Increase of Cell-Free Nuclear and Mitochondrial DNA Content in the Urine of Rats after X-ray Irradiation or Bleomycin Administration

2019 ◽  
Vol 64 (5) ◽  
pp. 5-8
Author(s):  
Г. Минкабирова ◽  
G. Minkabirova ◽  
С. Абдуллаев ◽  
S. Abdullaev
Keyword(s):  
Mitochondrial Dna ◽  
Linear Dependence ◽  
Dna Content ◽  
Nuclear Dna ◽  
The Body ◽  
Male Rats ◽  
Cytostatic Drug ◽  
Rat Urine ◽  
X Ray ◽  
Potential Biomarker

Purpose: To study the content of cell-free mitochondrial DNA (cf-mtDNA) and cell-free nuclear DNA (cf-nDNA) in urine of rats exposed to ionizing radiation, and after injection of a cytostatic drug bleomycin. Material and methods: Wistar male rats aged 3 months were used in the experiments. Rats were irradiated at a doses of 3, 5, and 8 Gy. Bleomycin was administered intraperitoneally in concentrations of 3, 7, and 10 mg/kg. The DNA content was measured by real-time PCR. Results: The results showed an increase in the level of the number of cf-nDNA and cf-mtDNA fragments in urine of irradiated rats. It was shown that the content of cf-nDNA and cf-mtDNA has a linear dependence on the X-ray dose. Thus, the maximum number of mtDNA and nDNA copies was recorded for 12–24th hours after irradiation. The number of PCR amplification products of cf-mtDNA is 2–3 times higher than those of cf-nDNA. Data analysis of the content of cf-nDNA and cf-mtDNA in rat urine after introduction of bleomycin also showed elevated levels compared with control animals. It was shown that the content of cf-nDNA and cf-mtDNA has a linear dependence on the dose of the chemotherapeutic drug. Conclusion: Thus, it has been shown that it is possible to overcome the transrenal (renal) barrier in animals with cf-mtDNA and cf-nDNA and pass them into the urine after X-ray irradiation, as well as after the administration of bleomycin. The dose dependence of the identified effects was found. The increased content of cell-free DNA in the urine can be considered as a potential biomarker for assessing the level of genotoxic load during radiation damage to the body, as well as when exposed to other genotoxic agents.

scholarly journals Endogenous Histamine Excretion in the Rat as Influenced by X-Ray Irradiation and Compound 48/80

1956 ◽  
Vol 187 (2) ◽  
pp. 307-311 ◽  
Author(s):  
James L. Leitch ◽  
Virginia G. Debley ◽  
Thomas J. Haley
Keyword(s):  
Chronic Administration ◽  
Female Rats ◽  
Male Rats ◽  
Whole Body ◽  
Rat Urine ◽  
Similar Treatment ◽  
X Ray ◽  
Endogenous Histamine ◽  
Liberation Of Histamine ◽  
Urinary Histamine

Studies have been made on a spasmogen from rat urine. This material has been identified as histamine. It has been shown that both acute whole body x-ray irradiation and compound 48/80 significantly increase the quantity of endogenous urinary histamine. Endogenous histamine excretion in female rats is approximately 9–10 times greater than in male rats. Neither x-ray irradiation nor compound 48/80 elevate the amount of urinary histamine of the males to that of the female. Histamine liberation is of little or no importance insofar as lethality from acute whole body irradiation is concerned. The amount of histamine liberated by such irradiation is independent of radiation dosage within the range 600–1200 r. After radiation injury, significant levels of urinary histamine were detected only during the first 24 hours. Histamine depletion by chronic administration of compound 48/80 did not prevent further liberation of histamine by acute whole body irradiation. Irradiated animals are much more susceptible to the toxic effects of compound 48/80 than normal animals. Gonadectomy followed by α-estradiol injection did not increase the output of urinary histamine in male rats. Similar treatment of female rats with testosterone did not reduce their urinary histamine output, but a reduction was observed 140 days after surgery. Administration of cortisone acetate to male rats did not increase their excretion of endogenous histamine. Inactivation of diamine oxidase with aminoguanidine had little or no effect on urinary histamine output in male rats, but caused a threefold increase in females. Further elevation in urinary histamine was produced by irradiation or compound 48/80.

scholarly journals Evidence of hybridization, mitochondrial introgression and biparental inheritance of the kDNA minicircles in Trypanosoma cruzi I

