Inherited Congenital Cataract: A Guide to Suspect the Genetic Etiology in the Cataract Genesis

Abstract

Cataracts are the principal cause of treatable blindness worldwide. Inherited congenital cataract (CC) shows all types of inheritance patterns in a syndromic and nonsyndromic form. There are more than 100 genes associated with cataract with a predominance of autosomal dominant inheritance. A cataract is defined as an opacity of the lens producing a variation of the refractive index of the lens. This variation derives from modifications in the lens structure resulting in light scattering, frequently a consequence of a significant concentration of high-molecular-weight protein aggregates. The aim of this review is to introduce a guide to identify the gene involved in inherited CC. Due to the manifold clinical and genetic heterogeneity, we discarded the cataract phenotype as a cardinal sign; a 4-group classification with the genes implicated in inherited CC is proposed. We consider that this classification will assist in identifying the probable gene involved in inherited CC.

Cataracts cause half of all cases of blindness and one-third of the visual impairment cases worldwide. The term cataract comes from the Latin word “cataracta” that itself derives from the Greek “katarráktés” and refers to a waterfall. Apparently, Constantine the African (a Carthaginian monk from the Monte Cassino monastery and a member of the Salerno school of translators) borrowed this term for lens opacity from the translation of Arab medical writings into Latin. Cataract is a term that is still used today [Chance, 1939]. Since cataracts are the most common cause of vision loss in the world, surgical management of this has been documented since ancient times. Possibly, the first written evidence is from 600 years B.C.: “Uttara-Tantra of Sushruta Samshita,” wherein the recline method technique is described [Bhishagratna, 2005].

A cataract is defined as an opacity of the lens resulting from a variation of the refractive index of the lens. This variation derives from changes in the lens structure resulting in light scattering, frequently due to a significant concentration of high-molecular-weight protein aggregates. Congenital cataract (CC), one of the leading causes of treatable blindness worldwide, has a prevalence of 1–15/ 10,000 live births with a greater presence in developing countries than in developed countries [Apple et al., 2000; Gilbert and Foster, 2001]. Inherited CC is a clinically and genetically heterogeneous disease normally associated with the breakdown of the lens architecture. Inherited CC presents all types of inheritance patterns in a syndromic and nonsyndromic form with more than 100 causative genes (http://cat-map.wustl.edu/). For example, hereditary CC may be inherited as autosomal dominant, autosomal recessive, or X-linked traits, with autosomal dominant being the most prevalent form of inheritance pattern. Hereditary CC shows an important intrafamilial and interfamilial variability; the same mutation in one gene can result in distinct cataract phenotypes, whereas expression of different genes can produce the same cataract pattern. Clinical variability can even be observed in the same patient [Hejtmancik and Smaoui, 2003].

Several terms have been used to describe the different types of hereditary CC in the literature. CC classification has not been homogenous. In some cases, the description corresponds to the name of the author who first described the cataract, i.e., Marner cataract, a cataract with sutural opacities [Marner et al., 1989]; to the name of the affected family such as Coppock cataract, an embryonic nuclear cataract [Nettleship and Ogilvie, 1906]; the Volkman cataract, a central or zonular variety with opacities in the embryonic, fetal, and juvenile nucleus [Eiberg 1995], or to the affected community such as the Hutterite cataract [Boone et al., 2015]. In other cases, anatomic localization is used to establish the cataract definition, i.e., subcapsular, nuclear, sutural, cortical, fetal, embryonic, or capsular cataract (Fig. 1).

Fig. 1.

Some examples of inherited cataracts: anterior subcapsular (A), fetal nuclear (B), punctate in lens cortex (C), and embryonic nuclear Y-sutural (D).

There are excellent reviews of inherited cataracts in the literature. The purpose of this review is to provide a guide to the suspected genetic causes of inherited CC. Due to the great clinical and genetic heterogeneity, we separated the genes implicated in the process of cataractogenesis into 4 groups (Tables 1, 2, 3, 4). These groups contain the genes involved in syndromic cataracts, the genes implicated in syndromic cataracts but with reports of congenital cataracts only, the genes that present cataracts only, and finally, the genes with cataract and eye anomalies. We consider that this classification will assist in identifying the probable gene involved in inherited CC.

Table 1.

