The role of fibrinogen A alpha chains in ADP-induced platelet aggregation in the presence of fibrinogen molecules containing gamma' chains

Human plasma fibrinogen (Fgn) is heterogenous with respect to the size of its gamma chains, which differ in that residues 408 to 411 of gammaA chains (93% of total) are replaced in gamma' chains by a unique 20 amino acid sequence (gamma408 to gamma427). In this study, we compared the contribution to adenosine diphosphate (ADP)-induced platelet aggregation of the A alpha chains in Fgn molecules containing predominantly (fraction 1-2) or exclusively (peak 1 Fgn) gammaA chains with that of molecules containing approximately 50% gamma' chains (peak 2 Fgn). Using washed human platelets, we confirmed that the number of peak 2 Fgn molecules binding to platelets in the presence of ADP was about half the number of peak 1 Fgn molecules (18,962 +/- 2,298 v 44,366 +/- 16,096 molecules per platelet), and that isolated S- carboxymethylated (SCM) gammaA chains supported ADP-induced platelet aggregation nearly as well as peak 1 Fgn. In contrast, SCM-gamma' chains alone supported aggregation poorly, whereas a mixture of SCM- gammaA and gamma' chains (1:1 ratio) gave intermediate results. Despite the findings with isolated SCM-gamma' chains, we found that peak 2 Fgn supported platelet aggregation nearly as well as peak 1 Fgn. However, peak 2 Fgn from which carboxy (COOH)-terminal A alpha chain segments had been removed by digestion with plasmin showed a markedly decreased platelet aggregation potential. Peak 1 Fgn core fraction from an 88% to 90% coagulable plasmin digest, or Fgn fraction 1-9, which has a high gammaA/gamma' chain ratio (93:7), but lacks COOH-terminal regions of A alpha chains, supported platelet aggregation to the same extent as did intact peak 2 Fgn. These findings indicate that Fgn molecules containing gamma' chains can approach the aggregation potential of Fgn molecules containing predominantly or exclusively gammaA chains only if intact A alpha chains are also present.     

Prognostic importance of structural chromosomal abnormalities in children with hyperdiploid (greater than 50 chromosomes) acute lymphoblastic leukemia

Approximately one fourth of children with newly diagnosed acute lymphoblastic leukemia (ALL) have hyperdiploid (greater than 50 chromosomes) blasts and a relatively favorable prognosis. Nonetheless, a substantial proportion of these patients fail therapy. We studied 138 children (70 male, 68 female) with hyperdiploid greater than 50 ALL to assess initial clinical and cytogenetic features that might predict treatment failure. In 85 of these cases (62%), structural chromosomal abnormalities were also present; clinical and laboratory features in this group did not differ from those of the 53 cases with only numeric abnormalities. However, of the 28 failures seen at a median follow-up of 4 years, 22 occurred in cases with structural chromosomal abnormalities (P = .03 by Breslow test). In a multivariate analysis, only the presence of structural chromosomal abnormalities and male gender were independently associated with treatment failure. Structural chromosomal abnormalities in cases of ALL with greater than 50 chromosomes may define a biologically different form of leukemia characterized by increased likelihood of drug resistance.     

Human interferon-inducible protein-10 induces mononuclear cell infiltration in mice and promotes the migration of human T lymphocytes into the peripheral tissues and human peripheral blood lymphocytes-SCID mice

