KIR genotypic diversity in Portuguese and analysis of KIR gene allocation after allogeneic hematopoietic stem cell transplantation.

The diversity of killer-cell immunoglobulin-like receptors (KIR) genes was evaluated in Portuguese and the observed genotypic profiles were found related to the ones reported in European populations. The KIR repertoire after hematopoietic stem cell transplantation is determined by these gene frequencies and the KIR group B motifs are the less common. We estimated donor-KIR/recipient-ligand interactions in transplants with related donors and unrelated donors found in a local registry or from abroad. A large fraction of transplants had all three ligands of inhibitory receptors, and therefore, in theory were not prone to natural killer cell (NK) mediated alloreactivity. Furthermore, the distribution of KIR alloreactive interactions was found independent of the donor-recipient genetic proximity, probably because of different gene segregation and comparable KIR frequencies in the donor pools.

CHK1 overexpression in T-cell acute lymphoblastic leukemia is essential for proliferation and survival by preventing excessive replication stress.

Checkpoint kinase 1 (CHK1) is a key component of the ATR (ataxia telangiectasia-mutated and Rad3-related)-dependent DNA damage response pathway that protect cells from replication stress, a cell intrinsic phenomenon enhanced by oncogenic transformation. Here, we show that CHK1 is overexpressed and hyperactivated in T-cell acute lymphoblastic leukemia (T-ALL). CHEK1 mRNA is highly abundant in patients of the proliferative T-ALL subgroup and leukemia cells exhibit constitutively elevated levels of the replication stress marker phospho-RPA32 and the DNA damage marker γH2AX. Importantly, pharmacologic inhibition of CHK1 using PF-004777736 or CHK1 short hairpin RNA-mediated silencing impairs T-ALL cell proliferation and viability. CHK1 inactivation results in the accumulation of cells with incompletely replicated DNA, ensuing DNA damage, ATM/CHK2 activation and subsequent ATM- and caspase-3-dependent apoptosis. In contrast to normal thymocytes, primary T-ALL cells are sensitive to therapeutic doses of PF-004777736, even in the presence of stromal or interleukin-7 survival signals. Moreover, CHK1 inhibition significantly delays in vivo growth of xenotransplanted T-ALL tumors. We conclude that CHK1 is critical for T-ALL proliferation and viability by downmodulating replication stress and preventing ATM/caspase-3-dependent cell death. Pharmacologic inhibition of CHK1 may be a promising therapeutic alternative for T-ALL treatment.

BK virus encephalitis with thrombotic microangiopathy in an allogeneic hematopoietic stem cell transplant recipient.

BK virus (BKV) infection occurs most often in immunocompromised hosts, in the setting of renal or bone marrow transplantation. Hemorrhagic cystitis is the commonest manifestation but in recent years infections in other organ systems have been reported. We report an unusual case of biopsy-proven BKV encephalitis in a hematopoietic stem cell transplant patient who subsequently developed thrombotic microangiopathy. As far as we know, this is the first report of such an association in a transplant patient.

La electromiografía en el diagnóstico y tratamiento odontológico / Electromyography in dental diagnosis and treatment

La electromiografía (EMG) permite registrar datos de la actividad eléctrica de los músculos en forma certera, reproducible y objetiva. En la actualidad, la actividad muscular puede estudiarse asociada a la función mandibular y la oclusión dentaria La actividad muscular se estudia no sólo en función dela fuerza de contracción, sino también en función de la frecuencia de contracciones. Esta última es quizá lavaloración más importante ya que un músculo fatigado, con menor frecuencia de contracciones producto de una inadecuada irrigación, déficit de energía y acumulación de productos del catabolismo celular, disminuye su capacidad de rendimiento, predisponiendo al paciente al dolor y disfunción miofascial. Estos registros pueden efectuarse tanto con el músculoen reposo como en actividad. En la práctica profesional odontológica la rehabilitación oral es un desafío permanente, rehabilitar la oclusiónde un paciente con músculos en un estado de hipertonicidad o de fatiga, perpetúa la patología existente.La EMG añade una nueva dimensión al tratamiento, tanto de los pacientes odontológicos sintomáticos comoasintomáticos, facilitando al odontólogo la capacidad de garantizar resultados previsibles y fisiológicos.

