Biomedical subjects

K A Mason

Publications and source records attributed to K A Mason.

At least 37 records · Page 2Linked to original sources

PubMed1997

Spatial and temporal patterns of expression of epidermal growth factor, transforming growth factor alpha and transforming growth factor beta 1-3 and their receptors in mouse jejunum after radiation treatment.

The goal of the present study was to assess changes in proliferation in the mouse jejunum after irradiation and the role of the growth factors EGF, TGF-alpha and TGF-beta 1-3 in the proliferative response. Our working hypothesis was that feedback signals from the villus to cells in the crypt regulate proliferation, and that the growth factors EGF and TGF-alpha with their common receptor EGF-R are involved in stimulation of proliferation, while the growth factors TGF-beta 1-3 with their receptors TGF-beta RI and TGF-beta RII are involved in inhibition of proliferation during this regulation. Immunohistochemical detection methods and automated image analysis were used for objective quantification of growth factor expression. The data indicate that, after 5 Gy irradiation, growth stimulation in the crypts takes place before major changes in the villi are observed. However, the combination of the reduction in the cell number, the number of cells expressing TGF-beta 1-3 and the reduction in the level of expression of TGF-beta 1-3 in the villi may cause the release of crypt cells from regulatory growth inhibition and initiate a proliferation-stimulating signal by an increase in the production of TGF-alpha and EGF. Regulation of proliferation after initiation of a proliferative response seems to be related more to the growth factors EGF, TGF-alpha and TGF-beta 3 in the crypts than to villus cellularity or growth factor expression, supporting the concept of stem cell autoregulation as a mechanism of cell regeneration in the intestinal crypt.

Animals↗

PubMed1997

The effect of proliferative status and clonogen content on the response of mouse jejunal crypts to split-dose irradiation.

The rate of cell production in hierarchical tissues is affected by the differentiation probability after each clonogen division and the frequency with which divisions take place. We have modified the latter by a high-fiber diet, which caused an increase in the BrdUrd labeling index (LI) in jejunal crypts of mice, and have tested for a change in tolerance using the in vivo colony assay. Clonogens were counted using the in vivo colony assay of crypt regeneration with Poisson correction. The LI was estimated by counting BrdUrd-labeled cells in longitudinal sections of complete crypts. Arrest in mitosis induced by injection of paclitaxel was used to test for a difference in the rate of crypt cell production in mice fed low- and high-fiber diets. Split-dose studies were used to test whether the change in proliferative status of the crypts was accompanied by changes in either the number of clonogens per crypt or their radiosensitivity, or an increased proliferative response to radiation-induced cell killing. We found an increase in the crypt LI induced by the high-fiber diet was 15-25% and was dependent on the time of day. The data on arrest in mitosis did not demonstrate a difference in cell production rates based on diet, possibly because of insufficient resolution of the assay. We conclude that the high-fiber diet had no effect on radiosensitivity, the number of clonogens per crypt (again, within the resolution of the assay) or the "repopulated dose," the dose represented by the shift in the dose-response curve for 2.5 days relative to that for 6 h. When the number of clonogens at the start of rapid proliferation was different (on account of different first doses), the repopulated dose was the same when 5 Gy X rays was given first but was higher for the animals on the high-fiber diet when 12 Gy was given first. The high-fiber diet caused an increase in the LI in the crypts that was not accompanied by any change in radiosensitivity or, within the resolution of the assay, numbers of clonogens per crypt. The increased LI also did not result in an increase in clonogen repopulation between split (and equal) doses. However, in split-dose experiments where the first dose was higher and as a consequence the number of clonogens at the start of the proliferative response was lower, there was evidence of a higher rate of clonogen production with the high-fiber diet than with the low-fiber diet.

Animals↗

PubMed1997

Docetaxel enhances tumor radioresponse in vivo.

