Irreversible loss of negative surface charge and loss of colony-forming ability in Burkitt lymphoma cells after x-irradiation

The electrokinetic behavior of Burkitt lymphoma P 3H-R1 cells were studied following X-irradiation. The surface charge of irradiated cells decreased progressively with time after X-ray treatment, as compared with a mean mobility value (−0.677 ± 0.045 μsec⁻¹ V⁻¹ cm) of non-irradiated cells. The frequency distribution of cells treated with 3 000 R 4 h previously showed quite a different peak of lower mobility than that of non-irradiated cells with the boundary of −0.550 μsec⁻¹ V⁻¹ cm. Percentages of cells which showed the higher mobility than the boundary value 4 h after irradiation with 30, 50, 65, 100, 200 or 3 000 R were in accord with the percentage of cells which showed colony-forming ability in soft agar, respectively. Following cell proliferation the population of electrokinetically intact cells increased with time after irradiation; these values also coincided with the population of colony-forming cells. Thus, the present results indicate that there is a close relation between loss of negative surface charge and loss of colony forming ability in irradiated cells, suggesting that loss of colony forming ability of the cells may be determined as early as 4 h after irradiation.     

Binucleate cell formation correlates to loss of colony-forming ability in X-irradiated cultured mammalian cells

The relationship between binucleate cell formation and the loss of colony-forming ability was examined in several cultured mammalian cell lines irradiated with X rays. The maximum fraction of binucleate cells after X irradiation increased dose-dependently within the range in which reproductive cell death might predominate over interphase cell death. When the logarithm of percentage survival was plotted against the percentage binucleate cells, a similar correlation was found for all cell lines tested, with the exception of mouse leukemia L5178Y cells, the most radiosensitive cells used. These observations suggest that the fraction of binucleate cells in the cell population can serve as a measure of cellular radiation damage.     

Relationship between nuclear and cytoplasmic RNA in Drosophilia cells.

Polyadenylated RNA was isolated from nuclei of cultured Drosophila cells, Schneider's line 2, and used as a template to synthesize a complementary DNA probe. Hybridization experiments were performed to study the relationship between nuclear and cytoplasmic RNA. About two-thirds of the nuclear polyadenylated RNA sequences exist in the cytoplasm. Experiments with fractionated cDNA probes demonstrated that RNA sequences that are frequent in the nucleus are also abundant in the cytoplasm. These findings are consistent with a precursor-product relationship in which some polyadenylated molecules in the nucleus are destined for the cytoplasm while other sequences are polyadenylated but not transferred.     

Effects of cholesterol- or 7-ketocholesterol-containing liposomes on colony-forming ability of cultured cells

Experiments with cultured Chinese hamster cells showed that incubation of the cells with (phosphatidylcholine + cholesterol + 7-ketocholesterol)-containing liposomes (4:3:1 by weight) during two hours led to a decrease in the colony-forming ability of cells down to zero, while (phosphatidylcholine + cholesterol)-containing liposomes (1:1 by weight) reduce this parameter by 90%. Furthermore, the cholesterol-containing liposomes (without 7-ketocholesterol) induce a decrease in the number of the maximal-site colonies accompanied by the corresponding increase in the number of the middle-size colonies.     

Relationship between fertilizing ability of frozen human spermatozoa and capacity for heparin binding and nuclear decondensation.

Nuclear decondensation of spermatozoa induced by heparin, reduced glutathione (GSH) or a mixture of heparin and GSH was studied using frozen-thawed human spermatozoa. The percentages of decondensed spermatozoa in controls and after treatment for 60 min with 30 mumol heparin l-1, 5 mmol GSH l-1, or heparin-GSH mixture were 1.5, 22.1, 4.3 and 37.6%, respectively. Most of the decondensed spermatozoa were eliminated by Percoll gradient centrifugation of samples previously treated with heparin or heparin-GSH mixture. However, comparable numbers of motile spermatozoa were recovered in the control and in each treated sample, demonstrating that a major proportion of motile spermatozoa was resistant to heparin (or heparin-GSH) effects on nuclear decondensation of spermatozoa. Fertilization of hamster oocytes was attempted using spermatozoa recovered in the 90% Percoll fraction and resistant to heparin-GSH decondensing mixture. Although insemination used a constant number of motile spermatozoa, fertilization rates were higher after treatments with heparin and GSH alone than in control or heparin-GSH-treated samples. In addition the number of spermatozoa that attached to the oocyte plasma membrane was a sixth or a half for sperm pretreated with heparin-GSH or heparin alone, respectively compared with untreated values. However, there was no evidence for induced acrosomal reaction by heparin and GSH, at least at the concentrations used. Qualitative analyses of heparin-binding sites were performed on untreated spermatozoa recovered in the 90% Percoll fraction by incubating spermatozoa in the presence of heparin covalently linked to albumin and coupled to colloidal gold (5 nm). Among this population of spermatozoa, 40.5% bound heparin-gold and labelling was mainly observed on the sperm head surface (88% of labelled spermatozoa) with (59.5%) or without (28.5%) tail labelling. Only a small proportion (23%) of spermatozoa that attached to the oocyte plasma membrane bound heparin-gold conjugate and only weak labelling was observed on the sperm head. Moreover, the proportion of spermatozoa that bound heparin-gold conjugate decreased (r = -0.77, P less than 0.0001) in relation to increasing concentrations of motile spermatozoa in the sample.(ABSTRACT TRUNCATED AT 400 WORDS)     

