hk2 sirna  (Santa Cruz Biotechnology)

Journal: Genes & Diseases

Article Title: lncRNA ADAMTS9-AS2/let-7a-5p axis regulates metabolic reprogramming by targeting HK2 in oral submucous fibrosis-associated oral squamous cell carcinoma

doi: 10.1016/j.gendis.2025.101670

Figure Lengend Snippet: ADAMTS9-AS2 regulates aerobic glycolysis via HK2. (A) The heatmap showing aerobic glycolysis-related genes differentially expressed among normal oral mucous, OSF, and OSCC-OSF samples. (B) Protein levels of metabolic enzymes were determined by western blot in CAL27 and SCC9 cells expressing ADAMTS9-AS2 and empty vector. GAPDH was used as a loading control. (C) HK2 protein levels were determined by western blot in HOK cells with ADAMTS9-AS2 knockdown. (D) Correlation between ADAMTS9-AS2 and HK2 analyzed using starBase/ENCORI database. (E) HK2 protein expression level in HOK cells with ADAMTS9-AS2 and/or HK2 knockdown. GAPDH was used as a loading control. (F) Cellular glucose uptake and lactate production in the medium of HOK cells with ADAMTS9-AS2 and/or HK2 knockdown were determined. The data were presented as means ± standard deviation of three independent experiments by student’s t -test. ∗ P < 0.05. A9-AS2, ADAMTS9-AS2; OSCC, oral squamous cell carcinoma; OSF, oral submucous fibrosis.

Article Snippet: siRNA oligos targeting ADAMTS9-AS2 were purchased from GenePharma (Suzhou, China). siRNAs targeting ADAMTS9-AS2 are as follows: si-AS2-1-sense: 5′- CAGAGACGCAGGUAUUUAUTT-3′; si-AS2-1-antisense: 5′- AUAAAUACCUGCGUCUCUGTT-3′; si-AS2-2-sense: 5′- CGGCUUUCAAGAUUGGAAUTT-3′; si-AS2-2-antisense, 5′- AUUCCAAUCUUGAAAGCCGTT-3′. siRNAs targeting HK2 ( SR302109 ) were purchased from Origene (Origene, Rockville, Maryland, USA). siRNAs were transfected into CAL27 and SCC9 cell lines using Lipofectamine 3000 (Invitrogen, USA) according to the manufacturer’s instructions.

Techniques: Western Blot, Expressing, Plasmid Preparation, Control, Knockdown, Standard Deviation

Journal: Genes & Diseases

Article Title: lncRNA ADAMTS9-AS2/let-7a-5p axis regulates metabolic reprogramming by targeting HK2 in oral submucous fibrosis-associated oral squamous cell carcinoma

doi: 10.1016/j.gendis.2025.101670

Figure Lengend Snippet: Let-7a-5p inhibits HK2 expression by directly targeting its 3′-UTR in oral squamous cell carcinoma cells. (A, B) Immunoblot analysis of HK2 expression in CAL27 and SCC9 cells transfected with non-targeting control (NTC), (A) let-7a-5p mimics, or (B) let-7a-5p antagomirs. Histograms show relative let-7a-5p expression by quantitative reverse transcription PCR. (C) HK2 mRNA levels were measured by quantitative reverse transcription PCR in CAL27 and SCC9 cells expressing NTC or let-7a-5p mimics. (D) The top panel indicates wild-type (WT) and mutated (Mut) forms of putative let-7a-5p target sequences of HK2 3′-UTR. The red font indicates the putative let-7a-5p binding sites within human HK2 3′-UTR. The blue font indicates the mutations introduced into the HK2 3′-UTR. Let-7a-5p mimics were co-transfected with pmirGLO-HK2-3′-UTR or pmirGLO-HK2-3′-UTR-mut into CAL27 and SCC9 cells, followed by dual-luciferase analysis. (E) NTC or let-7a-5p antagomirs were transfected into CAL27 and SCC9 cells with HK2 knockdown, followed by measurement of cellular glucose uptake and lactate production in culture media. The data were presented as mean ± standard deviation of three independent experiments by student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01.

