Comparative study of the cells of the freshwater planarian Polycelis tenuis (Iijima) using dissociated fragments cultivated in vitro: ultrastructural aspects and incorporation of (3H) leucine and (3H) uridine

Five cases of a rare type of malignant melanoma of the choroid with a typical clinical and histopathological picture are reported. This tumour is characterized by it location near the optic disk,its early growth through the retina, its structure with bloodless and blood-filled cavernous spaces, and its manifestation by a massive haemorrhage into the vitreous. "Historical" cases are mentioned, showing that this rare tumour, occurring in about 1:250 of malignant melanomas of the choroid, has a Scandinavian tradition. The clinical and histopathological details are discussed.     

Germ cell genotype controls cell cycle during spermatogenesis in the rat

Spermatogenesis is one of the most productive self-renewing systems in the body: on the order of 10(7) spermatozoa are produced daily per gram of testis tissue. In each mammalian species, the time required for completion of the process is unique and unalterable. Because the process is supported by somatic Sertoli cells, it has generally been thought that cell-cell interaction between germ and Sertoli cells controls the duration of cell cycles and cellular organization. We have used the newly developed technique of spermatogonial transplantation to examine which cell type(s) determines the rate at which germ cells proceed through spermatogenesis. Rat germ cells were transplanted into a mouse testis, and the mouse was killed 12.9-13 days after administration of a single dose of [3H]thymidine. The most advanced rat cell type labeled was the pachytene spermatocyte at stages VI-VIII of the spermatogenic cycle. In animals given only rat cells, some endogenous spermatogenesis of the mouse recovered. The most advanced labeled mouse cell types in recipients killed 12.9-13 days after administration of a single dose of [3H]thymidine were meiotic cells or young spermatids, which is consistent with a spermatogenic cycle length comparable to the 8.6 days reported for the mouse. The same results were obtained if a mixture of rat and mouse cells were transplanted. There existed two separate timing regimens for germ cell development in the recipient mouse testis; one of rat and one of mouse duration. Rat germ cells that were supported by mouse Sertoli cells always differentiated with cell cycle timing characteristic of the rat and generated the spermatogenic structural pattern of the rat, demonstrating that the cell differentiation process of spermatogenesis is regulated by germ cells alone.     

Interactions of rat repetitive sequence MspI8 with nuclear matrix proteins during spermatogenesis

Using the Southwestern blot analysis we have studied the interactions between rat repetitive sequence MspI8 and the nuclear matrix proteins of rat testis cells. Starting from 2 weeks the young to adult animals showed differences in type of testis nuclear matrix proteins recognizing the MspI8 sequence. The same sets of nuclear matrix proteins were detected in some fractions enriched in spermatocytes and spermatides and obtained after fractionation of testis cells of adult animals by the velocity sedimentation technique.     

Restoration of spermatogenesis in dibromochloropropane (DBCP)-treated rats by hormone suppression

The exposure of men to the nematocide dibromochloropropane (DBCP) has caused prolonged oligo- and azoospermia, which occasionally reverses spontaneously. We recently demonstrated that in testes of rats treated with a dose of DBCP sufficient to reduce the percentage of tubules producing differentiating germ cells (tubule differentiation index, TDI) to 20%, the tubules lacking differentiating cells contained type A spermatogonia. To determine whether these type A spermatogonia could be stimulated to differentiate, as had been demonstrated previously in other models of toxicant-induced sterility, we suppressed intratesticular testosterone and serum follicle stimulating hormone (FSH) levels with the GnRH agonist Lupron (leuprolide). When the GnRH agonist was given for 10 weeks starting immediately after DBCP exposure, the TDI was maintained at 94%. Even when GnRH-agonist treatment was stopped at week 10, the TDI remained between 65 and 80% 10 weeks later. Late spermatid counts averaged 10 x 10(6) per testis for the GnRH-agonist-treated rats at week 20 compared with 1.7 x 10(6) per testis in rats treated with only DBCP. To determine whether spermatogonial differentiation could be stimulated after the TDI had declined to below 30%, we initiated GnRH-agonist treatment 6 weeks after DBCP exposure. The GnRH treatment increased the TDI to 53% at week 16. These results indicate that, if the same principles apply to humans, suppression of testosterone may be applied to restore spermatogenesis in men rendered azoospermic by DBCP or other reproductive toxicants.     

Protein phosphatase inhibitors induce the release of serotonin from rat basophilic leukemia cells (RBL-2H3)

Oxidation capacities of laccase, manganese peroxidase (MnP) and lignin peroxidase (LiP) from Phlebia radiata were compared using non-phenolic (veratryl alcohol and ABTS) and phenolic (syringaldazine, vanillalacetone and Phenol red) compounds as reducing substrates. The effect of Mn(II) on enzyme reactions was also studied. Highest specific activities were recorded with laccase in the oxidation of phenolic compounds or ABTS and irrespective of Mn(II) concentration. LiP and MnP oxidized all these substrates but only the catalysis of MnP was dependent upon Mn(II). Only LiP clearly oxidized veratryl alcohol. However, Mn(II) interfered with this reaction by repressing veratraldehyde formation. These results point to multiple participation of manganese ions, either as a reducing (Mn(II)) or oxidizing (Mn(III)) agent in the enzymatic reactions.     

