Synthesis and preclinical evaluation of BOLD-100 radiolabeled with ruthenium-97 and ruthenium-103.

BOLD-100 (formerly IT-139, KP1339), a well-established chemotherapeutic agent, is currently being investigated in clinical trials for the treatment of gastric, pancreatic, colorectal, and bile duct cancer. Despite numerous studies, the exact mode of action is still the subject of discussions. Radiolabeled BOLD-100 could be a powerful tool to clarify pharmacokinetic pathways of the compound and to predict therapy responses in patients using nuclear molecular imaging prior to the therapy. In this study, the radiosyntheses of carrier-added (c.a.) [97/103Ru]BOLD-100 were performed with the two ruthenium isotopes ruthenium-103 (103Ru; ß-, γ) and ruthenium-97 (97Ru; EC, γ), of which in particular the latter isotope is suitable for imaging by single-photon emission computed tomography (SPECT). To identify the best tumor-to-background ratio for diagnostic imaging, biodistribution studies were performed with two different injected doses of c.a. [103Ru]BOLD-100 (3 and 30 mg kg-1) in Balb/c mice bearing CT26 allografts over a time period of 72 h. Additionally, ex vivo autoradiography of the tumors (24 h p.i.) was conducted. Our results indicate that the higher injected dose (30 mg kg-1) leads to more unspecific accumulation of the compound in non-targeted tissue, which is likely due to an overload of the albumin transport system. It was also shown that lower amounts of injected c.a. [103Ru]BOLD-100 resulted in a relatively higher tumor uptake and, therefore, a better tumor-to-background ratio, which are encouraging results for future imaging studies using c.a. [97Ru]BOLD-100.

Histopathological examination of characteristic brain MRI findings in acute hyperammonemic encephalopathy: A case report and review of the literature.

INTRODUCTION: Acute hyperammonemic encephalopathy is associated with distinct brain MRI findings, namely, hyperintensity in T2-weighted sequences as well as restricted diffusion in diffusion-weighted imaging with accentuation in the insular cortex and cingulate gyrus. The pathophysiology and the histopathological correlates of these characteristic MRI findings are largely unknown. CASE REPORT: We present a 57-year-old male with a history of chronic alcohol abuse, liver cirrhosis and portal hypertension, and a clinical syndrome (variceal bleeding, depression of consciousness, seizures), elevated plasma ammonia levels, and characteristic brain MRI abnormalities suggestive of acute hyperammonemic encephalopathy. A postmortem histopathological examination revealed extensive hypoxic ischemic encephalopathy without evidence for metabolic encephalopathy. No episodes of prolonged cerebral hypoxemia were documented throughout the course of the disease. We conducted a review of the literature, which exhibited no reports of hyperammonemic encephalopathy in association with characteristic brain MRI findings and a consecutive histopathological examination. CONCLUSION: This is the first report of a patient with acute hyperammonemic encephalopathy together with characteristic brain MRI findings and a histopathological correlation. Although characteristic MRI findings of acute hyperammonemic encephalopathy were present, a histopathological examination revealed only hypoxic pathology without signs of metabolic encephalopathy.

Development of peptide aptamer microarrays for detection of HPV16 oncoproteins in cell extracts.

Protein microarrays represent an emerging technology that promises to facilitate high-throughput proteomics. The major goal of this technology is to employ peptides, full-length proteins, antibodies, and small molecules to simultaneously screen thousands of targets for potential protein-protein interactions or modifications of the proteome. This article describes the performance of a set of peptide aptamers specific for the human papillomavirus (HPV) type 16 oncoproteins E6 and E7 in a microarray format. E6 and E7 peptide aptamer microarrays were probed with fluorescence-labeled lysates generated from HPV-infected cervical keratinocytes expressing both E6 and E7 oncoproteins. Peptide aptamer microarrays are shown to detect low levels of E6 and E7 proteins. Peptide aptamers specific for cellular proteins included on these microarrays suggested that expression of CDK2, CDK4, and BCL-6 may be affected by HPV infection and genome integration. We conclude that peptide aptamer microarrays represent a promising tool for proteomics and may be of value in biological and clinical investigations of cervical carcinogenesis.

