The metabolomic profile of gamma-irradiated human hepatoma and muscle cells reveals metabolic changes consistent with the Warburg effect

The two human cell lines HepG2 from hepatoma and HMCL-7304 from striated muscle were γ-irradiated with doses between 0 and 4 Gy. Abundant γH2AX foci were observed at 4 Gy after 4 h of culture post-irradiation. Sham-irradiated cells showed no γH2AX foci and therefore no signs of radiation-induced double-strand DNA breaks. Flow cytometry indicated that 41.5% of HepG2 cells were in G2/M and this rose statistically significantly with increasing radiation dose reaching a plateau at ∼47%. Cell lysates from both cell lines were subjected to metabolomic analysis using Gas Chromatography-Mass Spectrometry (GCMS). A total of 46 metabolites could be identified by GCMS in HepG2 cell lysates and 29 in HMCL-7304 lysates, most of which occurred in HepG2 cells. Principal Components Analysis (PCA) showed a clear separation of sham, 1, 2 and 4 Gy doses. Orthogonal Projection to Latent Structures-Discriminant Analysis (OPLS-DA) revealed elevations in intracellular lactate, alanine, glucose, glucose 6-phosphate, fructose and 5-oxoproline, which were found by univariate statistics to be highly statistically significantly elevated at both 2 and 4 Gy compared with sham irradiated cells. These findings suggested upregulation of cytosolic aerobic glycolysis (the Warburg effect), with potential shunting of glucose through aldose reductase in the polyol pathway, and consumption of reduced Glutathione (GSH) due to γ-irradiation. In HMCL-7304 myotubes, a putative Warburg effect was also observed only at 2 Gy, albeit a lesser magnitude than in HepG2 cells. It is anticipated that these novel metabolic perturbations following γ-irradiation of cultured cells will lead to a fuller understanding of the mechanisms of tissue damage following ionizing radiation exposure

A Metabolomic Analysis of Cirrhotic Ascites.

Ascites is a common complication of decompensated liver cirrhosis, and yet relatively little is known about its biochemical composition. We conducted two metabolomic investigations, comparing the profile of ascites from 33 cirrhotic patients and postoperative peritoneal drainage fluid from 33 surgical patients (Experiment 1). The profile of paired ascites and plasma was also compared in 17 cirrhotic patients (Experiment 2). Gas chromatography-mass spectrometry-based metabolomics identified 29 metabolites that significantly characterized ascites fluid, whether postoperative drainage fluid or plasma were used as controls. Ten elevated amino acids (glutamine, proline, histidine, tyrosine, glycine, valine, threonine, methionine, lysine, phenylalanine) and seven diminished lipids (laurate, myristate, palmitate, oleate, vaccenate, stearate, cholesterol) largely comprised the cirrhotic ascites metabolomic phenotype that differed significantly (adjusted p < 0.002 to 0.03) from peritoneal drainage fluid or plasma. The pattern of upregulated amino acids in cirrhotic ascites did not indicate albumin proteolysis by peritoneal bacteria. Bidirectional clustering showed that the more severe the cirrhosis, the lower the lipid concentration in ascitic fluid. The metabolomic compartment of ascites in patients with decompensated cirrhosis is characterized by increased amino acids and decreased lipids. These novel findings have potential relevance for diagnostic purposes

Türkiye’nin çeşitli bölgelerinden temin edilen bal örneklerinde naftalin aranması ve miktar tayini

