An anti-inflammatory protein secreted from the rat seminal vesicle epithelium inhibits the synthesis of platelet-activating factor and the release of arachidonic acid and prostacyclin

Platelet-activating factor (PAF), a 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine, is a mediator of inflammation and endotoxic shock produced by a variety of stimulated cells. Since the main biosynthetic pathway of PAF involves acetylation of 1-O-alkyl-sn-glycero-3-phosphocholine (lyso-PAF) generated from 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine by phospholipase A2, we suggest a general physiological role played by steroid-induced anti-(phospholipase A2) proteins in the modulation of PAF synthesis. The results of the present study support this hypothesis since an androgen-induced anti-inflammatory protein, SV-IV, secreted from rat seminal vesicles, inhibits PAF synthesis in stimulated polymorphonuclear neutrophils, macrophages and endothelial cells. SV-IV impairs PAF synthesis by inhibiting the activation of phospholipase A2, that also results in the inhibition of arachidonic acid and prostacyclin release, and of acetyl-CoA:lyso-PAF acetyltransferase.     

Assessment of the Chemical Quality of Recreational Bathing Water in Argentina by Health Risk Analysis

Del Azul creek (Argentina) is a natural water body used for recreational bathing in which heavy metals and pesticides have been detected. The aim of the study is to estimate the probabilistic non-cancer and cancer risks by recreational bathing, applying U.S. Environmental Protection Agency models for aggregated (exposure through accidental oral intake of water and dermal contact) and cumulative risks (combined exposure to groups of substances) for bathers of 5, 10, 15, and 20 years old. Bathing in Del Azul creek does not generate risks at the concentrations and the exposure scenarios considered. Cypermethrin and arsenic and heptachlor were the riskiest non-cancer and cancer substances, respectively. Our study highlights the importance of considering both routes of exposure because of the great significance of the dermal route and because of the variability of population characteristics, as it has been stated in other studies. These considerations are highly significant for Argentina, where the quality of recreational water for other than microbiological causes is frequently evaluated based only on the harmful oral intake and applied to a hypothetical individual representative of the population.     

Secretion of a TNFR:Fc fusion protein following pulmonary administration of pseudotyped adeno-associated virus vectors

This study evaluated and compared delivery of the tumor necrosis factor alpha receptor (TNFR)-immunoglobulin G1 (IgG1) Fc fusion (TNFR:Fc) gene to the lung by single and repeat administrations of multiple pseudotyped adeno-associated virus (AAV) vectors as a means for achieving systemic distribution of the soluble TNFR:Fc protein. A single endotracheal administration of AAV[2/5]cytomegalovirus (CMV)-TNFR:Fc vector (containing the AAV2 inverted terminal repeats and AAV5 capsid) to the rat lung resulted in long-term, high levels of serum TNFR:Fc protein that gradually declined over a period of 8 months. Endotracheal delivery of AAV[2/1]CMV-TNFR:Fc resulted in serum TNFR:Fc protein levels that were detectable for at least 4 months but were 10-fold lower than that of the AAV[2/5] vector. In contrast, secretion of the TNFR:Fc protein following pulmonary delivery of AAV[2/2]CMV-TNFR:Fc vector was very inefficient, and the protein was detected in the blood only when an airway epithelial cell-specific promoter, CC10, was substituted for the CMV enhancer/promoter to control transgene expression. In the context of AAV[2/5], the CC10 promoter was as efficient as CMV enhancer/promoter in generating similar levels of systemic TNFR:Fc protein, suggesting that this protein is secreted primarily from the airway epithelium. In mice, comparable long-term secretion of TNFR:Fc protein was demonstrated after AAV[2/2] and AAV[2/5] delivery, although the kinetics of transduction appeared to be different. All pseudotyped AAV vectors elicited serum anti-AAV capsid-neutralizing antibody responses, but these did not prevent lung transduction and efficient secretion of TNFR:Fc protein to the circulation following readministration with AAV[2/5]. These results highlight the potential utility of AAV vectors containing serotype 5 capsid to deliver and redeliver genes of secreted proteins to the lung to achieve long-term systemic protein expression.     

Expression and function of PAIRBP1 within gonadotropin-primed immature rat ovaries: PAIRBP1 regulation of granulosa and luteal cell viability

The protein PAIRBP1, which was initially referred to as RDA288, is involved in mediating the antiapoptotic action of progesterone (P4) in spontaneously immortalized granulosa cells (SIGCs). The present studies were designed to assess the expression and function of PAIRBP1 in the different cell types within the immature rat ovary. Western blot analysis detected PAIRBP1 within whole-cell lysates of immature rat ovaries. Equine gonadotropin (eCG) induced a 3-fold increase in ovarian levels of PAIRBP1. Moreover, human chorionic gonadotropin (hCG), given 48 h after eCG, maintained these elevated levels for up to 4 days. Immunohistochemical analysis confirmed this and further demonstrated that interstitial, thecal, and surface epithelial cells also expressed PAIRBP1. The level of PAIRBP1 in these cells was not influenced by gonadotropin treatment. In contrast, eCG stimulated an increase in PAIRBP1 within the granulosa cells of the developing follicles. Treatment with hCG induced ovulation and ultimately the formation of corpora lutea (CL). High levels of PAIRBP1 expression were also observed within the luteal cells. Immunocytochemical studies on living, nonpermeabilized granulosa and luteal cells revealed that some PAIRBP1 localized to the extracellular surface of these cells. The presence of PAIRBP1 on the extracellular surface was consistent with the observation that an antibody to PAIRBP1 attenuated P4's antiapoptotic action in both granulosa and luteal cells. Although the PAIRBP1 antibody attenuated P4's action, it did not reduce the capacity of cells to specifically bind (3)H-P4. Immunoprecipitation with the PAIRBP1 antibody pulled down the membrane P4 binding protein known as progesterone receptor membrane complex-1 (PGRMC1; rat homolog accession number AJ005837). Taken together, these findings suggest that gonadotropins regulate the expression of PAIRBP1 in granulosa and luteal cells and that PAIRBP1 plays an important role in mediating P4's antiapoptotic action in these ovarian cell types. The exact mechanism of PAIRBP1's action remains to be elucidated, but it may involve an interaction with PGRMC1.     

