Essential role for the Legionella pneumophila rep helicase homologue in intracellular infection of mammalian cells

We have previously isolated 32 mutants of Legionella pneumophila that are defective in the infection of mammalian cells but not protozoa. The mutated loci have been designated macrophage-specific infectivity (mil) loci. In this study we characterized the mil mutant GK11. This mutant was incapable of growth within U937 macrophage-like cells and WI-26 alveolar epithelial cells. This defect in intracellular replication correlated with a defect in cytopathogenicity to these cells. Sequence analysis of the GK11 locus revealed it to be highly similar to rep helicase genes of other bacteria. Since helicase mutants of Escherichia coli are hypersensitive to thymine starvation, we examined the sensitivity of GK11 to thymineless death (TLD). In the absence of thymine and thymidine, mutant GK11 did not undergo TLD but was defective for in vitro growth, and the defect was partially restored when these compounds were added to the growth medium. In addition, supplementation with thymidine or thymine partially restored the ability of GK11 to grow within and kill U937 macrophage-like cells. The data suggested that the low levels of thymine or thymidine in the L. pneumophila phagosome contributed to the defect of GK11 within macrophages. Using confocal laser scanning microscopy, we determined the effect of the mutation in the Rep helicase homologue on the intracellular trafficking of GK11 within macrophages. In contrast to the wild-type strain, phagosomes harboring GK11 colocalized with several late endosomal/lysosomal markers, including LAMP-1, LAMP-2, and cathepsin D. In addition, only 50% of the GK11 phagosomes colocalized with the endoplasmic reticulum marker BiP 4 h postinfection. Colocalization of BiP with GK11 phagosomes was absent 6 h postinfection, while 90% of the wild-type phagosomes colocalized with this marker at both time points. We propose that the low level of thymine within the L. pneumophila phagosome in combination with simultaneous exposure to multiple stress stimuli results in deleterious mutations that cannot be repaired in the rep helicase homologue mutant, rendering it defective in intracellular replication.     

Distinguishing characteristics of a new neuroblastoma cell line

The characteristics of a new neuroblastoma cell line (MC-NB-1) established from the bone marrow of a 2-year-old male are described. Morphologically, the cells appear as flattened and epithelial-like or as small and spherical. Electron microscopy demonstrated microtubules and dense core secretory granules. The doubling time was approximately 35 hr. Isoenzyme patterns and catecholamine secretion indicated a human line of neuronal origin. The soft agar tumor colony forming system demonstrated drug resistance in vitro comparable to in vivo nonresponsiveness. The stemline karyotype of MC-NB-1 is 44,Y,del(1) (p22:), -4, -7, +del(7)(q22:), -16, +t(7;16)(16pter leads to 16q24::7q22 leads to 7q32), -17. Additionally, double-minute bodies were observed. However, no evidence of homogeneous staining regions (HSRs) were detected.     

Coloanal anastomosis after resection of low rectal cancer in the elderly

Twenty-two patients had coloanal anastomoses performed after resection of low rectal tumors (16 adenocarcinomas and 6 large degenerated villous adenomas). The patients had an average age of 72 years (range: 62–85). A hand-sewn anastomosis was performed between a J-shaped colonic reservoir and the anal sphincter, except in 3 patients who underwent a straight end-to-end coloanal suture without pouch. A defunctionalized stoma was made in 18 cases. Mortality within 30 days of operation was 9%. Specific complications occurred in 2 patients (9%). Mean follow-up in this series was 20.2±11.7 months (± standard deviation). Fifteen patients were alive and free of disease up to 54 months after surgery. One patient was found to have local recurrence at 19 months. The actuarial 3-year survival rate was 73% in the whole group (operative mortality included). Functional results were good in 16 of 19 patients, with an average 1.8–2.9 stools/24 hours and the continence rate was good. Three patients had complications requiring a permanent colostomy. From these preliminary results, coloanal anastomosis appears to be satisfying in regard to oncologic results and affords good anal function, even in the elderly patient (over 62 years of age).

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Resumen Veintidos pacientes fueron sometidos a anastomosis después de la resecciôn de tumores rectales bajos (16 adenocarcinomas y 6 grandes adenomas vellosos con degeneración carcinomatosa), con edad promedio de 72 años (rango: 62–85). Se practicó anastomosis manual entre un reservorio colónico construído en forma de J y el esfínter anal, excepto en 3 pacientes sometidos a sutura colo-anal términoterminal sin construcción de bolsa. Se practicó un estoma de funcionante en 18 casos. La mortalidad en los primeros 30 días después de la operación fue 9%. Se presentaron complicaciones específicas en 2 pacientes (9%). El seguimiento promedio en la serie fue de 20.2±11.7 meses (± de). Quince pacientes se encuentran vivos y libres de enfermedad hasta por 54 meses. Un paciente demostró recurrencia local a los 19 meses. La tasa actuarial de sobrevida a 3 anos es de 73% para el grupo total (incluyendo la mortalidad operatoria). Los resultados funcionales fueron buenos en 16 de 19 pacientes, con un promedio de 1.8 a 2.9 deposiciones/24 horas y buena continencia. Tres pacientes presentaron complicaciones que requirieron colostomía permanente. Según estos resultados preliminares, la anastomosis colo-anal aparece como un procedimiento satisfactorio en relación con los resultados oncológicos y provee buena función anal aun en el paciente anciano (mayor de 62 años).

