Journal of Medical Microbiology Papers in Press. Published June 11, 2015 as doi:10.1099/jmm.0.000099

Journal of Medical Microbiology Infections and cardiovascular disease: is Bartonella henselae contributing to this matter? --Manuscript Draft-Manuscript Number:

JMM-D-14-00238

Full Title:

Infections and cardiovascular disease: is Bartonella henselae contributing to this matter?

Short Title:

Infectious diseases, Bartonella henselae and cardiovascular disease

Article Type:

Review

Section/Category:

Clinical microbiology and virology

Keywords:

Infectious diseases; Bartonella henselae; cardiovascular disease; inflammation; atherosclerosis

Corresponding Author:

Francesco Paolo Mancini, M.D., Ph.D. University of Sannio Benevento, ITALY

First Author:

Paola Salvatore, M.D., Ph.D.

Order of Authors:

Paola Salvatore, M.D., Ph.D. Alberto Zullo, Ph.D. Linda Sommese, Biol.D. Roberta Colicchio, Ph.D. Antonietta Picascia, Biol.D. Concetta Schiano, Ph.D. Francesco Paolo Mancini, M.D., Ph.D. Claudio Napoli, M.D., Ph.D.

Manuscript Region of Origin:

ITALY

Abstract:

Cardiovascular disease is still the major cause of death worldwide despite the remarkable progress in its prevention and treatment. Endothelial progenitor cells have recently emerged as key players of vascular repair and regenerative medicine applied to cardiovascular disease. A large amount of effort has been put into discovering the factors that could aid or impair the number and function of endothelial progenitor cells, and also characterizing at molecular level these cells in order to facilitate their therapeutic applications in vascular disease. Interestingly, the major cardiovascular risk factors have been associated with reduced number and function of endothelial progenitor cells. The bacterial contribution cardiovascular disease represents a longstanding controversy. The discovery that Bartonella henselae can infect and damage endothelial progenitor cells revitalizes the enduring controversy about the microbiological contribution to atherosclerosis, thus allowing the hypothesis that this infection could impair the cardiovascular regenerative potential increasing the risk for cardiovascular disease. In this review, we summarize the rationale to suggest that Bartonella henselae could favor atherogenesis by infecting and damaging endothelial progenitor cells, thus reducing their vascular repair potential. These mechanisms suggest a novel link between communicable and non-communicable human diseases, and put forward the possibility that Bartonella henselae could enhance the susceptibility and worsen the prognosis in cardiovascular disease.

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1

Infections and cardiovascular disease: is Bartonella henselae contributing to this matter?

2 3

Paola Salvatore1,2, Alberto Zullo2,3, Linda Sommese4,5, Roberta Colicchio1, Antonietta Picascia1,4,

4

Concetta Schiano6, Francesco Paolo Mancini3* and Claudio Napoli4,6

5 6 7

1

8

II, Naples, Italy

9

2

CEINGE-Advanced Biotechnologies, Naples, Italy

10

3

Department of Sciences and Technologies, University of Sannio, Benevento, Italy

11

4

U.O.C. Division of Immunohematology, Transfusion Medicine and Transplant Immunology

12

[SIMT], Regional Reference Laboratory of Transplant Immunology [LIT], Azienda Universitaria

13

Policlinico (AOU) and Department of Biochemistry, Biophysics and General Pathology, Second

14

University of Naples, Naples, Italy

15

5

16

Naples, Italy

17

6

Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico

Department of Experimental Medicine, Section of Microbiology, Second University of Naples,

Foundation SDN, Institute of Diagnostic and Nuclear Development, IRCCS, Naples, Italy

18 19

Contents Category: Review article

20 21

Running title: Infectious diseases, Bartonella henselae and cardiovascular disease

22 23

*Corresponding author: Francesco P. Mancini, via Port’Arsa, 11, 82100, Benevento, Italy - Tel.

