Fraction Of Cells In S-phase Research Articles

Interleukin-4 enhances proliferation of human pancreatic cancer cells: evidence for autocrine and paracrine actions

Interleukin-4 (IL-4) is an immunomodulatory cytokine, which can inhibit the growth of tumour cells. Pancreatic cancer cells and tissues express high levels of IL-4 receptors. The aim of this study was to characterise the effects of IL-4 on the growth and signalling pathways of pancreatic cancer cells. Cell growth was determined by cell counting and MTT assays in association with fluorescence-activated cell sorter analysis, IL-4 expression using ELISA and real-time PCR techniques, and signal transduction using immunoprecipitation or immunoblot analysis. We now report for the first time that IL-4 significantly enhanced the growth of five out of six cultured pancreatic cancer cell lines in a dose-dependent manner in association with an increased fraction of cells in S-phase. Surprisingly, all six cell lines expressed endogenous IL-4, and IL-4 was detectable in the supernatant. Incubating cells with neutralising IL-4 antibodies resulted in a significant inhibition of basal growth in three cell lines, including IL-4-unresponsive MIA PaCa-2 cells, which however expressed the highest endogenous IL-4 levels. Interleukin-4 enhanced activity of MAPK, Akt-1, and Stat3 in IL-4-responsive, but not in IL-4-unresponsive MIA PaCa-2 cells; however, IL-4 enhanced tyrosine phosphorylation of insulin receptor substrate-1 and -2 in all cell lines. Our results demonstrate for the first time that pancreatic cancer cells produce IL-4 and that IL-4 can act as a growth factor in pancreatic cancer cells. Together with the observation that neutralising IL-4 antibodies can inhibit the growth of these cells, our results suggest that IL-4 may act as an autocrine growth factor in pancreatic cancer cells and also give rise to the possibility that cancer-derived IL-4 may suppress cancer-directed immunosurveillance in vivo in addition to its growth-promoting effects, thereby facilitating pancreatic tumour growth and metastasis.

Hedgehog signaling and cell cycle control in differentiating erythroid progenitors

Hedgehog (Hh) signaling regulates differentiation in numerous systems, but its functions in the control of hematopoietic differentiation have not been extensively explored. Initial studies have indicated that hedgehog signaling affects the proliferation and differentiation of erythroid progenitors (Detmer, K., et al., Erythroid differentiation in vitro is blocked by cyclopamine, an inhibitor of hedgehog signaling. Blood Cells Mol. Dis. 26(4) (2000) 360-372). To examine the effect of Hh signaling on the erythroid developmental program at the molecular level, Hh signaling in committed erythroid progenitors differentiating in vitro was inhibited, and the appearance/disappearance of molecular markers of erythroid differentiation was monitored. The expression timetable for CD34, CD36, the erythropoietin receptor, and glycophorin A was retarded in the absence of Hh signaling. Hemoglobinization was delayed and decreased relative to controls. Morphological changes of erythroid maturation were also delayed. The fraction of cells in S-phase was decreased during the initial period of exponential expansion as assessed by propidium iodide staining and flow cytometry, as was the rate of tritiated thymidine incorporation. A modest decrease in the proliferation rate was observed. These results suggest that Hh signaling is one of the mechanisms in the regulation of erythroid proliferation and differentiation.

Cell apoptosis and regeneration of hepatocellular carcinoma after transarterial chemoembolization.

