Expression of alpha-fetoprotein and albumin genes in a rat hepatoma cell line SY/1/80: Evidence for the need of species specific serum factors

The expression of alpha-fetoprotein and albumin genes has been studied in a rat hepatoma cell line (SY/1/80) developed from a liver cell tumor induced with di-ethylnitrosamine. This original tumor produces both proteins. However, in $\text{in vitro}$ propagated hepatoma cells, after passages in growth medium containing new born calf serum, the mRNAs of both proteins were undetectable. Supplementation with rat serum, but not serum from calf, horse or human, in growth medium for this cell line led to resynthesis of albumin and AFPmRNAs. These findings suggest that species specific serum factor(s) play an important role in the regulation of gene expression. Although the nature of the factor(s) and of the mechanism of action remains to be elucidated, this phenomenon may explain the general feature of diminshing abilities of cells to produce specific proteins in continuous subculture using standard calf serum.     

Effect of passage in culture on a clone of BALB/c 3T3 cells transformed by simian virus 40.

Most simian virus 40 (SV40)-transformed BALB/c 3T3 clones employed for biochemical studies have been used without regard to passage level. To determine whether virus-induced properties are stable as a function of passage, we have extensively characterized one transformed clone, FNE, which was isolated after SV40 infection BALB/c 3T3 cells in factor-free medium. From the initial testing at passage 5 and for at least 50 subsequent subcultures, the cells stably maintained many transformed growth properties, including high saturation density, morphology, colony formation on contact-inhibited monolayers, tumorigenicity, and synthesis of viral-specific RNA. However, other properties varied as a function of passage. There was a slight decrease in viral genome equivalents per cell from 1.1 copy/cell at passage 5 to 0.7 copies at passage 40. Initially, the cells were negative for all type C virus; however, cells carried at low density for 13 to 20 passages (65 to 100 generations) began to release an endogenous type C virus that then persisted in the culture. Spontaneous release of type C virus did not occur in control BALB/c 3T3 cells carried under identical culture conditions for 90 passages. When the cultures were releasing type C viruses they stained uniformly and brightly positive for SV40 tumor (T) antigen by immunofluorescence, whereas T antigen staining was variable at early passage. These experiments suggest that subtle but perhaps important differences in viral gene expression can occur as a function of passage; they also demonstrate the importance of evaluating the interactions between SV40 and endogenous type C viruses.     

Mechanisms of siderophore iron transport in enteric bacteria.

Uptake of 55Fe- and 3H-labeled siderophores and their chronic analogues have been studied in Salmonella typhimurium LT-2 and Escherichia coli K-12. In S. typhimurium LT-2, at least two different mechanisms for siderophore iron transport may be operative. Uptake of 55Fe- and 3H-labeled ferrichrome and kinetically inert lambda-cis-chromic [3H]deferriferrichrome by the S. typhimurium LT-2 enb7 mutant, which is defective in the production of its native siderophore, enterobactin, appears to occur by two concurrent mechanisms. The first mechanism is postulated to involve either rapid uptake of iron released from the ferric complex by cellular reduction without penetration of the complex or ligand or dissociation of the complex and simultaneous uptake of both ligand and iron coupled with simultaneous expulsion of the ligand. The second mechanism appears to consist of slower uptake of the intact ferric complex.     

Long-Term Evolution of the Hypervariable Region of Hepatitis C Virus in a Common-Source-Infected Cohort

The long-term evolution of the hepatitis C virus hypervariable region (HVR) and flanking regions of the E1 and E2 envelope proteins have been studied in a cohort of women infected from a common source of anti-D immunoglobulin. Whereas virus sequences in the infectious source were relatively homogeneous, distinct HVR variants were observed in each anti-D recipient, indicating that this region can evolve in multiple directions from the same point. Where HVR variants with dissimilar sequences were present in a single individual, the frequency of synonymous substitution in the flanking regions suggested that the lineages diverged more than a decade previously. Even where a single major HVR variant was present in an infected individual, this lineage was usually several years old. Multiple lineages can therefore coexist during long periods of chronic infection without replacement. The characteristics of amino acid substitution in the HVR were not consistent with the random accumulation of mutations and imply that amino acid replacement in the HVR was strongly constrained. Another variable region of E2 centered on codon 60 shows similar constraints, while HVR2 was relatively unconstrained. Several of these features are difficult to explain if a neutralizing immune response against the HVR is the only selective force operating on E2. The impact of PCR artifacts such as nucleotide misincorporation and the shuffling of dissimilar templates is discussed.     

