Measurement Of Cellular Proliferation Research Articles

Adult IDH Wild-Type Glioblastoma Ultrastructural Investigation Suggests a Possible Correlation between Morphological Biomarkers and Ki-67 Index.

Glioblastoma is an aggressive brain tumor with an average life expectancy between 14 and 16 months after diagnosis. The Ki-67 labeling index (LI), a measure of cellular proliferation, is emerging as a prognostic marker in GBM. In this study, we investigated the ultrastructure of glioblastoma tissue from 9 patients with the same molecular profile (adult IDH wild-type glioblastoma, wild-type ATRX, and positive for TP53 expression, GFAP expression, and EGFR overexpression) to find possible ultrastructural features to be used as biomarkers and correlated with the only parameter that differs among our samples, the Ki-67 LI. Our main results were the visualization of the anatomical basis of astrocyte-endothelial cells crosstalk; the ultrastructural in situ imaging of clusters of hyperactivated microglia cells (MsEVs); the ultrastructural in situ imaging of microglia cells storing lipid vesicles (MsLVs); the ultrastructural in situ imaging of neoplastic cells mitophagy (NCsM). The statistical analysis of our data indicated that MsEVs and MsLVs correlate with the Ki-67 LI value. We can thus assume they are good candidates to be considered morphological biomarkers correlating to Ki-67 LI. The role of NCsM instead must be further evaluated. Our study findings demonstrate that by combining ultrastructural characteristics with molecular information, we can discover biomarkers that have the potential to enhance diagnostic precision, aid in treatment decision-making, identify targets for therapy, and enable personalized treatment plans tailored to each patient. However, further research with larger sample sizes is needed to validate these findings and fully utilize the potential of ultrastructural analysis in managing glioblastoma.

18F-3′-Deoxy-3′-Fluorothymidine Positron Emission Tomography Imaging for the Prediction of Acute Graft-Versus-Host Disease in Mouse Hematopoietic Stem Cell Transplant Models

Acute graft-versus-host disease (aGVHD) remains a barrier to the success of allogeneic hematopoietic stem cell transplantation. In mice, studies have demonstrated that donor conventional T cells traffic into host secondary lymphoid tissues early after transplant, and that this process is critical for the development of disease. As a result, the measurement of cellular proliferation within lymphoid sites early after transplant might be a useful approach for predicting aGVHD in humans. 18F-3′-deoxy-3′-fluorothymidine (FLT) positron emission tomography (PET) imaging has recently emerged as a functional imaging modality in oncology patients. FLT, a thymidine analog, is incorporated into replicating DNA and is thus an indirect marker of cellular proliferation. Here we report that FLT PET imaging can differentiate mice receiving alloreactive T cells and destined to develop lethal aGVHD from control mice. Mice receiving allogeneic T cells demonstrated a stronger FLT signal within the peripheral lymph nodes compared with control mice at all time points after transplant. In addition, allogeneic T cell recipients transiently demonstrated stronger FLT uptake within the spleen. Importantly, these differences were apparent before the development of clinical disease. In contrast, the FLT signal within the host bowel, an important aGVHD target organ, was more variable after transplant and was not consistently different between aGVHD mice and control mice. Collectively, these findings suggest that the imaging of patient lymphoid sites using existing FLT PET technology might be useful for predicting aGVHD in the clinical setting.

Click Chemistry for Analysis of Cell Proliferation in Flow Cytometry.

