Peripheral neural activity recording and stimulation system.

This paper presents a portable, embedded, microcontroller-based system for bidirectional communication (recording and stimulation) between an electrode, implanted in the peripheral nervous system, and a host computer. The device is able to record and digitize spontaneous and/or evoked neural activities and store them in data files on a PC. In addition, the system has the capability of providing electrical stimulation of peripheral nerves, injecting biphasic current pulses with programmable duration, intensity, and frequency. The recording system provides a highly selective band-pass filter from 800 Hz to 3 kHz, with a gain of 56 dB. The amplification range can be further extended to 96 dB with a variable gain amplifier. The proposed acquisition/stimulation circuitry has been successfully tested through in vivo measurements, implanting a tf-LIFE electrode in the sciatic nerve of a rat. Once implanted, the device showed an input referred noise of 0.83 µVrms, was capable of recording signals below 10 µ V, and generated muscle responses to injected stimuli. The results demonstrate the capability of processing and transmitting neural signals with very low distortion and with a power consumption lower than 1 W. A graphic, user-friendly interface has been developed to facilitate the configuration of the entire system, providing the possibility to activate stimulation and monitor recordings in real time.

[Doppler color ultrasonography in patients with erectile dysfunction]. / Eco-color Doppler in pazienti affetti da disfunzione erettiva.

The introduction of new oral therapy doesn't change the rule of color Doppler US in the screening modality of the diagnosis of vasculogenic impotence. The hemodynamic changes can be non invasively evaluated with color and energy Doppler US and with spectral analysis following injection of a vaso-active pharmacological agent. The new high sensitive US equipment allows an easy detection of the cavernous artery. Duplex Doppler can quantify the systolic-diastolic flow changes after the intra-cavernous vaso-active drugs injection. It is then possible to establish the diagnosis of arterial or venous vasculogenic impotence. The organic cause can also be excluded. The pick systolic velocity less than 0.30 m/sec is indicative of arterial origin. The absence of flow-reversal in the diastolic phase, 30 minutes after the injection, is significant for a venous type problem. Elicine arteries can be visualised and analysed using high sensitive color Doppler energy US equipment.

3q21 and 3q26 cytogenetic abnormalities in acute myeloblastic leukemia: biological and clinical features.

BACKGROUND AND OBJECTIVE: Acute myeloblastic leukemia (AML) with features of myelodysplastic syndrome (MDS) and abnormalities of megakaryocytopoiesis is often characterized by cytogenetic aberrations of the 3q21 and 3q26 bands involving inv(3)(q21q26) and (3;3)(q21;q26). These aberrations have been described in all FAB subtypes with the exception of M3, and in MDS and in megakaryoblastic crisis of chronic myeloid leukemia. We reviewed the biological and clinical features of 10 cases of AML with inv(3)(q21q26) and t(3;3)(q21;q26). DESIGN AND METHODS: Four hundred and sixteen patients with AML were studied in our Institute by cytogenetic analysis and 10 (2.4%) showed inv(3)(q21q26) (7 patients) or t(3;3)(q21;q26) (3 patients): 7 males, 3 females; median age, 43.5 yrs. We also used RT-PCR to investigate the pattern of expression of the EVI-1 gene in 5 patients. RESULTS: Additional chromosomal changes were demonstrated in 6 patients. In 5/10 cases a preceding MDS had been observed. A possible occupational exposure was established in 2 patients (a farmer and an histologist employing organic solvents) and another patient had a therapy-related leukemia. AML subtype was M1 in 9 patients and M2 in 1. A variable excess of micromegakaryocytes was observed in all the patients. In 5 patients the platelet count was normal or increased (median number: 172. 5x10(9)/L; range 55-440). Expression of EVI-1 gene was present in all the 5 patients studied. The clinical course and outcome was extremely poor: 9/10 patients were resistant and 1 patient showed a partial remission after induction therapy. Of the 9 patients resistant to the first line chemotherapy, 7 were also resistant to the second line chemotherapy. Three patients obtained a morphologic complete remission after third line chemotherapy (duration 1, 3 and 6 months); 2 of them were submitted to autologous bone marrow transplantation, but relapsed after 1 and 3 months. The median overall survival was 5.5 months. INTERPRETATION AND CONCLUSIONS: Our findings evidence a strong correlation between 3q21q26 chromosomal aberrations, abnormalities of megakaryocytopoiesis and lack of response to conventional chemotherapy and support the diagnostic and prognostic relevance of chromosome characterization in the classification of AML.

Detection and monitoring of trisomy 8 by fluorescence in situ hybridization in acute myeloid leukemia: a multicentric study.

