The adult fast isozyme of myosin is present in a nerve-muscle tissue culture system

Abstract. Organotypic nerve-muscle cultures were prepared from foetal mouse spinal cord and adult mouse muscle fibres. In this system, the adult fibres degenerate and new myotubes form. The regenerated muscle fibres become innervated, develop cross-striations, and survive for several months. We have investigated the isozymes of myosin present in these muscle fibres using histochemistry and immunocytochemistry with antibodies to rat embryonic, neonatal, and adult fast myosins. We demonstrate that some of the regenerated fibres contain adult fast but not embryonic or neonatal myosin. This is the first demonstration of the production of adult myosin heavy chain in tissue culture. This system therefore offers the possibility for further study of the development of adult myosin isoforms in vitro.     

Cloning of the rat insulin-like growth factor binding protein-1 gene and analysis of its 5' promoter region.

To understand specific mechanisms involved in the regulation of insulin-like growth factor binding protein-1 (IGFBP-1), an important modulator of IGF bioactivity, we cloned the rat IGFBP-1 gene and sequenced a 1.5 kb Sph1-Sph1 fragment containing 1110 bases upstream from the translation start site. Computer analysis reveals the presence of ATA, CACCC, and CCAAT elements, and putative homeodomain, AP-1, insulin and glucocorticoid response elements in the 5' promoter. Primer extension and ribonuclease protection studies reveal a single cap site in RNA from rat hepatoma cells and both control and diabetic rat liver.     

Murine hybridomas secreting natural monoclonal antibodies reacting with self antigens

Spleen cells from nonimmunized BALB/c mice were fused with two nonsecreting myeloma lines. The hybrids were selected in HAT medium and screened for Ig production and for antibody activity against actin, tubulin, myosin, thyroglobulin, myoglobin, spectrin, dsDNA, fetuin, and transferrin. Among 161 hybrids secreting Ig, three were found to react with DNA, one with thyroglobulin, and one mainly with myosin. Two of these hybrids could be propagated and further characterized. On the basis of inhibition experiments, one was found to be directed against dsDNA; the other was directed mainly against myosin but at the same time reacted significantly with actin, tubulin, spectrin, and dsDNA. Reactivity with myosin seemed to be concentrated in the light meromyosin subfragment, known to be rich in alpha-helical structure. These results indicate: 1) There are reactive B cell clones directed against self antigens. 2) The antibody specificities found for these antibodies are very similar to those found for natural antibodies in normal human serum and for human monoclonal Ig. 3) The widespread reactivity found for the clone mainly reacting with myosin raises the possibility that the determinant recognized by this antibody is a conformational structure that possibly is associated with alpha-helical structures.     

Modeling the Potential Impact of Host Population Survival on the Evolution of M. tuberculosis Latency

Tuberculosis (TB) is an infectious disease with a peculiar feature: Upon infection with the causative agent, Mycobacterium Tuberculosis (MTB), most hosts enter a latent state during which no transmission of MTB to new hosts occurs. Only a fraction of latently infected hosts develop TB disease and can potentially infect new hosts. At first glance, this seems like a waste of transmission potential and therefore an evolutionary suboptimal strategy for MTB. It might be that the human immune response keeps MTB in check in most hosts, thereby preventing it from achieving its evolutionary optimum. Another possible explanation is that long latency and progression to disease in only a fraction of hosts are evolutionary beneficial to MTB by allowing it to persist better in small host populations. Given that MTB has co-evolved with human hosts for millenia or longer, it likely encountered small host populations for a large share of its evolutionary history and had to evolve strategies of persistence. Here, we use a mathematical model to show that indeed, MTB persistence is optimal for an intermediate duration of latency and level of activation. The predicted optimal level of activation is above the observed value, suggesting that human co-evolution has lead to host immunity, which keeps MTB below its evolutionary optimum.     

Pertussis Prevalence and Its Determinants among Children with Persistent Cough in Urban Uganda

Background

We determined prevalence of pertussis infection and its associated host and environmental factors to generate information that would guide strategies for disease control.

Methods

In a cross-sectional study, 449 children aged 3 months to 12 years with persistent cough lasting ≥14 days were enrolled and evaluated for pertussis using DNA polymerase chain reaction (PCR) and ELISA serology tests.

Results

Pertussis prevalence was 67 (15% (95% Confidence Interval (CI): 12–18)) and 81 (20% (95% CI: 16–24)) by PCR and ELISA, respectively among 449 participating children. The prevalence was highest in children with >59 months of age despite high vaccination coverage of 94% in this age group. Study demographic and clinical characteristics were similar between pertussis and non-pertussis cases. Of the 449 children, 133 (30%) had a coughing household member and 316 (70%) did not. Among 133 children that had a coughing household member, sex of child, sharing bed with a coughing household member and having a coughing individual in the neighborhood were factors associated with pertussis. Children that had shared a bed with a coughing household individual had seven-fold likelihood of having pertussis compared to children that did not (odds ratio (OR) 7.16 (95% CI: 1.24–41.44)). Among the 316 children that did not have a coughing household member, age <23 months, having or contact with a coughing individual in neighborhood, a residence with one room, and having a caretaker with >40 years of age were the factors associated with pertussis. Age <23months was three times more likely to be associated with pertussis compared to age 24–59 months (OR 2.97 (95% CI: 1.07–8.28)).