Congenital amegakaryocytic thrombocytopenia--report of a new c-mpl gene missense mutation.

A 44-month old girl with congenital amegakaryocytic thrombocytopenia, already with pancytopenia, underwent an unrelated allogeneic cord blood transplantation with recovery of normal blood cell counts. The patient was a compound heterozygote for two c-mpl missense mutations inherited from both parents, one of them, a G578A exon 4 mutation leading to a cysteine to tyrosine replacement of codon 193, previously unreported.

Ablation of pulmonary vein foci for the treatment of atrial fibrillation; percutaneous electroanatomical guided approach.

AIMS: To evaluate the usefulness of three-dimensional (3D) electroanatomical mapping of the pulmonary veins (PV) for guiding radiofrequency (RF) ablation of focal atrial fibrillation (AF) in a single session and to correlate the electrophysiological results with the six month clinical outcome. METHODS AND RESULTS: Sixteen consecutive patients with idiopathic paroxysmal AF (more than 1 episode/month) were studied. A non-fluoroscopic mapping system was used to generate 3D electroanatomic maps of the left atrium and deliver RF energy. In patients with frequent ectopies, mapping was performed using the 'hot-cold' approach (looking for the earliest electrogram in the 3D reconstruction). In patients with infrequent/no ectopies, double/ multiple potentials recorded at the PV were tagged. Pacing at these sites to test for inducibility of ectopy or atrial fibrillation was used to define PV foci. The therapeutic endpoint was defined as suppression of premature beats, dissociation of PV potentials and inability to induce AF. Twenty-five foci were identified (multiple foci in 38%). In the 4 pts with frequent ectopies, Group A, these were suppressed by 4 +/- 4.7 applications. In the 12 pts with infrequent/no ectopies, Group B, an average 4.7 +/- 1.8 applications were delivered per focus; the endpoint was achieved in eight of the patients (13 of 21 foci). By 180 days follow-up, 11 patients were free of symptoms and in sinus rhythm, two had paroxysmal AF episodes and 3 have symptomatic ectopies and are receiving antiarrhythmic drugs. The overall success rate at six months was thus 69%, 100% for group A and 58% for group B. CONCLUSION: Electroanatomic guided RF ablation of paroxysmal AF was highly successful in patients with frequent ectopies. The use of electroanatomical mapping for precise anatomical localization of multiple potentials and for guiding the PV ostia isolation allowed successful RF ablation in 50% of pts with infrequent/no ectopies.

Follow-up of anti-Aspergillus IgG and IgA antibodies in bone marrow transplanted patients with invasive aspergillosis.

A total of 89 patients at risk for, or with invasive aspergillosis (IA) were recruited from bone marrow transplantation (BMT) units in two Lisbon hospitals, and followed for 2(1/2) years to monitor their immune response. Of these patients, six developed probable IA, from which five died. The presence of serum IgG or IgA antibodies against seven Aspergillus recombinant antigens was assessed in patients with IA, using an enzyme-linked immunosorbent assay (ELISA). In parallel, the serum levels of galactomannan (GM) were also monitored, using the Platelia Aspergillus kit (Sanofi Pasteur, Marnes-la-Coquette, France). Superoxide dismutase (Sod) and 94 kDa were the most immunogenic antigens for IgA, while the IgG pattern of recognition changed from patient to patient. From our results we conclude that although follow-up of antibodies against these antigens should not be used as a diagnostic method, patients with IA do produce an immune response that may influence disease outcome.

Factors associated with outcome after cord blood transplantation in children with acute leukemia. Eurocord-Cord Blood Transplant Group.

