Transgenic, inducible RNAi in megakaryocytes and platelets in mice.

BACKGROUND: RNA interference (RNAi) is a powerful tool for suppressing gene function. The tetracycline (tet)-regulated expression system has recently been adapted to allow inducible RNAi in mice, however its efficiency in a particular cell type in vivo depends on a transgenic tet transactivator expression pattern and is often highly variable. OBJECTIVE: We aimed to establish a transgenic strategy that allows efficient and inducible gene knockdown in particular hematopoietic lineages in mice. METHODS AND RESULTS: Using a tet-regulated reporter gene strategy, we found that transgenic mice expressing the rtTA (tet-on) transactivator under control of the cytomegalovirus (CMV) promoter (CMV-rtTA mice) display inducible reporter gene expression with unusual and near-complete efficiency in megakaryocytes and platelets. To test whether the CMV-rtTA transgene can drive inducible and efficient gene knockdown within this lineage, we generated a novel mouse strain harboring a tet-regulated short hairpin RNA (shRNA) targeting Bcl-x(L) , a pro-survival Bcl-2 family member known to be essential for maintaining platelet survival. Doxycycline treatment of adult mice carrying both transgenes induces shRNA expression, depletes Bcl-x(L) in megakaryocytes and triggers severe thrombocytopenia, whereas doxycycline withdrawal shuts off shRNA expression, normalizes Bcl-x(L) levels and restores platelet numbers. These effects are akin to those observed with drugs that target Bcl-x(L) , clearly demonstrating that this transgenic system allows efficient and inducible inhibition of genes in megakaryocytes and platelets. CONCLUSIONS: We have established a novel transgenic strategy for inducible gene knockdown in megakaryocytes and platelets that will be useful for characterizing genes involved in platelet production and function in adult mice.

The relationship between caries, fluoridation and material deprivation in five-year-old children in Country Durham.

An investigation to assess the impact of fluoridation on the caries experience of young children at high and low levels of material deprivation measured by the Townsend Index was undertaken in County Durham. Electoral wards were ranked for deprivation from 1991 Census data. Dental caries data for 2,751 five-year-old children resident in electoral wards in the upper and lower quartiles of material deprivation in the County were obtained from a dental survey conducted in 1991-92. Wards were stratified for fluoridated and non-fluoridated water supply and the dental data for the resultant four populations, defined by high and low material deprivation and the presence or absence of fluoridation, were analysed. The mean dmft values in the four groups were 0.8 in the fluoridated low deprivation group, 1.2 in the non-fluoridated low deprivation group, 1.2 in the fluoridated high deprivation wards and 2.1 in the non-fluoridated high deprivation wards. Thus, with fluoridation the variation in caries between high and low deprivation groups was significantly reduced but not eliminated, leaving the level significantly lower in the least deprived group, to the continuing disadvantage of that deprived group.

The use of an index of material deprivation to identify groups of children at risk to dental caries in County Durham.

An investigation of the potential for an index of material deprivation to indicate groups of young children at risk to dental caries and to assess caries experience in their age group at three levels of deprivation, measured by the Townsend Index, was undertaken in County Durham. Electoral wards were ranked on an index of material deprivation derived from 1991 Census data. Dental caries data for the upper quartile, inter-quartile range and lower quartile of material deprivation in the County were obtained from a dental survey of 6052 five-year-old children in 1991-92. The number of children in each group ranged from 1145 to 3058. Significant differences in dental caries experience between high, middle and low ranges of material deprivation existed. The index of material deprivation can indicate groups of children in the community at high and low risk of dental caries.

Dental disease and current treatment needs in a group of physically handicapped children.

One hundred and twenty-nine physically handicapped children (89 males, 40 females) were dentally examined in a local authority residential special school taking pupils from a wide geographical area in the Northern Region, for caries, periodontal disease, malocclusions and treatment need using World Health Organization criteria. The children were aged between 3 and 17 years with a mean age of 10.7 years. Mean deciduous caries experience (dft) was 0.9 and the mean permanent caries experience (DMFT) was 2.0. The mean dfs/DMFS values were 2.5 and 3.4 respectively. A higher experience of deciduous caries was found in girls, dft = 1.3, than in boys who had a mean dft value of 0.8 and in the permanent dentition girls had a mean DMFT value of 2.8 compared with 1.6 for boys. Each component of the index was also higher for girls in both dentitions. The gender difference identified in caries experience extended to treatment need for the permanent dentition, where more girls were recorded as requiring treatment than boys. Of the 129 children, 126 were assessed for treatment co-operation. Only 18 (14.3 per cent) were assessed as being amenable to routine dental care, 66 (52.4 per cent) were thought likely to present some management problems and the remaining 42 (33.3 per cent) were thought to require all their treatment under a general anaesthetic.

A 10-year comparison of General Dental Service care in the Northern Region 1979-1989.

The 1979/1989 patterns of General Dental Service provision in the Northern Region and its nine FPC/FHSA areas have been examined in relation to intraregional variations and assessed against the national average and other regions. Attention is drawn to the Northern Region's failure to reach national averages in a number of key measures of dental care, owing to the low numbers of practitioners working within the Region. It is clear that redistribution of dental manpower will not be generated under the present system. Alternatives are required if dentistry is to continue to be an essential part of a comprehensive health service for all.

The relationship between fluoridation, social class and caries experience in 5-year-old children in Newcastle and Northumberland in 1987.

The dental health of 457 5-year-old children who have lived continuously in fluoridated (at 1.0 mg F/litre) Newcastle and 370 children of the same age in non-fluoridated (less than 0.1 mg F/litre) South Northumberland has been reported. This paper examines in detail the caries prevalence in social class groups I + II, III, IV + V, and the social class/fluoridation relationship in 1987. The prevalence of dental caries in the three social class groupings I + II, III, and IV + V (and the mean dmft), respectively, was 35% (1.1), 46% (1.7) and 67% (2.4) in the fluoridated area, and 59% (2.2), 67% (3.7) and 77% (5.0) in the non-fluoridated area. Fluoridation was effective in all social class groupings and, because caries levels were higher in social classes IV + V, fluoridation brought about greater savings for these children than for those in social classes I + II. Fluoridation reduces but does not eliminate social inequalities, leaving social disadvantage/social background/social class as the major factors in caries prevalence for this age group. Further research into the fluoridation/social class relationship is required, particularly since the understanding of occupational class, now considered an imperfect representation of social class, is being superseded by other criteria which have stronger associations with measures of poor health.