Administration Of 3H-thymidine Research Articles

Isolation and Characterization of a Putative Intestinal Stem Cell Fraction From Mouse Jejunum

Although there have been many recent advances regarding the biology of intestinal stem cells, the field has been hampered significantly by the lack of a method to isolate these cells. Therefore, the aim of this study was to explore the hypothesis that viable intestinal stem cells can be isolated as a side population (SP) by fluorescence-activated cell sorting after staining with the DNA-binding dye Hoechst 33342. Preparations of individual cells from either whole mucosa or epithelium of mouse jejunum were stained with Hoechst 33342 and propidium iodide and then sorted using fluorescence-activated cell sorting. Cells were characterized using fluorochrome-labeled antibodies to surface markers, intracellular markers, and annexin V to detect early apoptosis. Total RNA was isolated from sorted fractions and used for quantitative real-time reverse-transcription polymerase chain reaction to evaluate the expression of cell lineage markers and the intestinal stem-cell marker, Musashi-1. Adult and neonatal jejunum contain a viable population of cells that shows the SP phenotype and is sensitive to verapamil. This population of cells (from both mucosal and epithelial preparations) includes a CD45-negative fraction corresponding to nonhematopoietic cells, which shows minimal expression of surface markers typically found on stem cells from other tissues and of intracellular markers found in mesenchymal cells. Additionally, these cells were enriched for Musashi-1 and beta1-integrin, were cytokeratin positive, and survived in culture for up to 14 days. The CD45-negative SP fraction, although not pure, represents the successful isolation of a viable population significantly enriched in small intestinal epithelial stem cells.

Regeneration of adrenal zone reticularis during hyperthermia

The appearance of cells with initial corticocyte differentiation signs beneath the internal capsule of the adrenal gland after exposure to exogenous hyperthermia is demonstrated. Ultrastructure of these cells and transformation of their organelles during further differentiation to typical zona reticularis corticocytes are described. The labeling index (administration of3H-thymidine) in these cells is considerably higher that in typical zona reticularis corticocytes.

Uncoupling of 2-fluoro-2-deoxyglucose transport and phosphorylation in rat hepatoma during gene therapy with HSV thymidine kinase.

This animal study investigates the application of positron emission tomography (PET) with tracers of tumour metabolism for monitoring suicide gene therapy with herpes simplex virus thymidine kinase (HSVtk). After transplantation of HSVtk-expressing Morris hepatoma cells into ACI rats, dynamic PET measurements of 18F-labeled 2-fluoro-2-deoxyglucose (FDG) uptake were performed in animals 2 days (n = 7) and 4 days (n = 5) after the onset of therapy with 100 mg ganciclovir (GCV)/kg body weight as well as after administration of sodium chloride (n = 8). The arterial FDG plasma concentration was measured dynamically in an extracorporeal loop and the rate constants for FDG transport (K1, k2) and FDG phosphorylation (k3) were calculated using a three-compartment model modified for heterogeneous tissues. Also, quantification using the metabolic rate of FDG turnover and the standardized uptake value (SUV) was done. Furthermore, the thymidine incorporation into the tumour DNA was determined after i.v. administration of 3H-thymidine. An uncoupling of FDG transport and phosphorylation was found with enhanced K1 and k2 values and a normal k3 after 2 days of GCV treatment. The increase in FDG transport normalized after 4 days whereas the phosphorylation rate k3 increased. Quantification using the metabolic rate or the SUV showed congruent but less sensitive results compared with the modeling approach. The thymidine incorporation into the DNA of the tumours declined to 10.5% of the controls after 4 days of GCV treatment. The data indicate that PET with 18FDG and 11C-thymidine may be applied for monitoring of gene therapy with the HSVtk/GCV suicide system. Increased transport rates are evidence of stress reactions early after therapy. The measurement of thymidine incorporation into the tumour DNA can be used as an indicator of therapy efficacy.

