Lymphocyte DNA damage in patients with acute coronary syndrome and its relationship with severity of acute coronary syndrome

OBJECTIVE: The aim of this study was to investigate the association between lymphocyte DNA damage and acute coronary syndromes (ACS). METHODS: The study population contained 53 patients with ACS, 48 patients with stable angina and 35 voluntary healty subjects. DNA damage was assessed by alkaline comed assay in peripheral lymphocyte and plasma levels of total antioxidant capacity (TAC) were determined using a novel automated measurement method. RESULTS: In ACS patients, DNA damage was significantly higher than in patients with stable angina and control subjects (144+/-52 AU, 116+/-37, 68+/-34 AU; for three p<0.001, respectively). The TAC levels in patients with ACS were lower than the other groups (1.24+/-0.31 mmol Trolox equiv./l, 1.46+/-0.29 mmol Trolox equiv./l, p<0.05, respectively). DNA damage values in patients with acute miyocardial infarction were significantly higher than in patients with unstable angina (159.8+/-53.0 AU versus 131.8+/-48.4 AU; p<0.05, respectively). Lymphocyte DNA damage values in patients with ACS showed positive correlation with d-dimer (r=0.880, p<0.001) troponin I (r=538, p<0.001) and C-reactive protein (r=0.544, p<0.001) and negative correlation with TAC (r=-0.346, p=0.011). In multiple linear regression analysis, TAC (beta=-0.213, p=0.001) and d-dimer (beta=0.697, p<0.001) were independent predictors of DNA damage in patients with ACS. CONCLUSIONS:These findings indicate that lymphocyte DNA damage level increases in patients with ACS. Elevated DNA damage may be related with plaque instability and be useful for the identification of patients with acute coronary syndromes.     

Simvastatin treatment prevents oxidative damage to DNA in whole blood leukocytes of dyslipidemic type 2 diabetic patients

Type 2 diabetes (T2D) is associated with increased oxidative stress as indicated by elevated levels of lipid peroxidation and protein oxidation products. Since reactive oxygen species (ROS) can cause damage to biological macromolecules including DNA, this study investigated oxidative damage to DNA using the alkaline (pH > 13) comet assay in peripheral whole blood leukocytes sampled from 15 dyslipidemic T2D patients treated with simvastatin (20 mg/day), 15 dyslipidemic T2D patients not treated with simvastatin, 20 non‐dyslipidemic T2D patients, and 20 healthy individuals (controls). Our results showed a greater DNA migration in terms of damage index (DI) (p < 0.01) in the dyslipidemic T2D patients not treated with statin (DI = 67.70 ± 10.89) when compared to the dyslipidemic T2D patients under statin treatment (DI = 47.56 ± 7.02), non‐dyslipidemic T2D patients (DI = 52.25 ± 9.14), and controls (DI = 13.20 ± 6.40). Plasma malondialdehyde (MDA) and C‐reactive protein (CRP) levels were also increased and total antioxidant reactivity (TAR) and paraoxonase activity (PON1) decreased in non‐dyslipidemic T2D patients and dyslipidemic T2D non‐treated with simvastatin. We also found that DI was inversely correlated with TAR (r = ?0.61, p < 0.05) and PON1 (r = ?0.67, p < 0.01). In addition, there was a significant positive correlation between DI and CRP (r = 0.80, p < 0.01). Our results therefore indicate that simvastatin treatment plays a protective role on oxidative damage to DNA in dyslipidemic T2D patients probably reflecting a general decrease in oxidative stress in these patients. Copyright © 2010 John Wiley & Sons, Ltd.     

