Can SCSA and TUNEL forecast apoptosis-related motility depletion in Asthenozoospermia?

This study is an attempt to determine the power of SCSA and TUNEL for the evaluation of apoptosis status and apoptosis-related motility depletion in Asthenozoospermia. Fifty-one semen samples from Asthenozoospermic and 20 samples from fertile men participated in this study. SCSA and TUNEL were applied for the assessment of DNA integrity by flow cytometry. Annexin V conjugated with FITC labelling and FLICA method were used for the assessment of externalisation of phosphatidylserine and spermatozoon with active Caspase 3 respectively. SCSA results were shown to have a significant correlation with EPS in live spermatozoon (r = .85, p value = .00) and spermatozoon with active Caspase 3 (r = .633, p value = .00). TUNEL result was revealed to have a nonsignificant positive correlation with them. Then, Asthenozoospermic individuals were divided into two groups, SCSA higher and SCSA lower than 27%. Results interestingly indicated that the two groups significantly differed from each other in terms of TUNEL, EPS in live spermatozoon, spermatozoon with active Caspase 3 and sperm vitality (p value = .00). Both SCSA and TUNEL were correlated with apoptosis-related motility depletion in Asthenozoospermia. However, SCSA might be more powerful than TUNEL and could provide reliable information about DNA, chromatin integrity and apoptosis status in Asthenozoospermia.

Effects of exenatide on weight and appetite in overweight adolescents and young adults with Prader-Willi syndrome.

BACKGROUND: Prader-Willi syndrome (PWS) is associated with hyperphagia and hyperghrelinemia with major morbidity because of obesity without effective medical treatment targeting hyperphagia. Exenatide (Byetta [synthetic Exendin-4]; AstraZeneca, Wilmington DE) is a GLP-1 receptor agonist which reduces appetite and weight and may be an effective treatment in PWS. OBJECTIVE: The objective of this study is to determine the effect of a 6-month trial of exenatide on appetite, weight and gut hormones in youth with PWS. METHODS: Ten overweight and obese subjects with PWS (13-25 years) were recruited for a 6-month open-label, non-randomized, longitudinal study conducted at Children's Hospital Los Angeles. Exenatide was given using standard diabetes dosing without dietary modifications. Weight, body mass index (BMI), truncal fat, appetite and plasma acylated ghrelin were measured over 6 months. Mixed meal tolerance tests were performed at 0 and 6 months. RESULTS: Appetite scores significantly decreased from baseline (32.2 ± 8.7) after 1, 3 and 6 moths of treatment (27.5 ± 8.8, 25.4 ± 9.3, and 25.4 ± 7.2 respectively; p = 0.004). Hemoglobin A1c decreased significantly after treatment, but weight, BMI z-score and adiposity did not. There was no significant change in ghrelin. CONCLUSIONS: This is the first longitudinal investigation of the effects of exenatide in subjects with PWS. It was effective in decreasing appetite, without change in weight or BMI in the short term. Larger, controlled, longer-term trials in patients with PWS are needed to confirm the efficacy and safety of exenatide and to evaluate whether its use might induce weight loss when given in conjunction with behavioural modification.

Accuracy of computerized automatic identification of cephalometric landmarks by a designed software.

OBJECTIVES: The purpose of this study was to design software for localization of cephalometric landmarks and to evaluate its accuracy in finding landmarks. METHODS: 40 digital cephalometric radiographs were randomly selected. 16 landmarks which were important in most cephalometric analyses were chosen to be identified. Three expert orthodontists manually identified landmarks twice. The mean of two measurements of each landmark was defined as the baseline landmark. The computer was then able to compare the automatic system's estimate of a landmark with the baseline landmark. The software was designed using Delphi and Matlab programming languages. The techniques were template matching, edge enhancement and some accessory techniques. RESULTS: The total mean error between manually identified and automatically identified landmarks was 2.59 mm. 12.5% of landmarks had mean errors less than 1 mm. 43.75% of landmarks had mean errors less than 2 mm. The mean errors of all landmarks except the anterior nasal spine were less than 4 mm. CONCLUSIONS: This software had significant accuracy for localization of cephalometric landmarks and could be used in future applications. It seems that the accuracy obtained with the software which was developed in this study is better than previous automated systems that have used model-based and knowledge-based approaches.

