[New nanofibrous scaffold for corneal tissue engineering]. / Eine neue Nanofaser-Matrix für den kornealen Gewebeersatz.

BACKGROUND: An estimated 10 million people suffer worldwide from vision loss caused by corneal damage. For the worst cases, the only available treatment is transplantation with human donor corneal tissue. However, in numerous countries there is a considerable shortage of corneal tissue of good quality, leading to various efforts to develop tissue substitutes. The present study aims to introduce a nanofibrous scaffold of poly(glycerol sebacate) PGS as a biodegradable implant, for the corneal tissue engineering. MATERIALS AND METHODS: Nanofibrous scaffolds were produced from PGS and poly(ε-caprolactone) (PCL) by a modified electro-spinning process. The biocompatibility of the material was tested in vitro by colorimetric MTT assay on days 3, 5, and 7 to test the cell viability of human corneal endothelium cells (HCEC). To examine a potential immunological reaction of the scaffolds, samples were exposed to mononuclear cells derived from peripheral blood (PBMCs). After an incubation period of 3 days, supernatants were assayed for apoptotic assessment and immunogenic potentials by annexin V FITC//propidium iodide and flow-cytometric analysis. RESULTS: We could successfully demonstrate that cultivation of HCECs on PGS/PCL scaffolds was possible. Compared to day 3, cell density determined by microplate absorbance was significantly higher after 7 days of cultivation (p < 0.0001). According to the MTT data, none of the samples showed toxicity. Apoptotic assessments by FACS analysis showed that no composition stimulated apoptosis or activated PBMCs occurred. All the compositions were inert for native as well as activated T/B/NK cells and monocytes. It can be concluded that leukocytes and their activity was not affected by the scaffolds. CONCLUSION: A tissue-like scaffold mimicking the human stroma could be developed. The results indicate that PGS/PCL scaffolds could be considered as ideal candidates for corneal tissue engineering as they are biocompatible in contact to corneal endothelial cells and blood cells.

Removal of indigenous coliphages and enteric viruses during riverbank filtration from highly polluted river water in Delhi (India).

Emerging countries frequently afflicted by waterborne diseases require safe and cost-efficient production of drinking water, a task that is becoming more challenging as many rivers carry a high degree of pollution. A study was conducted on the banks of the Yamuna River, Delhi, India, to ascertain if riverbank filtration (RBF) can significantly improve the quality of the highly polluted surface water in terms of virus removal (coliphages, enteric viruses). Human adenoviruses and noroviruses, both present in the Yamuna River in the range of 10(5) genomes/100 mL, were undetectable after 50 m infiltration and approximately 119 days of underground passage. Indigenous somatic coliphages, used as surrogates of human pathogenic viruses, underwent approximately 5 log10 removal after only 3.8 m of RBF. The initial removal after 1 m was 3.3 log10, and the removal between 1 and 2.4 m and between 2.4 and 3.8 m was 0.7 log10 each. RBF is therefore an excellent candidate to improve the water situation in emerging countries with respect to virus removal.

Comparison of functional and morphological diagnostics in glaucoma patients and healthy subjects.

PURPOSE: To evaluate the diagnostic value of microperimetry (MP), blue-on-yellow perimetry (B/YP), confocal scanning laser ophthalmoscopy (Heidelberg Retina Tomograph, HRT, III) and optical coherence tomography (OCT) in discriminating eyes with early glaucoma from healthy subjects. MATERIAL AND METHODS: Prospective examination of 22 eyes of subjects with early primary open-angle glaucoma and 24 eyes of healthy control subjects. After a complete ophthalmological examination, B/YP, MP, OCT and HRT III were determined. Morphological and functional parameters were analysed. RESULTS: Mean sensitivity threshold values obtained with B/YP and MP did not show significant differences between glaucoma patients and the control group (p = 0.321 and p = 0.281). Retinal nerve fibre layer (RNFL) thickness was significantly decreased in patients with glaucoma with both HRT III and OCT (p = 0.018 and p < 0.001). CONCLUSIONS: While B/YP and MP had no ability to discriminate between subjects with early glaucoma and healthy subjects, RNFL thickness measured with HRT III and OCT showed a significant difference. In early primary open-angle glaucoma, morphological changes like RNFL thickness seem to occur prior to functional defects in the visual field.

