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T G Pretlow

Publications and source records attributed to T G Pretlow.

At least 19 recordsLinked to original sources
PubMed1992

Aberrant crypts in human colonic mucosa: putative preneoplastic lesions.

Aberrant crypts are recognized in methylene blue-stained, unsectioned, colonic mucosa by their increased size, elliptical lumenal opening, thicker epithelial layer, and increased pericryptal region. Aberrant crypt foci in rodents are observed as early as 2 weeks and for at least 9 months after a single dose of carcinogen, have a distribution that parallels that of tumors, and have an increased number of aberrant crypts per focus with time after the carcinogen dose. The ability to quantify these lesions in the entire colon of rodents in less than an hour suggests that aberrant crypts may provide a highly efficient in vivo bioassay for colon carcinogens. Since aberrant crypt foci appear to be the earliest identifiable putative precursors of colon cancer, they represent lesions that can be characterized further for the earliest genetic and biochemical alterations. In F344 rats, we have demonstrated that aberrant crypts have multiple histochemically-detectable enzyme alterations. Using similar techniques, we were the first to demonstrate aberrant crypts in unsectioned human mucosa. After embedding and sectioning, these microscopic aberrant crypts resemble rare lesions described earlier in the literature after extensive serial sectioning. In rats and humans, aberrant crypts may be histologically normal or display varying degrees of dysplasia and histochemically-detectable altered enzyme activities. These putative, preneoplastic lesions should reveal early changes that precede colon cancer and ways to alter their progression.

5'-Nucleotidase↗
PubMed1992

Aberrant crypts correlate with tumor incidence in F344 rats treated with azoxymethane and phytate.

Aberrant crypts are putative preneoplastic lesions that have been proposed as intermediate biomarkers for colon cancer. The goals of these studies were to determine (i) if the colon cancer chemopreventive agent, sodium phytate, when started 1 week after a single dose of carcinogen, has any effect on the development of aberrant crypt foci (ACF) in treated rats; and (ii) if ACF at an early time period under these conditions correlate with the later formation of tumors in similarly treated animals. The number of ACF with four or more crypts was greater (P = 0.02, Mann-Whitney test) in rats with tumors compared with rats without tumors killed at 36 weeks after the injection of azoxymethane (AOM); the total number of ACF was not significantly different in these two groups. The incidence of tumors in F344 rats treated with AOM without phytate was 83% (10/12) compared to 25% (3/12) in rats treated with AOM plus phytate (P = 0.0045, two-tail Fisher's exact test). The finding of more (P = 0.005, Mann-Whitney test) ACF with four or more crypts in rats without phytate than in rats with phytate at 12 weeks after the injection of AOM is consistent with the hypothesis that the development of larger ACF (with four or more crypts) is predictive of the tumor incidence. These results validate the use of this parameter, i.e. ACF with four or more crypts, as an intermediate biomarker for tumor incidence in this system.

Animals↗
PubMed1992

High-resolution analyses of two different classes of tumor cells in situ tagged with alternative histochemical marker genes.

To evaluate interactions of two different tumor cell classes during the establishment of micrometastases at the single-cell level, two different BALB/c 3T3 tumor cell derivatives were established that harbor different histochemical marker genes: bacterial lacZ in a EJ-Harvey ras transformant (abbreviated LZEJ cells) and human placental alkaline phosphatase (ALP) gene in a human c-sis transformant (APSI cells). Several different histochemical staining methods were evaluated, using the distinctiveness of lacZ and ALP gene activities, for identification of these cell classes singly or together in the lung after their intravenous injection into nude mice. LZEJ and APSI cells could readily be distinguished from each other after co-injection by using specific and sequential staining protocols of whole organs or sections; staining of host organ cells was minimized. Co-injection of the two tumor cell classes resulted in similar numbers of homogeneous microfoci in lungs of LZEJ or APSI cells within minutes after injection that persisted for several hours before clearance of most of them. Furthermore, a significant percentage of foci could be identified containing both classes of tumor cells on whole-organ or section evaluations; these cohabiting foci resisted clearance from lungs. Therefore, use of two different histochemical marker genes to tag different classes of tumor cells provides a powerful approach for determining their in situ co-localization, cooperation, or interference with the establishment and development of micrometastases, as well as an opportunity to evaluate gene regulation in situ at the single-cell level.