Relationship between nuclear remodeling and development in nuclear transplant rabbit embryos.

The present study characterized the profile of nuclear remodeling in nuclear transplant rabbit embryos and investigated the relationship between chromatin behavior after transfer and embryo development. The developmental potential and pattern of remodeling of donor nuclei from cleavage-, morula-, and blastocyst- (inner cell mass ICM, and trophectoderm, TE) stage donors were evaluated. In addition, we determined whether a modification in the synchrony between blastomere fusion and oocyte activation altered the profile of nuclear remodeling and affected development of reconstituted embryos. Development to blastocysts was similar with 8- and 32-cell-stage donor nuclei (42% and 33%, respectively, p greater than 0.1). However, it was reduced with ICM transplants (17%, p less than 0.05), and development of TE transplants did not progress beyond the 8-cell stage. Upon blastomere fusion into nonactivated oocyte cytoplasm, nuclear remodeling was characterized by premature chromosome condensation (PCC), followed by pronuclear (PN) formation and swelling. PCC occurred synchronously within 1.2-1.5 h post-fusion with all stages of donor nuclei (p greater than 0.1). PN formation in 8- and 32-cell transplants occurred approximately 4 h after fusion, and was synchronous to that of female pronuclei in activated oocytes; however, it was delayed in ICM and TE transplants (p less than 0.01). With all stages of donor nuclei, final nuclear diameter was similar to, or larger than, that of female pronuclei. Fusion to activated oocyte cytoplasm, as opposed to nonactivated cytoplasm, prevented PCC and extensive nuclear swelling (16.0 +/- 0.7 vs. 30 +/- 0.7 microns, respectively, p less than 0.01). Nuclear diameter in early embryos was smaller (p less than 0.01), and development to blastocysts was reduced (p less than 0.05). The results indicate that remodeling of the donor nucleus is not essential for development to blastocysts; however, it is beneficial. Furthermore, complete reprogramming seems possible only after remodeling of the donor nucleus, i.e., PCC in nonactivated cytoplasm, followed by nuclear swelling upon activation of the oocyte.     

Relationship between membrane potential and external potassium in human glioblastoma cells in tissue culture.

Cells from a human glioblastoma (TC 526) maintained in tissue culture for ten years had a mean membrane potential of 27 +/- 0.9 mV at an external potassium concentration [Ko] of 5.3 mM. When [Ko] was varied between 2.5 and 5.3 mM, membrane potential changes were close to those predicted by the Nernst equation. At higher [Ko], the Nernstian slope was approached only in the presence of 10(-5) M ouabain, which did not affect membrane potential at a [Ko] of 5.3 mM. An electrogenic sodium pump activated by high [Ko] could explain these findings; such a mechanism has been demonstrated in other tissues.     

Effect of UV-irradiated donor blood on the colony-forming ability of human bone marrow cells in culture

A possible mechanism of the hemopoiesis stimulation by UV and solar radiation was studied. The UV irradiation of leucocyte-enriched donor serum results in increasing the colony formation activity of the serum. The addition of such a serum but depleted of leucocytes to human bone marrow cells in semisolid agar culture stimulates its colony formation by 1.5-3 times. A serum depleted of leucocytes before the irradiation has not this property. It is supposed that cell surface glycoproteins, desorbing into the serum under UV irradiation, induce the colony formation activity of UV-irradiated blood.     

Increase of the negative charge of erythrocyte membrane proteins in hereditary neuromuscular diseases

The parameters of erythrocyte ghost protein's fluorophores by nitrate's anions were studied in patients with various hereditary myodystrophy. In all the groups under examination the share of fluorophores accessible to a quencher was close to 1. In erythrocyte membranes of healthy donors the relevant constant quenchering was about 17.3 +/- 1.9 M-1 while those of patients were decreased by 3.1 (Duchenne's myodystrophy) and by about 2.0 (other forms of primary and secondary progressive muscular dystrophies). The most probable reason for the decreasing constant of quenchering is the increase of negative charges on the erythrocyte membrane proteins.     

