Journal: Molecular Human Reproduction

Article Title: Metformin: a novel promising option for fertility preservation during cyclophosphamide-based chemotherapy

doi: 10.1093/molehr/gaaa084

Figure Lengend Snippet: The timeline of experiments and the number of ovarian follicles in different stages in each group . ( A ) The timeline of experiments was shown. Metformin (MET), sirolimus (SIRO) and everolimus (EVE) were administered by gavage on Days 1 to 5 for 4 weeks. Cyclophosphamide (CP) was provided after gavage on Day 1 by intraperitoneal injection weekly for 3 weeks. Other experimental details were described in Materials and methods section. ( B – F ) The numbers of ovarian follicles in different stages during folliculogenesis in C57BL/6 mice are shown. The mice were treated with CP-alone, MET-alone, SIRO-alone, or CP in combination with MET, SIRO or EVE. After 4 weeks of treatment, the ovaries were processed into paraffin blocks, sectioned, mounted and hematoxylin and eosin (H&E) stained for follicular counting. The number of primordial and tertiary follicles, and corpus luteum decreased in the CP-alone group compared with the control group ( P = 0.0014, 0.00003, 0.073, respectively). The deleterious effects of CP on follicular counts were diminished when oral MET was given to mice (Primordial follicle: P = 0.0274). The other two specific mTOR inhibitors, SIRO and EVE, also exhibited significant protective effects against CP damage (primary follicles: CP-alone versus CP + SIRO: P = 0.024; tertiary follicles: CP-alone versus CP + EVE: P = 0.0046). (B) and (C) The Y axis represented the average follicular counts per high-power field (HPF). N = 10 mice in the control and CP-alone group, while n = 5 mice in the other groups. Each value represents the average of 2–3 HPF per animal. (D)–(F) The Y axis represented the total follicular counts per ovarian section. N = 10 mice in the control and CP-alone group, while n = 5 mice in the other groups. Data are expressed as the mean ± standard deviation. Statistical analyses were performed by nonparametric Kruskal–Wallis test with Dunn's post-hoc for multiple comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001. Note: The double slash mark on the X axis separated the MET-alone and SIRO-alone group from other groups because these two control groups were run in a separate experiment.

Article Snippet: For in-vitro studies, metformin hydrochloride (dissolved in PBS) and CP (dissolved in DMSO) was purchased from Sigma (St. Louis, MO, USA) and Tocris Bioscience (Bristol, UK), respectively.

Techniques: Injection, Staining, Control, Standard Deviation

Journal: Molecular Human Reproduction

Article Title: Metformin: a novel promising option for fertility preservation during cyclophosphamide-based chemotherapy

doi: 10.1093/molehr/gaaa084

Figure Lengend Snippet: The serum hormone concentration and the number of offspring in each group. C57BL/6 mice were treated with CP-alone, metformin (MET)-alone, sirolimus (SIRO)-alone, or CP in combination with MET, SIRO or everolimus (EVE). After 4 weeks of treatment, the mice were sacrificed and serum was collected for hormonal determination. ( A ) The serum levels of anti-Müllerian hormone (AMH) were significantly decreased in the CP-alone group (compared with control group: P < 0.0001). Although the AMH levels increased in CP + MET, CP + SIRO and CP + EVE group, the data did not reach statistical significance probably due to limited case number. ( B ) The serum levels of estradiol were significantly decreased in the CP-alone group ( P < 0.0001), and tended to increase in CP + MET group, CP + SIRO group and CP + EVE group. ( C ) The serum levels of progesterone were significantly decreased in the CP-alone group ( P < 0.0001) and tended to increase in CP + MET group and CP + EVE group. ( D ) A breeding test was conducted 1 week after the 4-week treatment. Only one round of timed mating was conducted per female mouse and the outcome of the first pregnancy in each mouse was evaluated. The number of the offspring was significantly decreased in the CP-alone group ( P < 0.0001) and tended to increase in the CP + MET group. In (A)–(C), n = 10 mice in the control and CP-alone group, while n = 5 mice in the other groups. In (D), n = 6 mice per group. Data are expressed as the mean ± standard deviation. Statistical analyses were performed by nonparametric Kruskal–Wallis test with Dunn's post-hoc for multiple comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. Note: The double slash mark on the X axis separated the MET-alone and SIRO-alone group from other groups because these two control groups were run in a separate experiment.

