Sandra Bucciantini

Human striatal neuroblasts develop and build a striatal-like structure into the brain of Huntington's disease patients after transplantation.

Rebuilding brain structure and neural circuitries by transplantation of fetal tissue is a strategy to repair the damaged nervous system and is currently being investigated using striatal primordium in Huntingtons disease (HD) patients. Four HD patients underwent bilateral transplantation with human fetal striatal tissues (9-12 week gestation). Small blocks of whole ganglionic eminencies were processed to obtain cell suspension and then stereotactically grafted in the caudate head and in the putamen. Follow-up period ranged between 18 and 34 months (mean, 24.7 months). Surgery was uneventful. Starting from the fourth month after grafting, neo-generation of metabolically active tissue with striatal-like MRI features was observed in 6 out of 8 grafts. The increase in D2 receptor binding suggested striatal differentiation of the neo-generated tissue in 3 patients. New tissue, connecting the developing grafts with the frontal cortex and, in one case, with the ventral striatum, was also observed. The new tissue growth halted after the ninth month post transplantation. All patients showed stabilization or improvement in some neurological indices. No clinical and imaging signs, suggestive of graft uncontrolled growth, were seen. This study provides the first evidence in humans that neuroblasts of a striatal primordium can develop and move into the brain after neurotransplantation. Primordium development resulted in the building of a new structure with the same imaging features as the corresponding mature structure, combined with short- and long-distance targeted migration of neuroblasts. The results of this study support both the reconstructive potential of fetal tissue and the remarkably retained plasticity of adult brain. Further studies are necessary to assess the clinical efficacy of the human fetal striatal transplantation.

Fetal striatal grafting slows motor and cognitive decline of Huntington's disease

Objective To assess the clinical effect of caudate-putaminal transplantation of fetal striatal tissue in Huntingtons disease (HD). Methods We carried out a follow-up study on 10 HD transplanted patients and 16 HD not-transplanted patients. All patients were evaluated with the Unified HD Rating Scale (UHDRS) whose change in motor, cognitive, behavioural and functional capacity total scores were considered as outcome measures. Grafted patients also received morphological and molecular neuroimaging. Results Patients were followed-up from disease onset for a total of 309.3 person-years (minimum 5.3, median 11.2 years, maximum 21.6 years). UHDRS scores have been available since 2004 (median time of 5.7 years since onset, minimum zero, maximum 17.2 years). Median post-transplantation follow-up was 4.3 years, minimum 2.8, maximum 5.1 years. Adjusted post-transplantation motor score deterioration rate was reduced compared to the pretransplantation period, and to that of not-transplanted patients by 0.9 unit/years (95% CI 0.2 to 1.6). Cognitive score deterioration was reduced of 2.7 unit/years (95% CI 0.1 to 5.3). For grafted patients the 2-year post-transplantation [18F]fluorodeoxyglucose positron emission tomography (PET) showed striatal/cortical metabolic increase compared to the presurgical evaluation; 4-year post-transplantation PET values were slightly decreased, but remained higher than preoperatively. [123I]iodobenzamide single photon emission CT demonstrated an increase in striatal D2-receptor density during postgrafting follow-up. Conclusions Grafted patients experienced a milder clinical course with less pronounced motor/cognitive decline and associated brain metabolism improvement. Life-time follow-up may ultimately clarify whether transplantation permanently modifies the natural course of the disease, allowing longer sojourn time at less severe clinical stage, and improvement of overall survival.

Fetal cells in a transcervical cell sample collected at 5 weeks of gestation.

Transcervical cell (TCC) sampling is being investigated as a promising method for obtaining fetal cells for prenatal genetic diagnosis. The present case report describes the identification of fetal cells by both fluorescent in situ hybridisation (FISH) and quantitative fluorescent polymerase chain reaction (QF-PCR) analyses in a TCC sample collected by intrauterine lavage at 5 + 0 weeks. This finding underscores the possible relevance of TCC sampling for extremely early prenatal genetic diagnosis.

