Transcritos de fusión del gen PML/RARα en pacientes con leucemia promielocítica / Fusion transcripts of the PML/RARα gene in patients with promyelocytic leukemia

Introducción: La leucemia promielocítica se considera una enfermedad bien definida por sus peculiares características clínicas, morfológicas, citogenéticas y moleculares. El descubrimiento de los mecanismos oncogenéticos implicados en la génesis de la enfermedad hacen de esta variante de leucemia uno de los modelos más relevantes de investigación traslacional. Objetivo: Caracterizar los transcritos de fusión del gen PML/RARα en pacientes con leucemia promielocítica. Métodos: Se realizó una investigación observacional, ambispectiva, descriptiva, longitudinal, en pacientes con diagnóstico de leucemia promielocítica, en el Instituto de Hematología e Inmunología, entre enero de 2001 y diciembre de 2020. El universo estuvo constituido por 105 pacientes que cumplieron los criterios de inclusión y exclusión. Resultados: No existe relación entre los transcritos y la edad, sexo, color de piel y las características clínicas. La presencia del transcrito de fusión bcr3 se asoció a mayores cifras de hemoglobina y menor valor de plaquetas. La incidencia de recaída no se relacionó con los transcritos de fusión y no se comprobó que existiera influencia de éstos, sobre la supervivencia global en pacientes con leucemia promielocítica. Conclusiones: Las características de los transcritos de fusión del gen PML/RARα son similares a los reportes internacionales, sobre todo en poblaciones de origen latino(AU)

Introduction: Promyelocytic leukemia is considered a well-defined entity due to its peculiar clinical, morphological, cytogenetic and molecular characteristics. The discovery of the oncogenetic mechanisms involved in the genesis of the disease makes this variant of leukemia one of the most relevant models for translational research. Objective: To characterize the fusion transcripts of the PML/RARα gene in patients with promyelocytic leukemia. Methods: An observational, ambispective, descriptive, longitudinal investigation was carried out in patients diagnosed with promyelocytic leukemia at the Institute of Hematology and Immunology, between January 2001 and December 2020. The sample consisted of 105 patients who met the criteria for inclusion and exclusion. Results: There is no relationship between the transcripts and age, sex, skin color and clinical characteristics. The presence of the bcr3 fusion transcript was associated with higher hemoglobin levels and lower platelet counts. The incidence of relapse was not related to fusion transcripts and their influence on overall survival in patients with promyelocytic leukemia was not proven. Conclusions: The fusion transcripts´scharacteristicsof the PML/RARα gene are similar to international reports, especially from populations of Latin origin(AU)

¿Es importante conocer el tipo de transcripto BCR-ABL en la leucemia mieloide crónica? / Is it important to know the type of BCR-ABL transcript in chronic myeloid leukemia?

La leucemia mieloide crónica se caracteriza por la ocurrencia de una translocación recíproca entre los cromosomas 9 y 22; que da origen a un cromosoma 22 derivativo conocido como Filadelfia. En el sitio de unión se forma el gen de fusión BCR-ABL que conlleva a la síntesis de una proteína híbridacon propiedades oncogénicas. El sitio de unión entre los cromosomas 9 y 22 es variable y da lugar a transcritos diferentes; los conocidos como e13a2 y e14a2 son los más frecuentes y estudiados. El análisis de las características clínico-hematológicas de presentación y la respuesta al tratamiento entre los pacientes portadores de e13a2 o e14a2 ha revelado diferencias que pueden ser útiles para la predicción del pronóstico. Se realizó una revisión de la literatura científica a través de PUBMED. Se analizó y resumió la información. Se evidencian diferentes características de presentación, pero no existe coincidencia entre todos los autores. Respecto al comportamiento de la respuesta al tratamiento con inhibidores de tirosina quinasa, algunos autores encuentran diferencias y algunos sugieren que puede tratarse de dos enfermedades diferentes. Puede ser importante conocer el tipo de transcripto BCR-ABL en la LMC ya que, al menos entre los dos más frecuentes, existen diferencias que pueden ser útiles en la predicción del pronóstico para el paciente, así como para el manejo del tratamiento(AU)

Chronic myeloid leukemia is characterized by the occurrence of a reciprocal translocation between chromosomes 9 and 22; which gives rise to a derivative chromosome 22 known as Philadelphia. At the binding site, the BCR-ABL fusion gene is formed, which leads to the synthesis of a hybrid protein with oncogenic properties. The binding site between chromosomes 9 and 22 is variable and gives rise to different transcripts; those known as e13a2 and e14a2 are the most frequent and studied. The analysis of the clinical-hematological characteristics of presentation and the response to treatment among patients with e13a2 or e14a2 has revealed differences that may be useful for the prediction of prognosis. To describe the different characteristics reported for one or another transcript and to know if it is important to know the type of transcript in the CML. A review of the scientific literature was carried out through PUBMED. The information was analyzed and summarized. Different presentation characteristics are evident but there is no coincidence between all the authors. Regarding the behavior of the response to treatment with tyrosine kinase inhibitors, some authors find differences and some suggest that it may be two different entities. It may be important to know the type of BCR-ABL transcript in CML cause, at least between the two most frequent, there are differences that may be useful in predicting the prognosis for the patient as well as for the management of treatment(AU)