[Removal of microorganisms by soil filters for bathing ponds]. / Entfernung von Mikroorganismen durch Bodenfilter für Kleinbadeteiche.

"Bathing ponds" are artificial outdoor water pools without disinfection. Whereas in conventional pools, chlorine promptly kills pathogens shed by bathers, such quick inactivation is missing in bathing ponds. We have explored the retention of indicator bacteria and viruses by a vertically operated, reed grown soil filter. After continuously running the filter with wastewater-spiked surface water, we found that the filter retains more than 99 % of the indicator organisms. It has been reported in the literature that the "spontaneous" inactivation of pathogens in water might be very variable depending on sunlight irradiation, water turbidity, etc. On the contrary, the performance of a filter like the one reported here allows filtering the water so as to reliably eliminate 90 % of the spiked microorganisms from the pool water within 24 hours.

Development and application of a one-step low cost procedure to concentrate viruses from seawater samples.

A novel and simple procedure for concentrating adenoviruses from seawater samples is described. The technique entails the adsorption of viruses to pre-flocculated skimmed milk proteins, allowing the flocs to sediment by gravity, and dissolving the separated sediment in phosphate buffer. Concentrated virus may be detected by PCR techniques following nucleic acid extraction. The method requires no specialized equipment other than that usually available in routine public health laboratories, and due to its straightforwardness it allows the processing of a larger number of water samples simultaneously. The usefulness of the method was demonstrated in concentration of virus in multiple seawater samples during a survey of adenoviruses in coastal waters.

Effects of antioxidants on glutathione levels and clinical recovery from the malnutrition syndrome kwashiorkor--a pilot study.

Kwashiorkor is a severe edematous form of malnutrition with high prevalence and lethality in many African countries, and repeatedly has been reported to be associated with oxidative stress. The therapy of kwashiorkor is still ineffective. In this pilot study, we tested the hypothesis that oral application of thiol-containing antioxidants increases glutathione status and is beneficial for the clinical recovery of kwashiorkor patients. The longitudinal clinical intervention study was carried out at St Joseph's Hospital, Jirapa, Ghana. Children with severe kwashiorkor were randomly assigned to either a standard treatment (ST) receiving a therapeutic protocol based on the recommendations of the WHO or to one of three study groups receiving in addition 2 x 600 mg reduced glutathione or 2 x 50 mg alpha-lipoic acid or 2 x 100 mg N-acetylcysteine per day. Patients were followed up clinically and biochemically for 20 days and compared with 37 healthy controls. Both glutathione and alpha-lipoic acid supplementation had positive effects on survival. Also, the blood glutathione concentrations correlated positively with survival rates. Furthermore, the initial skin lesions, glutathione and total protein concentrations were found to be strong predictors of survival. The data strongly suggest that a therapy restoring the antioxidative capacity by applying cysteine equivalents in the form of glutathione and/or alpha-lipoic acid is beneficial for biochemical and clinical recovery of kwashiorkor patients.

Oxidized low-density lipoproteins bind to the scavenger receptor, CD36, of hepatic stellate cells and stimulate extracellular matrix synthesis.

Cumulating evidence suggests that oxidative stress resulting in lipid peroxidation and protein modification is involved in the pathogenesis of chronic liver injury and fibrogenesis. We investigated the effects of oxidized low-density lipoproteins (oxLDL) on collagen and fibronectin synthesis of cultured human and rat hepatic stellate cells (HSC). As shown on protein and mRNA levels, oxLDL dose-dependently stimulated the synthesis of collagen types I and III and fibronectin of cultured HSC. The effect was biphasic, with a maximum between 5 and 25 microg/mL oxLDL (c-fibronectin concentration in HSC supernatants increased 3.9-fold; collagen type I increased 4-fold). Higher oxLDL concentrations were cytotoxic. LDL modified with malondialdehyde (MDA) was not toxic, but stimulated extracellular matrix synthesis as well. As demonstrated by immunofluorescence microscopy (double staining of CD36 and iso-alpha-smooth muscle actin [iso-alpha-sm actin]), immunoblot, and reverse-transcription polymerase chain reaction (RT-PCR), respectively, cultured human HSC express the oxLDL receptor, CD36 (glycoprotein IIIb). Colocalization of CD36 and iso-alpha-sm actin on sinusoidal lining cells was further demonstrated using sections of human fibrotic liver. Preincubation of cultured human HSC with the monoclonal antibody, OKM5, known to block CD36-mediated oxLDL uptake, resulted in a reduction of the oxLDL-stimulated collagen type I synthesis by 56%. In summary, our results demonstrate that low concentrations of modified lipoproteins (oxLDL and MDA-LDL) represent fibrogenic mediators that bind to CD36 and stimulate matrix synthesis of HSC.

