Fluorescence in situ hybridization: a rapid analysis to verify chromosome aberrations.

We applied fluorescence in situ hybridization (FISH) to assess the presence of structural rearrangement and numerical chromosome aberrations in both metaphase chromosome and interphase nuclei. For this purpose, the biotinylated repetitive alpha-satellite DNA probes for chromosome 1, 18 and 8 (pUC1.77, L1.84 and pJM128) were used to identify tetraploid mosaicism, ring chromosome 18 and trisomy 8 mosaicism for pre-, post-natal and tumor diagnosis respectively. Utilizing this approach, we showed the usefulness of FISH for routine clinical cytogenetics in addition to chromosome banding techniques. The chromosome aberrations with unknown or unclear origin, detected by chromosome analysis, could be confirmed accurately and rapidly.

Y chromosome haplotypes reveal prehistorical migrations to the Himalayas.

By using 19 Y chromosome biallelic markers and 3 Y chromosome microsatellite markers, we analyzed the genetic structure of 31 indigenous Sino-Tibetan speaking populations (607 individuals) currently residing in East, Southeast, and South Asia. Our results showed that a T to C mutation at locus M122 is highly prevalent in almost all of the Sino-Tibetan populations, implying a strong genetic affinity among populations in the same language family. Furthermore, the extremely high frequency of H8, a haplotype derived from M122C, in the Sino-Tibetan speaking populations in the Himalayas including Tibet and northeast India indicated a strong bottleneck effect that occurred during a westward and then southward migration of the founding population of Tibeto-Burmans. We, therefore, postulate that the ancient people, who lived in the upper-middle Yellow River basin about 10,000 years ago and developed one of the earliest Neolithic cultures in East Asia, were the ancestors of modern Sino-Tibetan populations.

Diagnosing dengue virus infection in archived autopsy tissues by means of the in situ PCR method: a case report.

BACKGROUND: The pathogenesis of the severe form of dengue virus infection, dengue hemorrhagic fever, is still obscure. A major research objective has been to determine which body organs are being damaged by dengue virus in this form of dengue. Research has been difficult because dengue hemorrhagic fever is sporadic and tends to occur in parts of the world where modern facilities are scarce and fresh or frozen patient materials are not available. However, major hospitals in these areas have accumulated libraries of paraffin-embedded surgical and autopsy tissues over the years. These tissues may have been subjected to less than optimal fixation and storage. Attempts to localize dengue virus using antigen detection in the stored tissue have encountered many difficulties. OBJECTIVE: Since viral nucleic acid may be preserved under circumstances which destroy protein antigens, our objective was to detect dengue viral RNA in situ in histologic sections of tissues from patients dying of dengue hemorrhagic fever in Thailand. STUDY DESIGN: Tissues from an 11-year-old boy who died at Ramathibodi Hospital, Bangkok, Thailand in November, 1987 with the clinical diagnosis of dengue hemorrhagic fever were treated by transcribing the dengue viral RNA to DNA followed by amplification using the polymerase chain reaction with subsequent in situ hybridization in order to visualize the cells infected with dengue virus. RESULTS: Viral RNA was detected in hepatocytes in the mid-zonal region of the liver, as well as scattered macrophages in skin and lymph nodes. CONCLUSION: Dengue virus infection can be detected in paraffin-embedded autopsy tissues which have been stored for five years. The same procedure can be used for diagnosing dengue viral infection and for studying the pathogenesis of dengue hemorrhagic fever.

Detection of BCR/ABL fusion gene in CML: a preliminary report.

The BCR/ABL fusion gene in 31 patients with chronic myeloid leukemia (CML) was detected by RT/PCR. In 18 cases of Ph' positive patients, 13 had BCR 3/ABL II rearrangement, 1 had BCR 2/ABL II rearrangement and 4 had both rearrangements. One case with complex translocation: 46,XY,t(9;9;22), had BCR 3/ABL II rearrangement. In 8 cases of Ph' negative patients, 4 had BCR 3/ABL II rearrangement, 3 had both rearrangements while 1 had no BCR/ABL rearrangement. Interestingly, in 4 patients who had no cytogenetic result, we could observe BCR 3/ABL II rearrangement in 3 cases and both rearrangements in 1 case. The results suggest that this procedure is sensitive and independent of the presence or absence of an identifiable Ph' chromosome.

Identification of dengue virus-infected cells in paraffin-embedded tissue using in situ polymerase chain reaction and DNA hybridization.

