Journal: International Journal of Molecular Sciences

Article Title: Selective Expression of a SNARE-Cleaving Protease in Peripheral Sensory Neurons Attenuates Pain-Related Gene Transcription and Neuropeptide Release

doi: 10.3390/ijms22168826

Figure Lengend Snippet: Pirt promoter drove exogenous gene expression selectively in sensory neurons. ( A ) Cultured mouse DRGs were incubated with 5.4 × 10 4 TU/well for 5 days before performing double immunofluorescent staining with antibodies for the LCA cleaved SNAP-25 A (green) and TRPV1 (red). The nuclei were stained blue with 4′, 6-Diamidino-2-phenylindole dihydrochloride. Bars, 50 μm. Cultured mouse DRGs ( B ) and spinal cord neurons (SCNs) ( C ) were infected by lentiPirtLCA with the indicated doses for 5 days before probing the SNAP-25 cleavage by the expressed LCA. Intact SNAP-25 and LCA cleaved form were probed by Western blotting with an antibody recognizing both intact and cleaved forms. ( D ) Quantified data derived from densitometric scanning of 3 replicate gels. Empty bars, virus free; hatched bars, 0.2 × 10 4 TU; filled bars, 5.4 × 10 4 TU. Data presented are mean ± SEM, from three independent cultures. One-way ANOVA with Bonferroni’s multiple comparison test demonstrated significant differences between the groups in DRGs: virus free vs. 0.2 × 10 4 TU p = 0.00003; virus free vs. 5.4 × 10 4 TU p < 0.00001; 0.2 × 10 4 TU vs. 5.4 × 10 4 TU p = 0.00119. In SCNs: virus free vs. 5.4 × 10 4 TU p = 0.01327. * p < 0.05; ** p < 0.01; *** p < 0.001, p > 0.05. Mouse DRGs ( E ), primary human keratinocytes (phKCs) ( F ) and Raw 264.7 cells ( G ) were infected with lentiPirtGFP for 5 days before probing the expression of GFP protein by Western blotting. β-tubulin III and β-actin proteins were probed as loading controls.

Article Snippet: RAW264.7 (Cat. No. CL-0190, Procell Life Science & Technology, Wuhan, China) and J774A.1 macrophage cells (Cat. No. CL-0370, Procell Life Science & Technology) and phKCs (Cat. No. 00192907, Lonza, Guangzhou, China) were cultured as previously described [ , ].

Techniques: Gene Expression, Cell Culture, Incubation, Staining, Infection, Western Blot, Derivative Assay, Virus, Comparison, Expressing

Journal: Nature Communications

Article Title: Hypomyelination in autism-associated neuroligin-3 mutant mice impairs parvalbumin interneuron excitability, gamma oscillations, and sensory discrimination

doi: 10.1038/s41467-025-61455-0

Figure Lengend Snippet: a Primary cultures of different cell types derived from the cortex of P0 mice by the mixed glial method or neurons from E16 mice. b qRT-PCR analysis of NL3 expression in different cell types in primary cultures (6 wells/samples from 3 mice cultures). c – e Immunostaining for NL3 across different stages of oligodendrocyte development in vitro revealed its expression in OPC (marked by NG2 (red)), immature OL (marked by O4 (red)), and mature OL (marked by MBP (red)). NL3 (cyan) staining was observed in the cell bodies (yellow arrows) and processes (white arrows) of NG2+ and O4+ cells, as well as on MBP+ myelin sheaths (blue arrows). f Immunostaining for NL3 (green) and PDGFRα (red) displaying the expression of NL3 in OPCs of white matter. NL3 staining is distributed along the proximal processes of OPCs (yellow arrow) as well as the distal processes (white arrow). g Immunostaining for NL3 (green) and CC1 (blue) displaying the expression of NL3 in mature OL in white matter. NL3 staining is mainly localized around the cytoplasm (white arrow). h Schematic illustration showing the time course for tamoxifen induction and mGFP expression in the PDGFRα-CreER T2 ; Tau-mGFP mice. i Immunostaining for NL3 (cyan) and mGFP (white) displaying the expression of NL3 in mature OL of white matter. Magnified images (lower) showing expression of NL3 in the cell body (red arrow), proximal processes (white arrow), and myelin sheaths (yellow arrow). j – l Immunostaining for NL3 (green) and PDGFRα (red) displaying the expression of NL3 in OPC of white matter in WT and KI mice ( j ). Quantification of NL3 + PDGFRα+ area ( k ) and PDGFRα+ area ( l ) in white matter at the age of P60 (WT/KI = 4/5 mice). The co-localization area of NL3 and PDGFRα in KI mice decreased, thereby decreasing NL3 protein expression in OPC. All data are shown as mean ± SEM; P -values were obtained by 2-tailed unpaired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: For conditional knockout (cKO) of NL3 in OPCs, Pdgfra-CreER T2 mice (predominantly C57BL/6) , were crossed with NL3 lox mice (C57BL/6, Cat#: S-CKO-08679, Cyagen Biosciences, Inc., Guangzhou, China), which had exon 6 of NL3 flanked by loxP sites.

