Simultaneous targeting of VEGF-receptors 2 and 3 with immunoliposomes enhances therapeutic efficacy

Background: Tumor progression depends on angiogenesis. Vascular endothelial growth factor (VEGF) receptors (VEGFRs) are the main signal transducers that stimulate endothelial cell migration and vessel sprouting. At present, only VEGFR2 is targeted in the clinical practice.

Purpose: To develop new, anti-angiogenic nanoparticles (immunoliposomes, ILs), that redirect cytotoxic compounds to tumor-associated vascular cells.

Methods: Pegylated liposomal doxorubicin (PLD) was targeted against VEGFR2- and VEGFR3-expressing cells by inserting anti-VEGFR2 and/or anti-VEGFR3 antibody fragments into the lipid bilayer membrane of PLD. These constructs were tested in vitro, and in vivo in the Rip1Tag2 mouse model of human cancer.

Results: The combination treatment with anti-VEGFR2-ILs-dox and anti-VEGFR3-ILs-dox was superior to targeting only VEGFR2 cells and provides a highly efficient approach of depleting tumor-associated vasculature. This leads to tumor starvation and pronounced reduction of tumor burden.

Conclusion: Nanoparticles against VEGFR2 and ?3 expressing tumor-associated endothelial cells represent a promising and novel anti-cancer strategy.     

Effects of rosiglitazone added to submaximal doses of metformin compared with dose escalation of metformin in type 2 diabetes: the EMPIRE Study

ABSTRACT

Objective: This study was designed to compare the efficacy, safety and tolerability of rosiglitazone (RSG) added to submaximal doses of metformin (MET) with dose escalation to the maximal effective dose of MET monotherapy in type 2 diabetes mellitus.

Research design and methods: In this multi-center, double-blind, randomized, parallel-group study, 766 subjects with a baseline MET dose of 1000?mg/day were randomized to receive either RSG 4?mg/day (4?mg/1000?mg) or MET 500?mg/day (1500?mg/day total dose) for 8 weeks. Only the RSG dose was increased in the combination group – to 8?mg/day (8?mg/1000?mg) – and only the MET dose was increased in the MET monotherapy group – to 2000?mg/day for the remaining 16 weeks.

Results: After 24 weeks, RSG added to MET (8?mg/1000?mg/day) was at least as effective as 2000?mg/day of MET in improving HbA_1c’, with mean reductions of –0.93% (95% CI: –1.06%, –0.80%) and –0.71% (95% CI: –0.83%, –0.60%), respectively, from baseline in subjects that completed the study according to the investigator (mean treatment effect/difference of –0.20% [95% CI: –0.36%,–0.04%]). In addition, a higher percentage of subjects in the RSG + MET group achieved American Diabetes Association target levels of HbA_1c < 7% (58.1% versus 48.4%) and American Association of Clinical Endocrinologists target levels of HbA_1c ≤ 6.5% (40.9% versus 28.2%). This combination provided significantly greater reductions from baseline in fasting plasma glucose (FPG; –2.29?mmol/L [± 2.37?mmol/L] and –1.12?mmol/L [± 2.41?mmol/L], respectively), with a treatment difference of –0.85?mmol/L (95% Cl: –1.23?mmol/L, –0.47?mmol/L). For the intent-to-treat (ITT) population, the percentage of subjects experiencing a gastrointestinal side-effect was 27.9% and 38.7% for the RSG + MET and MET groups, respectively (OR = 1.63, 95% CI: 1.19, 2.24). Mean body weight (± SD) increased in all randomized subjects treated with the combination therapy (+ 1.79 ± 4.15?kg) compared with a mean weight loss in the up-titrated MET group (–1.78 ± 3.50?kg).

Conclusions: This study suggests that addition of RSG to submaximal doses of MET may be a suitable alternative to the maximal effective dose of MET monotherapy.     

Efficacy and safety of the single pill combination of amlodipine 10 mg plus valsartan 160 mg in hypertensive patients not controlled by amlodipine 10 mg plus olmesartan 20 mg in free combination

ABSTRACT

Background and objective: For patients with moderate hypertension (grade 2, defined as systolic blood pressure [SBP] 160–179?mmHg and/or diastolic blood pressure [DBP] 100–109?mmHg), current guidelines recommend initial combination therapy and rapid dose-adjustment to achieve blood pressure goal. In this study we investigated the efficacy and tolerability of the single pill combination of amlodipine 10?mg plus valsartan 160?mg (A?10?+?Val 160) in patients not controlled by the free combination of amlodipine 10?mg plus olmesartan 20?mg (A?10?+?O 20).

Methods: In this prospective, open-label, non-randomized trial, 257 patients with mean sitting DBP of 100–109?mmHg at trough entered a 4 week treatment phase with A?10?+?O 20 in free combination once daily. Patients in whom DBP remained uncontrolled were switched in a second 4 week treatment phase to A?10?+?Val 160. The primary efficacy variable was the reduction in DBP at week 8 compared to week 4 in the intent-to-treat population.

