Genetic advances in Meniere Disease

Meniere Disease (MD) is an idiopathic inner ear disease with complex etiology and pathogenesis, which is still unclear. With the development in gene analysis technology, the genetic research of MD has attracted extensive attention, resulting in a large number of studies on the research of the relationship between human genes and MD. This paper aims to review the studies on this topic in recent years. The studies mainly focused on the genetics of familial MD and the correlation between MD and potentially related functional genes. The results of these studies have demonstrated the complexity and diversity of the pathogenesis of MD with both genetic and epigenetic alterations, suggesting that MD might be related to inflammation, immunity, aqua and ion balance in the lymphatic fluid, virus infection, metabolism, and abnormal function of nerve conduction. The finding of rare mutations in TECTA, MYO7A and OTOG genes and other genes such as CDH23, PCDH15 and ADGRV1 in the same families suggest that the integrity of the stereocilia and their interaction with the tectorial and otolithic membranes could be involved in the pathophysiology of familial MD.


Introduction
Meniere Disease (MD) is one of the most common disorders in otolaryngology, audiology and neurotology, first reported by Meniere Prosper in 1861, with endolymphatic hydrops as fundamental pathological basis. MD is characterized by intermittent paroxysm, fluctuating sensorineural hearing loss, tinnitus, and ear fullness, with unknown etiology and pathogenesis, and is associated with autoimmunity, mental stress, viral infection, anatomical factors, trauma, and genetic factors. One of the characteristics of MD, especially familial Meniere Disease (FMD) is the differences in incidence among races, regions, and family aggregation, suggesting that this disease may be related to genetic factors [1][2][3][4][5][6][7]. Many scholars believe that MD may be a multifactorial disease caused by interactions between one or more genes and environmental factors. Therefore, lots of researches have been done exploring genetics of MD, which mainly focused on FMD and the correlations between MD and functional genes. In this review paper, we focus on the progress in understanding the genetics of MD in details. Qingqing

Genetic studies of FMD
Gene mapping analysis Gabrikova et al. conducted a gene linkage analysis of a Swedish autosomal dominant FMD family and located the MD gene at 12p2.3 [8], but the results were inconsistent with those of a Finnish FMD linkage analysis [9]. The team used allele and haplotype association analysis to further locate MD to a 1.48 Mb region on 12p2.3 containing RERGL and PIK3C2G genes. RERGL gene is a member of the Ras family and is involved in cell signaling. As a member of the PI3Ks family, the PIK3C2G gene is involved in cell proliferation and other regulatory processes. The roles of the two genes in MD need to be further studied [10]. Arweiler et al. [11] located the MD gene on chromosome 5 based on linkage analysis results of 17 MD families. Fung et al. [12] located FMD-related genes in the HLA gene group of chromosome 6p through linkage analysis of two families. Frejo et al. [13] used immune genotyping array analysis to locate the MD target gene at 6p21.33, and the leading signal in the locus 6p21.33 was rs4947296. This region is a trans-expressed quantitative trait locus, possibly mediating the inflammatory response of MD by increasing NF-κB translation.

FMD candidate genes
Lopez-Escamez and his team sequenced the whole exome of FMD patients in Spain, and reported five candidate genes in four autosomal dominant FMD families. They described variants in two candidate genes, FAM136A and DTNA in a single family [2], a missense variant in gene PRKCB in another family that segregated the hearing loss phenotype [3], and rare heterozygous variants in the genes DPT and SEMA3D in another 2 families [4]. In addition, multiple rare missense mutations of the OTOG gene were found in 33% of familial MD, suggesting multiple allelic inheritances [5]. Mutation of FAM136A, DTNA, and DPT genes also appeared in some sporadic Meniere Disease (SMD) patients in South Korea [14]. Lopez-Escamez and his team also observed multiple families carrying rare variants in genes encoding proteins involved in the structure of the hair cells stereocilia and their attachment to the tectorial membrane (TM): six FMD families carrying rare missense heterozygous variants or a short deletion in the coding region of the TECTA gene encoding protein α-tectorin, which is one of the main non-collagenous proteins of the TM [5]; mutations in the MYO7A gene encoding cilia-motor proteins in inner ear hair cells co-segregated with some novel and rare variants in other genes involved in the organization of the stereocilia links such as CDH23, PCDH15 or ADGRV1 in nine Spanish and Swiss MD families [6]; 15 unrelated MD families carrying an enrichment of rare missense variants in the OTOG gene [5]. COCH gene, located at 14q12-13, encoding glycoprotein specific to acellular membranes of the inner ear and maybe involved in the organization and/or stabilization of the fibrillar network that compose the TM in the cochlea is an autosomal dominant gene for non-syndromic deafness. This gene mutation can cause the disease DFNA9 [15]. At least 10% of autosomal negative inherited non-syndromic sensorineural hearing loss is caused by DFNB16B gene mutation in STRC [16]. Both of them were accompanied by vestibular dysfunction and could show MD symptoms.
In addition, point mutations of EGFLAM and ITGA8 genes in autosomal dominant FMD families in China, P.y273N and P.L229F mutations of HMX2 genes in Finnish FMD families, and meaningless mutations of LSAMP genes in Iranian autosomal recessive FMD families have been reported [17][18][19].
FMD is inherited in an autosomal dominant, autosomal recessive, or mitochondrial genetic manner, and the differences in these findings suggest genetic heterogeneity among families of MD. These candidate genes are mostly expressed in nerve or inner ear tissues. Therefore, the pathogenesis of FMD may be to hinder nerve development, maintenance, or functional recovery by reducing the nutrient support of nerve endings to hair cells; influence neurite formation and growth; damage the inner ear cell structure; or interact with various proteins to affect hearing through changing lysosomal dynamics.

