C.S. Teixeira

Caracterização genética de seis plantéis comerciais de tilápia (Oreochromis) utilizando marcadores microssatélites

Two hundred and thirty five individuals from six commercial stocks of tilapias (Ceara, Chitralada, Israel, Nilotica, Taiwan and Red) from the Southeastern region of the country were genetically characterized using five microsatellite loci. The results suggest large genetic difference among the stocks, estimated through the fixation allele index (Fst = 0.3263), and a considerable loss of heterozigosity accurs in most of the stocks, according to the population inbreeding coefficient (Fis=0.0486). The Israel and Nilotica stocks were genetically similar (Ig=0.6663), while Chitralada and Taiwan showed less genes in common (Ig=0.2463). The Red stock was the most distinct stock.

Preparation, physicochemical characterization, and cell viability evaluation of long-circulating and pH-sensitive liposomes containing ursolic acid.

Cancer is one of the leading causes of death worldwide. Although several drugs are used clinically, some tumors either do not respond or are resistant to the existing pharmacotherapy, thus justifying the search for new drugs. Ursolic acid (UA) is a triterpene found in different plant species that has been shown to possess significant antitumor activity. However, UA presents a low solubility in aqueous medium, which presents a barrier to its biological applications. In this context, the use of liposomes presents a promising strategy to deliver UA and allow for its intravenous administration. In this work, long-circulating and pH-sensitive liposomes containing UA (SpHL-UA) were developed, and their chemical and physicochemical properties were evaluated. SpHL-UA presented adequate properties, including a mean diameter of 191.1 ± 6.4 nm, a zeta potential of 1.2 ± 1.4 mV, and a UA entrapment of 0.77 ± 0.01 mg/mL. Moreover, this formulation showed a good stability after having been stored for 2 months at 4°C. The viability studies on breast (MDA-MB-231) and prostate (LNCaP) cancer cell lines demonstrated that SpHL-UA treatment significantly inhibited cancer cell proliferation. Therefore, the results of the present work suggest the applicability of SpHL-UA as a new and promising anticancer formulation.

Development of a bone-targeted pH-sensitive liposomal formulation containing doxorubicin: physicochemical characterization, cytotoxicity, and biodistribution evaluation in a mouse model of bone metastasis

Background Despite recent advances in cancer therapy, the treatment of bone tumors remains a major challenge. A possible underlying hypothesis, limitation, and unmet need may be the inability of therapeutics to penetrate into dense bone mineral, which can lead to poor efficacy and high toxicity, due to drug uptake in healthy organs. The development of nanostructured formulations with high affinity for bone could be an interesting approach to overcome these challenges. Purpose To develop a liposomal formulation with high affinity for hydroxyapatite and the ability to release doxorubicin (DOX) in an acidic environment for future application as a tool for treatment of bone metastases. Materials and methods Liposomes were prepared by thin-film lipid hydration, followed by extrusion and the sulfate gradient-encapsulation method. Liposomes were characterized by average diameter, ζ-potential, encapsulation percentage, X-ray diffraction, and differential scanning calorimetry. Release studies in buffer (pH 7.4 or 5), plasma, and serum, as well as hydroxyapatite-affinity in vitro analysis were performed. Cytotoxicity was evaluated by MTT assay against the MDA-MB-231 cell line, and biodistribution was assessed in bone metastasis-bearing animals. Results Liposomes presented suitable diameter (~170 nm), DOX encapsulation (~2 mg/mL), controlled release, and good plasma and serum stability. The existence of interactions between DOX and the lipid bilayer was proved through differential scanning calorimetry and small-angle X-ray scattering. DOX release was faster when the pH was in the range of a tumor than at physiological pH. The bone-targeted formulation showed a strong affinity for hydroxyapatite. The encapsulation of DOX did not interfere in its intrinsic cytotoxicity against the MDA-MB-231 cell line. Biodistribution studies demonstrated high affinity of this formulation for tumors and reduction of uptake in the heart. Conclusion These results suggest that bone-targeted pH-sensitive liposomes containing DOX can be an interesting strategy for selectively delivering this drug into bone-tumor sites, increasing its activity, and reducing DOX-related toxicity.

Synthesis of a novel series of 2,3,4-trisubstituted oxazolidines designed by isosteric replacement or rigidification of the structure and cytotoxic evaluation

We have previously reported on a study of the structure–activity relationship in a series of 2,3,4-substituted oxazolidines recently discovered by our group varying the substituent at the ring or stereochemistry of the oxazolidine ring. We discovered the cytotoxic and pro-apoptotic potential of compounds 1 and 2 with good selectivity against cancer cell lines. In the present study we describe the synthesis and cytotoxic evaluation against cancer cell lines (HL60, JURKAT, MDA-MB-231 and LNCaP) of a series of oxazolidines designed by isosteric replacement or rigidification of the oxymethylene spacer of compounds 1 and 2. Alkenes 3 and 4 retained the activity against MDA-MB-231 cells and they were more active on HL60, JURKAT and LNCaP cells. Considering LNCaP cells, E-isomer 4 was at least 7 times and about 3 times more potent than lead 1 and Z-isomer 3, respectively. Compound 4 exerted significant activity against LNCaP with IC50 in the low micromolar range (11 μM) without affecting VERO cells and PBMC proliferation (IC50 > 100 μM) indicating its low toxicity to normal cells.

