Detection of human cytomegalovirus DNA in various blood components after liver transplantation

The quantification of human cytomegalovirus (HCMV DNA) by real-time PCR is currently a primary option for laboratory diagnosis of HCMV infection. However, the optimal sample material remains controversial due to the use of different PCR assays. To explore the best blood component for HCMV DNA surveillance after liver transplantation, whole blood (WB), serum (SE), and plasma (PL) specimens were collected simultaneously from targeted patients and examined for HCMV DNA using one commercially available assay. The HCMV DNA-positive rate with WB (16.67%) was higher than that with either SE or PL (8.33%, both P<0.01). Quantitative DNA levels in WB were of greater magnitude than those in SE (WB-SE mean log-transformed difference, 0.99; 95%CI=0.74-1.25; P<0.0001) and PL (WB-PL mean log-transformed difference, 1.37; 95%CI=1.07-1.66; P<0.0001). Dynamic monitoring revealed that HCMV DNA in WB was positive sooner and had higher values for a longer period of time during therapy. With earlier positive detection, higher sensitivity, and yield of greater viral loads, WB compared favorably to SE or PL and hence is recommended as the superior material for HCMV DNA surveillance after liver transplantation. In addition, infant recipients require more intensive monitoring and prophylactic care because of their higher susceptibility to primary HCMV infection.

Successful domino liver transplantation in maple syrup urine disease using a related living donor

Maple syrup urine disease (MSUD) is an autosomal recessive disease associated with high levels of branched-chain amino acids. Children with MSUD can present severe neurological damage, but liver transplantation (LT) allows the patient to resume a normal diet and avoid further neurological damage. The use of living related donors has been controversial because parents are obligatory heterozygotes. We report a case of a 2-year-old child with MSUD who underwent a living donor LT. The donor was the patient's mother, and his liver was then used as a domino graft. The postoperative course was uneventful in all three subjects. DNA analysis performed after the transplantation (sequencing of the coding regions of BCKDHA, BCKDHB, andDBT genes) showed that the MSUD patient was heterozygous for a pathogenic mutation in the BCKDHB gene. This mutation was not found in his mother, who is an obligatory carrier for MSUD according to the family history and, as expected, presented both normal clinical phenotype and levels of branched-chain amino acids. In conclusion, our data suggest that the use of a related donor in LT for MSUD was effective, and the liver of the MSUD patient was successfully used in domino transplantation. Routine donor genotyping may not be feasible, because the test is not widely available, and, most importantly, the disease is associated with both the presence of allelic and locus heterogeneity. Further studies with this population of patients are required to expand the use of related donors in MSUD.

Cytokine expression patterns and mesenchymal stem cell karyotypes from the bone marrow microenvironment of patients with myelodysplastic syndromes

The purpose of this study was to explore cytokine expression patterns and cytogenetic abnormalities of mesenchymal stem cells (MSCs) from the bone marrow microenvironment of Chinese patients with myelodysplastic syndromes (MDS). Bone marrow samples were obtained from 30 cases of MDS (MDS group) and 30 healthy donors (control group). The expression pattern of cytokines was detected by customized protein array. The karyotypes of MSCs were analyzed using fluorescence in situ hybridization. Compared with the control group, leukemia inhibitory factor, stem cell factor (SCF), stromal cell-derived factor (SDF-1), bone morphogenetic protein 4, hematopoietic stem cell (HSC) stimulating factor, and transforming growth factor-β in the MDS group were significantly downregulated (P<0.05), while interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and programmed death ligand (B7-H1) were significantly upregulated (P<0.05). For chromosome abnormality analysis, the detection rate of abnormal karyotypes (+8, -8, -20, 20q-, -Y, -7, 5q-) was 30% in the MDS group and 0% in the control group. In conclusion, the up- and downregulated expression of these cytokines might play a key role in the pathogenesis of MDS. Among them, SCF and SDF-1 may play roles in the apoptosis of HSCs in MDS; and IFN-γ, TNF-α, and B7-H1 may be associated with apoptosis of bone marrow cells in MDS. In addition, the abnormal karyotypes might be actively involved in the pathogenesis of MDS. Further studies are required to determine the role of abnormal karyotypes in the occurrence and development of MDS.

Manipulation of a quasi-natural cell block for high-efficiency transplantation of adherent somatic cells

Recent advances have raised hope that transplantation of adherent somatic cells could provide dramatic new therapies for various diseases. However, current methods for transplanting adherent somatic cells are not efficient enough for therapeutic applications. Here, we report the development of a novel method to generate quasi-natural cell blocks for high-efficiency transplantation of adherent somatic cells. The blocks were created by providing a unique environment in which cultured cells generated their own extracellular matrix. Initially, stromal cells isolated from mice were expanded in vitro in liquid cell culture medium followed by transferring the cells into a hydrogel shell. After incubation for 1 day with mechanical agitation, the encapsulated cell mass was perforated with a thin needle and then incubated for an additional 6 days to form a quasi-natural cell block. Allograft transplantation of the cell block into C57BL/6 mice resulted in perfect adaptation of the allograft and complete integration into the tissue of the recipient. This method could be widely applied for repairing damaged cells or tissues, stem cell transplantation, ex vivo gene therapy, or plastic surgery.