Using tissue from suckling mice infected with Dengue virus, the following improved method for detecting Dengue viral RNA in tissue sections was devised. Reverse transcription of the viral RNA to DNA was followed by the in situ polymerase chain reaction to amplify the viral nucleic acid. This was followed by DNA hybridization with an alkaline phosphatase-labelled probe. The enzyme was then reacted with Fast red and counterstained with hematoxylin. Viral nucleic acid was readily demonstrated within glial cells and macrophages in brains of animals infected with two different serotypes of Dengue.

Evolution of complex chromosomal rearrangements in a case of biphenotypic pre-B/myeloid acute leukemia.

A case of acute biphenotypic leukemia, in which blast cells coexpress antigens of both lymphoid and myeloid lineages, was seen in a 24-year-old female. The clonal karyotype was complex but did not include any of the abnormalities previously found in biphenotypic leukemia. Chromosome abnormalities included breakpoints seen in both acute lymphoblastic and acute myeloid leukemias as well as a number of known fragile sites, namely the rare heritable fragile site at 10q24, and the common, aphidicolin-induced sites on chromosome 7.

Human colorectal cancer: high frequency of deletions at chromosome 1p35.

Cytogenetic analyses of human colon cancer cells have revealed a high frequency of chromosome 1p deletions among other chromosomal abnormalities. In order to find out whether these chromosomal alterations are manifestations of loss of genetic material, we surveyed DNA of 62 primary tumors, 7 metastases, and matching peripheral blood cells with a panel of polymorphic DNA probes that detect different loci on chromosome 1p. A portion of the probes was derived from a microclone bank generated by microdissection and microcloning of 1p35----pter DNA. In 42% of the colon carcinomas allelic loss was observed with at least one probe. The deletions were of different sizes but always included a region involving band 1p35, except for two tumors in which allelic loss was detected more proximally. The frequency of 1p deletion in the metastases was higher than in the primary tumors. These data indicate that genetic information related to tumorigenesis is located within or nearby region 1p35 and that deletion of this region occurs later in tumor development. Our results add to the number of genetic changes presumably involved in colon cancerogenesis.

Efficiency of modified medium for direct chromosome preparation of the chorionic villi.

The direct chromosome preparation from the CV is a reliable method for diagnosis of the gross chromosomal anomalies and could be completed in the shortest possible time. However, direct villi preparation does not always provide sufficient or good quality metaphases. The culture medium plays an important role in achieving a good success rate. We report the efficacy of our modified medium and compared it with Chang's. The modified medium showed slightly higher successful karyotyping (68.4%) than the Chang medium (65.4%). This success rate showed no statistically significant difference. The success rate enhanced up to 92.3 per cent using our modified medium when the CV was of good quality. The higher amount of CV did not lead to a higher success rate. Any amount more than 5 mg of the CV was sufficient for direct chromosome preparation.

Induction of sister-chromatid exchange in human blood lymphocytes by aqueous extract of palmyrah (Borassus flabellifer) flour.

pPalmyrah palm (Borassus flabellifer) is widely consumed by people in certain tropical countries. The incidence of human malignant lymphomas, mutagenicity and toxicity in rats and bacteria encouraged us to study the potency of palmyrah crude aqueous extracts in inducing sister-chromatid exchanges (SCEs) in human blood lymphocytes in vitro. The extracts induced SCEs in a dose-related manner in both females and males. These effects apparently showed no consistency between batches. This result may be due to the intrinsic variation of different donors in their response to the induction of SCEs by palmyrah extracts. SCE frequency was proportional to chromosome length and SCEs at the centromeric region showed no difficulty in being scored. Concerning methods of short-term cytogenetic testing for detecting mutagenic and carcinogenic chemicals, we found that the SCE test was not more sensitive than the classic chromosome-breakage test.

Clastogenic effect of aqueous extracts of palmyrah (Borassus flabellifer) flour on human blood lymphocytes.

Palmyrah (Borassus flabellifer) flour is consumed by humans in some Asian and African countries. In view of its toxic and carcinogenic effects, including the induction of malignant lymphomas in rats, we examined the mutagenic effects of aqueous extracts of the flour on human blood lymphocytes in vitro. The extracts were clastogenic, mainly to group A chromosomes, producing chromatid and chromosome gaps, and chromatid and chromosome breaks with some formation of large and small acentric fragments. Dicentric chromosomes and chromatid interchanges were rare. No ring chromosome was detected. These effects were dose-dependent and were consistently produced by crude extracts of different batches of flour, but were less frequent than the clastogenic effects of mitomycin C which was used as a positive control. No mitogenic effect of the extracts alone was observed.