Techniques: Derivative Assay, Quantitative RT-PCR, Expressing, Immunostaining, In Vitro, Staining

Journal: Nature Communications

Article Title: Hypomyelination in autism-associated neuroligin-3 mutant mice impairs parvalbumin interneuron excitability, gamma oscillations, and sensory discrimination

doi: 10.1038/s41467-025-61455-0

Figure Lengend Snippet: a , b Illustration of the NL3 -flox mouse line, showing that loxP sites flank exon 6. Cre-recombinase deletes exon 6 ( a ). Experimental schedule for OPC-specific and temporal control of NL3 deletion ( b ). c – e Immunostaining for NL3 (green) and PDGFRα (red) displaying the expression of NL3 in OPC ( c ). The co-localization area of NL3 and PDGFRα in NL3 cKO mice decreased, thereby effectively producing a conditional knockout ( NL3 cKO). Quantification of NL3 + PDGFRα + area ( d ) and PDGFRα + area ( e ) in white matter at the age of P60 ( n = 4 mice/group). f , g Immunostaining showing CC1 (red) co-localizing with SOX10 (green), displaying mature OLs in layer II/III BC ( f ). Quantification ( g ) of CC1 + OLs in layer II/III at the age of P60 (Control/ NL3 cKO = 5/4 mice). h , i Representative IHC images ( h ) displaying MBP (green) expression in BC layer II/III at P60. Quantification ( i ) of MBP length and density in BC layer II/III ( n = 4 mice/group). j , k IHC (PV, magenta) and quantification of PV + interneurons in layer II/III BC at the age of P60 ( n = 4 mice/group). l– n Detailed view of representative subregions of a single section (MBP/PV, green/red) in layer II/III BC of adult (P60) in Control mice and NL3 cKO mice ( l ). Quantitative analysis of total PV + myelin length ( m ) and PV + immunostaining area ( n ) in layer II/III BC. Control/ NL3 cKO = 5/4 mice. o Experimental timeline for OPC-specific and temporal control of NL3 deletion. The OPC- NL3 -cKO mice were generated by crossing NL3 Flox G mice (female) with Pdgfrα -creERT2 G mice (male). p Representative traces of action potential firing in response to a 450-pA current step from PV + interneurons in 60- to 70-day-old Control and NL3 cKO mice. q Histogram of firing frequency (spikes/s) revealed differences from 150 pA to 700 pA (15 neurons/group from 4 mice). P = 0.008117/0.01069/0.002376/0.002974/0.004534/0.01473/0.01289/0.02273, at 350–700 pA. r Patch-clamp recordings of mIPSCs in pyramidal neurons of layer II/III of BC. s , t Quantification of frequencies ( s ) and amplitudes ( t ) of mIPSC (15 neurons/group from 4 mice). All data are shown as mean ± SEM; P -values were obtained by 2-tailed unpaired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: For conditional knockout (cKO) of NL3 in OPCs, Pdgfra-CreER T2 mice (predominantly C57BL/6) , were crossed with NL3 lox mice (C57BL/6, Cat#: S-CKO-08679, Cyagen Biosciences, Inc., Guangzhou, China), which had exon 6 of NL3 flanked by loxP sites.