To evaluate whether cell apoptosis and regeneration were existed in normal liver cells adjacent to carcinoma after transarterial chemoembolization (TACE). Fifty rabbits with hepatic carcinoma were divided into 5 groups at random: group A (control group), groups B and C (TACE treatment groups), groups D and E (partial hepatectomy groups). There were 10 rabbits in each group. Rabbits in groups B-E were treated by transarterial chemoembolization (TACE) and partial hepatectomy (PH) respectively. The changes of S-phase cell fraction (SPF), proliferation index (PI) and cell apoptosis in the normal liver tissue were determined with flow cytometry (FCM) after operations on the first and third days. We determined the mitosis index (MI) with histo-pathological method and the apoptosis index (AI) with TUNEL method at the same time. Twenty-four hours after operations, compared with control group, the rabbits in TACE group had much higher index of SPF, PI and MI (MI: t=4.89, P<0.001; SPF:t=5.27, P<0.001; PI: t=4.87, P<0.001). Moreover, the proliferation of liver cells in TACE group was much weaker than that of the cells treated by partial hepatectomy, and the differences were significant (MI: t=7.02, P<0.001; SPF: t=4.06, P<0.001; PI: t=2.70, P<0.05). Seventy-two h after operations, FCM showed a small sub-G1 peak in TACE group and PH group, compared with the control group, but there was no difference between them (t=0.41, P>0.05). TACE showed that AI in the treated rabbits was higher than that in control group (t=3.07, P<0.05), and there were no differences between TACE group and PH group, either (t=0.93, P>0.05). Cell apoptosis and regeneration exist in rabbit liver tissues after TACE in some degree, which may be associated with the selective embolization of iodised oil, chemotherapeutic drug and free radical damage.

Relation between the rate of tumour cell proliferation and latency time in radiation associated breast cancer.

BackgroundPatients with possible radiation induced cancer could be used to study if the rate of tumour cell proliferation is related to latency time. Such a finding could help researcher to find time periods when other initiating risk factors operate.MethodsSeventeen women with breast cancer, with a prior history of radiation treatment towards the parts or the whole breast, exclusive of the primary treatment of a breast cancer were identified. Most women had received treatment for benign disorders as hemangiomas, shoulder pain or skin infections. Three patients had been treated with mantle radiation for Hodgkin's disease prior to developing breast cancer. DNA analysis were performed, on remaining tumour tissue after hormone receptor analysis had been done, measuring the fraction of tumour cells in S-phase. Latency time (time between diagnosis and previous radiation treatment) was calculated and related to the S-phase fraction.ResultsA significant inverse relationship between latency time and S-phase was found (p < 0.0025), indicating that tumours with a high S-phase had a short latency time and vice versa. Among the possible radiation induced tumours, median S-phase was 14%, comparable with a median latency time of 22 years. Very high S-phase values were associated with short latency times (eg a S-phase of 35% would be compatible with a latency time of 7 years).ConclusionOur preliminary results indicate that S-phase is related to latency time and that the median latency time maybe as long as 22 years. Our data may also explain why breast cancer is rare before 30 years of age and if patients are diagnosed at early ages, tumours often show high S-phase values and bad prognostic signs. We postulate that these results from radiation induced breast cancer may be used to extrapolate possible latency times in patients with non radiation induced breast tumours in order to isolate possible time periods for research after other initiating events.

Potentiation of Antiproliferative Drug Activity by Lonidamine in Hepatocellular Carcinoma Cells

The ability of lonidamine (LND), a derivative of indazole-carboxylic acid, to modulate the cytotoxic activity of anticancer drugs was investigated in two human hepatocarcinoma (HCC) cell lines. The cytotoxicity of drugs used singly, in association or in sequence was evaluated using the Sulforhodamine B (SRB) assay. LND did not appreciably potentiate the effect of antitumor drugs when given before or simultaneously, in either cell line. Conversely, a synergistic interaction was observed in both cell lines when LND was given after conventional drugs. LND produced a moderate decrease in S-phase cell fraction and did not induce apoptosis. Conversely, paclitaxel (TAX) induced an important block in G2 and an increase in apoptosis. Following a 48-h TAX wash out, a progressive passage of cells from G2 to M phase was observed with a corresponding increase in apoptotic cells. Post-treatment with LND increased the cytotoxicity of some antitumor drugs, especially TAX, in hepatocarcinoma cells, possibly by preventing, as an energolytic drug, cell damage repair or by producing an additional effect on microtubule stabilization.

A congenic strain of rat for investigation of control of estrogen-induced growth.

A congenic strain of rat for investigation of control of estrogen-induced growth.

