Nuclear scaffold organization in the X-ray sensitive Chinese hamster mutant cell line, xrs-5.

Nuclear organization was probed in the radiation-sensitive Chinese hamster ovary CHO) cell line, xrs-5, and compared with parental CHO K1 cells using the resinless section technique and DNase I digestions. The resinless section data showed no gross morphological differences in core filaments from the nuclear scaffolds of unirradiated CHO K1 and xrs-5 cells. However, the nuclear scaffolds of irradiated xrs-5 cells (1 Gy) had significantly increased ground substance. Irradiated and unirradiated CHO K1 cell nuclear scaffolds were morphologically identical. These data suggest that both CHO K1 and xrs-5 cell nuclear scaffolds had internal nuclear scaffolding networks that could provide DNA attachment sites. The rate of DNase I digestion of unirradiated CHO K1 and xrs-5 was not significantly different, but the extent of digestion was greater in unirradiated CHO K1 cells that in xrs-5 cells suggesting that less xrs-5 cell chromatin at DNA attachment points is accessible to the enzyme DNase I. The extents of digestion in irradiated (1 Gy) CHO K1 and xrs-5 cell nuclei also differed but the relationship was reversed. The irradiated xrs-5 cell samples were digested to a greater extent compared with CHO K1 cells. These chromatin digestion data suggest that the matrix attachment regions in xrs-5 cells are different from CHO K1 cells. The different DNA attachment organization in the xrs-5 cells may play a role in modulating radiation sensitivity.

Changes in the nuclear structure in the radiation-sensitive CHO mutant cell, xrs-5.

The structural organization of the cell nucleus was investigated by transmission electron microscopy in the radiosensitive Chinese hamster ovary (CHO) cell mutant, xrs-5 (D0 = 45 cGy), relative to parental K1 cells (D0 = 200 cGy). In 99% of all xrs-5 cells, the outer layer of the nuclear envelope was separated from the inner layer, while 96% of K1 cells had closely apposed layers. This separation of the inner and outer layers of the nuclear envelope in xrs-5 cells was not explained by an increased susceptibility of xrs-5 cells to osmotically induced changes because (1) xrs-5 cells retained the altered nuclear periphery even when several different fixation protocols were used and (2) xrs-5 cells were not more susceptible to cell lysis as measured by trypan blue dye exclusion or by the extracellular presence of lactate dehydrogenase. The difference in the morphological organization in the nuclear periphery of xrs-5 cells correlated with the radiation sensitivity of the cells; xrs-5 cells which spontaneously reverted to a radiation sensitivity similar to that of K1 cells also reverted to a nuclear morphology similar to that of K1 cells. The inner and outer layers of the nuclear envelope were retained in nuclear scaffolds isolated from K1 and xrs-5 cells, indicating that components of the nuclear periphery are part of the nuclear scaffold. These data show that xrs-5 cells have an altered nuclear periphery which correlates with the radiation sensitivity of the cells. The separation of the layers of the nuclear envelope may represent an altered template for repair of DNA damage at the nuclear scaffold and thus may play a role in the defective repair of X-ray-induced DNA double-strand breaks in xrs-5 cells.

Thermal sensitivity of CHO cells with different shapes.

Confirming previous reports, we observed that rounded Chinese hamster ovary (CHO) cells from suspension culture were more resistant to heat kill than flattened CHO cells from monolayer culture. Cell survival was quantitatively described by determining full cell survival curves for flattened versus rounded CHO cells after exposure to 43, 44, 45 and 46 degrees C and calculating the To values. The cell survival responses of the rounded cells were significantly different from those of the flattened cells. Based on the ratio of the To values, the rounded cells were consistently twice as resistant to heat-induced cell death than flattened cells. This difference in cell survival was not explained by a trypsin effect in monolayer cells, or by the amount of time rounded cells were maintained in suspension culture, or technical differences in heat-up and cool-down kinetics, or differences in extracellular pH. The two groups also showed no differences in cell generation times or whole cell protein contents. However, we found that the rounded cells had a lower percentage of cells in the S phase of the cell cycle (40% versus 52%) than the flattened cells, which could account for part of the differential heat-induced cell kill in the flattened versus rounded cells. These data suggest there is a significant association between cell shape and heat-induced cell killing.

Chloramphenicol: an impatient study of use and abuse.

During a six-month period, chloramphenicol was administered systemically to 100 patients and topically to 24 patients who were admitted to the Medical Center Hospital of Vermont, Burlington, Vermont. Seventy-nine percent of the patients receiving the drug systemically were on surgical services, 13% were on the pediatric service, and 8% were on the medical service. In this group, chloramphenicol was administered appropriately in 60.4% of cases as follows: 57.5% on the surgical, 50% on the medical, and 84.6% on the pediatric services. In 19 of 35 instances of inappropriate use on the surgical services, a potentially less toxic antibiotic could have been chosen. Eight cases were treated inappropriately because of dosage errors, and in seven cases no antibiotic should have been used. On the medical services all inappropriate use was due to underdosage, and on the pediatric service there was one case of incorrect dosage and one instance in which a less toxic antibiotic could have been chosen. Ophthalmological use of the drug accounted for all topical use. Of 24 patients treated, 21 received the drug for prophylaxis and three for therapy.

Side effects of minocycline: different dosage regimens.

The incidence of side effects due to two dosage regimens of minocycline was examined over a 5-day period. A total of 60 normal women volunteers were randomly assigned in a double-blind manner to either a group who took 100 mg of minocycline twice a day or a group who took 75 mg of minocycline twice a day for 5 days. Both groups were comparable from the standpoints of age, size, race, and the use of oral contraception, nicotine, and ethanol. They were seen on a daily basis, and symptoms were evaluated by both volunteers (from diaries) and physicians. Minocycline serum concentrations were determined on blood samples taken 2 h after the a.m. dose. Volunteers taking 150 mg of minocycline per day had significantly lower serum antibiotic concentrations than those taking 200 mg per day. However, both low- and high-dose groups exhibited similar incidence and prevalence of recorded symptoms, with the single exception of nausea, where the low-dose group had fewer symptoms than the high-dose group (P = 0.035). Symptomatic volunteers did not have higher serum concentrations of minocycline than their asymptomatic counterparts. When either weight or surface area was examined with antibiotic serum concentration there was a significant inverse correlation between the two on day 2 for both groups and also on day 4 for the low-dose group. It is concluded that, in women, a dose of 150 mg of minocycline per day is associated with the same degree of side effects as a dose of 200 mg per day.