[Show abstract][Hide abstract] ABSTRACT: Soil acidity limits crop yields worldwide and is a common result of aluminum (Al) phytotoxicity, which is known to inhibit root growth. Here, we compared the transcriptome of leaves from maize seedlings grown under control conditions (soil without free Al) and under acidic soil containing toxic levels of Al. This study reports, for the first time, the complex transcriptional changes that occur in the leaves of maize plants grown in acidic soil with phytotoxic levels of Al. Our data indicate that 668 genes were differentially expressed in the leaves of plants grown in acidic soil, which is significantly greater than that observed in our previous work with roots. Genes encoding TCA cycle enzymes were upregulated, although no specific transporter of organic acids was differentially expressed in leaves. We also provide evidence for positive roles for auxin and brassinosteroids in Al tolerance, whereas gibberellin and jasmonate may have negative roles. Our data indicate that plant responses to acidic soil with high Al content are not restricted to the root; tolerance mechanisms are also displayed in the aerial parts of the plant, thus indicating that the entire plant responds to stress.
[Show abstract][Hide abstract] ABSTRACT: Three porphyrins, (5,10,15,20-tetra(3-hydroxyphenyl)porphyrin, 5-hexyl-10,20-bis(3-hydroxyphenyl)-porphyrin and 5-hexyl-10,15,20-tris(3-hydroxyphenyl)porphyrin), with different amphiphilicities and
equal singlet oxygen quantum yields in ethanol, were encapsulated into 50:50 poly(lactide-coglycolide), nanoparticles prepared by the emulsion/evaporation technique. A factorial design was
utilized to evaluate the influence of the porphyrin/polymer mass ratio and the percentage of ethanol
in the aqueous phase on the size and zeta potential of the nanoparticles. Increasing both the
amount of ethanol and the porphyrin/polymer ratio decreases the size and increases zeta potential for the photosensitizers studied, except for hex-m-trisHPP. Entrapment efficiency depended on
the individual m-hydroxyphenylporphyrin and ranged from 69 to 97%. After 1.5 h incubation with
m-hydroxyphenylporphyrin-loaded nanoparticles the percentages of intracellular uptake were the
same for all porphyrins since the molecules are confined in the nanoparticles, hampering the interaction of the amphiphilic photosensitizers with the cellular membrane. All encapsulated porphyrins
caused the same decrease of cell viability and always localized in the perinuclear region of the
cells. Results show that thesem-hydroxyphenylporphyrins, although with different amphiphilicities,
have equal photodynamic efficacies.
Journal of Nanoscience and Nanotechnology 08/2014; 14(8):6274-6286. · 1.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Three clinical trials are being conducted in Brazil by our research group on the use of autologous, BM-derived, hematopoietic SCT (auto-BMHSCT) for the treatment of retinitis pigmentosa (RP), dry age-related macular degeneration and ischaemic retinopathy (including diabetic retinopathy with macular ischaemia). These studies are registered with ClinicalTrials.gov, numbers NCT01068561, NCT01518127 and NCT01518842, respectively. 1 We published the results of phase I of the clinical trial using an intravitreal injection of 0.1 mL of a cell suspension containing 0.92 Â 10 4 –2.91 Â 10 4 (mean, 1.68 Â 10 4) BM-derived hemato-poietic stem cells (CD34 þ) for retinal dystrophy. The second phase of this study (NCT01560715) has already begun, and now we describe the case of a participant in this study who had cystoid macular oedema (CMO) associated with RP. After this, patient underwent treatment with an intraocular injection (intravitreal) of auto-BMHSCT in which the patient showed complete resolution of the oedema 7 days after injection, and this effect remained for 1 month of follow-up, which was observed through examination using optical coherence tomography (Figure 1). 2, 3 The improvement of the oedema led to an improvement in visual acuity (20/50–20/32) and an improvement in macular sensitivity, as measured by the microperimetry test (Figure 2). This patient had macular oedema associated with RP that started 2 years ago and had already undergone treatment with systemic carbonic anhydrase inhibitors (acetazolamide) 250 mg three times daily for 3 months and dorzolamide eye drops three times daily for 1 year without any sign of improvement in macular oedema. This group of drugs is commonly used to treat macular oedema associated with RP. 4–6 Acetazolamide increases fluid transport through the retinal pigment epithelium (RPE) by active ion transport and acidification of the RPE. 7 Macular oedema owing to low-grade inflammation may respond better to this mechanism than the diffuse loss of endothelial tight junctions that occurs in diabetes. Topical carbonic anhydrase inhibitors have a lower rate of systemic side effects than acetazolamide but are less efficacious. 4 CME that is refractory to acetazolamide treatment usually leads to permanent visual acuity loss. 5,6 Thus, the short time of response after the injection of auto-BMHSCT is strong evidence for the action of this therapy. The pathophysiology of macular oedema associated with retinitis is still controversial, but it may possibly be due to the blood–retinal barrier breakdown. However, the mechanism by which auto-BMHSCT led to improvement of the oedema is still Baseline