Islet transplantation for brittle type 1 diabetes: the UIC protocol.

This prospective phase 1/2 trial investigated the safety and reproducibility of allogeneic islet transplantation (Tx) in type I diabetic (T1DM) patients and tested a strategy to achieve insulin-independence with lower islet mass. Ten C-peptide negative T1DM subjects with hypoglycemic unawareness received 1-3 intraportal allogeneic islet Tx and were followed for 15 months. Four subjects (Group 1) received the Edmonton immunosuppression regimen (daclizumab, sirolimus, tacrolimus). Six subjects (Group 2) received the University of Illinois protocol (etanercept, exenatide and the Edmonton regimen). All subjects became insulin- independent. Group 1 received a mean total number of islets (EIN) of 1460 080 +/- 418 330 in 2 (n = 2) or 3 (n = 2) Tx, whereas Group 2 became insulin- independent after 1 Tx (537 495 +/- 190 968 EIN, p = 0.028). All Group 1 subjects remained insulin free through the follow-up. Two Group 2 subjects resumed insulin: one after immunosuppression reduction during an infectious complication, the other with exenatide intolerance. HbA1c reached normal range in both groups (6.5 +/- 0.6 at baseline to 5.6 +/- 0.5 after 2-3 Tx in Group 1 vs. 7.8 +/- 1.1 to 5.8 +/- 0.3 after 1 Tx in Group 2). HYPO scores markedly decreased in both groups. Combined treatment of etanercept and exenatide improves islet graft function and facilitates achievement of insulin-independence with less islets.

Lower third molar eruption following orthodontic treatment.

This study assessed the effect of extraction and preservation of the 1st premolar on lower 3rd molar eruption. Orthodontic clinic records from 1993 to 1995 were evaluated before and after treatment and 8-9 years after treatment for 3 groups of patients: 32 with extraction of 1st premolars in both jaws, 32 with no extraction but orthodontic treatment and 48 controls with no extraction but orthodontic treatment in the upper jaws only. Successful eruption of 3rd molars was evaluated. There was a significant difference in the rates of successful eruptions in the extraction (42%), non-extraction (12%) and control (20%) groups. The findings indicate that 1st premolar extraction may increase the chance of 3rd molar eruption, leading to a lower incidence of health and economic complications.

Lower third molar eruption following orthodontic treatment

This study assessed the effect of extraction and preservation of the 1st premolar on lower 3rd molar eruption. Orthodontic clinic records from 1993 to 1995 were evaluated before and after treatment and 8-9 years after treatment for 3 groups of patients: 32 with extraction of 1st premolars in both jaws, 32 with no extraction but orthodontic treatment and 48 controls with no extraction but orthodontic treatment in the upper jaws only. Successful eruption of 3rd molars was evaluated. There was a significant difference in the rates of successful eruptions in the extraction [42%], non-extraction [12%] and control [20%] groups. The findings indicate that 1st premolar extraction may increase the chance of 3rd molar eruption, leading to a lower incidence of health and economic complications

Orthodontic treatment needs of 12-15-year-old students in Shiraz, Islamic Republic of Iran.

The study used the Dental Aesthetic Index to evaluate the orthodontic treatment needs of adolescents from state junior high schools in Shiraz city, Islamic Republic of Iran. A random sample of 900 pupils (450 girls, and 450 boys) aged 12-15 years old, who received no orthodontic treatment before or during the study, were selected randomly over 2 months in 2004. Most of the pupils (70.1%) had normal or minor malocclusion indicating no need for orthodontic treatment. Only 4.2% had disabling malocclusion that required treatment. Severe and very severe grades of malocclusion were more common in boys than girls. Iranian youth from Shiraz had better dental appearance and needed less orthodontic treatment than other populations.

