Current approaches to the diagnosis and treatment - Cambridge ...

REVIEW

Expert Reviews in Molecular Medicine, Vol. 17; e9; 1 of 9. © Cambridge University Press, 2015 doi:10.1017/erm.2015.7

Current approaches to the diagnosis and treatment of white sponge nevus WENPING CAI1†, BEIZHAN JIANG2†, FANG YU2, JIANHUA YANG3, ZHENGHU CHEN3, JUNJUN LIU3, RONGBIN WEI3, SHOULIANG ZHAO1*, XIAOPING WANG2*, SHANGFENG LIU1* 1

Department of Stomatology, Huashan Hospital, Fudan University, Shanghai 200040, People’s Republic of China, Laboratory of Oral Biomedical Science and Translational Medicine, School of Stomatology, Tongji University, Shanghai 200072, People’s Republic of China, and 3Department of Ophthalmology, Shanghai Tenth People’s Hospital, Tongji University School of Medicine, Shanghai 200072, People’s Republic of China 2

White sponge nevus (WSN) in the oral mucosa is a rare autosomal dominant genetic disease. The involved mucosa is white or greyish, thickened, folded and spongy. The genes associated with WSN include mutant cytokeratin keratin 4 (KRT4) and keratin 13 (KRT13). In recent years, new cases of WSN and associated mutations have been reported. Here, we summarise the recent progress in our understanding of WSN, including clinical reports, genetics, animal models, treatment, pathogenic mechanisms and future directions. Gene-based diagnosis and gene therapy for WSN may become available in the near future and could provide a reference and instruction for treating other KRT-associated diseases.

Introduction White sponge nevus (WSN) is a rare hereditary leucokeratosis that was first described by Hyde in 1909 (Ref. 1); the term white sponge nevus was first coined by Cannon in 1935 (Ref. 2). The onset of WSN is early in life, and both sexes are affected equally. Lesions of WSN are easily recognised and clinically important; the lesions appear as bilateral white spongy plaques, typically found on the buccal mucosa, and the patients report no painful symptomatology. The lesion can be found in other common sites, including the tongue, floor of the mouth and alveolar mucosa (Ref. 3). The disease are characterised by white, thickened, folded and spongy lesions of the oral mucosa, although the oesophageal, laryngeal, nasal and anogenital mucosa can also be affected (Ref. 4). The WSN plaques are considered benign because the lesions are asymptomatic and painless in many cases, although they may undergo alternate periods of remission and exacerbation due to infections. This disorder often manifests in early childhood and exhibits no gender preference. WSN occur on the surface of the skin, including the oral mucosa and anal mucosa, among other areas. The pathogenesis of WSN is linked to a disorder of the epithelium and epithelial cells. Electron microscopic imaging of the oral mucosa of patients diagnosed with WSN reveals cellular keratinisation in the mucosa. Furthermore, McGininis observed keratinising epithelium in those tissues by focusing on the ultrastructural features of †

These authors equally contributed to this work.

the tissues (Ref. 5). Richard et al. and Rugg et al. first reported that WSN was related to abnormalities in genes encoding keratin (KRT) (Refs 6, 7). KRT is an intermediate filament (IF) cytoskeletal protein. The human genome contains 54 genes encoding functional KRTs, 37 of which encode epithelial KRTs and 17 of which encode hair KRTs. KRTs can be divided into acidic and basic forms. Both types are coexpressed during the differentiation of epithelial tissues and are arranged in heterotypic pairs to form chains of laterally aligned coiled-coil structures (Ref. 8). KRTs are types I and II IF proteins that form cytoskeletal networks within all epithelial cells. KRTs are expressed in pairs in a tissue and differentiation-specific fashion. KRTs, which play an important role in the cellular cytoskeleton, are the largest subgroup of IF proteins and are typically found in skin and other epithelial tissues. To date, KRTs have been shown to play regulatory roles in electrolyte transport, post-translational modifications and to confer protection against degradation (Ref. 9). KRTs can provide epithelia with mechanical support. Many KRT-related diseases have been reported (Table 1). KRT4 and KRT13 are the major differentiation-specific KRTs of the buccal mucosa and nasal, oesophageal and anogenital epithelia. The tissue distribution and nature of the lesions in patients affected by WSN suggest that mutations in KRT4 and/or KRT13 might be responsible for this disorder. Moreover, abnormal KRT mutations, such as KRT4

2

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

TABLE 1 KRT-RELATED DISEASE KRT KRT1, KRT10 KRT2e KRT2p KRT3, KRT12 KRT4, KRT13 KRT5, KRT14 KRT6a, KRT6c-f, KRT16 KRT6b, KRT17 KRT7 KRT8, KRT18 KRT9 KRT15 KRT19 KRT20 hHb6, hHb1 KRT6hf KRT6rs