Article Snippet: For in-vitro studies, metformin hydrochloride (dissolved in PBS) and CP (dissolved in DMSO) was purchased from Sigma (St. Louis, MO, USA) and Tocris Bioscience (Bristol, UK), respectively.

Techniques: Concentration Assay, Control, Standard Deviation

Journal: Molecular Human Reproduction

Article Title: Metformin: a novel promising option for fertility preservation during cyclophosphamide-based chemotherapy

doi: 10.1093/molehr/gaaa084

Figure Lengend Snippet: Immunohistochemical (IHC) staining of the proteins involved in mTOR signaling pathway. C57BL/6 mice were treated with CP-alone, metformin (MET)-alone, sirolimus (SIRO)-alone or CP in combination with MET or SIRO. After 4 weeks of treatment, the ovaries were processed into paraffin sections for the IHC detection of p-mTOR ( A , B ) and p-AMPK ( C , D ) proteins. The percentage of granulosa cells with positive staining was calculated by dividing the number of positive stained cells with the total number of granulosa cells in ovarian follicles under a microscopy at ×400 magnification. One largest tertiary follicle in each section was selected to calculate the number of positively stained cells. Five sections per mice were counted from a total of 5 mice in each group (n = 5 mice). Data are expressed as the mean (%) ± standard deviation. Statistical analyses were performed by nonparametric Kruskal–Wallis test with Dunn's post-hoc for multiple comparisons. * P < 0.05, ** P < 0.01. The scale bar is 50 μm. Note: The double slash mark on the X axis separated the MET-alone and SIRO-alone group from other groups because these two control groups were run in a separate experiment.

Article Snippet: For in-vitro studies, metformin hydrochloride (dissolved in PBS) and CP (dissolved in DMSO) was purchased from Sigma (St. Louis, MO, USA) and Tocris Bioscience (Bristol, UK), respectively.

Techniques: Immunohistochemical staining, Immunohistochemistry, Staining, Microscopy, Standard Deviation, Control

Journal: Molecular Human Reproduction

Article Title: Metformin: a novel promising option for fertility preservation during cyclophosphamide-based chemotherapy

doi: 10.1093/molehr/gaaa084

Figure Lengend Snippet: IHC staining of the cellular apoptotic and proliferative markers. C57BL/6 mice were treated with CP-alone, metformin (MET)-alone, sirolimus (SIRO)-alone or CP in combination with MET or SIRO. After 4 weeks of treatment, the ovaries were processed into paraffin sections for the TUNEL assay (A, B) and Ki67 staining (C, D) to separately evaluate the degree of cellular apoptosis and proliferation. The percentage of granulosa cells with positive staining was calculated by dividing the number of positive stained cells with the total number of granulosa cells in ovarian follicles under a microscopy at ×400 magnification. One largest tertiary follicle in each section was selected to calculate the number of positively stained cells. Five sections per mice were counted. N = 10 mice in the control and CP-alone group, while n = 5 mice in the other groups. Data are expressed as the mean (%) ± standard deviation. Statistical analyses were performed by nonparametric Kruskal–Wallis test with Dunn's post-hoc for multiple comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. The scale bar is 50 μm. Note: The double slash mark on the X axis separated the MET-alone and SIRO-alone group from other groups because these two control groups were run in a separate experiment.

Article Snippet: For in-vitro studies, metformin hydrochloride (dissolved in PBS) and CP (dissolved in DMSO) was purchased from Sigma (St. Louis, MO, USA) and Tocris Bioscience (Bristol, UK), respectively.

