scholarly journals A Novel RHCE*ce Variant Allele Is Highly Frequent in Chinese D-Negative Individuals and Results in C-Expression

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2892-2892
Author(s):  
Tamara C Stegmann ◽  
Y L Ji ◽  
Lonneke Haer-Wigman ◽  
L Wei ◽  
Y Zhao ◽  
...  
Keyword(s):  
Conflicts Of Interest ◽  
Lentiviral Vector ◽  
Variant Allele ◽  
The Novel ◽  
Facs Analysis ◽  
Exon 2 ◽  
Chinese Populations ◽  
New Variant ◽  
Novel Variant ◽  
Dependent Probe

Abstract Background: The RHCE allele is highly polymorphic and many variants have been described, especially in individuals of African origin. Donors carrying these variants can be falsely typed and elicit transfusion reactions, and patients carrying such a variant may be at risk to develop allo-antibodies in response to mismatched transfusions. Not much is known about the frequency of RHCE variants in Chinese populations, whereas in China genotyping assays are increasingly applied for typing of blood donors and patients. Methods: Standard column agglutination was used to serologically type for C/c and E/e expression in 200 serologically D-negative and 200 serologically D-positive Chinese donors. The RH Multiplex Ligation-dependent Probe Amplification (MLPA) genotyping assay was used for genotyping the RHCE status (Transfusion 2013;53:1559). In donors with discrepant results of genotyping and phenotyping all 10 exons of RHCE were amplified and directly sequenced. A lentivirus containing the novel RHce variant was created to transduce human erythroblasts cultured from peripheral blood from 3 ccDee and 3 CCDee donors as previously described (Haematologica 2010;95:1594). FACS analysis was used to assess the c- and C-expression caused by the variant. Results: In 6 out of 200 Chinese D-negative donors the results of the RH-MLPA indicated the presence of only one copy of exon 2 of the CE*c-allele, and no copy of exon 2 of the CE*C-allele, whereas these donors were serologically typed as Cc. Sequencing of all 10 RHCE exons revealed a novel RHCE*ce allele defined by 308C>T (p.103Pro>Leu) mutation next to a normal RHCE*ce allele. The variant allele was not found in the 200 Chinese D-positive donors, indicating the linkage of this new variant RHCE*ce allele with the D-negative haplotype. Wild type Rhce cDNA was mutated to create the RHce*308C>T mutation and subsequently cloned into a lentiviral vector. Transduction of human ccDee erythroblasts with this vector resulted in C expression, whereas virtually no c-expression was induced by transduction of human CCDee erythroblasts as assessed by FACS analysis with the monoclonals MS33, MS35 and MS42 to detect c expression and monoclonals MS24 and MS273 to detect C. Discussion: A new RHCE variant (RHCE*ce308T) is identified, which is present in 3% of D-negative Chinese individuals. The 308C>T mutation in the triplet encoding the 103Pro results in the expression of 103Leu. Position 103 is one of the 4 aminoacid differences between the c- and C-carrying polypeptides (Pro and Ser, respectively). The Pro>Leu mutation in the novel variant leads to C-expression and loss or strongly diminished c-expression. Most Rhc-genotyping assays target the c-specific-307C nucleotide and most RhC-genotyping assays target the C-specific-intron 2, which are respectively present and absent in this variant allele. Therefore, when individuals carrying this allele are genotyped, the predictive phenotype will be falsely C-negative. Conclusion: RHC-genotyping assays applied in Chinese populations, should be adapted to recognize the presence of this new RHCE*ce308T allele to prevent C-mismatched transfusions. Disclosures No relevant conflicts of interest to declare.

