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Ronquist, Gunnar
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Publications (10 of 29) Show all publications
Dubois, L., Löf, L., Larsson, A., Hultenby, K., Waldenström, A., Kamali-Moghaddam, M., . . . Ronquist, K. G. (2018). Human erythrocyte-derived nanovesicles can readily be loaded with doxorubicin and act as anticancer agents. Cancer Research Frontiers, 4(1), 13-26
Open this publication in new window or tab >>Human erythrocyte-derived nanovesicles can readily be loaded with doxorubicin and act as anticancer agents
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2018 (English)In: Cancer Research Frontiers, ISSN 2328-5249, Vol. 4, no 1, p. 13-26Article in journal (Refereed) Published
Abstract [en]

Purpose: In future therapeutics new formulas are needed that assure lower doses, fewer side effects, targeted administration and protection of the drug from degradation. In a first step to fulfil the requirements defined above, we carried out an in vitro study by developing a new procedure to encapsulate drugs using native vesicles first from prostasomes and then from erythrocyte membranes known to be well tolerated. The new method for production of drug delivery vesicles utilized osmotic loading of detergent resistant membranes (DRMs).

Materials and methods: DRMs of prostasomes and prepared human erythrocyte membranes were extracted and separated in a sucrose gradient at a density of 1.10 g/mL containing 1% Triton X-100. These DRMs were characterized by electron microscopy (transmission and scanning EM) and loaded with low and high molecular compounds. PC3 prostate cancer cells were treated with doxorubicin loaded DRMs in triplicate. DAPI (nuclear fluorescent stain) was included and fluorescence microscopic pictures were taken before the cells were trypsinized and counted after 48h.

Results: The content of the well separated band was observed ultrastructurally as small spherical, double layered membrane vesicles, (DRM vesicles) which harbored hyperosmolar sucrose of the gradient. Encapsulated hyperosmolar sucrose induced a transient osmotic lysis of the DRM vesicles when suspended in isotonic buffer containing loading molecules allowing vesicular inclusion. After this proof of concept, the method was finally employed for doxorubicin loading of DRM vesicles from human erythrocytes. When incubating such vesicles with PC3 cells a complete arrest of growth was observed in sharp contrast to PC3 cells incubated with plain doxorubicin in similar conditions.

Conclusion: The present results open up new possibilities for using DRM vesicles as drug delivery vesicles.

National Category
Clinical Laboratory Medicine
Identifiers
urn:nbn:se:uu:diva-364780 (URN)10.17980/2018.13 (DOI)
Note

Louise Dubois and Liza Löf contributed equally to this work.

Available from: 2018-11-02 Created: 2018-11-02 Last updated: 2019-01-31Bibliographically approved
Larssen, P., Wik, L., Czarnewski, P., Eldh, M., Löf, L., Ronquist, G., . . . Kamali-Moghaddam, M. (2017). Tracing Cellular Origin of Human Exosomes Using Multiplex Proximity Extension Assay. Molecular & cellular proteomics (online), 16(3), 502-511
Open this publication in new window or tab >>Tracing Cellular Origin of Human Exosomes Using Multiplex Proximity Extension Assay
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2017 (English)In: Molecular & cellular proteomics (online), ISSN 1535-9476, E-ISSN 1535-9484, Vol. 16, no 3, p. 502-511Article in journal (Refereed) Published
Abstract [en]

Extracellular vesicles (EVs) are membrane-coated objects such as exosomes and microvesicles, released by many cell-types. Their presence in body fluids and the variable surface composition and content render them attractive potential biomarkers. The ability to determine their cellular origin could greatly move the field forward. We used multiplex proximity extension assays (PEA) to identify with high specificity and sensitivity the protein profiles of exosomes of different origins, including seven cell lines and two different body fluids. By comparing cells and exosomes, we successfully identified the cells originating the exosomes. Furthermore, by principal component analysis of protein patterns human milk EVs and prostasomes released from prostate acinar cells clustered with cell lines from breast and prostate tissues, respectively. Milk exosomes uniquely expressed CXCL5, MIA and KLK6, while prostasomes carried NKX31, GSTP1 and SRC, highlighting that EVs originating from different origins express distinct proteins. In conclusion, PEA provides a powerful protein screening tool in exosome research, for purposes of identifying the cell source of exosomes, or new biomarkers in diseases such as cancer and inflammation.