Article Snippet: siRNA oligos targeting ADAMTS9-AS2 were purchased from GenePharma (Suzhou, China). siRNAs targeting ADAMTS9-AS2 are as follows: si-AS2-1-sense: 5′- CAGAGACGCAGGUAUUUAUTT-3′; si-AS2-1-antisense: 5′- AUAAAUACCUGCGUCUCUGTT-3′; si-AS2-2-sense: 5′- CGGCUUUCAAGAUUGGAAUTT-3′; si-AS2-2-antisense, 5′- AUUCCAAUCUUGAAAGCCGTT-3′. siRNAs targeting HK2 ( SR302109 ) were purchased from Origene (Origene, Rockville, Maryland, USA). siRNAs were transfected into CAL27 and SCC9 cell lines using Lipofectamine 3000 (Invitrogen, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Western Blot, Transfection, Control, Reverse Transcription, Binding Assay, Luciferase, Knockdown, Standard Deviation

Journal: Genes & Diseases

Article Title: lncRNA ADAMTS9-AS2/let-7a-5p axis regulates metabolic reprogramming by targeting HK2 in oral submucous fibrosis-associated oral squamous cell carcinoma

doi: 10.1016/j.gendis.2025.101670

Figure Lengend Snippet: The ADAMTS9-AS2/let-7a-5p/HK2 axis regulates glycolysis in oral squamous cell carcinoma cells. (A) Representative immunoblot reveals the expression level of HK2 in ADAMTS9-AS2-expressing CAL27 and SCC9 cells transfected with HK2 expression or control plasmids. (B) Glucose uptake, lactate production, cellular ATP levels, and oxygen consumption were measured in ADAMTS9-AS2-expressing CAL27 and SCC9 cells transfected with HK2 expression plasmid, compared with controls. (C) Cell viability in CAL27 and SCC9 cells with both ADAMTS9-AS2 and HK2 expression was determined by CCK8 assay. The data were presented as mean ± standard deviation of three independent experiments by one-way ANOVA with multiple comparison post hoc analysis. ∗ P < 0.05, ∗∗ P < 0.01. A9-AS2, ADAMTS9-AS2.

Article Snippet: siRNA oligos targeting ADAMTS9-AS2 were purchased from GenePharma (Suzhou, China). siRNAs targeting ADAMTS9-AS2 are as follows: si-AS2-1-sense: 5′- CAGAGACGCAGGUAUUUAUTT-3′; si-AS2-1-antisense: 5′- AUAAAUACCUGCGUCUCUGTT-3′; si-AS2-2-sense: 5′- CGGCUUUCAAGAUUGGAAUTT-3′; si-AS2-2-antisense, 5′- AUUCCAAUCUUGAAAGCCGTT-3′. siRNAs targeting HK2 ( SR302109 ) were purchased from Origene (Origene, Rockville, Maryland, USA). siRNAs were transfected into CAL27 and SCC9 cell lines using Lipofectamine 3000 (Invitrogen, USA) according to the manufacturer’s instructions.

Techniques: Western Blot, Expressing, Transfection, Control, Plasmid Preparation, CCK-8 Assay, Standard Deviation, Comparison

Journal: Cancer Medicine

Article Title: SPARC Promotes Aerobic Glycolysis and 5‐Fluorouracil Resistance in Colorectal Cancer Through the STAT3 / HK2 Axis

doi: 10.1002/cam4.70972

Figure Lengend Snippet: SPARC promotes aerobic glycolysis via HK2 in CRC. (A) Glucose consumption was measured in OE‐SPARC RKO and HCT116 cells compared with OE‐NC cells (5‐FU: 20 umol/L). (B) Lactate production was measured in OE‐SPARC RKO and HCT116 cells compared with OE‐NC cells. (5‐FU: 20 umol/L) (C) Glucose consumption was measured in sh‐SPARC RKO and HCT116 cells compared with sh‐NC cells (5‐FU: 20 umol/L). (D) Lactate production was measured in sh‐SPARC RKO and HCT116 cells compared with sh‐NC cells (5‐FU: 20 umol/L). (E) RT‐qPCR assays were used to analyze the mRNA expression of key genes in aerobic glycolysis in OE‐SPARC RKO and HCT116 cells compared with OE‐NC cells (5‐FU: 20 umol/L). (F) The protein expression of SPARC and HK2 in sh‐SPARC RKO and HCT116 cells was compared with sh‐NC cells (5‐FU: 20 umol/L). (G) The protein expression of SPARC and HK2 in OE‐SPARC RKO and HCT116 cells after treatment with HK2 siRNA (5‐FU: 20 umol/L). (H) Glucose consumption was measured in OE‐SPARC RKO and HCT116 cells after treatment with HK2 siRNA (5‐FU: 20 umol/L). (I) Lactate production was measured in OE‐SPARC RKO and HCT116 cells after treatment with HK2 siRNA (5‐FU: 20 umol/L). Non‐significant results were denoted as “NS” while statistical significance was indicated as * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: A commercial HK2 siRNA (5′‐GCAGAAGGUUGACCAGUAUTT‐3′) was purchased from Shanghai GenePharma Co. Ltd. (Shanghai, China).

