recombinant globular adiponectin  (PeproTech)

Journal: International Journal of Obesity (2005)

Article Title: Identification of circulating apolipoprotein M as a new determinant of insulin sensitivity and relationship with adiponectin

doi: 10.1038/s41366-024-01510-w

Figure Lengend Snippet: Spearman correlation coefficients of adiponectin, apoM and HOMA-IR with anthropometric parameters, biological markers and environmental factors in cohort A.

Article Snippet: After 10 days of differentiation, the hMADS cells were treated with recombinant globular adiponectin (PeproTech), or CRP (Merck Millipore) or exposed to 50/50 conditioned media from M1-like or M2-like polarized ThP-1.

Techniques: Activity Assay

Journal: International Journal of Obesity (2005)

Article Title: Identification of circulating apolipoprotein M as a new determinant of insulin sensitivity and relationship with adiponectin

doi: 10.1038/s41366-024-01510-w

Figure Lengend Snippet: Sera from 11 individuals with obesity (cohort B) were tested for apoM (panel A ) and adiponectin (panel B ) levels before and one year after sleeve gastrectomy. Changes in adipokine concentrations were correlated to the changes in the Homeostasis Model Assessment of Insulin Resistance (HOMA-IR) index. Data are expressed as percentage change from baseline. Linear correlation coefficients and P- values are displayed in each graph. The dotted lines represent 95% confidence intervals.

Article Snippet: After 10 days of differentiation, the hMADS cells were treated with recombinant globular adiponectin (PeproTech), or CRP (Merck Millipore) or exposed to 50/50 conditioned media from M1-like or M2-like polarized ThP-1.

Techniques:

Journal: International Journal of Obesity (2005)

Article Title: Identification of circulating apolipoprotein M as a new determinant of insulin sensitivity and relationship with adiponectin

doi: 10.1038/s41366-024-01510-w

Figure Lengend Snippet: APOM mRNA level upon 48 h of globular adiponectin treatment in hMADS adipocytes (panel A ) and in HepG2 hepatocytes (panel B ). Results are expressed as percentage of control and are presented as mean ± SD of 3 independent experiments or 1 experiment, respectively. Data were analyzed by Kruskal–Wallis’ test. * P < 0.05, ** P < 0.01.

Article Snippet: After 10 days of differentiation, the hMADS cells were treated with recombinant globular adiponectin (PeproTech), or CRP (Merck Millipore) or exposed to 50/50 conditioned media from M1-like or M2-like polarized ThP-1.

Techniques: Control

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Effects of globular adiponectin on the immunomodulatory function of ADSC. (A) ADSC were seeded in a 24-well plate at different densities (0.125 × 10 5 to 2 × 10 5 cells/well) and pretreated with globular adiponectin (gAcrp) for 6 h, followed by coculturing with CFSE-labeled splenocytes (10 6 cells/well). Proliferation of lymphocytes was induced by supplementation of 1 μg/mL of PHA-l into coculture media. After 4 days of coculture, non-adherent cells were stained with an APC-conjugated anti-CD3 antibody and subjected to FACS analysis. Live and CD3-positive cell populations were selected for CFSE dilution analysis. (B) ADSC were preconditioned with different concentrations of gAcrp before coculturing with splenocytes at ADSC/splenocytes of 1:20 and 1:40. Proliferation of splenocytes was measured as described above. (C) ADSC were treated with 1 μg/mL of gAcrp for different time durations. At the end of treatment period, splenocytes were added into culture wells containing ADSC with a fixed ADSC/splenocytes ratio of 1:20. Lymphocyte proliferation was determined by CFSE dilution assay. (D) ADSC were incubated with 1 μg/mL of gAcrp for 6 h, followed by coculture with CFSE-stained mouse CD3 + pan T cells (ADSC/splenocytes ratio of 1:20). T cells were activated with anti-CD3/CD28 beads in presence of IL-2 (10 ng/mL). The growth of T cells was examined after 4 days of coculture. (E) Mouse ADSC were stimulated with gAcrp (1 μg/mL, 6 h), followed by coculturing with non-labeled splenocytes for 4 days. Subsequently, splenocytes were subjected to immunofluorescent staining for anti-CD4 (FITC), CD25 (PE) and Foxp3 (APC). Dead cells were excluded by staining with a fixable viability dye. The proportion of CD25/Foxp3 double positive cell population among live and CD4-positive cells was determined. (F) C57BL/6 mice were fed with high-fat diet (HFD) for 12 weeks. After collection of blood samples from obese and lean mice, serum adiponectin was measured by ELISA. (G, H) ADSC derived from lean and obese mice were pretreated with gAcrp (1 μg/mL, for 6 h) and cocultured with CFSE-labeled splenocytes. PHA-stimulated lymphocyte proliferation was analyzed using CFSE dilution assay. ∗ P < 0.05 compared to indicated groups; n = 3 or n = 13 for Fig. 1 F.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Labeling, Staining, Dilution Assay, Incubation, Enzyme-linked Immunosorbent Assay, Derivative Assay