National Category
Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-314142 (URN)10.1074/mcp.M116.064725 (DOI)000395670900013 ()28111361 (PubMedID)
Funder
Swedish Research CouncilEU, FP7, Seventh Framework Programme, 259796 294409Swedish Cancer Society, 2013/867Stockholm County Council, 20140405Swedish Heart Lung Foundation, 20140497The Karolinska Institutet's Research FoundationCancer and Allergy Foundation
Available from: 2017-01-28 Created: 2017-01-28 Last updated: 2017-04-20Bibliographically approved
Ronquist, K. G., Sanchez, C., Dubois, L., Chioureas, D., Fonseca, P., Larsson, A., . . . Panaretakis, T. (2016). Energy-requiring uptake of prostasomes and PC3 cell-derived exosomes into non-malignant and malignant cells. Journal of Extracellular Vesicles, 5, Article ID 29877.
Open this publication in new window or tab >>Energy-requiring uptake of prostasomes and PC3 cell-derived exosomes into non-malignant and malignant cells
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2016 (English)In: Journal of Extracellular Vesicles, ISSN 2001-3078, E-ISSN 2001-3078, Vol. 5, article id 29877Article in journal (Refereed) Published
Abstract [en]

Epithelial cells lining the prostate acini release, in a regulated manner (exocytosis), nanosized vesicles called prostasomes that belong to the exosome family. Prostate cancer cells have preserved this ability to generate and export exosomes to the extracellular space. We previously demonstrated that human prostasomes have an ATP-forming capacity. In this study, we compared the capacity of extracellular vesicles (EVs) to generate ATP between normal seminal prostasomes and exosomes secreted by PC3 cells (PC3 exosomes), a prostate cancer cell line. Proteomic analyses identified enzymes of the glycolytic chain in both prostasomes and PC3 exosomes, and we found that both of them were capable of generating ATP when supplied with substrates. Notably, the net production of extracellular ATP was low for prostasomes due to a high ATPase activity contrary to an elevated net ATP level for PC3 exosomes because of their low ATPase activity. The uptake of the 2 types of EVs by normal prostate epithelial cells (CRL2221) and prostate cancer cells (PC3) was visualized and measured, demonstrating differential kinetics. Interestingly, this uptake was dependent upon an ongoing glycolytic flux involving extracellular ATP formation by EVs and/or intracellular ATP produced from the recipient cells. We conclude that the internalization of EVs into recipient cells is an energy-requiring process also demanding an active V-ATPase and the capacity of EVs to generate extracellular ATP may play a role in this process.

Keywords
extracellular ATP; prostate cancer; extracellular vesicles; exosomes; prostasomes; ATPase; glycolysis; energy metabolism; exosome uptake
National Category
Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-281232 (URN)10.3402/jev.v5.29877 (DOI)000375921700001 ()26955882 (PubMedID)
External cooperation:
Available from: 2016-03-21 Created: 2016-03-21 Last updated: 2017-11-30Bibliographically approved
Carlsson, L., Ronquist, G., Elisasson, R., Dubois, L., Ronquist, K. G. & Larsson, A. (2016). High Concentrations of the Angiogenic Peptide VEGF-A in Seminal Fluid and its Association to Prostasomes. Clinical Laboratory, 62(8), 1515-1520
Open this publication in new window or tab >>High Concentrations of the Angiogenic Peptide VEGF-A in Seminal Fluid and its Association to Prostasomes
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2016 (English)In: Clinical Laboratory, ISSN 1433-6510, Vol. 62, no 8, p. 1515-1520Article in journal (Refereed) Published
Abstract [en]

Background: Angiogenesis is the formation of new blood vessels by capillary sprouting from pre-existing vessels. This process is associated with increased expression of angiogenic factors like vascular endothelial growth factor (VEGF). The VEGF family consists of five members denoted VEGF-A, B, C, D and placenta growth factor (PlGF). Prostasomes are exosome-like extracellular vesicles existing in seminal plasma. The present study aimed at investigating the possible relationship between VEGF-A in seminal fluid and blood plasma and the prostasomal association of VEGF-A.

