This prospective follow-up study of 422 19-yr-old subjects born very preterm in The Netherlands was performed to determine whether intrauterine growth retardation (IUGR) predisposes to abnormal GFR and microalbuminuria in adolescents. GFR (ml/min per 1.73 m2) was estimated using the Cockcroft-Gault equation, and albumin-creatinine ratio (mg/mmol) was calculated in a cohort of 19-yr-old subjects born very preterm (gestational age <32 wk) in 1983. Birth weights were adjusted for gestational age and expressed as standard deviation scores (sds) as a measure of IUGR. All subjects had normal renal function. Birth weight (sds) was associated negatively with serum creatinine concentration (micromol/L) (beta = -1.0 micromol/L, 95% confidence interval [CI]: -1.9 to -0.2), positively with GFR (beta = 3.0, 95% CI: 1.7 to 4.2), and negatively with the logarithm of albumin-creatinine ratio (beta = -0.05, 95% CI: -0.09 to -0.01) in young adults born very preterm. IUGR is associated with unfavorable renal functions at young adult age in subjects born very premature. These data suggest that intrauterine growth-retarded subjects born very premature have an increased risk to develop progressive renal failure in later life.

OBJECTIVE

The neurotoxic effects of cyclosporine therapy are well known but poorly understood. Imaging studies typically show subcortical edema predominantly affecting the posterior regions of the brain. We sought to determine the causes for these findings by comparing radiographic data with various clinical parameters.

METHODS

In a 3-year period, 16 patients with neurologic findings attributed to cyclosporine therapy were examined with CT, MR imaging, or both. In most cases, imaging was performed both at the onset of the neurologic syndrome and after it had resolved. The radiographic findings were evaluated with respect to lesion location and changes over time. Various clinical and laboratory data obtained throughout the patients' hospital course were also reviewed, including cyclosporine levels, blood pressure values, hematologic data, and serum levels of cholesterol, magnesium, creatinine, and albumin.

RESULTS

The only major factor associated with the neurotoxic effects of cyclosporine in all patients was systemic hypertension. Microangiopathic hemolytic anemia, thrombocytopenia, and hypoalbuminemia were also common, and patients usually displayed signs of sympathetic overactivation. The onset of neurologic symptoms was unrelated to serum levels of creatinine, magnesium, cholesterol, or cyclosporine. The clinical and radiographic findings of these patients were identical to those previously reported in patients with hypertensive encephalopathy. Findings resolved in all but one patient after reduction of blood pressure, with or without reduction in cyclosporine dose. In four patients, intracranial hemorrhages occurred during the hypertensive episode, resulting in one fatality.

CONCLUSIONS

The clinical and radiologic findings in patients showing the neurotoxic effects of cyclosporine appear to be identical to those with hypertensive encephalopathy. Other associated factors, such as cyclosporine-induced vasculopathy or hypoalbuminemia may also play a role in the condition, and intracranial hemorrhage may occur owing to associated thrombocytopenia. Symptoms generally resolve after reduction of blood pressure, and follow-up is usually unnecessary in uncomplicated cases.

The second National Health and Nutrition Examination Survey, 1976 to 1980, incorporated medical history, physical examination, anthropometric measurements, dietary information (24-hour recall and food frequency), laboratory tests, and radiographs. In linear regressions of adjusted data from 2,695 children aged 7 years and younger, 91% of the variance in height, 72% of the variance in weight, and 58% of the variance in chest circumference were explained by six variables: age, race, sex, blood lead level, total calories or protein, and hematocrit or transferrin saturation level. Variables that did not significantly improve the models predicting growth included family income, degree of urbanization, serum albumin, copper, iron, and zinc levels, dietary carbohydrate, fat, calcium, potassium, phosphorus, vitamin A, vitamin C, niacin, riboflavin, and thiamine. The highly significant correlation of blood lead level with growth does not contradict the established association of childhood deprivation with increased lead exposure and with nutritional deficiences known to enhance lead absorption. Blood lead level may also represent a composite marker for unidentified genetic, ethnic, environmental, and sociocultural variables, other than race, sex, and nutrition, that affect growth. However, the correlation of stature, particularly height, with blood lead levels in the range of 5 to 35 micrograms/dL is so statistically significant that it merits investigation in other surveys and consideration of the multiple biologic mechanisms by which low-level lead exposure could impair the growth of children.

