Deoxyribonucleic Acid Probes Research Articles

Vascular endothelial growth factor gene expression in ovine placenta and fetal membranes

OBJECTIVE: The purpose of this study was to explore the gene expression of vascular endothelial growth factor (VEGF) in placental cotyledon, chorion, and amnion of the ovine fetus. STUDY DESIGN: Time-dated pregnant sheep with singleton or twin fetuses at a gestational age ranging from 100 to 140 days were used for the study. Placental cotyledonary, chorionic, and amniotic tissues were collected and processed for messenger ribonucleic acid analysis by Northern blotting and reverse transcription-polymerase chain reaction. RESULTS: By use of a phosphorus 32-labeled human VEGF complementary deoxyribonucleic acid probe, a prominent VEGF messenger ribonucleic acid transcript of 3.7 kb was detected in the cotyledon, chorion, and amnion. A minor band of 1.7 kb was also found but only in the cotyledon and chorion. The abundance of mesenger ribonucleic acid encoding VEGF was highest ( p < 0.001) in the cotyledon and lowest in the amnion. In these tissues polymerase chain reaction-amplified products corresonding to VEGF 121, VEGF 165, VEGF 189, and VEGF 206 were identified by ethidium bromide. In addition, a polymerase chain reaction fragment corresponding to VEGF 145 was observed. These fragments produced specific hybridization signals with the human VEGF radioactive probe where the intensity of the signal was strongest for VEGF 165 and weakest for VEGF 189. CONCLUSIONS: VEGF gene expression was detected in the cotyledon, chorion, and amnion of the near-term ovine fetus. These findings suggest that vascular endothelial growth factor may play a role in the induction of angiogenesis and promotion of permeability in the microvessels that perfuse the placental and fetal membranes.

Kininogen present in rat reproductive tissues is apparently synthesized by the liver, not by the reproductive system

OBJECTIVE: The purpose of this study was to determine the source(s) of the reproductive tract kininogen and to assess whether kininogen transcription is influenced by reproductive conditions. STUDY DESIGN: Rats in various reproductive states (immature, mature, ovulatory, luteal phase, pregnancy, parturition, postpartum) were used to obtain reproductive tissues (follicles, corpora lutea, oviduct, uterus, testes) and liver. Complementary deoxyribonucleic acid probes for rat prekininogens were used to quantify kininogen messenger ribonucleic acid synthesis. RESULTS: The T-prekininogen complementary deoxyribonucleic acid probe detected a single 1.6 kb message, whereas the k-prekininogen complementary deoxyribonucleic acid probe identified two messages, an abundantly expressed 1.6 kb band and a 2.2 kb band. The source of all the three prekininogen messages appears to be the liver. Naturally occurring reproductive conditions such as ovulation, implantation, and parturition, did not turn on prekininogen message transcription in the rat gonad or genital tract. Only decidualization of the uterus was associated with the induction of kininogen transcription in the liver. CONCLUSION: There appears to be little, if any, contribution of local gene expression to the kininogen present in the reproductive tissues. Apparently, the reproductive tract increases uptake of kininogen from plasma as needed.

Helicobacter pyrlori infection affects manganese- and copper/zinc-superoxide dismutase levels in the gastric mucosa

Oxidative stress may contribute to the progression of Parkinson’s disease, and while the status of antioxidant enzymes is thus important, little data on their regional distribution in basal ganglia exist. We now report on the distribution and levels of messenger ribonucleic acid (m-RNA) for the antioxidant enzymes copper, zinc-superoxide dismutase (Cu,Zn-SOD), manganese-superoxide dismutase (Mn-SOD), and glutathione peroxidase in rat basal ganglia using in situ hybridisation histochemistry with complementary deoxyribonucleic acid probes specific for these enzymes. The m-RNA for Cu,Zn-SOD, Mn-SOD, and glutathione peroxidase was expressed throughout basal ganglia. Levels of m-RNA were significantly higher in substantia nigra pars compacta than in all other regions of basal ganglia for both Cu,Zn-SOD (53–62%, P<0.001) and Mn-SOD (37–45%, P<0.05). Mn-SOD m-RNA levels were also significantly higher in SN pars reticulata than in the nucleus accumbens (10%, P<0.05) and striatum (12%, P<0.01). In contrast, glutathione peroxidase m-RNA levels were only significantly higher in SN pars compacta when compared with SN pars reticulata (23%, P<0.05), and in the striatum when compared with the nucleus accumbens (21%, P<0.05). The data suggest that SN pars compacta may be vulnerable to oxidative stress and thus dependent on the high antioxidant capacity provided by these cytoprotective enzymes. In conclusion, this study demonstrates the relative distribution of antioxidant enzymes in rat basal ganglia and forms the basis for further study in rodent models of Parkinson’s disease.

