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  • Guimond, Marie-Odile, et al. (författare)
  • Saralasin and Sarile Are AT2 Receptor Agonists
  • 2014
  • Ingår i: ACS Medicinal Chemistry Letters. - 1948-5875 .- 1948-5875. ; 5:10, s. 1129-1132
  • Tidskriftsartikel (refereegranskat)abstract
    • Saralasin and sarile, extensively studied over the past 40 years as angiotensin II (Ang II) receptor blockers, induce neurite outgrowth in a NG108-15 cell assay to a similar extent as the endogenous Ang II. In their undifferentiated state, these cells express mainly the AT2 receptor. The neurite outgrowth was inhibited by preincubation with the AT2 receptor selective antagonist PD 123,319, which suggests that the observed outgrowth was mediated by the AT2 receptor. Neither saralasin nor sarile reduced the neurite outgrowth induced by Ang II proving that the two octapeptides do not act as antagonists at the AT2 receptor and may be considered as AT2 receptor agonists.
  • Hall, Håkan, et al. (författare)
  • In vitro autoradiography of carcinoembryonic antigen in tissue from patients with colorectal cancer using multifunctional antibody TF2 and 67/68Ga-labeled haptens by pretargeting
  • 2012
  • Ingår i: American Journal of Nuclear Medicine and Molecular Imaging. - 2160-8407. ; 2:2, s. 141-150
  • Tidskriftsartikel (refereegranskat)abstract
    • The carcinoembryonic antigen (CEA) was visualized in vitro in tissue from patients with colorectal cancer with trivalent bispecific antibody TF2 and two hapten molecules, [(67/68)Ga]Ga-IMP461 and [(67/68)Ga]Ga-IMP485 by means of pretargeting. Colorectal cancer tissue samples obtained from surgery at Uppsala University Hospital, were frozen fresh and cryosectioned. The two hapten molecules comprising 1,4,7-triazacyclononanetriacetic acid chelate moiety (NOTA) were labeled with (67)Ga or (68)Ga. The autoradiography was conducted by incubating the tissue samples with the bispecific antibody TF2, followed by washing and incubation with one of the radiolabeled hapten molecules. After washing, drying and exposure to phosphor imager plates, the autoradiograms were analyzed and compared to standard histochemistry (hematoxylin-eosin). Pronounced binding was found in the tissue from colorectal cancer using the bispecific antibody TF2 and either of the haptens [(67/68)Ga]Ga-IMP461 and [(67/68)Ga]Ga-IMP485. Distinct binding was also detected in the epithelium of most samples of neighboring tissue, taken at a minimum of 10 cm from the site of the tumor. It is concluded that pretargeting CEA with the bispecific antibody TF2 followed by the addition of (67/68)Ga-labeled hapten is extremely sensitive for visualizing this marker for colorectal cancer. This methodology is therefore a very specific complement to other histochemical techniques in the diagnosis of biopsies or in samples taken from surgery. Use of the pretargeting technique in vivo may also be an advance in diagnosing patients with colorectal cancer, either using (67)Ga and SPECT or (68)Ga and PET.
  • Hammarlund-Udenaes, Margareta (författare)
  • In vivo approaches to assessing the blood-brain barrier
  • 2014
  • Ingår i: The blood-brain barrier (BBB). - Heidelberg New York Dordrecht London : Springer. - 9783662437865 ; , s. 21-48
  • Bokkapitel (refereegranskat)abstract
    • Methods for in vivo assessment of blood-brain barrier (BBB) transport are presented, with their advantages and disadvantages. The methods described are brain uptake index, the i.v. injection technique, in situ brain perfusion, brain efflux index, % injected dose, microdialysis, CSF sampling and positron emission tomography, and the combinatorial mapping of unbound drug partitioning across the BBB. The methods are put into a pharmacokinetic context by delineating the type of readings that they give, be it the rate of transport across the BBB or the extent of transport of total drug (unbound and bound), or of the unbound drug.