Syndromic cataract genes

Locus	Gene	Inheritance	OMIM	Disease	Gene product
1p36.32	PEX10	AR	614870	neonatal adrenoleukodystrophy, Zellweger syndrome	protein of peroxisomal matrix
1p36.22	PEX14	AR	614887	Zellweger syndrome	peroxisomal import machinery
1p36.1p34	HSPG2	AR	224410/25800	Schwartz-Jampel syndrome type 1/dyssegmental dysplasia	perlecan protein
1p36p35	GALE	AR	230350	galactose epimerase deficiency	UDP-galactose-4-epimerase
1p34.1	POMGNT1	AR	253280	muscular dystrophy-dystroglycanopathy	type 2 transmembrane protein that resides in the Golgi apparatus
1p22p21	ABCD3	AR	616278	Zellweger syndrome 2	member of the superfamily of ATP-binding cassette transporters
1p21	COL11A1	AD	604841/154780	Stickler syndrome type 2, Marshall syndrome cataracts, growth hormone deficiency, sensory neuropathy, sensorineural hearing loss, skeletal dysplasia	one of the 2 alpha chains of type XI collagen
1q41	IARS2	AR	616007		Aminoacyl-tRNA synthetase
1q41	RAB3GAP2	AR	212720/614225	Martsolf syndrome, Warburg micro syndrome	RAB3 protein regulates exocytosis of neurotransmitters and hormones
1q42	GNPAT	AR	222765	rhizomelic chondrodysplasia punctata type 2	enzyme in synthesis of ether phospholipids
2p14p16	PEX13	AR	614883	Zellweger syndrome	peroxisomal membrane protein
2q21.3	RAB3GAP1	AR	600118	Warburg micro syndrome	catalytic subunit of a Rab GTPase activating protein
2q24q31	LRP2	AR	222448	Donnai-Barrow syndrome	low density lipoprotein-related protein 2
2q33qter	CYP27A1	AR	213700	cerebrotendinous xanthomatosis	member of the cytochrome P450 superfamily
2q37	KCNJ13	AD	193230	snowflake vitreoretinal degeneration	member of the inwardly rectifying potassium channel family
3p21.1	COL7A1	AR	226600	epidermolysis bullosa dystrophica	alpha chain of type VII collagen
3p14.3	FLNB	AD, AR	150250/272460	Larsen syndrome/spondylocarpotarsal synostosis syndrome	member of the filamin family
3q21q22	CNBP	AD	602668	myotonic dystrophy type 2	a nucleic-acid binding protein with 7 zinc-finger domains
3q25	SLC33A1	AR	614482	congenital cataracts, hearing loss, neurodegeneration	required for the formation of O-acetylated (Ac) gangliosides
4p16.1	HMX1	AR	612109	oculoauricular syndrome	transcription factor that belongs to the H6 family of homeobox proteins
4p15.32	CC2D2A	AR	612285	Joubert syndrome 9	play a critical role in cilia formation
4p12q12	SRD5A3	AR	612713	Kahirazi syndrome	steroid 5-alpha reductase family
4q32q35	ETFDH	AR	231680	glutaric acidemia	component of the electron-transfer system in mitochondria
4q35.1	TRAPPC11	AR	615356	muscular dystrophy limb girdle 2S	a subunit of the TRAPP (transport protein particle) tethering complex
5q12.1	ERCC8	AR	216400	Cockayne syndrome type A	a WD repeat protein
5q14.3	VCAN	AD	143200	Wagner syndrome 1	a member of the aggrecan/versican proteoglycan family
5q31	SIL1	AR	248800	Marinesco-Sjögren syndrome	resident endoplasmic reticulum N-linked glycoprotein
6p24	TFAP2A	AD	113620	branchiooculofacial syndrome	a transcription factor
6p23	GCM2	AD	146200	hypoparathyroidism familial isolated	a homolog of the Drosophila glial cells missing gene
6p21.3	NEU1	AR	256550	sialidosis type 2	a lysosomal enzyme that cleaves terminal sialic acid residues
6q21q23.2	GJA1	AD	164200	oculodentodigital dysplasia	a member of the connexin gene family
6q22q24	PEX7	AR	215100	rhizomelic chondrodysplasia punctata type 1	cytosolic receptor for the set of peroxisomal matrix enzymes
6q24.2	PEX3	AR	614882	Zellweger syndrome	involved in peroxisome biosynthesis and integrity
7p15.3	FAM126A	AR	610532	hypomyelinating leukodystrophy 5	part in the beta-catenin/Lef signaling pathway
7q21.2	PEX1	AR	214100	Zellweger syndrome	a member of the AAA ATPase family
7q31.1	CAV1	AD	606721	partial lipodystrophy, congenital cataracts, neurodegeneration syndrome	main component of the caveolae plasma membranes
7q34	AGK	AR	212350	Sengers syndrome	a mitochondrial membrane protein involved in lipid and glycerolipid metabolism