Orthodontic treatment needs of 12-15-year-old students in Shiraz, Islamic Republic of Iran

The study used the Dental Aesthetic Index to evaluate the orthodontic treatment needs of adolescents from state junior high schools in Shiraz city, Islamic Republic of Iran. A random sample of 900 pupils [450 girls, and 450 boys] aged 12-15 years old, who received no orthodontic treatment before or during the study, were selected randomly over 2 months in 2004. Most of the pupils [70.1%] had normal or minor malocclusion indicating no need for orthodontic treatment. Only 4.2% had malocclusion that required treatment. Severe and very severe grades of malocclusion were more disabling common in boys than girls. Iranian youth from Shiraz had better dental appearance and needed less orthodontic treatment than other populations

Improved outcomes in islet isolation and transplantation by the use of a novel hemoglobin-based O2 carrier.

During isolation, islets are exposed to warm ischemia. In this study, intraductal administration of oxygenated polymerized, stroma-free hemoglobin-pyridoxalated (Poly SFH-P) was performed to improve O2 delivery. Rat pancreata subjected to 30-min warm ischemia were perfused intraductally with collagenase in oxygenated Poly SFH-P/RPMI or RPMI (control). PO2 was increased by Poly SFH-P (381.7 +/- 35.3 mmHg vs. 202.3 +/- 28.2, p = 0.01) and pH maintained within physiological range (7.4-7.2 vs. 7.1-6.6, p = 0.009). Islet viability (77% +/- 4.6 vs. 63% +/- 4.7, p = 0.04) was improved and apoptosis lower with Poly SFH-P (caspase-3: 34,714 +/- 2167 vs. 45,985 +/- 1382, respectively, p = 0.01). Poly SFH-P improved islet responsiveness to glucose as determined by increased intracellular Ca2+ levels and improved insulin secretion (SI 5.4 +/- 0.1 vs. 3.1 +/- 0.2, p = 0.03). Mitochondrial integrity was improved in Poly SFH-P-treated islets, which showed higher percentage change in membrane potential after glucose stimulation (14.7% +/- 1.8 vs. 9.8 +/- 1.4, respectively, p < 0.05). O2 delivery by Poly SFH-P did not increase oxidative stress (GSH 7.1 +/- 2.9 nm/mg protein for Poly SFH-P vs. 6.8 +/- 2.4 control, p = 0.9) or oxidative injury (MDA 1.8 +/- 0.9 nmol/mg protein vs. 6.2 +/- 2.4, p = 0.19). Time to reach normoglycemia in transplanted diabetic nude mice was shorter (1.8 +/- 0.4 vs. 7 +/- 2.5 days, p = 0.02), and glucose tolerance improved in the Poly SFH-P group (AUC 8106 +/- 590 vs. 10,863 +/- 946, p = 0.03). Oxygenated Poly SFH-P improves islet isolation and transplantation outcomes by preserving mitochondrial integrity.

Distribution of malocclusion types in 7-9-year-old Iranian children.

This study assessed the malocclusion types, very severe crowding and need for serial extraction among a random sample of 7-9-year-old children in Shiraz, Islamic Republic of Iran. Of the 3776 children 30.6% had normal occlusion, 47.4% class I malocclusion, 13.7% class II division 1 malocclusion (male/female ratio 3:2), 1.0% class II division 2 malocclusion (male/female ratio 3:1) and 2.1% class III malocclusion. Among the children examined, 47.9% had crowding problems and 14.7% of them had class I malocclusion with very severe crowding-more girls (17.3%) than boys (12.1%). No correlation was observed between the types of malocclusion and family size, parents' occupation or level of education.

Blood pressure distribution in an Iranian adolescent population: "Tehran Lipid and Glucose study".