24

+39.0824.305107, fax +39.0824.23013, e-mail: [email protected]

25 26 27 28 29 30 31

Abbreviations: CVD, cardiovascular disease; EPCs, endothelial progenitor cells; ECs, endothelial cells; vascular endothelial growth factor, VEGF; angiopoietin-1,Ang-1; vascular endothelial growth factor receptor, VEGFR; myocardial infarction, MI; bacillary angiomatosis, BA; bacillary peliosis, BP; infectious endocarditis, IE; blood-culture-negative endocarditis, BCNE; hematopoietic stem cells, HSCs;

32 33 34 1

1 2

Abstract

3

Cardiovascular disease is still the major cause of death worldwide despite the remarkable progress

4

in its prevention and treatment. Endothelial progenitor cells have recently emerged as key players of

5

vascular repair and regenerative medicine applied to cardiovascular disease. A large amount of

6

effort has been put into discovering the factors that could aid or impair the number and function of

7

endothelial progenitor cells, and also characterizing at molecular level these cells in order to

8

facilitate their therapeutic applications in vascular disease. Interestingly, the major cardiovascular

9

risk factors have been associated with reduced number and function of endothelial progenitor cells.

10

The bacterial contribution cardiovascular disease represents a long-standing controversy. The

11

discovery that Bartonella henselae can infect and damage endothelial progenitor cells revitalizes the

12

enduring controversy about the microbiological contribution to atherosclerosis, thus allowing the

13

hypothesis that this infection could impair the cardiovascular regenerative potential increasing the

14

risk for cardiovascular disease. In this review, we summarize the rationale to suggest that

15

Bartonella henselae could favor atherogenesis by infecting and damaging endothelial progenitor

16

cells, thus reducing their vascular repair potential. These mechanisms suggest a novel link between

17

communicable and non-communicable human diseases, and put forward the possibility that

18

Bartonella henselae could enhance the susceptibility and worsen the prognosis in cardiovascular

19

disease.

20 21

2

1

Introduction

2

Cardiovascular disease (CVD) is the primary cause of death all around the world and in 2008

3

accounted for 30% of all-cause mortality (World Health Organization, 2011). CVD, and, more

4

precisely atherosclerotic CVD, is predicted to remain the single leading cause of death (23.3 million

5

deaths by 2030) (Mathers & Loncar, 2006). Despite major risk factors for CVD have been firmly

6

established, still a quarter of people presenting the disease do not show any of the known

7

cardiovascular risk factors (Magnus & Beaglehole, 2001). Therefore, there is considerable interest

8

to look for novel components affecting cardiovascular health, especially for those that could

9

improve global cardiovascular risk prediction. In fact, the lipid hypothesis, which recognizes

10

elevated plasma levels of low-density lipoporotein cholesterol as the primary cause of

11

atherosclerosis and consequent CVD, is evolving and in the last couple of decades a large credit is

12

given to the potential role of inflammation in atherogenesis (Ross, 1999).

13

Based upon the immuno-inflammatory response elicited by infectious disease, the microbiological

14

contribution to atherosclerosis, the pathological basis of the vast majority of CVD, represents a

15

long-standing controversy (Nieto, 2002; Rosenfeld & Campbell, 2011). An abundance of data

16

suggests that several infectious agents, as Chlamydophila pneumoniae, Helicobacter pylori,

17

cytomegalovirus and periodontal pathogens, might contribute to atherosclerotic vascular diseases

18

(Chatzidimitriou et al., 2012). However, lowering plasma cholesterol reduces overall and

19

cardiovascular mortality (Steinberg, 2006), antibiotic treatments failed to do so (Cannon et al.,

20

2005; Grayston et al., 2005; O'Connor et al., 2003). Nevertheless, the finding that the bacterium

21

Bartonella henselae can infect endothelial progenitor cells (EPCs) (Salvatore et al., 2008) suggests

22

an alternative mechanism for infectious diseases to promote atherosclerotic CVD. Indeed, B.

23

henselae-infected EPCs could reduce the endothelial repair and replacement potential with the

24

consequent enhancement of vascular inflammation and degeneration. Therefore, this recent

25

evidence could relate infective agents, regenerative medicine and CVD.

26

Regenerative medicine aims at restoring the original structure and function of damaged tissues by

27

using tissue-specific progenitor cells and avoiding fibrosis and scar formation. The societal burden 3

1

imposed by CVD could particularly benefit from regenerative medicine, which might help both in

2

preventing and curing the disease.

3

Presently, it is prudent to indicate vascular maintenance and repair as the most likely possibility to

4

counteract the onset and development of CVD, while de novo vascularization of ischemic tissue

5

remains still a distant chimera.

