64-slice MDCT arthrography in shoulder instability: our experience.

PURPOSE: This study was performed to assess the diagnostic accuracy of air-contrast 64-slice multidetector computed tomography (MDCT) arthrography in the evaluation of glenohumeral joint instability by comparison with conventional arthroscopy. MATERIALS AND METHODS: Fifty patients with a history of shoulder instability underwent MDCT arthrography with thin collimation scans. The raw data were transferred to a workstation and processed using multiplanar reformation (MPR) and volume rendering (VR) algorithms. All patients subsequently underwent conventional arthroscopy. The results of the two techniques were compared and their sensitivity and specificity calculated. RESULTS: We diagnosed eight anterosuperior labrum lesions (group 1), 32 anteroinferior labrum lesions (group 2) and two posterior labrum lesions (group 3). Overall sensitivity and specificity (groups 1, 2, 3) were 88% and 100%, respectively. In group 1, sensitivity was only 66% (four false negatives), whereas in groups 2 and 3, it was 94% (two false negatives) and 100%, respectively. The labrum lesions were also found to be associated, with 100% sensitivity and specificity, with 20 lax capsules, 17 Hill-Sachs lesions, five Bankart lesions, two Perthes lesions and three complete rotator-cuff tears. CONCLUSIONS: Air-contrast MDCT arthrography is fast, reproducible, well tolerated and very accurate in the evaluation of lesions causing shoulder instability.

SARS Surveillance Project--Internet-enabled multiregion surveillance for rapidly emerging disease.

INTRODUCTION: On March 15, 2003, CDC requested health-care and public health agencies to conduct surveillance for severe acute respiratory syndrome (SARS). The SARS Surveillance Project (SARS-SP) was established to rapidly implement multiregional SARS surveillance in emergency departments (EDs) by using existing Internet-based tools. OBJECTIVES: The objectives of SARS-SP were to 1) disseminate and update SARS screening forms for ED triage, 2) establish surveillance for SARS syndrome elements by using Regional Emergency Medicine Internet (REMI), 3) expand surveillance to multiple regions, and 4) evaluate the usefulness of Internet tools for agile surveillance during a rapidly emerging global epidemic. METHODS: SARS-SP developed, distributed, and updated an Internet-based triage form to identify patients for infection control and public health reporting. EDs then were invited to report visit frequencies with various SARS syndrome elements to local public health authorities by using the REMI Internet application (first in one metropolitan area, and later in four). After pilot-testing in one metropolitan area, the surveillance system was implemented in three others. RESULTS: Active syndromic surveillance was established by health departments in Milwaukee, Wisconsin; Denver, Colorado; Akron, Ohio; and Fort Worth, Texas. A total of 27 EDs reported syndrome frequencies from >146,000 patient encounters. CONCLUSIONS: ED and public health partners reported being satisfied with the system, confirming the usefulness of Internet tools in the rapid establishment of multiregion syndromic surveillance during an emerging global epidemic.

Effect of poly(ADP-ribosyl)ation and Mg2+ ions on chromatin structure revealed by scanning force microscopy.

Poly(ADP-ribosyl)ation of nuclear proteins is responsible for major changes in the high-order chromatin structure. The effects of this post-translation modification on nuclear architecture were examined at different Mg2+ concentrations using scanning force microscopy. A quantitative analysis of the internucleosomal distance, the width, and the volume of chromatin fibers imaged in tapping mode reveals that poly(ADP-ribosyl)ation induces a complete relaxation and decondensation of the chromatin structure. Our data, on the center-to-center distance between adjacent nucleosomes and on the fiber width, indicate that the poly(ADP-ribosyl)ated fibers remain significantly decondensed even in the presence of Mg2+. Our results also show that the Mg2+ assumes an important role in the folding of chromatin structure, but Mg2+ is not able to restore the native feature of chromatin, when the fibers are depleted of H1/H5 histones. The combined effect of post-translation modification and cation ions on the chromatin structure shows that poly(ADP-ribosyl)ation could promote accessibility to DNA even in those nuclear processes that require Mg2+.

Interactions of Epstein-Barr virus origins of replication with nuclear matrix in the latent and in the lytic phases of viral infection.