Reduced C/EBPα Expression Favors Monopoiesis Over Granulopoiesis

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1230-1230
Author(s):  
Alan D. Friedman ◽  
Ou Ma ◽  
SunHwa Hong
Keyword(s):  
Liquid Culture ◽  
Leucine Zipper ◽  
Conflicts Of Interest ◽  
Lentiviral Vector ◽  
Facs Analysis ◽  
Lineage Specification ◽  
Dna Elements ◽  
Marrow Cells ◽  
Myeloid Progenitors

Abstract Abstract 1230 C/EBPα is required for the formation of the granulocyte-monocyte progenitor; however, its role in subsequent lineage specification remains uncertain. We previously compared G-CSF with M-CSF signaling in murine marrow, finding that G-CSF more potently induces phosphorylation of SHP2 whereas M-CSF more potently activates ERK. Knockdown (KD) of SHP2 impaired granulopoiesis relative to monopoiesis and unexpectedly led to reduced C/EBPα RNA. A. Cantor and colleagues recently reported that SHP2 dephosphorylates Src kinase-modified RUNX1 and increases the ability of RUNX1 to induce megakaryopoiesis, and we demonstrated that in myeloid cells RUNX1 binds and activates the CEBPA gene, via its promoter and via a conserved 450 bp enhancer located at +37 kb containing 4 RUNX1-binding sites. Thus, G-CSF or other cytokines potentially direct SHP2 to activate RUNX1 which then increases C/EBPα transcription in myeloid progenitors to favor granulopoiesis. As a test of this model, we have now transduced lineage-depleted murine marrow cells, cultured in SCF, TPO, and Flt3L to minimize myeloid differentiation, with shRNAs targeting CEBPA or with a control lentiviral vector. Western blotting indicated 2-fold KD by shRNAs B9 or B11 after 2 days of transduction followed by 2 days of puromycin selection. As this analysis was done on a mixed population, greater KD may occur in rare progenitors. Upon transfer to IL3, IL6, and SCF to favor myeloid maturation, vector-transduced cells produced 4-fold more CFU-G than CFU-M in methylcellulose, whereas CEBPA KD resulted in 4- or 5-fold more CFU-M than CFU-G. FACS analysis of pooled CFUs demonstrated significant reduction of Mac-1+Gr-1+ granulocytes and an increase in Mac-1+Gr-1− monocytes, and examination of cell morphology after Wright's-Giemsa staining further confirmed a shift towards monopoiesis in response to reduced C/EBPα. Similar findings were obtained for cells placed in liquid culture. In addition, on day 3 of liquid culture Q-RTPCR analysis showed that CEBPA shRNAs induced 2- or 3-fold CEBPA RNA KD, reduced granulocytic MPO, NE, PR3, and GCSFR RNAs, increased monocytic MCSFR and CD14 RNAs, and increased expression of EGR1, IRF8, and c-Jun RNAs, encoding transcription factors that help direct monopoiesis. RUNX1 RNA was unchanged, and PU.1 RNA was reduced <2-fold. To determine the effect of CEBPA KD in vivo, transduced, puromycin-selected CD45.2+ marrow cells were transplanted into lethally irradiated, syngeneic CD45.1+ recipients together with CD45.1+ carrier cells. At day 28, FACS analysis indicated a shift towards monopoiesis amongst shRNA B9- or B11-transduced marrow cells, and CFU assay of sorted CD45.2+ cells demonstrated significantly reduced CFU-G with increased CFU-M. Besides affecting myeloid lineage determination, CEBPA KD increased the rate of cell proliferation in liquid culture, associated with blasts evident upon morphologic analysis and an increase in Mac-1−Gr-1− cells, which mainly consisted of c-Kit+Sca-1− myeloid progenitors. In addition, CEBPA KD by shRNA B9, which impaired granulopoiesis more effectively than shRNA B11, increased CFU replating to 8 generations, compared with 4 generations for control cells. Analysis on day 3 of liquid culture did not reveal an alteration in the proportion of cells in G1, S, or G2/M or increased apoptosis, and c-Myc and Bcl-2 RNA levels were unaffected. We propose that CEBPA KD not only strongly impairs granulopoiesis, but also modestly impedes monopoiesis to produce a transient accumulation of cells with blast morphology and the ability to proliferate. Overall, our findings suggest that relative to monopoiesis, granulopoiesis requires increased level or increased activity of C/EBPα. Offering a potential mechanism, we previously found that C/EBPα :AP-1 leucine zipper heterodimers are more abundant than C/EBPα homodimers during monopoiesis compared with granulopoiesis, in part due to c-Fos, c-Jun, and JunB induction, and that C/EBPα :AP-1 heterodimers can direct marrow monopoiesis. Perhaps higher levels of C/EBPα favor homodimer formation, with reduced levels still able to heterodimerize with AP-1 proteins. ERK modification of C/EBPα on S21, as occurs downstream of M-CSF, reduces its ability to direct granulopoiesis; phospho-C/EBPα (S21) potentially retains the ability to direct AP-1 proteins to hybrid DNA elements recently identified in numerous monocytic enhancers. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Alternative 3' acceptor site in the exon 2 of human PAX8 gene resulting in the expression of unknown mRNA variant found in thyroid hemiagenesis and some types of cancers.

