Improved clonality assessment in germinal centre/post-germinal centre non-Hodgkin's lymphomas with high rates of somatic hypermutation.

BACKGROUND: PCR detects clonal rearrangements of the Ig gene in lymphoproliferative disorders. False negativity occurs in germinal centre/post-germinal centre lymphomas (GC/PGCLs) as they display a high rate of somatic hypermutation (SHM), which causes primer mismatching when detecting Ig rearrangements by PCR. AIMS: To investigate the degree of SHM in a group of GC/PGCLs and assess the rate of false negativity when using BIOMED-2 PCR when compared with previously published strategies. METHODS: DNA was isolated from snap-frozen tissue from 49 patients with GC/PGCL (23 diffuse large B cell lymphomas (DLBCLs), 26 follicular lymphomas (FLs)) and PCR-amplified for complete (VDJH), incomplete (DJH) and Ig kappa/lambda rearrangements using the BIOMED-2 protocols, and compared with previously published methods using consensus primers. Germinal centre phenotype was defined by immunohistochemistry based on CD10, Bcl-6 and MUM-1. RESULTS: Clonality detection by amplifying Ig rearrangements using BIOMED-2 family-specific primers was considerably higher than that found using consensus primers (74% DLBCL and 96% FL vs 69% DLBCL and 73% FL). Addition of BIOMED-2 DJH rearrangements increased detection of clonality by 22% in DLBCL. SHM was present in VDJH rearrangements from all patients with DLBCL (median (range) 5.7% (2.5-13.5)) and FL (median (range) 5.3% (2.3-11.9)) with a clonal rearrangement. CONCLUSIONS: Use of BIOMED-2 primers has significantly reduced the false negative rate associated with GC/PGCL when compared with consensus primers, and the inclusion of DJH rearrangements represents a potential complementary target for clonality assessment, as SHM is thought not to occur in these types of rearrangements.

Spectroscopic and structural analysis of somatic and N-domain angiotensin I-converting enzyme isoforms from mesangial cells from Wistar and spontaneously hypertensive rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Phylum-wide transcriptome analysis of oogenesis and early embryogenesis in selected nematode species

Oogenesis is a prerequisite for embryogenesis in Metazoa. During both biological processes important decisions must be made to form the embryo and hence ensure the next generation: (1) Maternal gene products (mRNAs, proteins and nutrients) must be supplied to the embryo. (2) Polarity must be established and axes must be specified. While incorporation of maternal gene products occurs during oogenesis, the time point of polarity establishment and axis specification varies among species, as it is accomplished either prior, during, or after fertilisation. But not only the time point when these events take place varies among species but also the underlying mechanisms by which they are triggered. For the nematode model Caenorhabditis elegans the underlying pathways and gene regulatory networks (GRNs) are well understood. It is known that there the sperm entry point initiates a primary polarity in the 1-celled egg and with it the establishment of the anteroposterior axis. However, studies of other nematodes demonstrated that polarity establishment can be independent of sperm entry (Goldstein et al., 1998; Lahl et al., 2006) and that cleavage patterns, symmetry formation and cell specification also differ from C. elegans. In contrast to the studied Chromadorea (more derived nematodes including C. elegans), embryos of some marine Enoplea (more basal representatives) even show no discernible early polarity and blastomeres can adopt variable cell fates (Voronov and Panchin 1998). The underlying pathways controlling the obviously variant embryonic processes in non-Caenorhabditis nematodes are essentially unknown. In this thesis I addressed this issue by performing a detailed unbiased comparative transcriptome analysis based on microarrays and RNA sequencing of selected developmental stages in a variety of nematodes from different phylogenetic branches with C. elegans as a reference system and a nematomorph as an outgroup representative. In addition, I made use of available genomic data to determine the presence or absence of genes for which no expression had been detected. In particular, I focussed on components of selected pathways or GRNs which are known to play essential roles during C. elegans development and/or other invertebrate or vertebrate model systems. Oogenesis must be regulated differently in non-Caenorhabditis nematodes, as crucial controlling components of Wnt and sex determination signaling are absent in these species. In this respect, I identified female-specific expression of potential polarity associated genes during gonad development and oogenesis in the Enoplean nematode Romanomermis culicivorax. I could show that known downstream components of the polarity complexes PAR-3/-6/PKC-3 and PAR-1/-2 are absent in non-Caenorhabditis species. Even PAR-2 as part of the polarity complex does not exist in these nematodes. Instead, transcriptomes of nematodes (including C. elegans), show expression of other polarity-associated complexes such as the Lgl (Lethal giant larvae) complex. This result could pose an alternative route for nematodes and nematomorphs to initiate polarity during early embryogenesis. I could show that crucial pathways of axis specification, such as Wnt and BMP are very different in C. elegans compared to other nematodes. In the former, Wnt signaling, for instance, is mediated by four paralogous beta-catenins, while other Chromadorea have fewer and Enoplea only one beta-catenin. The transcriptomes of R. culicivorax and the nematomorph show that regulators of BMP (e.g. Chordin), are specifically expressed during early embryogenesis only in Enoplea and the close outgroup of nematomorphs. In conclusion, my results demonstrate that the molecular machinery controlling oogenesis and embryogenesis in nematodes is unexpectedly variable and C. elegans cannot be taken as a general model for nematode development. Under this perspective, Enoplean nematodes show more similarities with outgroups than with C. elegans. It appears that certain pathway components were lost or gained during evolution and others adopted new functions. Based on my findings I can conjecture, which pathway components may be ancestral and which were newly acquired in the course of nematode evolution.