This work characterizes the thermodynamic processes involved in the binding and separation of (R)- and (S)-warfarin on a high-performance human serum albumin (HSA) column. Frontal analysis was used to determine the strength and degree of binding for each enantiomer. (R)- and (S)-warfarin were found to bind at the same region on HSA; however, (R)-warfarin had a larger number of column binding sites. The number of binding sites for both enantiomers showed a slight increase with temperature. The total changes in free energy for (R)- and (S)-warfarin binding were similar at 37 degrees C, but the contribution due to entropy was greater for the R-enantiomer. These results suggested that (R)-warfarin was interacting mainly with the binding site interior, while (S)-warfarin interacted more with the site's outer surface. This model was confirmed by examining the retention of (R)- and (S)-warfarin on the HSA column under various pH, ionic strength, and organic modifier conditions. The different changes in entropy for these solutes made it possible to vary their separation by changing column temperature. Both thermodynamic properties and column binding capacities were found to be important in determining the degree of separation obtained for these compounds.

To identify individual risk factors and to establish an index of risk in biliary tract surgery, data on 16 potential predictive factors were compiled from a series of 186 biliary tract operations excluding simple cholecystectomy. Eight factors had a significant association with postoperative mortality. Linear discriminant analysis showed that serum creatinine, serum albumin and serum bilirubin levels in the week before surgery had independent significance in predicting postoperative mortality. The discriminant function derived identified a high risk group of patients and the predictive value was confirmed in an independent series of 54 biliary tract operations carried out in another surgical unit. The discriminant function derived for patients jaundiced before surgery also defined a high and low risk group and was similarly validated. Identification of high risk patients undergoing surgery for obstructive jaundice may be useful in defining a group of patients to be considered for trials of preliminary biliary drainage.

The present in vitro microperfusion study examined whether insulin affects volume absorption (Jv) in the proximal convoluted tubule (PCT). PCT were perfused with an ultrafiltrate-like solution and were bathed in a serum-like albumin solution. Addition of a physiologic concentration of 10(-10) M insulin to the bathing solution resulted in a stimulation of Jv and a more negative transepithelial potential difference (PD). There was a progressive stimulation of the lumen negative PD and Jv with higher insulin concentrations. Maximal stimulation occurred at 10(-8) M bath insulin. The insulin-induced stimulation of volume reabsorption was also observed when glucose and amino acids were removed from the luminal perfusate. Direct examination of the effect of insulin on glucose, chloride, and bicarbonate absorption demonstrated that the transport of all these solutes was stimulated by insulin. Addition of insulin to the luminal perfusate had no affect on Jv. These data show that insulin has a direct effect to stimulate Jv in the proximal tubule.

BACKGROUND

Although differences between black and white persons in hemoglobin A(1c) (HbA(1c)) values are well established, recent studies suggest that this might not reflect differences in glycemia.

OBJECTIVE

To investigate racial disparities in glycemic markers, including those that reflect biological processes independent of hemoglobin glycation and erythrocyte turnover.

METHODS

Cross-sectional.

METHODS

Community-based.

METHODS

1376 nondiabetic and 343 diabetic adults in a substudy of the Atherosclerosis Risk in Communities Study.

METHODS

Hemoglobin A(1c), fasting glucose, glycated albumin, fructosamine, and 1,5-anhydroglucitol levels.

RESULTS

Among persons with and without diabetes, black persons had significantly higher HbA(1c), glycated albumin, and fructosamine levels than white persons before and after adjustment for covariates and fasting glucose concentration. Serum 1,5-anhydroglucitol levels, which are reduced in the setting of hyperglycemia-induced glycosuria, were lower in black persons than in white persons, although this difference was statistically significant only in nondiabetic adults.