High-level expression of the retinoic acid receptor beta gene in normal cells of the uterine cervix is regulated by the retinoic acid receptor alpha and is abnormally down-regulated in cervical carcinoma cells.

Retinoic acid (RA) is essential for regulation of epithelial cell differentiation. The intracellular effects of RA are mediated by RA-binding nuclear receptors, including the RA receptors (RARs) alpha, beta, and gamma. The ligand-activated receptors induce the transcription of target genes by binding to RA-responsive elements in the promoter regions. One target gene is the RAR beta gene, which encodes a potential tumor suppressor. Loss of RA inducibility of RAR beta gene expression is assumed to play a role in the development of several types of human carcinomas, including carcinomas of the uterine cervix. We have analyzed RAR beta gene expression in normal cervical cells and in cervical carcinoma cell lines. The results show that the RAR beta mRNA levels are high and RA inducible in the primary keratinocytes, whereas they are low and not inducible or only slightly inducible by RA in all of the cervical carcinoma cell lines analyzed. The basal and the RA-induced RAR beta mRNA levels tend to increase with senescence of the normal cells. Fusion of primary ectocervical keratinocytes with HeLa cervical carcinoma cells revealed that the characteristics of RAR beta gene expression of the normal cells are dominant over that of the tumor cells. Using synthetic retinoids with receptor-preferential agonist activities and a RAR alpha-specific antagonist, we show that RAR alpha is the major endogenous RAR subtype for induction of RA-dependent RAR beta gene expression. Taken together, our results indicate that abnormal downregulation of RAR beta gene expression may be an important step in the multifactorial process of cervical carcinogenesis.

Peptide aptamers targeting the hepatitis B virus core protein: a new class of molecules with antiviral activity.

A substantial proportion of the worldwide liver cancer incidence is associated with chronic hepatitis B virus (HBV) infection. The therapeutic management of HBV infections is still problematic and novel antiviral strategies are urgently required. Using the peptide aptamer screening system, we aimed to isolate new molecules, which can block viral replication by interfering with capsid formation. Eight peptide aptamers were isolated from a randomized expression library, which specifically bound to the HBV core protein under intracellular conditions. One of them, named C1-1, efficiently inhibited viral capsid formation and, consequently, HBV replication and virion production. Hence, C1-1 is a novel model compound for inhibiting HBV replication by blocking capsid formation and provides a new basis for the development of therapeutic molecules with specific antiviral potential against HBV infections.

Induction of the p53-target gene GADD45 in HPV-positive cancer cells.

The E6 oncoprotein of human papillomaviruses (HPVs) has the potential to functionally antagonize p53. In several experimental model systems, ectopic expression of E6 can block the genotoxic induction of the growth inhibitory p53 target gene gadd45, suggesting that the inactivation of this pathway may play a major role for HPV-associated cell transformation. Here, we investigated whether this reflects the regulation of gadd45 expression in carcinoma-derived HPV-positive cells. We found that the gadd45 gene is efficiently induced by mitomycin C, cisplatin, and UV irradiation in a series of HPV-positive cervical cancer cell lines. Moreover, clear induction of gadd45 gene expression was also observed following treatment with gamma-irradiation, a pathway that is strictly dependent on functional p53. This contrasted with findings in human foreskin keratinocytes experimentally immortalized by expressing the HPV16 E6, E7, or E6/E7 oncogenes from the heterologous CMV promoter, where expression of the E6 gene was linked to a lack of gadd45 induction following gamma-irradiation. These results indicate (1) that the tumorigenic phenotype of HPV-positive cancer cells is not linked to an inability to induce the gadd45 gene following DNA damage, (2) that experimental model systems in which the E6 gene is expressed ectopically and/or in a different cellular context do not necessarily reflect the regulation of p53-associated pathways in HPV-positive cancer cells and (3) that a pathway strictly depending on functional p53 is inducible in HPV-positive cancer cells, providing direct evidence that the endogenous p53 protein in these cells is competent to activate a cellular target gene, despite coexpression of the viral E6 oncogene.

p53 mutations are rare events in recurrent cervical cancer.

