Identification of Medieval Human Soft Tissue Remains in an Advanced State of Decomposition

Naturally preserved human soft tissue remains from mediaeval burials (ll-13th century A. D.) were investigated histologically after azocarmine/aniline alcohol (AZAN) or keratin-prekeratin-mucin (KPM) staining. The tissue remnants were in an advanced state of decomposition; they were completely collapsed and had lost their macroscopic characteristics. After rehydration, thin sectioning, and staining, microscopic properties permitted tissue identification, although differential staining of tissue components did not necessarily correspond with the expected results based on fresh tissue. The techniques and results presented in this paper are relevant for both anthropological and forensic purposes.     

Nephronophthisis-Associated CEP164 Regulates Cell Cycle Progression,Apoptosis and Epithelial-to-Mesenchymal Transition

We recently reported that centrosomal protein 164 (CEP164) regulates both cilia and the DNA damage response in the autosomal recessive polycystic kidney disease nephronophthisis. Here we examine the functional role of CEP164 in nephronophthisis-related ciliopathies and concomitant fibrosis. Live cell imaging of RPE-FUCCI (fluorescent, ubiquitination-based cell cycle indicator) cells after siRNA knockdown of CEP164 revealed an overall quicker cell cycle than control cells, although early S-phase was significantly longer. Follow-up FACS experiments with renal IMCD3 cells confirm that Cep164 siRNA knockdown promotes cells to accumulate in S-phase. We demonstrate that this effect can be rescued by human wild-type CEP164, but not disease-associated mutants. siRNA of CEP164 revealed a proliferation defect over time, as measured by CyQuant assays. The discrepancy between accelerated cell cycle and inhibited overall proliferation could be explained by induction of apoptosis and epithelial-to-mesenchymal transition. Reduction of CEP164 levels induces apoptosis in immunofluorescence, FACS and RT-QPCR experiments. Furthermore, knockdown of Cep164 or overexpression of dominant negative mutant allele CEP164 Q525X induces epithelial-to-mesenchymal transition, and concomitant upregulation of genes associated with fibrosis. Zebrafish injected with cep164 morpholinos likewise manifest developmental abnormalities, impaired DNA damage signaling, apoptosis and a pro-fibrotic response in vivo. This study reveals a novel role for CEP164 in the pathogenesis of nephronophthisis, in which mutations cause ciliary defects coupled with DNA damage induced replicative stress, cell death, and epithelial-to-mesenchymal transition, and suggests that these events drive the characteristic fibrosis observed in nephronophthisis kidneys.     

Structure Elucidation,Antimicrobial and Cytotoxic Activities of a Halimane Isolated from Vellozia kolbekii Alves (Velloziaceae)

A new halimane diterpene was isolated from Vellozia kolbekii Alves (Velloziaceae) and identified as (5R,8R,9S,13R)‐halim‐1,10‐ene‐15,16‐diol ( 1 ). It showed cytotoxicity against three human cancer cell lines, SF‐295 (glioblastoma), MDA‐MB‐435 (melanoma), and HCT‐8 (colon adenocarcinoma). In the mechanism of cytotoxic action, halimane 1 interferes in two major phases of the cell cycle: in S phase, in which DNA synthesis occurs and where it is very sensitive to damage, and G2M phase which is the phase of preparation for mitosis and mitosis itself, showing apoptosis‐inducing properties. Antimicrobial activity towards Gram‐positive and Gram‐negative bacteria was studied and, against Bacillus cereus, B. subtilis, Escherichia coli, and Pseudomonas aeruginosa, a MIC value of 0.025 μM was observed for halimane 1 , which is more active than the positive control chloramphenicol.     

Mycorrhizal response of <Emphasis Type="Italic">Clusia pusilla</Emphasis> growing in two different soils in the field

Arbuscular mycorrhizas (AM) are important for promoting the mineral nutrition, growth and survival of plants used to rehabilitate degraded areas. Clusia pusilla is an evergreen shrub which is tolerant of high irradiance, germinates readily and can be easily reproduced by cuttings. All these characteristics make this species useful in the recovery of deforested areas. The aim of this work was to explore the response of C. pusilla to AM in the field, in two types of soil: the shrubland soil in which the species naturally grows and in a soil of a riparian forest. Eight treatments were performed in each type of soil. The treatments consisted of a non-mycorrhizal control and mycorrhizal plants colonized by one of the three AM inocula tested in the presence or absence of triple superphosphate (150 kg ha^-1). After 11 months of growth in the shrubland soil, C. pusilla seedlings showed an increase in height and dry weight in response to the fertilizer but not to mycorrhizas. In contrast, in the forest soil the arbuscular mycorrhizal fungi (AMF) effect was equivalent to the fertilizer effect, and the two effects interacted positively. The lack of response to AM in shrubland soil was caused by its high sand content, which hinders the retention of the inocula. Due to a higher clay content, the forest soil binds inocula more tightly than shrubland soil. In conclusion, C. pusilla appears to benefit greatly from the addition of AMF in forest soil, though it requires an additional P source for such benefits in shrubland soil. This P source must be organic so that phosphorus is not lost by leaching. Although the growth rate of this species is very low, its survival can be guaranteed with the application of AMF inocula together with P-fertilizer applied at a low rate.     

Glutathione S transferase mu polymorphism and gastric cancer in the Portuguese population

The glutathione S-transferases appear to form part of a protective mechanism against the development of cancer where environmental chemical carcinogens are involved. In humans one member of the mu class gene family (GSTM1) has been shown to be polymorphic and is only expressed in ~50% of individuals. Previous studies have shown a possible link between the null phenotype and susceptibility to cancer but have been equivocal regarding stomach cancer. To evaluate any association in Portuguese gastric cancer individuals with GSTM1 variability, we performed GST M 1 polymorphism by PCR amplification in 148 gastric cancer patients and in 84 healthy control individuals. We found no statistical differences between the gastric cancer and control populations (wild type phenotype: 52%, 48%; null phenotype: 48%, 52%, respectively). A subset analysis into site of tumour also revealed no significant differences between the groups, although we found a slight increase of the wild type phenotype in the samples of the antrum compared with the control population (57% vs 48%, respectively; 2= 1.18; p 0.28) and a slight increase of the null phenotype in the signet ring cells/mucocellular group ( 2= 1.05; p 0.3). However, in both cases it did not reach statistical significance. A subset analysis of the histological groups following the WHO criteria revealed a statistically significant difference ( 2= 3.704; p 0.05) between the moderately differentiated gastric adenocarcinoma and the presence of the wild type phenotype. These results do not support the hypothesis that the GSTM1 null phenotype predisposes to gastric cancer in the Portuguese population and the moderately differentiated gastric adenocarcinoma seems to be associated with the presence of the G STM 1 wild type phenotype.