| 1. |
- Almquist, Joachim, et al.
(författare)
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Model-Based Analysis Reveals a Sustained and Dose-Dependent Acceleration of Wound Healing by VEGF-A mRNA (AZD8601)
- 2020
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Ingår i: CPT. - : WILEY. - 2163-8306. ; 9:7, s. 384-394
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Tidskriftsartikel (refereegranskat)abstract
- Intradermal delivery of AZD8601, an mRNA designed to produce vascular endothelial growth factor A (VEGF-A), has previously been shown to accelerate cutaneous wound healing in a murine diabetic model. Here, we develop population pharmacokinetic and pharmacodynamic models aiming to quantify the effect of AZD8601 injections on the dynamics of wound healing. A dataset of 584 open wound area measurements from 131 mice was integrated from 3 independent studies encompassing different doses, dosing timepoints, and number of doses. Evaluation of several candidate models showed that wound healing acceleration is not likely driven directly by time-dependent VEGF-A concentration. Instead, we found that administration of AZD8601 induced a sustained acceleration of wound healing depending on the accumulated dose, with a dose producing 50% of the maximal effect of 92 mu g. Simulations with this model showed that a single dose of 200 mu g AZD8601 can reduce the time to reach 50% wound healing by up to 5 days.
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| 2. |
- Beverborg, Niels Grote, et al.
(författare)
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Phospholamban antisense oligonucleotides improve cardiac function in murine cardiomyopathy
- 2021
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Ingår i: Nature Communications. - Stockholm : Karolinska Institutet, Dept of Cell and Molecular Biology. - 2041-1723.
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Tidskriftsartikel (refereegranskat)abstract
- Heart failure (HF) is a major cause of morbidity and mortality worldwide, highlighting an urgent need for novel treatment options, despite recent improvements. Aberrant Ca2+ handling is a key feature of HF pathophysiology. Restoring the Ca2+ regulating machinery is an attractive therapeutic strategy supported by genetic and pharmacological proof of concept studies. Here, we study antisense oligonucleotides (ASOs) as a therapeutic modality, interfering with the PLN/SERCA2a interaction by targeting Pln mRNA for downregulation in the heart of murine HF models. Mice harboring the PLN R14del pathogenic variant recapitulate the human dilated cardiomyopathy (DCM) phenotype; subcutaneous administration of PLN-ASO prevents PLN protein aggregation, cardiac dysfunction, and leads to a 3-fold increase in survival rate. In another genetic DCM mouse model, unrelated to PLN (Cspr3/Mlp−/−), PLN-ASO also reverses the HF phenotype. Finally, in rats with myocardial infarction, PLN-ASO treatment prevents progression of left ventricular dilatation and improves left ventricular contractility. Thus, our data establish that antisense inhibition of PLN is an effective strategy in preclinical models of genetic cardiomyopathy as well as ischemia driven HF.
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| 3. |
- De Genst, Erwin, et al.
(författare)
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Blocking phospholamban with VHH intrabodies enhances contractility and relaxation in heart failure
- 2022
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Ingår i: Nature Communications. - Stockholm : Karolinska Institutet, Dept of Cell and Molecular Biology. - 2041-1723.
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Tidskriftsartikel (refereegranskat)abstract
- The dysregulated physical interaction between two intracellular membrane proteins, the sarco/endoplasmic reticulum Ca2+ ATPase and its reversible inhibitor phospholamban, induces heart failure by inhibiting calcium cycling. While phospholamban is a bona-fide therapeutic target, approaches to selectively inhibit this protein remain elusive. Here, we report the in vivo application of intracellular acting antibodies (intrabodies), derived from the variable domain of camelid heavy-chain antibodies, to modulate the function of phospholamban. Using a synthetic VHH phage-display library, we identify intrabodies with high affinity and specificity for different conformational states of phospholamban. Rapid phenotypic screening, via modified mRNA transfection of primary cells and tissue, efficiently identifies the intrabody with most desirable features. Adeno-associated virus mediated delivery of this intrabody results in improvement of cardiac performance in a murine heart failure model. Our strategy for generating intrabodies to investigate cardiac disease combined with modified mRNA and adeno-associated virus screening could reveal unique future therapeutic opportunities.
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| 4. |
- Hansson, Kenny, 1972-, et al.
(författare)
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Comparative studies with surface plasmon resonance and free oscillation rheometry on the inhibition of platelets with cytochalasin E and monoclonal antibodies towards GPIIb/IIIa
- 2002
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Ingår i: Biosensors & bioelectronics. - 0956-5663 .- 1873-4235. ; 17:9, s. 761-771
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Tidskriftsartikel (refereegranskat)abstract
- In the haemostatic system a multitude of processes are intertwined in fine-tuned interactions that arrest bleeding, keep the circulatory system open, and the blood flowing. The occurrence of both surface and bulk interactions adds an additional dimension of complexity. These insights have led to the belief that global overall procedures can inform on the likely behaviour of the system in health and disease. Two sensing procedures: surface plasmon resonance (SPR), which senses surface interactions, and free oscillation rheometry (FOR), which senses interactions within the bulk, have been combined and evaluated. The contribution of blood cells, mainly platelets, to the SPR and FOR signals was explored by simultaneous SPR and FOR measurement during native whole blood coagulation, accelerated via the platelets through addition of SFLLRN peptide and inhibition of platelet aggregation with abciximab (ReoPro®) and of shape change with cytochalasin E. The SPR technique was found to be sensitive to inhibition of blood cell functions such as adhesion to and spreading on surfaces, as well as platelet aggregation. SPR seemed not to be directly sensitive to fibrin polymerisation in coagulating whole blood. The FOR technique detected the coagulation as a bulk phenomenon, i.e. the gelation of the blood due to fibrin formation was detected. The combination of SPR and FOR may therefore be suitable for studies on blood cell functions during coagulation.
