Genistein effects on quality of life and depression symptoms in osteopenic postmenopausal women: a 2-year randomized, double-blind, controlled study.

SUMMARY: Postmenopausal estrogen decline is implicated in several age-related physical and psychological changes in women, including decreases in perceived quality of life. The phytoestrogen genistein at a dose of 54 mg daily in osteopenic postmenopausal women after 2 years implies an improvement on quality of life and depression symptoms. INTRODUCTION: Postmenopausal estrogen decline is implicated in several age-related physical and psychological changes in women, including decreases in perceived quality of life (QoL). A number of trials with hormone therapy showed beneficial effects of the intervention on quality of life parameters. However, because of known or suspected serious side effects of conventional hormone therapy, there is a need for alternatives. METHODS: We conducted a double-blind randomized placebo-controlled trial using the isoflavone genistein, 54 mg, or placebo for 2 years. In this trial, we recruited 262 postmenopausal women aged 49 to 67 years. RESULTS: At baseline, after 1 year, and at final visit, participants filled in the Short Form of 36 questions (SF-36) and the Zung Self-rating Depression Scale (ZSDS). For the placebo group, scores on all dimensions of the SF-36 decreased after 1 and 2 years. The genistein group showed increases on all dimensions of the SF-36 at the end of the study. There were, however, statistically significant differences in changes of scores between the two intervention groups. For the ZSDS, similarly, significant differences were found between groups. CONCLUSION: In conclusion, the findings of this randomized trial showed that genistein improves quality of life (health status, life satisfaction, and depression) in osteopenic postmenopausal women.

Efficacy of genistein aglycone on some cardiovascular risk factors and homocysteine levels: A follow-up study.

BACKGROUND AND AIM: Recent evidence suggests that genistein aglycone may act beneficially on surrogate cardiovascular risk markers in postmenopausal women. We assessed the effects of genistein aglycone on some cardiovascular risk factors and homocysteine levels after 3-years of continued therapy in a cohort of osteopenic, postmenopausal women. METHODS AND RESULTS: The parent study was a randomized, double-blind, placebo-controlled trial involving 389 postmenopausal women with low bone mass for 24 months. Subsequently, a subcohort (138 patients) continued therapy for an additional year. Participants received 54mg of genistein aglycone (n=71) or placebo (n=67), daily. Both arms received calcium and vitamin D(3) in therapeutic doses. Moreover, 4 weeks before randomization procedures and during our follow-up study, all patients received dietary instructions in an isocaloric fat-restricted diet. Blood lipid profiles, fasting glucose and insulin, insulin resistance (HOMA-IR), fibrinogen, osteoprotegerin (OPG) and homocysteine at baseline and after 24 and 36 months of treatment were measured. Compared to placebo, genistein significantly decreased fasting glucose and insulin, HOMA-IR, fibrinogen and homocysteine after 24 and 36 months of treatment. By contrast, isoflavone administration did not affect high-density lipoprotein cholesterol and triglycerides though serum OPG was higher in the genistein recipients. There were no differences in adverse events or discomfort between groups. Results on routine biochemical, liver function, and hematologic testing did not change over time in placebo or genistein group. CONCLUSIONS: After 3-years of treatment, genistein aglycone plus calcium, vitamin D(3) and a healthy diet showed positive effects on some cardiovascular risk factors and homocysteine levels in a cohort of postmenopausal women with low bone mass.

Genistein effects on quantitative ultrasound parameters and bone mineral density in osteopenic postmenopausal women.

UNLABELLED: This study aimed at evaluating the effects of genistein (54 mg/die) on calcaneus and phalanges ultrasound (QUS) parameters and bone mineral density in osteopenic postmenopausal women. We concluded that genistein prevented bone loss in the osteopenic postmenopausal women and improves QUS parameters at the calcaneus and phalanges. INTRODUCTION: The purpose of the study was to evaluate the effects of genistein (54 mg/die) on quantitative ultrasound (QUS) parameters of the calcaneus and hand phalange and on bone mineral density (BMD) in osteopenic postmenopausal women. METHODS: One hundred thirty-eight women (age 49-67 years) were assigned to receive genistein or placebo. Bone status was assessed by measuring the anteroposterior lumbar spine and femoral neck BMD by dual energy X-ray absorptiometry and by ultrasound of the calcaneus (Achilles Plus, GE, Lunar) and of the phalanges (Bone Profiler. IGEA) at baseline and after a 1- and 2-year treatment. RESULTS: At the end of the experimental period, genistein had significantly increased BMD in the femur and lumbar spine (p < 0.001). The stiffness index, amplitude-dependent speed of sound, and bone transmission time in the genistein group had increased significantly at the end of study (+5.3, p < 0.001; +3.6%, p < 0.001; +4.6, p < 0.001, respectively). CONCLUSIONS: This study confirms that genistein prevented bone loss in the osteopenic postmenopausal women and it improves the QUS parameters.