2013 ◽  
Vol 60 (4) ◽  
Author(s):  
Ewelina Szczepanek-Parulska ◽  
Witold Szaflarski ◽  
Katarzyna Piątek ◽  
Bartłomiej Budny ◽  
Karolina Jaszczyńska-Nowinka ◽  
...  
Keyword(s):  
Acceptor Site ◽  
The Novel ◽  
Exon 2 ◽  
Thyroid Hemiagenesis ◽  
Mrna Variant ◽  
Different Types ◽  
Novel Variant ◽  
Pax8 Gene ◽  
Alternatively Spliced ◽  
First Time

PAX8 gene encodes one of the transcription factors engaged in the regulation of proper development of thyroid gland as well as Müllerian and renal/upper urinary tracts. So far, six alternatively spliced transcripts were reported, however, sequences of only four were deposited in the NCBI database. Here, we evaluate a fragment of a novel variant of PAX8 mRNA formed by an alternative 3' acceptor site located in the second exon. The molecular outcome encompasses extension of the 5' untranslated region of exon two by 97 nucleotides as is evident from mRNA. This new insert may impair binding of mRNA to the ribosome and in consequence significantly decrease expression of the PAX8 protein. Here, we show for the first time that the novel insert in exon two might be associated with congenital thyroid hemiagenesis and influence development of different types of cancer.

scholarly journals A novel multi-word paradigm for investigating semantic context effects in language production

2019 ◽  
Author(s):  
Cornelia van Scherpenberg ◽  
Rasha Abdel Rahman ◽  
Hellmuth Obrig
Keyword(s):  
Eye Tracking ◽  
Picture Naming ◽  
Language Production ◽  
Context Effects ◽  
Semantic Context ◽  
The Novel ◽  
Target Picture ◽  
Novel Variant ◽  
Lexical Interference ◽  
Respective Category

Semantic context modulates precision and speed of language production. Using different experimental designs including the Picture-Word-Interference (PWI) paradigm, it has consistently been shown that categorically related distractor words (e.g., cat) inhibit retrieval of the target picture name (dog). Here we introduce a novel variant of the PWI paradigm in which we present 8 words prior to a to be named target picture. Within this set, the number of words categorically related was varied between 3 and 5, and the picture to be named was either related or unrelated to the respective category. To disentangle interacting effects of semantic context we combined different naming paradigms manipulating the number of competitors, and assessing the effect of repeated naming instances. Evaluating processing of the cohort by eye-tracking provided us with a metric of the (implicit) recognition of the semantic cohort. Results replicate the interference effect in that overall naming of pictures categorically related to the distractor set was slower compared to unrelated pictures. However, interference did not increase with increasing number of distractors. Tracking this effect across naming repetitions, we found that interference is prominent at the first naming instance of every picture only, whereby it is stable across distractor conditions, but dissipates across the experiment. Regarding eye-tracking our data show that participants fixated longer on semantically related items, indicating the identification of the lexico-semantic cohort. Our findings confirm the validity of the novel paradigm and indicate that besides interference during first exposure, repeated exposure to the semantic context may facilitate picture naming and counteract lexical interference.

scholarly journals Overexpression of Protein Kinase C Isoform  but not δ in Human Interleukin-3–Dependent Cells Suppresses Apoptosis and Induces bcl-2 Expression