6

The aim of this review is to investigate the possibility that some types of infection, by modifying

7

the susceptibility to and/or the course of CVD, could represent novel risk or prognostic factors of

8

CVD.

9 10

Infectious disease and CVD

11

The pathological evidence of inflammatory changes within the arterial wall has prompted scientists

12

to recognize the inflammatory component of the atherosclerotic CVD since 150 years ago (Mayerl,

13

2006). Indeed, accumulation of macrophages and activated T lymphocytes, cytokine production and

14

consequent increase of the expression of adhesion molecule on the surface of the overlaying

15

dysfunctional endothelium, are all characteristics that were identified in the atherosclerotic plaque

16

(Ross, 1999). The lipid-induced transformation of macrophages into foam cells, the proliferation of

17

smooth muscle cell and the production of matrix metalloproteinases (MMPs) are also crucial events

18

in the evolution of the plaque. In particular, MMPs produce fissuring of the arterial wall by

19

breaking down intramural deposits of collagen and elastin; this, in turn, causes the adherence of

20

platelets and activation of the coagulation cascade at the lesion site leading to thrombosis, major

21

occlusion of the atherosclerotic vessel, and consequent myocardial infarction or cerebrovascular

22

accident (Libby et al., 2009). This novel basic knowledge has translated into clinical application:

23

indeed, enhancement of the assessment of the cardiovascular risk was obtained by taking into

24

account also the plasma level of C-reactive protein (CRP), a general inflammatory marker secreted

25

by the liver in response to interleukin-6 stimulation (Ridker et al., 2008).

26

The inflammatory hallmarks of atherosclerosis have driven many researchers to study the

27

hypothesis that microbial infections could cause, accelerate, or aggravate the atherosclerotic CVD. 4

1

About four decades ago, virus-induced cholesterol crystals were observed in cultured cells and such

2

crystals are known to form when cholesterol accumulate in the plaque and to activate

3

inflammasomes (Fabricant et al., 1973; Duewell et al., 2010). Moreover viral infection was shown

4

to induce experimental atherosclerosis and also to alter aortic cholesterol metabolism and

5

accumulation (Fabricant et al., 1978; Hajjar et al., 1986).

6

Interestingly, repeated inoculations of C. pneumoniae increased lipid accumulation in the aortic

7

sinus of normocholesterolemic mice (Törmäkangas et al., 2005). A number of epidemiological

8

studies have demonstrated an association between infections with C. pneumoniae, cytomegalovirus

9

and Helicobacter pylori, and the presence of coronary and carotid atherosclerosis (Grayston, 2000;

10

Mayr et al., 2000; Melnick et al., 1993; Ameriso et al., 2001; Franceschi et al, 2002; He et al.,

11

2014). Very impressive data were obtained in a large longitudinal study on 572 patients with

12

atherosclerosis; in this setting, the authors showed that the extent of the atherosclerotic disease was

13

associated with elevated IgA and IgG titers to infectious agents and CVD mortality was increased

14

by the number of infectious pathogens (Espinola-Klein et al., 2002). Another large human study,

15

involving 657 subjects with no history of CVD demonstrated that the mean carotid intima-media

16

thickness was related to the magnitude of the periodontal bacterial infection (Desvarieux et al.,

17

2005).

18

Indeed, raised serum titers of antibody to Chlamydia were observed in patients with acute

19

myocardial infarction or chronic coronary heart disease (Saikku, 1988). In addition, elevated

20

concentrations of lipoplysaccharides (also known as endotoxins, the large molecules of the outer

21

wall of Gram-negative bacteria) have been considered a risk factor for experimental and, possibly,

22

human atherosclerosis (Prasad et al., 2002). Among other findings suggesting an association

23

between infectious agents and atherosclerosis, bacterial DNA has been detected in atheromatous

24

plaques by quantitative PCR (Ott et al., 2006; Kozarov et al., 2006). Very recently, 16S RNA

25

sequencing detected 17 identical phylotypes in atherosclerotic plaques and subgingival samples

26

from CVD patients suggesting bacterial transfer between periodontal pockets and coronary arteries

5

1

(Serra E Silva Filho et al., 2014). In addition, some bacterial strains were also isolated and

2

cultivated from atheromas (Fiehn et al., 2005; Kozarov et al., 2005).

