Disorders of Phagocyte Function

Neutrophils are the first line of defense against microbial invasion. They arrive at the site of inflammation during the critical 2-4 hr after microbial invasion to contain the infection and prevent hematogenous dissemination. This well-orchestrated process is one of the most interesting stories in modern cell biology. In fact, much of our knowledge about neutrophil function derives from studies done in patients with genetic errors in neutrophil function. These critical functions and their associated disorders are depicted in Fig. 153.2 . Children with phagocytic dysfunction present at a young age with recurrent infections that often involve unusual organisms and are poorly responsive to treatment.

Primary defects of phagocytic function comprise <20% of immunodeficiencies, and there is significant overlap in the presenting signs and symptoms between phagocytic disorders and lymphocyte and humeral disorders. Children with phagocytic defects present with deep tissue infection, pneumonia, adenitis, or osteomyelitis rather than bloodstream infections ( Tables 156.1 and 156.2 and Fig. 156.1 ). A few clinical features point to phagocyte defects rather than other immunodeficiencies, but correct diagnosis relies on highly specialized laboratory tests.

Table 156.1

Infections and White Blood Cell Defects: Features That Can Be Seen in Phagocyte Disorders

From Leung DYM: Pediatric allergy principles and practice , ed 2, Philadelphia, 2010, Saunders, p 134.

SEVERE INFECTIONS		RECURRENT INFECTIONS		SPECIFIC INFECTIONS		UNUSUALLY LOCATED INFECTIONS
Type of Infection	Diagnosis to Consider	Site of Infection	Diagnosis to Consider	Microorganism	Diagnosis to Consider	Site of Infection	Diagnosis to Consider
Cellulitis	Neutropenia, LAD, CGD, HIES	Cutaneous	Neutropenia, CGD, LAD, HIES	Staphylococcus epidermidis	Neutropenia, LAD	Umbilical cord	LAD
Colitis	Neutropenia, CGD	Gums	LAD, neutrophil motility disorders	Serratia marcescens, Nocardia, Burkholderia cepacia	CGD	Liver abscess	CGD
Osteomyelitis	CGD, MSMD pathway defects	Upper and lower respiratory tract	Neutropenia, HIES, functional neutrophil disorders	Aspergillus	Neutropenia, CGD, HIES	Gums	LAD, neutrophil motility disorders
		Gastrointestinal tract	CGD, MSMD pathway defects (salmonella)	Nontuberculous mycobacteria, BCG	MSMD pathway defects, SCID, CGD
		Lymph nodes	CGD, MSMD pathway defects (mycobacteria)	Candida	Neutropenia, CGD, MPO
		Osteomyelitis	CGD, MSMD

BCG, Bacille Calmette-Guérin; CGD, chronic granulomatous disease; HIES, hyper-IgE syndrome; LAD, leukocyte adhesion deficiency; MSMD, mendelian susceptibility to mycobacterial disease; SCID, severe combined immunodeficiency.

Table 156.2

Clinical Disorders of Neutrophil Function

Adapted from Curnutte JT, Boxer LA: Clinically significant phagocytic cell defects. In Remington JS, Swartz MN, editors: Current clinical topics in infectious disease , ed 6, New York, 1985, McGraw-Hill, p 144.