Increased DNA damage in patients with complete hydatidiform mole

The pathologic mechanisms underlying the gestational trophoblastic diseases are largely unexplored, but are thought to involve oxidative damage to the maternal vasculature and also to the placenta. In this study we have assessed the plasma levels of total antioxidant response (TAR) and the levels of endogenous DNA damage--determined by the comet assay--in peripheral blood lymphocytes from 13 women with complete hydatidiform mole (CHM) and compared these with those of 12 healthy pregnant controls and 10 healthy non-pregnant controls. Significantly lower mean levels of plasma TAR were found in patients with CHM compared with healthy pregnant controls (1.08+/-0.29 versus 1.17+/-0.14 mmol Trolox Eq/L, p<0.05) and with healthy non-pregnant controls (1.08+/-0.29 versus 1.38+/-0.12 mmol Trolox Eq/L, p<0.05). Significantly higher mean levels of endogenous DNA damage were observed in patients with CHM than in healthy pregnant controls (234.5+/-50.74 versus 125.7+/-45.4 AU, p<0.05) or in healthy non-pregnant controls (234.5+/-50.74 versus 104.0+/-49.6 AU, p<0.05). We observed an inverse correlation between the plasma TAR and the levels of endogenous DNA damage (r=-0.64, p<0.01), in that the levels of oxidative damage to the DNA were found to parallel the decrease in the plasma TAR in the CHM group. These results reveal a relationship between the extracellular and intracellular (as reflected by damage to the DNA) levels of oxidation. Our observations suggest that there is a link between the increased levels of oxidative stress and the increase in endogenous DNA damage seen in patients with CHM, as compared with those seen in normal pregnancy. However, the nature of this link, and whether it is direct or indirect, remains to be explored.     

Relationship between DNA damage, total antioxidant capacity and coronary artery disease

OBJECTIVE: It has been known that there was a relation between the levels of DNA damage and the severity of the coronary artery disease (CAD). However, little is known about association of DNA damage with total antioxidant capacity (TAC) and CAD. The aim of this study is to evaluate the relationship between DNA damage, TAC and CAD. METHODS: We used the comet assay to measure DNA damage from 53 patients with angiographically documented CAD and 42 patients with angiographically documented normal coronary vessel. The extent and severity of CAD was calculated to Gensini score index. TAC of plasma was determined using a novel automated measurement method. RESULTS: Mean values of DNA damage were significantly higher in CAD patients than in the control group (p<0.001). There was a positive correlation between Gensini score index and DNA damage (r=0.590, p<0.001). Additionally, significantly positive correlations between score of DNA damage, and diabetes, smoking, obesity and hyperlipidemia were found (p<0.05). There was also a negative correlation between TAC and DNA damage (r=-0.711, p<0.001). The DNA damage was significantly higher in diabetic, smoker, hyperlipidemic and obese individuals than those without these conditions (p=0.001, p=0.006, p=0.001, p=0.004, respectively). CONCLUSIONS: These data indicate that the level of DNA damage is increased and TAC level is decreased in CAD. DNA damage is correlated with the severity of the CAD, and levels of TAC.     

Oxidative DNA damage and augmentation of poly(ADP-ribose) polymerase/nuclear factor-kappa B signaling in patients with type 2 diabetes and microangiopathy

Although oxidative stress and the subsequent DNA damage is one of the obligatory signals for poly(ADP-ribose) polymerase (PARP) activation and nuclear factor-kappa B (NFkappaB) alterations, these molecular aspects have not been collectively examined in epidemiological and clinical settings. Therefore, this study attempts to assess the oxidative DNA damage and its downstream effector signals in peripheral blood lymphocytes from Type 2 diabetes subjects without and with microangiopathy along with age-matched non-diabetic subjects. The basal DNA damage, lipid peroxidation and protein carbonyl content were significantly (p<0.05) higher in patients with and without microangiopathy compared to control subjects. Formamido Pyrimidine Glycosylase (FPG)-sensitive DNA strand breaks which represents reliable indicator of oxidative DNA damage were also significantly (p<0.001) higher in diabetic patients with (19.41+/-2.5) and without microangiopathy (16.53+/-2.0) compared to control subjects (1.38+/-0.85). Oxidative DNA damage was significantly correlated to poor glycemic control. PARP mRNA expression and PARP activity were significantly (p<0.05) increased in cells from diabetic patients with (0.31+/-0.03 densitometry units; 0.22+/-0.02PARPunits/mgprotein, respectively) and without (0.35+/-0.02; 0.42+/-0.05) microangiopathy compared to control (0.19+/-0.02; 0.11+/-0.02) subjects. Diabetic subjects with and without microangiopathy exhibited a significantly (p<0.05) higher (80%) NFkappaB binding activity compared to control subjects. In diabetic patients, FPG-sensitive DNA strand breaks correlated positively with PARP gene expression, PARP activity and NFkappaB binding activity. This study provides a comprehensive molecular evidence for increased oxidative stress and genomic instability in Type 2 diabetic subjects even prior to vascular pathology and hence reveals a window of opportunity for early therapeutic intervention.     