Identification of mediators stimulating proliferation and matrix synthesis of rat pancreatic stellate cells.

The aim of this study was to identify fibrogenic mediators stimulating activation, proliferation, and/or matrix synthesis of rat pancreatic stellate cells (PSC). PSC were isolated from the pancreas of normal Wistar rats and from rats with cerulein pancreatitis. Cell activation was demonstrated by immunofluorescence microscopy of smooth muscle alpha-actin (SMA) and real-time quantitative RT-PCR of SMA, fibronectin, and transforming growth factor (TGF)-beta(1). Proliferation was measured by bromodeoxyuridine incorporation. Matrix synthesis was demonstrated on the protein and mRNA level. Within a few days in primary culture, PSC changed their phenotype from fat-storing to SMA-positive myofibroblast-like cells expressing platelet-derived growth factor (PDGF) alpha- and PDGF beta-receptors. TGF-beta(1) and tumor necrosis factor (TNF)-alpha accelerated the change in the cells' phenotype. Addition of 50 ng/ml PDGF and 5 ng/ml basic fibroblast growth factor (bFGF) to cultured PSC significantly stimulated cell proliferation (4.37 +/- 0.49- and 2.96 +/- 0.39-fold of control). Fibronectin synthesis calculated on the basis of DNA was stimulated by 5 ng/ml bFGF (3.44 +/- 1.13-fold), 5 ng/ml TGF-beta(1) (2.46 +/- 0.89-fold), 20 ng/ml PDGF (2.27 +/- 0.68-fold), and 50 ng/ml TGF-alpha (1.87 +/- 0.19-fold). As shown by RT-PCR, PSC express predominantly the splice variant EIII-A of fibronectin. Immunofluorescence microscopy and Northern blot confirmed that in particular bFGF and TGF-beta(1) stimulated the synthesis of fibronectin and collagens type I and III. In conclusion, our data demonstrate that 1) TGF-beta(1) and TNF-alpha accelerate the change in the cell phenotype, 2) PDGF represents the most effective mitogen, and 3) bFGF, TGF-beta(1), PDGF, and, to a lesser extent, TGF-alpha stimulate extracellular matrix synthesis of cultured rat PSC.

Characterization of modified low density lipoprotein subfractions by capillary isotachophoresis.

Oxidative modification of low density lipoproteins (LDLs) is an important pathogenetic factor in atherosclerosis. The various steps in oxidative modifications of LDL can be monitored using different methodologies with varying degrees of complexity. In this study, we propose capillary isotachophoresis (CITP) as a suitable tool to detect and measure the degree of oxidation of LDL. LDL was isolated from pooled plasma of healthy volunteers by sequential ultracentrifugation, and oxidation was performed in vitro as well as in cell culture experiments. Native LDL and oxidatively modified LDL were characterized by apo B-100 fluorescence and conjugated diene formation. Samples were separated by CITP combined with sudan black B staining. To underline the inherent advantages of this approach, CITP was compared with classical lipoprotein electrophoresis using agarose gel. We demonstrate the CITP method to be highly sensitive, as changes in peak area of the separated LDL subfractions were detected after only 2 h of oxidation. The leading LDL peaks increased, while the terminating LDL peaks decreased in parallel throughout the duration of oxidation. The LDL samples, oxidized for 4-24 h, also exhibited an increased migration velocity of the fractions. In summary, we present the first study investigating LDL-subfractions separated by CITP and the alterations of these LDL-subfractions after gradual in vitro oxidation and after oxidative modification by monocyte-derived macrophages and vascular smooth muscle cells.

2nd Sino-European Congress of Laboratory Medicine--strategies for laboratory diagnostics in medicine.