Cryopreservation of avocado (Persea Americana Mill.) using somatic embryos

Avocado genetic resources are currently maintained in the form of field repositories at great cost and risk of natural disasters, pest and diseases. Cryopreservation offers a necessary, complimentary method that is safe, cost-effective and long-term. However, long-term maintenance and regeneration of plantlets from avocado somatic embryos has been a major barrier in the development of new avocado cultivars. In this study, two protocols for vitrification-based cryopreservation of avocado somatic embryos were investigated. Globular somatic embryos of two avocado cultivars were tested, revealing cultivar-dependent differences in desiccation tolerance and subsequent freezing resistance, possibly attributed to their size and culture age. A two-step regeneration system, involving an intermediate liquid phase step between subcultures in solid medium, significantly enhanced shoot development from somatic embryo tissue. This work will add considerable value towards cryopreservation of avocado somatic embryos for germplasm conservation and the generation of new and improved avocado cultivars.

Biofloc contribution to antioxidant defence status, lipid nutrition and reproductive performance of broodstock of the shrimp Litopenaeus stylirostris: Consequences for the quality of eggs and larvae

The aim of this study was to determine biofloc contributions to the antioxidant status and lipid nutrition of broodstock of Litopenaeus stylirostris in relationship with their reproductive performance and the health of larvae produced. Shrimp broodstock reared with Biofloc technology (BFT) compared to Clear water (CW) exhibited a higher health status with (i) a better final survival rate during the reproduction period (52.6% in CW against 79.8% in BFT); (ii) higher glutathione level (GSH) and total antioxidant status (TAS), reduced oxidized/reduced glutathione ratio and a higher spawning rate and frequency as well as higher gonado-somatic index and number of spawned eggs. Finally, larvae from broodstock from BFT exhibited higher survival rates at the Zoe 2 (+ 37%) and Post Larvae 1 (+ 51%) stages when compared with those from females from CW treatment. The improved reproductive performance of the broodstock and higher larvae survival rate resulting from BFT treatment may be linked to the dietary supplement obtained by the shrimp from natural productivity during BFT rearing. Indeed, our study confirms that biofloc particulates represent a potential source of dietary glutathione and a significant source of lipids, particularly essential phospholipids and n-3 highly unsaturated fatty acids (HUFA) for shrimps. Thus, broodstock from BFT treatment accumulated phospholipids, n-3 HUFA and arachidonic acid, which are necessary for vitellogenesis, embryogenesis and pre-feeding larval development. The predominant essential fatty acids, arachidonic acid (ARA), eicopentaeonic acid (EPA) and docosahexaenoic acid (DHA), had levels in the eggs that were, respectively, 2.5, 2.8 and 3 fold higher for BFT compared to the CW treatment. Statement of Relevance Today, the influence of biofloc technology on shrimp broodstock is not enough described and no information was available on the larvae quality. Moreover, two key pieces of new information emerge from the present study. Firstly, biofloc is a source of further dietary lipids that can act as energetic substrates, but also as a source of phospholipids and essential fatty acids necessary to sustain reproduction, embryonic and larval development. Second, improving the reproduction of the broodstock also leads to an improvement in the quality of the larvae. We think that our research is new and important to increase knowledge on biofloc topic. We believe the paper will contribute to the development of more efficient and therefore more sustainable systems.

Data supporting the co-expression of PDHA1 gene and of its paralogue PDHA2 in somatic cells of a family

This article presents a dataset proving the simultaneous presence of a 5′UTR-truncated PDHA1 mRNA and a full-length PDHA2 mRNA in the somatic cells of a PDC-deficient female patient and all members of her immediate family (parents and brother). We have designed a large set of primer pairs in order to perform detailed RT-PCR assays allowing the clear identification of both PDHA1 and PDHA2 mRNA species in somatic cells. In addition, two different experimental approaches were used to elucidate the copy number of PDHA1 gene in the patient and her mother. The interpretation and discussion of these data, along with further extensive experiments concerning the origin of this altered gene expression and its potential therapeutic consequences, can be found in “Complex genetic findings in a female patient with pyruvate dehydrogenase complex deficiency: null mutations in the PDHX gene associated with unusual expression of the testis-specific PDHA2 gene in her somatic cells” (A. Pinheiro, M.J. Silva, C. Florindo, et al., 2016).

