RHCE variant alleles and risk of alloimmunization in Brazilians.

Variant RHCE alleles are found mainly in Afro-descendant individuals, as well as in patients with sickle cell disease (SCD). The most common variants are related to the RHCE*ce allele, which can generate partial e and c antigens. Although RHCE variant alleles have been extensively studied, defining their clinical significance is a difficult task. We evaluated the risk of RhCE alloimmunization as a consequence of partial antigens in patients with a positive phenotype transfused with red blood cell (RBC) units with the corresponding antigen. A retrospective study was performed with Brazilian patients, evaluating the number of antigen-positive transfused RBC units (incompatible due to partial antigen) in 27 patients with SCD carrying RHCE variant alleles who did not develop antibodies as well as evaluating the variants present in 12 patients with partial phenotype and correlated antibody (one patient with SCD and 11 patients with other pathologies). Two patients showed variant alleles with molecular changes that had not yet been described. Variant RHCE alleles were identified in a previous study using molecular methods. RHCE*ceVS.01 was the most frequent allele found among the patients without antibodies. Six patients with partial c antigen had a mean of 3.8 c+ RBC units transfused, and 10 patients with partial e antigen were exposed for a mean of 7.2 e+ RBC units. Among the variant alleles found in alloimmunized patients, the most frequent was RHCE*ceAR, which was found in five patients; the antibodies developed were anti-hrS and/or anti-c. Our results showed that RHCE*ceVS.01 is indeed the most frequent variant allele in our cohort of patients with SCD, but the partial antigens that were identified have low risk of alloimmunization. RHCE*ceAR is the most impactful variant in the Brazilian population with high risk of alloimmunization and clinically significant anti-hrS formation.

RHCE variants inherited with altered RHD alleles in Brazilian blood donors.

BACKGROUND: The high homology and opposite orientation of RH genes promote rearrangements between them and generate a large number of RHD and RHCE variants which can be inherited together. Searching of RHD-CE genotypes predicting partial antigens in donors is of interest in order to find more closely matched donors for African descent patients. This study aimed to evaluate a molecular approach to search for RhCE variants in a cohort of individuals with altered expression of D antigen and determine the association of RH variant alleles in Brazilian blood donors. METHODS: From 80,961 blood samples tested, 421 with atypical D typing results were studied. The samples were phenotyped for C, c, E, e antigens. Rh variants were identified using molecular techniques. RESULTS: All 421 samples had altered RHD alleles, being 56·3% of them partial D. Among them, 94·9% presented variant RHCE*ce and the most common associations were: RHD*weak D type 4.2.2 with RHCE*ceAR; RHD*DAR linked to RHCE*ceVS.02; RHD*weak D type 4.0 linked to RHCE*ceVS.02 and RHCE*ce (c.48C, c.105T, c.733G, c.744C, c.1025T). Among the samples with RhCE variants, 10·6% predict partial c, partial e, hr(B) - and/or hr(S) - and 100% express low prevalence antigens. CONCLUSION: Targeting individuals with altered expression of D antigen can be a good strategy for finding donors with RhCE variants. In our study 94·9% of the partial D samples revealed altered RHCE variant alleles and 5·7% of the samples with altered RHD allele predicted partial c, partial e and the lack of the high prevalence hr(B) and hr(S) antigens.

An overview of the use of SNaPshot for predicting blood group antigens.

The use of SNaPshot (Applied Biosystems, Foster City, CA) for predicting blood group antigens has emerged as an alternative to hemagglutination testing and also to the current low- and highthroughput blood group genotyping methods. Several groups have developed multiplex-polymerase chain reaction SNaPshot assays to determine single nucleotide polymorphisms (SNPs) in blood group genes with the purpose of identifying clinically relevant antigens and rare alleles. The selection of SNPs is based on the population or laboratory reality and the purpose of the genatyping. Unlike high-throughput genotyping strategies that are provided as commercial platforms, the SNPs can be chosen to best meet the needs of the user, and the interpretation of the results do not depend on the manufacturer.

Five-year follow-up of 10 patients treated with globus pallidus internus deep brain stimulation for segmental or multisegmental dystonia.

