Normal Ciliary Ultrastructure and Function

The upper and lower respiratory tracts are continuously exposed to inhaled pathogens, and local defenses have evolved to protect the airway. The respiratory epithelium in the nasopharynx, middle ear, paranasal sinuses, and larger airways are lined by a ciliated, pseudostratified columnar epithelium that is essential for mucociliary clearance (Fig. 396-1). Motile cilia are hairlike organelles that move fluids, mucus, and inhaled particulates vectorially from conducting airways, paranasal sinuses, and eustachian tubes.

Figure 396-1 Scanning electron photomicrograph of an airway epithelium grown in primary culture showing ciliated and non-ciliated cells.

A mature ciliated epithelial cell has approximately 200 uniform motile cilia that are functionally and anatomically oriented in the same direction, moving with intracellular and intercellular synchrony. Anchored by a basal body to the apical cytoplasm and extending from the cell surface into the extracellular space, each cilium is a complex, specialized structure, composed of roughly 250 proteins. It contains a cylinder of microtubule doublets, arranged around a central pair of microtubules (Fig. 396-2), the characteristic “9+2” arrangement as viewed by cross-sectional views on electron microscopy. Multiple, different adenosine triphosphatases (ATPases), called dyneins, serve as “motors” of the cilium. Attached to the microtubules as distinct inner and outer dynein arms, dyneins cleave ATP to promote microtubule sliding, which is converted into bending. Nexin links and radial spokes act to restrict the degree of sliding between microtubules, allowing the cilium to bend.

Figure 396-2 Electron photomicrograph of a normal cilium demonstrating the complex structure and arrangement of the ciliary axoneme. Each inner and outer dynein arm is composed of numerous proteins and multiple dyneins.

The ciliary axoneme is highly conserved across species, and the structural elements of simple protozoan flagella and the mammalian cilium are similar. The inner dynein arm influences the bend shape of the cilium, whereas the outer dynein arm controls beat force and frequency. Ciliary beat frequency is constant throughout the airway, 8 to 20 beats/seconds, but can be negatively affected by several factors, such as anesthetics and dehydration. Alternatively, beat frequency may be accelerated by exposure to irritants or bioactive molecules, including β-adrenergic agents, acetylcholine, and serotonin. Cilia beat frequency can also be increased through the activity of nitric oxide synthases that are localized in the apical cytoplasm. The coordinated wavelike pattern of ciliary motion has important functions in fluid and cell movement, and any disturbance in the precise, orchestrated movement of the cilia can lead to disease.

In contrast to motile cilia, sensory or primary cilia lack a central microtubule doublet and outer dynein arms, thus creating a “9+0” arrangement and leaving these structures immotile. Primary cilia have specialized sensory functions and are present in the retina, renal nephron, bile duct, hypothalamus, and inner ear. Defects in primary cilia have been linked to wide-ranging conditions, including retinitis pigmentosa, polycystic kidney disease, polycystic liver disease, nephronophthisis, Bardet-Biedl syndrome, Meckel-Gruber syndrome, Joubert syndrome, Alström syndrome, and Jeune syndrome.

A third distinct classification of cilia also exists, but only during embryonic development. These nodal cilia have a “9+0” microtubule arrangement similar to that of primary cilia, but they exhibit rotational movement, resulting in leftward flow of extracellular fluid that establishes body sidedness. Nodal cilia defects result in left-right body orientation abnormalities, such as situs ambiguus (Chapter 425.11).

Genetics of Primary Ciliary Dyskinesia

Primary ciliary dyskinesia is considered to have autosomal recessive patterns of inheritance, though rare cases of autosomal dominant and X-linked inheritance have been reported. The calculated frequency of PCD ranges from 1/12,000 to 1/20,000 live births, but these measures likely underestimate disease incidence in the general population.