Results: In the total cohort, baseline SBP/DBP of 164.2?±?9.8/103.6?±?2.1?mmHg decreased by 19.2?±?12.4/14.1?±?7.4?mmHg at week 4. In patients who did not achieve BP control (n?=?175), subsequent treatment with A?10?+?Val 160 for 4 weeks reduced SBP from 149.6?±?11.1 at week 4 by 7.9?mmHg at week 8 (95% CI: 6.1–9.6, p?<?0.0001) and DBP from 93.4?±?3.9?mmHg by 9.1?mmHg (95% confidence interval: 8.1–10.2, p?<?0.0001). The combination of A?10?+?Val 160 was well tolerated, and the observed adverse events (15.3% of patients in phase 2) were consistent with the known drug profiles.

Conclusions: In a study designed to reflect typical clinical practice, in patients not controlled by the free combination of A?10?+?O 20, the single pill combination of A?10?+?Val 160 produced a statistically and clinically significant additional BP reduction and was well tolerated. Potential limitations of the design (open-label, non-controlled design, short term treatment) have to be taken into account.     

The effects of Melissa officinalis (lemon balm) pretreatment on the resistance of the heart to myocardial injury

Context: The antihyperlipidemic, antiarrhythmic, neuroprotective and hepatoprotective effects of Melissa officinalis L. (Lamiaceae) have been reported. However, no study has examined its effects on the resistance of the heart to stressful conditions.

Objective: The objective of this study is to evaluate the effects of aqueous extract of M. officinalis aerial parts on Wistar rat heart with/without cardiac injury.

Materials and methods: Animals were grouped as control, isoproterenol (ISO), M. officinalis without (M50, M100, and M200) and with isoproterenol (M50?+?ISO, M100?+?ISO, and M200?+?ISO). The aqueous extract of M. officinalis was orally administered at dosages of 50, 100, and 200?mg/kg/d, respectively, for 7 consecutive days. On the 6th and 7th day, ISO, M50?+?ISO, M100?+?ISO, and M200?+?ISO groups received 85?mg/kg of isoproterenol for myocardial injury induction. On day 8, hemodynamic parameters were recorded and samplings were done.

Results: The extract (50, 100, and 200?mg/kg) significantly reduced the heart rate (264?±?5, 259?±?5 and 281?±?3 versus 377?±?13 in control group, p?<?0.01). Blood pressure was significantly decreased in M50?+?ISO (75?±?5) versus M50 (110?±?6) and M100?+?ISO (72?±?6) versus M100 (105?±?5?mmHg, p?<?0.01). The malondialdehyde levels of the injured hearts were lower in M50?+?ISO and M100?+?ISO groups than in the ISO group (p?<?0.05). Serum cardiac troponin I was higher in the M200?+?ISO group (5.1?±?1.7) than in the ISO group (2.7?±?0.7?ng/ml, p?<?0.05).

Conclusion: The lower dose of extract, by improving the balance of the redox system and by reducing the heart rate, may increase the heart resistance to injury. However, the higher doses of extract may intensify the injury of ischemic heart.     

Reductions in biomarkers of cardiovascular risk in type 2 diabetes with rosiglitazone added to metformin compared with dose escalation of metformin: an EMPIRE trial sub-study*

ABSTRACT

Objective: To compare the effects of rosiglitazone added to metformin with dose escalation of metformin on cardiovascular risk biomarkers in type 2 diabetes mellitus.

Research design and methods: Cardiovascular biomarkers were assessed in a sub-population of 122 subjects with type 2 diabetes mellitus (mean age 54.6 and 56.0 years, BMI 34.7 and 32.1?kg/m² for the rosiglitazone plus metformin and metformin groups, respectively) from the multicenter (63 centers in the USA), double-blind, randomized parallel-group Escalation of Metformin theraPy vs. Initiation of Rosiglitazone Early (EMPIRE) study. Treatment group sizes were slightly imbalanced owing to central, rather than local, randomization. Subjects receiving metformin 1000?mg/day at baseline were randomized to rosiglitazone 4?mg/day plus metformin 1000?mg/day (RSG + MET) or metformin 1500?mg/day (up-titrated MET) for 24 weeks. At 8‐weeks, rosiglitazone was increased to 8?mg/day in RSG + MET recipients and metformin to 2000?mg/day in up-titrated MET recipients.

Results: Reductions from baseline in HbA_1c at week 24 (mean ± SD) occurred in both groups (RSG + MET: –0.61% ± 1.16%; up-titrated MET: –0.65% ± 1.18%). Post-prandial glucose levels (AUC_0–3h) decreased with RSG + MET (–3.5?mmol/L.h; 95% confidence interval [CI]: –5.2 to –1.8) and up-titrated MET (–1.3?mmol/L.h; 95% CI: –3.8 to 1.1). Homeostasis Model Assessment (HOMA)-estimated insulin sensitivity increased by 37.7% (95% CI: 22.8 to 54.5) in RSG + MET and 6.9% (95% CI: –6.2 to 21.9) in up-titrated MET recipients. RSG + MET reduced C‐reactive protein (CRP; –23.9%; 95% CI: –40.4 to –2.8), plasminogen activator inhibitor‐1 (PAI‐1) activity (–30.1%; 95% CI: –44.5 to –11.9), PAI‐1 antigen (–15.5%; 95% CI: –28.3 to –0.3) and matrix metalloproteinase‐9 (MMP‐9; –13.8%; 95% CI: –25.1 to –0.9), but increased tumor necrosis factor‐α (TNF‐α; 27.0%; 95% CI: 6.8 to 50.9). Corresponding values for up-titrated MET were CRP –9.3% (95% CI: –36.9 to 30.2), PAI‐1 activity –7.2% (95% CI: –28.2 to 20.0), PAI‐1 antigen –1.5% (95% CI: –17.4 to 17.5), MMP‐9 29.0% (95% CI: –1.3 to 68.6) and TNF‐α –6.0% (95% CI: –22.0 to 13.2).