Türkiye çok zengin bir bitki örtüsü ve farklı iklim kuşaklarına sahip olup arıcılığın yaygın olarak yapıldığı bir ülkedir. Pestlerle mücadelede kullanılan pestisitlerin arıcılar tarafından yanlış ve bilinçsiz bir şekilde uygulanması sonucu bal ve balmumunda kalıntı bıraktığı bilinmektedir. Kovandan güveleri uzaklaştırmak amacıyla yapılan naftalin uygulama işlemi sonucu bal naftalinle kontamine olur. Naftalin, uçucu özellikte bir hidrokarbon olup genellikle güvelere karşı ve tuvalet deodorantları olarak, lubrikant ve boya üretiminde kullanılmaktadır. Naftalinin oral, inhalasyon ve dermal yollarla absorbsiyonu insanlarda sistemik toksik etkilere yol açmaktadır. Akciğerler ve gözler toksik açıdan en çok etkilenen organlardır. Bunların dışında diğer dokulardan, karaciğer, beyin ve böbrekte de toksik etkiler görülmektedir. Çalışmamızda, marketlerden ve pazarlardan temin edilmiş bal örneklerinde naftalin, diode dizi detektörlü (DAD) yüksek basınçlı sıvı kromatografisi (YBSK) ile tayin edildi. Bu çalışmada 100 tane bal örneğinde naftalin analizi yapıldı. Türkiye’nin çeşitli bölgelerinden temin edilmiş toplam 100 örneğin 53 tanesi marketlerden, 47 tanesi pazarlardan satın alındı. Naftalin şüpheli bir bal örneği gaz kromatografisi-kütle spektrometrisi (GC-MS) ile analiz edildi ve naftalin olduğu doğrulandı. Ülkemiz beslenmesinde çok önemli bir yer tutan ve yurt dışına giderek artan miktarlarda ihraç edilen ballar, aynı zamanda ekonomimiz açısından da önemlidir. Bu nedenle besinlerin tüm besin zinciri boyunca geçirdiği aşamalarda, naftalinle ve diğer kalıntılarla kontaminasyonun önlenmesi, hijyen bakımından kontrol ve analizlerinin yapılması gerekmektedir. Beslenmeyle ilgili yapılacak kalıntı çalışmaları sağlıklı bir toplum kazanmak ve sağlıklı nesillerin yetişmesi bakımından büyük önem taşır. SUMMARY Turkey having a wealthy flora with different climate conditions, beekeeping is widely spread all over the country. Due to wrong and/or unconscious pesticide treatments against pests generally causes residues in honey and beeswax. Treatment of naphthalene to beeswax is preferred for removing the moths from the beehive and as a result of it, honey is contaminated with naphthalene. Naphthalene, a volatile aromatic hydrocarbon, is widely used in mothballs and toilet bowl deodorants, in the production of dyes and lubricants. Naphthalene absorption by oral, inhalation and dermal contact causes systemic toxic effects for human. The lungs and eyes appear to be most susceptible to toxicity, although toxic effects can be demonstrated in other tissues, such as the liver, brain and kidney. In our study, naphthalene was detected in honey specimens which are provided from various regions of Turkey using high pressure liquid chromatography (HPLC) with diode array detector (DAD). In this study, naphthalene analysis was carried out in 100 honey specimens. Out of 100 specimens 53 of them were purchased from markets and 47 of them were from street bazaars existing in various regions of Turkey. Naphthalene-suspicious one honey specimen was analysed and confirmed by gas chromatography-mass spectrometry (GC-MS). Honey has an important role in our food chain and in our country’s economy. Therefore, fooodstuffs are needed to be hygienic controlled and analyzed during food process and prevent the contamination of naphthalene and other residues. All residue analyses for the entire food chain have a lot of importance for healthy people and healthy generation

İstanbul’da satılan işlenmiş hububat ve bakliyat ürünlerinde deoksinivalenol (vomitoksin) aranması.