BACKGROUND AND OBJECTIVE: The role of fluorescence in situ hybridization (FISH) in the detection and monitoring of trisomy 8 (+8) in acute myelogenous leukemia (AML) has not been defined exactly. This multicentric study was performed in order to: i) analyze the sensitivity of interphase FISH with respect to conventional chromosome analysis (CCA) in detecting +8; ii) compare the results of FISH and CCA in the quantitation of the frequency of +8-positive cells; iii) analyze the possible role of FISH in the cytogenetic follow-up of patients with +8. DESIGN AND METHODS: One hundred and ninety-eight nonconsecutive patients with a diagnosis of AML seen at five centers over a 3-year period were studied by CCA and FISH with a chromosome 8-specific centromeric probe. Two hundred interphase cells were scored in each test and the cut-off for the recognition of +8 was set at 3%. An irrelevant pericentromeric probe was used as negative control in those cases with an apparently normal karyotype and trisomy 8 in interphase cells. FISH studies were conducted at diagnosis and, in 14 cases with +8, on 1.5 occasions during follow-up. RESULTS: Karyotype aberrations were seen in 121 cases (61.1%), with +8 being present in 38 of them (16 as the sole aberration). Interphase FISH detected +8 in 37/38 cases; in a patient with 1/10 metaphases with +8, 2.3% interphase cells with 3 signals were seen. Fourteen additional cases with occult +8 were detected by FISH, which showed 4-22% interphase cells with three signals; 6 patients had an abnormal karyotype without +8, 3 had a normal karyotype, 5 had no analyzable mitoses. In 24 cases with > 15 analyzable metaphases, percent variations between CCA and FISH in the estimation of the size of the trisomic clone ranged between 0.4% and 51%, median value 22%. Underestimation of the percent of trisomy 8 by FISH occurred in all 10 cases with > 90% +8 metaphases. In 7/14 cases investigated sequentially, FISH detected 5-35% trisomic cells in the BM after induction therapy (4 CR, 3 PR); 4 cases relapsed with +8 at 8-15 months. The absence of +8 in remission marrows was documented in the remaining 7 cases, 4 of which relapsed at 20-32 months. INTERPRETATION AND CONCLUSIONS: It is concluded that FISH was a valuable method in this multicentric study since it showed greater sensitivity than CCA in detecting minor clones with +8, in patients with both normal and abnormal karyotypes. The role of FISH in the cytogenetic follow-up of trisomies in AML patients may be promising.

Autologous peripheral blood stem cell transplantation in acute myeloblastic leukaemia and myelodysplastic syndrome patients: evaluation of tumour cell contamination of leukaphereses by cytogenetic and molecular methods.

We evaluated 18 acute myeloblastic leukaemia (AML) and myelodysplastic syndrome (MDS) patients with abnormal karyotype at diagnosis who underwent peripheral blood stem cell (PBSC) transplantation. To evaluate the presence of residual tumour cells, bone marrow (BM) samples and PBSC collections were analysed by cytogenetics and in selected cases also by fluorescence in situ hybridisation (FISH) and molecular studies. All patients were considered to be in morphologic and cytogenetic complete remission (CR) at the time of mobilisation. Seven patients showed neoplastic cells in PBSC harvest and/or BM specimen before reinfusion. Cytogenetic studies revealed contamination in apheretic collections in one patient only, while three patients had BM but not PBSC contamination. Three more patients had leukaemic cells both in the BM and PBSC. All but one (with only BM contamination) of these patients relapsed within 9 months. However, five more patients relapsed after transplantation: in four cases there was no cytogenetic sign of contamination either in PBSC or BM cells and in one case no molecular evidence was revealed either. This study suggests that, whereas the presence of leukaemic cells in autologous grafts correlates with a poor prognosis, the lack of detection of tumour cells is not always predictive of long-term disease-free survival. More importantly, PBSC collections from AML patients are not contaminated by leukaemic cells if the BM is disease-free.

A new method of "in-cell reverse transcriptase-polymerase chain reaction" for the detection of BCR/ABL transcript in chronic myeloid leukemia patients.

Methods of detecting minimal residual disease (MRD) in chronic myeloid leukemia (CML) include chromosome analysis, Southern blotting, polymerase chain reaction (PCR) and fluorescent in situ hybridization (FISH) techniques. We report a novel method to detect intracellular messenger RNA (mRNA) by combining the techniques of reverse transcription (RT) and PCR performed directly inside the cells, without extraction of the nucleic acid. We applied this method, which we call "in-cell RT-PCR", to detect hybrid BCR/ABL transcript within single cells. After cellular permeabilization and fixation of single cells in suspension, the neoplastic mRNA was reverse transcribed into cDNA, and the cDNA was amplified by PCR with fluorescent primers, specific for bcr/abl. Flow cytometry was used to detect cells positive for the amplified DNA within the cell cytoplasm. After transferring the amplified cells onto slides by cytospin, the positive cells for BCR/ABL cDNA were observed by fluorescent microscopy. The technique was capable of detecting low abundancy signals and distinguishing different levels of gene expression. The amplification products were found in the cells and supernatants. The distribution was critically affected by the protease digestion condition. The specificity of amplification was confirmed by a nested RT-PCR of BCR/ABL performed on extracted mRNA from the same sample, and by reamplification of supernatants. We have used the technique to study 10 Ph+ CML patients and three normal subjects as controls. Four patients were 100% Ph+ at diagnosis time and RT-PCR+ at cytogenetic and molecular analysis, respectively. In-cell RT-PCR showed that the residual non-neoplastic cells could be observed in all cases. In two patients undergoing interferon-alpha (IFN-alpha) therapy and in four bone-marrow transplanted patients, the in-cell RT-PCR was used to compare the level of Ph+ positivity detected by cytogenetic analysis with the number of cells expressing BCR/ABL transcript. In this manner, we could estimate the MRD. Our preliminary application of the technique suggests that it is capable of accurately identifying cells transcribing bcr/abl, and that it may have significant clinical applications in the detection of MRD.