Lymphocyte DNA damage and oxidative stress in patients with iron deficiency anemia

Oxidant stress has been shown to play an important role in the pathogenesis of iron deficiency anemia. The aim of this study was to investigate the association between lymphocyte DNA damage, total antioxidant capacity and the degree of anemia in patients with iron deficiency anemia. Twenty-two female with iron deficiency anemia and 22 healthy females were enrolled in the study. Peripheral DNA damage was assessed using alkaline comet assay and plasma total antioxidant capacity was determined using an automated measurement method. Lymphocyte DNA damage of patients with iron deficiency anemia was significantly higher than controls (p<0.05), while total antioxidant capacity was significantly lower (p<0.001). While there was a positive correlation between total antioxidant capacity and hemoglobin levels (r=0.706, p<0.001), both total antioxidant capacity and hemoglobin levels were negatively correlated with DNA damage (r=-0.330, p<0.05 and r=-0.323, p<0.05, respectively). In conclusion, both oxidative stress and DNA damage are increased in IDA patients. Increased oxidative stress seems as an important factor that inducing DNA damage in those IDA patients. The relationships of oxidative stress and DNA damage with the severity of anemia suggest that both oxidative stress and DNA damage may, in part, have a role in the pathogenesis of IDA.     

Oxidative DNA damage in rats exposed to extremely low frequency electro magnetic fields

Extremely low frequency (ELF) electromagnetic field (EMF) is thought to prolong the life of free radicals and can act as a promoter or co-promoter of cancer. 8-hydroxy-2'-deoxyguanosine (8OHdG) is one of the predominant forms of radical-induced lesions to DNA and is a potential tool to asses the cancer risk. We examined the effects of extremely low frequency electro magnetic field (ELF-EMF) (50 Hz, 0.97 mT) on 8OHdG levels in DNA and thiobarbituric acid reactive substances (TBARS) in plasma. To examine the possible time-dependent changes resulting from magnetic field, 8OHdG and TBARS were quantitated at 50 and 100 days. Our results showed that the exposure to ELF-EMF induced oxidative DNA damage and lipid peroxidation (LPO). The 8OHdG levels of exposed group (4.39+/-0.88 and 5.29+/-1.16 8OHdG/dG.10(5), respectively) were significantly higher than sham group at 50 and 100 days (3.02+/-0.63 and 3.46+/-0.38 8OHdG/dG.10(5)) (p<0.001, p<0.001). The higher TBARS levels were also detected in the exposure group both on 50 and 100 days (p<0.001, p<0.001). In addition, the extent of DNA damage and LPO would depend on the exposure time (p<0.05 and p<0.05). Our data may have important implications for the long-term exposure to ELF-EMF which may cause oxidative DNA damage.     

Effects of hydroxyurea and aphidicolin on phosphorylation of ataxia telangiectasia mutated on Ser 1981 and histone H2AX on Ser 139 in relation to cell cycle phase and induction of apoptosis.

