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Biomedical subjects

C E Lincoln

Publications and source records attributed to C E Lincoln.

13 recordsLinked to original sources
PubMed1996

Wild-type and mutant retinoblastoma protein in paraffin sections.

Inactivation of the retinoblastoma susceptibility (RB) gene plays a role in the pathogenesis of a variety of human malignancies. Recently, it has become feasible to study RB expression in archival tissues, and it is expected that immunohistochemical studies on routinely processed tumors will further elucidate the biologic and clinical significance of RB mutations. Our study was designed to address two issues that are critical for the interpretation of such studies, i.e., whether mutant RB protein (pRB) can reliably be distinguished from normal pRB and whether there are significant differences in the performance characteristics of various anti-RB antibodies. We studied cell blocks of 26 mutant RB cell lines (11 lines without any RB expression, nine lines expressing truncated mRNA/pRB, six lines carrying missense mutations) with five different anti-RB monoclonal antibodies, using a recently described procedure that includes an antigen retrieval step. The specific staining pattern for pRB was nuclear. Cytoplasmic staining was found to be nonspecific and could be strong. Some truncated and all full-length mutant pRBs localized to the nucleus, creating positive nuclear staining that might be indistinguishable from the staining pattern of cells carrying wild-type RB. The five antibodies tested showed significant differences in sensitivity, specificity, and background reactivity. Our data suggest that a significant subset of mutant pRB has preserved nuclear translocation capacity, that not all anti-RB antibodies are equally suitable for immunohistochemical analysis of RB expression, and that any such analysis is bound to include a certain, albeit probably small, number of positive stains, despite the absence of functional pRB.

Humans↗
PubMed1995

Immunohistochemical analysis of the p16INK4 cyclin-dependent kinase inhibitor in malignant mesothelioma.

BACKGROUND: The identification in 1994 of the CDKN2 gene as a target for mutations in a wide range of human cancers, including malignant mesothelioma, has been controversial because subsequent studies have detected a lower frequency of CDKN2 gene mutations in primary tumors than in cultured cell lines. These reports raised the hypothesis that another gene, distinct from CDKN2, might be the target of the chromosome 9p21 deletions frequently observed in these tumors. PURPOSE: To address whether inactivation of CDKN2 function is an essential event in the etiology of malignant mesothelioma, we examined p16INK4 protein expression in primary thoracic mesotheliomas, in nonmalignant pleural tissues, and in independent mesothelioma cell lines. We also studied the growth rate of tumor cell lines following stable transfection of CDKN2 gene. METHODS: Retinoblastoma (Rb) and p16INK4 protein expression was determined by immunohistochemical analysis from archival paraffin specimens of 12 primary thoracic mesotheliomas and a nonmalignant pleural biopsy specimen. In addition, protein immunoblot analysis for Rb and p16INK4 expression was conducted on 15 independent mesothelioma cell lines, and the ability of a transfected CDKN2 gene to suppress the growth of the mesothelioma cell lines H2373 and H2461 in vitro was examined. RESULTS: We demonstrated abnormal p16INK4 expression in 12 of 12 primary mesothelioma specimens and in 15 of 15 mesothelioma cell lines. All tumor specimens and the tumor cell lines showed expression of wild-type Rb protein. In addition, we have confirmed the ability of a transfected CDKN2 gene to suppress growth of two independent mesothelioma cell lines. CONCLUSIONS: Immunohistochemical analysis of the p16INK4 gene product is feasible in archival biopsy samples. With this analysis, CDKN2 gene inactivation can be determined in tumors that are contaminated with nonmalignant cells. Furthermore, since loss of p16INK4 protein expression can result from both genetic (gene mutations) and epigenetic (abnormal DNA hypermethylation) mechanisms, as we and others have shown recently, examination of protein expression is a highly sensitive method for analyzing the CDKN2 status in large numbers of tumor samples. IMPLICATIONS: This study suggests that inactivation of the CDKN2 gene is an essential step in the etiology of malignant mesotheliomas. Defining the role of the p16INK4:Rb tumor suppressor pathway and its immediate downstream substrates will be an important goal in designing future therapeutic strategies.