CONCLUSIONS

The design was cross-sectional, a limited number of participants with a history of diabetes was included, and the study did not include integrated measures of circulating nonfasting glycemia.

CONCLUSIONS

Differences between black and white persons in glycated albumin, fructosamine, and 1,5-anhydroglucitol levels parallel differences between these groups in HbA(1c) values. Racial differences in hemoglobin glycation and erythrocyte turnover cannot explain racial disparities in these serum markers. The possibility that black persons have systematically higher levels of nonfasting glycemia warrants further study.

BACKGROUND

National Institutes of Health, National Institute of Diabetes and Digestive and Kidney Diseases.

The diffusion of macromolecules introduced into the cytoplasm of human fibroblasts by erythrocyte-mediated microinjection was measured by the fluorescence recovery after photobleaching technique. The apparent diffusion coefficients for fluorescein-labeled IgG and fluorescein-labeled bovine serum albumin were approximately 10(-8) cm2/sec at 22 degrees C, consistent with the kinetics of spreading of the fluorescent probe following microinjection and approximately 1/70 the values in aqueous buffer. The diffusion of labeled bovine serum albumin was shown to be strongly dependent on temperature and, in fact, similar to that expected in a 61% aqueous sucrose solution. However, the marked reduction in diffusion at 5 degrees C could be fully reversed by incubation with 0.1 mM colchicine. These findings suggest that cytoplasmic diffusion rates are reduced relative to rates in aqueous media as a result of increased aqueous phase viscosity or the impedence provided by structural elements. Several simple models to account for the data are presented.

Immunocytochemical techniques have localized a large protein which is an intrinsic membrane component of isolated frog rod outer segments (ROS). This large protein whose apparent mol wt is 290,000 daltons comprises about 1--3% of the ROS membrane mass. Its molar ratio to opsin is between 1:300 and 1:900. Adequate immune responses were obtained with less than 30 microgram (100 pmol) of antigen per rabbit. Antibodies to the large protein were used for its localization on thin sections of frog retina embedded in glutaraldehyde cross-linked bovine serum albumin (BSA). Specifically bound antibodies were detected by an indirect sequence with ferritin-conjugated antibodies. This technique detected the protein which is represented by 1,000--3,000 molecules per disk. This indicates that the procedure is sufficiently sensitive for analysis of membrane components in low molar proportions. The large protein was specifically localized to the incisures of ROS disks which divide the disks into lobes and to the disk margin. Thus, opsin is mobile within the membrane of the disk while the large protein is apparently constrained to the disk edges. This finding raises the possibility that special functions are also localized ot his unusual region of high curvature, and that collisions of bleached opsin with these edges are physiologically important in couter segment function.

Protein-bound and unbound fatty acids can be efficiently separated at 0 degree C using a hydrophobic column-packing material (Lipidex 1000) similar to the separation of protein-bound and unbound steroids (E. Dahlberg, M. Snochowski, and J.-A. Gustafsson (1980) Anal Biochem. 106, 380-388). Protein-bound fatty acids are also removed by Lipidex 1000 when treatment is performed at 37 degrees C. Lipidex 1000 does not exhibit binding properties for soluble proteins at 0 and 37 degrees C, in contrast to dextran-coated charcoal. Lipidex 1000 appeared to be useful for the delipidation of protein samples at 37 degrees C and for a radiochemical assay of fatty acid-binding by microgram amounts of protein at 0 degree C. With this assay we obtained results on palmitate binding to serum albumin similar to those reported on the basis of equilibrium dialysis. Delipidated proteins from dealbuminized rat liver cytosol maximally bind about 4 nmol palmitate/mg protein.