The measurement of cellular proliferation is fundamental to the assessment of cellular health, genotoxicity, and the evaluation of drug efficacy. Labeling, detection, and quantification of cells in the synthesis phase of cell cycle progression are not only important for characterizing basic biology, but also in defining cellular responses to drug treatments. Changes in DNA replication during S-phase can provide valuable insights into mechanisms of cell growth, cell cycle kinetics, and cytotoxicity. A common method for detection of cell proliferation is the incorporation of a thymidine analog during DNA synthesis. This chapter presents a pulse labeling method using the thymidine analog, 5-ethynyl-2'-deoxyuridine (EdU), with subsequent detection by click chemistry. EdU detection using click chemistry is bio-orthogonal to most living systems and does not non-specifically label other biomolecules. Live cells are first pulsed with EdU. After antibody labeling cell surface markers, fixation, and permeabilization, the incorporated EdU is covalently labeled using click chemistry thereby identifying proliferating cells. Improvements in click chemistry allow for labeling in the presence of fluorescent proteins and phycobiliproteins without quenching due to copper. Measuring DNA replication during cell cycle progression has cell health applications in flow cytometry, fluorescence microscopy, and high content imaging. This protocol has been developed and optimized for research use only and is not suitable for use in diagnostic procedures. © 2017 by John Wiley & Sons, Inc.

PETを用いた癌分子イメージング：頭頸部がん領域への応用

The imaging of specific molecular targets associated with cancer can be expected to allow earlier diagnosis and better management of oncology cancer patients. Positron emission tomography (PET) is a highly sensitive non-invasive technology that is ideally suited to clinical imaging of cancer biology. 2’-Deoxy-2’-18F-fluorodeoxy glucose (FDG) PET is now widely used for the diagnosis of malignant tumors and assessment of the therapeutic response. However, FDG is also metabolized at sites of inflammation and in other physiologically reactive organs, so that development of newer imaging probes with specificity for cancer are needed. The amount of DNA synthesis is a measure of cellular proliferation, and increased cellular proliferation is a feature of cancer. Thymidine analogs, 3’-deoxy-3’-18F-fluorothymidine (FLT) and 4’-[methyl-11C]-thiothymidine (4DST), have been introduced as stable cell proliferation imaging agents. Our recent researches have shown that FLT and 4DST PET show high sensitivity for the detection of head and neck cancer, despite the low uptake level of the tracers. In regard to assessment of the therapeutic response, FLT PET during treatment and early follow-up has the potential to predict therapeutic responses, with the ability to discriminate residual tumors from inflammatory changes. Volume-based parameters of pretreatment 4DST-PET can provide important prognostic information in patients with head and neck cancer.A depth-of interaction (DOI) detector is a key device to obtain any significant improvement of the sensitivity while maintaining high spatial resolution. DOI measurement also has the potential to expand the field of application of PET, because it allows for more flexible detector arrangement.New molecular imaging probes have also been developed for visualization of the brain and nervous system functions. Future applications are expected to allow analysis of the functions of sensory areas and diagnosis of sensory nerve disorders.

Biological basis of [11C]choline-positron emission tomography in patients with breast cancer

The biological significance of [¹¹C]choline (CHO) uptake in human tumours is unclear and probably linked to choline kinase-α (CHKα) expression and cell proliferation. We directly compared CHO with [¹⁸F]fluorothymidine (FLT), an imaging biomarker of proliferation, by positron emission tomography (PET) in patients with breast cancer to investigate whether cell proliferation is an important determinant of CHO uptake. Furthermore, we evaluated CHKα and the Ki67-labelling index (LIKi67) in tumour biopsies. Sequential CHO-PET and FLT-PET within the same imaging session were performed in 21 patients with oestrogen receptor (ER)-positive breast cancer (28 lesions). Average and maximum CHO standardized uptake values (SUV) at 60 min: SUV60,av, and SUV60,max, and the rate constant of net irreversible uptake, Ki, were compared with FLT uptake at 60 min: SUV60,av and SUV60,max. Biopsies were stained for CHKα and LIKi67 in eight cases. Tumours were equally visible on CHO-PET and FLT-PET imaging. Tumour CHO-PET strongly correlated with FLT imaging variables (Pearson's r=0.83; P<0.0001 for CHO-SUV60,max vs. FLT-SUV60,max). A statistically significant association was found between CHO-PET variables and categorical scores of cytoplasmic CHKα intensity and between FLT-PET and LIKi67 (P<0.05, one-way analysis of variance). Choline metabolism and proliferation as assessed by PET were correlated in ER-positive breast cancer, indicating that high CHO uptake is a measure of cellular proliferation in this setting. CHO uptake was also found to be related to cytoplasmic CHKα expression.

