Feulgen-DNA cytofluorometry is a very useful methodology for the quantitative analysis of individual cellular DNA content. This method, therefore, has been applied to cytofluorometrical studies of various human organs in our laboratory. In our recent studies including this paper, we have attempted to apply this methodology to the cellular analysis of chondrocytes of the hyaline cartilages such as the articular and epiphyseal cartilages, and in addition, we have quantitatively analyzed the changes of nuclear DNA content of these chondrocytes from the rats in their process of the growth and ageing. However, the most difficult problem encountered in these experiments was how one can isolate many intact chondrocytes from such a hard tissue as the hyaline cartilage in a reasonably short time, to make, without cellular damage, smear preparations appropriate for high-sensitive Feulgen-DNA cytofluorometry. So, we have carried out in this paper some test experiments to search for the most suitable method of cell isolation and its smear preparation for the Feulgen-DNA cytofluorometry of chondrocytes. As the result of extensive studies, we found that both the cell isolation by enzymatic digestions of the cartilage matrix with papain and collagenase followed by mechanical separation and its smear preparation with PBS, are the most suitable procedures for the Feulgen-DNA cytofluorometry of the rat chondrocytes. Further, using these methodologies of the cell isolation and the smear preparation, we have quantitatively analyzed, on the basis of nuclear DNA content per cell, the ploidy patterns of the individual chondrocytes from the articular and the epiphyseal cartilages in the growing rats. It has then been shown that the chondrocytes of the epiphyseal and articular cartilages in the growing rats, consist of many mononuclear diploid cells and a few mononuclear tetraploid cells, and there are noticeably some DNA systhetic cells with intermediate DNA values between diploid and tetraploid levels in the epiphyseal chondrocytes.
Experimental calcification called ‘mastocalcergy’ was elicited in mice. Each mouse was injected with 1.0mg of lead acetate intra-venously followed by subcutaneonu injection of 0.5mg of compound 48/80 (a mast cell discharger). It is generally believed that mast cells play an essential role in ‘mastocalcergy’. We used mast-cell-deficient W/Wv mice in order to evaluate the role of mast cells in this form of calcification. Despite the deficiency of mast cells, the amount of calcification in W/Wv mice was comparable with that in normal congeneic mice. Therefore, it can be concluded that mast cells are not necessarily essential in ‘mastocalcergy’.
Vitamin D-resistant hypophosphatemic osteomalacia associated with 1, 25(OH)2D3 deficiency, hyperphosphaturia, renal pan-aminoaciduria and glucosuria in a 27 year old woman was cured by resection of a benign osteoblastoma in her knee, with concomitant correction of these abnormalities. Correction of serum 1, 25(OH)2D concentration by oral 1α(OH)D3 administration alone raised serum phosphorus level and %TRP, but not to normal levels. A tumor-derived substance is postulated to cause reversible impairment of proximal renal functions, leading to hypophosphatemia due to impaired reabsorption and to 1, 25(OH)2D3 deficiency due to impaired formation, both of which caused osteomalacia. Both intramuscular injections of tumor-homogenate and intravenous infusion of the patient's pre-operative urine in rats increased urinary phosphate excretion significantly. Thus, phosphaturic substance was proved to exist both in the tumor and urine, which supports the above mentioned hypothesis.