INTRODUCTION: Globus pallidus internus (GPi) deep brain stimulation (DBS) represents a validated, effective, and safe treatment for patients affected by generalized dystonia resistant to conservative treatment. Segmental and multisegmental dystonia have more recently been proposed as further indications for GPi DBS despite the lack of long-term homogenous follow-up. Here we present an original and detailed long-term follow-up (5 years) of a homogeneous population of 11 patients affected by segmental or multisegmental dystonia. MATERIALS AND METHODS: Ten patients underwent bilateral GPi DBS electrode implantations under a Leksell stereotactic guide, with intraoperative neurophysiological monitoring. The follow-ups at 1, 3 and 5 years were collected using video-BFMDRS for motor and disability scores. The statistical analysis of the results is provided. RESULTS: We reported a statistically significant improvement in motor and disability overall scores until 5 years after treatment. At the last follow-up, even the single motor subitems were statistically improved. DISCUSSION: We observed a continuous and statistically significant improvement in all of the motor subitems and in the overall disability score until the 3-year follow-up. These results did not improve any further but they appeared steady at the last follow-up. We also report a significant improvement in the cranial-cervical subitems. CONCLUSIONS: GPi DBS should definitely be considered a safe and effective treatment also for segmental and multisegmental dystonia even in cases of relevant or prevalent cranial-cervical involvement.

Spinal anesthesia and minimal invasive laminotomy for paddle electrode placement in spinal cord stimulation: technical report and clinical results at long-term followup.

OBJECT: We arranged a mini-invasive surgical approach for implantation of paddle electrodes for SCS under spinal anesthesia obtaining the best paddle electrode placement and minimizing patients' discomfort. We describe our technique supported by neurophysiological intraoperative monitoring and clinical results. METHODS: 16 patients, affected by neuropathic pain underwent the implantation of paddle electrodes for spinal cord stimulation in lateral decubitus under spinal anesthesia. The paddle was introduced after flavectomy and each patient confirmed the correct distribution of paresthesias induced by intraoperative test stimulation. VAS and patients' satisfaction rate were recorded during the followup and compared to preoperative values. RESULTS: No patients reported discomfort during the procedure. In all cases, paresthesias coverage of the total painful region was achieved, allowing the best final electrode positioning. At the last followup (mean 36.7 months), 87.5% of the implanted patients had a good rate of satisfaction with a mean VAS score improvement of 70.5%. CONCLUSIONS: Spinal cord stimulation under spinal anesthesia allows an optimal positioning of the paddle electrodes without any discomfort for patients or neurosurgeons. The best intraoperative positioning allows a better postoperative control of pain, avoiding the risk of blind placements of the paddle or further surgery for their replacement.

Awake surgery in low-grade gliomas harboring eloquent areas: 3-year mean follow-up.

Low-grade gliomas are slow-growing tumors invading eloquent areas and white matter pathways. For many decades these tumors were considered inoperable because of their high tropism for eloquent areas. However, the young age of the patients and the inescapable anaplastic transformation have recently suggested more aggressive treatments. We analyzed the neurological and neuro-oncological outcome of 12 patients who underwent surgery fully awake for the resection of LGG, harboring eloquent areas. 10 right- and 2 left-handed patients underwent pre-operative assessment: Karnofsky Performance Status, Edinburgh Handedness Inventory Score; neuropsychological and neurophysiological evaluations, according to the tumor location. During surgery we performed: sensory-motor-evoked potentials, continuous electro-corticography and bipolar/monopolar cortico-subcortical mapping during neuropsychological tests. The resection rate was calculated with neuro-imaging elaboration software. No permanent post-operative deficits were reported; 2 patients improved after surgery. No impairment of cognitive functions was reported. The KPS improved in 8 patients and was steady in the others. The mean resection rate was 78.3%. The resection allowed the control of pre-operative seizures without increasing the drug intake. Awake surgery allowed a good resection rate despite the eloquent location of the tumors, without post-operative deficit. The neuropsychological outcome was unchanged after surgery. The resection seems to improve seizure control. All the patients came back to normal life and work. In conclusion, awake surgery is reliable and feasible in removal of LGG, even if invading the main eloquent areas and networks. All the patients experienced a normal life after surgery, without permanent deficits.

Development of a primary standard for calibration of (64)Cu activity measurement systems.