Eukaryotic DNA is organized into domains or loops generated by the attachment of chromatin fibers to the nuclear matrix via specific regions called scaffold or matrix attachment regions. The role of these regions in DNA replication is currently under investigation since they have been found in close association with origins of replication. Also, viral DNA sequences, containing the origins of replication, have been found attached to the nuclear matrix. To investigate the functional role of this binding we have studied, in Raji cells, the interaction between Epstein-Barr virus (EBV) origins of replication and the nuclear matrix in relation to the viral cycle of infection. We report here that both the latent (ori P) and the lytic (ori Lyt) EBV origins of replication are attached to the nuclear matrix, the first during the latent cycle of infection and the second after induction of the lytic cycle. These findings suggest that the binding of the origins of replication with the nuclear matrix modulates viral replication and expression in the two different phases of infection.

High glucose modifies heparansulphate synthesis by mouse glomerular epithelial cells.

BACKGROUND: Alterations in proteoglycan metabolism are involved in the pathogenesis of diabetic nephropathy. The aim of this study is to evaluate the effects of high glucose on proteoglycan production and to find a reliable in vitro model for the study of diabetic nephropathy. METHODS: A clone of mouse glomerular epithelial cells was cultured in media containing elevated (30 mmol) and physiological (5 mmol) glucose, or iso-osmolar (30 mmol) mannitol concentrations. We evaluated the synthesis of 35SO4-labeled molecules and the amount of proteoglycans by Sepharose CL6B and DEAE-Sephacel chromatographies. RESULTS: A clear decrease (56%) in total cell-layer proteoglycan synthesis was induced by 30 mmol glucose, in comparison with normal glucose. A reduction of 25% in medium associated proteoglycan synthesis was observed in high glucose cultured cells. After Sepharose CL6B, in cells cultured in high glucose, cell layer heparansulphate proteoglycan-I (Kav 6B 0. 04) synthesis was reduced by about 81%, heparansulphate proteoglycan-II (Kav 6B 0.21) by about 87% and heparansulphate glycosaminoglycan (Kav 0.4-0.8) by about 91%, respectively. In mannitol-incubated cells the reductions observed were less evident and not significantly different from those in normal glucose. CONCLUSIONS: These results indicate that (1) glomerular epithelial cells play a central role in proteoglycan synthesis, (2) high glucose modifies the amount and influences the different species production of these macromolecules, while osmotic forces seem to be only partially involved in these effects, and (3) this cellular clone of glomerular epithelial cells can represent a reliable in vitro model for the study of the mechanisms involved in diabetic nephropathy.

[Ozone therapy in lumbar sciatic pain]. / Ozonoterapia intradiscale nella lombosciatalgia.

INTRODUCTION: Medical ozone is a mixture of oxygen and ozone which can be used for several medical applications. Ozone was first applied clinically to the treatment of lumbar sciatic pain peridurally, while Pietrogrande was the first in Italy to report on its intradiscal administration to treat nucleus polposus herniation. On account of these considerations, we have decided to introduce this method in our Institute (I.C.O.T. Latina) as an alternative to surgery in the treatment of lumbar sciatic pain supported by an intradiscal hernia. MATERIAL AND METHODS: September, 1995, to April, 1997, we treated more than 1000 patients with intradiscal ozone infiltration. We prospectively analyzed the first 50 patients, with 6 months' follow-up at least; all of them were preliminarily submitted to clinical examination, electromyography, CT and MRI. After local anesthesia, we injected the disk, with 18-20 G needles and under CT or fluoroscopic guidance, with 12 ml of a mixture of oxygen and ozone at a concentration of 20-30 micrograms/ml. The treatment was repeated two or three more times at intervals of 3, 15 or, when necessary, 30 days. After each treatment, CT follow-ups were carried out and the final follow-up was made 3 months later. RESULTS: We divided our results into clinical and instrumental. As for clinical response, we had 68% positive results (40% excellent, 28% good) and 32% negative results (10% of patients underwent surgery and 22 are under medical and physical treatment). As for CT response, we had 82% positive results (36% excellent, 46% good), while no major changes between pre- and post-treatment CT findings in the remaining 18% of cases. CONCLUSIONS: Ozone therapy, thanks to its ease of execution and noninvasiveness, permits the successful outpatient treatment of lumbar sciatic pain. Moreover, the lack of major complications and the good results obtained compared to other methods, such as chemonucleolysis, percutaneous automated discectomy, microsurgery and conventional surgery, suggest that ozone therapy can be considered the treatment of choice for lumbar sciatic pain and a valid alternative to surgery in many cases.

Does poly(ADP-ribosyl)ation regulate the DNA methylation pattern?