The human cytokine, interferon-inducible protein-10 (IP-10), is a small glycoprotein secreted by activated monocytes, T cells, keratinocytes, astrocytes, and endothelial cells and is structurally related to the alpha subfamily of chemotactic cytokines called chemokines (Taub and Oppenheim, Cytokine 5:175, 1993). However, in contrast to other alpha chemokines that induce neutrophil migration, IP-10 has been shown to chemoattract monocytes and T lymphocytes in vitro, suggesting a role in T-cell-mediated immune responses. We therefore examined the effects of human IP-10 after in vivo administration. IP-10 induces significant mononuclear cell infiltration after subcutaneous injections in normal mice. In an effort to study the in vivo effects of IP-10 on human leukocyte migration, we then examined the ability of recombinant human IP-10 (rhIP-10) to induce human-T-cell infiltration using a human/severe combined immune deficiency (SCID) mouse model. SCID mice received an intraperitoneal injection of human peripheral blood lymphocytes (10(8) cells), followed by a subcutaneous injection of rhIP- 10 (1 micrograms/injection) in the hind flank for 4 hours or sequential injections for 3 days. The skin and underlying tissue from the rhIP-10 injection site were then biopsied and examined for the extent of mononuclear cell infiltration. rhIP-10 again induced significant mononuclear cell accumulation 72 hours after injection. Immunohistologic evaluation determined that a significant number of human CD3+ T cells were recruited in response to rhIP-10 injections. These results show that rhIP-10 is capable of inducing human T-cell migration in vivo and may play an important role in monocyte and lymphocyte recruitment into inflammatory sites.     

Augmentation of donor bone marrow engraftment in histoincompatible murine recipients by granulocyte/macrophage colony-stimulating factor

T cell depletion of donor bone marrow can prevent graft v host disease (GVHD) in human and murine marrow graft recipients. However, engraftment in the recipient may be compromised as a consequence of donor marrow T cell depletion. The effect of recombinant murine granulocyte/macrophage colony-stimulating factor (rmu GM-CSF) on engraftment and hematologic reconstitution was evaluated in a murine allogeneic bone marrow transplantation (BMT) model involving T cell depletion of marrow. Before transplantation into irradiated mice differing at major and minor histocompatibility loci, rmu GM-CSF was preincubated with T cell-depleted donor marrow. When low degrees of engraftment were noted in control recipients, treatment of donor marrow with high concentrations of rmu GM-CSF led to enhanced engraftment. Ex vivo donor graft incubation with rmu GM-CSF or a single in vivo injection of rmu GM-CSF were both effective means of promoting engraftment. When the engraftment rate in control recipients was high, rmu GM-CSF did not have an identifiable effect. Only slight increases in hematologic recovery were detected regardless of the rate of engraftment. Neither post-BMT survival nor marrow stem cell capacity was affected by rmu GM-CSF incubation. Furthermore, growth factor administration did not have a significant effect on the incidence of GVHD in recipients of non-T cell-depleted bone marrow splenocyte preparations. In vitro natural killer-mediated target cell lysis was not altered by incubation of effector cells with rmu GM-CSF. These results demonstrate the potential of ex vivo rmu GM-CSF treatment of donor marrow to facilitate engraftment across extensive histo- compatibility barriers.     

Cloning of thermostable DNA polymerases from hyperthermophilic marine Archaea with emphasis on Thermococcus sp. 9 degrees N-7 and mutations affecting 3''-5'' exonuclease activity.

Five extremely thermophilic Archaea from hydrothermal vents were isolated, and their DNA polymerases were cloned and expressed in Escherichia coli. Protein splicing elements (inteins) are present in many archaeal DNA polymerases, but only the DNA polymerase from strain GB-C contained an intein. Of the five cloned DNA polymerases, the Thermococcus sp. 9 degrees N-7 DNA polymerase was chosen for biochemical characterization. Thermococcus sp. 9 degrees N-7 DNA polymerase exhibited temperature-sensitive strand displacement activity and apparent Km values for DNA and dNTP similar to those of Thermococcus litoralis DNA polymerase. Six substitutions in the 3'-5' exonuclease motif I were constructed in an attempt to reduce the 3'-5' exonuclease activity of Thermococcus sp. 9 degrees N-7 DNA polymerase. Five mutants resulted in no detectable 3'-5' exonuclease activity, while one mutant (Glul43Asp) had <1% of wild-type activity.     