Bone marrow transplantation 09/2012; · 3.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A 2(4-1) fractional factorial design was utilized to evaluate the influence of four preparation conditions on six characteristics of poly(lactide-co-glycolide) nanospheres loaded with chloro(5,10,15,20-tetraphenylporphyrinato)indium(III). Ethanol in the aqueous phase and the stirring rate were the factors that most influenced the nanosphere characteristics. An increase in these factors caused a decrease in nanosphere size, recovery yield and residual chloroform and an increase in the percent of residual poly(vinyl alcohol). The synergic interaction between these two factors caused an increase in the percent residual chloroform. The entrapment efficiency was increased by an increase of ethanol in the aqueous phase or an increase in the percent poly(vinyl alcohol), but an overall decrease was obtained due to a synergic interaction between these factors. The stirring rate was the only parameter that caused an increase of the zeta potential. Evolutionary operations were then carried out based on the results from the fractional factorial design and nanospheres were obtained with sizes smaller than 200 nm.
Journal of Nanoscience and Nanotechnology 06/2011; 11(6):5234-46. · 1.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To describe the presence of iris neovascularization in a rabbit-model of retinal neovascularization induced by the intravitreal injection of latex-derived angiogenic fraction microspheres (LAF).
Eight New Zealand rabbits received one intravitreal injection of PLGA (L-lactide-co-glycolide) microspheres with 50 ug of LAF in the right eye (Group A). Microspheres without the LAF (0.1 ml) were injected in controls (Group B; n = 8). Follow-up with clinical evaluation and iris fluorescein angiography was performed after 4 weeks when eyes were processed for light microscopy.
All eyes from Group A showed significant vascular dilation, conjunctival hyperemia and neovascularization on the iris surface, after LAF injection. No vascular changes were observed in Group B.
The intravitreal injection of microspheres containing the LAF can induce rubeosis iridis in rabbits and could be used as a simple experimental model for iris neovascularization.
Current eye research 05/2011; 36(9):857-9. · 1.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To create a retinal neovascularization experimental model using intravitreal injection of microspheres loaded with latex-derived angiogenic fraction.
Thirty-two albino New Zealand rabbits, divided in 4 groups of 8 animals, were enrolled in this study. Rabbits in groups I, II, and III received one intravitreal injection of PLGA (L-lactide-co-glycolide) microspheres with 10, 30, and 50 microg of latex-derived angiogenic fraction into their right eyes, respectively, and group IV received 0.1 ml of microspheres without the angiogenic fraction. Weekly follow-up with ophthalmoscopy and fluorescein angiography was performed; the rabbits were sacrificed in the 4th week and their eyes processed for light microscopy.
All eyes from group I demonstrated increased retinal vascular tortuosity, observed from 14 days after injection and maintained for 28 days, otherwise without new vessels detection. All group II eyes showed vascular changes similar to group I. Fifty percent of the eyes from group II rabbits developed retinal neovascularization 21 days after injection. All eyes from group III demonstrated significant vascular tortuosity and retinal new vessels 2 weeks after injection, progressing to fibrovascular proliferation and tractional retinal detachment. No vascular changes or retinal new vessels were observed in group IV eyes. Light microscopy confirmed the existence of new vessels previously seen on fluorescein angiography, in retinal sections adjacent to the optic disc, not observed in sections at the same area in the control group.
Thirty- and 50-microg microspheres containing latex-derived angiogenic fraction injected into the vitreous cavity induced retinal neovascularization in rabbits.
Current eye research 01/2010; 35(1):56-62. · 1.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A new technique using diffuse reflectance spectroscopy was developed that enables the
monitoring of encapsulated drug release without particle separation from the assayed medium.
