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PubMedDate not supplied

Microdissection study of the myenteric plexus in acardia, ataxia-telangiectasia, cystic fibrosis, extrahepatic biliary atresia, pediatric AIDS and Werdnig-Hoffmann disease.

Microdissection-point count morphometric study of the myenteric (Auerbach) plexus or esophagus, small intestine, and colon was done for infants and children with acardia (2), ataxia-telangiectasia (5), cystic fibrosis of the pancreas (CFP) (25), extrahepatic biliary atresia (EBA) (17), pediatric AIDS (10), and Werdnig-Hoffmann disease (WHD) (8). Values for fractional area of neural tissue in the plane of the plexus were compared to those of control patients in same age range as those in each disease category by t-test. Statistically abnormal values included low values for small intestine and colon in Werdnig-Hoffmann disease, high values for small intestine and colon in biliary atresia, and high value for colon but a low value for small intestine in cystic fibrosis. Values for all three loci were within the normal range for ataxia telangiectasia and pediatric AIDS. The mechanisms of the low value for small and large intestines in WHD, which causes chronic constipation as a result of skeletal muscle weakness, and of the high values for colon in CFP and EBA, both causing malabsorption with bulky stools, are unclear. The value for small intestine in acardia was normal for term but lower than expected for fetal bowel of the same size, possibly because of reduced neural crest inflow to the fetal bowel.

Acquired Immunodeficiency Syndrome↗
PubMed1998

Clonality analysis of cervical cancer on microdissected archival materials by PCR-based X-chromosome inactivation approach.

Clonality of invasive human cervical cancer was assessed in microdissected archival formaldehyde-fixed, paraffin-embedded tissues by PCR-based analysis of X-chromosome inactivation of the androgen receptor gene. In 18 informative cases, cancers from 12 cases including 2 early-invasive cancers were scored as monoclonal. Among them, 8 cases had the combination of random X-chromosome inactivation in normal cervical tissue and non-random in the cancer and in other 4 cases normal and cancer tissue were both non-random but had discordant X-chromosome inactivation. Six cases showed that the non-random inactivation affected the same allele in cancer and normal tissue and thus were considered to be inconclusive. The results suggest the monoclonal origin of cervical cancer and indicate that genetic events are critical in transition of pre-malignant epithelia to invasive cancer in cervical carcinogenesis. Finding of monoclonal early invasive cancer also argues that monoclonality of human tumors is not a late event due to clonal competition or selection. X-chromosome inactivation patterns in normal cervical epithelia and tissues were analyzed in 21 informative cases. When a mixture of normal stroma and epithelium was analyzed, 38% (8/21) of cases showed non-random inactivation pattern, whereas using pure epithelium it was 64% (11/17). The X-chromosome inactivation pattern between mixed epithelium/stroma and matched epithelium differed in 2/8 cases. These results point to a certain amount of overlooked source of error in X-inactivation-based tumor clonality analysis in which peripheral blood monocytes were chosen as control and strongly recommend that control tissues which share close histological origin with analyzed tumors should be selected in any clonality study of human neoplasm. Post-embryological mechanisms of clonal patch in normal cervical epithelia are also discussed.

Carcinoma, Squamous Cell↗
PubMedDate not supplied

Loss of heterozygosity of nucleotide excision repair factors in sporadic oral squamous cell carcinoma using microdissected tissue.

The deficiencies of nucleotide excision repair (NER) factors are involved in rare genetic diseases such as xeroderma pigmentosum (XP) with increased risk of developing cancer on sun-exposed areas of the skin. However, the abnormality of NER factors in human sporadic carcinoma remains unclear. Loss of heterozygosity (LOH) analysis, using the microdissected tissues, for the XPA, XPB, XPC, XPD, XPE, XPF, XPG and the transcription-coupled repair factor, Cockayne syndrome B (CSB) revealed that NER factors were abnormal in 30.0% (3/10 cases) of oral squamous cell carcinomas. Furthermore, 10.0% of oral carcinomas exhibited LOH for NER factors without LOH for tumor suppressor genes such as p53, FHIT, APC, BRCA1, BRCA2 and DCC. These observations raise the possibility that alterations of NER factors may be involved in carcinogenesis in human oral squamous cell carcinoma.

Carcinoma, Squamous Cell↗
PubMed2003

Primary duodenal carcinoma showing divergent growth patterns as determined by microdissection-based mutational genotyping.

