HeLa cells as a model to study the invasiveness and biology of Legionella pneumophila

HeLa cells were established as a model system to study the invasiveness and biology of Legionella pneumophila. In this model, invasion could be distinguished from adherence; virulent strains of L. pneumophila were adherent and invasive, whereas nonvirulent strains were adherent but poorly invasive. Invasion was rapid and did not require de novo bacterial protein synthesis, suggesting that the invasion factor is constitutively expressed by virulent strains. Entry into HeLa cells required actin polymerization and an intact microtubule cytoskeleton and was only moderately inhibited by the presence of 100 mM glucose or galactose. Intracellular replication of virulent L. pneumophila took place in ribosome-studded complex endosomes and led to the formation of free bacteria-laden vesicles presumably released from lysed HeLa cells. These free vesicles (referred to as mature vesicles) were isolated in continuous density gradients of Percoll. The bacteria contained in the isolated mature vesicles had a unique envelope structure and were highly adherent to HeLa cells, characteristics that correlated with a bright red appearance after the Giménez stain (Giménez positive). Plate-grown legionellae and replicating legionellae, harboured in complex endosomes, displayed a typical Gram-negative envelope and stained green after the Giménez stain (Giménez negative). Chronically infected cultures of HeLa cells were also established that may be a useful tool for studying long-term interactions between virulent L. pneumophila and mammalian cells. HeLa cells constitute a valuable model system that offers unique opportunities to study parasite-directed endocytosis, as well as stage specific-parasite interactions.     

Role of heat shock proteins in heart transplant rejection

STUDY OBJECTIVE: To study secular trends of placental abruption (PA), the effects of demographic variables and the use of cesarean section (CS) associated with PA. DESIGN: A population based cohort study. SETTING: The Medical Birth Registry of Norway. PATIENTS: 9,592 cases of PA of a total of 1,446,154 births notified, i.e. all births in Norway 1967-1991. MAIN RESULTS: The PA proportion was 6.6 per 1000 births of a gestational age of 16 weeks or more, ranging from 5.3 in 1971 to 9.1 in 1990. Birth order two had the lowest proportion and it increased by maternal age. The PA proportion decreased by gestational age from 86.4 per 1000 below 28 weeks to 3.4 in term pregnancies. The PA proportion per 10,000 pregnancies at risk increased from 1.3 in the 28th week to 14.1 in the 42nd week. A secular trend of a decreasing but still high relative risk of PA in SGA-births at any gestational age increased from 1967 through 1991. The relative risk of PA of Apgar score <7 after five minutes, adjusted for gestational age, was 7.8. CONCLUSIONS: Inspite of an increasing CS rate, an increasing proportion of PA was noted from 1967 through 1991. The proportion was lowest for birth order two and increased by maternal age. To an increasing extent, PA births were centralized. SGA, prematurity and asphyxia were major problems associated with PA. A tendency towards larger infants and a decreasing relative risk of PA in SGA-births might be attributable to improvements in antenatal care.     

Variation in the ratio of physical to genetic distance in intervals adjacent to the Mla locus on barley chromosome 1H

Variants of the pulsed-field gel electrophoresis technique were used in conjunction with two-dimensional DNA gel electrophoresis (2-DDGE) to determine the ratio of physical to genetic distance in two genetically defined intervals on barley chromosome 1H.2-DDGE analysis demonstrated that two loci that define a 0.3 cM interval, as determined by hybridization with BCD249, reside on a single 450-kb MluI fragment. This result indicates a maximum ratio of physical to genetic distance in this interval of 1500 kb/cM as compared to 3.7-4.2 Mb/cM for the barley genome as a whole. High molecular weight (HMW) DNA restricted with NotI and probed sequentially with MWG068 and BCD249 yield diffuse bands at approximately 2.8 Mb and 3.0 Mb in the C.I. 16151 and C.I. 16155 parental lines, respectively. These results suggest the maximum ratio of physical to genetic distance in the interval defined by these probes is 7.8 Mb/cM. Unique HMW DNA restriction fragment length polymorphisms (RFLP) were attributed to the presence of recombination breakpoints. Data from the recombination breakpoint analysis were used to estimate a ratio of physical to genetic distance of 2.5 Mb/cM in the Xbcd249.2-Xmwg068 interval and 0.465 Mb/cM in the Xbcd249.1-Xbcd249.2 interval. Both physical linkage and recombination breakpoint analysis indicate the Xbcd249.1-Xbcd249.2 interval is approximately five-fold smaller, physically, than the Xbcd249.2-Xmwg068 interval.     

Isolation of genomic fragments from polymorphic regions by representational difference analysis

Representational difference analysis is an effective technique for isolating the differences between two nearly identical genomes. We have found the technique to be extremely valuable in our analyses of mouse germline mutations. We also present several technical improvements in the procedure that make it more efficient and reliable.     

Left ventricular geometry in presenting untreated hypertension

In order to investigate the spectrum of geometry in our patient population, 63 untreated hypertensives underwent two-dimensional echocardiography. Left ventricular (LV) mass index and relative wall thickness, a measure of wall thickness in relation to cavity size, were calculated from the M-mode strip. In addition, to assess the sphericity of the left ventricle the ratio of LV minor to major hemiaxis was calculated. The subjects comprised 41 men (17 Caucasian, 22 Afro-Caribbean and two Oriental), and 21 women (five Caucasian, 12 Afro-Caribbean and two Oriental). Concentric hypertrophy was present in 46% of subjects, concentric remodelling in 32% of subjects, eccentric hypertrophy in only 6% of subjects and a normal left ventricular shape in 16% of subjects. The degree of sphericity of the left ventricle was similar among the four groups, suggesting that it does not change in uncomplicated hypertension. In contrast to the previously published combined series from Sassari and New York we had a low proportion of patients with either eccentric hypertrophy or normal left ventricular geometry. This is probably due to the high proportion of Afro-Caribbean subjects in our clinic population who are more likely to have left ventricular hypertrophy.