Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Tissue microdissection”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 559 records · Page 31Linked to original sources
PubMed2001

Frequent p16/MTS1 inactivation in early stages of urothelial carcinoma of the bladder is not associated with tumor recurrence.

OBJECTIVE: p16, located at chromosome 9p21, is a negative regulator of G1 cell checkpoint and functions as tumor suppressor gene. Only few data are available on the frequency and clinical relevance of p16 alterations in Ta, T1 transitional cell carcinoma (TCC) of the bladder. We investigated 40 patients with Ta, T1 TCC of the bladder for p16 alterations (mutations, homozygote deletions, allelic loss) or reduced p16 immunoreaction. PATIENTS AND METHODS: DNA was prepared from microdissected tumor tissue from 40 patients with pTa, pT1 TCC of the bladder (pTa: 18 patients; pT1: 22 patients; grade 1: 7 patients; grade 2: 28 patients; grade 3: 5 patients). Mutation screening was performed using polymerase chain reaction (PCR), single-strand conformation polymorphism (SSCP) and direct sequencing at exon 1 and exon 2. Detection of homozygote deletions was performed using multiplex PCR. Immunohistochemistry (IHC) was performed using an anti-human monoclonal antibody (p16, Pharmingen). Allelic loss was detected by PCR using three different microsatellite markers (D9S161, D9S171, D9S319). RESULTS: SSCP and direct sequencing revealed 3 cases of base substitution which turned out to be natural polymorphisms. Homozygote deletions were not detected in any case. p16 IHC revealed reduced p16 expression (<5% positive nuclei) in 10 patients; 30 patients had a positive reaction (> or =5% positive nuclei) and 10 patients a strong positive reaction (> or =50% positive nuclei). Thirteen of 37 informative cases revealed loss of heterozygosity (LOH) with at least one marker. After a median follow-up of 23 months, 15 patients suffered from disease recurrence. Statistical analysis using Kaplan-Meier analysis and the log-rank test did not reveal significant association of recurrence-free interval and detection of LOH (p = 0.34) or p16 IHC (p = 0.9). CONCLUSIONS: We present a comprehensive evaluation of chromosome 9p21 alterations including p16 analysis and clinical follow-up data. Although p16 mutations and homozygote deletions are rarely detectable in Ta, T1 TCC, the reduction of p16 expression and the frequent hemizygote deletions at 9p21 suggest an early involvement of chromosome 9p and p16 in superficial TCC.

Carcinoma, Transitional Cell↗
PubMed1987

Intranephron distribution of epidermal growth factor immunoreactivity in the mouse.

Using a specific and sensitive epidermal growth factor (EGF) radioimmunoassay, the distribution of EGF in mouse kidneys was studied. In five portions (outer cortex, inner cortex, outer medulla I, II, and papilla) of kidney slices, the outer medulla revealed the highest immunoreactive EGF (iEGF) per wet weight of tissue. In microdissected nephron segments, the predominant iEGF per millimeter length was observed in the medullary and cortical thick ascending limbs of Henle's loop compared with other segments. It became more apparent when expressed per milligram protein. Fluorescence microscopy showed prominent staining for EGF in the thick ascending limbs of Henle's loop. The significance of the nephron heterogeneity in EGF contents is discussed.

Animals↗
PubMed2004

Microsatellite instability and MLH1 promoter methylation in human retinoblastoma.

