Structure of N-linked oligosaccharides attached to chlorovirus PBCV-1 major capsid protein reveals unusual class of complex N-glycans

The major capsid protein Vp54 from the prototype chlorovirus Paramecium bursaria chlorella virus 1 (PBCV-1) contains four Asn-linked glycans. The structure of the four N-linked oligosaccharides and the type of substitution at each glycosylation site was determined by chemical, spectroscopic, and spectrometric analyses. Vp54 glycosylation is unusual in many ways, including: (i) unlike most viruses, PBCV-1 encodes most, if not all, of the machinery to glycosylate its major capsid protein; (ii) the glycans are attached to the protein by a β-glucose linkage; (iii) the Asn-linked glycans are not located in a typical N-X-(T/S) consensus site; and (iv) the process probably occurs in the cytoplasm. The four glycoforms share a common core structure, and the differences are related to the nonstoichiometric presence of two monosaccharides. The most abundant glycoform consists of nine neutral monosaccharide residues, organized in a highly branched fashion. Among the most distinctive features of the glycoforms are (i) a dimethylated rhamnose as the capping residue of the main chain, (ii) a hyperbranched fucose unit, and (iii) two rhamnose residues with opposite absolute configurations. These glycoforms differ from what has been reported so far in the three domains of life. Considering that chloroviruses and other members of the family Phycodnaviridae may have a long evolutionary history, we suggest that the chlorovirus glycosylation pathway is ancient, possibly existing before the development of the endoplasmic reticulum and Golgi pathway, and involves still unexplored mechanisms.     

A novel animal model to evaluate the ability of a drug delivery system to promote the passage through the BBB

The purpose of this investigation was to explore the potentiality of a novel animal model to be used for the in vivo evaluation of the ability of a drug delivery system to promote the passage through the blood–brain barrier (BBB) and/or to improve the brain localization of a bioactive compound. A Tween 80^®-coated poly-l-lactid acid nanoparticles was used as a model of colloidal drug delivery system, able to trespass the BBB. Tacrine, administered in LiCl pre-treated rats, induces electrocorticographic seizures and delayed hippocampal damage. The toxic effects of tacrine-loaded poly-l-lactid acid nanoparticles (5 mg/kg), a saline solution of tacrine (5 mg/kg) and an empty colloidal nanoparticle suspension were compared following i.p. administration in LiCl-pre-treated Wistar rats. All the animals treated with tacrine-loaded nanoparticles showed an earlier outcome of CNS adverse symptoms, i.e. epileptic onset, with respect to those animals treated with the free compound (10 min vs. 22 min respectively). In addition, tacrine-loaded nanoparticles administration induced damage of neuronal cells in CA1 field of the hippocampus in all treated animals, while the saline solution of tacrine only in 60% of animals. Empty nanoparticles provided similar results to control (saline-treated) group of animals. In conclusion, the evaluation of time-to-onset of symptoms and the severity of neurodegenerative processes induced by the tacrine–lithium model of epilepsy in the rat, could be used to evaluate preliminarily the capability of a drug delivery system to trespass (or not) the BBB in vivo.     

Treatment of Isolated Below the Knee Deep Vein Thrombosis

The natural history of isolated distal deep-vein thrombosis (IDDVT) is still uncertain, as well as the real clinical risks associated with the disease and the need for its diagnosis and treatment. While more and more IDDVTs are diagnosed in everyday clinical practice, their appropriate therapeutic management is, unfortunately, far from straightforward, and different recommendations on how patients with diagnosed IDDVT should be treated are present between expert professionals and even among international guidelines. The present article aims at briefly reviewing the issue of IDDVT therapy in general, particularly focusing on the different approaches to the treatment of the disease that have been suggested by recent guidelines, those that are currently adopted in clinical practice, and necessary future directions.     

Muscle glycogenosis and mitochondrial hepatopathy in an infant with mutations in both the myophosphorylase and deoxyguanosine kinase genes

OBJECTIVES: To document 2 apparently incongruous clinical disorders occurring in the same infant: congenital myopathy with myophosphorylase deficiency (McArdle disease) and mitochondrial hepatopathy with liver failure and mitochondrial DNA depletion. METHODS: An infant girl born to consanguineous Moroccan parents had severe congenital hypotonia and hepatomegaly, developed liver failure, and died at 5 months of age. We studied muscle and liver biopsy specimens histochemically and biochemically, and we sequenced the whole coding regions of the deoxyguanosine kinase (dGK) and myophosphorylase (PYGM) genes. RESULTS: Muscle biopsy specimens showed subsarcolemmal glycogen accumulation and negative histochemical reaction for phosphorylase. Liver biopsy specimens showed micronodular cirrhosis and massive mitochondrial proliferation. Biochemical analysis showed phosphorylase deficiency in muscle and cytochrome c oxidase deficiency in liver. We identified a novel homozygous missense G-to-A mutation at codon 456 in exon 11 of PYGM, as well as a homozygous 4-base pair GATT duplication (nucleotides 763-766) in exon 6 of dGK, which produces a frame shift and a premature TGA stop codon at nucleotides 766 to 768, resulting in a truncated 255-amino acid protein. Both mutations were absent in 100 healthy individuals. CONCLUSIONS: Our data further expand the genetic heterogeneity in patients with McArdle disease; confirm the strong relationship between mitochondrial DNA depletion syndrome, liver involvement, and dGK mutations; and suggest that genetic "double trouble" should be considered in patients with unusual severe phenotypes.     