PCD is a genetically heterogeneous disorder involving multiple genes; mutations in any of over 250 proteins that are involved in ciliary assembly or structure could theoretically cause disease. Linkage analyses have shown substantial locus heterogeneity, making correlations between ciliary defects and the underlying mutations difficult.

Investigations into the genetic basis of PCD have focused on dynein proteins; candidates genes involved in PCD include DNAI1 (IC78) and DNAH5 (γ-heavy chain). DNAI1, a large gene located on chromosome 9 (9p13-21) and highly expressed in trachea and testes, encodes for an intermediate chain found in the outer dynein arms. Mutations in DNAI1 have been found in PCD patients with outer dynein arm defects, and have been estimated to occur in 10% of PCD patients. Several different mutations in another gene, DNAH5, an axonemal dynein heavy-chain gene localized to chromosome 5p (5p14-5p15) expressed in lung, kidney, brain, and testis, have also been found in families with PCD. More than half of PCD patients with known outer dynein arm defects have DNAH5 mutations. Additional genes (DNAH11, TXNDC, and DNAI2) are also implicated in PCD. DNAH11 is particularly interesting because mutations in this gene have been shown to cause typical clinical phenotypes without apparent axonemal ultrastructural defects.

Clinical Manifestations of Primary Ciliary Dyskinesia (Table 396-1)

Most patients with PCD present in the newborn period shortly after birth with respiratory distress, which manifests as tachypnea, hypoxemia, or even respiratory failure requiring mechanical ventilation. The association of respiratory distress in term neonates with PCD has been underappreciated. Chronic cough and persistent rhinosinusitis have frequently been present since early infancy. In relation to upper respiratory involvement, the infant may have poor feeding and growth delays, similar to the effects of cystic fibrosis (Chapter 395), which can confound the diagnosis. Indeed, the upper respiratory tract is almost universally involved in PCD. Inadequate innate mucus clearance manifests as chronic sinusitis (Chapter 372) and nasal polyposis. Middle ear disease is common, with varying degrees of chronic otitis media leading to conductive hearing loss and myringotomy tube placement, which is often complicated by recurrent otorrhea.

Impaired mucociliary clearance of the lower respiratory tract leads to chronic cough secondary to recurrent pneumonia or bronchitis. Bacterial cultures of sputum or bronchial aspirates commonly yield nontypable Haemophilus influenzae (Chapter 186), Staphylococcus aureus (Chapter 174.1), Streptococcus pneumoniae (Chapter 175), and Pseudomonas aeruginosa (Chapter 197.1). Persistent airway infection and inflammation lead to bronchiectasis, even in preschool children. Clubbing is a sign of long-standing pulmonary involvement.

Left-right laterality defects are found in PCD; 50% of patients have situs inversus totalis with transposition of the thoracic and abdominal organs. Without functional nodal cilia in the embryonic period, thoracoabdominal orientation is random. These patients have Kartagener triad, defined as situs inversus totalis, chronic sinusitis, and bronchiectasis. Approximately 25% of patients with situs inversus totalis have PCD, but situs inversus totalis alone does not establish the diagnosis of PCD. Other forms of laterality defects, such as heterotaxy, have been reported with PCD and may coexist with congenital cardiac defects, asplenia, or polysplenia.

Most men with PCD have immotile spermatozoa because the ultrastructures of the central machinery of the flagellum and motile cilium are similar. Male infertility is typical, but not always found in this disease. Fertility issues in women have also been reported; they are likely due to ciliary dysfunction in the fallopian tubes.

A few case reports have associated neonatal hydrocephalus with PCD. The ependyma of the brain ventricles are lined by ciliated epithelium and are important for cerebrospinal fluid flow through the ventricles and aqueduct of Sylvius. The finding of enlarged brain ventricles on sonograms, when linked with situs inversus totalis, is proposed as a prenatal diagnostic marker for PCD.

Retinitis pigmentosa has been linked with PCD. Intraflagellar transport proteins are essential for photoreceptor assembly, and when mutated, lead to their apoptosis within the retinal pigment epithelium (Chapter 622). X-linked retinitis pigmentosa has been associated with recurrent respiratory infections in families with RPGR gene mutations.