  • Harandi, Vahid M., et al. (författare)
  • Antioxidants reduce muscular dystrophy in the dy2J/dy2J mouse model of laminin α2 chain-deficient muscular dystrophy
  • 2020
  • Ingår i: Antioxidants. - : MDPI AG. - 2076-3921. ; 9:3
  • Tidskriftsartikel (refereegranskat)abstract
    • Congenital muscular dystrophy with laminin α2 chain-deficiency (LAMA2-CMD) is a severe neuromuscular disorder without a cure. Using transcriptome and proteome profiling as well as functional assays, we previously demonstrated significant metabolic impairment in skeletal muscle from LAMA2-CMD patients and mouse models. Reactive oxygen species (ROS) increase when oxygen homeostasis is not maintained and, here, we investigate whether oxidative stress indeed is involved in the pathogenesis of LAMA2-CMD. We also analyze the effects of two antioxidant molecules, N-acetyl-L-cysteine (NAC) and vitamin E, on disease progression in the dy2J/dy2J mouse model of LAMA2-CMD. We demonstrate increased ROS levels in LAMA2-CMD mouse and patient skeletal muscle. Furthermore, NAC treatment (150 mg/kg IP for 6 days/week for 3 weeks) led to muscle force loss prevention, reduced central nucleation and decreased the occurrence of apoptosis, inflammation, fibrosis and oxidative stress in LAMA2-CMD muscle. In addition, vitamin E (40 mg/kg oral gavage for 6 days/week for 2 weeks) improved morphological features and reduced inflammation and ROS levels in dy2J/dy2J skeletal muscle. We suggest that NAC and to some extent vitamin E might be potential future supportive treatments for LAMA2-CMD as they improve numerous pathological hallmarks of LAMA2-CMD.
  • Hedeland, Mikael, et al. (författare)
  • Confirmation of botulism in birds and cattle by the mouse bioassay and Endopep-MS
  • 2011
  • Ingår i: Journal of Medical Microbiology. - 0022-2615 .- 1473-5644. ; 60:9, s. 1299-1305
  • Tidskriftsartikel (refereegranskat)abstract
    • There have been several outbreaks of botulism among poultry and wild birds in Sweden in recent years. The National Veterinary Institute of Sweden (SVA) has identified botulinum neurotoxin (BoNT)/C1 or the mosaic BoNT/C1D using the mouse bioassay. This is believed to be the first report on the application of the Endopep mass spectrometry (Endopep-MS) method to selected clinical animal (serum and liver) samples and a feed sample that had previously given positive test results with the mouse bioassay. In the mouse bioassay eight of the eleven samples were found to be neutralized by both BoNT/C1 and /D antitoxins; the other three were neutralized only by BoNT/C1 antitoxin, but the mice showed a prolonged survival time when the samples had been treated with /D antitoxin. The Endopep-MS analysis, on the other hand, demonstrated only BoNT/C1 activity for all eleven samples. This suggests that at least eight of the samples were of the chimeric toxin type BoNT/C1D, where the enzymically active site is identical to that of BoNT/C1, while other parts of the protein contain sequences of BoNT/D. This is the first step of a cross-validation between the established mouse bioassay and the Endopep-MS of serotypes BoNT/C1 and /C1D. Endopep-MS is concluded to have potential as an attractive alternative to the mouse bioassay.
  • Hillarp, Andreas, et al. (författare)
  • Unveiling the complex effects of direct oral anticoagulants on dilute Russell's viper venom time assays
  • 2020
  • Ingår i: Journal of Thrombosis and Haemostasis. - : Wiley-Blackwell. - 1538-7933 .- 1538-7836. ; 18:8, s. 1866-1873
  • Tidskriftsartikel (refereegranskat)abstract
    • Introduction: Dilute Russell viper venom time (dRVVT) assays can be affected by direct oral anticoagulants (DOACs), which may cause false-positive results. However, there are conflicting results indicating significant differences between different reagents and DOACs. Objectives: To evaluate the effect of DOACs on dRVVT assays. Material and Methods: Samples were prepared by adding DOAC (dabigatran, rivaroxaban, apixaban, or edoxaban) to pooled normal plasma in the concentration range 0 to 800 µg/L. Six integrated dRVVT reagents were used, all composed of a screen assay (low phospholipid content) and a confirm assay (high phospholipid content). The screen/confirm dRVVT results were expressed as normalized ratios. To further evaluate the observed differences between tests and DOACs, addition of synthetic phospholipids was used. Results: The dRVVT ratios increased dose dependently for all DOACs, with four of the six tests and the DOAC rivaroxaban having the greatest effect. With one test, the ratios were almost unaffected with increasing DOAC concentration, whereas another test revealed a negative dose dependency for all DOACs. Variable DOAC effects can be explained by different effects on dRVVT screen and confirm clotting time. Adding synthetic phospholipids to samples containing rivaroxaban resulted in greatly reduced screen clotting times and thereby lower calculated dRVVT ratios. Conclusions: There is a great variability in the dRVVT test result with different DOACs. The dRVVT ratios are unaffected for some reagents and this can be explained by an equal dose-dependent effect on both screen and confirm assays. The phospholipid type and content of the different reagents may contribute to the observed differences.
