Disordered vascular compliance in haemochromatosis.

BACKGROUND: A relationship may exist between body iron stores, endothelial dysfunction and overall cardiovascular risk. AIMS: To compare vascular compliance, biochemical endothelial function and antioxidant status between patients with homozygous hereditary haemochromatosis and healthy controls. METHODS: Haemochromatosis patients and healthy controls were recruited. Measures of vascular compliance were assessed by applanation tonometry. Serological markers of endothelial function (plasma lipid hydroperoxides, cell adhesion molecules), antioxidant levels (ascorbate, lipid soluble antioxidants) and high-sensitivity C-reactive protein (CRP) were also measured. RESULTS: Thirty-five hereditary haemochromatosis patients (ten females, mean age 54.6) and 36 controls (27 female, mean age 54.0) were recruited. Haemochromatosis patients had significantly higher systolic and diastolic blood pressures. Pulse wave velocity (PWV) was significantly higher in male haemochromatosis patients (9.90 vs. 8.65 m/s, p = 0.048). Following adjustment for age and blood pressure, male haemochromatosis patients continued to have a trend for higher PWVs (+1.37 m/s, p = 0.058). Haemochromatosis patients had significantly lower levels of ascorbate (46.11 vs. 72.68 µmol/L, p = 0.011), retinol (1.17 vs. 1.81 µmol/L, p = 0.001) and g-tocopherol (2.51 vs. 3.14 µmol/L, p = 0.011). However, there was no difference in lipid hydroperoxides (0.46 vs. 0.47 nmol/L, p = 0.94), cell adhesion molecule levels (ICAM: 348.12 vs. 308.03 ng/mL, p = 0.32 and VCAM: 472.78 vs. 461.31 ng/mL, p = 0.79) or high-sensitivity CRP (225.01 vs. 207.13 mg/L, p = 0.32). CONCLUSIONS: Haemochromatosis is associated with higher PWVs in males and diminished antioxidants across the sexes but no evidence of endothelial dysfunction or increased lipid peroxidation.

Adjuvant therapy used in conjunction with combination therapy for chronic hepatitis C improves sustained virus response rates in genotype 1 patients.

Combination treatment with pegylated interferon (Peg-IFN) and ribavirin remains the gold standard in the treatment of chronic hepatitis C. This therapy is limited by many side-effects including anaemia, neutropenia and reduced quality of life. The use of adjuvant agents to reduce the frequency of dose reductions because of haematological side-effects has been proven to be effective but there are few reports of what effect the use of these adjuvant therapies is having on sustained virological response (SVR). The aim of the study was to assess the clinical impact on sustained virological response of adjuvant therapies during combination therapy with Peg-IFN and ribavirin for chronic hepatitis C. A total of 132 patients, 96 males, were included in the study. The overall SVR was 66.7%, with 50% of genotype 1/4/6 (n = 27/54) patients achieving SVR and 78.2% of genotypes 2/3. The overall SVR of the treatment naïve patients (83/121) was 68.6%. Fifty-one of these patients were genotype 1 with 49.0% (25/51) of this group achieving SVR. The genotype 2/3 group of treatment naïve patients reached an SVR of 82.9% (58/70). Adjuvant therapy was used in 57 patients (43.8%). With the use of supportive adjuvant therapy, we achieved an overall SVR of 66.7% and in treatment naïve patients 68.6%. In genotype 1 patients, SVR rates of up to 46% have been reported in previous studies without the use of erythropoietin and granulocyte colony stimulating factor. We have demonstrated the SVR for genotype 1 can be improved to 50% overall.

Current concepts in the assessment and treatment of hepatic encephalopathy.

Hepatic encephalopathy (HE) is defined as a metabolically induced, potentially reversible, functional disturbance of the brain that may occur in acute or chronic liver disease. Standardized nomenclature has been proposed but a standardized approach to the treatment, particularly of persistent, episodic and recurrent encephalopathy associated with liver cirrhosis has not been proposed. This review focuses on the pathogenesis and treatment of HE in patients with cirrhosis. The pathogenesis and treatment of hepatic encephalopathy in fulminant hepatic failure is quite different and is reviewed elsewhere.