2019 ◽  
Author(s):  
Fanny Rusman ◽  
Noelia Floridia-Yapur ◽  
Paula G. Ragone ◽  
Patricio Diosque ◽  
Nicolás Tomasini
Keyword(s):  
Mitochondrial Dna ◽  
Trypanosoma Cruzi ◽  
Dna Sequences ◽  
Dna Content ◽  
Nuclear Dna ◽  
Genetic Exchange ◽  
Natural Hybridization ◽  
Biparental Inheritance ◽  
Hybrid Strain ◽  
Mitochondrial Introgression

AbstractBackgroundGenetic Exchange in Trypanosoma cruzi is controversial not only in relation to its frequency but also in relation to its mechanism. A mechanism of parasexuality has been proposed based on laboratory hybrids, but population genomics strongly suggests meiosis. In addition, mitochondrial introgression has been reported several times in natural isolates although its mechanism is not clear. Moreover, hybrid DTUs (TcV and TcVI) have inherited at least part of the kinetoplastic DNA (kDNA = mitochondrial DNA) from both parents.Methodology/Principal findingsIn order to address such topics, we sequenced and analyzed fourteen nuclear DNA fragments and three kDNA maxicircle genes in three TcI stocks which are natural clones potentially involved in events of genetic exchange. We also deep-sequenced (a total of 6,146,686 paired-end reads) the hypervariable region of kDNA minicircles (mHVR) in such three strains. In addition, we analyzed the DNA content by flow cytometry to address cell ploidy. We observed that most polymorphic sites in nuclear loci showed a hybrid pattern in one cloned strain and the other two cloned strains were compatible as parental strains (or nearly related to the true parents). The three clones have almost the same ploidy and the DNA content was similar to the reference strain Sylvio (an almost diploid strain). Despite maxicircle genes evolve faster than nuclear housekeeping ones, we did not detect polymorphism in the sequence of three maxicircle genes showing mito-nuclear discordance. In addition, the hybrid stock shared 66% of its mHVR clusters with one putative parental and 47% with the another one. In contrast, the putative parental stocks shared less than 30% of the mHVR clusters among them.Conclusions/significanceThe results suggest a reductive division, a natural hybridization, biparental inheritance of the minicircles in the hybrid and maxicircle introgression. The models including such phenomena and that would explain the relationships between these three clones are discussed.Author summaryChagas disease, an important public health problem in Latin America, is caused by the parasite Trypanosoma cruzi. Despite it is a widely studied parasite, several questions about the biology of genetic exchange remain. Meiosis has not been yet observed in laboratory, although inferred from population genomic studies. In addition, previous results suggest that the mitochondrial DNA (called kDNA) may be inherited from both parents in hybrids. Here, we analyzed a hybrid strain and the potential parents to address about the mechanisms of genetic exchange at nuclear and mitochondrial level. We observed that the hybrid strain has heterozygous patterns and DNA content compatible with an event of meiosis. In addition, we observed that the evolutionary histories of nuclear DNA and maxicircles (a part of the kDNA) were discordant and the three strains share identical DNA sequences. Mitochondrial introgression of maxicircle DNA from one genotype to another may explain this observation. In addition, we detected that the hybrid strain shared minicircles (another part of the kDNA) with both parental strains. Our results suggest that hybridization implied meiosis and biparental inheritance of the kDNA. Further research is required to address such phenomena in detail.

scholarly journals Mitochondrial growth and DNA synthesis occur in the absence of nuclear DNA replication in fission yeast

1990 ◽  
Vol 97 (3) ◽  
pp. 509-516 ◽  
Author(s):  
S. Sazer ◽  
S.W. Sherwood
Keyword(s):  
Mitochondrial Dna ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Fission Yeast ◽  
Dna Content ◽  
Nuclear Dna ◽  
Equal Distribution ◽  
Daughter Cells ◽  
Mitochondrial Dna Content ◽  
Mitochondrial Volume

Cell growth and division require the doubling of cellular constituents followed by their equal distribution to the two daughter cells. Within a growing population, the ratio of mitochondrial to cellular volume is maintained, as is the number of mitochondrial genomes per cell. The mechanisms responsible for coordinating nuclear and mitochondrial DNA synthesis, and for balancing increases in cell and mitochondrial size are not well understood. In studies of the fission yeast Schizosaccharomyces pombe we quantified cellular and mitochondrial DNA content by both Southern blot analysis and flow cytometry of cells stained with a variety of DNA-binding fluorochromes, which we show are able to detect nuclear and mitochondrial DNA with different efficiencies. In the conditional cell division cycle mutant cdc10, which is unable to initiate nuclear DNA synthesis, we found that there was an increase in the mitochondrial DNA content in the absence of nuclear DNA replication. This demonstrates that mitochondrial and nuclear DNA synthesis are not obligately linked. We also show that mitochondrial DNA replication is not required for the increase in mitochondrial size that occurs as cells elongate, although this results in a decrease in the ratio of mitochondrial DNA to mitochondrial volume.

