Volume 24, Issue 3-4

Pro-inflammatory cytokines like interferon-γ (IFN-γ) play dominant roles in pathophysiologic conditions like infections, cardiovascular and neurodegenerative disorders and autoimmune syndromes. As part of its antimicrobial and immunomodulatory armature, the tryptophan-degrading enzyme indoleamine (2,3)-dioxygenase (IDO) is mainly up-regulated in dendritic cells (DCs) and phagocytes by pro-inflammatory stimuli, most notably IFN-γ. By the breakdown of the essential amino acid l -tryptophan along the kynurenine pathway, IDO plays a key role in the inhibition of cell proliferation including that of activated T cells, thereby supporting immune tolerance in mammalian pregnancy, tumor development, allergic inflammation and allotransplantation. IFN-γ-induced tryptophan deprivation also seems to be involved in the pathogenesis of anemia and cachexia when erythroid progenitor cells suffer from insufficient amino acid supply or when protein biosynthesis of the organism is restricted by diminished tryptophan availability. This biochemical cascade seems also to be involved in the production of potentially neurotoxic tryptophan catabolites such as quinolinic acid, which ultimately leads to the development of neuropsychiatric symptoms like cognitive impairment and depression especially in patients suffering from severe and chronic infections.

The need for sufficient quantities of folic acid for normal embryogenesis and fetal development is well known. Women of childbearing age must be sure to have an adequate intake of folic acid periconceptionally prior to and during pregnancy. Folate plays an essential role in the biosynthesis of RNA and DNA, phospholipid and neurostransmitters synthesis, using S-adenosylmethionine (SAM) as the primary methyl group donor. In some countries, the knowledge that folic acid is preventable for the reduction of the risk for neural tube defects (NTD) and in lowering the high amount of homocysteine (Hcy) led to the recommendation that all women capable of becoming pregnant should consume 400 µg of folic acid daily. Later, in 1998, folic acid fortification of all enriched cereal grain product flour was implemented in some countries. In addition to careful monitoring of adverse effects, the studies are warranted in order to determine the potentially positive and deleterious effects of folic acid fortification and supplementation on the human health or human diseases. To further understand the metabolic action of folates we asked ourselves the question: Is the intake of a sufficient amount of this vitamin a benefit or risk for human health?

The effects of cytokine and mitogen stimulation on homocysteine (HCY) metabolism in different cells were investigated: in human dermal microvascular endothelial cells (HDMEC), T lymphocytes, mature and immature dendritic cells, and myelomonocytic (THP-1) and monocytic cell lines (U-937). Furthermore, the influence of supplementation of cells with folate acid, methionine and the combination of both on HCY metabolism was investigated. Unstimulated HDMEC and dendritic cells only produced very little amounts of HCY, and stimulation did not change HCY formation significantly either. However, higher HCY concentrations were detected in HDMEC and dendritic cells under supplementation with methionine and slightly less under supplementation with folate. Proliferating T lymphocytes showed an increase in HCY production on stimulation with increasing doses of mitogens; proliferative activity was associated with HCY formation. THP-1 and U-937 cells produced significantly more HCY than endothelial cells; U-937 cells produced most HCY, which was mainly due to their high proliferation rate. Stimulation of both cell lines with lipopolysaccharide and interferon-γ, respectively, showed a significant effect on HCY production of cells; in THP-1 cells, stimulation with IFN-γ and lipopolysaccharide induced neopterin formation. Methionine supplementation strongly increased and folate supplementation slightly decreased HCY formation in both cell lines. Thus, inflammation may play a role in moderate hyperhomocysteinemia.

The objective of the present study was to extract ursolic acid (UA) from Ocimum sanctum , to synthesize its bioactive derivatives, evaluate the anti-cancer effect of its derivatives and to establish the possible mechanism of action. In the present report, we extracted UA from whole plant of O. sanctum , synthesized its novel derivatives and investigated their effect on homocysteine metabolism and dihydrofolate reductase (DHFR) activity of HepG2 cells. UA and its derivatives UA-1, UA-2 and UA-3 down-regulated DHFR activity and increased extracellular homocysteine. UA-2 showed significant anti-proliferation activity in cancer cells. Cancer cells have increased the requirement of pyrimidine base thymidylate due to rapid cell division. Thymidylate biosynthesis depends on sufficient pools of folate dependent enzymes like DHFR. In the present study, we examined the UA and its derivatives mediated perturbation of DHFR activity and extracellular homocysteine in HepG2 human hepatoma cells. After incubation with UA-2, a potent inhibition of DHFR activity was observed. Our results showed that derivatization of UA might adversely affect DHFR activity. Measurement of extracellular homocysteine indicated impaired one-carbon metabolism in cells treated with UA derivatives. In conclusion, our data suggest an anti-cancer role of UA and its derivatives via inhibition of one-carbon metabolism.