Study on the relationship between MD and genes
Immunity-, inflammation-, and stress-related protein genes

MD and histocompatibility leukocyte antigen genes
Major histocompatibility complex (MHC) is a group of genes encoding major histocompatibility antigens in animals. Human MHC is called histocompatibility leukocyte antigen (HLA). HLA gene complex located at 6p21.31 has a complex genetic polymorphism and is closely related to the immune response. The gene group can be divided into three types: Class I genes include HLA-A, B, C, E, and F, which exist on the surface of nucleated cells; Class II gene HLA-D consists of HLA-DR, DQ, and DP subregions, which mainly located on the surface of antigen-presenting cells; Class III genes are mostly complement-related genes, tumor necrosis factor (TNF), heat shock proteins (HSP), and transcription factor genes, which are located between class I and II genes. HLA-II genes are known to play important roles in most autoimmune diseases. Major Histocompatibility Complex Class I chain-related gene A (MICA) genes are atypical Class I MHC genes. MICA binds to receptors of some immune cells as a ligand, participating in immune response, and may playing an essential role in autoimmune diseases. Immunogical factors are considered to play an important role in the pathogenesis of MD, so there are many studies on the correlation between HLA and MD [20][21][22][23][24][25][26][27][28][29] (Table 1).
HLA association studies have not been replicated and they are pending confirmation. The differences in HLA expression among ethnic groups have not been ruled out. Furthermore, there are few large sample studies. Therefore, no significant breakthrough has been made. Whether HLA gene mutation is one of the causes of MD needs to be confirmed.

Other immunity-, inflammation-, and stress-related protein genes
In addition to HLA, a large number of other genes related to immunity, inflammation, and oxidative stress have been studied on the relationship with MD by scholars [7,[30][31][32][33][34][35][36][37][38][39][40] ( Table 2). Lopez-Escamez et al. [31] found that protein tyrosine phosphatase non-receptor type 22 (PTPN22) gene 1858 C/T, which strongly negatively affects T cell activation may be responsible for BMD among the Hispanic population. This finding supports the hypothesis that autoimmunity is one of the etiological factors of BMD. It was found that interleukin-1 A (IL-1 A) gene polymorphism, MIF-173 G/C gene polymorphism, and HSPA1A gene polymorphism were associated with MD susceptibility in the Japanese population [32,38,41]. In addition, Toll-like receptor 10 (TLR10) is involved in non-specific immune responses and the variant rs11096955 of TLR10 gene was associated with a better hearing prognosis in Spanish and Italian patients. NF-κB pathway gene NFKB1, histamine H4 receptor gene (HRH4), mediating the regulation of pro-inflammatory factors, and inflammatory disease-related gene RANTES were also found to be associated with MD [7,[35][36][37]. However, no studies have replicated these results.

Aquaporin and ion channel protein-related genes
Aquaporin subtypes 1-5, 7, and 9 are expressed in inner ear tissues to transport water and small soluble molecules such as glycerol. Potassium channel protein KCNE plays an important role in transmembrane ion and water transport in the inner ear. The pathological basis of MD is endolymphatic hydrops, suggesting that aquaporin and ion channel protein-related genes may be associated with MD. However, studies on AQP and KCNE gene polymorphisms showed opposite results in different populations (Table 3) [1,[42][43][44][45][46][47][48][49]. A meta-analysis based on current published studies showed that the KCNE1 rs1805127 and KCNE3 rs2270676 variants are not associated with the risk of MD [49]. Further replication studies in distinct populations are  Not correlated with MD [40] Chinese