scholarly journals Changes in circulating cell-free nuclear DNA and mitochondrial DNA of patients with adolescent idiopathic scoliosis

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Jiong Li ◽  
Longjie Wang ◽  
Guanteng Yang ◽  
Yunjia Wang ◽  
Chaofeng Guo ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Adolescent Idiopathic Scoliosis ◽  
Idiopathic Scoliosis ◽  
Sensitivity And Specificity ◽  
Dna Analysis ◽  
Nuclear Dna ◽  
Predictive Accuracy ◽  
Lower Sensitivity ◽  
Mt Dna ◽  
Potential Biomarker

Abstract Background Adolescent idiopathic scoliosis (AIS) which characterized by complex three-dimensional deformity of spine has been difficult to cure because of the unknown etiopathology and uncertainty of progression. Nowadays, circulating cell-free (ccf) DNA was found to be a potential biomarker for several benign and malignant diseases. However, whether ccf DNA can be a biomarker for AIS has not been reported yet. In this study, we investigate the circulating cell-free nuclear DNA (ccf n-DNA) and mitochondrial DNA (ccf mt-DNA) concentrations in the plasma of patients with AIS and controls (CT), and the changed plasma ccf n-DNA and ccf mt-DNA levels and their association with clinical parameters were assessed. Methods The plasma of peripheral blood from 69 AIS patients and 21 age-matched CT was collected for ccf DNA analysis. Quantitative PCR was used to detect ccf n-DNA and ccf mt-DNA levels, and correlation analyses between the ccf n-DNA and ccf mt-DNA levels and clinical characteristics were conducted. Receiver operator curves (ROC) were used to analyze the sensitivity and specificity of ccf n-DNA and ccf mt-DNA levels to different characteristics. Results The plasma ccf n-DNA levels of both GAPDH and ACTB were significantly decreased in AIS patients compared with those in controls, while the plasma ccf mt-DNA levels did not changed. According to sex-related analyses, the ccf n-DNA levels in male CT-M was higher than that in female CT and male AIS, but the ccf n-DNA levels in female AIS was not significantly changed when compared with male AIS or female CT. However, the concentration of ccf mt-DNA in female AIS increased significantly when compared with male AIS. Surprisingly, Lenke type-related analyses suggested that Lenke type 1 patients had lower ccf n-DNA levels, whereas Lenke type 5 patients had higher ccf mt-DNA levels compared with those of controls. However, a lower sensitivity and specificity of AIS predicted by ccf n-DNA or ccf mt-DNA levels was observed, whether in total, by sex, or by Lenke type. Conclusion Although with no/little predictive accuracy of AIS/progressed AIS by ccf DNA levels, significantly changed plasma ccf DNA levels were observed in AIS patients compared with those in controls.

scholarly journals Chasing a moving target: Detection of mitochondrial heteroplasmy for clinical diagnostics

2017 ◽  
Author(s):  
Eric J. White ◽  
Tristen Ross ◽  
Edgardo Lopez ◽  
Anastasia Nikiforov ◽  
Christopher Gault ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Nuclear Dna ◽  
De Novo ◽  
Clinical Diagnostics ◽  
The Body ◽  
Sequencing Data ◽  
Simple Method ◽  
Large Deletions ◽  
Mucosal Cells ◽  
Mtdna Variants