The aim of the present study was to evaluate the prevalence of the risk factor of atherosclerosis in patients with history of breast cancer. C-reactive protein, lipoprotein (a), cholesterol, high-density lipoprotein (HDL) cholesterol, low-density lipoprotein cholesterol, triglycerides, homocysteine, fibrinogen, glucose, magnesium, uric acid, and urinary albumin were determined by routine methods in 61 patients with history of breast cancer and 74 control subjects. Urinary neopterin and creatinine concentrations were measured by high-performance liquid chromatography, and intima-media thickness was determined sonographically. Breast cancer patients were significantly older and also had higher systolic blood pressure, glycosylated hemoglobin, lipoprotein (a), serum neopterin, and intima-media thickness. Serum HDL cholesterol and magnesium concentrations were significantly higher in controls compared with patients. Differential patterns of associations between the risk factors of atherosclerosis were observed in patients with history of breast cancer compared with controls. In conclusion, the prevalence of risk factors of atherosclerosis is high in patients with history of breast cancer. Differential associations between risk factors suggest possible differences in the pathogenesis of atherosclerosis in breast cancer patients and controls.

The aim of the present study was to evaluate the association between changes in peripheral blood cell count and neopterin concentrations in patients with a history of breast cancer. Peripheral blood cell count, serum ferritin, serum neopterin and urinary neopterin concentrations were determined in 61 patients with a history of breast cancer and 74 control subjects. Hemoglobin, relative lymphocyte count and absolute lymphocyte count were lower, and relative neutrophil count, ferritin and serum neopterin concentrations were significantly higher in breast cancer patients than in controls. Compared to controls, the difference in hemoglobin concentration was statistically significant only in patients with active disease. Significant negative correlations were observed between urinary neopterin and hemoglobin as well as between serum neopterin and relative lymphocyte counts in breast cancer patients. Increased ferritin concentrations were associated with a history of hypertension, but higher absolute lymphocyte counts were associated with hypertension only in subjects without history of cancer. In conclusion, in patients with a history of breast cancer, anemia is associated with disease activity and systemic immune activation. Ferritin concentrations are increased in subjects with hypertension.

End-stage renal disease (ESRD) is the inability of the kidneys to remove waste products from the blood. The most important factors causing ESRD that require hemodialysis are diabetes and hypertension. There are limited numbers of studies to evaluate tetrahydrobiopterin pathway in these patients. The aim of the study was to evaluate tetrahydrobiopterin pathway by measuring its important components, biopterin to creatinine concentrations and dihydropteridine reductase activities in diabetes and hypertension patients treated with/without hemodialysis. The patients undergoing hemodialysis were classified as diabetic nephropathy (n=21), hypertensive nephropathy (n=20) and others (n=30), while the controls consisted of healthy subjects (n=21), diabetic subjects (n=23) and hypertensive subjects (n=22) without any renal disorder. It was found that urinary biopterin to creatinine concentrations significantly increased in kidney failure patients undergoing hemodialysis compared to the healthy control group (p<0.05). Additionally, there were significant differences in urinary biopterin to creatinine concentrations between diabetes or hypertension patients and their hypertensive or diabetic control counterparts (both p<0.05). Our results indicated an alteration in tetrahydrobiopterin pathway in ESRD, and in the presence of secondary pathologies such as diabetes and hypertension in the patients undergoing hemodialysis, more considerable changes are observed in the pathway.

Tetrahydrobiopterin (BH 4 ) and 4-amino-tetrahydrobiopterin (ABH 4 ) prevent acute rejection after solid organ transplantation. Moreover, BH 4 also attenuates ischemia-reperfusion injury (IRI). The mechanisms underlying these protective effects are poorly defined. Activation of intracellular signaling proteins, including the mitogen-activated protein kinases (MAPKs) ERK, p38, and JNK, and the excessive production of mitochondrial reactive oxygen species (ROS) are observed mainly during early reperfusion. While the role of ROS in the initiation and progression of IRI is well understood, the contribution of individual signaling pathways is less clear. Here, we tested the potential effects of BH 4 and ABH 4 on MAPK activity and mitochondrial ROS levels. During hypoxia and reoxygenation (H/R), all three MAPKs were activated during early reoxygenation in cardiomyocytes and endothelial cells. p38 and JNK activation were further augmented by BH 4 and ABH 4 , whereas ERK activation was not affected. Pretreatment with BH 4 and ABH 4 reduced the basal mitochondrial ROS levels as well as the H/R-induced increase in ROS. Prolonged incubation with ABH 4 , however, showed pro-apoptotic effects in cardiomyocytes. These data suggest that a protective effect of BH 4 and ABH 4 pretreatment may be attributed mainly to their antioxidant capacity. The effects on intracellular signaling are complex and warrant further investigations.