Diagnosis of Primary Ciliary Dyskinesia

Currently the diagnosis of PCD requires the presence of characteristic clinical phenotype and ultrastructural defects of cilia. It should be suspected in children with chronic or recurring upper and lower respiratory tract symptoms. The clinical presentation of PCD can be variable and subtle in milder forms of this disease, in which there is partial preservation of ciliary function. Even in children who have classic clinical features, such as chronic rhinitis in infancy, persistent otitis media, or even situs inversus totalis, the diagnosis is often delayed. The average age of PCD diagnosis is >4 yr; a high index of suspicion is necessary.

Imaging studies show extensive involvement of the paranasal sinuses. Chest radiographs frequently demonstrate bilateral lung overinflation, peribronchial infiltrates, and lobar atelectasis. CT of the chest often reveals bronchiectasis, even in young children. Situs inversus totalis in a child who has chronic respiratory tract symptoms is virtually diagnostic of PCD, but this configuration occurs in only 50% of patients with PCD. Pulmonary function testing of older children typically shows a progressive intrathoracic airway obstruction.

Transmission electron microscopy is the current gold standard to assess ultrastructural defects within the cilium. Curettage from the nasal epithelium or bronchial brushing can provide an adequate specimen for review. Identification of a discrete, consistent defect in any aspect of the ciliary structure with concurrent phenotypic features is sufficient to make the diagnosis. Shortening or absence of dynein arms is the most common abnormality seen in PCD, accounting for 90% of cases with defined ultrastructural defects (Fig. 396-3). Other axonemal changes consistent with PCD include microtubular transposition, radial spoke and nexin link defects, and ciliary agenesis. Unfortunately, ultrastructural examination of cilia as a diagnostic test for PCD has significant drawbacks. Careful interpretation of the ultrastructural findings is necessary, because nonspecific, secondary changes may be seen in relation to exposure to environmental pollutants or infection. Ciliary defects can be acquired. Acute airway infection or inflammation can result in ultrastructural changes (compound cilia or blebs). Ciliary disorientation has also been proposed as a form of primary ciliary dyskinesia, but this phenomenon may be the result of airway injury. Frequently, the diagnosis of PCD can be delayed or missed because of inadequate tissue collection or sample processing as well as the lack of an experienced pathologist who can distinguish between primary and acquired ciliary defects. Several reviews have advocated culturing of airway epithelial cells and allowing the secondary changes to resolve. Nevertheless, the presence of normal axonemal ultrastructure does not exclude PCD.

Figure 396-3 Electron photomicrograph of cilia collected from a patient with primary ciliary dyskinesia (Kartagener syndrome). Although the ciliary axoneme has the normal “9+2” conformation, both inner and outer dynein arms are absent in all cilia.

Qualitative tests to assess ciliary function have been used to screen for PCD. The saccharin test has been applied to qualitatively assess mucociliary function. This approach has several limitations. It is not standardized, does not distinguish between primary and secondary forms of ciliary dyskinesia, and cannot be used in young children. Ciliary beat frequency measurements that use conventional microscopic techniques, have been employed as a screen. This method alone will miss some cases of PCD, such as ciliary transposition, since these defects can have normal beat frequency. High-resolution, high-speed, digital imaging of ciliary motion in multiple planes has permitted more comprehensive analysis of abnormal ciliary beat patterns. Immunofluorescence imaging has been used to show mislocalization of dynein arm proteins. Such techniques are research tools and not widely available.

Another approach exploits the observation that nasal nitric oxide (NO) concentrations are reduced in subjects with PCD. Because nasal NO measurements are relatively easy to perform and noninvasive, this method is a promising screen for PCD in patients >5 yr of age, provided that cystic fibrosis has been excluded (Chapter 395). Few studies in younger children have been reported, and the accuracy of nasal NO measurements in infants has not been established.