Inhibition of gene expression in Entamoeba histolytica with antisense peptide nucleic acid oligomers.

Peptide nucleic acids (PNAs) may be a potent tool for gene function studies in medically important parasitic organisms, especially those that have not before been accessible to molecular genetic knockout approaches. One such organism is Entamoeba histolytica, the causative agent of amebiasis, which infects about 500 million people and is the cause of clinical disease in over 40 million each year, mainly in the tropical and subtropical world. We used PNA antisense oligomers to inhibit expression of an episomally expressed gene (neomycin phosphorotransferase, NPT) and a chromosomal gene (EhErd2, a homolog of Erd2, a marker of the Golgi system in eukaryotic cells) in axenically cultured trophozoites of E. histolytica. Measurement of NPT enzyme activity and EhErd2 protein levels, as well as measurement of cellular proliferation, revealed specific decreases in expression of the target genes, and concomitant inhibition of cell growth, in trophozoites treated with micromolar concentrations of unmodified antisense PNA oligomers.

Low Temperature Antigen Restoration of MIB-5 Immunoreactivity in Rat Colon

Although the monoclonal antibody Ki-67 has obviated the need for counting mitoses as a measure of cellular pro liferation, it is formalin sensitive and works only in fresh frozen sections. The MIB antibodies recognize the same epitope and can be used in both frozen and fixed tissues; however, high temperature antigen retrieval is necessary to unmask the effects of formalin fixation. In this study, we examined the use of low temperature antigen retrieval (LTAR) for unmasking the Ki-67 epitope in rat tissues to restore immunoreactivity of MIB-5. LTAR reduced section detachment without compromising sensitivity. (The J Histotechnol 22:3 19, 1999)

Low Temperature Antigen Restoration of MIB-5 Immunoreactivity in Rat Colon

Although the monoclonal antibody Ki-67 has obviated the need for counting mitoses as a measure of cellular pro liferation, it is formalin sensitive and works only in fresh frozen sections. The MIB antibodies recognize the same epitope and can be used in both frozen and fixed tissues; however, high temperature antigen retrieval is necessary to unmask the effects of formalin fixation. In this study, we examined the use of low temperature antigen retrieval (LTAR) for unmasking the Ki-67 epitope in rat tissues to restore immunoreactivity of MIB-5. LTAR reduced section detachment without compromising sensitivity. (The J Histotechnol 22:3 19, 1999)

Breathing pattern response and epithelial labeling in ozone-induced airway injury in neutrophil-depleted rats.

To test the hypothesis that neutrophils enhance the repair of ozone (O3)-injured airway epithelium, we investigated breathing pattern responses and airway epithelial injury and repair in rats depleted of neutrophils using rabbit antirat neutrophil serum (ANS) and control rats treated with normal rabbit serum (NRS). Thirty-seven Wistar rats were exposed to O3 (1 ppm) or filtered air (FA) for 8 h followed by 8 h in FA. O3-exposed NRS- and ANS-treated rats showed similar progressive decreases in tidal volume and increase in breathing frequency, with maximal changes occurring at 8 h of exposure, whereas FA-exposed rats showed no significant changes. O3-exposed ANS-treated rats showed more epithelial necrosis in the nasal cavity, bronchi, and distal airways than did O3-exposed NRS-treated rats. Incorporation of 5-bromo-2-deoxyuridine (BrdU), a measure of cellular proliferation, was assessed using an optical disector to count BrdU- labeled terminal bronchiolar epithelial cells. O3-exposed ANS-treated rats had significantly less BrdU- labeled epithelial cells than did O3-exposed NRS-treated rats. We conclude that neutrophils contribute to the repair process by enhancing the proliferation of O3-injured airway epithelial cells.