Simulated beetle defoliation on willow genotypes in mixture and monotype plantations

The effect of simulated beetle damage (0%, 25%, 50% and 75% mechanical defoliation) on 12 willow genotypes, grown in short‐rotation coppice, was studied in a modified criss‐cross experimental design. The design enabled the above‐ground effects of monoculture and mixed planting to be assessed. Repeated measurements were modelled to produce derived variables in terms of time or, more appropriately, in terms of accumulated day length (i.e. ‘developmental time’) units. These derived variables were then analysed using the REsidual Maximum Likelihood (REML) method implemented in GenStat? (2001) . No significant competition effect between the genotypes due to planting regime was detected. Genotypes Salix viminalis × Salix schwerinii‘Beagle’ and S. viminalis × S. schwerinii‘Torhild’ were found to have the greatest rate of increase in leaves regardless of defoliation and also the greatest height prior to defoliation. Genotype Salix dasyclados‘Loden’ showed the highest rate of growth under the stress of defoliation. When assessing height at the end of the growing season, S. viminalis × S. schwerinii‘Olof’ was the highest genotype for 25% and 75% levels of defoliation, but genotypes Salix aurita × Salix cinerea‘Delamere’, Loden and S. viminalis × Salix burjatica‘Ashton Parfitt’ appeared to be most tolerant by having consecutively lower base day lengths (i.e. increasing the accumulation of developmental units and the length of the growing season) for increasing defoliation. Shorter genotypes tended to be more tolerant, but of the higher genotypes reaching a control height of greater than 3 m by the end of the growing season, S. viminalis × S. schwerinii‘Tora’ and Beagle performed best to 50% defoliation.     

HIV/AIDS in Asia： The Shape of Epidemics and Their Molecular Epidemiology

The Asia-Pacific region is a home to 60% of the population in the world and to approximately one quarter of people with HIV/AIDS. Close to a million of people has been infected and a half million people died of AIDS annually in Asia,becoming the second largest epicenter of global AIDS epidemic. Molecular epidemiology has been useful tool to track a course of HIV spread. In-depth knowledge from the studies on molecular epidemiology elucidates the dynamics of HIV spread and the interrelationship of epidemics in the different regions in Asia.     

Coumarins from Malaysian Micromelum minutum

In a continuation of our study of the Rutaceae, detailed chemical investigation on Micromelum minutum (Rutaceae) collected from Sepilok, Sabah, Malaysia gave four new coumarins. The structures of the coumarins have been fully characterised by spectroscopic methods as 3",4"-dihydrocapnolactone 1, 2',3'-epoxyisocapnolactone 2, 8-hydroxyisocapnolactone-2',3'-diol 3 and 8-hydroxy-3",4"-dihydrocapnolactone-2',3'-diol 4.     

The congenital bicuspid aortic valve can experience high-frequency unsteady shear stresses on its leaflet surface

The bicuspid aortic valve (BAV) is a common congenital malformation of the aortic valve (AV) affecting 1% to 2% of the population. The BAV is predisposed to early degenerative calcification of valve leaflets, and BAV patients constitute 50% of AV stenosis patients. Although evidence shows that genetic defects can play a role in calcification of the BAV leaflets, we hypothesize that drastic changes in the mechanical environment of the BAV elicit pathological responses from the valve and might be concurrently responsible for early calcification. An in vitro model of the BAV was constructed by surgically manipulating a native trileaflet porcine AV. The BAV valve model and a trileaflet AV (TAV) model were tested in an in vitro pulsatile flow loop mimicking physiological hemodynamics. Laser Doppler velocimetry was used to make measurements of fluid shear stresses on the leaflet of the valve models using previously established methodologies. Furthermore, particle image velocimetry was used to visualize the flow fields downstream of the valves and in the sinuses. In the BAV model, flow near the leaflets and fluid shear stresses on the leaflets were much more unsteady than for the TAV model, most likely due to the moderate stenosis in the BAV and the skewed forward flow jet that collided with the aorta wall. This additional unsteadiness occurred during mid- to late-systole and was composed of cycle-to-cycle magnitude variability as well as high-frequency fluctuations about the mean shear stress. It has been demonstrated that the BAV geometry can lead to unsteady shear stresses under physiological flow and pressure conditions. Such altered shear stresses could play a role in accelerated calcification in BAVs.