Casos atípicos e14a3 (b3a3) en el gen de fusión BCR-ABL de la leucemia mieloide crónica en Cuba / Rare cases of e14a3 (b3a3) BCR-ABL fusion in chronic myeloid leukemia in Cuba

Introducción: La leucemia mieloide crónica es un desorden clonal maligno de células madres hematopoyéticas pluripotentes que se caracteriza por la presencia del cromosoma Filadelfia, consecuencia de la traslocación cromosómica recíproca entre los brazos largos de los cromosomas 9 y 22. El resultado de esta alteración cromosómica es un gen de fusión que contiene las uniones b2a2 (e13a2) o b3a2 (e14a2). En la mayor parte de los casos, las células de la leucemia mieloide crónica expresan uno de los dos transcritos (b2a2 o b3a2); sin embargo, el 5 por ciento de los pacientes tienen ambos tipos de ARNm como resultado de empalmes alternativos. Se han encontrado otros transcriptos como e19a2, e2a2, e1a3, e6a2, e13a3(b2a3), y e14a3(b3a3), que ocurren con menos frecuencia. Objetivo: Describir el comportamiento de dos pacientes con leucemia mieloide crónica que presentan un trascripto BCR/ABL atípico. Casos clínicos: En el estudio molecular por reacción en cadena de la polimerasa cualitativo realizado a los dos pacientes, se observó un punto de ruptura del gen de fusión BCR/ABL poco frecuente, el cual se correspondía al transcripto e14a3 (b3a3). Estos pacientes iniciaron tratamiento con mesilato de imatinib a dosis de 400 mg diarios. Al primer paciente a los dos meses de tratamiento se le detectó crisis blástica, por lo que se le cambió el tratamiento a nilotinib 400 mg diarios que mantiene hasta la actualidad. La segunda paciente mantuvo igual tratamiento, aunque en ocasiones ha sido necesario incorporar tratamiento citorreductor con hidroxiurea por presentar leucocitosis. Conclusiones: Los pacientes con BCR/ABL a3 presentan un curso más benigno de la enfermedad. Aunque en los pacientes estudiados no se observó una respuesta satisfactoria al tratamiento pues presentaron diversas complicaciones(AU)

Introduction: Chronic myeloid leukemia is a malignant clonal disorder of pluripotent hematopoietic stem cells and characterized by the presence of the Philadelphia chromosome, which is the product of a reciprocal translocation between the long arms of chromosomes 9 and 22. The result of this chromosomal alteration is a fusion gene that contains the e13a2 (b2a2) and e14a2 (b3a2) junctions. In most cases, chronic myeloid leukemia cells express one of the two transcripts (b2a2 or b3a2); however, 5 percent of patients have both types of mRNA, as a result of alternative junctions. Other transcripts have been identified, such as e19a2, e2a2, e1a3, e6a2, e13a3 (b2a3), and e14a3 (b3a3), which occur less frequently. Objective: To describe the behavior of two patients with chronic myeloid leukemia who have an atypical BCR-ABL transcript. Clinical cases: In a qualitative molecular study of polymerase chain reaction carried out with two patients, a BCR-ABL fusion gene breakpoint was observed, which corresponded to the e14a3 (b3a3) transcript. These patients started treatment with imatinib mesylate at a dose of 400mg/d. At two months, the first patient had a diagnose of blast crisis, so the treatment was changed to nilotinib at a dose of 400mg/d, which the patient maintained to date. The second patient maintained the same treatment, although it was sometimes necessary to incorporate cytoreductive treatment with hydroxyurea due to leukocytosis. Conclusions: Patients with BCR-ABL a3 present a more benign evolution of the disease. However, a satisfactory response to treatment was not observed in the patients studied, as long as they presented various complications(AU)

Development of transgenic cotton (Narasimha) using triple gene Cry2Ab-Cry1F-Cry1Ac construct conferring resistance to lepidopteran pest

