Specific nail alterations in cutaneous T-cell lymphoma: successful treatment with topical mechlorethamine

Cutaneous T-cell lymphoma can be associated with clinically significant nail alterations, the presentation of which can be protean and misleading. To date, only a few reports have demonstrated direct specific tumor infiltration of the nail bed, while little is known about the efficacy of topical treatments.

Stress-induced alterations in coagulation: assessment of a new hemoconcentration correction technique

For the examination of psychological stress effects on coagulation, the Dill and Costill correction (DCC) for hemoconcentration effects has been used to adjust for stress-induced plasma volume changes. Although the correction is appropriate for adjusting concentrations of various large blood constituents, it may be inappropriate for time-dependent or functional coagulation assays. Two new plasma reconstitution techniques for correcting hemoconcentration effects on stress-induced changes in coagulation were compared with the DCC.

Serum amino acid profiles and their alterations in colorectal cancer

Mass spectrometry-based serum metabolic profiling is a promising tool to analyse complex cancer associated metabolic alterations, which may broaden our pathophysiological understanding of the disease and may function as a source of new cancer-associated biomarkers. Highly standardized serum samples of patients suffering from colon cancer (n = 59) and controls (n = 58) were collected at the University Hospital Leipzig. We based our investigations on amino acid screening profiles using electrospray tandem-mass spectrometry. Metabolic profiles were evaluated using the Analyst 1.4.2 software. General, comparative and equivalence statistics were performed by R 2.12.2. 11 out of 26 serum amino acid concentrations were significantly different between colorectal cancer patients and healthy controls. We found a model including CEA, glycine, and tyrosine as best discriminating and superior to CEA alone with an AUROC of 0.878 (95% CI 0.815-0.941). Our serum metabolic profiling in colon cancer revealed multiple significant disease-associated alterations in the amino acid profile with promising diagnostic power. Further large-scale studies are necessary to elucidate the potential of our model also to discriminate between cancer and potential differential diagnoses. In conclusion, serum glycine and tyrosine in combination with CEA are superior to CEA for the discrimination between colorectal cancer patients and controls.

Profiling of alterations in platelet proteins during storage of platelet concentrates

BACKGROUND: The quality of platelet concentrates (PCs) is primarily determined in vitro by selective methods (e.g., pH, aggregometry), which provide only limited information on certain platelet (PLT) characteristics. In contrast, proteomic technologies provide a comprehensive overview of the PLT proteome. High interassay variability, however, limits meaningful assessment of samples taken from the same product over time or before and after processing. STUDY DESIGN AND METHODS: Differential in-gel electrophoresis (DIGE) and mass spectrometry were applied to analyze changes in the PLT proteome during storage of PCs. RESULTS: DIGE provides a comprehensive and reproducible overview of the cytoplasmic PLT proteome (median standard deviation of protein spot intensities, 5%-9%). Although 97 percent of cytosolic PLT proteins remained unchanged over a 9-day storage period, septin 2 showed characteristic alterations that preceded by several days more widespread alterations affecting numerous other proteins. Also beta-actin and gelsolin are potential marker proteins for changes in the PLT proteome. Interestingly septin 2 and gelsolin are affected during apoptosis, indicating that apoptosis in PCs may have an impact on PLT storage. CONCLUSION: DIGE is a tool for comprehensively assessing the impact of storage on the global proteome profile of therapeutic PCs. Most of the changes detected are in high-abundance PLT proteins.

Cerebral vasculature is the major target of oxidative protein alterations in bacterial meningitis