Methods: Measurement of VEGF-A concentrations was carried out in seminal plasma from 40 males and in blood plasma from 40 male blood donors utilizing commercial ELISA kits. The prostasomal association of VEGF-A was investigated by flow cytometry.

Results: We found highly elevated concentrations of VEGF-A in seminal fluid (median value 150000 pg/mL) compared with those of blood plasma. Flow cytometric analysis showed that VEGF-A is bound to the surface of prostasomes.

Conclusions: Prostasomes and seminal plasma contain the angiogenic factor VEGF-A in high concentrations exceeding that of blood plasma by 1000 times.

Keywords
angiogenesis factor, ELISA, prostasomes, seminal fluid, vascular endothelial growth factor
National Category
Urology and Nephrology
Identifiers
urn:nbn:se:uu:diva-303148 (URN)10.7754/Clin.Lab.2016.151229 (DOI)000381716800017 ()28164613 (PubMedID)
Available from: 2016-09-15 Created: 2016-09-15 Last updated: 2017-11-21Bibliographically approved
Nickel, K. F., Ronquist, G., Langer, F., Labberton, L., Fuchs, T. A., Bokemeyer, C., . . . Renne, T. (2015). The polyphosphate-factor XII pathway drives coagulation in prostate cancer-associated thrombosis. Blood, 126(11), 1379-1389
Open this publication in new window or tab >>The polyphosphate-factor XII pathway drives coagulation in prostate cancer-associated thrombosis
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2015 (English)In: Blood, ISSN 0006-4971, E-ISSN 1528-0020, Vol. 126, no 11, p. 1379-1389Article in journal (Refereed) Published
Abstract [en]

Cancer is a leading cause of thrombosis. We identify a new procoagulant mechanism that contributes to thromboembolism in prostate cancer and allows for safe anticoagulation therapy development. Prostate cancer-mediated procoagulant activity was reduced in plasma in the absence of factor XII or its substrate of the intrinsic coagulation pathway factor XI. Prostate cancer cells and secreted prostasomes expose long chain polyphosphate on their surface that colocalized with active factor XII and initiated coagulation in a factor XII-dependent manner. Polyphosphate content correlated with the procoagulant activity of prostasomes. Inherited deficiency in factor XI or XII or high-molecular-weight kininogen, but not plasma kallikrein, protected mice from prostasome-induced lethal pulmonary embolism. Targeting polyphosphate or factor XII conferred resistance to prostate cancer-driven thrombosis in mice, without increasing bleeding. Inhibition of factor XII with recombinant 3F7 antibody reduced the increased prostasome-mediated procoagulant activity in patient plasma. The data illustrate a critical role for polyphosphate/factor XII-triggered coagulation in prostate cancer-associated thrombosis with implications for anticoagulation without therapy-associated bleeding in malignancies.

National Category
Cardiac and Cardiovascular Systems Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-267337 (URN)10.1182/blood-2015-01-622811 (DOI)000363479500019 ()26153520 (PubMedID)
Funder
Swedish Cancer Society, 100615Swedish Research Council, K2013-65X-21462-014-5Swedish Heart Lung Foundation, 20140741Stockholm County Council, 20140464German Research Foundation (DFG), SFB877German Research Foundation (DFG), SFB841EU, European Research Council, ERC-StG-2012-311575_F-12
Available from: 2015-11-24 Created: 2015-11-20 Last updated: 2017-12-01Bibliographically approved
Ronquist, G. (2012). Prostasomes are mediators of intercellular communication: from basic research to clinical implications. Journal of Internal Medicine, 271(4), 400-413
Open this publication in new window or tab >>Prostasomes are mediators of intercellular communication: from basic research to clinical implications
2012 (English)In: Journal of Internal Medicine, ISSN 0954-6820, E-ISSN 1365-2796, Vol. 271, no 4, p. 400-413Article, review/survey (Refereed) Published
Abstract [en]