Deoksinivalenol (DON, vomitoksin), hububat ve bakliyat gibi besin ürünlerinde F.graminearium, F.culmorum gibi fungal Fusarium cinsinin çeşitli türleri tarafından sentezlenen toksik kimyasal bir maddedir. Bu mikotoksinin oluşumu sıcaklık, nem, besin üretim aşamaları ve üretilen besinin tipi gibi birçok faktöre bağlıdır. Tarım ürünlerinde hasattan önce veya depolama sırasında oluşan, yemlerde de rastlanan DON'la kontaminasyon sonucu insan ve hayvanlarda vomitus ve besin reddi, immun sistem faaliyetinin yavaşlaması, gastrointestinal bozukluklar gibi pekçok hastalığa rastlanmaktadır.Çalışmamızda, marketlerden, pazarlardan temin edilmiş hububat ve bakliyat ürünlerinde DON araştırıldı. DON, UV detektörlü yüksek basınçlı sıvı kromatografisi (YBSK) ve ince tabaka kromatografisi (İTK) ile tayin edildi. DON aranmasında kullanılan iki yöntem de büyük kolaylıklar sağlamaktadır. İnce tabaka kromatografisi basit, pratik, hızlı ve ucuz olması nedeniyle tercih edilirken, yüksek basınçlı sıvı kromatografisi de hassasiyeti ve teşhis kolaylığı sebebiyle kullanılmaktadır.Bu çalışmada 68 tane işlenmiş hububat ve 15 tane bakliyat örneğinde DON analizi yapıldı. İstanbul'dan temin edilmiş bu toplam 83 örneğin 66 tanesi marketlerden, 17 tanesi semt pazarlarından satın alındı. YBSK ile DON tespit edilen örneklerden ý0.60 ppm miktarında bulunanların aynı zamanda İTK ile de analizleri yapıldı. En yüksek DON düzeyi pazardan temin edilen bir mısır unu örneğinde 2.67 ppm olarak saptandı. İTK ve YBSK çalışmalarının sonuçları birbirleriyle uyumlu bulundu. DON, hububat örneklerinin %8.82'sinde tespit edilmiş olup, bakliyat örneklerin de DON'a rastlanmamıştır.Ülkemiz beslenmesinde çok önemli bir yer tutan ve yurt dışına giderek artan miktarlarda ihraç edilen işlenmiş hububat ve bakliyat ürünleri aynı zamanda ekonomimiz açısından da önemlidir. Bu nedenle besinlerin tüm besin zinciri boyunca geçirdiği aşamalarda, mikotoksinlerle kontaminasyonun önlenmesi, hijyen bakımından kontrol ve analizlerinin yapılması gerekmektedir. Beslenmeyle ilgili yapılacak mikotoksin çalışmaları sağlıklı bir toplum kazanmak ve sağlıklı nesillerin yetişmesi bakımından büyük önem taşır. 2. SUMMARYDETERMINATION OF DEOXNIVALENOL IN PROCESSED CEREAL AND PULSE PRODUCTS PROVIDED FROM ISTANBUL MARKETDeoxynivalenol (DON, vomitoxin) is chemical compound which is synthesized by various species of the fungal genus Fusarium such as F. graminearium, F.culmorum, on foods like cereal, pulse and their products. The occurrence of this mycotoxin depends on such factors as temperature, humidity, food processing methods, and type of food product. DON which is capable of growing on agricultural products before harvest and during storage was also seen on feedstuffs. Contamination with DON causes a number of illness in man and animals such as decrease in food consumption (anorexia), depression or inhibition on immun system function and irritation of the gastrointestinal tract.In our study, DON was investigated with processed the cereal and pulse products which are provided from markets and street bazaars. DON was detected using thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) with UV detector. Two methods used for the detection of DON provide lots of facilities. TLC was preferred because of its easy, fast and inexpensive application and HPLC was used because of its facility in detection.In this study, DON analysis was carried out in 68 processed cereal and 15 pulse samples. Out of 83 samples, 66 of them were purchased from markets and 17 of them were from street bazaars existing in Istanbul. DON detected samples by HPLC at the level of 0.60 ppm were also analysed by TLC. The highest level of DON was determined as 2.67 ppm in a corn flour sample. DON was detected in 8.82% of cereal and none in pulse products.Cereal and pulse products which were also exported are very important in our food chain and in our country's economy. Therefore, foodstuffs are needed to be controlled and analysed during food process. All mycotoxin analyses for the entire food chain have a lot of importance for healthy people and healthy generations

Türkiye’nin çeşitli bölgelerinden temin edilen bal örneklerinde naftalin aranması ve miktar tayini