Conclusions: These results suggest that rosiglitazone plus metformin has positive cardiovascular effects against a background of similar glycemic improvements.     

Quercetin and vitamin E attenuate Bonny Light crude oil-induced alterations in testicular apoptosis,stress proteins and steroidogenic acute regulatory protein in Wistar rats

Studies have shown the reproductive effects of Bonny Light crude oil (BLCO) via the mechanism of oxidative stress and testicular apoptosis. We investigated the protective role of quercetin and vitamin E on BLCO-induced testicular apoptosis. Experimental rats were divided into four groups of four each. Animals were orally administered 2?ml/kg corn oil (control: group 1), BLCO-800?mg/kg body weight?+?10?mg/kg quercetin (group 2), BLCO-800?mg/kg body weight?+?50?mg/kg vitamin E (group 3) and BLCO-800?mg/kg body weight only (group 4) for 7 d. Protein levels of caspase 3, FasL, NF-kB, steroidogenic acute regulatory protein and stress response proteins were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Immunofluorescence staining was used to quantify the expression of caspase 3, FasL and NF-kB. Apoptosis was quantified by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. Administration of BLCO resulted in a significant increase in the levels of stress response proteins and apoptosis-related proteins by 50% and above after 7 d following BLCO exposure and a concomitant increase in expression of caspase 3, FasL and NF-kB expression by immunofluorescence staining. Apoptosis showed a significant increase in TUNEL positive cells. Co-administration with quercetin or vitamin E reversed BLCO-induced apoptosis and levels of stress protein, relative to control. These findings suggest that quercetin and vitamin E may confer protection against BLCO-induced testicular oxidative stress-related apoptosis.     

Co-administration of walnut (Juglans regia) prevents systemic hypertension induced by long-term use of dexamethasone: a promising strategy for steroid consumers

Context: The long-term consumption of glucocorticoids (GCs) may induce serious adverse effects such as hypertension. There is sufficient evidence related to the benefit of walnuts on the cardiovascular system.

Objective: This study assesses the effect of methanol extract of walnut [Juglans regia L. (Juglandaceae)] on dexamethasone-induced hypertension and the possible mechanisms in Wistar rats.

Material and methods: Animals were randomized into control, kernel extract (100 and 200?mg/kg/d, orally), dexamethasone (0.03?mg/kg/d, subcutaneously), dexamethasone?+?kernel (100 and 200?mg/kg/d, separately), and dexamethasone?+?captopril (25?mg/kg/d, orally) groups. Animals were treated with water, kernel extract or captopril by gavage 4 d before and during 11 d of saline or dexamethasone treatment. On the 16th day, blood pressure (BP) was recorded and blood samples were collected to measure nitric oxide (NO). Animal hearts were frozen for measurement of malondialdehyde (MDA) and glutathione peroxidase (GPX).

Results: Dexamethasone increased the diastolic BP and MDA/GPX ratio in comparison with control group (128?±?7 vs. 105?±?3?mmHg, p?p?p?p?Conclusion: Similar to captopril, walnut extract normalized dexamethasone-induced hypertension. A part of this beneficial effect apparently involves maintaining balance of the redox system and NO production.     

Hepatotoxicity and nephrotoxicity in rats induced by carbon tetrachloride and the protective effects of Teucrium polium and vitamin C

This work investigated the protective effects of Teucrium polium (T. polium) and vitamin C (Vit C) against carbon tetrachloride (CCl₄) induced hepatotoxicity and nephrotoxicity in rats. T. polium reduced the Fer reduced antioxidant power (FRAP) (IC₅₀?=?0.89?mg/ml) and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) (IC₅₀?=?0.049 µg/ml) than Vit C, FRAP (IC₅₀?=?0.71?mg/ml) and DPPH (IC₅₀?=?0.029 µg/ml). Male albino Wistar rats were divided into six groups: Group I was used as controls, Group II received CCl₄ in olive oil (0.5?ml/kg) by gavage, Group III received CCl₄ in olive oil (0.5?ml/kg) by gavage after 3 d of receiving T. polium (5?g/l), orally, Group IV received T. polium (5?g/l) alone, by gavage, for 7 d, Group V received CCl₄ in olive oil (0.5?ml/kg) by gavage after 3 d of receiving Vit C (250?mg/kg) by gavage and Group VI received Vit C (250?mg/kg) alone by gavage. CCl₄ showed an increase of serum hepatic and renal markers aspartate aminotransferase (AST), alanine aminotransferase (ALT), urea, and creatinine. Moreover, we noted an increase of lipid peroxidations and a decrease in antioxidants enzymes: superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) of CCl₄ rats compared to controls. The pretreatment with (200?mg/kg) of T. polium and with Vit C (250?mg/kg) by gavage, for 7 d, displayed their ability to protect against oxidative damage and biochemical changes induced by CCl₄. Our results were in accordance with histopathological observations.     