Techniques: Control, Immunostaining, Expressing, Knock-Out, Generated, Patch Clamp

Journal: Nature Communications

Article Title: Hypomyelination in autism-associated neuroligin-3 mutant mice impairs parvalbumin interneuron excitability, gamma oscillations, and sensory discrimination

doi: 10.1038/s41467-025-61455-0

Figure Lengend Snippet: a Experimental schedule. b , c Heat maps ( b ) and scatter plots ( c ) depicting time spent exploring identically textured objects during the learning phase and time spent exploring the novel (T2) and the familiar (T1) object for Control (left) and NL3 cKO (right), Control/ NL3 cKO = 15/12 mice. d Scatter plot showing exploration index for animals in ( c ). Control/ NL3 cKO = 15/12 mice. e – h IHC (CUX1/c-Fos, magenta/green) and quantification of CUX1 + neurons expressing c-Fos in the BC layer II/III after texture discrimination task (Control/ NL3 cKO = 5/4 mice). i Averaged relative BC layer II/III LFP power (1–90 Hz; Inset: 55–90 Hz) when test mice explored the sandpaper (Shaded areas indicate SEM). j Theta and gamma oscillations of the original signal and the corresponding phase and amplitude. k Statistics of LFP power at different frequency bands during the exploring period (Control/ NL3 cKO = 5/4 mice). l , m Standard comodulogram of PAC ( l ). Average PAC-MVL ( m ) in high-frequency bands coupled to theta bands (Control/ NL3 cKO = 5/4 mice). n , o The results from employing the OTC method to measure theta-high gamma phase-amplitude coupling during the texture discrimination task, summing 400 raw LFPs centered at events identified as the large gamma oscillations. n The degree of coupling strength; o preferred phase of coupling. Control/ NL3 cKO = 5/4 mice. All data are shown as mean ± SEM. Statistical analyses of results in ( d ), ( f ), ( g ), ( h ), ( k ), ( m ), ( n ) and ( o ) were evaluated with 2-tailed unpaired Student’s t -test. Statistical analyses of results in ( c ) were evaluated with 2-tailed paired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: For conditional knockout (cKO) of NL3 in OPCs, Pdgfra-CreER T2 mice (predominantly C57BL/6) , were crossed with NL3 lox mice (C57BL/6, Cat#: S-CKO-08679, Cyagen Biosciences, Inc., Guangzhou, China), which had exon 6 of NL3 flanked by loxP sites.

Techniques: Control, Expressing

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-301a-3p increases oxidative stress, inflammation and apoptosis in ox-LDL-induced HUVECs by targeting KLF7

doi: 10.3892/etm.2021.10001

Figure Lengend Snippet: miR-301a-3p is highly expressed in ox-LDL-induced HUVECs. (A) The viability of HUVECs induced by different concentrations of ox-LDL was detected by Cell Counting Kit-8 assay. * P<0.05 and *** P<0.001 vs. CTL group. (B) miR-301a-3p expression in HUVECs induced by different concentrations of ox-LDL was analyzed by RT-qPCR. ** P<0.01 and *** P<0.001 vs. CTL group. (C) miR-301a-3p expression in HUVECs transfected with miR-301a-3p inhibitor was analyzed by RT-qPCR. *** P<0.001 vs. control group; ### P<0.001 vs. inhibitor-NC group. (D) miR-301a-3p expression in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was analyzed by RT-qPCR. ** P<0.01 and *** P<0.001 vs. CTL group. ## P<0.01 vs. oxLDL group. ∆∆ P<0.01 vs. oxLDL + miR-NC group. miR, microRNA; HUVEC, human umbilical vein endothelial cells; oxLDL, oxidized low-density lipoprotein; CTL, control; NC, negative control; RT-qPCR, reverse transcription-quantitative PCR.

Article Snippet: HUVECs were respectively transfected with miRNA inhibitor-negative control (NC; 5 nM; cat. no. miR2N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p inhibitor (5 nM; cat. no. miR20000688-1-5; Guangzhou RiboBio Co., Ltd.), miRNA mimic-NC (5 nM; cat. no. miR1N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p mimic (5 nM; cat. no. miR10000688-1-5; Guangzhou RiboBio Co., Ltd.), small interfering RNA (si) R-NC (5 nM; cat. no. siN0000002-1-5; Guangzhou RiboBio Co., Ltd.) and siKLF7 (5 nM; cat. no. siG000008609A-1-5; Guangzhou RiboBio Co., Ltd.) using Lipofectamine ® 2000 transfection reagent (Invitrogen; Thermo Fisher Scientific, Inc.) for 48 h at 37 ̊C.