PURPOSE: To investigate the link between microsatellite instability and epigenetic silencing of the MLH1 gene in the human retinoblastoma genome. METHODS: Methylation at the 5' region of MLH1 was studied, along with its protein expression level by using immunohistochemical staining in 51 retinoblastoma tumors and 2 retinoblastoma cell lines. Also assessed was the genomic stability of 26 retinoblastoma DNAs from microdissected tumor tissue and matched normal retina tissue obtained from the same patient by microsatellite instability (MSI) analysis. The National Cancer Institute-designed reference panel, and 85 markers on chromosomes 1, 6, 9, and 13 were used. RESULTS: Hypermethylation of the MLH1 promoter was detected in the WERI-Rb1 cell line and in 34 (67%) of the 51 tumors, but not in cell line Y79 and the other 17 tumors. MLH1 hypermethylation was associated with null MLH1 protein expression (P < 0.0005) and with well-differentiated histology (P < 0.05). MSI at three markers (D2S123, D6S470, and D13S265) was frequently identified among 26 retinoblastoma specimens with matched normal DNA. Among these 26 retinoblastomas, high-frequency MSI (MSI-H) tumors were detected in 19% (5/26) and low-frequency MSI (MSI-L) in another 19% (5/26). The remaining 62% (15/26) were genetically stable (MSS). MSI status (MSS, MSI-L, and MSI-H) was not associated with MLH1 promoter hypermethylation (P = 0.088; Kruskal-Wallis test). CONCLUSIONS: Epigenetic silencing of the DNA repair gene MLH1 by promoter hypermethylation is a frequent event in retinoblastoma. The results showed that somatic genetic changes involving MSI occur in a subset of retinoblastoma and implicated the presence of a defective DNA mismatch repair pathway resulting in MSI in retinoblastoma.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Clonal analysis of palmar fibromatosis: a study whether palmar fibromatosis is a real tumor.

BACKGROUND: Palmar fibromatosis that arises in the palmar soft tissue is characterized by infiltrative growth with a tendency toward local recurrence but does not metastasize. This study investigated the clonality of this process in twelve female patients, each with a single lesion, by examining the pattern of X-chromosome inactivation. METHODS: Hematoxylin and eosin stained sections of formalin-fixed, paraffin-embedded tissues were microdissected by laser capture microdissection to obtain the proliferative spindle cells. Tumor cells were isolated from the sections of rectum adenocarcinoma, and used for positive control. The genomic DNAs was extracted with phenol-chloroform, digested with a methylation-sensitive restriction endonuclease HpaII, and amplified by polymerase chain reaction (PCR), using primers targeted to a highly polymorphic short tandem repeat (STR) of the human androgen receptor gene (HUMARA). RESULTS: Among the twelve samples, three samples failed amplification, one sample showed homozygosity which was not suitable for further analysis, eight samples were successfully amplified, and showed a random X chromosome inactivation pattern, suggesting polyclonality of these lesions. CONCLUSION: The current findings suggest that palmar fibromatosis is a reactive proliferation rather than a clonal neoplasm.

Journal Article↗
PubMed1995

Expression of nuclear hormone receptors in the rat supraoptic nucleus.

To investigate the role of nuclear hormone receptors on neuropeptide gene expression in the hypothalamo-neurohypophyseal system (HNS) of the rat, a survey was made of members of the nuclear hormone receptor superfamily that are expressed in the supraoptic nucleus (SON). A polymerase chain reaction cloning strategy based on homologies in the DNA-binding domain of AGGTCA-binding factors was devised for the identification of receptors in microdissected SON tissue. Cloning of the amplified products led to the identification of five true receptors, thyroid hormone receptor-alpha (THR alpha), retinoic acid receptor-alpha, retinoic acid receptor-gamma, retinoid X receptor-alpha, and retinoid X receptor-gamma, as well as four orphan receptors, apolipoprotein AI regulatory protein (ARP-1), chicken ovalbumin upstream promoter transcription factor I (COUP-TF I), estrogen-related receptor 2, and testis receptor 4 (TR4). Dot-blot screening of amplified gene fragment analysis showed that THR alpha, ARP-1, TR4, and COUP-TF I were the most abundant factors expressed in the SON region, in the order THR alpha > ARP-1 > TR4 approximately COUP-TF I. THR alpha has previously been localized to HNS neurons. In situ hybridization analysis showed that ARP-1, COUP-TF I, and TR4 were not expressed in magnocellular neurons at appreciable levels, but rather in surrounding structures. Furthermore, in lactating female rats there were no significant differences in the composition of the nine identified nuclear hormone receptors in the SON region compared with control animals. From these experiments, it is concluded that there is a multitude of hypothalamically expressed nuclear hormone receptors, but that only THR alpha is expressed at relatively high abundance in HNS neurons. This indicates that the peptide-producing magnocellular neurons of the SON express a specific set of transcription factors of the nuclear hormone receptor family.