Primary duodenal adenocarcinoma accounts for less than 0.5% of all gastrointestinal cancers. We report a case of duodenal adenocarcinoma with highly divergent growth patterns consisting of poorly differentiated adenocarcinoma and neuroendocrine carcinoma proven to arise as a single neoplasm of monoclonal origin, as demonstrated by microdissection-based mutational profiling. Multicomponent growth patterns, as seen in this case, can occasionally be encountered in gastrointestinal malignancies and have led to speculation about the pathogenesis. The methods used to clearly establish monoclonal origin based on the unique profiling of mutational damage can address fundamental issues related to tumor development and progression, while providing cogent clinical information. Our findings confirm the great potential for intestinal epithelial cells to differentiate along different histogenetic lines during tumor progression.

Adenocarcinoma↗
PubMed1999

Chronic inflammatory ear disease and cholesteatoma: creation of auxiliary attic aeration pathways by microdissection.

HYPOTHESIS: The attic compartments, except for Prussak's space, are aerated through the tympanic isthmus. The aim of this study was to develop aeration pathways that would bypass the isthmus in surgery for chronic inflammatory ear disease and cholesteatoma. BACKGROUND: Microdissection of the epitympanum has shown that the anterior attic and the supratubal recess are separated by the tensor fold, the excision of which creates a large new aeration pathway. METHODS: Earlier surgical experience was reexamined as to the access to the tensor fold. Twenty temporal bones were dissected to create clinically useful new surgical routes for tensor fold removal in the presence of an intact ossicular chain. RESULTS: An endaural atticotomy, extended to the supratubal recess, allows excision of the tensor fold; however, the excision must be performed blindly. Cutting the neck of the malleus to allow lateral lifting of the manubrium exposes the tensor tendon and allows rapid excision of the fold. The elasticity of the tendon assists in approximation of the cut edges. In canal wall up surgery, removal of the lateral attic bone until the root of the zygoma exposes the anterior surface of the head of the malleus and the lateral portion of the transverse crest. Drill-out of the crest leads directly to the posterior side of the tensor fold, allowing its excision under direct vision. Thinning of the attic bone lateral to the body and short process of the incus allows simultaneous removal of the lateral incudomalleal fold. CONCLUSIONS: When the ossicular chain is discontinuous, tensor fold resection can be made under direct vision. With an intact chain, cutting of the neck of the malleus used in tympanic glomus tumors causes no hearing changes, allows complete fold excision, and is adaptable to chronic ear surgery. The frontolateral attic route for removal of tensor fold, together with the lateral incudomalleal fold, can be used in the canal wall up surgery to improve attic aeration.

Cholesteatoma, Middle Ear↗
PubMed1999

Chromosome 16 allelic loss analysis of a large set of microdissected prostate carcinomas.

PURPOSE: To perform loss of heterozygosity (LOH) analysis on chromosome 16 in 102 highly purified DNA samples isolated from one or more adenocarcinomas, prostatic intraepithelial neoplasia (PIN), and matched benign prostatic epithelium from 95 radical prostatectomy patients. MATERIALS AND METHODS: Specimens were procured by microdissection of frozen tissue samples, thus ensuring that highly select pure populations of cells were obtained for DNA extraction and LOH analysis. Multiple microsatellite markers were used to determine allelic loss on chromosome 16q. RESULTS: Overall loss on 16q was seen in 31% of the cancers, and occurred more frequently in high stage cancers than low stage cancers. In contrast, allelic loss in PIN failed to exceed 6% at any of the loci that were examined. CONCLUSIONS: These results suggest that inactivation of a putative tumor suppressor gene on 16q may be involved in the progression of some prostate cancers.

Adenocarcinoma↗
PubMed1999

Loss of heterozygosity is detected at chromosomes 1p35-36 (NB), 3p25 (VHL), 16p13 (TSC2/PKD1), and 17p13 (TP53) in microdissected apocrine carcinomas of the breast.

