Scielo RSS <![CDATA[Biological Research]]> vol. 42 num. 4 lang. en <![CDATA[SciELO Logo]]> <![CDATA[<b>Some ultrastructural observations of the vegetative, resting and excysting ciliate, <i>urostyla grandis </i>(urostylidae, hypotrichida)</b>]]> In order to reveal the differentiation characteristics of organelles of ciliates under different physiological status, the cellular ultrastructure of Urostyla grandis was studied by transmission electron microscopy. In the resting cells most ciliary shafts, kinetosomes and sub-pellicle microtubules were resorbed, and the endoplasmic reticulum (ER) disappeared with the autophagocytosis taking place within the cytoplasm. As well, the nuclear matrix of the macronucleus was extruded into the cytoplasm, forming pseudopodia-like structures with large quantities of heterochromatin (CH) attached to the inner nuclear membrane. During excystment, membraneous structures developed and gradually increased in number to form the ER. Autophagic vacuoles (AVs) appeared containing mitochondria, paraglycogen particles (PGP), membranous structures, etc. Moreover, the number of nucleoli decreased with the chromatin, condensing in parallel with the process of recombination. Based on these observations, it could be concluded that the de-differentiation of microtubular organelles and the changes occurring in macronuclei in the resting Urostyla grandis, as well as the differentiation of cytoplasmic organelles, digestion by AVs, and the recombination of chromatin during excystment, are not similar to events that occur in non-kinetosome-resorbing cysts (NKR). <![CDATA[<b>Bioactivity of crude ethanol extract and fractions of <i>Eugenia uniflora </i>(Myrtaceae) in the hepatopancreas of <i>Oreochromis niloticus </i>L</b>]]> This study evaluates the bioactivity of the crude ethanol extract and ethyl acetate, hexane and chloroform fractions obtained from Eugenia uniflora leaves using the hepatopancreas of Oreochromis niloticus L. as an experimental model. The ethanol extract and fractions were administered to the fish orally with their feed. Twenty-four hours later, the fish were sacrificed and their livers dissected, fixed in neutral formalin, embedded in paraffin and sectioned. Histological analyses were performed using Masson's trichrome and Haematoxylin-Eosin. Histochemical studies were performed using Feulgen, PAS (Periodic Acid Schiff) and PAS + salivary amylase and Sudan IV stain. The qualitative analysis of the material showed that the crude extract and the ethyl, chloroform and hexane fractions induced vasodilation, vascular congestion and toxicity due to the presence of eosinophilic granular cells, rodlet cells, some leukocytic infiltrate and rare focal necroses. The Nile tilapia proved to be a satisfactory model for screening plant products. <![CDATA[<b>Effect of the monocyte locomotion inhibitory factor (MLIF) produced by <i>E. histolityca </i>on cytokines and chemokine receptors in T CD4+ lymphocytes</b>]]> Entamoeba histolytica produces Monocyte Locomotion Inhibitory Factor (MLIF), which may contribute to the delayed inflammation observed in amoebic hepatic abscesses. Leukocytes are affected through the modulation of cytokine expression and/or production. We evaluated the effects of MLIF on the activation and production of intracellular cytokines in human CD4+ T lymphocytes by flow cytometry. Cells were stimulated for 24 h with PMA, MLIF, or PMA+MLIF. Cellular activation was measured using anti-CD69. Th1/Th2 production was studied by the expression of intracellular cytokines and cytokine/chemokine receptors. MLIF increased CD69 and induced the over-expression of the IL-lß, IFN-γ, IL-2, IL-4, and IL-10 intracellular cytokines; PMA+MLIF inhibited Th1 cytokine (IFN-γ) and increased Th2 cytokines (IL-4 and IL-10). The co-expression of the cytokine and chemokine receptors IFN-γ/CCR5 and IL-1ß/CCR5 was inhibited by PMA+MLIF and Th2 co-expression was increased. MLIF effects varied depending on the conditions. MLIF alone activated the Th1 and Th2 cytokines and cytokine/receptor expression; however, PMA+MLIF increased the expression of Th2 but inhibited it in Th1. <![CDATA[<b>Biofilm formation and acyl homoserine lactone production in <i>Hafnia alvei </i>isolated from raw milk</b>]]> The objective of this study was to detect the presence of acyl homoserine lactones (AHLs), signal molecules of the quorum sensing system in biofilm formed by Hafnia alvei strains. It also evaluated the effect of synthetic quorum sensing inhibitors in biofilm formation. AHLs were assayed using well diffusion techniques, thin layer chromatography (TLC) and detection directly in biofilm with biomonitors. The extracts obtained from planktonic and sessile cell of H. alvei induced at least two of three monitor strains evaluated. The presence of AHLs with up to six carbon atoms was confirmed by TLC. Biofilm formation by H. alvei was inhibited by furanone, as demonstrated by 96-well assay of crystal violet in microtitre plates and by scanning electron microscopy. The H. alvei 071 hall mutant was deficient in biofilm formation. All these results showed that the quorum sensing system is probably involved in the regulation of biofilm formation by H. alvei. <![CDATA[<b>Mycophenolate mofetil versus Rapamycin in Han</b>: <b>SPRD rats with Polycystic Kidney Disease</b>]]> Objective: We aimed to explore the effect of Mycophenolate mofetil (MMF) on loss of renal function and cyst progression compared to rapamycin in Han: SPRD rats. We also sought to assess whether the effect of combination therapy of MMF plus rapamycin was better than that of monotherapy. Methods: Sixty heterozygous (Cy/+) and littermate control (+/+) male Han: SPRD rats were weaned at 4 weeks of age, then divided into four groups randomly to receive different treatments by intragastric administration for 2 months: vehicle-treated group as control, MMF-treated group (20mg/kg/day), rapamycin-treated group (2mg/kg/day), and MMF+Rapa- treated group (MMF 20mg/kg/day plus Rapamycin 2mg/kg/day). Resulls: After 2 months of treatment, rapamycin caused a 22 % decrease in body weight in comparison to the control group, whereas MMF had no significant effect on weight gain. The steady increase of BUN in Cy/+ rats was reduced by 15% in MMF-treated Cy/+ rats. However, rapamycin and combination therapy reduced BUN by 42% and 43%, respectively. CCr was 0.93±0.11ml/min in vehicle-treated Cy/+ rats, 1.67±0.23 ml/min in MMF-treated Cy/+ rats (P<0.05), 1.72±0.44 ml/min and 1.83±0.21 ml/min in rapamycin- and MMF+Rapa-treated Cy/+ rats, respectively (.P<0.01). Cyst volume density was 57.1 % in vehicle-treated Cy/+ rats, 45.2% in MMF-treated Cy/+ rats (P<0.05), 32.9% and 37.7% in rapamycin- and MMF+Rapa-treated Cy/+ rats, respectively (P<0.01). MMF markedly ameliorated interstitial inflammation and fibrosis. Rapamycin showed a similar effect on interstitial inflammation and fibrosis, but to a lesser degree. Conclusion : MMF is more tolerable than rapamycin and can retard deterioration of renal function in Han: SPRD rats, though its effect is weaker than that of rapamycin. Combination therapy does not exert more favorable effect than monotherapy. <![CDATA[<b>Alterations in core histone variant ratios during maize root differentiation, callus formation and in response to plant hormone treatment</b>]]> Although several histone variants have been studied in both animal and plant organisms, little is known about their distribution during processes that involve alterations in chromatin function, such as differentiation, dedifferentiation and hormone treatment. In this study we evaluated the ratio of each histone variant in each of the four core histone classes in the three developmental zones of maize (Zea mays L.) root and in callus cultures derived from them, in order to define possible alterations either during plant cell differentiation or dedifferentiation. We also evaluated core histone variant ratios in the developmental zones of roots treated with auxin and gibberellin in order to examine the effects of exogenously applied plant hormones to histone variant distribution. Finally, immunohistochemical detection was used to identify the root tissues containing modified forms of core histones and correlates them with the physiological status of the plant cells. According to the results presented in this study, histone variant ratios are altered in all the cases examined, i.e. in the developmental zones of maize root, in callus cultures derived from them and in the developmental zones of roots treated either with auxin or gibberellin. We propose that the alterations in linker histone variant ratios are correlated with plant cell differentiation and physiological status in each case. <![CDATA[<b>VEGFA polymorphisms and cardiovascular anomalies in 22q11 microdeletion syndrome</b>: <b>a case-control and family-based study</b>]]> Microdeletion 22q11 in humans causes velocardiofacial and DiGeorge syndromes. Most patients share a common 3Mb deletion, but the clinical manifestations are very heterogeneous. Congenital heart disease is present in 50-80% of patients and is a significant cause of morbidity and mortality. The phenotypic variability suggests the presence of modifiers. Polymorphisms in the VEGFA gene, coding for the vascular endothelial growth factor A, have been associated with non-syndromic congenital heart disease, as well as with the presence of cardiovascular anomalies in patients with microdeletion 22q11. We evaluated the association of VEGFA polymorphisms c.