Although the radiosensitizing potential of paclitaxel has been investigated extensively in cancer treatment, a sister taxane, docetaxel, has been studied rarely. We investigated the ability of docetaxel to enhance in vivo tumor radioresponse and influence radiation injury to normal tissue. In addition, mitotic arrest and apoptosis in tumors and normal tissues were assessed after docetaxel administration to determine whether these cellular effects underly its radio-modifying action. Mice bearing in their legs 8-mm isotransplants of a murine mammary carcinoma, designated MCA-4, were treated with 33 mg/kg docetaxel i.v., 9-21 Gy single-dose local tumor irradiation, or both (in which case radiation was given 9 or 48 h after docetaxel). Tumor growth delay was the end point of the treatments. Mitotic arrest and apoptosis were assayed 1-72 h after treatment with docetaxel. Normal tissue radioresponse was determined using jejunal crypt cell survival 3.5 days after mice were exposed to 9.2-14.8 Gy single-dose, total-body irradiation; the mice were treated with 33 mg/kg docetaxel i.v. 3, 9, or 48 h before irradiation. Docetaxel was assessed for its ability to induce mitotic arrest and apoptosis in jejunum 1-72 h after treatment. Docetaxel induced both mitotic arrest and apoptosis in both tumor and jejunum. Mitotic arrest preceded apoptosis and peaked in the tumor at 9-12 h after treatment; it peaked at 3 h in jejunum. Docetaxel enhanced tumor radioresponse by a factor of 1.45 when the drug was given 9 h before radiation and 2.33 when it was given 48 h before. In contrast, it only slightly enhanced radiation-induced damage of the jejunum and only when given 3 or 9 h before irradiation. Thus, docetaxel given within 2 days before irradiation acted as a potent enhancer of tumor radioresponse and increased the therapeutic gain of irradiation.

Animals↗

PubMed1996

Apoptosis and mitosis as prognostic factors in pathologically staged N1 nonsmall cell lung cancer.

PURPOSE: This study aimed to established whether spontaneous apoptosis or mitosis has prognostic value among patients with pathologically staged N1 nonsmall cell lung carcinoma (NSCLC) treated with surgical resection with or without adjuvant therapy. METHODS AND MATERIALS: Material from 173 patients who had resections between 1970 and 1988 was analyzed for apoptosis and mitosis. There were 128 men and 45 women, with a median age of 61 years. There were 86 squamous cell carcinomas (SQ), 73 adenocarcinomas (AC), 3 large-cell carcinomas (LC), 6 SQ-AC, and 5 unclassified. Patients were observed from 2 to 209 months (median 27). Actuarial methods were used to assess survival and freedom from distant metastasis. RESULTS: In NSCLC, apoptosis was found to range from 0.2% to 2.8% (median 1.0%) and mitosis from 0 to 1.8% (median 0.4%). Tumors having higher levels of apoptosis also had higher levels of mitosis (p = 0.001). The values of neither apoptosis nor mitosis depended on size, location, differentiation of tumors, age, performance status, or weight loss of patients. However, the values of apoptosis depended on tumor histology in that high values (greater than or equal to the median) were more frequent in SQ (49%) than in AC/LC (29%) (p = 0.01). The overall survival for NSCLC patients, which was 33% at 5 years, did not depend on the level of either apoptosis or mitosis. The 5-year survival of patients having SQ was higher (43%) than that of patients having AC/LC (21%) (p = 0.03). Patients with high apoptosis showed significantly better 5-year overall (p = 0.008) and DMF (p = 0.0012) survivals in the SQ group compared to the AC/LC group. High mitosis compared to low mitosis was a significantly better predictor for 5-year survival (62% vs. 29%, respectively) (p = 0.035) in the SQ. However, high mitosis was a significantly worse 5-year DMF survival predictor compared to low mitosis: 13% vs. 56%, respectively (p = 0.05) in AC/LC. In the multivariate models for AC/LC, mitosis remained a significant predictor of 5-year distant metastasis (p = 0.025) controlling for treatment groups (p = 0.042), whereas apoptosis was an independently significant predictor of 5-year distant metastasis (p = 0.010). CONCLUSION: Squamous cell histology predicted significantly better 5-year overall and DMF survivals compared to AC/LC. Apoptosis was correlated with mitosis. Although apoptosis or mitosis did not predict survival or DM, high apoptosis or mitosis predicted significantly better survival in SQ and significantly worse survival in AC/LC with regard to overall and DMF survivals. In the multivariate models for AC/LC, apoptosis alone or mitosis with variable treatment was a significant predictor of 5-year distant metastasis. Thus, pretreatment levels of apoptosis or mitosis might be useful for predicting treatment outcome of SQ and AC/LC subsets of NSCLC when analyzed separately and for predicting metastatic incidence of AC/LC.