Conclusion

Findings suggest high prevalence of pertussis among children with persistent cough at a health facility and it was marked in children >59 months of age, suggesting the possibility of waning immunity. The factors associated with pertussis varied by presence or absence of a coughing household member.     

A comparative analysis of intermediate filament proteins in bovine heart Purkinje fibres and gastric smooth muscle

The intermediate filament (IF) composition of muscle cells of various sources is still a controversial issue. In the present study, the IF composition of bovine Purkinje fibres (PFs), atrial and ventricular myocardium, and gastric smooth muscle (SM) has been compared using biochemical and immunocytochemical methods. The Mr of the major IF subunit protein in all four tissues was 55,000. In two-dimensional (2-D) electrophoresis gels of Triton-treated ordinary atrial and ventricular myocardium and the gastric muscular wall, two or three isoelectric isoforms were seen, whereas in PFs up to seven isoforms caused by phosphorylation were observed. In immunofluorescence studies antibodies against the Mr 55,000 subunit of PFs and gastric SM, respectively, both showed identical reactivity with PFs, atrial and ventricular myocytes, gastric SM cells and some SM cells in intramyocardial and gastric muscular wall blood vessels. A small amount of vimentin (Mr 57,000) was also detected in 2-D gel electrophoresis in all four tissues as well as in immunoblotting of PFs with antibodies to vimentin. Immunofluorescence studies using both polyclonal and monoclonal antibodies to vimentin showed that vimentin was present in the endothelium and SM cells of both intramyocardial and gastric muscular wall vessels, sometimes together with desmin in the vascular SM cells, but was never seen in PF, atrial, ventricular or gastric SM cells proper. As expected, vimentin was present in interstitial tissue, i.e., fibroblasts and capillaries. However, interestingly, the monoclonal antibodies, which recognized different antigenic determinants of vimentin, did not give identical staining patterns. Especially the staining of the vascular SM cells differed. Since this staining pattern did not change upon denaturation and unmasking experiments, it seems that the organization of vimentin in different mesenchymal cell types varies. Vimentin was also detected in isolated PFs but here it was located solely in the contaminating interstitial tissue. Thus, desmin is the sole IF protein expressed in PFs, in atrial and ventricular myocytes and in gastric SM cells proper; vimentin alone being present in the interstitial tissue cells, whilst in vascular SM cells desmin and vimentin are coexpressed in various proportions. The variation in number of isoforms of desmin and the heterogeneity in staining of mesenchymal tissues with monoclonal vimentin antibodies probably indicates that the IF cytoskeletons are differently organized in various cell types, even though they contain IFs of the same class.     

Purification, characterization and antimicrobial properties of an antibiotic (wassumycin) from Streptomyces species 215

A yellow amorphous compound, wassumycin, was isolated from an unidentified strain of Streptomyces. Wassumycin inhibited the growth of several species of human and animal pathogenic bacteria especially Clostridia. Wassumycin also inhibited the growth of HeLa cells, and showed good antitumour activity against experimental mouse tumours such as L1210, P388 leukaemias, and B16 melanoma. On the basis of the chemical and antimicrobial properties, the antibiotic appeared similar to the chromomycins and olivomycins, but its ultraviolet, infrared, and proton magnetic resonance spectra differed from theirs significantly.     

Combining distances of ballistic and myrmecochorous seed dispersal in Adriana quadripartita (Euphorbiaceae)

The separate contributions of different vectors to net seed dispersal curves of diplochorous systems have rarely been characterised. In Australia, myrmecochory is a common seed dispersal syndrome and in the majority of such systems, seeds are initially dispersed ballistically. We measured ballistic and myrmecochorous seed dispersal distances in relation to canopies of Adriana quadripartita (Euphorbiaceae) and used a simulation model to estimate the net dispersal curve. We also compared seed removal rates and ant abundances under, and outside, plant canopies to examine how foraging patterns by ants may affect net dispersal.Overall ant abundance did not show a significant numerical response to seedfall; however, the abundance of the main seed dispersing ant, Rhytidoponera ‘metallica’ did. Despite this, seed removal rates did not differ significantly between canopy and open locations. Rhytidoponera ‘metallica’ account for 93% of observed seed dispersal events. On average, the ants dispersed seeds 1.54 m and in doing so, moved seed a mean radial distance of 0.76 m away from canopy edges. This contribution to net dispersal distance by ants is considerable since ballistic dispersal moved seeds a median distance of 7.5 cm. Our simulation model indicated that the combination of ballistic and ant seed dispersal is expected to result in seeds being transported a median net radial dispersal distance of 1.05 m from the canopy edge.Thus in this system, an important function of diplochory may simply be to move a higher proportion of seeds from under the canopy of parent plants than is possible by ballistic dispersal alone. This ‘dispersal-for-distance’ may result in reduced parent–offspring competition or may increase the probability that seeds reach rare safe sites for germination and recruitment.