Determination Of Labeling Index Research Articles

Nuclear β-catenin positivity as a predictive marker of long-term survival in advanced epithelial ovarian cancer

BackgroundClassical features as histomorphology, grade, FIGO stage, and residual tumour mass have strong prognostic value in advanced epithelial ovarian carcinomas (AEOC). Most AEOCs are associated with early recurrence and poor overall survival (OS). Despite of early recurrence, general poor outcome, both high grade tumours or tumours with advanced FIGO stage at the time of diagnosis, in some of such cases, long-term survival (LTS) has been recorded . The aim of this study was to compare the utility of “classical” prognostic factors to molecular factors such as β-catenin- E-cadherin-, mutated TP53-, and MiB-1 (Ki-67) labelling index determination in predicting long-term survival. MethodsThe expression of β-catenin, E-cadherin, Ki-67, and p53 was determined by immunohistochemistry (IHC) in AEOC. Correlation was sought for between expression of these proteins and the status of classical features vis-á-vis overall survival of patients. Statistical evaluation of the data included Kaplan–Meier analysis, the log-rank test and Cox proportional hazards model. ResultsAs expected, residual tumour size was an independent adverse prognostic factor for OS (univariate analysis: p=0.003, multivariate analysis: p=0.005). Nuclear expression of β-catenin in advanced ovarian cancer of LTS patients proved to be not only an independent favourable predictor of OS (univariate analysis: p=0.025, multivariate analysis: p=0.041) but also showed strong correlation with platinum sensitivity and platinum re-induction. ConclusionsTranslocation of stabilized β-catenin from cytoplasm to the nucleus plays an important role in predicting platinum sensitivity. It also seems to support the chance for platinum re-induction in AEOC and thus enhances long-term survival.

Cell proliferation in the gastric epithelium of the ulcer rat

Objective. Cell division is brisk in the ulcer margin and many of the new cells will migrate over and cover the ulcer bed. The aim of this study was to determine how agents that promote or delay gastric ulcer healing influence cell proliferation in the gastric epithelium. Material and methods. Acetic acid ulcers were produced in the rat gastric corpus; non-ulcer rats served as controls. All rats were given a continuous infusion of 3H-thymidine. Some rats were also given gastrin or indomethacin, or infected with Helicobacter pylori. The rats were killed after 1, 2, 6 or 13 days, and the ulcer margin and undamaged corpus were excised for determination of labeling index (LI) by autoradiography. Antrum, duodenum and colon were also studied. Silver grain counting was carried out in some groups. Results. LI in the ulcer margin grew exponentially, reaching 84% after 6 days; gastrin increased, and indomethacin decreased LI significantly. In 6-day ulcer rats that were given 3H-thymidine only during the first day LI was 5%, while in those given 3H-thymidine only during the last day LI was 27%. LI and silver grain counting results indicated that during the first 6 days of healing the epithelial cells in the ulcer margin divide twice. In the undamaged epithelium of the 1-day ulcer rats LI was <one-third, and in the antral epithelium one-half of that in the non-ulcer controls. Conclusions. During the first 6 days of healing, the LI of the non-parietal epithelial cells increases exponentially in the ulcer margin; this is enhanced by gastrin. The unexpected finding in the ulcer rats of decreased LI in undamaged oxyntic and antral epithelium may be caused by a block of the G1 restriction point in the cell cycle. The reason for this remains obscure.

Long-term follow-up of germinoma after stereotactic biopsy and brain radiotherapy: a cell kinetics study.

The primary aim of this study is to report the long-term outcome of pineal and suprasellar germinoma after stereotactic biopsy and whole brain radiotherapy. The second purpose is to report an investigation of the biological features and cell kinetics of this peculiar and enigmatic brain tumour. Of 34 supratentorial germ cell tumours diagnosed and treated between 1980 and 1993, 20 patients were found to be affected by true germinoma localized in the pineal and/or suprasellar regions. The diagnosis was achieved by stereotactic biopsy in all cases. In 14 patients, the potential proliferative activity of the tumour was investigated by (3H)thymidine in vitro binding and labelling index determination. Chorionic gonadotropin, alpha-fetoprotein and embryonal carcinoma antigen were negative in the cerebrospinal fluid of these patients. All but 1 patient underwent whole brain radiotherapy. Clinical and neuroradiological follow-up ranged between 3 and 13 years (mean 8). Complete clinical and neuroradiological recovery was achieved in all patients after treatment. Fatal recurrences owing to neuraxis dissemination occurred in three cases. The labelling index in the whole series ranged between 0.1 and 5% (median 2.5). Only syncytiotrophoblastic cells had proliferative activity, while none of the lymphoid-like cells showed thymidine labelling.

