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The influence of the fixation procedure on the localization of albumin and transferrin in adult rat liver has been carried out using an indirect immunoperoxidase technique at the light and electron microscopic levels. Perfusion and immersion fixations with different concentrations of paraformaldehyde (with or without addition of glutaraldehyde) have been investigated. According to the mode of fixation (perfusion versus immersion) and the concentration of the fixative, the number of albumin and transferrin containing hepatocytes could vary from 10% to 100%, and different labeling patterns could be observed at the electron microscopic level. For the same concentration of fixative, a perfusion fixation induces a less intense labeling than an immersion fixation. Thus similar results are obtained after immersion fixation in 6% paraformaldehyde + 0.25% glutaraldehyde or after perfusion fixation in 4% paraformaldehyde + 0.025% glutaraldehyde. Similar data are noticed after immersion fixation in 4% paraformaldehyde or after perfusion fixation in 1% paraformaldehyde + 0.025% glutaraldehyde. Moreover, perfusion fixation induced a more fine cell structure preservation than immersion fixations and avoided the appearance of zones of fixation.

The synthesis of transferrin has been investigated in rat hepatocytes during pre- and postnatal development of the liver by ultrastructural immunoperoxidase cytochemistry. Transferrin was detected in fetal and postnatal hepatocytes on the rough endoplasmic reticulum and in the Golgi apparatus, but the amount and distribution of the protein varied according to the stage of the hepatocyte maturation. Between days 13 and 19 of gestation, nearly all hepatocytes exhibited transferrin. After day 20 of pregnancy more and more unstained hepatocytes were detected. Two periods (19th day of gestation and one day after birth) were characterized by an intense labeling of the Golgi apparatus; in contrast, only a small amount of transferrin was located in this organelle at other stages of the rat liver development. The accumulation of transferrin in the Golgi apparatus is interpreted as a transitory storage of the protein. Transferrin could be produced by immature as well as mature hepatocytes.

The location of three plasma proteins (transferrin, hemopexin, and albumin) in hepatocytes was investigated in adult rats. The synthesis of transferrin anf hemopexin has been established by ultrastructural studies showing a labeling of the rough endoplasmic reticulum (RER). By using an indirect immunoenzymatic method with monospecific antibody solutions, the three proteins were detected in the same hepatocyte. The simultaneous presence of different plasma proteins in hepatocytes seems to point to the fact that the synthesis in these cells could be a non-specialized type.

Qualitative and quantitative changes in the liver tissue composition have been studied during prenatal development of the Wistar rat by electron microscopy and stereologic methods. The absolute volume of the fetal liver is multiplied by 84 between days 13 and 20 of gestation. In the meantime, the average hepatocyte volume is multiplied by 1.5 between days 12 and 20. The volumetric fraction of hepatocytes increases from 35% of the volumetric fraction of the liver on day 12 to 66% on day 20 of gestation. The non-hepatocyte cells decrease from 49% on day 12 to 25% on day 20. By days 12 and 13, the rough endoplasmic reticulum and the Golgi apparatus are well differentiated, indicating that young fetal hepatocytes are able to synthesize and export plasma proteins. The volumetric fraction of free ribosomes decreases from 38% of the hepatocytic cytoplasm on days 12 and 13 to 6% on day 20. The mitochondrial compartment occupies about 10% of the hepatocyte cytoplasm. The mitochondria, small and round on days 12, 13 and 14, become oblong from day 18 of gestation. The shape of hepatocytes changes during the prenatal development, from potato-like on days 12, 13, 14 to cubic on day 20, with an intermediate, more spheric, stage on day 18.

Nuclear enlargement, usually due to the existence of abnormally high DNA (ploidy) values, is one of the alterations most often encountered in pathological epithelia, especially in cancerous and precancerous states. Direct DNA measurements on microscopic sections are impossible, and karyometric studies must take the difficult stereologic problem of spheroid sectioning into account. A mathematical model is proposed to describe an aggregate of separate parallel anisotropic prolate spheroids of variable size and eccentricity. This model appears suitable for the study of epithelial nuclei. After a tentative estimation of the biopsy angle, the application of the model to nuclear profile measurements produces the stereologic data of the mean spheroid nucleus: major and minor axes (and their ratio), volume, surface area (and the V/S ratio), and numerical density. Considerable time may be saved by the use of an image analyser with a specific algorithm. Most of the steps are entirely automated, but the user retains the interactive choice of the nuclei to be studied, for instance by separating the different epithelial layers. The profile data are computer-filed and several programs can yield the stereologic estimations and plots of nuclear profile areas. Poolings of cases of similar histologic staging and nearly the same sectioning angle can be used for the profile area plots. The model has been applied to pseudostratified and altered nasal mucosa of nickel workers. The stereologic estimates, mainly the nuclear volume, correlate well with histologic gradings. The nuclear profile area plots are considered useful for an assessment of the nuclear polyploidism. The complete model should be applicable to many other types of functionally or pathologically altered epithelia, particularly for cancer and precancer screenings, when the use of sectioned material is a prerequisite.

Studies were made to compare the impact of immersion fixations with different concentrations of paraformaldehyde (10%, 6% + 0,25% glutaraldehyde, 4% and 1%) on the cellular and subcellular structure preservation. The study was performed on 3 animals. Similar conditions existed for all the preparative steps, since they were done in one operation. Cellular parameters as the volume densities of hepatocytes (VVH), nuclei (VVNH), cytoplasm (VVCYT) and extra-hepatocytic space (VVEX) were assessed by light microscopy on toluidine blue stained semi-thin sections at X 1 000. The volume (VVM), the surface density of mitochondria (SVMO) and their mean profile size (am) were measured at the electron microscopic level (X 15 000). The most striking differences were observed in the volume density (VVM), surface to volume ratio (SV) and mean profile area of the mitochondria (am). Qualitative results revealed that the zone of good electron microscopic observation varied according to the concentrations of the fixative. In conclusion, the best qualitative and quantitative preservation is assumed with 4% paraformaldehyde and at a less degree with 6% paraformaldehyde + 0,25% glutaraldehyde.

The difficulty in taking sweat during heavy physical exercise has drawn the authors into testing a technique of sampling generally used in paediatrics. The fact that the results, which have been dealt with statistically, should coincide with the physiological facts already published allows us to consider a use of the technique in order to investigate the physiological mechanisms in action during sweating under different metabolic conditions.