INTRODUCTION: Apocrine carcinomas of the breast are an unusual special category of predominantly AR+, ER-, and PR- breast cancer, characterized by cells with abundant, eosinophilic cytoplasm and nuclei with often prominent nucleoli. To further investigate these lesions, loss of heterozygosity (LOH) was evaluated at multiple chromosomal loci, including loci frequently mutated in breast cancer. MATERIALS AND METHODS: Twenty-five intraductal apocrine carcinomas, 11 invasive apocrine carcinomas, and six apocrine hyperplasias were retrieved from the files of the Armed Forces Institute of Pathology (Washington, DC) and Fairfax Hospital (Fairfax, VA). Cells from lesional as well as normal tissues were microdissected. LOH was performed at a number of chromosomal loci, including loci commonly altered in breast cancer: 1p35-36 (NB), 3p25.5 (VHL), 8p12 (D8S136), 9p21 (p16), 11p13 (D11S904), 11q13 (INT-2 and PYGM), 16p13.3 (TSC2/PKD1 gene region), 17p13 (TP53), 17q13 (NM23), and 22q12 (D22S683). RESULTS: Among informative in situ and invasive apocrine carcinomas, LOH was present in 33% of 15 cases for 17p13 (TP53), as well as 36% of 14 cases for 3p25 (VHL), 30% of 10 cases for 1p35-36 (NB), and 27% of 11 cases for 16p13.3 (TSC2/PKD1). A higher frequency of LOH was noted among invasive apocrine carcinomas (30 to 50%) compared with in situ apocrine carcinomas (23 to 33%) at these loci. LOH was present simultaneously for TP53 and either VHL or NB in five cases. Infrequent (< or =12%) or absent LOH was detected at the remaining loci, including several loci commonly mutated in breast cancer (i.e., INT2, PYGM, and NM23). LOH was not detected in any of the six apocrine hyperplasias. CONCLUSION: An intermediate frequency of allelic loss was detected at multiple tumor suppressor gene loci, including 17p13 (TP53), as well as 1p35-336 (NB), 3p25 (VHL), and 16p13 (PKD1/ TSC2), in apocrine carcinomas of the breast, with a higher overall frequency of LOH noted among invasive tumors compared with in situ tumors. Aside from LOH at p53, LOH was infrequent or absent at several other loci commonly mutated in breast cancer. This preliminary molecular evidence supports immunohistochemical data that apocrine carcinomas of the breast may possess unique mechanisms of carcinogenesis, compared with ordinary ductal carcinomas. However, further study is needed to support this assertion and to determine if the LOH detected is truly etiologic or if it is the result of genetic progression.

Alleles↗
PubMed2000

Microdissection based comparative genomic hybridization analysis (micro-CGH) of secondary acute myelogenous leukemias.

Comparative genomic hybridization (CGH) is a well established technique in molecular cytogenetics. However, leukemias, and especially secondary acute myelogenous leukemias (sAML) are not very well analyzed by this technique, even though such diseases are often characterized by complex karyotypic changes, not resolvable by conventional cytogenetic banding analysis. This lack of CGH-studies might be due to the fact, that in most cases bone marrow aspirate is too limited to do DNA-extraction additionally to the cytogenetic analysis. To circumvent this problem a new CGH technique has been applied to analyze 10 AML cases with complex karyotypic changes. In each case 15 interphase nuclei of the harvested and fixed bone marrow cell-suspension have been microdissected from the coverslip surface and collected in a tube. Subsequently, DNA was amplified by DOP-PCR. With this micro-CGH technique additional cytogenetic information from 10 highly aberrant AML cases was obtained and confirmed by FISH on metaphase of the corresponding AML case.

Aged↗
PubMed2000

Characterization of intracellular prostate-specific antigen from laser capture microdissected benign and malignant prostatic epithelium.

The proportion of unbound serum prostate-specific antigen (PSA; percent-free PSA) is reported to be lower in men with prostate cancer compared to men with benign prostates (U. H. Stenman et al., Cancer Res., 51: 222-226, 1991; H. Lilja et al., Clin. Chem., 37: 1618-1625, 1991; D. L. Woodrum et al., J. Urol., 159: 5-12, 1998; W. J. Catalona et al., J. Am. Med. Assoc., 279: 1542-1547, 1998). The majority of immunoreactive PSA in serum is complexed to alpha-1-antichymotrypsin (ACT). Two major mechanistic questions have previously been unknown: (a) Does PSA in human prostate cancer cells in tissue exist in a free or bound form? and (b) Is PSA produced by malignant cells in the free form because it has lost the ability to form a complex with ACT? Laser capture microdissection (LCM) enables the acquisition of pure populations of defined cell types from tissue (M. R. Emmert-Buck et al., Science, 274: 998-1001, 1996; R. F. Bonner et al., Science, 278: 1481-1483, 1997). This technology provides a unique opportunity to study intracellular protein composition and structure from human cells. In this study, we used LCM to assess the bound versus free form of intracellular PSA in both benign and malignant epithelium procured from prostate tissue. One-dimensional and two-dimensional PAGE were performed on cellular lysates from LCM-procured benign and malignant prostate epithelium from frozen tissue specimens. Western blotting analysis of one-dimensional PAGE gels revealed a strong band at M(r) 30,000 (expected molecular weight of unbound PSA) in all cases demonstrating that the vast majority of intracellular tumor and normal PSA exists within cells in the "free" form. Binding studies showed that PSA recovered from LCM-procured cells retained the full ability to bind ACT, and two-dimensional PAGE Western analysis demonstrated that the PSA/ACT complex was stable under strong reducing conditions. We conclude that intracellular PSA exists in the "free" form and that binding to ACT occurs exclusively outside of the cell.

