Allogenic peripheral stem cell transplantation from HLA-matched related donors for adult sickle cell disease: remarkable outcomes from a single-center trial.

Adult patients with sickle cell disease (SCD) are highly susceptible to stem cell transplant complications, including drug toxicity, graft versus host disease (GVHD), and graft rejection due to SCD-related tissue damage, endothelial activation, and inflammation. The scarcity of compatible stem cells for transplantation further limits treatment options, with only 43 cases of adult allogeneic peripheral blood stem cell transplantation (allo-PSCT) from human leukocyte antigen (HLA)-identical sibling donors reported in the international registry for the period 1986-2013. Herein we report remarkable outcomes in a cohort of adult SCD patients who underwent allo-PSCT using a fludarabine (Flu), busulfan (Bu), and anti-T-cell lymphocyte globulin (ATG)-based conditioning regimen in combination with very low dose total body irradiation (TBI), followed by post-transplant cyclophosphamide (Cy) and sirolimus as GVHD prophylaxis. We performed a single-center, retrospective study consisting of 20 consecutive patients (mean age 33.4 years) who underwent allo-PSCT from HLA-matched related donors with a conditioning regimen of Flu 150/Bu 3.2/Cy 29/ATG 30 (Fresenius)/TBI 200 between September 2013 and September 2017. Data were validated by an independent data audit group of the affiliated JACIE-accredited transplantation center. All patients experienced a sustained donor cell engraftment. Full donor chimerism (total cell) occurred within 180 days in all patients. Mean duration of follow-up was 13.8 months (range: 0.3-50 months), with 12 (60%) patients completing 12 months. No non-relapse mortality or graft rejection occurred. Successful treatment was achieved without the presence of graft loss, grade III-IV acute GVHD, extensive chronic GVHD, or other major complications. Allo-PSCT in combination with Flu 150/Bu 3.2/Cy 29/ATG 30(Fresenius)/TBI 200- Cy/Sirolimus therapy yielded encouraging outcomes with no mortality and low incidence of GVHD. Further controlled studies will be necessary to compare transplant protocols and long-term outcomes.

C-C coupling between trinitrothiophenes and triaminobenzenes: zwitterionic intermediates and new all-conjugated structures.

The reactions of 1,3,5-triaminobenzene derivatives with 2,3,4-trinitrothiophene and 2-bromo-3,4,5-trinitrothiophene gave new all-conjugated compounds bearing both an electron-withdrawing and an electron-donor moiety on the same unit. The reactions with 2,3,4-trinitrothiophene offered evidence, by NMR spectroscopy at low temperature, of the formation of new labile Wheland-Meisenheimer intermediates whereas at room temperature stable unexpected products derived from the attack of the nucleophile at C-4 with replacement of the nitro group were isolated. Their formation caused, in turn, the obtainment of a salt between 1-nitroso-2,4,6-triaminobenzenes and 2,4-dinitrothiophen-3-ol. The reactions with 2-bromo-3,4,5-trinitrothiophene produced in good yields the SNAr substitution product with the displacement of the bromide. All the new coupling products obtained are of applicative interest, considering the increasing concern for highly conjugated π-systems in solar energy conversion or optoelectronic devices.

New electron-donor and -acceptor architectures from benzofurazans and sym-triaminobenzenes: intermediates, products and an unusual nitro group shift.

New all-conjugated C-C coupling products bearing both an electron-poor and an electron-rich aromatic moiety have been obtained from the reaction between sym-triaminobenzene derivatives and a series of isomeric chloro-nitrobenzofurazans. The reactions occur under mild reaction conditions, and in some cases a different behaviour depending on the presence, or not, of triethylamine was observed. From 1,3,5-tris(N-morpholinyl)benzene and 5-chloro-4-nitrobenzofurazan in the presence of triethylamine an unexpected product derived from the shift of the nitro group from C-4 to C-5 of the electrophile and bearing the nucleophile at position 4 was obtained. Moreover, from the coupling between 1,3,5-tris(N-pyrrolidinyl)benzene and 4-chloro-7-nitrobenzofurazan a highly stable Wheland intermediate was isolated.

Effectiveness of fludarabine- and busulfan-based conditioning regimens in patients with acute myeloblastic leukemia: 8-year experience in a single center.