HRH4
Highly expressed in the immune system, mediates the regulation of pro-inflammatory factors rs77485247 Associated with vertigo in MD and pro-inflammatory factor levels in blood [36] Iranian

Associated with inflammatory disease -403 A
Protective for male MD [37] TNF-α Pro-inflammatory factor gene -238 A/G Susceptible for MD [30] required to confirm the ethnic stratification of the association. Whether these two genes are related to MD needs to be confirmed by further large-sample studies.
Besides the genes mentioned above, Na+, K+-ATPase and sodium-calcium exchanger are vital to maintaining the ion balance of endolymph. In addition, Adducin is a kind of cytoskeleton protein, which contain subunits α, β, and γ, encoded by genes ADD1, ADD2, and ADD3, respectively. We already know that the gene polymorphism of ADD1 is related to salt-sensitive hypertension. Furthermore, the SIK1 gene encodes salt-induced kinase 1, which is related to Na+, K+-ATPase. Moreover, the SLC8A1 gene encodes a sodium-calcium exchanger. Teggi et al. found that ADD1, SIK1, and SLC8A1 gene polymorphisms were associated with MD. These results support the hypothesis that ion transport decompensation may cause MD (Table 3) [50,51].

Virus-associated genes
Host cytokine C1 (HCFC1) interacts with herpes simplex virus proteins and is involved in viral replication in nerve cells. Vrabec et al. [52] found that the frequency of the primary allele of HCFC1 SNPs increased in MD patients, and the secondary allele was a protective gene, suggesting that herpes virus infection might be a potential cause of MD.

Other candidate genes
Lopez-Escamez's team studied hundreds of patients with SMD in Spain and found that they had a high concentration of sensorineural deafness mutations in genes including GJB2, USH1G, SLC26A4, ESRRB and CLDN14. In addition, a rare synonymous mutation was found in another nonsyndromic deafness-related gene MARVELD2 among several unrelated MD patients, whose role in MD was unknown [53]. Missense mutations of axon-oriented signaling pathway-related genes NTN4 and NOX3 were also found [54]. We found low frequency of PARP-1 long allele (CA) < SUB > 17-20</sub > in BMD patients, suggesting its protective role in BMD [55].
Genetic variations of mitochondrial ribosomal genes TFB1M and MRPS12 are not associated with hearing loss in MD [56]. Polymorphisms of MTHFR C677T and A1298C genes associated with folic acid metabolism are related to susceptibility for MD in the Japanese population [57]. Polymorphism of Caveolin 1 (CAV1) gene, which encodes protein interacting with estrogen, was found to be significantly correlated with MD, suggesting that estrogen may be associated with the pathogenesis of MD [58].
In conclusion, SMD genetic variations are more complex than FMD. The probably related genes are inflammation-, autoimmune-related genes, ion channel-related genes, virus related genes, and nerve, metabolism-related genes, prompt that its pathogenesis may be related to inflammation,

Epigenetic studies of MD
Shew et al. [59] studied the miRNA in lymphatic fluid and serum of 10 patients with MD. They found that the levels of aquaporin, and inflammation & autoimmunity pathwayrelated proteins were higher than those in the control group, suggesting that the pathogenesis of MD may be related to both water imbalance and dysfunction of the immune system. Flook et al. [60] detected many differentially expressed methylated CpG islands in blood monocytes of Spanish SMD patients, some of which existed in causative genes of hearing loss, such as PCDH15, ADGRV1, and CDH23, while CpG island methylation deficiency existed in PHB genes. Combined with bioinformatic analysis, These epigenetic changes are considered to be associated with abnormal nerve electrical activity and inflammation in the inner ear.

Conclusion
The results of genetic studies on MD have demonstrated the complexity and diversity of the pathogenesis of MD, suggesting that MD might be related to inflammation, immunity, aqua and ion balance in the lymphatic fluid, virus infection, metabolism, and abnormal function of nerve conduction. The genetic study of MD is still facing various difficulties and challenges. Due to the low incidence of MD in many countries and races, the main subject of genetic research is the Caucasian population with a high incidence of MD. The period of collecting FMD patient samples is quite long due to the middle-aged onset of MD. In addition, due to the limitation of medical conditions and levels, so as the lack of understanding of MD, it is difficult to diagnose MD in many hospitals, hinding MD sample collection. Furthermore, few epigenetic studies have been performed on MD.
In conclusion, in the future, more races, more large samples of the disease, and more special testing techniques are needed to study the target genes at both genetic and epigenetic levels, in order to achieve more extensive and in-depth research.
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