Histologic variants of idiopathic focal segmental glomerulosclerosis (FSGS) may have prognostic value. A recent working classification system has distinguished five FSGS variants. We evaluated a cohort of adult patients with biopsy-proven FSGS diagnosed between March 1982 and July 2001 to determine if subtypes were associated with renal outcome. Renal biopsies were reviewed by two pathologists. Demographic and clinical data were obtained from charts. Outcomes were partial and complete remission of the nephrotic syndrome, and renal failure. The frequency of FSGS variants was: 3% cellular (N=6), 11% collapsing (N=22), 17% tip lesion (N=34), 26% perihilar (N=52), and 42% not otherwise specified (NOS) (N=83). Collapsing FSGS affected younger and more often black patients. Black race was uncommon in tip variant. Collapsing and tip variants had higher proteinuria and lower serum albumin than perihilar and NOS variants. Better renal function and less severe tubulointerstitial injury were observed in patients with tip variant. These patients were more likely to receive steroids and more often achieved complete remission (50%). After a median follow-up of 1.8 years, 23% of patients were on dialysis and 28% had renal failure. Collapsing FSGS had worse 1-year (74%) and 3-year (33%) renal survival compared to other variants (overall cohort renal survival at 1 and 3 years: 86 and 67%). Different histologic variants of FSGS have substantial differences in clinical features at the time of biopsy diagnosis and substantial differences in renal outcomes.

Murine thioglycollate-elicited peritoneal macrophages were cultured in the presence of a variety of fatty acids added as complexes with bovine serum albumin. All fatty acids tested were taken up readily by the cells and both neutral and phospholipid fractions were enriched with the fatty acid provided in the medium. This generated a range of cells enriched in saturated, monounsaturated or polyunsaturated fatty acids, including n-3 acids of fish oil origin. Saturated fatty acid enrichment enhanced macrophage adhesion to both tissue culture plastic and bacterial plastic compared with enrichment with polyunsaturated fatty acids. Macrophages enriched with the saturated fatty acids myristate or palmitate showed decreases of 28% and 21% respectively in their ability to phagocytose unopsonized zymosan particles. Those enriched with polyunsaturated fatty acids showed 25-55% enhancement of phagocytic capacity. The greatest rate of uptake was with arachidonate-enriched cells. Phagocytic rate was highly correlated with the saturated/unsaturated fatty acid ratio, percentage of polyunsaturated fatty acid and index of unsaturation, except for macrophages enriched with fish-oil-derived fatty acids; they showed lower phagocytic activity than expected on the basis of their degree of unsaturation. These results suggest that membrane fluidity is important in determining macrophage adhesion and phagocytic activity. However, in the case of phagocytosis, this effect may be partially overcome if the cells are enriched with fish-oil-derived fatty acids. Thus it may be possible to modulate the activity of cells of the immune system, and so an immune response, by dietary lipid manipulation.

The surface charges on biodegradable albumin nanoparticles were introduced by covalent coupling different primary amines to examine their influence on phagocytosis by macrophages under in vitro conditions. Albumin particles with a zeta potential close to zero showed a reduced phagocytic uptake in comparison with charged particles, especially nanoparticles with a positive zeta potential. The phagocytic uptake in the present study was examined using an established cell culture model based on primary mouse peritoneal macrophages and a human hematopoietic monocytic cell line (U-937) treated with phorbol-12-myristic-13-acetate to induce cell differentiation. The influence of opsonins on in vitro phagocytosis experiments was characterized using carriers pre-treated with human serum. In the presence of human serum the phagocytic activity of U-937 cells was found to be similar to primary mouse macrophages without serum. In contrast to peritoneal macrophages, U-937 cells showed no phagocytic activity in the absence of serum. In particular, only the C3b- complement deposition on the particle surface seems to promote the phagocytic process. The in vivo distribution of albumin carriers in rats was investigated using magnetic resonance imaging (MRI). No differences in blood circulation times and organ accumulation between different nanoparticle preparations with positive, neutral and negative surface charges could be observed in rats, suggesting that the in vivo fate of albumin nanoparticles is significantly influenced by factors not reflected in the in vitro cell culture models.