Sites of primary expression

Diseases

Suprabasal cells of stratified cornified epithelia Late suprabasal cells of stratified cornified epithelia Hard palate-specific keratin Cornea-specific keratin Mucosa, stratified non-cornified epithelia, Basal keratinocytes of epidermis and stratified epithelia Palmoplantar, mucosa, wound healing, epidermal appendages Epidermal appendages Myopithelial cells, simple epithelia Simple epithelia Palmoplantar epidermis Basal keratinocytes Simple epithelia, epidermal appendages Gastrointestinal tract epithelia Hair follicle cells in the cortex Follicle cells with layers Inner root sheath of hair follicles

BCIE,DNEPPK IBS ? MCD WSN EBS PC-1, FNEPPK PC-2, SCM ? Occult patients with cirrhosis EPK ? ? ? Moniliform hair ? ?

and KRT13, are closely linked to WSN (Ref. 10). Both the oral and anogenital mucosae express type II KRT4 and its type I partner, KRT13. Furthermore, mutations of KRT4 and KRT13 were recently demonstrated to be the underlying cause of WSN. WSN in the oral mucosa is an autosomal dominant genetic disease. In recent years, there have been many reports of new cases and mutations. Here, we summarise recent research concerning WSN in the oral mucosa, including clinical pathology, genetics, animal models, treatment, prognosis and future directions of WSN research.

Clinical report To date, several hundreds of cases of WSN have been reported worldwide. WSN present as corrugated or velvety plaques on the oral, oesophageal or genital mucosa. We examined two WSN patients who presented with white lesions on the lips, tongue and buccal mucosa (Fig. 1). Maahs found to exhibit bilateral asymptomatic lesions. The tissue changes associated with WSN can be present at birth or can manifest during childhood or adolescence, and this disorder occurs in females more frequently than in males (Ref. 11). Jornet

a

WSN-patient 1

b

WSN-patient 2

Clinical symptom of two WSN patients Expert Reviews in Molecular Medicine © 2015 Cambridge University Press FIGURE 1. Clinical symptom of two WSN patients. (a) WSN patient-1 exhibited white lesions on the tongue and buccal mucosa; (b) WSN patient-2 exhibited white lesions on the lips and buccal mucosa.

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

presented a Spanish family consisting of a father and three boys affected with WSN. The proband, a 6year-old boy, exhibited white lesions on the buccal mucosa. His 11-year-old brother exhibited lesions in the vestibule floor and gums, and the 4-month-old brother exhibited lesion on the lip frenulum and oral buccal mucosa (Ref. 12). The most frequent sites of WSN are the buccal and labial mucosa, with a rare appearance on the palate. No extra-oral lesions were found. Lucchese described three cases diagnosed as WSN following the clinical and histological criteria that exhibited unusual appearances. In one case, the lesion was also present in the vaginal area (Ref. 13). Cheong reported the case of a 50-year-old Chinese woman with WSN signs on the buccal mucosa and genital mucosa (Ref. 14). Another case described a 28-year-old female with white plaques on the vulvar mucosa (Ref. 15). A few other cases have been reported independently without familial backgrounds. For example, in one case, the parent reported that the family history was notable for a 7-year-old cousin with similar lesions, but nobody else in the family exhibited such findings (Ref. 16). A 33-year-old male with a 13-year history of white, soft and folded plaques on the buccal mucosae and surfaces of the tongue was reported by Dadlani et al. (Ref. 17). Aghbali et al. report a case of WSN in a 21-year-old healthy Iranian male with no familial history who exhibited white bilateral lesion in the oral mucosa (Ref. 18). To date, only a few reports of WSN have been related to other inherited diseases. Dalben et al. first report the case of a 10-year-old boy diagnosed WSN and Ectrodactyly–Ectodermal Dysplasia–Clefting (EEC) syndrome. Subsequent studies focused on the pathogenesis of both WSN and EEC syndromes (Ref. 19). Legler et al. report a rare case of a 16-yearold American boy with a woolly hair nevus on his left parietal scalp, an associated epidermal nevus along Blaschko’s lines, and a white plaque on the central third of his ventral tongue who was clinically diagnosed as a WSN patient (Ref. 20). Further WSN research has revealed an increasing number of familial cases. These cases confirm that WSN is an autosomal dominant hereditary condition. DeTomas et al. reported 16 WSN cases from six families (Ref. 21). Martelli et al. report a three-generation family with WSN; clinical examination of family members revealed that of 23 descendants, 34.78% exhibited WSN features (Ref. 22). Another report presented a familial hereditary case of WSN in which two sisters (13 and 15 years old) exhibited white patches in the oral mucosa, tongue, lip and gum. Their mother exhibited similar lesions in her mouth when she was an adolescent (Ref. 23). Samatha et al. reported a familial WSN case in which oral lesions were found in three generations of a family (Ref. 24). Kimura et al. described a Japanese family; the proband was an 11year-old boy, and three other members of the family were affected by WSN (Ref. 25). Both genetic and environmental factors contribute to the development of WSN;