Sorafenib Decreases Tumor Exposure to an Anti-carcinoembryonic Antigen Monoclonal Antibody in a Mouse Model of Colorectal Cancer

In this investigation, we test the hypothesis that treatment with sorafenib, an anti-angiogenic agent, decreases tumor vascularization and, consequently, hinders the delivery of monoclonal antibodies (mAb) to xenograft tumors. Severe combined immunodeficiency mice bearing carcinoembryonic antigen (CEA) expressing tumor xenografts were divided into control and sorafenib-treated groups. Sorafenib was administered to the latter group at 50 mg/kg IP every 48 h, starting 4 days post-tumor implantation. When tumors attained a size of 200–300 mm³, mice were evaluated for (a) tumor microvessel density (using immunohistochemical analysis), (b) tumor macromolecular extravasation (using Evans Blue Dye (EBD)), (c) pharmacokinetics of an anti-CEA mAb, T84.66, following an intravenous dose of 10 mg/kg, and (d) intra-tumoral spatial distribution of T84.66 (using autoradiography). Sorafenib treatment resulted in a substantial reduction in tumor growth rate, a visible reduction in tumor microvessel density, and in a 46.4% decrease in EBD extravasation in tumor tissue (p?<?0.0455). For control and treated mice, no significant difference was found for the area under the mAb plasma concentration-time curve (AUC_(0–7d): 1.67?×?10³?±?1.28?×?10² vs. 1.76?×?10³?±?1.75?×?10² nM?×?day, p?=?0.51). However, tumor AUC_(0–7d) was reduced by 40.8% in sorafenib-treated mice relative to that observed in control mice (5.61?×?10²?±?4.27?×?10¹ vs. 9.48?×?10²?±?5.61?×?10¹ nM?×?day, p?<?0.001). Sorafenib therapy was also found to markedly alter mAb tumor spatial distribution. The results collectively suggest that sorafenib treatment causes a significant reduction in mAb delivery to, and distribution within, solid tumors.     

Effect of tesaglitazar,a dual PPARα/γ agonist,on glucose and lipid abnormalities in patients with type 2 diabetes: a 12‐week dose-ranging trial

ABSTRACT

Objective: The Glucose and Lipid Assessment in Diabetes (GLAD) trial examined the dose-response relationship of the dual peroxisome proliferatoractivated receptor (PPAR) α/γ agonist tesaglitazar in type 2 diabetic patients.

Study design: GLAD was a 12‐week, multicenter, international, randomized, parallel-group trial. Five-hundred men and women aged 30–80 years with type 2 diabetes (fasting plasma glucose [FPG] ≥ 126?mg/dL [≥ 7.0?mmol/L]) received once-daily, double-blind placebo or tesaglitazar (0.1?mg, 0.5?mg, 1.0?mg, 2.0?mg, or 3.0?mg) or open-label pioglitazone (45?mg), included as a therapeutic benchmark.

Main outcome measures: Placebo-corrected changes from baseline in FPG (primary end point), plasma lipids, and insulin-resistance measures.

Results: At baseline, the mean patient age was 56.1 years, 57.5 years, and 58.9 years for placebo, across tesaglitazar groups, and for pioglitazone, respectively. For the corresponding groups, mean body mass index was 30.6?kg/m², 30.9?kg/m², and 29.7?kg/m², and mean HbA_1c was 7.0%, 7.2%, and 7.0%, respectively. At 12 weeks, tesaglitazar 0.5?mg, 1.0?mg, 2.0?mg, and 3.0?mg produced statistically significant reductions in FPG (–30.3?mg/dL, –41.1?mg/dL, –55.0?mg/dL, –60.9?mg/dL; p < 0.0001), triglycerides (–17.2%, –32.9%, –41.0%, –40.9%; p < 0.01), and apolipoprotein B (–15.0%, –15.7%, –21.0%, –22.3%, respectively; p < 0.0001). Tesaglitazar at doses ≥ 1.0?mg significantly increased high-density lipoprotein-cholesterol (HDL‐C) (15.0%, 13.0%, 12.9%; p < 0.001), and reduced non-HDL‐C (–13.2%, –22.2%, –25.0%; p < 0.0001), very-low-density lipoprotein-cholesterol (VLDL‐C) (–36.9%, –49.8%, –52.5%; p < 0.0001), and total cholesterol (–6.8%, –14.1%, –15.5%, respectively; p < 0.01). Tesaglitazar ≥ 0.5?mg improved insulin-resistance measures. Although no formal statistical analyses were performed between active treatments, improvements in efficacy measures with tesaglitazar 1.0?mg were numerically similar to or greater than those with pioglitazone. Similar numbers of adverse events occurred in the tesaglitazar ≤ 1.0?mg, placebo, and pioglitazone arms, but there was an increasing frequency of discontinuations due to pre-specified hematologic and clinical-chemistry criteria with tesaglitazar doses ≥ 1.0?mg.

Conclusions: In type 2 diabetic patients, tesaglitazar dose-dependently reduced FPG levels at doses ≥ 0.5?mg. Other markers of glycemic control, atherogenic dyslipidemia, and measures associated with insulin resistance were improved at doses ≥ 0.5?mg or ≥ 1.0?mg. Study limitations included that the majority of patients were white, patients had good glycemic control at baseline, and the increased number of early withdrawals in the tesaglitazar 2.0?mg and 3.0?mg doses limits conclusions about the efficacy of these doses. The 0.5?mg and 1.0?mg tesaglitazar doses were identified for further investigation.     

