There is a growing demand for in vitro assays for toxicity screening in three-dimensional (3D) environments. In this study, 3D cell culture using magnetic levitation was used to create an assay in which cells were patterned into 3D rings that close over time. The rate of closure was determined from time-lapse images taken with a mobile device and related to drug concentration. Rings of human embryonic kidney cells (HEK293) and tracheal smooth muscle cells (SMCs) were tested with ibuprofen and sodium dodecyl sulfate (SDS). Ring closure correlated with the viability and migration of cells in two dimensions (2D). Images taken using a mobile device were similar in analysis to images taken with a microscope. Ring closure may serve as a promising label-free and quantitative assay for high-throughput in vivo toxicity in 3D cultures.
The present study aimed to define the natural history, World Health Organization (WHO) classification, prognostic factors, and treatment outcome of 87 patients with primary lymphoma of the palatine tonsil and literature review and analysis. Between 1990 and March 2008, 87 consecutive patients diagnosed with primary lymphoid malignancy of the palatine tonsil. All pathologic specimens were reviewed and reclassified according to the recent WHO classification. To investigate the association of tonsillar lymphomas with Epstein-Barr virus (EBV), in situ hybridization was performed for 24 tonsillar lymphomas (23 diffuse large B-cell lymphoma (DLBC) and one classic Hodgkin's disease) and ten normal tonsils as control group. In literature review, we found 26 major related series including 1,602 patients with primary tonsillar lymphoma. The median age of our patients was 52 years (range 11-86 years). There were 39 women and 48 men with a median follow-up of 67 months for living patients. The vast majority (95%) of patients had B-cell phenotype. DLBC was the most frequent histology. In situ hybridization revealed none of 23 DLBC to be positive for EBV. The 5-year disease-free and overall survival rates were 78.9% and 86%, respectively. In the literature review and by analyzing the data collection from 26 major reported series, the median age was 55 years and male/female ratio was 1.3:1. Intermediate grade tumors consisted of 72% of all tonsillar lymphomas and B-cell lymphomas constituted 82% of all cell immunophenotypes. The 5-year disease-free and overall survival rates were 61% and 67%, respectively. The vast majority of tonsillar lymphomas are of B-cell origin and with intermediate to high-grade histology. These neoplasms tend to present in early stage disease and to have favorable outcome. WHO classification predicts more accurately treatment outcome of patients with tonsillar lymphoma. The association of DLBC in the palatine tonsil with EBV infection is infrequent.
Accurate cause of death coding leads to organised and usable death information but there are some factors that influence documentation on death certificates and therefore affect the coding. We reviewed the role of documentation errors on the accuracy of death coding at Shahid Mohammadi Hospital (SMH), Bandar Abbas, Iran. We studied the death certificates of all deceased patients in SMH from October 2010 to March 2011. Researchers determined and coded the underlying cause of death on the death certificates according to the guidelines issued by the World Health Organization in Volume 2 of the International Statistical Classification of Diseases and Health Related Problems-10th revision (ICD-10). Necessary ICD coding rules (such as the General Principle, Rules 1–3, the modification rules and other instructions about death coding) were applied to select the underlying cause of death on each certificate. Demographic details and documentation errors were then extracted. Data were analysed with descriptive statistics and chi square tests. The accuracy rate of causes of death coding was 51.7%, demonstrating a statistically significant relationship (p=.001) with major errors but not such a relationship with minor errors. Factors that result in poor quality of Cause of Death coding in SMH are lack of coder training, documentation errors and the undesirable structure of death certificates.
Several clinical trials have shown the superiority of high-dose therapy (HDT) and autologous stem cell transplantation (ASCT) over conventional dose therapy for patients with multiple myeloma. There is limited experience with non-cryopreserved autologous hematopoietic stem cell transplantation. Between January 2004 and February 2010, we treated 38 patients with myeloma (mean age = 50.6) with a preparative regimen of Melphalan 140-200 mg/m(2) and non-cryopreserved ASCT. All the apheresis products were kept in a conventional blood bank refrigerator at 4°C for 2 d before infusion. The median time to platelet count of >20 × 10(9) /L was 13 d (range 10-31). Also, the median time to absolute neutrophil count >0.5 × 10(9) /L was 11 d (range 9-21). All the 38 patients were engrafted and there was not graft failure in this study group. Twenty-nine of 38 (76.3%) patients are alive and without disease activity after a median follow-up of 31 months (range 6-77). Responses (complete and partial response) were seen in all the 38 patients. The median progression-free survival was 27 months with 95% confidence interval 23.52-30.48, whereas the median overall survival was 30 months. One hundred days transplant-related mortality was 0%. HDT and autologous transplantation without cryopreservation is an effective and safe method which simplifies the procedure and is feasible and cost saving in our patients.