An increase in expression of the lipocalin 24p3 is found in mouse uterus coincident with birth

OBJECTIVE: To provide novel insights into the molecular events associated with parturition, a differential screen was made of a mouse uterine complementary deoxyribonucleic acid library to identify selectively expressed genes in late pregnancy.STUDY DESIGN: A differential hybridization was used to screen a mouse complementary deoxyribonucleic acid library prepared from late pregnancy uterus. A 1 kb clone was isolated that was subsequently identified as 24p3, a member of the lipocalin family. By use of radiolabeled complementary deoxyribonucleic acid probes prepared from this clone Northern hybridization were conducted against total ribonucleic acid purified from mouse uterus collected during late pregnancy and the first week after birth and a variety of other mouse tissues to determine whether this gene is selectively expressed in the uterus coincident with parturition.RESULTS: Low levels of message for the 24p3 gene could be detected in uterine ribonucleic acid, but there was a massive increase in the level of message for this gene on the days surrounding birth. Northern hybridizations conducted against additional tissues collected from both pregnant and nonpregnant mice did not detect message to a similar degree as found in the uterus.CONCLUSIONS: The uterus constitutes a major site of expression of this gene, particularly near birth. The expression of this gene coincident with birth suggests a potential physiologic role of the neutrophil with the induction of labor.

In situ hybridization: A new technique to determine the origin of fibroblasts in cryopreserved aortic homograft valve explants

Tissue degeneration reduces the durability of cryopreserved homografts. Earlier studies indicated that the presence of fibroblasts in homograft leaflets may contribute to increased valve longevity. These fibroblasts may be of recipient origin or represent surviving donor cells. We developed a method, based on in situ hybridization, to determine the origin of fibroblasts in homograft explants. In young pigs we performed aortic valve replacement with a cryopreserved porcine aortic homograft. A male homograft was implanted in a female pig, whereas two male recipients received a female homograft. After 3 to 4 months the homografts were explanted. Frozen sections were made and alternately examined with hematoxylin-eosin staining and in situ hybridization. With a biotinylated porcine Y chromosome-specific deoxyribonucleic acid probe, male fibroblasts could be clearly distinguished from female fibroblasts. In all leaflets we observed both donor and recipient fibroblasts. The distribution of these populations was marked in schematic drawings. Recipient fibroblasts mostly spread onto the leaflet surface but also penetrated the leaflet tissue. Remaining donor fibroblasts did not show morphologic signs of decreased viability on hematoxylin-eosin staining. In situ hybridization may become a useful technique in homograft research. In this porcine model, the fibroblasts in the aortic homograft explants were of both donor and recipient origin.

Assessment of numeric abnormalities of X, Y, 18, and 16 chromosomes in preimplantation human embryos before transfer