We have recently demonstrated that induction of Th2 responses by Schistosoma mansoni egg Ag is largely due to carbohydrates on the Ag functioning as adjuvants. Lacto-N-fucopentaose III (LNFPIII), a polylactosamine sugar, is the predominant carbohydrate found in S. mansoni egg Ag. Therefore, using neoglycoprotein, we investigated whether LNFPIII induces in vivo Th2 response and functions as an adjuvant. Following intranasal immunization with LNFPIII linked to human serum albumin (HSA) (HSA-LNFPIII), BALB/c mice mounted a strong Th2 response and produced significantly higher levels of total IgE as well as HSA-specific IgG, IgG1, and IgE. HSA-LNFPIII was over 1000-fold more potent in inducing Ab production as compared with HSA alone. Although LNFPIII itself did not function as an epitope for either IgG or IgE, its conjugation with protein was essential for the adjuvant activity. Moreover, fucose residue on LNFPIII was crucial for induction of Ab production. Nasal lymphocytes from mice immunized with HSA-LNFPIII produced IL-4, IL-5, and IL-10, but not IFN-gamma following in vitro stimulation with HSA or HSA-LNFPIII. In addition, these activated nasal lymphocytes also showed a significant increase of B7-2 expression on B220-positive cells. Furthermore, not only intranasal but also both i.p. and s.c. immunization with HSA-LNFPIII induced significant production of HSA-specific Abs compared with the immunization with HSA alone, suggesting that the activity of LNFPIII was not restricted on particular route of immunization. These results demonstrate that Lewis type carbohydrate LNFPIII can function as an adjuvant by their ability to induce a Th2 response.

BACKGROUND

The development of diabetic nephropathy has been linked to the release of vasoactive hormones and growth factors. Currently the role of inflammatory cytokines in this pathogenic process is not clear.

METHODS

We utilized the microdialysis technique to monitor early changes in tumor necrosis-alpha (TNF-alpha) levels in the renal interstitial fluid and urine of conscious Sprague-Dawley rats (N = 8) before and after induction of diabetes with streptozotocin (STZ). Measurement of the urinary albumin excretion (UAE) was utilized to monitor the development and progression of diabetic nephropathy.

RESULTS

UAE increased from 0.56 +/- 0.20 microg/min to 8.14 +/- 2.98 microg/min 17 days after induction of diabetes (P = 0.01). Renal interstitial fluid TNF-alpha increased from 11.96 +/- 5.32 pg/mL at baseline to 45.02 +/- 11.69 pg/mL 5 days after induction of diabetes (P = 0.03). Renal interstitial fluid TNF-alpha levels remained elevated throughout the remainder of the study period. Urinary TNF-alpha also increased significantly compared to baseline 3 days after induction of diabetes (294.18 +/- 36.94 pg/mL vs. 16.05 +/- 6.07 pg/mL, P < 0.002). There was a second significant rise in urinary TNF-alpha concentration to 638.16 +/- 36.94 pg/mL 21 days after induction of diabetes (P < 0.001). Serum TNF-alpha levels were undetectable before STZ injection and remained undetectable by the end of the study. Urinary and renal interstitial fluid TNF-alpha in the control rats (N = 5) did not change throughout the study.

CONCLUSIONS

We found an early rise in renal TNF-alpha levels after induction of diabetes with STZ, which precedes the rise in UAE by about 2 weeks. These findings suggest a possible contribution of TNF-alpha in the complicated pathogenic process resulting in microalbuminuria in diabetes.

BACKGROUND

As the population ages, the number of injured elderly is increasing. We sought to determine if low skeletal muscle mass adversely affected outcome in elderly patients following trauma.

METHODS

Patients ≥ 65 years of age with an admission abdominal computed tomography scan and requiring intensive care unit (ICU) stay at a Level I trauma center in 2009-2010 were reviewed. Muscle cross-sectional area at the 3rd lumbar vertebra was quantified and muscle index, a normalized measure of muscle mass, was calculated and related to clinical parameters including ventilator-free days, ICU-free days, and mortality. Using previously established sex-specific, muscle index cut-points, patients were then categorized as sarcopenic or non-sarcopenic and differences in clinical outcomes between these two groups were also compared. We also examined muscle index as a continuous variable relative to the same clinical outcomes.

