Cytotoxic activities of trichothecenes isolated from an endophytic fungus belonging to order hypocreales

Bioassay-guided fractionation of the extract of the endophytic fungus KLAR 5 belonging to order Hypocreales, which was isolated from the twig of Knema laurina (Blume) Warb., resulted in the isolation of brefeldin A (1), 8-deoxy-trichothecin (2), trichothecolone (3), 7alpha-hydroxytrichodermol (4), and 7alpha-hydroxyscirpene (5). Compound 5 was isolated from natural source for the first time. Compound 1 was very highly active against human epidermoid carcinoma of the mouth, human breast cancer (BC-1), and human small cell lung cancer (NCI-H187) cells whereas compounds 2 and 4 were selectively active against BC-1 and NCI-H187 cells. Compounds 3 and 5 were moderately active against these three cancer cell lines.     

Cytotoxic constituents fromSolidago virga-aurea var.gigantea MIQ

Activity-guided fractionation of the whole plant of Solidago virga-aurea var. gigantea M(IQ). (Compositae) has led to the isolation of three cytotoxic compounds, erythrodiol-3-acetate (1), alpha-tocopherol-quinone (2), and trans-phytol (3) from the hexane soluble fraction. It is the first report of those compounds from the genus.     

Inhibitory activity of nitric oxide production in RAW 264.7 cells of daldinals A–C from the fungus Daldinia childiae and other metabolites isolated from inedible mushrooms

Three benzophenone derivatives, daldinals A–C, from the Japanese fungus Daldinia childiae were evaluated for their inhibitory activity of nitric oxide (NO) production stimulated by lipopolysaccharide (LPS) in RAW 264.7 cells. They strongly suppressed the LPS-induced production of NO with IC₅₀ values of 15.2, 4.6 and 6.4 μM, respectively. To clarify the mechanism involved, total RNA extraction, followed by reverse-transcribed, polymerase chain reaction (PCR) for inducible nitric oxide synthase (iNOS) mRNA and finally electrophoresis on agarose gel were performed. These experimental results suggest that the inhibition of the LPS-induced NO production of daldinal B is due to the inhibition of iNOS mRNA synthesis. Further, their biological activities were compared with those of other metabolites obtained during our studies on biologically active substances from inedible fungi in recent years.     

Cytotoxic activities of diarylheptanoids from Alnus japonica

The diarylheptanoids (1–10) 1,7-bis-(3,4-dihydroxyphenyl)-heptane-3-O-β-D-glucopyranosyl(1→3)-β-D-xylopyranoside (1), 1,7-bis-(3,4-dihydroxyphenyl)-heptane-3-O-β-D-apiofuranosyl(1→6)-β-D-glucopyranoside (2), 1,7-bis-(3,4-dihydroxyphenyl)-heptane-5-O-β-D-glucopyranoside (3), 1,7-bis-(3,4-dihydroxyphenyl)-5-hydroxyheptane (4), 1,7-bis-(3,4-dihydroxyphenyl)-heptane-3-one-5-O-β-D-glucopyranoside (5), oregonin (6), hirsutanonol (7), hirsutenone (8), 1,7-bis-(3,4-dihydroxyphenyl)-5-hydroxyheptane-3-O-β-D-xylopyranoside (9), and platyphylloside (10), isolated from the bark of Alnus japonica, were analyzed for their cytotoxic activities on various human and mouse cancer cell lines. The cytotoxic activities of these ten compounds were evaluated against murine B16 melanoma, human SNU-1 gastric cancer, human SNU-354 hepatoma cancer and human SNU-C4 colorectal cell lines. The diarylheptanoids showed potent cytotoxic activities against murine B16 melanoma cells and human SNU-C1 gastric cancer cell when the cell viability was analyzed by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide) assay.     

In vitro anticomplementary activity of hederagenin saponins isolated from roots ofDipsacus asper

Anticomplementary activity of hederagenin and related saponins isolated from Dipsacus asper was investigated in vitro. HN saponin F (3) was most potent with IC50 value of 3.7x10(-5) M followed by 3-O-beta-D-glucopyranosyl-(1->3)-alpha-L-rhamnopyranosyl-(1->2)-beta-L-+ ++arabi nopyranosyl hederagenin 28-O-beta-D-glucopyranosyl-(1->6)-beta-D-glucopyrano side (8), 3-O-beta-L-arabinopyranosyl hederagenin 28-O-beta-D-glucopyranosyl-(1->6)-beta-D-glucopyranoside (5), dipsacus saponin A (4), and hederagenin (1) on the classical pathway (CP) of complement system, while the saponins 3-5 did not show the inhibition of hemolysis and rather increase the hemolysis on the alternative pathway (AP). However, all of C-3 monodesmosides [prosapogenin CP (2), dipsacus saponin B (6), and dipsacus saponin C (7)] evoked hemolysis directly on the erythrocytes.     