In vitro , large amounts of neopterin are released from human monocyte-derived macrophages and dendritic cells primarily upon stimulation with Th1-type cytokine interferon-γ (IFN-γ). IFN-γ also induces the enzyme indoleamine 2,3-dioxygenase (IDO), which degrades tryptophan (TRP) to form kynurenine (KYN). IDO-mediated TRP catabolism is very effective in suppressing the proliferation of T lymphocytes as well as of pathogens in vitro and in vivo . In this study, we investigated whether exogenously added neopterin may influence IDO activity in resting and in stimulated peripheral blood mononuclear cells (PBMC). PBMC were isolated from healthy donors, and neopterin was added in a concentration range from 0.01 to 50 μmol/L. After 30 min, PBMC were stimulated or not with 10 μg/mL of mitogen phytohemagglutinin (PHA). After 48 h, culture supernatants were collected, KYN and TRP concentrations were measured by high-performance liquid chromatography, and the ratio of KYN vs. TRP was calculated as an estimate of IDO activity. Spontaneous as well as PHA-induced TRP breakdown was suppressed by exogenously added neopterin in a dose-dependent way; the lowest active concentration of neopterin was <100 nmol/L. As neopterin concentrations in the nanomolar range are commonly observed in patients suffering from infections, sepsis, or uremia, our results suggest that neopterin formation might also serve as a feedback mechanism to slow down TRP degradation in vivo .

Survival of neurons is dependent on the presence of trophic and non-trophic factors. We have previously observed the protective capacity of purine nucleosides in hypoxic neuronal cell cultures. Guanosine appeared especially interesting with respect to its remarkable neurite-stimulating aptitude. Here we report the effect of ischemic stress and guanosine on the concentration of the essential amino acids phenylalanine and tyrosine, in culture supernatants of PC12 cells. In ischemic neuronal cultures, a substantial rise of phenylalanine and tyrosine levels was observed, indicating inhibition of cellular metabolism due to neuronal stress. Elevated phenylalanine and tyrosine concentrations were reduced by the addition of guanosine. Guanosine even down-regulated the ratio of phenylalanine to tyrosine. Results may support current efforts to propagate guanosine as a neuroprotective and neuroregenerative substance.

α-Thalassemia (α-thal) is one of the most common genetic disorders in the world. It is characterized by the absence or reduced expression of α-globin genes. This study was carried out to evaluate the allelic frequency of α-thal defects in a patient for the first time in Sétif (Algeria). One hundred and two patients with hemoglobinopathies from Sétif region, Algeria, presenting thalassemia were included in this study. Genomic DNA isolation was carried out according to standard methods. For identifying the α-thal genotype, investigation of α-globin gene deletions (-α3.7, -α4.2, - MED and -α20.5) was performed by using multiplex-polymerase chain reaction (PCR). Among the three deletions found, the most mutations were the -α3.7 (10.78%), followed by the - MED (5.88%) and -α20.5 (0.98%), whereas the -α4.2 deletion was not observed (0.0%). The allele frequency is 0.054 (11/204) for the 3.7 deletion, 0.029 (6/204) for the MED and 0.005 (1/204) for the 20.5. Molecular heterogeneity of mutations is typical of α-thal in Algeria. Our findings will be valuable and essential for the molecular diagnosis and prevention strategies of hemoglobinopathy gene mutations in the Algerian population.

The aim of this study was to identify the types of abnormalities associated with neural tube defects (NTDs) and the magnitude of the risk for their expression under the influence of the following factors: maternal age >35 years, consanguinity and season of conception. One hundred and fifty fetuses were autopsied during the period 2006–2009 at the Center for Maternity and Neonatology, Tunisia. A mother’s age of >35 years increases the probability of intrauterine growth retardation by two-fold [odds ratio (OR) 2.043, confidence interval (CI) 0.880–4.741]. Consanguinity increases the relative risk for abnormalities in the facial shape (OR 3.031, CI 1.279–7.183) and adrenal hypoplasia (OR=2.787, CI 1.140–6.814). The autumn-winter period of conception increases the relative risk for the expression of cleft palate by more than nine times (OR 9.035, CI 1.161–70.258) and by about three times for abnormalities of the excretory tract (OR 2.935, CI 0.954–9.141). The prenatal ultrasound diagnosis of NTDs with risk factors such as maternal age >35 years, consanguinity and conception during the autumn-winter period should be targeted to the search for lower-than-normal fetal weight and abnormalities of the excretory tract and the adrenal glands. The head should be examined with special care for deviations in the cranial perimeter and cleft palate.