Rutin ameliorates oxidative stress and preserves hepatic and renal functions following exposure to cadmium and ethanol

Context: Rutin (RUT) is an antioxidant flavonoid with well-known metal chelating potentials.

Objective: This study was designed to evaluate the protective effects of RUT against cadmium (Cd)?+?ethanol (EtOH)-induced hepatic and renal toxicity in rats.

Materials and methods: Wistar rats were treated with Cd (50?mg/kg) alone or in combination with EtOH (5?mg/kg) and RUT (25, 50 and 100?mg/kg) for 15?days. After treatment, the liver, kidney and serum were removed for biochemical assays by spectrophotometric methods.

Results: Serum, hepatic and renal malondialdehyde (MDA) levels were highest in the Cd?+?EtOH group and lowest in Cd?+?EtOH animals co-treated with the highest dose of RUT (2.98?±?0.34, 10.08?±?2.32, 4.99?±?1.21 vs. 1.69?±?0.33, 6.13?±?0.28, 3.66?±?1.12?μmol MDA/mg protein, respectively). The serum level of Cd was increased in the Cd?+?EtOH treated animals compared to Cd?+?EtOH animals co-treated with 100?mg/kg RUT (2.54?±?0.08 vs. 1.28?±?0.04?ppm). Furthermore, RUT at the highest dose protected against Cd?+?EtOH-induced elevation of bilirubin and uric acid levels as well as activities of lactate dehydrogenase and γ-glutamyl transferase (62.86?±?2.74 vs. 122.52?±?6.35?µmol/L; 1.77?±?0.35 vs. 3.23?±?0.55?mmol/L; 9.56?±?1.22 vs. 16.21?±?1.64?U/L; 288.92?±?40.12 vs. 159.8?±?18.01?U/L). The histo-pathological changes in the liver and kidney were also reduced in the Cd?+?EtOH animals co-treated with RUT in a dose-dependent manner.

Discussion and conclusion: RUT protected against the combined effects of Cd?+?EtOH on hepatic and renal functions and improved the antioxidant defence system in the blood.     

Evaluation of herbal antimalarial MAMA decoction-amodiaquine combination in murine malaria model

Context: Co-administration of amodiaquine with MAMA decoction (MD), an herbal antimalarial drug comprising the leaves of Mangifera indica L. (Anacardiaceae), Alstonia boonei De Wild (Apocynaceae), Morinda lucida Benth (Rubiaceae) and Azadirachta indica A. Juss (Meliaceae) was investigated. The practice of concurrent administration of herbal medicines with orthodox drugs is currently on the increase globally.

Objective: The study was designed to investigate the possible enhancement of the antimalarial potency as well as possible herb–drug interaction resulting from concurrent administration of MAMA decoction with amodiaquine (AQ).

Materials and methods: Combinations of MD with AQ were investigated in chloroquine (CQ)-sensitive Plasmodium berghei NK 65 in varying oral doses (mg/kg) at: sub-therapeutic [MD30?+?AQ1.25], therapeutic [MD120?+?AQ10] and median effective [MD40?+?AQ3.8], using chemosuppressive and curative antimalarial test models. Secondly, P. berghei ANKA (CQ-resistant)-infected mice were orally treated with MD 120, 240, [MD120?+?AQ10] and [MD240?+?AQ10] mg/kg, using both models. The survival times of mice were monitored for 28 d.

Results: ED₅₀ values of MD and AQ were 48.8 and 4.1?mg/kg, respectively. A total parasite clearance of CQ-sensitive P. berghei NK65 was obtained with the therapeutic combination dose in the curative test giving an enhanced survival time. In CQ-resistant P. berghei ANKA-infected mice, [MD120?+?AQ10] and [MD240?+?AQ10] mg/kg gave comparable activities with AQ (10?mg/kg) in both models.

Conclusion: The therapeutic combination dose gave total parasite clearance of CQ-sensitive P. berghei NK65, whereas none of the doses tested showed notable activity against CQ-resistant P. berghei ANKA.     

Comparison of protective and curative potential of Daucus carota root extract on renal ischemia reperfusion injury in rats

Abstract

Context: Daucus carota Linn (Apiaceae), a useful vegetable, is traditionally used in treating kidney and hepatic dysfunctions.

Objective: To evaluate the protective and curative potential of D. carota root extract on renal ischemia reperfusion injury in rats.

Materials and methods: Wistar rats were selected with 8?+?8 groups (n?=?6). Renal pedicles of rats were occluded for 45?min and allowed for reperfusion period. In protective and curative studies, 14 days prior and 14 days after the induction of ischemia/reperfusion (I/R), rats received petroleum ether extract (PEE 250 and 500?mg/kg), fractional methanol extract (FME 250 and 500?mg/kg) and direct methanol extract (DME 250 and 500?mg/kg) of Daucus carota root, orally, once daily.

Results: PEE at a dose of 500?mg/kg significantly (p?<?0.001) reduced the levels of serum creatinine (0.853–3.090?mg/dl), uric acid (1.300–3.500?mg/dl) and urea (58.26–132.00?mg/dl) compared to disease control. FME at a dose of 500?mg/kg body weight significantly (p?<?0.001) reduced the levels of serum creatinine (0.960–3.090?mg/dl), uric acid (1.700–3.500?mg/dl) and urea (77.17–132.00?mg/dl) compared to disease control. DME at a dose of 500?mg/kg body weight significantly (p?<?0.001) reduced the levels of serum creatinine (1.173–3.090?mg/dl), uric acid (2.267–3.500?mg/dl) and urea (84.75–132.00?mg/dl) compared to disease control.