2009年美国血液病学年会造血干细胞移植热门临床研究报告

<正>2009年美国血液病学年会上研究者对异基因造血干细胞移植(allogeneic hematopoietic stem cell transplantation,allo-HSCT)予以极大的关注,相关会议摘要超过1200篇。本文就一些涉及重要的、被反     

Delayed diagnosis of Gorlin's syndrome in a renal transplant recipient

A 35-year-old woman was referred to the dedicated dermatology clinic for RTR. She underwent her first renal transplant at 18 years of age due to chronic renal failure following reflux nephropathy of the single right kidney (left kidney agenesis). She has since then had two further transplants. During clinical examination she was noted to have at least 16 basal cell carcinomas (BCC) and there are records in the case notes of 10 BCC having being excised and confirmed histologically in the past. By contrast, she had only had two squamous cell carcinomas (SCC) excised. She was also noted to have distinctive facial features, a kyphoscoliosis and palmar pits, and a diagnosis of Gorlin's (naevoid BCC) syndrome (GS) was made. Although immunosuppression may have contributed to the multiplicity of her BCC, the contrasting very small number of SCC is unusual in an organ transplant recipient and this alerted the authors to the diagnosis of GS.     

Phospholipid/deoxycytidine analogue prodrugs for the treatment of cancer

We synthesized thioether phospholipid carrier molecules, conjugated each of them to 1-b-D-arabinofuranosylcytosine (ara-C), and synthesized amido containing phospholipid carriers conjugated to gemcitabine. Changing the alkyl chain at the C1- and C2-positions of the phospholipid increased the conjugates’ cytotoxicity over previous conjugates. Dipyridamole increased ara-C’s and gemcitabine’s IC₅₀ value while the IC₅₀ values for the phospholipid conjugates were relatively unchanged suggesting that phospholipid conjugates do not require a transporter for entry into the cell. The phospholipid conjugates were cytotoxic to MCF-7 cells and its multidrug resistance-1 (MDR-1) overexpressing cell line derivative (BC-19). Ara-C had no effect on either cell line. Therefore, these novel phospholipid/nucleoside analogue conjugates could be used for the treatment of tumor cells that express certain resistance phenotypes such as a loss of transporter activity and/or MDR-1 overexpression. In vivo the gemcitabine-phospholipid conjugate was well tolerated and prolonged the survival of tumor bearing mice compared to control mice.     

Cyclooxygenase inhibition and hyperthermia for the potentiation of the cytotoxic response in ovarian cancer cells

OBJECTIVES: The progression of chemotherapy-resistant cancer confers poor prognosis and decreases overall survival in ovarian cancer patients. Adjuvants to traditional chemotherapy regimens have become attractive modalities for the clinical treatment of refractory or resistant ovarian cancer. We evaluated whether the addition of NSAID to hyperthermic chemotherapy would increase cytotoxicity in cisplatin- and taxane-treated ovarian cancer cells. METHODS: Western blot analysis was utilized to determine COX-2 protein expression levels in the 2008, cisplatin-sensitive, and C13*, cisplatin-resistant, cell lines. PGE2 levels were determined and analyzed as a function of cyclooxygenase activity by LC/MS/MS. Cells were treated with cisplatin, docetaxel or paclitaxel in combination with either NS-398 or sulindac sulfide at 37 degrees C, 41 degrees C or 43 degrees C. Cell viability was determined by a MTS cell proliferation assay. RESULTS: Both cell lines expressed COX-2 protein, and NS-398 and sulindac sulfide effectively blocked PGE2 production. The addition of a NSAID to cisplatin treatment in 2008 and C13* cells offered enhanced cytotoxicity and this effect was further enhanced at 41 degrees C. In docetaxel-treated 2008 cells, both NS-398 and sulindac sulfide offered enhanced cell kill; however, this result was not observed in paclitaxel-treated cells. Hyperthermia appeared to play no additional role in taxane cytotoxicity enhancement, however no antagonism was detected. CONCLUSIONS: Our results suggest that the combination treatment (cisplatin or docetaxel in combination with NSAID) cause a dose-dependent enhancement of cytotoxicity. Hyperthermia may improve the results of intraperitoneal cisplatin therapy, thus warranting further evaluation in clinical studies.