8p21.1	ESCO2	AR	268300	Roberts syndrome	acetyltransferase activity may be required for the establishment of sister chromatid cohesion
8q13.3	EYA1	AD	601653	branchiootorenal syndrome 1	a member of the eyes absent (EYA) family of proteins
8q21.1	PXMP3	AR	614866	Zellweger syndrome	an integral peroxisomal membrane protein required for peroxisome biogenesis
8q21q22	CNGB3	AR	262300	achromatopsia 3	the beta subunit of a cyclic nucleotide-gated ion channel
8q24.3	RECQL4	AR	268400	Rothmund Thompson syndrome	DNA helicase that belongs to the RecQ helicase family
9p13.13	GALT	AR	230400	galatosemia	galactose-1-phosphate uridyl transferase
9p24	VLDLR	AR	224050	cerebellar hypoplasia and mental retardation with or without quadrupedal locomotion 1	low-density lipoprotein receptor
9q31q33	FKTN	AR	253800	muscular dystrophy-dystroglycanopathy	a putative transmembrane protein localized to the cis-Golgi compartment
9q34	LMX1B	AD	161200	nail-patella syndrome	a member of LIM-homeodomain family of proteins
9q34.1	POMT1	AR	236670	muscular dystrophy-dystroglycanopathy A1	an O-mannosyltransferase
10q11.23	ERCC6	AR	214150/133540	cerebrooculofacioskeletal syndrome I/Cockayne syndrome type B	a DNA-binding protein that is important in transcription-coupled excision repair
10q23.31	PTEN	AD	158350	Cowden disease	a tumor suppressor
10q24.3	ALDH18A1	AD/AR	616603/219150	cutis laxa AD/cutis laxa AR	a member of the aldehyde dehydrogenase family
10q26	OAT	AR	258870	gyrate atrophy of choroid and retina	mitochondrial enzyme ornithine aminotransferase
11p15.3p15.1	PTH	AD/AR	146200	familial isolated hypoparathyroidism	a member of the parathyroid family of proteins
11q13.4	LRP5	AR, AD	601813/607634	exudative vitreoretinopathy 4/osteopetrosis, autosomal dominant 1	a transmembrane low-density lipoprotein receptor
11q13.2q13.5	DHCR7	AR	270400	Smith-Lemli-Opitz syndrome	an enzyme that removes the C(7–8) double bond in the B ring of sterols
11q13.4	CLPB	AR	616271	3-methylglutaconic aciduria with cataracts, neurologic involvement, neutropenia	member of the ATPases associated with diverse cellular activities (AAA+) superfamily
11q14.2	FZD4	AD	133780	retinopathy of prematurity	a member of the frizzled gene family
11q22.3	MMP1	AR	226600	epidermolysis bullosa dystrophic	a member of the peptidase M10 family of matrix metalloproteinases
11q22.1q23.2	CRYAB	AD	608810	myofibrillar myopathy	members of the small heat shock protein (HSP20) family
11q23.3	SC5DL	AR	607330	lathosterolosis	enzyme of cholesterol biosynthesis
11q25	JAM3	AR	613730	hemorrhagic destruction of brain, subependymal calcification	immunoglobulin superfamily gene member
12q13.12	TUBA1A	AD	611603	lissencephaly	microtubule constituents of to the tubulin superfamily
12p13.3	PEX5	AR	214110	Zellweger syndrome	protein essential for the assembly of functional peroxisomes
12q24	MVK	AR	610377	mevalonic aciduria	peroxisomal enzyme mevalonate kinase
13q12	GJB6	AD	129500	Clouston syndrome	one of the connexin proteins
13q12.3	B3GALTL	AR	261540	Peters-plus syndrome	a beta-1,3-glucosyltransferase that transfers glucose to O-linked fucosylglycans on thrombospondin type-1 repeats
13q14.3	ITM2B	AD	117300	cerebral amyloid angiopathy	a transmembrane protein
13q34	COL4A1	AD	607595	cerebral small vessel disease	a type IV collagen alpha protein
14q21.1	SEC23A	AR	607812	craniolenticulosutural dysplasia	a member of the SEC23 subfamily of the SEC23/SEC24 family
14q24	POMT2	AR	613150	muscular dystrophy-dystroglycanopathy	an O-mannosyltransferase
15q15	BUB1B	AR	257300	mosaic variegated aneuploidy syndrome 1	a kinase involved in spindle checkpoint function
15q21.1	FBN1	AD	608328	Weill-Marchesani syndrome	a member of the fibrillin family of proteins
15q25	POLG	AD/AR	157640	progressive external ophthalmoplegia with mitochondrial DNA deletions 1	the catalytic subunit of mitochondrial DNA polymerase
