Trinucleated Cells Research Articles

Sampling, composition, and biological effects of Mexico City airborne particulate matter from multiple periods

Air pollution is a worldwide environmental problem with an impact on human health. Particulate matter of ten micrometers or less aerodynamic diameter (PM10) as well as its fine fraction (PM2.5) is related to multiple pulmonary diseases. The impact of air pollution in Mexico City, and importantly, particulate matter has been studied and considered as a risk factor for two decades ago. Previous studies have reported the composition of Mexico City particulate matter, as well as the biological effects induced by this material. However, material collected and used in previous studies is a limited resource, and sampling and particle recovery techniques have been improved. In this study, we describe the methods used in our laboratory for Mexico City airborne particulate matter PM10 and PM2.5 sampling, considering the years 2017, 2018 and 2019. We also analyzed the PM10 and PM2.5 samples obtained to determine their composition. Finally, we exposed lung cell line cultures to PM10 and PM2.5 to evaluate the biological effect of the material in terms of cell viability, cell death, inflammatory response, and cytogenetic alterations. Our results showed that PM10 composition includes inorganic, organic and biological compounds, while PM2.5 is a mixture of more enriched organic compounds. PM10 and PM2.5 treatment in lung cells does not significantly impact cell viability/cell death. However, PM10 and PM2.5 increase the secretion levels of IL-6. Moreover, PM10 as well as PM2.5 induce cytogenetic alterations, such as micronuclei, anaphase bridges, trinucleated cells and apoptotic cells in lung cells. Our results update the evidence of the composition and biological effects of Mexico City particulate matter and provide us a reliable basis for future approaches.

Airborne particulate matter induces mitotic slippage and chromosomal missegregation through disruption of the spindle assembly checkpoint (SAC)

Particulate matter (PM) is a risk factor for lung cancer development and chromosomal missegregation and cell cycle disruptions are key cellular events that trigger tumorigenesis. We aimed to study the effect of PM10 (PM with an aerodynamic diameter ≤10 μm) on mitotic arrest and chromosomal segregation, evaluating the spindle assembly checkpoint (SAC) protein dynamics in the human lung A549 adenocarcinoma cell line. For this purpose, synchronized cells were exposed to PM10 for 24 h to obtain the frequency of micronucleated (MN) and trinucleated (TN) cells. Then, the efficiency of the mitotic arrest after PM10 exposure was analyzed. To elucidate the effect of PM10 in chromosomal segregation, the levels and subcellular localization of essential SAC proteins were evaluated. Results indicated that A549 cells exposed to PM10 exhibited an increase in MN and TN cells and a decrease in mitotic indexes and G2/M phase. A549 cells treated with PM10 showed reduced protein levels of MDC1 and NEK2 (38% and 35% respectively), which is required for MAD2 recruitment to kinetochores, MAD2 and BUBR1, effectors of the SAC (25% and 18% respectively), and CYCLIN B1, required during G2/M phase (35%). Besides, PM10 exposure increase the levels of AURORA B and SURVIVIN, required for SAC activation through chromosome-microtubule attachment errors (85% and 74% respectively). We suggest that PM10 causes mitotic slippage due to alterations in MAD2 localization. Thus, PM10 causes inadequate chromosomal segregation and deficient mitotic arrest by altering SAC protein levels, predisposing A549 cells to chromosomal instability, a common feature observed in cancer.

Lymphocyte Proliferation Kinetics in Inhabitant of Takandeang Village, Mamuju: A High Background Radiation Areas in Indonesia