Carrier Proteins↗
PubMed1995

Immunohistochemical detection of the cyclin-dependent kinase inhibitor 2/multiple tumor suppressor gene 1 (CDKN2/MTS1) product p16INK4A in archival human solid tumors: correlation with retinoblastoma protein expression.

The retinoblastoma (RB) and cyclin-dependent kinase inhibitor 2/multiple tumor suppressor gene 1 (CDKN2/MTS1) tumor suppressor genes play important roles in the regulation of the cell cycle. The protein products of these two genes, pRB and p16INK4A ("p16"), respectively, inhibit progression from G1 to S phase. Moreover, p16 has been shown to exert its function through inhibition of CDK4-mediated phosphorylation of pRB. Both genes have been found to be mutated or deleted in a wide range of primary human tumors and tumor cell lines. However, the presence of CDKN2/MTS1 containing nonneoplastic elements in every tumor specimen may contribute to the apparent lower deletion detection rate in resected neoplasms compared to cell lines. We have developed an immunohistochemical assay that allows us to assess p16 expression in formalin-fixed, paraffin-embedded tissues. As controls, we used paraffin-embedded pellets of cell lines with well-defined p16 status (four positive and four negative lines), as well as routinely processed nude mouse xenografts of two p16-positive cell lines. p16-negative cells were characterized by the absence of nuclear staining, whereas cytoplasmic staining was variable. In neoplastic and normal tissues, the level of p16 generally appeared to be low. We tested 75 random human malignancies from 4 different anatomic sites for p16 expression and correlated the findings with the immunohistochemical presence or absence of pRB. Twenty % of tumors selectively lacked pRB, while 37% of neoplasms had undetectable p16. In 43% of all carcinomas, both pRB and p16 could be detected. Significant differences existed in the expression of both tumor suppressor genes between carcinomas from different sites. Breast cancers had the highest rate of p16 negativity (13 of 20). Our data show that: (a) immunohistochemistry may be a suitable modality to screen for RB and CDKN2/MTS1 abnormalities in paraffin-embedded tissues; (b) undetectable levels of p16 expression occur at a relatively high frequency; (c) p16 and pRB expression in common human malignancies is not mutually exclusive; (d) loss of function of both tumor suppressor genes appears to be a distinctly uncommon phenomenon; and (e) different types of carcinomas have variable rates of disturbance in the p16/pRB pathway.

Carrier Proteins↗
PubMed1994

Aberrant RB gene expression in routinely processed, archival tumor tissues determined by three different anti-RB antibodies.

The retinoblastoma (RB) susceptibility gene encodes a nuclear phosphoprotein which is likely involved in cell cycle control and cell differentiation. The RB protein is mutated or absent in a variety of human malignancies. Its role as a molecular marker for clinical tumor behavior is under extensive investigation. However, studies on the status of the RB protein in primary or metastatic tumors and their precursor lesions have been slowed by the lack of availability of a sensitive, reliable assay which allows examination of RB expression in selected cell populations within archival tissues. Thus far, meaningful immunohistochemical analysis of RB protein in formalin-fixed, paraffin-embedded specimens has been achieved only with the polyclonal antibody RB-WL-I. We now describe a method which produced excellent staining results in formalin-fixed, routinely processed tissues, using commercially available monoclonal antibodies (MAbs) in conjunction with an antigen-retrieval step. The resulting stains were superior to those on frozen sections and comparable to those obtained with RB-WL-I. Twelve of 51 random invasive bladder cancers (24%) had abnormal expression of the RB gene, as determined by immunohistochemistry. Smaller cohorts of breast, prostate and lung carcinomas had incidences of aberrant RB gene expression ranging from 9% to 24%. Since the staining method was widely applicable to essentially all formalin-fixed, archival tissues, it may expedite studies on the biological and clinical significance of altered RB expression in human neoplasia.