16p13.3p13.2	GFER	AR?	613076	progressive mitochondrial myopathy, sensorineural hearing loss, developmental delay	structural and functional homolog of the yeast scERV1 gene
16q21	NOD2	AD	186580	Blau syndrome	a member of the Nod1/Apaf-1 family
16q23	ADAMTS18	AR	267750	Knobloch syndrome	a member of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) protein family
16q22q23	MAF	AD	601088	Aymé-Gripp syndrome	a DNA-binding, leucine zipper-containing transcription factor
17p13.3	YWHAE	AD	247200	Miller-Dieker lissencephaly syndrome	member of the 14-3-3 family of proteins which mediate signal transduction by binding to phosphoserine-containing proteins
17q12	PEX12	AR	614859	Zellweger syndrome	member of the peroxin-12 family
17q21	WNT3	AR	273395	tetraamelia syndrome	secreted signaling proteins implicated in oncogenesis
17q21.33	XYLT2	AR	605822	spondyloocular syndrome	an isoform of xylosyltransferase, which belongs to a family of glycosyltransferases
18q12.3	EPG5	AR	242840	Vici syndrome (immunodeficiency, cleft lip/palate, cataract, hypopigmentation, absent corpus callosum)	a large coiled-coil domain-containing protein that functions in autophagy
18q23	CTDP1	AR	604168	congenital cataracts, facial dysmorphism, neuropathy	a protein which interacts with the carboxy-terminus of the RAP74 subunit of transcription initiation factor TFIIF
19q13.1	MAN2B1	AR	248500	alpha-mannosidosis	an enzyme that hydrolyzes terminal, nonreducing alpha-D-mannose residues in alpha-D-mannosides
19q13.3	DMPK	AD	160900	myotonic dystrophy 1	a serine-threonine kinase
19q13.32	FKRP	AR	613153	muscular dystrophy-dystroglycanopathy	a protein which is targeted to the medial Golgi apparatus and is necessary for posttranslational modification of dystroglycan
20p11.21q12	ABHD12	AR	612674	polyneuropathy, hearing loss, ataxia, retinitis pigmentosa, cataract	an enzyme that catalyzes the hydrolysis of 2-arachidonoyl glycerol
20q13.13q13.2	SALL4	AD	607323	Duane-radial ray syndrome	a zinc finger transcription factor thought to play a role in the development of abducens motor neurons
20q13.3	GNAS	AD	103580/612462/612463	pseudohypoparathyroidism type 1A/pseudohypoparathyroidism type 1C/pseudohypoparathyroidism	multiple transcript variants encoding different isoforms
21q22.3	COL18A1	AR	267750	Knobloch syndrome 1	alpha chain of type XVIII collagen
22q11.21	PEX26	AR	614872	Zellweger syndrome	member of the peroxin-26 gene family
22q12.2	NF2	AD	101000	neurofibromatosis type 2	a protein similar to ezrin, radixin, moesin family of proteins
22q12.3	LARGE	AR	613154	muscular dystrophy-dystroglycanopathy	a member of the N-acetylglucosaminyltransferase gene family
22q13.1	MYH9	AD	153640	Fechtner syndrome	a conventional non-muscle myosin
Xp11.4	BCOR	XL	300166	microphthalmia syncromic 2	an interacting corepressor of BCL6
Xp11.23	PQBP1	XL	309500	Renpenning syndrome	a nuclear polyglutamine-binding protein that is involved with transcription activation
Xp11.23p11.22	EBP	XL	302960	chondrodysplasia punctata 2	an integral membrane protein of the endoplasmic reticulum
Xp22.3	ARSE	XL	302950	chondrodysplasia punctata 1	a member of the sulfatase family
Xp22.3	HCCS	XL	309801	linear skin defects with multiple congenital anomalies	an enzyme that covalently links a heme group to the apoprotein of cytochrome c
Xp22	AIC	XL	304050	Aicardi syndrome	unknown
Xp22.13	NHS	XL	302350	Nance-Horan syndrome	a protein containing 4 conserved nuclear localization signals
Xq22	GLA	XL	301500	Fabry disease	a homodimeric glycoprotein that hydrolyzes the terminal alpha-galactosyl moieties from glycolipids and glycoproteins
Xq22	COL4A5	XL	301050	Alport syndrome	one of the 6 subunits of type IV collagen
Xq25q26.1	OCRL	XL	309000	Lowe oculocerebrorenal syndrome	an inositol polyphosphate 5-phosphatase
Xq28	IKBKG	XL	308300	incontinentia pigmenti	regulatory subunit of the inhibitor of kappaB kinase complex