BACKGROUND: Mamuju area in West Sulawesi considered as the high natural background radiation area in Indonesia. Our previous study showed that the mean mitotic index (MI) and nuclear division index (NDI) in lymphocytes of Botteng Village, Mamuju inhabitants was lower compared to control samples. To validate our previous study results, here in this study the evaluation of cell proliferation markers which were MI and NDI in peripheral blood lymphocytes (PBL) of Takandeang Village inhabitants was conducted.METHODS: A total 60 people were enrolled in this study, consisted of 35 samples from Takandeang Sub-Village and 25 from normal background radiation area. MI was calculated manually and automatically using Metafer 3.11.2 imaging system. The NDI defined as proportion of mononucleated, binucleated, trinucleated and tetranucleated cells were conducted using cytokinesis block micronucleus (CBMN) assay.RESULTS: The results of this study showed that the mean manual MI in Takandeang Sub-Village inhabitants was lower compared to control group (4.96±2.25 vs. 5.93±2.14). In contrast, the mean automatic MI (20.37±10.49 vs. 18.87±7.49) and NDI (1.555±0.174 vs. 1.523±0.112) in Takandeang Sub-Village inhabitants was higher compared to the control group. Statistical analysis revealed that the difference of mean manual MI, automatic MI and NDI in Takandeang Sub-Village inhabitants was not significantly different compared to the control group(p>0.05).CONCLUSION: It can be concluded that based on this study the chronic low radiation dose exposure in Takandeang Sub-Village, Mamuju has no significant effect on the lymphocytes proliferation.KEYWORDS: lymphocytes, mitotic index, nuclear division index, high background radiation

Increased genome instability and oxidative DNA damage and their association with IGF-1 levels in patients with active acromegaly

ObjectiveThe objectives of this study were to assess cytokinesis-block micronucleus cytome (CBMN Cyt) assay parameters and also oxidative DNA damage in patients with active acromegaly and controls and to assess the relationship between age, serum insulin-like growth factor 1 (IGF-1) levels, pituitary adenoma diameters, 8-hydroxy-2′-deoxyguanosine (8-OHdG) levels and CBMN Cyt assay parameters in patients with active acromegaly. DesignThe study population included 30 patients with active acromegaly and 30 age- and sex-matched healthy controls. CBMN Cyt assay parameters in peripheral blood lymphocytes of patients with active acromegaly and controls were evaluated and plasma 8-OHdG levels were measured. ResultsFrequencies of micronucleus (MN), nucleoplasmic bridges (NPBs) and nuclear buds (NBUDs) in lymphocytes of patients with acromegaly were found to be significantly higher than those in controls (p<0.001, p<0.001, p<0.001, respectively). The frequencies of apoptotic and necrotic cells in lymphocytes of patients with acromegaly were found to be significantly higher than those in controls (p<0.001 and p<0.001 respectively). No statistically significant differences in the number of cells in metaphase, the number of bi-nucleated cells (M2), the number of tri-nucleated cells (M3), the number of tetra-nucleated cells (M4) and nuclear division index (NDI) values were observed between patients and controls (p>0.05). Plasma 8-OHdG (ng/ml) levels in patients with acromegaly were found to be significantly higher than those in controls (p<0.005). MN frequency in the lymphocytes of patients with acromegaly increased with elevated serum IGF-1 levels (p<0.05), whereas the number of NPBs and the frequency of apoptotic cells decreased with elevated serum IGF-1 levels (p<0.01 and p<0.05 respectively). ConclusionsBoth the increase in chromosomal/oxidative DNA damage and the positive association between MN frequency and serum IGF-1 levels may predict an increased risk of malignancy in acromegalic patients. Long-term follow-up of patients with acromegaly will be necessary to establish the degree of cancer risk in this population.

Gross morphology and histology of head and salivary apparatus of the predatory bug, Rhynocoris marginatus.

Rhynocoris marginatus Fabricius (Hemiptera: Reduviidae) is an important biological control agent against more than 25 insect pests in India. For a better understanding of the feeding adaptation of this bug, the gross morphology and histology of its head and salivary apparatus were studied using both a light microscope and scanning electron microscope. The head is more or less elongate, mobile, and immersed into the eyes. R. marginatus has a three-segmented curved rostrum; the middle segment is longer than the other two segments. The terminal rostral segment bears spines and trichobothria externally. Stylet bundles bear two pairs of maxillary and mandible stylets in the curved rostrum with serrations. The stylets help to penetrate into the tissue and directly pump the toxic venomous saliva deep into the prey. The principal gland is bi-lobed (anterior lobe and posterior lobe), whereas the accessory gland is uni-lobed, exhibiting distinct functional and histological differences. These glands receive tracheal and nerve supply. Mononucleated, binucleated, trinucleated and polynucleated cells are distributed both in anterior and posterior lobes of the principal gland. The cytoplasm has collecting vacuoles with secretions. Therefore, this predator is highly equipped with well-developed mouthparts that are attached to the salivary apparatus.

