Significantly elevated Helicobacter pylori density and different genotype distribution in erosions as compared with normal gastric biopsy specimen detected by quantitative real-time PCR

BACKGROUND AND AIM: Determination of the local densities of Helicobacter pylori and its genotypic variations in gastric biopsy specimens by using novel real-time PCR-based methods could support the precise diagnosis and understanding of H. pylori infections. METHODS: Serial dilutions of H. pylori (0.016-16 microg/microl), control, bacterial, and human DNA samples were prepared. Fresh-frozen gastric biopsy specimens were taken from 103 patients, and the DNA was isolated. Quantitative determination of the ureaseA gene using hybridization probes with parallel evaluation of an internal human control gene (beta-globin) was performed by real-time PCR. CagA and VacA s1 genotypic characterizations were also performed. The data were compared with urea breath test (UBT), histology, and serological testing. RESULTS: The presence of H. pylori could be detected by ureaseA-fluorescence energy transfer (53%), UBT (51%), serological testing (48%), and histology (52%) when compared with the gold standard (54%). A significant correlation was found between the quantitative real-time ureaseA/beta-globin ratio-based H. pylori frequency and the UBT results (P<0.01). Significantly increased bacterial density was found in the erosions when compared with the healthy part of the antrum and corpus (P<0.01). Real-time PCR VacA s1 results were in significant correlation (P<0.01) with those of serological tests, but CagA results were not. The genomic profiles (VAC/GAC) were different in 13.7% of the cases, which involved three different locations in the stomach. CONCLUSION: Real-time PCR was the most reliable method for H. pylori diagnosis. Furthermore, quantification and genotyping could also be performed using this technique. The density of H. pylori was significantly increased in macroscopic erosions.     

Effects of estrogen and medroxyprogesterone acetate on subpopulations of triglyceride-rich lipoproteins and high-density lipoproteins

Hormonal replacement therapy (HRT) in postmenopausal women has been shown to increase both triglyceride (TG) and high-density lipoprotein (HDL) cholesterol levels. To better understand the effects of conjugated equine estrogen (CEE) and medroxyprogesterone acetate (MPA), the 2 most commonly prescribed hormones in HRT, on the different subpopulations of TG-rich and HDL lipoproteins, we conducted a placebo-controlled, double-blind, randomized, crossover study consisting of 3 different phases in 14 postmenopausal women. The 3 phases, each 8-week long, included: (1) placebo, (2) CEE 0.625 mg/d, and (3) CEE 0.625 mg/d and MPA 2.5 mg/d. Slight and statistically nonsignificant elevations in TG levels were observed during the CEE treatment. While very-low-density lipoprotein (VLDL) cholesterol levels were not significantly affected by CEE and CEE + MPA, both HRT treatments lowered remnant lipoprotein (RLP) cholesterol (-14% and -37%, respectively). Compared with placebo, CEE caused a significant increase in HDL, HDL(2), apolipoprotein (apo) A-I, LpAI, alpha1, and prealpha1 levels (12%, 27%, 17%, 26%, 60%, and 102%, respectively). The combination therapy blunted the CEE effect on all HDL parameters, resulting in HDL, HDL(2), and LpAI levels being no longer significantly different from placebo. Apo A-I levels and alpha1, and prealpha1 levels were still significantly higher than placebo (+11%, +50%, and +112%, respectively). These results indicate that HRT has beneficial effects on RLP levels and that, while the estrogen component of HRT has a beneficial effect on the HDL subpopulations mostly associated with coronary heart disease (CHD) protection, MPA partially inhibits this effect.     

Involvement of Oxygen-Derived Free Radicals in L-Arginine-Induced Acute Pancreatitis