Techniques: Quantitative RT-PCR, Expressing

Journal: Cancer Medicine

Article Title: SPARC Promotes Aerobic Glycolysis and 5‐Fluorouracil Resistance in Colorectal Cancer Through the STAT3 / HK2 Axis

doi: 10.1002/cam4.70972

Figure Lengend Snippet: SPARC promotes resistance to 5‐FU chemotherapy as mediated by HK2 in CRC. (A) Colony formation assays were used to measure the proliferation ability of OE‐SPARC RKO and HCT116 cells after being treated with HK2 siRNA (5‐FU: 20 umol/L). (B) CCK8 assays were used to detect the cell viability of OE‐SPARC RKO and HCT116 cells after being treated with HK2 siRNA (5‐FU: 20 umol/L). (C) Flow cytometry was used to determine the apoptosis rate of OE‐SPARC RKO and HCT116 cells after being treated with HK2 siRNA (5‐FU: 20 umol/L). Non‐significant results were denoted as “NS” while statistical significance was indicated as * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: A commercial HK2 siRNA (5′‐GCAGAAGGUUGACCAGUAUTT‐3′) was purchased from Shanghai GenePharma Co. Ltd. (Shanghai, China).

Techniques: Flow Cytometry

Journal: Cancer Medicine

Article Title: SPARC Promotes Aerobic Glycolysis and 5‐Fluorouracil Resistance in Colorectal Cancer Through the STAT3 / HK2 Axis

doi: 10.1002/cam4.70972

Figure Lengend Snippet: SPARC promotes resistance to 5‐FU chemotherapy as mediated by the STAT3/HK2 axis in CRC. (A) The protein expression of P‐STAT3 and STAT3 in sh‐SPARC RKO and HCT116 cells compared with sh‐NC cells (5‐FU: 20 umol/L). (B) The protein expression of HK2, P‐STAT3, and STAT3 in OE‐SPARC RKO and HCT116 cells after being treated with HK2 siRNA (5‐FU: 20 umol/L, stattic: 2 μmol/L). (C) Colony formation assays were used to measure the proliferation ability of OE‐SPARC RKO and HCT116 cells after being treated with stattic (5‐FU: 20 umol/L, stattic: 2 μmol/L). (D) CCK8 assays were used to detect the cell viability of OE‐SPARC RKO and HCT116 cells after being treated with stattic (5‐FU: 20 umol/L, stattic: 2 μmol/L). (E) Flow cytometry was used to determine the apoptosis rate of OE‐SPARC RKO and HCT116 cells after treated with stattic (5‐FU: 20 umol/L, stattic: 2 μmol/L). Non‐significant results were denoted as “NS” while statistical significance was indicated as * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: A commercial HK2 siRNA (5′‐GCAGAAGGUUGACCAGUAUTT‐3′) was purchased from Shanghai GenePharma Co. Ltd. (Shanghai, China).

Techniques: Expressing, Flow Cytometry

Journal: Cancer Medicine

Article Title: SPARC Promotes Aerobic Glycolysis and 5‐Fluorouracil Resistance in Colorectal Cancer Through the STAT3 / HK2 Axis

doi: 10.1002/cam4.70972

Figure Lengend Snippet: The SPARC/STAT3/HK2 axis promoted resistance to 5‐FU chemotherapy of CRC cells in vivo. (A) Representative images of subcutaneous tumour tissue were acquired from the OE‐NC, OE‐SPARC, and 2‐DG mouse models (5‐FU:23 mg/kg, 2‐DG: 50 mg/kg). (B) The volume and growth rate of subcutaneous tumours in the OE‐NC, OE‐SPARC, and 2‐DG groups were calculated. (C) The expression of SPARC, STAT3, P‐STAT3, and HK2 in subcutaneous tumour tissues in the OE‐NC, OE‐SPARC, and 2‐DG groups were determined by IHC experiments. Scale bar, 50 μm. Non‐significant results were denoted as “NS” while statistical significance was indicated as ** p < 0.01 and *** p < 0.001.

