D. R. Appleton

The metaphase arrest technique. A critical review.

The accuracy and precision of the stathmokinetic experiment for the determination of cell birth rate are discussed from practical and theoretical viewpoints. Topics covered include the determination of the optimal dosage of the metaphase arresting agent, the times at which observations should be taken and which cells should be counted, the correct method of estimating cell birth rate and the dependence of any derived estimate of the turnover time on the age distribution of cycling cells. Failure to consider any of these points can lead to considerable inaccuracy. It is further shown that the stathmokinetic method is a rather imprecise one for measuring the turnover time and the duration of mitosis.

Cell kinetics in flat (avillous) mucosa of the human small intestine

A new method for the analysis of small-intestinal crypt-cell kinetics using routine peroral diagnostic biopsies is described. Untreated patients with childhood and adult coeliac disease and adults with the gluten-sensitive enteropathy of dermatitis herpetiformis were studied, together with groups of adult and childhood controls. In the classical flat avillous mucosae the increase in crypt size was found to be three-dimensional. The number of proliferating cells per crypt was shown to be markedly increased, and an even greater rise in the crypt-cell production rate was demonstrated. A significant increase in the mitotic index was also confirmed in the avillous mucosae. On the basis of these findings it is suggested that the characteristic crypt morphology in glutensensitive enteropathy can be explained as an adjustment to accommodate the expanded mass of proliferating and maturing cells necessary to support the augmented cell production rate. We may speculate that this in turn is a response to a pathologically rapid loss of cells from the mucosal surface.

Nutritional intake, height and weight of 11-12-year-old northumbrian children in 1990 compared with information obtained in 1980

There is little age-specific information on changes in dietary intake over time in this country, yet this is valuable in assessing the effectiveness of health education programmes particularly in vulnerable groups such as adolescents. In 1990, 379 children aged 12 years completed two 3 d dietary records. They were interviewed by one dietitian on the day after completion of each diary to verify and enlarge on the information provided and, with the aid of food models, obtain a quantitative record of food intake. Nutrient intake was calculated using computerized food tables. These children attended the same seven Middle schools in Northumberland as 405 children of the same age who recorded their diet using the same method, 10 years previously. Heights and weights were also recorded in both studies in the same manner. Comparing the nutrient intakes in 1990 with 1980, energy intake fell in the boys (to 8.6 MJ) but not in the girls (8.3 MJ). The contribution of fat to energy intake was unchanged at about 40% (about 90 g/d). Likewise, intake of sugars was unchanged at about 22% of energy (about 118 g/d). Calcium intake remained the same in the girls (763 mg/d in 1990) but fell in the boys (786 mg/d in 1990). Iron, vitamin C and unavailable carbohydrate intakes increased in both sexes, and the nutrient density of the diet improved in all sex and social-class groups. However, a social trend evident in 1980 still existed in 1990 with low social groups having the poorest-quality diet. It is concluded that there is little evidence of substantial progress towards improving the diet of adolescents in this country.

VARIATION IN THE DURATION OF MITOSIS IN THE CRYPTS OF LIEBERKUHN OF THE RAT; A CYTOKINETIC STUDY USING VINCRISTINE

The variation in the duration of mitosis (tm) with cell position in the small intestinal crypts of the adult rat has been measured by a stathmokinetic technique using vincristine. The value for the whole crypt column was 0.43 hr, or 26 min. At the bottom of the crypt tm was in excess of 1 hr, but rapidly decreased and throughout the greater part of the proliferative compartment was between 0.40 and 0.50 hr. At the top of the proliferative compartment an increase in tm was demonstrated.

A 2-year longitudinal nutritional survey of 405 Northumberland children initially aged 11.5 years.

Children (405), initially of average age 11.5 years, recorded all food and drinks consumed for three consecutive days (with an interview on the fourth day) on five separate occasions over a 2-year period. Food tables (Paul & Southgate, 1978) enabled nutrient intakes to be calculated. The data collected were found to be of high reliability (Hackett et al. 1983). The mean energy intakes showed broad agreement with other recent British dietary surveys but were well below those recorded in the prewar study by Widdowson (1947) and the FAO/WHO (1973) recommended levels. They were slightly below the current Department of Health and Social Security (DHSS; 1979) recommended intakes. Over the 2-year period, the energy intake of the boys increased by 13% compared with an increase of only 7% in the girls. The iron and vitamin A intakes of all groups of children were low compared with current recommendations (DHSS, 1979). This seems to be a result of falling energy intake. Mean calcium intakes were also only marginally in excess of the recommended intake, and those of most of the girls would fall below the recommendation if the proposal to end the fortification of flour (DHSS, 1981) is implemented.