BACKGROUND: DNA replication stress often induces DNA damage. The antitumor drug hydroxyurea (HU), a potent inhibitor of ribonucleotide reductase that halts DNA replication through its effects on cellular deoxynucleotide pools, was shown to damage DNA inducing double-strand breaks (DSBs). Aphidicolin (APH), an inhibitor of alpha-like DNA polymerases, was also reported to cause DNA damage, but the evidence for induction of DSBs by APH is not straightforward. Histone H2AX is phosphorylated on Ser 139 in response to DSBs and one of the protein kinases that phosphorylate H2AX is ataxia telangiectasia mutated (ATM); activation of ATM is through its phosphorylation of Ser 1981. The present study was undertaken to reveal whether H2AX is phosphorylated in cells exposed to HU or APH and whether its phosphorylation is mediated by ATM. MATERIALS AND METHODS: HL-60 cells were treated in cultures with 0.1-5.0 mM HU or 1-4 muM APH for up to 5 h. Activation of ATM and H2AX phosphorylation was detected immunocytochemically using Ab specific to Ser1981-ATM or Ser 139-H2AX epitopes, respectively, concurrent with measurement of cellular DNA content. RESULTS: While exposure of cells to HU led to H2AX phosphorylation selectively during S phase and the cells progressing through the early portion of S (DI = 1.1-1.4) were more affected than late-S phase (DI = 1.6-1.9) cells, ATM was not activated by HU. In fact, the level of constitutive ("programmed") ATM phosphorylation was distinctly suppressed, in all phases of the cell cycle, at 0.1-5.0 mM HU. Cells' exposure to APH also resulted in H2AX phosphorylation at Ser139 with no evidence of ATM activation, and as in the case of HU, the early-S cells were more affected than the late-S phase cells. The rise in frequency of apoptotic cells became apparent after 2 h of exposure to HU or APH, and all apoptotic cells had markedly elevated levels of both H2AX-Ser139 and ATM-Ser1981 phosphorylation. CONCLUSIONS: The lack of correlation between H2AX phosphorylation and ATM activation indicates that protein kinase(s) other than ATM (ATR and/or DNA-dependent protein kinase) are activated by DSBs induced by replication stress. Interestingly, HU inhibits the constitutive ("programmed") level of ATM phosphorylation in untreated cells. However, DNA fragmentation during apoptosis activates ATM and dramatically increases level of H2AX phosphorylation.     

Soluble tumour necrosis factor-alpha receptor type 1 as a biomarker of response to phototherapy in patients with psoriasis.

The purpose of the study was to analyze the relationship between the serum concentration of soluble tumour necrosis factor-alpha type 1 (sTNF-R1), the severity of plaque-type psoriasis and therapeutic response. We compared sTNF-R1 in 25 patients treated with narrowband ultraviolet B (NB-UVB) radiation and 25 patients treated with systemic photochemotherapy (psoralen plus UVA-PUVA). The pretreatment Psoriasis Area and Severity Index (PASI) score and sTNF-R1 concentration were 16.32+/-5.26 and 1.99+/-0.40 ng ml(-1), respectively, in the group treated with NB-UVB, and 17.22+/-3.48 and 2.07+/-0.31 ng ml(-1), respectively, in the group treated with PUVA. The concentration of sTNF-R1 in healthy controls was 1.49+/-0.34 ng ml(-1) (p<0.05 compared with patients with psoriasis). The pretreatment PASI score correlated with sTNF-R1 in both treatment groups (r=0.46 and r=0.44, p<0.05). NB-UVB and PUVA gave similar therapeutic effects (the PASI score after 20 treatments was 4.42+/-1.67 in the NB-UVB-treated group and 5.55+/-2.10 in PUVA-treated patients); however, the sTNF-R1 concentration at the same time differed significantly: 1.52+/-0.37 ng ml(-1) and 1.98+/-0.39 ng ml(-1) (p<0.001), respectively. Moreover, the decline in sTNF-R1 in both treatment groups was significant only in patients in whom the duration of skin lesions was less than 3 months. The results suggest that the value of serum sTNF-R1 concentration as a marker of response to phototherapy may depend on duration of skin lesions and the treatment method.     

Increased levels of C-type natriuretic peptide in patients with idiopathic left ventricular dysfunction

C-type natriuretic peptide (CNP) is expressed in the vascular endothelium. It is not known whether CNP is specifically increased in patients with idiopathic left ventricular systolic dysfunction (ILVDys) with or without overt heart failure, and whether in these patients it is related with indicators of myocardial and/or endothelial/microvascular impairment. We determined plasma CNP levels in 51 ILVDys and in 60 controls. We observed a significant increase in patients with (7.0+/-0.9 pg/ml) or without (6.1+/-0.53 pg/ml) overt heart failure (p<0.001) in respect to controls (2.5+/-0.12 pg/ml). CNP was significantly correlated with LVEF (p<0.001), end-diastolic dimension (p<0.05), ANP (p<0.001) and BNP (p<0.001), interleukin-6 (p<0.001), total cholesterol (p<0.05), low-density lipoprotein (p=0.05), ratio total cholesterol/ high-density lipoprotein (p=0.05) and, in a subgroup of patients, with abnormal vasodilating capacity of the coronary microcirculation. In conclusion, CNP is activated in patients with LV dysfunction but without coronary artery disease, independently of the presence of overt heart failure and in tune with the extent of myocardial functional involvement. In these patients CNP is also related with both systemic and coronary indicators of endothelial/microvascular damage.     