Haemostatic effects of phytoestrogen genistein in postmenopausal women.

INTRODUCTION: Genistein is an isoflavone phytoestrogen derived from the soybean which acts as natural selective estrogen receptor modulator. Various studies have pointed out its cardioprotective role. The aim of the study was to evaluate the haemostatic effects of genistein in postmenopausal women. MATERIAL AND METHODS: In this double-blind placebo-controlled trial we enrolled 104 healthy postmenopausal women with osteopenia. 53 patients (mean age 54.9+/-4.2 yr; BMI 23.4+/-3.2 Kg/m(2)) received genistein (54 mg/day) and 51 patients (mean age 55.4+/-4.3 yr; BMI 23.6+/-3.6 Kg/m(2)) received an identical placebo-tablet. Both groups received a calcium and vitamin D supplement. Plasma levels of D-dimer (DD), plasminogen activator inhibitor-1 (PAI-1) and prothrombin fragment 1+2 (F1+2) were measured at baseline and after 6 and 12 months of treatment. RESULTS: Baseline characteristics of the two groups were similar. Compared with placebo, genistein decreased significantly DD (p<0.001), but did not affect PAI-1 and F 1+2 plasma levels. CONCLUSION: The results of our study do not confirm effects of genistein on activation of the haemostatic system, but on the contrary the significant decrease of DD could indicate a possible cardioprotective role of genistein in postmenopausal women.

Bisphosphonates in the treatment of thalassemia-associated osteoporosis.

Thalassemia major is a common cause of skeletal morbidity, as shown by the increased fracture risk in thalassemic patients. The etiology of this bone disease is multifactorial and culminates in a state of increased bone turnover with excessive bone resorption and remodeling. Despite hormonal replacement therapy, calcium and vitamin D administration, effective iron chelation, and normalization of hemoglobin levels, patients with thalassemia major continue to lose bone mass. The increased bone turnover rate observed in thalassemic patients justifies the use of powerful anti-resorption drugs, such as bisphosphonates. To date, alendronate, pamidronate, and zoledronate seem to be effective in increasing bone mineral density and normalizing bone turnover, but more trials are necessary to evaluate their efficacy in reducing fracture risks in larger thalassemic populations.

Pulmonary hypertension and erythropoietin.

Numerous uremic patients on hemodialysis have pulmonary hypertension attributable to the presence of arteriovenous fistulas, vascular calcification, and endothelial dysfunction due to alterations in the balance between vasoconstrictive and vasodilatory substances. For these reasons, the effects of recombinant human erythropoietin, a drug widely used in patients on dialysis, on the pulmonary circulation were studied. Some authors maintain that recombinant human erythropoietin has an antihypertensive effect, while others have observed that this hormone induces a reduction in pulmonary arterial pressure due to its vasoactive and stimulatory effects on endothelial and smooth muscle cell precursors.

Randomized, double-blind, placebo-controlled study on effects of raloxifene and hormone replacement therapy on plasma no concentrations, endothelin-1 levels, and endothelium-dependent vasodilation in postmenopausal women.