3

It is also worth mentioning that an antimicrobial activity has been attributed to statins, the most

4

powerful drugs for lowering plasma cholesterol and cardiovascular mortality (Falagas et al., 2008;

5

Sun & Singh, 2009). These data indirectly lend support to the infective hypothesis of CVD,

6

implying that reduction of cardiovascular mortality following statin treatment could be due, at least

7

in part, to the statin antimicrobial effect (Kozarov et al., 2014).

8

As far as cellular and molecular mechanisms that could mediate the pro-atherogenic activity of

9

infectious agents, the development of the atherosclerotic plaque could be favored by the infection-

10

associated chronic inflammation of the vessel wall and endothelial dysfunction (Ridker et al., 2002;

11

Epstein et al., 2009). Indeed, bacteria seem to activate the most relevant cell types in the

12

atherosclerotic plaque development: leukocytes, endothelial cells (ECs), and smooth muscle cells.

13

Indeed, bacteria stimulate the toll-like receptor-mediated cytokine release in leukocytes, the

14

expression of adhesion molecules in ECs, and a prothrombotic response in smooth muscle cells

15

(Stassen et al., 2008; Kozarov, 2012; Roth et al., 2009). This evidence was further supported by

16

transcriptomic analysis of human aortic endothelial cells (HAECs) after infection with

17

Porphyromonas gingivalis (Chou et al., 2005). In addition, P. gengivalis has also been shown to

18

induce apoptosis in HAECs and the secreted proteins of the same bacterium to disrupt and alter

19

trans-endothelial resistance in polarized ECs (Roth et al., 2007; Kozarov et al., 2003).

20 21

The role of endothelial progenitor cells in vascular integrity

22

A plethora of endothelium-related cells have been attributed the capacity to replace damaged

23

endothelium (Table 1). Firstly, intact mature ECs, adjacent to injured intima, were considered one

24

of the major components of the repair process (Carmeliet, 2000). Also circulating ECs (mature ECs

25

that may have detached from the vascular intimal layer following some injury and increases in

26

number in the presence of vessel damage) have been ascribed some kind of reendothelization

27

capability (Duda et al., 2007; Moroni et al., 2005; Yoder, 2010). However, besides these mature 6

1

ECs, with very limited proliferative capacity, a major step forward was taken when it was observed

2

that a subtype of peripheral blood mononuclear cell, expressing surface markers of both

3

hematopoietic stem cells and primary embryonic hemangioblast (CD34, and VEGFR2), was able to

4

differentiate into ECs and home at ischemic angiogenic sites (Asahara et al., 1997). Later on,

5

CD34+ cells with similar capabilities were isolated from the bone marrow and were termed EPCs

6

(Shi et al., 1998), but the group of Rafii suggested that true EPCs should coexpress CD133, a

7

recognized marker of pluripotent cells (Mahmood et al., 2011; Peichev et al., 2000). Interestingly,

8

further studies associated the presence/absence of surface markers with specific functionalities of

9

EPCs, like vascular homing and repair competence, incorporation into the damaged endothelium,

10

and in vivo vessel formation (Asahara et al., 1997; Becher et al., 2010, Friedrich et al., 2006; Hur et

11

al., 2004; Sieveking et al., 2008).

12

EPCs are being increasingly recognized as key players in the maintenance of endothelium integrity,

13

injury repair, and postnatal neovascularization (Asahara et al., 2011, Hirschi et al., 2008; Napoli et

14

al., 2008; Sen et al., 2011; Yoder et al., 2007). An indirect contribution of EPCs to

15

neovascularization has been proposed, namely a paracrine effect of EPCs, due to the secretion of

16

pro-angiogenic cytokines and growth factors that induce proliferation and migration of pre-existing

17

ECs (Asahara et al., 2011). Indeed, it has been demonstrated that EPCs migrate to sites of vascular

18

injury and secrete vascular endothelial growth factor (VEGF), hepatic growth factor, angiopoietin-1

19

(Ang-1), and many more vasculogenic factors (Miyamoto et al., 2007). These observations

20

emphasized that these cells could allow therapeutic vasculogenesis in ischemic tissues. Moreover,

