Analyses of Synthetic N-Acyl Dopamine Derivatives Revealing Different Structural Requirements for Their Anti-inflammatory and Transient-Receptor-Potential-Channel-of-the-Vanilloid-Receptor-Subfamily-Subtype-1 (TRPV1)-Activating Properties.

We studied the chemical entities within N-octanoyl dopamine (NOD) responsible for the activation of transient-receptor-potential channels of the vanilloid-receptor subtype 1 (TRPV1) and inhibition of inflammation. The potency of NOD in activating TRPV1 was significantly higher compared with those of variants in which the ortho-dihydroxy groups were acetylated, one of the hydroxy groups was omitted ( N-octanoyl tyramine), or the ester functionality consisted of a bulky fatty acid ( N-pivaloyl dopamine). Shortening of the amide linker (ΔNOD) slightly increased its potency, which was further increased when the carbonyl and amide groups (ΔNODR) were interchanged.

Donor Preconditioning After the Onset of Brain Death With Dopamine Derivate n-Octanoyl Dopamine Improves Early Posttransplant Graft Function in the Rat.

Heart transplantation is the therapy of choice for end-stage heart failure. However, hemodynamic instability, which has been demonstrated in brain-dead donors (BDD), could also affect the posttransplant graft function. We tested the hypothesis that treatment of the BDD with the dopamine derivate n-octanoyl-dopamine (NOD) improves donor cardiac and graft function after transplantation.

Evaluation of Perfluorohexyloctane/Polydimethylsiloxane for Pancreas Preservation for Clinical Islet Isolation and Transplantation.

This study aimed to evaluate a 50:50 mix of perfluorohexyloctane/polydimethylsiloxane 5 (F6H8S5) preservation of pancreases in a clinical setting compared with standard solutions for 1) cold ischemia time (CIT) 10 h and 2) an extended CIT 20 h. Procured clinical-grade pancreases were shipped in either F6H8S5 or in standard preservation solutions, that is, University of Wisconsin (UW) or Custodiol. F6H5S5 was preoxygenated for at least 15 min.

F Oximetry with semifluorinated alkanes.

This work examines the variation of longitudinal relaxation rate R1(= 1/T1) of the F-CF-resonance of semifluorinated alkanes (SFAs) with oxygen tension (pO), temperature (T) and pH in vitro. Contrary to their related perfluorocarbons (PFCs), SFA are amphiphilic and facilitate stable emulsions, a prerequisite for clinical use. A linear relationship between R1 and pO was confirmed for the observed SFAs at different temperatures.

Pancreatic L-Glutamine Administration Protects Pig Islets From Cold Ischemic Injury and Increases Resistance Toward Inflammatory Mediators.

The isolation and transplantation of porcine islets represent a future option for the treatment of type 1 diabetic patients. Stringent product release criteria and limited availability of transgenic and specific pathogen-free pigs will essentially require processing of explanted pig pancreata in specialized, possibly remote isolation facilities, whereby pancreata are exposed to cold ischemia due to prolonged tissue transit time. In the present study we investigated whether pancreas oxygenation can be efficiently combined with an antioxidant strategy utilizing intraductal L-glutamine administration.

N-Octanoyl Dopamine for Donor Treatment in a Brain-death Model of Kidney and Heart Transplantation.

Background: This study investigated the potential use of N-octanoyl dopamine (NOD) in donor management to ameliorate the damage caused by brain death and ischemia-reperfusion injury in a rat model of kidney and heart transplantation.

Methods: Brain-dead Fisher rats were treated for 6 hours with either saline or saline plus NOD. Orthotopic kidney and heterotopic heart transplantation were performed in different Lewis recipient rats.

Quality of isolated pig islets is improved using perfluorohexyloctane for pancreas storage in a split lobe model.

Pancreas transportation between donor center and islet production facility is frequently associated with prolonged ischemia impairing islet isolation and transplantation outcomes. It is foreseeable that shipment of pig pancreases from distant centralized biosecure breeding facilities to institutes that have a long-term experience in porcine islet isolation is essentially required in future clinical islet xenotransplantation. Previously, we demonstrated that perfluorohexyloctan (F6H8) is significantly more efficient to protect rat and human pancreata from ischemically induced damage compared to perfluorodecalin (PFD).

N-octanoyl-dopamine is a potent inhibitor of platelet function.