OBJECTIVE: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for acute myeloblastic leukemia (AML). Because the conditioning regimen of busulfan plus cyclophosphamide carries significant risks of toxicity, we evaluated the factors affecting survival after fludarabine replacement instead of cyclophosphamide. METHODS: The study included 55 patients who underwent allo-HSCT for AML and received busulfan, fludarabine, and antithymocyte globulin (ATG). RESULTS: Forty-eight patients received a myeloablative regimen; 7 patients received a reduced-intensity conditioning regimen. The neutrophil and platelet engraftment times were 12 days (range 9 to 20) and 12 days (range 7 to 19), respectively. Graft-vs-host disease (GvHD) developed in 10% and 50% of the patients, respectively. Seven patients received donor lymphocyte infusion. Of them, 5 patients developed grade I or II GvHD, one grade IV GvHD. The median follow-up period was 20.6 months. The predicted progression-free survival (PFS) at 1 and 3 years after transplantation was 78% and 74%, respectively. The overall survival (OS) at 1, 3, and 5 years was 76%, 74%, and 62%, respectively. Treatment-related mortality (infection in 1 patient, GvHD in 2 patients) occurred in 3 patients (5.5%). Multivariate analysis revealed that OS and PFS were not influenced by age, dose of busulfan or ATG, or presence of cytomegalovirus antigenemia. Acute GvHD and pretransplantation minimal residual disease positivity negatively affected the transplant outcome. The presence of active disease at the time of transplantation was found as an independent risk factor for AML. CONCLUSIONS: Busulfan- and fludarabine-based conditioning regimens are effective for AML, and have acceptable toxicity, morbidity, and mortality.

Formaldehyde replacement with glyoxylic acid in semipermanent hair straightening: a new and multidisciplinary investigation.

OBJECTIVE: Formaldehyde is an effective and popular semipermanent hair straightener, but the severe consequences for human health due to its toxicity have prompted the search for safer alternatives. Different carbonyl compounds, including glyoxylic acid, have recently been proposed as promising candidates. Despite the interest in this topic, there is a lack of information about the interactions between hair keratin and straightener agents. This study addresses this issue to gain new insights useful in the development of new products for safe, semipermanent hair deformation. METHODS: The possible reactions occurring between carbonyl groups and nucleophilic sites on amino acid residues belonging to the keratin were investigated using as model compounds some aldehydes and amino acid derivatives. Raman and IR analyses on yak hair subjected to the straightening treatment with glyoxylic acid in different conditions were carried out. Scanning electron microscope (SEM) analyses were carried out on yak and curly human hair after each step of the straightening procedure. RESULTS: The reactions between aldehydes and N-α-acetyl-L-lysine revealed the importance of the carbonyl electrophilicity and temperature to form imines. Raman and IR analyses on yak hair subjected to the straightening treatment evidenced rearrangements in the secondary structure distribution, conformational changes to the disulphide bridges, a decrease of the serine residues and formation of imines. It was also indicated that straightening produced major conformational rearrangements within the hair fibre rather than on the cuticle. CONCLUSION: This investigation revealed the role played by the electrophilicity of the carbonyl on the straightener agent and of the temperature, closely related to the dehydration process. Raman and IR studies indicated the involvement of imine bonds and the occurrence of a sequence of conformational modifications during the straightening procedure. SEM analyses showed the effectiveness of the treatment at the cuticular level.

Alterations of circulating endothelial cells after apheresis in patients with sickle cell disease: a potential clue for restoration of pathophysiology.

OBJECTIVES: The potential influence of automated red cell exchange (ARCE) on endothelial activation is not well established. This study was intended to assess whether ARCE influences circulating endothelial cells (CECs) in patients with sickle cell disease. BACKGROUND: Automated red cell exchange (ARCE) has been used to protect the patient from complications of sickle cell disease. However, the expected benefits vary in different patients. CECs reflect endothelial activation. We hypothesize that suppression of endothelial activation may be an important mechanism of ARCE. METHODS: The study included 20 patients with sickle cell disease who underwent 30 apheresis procedures. We used flow cytometry to directly compare pre- and post-apheresis CEC number (prior to ARCE and 5 days after ARCE) during the steady state and painful crisis. We also determined if independent variables (the level of plasma nitrite concentration, the percentage of circulating hemoglobin S, and painful crisis) significantly contributed to the CEC level. RESULTS: The mean CEC number decreased (P = 0.04), while progenitor CECs did not change in patients with sickle cell disease after ARCE compared with pre-ARCE values (P>0.05). Clinical factors such as the volume of replacement fluid and the citrate infusion rate did not correlate with post-apheresis CECsand progenitor CEC numbers. The independent variables were not significantly associated with CEC and progenitor CEC numbers. CONCLUSIONS: ARCE can alter the CEC number, suggesting the possibility of suppression of endothelial activation. This may highlight the efficacy of ARCE for prevention or management of sickle cell vaso-occlusive crisis.