Silkworm silk protein fibroin is widely exploited to develop novel silk-based biomaterials due to its stable b-sheetstructure, providing high crystallinity and tensile strength. The polymorphic behaviour of silk fibroin provides awindow to modulate its structural transitions during self-assembly for different functional outcomes. Most studiesare therefore mainly focused on formation of well-developed b-sheet structure and self-assembly of silk fibroinwhich are regulated by many parameters. Glyoxal, a highly reactive a-oxoaldehyde, reacts with different proteinsto form advanced glycation end products (AGEs) following Maillard-like reaction. Considering the significanceof protein modification by glyoxal-derived AGEs, in the present study the effect of glyoxal (250, 500 and1000 lM) on the structure of silk fibroin has been investigated. CD and fluorescence studies reveal that higherconcentrations of the a-oxoaldehyde induce considerable alterations of secondary and tertiary structure of theprotein leading to aggregation following incubation with for 3 weeks. The aggregates exhibit fibrillar morphologywith amyloidal nature as evident from SEM, FTIR and XRD experiments. The findings highlight that glycationinduced modification can be a possible approach for modulating the conformation of the silk protein which may berelevant in connection to clinical, biomedical or synthetic biology based applications.

Analysis of the transcripts encoding for antigenic proteins of bovine gammaherpesvirus 4

Transcriptome profiling and digital gene expression analysis of genes associated with salinity resistance in peanut

Background: Soil salinity can significantly reduce crop production, but the molecular mechanism of salinity tolerance in peanut is poorly understood. A mutant (S1) with higher salinity resistance than its mutagenic parent HY22 (S3) was obtained. Transcriptome sequencing and digital gene expression (DGE) analysis were performed with leaves of S1 and S3 before and after plants were irrigated with 250 mM NaCl. Results: A total of 107,725 comprehensive transcripts were assembled into 67,738 unigenes using TIGR Gene Indices clustering tools (TGICL). All unigenes were searched against the euKaryotic Ortholog Groups (KOG), gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases, and these unigenes were assigned to 26 functional KOG categories, 56 GO terms, 32 KEGG groups, respectively. In total 112 differentially expressed genes (DEGs) between S1 and S3 after salinity stress were screened, among them, 86 were responsive to salinity stress in S1 and/or S3. These 86 DEGs included genes that encoded the following kinds of proteins that are known to be involved in resistance to salinity stress: late embryogenesis abundant proteins (LEAs), major intrinsic proteins (MIPs) or aquaporins, metallothioneins (MTs), lipid transfer protein (LTP), calcineurin B-like protein-interacting protein kinases (CIPKs), 9-cis-epoxycarotenoid dioxygenase (NCED) and oleosins, etc. Of these 86 DEGs, 18 could not be matched with known proteins. Conclusion: The results from this study will be useful for further research on the mechanism of salinity resistance and will provide a useful gene resource for the variety breeding of salinity resistance in peanut.

Heat-Killed Saccharomyces cerevisiae, A Dectin-1 Agonist, Selectively Induces IgG4 Production by Human B Cells

Dectin-1 is a major receptor that recognizes fungal cell wall β-glucan. We previously reported that heat-killed Saccharomyces cerevisiae (HKSC), a Dectin-1 agonist, selectively induces IgG1 class switching in mouse B cells. Dectin-1 is also expressed on human B cells; however, Dectin-1 function in human B cells remains unknown. This study aimed to investigate the direct effect of in vitro stimulation using HKSC on Ig class switching in human B cells. HKSC selectively induced the expression of germline γ4 transcripts (GLTγ4) by human B cell line 2E2, and HKSC significantly augmented GLTγ4 promoter activity. Moreover, HKSC selectively enhanced GLTγ4 expression and IgG4 production by anti-CD40-activated human tonsillar resting B cells. Thus, these results suggest that Dectin-1 maybe involved in selective IgG4 class switching by human B cells.

Sperm chromatin and DNA integrity, methyltransferase mRNA levels, and global DNA methylation in oligoasthenoteratozoospermia / 대한생식의학회지

OBJECTIVE: To investigate sperm chromatin/DNA integrity, global DNA methylation, and DNMT mRNA transcription in men with oligoasthenoteratozoospermia (OAT) compared with normozoospermic men. METHODS: Semen samples from 32 OAT patients who comprised the case group and 32 normozoospermic men who comprised the control group were isolated and purified using a standard gradient isolation procedure according to World Health Organization criteria. DNMT1, DNMT3A, and DNMT3B transcripts were then compared between groups using real-time quantitative reverse-transcription polymerase chain reaction. Global DNA methylation in sperm was determined by an enzyme-linked immunosorbent assay. Protamine deficiency and the proportion of apoptotic spermatozoa were evaluated using chromomycin A3 (CMA3), aniline blue (AB), and toluidine blue (TB) staining, as well as the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. The p-values < 0.05 were considered to indicate statistical significance. RESULTS: Significantly higher proportions of AB+, TB+, CMA3+, and TUNEL+ spermatozoa, as well as DNMT3A and DNMT3B transcription, were found in the OAT group. Positive correlations were detected between sperm parameters, DNA/chromatin damage, and DNMT3A and DNMT3B transcripts. Global DNA methylation was significantly higher in the OAT patients and had a significant correlation with abnormal results of all sperm chromatin integrity tests, but was not associated with DNMT1, DNMT3A, or DNMT3B expression. CONCLUSION: Oligoasthenoteratozoospermic men showed abnormal sperm parameters, abnormal chromatin/DNA integrity, and a higher global DNA methylation rate, as well as overexpression of DNMT mRNA.