RESULTS

There were 149 severely injured elderly patients (median age 79 years) enrolled in this study of which 71% were sarcopenic. Of the patients who were sarcopenic, 9% were underweight, 44% normal weight, and 47% overweight/obese as per body mass index (BMI) classifications. The overall mortality rate was 27% and univariate analysis demonstrated higher mortality among those who were sarcopenic (32% vs. 14%, P = 0.018). After controlling for age, sex, and injury severity, multiple logistic regression demonstrated that increased muscle index was significantly associated with decreased mortality (OR per unit muscle index = 0.93, 95% CI: 0.875-0.997, P = 0.025). In addition, multivariate linear regression showed that sarcopenia, but not muscle index, was associated with decreased ventilator-free (P = 0.004) and ICU-free days (P = 0.002). Neither BMI, serum albumin nor total adipose tissue on admission were indicative of survival, ventilator-free or ICU-free days.

CONCLUSIONS

Sarcopenia is highly prevalent in the elderly population with traumatic injuries. Traditional measures of nutritional assessment, such as BMI and serum albumin, do not accurately predict outcome in the injured elderly. Sarcopenia, however, represents a potential new predictor for mortality, discharge disposition, and ICU utilization. Measurement of muscularity allows for the early identification of at-risk patients who may benefit from aggressive and multidisciplinary nutritional and rehabilitative strategies.

Four methods for measuring diffusion coefficients were compared on a microfabricated fluidic device using rhodamine 6G as the analyte. The measurements were made using a static imaging method and three dynamic methods-stopped flow, varying the applied potential (E-field method), and varying the detection length (length method). Under conditions where analyte-wall interactions (adsorption) are minimized, e.g. in a 50/50 (v/v) methanol/aqueous buffer, the stopped flow (2.71+/-0.09x10(-6) cm(2) s(-1)), E-field (2.684+/-0.005x10(-6) cm(2) s(-1)) and the static imaging (2.69+/-0.02x10(-6) cm(2) s(-1)) measurements were all within experimental error of one another and previously reported values. Under 100% aqueous conditions, however, the diffusion coefficient measured dynamically was 11% larger than that measured statically. Diffusion coefficients for rhodamine B, fluorescein, 2',7'dichloro-fluorescein (DCF), rhodamine 6G, tetramethylrhodamine labeled glutamic acid and isoleucine, and fluorescein conjugated bovine serum albumin and ovalbumin were also measured using the static imaging method.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that destroys patient memory and cognition, communication ability with the social environment and the ability to carry out daily activities. Despite extensive research into the pathogenesis of AD, a neuroprotective treatment - particularly for the early stages of disease - remains unavailable for clinical use. In this review, we advance the suggestion that lipoic acid (LA) may fulfil this therapeutic need. A naturally occurring cofactor for the mitochondrial enzymes pyruvate dehydrogenase and alpha-ketoglutarate dehydrogenase, LA has been shown to have a variety of properties which can interfere with the pathogenesis or progression of AD. For example, LA increases acetylcholine (ACh) production by activation of choline acetyltransferase and increases glucose uptake, thus supplying more acetyl-CoA for the production of ACh. LA chelates redox-active transition metals, thus inhibiting the formation of hydroxyl radicals and also scavenges reactive oxygen species (ROS), thereby increasing the levels of reduced glutathione. In addition, LA down-regulates the expression of redox-sensitive pro-inflammatory proteins including TNF and inducible nitric oxide synthase. Furthermore, LA can scavenge lipid peroxidation products such as hydroxynonenal and acrolein. In human plasma, LA exists in an equilibrium of free and plasma protein bound form. Up to 150 muM, it is bound completely, most likely binding to high affinity fatty acid sites on human serum albumin, suggesting that one large dose rather than continuous low doses (as provided by "slow release" LA) will be beneficial for delivery of LA to the brain. Evidence for a clinical benefit for LA in dementia is yet limited. There are only two published studies, in which 600 mg LA was given daily to 43 patients with AD (receiving a standard treatment with choline-esterase inhibitors) in an open-label study over an observation period of up to 48 months. Whereas the improvement in patients with moderate dementia was not significant, the disease progressed extremely slowly (change in ADAScog: 1.2 points=year, MMSE: -0.6 points=year) in patients with mild dementia (ADAScog<15). Data from cell culture and animal models suggest that LA could be combined with nutraceuticals such as curcumin, (-)-epigallocatechin gallate (from green tea) and docosahexaenoic acid (from fish oil) to synergistically decrease oxidative stress, inflammation, Abeta levels and Abeta plaque load and thus provide a combined benefit in the treatment of AD.

Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality worldwide and ranks second in China. The prognosis of HCC remains dismal mainly because of its late diagnosis, especially in patients with coexisting chronic liver diseases. To identify serum biomarkers for HCC, sera from 20 healthy volunteers, 20 hepatitis B virus (HBV) infected patients and 20 HCC patients were selected for screening study and same number of sera into the same three groups were used for validation study. A strategy including sonication, albumin and immunoglobulin G (IgG) depletion and desalting was optimized for screening differentially expressed proteins of low abundance in serum. By 2-DE image analysis and MALDI-TOF-MS/MS identification, eight proteins including heat-shock protein 27 (HSP27), alpha-fetoprotein (AFP), alpha-1 antitrypsin, clusterin, caeruloplasmin, haptoglobin alpha2 chain, tranferrin and transthyretin were found significantly changed among the healthy, HBV and HCC groups. Further validation study by Western blot showed the detection of HSP27 in 90% HCC sera and two HBV sera, but in none of normal sera. Thus, 2-DE based serum proteome analysis can be useful in the screening of serum biomarkers for HCC and HSP27 could aid in the diagnosis of HCC though further validation is needed.

We have studied the effects of macromolecular crowding on protein folding kinetics by studying the oxidative refolding of hen lysozyme in the absence and presence of high concentrations of bovine serum albumin and Ficoll 70. The heterogeneity characteristic of the lysozyme refolding process is preserved under crowded conditions. This, together with the observation that the refolding intermediates that accumulate to significant levels are very similar in the absence and presence of Ficoll, suggests that crowding does not alter substantially the energetics of the protein folding reaction. However, the presence of high concentrations of macromolecules results in the acceleration of the fast track of the refolding process whereas the slow track is substantially retarded. The results can be explained by preferential excluded volume stabilization of compact states relative to more unfolded states, and suggest that, relative to dilute solutions, the rates of many protein folding processes are likely to be altered under conditions that more closely resemble the intracellular environment.

Glycosylation is one the most common post-translational modifications (PTM) and glycoproteins play fundamental roles in a diversity of biological processes. The development of an analytical approach to the study of variation of glycosylation patterns in serum samples has been hindered by the structural heterogeneity of this post-translational modification and the complexity of serum proteome. We have used the ability of different lectins to recognize specific glycosylation motifs to develop a specific affinity system that can achieve a comprehensive capture of serum glycoproteins. In a preliminary investigation, we evaluated the ability of five commonly used immobilized lectins to capture glycoproteins from human serum. SDS-PAGE analysis showed each lectin was able to enrich a subset of the serum glycoproteome and overlaps in lectin specificity were indeed observed. Based on these results and with the goal of studying the extent of the human serum glycoproteome, we then developed a multi-lectin affinity column containing Concanavalin A (Con A), Wheat germ and Jacalin lectin. The selection of lectins was also based on the known N-linked and O-linked glycan structures that are considered representative of the serum proteome. We then demonstrated that the capture of glycoproteins was specific, efficient and reproducible with this multi-lectin column. The results obtained with this affinity step indicated that about 10% of human serum proteins are glycosylated (weight/weight) and, after removal of six high abundance proteins, including albumin, at least 50% of the remaining proteins were glycosylated. We then evaluated the use of this affinity column to monitor changes in the pattern of glycosylation in serum samples by a controlled, stepwise release of the captured proteins from the multi-lectin affinity column with specific displacers.