Techniques: Immunohistochemistry, TUNEL Assay, Staining, Microscopy, Control, Standard Deviation

Journal: Molecular Human Reproduction

Article Title: Metformin: a novel promising option for fertility preservation during cyclophosphamide-based chemotherapy

doi: 10.1093/molehr/gaaa084

Figure Lengend Snippet: Cell cycle phase analysis by flow cytometry using cultured mouse granulosa cells. Mouse granulosa cells were treated with CP-alone (1ug/ml) or with CP + metformin (MET) (10 mM) or with MET-alone for 72 h. Fresh granulosa cells were collected from five mice, and then mixed and cultured together. ( A ) The cell cycles, including subG1 (apoptotic phase), G1, S and G2/M of the assayed cells were determined by quantifying their DNA contents with propidium iodide (PI) staining and flow cytometry. The M1, M2, M3 and M4 bars represent the subG1, G1, S and G2/M phases, respectively. ( B ) The quantitative results of the cell cycle phases in each group were shown. The percentage of cells in the subG1 phase was significantly increased in the CP group ( P = 0.0009) and tended to decrease when cells were cotreated with MET. There was no significant difference between control group and MET-alone group. All the experiments were repeated five times and therefore N = 5 per group. Statistical analyses were performed by nonparametric Kruskal–Wallis test with Dunn's post-hoc for multiple comparisons. * P < 0.05, *** P < 0.001.

Article Snippet: For in-vitro studies, metformin hydrochloride (dissolved in PBS) and CP (dissolved in DMSO) was purchased from Sigma (St. Louis, MO, USA) and Tocris Bioscience (Bristol, UK), respectively.

Techniques: Flow Cytometry, Cell Culture, Staining, Control

Journal: Molecular Human Reproduction

Article Title: Metformin: a novel promising option for fertility preservation during cyclophosphamide-based chemotherapy

doi: 10.1093/molehr/gaaa084

Figure Lengend Snippet: Metformin exerted AMPK/p53/p21-mediated anti-apoptotic effect on cultured mouse granulosa cells. ( A ) Mouse granulosa cells were treated with p53 siRNA (25 nM) or control siRNA for 24 h prior CP (1ug/ml) or metformin (MET) (10 mM) treatment, after 72 h. Apoptotic cells were determined by flow cytometry with propidium iodide staining and quantified by the subG1 ratio. The experiments were repeated five times and therefore N = 5 per group. The apoptosis rate significantly increased in the CP-alone group ( P = 0.0029) and tended to decrease after cotreating the cells with MET. The anti-apoptosis effect of MET was diminished after blocking the p53 activity with siRNA. ( B ) The experimental conditions were the same as A, except that p21 siRNA (25 nM) was applied instead of p53 siRNA. ( C ) Mouse granulosa cells were treated with AMPK inhibitor BML275(10 μM) for 30 min prior MET (10 mM) treatment. At indicated time periods, the expression of p53 mRNA was determined by qRT-PCR. The experiments were repeated four times and therefore N = 4 per group. Sequential measurement of p53 mRNA expression in culture granulosa cells significantly increased 8 h after the addition of MET ( P = 0.0098). The expression of p53 mRNA was blocked with the addition of cell-permeable AMPK inhibitor BML. ( D ) The expression of p21 mRNA was determined by qRT-PCR and sequential measurement of p21 mRNA expression in cultured granulosa cells significantly increased 24 h after the addition of MET ( P = 0.0095). The experiments were repeated five times and therefore N = 5 per group. ( E ) C57BL/6 mice were treated with CP-alone or in combination with MET or MET-alone. After 4 weeks of treatment, the ovaries were processed into paraffin sections for the p53 and p21 staining. Both p53 and p21 activity were generally low in normal untreated ovarian tissue but was elevated after CP treatment (p53: P = 0.6529; p21: P = 0.0452) as determined with IHC staining under microscopic examination at ×400 magnification. The coadministration of MET with CP increased p53 and p21 protein expression in the ovarian tissue even higher ( P < 0.0004 in both). Statistical analyses were performed by nonparametric Kruskal–Wallis test with Dunn's post-hoc for multiple comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001. ( F ) Representative p53 and p21 IHC images of each group were shown. The scale bar is 50 μm.

Article Snippet: For in-vitro studies, metformin hydrochloride (dissolved in PBS) and CP (dissolved in DMSO) was purchased from Sigma (St. Louis, MO, USA) and Tocris Bioscience (Bristol, UK), respectively.