A phenolic glucoside isolated fromPrunus serrulata var.spontanea and its peroxynitrite scavenging activity

A new phenolic glucoside (1), pursargentoside, was isolated from the leaves of Prunus serrulata var. spontanea, along with three other known compounds, orobol 7-omicron-glucoside (2), 1beta, 2alpha, 3alpha, 24-tetrahydroxy-urs-12-en-28-oic acid (3), and chlorogenic acid (4). The structure of pursargentoside (1) was identified by spectroscopic data analysis including 1D and 2D NMR spectroscopy, as 2-omicron-beta-(6'-benzoyl)-glucopyranosyl omicron-(Z)-coumaric acid. Compounds 1, 2, and 4 exhibited ONOO scavenging activity, whereas compound 3 was determined to be virtually inactive.     

Cytotoxic evaluation of Melia azedarach in comparison with,Azadirachta indica and its phytochemical investigation

Background

Melia azedarach L. is an important medicinal plant that is used for variety of ailments in Iranian traditional medicine. Azadirachta indica A. Juss is its allied species and possesses similar properties and effects. The present study was undertaken to investigate anticancer activity of these M. azedarach in comparison with A. indica on cancer cell lines and also to evaluate their safety in humans by testing them on normal cell line. The study also aimed to determine the active components that are responsible for medicinal effects of M. azedarach in traditional usages.

Methods

In this study, the cytotoxic activity of crude extracts from M. azedarach and A. indica leaves, pulps and seeds as well as three main fractions of their leaf extracts were assayed against HT-29, A-549, MCF-7 and HepG-2 and MDBK cell lines. MTT assay was used to evaluate their cytotoxic activities. Methanol leaf fraction of M. azedarach as the safest leaf fraction in terms of cytotoxicity was subjected for phytochemical study.

Results

Results of the present study indicated that seed kernel extract of M. azedarach had the highest cytotoxic activity and selectivity to cancer cell lines (IC₅₀ range of 8.18- 60.10 μg mL^-1). In contrast to crude seed extract of A. indica, crude pulp and crude leaf extracts of this plant showed remarkably stronger anti-prolifrative activity (IC₅₀ ranges of 83.45 - 212.16 μg mL^-1 and 34.11- 95.51 μg mL^-1 respectively) than those of M. azedarach (all IC₅₀ values of both plants > 650 μg mL^-1). The phytochemical analysis led to the isolation of four flavonol 3-O-glycosides including rutin, kaempferol-3-O-robinobioside, kaempferol-3-O-rutinoside and isoquercetin along with a purin nucleoside, β-adenosine.

Conclusions

The anti-prolifrative potentials of extracts from different parts of M. azedarach and A. indica were determined. By comparison, methanol leaf fraction of M. azedarach seems to be safer in terms of cytotoxicity. Our study shows that flavonols are abundant in the leaves of M. azedarach and these compounds seem to be responsible for many of medicinal effects exploited in the traditional uses.     

Cytotoxic and antimitotic activities in aqueous extracts of eight cyanobacterial strains isolated from the marine sponge Petrosia ficiformis

Marine cyanobacteria are photosynthetic prokaryotes of significant ecological interest, living free or in association with invertebrates. They are also considered as excellent sources of antineoplastic, antibacterial, antiviral and antifungal compounds. In this work, aqueous extracts from eight cyanobacterial strains isolated from the Mediterranean sponge Petrosia ficiformis have been investigated for their bioactive properties. Bioassays with human erythrocytes, Artemia salina nauplii, and Paracentrotus lividus gametes and embryos were performed. Some aqueous extracts exhibited citolytic effect on human erythrocytes and toxic activity against A. salina nauplii. Furthermore antimitotic activity was evidenced during sea urchin embryos development and disorganization of blastomeres with altered cell-cell contact was also induced. Some of the isolated cyanobacterial strains, belonging to Leptolyngbya and Synechococcus genera with an high citotoxic activity, should be further investigated to better characterize their bioactive molecules. Our data confirm cyanobacteria as an interesting source of novel bioactive compounds with potential applications in pharmaceutics.     