Article Snippet: A commercial HK2 siRNA (5′‐GCAGAAGGUUGACCAGUAUTT‐3′) was purchased from Shanghai GenePharma Co. Ltd. (Shanghai, China).

Techniques: In Vivo, Expressing

Journal: Cancer Medicine

Article Title: SPARC Promotes Aerobic Glycolysis and 5‐Fluorouracil Resistance in Colorectal Cancer Through the STAT3 / HK2 Axis

doi: 10.1002/cam4.70972

Figure Lengend Snippet: Schematic representation of the mechanism by which SPARC promotes aerobic glycolysis and 5‐fluorouracil resistance in colorectal cancer through the STAT3/HK2 axis.

Article Snippet: A commercial HK2 siRNA (5′‐GCAGAAGGUUGACCAGUAUTT‐3′) was purchased from Shanghai GenePharma Co. Ltd. (Shanghai, China).

Techniques:

Journal: Theranostics

Article Title: Glucose homeostasis controls N-acetyltransferase 10-mediated ac4C modification of HK2 to drive gastric tumorigenesis

doi: 10.7150/thno.104310

Figure Lengend Snippet: NAT10-mediated ac4C modification of HK2 mRNA maintains its stability. (A) The distribution of ac4C-containing peaks across mRNAs in NAT10-overexpressing (left panel) and NAT10 knockout (right panel) GC cells is displayed in a metagene plot. (B) Representative pie chart of the peak distribution showing the proportions of total ac4C peaks in the indicated regions, including the 5′-untranslated region (5′-UTR), coding sequence (CDS), 3′- untranslated region (3′-UTR), start codon, and stop codon. (C) Consensus motif in AGS WT and NAT10-knockout cells identified by HOMER. (D) Venn diagram showing HK2 as the selected candidate target gene of NAT10. (E) The mRNA level of HK2 in NAT10-overexpressing and NAT10-KO GC cells was measured by qRT-PCR. (F) The protein level of HK2 in NAT10-overexpressing and NAT10-KO GC cells was measured by Western blotting. (G) Sections of xenograft tumors derived from NAT10-overexpressing (upper panel) and NAT10-knockdown (bottom panel) GC cells subcutaneously injected into nude mice were stained with an anti-HK2 antibody for IHC analysis (scale bars = 50 μm). (H) Sections of organoids formed from cells transduced with the NAT10 overexpression or control lentiviral vector were subjected to H&E staining or IHC staining with an anti-HK2 antibody (scale bars = 50 μm). (I) Sections of stomachs from NAT10 WT and cKO mice were stained with an anti-HK2 antibody for IHC analysis (scale bars = 100 μm). (J) Integrative Genomics Viewer (IGV) tracks revealing the ac4Cseq read distribution on HK2 mRNA in NAT10-overexpressing and NAT10-knockout GC cells. (K) acRIP-qPCR analysis was employed to demonstrate the presence of NAT10-mediated HK2 ac4C modifications. ac4C modification of HK2 was increased upon overexpression of NAT10, while it was decreased upon knockout of NAT10. (L) The level of HK2 expression in NAT10-overexpressing, NAT10-knockout and the corresponding control GC cells treated with actinomycin D (2 μg/mL) at the indicated time points was measured by qRT-PCR. (M) Schematic representation of the construction of the luciferase reporter vectors containing the HK2 WT, Mut1, Mut2, or Both Mut sequences (left panel). The relative luciferase activities of the HK2 WT, Mut1, Mut2, and Both Mut luciferase reporters in AGS cells with NAT10 knockout and the corresponding control cells were measured (right panel). H&E, haematoxylin and eosin; IHC, immunohistochemical; GC, gastric cancer; WT, wild type; KO, knockout. The statistical data in this figure are presented as the mean values ± SDs of three independent experiments. Statistical significance was determined by a two-tailed t test. * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: siRNAs targeting NAT10 or HK2 were designed and synthesized by RiboBio (Guangzhou, China).