During the 2nd Sino-European Congress in Shanghai in April 2001 the aim of the presentation was an overview on strategies and systems of laboratory diagnostic work. The main focus was to define the prerequisites and the different approaches in the diagnostic work in patients' treatment. In this paper, besides the area of emergency medicine with definition of vital functions and the routine work in deciding between healthy and ill individuals and in the control of therapy, the different aspects in the field of research and teaching are specified. Some aspects of workflow definition and optimization are discussed.

Platelet-derived growth factors stimulate proliferation and extracellular matrix synthesis of pancreatic stellate cells: implications in pathogenesis of pancreas fibrosis.

At present, the cell-cell interactions and molecular mechanisms of pancreas fibrogenesis are largely unknown. The purpose of this study was to investigate paracrine stimulatory loops between platelets and pancreatic stellate cells (PSC). Human PSC were obtained by outgrowth from fibrotic human pancreas. Native platelet lysate (nPL) and transiently acidified platelet lysate (aPL) were added to cultured PSC (passage 4 to 7) in the absence of serum. The synthesis of collagen types I and III and c-fibronectin (cFN) was demonstrated on protein (immunofluorescence and quantitative immunoassay) and mRNA (Northern blot) level. Using sections of human pancreas with acute pancreatitis, platelet aggregates in capillaries were demonstrated by transmission electron microscopy. nPL, and to an even greater extent aPL, significantly increased the synthesis of collagen types I and III and of c-FN (120 microl/ml aPL increased collagen type I concentration in PSC supernatants by 1.99 +/- 0.17 times and c-FN of 2.49 +/- 0.28 times, mean +/- SD, n = 3). nPL and aPL also significantly stimulated cell proliferation (increased bromodeoxyuridine (BrdU) incorporation by 6.4 +/- 0.78 times and 10 +/- 0.29 times, respectively). By preincubating aPL with transforming growth factor beta (TGFbeta)- and platelet-derived growth factor (PDGF)-neutralizing antibodies and the TGFbeta-latency associated peptide, respectively, TGFbeta1 was identified as the main mediator stimulating matrix synthesis and PDGF as the responsible mitogen. Our data demonstrate that platelets contain fibrogenic mediators that stimulate proliferation (PDGF) and matrix synthesis (TGFbeta1) of cultured PSC. We suggest that platelets and PSC cooperate in the development of pancreas fibrosis.

Serum amyloid A versus C-reactive protein in acute pancreatitis: clinical value of an alternative acute-phase reactant.

OBJECTIVES: The acute-phase reactant C-reactive protein (CRP) is currently the serum variable of choice for an early, accurate, and cost-effective severity assessment of acute pancreatitis in the daily clinical routine. Serum amyloid A (SAA) proteins comprise a family of apolipoproteins that constitute another major acute-phase reactant and thus could be a potential alternative to CRP assessment. In the present study we investigated the clinical usefulness of SAA determinations in acute pancreatitis using an automated immunoassay technique. DESIGN: Cohort study, comparing patients with complicated and mild acute pancreatitis; control groups included individuals with further abdominal disorders and healthy volunteers. SETTING: A collaborative study between the department of general surgery and the routine laboratory of the department of clinical chemistry/pathobiochemistry. PATIENTS: We enrolled 66 patients with acute pancreatitis in the present study. Control groups consisted of healthy subjects (n = 30), patients with chronic pancreatitis (n = 20), patients with pancreatic carcinoma (n = 20), and patients with acute appendicitis (n = 20). INTERVENTIONS: Blood samples were collected during 14 consecutive days in patients with acute pancreatitis. A single blood specimen was taken in all control groups after the diagnosis was established. MEASUREMENTS AND MAIN RESULTS: SAA concentrations were 3 mg/L (median; range, 3-93) in healthy subjects. Although SAA and CRP both reached their maximum within 4 days after onset of symptoms in patients with acute pancreatitis, SAA concentrations rose faster above normal ranges and reached 676 mg/L (median; range, 12-1880), higher than CRP, which reached 313 mg/L (median; range, 29-613). As observed for CRP, SAA was significantly higher in patients who developed complications such as necrosis, infection of necrosis, or multiple organ dysfunction syndrome or in patients who died. SAA achieved best results in discriminating between necrotizing pancreatitis and interstitial edematous pancreatitis. However, CRP provided an earlier differentiation between both entities and a significantly better overall accuracy, as shown by receiver operating characteristics analysis. SAA concentrations in patients with chronic pancreatitis were 6 mg/L (median; range, 3-756). In patients with pancreatic carcinoma, SAA concentrations were 7 mg/L (median; range, 3-492), and in patients with acute appendicitis, they were 50 mg/L (median; range, 3-2140). CONCLUSION: SAA is a nonspecific and rapidly produced variable in inflammatory abdominal disorders with a wider dynamic range than CRP. The current assay technique renders SAA an applicable and readily available variable under clinical routine conditions. In cases of acute pancreatitis, however, CRP is still superior to SAA for early and accurate stratification of patients with a complicated course.