Prostasomes are nanosized microvesicles secreted by acinar epithelial cells of the prostate gland. Furthermore, they are intracellular microvesicles inside another larger vesicle, a so-called storage vesicle, equivalent to multivesicular bodies of late endosomal origin. Prostasomes are thought to play an important role in intercellular communication by direct interaction primarily between the immobile acinar cells of the prostate gland and the mobile spermatozoa. Prostasomes transfer not only membrane components but also genetic material to spermatozoa. They are rich in various transferable bioactive molecules (e.g. receptors and enzymes) that promote the fertilizing ability of spermatozoa. In this review, the pleiotropic biological effects of prostasomes that are relevant for successful fertilization will be discussed. The ability to synthesize and export prostasomes to the extracellular space is observed not only in normal prostate epithelial cells but also in malignant prostate cells. Release of prostasomes by prostate cancer cells suggests a role in malignant cell growth and proliferation. These findings may provide new therapeutic and diagnostic strategies.

Keywords
exosomes, intercellular communication, microvesicles, prostasomes, prostate cancer, reproduction, spermatozoa
National Category
Urology and Nephrology
Identifiers
urn:nbn:se:uu:diva-167283 (URN)10.1111/j.1365-2796.2011.02487.x (DOI)000301927100010 ()22112042 (PubMedID)
Available from: 2012-01-24 Created: 2012-01-24 Last updated: 2017-12-08Bibliographically approved
Branth, S., Hambraeus, L., Piehl-Aulin, K., Essén-Gustavsson, B., Åkerfeldt, T., Olsson, R., . . . Ronquist, G. (2009). Metabolic stress-like condition can be induced by prolonged strenuous exercise in athletes. Upsala Journal of Medical Sciences, 114(1), 12-25
Open this publication in new window or tab >>Metabolic stress-like condition can be induced by prolonged strenuous exercise in athletes
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2009 (English)In: Upsala Journal of Medical Sciences, ISSN 0300-9734, E-ISSN 2000-1967, Vol. 114, no 1, p. 12-25Article in journal (Refereed) Published
Abstract [en]

Few studies have examined energy metabolism during prolonged, strenuous exercise. We wanted therefore to investigate energy metabolic consequences of a prolonged period of continuous strenuous work with very high energy expenditure. Twelve endurance-trained athletes (6 males and 6 females) were recruited. They performed a 7-h bike race on high work-load intensity. Physiological, biochemical, endocrinological, and anthropometric muscular compartment variables were monitored before, during, and after the race. The energy expenditure was high, being 5557 kcal. Work-load intensity (% of VO2 peak) was higher in females (77.7%) than in men (69.9%). Muscular glycogen utilization was pronounced, especially in type I fibres (>90%). Additionally, muscular triglyceride lipolysis was considerably accelerated. Plasma glucose levels were increased concomitantly with an unchanged serum insulin concentration which might reflect an insulin resistance state in addition to proteolytic glyconeogenesis. Increased reactive oxygen species (malondialdehyde (MDA)) were additional signs of metabolic stress. MDA levels correlated with glycogen utilization rate. A relative deficiency of energy substrate on a cellular level was indicated by increased intracellular water of the leg muscle concomitantly with increased extracellular levels of the osmoregulatory amino acid taurine. A kindred nature of a presumed insulin-resistant state with less intracellular availability of glucose for erythrocytes was also indicated by the findings of decreased MCV together with increased MCHC (haemoconcentration) after the race. This strenuous energy-demanding work created a metabolic stress-like condition including signs of insulin resistance and deteriorated intracellular glucose availability leading to compromised fuelling of ion pumps, culminating in a disturbed cellular osmoregulation indicated by taurine efflux and cellular swelling.