Use of the Quantitative Fluorescent-PCR Assay in the Study of Fetal DNA from Micromanipulated Transcervical Samples

AbstractAim: The purpose of this study was to evaluate the validity of the combined use of micromanipulation and quantitative fluorescent (QF)-PCR assay for the identification of fetal elements in transcervical cell (TCC) samples collected in early pregnancy. Methods: TCC samples were obtained by intrauterine lavage (IUL) in 113 pregnant women who were between 7 and 12 weeks pregnant before termination of pregnancy. All IUL samples were screened under an inverted microscope, at which time the isolation of fetal cells by micromanipulation was attempted. QF-PCR assay, using 9 small tandem repeat (STR) markers for chromosomes 13, 18, 21, X, and Y, was performed in all specimens to identify fetal cells in TCC samples and the corresponding placental tissue and blood specimens. TCC samples from male fetuses in which either the micromanipulation or QF-PCR analysis were unsuccessful, were studied with fluorescent in situ hybridization (FISH), using probes for X and Y chromosomes. Results: Isolation of supposed fetal material from IUL samples was carried out by means of micromanipulation in 93 cases (82.3%), where discernible chorionic villous filaments or cell clumps of probable trophoblastic origin were present. The QF-PCR analysis was performed in all 93 IUL samples and paternal peaks could be documented in 88 cases (94.6%) thus confirming the presence of fetal cells. Thirteen cases negative to micromanipulation and derived from male fetuses and four male cases not informative with QF-PCR analysis, after micromanipulation, were then tested with FISH assay using probes for sexual chromosomes. In six samples, rare (2–3%) male fetal cells were detected. Considering the combined results obtained from QF-PCR and FISH assays, the overall fetal sexing was correct in 83.2% of cases (94 of 113). Conclusion: This study provides evidence that fetal cells are present in a high proportion of IUL samples. Micromanipulation appears to be an extremely efficient method for the isolation of trophoblastic elements. This study also confirms the potential of IUL as a possible alternative to the traditional prenatal diagnostic procedures for the recovery of fetal cells in precocious stage of gestation, and validates the combination of the isolation of such fetal elements by means of micromanipulation and analysis with the QF-PCR assay for the identification of the most frequent prenatal chromosomal aneuploidies.

Premature ovarian insufficiency in young girls: repercussions on uterine volume and bone mineral density.

Abstract Study objective: To evaluate biological differences among young subjects with premature ovarian insufficiency (POI) commencing at different stages of life. Design: Retrospective observational study. Setting: Careggi University Hospital Participants: One hundred sixty-two females aged between 15 and 29 years with premature ovarian insufficiency. Methods: Data were collected as a retrospective chart review of baseline evaluation at diagnosis of premature ovarian insufficiency (POI). About 162 participants were divided into four groups based on gynecological age. Two primary outcome variables (uterine development and bone mineral density (BMD)) were analyzed in terms of differences among groups and in a multivariate logistic regression analysis. Results: Uterine development was clearly jeopardized when estrogen insufficiency started at a very young age. Total body BMD showed significant differences among the four groups studied, clearly corresponding to the duration of ovarian function. Data were discussed in relation to the choice of hormone replacement therapy regimens. Chinese abstract 评估年轻患者不同阶段发生卵巢功能不全（POI）的生物学差异。回顾性观察研究。设置：Careggi大学医院。对象：162名15岁～29岁卵巢功能不全女性。对诊断为卵巢功能不全（POI）的基线数据进行了回顾性分析。162名参与者根据妇科年龄分为四组。分析不同组的两个主要指标（子宫发育和骨密度（BMD）），并进行多因素logistic回归分析。雌激素不足开始于非常年轻的年龄时子宫发育明显受影响。不同四组的总BMD有显著差异，与卵巢功能的持续时间有明确的相关性。数据讨论了相关的激素治疗方案的选择。

Donor-Specific Anti-HLA Antibodies in Huntington's Disease Recipients of Human Fetal Striatal Grafts.

