Value of transmission electron microscopy for primary ciliary dyskinesia diagnosis in the era of molecular medicine: Genetic defects with normal and non-diagnostic ciliary ultrastructure

Abstract

Primary Ciliary Dyskinesia (PCD) is a genetic disorder causing chronic oto-sino-pulmonary disease. No single diagnostic test will detect all PCD cases. Transmission electron microscopy (TEM) of respiratory cilia was previously considered the gold standard diagnostic test for PCD, but 30% of all PCD cases have either normal ciliary ultrastructure or subtle changes which are non-diagnostic. These cases are identified through alternate diagnostic tests, including nasal nitric oxide measurement, high speed videomicroscopy analysis, immunofluorescent staining of axonemal proteins, and/or mutation analysis of various PCD causing genes. Autosomal recessive mutations in DNAH11 and HYDIN produce normal TEM ciliary ultrastructure, while mutations in genes encoding for radial spoke head proteins result in some cross-sections with non-diagnostic alterations in the central apparatus interspersed with normal ciliary cross-sections. Mutations in nexin link and dynein regulatory complex genes lead to a collection of different ciliary ultrastructures; mutations in CCDC65, CCDC164, and GAS8 produce normal ciliary ultrastructure, while mutations in CCDC39 and CCDC40 cause absent inner dynein arms and microtubule disorganization in some ciliary cross-sections. Mutations in CCNO and MCIDAS cause near complete absence of respiratory cilia due to defects in generation of multiple cellular basal bodies; however, the scant cilia generated may have normal ultrastructure. Lastly, a syndromic form of PCD with retinal degeneration results in normal ciliary ultrastructure through mutations in the RPGR gene. Clinicians must be aware of these genetic causes of PCD resulting in non-diagnostic TEM ciliary ultrastructure and refrain from using TEM of respiratory cilia as a test to rule out PCD.

DRC2/CCDC65 is a central hub for assembly of the nexin–dynein regulatory complex and other regulators of ciliary and flagellar motility

“…, 2013 ; Wirschell et al. , 2013 ; Shapiro and Leigh, 2017 ). We predict that the PCD causing mutations in DRC1 and DRC2 likely disrupts the assembly of multiple axonemal substructures in human cilia similarly to that described here in Chlamydomonas .…”

Sperm defects in primary ciliary dyskinesia and related causes of male infertility

“…Nasal nitric oxide gas is reduced in most patients with PCD [10]. Analysis of the respiratory epithelium by nasal brushing can be used for identification of ciliary structural defects by transmission electron microscopy (TEM) [113,118] and defects of the ciliary beat pattern and frequency can be identified by high-speed video microscopy analysis (HSVMA) [107]. HSVMA has excellent sensitivity and specificity for PCD and particular beat patterns have been linked to specific ultrastructural defects [15].…”

Biallelic Mutations in LRRC56, Encoding a Protein Associated with Intraflagellar Transport, Cause Mucociliary Clearance and Laterality Defects

“…For example, DNAH11 nonsense mutations are associated with subtle dynein arm modifications that can be detected only by advanced TEM imaging and tomography. 11 In this regard, approximately 30% of PCDaffected individuals showing ciliary dysfunction have no or very subtle ciliary ultrastructure abnormalities when investigated with standard transmission electron microscopy 58 Overall, our findings underline the evolutionary complexity of outer dynein arms assembly and trafficking. We have shown that bi-allelic mutations in LRRC56 are responsible for laterality defect in three unrelated families.…”