Techniques: Cell Culture, Control, Flow Cytometry, Staining, Blocking Assay, Activity Assay, Expressing, Quantitative RT-PCR, Immunohistochemistry

Journal: International Journal of Applied and Basic Medical Research

Article Title: Formulation and design of sustained release matrix tablets of metformin hydrochloride: Influence of hypromellose and polyacrylate polymers

doi: 10.4103/2229-516x.112242

Figure Lengend Snippet: Figure 2: Infrared spectra of Metformin hydrochloride

Article Snippet: Metformin HCl was procured from Tocris bioscience (USA).

Techniques:

Journal: International Journal of Applied and Basic Medical Research

Article Title: Formulation and design of sustained release matrix tablets of metformin hydrochloride: Influence of hypromellose and polyacrylate polymers

doi: 10.4103/2229-516x.112242

Figure Lengend Snippet: Figure 4: Differential scanning calorimetry study of (a) Pure Metformin hydrochloride (b) Anhydrous dicalcium phosphate (c) (HPMC) K200M (d) Eudragit RS100 (e) Kollidon 30 (f) HPMC K15M (g) Formulation F6

Article Snippet: Metformin HCl was procured from Tocris bioscience (USA).

Techniques: Differential Scanning Calorimetry, Formulation

Journal: Diabetes

Article Title: Beneficial Endocrine but Adverse Exocrine Effects of Sitagliptin in the Human Islet Amyloid Polypeptide Transgenic Rat Model of Type 2 Diabetes

doi: 10.2337/db09-0058

Figure Lengend Snippet: Fasting plasma glucose ( A ), body weight ( B ), mean daily food intake ( C ), and mean daily drug consumption ( D ) after 12-week treatment with 60% high-fat chow diet in wild-type (WT) rats ( n = 7), HIP rats ( n = 8), HIP rats treated with sitagliptin (HIP+SIT; n = 8), HIP rats treated with metformin (HIP+MET; n = 9), and HIP rats treated with combination therapy (HIP+SIT+MET; n = 8). Data are means ± SE.

Article Snippet: Sitagliptin was provided by Merck Research (Rahway, NJ), and metformin was purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques: Clinical Proteomics

Journal: Diabetes

Article Title: Beneficial Endocrine but Adverse Exocrine Effects of Sitagliptin in the Human Islet Amyloid Polypeptide Transgenic Rat Model of Type 2 Diabetes

doi: 10.2337/db09-0058

Figure Lengend Snippet: A : Typical islets from wild-type (WT) rats, HIP rats, HIP rats treated with sitagliptin (HIP+SIT), and HIP rats treated with sitagliptin and metformin (HIP+SIT+MET) stained for insulin (pink) and hematoxylin (blue). β-Cell area ( B ) and mean β-cell mass ( C ) after 12-week treatment with 60% high-fat diet in wild-type rats ( n = 7), HIP rats ( n = 8), HIP rats treated with sitagliptin (HIP+SIT; n = 8), HIP rats treated with metformin (HIP+MET; n = 9), and HIP rats treated with combination therapy (HIP+SIT+MET; n = 8). Data are the means ± SE. (A high-quality digital representation of this figure is available in the online issue.)

Article Snippet: Sitagliptin was provided by Merck Research (Rahway, NJ), and metformin was purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques: Staining

Journal: Diabetes

Article Title: Beneficial Endocrine but Adverse Exocrine Effects of Sitagliptin in the Human Islet Amyloid Polypeptide Transgenic Rat Model of Type 2 Diabetes

doi: 10.2337/db09-0058

Figure Lengend Snippet: A : Examples of islets stained for insulin (pink) and replication marker Ki-67 (brown) and nuclear stain hematoxylin (blue) imaged at 20×. B : Frequency of β-cell replication in wild-type (WT) rats ( n = 7), HIP rats ( n = 8), HIP rats treated with sitagliptin (HIP+SIT; n = 8), HIP rats treated with metformin (HIP+MET; n = 9), and HIP rats treated with combination therapy (HIP+SIT+MET; n = 8). C : Examples of islets stained for insulin (green) and apoptosis marker (TUNEL; red) and nuclear stain (DAPI; blue) imaged at ×20. D : Frequency of β-cell apoptosis in wild-type rats ( n = 7), HIP rats ( n = 8), HIP rats treated with sitagliptin ( n = 8), HIP rats treated with metformin ( n = 9), and HIP rats treated with combination therapy ( n = 8). Data are means ± SE. * P < 0.05 vs. wild type, HIP, and HIP plus metformin. Arrows indicate examples of insulin-positive Ki-67 and TUNEL-positive cells. (A high-quality digital representation of this figure is available in the online issue.)