DISORDER	ETIOLOGY	IMPAIRED FUNCTION	CLINICAL CONSEQUENCE
DEGRANULATION ABNORMALITIES
Chédiak-Higashi syndrome (CHS)	Autosomal recessive; disordered coalescence of lysosomal granules; responsible gene is CHSI/LYST , which encodes a protein hypothesized to regulate granule fusion	Decreased neutrophil chemotaxis, degranulation, and bactericidal activity; platelet storage pool defect; impaired NK function, failure to disperse melanosomes	Neutropenia; recurrent pyogenic infections; propensity to develop marked hepatosplenomegaly as a manifestation of hemophagocytic syndrome
Specific granule deficiency	Autosomal recessive; functional loss of myeloid transcription factor arising from a mutation or arising from reduced expression of Gfi-1 or C/EBPε , which regulates specific granule formation	Impaired chemotaxis and bactericidal activity; bilobed nuclei in neutrophils; defensins, gelatinase, collagenase, vitamin B ₁₂ –binding protein, and lactoferrin	Recurrent deep-seated abscesses
ADHESION ABNORMALITIES
Leukocyte adhesion deficiency 1 (LAD-1)	Autosomal recessive; absence of CD11/CD18 surface adhesive glycoproteins (β ₂ -integrins) on leukocyte membranes most commonly arising from failure to express CD18 messenger RNA	Decreased binding of iC3b to neutrophils and impaired adhesion to ICAM-1 and ICAM-2	Neutrophilia; recurrent bacterial infection associated with a lack of pus formation
Leukocyte adhesion deficiency 2 (LAD-2)	Autosomal recessive; loss of fucosylation of ligands for selectins and other glycol conjugates arising from mutations of GDP-fucose transporter	Decreased adhesion to activated endothelium expressing ELAM	Neutrophilia; recurrent bacterial infection without pus
Leukocyte adhesion deficiency 3 (LAD-1 variant syndrome)	Autosomal recessive; impaired integrin function arising from mutations of FERMT3 , which encodes kindlin-3 in hematopoietic cells; kindlin-3 binds to β-integrin and thereby transmits integrin activation	Impaired neutrophil adhesion and platelet activation	Neutrophilia, recurrent infections, bleeding tendency
DISORDERS OF CELL MOTILITY
Enhanced motile responses; FMF	Autosomal recessive gene responsible for FMF on chromosome 16, which encodes for a protein called pyrin; pyrin regulates caspase-1 and thereby IL-1β secretion; mutated pyrin may lead to heightened sensitivity to endotoxin, excessive IL-1β production, and impaired monocyte apoptosis	Excessive accumulation of neutrophils at inflamed sites, possibly the result of excessive IL-1β production	Recurrent fever, peritonitis, pleuritis, arthritis, amyloidosis
DEPRESSED MOTILE RESPONSES
Defects in the generation of chemotactic signals	IgG deficiencies; C3 and properdin deficiency can arise from genetic or acquired abnormalities; mannose-binding protein deficiency predominantly in neonates	Deficiency of serum chemotaxis and opsonic activities	Recurrent pyogenic infections
Intrinsic defects of the neutrophil, e.g., LAD, CHS, specific granule deficiency, neutrophil actin dysfunction, neonatal neutrophils	In the neonatal neutrophil there is diminished ability to express β ₂ -integrins, and there is a qualitative impairment in β ₂ -integrin function	Diminished chemotaxis	Propensity to develop pyogenic infections
Direct inhibition of neutrophil mobility, e.g., drugs	Ethanol, glucocorticoids, cyclic AMP	Impaired locomotion and ingestion; impaired adherence	Possible cause for frequent infections; neutrophilia seen with epinephrine arises from cyclic AMP release from endothelium
Immune complexes	Bind to Fc receptors on neutrophils in patients with rheumatoid arthritis, systemic lupus erythematosus, and other inflammatory states	Impaired chemotaxis	Recurrent pyogenic infections
Hyper-IgE syndrome	Autosomal dominant; responsible gene is STAT3	Impaired chemotaxis at times; impaired regulation of cytokine production	Recurrent skin and sinopulmonary infections, eczema, mucocutaneous candidiasis, eosinophilia, retained primary teeth, minimal trauma fractures, scoliosis, and characteristic facies
Hyper-IgE syndrome–AR	Autosomal recessive; more than 1 gene likely contributes to its etiology	High IgE levels, impaired lymphocyte activation to staphylococcal antigens	Recurrent pneumonia without pneumatoceles sepsis, enzyme, boils, mucocutaneous candidiasis, neurologic symptoms, eosinophilia
MICROBICIDAL ACTIVITY
Chronic granulomatous disease (CGD)	X-linked and autosomal recessive; failure to express functional gp91 ^phox in the phagocyte membrane in p22 ^phox (AR) Other AR forms of CGD arise from failure to express protein p47 ^phox or p67 ^phox	Failure to activate neutrophil respiratory burst, leading to failure to kill catalase-positive microbes	Recurrent pyogenic infections with catalase-positive microorganisms
G6PD deficiency	<5% of normal activity of G6PD	Failure to activate NADPH-dependent oxidase; hemolytic anemia	Infections with catalase-positive microorganisms
Myeloperoxidase deficiency	Autosomal recessive; failure to process modified precursor protein arising from missense mutation	H ₂ O ₂ -dependent antimicrobial activity not potentiated by myeloperoxidase	None
Rac2 deficiency	Autosomal dominant; dominant negative inhibition by mutant protein of Rac2-mediated functions	Failure of membrane receptor–mediated O ₂ ⁻ generation and chemotaxis	Neutrophilia, recurrent bacterial infections
Deficiencies of glutathione reductase and glutathione synthetase	AR; failure to detoxify H ₂ O ₂	Excessive formation of H ₂ O ₂	Minimal problems with recurrent pyogenic infections