Dynamics of epithelial cells in the corpus of the mouse stomach. III. Inward migration of neck cells followed by progressive transformation into zymogenic cells

The neck cells (or mucous neck cells) present in the neck region and the zymogenic cells (or chief cells) present in the base region of the units in the mouse corpus were examined in the electron microscope (EM) and in radioautographs prepared after administration of 3H-thymidine by single or multiple injections or by continuous infusion for 1-52 days. For these studies, the neck region of the units has been subdivided into three equal segments, respectively named high neck, mid neck, and low neck, while the base region has been similarly subdivided into high base, mid base, and low base. The neck region includes an average of 12.6 neck cells, characterized in the EM by dark, mucous secretory granules that frequently exhibit a light, pepsinogenic core. Continuous 3H-thymidine infusion reveals that neck cells come from pre-neck cells, which are believed to arise in the isthmus region from the undifferentiated granule-free cells through a pre-neck cell precursor stage. The pre-neck cells, characterized by the presence of a few cored secretory granules, migrate inward (i.e., in the direction of the blind end of the units) and enter the neck region to become neck cells. It is estimated that 59% of the neck cells arise from differentiation of pre-neck cells, whereas the other 41% are derived from their own mitoses. Neck cells migrate inward in 1-2 weeks from the high through the mid and low neck segments, while they keep on producing more and larger secretory granules and thus further differentiate as mucus-producing cells. When neck cells reach the high base segment, they become pre-zymogenic cells that produce secretory granules in which appear light, irregular, pepsinogenic patches which encroach on the dark mucous content. With time, the pre-zymogenic cells, of which there are 5.0 per unit on the average, keep on producing new granules with larger and larger light patches, so that in the end the cells produce granules which are entirely filled by light, pepsinogenic material. At this stage, the cells are zymogenic cells. Zymogenic cells, which average 67.5 per unit, further migrate inward, while gradually enlarging and producing pepsinogenic granules of increasing size. In the low base segment, some zymogenic cells show signs of degeneration leading to death by either necrosis or apoptosis.(ABSTRACT TRUNCATED AT 400 WORDS)

Cell proliferation rate and tumor volume in human osteosarcoma during exposure to methotrexate

The growth inhibitory effect of methotrexate (MTX) on osteosarcoma cells was studied in dysthymic nude mice bearing tumor transplants obtained from a patient before (PRE-CHEM) and after (POST-CHEM) preoperative chemotherapy for osteosarcoma of the distal femur. Cell proliferation was analyzed by autoradiographic evaluation of the fraction of labeled cells after continuous administration of 3H-thymidine for seven days. Histomorphometric analysis of the tissue distribution of cells in the partly ossified tumors was performed. The PRE-CHEM sarcoma transplants showed a significant reduction of labeled interphases from 52 to 1.7 percent upon daily MTX treatment of the mice as compared to controls. In contrast, MTX treatment did not inhibit cell proliferation in the POST-CHEM tumor transplants in which approximately 70 percent of the cells were labeled. Tumor volume increased by 65 and 54 percent in the MTX-treated PRE- and POST-CHEM groups, respectively. During the same eight-day period, control transplant volume increased by 30 percent (PRE-CHEM) and 20 percent (POST-CHEM). Tumor cell densities in the MTX-treated groups were reduced by a factor of approximately 11 in the PRE-CHEM transplants and by a factor of approximately 1.5 in the POST-CHEM transplants. The results show that in this patient the osteosarcoma cells had changed their responsiveness to MTX during the preoperative chemotherapy period. In both the MTX-sensitive and non-sensitive tumor lines, exposure to MTX induced increased tumor volume by increasing the extra cellular matrix volume, irrespective of the neoplastic cell proliferation rate. This effect of MTX was most pronounced in the MTX-sensitive tumor line. These results indicate that in the clinical situation it is difficult to judge the response to chemotherapy even from morphologic parameters.