Dopamine (DA) is a co-agonist for platelet activation; yet, donor DA treatment is associated with improved transplantation outcome in renal and heart recipients. Recently, N-octanoyl-dopamine (NOD) was developed which displays superior effects compared to DA in terms of graft protecting properties. Whereas DA is a known platelet co-agonist, the effect of NOD on platelet function is unknown.

N-octanoyl-dopamine is an agonist at the capsaicin receptor TRPV1 and mitigates ischemia-induced [corrected] acute kidney injury in rat.

Since stimulation of transient receptor potential channels of the vanilloid receptor subtype 1 (TRPV1) mitigates acute kidney injury (AKI) and endogenous N-acyl dopamine derivatives are able to activate TRPV1, we tested if synthetic N-octanoyl-dopamine (NOD) activates TRPV1 and if it improves AKI. These properties of NOD and its intrinsic anti-inflammatory character were compared with those of dopamine (DA). TRPV1 activation and anti-inflammatory properties of NOD and DA were tested using primary cell cultures in vitro.

Improvement of oxygen supply by an artificial carrier in combination with normobaric oxygenation decreases the volume of tissue hypoxia and tissue damage from transient focal cerebral ischemia.

Tissue hypoxia may play an important role in the development of ischemic brain damage. In the present study we investigated in a rat model of transient focal brain ischemia the neuroprotective effects of increasing the blood oxygen transport capacity by applying a semifluorinated alkane (SFA)-containing emulsion together with normobaric hyperoxygenation (NBO). The spread of tissue hypoxia was studied using pimonidazole given prior to filament-induced middle cerebral artery occlusion (MCAO, 2 h).

Aerosolized semifluorinated alkanes as excipients are suitable for inhalative drug delivery--a pilot study.

Semifluorinated alkanes (SFAs) have been described as potential excipients for pulmonary drug delivery, but proof of their efficacy is still lacking. We tested whether SFA formulations with the test drug ibuprofen can be nebulised and evaluated their pharmacokinetics. Physico-chemical properties of five different ibuprofen formulations were evaluated: an aqueous solution (H2O), two different SFAs (perfluorohexyloctane (F6H8), perfluorobutylpentane (F4H5)) with and without ethanol (SFA/EtOH).

UW is superior compared with HTK after prolonged preservation of renal grafts.

Background: In recent clinical studies, the efficacy of histidine-tryptophan-ketoglutarate (HTK) in kidney transplantation was questioned. This study compares the efficacy of University of Wisconsin (UW) and HTK solutions on transplantation outcome.

Materials And Methods: Rat kidneys were preserved for different periods of cold ischemia (CIT).

Semifluorinated alkanes--a new class of excipients suitable for pulmonary drug delivery.

Introduction: Semifluorinated alkanes (SFAs) are considered as diblock molecules with fluorocarbon and hydrocarbon segments. Unlike Perfluorocarbons (PFCs), SFAs have the potential to dissolve several lipophilic or water-insoluble substances. This makes them possibly suitable as new excipients for inhalative liquid drug carrier systems.

A new oxygen carrier for improved long-term storage of human pancreata before islet isolation.

Background: Pancreas oxygenation during cold storage has been established in islet isolation and transplantation to prevent ischemic tissue damage using perfluorodecalin (PFD) as hyperoxygen carrier. However, studies in humans and pigs provided conflicting results about the efficiency of PFD for pancreas oxygenation. The aim of this study was to compare PFD with a newly developed oxygen carrier composed of perfluorohexyloctane and polydimethylsiloxane 5 (F6H8S5) for long-term storage of human pancreata.

Perfluorohexyloctane improves long-term storage of rat pancreata for subsequent islet isolation.

Pancreas oxygenation by means of the hyperoxygen carrier perfluorodecalin (PFD) has been established to prevent ischemically induced damage from cold-stored pancreata. However, large-scale studies did not confirm the promising results that had been observed in smaller donor populations. This study assessed whether islet isolation from pancreata stored for prolonged periods can be improved by utilizing the new oxygen carrier perfluorohexyloctane (F6H8) characterized by lower gravity and higher lipophilicity than PFD.

A cleaning solution for silicone intraocular lenses: "sticky silicone oil".

Aim: The aim of the study was to compare the efficacy of perfluorobutylpentane (F4H5) and perfluorohexyloctane (F6H8) in dissolving silicone oil from the surface of silicone intraocular lenses (IOL).

Methods: Droplets of stained silicone oil were applied to an object slide either lying flat or tilted by 30 degrees . Mixing with H(2)O, F4H5 or F6H8 was documented by a digital camera.