Techniques: Modification, Knock-Out, Sequencing, Quantitative RT-PCR, Western Blot, Derivative Assay, Knockdown, Injection, Staining, Transduction, Over Expression, Control, Plasmid Preparation, Immunohistochemistry, Expressing, Luciferase, Immunohistochemical staining, Two Tailed Test

Journal: Theranostics

Article Title: Glucose homeostasis controls N-acetyltransferase 10-mediated ac4C modification of HK2 to drive gastric tumorigenesis

doi: 10.7150/thno.104310

Figure Lengend Snippet: NAT10 accelerates malignant progression of GC by upregulating HK2. (A) The HK2 protein level was measured in GC tissues and paired normal gastric mucosal tissues by Western blotting (n = 5). (B) Representative IHC images of the tissue microarray analysed with the anti-HK2 antibody (scale bars = 200 or 100 μm) are shown (left panel). The distribution of the difference in the HK2 immunoreactivity score (IRS) (△IRS = IRST - IRSN) is shown. The IRS for HK2 staining was available for 192 pairs of tissues. (C) Representative images of HK2 IHC staining in tissues from GC patients in the SUVmax-high and SUVmax-low groups (left panel). Scale bars = 50 μm; n = 22; statistical analysis of the difference in HK2 expression between the SUVmax-high group and the SUVmax-low group (right panel). (D) Correlation analysis between the SUVmax and HK2 protein expression level based on the IHC score in GC tissue microarrays with associated PET/CT data. R, Pearson correlation coefficient; n = 22. (E) Kaplan-Meier analysis of OS in GC patients stratified by HK2 expression (n = 192, P = 6.4e-15; log-rank test). (F) Multivariate analyses were performed for the GC cohort. (G) Glucose uptake (left panel) and lactate production (right panel) in HK2-knockdown AGS cells were measured. (H) A colony formation assay was performed in stable HK2-knockdown AGS cells (left panel). Quantitative analysis of the colony formation assay results (right panel). (I) NAT10 expression was positively correlated with HK2 protein expression in GC tissues (linear regression) according to analysis of the IHC score from the TMA data (n = 192). (J) Correlation analysis of NAT10 and HK2 protein expression based on the IHC score in GC tissue microarrays with associated PET/CT data (n = 22). (K) The colony formation ability was evaluated in NAT10-overexpressing BGC823 cells with or without stable HK2 knockdown or the corresponding controls. Representative images (left panel) and quantitative results (right panel) are shown. (L) Lactate production was measured in NAT10-overexpressing BGC823 cells with or without stable HK2 knockdown or the corresponding controls. CI, confidence interval; GC, gastric cancer; HR, hazard ratio; IHC, immunohistochemical; OS, overall survival; TNM, tumor, node, metastasis. The statistical data in this figure are presented as the mean value ± SD of three independent experiments. Statistical significance was determined by a two-tailed t test. * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: siRNAs targeting NAT10 or HK2 were designed and synthesized by RiboBio (Guangzhou, China).

Techniques: Western Blot, Microarray, Staining, Immunohistochemistry, Expressing, Positron Emission Tomography-Computed Tomography, Knockdown, Colony Assay, Immunohistochemical staining, Two Tailed Test

Journal: Theranostics

Article Title: Glucose homeostasis controls N-acetyltransferase 10-mediated ac4C modification of HK2 to drive gastric tumorigenesis

