Contribution of heme oxygenase 2 to blood pressure regulation in response to swimming exercise and detraining in spontaneously hypertensive rats

Background: We aimed to determine the effects of exercise followed by detraining on systolic blood pressure (SBP), heme oxygenase 2 (HO-2) expression, and carboxyhemoglobin (COHb) concentration in spontaneously hypertensive rats (SHR) to explain the role of carbon monoxide (CO) in this process. Material/Methods: Animals were randomized into exercised and detrained groups. Corresponding sedentary rats were grouped as Time 1–2. Swimming of 60 min/5 days/week for 10 weeks was applied. Detraining rats discontinued training for an additional 5 weeks. Gene and protein expressions were determined by real-time PCR and immunohistochemistry. Results: Aorta HO-2 histological scores (HSCORE) of hypertensive rats were lower, while SBP was higher. Swimming caused enhancement of HO-2 immunostaining in aorta endothelium and adventitia of SHR. Exercise induced elevation of blood COHb index in SHR. Synchronous BP lowering effect of exercise was observed. HO-2 mRNA expression, HSCORE, and blood COHb index were unaltered during detraining, while SBP was still low in SHR. Conclusions: CO synthesized by HO-2 at least partly plays a role in SBP regulation in the SHR-and BP-lowering effect of exercise. Regular exercise with short-term pauses may be advised to both hypertensives and individuals who are at risk. © Med Sci Monit

Possible role of GADD45γ methylation in diffuse large B-cell Lymphoma: Does it affect the progression and tissue involvement?

Objective: Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma among adults and is characterized by heterogeneous clinical, immunophenotypic, and genetic features. Different mechanisms deregulating cell cycle and apoptosis play a role in the pathogenesis of DLBCL. Growth arrest DNA damage-inducible 45 (GADD45γ) is an important gene family involved in these mechanisms. The aims of this study are to determine the frequency of GADD45γ methylation, to evaluate the correlation between GADD45γ methylation and protein expression, and to investigate the relation between methylation status and clinicopathologic parameters in DLBCL tissues and reactive lymphoid node tissues from patients with reactive lymphoid hyperplasia. Materials and Methods: Thirty-six tissue samples of DLBCL and 40 nonmalignant reactive lymphoid node tissues were analyzed in this study. Methylation-sensitive high-resolution melting analysis was used for the determination of GADD45γ methylation status. The GADD45γ protein expression was determined by immunohistochemistry. Results: GADD45γ methylation was frequent (50.0%) in DLBCL. It was also significantly higher in advanced-stage tumors compared with early-stage (p=0.041). In contrast, unmethylated GADD45γ was associated with nodal involvement as the primary anatomical site (p=0.040). Conclusion: The results of this study show that, in contrast to solid tumors, the frequency of GADD45γ methylation is higher and this epigenetic alteration of GADD45γ may be associated with progression in DLBCL. In addition, nodal involvement is more likely to be present in patients with unmethylated GADD45γ. © 2015 Turkish Society of Hematology. All rights reserved

Comparıson of exosomal and nuclear dna methylatıon paterns ın dıffuse large b-cell lymphoma