Cryopreservation of somatic embryos for avocado germplasm conservation

Presently avocado germplasm is conserved ex situ in the form of field repositories across the globe including Australia. The maintenance of germplasm in the field is costly, labour and land intensive, exposed to natural disasters and always at the risk of abiotic and biotic stresses. The aim of this study was to overcome these problems using cryopreservation to store avocado (Persea americana Mill.) somatic embryos (SE). Two vitrification-based methods of cryopreservation were optimised (cryovial and droplet-vitrification) using four avocado cultivars (‘A10′, ‘Reed’, ‘Velvick’ and ‘Duke-7′). SE of the four cultivars were stored for short-term (one hour) in liquid nitrogen using the cryovial-vitrification method and showed a viability of 91%, 73%, 86% and 80% respectively. While when using the droplet vitrification method viabilities of 100%, 85% and 93% were recorded for ‘A10′, ‘Reed’ and ‘Velvick’. For long-term storage, SE of cultivars ‘A10′, ‘Reed’ and ‘Velvick’ were successfully recovered with viability of 65–100% after 3 months of LN storage. For cultivar ‘Reed’ and ‘Velvick’ SE were recovered after 12 months of LN storage with viability of 67% and 59%, respectively. The outcome of this work contributes towards the establishment of a cryopreservation protocol that is applicable across multiple avocado cultivars.

Cow, farm, and herd management factors in the dry period associated with raised somatic cell counts in early lactation

This study investigated cow characteristics, farm facilities, and herd management strategies during the dry period to examine their joint influence on somatic cell counts (SCC) in early lactation. Data from 52 commercial dairy farms throughout England and Wales were collected over a 2-yr period. For the purpose of analysis, cows were separated into those housed for the dry period (6,419 cow-dry periods) and those at pasture (7,425 cow-dry periods). Bayesian multilevel models were specified with 2 response variables: ln SCC (continuous) and SCC >199,000 cells/mL (binary), both within 30 d of calving. Cow factors associated with an increased SCC after calving were parity, an SCC >199,000 cells/mL in the 60 d before drying off, increasing milk yield 0 to 30 d before drying off, and reduced DIM after calving at the time of SCC estimation. Herd management factors associated with an increased SCC after calving included procedures at drying off, aspects of bedding management, stocking density, and method of pasture grazing. Posterior predictions were used for model assessment, and these indicated that model fit was generally good. The research demonstrated that specific dry-period management strategies have an important influence on SCC in early lactation.

Growth And Body Composition In Brazilian Female Rhythmic Gymnastics Athletes.

The aim was to analyse the physical growth and body composition of rhythmic gymnastics athletes relative to their level of somatic maturation. This was a cross-sectional study of 136 athletes on 23 teams from Brazil. Mass, standing height and sitting height were measured. Fat-free and fat masses, body fat percentages and ages of the predicted peak height velocity (PHV) were calculated. The z scores for mass were negative during all ages according to both WHO and Brazilian references, and that for standing height were also negative for all ages according to WHO reference but only until 12 years old according to Brazilian reference. The mean age of the predicted PHV was 12.1 years. The mean mass, standing and sitting heights, body fat percentage, fat-free mass and fat mass increased significantly until 4 to 5 years after the age of the PHV. Menarche was reached in only 26% of these athletes and mean age was 13.2 years. The mass was below the national reference standards, and the standing height was below only for the international reference, but they also had late recovery of mass and standing height during puberty. In conclusion, these athletes had a potential to gain mass and standing height several years after PHV, indicating late maturation.

Genomic Copy Number Alterations Of Primary And Secondary Metastasizing Pleomorphic Adenomas.

Metastasizing pleomorphic adenoma (MPA) is a rare tumour, and its mechanism of metastasis still is unknown. To date, there has been no study on MPA genomics. We analysed primary and secondary MPAs with array comparative genomic hybridization to identify somatic copy number alterations and affected genes. Tumour DNA samples from primary (parotid salivary gland) and secondary (scalp skin) MPAs were subjected to array comparative genomic hybridization investigation, and the data were analysed with NEXUS COPY NUMBER DISCOVERY. The primary MPA showed copy number losses affecting 3p22.2p14.3 and 19p13.3p123, and a complex pattern of four different deletions at chromosome 6. The 3p deletion encompassed several genes: CTNNB1, SETD2, BAP1, and PBRM1, among others. The secondary MPA showed a genomic profile similar to that of the primary MPA, with acquisition of additional copy number changes affecting 9p24.3p13.1 (loss), 19q11q13.43 (gain), and 22q11.1q13.33 (gain). Our findings indicated a clonal origin of the secondary MPA, as both tumours shared a common profile of genomic copy number alterations. Furthermore, we were able to detect in the primary tumour a specific pattern of copy number alterations that could explain the metastasizing characteristic, whereas the secondary MPA showed a more unbalanced genome.

Avaliação do crescimento físico e maturação somática de escolares da rede de ensino fundamental de Campinas, SP

Universidade Estadual de Campinas . Faculdade de Educação Física

Survey regional sobre crescimento e desenvolvimento : relações com maturação, sobrepeso e obesidade, nível socioeconômico e desempenho motor

Universidade Estadual de Campinas . Faculdade de Educação Física

"Crescimento humano, composição corporal e desenvolvimento das caracteristicas sexuais secundarias de adolescentes de 11 a 14 anos"

Universidade Estadual de Campinas . Faculdade de Educação Física