MV-mimicking micelles loaded with PEG-serine-ACP nanoparticles to achieve biomimetic intra/extra fibrillar mineralization of collagen in vitro

Since their discovery, matrix vesicles (MVs) containing minerals have received considerable attention for their role in the mineralization of bone, dentin and calcified cartilage. Additionally, MVs' association with collagen fibrils, which serve as the scaffold for calcification in the organic matrix, has been repeatedly highlighted. The primary purpose of the present study was to establish a MVs–mimicking model (PEG-S-ACP/micelle) in vitro for studying the exact mechanism of MVs-mediated extra/intra fibrillar mineralization of collagen in vivo. In this study, high-concentration serine was used to stabilize the amorphous calcium phosphate (S-ACP), which was subsequently mixed with polyethylene glycol (PEG) to form PEG-S-ACP nanoparticles. The nanoparticles were loaded in the polysorbate 80 micelle through a micelle self-assembly process in an aqueous environment. This MVs–mimicking model is referred to as the PEG-S-ACP/micelle model. By adjusting the pH and surface tension of the PEG-S-ACP/micelle, two forms of minerals (crystalline mineral nodules and ACP nanoparticles) were released to achieve the extrafibrillar and intrafibrillar mineralization, respectively. This in vitro mineralization process reproduced the mineral nodules mediating in vivo extrafibrillar mineralization and provided key insights into a possible mechanism of biomineralization by which in vivo intrafibrillar mineralization could be induced by ACP nanoparticles released from MVs. Also, the PEG-S-ACP/micelle model provides a promising methodology to prepare mineralized collagen scaffolds for repairing bone defects in bone tissue engineering.     

Paleolimnological evidence for recent acidification of Big Moose Lake,Adirondack Mountains,N.Y. (USA)

Big Moose L. has become significantly more acidic since the 1950s, based on paleolimnological analyses of sediment cores. Reconstruction of past lakewater pH using diatom assemblage data indicates that from prior to 1800 to ca. 1950, lakewater pH was about 5.8. After the mid-1950s, the inferred pH decreased steadily and relatively quickly to about 4.6. Alkalinity reconstructions indicate a decrease of about 30 eq · l^-1 during the same period. There was a major shift in diatom assemblage composition, including a nearly total loss of euplanktonic taxa. Chrysophyte scale assemblages and chironomid (midge larvae remains also changed in a pattern indicating decreasing lakewater pH starting in the 1950s. Accumulation rates of total Ca, exchangeable and oxide Al, and other metals suggest recent lake-watershed acidification. Cores were dated using²¹⁰Pb, pollen, and charcoal. Indicators of watershed change (deposition rates of Ti, Si, Al) do not suggest any major erosional events resulting from fires or logging. Accumulation rates of materials associated with combustion of fossil fuels (polycyclic aromatic hydrocarbons, coal and oil soot particles, some trace metals, and sulfur) are low until the late 1800s-early 1900s and increase relatively rapidly until the 1920s–1930s. Peak rates occurred between the late 1940s and about 1970, when rates declined.The recent decrease in pH of Big Moose L. cannot be accounted for by natural acidification or processes associated with watershed disturbance. The magnitude, rate and timing of the recent pH and alkalinity decreases, and their relationship to indicators of coal and oil combustion, indicate that the most reasonable explanation for the recent acidification is increased atmospheric deposition of strong acids derived from combustion of fossil fuels.     

Genotoxicity studies with mineral oils; effects of oils on the microbial mutagenicity of precursor mutagens and genotoxic metabolites