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| 5. |
- Hansson, Kenny M., et al.
(författare)
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Abnormalities in coagulum lysis and structure are associated with deep venous thrombosis
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The present study aimed to investigate the relationship between deep venous thrombosis (DVT) and fibrinolytic susceptibility of plasma coagulum, including the possible role of this property in laboratory diagnosis. From 276 patients consecutively admitted to hospital for suspected deep venous thrombosis (DVT), 75 patients and 47 controls were selected. With certainty, the patients and controls either had or did not have DVT. Fibrinolytic susceptibility was assayed by reacting plasma with thromboplastin and tissue plasminogen activator and recording a nephelometric signal. Coagulation time (CT), coagulum lysis time (CLT) and maximal increase in coagulum light scatter (CLS) were determined. Increase in D-dimer levels caused by coagulum lysis was also determined. This was viewed as a fibrinogen measure. CL T and CLS were interpreted as measures of fibrinolytic susceptibility and coagulum structure, respectively. CL T and CLS for patients and controls differed, p<0.025 and p<0.001, respectively. Compared to 5% for controls, 24% and 43% of the patients showed CL T and CLS outside the reference range. High fibrinogen levels could not explain the findings, since these were normal in most patients with abnormal CL T and CLS. Abnormal coagulum lysis and abnormal coagulum structure were thus found to be associated with DVT. Possible laboratory diagnostic role of CL T and CLS was investigated with bivariate reference ranges that excluded 5% and 0.3%. These ranges excluded significantly (p<0.0001) more patients, 47% and 27%, respectively. Tests for abnormal fibrinolytic susceptibility and coagulum structure may thus have a role in laboratory diagnosis of thrombotic disorders.
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| 6. |
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| 7. |
- Hansson, Kenny M., et al.
(författare)
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Surface plasmon resonance and free oscillation rheometry in combination : A new approach forstudies on haemostasis and biomaterials
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- In haemostasis and biomaterial research it is important to be able to study biological processes at surfaces and in the bulk. Surface plasmon resonance (SPR) is sensitive to changes at surface and free oscillation rheometry (FOR) probes the bulk. The present work demonstrates the usefulness of the combination of the techniques for simultaneous real-time measurements on coagulation and fibrinolysis of blood plasma, as well as coagulation of whole blood. SFLLRN stimulated coagulation of native whole blood presented a higher SPR signal with a different appearance than for plasma coagulation, while the FOR signals corresponding to plasma and whole blood coagulation were similar. This result indicated that the SPR technique was more sensitive to cell-surface interactions than to fibrin formation in whole blood, while the FOR technique were equally sensitive to coagulation in whole blood and plasma. Spontaneous coagulation of native whole blood in contact with methyland hydroxyl-terminated self-assembled monolayers on gold and gold surfaces regenerated after coagulation by degradation of adsorbed proteins with trypsin and SOS were also studied. The regenerated gold surfaces displayed the shortest coagulation times, although the contact-activation of blood coagulation was found to be low. The methylated and hydroxylated surfaces were comparable in terms of coagulation activation, while the hydroxylated surfaces presented FOR signals that indicated difficulties for the coagulum to attach to the surface. The combination of SPR and FOR may be suited for studies of cell-surface interactions, and may find applications in studies of blood cell defects in patients and testing of medical substances.
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| 8. |
- Hansson, Kenny, 1972-
(författare)
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Real-time analysis of blood coagulation and fibrinolysis : new rheological and optical sensing techniques for diagnosis of haemostatic disorders.
