51 research outputs found
Imaging of RNA in situ hybridization by atomic force microscopy
In this study we investigated the possibility of imaging internal cellular molecules after cytochemical detection with atomic force microscopy (AFM). To this end, rat 9G and HeLa cells were hybridized with haptenized probes for 28S ribosomal RNA, human elongation factor mRNA and cytomegalovirus immediate early antigen mRNA. The haptenized hybrids were subsequently detected with a peroxidase-labelled antibody and visualized with 3,3'-diaminobenzidine (DAB). The influence of various scanning conditions on cell morphology and visibility of the signal was investigated. In order to determine the influence of ethanol dehydration on cellular structure and visibility of the DAB precipitate, cells were kept in phosphate-buffered saline (PBS) and scanned under fluid after DAB development or dehydrated and subsequently scanned dry or submerged in PBS. Direct information on the increase in height of cellular structures because of internally precipitated DAB and the height of mock-hybridized cells was available. Results show that internal DAB precipitate can be detected by AFM, with the highest sensitivity in the case of dry cells. Although a relatively large amount of DAB had to be precipitated inside the cell before it was visible by AFM, the resolution of AFM for imaging of RNA–in situ hybridization signals was slightly better than that of conventional optical microscopy. Furthermore, it is concluded that dehydration of the cells has irreversible effects on cellular structure. Therefore, scanning under fluid of previously dehydrated samples cannot be considered as a good representation of the situation before dehydration.\ud
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Comparative genomic hybridization of cancer of the gastroesophageal junction: deletion of 14Q31-32.1 discriminates between esophageal (Barrett's) and gastric cardia adenocarcinomas
Incidence rates have risen rapidly for esophageal and gastric cardia
adenocarcinomas. These cancers, arising at and around the gastroesophageal
junction (GEJ), share a poor prognosis. In contrast, there is no consensus
with respect to clinical staging resulting in possible adverse effects on
treatment and survival. The goal of this study was to provide more insight
into the genetic changes underlying esophageal and gastric cardia
adenocarcinomas. We have used comparative genomic hybridization for a
genetic analysis of 28 adenocarcinomas of the GEJ. Eleven tumors were
localized in the distal esophagus and related to Barrett's esophagus, and
10 tumors were situated in the gastric cardia. The remaining seven tumors
were located at the junction and could not be classified as either
Barrett-related, or gastric cardia. We found alterations in all 28
neoplasms. Gains and losses were distinguished in comparable numbers.
Frequent loss (> or = 25% of all tumors) was detected, in decreasing order
of frequency, on 4pq (54%), 14q (46%), 18q (43%), 5q (36%), 16q (36%), 9p
(29%), 17p (29%), and 21q (29%). Frequent gain (> or = 25% of all tumors)
was observed, in decreasing order of frequency, on 20pq (86%), 8q (79%),
7p (61%), 13q (46%), 12q (39%), 15q (39%), 1q (36%), 3q (32%), 5p (32%),
6p (32%), 19q (32%), Xpq (32%), 17q (29%), and 18p (25%). Nearly all
patients were male, and loss of chromosome Y was frequently noted (64%).
Recurrent high-level amplifications (> 10% of all tumors) were seen at
8q23-24.1, 15q25, 17q12-21, and 19q13.1. Minimal overlapping regions could
be determined at multiple locations (candidate genes are in parentheses):
minimal regions of overlap for deletions were assigned to 3p14 (FHIT,
RCA1), 5q14-21 (APC, MCC), 9p21 (MTS1/CDKN2), 14q31-32.1 (TSHR), 16q23,
18q21 (DCC, P15) and 21q21. Minimal overlapping amplified sites could be
seen at 5p14 (MLVI2), 6p12-21.1 (NRASL3), 7p12 (EGFR), 8q23-24.1 (MYC),
12q21.1, 15q25 (IGF1R), 17q12-21 (ERBB2/HER2-neu), 19q13.1 (TGFB1, BCL3,
AKT2), 20p12 (PCNA), 20q12-13 (MYBL2, PTPN1), and Xq25. The distribution
of the imbalances revealed similar genetic patterns in the three GEJ tumor
groups. However, loss of 14q31-32.1 occurred significantly more frequent
in Barrett-related adenocarcinomas of the distal esophagus, than in
gastric cardia cancers (P = 0.02). The unclassified, "pure junction" group
displayed an intermediate position, suggesting that these may be in part
gastric cardia tumors, whereas the others may be related to
(short-segment) Barrett's esophagus. In conclusion, this study has, fist,
provided a detailed comparative genomic hybridization-map of GEJ
adenocarcinomas documenting new genetic changes, as well as candidate
genes involved. Second, genetic divergence was revealed in this poorly
understood group of cancers
Immune reactivity in relationto cytomegalovirusinfection after allogeneic bone marrow transplantation
Contains fulltext :
4445.pdf (publisher's version ) (Open Access
Do senescence markers correlate in vitro and in situ within individual human donors?
Molecular Epidemiolog
Molecular cytogenetic analysis of prostatic adenocarcinomas from screening studies : early cancers may contain aggressive genetic features
No objective parameters have been found so far that can predict the
biological behavior of early stages of prostatic cancer, which are
encountered frequently nowadays due to surveillance and screening
programs. We have applied comparative genomic hybridization to routinely
processed, paraffin-embedded radical prostatectomy specimens derived from
patients who participated in the European Randomized Study of Screening
for Prostate Cancer. We defined a panel consisting of 36 early cancer
specimens: 13 small (total tumor volume (Tv) < 0.5 ml) carcinomas and 23
intermediate (Tv between 0.5-1.0 ml) tumors. These samples were compared
with a set of 16 locally advanced, large (Tv > 2.0 ml) tumor samples, not
derived from the European Randomized Study of Screening for Prostate
Cancer. Chromosome arms that frequently (ie, > or = 15%) showed loss in
the small tumors included 13q (31%), 6q (23%), and Y (15%), whereas
frequent (ie, > or = 15%) gain was seen of 20q (15%). In the intermediate
cancers, loss was detected of 8p (35%), 16q (30%), 5q (26%), Y (22%), 6q,
and 18q (both 17%). No consistent gains were found i
Nonmuscular involvement in merosin-negative congenital muscular dystrophy.
Item does not contain fulltextThe spectrum of nonmuscular involvement in six children with merosin-negative congenital muscular dystrophy is described. In all children, biochemical, neuroradiologic, cardiac, and neurophysiologic studies were performed. Cerebral structures that were myelinated at gestation, including internal capsule, corpus callosum, brainstem, and cerebellar white matter, demonstrated no abnormalities, whereas the periventricular and subcortical white matter, which were myelinated in the first postnatal year, demonstrated signs of leukoencephalopathy. Cerebrospinal fluid analysis revealed an elevated albumin cerebrospinal fluid to serum ratio in the younger children. Electroencephalogram results were abnormal in the two elder children. One child suffered from congestive cardiomyopathy. The increase in nerve conduction velocity in these children over the years lagged behind those of healthy patients, pointing to a demyelinating neuropathy. We conclude that in merosin-negative congenital muscular dystrophy patients, nonmuscular involvement includes the central and peripheral nervous system and the heart. The pattern of myelination of the brain and nerve conduction slowing suggests a myelination arrest. Merosin deficiency can give rise to a congestive cardiomyopathy, which is of no clinical relevance in the majority of children
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