Antisense transcription regulates the expression of sense gene via alternative polyadenylation

Natural antisense transcripts (NAT) and alternative polyadenylation (APA) of messenger RNA (mRNA) are important contributors of transcriptome complexity, each playing a critical role in multiple biological processes. However, whether they have crosstalk and function collaboratively is unclear. We discovered that APA enriched in human sense-antisense (S-AS) gene pairs, and finally focused on RNASEH2C-KAT5 S-AS pair for further study. In cis but not in trans over-expression of the antisense KAT5 gene promoted the usage of distal polyA (pA) site in sense gene RNASEH2C, which generated longer 3' untranslated region (3'UTR) and produced less protein, accompanying with slowed cell growth. Mechanistically, elevated Pol II occupancy coupled with SRSF3 could explain the higher usage of distal pA site. Finally, NAT-mediated downregulation of sense gene's protein level in RNASEH2C-KAT5 pair was specific for human rather than mouse, which lacks the distal pA site of RNASEH2C. We provided the first evidence to support that certain gene affected phenotype may not by the protein of its own, but by affecting the expression of its overlapped gene through APA, implying an unexpected view for understanding the link between genotype and phenotype.

Expression of androgen receptor and embryonic stem cell associated transcript 4 in breast cancer patients with over-expression of human epidermal growth factor receptor 2 and their clinical significances / 肿瘤研究与临床

Objective To investigate the expression and clinical significance of androgen receptor (AR) and embryonic stem cell associated transcripts 4 (NANOG) in breast cancer patients with human epidermal growth factor receptor 2 (HER-2) over-expression, and to analyze its relationship with clinicopathologic features of breast cancer. Methods 143 breast cancer patients with HER-2 over-expression were selected from the screening of 1052 cases of invasive breast cancer according to estrogen receptor (ER), progesterone receptor (PR) and HER-2 status. The protein expression of AR and NANOG was assayed by using immunohistochemistry.The relationship between AR expression and clinicopathological features was analyzed by χ2 test. The correlation between AR expression and NANOG expression was analyzed by Spearman correlation analysis. Results The positive expression of AR was 35.7 % (51/143). The AR expression was not associated with age and menstruation status (both P>0.05), and was associated with tumor size, clinical TNM staging and lymphatic metastasis (all P< 0.05). The positive rate of NANOG were 53.1 %(76/143), and NANOG proteins were negative in adjacent normal breast tissue and benign breast lesions. The positive rate of AR was 27.6%(21/76) in NANOG-positive cases, whereas the positive rate of AR was 44.8%(30/67) in NANOG-negative cases, and the difference was statistically significant (χ2=4.526, P=0.033). There was an inverse correlation between NANOG and AR expressions (r= -0.255, P= 0.002). Conclusion AR and NANOG may be new targets for endocrine therapy and molecular biological therapy.

Effect of downregulation of FBI-1 on proliferation of human breast carcinoma cell line and its mechanism / 中国癌症杂志

Background and purpose: Factor that binds to the inducer of short transcripts of human immuno-deficiency virus-1 (FBI-1) in a variety of malignant tumors showed high expression levels, which may be closely related to tumor proliferation and differentiation, angiogenesis, metastasis, but its relationship with breast cancer has not been fully elucidated. The purpose of this study was to investigate the expression of FBI-1 in breast cancer cells, and to study the effect of FBI-1 gene expression on the proliferation of breast cancer cells and its possible mechanism. Methods:Real-time fluorescent quantitative polymerase chain reaction (RTFQ-PCR) and Western blot analysis were applied to detect FBI-1 expression in normal human mammary epithelial cell line MCF-10A and breast cancer cell MCF-7. RNA interference method was used to down-regulate FBI-1 expression in MCF-7 cells. The cell proliferation was measured by CCK-8 kit and colony formation assay. RTFQ-PCR and Western blot were used to detect the expression of FBI-1 and NF-κBp65 in MCF-7 cells before and after the interference of FBI-1 expression. Results: The expression of FBI-1 was higher in breast cancer cells than that in normal human mammary epithelial cells (P<0.05). The effects of FBI-1 down-regulation inhibited proliferation in MCF-7 cells (P<0.05). At the same time, after inhibition of FBI-1, the NF-κBp65 mRNA and protein expression levels were significantly decreased (P<0.05). Conclusion: FBI-1 is highly expressed in breast cancer cells. Down-regulated FBI-1 expression can inhibit the proliferation of breast cancer cells,and its mechanism may be related to the inhibition of NF-κB signaling pathway.