3

Marrelli et al. described a father (38 years old) and son affected by WSN whose oral diseases were related to a Staphylococcus aureus infection (Ref. 26). Lin et al. described two siblings affected by WSN who exhibited different clinical manifestations based on their smoking status. Theoretically, smoking may initiate exacerbation of WSN symptoms, but the authors reported that the degree of oral epithelial keratinisation among smokers was less than among non-smokers (Ref. 27). All of these clinical reports describe the clinical manifestations of the disease and provide a solid foundation for aetiology research.

Mutation report All of the reported mutations to date in exon 1A of the KRT4 and KRT13 gene are listed in Table 1. This disease may be hereditary. However, the pathogenesis of WSN remains unclear. Although the disease name refers to moles, the disorder is not associated with features of moles. Furthermore, the symptoms are caused by normal keratinisation (Ref. 7). Researchers have found that the abnormal expressions of several KRTs, mainly KRT4 or KRT13, play an important role in the generation of WSN (Ref. 28). In Terrinoni’s study, the authors investigated a new KRT4 gene mutation (3 bp (ACA) insertion) and detected an amino acid insertion in exon 1A (Ref. 29). Another mutation identified was a heterozygous missense mutation 1345G > A in the KRT4 gene, which caused an amino acid substitution Glu449Lys in exon 2B (Ref. 30). Zhang et al. described WSN in a Chinese family and identified two new KRT4 mutations. One mutation was 1829G > A, which is predicted to cause the amino acid substitution Glu520Lys. The second mutation was 2324A > G in the non-coding region (Ref. 31). Several other mutations of the KRTs have been close linked to WSN. The KRT4 gene mutation 1558G > A in the 2B region is predicted to change amino acid Glu520 to a Lys (Ref. 32). Kimura et al. detected a mutation (3 bp AAC deletion) in the KRT4 gene (Asn160Del) (Ref. 25). Richard et al. first identified the missense mutation 356T > C in the 1A region of the helical rod domain of KRT13 that led to Leu115Pro, which is linked to WSN (Ref. 33). Rugg et al. identified two novel KRT13 mutations: a familial 344T > C mutation causing a Leu115Pro substitution and a familial 323T > C mutation causing a Met118Thr substitution (Ref. 34). The heterozygous missense mutation 335A > G was detected in exon 1 of the KRT13 gene and was predicted to cause the amino acid change Asn112Ser in the 1A domain of the KRT13 polypeptide (Ref. 35). Shibuya et al. identified a mutation at 332T > C, resulting in a Leu111Pro substitution (Ref. 36). Nishizawa et al. analysed the KRT13 mutation 341G > A, which was predicted to cause the amino acid substitution Arg114His (Ref. 37). Liu et al. described a mutational analysis in familial and sporadic patients with WSN and identified the novel

4

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

mutation 340C > T, causing the amino acid substitution Arg114Cys (Ref. 38). In our study, we identified two heritable mutations in the KRT13 gene that were associated with the development of WSN in two

insert ACA at 419 and 420 Shift frame

a

436–438 delAAC N146del

Chinese families: 332T > C (Leu111Pro) and 340C > T (Arg114Cys) (Ref. 39). We have described five mutations in the KRT4 gene (Fig. 2a) and seven heritable mutations in the KRT13

478–480 delCAA Q160del

G1303A E435K

G1336A E446K

1 bp 5’

KRT4 Exon 1

b

2

3

6

5

4

7

8

Exon 9

C340T R114C A335G N112S T332C L111P

G341A R114H T344C L115P

T323C M108T

T356C L119P

1688 bp

1 bp 5’

c

2147 bp 3’

KRT13 Exon 1

2

3

4

5

6

7

Exon 8

3’

