尖叶清风藤化学成分的研究

从尖叶清风藤(Sabia swinhoei Hmsl.exForb.etHemsl.)的地上部分分离出8个化合物,经理化常数测定和光谱解析,证明其中2个为新成分,分别命名为清风藤内酯(I)和清风藤酮(II),另外6个为已知化合物,即没药烯(III)、三十烷醇(IV)、β-谷甾醇(V)、二十八烷酸(VI)、齐墩果酸(VII)和胡萝卜甙(VIII)。     

Human erythrocyte protein 4.2 deficiency associated with hemolytic anemia and a homozygous 40glutamic acid-->lysine substitution in the cytoplasmic domain of band 3 (band 3Montefiore)

Red blood cell (RBC) protein 4.2 deficiency is often associated with a moderate nonimmune hemolytic anemia, splenomegaly, and osmotically fragile RBCs resembling, but not identical to, hereditary spherocytosis (HS). In the Japanese type of protein 4.2 deficiency (protein 4.2Nippon), the anemia is associated with a point mutation in the protein 4.2 cDNA. In this report, we describe a patient with moderate and apparently episodic nonimmune hemolytic anemia with splenomegaly, spherocytosis, osmotically fragile RBCs, reduced whole cell deformability, and abnormally dense cells. Sodium dodecyl sulfate- polyacrylamide gel electrophoresis analysis of the proposita's RBC membrane proteins showed an 88% deficiency of protein 4.2 and a 30% deficiency of glyceraldehyde-3-phosphate dehydrogenase (band 6). Structural and molecular analyses of the proposita's protein 4.2 were normal. In contrast, limited tryptic digestion of the proposita's band 3 showed a homozygous abnormality in the cytoplasmic domain. Analysis of the pedigree disclosed six members who were heterozygotes for the band 3 structural abnormality and one member who was a normal homozygote. Direct sequence analysis of the abnormal band 3 tryptic peptide suggested that the structural abnormality resided at or near residue 40. Sequence analysis of the proposita's band 3 cDNA showed a 232G-->A mutation resulting in a 40glutamic acid-->lysine substitution (band 3Montefiore). Allele-specific oligonucleotide hybridization was used to probe for the mutation in the pedigree, showing that the proposita was homozygous, and the pedigree members who were heterozygous for the band 3 structural abnormality were also heterozygous for the band 3Montefiore mutation. The band 3Montefiore mutation was absent in 26 chromosomes from race-matched controls and in one pedigree member who did not express the band 3 structural abnormality. In coincidence with splenectomy, the proposita's anemia was largely corrected along with the disappearance of most spherocytes and considerable improvements of RBC osmotic fragility, whole cell deformability, and cell density. We conclude that this hereditary hemolytic anemia is associated with the homozygous state for band 3Montefiore (40glutamic acid-->lysine) and a decreased RBC membrane content of protein 4.2. We speculate that band 3 structural abnormalities can result in defective interactions with protein 4.2 and band 6, and in particular, that the region of band 3 containing 40glutamic acid is involved directly or indirectly in interactions with these proteins.     

Cytogenetic and histologic correlations in malignant lymphoma

Although a number of studies have indicated correlations between histologic subtypes of tumors and certain nonrandom chromosome changes, cytogenetic studies of lymphoma are in an early stage compared to those of leukemia. No comprehensive analysis of available data has so far been attempted in the literature either. Here we present an analysis of chromosome changes and their correlation with subtypes of lymphoma studied by conventional histology and cell surface markers, as observed in two sets of data: a group of 65 karyotypically abnormal tumors sequentially ascertained and studied by us during the period January 1, 1984 to April 30, 1985, and a larger data set derived by combining our data with those from two published series from the University of Minnesota that are comparable to our data. These combined data, which comprise the largest data set on the cytogenetics of lymphomas assembled so far, enabled a comprehensive analysis of correlation between chromosome change and tumor histology and the patterns of chromosome instability in these tumors. We found several significant associations, some previously described and others now recognized, between nonrandom chromosome gains, breaks, translocations, and deletions and histologic subtypes of tumors that characterize lymphomas. The data indicate that finding of chromosome breaks at certain sites (eg, 8q24, 14q32, 18q21) is of diagnostic value in dealing with cases of unusual lymphoma. Furthermore, nonrandom chromosome breakage exhibited three distinct patterns that reflected three levels of etiologically relevant genetic change.     

Development of a Novel Human Cell-Derived Tissue-Engineered Heart Valve for Transcatheter Aortic Valve Replacement: an In Vitro and In Vivo Feasibility Study

Transcatheter aortic valve replacement (TAVR) is being extended to younger patients. However, TAVR-compatible bioprostheses are based on xenogeneic materials with limited durability. Off-the-shelf tissue-engineered heart valves (TEHVs) with remodeling capacity may overcome the shortcomings of current TAVR devices. Here, we develop for the first time a TEHV for TAVR, based on human cell-derived extracellular matrix and integrated into a state-of-the-art stent for TAVR. The TEHVs, characterized by a dense acellular collagenous matrix, demonstrated in vitro functionality under aortic pressure conditions (n?=?4). Next, transapical TAVR feasibility and in vivo TEHV functionality were assessed in acute studies (n?=?5) in sheep. The valves successfully coped with the aortic environment, showing normal leaflet motion, free coronary flow, and absence of stenosis or paravalvular leak. At explantation, TEHVs presented full structural integrity and initial cell infiltration. Its long-term performance proven, such TEHV could fulfill the need for next-generation lifelong TAVR prostheses.     