Determination of stress responses induced by aluminum in maize(Zea mays)

To assess the alternative responses to aluminum toxicity, maize (Zea mays L. cv Karadeniz yıldızı) roots were exposed to different concentrations of AlCl3 (150, 300 and 450 μM). Aluminum reduced the root elongation by 39.6% in 150 μM, 44.1% in 300 μM, 50.1% in 450 μM AlCl3 after 96 h period. To correlate the root elongation with the alternative stress responses including aluminum accumulation, lipid peroxidation, mitotic abnormalities, reduction of starch content, intracellular Ca2+ accumulation, callose formation, lignin deposition and peroxidase activity, cytochemical and biochemical tests were performed. The results indicated that aluminum accumulation and lipid peroxidation were observed more densely on the root cap and the outer cortex cells. In addition to morphological deformations, cytochemical analysis displayed cellular deformations. Furthermore, mitotic abnormalities were observed such as c-mitosis, micronuclei, bi- and trinucleated cells in aluminum treated root tips. Aluminum treatment induced starch reduction, callose formation, lignin accumulation and intracellular Ca2+ increase. Moreover, the peroxidase activity increased significantly by 3, 4.4 and 7.7 times higher than in that of control after 96 h, respectively. In conclusion, aluminum is significantly stressful in maize culminating in morphological and cellular alterations.

The materno-fetal interface in llama (Lama guanicoe glama)

Samples from 9 llamas (28 through 36 weeks of gestation) were collected and fixed in 4% buffered paraformaldehyde (light microscopy) and in 2.5% buffered glutaraldehyde (transmission and scanning electron microscopy). The material was processed in paraplast and slides (5mm) were stained with HE, PAS, Masson-Trichrome, acid phosphatase and Perl's. The uteroferrin was immunolocalized. The results show that llama placenta is chorioallantoic, diffuse, folded and epitheliochorial, and the fetus is covered with an epidermal membrane. The trophoblast cells have variable morphology: cubic, rounded and triangular cells, with cytoplasm containing PAS-positive granules. Binucleated cells with large cytoplasm and rounded nuclei, as well as giant trophoblastic cells with multiple nuclei were also observed. Numerous blood vessels were observed beneath the cells of the uterine epithelium and around the chorionic subdivided branches. Glandular activity was shown by PAS, Perl's, and acid phosphatase positive reactions in the cytoplasm and glandular lumen, and by immunolocalization of the uteroferrin in the glandular epithelium. The uterine glands open in spaces formed by the areoles, which are filled by PAS-positive material. The llama fetus was covered by the epidermal membrane, composed of stratified epithelium, with up to seven layers of mono-, bi- or trinucleated cells. The high level of maternal and fetal vascularization surfaces indicates an intense exchange of substances across both surfaces. The metabolic activity shown in the uterine glands suggests an adaptation of the gestation to the high altitudes of the natural habitat of this species.