The clinical value of procalcitonin in the prediction of infected necrosis in acute pancreatitis.

OBJECTIVE: Infection of pancreatic necrosis (IN) has a major impact on management and outcome in acute pancreatitis (AP). Currently, guided fine-needle aspiration (FNA) is the only means for an accurate diagnosis of IN. Procalcitonin (PCT), a 116 amino acid pro-peptide of calcitonin has been found in high concentrations in patients with sepsis. In the present study we analyzed the clinical value of serum PCT for predicting IN in AP and compared the results to guided FNA. DESIGN: Clinical study. SETTING: A collaborative study between the Departments of General Surgery and Clinical Chemistry/ Pathobiochemistry of the University of Ulm, Germany. PATIENTS: 61 patients with AP entered this study and were stratified into three groups according to morphological and bacteriological data: I. 22 patients with edematous pancreatitis (AIP), II. 18 patients with sterile necrosis (SN), III. 21 patients with IN. MEASUREMENTS AND RESULTS: During an observation period of 14 days PCT was measured by immunoluminometry, CRP was determined by lasernephelometry on a routine base. In patients with IN overall PCT concentrations were significantly higher than in those with SN, whereas CRP levels did not differ in both groups. In contrast, only low concentrations of both parameters were found in patients with AIP. By ROC analysis the best PCT cut-off level for predicting IN or persisting pancreatic sepsis was obtained at > or =1.8 ng/ml. If this cut-off was reached on at least two consecutive days, IN could be predicted with a sensitivity of 95%, a specificity, of 88%, and an accuracy of 90%. Guided FNA achieved a sensitivity, specificity, and accuracy of 91%. 79%, and 84% in differentiating IN from SN, respectively. After surgical treatment of IN median PCT values continued to be significantly higher in patients with persisting pancreatic sepsis (n=12) compared to those with an uneventful postoperative course (n=7). Our results demonstrate that monitoring of serum PCT could serve as a noninvasive and accurate method to predict IN in AP as well as to select patients with persisting septic complications after surgical debridement.

Lipopolysaccharide-activated macrophages stimulate the synthesis of collagen type I and C-fibronectin in cultured pancreatic stellate cells.

We have recently identified and characterized pancreatic stellate cells (PSC) in rats and humans (Gastroenterology 1998, 15:421-435). PSC are suggested to represent the main cellular source of extracellular matrix in chronic pancreatitis. Now we describe a paracrine stimulatory loop between human macrophages and PSC (rat and human) that results in an increased extracellular matrix synthesis. Native and transiently acidified supernatants of cultured macrophages were added to cultured PSC in the presence of 0.1% fetal calf serum. Native supernatants of lipopolysaccharide-activated macrophages stimulated the synthesis of collagen type I 1.38 +/- 0.09-fold of control and c-fibronectin 1.89 +/- 0.18-fold of control. Transiently acidified supernatants stimulated collagen type I and c-fibronectin 2.10 +/- 0.2-fold and 2.80 +/- 0.05-fold of control, respectively. Northern blot demonstrated an increased expression of the collagen-I-(alpha-1)-mRNA and fibronectin-mRNA in PSC 10 hours after addition of the acidified macrophage supernatants. Cell proliferation measured by bromodeoxyuridine incorporation was not influenced by the macrophage supernatants. Unstimulated macrophages released 1.97 pg TGFbeta1/microgram of DNA over 24 hours and lipopolysaccharide-activated macrophages released 6.61pg TGFbeta1/microgram of DNA over 24 hours. These data together with the results that, in particular, transiently acidified macrophage supernatants increased matrix synthesis, identify TGFbeta as the responsible mediator. In conclusion, our data demonstrate a paracrine stimulation of matrix synthesis of pancreatic stellate cells via TGFbeta1 released by activated macrophages. We suggest that macrophages might play a pivotal role in the development of pancreas fibrosis.