Hepatic encephalopathy as an unusual late complication of transjugular intrahepatic portosystemic shunt insertion for non-cirrhotic portal hypertension caused by nodular regenerative hyperplasia in an HIV-positive patient on highly active antiretroviral therapy.

Transjugular intrahepatic portosystemic shunt (TIPS) is an artificially created conduit between the portal and systemic vascular system in the liver performed percutaneously via radiological guidance. It is used mainly in conditions causing portal hypertension and its resulting complications. It reduces portal pressure by diverting portal blood flow into the systemic circulation. Hepatic encephalopathy is the most common complication following TIPS insertion and tends to present fairly early. We describe a case of hepatic encephalopathy as an unusual late complication of TIPS insertion (first presenting six years after) for non-cirrhotic portal hypertension caused by nodular regenerative hyperplasia in an HIV-positive patient on highly active antiretroviral therapy.

Treatment intensification does not reduce residual HIV-1 viremia in patients on highly active antiretroviral therapy.

In HIV-1-infected individuals on currently recommended antiretroviral therapy (ART), viremia is reduced to <50 copies of HIV-1 RNA per milliliter, but low-level residual viremia appears to persist over the lifetimes of most infected individuals. There is controversy over whether the residual viremia results from ongoing cycles of viral replication. To address this question, we conducted 2 prospective studies to assess the effect of ART intensification with an additional potent drug on residual viremia in 9 HIV-1-infected individuals on successful ART. By using an HIV-1 RNA assay with single-copy sensitivity, we found that levels of viremia were not reduced by ART intensification with any of 3 different antiretroviral drugs (efavirenz, lopinavir/ritonavir, or atazanavir/ritonavir). The lack of response was not associated with the presence of drug-resistant virus or suboptimal drug concentrations. Our results suggest that residual viremia is not the product of ongoing, complete cycles of viral replication, but rather of virus output from stable reservoirs of infection.

Early experience with chronic hepatitis C in Northern Ireland: epidemiology and response to monotherapy.

UNLABELLED: Chronic hepatitis C virus (HCV) infection has become a major health problem affecting an estimated 170 million people worldwide. The epidemiology of HCV and its response to treatment in Northern Ireland has not been described before. Our aims were to determine the epidemiology, histological stage, suitability for treatment and response to treatment in patients with hepatitis C presenting to one clinic in Northern Ireland. All patients were prospectively recruited with hepatitis C attending the Liver Clinic, Royal Victoria Hospital during the period December 1992 to June 1997. Sixty patients (33 male, mean age 44 years, range 19-84 years) who tested anti-HCV antibody positive were identified. The predominant genotypes were 1b (33%), 3a (28%) and 1a (26%). Most patients (78%) were asymptomatic at the time of detection and only four (7%) gave a history of jaundice. The most common modes of transmission were i.v. drug use in 30 (50%) and blood products in 20 (33%) patients. Forty-eight (86%) of the 56 patients tested were PCR positive for HCV RNA. Fifty-one patients (85%) underwent liver biopsy of whom 13 had cirrhosis (22% of original group). Twenty-nine patients were suitable for treatment, but three declined treatment and only 26 (43%) started interferon-alpha. During treatment 17 (65%) patients became PCR negative and eight (31%) remained PCR negative 12 months after completion of therapy. Liver histology was assessed before and after interferon treatment in 17 patients and showed no change in total necroinflammatory scores (p = 0.1) or staging of architectural change (p = 0.55). CONCLUSIONS: The epidemiology and response to therapy of HCV in Northern Ireland appear comparable to elsewhere in the UK. Only a minority of anti-HCV positive non-haemophiliac patients progress to have interferon therapy suggesting that the cost of treating chronic HCV may not be as great as initially thought.

Expression of type 2 nitric oxide synthase and p21 in oral squamous cell carcinoma.