Expression and activity of L-Myc in normal mouse development.

To determine the role of L-Myc in normal mammalian development and its functional relationship to other members of the Myc family, we determined the normal patterns of L-myc gene expression in the developing mouse by RNA in situ hybridization and assessed the phenotypic impact of L-Myc deficiency produced through standard gene targeting methodology. L-myc transcripts were detected in the developing kidney and lung as well as in both the proliferative and the differentiative zones of the brain and neural tube. Despite significant expression of L-myc in developing mouse tissue, homozygous null L-myc mice were found to be viable, reproductively competent, and represented in expected frequencies from heterozygous matings. A detailed histological survey of embryonic and adult tissues, characterization of an embryonic neuronal marker, and measurement of cellular proliferation in situ did not reveal any congenital abnormalities. The lack of an apparent phenotype associated with L-Myc deficiency indicates that L-Myc is dispensable for gross morphological development and argues against a unique role for L-Myc in early central nervous system development as had been previously suggested. Although overlapping expression patterns among myc family members raise the possibility of complementation of L-Myc deficiency by other Myc oncoproteins, compensatory changes in the levels of c- and/or N-myc transcripts were not detected in homozygous null L-myc mice.

Prognostic Value of Cellular Proliferation and Histologic Grade in Follicular Lymphoma

The clinical usefulness of histologic grading in follicular lymphoma (FL) is controversial and is further compromised by the subjective nature and poor reproducibility of most systems in current use. Therefore, we decided to objectively evaluate the importance of cellular proliferation in FL, along with the current grading systems. We studied 106 patients with FL who were uniformly staged and aggressively treated. A proliferative index (PI) was determined quantitatively using an automated image analyzer and a new Ki-67 antibody that stains archival paraffin tissues. The cases were also subclassified according to the Berard, Rappaport, Luke-Collins, and Jaffe methods, and survival analysis was performed. Patients with a low PI (< 40%) had a significantly longer overall survival (OS) than those with a high PI (> or = 40%), but the PI did not predict failure-free survival (FFS). The mean PI correlated well with the subgroups in each of the various classifications. All four of the classification methods were predictive of OS, but only the Berard method appeared to predict FFS and suggest that a proportion of patients with FL may be curable. In multivariate analysis, histologic classification was the only independent predictor of OS (Berard method: relative risk, 3.1) and the International Prognostic Index was the only independent predictor of FFS (relative risk, 2.3). We conclude that the Berard method for grading of FL is clinically useful and, along with the International Prognostic Index, should be included in future clinical studies of FL. The measurement of cellular proliferation does not appear to add additional useful information in FL.

Cellular proliferation in prostatic adenocarcinoma as assessed by bromodeoxyuridine uptake and Ki‐67 and PCNA expression

Prostatic carcinomas vary in their biological potential, even when stratified by grade and stage. Measurement of cellular proliferation by various methods has been shown to correlate with outcome for several human cancers, including prostatic carcinoma. Uptake of bromodeoxyuridine (BrdUrd), a thymidine analogue, has been accepted as a measure of cellular proliferative rate. However, the technique is somewhat complex, requiring incubation with fresh tissue. We compared cellular proliferation as measured by BrdUrd uptake with two more simple immunohistochemical methods in 44 prostatic adenocarcinoma specimens and correlated the results with standard clinical parameters. The tissue was obtained via needle biopsy, channel transurethral resection, and radical prostatectomy. Specimens were incubated in vitro with BrdUrd and then fixed and paraffin embedded. Sections were immunohistochemically stained with antibodies to BrdUrd, proliferating cell nuclear antigen (PCNA), and Ki-67. At least 1,000 cells were scored, and a labeling index (LI) was calculated (number of positive cells/total number of cells). The mean LI determined by all three indices was low (BrdUrd = 3.0, PCNA = 7.0, Ki-67 = 3.4), consistent with the knowledge that prostatic tumors grow slowly. In 36 patients who had not been treated at the time of analysis, the LI as determined by all three methods correlated well with clinical stage and pathological grade. Furthermore, the LIs discriminated between those with tumor confined to the prostate and those with extension to the seminal vesicles, lymph nodes, or bone (P = 0.003, 0.004, 0.008 for BrdUrd, PCNA, and Ki-67, respectively). The LIs for PCNA and Ki-67 correlated well with that for BrdUrd (r = 0.84; r = 0.85), while the LIs for Ki-67 and PCNA correlated slightly less well with each other (r = 0.78). PCNA and Ki-67 expression appear to reflect essentially the same biological process as BrdUrd uptake. Either can substitute for BrdUrd as a measure of cellular proliferation, and Ki-67 seems to offer the fewest technical problems.

