The influence of protein adsorption on interactions of cultured human endothelial cells with polymers

A systematic study of the effects of polymer surface properties on the interaction with human endothelial cells (HEC) may lead to the development of small-diameter vascular grafts. HEC, suspended in culture medium containing 20% serum adhered and spread onto moderately wettable polymers such as TCPS (tissue culture polystyrene). Reduced or no adhesion of HEC was observed upon the hydrophobic polymers PETP (polyethyleneterephthalate, Dacron) and FEP (fluoroethylenepropylene copolymer, Teflon). Polymers precoated with the proteins albumin (Alb), high density lipoprotein (HDL), and immunoglobulin G (IgG) inhibited the adhesion of HEC, whereas fibronectin (Fn) coátings promoted cell adhesion. Endothelialization of PETP and FEP only occurred after precoating of these materials with Fn. The adsorption of Fn, Alb, HDL, and IgG from solutions of different serum concentrations onto TCPS, PETP, and FEP was related to the adhesion of HEC. Serum Fn only adsorbed onto TCPS, with the maximum at 0.1% serum concentration. Maximal cell adhesion onto TCPS was also observed after pretreatment with a solution containing 0.1% serum. The cell adhesion inhibiting proteins Alb and HDL preferentially adsorbed at higher serum concentrations. Desorption of these proteins and exchange for, e. g., cellular Fn may result in cell spreading and proliferation of HEC upon TCPS

Characterization of eukaryotic cell surfaces prior to and after serum protein adsorption by X-ray photoelectron spectroscopy - Fibroblasts, HELA epithelial, and smooth muscle cells

Elemental surface concentration ratios N/C, O/C, and P/C of fibroblasts, HELA epithelial cells, and smooth muscle cells, prior to and after washing in the absence or presence of serum proteins, were determined by X-ray photoelectron spectroscopy. Cell surfaces appeared to adsorb hardly any serum proteins, and the relatively high P/C, as compared to N/C and O/C, elemental surface concentration ratio indicated that the cell surfaces consisted mainly of the phospholipid bilayer, with little or no proteins present. The lack of adsorption of serum proteins to the cell surfaces seems at odds with the common notion that cells require adhesive proteins in order to adhere and spread. However, the adsorption behavior of cellularly produced proteins may be completely different, particularly since they seem to be able to displace adsorbed serum proteins from biomaterials surfaces. Interestingly, only HELA epithelial cells (a tumor cell line) appeared to adsorb a very small amount of proteins.</p

Pathophysiology of elevated ascites fluid cholesterol in malignant ascites

The existence of marked elevations of ascitic fluid cholesterol has been observed in patients with peritoneal carcinomatosis compared to patients with cirrhosis and has been found useful in differential diagnosis. This finding could be caused by an enhanced movement of plasma lipoproteins into the peritoneal cavity. To test this hypothesis we determined the fasting concentrations of total, high density lipoprotein (HDL)- and low density lipoprotein (LDL)-cholesterol, apolipoprotein-A1 (apo-A1) and apolipoprotein-B (apo-B) in serum and ascites of 17 patients with cirrhosis and 16 patients with peritoneal carcinomatosis. The movement of proteins from plasma to ascites was calculated from the ascites/serum concentration ratios of six different sized proteins with a molecular mass ranging from 54 kDa to 971 kDa. Mean values (mg/dl) for total cholesterol (92.6 vs. 21.0), HDL-cholesterol (15.6 vs. 1.8), LDL-cholesterol (63.4 vs. 16.1), apo-A1 (50.2 vs. 13.6) and apo-B (41.2 vs. 12.9) in ascites were significantly higher in peritoneal carcinomatosis than in cirrhosis. These differences could only partially be explained by the higher serum concentrations of these parameters in peritoneal carcinomatosis, but were mainly due to a lower selectivity for the movement of plasma proteins and lipoproteins into ascites (mean ascites/serum (A/S) ratio: 0.30–0.77) in peritoneal carcinomatosis as compared to cirrhosis (mean ascites/serum ratio: 0.11–0.21). In both groups about 85% of the total cholesterol in serum and ascites consisted of HDL- and LDL-cholesterol. These findings support the hypothesis that elevations in ascitic cholesterol in peritoneal carcinomatosis compared to cirrhosis are mainly caused by the increased movement of plasma HDL and LDL into the peritoneal cavity