Amino Acid Sequence↗
PubMed2001

Molecular profiling of tissue samples using laser capture microdissection.

The management of cancer and other genetically based diseases is far from optimal in even our most advanced medical centers. There is still uncertainty regarding how diseases will progress in certain patients, toxicity that must be tolerated with imprecise treatment regimens and significant potential for treatment failure. As our understanding of the complexity of these diseases has increased, it has become clear that we must move toward precisely tailored approaches to treating each individual patient. To that end, a major goal of current medical research is the rapid identification of the specific molecular alterations in each patient's disease. This will enable the design of optimal diagnosis, prognosis and treatment, significantly improving survival. This review describes one important approach to the genetic analysis of disease--molecular profiling--and the tenets and technologies necessary for its success.

DNA Fingerprinting↗
PubMed2001

In vitro models to study cellular differentiation and function in human prostate cancers.

In Vitro Models to Study Cellular Differentiation and Function in Human Prostate Cancers. To augment the currently available models of human prostate cancer in vitro, we have established extended life-span epithelial cultures from biopsies of well-differentiated prostate cancers. The genetic identity of the target cells was assessed by allelotyping, using microsatellites located on chromosome 8p, and microdissection of tissues and primary cell cultures. Cells with an extended life span (PxE6) were derived by recombinant retrovirus infection to introduce the human papilloma virus E6 gene (epithelial cells). Immunophenotyping of the resultant cell strains confirmed retention of differentiated cell functions, and the genotype of the E6-expressing epithelial cells was stable, while SV40-immortalized cultures were more unstable, leading to tetraploidy. All PxE6 cells eventually senesced, but an immortalized epithelial culture, P4E6, was derived from one of the epithelial cultures. The properties of this cell line, which remains close to diploid, are similar to those of early prostate cancer cells, and it retains expression of many prostate-associated antigens, such as prostate-specific antigen (PSA).

Allelic Imbalance↗
PubMed2000

Laser capture microdissection-generated target sample for high-density oligonucleotide array hybridization.

Current advances in biomolecular technology allow precise genetic fingerprinting of specific cells responsible for the pathogenesis of human diseases. This study demonstrates the feasibility of generating target samples from laser capture microdissection (LCM) tissues suitable for hybridization of high-density oligonucleotide arrays for gene expression profiling. RNA was successfully isolated by LCM from three paired specimens of oral cancer and linearly amplified using T7 RNA polymerase. Evaluation of the cDNA revealed that five of five cellular maintenance transcripts are detected. Biotinylated cRNA was generated and hybridized to the human Test 1 GeneChip probe arrays, which demonstrated that the RNA is of sufficient quality and integrity to warrant further analysis. Subsequent hybridization of the samples to the HuGenFL GeneChip probe arrays revealed that 26.5%-33.0% of the approximately 7000 represented genes are expressed in each of the six samples. These results demonstrate that LCM-generated tissues can generate sufficient quality cRNA for high-density oligonucleotide microarray analysis, an important step in determining comprehensive gene expression profiling using this high-throughput technology.

Biotinylation↗
PubMed2006

Nerve damage in Mycobacterium ulcerans-infected mice: probable cause of painlessness in buruli ulcer.