Discussion and conclusion: Findings demonstrate that postconditioning with the D. carota root extract significantly improves kidney function in I/R rats.     

3-Aminobenzamide – a PARP inhibitor enhances the sensitivity of peripheral blood micronucleus and comet assays in mice

Context: DNA repair is an essential outcome of DNA damage, which may compromise the end point of various in vitro and in vivo test systems of the genotoxicity evaluation. poly(ADP-ribose) polymerase (PARP) enzymes have an essential role in DNA repair. Here, we investigated the effect of 3-AB, a PARP inhibitor on the sensitivity of comet and PBMN assays.

Objective: This study aimed to enhance the sensitivity of the comet and peripheral blood micronucleus (PBMN) assays using 3-aminobenzamide (3-AB), a well-characterized PARP inhibitor.

Materials and methods: Cyclophosphamide (CP, 50?mg/kg), 5-flourouracil (5-FU, 25?mg/kg), zidovudine (AZT, 400?mg/kg) and furosemide (FUR, 60?mg/kg) were selected as genotoxins. 3-AB was given every 8?h with the first dose given 2?h before the genotoxin treatment. For the PBMN assay, small amount of blood was taken from the tail tip of each animal and smears were prepared. The comet assay was performed in PBL, bone marrow and liver.

Results: In the comet as well as PBMN assay, 3-AB pre-treatment enhanced the extent of DNA damage in all the combination groups (3-AB?+?CP, 3-AB?+?5-FU and 3-AB?+?AZT) compared to CP, 5-FU and AZT per se. 3-AB also enhanced the DNA damage caused by FUR in the bone marrow and liver.

Discussion: This study results clearly demonstrate that the pretreatment with 3-AB (30?mg/kg) significantly enhances the sensitivity of the PBMN and comet assays. This model may be useful for the detection of marginally active DNA damaging agents.     

Additional therapy for cholesterol lowering in ezetimibe-treated,statin-intolerant patients in clinical practice: results from an internal audit of a university lipid clinic

Objective: The aim of our study was to evaluate the tolerability and efficacy of alternative approaches to improve cholesterolemia control in patients with statin-related myalgia treated with ezetimibe.

Research design and methods: We retrospectively evaluated 3534 Clinical Report Forms (CRFs) filled in the period June 2012–June 2015 for first visits to the lipid clinic of the University of Bologna. For this study, we selected 252 CRFs based on the following criteria: statin-related myalgia, previous failed treatment with at least two low-dosed statins, well tolerated treatment with ezetimibe. Then, the following lipid-lowering treatments were added in order to improve the ezetimibe low density lipoprotein cholesterol (LDL-C) lowering efficacy, based on clinical judgment: fenofibrate 145?mg, rosuvastatin 5?mg 1 tablet/week, rosuvastatin 5?mg 2 tablets/week, red yeast rice (standardized in monacolin K 3?mg) + berberine 500?mg, berberine 500?mg b.i.d., phytosterols 900?mg?+?psyllium fiber 3.5?g b.i.d. Patients continuing to claim a tolerable myalgia were then treated with coenzyme Q10 nanoemulsions 200?mg/day.

Results: The treatment with standard lipid-lowering diet plus ezetimibe alone was associated with a mean LDL-C reduction of 17?±?2%. The additive LDL-lowering effect with the various tested treatment was: ?16?±?2% with fenofibrate 145?mg/day, ?13?±?1% with rosuvastatin 5?mg 1 tablet/week, ?17?±?3% with rosuvastatin 5?mg 2 tablets/week, ?19?±?4% with red yeast rice?+?berberine, ?17?±?4% with berberine b.i.d. and ?10?±?3% with phytosterols?+?psyllium b.i.d.; 11% of the patients treated with fenofibrate required treatment modification because of myalgia recurrence, while the percentage was negligible for the other tested treatments. In patients with residual tolerable myalgia, treatment with coenzyme Q10 for 8 weeks was associated with a mean improvement of the graduated myalgia score from 4.8?±?1.9 to 2.9?±?1.3 (p?=?0.013).

Conclusions: Some alternative treatments seems to be effective and well tolerated, thus improving the ezetimibe effect on cholesterolemia.     

Metabolism and disposition of 2-ethylhexyl-p-methoxycinnamate following oral gavage and dermal exposure in Harlan Sprague Dawley rats and B6C3F1/N mice and in hepatocytes in vitro

1. 2-Ethylhexyl-p-methoxycinnamate (EHMC) is commonly used as an ingredient in sunscreens, resulting in potential oral and dermal exposure in humans.

2. Clearance and metabolism of EHMC in hepatocytes and disposition and metabolism of EHMC in rodents following oral (8–800?mg/kg) intravenous (IV) (8?mg/kg) or dermal (0.8–80?mg/kg representing 0.1–10% formulation concentration) exposure to [¹⁴C]EHMC were investigated in rats and mice.

3. EHMC was rapidly cleared from rat and mouse hepatocytes (half-life ≤3.16?min) and less rapidly (half-life ≤48?min) from human hepatocytes.