Mutations of the p53 gene have been shown to be associated with aggressive growth behavior and increased recurrence rates for certain tumors. Primary cervical cancers contain oncogenic human papillomaviruses (HPV) in more than 90% of cases and usually possess wild-type p53 alleles. Cervical cancer cells contain detectable levels of functional p53 protein despite of the expression of the HPV E6 protein, which can induce p53 degradation. Thus, inactivation of p53 by somatic mutation should have functional consequences in HPV-positive cancers. We investigated whether p53 mutations play a role in the recurrence of the disease by analyzing p53 status in 18 biopsy specimens from recurrent cervical cancers. Only one of these (5.6%) contained a p53 mutation, as assessed by a sensitive yeast functional assay that detects mutations of the p53 mRNA between codons 52 and 364. These results indicate that p53 mutations are rare events in recurrent cervical carcinomas, and that somatic mutations of p53 do not provide cervical cancer cells with a selective growth advantage for recurrence.

Expression pattern of AP-2 transcription factors in cervical cancer cells and analysis of their influence on human papillomavirus oncogene transcription.

The AP-2 family of transcription factors consists of three known members, namely AP-2alpha, AP-2beta, and AP-2gamma. In experimental systems AP-2 factors possess tumor suppressor-like activities, and alterations in the AP-2 expression pattern have been described for some tumor entities. In addition, AP-2 has been implicated in the transcriptional control of human papillomaviruses (HPVs). We investigated here the expression pattern of AP-2alpha, AP-2beta, and AP-2gamma, as well as that of the cellular AP-2 target gene c-erbB-2, in a series of cervical cancer cell lines. In addition, we analyzed the influence of AP-2 factors on the activity of the HPV16 and HPV18 E6/E7 oncogene promoter. We found that, with the exception of HPV-negative C33A cells, all investigated cervical cancer cell lines expressed all three AP-2 family members, although at varying levels. No linear correlation between AP-2 and c-erbB-2 levels was observed. Although AP-2alpha, AP-2beta, and AP-2gamma can activate the c-erbB-2 promoter in reporter gene assays, they do not stimulate the HPV16 or HPV18 E6/E7 promoter. These results indicate that, although a rare event, loss of AP-2 expression occurs in cervical cancer cells. Moreover, AP-2alpha, AP-2beta, and AP-2gamma are neither sufficient nor required to activate the viral E6/E7 promoter.

Cell cycle regulation of deoxycytidine kinase. Evidence for post-transcriptional control.

Deoxycytidine kinase enzyme activity and deoxycytidine kinase mRNA content were determined at different positions of the cell cycle in permanent cell lines. There was no variation of deoxycytidine kinase activity during the cell cycle in two cell lines, whereas in two other lines the enzyme activity was induced (10- and 15-fold) at the G1/S boundary. In contrast, the level of deoxycytidine kinase mRNA never displayed any cell cycle-dependent changes. The decay of enzymatic activity was measured after addition of cycloheximide in different phases of the cell cycle; the enzyme was much more stable in cells with constant activity. We suggest that post-transcriptional mechanisms account for the periodic behaviour of the enzyme activity, and that whether this regulation can be detected depends on the half-life of deoxycytidine kinase.

Reduced gene expression of E-cadherin and associated catenins in human cervical carcinoma cell lines.

Cell-cell adhesion mediated by E-cadherin is often lost or disturbed in human carcinomas. For regular adhesive function, E-cadherin has to form complexes with peripheral cytoplasmic catenins which are multifunctional proteins that are also involved in signal transduction and growth regulation. We have analyzed the expression levels of the genes encoding alpha-catenin, beta-catenin and plakoglobin in correlation to the E-cadherin expression levels in cell lines derived from human cervical carcinomas. Reduced mRNA and protein levels were detected for plakoglobin, whereas alpha- and beta-catenin showed only reduced protein (but not mRNA) levels. The alterations in catenin gene expression were often associated with absent or reduced E-cadherin. The findings indicate that a reduction of catenin gene expression may contribute to the development of cervical carcinomas.