Adenocarcinoma↗

PubMed1996

Relationship of mitotic arrest and apoptosis to antitumor effect of paclitaxel.

BACKGROUND: Microtubules are cellular organelles with functions that include control of cell division by mitosis, cell morphology, and transport of material within the cell. The anticancer drug paclitaxel (Taxol) promotes accelerated assembly of excessively stable microtubules. Consequently, treated cells tend to become arrested in mitosis. The drug also induces apoptotic cell death in vitro and in vivo. Prior to this study, the relative contributions of mitotic arrest and apoptosis to the in vivo antitumor effect and the relationship between the two factors had not been established; moreover, it is not known whether paclitaxel-induced mitotic arrest inevitably results in cell death. PURPOSE: Our aim was to quantify the mitotic arrest and apoptosis induced by paclitaxel in 16 murine tumors in vivo and to correlate these two factors with the drug's antitumor effect. METHODS: Inbred C3Hf/Kam mice were implanted with one of the following 16 syngeneic tumors: seven adenocarcinomas (MCa-4, MCa-29, MCa-35, MCa-K, OCa-I, ACa-SG, and HCa-I), two squamous cell carcinomas (SCC-IV and SCC-VII), six sarcomas (FSa, FSa-II, Sa-IIa, Sa-NH, NFSa, and Sa-4020), and one lymphoma (Ly-TH). The tumor growth delay induced by paclitaxel (40 mg/kg body weight given intravenously) was measured in 163 control and 163 treated mice, and its significance was assessed by Student's t test. In a separate group of 439 mice, the percentage of cells in mitosis or apoptosis was scored micromorphometrically at various times after paclitaxel administration. The significance of correlations between paclitaxel-induced tumor growth delay and paclitaxel-induced levels of mitosis or apoptosis was determined by simple correlation and Spearman's rank correlation. P values reported represent two-sided tests of statistical significance. RESULTS: Statistically significant tumor growth delays were found in response to paclitaxel treatment of mice for three of four murine mammary carcinomas (all P < or = .010), an ovarian carcinoma (P = .00003), a salivary gland adenocarcinoma (P = .0002), a lymphoma (P = .0002), and two of six sarcomas (both P < or = .034), but not for either of two squamous cell carcinomas or for the hepatocellular carcinoma. Paclitaxel-induced mitotic arrest was apparent in all tumor types, but to various degrees, and was not significantly correlated with growth delay (R2 = .16; P = .124). In contrast, apoptotic cell death in response to paclitaxel was not ubiquitous, but it was strongly correlated with growth delay (R2 = .59; P = .001). The pretreatment level of apoptosis was correlated with both paclitaxel-induced apoptosis (R2 = .71; P = .00004) and tumor growth delay (R2 = .55; P = .001). CONCLUSION: The antitumor effect of paclitaxel was correlated with paclitaxel-induced apoptosis and base-line apoptosis, but not with mitotic arrest. IMPLICATIONS: Apoptosis is an important mechanism of cell death in response to paclitaxel treatment of in vivo murine tumors. An underlying tumor type-specific propensity for apoptosis is implied by the correlation between pretreatment and paclitaxel-induced apoptosis. Both the extent of pretreatment apoptosis and the paclitaxel-induced percentage of apoptosis may be useful predictors of response to the drug.

Adenocarcinoma↗

PubMed1996

Accelerated repopulation during fractionated irradiation of a murine ovarian carcinoma: downregulation of apoptosis as a possible mechanism.