A detachment technique based on the thermophysiologic responses of cultured mesenchymal cells exposed to cold.

Trypsinization has generally been used as a technique to detach adherent mesenchymal stromal cells (MSC). However, this technique involves chemical manipulation. This study was designed to identify whether detachment of MSC can be induced by cold without using trypsin. MSC isolated from bone marrow were detached via trypsin or exposed to -20 degrees C for 1, 5 or 10 min at all passages. Compared with trypsinization, exposing MSC to -20 degrees C for 10 min resulted in a significant decrease in MSC number and viability. In conclusion, although detachment of adhered MSC on culture dishes via exposure to cold may allow structurally and functionally intact detached cells, the technique requires improvement of the thermotolerance of MSC.

Assessment of corrected QT interval in sickle-cell disease patients who undergo erythroapheresis.

Extension of the QT interval is characterized by syncope and cardiac arrest and often occurs in association with medical therapies and procedures. Whether erythroapheresis (EPH) could influence the QT interval duration in patients with sickle cell disease (SCD) is not known. We aimed to investigate the effects of EPH on the heart rate-corrected QT (QTc) interval. The study included 25 patients with SCD who underwent 34 EPH procedures. Two independent observers measured QTc interval duration from electrocardiograms performed continuously for 3 min at three different points during the EPH procedures (prior to EPH, after completion of 50% EPH and 15 min after EPH). Multiple regression analysis was used to determine if the ionized plasma calcium, the level of plasma magnesium, citrate infusion rate and painful crisis significantly contributed to the QTc interval. There was a non-significant trend (P = 0.184) towards increased QTc in sickle cell patients during EPH compared with pre-EPH values. QTc prolongation (>440 ms) occurred in 72% of the procedures. Fifty percent QTc values returned to baseline after the procedure. The independent variables were not significantly associated with QTc interval. Exchange procedures can induce QTc prolongation in patients with SCD.

Cytogenetic findings and clinical outcomes of adult acute myeloid leukaemia patients.

The role of cytogenetic findings in determining the diagnosis, therapy and prognosis of acute myeloid leukaemia (AML) has become more valuable by the day. In this study, the results of conventional and molecular cytogenetic analyses and clinical outcomes of 66 AML patients of different subgroups aged between 16 and 82 were evaluated. Chromosomal abnormalities were detected in 17 (25.7%) patients cytogenetically at the time of diagnosis, whereas molecular cytogenetic abnormalities were detected in 21 (31.8%) patients by fluorescence in situ hybridisation (FISH). Thirty-eight (57.6%) patients had a normal karyotype. In 8 patients, we did not obtain suitable chromosomes for karyotype analysis and in 3 patients conventional cytogenetics were not requested by the physician. During clinical follow-up, 21 patients (31.8%) achieved complete remission (CR), 2 had partial remission (PR) (3.0%) and 4 patients had progressive disease (6.06%). Twenty-eight (42.4%) patients died during treatment and no follow-up data were available in 7 cases. Among patients with chromosome abnormalities detected by either one of the two cytogenetic methods (n=28), 12 had achieved CR, 9 of whom were already categorised in the good prognostic group with t(15;17), inv16 or t(8;21). As for the normal karyotype, each patient displayed a different clinical course, which is probably due to the molecular changes in leukaemia-related genes. Here we report our findings, which correlate with previous reports and conclude that cytogenetics is a crucial marker in leukaemia diagnosis and conventional and molecular cytogenetics should be performed as well as molecular genetic diagnostic methods.

Plasma exchange in critically ill patients with sickle cell disease.