Bioactive Terpenoids and Flavonoids from Daucus littoralis Smith subsp. hyrcanicus Rech.f,an Endemic Species of Iran

Background

Daucus littoralis Smith subsp. hyrcanicus Rech.f. (Apiaceae) is an endemic species in northern parts of Iran where it is commonly named Caspian carrot. The fruits have been used as condiment.

Methods

In a series of in vitro assays, antioxidant (DPPH and FRAP assays), cytotoxic and antimicrobial activities of different extracts of roots and fruits were evaluated for the first time. The separation and purification of the compounds were carried out on the most potent extracts using various chromatographic methods and identified by spectroscopic data (¹H and ¹³C NMR).

Results

The results showed that among the extracts only fruit methanol extract (FME) has significant antioxidant activity (IC₅₀ = 145.93 μg.ml^-1 in DPPH assay and 358 ± 0.02 mmol FeII/g dry extract in FRAP assay). The radical scavenging activity of FME at 400 μg.ml^-1 was comparable with α-tocopherol (40 μg.ml^-1) and with BHA (100 μg.ml^-1) (p > 0.05). FME did not show any toxicity against cancerous and normal cell lines. Fruit ethyl acetate extract (FEE) had cytotoxic activity against breast carcinoma and hepatocellular carcinoma cells (IC₅₀ 168.4 and 185 μg.ml^-1, respectively), while it did not possess antioxidant activity in comparison with α-tocopherol and BHA as standard compounds. Ethyl acetate and methanol extract of fruits showed antimicrobial activity against Staphylococcus aureus (MIC: 3.75 mg.ml^-1) and Candida albicans (MIC: 15.6 and 7.8 mg.ml^-1, respectively). Four terpenoids were isolated form FEE including: β-sitosterol (1), stigmasterol (2), caryophyllene oxide (3), β-amyrin (4). Also, three flavonoids namely quercetin 3-O-β-glucoside (5), quercetin 3-O-β-galactoside (6) and luteolin (7) were isolated from FME.

Conclusion

This study showed that FEE and FME of D. littoralis Smith subsp. hyrcanicus Rech.f. had the highest biological activities which may be correlated with in vitro cytotoxic, antimicrobial and antioxidant activities of terpenoids and flavonoids components of the extracts.     

Mode of antiviral activity of water soluble components isolated fromElfvingia applanata on vesicular stomatitis virus

A preparation of water soluble components (EA) was made from carpophores of Elfvingia applanata (Pers.) Karst and its in vitro antiviral activity on vesicular stomatitis virus [(Indiana serotype, VSV(IND)] was investigated by plaque reduction assay. EA exhibited potent antiviral activity on VSV(IND) growth and negligible cytotoxicity on Vero cells, 50% effective concentration (EC50) of 104 microg/ml and 50% cytotoxic concentration (CC50) of 3,793 microg/ml, respectively. Selectivity index (SI, CC50/EC50) of EA on Vero cell and VSV(IND) was about 36.5. EA did not display either a direct virucidal effect on VSV(IND) or induction of antiviral substance by Vero cells upon its treatment. Thus, the mode of antiviral activity of EA was studied at steps of viral adsorption onto cell. When both EA and virus were added to cell monolayers, titer of cell-free virus in culture supernatant increased in ca. 30-40% compared with that of control group and titer of cell-associated virus was 60-100% higher than that of control group. These results suggested that antiviral activity of EA on VSV(IND) might be due to the hindrance of viral entry to cells at either endocytosis or loss of envelope.     

Calcium-antagonizing activity of S-petasin, a hypotensive sesquiterpene from Petasites formosanus, on inotropic and chronotropic responses in isolated rat atria and cardiac myocytes