AMP, Adenosine monophosphate; AR, autosomal recessive; C, complement; CD, cluster of differentiation; ELAM, endothelial-leukocyte adhesion molecule; FMF, familial Mediterranean fever; G6PD, glucose-6-phosphate dehydrogenase; GDP, guanosine diphosphate; ICAM, intracellular adhesion molecule; IL-1, interleukin-1; NADPH, nicotinamide adenine dinucleotide phosphate; NK, natural killer.

Chemotaxis , the direct migration of cells into sites of infection, involves a complex series of events (see Chapter 153 ). Disorders of adhesion or granule abnormalities can have intermediate or profound motility defects, and the propensity to infections is related to a combination of these functional deficits. One family with recessively inherited neutrophil actin dysfunction demonstrated that a pure severe chemotactic defect can result in fatal recurrent infection. Defective in vitro chemotaxis of neutrophils can be detected in children with various clinical conditions. However, unless chemotaxis is essentially absent, it is difficult to establish whether frequent infections arise from a primary chemotactic abnormality or occur as secondary medical complications of the underlying disorder. Dental infection with Capnocytophaga is associated with a clear neutrophil motility defect that resolves when the infection is eliminated.

Motility defects present with significant skin and mucosal infections. Tender cutaneous nodular lesions may also be present and characteristically do not contain neutrophils. In fact, presence of a true abscess makes the diagnosis of a significant chemotactic defect less likely.

Laboratory tests of chemotaxis are biologic assays and have high variability except in the most experienced hands. The assays must be done on freshly obtained blood and are affected by many factors related to blood sampling itself. It is best to assay other features of the suspected disorder, such as surface marker expression, to establish a specific diagnosis.

Leukocyte Adhesion Deficiency

Leukocyte adhesion deficiency types 1 (LAD-1), 2 (LAD-2), and 3 (LAD-3) are rare autosomal recessive disorders of leukocyte function. LAD-1 affects about 1 per 10 million individuals and is characterized by recurrent bacterial and fungal infections and depressed inflammatory responses despite striking blood neutrophilia ( Table 156.3 ). The neutrophils have significant defects in adhesion, motility, and ability to phagocytose bacteria.

Table 156.3

Leukocyte Adhesion Deficiency Syndromes

From Leung DYM: Pediatric allergy principles and practice , ed 2, Philadelphia, 2010, Saunders, p 139.

LEUKOCYTE ADHESION DEFICIENCY (LAD)	TYPE 1 (LAD-1)	TYPE 2 (LAD-2 or CDG-IIc)	TYPE 3 (LAD-3)	E-SELECTIN DEFICIENCY	Rac2 DEFICIENCY
OMIM	116920	266265	612840	131210	602049
Inheritance pattern	Autosomal recessive	Autosomal recessive	Autosomal recessive	Unknown	Autosomal dominant
Affected protein(s)	β ₂ -Integrin common chain (CD18)	Fucosylated proteins (e.g., sialyl-Lewis ^x , CD15s)	Kindlin 3	Endothelial E-selectin expression	Rac2
Neutrophil function affected	Chemotaxis, tight adherence	Rolling, tethering	Chemotaxis, adhesion, superoxide production	Rolling, tethering	Chemotaxis, superoxide production
Delayed umbilical cord separation	Yes (severe phenotype only)	Yes	Yes	Yes	Yes
Leukocytosis/neutrophilia	Yes	Yes	Yes	No (mild neutropenia)	Yes

CDG-IIc, Congenital disorder of glycosylation IIc, OMIM , Online Mendelian Inheritance in Man.