The endothelium is thought to play an important role in the genesis of atherosclerosis, and several lines of evidence suggest that the effect of an intervention on endothelial function might predict its involvement in coronary disease progression and in the rate of cardiovascular events. Estrogen has direct effects on the blood vessel wall, indicating that vascular endothelium may play a key role in the cardiovascular protective effects of hormone replacement therapy (HRT). Raloxifene relaxes coronary arteries in vitro by an estrogen receptor-dependent and NO-dependent mechanism, thus suggesting that this selective estrogen receptor modulator could also have beneficial effects on endothelial function. This study compared the effects of HRT and raloxifene on NO products, endothelin-1 plasma levels, and endothelium-dependent vasodilatation in postmenopausal women. Healthy postmenopausal women (n=90) were enrolled in a double-blind, randomized, placebo-controlled, 6-month trial. Women were randomly assigned to receive continuous HRT (1 mg 17beta-estradiol combined with 0.5 mg norethisterone acetate), raloxifene (60 mg/d), or placebo for 6 months. Flow-mediated endothelium-dependent vasodilation of the brachial artery, plasma NO concentrations, and endothelin levels were measured at baseline and after 6 months of therapy. The mean baseline level of NO breakdown products was 26.5+/-10.7 micromol/L and increased to 36.3+/-11.4 micromol/L after 6 months of treatment with raloxifene. The mean baseline plasma endothelin level was 17.3+/-8.9 pg/mL and decreased to 11.5+/-2.1 pg/mL after 6 months of treatment with the selective estrogen receptor modulator. The mean baseline ratio of NO (breakdown products) to endothelin was also significantly increased at the end of treatment with raloxifene. Postmenopausal women treated with HRT had similar changes in plasma nitrites/nitrates and endothelin levels as well as in the ratio of NO to endothelin. In contrast, these markers of endothelial function did not change in the placebo-treated women. Flow-mediated endothelium-dependent vasodilation of the brachial artery was 8.3+/-2.1% at baseline and increased to 12.3+/-2.1% after 6 months of treatment with raloxifene. HRT also caused a significant and similar increase in flow-mediated endothelium-dependent vasodilation. No change in flow-mediated vasodilation was observed in the participants treated with placebo. We conclude that raloxifene therapy and HRT influence endothelial function and improve flow-mediated endothelium-dependent vasodilation to a comparable extent in healthy postmenopausal women at least after a 6-month treatment period. However, further investigation is warranted to enhance our understanding of the mechanisms of the effect of raloxifene on vascular function and to determine whether its effect on endothelial function may contribute to the reduction in cardiovascular-related morbidity and mortality.

The pleiotropic effects of erythropoietin in the central nervous system.

Erythropoietin (Epo) is a hydrophobic sialoglycoproteic hormone produced by the kidney and responsible for the proliferation, maturation, and differentiation of the precursors of the erythroid cell line. Human recombinant erythropoietin (rHuEpo) is used to treat different types of anemia, not only in uremic patients but also in newborns with anemia of prematurity, in patients with cancer-related anemia or myeloproliferative disease, thalassemias, bone marrow transplants, or those with chronic infectious diseases. The pleiotropic functions of Epo are well known. It has been shown that this hormone can modulate the inflammatory and immune response, has direct hemodynamic and vasoactive effects, could be considered a proangiogenic factor because of its interaction with vascular endothelial growth factor, and its ability to stimulate mitosis and motility of endothelial cells. The multifunctional role of Epo has further been confirmed by the discovery in the central nervous system of a specific Epo/Epo receptor (EpoR) system. Both Epo and EpoR are expressed by astrocytes and neurons and Epo is present in the cerebrospinal fluid (CSF). Therefore, novel functions of Epo, tissue-specific regulation, and the mechanisms of action have been investigated. In this review we have tried to summarize the current data on the role of Epo on brain function. We discuss the different sites of cerebral expression and mechanisms of regulation of Epo and its receptor and its role in the development and maturation of the brain. Second, we discuss the neurotrophic and neuroprotective function of Epo in different conditions of neuronal damage, such as hypoxia, cerebral ischemia, and subarachnoid hemorrhage, and the consequent possibility that rHuEpo therapy could soon be used in clinical practice to limit neuronal damage induced by these diseases.

Cold pressor test raises serum concentrations of ICAM-1, VCAM-1, and E-selectin in normotensive and hypertensive patients.

In patients with essential hypertension, elevated soluble E-selectin (sE-selectin) levels may indicate endothelial cell injury or activation. We therefore sought to ascertain whether arterial blood pressure increased by the cold pressor test can modify serum concentrations of sE-selectin and other soluble forms of adhesion molecules, such as soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1), or the expression of any adhesion molecules in circulating monocytes and lymphocytes. Our findings show that levels of sE-selectin, sVCAM-1, and sICAM-1 are higher in patients with essential hypertension than in normotensive subjects (sICAM-1, 380 +/- 52 versus 262 +/- 96 ng/mL, P<.05; sVCAM-1, 720 +/- 52 versus 625 +/- 38 ng/mL, P<.05; and sE-selectin, 75 +/- 21 versus 61 +/- 22 ng/mL, P<.05). Furthermore, in normotensive and hypertensive patients, the cold pressor test caused an increase in serum concentrations of sICAM-1, sVCAM-1, and sE-selectin, but it did not cause changes in the expression of adhesion molecules in circulating monocytes and lymphocytes. High arterial blood pressure may therefore increase the production of serum adhesion molecules, probably through endothelial activation.