Keywords
Cellular swelling, energy expenditure, lipid peroxidation, metabolism, myocytes, taurine efflux
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-120869 (URN)10.1080/03009730802579778 (DOI)000263095500003 ()19242868 (PubMedID)
Available from: 2010-03-16 Created: 2010-03-16 Last updated: 2017-12-12Bibliographically approved
Ronquist, G., Carlsson, L., Ronquist, G., Semjonow, A., Wülfing, C. & Larsson, A. (2008). Serum antibodies against prostasomal clusterin in prostate cancer patients. Scandinavian Journal of Clinical and Laboratory Investigation, 68(3), 219-227
Open this publication in new window or tab >>Serum antibodies against prostasomal clusterin in prostate cancer patients
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2008 (English)In: Scandinavian Journal of Clinical and Laboratory Investigation, ISSN 0036-5513, E-ISSN 1502-7686, Vol. 68, no 3, p. 219-227Article in journal (Refereed) Published
Abstract [en]

Objective. Clusterin is a ubiquitous secretory sulphated glycoprotein present in prostasomes. It is an antiapoptotic mediator in prostate cancer and is among the most frequently occurring prostasomal proteins immunogenic in prostate cancer patients. The aim of the present study was to investigate the occurrence of anticlusterin antibodies in the serum of patients with prostate cancer and whether there is a relationship between anticlusterin antibody titres and other clinico-pathological variables. Material and methods. Serum samples were collected from 391 consecutive patients with suspected prostate cancer (150 benign prostate and 241 prostate cancer). The patients’ serum samples were used in an ELISA where microtitre wells were coated with purified clusterin from serum of a healthy volunteer. Flow cytometric studies of clusterin and prostasomes were performed. Results. Flow cytometric analyses revealed the presence of clusterin on the surface of seminal prostasomes. Anti-clusterin ELISA titres in sera of patients did not differ significantly from those of a control group. A significant ‘‘inverse’’ correlation existed between anti-clusterin ELISA titres and lymph node metastases (p50.047), but only 11 out of 161 patients had metastases. These titres correlated significantly with total prostate (p50.021) and transitional zone (p50.015) volumes of the patients. Conclusions. The correlation between serum anti-clusterin antibody titres and other clinico-pathological variables was generally weak in prostate cancer patients, although clusterin has been assigned an important role in tumourigenesis and progression of prostate cancer. However, the anti-clusterin antibody titre appeared to be related to prostate volume, correlating to both transitional zone volume and total volume of the prostate.

Keywords
Antibodies, clusterin, prostasomes, prostate cancer
National Category
Medical and Health Sciences
Research subject
Clinical Chemistry
Identifiers
urn:nbn:se:uu:diva-100731 (URN)10.1080/00365510701604602 (DOI)000255454500006 ()17926197 (PubMedID)
Available from: 2009-04-07 Created: 2009-04-06 Last updated: 2017-12-13Bibliographically approved
Branth, S., Ronquist, G., Stridsberg, M., Hambraeus, L., Kindgren, E., Olsson, R., . . . Arnetz, B. (2007). Development of abdominal fat and incipient metabolic syndrome in young healthy men exposed to long-term stress. NMCD. Nutrition Metabolism and Cardiovascular Diseases, 17(6), 427-435
Open this publication in new window or tab >>Development of abdominal fat and incipient metabolic syndrome in young healthy men exposed to long-term stress
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2007 (English)In: NMCD. Nutrition Metabolism and Cardiovascular Diseases, ISSN 0939-4753, E-ISSN 1590-3729, Vol. 17, no 6, p. 427-435Article in journal (Refereed) Published
Abstract [en]