We have previously shown that antioxidants such as a-phenyl-tert-butyl nitrone or N-acetylcysteine attenuate cortical neuronal injury in infant rats with bacterial meningitis, suggesting that oxidative alterations play an important role in this disease. However, the precise mechanism(s) by which antioxidants inhibit this injury remain(s) unclear. We therefore studied the extent and location of protein oxidation in the brain using various biochemical and immunochemical methods. In cortical parenchyma, a trend for increased protein carbonyls was not evident until 21 hours after infection and the activity of glutamine synthetase (another index of protein oxidation) remained unchanged. Consistent with these results, there was no evidence for oxidative alterations in the cortex by various immunohistochemical methods even in cortical lesions. In contrast, there was a marked increase in carbonyls, 4-hydroxynonenal protein adducts and manganese superoxide dismutase in the cerebral vasculature. Elevated lipid peroxidation was also observed in cerebrospinal fluid and occasionally in the hippocampus. All of these oxidative alterations were inhibited by treatment of infected animals with N-acetylcysteine or alpha-phenyl-tert-butyl nitrone. Because N-acetylcysteine does not readily cross the blood-brain barrier and has no effect on the loss of endogenous brain antioxidants, its neuroprotective effect is likely based on extraparenchymal action such as inhibition of vascular oxidative alterations.

Post-prandial alterations in LDL size and subclasses in patients with growth hormone deficiency

Several studies have suggested that lipoproteins generated during the post-prandial phase are highly atherogenic, with modifications in low-density lipoproteins (LDL) size and density. In the present study we assessed post-prandial variations in LDL size and subclasses in patients with growth hormone deficiency (GHD).

Proteomic alterations in heat shock protein 27 and identification of phosphoproteins in ascending aortic aneurysm associated with bicuspid and tricuspid aortic valve

Whether or not there are molecular differences, at the intra- and extracellular level, between aortic dilatation in patients with bicuspid (BAV) and those with a tricuspid aortic valve (TAV) has remained controversial for years. We have performed 2-dimensional gel electrophoresis and mass spectrometry coupled with dephosphorylation and phosphostaining experiments to reveal and define protein alterations and the high abundant structural phosphoproteins in BAV compared to TAV aortic aneurysm samples. 2-D gel patterns showed a high correlation in protein expression between BAV and TAV specimens (n=10). Few proteins showed significant differences, among those a phosphorylated form of heat shock protein (HSP) 27 with significantly lower expression in BAV compared to TAV aortic samples (p=0.02). The phosphoprotein tracing revealed four different phosphoproteins including Rho GDP dissociation inhibitor 1, calponin 3, myosin regulatory light chain 2 and four differentially phosphorylated forms of HSP27. Levels of total HSP27 and dually phosphorylated HSP27 (S78/S82) were investigated in an extended patient cohort (n=15) using ELISA. Total HSP27 was significantly lower in BAV compared to TAV patients (p=0.03), with no correlation in levels of phospho-HSP27 (S78/S82) (p=0.4). Western blots analysis showed a trend towards lower levels of phospho-HSP27 (S78) in BAV patients (p=0.07). Immunohistochemical analysis revealed that differences in HSP27 occur in the cytoplasma of VSMC's and not extracellularly. Alterations in HSP27 may give early evidence for intracellular differences in aortic aneurysm of patients with BAV and TAV. Whether HSP27 and the defined phosphoproteins have a specific role in BAV associated aortic dilatation remains to be elucidated.

Characterization of white matter alterations in phenylketonuria by magnetic resonance relaxometry and diffusion tensor imaging

A multimodal MR study including relaxometry, diffusion tensor imaging (DTI), and MR spectroscopy was performed on patients with classical phenylketonuria (PKU) and matched controls, to improve our understanding of white matter (WM) lesions. Relaxometry yields information on myelin loss or malformation and may substantiate results from DTI attributed to myelin changes. Relaxometry was used to determine four brain compartments in normal-appearing brain tissue (NABT) and in lesions: water in myelin bilayers (myelin water, MW), water in gray matter (GM), water in WM, and water with long relaxation times (cerebrospinal fluid [CSF]-like signals). DTI yielded apparent diffusion coefficients (ADCs) and fractional anisotropies. MW and WM content were reduced in NABT and in lesions of PKU patients, while CSF-like signals were significantly increased. ADC values were reduced in PKU lesions, but also in the corpus callosum. Diffusion anisotropy was reduced in lesions because of a stronger decrease in the longitudinal than in the transverse diffusion. WM content and CSF-like components in lesions correlated with anisotropy and ADC. ADC values in lesions and in the corpus callosum correlated negatively with blood and brain phenylalanine (Phe) concentrations. Intramyelinic edema combined with vacuolization is a likely cause of the WM alterations. Correlations between diffusivity and Phe concentrations confirm vulnerability of WM to high Phe concentrations.

