The Clinical Impact and Molecular Biology of del(17p) in Multiple Myeloma Treated with Conventional or Thalidomide-Based Therapy

Hemizygous deletion of 17p (del(17p)) has been identified as a variable associated with poor prognosis in myeloma, although its impact in the context of thalidomide therapy is not well described. The clinical outcome of 85 myeloma patients with del(17p) treated in a clinical trial incorporating both conventional and thalidomide-based induction therapies was examined. The clinical impact of deletion, low expression, and mutation of TP53 was also determined. Patients with del(17p) did not have inferior response rates compared to patients without del(17p), but, despite this, del(17p) was associated with impaired overall survival (OS) (median OS 26.6 vs. 48.5 months, P <0.001). Within the del(17p) group, thalidomide induction therapy was associated with improved response rates compared to conventional therapy, but there was no impact on OS. Thalidomide maintenance was associated with impaired OS, although our analysis suggests that this effect may have been due to confounding variables. A minimally deleted region on 17p13.1 involving 17 genes was identified, of which only TP53 and SAT2 were underexpressed. TP53 was mutated in <1% in patients without del(17p) and in 27% of patients with del(17p). The higher TP53 mutation rate in samples with del(17p) suggests a role for TP53 in these clinical outcomes. In conclusion, del(17p) defined a patient group associated with short survival in myeloma, and although thalidomide induction therapy was associated with improved response rates, it did not impact OS, suggesting that alternative therapeutic strategies are required for this group. (C) 2011 Wiley-Liss, Inc.

Potential pathogenic fungi assessment through molecular biology in cork industry

Portugal has been the world leader in the cork sectr in terms of exports, employing ten thousands of workers. In this working activity, the permanent contact with cork may lead to the exposure to fungi raising concerns as occupational hazards in cork industry. A study was developed aiming at assessing fungal contamination due to Aspergillus fumigatus complex and Penicillium glabrum complex by molecular methods in three cork industries in the outskirt of Lisbon city. The chosen fungal species are the ones most frequently associated with respiratory problems in workers from these industries.

Cholangiocarcinoma: from molecular biology to treatment

Cholangiocarcinoma is a rare tumor originating in the bile ducts, which, according to their anatomical location, is classified as intrahepatic, extrahepatic and hilar. Nevertheless, incidence rates have increased markedly in recent decades. With respect to tumor biology, several genetic alterations correlated with resistance to chemotherapy and radiotherapy have been identified. Here, we highlight changes in KRAS and TP53 genes that are normally associated with a more aggressive phenotype. Also IL-6 and some proteins of the BCL-2 family appear to be involved in the resistance that the cholangiocarcinoma presents toward conventional therapies. With regard to diagnosis, tumor markers most commonly used are CEA and CA 19-9, and although its use isolated appears controversial, their combined value has been increasingly advocated. In imaging terms, various methods are needed, such as abdominal ultrasound, computed tomography and cholangiopancreatography. Regarding therapy, surgical modalities are the only ones that offer chance of cure; however, due to late diagnosis, most patients cannot take advantage of them. Thus, the majority of patients are directed to other therapeutic modalities like chemotherapy, which, in this context, assumes a purely palliative role. Thus, it becomes urgent to investigate new therapeutic options for this highly aggressive type of tumor.

Characterization of Borrelia burgdorferi Gene Product BBD18 for Its Role(s) in Pathogen Biology and Infectivity

bbd18 is a differentially expressed Borrelia burgdorferi gene that is transcribed at almost undetectable levels in spirochetes grown in vitro but dramatically upregulated during tick infection. The gene also displays low yet detectable expression at various times in tissues of murine hosts. As the gene product bears no homology to known proteins, its biological significance remains enigmatic. To understand the gene function, we created isogenic bbd18-deletion mutants as well as genetically-complemented isolates from an infectious wild-type B. burgdorferi strain. Compared to parental isolates, bbd18 mutants - but not complemented spirochetes - displayed slower in vitro growth. The bbd18 mutants also reflect significantly reduced ability to persist or remain undetectable both in immunocompetent and SCID mice, yet were able to survive in ticks. This suggests BBD18 function is essential in mammalian hosts but redundant in the arthropod vector. Notably, although bbd18 expression and in vitro growth defects are restored in the complemented isolates, their phenotype is similar to the mutants - being unable to persist in mice but able to survive in ticks. Despite low expression in cultured wild-type B. burgdorferi, bbd18 deletion downregulated several genes. Interestingly, expression of some, including ospD and bbi39, could be complemented, while that of others could not be restored via bbd18 re-expression. Correspondingly, bbd18 mutants displayed altered production of several proteins, and similar to RNA levels, some were restored in the bbd18 complement and others not. To understand how bbd18 deletion results in apparently permanent and noncomplementable phenotypic defects, we sought to genetically disturb the DNA topology surrounding the bbd18 locus without deleting the gene. Spirochetes with an antibiotic cassette inserted downstream of the gene, between bbd17 and bbd18, were significantly attenuated in mice, while a similar upstream insertion, between bbd18 and bbd19, did not affect infectivity, suggesting that an unidentified cis element downstream of bbd18 may encode a virulence-associated factor critical for infection.