Buruli ulcer is an emerging chronic painless skin disease found in the tropics and caused by Mycobacterium ulcerans; however, it remains unknown why the large and deep ulcers associated with this disease remain painless. To answer this question, we examined the pathology of BALB/c mice inoculated in the footpads with M. ulcerans African strain 97-107. On days 54 to 70 after inoculation, extensive dermal ulcers, subcutaneous edema, and numerous acid-fast bacilli were noted at the inoculate region. Nerve invasion occurred in the perineurium and extended to the endoneurium, and some nerve bundles were swollen and massively invaded by acid-fast bacilli. However, Schwann cell invasion, a characteristic of leprosy, was not observed. Vacuolar degeneration of myelin-forming Schwann cells was noted in some nerves which may be induced by mycolactone, a toxic lipid produced by M. ulcerans. Polymerase chain reaction analysis of microdissected nerve tissue sections showed positive amplification of M. ulcerans-specific genomic sequences but not of Mycobacterium leprae-specific sequences. Behavioral tests showed decrease of pain until edematous stage, but markedly ulcerated animals showed ordinary response against stimulation. Our study suggests that the painlessness of the disease may be partly due to intraneural invasion of bacilli. Further studies of nerve invasion in clinical samples are urgently needed.

Animals↗
PubMed2004

Proteomic analysis of human breast cancer tissue with laser-capture microdissection and reverse-phase protein microarrays.

Despite recent advances in breast cancer therapy, women with similar types of breast cancers may respond very differently to standard treatments. The emerging field of clinical proteomics has the potential to revolutionize breast cancer therapy. The ultimate goal of clinical proteomics is to characterize information flow through protein cascades for individual patients. After the protein networks have been elucidated, drug therapies may be specially designed for each patient. The following review describes the proteomic technologies of laser-capture microdissection (LCM) and reverse-phase protein arrays (RPPAs). These technologies allow scientists to analyze relative abundances of key cellular signaling proteins from pure cell populations. Cell survival and apoptotic protein pathways are currently being monitored with LCM and RPPAs at the National Institutes of Health, in phase II clinical trials of metastatic breast and ovarian cancers. Ultimately, proteomics will become an integral component of tracking and managing individualized breast cancer therapy.

Breast↗
PubMed2000

Genetic alterations in metastatic renal cell carcinoma detected by comparative genomic hybridization: correlation with clinical and histological data.

In order to optimize the management of patients with renal cell carcinoma (RCC) it is important to define the genetic risk for metastatic disease. In this study we performed comparative genomic hybridization (CGH) on metastatic tumors aiming at the identification of genetic alterations associated with metastatic disease. We analyzed 46 renal tumors along with their metastases, and 15 non-metastatic renal tumors. Tumors were classified pathologically according to the Heidelberg classification of RCC, and staged according to the TNM-system. Standard CGH was performed using microdissected archival tissues and DOP-PCR. The average numbers of chromosomal aberrations per tumor were 3.0, 2.1 and 3.9 in patients without metastasis, in patients who developed metastases after a two-year latency period (late onset of metastatic disease) and in patients who developed metastases within two years after therapy of the primary tumor (early onset of metastatic disease). CGH revealed chromosomal aberrations in 91% of primary metastatic tumors. Deletions or losses of chromosomes 9 (26% vs 6%), 10 (21% vs 6%) and 18 (23% vs 0) and 17 (28% vs 7%) occurred more often in metastatic tumors than in non-metastatic tumors. Furthermore, these aberrations were more common in patients with early metastases. CGH analysis of 40 pairs of primary RCCs and their corresponding metastasis revealed similar aberrations in 70% of cases. In 30%, however, metastases showed additional chromosomal aberrations not detected in the corresponding primary tumors. In conclusion, we identified genetic alterations associated with metastatic disease in RCC which could be useful for predicting prognosis. Genetic changes leading to metastases occurred early in tumorigenesis of metastatic tumors.

Carcinoma, Renal Cell↗
PubMed2000

Update of diagnostic and prognostic markers in cutaneous malignant melanoma.

The biomolecules described in this article generally have been studied as possible diagnostic or clinically prognostic markers in the context of melanoma disease progression as measured by the gold standards of tumor thickness and development of metastasis. Most of the markers showed variations in expression phenotype only during the deeply invasive or metastatic stage of tumor progression and were thus predictive of clinical outcome only for these subgroups of patients. Some of the markers may have utility in identifying patients with deeply invasive primary tumors who are likely to develop metastasis and thus should receive earlier, more aggressive treatments. In addition, some of the markers may identify patients likely to respond better to a new type of therapy (e.g., anti-angiogenic therapy in a patient whose tumor is overexpressing VEGF or immunotherapy for a patient whose tumor is expressing high levels of MART-1). In the future, it will probably be possible to employ new techniques, such as laser-guided microdissection of tissues, to isolate individual melanocytes in order to identify the earliest stage-specific defects that contribute to an aggressive biological behavior. Identifying the subset of patients with superficially invasive melanomas who will develop metastatic disease will continue to provide a challenge.