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Therapeutic effectiveness of lean and obese ADSC modified with globular adiponectin against DSS-induced colitis. (A) Experiment scheme for the in vivo study. Colitis was induced in C57BL/6 mice by administration of 3% DSS in drinking water for 7 days. Colitis mice received one of the following treatments: PBS (DSS control), lean ADSC (L-DSC), gAcrp-modified lean ADSC (gAcrp-L-ADSC), obese ADSC (Ob-ADSC), and gAcrp-modified obese ADSC (gAcrp-Ob-ADSC). All treatments were administered two times on Days 1 and 3 (3 × 10 6 cells each dose) via intraperitoneal injection. On Day 9, mice were sacrificed, and colon tissues were collected for further investigations. (B) Body weight was monitored throughout the study and changes in body weight compared to Day 0 were presented. (C) Disease activity index was calculated based on the status of stool, bleeding, and body weight loss, as indicated in Methods. (D) Representative images of colon from each group were presented (left panel). Colon lengths in each group were indicated in the bar graph (right panel). (E) MPO activity in colon tissues was determined. (F) Colon tissues were subjected to H&E staining. Representative images for each group were presented (left panel) along with histological injury scores determined as described in Methods (right panel). (G) Single cells were isolated from spleens and mesenteric lymph nodes (MLN) of all mice at the end of the study. Cells were then stained with anti-CD4 (FITC), anti-CD25 (PE), and anti-Foxp3 (APC) antibody. Representative images for CD25 + Foxp3 hi population among CD4 + cells from each group were shown (left panel) along with the proportion of Treg (right panel). (H, I) Colon tissues were digested with collagenase to obtain single cells. Cells were stained either with anti-CD3 (APC), anti-CD4 (PE), and anti-CD8 (Percp-Cy5.5) antibody cocktail (H), or with anti-MHC-II (Percp-Cy5.5), anti-CD11b (APC-Cy7), anti-CD11c (APC), and anti-F4/80 (PE) antibody cocktail (I). The percentage of CD4 + T cells (CD3 + CD4 + population), CD8 + T cells (CD3 + CD8 + population), macrophages (CD11b + F4/80 + population), and dendritic cells (MHC-II + CD11c + population) among colon-derived single cells was determined using FACS analysis. (J) The messenger RNA levels of indicated genes were measured in colon tissues by RT-qPCR and demonstrated in a figure representing fold changes. The quantification data for each gene are presented in Fig. S6 . ∗ P < 0.05 compared to indicated groups; n = 6.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Modification, In Vivo, Control, Injection, Activity Assay, Staining, Isolation, Derivative Assay, Quantitative RT-PCR