Comodulation of cellular polyamines and proliferation: biomarker application to colorectal mucosa.

Polyamines are low molecular weight aliphatic amines required for normal cellular growth which are ubiquitously found in all living tissues. Polyamine biosynthesis is known to increase with mitogenesis, and elevated polyamine concentrations are found in hyperproliferative tissues. Quantitation of tissue polyamine content may thus provide a biochemical measure of proliferation, with potential biomarker application to the colonic mucosa where dysregulated epithelial proliferation is associated with cancer risk. This study was performed to validate polyamine analyses as a measure of cellular proliferation, and to preliminarily assess polyamine characteristics when applied to clinical samples. Using FHC, a human colonic epithelial cell line, for in vitro experimentation, deoxycholic acid or retinol was added to freshly passaged cultures to either stimulate or inhibit proliferation, respectively. Parallel cultures were then assayed for (1) proliferation by sulforhodamine B staining; and (2) polyamine content by a high-performance liquid chromatographic method. Deoxycholic acid stimulated, and retinol inhibited proliferation in dose-dependent fashion. Polyamine content, specifically the spermidine content and the spermidine/spermine ratio, also increased or decreased in response to culture with deoxycholic acid or retinol, respectively. Significant linear correlations between proliferation and spermidine (r = 0.858, P < 0.001), and with the spermidine/spermine ratio (r = 0.574, P < 0.05) were observed. When quantitative polyamine analyses were applied to human colonic specimens, replicate mucosal sampling revealed a high degree of intra-individual variability, indicating a heterogeneous distribution of polyamines within anatomically confined colonic segments. The results support a role for quantitative polyamine analyses as a correlative measure of colonic epithelial proliferation; however, intraindividual variability may limit the utility of colorectal biomarker measurements.

Sulindac and polyp regression.

Sulindac is useful in regression of adenomatous polyps. In addition to orally administered sulindac, rectal preparations also appear to be efficacious [32]. However, further studies are necessary to determine whether regression of adenomas, the precursor of colorectal cancer, will cause decrease in colorectal cancer risk in both FAP and non FAP patients. Moreover, clinical studies are needed to test the application of this potential chemopreventative drug in several other patient populations. Several interesting observations have been made concerning the use of sulindac. Indomethacin, a related NSAID to sulindac, did not cause polyp regression. Also, upper gastrointestinal tract polyps in the stomach and duodenum appear not to be affected by sulindac therapy. These observations might be explained by the metabolism of sulindac in which the pharmacologically active sulfide metabolite is generated and distributed in the large intestine. Also, investigators noted that after discontinuation of sulindac adenomas recurred. Sulindac treatment was well tolerated at the usual clinical doses. Although some investigators have speculated on the effect of prostaglandin inhibition sulindac on cAMP-dependent mechanisms which control cellular proliferation [33], the cause of adenoma regression is unknown. There is evidence that colorectal endogenous prostaglandin levels decrease with sulindac. Evaluation of colorectal mucosal cellular proliferation in patients treated with sulindac has revealed a decrease in labelling index by bromodeoxyuridine labeling in one study [28] but no change in labelling index by [3H]thymidine incorporation in another investigation [34]. Also, ki 67 labelling index, another measure of cellular proliferation, was not affected in the colorectal mucosa of patients taking sulindac.(ABSTRACT TRUNCATED AT 250 WORDS)