Armadillo KRT13 Rabbit KRT13 Cow KRT13 Sheep KRT13 Dolphin KRT13 Elephant KRT13 Ferret KRT13 Panda KRT13 Dog KRT13 Cat KRT13 Chimpanzee KRT13 Human KRT13 Gorilla KRT13 Gibbon KRT13 Marmoset KRT13 Bushbaby KRT13 Squirrel KRT13 Mouse KRT13 Rat KRT13 Kangaroo KRT13 Pig KRT13 Hedgehog KRT13 Microbat KRT13 Opossum KRT13 Wallaby KRT13 Tasmanian KRT13 Platypus KRT13 Zebrafish KRT13

WSN-related gene mutations and homology analysis of KRT13 Expert Reviews in Molecular Medicine © 2015 Cambridge University Press FIGURE 2. WSN-related gene mutations and homology analysis of KRT13. (a) There were five mutations in the KRT4 gene on exons 1, 2 and 7; (b) There were seven heritable mutations in the KRT13 gene on exon 1; (c) The homology analysis of the KRT13 protein sequence from 29 species. Some amino acids from exon 1 of the KRT13 sequences of the 29 species were relatively conserved.

5

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

TABLE 2 WSN-RELATED GENE MUTATIONS Genes

KRT13

KRT4

Nucleotide changes T323C T332C A335G C340T G341A T344C T356C Insert ACA at 419 and 420 436–438 del AAC 478–480 del CAA G1336A G1303A

Chromosome location

Exon position

Amino acid changes

References

17q21.2 17q21.2 17q21.2 17q21.2 17q21.2 17q21.2 17q21.2 12q13.13 12q13.13 12q13.13 12q13.13 12q13.13

Exon 1 Exon 1 Exon 1 Exon 1 Exon 1 Exon 1 Exon 1 Exon 1 Exon 1 Exon 2 Exon 7 Exon 7

M108T L111P N112S R114C R114H L115P L119P Shift frame N146del Q160del E446K E435K

(34) (36) (35) (38) (37) (34) (6) (29) (7) (38) (31) (30)

gene (Fig. 2b) that are associated with WSN (Table 2). We analysed the homology of the KRT4 protein sequence from 43 species and the KRT13 protein sequence from 29 species, and we performed a multiple alignment of the KRT13 protein sequence from 29 species using the ClustalW program (http://www. genome.jp/tools/clustalw/). Some of the amino acids from exon1 of the KRT13 sequences of 29 species were relatively conserved (Fig. 2c). However, there were no highly similar fragments of the KRT4 protein sequences among the 43 species.

WSN animal model The WSN animal model focuses on KRT4. Ness used KRT4-deficient mice to confirm that KRT4 is an important protein for internal epithelial integrity. In humans, KRT4 mutations lead to WSN, and KRT4-deficient mice may serve as ideal models for WSN to examine the changes in KRT4 when cells proliferate and differentiate (Ref. 40). McGowan et al. identified a missense mutation in mutant animals (bright coat colour 1 (Bcc1)) that predicts an Asn154Ser amino acid substitution in the 1A domain of KRT4. Bcc1 recapitulates the histopathology and human genetics of WSN (Ref. 41). We should improve the development of animal models associated with WSN (KRT4 and KRT13). It is necessary to establish an ideal animal model for further research on WSN, which will provide a platform for drug screening and clinical treatment. Treatment and prognosis Although WSN patients experience no significant physical pain, they often complain of an altered texture to their mucosa or dissatisfaction with the appearance of the lesions. Many WSN patients undergo therapeutic treatments with nystatin, antihistamines, vitamins and mouth rinses. Azithromycin, tetracycline and penicillin have exhibited some clinical effects (Ref. 42). However, there is no standard treatment protocol for WSN to date. A case of WSN exhibiting significant improvement following penicillin administration has been reported (Ref. 43). McDonagh et al. first reported that tetracycline

medicines were effective in four WSN patients (Ref. 44). Another report stated that oral tetracycline rinse improved the symptoms of WSN (Ref. 45). Subsequently, WSN was successfully treated with a tetracycline mouth rinse (Ref. 46). Long-term lowdose systemic antibiotic therapy maintained the remission of WSN (Ref. 47). Victoria A acid can also inhibit keratosis formation (Ref. 48). Quintella et al. described a WSN patient who received orthodontic treatment in parallel with a sibling (Ref. 49). Otobe successfully treated a WSN patient with concurrent systemic lupus erythematosus using a topical tetracycline (Ref. 50). Four cases also improved with tetracycline mouth rinse (Ref. 51). To date, systemic antibiotics or local applications of retinoic acid have provided limited benefits, but both are poorly effective. Dufrasne et al. described a case of effective surgical resection, and the patient was free of recurrence 2 years later (Ref. 52). These methods together with the iPS technology may point to a treatment for this rare disease. WSN may be treated with chlorhexidine (Ref. 53). Meanwhile, WSN patients should perform a careful oral hygiene to reduce infection in the oral cavity. The proper diagnosis and treatment of this rare disease will require the combination of clinical history, clinical examination and pathologic findings.