doi: 10.7150/thno.104310

Figure Lengend Snippet: Targeting the NAT10-HK2 axis in GC cells has clinical value. (A) Different GC cell lines were treated with 10 μM remodelin for 24 h. Then, the protein level of NAT10 was measured by Western blot analysis (upper panel). mRNA isolated from GC cells were subjected to dot blot analysis with an anti-ac4C antibody (bottom panel). MB staining served as a loading control. (B) Glucose uptake (left panel) and lactate production (right panel) were measured in MKN45 cells after treatment with 10 μM Remodelin for 24 h. (C) Remodelin inhibited subcutaneous tumor growth in nude mice (n = 6, left panel). The tumor volume was monitored every other day, and tumor growth curves were generated (middle panel). The tumors were extracted and weighed after 25 days (right panel). (D) Sections of tumors were stained with anti-Ki-67, anti-HK2, and anti-NAT10 antibodies for IHC analysis (scale bars = 50 μm). (E) Representative images of GC organoids treated with 10 μM Remodelin for the indicated durations (scale bars = 100 μm, left panel) and quantification of organoid diameters (right panel). (F) Sections of organoids treated with Remodelin were subjected to H&E staining or stained with anti-Ki-67, anti-HK2, and anti-NAT10 antibodies for IHC analysis (scale bars = 50 μm). (G) The combination of NAT10 and HK2 was evaluated as a new two-gene risk signature, and multivariate analyses were performed for the GC cohort. (H) GC patients were divided into three subgroups according to the median expression level of each protein: high expression of both NAT10 and HK2, low expression of both NAT10 and HK2, and other expression patterns (NAT10 high and HK2 low OR NAT10 low and HK2 high). Kaplan-Meier analysis of survival in the three subgroups of GC patients. (I) Time-dependent receiver operating characteristic (ROC) curve analysis of the NAT10 risk score, the HK2 risk score, and the combined NAT10/HK2 score in the GC cohort. AUC, area under the curve; CI, confidence interval; GC, gastric cancer; HR, hazard ratio; IHC, immunohistochemical; OS, overall survival; TNM, tumor, node, metastasis. (J) Graphical illustration of the mechanism by which NAT10 modulates glycolysis, promoting GC growth. The statistical data in this figure are presented as the mean value ± SD of three independent experiments. Statistical significance was determined by a two-tailed t test. * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: siRNAs targeting NAT10 or HK2 were designed and synthesized by RiboBio (Guangzhou, China).

Techniques: Western Blot, Isolation, Dot Blot, Staining, Control, Generated, Expressing, Immunohistochemical staining, Two Tailed Test

Journal: Journal of Cellular and Molecular Medicine

Article Title: Upregulation of circ‐ IGF1R increased therapeutic effect of hypoxia‐pretreated ADSC ‐derived extracellular vesicle by regulating miR ‐503‐5p/ HK2 / VEGFA axis

doi: 10.1111/jcmm.18471

Figure Lengend Snippet: Both HK2 and VEGFA were the downstream target of miR‐503‐5p. (A and B) Bioinformatics analysis and luciferase report analysis show the interactive relationship between miR‐503‐5p and 3'‐UTR‐HK2. *** p < 0.001. (C and D) Bioinformatics analysis and luciferase report analysis show the interactive relationship between miR‐503‐5p and 3'‐UTR‐VEGFA. *** p < 0.001. (E and F) Representative figures of wound‐healing procedures at various time points in various groups. The wound healing rate quantification. *** p < 0.001 versus HEV. ### p < 0.001 versus si‐circ‐IGF1R‐HEV. (G and H) Immunohistochemicals for Tunel staining show the apoptosis in skin tissue from the ulcer. *** p < 0.001 versus HEV. ### p < 0.001 versus si‐circ‐IGF1R‐HEV. (I and J) Immunohistochemicals for CD31 staining show the angiogenesis in skin tissue from the ulcer. ** p < 0.01, *** p < 0.001 versus HEV. ### p < 0.001 versus si‐circ‐IGF1R‐HEV.

Article Snippet: Our team synthesized miR‐503‐5p mimics and HK2/VEGFA siRNA via Genepharma (Suzhou, China).

Techniques: Luciferase, TUNEL Assay, Staining

Journal: Journal of Cellular and Molecular Medicine

Article Title: Upregulation of circ‐ IGF1R increased therapeutic effect of hypoxia‐pretreated ADSC ‐derived extracellular vesicle by regulating miR ‐503‐5p/ HK2 / VEGFA axis

doi: 10.1111/jcmm.18471

Figure Lengend Snippet: Overexpression miR‐503‐5p or silence HK2/VEGFA reversed the protective effect of circ‐IGF1R on vascular endothelial cell function under hyperglycemic microenvironment. (A–D) RT‐qPCR detection shows the expression of circ‐IGF1R, miR‐503‐5p, HK2 and VEGFA. ** p < 0.01, *** p < 0.001 versus NC. ### p < 0.001 versus circ‐IGF1R. (E and G) MLMECs apoptosis was assayed by flow cytometry after Annexin V‐FITC staining. *** p < 0.001 versus NC. ### p < 0.001 versus circ‐IGF1R. (F and H) In vitro tube formation of MLMECs. The total branching was analysed. ** p < 0.01, *** p < 0.001 versus NC. ### p < 0.001 versus circ‐IGF1R. mimic, miR‐503‐5p mimic. NC, negative control.

Article Snippet: Our team synthesized miR‐503‐5p mimics and HK2/VEGFA siRNA via Genepharma (Suzhou, China).