Antibodies, Monoclonal↗
PubMed1990

Seasonal cycles in the blood plasma concentration of FSH, inhibin and testosterone, and testicular size in rams of wild, feral and domesticated breeds of sheep.

Seasonal cycles in testicular activity in rams were monitored in groups of wild (mouflon), feral (Soay) and domesticated breeds of sheep (Shetland, Blackface, Herdwick, Norfolk, Wiltshire, Portland and Merino) living outdoors near Edinburgh (56 degrees N). The changes in the blood plasma concentrations of FSH, inhibin and testosterone, and the diameter of the testis were measured every half calendar month from 1 to 3 years of age. There were significant differences between breeds in the magnitude and timing of the seasonal reproductive cycle. In the mouflon rams, the seasonal changes were very pronounced with a 6-15-fold increase in the plasma concentrations of FSH, inhibin and testosterone from summer to autumn, and a late peak in testicular diameter in October. In the Soay rams and most of the domesticated breeds, the seasonal increase in the reproductive hormones occurred 1-2 months earlier with the peak in testicular size in September or October. In the two southern breeds (Portland and Merino), the early onset of testicular activity was more extreme with the seasonal maximum in August. In cross-bred rams, produced by mating Soay ewes (highly seasonal breed) with Portland or Merino rams (less seasonal breeds), there was a seasonal reproductive cycle that was intermediate compared to that of the parents. A comparison between all 11 breeds showed a significant correlation between the timing of the seasonal cycle in plasma FSH concentration and testicular diameter (time of peak FSH vs testis, r = 0.95). The overall results in the rams are consistent with a primary role of FSH in dictating the seasonal cycle in testicular size and the secretion of inhibin. The earlier seasonal onset in the testicular cycle in the southern breeds of domesticated sheep, and the differences from the wild type, are taken to represent the effects of genetic selection for a longer mating season.

Animals↗
PubMed1982

Pictorial similarity judgments and the organization of visual memory in the rhesus monkey.

The organization of visual memory for pictures was studied in the rhesus monkey. Two monkeys were tested in a same/different task in which sequentially presented pictures were compared to each other in a pair-wise fashion. The resulting confusion matrixes were analyzed using a multidimensional scaling procedure to obtain two- and three-dimensional graphic representations of the stimulus space. In Experiment 1, the monkeys' confusion errors caused pictures of human and rhesus monkey faces to fall in the same region of multidimensional space, which suggested that the monkeys categorized facial stimuli. A similar effect was found for pictures of different types of fruit. Experiment 2 replicated the categorization of faces with a more diverse collection of human and nonhuman primate faces. Experiment 3 explored the fruit category by varying stimulus attributes orthogonally. The results from this experiment showed that both monkeys encoded the pictures in this category by type of fruit (apples or grapes) and color (red or yellow). Taken together, these studies indicate that rhesus monkeys will treat some classes of pictorial stimuli categorically in visual memory.

Animals↗
PubMed1982

Age-related changes in peripheral nerves of the dog. I. A morphologic and morphometric study of single-teased fibers.

Incidence of lesions in single-teased myelinated fibers and the relationship between internodal length and fiber diameter were studied by morphologic and morphometric techniques on common peroneal and ulnar nerves of 28 dogs between three months and 15 years of age that were free of neuromuscular disease. Minimal lesions were seen in nerves of dogs under ten years of age, usually involving less than 3% of the teased fibers; however, in older dogs, the mean incidence of lesions that included axonal degeneration and segmental demyelination and remyelination, increased to 6% and 9% in ulnar and common peroneal nerves, respectively. A positive correlation (p less than 0.0001) between internodal length and fiber diameter was found in both nerves from immature and mature dogs, but slopes of regression lines were reduced in older dogs.

Aging↗
PubMed1982

Age-related changes in peripheral nerves of the dog. II. A morphologic and morphometric study of cross-sectional nerve.

