Farzin Farzaneh

Investigation of maternal blood enriched for fetal cells : Role in screening and diagnosis of fetal trisomies

Prenatal diagnosis of chromosomal abnormalities relies on assessment of risk followed by invasive testing in the group with highest risk. Assessment of risk by a combination of maternal age and fetal nuchal translucency and invasive testing in the 5% of the population with the highest risk would identify about 80% of trisomy 21 pregnancies. Preliminary reports suggest that chromosomal abnormalities can also be diagnosed by fluorescent in situ hybridization (FISH) in maternal blood enriched for fetal cells. This study examines the potential role of this method on the prenatal diagnosis of fetal trisomies. Maternal blood was obtained before invasive testing in 230 pregnancies at 10-14 weeks of gestation. After enrichment for fetal cells, by triple density centrifugation and anti-CD71 magnetic cell sorting, FISH was performed and the proportion of cells with positive signals in the chromosomally normal and abnormal groups was determined. Fetal karyotype was normal in 150 cases and abnormal in 80 cases, including 36 with trisomy 21. Using a 21 chromosome-specific probe, three-signal nuclei were present in at least 5% of the enriched cells from 61% of the trisomy 21 pregnancies and in none of the normal pregnancies. For a cut-off of 3% of three-signal nuclei the sensitivity for trisomy 21 was 97% for a false positive rate of 13%. Similar values were obtained in trisomies 18 and 13 using the appropriate chromosome-specific probe. Examination of fetal cells from maternal blood may provide a noninvasive prenatal diagnostic test for trisomy 21 with the potential of identifying about 60% of affected pregnancies. Alternatively, this technique can be combined with maternal age and fetal nuchal translucency as a method of selecting the high-risk group for invasive testing. Potentially, 80% of trisomy 21 pregnancies could be identified after invasive testing in less than 1% of the pregnant population.

ADP-ribosyl transferase activity in Trypanosoma brucei.

Nuclear adenosine diphosphoribosyl transferase (ADPRT) catalyses the covalent modification of chromatin proteins by (ADP-ribose)n. This activity, which is entirely dependent on DNA containing strand breaks, is required for efficient DNA excision repair possibly because it regulates DNA ligation. ADPRT activity is also required for cytodifferentiation in a number of different cell types. We report here the presence of ADPRT activity in the blood-stream form of Trypanosoma brucei and its activation by DNA strand breaks formed by exposure to, either exogenously supplied deoxyribonuclease I, or treatment with the methylating agent, dimethylsulphate. 3-Aminobenzamide, but not its chemical analogue 3-aminobenzoic acid, is a competitive inhibitor of ADPRT activity in T. brucei. Intact trypanosomes are readily permeable to this competitive inhibitor of ADPRT activity.

Transient Formation of DNA Strand Breaks During DMSO-Induced Differentiation of Promyelocytic Cell Line, HL-60

We have previously shown that during the spontaneous induction of differentiation in primary avian myoblasts several hundred single-strand DNA breaks are formed in the genome [1]. This increase in the detectable number of DNA strand breaks is not due to a general deficiency in DNA repair mechanisms and γ-radiation induced breaks are as efficiently repaired in the post-mitotic nuclei of terminally differentiating myotubes as they are in the proliferating myoblasts. The formation of DNA strand breaks during myoblast differentiation is accompanied by an increase in ADPRT activity [1]. The nuclear ADPRT activity, which is totally dependent on the presence of DNA strand breaks, is required for the efficient ligation of DNA strand breaks [2]. Inhibition of this activity, either by enzyme inhibitors or substrate deprivation, blocks myoblast differentiation but not their proliferation [1]. Here, we report the transient formation of DNA strand breaks during the induced differentiation of the promyelocytic cell line, HL-60. Inhibition of ADPRT activity blocks the religation of these breaks which are formed in the face of a proficient DNA repair mechanism. ADPRT activity is also required for the excision repair of DNA strand breaks formed by either γ-irradiation or exposure to the mono-functional methylating agent dimethylsulphate (DMS).