Techniques: Cell Counting, Expressing, Quantitative RT-PCR, Transfection, Control, Negative Control, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-301a-3p increases oxidative stress, inflammation and apoptosis in ox-LDL-induced HUVECs by targeting KLF7

doi: 10.3892/etm.2021.10001

Figure Lengend Snippet: Inhibition of miR-301a-3p inhibits inflammation and oxidative stress in ox-LDL-induced HUVECs. (A) The expression of IL-6 and MCP-1 in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was analyzed by western blot analysis. (B) The levels of IL-6 and MCP-1 in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was analyzed by ELISA. (C) ROS level in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was determined by a 2',7'-dichlorofluorescin diacetate assay (magnification, x200). (D) The SOD levels in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was determined by a SOD assay kit. * P<0.05, ** P<0.01 and *** P<0.001 vs. CTL group. ### P<0.001 vs. oxLDL group. ∆∆ P<0.01 and ∆∆∆ P<0.01 vs. oxLDL + miR-NC group. miR, microRNA; HUVEC, human umbilical vein endothelial cells; oxLDL, oxidized low-density lipoprotein; CTL, control; NC, negative control; MCP-1, monocyte chemoattractant protein-1; ROS, reactive oxygen species; SOD, superoxide dismutase.

Article Snippet: HUVECs were respectively transfected with miRNA inhibitor-negative control (NC; 5 nM; cat. no. miR2N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p inhibitor (5 nM; cat. no. miR20000688-1-5; Guangzhou RiboBio Co., Ltd.), miRNA mimic-NC (5 nM; cat. no. miR1N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p mimic (5 nM; cat. no. miR10000688-1-5; Guangzhou RiboBio Co., Ltd.), small interfering RNA (si) R-NC (5 nM; cat. no. siN0000002-1-5; Guangzhou RiboBio Co., Ltd.) and siKLF7 (5 nM; cat. no. siG000008609A-1-5; Guangzhou RiboBio Co., Ltd.) using Lipofectamine ® 2000 transfection reagent (Invitrogen; Thermo Fisher Scientific, Inc.) for 48 h at 37 ̊C.

Techniques: Inhibition, Expressing, Transfection, Western Blot, Enzyme-linked Immunosorbent Assay, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-301a-3p increases oxidative stress, inflammation and apoptosis in ox-LDL-induced HUVECs by targeting KLF7

doi: 10.3892/etm.2021.10001

Figure Lengend Snippet: Inhibition of miR-301a-3p inhibits apoptosis in ox-LDL-induced HUVECs. (A) LDH leakage in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was detected by an LDH assay kit. (B) The viability of HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was detected by Cell Counting Kit-8 assay. (C) The expression of apoptosis-related proteins in HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was determined by western blot analysis. (D and E) The apoptosis of HUVECs treated with ox-LDL and transfected with miR-301a-3p inhibitor was detected by flow cytometry analysis. ** P<0.01 and *** P<0.001 vs. CTL group. ### P<0.001 vs. oxLDL group. ∆∆∆ P<0.001 vs. oxLDL + miR-NC group. miR, microRNA; HUVEC, human umbilical vein endothelial cells; oxLDL, oxidized low-density lipoprotein; CTL, control; NC, negative control; LDH, lactate dehydrogenase; PARP, poly (ADP-ribose) polymerase.

Article Snippet: HUVECs were respectively transfected with miRNA inhibitor-negative control (NC; 5 nM; cat. no. miR2N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p inhibitor (5 nM; cat. no. miR20000688-1-5; Guangzhou RiboBio Co., Ltd.), miRNA mimic-NC (5 nM; cat. no. miR1N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p mimic (5 nM; cat. no. miR10000688-1-5; Guangzhou RiboBio Co., Ltd.), small interfering RNA (si) R-NC (5 nM; cat. no. siN0000002-1-5; Guangzhou RiboBio Co., Ltd.) and siKLF7 (5 nM; cat. no. siG000008609A-1-5; Guangzhou RiboBio Co., Ltd.) using Lipofectamine ® 2000 transfection reagent (Invitrogen; Thermo Fisher Scientific, Inc.) for 48 h at 37 ̊C.

