Quantitation of intrathecal measles virus IgG antibody synthesis rate: subacute sclerosing panencephalitis and multiple sclerosis

A method for quantitating specific anti-viral antibodies in serum and cerebrospinal fluid (CSF) is established using enzyme-linked immunosorbent assay (ELISA). Quantitated antibody levels are used to determine intrathecal specific IgG synthesis rate for the particular antibody. Measles virus was used as a model for validating this quantitative technique: a mutated form of measles virus is a cause of subacute sclerosing panencephalitis (SSPE) and there is a possibility that measles virus is related to the cause of multiple sclerosis (MS). Matched serum and CSF samples were assayed. Concentration of anti-measles IgG was determined and intrathecal measles-specific IgG synthesis rate was calculated. For the SSPE samples, measles-specific IgG synthesis rate was elevated and comprised > 20% of the total intrathecal IgG synthesis rate; these results are consistent with the literature. The ELISA method can be performed routinely, providing a quick, simple, reproducible means of quantitating specific antibody concentrations, with sensitivity greater than 1 nanogram per milliliter. With this method, quantitation of IgG antibodies to any other viral antigen can be reliably and precisely determined.     

Risk stratification using the Society of Thoracic Surgeons Program

In an era of progressive cost containment and public scrutiny, the wisdom of aggressive surgical therapy for high-risk candidates has been questioned. At our center in the previous 24 months, 728 patients with coronary artery disease were entered into The Society of Thoracic Surgeons national database, and the hospital outcomes plus length of stay were analyzed. Patients were separated according to the predicted mortality based on the groupings in The Society of Thoracic Surgeons database: 0 to 5% (453 patients); 5% to 10% (126 patients); 10% to 20% (96 patients); 20% to 30% (17 patients); and 30% and greater (36 patients). There was a close correlation with the predicted rates of mortality. Importantly, the preoperative risk stratification demonstrated a strong correlation with the significant morbidity and excessive length of stay in the highest-risk groups (predicted risk of 20% to > or = 30%). The incidences of the most common complications in the group with the highest predicted risk (> or = 30%) were 28%, renal failure; 33%, ventilator dependence; and 17%, cardiac arrest. In addition, at short-term follow-up (6 to 8 months), a 24.3% mortality was identified in patients with a predicted mortality that exceeded 20%. These data quantify the risks and morbidities associated with the care of seriously ill patients with coronary artery disease and demonstrate the need for professional and public discussions focusing on the association of a high preoperative risk status and the consumption of resources.     

Discontinuation of maintenance therapy in patients with quiescent cytomegalovirus retinitis and elevated CD4+ counts

OBJECTIVE: To determine whether maintenance therapy can be discontinued safety in patients with quiescent cytomegalovirus retinitis (CMVR) and increased CD4+ counts after treatment with highly active antiretroviral therapy (HAART). DESIGN: A prospective observational case series. PARTICIPANTS: Eight human immunodeficiency virus (HIV)-positive patients with quiescent CMVR who were taking HAART and had CD4+ counts above 100 cells/microliter elected to discontinue anti-CMV maintenance treatment. INTERVENTION: Biweekly-to-monthly indirect ophthalmoscopy and fundus photographs, monthly-to-quarterly CD4+ counts, and quarterly HIV viral loads were ordered. MAIN OUTCOME MEASURES: Twelve previously affected eyes were examined for evidence of recurrent retinitis, which was defined as any retinal whitening, border opacification, or expansion of areas of retinal pigment epithelial (RPE) atrophy greater than 750 microns. Four previously unaffected fellow eyes were observed for new CMVR. RESULTS: There was no reactivation or progression of retinitis in any patient during the mean follow-up interval of 11.4 months (range, 3-16 months). No previously unaffected eye developed CMVR. CD4+ remained elevated in all patients (range, 70-725; mean, 255). The HIV viral load ranged from undetectable to 139,000 copies. CONCLUSION: Discontinuation of maintenance therapy may be considered in patients with HAART-induced elevated CD4+ counts above 100 cells/microliter, prolonged relapse-free intervals during the reconstitution period before CD4+ counts rise above 100 cells/microliter, and completely quiescent retinitis characterized by RPE scarring only. Reduced risks of drug toxicity and drug-resistant organisms are potential benefits. Close observation for evidence of recurrent retinitis is indicated.     

Expansion and length-dependent fragility of CTG repeats in yeast

Expansion of DNA trinucleotide repeats (TNRs) is the causative mutation in a growing number of human genetic diseases. Large expansions of a CTG tract were obtained and shown by genetic and physical assays to be length-dependent sites of chromosome breakage in Saccharomyces cerevisiae. Deletion of RAD27, which encodes a nuclease involved in Okazaki fragment processing, caused length-dependent destabilization of CTG tracts and a substantial increase in expansion frequency. The genetic assay described here can be used to evaluate other factors that induce TNR expansion or chromosome fragility in humans.     