Simple multiplex RT-PCR for identifying common fusion BCR-ABL transcript types and evaluation of molecular response of the a2b2 and a2b3 transcripts to Imatinib resistance in North Indian chronic myeloid leukemia patients.

INTRODUCTION: Chronic myeloid leukemia (CML) is characterized by the Philadelphia chromosome, an abnormally shortened chromosome 22. It is the result of a reciprocal translocation of chromosomes 9 and 22, creating BCR‑ABL fusion transcripts, b3a2, b2a2, and e1a2. The aim of our study was to determine the type of BCR‑ABL fusion transcripts for molecular diagnosis and investigate the frequency of BCR‑ABL fusion transcripts in CML patients by multiplex RT‑PCR in CML. MATERIALS AND METHODS: A single reaction with multiple primers multiplex PCR was used to detect and investigate the type and frequency in 200 CML patients among which 116, 33, and 51 were in CP, AP, and BC phase, respectively. RESULTS: The study included 200 CML patients, among whom breakpoints in b3a2, b2a2 transcripts were detected in 68% and 24%, respectively, while 8% of the patients showed both b3a2/b2a2. A statistically significant difference was seen between frequency of BCR‑ABL fusion transcripts and gender (P = 0.03), molecular response (P = 0.04), and hematological response (P = 0.05). However, there was no correlation found between frequencies of BCR‑/ABL fusion transcripts and other clinicopathological parameters like age, type of therapy, thrombocytopenia, and white blood cell count. CONCLUSION: Multiplex reverse transcriptase‑polymerase chain reaction is useful and saves time in the detection of BCR‑ABL variants; the occurrence of these transcripts associated with CML can assist in prognosis and treatment of disease.

DDRT-PCR approaches applied for preeminent results in the isolation of DETs from fish brain tissues / Aplicação de abordagens de DDRT-PCR para aprimoramento de resultados no isolamento de DETs a partir de tecido cerebral de peixes

Differential Display (DD) is a technique widely used in studies of differential expression. Most of these analyses, especially those involving fish species, are restricted to species from North America and Europe or to commercial species, as salmonids. Studies related to South American fish species are underexplored. Thus, the present work aimed to describe DD technique modifications in order to improve outcomes related to the isolation of DETs (Differentially Expressed Transcripts), using Leporinus macrocephalus, a large commercially exploited South American species, as a fish design. Different DDRT-PCR approaches were applied to brain samples and the products of the reactions were analyzed on 6% polyacrylamide gels stained with 0.17% Silver Nitrate (AgNO3). The use of PCR reactions under high stringency conditions and longer oligonucleotides based on VNTR (Variable Number of Tandem Repeats) core sequences led to better results when compared to low stringency PCR conditions and the use of decamer oligonucleotides. The improved approach led to the isolation of differentially expressed transcripts on adult males and females of L. macrocephalus. This study indicates that some modifications on the DDRT-PCR method can ensure isolation of DETs from different fish tissues and the development of robust data related to this approach.

Display Diferencial (DD) é uma técnica amplamente utilizada em estudos de expressão diferencial. A maioria desses estudos envolvendo espécies de peixes está restrita a espécies da América do Norte e Europa ou a espécies comerciais, como os salmoniformes. Estudos relacionados a peixes da América do Sul são ainda pouco explorados. Desse modo, o presente trabalho teve como objetivo descrever modificações na técnica de DD, a fim de melhorar os resultados relacionados ao isolamento de DETs (Transcritos Diferencialmente Expressos), utilizando Leporinus macrocephalus, peixe explorado comercialmente na América do Sul, como espécie para tal delineamento. Diferentes abordagens de DDRT-PCR foram desenvolvidas a partir de amostras de tecido cerebral e os produtos das reações foram analisados em gel de poliacrilamida 6% corados com 0,17% de nitrato de Prata (AgNO3). A utilização de reações de PCR sob condições de elevada estringência e oligonucleotídeos mais longos, com base em sequências cerne de VNTR (Número Variável de Repetições em Tandem), mostrou melhores resultados quando comparada a condições de baixa estringência e ao uso de oligonucleotídeos decâmeros. A estratégia empregada permitiu o isolamento de transcritos diferencialmente expressos em machos e fêmeas adultos de L. macrocephalus. Este estudo evidencia que modificações no método de DDRT-PCR garantem o melhor isolamento de DETs a partir de diferentes tecidos de peixes e asseguram a obtenção de dados mais sólidos relacionados a essa abordagem.