Red cell exchange transfusion is the recommended therapy for patients with sickle cell disease who have complicated vaso-occlusive episodes. However, the role of the therapeutic plasma exchange in the management of the potentially life-threatening complications in patients with sickle cell disease is not well known. To determine whether plasma exchange had a cumulative effect on the red cell exchange in patients with sickle cell disease who developed multi-organ failure during the post red cell exchange period, we performed plasma exchange in the nine episodes of multi-organ failure of 7 patients with sickle cell anemia. The median age of those patients was 21 years (range, 9-50 years). The criterion of the multi-organ failure was defined as organ failure of two or more organs i.e. lung, liver, or renal, established according to Acute Physiological and Chronic Health Evaluation-II (APACHE-II) criteria. The average total plasma exchange volume was 1.0 times the patient's plasma volume. The patients had a good outcome, with a survival rate at 86% after one year of follow-up. Plasma exchange may have cumulative benefits in the treatment of severe illness in patients with sickle cell disease who underwent automatic red cell exchange therapy.

The first 2 years of clinical experience with peripheral blood stem cell transplantation for various hematological malignancies: results from a single Baskent University Center.

Autologous stem cell transplantation is the current standard approach for patients with multiple myeloma and relapsed or refractory lymphoma. Nonmyeloablative allogeneic stem cell transplantation has been applied worldwide. We analyzed the results of transplantation activity from 2004 to 2006. Seven evaluable patients younger than 65 years old with stage II/III multiple myeloma were treated with high-dose melphalan therapy (140 mg/m(2)) plus autologous peripheral blood stem cell transplantation. Complete responses or tumor reductions of more than 75% were obtained in all patients. At a median follow-up of 10 months, all patients remained disease-free. Four patients with acute myeloblastic leukemia underwent nonmyeloablative allogeneic peripheral stem cell transplantation. Their median age was 30 years. One patient was refractory and the others were in hematological remission. The patients received fludarabine-based preparative regimens. All patients received fully matched blood from a related donor 2 days after chemotherapy in conjunction with graft-versus-host disease prophylaxis. One refractory patient with >90% engraftment had late autologous reconstitution at 3 months with evidence of relapse. All other patients in remission remained with >90% donor cell engraftment. These patients are disease-free at 13, 10, and 2 months. Toxicity was minimal. These results showed promise due to the minimal toxicity observed with the conditioning regimens which indicated the feasibility of these procedures.

Modulation of apoptotic signalling by 9-hydroxystearic acid in osteosarcoma cells.

9-hydroxystearic acid (9-HSA) belongs to the class of endogenous lipid peroxidation by-products that greatly diminish in tumors, causing as a consequence the loss of one of the control mechanisms on cell division. We have previously shown that 9-HSA controls cell growth and differentiation by inhibiting histone deacetylase 1 (HDAC1) activity. In this paper our attention has not only been focused on HDAC1 inhibition but also on the hyperacetylation of other substrates such as p53, that is involved in inducing cell cycle arrest and/or apoptosis, and whose activity and stability are known to be regulated by posttranslational modifications, particularly by acetylation at the C-terminus region. 9-HSA administration to U2OS, an osteosarcoma cell line p53 wt, induces a growth arrest of the cells in G2/M and apoptosis via a mitochondrial pathway. In particular hyperacetylation of p53 induced by the HDAC1 inhibitory activity of 9-HSA has been demonstrated to increase Bax synthesis both at the transcriptional and the translational level. The subsequent translocation of Bax to the mitochondria is associated to a significant increase in caspase 9 activity. Our data demonstrate that the effects of 9-HSA on U2OS correlate with posttranslational modifications of p53.

Conventional and molecular cytogenetic findings of myelodysplastic syndrome patients.

Myelodysplastic syndrome (MDS) involves myeloid cells of the bone marrow, which is important in progressive bone marrow insufficiency. Of all MDS patients, 40%-50% have at least one chromosomal rearrangement. Loss of specific chromosomal regions like 5q- and 7q- are usually the secondary cytogenetic abnormalities associated with MDS. In order to detect chromosome abnormalities associated with MDS, bone marrow samples from 26 patients diagnosed as MDS were obtained prior to chemotherapy. Both conventional cytogenetic analyses and fluorescence in situ hybridisation (FISH) methods were performed and locus-specific probes for 5q and 7q were used. Results obtained were compared. Twenty-one patients had normal karyotypes and four patients had abnormal karyotypes, while in one patient we could not obtain metaphases from cultures. Three patients with normal karyotypes revealed del (5q), two patients had del (7q) and one patient had monosomy (7). A total of 10 of 26 patients had chromosome changes visualised by either conventional or molecular cytogenetics (approximately 38.5%). Our results show that both methods are important in diagnosis and follow up of MDS patients. When used together, conventional cytogenetics and FISH detect clinically significant chromosome abnormalities in MDS patients.