Biomarkers, Tumor↗
PubMed2001

Patterns of CDKN2A gene loss in sequential oral epithelial dysplasias and carcinomas.

The CDKN2A gene locus encodes two different proteins derived from alternative splicing. p16 (exons 1alpha, 2, and 3) acts as a G1 cell cycle regulator, and p14ARF (exons 1beta, 2, and 3) acts to modulate MDM2-mediated degradation of p53. Inactivation of p16 is a common finding in many cancers; however, there is little data on CDKN2A gene abnormalities in oral precancer. In this longitudinal study, we examined changes in the CDKN2A gene locus in sequential epithelial dysplasias and oral carcinomas from 11 patients. Genomic DNA was extracted from laser-microdissected lesional tissue, and exons 1alpha, 1beta, and 2 were analyzed by duplex PCR. Immunohistochemistry was done to identify p16 and p14ARF protein expression. Two adjacent polymorphic microsatellite markers were used for allelotyping. Homozygous deletion of exon 1alpha was identified in 2 of 17 (12%) precancerous lesions. Loss of either exon 1alpha, exon 2, or both was seen in seven of nine (78%) carcinomas. In five of these carcinomas, there was loss of only exon 1alpha. No case showed deletion of exon 1beta. In 5 of 11 patients, microsatellite markers showed differing patterns of allelic imbalance in the precancerous lesions and the subsequent carcinoma, suggesting a complex genetic pattern of progression from dysplasia to carcinoma. We conclude that during oral carcinogenesis homozygous deletion of exon 1alpha of the CDKN2A gene is common but that deletion of exon 2 and 1beta is less frequent. Moreover, our results suggest that the progression from oral precancer to cancer, in some cases, is more complex genetically than predicted by linear models of carcinogenesis.

Actins↗
PubMed2001

[Role of the pathologist in management of familial cancers].

Pathologists have long been in charge of the morphological analysis of human cancers, and have been used to be at the end of a chain in the diagnostic procedure. With the search for somatic mutations and new prognostic markers, the pathologists have become key-members in oncogenetics. New modalities for tissue fixation, microdissection of tumors, and forwarding of specimens to the molecular biologists are therefore requested. Two examples are presented to illustrate the need for tumor DNA, with the diagnostic utility of a microsatellite testing as a prescreening test in HNPCC tumors, and the search for new prognostic markers in some inherited predispositions to breast cancers.

Acetic Acid↗
PubMed2003

Loss of heterozygosity of chromosome 1q in gastrinomas: occurrence and prognostic significance.

A proportion of gastrinomas demonstrates aggressive growth, and most deaths occur in this group. Little is known about the molecular pathogenesis of growth of this tumor, and there are no predictive factors that are useful in an individual patient. Chromosome 1 (Chr 1) loss of heterozygosity (LOH) is frequent in a number of nonendocrine tumors and in a few endocrine tumors, and its presence can correlate with tumor aggressiveness and survival. In gastrointestinal endocrine tumors including gastrinomas, little data are available on Chr 1 LOH, and the limited results are contradictory. In the present study we determine whether Chr 1 LOH occurs in gastrinomas and is associated with aggressive growth by performing Chr 1 allelotyping with microsatellite markers in microdissected tumor tissue from 27 human gastrinomas and the leukocyte DNA of the patients. Detailed clinical pathological correlations were possible, because tumor growth in all of the patients was prospectively assessed with yearly imaging studies. Twelve gastrinomas (44%) had Chr 1 LOH, and in all of the cases 1q LOH occurred. 1q LOH was associated with aggressive growth (P = 0.0004), presence of liver metastases (P = 0.019), and postoperative development of hepatic metastases (P = 0.017). Eight (75%) of the 12 tumors with 1q LOH had 1q31-32 LOH over a 17.3 cM region, whereas LOH in 6 tumors (50%) occurred at 1q21-23 over a 12.3 cM area. The presence of 1q31-32 LOH and 1q21-23 LOH correlated with aggressive tumor growth (P = 0.0056 and P = 0.0031, respectively), and with postoperative development of liver metastases (P = 0.0114 and P = 0.011, respectively). These data suggest that 1q LOH is not infrequent in gastrinomas and could be a molecular/genetic prognostic factor for aggressive growth that could be useful clinically. The high frequent allelic loss at 1q31-32 as well as 1q21-23, which was associated with tumor aggressive growth, suggests these two regions harbor putative tumor suppressor gene(s) that are important for aggressive growth of this tumor.