21

ex vivo isolated EPCs improved blood flow and tissue function in animal models of myocardial

22

infarction (MI) and hind limb ischemia (de Nigris et al., 2005; Kawamoto et al., 2003; Urbich et al.,

23

2003). More importantly, several clinical studies indicate the possibility to use EPCs as therapeutic

24

strategies for myocardial neovascularization and endothelial regeneration (Hristov & Weber, 2006;

25

Shantsila et al., 2008; Vanderheyden et al., 2007). Recent studies suggest also the possibility of

26

using new stents to attract EPCs. The EPC capturing technology has been shown to promote the

7

1

arterial healing response by binding circulating EPCs, resulting in the rapid establishment of a

2

functional endothelial monolayer (Aoki et al., 2005; Barsotti et al., 2009).

3

Several other studies showed that bone marrow-derived progenitor cells significantly improved

4

blood supply in patients with peripheral artery disease and MI (Assmus et al., 2002; Casamassimi et

5

al., 2012; Tateishi-Yuyama et al., 2002). However, further human studies, aimed at evaluating the

6

therapeutic efficacy of EPC infusion, gave contradictory results with significant improvement of

7

limb ischemia, but only very modest benefits of left ventricular function after MI (Lasala &

8

Minguel, 2009; Martin-Rendon et al., 2008; Schächinger et al., 2004). In this regard, differences in

9

the route of administration of the progenitor cells could contribute to conflicting evidence. Indeed,

10

either intramuscular or intra-arterial administration of progenitor cells have been used to treat

11

peripheral artery disease, while intra-coronary injection has been used to treat MI.

12 13

Bartonella henselae: mechanisms of infection and cardiovascular localization

14

B. henselae, a facultative intracellular gram-negative bacterium, causes cat-scratch disease, a benign

15

infection often characterized by lymphadenopathy or by an asymptomatic course in

16

immunocompetent patients. However, B. henselae infections occur more frequently and cause

17

poorer-prognosis diseases in immunocompromised patients (Fig.1) (Picascia et al., 2015; Maguiña

18

et al., 2009). In these patients, B. henselae can develop bacillary angiomatosis (BA) or peliosis

19

(BP), vasoproliferative tumor lesions of the skin or the inner organs, respectively (Mosepele et al.,

20

2012; Relman et al., 1990), which derive from bacterial colonization and activation of human ECs

21

inducing bacteremia and fever of unknown cause (Chomel et al., 2009). In addition, B. henselae, as

22

also B. quintana, causes blood-culture-negative endocarditis (BCNE), as indicated by its isolation

23

from native aortic valve tissue of people affected by infectious endocarditis (IE) (Lamas et al.,

24

2013; González et al., 2014; Que et al., 2011; Katsouli et al., 2013; Fournier et al., 2001).

25

Furthermore, because of the negativity of the blood culture, the diagnosis of BCNE is usually

26

considerably delayed; this could be the reason why most patients present acute cardiac failure,

27

cardiac murmur, dyspnea, and bibasilar rales, thus suggesting global cardiac failure (Brouqui & 8

1

Raoult, 2001; Dimopoulos et al., 2012). Therefore, the possibility of Bartonella spp. infection must

2

be considered in every patient with IE.

3

In the mammalian reservoir, B. henselae initially infects a yet unrecognized primary niche, which

4

disseminates microorganisms into the bloodstream leading to the establishment of a long-lasting

5

intraerythrocytic bacteremia (Chomel et al., 2009). Recently, a murine model of chronic infection in

6

immunocompromised SCID/Beige mice showed the ability of bacteria to recapitulate human

7

pathologies. Indeed, in this model, bacteria grow in extracellular aggregates, embedded within

8

collagen matrix, similar to the observations in BP, BA and catch-scratch disease (Chiaraviglio et al.,

9

2010). Bacterial VirB type IV secretion systems (T4SS) represent crucial pathogenic factors that

10

have contributed to the B. henselae expansion in nature; in fact, T4SS facilitate adaptation to

11

mammalian hosts (Schmid et al., 2004; Schülein et al., 2001) by translocation of a cocktail of

12

Bartonella effector proteins (Beps) into host cells (Okujava et al., 2014). The ability to survive in

13

these cells helps B. henselae to escape the host immune response, with its low endotoxic-potency

14

lipopolysaccaride (Zähringer et al., 2004). Until now, cellular immune response was considered the

15

only hallmark of immunity to intracellular pathogens. Even so, the mechanisms underlying B.