A (64)Cu national primary standard, was developed by the National Institute for Ionising Radiation Metrology (INMRI) of the ENEA (ENEA-INMRI) using the CIEMAT/NIST method of 4pibeta liquid scintillation spectrometry with (3)H-standard efficiency tracing. Relatively short (64)Cu half-life is required for the work to be performed at the production site. It was produced at the Scanditronix MC40 Cyclotron of the Joint Research Centre (JRC) of the European Commission (Ispra, Italy) through the (64)Zn(d,2p) (64)Cu reaction. Significant efforts were made to identify and quantify the impurities of (61)Cu and (65)Zn in the mother solution, which were activated through the (64)Zn(d,alphan) (61)Cu and (64)Zn(d,p) (65)Zn reactions, respectively. To this purpose, a new procedure for the determination of pure beta-emitter impurities by the CIEMAT/NIST method has been applied. A transfer standard portable well-type ionisation chamber was also calibrated with minimum uncertainty.

Rapid and Accurate MRI Segmentation of Peritumoral Brain Edema in Meningiomas.

PURPOSE: The extent of peritumoral brain edema (PTBE) in meningiomas commonly affects the clinical outcome. Despite its importance, edema volume is usually highly inaccurately approximated to a spheroid shape. We tested the accuracy and the reproducibility of semiautomatic lesion management software for the analysis of PTBE in a homogeneous case series of surgically confirmed intracranial meningiomas. METHODS: PTBE volume was calculated on magnetic resonance images in 50 patients with intracranial meningiomas using commercial lesion management software (Vue PACS Livewire, Carestream, Rochester, NY, USA). Inter and intraobserver agreement evaluation and a comparison between manual volume calculation, the semiautomatic software and spheroid approximation were performed in 22 randomly selected patients. RESULTS: The calculation of edema volume was possible in all cases irrespective of the extent of the signal changes. The median time for each calculation was 3 min. Interobserver and intraobserver agreement confirmed the reproducibility of the method. Comparison with standard (fully manual) calculation confirmed the accuracy of this software. CONCLUSIONS: Our study showed a high level of reproducibility of this semiautomatic computational method for peritumoral brain edema. It is rapid and easy to use after relatively short training and is suitable for implementation in clinical practice.

Erythropoietin and granulocyte-macrophage colony-stimulating factor allow acceleration and dose escalation of cyclophosphamide/epidoxorubicin/5-fluorouracil chemotherapy: a dose-finding study in patients with advanced breast cancer.

To verify whether the association of granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (EPO) would allow both the acceleration and the dose escalation of the cyclophosphamide/epidoxorubicin/5-fluorouracil (CEF) regimen as first-line therapy in advanced breast cancer patients, we conducted a dose-finding study. Cohorts of three consecutive patients received cyclophosphamide (Ctx, dose range 800-1400 mg/m2), epidoxorubicin (Epidx, dose range 70-100 mg/m2), and 5-fluorouracil (5-Fu, 600 mg/m2, fixed dose) given as an intravenous bolus on day 1 every 14 days; GM-CSF at 5 micrograms/kg given as a subcutaneous injection from day 4 to day 11; and EPO at 150 IU/kg given as a subcutaneous injection three times a week. In no single patient was any dose escalation allowed. A total of 14 patients entered the study. At the 4th dose level (Ctx 1400 mg/m2, Epidx 100 mg/m2, 5-Fu 600 mg/m2), two patients had dose-limiting mucositis and one patient developed dose-limiting neutropenia. Therefore, the 3rd cohort received the maximum tolerated dose, i.e. Ctx at 1200 mg/m2, Epidx at 90 mg/m2, and 5-Fu at 600 mg/m2, given every 18.5 (+/-2.5) days. Toxicity was moderate and manageable in an outpatient setting. Only 1 admission at the 4th dose level was required. Throughout the 4 dose levels there was no toxicity-related death; grade IV leukopenia ranged from 24% to 75% of cycles and grade IV thrombocytopenia ranged from 6% to 8%. No grade IV anemia was recorded. Increasing the doses of Ctx and Epidx while maintaining a fixed dose of 5-Fu with the support of both EPO and GM-CSF allows safe acceleration and dose escalation of CEF chemotherapy. Further controlled studies will evaluate the activity and efficacy of this strategy.