Effect of fluvastatin on proteinuria in patients with immunoglobulin A nephropathy.

3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors are established drugs for the treatment of hypercholesterolemia, but several studies have shown that benefits obtained with these drugs are not causally related only to regression of cholesterol lowering. Moreover, in experimental models of progressive renal disease, statins have reduced the extent of glomerulosclerosis. This study evaluated the antiproteinuric effect of a daily dose of 40 mg fluvastatin for 6 months in moderately proteinuric patients with immunoglobulin A nephropathy, stable renal function, and no indicators of poor long-term prognosis. The effects of therapy were evaluated on the basis of 24-hour proteinuria (total proteinuria and albuminuria), albuminemia, creatinine clearance, cholesterol, and triglyceride values. Renal function remained stable in all patients. A significant decrease in proteinuria was observed after 6 months of therapy and persisted for all the observations. An increase in serum albumin was observed after 6 months of therapy. This study suggests that there is an antiproteinuric effect of HMG-CoA reductase inhibitors in moderately proteinuric patients with immunoglobulin A nephropathy.

Effects of lisinopril administration on blood bcl-2 concentrations in patients with immunoglobulin A nephropathy.

We evaluated blood concentrations of bcl-2, a proto-oncogene that can inhibit apoptotic phenomena, in a group of patients with immunoglobulin A (IgA) nephropathy. Concentrations of bcl-2 were higher in patients with proteinuria than in those without proteinuria. A 6-month course of 5 mg/day lisinopril given to subjects with proteinuria significantly reduced blood bcl-2 concentrations and caused a reduction in proteinuria. Therefore increased blood bcl-2 concentrations may be considered an index of risk in subjects with IgA nephropathy, and the positive effects of angiotensin-converting enzyme inhibitors on proteinuria in patients with IgA nephropathy may be attributed, at least in part, to their effect on the mechanisms that regulate apoptosis. This is of fundamental importance in resolving glomerular hypercellularity in the course of glomerulonephritis.

Effects of long-lasting raloxifene treatment on serum prolactin and gonadotropin levels in postmenopausal women.

OBJECTIVE: To evaluate the effects of a 6 month administration of raloxifene hydrochloride, a selective estrogen receptor modulator which was recently approved for the prevention of osteoporosis, on serum gonadotropin and prolactin (PRL) levels and on TRH-stimulated PRL responsiveness in postmenopausal women who have not undergone estrogen replacement therapy. DESIGN AND METHODS: Sixteen healthy postmenopausal women were divided into two groups on the basis of their bone status, evaluated by dual energy X-ray absorptiometry at the lumbar level. Eight women (chronological age 52.4+/-4.1 (s.d.) years, menopausal age 42.4+/-3.9 years), in whom T-score L2-L4 was less than -2.5 s.d., were treated with raloxifene (60 mg p.o.) administered daily for 6 months (group 1), while the other eight women (chronological age 52.6+/-2.5 years, menopausal age 42.1+/-3.6 years), in whom the T-score L2-L4 ranged between -1 and -2.5 s.d., were used as a control group (group 2). Serum PRL, FSH, LH and 17beta-estradiol (E2) levels were evaluated at baseline and after 3 and 6 months of treatment. In all subjects, PRL responsiveness to TRH (200 microg i.v.) administration was evaluated at baseline and at the end of the study. RESULTS: At baseline, mean PRL, LH and FSH levels were not significantly different in the two groups (PRL 133.6+/-21.7 vs 136.7+/-28.1 mIU/l (NS), LH 25.1+/-6.8 vs 24.4+/-6.7 mIU/ml (NS), FSH 74.4+/-25.0 vs 71.1+/-24.1 mIU/ml (NS), in group 1 and group 2 respectively). No significant variations in serum FSH and LH values, in either group, or in serum PRL levels in group 2, were observed at the 3 and 6 month examinations. On the contrary, serum PRL values decreased significantly in group 1 after 3 months (100.1+/-47.7 mIU/l, P<0.05) and 6 months (81.5+/-30.2 mIU/l, P<0.001). At baseline, no significant differences were observed in the TRH-stimulated serum PRL peak between the groups (1015.4+/-30.5 vs 1030.2+/-25.7 mIU/l in group 1 and in group 2 respectively), while it decreased significantly at the 6 month examination in group 1 (770.5+/-47.4 mIU/l, P<0.001) and it was significantly lower than in group 2 (1068.1+/-301.8 mIU/l, P=0.02). Serum E2 was not detected at baseline and at each examination, in all patients. CONCLUSIONS: The decrease of PRL values induced by long-term raloxifene administration in postmenopausal women could be explained by a direct antiestrogenic effect of raloxifene on lactotrope cells or by the recently suggested increase of opiatergic tone on the hypothalamic-pituitary region.