Homeostasis maintenance of encapsulated cells

Cell niche homeostasis plays a critical role in many bodily functions including tissue functionality, stem cell maintenance and differentiation, wound healing, cancer development and propagation, and many others. Many tissue engineering approaches overlook the importance of engineered constructs homeostasis, in particular for transplantation purposes. Here, we present a study of the effect of encapsulation duration on engineered tissue maturation and provide a protocol for the evaluation of critical conditions for transplantation purposes. In brief, SHSY5Y human neuroblastoma cells were encapsulated in 2% alginate by electrohydrodynamic jetting method for up to 4 weeks. We evaluated extracellular matrix niche formation and tissue maturation in situ through COL1A1 expression. In in vitro conditions, we studied the ability of cells to maintain their critical functions after being released from alginate beads. Cellular viability was evaluated via an apoptosis/necrosis detection kit and AlamarBlue assay, and functionality via immunocytochemistry. We proved the importance of engineered tissue homeostasis stabilization for future cell recovery, in particular, for our system cells encapsulated for 28 days met all critical requirements for successful tissue transplantation. Maturation of engineered tissue constructs could be accelerated by enriching alginate with growth factors or extracellular matrix molecules.     

Sensitivity of Ph1+ CFU-GM to human recombinant interferon α and γ alone and in combination

Summary The in vitro effect of human recombinant interferon (IFN) alone and in combination were studied on granulomonocytic colony forming units (CFU-GM) from the peripheral blood of 10 Ph1+ chronic myeloid leukemia (CML) patients and from the marrow of 5 normal or non-leukemic subjects. - and -IFN alone determined a slight inhibition on colony growth with a preferential effect on pure macrophagic colonies. At maximum concentration (10⁴ U/ml) leukemic colony inhibition was 46±34% for IFN and 43±19% for IFN. Culture growth with + IFN in combination were significantly inhibited (up to 96±4%) with a concentration-related effect. Similar results were obtained with normal CFU-GM. The synergism that was found in vitro is probably relevant for the in vivo therapeutic effects of these compounds in CML and suggest that the combination is worth testing in vivo.This work was supported by CNR, Oncology Finalized Project, contract n. 8701169.44. and n. 8600603.44. and by AIRC     

Clinical and molecular follow-up by amplification of the CDR-III IgH region in multiple myeloma patients after autologous transplantation of hematopoietic CD34+ stem cells.

BACKGROUND AND OBJECTIVE: Autologous blood stem cell transplantation (ABSCT) using chemotherapy-induced mobilization of peripheral blood stem cells (PBSC) is being increasingly used in the treatment of multiple myeloma (MM). We report the clinical and molecular follow-up of 10 MM patients who underwent autologous stem cell transplantation with peripheral blood selected CD34+ cells, as support therapy following a myeloablative conditioning regimen. DESIGN AND METHODS: The CDR-III coding region of the IgH gene was studied by a) consensus PCR applied to 8 MM patients, or b) by direct sequencing of PCR product generated by family-specific primers in the remaining two patients (who became immunofixation analysis (IF) negative). In this case, two patient-specific primers were generated, thus obtaining a high PCR assay sensitivity and specificity (ASO PCR). RESULTS: Seven patients are alive: 4 of them have serum M protein assessable by IF, while 1 was not a secretor and 2 converted from serum IF positivity to negativity 6 and 12 months after ABSCT. Three patients died: 1 from disease progression and 2 from infective complications during clinical remission. The molecular analysis during the follow-up showed that the bone marrow samples from the two patients who obtained IF negativity were persistently PCR positive for the presence of rearranged CDR-III region. Moreover, despite the remarkable reduction of myeloma burden, a minimal level of residual myeloma cells was still detectable by molecular analysis. INTERPRETATION AND CONCLUSIONS: These results confirm that although positive selection of CD34+ cells markedly reduces the contamination of myeloma cells from apheresis products by up to 3 log, and provides a cell suspension capable of restoring normal hematopoiesis after ablative conditioning regimen, it does not abrogate myeloma cell contamination in most of the apheresis products.     

Ultrastructural changes of coronary artery endothelium induced by cardioplegic solutions

Coronary artery endothelial and myocardial ultrastructure was studied in guinea-pig heart-lung preparations (HLP) subjected to ischemic cardiac arrest induced by three hypothermic solutions. Two of the solutions used had high potassium chloride concentration ("Alabama" and "St. Thomas") while the third, instead, was a bicarbonate buffer (Kreb's solution). Five experimental groups were studied. In group 1 (control) the HLP were not subjected to cardiac arrest. Groups 2, 3, and 4 were subjected to a period of cardiac arrest of 30, 60, and 120 minutes respectively. In group 5, HLP were reperfused with blood for 30 minutes after 60 minutes of cardiac arrest. A thin ring of the left anterior descending coronary artery and myocardial fragments were obtained at the end of each experiment and were analyzed by means of transmission electron microscopy (TEM). Functional parameters were recorded in group 5. HLP perfused with Alabama solution showed a well-preserved endothelium and myocardium. HLP perfused with Krebs solution showed slight changes of the endothelial glycocalix only in group 4. Further, HLP perfused with Krebs solution showed extensive myocardial lesions (groups 3 and 4). These ischemic changes were not completely reversed after reperfusion (group 5). HLP perfused with St. Thomas solution showed only endothelial changes. These lesions were mainly characterized by: disappearance of the glycocalix and pynocytotic vesicles, endothelial cell bulging (group 2), and loss of the endothelial continuity (groups 3, 4, and 5). Hemodynamic parameters were significantly changed only in the Krebs-perfused HLP which showed a deterioration of the cardiac function related to the ischemic damage.(ABSTRACT TRUNCATED AT 250 WORDS)