Inoculation of VacA− and CagA− Helicobacter pylori Delays Gastric Ulcer Healing in the Rat

Background: Helicobacter pylori is associated with peptic ulcer disease. In the present study, the influence of VacA and CagA− H. pylori on gastric ulcer healing was studied in the rat. Methods: Twenty-four rats with acetic-acid-induced gastric ulcer were divided into two groups and given either vehicle (Brucella broth) or H. pylori suspension by gavage every 12 h for 7 days. The animals were killed 1 and 8 days after the last H. pylori gavage (i.e. 10 and 17 days after the induction of the ulcer). One hour before death, 3H-thymidine was given intraperitoneally. Tissue samples from the stomach (including the ulcer area) and the duodenum were processed for determination of labelling index and apoptotic cells. Results: Compared with the vehicle-treated controls, the ulcer area in H. pylori-inoculated rats was significantly larger, the epithelial apoptotic cells in the ulcer margin and intact corpus were more numerous, while the cell proliferation of gastroduodcnal epithelium was slightly, but not significantly, increased by H. pylori gavage. Conclusion: Gastric ulcer healing was delayed after the inoculation of VacA− and CagA H. pylori in the rat, possibly as a result of excess cell loss by apoptosis.

2097 Labelling index determination in carcinoma of the uterine cervix treated by brachytherapy and hysterectomy

2097 Labelling index determination in carcinoma of the uterine cervix treated by brachytherapy and hysterectomy

Randomized, Double-Blinded, Placebo-Controlled Intervention Study With Supplemental Calcium in Families With Hereditary Nonpolyposis Colorectal Cancer

A high-fat diet has been recognized for some time as a major risk factor for colorectal cancer. It is thought that fat promotes this disease by increasing the levels of fatty and bile acids within the colon. These acids irritate and damage the epithelial cells of the colon. As a result of this cellular destruction, an increase in the rate of cellular proliferation occurs. Oral calcium supplementation has been proposed as a dietary intervention for individuals at high risk of colorectal cancer because of its ability to reduce rectal epithelial cell proliferation through the binding of fatty and bile acids. Placebo-controlled studies, however, have yielded varying results. We conducted a randomized, double-blinded, placebo-controlled trial to test oral calcium supplementation in patients at high risk of developing hereditary nonpolyposis colorectal cancer. Thirty subjects at risk for this cancer, with an increased epithelial cell proliferation along the colon and rectum, were randomly assigned to either a placebo group (n = 15) or a treatment group (n = 15). They received either oral calcium carbonate (CaCO3) supplements (1.5 g) or placebo (cellulose and starch) three times a day during a 12-week period. Colonic biopsy specimens (rectal, sigmoidal, and descending) were obtained prior to and after the intervention trial, during endoscopy, for determination of labeling index (LI) of whole crypts and crypt compartments by 5-bromo-2'-deoxyuridine incorporation and immunohistochemistry. Proportional bile acid compositions in duodenal bile and cytolytic activity of fecal water were also determined. All P values represent two-tailed tests of statistical significance. Statistically significant reductions, comparing before with after intervention, in rectal whole-crypt LI after receiving either calcium supplements (from 10.9% +/- 5.2% [mean +/- SD] to 6.2% +/- 1.5%; P < .02) or placebo (from 11.7% +/- 4.7% to 8.2% +/- 3.1%; P < .05) were observed. In the three bowel segments, no statistically significant differences were observed between the supplemental calcium and placebo groups. A statistically significant reduction in cytolytic activity was determined during calcium supplementation (from 57% +/- 41% to 32% +/- 30%; P < .05), whereas in the placebo group, it did not change (from 42% +/- 41% to 36% +/- 27%; P > .10). Oral calcium supplementation was shown to cause only a minor nonstatistically significant reduction of epithelial cell proliferation in the rectum, compared with placebo, and to have no effect on the same parameter in the sigmoid and descending colon in first-degree relatives of hereditary nonpolyposis colorectal cancer patients. These results cast doubt on the value of calcium supplementation in the prevention of colorectal cancer, especially in individuals already consuming an adequate amount of dietary calcium.

In vivo determinations of labelling index of metastatic colorectal carcinoma and normal colonic mucosa using intravenous infusions of bromodeoxyuridine.