Discussion To date, several hundreds of cases of WSN have been reported worldwide, including in China (Refs 30, 31, 39), Japan (Refs 25, 32), Italy (Refs 13, 26, 29), Spain (Ref. 12), Iran (Ref. 18), Caucasia (Refs 15, 44), Brazil (Ref. 23), Turkey (Ref. 42) and Scotland (Ref. 54). The KRT4 and KRT13 gene mutations have been demonstrated to be involved in the pathogenesis of WSN. Because WSN in the oral mucosa is an autosomal dominant genetic disease, the percentage of the affected population in a family is generally greater than 30%. The hazards of this disease are much larger and more widespread than we previously appreciated. In Table 2, we summarise the reported mutations of KRT4 and KRT13 in WSN patients. Because most KRT-related mutations occur in pairs (Refs 55, 56),

6

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

cytoskeleton in mucosal epithelial cells leads to the massive hyper-proliferative changes observed in WSN and the KRT4 knockout mice is unclear. Previous work revealed that when one member of a pair of KRTs is absent, its natural partner tends to be degraded. Due to differences between humans and mice, the regulation of the members of the KRT4 and KRT13 pair may be different. Studies of WSN disease and drugs are largely dependent on animal models. Genetically modified animal models provide important insights into the pathogenesis of human diseases. These models are also essential in developing new drugs. Classical mouse models have promoted significant developments in biomedicine. However, mice and humans are different in many aspects, including physiological traits and gene expression. Thus, mouse models cannot sufficiently mimic human diseases in some cases, and large animal models remain

we expect that WSN-related genes will be limited to KRT4 and KRT13. However, new mutations in KRT4 and KRT13 are likely to be found in the near future. Our recent work suggests that the sequences of the KRT13 protein are highly conserved in the α-helix 1A domain from G102 to I136 in 23 diverse species, which represents the region in which all mutations of KRT13 have been found. Thus, this area represents a ‘mutation hotspot’ in the KRT13 gene. However, KRT4 does not exhibit a ‘mutation hotspot’. New mutations in this region will undoubtedly be identified, and the majority of genetic diagnosis will focus on this hotspot. In a recently reported mouse model, the murine KRT4 gene was ablated by gene targeting. These animals exhibited a marked increase in mucosal cell division, as shown by staining for proliferating cell nuclear antigen (Ref. 40). Precisely how the defective

Molecular biology

Pedigree collection

Clinical detection

Cell modle platform

Establish cell line

WSN-fibroblast

Histopathologic

Animal modle platform

Pathogenic mechanism

iPSC

Drug screening

RNA-seq

Yamanaka’s

Animal model

iPSC Lentivirus

WT-fibroblast

Blood

PCR-Seq

Expression profiling

RT-PCR-seq

New drug screening

Oral epithelium cells

Western Blot Co-IP-MS

Rescue experiments

Mutation analysis

Signal pathway

Further vetify the pathogenic mechanism

Research methods, thoughts and future direction of WSN Expert Reviews in Molecular Medicine © 2015 Cambridge University Press FIGURE 3. Research methods, thoughts and future direction of WSN.

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

urgently needed (Refs 57, 58, 59). Pigs have been recently used as models for human diseases because they are more similar to humans than mice in terms of anatomy, neurobiology, cardiac vasculature, gastrointestinal tract and genome (Ref. 60). Pigs also present fewer ethical and economic concerns compared with primates. To date, several genetically modified pig models for human diseases have been produced, including models of Huntington’s disease, Alzheimer’s disease, spinal muscle atrophy, cardiovascular disease, diabetes mellitus (MODY3), diabetes mellitus type 2, retinitis pigmentosa, Stargardtlike macular dystrophy type 3 and breast cancer (Ref. 61). With respect to the WSN disease animal model, further considerations are necessary because the molecular mechanism of this process in vivo remain incompletely understood. In our future work, we will use the CRISPR/Cas9 system to build a KRT13 T332C WSN transgenic pig model and explore the pathogenesis of WSN. We will apply phenotype reversal comprehensive treatment and drug screening to interpret and validate the molecular pathogenesis of WSN in vivo. This approach will provide an important reference for further exploration of the molecular mechanism of WSN as well as its clinical treatment and drug screening. Lentiviruses using HIV-1 as a resource exhibit superiority over other transgenic technologies with respect to infection efficiency, stable expression, weak antigenicity, high degree of manoeuvrability, capability of containing large sizes of gene fragment, and infection of cells either in dividing and non-dividing phases (Refs 62, 63). In gene therapy, numerous studies have confirmed that lentiviral vectors can be used for robust and highly efficient gene transfer into organs and tissues that are out of reach for many other gene delivery systems. However, there has been no related report about the therapy or study of WSN. To develop better therapeutic trials for WSN, it is important to understand the consequences of the genetic mutations and molecular changes associated with WSN. Human-induced pluripotent stem cells (iPSCs) represent an excellent tool for this work with many advantages for research (Ref. 64). With the development of iPSC, many completed clinical trials have demonstrated the efficacy of mesenchymal stem cell (MSC) infusion for diseases, including acute myocardial ischaemia, stroke, liver cirrhosis, amyotrophic lateral sclerosis and graft-versus-host disease (Ref. 65). However, there has been no related report about the application of WSN. We obtained WSN iPSC by transforming human WSN fibroblast cells and wild-type fibroblasts into iPSC, and the iPSC could be transformed into human epidermal cells for drug screening or mechanism research. Another cell research platform for WSN is MSCs. Recently, we have succeeded in isolating MSCs from patient gum tissue with the KRT13 point mutation (Ref. 39). Many other biotechnologies