We have analyzed factors influencing the outcome of 102 children with acute leukemia given a cord blood transplantation (CBT) and reported to the Eurocord Registry. Seventy patients with acute lymphoblastic and 32 with acute myeloid leukemia were given either a related (n = 42) or an unrelated (n = 60) CBT. Children given CBT during first or second complete remission were considered as belonging to the good-risk group (n = 66), whereas those who received a transplant in a more advanced stage of disease were assigned to the poor-risk group (n = 36). In the related group (RCBT), 12 of 42 patients received transplantation from an HLA-disparate donor, whereas in the unrelated group (UCBT) 54 of 60 received an HLA mismatched CBT. Kaplan-Meier estimates for neutrophil recovery at day 60 were 84% +/- 7% in RCBT and 79 +/- 6% in UCBT (P =.16). In multivariate analysis, the most important factor influencing neutrophil engraftment in UCBT was a nucleated cell dose infused greater than 3.7 x 10(7)/kg (P =.05, relative risk [RR] of 1.85, 95% confidence interval [CI]: 0.98-3.4). The incidence of grade II through IV acute graft-versus-host disease was 41% +/- 8% in the RCBT group and 37% +/- 6% in the UCBT group (P =.59). Kaplan-Meier estimates of 2-year event-free survival (EFS) after RCBT or UCBT were 39% +/- 8% and 30% +/- 7%, respectively (P =.19). In multivariate analysis, the most important factor influencing EFS was disease status at time of transplantation: good-risk patients had a 2-year EFS of 49% +/- 7% as compared to 8% +/- 5% in patients with more advanced disease (P =.0003, RR: 0.40, 95% CI: 0.24 to 0. 65). This was a consequence of both an increased 1-year transplant related mortality and a higher 2-year relapse rate in the poor-risk group (65% +/- 9% and 77% +/- 14%, respectively), as compared with good risk patients (34% +/- 6% and 31% +/- 9%, respectively). These data confirm that allogeneic CBT from either a related or an unrelated donor is a feasible procedure able to cure a significant proportion of children with acute leukemia, especially if transplanted in a favorable phase of disease.

Defibrotide as salvage therapy for refractory veno-occlusive disease of the liver complicating allogeneic bone marrow transplantation.

A 30-year-old woman developed veno-occlusive disease of the liver during an allogeneic BMT for acute leukemia. Treatment with recombinant human tissue plasminogen activator and heparin resulted in an incomplete and transient response followed by progressive disease. The patient was then given defibrotide (DF), a mammalian tissue-derived polydeoxyribonucleotide developed for the treatment of a number of vascular disorders, which has thrombolytic and anti-thrombotic properties. No significant bleeding or other major toxicities were observed during treatment and she made a full recovery. At 6 months after the onset of VOD her liver function tests and color flow Doppler ultrasound scan are normal. Our experience supports the preliminary results already obtained with DF. Its efficacy should be evaluated in a prospective randomized fashion.

Heterogeneous spectrum of mutations in the Fanconi anaemia group A gene.

Fanconi anaemia (FA) is a genetically heterogeneous autosomal recessive disorder associated with chromosomal fragility, bone-marrow failure, congenital abnormalities and cancer. The gene for complementation group A (FAA), which accounts for 60-65% of all cases, has been cloned, and is composed of an open reading frame of 4.3 kb, which is distributed among 43 exons. We have investigated the molecular pathology of FA by screening the FAA gene for mutations in a panel of 90 patients identified by the European FA research group, EUFAR. A highly heterogeneous spectrum of mutations was identified, with 31 different mutations being detected in 34 patients. The mutations were scattered throughout the gene, and most are likely to result in the absence of the FAA protein. A surprisingly high frequency of intragenic deletions was detected, which removed between 1 and 30 exons from the gene. Most microdeletions and insertions occurred at homopolymeric tracts or direct repeats within the coding sequence. These features have not been observed in the other FA gene which has been cloned to date (FAC) and may be indicative of a higher mutation rate in FAA. This would explain why FA group A is much more common than the other complementation groups. The heterogeneity of the mutation spectrum and the frequency of intragenic deletions present a considerable challenge for the molecular diagnosis of FA. A scan of the entire coding sequence of the FAA gene may be required to detect the causative mutations, and scanning protocols will have to include methods which will detect the deletions in compound heterozygotes.