Studies of chloro(5,10,15,20-tetraphenylporphyrinato)indium(III) (InTPP) release from
microspheres and nanospheres of poly(lactide-co-glycolide) (PLGA) were performed using this
new technique. The release of InTPP was biphasic, with an initial fast release followed by a second
slower release. Mathematical models applied to the release profiles showed that the release of
InTPP from the nanospheres was controlled by diffusion, which is to be expected for a substance
homogeneously dispersed within the spheres. However, due to the large size distribution of the
microspheres loaded with InTPP, the release profiles were irregular, hampering an adequate fit
to mathematical models. Confocal analysis of microparticles showed that the InTPP appeared to
be homogenously distributed within the microspheres and no preferential distribution of InTPP
towards the interior or towards the surface of the spheres was observed.
Journal of the Brazilian Chemical Society 01/2010; 21(2):214. · 1.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Aluminum (Al) toxicity is one of the most important yield-limiting factors of many crops worldwide. The primary symptom of Al toxicity syndrome is the inhibition of root growth leading to poor water and nutrient absorption. Al tolerance has been extensively studied using hydroponic experiments. However, unlike soil conditions, this method does not address all of the components that are necessary for proper root growth and development. In the present study, we grew two maize genotypes with contrasting tolerance to Al in soil containing toxic levels of Al and then compared their transcriptomic responses.
When grown in acid soil containing toxic levels of Al, the Al-sensitive genotype (S1587-17) showed greater root growth inhibition, more Al accumulation and more callose deposition in root tips than did the tolerant genotype (Cat100-6). Transcriptome profiling showed a higher number of genes differentially expressed in S1587-17 grown in acid soil, probably due to secondary effects of Al toxicity. Genes involved in the biosynthesis of organic acids, which are frequently associated with an Al tolerance response, were not differentially regulated in both genotypes after acid soil exposure. However, genes related to the biosynthesis of auxin, ethylene and lignin were up-regulated in the Al-sensitive genotype, indicating that these pathways might be associated with root growth inhibition. By comparing the two maize lines, we were able to discover genes up-regulated only in the Al-tolerant line that also presented higher absolute levels than those observed in the Al-sensitive line. These genes encoded a lipase hydrolase, a retinol dehydrogenase, a glycine-rich protein, a member of the WRKY transcriptional family and two unknown proteins.
This work provides the first characterization of the physiological and transcriptional responses of maize roots when grown in acid soil containing toxic levels of Al. The transcriptome profiles highlighted several pathways that are related to Al toxicity and tolerance during growth in acid soil. We found several genes that were not found in previous studies using hydroponic experiments, increasing our understanding of plant responses to acid soil. The use of two germplasms with markedly different Al tolerances allowed the identification of genes that are a valuable tool for assessing the mechanisms of Al tolerance in maize in acid soil.
[Show abstract][Hide abstract] ABSTRACT: cDNA arrays are a powerful tool for discovering gene expression patterns. Nylon arrays have the advantage that they can be re-used several times. A key issue in high throughput gene expression analysis is sensitivity. In the case of nylon arrays, signal detection can be affected by the plastic bags used to keep membranes humid. In this study, we evaluated the effect of five types of plastics on the radioactive transmittance, number of genes with a signal above the background, and data variability. A polyethylene plastic bag 69 μm thick had a strong shielding effect that blocked 68.7% of the radioactive signal. The shielding effect on transmittance decreased the number of detected genes and increased the data variability. Other plastics which were thinner gave better results. Although plastics made from polyvinylidene chloride, polyvinyl chloride (both 13 μm thick) and polyethylene (29 and 7 μm thick) showed different levels of transmittance, they all gave similarly good performances. Polyvinylidene chloride and polyethylene 29 mm thick were the plastics of choice because of their easy handling. For other types of plastics, it is advisable to run a simple check on their performance in order to obtain the maximum information from nylon cDNA arrays.