Rectal diazepam: a clinical and EEG study after a single dose in children.

EEG monitoring was performed on a group of ten children, aged 3 days to 13 years, during admission to the Paediatric Clinic at the University of Bologna for seizures. Diazepam was injected by rectal route as an anticonvulsant, using a dose of 0.5-1 mg/kg in the first nine children, whereas an intravenous bolus was administered to the last child. The main aim of this study was to document the time taken for the drug to reach the brain and to modify the electrical activity. We observed significant EEG changes between 1 and 9.30 min and, in particular, the appearance of fast activity over one or both hemispheres after rectal diazepam. The EEG results confirm the clinical efficacy of rectal diazepam.

Screening of thalassaemia carriers in intermediate school of Latium: results of four years' work.

From 1975 to 1979 the Rome Microcythaemia Centre carried out four health education campaigns and thalassaemia screenings among students of the intermediate schools throughout Latium, under the auspices and with the financial support of the Health Authorities of the Latium region. This project is aimed at avoiding reproduction by pairs of thalassaemia carriers and the birth of homozygous children. During the four campaigns 138 501 students were examined, that is, 70 to 76% of those enrolled. Of these 3343 were found to be thalassaemic. Thus the overall prevalence of thalassaemia in Latium is 2.41%, with minor fluctuations from one province to another and, above all, a slight, though definite, trend towards higher values in the southern part of Latium bordering on Campania. The screening was welcomed by the population and the thalassaemic families, there were no detectable negative side-effects, and it resulted in an increased awareness of the problem of the thalassaemias.

First premarital screening of thalassaemia carriers in intermediate schools in Latium.

In the 1975 to 1976 school year, under the auspices of the Health Authorities of the Latium Region, the Rome Microcythaemia Centre carried out for the first time a partial screening survey of thalassaemia carriers among the students of the compulsory intermediate school in Latium. This work was the beginning of a new preventive school health service aimed at the prophylaxis of Cooley's disease. In 23 places investigated in Latium, 17724 students were examined, 13354 of whom were in Rome and 4370 elsewhere. The mean percentage of co-operation was 70% and the mean percentage of thalassaemia 2.42%. Thalassaemic students were invited to attend the centre for a check-up along with their families: about half had already come in by the end of June 1976. All students examined, whether normal or thalassaemic, have received written results of the tests. The screening survey aroused notable interest and obtained wide approval both at school and at home. The news of being thalassaemia carriers, even if not welcome, was never the cause of family tragedy.

Heterozygous beta-thalassaemia with normal haemoglobin pattern. Haematologic, haemoglobin and biosynthesis study of 4 families.

Thalassaemia with normal levels of Hb A2 and Hb F and with an alpha/beta ratio higher than 1 is described in 4 families. 3 of these families show direct or indirect signs of the presence of the delta-thalassaemia gene along with the beta-thalassaemia gene. The fourth family leaves the question as to whether there is a single mutation of the deltabeta tract or a beta + delta-thalassaemia in coupling unanswered. The necessity of knowing of the existence of this thalassaemia which conceals the presence of a beta-thalassaemia gene, is stressed, above all in view of the danger that mating between a carrier of this thalassaemia and a carrier of classical beta-thalassaemia could result in the birth of children with Cooley's disease.

Genetic patterns in thalassemia intermedia (constitutional microcytic anemia). Familial, hematologic and biosynthetic studies.

Globin chain synthesis has been studied in 17 patients with thalassemia intermedia, and their relatives, also investigated by routine hematologic and hemoglobinic tests. The mean a/non a ratio was always around 2.20-2.30. In patients with severe thalassemia major, used as a control, the mean a/non a ratio was significantly higher, that is 3.11-3.07. Therefore, the hypothesis that the cause of the lesser severity of the thalassemia intermedia is a lesser imbalance of globin chain synthesis, is suggested. One or both the parents of patients with thalassemia intermedia have mild beta-thalassemia and normal alpha/beta ratio, whereas the parents of patients with severe thalassemia major show a marked beta-thalassemia and a mean alpha/beta ration of 1.76. These data suggest that genes for beta +-thalassemia are responsible for thalassemia intermedia, and genes for beta o-thalassemia are responsible for thalassemia major. In two patients with thalassemia intermedia, the association of an alpha-thalassemia gene with homozygous beta-thalassemia that it is well known to reduce the globin chain imbalance typical of the beta-thalassemia, has also been observed.