AD, autosomal dominant; AR, autosomal recessive; XL, X linked.

Table 2.

Syndromic genes only with cataract

Locus	Gene	OMIM	Cataract inheritance	Syndrome inheritance	Syndrome	Reference of only cataract affection
4p16.1	WFS1	614296	AD	AD	Wolfram-like syndrome (no cataract is associated with this syndrome)	Berry et al., 2013
7q34	AGK	212350	AR	AR	Sengers syndrome	Aldahmesh et al., 2012
8q13.3	EYA1	601653	AD	AD	brachio-oto-renal syndrome-1, nystagmus	Azuma et al., 2000
11q22.1q23.2	CRYAB	608810	AR/AD	AD	myofibrillar myopathy (alpha-B crystallinopathy)	Berry et al., 2001; Liu et al., 2006; Devi et al., 2008; Chen et al., 2009; Safieh et al., 2009; Sun et al., 2011b; Xia et al., 2014b; Jiaox et al., 2015; Khan et al., 2015; Ma AS et al., 2016
13q34	COL4A1	607595	AD	AD	familial porencephaly, brain small vessel disease with hemorrhage/Axenfeld-Rieger anomalies, hereditary angiopathy with nephropathy, aneurysms, and muscle cramps	Xia et al., 2014a
15q21.1	FBN1	608328	AD	AD	Marfan syndrome, Weill-Marchesani syndrome	Li D et al., 2016
16q22q23	MAF	601088	AD	AD	Aymé-Gripp syndrome	Jamieson et al., 2002; Vanita et al., 2006; Hansen et al., 2007, 2009; Narumi et al., 2014; Sun et al., 2014; Ma AS et al., 2016
Xp22.13	NHS	302350	XL	XL	Nance-Horan syndrome	Ramprasad et al., 2005; Florijn et al., 2006; Sharma et al., 2008; Coccia et al., 2009; Sun et al., 2014; Li D et al., 2016; Ma AS et al., 2016

AD, autosomal dominant; AR, autosomal recessive; XL, X linked.

Table 3.