7

can be used based on this cell platform, including mass spectrometry (Co-IP-MS), RNA-seq and weighted gene co-expression network analysis (Ref. 66). The results would provide a valuable resource to dissect the gene regulatory mechanisms underlying the progressive development of WSN. This approach could identify genes downstream of KRT4 and KRT13, clarifying the pathogenesis of WSN. Meanwhile, our work suggests that the abnormal degradation of the KRT13 protein in WSN is likely associated with an abnormal ubiquitination process. Together, these efforts may contribute to molecular therapy for WSN. To the best of our knowledge, there remains no appropriate animal model or effective therapeutic drugs for WSN. The mechanism underlying WSN remains unclear. Almost all studies have focused on case reports or mutation reports. Prenatal diagnosis seems to be the only method to reverse the phenotype in affected families. We have described the research methods, thoughts and future research directions concerning WSN (Fig. 3). With further research into WSN, there is optimism that the problem will be solved in the next decade.

Acknowledgements This work was supported, in whole or in part, by the National Basic Research Programme (973 Programme, No. 2013CB967501), the Natural Science Foundation of Shanghai (No. 12ZR1434200), the Fundamental Research Funds for the Central Universities (No. 20120072110016), the Key Programme of Shanghai (No. 074119614) and the Shanghai Science and Technology Commission Programme (Nos124119A7400 and114119A3400).

Competing interests The authors declare that they have no competing interests. References 1. Hyde J. (1909) An unusual naevus of the tongue in a five-yearold boy. Journal of Cutaneous Diseases 27, 256 2. Cannon A. (1935) White sponge nevus of the mucosa (naevus spongiosus albus mucosae). Archives of Dermatology 31, 365 3. Frithiof L. and Bánóczy J. (1976) White sponge nevus (leukoedema exfoliativum mucosae oris): ultrastructural observations. Oral Surgery, Oral Medicine, Oral Pathology 41, 607-622 4. Jorgenson R.J. and Levin L.S. (1981) White sponge nevus. Archives of Dermatology 117, 73 5. McGinnis J.P. and Turner J.E. (1975) Ultrastructure of the white sponge nevus. Oral Surgery, Oral Medicine, Oral Pathology 40, 644-651 6. Richard G. et al. (1995) Keratin 13 point mutation underlies the hereditary mucosal epithelia disorder white sponge nevus. Nature Genetics 11, 453-455 7. Rugg E. et al. (1995) A mutation in the mucosal keratin K4 is associated with oral white sponge nevus. Nature Genetics 11, 450-452 8. Moll R. et al. (2008) The human keratins: biology and pathology. Histochemistry & Cell Biology 129, 705-733 9. Majumdar D. et al. (2012) Keratins in colorectal epithelial function and disease. International Journal of Experimental Pathology 93, 305-318 10. Smith F.J. (2003) The molecular genetics of keratin disorders. American Journal of Clinical Dermatology 4, 347-364 11. Maahs M. et al. (2007) White sponge nevus. A case report. Minerva Stomatologica 56, 649