Article Snippet: Sitagliptin was provided by Merck Research (Rahway, NJ), and metformin was purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques: Staining, Marker, TUNEL Assay

Journal: Diabetes

Article Title: Beneficial Endocrine but Adverse Exocrine Effects of Sitagliptin in the Human Islet Amyloid Polypeptide Transgenic Rat Model of Type 2 Diabetes

doi: 10.2337/db09-0058

Figure Lengend Snippet: Mean glucose infusion rates during the hyperinsulinemic-euglycemic clamp after 12-week treatment with 60% high-fat diet (HFD) in wild-type (WT) rats ( n = 5), HIP rats ( n = 6), HIP rats treated with sitagliptin (HIP+SIT; n = 7), HIP rats treated with metformin (HIP+MET; n = 6), and HIP rats treated with combination therapy (HIP+SIT+MET; n = 6). Data are means ± SE.

Article Snippet: Sitagliptin was provided by Merck Research (Rahway, NJ), and metformin was purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques:

Journal: Diabetes

Article Title: Beneficial Endocrine but Adverse Exocrine Effects of Sitagliptin in the Human Islet Amyloid Polypeptide Transgenic Rat Model of Type 2 Diabetes

doi: 10.2337/db09-0058

Figure Lengend Snippet: Mean first-phase insulin response during the hyperglycemic clamp ( A ), mean disposition index ( B ), mean first-phase insulin response to arginine ( C ), and the relationship between β-cell mass and first-phase insulin response to arginine ( D ) after 12-week treatment with 60% high-fat diet in wild-type (WT) rats ( n = 6), HIP rats ( n = 8), HIP rats treated with sitagliptin (HIP+SIT; n = 8), HIP rats treated with metformin (HIP+MET; n = 6), and HIP rats treated with combination therapy (HIP+SIT+MET; n = 6). Data are means ± SE.

Article Snippet: Sitagliptin was provided by Merck Research (Rahway, NJ), and metformin was purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques:

Journal: Diabetes

Article Title: Beneficial Endocrine but Adverse Exocrine Effects of Sitagliptin in the Human Islet Amyloid Polypeptide Transgenic Rat Model of Type 2 Diabetes

doi: 10.2337/db09-0058

Figure Lengend Snippet: Increased ductal cell turnover in HIP rats treated with sitagliptin. Quantification of ductal cell replication ( A ) and apoptosis ( B ) in wild-type rats, HIP rats, and HIP rats treated with either sitagliptin (HIP+SIT), metformin (HIP+MET), or combination therapy of sitagliptin and metformin (HIP+SIT+MET) for 12 weeks. C : Regression analysis of the relationships between ductal cell proliferation versus fasting plasma glucose. Note that ductal cell replication in sitagliptin-treated rats was quantified only in metaplasia and pancreatitis–free areas of the exocrine pancreas. * P < 0.05.

Article Snippet: Sitagliptin was provided by Merck Research (Rahway, NJ), and metformin was purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques: Clinical Proteomics

Journal: Cancer Medicine

Article Title: Biguanides in combination with olaparib limits tumorigenesis of drug‐resistant ovarian cancer cells through inhibition of Snail

doi: 10.1002/cam4.2738

Figure Lengend Snippet: Biguanides alone and/or in combination with olaparib inhibit OC cell proliferation. A2780PAR and A2780CR ovarian cancer cells were treated for 72 hours and cell proliferation was determined using alamarBlue assays. (A) Phenformin (0‐1 mmol/L), (B) metformin (0‐5 mmol/L), (C) olaparib (0‐2 µmol/L), phenformin‐olaparib (D and E) and metformin‐olaparib (F and G). Data represent the mean ± SEM of triplicate of three independent experiments, statistical significance is represented by *( P < .05) and **( P < .01)

Article Snippet: Phenformin Hydrochloride and Metformin Hydrochloride were bought from Toronto Research Chemicals.