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Involvement of both soluble factors and cell-to-cell contact in modulating immunosuppressive potency of ADSC by globular adiponectin. (A) ADSC were pretreated with gAcrp for 6 h, followed by coculturing with splenocytes (ADSC/splenocytes of 1:20) directly or through a transwell insert to prevent cell-to-cell contact. PHA-stimulated lymphocyte proliferation was examined by CFSE dilution assay as described above. (B) ADSC were treated with 1 μg/mL of gAcrp for indicated periods. The messenger RNA levels of immunomodulation-related genes were measured by RT-qPCR and demonstrated in a figure representing the fold changes. Quantification data for each gene are presented in Fig. S7 . (C–E) ADSC were pretreated with gAcrp for 6 h, followed by further stimulation with TNF α (10 ng/mL) and IFN γ (20 ng/mL) for 24 h (C) or 48 h (D, E). (C) Expression levels of Nos2 , Cox2 , Ptges , Icam1 , and Cd274 were determined using RT-qPCR. (D, E) Nitrite/nitrate (D) and prostaglandin E2 (PGE2) (E) levels in culture media were measured as described in the Methods. (F, G) ADSC were preconditioned with gAcrp for 6 h, followed by coculturing with PHA-activated splenocytes. The concentrations of nitrate/nitrite (F) and PGE2 (G) in coculture media were examined after 3 days of coculture. (H, I) ADSC were pretreated with gAcrp for 6 h and further stimulated with TNF α /IFN γ for 24 h. Membrane ICAM-1 (H) and PD-L1 (I) were determined by labelling with respective fluorophore-conjugated antibodies, followed by flow cytometry analysis. (J) ADSC were treated with gAcrp for 6 h, followed by coculturing with PHA-activated splenocytes. NS-398 (10 μmol/L), a COX-2 inhibitor, and l -NAME, an inhibitor of iNOS, were supplemented to coculture media. Lymphocyte proliferation rate was measured at Day 4 of coculture. (K) ADSC were transfected with siRNA targeting Icam1 or Cd274 . After 24 h, cells were treated with gAcrp for further 6 h and cocultured with PHA-activated splenocytes for 4 days. Lymphocyte proliferation rate was finally monitored as described in Methods. (L) The mRNA levels of Nos2 , Cox2 , Ptges , Icam1 , and Cd274 were determined in ADSC derived from lean and obese mice with or without TNF α /IFN γ stimulation. (M) Obese ADSC were treated with gAcrp (1 μg/mL) for 6 h and the expression of indicated genes was measured by RT-qPCR. The knockdown efficiency of ICAM-1 and PD-L1 was examined by flow cytometry analysis ( Fig. S9 ). ∗ P < 0.05 compared to indicated groups; n = 3.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Dilution Assay, Quantitative RT-PCR, Expressing, Membrane, Flow Cytometry, Transfection, Derivative Assay, Knockdown

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Pro-survival effects of globular adiponectin on ADSC under inflammatory microenvironment. (A) After treatment with gAcrp for different time periods, viability of ADSC was examined by MTS assay. (B) ADSC were treated with gAcrp for 48 h. The proportion of apoptotic cells was determined by Annexin V binding assay. (C, D) ADSC were treated with gAcrp for different time periods. (C) Expression levels of BCL2, BAX, and cleaved caspase-3 (c-CASP3) were measured by Western blot analysis. (D) The caspase-3 activity was measured as described in Methods. In experiments A–D, cell death was induced by culturing in low-serum (0.5% FBS) containing media. (E) ADSC derived from obese mice were treated with gAcrp for 48 h. Proportion of apoptotic cells was determined by Annexin V binding assay. (F) ADSC were treated with gAcrp for 6 h, followed by coculturing with anti-CD3/CD28-activated splenocytes. After 3 days of coculture, adherent cells were collected by trypsinization, and then sequentially labeled with anti-CD45 (APC) antibody and FITC-Annexin V/7-AAD. Proportion of apoptotic cell death in CD45-negative cells was determined by flow cytometry analysis. (G, H) Colitis was induced in C57BL/6 by supplementation of 3% DSS in drinking water. After 3 days of colitis induction, mice were intraperitoneally injected with 3 × 10 6 DiR-labeled ADSC from lean and obese mice with or without gAcrp preconditioning. PBS was used as negative control. On Day 5, colon tissues were collected for fluorescent imaging analysis, followed by preparation of single cells for flow cytometry analysis. (G) Representative images of colon tissues were shown along with the quantitation of relative DiR fluorescent intensity in colon tissues from all mice. (H) Single cells were subjected to FACS analysis for determining the proportion of DiR-positive live cells. ∗ P < 0.05 compared to indicated groups; n = 3 (A–F) or 4 (G, H).

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: MTS Assay, Binding Assay, Expressing, Western Blot, Activity Assay, Derivative Assay, Labeling, Flow Cytometry, Injection, Negative Control, Imaging, Quantitation Assay