Ciência e Agrotecnologia 01/2009; 33. · 0.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In(III)-meso-tetraphenylporphyrin (InTPP) was encapsulated into nanoparticles (smaller than 200 nm) of poly(d,l-lactide-co-glycolide) (PLGA) using the emulsification-evaporation technique. The photodynamic efficacy of InTPP-loaded nanoparticles and its cellular uptake was investigated with LNCaP prostate tumour cells, in comparison with the free InTPP. The effects of incubation time (1-3h), drug concentration (1.8-7.7 micromol/L) and incident light dose (15-45 J/cm(2)) with both encapsulated and free InTPP were studied. The type of cell death induced by the photochemical process using both encapsulated and free InTPP was also investigated. Cell viability was reduced more significantly with increasing values of these effects for InTPP-loaded nanoparticles than with the free drug. The cellular death induced by both encapsulated and free InTPP was preponderantly apoptotic. Confocal laser scanning microscopy data showed that the InTPP-loaded nanoparticles, as well free InTPP, were localized in the cells, and always in the perinuclear region. Encapsulated InTPP was measured by the intensity of fluorescence intensity of cell extracts and was three times more internalized into the cells than was the free InTPP. Electron paramagnetic resonance experiments corroborated the participation of singlet oxygen in the photocytotoxic effect of nanoparticles loaded with InTPP.
Journal of Photochemistry and Photobiology B Biology 12/2008; 94(2):101-12. · 3.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This in vivo study assessed and compared the effectiveness of an aqueous indocyanine green (ICG) formulation (R-ICG) and a lipid ICG formulation (L-ICG) in occluding the rabbit choriocapillaris, and determined the singlet oxygen quantum yields and aggregation properties of both formulations in vitro.
Singlet oxygen production and aggregation were compared. The eye fundus of 30 albino rabbits was irradiated 0-15 min after dye injection using an 810 nm diode laser. Fluorescein angiography and light microscopy were used to evaluate the safety and efficacy of R-ICG and L-ICG.
L-ICG decreased the dimerisation constant and the tendency of ICG to form aggregates, and increased the efficiency of ICG in generating singlet oxygen (R-ICG, PhiDelta = 0.120 and L-ICG, PhiDelta = 0.210). Using a 10 mg/kg dose, choriocapillaris occlusion was achieved at a light dose of 35.8 J/cm(2) with L-ICG and 71.6 J/cm(2) with R-ICG with minimal damage to the neurosensory retina.
Restrictions to the use of ICG in aqueous solution, low singlet oxygen quantum yields and high aggregation tendency, were overcome with L-ICG. The lower laser irradiance required to obtain choriocapillaris occlusion may suggest that L-ICG is a more potent and selective photosensitiser than R-ICG.
The British journal of ophthalmology 03/2008; 92(2):276-80. · 2.92 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to compare the morphological and visual acuity outcomes associated with a single intravitreal injection of triamcinolone acetonide versus bevacizumab for the treatment of refractory diffuse diabetic macular oedema.
Twenty-eight patients were randomly assigned to receive a single intravitreal injection of either 4 mg/0.1 ml triamcinolone acetonide or 1.5 mg/0.06 ml bevacizumab. Comprehensive ophthalmic evaluation was performed at baseline and at weeks 1, 4, 8 (+/-1), 12 (+/-2) and 24 (+/-2) after treatment. Main outcome measures included central macular thickness measured with optical coherence tomography (OCT) and best corrected Early Treatment Diabetic Retinopathy Study (ETDRS) visual acuity.
Twenty-six patients (26 eyes) completed all study visits (two patients missed two consecutive study visits). Central macular thickness was significantly reduced in the intravitreal triamcinolone group compared with the bevacizumab group at weeks 4, 8, 12 and 24 (p<0.05). Logarithm of the minimum angle of resolution (LogMAR) best-corrected visual acuity was significantly higher at weeks 8 (0.69; approximately 20/100(+1)) and 12 (0.74; 20/100(-2)) in the intravitreal triamcinolone group compared with the bevacizumab group (weeks 8 (0.83; approximately 20/125(-1)) and 12 (0.86; 20/160(+2))) (p<0.05). Significant change from baseline in mean intraocular pressure (mmHg) was seen at week 4 (+2.25) only in the intravitreal triamcinolone group (p<0.0001). No patient had observed cataract progression during the study.
One single intravitreal injection of triamcinolone may offer certain advantages over bevacizumab in the short-term management of refractory diabetic macular oedema, specifically with regard to changes in central macular thickness. The actual clinical relevance of our preliminary findings, however, remains to be determined in future larger studies.