Cutaneous Rosai-Dorfman disease: clinicopathological profiles, spectrum and evolution of 21 lesions in six patients

An uncommon histiocytosis primarily involving the lymph nodes, Rosai-Dorfman disease (RDD, originally called sinus histiocytosis with massive lymphadenopathy) involves extranodal sites in 43% of cases; cutaneous RDD (C-RDD) is a rare form of RDD limited to the skin. The clinicopathological diagnosis of C-RDD may sometimes be difficult, with different clinical profiles from those of its nodal counterpart, and occasionally misleading histological pictures. There have been few multipatient studies of C-RDD and documentation of its histological spectrum is rare. To identify the clinical and histopathological profiles, associated features, and the chronological changes of this rare histiocytosis. From 1991 to 2002, patients diagnosed as having C-RDD were collected in four academic hospitals. Clinical presentations, treatments, and courses of each case were documented. In total, 21 biopsy specimens obtained from these patients were re-evaluated and scored microscopically with attention to the uncommon patterns and chronological evolution both clinically and histologically. We examined six patients with C-RDD, three men and three women. The mean age at the first visit was 43.7 years. The clinical presentations were mostly papules, nodules and plaques, varying with the duration and depth of lesions. Although the anatomical distribution was wide, the face was most commonly involved. Evolutional changes were identified clinically, as the lesions typically began with papules or plaques and grew to form nodules with satellite lesions and resolved with fibrotic plaques before complete remission. No patient had lymphadenopathy or extracutaneous lesions during follow-up (mean 50.5 months). At the end of follow-up, the lesions in four patients had completely resolved irrespective of treatment; two patients had persistent lesions. The histopathological pattern of the main infiltrate, the components of cells and the stromal responses showed dynamic changes according to the duration of lesions. The characteristic Rosai-Dorfman cells (RD cells) were found in association with a nodular or diffuse infiltrate in 15 lesions (71%). Four lesions (19%) demonstrated a patchy/interstitial pattern. One lesion (5%) assumed the pattern of a suppurative granuloma. RD cells were less readily found in these atypical patterns. Conspicuous proliferation of histiocytes associated with RD cells was found in three lesions, including xanthoma, localized Langerhans cell histiocytosis and xanthogranuloma. Along with lymphocytes, plasma cells were present in all lesions, often in large numbers with occasional binucleated or trinucleated cells. Variably found in the lesions were neutrophils (nine lesions, 43%) and eosinophils (13 lesions, 62%). The former occasionally formed microabscesses, while the latter were often few in number. Vascular proliferation was a relatively constant feature (90%). Fibrosis was found in 10 lesions (48%). Our study further confirms that C-RDD is a distinct entity with different age and possibly race distributions from RDD. Compared with its nodal counterpart, C-RDD demonstrates a wider histopathological spectrum with different clinicopathological phases depending on duration of the lesions. Awareness of these features is helpful in making a correct diagnosis. The associations of C-RDD with other histiocytoses may have important implications for the pathogenesis of this rare histiocytosis.

Lavagem traqueobrônquica por sondagem nasotraqueal em bezerros

Avaliou-se a técnica de lavagem traqueobrônquica por sondagem nasotraqueal e caracterizou-se a população celular em 10 bezerros clinicamente sadios. Após a contenção dos animais em decúbito lateral e auxílio de sonda guia, foi introduzida uma sonda de menor diâmetro até a bifurcação da traquéia, para produzir tosse e obter o lavado traqueobrônquico. A média de células totais nas amostras de lavado foi de 133.750 células/ml. À citologia, foram observados na contagem diferencial: 77,2% macrófagos, 14,9% células epiteliais cilíndricas, 6,0% neutrófilos e 1,8% linfócitos. Das células epiteliais cilíndricas, 79,0% eram do tipo ciliadas e 21,0% não-ciliadas. A média de contagem de macrófagos binucleados foi de 78,5 células/lâmina, a de macrófagos trinucleados de 20,5/lâmina e a de células gigantes 28,5/lâmina. Concluiu-se que o método de colheita por sondagem nasotraqueal é eficiente para caracterizar a citologia do lavado traqueobrônquico de bezerros clinicamente sadios.