The existence of a possible correlation between poly(ADP-ribosyl)ation and DNA methylation processes was investigated. In vivo and in vitro experiments were carried out on L929 mouse fibroblasts preincubated for 24 h with or without 3-aminobenzamide, a well-known inhibitor of poly(ADP-ribose) polymerase. Both experimental approaches evidenced a close relationship between these two important nuclear enzymatic mechanisms, suggesting that the poly(ADP-ribosyl)ated isoform of H1 histone and/or long and branched protein-free ADP-ribose polymers could act as protecting agents against full methylation of the CpG dinucleotides in genomic DNA.

H1-H1 cross-linking efficiency depends on genomic DNA methylation.

Oligonucleosomal DNA preparations from condensed-inactive chromatin were examined, before and after artificial methylation by bacterial SssI methylase, for their ability to allow cooperative H1-H1 interactions under conditions of different ionic strength. Our results support the conclusion that, within the highly methylated genomic DNA, there are some CpG's whose unmethylated state is critical for chromatin folding. Circular dichroism spectra indicate that artificial overmethylation of native oligonucleosomal DNA reduces its efficiency in inducing an ordered conformation of H1 histone. Temperature melting profiles confirm on the other hand that the native and the artificially overmethylated forms of oligonucleosomal DNA are both able to bind H1 histone.

Autoimmunity to the GM2-1 islet ganglioside before and at the onset of type I diabetes.

Recently, the GM2-1 pancreatic islet ganglioside, proposed as a potential autoantigen in type I diabetes autoimmunity, has been biochemically characterized and found to be a novel ganglioside structure. In the present study, we aimed to determine whether an autoimmune response toward this novel islet molecule is 1) present in type I diabetes and is specifically directed against this molecule and not to gangliosides in general and 2) predictive of disease in high-risk subjects. To this end, the following patients have been studied: 1) 24 newly diagnosed type I diabetic subjects, 20 islet cell autoantibody (ICA)-negative first-degree relatives of type I diabetic subjects, and 25 age-matched normal control individuals; and 2) 31 prospectively evaluated ICA+ first-degree relatives of type I diabetic subjects who were followed for up to 10 years, during which 14 of them developed type I diabetes. A direct assay for autoantibodies to GM2-1 and to other pancreatic gangliosides (GM3, GD3, GD1a) was developed using an indirect immunoperoxidase technique performed directly on thin layer chromatography plates. Anti-GM2-1 autoantibodies (all belonging to the IgG class) were expressed in a high percentage of newly diagnosed type I diabetic subjects (71%), while no significant difference was found in the expression of antibodies directed against other pancreatic gangliosides (GM3, GD3, GD1a) among the different groups studied. Anti-GM2-1 autoantibodies were also present in ICA+ relatives (64%) (P < 0.001 vs. control subjects and ICA-relatives): in this group, life table analysis of progression to diabetes showed that anti-GM2-1 autoantibodies were significantly (P < 0.001) associated with disease, occurring in all relatives developing type I diabetes within 5 years and thus identifying a cohort of ICA+ subjects with markedly increased diabetes risk.

[Lesions of the collateral ligaments of the ankle: diagnosis and follow-up with magnetic resonance and ultrasonography]. / Le lesioni dei legamenti collaterali del collo del piede: diagnosi e follow-up con risonanza magnetica ed ecografia.

Acute sprains are one of the most frequent ankle conditions; the lateral collateral ligaments are often involved. Currently, plain radiography and clinical examination are the methods of choice in the management of these injuries. This study was aimed at describing the MR findings of acute ankle sprains, to assess which ligaments are involved, to study the repair process during conservative treatment and, finally, to compare MR and US findings in acute and chronic injuries. We divided our study into a prospective and a retrospective parts. In the prospective study, MRI was performed in 20 consecutive patients with acute ankle sprain diagnosed at the emergency care unit of our institute and treated conservatively with braces. The patients with fractures were excluded. Follow-up was based on a series of MR exams performed every 30 days for 6 months. We diagnosed 18 injuries of the lateral collateral ligaments, 5 of the anterior talofibular ligament (ATFL) (2 partial and 3 complete tears) and 13 of both the ATFL and the calcaneofibular (CFL) (6 partial and 7 complete tears). The follow-up showed complete edema resorption at 30 days in 2 patients with no ligament injuries; the persistence of a medium-large amount of fluid in partial tears at 30 days and its complete resorption at 60 days; and, finally, the persistence of a medium-large amount of fluid at 180 days in 2 complete tears. No scar could be identified in all the patients with ligament injuries. The retrospective study was based on the comparison of MR (gold standard) and US findings in 78 patients with ankle sprain, 28 in the acute and 50 in the chronic phase. In the first group we found 9 ATFL injuries (6 partial and 3 complete tears), 5 ATFL and CFL injuries (3 partial and 2 complete tears), 2 complete tears of lateral collateral ligaments, 3 deltoid ligament (DL) injuries, 2 ATFL and DL injuries and 2 injuries of both lateral and medial collateral ligaments. US was in agreement with MRI in 85% of ATFL, 67% of CFL and 28% of DL injuries; US also yielded 2 false positives in PAA. In the second group of 50 patients, MRI showed 11 ATFL and 8 ATFL and CFL injuries, 3 injuries of lateral collateral ligaments, 6 DL and 8 ATFL, CFL and DL injuries and 5 complete tears of both internal and external collateral ligaments; 10 exams were negative. US had 58% agreement with MRI in ATFL, 46% in CFL and 21% in DL injuries. In this series, US yielded 3 false positives in PAA injuries.