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Critical roles of HIF1 α signaling in the enhanced cell viability and immunosuppressive function of ADSC by globular adiponectin. (A, B) ADSC were treated with gAcrp for indicated time periods (A) or pretreated with gAcrp for 6 h and then stimulated with TNF α /IFN γ for additional 24 h (B). HIF1 α expression was measured by Western blot analysis. (C) ADSC were treated with gAcrp alone or in combination with TNF α /IFN γ for 24 h. Cells were immunolabeled with an anti-HIF1 α primary antibody and an Alexa fluor 488-conjugated secondary antibody (green), followed by counterstaining with DAPI (red). Subcellular distribution of HIF1 α was analyzed using a confocal microscope. Representative images from three independent experiments were presented in upper panel. The nuclear integrated intensity of HIF1 α was analyzed using Image J software and presented in the bar diagram (lower panel). Scale bar: 20 μm. (D, E) Expression levels of HIF1 α in lean and obese ADSC in the absence (D) or presence (E) of gAcrp were determined by Western blot analysis. (F, G) (F) ADSC were transfected with siRNA targeting Hif1a (20 nmol/L) for 24 h, followed by treatment with gAcrp for additional 6 h. Gene silencing efficiency was monitored by Western blot analysis (upper panel). (G) Cells were pretreated with PX-478 (50 μmol/L) (G), a pharmacological inhibitor of HIF1 α , for 2 h, followed by treatment with gAcrp for further 6 h. ADSC were then cocultured with CFSE-labeled splenocytes in presence of mitogen PHA-l. Lymphocyte proliferation was examined on Day 4 of coculture using FACS analysis. (H) ADSC were transfected with Hif1a siRNA (20 nmol/L) for 24 h, and then treated with gAcrp for 6 h. The expression levels of Nos2 , Cox2 , Ptges , Icam1 , and Cd274 were examined by RT-qPCR. (I, J) ADSC were transfected with Hif1a siRNA. After 24 h of incubation, cells were further stimulated with gAcrp for 48 h. (I) The percentage of apoptotic cells was determined using Annexin V binding assay. (J) Expression levels of apoptosis-related proteins were evaluated by Western blot analysis. ∗ P < 0.05 compared to indicated groups; n = 3.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Expressing, Western Blot, Immunolabeling, Microscopy, Software, Transfection, Labeling, Quantitative RT-PCR, Incubation, Binding Assay

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Involvement of HIF1 α -driven glycolytic induction in the enhanced survival and function of ADSC by globular adiponectin. (A–D) ADSC were pretreated with gAcrp for 6 h, followed by stimulation with TNF α /IFN γ (10 and 20 ng/mL, respectively) for additional 24 h, or treated with gAcrp alone for 24 h. Extracellular acidification rate (ECAR) (A) and oxygen consumption rate (OCR) (B) were measured as described in Methods. (C) Relative ECAR and OCR compared to the control were shown. (D) Expression levels of Slc2a1 , Pdk1 , and Ldha were examined by RT-qPCR assay. (E, F) ADSC were transfected with Hif1a siRNA (20 nmol/L) for 24 h, followed by treatment with gAcrp for further 24 h. (E) ECAR and OCR were measured as described above and their values relative to control group were presented. (F) The messenger RNA levels of Slc2a1 , Pdk1 , and Ldha were determined by RT-qPCR. (G) ECAR and OCR were examined in obese ADSC treated with gAcrp in comparison with those in lean counterparts. (H–J) ADSC were pretreated with oligomycin (Omy) or 2-deoxy- d -Glucose (2-DG) at indicated concentrations for 2 h, followed by treatment with gAcrp (1 μg/mL) for additional 24 h. (H, I) The caspase-3 activity was determined as described above. (J) The apoptotic levels were measured using Annexin V binding assay. (K) ADSC were pretreated with Omy and 2-DG as indicated, followed by further incubation with 1 μg/mL of gAcrp for 6 h. ADSC were then cocultured with CFSE-labeled splenocytes in presence of PHA-l. Lymphocyte proliferation was analyzed by CFSE dilution assay. ∗ P < 0.05 compared to indicated groups; n = 3.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Control, Expressing, Quantitative RT-PCR, Transfection, Comparison, Activity Assay, Binding Assay, Incubation, Labeling, Dilution Assay