Significant hypertension in adolescence is defined according to Blood Pressure (BP) distribution among general population. The present study was conducted to determine the BP distribution in Iranian adolescents. In a population-based study held in Tehran, BP data for 2560 participants aged 10-17 years (1247 boys and 1313 girls) were collected. The 90th and 95th percentiles of systolic (SBP) and diastolic BP (DBP) were obtained for different age and height subgroups of boys and girls. While the 90th and 95th percentiles of SBP obtained in our study were generally lower than international guidelines, the corresponding percentiles of DBP were higher. Significant associations were found between BP and age, height, weight and BMI. According to our findings, it is necessary to prepare BP reference tables according to regional surveys.

Distribution of malocclusion types in 7-9-year-old Iranian children

This study assessed the malocclusion types, very severe crowding and need for serial extraction among a random sample of 7-9-year-old children in Shiraz, Islamic Republic of Iran. Of the 3776 children 30.6% had normal occlusion, 47.4% class I malocclusion, 13.7% class II division 1 malocclusion [male/female ratio 3:2], 1.0% class II division 2 malocclusion [male/female ratio 3:1] and 2.1% class III malocclusion. Among the children examined, 47.9% had crowding problems and 14.7% of them had class I malocclusion with very severe crowding-more girls [17.3%] than boys [12.1%]. No correlation was observed between the types of malocclusion and family size, parents' occupation or level of education

Cephalometric evaluation of class-III patients with chin cap and tongue guard.

The purpose of this study was to determine the effect of chin cap therapy combined with an upper tongue guard in the early treatment of class-III malocclusion on the nasomaxillary complex and mandible. The subjects of this study consisted of 40 patients aged 5-13 years (mean age of 8.5 +/- 2). All of them possessed an anterior cross bite and/or concave profile. The mean force of chin cap was determined to be 200 g on each side for 18 h/day and the mean treatment period was 22 months. The cephalometric analyses including skeletal, dental, and soft tissue analysis were carried out before and after treatment. The analysis of the cephalometric measurements revealed a negative correlation between the combination effects of chin cap therapy upper tongue guard. This effect appeared in the early stages of treatment. It was a case-control study and Wilcoxon test was used for statistical analysis. The comparison of dependent variables revealed that skeletal effects of chin cap therapy were more than dental and soft tissue effects. Changes in the upper and lower pharyngeal spaces were not significant. A reduction of nasolabial angle occurred due to the protrusion of upper incisors. Finally, it was shown that the combination of chin cap and upper tongue guard could be more effective in the early treatment.

Serum lipid levels in an Iranian adults population: Tehran Lipid and Glucose Study.

Data from 6246 participants aged 20-64 years (2339 males and 3907 females) in the cross-sectional phase of Tehran Lipid and Glucose Study (February 1999-May 2000) were used to determine distribution of serum lipid levels after 12-14 hour overnight fast. Mean total cholesterol (TC) concentration was 210 mg/dl. TC was significantly greater in females than males, 213 and 206 mg/dl, respectively (p < 0.0001). Thirty-one percent of population had TC values between 200 and 239 and 24% had values of 240 mg/dl or greater. Mean low-density lipoprotein cholesterol (LDL-C) was 129 and 135 mg/dl in males and females, respectively (p < 0.0001). Twenty-seven percent had LDL-C values between 130 and 159 and 23% had values 160 mg/dl or greater. The mean triglycerides (TGs) values were 190 and 162 mg/dl for males and females, respectively (p < 0.0001). The mean high-density lipoprotein cholesterol (HDL-C) was 39 in males and 45 mg/dl in females (p < 0.0001). The results showed higher levels of TC, LDL-C and TGs and slightly lower HDL-C in Tehranian adults than other studies in the industrialized countries.

Improvement of pancreatic islet isolation outcomes using glutamine perfusion during isolation procedure.