In vitro genotoxicity assays are extensively used to predict carcinogenic activity in vivo. The standard microbial mutagenicity assays however often fail to yield positive results with mineral oils which are carcinogenic to mice in long-term skin-cancer studies. A comprehensive programme of studies has therefore investigated the basis of this apparently anomalous behaviour. This investigation has addressed the possible effects of oils on the bioactivation of precursor mutagens and the disposition of mutagenic metabolites by studying the microbial mutagenicity of selected precursor mutagens (benzo[a]pyrene, benzo[a]anthracene, 2-aminoanthracene and 2-naphthylamine) and intrinsically reactive mutagens [+/- )-benzo[a]pyrene-4,5-oxide and (+/-)-7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene) in the presence and absence of mineral oils. Notably the mutagenicity associated with the deliberate additions of these mutagens or precursor mutagens to oils was readily detected by the microbial assays. The mutagenicity of only one of the precursor mutagens, benzo[a]pyrene, was significantly reduced by the oils, and then only in the standard plate-incorporation assay. Interestingly the degree of suppression appeared to be related to the polycyclic aromatic hydrocarbon content of the oils. In the case of 2-aminoanthracene large enhancements in its mutagenicity were observed in the presence of oils. These latter findings appear to be due to effects of oils on the bioactivation of precursor mutagens rather than on the disposition of their bioactivation products. The mutagenicity of intrinsically reactive mutagens, of a type generated by bioactivation of polycyclic aromatic hydrocarbons, was not significantly reduced in the presence of mineral oils. This indicates that it is unlikely that components in oils trap or facilitate the deactivation of ultimate mutagens whether these pre-exist in the oil or are formed from precursors by bioactivation in the in vitro test system. Viewed overall these results suggest that mineral oils judged to be carcinogenic on the basis of in vivo studies in mouse skin may possess only very weak genotoxic potential. While this potential is likely to be a prerequisite for carcinogenic action, the current results cause attention to be focussed on other factors, e.g. promotion, as potentially important determinants of the carcinogenic potencies of mineral oils in mouse skin.     

In vitro metabolism of N-methyl-dibenzo [c,g]carbazole a potent sarcomatogen devoid of hepatotoxic and hepatocarcinogenic properties

The metabolism of N-methyl substituted 7H-dibenzo[c,g]carbazole (N-Me DBC) was investigated in vitro using liver microsomes from 3-methylcholanthrene (MC)-, benzo[c]carbazole (BC) and Arochlor-pretreated mice and rats. N-Me DBC is a potent sarcomatogen devoid of hepatotoxicity and liver carcinogenic activity. The ethyl acetate-extractable metabolites were separated by high performance liquid chromatography (HPLC) and most of them were identified by proton magnetic resonance (PMR), mass spectrometry (MS) and comparison with synthetically prepared specimens. Mouse and rat microsomes gave rise to the same metabolites. The major metabolites were 5-OH-N-Me DBC (50%), N-hydroxymethyl (HMe) DBC (25-30%) and 3-OH-N-Me DBC (10%). Addition of 1,1,1-trichloropropene-2,3-oxide (TCPO) to the standard incubation medium permitted the identification of two dihydrodiols among the minor metabolites. No metabolite of DBC was observed after incubation of N-Me DBC, or its major metabolite N-HMe DBC, with either mouse or rat microsomes, but the possibility of a slight demethylation cannot be totally excluded. The lack of biotransformation at the nitrogen atom site may explain the lack of hepatotoxicity and liver carcinogenic activity of N-Me DBC. The modulation of metabolism by epoxide hydrolase, cytosol and glutathione was also investigated. The results are discussed in the light of data previously obtained with hepatotoxic and hepatocarcinogenic DBC.     

The formation and biotransformation of cysteine conjugates of halogenated ethylenes by rabbit renal tubules

The nephrotoxicity of chlorotrifluoroethylene (CTFE) was examined using isolated rabbit renal tubules suspensions. Exposure of the tubules to CTFE resulted in consumption of CTFE, formation of a glutathione conjugate and inhibition of active organic acid transport. Synthetic cysteine, N-acetylcysteine or glutathione conjugates of CTFE inhibited transport indicating S-conjugation as a possible toxic pathway. 1,2-dichlorovinyl glutathione (DCVG), a model synthetic glutathione conjugate, was used to examine the degradation and toxicity of these conjugates. DCVG inhibited rabbit renal tubule transport in vivo and in vitro. The DCVG was found to be degraded with the evolution of glutamine and glycine to produce the ultimate nephrotoxicant, dichlorovinyl cysteine. Dichlorovinyl cysteine is then bioactivated with the release of ammonia. This sequential degradation explains the latency of DCVG-induced renal transport inhibition relative to dichlorovinyl cysteine. It is now evident that certain halogenated ethylenes are capable of being biotransformed to glutathione conjugates in the kidney with their subsequent hydrolysis to nephrotoxic cysteine conjugates.     

Inhibition of epidermal metabolism and DNA-binding of benzo[a]pyrene by ellagic acid

Ellagic acid, a common plant phenol, was shown to be a potent inhibitor of epidermal microsomal aryl hydrocarbon hydroxylase (AHH) activity in vitro, and of benzo[a]pyrene (BP)-binding to both calf thymus DNA in vitro and to epidermal DNA in vivo. The in vitro addition of ellagic acid (0.25-2.0 microM) resulted in a dose-dependent inhibition of AHH activity in epidermal microsomes prepared from control or carcinogen-treated animals. The I50 of ellagic acid for epidermal AHH was 1.0 microM making it the most potent inhibitor of epidermal AHH yet identified. In vitro addition of ellagic acid to microsomal suspensions prepared from control or coal tar-treated animals resulted in 90% inhibition of BP-binding to calf thymus DNA. Application of ellagic acid to the skin (0.5-10.0 mumol/10 gm body wt) caused a dose-dependent inhibition of BP-binding to epidermal DNA. Our results suggest that phenolic compounds such as ellagic acid may prove useful in modulating the risk of cutaneous cancer from environmental chemicals.     

