Scanning electron microscopic assessment on surface morphology of preserved human amniotic membrane after gamma sterilisation

Human amniotic membrane that has been processed and sterilised by gamma irradiation is widely used as a biological dressing in surgical applications. The morphological structure of human amniotic membrane was studied under scanning electron microscopy (SEM) to assess effects of gamma radiation on human amniotic membrane following different preservation methods. The amniotic membrane was preserved by either air drying or submerged in glycerol before gamma irradiated at 15, 25 and 35 kGy. Fresh human amniotic membrane, neither preserved nor irradiated was used as the control. The surface morphology of glycerol preserved amnion was found comparable to the fresh amniotic membrane. The cells of the glycerol preserved was beautifully arranged, homogonous in size and tended to round up. The cell structure in the air dried preserved amnion seemed to be flattened and dehydrated. The effects of dehydration on intercellular channels and the microvilli on the cell surface were clearly seen at higher magnifications (10,000×). SEM revealed that the changes of the cell morphology of the glycerol preserved amnion were visible at 35 kGy while the air dried already changed at 25 kGy. Glycerol preservation method is recommended for human amniotic membrane as the cell morphological structure is maintained and radiation doses lower than 25 kGy for sterilization did not affect the appearance of the preserved amnion.     

The microscopic biological response of human chondrocytes to bovine bone scaffold

This study was performed to determine the microscopic biological response of human nasal septum chondrocytes and human knee articular chondrocytes placed on a demineralized bovine bone scaffold. Both chondrocytes were cultured and seeded onto the bovine bone scaffold with seeding density of 1 × 105 cells per 100 μl/scaffold and incubated for 1, 2, 5 and 7 days. Proliferation and viability of the cells were measured by mitochondrial dehydrogenase activity (MTT assay), adhesion study was analyzed by scanning electron microscopy and differentiation study was analyzed by immunofluorescence staining and confocal laser scanning electron microscopy. The results showed good proliferation and viability of both chondrocytes on the scaffolds from day 1 to day 7. Both chondrocytes increased in number with time and readily grew on the surface and into the open pores of the scaffold. Immunofluorescence staining demonstrated collagen type II on the scaffolds for both chondrocytes. The results showed good cells proliferation, attachment and maturity of the chondrocytes on the demineralized bovine bone scaffold. The bovine bone being easily resourced, relatively inexpensive and non toxic has good potential for use as a three dimensional construct in cartilage tissue engineering.     

Fishing through time: population dynamics of plaice (Pleuronectes platessa) in the Kattegat–Skagerrak over a century

Over the past century, advances in technology and historical events such as climate change have resulted in significant changes in the exploitation pattern, population sizes and the potential yield of fish stocks. These variations provide contrast in the data that improves our knowledge on population dynamics and our ability to develop management strategies for long-term sustainable exploitation. In this study, we use a standardized scientific trawl survey to obtain a historical time series (1901–2007) of relative abundance, recruitment and size structure for plaice in the Kattegat–Skagerrak. Our work extends the available time series by more than 80 years so that the evaluation of trends is more informative than is possible from the current assessment. We show that the current adult biomass is approximately 40% of the maximum observed at the beginning of the century and during the 1960s. The average maximum individual length has been reduced by 10 cm over the studied time period. An analysis of trends in mean length indicates that fishing mortality was variable during the first half of the century and has increased steadily over the past 20 years. Recruitment has been the highest on record during recent years, suggesting that the alleged link between coastal environmental degradation and juvenile survival is of low importance. The overall findings of our work will provide managers with a historical perspective on the population dynamics of the stock, which will support the long-term management of plaice in the Kattegat–Skagerrak.     