Effect of remote ischaemic preconditioning in cardiac dysfunction and end-organ injury following cardiac surgery with cardiopulmonary bypass in children: A translational approach investigating clinical outcome and myocardial molecular biology

Congenital heart disease (CHD) is the most common birth defect, causing an important rate of morbidity and mortality. Treatment of CHD requires surgical correction in a significant percentage of cases which exposes patients to cardiac and end organ injury. Cardiac surgical procedures often require the utilisation of cardiopulmonary bypass (CPB), a system that replaces heart and lungs function by diverting circulation into an external circuit. The use of CPB can initiate potent inflammatory responses, in addition a proportion of procedures require a period of aortic cross clamp during which the heart is rendered ischaemic and is exposed to injury. High O2 concentrations are used during cardiac procedures and when circulation is re-established to the heart which had adjusted metabolically to ischaemia, further injury is caused in a process known as ischaemic reperfusion injury (IRI). Several strategies are in place in order to protect the heart during surgery, however injury is still caused, having detrimental effects in patients at short and long term. Remote ischaemic preconditioning (RIPC) is a technique proposed as a potential cardioprotective measure. It consists of exposing a remote tissue bed to brief episodes of ischaemia prior to surgery in order to activate protective pathways that would act during CPB, ischaemia and reperfusion. This study aimed to assess RIPC in paediatric patients requiring CHD surgical correction with a translational approach, integrating clinical outcome, marker analysis, cardiac function parameters and molecular mechanisms within the cardiac tissue. A prospective, single blinded, randomized, controlled trial was conducted applying a RIPC protocol to randomised patients through episodes of limb ischaemia on the day before surgery which was repeated right before the surgery started, after anaesthesia induction. Blood samples were obtained before surgery and at three post-operative time points from venous lines, additional pre and post-bypass blood samples were obtained from the right atrium. Myocardial tissue was resected during the ischaemic period of surgery. Echocardiographic images were obtained before the surgery started after anaesthetic induction and the day after surgery, images were stored for later off line analysis. PICU surveillance data was collected including ventilation parameters, inotrope use, standard laboratory analysis and six hourly blood gas analysis. Pre and post-operative quantitation of markers in blood specimens included cardiac troponin I (cTnI) and B-type natriuretic peptide (BNP), inflammatory mediators including interleukins IL-6, IL-8, IL-10, tumour necrosis factor (TNF-α), and the adhesion molecules ICAM-1 and VCAM-1; the renal marker Cystatin C and the cardiovascular markers asymmetric dymethylarginine (ADMA) and symmetric dymethylarginine (SDMA). Nitric oxide (NO) metabolites and cyclic guanosine monophosphate (cGMP) were measured before and after bypass. Myocardial tissue was processed at baseline and after incubation at hyperoxic concentration during four hours in order to mimic surgical conditions. Expression of genes involved in IRI and RIPC pathways was analysed including heat shock proteins (HSPs), toll like receptors (TLRs), transcription factors nuclear factor κ-B (NF- κ-B) and hypoxia inducible factor 1 (HIF-1). The participation of hydrogen sulfide enzymatic genes, apelin and its receptor were explored. There was no significant difference according to group allocation in any of the echocardiographic parameters. There was a tendency for higher cTnI values and inotropic score in control patients post-operatively, however this was not statistically significant. BNP presented no significant difference according to group allocation. Inflammatory parameters tended to be higher in the control group, however only TNF- α was significantly higher. There was no difference in levels of Cystatin C, NO metabolites, cGMP, ADMA or SDMA. RIPC patients required shorter PICU stay, all other clinical and laboratory analysis presented no difference related to the intervention. Gene expression analysis revealed interesting patterns before and after incubation. HSP-60 presented a lower expression at baseline in tissue corresponding to RIPC patients, no other differences were found. This study provided with valuable descriptive information on previously known and newly explored parameters in the study population. Demographic characteristics and the presence of cyanosis before surgery influenced patterns of activity in several parameters, numerous indicators were linked to the degree of injury suffered by the myocardium. RIPC did not reduce markers of cardiac injury or improved echocardiographic parameters and it did not have an effect on end organ function; some effects were seen in inflammatory responses and gene expression analysis. Nevertheless, an important clinical outcome indicator, PICU length of stay was reduced suggesting benefit from the intervention. Larger studies with more statistical power could determine if the tendency of lower injury and inflammatory markers linked to RIPC is real. The present results mostly support findings of larger multicentre trials which have reported no cardiac benefit from RIPC in paediatric cardiac surgery.

