Role of immune cells in protection against and control of reovirus infection in neonatal mice.

We studied the role of T cells in resistance to reovirus intestinal and central nervous system infection. Transfer of reovirus-immune adult spleen cells protected neonatal mice from (i) lethal infection with reovirus serotype 3 Dearing (T3D, footpad inoculation) and serotype 3 clone 9 (T3C9, oral inoculation) and (ii) hydrocephalus caused by serotype 1 Lang (T1L, intracranial [i.c.] inoculation). Cell-mediated protection was not serotype specific. While immune cells protected against T1L i.c., they failed to protect against 1/5,000 of the dose of T3D i.c. Two types of experiments showed that both CD4 and CD8 T cells are involved in reovirus resistance. First, immune cell-mediated protection against T3D was abrogated by in vivo treatment with anti-CD4 monoclonal antibody (MAb) and significantly inhibited by in vivo treatment with anti-CD8 MAb. Second, T3C9-infected neonatal mice treated with anti-CD4 and/or anti-CD8 developed a novel disease phenotype, an oily hair syndrome, associated with severe hepatobiliary pathology and increased viral titer in heart and liver. Immune cells and an MAb to the cell attachment protein sigma 1 (MAb G5) protected by different mechanisms. Immune cells were more effective than sigma 1 MAb G5 at controlling primary replication, while sigma 1 MAb G5 was more effective than immune cells at inhibiting neural spread of virus. We conclude that both CD4 and CD8 T cells are important for reovirus resistance, that cells and antibody act preferentially at different stages in pathogenesis in vivo, and that adoptively transferred immune cells can protect both the central nervous system and intestine.     

Genetic analysis of a mutant class of Physcomitrella patens in which the polarity of gravitropism is reversed.

Summary In the moss Physcomitrella patens, single-cell protonemata and multicellular gametophores respond to reorientation relative to the gravity vector by growing negatively gravitropically. A mutant class in which the protonemata, but not the gametophores, respond by growing towards gravity has been identified. In this paper, we describe the isolation of additional mutants of this class. Complementation and segregation ratio analyses were carried out on these mutants, which indicate that a single gene may mutate to switch the polarity of gravitropism.     

Stable transformation of the moss Physcomitrella patens

Summary We report the stable transformation of Physcomitrella patens to either G418 or hygromycin B resistance following polyethylene glycol-mediated direct DNA uptake by protoplasts. The method described in this paper was used successfully in independent experiments carried out in our two laboratories. Transformation was assessed by the following criteria: selection of antibiotic-resistant plants, mitotic and meiotic stability of phenotypes after removal of selective pressure and stable transmission of the character to the offspring; Southern hybridisation analysis of genomic DNA to show integration of the plasmid DNA; segregation of the resistance gene following crosses with antibiotic-sensitive strains; and finally Southern hybridisation analysis of both resistant and sensitive progeny. In addition to stable transformants, a heterogeneous class of unstable transformants was obtained.     

Oral vaccination of skunks with raccoon poxvirus recombinants expressing the rabies glycoprotein or the nucleoprotein.

Twenty nine skunks (Mephitis mephitis) were vaccinated orally with raccoon poxvirus (RCN) recombinants: 10 with a recombinant expressing the rabies virus glycoprotein (RCNRG), 10 with RCNRG mixed with a recombinant expressing the rabies virus nucleoprotein (RCNRN) and nine with RCN alone. Rabies virus neutralizing antibodies were detected in six of the 20 skunks; five skunks (three given RCNRG, two given a mixture of recombinants) survived a rabies challenge that was lethal for nine skunks vaccinated with RCN alone.     

Presymptomatic testing for Huntington's disease in the United Kingdom. The United Kingdom Huntington's Disease Prediction Consortium.

OBJECTIVE--To evaluate the United Kingdom Huntington''s disease presymptomatic testing programme. DESIGN--Postal questionnaire survey to collect data on all tests performed by clinical genetics centres between 1987 and 1990. SETTING--Genetic centres providing presymptomatic testing in the United Kingdom. SUBJECTS--248 subjects at risk of Huntington''s disease who had presymptomatic testing at their request. MAIN OUTCOME MEASURES--Sex, age, prior risk, and risk after testing. RESULTS--The risk of carrying the Huntington disease gene was reduced for 151 (61%) of the applicants and raised for 97 (39%). 158 (64%) of the subjects were female and 90 (36%) male. The median age at which the results were given was 32.5 years. CONCLUSIONS--The demand for testing was lower than expected and may have reached its peak in 1990. The excess of low risk results was not fully explained by the age effect. All the genetics centres concerned have agreed a common service protocol which requires extensive pre-test counselling and post-test follow up. The worth of the procedure remains to be decided. The availability of a large body of pooled data from all the United Kingdom testing centres, which individually are likely to have only a few results, will form a valuable resource for monitoring the long term psychosocial impact of testing.     