OBJECTIVE: Our purpose was to determine the feasibility of ascertaining aneuploidy for chromosomes X, Y, 18, and 16 by use of multiple-probe fluorescence in situ hybridization in blastomeres from preimplantation human embryos. STUDY DESIGN: A short fluorescence in situ hybridization procedure involving the simultaneous use of four deoxyribonucleic acid probes detected with red, green, blue, or a mixture of red and green fluorochromes was developed to determine numeric abnormalities of chromosomes X, Y, 18, and 16. Embryos underwent biopsy, and all or most cells were analyzed to distinguish true aneuploidy from mosaicism and to assess technique variations within the same embryo ( n = 64). RESULTS: The analysis of all the blastomeres of an embryo was achieved in 91% of the embryos. Successful analyses including biopsy, fixation, and fluorescence in situ hybridization were achieved in 87.8% of the blastomeres. Of the four chromosomes tested, numeric aberrations were found in 23% and 42% of normally and abnormally developing embryos, respectively, includig aneuploidy, polyloidy, haploidy, and mosaicism. When diploid embryos containig one or several tetraploid cells are counted as chromosomally abnormal, then 49% and 61% of normally and abnormally developing embryos, respectively, were chromosomally abnormal. Aneuploid embryos consisted of two monosomies for chromosome 16, one for chromosome 18, and a trisomy for chromosome 16. There was a tendency for aneuploidy to increase with maternal age. CONCLUSIONS: Fluorescence in situ hybridization is a more efficient method than cytogenetic analysis to study specific aneuploidies at preimplantation stages of development in human embryos. In addition, the preimplantation genetic diagnosis of two blastomeres per eight-cell embryo may be sufficient to ensure successful analysis of polyploidy, haploidy, and specific aneuploidies without endangering the survival of the embryo. The technique can be easily modified to consider other chromosomes, including 13 and 21. Because most chromosomally abnormal embryos do not develop to term, the use of this technique may increasethe delivery rate per embryo by allowing only transfer of embryos normal for the tested chromosomes. This technique would be most useful for older women undergoing in vitro fertilization, because aneupoidy appears to increase with advancing maternal age.

46, XY Female Karyotype Containing SRY

We report the case of a girl with a 46, XY karyotype containing the sex determining region Y gene (SRY). She was 14 years old, with the height of 131.2 cm (-4.5 SD). She had a low hair line, shield-shaped chest and multiple pigmented naevi on her face. The external genitalia were normal female. No inguinal or labial masses were found. Chromosomal analysis from blood lymphocytes and skin fibroblasts revealed a 46, XY karyotype. The basal levels of serum luteinizing hormone (LH) and follicle stimulating hormone (FSH) and their responses to LH-releasing hormone (RH) stimulation were elevated; the insulin-like growth factor-I (IGF-I), thyroid function and growth hormone (GH) secretory responses to insulin and glucagon were normal. SRY was identified on the short arm of the Y chromosome using Y-specific deoxyribonucleic acid (DNA) probes. There were no mutations in the SRY by direct sequencing analysis. Pelvic ultrasound and magnetic resonance imaging (MRI) did not reveal internal genitalia. Gonadectomy may be necessary because she has a substantial risk of developing malignant germ cell neoplasia. Two questions arise: 1) why was male sexual differentiation inhibited although she had a normal SRY, and 2) why did she have the appearance of Turner syndrome?

Control of peripartal collagenolysis in the human chorion-decidua

OBJECTIVE: This study was designed to observe the in vivo status of chorion-decidua gene expression for some major metalloproteinases, an inhibitor (tissue inhibitor of metalloproteinase-1), and an activator (tissue plasminogen activator) and the hormone relaxin at accessible time points in the peripartial period. STUDY DESIGN: Chorion-decidua from patients at cesarean section with and without labor and after spontaneous labor and delivery was used for preparation of poly (A) + ribonucleic acid and quantitative Northern analyses with a series of oligo and complementary deoxyribonucleic acid probes. RESULTS: In the period designated as the period before parturition, relaxin and matrix metalloproteinase-1 (interstitial collagenase) gene expression were relatively high, reflecting the controlled loss of amniotic collagen necessary for fetal membrane expansion without rupture as the uterine volume increases. When active labor has begun, but before delivery, matrix metalloproteinase-3 (stromelysin) and matrix metalloproteinase-9 (type V collagenase) gene levels significantly increased. After normal spontaneous labor and delivery, tissue plasminogen activator and tissue inhibitor of metalloproteinase-1 messenger ribonucleic acids significantly increased, together with a marginally increased expression of the genes for matrix metalloproteinase-1, matrix metalloproteinase-2 (type IV collagenase), and relaxin. CONCLUSION: The expression of the genes for some major collagenolytic enzymes, an inhibitor, activator, and relaxin in the chorion-decidua, is different at the different stages of parturition.