PURPOSE: To test whether accelerated tumor clonogen repopulation occurs during continuous fractionated radiotherapy of a slow-growing mouse ovarian tumor, and if so whether the accelerated rate of repopulation is predicted by the pretreatment potential doubling time, and whether changes in apoptotic response are a possible mechanism for this change. METHODS AND MATERIALS: The rate of clonogen production during fractionated radiotherapy was followed using the tumor-control assay, with an independent determination of the sensitivity to repeated dose fractions in vivo in the absence of repopulation. The pretreatment potential doubling time was measured by bromodeoxyuridine (BrdUrd) labeling and fluorescence measurements. The apoptotic and mitotic indices at various times during treatment were scored histologically. RESULTS: The slow-growing (pretreatment volume doubling time 6 days) ovarian tumor OCA responds to daily irradiation with 6 Gy under hypoxia by negligible tumor clonogen production in the first few days, followed by a change at about 9 days to accelerated repopulation, after which the effective clonogen doubling time Tclon was about 2 days, near the pretreatment Tpot of 1.7 days. Alternative interpretations of the data, such as a change in radiosensitivity vs. a change in the repopulation rate or acceleration at 3 days as opposed to 9 days, were shown to be unlikely. This change was accompanied by a reduced apoptotic response (measured morphometrically). CONCLUSIONS: When sensitivity to fractionated doses has been corrected for in vivo, this slow-growing mouse tumor exhibits a change to accelerated clonogen production during a continuous radiotherapy regimen that is accompanied or preceded by a reduced histologic apoptotic response. Tclon during accelerated repopulation was slightly longer than the pretreatment Tpot.

Animals↗

PubMed1996

Radiation-induced apoptosis in normal and pre-neoplastic mammary glands in vivo: significance of gland differentiation and p53 status.

The tumor suppressor gene p53 maintains the integrity of the genome by stimulating apoptosis in cells that have sustained DNA damage. The p53 gene is frequently altered in human cancers, including breast cancer, and such alterations are thought to result in genomic instability and aneuploidy, 2 hallmarks of anaplastic cells. We used radiation as a DNA-damaging agent to test the role of p53 in controlling apoptosis propensity in pre-neoplastic mammary lesions in mice. Four different pre-neoplastic mammary outgrowth lines, D1, TM2H, TM4 and TM12, were maintained by serial transplantation in the cleared mammary fat pads of syngeneic BALB/c mice. These lines have known alterations in p53 expression: TM12 has normal expression, D1 over-produces wild-type protein, TM2H contains a deletion resulting in a null phenotype and TM4 produces a mutant protein. Mice bearing the various outgrowths were irradiated with 5 Gy, and apoptosis was scored by examination of histological sections prepared from the outgrowths 6 hr after irradiation. The TM12 outgrowths, but not the TM2H outgrowths, exhibited radiation-induced apoptosis. The D1 and TM4 lines expressed radiation-induced apoptosis while the fat pads were being repopulated; however, the induced apoptosis declined to low levels as the mice aged. These results are consistent with the hypothesis that normal p53 function is important if mammary cells with DNA damage are to be deleted by apoptosis.

Animals↗

PubMed1996

Therapeutic potential of paclitaxel-radiation treatment of a murine ovarian carcinoma.