Alkaline Phosphatase↗
PubMed1991

Tissue concentrations of prostate-specific antigen in prostatic carcinoma and benign prostatic hyperplasia.

Prostate-specific antigen (PSA), as measured in peripheral blood, is currently the most widely used marker for the assessment of tumor burden in the longitudinal study of patients with carcinoma of the prostate (PCA). Studies from other laboratories have led to the conclusion that a given volume of PCA causes a much higher level of PSA in the peripheral circulation of patients than a similar volume of prostate without carcinoma. We have evaluated PSA in the resected tissues immunohistochemically and in extracts of PCA and of prostates resected because of benign prostatic hyperplasia (BPH) with an enzyme-linked immunosorbent assay. Immunohistochemical results were less quantitative than but consistent with the results of the ELISA of tissue extracts. Immunohistochemically, there was considerable heterogeneity in the expression of PSA by both PCA and BPH both within and among prostatic tissues from different patients. While the levels of expression of PSA in these tissues overlap broadly, PSA is expressed at a lower level in PCA than in BPH when PSA is expressed as a function of wet weight of tissue (p = 0.0095), wet weight of tissue/% epithelium (p less than 0.0001), protein extracted from the tissue (p = 0.0039), or protein extracted/% epithelium (p less than 0.0001).

Adult↗
PubMed1991

Differences in the leucine aminopeptidase activity in extracts from human prostatic carcinoma and benign prostatic hyperplasia.

Extracts of tissue showed that prostatic carcinomas contain less leucine aminopeptidase activity than benign prostatic hyperplasia. This is true when activity is expressed as specific activity (P = 0.0033), specific activity/% epithelium (P = 0.0007), activity/wet weight of tissue (P = 0.0028), or activity/wet weight of tissue/% epithelium (P = 0.0005). Almost all histochemically demonstrable activity is located in the epithelium. Enzymatic activities in extracts and in histochemical preparations showed similar differences between carcinoma and benign prostatic hyperplasia and were related (R = -0.38, P = 0.0400) to Gleason's grades. The transurethrally resected prostate cancers studied contained no well-differentiated tumors and a high proportion of poorly differentiated tumors. Histochemical activity is absent in most prostatic carcinomas and decreased in others. This observation is particularly interesting in view of the growing knowledge of tumor suppressor genes.

Aged↗
PubMed1991

Transplantation of human prostatic carcinoma into nude mice in Matrigel.

Previous successful transplantation of human primary prostatic carcinomas into nude mice has been described as "close to zero." When injected in Matrigel instead of culture medium, 25,000-fold fewer cells of the PC-3 human prostatic carcinoma cell line were required for the growth of tumors in nude mice during a 3-month period of observation; similar enhancement was observed with two other human prostatic carcinoma cell lines. Six of ten primary human prostatic carcinomas were transplanted successfully into nude mice when Matrigel was used as the vehicle.

Animals↗
PubMed1991

Aberrant crypts: putative preneoplastic foci in human colonic mucosa.

Aberrant crypts were identified for the first time in whole-mount preparations of normal-appearing human colonic mucosa after staining with methylene blue. The foci of aberrant crypts varied from single altered glands to plaques of greater than 30 crypts. The mean proportion of colonic mucosa altered and the number of foci with aberrant crypts per cm2 of colonic mucosa were (a) higher in patients with colon cancer than in patients without colon cancer or predisposing conditions and (b) highest in our single case of Gardner's syndrome. Aberrant crypts are postulated to be the earliest identifiable potential precursors of colon cancer. Analysis of aberrant crypts may facilitate the study of the early pathological and molecular changes that precede colon cancer.