Validation Studies on the Forensic Analysis of Restriction Fragment Length Polymorphism (RFLP) on LE Agarose Gels Without Ethidium Bromide: Effects of Contaminants, Sunlight, and the Electrophoresis of Varying Quantities of Deoxyribonucleic Acid (DNA)

This study was designed to analyze the effects of sunlight, various contaminants (those found typically in forensic samples) and the electrophoresis of varying quantities of DNA on the restriction fragment length polymorphism (RFLP) patterns produced from DNA isolated from blood and semen stains. The DNA RFLP patterns were obtained following Hae III restriction enzyme digestion, low electroendosmotic (LE) agarose gel electrophoresis (in the absence of ethidium bromide), Southern transfer, hybridization with DNA probes detecting highly polymorphic variable number of tandem repeats (VNTRs) and autoradiography. Computer assisted image analyses were used to detect variations in RFLP band sizes in relation to control samples. Comparisons between the samples were made for the presence of high molecular weight DNA, the ability to achieve a complete restriction digestion, and the RFLP fragment sizes obtained. The results demonstrate that high molecular weight DNA can be obtained when blood and semen stains are subjected to environmental and contaminating factors. The RFLP allele sizes were not significantly affected by environmental conditions, contamination factors or by loading varying amounts of DNA. This study serves to further document the reliability and validity of DNA typing for forensic applications.

Development and Evaluation of Oligonucleotide DNA Probes for Detection and Genotyping of Shiga-like Toxin Producing Escherichia coli

Use of antisense oligonucleotide probes (OP) for detection and genotyping of Shiga-like toxin producing Escherichia coli (SLTEC) was evaluated. Based on published deoxyribonucleic acid (DNA) sequences of the A subunits of Shiga-like toxin (SLT) I and II genes, three synthetic antisense OP were constructed (OP-1, -2 and -3). Their use for detection and genotyping of SLTEC was evaluated and the results were compared to those obtained using cloned toxin-gene probe fragments. Both the OP-1 and OP-2 hybridized with all 69 SLT-I and II producing strains. Furthermore, the OP-1 hybridized only with all 18 SLT-I-only producing strains, and the OP-2 hybridized only with all 48 SLT-II-only producing strains. OP-3, a pool of four OP, detected all SLTEC strains regardless of toxin genotype. None of the OP hybridized to any of the 91 SLT-negative strains. These results demonstrate the three OP to be unique reagents for SLTEC epidemiological studies.

Deoxyribonucleic Acid Hybridization Method for the Detection of Listeria in Dairy Products, Seafoods, and Meats: Collaborative Study

The method is based on the hybridization of synthetic deoxyribonucleic acid probes to ribosomal ribonucleic acid sequences unique to Listeria. This method was compared to 2 culture methods: the U.S. Food and Drug Administration method for the detection of Listeria in dairy products and seafoods and the U.S. Department of Agriculture, Food Safety and Inspection Service method for Listeria in meats. Six food types with replicate samples containing various concentrations of Listeria were analyzed by the collaborating laboratories. Listeria was detected in 774 samples using the DNAH method and in 772 samples using a culture method. The DNAH and culture methods were in agreement for 668 samples containing Listeria and 306 samples without Listeria. The overall rate of agreement between methods was 82.3%. The method has been adopted first action by AOAC INTERNATIONAL.