BACKGROUND AND AIM: The sympathetic nervous system may be involved in the pathophysiology of insulin resistance and metabolic cardiovascular syndrome in young men. The aim was to study the effects of long-term stress on different features of the metabolic syndrome (MES) in formerly non-obese healthy young males during 5 months of defined conditions. METHODS AND RESULTS: Sixteen healthy male sailors (mean age 36.5 (SD)+/-7 years) participating in a sailing race around the world were recruited for the study. Investigations were done before the start and at stop overs after finishing laps 1, 2 and 4 (1, 2(1/2) and 5 months, respectively). Anthropometric and blood pressure data as well as biochemical data associated with MES were substantiated. Food intake and exercise were chartered and largely controlled. A mean weight loss of 4.5+/-2 kg (P<0.005), comprising both fat and lean body mass, was recorded during the first lap. Subsequently after 5 months, a weight gain, mainly consisting of 1.2+/-1.1 kg body fat (P<0.05), took place, concomitantly with a protein mass drop of 0.6+/-1.1 kg (P<0.05). The body fat gain accumulated on the abdominal region. Elevated blood levels of HbA1c, insulin and the triglycerides/high-density lipoprotein ratio were also observed during the race. Likewise heart rate and systolic blood pressure increased slightly but to a statistically significant extent. CONCLUSIONS: Non-obese healthy young men exposed to long-term stress developed abdominal obesity and signs of a metabolic syndrome in embryo, also emphasized by biochemical and blood pressure alterations. It is suggested that long-term and sustained stress activation might be an additional risk factor for the development of MES, even after control of dietary and exercise habits.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-15959 (URN)10.1016/j.numecd.2006.03.001 (DOI)000249711700003 ()17134957 (PubMedID)
Available from: 2008-04-15 Created: 2008-04-15 Last updated: 2017-12-08Bibliographically approved
Wiklund, L., Basu, S., Miclescu, A., Wiklund, P., Ronquist, G. & Sharma, H. S. (2007). Neuro- and cardioprotective effects of blockade of nitric oxide action by administration of methylene blue. In: Slikker W; Andrew RJ; Trembly B (Ed.), Neuroprotective agents: Eighth international neuroprotection society meeting. Paper presented at 8th International Conference on Neuroprotective Agents Mackinac Isl, MI, SEP 18-20, 2006 (pp. 231-244). , 1122(1)
Open this publication in new window or tab >>Neuro- and cardioprotective effects of blockade of nitric oxide action by administration of methylene blue
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2007 (English)In: Neuroprotective agents: Eighth international neuroprotection society meeting / [ed] Slikker W; Andrew RJ; Trembly B, 2007, Vol. 1122, no 1, p. 231-244Conference paper, Published paper (Refereed)
Abstract [en]

Methylene blue (MB), generic name methylthioninium (C16H18ClN3 S · 3H2O), is a blue dye synthesized in 1876 by Heinrich Caro for use as a textile dye and used in the laboratory and clinically since the 1890s, with well-known toxicity and pharmacokinetics. It has experimentally proven neuroprotective and cardioprotective effects in a porcine model of global ischemia–reperfusion in experimental cardiac arrest. This effect has been attributed to MB's blocking effect on nitric oxide synthase and guanylyl cyclase, the latter blocking the synthesis of the second messenger of nitric oxide. The physiological effects during reperfusion include stabilization of the systemic circulation without significantly increased total peripheral resistance, moderately increased cerebral cortical blood flow, a decrease of lipid peroxidation and inflammation, and less anoxic tissue injury in the brain and the heart. The last two effects are recorded as less increase in plasma concentrations of astroglial protein S-100β, as well as troponin I and creatine kinase isoenzyme MB, respectively.

Series
Annals of the New York academy of sciences, ISSN 0077-8923 ; 1122
Keywords
cardiac arrest, hemodynamics, methylene blue
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-15742 (URN)10.1196/annals.1403.016 (DOI)000252267100016 ()18077576 (PubMedID)978-1-57331-685-9 (ISBN)
Conference
8th International Conference on Neuroprotective Agents Mackinac Isl, MI, SEP 18-20, 2006
Available from: 2008-06-02 Created: 2008-06-02 Last updated: 2011-03-24Bibliographically approved
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