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Ig-binding capacity of carp macrophages
The Scientific & Social Program: Vth ISDCI Congress
$85
P17 Ig-BINDING CAPACITY OF CARP M A C R O P H A G E S .
"J.H.W.M.Rombout,J.C.E. van Diepen, M.H.M. van Lisdonk,JJ. Taverne-Thiele& B.M.Verburg-vanKemenade. Department of Experimental Animal Morphology and Cell Biology, Agricultural University, P.O. Box 338, 6700 AH Wageningen, The Netherlands. Fc-receptor-mediated Ig-bindingis generallyaccepted for macrophagesof higher vertebrates. In teleo~ts, only indications are available for this phenomenon. In carp, melanomacrophages and intestinal macrophages appeared to be immunoreactive with a monoclonal antibody against carp serum Ig (WCI 12). This study deals with the Ig-bindingcapacity of carp macrophages,in order to explain their Ig-immunoreaetivity.Isolated intestinal lymphocytes and macrophages challenged with carp Ig-coated SRBC revealed much more rosettes compared with a control challenge (uncoated SRBC). As characterization of rosette-formingcells was laborious, we developed a procedure that allowed us to study the morphology of Ig-immunoreaetivecells in combination with their Ig-binding capacity. Firstly lymphocytes and macrophages were labelled in suspension with WCI 12 and subsequentlywith a fluorescent or gold marker. After intemalisation of the label (lh at room temperature in a medium without azide) cells were incubated with carp Ig. Finally, cells were immunostained again with WCI 12 and subs~xl,~ntlywith a ditIerent lluoresceat or gold marker. The whole procedureexcept the intemalisatioa step was carried out on ice in azide-containino medium. Cells were studied with the fluorescence microscope or electron microscope.Ig-bindingwas only demonstrated for macrophagesby the presence of bothinternalized and surface labels. Reactions without the addition of carp Ig revealed only internalised labels in macrophages and a minority of lymphocytes.The proportion of Ig-bindingmacrophages was much hi~her in cell suspensions of gut than of spleen or head kidney.
P18 P H A G O C Y T O S I S OF YEAST G L U C A N PARTICLES BY A T L A N T I C SALMON MACROPHAGES. *J.Jorgensen, B.Robertsen & R.Engstad.The N o r w e g i a n College of Fishery Science, University of Tromso, N-9000 Tromso, Tromsa, N o r w a y . Yeast glucan particles have been s h o w n to enhance the non-specific disease resistance of Atlantic salmon, Salmo salar L. (B. Robertsen et a1,1990, J.Fish Diseases 13, 391-400). As m a c r o p h a g e s are t h o u g h t to be i m p o r t a n t in this process, w e d e c i d e d to s t u d y u p t a k e of glucan particles b y s a l m o n m a c r o p h a g e s . P r o n e p h r o s m a c r o p h a g e s w e r e isolated, cultured on a s e r u m free m e d i u m a n d incubated with glucan particles u n d e r various conditions. At the end of the incubation the m a c r o p h a g e s w e r e fixed, stained with Giemsa a n d the percentage of m a c r o p h a g e s containing glucan particles estimated microscopically. Fifty to sixty % of the m a c r o p h a g e s w e r e f o u n d to contain glucan particles after 30 rain of incubation. The u p t a k e of particles was inhibited b y a range of different soluble neutral polysaccharides w h i c h indicates that the majority of glucan particles are p h a g o c y t i z e d b y non-specific u p t a k e mechanism. O n the other hand, the most effective inhibitor of glucan particle u p t a k e was a soluble glucan obtained b y partial formolysis of glucan particles. This indicate that the m a c r o p h a g e s m a y also h a v e specific receptors for yeast glucans.
$85
P17 Ig-BINDING CAPACITY OF CARP M A C R O P H A G E S .
"J.H.W.M.Rombout,J.C.E. van Diepen, M.H.M. van Lisdonk,JJ. Taverne-Thiele& B.M.Verburg-vanKemenade. Department of Experimental Animal Morphology and Cell Biology, Agricultural University, P.O. Box 338, 6700 AH Wageningen, The Netherlands. Fc-receptor-mediated Ig-bindingis generallyaccepted for macrophagesof higher vertebrates. In teleo~ts, only indications are available for this phenomenon. In carp, melanomacrophages and intestinal macrophages appeared to be immunoreactive with a monoclonal antibody against carp serum Ig (WCI 12). This study deals with the Ig-bindingcapacity of carp macrophages,in order to explain their Ig-immunoreaetivity.Isolated intestinal lymphocytes and macrophages challenged with carp Ig-coated SRBC revealed much more rosettes compared with a control challenge (uncoated SRBC). As characterization of rosette-formingcells was laborious, we developed a procedure that allowed us to study the morphology of Ig-immunoreaetivecells in combination with their Ig-binding capacity. Firstly lymphocytes and macrophages were labelled in suspension with WCI 12 and subsequentlywith a fluorescent or gold marker. After intemalisation of the label (lh at room temperature in a medium without azide) cells were incubated with carp Ig. Finally, cells were immunostained again with WCI 12 and subs~xl,~ntlywith a ditIerent lluoresceat or gold marker. The whole procedureexcept the intemalisatioa step was carried out on ice in azide-containino medium. Cells were studied with the fluorescence microscope or electron microscope.Ig-bindingwas only demonstrated for macrophagesby the presence of bothinternalized and surface labels. Reactions without the addition of carp Ig revealed only internalised labels in macrophages and a minority of lymphocytes.The proportion of Ig-bindingmacrophages was much hi~her in cell suspensions of gut than of spleen or head kidney.
P18 P H A G O C Y T O S I S OF YEAST G L U C A N PARTICLES BY A T L A N T I C SALMON MACROPHAGES. *J.Jorgensen, B.Robertsen & R.Engstad.The N o r w e g i a n College of Fishery Science, University of Tromso, N-9000 Tromso, Tromsa, N o r w a y . Yeast glucan particles have been s h o w n to enhance the non-specific disease resistance of Atlantic salmon, Salmo salar L. (B. Robertsen et a1,1990, J.Fish Diseases 13, 391-400). As m a c r o p h a g e s are t h o u g h t to be i m p o r t a n t in this process, w e d e c i d e d to s t u d y u p t a k e of glucan particles b y s a l m o n m a c r o p h a g e s . P r o n e p h r o s m a c r o p h a g e s w e r e isolated, cultured on a s e r u m free m e d i u m a n d incubated with glucan particles u n d e r various conditions. At the end of the incubation the m a c r o p h a g e s w e r e fixed, stained with Giemsa a n d the percentage of m a c r o p h a g e s containing glucan particles estimated microscopically. Fifty to sixty % of the m a c r o p h a g e s w e r e f o u n d to contain glucan particles after 30 rain of incubation. The u p t a k e of particles was inhibited b y a range of different soluble neutral polysaccharides w h i c h indicates that the majority of glucan particles are p h a g o c y t i z e d b y non-specific u p t a k e mechanism. O n the other hand, the most effective inhibitor of glucan particle u p t a k e was a soluble glucan obtained b y partial formolysis of glucan particles. This indicate that the m a c r o p h a g e s m a y also h a v e specific receptors for yeast glucans.