- 2001
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The haemostatic system has a dual paradoxical function in the body. It should arrest bleeding whenever needed, but also keep the blood flowing in the circulatory system without any obstructing blood clots. The system is complex with maoy intertwined processes that interact to produce a fine-tuned regulation of the performance. In case of malfunction in this regulation there may be an excessive coagulation ability, thrombophilia, or bleeding tendency, haemophilia. These are common disorders in the Western societies and may be lethal. The long-term airo of this work is therefore to improve the laboratory diagnosis of haemostatic disorders, for thrombophilia in particular. To achieve this goal a global approach has been chosen, meaning that the environment in which a blood sample is analysed should mimic the physiology of the haemostatic system to better elucidate the overall situation in a particular individual. A first attempt to assess the susceptibility for tissue plasminogen activator induced lysis and coagulum structure in plasma as markers for deep vein thrombosis showed promising results with 47% abnormals among the DVT patients included in the study. To improve this assay new sensing techniques were needed, since one of the most important conditions included in a global assay is analysis of whole blood, i.e. blood with all types of blood cells present. Whole blood is opaque and excludes the traditional optical methods that have been used for coagulation analysis. Several candidate techniques have been identified and surface plasmon resonance (SPR), quartz crystal microbalance-dissipation (QCM-D), and free oscillation rheometry (FOR) have been evaluated for haemostatic studies in this thesis. SPR is an optical surface sensitive technique that has showed promising results for measurements in blood plasma during coagulation and fibrinolysis and for whole blood coagulation. The SPR responses were sensitive to treatments with heparin and oral anticoagulants, which are substances used to treat thrombosis. QCM-D that is sensitive to mass deposition and viscoelastic changes in the sample at the quartz crystal surface has been tested in combination with SPR and provided new information about the viscoelastic properties of the coagulum, although with similar sensing depth as SPR. The idea of combined sensing techniques was reconsidered and resulted in a combination of SPR and FOR for siroultaneous real-time measurements in a blood sample. FOR is bulk sensitive and probes rheological changes in the sample. The combination was applied in studies of plasma and whole blood coagulation as well as plasma fibrinolysis. Coagulation studies including chemical surface modifications by using thiol-based self-assembled monolayers were also attempted. Finally, the FOR/SPR combination was found to be sensitive to inhibition of platelet aggregation and blood cell shape changes iroplying that studies on the cellular component of the blood is possible. In conclusion, the combination of FOR and SPR is a promising sensing system for an improved global assay for haemostatic disorders.
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| 9. |
- Hansson, Kenny, 1972-, et al.
(författare)
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Surface plasmon resonance and free oscillation rheometry in combination : a useful approach for studies on haemostasis and interactions between whole blood and artificial surfaces
- 2002
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Ingår i: Biosensors & bioelectronics. - 0956-5663 .- 1873-4235. ; 17:9, s. 747-759
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Tidskriftsartikel (refereegranskat)abstract
- In haemostatic and biomaterial research biological processes at surfaces and in the bulk phase of the surface-contacting medium are important. The present work demonstrates the usefulness of the combination of surface plasmon resonance (SPR), sensitive to changes in refractive index at surfaces, and free oscillation rheometry (FOR), sensitive to rheological properties of the bulk, for simultaneous real-time measurements on coagulation and fibrinolysis of blood plasma and coagulation of whole blood. SFLLRN stimulated coagulation of native whole blood presented a higher SPR signal with different appearance than plasma coagulation, while the FOR signals corresponding to plasma and whole blood coagulation were similar. This indicated that the SPR technique was more sensitive to cell-surface interactions than to fibrin formation in whole blood during coagulation, while the FOR technique were equally sensitive to coagulation in whole blood and plasma. Spontaneous coagulation of native whole blood in contact with methyl- and hydroxyl-terminated self-assembled monolayers (SAM) on gold and gold surfaces regenerated after coagulation were also studied. The regenerated gold surfaces displayed the shortest coagulation times, although the contact-activation of blood coagulation for these surfaces was low. The methylated and hydroxylated surfaces were comparable in terms of coagulation activation, while the hydroxylated surfaces presented FOR signals that indicated detaching of the coagulum from the surface. The combination of SPR and FOR is well suited for studies of cell– and protein–surface interactions and simultaneous bulk processes. Possible applications are investigations of blood cell defects in patients and monitoring of native whole blood interactions with artificial surfaces.
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| 10. |
- Hansson, Kenny, 1972-, et al.
(författare)
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Surface plasmon resonance detection of blood coagulation and platelet adhesion under venous and arterial shear conditions.
- 2007
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Ingår i: Biosensors & bioelectronics. - : Elsevier BV. - 0956-5663 .- 1873-4235. ; 23:2, s. 261-8
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Tidskriftsartikel (refereegranskat)abstract
- A surface plasmon resonance (SPR) based flow chamber device was designed for real time detection of blood coagulation and platelet adhesion in platelet rich plasma (PRP) and whole blood. The system allowed the detection of surface interactions throughout the 6mm length of the flow chamber. After deposition of thromboplastin onto a section of the sensor surface near the inlet of the flow chamber, coagulation was detected downstream of this position corresponding to a SPR signal of 7 to 8 mRIU (7 to 8 ng/mm2). A nonmodified control surface induced coagulation 3.5 times slower. Platelet adhesion to gold and fibrinogen coated surfaces in the magnitude of 1.25 and 1.66 mRIU was also shown with platelets in buffer, respectively. SPR responses obtained with PRP and whole blood on surfaces that were methylated or coated with von Willebrand factor (vWF), fibrinogen, or collagen, coincided well with platelet adhesion as observed with fluorescence microscopy in parallel experiments. The present SPR detection equipped flow chamber system is a promising tool for studies on coagulation events and blood cell adhesion under physiological flow conditions, and allows monitoring of short-range surface processes in whole blood.
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