During procurement, isolation, and transplantation, islets are exposed to high levels of oxidative stress triggering a variety of signaling pathways that can ultimately lead to cell death. Glutamine is an important cellular fuel and an essential precursor for the antioxidant glutathione. The aim of this study was to examine the role of intraductal glutamine administration in facilitating recovery of isolated rat islets from pancreases subjected to a clinically relevant period of warm ischemia. Islets were isolated in Sprague-Dawley (SD) rats (n = 18 per group). Pancreata in groups 1 and 2 were procured immediately while groups 3 and 4 were subjected to 30-min warm ischemia. Groups 2 and 4 were treated intraductally with 5 mM glutamine prior to pancreatectomy. Exposure to 30-min warm ischemia significantly reduced islet yield [groups 1 & 2 (nonischemia): 503 +/- 29 islets/rat vs. groups 3 & 4 (ischemia): 247 +/- 26 islets/rat; p < 0.05]. Intraductal glutamine treatment significantly improved islet yield when pancreata were subjected to 30-min warm ischemia [144 +/- 16 islets/rat without glutamine (group 3) vs. 343 +/- 36 islets/rat with glutamine (group 4), p < 0.05]. Glutamine also significantly improved islet viability (values were 50 +/- 4% in group 4 vs. 27 +/- 3% in group 3, p < 0.05). Similarly, glutathione (reduced) levels were significantly elevated in both glutamine-treated groups; however, this increase was greatest in tissues exposed to ischemia (2.76 +/- 0.04 nmol/mg protein in group 4 vs. 1.66 +/- 0.04 nmol/mg protein in group 3, p < 0.05). Intraductal glutamine administration considerably improves the islet yield, viability, and augments endogenous glutathione levels in pancreata procured after a clinically relevant period of ischemia. Intraductal administration of glutamine at the time of digestive enzyme delivery into the harvested pancreas may represent a simple yet effective tool to improve islet yields in clinical isolations.

Improvement of Pancreatic Islet Isolation Outcomes Using Glutamine Perfusion during Isolation Procedure.

During procurement, isolation, and transplantation, islets are exposed to high levels of oxidative stress triggering a variety of signaling pathways that can ultimately lead to cell death. Glutamine is an important cellular fuel and an essential precursor for the antioxidant glutathione. The aim of this study was to examine the role of intraductal glutamine administration in facilitating recovery of isolated rat islets from pancreases subjected to a clinically relevant period of warm ischemia. Islets were isolated in Sprague-Dawley (SD) rats (n= 18 per group). Pancreata in groups 1 and 2 were procured immediately while groups 3 and 4 were subjected to 30-min warm ischemia. Groups 2 and 4 were treated intraductally with 5 mM glutamine prior to pancreatectomy. Exposure to 30-min warm ischemia significantly reduced islet yield [groups 1 & 2 (nonischemia): 503 ± 29 islets/rat vs. groups 3 & 4 (ischemia): 247 ± 26 islets/rat; p < 0.05]. Intraductal glutamine treatment significantly improved islet yield when pancreata were subjected to 30-min warm ischemia [144 ± 16 islets/rat without glutamine (group 3) vs. 343 ± 36 islets/rat with glutamine (group 4), p < 0.05]. Glutamine also significantly improved islet viability (values were 50 ± 4% in group 4 vs. 27 ± 3% in group 3, p < 0.05). Similarly, glutathione (reduced) levels were significantly elevated in both glutamine-treated groups; however, this increase was greatest in tissues exposed to ischemia (2.76 ± 0.04 nmol/mg protein in group 4 vs. 1.66 ± 0.04 nmol/mg protein in group 3, p < 0.05). Intraductal glutamine administration considerably improves the islet yield, viability, and augments endogenous glutathione levels in pancreata procured after a clinically relevant period of ischemia. Intraductal administration of glutamine at the time of digestive enzyme delivery into the harvested pancreas may represent a simple yet effective tool to improve islet yields in clinical isolations.

Determinants of serum HDL-C level in a Tehran urban population: the Tehran Lipid and Glucose Study.