Peptide aptamers: new tools to study protein interactions.

The ability to specifically interfere with the function of proteins of pathological significance has been a goal for molecular medicine for many years. Peptide aptamers comprise a new class of molecules, with a peptide moiety of randomized sequence, which are selected for their ability to bind to a given target protein under intracellular conditions. They have the potential to inhibit the biochemical activities of a target protein, can delineate the interactions of the target protein in regulatory networks, and identify novel therapeutic targets. Peptide aptamers represent a new basis for drug design and protein therapy, with implications for basic and applied research, for a broad variety of different types of diseases.

Do consumers know how their health plan works?

Expanding consumer choice of plans is beneficial only to the extent that consumers make informed choices. Using data from the 1996-97 Community Tracking Study (CTS), this study compares consumers' responses on four key attributes of their health plan with information provided directly by the plan. Plan attributes relate to choice of providers and access to specialists. Although the accuracy of reporting some individual attributes was fairly high, fewer than one-third of consumers accurately reported all four health plan attributes. In general, consumers tended to overreport plan restrictions, especially the need for approval to see specialists.

Public opinion on alcohol policies in the United States: results from a national survey.

We surveyed the U.S. non-institutionalized population age 18+ on opinions regarding 23 alcohol control policies (N = 7,021). The cooperation rate among contacted households was 70% and the overall response rate was 54%. Results showed high levels of public support for most alcohol control policies. Over 80% support restrictions on alcohol use in public places, such as parks, beaches, concert venues, and on college campuses. Eighty-two percent support increased alcohol taxes, provided the funds are used for treatment or prevention programs. Over 60% support alcohol advertising and promotion restrictions, such as banning billboard advertising, banning promotion at sporting events, or banning liquor and beer advertising on television. Multivariate regression analyses indicated significant relationships between alcohol policy opinions and a variety of sociodemographic, political orientation, and behavioral measures. However, the absolute differences in alcohol policy support across groups is small. There is a strong base of support for alcohol control policies in the U.S., and such support is found among whites and ethnics of color, young and old, rich and poor, and conservatives, moderates, and liberals.

Risk factors for late relapse in male and female ex-smokers.

Three hundred twenty-nine ex-smokers (149 males and 180 females) who had maintained abstinence for at least 3 months prior to intake were followed for a period of 1 year to ascertain individual characteristics from cognitive, behavioral, environmental, and health domains that were predictive of later relapse. Multiple logistic regression analysis revealed significant associations between relapse, family history, and stress variables in males and between relapse, family history, stress, pre- and postcessation smoking characteristics, work-related variables, and physical activity in females. Within the predictor sets, both risk and protective factors were identified. The male and female models are discussed separately, compared, and related to previous research. Implications for further model development are specified; most notably, the utility of conceptualizing relapse independent of gender is questioned. Future research efforts could focus on the role played by family smoking history in adult relapse behavior. Smoking cessation interventions may need to tailor their efforts specifically to men and to women. Along with techniques to manage stress both during and after cessation, women may benefit especially from interventions that provide nicotine replacement and counseling for work-related difficulties.

Light and electron microscopic examination of human subungual tissue. Glomus and lamellated bodies.

OBJECTIVE: There is only limited data related to the subungual glomus body. We therefore studied the structure of this organ, aiming to obtain further evidence. Additionally, we encountered undefined receptor like structures in close association with these glomus cells, named them as lamellated bodies and examined both of the structures at light and electron microscopic levels. METHODS: This study was carried out at the Faculty of Medicine, Hacettepe University, Ankara, Turkey, during the time period May 2001 to March 2002. In this study, the subungual tissues of 4 patients were examined. RESULTS: Within subungual tissue, 2 groups of morphologically significant structures were determined by light microscopy. The first structure was described as glomus body. It was characterized as an encapsulated structure, rich in rounded clear cells filling its central compartment. The latter structure having a lamellated appearance was described as lamellated body. In the electron microscopic examination, lamellated bodies were characterized by central filament rich large cells and surrounding cytoplasmic processes of ensheathing cells, some of which were vacuolated. Glomus bodies were surrounded by a capsule and centrally located numerous rounded cells which reflected the structural features of an active cell. CONCLUSION: The lamellated bodies are very unusual structures and they are not found in any other part of the body. The structural organization of the ensheathing cells in the lamellated bodies greatly resembles many skin associated receptors. Therefore, we planned future studies by using immunohistochemistry, to reveal nervous elements for structural contribution.