ÖZET Türkiye çok zengin bir bitki örtüsü ve farklı iklim kuşaklarına sahip olup arıcılığın yaygın olarak yapıldığı bir ülkedir. Pestlerle mücadelede kullanılan pestisitlerin arıcılar tarafından yanlış ve bilinçsiz bir şekilde uygulanması sonucu bal ve balmumunda kalıntı bıraktığı bilinmektedir. Kovandan güveleri uzaklaştırmak amacıyla yapılan naftalin uygulama işlemi sonucu bal naftalinle kontamine olur. Naftalin, uçucu özellikte bir hidrokarbon olup genellikle güvelere karşı ve tuvalet deodorantları olarak, lubrikant ve boya üretiminde kullanılmaktadır. Naftalinin oral, inhalasyon ve dermal yollarla absorbsiyonu insanlarda sistemik toksik etkilere yol açmaktadır. Akciğerler ve gözler toksik açıdan en çok etkilenen organlardır. Bunların dışında diğer dokulardan, karaciğer, beyin ve böbrekte de toksik etkiler görülmektedir.Çalışmamızda, marketlerden ve pazarlardan temin edilmiş bal örneklerinde naftalin, diode dizi detektörlü (DAD) yüksek basınçlı sıvı kromatografisi (YBSK) ile tayin edildi. Bu çalışmada 100 tane bal örneğinde naftalin analizi yapıldı. Türkiye’nin çeşitli bölgelerinden temin edilmiş toplam 100 örneğin 53 tanesi marketlerden, 47 tanesi pazarlardan satın alındı. Naftalin şüpheli bir bal örneği gaz kromatografisi-kütle spektrometrisi (GC-MS) ile analiz edildi ve naftalin olduğu doğrulandı. Ülkemiz beslenmesinde çok önemli bir yer tutan ve yurt dışına giderek artan miktarlarda ihraç edilen ballar, aynı zamanda ekonomimiz açısından da önemlidir. Bu nedenle besinlerin tüm besin zinciri boyunca geçirdiği aşamalarda, naftalinle ve diğer kalıntılarla kontaminasyonun önlenmesi, hijyen bakımından kontrol ve analizlerinin yapılması gerekmektedir. Beslenmeyle ilgili yapılacak kalıntı çalışmaları sağlıklı bir toplum kazanmak ve sağlıklı nesillerin yetişmesi bakımından büyük önem taşır.SUMMARYTurkey having a wealthy flora with different climate conditions, beekeeping is widely spread all over the country. Due to wrong and/or unconscious pesticide treatments against pests generally causes residues in honey and beeswax. Treatment of naphthalene to beeswax is preferred for removing the moths from the beehive and as a result of it, honey is contaminated with naphthalene. Naphthalene, a volatile aromatic hydrocarbon, is widely used in mothballs and toilet bowl deodorants, in the production of dyes and lubricants. Naphthalene absorption by oral, inhalation and dermal contact causes systemic toxic effects for human. The lungs and eyes appear to be most susceptible to toxicity, although toxic effects can be demonstrated in other tissues, such as the liver, brain and kidney. In our study, naphthalene was detected in honey specimens which are provided from various regions of Turkey using high pressure liquid chromatography (HPLC) with diode array detector (DAD). In this study, naphthalene analysis was carried out in 100 honey specimens. Out of 100 specimens 53 of them were purchased from markets and 47 of them were from street bazaars existing in various regions of Turkey. Naphthalene-suspicious one honey specimen was analysed and confirmed by gas chromatography-mass spectrometry (GC-MS). Honey has an important role in our food chain and in our country’s economy. Therefore, fooodstuffs are needed to be hygienic controlled and analyzed during food process and prevent the contamination of naphthalene and other residues. All residue analyses for the entire food chain have a lot of importance for healthy people and healthy generation

Metabolic Rewiring and the Characterization of Oncometabolites

The study of low-molecular-weight metabolites that exist in cells and organisms is known as metabolomics and is often conducted using mass spectrometry laboratory platforms. Definition of oncometabolites in the context of the metabolic phenotype of cancer cells has been accomplished through metabolomics. Oncometabolites result from mutations in cancer cell genes or from hypoxia-driven enzyme promiscuity. As a result, normal metabolites accumulate in cancer cells to unusually high concentrations or, alternatively, unusual metabolites are produced. The typical oncometabolites fumarate, succinate, (2R)-hydroxyglutarate and (2S)-hydroxyglutarate inhibit 2-oxoglutarate-dependent dioxygenases, such as histone demethylases and HIF prolyl-4-hydroxylases, together with DNA cytosine demethylases. As a result of the cancer cell acquiring this new metabolic phenotype, major changes in gene transcription occur and the modification of the epigenetic landscape of the cell promotes proliferation and progression of cancers. Stabilization of HIF1α through inhibition of HIF prolyl-4-hydroxylases by oncometabolites such as fumarate and succinate leads to a pseudohypoxic state that promotes inflammation, angiogenesis and metastasis. Metabolomics has additionally been employed to define the metabolic phenotype of cancer cells and patient biofluids in the search for cancer biomarkers. These efforts have led to the uncovering of the putative oncometabolites sarcosine, glycine, lactate, kynurenine, methylglyoxal, hypotaurine and (2R,3S)-dihydroxybutanoate, for which further research is required

Metabolomic and Lipidomic Biomarkers for Premalignant Liver Disease Diagnosis and Therapy