The British journal of ophthalmology 02/2008; 92(1):76-80. · 2.92 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A atividade fotodinâmica do cloro(5,10,15,20-tetrafenilporfirinato) de índio(III) (InTPP) in vitro foi investigado para possível uso em terapia fotodinâmica (PDT). O rendimento quântico de oxigênio singlete do InTPP (Φ ∆ = 0,72) em DMSO foi maior que da 5,10,15,20-tetrafenilporfirina (TPP) (Φ ∆ = 0,52). Os sítios de ligação entre os fotossensibilizadores e albumina bovina (BSA) são independentes e com células vermelhas de sangue humano (RBC) são cooperativos, com um e quatro sítios de ligação por molécula, respectivamente. As constantes de associação com BSA são (1,15 ± 0,07) × 10 5 e (2,6 ± 0,1) × 10 4 L mol -1 e com RBC são (2,40 ± 0,05) × 10 7 L mol -1 e (7,2 ± 0,2) × 10 4 L mol -1 para InTPP e Photofrin®, respectivamente. O InTPP foi mais eficiente do que Photofrin® em fotooxidar L-triptofano (Trp) e BSA quando maiores concentrações dos fotossensibilizadores foram utilizadas (acima de 14 µmol L −1). O InTPP foi 1,37 a 1,5 vezes mais eficaz em fotooxidar as RBC do que Photofrin®. Nossos resultados indicam que o InTPP pode ser usado para estudos futuros de PDT. Photodynamic activity of chloro(5,10,15,20-tetraphenylporphyrinato)indium(III) (InTPP) in vitro was investigated for possible use in photodynamic therapy (PDT). The quantum yield of singlet oxygen generation in DMSO of InTPP (Φ ∆ = 0.72) was higher than 5,10,15,20-tetraphenylporphyrin (TPP) (Φ ∆ = 0.52). Binding sites between photosensitizers and bovine serum albumin (BSA) are independent while binding sites with human red blood cells (RBC) are cooperatives, with one and four binding sites per molecule, respectively. Binding constants with BSA are (1.15 ± 0.07) × 10 5 and (2.6 ± 0.1) × 10 4 L mol -1 and with RBC are (2.40 ± 0.05) × 10 7 L mol -1 e (7.2 ± 0.2) × 10 4 L mol -1 for InTPP and Photofrin®, respectively. InTPP was more efficient than Photofrin® in the photooxidation of L-tryptophan(Trp) and BSA when higher concentrations (14 µmol L −1) of photosensitizers were used. InTPP was 1.37-1.5 times more effective in the photooxidation of RBC than Photofrin®. Our results indicate that InTPP should be used in future studies of PDT.
[Show abstract][Hide abstract] ABSTRACT: Photodynamic activity of chloro(5,10,15,20-tetraphenylporphyrinato)indium(III) (InTPP) in vitro was investigated for possible use in photodynamic therapy (PDT). The quantum yield of singlet oxygen generation in DMSO of InTPP (FD = 0.72) was higher than 5,10,15,20-tetraphenylporphyrin (TPP) (FD = 0.52). Binding sites between photosensitizers and bovine serum albumin (BSA) are independent while binding sites with human red blood cells (RBC) are cooperatives, with one and four binding sites per molecule, respectively. Binding constants with BSA are (1.15 ± 0.07) × 105 and (2.6 ± 0.1) × 104 L mol-1 and with RBC are (2.40 ± 0.05) × 107 L mol-1 e (7.2 ± 0.2) × 104 L mol-1 for InTPP and Photofrin®, respectively. InTPP was more efficient than Photofrin® in the photooxidation of L-tryptophan(Trp) and BSA when higher concentrations (14 µmol L1) of photosensitizers were used. InTPP was 1.37-1.5 times more effective in the photooxidation of RBC than Photofrin®. Our results indicate that InTPP should be used in future studies of PDT.
Journal of the Brazilian Chemical Society 01/2008; 19(3). · 1.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the molecular mechanisms of Al toxicity, cross-species cDNA array approach was employed to identify expressed
sequence tags (ESTs) regulated by Al stress in root tips of Al-tolerant maize (Zea mays) genotype Cat100-6 and Al-sensitive genotype S1587-17. Due to the high degree of conservation observed between sugarcane
and maize, we have analyzed the expression profiling of maize genes using 2 304 sugarcane (ESTs) obtained from different libraries.
We have identified 85 ESTs in Al stressed maize root tips with significantly altered expression. Among the up-regulated ESTs,
we have found genes encoding previously identified proteins induced by Al stress, such as phenyl ammonia-lyase, chitinase,
Bowman-Birk proteinase inhibitor, and wali7. In addition, several novel genes up-and downregulated by Al stress were identified
in both genotypes.
Biologia Plantarum 01/2008; 52(3):475-485. · 1.69 Impact Factor