The role of tumor-infiltrating T cell subsets in the prognosis of non-Hodgkin's lymphoma (NHL) has previously been reported. In the present study, we investigated the prognostic significance of different T cell subsets in the peripheral blood of NHL patients. Immunophenotyping was performed on the peripheral blood samples of 45 patients with newly diagnosed B cell NHL using flow cytometry. The relationship between T cell subsets of CD4+, CD8+, CD3+CD25+, CD4+CD25+, CD4+CD25high [as T regulatory cells (T reg)], and the CD4/CD8 ratio with international prognostic index (IPI) and response to therapy was determined. The percentages of CD3+, CD4+, and CD8+ T cells in the peripheral blood of the patients were 49.1±20.3%, 23.6±11%, and 31.4±14.4%, respectively (CD4/CD8 ratio: 0.92±0.6). There were 4.2±3.2% T reg cells. A study of the percentage of T cells in relation to IPI score showed a higher proportion of CD3+CD25+, CD4+, and CD4+CD25+ cells in low-risk patients compared with intermediate/high risk groups (p<0.05). The above cells, as well as CD4+CD25high T reg cells, indicated a positive correlation with complete remission (CR) and survival. CD4 positivity correlated significantly with survival and CR durations, which were longer in patients with ≥20% CD4+ cells than those with <20% CD4+ cells, thus indicating the prognostic value of CD4+ T cells in NHL patients. There was no significant data on CD8+ cells as well as the CD4/CD8 ratio between distinct IPI groups and response to therapy. These data indicated the importance of CD4+ cells and the activation status of T cells in immunity against lymphoma and the prognostic implication of enumeration of these cells in NHL patients.
Objective:To compare the healing process of pressure ulcers treated with cryopreserved human amniotic membrane allograft and routine pressure ulcer care in our hospital. Methods: From January 2012 to December 2013, in a prospective randomized clinical trial (IRCT201612041335N2), 24 patients with second and third stage of pressure ulcers were enrolled in this study. All patients needed split thickness skin grafts for pressure ulcer-wound coverage. Selected patients had symmetric ulcers on both upper and lower extremities. The patients were randomly divided into two groups: amnion and control. In the amnion group, the ulcer was covered with cryopreserved amniotic membrane and in the control group it was treated with local Dilantin powder application. The duration and success rate of complete healing was compared between the two groups. Results: The study group was composed of 24 pressure ulcers in 24 patients (19 males and 5 females) with a mean age of 44±12.70 years. The demographic characteristics, ulcer area, and underlying diseases were similar in both groups. The early sign of response, such as decrease in wound discharge, was detected 12-14 days after biological dressing. Complete pressure ulcer healing occurred only in the amnion group (p<0.001). Partial healing was significantly higher in the amnion group (p<0.03). Healing time in this group was faster than that the control group (20 days versus 54 days). No major complication was recorded with amniotic membrane dressing. Conclusion: Cryopreserved amniotic membrane is an effective biologic dressing that promotes reepithelialization in pressure ulcers.
TaqMan™ genotyping assays are widely used to genotype CYP2D6, which encodes a major drug metabolizing enzyme. Assay design for CYP2D6 can be challenging owing to the presence of two pseudogenes, CYP2D7 and CYP2D8, structural and copy number variation and numerous single nucleotide polymorphisms (SNPs) some of which reflect the wild-type sequence of the CYP2D7 pseudogene. The aim of this study was to identify the mechanism causing false-positive CYP2D6*15 calls and remediate those by redesigning and validating alternative TaqMan genotype assays. Among 13,866 DNA samples genotyped by the CompanionDx® lab on the OpenArray platform, 70 samples were identified as heterozygotes for 137Tins, the key SNP of CYP2D6*15. However, only 15 samples were confirmed when tested with the Luminex xTAG CYP2D6 Kit and sequencing of CYP2D6-specific long range (XL)-PCR products. Genotype and gene resequencing of CYP2D6 and CYP2D7-specific XL-PCR products revealed a CC>GT dinucleotide SNP in exon 1 of CYP2D7 that reverts the sequence to CYP2D6 and allows a TaqMan assay PCR primer to bind. Because CYP2D7 also carries a Tins, a false-positive mutation signal is generated. This CYP2D7 SNP was also responsible for generating false-positive signals for rs769258 (CYP2D6*35) which is also located in exon 1. Although alternative CYP2D6*15 and *35 assays resolved the issue, we discovered a novel CYP2D6*15 subvariant in one sample that carries additional SNPs preventing detection with the alternate assay. The frequency of CYP2D6*15 was 0.1% in this ethnically diverse U.S. population sample. In addition, we also discovered linkage between the CYP2D7 CC>GT dinucleotide SNP and the 77G>A (rs28371696) SNP of CYP2D6*43. The frequency of this tentatively functional allele was 0.2%. Taken together, these findings emphasize that regardless of how careful genotyping assays are designed and evaluated before being commercially marketed, rare or unknown SNPs underneath primer and/or probe regions can impact the performance of PCR-based genotype assays, including TaqMan. Regardless of the test platform used, it is prudent to confirm rare allele calls by an independent method.
Conclusions: These data demonstrate the safety and feasibility of BEAM-like regimen as a new and modified regimen; longer follow-up is required to evaluate fully efficacy and long-term safety of our method.
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