In recent years, there has been a plethora of attempts to discover biomarkers that are more reliable than α-fetoprotein for the early prediction and prognosis of hepatocellular carcinoma (HCC). Efforts have involved such fields as genomics, transcriptomics, epigenetics, microRNA, exosomes, proteomics, glycoproteomics, and metabolomics. HCC arises against a background of inflammation, steatosis, and cirrhosis, due mainly to hepatic insults caused by alcohol abuse, hepatitis B and C virus infection, adiposity, and diabetes. Metabolomics offers an opportunity, without recourse to liver biopsy, to discover biomarkers for premalignant liver disease, thereby alerting the potential of impending HCC. We have reviewed metabolomic studies in alcoholic liver disease (ALD), cholestasis, fibrosis, cirrhosis, nonalcoholic fatty liver (NAFL), and nonalcoholic steatohepatitis (NASH). Specificity was our major criterion in proposing clinical evaluation of indole-3-lactic acid, phenyllactic acid, N-lauroylglycine, decatrienoate, N-acetyltaurine for ALD, urinary sulfated bile acids for cholestasis, cervonoyl ethanolamide for fibrosis, 16α-hydroxyestrone for cirrhosis, and the pattern of acyl carnitines for NAFL and NASH. These examples derive from a large body of published metabolomic observations in various liver diseases in adults, adolescents, and children, together with animal models. Many other options have been tabulated. Metabolomic biomarkers for premalignant liver disease may help reduce the incidence of HCC

The glycine deportation system and its pharmacological consequences

The glycine deportation system is an essential component of glycine catabolism in man whereby 400 to 800mg glycine per day are deported into urine as hippuric acid. The molecular escort for this deportation is benzoic acid, which derives from the diet and from gut microbiota metabolism of dietary precursors. Three components of this system, involving hepatic and renal metabolism, and renal active tubular secretion help regulate systemic and central nervous system levels of glycine. When glycine levels are pathologically high, as in congenital nonketotic hyperglycinemia, the glycine deportation system can be upregulated with pharmacological doses of benzoic acid to assist in normalization of glycine homeostasis. In congenital urea cycle enzymopathies, similar activation of the glycine deportation system with benzoic acid is useful for the excretion of excess nitrogen in the form of glycine. Drugs which can substitute for benzoic acid as substrates for the glycine deportation system have adverse reactions that may involve perturbations of glycine homeostasis. The cancer chemotherapeutic agent ifosfamide has an unacceptably high incidence of encephalopathy. This would appear to arise as a result of the production of toxic aldehyde metabolites which deplete ATP production and sequester NADH in the mitochondrial matrix, thereby inhibiting the glycine deportation system and causing de novo glycine synthesis by the glycine cleavage system. We hypothesize that this would result in hyperglycinemia and encephalopathy. This understanding may lead to novel prophylactic strategies for ifosfamide encephalopathy. Thus, the glycine deportation system plays multiple key roles in physiological and neurotoxicological processes involving glycine

(2R,3S)-Dihydroxybutanoic Acid Synthesis as a Novel Metabolic Function of Mutant Isocitrate Dehydrogenase 1 and 2 in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) frequently harbors mutations in isocitrate 1 (IDH1) and 2 (IDH2) genes, leading to the formation of the oncometabolite (2R)-hydroxyglutaric acid (2R-HG) with epigenetic consequences for AML proliferation and differentiation. To investigate if broad metabolic aberrations may result from IDH1 and IDH2 mutations in AML, plasma metabolomics was conducted by gas chromatography-mass spectrometry (GC-MS) on 51 AML patients, 29 IDH1/2 wild-type (WT), 9 with IDH1R132, 12 with IDH2R140 and one with IDH2R172 mutations. Distinct metabolic differences were observed between IDH1/2 WT, IDH1R132 and IDH2R140 patients that comprised 22 plasma metabolites that were mainly amino acids. Only two plasma metabolites were statistically significantly different (p < 0.0001) between both IDH1R132 and WT IDH1/2 and IDH2R140 and WT IDH1/2, specifically (2R)-hydroxyglutaric acid (2R-HG) and the threonine metabolite (2R,3S)-dihydroxybutanoic acid (2,3-DHBA). Moreover, 2R-HG correlated strongly (p < 0.0001) with 2,3-DHBA in plasma. One WT patient was discovered to have a D-2-hydroxyglutarate dehydrogenase (D2HGDH) A426T inactivating mutation but this had little influence on 2R-HG and 2,3-DHBA plasma concentrations. Expression of transporter genes SLC16A1 and SLC16A3 displayed a weak correlation with 2R-HG but not 2,3-DHBA plasma concentrations. Receiver operating characteristic (ROC) analysis demonstrated that 2,3-DHBA was a better biomarker for IDH mutation than 2R-HG (Area under the curve (AUC) 0.861; p < 0.0001; 80% specificity; 87.3% sensitivity). It was concluded that 2,3-DHBA and 2R-HG are both formed by mutant IDH1R132, IDH2R140 and IDH2R172, suggesting a potential role of 2,3-DHBA in AML pathogenesis