Increased radiosensitivity with chronic hypoxia in four human tumor cell lines

Purpose : It is well known that the radiosensitivity of tumor cells can be significantly reduced under hypoxic conditions. However, most of the reports in the literature refer to an experimental setup in which the supply of oxygen is kept low for a short period of time only. In tumors, chronic hypoxia would seem to be the more typical situation, because of an insufficient vascularization and the limited diffusion of oxygen into the tissue. Under such conditions, certain changes in the proliferation patterns of tumor cells, in which the cell cycle checkpoint protein p53 seems to play a role, have been shown to occur. We therefore decided to study radiosensitivity and cell cycle progression under conditions of chronic hypoxia in several human tumor cell lines differing in their p53 status. Methods and Materials : Four human tumor cell lines (melanomas Be11 and MeWo and squamous carcinomas 4197 and 4451) were incubated for 3 h, 24 h, and 72 h under either oxic or hypoxic conditions and subsequently exposed to graded doses of X-rays. In some cases, cells were kept under hypoxia for the same periods of time, but then reoxygenated immediately before irradiation. Cell survival was assessed with the usual colony formation assay, and cell cycle distributions were determined by two-parameter flow cytometry after labeling with bromodeoxyuridine (BrdU). Results : As expected, the oxygen enhancement ratio at 3 h was 2.0 or more in all cases. Differences, however, became evident with longer incubation times. At 24 h, the sensitivity of cells kept under hypoxic conditions both before and during irradiation was practically unchanged with cell lines Be11, 4197, and 4451, but clearly increased with MeWo. This resulted in an oxygen enhancement ratio of only 1.1 for the latter cell line when the sensitivity of aerated cells was used as reference. Cells kept under hypoxia for 24 h and reoxygenated shortly before irradiation, however, also showed an increase in sensitivity, so that the oxygen enhancement ratio based on differences in irradiation atmosphere alone was still around 2.0. At 72 h, the two p53 wild-type cell lines were not available for experiments, because they quickly degenerated under hypoxic conditions. Both mutant cell lines now showed similar results, the sensitivity being increased with irradiation under continued hypoxia as well as after reoxygenation. The oxygen enhancement ratios with reference to aerated cells were 1.3 and 1.5 for MeWo and 4451, respectively. Flow cytometric measurements after labeling with BrdU revealed that in all cell lines, the fraction of active S-phase cells during incubation tended to decrease under hypoxic conditions. Only in the p53 mutant cell lines, however, was this accompanied by an increase of the percentage of S-phase cells that were not actively incorporating BrdU. Conclusions : It is suggested that these quiescent cells in the S-phase compartment develop because of a general breakdown of cellular energy metabolism. In the p53 mutant cells, this may lead to a cessation of cell cycle progression in all phases alike, because checkpoint control has been lost; p53 wild-type cells, on the other hand, settle down preferentially in G 1 under the same conditions. Independently of the p53 status, however, energy depletion may be the cause of a decreased ability to cope with radiation damage and thus the cause of the observed increase in radiosensitivity. This would become more easily apparent in the p53 mutant cell lines, because they are less sensitive than the p53 wild types to hypoxia as such.

Determination of the fraction of S-phase cells in root meristems using bromodeoxyuridine labeling

The use of bromodeoxyuridine (BrdU) and subsequent immunocytochemical visualization for studying cell proliferation in plant meristems was investigated in Allium cepa L. root-tips. We describe the optimization of an indirect immunoperoxidase method for detecting incorporation of this DNA precursor in pulse-labeled cells. The basic object of this study is to quantify the extent to which the fraction of S-phase cells can reliably be estimated in asynchronous populations. A matrix of parallel labeling schedules with tritiated-thymidine or BrdU was developed, and the labeling indices provided by autoradiography or immunocytochemistry were compared. Thus, 0.5 mM BrdU assured saturation S-phase labeling after an exposure time of 30 min, and the mean length of the S-phase determined under such conditions was similar to that previously reported for this plant system. Interestingly, Feulgen staining did not interfere with subsequent detection of the BrdU probe. This allowed comparative evaluations of the nuclear DNA content by Feulgenmicrodensitometry and the position of a given cell in G1, S or G2 compartments. We also explored the possibility of quantifying BrdU-incorporation in single nuclei by densitometry measurement of the peroxidase label.