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Induction of HIF1 α and enhancement of ADSC potency by globular adiponectin are mediated via AdipoR1/p38 MAPK signaling pathways. (A) ADSC were transfected with siRNA targeting Adipor1 or Adipor2 (20 nmol/L) for 36 h, followed by treatment with gAcrp for further 6 h. Gene silencing efficiency was monitored by Western blot analysis (upper panel). Adipor1 - or Adipor2 -deficient ADSC were cocultured with CFSE-labeled splenocytes in presence of PHA-l. The proliferation rate of lymphocytes was measured by flow cytometry analysis (lower panel). (B) The mRNA expression levels of Nos2 , Cox2 , Ptges , Icam1 , and Cd274 were determined by RT-qPCR assay. (C) After treatment with gAcrp as indicated, total and phosphorylated levels of p38, ERK, and JNK were examined by Western blot analysis. (D) ADSC were pretreated with MAPK inhibitors, including U0126 (MEK inhibitor, 10 μmol/L), SB203580 (p38 MAPK inhibitor, 20 μmol/L), and SP600125 (JNK inhibitor, 20 μmol/L) for 2 h, followed by stimulation with gAcrp (1 μg/mL) for further 6 h. ADSC were then cocultured with PHA-activated splenocytes and lymphocyte proliferation was measured by CFSE dilution assay. (E) ADSC were pretreated with SB203580 for 2 h, followed by further treatment with gAcrp for 6 h. The mRNA levels of immunomodulation-related genes were determined by RT-qPCR. (F) ADSC were transfected with Adipor1 siRNA for 36 h, followed by further incubation with gAcrp for 1 h (F). Total and phospho-p38MAPK levels were analyzed by Western blot analysis. (G, H) ADSC were transfected with Adipor1 siRNA for 36 h (G) or pretreated with SB203580 for 2 h (H). The protein expression levels of HIF1 α were determined by Western blot. ∗ P < 0.05 compared to indicated groups; n = 3.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Protein-Protein interactions, Transfection, Western Blot, Labeling, Flow Cytometry, Expressing, Quantitative RT-PCR, Dilution Assay, Incubation

Journal: Acta Pharmaceutica Sinica. B

Article Title: Adiponectin restores the obesity-induced impaired immunomodulatory function of mesenchymal stromal cells via glycolytic reprogramming

doi: 10.1016/j.apsb.2023.10.019

Figure Lengend Snippet: Proposed model for the modulation of immunomodulatory and therapeutic potentials of ADSC by adiponectin under physiological and obese conditions. ADSC isolated from lean mice exert potent immunosuppressive function by regulating T cell proliferation, but inducing differentiation of Treg. In contrast, ADSC derived from obese mice exhibit a compromised immunomodulatory potency, partly due to altered metabolic profile, such as enhanced OXOPHOS, but suppressed glycolysis, which are promoted by obese environment. Pretreatment with adiponectin stimulates the immunomodulatory properties of lean ADSC via glycolytic metabolic reprogramming and enhances the therapeutic efficacy against colitis. Interestingly, adiponectin also results in recovery of dysregulated function of ADSC derived from the obese mice. Mechanistically, adiponectin activates AdipoR1/p38MAPK/HIF1 α signaling pathway and thereby boosts glycolytic capacity in lean ADSC and recovers metabolic profile of obese ADSC. Given that serum adiponectin is significantly lowered in obese individuals, these findings suggest that dysfunction of obese ADSC may be ascribed to impaired adiponectin signaling by excess adiposity and re-activation of adiponectin signaling would be a promising strategy to enhance therapeutic potentials of these multipotent cells.

Article Snippet: Recombinant globular adiponectin (gAcrp; #450–21) was acquired from PeproTech (Rocky Hill, NJ, USA).

Techniques: Isolation, Derivative Assay, Drug discovery, Activation Assay

Journal: Journal of Translational Medicine

Article Title: Adiponectin suppresses tumor growth of nasopharyngeal carcinoma through activating AMPK signaling pathway

doi: 10.1186/s12967-022-03283-0

Figure Lengend Snippet: Circulating adiponectin levels and risk of nasopharyngeal carcinoma in retrospective and prospective cohorts

Article Snippet: Recombination human full-length adiponectin was dissolved in deionized water to prepare a working stock solution of approximately 0.5 mg/mL (BioVendor, Brno, Czech Republic).