-2578C>A (rs699947), c.-1154G>A (rs1570360) and c.-634C>G (rs2010963) with congenital heart disease in Chilean patients with microdeletion 22q11. The study was performed using case-control and family-based association designs. We evaluated 122 patients with microdeletion 22q11 and known anatomy of the heart and great vessels, and their parents. Half the patients had congenital heart disease. We obtained no evidence of association by either method of analysis. Our results provide further evidence of the incomplete penetrance of the cardiovascular phenotype of microdeletion 22ql 1, but do not support association between VEGFA promoter polymorphisms and the presence of congenital heart disease in Chilean patients with this syndrome. <![CDATA[<b>Donepezil reverses buprenorphine-induced central respiratory depression in anesthetized rabbits</b>]]> Buprenorphine is a mixed opioid receptor agonist-antagonist used in acute and chronic pain management. Although this agent's analgesic effect increases in a dose-dependent manner, buprenorphine-induced respiratory depression shows a marked ceiling effect at higher doses, which is considered to be an indicator of safety. Nevertheless, cases of overdose mortality or severe respiratory depression associated with buprenorphine use have been reported. Naloxone can reverse buprenorphine-induced respiratory depression, but is slow-acting and unstable, meaning that new drug candidates able to specifically antagonize buprenorphine-induced respiratory depression are needed in order to enable maximal analgesic effect without respiratory depression. Acetylcholine is an excitatory neurotransmitter in central respiratory control. We previously showed that a long-acting acetylcholinesterase inhibitor, donepezil, antagonizes morphine-induced respiratory depression. We have now investigated how donepezil affects buprenorphine-induced respiratory depression in anesthetized, paralyzed, and artificially ventilated rabbits. We measured phrenic nerve discharge as an Índex of respiratory rate and amplitude, and compared discharges following the injection of buprenorphine with discharges following the injection of donepezil. Buprenorphine-induced suppression of the respiratory rate and respiratory amplitude was antagonized by donepezil (78.4 ± 4.8 %, 92.3% ± 22.8 % of control, respectively). These findings indicate that systemically administered donepezil restores buprenorphine-induced respiratory depression in anesthetized rabbits. <![CDATA[<b>DNA damage and related modifier genes in Italian Cystic fibrosis patients</b>]]> Cystic Fibrosis (CF) is an autosomal recessive multisystemic disorder showing a highly heterogeneous phenotype, even among siblings carrying identical CFTR mutations. Moreover, oxidative stress is of central importance in the pathogenesis of cystic fibrosis. The present study seeks to value the presence of oxidative damage in CF patients and the possible modifier effect of repair and glutathione-S-transferase genes. We analysed the presence of DNA damage in leukocytes of 63 CF patients at an Italian CF centre and 63 controls, through the alkaline Comet assay to detect DNA strand breaks. Furthermore, controls and 93 CF subjects were genotyped for 5 genes by RFLP-PCR (XRCC1,0GG1,GSTP1) and PCR assay (GSTM1, GSTT1). No difference in Comet assay values was observed comparing controls to CF patients, although CF subjects showed slightly higher mean values. The crude Odds-Ratio (OR) was higher than one for XRCC1 and GSTP1 genotypes and liver status and for XRCC1 and OGG1 genotypes and pancreatic insufficiency, but in all cases the p-values were not significant. In this case-control study, neither DNA damage ñor gene polymorphisms seem to influence CF manifestation. <![CDATA[<b>Upregulation of liver inducible nitric oxide synthase following thyroid hormone preconditioning</b>: <b>suppression by N-acetylcysteine</b>]]> 3,3-5-L-Triiodothyronine (T3) exerts significant protection against ischemia-reperfusion (IR) liver injury in rats. Considering that the underlying mechanisms are unknown, the aim of this study was to assess the involvement of inducible nitric oxide synthase (iNOS) expression and oxidative stress in T3 preconditioning (PC). Male Sprague-Dawley rats given a single dose of 0.1 mg of T3/kg were