Cultured primary human keratinocytes are frequently employed for studies of immunological and inflammatory responses; however, interpretation of experimental data may be complicated by donor to donor variability, the relatively short culture lifetime, and variations between passages. To standardize the in vitro studies on keratinocytes, we investigated the use of HaCaT cells, a long-lived, spontaneously immortalized human keratinocyte line which is able to differentiate in vitro, as a suitable model to follow the release of inflammatory and repair mediators in response to TNF伪 or IL-1尾. Different treatment conditions (presence or absence of serum) and differentiation stimuli (increase in cell density as a function of time in culture and elevation of extracellular calcium) were considered. ELISA and Multiplex measurement technologies were used to monitor the production of cytokines and chemokines. Taken together, the results highlight that Ca2+ concentration in the medium, cell density, and presence of serum influences at different levels the release of proinflammatory mediators by HaCaT cells. Moreover, HaCaT cells maintained in low Ca2+ medium and 80% confluent are similar to normal keratinocytes in terms of cytokine production suggesting that HaCaT cells may be a useful model to investigate anti-inflammatory interventions/therapies on skin diseases.
We have cloned and sequenced a cDNA encoding human liver carnitine palmitoylransferase (CPTase; pahnitoyl-CoA:L-carnitine O-palmitoyltransferase, EC 2.3.1.21), an inner mitochondrial membrane enzyme that plays a major role in the fatty acid oxidation pathway. Mixed oligonudeotide primers whose sequences were deduced from one tryptic peptide obtained from purified CPTase were used in a polymerase chain reaction, allowing the amplification of a 0. A major source for energy production in the cell is the mitochondrial p-oxidation of long-chain fatty acids. In order to cross the mitochondrial membranes, long-chain fatty acids are first activated by coenzyme A and then reversibly conjugated with L-carnitine, a reaction that is catalyzed by the enzyme carnitine palmitoyltransferase (CPTase; palmitoylCoA:L-carnitine O-palmitoyltransferase, EC 2.3.1.21) (1). One part of CPTase activity, conventionally referred to as CPTase I, is associated with the outer mitochondrial membrane, and another significant pool of CPTase activity, conventionally referred to as CPTase II, is in close relationship with the inner mitochondrial membrane (2). However, all the attempts to purify CPTase led to the identification of a single protein with a subunit of Mr 70 kDa (3)(4)(5)42).An important role in the regulation of CPTase activity is played by malonyl-CoA, an intermediate in fatty acid biosynthesis that inhibits the activity of outer (CPTase I) but not of inner (CPTase II) CPTase (6). The malonyl-CoA binding domain has been proposed to be located either on CPTase I (4) or on a regulatory protein without CPTase activity (7).CPTase activity is also under hormonal control of estrogen (8), insulin, and glucagon. Diabetes causes increased CPTase activity and decreased affinity for malonyl-CoA, both reversed by insulin (9). Glucagon has been reported to stimulate CPTase through phosphorylation by a cAMP-dependent protein kinase (10). Interestingly, hypoglycemic agents, such as sulfonylureas, exercise their pharmacological action by strongly inhibiting CPTase activity (11).CPTase seems also to play a role in the regulation of the hepatic synthesis of very low density lipoproteins (VLDL), since its inhibition increases VLDL production, whereas activation following depletion of malonyl-CoA levels decreases VLDL synthesis (12, 13). Accordingly, the hypolipidemic drug lovastatin causes a 2-fold increase in the activity of hepatic CPTase (14).From these observations it appears that CPTase has a role in the development of metabolic alterations in very common diseases such as diabetes and hyperlipoproteinemias. Furthermore, recessively inherited deficiency of CPTase in man has been described with two distinct phenotypes. In adults, CPTase deficiency causes a muscle disease with weakness, cramps, and myoglobinuria (15). In infants, CPTase deficiency is characterized by hyperammonemia, increased levels of serum transaminases and plasma-free fatty acids, hepatomegaly, nonketotic hypoglycemia, and coma (16). This life-threatening and often fatal d...
These experiments suggest that human tumors expressing UK114 on the surface of the cell membrane may undergo antibody-mediated cytolysis. Thus, UKll4 is thought to be the main significant component of UK101 with potential pharmacological activity. Preliminary structural results suggested that UK114 might be a protein not yet characterized [6]. In this paper we present the complete primary structure of UK114 as a preliminary step to investigation of its biological activity. Material and methods
Sphingolipids are widespread membrane components that are found in all eukaryotic cells. They consist of a long chain sphingoid-base, usually sphingosine, which is acylated at the 2-amino position, forming a ceramide. All together, sphingolipids may represent the most structurally diverse category of lipids in nature. There is no known nutritional requirement for sphingolipids. Nonetheless, studies with experimental animals have shown that consumption of sphingolipids inhibits colon carcinogenesis, reduces serum low-density lipoprotein cholesterol and elevates high-density lipoproteins, which suggest that they are 'functional' components of food. In animal models (CF1 mice) sphingomyelin supplementation reduces the number of aberrant colonic crypt foci by approximately 70% and, with longer feeding, reduces the number of colonic adenocarcinomas. A possible mechanism of action of sphingolipids in suppressing colon carcinogenesis is that exogenously supplied sphingolipids bypass a sphingolipid signalling defect that is important in cancer (for example, a loss of cellular sphingomyelin turnover to produce ceramide and sphingosine). Indirect evidence suggests that sphingolipids can inhibit colon cancer in humans: sphingosine and ceramide induce apoptosis in a human adenocarcinoma cell line and feeding sphingolipids to Min mice reduces the number of colon tumours.
In recent years, there has been a growing interest in natural antioxidants as replacements of synthetic compounds because of increased safety concerns and worldwide trend toward the usage of natural additives in foods. One of the richest sources of natural antioxidants, nowadays largely studied for their potential to decrease the risk of diseases and to improve oxidative stability of food products, are edible brown seaweeds. Nevertheless, their antioxidant mechanisms are slightly evaluated and discussed. The aims of this study were to suggest possible mechanism(s) of Fucus vesiculosus antioxidant action and to assess its bioactivity during the production of enriched rye snacks. Chemical and cell-based assays indicate that the efficient preventive antioxidant action of Fucus vesiculosus extracts is likely due to not only the high polyphenol content, but also their good Fe 2+ -chelating ability. Moreover, the data collected during the production of Fucus vesiculosus-enriched rye snacks show that this seaweed can increase, in appreciable measure, the antioxidant potential of enriched convenience cereals. This information can be used to design functional foods enriched in natural antioxidant ingredients in order to improve the health of targeted consumers.
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