Adult↗
PubMed1990

Recovery of nuclei from glycol-methacrylate-embedded tissue.

The analysis of antigens, enzyme histochemical markers, and DNA has become an important part of the classification of some leukemias, lymphomas, and other neoplastic diseases. Many of the relevant antigens and most of the relevant enzyme histochemical activities are destroyed and others are less than optimally preserved in tissues embedded in hot paraffin. Most enzymatic activities and antigens are well preserved in tissues embedded at 4 degrees C in glycol methacrylate (GMA). The measurement of DNA content in neoplastic cells with the most commonly employed techniques depended on the availability of fresh suspensions of cells until the development by Hedley of methods that permit the recovery of nuclei from paraffin blocks for this purpose. In order to facilitate the analysis of antigens, enzymatic markers, and DNA from the same sample of tissue, we have developed a means of recovery of nuclei from GMA-embedded tissues. Twenty-microns-thick sections of GMA-embedded tonsil were either pretreated with an organic solvent (absolute ethanol or 2-ethoxyethanol) followed by rehydration in phosphate buffered saline (PBS) or directly rehydrated in PBS. The suspensions were formed mechanically by gentle sonication. The type of fixative and length of PBS rehydration were varied. Tissue fixed in 100% acetone, embedded in GMA, and rehydrated directly in PBS for six days gave the highest average yield of nuclei, 3.7 x 10(7) nuclei per gram tissue. In order to assess DNA binding of fluorescent dyes, 2-microns-thick GMA sections were stained with chromomycin, Hoechst 33342 (Sigma Chemical, St Louis, MO), and propidium iodide. Hoechst 33342 bound specifically to the nuclei with low background staining.

Benzimidazoles↗
PubMed1990

Development of micrometastases: earliest events detected with bacterial lacZ gene-tagged tumor cells.

For the study of micrometastases at their earliest stages, we transfected the lacZ gene, which codes for beta-D-galactosidase in Escherichia coli, into BALB/c 3T3 cells transformed by the Ha-ras oncogene (also known as HRAS1) of a human EJ bladder carcinoma. These cells were subsequently injected into 6-week-old, female athymic NCR-NU nude mice by several routes. With chromogenic detection of the product of the lacZ gene (a heterologous gene not observed in animal cells) by use of 5-bromo-4-chloro-3-indoyl-beta-D-galactopyranoside, we easily identified tumor cells implanted in the lungs minutes after intravenous injection by the intensely blue staining of the cells harboring the lacZ gene. The number of lung-associated tumor cells remained constant for several hours after intravenous injection but then decreased to a stable level by 24 hours. At most sites of lung invasion, multiple tumor cells, rather than single cells, were identified; this finding suggests that cooperation among multiple cells may be important in the early stages of micrometastasis development. Within several days, a few foci of micrometastases were expanding by proliferation and/or migration of individual tumor cells among host lung cells. These results confirm that the lacZ gene is an ultrasensitive histochemical marker for analyzing both qualitatively and quantitatively the earliest stages of micrometastasis development in the lung and in other organs where micrometastases may ensue.

Animals↗
PubMed1990

Bacterial lacZ gene as a highly sensitive marker to detect micrometastasis formation during tumor progression.

