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Isolation of murine intrahepatic immune cells employing a modified procedure for mechanical disruption and functional characterization of the B, T and natural killer T cells obtained.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
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2009 (English)In: Clinical and experimental immunology, ISSN 1365-2249, Vol. 155, no 2, 320-9 p.Article in journal (Refereed) Published
Abstract [en]

Intrahepatic immune cells (IHIC) are known to play central roles in immunological responses mediated by the liver, and isolation and phenotypic characterization of these cells is therefore of considerable importance. In the present investigation, we developed a simple procedure for the mechanical disruption of mouse liver that allows efficient isolation and phenotypic characterization of IHIC. These cells are compared with the corresponding cells purified from the liver after enzymatic digestion with different concentrations of collagenase and DNase. The mechanical disruption yielded viable IHIC in considerably greater numbers than those obtained following enzymatic digestion. The IHIC isolated employing the mechanical disruption were heterogeneous in composition, consisting of both innate and adaptive immune cells, of which B, T, natural killer (NK), NK T cells, granulocytes and macrophages were the major populations (constituting 37.5%, 16.5%, 12.1%, 7.9%, 7.9% and 7.5% of the total number of cells recovered respectively). The IHIC obtained following enzymatic digestion contained markedly lower numbers of NK T cells (1.8%). The B, T and NK T cells among IHIC isolated employing mechanical disruption were found to be immunocompetent, i.e. they proliferated in vitro in response to their specific stimuli (lipopolysaccharide, concanavalin A and alpha-galactosylceramide respectively) and produced immunoglobulin M and interferon-gamma. Thus, the simple procedure for the mechanical disruption of mouse liver described here results in more efficient isolation of functionally competent IHIC for various types of investigation.

Place, publisher, year, edition, pages
2009. Vol. 155, no 2, 320-9 p.
URN: urn:nbn:se:su:diva-34717DOI: 10.1111/j.1365-2249.2008.03815.xISI: 000262282400024PubMedID: 19040612OAI: diva2:285339
Available from: 2010-01-11 Created: 2010-01-11 Last updated: 2011-10-17Bibliographically approved
In thesis
1. Immunohepatotoxicity of the persistent environmental pollutants perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS)
Open this publication in new window or tab >>Immunohepatotoxicity of the persistent environmental pollutants perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS)
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS), manufactured for a variety of industrial and consumer applications, are ubiquitous environmental pollutants. Their accumulation in humans and wildlife raises serious health concerns.

Here, we examined the potential effects of PFOA and PFOS on the innate immune system in mice. Short-term dietary exposure to high doses reduces the total number and subpopulations of circulating white blood cells. Moreover, production of proinflammatory cytokines by macrophages in the peritoneal cavity and bone marrow, but not in the spleen following exposure to in vitro or in vivo stimulation by bacterial lipopolysaccharides is enhanced. With respect to adaptive immunity, PFOS reduces the total numbers of thymocytes and splenocytes and subpopulations thereof in a dose dependent fashion. Furthermore, comparison of wild-type mice and the corresponding knock-out strain lacking peroxisome proliferator-activated receptor-alpha revealed that these immunological changes are partially dependent on this receptor. Our further studies also show that sub-chronic dietary exposure to an environmentally relevant dose of PFOS does not alter the cellularity of the thymus and spleen and exerts no influence on humoral immune responses.

To facilitate examination of the effects of PFOA and PFOS on the hepatic immune system, we developed a procedure for mechanical disruption that yields a larger number of functionally competent immune cells from this organ. In our last study, lower doses of PFOA or PFOS induced hypertrophy of hepatocytes and altered the hepatic immune status. Thus, we find that short-term, high- and low-dose exposure of mice to these fluorochemicals is immunohepatotoxic.

Abstract [sv]

Perfluorooktanat (PFOA) och perfluorooktansulfonat (PFOS) som tillverkas för många olika industri och konsumentprodukter, är globalt förekommande miljögifter. Deras ackumulering i människor och djur ger upphov till en stark oro för hälsoproblem.

Vi har granskat effekterna av PFOA och PFOS på det medfödda, ospecifika immunförsvaret. Exponering för höga doser via maten under kort tid minskar det totala antalet cirkulerande vita blodkroppar samt delpopulationerna.. Immunsvaret ökar dock efter stimulering med bakteriella lipopolysaccharider både in vitro och in vivo , dvs produktionen av proinflammatoriska cytokiner av makrofager i bukhålan och benmärgen, men inte i mjälten ökar.. När det gäller adaptiv, specifik immunitet minskar PFOS det totala antalet tymocyter och splenocyter och deras olika subpopulationer. Vid exponering för lägre doser av PFOS induceras hepatomegali utan att påverka tymus eller mjälten.   Vi kunde visa att peroxisomal proliferator-aktiverad receptor-alfa medierar effekterna utav PFOS i tymus samt delar av effekterna av PFOS i mjälten genom att använda möss som saknade denna receptor. . Dettastöds av vår studie med subkronisk exponering för en miljömässig dos av PFOS vilken inte ändrade den cellulära sammansättningen i vare sig  tymus eller mjälte och inte hade  något inflytande på det humorala immunsvaret.

För att underlätta studier av hur PFOA och PFOS påverkar immunsystemet i levern utvecklade vi en metod för framrening av immunceller via mekanisk sönderdelning av levern, vilket gavett större antal av funktionella  immunceller från detta organ. I vår sista studie kunde vi påvisa att lägre doser av PFOA eller PFOS inducerade hypertrofi av hepatocyter samt en påverkan av leverns immunförsvar.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2011. 126 p.
Perfluorooctanoate, Perfluorooctane sulfonate, Peroxisome proliferator-activated receptor-alpha, Hepatomegaly, Immunotoxicity, Thymus, Spleen
National Category
Natural Sciences
Research subject
urn:nbn:se:su:diva-63180 (URN)978-91-7447-381-0 (ISBN)
Public defence
2011-11-21, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Available from: 2011-10-30 Created: 2011-10-11 Last updated: 2011-11-15Bibliographically approved

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Qazi, M RahmanNelson, B DDePierre, J WAbedi-Valugerdi, M
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