Genetics and Pathogenesis

LAD-1 results from mutations of the gene on chromosome 21q22.3 encoding CD18, the 95-kDa β ₂ -leukocyte transmembrane integrin subunit. Normal neutrophils express 4 heterodimeric adhesion molecules: LFA-1 (CD11a/CD18), Mac-1 (CD11b/CD18, also known as CR3 or iC3b receptor), p150,95 (CD11c/CD18), and α ₁ β ₂ (CD11d/CD18). These 4 transmembrane adhesion molecules are composed of unique extracellular α ₁ encoded on chromosome 16, and they share a common β ₂ subunit (CD18) that links them to the membrane and connects them to intracellular signal transduction machinery. This group of leukocyte integrins is responsible for the tight adhesion of neutrophils to the endothelial cell surface, egress from the circulation, and adhesion to iC3b-coated microorganisms, which promotes phagocytosis and particulate activation of the phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Some mutations of CD11/CD18 allow a low level of assembly and activity of integrin molecules, resulting in retention of some neutrophil integrin adhesion function and a moderate phenotype.

Because of their inability to adhere firmly to intercellular adhesion molecules 1 (ICAM-1) and 2 (ICAM-2) expressed on inflamed endothelial cells (see Chapter 153 ), neutrophils cannot transmigrate through the vessel wall and move to the site infection. Furthermore, neutrophils that do arrive at inflammatory sites fail to recognize microorganisms opsonized with complement fragment iC3b , an important stable opsonin formed by the cleavage of C3b. Therefore, other neutrophil functions such as degranulation and oxidative metabolism normally triggered by iC3b binding are also greatly compromised in LAD-1 neutrophils, resulting in impaired phagocytic function and high risk for serious and recurrent bacterial infections.

Monocyte function is also impaired, with poor fibrinogen-binding function, an activity that is promoted by the CD11/CD18 complex. Consequently, such cells are unable to participate effectively in wound healing.

Children with LAD-2 share the clinical features of LAD-1 but have normal CD11/CD18 integrins. Features unique to LAD-2 include neurologic defects, cranial facial dysmorphism, and absence of the erythrocyte ABO blood group antigen ( Bombay phenotype). LAD-2 (also known as congenital disorder of glycosylation IIc (CDG-IIc) derives from mutations in the gene encoding a specific guanosine diphosphate (GDP)- l -fucose transporter of the Golgi apparatus. This abnormality prevents the incorporation of fucose into various cell surface glycoproteins, including the carbohydrate structure sialyl Lewis X that is critical for low-affinity rolling adhesion of neutrophils to vascular endothelium. This is an important initial step necessary for subsequent integrin-mediated activation, spreading, and transendothelial migration. Infections in LAD-2 are milder than that in LAD-1.

LAD-3 is characterized by a Glanzmann thrombasthenia –like bleeding disorder, delayed separation of the umbilical cord, and serious skin and soft tissue infections similar to those seen in LAD-1, and failure of leukocytes to undergo β ₂ - and β ₁ -integrin–mediated adhesion and migration. Mutations in KINDLIN3 affect integrin activation.

Clinical Manifestations

Patients with the severe clinical form of LAD-1 express <0.3% of the normal amount of the β ₂ -integrin molecules, whereas patients with the moderate phenotype may express 2–7% of the normal amount. Children with severe forms of LAD present in infancy with recurrent, indolent bacterial infections of the skin, mouth, respiratory tract, lower intestinal tract, and genital mucosa. Significant neutrophilic leukocytosis, often >25,0000/mm ³ , is a prominent feature. They may have a history of delayed separation of the umbilical cord, usually with associated infection of the cord stump. The presence of significant omphalitis is an important feature that distinguishes these rare patients from the 10% of healthy infants who can have cord separation at age 3 wk or later. Skin infection may progress to large chronic ulcers with polymicrobial infection, including anaerobic organisms ( Fig. 156.2 ). The ulcers heal slowly, need months of antibiotic treatment, and often require plastic surgery grafting. Severe gingivitis can lead to early loss of primary and secondary teeth ( Fig. 156.3 ). Infected areas characteristically have very little neutrophilic infiltration.