Disruption of the blood-brain barrier (BBB) is a characteristic finding in common neurological disorders. Human data suggest BBB disruption may underlie cerebral dysfunction. Animal experiments show the development of epileptiform activity following BBB breakdown. In the present study we investigated the neurophysiological, structural and functional consequences of BBB disruption. Adult rats underwent focal BBB disruption in the rat sensory-motor cortex using the bile salt sodium deoxycholate (DOC). Magnetic resonance imaging in-vivo showed an early BBB disruption with delayed reduction in cortical volume. This was associated with a reduced number of neurons and an increased number of astrocytes. In-vitro experiments showed that the threshold for spreading depression and the propagation velocity of the evoked epileptic potentials were increased 1 month after treatment. Furthermore, animals' motor functions deteriorated during the first few weeks following BBB disruption. Treatment with serum albumin resulted in a similar cell loss confirming that the effect of DOC was due to opening of the BBB. Our findings suggest that delayed neurodegeneration and functional impairment occur following the development of the epileptic focus in the BBB-permeable cerebral cortex.

The effects of various experimental conditions on in vitro drug permeability to Caco-2 monolayers were investigated to determine the optimized conditions for the prediction of intestinal drug absorption. Concerning the pH of the transport medium in the Caco-2 study, two different pH values, 6.0 and 7.4, were tested for the apical medium with the pH of the basolateral medium fixed to 7.4. The change in the apical pH showed pronounced effects on the permeability of both passively and actively transported drugs. It was found that the transport study under the condition of an apical pH value of 6.0 showed a better prediction of in vivo drug absorption in human. The appropriate conditions for determining the permeability of poorly soluble drugs were also examined. First, the effects of bile acids, surfactant and some agents used for solubilizing drugs on the permeability and transepithelial electrical resistance (TEER) of Caco-2 monolayers were investigated. Taurocholic and cholic acid showed no effects on the permeability of 3H-Dexamethasone (DEX) and TEER at 10 mM concentration, suggesting the possibility of use in the Caco-2 study. Polyethyleneglycol-400 and dimethylsulfoxide reduced the permeability of DEX concentration dependently, whereas ethanol induced no significant changes in the permeability. Furthermore, it was demonstrated that the addition of plasma protein (bovine serum albumin) to the basolateral medium apparently facilitated the transport of poorly soluble drugs with high lipophilicity across Caco-2 monolayers. These findings clearly suggest the importance of considering the physiological conditions of in vivo drug absorption in optimizing the in vitro experimental conditions for transport study using Caco-2 cells, in order to obtain a satisfactory in vitro-in vivo correlation.

Approximately 10.2 million persons in the United States sometimes or often do not have enough food to eat, a condition known as food insufficiency. Using cross-sectional data from the Third National Health and Nutrition Examination Survey (NHANES III), we examined whether dietary intakes and serum nutrients differed between adults from food-insufficient families (FIF) and adults from food-sufficient families (FSF). Results from analyses, stratified by age group and adjusted for family income and other important covariates, revealed several significant findings (P < 0.05). Compared with their food-sufficient counterparts, younger adults (aged 20-59 y) from FIF had lower intakes of calcium and were more likely to have calcium and vitamin E intakes below 50% of the recommended amounts on a given day. Younger adults from FIF also reported lower 1-mo frequency of consumption of milk/milk products, fruits/fruit juices and vegetables. In addition, younger adults from FIF had lower serum concentrations of total cholesterol, vitamin A and three carotenoids (alpha-carotene, beta-cryptoxanthin and lutein/zeaxanthin). Older adults (aged>> or =60 y) from FIF had lower intakes of energy, vitamin B-6, magnesium, iron and zinc and were more likely to have iron and zinc intakes below 50% of the recommended amount on a given day. Older adults from FIF also had lower serum concentrations of high-density lipoprotein cholesterol, albumin, vitamin A, beta-cryptoxanthin and vitamin E. Both younger and older adults from FIF were more likely to have very low serum albumin (<35 g/L) than were adults from FSF. Our findings show that adults from FIF have diets that may compromise their health.