Co-operative interactions of oligonucleosomal DNA with the H1e histone variant and its poly(ADP-ribosyl)ated isoform.

H1 histone somatic variants from L929 mouse fibroblasts were purified by reverse-phase HPLC. We analysed the ability of each H1 histone variant to allow the H1-H1 interactions that are essential for the formation of the higher levels of chromatin structure, and we investigated the role played by the poly(ADP-ribosyl)ation process. Cross-linking analysis showed that H1e is the only somatic variant which, when bound to DNA, is able to produce H1-H1 polymers; the size of polymers was decreased when H1e was enriched in its poly(ADP-ribosyl)ated isoform. Measurement of the methyl-accepting ability in native nuclei compared with nuclei in which poly(ADP-ribosyl)ation was induced showed that the poly(ADP-ribosyl)ated H1 histone had not been removed from linker regions, in spite of its different interaction with DNA.

Specific inhibitory effect of H1e histone somatic variant on in vitro DNA-methylation process.

H1e and H1c histone variants were purified from mouse L929 fibroblasts using a reverse phase HPLC, and their effect on in vitro DNA methylation was investigated, together with their ability to bind unmethylated or methylated CpG-rich 44bp oligonucleotides. In a "physiological" range of H1:DNA ratios only H1e, at variance from H1c, was found to cause a marked inhibition of in vitro enzymic DNA methylation. It was also shown that both variants have a similar affinity in binding a methylated CpG-rich oligonucleotide, but that the binding to the same oligonucleotide in the unmethylated form occurs preferentially with H1e rather than with H1c. H1e is therefore likely to be directly involved in maintaining CpG-rich sequences in the unmethylated state.

GM2-1 pancreatic islet ganglioside: identification and characterization of a novel islet-specific molecule.

Recent studies have indicated that GM2-1, a pancreatic islet monosialo-ganglioside, is an islet-specific component whose expression is metabolically regulable and represents one of the target antigens of cytoplasmic islet cell antibodies. In the present study we aimed to biochemically characterize this molecule using a panel of biochemical techniques including gas chromatography, thin layer chromatography, enzymatic digestion and mass spectrometry. GM2-1 ganglioside was extracted from human pancreas and purified by thin-layer chromatography. Fatty acids in the ceramide (the hydrophobic portion of the molecule), identified by gas chromatography ranged from C16:1 to C24:1. The oligosaccharide chain was enzymatically digested by the sequential application of various exoglycosidases (neuraminidase followed by beta-galactosidase, followed by beta-hexosaminidase) and characterized by gas chromatography identification of the liberated sugars. The following structure was deducted from enzymatic studies and confirmed by mass spectrometry analysis: N-acetyl neuraminic acid-galactose-galactosamine-galactosamine-glucose-ceramide. This is a novel ganglioside structure, not yet described, which shares characteristics with a neuronal glycolipid autoantigen: the LM1 ganglioside. Both GM2-1 and LM1 have a single sialic acid residue in the terminal position, the same migration position on thin layer chromatography and the same number of carbohydrate moieties. In conclusion, we have characterized a novel islet-specific ganglioside molecule with unusual characteristics, such as the terminal sialic acid and the galactosamine residues, which may facilitate both its antigenicity and its involvement in beta-cell autoimmunity.

[CT-guided percutaneous treatment of osteoid osteoma]. / Trattamento percutaneo dell'osteoma osteoide sotto guida con Tomografia Computerizzata.