BACKGROUND: Paclitaxel has been shown to radiosensitize tumor cells in culture by arresting them in the most radiosensitive G2 and M cell cycle phases. In vivo preclinical studies are now necessary to obtain full insight into the radiopotentiating potential of this drug and its ability to increase the therapeutic gain of radiotherapy. We tested its ability to enhance the tumor radioresponse of an ovarian carcinoma and to influence the normal tissue radioresponse of recipient mice. METHODS: Mice bearing 8-mm isotransplants of a syngeneic ovarian carcinoma, designated OCA-I, in their legs were treated with 40 mg/kg paclitaxel i.v., 14-60 Gy single-dose local tumor irradiation, or both; radiation was given under ambient conditions 1-96 h after paclitaxel. Tumor growth delay, tumor cure rate (TCD50 assay), and delay in tumor recurrences were measured. Normal tissue radioresponse was determined using jejunal crypt cell survival at 3.5 days after exposure of mice to 9-14 Gy single dose of total body irradiation; the mice were untreated or treated with 40 mg/kg i.v. paclitaxel 4-96 h before irradiation. RESULTS: Paclitaxel alone was effective against OCA-I, but its combination with irradiation produced supra-additive tumor growth delay. It also reduced TCD50 values and delayed tumor recurrences. The enhancement of tumor radioresponse ranged from 1.33 to 1.96; the value increased as the time between paclitaxel administration and tumor irradiation increased up to 48 h, but then decreased again at 96 h. In contrast, paclitaxel protected jejunum against radiation damage by factors of 1.03 to 1.07 when given 24-96 h before irradiation. It showed some potentiation of damage (by a factor of 1.07), but only when given 4 h before irradiation. CONCLUSIONS: Paclitaxel potentiated tumor radioresponse if given within 4 days before irradiation, whereas it caused radioprotection of normal tissue (jejunum) at that time. Therefore, paclitaxel significantly increased therapeutic gain and so has potential for use in combination with radiotherapy for pelvic malignancies.

Animals↗

PubMed1996

Strain difference in jejunal crypt cell susceptibility to radiation-induced apoptosis.

Levels of radiation-induced jejunal crypt cell apoptosis were compared in C57BL/6J, C3Hf/Kam and C3H/HeJ mice. Apoptosis levels were consistently lower in the C3H strains than in C57BL/6J. Although other explanations are possible, the strain difference is most likely to have a genetic basis, and in fact a preliminary analysis of the F2 progeny of C3H/HeJ and C57BL/6J mice indicates that more than one gene is involved. Both C3H strains also had lower levels of radiation-induced thymocyte apoptosis than C57BL/6J mice. Jejunal crypt cell apoptosis levels did not co-segregate with thymocyte apoptosis levels in the F2 progeny of C57BL/6J and C3H/HeJ mice. These results imply that the genes responsible for the difference in radiation-induced thymocyte apoptosis levels between these two strains are not the same as those responsible for the strain difference in radiation-induced jejunal crypt cell apoptosis levels. The experiments reported here identify strain-specific differences in levels of radiation-induced crypt cell apoptosis and are a first step towards identifying genetic polymorphisms that influence sensitivity of the small intestine to damage from ionizing radiation.

Analysis of Variance↗

PubMed1996

Changes in clonogen number and radiation sensitivity in mouse jejunal crypts after treatment with dimethylsulfoxide and retinoic acid.

Retinoic acid (RA) and the drug carrier dimethylsulfoxide (DMSO) have been shown to reduce cellular radiation sensitivity in vitro because of their hydroxyl radical scavenging properties. Both agents have also been shown to induce differentiation in vitro and in vivo. As intestinal crypts are multicellular systems, crypt survival after irradiation depends not only on the cellular sensitivity of the clonogenic cells, but also on the number of clonogenic cells in each crypt, which may be changed by treatments with agents which induce differentiation. In the present experiments we examined the effects of DMSO and RA on the radiosensitivity of mouse jejunal crypts in vivo using the microcolony assay. Mice were treated with five daily intraperitoneal doses of 0-500 microgram RA in 0.1 ml DMSO per mouse, the last dose applied 4 h before the start of irradiation. The results showed a clear protection by 100 and 500 micrograms/day RA in 0.1 ml DMSO for crypt survival over the dose range of 9-16 Gy. The D0 was increased from 1.30 Gy for untreated controls to 1.59 Gy after treatment with DMSO alone, and to 1.85 Gy after treatment with 100 micrograms/day RA in DMSO. Split-dose experiments showed a reduction in the number of clonogens by a factor of about 2 from DMSO treatment alone, with no additional effect of RA on the number of clonogens. Despite this reduction, the number of BrdUrd-labeled cells per crypt remained roughly the same, as did the count of cells per longitudinal villus section. We conclude that, in addition to the protective effects of RA in DMSO, there is induced differentiation of crypt clonogens which is counteracted by increased proliferative activity of transit cells with the result that villus cellularity is maintained.