subjected to 1-hour ischemia followed by 20 hours reperfusion, in groups of animals pretreated with 0.5 g of N-acetylcysteine (NAC)/kg 0.5-hour prior to T3 or with the respective control vehicles. At the end of the reperfusion period, liver samples were taken for analysis of iNOS mRNA levels (RT-PCR), liver NOS activity, and hepatic histology. T3 protected against hepatic IR injury, with 119% enhancement in liver iNOS mRNA/18S rRNA ratios (p<0.05) and 12.7-fold increase (p<0.05) in NOS activity in T3-treated animals subjected to IR over values in control-sham operated rats, with a net 7.7-fold enhancement (p<0.05) in the net effect of T3 on liver iNOS expression and a net enhancement of 0.58 units in NOS activity, changes that were abolished by NAC treatment before T3. It is concluded that T3-induced liver PC is associated with upregulation of iNOS expression as a protective mechanisms against IR injury, which is achieved through development of transient and reversible oxidative stress. <![CDATA[<b>Y Chromosomal STR Polymorphism in Northern Chinese Populations</b>]]> Y chromosomal STRs show sufficient variability among individduals in a population and a high degree of geographical differentiation, such that their polymorphic character makes them especially suited for population genetic studies. To investígate the polymorphism of a set of 17 Y-STR loci in northern China, we genotyped the 17 Y chromosomal STR loci in a population sample of 377 unrelated males from eight ethnic populations in northern China. We calculated the haplotype frequencies, Rst value and carried out the analysis of molecular variance (AMOVA). We then drew the multidimensional scaling analysis (MDS) plot and phylogenetic tree based on the Rst value. All populations showed a high level of haplotype diversity, with low inter-population variance as measured by an analysis of molecular variance. However, the genetic distances were significant when the eight populations were compared to other populations. By MDS and the phylogenetic tree, we found that the eight populations had a close relationship and Xibo had a northeast origination. <![CDATA[<b>Characterization of Calreticulin Expression in Mouse Endometrium during Embryo Implantation</b>]]> Calreticulin (CRT), a Ca2+-binding storage protein and chaperone in the endoplasmic reticulum, modulates cell adhesiveness and integrin-dependent Ca2+ signaling. However, the role of CRT during implantation remains poorly understood. In the present study, we characterized the expression of CRT mRNA and the protein in mouse endometria from pregnancy DI to D7. Real-Time PCR and in situ hybridization results showed that the levels of CRT mRNA in the endometria of pregnant mice were significantly higher than those of non-pregnant mice (P<0.05), and increased gradually from pregnancy DI to D4, reaching the máximum level on D4, followed by a plateau from D4 to D7. Using immunofluorescence histochemistry and western blot, changes of CRT expression in the endometria of pregnant mice were consistent with the expression of CRT mRNA. Furthermore, antisense CRT oligodeoxynucleotide was injected into the uterus horns of pregnant mice (D3) to investígate its effect on embryo implantation. The result showed that the number of implanted embryos markedly decreased in the side of uterine horns receiving antisense CRT oligodeoxynucleotide(í><0.05). These findings suggest that CRT may play an important role in embryo implantation in mice. <![CDATA[<b>Induction of programmed cell death by <i>Prangos uloptera, </i>a medicinal plant</b>]]> Inhibition of the cell growth or induction of cell death is the most promising area in cancer therapy. The induction of apoptosis by dichloromethane extract of Prangos uloptera was evaluated on the McCoy cell line. This plant's roots, aerial parts and fruit have medicinal value. Cell growth inhibitory and cell cytotoxicity effects of the extract were assayed by MTT and Trypan-blue tests, respectively. Morphological changes and DNA fragmentation were also evaluated. The viability tests showed 0.49 and 0.3 mg/ml as 50% inhibition concentration and 50% cytotoxicity concentration after 24 hours of treatment, respectively. Fluorescent microscopy analysis revealed chromatin fragmentation and scanning electron microscopy showed cell shrinkage and cytoplasmic blebbing. These findings were confirmed by DNA fragmentation analysis. The results demonstrated efficient induction of apoptosis by the plant extract in moderate concentrations, but administration of higher concentrations showed that the primary manner of cell death was necrosis.