Determination of the fraction of S-phase cells in root meristems using bromodeoxyuridine labeling

The use of bromodeoxyuridine (BrdU) and subsequent immunocytochemical visualization for studying cell proliferation in plant meristems was investigated in Allium cepa L. root-tips. We describe the optimization of an indirect immunoperoxidase method for detecting incorporation of this DNA precursor in pulse-labeled cells. The basic object of this study is to quantify the extent to which the fraction of S-phase cells can reliably be estimated in asynchronous populations. A matrix of parallel labeling schedules with tritiated-thymidine or BrdU was developed, and the labeling indices provided by autoradiography or immunocytochemistry were compared. Thus, 0.5 mM BrdU assured saturation S-phase labeling after an exposure time of 30 min, and the mean length of the S-phase determined under such conditions was similar to that previously reported for this plant system. Interestingly, Feulgen staining did not interfere with subsequent detection of the BrdU probe. This allowed comparative evaluations of the nuclear DNA content by Feulgenmicrodensitometry and the position of a given cell in G1, S or G2 compartments. We also explored the possibility of quantifying BrdU-incorporation in single nuclei by densitometry measurement of the peroxidase label.

Vascular endothelial growth factor and prognosis in patients with node-negative breast cancer.

Vascular endothelial growth factor (VEGF) has emerged as one of the most important angiogenic growth factors from experimental in vitro and in vivo studies. In the present study, we investigated the relationship between VEGF expression and microvessel density (MVD) and defined their prognostic relevance on a series of 242 patients with node-negative breast cancer, using immunohistochemical methods. In parallel, estrogen and progesterone receptors were quantitatively assessed using the dextran-charcoal technique and cell proliferation was evaluated as S-phase cell fraction according to (3)H-thymidine-labeling index (TLI). The percentage of VEGF-expressing cells varied from 0-95% in the different tumors and was unrelated to menopausal status, tumor size or steroid receptor status. Conversely, a significant inverse relation was observed with patient age or tumor cell proliferation, albeit with very poor correlation coefficients. A significant relation was observed between VEGF expression and MVD (r(s) = 0.55, p < 0.001). Clinical outcome analyzed as a function of high and low VEGF expression showed slight differences in terms of both disease-free survival (DFS) and overall survival (OS) that never reached statistical significance. Moreover, the trend was paradoxically in favor of patients with highly VEGF-expressing tumors. Finally, DFS and OS curves, when analyzed as a function of VEGF expression or MVD, were superimposable. In conclusion, our study did not highlight a prognostic relevance of VEGF expression in patients with node-negative breast cancer, as already observed for MVD.

Cell proliferation markers in human solid tumors: Assessing their impact in clinical oncology

Publisher Summary Proliferation, a fundamental biological process, is involved in determining growth and maintenance of tissue homeostasis. This chapter describes several approaches used to evaluate the proliferative activity of human solid tumors. It examines the relevance of S-phase markers in breast, colorectal, and lung cancers. Cell proliferation of primary breast and non-small cell lung cancers is only weakly related to clinical and pathologic stage, and such a relation is generally observed in large case series in which minimal differences in mean or median thymidine labeling index (TLI) or flow cytometry within stage subsets reached statistical significance. For colorectal cancer, as for other solid tumors, surgical resection represents the best treatment option, but it does not prevent the risk of relapse, which depends on degree of local invasion, spread to regional lymph nodes, and tumor site and differentiation. Alterations in cell proliferating compartments have been shown to be the early event of colon mucosa transformation, and an increase in proliferating or S-phase cell fraction has been generally shown to be related to poor prognosis.

