An improved 3-(4,5-dmethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl )-2H-tetrazolium proliferation assay to overcome the interference of hydralazine
Description:
The MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay is one of the most commonly used assays to assess cell proliferation and cytotoxicity, but is subject to interference by testing compounds. Hydralazine, an antihypertensive drug, is commonly investigated in multiple fields such as heart failure, cancer, and blood pressure research. This study reported interference of the MTS assay by hydralazine and a simple modification overcoming this interference. Vascular smooth muscle cells were cultured in the presence or absence of hydralazine (0, 10, 50,100, and 500 mu M) for 2 or 24 h. Cell numbers were analyzed using MTS, trypan blue exclusion, or microscopic assays. A modified version of the standard MTS assay was established, in which an additional step was added replacing the test medium, containing hydralazine, with fresh culture medium immediately before the addition of the MTS reagent. Culture with hydralazine at concentrations of 50, 100, and 500 mu M for 2 h increased absorbance (p< 0.05) in the standard MTS assay, whereas microscopy suggested no change in cell numbers. Culture with 500 mu m hydralazine for 24 h increased absorbance (p< 0.05) in the standard MTS assay, however, trypan blue exclusion and microscopy suggested a decrease in cell numbers. In a cell-free system, hydralazine (>= 10 mu M) increased absorbance in a concentration-dependent manner. The modified MTS assay produced results consistent with trypan blue exclusion and microscopy. In conclusion, a simple modification of the standard MTS assay overcame the interference of hydralazine and may be useful to avoid interference from other tested compounds.
Description:
The MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay is one of the most commonly used assays to assess cell proliferation and cytotoxicity, but is subject to interference by testing compounds. Hydralazine, an antihypertensive drug, is commonly investigated in multiple fields such as heart failure, cancer, and blood pressure research. This study reported interference of the MTS assay by hydralazine and a simple modification overcoming this interference. Vascular smooth muscle cells were cultured in the presence or absence of hydralazine (0, 10, 50,100, and 500 mu M) for 2 or 24 h. Cell numbers were analyzed using MTS, trypan blue exclusion, or microscopic assays. A modified version of the standard MTS assay was established, in which an additional step was added replacing the test medium, containing hydralazine, with fresh culture medium immediately before the addition of the MTS reagent. Culture with hydralazine at concentrations of 50, 100, and 500 mu M for 2 h increased absorbance (p< 0.05) in the standard MTS assay, whereas microscopy suggested no change in cell numbers. Culture with 500 mu m hydralazine for 24 h increased absorbance (p< 0.05) in the standard MTS assay, however, trypan blue exclusion and microscopy suggested a decrease in cell numbers. In a cell-free system, hydralazine (>= 10 mu M) increased absorbance in a concentration-dependent manner. The modified MTS assay produced results consistent with trypan blue exclusion and microscopy. In conclusion, a simple modification of the standard MTS assay overcame the interference of hydralazine and may be useful to avoid interference from other tested compounds.