Tissue distribution, quantitation and proliferation kinetics of fat-storing cells in carbon tetrachloride-injured rat liver

In this study, we have investigated the cell population kinetics of fat-storing cells in livers of rats intoxicated with CCl4. Fat-storing cells were identified in cryostat sections by immunoperoxidase staining of desmin. The peroxidase label was visualized using diaminobenzidine/hydrogen peroxide containing Ni2+ and Co(2+)-ions (Nico/diaminobenzidine method). In normal rats, we found 12.8 fat-storing cells/0.1 mm2 in periportal areas vs. 9.4 in pericentral fields. After one injection of CCl4, the number of pericentral cells increased gradually to reach a maximum of 39.4 cells/0.1 mm2 96 hr after injection. The desmin staining intensity of the pericentral fat-storing cells increased from 48 hr onward. At 72 to 120 hr, strongly stained cells were observed in pericentral areas and in bands of tissue between adjacent central veins, reminiscent of the connective tissue septa in fibrotic livers. In the periportal areas the number of fat-storing cells was not altered. After a second and third injection of CCl4, the number of cells increased further in the pericentral areas. When more than three injections were given, the pericentral fat-storing cell population reached a new steady state with the cell number being seven times higher than in control animals. Proliferation of fat-storing cells at different stages of CCl4 intoxication was studied by intravenous administration of 3H-thymidine, followed by combined desmin staining and autoradiography. Autoradiographical labeling of fat-storing cells was nearly absent in control animals and at 24 hr after a single CCl4 injection. At 48 to 96 hr, labeling indices of pericentral fat-storing cells were significantly higher than in control animals, with a maximum at 72 hr when 22.9% of the cells were labeled. After multiple injections of CCl4, labeling indices between 4.9% and 8.4% were found. We conclude that fibrogenesis is preceded by a strong expansion of the fat-storing cell population in the pericentral areas of the liver lobules and in bands of tissue between adjacent central veins. Local proliferation is an important mechanism underlying the expansion of this cell population.

Examination of potential mechanisms of carcinogenicity of 1,4-dioxane in rat nasal epithelial cells and hepatocytes

Several long-term studies with 1,4-dioxane (dioxane) have shown it to induce liver tumors in mice, and nasal and liver tumors in rats when administered in amounts from 0.5 to 1.8% in the drinking water (Argus et al. 1965; Kociba et al. 1974; National Cancer Institute, 1978). In order to examine potential mechanisms of action, chemically-induced DNA repair (as an indicator of DNA reactivity) and cell proliferation (as an indicator of promotional activity) were examined in nasal turbinate epithelial cells and hepatocytes of male Fischer-344 rats treated with dioxane. Neither dioxane nor 1,4-dioxane-2-one, one of the proposed metabolites, exhibited activity in the in vitro primary rat hepatocyte DNA repair assay, even from cells that had been isolated from animals given either 1 or 2% dioxane in the drinking water for 1 week to induce enzymes that might be responsible for producing genotoxic metabolites. No activity was seen in the in vivo hepatocyte DNA repair assay in animals given a single dose of up to 1000 mg/kg dioxane or up to 2% dioxane in the drinking water for 1 week. Treatment of rats with 1.0% dioxane in the drinking water for 5 days yielded no increase in liver/body weight nor induction of palmitoyl CoA oxidase, indicating that dioxane does not fit into the class of peroxisomal proliferating carcinogens. The percentage of cells in DNA synthesis phase (S-phase) was determined by administration of 3H-thymidine and subsequent quantitative histoautoradiography. The hepatic labeling index (LI) did not increase at either 24 or 48 h following a single dose of 1000 mg/kg dioxane.(ABSTRACT TRUNCATED AT 250 WORDS)

Proliferative Response of Different Exocrine Pancreatic Cells after Surgical Pancreaticobiliary Diversion in the Rat

Pancreaticobiliary diversion (PBD) is known to induce chronic, endogenous hypercholecystokininemia causing pancreatic growth in rats. In the present study the proliferative response of the different exocrine pancreatic cells was studied by administration of 3H-thymidine, 1 mCi/kg, given 1 h before the rats were killed and 5, 10, 20, and 40 days after PBD. DNA and 3H-thymidine uptake, both expressed per 1 mg of pancreatic tissue, were significantly increased on day 5. The nuclear labeling index was increased fivefold in both the acinar and ductal cell group. In the centroacinar cell group the labeling index was increased on day 10. In conclusion, we found that the proliferative activity after PBD occurred during the first 10 days and that the ductal cells were forced into proliferation to the same extent as the acinar ones. These findings are of interest for future studies of hormonal influences on the development of pancreatic carcinoma.