EFFECT OF AMPICILLIN AND CHLORAMPHENICOL ON CHICK SERUM

  Objective: Cystatin protein present in chick serum exhibits antimicrobial activity. The present study focuses on the effect of ampicillin and chloramphenicol antibiotics on chick serum and thus verifies Beer–Lambert's law.Methods: The serum is separated from the cellular matter with the help of a micropipette to get a clean serum sample. The quantification of protein was done by Lowry's method. The antibiotics ampicillin and chloramphenicol stock solution were prepared by 10 mg of antibiotic powder in 10 ml of sterilized water. The statistical analysis of the values obtained was done by SPSS logistics software.Results: The different values of concentration of serum with absorption showed a linear relationship which verifies Beer–Lambert's law. With an increase in the concentration of protein in chick serum, the absorption also increases, which gives a range of concentration of protein at which ampicillin and chloramphenicol act.Conclusion: The rise and fall in the absorbance rate of proteins after addition of different antibiotics represent the increase and decrease in the concentration of proteins, respectively. This shows that every antibiotic acts at a particular concentration on the protein of the serum. Therefore, proper doses of antibiotics are recommended by the doctors

The serum proteome of Atlantic salmon, Salmo salar, during pancreas disease (PD) following infection with salmonid alphavirus subtype 3 (SAV3)

Salmonid alphavirus is the aetological agent of pancreas disease (PD) in marine Atlantic salmon, Salmo salar, and rainbow trout, Oncorhynchus mykiss, with most outbreaks in Norway caused by SAV subtype 3 (SAV3). This atypical alphavirus is transmitted horizontally causing a significant economic impact on the aquaculture industry. This histopathological and proteomic study, using an established cohabitational experimental model, investigated the correlation between tissue damage during PD and a number of serum proteins associated with these pathologies in Atlantic salmon. The proteins were identified by two-dimensional electrophoresis, trypsin digest and peptide MS/MS fingerprinting. A number of humoral components of immunity which may act as biomarkers of the disease were also identified. For example, creatine kinase, enolase and malate dehydrogenase serum concentrations were shown to correlate with pathology during PD. In contrast, hemopexin, transferrin, and apolipoprotein, amongst others, altered during later stages of the disease and did not correlate with tissue pathologies. This approach has given new insight into not only PD but also fish disease as a whole, by characterisation of the protein response to infection, through pathological processes to tissue recovery. Biological significance: Salmonid alphavirus causes pancreas disease (PD) in Atlantic salmon, Salmo salar, and has a major economic impact on the aquaculture industry. A proteomic investigation of the change to the serum proteome during PD has been made with an established experimental model of the disease. Serum proteins were identified by two-dimensional electrophoresis, trypsin digest and peptide MS/MS fingerprinting with 72 protein spots being shown to alter significantly over the 12 week period of the infection. The concentrations of certain proteins in serum such as creatine kinase, enolase and malate dehydrogenase were shown to correlate with tissue pathology while other proteins such as hemopexin, transferrin, and apolipoprotein, altered in concentration during later stages of the disease and did not correlate with tissue pathologies. The protein response to infection may be used to monitor disease progression and enhance understanding of the pathology of PD

Binding of bromocresol green and bromocresol purple to albumin in hemodialysis patients

BACKGROUND: Colorimetric albumin assays based on binding to bromocresol purple (BCP) and bromocresol green (BCG) yield different results in chronic kidney disease. Altered dye binding of carbamylated albumin has been suggested as a cause. In the present study, a detailed analysis was carried out in which uremic toxins, acute phase proteins and Kt/V, a parameter describing hemodialysis efficiency, were compared with colorimetrically assayed (BCP and BCG) serum albumin. METHODS: Albumin was assayed using immunonephelometry on a BN II nephelometer and colorimetrically based on, respectively, BCP and BCG on a Modular P analyzer. Uremic toxins were assessed using high-performance liquid chromatography. Acute phase proteins (C-reactive protein and α1-acid glycoprotein) and plasma protein α2-macroglobulin were assayed nephelometrically. In parallel, Kt/V was calculated. RESULTS: Sixty-two serum specimens originating from hemodialysis patients were analyzed. Among the uremic toxins investigated, total para-cresyl sulfate (PCS) showed a significant positive correlation with the BCP/BCG ratio. The serum α1-acid glycoprotein concentration correlated negatively with the BCP/BCG ratio. The BCP/BCG ratio showed also a negative correlation with Kt/V. CONCLUSIONS: In renal insufficiency, the BCP/BCG ratio of serum albumin is affected by multiple factors: next to carbamylation, uremic toxins (total PCS) and α1-acid glycoprotein also play a role