Techniques:

Journal: Cancer Medicine

Article Title: Biguanides in combination with olaparib limits tumorigenesis of drug‐resistant ovarian cancer cells through inhibition of Snail

doi: 10.1002/cam4.2738

Figure Lengend Snippet: Biguanides alone or in combination with olaparib inhibit OC cell colony formation. A2780PAR and A2780CR ovarian cancer cells were treated for 7 days and survival was determined using clonogenic assays. (A) Phenformin, (B) metformin, (C) olaparib, (D and E) phenformin‐olaparib (F and G) and metformin‐olaparib. The evaluation of combination index for A2780PAR (H) or A2780CR (I) treated with phenformin or metformin and olaparib was calculated where CI < 1 indicates synergy between the two drugs and CI > 1 indicates an additive effect. Results are presented as mean ± SEM for triplicate of three independent experiments

Article Snippet: Phenformin Hydrochloride and Metformin Hydrochloride were bought from Toronto Research Chemicals.

Techniques:

Journal: Cancer Medicine

Article Title: Biguanides in combination with olaparib limits tumorigenesis of drug‐resistant ovarian cancer cells through inhibition of Snail

doi: 10.1002/cam4.2738

Figure Lengend Snippet: Biguanides alone or in combination with olaparib inhibit OC cell migration. A2780PAR and/or A2780CR ovarian cancer cells were treated for 24 hours and wound healing assays were performed to evaluate cell migration. (A) Phenformin, (B) metformin, (C) olaparib (upper panel depicts wound healing assay after treatments and the graph below its quantification), (D and E) phenformin‐olaparib (F and G) and metformin‐olaparib (left panel depicts wound healing assay after treatments and the right graph its quantification). Data represent the mean ± SEM of triplicate of three independent experiments, statistical significance is represented by *( P < .05)

Article Snippet: Phenformin Hydrochloride and Metformin Hydrochloride were bought from Toronto Research Chemicals.

Techniques: Migration, Wound Healing Assay

Journal: Cancer Medicine

Article Title: Biguanides in combination with olaparib limits tumorigenesis of drug‐resistant ovarian cancer cells through inhibition of Snail

doi: 10.1002/cam4.2738

Figure Lengend Snippet: Biguanide treatment inhibits the expression of EMT markers. Protein expression levels of mesenchymal markers (Slug, Snail, Vimentin, N‐cadherin, Twist and Fibronectin) and the epithelial marker E‐cadherin in A2780PAR and A2780CR ovarian cancer cells treated for 72 hours with (A) phenformin, (B) metformin, (C) phenformin‐olaparib, (D) metformin‐olaparib. Bands were quantified using Image J software and normalized to β‐actin. Data represent the mean ± SEM of triplicate of three independent experiments

Article Snippet: Phenformin Hydrochloride and Metformin Hydrochloride were bought from Toronto Research Chemicals.

Techniques: Expressing, Marker, Software

Journal: Cancer Medicine

Article Title: Biguanides in combination with olaparib limits tumorigenesis of drug‐resistant ovarian cancer cells through inhibition of Snail

doi: 10.1002/cam4.2738

Figure Lengend Snippet: Cell migration is inhibited upon Snail knock‐down and enhanced by biguanide treatment. (A) Protein expression levels of mesenchymal markers (Snail, Vimentin, N‐cadherin and Twist) and E‐cadherin in A2780CR, A2780CR/shVector and A2780CR/shSnail 10‐2 ovarian cancer cells. A2780CR/shVector, A2780CR/shSnail 10‐2 ovarian cancer cells were treated for 24 hours and wound healing assay was performed to evaluate cell migration: (B) phenformin, phenformin‐olaparib 0.1 µmol/L and phenformin‐olaparib 0.5 µmol/L. (C) Metformin, metformin‐olaparib 0.1 µg, and metformin‐olaparib 0.5 µmol/L. Data represent the mean ± SEM for triplicate of three independent experiments, statistical significance is represented by *( P < 0.05) and **( P < 0.01).