Fig. 156.2, Skin infection of a patient with leukocyte adhesion deficiency type 1.

Fig. 156.3, Oral pathology in a patient with leukocyte adhesion deficiency type 1.

The pathogens infecting patients with LAD-1 are similar to those affecting patients with severe neutropenia (see Chapter 157 ) and include Staphylococcus aureus and enteric gram-negative organisms such as Escherichia coli. These patients are also susceptible to opportunistic infection by fungi such as Candida and Aspergillus. Typical signs of inflammation, such as swelling, erythema, and warmth, may be absent. Pus does not form, and few neutrophils are identified microscopically in biopsy specimens of infected tissues. Despite the paucity of neutrophils within the affected tissue, the circulating neutrophil count during infection typically exceeds 30,000/µL and can surpass 100,000/µL. During intervals between infections, the peripheral blood neutrophil count may chronically exceed 12,000/µL. LAD-1 genotypes with only moderate, rather than absent, amounts of functional integrins at the surface of the neutrophil have significantly reduced severity and frequency of infections compared to children with the severe form, although gingival disease is still a prominent feature.

Laboratory Findings

The diagnosis of LAD-1 is established most readily by flow cytometric measurements of surface CD11b/CD18 in stimulated and unstimulated neutrophils. Neutrophil and monocyte adherence, aggregation, chemotaxis, and iC3b-mediated phagocytosis demonstrate striking abnormalities. However, these assays are not clinically available. Delayed-type hypersensitivity reactions are normal, and most individuals have normal specific antibody synthesis, although some patients have impaired T-lymphocyte–dependent antibody responses. The diagnosis of LAD-2 is established by flow cytometric measurement of sialyl Lewis X (CD15) on neutrophils. It is important to note that the flow cytometric assays are not done the same as the more common lymphocyte subset analysis and require specialized approaches to detect levels of surface expression, especially to detect milder phenotypes.

Treatment

Treatment of LAD-1 depends on the phenotype, as determined by the level of expression of functional CD11/CD18 integrins. Early allogeneic hematopoietic stem cell transplantation (HSCT) is the treatment of choice for severe LAD-1 (and LAD-3). One patient was successfully treated with ustekinumab, an inhibitor of interleukins 12 and 23. Other treatment is largely supportive. Patients can be maintained on prophylactic trimethoprim/sulfamethoxazole (TMP/SMX) and should have close surveillance for early identification of infections and initiation of empirical treatment with broad-spectrum antibiotics. Specific determination of the etiologic agent by culture or biopsy is important because of the prolonged antibiotic treatment required in the absence of neutrophil function.

Some LAD-2 patients have responded to fucose supplementation, which induced a rapid reduction in the circulating leukocyte count and appearance of the sialyl Lewis X molecules, accompanied by marked improvement in leukocyte adhesion.

Prognosis

The severity of infectious complications correlates with the degree of β ₂ -integrin deficiency. Patients with severe deficiency may die in infancy, and those surviving infancy have a susceptibility to severe life-threatening systemic infections. Patients with moderate deficiency have infrequent life-threatening infections and relatively long survival.

Chédiak-Higashi Syndrome

Chédiak-Higashi syndrome (CHS) is a rare autosomal recessive disorder characterized by increased susceptibility to infection caused by defective degranulation of neutrophils, a mild bleeding diathesis, partial oculocutaneous albinism, progressive peripheral neuropathy, and a tendency to develop a life-threatening form of hemophagocytic lymphohistiocytosis (see Chapter 534.2 ). CHS is caused by a fundamental defect in granule morphogenesis that results in abnormally large granules in multiple tissues. Pigmentary dilution involving the hair, skin, and ocular fundi results from pathologic aggregation of melanosomes. Neurologic deficits are associated with a failure of decussation of the optic and auditory nerves. Patients exhibit an increased susceptibility to infection that can be explained only in part by defects in neutrophil function. The patients have progressive neutropenia as well as abnormalities in natural killer (NK) function, again related to granule dysfunction.

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