Bootsma, HJ

De Groot, R

De Jonge, MI

Langereis, JD

Pickering, MC

Ruseva, MM

Van de Kar, NC

Van den Heuvel, LP

Van der Flier, M

Van der Maten, E

Van Opzeeland, FJ

Van Selm, S

Westra, D

The Journal of Infectious Diseases

Maten, E. van der

Westra, D.

Selm, S. van

Langereis, J.D.

Bootsma, H.J.

Opzeeland, F.J.H. van

Groot, R. de

Ruseva, M.M.

Pickering, M.C.

Heuvel, L.P. van den

Kar, N.C.A.J. van de

Jonge, M.I. de

Flier, M. van der

NARCIS 

Complement Factor H Serum Levels Determine Resistance to Pneumococcal Invasive Disease

Radboud Repository

Spiral - Imperial College Digital Repository

Figure	1.	Clearance	of	Streptococcus	pneumoniae	from	plasma	is	enhanced	in	mice	heterozygous	for	the	gene	encoding	factor	H	(FH).	A–C,	Wild-type	(CA	+/+;	n	=	10),	heterozygous	(CA	+/−;	n	=	10),	and	homozygous	(CA	−/−	n	=	10)	FH-deficient	C57BL/6	mice	were	infected	with	1	×	107	 colony-forming	units	 (CFU)	of	S.	 pneumoniae	 (TIGR4)	 and	 sacrificed	19	hours	 later.	 #Four	of	 the	10	CA	−/−	animals	 were	 sacrificed	 before	 this	 because	 the	 predefined	 humane	 end	 point	 was	 reached.	 Nineteen	 hours	 aZer	 infec[on,	 the	number	of	CFU	per	milliliter	was	determined	in	blood,	and	levels	of	proinflammatory	cytokines	interleukin	6	(IL-6)	and	macrophage	inflammatory	 protein	 2	 (MIP-2)	 were	 measured	 in	 serum.	 Each	 point	 represents	 1	 mouse.	 Cytokine	 values	 were	 analyzed	 aZer	logarithmic	 transforma[on;	 horizontal	 line	 represents	 the	 median.	 Comparisons	 between	 groups	 were	 performed	 using	 the	nonparametric	Mann–Whitney	test	with	the	Bonferroni	correc[on.	*P	<	.05,	**P	<	.01,	and	***P	<	.001.				Van	der	Maten	et	al		Complement	Factor	H	Serum	Levels	Determine	Resistance	to	Pneumococcal	Invasive	Disease		J	Infect	Dis.	2016	Jun	1;213(11):1820-7.	doi:	10.1093/infdis/jiw029.	Epub	2016	Jan	21	Figure	2.	Administra[on	of	human	factor	H	(FH)	to	wild-type	mice	impaired	plasma	clearance	of	Streptococcus	pneumoniae.	A–C,	Wild-type	C57BL/6	mice	were	intravenously	infected	with	1	×	107	colony-forming	units	of	S.	pneumoniae	(TIGR4)	and	injected	with	purified	human	FH	(n	=	10)	or	phosphate-buffered	saline	(n	=	10)	and	sacrificed	21	hour	later.	Twenty-one	hours	aZer	infec[on,	the	number	of	colony-forming	units	(CFU)	per	milliliter	was	determined	in	blood,	and	levels	of	the	proinflammatory	cytokines	IL-6	and	MIP-2	were	measured	in	serum.	Each	point	represents	one	mouse;	cytokine	values	were	analyzed	aZer	logarithmic	transforma[on;	horizontal	lines	represents	the	median.	Comparisons	between	groups	were	performed	using	the	nonparametric	Mann-Whitney	test	with	the	Bonferroni	correc[on.	*P	<	.05,	**P	<	.01,	and	***P	<	.001.			Van	der	Maten	et	al		Complement	Factor	H	Serum	Levels	Determine	Resistance	to	Pneumococcal	Invasive	Disease		J	Infect	Dis.	2016	Jun	1;213(11):1820-7.	