Protein phosphatases and the regulation of MAP kinase activity

A family with hereditary factor X deficiency is presented. One member, a 25-year-old man, showed a mild bleeding tendency. His factor X activity (extrinsic: 56%; intrinsic: 55%; Russell's viper venom: 57%) and his level of circulating factor X antigen (55% of normal) were markedly reduced. Analysis of his factor X gene revealed a single point mutation within exon II resulting in the substitution of +25 Gla (GAA) by Lys (AAA). The mutation was determined by gene analysis to be heterozygous in this patient, his mother and one of his brothers. Clotting assays of factor X purified from the plasma of the index patient revealed an activity of 89% of normal upon activation with Russell's viper venom, 77% of normal in the intrinsic and 81% of normal in the extrinsic coagulation pathway. The mutation responsible for the substitution of Lys for Gla+25 was introduced into an expression plasmid containing a wild type factor X cDNA and expressed in a mammalian cell line. Factor X antigen levels in the cell lysates and in the supernatant were identical in the mutant and wild type constructs. The specific activity of the factor X expressed from the mutant construct was 3% compared with the wild type construct. These data demonstrate that the substitution of Lys for Gla+25 results not only in a reduced level of factor X in the affected family members, but also in a substantial loss of specific factor X activity.     

Learning curve for oesophageal cancer surgery

BACKGROUND: Surgical training and experience are frequently claimed to influence early and late outcome measures. The aim of this study was to examine any improvement in an individual surgeon's performance in one operation over a period of 7 years from initial appointment to date. METHODS: Patients undergoing Ivor Lewis subtotal oesophagectomy performed by a single surgeon between April 1990 and December 1996 were identified from a prospectively compiled oesophageal cancer database. Operating time (abdominal, thoracic and 'one-lung time'), blood loss, transfusion requirements (intraoperative and total), extent of lymphadenectomy (number of lymph nodes sampled), intensive treatment unit (ITU) stay, hospital stay, postoperative morbidity and mortality, pathological stage, grade and survival were recorded. RESULTS: The records of 150 patients were identified for analysis. The cohort was split into five groups, each of 30 patients operated on consecutively. Each of the groups was comparable for age, sex, smoking history, preoperative haemoglobin and creatinine levels, weight loss, American Society of Anesthesiologists' grade, and histological stage and grade of disease. Analysis of the variables pertaining to operation revealed a significant improvement with time including reduced single-lung operating time (P=0.01), reduced blood loss (P=0.03), reduced transfusion requirement (P < 0.0001), reduced ITU stay (P< 0.0001), reduced inpatient stay (P< 0.0001) and an increased yield of lymph nodes (P < 0.0001). CONCLUSION: This study showed a continuing improvement in a surgeon's performance over a 7-year period. With the current trend to shorter training periods there is a case for continuing supervision of the 'fully trained' surgeon within highly specialist units.     

Current options in prosthetic vascular graft infection

Debate continues over which procedure is the best treatment for prosthetic graft infections. We retrospectively reviewed the medical records at our institution for all vascular graft infections that occurred from 1985 to 1995 to evaluate their occurrence, treatment, and outcome. Twenty-four patients had prosthetic graft infections. The average patient age was 62 years, and 67 per cent of the patients studied were men. The initial operation was for treatment of occlusive disease in 92 per cent of the patients, and aortofemoral bypasses were the most common procedures performed (15 of 24 patients, 63%). The average interval from graft implantation to presentation of infection was 29 months. In lower-extremity bypasses, the site of infection was most commonly in the groin (87%). Gram-positive organisms, including coagulase-negative Staphylococcus (32%) and Staphylococcus aureus (28%), were the most frequently isolated bacteria. Thirty procedures were performed for management of the graft infections. Extra-anatomic bypass was associated with no recurrent graft infections. Graft preservation was successful in two cases of early S. aureus infection (less than 1 year after original procedure), and in situ graft replacement was successful in all four cases of late-appearing coagulase-negative Staphylococcus infection (more than 1 year after original procedure). Both treatments failed in all five cases of Gram-negative infection (P = 0.008 by Fisher's exact test). The overall mortality and amputation rates were 17 per cent and 21 per cent, respectively, without significant differences between the treatment modalities. Extra-anatomic bypass remains the best treatment for prosthetic graft infection. In situ replacement and graft preservation treatments should be selective and based on presentation of the infection and the type of pathogenic organism.     

The detection of oligonucleotide N3'-->P5' phosphoramidate/RNA duplexes with capillary gel electrophoresis

Oligonucleotide N3'-->P5' phosphoramidates (3'-phosphoramidates) are DNA analogs that are presently under investigation as potential therapeutic agents. These compounds may also hold promise as a diagnostic tool. Here, we describe a rapid method for the analysis of single-stranded RNA fragments utilizing 3'-phosphoramidate oligonucleotides as probes in conjunction with capillary gel electrophoresis (CGE). 3'-Phosphoramidate 9-mers were mixed with complimentary RNA, and CGE was used to monitor duplex formation. Complimentary strands of RNA and 3'-phosphoramidate formed duplexes that gave unique relative mobilities based on an internal standard. The ability of CGE to discriminate between perfect duplexes and duplexes that contain a base mismatch was also investigated. However, the primary focus of this work was to determine CGE's ability to detect the presence of the 3'-phosphoramidates/RNA duplex under routine electrophoretic running conditions. Polyacrylamide gel electrophoresis analysis was utilized to verify duplex formation.