Blood ◽  
1998 ◽  
Vol 91 (3) ◽  
pp. 823-829 ◽  
Author(s):  
E. Gubina ◽  
M.S. Rinaudo ◽  
Z. Szallasi ◽  
P.M. Blumberg ◽  
R.A. Mufson
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
The Novel ◽  
Facs Analysis ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Kinase C ◽  
Intracellular Ca ◽  
Cellular Dna ◽  
Stimulating Factor

Hematopoietic progenitor cells die by apoptosis after removal of the appropriate colony-stimulating factor (CSF). Recent pharmacologic data have implicated protein kinase C (PKC) in the suppression of apoptosis in interleukin-3 (IL-3) and granulocyte-macrophage (GM)-CSF–dependent human myeloid cells. Because IL-3 and GM-CSF induce increases in diacylglycerol without mobilizing intracellular Ca++, it seemed that one of the novel Ca++ independent isoforms of PKC was involved. We report here that overexpression of PKC in factor-dependent human TF-1 cells extends cell survival in the absence of cytokine. Overexpression of PKCδ does not have this effect. By 72 to 96 hours after cytokine withdrawal, the PKC transfectants remain distributed in all phases of the cell cycle, as shown by fluorescence-activated cell sorting (FACS) analysis, while little intact cellular DNA is detectable in vector or PKCδ transfectants. PKC induces bcl-2 protein expression fivefold to sixfold over the levels in empty vector transfectants, whereas the levels in PKCδ transfectants are similar to those in vector controls.

scholarly journals A Case Series: The Novel Use of Emapalumab and Ruxolitinib in Acquired Malignancy-Associated HLH in Pediatric Patients

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 31-31
Author(s):  
Kenny Hung Vo ◽  
Jong Chung ◽  
Arun Ranjan Panigrahi
Keyword(s):  
Induction Chemotherapy ◽  
Pediatric Patients ◽  
Conflicts Of Interest ◽  
Case Series ◽  
The Novel ◽  
Ifn Gamma ◽  
Novel Therapeutics ◽  
Genetic Components ◽  
Organ System Failure ◽  
Concurrent Processes

Hemophagocytic lymphohistiocytosis (HLH) is a potentially fatal hyperinflammatory syndrome that is classified into primary and acquired forms. Primary-HLH has genetic components related to defects in cytotoxic T and NK cells. The acquired form of HLH is classified based on the underlying triggers of immune dysregulation through malignancy, infections, and/or autoimmune processes. The standard of care is established with etoposide and dexamethasone, but there are currently no guidelines for refractory HLH or cases with confounding presentations with infection or malignancy. We describe cases of malignancy-associated HLH (m-HLH) in pediatric patients to discuss the intricacies in the initial diagnostic considerations, the balance of therapeutic regimens and their toxicities, and the novel use of emapalumab and ruxolitinib in refractory patients. We discuss a 16-year-old female with NK/T-cell lymphoma of the nasopharynx who presented in septic shock with pancytopenia and DIC. Diagnostic considerations included progressive lymphoma, bacteremia, and EBV viremia as possible explanations for her shock physiology. HLH was a consideration on admission given her presentation, but she did not meet criteria until HD#7 with ferritin of 740ng/mL (ref 10-291ng/mL), soluble IL-2 receptor (CD25) of 10,600pg/mL (ref &lt;1033pg/mL), sCD163 of 5,636ng/mL (ref 387-1,785ng/mL) and an abnormal NK cell function. Standard treatment with etoposide and dexamethasone showed minimal effect, as did the introduction of rituximab. Gemcitabine and oxaliplatin were started as treatment for her lymphoma as the driver of her HLH, but she developed acute decompensation with hemodynamic instability and multi-organ system failure. Her systemic inflammation worsened as evidenced by a rise in her ferritin to 15,462ng/mL and sIL-2 receptor to 28,700pg/mL. Her CXCL9, a downstream chemokine and marker of IFN-gamma activity shown to be elevated in HLH, had a dramatic increase to 106,918 pg/mL (ref &lt;121pg/mL). A decision to hold treatment for her lymphoma was made to reduce toxicities and better manage her HLH. Compassionate use of emapalumab, an IFN-gamma inhibitor indicated in primary HLH, led to dampening of her systemic inflammatory processes with resolution of fevers and signs of recovery of her hepatobiliary, renal, and gastrointestinal systems. We also describe a 17-year-old male who presented with T-ALL and bacteremia who developed HLH during induction-chemotherapy with complications from bacterial sepsis and fulminant fungemia. He represented a diagnostic conundrum throughout his induction chemotherapy with persistent fevers despite appropriate treatment and improvement of his bacteremia and fungemia. It was not until HD#50 that he met criteria for HLH with the addition of a ferritin level of 6,073ng/mL. He had limited response to treatment and had multi-system toxicities from the therapies for his HLH, ALL, and fungemia that necessitated frequent adjustments to his treatment plans. The treatment for his HLH was not standard given the toxicities. Etoposide was never in consideration given concerns for myelosuppression and dexamethasone was not given at full doses due to intolerance but was given over a prolonged duration of months. His course included a waxing and waning response to a combination of steroids and IVIG with recurrent admissions for presumed HLH flares. A trial of ruxolitinib, a JAK inhibitor, demonstrated response with stabilization of his hyperinflammatory state. Although not curative, as evidenced by his persistent hyperferritinemia, the use of ruxolitinib has allowed him to better tolerate his chemotherapy and improve his overall quality of life with a decrease in frequency of flares and hospitalizations. These cases highlight the intricacies of the management of m-HLH in pediatric patients with the spectrum of disease severity and complex diagnostic evaluations one must consider at presentation. Due to the toxicity of treatment options for often concurrent processes of malignancy, infection, and HLH, novel therapeutics such as emapalumab and ruxolitinib are required and should be evaluated in larger studies. Furthermore, guidelines for the management of pediatric m-HLH are required with inclusion of adjustments to standard therapy based on toxicity, inciting factors, concurrent processes, and the incorporation of novel therapeutics to achieve improved outcomes. Disclosures No relevant conflicts of interest to declare. OffLabel Disclosure: Ruxolitinib and emapalumab for refractory acquired HLH in Pediatric patients.

