Alpha-1-antitrypsin (AAT), an acute-phase protein encoded by the SERPINA1 gene, is a member of the serine protease inhibitor (SERPIN) superfamily. Its primary function is to protect tissues from enzymes released during inflammation, such as neutrophil elastase and proteinase 3. In addition to its antiprotease activity, AAT interacts with numerous other substances and has various functions, mainly arising from the conformational flexibility of normal variants of AAT. Therefore, AAT has diverse biological functions and plays a role in various pathophysiological processes. This review discusses major molecular forms of AAT, including complex, cleaved, glycosylated, oxidized, and S-nitrosylated forms, in terms of their origin and function.

Encephalitis is a severe neurological syndrome usually caused by viruses. Despite significant progress in diagnostic techniques, the causative agent remains unidentified in the majority of cases. The aim of the present study was to test an alternative approach for the detection of putative pathogens in encephalitis using next-generation sequencing (NGS). RNA was extracted from cerebrospinal fluid (CSF) from a 60-year-old male patient with encephalitis and subjected to isothermal linear nucleic acid amplification (Ribo-SPIA, NuGen) followed by next-generation sequencing using MiSeq (Illumina) system and metagenomics data analysis. The sequencing run yielded 1,578,856 reads overall and 2579 reads matched human herpesvirus I (HHV-1) genome; the presence of this pathogen in CSF was confirmed by specific PCR. In subsequent experiments we found that the DNAse I treatment, while lowering the background of host-derived sequences, lowered the number of detectable HHV-1 sequences by a factor of 4. Furthermore, we found that the routine extraction of total RNA by the Chomczynski method could be used for identification of both DNA and RNA pathogens in typical clinical settings, as it results in retention of a significant amount of DNA. In summary, it seems that NGS preceded by nucleic acid amplification could supplement currently used diagnostic methods in encephalitis.

Hearing loss and ovarian dysfunction are key features of Perrault syndrome (PRLTS) but the clinical and pathophysiological features of hearing impairment in PRLTS individuals have not been addressed. Mutations in one of five different genes HSD17B4, HARS2, LARS2, CLPP or TWNK (previous symbol C10orf2) cause the autosomal recessive disorder but they are found only in about half of the patients. We report on two siblings with a clinical picture resembling a severe, neurological type of PRLTS. For an exhaustive characterisation of the phenotype neuroimaging with volumetric measurements and objective measures of cochlear hair cell and auditory nerve function (otoacustic emissions and auditory brainstem responses) were used. Whole exome sequencing was applied to identify the genetic cause of the disorder. Co-segregation of the detected mutations with the phenotype was confirmed by Sanger sequencing. In silico analysis including 3D protein structure modelling was used to predict the deleterious effects of the detected variants on protein function. We found two rare biallelic mutations in TWNK, encoding Twinkle, an essential mitochondrial helicase. Mutation c.1196A>G (p.Asn399Ser) recurred for the first time in a patient with PRLTS and the second mutation c.1802G>A (p.Arg601Gln) was novel for the disorder. In both patients neuroimaging studies showed diminished cervical enlargement of the spinal cord and for the first time in PRLTS partial atrophy of the vestibulocochlear nerves and decreased grey and increased white matter volumes of the cerebellum. Morphological changes in the auditory nerves, their desynchronized activity and partial cochlear dysfunction underlay the complex mechanism of hearing impairment in the patients. Our study unveils novel features on the phenotypic landscape of PRLTS and provides further evidence that the newly identified for PRLTS TWNK gene is involved in its pathogenesis.

POU3F4 mutations (DFNX2) are the most prevalent among non-syndromic X-linked hearing loss (HL) identified to date. Clinical manifestations of DFNX2 usually comprise congenital HL either sensorineural or mixed, a tendency towards perilymphatic gusher during otologic surgery and temporal bone malformations. The aim of the present study was to screen for POU3F4 mutations in a group of 30 subjects with a suggestive clinical phenotype as well as a group (N = 1671–2018) of unselected hearing loss patients. We also planned to analyze audiological and radiological features in patients with HL caused by POU3F4 defects. The molecular techniques used to detect POU3F4 mutations included whole exome sequencing (WES), Sanger sequencing and real-time polymerase chain reaction. Hearing status was assessed with pure-tone audiometry and auditory brainstem response. Computer tomography scans were evaluated to define the pattern of structural changes in the temporal bones. Six novel (p.Gln27*, p.Glu187*, p.Leu217*, p.Gln275*, p.Gln306*, p.Val324Asp) and two known (p.Ala116fs141*, p.Leu208*) POU3F4 mutations were detected in the studied cohort. All probands with POU3F4 defects suffered from bilateral, prelingual, severe to profound HL. Morphological changes of the temporal bone in these patients presented a similar pattern, including malformations of the internal auditory canal, vestibular aqueduct, modiolus and vestibule. Despite different localization in the POU3F4 gene all mutations severely impair the protein structure affecting at least one functional POU3F4 domain, and results in similar and severe clinical manifestations. Sequencing of the entire POU3F4 gene is recommended in patients with characteristic temporal bone malformations. Results of POU3F4 mutation testing are important not only for a proper genetic counseling, but also for adequate preparation and conduction of a surgical procedure.

Implementation of whole exome sequencing has provided unique opportunity for a wide screening of causative variants in genetically heterogeneous diseases, including nonsyndromic hearing impairment. TRIOBP in the inner ear is responsible for proper structure and function of stereocilia and is necessary for sound transduction. Whole exome sequencing followed by Sanger sequencing was conducted on patients derived from Polish hearing loss family. Based on whole exome analysis, we identified two TRIOBP pathogenic variants (c.802_805delCAGG, p.Gln268Leufs*610 and c.5014G>T, p.Gly1672*, the first of which was novel) causative of nonsyndromic, peri- to postlingual, moderate-to-severe hearing loss in three siblings from a Polish family. Typically, TRIOBP pathogenic variants lead to prelingual, severe-to-profound hearing loss, thus the onset and degree of hearing impairment in our patients represent a distinct phenotypic manifestation caused by TRIOBP variants. The pathogenic variant p.Gln268Leufs*610 disrupts the TRIOBP-4 and TRIOBP-5 isoforms (both expressed exclusively in the inner ear and retina) whereas the second pathogenic variant c.514G>T, p.Gly1672* affects only TRIOBP-5. The onset and degree of hearing impairment, characteristic for our patients, represent a unique phenotypic manifestation caused by TRIOBP pathogenic variants. Although TRIOBP alterations are not a frequent cause of hearing impairment, this gene should be thoroughly analyzed especially in patients with a postlingual hearing loss. A delayed onset of hearing impairment due to TRIOBP pathogenic variants creates a potential therapeutic window for future targeted therapies.