AbstractClinical interpretation of human mitochondrial DNA (mtDNA) variants has been challenging for technical and biological reasons but the involvement of dysfunctional mitochondria in many diseases makes it imperative to have a validated assay for detecting pathogenic variants. We have tested several methods to identify those best suited to detect and confirm mtDNA variants. The choice of methods is dependent on the amount of DNA available for testing and the sensitivity required for detecting low-level heteroplasmies. There is a tradeoff between a polymerase’s ability to amplify small amounts of DNA and its ability to generate accurate sequence. We report a simple method to measure heteroplasmy levels of large deletions from NGS data alone without need for qPCR or other methods. Use of HapMap samples for standardization needs to be done with caution as most have novel heteroplasmic sites that have arisen during immortalization/cell culture processes. Different batches of DNA can have variable sequence. In contrast, we observed no de novo heteroplasmies in healthy mother-child pairs studied using blood or saliva though the frequency of pre-existing heteroplasmies often changed dramatically across generations. Long-read nanopore sequencing of individuals with two heteroplasmies suggested a random distribution of variants on single molecules but technical artifacts prevent certainty on this finding. Urine provides an additional readily accessible source of mtDNA that can be used for bone marrow transplant recipients whose saliva/blood mtDNA may be contaminated by the BMT donor’s mtDNA. We have characterized cells suspended in urine via expression profiling and shown them to be primarily mucosal cells that are independent of blood. Understanding the pitfalls of the various mtDNA sequencing methods allows development of reliable and accurate tests suitable for clinical diagnostics.Author SummaryMitochondrial DNA is important for many diseases but it is present at many copies per cell so is harder to check for mutations compared to nuclear DNA. We have studied mitochondrial DNA in different ways to see how it changes across generations and in different locations in the body. The tests need to be much more sensitive than nuclear DNA tests so that we can detect mutations down to 1%. We have shown that mitochondrial DNA changes when cell lines are used but saliva, blood and cells in the urine can all be used for testing. Cells in the urine originate as mucosal cells and are independent of blood. We developed a new method for analyzing large deletions that means sequencing data alone can be used for measuring the frequency of deletions. We also followed a family with two variable sites to better understand how mitochondrial DNA changes from mother to child. In some children, the variants stayed the same while, in others, variants disappeared.

X-ray micro tomography in the scanning electron microscope

1978 ◽  
Vol 36 (1) ◽  
pp. 106-107 ◽  
Author(s):  
W. Brünger
Keyword(s):  
Electron Beam ◽  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Target Surface ◽  
The Body ◽  
X Rays ◽  
Cross Sectional ◽  
X Ray ◽  
Micro Tomography ◽  
Scanning Electron

Reconstructive tomography is a new technique in diagnostic radiology for imaging cross-sectional planes of the human body /1/. A collimated beam of X-rays is scanned through a thin slice of the body and the transmitted intensity is recorded by a detector giving a linear shadow graph or projection (see fig. 1). Many of these projections at different angles are used to reconstruct the body-layer, usually with the aid of a computer. The picture element size of present tomographic scanners is approximately 1.1 mm2.Micro tomography can be realized using the very fine X-ray source generated by the focused electron beam of a scanning electron microscope (see fig. 2). The translation of the X-ray source is done by a line scan of the electron beam on a polished target surface /2/. Projections at different angles are produced by rotating the object.During the registration of a single scan the electron beam is deflected in one direction only, while both deflections are operating in the display tube.

Great Increase of Numbers of Virus Particles by Irradiating C3H Mouse Mammary Tumors Transplanted into Male Mice

1972 ◽  
Vol 30 ◽  
pp. 272-273
Author(s):  
P. J. Melnick ◽  
J. W. Cha ◽  
E. Samouhos
Keyword(s):  
Subcutaneous Tissue ◽  
Mammary Tumors ◽  
The Body ◽  
X Ray ◽  
Virus Particles ◽  
Mouse Mammary Tumors ◽  
Transplantable Tumors ◽  
C3h Mice ◽  
Cobalt Radiation ◽  
Enzyme Changes

Spontaneous mammary tumors in females of a high tumor strain of C3H mice were cut into small fragments that were Implanted into the subcutaneous tissue of the back of males of the same strain, where they grew as transplantable tumors. When about Cm. In diameter daily fractional radiation was begun, applied to the tumors, the rest of the body being shielded by a lead shield. Two groups were treated with 150 and 200 r X-ray dally, of half value layer 0.6mm. copper; a third group was treated with 500 r cobalt radiation dally. The primary purpose was to examine the enzyme changes during radiation, with histochemlcal technics.

Pathophysiological mechanisms of hepatic immune reactivity in prolonged experimental exposure of the body to sodium fluoride

2019 ◽  
pp. 64-72
Author(s):  
А.С. Казицкая ◽  
Т.К. Ядыкина ◽  
М.С. Бугаева ◽  
А.Г. Жукова ◽  
Н.Н. Михайлова ◽  
...  
Keyword(s):  
Sodium Fluoride ◽  
Histological Study ◽  
The Body ◽  
Male Rats ◽  
Free Access ◽  
Immune Reactivity ◽  
Organ Protection ◽  
Pathophysiological Mechanisms ◽  
Subchronic Exposure ◽  
Study Results