Nitric oxide (NO) has a complex role in tumour biology. Most cancer research has focused on the enzyme nitric oxide synthase-2 (NOS2), an inducible isoform responsible for prolonged NO production. In normal cells exposed to high NO concentrations, the tumour-suppressor gene, p53, promotes apoptosis via the p21 pathway, in an attempt to safeguard against potential NO-mediated DNA damage. In cancer cells with mutant p53, this pathway is unlikely to occur directly, although, p53-independent p21 expression and subsequent apoptosis can occur at higher NO concentrations. In this study, the possible direct association between NOS2 and p21 was assessed in oral squamous cell carcinoma. Immunohistochemistry was performed for NOS2 and p21 on 56 cases, and NOS2 activity was determined with citrulline assays in selected cases. A significant relationship was demonstrated between the immunohistochemical expression of NOS2 and its activity (P<0.001), but not between NOS2 and p21 expression (P=0.76). It is unlikely that the NO concentrations found in oral cancer (up to 10.3 pmol NO min(-1) mg protein(-1)) are sufficient to cause direct (p53-independent) p21 accumulation and subsequent apoptosis. As with many other tumours, since NO production has a detrimental role, its pharmacological inhibition in oral cancer represents an exciting area for possible future therapeutic manipulation.

Does type II nitric oxide synthase expression correlate with cellular proliferation in oral squamous cell carcinoma and dysplasia?

BACKGROUND: Nitric oxide (NO) has been implicated both in tumor progression and inhibition. This study investigated whether type II nitric oxide synthase (NOS2) expression correlated with cell proliferation in oral squamous cell carcinoma (OSCC) and dysplasia. METHODS: Paraffin-embedded tissue samples of normal oral mucosa, OSCC, and dysplasia were assessed immunohistochemically using monoclonal antibodies to NOS2 and Ki-67 antigen. We used Western blotting to confirm NOS2 antibody specificity and protein expression in select cases. RESULTS: NOS2 staining was increased in OSCC relative to normal oral mucosa, in which no expression was found. Both NOS2 expression and Ki-67 indices independently correlated with grade of dysplasia (p < .001) but not with the degree of tumor differentiation. A positive correlation was found between NOS2 expression and Ki-67 in cases of mild and moderate dysplasia (p < .001), but not in severe dysplasia and OSCC. CONCLUSIONS: No correlation exists between Ki-67 and NOS2 expression in severe dysplasia and OSCC. The findings suggest that the level of NO produced by NOS2 is insufficient to affect cellular proliferation in these conditions. The mechanism of NOS2 activation and the consequences of its expression remain to be fully explained.

Biphasic regulation of NF-kappa B activity underlies the pro- and anti-inflammatory actions of nitric oxide.

Expression of inducible NO synthase (iNOS) by macrophages is a prerequisite for the production of high output NO, which mediates many bactericidal and tumoricidal actions of these immune cells. The expression of iNOS in mammalian cells is governed predominantly by the transcription factor, NF-kappa B, which regulates the expression of many host defense proteins. In the present study, we characterize a novel, biphasic effect of NO on NF-kappa B activity in murine macrophages. This mechanism depends on the local concentration of NO and enables it both to up- and down-regulate the expression of host defense proteins including iNOS, cyclooxygenase-2, and IL-6. This biphasic activity of NO appears to play a pivotal role in the time course of activation of these immune cells and, by inference, in facilitating the initiation of a defense response against pathogenic stimuli and in its termination to limit tissue damage. This mechanism may explain at least in part the reported ability of NO to act in both a pro- and anti-inflammatory manner.

Type II nitric oxide synthase (NOS2) expression correlates with lymph node status in oral squamous cell carcinoma.