Techniques: Inhibition, Transfection, Lactate Dehydrogenase Assay, Cell Counting, Expressing, Western Blot, Flow Cytometry, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-301a-3p increases oxidative stress, inflammation and apoptosis in ox-LDL-induced HUVECs by targeting KLF7

doi: 10.3892/etm.2021.10001

Figure Lengend Snippet: miR-301a-3p targets KLF7 in ox-LDL-induced HUVECs. (A) KLF7 mRNA expression in ox-LDL-induced HUVECs was analyzed by RT-qPCR. ** P<0.01 vs. CTL group. (B) KLF7 protein expression in ox-LDL-induced HUVECs was analyzed by western blot analysis. ** P<0.01 vs. CTL group. (C) miR-301a-3p expression in ox-LDL-induced HUVECs following miR-301a-3p mimic transfection was analyzed by RT-qPCR. *** P<0.001 vs. CTL group. ### P<0.001 vs. Mimic-NC group. (D/E) KLF7 expression in ox-LDL-induced HUVECs following miR-301a-3p mimic transfection was analyzed by western blot analysis. ** P<0.001 vs. Mimic-NC group. (F) Binding sites between KLF7 and miR-301a-3p. (G) Luciferase reporter assay was performed to confirm the direct binding relationship between KLF7 and miR-301a-3p. *** P<0.001 vs. Mimic-NC group. miR, microRNA; HUVEC, human umbilical vein endothelial cells; oxLDL, oxidized low-density lipoprotein; CTL, control; NC, negative control; KLF7, Krueppel-like factor 7; WT, wild-type; Mut, mutant; RT-qPCR, reverse transcription-quantitative PCR.

Article Snippet: HUVECs were respectively transfected with miRNA inhibitor-negative control (NC; 5 nM; cat. no. miR2N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p inhibitor (5 nM; cat. no. miR20000688-1-5; Guangzhou RiboBio Co., Ltd.), miRNA mimic-NC (5 nM; cat. no. miR1N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p mimic (5 nM; cat. no. miR10000688-1-5; Guangzhou RiboBio Co., Ltd.), small interfering RNA (si) R-NC (5 nM; cat. no. siN0000002-1-5; Guangzhou RiboBio Co., Ltd.) and siKLF7 (5 nM; cat. no. siG000008609A-1-5; Guangzhou RiboBio Co., Ltd.) using Lipofectamine ® 2000 transfection reagent (Invitrogen; Thermo Fisher Scientific, Inc.) for 48 h at 37 ̊C.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Transfection, Binding Assay, Luciferase, Reporter Assay, Control, Negative Control, Mutagenesis, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-301a-3p increases oxidative stress, inflammation and apoptosis in ox-LDL-induced HUVECs by targeting KLF7

doi: 10.3892/etm.2021.10001

Figure Lengend Snippet: Inhibition of KLF7 weakens the inhibitory effects of miR-301a-3p inhibition on inflammation and oxidative stress in ox-LDL-induced HUVECs. (A) KLF7 protein expression in ox-LDL-induced HUVEC following transfection of miR-301a-3p inhibitor transfection and siKLF7 was analyzed by western blot analysis. *** P<0.001 vs. Control group. ### P<0.001 vs. siR-NC group. (B) The expression of KLF7 in ox-LDL-induced HUVECs following transfection of siKLF7 was analyzed by western blot analysis. ** P<0.01 vs. siR-NC group. (C) The levels of IL-6 and MCP-1 in ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was analyzed by western blot analysis. (D) The levels of IL-6 and MCP-1 in ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was analyzed by ELISA. (E and F) The ROS level in ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was determined by a 2',7'-dichlorofluorescin diacetate assay. (G) The SOD levels in ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was determined by a SOD assay kit. * P<0.05, ** P<0.01 and *** P<0.001 vs. miR-NC group. # P<0.05 and ## P<0.01 vs. miR-301a-3p inhibitor group. ∆ P<0.05 and ∆∆ P<0.01 vs. miR-301a-3p inhibitor + siR-NC group. miR, microRNA; HUVEC, human umbilical vein endothelial cells; oxLDL, oxidized low-density lipoprotein; CTL, control; NC, negative control; KLF7, Krueppel-like factor 7; MCP-1, monocyte chemoattractant protein-1; ROS, reactive oxygen species; SOD, superoxide dismutase; si, small interfering.

