Cellular response to irinotecan in colon cancer cell lines showing differential response to 5-fluorouracil.

BACKGROUND: Use of irinotecan (CPT-11) as second-line therapy for metastatic colorectal cancer has shown some promise in cases where 5-fluorouracil (5-FU) has failed. Cross-resistance to both drugs may however be a potential clinical problem. The cellular response to CPT-11 was investigated in two human colon cancer cell lines that demonstrate a differential response to 5-FU. MATERIALS AND METHODS: Cell cycle progression, clonogenic survival, DNA damage checkpoint activation, apoptosis induction and senescence development were assessed during 48 hours of treatment and 72 hours of recovery. RESULTS: Both cell lines had similar cellular response patterns to CPT-11. Growth inhibition, loss of clonogenicity, ataxia telangiectasia mutated (ATM) activation, H2AX phosphorylation, TP53 stabilization, CDKN1A induction, G2/M arrests, endoreduplication, negligible cell death and appearance of a senescence-associated beta-galactosidase phenotype were observed. CONCLUSION: Cross-resistance to 5-FU and CPT-11 was not demonstrated. The appearance of a senescence phenotype in response to CPT-11 treatment may have potential clinical relevance for treatment regimens.

Regeneration of epithelial defects in corneas previously treated with excimer laser. A study of cell kinetics in the rat corneal epithelium.

PURPOSE: The present study was to investigate, using cell kinetic methods, whether previous excimer laser treatment affected healing of later corneal epithelial erosions. METHODS: The right eyes of rats underwent central excimer corneal stromal-epithelial ablations (ArF 193 nm, 228 pulses, diameter 3,5 mm, 17,3 mJ per pulse, depth about one third of corneal thickness) and were allowed to heal for 15 weeks. Then central circular epithelial abrasions (diameter 3 mm), using N-Heptanol and mechanical debridement, were made on both corneas. The rats were killed 1,2,4 and 6 days after the last treatment. The central stromal thickness, the epithelial cell density, the labelling index (LI) and the mitotic rate (MR) of the peripheral, the midperipheral and the central areas of the corneal epithelium were calculated for each timepoint. RESULTS: The central stromal thickness increased equally in the two groups the first day after making the erosion, normalising in both groups during the following days. The corneal epithelium was restored at the same rate in both groups. The cell number per microscopic visual field, the LI and the MR were very similar for the two groups at all timepoints. CONCLUSION: Previous excimer laser treatment does not seem to interfere with healing of later epithelial erosions, when studied with cell kinetic methods.

Epithelial wound healing of the rat cornea after excimer laser ablation.

Rats with excimer corneal ablations (ArF 193 nm, 228 pulses, diameter 3,5 mm, 17.3 mJ per pulse, depth about 1/2 of corneal thickness) in one eye were killed 1, 2, 4, 6 and 13 days after treatment. The other eye served as control. The cell number per microscopic vision field, the labelling index (LI) and the mitotic rate (MR) were calculated for the peripheral, midperipheral and central areas of the corneal epithelium. The cell number showed a uniform depression in the remaining corneal epithelium at Day 1, normalizing from the centre to the periphery. The LI was only significantly increased at Day 1, while the MR was statistically significantly increased peripherally at Day 2 and in all areas at Day 6. However, when the corneal epithelium was evaluated as a whole, the MR was significantly increased at days 1, 2 and 6. The proliferative response of the epithelium was very homogenous irregardless of the distance to the original lesion. Both the migratory and the proliferative phases of the healing process seemed to be delayed when compared to the healing of pure epithelial wounds. However, the initiation of an increase in DNA synthesis seems not to be delayed, indicating that it is primarily the G2 phase that has been prolonged with this ablation procedure. The stromal thickness was increased from about one half of the normal values immediately after the treatment to above normal values at Day 4, thereafter decreasing to normal values at Day 13. Thus, the regenerative ability of the stroma is more pronounced in rats than in humans, but also in humans regeneration of the stroma can possibly explain the regression of the myopic shift seen some time after excimer laser treatment.

Cell kinetics of conjunctival and corneal epithelium during regeneration of different-sized corneal epithelial defects.

Rats with small (diam. 1.7 mm), medium sized (diam. 3.5 mm) or large (diam. 5.5 mm) corneal epithelial erosions in one eye were killed 1, 2 or 4 days after the injury. The proliferative response was evaluated by measuring the labelling index and the mitotic rate in the corneal epithelium and in the adjacent conjunctiva. The small erosions triggered a proliferative response in the cornea only with the maximum response occurring midperipherally. The medium sized erosions induced a higher and more extensive response in the cornea and also a slight increase of the labelling index in the limbal area. The large erosions induced an even more pronounced response in the peripheral cornea and an increase both of the labelling index and the mitotic rate well beyond the limbal part of the conjunctiva. It is concluded that the magnitude and the extent both of the conjunctival and the corneal regenerative response to a corneal abrasion is correlated to the size of the corneal defect. Temporary reduction in the conjunctival epithelial cell number shows that both cells in the limbal and the extralimbal conjunctiva migrate centripetally during healing of large corneal wounds. It is suggested that the stem cell theory should be modified. The limbal area is probably an area in which conjunctival epithelial cells or conjunctiva-derivated cells transform or differentiate to corneal epithelial cells.

Early cell kinetic effects of a single dose of narrow-banded ultraviolet B irradiation on the rat corneal epithelium.

The right eyes of 40 rats were exposed to a single erythemogenic dose of ultraviolet B irradiation (UVB) at 297 nm. The irradiation was directed perpendicular to the center of the cornea. The left eyes served as controls. The animals were randomly assigned into 10 groups. The labeling index (LI) after pulse labeling with tritiated thymidine and the mitotic rate (MR) after Colcemid administration were registered in the corneal epithelium at predetermined intervals up to 96 h after the irradiation. A mathematical method was used to correlate corresponding corneal areas from the different animals. In the central cornea the LI was considerably reduced up to 36 h after the irradiation. The LI increased toward the peripheral cornea and reached normal values at the limbal area. The MR was also reduced up to 36 h. However, this reduction was over the entire epithelium. The block in cell proliferation was followed by increased proliferation.

The early cell kinetic response during healing of corneal epithelial wounds.

The labeling index and the mitotic rate were measured at 4-hour intervals during the first 24 hours after a central abrasion had been made in the corneal epithelium of six groups of four rats each. The proliferative response was noted in the conjunctival, the limbal, and the corneal epithelium. After 24 hours, the density of epithelial cells was equal throughout the corneal epithelium, but there was only half the normal number of cells. Physiological mechanisms seem strongly to regulate the total number of cells per square unit throughout the healing corneal epithelium, but the nature of these mechanisms is unknown.