Article Snippet: Phenformin Hydrochloride and Metformin Hydrochloride were bought from Toronto Research Chemicals.

Techniques: Migration, Knockdown, Expressing, Wound Healing Assay

Journal: Cancer Medicine

Article Title: Biguanides in combination with olaparib limits tumorigenesis of drug‐resistant ovarian cancer cells through inhibition of Snail

doi: 10.1002/cam4.2738

Figure Lengend Snippet: Cell survival is inhibited upon Snail knock‐down and enhanced by biguanide treatment. A2780CR/shVector, A2780CR/shSnail 10‐2 ovarian cancer cells were treated for 7 days with biguanides alone or in combination with olaparib and survival was determined using clonogenic assays. (A) Phenformin, phenformin‐olaparib 0.1 µmol/L and phenformin‐olaparib 0.5 µmol/L. (B) Metformin, metformin‐olaparib 0.1 µmol/L, and metformin‐olaparib 0.5 µmol/L. The evaluation of combination index for A2780CR/shVector (C) or A2780CR/shSnail 10‐2, (D) treated with phenformin or metformin and olaparib was calculated where CI < 1 indicates synergy between the two drugs and CI > 1 indicates an additive effect. Data represent the mean ± SEM for triplicate of three independent experiments, statistical significance is represented by *( P < .05) and **( P < .01)

Article Snippet: Phenformin Hydrochloride and Metformin Hydrochloride were bought from Toronto Research Chemicals.

Techniques: Knockdown

Journal: Molecular Cancer Therapeutics

Article Title: Simvastatin-Induced Apoptosis in Osteosarcoma Cells: A Key Role of RhoA-AMPK/p38 MAPK Signaling in Antitumor Activity

doi: 10.1158/1535-7163.mct-16-0499

Figure Lengend Snippet: Figure 5. Additive effects of simvastatin and metformin on AXT cell proliferation as well as on RhoA, AMPK, and p38 MAPK signaling. A and B, Viability of AXT cells treated with the indicated concentrations of simvastatin and metformin (Met; A) or with simvastatin, metformin, and mevalonate (B) for 2 days. C, Immunoblot analysis of RhoA and RhoA-GTP in AXT cells treated with the indicated reagents for 12 hours. D, Immunoblot analysis of RhoA in membrane and cytosolic fractions of AXT cells treated with simvastatin and metformin for 12 hours. E and F, Immunoblot analysis of AMPK and p38 MAPK phosphorylation as well as of caspase-3 cleavage in AXT cells treated with 0.5 mmol/L simvastatin and the indicated concentrations of metformin for 12 hours (E) or 24 hours (F). The arrow indicates p-p38 MAPK.

Article Snippet: Simvastatin was obtained from Combi-Blocks, metformin from LKT Laboratories, rosuvastatin and atorvastatin fromWako, fluvastatin from Tocris Bioscience, and 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) from Cell Signaling.

Techniques: Western Blot, Membrane, Phospho-proteomics

Journal: Molecular Cancer Therapeutics

Article Title: Simvastatin-Induced Apoptosis in Osteosarcoma Cells: A Key Role of RhoA-AMPK/p38 MAPK Signaling in Antitumor Activity

doi: 10.1158/1535-7163.mct-16-0499

Figure Lengend Snippet: Figure 6. Antitumor effect of simvastatin in vivo. A and B, Immunoblot analysis of ACC phosphorylation in AXT cells treated with the indicated concentrations of simvastatin for 24 hours (A) or with 10 mmol/L simvastatin in the absence or presence of mevalonate, GGPP, or FPP for 20 hours (B). C, Immunoblot analysis of AMPK and ACC phosphorylation in AXT cells treated with 5 mmol/L simvastatin either alone or together with the indicated concentrations of 5-iodotubercidin for 20 hours. D, Viability of AXT cells treated with simvastatin or TOFA as indicated. E and F, Weight of AXT cell–derived tumors formed in mice fed a fat-free diet and injected intraperitoneally with saline (control), simvastatin (50 mg/kg), metformin (100 mg/kg), or both simvastatin (50 mg/kg) and metformin (50 mg/kg). Data are means SD (n ¼ 10), with the P values being for comparison with control (Student t test). The excised tumors are also shown (F). G, Immunoblot analysis of AMPK and p38 MAPK phosphorylation in 5 randomly selected tumors of each group in E.