In tumour biology, nitric oxide (NO) has a complex array of concentration-dependent actions, including both inhibitory and promoting effects. It is thought that the levels of NO found in many human cancers lead to enhanced angiogenesis and tumour dissemination. In the current study, we assessed the immunohistochemical expression of the enzyme type II nitric oxide synthase (NOS2) in 41 cases of oral squamous cell carcinoma and correlated the findings with lymph node status. A significant relationship was found between NOS2 expression and lymph node metastasis (P<0.0002). Furthermore, lymph node metastasis correlated with the degree and intensity of staining seen (P<0.001). No correlation was found between the size of the primary tumour, degree of tumour differentiation or smoking status and NOS2 staining. Western blotting confirmed NOS2 protein expression in select cases. As with many other human tumours, NOS2 is not a ubiquitous finding in oral cancer. Its expression may be of value in assessing lymph node status prior to surgery, and it represents a target for possible therapeutic manipulation.

Enhanced peroxynitrite formation is associated with vascular aging.

Vascular aging is mainly characterized by endothelial dysfunction. We found decreased free nitric oxide (NO) levels in aged rat aortas, in conjunction with a sevenfold higher expression and activity of endothelial NO synthase (eNOS). This is shown to be a consequence of age-associated enhanced superoxide (.O(2)(-)) production with concomitant quenching of NO by the formation of peroxynitrite leading to nitrotyrosilation of mitochondrial manganese superoxide dismutase (MnSOD), a molecular footprint of increased peroxynitrite levels, which also increased with age. Thus, vascular aging appears to be initiated by augmented.O(2)(-) release, trapping of vasorelaxant NO, and subsequent peroxynitrite formation, followed by the nitration and inhibition of MnSOD. Increased eNOS expression and activity is a compensatory, but eventually futile, mechanism to counter regulate the loss of NO. The ultrastructural distribution of 3-nitrotyrosyl suggests that mitochondrial dysfunction plays a major role in the vascular aging process.

Correlation between type II nitric oxide synthase and p53 expression in oral squamous cell carcinoma.

Prolonged nitric oxide (NO) production by the enzyme type II nitric oxide synthase (NOS2) has been implicated in angiogenesis and metastasis of human cancers. In animal models, wild-type p53 (but not mutant) protein results in down-regulation of NOS2 expression, which reduces both tumour growth and dissemination. In the current study, we aimed to find out whether a correlation was present in oral squamous cell carcinoma. Fifty-six cases of squamous cell carcinoma were assessed immunohistochemically using antibodies to NOS2 and p53 (clone DO-7). We also confirmed NOS2 protein expression in selected cases using immunoblotting. The results were correlated with clinicopathological findings. Statistical analysis showed a significant relationship between p53 and NOS2 expression (P= 0.001). No relationship was found between size of tumour or histological degree of differentiation, and NOS2 expression in the primary tumour.

A study to assess inducible nitric oxide synthase expression in oral lichen planus.

Nitric oxide (NO) has been implicated in a variety of diseases but has not been previously studied in oral lichen planus (OLP). Since OLP has a complex immunogenesis with abundant macrophage infiltration, this study determined by immunohistochemistry whether or not the expression of the inducible form of nitric oxide synthase (iNOS) was increased in this condition relative to normal mucosa. Thirty cases of OLP and 10 normal buccal mucosa biopsies were studied utilising primary antibodies to iNOS and CD68, a myelomonocytic marker. iNOS activity was additionally assessed using a [(14-)C]-labelled arginine to citrulline assay. CD68 expression was significantly increased in the cellular infiltrate of all 30 cases of OLP compared with normal mucosa (P<0.009). Although iNOS staining was seen in a minority of cells in nine cases, this was not statistically significant when compared with the absent staining in normal oral mucosa (P=0.26). Furthermore, the minimal iNOS activity found in OLP was similar to that in normal mucosa. We conclude that expression of iNOS by macrophages is downregulated in OLP and discuss the possible reasons for this finding.

Does expression of inducible nitric oxide synthase correlate with severity of oral epithelial dysplasia?