Proliferative Response of Different Exocrine Pancreatic Cell Types to Hormonal Stimuli

The trophic effect on the exocrine pancreas of the cholecystokinin analogue cerulein was studied in a long-term experiment (20 or 160 micrograms/kg/24 h for 14 days) in mice by measuring changes in pancreatic weight and protein, amylase, and DNA content. Further, the selective cell growth stimulation exerted by various doses of cerulein (4, 20, 54, 160 micrograms/kg/24 h) on different exocrine pancreatic cell types was studied by continuous administration of 3H-thymidine. In the first experiment animals given 20 micrograms/kg/24 h of cerulein had increased pancreatic weight and amylase and protein content, whereas the animals given the higher dose had unchanged weight and a less pronounced increase in amylase and protein content. The pancreatic DNA content was unaffected in the 20-micrograms group but was clearly decreased by the higher dose. In the second experiment a statistically significant increase over controls was found in the fraction of labeled ductal cells when 20, 54, and 160 micrograms of cerulein was administered. However, in the acinar cell population an increase was measured only in the 160-micrograms group. A tendency to nadir in cell labeling was observed in both acinar and ductal cell groups at less stimulation. Labeling of centroacinar cells increased in all cerulein-treated groups. The results show that all cell types of the exocrine pancreas can be forced into proliferation by the cholecystokinin analogue used and that there is preferential cell growth stimulation in the ductal and centroacinar cell populations.

Interstitial cells of the adult neocortical white matter are the remnant of the early generated subplate neuron population

The postnatal fate of the first-generated neurons of the cat cerebral cortex was examined. These neurons can be identified uniquely by 3H-thymidine exposure during the week preceding the neurogenesis of cortical layer 6. Previous studies in which 3H-thymidine birthdating at embryonic day 27 (E27) was combined with immunohistochemistry have shown that these neurons are present in large numbers during fetal and early postnatal life within the subplate (future white matter), that they are immunoreactive for the neuron-specific protein MAP2 and for the putative neurotransmitters GABA, NPY, SRIF, and CCK. Here, the same techniques were used to follow the postnatal location and disappearance of the early generated subplate neuron population. At birth (P0), subplate neurons showing immunoreactivity for GABA, NPY, SRIF, or CCK are present in large numbers and at high density within the white matter throughout the neocortex, and the entire population can be observed as a dense MAP2-immunoreactive band situated beneath cortical layer 6. Between P0 and P401 (adulthood), the MAP2-immunostained band disappears so that comparatively few MAP2-immunoreactive neurons remain within the white matter. There is a corresponding decrease in the number and density of neurons stained with antibodies against neurotransmitters. In each instance, these neurons could be double-labeled by the administration of 3H-thymidine at E27, indicating that they are the remnants of the early generated subplate neuron population. The major period of decrease occurs during the first 4 postnatal weeks, and adult values are attained by 5 months. Within the white matter of the lateral gyrus (visual cortex), the density of immunostained neurons decreases dramatically: MAP2, 82%, SRIF, 81%, and NPY, 96%. While SRIF-immunoreactive neurons compose a nearly constant percentage of MAP2-immunoreactive neurons in the white matter between P0 (22%) and P401 (23%), those immunoreactive for NPY decline from 18 to 4%. These changes occur during the same period in which there is less than a twofold increase in white matter area. These observations indicate that the interstitial neurons of the adult neocortical white matter are the oldest neurons of the cerebral cortex since most if not all are derived from the subplate neuron population. In addition, a quantitative analysis suggests that the postnatal decline in subplate neuron density cannot be accounted for solely through dilution by differential growth of the white matter and most likely reflects an absolute decrease in subplate neuron number.(ABSTRACT TRUNCATED AT 400 WORDS)

The effect of sofalcone on the kinetics of the generative cells and superficial epithelial cells in mouse gastric mucosa