Petasites formosanus, an indigenous species of Petasites, has been used to treat cardiovascular diseases such as hypertension for years. We have suggested recently that S-petasin, a major sesquiterpene from P. formosanus, inhibits vascular smooth muscle contraction through inhibition of voltage-dependent Ca²⁺ channels, a phenomenon possibly responsible for the hypotensive effect of P. formosanus. This study was designed to examine the chronotropic and inotropic actions of S-petasin in the heart in vivo and in vitro. Administration of S-petasin (0.1–1.5 mg/kg i.v.) in anesthetized rats reduced heart rate dose-dependently. This response was consistent with significant suppression of both contractile amplitude and spontaneous firing rate of isolated atria, responses that were not antagonized by atropine (1 µM). Mechanical evaluation in isolated ventricular myocytes showed that S-petasin (0.1 to 100 µM) depressed peak myocyte contraction and intracellular Ca²⁺ transients concentration-dependently. The duration of myocyte contraction was not affected. Whole-cell voltage clamp analysis revealed that S-petasin inhibited the L-type Ca²⁺ current (I_Ca,L) concentration-dependently and shifted the steady-state inactivation curve of I_Ca,L to more negative potentials. However, a receptor-binding assay failed to identify any significant interaction between S-petasin (0.1–300 µM) and the dihydropyridine binding sites of L-type voltage-dependent Ca²⁺ channels. Taken together, these data show that the negative chronotropic and inotropic properties of S-petasin that can be ascribed mainly to I_Ca,L inhibition, but not to blockade of dihydropyridine binding sites of L-type Ca²⁺ channel or to muscarinic receptor activation.     

Cytotoxic peroxides fromArtemisia stolonifera

Two sesquiterpene endoperoxides, 1S, 4R, 6R-1, 4-endoperoxy-bisabola-2, 10-diene (I), 1R, 4S, 6R-1, 4-endoperoxy-bisabola-2, 10-diene (II), and a sesquiterpene hydroperoxide, 1beta-hydroperoxygermacra-4 (15), 5, 10 (14)-triene (III) were isolated from the aerial parts of Artemisia stolonifera (Compositae). Their chemical structures were assigned by spectral evidences. Compounds I and II exhibited cytotoxicity against five human tumor cell lines with their ED50 values ranging from 0.20 to 5.43 microg/ml and from <0.1 to 0.87 microg/ml, respectively.     

白豆中α-淀粉酶抑制剂的分离及其活性研究

采用乙醇分级沉淀、CMII纤维素离子交换柱色谱及凝胶柱色谱，从白豆中纯化得到一种α- 淀粉酶抑制剂(α-AI)，经SDS-PAGE及Sepharose CL-6B柱色谱鉴定其为结构均一的糖蛋白。该糖蛋白中蛋白质含量为88.2%，氨基酸组成主要为天冬氨酸、谷氨酸、苏氨酸及丝氨酸。糖链部分单糖组成为甘露糖、葡萄糖、半乳糖和木糖，其摩尔比为2.42∶1.50∶1.52∶1.00。糖和蛋白质结合的糖肽键类型为O-糖肽键。白豆α-AI使用剂量为150 mg·kg^-1体重，连续使用7 d时，α-AI可明显降低高血糖大鼠的空腹血糖；使用剂量为300 mg·kg^-1时，对高血糖大鼠的糖耐量具有明显的改善作用。研究结果表明，从白豆中分离得到的淀粉酶抑制剂对高血糖大鼠具有明显的降血糖功能，其作为一种安全、天然的降糖药物具有良好的开发前景。     

民族药四数九里香药效部位筛选及咔唑生物碱含量测定研究

目的 筛选民族药四数九里香抑制肿瘤细胞增殖作用的药效部位及测定质量标准控制物含量。方法 采用MTS法筛选萃取馏分及单体化合物的细胞毒活性,利用高效液相色谱法测定质量标准物的含量。结果 在初试浓度为100 μg•mL^-1时,5个馏分中乙酸乙酯萃取馏分对5株肿瘤细胞（HL-60、A-549、SMMC-7721、MCF-7、SW-480）的抑制率（%）最大,故确定其为民族药四数九里香的有效部位,IC₅₀（μg•mL^-1）依次为 52.84 ± 1.63、78.34 ± 4.79、43.29 ± 1.74、64.19 ± 1.24、43.75 ± 2.20;从药效部位分离和鉴定7个咔唑生物碱化合物,并研究了这些化合物的细胞毒活性,分析得化合物7的抑制作用最大,其抑制肿瘤细胞HL-60、A-549、MCF-7、SW-480的IC50(μmol•L-1)值依次为 15.11 ± 1.14、18.11 ± 1.11、24.13 ± 1.32、31.12 ± 1.12;选择咔唑生物碱mahanimbine（化合物7）作为民族药四数九里香的质量标准控制物,分析得到化合物7在四数九里香中的含量为61.08 μg•g^-1。结论 乙酸乙酯萃取部位为民族药四数九里香醇提物抑制肿瘤细胞增殖的活性部位、以从中分离鉴定的咔唑化合物7为质量控制标准物,采用高效液相色谱仪建立快速分析四数九里香中mahanimbine测定分析方法,本法对样品前期处理要求不高,设备条件简便,结果可靠,为民族药四数九里香抑制肿瘤细胞增殖作用质量标准建立提供理论支撑。     

