21 research outputs found
An Epstein-Barr Virus-Associated Leukemic Lymphoma in a Patient Treated with Rabbit Antithymocyte Globulin and Cyclosporine for Hepatitis-Associated Aplastic Anemia
Lymphoproliferative disorders (LPDs) are generally caused by uncontrolled B-cell proliferation induced by the Epstein-Barr virus (EBV) in the setting of impaired EBV-specific T-cell immunity, particularly when there is pharmacological immunosuppression including antithymocyte globulin. We herein present an unusual case of EBV associated with LPD (EBV-LPD) in which LPD occurred 3 weeks after the use of rabbit antithymocyte globulin administered for severe hepatitis-associated aplastic anemia; the patient died of fulminant leukemic lymphoma 5 days after the onset. We also review the pertinent literature on EBV-LPD after immunosuppressive therapy and document the efficacy of EBV viral load monitoring and the need for preemptive therapy. Copyright © 2011 S. Karger AG, Basel
Intensive chemotherapy for a relapsed ALL patient who received living-donor lobar lung transplantation
Graft rejection and hyperacute graft-versus-host disease in stem cell transplantation from non-inherited maternal antigen complementary HLA-mismatched siblings
金沢大学大学院医学系研究科機能再生学Human leukocyte antigen (HLA)-mismatched stem cell transplantation from non-inherited maternal antigen (NIMA)-complementary donors is known to produce stable engraftment without inducing severe graft-versus-host disease (GVHD). We treated two patients with acute myeloid leukemia (AML) and one patient with severe aplastic anemia (SAA) with HLA-mismatched stem cell transplantation (SCT) from NIMA-complementary donors (NIMA-mismatched SCT). The presence of donor and recipient-derived blood cells in the peripheral blood of recipient (donor microchimerism) and donor was documented respectively by amplifying NIMA-derived DNA in two of the three patients. Graft rejection occurred in the SAA patient who was conditioned with a fludarabine-based regimen. Grade III and grade IV acute GVHD developed in patients with AML on day 8 and day 11 respectively, and became a direct cause of death in one patient. The findings suggest that intensive conditioning and immunosuppression after stem cell transplantation are needed in NIMA-mismatched SCT even if donor and recipient microchimerisms is detectable in the donor and recipient before SCT. © 2007 The Authors
An Epstein-Barr Virus-Associated Leukemic Lymphoma in a Patient Treated with Rabbit Antithymocyte Globulin and Cyclosporine for Hepatitis-Associated Aplastic Anemia
Efficient Multiple Genome Modifications Induced by the crRNAs, tracrRNA and Cas9 Protein Complex in Zebrafish
<div><p>The type II clustered regularly interspaced short palindromic repeats (CRISPR) associated with Cas9 endonuclease (CRISPR/Cas9) has become a powerful genetic tool for understanding the function of a gene of interest. In zebrafish, the injection of Cas9 mRNA and guide-RNA (gRNA), which are prepared using an <i>in vitro</i> transcription system, efficiently induce DNA double-strand breaks (DSBs) at the targeted genomic locus. Because gRNA was originally constructed by fusing two short RNAs CRISPR RNA (crRNA) and <i>trans</i>-activating crRNA (tracrRNA), we examined the effect of synthetic crRNAs and tracrRNA with Cas9 mRNA or Cas9 protein on the genome editing activity. We previously reported that the disruption of <i>tyrosinase</i> (<i>tyr</i>) by tyr-gRNA/Cas9 mRNA causes a retinal pigment defect, whereas the disruption of <i>spns2</i> by spns2-gRNA1/Cas9 mRNA leads to a cardiac progenitor migration defect in zebrafish. Here, we found that the injection of spns2-crRNA1, tyr-crRNA and tracrRNA with Cas9 mRNA or Cas9 protein simultaneously caused a migration defect in cardiac progenitors and a pigment defect in retinal epithelial cells. A time course analysis demonstrated that the injection of crRNAs and tracrRNA with Cas9 protein rapidly induced genome modifications compared with the injection of crRNAs and tracrRNA with Cas9 mRNA. We further show that the crRNA-tracrRNA-Cas9 protein complex is functional for the visualization of endogenous gene expression; therefore, this is a very powerful, ready-to-use system in zebrafish.</p></div
Intensive chemotherapy for a relapsed ALL patient who received living-donor lobar lung transplantation
Detection of heteroduplexes at <i>spns2</i> and <i>tyr</i> loci by HMA in F1 embryos.