Effects of an anti-ulcer drug, 2'-carboxymethoxy-4,4'-bis(3-methyl-2-butenyloxy) chalcone (sofalcone), on the generative cells and kinetics of superficial epithelial cells in mouse gastric mucosa, including the effect of hydrocortisone on them, were investigated by the use of 3H-thymidine autoradiography. The labeling indices and the width of the generative cell zone in the fundic mucosa were significantly decreased by administration of hydrocortisone. The decrease in the labeling indices and the width of the generative cell zone induced by administration of hydrocortisone were not inhibited by sofalcone. Hydrocortisone had no significant influence on the labeling indices of the generative cell zone in the pyloric mucosa, but decreased the width of it. The decrease in the width of the generative cell zone induced by hydrocortisone was inhibited by sofalcone. Concerning the influence on the kinetics of superficial epithelial cells, hydrocortisone inhibited the increase in the labeling indices of the superficial epithelial cells after continuous administration of 3H-thymidine in both the fundic mucosa and pyloric mucosa. In the fundic mucosa, sofalcone showed no influence on the inhibition by hydrocortisone, but in the pyloric mucosa, the effect of hydrocortisone was abolished by sofalcone. These results suggest that sofalcone inhibits the reduction of the cell production and the prolongation of the life span of superficial epithelial cells caused by hydrocortisone in the pyloric mucosa.

Development of intracerebral dopaminergic grafts: a combined immunohistochemical and autoradiographic study of its time course and environmental influences.

The aim of the study was to obtain a description of some aspects of the development of intracerebral dopaminergic grafts, namely, the time course of the glial reaction and its relation to cell division on one hand, and the development of graft-originated innervation and its dependence on adequate matching of the implanted neurons and target site on the other hand. Cell suspensions obtained from the mesencephalon or hypothalamus of embryonic day (ED) 14 rat embryos were implanted into the striatum or lateral hypothalamus of adult rats following the destruction of the nigrostriatal system of the hosts. Animals were sacrificed at different postimplantation times, and the development of the graft was followed by immunohistochemistry by using antisera directed against tyrosine hydroxylase (TH) or glial fibrillary acidic protein (GFA). Furthermore, the existence of cell division at various times following implantation was examined by performing autoradiography on immunostained sections after prior intraventricular administration of 3H-thymidine to the host. The first stage of the development of intracerebral grafts was characterized by the existence of intense cell division within the grafted tissue, lasting about 2 weeks, and also in the host tissue surrounding the graft, lasting only about 6 days. The cell division in the host tissue was paralleled by the existence of a strong glial reaction which, however, did not extend into the graft itself. Glial reaction in the host tissue gradually decreased at later times and disappeared by 4 weeks postimplantation without leaving behind a noticeable glial scar. The graft itself was, however, transiently filled with a population of reactive astroglial cells between 3 and 6 weeks postimplantation. Within grafts of mesencephalic tissue located in the striatum TH-positive neurons were distributed evenly at short times postimplantation (2-6 days). At later time a compartmentation could be observed, with TH-positive neurons being aligned along the graft-host interface or clustered within the graft itself. Innervation of the host tissue by TH-positive fibers increased between 1 and 6 weeks postimplantation. On the other hand, no compartmentation and reinnervation of surrounding host tissue was observed for intrahypothalamic grafts of mesencephalic tissue or intrastriatal grafts of hypothalamic tissue. This last observation indicates that adequate matching of implanted neurons and target tissue plays an important role in the development of intracerebral dopaminergic grafts.

Mechanistic considerations for carcinogenic risk estimation: chloroform.

Chloroform has been reported to induce cancer in rodents after chronic administration of high doses by gavage. However, the interpretation of these findings is hampered by a lack of knowledge concerning the relative roles of genetic and nongenetic mechanisms in these bioassays. The present studies were carried out in male B6C3F1 mice in order to investigate the potential of chloroform to induce genetic damage and/or organ toxicity at the sites where tumors have been observed in the various bioassays. These studies revealed that carcinogenic doses of chloroform produced severe necrosis at the sites where tumors later developed. This was demonstrated by light microscopy as well as by determination of the cellular regeneration index following administration of 3H-thymidine. Noncarcinogenic doses of chloroform failed to induce these responses. In contrast, studies of DNA alkylation and DNA repair in vivo failed to give any indication that chloroform had produced the type of genetic alterations associated with known genotoxic chemicals. These data suggest that the primary mechanism of chloroform-induced carcinogenesis is nongenetic in nature. If the same mechanism predominates in man, there should be little to no carcinogenic risk associated with exposure to noncytotoxic levels of chloroform.