Blotting, Western↗
PubMed2000

Relationship between cytogenetic aberrations by CGH coupled with tissue microdissection and DNA ploidy by laser scanning cytometry in head and neck squamous cell carcinoma.

BACKGROUND: The relationship between DNA sequence copy number aberrations (DSCNAs) and DNA ploidy in head and neck squamous cell carcinomas (HNSCCs) is still controversial. Materials and Methods We analyzed DSCNAs by comparative genomic hybridization (CGH) combined with microdissection and DNA ploidy by laser scanning cytometry (LSC) in 18 surgically removed HNSCCs and compared the data. RESULTS: Copy number increases were most frequently observed on chromosomes 3q (16 cases), 8q (13 cases), and 12p (11 cases). Copy number decreases were observed on chromosome 3p (14 cases). LSC revealed DNA aneuploidy in 10 of the 18 cases. All DNA aneuploid tumors exhibited gain or amplification of DNA copy number at 12p11-12.1, whereas gain of DNA copy number was found in only 1 of 8 diploid tumors. DSCNAs were more frequent in DNA aneuploid tumors than in diploid tumors (P < 0.005). CONCLUSIONS: The present observations indicate a close relationship between DSCNAs and DNA ploidy in HNSCCs.

Aged↗
PubMedDate not supplied

[Structural organization of nephrons in the kideny and proximal reabsorption in representatives of different vertebrate classes (according to microdissection and micropuncture studies)].

By microdissection method, single nephrons have been isolated from the river lamprey Lampetra fluviatilis, teleost fish Myoxocephalus scorpius (White sea), frog Rana temporaria, lizard Agama caucasica, hen Gallus domesticus and albino rat Rattus norvegicus (Wistar strain). Tubular reabsorptive capacity was measured in kidneys of lampreys and rats by the split oil droplet method. In the animals studied, each of the nephrons consists of the glomerulus, proximal, thin and distal segments. Relative length of the proximal segment does not depend on phylogenetic position of a species. Transtubular isotonic water reabsorption from the proximal tubule in rats is significantly higher than in lampreys. The level of proximal reabsorption is not related to the length of the proximal tubule in the Verterbrates studied but depends on the intensity of tubular transport. The structure which is similar to Henle's loop is present already in Cyclostomes [12]. Further stage of the progressive development of the uriniferous tubules is presented by uneven localization of nephron populations within the kideney in Reptiles. These data suggest that the effective system of osmotic concentration appeared in evolution not as the result of Henle's loop formation, but on the basis of the development of the medullar substance formed mainly by intercortical and juxtamedullar nephron populations.

Absorption↗
PubMed2000

High resolution allelotype of microdissected primary nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is a common cancer in South China but is rare in other parts of the world. To better understand the molecular basis of this cancer, we performed high-resolution allelotyping on 27 microdissected primary tumors using 382 microsatellite markers. We have detected high frequencies of allelic imbalance on 3p (96.3%), 9p (85.2%), 9q (88.9%), 11q (74.1%), 12q (70.4%), 13q (55.6%), 14q (85.2%), and 16q (55.6%). Nonrandom allelic changes of 12q and 16q were revealed for the first time. In addition, loss of heterozygosity on chromosomal arms 1p (37.0%), 5q (44.4%), and 12p (44.4%) were also common in NPC. Multiple minimally deleted regions, 7-40 cM, were identified at 3p14-24.2, 11q21-23, 13q12-14, 13q31-32, 14q24-32, and 16q22-23. Frequent deletions of these minimally deleted regions implied the presence of tumor suppressor genes that may be involved in the development of NPC. Consistent loss of heterozygosity on 3p, 9p, and 14q in almost all tumors suggested that such changes are critical events in NPC tumorigenesis.

Alleles↗
PubMed2000

Genome-wide search for loss of heterozygosity using laser capture microdissected tissue of breast carcinoma: an implication for mutator phenotype and breast cancer pathogenesis.