16

henselae evasion of the human immune system response are still not fully understood (Mosepele et

17

al., 2012). Invasive bacterial pathogens, during the infection, exploit various cellular processes to

18

facilitate their uptake into an intracellular compartment of the host cells. The endocardium is one of

19

the most severe localization of Bartonella infection and is an optimal point to transfer the microbe

20

to blood cells.

21

The interactions between human pathogenic bacteria and human vascular progenitor cells is a

22

matter of increasing attention and many research lines aim at clarifying how infective events can

23

contribute to worsen vascular pathologies.

24 25

Endothelial cells and Bartonella henselae

26

It is known that erythrocytes and ECs are target cells of Bartonella infection (Dehio, 2001) and the

27

location within these cells is also believed to protect Bartonella spp. from antimicrobial agents 9

1

(Rolain et al., 2001; Rolain et al., 2003). Intraerythrocytic and endothelial/periendothelial

2

persistence

3

immunocompromised hosts, respectively (Mosepele et al., 2012).

4

Mechanisms required to infect human erythrocytes and ECs are: (i) massive rearrangements of actin

5

cytoskeleton with formation of bacterial aggregates (invasome) (Dehio, 2001); (ii) nuclear factor

6

kB-dependent proinflammatory activation, leading to adhesion molecules expression and

7

chemokine secretion (Kempf et al., 2001; Resto-Ruiz, 2002); (iii) inhibition of apoptosis (Kirby &

8

Nekorchuk, 2002); (iv) two T4SSs (Trw and VirB) to adapt to a wide range of mammalian hosts,

9

both essential for the interaction with the host but at different stages of the infection cycle (Okujava

of

Bartonella

are

distinguishing

features

in

immunocompetent

and

10

et al., 2014).

11

Inihibition of apoptosis, together with induction of cell proliferation, are pivotal mechanisms

12

allowing Bartonella permanence into the host cell. Indeed, Bartonella induces directly EC

13

proliferation and inhibits human umbilical vein endothelial cell apoptosis, thus suggesting a direct

14

and indirect role of the pathogen in vasoproliferation (Kirby & Nekorchuk, 2002; Pullianinen &

15

Dehio, 2009; Schmid et al., 2004). B. henselae-triggered vasculoproliferative lesions resemble

16

tumor angiogenesis, the pathological process of new capillary formation out of pre-existing blood

17

vessels (Varanat et al., 2013). Components of the VEGF family are essential regulators of neo-

18

angiogenesis under physiological as well as pathological conditions (Takahashi & Shibuya, 2005).

19

VEGF is secreted from B. henselae-infected macrophages (Resto-Ruiz et al., 2002), recruited to the

20

site of infection as a result of proinflammatory response in ECs (Fuhrmann et al., 2001; Schmid et

21

al., 2004), or from infected epithelial cells surrounding the blood vessels (Kempf et al., 2008). This

22

mechanism has been proposed to contribute to BA (cutaneous manifestations) or BP (hepatic

23

manifestations) by promoting EC proliferation in a paracrine manner (Deng et al., 2012; Harms &

24

Dehio,

25

immunocompromised individuals (Resto-Ruiz et al., 2002; Schmid et al., 2006). Nevertheless,

26

hepatosplenic complications among immunocompetent adults with cat-scratch disease have been

27

recently reviewed and three new cases have been communicated in Spain (García et al., 2014).

2012).

These

lesions

have

been

reported

as

a

common

characteristic

in

10

1

When Bartonella gains access to the human circulatory system, it disseminates from the point of

2

inoculation and colonizes secondary foci showing preference for highly vascularized tissues like

3

heart valves, liver and spleen (Chiaraviglio et al., 2010). Immediately after, bacteria are released

4

into the blood stream, where they efficiently infect erythrocytes and also reinfect the primary niche

5

(Fig. 1).