The relationship between colony-forming ability, chromosome aberrations and incidence of micronuclei in V79 Chinese hamster cells exposed to microwave radiation

Cultured V79 Chinese hamster fibroblast cells were exposed to continuous radiation, frequency 7.7 GHz, power density 0.5 mW/cm2 for 15, 30 and 60 min. The effect of microwave radiation on cell survival and on the incidence and frequency of micronuclei and structural chromosome aberrations was investigated. The decrease in the number of irradiated V79 cell colonies was related to the power density applied and to the time of exposure. In comparison with the control samples there was a significantly higher frequency of specific chromosome aberrations such as dicentric and ring chromosomes in irradiated cells. The presence of micronuclei in irradiated cells confirmed the changes that had occurred in chromosome structure. These results suggest that microwave radiation can induce damage in the structure of chromosomal DNA.     

Effect of polyinosinic-polycytidylic acid on the colony-forming ability of hematopoietic stem cells in conditions of allogeneic inhibition]

It was found that the colony-forming capacity of parental bone marrow transplant (C57BL/6) was partially restored in the (CBA X C57BL/6) hybrid recipient irradiated with 800 rad when poly I -- poly C preparation was injected. The effect of poly I -- poly C injection on the colony formation was equivalent to addition of the thymus cells syngeneic with the marrow. In either case the number of splenic colonies was more than double that in the control. On the other hand, it was found that in a completely syngeneic system the number of splenic colonies was not influenced by the thymus cells and poly I -- poly C preparation. Poly I -- poly C doses ranging from 50 to 100 mug and thymus cell doses ranging from 4-10(6) to 8-10(6) did not increase the efficiency of the colony formation with a stable bone marrow dose transplant.     

Nano-chemotherapy using cationic liposome that strategically targets the cell membrane potential of pancreatic cancer cells with negative charge

Negatively charged phosphatidylserine (PS) and sialic acid-containing glycosphingolipids (GM1) were observed to be over represented on the cell membranes of pancreatic cancer cells (BxPC-3) as opposed to normal pancreatic cells. Cationic liposomes (CL) were also found to selectively accumulate into the negatively charged cell membranes of BxPC-3 cells and inhibited their growth but have no effect on the viability of normal pancreatic cells. CL induced apoptosis in BxPC-3 cells via activation of caspase-3, -8, and -9 and mitochondrial events and inhibited tumor enlargement in xenograft mouse models of pancreatic cancer.     

Relationship between membrane lipid mobility and spectrin distribution in lymphocytes.

We have previously established that T and B lymphocytes in situ are remarkably heterogeneous with respect to the cytoskeletal protein spectrin. Since in erythrocytes spectrin is known to play an important role in the regulation of membrane fluidity, lipid organization and lateral mobility of membrane proteins, we have sought to determine if the heterogeneous patterns of spectrin distribution that we have observed are related to possible differences in membrane lipid organization in these various subsets. To this end, we have utilized a fluorescent pyrene-labelled phospholipid as a probe of the lipid lateral mobility and have examined two related T cell systems maintained in vitro, DO.11.10 cells and a spontaneously arising variant, DO.11.10V. In these (and other cloned in vitro systems) we have previously observed that the cells homogeneously express one of the kinds of spectrin distribution patterns observed in situ. Thus the uniformity of staining of these systems permits us to address whether the various patterns of spectrin distribution may be predictive of differences in membrane lipid properties. Here we show that in cells in which there is little or nor spectrin at the plasma membrane (DO.11.10) that the lipids in the plasma membrane are considerably less mobile than in its related variant in which spectrin is diffusely distributed within the cell and at the plasma membrane. From this and previous results, we conclude that differences in the distribution of the cytoskeletal protein spectrin among lymphocytes may be a useful parameter in helping to predict the status of membrane lipid organization.     

Relationship between membrane potential and external potassium in human glioblastoma cells in tissue culture

Cells from a human glioblastoma (TC 526) maintained in tissue culture for ten years had a mean membrane potential of 27 ± 0.9 mV at an external potassium concentration [K₀] of 5.3 mM. When [K₀] was varied between 2.5 and 5.3 mM, membrane potential changes were close to those predicted by the Nernst equation. At higher [K₀], the Nernstian slope was approached only in the presence of 10^?5 M ouabain, which did not affect membrane potential at a [K₀] of 5.3 mM. An electrogenic sodium pump activated by high [K₀] could explain these findings; such a mechanism has been demonstrated in other tissues.