Flow cytometric DNA ploidy, p53, PCNA, and c-erbB-2 protein expressions as predictors of survival in surgically resected gastric cancer patients.

In order to determine retrospectively the impact of some cytometric and immunohistochemical parameters on the overall survival of gastric cancer patients treated with surgery alone, paraffin-embedded tumor samples from 137 gastric carcinoma patients undergoing curative resection from 1987-1993 were analyzed by flow cytometry (FCM) and immunohistochemistry (p53, c-erbB-2, and PCNA expression). FCM-derived parameters were DNA ploidy and fraction of S-phase cells (SPF). Multiple regression analysis was applied to determine the prognostic significance of the conventional clinicopathologic findings together with the flow cytometric and immunohistochemical parameters on overall survival. When all parameters were entered simultaneously into the Cox regression model, stage and DNA ploidy (DNA index >1.35) clearly emerged as the only independent prognostic factors. When the stages were analysed separately, the independent prognostic factors resulted DNA ploidy in early stages (I-II) and grading in stage IIIA tumors. For stage IIIB tumors, no independent prognostic factor was found. These results indicate that the DNA ploidy pattern is a valuable predictor of survival in curatively resected gastric cancer patients, especially when less advanced tumors are taken into consideration.

Total parenteral nutrition and tumor growth in malnourished patients with gastric cancer.

Evidence that total parenteral nutrition (TPN) can stimulate tumor growth in humans is scanty and contradictory. The purpose of this study was to evaluate the impact of TPN on tumor cell proliferation in malnourished patients receiving preoperative TPN. We evaluated variations in the S-phase cell fraction, defined as 3H-thymidine labeling index (TLI), before and after 10 days of TPN or non-administration of nutritional support in 19 malnourished patients (weight loss, > or = 10%) with gastric cancer. TLI was determined on endoscopic biopsies at the time of diagnosis, and subsequently on the operative specimen or through intraoperative sampling. At diagnosis, a higher median TLI value was observed in the control than in the TPN group. Administration of TPN enhanced tumor cell proliferation in 50% of patients; however, at surgery there was no difference in the median TLI value of the two groups. The TPN regimen seems to be associated with increased tumor cell proliferation, even though this stimulating effect was moderate and at surgery the TLIs of TPN patients and controls were not different. Although the potential stimulation probably has little consequence for patients receiving short-term preoperative nutrition, it may call for further investigation in cancer patients undergoing long-term home TPN.

Measurement of proliferation activity in human melanoma xenografts by magnetic resonance imaging

Tumor proliferation may be predictive for malignant progression and response to fractionated therapy of cancer. The purpose of the present work was to investigate whether the proliferation activity of solid tumors can be assessed in vivo from the proton relaxation times, T 1 and T 2. Tumors of four amelanotic human melanoma xenograft lines were studied. Three parameters were used to represent tumor proliferation activity; the volume doubling time, T vol, the potential doubling time, T pot, and the fraction of cells in S-phase. T vol was determined from volumetric growth data. T pot and S-phase fraction were determined by flow cytometric analysis of tumor cells after bromodeoxyuridine (BrdU) incorporation in vivo. T 1 and T 2 were measured by 1H-MRI in vivo, using spin-echo pulse sequences. The proliferation parameters and relaxation times differed considerably among the tumor lines. Significant correlations were found between the proliferation parameters and the relaxation times, regardless of whether T vol, T pot, or S-phase fraction was considered. Tumors with short T vol and T pot and high S-phase fraction had long T 1 and T 2 compared to tumors with long T vol and T pot and low S-phase fraction. The elongated T 1 and T 2 of fast growing tumors were probably due to increased interstitial and/or intravascular water content. The present results suggest that in vivo spin-echo 1H-MRI can be used to discriminate between tumors of high and low proliferation activity.

Reduced telomere DNA content is correlated with genomic instability and metastasis in invasive human breast carcinoma.