In vivo determination of the labelling index of metastatic colon carcinoma was performed in 14 patients following intravenous infusion of bromodeoxyuridine (BrdU), at doses of 100 and 50 mg/M2. No toxicity was seen. Labelled cells were detected by an immunohistochemical technique using a monoclonal antibody to BrdU. Labelling was uniform and highly specific with good preservation of morphology. Labelling index values ranged from 15.0-40.6% for metastatic lesions and 3.6%-20% for normal colonic mucosa, with mean values of 24% and 11%, respectively. This represents a useful new method for in vivo labelling index determination in solid tumors.

In VitroBrdUrd Labeling of Solid Tumors and Immunological Determination of Labeling Index

AbstractThe use of radioisotope-labeled DNA precursors like tritiated thymidine has made research into cell kinetics of human cancer difficult because of technical problems and the ethical problems involved in the use of human subjects. Once incorporated into the DNA of patients, these compounds remain there permanently. We have developed a simple and reliable combination of in vitro bromodeoxyuridine (BrdUrd) labeling and monoclonal immunocytochemical staining of biopsy and surgical specimens that provides an accurate estimate of DNA synthesizing cells within several hours after sampling. This method opens the way to realization of “dynamic” histopathology because neither radioisotopes nor time consuming autoradiography is required.

Immunological determination of labeling index on human tumor tissue sections using monoclonal anti-BrdUrd antibody.

The use of a monoclonal antibody against the thymidine analogue bromodeoxyuridine together with an in vitro labeling technique allowed rapid determination of the labeling index in human tumors. The labeling index estimated by these relatively simple immunofluorescence or immunoenzymatic staining methods was equivalent to that obtained by autoradiography. The interpretation of the preparations is easy since there is a minimum of background staining. This immunohistochemical technique combined with in vitro labeling provides a suitable alternative for determining the labeling index of human tumors.

Comparative study of the cellular replication kinetics of rat mammary epithelial cells in vivo and in vitro

Differential staining techniques based on the incorporation of 5-bromo-2′-deoxyuridine (BrdU) into DNA permit the unequivocal identification of metaphrase cells which have replicated once, twice and three or more times. This technique is used to analyze the cell replication kinetics of mammary epithelial cells from young virgin rats in culture. Comparison is made between these in vitro observations and expected results derived from in vivo determination of labelling index, growth fraction, cell cycle length and distribution for the different structures and cell types of the rat mammary gland. The correlation coefficient for the comparison between in vivo and in vitro results is 0.84 suggesting that the heterogeneity of cell population and replication kinetics of the rat mammary gland is preserved when cells are placed in culture. Furthermore, the BrdU incorporation system for the analysis of cell replication kinetics is accurate and acceptable for the study of rat mammary epithelial cell kinetics.

Control of replication by dietary lipids and namely by linoleic acid in liver and adipose tissue of developing rats

Pulse labeling of adipose tissue and liver by injection of [Me 3H]thymidine in developing rats receiving 20% dietary lipids suggests a dual function of this molecule in nuclei: tracer incorporation into newly formed DNA, and tracer incorporation into RNA linked to nascent DNA. This conclusion is based on (1) determination of the specific radioactivity (SRA) of long alkali-stable nucleotide sequences which increased dramatically during 1 hr after radioactive injection when rats were given 20% lard (L) containing little linoleic acid, and decreased sharply thereafter; (2) isopycnic centrifugation of the native DNA extracted from nuclei pulse labeled in vivo with [Me 3H]thymidine and [6- 14C]orotic acid, which indicated the presence of doubly-labeled constituents with the buoyant density of DNA; (3) isopycnic centrifugation after freeze-thawing of the same samples, which indicated the disappearence of 14C-labeled long alkali-stable nucleotide sequences in the tritium labeled material with the buoyant density of DNA. A transient elevated level of radioactivity was also observed by autoradiography and determination of labeling index in adipose cell nuclei. The labeling index was 2.3-fold higher 1 hr after injection of [Me- 3H]thymidine in (L) group adipose tissues than in (SO) adipose tissues, when rats were given 20% sunflower oil containing large proportion of linoleic acid. In contrast, the labeling index in (SO) nuclei was 1.8-fold higher than in (L) nuclei 2 hr after injection. The labeling index was much lower in nuclei of both groups 3 hr after injection. Moreover, the striking increase in SRA was not observed in nuclei after injection [6- 3H]thymidine instead of [Me- 3H]thymidine, and was never observed in mitochondria, whichever labeled thymidine was injected. Therefore, high lipid intake appears to increase methyl group requirement, and this effect seems to be more pronounced when essential fatty acid is given in small proportion. This relative methyl deficit seemed to be filled up in RNA linked to nascent DNA, by incorporation of labeled thymine during the early process of replication in adipose cells and liver of developing rats.