During tumor progression, micrometastases at their earliest stages have been difficult to analyze qualitatively or quantitatively because of a lack of suitably sensitive markers to discriminate small numbers of tumor cells from normal tissue cell populations. To overcome this problem, the Escherichia coli beta-galactosidase (lacZ) gene was introduced into human EJ Ha-ras oncogene-transfected BALB/c 3T3 cells with subsequent injection of transfected cells into athymic nude mice. Using a chromogenic substrate (5-bromo-4-chloro-3-indoyl-beta-D-galactopyranoside), the lacZ-bearing tumor cells at primary tumor sites as well as at secondary organs stain intensely blue and can be easily distinguished from the host tissue cells hours, days, or weeks postinjection. Staining of lacZ-bearing tumor cells is specific and extremely sensitive in detecting micrometastatic foci in lungs and other organs, including brain and kidney for the first time. Stable integration of the lacZ and ras genes into cultured cells and subsequent tumor cells was verified by Southern blot analyses. The lacZ gene appears to be a stable marker during tumor progression in vivo based both on phenotypic (5-bromo-4-chloro-3-indoyl-beta-D-galactopyranoside staining) and on genotypic (Southern blot analysis) evidence. Furthermore, 5-bromo-4-chloro-3-indoyl-beta-D-galactopyranoside staining of tumor cells can also be used together with alkaline phosphatase staining relatively specific for endothelial cells to relate the topographies of metastatic cells and host blood vessels in embedded sections. By using the lacZ gene as a sensitive quantitative marker, analyses of micrometastasis development in the lung indicate that the ras oncogene contributes to the metastatic phenotype in this EJ Ha-ras model system, although further genetic and/or phenotypic alterations appear to be necessary for long-term growth and development into overt metastases. These findings demonstrate the effectiveness and sensitivity of the bacterial lacZ gene as a phenotypic marker in tumor progression studies, providing both a qualitative and a quantitative tool in virtually any tumor system for examining micrometastasis formation in target organs and the relationship of tumor cells to host organ microenvironments.

Animals↗
PubMed1990

Enzymatic activities in extracts of small (20 +/- 5 mg) samples of prostatic carcinoma.

Previously, we reported that the activities of several enzymes extracted from prostatic cancers were closely related to the histologic grades (Gleason's grades) of the cancers. The survival of patients was more closely correlated with certain enzymatic activities than with Gleason's grades. Unfortunately, because our assays required relatively large amounts of tissue, the increasing proportion of patients who have needle biopsies as their only surgical procedure had to be excluded from our previous study. We now report a method that permits multiple enzyme assays to be carried out with much smaller amounts of tissue. This method will enable us to avoid the selection inherent in the exclusion of patients who have only needle biopsies and should permit the prospective study of most patients with prostatic carcinoma.

Arginase↗
PubMed1989

Correlation of stromal cells by morphometric analysis with metastatic behavior of human colonic carcinoma.

The cellular compositions of the inflammatory infiltrates in human colonic carcinoma in well-defined compartments have not been quantified previously. Morphometric analysis of this tissue revealed several relationships between the concentrations of different kinds of cells that may be an important first approach to understanding the nature of the host reaction in different groups of patients. Primary tumors without metastases differ from those with metastases in that those without metastases contain higher concentrations of plasma cells (P = 0.0019) and eosinophils (P = 0.0098) in sections taken at a location remote from the margin (tissue located greater than 1 cm from the margin) and increased concentrations of eosinophils (P = 0.0224) in sections of tumor contiguous to the margin (tissue 0-4.5 mm from the margin). In sections contiguous to the margin, the concentration of plasma cells is related to the concentrations of lymphocytes (R = 0.55, P = 0.0014), eosinophils (R = 0.46, P = 0.0085), fibroblasts (R = 0.47, P = 0.0075), and neutrophils (R = 0.63, P = 0.0001). In sections remote from the margin, the concentration of plasma cells is related to the concentration of lymphocytes (R = 0.36, P = 0.0442), eosinophils (R = 0.36, P = 0.0457), mast cells (R = 0.38, P = 0.0375), and neutrophils (R = 0.38, P = 0.0371).

Colon↗
PubMed1989

Beta-glucuronidase activity in prostatic carcinoma and benign prostatic hyperplasia.

