Interaction of serum amyloid A with human cystatin C—assessment of amino acid residues crucial for hCC–SAA formation (part II)

Secondary amyloid A (AA) amyloidosis is an important complication of some chronic inflammatory diseases, primarily rheumatoid arthritis (RA). It is a serious, potentially life‐threatening disorder caused by the deposition of AA fibrils, which are derived from the circulatory, acute‐phase‐reactant, serum amyloid A protein (SAA). Recently, a specific interaction between SAA and the ubiquitous inhibitor of cysteine proteases—human cystatin C (hCC)—has been proved. Using a combination of selective proteolytic excision and high‐resolution mass spectrometry, the binding sites in the SAA and hCC sequences were assessed as SAA(86–104) and hCC(96–102), respectively. Here, we report further details concerning the hCC–SAA interaction. With the use of affinity tests and florescent ELISA‐like assays, the amino acid residues crucial for the protein interaction were determined. It was shown that all amino acid residues in the SAA sequence, essential for the formation of the protein complex, are basic ones, which suggests an electrostatic interaction character. The idea is corroborated by the fact that the most important residues in the hCC sequence are Ser‐98 and Tyr‐102; these residues are able to form hydrogen bonds via their hydroxyl groups. The molecular details of hCC–SAA complex formation might be helpful for the design of new compounds modulating the biological role of both proteins. Copyright © 2013 John Wiley & Sons, Ltd.     

Amniotic membrane-derived cells inhibit proliferation of cancer cell lines by inducing cell cycle arrest

Cells derived from the amniotic foetal membrane of human term placenta have drawn particular attention mainly for their plasticity and immunological properties, which render them interesting for stem-cell research and cell-based therapeutic applications. In particular, we have previously demonstrated that amniotic mesenchymal tissue cells (AMTC) inhibit lymphocyte proliferation in vitro and suppress the generation and maturation of monocyte-derived dendritic cells. Here, we show that AMTC also significantly reduce the proliferation of cancer cell lines of haematopoietic and non-haematopoietic origin, in both cell-cell contact and transwell co-cultures, therefore suggesting the involvement of yet-unknown inhibitory soluble factor(s) in this 'cell growth restraint'. Importantly, we provide evidence that the anti-proliferative effect of AMTC is associated with induction of cell cycle arrest in G0/G1 phase. Gene expression analyses demonstrate that AMTC can down-regulate cancer cells' mRNA expression of genes associated with cell cycle progression, such as cyclins (cyclin D2, cyclin E1, cyclin H) and cyclin-dependent kinase (CDK4, CDK6 and CDK2), whilst they up-regulate cell cycle negative regulator such as p15 and p21, consistent with a block in G0/G1 phase with no progression to S phase. Taken together, these findings warrant further studies to investigate the applicability of these cells for controlling cancer cell proliferation in vivo.     

A new methodology combining PCR, cloning, and sequencing of clones discriminated by RFLP for the study of microbial populations: application to an UASB reactor sample

This work describes a methodology combining DNA extraction, polymerase chain reaction amplification with primers targeting 16S ribosomal RNA genes, cloning, and sequencing of clones previously analyzed by restriction fragment length polymorphism (RFLP), which can be applied to study the microbial diversity in a given habitat. The methodology allows the minimization of the sequencing effort, which is particularly relevant when analyzing large numbers of clones. The methodology does not require particularly skilled personnel and can easily be adaptable to the molecular characterization of virtually any particular microbial population, provided that both adequate primers and suitable restriction enzymes for RFLP analysis of the clone library have been chosen. An example of application is presented, in which a sample taken from a continuously operating upflow anaerobic sludge blanket reactor was analyzed. RFLP analysis of the initial 162 clones with HaeIII allowed the identification of only 28 distinct profiles. As expected, identical RFLP profiles corresponded to identical nucleotide sequences.     

The importance of ‘tooglies’ to the ethnography of F.E. Williams,government anthropologist

The theoretical orientation, encapsulated in the semi‐serious concept of ‘tooglies'of F.E. Williams, government anthropologist for the Territory of Papua in the inter‐war period, is considered. His ideas about culture change are contrasted with those of Bronislaw Malinowski, who acted as his mentor at one time. His treatment of the social organisational anomaly of Sogeri Koiari ‘sex affiliation’, often cited as a case of parallel bilineal descent, is compared with Margaret Mead's analysis of the ‘Mundugumor ropes’, which is classed as the opposite, cross‐sex bilineal descent. It is shown that Williams was able to get a clearer insight into this anomalous data than Mead and Fortune, and, on the whole, worked with an understanding of culture and Papuan social organisation that presages the relational approach of today.     

Diversification and biogeographic history of the Western Palearctic freshwater flatworm genus Schmidtea (Tricladida: Dugesiidae), with a redescription of Schmidtea nova

The freshwater flatworm genus Schmidtea is endemic in the Western Palearctic region, where it is represented by only four species, thus contrasting with the high species diversity of the closely related genus Dugesia within Europe. Although containing an important model species in developmental and regeneration research, viz. Schmidtea mediterranea, no evolutionary studies on the genus Schmidtea have been undertaken. For the first time, we present a well‐resolved molecular phylogenetic tree of the four species of the genus, inferred on the basis of two molecular markers, and provide also the first detailed morphological account of Schmidtea nova. The phylogenetic tree generated corroborates an earlier speciation hypothesis based on karyological data and points to chromosomal rearrangements as the main drivers of speciation in this genus. The high genetic divergence between the four species, in combination with previous dating studies and their current geographic distribution, suggests that Schmidtea could have originated in Laurasia but lost most of its diversity during the Oligocene. Thus, its present distribution pattern may be the result of the expansion of three of its four relictual species over Europe, probably after the Pleistocene glaciations. Our detailed morphological study of S. nova revealed that it shows a number of remarkable features: interconnected testis follicles, parovaria, an ejaculatory duct exiting into the primary as well as the secondary seminal vesicle by means of a nipple, and the wall of the distal section of the ejaculatory duct being sclerotic or chitinized.