Bu çalışma, PAÜ Bilimsel Araştırma Projeleri Koordinasyon Birimi tarafından desteklenmiştir (Proje No: 2016SABE005).Diffüz Büyük B-Hücreli Lenfoma (DBBHL) agresif lenfomaların en yaygın tipidir ve Hodgkin-dışı lenfomaların yaklaşık %30-40’ını oluşturmaktadır. DBBHL’nin moleküler patogenezi kompleks ve çok basamaklı mekanizmaları içermektedir. Bu mekanizmalardan bir bölümü aydınlatılmış olsa da, henüz hastalığın tam olarak patogenezi bilinmemektedir.Kanser hücrelerinden köken alan eksozomların tümör oluşumu, progresyonu ve yayılımına neden olacak önemli modülatörleri içerdikleri iyi bilinmektedir. Bu çerçevede, bu çalışma DBBHL’de primer tümörde EZH2-hedef genlerinde gözlenen epigenetik değişimlerin sirkülasyondaki eksozomlarda da belirlenebilirliğini değerlendirmek amacı ile dizayn edildi.Çalışmada 21 DBBHL hastası ve 21 sağlıklı gönüllü bireyin plazma eksozomları izole edildi. Eksozomlardan ve primer tümör dokularından DNA izolasyonu yapılarak, metilasyon-spesifik PCR ile hedef genlerdeki metilasyon durumları belirlendi. Aynı zamanda izole edilen RNA örnekleri kullanılarak, eksozomların ve primer tümör doku örneklerinin hedef genlere ait transkriptleri içerip içermedikleri belirlendi. EZH2’de mutasyonun varlığı DNA dizi analizi ile incelendi.Primer tümörlerle uyumlu olarak DBBHL eksozom örneklerinin CDKN1A ve CDKN1B unmetile DNA’sını ve CDKN2A ve CDKN2B metile DNA’sını içerdikleri belirlendi. DBBHL FFPE doku örneklerinin tümünde EZH2 mRNA'sının varlığı gözlenmişken karşılıkları olan eksozom örneklerinde söz konusu transkriptin varlığına rastlanmadı. FFPE doku örneklerinin 12’sinde (%57) CDKN1A; 9’unda (%43) CDKN2A mRNA'sının varlığı belirlenirken, aynı örneklerin eksozom karşılıklarında CDKN1A ve CDKN2A mRNA’larına rastlanmadı. FFPE doku örneklerinin tümünde CDKN1B; 3’ünde (%14) CDKN2B transkriptinin varlığı belirlenmişken karşılıkları olan eksozom örneklerinin %38’inin CDKN1B transkriptini içerdikleri ve hiçbirinin CDKN2B transkripti içermediği belirlendi. Ayrıca, primer tümörle uyumlu olarak eksozom örneklerinin EZH2 Y641 mutasyonunu taşımadığı saptandı.Bu çalışma, DBBHL-kökenli eksozomlarda DNA’nın varlığını gösteren ilk çalışmadır. Bu çalışma DBBHL’de plazma eksozomlarının primer tümörle uyumlu DNA fragmanlarını içerdiklerini gösteren ilk çalışmadır. DBBHL’de plazma eksozomlarının primer tümörden tercihli olarak özgün hedef moleküllerini paketlediklerini ve CDKN1B ile CDKN2A ve CDKN2B’nin rol oynadığı yolaklar üzerinden lenfomageneze katıldıklarını düşünmekteyiz.Diffuse Large B-Cell Lymphoma (DLBCL) is the most common type of aggressive lymphoma, and accounts for approximately 30-40% of non-Hodgkin's lymphomas. The molecular pathogenesis of DLBCL is a complex, multistep process. Although recent advances in our understanding of the process have been significant, the pathogenesis still remains to be elucidated.It is well known that exosomes derived from cancer cells are able to transfer important modulators for tumor formation, progression and spread. In this respect, this study was designed to evaluate if epigenetic changes in EZH2-targeted genes in primary tumor are also observed in circulating exosomes.This study included 21 healthy volunteers and 21 DLBCL patients and then plasma exosomes were isolated. After DNA isolation from the exosomes and primary tumor tissue samples, methylation-specific PCR was used to determine methylation status of the target genes. It was also determined whether exosomes and primary tumor tissue samples contained transcripts of the target genes using isolated RNA samples. DNA sequencing was used to determine the presence of mutation in EZH2.In concordance with the primary tumors, CDKN1A and CDKN1B unmetile DNAs and CDKN2A and CDKN2B methylated DNAs were determined in the exosomes isolated from DLBCL patients. EZH2 transcript was found in all primary tumor samples of DLBCL patients but no in the exosome counterpart. CDKN1A and CDKN2A transcrits were determined in 12 (57%) and 9 (43%) of the FFPE tumor samples, respectively whereas these transcripts were not found in the exosomes. CDKN1B transcript was determined in all FFPE tumor samples while CDKN2B transcript was determined in 3(14%) of the tumor samples. In the exosome counterpart, the presence of CDKN1B transcript was observed in 38% of the samples while there was no transcript for CDKN2B. In addition to, EZH2 Y641 mutation was not detected in both exosome samples and primary tumor samples counterparts. This study is the first to show that exosomes included DNA fragments which is in concordance with primary tumors. We thought that specific target molecules were preferentially sorted into exosomes and plasma exosomes may contibute to lymphomagenesis, at least in part, by pathogenic pathways including CDKN1B, CDKN2A and CDKN2B

Determination of GADD45gamma methylation by high resolution melting analysis in diffuse large B-Cell Lymphoma and evaluation of the possible correlation with apoptosis