Techniques: Over Expression, Cell Function Assay, Quantitative RT-PCR, Expressing, Flow Cytometry, Staining, In Vitro, Negative Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: Upregulation of circ‐ IGF1R increased therapeutic effect of hypoxia‐pretreated ADSC ‐derived extracellular vesicle by regulating miR ‐503‐5p/ HK2 / VEGFA axis

doi: 10.1111/jcmm.18471

Figure Lengend Snippet: Circ‐IGF1R upregulation increased therapeutic effect of ADSC‐HEV on wound healing in diabetic mice. (A, B) Representative figures of wound‐healing procedures at various time points in various groups. The wound‐healing rate quantification. *** p < 0.001 versus NC. # p < 0.05 versus HEV. (C and D) Immunohistochemicals for Tunel staining show the apoptosis in skin tissue from the ulcer. ** p < 0.01, *** p < 0.001 versus NC. ## p < 0.01 versus HExo. (E and F) Immunohistochemicals for CD31 staining show the angiogenesis in skin tissue from the ulcer. *** p < 0.001 versus NC. # p < 0.05 versus HEV. (G–I) RT‐qPCR detection shows the expression of miR‐503‐5p, HK2 and VEGFA. ** p < 0.01, *** p < 0.001 versus NC. # p < 0.05, ## p < 0.01, ### p < 0.001 versus HEV. ADSC‐HEV, hypoxia‐pretreated adipose‐derived stem cells‐extracellular vesicle; NC, negative control.

Article Snippet: Our team synthesized miR‐503‐5p mimics and HK2/VEGFA siRNA via Genepharma (Suzhou, China).

Techniques: TUNEL Assay, Staining, Quantitative RT-PCR, Expressing, Derivative Assay, Negative Control

Journal: Stem Cell Research & Therapy

Article Title: MSCs promote the efferocytosis of large peritoneal macrophages to eliminate ferroptotic monocytes/macrophages in the injured endometria

doi: 10.1186/s13287-024-03742-z

Figure Lengend Snippet: MSCs promote the efferocytosis of LPMs by secreting STC-1 in vitro and in vivo . ( A , B ) The mRNA ( A ) and protein ( B ) levels of STC-1 in MSCs cocultured with LPMs stimulated with LPS for 24 h were determined by qRT-PCR and ELISA (normalised to GAPDH). ( C , D ) LPMs were incubated with nontreated or Erastin-treated THP-1 cells in the presence of rhSTC-1 protein (100 ng/mL) for 4 h. Efferocytosis of LPMs from each group was determined by ( C ) flow cytometry and ( D ) confocal microscope (scale bar: 40 μm). ( E , F ) qRT-PCR analysis ( E ) and ELISA ( F ) showed the mRNA and protein levels of STC-1 in MSCs transfected with three interfering fragments of STC-1 (si-STC-1) for 24 h (normalised to GAPDH). ( G , H ) MSCs were transfected with siSTC-1 for 24 h before cocultured with LPMs, then LPMs were incubated with Erastin-treated THP-1 cells stained by CFSE for 4 h. Efferocytosis of LPMs from each group was determined by ( G ) flow cytometry and ( H ) confocal microscope (scale bar: 40 μm). ( I ) Schematic of in vivo experimental model design ( n = 5–7). ( J ) Flow cytometry analysis for LPMs isolated from peritoneal cavity harvested after 7-days injury. Cells were pregated on CD11b + . ( K ) Flow cytometry analysis for LPMs isolated from uteri harvested after 7-days injury. ( L ) Flow cytometry analysis for GPX4 expression of CD11b + F4/80 med subsets in the uteri harvested after 7-days injury. ( M ) HE staining, Masson’s trichrome staining, and MDA staining of endometrial tissues obtained from mice of four groups. Scale bar indicates 100 μm. ( N ) The mRNA expression levels of TNF-α, IL-1β, and IL-6 in the endometria of mice were determined by qRT-PCR (normalised to β-actin). ( O ) Serum concentrations of TNF-α, IL-1β, and IL-6 were measured by ELISA. Values are mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, ns denotes p > 0.05 (by unpaired Student’s t test)

Article Snippet: SiRNAs targeting HK2 and STC-1 were purchased from RiboBio (Guangzhou, China).

Techniques: In Vitro, In Vivo, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Incubation, Flow Cytometry, Microscopy, Transfection, Staining, Isolation, Expressing