9-Hydroxystearic acid upregulates p21(WAF1) in HT29 cancer cells.

Growing evidence supports the critical role of lipid peroxidation products in the control of cell proliferation. In previous studies we demonstrated the efficient restriction of the proliferation rate in several cell lines resulting from the in vitro treatment with endogenous lipid polar components of cell membranes. Among these, 9-hydroxystearic acid (9-HSA), a primary intermediate of lipid peroxidation, induced a significant arrest in G0/G1 in HT29 colon cancer cells. In response to 9-HSA treatment of HT29 we observed cell growth arrest and increase in p21(WAF1) expression both at the transcriptional and the translational levels. Growth of p21(WAF1)-deleted HCT116 human colon carcinoma cells was not inhibited by 9-HSA. We present evidence that p21(WAF1) is required for 9-HSA mediated growth arrest in human colon carcinoma cells.

Cytotoxic and cytostatic effects induced by 4-hydroxynonenal in human osteosarcoma cells.

Several studies point to the existence of an inverse correlation between cellular lipid peroxidation and both cell proliferation and neoplastic transformation. Furthermore, numerous results demonstrate that lipid peroxidation products affect central biochemical pathways and intracellular signalling at physiological concentrations. 4-Hydroxynonenal (HNE) is one of the most active products of lipid peroxidation. This work has focused on the evaluation of HNE nuclear content, so far never directly measured, by electrospray-ionization-mass-spectrometry (ESI/MS) and on the correlation between its concentration and the induced effects after exogenous administration. In a human osteosarcoma cell line (SaOS2), HNE exhibited an early cytotoxic effect characterized by apoptosis, cytostatic and differentiating effects characterized by slow growth, increase in alkaline phosphatase (ALP), and alpha5 integrin subunit content with decrease in tumorigenicity.

Gas chromatography/mass spectrometric assay of endogenous cellular lipid peroxidation products: quantitative analysis of 9- and 10-hydroxystearic acids.

A sensitive, specific, accurate and reproducible gas chromatography/mass spectrometry method was developed for the assay of 9- and 10-hydroxystearic acids in samples obtained as cell extracts. The preparation of the samples required specific procedures to allow the analysis of both the free and the conjugated hydroxy acids as the corresponding methyl esters. The quantification used propyl-paraben as the internal standard and monitoring of a specific fragment of each isomeric hydroxy acid methyl ester, and allowed quantification of the conjugate and the free fractions of both 9- and 10-hydroxystearic acids. This method is suitable for identification and quantification (LOQ 1.8 and 4.4 ng, respectively) of these important metabolites of lipid peroxidation. In particular the development of an assay for the free 9-hydroxystearic acid methyl ester makes the method a reliable analytical tool for investigations of the role of this metabolite in the mechanisms of tumour cell proliferation.

Determination of 4-hydroxy-2-nonenal at cellular levels by means of electrospray mass spectrometry.

trans-4-Hydroxy-2-nonenal (HNE) is an end-product of lipid peroxidation in biological systems which produces a variety of powerful biological effects. A method based on electrospray mass spectrometry was developed for the determination of 4-HNE at cellular levels. Quantification was carried out by using HNE-d(11) as internal standard; the mass chromatograms were acquired in the single ion monitoring mode (SIM) on the [M + H](+) monoisotopic species for HNE and HNE-d(11). With this approach a higher precision and lower detection limit and biological sample size than those typical of the methods so far employed are achieved. Furthermore the determination of the analyte from the cell extract is directly performed without the need of any HNE derivatization. As a first application the method was used to identify and quantify HNE in human T cell leukemia extracts.

Relations between accelerated sexual maturation and menstrual disorders.

The author demonstrates that the phenomenon of acceleration of sexual maturation is more frequent in urban areas. The factors having a part in determining this process are multiple: psychoemotional and audio-visual ones, and have an important role. The acceleration of sexual maturation has rather negative consequences, due to the significant increase of puberal menstrual troubles.