<p>F0 founder male 3 and female 5 were mated with wild-type fish and genomic DNA from individual F1 embryos was isolated at 1 dpf. PCR amplicons for <i>spns2</i> or <i>tyr</i> were electrophoresed on a 15% polyacrylamide gel. We observed heteroduplex bands containing mutant alleles (brackets) at <i>spns2</i> and <i>tyr</i> loci. The positions of the expected homoduplexes are indicated by the arrowheads. The sizes of the DNA ladder markers are indicated by the base pairs at the left.</p
Phenotypic analysis in the embryos injected with two crRNAs, tracrRNA and Cas9 protein.
<p>(A, F) An uninjected embryo derived from Tg(<i>cmlc2</i>:<i>eGFP</i>) expressing eGFP in the cardiac cells. (B, G) Tg(<i>cmlc2</i>:<i>eGFP</i>)-derived embryos injected with spns2-crRNA1 (25 pg), tyr-crRNA (25 pg), tracrRNA (100 pg) and Cas9 protein (400 pg). (C, H) Tg(<i>cmlc2</i>:<i>eGFP</i>)-derived embryos injected with spns2-gRNA1 (25 pg), tyr-gRNA (25 pg) and Cas9 protein (400 pg). (D, I) Tg(<i>cmlc2</i>:<i>eGFP</i>)-derived embryos injected with spns2-crRNA1 (25 pg), tyr-crRNA (25 pg), tracrRNA (100 pg) and Cas9 mRNA (250 pg). (E, J) Tg(<i>cmlc2</i>:<i>eGFP</i>)-derived embryos injected with spns2-gRNA1 (25 pg), tyr-gRNA (25 pg) and Cas9 mRNA (250 pg). (A-E) The injection of the two crRNAs and tracrRNA with Cas9 protein or Cas9 mRNA (B and C) as well as the injection of two gRNAs with Cas9 protein or Cas9 mRNA (D and E) caused the cardiac progenitor migration defects at 1 day post-fertilization (dpf), whereas an uninjected embryo had a normal heart (A). White arrowheads indicate the position of the developing heart. (F–J) The injection of the two crRNAs and tracrRNA with Cas9 protein or Cas9 mRNA (G and H) as well as the injection of two gRNAs with Cas9 protein or Cas9 mRNA (I and J) caused pigmentation defects in the retinal epithelium at 2 dpf, whereas an uninjected embryo had retinal epithelial cells with normal pigmentation (F). Black arrowheads indicate the position of the eye. The embryos in (A), (B), (C), (D) and (E) correspond to the embryos in (F), (G), (H), (I) and (J), respectively. (A-E) Ventral view with anterior at the top. (F-J) Lateral view with anterior to the left and dorsal at the top.</p
Visualization of endogenous gene expression by knock-in of the eGFP reporter using the crRNA-tracrRNA-Cas9 protein complex.
<p>(A) Whole-mount <i>in situ</i> hybridization with an <i>epdr1</i> probe. (B) An uninjected embryo. (C) The embryo injected two crRNAs (epdr1-crRNA; 25 pg + Mbait-crRNA; 25 pg), tracrRNA (100 pg) and Mbait-hs-eGFP (25 pg) with Cas9 protein (400 pg). The expression of eGFP was detected in the anterior central nervous system, including neurons.</p
Comparison of the crRNAs and gRNAs on the Cas9 protein-mediated genome editing activity.
<p>Three crRNAs (spns2-crRNA2; 25 pg + s1pr2-crRNA1; 25 pg + s1pr2-crRNA2; 25 pg) plus tracrRNA (100 pg) or three gRNAs (spns2-gRNA2; 25 pg + S1PR2-gRNA1; 25 pg + S1PR2-gRNA2; 25 pg) were co-injected with Cas9 protein (400 pg) into zebrafish embryos and the genomic DNA was prepared from 24 hpf embryos. The genome modifications induced by the Cas9 protein were assessed by a HMA. Target sites for S1PR2-gRNA1 and S1PR2-gRNA2 are located within the binding sites of S1PR2-specific primers. The positions of the expected homoduplexes are indicated by the asterisks. The sizes of the DNA ladder markers are indicated by the base pairs at the left.</p