Animals↗

PubMed1996

What insights into vertebrate pigmentation has the axolotl model system provided?

Amphibians have been judiciously exploited by developmental biologists for many years for studying basic developmental mechanisms in vertebrates. In this review, the contributions that have been made by urodeles, in particular the axolotl (Ambystoma mexicanum), to the study of pigment cell biology are elaborated. Pigment cell differentiation is described, and the wild-type pigment phenotype is contrasted to pigment mutants such as albino, axanthic, melanoid, and white. Methods used for studying pigmentation, including recently developed molecular biological tools, are included to illustrate the significance of the axolotl as a model system for studying vertebrate pigmentation.

Ambystoma↗

PubMed1996

Genetic basis of strain variation in levels of radiation-induced apoptosis of thymocytes.

Levels of radiation-induced apoptosis of thymocytes were compared in C57BL/6J and C3Hf/Kam mice. For up to 8 h after irradiation, levels of apoptosis were higher in the C57BL/6J strain for all doses assayed. The heritability of the strain difference in the extent of apoptosis resulting from irradiation was investigated using a breeding study. Analysis of the F1 and F2 intercross progeny of these two strains provided strong evidence for a maternal effect but little evidence of sex linkage. The results indicate that in this strain combination the level of apoptosis of thymocytes after irradiation is a heritable trait that is likely to be controlled by few genes. These genes should be detectable in a mapping study.

Animals↗

PubMed1995

Sequence-dependent antitumor activity of paclitaxel (taxol) and cisplatin in vivo.

The established antitumor efficacy of paclitaxel and cisplatin as single agents and their distinctly different mechanisms of action have prompted laboratory and clinical research into their use in combination. Our in vivo study was performed to investigate the importance of sequence of administration and inter-agent interval. C3Hf/Kam mice bearing OCa-I tumors received paclitaxel and cisplatin. The antitumor efficacy of the combination, measured as re-growth delay and expressed as the enhancement factor (EF), was determined for inter-agent intervals of 1, 9, 24, 48 and 72 hr. Morphometric analysis was used to determine the contribution of induced apoptosis. Our findings showed an additive effect when cisplatin preceded paclitaxel by 1 and 24 hr, producing EF of 1.1 and 1.0, respectively, and a greater than additive effect for 9 and 48 hr, producing EF of 1.3 and 1.8, respectively. This sequence, however, was associated with significant morbidity and mortality. When paclitaxel preceded cisplatin the effect was greater than additive with the EF for 1, 9 and 24 hr, being 1.2, 1.5 and 1.5, respectively, and increasing to a maximum of 1.9 at 48 hr. Thus, for this combination, the therapeutic ratio was improved when paclitaxel preceded cisplatin and was greatest when a 48 hr interval was allowed between drugs. We were unable to attribute the efficacy of the drug combination to increased induction of apoptosis and suggest other possible mechanisms.

Animals↗

PubMed1995

Role of reoxygenation in induction of enhancement of tumor radioresponse by paclitaxel.