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Résumé Vingt-deux patients ont eu une anastomose coloanale après résection d'une tumeur rectale basse (16 adénocarcinomes et 6 tumeurs villeuses larges dégénérées). L'âge moyen des patients était de 72 ans (extrêmes 62 et 85 ans). Dix-neuf patients ont eu une anastomose manuelle entre un réservoir colique en J et le canal anal, alors que 3 ont eu une anastomose manuelle termino-terminale coloanale sans réservoir. Une colostomie de décharge a été ajoutée dans 18 cas. La mortalité des 30 premiers jours postopératoires a été de 9%. Deux patients (9%) ont eu des complications spécifiques. La survie moyenne a été de 20.2±11.7 mois (± ET). A 54 mois, 15 patients sont en vie et sans maladie. Une récidive locale a été décelée chez un patient à 19 mois. La survie actuarielle à 3 ans est de 73% pour l'ensemble des patients (mortalité opératoire incluse). Les résultats fonctionnels étaient bons chez 16 des 19 patients. Ces patients avaient en moyenne 1.8 à 2.9 selles par 24 heures et la continence était bonne. Trois patients avaient des complications nécessitant une colostomie permanente. D'après ces premiers résultats, il semble que l'anastomose coloanale donne de bons résultats du point de vue oncologique avec conservation d'une fonction sphinctérienne satisfaisante, même chez le patient âgé (plus de 62 ans).

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Presented at the Société Internationale de Chirurgie in Toronto, Ontario, Canada, September, 1989.     

Synthesis of some heterocycles of potential biological activity

A convenient method for the preparation of imidazobenzimidazole3, imidazoimidazole5, imidazotriazole6 and pyrano [2, 3-c] oxazole7 derivatives is described. This depends on interaction of 2-methyl-4-arylidene-2-oxazolin-5-ones1 with o-diamines, thiosemicarbazide and/or ethylcyanoacetate. The effect of alcoholic potassium cyanide on oxazolinone1 was studied. Antibacterial activity of the obtained products was studied.     

Ischaemic peripheral polyneuropathy in the upper limb as a result of steal phenomenon in an arteriovenous malformation

We present the case of a 38-year-old right-hand-dominant female with a longstanding extensive high flow arteriovenous malformation (AVM) affecting the left hand and forearm. The patient’s main complaints were altered sensation in the left hand and difficulty with co-ordination. Clinical examination revealed reduced global sensibility in the hand and evidence of a vascular steal phenomenon resulting in a cold thumb and small finger without trophic changes. Subsequent nerve conduction studies identified peripheral neuropathy affecting the median, ulnar and radial nerves. There was no evidence of entrapment neuropathy and the findings were attributed to nerve ischemia as a result of the steal phenomenon. This case represents an unusual complication of an AVM of the upper limb which has not been reported previously to our knowledge. Patients with complex AVMs presenting with atypical peripheral neuropathies should be assessed for the possibility of steal phenomenon.Level of Evidence: Level V, risk/prognostic study.     

Two Different Presentations of Sinus of Valsalva Aneurysm

Sinus of Valsalva aneurysm (SVA) is a rare cardiac anomaly that can be congenital or acquired. We report 2 cases of SVA. The first case involves a 59‐year‐old male presenting with frequent syncope. Echocardiogram revealed a large right SVA obstructing the right ventricular outflow tract (RVOT). The second case involves a 21‐year‐old female presenting with sudden onset chest pain and a continuous machinery murmur. Echocardiogram revealed a ruptured right SVA into the right atrium. Although advanced percutaneous techniques have been implemented in the correction of this anomaly, open‐heart surgery with or without aortic valve replacement remains the treatment of choice.     

The clinimetric properties of the World Health Organization Disability Assessment Schedule II in early inflammatory arthritis

OBJECTIVE: To assess the clinimetric properties of a new health-related quality of life (HRQOL) instrument, the World Health Organization Disability Assessment Schedule II (WHODAS II), in patients with early inflammatory arthritis. METHODS: Internal consistency as well as criterion, construct, and discriminative validity of the WHODAS II were assessed in 172 patients with early inflammatory arthritis who completed the WHODAS II, the Medical Outcomes Study Short Form 36 (SF-36), and other measures of disease severity, functioning, pain, depression, and resource use. Test-retest reliability of the WHODAS II was assessed by having a subset of 20 patients complete the WHODAS II a second time, 1 week after the first assessment. RESULTS: The WHODAS II had high internal consistency (Cronbach's alpha = 0.96 for patients working or in school and 0.93 for patients not working or in school). Test-retest intraclass correlation coefficients of the WHODAS II total score and subscales ranged from 0.82-0.96. The WHODAS II total score was strongly correlated with the SF-36 physical component score (Kendall's tau-b 0.51, P < 0.001) and moderately correlated with the SF-36 mental component score (tau-b 0.43, P < 0.001). WHODAS II correlations with disease outcomes ranged from Kendall's tau-b 0.15-0.55. The WHODAS II significantly differentiated between every aspect of disease severity assessed with the exception of measures of health resource use. CONCLUSION: The WHODAS II is a valid and reliable measure of HRQOL in cross-sectional studies of patients with early inflammatory arthritis. Research is still required to investigate potential item redundancy and determine its usefulness in longitudinal studies.