Breast cancer is considered to display a high degree of intratumor heterogeneity, without any obvious morphological and pathological steps to define sequential evolution, and its progression may vary among individual tumors. In an attempt to elucidate these etiological and phenotypic complexities, the present study, based on the fundamental concept that genomic instability is the engine of both tumor progression and tumor heterogeneity, was conducted to test the hypothesis that breast cancer pathogenesis is driven by double-strand break (DSB)-initiated chromosome instability (CIN). The rationale underlying this hypothesis is derived from the clues provided by family breast cancer syndromes, in which susceptibility genes, including p53, ATM, BRCA1 and BRCA2, are involved within the common functional pathway of DSB-related checkpoint/ repair. Because genomic deletion caused by DSB is reflected in the genetic mechanism of loss of heterozygosity (LOH), this genome-wide LOH study was conducted, using 100 tumors and 400 microsatellite markers. To minimize the effect of heterogeneity within tumors, the experimental technique of laser capture microdissection was used to ensure that genetic and phenotypic examinations were based on the same tumor cells. Support for our hypothesis comes from the observations that: (a) the extent of DSB-initiated CIN in tumors significantly increased as tumors progressed to poorer grades or later stages; (b) in the sequential steps toward CIN, the loci of p53 and ATM, the key checkpoint genes against DSB, were lost at the earliest stage; and (c) many loci identified to be important in breast tumorigenesis were the genomic sites possibly harboring the genes involved in DSB-related checkpoint/repair (including RAD51, RAD52, and BRCA1) or CIN (including FA-A, FA-D, and WRN), and a higher number of these loci showing LOH was significantly associated with increased level of DSB-initiated CIN (P < 0.0001). Breast cancers are thus considered to be sequentially progressive with CIN. However, CIN might also cause genetic heterogeneity, which was revealed by the findings that LOH at some markers was observed only in the component of ductal carcinoma in situ but not in the invasive component of the same tumors. In addition, some markers were found to preferentially lose at specific tumor grades, implying their contribution to genetic heterogeneity during tumor development. Therefore, this study suggests that breast cancer progression is clonal with regard to CIN, but different breast cancers would present distinct molecular profiles resulting from genetic heterogeneity caused by CIN.

Adult↗
PubMed2000

Lung hypoplasia in a patient with del(2)(q33-q35) demonstrated by chromosome microdissection.

We report on a 17-month-old girl with multiple malformations, including lung hypoplasia, multiple ventricular septal defects, craniofacial anomalies, and malrotation of the intestine. Moreover, the patient showed Robin sequence, developmental delay, as well as pre- and postnatal growth retardation. Postnatal cytogenetic analysis revealed an interstitial deletion on the long arm of chromosome 2. Microdissection and reverse chromosome painting of the aberrant chromosome 2 as well as FISH with a panel of chromosome 2q band-specific YACs mapped the deletion to 2q33-q35. Lung hypoplasia has not been described so far in patients with del(2)(q33-q35). A review of previously reported patients showed variable phenotypes apparently due to different deleted chromosomal segments.

Abnormalities, Multiple↗
PubMed1997

[Isolation of the expression fragments with the probe pool of human chromosome 14 q 24.3 generated by microdissection].

The strategy of isolating the band-specific expression fragments from the probe pool of human chromosome generated by microdissection was reported in present paper. A chromosome 14 q 24.3 band-specific single copy DNA library was constructed based on this probe pool. Using this pool DNA as probe to hybridize the human bone marrow cell cDNA library, 68 primary positive clones were selected from 5 x 10(5) cDNA clones. Of them 32 clones were got in second-round screening and designed as cFD 14-1-32. Finally, 24 bandspecific expression fragments were identified from these 32 positive clones by analysing the results of DNA hybridization. Those band-specific clones can hybridize to both 14 q 24.3 DNA and human genomic DNA, but have no hybridization signal with 17 q 11-12 DNA. Partial sequences of 13 fragments of them were sequenced and were identified as novel cDNA sequences as well as have some homology with known genes in NCBI database. Analysis of expression spectrum of cFD 14-1 suggested that the cDNA fragments thus obtained can be used to isolate the genes not yet be cloned in 14 q 24.3 region.

Base Sequence↗
PubMed2000

Presence of genetic alterations in microdissected stroma of human colon and breast cancers.

We show by laser-assisted microdissection that frequent genetic alterations in non-hereditary invasive human colon and breast cancers (loss of heterozygosity and TP53 mutations) occur not only in the neoplastic epithelial cells, but also in the adjacent fibroblastic tumor stroma and that both components can share clonal features. Tumor cell-mesenchyme transitions are among the possible explanations for these findings.

Breast Neoplasms↗
PubMed2000

[Microdissection of chromosomes].

The technique of deciphering different chromosome abnormalities by the employment of specific DNA probes prepared by microdissection of normal or abnormal chromosomes is described. It is emphasised that in some families the diagnosis is possible only by "family specific" probes.

Chromosome Aberrations↗