Morphogenesis of the respiratory bronchiole in rhesus monkey lungs

The epithelium of the respiratory bronchiole in the adult rhesus monkey consists of two populations: a pseudostratified epithelium with basal, mucous goblet, and ciliated cells located near the pulmonary artery (PA); and a simple cuboidal epithelium composed only of nonciliated bronchiolar epithelial (or Clara) cells in areas away from the PA. This study describes the pattern of differentiation of these two epithelial populations, and their relationship to the PA and to the time of appearance of alveoli in the respiratory bronchiole of the rhesus monkey during the period of 90-125 days gestational age (DGA). These events were related to changes in the adjacent parenchyma. Dissected airways of infusion-fixed, critical-point-dried lungs were evaluated by scanning microscopy followed by light microscopy of the same airways. At 54% of gestation (90 DGA), the distal airway was lined by a mixture of ciliated and nonciliated cells. By 67% of gestation (110 DGA), the ciliated cells were confined to the epithelium over the PA. The underlying connective tissue initially was cellular containing few fibers but was fibrous by 76% of gestation (125 DGA). Alveolarization began near the most distal cartilage at 57% of gestation (95 DGA), the same period at which secondary septation occurred in the distal acinus. Thus, alveolarization occurred simultaneously in two centers: 1) the proximal centriacinar region in the vicinity of the most distal cartilage and 2) the distal lung parenchyma. The duration of centriacinar alveolarization was short, approximately 5 days.     

Localization of vitamin D3-responsive alkaline phosphatase in cultured chondrocytes

Alkaline phosphatase activity appears to be altered when chondrocyte cultures are incubated with 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3). This study examined whether the hormone-responsive enzyme activity is associated with alkaline phosphatase-enriched extracellular membrane organelles called matrix vesicles. Confluent, third passage cultures of rat costochondral growth cartilage (GC) or resting zone chondrocytes (RC) were incubated with 1,25-(OH)2D3 or 24,25-dihydroxyvitamin D3 (24,25-(OH)2D3) and enzyme specific activity was assayed in the cell layer or in isolated matrix vesicle and plasma membrane fractions. Alkaline phosphatase-specific activity in the matrix vesicles was enriched at least 2-fold over that of the plasma membrane and 10-fold over that of the cell layer. Matrix vesicle alkaline phosphatase was stimulated by 1,25-(OH)2D3 in GC cultures and by 24,25-(OH)2D3 in RC cultures. The cell layer failed to reveal these subtle differences. 1,25-(OH)2D3 increased GC enzyme activity but the effect was one-half that observed in the matrix vesicles alone. No effect of 1,25-(OH)2D3 on enzyme activity of the RC cell layer or of 24,25-(OH)2D3 on either GC or RC cell layers was detected. Thus, response to the metabolites is dependent on chondrocytic differentiation and is site specific: the matrix vesicle fraction is targeted and not the cells per se.     

Ultrastructure of the frontal organ in Convoluta and Macrostomum spp.: significance for models of the turbellarian archetype

Present models of turbellarian evolution depict the organism with a frontal organ — a complex of glands whose necks emerge at the anterior tip of the body — and therefore imply that this organ is homologous throughout the Turbellaria. However, comparisons of representatives of the Acoela and Macrostomida, two putatively primitive orders of the Turbellaria, show that frontal organs in these two are not similar in ultrastructure or histochemistry. The acoel Convoluta pulchra had a prominent cluster of frontal mucous glands whose necks emerged together in a frontal pore at the exact apical pole of the organism, and an array of smaller glands of at least five other types opened at the anterior end, separately from and ventral to this pore. The frontal organs (Stirndrüsen) of two species of Macrostomum on the other hand, comprised an array of discretely emerging necks of at least two gland types including one with rhabdiform (rhammite) and one with globular mucous secretion granules neither of which emerge at the apical pole. In neither species did the organ appear to be sensory. Our findings indicate a low probability of homology between the frontal glands of the Acoela and Macrostomida.     

Metabolic adaptations in goat mammary tissue during pregnancy and lactation

Metabolic adaptations of goat mammary tissue during pregnancy and lactation were monitored in serial biopsies of the tissue. Changes in the synthetic capacity of secretory cells were studied by combining measurements of enzyme activities with short-term culture of mammary explants to measure lactose, casein and total protein synthesis. By these criteria, the main phase of mammary differentiation began in late pregnancy and was essentially complete by Week 5 of lactation, coinciding with the achievement of peak milk yield. While milk yield declined after Week 5, the activities of key enzymes expressed per mg DNA and the rates of lactose and casein synthesis in mammary explants were maintained over a considerable period. The results suggest that changes in the synthetic capacity of epithelial cells may account for much of the rise in milk yield in early lactation, but are not responsible for the declining phase of milk production characteristic of lactation in ruminants.