The small molecule nitric oxide (NO) has generated an exponential amount of research since its discovery as a biological messenger in 1987. It has a vast number of actions, many of which are poorly understood. It has been studied in a variety of human cancers and has been implicated both in tumour promotion and inhibition. Although NO is produced by three distinct isoforms of the enzyme nitric oxide synthase (NOS), most cancer research is directed towards the calcium-independent form, iNOS which following induction, produces much higher quantities of NO than the other two. In this study the expression of iNOS is assessed by immunohistochemistry in 26 cases of oral epithelial dysplasia ranging in severity from mild to severe. iNOS staining was found in all 26 cases of dysplasia with the degree of staining correlating to the severity of dysplasia (p < 0.001). There was no iNOS staining seen in adjacent normal epithelium. The possible role of iNOS in the complex transformation from dysplasia to invasive oral cancer and the clinical applications are discussed.

Afocal magnification does not influence chick eye development.

In defocus-induced ametropia experiments, retinal blur circles are a likely source of information as to the magnitude but not the sign of the defocus. However, magnification (and minification) produced by the lenses may be a cue. In this study, 1-day-old broiler chicks (N = 13) were treated monocularly for 7 days with special goggles containing approximately afocal iseikonic lenses which were designed to produce 10% retinal image magnification. This is a little less than the magnification produced by +10 D defocusing lenses used to produce about 10 D of hyperopia in earlier work. Intraocular dimensions of both eyes were measured by A-scan ultrasonography on the first and last day. Refractive states of both eyes were measured daily with a retinoscope and trial lenses. After the birds were sacrificed, the eyes were enucleated, weighed, and measured with calipers. Before the treatment there was no difference in the refractive state or dimensions of the right and left eyes. After 1 week of goggle wear there was still no significant difference between the eyes in spite of the magnification produced by the goggles. These data suggest that factors other than magnification are responsible for the ability of the eye to respond to the sign of defocus.

High incidence of the Cys 282 Tyr mutation in the HFE gene in the Irish population--implications for haemochromatosis.

A simple PCR-SSOP approach based on a single PCR product has been developed to screen the HFE gene for the haemochromatosis-associated mutations Cys 282 Tyr and His 63 Asp. Using this approach the prevalence of these mutations in a cohort (30) of haemochromatosis patients and normal controls (404) was determined. Ninety percent of the haemochromatosis patients were homozygous for the Cys 282 Tyr mutation. In the normal population we found an increased incidence of the Cys 282 Tyr mutation (17.3%; 95% confidence limits 0.136-0.209) which was also reflected in the higher frequency of Cys 282 Tyr homozygotes (1.24%; 95% confidence limits 0.0016-0.0232). Linkage disequilibrium analysis confirmed the association between A*03 and Cys 282 Tyr. However, strong linkage disequilibrium occurred with the HLA-A*03-associated allele HLA-B*14 but not the HLA-A*03-associated allele HLA-B*07. The His 63 Asp was found to be in linkage disequilibrium with HLA-A*29.

Orthotopic liver transplantation for hepatitis C infection: the best Christmas present?

We report the case of a 60-year-old man with mild Christmas disease, Factor IX 10% of normal, who developed chronic hepatitis C infection after receiving coagulation factor concentrates. Subsequently he developed encephalopathy and liver failure and was referred for liver transplantation. Following transplantation, Factor IX levels rapidly normalised and have remained so, representing a phenotypic cure of his Christmas disease.

High prevalence of celiac sprue among patients with primary biliary cirrhosis.

Although coexisting primary biliary cirrhosis (PBC) and celiac sprue have been described, celiac sprue is sufficiently common in western Europe for chance to explain isolated cases. We screened our patients with PBC for celiac sprue using serum immunoglobulin A endomysial antibody (EmA), with confirmation by duodenal biopsy in EmA-positive patients. Of 57 patients, 6 (11%) had EmA. Four agreed to have a biopsy taken, and all had villous atrophy, yielding a minimum prevalence of 1:14 (7%). Apart from anemia in one patient, none of the four had symptoms or routine laboratory abnormalities suggestive of celiac sprue. None had improvement in liver biochemical tests after 12 to 24 months on gluten-free diets despite the disappearance of EmA. Celiac sprue is common among patients with PBC and they should be routinely screened for this condition. Symptoms wrongly attributed to PBC may respond to gluten exclusion, and both conditions are potent risk factors for osteoporosis.