4. [¹⁴C]EHMC was extensively absorbed and excreted primarily in urine by 72?h after oral administration to rats (65–80%) and mice (63–72%). Oral doses to rats were excreted to a lesser extent (3–8%) in feces and as CO₂ (1–4%). Radioactive residues in tissues were <1% of the dose. There were no sex or species differences in disposition in rats.

5. Following dermal application, 34–42% of an 8–mg/kg dose was absorbed in rats, and 54–62% in mice in 72–h.

6. Among numerous urinary metabolites associated with hydrolysis of the ester, two potential reproductive and developmental toxicants, 2-ethylhexanol and 2-ethylhexanoic acid were produced by metabolism of EHMC.     

Potential mechanisms of neurobehavioral disturbances in mice caused by sub-chronic exposure to low-dose VOCs

Abstract

To investigate effects of neurobehavioral disturbances in mice caused by sub-chronic exposure to low-dose volatile organic compounds (VOCs) and the possible mechanism for these effects, 60 male Kunming mice were exposed in 5 similar static chambers, 0 (control) and 4 different doses of VOCs mixture (G1–4) for consecutively 90?d at 2?h/d. The concentrations of VOCs mixture were as follows: formaldehyde, benzene, toluene, and xylene 0.05?+?0.05?+?0.10?+?0.10?mg/m³, 0.10?+?0.11?+?0.20?+?0.20?mg/m³, 0.50?+?0.55?+?1.00?+?1.00?mg/m³, 1.00?+1.10?+?2.00?+?2.00?mg/m³, respectively, which corresponded to 1/2, 1, 5, and 10 times of indoor air quality standard in China. Morris water maze (MWM) and Grip strength (GS) test were performed in the last 7 weeks. One day following VOCs exposure, oxidative stress markers, neurotransmitters, and cholinergic system enzymes in brain were examined. In addition, the expressions of N-methyl-d-aspartate (NMDA) receptor in hippocampus were determined. VOCs exposure induced behavioral impairment of mice in MWM and GS test. The levels of reactive oxygen species (ROS), malondialdehyde (MDA) and glutamic acid (Glu) were significantly increased, while the acetylcholinesterase (AChE), choline acetyltransferase (ChAT) and acetylcholine (ACh) levels, and the expression of NMDA receptor were significantly decreased in VOCs exposed groups. Results showed that sub-chronic exposure to low-dose VOCs induced damage on physique and motor function, as well as impairment on learning and memory capacity of mice. Oxidative damage, abnormal metabolism of neurotransmitters and cholinergic system enzymes, and the alternation of NMDA receptor expression may be the possible mechanism for VOCs-induced neurotoxicity.     

Beneficial effect of Citrus limon peel aqueous methanol extract on experimentally induced urolithic rats

Context: Citrus limon (L.) Burm.f. (Rutaceace) is a commonly available fruit variety with high medicinal and industrial values.

Objective: Lemon peel (LP) extract was studied as a potent preventive and curative agent for experimentally induced hyperoxaluric rats.

Materials and methods: Gas chromatography–mass spectrometry (GC–MS) analyses and toxicity study were performed for aqueous methanol LP extract. Twenty-four Wistar rats were segregated into four groups. Group 1: Control; Group 2: Urolithic (ethylene glycol (EG) – 0.75%); Group 3: Preventive study (EG?+?LP extract administration from 0th to 7th week); Group 4: Curative study (EG?+?LP extract administration from 4th to 7th week). Animals received LP extract daily by oral administration (100?mg/kg body weight) for 7 weeks.

Results and discussion: GC–MS analyses revealed that compound 6 was abundant in the LP extract (32%) followed by compound 1 (～21%). The LD₅₀ value of LP extract was found to be >5000?mg/kg of body weight. Urolithic rats showed significantly higher urinary calcium and oxalate (4.47?±?0.44 and 18.86?±?0.55?mg/24 h, respectively) excretion compared with control and experimental rats. Renal function parameters like urea (84?±?8.5 and 96.1?±?3.6?mg/dL), creatinine (1.92?±?0.27 and 1.52?±?0.22?mg/dL), and urinary protein (2.03?±?0.02 and 2.13?±?0.16?mg/24 h) were also reduced by LP extract (p?<?0.001) and corroborated with tissue analyses (SOD, catalase, and MDA levels) and histological studies in normal and experimental animals. Immunohistochemical staining of THP and NF-κB in urolithic animals showed elevated expression than the control, while LP extract suppressed the expression of these proteins.

Conclusion: In conclusion, lemon peel is effective in curing kidney stone disease and also can be used to prevent the disease and its recurrence.     

DMP 504, a novel hydrogel bile acid sequestrant: III. Safety,tolerability, and cholesterol-lowering in healthy hypercholesterolemic subjects