This study was aimed at an assessment of the role of oxygen-derived free radicals in the pathogenesis of L-arginine (Arg)-induced acute pancreatitis in rat, by measuring the levels of malonyl dialdehyde (MDA), glutathione peroxidase (GPx), catalase, and superoxide dismutase (Mn- and Cu,Zn-SOD) in the pancreatic tissue, and evaluating the protective effect of the xanthine oxidase inhibitor allopurinol. Acute pancreatitis was induced in male Wistar rats by injecting 2 × 250 mg/100 g body weight of Arg intraperitoneally in a 1-hr interval, as a 20% solution in 0.15 M NaCl. Control rats received the same quantity of glycine. Allopurinol, 100 or 200 mg/kg, was administered subcutaneously 30 min before the first Arg injection. Rats were killed at 6, 12, 24, and 48 hr following Arg administration, and acute pancreatitis was confirmed by a serum amylase level elevation and typical inflammatory features observed microscopically. The serum level of amylase reached the peak level at 24 hr after the Arg injection (30,800 ± 3813 vs 6382 ± 184 units/liter in the control) and normalized at 48 hr. The tissue concentration of MDA was significantly elevated at 24 hr and reached the peak value at 48 hr (5.00 ± 1.75 vs 0.28 ± 0.05 nM/mg protein in the control). The catalase and Mn-SOD activities were significantly decreased throughout the study, while the GPx activity was significantly reduced at 6 and 12 hr, and the Cu,Zn-SOD activity was significantly lower at 12 hr after the Arg injection as compared with the controls. Allopurinol treatment markedly reduced the serum amylase elevation (12.631 ± 2.257 units/liter at 24 hr) and prevented the increase in tissue MDA concentration (0.55 ± 0.09 nM/mg protein at 48 hr). Both doses of allopurinol significantly ameliorated the pancreatic edema, necrosis, and inflammation at 48 hr after Arg administration. Oxygen-derived free radicals are generated at an early stage of Arg-induced acute pancreatitis. Prophylactic allopurinol treatment prevents the generation of reactive oxygen metabolites, reduces the serum amylase concentration, and exerts a beneficial effect on the development of histopathological changes.     

Prevalent diagnosed HIV in England, Wales and Northern Ireland: adjusted totals 1996 to 2001 and extrapolations to 2004

OBJECTIVE: To predict trends in diagnosed HIV prevalence by extrapolation to 2004 using data from the annual surveys of individuals receiving HIV-related care in England, Wales and Northern Ireland from 1996 to 2001. METHODS: Data from the annual surveys of prevalent HIV infections diagnosed (SOPHID) were adjusted for under-reporting and non-attendance and separately extrapolated for infections acquired homosexually, heterosexually and by other routes. The data were extrapolated using negative binomial and linear regression models based on the 1996 to 2001 annual surveys. RESULTS: The negative binomial model predicted an increase of 56% in diagnosed HIV prevalence in England, Wales and Northern Ireland between 2001 and 2004. The linear model predicted an increase of 25% for the same time period. The predicted increases are mostly driven by the large rise in the number of new diagnoses, in particular in individuals infected heterosexually. CONCLUSION: Increases in HIV prevalence in England, Wales and Northern Ireland have diverged from a linear trend. Negative binomial modelling of the data predicts that large rises in prevalence will continue during the early 2000s.     

Inhibition of EGF Signaling Protects the Diabetic Retina from Insulin-Induced Vascular Leakage