We reported previously (L. Milas et al., Cancer Res., 54: 3506-3510, 1994) that paclitaxel greatly enhances the response of a murine mammary carcinoma to subsequent irradiation and hypothesized that the enhanced radioresponse was mediated by tumor cell reoxygenation caused by treatment with paclitaxel. Because paclitaxel induced massive tumor cell destruction by apoptosis, it was reasoned that as apoptotic cells were removed from the tumor more hypoxic cells would have access to oxygen, be reoxygenated, and, thus, become more sensitive to radiation. The present study tested this hypothesis by assessing the effect of 60 or 40 mg/kg paclitaxel on radioresponse of an 8-mm MCA-4 tumor irradiated under air-breathing or hypoxic conditions 9, 24, 48, or 72 h after paclitaxel administration. If the hypothesis was correct, paclitaxel would enhance tumor radioresponse more under air breathing than under hypoxic conditions, and the enhancement would increase as the time between paclitaxel administration and tumor irradiation increased within a few days after paclitaxel treatment but only when radiation was given under air-breathing conditions. The effect of the treatments was determined by tumor growth delay and the radiation dose required to control 50% of the tumors (TCD50). Paclitaxel greatly enhanced tumor radioresponse under air-breathing (and not hypoxic) conditions, increasing tumor growth delay, and reducing TCD50. These effects increased as the time interval between paclitaxel administration and tumor irradiation increased within the observation period of 72 h after paclitaxel treatment. The enhancement factors for tumor growth delay ranged from 1.19 at 9 h to 1.86 at 48 h and for TCD50, from 1.16 at 9 h to 1.47 at 72 h after paclitaxel. Direct measurements of tumor pO2 showed a median value in untreated tumors of 6.2 mmHg, which increased to 10.5 mmHg at 24 h and to 31.2 mmHg at 48 h after paclitaxel administration. Overall, these results show that paclitaxel is a potent enhancer of tumor radioresponse and that its effect is mediated by reoxygenation of hypoxic tumor cells.

Animals↗

PubMed1995

Effect of paclitaxel (taxol) alone and in combination with radiation on the gastrointestinal mucosa.

PURPOSE: Paclitaxel is a potentially useful drug for augmenting the cytotoxic action of radiotherapy because it has independent cytotoxic activity against certain cancers and blocks cells in the radiosensitive mitotic phase of the cell cycle. However, all rapidly proliferating tissues, both normal and neoplastic, may be affected by this therapeutic strategy. The aim of this study was to define the in vivo response of rapidly dividing cells of the small bowel mucosa to paclitaxel given alone and in combination with radiation. METHODS AND MATERIALS: Mice were given single IV doses of 10 or 40 mg/kg paclitaxel or four doses of 10 mg/kg paclitaxel at 6, 12, or 24 h intervals. The kinetics of mitotic arrest and apoptosis in jejunal crypts of mice at 1-24 h after treatment were defined histologically. An in vivo stem cell microcolony assay was used to assess the radiosensitizing potential of paclitaxel when radiation was delivered at the peak of mitosis and at 24 h after drug treatment. RESULTS: Paclitaxel blocked jejunal crypt cells in mitosis and induced apoptosis in a dose-dependent manner. Fractionating the paclitaxel dose over 1-4 days did not result in any greater accumulation of mitotically blocked cells than did a single dose. Mitosis peaked 2-4 h after paclitaxel and returned to near normal by 24 h. Apoptosis lagged several hours behind mitosis and peaked about 6 h later than mitosis. Despite these kinetic perturbations, there was little or no enhancement of radiation effect when single doses were delivered 2-4 h after paclitaxel administration. The maximum sensitizer enhancement ratio of 1.07 observed after a single paclitaxel dose of 40 mg/kg is consistent with independent crypt cell killing. Conversely, when radiation was given 24 h after paclitaxel, a significant protective effect of the drug (SER 0.89-0.92), most probably due to a regenerative overshoot induced by paclitaxel, was observed. CONCLUSION: Stem cells of the jejunal mucosa determining radiation response were not radiosensitized by paclitaxel with the drug concentrations and dose delivery schedules used, although additive cytotoxicity was observed with the highest drug dose. A radioprotective effect was observed when radiation was given 24 h after paclitaxel administration.

Animals↗

PubMed1995

Kinetics of mitotic arrest and apoptosis in murine mammary and ovarian tumors treated with taxol.

