Effects of submaximal exercise with water ingestion on intraocular pressure in healthy human males

The effects of exercise and water replacement on intraocular pressure (IOP) have not been well established. Furthermore, it is not known whether the temperature of the fluid ingested influences the IOP response. In the present study we determined the effect of water ingestion at three temperatures (10, 24 and 38ºC; 600 ml 15 min before and 240 ml 15, 30 and 45 min after the beginning of each experimental session) on the IOP of six healthy male volunteers (age = 24.0 ± 3.5 years, weight = 67.0 ± 4.8 kg, peak oxygen uptake (VO2peak) = 47.8 ± 9.1 ml kg-1 min-1). The subjects exercised until exhaustion on a cycle ergometer at a 60% VO2peak in a thermoneutral environment. IOP was measured before and after exercise and during recovery (15, 30 and 45 min) using the applanation tonometry method. Skin and rectal temperatures, heart rate and oxygen uptake were measured continuously. IOP was similar for the right eye and the left eye and increased post-water ingestion under both exercising and resting conditions (P<0.05) but did not differ between resting and exercising situations, or between the three water temperatures. Time to exhaustion was not affected by the different water temperatures. Rectal temperature, hydration status, heart rate, oxygen uptake, carbon dioxide extraction and lactate concentration were increased by exercise but were not affected by water temperature. We conclude that IOP was not affected by exercise and that water ingestion increased IOP as expected, regardless of water temperature.

Sm14 gene expression in different stages of the Schistosoma mansoni life cycle and immunolocalization of the Sm14 protein within the adult worm

Sm14 is a 14-kDa vaccine candidate antigen from Schistosoma mansoni that seems to be involved in cytoplasmic trafficking of fatty acids. Although schistosomes have a high requirement for lipids, they are not able to synthesize fatty acids and sterols de novo. Thus, they must acquire host lipids. In order to determine whether Sm14 is present in different stages of the life cycle of the parasite, we performed RT-PCR. Sm14 mRNA was identified in all stages of the life cycle studied, mainly schistosomulum, adult worm and egg. Additionally, we used a rabbit anti-Sm14 polyclonal antibody in an indirect immunofluorescence assay to localize Sm14 in adult worm sections. The basal lamella of the tegument and the gut epithelium were strongly labeled. These tissues have a high flow of and demand for lipids, a finding that supports the putative role of Sm14 as an intracellular transporter of fatty acids from host cells.

Interruption of the blood-stage cycle of the malaria parasite, Plasmodium chabaudi, by protein tyrosine kinase inhibitors

Malaria is a devastating disease caused by a unicellular protozoan, Plasmodium, which affects 3.7 million people every year. Resistance of the parasite to classical treatments such as chloroquine requires the development of new drugs. To gain insight into the mechanisms that control Plasmodium cell cycle, we have examined the effects of kinase inhibitors on the blood-stage cycle of the rodent malaria parasite, Plasmodium chabaudi. In vitro incubation of red blood cells for 17 h at 37ºC with the inhibitors led to a decrease in the percent of infected cells, compared to control treatment, as follows: genistein (200 µM - 75%), staurosporine (1 µM - 58%), R03 (1 µM - 75%), and tyrphostins B44 (100 µM - 66%) and B46 (100 µM - 68%). All these treatments were shown to retard or prevent maturation of the intraerythrocytic parasites. The diverse concentration ranges at which these inhibitors exert their effects give a clue as to the types of signals that initiate the transitions between the different developmental stages of the parasite. The present data support our hypothesis that the maturation of the intraerythrocytic cycle of malaria parasites requires phosphorylation. In this respect, we have recently reported a high Ca2+ microenvironment surrounding the parasite within red blood cells. Several kinase activities are modulated by Ca2+. The molecular identification of the targets of these kinases could provide new strategies against malaria.