Comparação entre métodos de estocagem de DNA extraído de amostras de sangue, sêmen e pêlos e entre técnicas de extração

DNA samples of six bovines obtained from three tissues (blood, semen and hair) were extracted using two different techniques. After the extraction procedures the samples were divided in six fractions. Three were stored at -20° C and three at 4° C. Every three months one sample of each tissue/extraction procedure was analyzed in spectrophotometer, to determine the quantity of the DNA and the extract was amplified using the primer RM 29. No differences in the DNA quantity or in the level of protein contamination among the three periods of analyses were observed. All the DNA extracted by quick extraction technique showed good amplification patterns during the nine months, meaning that this technique can be used in laboratory routine instead of the permanent extraction technique. The extract obtained from blood, using the permanent extraction technique, showed the higher quantity of DNA with the smaller index of protein contamination. The high quantity of protein contamination found in the semen samples preserved in egg yolk demanded modifications in both extraction techniques. After that the results were positive, showing good amplification patterns.

Prevalência da imunodeficiência severa combinada em cavalos da raça Árabe em plantéis de Minas Gerais e São Paulo

Neste estudo foram genotipados 205 cavalos da raca Arabe, criados nos estados de Minas Gerais e Sao Paulo, via DNA por meio de PCR, para determinacao da presenca do gene mutante SCID. Os resultados mostraram 98,5% de animais normais (202/205) e 1,5% de portadores (3/205). Pela analise da genealogia dos portadores identificados pode-se, ainda, confirmar a participacao de um garanhao anteriormente identificado como provavel disseminador da doenca.

Reduced expression of the murine HLA-G homolog Qa-2 is associated with malignancy, epithelial-mesenchymal transition and stemness in breast cancer cells

Qa-2 is believed to mediate a protective immune response against cancer; however, little is known about the role of Qa-2 in tumorigenesis. Here, we used 4T1 breast cancer cells to study the involvement of Qa-2 in tumor progression in a syngeneic host. Qa-2 expression was reduced during in vivo tumor growth and in cell lines derived from 4T1-induced tumors. Tumor-derived cells elicited an epithelial-mesenchymal transition associated with upregulation of Zeb1 and Twist1/2 and enhanced tumor initiating and invasive capacities. Furthermore, these cells showed increased stem characteristics, as demonstrated by upregulation of Hes1, Sox2 and Oct3/4, and enrichment of CD44high/CD24median/low cells. Remarkably, Qa-2 cell-surface expression was excluded from the CD44high/CD24median/low subpopulation. Tumor-derived cells showed increased Src activity, and treatment of these cells with the Src kinase inhibitor PP2 enhanced Qa-2 but reduced Sox2 and CD44high/CD24median/low expression levels, suggesting that Src signaling, while positively associated with stemness, negatively regulates Qa-2 expression in breast cancer. Finally, overexpression of the Qa-2 family member Q7 on the cell surface slowed down in vivo tumor growth and reduced the metastatic potential of 4T1 cells. These results suggest an anti-malignant role for Qa-2 in breast cancer development, which appears to be absent from cancer stem cells.

Synthesis of novel 2,3,4-trisubstituted-oxazolidine derivatives and in vitro cytotoxic evaluation.

We have previously reported the discovery of cytotoxic and pro-apoptotic hit compound 1,1-dimethylethyl (S)- 2,2-dimethyl-4-[(3-nitrophenoxy)methyl]-3-oxazolidinecarboxylate 1 against leukemia cells. In the present work we describe the synthesis of 25 derivatives of this hit varying the substituent at ring or stereochemistry of the oxazolidine ring and evaluated them against human cancer cells lines. Six compounds exerted significant activity against HL60 promyelocytic leukemia cells with IC50 in low micromolar range (4-18 μM) and three compounds displayed activity against MDA-MB231 breast cancer cells (25-37 μM). In vitro cytotoxicity on normal cells PBMC (human peripheral blood mononuclear cells) was also evaluated. Compounds 7e (p-NO2, S) and 7m (p-COOCH3, S) showed good antiproliferative activity against HL60 (4 and 5 μM) and MDA-MB231 (37 and 25 μM) without affecting lymphocyte proliferation in PBMC, indicating low toxicity to normal cells. Besides, compound 7e induced DNA fragmentation on about 100% of HL60 cells at 50 μM. In this case, it was more potent than 7m and lead 1. This indicated that compound 7e has a great pro-apoptotic potential.

Microssatélites BM2113, ILSTS005, ILSTS008, ETH131 e RM88 em testes de verificação de parentesco para bovinos da raça Gir

Forty six animals of the Gir breed, registered at the Brazilian Association of Zebu Breeders, coming from five farms located in Minas Gerais State, were used to analyze the efficiency of the microsatellites BM2113, ILSTS005, ILSTS008, ETH131 and RM88 in parentage tests. The loci BM2113, ILSTS005, ETH131 and RM88 showed to be efficient, presenting values of PE2 (exclusion probability when both parents are genotiped) between 0.62 and 0.69 and PIC2 (polymorphic information contents when both parents are genotiped) between 0.78 and 0.83. The same was not observed for the locus ILSTS008 that showed low values of PE2 (0.24) and PIC2 (0.41)

Freqüência do gene Miostatina (GDF-8) em rebanhos brasileiros da raça Marchigiana

The frequency of the normal myostatin gene (GDF-8) and the mutant allele in Marchigiana herds was detected. Three hundred and seventy-seven animals of Marchigiana breed raised in Sao Paulo and Parana States, Brazil, were tested. The results showed that 37.9% were homozygous normal animals, 55.2% heterozygous and 6.9% homozygous double muscling. The results suggest the interest of the breeders in having interest in the character double muscling, randomly promoting, the selection in favor of the mutation.