Article Snippet: Simvastatin was obtained from Combi-Blocks, metformin from LKT Laboratories, rosuvastatin and atorvastatin fromWako, fluvastatin from Tocris Bioscience, and 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) from Cell Signaling.

Techniques: In Vivo, Western Blot, Phospho-proteomics, Derivative Assay, Injection, Saline, Control, Comparison

Journal: Oncology Research

Article Title: Glutamic Acid–Chelated Cobalt Stabilizes G-Quadruplexes and Selectively Suppresses Hepatocellular Carcinoma Growth

doi: 10.32604/or.2026.074144

Figure Lengend Snippet: (AI-PRS optimization of combination therapies: GACC, metformin, and regorafenib. ( A–C ) Viability responses for the tested 3 × 3 × 3 dose grid of GACC (9.54, 28.61, 85.85 µM), metformin (0.85, 2.55, and 7.65 mM), and regorafenib (0.5, 1.5, 4.5 µM) in hepatoma cells (PLC5) and non-transformed hepatocytes (THLE-2), as indicated. ( D ) AI-phenotypic response surface (AI-PRS) fit across the dose grid used to identify dose regions with improved efficacy–toxicity balance. ( E ) Observed vs. AI-PRS–predicted viability for the off-grid optimal dose combination in PLC/PRF/5 and THLE-2. Data are presented as mean ± SEM; n values and statistical methods are provided in . Statistical annotations: ns, not significant; * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.)

Article Snippet: Metformin hydrochloride (MedChemExpress, HY-17471A/CS-1851; CAS 1115-70-4; MedChemExpress, Monmouth Junction, NJ, USA), regorafenib (TargetMol, T1792; CAS 755037-03-7, TargetMol, Boston, MA, USA), and pyridostatin (RR82) trifluoroacetate salt (PDS, Selleck Chemicals, S7444; Houston, TX, USA) were dissolved in dimethyl sulfoxide (DMSO, Sigma-Aldrich, D4540; St. Louis, MO, USA) to prepare stock solutions and diluted into assay media immediately before use.

Techniques: Transformation Assay

Journal: Oncology Research

Article Title: Glutamic Acid–Chelated Cobalt Stabilizes G-Quadruplexes and Selectively Suppresses Hepatocellular Carcinoma Growth

doi: 10.32604/or.2026.074144

Figure Lengend Snippet: Combination therapies of GACC, metformin, and regorafenib for Hepatocellular Carcinoma (HCC) treatment in tert transgenic zebrafish larvae and HBx, src, p53-, RPIA (HSPR) adult zebrafish model. ( A ) Evaluation of drug-induced side effects by measuring body length in tert transgenic zebrafish larvae before and after treatment, assessing potential toxicity and systemic impact. ( B – D ) qPCR analysis of cell proliferation markers ( ccne1 , cdk1 , cdk2 ) in tert transgenic zebrafish larvae, demonstrating significant downregulation following combination therapy. ( E , F ) Histological examination of liver cancer hallmarks in HSPR transgenic zebrafish using hematoxylin and eosin (H&E) staining, revealing a significant reduction in mitotic figures, karyomegalic cells, and hepatocellular dysplasia following combination therapy. n = 10 embryos/group; one-way ANOVA with Dunnett’s post-hoc vs. vehicle; p thresholds as shown. * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.001, ns, not significant.

Article Snippet: Metformin hydrochloride (MedChemExpress, HY-17471A/CS-1851; CAS 1115-70-4; MedChemExpress, Monmouth Junction, NJ, USA), regorafenib (TargetMol, T1792; CAS 755037-03-7, TargetMol, Boston, MA, USA), and pyridostatin (RR82) trifluoroacetate salt (PDS, Selleck Chemicals, S7444; Houston, TX, USA) were dissolved in dimethyl sulfoxide (DMSO, Sigma-Aldrich, D4540; St. Louis, MO, USA) to prepare stock solutions and diluted into assay media immediately before use.

Techniques: Transgenic Assay, Staining