doi:	10.1093/infdis/jiw029.	Epub	2016	Jan	21	Figure	3.	Pneumococcal	C3	opsoniza[on	is	enhanced	in	mice	heterozygous	for	the	gene	encoding	factor	H	(FH).	C3	deposi[on	on	the	surface	of	Streptococcus	pneumoniae	strain	TIGR4	in	pooled	mouse	serum	derived	from	CA	+/+,	CA	+/−,	or	CA	−/−	mice.	A	total	of	5	×	 107	 colony-forming	 units/mL	 S.	 pneumoniae	 TIGR4	 were	 incubated	 in	 20%	mice	 serum	 in	 veronal	 buffer	 containing	Mg2+	 and	ethylene	 glycol	 tetraace[c	 acid.	 AZer	 90	 minutes	 of	 incuba[on	 at	 37°C,	 pneumococcal	 C3	 deposi[on	 was	 measured	 by	 flow	cytometry.	 A,	 A	 representa[ve	 example	 of	 a	 flow	 cytometry	 histogram	 for	 bacteria	 incubated	 in	 wild-type	 (FH+/+;	 white	 peak),	heterozygous	 (FH+/−;	 gray	 peak),	 and	 homozygous	 (FH−/−;	 black	 peak)	 FH-deficient	 pooled	 mouse	 serum.	 B,	 Intensity	 of	 C3	deposi[on	on	bacteria.	C,	Propor[on	of	C3-posi[ve	bacteria.	For	panels	B	and	C,	each	bar	represents	the	mean	±	standard	error	of	the	mean	for	results	obtained	from	5	separate	experiments.	Comparisons	between	groups	were	performed	using	a	paired	Student	t	test.*P	<	.05,	**P	<	.01,	***P	<	.001.	Abbrevia[on:	NS,	not	significant.				Van	der	Maten	et	al		Complement	Factor	H	Serum	Levels	Determine	Resistance	to	Pneumococcal	Invasive	Disease		J	Infect	Dis.	2016	Jun	1;213(11):1820-7.	doi:	10.1093/infdis/jiw029.	Epub	2016	Jan	21	Figure	4.	Pneumococcal	C3	opsoniza[on	in	wild-type	mice	serum	is	 impaired	by	excess	human	factor	H	(FH).	C3	deposi[on	on	the	surface	of	S.	pneumoniae	strain	TIGR4	 in	pooled	wild-type	mouse	serum	with	addi[onal	human	FH.	S.	pneumoniae	TIGR4,	5	×	107	colony-forming	units/mL,	were	incubated	in	20%	mice	serum	in	Veronal	buffer	containing	Mg2+	and	ethylene	glycol	tetraace[c	acid.	AZer	90	minutes	of	incuba[on	at	37°C,	pneumococcal	C3	deposi[on	was	measured	by	flow	cytometry.	A,	A	representa[ve	example	of	a	flow	cytometry	histogram	for	bacteria	incubated	in	wild-type	(CA+/+;	white	peak),	wild-type	with	exogenous	human	FH	(CA	+/+	[+FH];	gray	peak),	and	heat-inac[vated	mouse	serum	(HI)	as	a	nega[ve	control	(black	peak).	B,	Intensity	of	C3	deposi[on	on	bacteria.	C,	Propor[on	of	C3-posi[ve	bacteria.	The	bars	 in	panels	B	and	C	 represent	 the	mean	±	standard	error	of	 the	mean	of	2	 separate	experiments	in	duplicate.	Comparisons	between	groups	were	performed	using	a	paired	Student	t	test.	*P	<	.05,**P	<	.01,	***P	<	.001.	Abbrevia[on:	NS,	not	significant.			Van	der	Maten	et	al		Complement	Factor	H	Serum	Levels	Determine	Resistance	to	Pneumococcal	Invasive	Disease		J	Infect	Dis.	2016	Jun	1;213(11):1820-7.	doi:	10.1093/infdis/jiw029.	Epub	2016	Jan	21	Figure	5.	Alterna[ve	pathway	modula[on	by	human	factor	H	(FH)	levels	determines	pneumococcal	C3	opsoniza[on.	