Incorporation of 3H-thymidine into the cells of the heart conduction system (sinoatrial and atrioventricular nodes, bundle of His) during cardiogenesis in mice

The number of DNA synthesizing myocytes in the conductive system, as compared with the working myocardium, was determined by means of light autoradiography. At the same time the number of mitosing cells was counted. The indices of labelled nuclei in the sinoatrial and atrioventricular nodes of the embryos after the single and repeated 3H-thymidine injections were significantly lower than in the working ventricle and atrium. During the postnatal development (3-11 days) only in the sinoatrial node the index of labelled nuclei was significantly lower than in the working ventricle. The proliferative activity in all the myocardium parts fell markedly by the 13th day of development. Following 10 3H-thymidine injections with 12 hrs intervals, the indices of labelled nuclei varied from 0.03 to 0.67% in the working ventricle and atrium and from 0.13 to 1.0% in the conductive system of adult mice. During the embryogenesis the index of labelled nuclei in the sinoatrial node was significantly higher than in the atrioventricular one and during the postnatal development-vice versa.     

Nuclear DNA damage during NAD(P)H oxidation by membrane redox chains

The objective of this study was to develop a valid and reliable discriminative index that measures parent satisfaction with the medical care of their infant in the NICU. We developed an initial questionnaire (Item Reduction Questionnaire) by reviewing the literature, surveying 63 NICU clinicians, and interviewing 125 parents of infants in 2 tertiary level NICUs regarding what they liked and disliked about the medical care of their infants. We administered the Item Reduction Questionnaire, which included 154 items, to 60 parents, who rated the frequency and importance of these items. We included the items identified most frequently as sources of dissatisfaction and rated most important in a second, briefer instrument, the Neonatal Index of Parent Satisfaction (NIPS). To measure reliability we administered the NIPS to 47 parents twice, separated by a 1-week interval. We assessed validity by comparing actual to predicted correlations between NIPS scores and other measures: parent's global rating of satisfaction, medical caregiver ratings of mother's satisfaction, medical caregiver ratings of father's satisfaction, and parents' perception of their infant's health status. We also compared mean NIPS scores for parents who did and who did not report incidents when errors occurred in the medical care of the infant. Of 154 items generated, we included 27 in the NIPS. The intraclass correlation between two administrations of the NIPS to the same 47 parents was 0.71. As predicted, there was a high correlation (0.61) between the NIPS score and parent global rating of satisfaction, and much lower correlations with other variables. Mean NIPS scores for parents who did and who did not report errors differed significantly (difference, 14.6; 95% CI around difference, 5.8-23.5; p < 0.001). The NIPS is likely to be a useful measure for discriminating between parents who differ in terms of their satisfaction with the medical care of their infant in the NICU.     

Negative regulation of MAP kinase by diacylglycerol-dependent mechanisms via G protein-coupled receptors in rat basophilic RBL-2H3 (ml) cells

Carbachol and 5'-(N-ethylcarboxamido)-adenosine (NECA), stimulants of G protein-coupled receptors, induce MAP kinase activation in the muscarinic ml receptor-transfected mast cell line, RBL-2H3 (ml) cells. The phospholipase C inhibitor neomycin and the phosphatidate phosphohydrolase inhibitor propranolol augmented MAP kinase activation induced by carbachol and NECA without affecting the antigen-induced MAP kinase activation. Furthermore, the duration of MAP kinase activation induced by carbachol or NECA was also prolonged by neomycin and propranolol. The specific protein kinase C inhibitor Ro 31-8425 enhanced the carbachol- or NECA-induced MAP kinase activation. These findings suggest that the MAP kinase activation mediated by the G protein-coupled receptors is negatively regulated by diacylglycerol and activated protein kinase C(s).     

Inhibition of peroxisomal degradation by 3-methyladenine (3MA) in primary cultures of rat hepatocytes

BACKGROUND: A significant reduction in peroxisomes has been demonstrated in primary cultures of rat hepatocytes. This report demonstrates that 3-methyladenine (3MA), a potent inhibitor of autophagy, inhibits this effect. METHODS: Hepatocytes from male Wistar rats were isolated by a two-step in situ perfusion technique using collagenase and were cultured in Williams E medium. After a 2-hr attachment period (day 0 of culture), the cells were treated with 200 microM bezafibrate (BF), a peroxisome proliferator, and 5 mM 3MA for 3 days. The cells in the culture dish were fixed in situ, stained for catalase, and embedded in Poly/Bed 812. The number and size of peroxisomes in electron micrographs were analyzed morphometrically. RESULTS: After 3 days of culture, the number of peroxisomes had decreased to 30% of the day 0 level. However, the day 0 level was maintained by treatment with 3MA. In BF-treated cells, many autophagosomes were observed, and peroxisomes had proliferated significantly, although they did not exceed the day 0 level. In cells treated with a combination of 3MA and BF, the number and size of peroxisomes had increased remarkably. CONCLUSIONS: These results suggest that 3MA is effective in maintaining both the number and size of peroxisomes in the course of primary cultures of rat hepatocytes. Suppression of peroxisome proliferation by treatment with BF may be regulated by autophagic/lysosomal degradation.     

Effect of Ambilhar (niridazole) on spermatogenesis in guinea-pigs

Niridazole (Ambilhar), a new antibilharzial drug was fed for 5 days to 20 male guinea pigs at 50 mg/kg while testicular and epididymal biopsies were examined histologically for 10 weeks. Only 10 animals survived the treatment. On the 5th day sperm numbers were reduced in testis tubules. On Day 8 variations in tubule size and in degrees of inhibition of spermatogenesis were at their maximum. Some tubules had recovered spermatogenesis at 15 days, and all had at 3 weeks. Guinea pigs are more sensitive to niridazole than are rats.