Enhanced secretory production of hemolysin-mediated cyclodextrin glucanotransferase in Escherichia coli by random mutagenesis of the ABC transporter system

The hemolysin transport system was found to mediate the release of cyclodextrin glucanotransferase (CGTase) into the extracellular medium when it was fused to the C-terminal 61 amino acids of HlyA (HlyAs(61)). To produce an improved-secretion variant, the hly components (hlyAs, hlyB and hlyD) were engineered by directed evolution using error-prone PCR. Hly mutants were screened on solid LB-starch plate for halo zone larger than the parent strain. Through screening of about 1 × 10(4) Escherichia coli BL21(DE3) transformants, we succeeded in isolating five mutants that showed a 35-217% increase in the secretion level of CGTase-HlyAs(61) relative to the wild-type strain. The mutation sites of each mutant were located at HlyB, primarily along the transmembrane domain, implying that the corresponding region was important for the improved secretion of the target protein. In this study we describe the finding of novel site(s) of HlyB responsible for enhancing secretion of CGTase in E. coli.     

Interleukin-27 Induces a STAT1/3- and NF-κB-dependent Proinflammatory Cytokine Profile in Human Monocytes

IL-27 is a heterodimeric cytokine bridging innate and adaptive immunity by playing a role in the activation of naive T cells and in development of Th1 cells. Additionally, recent evidence supports a role for IL-27 in the activation of monocytic cells. Both pro-inflammatory and anti-inflammatory activities have been attributed to IL-27; however, the role played by IL-27 in the activation of human monocytic cells in terms of cytokine production has not been well described. Our results show that IL-27 is a strong inducer of proinflammatory cytokine and chemokine expression, including enhancement of IL-6, IP-10, MIP-1α, MIP-1β, and TNF-α expression in human primary monocytes. Furthermore, we observed that IL-27-induced cytokine and chemokine production was mediated by STAT1, STAT3, and NF-κB activation. Understanding how IL-27 exerts its effects on monocytic cells will identify important molecular mechanisms in the regulation of immune responses, particularly in the modulation of monocyte activation.     

Functional Characterisation of the Oil Palm Type 3 Metallothionein-like Gene (<Emphasis Type="Italic">MT3-B</Emphasis>) Promoter

In silico analysis showed that the differentially expressed type 3 oil palm metallothionein-like genes MT3-A and MT3-B share at least 11 common putative promoter regulatory elements. The identified motifs include W-boxes, TATCCA element, binding element for cytokinin response regulators and pollen-specific elements. A high degree of conservation was observed in their genomic organisation where the coding regions are divided at two identical positions in both genes by two AT-rich introns. Promoter activity of the MT3-B gene was analysed using a transient assay by bombarding oil palm tissue slices with a β-glucuronidase (GUS) gene construct and a stable reporter assay by analysing GUS expression in transformed Arabidopsis thaliana plants. Transient expression analysis revealed MT3-B promoter activity in oil palm root tissues but not in fruit mesocarp at 12 weeks after anthesis and spear leaves. The T3 homozygous transgenic Arabidopsis plants, harbouring the MT3-B promoter/GUS construct, showed reporter activity in cotyledons and mature leaves with lower expression levels in root tissues. The expression levels in the roots of the T3 homozygous transgenic plants increased five- and 2.5-folds when treated with 80 μM of Zn²⁺ and Fe²⁺, respectively. Altogether, these results indicate that the MT3-A and MT3-B promoter activities may be regulated by a variety of abiotic factors and MT3-B promoter may potentially be manipulated for use in plant genetic engineering for induced synthesis of gene product.     