Diffüz Büyük B-Hücreli Lenfoma (DBBHL), yetişkin bireylerde Hodgkin dışı lenfomaların en yaygın tipi olmakla birlikte klinik, immunofenotipik ve genetik özellikler açısından heterojen bir yapı göstermektedir. DBBHL'nın patogenezinde, hücre döngüsü ve apoptotik yollarda düzensizliğe neden olan farklı mekanizmalar rol oynamaktadır. 'Growth Arrest DNA Damage-Inducible 45(GADD45)' gen ailesi bu mekanizmalarda yer alan önemli bir gen ailesidir. Bu çalışmanın amaçları DBBHL doku örnekleri ve reaktif lenfoid doku örneklerinde i) GADD45gama'nın metilasyon sıklığını belirlemek, ii) metilasyon durumu ile protein ekspresyon düzeyi arasındaki korelasyonu değerlendirmek, ve iii) GADD45gama'nın metilasyon ve ekspresyon durumlarının apoptozla olan ilişkisi belirlemektir. Bu çalışmada, 40 DBBHL ve 40 reaktif lenfoid doku (RLD) örnekleri değerlendirildi. GADD45gama metilasyon durumu HRM analiziyle, GADD45gama, Bcl-2 ve Bcl-XL ekspresyonları immunohistokimya ve apoptotik indeks (AI) değerleri TUNEL yöntemi ile belirlendi. DBBHL'de GADD45gama metilasyon sıklığı yüksek (%55) bulundu. Aynı zamanda, ileri evre olgularda GADD45gama metilasyonunun, erken evre olgulara oranla yüksek olduğu belirlendi (P=0.041). GADD45gama aşırı ekspresyonu DBBHL?li ve RLD?li olgularda sırası ile %45 ve %5 olarak belirlendi ve fark anlamlı bulundu (P =0.000). İki olgu grubu arasında Bcl-2 ekpresyonu açısından anlamlı fark belirlenirken (P =000), Bcl-xL protein ekspresyonu açısından anlamlı bir ilişki bulunamadı (P =0.154). Aynı zamanda, iki grup arasında AI değerleri açısından anlamlı bir farklılık yoktu (P =1.000). Bu çalışmadan elde edilen veriler; i) solid tümörlerin aksine, DBBHL'de GADD45gama metilasyon sıklığının yüksek olduğunu, ii) GADD45gama geninde gözlenen bu epigenetik değişimin DBBHL'nin progresyonunda rol oynayabileceğini ve iii) DBBHL'de GADD45gama'nın, tek başına hücreleri apoptoza yönlendirmediğini göstermektedir.Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma among adults and is characterized by heterogeneous clinical, immunophenotypic and genetic features. Different mechanisms deregulating cell cycle and apoptosis play a role in the pathogenesis of DLBCL. 'Growth Arrest DNA Damage-Inducible 45(GADD45)' is an important gene family involved in these mechanisms. The aims of this study are; i) to determine the frequency of GADD45gamma promoter methylation, ii) to evaluate the correlation between GADD45gamma methylation and protein expression and, iii) to investigate the relation between apoptosis and GADD45gamma methylation and protein expression in DLBCL tissues and reactive lymphoid node tissues (RLT). Forty tissue samples from patients with DLBCL and 40 RLT from patient with reactive lymphoid hyperplasia were investigated in this study. It was used HRM analysis for determination of GADD45gamma methylation status, immunohistochemistry for protein expression of GADD45gamma, Bcl-2 and Bcl-xL, and TUNEL method for apoptotic index (AI). It was found that the frequency of GADD45gamma methylation was high (55%) in DLBCL. The methylation frequency for GADD45gamma was also significantly higher in advanced-stage compared with early-stage (P=0.041). Overexpression of GADD45gamma was found 45% and 5% in DLBCL and RLT groups, respectively and the significant association was observed (P=0.000). There was a significant association of Bcl-2 expression between both groups (P=0.000) while no significant difference was observed in expression of Bcl-xL (P=0.154). In addition, there was no significant association of AI between both groups. The results of this study show that i) in contrast to solid tumors, the frequency of GADD45gamma methylation is higher in DLBCL, ii) this epigenetic alteration of GADD45gamma may associate with DLBCL progression, and iii) GADD45gamma is not the only key factor for the cells to direct into the apoptotic process

GCN5 participates in ADA3 partnership as revealed with yeast hybrid

41st FEBS Congress on Molecular and Systems Biology for a Better Life -- SEP 03-08, 2016 -- Kusadasi, TURKEYWOS: 000383616900265FEB

Mutational status of EZH2 and CD79B hot spots in mature B-cell non-Hodgkin's lymphomas: Novel CD79B variations have been revealed

OBJECTIVE: We aimed to determine the hot spot mutational frequencies of Enhancer of Zeste homolog 2 (EZH2) and cluster of differentiation 79B (CD79B) genes in a cohort of mature B-cell non-Hodgkin's lymphomas. PATIENTS AND METHODS: DNA samples from formalin-fixed and paraffin embedded (FFPE) tissues from a total of 37 patients with mature B-cell non-Hodgkin lymphomas were included in the study. Molecular genetic analysis was performed by direct sequencing of the DNA samples. RESULTS: We analyzed formaldehyde fixed-paraffin embedded (FFPE) tumor tissue samples from 17 female and 20 male patients with a median age of 63.7 years at the time of diagnosis. None of the patients had previously reported hot spot mutations in EZH2 and CD79B, but previously unreported single nucleotide variations of CD79B were present in nine patients. rs779833118 was the most frequent variation (7/37 patients, 18.9%). A non-synonymous variation rs757407417, which could have a potentially damaging outcome, was detected in two patients. CONCLUSIONS: None of the patients had well-known hot spot mutations in EZH2 and CD79B. However, we detected novel CD79B variations in mature B-cell non-Hodgkin's lymphoma patients. © 2016, Verduci Editore. All rights reserved