No changes of peripheral insulin resistance in polycystic ovary syndrome after long-term reduction of endogenous androgens with leuprolide.

The aim of this study was to investigate the relationship between plasma insulin levels, peripheral insulin sensitivity and androgen secretion in ten patients with polycystic ovary syndrome and in six obese women as compared with six normal-weight control subjects. During a euglycemic-hyper-insulinemic clamp no significant change of testosterone, androstenedione or dehydroepiandrosterone sulfate plasma levels was observed in the two groups of patients or in the control subjects; insulin sensitivity was clearly reduced and was similar in polycystic ovary patients and in obese women, in spite of the different plasma androgen levels. A long-term (5 months) androgen suppression with the gonadotropin-releasing hormone agonist leuprolide was not able to improve significantly the insulin sensitivity. These results demonstrate that the short-term hyperinsulinemia achieved with the clamp technique does not affect androgen secretion and that insulin resistance, measured with the same technique, is not influenced by long-term suppression of plasma androgen levels in polycystic ovary syndrome.

Metabolic effects of dehydroepiandrosterone replacement therapy in postmenopausal women.

OBJECTIVE: To investigate whether long-term treatment with dehydroepiandrosterone (DHEA) in postmenopausal women can modify insulin sensitivity and plasma lipid profile. DESIGN AND METHODS: Twenty healthy postmenopausal women with serum dehydroepiandrosterone sulfate (DHEA-S) concentrations <2.5 micromol/l were enrolled and randomly assigned to two different treatment groups: group 1 were treated with micronized DHEA, 25 mg/day at 0800 h for 12 months; group 2 were treated with an identical placebo tablet. At the beginning and at the end of the study, plasma lipid profile, glucose tolerance (oral glucose tolerance test) and insulin sensitivity (euglycemic hyperinsulinemic clamp: M index) were assessed. RESULTS: After 12 months, the group treated with DHEA showed a considerable improvement of insulin sensitivity (M index +29.55%, P=0.01) and lipid pattern (high-density lipoprotein cholesterol +11.61%, P=0.03; low-density lipoprotein cholesterol -11.07%, P=0.04; triglycerides -19.60%, P=0.03), but glucose tolerance did not change. No modifications were observed in the placebo group. CONCLUSIONS: Long-term treatment with DHEA ameliorates some metabolic parameters that are linked to increased cardiovascular risk and, consequently, this seems to be an interesting therapeutic tool in the management of the postmenopausal syndrome.

Cardiovascular remodeling, apoptosis, and drugs.

Apoptosis, a form of programmed cell death, mediates the controlled deletion of so-called "unwanted" cells. This review deals with the key features of this cell death program, showing that apoptosis is regulated by factors extrinsic and intrinsic to the dying cell. The elucidation of the possible interactions between these factors may be of major interest in preventing the progression to cardiovascular remodeling in patients with hypertensive disease. New pathways of research are emerging for drugs, such as beta-blockers, ACE inhibitors, the calcium-antagonists, and the receptor antagonist of angiotensin II, all of which have beneficial effects on cardiovascular remodeling. This may be due to the direct effect of these drugs on the cell proliferation/apoptosis balance.

Reduced bcl-2 concentrations in hypertensive patients after lisinopril or nifedipine administration.

In 30 patients with essential hypertension and 30 healthy control subjects, we evaluated blood concentrations of B cell leukemia-2 (bcl-2), a protooncogene that can reduce apoptosis. Bcl-2 concentrations were higher in hypertensive than in normotensive subjects. The increase in pressure due to a cold pressor test caused a further increase in blood bcl-2 concentrations, in both hypertensive and normotensive subjects. Treatment of hypertensive patients with hypotensive drugs caused a reduction in bcl-2 concentrations, which was more marked after administration of lisinopril than of nifedipine. The results suggest that concentrations of bcl-2 are increased in patients with hypertension, which could be an important factor in cell proliferation underlying posthypertensive vascular remodeling. Moreover, lisinopril and nifedipine appear to be capable of reducing bcl-2 concentrations, with potentially beneficial effects on vascular modifications in patients with hypertension.