This multiple-dose study of DMP 504, a hydrogel bile acid sequestrant, employed a randomized, double-blind, sequential-cohort design with placebo (blinded) and open-label cholestyramine (CS) controls. Ninety-three healthy primary hypercholesterolemic subjects (serum LDL cholesterol 130 to 200 mg/dL; triglycerides ≤280 mg/dL) maintaining a stable diet (NCEP Step One) for 3 weeks received either DMP 504 or placebo DMP 504 in capsules, or CS powder for suspension (16 g/d). The DMP 504 dose (0.9, 1.8, 2.7, 3.6, 5.4, 7.2 g/d) escalated with sequential enrollment of cohorts (DMP 504 n = 10, placebo n = 2, and CS n = 3 for Cohorts I–V; DMP 504 n = 8, placebo n = 2 for Cohort VI). An additional cohort (VII) included DMP 504 3.6 g/d, n = 9, and placebo, n = 2. LDL cholesterol (LDL-C) decreased in a dose-dependent manner (–15.8% to –34.1%; r = –0.56; P < 0.001) over the dose range 0.9 g/d to 7.2 g/d after 2 weeks of dosing with DMP 504 and +0.8% to –19.7% (r = –0.48; P < 0.001) over doses 0.9 g/d to 5.4 g/d after 6 weeks. LDL-C did not change with placebo but declined with cholestyramine by –28.3% at 2 weeks and –23.9% at 6 weeks. Plasma 7-α-hydroxy-4-cholesten-3-one, a marker of hepatic cholesterol 7-α-hydroxylase activity, increased dose-dependently with DMP 504 and with CS; change in LDL-C correlated inversely with 7-α-hydroxy-4-cholesten-3-one (r = –0.46; P < 0.001). There was no dose-limiting toxicity with DMP 504. One subject (DMP 504 2.7 g/d) withdrew because of gastrointestinal disturbance. Subject complaints were largely gastrointestinal in nature (flatulence, abdominal discomfort, diarrhea/constipation) occurring in 53% of the 64 DMP 504, 60% of 15 CS, and 29% of 14 placebo subjects. The novel hydrogel DMP 504 can induce significant LDL cholesterol lowering at doses of several grams/day with an acceptable side effect profile. Drug Dev. Res. 41:76–84, 1997. © 1997 Wiley-Liss, Inc.     

Efficacy and safety of initial combination therapy with sitagliptin and metformin in patients with type 2 diabetes: a 54-week study

ABSTRACT

Objective: To assess the 54-week efficacy and safety of initial combination therapy with sitagliptin and metformin in patients with type 2 diabetes and inadequate glycemic control (HbA_1c 7.5–11%) on diet and exercise.

Methods and materials: This was multinational study conducted at 140 clinical sites in 18 countries. Following an initial 24-week, double-blind, placebo-controlled period, patients entered a double-blind continuation period for an additional 30 weeks. Following the week 24 evaluation, patients remained on their previously assigned active, oral treatments: sitagliptin 50?mg b.i.d.?+?metformin 1000?mg b.i.d. (S100?+?M2000), sitagliptin 50?mg b.i.d.?+?metformin 500?mg b.i.d. (S100?+?M1000), metformin 1000?mg b.i.d. (M2000), metformin 500?mg b.i.d. (M1000), and sitagliptin 100?mg q.d. (S100). Patients initially randomized to placebo were switched to M2000 (designated PBO/M2000) at week 24. This report summarizes the overall safety and tolerability data for the 54-week study and presents efficacy results for patients randomized to continuous treatments who entered the 30-week continuation period.

Results: Of the 1091 randomized patients, 906 completed the 24-week placebo-controlled phase and 885 patients continued into the 30-week continuation period (S100?+?M2000 n?=?161, S100+M1000 n?=?160, M2000 n?=?153, M1000 n?=?147, S100 n?=?141, PBO/M2000 n?=?123). At baseline, patients included in the efficacy analysis had mean age of 54 years, mean BMI of 32?kg/m², mean HbA_1c of 8.7% (8.5–8.8% across groups), and mean duration of type 2 diabetes of 4 years. At week 54, in the all-patients-treated analysis of continuing patients, least-squares (LS) mean changes in HbA_1c from baseline were ?1.8% (S100?+?M2000), ?1.4% (S100?+?M1000), ?1.3% (M2000), ?1.0% (M1000), and ?0.8% (S100). The proportions of continuing patients with an HbA_1c?<?7% at week 54 were 67% (S100?+?M2000), 48% (S100?+?M1000), 44% (M2000), 25% (M1000), and 23% (S100). For the patients completing treatment through week 54, LS mean changes in HbA_1c from baseline were ?1.9% (S100?+?M2000), ?1.7% (S100?+?M1000), ?1.6% (M2000), ?1.2% (M1000), and ?1.4% (S100). Glycemic response was generally durable over time across treatments. All treatments improved measures of β-cell function (e.g., HOMA-β, proinsulin/insulin ratio). Mean body weight decreased from baseline in the combination and metformin monotherapy groups and was unchanged from baseline in the sitagliptin monotherapy group. The incidence of hypoglycemia was low (1–3%) across treatment groups. The incidence of gastrointestinal adverse experiences with the co-administration of sitagliptin and metformin was similar to that observed with metformin alone.

Limitations: The patient population evaluated in the 54-week efficacy analysis was a population of patients who entered the continuation period without receiving glycemic rescue therapy in the 24-week placebo-controlled period. Because the baseline HbA_1c inclusion criteria ranged from 7.5 to 11% and the glycemic rescue criterion was an HbA_1c?>?8% after week 24, there was a greater likelihood of glycemic rescue in the monotherapy groups; this led to more missing data in the continuation all-patients-treated population(CAPT) analysis and fewer patients contributing to the completers analysis in the monotherapy groups.

Conclusions: In this study, initial treatment with sitagliptin, metformin, or the combination therapy of sitagliptin and metformin provided substantial and durable glycemic control, improved markers of β-cell function, and was generally well-tolerated over 54 weeks in patients with type 2 diabetes.