Telomere shortening leads to genomic instability and has been correlated with poor outcome in several types of cancer. A recently described, robust titration assay was used to quantify telomere DNA content in frozen and paraffin-embedded specimens of 49 invasive human breast carcinomas, including tumors with normal or abnormal contents of genomic DNA, which produced regional, distant, or local disease. Telomere DNA contents ranged from 53% to 370% of the content in a reference DNA purified from normal placenta. Tumors were divided into three groups of approximately equal size based on increasing telomere DNA content. All of 16 tumors in the group with the least telomere DNA (Group I), were aneuploid compared to 9/17 tumors in the group with the most telomere DNA (Group III). The Chi-square test for trend indicated that tumors with the least telomere DNA were significantly more likely to be aneuploid than tumors with the most telomere DNA (p < 0.002). Twelve of 14 tumors in Group I also produced metastatic disease compared to 8/15 tumors in Group III. The Fischer Exact Test indicated that tumors with the least telomere DNA were significantly more likely to be metastatic than tumors with the most telomere DNA (p < 0.05). There was no association between telomere DNA content and patients' age, tumors' size, grade, stage, or fraction of cells in S-phase. The correlation of reduced telomere DNA content with aneuploidy and metastasis, both of which are associated with poor outcome in invasive breast carcinoma, implies that telomere DNA content also could have prognostic value.

Assessment of cell proliferation by Ki-67 staining and flow cytometry in fine needle aspirates (FNAs) of reactive lymphadenitis and non-Hodgkin's lymphomas.

This study was undertaken to assess cell proliferation in FNAs from a series of 57 non-Hodgkin's lymphomas (NHL) and 11 cases of reactive lymphadenitis using Ki-67 staining and flow cytometry (FCM). The results were compared and correlated to the cytomorphological subgrouping according to Kiel classification. The mean percentages of Ki-67 positivity were 16.6% and 61.1% for low and high grade lymphomas, respectively (P < 0.001). The mean S-phase fraction (SPF) determined by FCM was 4.61% for low grade and 12.9% for high grade lymphomas (P < 0.001). The figures for Ki-67 positivity and S-phase fraction in reactive lymphadenitis were 16.8% and 40%, respectively. We observed a strong correlation in low grade lymphomas between Ki-67 and SPF. A good correlation was also found in reactive lymphadenitis. In high grade lymphomas, however, with highly scattered Ki-67 and S-phase values, this correlation was lost. In some cases this discrepancy can be explained by a rich admixture of non-neoplastic, non-proliferating cells in aspirates from diploid tumours. In addition, the existence of a minor aneuploid tumour cell population of high proliferation such as that in Ki-1 lymphomas will not be accurately analysed by FCM but is easily assessed by Ki-67 staining. However, the main reason seems to be a high variability between the fraction of cells in S-phase and the total number of cells in G1, S and G2 in individual tumours.

Low levels of cathepsin D are associated with a poor prognosis in endometrial cancer.

Total cytosolic cathepsin D (Cat D) levels were estimated by an immunoradiometric assay in a series of 156 consecutive patients with surgical stages I–III primary endometrial adenocarcinoma. Simultaneously, the tissue content of both oestrogen (ER) and progesterone (PR) receptors, and p185HER-2/neu, DNA content (ploidy), and the fraction of S-phase cells (S-phase) were also estimated. Tumoral Cat D content ranged from 0 to 243 pmol mg−1 protein (median 44 pmol mg−1 protein) and was not associated with any of the established clinicopathological and biological prognostic variables, with the exception of a weak positive correlation with the tumoral p185HER-2/neu levels. Univariable analysis performed on a subset of 97 patients, followed for a minimum of 2 years or until death, showed that patient age at diagnosis, high histological grade, advanced surgical stage, vascular invasion, positive peritoneal cytology, low levels of Cat D, negative ER and PR status, aneuploidy, and high S-phase were predictive of the presence of persistent or recurrent disease. However, multivariable analysis revealed that only histological grade, surgical stage, Cat D and PR were significantly associated with the patient's outcome. From these findings, we conclude that Cat D is an independent prognostic factor in endometrial adenocarcinoma, its low levels being associated with a worse clinical outcome. © 1999 Cancer Research Campaign