Salvaged single-unit cord blood transplantation for 26 patients with hematologic malignancies not in remission

Treatments for patients with hematologic malignancies not in remission are limited, but a few clinical studies have investigated the effects of salvaged unrelated cord blood transplantation (CBT). We retrospectively studied 19 patients with acute leukemia, 5 with myelodysplastic syndrome (MDS with refractory anemia with excess blasts [RAEB]), and 2 with non-Hodgkin's lymphoma who received 1 CBT unit ≤2 loci human leukocyte antigen (HLA)-mismatched after undergoing myeloablative conditioning regimens between July 2005 and July 2014. All of them were in non-remission before transplantation. The infused total nucleated cell (TNC) dose was 4.07 (range 2.76-6.02)×107/kg and that of CD34+ stem cells was 2.08 (range 0.99-8.65)×105/kg. All patients were engrafted with neutrophils that exceeded 0.5×109/L on median day +17 (range 14-37 days) and had platelet counts of >20×109/L on median day +35 (range 17-70 days). Sixteen patients (61.5%) experienced pre-engraftment syndrome (PES), and six (23.1%) patients progressed to acute graft-versus-host disease (GVHD). The cumulative incidence rates of II-IV acute GVHD and chronic GVHD were 50% and 26.9%, respectively. After a median follow-up of 27 months (range 5-74), 14 patients survived and 3 relapsed. The estimated 2-year overall survival (OS), disease-free survival (DFS), and non-relapse mortality (NRM) rates were 50.5%, 40.3%, and 35.2%, respectively. Salvaged CBT might be a promising modality for treating hematologic malignancies, even in patients with a high leukemia burden.

Transplantation and survival of mouse inner ear progenitor/stem cells in the organ of Corti after cochleostomy of hearing-impaired guinea pigs: preliminary results

In mammals, damage to sensory receptor cells (hair cells) of the inner ear results in permanent sensorineural hearing loss. Here, we investigated whether postnatal mouse inner ear progenitor/stem cells (mIESCs) are viable after transplantation into the basal turns of neomycin-injured guinea pig cochleas. We also examined the effects of mIESC transplantation on auditory functions. Eight adult female Cavia porcellus guinea pigs (250-350g) were deafened by intratympanic neomycin delivery. After 7 days, the animals were randomly divided in two groups. The study group (n=4) received transplantation of LacZ-positive mIESCs in culture medium into the scala tympani. The control group (n=4) received culture medium only. At 2 weeks after transplantation, functional analyses were performed by auditory brainstem response measurement, and the animals were sacrificed. The presence of mIESCs was evaluated by immunohistochemistry of sections of the cochlea from the study group. Non-parametric tests were used for statistical analysis of the data. Intratympanic neomycin delivery damaged hair cells and increased auditory thresholds prior to cell transplantation. There were no significant differences between auditory brainstem thresholds before and after transplantation in individual guinea pigs. Some mIESCs were observed in all scalae of the basal turns of the injured cochleas, and a proportion of these cells expressed the hair cell marker myosin VIIa. Some transplanted mIESCs engrafted in the cochlear basilar membrane. Our study demonstrates that transplanted cells survived and engrafted in the organ of Corti after cochleostomy.

Cytotoxicity and mutagenicity of cola and grape flavored soft drinks in bone marrow cells of rodents

Due to the large consumption of soft drinks in Brazil and worldwide in recent years and considering that some of the components present in their composition pose potential risks to human health, the aim of this study was to evaluate the cytotoxic and mutagenic potential of specific cola and grape-flavored soft drink brands. Bone marrow cells of Wistar rats were initially treated by gavage with one single dose of Cola or Grape soft drink, which was next offered ad libitum (instead of water) for 24 hours. A negative control treatment was performed by administering one single dose of water and a positive control administering cyclophosphamide intraperitoneally. Statistical analysis showed that the Cola and Grape soft drinks studied were not cytotoxic. However, the Cola soft drink proved mutagenic in this experiment treatment time. Therefore, this study serves as a warning about the consumption of Cola-flavored soft drink and for the need for further subchronic and chronic studies on soft drinks in order to evaluate the long term mutagenic and cytotoxic effects of these substances.

Histopathological analysis of pre-implantation donor kidney biopsies: association with graft survival and function in one year post-transplantation

Introduction: Pre-implantation kidney biopsy is a decision-making tool when considering the use of grafts from deceased donors with expanded criteria, implanting one or two kidneys and comparing this to post-transplantation biopsies. The role of histopathological alterations in kidney compartments as a prognostic factor in graft survival and function has had conflicting results. Objective: This study evaluated the prevalence of chronic alterations in pre-implant biopsies of kidney grafts and the association of findings with graft function and survival in one year post-transplant. Methods: 110 biopsies were analyzed between 2006 and 2009 at Santa Casa de Porto Alegre, including live donors, ideal deceased donors and those with expanded criteria. The score was computed according to criteria suggested by Remuzzi. The glomerular filtration rate (GFR) was calculated using the abbreviated MDRD formula. Results: No statistical difference was found in the survival of donors stratified according to Remuzzi criteria. The GFR was significantly associated with the total scores in the groups with mild and moderate alterations, and in the kidney compartments alone, by univariate analysis. The multivariate model found an association with the presence of arteriosclerosis, glomerulosclerosis, acute rejection and delayed graft function. Conclusion: Pre-transplant chronic kidney alterations did not influence the post-transplantation one-year graft survival, but arteriosclerosis and glomerulosclerosis is predictive of a worse GFR. Delayed graft function and acute rejection are independent prognostic factors.