Alterations in nitric oxide synthase isoforms in acute lower limb ischemia and reperfusion

Alterations in nitric oxide synthase (NOS) are implicated in ischemia and ischemia-reperfusion injury. Changes in the 3 NOS isoforms in human skeletal muscle subjected to acute ischemia and reperfusion were studied. Muscle biopsies were taken from patients undergoing total knee replacement. Distribution of the specific NOS isoforms within muscle sections was studied using immunohistochemistry. NOS mRNA levels were measured using real-time reverse transcription-polymerase chain reaction and protein levels studied using Western blotting. NOS activity was also assessed using the citrulline assay. All 3 NOS isoforms were found in muscle sections associated with muscle fibers and microvessels. In muscle subjected to acute ischemia and reperfusion, NOS I/neuronal NOS mRNA and protein were elevated during reperfusion. NOS III/endothelial NOS was also upregulated at the protein level during reperfusion. No changes in NOS II/inducible NOS expression or NOS activity occurred. In conclusion, alterations in NOS I and III (neuronal NOS and endothelial NOS) at different levels occurred after acute ischemia and reperfusion in human skeletal muscle; however, this did not result in increased NOS activity. In the development of therapeutic agents based on manipulation of the NO pathway, targeting the appropriate NOS isoenzymes may be important.

Nitric oxide synthase in critically ischaemic muscle and alterations in isoform expression during revascularization surgery

BACKGROUND: Dysfunction of the nitric oxide pathway is implicated in peripheral arterial disease. Nitric oxide synthase (NOS) isoforms and NOS activity were studied in muscle from patients with critical leg ischaemia (CLI). Alterations in NOS during revascularization surgery were also assessed. METHODS: Muscle biopsies were taken from patients with CLI undergoing amputation and also from patients undergoing femorodistal bypass at the start of surgery, after arterial clamping and following reperfusion. The presence of NOS within muscle sections was confirmed using reduced nicotinamide adenine dinucleotide phosphate diaphorase histochemistry. NOS isoform distribution was studied by immunohistochemistry. NOS mRNA and protein levels were measured using real-time reverse transcriptase-polymerase chain reaction and western blotting. NOS activity was assessed with the citrulline assay. RESULTS: All three NOS isoforms were found in muscle, associated with muscle fibres and microvessels. NOS I and III protein expression was increased in CLI (P = 0.041). During revascularization, further ischaemia and reperfusion led to a rise in NOS III protein levels (P = 0.008). NOS activity was unchanged. CONCLUSION: Alterations in NOS I and III occurred in muscle from patients with CLI and further changes occurred during bypass surgery.

Mice with a targeted disruption of the Fgfrl1 gene die at birth due to alterations in the diaphragm

FGFRL1 is a recently discovered member of the fibroblast growth factor receptor family that is lacking the intracellular tyrosine kinase domain. To elucidate the function of the novel receptor, we created mice with a targeted disruption of the Fgfrl1 gene. These mice develop normally until term, but die within a few minutes after birth due to respiratory failure. The respiratory problems are explained by a significant reduction in the size of the diaphragm muscle, which is not sufficient to inflate the lungs after birth. The remaining portion of the diaphragm muscle appears to be well developed and innervated. It consists of differentiated myofibers with nuclei at the periphery. Fast and slow muscle fibers occur in normal proportions. The myogenic regulatory factors MyoD, Myf5, myogenin and Mrf4 and the myocyte enhancer factors Mef2A, Mef2B, Mef2C and Mef2D are expressed at normal levels. Experiments with a cell culture model involving C2C12 myoblasts show that Fgfrl1 is expressed during the late stages of myotube formation. Other skeletal muscles do not appear to be affected in the Fgfrl1 deficient mice. Thus, Fgfrl1 plays a critical role in the development of the diaphragm.