Techniques:

Journal: Journal of Translational Medicine

Article Title: Adiponectin suppresses tumor growth of nasopharyngeal carcinoma through activating AMPK signaling pathway

doi: 10.1186/s12967-022-03283-0

Figure Lengend Snippet: Adiponectin suppresses nasopharyngeal carcinoma growth. A , B 1 × 10 6 CNE-2 cells were injected subcutaneously into 5- to 6- week-old adiponectin-deficient nude mice, or the control nude mice ( n = 6 per group). Tumor growth were monitored by measuring the tumor volume for 10 days. Next, the mice were sacrificed, and tumors were collected, measured, weighed. C The plate colony assay was performed to determine colony-formation ability. CNE-2 and C666-1 cells were treated with various concentrations of adiponectin for 7 days. Graphs show the number of colonies. D EdU incorporation assay was performed to determine cell proliferation. CNE-2 and C666-1 cells were treated with various concentrations of adiponectin for 48 h. Bars: 50 μm. Graphs show the relative cell proliferation percentage. E , F CNE-2 and C666-1 cells were incubated with adiponectin (40 μg/mL) for 48 h. At the end of incubation, the cells were collected for FACS analysis. G Western blot analysis of p-AMPKα (T172), p-LKB1 and p-ERk1/2 in cultured CNE-2 and C666-1 cells after the treatment with adiponectin (40 μg/mL) for 30 min. Results are presented as mean ± SD of three independent experiments performed in triplicate. * P < 0.05, ** P < 0.01, *** P < 0.001, # P < 0.0001

Article Snippet: Recombination human full-length adiponectin was dissolved in deionized water to prepare a working stock solution of approximately 0.5 mg/mL (BioVendor, Brno, Czech Republic).

Techniques: Injection, Colony Assay, Incubation, Western Blot, Cell Culture

Journal: Journal of Translational Medicine

Article Title: Adiponectin suppresses tumor growth of nasopharyngeal carcinoma through activating AMPK signaling pathway

doi: 10.1186/s12967-022-03283-0

Figure Lengend Snippet: Adiponectin suppresses proliferation of NPC cells via AMPK activation. A , B CNE-2 and C666-1 cells were pretreated with compound C (10 mM) followed by treatment with adiponectin (40 μg/mL) for 24 h. Cell cycle was then analyzed using flow cytometer. # P < 0.05 and ## P < 0.01 compared to cells treated with adiponectin but not ComC; ** P < 0.01 compared with cells treated without adiponectin and ComC. C CNE-2 and C666-1 cells viability was determined after treatment with or without adiponectin for 48 h in the presence or absence of ComC (10 μM). ** P < 0.01, *** P < 0.001. D CNE-2 and C666-1 cells were pretreated with compound C (10 mM), p-AMPKα (Thr172) protein level was then determined by Western blot analysis after the treatment with adiponectin (40 μg/mL) for 30 min; cyclin D1, p21, and p27 protein level was then determined by Western blot analysis after the cells were exposed to adiponectin (40 μg/mL) for 48 h. E Quantitative analysis of p-AMPKα level was performed by densitometric analysis. Results are presented as mean ± SD of three independent experiments performed in triplicate. ** P < 0.01, *** P < 0.001

Article Snippet: Recombination human full-length adiponectin was dissolved in deionized water to prepare a working stock solution of approximately 0.5 mg/mL (BioVendor, Brno, Czech Republic).

Techniques: Activation Assay, Flow Cytometry, Western Blot

Journal: Journal of Translational Medicine

Article Title: Adiponectin suppresses tumor growth of nasopharyngeal carcinoma through activating AMPK signaling pathway

doi: 10.1186/s12967-022-03283-0

Figure Lengend Snippet: AdipoR1 and AdipoR2 mediate the anti-proliferative effect of adiponectin in NPC cells. A , B Expression of AdipoR1 and AdipoR2 was determined by qRT-PCR and Western blot in NPC cell lines. C , D CNE-2 and C666-1 cells were transfected with 50 μM siRNAs of NC, AdipoR1, or AdipoR2. The relative amounts of each AdipoR1/R2 mRNA against β-actin were measured with qRT-PCR. E Effect of the knockdown of AdipoR1 and AdipoR2 expression on the cell viability of CNE-2 and C666-1 cells treated with or without 40 μg/mL adiponectin for 48 h. F CNE-2 and C666-1 cells were transfected with AdipoR1, AdipoR2 siRNA or NC siRNA and treated with 40 μg/mL adiponectin for 30 min. AMPKα and p-AMPKα (T172) protein levels were determined by Western blot analysis. Quantitative analysis of p-AMPKα level was performed by densitometric analysis and shown in the below part. Results are presented as mean ± SD of three independent experiments performed in triplicate. ** P < 0.01, *** P < 0.001

Article Snippet: Recombination human full-length adiponectin was dissolved in deionized water to prepare a working stock solution of approximately 0.5 mg/mL (BioVendor, Brno, Czech Republic).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Transfection