C3	deposi[on	on	the	surface	of	Streptococcus	pneumoniae	strain	TIGR4	in	human	FH–depleted	serum	recons[tuted	with	various	FH	concentra[ons.	S.	pneumoniae	TIGR4,	5	×	107	colony-forming	units/mL,	were	 incubated	 in	10%	human	serum	 in	Veronal	buffer	containing	Mg2+	and	ethylene	 glycol	 tetraace[c	 acid.	 AZer	 30	 minutes	 of	 incuba[on	 at	 37°C,	 pneumococcal	 C3	 deposi[on	 was	 measured	 by	 flow	cytometry.	A,	A	representa[ve	example	of	a	flow	cytometry	histogram	for	bacteria	 incubated	 in	FH-depleted	serum	recons[tuted	with	the	physiological	FH	concentra[on	of	500	μg/mL	FH	(light	gray),	or	0	μg/mL	(black),	300	μg/mL	(white),	or	1000	μg/mL	(dark	gray)	FH.	B,	Propor[on	of	C3-posi[ve	cells.	Each	bar	represents	the	mean	±	standard	error	of	the	mean	of	results	obtained	from	4	separate	experiments.	Comparison	of	various	FH	concentra[ons	to	the	physiological	FH	concentra[on	of	500	μg/mL	(gray	bar)	were	performed	by	using	a	repeated	measures	analysis	of	variance	with	the	Bonferroni	correc[on.	*P	<	.05,	**P	<	.01,	and	***P	<	.001.	Abbrevia[on:	NS,	not	significant			Van	der	Maten	et	al		Complement	Factor	H	Serum	Levels	Determine	Resistance	to	Pneumococcal	Invasive	Disease		J	Infect	Dis.	2016	Jun	1;213(11):1820-7.	doi:	10.1093/infdis/jiw029.	Epub	2016	Jan	21	Figure	6.	Alterna[ve	pathway	modula[on	by	human	factor	H	(FH)	determines	pneumococcal	killing	in	blood.	Pneumococcal	killing	in	human	blood	containing	FH-depleted	serum	recons[tuted	with	various	FH	concentra[ons.	Streptococcus	pneumoniae	TIGR4,	1	×	106	colony-forming	 units/mL,	 were	 incubated	 in	 healthy	 donor	 blood	 in	 which	 plasma	 was	 replaced	 by	 human	 FH–depleted	 serum	recons[tuted	with	various	FH	concentra[ons	or	with	heat-inac[vated	(HI)	serum	as	a	control.	The	number	of	colony-forming	units	were	 determined	 by	 pla[ng	 serial	 dilu[ons	 before	 incuba[on	 and	 240	minutes	 aZer	 incuba[on	 at	 37°C,	 and	 the	 percentage	 of	surviving	bacteria	was	calculated.	Each	dot	represents	1	blood	donor.	Comparisons	of	various	FH	concentra[ons	to	the	physiological	FH	concentra[on	of	500	μg/mL	 (white	dots)	were	performed	using	a	 repeated	measures	analysis	of	 variance	with	 the	Bonferroni	correc[on.	*P	<	.05	was	considered	significant.	**P	<	.01	and	***P	<	.001.		Van	der	Maten	et	al		Complement	Factor	H	Serum	Levels	Determine	Resistance	to	Pneumococcal	Invasive	Disease		J	Infect	Dis.	2016	Jun	1;213(11):1820-7.	doi:	10.1093/infdis/jiw029.	Epub	2016	Jan	21	

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Complement factor H serum levels determine resistance to pneumococcal invasive disease

Complement factor H serum levels determine resistance to pneumococcal invasive disease

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