The Role of Brain-derived Neurotrophic Factor in Transient Forebrain Ischemia in the Rat Brain

Brain-derived neurotrophic factor (BDNF) may play a role in the pathophysiology of neuronal cell death after cerebral ischemia. We investigated alterations in BDNF gene expression and the effect of BDNF on neuronal death after transient forebrain ischemia in the rat brain. Transient forebrain ischemia was induced by occlusion of the bilateral common carotid arteries and by producing systemic hypotension for 8 minutes. The alterations in the BDNF messenger ribonucleic acid content in the hippocampus and the cerebral cortex were examined by Northern blot analysis, using a phosphorus-32-labeled mouse BDNF complementary deoxyribonucleic acid probe. Recombinant Chinese hamster ovary cells with BDNF-secreting capacity were established by expression vector transfection with BDNF complementary deoxyribonucleic acid. The effect of BDNF on neuronal death in the hippocampal CA1 region after ischemia was then examined by using a continuous intraventricular infusion of 200 microliters of normal (Group II, n = 6) or 30-times concentrated recombinant Chinese hamster ovary cell culture medium containing BDNF (Group IV, n = 6). Normal (Group I, n = 6) or 30-times concentrated (Group III, n = 6) Chinese hamster ovary cell culture medium, not including BDNF complementary deoxyribonucleic acid, was infused into the same ischemic brains, which served as controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular biological approaches to the epidemiology of diarrhoeal diseases in developing countries

Diarrhoea in developing countries is caused by an increasingly long list of bacterial, viral, and parasitic pathogens with rotavirus, enterotoxigenic Escherichia coli, Campylobacter, Shigella, and Salmonella heading the list. Using methods to detect most of the known enteropathogens, one or more enteropathogen(s) is isolated in two-thirds of diarrhoeal illnesses in the developing world. Many of these enteropathogens are also frequently isolated from children without diarrhoea. An aetiologic agent is more frequently isolated from cases of invasive diarrhoea than from those with secretory diarrhoea. Deoxyribonucleic acid probes have proved very useful in detecting pathogens such as enterotoxigenic (ETEC), enteroinvasive (EIEC), and enteropathogenic E. coli (EPEC), but have not yet proved to be particularly rapid or less expensive. Molecular biology has proved useful in epidemiological studies as a means of strain identification. Plasmids were initially used as convenient markers and proved useful in identifying epidemic strains of bacteria. Other molecular markers, such as ribotyping, are accurate enough to be used as taxonomic tools.

Human cutaneous leishmaniasis caused by Leishmania donovani s.l. in Kenya

Our laboratory is characterizing Leishmania stabilates and isolates from active leishmaniasis cases. Smears and cultures from aspirates made on different dates from a single lesion on the bridge of the nose of an 18 years old Kenyan male from Nyandarua District contained Leishmania. The isolates, NLB-271 and NLB-271-IA, were characterized by cellulose acetate electrophoresis (CAE) using 20 enzyme systems and by Southern analysis using 2 deoxyribonucleic acid (DNA) probes (pDK10 and pDK20) from a Dakar strain of L. major (MHOM/SN/00/DK1) and a third probe, p7-059 from L. infantum strain ITMAP-263. Digestion of the two Leishmania DNAs with endonucleases HindIII and PstI, followed by hybridization with the 3 probes, revealed DNA fragment banding patterns indistinguishable from those of the L. donovani species complex. The CAE isoenzyme profiles of these 2 Kenyan isolates were indistinguishable from those of Kenyan L. donovani strains we designated as zymodeme Z6. Excluding post-kala-azar dermal leishmaniasis, this constitutes the first human case of cutaneous leishmaniasis caused by L. donovani s.l. in Kenya. Previously, cutaneous leishmaniasis cases in Kenya have been due to L. aethiopica, L. major and L. tropica only.