BACKGROUND AND AIM: Decreased serum high-density lipoprotein cholesterol (HDL-C) is one of the most common lipid disorders in patients with coronary artery disease (CAD). Existing evidence suggests that every 1 mg/dL decrease in serum HDL-C increases the risk of CAD by 2-3%. This study was performed in the year 2000 to study HDL-C determinants in a Tehran population. METHODS AND RESULTS: We studied 9514 subjects (3942 men and 5572 women) aged 20-69 years, who participated in the Tehran Lipid and Glucose Study (TLGS), completed a personal history questionnaire (especially concerning physical activity and cigarette smoking), and underwent a clinical examination including anthropometric and blood pressure measurements. Serum total cholesterol, triglyceride and HDL-C levels were measured, and OGTT was used to define diabetic patients according to WHO criteria. The women had a significantly higher mean HDL-C level than the mean (45 +/- 11 vs 38 +/- 9 mg/dL; p < 0.001); low HDL-C levels (< 35 mg/dL) were observed in 31% of the men and 13% of the women (p < 0.001). Obese subjects (BMI > or = 30 kg/m2) had a significantly lower HDL-C level than the normal subjects (42 +/- 11 vs 44 +/- 11 mg/dL: p < 0.001), and those with truncal obesity (WHR > or = 0.95 in men and > or = 0.8 in women) lower HDL-C levels than the normal subjects (37 +/- 9 vs 39 +/- 10 mg/dL in men and 44 +/- 11 vs 42 +/- 11 mg/dL in women; p < 0.001 for both). Smokers had a significantly lower HDL-C level than non-smokers (38 +/- 10 vs 43 +/- 11 mg/dL; p < 0.001) and a low HDL-C level was twice as common (36.4 vs 18.2%). Passive smokers also had lower HDL-C levels (42 +/- 11 vs 43 +/- 11 mg/dL; p < 0.001). Mean serum HDL-C was significantly lower in hypertriglyceridemic than those with normal triglycerides levels (men: 4 +/- 8 vs 40 +/- 9 mg/dL, p < 0.001; women: 40 +/- 10 vs 47 +/- 11 mg/dL, p < 0.01). Mean HDL-C levels were similar in subjects with different degrees of physical activity, as well as between diabetics and non-diabetics and hypertensive and normotensive subjects. Multiple stepwise regression analysis showed that the determinants of serum HDL-C levels were, in order of entering the model: hypertriglyceridemia (OR 3.4, p < 0.001), male sex (OR 3.1, p < 0.001), cigarette smoking (OR 1.7, p < 0.001), obesity (OR 1.4, p < 0.01), age (OR 0.9, p < 0.05), high WHR (OR 1.2, p < 0.05), and passive smoking (OR 1.1, p < 0.05). Physical activity, hypertension, and diabetes mellitus did not enter the predictive model. CONCLUSION: Apart from age and sex which are constitutional, and unmodifiable variables, the determinants of HDL-C level (hypertriglyceridemia, obesity, truncal obesity, cigarette smoking, and passive smoking) can be used in community CAD prevention programmes.

Cardiovascular risk factors in an Iranian urban population: Tehran lipid and glucose study (phase 1).

OBJECTIVES: Coronary artery disease is becoming more prevalent in developing countries, particularly in urban areas. This study was conducted to determine the prevalence of cardiovascular risk factors among Tehran urban population. METHODS: The prevalence and distribution of high blood pressure, cigarette smoking, dyslipoproteinemia, diabetes mellitus, and obesity was determined in 15,005 subjects, aged three years and over, selected by cluster random sampling in Tehran urban district-13 between February 1999 to August 2001. Total energy intake, percent of energy derived from protein, carbohydrate, and fat were assessed in a subsidiary of 1,474 persons by means of two 24-hour dietary recalls. RESULTS: In adults, 78% of men and 80% of women presented at least one CVD risk factor. The percentage of adult women with two or more risk factors was significantly greater than the one for men. Prevalence of DM, hypertension, obesity, high TC, low HDL, high TGs, and smoking was 9.8, 20.4, 14.4, 19.3, 32, 5.3, and 22.3%, respectively. In children and adolescents, two or more CVD risk factors were found in 9% of boys and 7% of girls. Prevalence of hypertension, obesity, high TC, low HDL, and high TGs, was 12.7, 5.2, 5.1, 10.2, and 5%, respectively. The mean percentage values of energy intake derived from carbohydrate, protein, and fat were 57.8 +/- 6.9, 11.1 +/- 1.8, and 30.9 +/- 7.2, respectively. CONCLUSION: The prevalence of cardiovascular risk factors among Tehran urban population is high; particularly of high total cholesterol, low HDL cholesterol levels, and high waist to hip ratio. An effective strategy for life style modification is a cornerstone of a population approach to the cardiovascular risk factors. Moreover, these results will serve as a baseline for assessment of future trends in the risk factors studied.