Genetic testing for PCD is available, and commercial laboratories offer testing for specific DNAI1 and DNAH5 mutations.

Treatment

No therapies have been adequately studied to demonstrate their efficacy in PCD. Many of treatments applied to PCD patients are similar to those used in other suppurative lung diseases characterized by impaired airway clearance and bronchiectasis, such as cystic fibrosis. No treatments have been shown to correct ciliary dysfunction in PCD.

Strategies to enhance mucociliary clearance are central to PCD therapy, and routine airway clearance with postural drainage, percussion vests, positive expiratory pressure devices, or other techniques should be instituted on a daily basis. Because ciliary function is impaired, cough becomes a critical mechanism for mucus clearance and should not be suppressed. Exercise can enhance airway clearance in patients with PCD and should be encouraged. Inhaled mucolytic agents are often used in cystic fibrosis care, and few case reports have shown improvement in lung function in patients with PCD after treatment. Larger studies supporting these findings are needed before such agents can be routinely used as maintenance therapy for PCD.

When children with PCD develop increasing respiratory symptoms consistent with infection, antimicrobial therapy should be instituted on the basis of respiratory culture results and bacterial sensitivities. Early eradication strategies to clear bacteria from the PCD lung have not been studied. Maintenance therapy with inhaled or oral antibiotics can be used cautiously in patients with PCD who have bronchiectasis or frequent exacerbations, although current literature lacks evidence supporting long-term antimicrobial therapy. Immunizations against pertussis, influenza, and pneumococci are cornerstones of care. Additional preventive measures include avoidance of cigarette smoke and other airway irritants.

Although β-adrenergic agonists have been shown to increase ciliary beat frequency in epithelial cell models, there is no data showing that they improve function of dyskinetic cilia. Moreover, such agents do not necessarily provide bronchodilation in patients with PCD and obstructive airway disease.

Surgical resection of bronchiectatic lung has been performed on patients with PCD, typically in cases of localized disease with severe hemoptysis or recurrent febrile illnesses. It is unclear whether surgical interventions provide any long-term or survival benefit.

Progression to end-stage lung disease and respiratory failure has been reported in patients with PCD. Adult patients have undergone successful heart-lung, double lung, or living donor lobar lung transplantation. Situs inversus totalis complicates the procedure owing to anatomic considerations. Otherwise, survival is similar to that for other transplant recipients.

The treatment of chronic otitis media and middle ear effusions in patients with PCD is controversial. Myringotomy tubes are frequently used in children with PCD, but are not without complications, because they may lead to chronic mucoid otorrhea, permanent membrane perforation, and tympanosclerosis. Myringotomy tubes have not measurably improved hearing acuity. Although hearing tends to improve with time, it should be routinely screened, and hearing aids used when necessary.

Chronic rhinitis and sinusitis are frequent clinical manifestations of PCD. No treatments have been shown to be effective, although patients are frequently treated with sinus lavage and systemic antibiotics when they are symptomatic. As with any overuse of antimicrobial agents, the development of resistant organisms is a concern. When sinus symptoms are severe or refractory to medical management, endoscopic sinus surgery can be used to promote drainage or local delivery of medications.

Prognosis

Although signs and symptoms related to upper respiratory involvement predominate early in PCD, clinical manifestations of airway disease tend to increase with age and becomes the leading cause of morbidity and mortality in PCD patients. It is thought that progression and extent of lung disease can be slowed with early diagnosis and therapy. Thus, routine surveillance studies recommended that the care of children with PCD include regular spirometry to monitor lung function, chest radiographs, and sputum or oropharyngeal cultures to assess airway flora.

Patients with PCD typically have slower decline in pulmonary function than those with cystic fibrosis; prognosis and long-term survival are better. In fact, most patients with PCD have a normal or near normal life span, although some do experience progressive bronchiectasis and respiratory failure.