Relapsed chronic myeloid leukemia in accelerated phase 10 years after allogeneic bone marrow transplantation: full chimera reconversion with donor peripheral blood stem cells infusion.

We report the case of a 44-year-old male who relapsed in accelerated phase chronic myeloid leukemia 10 years after a successful bone marrow transplantation from his HLA-identical brother, and 3 years after 12 months treatment with interferon-alpha (IFN-alpha) for chronic active hepatitis C (CAH). The patient was infused with G-CSF-primed peripheral blood cells (PBSC) from the original bone marrow donor and a full donor reconstitution, with no detectable molecular disease, was obtained within 4 months without clinical aplasia or GVHD, nor help from other forms of chemotherapy or use of biological response modifiers. We speculate that IFN-alpha for CAH delayed the onset of a clinical recurrence of chronic myeloid leukemia and that in advanced disease PBSCs can provide an advantageous alternative to donor lymphocyte infusion (DLI).

IBMTR Severity Index for grading acute graft-versus-host disease: retrospective comparison with Glucksberg grade.

Acute graft-versus-host disease (GVHD) severity is graded by pattern of organ involvement and clinical performance status using a system introduced by Glucksberg and colleagues 21 years ago. We examined how well Glucksberg grade predicted transplant outcome and constructed a Severity Index not requiring subjective assessment of performance in 2881 adults receiving an HLA-identical sibling T-cell-depleted (n = 752) or non-T-cell-depleted (n = 2129) bone marrow transplant for leukaemia between 1986 and 1992. Relative risks (RR) of relapse, treatment-related mortality (TRM) and treatment failure (TF) (relapse or death) were calculated for patients with (Glucksberg Grade I, II or III/IV acute (GVHD) versus those without acute GVHD and for patients with distinct patterns of organ involvement regardless of Glucksberg grade. Using data for non-T-cell-depleted transplants, a Severity Index was developed grouping patients with patterns of organ involvement associated with similar risks of TRM and TF. Higher Glucksberg grade predicted poorer outcome; however, patients with the same grade but different patterns of skin, liver or gut involvement often had significantly different outcomes. The revised Severity Index groups patients in four categories, A-D. Compared to patients without acute GVHD, RRs (95% confidence interval) of TF were 0.85 (0.69, 1.05) for patients with Index A, 1.21 (1.02, 1.43) with B, 2.19 (1.78, 2.71) with C, and 5.69 (4.57, 7.08) with D. Prognostic utility of the Index was tested in patients receiving T-cell-depleted transplants; similar RRs of TF were observed. An acute GVHD Severity Index is proposed to enhance design and interpretation of clinical trials in the current era of allogeneic blood and bone marrow transplantation.

What are the limits for the Ross operation?

The Ross operation is physiologically the best approach for aortic valve replacement. At the Hospital de Santa Cruz 22 consecutive pulmonary autograft operations have been performed in patients with a mean age of 49 (range 17-65) years. Six patients had mitral valve disease, two had aortic aneurysms and one had a ventricular septal defect. Subcoronary implantation of the autograft was performed in 20 patients. A partial inclusion aortic root replacement technique was used in one and the aortic root was replaced in another. There were no hospital or late deaths. Two patients required autograft replacement at 3 and 9 months postoperatively because of regurgitation. One of these cases was caused by an abnormality of the pulmonary valve and since then echocardiographic assessments of this valve have been performed routinely and have detected significant pulmonary incompetence in four patients who otherwise would have been operated on using the Ross procedure.