The kinetics of taxol-induced mitotic arrest and apoptosis in murine mammary carcinoma MCA-4 and ovarian carcinoma OCA-I tumors were determined to establish a possible causative relationship between mitotic arrest and apoptosis and to see whether these cellular effects of taxol would correlate with the extent of its antitumor efficacy. Mice bearing 8-mm tumors in a hind leg were given taxol i.v. at a dose of 10-80 mg/kg. Both tumors responded to taxol by significant growth delay or transient regression; in general, the response was greater as the dose of taxol was increased. For kinetics studies the mice were treated with 60 mg/kg taxol given once when tumors were 8 mm in size or twice, with the second dose being given 3 days after the first. At various times ranging from 1 to 96 h after treatment with taxol, tumors were histologically analyzed to quantify mitotic and apoptotic activity. After a single dose of taxol, mitotic arrest was visible at 1 h, and the mitotic index increased with time to reach peak values of 36% in MCA-4 tumors and 22% in OCA-I tumors at 9 h. The index then declined to a baseline of 1%-3% at 3 days for MCA-4 tumors and 1 day for OCA-I tumors. Apoptosis followed mitotic arrest, beginning at the time of peak mitotic arrest, increasing to the highest level of about 20% at 18-24 h after treatment and gradually declining to the normal level of 3%-6% after 3-4 days. Nuclear material progressively condensed in mitotically arrested cells, culminating in the frank appearance of multiple apoptotic bodies. The change in cell morphology plus the dynamics of apoptosis development imply that a large percentage of tumor cells arrested in mitosis by taxol die by apoptosis. Kinetic analysis undertaken after the second dose of taxol showed a considerably lower percentage of cells arrested in mitosis as compared with that seen after a single dose, and the induction of apoptosis by the second dose was minimal. However, the antitumor efficacy of the second dose of taxol was similar to or better than that of the first dose, implying that in addition to mitotic arrest and apoptosis, there exist other mechanisms by which taxol exerts its antitumor action.

Animals↗

PubMed1995

The identification and partial cloning by PCR of the gene for tyrosinase-related protein-1 in the Mexican axolotl.

The tyrosinase gene family is currently composed of three members, tyrosinase and two tyrosinase-related proteins, TRP-1 and TRP-2. These three gene products have all been found to act in the synthesis of melanin pigments with the enzyme tyrosinase catalyzing the initial rate-limiting steps. Thus far these genes have primarily been analyzed in higher vertebrates. We have used degenerate PCR primers to isolate a large fragment of an axolotl tyrosinase-related protein. Sequence analysis of the entire 1,057-bp fragment isolated indicates a high degree of similarity to the mouse TRP-1, the product of the brown locus. Phylogenetic analysis supports the conclusion that the fragment isolated corresponds to the axolotl TRP-1 homolog. This is the first TRP-1 gene to be identified in an amphibian species.

Ambystoma mexicanum↗

PubMed1995

Tumor reoxygenation as a mechanism of taxol-induced enhancement of tumor radioresponse.

Paclitaxel is a novel chemotherapeutic agent that arrests cells in the radiosensitive G2 and M phases of the cell cycle and as such may act as a specific cell cycle radiosensitizer. We recently reported that paclitexel induces mitotic arrest in the MCA-4 murine mammary carcinoma and enhances radio-response of this tumor. However, the greatest enhancement was observed not when radiation was given at the time of peak mitotic arrest, which was 9 h after paclitaxel administration, but when it was given 24 h after paclitaxel. This implied the involvement of other mechanisms in radiosensitization; we hypothesized that tumor reoxygenation was a likely mechanism based on the observed massive loss of mitotically arrested cells at 24 h. The present study shows that paclitaxel greatly enhanced MCA-4 tumor radioresponse when radiation was given under air-breathing conditions (DMF = 1.74), but not when it was performed under hypoxic conditions. This observation supports the hypothesis of tumor reoxygenation as a mechanism of enhancement of tumor radioresponse. That reoxygenation occurred in tumors treated with paclitaxel 24 h earlier was confirmed by direct measurements of pO2 values, using the Eppendorf pO2 histograph. Median pO2 values increased from 6.2 mmHg in untreated tumors to 10.0 mmHg in tumors treated with paclitaxel. These observations emphasize the importance of timing of paclitaxel administration in relation to radiation treatment.

Animals↗