Hypospadias: incidence and effects on psychosexual development as evaluated with the Minnesota Multiphasic Personality Inventory test in a sample of 11,649 young Italian men

PURPOSE: We evaluated the incidence of hypospadias and its effects on psychosexual development in a sample of 11,649 young Italian males. Over the last 30 years only about 30 major publications have addressed these issues and the results of many studies have been contrasting. Some defects in methodology, such as low response rates, heterogeneity of age ranges and the choice of controls, have been the main limitations. Our study was designed to take these problems into account. MATERIALS AND METHODS: Forty-two hypospadic subjects and a random sample of 500 nonaffected males selected from the large sample of 11,649 young men (>90% of the 18-year-old males living in the Italian region of Tuscany) were screened by the Minnesota Multiphasic Personality Inventory (MMPI) test, psychological interview and clinical evaluation. RESULTS: The incidence of hypospadias in this representative group of Italian men is 3.6/1,000. No difference was noted in the percentage with altered MMPI compared with the control group. The age at surgical corrections and the number of operations are not related to an abnormal global psychological adjustment. Severity of disease influences a more negative genital appraisal and the number of operations is correlated only with more difficulty in initiating contact with the opposite sex. CONCLUSIONS: Surgery for hypospadias has to be strongly pursued in as many cases as possible. In addition, we strongly recommend following up all hypospadics, independently of the severity of their genital malformations, through adolescence to early adulthood, to ensure early detection of subjects with impaired psychological profiles.     

Symptomatic nonparasitic hepatic cysts: options for and results of surgical management

BACKGROUND: Management options for symptomatic nonparasitic hepatic cysts (SNHC) lack verification through comparative studies with respect to safety and long-term effectiveness. HYPOTHESIS: Open cystectomy is the treatment of choice for patients with SNHC. SETTING: University hospital department of surgery. PATIENTS: Data were retrospectively analyzed from the clinical charts of 34 patients (26 women and 8 men) undergoing surgery for SNHC from January 1, 1975, through January 1, 1999. Charts were obtained from the original hospital referral. MAIN OUTCOME MEASURES: Morbidity rates and long-term recurrence. We considered the following variables for analysis: age, sex, hepatic cyst location, diameter of the cyst at primary surgery, symptoms, surgical procedure, postoperative morbidity and mortality, length of postoperative hospital stay, and long-term outcome. RESULTS: The 34 patients underwent 47 operations for SNHC (mean diameter, 15.0 cm), with a mean follow-up of 50.0 months. Ten patients underwent open and 8, laparoscopic deroofing of the cyst. Enucleation of the cyst and hepatic resections were performed as primary procedures in 4 and 2 patients, respectively, and as secondary procedures in 6 and 7 patients, respectively. Two recurrences (25%) were found after laparoscopic deroofing and 3 (30%) after open deroofing. Two (50%) and 6 (100%) recurrences were found after cystojejunostomy and needle aspiration, respectively. No symptomatic recurrences occurred after 10 cystectomies and 9 hepatectomies. One operative death (3%) occurred; however, morbidity rates were 18% (6/34) and 15% (2/13) after primary and secondary surgery, respectively. CONCLUSIONS: These results support our policy of performing open radical procedures in the treatment of SNHC; cystectomy is performed for primary surgery and hepatic resections for recurrences and complications. Conservative procedures have shown higher rates of recurrence and the need for further surgery. Only further technological improvements will allow a systematic and safe use of laparoscopy for radical surgery for SNHC.     

Nutritional characteristics of a rural Southern Italy population: the Ventimiglia di Sicilia Project

OBJECTIVE: Knowledge of alimentary habits among populations permits a better definition of appropriate public health interventions. We designed the epidemiological project "Ventimiglia di Sicilia" to characterize the risk profile in a rural village with low total cholesterol levels and low early cardiovascular mortality but with a large prevalence of overweight and obesity, which previously have been significantly associated with total mortality. METHODS: 488 individuals of age 20 to 69 years were included in the dietary survey conducted by a seven-day food record. RESULTS: Alimentary habits were characterized by high consumption of total and complex carbohydrates (respectively 52.5 +/- 7.6% and 46.6 +/- 8.2% of daily energy) and by a low cholesterol intake (92.5 +/- 35.0 mg/1000 kcal/day). Fat intake was 34.7 +/- 7.7% of daily energy due to a higher consumption of monounsaturated fats in respect to saturated fats (respectively 20.5 +/- 5.1% and 10.2 +/- 2.9% of daily energy). In particular, in this population there was a large consumption of bread, pasta, fresh vegetables, olive oil and fruits. We also observed an excess of total calories (about 2900 kcal/day in men and 2100 kcal/day in women) not balanced by a high degree of physical activity levels. Furthermore we found a significant higher total and saturated fat consumption in the youngest individuals and in people with higher educational levels. CONCLUSIONS: Dietary habits of Ventimiglia di Sicilia still follow the nutritional characteristics typical of the Mediterranean diet. The high total calorie intake indicates a quantitative more than qualitative problem, which may account the large prevalence of overweight and obesity and may represent a public health issue that needs to be corrected in such a rural population.