VENTRICULAR EXPRESSION OF ATRIAL NATRIURETIC PEPTIDE AFTER ORTHOTOPIC CARDIAC TRANSPLANTATION

In order to determine if atrial natriuretic peptide is produced by the human ventricle after transplantation, specimens were obtained from 54 separate right ventricular endomyocardial biopsies from 14 transplant recipients (age range 10-19 years) and were analyzed with simultaneous plasma samples. Radioimmunoassay techniques were used to determine plasma and tissue levels of atrial natriuretic peptide in each biopsy specimen, and blot hybridizations with the atrial natriuretic peptide c deoxyribonucleic acid probe were performed once for each patient. Mean plasma levels of atrial natriuretic peptide were obtained from the right ventricle (106 +/- 12 pg/ml); there was a positive correlation with end diastolic pressure (r = 0.7, P < 0.0001). Plasma levels, however, were not significantly increased during rejection episodes. Atrial natriuretic peptide was detectable in all ventricular biopsy specimens--mean 989 +/- 164 pg/mg soluble protein. Mean tissue level during rejection episodes was 1163 +/- 17 and following treatment of rejection was 411 +/- 147--however, this difference was not significant (P = 0.55). In 6 patients biopsied within 1 month of transplantation initial myocardial levels were significantly higher than those obtained at the following biopsy (P < 0.05). Despite these initially high tissue levels no correlation was found with time posttransplantation. Ventricular tissue levels positively correlated with systolic ventricular pressure, r = 0.3, P < 0.05, but there was no significant correlation with other hemodynamic parameters. The normalized atrial natriuretic peptide messenger ribonucleic acid signal from the slot blot hybridization was 59 +/- 6 mm (peak height of signal intensity) range 20-110 mm. The expression of atrial natriuretic peptide gene was also confirmed by Northern blot analysis. The ventricular messenger ribonucleic acid signal contained a single hybridizing band of approximately 920 nucleotides. Thus, we have demonstrated that atrial natriuretic peptide production by the human ventricle is independent of innervation. The consistent atrial natriuretic peptide gene expression and detection by radioimmunoassay in apparently healthy ventricles was unexpected but may reflect the use of immunosuppressive drugs, or perhaps subclinical graft dysfunction.

Detection of Chlamydia trachomatis in semen by the polymerase chain reaction in male members of infertile couples

Our objective was to evaluate the presence of asymptomatic Chlamydia trachomatis infection by means of the polymerase chain reaction in male members of couples with previously undiagnosed infertility. Twenty-eight infertile-couples who had negative cultures or negative results when tested by deoxyribonucleic acid probe for Chlamydia trachomatis in semen and cervical samples were studied. Semen samples were tested for Chlamydia trachomatis by means of the polymerase chain reaction. Sera from both partners were diluted 1:128 and tested for immunoglobulin M antibodies to Chlamydia. Sera and ejaculated sperm were evaluated for the presence of antisperm antibodies. Semen analyses were also performed. Chlamydia trachomatis was identified in semen from 11 (39.3%) of the male partners. Its detection correlated with the presence in the ejaculate of motile sperm containing antisperm antibodies (p < 0.01). Either antisperm immunoglobulin G, immunoglobulin A, or both were located on sperm only from 5 (45.5%) of the 11 men whose results were positive when tested for Chlamydia trachomatis. Similarly, immunoglobulin G or immunoglobulin A antibodies to sperm were only detected in 5 (45.5%) of the spouses of men with Chlamydia in semen. Immunoglobulin M antibody to Chlamydia trachomatis was identified in only one of the men. However, antichlamydial immunoglobulin M antibodies were present in sera from 6 (54.5%) female partners of men with seminal Chlamydia trachomatis but in none of the other 17 women (p < 0.01). Although undetected by culture of deoxyribonucleic acid probe of semen samples, Chlamydia trachomatis was nevertheless identified in semen of some symptom-free men by the polymerase chain reaction. This is probably a result of the increased sensitivity of the polymerase chain reaction to detect Chlamydia trachomatis. The increased prevalence of an autoimmune response to sperm in men with this organism in their semen suggests that a subclinical chlamydial infection may activate an immune response to sperm. A similar association between Chlamydia trachomatis in semen and circulating antisperm antibodies in female partners indicates that Chlamydia may also induce an immune response to sperm in women. Infertility in these couples may be the result of a direct inflammatory response in the cervix or endometrium to repeated Chlamydia exposure or of the ability of Chlamydia to evoke an immune response to spermatozoa.