Caffeic acid, chlorogenic acid, and dihydrocaffeic acid metabolism: glutathione conjugate formation.

The antioxidant properties of the dietary dihydroxycinnamic acids [caffeic (CA), dihydrocaffeic (DHCA), and chlorogenic (CGA) acids] have been well studied but little is known about their metabolism. In this article, evidence is presented showing that CA, DHCA, and CGA form quinoids and hydroxylated products when oxidized by peroxidase/H(2)O(2) or tyrosinase/O(2). Mass spectrometry analyses of the metabolites formed with peroxidase/H(2)O(2)/glutathione (GSH) revealed that mono- and bi-glutathione conjugates were formed for all three compounds except CGA, which formed a bi-glutathione conjugate only when GSH was present. In contrast, the metabolism of the dihydroxycinnamic acids by tyrosinase/O(2)/GSH resulted in the formation of only mono-glutathione conjugates. In the absence of GSH, hydroxylated products and p-quinones of CA or CGA were formed by peroxidase/H(2)O(2). DHCA formed a hydroxylated adduct (even though GSH was present), as well as the corresponding p-quinone and dihydroesculetin, an intramolecular cyclization product. NADPH also supported rat liver microsomal-catalyzed CA-, CGA-, and DHCA-glutathione conjugate formation, which was prevented by benzylimidazole, a cytochrome P450 inhibitor. Furthermore, the cytotoxicity of CA, CGA, and DHCA toward isolated rat hepatocytes was markedly enhanced by hydrogen peroxide or cumene hydroperoxide-supported cytochrome P450 and was prevented by benzylimidazole. Cytotoxicity was also markedly enhanced by dicumarol, an NADPH/oxidoreductase inhibitor. These results suggest that dihydroxycinnamic acids were metabolically activated by P450 peroxidase activity to form cytotoxic quinoid metabolites.

Catechin metabolism: glutathione conjugate formation catalyzed by tyrosinase, peroxidase, and cytochrome p450.

The metabolic pathways of dietary flavonoids are still largely unknown. In the present work, mass spectrometry and UV-vis spectroscopy studies were used to show that the naturally occurring flavonoid catechin underwent enzymatic oxidation by tyrosinase in the presence of glutathione (GSH) to form mono-, bi-, and tri-glutathione conjugates of catechin and mono- and bi-glutathione conjugates of a catechin dimer. A hydroxylated catechin adduct was also detected. Using UV spectroscopy, it was shown that the catechol B-ring of catechin was oxidized by tyrosinase to form an o-quinone which could be reduced back to catechin with potassium borohydride or reacted with GSH to form glutathione conjugates. The catechin-glutathione conjugates formed had much lower distribution coefficient values than catechin itself. When peroxidase and hydrogen peroxide were used instead of tyrosinase, only mono-glutathione conjugates were formed but not bi-glutathione conjugates or hydroxylated adducts. (1)H NMR evidence showed that three different mono-glutathione conjugates on ring B of catechin were formed by peroxidase and hydrogen peroxide. Rat liver microsomes and NADPH or cumene hydroperoxide also catalyzed catechin-glutathione conjugate formation which was prevented by benzylimidazole, a P450 2E1 inhibitor. Catechin cytotoxicity toward isolated hepatocytes was also markedly enhanced by hydrogen peroxide or cumene hydroperoxide and was prevented by benzylimidazole, suggesting that catechin could be metabolically activated by P450 peroxidase activity to form cytotoxic quinoid species.