Proliferative activity in the rat epididymis during postnatal development.

The proliferative activity of the rat epididymis during postnatal development was investigated with the use of autoradiography. Animals at 14, 21, 28, 35 and 56 days of age were sacrificed 2 hours following the administration of 3H-thymidine. Cell types that showed a significant labeling index were columnar cells in 14- and 21-day-old animals; principal and basal cells in 28, 35, and 56 day old rats. During this period of development, the pattern of cellular activity in the initial segment differed from the middle and terminal segments in having a peak of activity on day 28. The middle and terminal segments had similar proliferative patterns. In two additional experiments, 10- and 21-day-old rats were given 3H-thymidine and killed 1 week later on day 16 and 28. Labeled narrow cells were present in day 16 animals, whereas labeled narrow, principal, and basal cells were found in day 28 rats. It was concluded that columnar cells are the precursor to narrow, principal and basal cells.

Anaphase and telophase labeling patterns following administration of 3H-thymidine as an indicator of cell cycle time in vivo.

According to the Taylor, Woods and Hughes model, incorporation of3H-thymidine (3H-TdR) into DNA of cells in S phase implies that chromatids are labeled equally and symmetrically. In autoradiographs, nuclei of prospective daughter (sister) cells produced at the first division (M1) after administration of the radioactive precursor, confirm this model, being covered by grain numbers which are equal or nearly equal. During the second division (M2) following pulse application of3H-TdR, chromosomes with labeled and newly synthesized but unlabeled chromatids are separated by what appears to be a random process. Correspondingly, a higher degree of asymmetry in grain counts over daughter (sister) nuclei is observed during and after M2 than during and after M1. This reasoning has been applied to dog erythroblasts (cell cycle time in vivo of approximately 10 h). It was noticed that the switch from the M1-pattern of grain distribution over anaand telophase figures to the M2-pattern takes place at exactly the expected time for a second cycle. Statistical analysis using a double square root plot showed that this phenomenon is highly significant. The method presented in this report may be useful in estimating intermitotic + G2 times in humans, particularly with regard to neoplastic disorders, since it involves less frequent sampling than for establishing labeled-mitosis-curves. For optimal flattening of cells and autoradiographic geometry, it is proposed that smears be prepared with the help of a cytocentrifuge.

The cellular reaction in glomeruli of rats with anti-glomerular basement membrane nephritis.

This work examines the cellular reaction in glomeruli of rats with an acute form of anti-glomerular basement membrane (GBM) nephritis. After intravenous injection of rabbit-derived anti-GBM serum, heterologous and, later, autologous IgG fixed to the glomerular capillary loops. From day 1 to 11 of nephritis, glomerular hypercellularity was not appreciable or modest by qualitative histology, whereas morphometric evaluation revealed mild but significant increases of glomerular cell counts as compared to untreated controls. Histoautoradiographic studies of renal tissue 2 h after pulse administration of 3H-thymidine served to determine the proportion of glomerular cells which synthesized DNA, indicating forthcoming cell division. The mean labeling index (LI) of epithelial cells of Bowman's capsule was 1.8% in controls, rose 4- to 5-fold on the first day of nephritis and fell gradually from day 2 to 11. LI to tuft cells was 1.9% in controls, peaked on day 4, and fell thereafter. Histochemical staining for nonspecific esterase, a marker of monocytes and macrophages, revealed that glomeruli of normal rats contain very few stained cells (mean: 0.2/tuft section). In the course of nephritis, a gradual increase of esterase-positive cells took place in glomeruli which reached a plateau on day 4 (mean: 5.5/tuft section), indicating an influx of migrating monocytes. It is concluded that the injury of this model of anti-GBM nephritis involves markedly enhanced proliferation of intrinsic glomerular cells associated with infiltration of monocytes. Such a prominent cellular response is revealed by the applied quantitative methods in the absence of histologically impressive glomerular hypercellularity.