Interleukin-1 type I receptor messenger ribonucleic acid expression in human endometrium throughout the menstrual cycle

To investigate the messenger ribonucleic acid (mRNA) expression of interleukin-1 (IL-1) type I receptor in the endometrial tissue of normal patients during the menstrual cycle. Prospective longitudinal study. Department of Obstetrics and Gynecology, Stanford University Medical Center, Stanford, California. Twenty fertile women between 19 and 41 years of age underwent hysterectomy for benign reasons (n = 9) and laparoscopy for tubal ligation (n = 11). In all cases, endometriosis was not visualized. Endometrial biopsy using the Novak curette was obtained at the time of surgery. Total RNA extracted from unfractioned endometrial tissue was analyzed on Northern blots by using specific complementary deoxyribonucleic acid probes. We found IL-1 type I receptor mRNA expression in endometrial tissue throughout the entire menstrual cycle. However, IL-1 type I receptor mRNA levels were significantly higher during both early and late luteal phases than follicular and midluteal phases. Our results demonstrate the presence of the IL-1 system in the human endometrium and that the receptor is regulated throughout the menstrual cycle with a 4.1-fold increased expression of the IL-1 receptor gene in the early luteal phase compared with preovulatory endometrium.

Tuberculous Spondylitis as A Complication of Intravesical Bacillus Calmette-Guerin Therapy

We report a case of tuberculous spondylitis following intravesical bacillus Calmette-Guerin (BCG) instillation. A 90-year-old male physician living in South Africa received an uncomplicated 6-week course of intravesical BCG (Japanese 172 strain) for high grade superficial bladder carcinoma. He experienced a sudden onset of debilitating lower back pain 16 months following this treatment. A lytic lesion involving the anterior Til and T12 vertebral bodies was diagnosed and subsequently biopsied. An acid-fast organism was isolated after 3 weeks of incubation and was confirmed through deoxyribonucleic acid probe hybridization as a mycobacterium. High performance liquid chromatography analysis speciated the organism as Mycobacterium bovis BCG, proving that it was acquired through the intravesical therapy.

Detection by in situ hybridization of HIV and c‐myc RNA in tumour cells of AIDS‐related B‐cell lymphomas

The incidence of acquired immune deficiency syndrome (AIDS)-related malignant lymphoma has increased since the disease was first described, but its pathogenesis is still not understood. There have been numerous molecular studies addressing the clonality of these proliferations, the presence of Epstein-Barr virus genome in the tumor cells, and rearrangements of the c-myc oncogene. However, very few in situ hybridization studies have been carried out. We analysed 24 cases of high-grade B-cell malignant lymphomas and two cases of polymorphic B-cell proliferation associated with human immunodeficiency virus (HIV) infection. Human immunodeficiency virus ribonucleic acids were detected in some of the tumour cells in 19 of the 24 cases of malignant lymphomas and in both cases of polymorphic B-cell proliferation, with the in situ hybridization technique and using a specific tritiated copy deoxyribonucleic acid probe. With the same technique, c-myc ribonucleic acid was detected in most of the tumour cells from all the 21 cases of malignant lymphomas tested but not in the polymorphic B-cell proliferation.

Genetic study of hydatidiform moles by restriction fragment length polymorphisms (RFLPs) analysis.

Twenty three hydatidiform moles (HMs) were studied using the techniques of "RFLPs" employing a minisatellite deoxyribonucleic acid probe. Among the 23 HMs, 17 were homozygous types resulting from a duplicated haploid sperm, and two were heterozygous types resulting from fertilization two independent sperms (dispermy). It was revealed that the four histopathologically diagnosed complete HMs (CHMs) were partial HMs (PHMs) with one maternal and 2 paternal chromosome contribution (diandry) or two maternal and 1 paternal alleles (digyny). The locus specific minisatellite probes were useful in classifying CHM into heterozygous and homozygous types as well as in diagnosing PHM. One heterozygous (50%) and 5 homozygous (29.4%) CHMs, and one PHM (25%) progressed to persistent gestational trophoblastic disease (p > 0.5).