The terrestrial Gastropoda Megalobulimus abbreviatus as a useful model for nociceptive experiments: effects of morphine and naloxone on thermal avoidance behavior

We describe the behavior of the snail Megalobulimus abbreviatus upon receiving thermal stimuli and the effects of pretreatment with morphine and naloxone on behavior after a thermal stimulus, in order to establish a useful model for nociceptive experiments. Snails submitted to non-functional (22ºC) and non-thermal hot-plate stress (30ºC) only displayed exploratory behavior. However, the animals submitted to a thermal stimulus (50ºC) displayed biphasic avoidance behavior. Latency was measured from the time the animal was placed on the hot plate to the time when the animal lifted the head-foot complex 1 cm from the substrate, indicating aversive thermal behavior. Other animals were pretreated with morphine (5, 10, 20 mg/kg) or naloxone (2.5, 5.0, 7.5 mg/kg) 15 min prior to receiving a thermal stimulus (50ºC; N = 9 in each group). The results (means ± SD) showed an extremely significant difference in response latency between the group treated with 20 mg/kg morphine (63.18 ± 14.47 s) and the other experimental groups (P < 0.001). With 2.5 mg/kg (16.26 ± 3.19 s), 5.0 mg/kg (11.53 ± 1.64 s) and 7.5 mg/kg naloxone (7.38 ± 1.6 s), there was a significant, not dose-dependent decrease in latency compared to the control (33.44 ± 8.53 s) and saline groups (29.1 ± 9.91 s). No statistically significant difference was found between the naloxone-treated groups. With naloxone plus morphine, there was a significant decrease in latency when compared to all other groups (minimum 64% in the saline group and maximum 83.2% decrease in the morphine group). These results provide evidence of the involvement of endogenous opioid peptides in the control of thermal withdrawal behavior in this snail, and reveal a stereotyped and reproducible avoidance behavior for this snail species, which could be studied in other pharmacological and neurophysiological studies.

NFAT transcription factors: from cell cycle to tumor development

The nuclear factor of activated T cells (NFAT) family of transcription factors has been primarily identified in immune cells; however, these proteins have been recently found to be functionally active in several other non-immune cell types. NFAT proteins are activated upon different stimuli that lead to increased intracellular calcium levels. Regardless of their widely known cytokine gene expression properties, NFATs have been shown to regulate other genes related to cell cycle progression, cell differentiation and apoptosis, revealing a broader role for these proteins in normal cell physiology. Several reports have addressed the participation of NFATs in many aspects of malignant cell transformation and tumorigenic processes. In this review, we will discuss the involvement of the different NFAT family members in the regulation of cell cycling, differentiation and tumor formation, and also its implications on oncogenesis. Better understanding the mechanisms by which NFATs regulate cell cycle and tumor-related events should be relevant for the development of rational anti-cancer therapies.

Effects of gamma-radiation on cell growth, cycle arrest, death, and superoxide dismutase expression by DU 145 human prostate cancer cells

Gamma-irradiation (gamma-IR) is extensively used in the treatment of hormone-resistant prostate carcinoma. The objective of the present study was to investigate the effects of 60Co gamma-IR on the growth, cell cycle arrest and cell death of the human prostate cancer cell line DU 145. The viability of DU 145 cells was measured by the Trypan blue exclusion assay and the 3(4,5-dimethylthiazol-2-yl)-2,5,diphenyltetrazolium bromide test. Bromodeoxyuridine incorporation was used for the determination of cell proliferation. Cell cycle arrest and cell death were analyzed by flow cytometry. Superoxide dismutase (SOD), specifically CuZnSOD and MnSOD protein expression, after 10 Gy gamma-IR, was determined by Western immunoblotting analysis. gamma-IR treatment had a significant (P < 0.001) antiproliferative and cytotoxic effect on DU 145 cells. Both effects were time and dose dependent. Also, the dose of gamma-IR which inhibited DNA synthesis and cell proliferation by 50% was 9.7 Gy. Furthermore, gamma-IR induced cell cycle arrest in the G2/M phase and the percentage of cells in the G2/M phase was increased from 15% (control) to 49% (IR cells), with a nonsignificant induction of apoptosis. Treatment with 10 Gy gamma-IR for 24, 48, and 72 h stimulated CuZnSOD and MnSOD protein expression in a time-dependent manner, approximately by 3- to 3.5-fold. These data suggest that CuZnSOD and MnSOD enzymes may play an important role in the gamma-IR-induced changes in DU 145 cell growth, cell cycle arrest and cell death.