Evaluation of level of DNA damage in blood leukocytes of non-diabetic and diabetic rat exposed to cigarette smoke

The objective of the present study was to use the comet assay to evaluate the steady-state level of DNA damage in peripheral blood leukocytes from diabetic and non-diabetic female Wistar rats exposed to air or to cigarette smoke. A total of 20 rats were distributed into four experimental groups (n=5 rats/group): non-diabetic (control) and diabetic exposed to filtered air; non-diabetic and diabetic exposed to cigarette smoke. A pancreatic beta (beta)-cytotoxic agent, streptozotocin (40 mg/kg b.w.) was used to induce experimental diabetes in rats. Rats placed into whole-body exposure chambers were exposed for 30 min to filtered air (control) or to tobacco smoke generated from 10 cigarettes, twice a day, for 2 months. At the end of the 2-month exposure period, each rat was anesthetized and humanely killed to obtain blood samples for genotoxicity analysis using the alkaline comet assay. Blood leukocytes sampled from diabetic rats presented higher DNA damage values (tail moment=0.57+/-0.05; tail length=19.92+/-0.41, p<0.05) compared to control rats (tail moment=0.34+/-0.02; tail length=17.42+/-0.33). Non-diabetic (tail moment=0.43+/-0.04, p>0.05) and diabetic rats (tail moment=0.41+/-0.03, p>0.05) exposed to cigarette smoke presented non-significant increases in DNA damage levels compared to control group. In conclusion, our data show that the exposure of diabetic rats to cigarette smoke produced no additional genotoxicity in peripheral blood cells of female Wistar rats.     

Donor heart preservation with iloprost supplemented St. Thomas Hospital cardioplegic solution in isolated rat hearts

This study was designed to assess the influence of St. Thomas Hospital cardioplegic solution (St. Th.) on heart preservation in rat hearts subjected to 6h ischemia when supplemented with iloprost. In the control group (n=8), nothing was added to St. Th., whereas 10 or 1000 nmol L(-1) iloprost was added in the second (n=7) and third (n=8) groups, respectively. Mechanical contraction parameters, cardiac tissue damage and oxidative stress markers were evaluated. The 10 nmol/L iloprost group peak systolic pressure (71.0+/-30.9 versus 41.0+/-9.4 mm Hg) and -dp/dtmax (1103.8+/-94.3 versus 678.6+/-156.8 mm Hg s(-1)) were significantly higher than control group at 30 min of reperfusion (p<0.05). Iloprost supplemented groups had higher GSH and catalase levels of coronary perfusate at reperfusion, in comparison with initial values (p<0.05). AST, CK, CK-MB values increased at 0 min of reperfusion and cTnI values at 45 min of reperfusion (p<0.05) in all groups with no difference between groups. According to our results, iloprost supplementation had mild but significant improvement in postischemic values in mechanical and oxidative stress parameters, resulting in better heart preservation.     