CT was used to localize and guide the percutaneous ablation of osteoid osteomas in 11 patients whose age ranged 5 to 63 years. The treatment was performed directly in the CT room ensuring maximum asepsis. General anesthesia was used in children and in vertebral and sacroiliac localizations, while the peripheral block was used in peripheral localizations. In the latter cases, an ischemic band was used to reduce bleeding. A dedicated drill resection system guided by a Kirschner guide wire was used for removal. The treatment was curative in the short period in all the patients, with complete symptom remission. Only one patient required retreatment after 6 months. In our series of patients no major complications, e.g., bleeding or infections, were observed. In 8 cases the resection yielded enough material for histology. To conclude, this technique can be considered a valuable alternative to surgery in the treatment of osteoid osteomas.

Does hypomethylation of linker DNA play a role in chromatin condensation.

The inhibitory effect that H1 histone exerts on the in vitro DNA methylation process, catalysed by mammalian DNA methyltransferase, together with the relative hypomethylation of linker DNA in eukaryotic cells chromatin, suggest that this hypomethylated state of linker DNA can be of importance in allowing or regulating H1-dependent chromatin condensation. In native oligonucleosomes (olnu), i.e., in chromatin fragments consisting of 5-20 nucleosomes each, there was a correlation between the effects of H1 on the DNA ellipticity at 280 nm and the in vitro assayed methyl-accepting ability. The same was true in H1-depleted or in H1-reconstituted preparations. Artificial methylation caused olnu DNA to lose its ability to allow cooperative H1-H1 interactions under ionic strength conditions similar to those known to affect the transition of the 10-nm filament to the 30-nm chromatin fiber. These results suggest that hypomethylation of linker DNA plays a role in the H1-H1 interactions that are needed for solenoid condensation.

Specific variants of H1 histone regulate CpG methylation in eukaryotic DNA.

Upon HPLC fractionation of human placenta or calf thymus H1 histone preparations, only some fractions enriched in the H1e-c variants were able to exert a severe inhibition on in vitro enzymatic DNA methylation. These fractions, though similar to the other variants in interacting with genomic DNA, were also the only ones which could bind CpG-rich ds-oligodeoxyribonucleotides (oligos). Both the 6-CpG ds-oligo and the DNA purified from chromatin fractions enriched in 'CpG islands' were good competitors for the binding of H1e-c to the 6meCpG ds-oligo. This ability to bind any DNA sequence and to suppress the enzymatic methylation in any sequence containing CpG dinucleotides suggests, for these particular H1 variants, a possible role in maintaining CpG island DNA and linker DNA at low methylation levels.

Binding of histone H1e-c variants to CpG-rich DNA correlates with the inhibitory effect on enzymic DNA methylation.

Within the H1 histone family, only some fractions enriched in the H1e-c variants are effective in causing a marked inhibition, in vitro, of enzymic DNA methylation and, in gel retardation and Southwestern blot experiments, in binding double-stranded (ds) CpG-rich oligonucleotides. Both the 6-CpG ds-oligonucleotide and the DNA purified from chromatin fractions enriched in 'CpG islands' are good competitors for the binding of H1e-c to 6-meCpG ds-oligonucleotide. Because of their ability to bind any DNA sequence and to suppress the enzymic methylation in any sequence containing CpG dinucleotides, these particular H1 variants could play some role in maintaining linker DNA at low methylation levels and even in preserving the unmethylated state of the CpG-rich islands which characterize the promoter regions of housekeeping genes.

[Ultra-low field magnetic resonance in the diagnosis of ovarian lesions]. / La risonanza magnetica a campo ultra basso nella diagnostica delle lesioni ovariche.

To investigate the diagnostic capabilities of ultra low field MRI in the characterization of adnexal conditions, 46 women (aged 17 to 83 years) with US diagnosis of tubo-ovarian diseases were examined with MRI. An 0.064 T permanent magnet was used (toshiba Access), with T1- and T2-weighted spin-echo (SE) sequences, 3D T1-weighted gradient-echo (GE) sequences and, in 5 patients, a STIR sequence. Compared with surgical (37 patients), needle biopsy (3 patients) and clinical follow-up (6 patients) findings, MR diagnostic accuracy was 86.9%, which is comparable with literature data at higher field strength. The most common diagnostic error in our series was in the differential diagnosis between dermoid cyst and endometriosis which exhibited similar MR patterns. This limitation seems to be overcome by using STIR sequences with 60 ms T1 which, at this field strength, can suppress fat signal completely, whereas blood signal intensity remains high. The MR signal patterns of each group of conditions are also reported: in some cases, such patterns differ from those reported at higher field strength due to different T1 and T2 tissue relaxation times.