Molecular biology of prolactin and prolactin receptor in a marine teleost: the sea bream(Sparus aurata)

Tese de dout. em Biologia, especialidade de Biologia Molecular, Unidade de Ciências e Tecnologias dos Recursos Aquáticos, Univ. do Algarve

The expression of genes coding for distinct types of glycine-rich proteins varies according to the biology of three metastriate ticks, Rhipicephalus (Boophilus) microplus, Rhipicephalus sanguineus and Amblyomma cajennense

Background: Ticks secrete a cement cone composed of many salivary proteins, some of which are rich in the amino acid glycine in order to attach to their hosts' skin. Glycine-rich proteins (GRPs) are a large family of heterogeneous proteins that have different functions and features; noteworthy are their adhesive and tensile characteristics. These properties may be essential for successful attachment of the metastriate ticks to the host and the prolonged feeding necessary for engorgement. In this work, we analyzed Expressed Sequence Tags (ESTs) similar to GRPs from cDNA libraries constructed from salivary glands of adult female ticks representing three hard, metastriate species in order to verify if their expression correlated with biological differences such as the numbers of hosts ticks feed on during their parasitic life cycle, whether one (monoxenous parasite) or two or more (heteroxenous parasite), and the anatomy of their mouthparts, whether short (Brevirostrata) or long (Longirostrata). These ticks were the monoxenous Brevirostrata tick, Rhipicephalus (Boophilus) microplus, a heteroxenous Brevirostrata tick, Rhipicephalus sanguineus, and a heteroxenous Longirostrata tick, Amblyomma cajennense. To further investigate this relationship, we conducted phylogenetic analyses using sequences of GRPs from these ticks as well as from other species of Brevirostrata and Longirostrata ticks. Results: cDNA libraries from salivary glands of the monoxenous tick, R. microplus, contained more contigs of glycine-rich proteins than the two representatives of heteroxenous ticks, R. sanguineus and A. cajennense (33 versus, respectively, 16 and 11). Transcripts of ESTs encoding GRPs were significantly more numerous in the salivary glands of the two Brevirostrata species when compared to the number of transcripts in the Longirostrata tick. The salivary gland libraries from Brevirostrata ticks contained numerous contigs significantly similar to silks of true spiders (17 and 8 in, respectively, R. microplus and R. sanguineus), whereas the Longirostrata tick contained only 4 contigs. The phylogenetic analyses of GRPs from various species of ticks showed that distinct clades encoding proteins with different biochemical properties are represented among species according to their biology. Conclusions: We found that different species of ticks rely on different types and amounts of GRPs in order to attach and feed on their hosts. Metastriate ticks with short mouthparts express more transcripts of GRPs than a tick with long mouthparts and the tick that feeds on a single host during its life cycle contain a greater variety of these proteins than ticks that feed on several hosts.

Wolbachia: Invasion biology in South Pacific butterflies

Wolbachia ensdosymbionts are well known for their ability to manipulate the population biology and development of their hosts. One of the less studied outcomes of Wolbachia infection with this symbiont is the selective killing of male embryos. Recent work on butterflies living on different South Pacific islands is beginning to help us understand the complexity of the co-evolutionary interactions between these partners.

The systematics and biology of the south African gall-inducing scale insect, Calycicoccus merwei brain (Hemiptera : Coccoidea : Eriococcidae)

The scale insect genus Calycicoccus Brain has a single described species, C. merwei Brain, which is endemic to southeastern South Africa. Females of C. merwei induce small, mostly conical galls on the foliage of their host tree, Apodytes dimidiata E. Meyer ex Arn. (Icacinaceae), which has a wider, mostly coastal distribution, than that currently known for the scale insect. Calycicoccus has been placed in the family Eriococcidae and may be related to the South American genus Aculeococcus Lepage. No other native eriococcid species have been described so far in South Africa, although the family is diverse in other Gondwanan regions. This paper summarizes the biology of C. merwei, redescribes the adult female, describes the adult male, the second-instar female and the first-instar nymphs for the first time, and reconsiders the phylogenetic relationships of the genus. The adult female is shown to have unusual abdominal segmentation, in that segment I is present both dorsally and ventrally, but a segment is absent ventrally on the middle abdomen. First-instar nymphs are sexually dimorphic; males have a larger and relatively narrower body, larger mouthparts, longer antennae and legs, and more thoracic dorsal setae compared with females. Molecular data from nuclear small-subunit ribosomal DNA (18S) and elongation factor 1 alpha (EF-1a) show C. merwei to have no close relatives among the Eriococcidae sampled to date. Instead, the Calycicoccus lineage is part of a polytomy near the base of the Eriococcidae. Molecular dating of the node suggests that the Calycicoccus lineage diverged from other eriococcids more than 100 Mya. These data support the placement of Calycicoccus as the only genus in the subfamily Calycicoccinae Brain.