Protoporphyrin IX enhancement by 5-aminolaevulinic acid peptide derivatives and the effect of RNA silencing on intracellular metabolism

Intracellular generation of the photosensitiser, protoporphyrin IX, from a series of dipeptide derivatives of the haem precursor, 5-aminolaevulinic acid (ALA), was investigated in transformed PAM212 murine keratinocytes, together with studies of their intracellular metabolism. Porphyrin production was substantially increased compared with equimolar ALA using N-acetyl terminated phenylalanyl, leucinyl and methionyl ALA methyl ester derivatives in the following order: Ac-L-phenylalanyl-ALA-Me, Ac-L-methionyl-ALA-Me and Ac-L-leucinyl-ALA-Me. The enhanced porphyrin production was in good correlation with improved photocytotoxicity, with no intrinsic dark toxicity apparent. However, phenylalanyl derivatives without the acetyl/acyl group at the N terminus induced significantly less porphyrin, and the replacement of the acetyl group by a benzyloxycarbonyl group resulted in no porphyrin production. Porphyrin production was reduced in the presence of class-specific protease inhibitors, namely serine protease inhibitors. Using siRNA knockdown of acylpeptide hydrolase (ACPH) protein expression, we showed the involvement of ACPH, a member of the prolyl oligopeptidase family of serine peptidases, in the hydrolytic cleavage of ALA from the peptide derivatives. In conclusion, ALA peptide derivatives are capable of delivering ALA efficiently to cells and enhancing porphyrin synthesis and photocytotoxicity; however, the N-terminus state, whether free or substituted, plays an important role in determining the biological efficacy of ALA peptide derivatives

Mating Behaviour in Laevicaudatan Clam Shrimp (Crustacea, Branchiopoda) and Functional Morphology of Male Claspers in a Phylogenetic Context: A Video-Based Analysis

Clam shrimps are freshwater branchiopod crustaceans which often present complicated breeding systems including asexual reproduction (parthenogenesis) and mixed mating systems (in androdioecious species both selfing and outcrossing occurs due to the co-presence of hermaphrodites and males). Reproductive patterns of Spinicaudata, which contains most clam shrimp species, have received much attention. Another group of clam shrimps, Laevicaudata, which holds a key position in branchiopod phylogeny, has practically not been studied. As a part of the mating process, males clasp to the carapace margin of the females with a pair (or two pairs) of anterior trunk limbs modified as claspers. Previous studies have shown that clasper morphology is important in a phylogenetic context, and that some parts of the claspers in Spinicaudata and Laevicaudata may have undergone a remarkable parallel evolution. Here we have used video microscopy to study aspects of the mating behaviour, egg extrusion, and fertilization in Lynceus brachyurus (Laevicaudata). It is shown that fertilization is likely to be external and that the peculiar tri-lobed lateral lamellae of female's hind body assist in guiding the egg mass to the exopodal egg carriers where they are collected by their distal setation. The functional morphology of the male claspers was studied in detail by close-up video recordings. The movable “finger” of the clasper bends around the female's carapace edge and serves to hold the female during mating. The larger palp grasps around the female carapace margin in a way very similar to the movable “finger”, possibly indirectly providing sensory input on the “finger” position. A brief comparative study of the claspers of a spinicaudatan clam shrimp showed both similarities and differences to the laevicaudatan claspers. The presence of two pairs of claspers in Spinicaudata seems to give males a better hold of the female which may play a role during extended mate guarding

Fish oil and the prevention and regression of atherosclerosis

Epidemiological studies in the seventies have put forward that dietary rather than genetic factors are responsible for the lower incidence of ischemic heart disease in Greenland Inuit and have generated a large body of both in vitro and in vivo experimental studies, exploring the putative favorable effects of fish (oil) on atherogenesis and its risk factors. The first part of this report reviews the in vivo animal studies, concentrating on the hypercholesterolemic models and the arterialized vein graft model. In the hypercholesterolemic animal studies, the results are inconclusive as the studies reporting a protective effect are matched by the number of studies showing no effect or an adverse effect. The diversity in species, dose of fish oil, duration of study, type of vessel studied and type of fish oil preparation (content of n-3 fatty acids, unesterified n-3 fatty acids, ethylesters or triglycerides) could all contribute. Furthermore, the definitions and criteria used in the literature to evaluate atherogenesis are diverse and it appears that while one parameter is affected, another is not necessarily modified in the same direction, stressing the importance of extending the analysis of the effects on atherogenesis to more than one parameter. We also believe that it is time to reach a consensus as to which animal model mimicks most closely a particular human situation. Only in appropriate models, investigating more than one atherosclerosis variable, can the effects of a putative anti-atherogenic drug or diet be verified. In the veno-arterial autograft model, mimicking the patient after coronary bypass grafting, dietary fish oil has been consistently effective in preventing accelerated graft intima proliferation. It could therefore be of interest to evaluate the effects of fish oil on graft patency in patients after coronary bypass surgery after a period of years. The results from studies on restenosis after percutaneous transluminal angioplasty are also reviewed and it is concluded that the two large scale trials, that are currently underway, might reliably answer the question whether fish oil is effective as a non-pharmacological adjuvants in the prevention of restenosis. Lastly, the studies on the effects of fish oil on the regression of experimental atherosclerosis are reviewed. In view of the small number of studies (i.e., four) investigating the effects of fish oil on the regression of atherosclerosis, it is premature to draw any conclusion, and therefore further experimental work is required