Haploidentical cord blood transplant contaminated with maternal T cells in a patient with advanced leukaemia.

Myeloablative treatment followed by lymphohaematopoietic reconstitution with stem cells from umbilical cord blood (UCB) can cure children with leukaemia. The clinical experience of UCB transplantation with HLA 2- and 3-antigen mismatched siblings is rather limited and there are no reports of such patient being given UCB significantly contaminated with maternal T lymphocytes. In this study, we report our experience in treating a child with chronic myeloid leukaemia in blast crisis who was transplanted using UCB cells from mismatched sibling donor containing a significant number of maternal T cells. The patient received 1.17 x 10(8) nucleated cells/kg after conditioning with Ara-C, busulphan, TBI and cyclophosphamide. GVHD prophylaxis was with cyclosporine and an anti-CD25 monoclonal antibody. Although engraftment was somewhat slow it was complete as documented by cytogenetic analysis and DNA studies. Results of minimal residual disease monitoring by RT-PCR for the hybrid BCR/ABL gene showed no evidence of leukaemic mRNA post-transplant. Acute GVHD, skin only, developed on day +14 but promptly responded to low-dose steroids. The technique used for UCB collection may have cell contamination found. In spite of these potential disadvantages: advanced disease, HLA antigen disparate donor and significant maternal T cell contamination, the transplant was successful and at a follow-up of 14 months the child is well with no evidence of chronic GVHD. Immune naivety of cord blood and lack of immunological reactivity of maternal T cells in this context may have played a significant role in the outcome of this case.

The large septal arteries in normal hearts, in aortic valve disease, and in tetralogy of Fallot.

BACKGROUND: Several surgical techniques such as the Ross operation or total correction of tetralogy of Fallot require incisions of the upper ventricular septum. Very few reports on the anatomy of the septal arteries of the pathologic heart can be found in the literature. To get a more precise knowledge of the large septal arteries in pathologic hearts, we have compared the anatomy of normal hearts with that of hearts with aortic valve disease and of tetralogy of Fallot. METHODS: Twenty-six normal heart specimens (group A), 11 with aortic valve disease (group B), and 4 with tetralogy of Fallot (group C) were dissected. RESULTS: In groups B and C a single large septal artery was always found. The large septal artery had the orientation previously described for normal hearts. Still, its course in the lower border of the anterior extension of the septomarginal trabecula was deeper. The anterior extension of the septomarginal trabecula was 4 +/- 3 mm deep in group A, 6 +/- 2 mm in group B, and 3 mm in group C. The interventricular septum was much thicker in groups B and C than in group A. CONCLUSIONS: The position of the large septal artery can be predicted from coronary angiography and from the morphology of the anterior extension of the septomarginal trabecula. Knowledge of its position can improve the safety of operations performed on the outflow of the interventricular septum.

Atrioventricular valve repair using externally adjustable flexible rings.

Initial results obtained with a new flexible ring, adjustable from outside of the heart after interruption of extracorporeal circulation, are presented. Twenty-five rings have been inserted in 20 patients, 14 in the mitral position and 11 in the tricuspid position. In 8 of the 14 patients receiving mitral annuloplasty, other standard mitral valve repair techniques were used. Adjustment, assisted by intraoperative transesophageal color Doppler echocardiography, was done for 10 (71%) of the mitral rings and for 8 (73%) of the tricuspid rings. Residual mitral regurgitation disappeared or became minimal in 9 (90%) patients, and a mild regurgitation remained in 1. In all patients who received tricuspid rings regurgitation was abolished after the adjustment. There was no hospital or late mortality. After a maximum follow-up of 6 months results are comparable in the tricuspid and mitral positions and echocardiographic evaluation revealed stable competent valves in all patients but one, who underwent reoperation because of failure of a mitral valve chordae shortening procedure. The use of externally adjustable rings for the mitral and tricuspid valves is a safe alternative for atrioventricular valve annuloplasty and has the additional advantage of reducing postrepair regurgitation.