Lymphocyte DNA damage is associated with increased aortic intima-media thickness

OBJECTIVES: To investigate whether there is an association between lymphocyte DNA damage and aortic intima-media thickness (IMT). METHODS: We studied 70 patients (mean age: 41.6+/-10 years) who underwent transesophageal echocardiography for various indications. Four different grades were determined according to intima-media thickness (IMT) of thoracic aorta measured by transesophageal echocardiography. DNA damage was assessed by alkaline single cell electrophoresis (comet) assay in peripheral lymphocytes. Plasma level of total antioxidant status (TAS) was determined by using automated measurement method. High sensitive C-reactive protein and other biochemical markers were studied in all subjects. RESULTS: The major increase in lymphocyte DNA damage was observed in patients with grade 4 IMT when compared with other groups (p<0.001, for all). Lymphocyte DNA damage of patients with grade 1 IMT was also lower than patients with grade 2 IMT (p=0.013) and patients with grade 3 IMT (p<0.001). Lymphocyte DNA damage of patients with grade 2 IMT was found at low level compared with patients with grade 3 IMT (p=0.012) as well. In multiple linear regression analysis, IMT was independently correlated with lymphocyte DNA damage (beta=0.515, p<0.001), TAS level (beta=-420, p<0.001), total cholesterol (beta=0.407, p<0.001) and LDL cholesterol level (beta=287, p=0.020). CONCLUSION: Lymphocyte DNA damage may be an independent predictor for the grade of thoracic IMT, and may play a role in pathogenesis of thoracic atherosclerosis besides TAS and cholesterol levels.     

Monitoring of early and advanced glycation in relation to the occurrence of microvascular complications in children and adolescents with type 1 diabetes mellitus

The authors aimed to evaluate if the monitoring of serum advanced glycation end-products (s-AGEs) could help to predict a development of diabetic complications. Clinical and biochemical parameters including fructosamine (FAM), glycated hemoglobin (HbA1c) and serum AGEs were investigated in children and adolescents with 1 type diabetes with (+DC) and without (-DC) complications. FAM levels (in mmol/l) were significantly elevated in +DC diabetic group compared to -DC one (3.043+/-0.459 vs. 2.614+/-0.430; p<0.001) or to controls (3.043+/-0.459 vs. 1.620+/-0.340; p<0.001) as well as in -DC compared to controls (2.614+/-0.430 vs. 1.620+/-0.340; p<0.001). HbA1c (in %) were significantly elevated in +DC diabetic group compared to -DC one (10.48+/-1.83 vs. 8.41+/-1.19; p<0.001) or to controls (10.48+/-1.83 vs. 5.0+/-0.38, p<0.001) and also in -DC compared to controls (8.41+/-1.19 vs. 5.0+/-0.38; p<0.001). Serum AGEs levels (in A. U.) were significantly higher in +DC group than in -DC (73.0+/-14.09 vs. 65.8+/-9.05; p<0.05) and in group +DC than in controls (73.0+/-14.09 vs. 60.17+/-13.78; p<0.05), whereas there was no difference between -DC and controls. FAM correlated with HbA1c in both diabetic groups (+DC: r=0.374; p<0.05; -DC: r=0.719; p<0.001), but not in controls. Serum AGEs were correlated with HbA1c (r=0.478; p=0.003) in +DC, but not in -DC or controls. Enhanced serum AGEs levels show that they could be not only an attendant phenomenon of microangiopathies, but also a predictor of their development.     

Correlation of ultrastructural changes of endothelial cells and astrocytes occurring during blood brain barrier damage after traumatic brain injury with biochemical markers of BBB leakage and inflammatory response

Focal cerebral contusion can be dynamic and expansive. It has been proved that subsequent expansive contusion is caused by brain parenchyma damage, especially BBB damage. We investigated a group of patients with traumatic brain injury. The patients (n=18) were divided into group I (n=7) of patients submitted to neurosurgery due to expansive contusion, and group II (n=11) of patients without surgery. Serum concentrations of NSE and S-100B protein were measured by electrochemiluminescence immunoassay, interleukin-6 (IL-6) was measured by chemiluminescent sequential immunometric assay and matrix metalloproteinases (MMP-9, MMP-2) were measured by immunoassays. Cortical biopsy specimens of brain were investigated by electron microscopy in patients with trauma brain injury submitted to neurosurgery. Biochemical investigation from first day up to third day after traumatic brain injury proved increased values of IL-6 (302.2+/-119.9 vs. 59.6+/-11.9 ng/l, p<0.02) and S-100B protein (3.064+/-1.064 vs. 0.649+/-0.182 microg/l, p<0.05) in patients with expansive lesion compared to patients without expansive contusion. Significantly higher levels of MMP-9 (150.4+/-28.46 vs. 74.11+/-13.16 ng/l, p<0.05) and of MMP-2 (814.5+/-126.3 vs. 523.1+/-25.28 ng/l, p<0.05) were found during first 3 days after admission in group I compared to group II. MMP-9 has also elevated in group II from lower values after admission (74.11+/-13.16 ng/l) up to high levels on the 10th day of hospitalization (225.1+/-49.35 ng/l). Ultrastructural investigation of endothelial cells and surrounded tissue revealed perivascular hemorrhage, increased pinocytic activity of endothelial cells, and cytotoxic edema of astroglial cells. Multivesical bodies were disclosed inside the endothelial cells. Higher levels of serum protein S-100B and IL-6 correlated with ultrastructural changes of endothelial cells, and with inflammatory response following TBI, respectively.     