Separation of lipoproteins by capillary isotachophoresis combined with enzymatic derivatization of cholesterol and triglycerides.

Combining specific enzymatic derivatization of cholesterol or triglycerides with capillary isotachophoresis (CITP), human serum lipoproteins are separated into 14 lipoprotein subfractions, monitored and quantitated by direct capillary UV detection. By comparing the separation patterns of human serum with the patterns of lipoprotein particles isolated by sequential ultracentrifugation it became evident that peaks 1-5 represent lipoproteins of the high density lipoprotein (HDL) fraction, peaks 6-8 embody the very low density lipoprotein (VLDL) fraction and chylomicrons, and peaks 7-14 represent the low density lipoprotein (LDL) fraction. Peaks 7 and 8 were found in the VLDL as well as in the LDL fraction. Using triglyceride-specific staining peaks 6-8 occurred prominently; and with cholesterol-specific staining, peaks 1-5 and 7-14 were prominent. The coefficient of variation, for the sum of the peak heights of a pooled serum, was 3.94 for triglyceride-specific staining and 2.32 for cholesterol-specific staining. A linearity range between 0.23 and 2.29 mM/L was found for triglyceride-specific staining and between 0.043 and 4.33 mM/L for cholesterol-specific staining. The practicability of the method was evaluated (i) using blood of humans before and 45 min after an oral fat load. Triglyceride-specific staining revealed a prominent increase in the VLDL fraction and chylomicrones containing peaks 6 and 7, and a minor increase in the HDL fraction containing peaks 3 and 4, and (ii) in patients with manifest hypothyroidism before and after thyroxine therapy. Cholesterol-specific staining demonstrated a massive decrease in the first peak of the HDL fraction and in peaks 9 and 11 of the LDL fraction regarding the hypo versus hyperthyroid state.

Depending on their concentration oxidized low density lipoproteins stimulate extracellular matrix synthesis or induce apoptosis in human coronary artery smooth muscle cells.

Various lines of evidence indicate that oxidative stress resulting in lipid peroxidation and protein modification is involved in the pathogenesis of atherosclerosis and coronary heart disease. We have investigated the effect of modified (oxidized) low-density lipoproteins (oxLDL) on collagen and fibronectin synthesis in cultured human coronary artery smooth muscle cells (HCA-SMC). As shown by immunofluorescence microscopy and time-resolved fluorescence immunoassay, oxLDL dose-dependently stimulated collagen type I and fibronectin synthesis in cultured HCA-SMC. The effect on matrix synthesis was biphasic, with a maximum effect at concentrations between 1 and 10 microg/ml oxLDL. Higher oxLDL concentrations (>25 microg/ml) were cytotoxic. Beside oxLDL, malondialdehyde-modified LDL also stimulated extracellular matrix synthesis. In the presence of 100 microg/ml ascorbic acid, 25, 50 and 100 microg/ml oxLDL induced apoptosis within 6-8 hours (demonstrated by TUNEL-reaction, annexin-V binding and APO-2.7-expression). Apoptosis was not induced by normal (unmodified) LDL and malondialdehyde-modified LDL. The radical scavengers and antioxidants TROLOX and probucol and the hydrogen peroxide eliminator catalase significantly reduced oxLDL-induced apoptosis. Our results demonstrate that low concentrations of oxLDL are profibrogenic by stimulating extracellular matrix synthesis, whereas higher oxLDL concentrations induce oxidative stress and apoptosis in coronary artery smooth muscle cells. The profibrogenic effect might be relevant in the formation of atherosclerotic plaques, and the proapoptotic effect might contribute to an increased plaque vulnerability.

Zinc status in patients with alveolar echinococcosis is related to disease progression.