8

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

12. Jornet P.L. (2008) White sponge nevus: presentation of a new family. Pediatric Dermatology 25, 116-117 13. Lucchese A. and Favia G. (2006) White sponge naevus with minimal clinical and histological changes: report of three cases. Journal of Oral Pathology & Medicine 35, 317-319 14. Cheong M.L. (2009) Hereditary mucosal leukokeratosis. International Journal of Dermatology 48, 1001-1002 15. Cutlan J. et al. (2010) White sponge nevus presenting as genital lesions in a 28-year-old female. Journal of Cutaneous Pathology 37, 386-389 16. Polcari I. and Chamlin S. (2006) A 6-month-old boy with white oral lesions. Pediatric Annals 35, 874-876 17. Dadlani C. et al. (2008) White sponge nevus. Dermatology Online Journal 14, 16 18. Aghbali A. et al. (2009) White sponge nevus: a case report. Journal of Dental Research, Dental Clinics, Dental Prospects 3, 70 19. Dalben G. et al. (2010) White sponge nevus in a patient with EEC syndrome. Dermatology Online Journal 16, 7 20. Legler A. et al. (2010) Woolly hair nevus with an ipsilateral associated epidermal nevus and additional findings of a white sponge nevus. Pediatric Dermatology 27, 100-101 21. DeTomas M. et al. (1999) White sponge nevus: presentation of sixteen cases corresponding to six families. Med Oral. 4, 494-502 22. Martelli H.J. et al. (2007) White sponge nevus: report of a three-generation family. Oral Surgery Oral Medicine Oral Pathology Oral Radiology Endodontics 103, 43 23. Martins F.P. et al. (2011) Familial case of oral white sponge nevus: a rare hereditary condition. Anais Brasileiros de Dermatologia 86, 39-41 24. Samatha Y. et al. (2012) Familial white sponge nevus of the oral mucosa: report of occurrence in three generations. Quintessence International 43, 319 25. Kimura M. et al. (2012) Mutation of keratin 4 gene causing white sponge nevus in a Japanese family. International Journal of Oral Maxillofac Surgery 42, 615-618 26. Marrelli M. et al. (2012) Oral infection by Staphylococcus aureus in patients affected by White Sponge Nevus: a description of two cases occurred in the same family. International Journal of Medical Science 9, 47 27. Lin D. et al. (2013) Siblings present different clinical manifestations in white sponge nevus. West China Journal of Stomatology 31, 432-433 28. Su L. et al. (1993) Expression of keratin 14 and 19 mRNA and protein in normal oral epithelia, hairy leukoplakia, tongue biting and white sponge nevus. Journal of Oral Pathology & Medicine 22, 183-189 29. Terrinoni A. et al. (2000) A glutamine insertion in the 1A alpha helical domain of the keratin 4 gene in a familial case of white sponge nevus. Journal of Investigative Dermatology 114, 388391 30. Chao S. et al. (2003) A novel mutation in the keratin 4 gene causing white sponge naevus. British Journal of Dermatology 148, 1125-1128 31. Zhang J. et al. (2009) Two new mutations in the keratin 4 gene causing oral white sponge nevus in Chinese family. Oral Diseases 15, 100-105 32. Shimizu A. et al. (2012) White sponge nevus caused by a missense mutation in the keratin 4 gene. European Journal of Dermatology 22, 571-572 33. Richard G. et al. (1995) Keratin-13 point mutation underlies the hereditary mucosal epithelia disorder white sponge nevus. Nature Genetics 11, 453-455 34. Rugg E. et al. (1999) Identification of two novel mutations in keratin 13 as the cause of white sponge naevus. Oral Diseases 5, 321-324 35. Terrinoni A. et al. (2001) A novel mutation in the keratin 13 gene causing oral white sponge nevus. Journal of Dental Research 80, 919-923 36. Shibuya Y. et al. (2003) Constitutional mutation of keratin 13 gene in familial white sponge nevus. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology and Endodontics 96, 561-565 37. Nishizawa A. et al. (2008) A de novo missense mutation in the keratin 13 gene in oral white sponge naevus. British Journal of Dermatology 159, 974-975