Fetal grafting in a human diseased brain was thought to be less immunogenic than other solid organ transplants, hence the minor impact on the efficacy of the transplant. How much prophylactic immune protection is required for neural allotransplantation is also debated. High-sensitive anti-HLA antibody screening in this field has never been reported. Sixteen patients with Huntingtons disease underwent human fetal striatal transplantation in the frame of an open-label observational trial, which is being carried out at Florence University. All patients had both brain hemispheres grafted in two separate robotic-stereotactic procedures. The trial started in February 2006 with the first graft to the first patient (R1). R16 was given his second graft on March 2011. All patients received triple immunosuppressive treatment. Pre- and posttransplant sera were analyzed for the presence of anti-HLA antibodies using the multiplexed microsphere-based suspension array Luminex xMAP technology. Median follow-up was 38.5 months (range 13-85). Six patients developed anti-HLA antibodies, which turned out to be donor specific. Alloimmunization occurred in a time window of 0–49 months after the first neurosurgical procedure. The immunogenic determinants were non-self-epitopes from mismatched HLA antigens. These determinants were both public epitopes shared by two or more HLA molecules and private epitopes unique to individual HLA molecules. One patient had non-donor-specific anti-HLA antibodies in her pretransplant serum sample, possibly due to previous sensitization events. Although the clinical significance of donor-specific antibodies is far from being established, particularly in the setting of neuronal transplantation, these findings underline the need of careful pre- and posttransplant immunogenetic evaluation of patients with intracerebral grafts.

Young Human Cholinergic Neurons Respond to Physiological Regulators and Improve Cognitive Symptoms in an Animal Model of Alzheimer’s Disease

The degeneration of cholinergic neurons of the nucleus basalis of Meynert (NBM) in the basal forebrain (BF) is associated to the cognitive decline of Alzheimer’s disease (AD) patients. To date no resolutive therapies exist. Cell-based replacement therapy is a strategy currently under consideration, although the mechanisms underlying the generation of stem cell-derived NBM cholinergic neurons able of functional integration remain to be clarified. Since fetal brain is an optimal source of neuronal cells committed towards a specific phenotype, this study is aimed at isolating cholinergic neurons from the human fetal NBM (hfNBMs) in order to study their phenotypic, maturational and functional properties. Extensive characterization confirmed the cholinergic identity of hfNBMs, including positivity for specific markers (such as choline acetyltransferase) and acetylcholine (Ach) release. Electrophysiological measurements provided the functional validation of hfNBM cells, which exhibited the activation of peculiar sodium (INa) and potassium (IK) currents, as well as the presence of functional cholinergic receptors. Accordingly, hfNBMs express both nicotinic and muscarinic receptors, which were activated by Ach. The hfNBMs cholinergic phenotype was regulated by the nerve growth factor (NGF), through the activation of the high-affinity NGF receptor TrkA, as well as by 17-β-estradiol through a peculiar recruitment of its own receptors. When intravenously administered in NBM-lesioned rats, hfNBMs determined a significant improvement in memory functions. Histological examination of brain sections showed that hfNBMs (labeled with PKH26 fluorescent dye prior to administration) reached the damaged brain areas. The study provides a useful model to study the ontogenetic mechanisms regulating the development and maintenance of the human brain cholinergic system and to assess new lines of research, including disease modeling, drug discovery and cell-based therapy for AD.

The presence of trophoblastic cells in intrauterine lavage samples : lack of correlation with maternal and obstetric characteristics

To investigate the correlation between maternal, obstetric and sample characteristics and the quality (i.e. yield of trophoblastic cells) of intrauterine lavage (IUL) samples.

From Primary Hypergonadotropic Amenorrhea to “POI”: Aetiology and Therapy

Premature ovarian damage with onset before menarche is expressed clinically with primary hypergonadotropic amenorrhea. The term premature ovarian insufficiency (POI) currently used in the literature worldwide substitutes the previously used POF (premature ovarian failure). The very concept of POI refers to the many, varied clinical expressions of ovarian insufficiency, including the possible albeit usually transitory recuperation of ovarian function.