Diabetes mellitus is a disease with considerable morbidity and mortality worldwide. Breakdown of the blood–retinal barrier and leakage from the retinal vasculature leads to diabetic macular edema, an important cause of vision loss in patients with diabetes. Although epidemiologic studies and randomized clinical trials suggest that glycemic control plays a major role in the development of vascular complications of diabetes, insulin therapies for control of glucose metabolism cannot prevent long-term retinal complications. The phenomenon of temporary paradoxical worsening of diabetic macular edema after insulin treatment has been observed in a number of studies. In prospective studies on non–insulin-dependent (type 2) diabetes mellitus patients, a change in treatment from oral drugs to insulin was often associated with a significant increased risk of retinopathy progression and visual impairment. Although insulin therapies are critical for regulation of the metabolic disease, their role in the retina is controversial. In this study with diabetic mice, insulin treatment resulted in increased vascular leakage apparently mediated by betacellulin and signaling via the epidermal growth factor (EGF) receptor. In addition, treatment with EGF receptor inhibitors reduced retinal vascular leakage in diabetic mice on insulin. These findings provide unique insight into the role of insulin signaling in mediating retinal effects in diabetes and open new avenues for therapeutics to treat the retinal complications of diabetes mellitus.Diabetic maculopathy, an important cause of vision loss in patients with type 2 diabetes, is characterized by hyperpermeability of retinal blood vessels and subsequent formation of macular edema and hard exudates. Although the increase in retinal vascular permeability occurs both diffusely and in focal regions, the basic physiological defect that causes retinal vascular leakage is unknown. The blood–retinal barrier (BRB) isolates the retina from the bloodstream, establishing a favorable environmental milieu with the regulation of ionic balance, nutrient availability, and blockage of potentially toxic molecules that allows for optimal retinal function. The BRB consists of an inner BRB, formed by endothelial cells lining the retinal blood vessels and the outer BRB formed by the retinal pigment epithelium (RPE), a layer of epithelial cells between the retina and the non-neuronal choroid.^1,2 Disruption of the BRB is an important feature of diabetic retinopathy.Based on data from the Wisconsin Epidemiologic Study of Diabetic Retinopathy (WESDR), a prospective population-based cohort study of patients with type 1 and 2 diabetes mellitus, the prevalence of clinically significant macular edema is 5.9% for type 1 and 7.5% for type 2 diabetes.³ Although epidemiologic studies and randomized clinical trials suggest that glycemic control plays a major role in the development of vascular complications of diabetes,⁴ insulin therapies for control of glucose metabolism may not prevent long-term complications.^5,6 Even though both laser photocoagulation and anti-VEGF therapies have shown significant promise in the treatment of proliferating vessels in proliferative diabetic retinopathy, diabetic macular edema (DME) appears to be more resistant to these treatment approaches, suggesting that other factors might contribute to this complication. We have recently reported the potential role of betacellulin (Btc) in inducing retinal vascular permeability in diabetes.⁷ Clinical trials and other studies have determined that initiation of acute intensive insulin therapy in patients with long-standing poor glycemic control results in a transient worsening of diabetic retinopathy.^8–13 A change in treatment from oral drugs to insulin in patients with non–insulin-dependent (type 2) diabetes mellitus was associated with a significantly increased risk of retinopathy progression and visual impairment.^14–16 In addition, it has been reported that patients who undergo total pancreatectomy for cancer develop severe diabetes because of the complete absence of insulin but rarely if ever develop proliferative diabetic retinopathy,¹⁷ even when they survive for more than one or two decades. These reports led us to model and evaluate the pathophysiological effects of insulin on the retinal vasculature and the potential crosstalk between insulin and Btc in the regulation of retinal vascular permeability.     

Protection of Chinese painted quails (Coturnix chinensis) against a highly pathogenic H5N1 avian influenza virus strain after vaccination

Chinese painted quails immunized with a single dose (6 μg HA) of inactivated H5N1 (clade 1) influenza vaccine NIBRG-14 and challenged with 100 LD₅₀ of the heterologous A/Swan/Nagybaracska/01/06(H5N1) (clade 2.2) strain were protected, whereas unvaccinated quails died after challenge. No viral antigens or RNA were detected in cloacal swabs from immunized animals. Sera obtained post-immunization gave low titres in serological assays against the vaccine and the challenge viruses. Our results demonstrate the protective efficacy of the NIBRG-14 strain against the challenge virus and the usefulness of these small birds in protection studies of influenza vaccines.     

Elevated antibody levels against Chlamydia pneumoniae, human HSP60 and mycobacterial HSP65 are independent risk factors in myocardial infarction and ischaemic heart disease

The relative significance of traditional risk factors, chronic infections and autoimmune processes in the development of acute myocardial infarction (AMI) has not been fully elucidated. We compared serum IgG antibody titres to various pathogens, i.e. Chlamydia pneumoniae (Cpn), cytomegalovirus (CMV) and herpes simplex virus type 1 (HSV-1), and to the potential autoantigens human heat shock protein 60 (hHSP60) and mycobacterial heat shock protein 65 (mHSP65), in serum samples obtained from patients 3-48 h after AMI (n = 40) or stable effort angina (SEA, n = 43), and from controls (n = 46). The strongest association was observed between AMI and the elevated level of hHSP60 antibodies. The association between AMI and the level of Cpn antibodies was also significant. High levels of hHSP60 and Cpn antibodies represented independent risk factors for the development of AMI, but the simultaneous presence of high levels of antibodies to Cpn and hHSP60 suggested a joint effect on the relative risk of AMI (OR = 12.0-21.1). The antibody titres to mHSP65 were higher in the SEA group than in the controls, and the simultaneous presence of high levels of Cpn and mHSP65 antibodies meant an increased risk among the SEA patients. The antibody titres to CMV or HSV-1 were similar in the three groups. In conclusion, these results demonstrate associations of AMI with high levels of anti-hHSP60 and anti-Cpn antibodies, and of SEA with the level of anti-mHSP65 antibodies, these being independent risk factors.