Studies on the mechanism of diurnal variation of proliferative indices in the small bowel mucosa of the rat.

In the rat small bowel mucosa significant variation was found in both the labelling and the mitotic indices with time of day. The zenith and the nadir of labelling and mitotic activity coincided at 15.00 and 02.00 hours respectively. Small changes were found in the ‘cut‐off’ position, but this variation in proliferative compartment size was insufficient to account for the comparatively wider fluctuations in proliferative indices. Measurements of the rate of entry into mitosis, using metaphase arrest with vincristine at three widely separated times during the day, showed no significant change.

Cell population kinetics in the mouse jejunal crypt

SummaryThe time parameters of the cell cycle were determined in the jejunal crypts of male Balb/c mice by an FLM experiment. The cell cycle time was 12.42 ±0.11 h, and the duration of DNA synthesis was 7.61 ±0.09 (mean values ± S.E.). A stathmokinetic technique using vincristine gave a value of Tc of 11.8 h.The growth fraction (Ip) calculated from cycle parameters and the labelling index was 0.61, while a value of Ip estimated from a labelling index distribution curve was 0.65.For the whole crypt a value of 0.86 h was obtained for the duration of mitosis, longer than that in the rat. The mitotic duration was found to vary with cell position, but values of between 0.8 and 1.2 h prevailed throughout the proliferative compartment; values in the basal cell positions appeared shorter. Apparent cell cycle times were longest in the basal cell positions.A value for crypt migration rate calculated from a cumulative birth rate curve was 1.48 cell positions per h, compared with a value of 1.8±0.27 cell positions per h as measured from movement of the 50% peak value on the labelling index distribution curve with time after tritiated thymidine.The crypt cell production rate was calculated from microdissected and squashed crypts to be about 14 cells per crypt per h. There was a total crypt population of 282± 65 (S.E.) cells, of which 172 were proliferative.

The cell cycle time in the flat (avillous) mucosa of the human small intestine

A hyperproductive mucosal state in gluten-sensitive enteropathy has been proposed on the basis of an elevated mitotic index, but this parameter is dependent on the mitotic duration when used as an index of proliferative status. The mitotic duration was therefore measured in two control patients with normal villous mucosae and in two patients with the flat avillous mucosa of untreated gluten-sensitive enteropathy, using two different stathmokinetic techniques with vincristine. No significant difference in mitotic duration was found but values obtained for cell cycle time showed a halving in the flat mucosae. An increased rate of cell production in the small bowel mucosa of untreated gluten-sensitive enteropathy is thus confirmed.

Disturbances in lactate metabolism in patients with liver damage due to paracetamol overdose

Six patients with liver damage following paracetamol overdose, one patient with viral hepatitis and six control subjects were infused with sodium L(+) lactate. In controls the results were analysed using a single compartment model while in paracetamol patients a two compartment system was used to derive the fractional rate removal constant and lactate distribution volume. Forearm arterio-venous differences of lactate were also determined in order to assess the role of voluntary muscle in removal of a lactate load. In paracetamol patients with fractional rate removal constant was decreased to less than half the control value (P less than 0.001) while total distribution volume was similar to the two groups. Fasting lactate concentrations were significantly increased in paracetamol patients due to diminished lactate removal since the endogenous production rate of lactate was not significantly different from controls. A greater proportion of the lactate load was removed in voluntary muscle in paracetamol patients (39%) than controls (17%). Since the balance of lactate removal occurs principally in the liver, the decrease in the fractional rate removal constant in patients following paracetamol overdose indicates a severe derangement of hepatic lactate metabolism with a compensatory increase in lactate metabolism in voluntary muscle.

Cell proliferation at different sites along the length of the rat colon

SummaryThis study has been undertaken in order to compare in detail cell proliferation in the mucosal crypts at several sites along the length of the large bowel of the rat. The techniques which have been used include simple morphometry, calculation of mitotic and tritiated thymidine labelling indices, metaphase arrest with vincristine, and the fraction of labelled mitoses method.Major differences exist in the size and shape of the crypts at different sites. In particular the distribution of the proliferating cells within the crypt varies. Mean cell cycle time ranges from 58 h in the descending colon to 25 h in the caecum; this variation appears to be brought about largely by changes in the duration of G1, the other phase durations remaining relatively constant. There is also variation in cell cycle time and growth fraction at different levels within the crypt; throughout the bowel cells appear to cycle more slowly at the bottom of the crypt, but changes in growth fraction do not display a similarly consistent pattern.Clearly the organisation of cell proliferation in the normal rat colon is very complex, and strict definition of anatomical location is required in any study of cell proliferation whether in normal or in diseased animals.