Acentromeric micronuclei are increased in peripheral blood lymphocytes of untreated cancer patients

Increased micronucleated cell rates, dicentric chromosomes, and other chromosomal damages have been reported in lymphocytes of cancer patients prior to the initiation of chemotherapy, and/or radiotherapy. The cause of these chromosomal damages in these lymphocytes remains unclear. In the present work, we investigated whether these micronuclei mainly reflect structural or numerical chromosomal aberrations by applying the cytokinesis-blocked micronucleus (CBMN) assay in combination with fluorescent in situ hybridization (FISH) of a DNA centromeric probe on blood samples of 10 untreated cancer patients (UCPs), and 10 healthy subjects (HSs). Micronucleated binucleated lymphocyte rate was significantly increased in patients (mean±S.D.: 19.0‰±14.1 versus 9.2‰±4.6 in controls). Trinucleated cytokinesis-blocked cells were not significantly higher in patients than in controls. Acentromeric, centromeric, and multicentromeric micronucleus levels were two-fold higher in patients than in controls, but the difference was significant only with acentromeric micronuclei. The percentage of micronuclei containing one or more centromeres averaged 69.2, and 71.5% in patients, and controls, respectively. The percentage of micronuclei containing several centromeres was 44.7% in patients, and 54.6% in controls. Among centromere-positive micronuclei, the percentage of micronuclei containing several centromeres averaged 59.7% in patients, and 75.4% in controls. These results indicate that genetic instability in peripheral blood lymphocytes of UCPs occurs because of enhanced chromosome breakage. However, a substantial proportion of this genetic instability occurs because of defects in chromosome segregation.

Regulation of DNA synthesis: age-dependent cooperation among G1 cells upon fusion.

The object of this study was to determine whether the inducer(s) of DNA synthesis in mammalian cells accumulates gradually throughout the G1 period or becomes available suddenly at the G1-S transition. HeLa cells, synchronized at various points in the G1 period, were fused by using UV-inactivated Sendai virus. Early G1 cells were fused with mid-G1 or late G1 cells and late G1 cells were fused with mid-G1 cells. The G1 traverse of mono-, bi-, and trinucleated cells was studied. The bi- and trinucleated cells of mid-G1 and late G1 parents traversed the G1 period significantly faster than did their mononucleated counterparts. The reduction in the duration of the G1 period was proportional to the number and age of nuclei at the time of fusion. There was no significant difference between the mono- and binucleated cells of the early G1 parent in their rates of entry into S period. In light of these findings, a model is proposed in which the inducer(s) of DNA synthesis accumulates gradually throughout the G1 period, reaching a critical level at the G1-S boundary when DNA replication is initiated; after reaching a peak during early or mid-S period, it declines to below the critical level when DNA synthesis ceases.

BINUCLEATE AND TRINUCLEATE OOCYTES IN POST-OVULATION OVARIES OF RANA PIPIENS

1. Fifty-six binucleated and two trinucleated cells were found among 249,616 young transparent primary oocytes in post-ovulation ovaries of 25 females of Rana pipiens (0.022%).2. Multinucleated oocytes were absent in 15 females, present in 10. Eight of these 10 produced only one to four binucleates each: one female was the source of 1 tri- and 14 binucleates; and another gave the remarkable number of 24 (0.175%), all binucleates.3. The multiple nucleated condition did not seem to be correlated in any way with the size of the female, the abundance of her eggs, nor the size of the egg.4. In none of the binucleates did the volume of either of the two nuclei approach that of the nucleus of a mononucleate of the same size. The one exception was egg #43 where one nucleus did have exactly the volume of the control, the other about half that.In 29 of the 31 measurable binucleates the two nuclei were of approximately the same size and the sum of the two volumes equalled that of the mononucleate, except in one case where it was half.The origin of binucleate oocytes remains uncertain; it may be connected with a final division of an oogonial nucleus that was not followed by cell division. In this case the two nuclei would both be diploid.5. The two trinucleates conformed in general to the same pattern as the binucleates as to their distribution, size of oocyte, and the volumes of their nuclei. The sum of the three nuclear volumes approximated that of the mononucleate.6. In two cases where the nuclei were markedly unequal in size, there was a definite difference in the appearance of their chromatic granules. These bodies were more abundant and finer in the larger nuclei, peripheral, larger and more distinct in the smaller nucleus.