Adult↗
PubMed2003

Retinoschisin, a photoreceptor-secreted protein, and its interaction with bipolar and muller cells.

Usually, photoreceptors interact with other retinal cells through the neurotransmitter glutamate. Here we describe a nonsynaptic interaction via a secreted protein, retinoschisin. Using in situ hybridization, we found that from early postnatal life retinoschisin mRNA is present only in the outer retina of the mouse, and with single-cell RT-PCR we demonstrated its localization in rod and cone photoreceptor cells but not in Müller cells. Western blot analyses of proteins from cultured ocular tissues and microdissected outer and inner retinas, as well as from the culture media of these samples, showed that retinoschisin is secreted from the photoreceptor cells. Immunostaining of permeabilized and nonpermeabilized dissociated retinal cells revealed that retinoschisin is mainly inside and outside the photoreceptors, outside bipolar cells, and associated with plasma membranes of Müller cells and inside their distal processes. Because we showed previously that retinoschisin is distributed all over the retina, our current data suggest that after synthesis and secretion by the photoreceptors, retinoschisin reaches the surface of retinal cells and mediates interactions/adhesion between photoreceptor, bipolar, and Müller cells, contributing to the maintenance of the cytoarchitectural integrity of the retina. These interactions may not occur when the gene encoding retinoschisin is mutated, as it occurs in X-linked juvenile retinoschisis, a disease that results in morphological and electrophysiological defects of the retina.

Age Factors↗
PubMed2005

Synchronous endometrial and ovarian carcinomas: analysis of genetic relationship of the tumors.

Simultaneous carcinomas of the endometrium and ovary may represent a diagnostic dilemma and the clinical management of such cases may be problematic. In surgical pathology practice, we classify them either as double primary tumors (DP) or a single primary tumor with metastasis (PM), according to the conventional clinicopathologic criteria. The distinction has important therapeutic and prognostic implications, but it can be sometimes difficult, especially in advanced cases. In this study, in addition to the clinicopatho-logic classification, we assessed tumor cell clonality in 13 cases with synchronous endometrial and ovarian endometrioid adenocarcinomas using PCR-based microsatellite analysis of microdissected archival tissues for loss of heterozygosity (LOH) and microsatellite instability (MSI). All paired endometrial and ovarian tumors demonstrated either MSI or/and LOH except for 1 case, and therefore the microsatellite analysis was informative in 92.3% of the cases. Nine of 26 tumors (34.6%) exhibited MSI-H and 15 of 26 (57.7%) showed LOH. In contrast to 4 DP cases and 9 PM cases classified according to clinicopathologic criteria, microsatellite analysis suggested 10 DP cases and 2 PM cases. The molecular analysis was not informative in 1 case. Thus, analysis of microsatellite abnormality is a helpful adjunct in the assessment of synchronous tumors, especially to differentiate DP from PM cases in advanced tumor cases. Moreover, the combination of conventional clinicopathologic evaluation and molecular analysis is important and helpful in distinction between the two groups of synchronous tumors.

Adenocarcinoma↗