  • Holmbäck, Jan, et al. (författare)
  • Preclinical development of sodium fusidate antibiotic cutaneous spray based on water-free lipid formulation system
  • 2022
  • Ingår i: European Journal of Pharmaceutical Sciences. - : Elsevier B.V.. - 0928-0987 .- 1879-0720. ; 176
  • Tidskriftsartikel (refereegranskat)abstract
    • Topical antibiotics are a key component in the management of mild to moderate skin and soft tissue infections. There are, however, concerns about the emerging bacterial resistance against topical antibacterial agents such as fusidic acid, due to the prolonged treatment period of its marketed dosage forms. Improving the efficacy of topical formulations could potentially shorten the treatment period and avoid the resistance growth. To provide a more effective drug delivery, a water-free lipid-based formulation system (AKVANO (R)) which can be applied by spraying, has been developed. In the current paper, different formulations containing sodium fusidate were evaluated for their in vitro skin permeability using artificial skin mimicking membranes and antibacterial properties using ex vivo and in vivo skin wound infection models. The novel formulations containing sodium fusidate showed a much higher skin permeation (up to 60% of nominal amount) than the commercially available Fucidin (R) cream (3%). These formulations also gave a significantly stronger antibacterial effect than Fucidin cream showing a clear dose-response relationship for the sodium fusidate content. A spray product based on the described formulation technology would therefore require a shorter treatment time and thereby lower the risk for the development of bacterial resistance. Spray administration of these formulations provides an even layer on the skin surface from which the solvent quickly evaporates and thereby facilitates a non-touch application where no rubbing is required.
  • Horvath, Istvan, et al. (författare)
  • Immunochemical Detection of alpha-Synuclein Autoantibodies in Parkinson's Disease : Correlation between Plasma and Cerebrospinal Fluid Levels
  • 2017
  • Ingår i: ACS Chemical Neuroscience. - : American Chemical Society (ACS). - 1948-7193 .- 1948-7193. ; 8:6, s. 1170-1176
  • Tidskriftsartikel (refereegranskat)abstract
    • Autoantibodies to Parkinson's disease (PD) amyloidogenic protein, a-synuclein, were recognized as a prospective biomarker for early disease diagnostics, yet there is inconsistency in previous reports, potentially related to PD status. Therefore, plasma and cerebrospinal fluid (CSF) of the cross-sectional cohort of 60 individuals, including recently diagnosed PD patients with mild and moderate PD and age-matched controls, were examined by enzyme-linked immunosorbent assay (ELISA). Nonparametric statistics was used for data analysis. We found significantly elevated levels of a-synuclein autoantibodies in both plasma and CSF in mild PD compared to controls, followed by some decrease in moderate PD. Receiver operating characteristic and effect size analyses confirmed the diagnostic power of a-synuclein antibodies in both plasma and CSF. For the first time, we showed the correlation between plasma and CSF a-synuclein antibody levels for mild, moderate, and combined PD groups. This indicates the potentiality of a-synuclein antibodies as PD biomarker and the increased diagnostic power of their simultaneous analysis in plasma and CSF.
  • Jankovskaja, Skaidre, et al. (författare)
  • Optimization of sample preparation for transporter protein quantification in tissues by LC–MS/MS
  • 2019
  • Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier. - 0731-7085 .- 1873-264X. ; 164, s. 9-15
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: Reproducible quantification of drug transporter protein expression in tissues is important for predicting transporter mediated drug disposition. Many mass-spectrometry based transporter protein quantification methods result in high variability of the estimated transporter quantities. Therefore, we aimed to evaluate and optimize mass spectrometry-based quantification method for drug transporter proteins in tissues. Materials and methods: Plasma membrane (PM) proteins from mouse tissues were isolated by applying three extraction protocols: commercial plasma membrane extraction kit, tissue homogenization by Potter-Elvehjem homogenizer in combination with sucrose-cushion ultracentrifugation, and PM enrichment with Tween 40. Moreover, five different protein digestion protocols were applied on the same PM fraction. PM isolation and digestion protocols were evaluated by measuring the amount of transporter proteins by liquid chromatography-tandem mass spectrometry in selected reaction monitoring mode. Results: Mouse liver homogenization by Potter-Elvehjem homogenizer in combination with sucrose-cushion ultracentrifugation and PM enrichment with Tween 40 resulted in two times higher transporter protein quantity (Breast cancer resistance protein (Bcrp) 18.0 fmol/μg protein) in comparison with the PM samples isolated by extraction kit (Bcrp 9.8 fmol/μg protein). The evaluation of protein digestion protocols revealed that the most optimal protocol for PM protein digestion is with Lys-C and trypsin, in combination with trypsin enhancer and heat denaturation. Overall, quantities of Bcrp and Na+/K + ATPase proteins evaluated in mouse liver and kidney cortex by using our optimized PM isolation method, as well as, established digestion protocol were two to three times higher than previously reported and coefficient of variation (CV) for technical replicates was below 10%. Conclusion: We have established an improved transporter protein quantification methodology by optimizing PM isolation and protein digestion procedures. The optimized procedure resulted in a higher transporter protein yield and improved precision.
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