Luteal phase of the menstrual cycle increases sweating rate during exercise

The present study evaluated whether the luteal phase elevation of body temperature would be offset during exercise by increased sweating, when women are normally hydrated. Eleven women performed 60 min of cycling exercise at 60% of their maximal work load at 32ºC and 80% relative air humidity. Each subject participated in two identical experimental sessions: one during the follicular phase (between days 5 and 8) and the other during the luteal phase (between days 22 and 25). Women with serum progesterone >3 ng/mL, in the luteal phase were classified as group 1 (N = 4), whereas the others were classified as group 2 (N = 7). Post-exercise urine volume (213 ± 80 vs 309 ± 113 mL) and specific urine gravity (1.008 ± 0.003 vs 1.006 ± 0.002) changed (P < 0.05) during the luteal phase compared to the follicular phase in group 1. No menstrual cycle dependence was observed for these parameters in group 2. Sweat rate was higher (P < 0.05) in the luteal (3.10 ± 0.81 g m-2 min-1) than in the follicular phase (2.80 ± 0.64 g m-2 min-1) only in group 1. During exercise, no differences related to menstrual cycle phases were seen in rectal temperature, heart rate, rate of perceived exertion, mean skin temperature, and pre- and post-exercise body weight. Women exercising in a warm and humid environment with water intake seem to be able to adapt to the luteal phase increase of basal body temperature through reduced urinary volume and increased sweating rate.

Protein-energy malnutrition halts hemopoietic progenitor cells in the G0/G1 cell cycle stage, thereby altering cell production rates

Protein energy malnutrition (PEM) is a syndrome that often results in immunodeficiency coupled with pancytopenia. Hemopoietic tissue requires a high nutrient supply and the proliferation, differentiation and maturation of cells occur in a constant and balanced manner, sensitive to the demands of specific cell lineages and dependent on the stem cell population. In the present study, we evaluated the effect of PEM on some aspects of hemopoiesis, analyzing the cell cycle of bone marrow cells and the percentage of progenitor cells in the bone marrow. Two-month-old male Swiss mice (N = 7-9 per group) were submitted to PEM with a low-protein diet (4%) or were fed a control diet (20% protein) ad libitum. When the experimental group had lost about 20% of their original body weight after 14 days, we collected blood and bone marrow cells to determine the percentage of progenitor cells and the number of cells in each phase of the cell cycle. Animals of both groups were stimulated with 5-fluorouracil. Blood analysis, bone marrow cell composition and cell cycle evaluation was performed after 10 days. Malnourished animals presented anemia, reticulocytopenia and leukopenia. Their bone marrow was hypocellular and depleted of progenitor cells. Malnourished animals also presented more cells than normal in phases G0 and G1 of the cell cycle. Thus, we conclude that PEM leads to the depletion of progenitor hemopoietic populations and changes in cellular development. We suggest that these changes are some of the primary causes of pancytopenia in cases of PEM.

Estrous cycle and stress: influence of progesterone on the female brain

The female brain operates in a constantly changing chemical milieu caused by cyclical changes in gonadal hormones during the estrous cycle (menstrual cycle in women). Such hormones are highly lipophilic and pass readily from the plasma to the brain where they can influence neuronal function. It is becoming clear that the rapid reduction in peripheral circulating progesterone, which occurs during the late diestrous phase of the cycle, can trigger a withdrawal-like response, in which changes in GABA A receptor expression render hyper-responsive certain brain areas involved in processing responses to stressful stimuli. The periaqueductal gray matter (PAG) is recognised as an important region for integrating anxiety/defence responses. Withdrawal from progesterone, via actions of its neuroactive metabolite allopregnanolone, triggers up-regulation of extrasynaptic GABA A receptors on GABAergic neurons in the PAG. As a consequence, ongoing GABAergic tone on the output cells decreases, leading to an increase in functional excitability of the circuitry and enhanced responsiveness to stressful stimuli during the late diestrous phase. These changes during late diestrus could be prevented by short-term neurosteroid administration, timed to produce a more gradual fall in the peripheral concentration of allopregnanolone than the rapid decrease that occurs naturally, thus removing the trigger for the central withdrawal response.