Qualitative histologic studies and quantitative measurements were made on cross-sectional preparations of common peroneal and ulnar nerves of 32 neuromuscular disease-free dogs from birth to 15 years of age, to provide normative data not available previously. Minimal lesions were seen in nerves of dogs from birth to seven years; however, in older dogs, the incidence of axonal degeneration and segmental demyelination and remyelination increased. Total fiber density of both nerves was over 40,000 fibers/mm2 at birth and declined rapidly during the first six to nine months to level off at about 10,000 fibers/mm2 by one year of age. Density of small (less than 5 micrometers) and large (greater than or equal to 5 micrometers) diameter fibers attained adult values by one year of age. The frequency distribution of the myelinated fibers was unimodal at birth and became bimodal between three and six months of age. The peak of the small and large fiber groups occurred at 3 micrometers and 6 micrometers, respectively. Larger diameter fibers (10 micrometers to 12 micrometers) reached adult values between nine months and one year of age. A 2.5 fold increase in mean fiber diameter occurred during the first year of life. There was no statistically significant change in any histometric parameter after maturity (approximately one year of age).

Aging↗
PubMed1982

Observations on normal skeletal muscle of mature dogs: a cytochemical, histochemical, and morphometric study.

Cytochemical, histochemical, and morphometric features of skeletal muscle were evaluated in 140 specimens from five appendicular muscles of 28 dogs which were free of neuromuscular disease. Dogs ranged from six months to 15 years of age. Evidence of degeneration and regeneration, cell reactions and architectural change was found but with an incidence of less than 1% of the total myofiber population in any muscle. The number of fibers with internal nuclei never exceeded 1%. The only changes that appeared to be age-related were fiber size variation associated with the presence of small angular type IIA fibers in muscles of dogs seven years of age and older. Percentages of type I, type IIA, and type IIC fibers remained relatively constant in the various muscles in the age range examined. Type I fibers predominated in medial head of triceps brachii and superficial digital flexor muscles, whereas type IIA fibers were more numerous in biceps femoris and long head of triceps brachii muscles. An approximately equal ratio was found in gastrocranius muscles. In any given muscle, type IIC fibers represented less than 2% of the fibers. Statistically significant differences were found in values for fiber size between groups of dogs weighing more than (mean diameter range = 40 to 50 microns) and less than (mean diameter range = 30 to 40 microns) 15 kilograms.

Animals↗
PubMed1981

Histochemical differentiation of fiber types in neonatal canine skeletal muscle.

Differentiation of fiber types in developing canine skeletal muscle was studied, using morphologic, morphometric, and histochemical techniques. Sample collections were made from 6 muscles from the pectoral and pelvic limbs of 16 healthy pups between 1 day and 12 weeks of age. In newborn pups, 90% to 95% of the fibers in the 6 muscles were classified as undifferentiated or type IIC; the remaining fibers were classified either normal or large-size type I. Large-size type I fibers usually accounted for 2% to 4% of the total population and were considered analogous with the B fiber of Wohlfart. These fibers were larger than all other fiber types and disappeared after pups reached 4 to 5 weeks of age. After 2 to 4 weeks, the number of undifferentiated fibers decreased with the appearance of, and the concomitant numerical increases of, normal size type I and type IIA fibers. The percentages of type I and IIA fibers approached proportions of the adult dog by 12 weeks, at which time a type IIA fiber predominance was present in biceps femoris, lateral head of the gastrocnemius, cranial tibial, and long head of the triceps. Type I fibers predominated in medial head of the triceps and superficial digital flexor after 4 to 5 weeks. The mean fiber diameters of type I and IIA fibers were similar to any given muscle throughout the postnatal development. All fiber types stained uniformly with the oxidative stain nicotinamide adeninedinucleotide-tetrazolium reductase during the first 12 weeks of life, whereas a distinction between type I and II fibers was evident after 3 to 4 weeks with the periodic acid-Schiff stain reaction.

Adenosine Triphosphatases↗