Beta-glucuronidase activity was extracted from prostatic carcinomas and hyperplastic prostates. It was 3.6-fold higher (P less than 0.0001) in prostatic carcinoma than in prostatic hyperplasia. Analysis for isoenzymes of prostatic beta-glucuronidase by disc-gel electrophoresis in nondenaturing polyacrylamide gels demonstrated only one form of the enzyme. Histochemically, enzymatic activity was observed almost entirely in epithelial cells in both prostatic carcinoma and benign prostatic hyperplasia.

Aged↗
PubMed1989

5'-nucleotidase activity in prostatic carcinoma and benign prostatic hyperplasia.

In light of previous reports of alterations in 5'-nucleotidase activity in neoplastic conditions, 5'-nucleotidase activity was examined histochemically in tissue sections and quantified biochemically in extracts of human hyperplastic prostates and prostatic carcinomas obtained surgically. The 5'-nucleotidase activities per mg protein in extracts of 29 prostatic carcinomas were lower (P less than 0.0005) than in extracts from 10 samples of benign prostatic hyperplasia. The 5'-nucleotidase activity per mg protein in extracts of prostatic carcinoma from the 29 patients correlated (R = -0.369, P = 0.049) with the degree of histological differentiation; the extracts of poorly differentiated carcinomas contained low levels of 5'-nucleotidase. When age and histological differentiation (Gleason's grade) were adjusted, the enzyme activity per mg protein correlated (R = 0.242, P = 0.004) with patient survival. When all three parameters were considered together, i.e., histological grade of the tumor, 5'-nucleotidase extracted from the tumor, and age of the patient, they were found to be mutually complementary for the prediction of patient survival (R = 0.388, P = 0.0001). To our knowledge, this is the first report that prostatic epithelium expresses 5'-nucleotidase; further work will be required to define the reasons for the high levels of activity observed in prostates without cancer and for the decrease in the activity in prostatic carcinoma.

5'-Nucleotidase↗
PubMed1989

Enzyme histochemistry of monocytes/macrophages: a study in a murine model of immune complex-mediated glomerulonephritis.

In a model of immune complex glomerulonephritis in BALB/c mice, cells of monocyte/macrophage lineage (M phi), identifiable by electron microscopy, infiltrate the glomerulus. In spite of this, no unequivocal nonspecific esterase activity can be demonstrated histochemically in the glomeruli. On the other hand, many mononuclear cells with strong acid phosphatase activity are consistently present. This observation is in line with other studies that have demonstrated the heterogeneity of enzyme profiles in different M phi populations. Despite the wide acceptance of nonspecific esterases as markers for M phi, the present study indicates that exclusive reliance on a single marker in investigating the participation of M phi in a pathological process can lead to erroneous conclusions. Use of multiple markers and preferably multiple investigative modalities is recommended.

Animals↗
PubMed1988

Velocity sedimentation of cells.

Velocity sedimentation provides a variety of methods for the separation of cells according to their rates of sedimentation. Many approaches are available.

Cell Fractionation↗
PubMed1987

Enzymatic histochemistry of mouse kidney in plastic.

Two-micrometer sections of methacrylate-embedded kidney were used to investigate the enzymatic activities of mouse kidney where the proximal tubule and Bowman's capsule from the same corpuscle were viewed in the same section. Alkaline phosphatase, acid phosphatase, 5'-nucleotidase, gamma-glutamyl transpeptidase, N-acetyl-beta-glucosaminidase, leucine aminopeptidase, alpha-naphthyl butyrate esterase, and adenosine triphosphatase activities were observed in the proximal tubule, but only 5'-nucleotidase, alpha-naphthyl butyrate esterase, and alkaline phosphatase were observed in the squamous portion of the parietal epithelium of Bowman's capsule. The use of methacrylate-embedded tissue allowed more precise localization of enzymatic activity than is possible with most frozen sections. This may provide interesting applications not only for characterization of kidney diseases but also for characterization of other normal and abnormal tissues.

5'-Nucleotidase↗