Selenium status in psoriasis and its relationship with alcohol consumption

The aim of the study was to examine the influence of alcohol consumption on the severity of psoriasis and selenium (Se) concentration and Se-dependent gluathione peroxidase activity in plasma (pl-GSH-Px) and in erythrocytes (RBC-GSH-Px) in psoriatic patients. Thirty-five in-patients with psoriasis lasting <10 mo and 42 with psoriasis lasting >3 yr constituted groups 1 and 2, respectively. The severity of psoriasis was assessed using the PASI scoring system and the consumption of alcohol, using a structured questionnaire. The Se concentration was 47.11±11.61 μg/L in group 1 and 38.69±13.22 μg/L in group 2 (p<0.05), the pl-GSH-Px was 0.15±0.04 U/mL and 0.14±0.04 U/mL (p>0.05), and the RBC-GSH-Px was 13.97±4.27 U/g Hb and 13.16±3.85 U/g Hb (p>0.05), respectively. In excessive drinkers (<10% of patients, all males), the Se concentration was 32.84±10.88 μg/L, the pl-GSH-Px was 0.15±0.03 U/mL, and the RBC-GSH-Px was 11.64±3.32 U/g Hb. A low RBC-GSH-Px correlated to the consumption of high-grade alcoholic beverages (R=−0.45, p<0.05) and to the PASI value (R=−0.37, p<0.05) in group 2. Depressed Se concentration and Se-dependent GSH-Px can be related to the severity and a duration of psoriasis. The excessive consumption of alcohol is associated with severity of the disease and with low activity of GSH-Px in erythrocytes in patients with long-lasting psoriasis.     

Development of porcine embryos and offspring after intracytoplasmic sperm injection with liposome transfected or non-transfected sperm into in vitro matured oocytes.

The objective of this study was to evaluate in vitro and in vivo development of porcine in vitro matured (IVM) porcine oocytes fertilised by intracytoplasmic sperm injection (ICSI) and the possibility of producing transgenic embryos and offspring with this procedure. Activated ICSI oocytes had a higher pronuclear formation than non-activated ICSI oocytes (mean 64.8+/-17.3% vs 28.5+/-3.4%, p<0.05). When the zygotes with two pronuclei were cultured to day 2, there was no difference (p<0.05) in the cleavage rate (mean 60.0+/-7.0% vs 63.3+/-12.7%) between the two groups. The blastocyst rate in the activation group was significantly higher than that in the non-activation group (mean 30.0+/-11.6% vs 4.6+/-4.2%, p<0.05). After injection of the sperm transfected with DNA/liposome complex, destabilised enhanced green fluorescent protein (d2EGFP) expression was not observed on day 2 in either cleaved or uncleaved embryos. But from day 3, some of the embryos at the 2-cell to 4-cell stage started to express d2EGFP. On day 7, about 30% of cleaved embryos, which were in the range of 2-cell to blastocyst stage, expressed d2EGFP. However, for the IVF oocytes inseminated with sperm transfected with DNA/liposome complex, and for oocytes injected with sperm transfected with DNA/liposome complex, and for oocytes injected with DNA/liposome complex following insemination with sperm not treated with DNA/liposome complex, none of the embryos expressed d2EGFP. Sixteen day 4 ICSI embryos derived from sperm not treated with DNA/liposome complex were transferred into a day 3 recipient. One recipient delivered a female piglet with normal birthweight. After transfer of the ICSI embryos derived from sperm transfected with DNA/liposome complex, none of the four recipients maintained pregnancy.     