Ginsenoside Rg1 and 20(S)-Rg3 Induce IgA Production by Mouse B Cells

Ginsenosides are the major components of ginseng, which is known to modulate blood pressure, metabolism, and immune function, and has been used to treat various diseases. It has been reported that ginseng and several ginsenosides have immunoregulatory effects on the innate and T cell-mediated immune response. However, their effects on the humoral immune response have not been fully explored. The present study examined the direct effects of red ginseng extract (RGE) and ginsenosides on mouse B cell proliferation and on antibody production and the expression of germline transcripts (GLT) by mouse B cells in vitro. RGE slightly reduced B cell proliferation, but increased IgA production by LPS-stimulated B cells. Furthermore, ginsenoside Rg1 and 20(S)-Rg3 selectively induced IgA production and expression of GLTalpha transcripts by LPS-stimulated B cells. Collectively, these results suggest that ginsenoside Rg1 and 20(S)-Rg3 can drive the differentiation of B cells into IgA-producing cells through the selective induction of GLTalpha expression.

SUMO Proteins are not Involved in TGF-beta1-induced, Smad3/4-mediated Germline alpha Transcription, but PIASy Suppresses it in CH12F3-2A B Cells

TGF-beta induces IgA class switching by B cells. We previously reported that Smad3 and Smad4, pivotal TGF-beta signal-transducing transcription factors, mediate germline (GL) alpha transcription induced by TGF-beta1, resulting in IgA switching by mouse B cells. Post-translational sumoylation of Smad3 and Smad4 regulates TGF-beta-induced transcriptional activation in certain cell types. In the present study, we investigated the effect of sumoylation on TGF-beta1-induced, Smad3/4-mediated GLalpha transcription and IgA switching by mouse B cell line, CH12F3-2A. Overexpression of small ubiquitin-like modifier (SUMO)-1, SUMO-2 or SUMO-3 did not affect TGF-beta1-induced, Smad3/4-mediated GLalpha promoter activity, expression of endogenous GLalpha transcripts, surface IgA expression, and IgA production. Next, we tested the effect of the E3 ligase PIASy on TGF-beta1-induced, Smad3/4-mediated GLalpha promoter activity. We found that PIASy overexpression suppresses the GLalpha promoter activity in cooperation with histone deacetylase 1. Taken together, these results suggest that SUMO itself does not affect regulation of GLalpha transcription and IgA switching induced by TGF-beta1/Smad3/4, while PIASy acts as a repressor.

Cytogenetic & molecular analyses in adult chronic myelogenous leukaemia patients in north India.

Background & objectives: Chronic myelogenous leukaemia (CML) is the commonest leukaemia in Asia. There is a paucity data on cytogenetic and molecular analyses of Indian CML patients. This apparently reflects the low availability of cytogenetic and molecular techniques in our country. This study aimed to document various types of BCR-ABL fusion transcripts in different phases of CML and to compare the Ph chromosome positivity/negativity vis-a-vis BCR-ABL fusion transcripts in adult CML patients. Methods: Between June 2004 and February 2009, 208 patients were diagnosed as CML in chronic phase (CP), accelerated phase (AP) and blast crisis (BC), according to standard criteria. Cytogenetic and molecular genetic analyses were performed in all patients. Various types of BCR-ABL hybrid transcripts were compared with phases of CML and cytogenetic abnormalities. Results: Among 208 CML patients, b3a2 BCR-ABL transcripts were most commonly detected (66.82%) followed by b2a2 (28.84%), b3a2 + b2a2 (3.36%), b3a2 + e19a2 (0.48%) and b2a2 + e19a2 (0.48%). b3a2 transcripts were more frequently detected than b2a2 transcripts, in the whole group of 208 as well as in 183 CML-CP patients (P<0.0001). Ph chromosome was positive in 135 of 139 patients with b3a2 transcripts and 56 of 60 patients with b2a2 transcripts, difference not being significant. Additional cytogenetic abnormalities detected in 3.8 per cent patients in CML-CP and 44 per cent patients in CMLAP/ BC, did not show predilection for any BCR-ABL transcript type. Interpretation & conclusions: This study documents higher Ph positivity (96.15%) by cytogenetic analysis among CML patients, as confirmed by qualitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis in a large patient group from north India. Both the techniques contribute towards understanding the disease biology, and have important implications for diagnosis and management of CML patients.