Description:
Thoracic aortic aneurysm (TAA) has a prevalence of 0.16–0.34% and an incidence of 7.6 per 100,000 person-years, accounting for 1–2% of all deaths in Western countries. Currently, no effective pharmacological therapies have been identified to slow TAA development and prevent TAA rupture. Large TAAs are treated with open surgical repair and less invasive thoracic endovascular aortic repair, both of which have high perioperative mortality risk. Therefore, there is an urgent medical need to identify the cellular and molecular mechanisms underlying TAA development and rupture to develop new therapies. In this review, we summarize animal TAA models including recent developments in porcine and zebrafish models: porcine models can assess new therapeutic devices or intervention strategies in a large mammal and zebrafish models can employ large-scale small-molecule suppressor screening in microwells. The second part of the review covers current views of TAA pathogenesis, derived from recent studies using these animal models, with a focus on the roles of the transforming growth factor-beta (TGF
Description:
Thoracic aortic aneurysm (TAA) has a prevalence of 0.16–0.34% and an incidence of 7.6 per 100,000 person-years, accounting for 1–2% of all deaths in Western countries. Currently, no effective pharmacological therapies have been identified to slow TAA development and prevent TAA rupture. Large TAAs are treated with open surgical repair and less invasive thoracic endovascular aortic repair, both of which have high perioperative mortality risk. Therefore, there is an urgent medical need to identify the cellular and molecular mechanisms underlying TAA development and rupture to develop new therapies. In this review, we summarize animal TAA models including recent developments in porcine and zebrafish models: porcine models can assess new therapeutic devices or intervention strategies in a large mammal and zebrafish models can employ large-scale small-molecule suppressor screening in microwells. The second part of the review covers current views of TAA pathogenesis, derived from recent studies using these animal models, with a focus on the roles of the transforming growth factor-beta (TGF
Description:
The association between constipation and cardiovascular risk is unclear. This population-level matched cohort study compared the association of constipation with hypertension and incident cardiovascular events in 541,172 hospitalized patients aged
Description:
The association between constipation and cardiovascular risk is unclear. This population-level matched cohort study compared the association of constipation with hypertension and incident cardiovascular events in 541,172 hospitalized patients aged
Description:
•The LDL-C reference interval was 1.48–3.77 mmol/L in Chinese adults.•Hypocholesterolemia was associated with a 57% higher risk for diabetes.•Hypercholesterolemia was associated with a 29% higher risk for diabetes. This study aimed to investigate whether both high and low levels of low-density lipoprotein cholesterol (LDL-C), i.e., hypercholesterolemia and hypocholesterolemia, were associated with diabetes in Chinese adults. This cross-sectional study included 22,557 Chinese adults. The LDL-C reference interval was determined from a healthy sub-cohort. Associations between hypocholesterolemia or hypercholesterolemia with diabetes were analyzed using binary logistic regression. The LDL-C reference interval was 1.48–3.77 mmol/L (57.23–145.78 mg/dL). Therefore, hypocholesterolemia, normocholesterolemia, and hypercholesterolemia were defined as an LDL-C concentration of <1.48, 1.48–3.77, and >3.77 mmol/L, respectively. Prevalence of diabetes was higher in people with hypocholesterolemia or hypercholesterolemia than that in people with normocholesterolemia. Hypocholesterolemia was associated with an increased multivariable-adjusted risk for diabetes diagnosis (odds ratio, 1.57 95% confidence interval, 1.18–2.08), and so was hypercholesterolemia (odds ratio, 1.29 95% confidence interval, 1.10–1.51). The results remained significant after exclusion of those who took lipid-lowering drugs from the analysis. This study demonstrated that both low and high levels of LDL-C were associated with a higher risk of diabetes diagnosis. Patients with either high or low LDL-C may need to be closely monitored for the risk of diabetes .
Description:
Several lines of evidence indicate that quercetin, a polyphenol derived in the diet from fruit and vegetables, contributes to cardiovascular health. We aimed to investigate the effects of dietary quercetin on endothelial function and atherosclerosis in mice fed a high-fat diet. Wild-type C57BL/6 (WT) and apolipoprotein E gene knockout (ApoE−/−) mice were fed: (i) a high-fat diet (HFD) or (ii) a HFD supplemented with 0.05% w/w quercetin (HFD+Q), for 14 weeks. Compared with animals fed HFD, HFD+Q attenuated atherosclerosis in ApoE−/− mice. Treatment with the HFD+Q significantly improved endothelium-dependent relaxation of aortic rings isolated from WT but not ApoE−/− mice and attenuated hypochlorous acid-induced endothelial dysfunction in aortic rings of both WT and ApoE−/− mice. Mechanistic studies revealed that HFD+Q significantly improved plasma F2-isoprostanes, 24h urinary nitrite, and endothelial nitric oxide synthase activity, and increased heme oxygenase-1 (HO-1) protein expression in the aortas of both WT and ApoE−/− mice (P<0.05). HFD+Q also resulted in small changes in plasma cholesterol (P<0.05 in WT) and plasma triacylglycerols (P<0.05 in ApoE −/−mice). In a separate experiment, quercetin did not protect against hypochlorite-induced endothelial dysfunction in arteries obtained from heterozygous HO-1 gene knockout mice with low expression of HO-1 protein. Quercetin protects mice fed a HFD against oxidant-induced endothelial dysfunction and ApoE−/− mice against atherosclerosis. These effects are associated with improvements in nitric oxide bioavailability and are critically related to arterial induction of HO-1. •Quercetin is an important dietary flavonoid that can reduce atherosclerosis in animal models.•Dietary quercetin improved oxidant-induced vascular dysfunction in mice.•Quercetin reduced oxidative stress, increased endothelial eNOS activity, and increased heme oxygenase-1 protein expression in aortic tissue.•Protection against oxidant-induced vascular dysfunction was critically related to arterial induction of HO-1 expression.