Prolactin secretion in patients with idiopathic diabetes insipidus.

It has been demonstrated that hyperprolactinemia is sometimes present even in patients with idiopathic diabetes insipidus (DI). In this study, we examined the responses of serum prolactin (PRL) to hypertonic saline infusion and TRH injection in 11 patients with idiopathic DI diagnosed by clinical examinations. Serum sodium in these patients (147.5 +/- 3.2 mEq/L) was significantly higher at baseline than in normal subjects (139.7 +/- 2.4 mEq/L). The plasma arginine vasopressin (AVP) level was significantly lower in DI (0.42 +/- 0.24 pg/ml) at baseline than in normal subjects (2.53 +/- 1.03 pg/ml). However, the serum PRL level in both groups did not differ significantly except in one patient with idiopathic DI (35.6 ng/ml). There was no significant correlation between the basal serum sodium and basal serum PRL in either group. After an infusion of hypertonic saline, the serum sodium level gradually increased to 155.6 +/- 3.4 mEq/L in DI and to 146.5 +/- 4.3 mEq/L in the normal subjects. However, this increase did not affect PRL secretion in either group. PRL response to TRH was essentially normal in all patients with idiopathic DI. These results indicate that the secretion of PRL is not generally affected by chronic mild hypernatremic hypovolemia in the patients with idiopathic DI.     

The isolation of reticulocyte-free human red blood cells

We depleted reticulocytes from erythrocytes of both sickle cell disease (SCD) subjects and healthy controls by four methods: fluorescence-activated cell sorting (FACS), Miltenyi immunomagnetic depletion (MACS), a combination of these methods (FACS + MACS) and Percoll density separation. The efficiency of these methods was assessed by new methylene blue staining and manual enumeration of the reticulocytes. FACS sorted erythrocytes from reticulocytes based on size and granularity, as well as the absence of dsDNA staining. MACS depleted reticulocytes from erythrocytes based on the immunoaffinity to CD36 and CD71. Reticulocytes from healthy controls were depleted to 相似文献   

The comparison of insulin sensitivity in non-diabetic hemodialysis patients treated with and without recombinant human erythropoietin.

BACKGROUND: Patients with end-stage renal disease (ESRD) are known to have insulin resistance. Treatment with EPO is associated with improvement in insulin sensitivity in uremic patients. The aim of this study was to compare insulin sensitivity and pancreatic B cell function in adult non-diabetic uremic hemodialysis patients treated with or without rHuEPO. SUBJECTS AND METHODS: Three groups of subjects were included to the study: hemodialysis patients treated with rHuEPO [EPO(+) group] or without rHuEPO [EPO(-) group], and healthy controls. Anthropometrical parameters, lipid levels, fasting glucose and insulin levels were measured in all subjects. Homeostasis Model Assessment (HOMA) was used to compare insulin sensitivity. ANOVA, independent t-test, and Pearson correlation were used for statistical analysis. RESULTS: Mean insulin level of control group (20.04 +/- 7.2 pmol/l) was significantly lower than EPO(+) group (p < 0.04) and EPO(-) group (p < 0.0001). HOMA-(%B) levels in the EPO(+) group were significantly lower than in the EPO(-) group (106 +/- 42, 140 +/- 63 respectively, p < 0.02). HOMA-(%B) levels in the control group (66 +/- 17) were significantly lower than in the EPO(+) and EPO(-) group (p < 0.005 and p < 0.0001 respectively). HOMA-(%S) levels in the EPO(+) groups was significantly higher than in the EPO(-) group (91 +/- 40, 56 +/- 26, respectively; p < 0.01). HOMA-(%S) levels of control group (125 +/- 24 ) was significantly higher than EPO(+) and EPO(-) groups (p < 0.02, p < 0.0001 respectively). We found a positive correlation between duration of erythropoietin treatment and insulin sensitivity (r = 0.484, p < 0.002). CONCLUSIONS: Firstly, patients treated with EPO are insulin sensitive compared to patients not treated with EPO. Secondly, duration of erythropoietin treatment is positively correlated with insulin sensitivity in hemodialysis patients.