Effects of herpes simplex virus 2 latency-associated transcript open reading frame 3 on the apoptosis in Vero cells / 中华皮肤科杂志

Objective To explore the effects of herpes simplex virus 2 (HSV-2) latency-associated transcript open reading frame 3 (LAT ORF3) gene on Vero cells against cisplatin-induced apoptosis.Methods Recombinant plasmid enhanced green fluorescent protein-open reading frame 3 (named pEGFP-ORF3) was constructed and transfected into Vero cells; then,reverse transcription (RT)-PCR was performed to detect the expression of the target gene.Cisplatin of 3 mg/L was selected to induce the apoptosis in Vero cells.Cultured Vero cells were transfected with empty plasmid and induced by cisplatin (pEGFP-C2 group),transfected with recombinant plasmid pEGFP-ORF3 and induced by cisplatin (pEGFP-ORF3 group),only induced by cisplatin (cisplatin-induced control group),or remained untreated (normal control group).Subsequently,fluorescence microscopy was conducted to observe apoptotic bodies,Giemsa stain to observe the morphology of cell nuclei,methyl thiazolyl tetrazolium (MTT) assay to evaluate cell proliferation,and flow cytometry to assess cell apoptosis.Data were assessed by using SPSS 13.0 software,and statistical analysis was carried out by one-way ANOVA and t test.Results HSV-2 333 LAT ORF3 gene was successfully cloned.The eukaryotic expression plasmid for LAT ORF3 was constructed,and the expression of LAT ORF3 gene in Vero cells was confirmed by RT-PCR.Giemsa stain showed blue-staining nuclei and pale cytoplasm in recombinant plasmid-transfected and cisplatin-induced Vero cells with a normal shape.The value of cell proliferation (absorbance at 490 nm) by MTT assay was 2.56 ± 0.21 in pEGFP-ORF3 group,similar to that in the normal control group (2.66 ± 0.13,P ＞ 0.05),but significantly higher than cisplatin-induced control group (1.65 ± 0.11,P ＜ 0.05) and pEGFP-C2 group (1.56 ± 0.18,P ＜ 0.05).As far as the apoptosis rate was concerned,no significant difference was observed between pEGFP-ORF3 group and normal control group (4.03％ ± 1.04％ vs.2.13％ ± 0.09％,P ＞ 0.05),but pEGFP-ORF3 group was statistically lower than pEGFP-C2 group (19.45％ ± 2.05％,P ＜ 0.05).Conclusion The transfected HSV-2 LAT ORF3 gene could protect Vero cells from cisplatin-induced apoptosis.

Expression of ILT3 and ILT4 in dendritic cells of kidney transplantation recipients and its significance / 中华器官移植杂志

Objective To study the expression of immunoglobulin-like transcripts 3 (ⅡT3) and ILT4 in peripheral blood dendritic cells (DC) of kidney transplantation recipients and to analyze its significance in immunity hyporesponsiveness of transplantation. Methods Twenty kidney allograft recipients who were survived more than five years were recruited to two groups: renal function stable groups, chronic rejection groups, and 10 healthy volunteers served as a control group. The peripheral blood mononuclear cells (PBMC) were stimulated with granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin 4 (IL-4) and immature DC were obtained. The expression of ILT3 and ILT4 was detected by using flow cytometry. The level of HLA-G5 in serum was determined by using enzyme linked immunosorbent assay. Results ILT3 expression in renal function stable group was increased and decreased in chronic rejection groups as compared with control group (P＜0.05),but ILT4 expression had no significant difference among all groups. HLA-G5 in serum was significantly increased in renal function stable group as compared with other groups. Conclusion Expression of ILT3 and HLA-G was increased in the kidney transplantation recipients with stable renal function and long-term survival, suggesting that they may play an important role in inducing and maintaining periphery immune tolerance.

Análise da expressão de RNAs não-codificadores intrônicos em câncer de rim / Expression analyses of intronic non-coding RNAs in renal cancer

O carcinoma de célula renal (RCC) subtipo célula clara é o câncer mais letal e prevalente do sistema urinário. O diagnóstico deste tipo de câncer frequentemente é tardio em conseqüência da falta de sintomas perceptíveis aos pacientes. Um dos objetivos deste trabalho é a identificação de novos marcadores moleculares para diagnóstico precoce, o que ajudaria a diminuir a mortalidade em função de complicações resultantes do avanço da doença. Outro objetivo é a identificação de um conjunto de marcadores moleculares de prognóstico, de modo à prever com acurácia a evolução clínica da doença e, por conseqüência, o tempo de sobrevida do paciente. As modificações transcricionais associadas à carcinogênese e à progressão do câncer de rim ainda não foram completamente elucidadas. Além dos oncogenes e genes supressores de tumor, RNAs não-codificadores (ncRNAs) recentemente foram apontados como importantes reguladores da expressão gênica em humanos, e podem ter um papel importante na transformação maligna do câncer de rim. Para analisar a expressão gênica de ncRNAs e de genes codificadores para proteína foram utilizados dois microarranjos desenvolvidos por nosso grupo, enriquecidos em sondas para ncRNAs. Uma das plataformas possui 4 mil sondas de cDNA, das quais 822 sondas são para ncRNAs mapeando em regiões intrônicas. Outra possui 44 mil elementos e combina sondas de oligonucleotídeos (60-mer) intrônicas e exônicas de um mesmo locus genômico. Análises estatísticas foram feitas com a ferramenta Significance Analysis of Microarrays (q ≤ 0,05) combinadas ou com a técnica de "patient leave-one-out" (genes com presença em 8 100% dos subconjuntos), ou alternativamente com o teste discriminante de Golub (p ≤ 0,01 ou p < 0,05). Com a plataforma de 4 mil sondas foram estudadas 30 amostras de tecido renal de 18 pacientes com RCC subtipo célula clara. Um conjunto de 36 ncRNAs foi identificado como diferencialmente expresso entre amostras tumorais e não-tumorais...