Influences on Measurement of Cellular Proliferation by Histology and Flow Cytometry in Mammary Carcinomas LabeledIn Vivowith Bromodeoxyuridine

Field-selection procedures strongly influence histologically determined bromodeoxyuridine (BrdU) labeling indices (HLIs): For a series of 52 breast carcinomas, "high-labeling" fields had mean 2,000-cell HLIs that were, on average, more than double those for random fields, but the ratio was extremely variable. Although there may be a priori reasons for preferring particular field-sampling techniques, nonstandardized kinetic data from different laboratories will be uninterpretable and therefore of limited clinical application. Also, if heterogeneity of tumor cell proliferation is neglected, it is possible that incomplete or even misleading data may be gained. Total flow cytometric BrdU labeling indices (TLIs) were not, on average, higher or lower than HLIs for the same cases, but individual discrepancies could be pronounced. The best correlation of flow cytometric LIs with HLIs was obtained for aneuploid carcinomas with a restricted labeling index derived for aneuploid (presumptively neoplastic) cells only. Although flow cytometric analysis avoids the statistical problems associated with accurately determining a small percentage from relatively few cells, it lacks precise definition of the character of the cells counted, but even histologic methods are not entirely free of this difficulty.

Measurement of cellular proliferation in human prostate by AgNOR, PCNA, and SPF.

Tumor differentiation and proliferative activity are important predictors of biological behavior. While routine histological evaluation is fairly adequate to assess differentiation, tumor proliferative activity is difficult to measure. Silver staining for nucleolar organizer regions (AgNORs) is reported to be helpful for assessing tumor proliferation. We investigated the AgNOR counts in 20 formalin fixed, paraffin embedded human prostate tissues in three microscopic fields of 330X, using an image analysis system. A total of 200-700 nuclei were evaluated on histologically controlled areas of nonneoplastic prostate tissue, prostatic intraepithelial neoplasia (PIN), and invasive carcinoma. The values were compared to flow cytometrically obtained synthesis phase fractions (SPF) and immunohistochemically semi-quantitated, proliferative cell nuclear antigen (PCNA) patterns. AgNOR counts were also compared to tumor stage and Gleason's score. The pattern of PCNA staining in formalin fixed specimens was widely variable, probably due to differences in preservation of antigen. The positive counts varied from 0 to 55%, with a mean value of 8.55 +/- 15.9. The SPF values ranged from 5 to 13% with a mean value of 8.50 +/- 2.37. Two of 20 tumors were aneuploid and 18 were of diploid range. The mean AgNOR values in nonneoplastic nuclei (1.836 +/- 0.299), PIN (3.129 +/- 0.295), and invasive tumor cell nuclei (4.737 +/- 0.369) were highly significant (P < 0.0001) when paired differences were compared. AgNOR counts correlated significantly with tumor Gleason's score (P < 0.0145). However, the correlation coefficient for SPF and AgNOR values was not significant (P > 0.24), possibly because of the small number of samples examined. The highest AgNOR counts were found in the two aneuploid tumors. We conclude that AgNOR count may be a potential indicator of cellular proliferation, and possibly a marker of tumor differentiation.

Nutrition and tumor promotion: in vivo methods for measurement of cellular proliferation and protein metabolism.