Competitive adsorption of plasma proteins at solid—liquid interfaces

The competitive adsorption of human serum albumin (HSA), human immuno-γ-globulin (HIgG) and human fibrinogen (HFb) onto polystyrene (PS) at 20° C and a pH of 7.35 (phosphate-buffered saline) was studied. Protein adsorption was studied using enzyme immunoassay. The results obtained with the immunoassay were compared with those obtained using radiolabelled proteins. Recent studies revealed that the adsorption behaviour of radiolabelled proteins onto surfaces differs from that of the non-labelled proteins, which may lead to misinterpretation of adsorption data. Differences in the adsorption behaviour of the labelled proteins as compared to non-labelled proteins can possibly be explained by the formation of modified proteins during the labelling procedure as shown by ion-exchange high-performance liquid chromatography (HPLC). The competitive adsorption of HSA, HIgG and HFb onto a PS latex was studied by measuring the depletion of proteins in solution. The decrease in protein concentration in solution was determined by HPLC techniques. A strong preferential adsorption of HFb was observed with maximum adsorption values of 0.6 μg/cm2

Serum C-reactive protein concentrations in dogs with idiopathic epilepsy

Inflammatory reactions in dogs are associated with systemic changes in serum, called the acute phase response; changes in the concentration of acute phase proteins in the serum take place. C-reactive protein (CRP) is a positive acute phase protein, which increases during inflammation. The role of inflammation in epilepsy remains unclear. In this study, the inflammatory response in dogs with idiopathic epilepsy (1E) was investigated. The aims of the study were: 1. to measure serum CRP concentrations in dogs with IE and in healthy dogs, 2. to measure serum CRP concentrations in dogs with acute cluster seizures and in dogs with isolated seizures and 3. to observe the evolution of serum CRP concentrations in time after the last seizure. This study showed no significant differences in serum CRP concentrations between dogs with IE (7.8 mg/I) and dogs of the control group (8.3 mg/I). Furthermore, the results showed higher mean serum CRP concentrations in dogs with IE exhibiting cluster seizures (11,8 mg/I) than in dogs with isolated seizures (5.7 mg/I). However, these results were not statistically significant (P = 0,077). Finally, no statistically significant decrease in serum CRP concentrations was seen with time after the last epileptic seizure in dogs with IE (P = 0,077)

Variation in blood serum proteins and association with somatic cell count in dairy cattle from multi-breed herds

Blood serum proteins are significant indicators of animal health. Nevertheless, several factors should be considered to appropriately interpret their concentrations in blood. Therefore, the objectives of this study were (1) to assess the effect of herd productivity, breed, age and stage of lactation on serum proteins and (2) to investigate association between serum proteins and somatic cell count (SCC) in dairy cattle. Milk and blood samples were collected from 1508 cows of six different breeds (Holstein Friesian, Brown Swiss, Jersey, Simmental, Rendena and Alpine Grey) that were housed in 41 multi-breed herds. Milk samples were analyzed for composition and SCC, while blood samples were analyzed for serum proteins (i.e.Total protein, albumin, globulin and albumin-To-globulin ratio (A : G)). Herds were classified as low or high production, according to the cow's average daily milk energy yield adjusted for breed, days in milk (DIM) and parity. Data were analyzed using a linear mixed model that included the fixed effects of DIM, parity, SCS, breed, herd productivity and the random effect of the Herd-Test date within productivity level. Cows in high producing herds (characterized also by greater use of concentrates in the diet) had greater serum albumin concentrations. Breed differences were reported for all traits, highlighting a possible genetic mechanism. The specialized breed Jersey and the two dual-purpose local breeds (Alpine Grey and Rendena) had the lowest globulin concentration and greatest A : G. Changes in serum proteins were observed through lactation. Total protein reached the highest concentration during the 4th month of lactation. Blood albumin increased with DIM following a quadratic pattern, while globulin decreased linearly. As a consequence, A : G increased linearly during lactation. Older cows had greater total protein and globulin concentrations, while albumin concentration seemed to be not particularly affected by age. A linear relationship between serum proteins and SCS was observed. High milk SCS was associated with greater total protein and globulin concentrations in blood. The rise in globulin concentration, together with a decrease in albumin concentrations, resulted in a decline in A : G as SCS of milk increased. In conclusion, such non-genetic factors must be considered to appropriately interpret serum proteins as potential animal welfare indicator and their evaluation represents an important first-step for future analysis based on the integration of metabolomics, genetic and genomic information for improving the robustness of dairy cows