PPARγ激动剂的设计、合成及其胰岛素增敏活性

目的寻找新的高效、低毒的PPARγ激动剂。方法以JTT-501和JTT-20993为先导化合物，设计并合成新的丙二酸类和异恶唑类化合物，并测定其胰岛素增敏活性。结果共合成了8个新化合物,用核磁共振、质谱和红外光谱进行结构确证，并用胰岛素筛选模型初步评价了这些化合物的胰岛素增敏活性。化合物1A-4A显示胰岛素增敏活性,其中化合物1A和3A有较强活性。结论化合物1A和3A值得进一步评价。     

CARDIOVASCULAR ACTIVITY OF 14-DEOXY-11, 12-DIDEHYDROANDROGRAPHOLIDE IN THE ANAESTHETISED RAT AND ISOLATED RIGHT ATRIA

The cardiovascular activity of 14-deoxy-11,12-didehydroandrographolide (DDA) fromAndrographis paniculata(Burm.f.) Nees (Acanthaceae) was elucidated in anaesthetised Sprague–Dawley (SD) rats and isolated rat right atria. In anaesthetised rats, DDA produced significant falls in mean arterial blood pressure (MAP) and heart rate in a dose-dependent manner with the maximum decrease of 37.6±2.6% and 18.1±4.8%, respectively. The ED₅₀value for MAP was 3.43 mmol kg⁻¹. Pharmacological antagonist studies were done using this dose. The hypotensive action of DDA was not mediated through effects on the α-adrenoceptor, muscarinic cholinergic and histaminergic receptors, for it was not affected by phentolamine, atropine as well as pyrilamine and cimetidine. However, it seems to work via adrenoceptors, autonomic ganglia receptor and angiotensin-converting enzyme, since the hypotensive effect of DDA was negated or attenuated in the presence of propranolol, hexamethonium and captopril. In the isolated right atria, DDA caused negative chronotropic action and antagonised isoproterenol-induced positive chronotropic actions in a non-competitive and dose-dependent manner. These results further supported the bradycardia-inducing and β-adrenoceptor antagonistic properties of DDAin vivo.     

Cytotoxic proteins of Amanita virosa Secr. mushroom: Purification, characteristics and action towards mammalian cells

A new hemolytic lectin was purified from the fruit bodies of Amanita virosa Secr. mushroom by the affinity chromatography on the cross-linked ovomucin. This lectin destroyed erythrocytes of human and animals of various species, and its hemolytic activity decreased in the row: rabbit > rat > human > dog. The erythrocytes of sheep, cow and carp were resistant to such hemolytic action of the lectin (1 mg/mL). The lectin-mediated hemolysis was blocked by the polyethylene glycol with molecular mass over 1350. A. virosa lectin, unlike Amanita phalloides lectin, did not interact with tested monosaccharides. However, the 4-nitrophenyl derivates of the monosaccharides inhibited the action of A. virosa lectin which did not prefer targeting O-type glycoproteins over the N-type glycoproteins. Murine leukemia cells of L1210 line and human leukemia T-cells of CEM T4 and Jurkat lines were shown to be sensitive to toxic effect of the lectin and another protein toxovirin isolated from A. virosa fruit bodies It was found that toxovirin possessed an enzymatic activity of l-amino acid oxidase. Since both toxic proteins - the lectin and toxovirin - are sensitive to an elevated temperature, it is suggested that they play a significant role in human poisoning only when the unbaked mushroom is eaten.     