В условиях непрерывного воздействия неблагоприятных факторов окружающей и производственной среды на человека особую актуальность приобретает изучение механизмов, поддерживающих гомеостаз организма. Длительное поступление фторидов в организм приводит к формированию хронической фтористой интоксикации, патогенез которой вызывает многочисленные споры и дискуссии. До сих пор недостаточно внимания уделяется изучению висцеральной патологии, обусловленной нарушениями иммунного статуса в условиях воздействия на организм соединений фтора. Практически отсутствуют исследования по изучению иммунной реактивности, определяющей морфофункциональный характер ответной реакции печени на ранних стадиях развития фтористой интоксикации. Цель работы - изучение действий патофизиологических механизмов иммунной реактивности печени при субхроническом действии на организм соединений фтора. Методика. Опыты проведены на 210 лабораторных крысах-самцах массой 180-220 г., разделенных на 2 группы: контрольную (n=80) и группу животных с субхроническим действием фторида натрия (n=130). Экспериментальные животные в течение 12 нед имели свободный доступ к водному раствору фторида натрия (концентрация 10 мг/л, что составляет суточную дозу фтора 1,2 мг/кг массы тела). Для изучения иммунологических и биохимических показателей забирали кровь из хвостовой вены через 1, 3, 6, 9, 12 нед от начала эксперимента. Для оценки состояния гуморального звена иммунитета определяли уровень сывороточных иммуноглобулинов (IgA, IgG, IgM) иммуноферментным анализом с помощью наборов реактивов ЗАО «Вектор-Бест» (Новосибирск). Уровень сывороточных цитокинов: TNF-α, IL-1β, 2, 4, 6, 10 определяли на анализаторе Multiskan EX методом иммуноферментного анализа с использованием наборов «Вектор Бест» (Новосибирск). Подсчет общего количества лейкоцитов произведен классическим способом в камере Горяева, анализ лейкоцитарной формулы - в окрашенных мазках периферической крови. Метаболические изменения оценивали по активности ферментов в ткани печени: щелочной фосфатазы (ЩФ), аланин- и аспартатаминотрансфераз (АЛТ, АСТ), лактатдегидрогеназы (ЛДГ), гаммаглутамилтранспептидазы (γ-ГТ). Активность ферментов определяли унифицированными методами с помощью наборов реактивов ЗАО «Вектор-Бест» (Новосибирск) на фотометре PM-750 (Германия). Гистологические исследования печени осуществляли после декапитации крыс, проводимой под эфирным наркозом. Результаты. Показано, что субхроническое воздействие фторида натрия сопровождается формированием внутриклеточных и внутрисосудистых повреждений печени. Активация медиаторов воспаления и развитие иммунологических нарушений в динамике эксперимента способствуют формированию системной воспалительной реакции, которая приводит к появлению стойких морфологических нарушений в печени и изменению активности ферментов основных метаболических путей. Заключение. Полученные результаты могут быть использованы при разработке и проведении профилактических мероприятий в условиях воздействия на организм высоких концентраций фтора с последовательным применением детоксикационной, иммуномодуляторной и органопротекторной коррекции. Studying mechanisms, which maintain the body homeostasis, is particularly important in the conditions of continuous impact of adverse environmental and manufacturing factors. Long-term exposure to fluorides leads to chronic fluoric intoxication, the pathogenesis of which is a subject of multiple controversy and discussions. Not enough attention is still paid to elucidating the visceral pathology associated with fluorine-induced immune disorders. There are virtually no studies of immune reactions that define the morphofunctional nature of the liver response to early stages of fluoric intoxication. Aim. To study pathophysiological mechanisms of hepatic immune reactivity in subchronic exposure of the body to fluorine compounds. Methods. Experiments were performed on 210 male rats weighing 180-220 g. The animals were divided into two groups: 1) control (n=80) and 2) subchronic exposure to sodium fluoride (n=130). The rats had free access to a 10 mg/l aqueous solution of sodium fluoride (daily dose, 1.2 mg/kg body weight) for 12 weeks. Blood was withdrawn from the caudal vein at 1, 3, 6, 9, and 12 weeks of the experiment for immunological and biochemical tests. Histological study of the liver was performed after decapitation of rats under ether anesthesia. Results. The subchronic exposure to sodium fluoride was associated with intracellular and intravascular damage of the liver. Activation of inflammatory mediators and development of immunological disorders during the experiment contributed to a systemic inflammatory reaction, which resulted in persistent morphological injuries of the liver and changes in enzyme activities in major metabolic pathways. Conclusion. The study results can be used for development and implementation of preventive measures against the effects of high fluorine concentrations, which would include a successive use of detoxification, immunomodulation and organ protection.

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