REGγ is a strong candidate for the regulation of cell cycle, proliferation and the invasion by poorly differentiated thyroid carcinoma cells

REGγ is a proteasome activator that facilitates the degradation of small peptides. Abnormally high expression of REGγ has been observed in thyroid carcinomas. The purpose of the present study was to explore the role of REGγ in poorly differentiated thyroid carcinoma (PDTC). For this purpose, small interfering RNA (siRNA) was introduced to down-regulate the level of REGγ in the PDTC cell line SW579. Down-regulation of REGγ at the mRNA and protein levels was confirmed by RT-PCR and Western blot analyses. FACS analysis revealed cell cycle arrest at the G1/S transition, the MTT assay showed inhibition of cell proliferation, and the Transwell assay showed restricted cell invasion. Furthermore, the expression of the p21 protein was increased, the expression of proliferating cell nuclear antigen (PCNA) protein decreased, and the expression of the p27 protein was unchanged as shown by Western blot analyses. REGγ plays a critical role in the cell cycle, proliferation and invasion of SW579 cells. The alteration of p21 and PCNA proteins related to the down-regulation of REGγ suggests that p21 and PCNA participate in the process of REGγ regulation of cell cycle progression and cell proliferation. Thus, targeting REGγ has a therapeutic potential in the management of PDTC patients.

A phthalide derivative isolated from endophytic fungiPestalotiopsis photiniae induces G1 cell cycle arrest and apoptosis in human HeLa cells

MP [4-(3′,3′-dimethylallyloxy)-5-methyl-6-methoxyphthalide] was obtained from liquid culture of Pestalotiopsis photiniaeisolated from the Chinese Podocarpaceae plant Podocarpus macrophyllus. MP significantly inhibited the proliferation of HeLa tumor cell lines. After treatment with MP, characteristic apoptotic features such as DNA fragmentation and chromatin condensation were observed in DAPI-stained HeLa cells. Flow cytometry showed that MP induced G1 cell cycle arrest and apoptosis in a dose-dependent manner. Western blotting and real-time reverse transcription-polymerase chain reaction were used to investigate protein and mRNA expression. MP caused significant cell cycle arrest by upregulating the cyclin-dependent kinase inhibitor p27KIP1 protein and p21CIP1 mRNA levels in HeLa cells. The expression of p73 protein was increased after treatment with various MP concentrations. mRNA expression of the cell cycle-related genes, p21CIP1, p16INK4a and Gadd45α, was significantly upregulated and mRNA levels demonstrated significantly increased translation ofp73, JunB, FKHR, andBim. The results indicate that MP may be a potential treatment for cervical cancer.

Early detection of metabolic and energy disorders by thermal time series stochastic complexity analysis

Maintenance of thermal homeostasis in rats fed a high-fat diet (HFD) is associated with changes in their thermal balance. The thermodynamic relationship between heat dissipation and energy storage is altered by the ingestion of high-energy diet content. Observation of thermal registers of core temperature behavior, in humans and rodents, permits identification of some characteristics of time series, such as autoreference and stationarity that fit adequately to a stochastic analysis. To identify this change, we used, for the first time, a stochastic autoregressive model, the concepts of which match those associated with physiological systems involved and applied in male HFD rats compared with their appropriate standard food intake age-matched male controls (n=7 per group). By analyzing a recorded temperature time series, we were able to identify when thermal homeostasis would be affected by a new diet. The autoregressive time series model (AR model) was used to predict the occurrence of thermal homeostasis, and this model proved to be very effective in distinguishing such a physiological disorder. Thus, we infer from the results of our study that maximum entropy distribution as a means for stochastic characterization of temperature time series registers may be established as an important and early tool to aid in the diagnosis and prevention of metabolic diseases due to their ability to detect small variations in thermal profile.