Renal cell carcinoma (RCC) is the most common malignancy of the adult kidney, and the clear cell subtype is the most prevalent and lethal cancer of the urinary system. Late diagnosis for this type of cancer is frequent, usually as a consequence of the lack of symptoms. One of the objectives of the present work is the identification of new molecular markers for the early diagnosis, which would help decrease mortality that develops as a function of disease progression. Another objective is the identification of a set of prognosis molecular markers, so as to accurately predict the clinical outcome of the disease, and consequently, patient survival. Transcriptional changes associated to carcinogenesis and to kidney cancer progression have not been entirely elucidated. Besides oncogenes and tumor suppressor genes, non-coding RNAs (ncRNAs) have been recently indicated as important regulators of gene expression in humans, and could have an important role in the malignant transformation in renal cancer. In order to measure ncRNA and protein-coding gene expression we have used two microarray platforms developed by our group, which are enriched in ncRNA probes. One of the platforms has 4 thousand cDNA probes, of which 822 are for ncRNAs that map to intronic regions. Another has 44 thousand elements and combines 60-mer oligonucleotide probes for intronic and exonic regions from the same genomic locus. Statistical analyses have been performed with the Significance Analysis of Microarrays tool (q ≤ 0.05) combined with a patient leave-one-out approach (genes present in 100% of the sub-sets), or alternatively with Golubs discriminant test (p ≤ 0.01 or p < 0.05). 11 With the 4-thousand probes platform we studied 30 samples from renal tissue of 18 RCC patients with clear cell subtype. A set of 36 ncRNAs has been identified as differentially expressed between tumor and non-tumor tissue...

Latencia del heppesvirus bovino-1: el papel de los transcritos relacionados con latencia (rl) / Bovine Herpesvirus-1: The Role of Latency-Related Genes

El herpesvirus bovino-1 es un virus de distribución mundial causante de graves pérdidas económicas debidas principalmente a la disminución de la eficiencia y en los indicadores de salud y productividad de cualquier hato ganadero infectado. Luego de la infección inicial del tracto respiratorio de los animales, el virus establece un estado de latencia viral en las neuronas sensoriales del ganglio trigémino y en los centros germinales de las tonsilas faríngeas. Periódicamente, el virus es reactivado y excretado en secreciones a través de las cuales puede infectar a otros animales susceptibles. Durante dicho estado de latencia hay disminución dramática de la expresión de genes virales, llevando solo a la expresión de dos transcritos: El RNA codificado por el gen relacionado con latencia (RL) y el ORF-E viral. Múltiples estudios demuestran como el RL y el ORF-E están involucrados en la regulación del complejo ciclo de latencia y reactivación de la infección. La presente revisión de literatura se enfocará en describir y analizar los distintos estudios que han llevado a dilucidar el papel jugado por el gen RL y el ORF-E, sus transcritos y sus productos proteicos en el establecimiento, mantenimiento y reactivación de la latencia del HVB-1.

Bovine herpesvirus-1 is a world wide spread virus that causes significant economic losses due mainly to a decrease in the efficiency and in the health and productivity indicators in all the infected herds. After a primary infection of the respiratory tract of the animals, the virus establishes viral latency state in sensory neurons of trigeminal ganglia and germinal centers of pharyngeal tonsils. Periodically, the virus reactivates from latency, is shed through secretions, and can infect other susceptible animals. During latency there is a dramatic reduction of viral gen expression; only two transcripts are abundantly expressed: the latency related (LR) RNA and the viral ORF-E. Multiple studies have shown LR and ORF-E role in the regulation of BHV-1 latencyreactivation cycle. This review focuses on the description and analysis of the litherature that had lead to incriminate LR gene and viral ORF-E, their transcripts and protein products in the establishment, maintenance and reactivation of BHV-1 latency state.