38. Liu X. et al. (2011) Mutational analysis in familial and sporadic patients with white sponge naevus. British Journal of Dermatology 165, 448-451 39. Cai W. et al. (2014) Keratin 13 mutations associated with oral white sponge nevus in two Chinese families. Meta Gene 2, 374-383 40. Ness S.L. et al. (1998) Mouse keratin 4 is necessary for internal epithelial integrity. Journal of Biological Chemistry 273, 23904-23911 41. McGowan K.A. et al. (2006) Identification of a Keratin 4 mutation in a chemically induced mouse mutant that models white sponge nevus. Journal of Investigative Dermatology 127, 60-64 42. Songu M. et al. (2012) White sponge nevus: clinical suspicion and diagnosis. Pediatric Dermatology 29, 495-497 43. Alinovi A. et al. (1982) White sponge nevus: successful treatment with penicillin. Acta Dermato-Venereologica 63, 83-85 44. McDonagh A. et al. (1990) White sponge naevus successfully treated with topical tetracycline. Clinical and Experimental Dermatology 15, 152-153 45. Lim J. and Ng S.K. (1992) Oral tetracycline rinse improves symptoms of white sponge nevus. Journal of the American Academy of Dermatology 26, 1003-1005 46. Becker L. et al. (1997) White sponge naevus successfully treated with tetracycline mouth rinse. Acta DermatoVenereologica 77, 413 47. Lamey P.J. et al. (1998) Oral white sponge naevus: response to antibiotic therapy. Clinical and Experimental Dermatology 23, 59-63 48. Irvine A. and McLean W. (1999) Human keratin diseases: the increasing spectrum of disease and subtlety of the phenotype–genotype correlation. British Journal of Dermatology 140, 815-828 49. Quintella C. et al. (2004) Orthodontic therapy in a patient with white sponge nevus. American Journal of Orthodontics and Dentofacial Orthopedics 125, 497-499 50. Otobe I.F. et al. (2006) Successful treatment with topical tetracycline of oral white sponge nevus occurring in a patient with systemic lupus erythematosus. International Journal of Dermatology 45, 1130-1131 51. Otobe I. et al. (2007) White sponge naevus: improvement with tetracycline mouth rinse: report of four cases. Clinical and Experimental Dermatology 32, 749-751 52. Dufrasne L. et al. (2011) Current therapeutic approach of the white sponge naevus of the oral cavity. Bulletin du Groupèment international pour la recherche scientifique en stomatologie & odontologie 50, 1-5 53. Satriano R.A. et al. (2012) White sponge nevus treated with chlorhexidine. Journal of Dermatology 39, 742-743 54. Terrinoni A. et al. (2001) A novel mutation in the keratin 13 gene causing oral white sponge nevus. Journal of Dental Research 80, 919 55. Rugg E.L. and Leigh I.M. (2004) The keratins and their disorders. American Journal of Medical Genetics C Seminars in Medical Genetics 131C, 4-11 56. Cordon L.D. and McLean W.H.I. (1996) Human keratin diseases. Experimental Dermatology 5, 297-307 57. Verma N. et al. (2011) Recent advances in the use of Sus scrofa (pig) as a model system for proteomic studies. Proteomics 11, 776-793 58. Walters E.M. et al. (2011) Animal models got you puzzled?: think pig. Annals of the New York Academy of Sciences 1245, 63-64 59. Li X.J. and Li W. (2012) Beyond mice: genetically modifying larger animals to model human diseases. Journal of Genetics & Genomics 39, 237-238 60. Bendixen E. et al. (2010) Advances in porcine genomics and proteomics e a toolbox for developing the pig as model organism for molecular biomedical research. Briefings in Functional Genomics 9, 208-219 61. Fan N. and Lai L. (2013) Genetically modified pig models for human diseases. Journal of Genetics & Genomics 40, 67-73 62. Wiznerowicz M. and Trono D. (2005) Harnessing HIV for therapy, basic research and biotechnology. Trends in Biotechnology 23, 42-47 63. Allie A. et al. (2010) Oral medicine case book 25. White sponge naevus. SADJ: Journal of the South African Dental Association = tydskrif van die Suid-Afrikaanse Tandheelkundige Vereniging 65, 130

CURRENT APPROACHES TO THE DIAGNOSIS AND TREATMENT OF WHITE SPONGE NEVUS

64. Takahashi K. and Yamanaka S. (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126, 663-676 65. Wang S. et al. (2012) Clinical applications of mesenchymal stem cells. Journal of Hematology & Oncology 5, 19 66. Xue Z. et al. (2013) Genetic programs in human and mouse early embryos revealed by single-cell RNA sequencing. Nature 500, 593-597

9

∗ Corresponding author: Shangfeng Liu, Department of Stomatology, Huashan Hospital, Fudan University, Shanghai 200040, People’s Republic of China Tel: 86-21-65986790; Fax: 86-21-65986790; E-mail: [email protected]

Xiaoping Wang, Laboratory of Oral Biomedical Science and Translational Medicine, School of Stomatology, Tongji University, Shanghai 200072, People’s Republic of China Tel: 86-21-65986073; Fax: 86-21-65986073; E-mail: [email protected] Shouliang Zhao, Department of Stomatology, Huashan Hospital, Fudan University, Shanghai 200040, People’s Republic of China Tel: 86-21-52887813; Fax: 86-21-52887813; E-mail: [email protected]