Analysis of heart rate control to assess thermal sensitivity responses in Brazilian toads

In anurans, changes in ambient temperature influence body temperature and, therefore, energy consumption. These changes ultimately affect energy supply and, consequently, heart rate (HR). Typically, anurans living in different thermal environments have different thermal sensitivities, and these cannot be distinguished by changes in HR. We hypothesized that Rhinella jimi (a toad from a xeric environment that lives in a wide range of temperatures) would have a lower thermal sensitivity regarding cardiac control than R. icterica (originally from a tropical forest environment with a more restricted range of ambient temperatures). Thermal sensitivity was assessed by comparing animals housed at 15° and 25°C. Cardiac control was estimated by heart rate variability (HRV) and heart rate complexity (HRC). Differences in HRV between the two temperatures were not significant (P=0.214 for R. icterica and P=0.328 for R. jimi), whereas HRC differences were. All specimens but one R. jimi had a lower HRC at 15°C (all P<0.01). These results indicate that R. jimi has a lower thermal sensitivity and that cardiac control is not completely dependent on the thermal environment because HRC was not consistently different between temperatures in all R. jimi specimens. This result indicates a lack of evolutive trade-offs among temperatures given that heart rate control at 25°C is potentially not a constraint to heart rate control at 15°C.

Cell cycle synchronization and BrdU incorporation as a tool to study the possible selective elimination of ErbB1 gene in the micronuclei in A549 cells

Lung cancer often exhibits molecular changes, such as the overexpression of the ErbB1 gene that encodes epidermal growth factor receptor (EGFR). ErbB1 amplification and mutation are associated with tumor aggressiveness and low response to therapy. The aim of the present study was to design a schedule to synchronize the cell cycle of A549 cell line (a non-small cell lung cancer) and to analyze the possible association between the micronuclei (MNs) and the extrusion of ErbB1 gene extra-copies. After double blocking, by the process of fetal bovine serum deprivation and vincristine treatment, MNs formation was monitored with 5-bromo-2-deoxyuridine (BrdU) incorporation, which is an S-phase marker. Statistical analyses allowed us to infer that MNs may arise both in mitosis as well as in interphase. The MNs were able to replicate their DNA and this process seemed to be non-synchronous with the main cell nuclei. The presence of ErbB1 gene in the MNs was evaluated by fluorescent in situ hybridization (FISH). ErbB1 sequences were detected in the MNs, but a relation between the MNs formation and extrusion of amplified ErbB1could not be established. The present study sought to elucidate the meaning of MNs formation and its association with the elimination of oncogenes or other amplified sequences from the tumor cells.

Antioxidant activity of rosemary and oregano ethanol extracts in soybean oil under thermal oxidation

Four experiments were conducted to measure the antioxidant activity of ethanol extracts of rosemary and oregano compared with synthetic antioxidants such as TBHQ and BHA/BHT. The antioxidant activity was determined and results differed from those of the Oven test at 63º C. Peroxide values and absorptivities at 232 nm of soybean oil under Oven test were lower in treatments with 25, 50, 75, 100 and 200 mg.Kg-1 TBHQ than in treatments with 1000 mg.Kg-1 oregano extract (O), 500 mg.Kg-1 rosemary extract (R) and their mixture R+O. All the treatments were effective in controlling the thermal oxidation of oils; the natural extracts were as effective as BHA+BHT and less effective than TBHQ. The natural extracts were mixed with 25, 50, 75 and 100 mg.Kg-1 TBHQ and then added to the oil. No improvement in antioxidative properties was observed. The best antioxidant concentration could be determined from polynomial regression and quadratic equation from the experimental data.