Height of normal pituitary gland on MRI: differences between age groups and sexes.

In this study, the coronal and sagittal heights of the pituitary glands were measured by magnetic resonance imaging (MRI) technique in 201 individuals. There was no sellar or parasellar region pathology in the study group. The data were evaluated according to age and sex groups. In all cases the coronal and sagittal heights of the pituitary glands were equal. The mean values of the coronal and sagittal heights in females (6.1 +/- 0.1 mm, 6.1 +/- 0.1 mm respectively) were higher than in males (5.7 +/- 0.2 mm, 5.6 +/- 0.2 mm respectively). The highest values for the coronal and sagittal heights were in the 11-20 years age group in both sexes. A gradual increase in the coronal and sagittal heights of the pituitary glands in the 0-10, 11-20 age groups was present in both sexes. Decrease in the heights of the pituitary glands was noted after 20 years of age onwards. Nevertheless there was a conspicuous increase in the mean value of the pituitary glands' heights in the 51-60 years age group in males. In females, a minimal increase in the mean value of the pituitary glands' heights was observed in the 61 years and over age group.

A unique anomaly of the fibrous sling of the digastric muscle.

Anomalies of digastric muscle are not very rare. During a routine dissection, it was found on a cadaver that the digastric muscles on both sides had no fibrous slings. It was also observed that the two intermediate tendons crossed over the stylohyoid muscles. Such anomalies should be considered during the evaluation of the floor of the mouth after CT examination of MR imaging, and during surgical procedures as well.

An accessory fissure in the lower lobe of the right lung.

We report here an accessory fissure in the right lung. Accessory fissures are described as clefts of various depths lined by two layers of visceral pleura. Anomalies of location of the lungs may be produced by the accessory fissures. In our case, an unusual accessory fissure in the right lung was found in a 50 years old male cadaver. The accessory fissure was located between the superior and basal segments of the lower lobe of the right lung. Its depth was around 3 cm as an average value. Its length was found to be 12 cm on the surface of the lung. In conclusion, a surgeon must always remember the anatomical variations of the location of the lungs especially in lobectomies and in segmental resection. From the radiological aspect, an accessory fissure is important in that it can be mistaken for lung pathologies.

CT-guided piriformis muscle injection for the treatment of piriformis syndrome.

AIM: Piriformis syndrome is a rare neuromuscular disorder that occurs when the piriformis muscle compresses or irritates the sciatic nerve. The treatment of piriformis syndrome includes injections into the piriformis muscle around the sciatic nerve. These invasive approaches have been used with various techniques to increase the safety of the procedure. Computed tomography (CT)-guided injection of the piriformis muscle and the clinical outcome of the patients are discussed. MATERIAL AND METHODS: The authors presented 10 consecutive patients that underwent CT-guided piriformis injection between March and December 2007. Three patients had a history of a severe fall on the buttocks, one had gluteal abscess formation following deep intramuscular injection, and another one had a habit of prolonged sitting on the carpet. Etiology was not identified in the other patients. Main complaints of the patients were pain and numbness in the legs. Hypesthesia was the major neurological finding. Magnetic resonance imaging (MRI) and electromyography (EMG) were performed in all patients. RESULTS: Nine patients had full and sustained recovery of their symptoms after piriformis injection. Only the patient who had gluteal abscess formation following deep intramuscular injection showed moderate improvement. Another patient was operated on in the 6th month after piriformis injection due to an extruded disc herniation. CONCLUSION: CT-guided piriformis injection is a safe and effective method in the treatment of piriformis syndrome.