Novel somatic mutations in UBA1 as a cause of VEXAS syndrome

Blood ◽  
2021 ◽  
Author(s):  
James A Poulter ◽  
Jason Charles Collins ◽  
Catherine Cargo ◽  
Ruth M de Tute ◽  
Paul Evans ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Somatic Mutations ◽  
Clinical Phenotype ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
The Novel ◽  
Disease Mechanism ◽  
Bone Marrow Biopsies ◽  
Novel Variant

Somatic mutations at methionine 41 (Met41) in UBA1, encoding the major E1 enzyme responsible for initiating ubiquitylation, were recently identified as the cause of a novel autoinflammatory disease, named VEXAS (Vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic). We sought to determine the prevalence of UBA1 mutations in a UK cohort of patients matching the VEXAS clinical phenotype. We identified 10 new patients with somatic mutations in UBA1, but only 8 had altered p.Met41. A novel variant, c.167C&gt;T; p.Ser56Phe was identified, which was present in myeloid, and not lymphoid lineages and led to preferential loss of the catalytic activity of cytoplasmic UBA1. An additional novel variant, c.118-1G&gt;C was identified at the splice acceptor site of exon 3 leading to altered splicing in vitro. Bone marrow biopsies from two patients with a Met41 substitution and the novel splice site variant were consistent with previously reported features of VEXAS. The bone marrow of the patient with the p.Ser56Phe variant was less similar, likely driven by a distinct but overlapping disease mechanism. Our study therefore confirms somatic p.Met41 substitutions in UBA1 as a major cause of VEXAS syndrome and identifies two new disease causing mutations.

