vs 0 h group.

Article Snippet: Rabbit polyclonal anti-aSMA (Proteintech, USA) was diluted at 1:500, Rabbit polyclonal anti -PTP1B and Col1α1 (Boster, China) were diluted 1:200; mouse monoclonal anti-β-actin was diluted 1:200.

Techniques: Expressing, Activation Assay, Real-time Polymerase Chain Reaction, Western Blot, Negative Control

Journal: Toxicology and applied pharmacology

Article Title: PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

doi: 10.1016/j.taap.2015.12.021

Figure Lengend Snippet: Fig. 4. Effect of PTP1B on cell proliferation, cell cycle and apoptosis in TGF- β1-treated HSC-T6 cells. a. Reduced expression of PTP1B in HSC-T6 cells by PTP1B-siRNA. The results are expressed as relative expression against control expression. Data represent the mean ± SD from three independent experiments. ⁎⁎p b 0.01 vs Scrambled-siRNA group. b. Inhibition of PTP1B significantly decreased cell proliferation of TGF-β1-treated HSC-T6 cells. Proliferation of HSC-T6 cells was tested by MTT assay. Data represent the mean ± SD from three independent experiments.⁎p b 0.05 vs TGF-β1 + Scrambled-siRNA group. c. Effect of PTP1B on cell cycle in TGF-β1-treated HSC-T6 cells. The results are expressed as relative expression against control expression without treatment. Representative images of three independent experiments are shown. ##p b 0.01vs TGF-β1 + Scrambled-siRNA group. d. Effect of PTP1B on apoptosis in TGF- β1-treated HSC-T6 cells. The results are expressed as relative expression against control expression without treatment. One representative experiment of the three independent experiments is demonstrated.

Article Snippet: Rabbit polyclonal anti-aSMA (Proteintech, USA) was diluted at 1:500, Rabbit polyclonal anti -PTP1B and Col1α1 (Boster, China) were diluted 1:200; mouse monoclonal anti-β-actin was diluted 1:200.

Techniques: Expressing, Control, Inhibition, MTT Assay

Journal: Toxicology and applied pharmacology

Article Title: PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

doi: 10.1016/j.taap.2015.12.021

Figure Lengend Snippet: Fig. 5. Blockade of PTP1B decreased TGF-β1-induced expressions of α-SMA and Col1 α1 in HSC-T6 cells. After PTP1B-siRNA or Scrambled-siRNA transfection, then exposure HSC-T6 cells to TGF-β1 (10 ng/ml) for 24 h. (a) Real-time PCR were performed to examine the mRNA level of PTP1B, Col1a1 and α-SMA. (b) Western blot was performed to assess the protein level of PTP1B, Col1a1 and α-SMA. The results are expressed as relative expression against negative control expression. Data represent the mean ± SD from three independent experiments. ⁎⁎p b 0.01 vs Control; ##p b 0.01 vs Scrambled-siRNA.

Article Snippet: Rabbit polyclonal anti-aSMA (Proteintech, USA) was diluted at 1:500, Rabbit polyclonal anti -PTP1B and Col1α1 (Boster, China) were diluted 1:200; mouse monoclonal anti-β-actin was diluted 1:200.

Techniques: Transfection, Real-time Polymerase Chain Reaction, Western Blot, Expressing, Negative Control, Control

Journal: Toxicology and applied pharmacology

Article Title: PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

doi: 10.1016/j.taap.2015.12.021

Figure Lengend Snippet: Fig. 6. Decreased expression of PTP1B in inactivated HSC-T6 cells. HSC-T6 cells were activated by TGF-β1 (10 ng/ml, 24 h), then they were treated with MDI for 48 h to be inactivated. The mRNA level of PTP1B, Col1a1 and α-SMA were examined by Real-time PCR (a); the protein level of PTP1B, Col1a1 and α-SMA were assessed by Western blot (b). The results are expressed as relative expression against negative control expression. Data represent the mean ± SD from three independent experiments. ⁎p b 0.05, ⁎⁎p b 0.01 vs control; #p b 0.05, ##p b 0.01 vs TGF-β1.

Article Snippet: Rabbit polyclonal anti-aSMA (Proteintech, USA) was diluted at 1:500, Rabbit polyclonal anti -PTP1B and Col1α1 (Boster, China) were diluted 1:200; mouse monoclonal anti-β-actin was diluted 1:200.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Negative Control, Control

Journal: Toxicology and applied pharmacology

Article Title: PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

doi: 10.1016/j.taap.2015.12.021

Figure Lengend Snippet: Fig. 7. Over-expression of PTP1B reversed the inactivation of HSC-T6 cells induced by MDI. HSC-T6 cells were activated by TGF-β1 (10 ng/ml, 24 h), then they were transfected with pCDNA3.1-PTP1B or pCDNA3.1-control plasmid. After that, cells were treated with MDI for 48 h to be inactivated. The mRNA level of PTP1B, Col1a1 and α-SMA were examined by Real-time PCR (a); the protein level of PTP1B, Col1a1 and α-SMA were assessed by Western blot (b). The results are expressed as relative expression against negative control expression. Data represent the mean ± SD from three independent experiments. ⁎p b 0.05, ⁎⁎p b 0.01 vs TGF-β1; #p b 0.05, ##p b 0.01 vs pCDNA3.1-control.

Article Snippet: Rabbit polyclonal anti-aSMA (Proteintech, USA) was diluted at 1:500, Rabbit polyclonal anti -PTP1B and Col1α1 (Boster, China) were diluted 1:200; mouse monoclonal anti-β-actin was diluted 1:200.

Techniques: Over Expression, Transfection, Control, Plasmid Preparation, Real-time Polymerase Chain Reaction, Western Blot, Expressing, Negative Control