Article Snippet: HUVECs were respectively transfected with miRNA inhibitor-negative control (NC; 5 nM; cat. no. miR2N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p inhibitor (5 nM; cat. no. miR20000688-1-5; Guangzhou RiboBio Co., Ltd.), miRNA mimic-NC (5 nM; cat. no. miR1N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p mimic (5 nM; cat. no. miR10000688-1-5; Guangzhou RiboBio Co., Ltd.), small interfering RNA (si) R-NC (5 nM; cat. no. siN0000002-1-5; Guangzhou RiboBio Co., Ltd.) and siKLF7 (5 nM; cat. no. siG000008609A-1-5; Guangzhou RiboBio Co., Ltd.) using Lipofectamine ® 2000 transfection reagent (Invitrogen; Thermo Fisher Scientific, Inc.) for 48 h at 37 ̊C.

Techniques: Inhibition, Expressing, Transfection, Western Blot, Control, Enzyme-linked Immunosorbent Assay, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-301a-3p increases oxidative stress, inflammation and apoptosis in ox-LDL-induced HUVECs by targeting KLF7

doi: 10.3892/etm.2021.10001

Figure Lengend Snippet: Inhibition of KLF7 weakens the inhibitory effects of miR-301a-3p inhibition on apoptosis in ox-LDL-induced HUVECs. (A) LDH leakage in ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was detected by an LDH assay kit. (B) The viability of ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was detected by Cell Counting Kit-8 assay. (C) The expression of apoptosis-related proteins in ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was determined by western blot analysis. (D and E) The apoptosis of ox-LDL-induced HUVECs following transfection of miR-301a-3p inhibitor and siKLF7 was detected by flow cytometry analysis. ** P<0.01 and *** P<0.001 vs. miR-NC group. ## P<0.01 and ### P<0.001 vs. miR-301a-3p inhibitor group. ∆∆ P<0.01 and ∆∆∆ P<0.001 vs. miR-301a-3p inhibitor + siR-NC group. miR, microRNA; HUVEC, human umbilical vein endothelial cells; oxLDL, oxidized low-density lipoprotein; CTL, control; NC, negative control; si, small interfering; LDH, lactate dehydrogenase; PARP, poly (ADP-ribose) polymerase.

Article Snippet: HUVECs were respectively transfected with miRNA inhibitor-negative control (NC; 5 nM; cat. no. miR2N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p inhibitor (5 nM; cat. no. miR20000688-1-5; Guangzhou RiboBio Co., Ltd.), miRNA mimic-NC (5 nM; cat. no. miR1N0000001-1-5; Guangzhou RiboBio Co., Ltd.), miR-301a-3p mimic (5 nM; cat. no. miR10000688-1-5; Guangzhou RiboBio Co., Ltd.), small interfering RNA (si) R-NC (5 nM; cat. no. siN0000002-1-5; Guangzhou RiboBio Co., Ltd.) and siKLF7 (5 nM; cat. no. siG000008609A-1-5; Guangzhou RiboBio Co., Ltd.) using Lipofectamine ® 2000 transfection reagent (Invitrogen; Thermo Fisher Scientific, Inc.) for 48 h at 37 ̊C.

Techniques: Inhibition, Transfection, Lactate Dehydrogenase Assay, Cell Counting, Expressing, Western Blot, Flow Cytometry, Control, Negative Control

Journal: Oncology Letters

Article Title: Identification of feature genes and molecular mechanisms involved in cell communication in uveal melanoma through analysis of single‑cell sequencing data

doi: 10.3892/ol.2024.14636

Figure Lengend Snippet: In vitro biofunction of PLXND1 and GNAI2 in C918 cells. Viability of C918 cells after (A) GNAI2 and (B) PLXND1 siRNA transfection. Gap closure rate of the (C) siGNAI2 and (D) siPLX1 groups after 20 h. (E) Representative images of the gap closure assay; scale bar, 200 µm. **P<0.01, ***P<0.001, ****P<0.0001. CON, control; GNAI2, G protein subunit α I2; PLXND1, plexin D1; si, small interfering (RNA).

Article Snippet: When the cells reached a 50–60% confluency, the SEC14L1, PLXND1, TPM4, GNAI2 or control siRNA (100 nmol/l) were transfected by riboFECT CP Transfection Kit (cat. no. C10511-05; Guangzhou RiboBio Co., Ltd.) according to the manufacturer's instructions.

Techniques: In Vitro, Transfection, Control, Small Interfering RNA