Soraphen A, an inhibitor of acetyl CoA carboxylase activity, interferes with fatty acid elongation

Acetyl CoA carboxylase (ACC1 and ACC2) generates malonyl CoA, a substrate for de novo lipogenesis (DNL) and an inhibitor of mitochondrial fatty acid β-oxidation (FAO). Malonyl CoA is also a substrate for microsomal fatty acid elongation, an important pathway for saturated (SFA), mono- (MUFA) and polyunsaturated fatty acid (PUFA) synthesis. Despite the interest in ACC as a target for obesity and cancer therapy, little attention has been given to the role ACC plays in long chain fatty acid synthesis. This report examines the effect of pharmacological inhibition of ACC on DNL and palmitate (16:0) and linoleate (18:2, n − 6) metabolism in HepG2 and LnCap cells. The ACC inhibitor, soraphen A, lowers cellular malonyl CoA, attenuates DNL and the formation of fatty acid elongation products derived from exogenous fatty acids, i.e., 16:0 and 18:2, n − 6; IC₅₀ ∼ 5 nM. Elevated expression of fatty acid elongases (Elovl5, Elovl6) or desaturases (FADS1, FADS2) failed to override the soraphen A effect on SFA, MUFA or PUFA synthesis. Inhibition of fatty acid elongation leads to the accumulation of 16- and 18-carbon unsaturated fatty acids derived from 16:0 and 18:2, n − 6, respectively. Pharmacological inhibition of ACC activity will not only attenuate DNL and induce FAO, but will also attenuate the synthesis of very long chain saturated, mono- and polyunsaturated fatty acids.     

Analgesic activity of myricetin isolated from <Emphasis Type="Italic">Myrica rubra Sieb. et Zucc.</Emphasis> leaves

Myrica rubra Sieb. et Zucc. leaves are commonly used as an astringent, antidiarrheic, and analgesics in folk medicine in China. In the present study, the analgesic activity of myricetin, a major compound in Myrica rubra Sieb. et Zucc. leaves was evaluated in vivo. The analgesic effect of myricetin was tested by a serial of models, such as acetic acid-induced writhing response, formalin-induced paw licking and hot plate test. The sedative activity was evaluated by pentobarbital-induced sleep time. Platelet aggregation induced by collagen and arachidonic acid was also performed in vitro. Myricetin showed a significant inhibition on chemical nociceptive models such as the acetic acid-induced writhing response and the licking time on the late phase in the formalin test in a dose-dependent manner, but did not manifest a signicant effect in hot plate test. Myricetin was also not able to increase the sleeping time induced by pentobarbital, which further indicated that the analgesic effect of myricetin was unrelated to sedation. In addition, myricetin inhibited the content of PGE2 in the peritoneal fluid and platelet aggregation induced by collagen and arachidonic acid in vitro. These results collectively demonstrated that myricetin possessed potent analgesic activity, which was related with peripheral analgesia, but, not with the opioid system. Myricetin may be a potent COX-1 inhibitor with anti-platelet activity.     

Endothelial cell-specific aryl hydrocarbon receptor knockout mice exhibit hypotension mediated, in part, by an attenuated angiotensin II responsiveness

Hypotension in aryl hydrocarbon receptor knockout mice (ahr^−/−) is mediated, in part, by a reduced contribution of angiotensin (Ang) II to basal blood pressure (BP). Since AHR is highly expressed in endothelial cells (EC), we hypothesized that EC-specific ahr^−/− (ECahr^−/−) mice would exhibit a similar phenotype. We generated ECahr^−/− mice by crossing AHR floxed mice (ahr^fx/fx) to mice expressing Cre recombinase driven by an EC-specific promoter. BP was assessed by radiotelemetry prior to and following an acute injection of Ang II or chronic treatment with an angiotensin converting enzyme inhibitor (ACEi). ECahr^−/− mice were hypotensive (ECahr^+/+: 116.1 ± 1.4; ECahr^−/−: 107.4 ± 2.0 mmHg, n = 11, p < 0.05) and exhibited significantly different responses to Ang II and ACEi. While Ang II increased BP in both genotypes, the increase was sustained in ECahr^+/+, whereas the increase in ECahr^−/− mice steadily declined. Area under the curve analysis showed that Ang II-induced increase in diastolic BP (DBP) over 30 min was significantly lower in ECahr^−/− mice (ECahr^+/+ 1297 ± 223 mmHg/30 min; ECahr^−/−_AUC: 504 ± 138 mmHg/30 min, p < 0.05). In contrast, while ACEi decreased BP in both genotypes, the subsequent rise in DBP after treatment was significantly delayed in the ECahr^−/− mice. ECahr^−/− mice also exhibited reduced vascular and adipose Ang II type 1 receptor (AT1R) expression, and reduced aortic Ang II-dependent vasoconstriction in the presence of vascular adipose. Taken together these data suggest that hypotension in ECahr^−/− mice results from reduced vascular responsiveness to Ang II that is influenced by AT1R expression and adipose.