Phase dependent proliferative response of lymphoid and erythroid cells in the pregnant mouse

Spontaneous mitotic activity in different maternal lymphoid organs was evaluated in pregnant CBA mice by in vivo and/or in vitro administration of 3H-thymidine. Thymidine uptake in thymus, spleen and inguinal lymph nodes increased transiently at the time of implantation, a response perhaps induced by a preimplantation peak of luteinizing hormone (LH). At mid-pregnancy there was a second, more pronounced increase in splenic uptake of 3H-thymidine, which may be due partly to a hormonal response induced by chorionic gonadotrophin (CG). Both B and T cells were involved in the proliferative response of spleen cells, but the B/T activity ratio increased in late pregnancy was also accompanied by formation of splenic erythroblasts.

Cell kinetic analyses of human malignant brain tumors (gliomas)

Administration of 3H-thymidine, either with or without mitostatic agents, to 24 patients with various gliomas revealed 1) in viable areas a labeling index (LI) for glioblastomas equal to 5-15%, 2) an LI of 1% or less in well-differentiated gliomas, and 3) the LI of anaplastic astrocytomas was intermediate and reflected the extent of anaplasia. Growth fraction was 0.14-0.44 in malignant gliomas (average 0.31 +/- 0.10), and cell cycle time obtained by means of stathmokinetic analysis was 75.6 +/- 45.7 hr. With few exceptions, the patients whose tumors showed an LI of 5% or more died within 1 year after the onset of disease, and no patients whose tumors at biopsy had an LI of over 5% survived more than 6 months. In contrast, there was a far better prognosis for patients whose tumors demonstrated an LI of less than 1%.

Radioautographic investigation of gliogenesis in the corpus callosum of young rats. I. Sequential changes in oligodendrocytes.

The corpus callosum of young rats was examined to clarify the behavior of the three subtypes of oligodendrocytes (the large organelle-rich "light oligodendrocytes," the smaller and more densely stained cells referred to as "medium oligodendrocytes," and the even smaller and denser "dark oligodendrocytes"). It was hoped to find out whether cells of the three subtypes undergo division and how they are related to one another. 3H-thymidine was given intraperitoneally as single or three shortly spaced injections to a first group of 19- to 20-day old rats weighing about 40 g, and to a second group of 25-day old rats weighing about 80 g. The animals were sacrificed at various time intervals from 2 hours to 35 days after 3H-thymidine administration. Pieces of corpus callosum were taken near the superior lateral angle of the lateral ventricles; and semithin sections were radioautographed and stained with toluidine blue. Two hours after 3H-thymidine injection, label is virtually absent from light, medium and dark oligodendrocytes, from microglia, and probably from astrocytes, but is present in about 10% of the immature glial cells, which include the poorly differentiated glioblasts and the partially differentiated oligodendroblasts and astroblasts. Hence, the cells undergoing DNA synthesis and mitosis in the corpus callosum are these three types of immature cells. During the week that follow the administration of 3H-thymidine, label appears in oligodendrocytes and astrocytes, which presumably have arisen from the initially labeled immature cells. The oligodendrocytes acquire label in a sequential manner: the light cells show label first and their labeling index reaches a peak at the seven-day interval; the medium oligodendrocytes become labeled next with a labeling peak toward the 14- and 21-day intervals and, finally, the dark oligodendrocytes with a peak around the 28-day interval. Analysis by the method of Zilversmit et al. ('42-'43) provides precise details on the sequence: immature cells presumed to be oligodendroblasts give rise to light oligodendrocytes which, after four to seven days, transform into medium oligodendrocytes which, after another 11 to 18 days, transform into dark oligodendrocytes. The dark cells may persist indefinitely or turn over at a very slow rate. It is concluded that oligodendrocytes arise from the last division of oligodendroblasts and develop in three main periods: a light stage lasting less than a week, a medium stage lasting about two weeks, and a very long lasting dark stage.

3H-thymidine incorporation (DNA synthesis) and radiotoxicity in the ovary of the Japanese quail before and during follicle formation.

No evidence was found for ribosomal DNA amplification in the oocytes of the Japanese quail, before or during folliculogenesis. DNA synthesis in the somatic cells, involved in follicle formation, starts at the medullar side of the basement membrane. The localized sterilization of the quail ovary after administration of 3H-thymidine (3H-TdR) seems to be due to radiation-induced lesions in the follicle forming somatic cells, rather than to direct radiation damage of the oocyte.