Description:
The rupture of an abdominal aortic aneurysm (AAA) causes about 200,000 deaths worldwide each year. However, there are currently no effective drug therapies to prevent AAA formation or, when present, to decrease progression and rupture, highlighting an urgent need for more research in this field. Increased vascular inflammation and enhanced apoptosis of vascular smooth muscle cells (VSMCs) are implicated in AAA formation. Here, we investigated whether hydralazine, which has anti-inflammatory and anti-apoptotic properties, inhibited AAA formation and pathological hallmarks. In cultured VSMCs, hydralazine (100
Description:
The rupture of an abdominal aortic aneurysm (AAA) causes about 200,000 deaths worldwide each year. However, there are currently no effective drug therapies to prevent AAA formation or, when present, to decrease progression and rupture, highlighting an urgent need for more research in this field. Increased vascular inflammation and enhanced apoptosis of vascular smooth muscle cells (VSMCs) are implicated in AAA formation. Here, we investigated whether hydralazine, which has anti-inflammatory and anti-apoptotic properties, inhibited AAA formation and pathological hallmarks. In cultured VSMCs, hydralazine (100
Establishment of sex difference in circulating uric acid is associated with higher testosterone and lower sex hormone-binding globulin in adolescent boys
Description:
Background and Aims: Both fasting and non-fasting levels of triglyceride have been shown positively associated with all-cause mortality. It is unknown whether fasting status modifies this association. This study aimed to address this question. Methods: This study included 34,512 US adults (27,036 fasting and 7476 nonfasting participants). All-cause mortality was ascertained by linkage to the National Death Index records. Cox proportional hazards models were used to estimate hazard ratios of triglyceride for mortality. Results: This cohort was followed up for a mean of 13.0 years. During the follow-up, 8491 all-cause deaths were recorded. A 1-natural-log-unit increase in triglyceride was associated with an 8% higher multivariate-adjusted risk of all-cause mortality. Interaction analyses showed that fasting status interacted with triglyceride in predicting all-cause mortality. Sub-analyses showed that a 1-natural-log-unit increase in triglyceride was associated with a 17% higher multivariate-adjusted risk of all-cause mortality in the nonfasting subcohort; however, there lacked such an association in the fasting sub-cohort. Similarly, high (200–499 mg/dL) and very high levels of triglyceride (
Description:
Background and Aims: Both fasting and non-fasting levels of triglyceride have been shown positively associated with all-cause mortality. It is unknown whether fasting status modifies this association. This study aimed to address this question. Methods: This study included 34,512 US adults (27,036 fasting and 7476 nonfasting participants). All-cause mortality was ascertained by linkage to the National Death Index records. Cox proportional hazards models were used to estimate hazard ratios of triglyceride for mortality. Results: This cohort was followed up for a mean of 13.0 years. During the follow-up, 8491 all-cause deaths were recorded. A 1-natural-log-unit increase in triglyceride was associated with an 8% higher multivariate-adjusted risk of all-cause mortality. Interaction analyses showed that fasting status interacted with triglyceride in predicting all-cause mortality. Sub-analyses showed that a 1-natural-log-unit increase in triglyceride was associated with a 17% higher multivariate-adjusted risk of all-cause mortality in the nonfasting subcohort; however, there lacked such an association in the fasting sub-cohort. Similarly, high (200–499 mg/dL) and very high levels of triglyceride (