A novel variant in PAX6 as the cause of aniridia in a Chinese family

2020 ◽  
Author(s):  
Xin Jin ◽  
Wei Liu ◽  
HouBin Huang
Keyword(s):  
Nonsense Mutation ◽  
Novel Mutation ◽  
Causative Mutation ◽  
Chinese Family ◽  
The Novel ◽  
Targeted Next Generation Sequencing ◽  
Iris Defect ◽  
The Family ◽  
Novel Variant ◽  
Pax6 Mutation

Abstract Background: Aniridia is a kind of congenital human panocular anomaly, which is related to PAX6 commonly. Methods: A Chinese Aniridia pedigree underwent ophthalmic examinations, including visual acuity, slit lamp and fundoscopy examination. The targeted next-generation sequencing of Aniridia genes was used to identify the causative mutation. Results: A novel heterozygous PAX6 nonsense mutation c.619A>T (p.K207*) was identified in the Chinese autosomal dominant family with aniridia. Phenotypes related to the novel mutation include nystagmus, iris defect, cataract and absence of macular fovea. Conclusion: The novel nonsense mutation in PAX6 was responsible for aniridia phenotype in the family. which expands the spectrum of the PAX6 mutation and its associated phenotype.

scholarly journals Approximation algorithms for tours of orientation-varying view cones

2020 ◽  
Vol 39 (4) ◽  
pp. 389-401
Author(s):  
Nikolaos Stefas ◽  
Patrick A Plonski ◽  
Volkan Isler
Keyword(s):  
Approximation Algorithm ◽  
Ground Plane ◽  
Orientation Angle ◽  
Apex Angle ◽  
Field Of View ◽  
Plane Normal ◽  
Polynomial Time Approximation Algorithm ◽  
New Variant ◽  
Angle Difference ◽  
Novel Variant

This article considers the problem of finding a shortest tour to visit viewing sets of points on a plane. Each viewing set is represented as an inverted view cone with apex angle [Formula: see text] and height [Formula: see text]. The apex of each cone is restricted to lie on the ground plane. Its orientation angle (tilt) [Formula: see text] is the angle difference between the cone bisector and the ground plane normal. This is a novel variant of the 3D Traveling Salesman Problem with Neighborhoods (TSPN) called Cone-TSPN. One application of Cone-TSPN is to compute a trajectory to observe a given set of locations with a camera: for each location, we can generate a set of cones whose apex and orientation angles [Formula: see text] and [Formula: see text] correspond to the camera’s field of view and tilt. The height of each cone [Formula: see text] corresponds to the desired resolution. Recently, Plonski and Isler presented an approximation algorithm for Cone-TSPN for the case where all cones have a uniform orientation angle of [Formula: see text]. We study a new variant of Cone-TSPN where we relax this constraint and allow the cones to have non-uniform orientations. We call this problem Tilted Cone-TSPN and present a polynomial-time approximation algorithm with ratio [Formula: see text], where [Formula: see text] is the set of all cone heights. We demonstrate through simulations that our algorithm can be implemented in a practical way and that by exploiting the structure of the cones we can achieve shorter tours. Finally, we present experimental results from various agriculture applications that show the benefit of considering view angles for path planning.

scholarly journals Identification of a Modified HOXB9 mRNA in Breast Cancer

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Ayako Nakashoji ◽  
Tetsu Hayashida ◽  
Yuko Kawai ◽  
Masayuki Kikuchi ◽  
Rurina Watanuki ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Human Breast Cancer ◽  
Cancer Cell Line ◽  
Clinical Samples ◽  
The Novel ◽  
Rna Seq ◽  
Human Breast ◽  
Aberrant Expression ◽  
Novel Variant

First identified as a developmental gene, HOXB9 is also known to be involved in tumor biological processes, and its aberrant expression correlates with poor prognosis of various cancers. In this study, we isolated a homeodomain-less, novel HOXB9 variant (HOXB9v) from human breast cancer cell line-derived mRNA. We confirmed that the novel variant was produced from variationless HOXB9 genomic DNA. RT-PCR of mRNA isolated from clinical samples and reanalysis of publicly available RNA-seq data proved that the new transcript is frequently expressed in human breast cancer. Exogenous HOXB9v expression significantly enhanced the proliferation of breast cancer cells, and gene ontology analysis indicated that apoptotic signaling was suppressed in these cells. Considering that HOXB9v lacks key domains of homeobox proteins, its behavior could be completely different from that of the previously described variationless HOXB9. Because none of the previous studies on HOXB9 have considered the presence of HOXB9v, further research analyzing the two transcripts individually is warranted to re-evaluate the true role of HOXB9 in cancer.

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