Cancer and anticancer therapy-induced modifications on metabolism mediated by carnitine system

An efficient regulation of fuel metabolism in response to internal and environmental stimuli is a vital task that requires an intact carnitine system. The carnitine system, comprehensive of carnitine, its derivatives, and proteins involved in its transformation and transport, is indispensable for glucose and lipid metabolism in cells. Two major functions have been identified for the carnitine system: (1) to facilitate entry of long-chain fatty acids into mitochondria for their utilization in energy-generating processes; (2) to facilitate removal from mitochondria of short-chain and medium-chain fatty acids that accumulate as a result of normal and abnormal metabolism. In cancer patients, abnormalities of tumor tissue as well as nontumor tissue metabolism have been observed. Such abnormalities are supposed to contribute to deterioration of clinical status of patients, or might induce cancerogenesis by themselves. The carnitine system appears abnormally expressed both in tumor tissue, in such a way as to greatly reduce fatty acid beta-oxidation, and in nontumor tissue. In this view, the study of the carnitine system represents a tool to understand the molecular basis underlying the metabolism in normal and cancer cells. Some important anticancer drugs contribute to dysfunction of the carnitine system in nontumor tissues, which is reversed by carnitine treatment, without affecting anticancer therapeutic efficacy. In conclusion, a more complex approach to mechanisms that underlie tumor growth, which takes into account the altered metabolic pathways in cancer disease, could represent a challenge for the future of cancer research.     

A plausible mechanism of electron transfer between quinones in photosynthetic reaction centers

The mechanism of long-range electron transfer between the primary and the secondary quinone of photosynthetic reaction centers has been investigated, with particular attention on the role of the iron-histidine bridge. Computations suggest that in such a system, where the molecular subunits are packed together by H-bonds, a mobile electron, injected on one end of the chain, can be carried to the other end by switching the positions of the H-bonded hydrogens. Energy estimates would suggest that the proposed mechanism is plausible and worthy of further experimental investigations.     

Use of nodulation pattern, stress tolerance, nodC gene amplification, RAPD-PCR and RFLP-16S rDNA analysis to discriminate genotypes of Rhizobium leguminosarum biovar viciae

Twenty-seven new Rhizobium isolates were obtained from root nodules of wild and crop legumes belonging to the genera Vicia, Lathyrus and Pisum from different agroecological areas in central and southern Italy. A polyphasic approach including phenotypic and genotypic techniques was used to study their diversity and their relationships with other biovars and species of rhizobia. Analysis of symbiotic properties and stress tolerance tests revealed that wild isolates showed a wide spectrum of nodulation and a marked variation in stress tolerance compared with reference strains tested in this study. All rhizobial isolates (except for the isolate CG4 from Galega officinalis) were presumptively identified as Rhizobium leguminosarum biovar viciae both by their symbiotic properties and the specific amplification of the nodC gene. In particular, we found that the nodC gene could be used as a diagnostic molecular marker for strains belonging to the bv. viciae. RFLP-PCR 16S rDNA analysis confirms these results, with the exception of two strains that showed different RFLP-genotypes from those of the reference strains of R. leguminosarum bv. viciae. Analysis of intraspecies relationship among strains by using the RAPD-PCR technique showed a high level of genetic polymorphism, grouping our isolates and reference strains into six different major clusters with a similarity level of 20%. Data from seven parameters of phenotypic and genotypic analyses were evaluated by using principal component analysis which indicated the differences among strains and allowed them to be divided into seven different groups.     

Total synthesis of ichthyotoxic macrolides from the skin of the marine mollusk <Emphasis Type="Italic">Aplysia depilans</Emphasis>

A convergent pathway is described for the synthesis of natural aplyolides A–E (1–5), ichthyotoxic macrolides isolated from the skin of the marine mollusk Aplysia depilans. Aplyolide A (1) was prepared using ethyl (S)-lactate as chiral source. Key diols for the preparation of aplyolides B–E (2–5) were obtained stereoselectively by Sharpless asymmetric dihydroxylation of eneyne precursors. An improved procedure, based on cesium carbonate as base for alkynide preparation, was used in key steps of the synthesis involving coupling reactions between copper (I) alkynides and propargylic halides for the formation of skipped diyne systems. Finally, the crucial step of macrocyclization was accomplished using the Yamaguchi methodology.