Transfer of polycyclic aromatic hydrocarbons between model membranes: relation to carcinogenicity

Polynuclear aromatic hydrocarbons (PAH), some of which are potent carcinogens, are common environmental pollutants. The transport processes for these hydrophobic compounds into cells and between intracellular membranes are diverse and are not well understood. A common mechanism of transport is by spontaneous desorption and transfer through the aqueous phase. From the partitioning parameters, we have inferred that the rate limiting step involves solvation of the transfer species in the interfacial water at the phospholipid surface. Transfer of 10 PAH (pyrene, 3,4-benzophenanthrene, triphenylene, chrysene, 1,2-benzanthracene, 1,1'-binaphthyl, 9-phenylanthracene, 2,2'-binaphthyl, m-tetraphenyl and 1,3,5-triphenylbenzene) out of phosphatidylcholine vesicles has been examined. Our results show that the molecular volume of the PAH is a rate-determining factor. Moreover, high performance liquid chromatography (HPLC) data confirms the hypothesis that the rate of transfer is correlated with the size of the molecule and with the partitioning of the molecule between a polar and hydrocarbon phase. The kinetics and characteristics of the spontaneous transfer of carcinogens are likely to have a major impact on the competitive processes of PAH metabolism within cells.     

Cutaneous metabolism of benzo[a]pyrene: comparative studies in C57BL/6N and DBA/2N mice and neonatal Sprague--Dawley rats

The metabolism of the polycyclic aromatic hydrocarbon (PAH) carcinogen benzo[a]pyrene (BaP) was studied using microsomes prepared from the skin of the mouse and rat. Topical application of the polychlorinated biphenyl (PCB) Aroclor 1254 or the PAH 3-methylcholanthrene (3-MC) to the skin of the C57BL/6N and DBA/2N mouse and the Sprague-Dawley rat caused statistically significant enhancement of cutaneous microsomal aryl hydrocarbon hydroxylase (AHH) activity in each animal. PCB was a more potent inducer of the enzyme than was 3-MC. BaP metabolism by skin microsomes from the same animals was assessed using high performance liquid chromatography (HPLC). The skin of untreated animals metabolized BaP into 9,10-, 7,8- and 4,5-dihydrodiols, phenols and quinones. Skin application of PCB caused greater than 16–18-fold enhancement of BaP metabolism in the C57BL/6N mouse and the rat and 2–5-fold enhancement in the DBA/2N mouse. Skin application of 3-MC enhanced BaP metabolism 2–8-fold in the C57BL/6N mouse and 5–10-fold in the rat and had no effect in the DBA/2N mouse. The formation of procarcinogenic metabolite BaP-7, 8-diol was greatly enhanced (4–12-fold) by treatment with the PCB and 3-MC in the tumor susceptible C57BL/6N mouse and in the tumor-resistant neonatal Sprague-Dawley rat. In contrast, the formation of BaP-7,8-diol was either slightly enhanced (2-fold) or unaffected by treatment with the PCB or 3-MC in the tumor-resistant DBA/2N mouse. Our data indicate that neither the patterns of metabolism nor the amount of BaP-7,8-diol formation in the skin are reliable predictors of tumor susceptibility to the PAH in rodent skin.     

九种罕见的人类染色体异常核型报告

ReportonNineRareSpeciesofHumanChromosomalAbnormalKaryotypesHanWeitian;QuOu;YuPing;JiangMiao(LiaoningResearchInstituteofFamilyPlanning,Shenyang110031)自1983年以来,我们对数千例不育及胚胎丢失等生殖异常患者进行细胞遗传学研究,发现大量异常染色体核型,而且异常种类繁多,已报道世界首报人类异常核型25种[1]。最近,在不良妊娠患者中又发现9种异常核型,经湖南医科大学医学细胞遗传学国家培训中心鉴定,为世界首次报道．现报告如下。1病例摘要及核型例1男,30岁,表型及智力正常,其妻子妊娠两次,均在无任何诱因情…