Translocation (5;10)(q13;q26) in acute monoblastic leukemia.

We report a case of acute monoblastic leukemia [French-American-British (FAB) M5a] observed in a 38-year-old man and associated at diagnosis with a t(5;10)(q13;q26) found in cells from a bone marrow culture. The patient survived only 2 months after diagnosis. t(5;10) as a solitary chromosome abnormality has not been previously reported in M5a, and, in the case that we describe, it appears to be correlated with a poor prognosis.

Erythropoietin protects against brain ischemic injury by inhibition of nitric oxide formation.

Erythropoietin prevents in vitro glutamate-induced neuronal death and could play a role in the central nervous system. We investigated the in vivo effects of recombinant human erythropoietin after intraperitoneal (i.p.; 25-100 U) or intracerebroventricular (i.c.v.; 0.25-25 U) administration on survival, brain malonildialdehyde (MDA) levels, brain edema, hippocampal neuronal death and brain nitric oxide (NO) synthesis after bilateral carotid occlusion (5 min), followed by reperfusion in the Mongolian gerbil. Peripheral posttreatment with recombinant human erythropoietin reduced postischemic MDA levels, brain edema and increased survival. Either peripheral or i.c.v. posttreatment with recombinant human erythropoietin significantly reduced hippocampal CA1 neuronal loss, observed 7 days after the ischemic event. Increase of nitrite and nitrate (as an index of NO formation) in the hippocampus, as observed after ischemia, was reduced in animals treated with recombinant human erythropoietin. These data suggest that in vivo recombinant human erythropoietin effects on brain ischemic injury could be due to inhibition of NO overproduction.

In vivo evidence that erythropoietin has a neuroprotective effect during subarachnoid hemorrhage.

To ascertain in vivo whether recombinant human erythropoietin has a neuroprotective effect on the cortex during subarachnoid hemorrhage, 56 rabbits were divided into the following groups: Group 1 control sham operated plus placebo (n=14; saline solution - NaCl 0.9%); Group 2 control sham operated plus recombinant human erythropoietin (n=14); Group 3 subarachnoid hemorrhage plus placebo (n=14); Group 4 subarachnoid hemorrhage plus recombinant human erythropoietin (n=14; intraperitoneal administration of recombinant human erythropoietin immediately after inducing subarachnoid hemorrhage). In none of the Groups 1 and 2 animals was subarachnoid hemorrhage induced. In Group 3 rabbits, an increase in locomotor activity (open field apparatus) was observed 24, 48 and 72 h after surgery, and the mortality rate was 42.9% within 72 h after surgery, and, no increase in locomotor activity was observed in Group 4 rabbits, which survived for at least 72 h. Our findings suggest that recombinant human erythropoietin may be of benefit in the treatment of subarachnoid hemorrhage.

Effects of homocysteine on proliferation, necrosis, and apoptosis of vascular smooth muscle cells in culture and influence of folic acid.

BACKGROUND: It is known that hyperhomocysteinemia is associated with an increased risk of vascular disease, yet little is known about the pathogenic mechanisms underlying the action of homocysteine (Hcy) itself. METHODS: We evaluated the effects of Hcy on cell proliferation, apoptosis, and necrosis in smooth muscle cells (SMCs) cultured for 24 h with different amounts of Hcy. The percentage of apoptotic and necrotic cells from the culture was evaluated using two different techniques: annexin V-FITC and propidium iodide (PI) fluorescence and apoptosis TUNEL assay. RESULTS: The addition of 10 microM/l of Hcy to the medium was followed by a significant increase in cell proliferation and death, through apoptosis and necrosis, respectively. Notwithstanding this apparent balance, a significant increase was found in the total number of cells present in Hcy-treated culture, thus demonstrating a positive dose-dependent correlation with Hcy concentrations in the culture medium. The addition of folic acid to the culture medium significantly reduced both Hcy concentrations in media and the effects of Hcy on the proliferation/apoptosis/necrosis balance of cells in culture. The percentages for apoptotic cells and for cells with a necrotic morphology continued to increase as Hcy concentrations increased, although the absolute values were lower in the culture treated than in that not treated with folic acid. CONCLUSIONS: In the presence of folic acid, at increasing concentrations of Hcy, the total number of cells in culture showed increases far less relevant with respect to the control. Also the percentage of apoptotic cells to that of cells with a necrotic morphology, although conserving the tendency to increase to growth of the concentrations of Hcy, have shown absolute values that were lower in the folic acid-treated cultures.