6

The evidence that B. henselae can infect and damage EPCs reducing the endothelium regenerating

7

potential (Salvatore et al., 2008) might suggest that this infection could increase the risk for CVD. It

8

has been reported that immunocompromised patients, such as those affected by AIDS, receiving

9

organ transplants, under immunosuppressive therapy, or affected by Down syndrome (DS), have

10

lower levels of EPCs, reduced angiogenesis, and higher risk of CVD (Brouqui & Raoult, 2001;

11

Costa et al., 2010; Dehio, 2001; Harms & Dehio, 2012; Kolb-Mäurer et al., 2002; Mändle, 2005;

12

Mosepele et al., 2012; Otsui et al., 2007; Que & Moreillon, 2011; Ryeom & Folkman, 2009; Teofili

13

et al., 2010; Vis et al., 2009). Interestingly, Costa et al. observed that B. henselae reduces EPC

14

number and angiogenesis in DS patients, possibly due to increased oxidative stress (Costa et al.,

15

2010). Furthermore, recent data establish that Bacteroides fragilis colonization protects mice from

16

B. henselae infection and exerts beneficial effects on EPCs and vascular system through

17

polysaccharide A (Sommese et al., 2012). The hypothesis that infectious agents could impair the

18

endothelial repair potential was reinforced by another study demonstrating an association between

19

cerebral malaria and low levels of circulating ECs in African children (Gyan et al., 2009). An in

20

vitro study has underlined that hematopoietic stem cells (HSCs) are resistant to infection with

21

Listeria monocytogenes, Salmonella enteritica, and Yersinia enterocolitica but not when these cells

22

are differentiated (Kolb-Mäurer et al., 2002). Differently, it has been demonstrated that HSCs play a

23

central role in the pathogenesis of B. henselae infection (Mändle et al., 2005).

24

Although several studies have highlighted the main pathogenetic mechanisms of B. henselae

25

infection (Schulein et al., 2001; Dehio, 2008; Okujava et al., 2014), to date it is still unclear what is

26

the primary niche of the bacterium in vivo. Three loci have been proposed as the primary sites of

27

infection: extracellular matrix, bone-marrow stem cells, and mature ECs (Mosepele et al., 2012). 11

1

Additionally, based on the involvement of lymph nodes in the progression of bartonellosis, it has

2

been proposed a role for the migratory cells of the lymphatic system (lymphocytes or mononuclear

3

phagocytes) in the establishment of the primary niche of Bartonella infection (Harms & Dehio,

4

2012). As previously mentioned, Salvatore et al. demonstrated that isolated EPCs internalize B.

5

henselae constituting probably an additional circulating niche of this pathogen (Salvatore et al.,

6

2008). Mobilized EPCs could carry the pathogen to other organs and, more importantly, to the

7

endothelium of the microcirculation.

8

On this basis, it is plausible that the colonization of ECs and EPCs by Bartonella could profoundly

9

alter the maintenance of vascular homeostasis either by impairing the potential of these cells to

10

regenerate the endothelial layer (Fig. 2), or by promoting the development of vasoproliferative

11

tumors. In fact, it has been reported that B. henselae could reduce the function of EPCs (Salvatore et

12

al., 2008), induce the hyper-proliferation of ECs in vitro, and be associated with tumors of the

13

vascular system in patients (Kaiser et al., 2011; Koehler et al., 1997; Leong et al., 1992; Pulliainen

14

& Dahio, 2009).

15 16

Conclusion

17

CVD remains a major burden on public health all over the world, and therefore the scientific and

18

societal interest in the field is still very strong.

19

Infectious agents can be detrimental to the cardiovascular apparatus through different mechanisms

20

such as direct microbial invasion of the vascular cells. This ability allows the microbes to escape the

21

cellular/humoral immune response, to survive in an intracellular environment, and to avoid the lysis

22

that can induce the chronicity of the vascular inflammations. In addition, bacteria can cause

23

endothelial activation/dysfuction and the secretion of inflammatory cytokines with stimulation and

24

migration of leukocytes, thus promoting atherogenesis. However, infectious diseases can be

25

detrimental to cardiovascular health not only because they directly promote atherogenesis, but also

26

because they could indirectly contribute to atherosclerosis by reducing the endothelial repair

27

potential of adult stem cells. To address this point we use as a paradigm the recent finding that B. 12

1

henselae is able to infect EPCs and reduce their number and functionality (Salvatore et al., 2008).