CAPN 8: isolation of a new mouse calpain-isoenzyme.

Recent molecular biological approaches indicate that calpain, also named CANP for calcium-activated neutral protease and originally characterized as an intracellular cytoplasmatic nonlysosomal cysteine protease that requires calcium ions for activity, constitutes a large superfamily consisting of ubiquitous and tissue specific homologues, which are widely distributed in cells of various organisms from human to fungus. Due to the increasing number of substrates along with the involvement of calpain isoenzymes in mammalian diseases, especially in malignancies, members of the calpain superfamily seem to be important biomodulators in physiological as well as pathological cell function. Here we report the characterisation of a new calpain, named CAPN 8 with a different C-terminal domain, implicating a putative new regulatory mechanism.

Expression of calpain I messenger RNA in human renal cell carcinoma: correlation with lymph node metastasis and histological type.

Calpain, also named CANP (for calcium-activated neutral protease), is an intracellular cytoplasmatic non-lysosomal cysteine endopeptidase that requires calcium ions for activity. Many substrates of the calpain isoenzymes, such as the transcription factors c-Fos and c-Jun, the tumor supressor protein p53, protein kinase C, pp60c-src and the adhesion molecule integrin, have been implicated in the pathogenesis of different human tumors, suggesting an important role of the calpains in malignant diseases. We now report differential expression of the calpain I gene (CL I) in a variety of tumors, extending our study to a larger series of renal cell carcinomas.

Glyceraldehyde-3-phosphate dehydrogenase and Nm23-H1/nucleoside diphosphate kinase A. Two old enzymes combine for the novel Nm23 protein phosphotransferase function.

We have recently discovered an alternative function of the putative metastasis suppressor protein Nm23, which is identical to nucleoside diphosphate kinase, as a protein phosphotransferase in vitro. While purified native Nm23 protein did not phosphorylate other proteins, we could purify a Nm23-associated protein that activates the protein phosphotransferase function; it was identified as a glyceraldehyde-3-phosphate dehydrogenase (GAPDH) isoenzyme. Co-expression and purification of (His)6-tagged GAPDH in combination with either Nm23-H1 or Nm23-H2 in baculovirus-infected Sf9 cells showed that only Nm23-H1, but not Nm23-H2, forms a stable complex with GAPDH.

NM23-H1 and NM23-H2 gene expression in human renal tumors.

The two nm23 genes, nm23-H1 and nm23-H2, are implicated in the metastatic process and tumor progression in some human tumors. Until now no data exist about nm23 expression in the different types of human renal tumors. To investigate if the nm23 genes play a central role in the progression of renal tumors, we have examined nm23-H1 and nm23-H2 gene expression using Northern-blot analysis and immunohistochemistry.

Isolation and characterization of new microsatellites at the nm23-H1 and nm23-H2 gene loci and application for loss of heterozygosity (LOH) analysis.

The nm23-H1 gene has been suggested to be a metastasis suppressor gene. Studies about the events of loss of heterozygosity (LOH) at the nm23 locus and its correlation to metastasis are controversially discussed. To optimize detection of LOH at the nm23 locus, we screened two P1 clones for additional microsatellites.

A second trefoil protein, ITF/hP1.B, is transcribed in human breast cancer.

Trefoil proteins form a specific group of stable secreted polypeptides. They are expressed in a lot of human cancers and during inflammatory processes of the gastrointestinal tract. Recently a new human trefoil protein, ITF/hP1.

A novel serine/threonine-specific protein phosphotransferase activity of Nm23/nucleoside-diphosphate kinase.

Two human nm23 genes have been identified, designated nm23-H1 and nm23-H2, which encode the 88% identical nucleoside-diphosphate kinase (NDPK) A and NDPK B polypeptides, respectively. The nm23-H1 gene product has been shown to play a functional role in the suppression of tumor metastasis. The Nm23 proteins/NDPK are highly conserved throughout evolution and are implicated in controlling cellular differentiation and development in various species, while the underlying mechanisms remain undefined.

Presence of regulatory sequences within intron 4 of human and murine c-myb genes.

The molecular mechanisms that modulate c-myb mRNA transcription in hematopoietic cells appear to involve intron regulatory sequences. We have characterized the fourth of ten introns from both human and murine c-myb genes in regard to nucleotide sequence and specific protein binding. For this approach complete genomic c-myb intron 4 fragments were isolated from mouse and human DNA using PCR amplification with flanking exon-primers derived from the mouse gene.