Biological markers as indicators of pathological response to primary chemotherapy in oral‐cavity cancers

The predictive role in terms of pathological response and prognostic role of biomarkers such as GST-π, p53, bcl-2 and bax expression, immuno-histochemically detected, and of the S-phase cell fraction, autoradiographically determined as thymidine labeling index (TLI), were investigated within a prospective randomized phase III clinical trial on squamous-cell carcinoma of the oral cavity, including surgery or primary chemotherapy (PCT), which foresaw the prospective determination of biological markers. Pathological response was defined as the achievement after PCT of a pathological complete remission or the presence of microresidual disease. The study was performed on tumors obtained from a series of 100 previously untreated patients with resectable T2–4N0–2M0 carcinoma. All biomarkers were unrelated, except for an inverse relation between TLI and GST-π and a direct relation between bcl-2 and bax expression. In patients treated with surgery alone, 3-year disease-free survival (DFS) appeared to be weakly, but not significantly, related only to GST-π and p53 expression. In patients treated with PCT, pathological response and DFS were independent of p53 expression and cell proliferation. Conversely, low GST-π and bax expression were indicative of pathological response but lost relevance as predictors of DFS, whereas absence of bcl-2 was associated with high probability of 3-year DFS in the overall series as well as in non-responding patients. Within this latter sub-set, all patients with bcl-2-positive tumors relapsed within 1 year of surgery, whereas a 60% probability of 3-year DFS was observed for patients with bcl-2-negative tumors (p= 0.02). This interim analysis appears to indicate that some biofunctional markers can provide information on pathological response to PCT and could help in understanding treatment efficacy at a cellular level. Int. J. Cancer (Pred. Oncol.) 79:619–623, 1998. © 1998 Wiley-Liss, Inc.

S-phase fraction assessed by a variant of the rectangular model adapted to the flow-cytometric DNA histogram profile.

Measurement of S-phase fraction by DNA flow cytometry is widely used in the analysis of human tumors. The calculation of fraction of S-phase cells is performed either by means of curve-fitting techniques or by more simple planimetric methods. Estimates by the latter often suffer from subjectivity or poor adaptation to complex DNA histograms. We describe a variant which is based on the rectangular model. The height of the rectangle is automatically determined from the lowest density value in the S-phase interval. The variant was tested on 141 DNA histograms, and the results were compared with those obtained with other variants based on the rectangular model. The new variant is thought to circumvent some of the problems concerning subjectivity and disturbances caused by aggregate peaks.

Prognostic and predictive value of thymidine labelling index in breast cancer.

In the last few decades, much effort has been directed towards identifying the phenotypic or functional aspect of tumor cells which can contribute to a biofunctional staging for improving the accuracy of pathologic staging used for identifying patients at different risk. Among known biologic factors, the proliferative capacity of the tumor cell population, a feature common to all tumors, has been widely investigated. Several approaches have been used to measure different aspects of the cell cycle. Among these, the thymidine labeling index (TLI) represents the fraction of cells in S-phase cell fraction and is based on the active incorporation of labelled thymidine into DNA. From basic studies conducted on several thousands of patients, the TLI of primary breast cancers appears closely related to steroid receptor status and generally unrelated to pathologic stage. Retrospective analyses performed on large series of patients treated with local regional therapy alone have consistently shown the relevance of TLI value to clinical aggressiveness in terms of relapse-free survival and overall survival. Moreover, TLI is a prognostic indicator which is independent of tumor size, steroid receptors, and p53 and bc12 protein expression, and which, together with patient age and tumor size, is able to identify patients at different risk of loco-regional or distant metastases. Recently, a direct relationship between TLI and response to polychemotherapy has been shown in patients with operable and advanced breast cancers. This finding, derived from retrospective and recently confirmed in prospective clinical studies, has led to the activation of cell kinetics based therapeutic protocols for patients with node-negative and one to three node-positive operable breast cancers.