2

Since circulating EPCs play an important role in accelerating endothelization at sites of vascular

3

damage, it is conceivable to hypothesize that the atherosclerotic plaque growth is enhanced and

4

plaque stability is reduced by a spoiled asset of endothelial precursors.

5

These evidences herein summarized lend support to the possibility that B. henselae infection could

6

represent a novel risk and/or prognostic factor for CVD. Further studies are needed in order to

7

support the involvement of Bartonella-endothelial interaction in the onset and evolution of CVD.

8 9

Acknowledgments

10

This work has been supported by POR Campania FSE 2007-2013 Project MODO - Model

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Organism, CUP B25B09000030007 and by POR Campania FSE 2007-2013 Project CRÈME.

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Conflict of interest

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The authors declare that they have no conflict of interest.

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1 2 3

Table 1. Molecular characterization of angiogenic cells

4 Cell Name

Molecular Phenotype

CFU-EC

CD31,CD105, KDR, vWF, Ac-LDL, CD14, CD45, CD115

Yoder et al., 2007

ECFC

CD31, CD105, CD144, CD146, KDR, vWF, UEA-I, Ac-LDL

Yoder et al., 2007

VEC

CD34, Flt1, KDR

George et al., 2011

Myelo/Monocytic Progenitors

Flt1, KDR

George et al., 2011

MEC

VE-Cadherin

Reference

George et al., 2011

MC

vWF

George et al., 2011

MAPC

Flt1, KDR

George et al., 2011

LEC

KDR

George et al., 2011

Hemangioblast

KDR, CD34

George et al., 2011

HC

CD133

George et al., 2011

EPC

CD133, KDR, CD34

George et al., 2011

EC

vWF

George et al., 2011

CHS

CD45

George et al., 2011

Early EPC

CD34, CD133, VEGFR2, CD14, CD45, vWF, CD31

Sen et al., 2011

Late Outgrowth EPC

vWF, VEGFR2, CD31, CD45, CD144, CD34

Sen et al. 2011

Mature EC

vWF, VEGFR2, CD31, CD144, CD34

Sen et al., 2011

CEC

CD31, CD34, CD105, CD144, CD146, CD202b, VEGFR2

Yoder, 2011

CAC

CD133, VEGFR2, CD115, CD31, UEA-1, e-NOS, vWF, acLDL, ALDH

Hirschi et al., 2008

ECFC

CD34, VEGFR2, CD31, UEA-1, e-NOS, vWF, ac-LDL

Hirschi et al., 2008

CD133, VEGFR-2, CD115, CD31, ALDH, ac-LDL, UEA-1, eHirschi et al., 2008 NOS, vWF CAC, circulating angiogenic cell; CEC, circulating endothelial cell; CFU-EC, colony forming unit endothelial cell; CFU-Hill, colony forming unit-Hill; CHS,cells of hematopoietic system; EC, endothelial cell; ECFC, endothelial colony forming cell; EPC, endothelial progenitor cell; HC, hematopoietic cell; LEC, lymphatic endhotelial cell; MAPC, multipotent adult progenitor cell; MC, megakaryocytes; MEC, mature endhotelial cell; VEC, vascular endothelial cell. CFU-Hill

5 6 7 8 9 23

1

LEGENDS TO FIGURES

2 3

Fig. 1. Natural history of B. henselae. It infects a variety of mammals (e.g., cats, dogs and humans)

4

and is transmitted by hematophagous insects (Ctenocephalides felis). B. henselae attaches human

5

erythrocytes causing a persistent infection that culminates in the potentially fatal cat-scratch

6

disease. The EPCs can internalize B. henselae, thus possibly establishing a circulating niche of this

7

pathogen.

8 9

Fig. 2. A proposed mechanism to explain the reduced vascular repair potential in people infected by

10

B. henselae. A low level of cardiovascular risk factors can induce minimal endothelial injury; the

11

intracellular bacterium reduces the number and function of EPCs that can not efficiently fix the

12

damaged endothelium, thus favouring the development of the atherosclerotic plaque.

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Figure 1 Click here to download Figure: Fig1.jpg

Figure 2 Click here to download Figure: Fig2.jpg