The notion that tumors act as "nitrogen traps" has led to the belief that nutrition support of the cancer-bearing patient can enhance tumor growth. Proponents of this theory consider the provision of energy and essential nutrients as well as the influence of hormones and growth factors as responsible for this effect. On the other hand, nutrition administration in the debilitated cancer patient may improve antitumor host defense mechanisms and reduce tumor growth. This paper reviews methodologic issues related to the study of nutrition and cancer growth with emphasis on in vivo methods for measuring tumor protein turnover and cytokinetics. Using this combined approach, we previously demonstrated that dietary fat may significantly regulate tumor growth during chronic feeding as well as with short-term intravenous nutrition support in experimental models. Although the mechanism of this effect remains unclear, we have reasoned that by altering arachidonic acid metabolism and prostaglandin synthesis, omega-3 fatty acids could change tumor protein breakdown rates and inhibit the proliferation potential of these tumors. Acknowledging alternative hypotheses, we now present cytokinetic evidence that intracellular protein degradation may regulate tumor cell proliferation. Additional studies relating dietary fat, tumor protein metabolism and tumor proliferation potential are currently in progress. We propose that the effect of nutrition administration on tumor growth is complex and involves several regulatory systems. Thus, based on available evidence, an a priori tumor-enhancing effect for nutrition support is clearly not warranted. Intracellular protein breakdown and host defense mechanisms, both of which are energy dependent, are important loci at which nutrition and tumor growth regulation could interact.(ABSTRACT TRUNCATED AT 250 WORDS)

Measurement of cellular proliferation within the vitreous during experimental proliferative vitreoretinopathy.

Cellular proliferation is an important component in the pathogenesis of complex retinal detachments caused by proliferative vitreoretinopathy (PVR). We used flow cytometry to measure the proliferation of cells recovered from the vitreous cavity in an experimental model of tractional retinal detachment induced by homologous fibroblast injection. Two distinct populations of cells were detected. One population comprised smaller, dense cells representing mostly leukocytes. A second population of larger cells contained not only the injected fibroblasts but also high concentrations of host-derived cells with significant proliferative activity, whose number increased for 3 days following injection. Flow cytometry was useful for analysis of the recovered cells for concentration, morphologic features and proliferation. This technique may be applicable in the identification of patients at high risk for the development of PVR.

Ploidy as a Prognostic Factor in Ovarian Cancer

The cellular DNA content of 50 ovarian common epithelial carcinomas was determined by flow cytometry, and tumours were classified as being either diploid or aneuploid. A significant association between tumour stage and ploidy was demonstrated, with all diploid tumours being of an early stage (p less than 0.001). Forty percent of early-stage tumours (FIGO stages I and II) and all late-stage tumours (FIGO stages III and IV) were aneuploid. This heterogeneity with respect to DNA content among tumours of a similar stage may allow the identification of neoplasms with a different natural history. The proportion of S-phase cells determined by flow cytometry is a measure of cellular proliferation and may also be of prognostic significance. Diploid tumours had a median S phase of 9.8% (2.4-14.1%), while aneuploid tumours had a significantly higher S phase of 19.6% (7-24.7%; p less than 0.05). In this study there was no relationship between histological grading of invasive carcinomas and ploidy, but in view of the relatively small numbers and limited follow-up, it was not possible to perform a multivariate analysis of all known prognostic factors in ovarian cancer. Our results suggest that ploidy reflects tumour behaviour, but prolonged follow-up and increased patient accrual is necessary to assess whether the flow cytometric analysis of DNA content will provide clinically important information in ovarian cancer.

Vascular Smooth Muscle Cell Kinetics: a New Assay for Studying Patterns of Cellular Proliferation in Vivo

A new quantitative assay for studying the kinetics of vascular smooth muscle cells in vivo is reported. The assay was used to determine the specific activity of DNA from rabbit aortic smooth muscle cells stimulated to grow by removal of the endothelial layer. The specific activity of the DNA was correlated with the rate of tritiated thymidine incorporation as measured by autoradiography and with the rate of DNA synthesis as estimated by direct measurement of cellular proliferation. Smooth muscle cells exhibit a 24-hour latent period in vivo prior to DNA synthesis; the synthesis peaks at 48 hours and then rapidly declines. The decline in DNA synthesis is not related to endothelial regrowth, and may be of homeostatic significance in limiting luminal stenosis. The assay offers a rapid and reliable alternative to autoradiographic and morphometric techniques for evaluating growth kinetics and growth regulation in vivo.