Phylogenetic relationships of species of Crenicichla (Teleostei: Cichlidae) from southern South America based on the mitochondrial cytochrome b gene

Phylogenetic analysis using Bayesian inference, likelihood and parsimony methods was conducted on 60 complete mitochondrial cytochrome b sequences from 21 species of Crenicichla, including all species known from Uruguay (Crenicichla celidochilus, Crenicichla lepidota Crenicichla minuano, Crenicichla missioneira, Crenicichla punctata, Crenicichla scottii, Crenicichla vittata), Crenicichla compressiceps, Crenicichla empheres, Crenicichla geayi, Crenicichla iguassuensis, Crenicichla macrophthalma, Crenicichla menezesi, Crenicichla notophthalmus, Crenicichla regani, Crenicichla cf. regani, Crenicichla semifasciata, Crenicichla sveni, Crenicichla tendybaguassu, two unidentified species, and also two species of Teleocichla. Bayesian analysis resulted in a trichotomy with three major groups: (1) The C. missioneira species group (C. celidochilus, C. empheres, C. minuano, C. missioneira, C. tendybaguassu, and an undescribed species analyzed); (2) a group of southern species (C. iguassuensis, C. punctata, C. scottii, C. vittata); and (3) a rather heterogeneous group comprising the type species C. macrophthalma, members of the Crenicichla reticulata species group (C. geayi, C. semifasciata), members of the Crenicichla wallacii species group (C. compressiceps, C. notophthalmus, C. regani, C. cf. regani), members of the Crenicichla saxatilis species group (C. lepidota, C. menezesi, C. sveni, C. sp.), and two species of Teleocichla. Parsimony jackknifing resulted in a quadritomy with: (1) C. macrophthalma, (2) Teleocichla, (3) the saxatilis + wallacii group species, and (4) the rest, which include C. geayi and C. semifasciata as sister group to a dichotomy with the C. missioneira group and the remaining southern species. The sequence variation within the C. missioneira group is remarkably minor despite considerable morphological differences, supporting the conclusion that it forms an endemic species flock in the Uruguay River basin. Previously proposed species groups within the speciose genus Crenicichla (more than 90 species known) are partly corroborated. However, C. celidochilus was not previously associated with the C. missioneira species group, and C. vittata has not previously been associated with C. scottii, C. iguassuensis, or C. punctata. Crenicichla lepidota, C. sveni, C. menezesi and C. sp. represent the C. saxatilis group. Species of small size, representing the C. wallacii species group and Teleocichla are characterized by very long branches, and the position of Teleocichla differed considerably between the Bayesian and parsimony trees. This finding does not invalidate Teleocichla but rather suggests that the several monophyletic major clades within Crenicichla may need nominal recognition. A putative hybrid specimen with a morphology combining components from C. vittata and C. scottii, but with a cytochrome b sequence from C. scottii was found in a sample from the Rio Quaraí/Cuareim. Another putative hybrid specimen with a unique morphology but a cytochrome b sequence agreeing with C. scottii was found in a sample from Maldonado, but no other Crenicichla species than C. scottii is known from that locality.     

4-Phenylcoumarins from Mesua elegans with acetylcholinesterase inhibitory activity

A significant acetylcholinesterase (AChE) inhibitory activity was observed for the hexane extract from the bark of Mesua elegans (Clusiaceae). Thus, the hexane extract was subjected to chemical investigation, which led to the isolation of nine 4-phenylcoumarins, in which three are new; mesuagenin A (1), mesuagenin C (3), mesuagenin D (4) and one new natural product; mesuagenin B (2). The structures of the isolated compounds were characterized by spectroscopic data interpretation, especially 1D and 2D NMR. Four compounds showed significant AChE inhibitory activity, with mesuagenin B (2) being the most potent (IC(50)=0.7μM).     

Haplotype and network analysis of island flying fox (Pteropus hypomelanus) using D-loop region of mitochondrial DNA to confirm subspecies designation

Mammal Research - Taxonomic confusion among island flying foxes (Pteropus hypomelanus) still in debate as in the lack of further genetic studies that describe subspecies validity needs...     

Proteasomal degradation of p130 facilitate cell cycle deregulation and impairment of cellular differentiation in high-risk Human Papillomavirus 16 and 18 E7 transfected cells

Molecular Biology Reports - The High-Risk Human Papillomaviruses (HR-HPVs) 16 and 18 are known to cause cervical cancer, which is primarily attributed to E6 and E7 oncoproteins. In addition, recent...