We have examined components of the preintegration complex of human immunodeficiency virus type 1 (H1V-1) and have analyzed features which govern the association of these components. HIV-1 nucleoprotein complexes, isolated from nuclear and cytoplasmic extracts of CD4+ cells after acute virus infection, contained viral RNA and DNA in association with viral matrix (MA), integrase (IN), and reverse ranscriptase (RT) antigens but not capsid (CA) antigens and posessed integration activity in vitro. Association of IN but not RT or MA antigens with viral DNA was detergent-stable. Analysis of viral DNA synthesis and nuclear import of viral nucleoprotein complexes in the presence of a reversible RT inhibitor demonstrated that reverse transcription of viral RNA could be completed entirely in the host cell nucleus. Our studies demonstrate structural and functional features of the nudeoprotein (preintegration) complex of HIV-1 which are pertinent to the understanding of early events in the lentiviral life cycle.

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The serum concentrations of haptoglobin, serum amyloid A and alpha1 acid glycoprotein were determined in serum collected from healthy dairy cows and cows with clinical mastitis, graded as mild (clots in milk) or moderate (clots in milk and visible signs of inflammation in the mammary gland/s) to assess their relative diagnostic value in detecting the disease. The concentrations of haptoglobin and serum amyloid A were also measured in milk collected from infected and uninfected quarters. The concentrations of haptoglobin and serum amyloid A were higher in the serum and milk from the cows with mild or moderate mastitis. The diagnostic value of haptoglobin in differentiating between healthy animals and those with mastitis gave sensitivities and specificities of 82 per cent and 94 per cent respectively with serum and 86 per cent and 100 per cent with milk. The diagnostic value of serum amyloid A in differentiating between healthy animals and those with mastitis gave sensitivities and specificities of 83 per cent and 90 per cent with serum and 93 per cent and 100 per cent with milk. The diagnostic value of serum alpha1 acid glycoprotein in differentiating between healthy animals and those with mastitis gave sensitivities and specificities of 62 per cent and 91 per cent.

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SummaryWe have engineered the chloroplast of eukaryotic algae to produce a number of recombinant proteins, including human monoclonal antibodies, but, to date, have achieved expression to only 0.5% of total protein. Here, we show that, by engineering the mammalian coding region of bovine mammary-associated serum amyloid (M-SAA) as a direct replacement for the chloroplast psbA coding region, we can achieve expression of recombinant protein above 5% of total protein. Chloroplast-expressed M-SAA accumulates predominantly as a soluble protein, contains the correct amino terminal sequence and has little or no post-translational modification. M-SAA is found in mammalian colostrum and stimulates the production of mucin in the gut, acting in the prophylaxis of bacterial and viral infections. Chloroplast-expressed and purified M-SAA is able to stimulate mucin production in human gut epithelial cell lines. As Chlamydomonas reinhardtii is an edible alga, production of therapeutic proteins in this organism offers the potential for oral delivery of gut-active proteins, such as M-SAA.

Creatine is widely used by both elite and recreational athletes as an ergogenic aid to enhance anaerobic exercise performance. Older individuals also use creatine to prevent sarcopenia and, accordingly, may have therapeutic benefits for muscle wasting diseases. Although the effect of creatine on the musculoskeletal system has been extensively studied, less attention has been paid to its potential effects on other physiological systems. Because there is a significant pool of creatine in the brain, the utility of creatine supplementation has been examined in vitro as well as in vivo in both animal models of neurological disorders and in humans. While the data are preliminary, there is evidence to suggest that individuals with certain neurological conditions may benefit from exogenous creatine supplementation if treatment protocols can be optimized. A small number of studies that have examined the impact of creatine on the immune system have shown an alteration in soluble mediator production and the expression of molecules involved in recognizing infections, specifically toll-like receptors. Future investigations evaluating the total impact of creatine supplementation are required to better understand the benefits and risks of creatine use, particularly since there is increasing evidence that creatine may have a regulatory impact on the immune system.

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Recent studies have shown that the alcohol metabolites malondialdehyde and acetaldehyde can combine to form a stable adduct (MAA) on proteins. This adduct has been detected in the livers of rats chronically consuming ethanol, and serum antibodies to MAA have been observed at significantly higher concentrations in ethanol-fed when compared with pair-fed or chow-fed control rats. More recently, preliminary studies have strongly suggested that the MAA adduct is capable of stimulating antibody responses to soluble proteins in the absence of adjuvants. The antibodies produced recognize either the MAA epitope or the carrier protein itself. Therefore, it was the purpose of this study to examine the potential immunogenicity of MAA-modified exogenous proteins in the absence of adjuvants. Balb/c mice were immunized in the presence or absence of adjuvant with different concentrations of unmodified or MAA-modified proteins. The antibody response to both the MAA epitope and unmodified protein epitopes were determined by ELISA. In the absence of adjuvant, significant antibody responses were induced to both the MAA epitope and nonmodified protein epitopes. Smaller immunizing doses of MAA-protein conjugate favored the production of antibodies to nonmodified proteins, whereas larger doses induced a strong anti-MAA response. In studies to begin determining a mechanism for the specificity of the response in the absence of adjuvants, peritoneal macrophages were found to bind and degrade MAA-adducted proteins through the use of a scavenger receptor. This indicated that MAA-adducted proteins may be specifically taken up and epitopes presented to the humoral immune system in the absence of adjuvants. Importantly, these are the first data showing that an alcohol-related metabolite can induce an antibody response in the absence of adjuvant and suggesting a mechanism by which antibody to the MAA adduct or its carrier (exogenous or endogenous) proteins may be generated in vivo.