Zinc is an essential trace element for immune function that plays a role in immune response against parasites. To determine a possible relationship between zinc level and disease status in alveolar echinococcosis (AE), we investigated serum concentrations of zinc, immunoglobulin (Ig)E, IgG, and C-reactive protein (CRP) in 40 AE patients and 20 controls. Patients were classified into three groups: group A: patients after curative surgery, group B: patients with stabilized disease, group C: patients with progressive disease. Patients showed significantly higher levels of IgE and IgG than controls. Amounts of IgE and IgG were related to disease severity, achieving highest levels in group C and lowest in group A. Zinc levels were comparable in patients and controls. However, there was an obvious association between zinc concentration and disease severity. Zinc was far below the normal range in group C (median 9.2 micromol/l) and significantly diminished compared to group B and controls. An inverse pattern was seen for CRP. In conclusion, lowered zinc concentration in progressive cases may be caused by enhanced immune activation but consumption of zinc by the growing parasite may also play a role. Furthermore, decreased zinc levels may contribute to the observed immunosuppression in AE.

Serum half-life time determination of free and total prostate-specific antigen following radical prostatectomy--a critical assessment.

OBJECTIVES: All studies investigating the elimination kinetics of serum total (tPSA) and free (fPSA) prostate-specific antigen (PSA) were carried out in men undergoing radical prostatectomy. Radical prostatectomy itself could, however, have a major influence on the serum concentration of these tumor markers (e.g., perioperative fluid shift or blood loss). The purpose of our study was to determine the half-life time of fPSA and tPSA with special regard to the influence of the radical prostatectomy on the serum concentration of these tumor markers. METHODS: Eleven men (mean age 63.2+/-7.2 years) with organ-confined prostate cancer who underwent radical prostatectomy were investigated (final pathologic Stage pT2pN0 or lower). Serum samples were obtained preoperatively and 0.25, 0.5, 1, 2, 4, 8, 12, 16, 24, 48, 72, 120, 168, and 240 hours after removal of the prostate. fPSA and tPSA and albumin and total protein serum concentrations were determined in all samples. RESULTS: During the first 120 minutes after removal of the prostate, albumin and total protein serum concentrations continuously declined, with a half-life time of -104.5+/-28 minutes and -129.7+/-32 minutes, respectively. Serum decline of fPSA and tPSA followed a biphasic kinetic. During the initial alpha-phase, fPSA and tPSA serum concentrations decreased, with a half-life time of -69+/-10.3 minutes and -87.3+/-18.1 minutes, respectively. During the terminal beta-phase, the half-life time of fPSA and tPSA was -1152.2 minutes (0.8 days) and -3916.1 minutes (2.7 days), respectively. Between the alpha-phase half-life time of fPSA or tPSA and the half-life time of the total protein or albumin concentration decline, significant correlations were found. CONCLUSIONS: These correlations indicate that the rapid decline of fPSA and tPSA directly after removal of the prostate (alpha-phase half-life time) is caused by the radical prostatectomy itself. The half-life time of the beta-phase reflects the biologic clearance of PSA. Therefore, the half-life time determination of PSA after radical prostatectomy is of limited value if the influence of the operation itself on the serum PSA concentration is not taken into account.

Identification, culture, and characterization of pancreatic stellate cells in rats and humans.

BACKGROUND & AIMS: Until now, the basic matrix-producing cell type responsible for pancreas fibrosis has not been identified. In this report, retinoid-containing pancreatic stellate cells (PSCs) in rat and human pancreas are described, and morphological and biochemical similarities to hepatic stellate cells are shown. METHODS: Electron and immunofluorescence microscopy (collagen types I and III, fibronectin, laminin, alpha-actin, and desmin) was performed using pancreatic tissue and cultured PSCs. Extracellular matrix synthesis was shown using quantitative immunoassay and Northern blot analysis. RESULTS: PSCs are located in interlobular areas and in interacinar regions. Early primary cultured PSCs contain retinol and fatty acid retinyl-esters. Addition of retinol to passaged cells resulted in retinol uptake and esterification. During primary culture, the cells changed from a quiescent fat-storing phenotype to a highly synthetic myofibroblast-like cell expressing iso-alpha-smooth muscle actin (>90%) and desmin (20%-40%) and showing strong positive staining with antibodies to collagen types I and III, fibronectin, and laminin. As determined on protein and messenger RNA level, serum growth factors stimulated the synthesis of collagen type I and fibronectin. CONCLUSIONS: The identification of PSCs, particularly in fibrotic areas, and the similarities of these cells to hepatic stellate cells suggest that PSCs participate in the development of pancreas fibrosis.