Only cataract genes

Locus	Gene	Inheritance	Cataract phenotype	Other phenotype	Gene product	Reference
1p36	EPHA2	AR/AD	nuclear, posterior, cortical, total, posterior polar, subcapsular and cortical, zonular	a few reports with microcornea or delay developmenta, b	ephrin receptor subfamily of the protein-tyrosine kinase family	Shiels et al., 2008; Zhang et al., 2009a; Kaul et al., 2010; Aldahmesh et al., 2012; Dave et al., 2013; Shentu et al., 2013; Gillespie et al., 2014; Reis et al., 2014; Sun et al., 2014; Bu et al., 2016; Li D et al., 2016
2q33q35	CRYGD	AD	punctate, progressive		account for approximately one-third of total lens proteins	Héon et al., 1999; Stephan et al., 1999; Kmoch et al., 2000; Santhiya et al., 2002; Nandrot et al., 2003; Burdon et al., 2004; Mackay et al., 2004; Shentu et al., 2004; Xu et al., 2004; Gu et al., 2005; Zenteno et al., 2005; Gu et al., 2006; Messina-Baas et al., 2006; Hansen et al., 2007; Plotnikova et al., 2007; Zhang et al., 2007; Devi et al., 2008; Li et al., 2008; Khan et al., 2009; Santana et al., 2009; Zhang et al., 2009b; Roshan et al., 2010; Wang et al., 2010; Sun et al., 2011b; Wang et al., 2011b; Vanita et al., 2012; Jia et al., 2013; Reis et al., 2013; Gillespie, et al., 2014; Mackay et al., 2014; Zhai et al., 2014; Zhuang et al., 2015; Gao et al., 2016; Ma AS et al., 2016; Yang et al., 2016
2q33q35	CRYGC	AD	central zonular pulverulent (Coppock-like)	some cases with microphthalmia and microcornea	constitute the major proteins of vertebrate eye lens	Héon et al., 1999; Ren et al., 2000; Santhiya et al., 2002; Gonzalez-Huerta et al., 2007; Devi et al., 2008; Yao et al., 2008; Zhang et al., 2009b; Kumar et al., 2011; Guo et al., 2012; Li et al., 2012; Kondo et al., 2013; Reis et al., 2013; Gillespie et al., 2014; Prokudin et al., 2014; Li D et al., 2016; Ma AS et al., 2016
2q33q35	CRYGB	AD	anterior polar and lamellar		account for approximately one-third of total lens proteins	AlFadhli et al., 2012
2q33q35	CRYGA	sporadic	Total		major protein components of the vertebrate eye lens	Li D et al., 2016
3p21.31	FYCO1	AR	posterior capsular, nuclea		plays a role in microtubule plus end-directed transport of autophagic vesicles	Chen et al., 2011; Aldahemesh et al., 2012; Gillespie et al., 2014; Khan et al., 2015
3q21q22	BFSP2	AD/AR	nuclear, pulverulent, cortical, Y-sutural, lamellar, opalescent, spok-like opacities, sutural		conforms the cytoskeletal element referred to as the beaded filament	Conley et al., 2000; Jakobs et al., 2000; Zhang et al., 2004; Zhang et al., 2006; Cui et al., 2007; Ma et al., 2008; Aldahmesh et al., 2011, 2012; Gillespie et al., 2014; Liu et al., 2014; Li D et al., 2016
3q25qter	CRYGS	AD	central pulverulent, opalescent, sutural, lamellar, Coppock		is expressed late abundantly in the ocular lens	Sun et al., 2005; Devi et al., 2008; Vanita et al., 2009; Sun et al., 2011a; Yang et al., 2013
5q22	WDR36	sporadic	posterior polar, nuclear		a member of the WD repeat protein family	Li D et al. 2016
6p24.2	GCNT2	AR	central, nuclear, polar	adult i blood group	enzyme responsible for formation of the blood group I antigen	Yu et al., 2001; Inaba et al., 2003; Pras et al., 2004; Wussuki-Lior et al., 2011; Aldahmesh et al., 2012; Borck et al., 2012; Happ et al., 2016
6p21.31	LEMD2	AR	Hutterite type		a transmembrane protein of the inner nuclear membrane that is involved in nuclear structure organization	Boone et al., 2015
9p22.1	RRAGA	AD	nuclear		a Ras-related GTP-binding A	Chen et al., 2016
9q13q22	CTPL1	AR	progressive pulverulent		unknown	Héon et al., 2001; Forshew et al., 2005
9q22.33	TDRD7	AR	polar		a member of the Tudor family of proteins	Lachke et al., 2011; Li D et al., 2016
10p13	VIM	AD	pulverulent		a member of the intermediate filament family	Müller et al., 2009; Ma AS et al., 2016
12q13	MIP	AD	lamellar, sutural, cerulean, punctate, nuclear, pulverulent, snail-like, cortical, Y-sutural		a member of the water-transporting aquaporins as well as the original member of the MIP family of channel proteins	Berry et al., 2000; Geyer et al., 2006; Gu et al., 2007; Lin et al., 2007; Jiang et al., 2009; Wang et al., 2010; Wang et al. 2011a; Xiao et al., 2011; Yang et al., 2011b; Reis et al., 2013; Senthil Kumar et al., 2013; Zeng et al., 2013; Ding et al., 2014; Sun et al., 2014; Yu et al., 2014; Shentu et al., 2015; Song et al., 2015; Ma AS et al., 2016; Qin et al., 2016
13q11q12	GJA3	AD	nuclear, pulverulent, Y-sutural, lamellar, lamerllar sutural, pearl box, coralliform, punctate, Coppock-like		gap junction protein alpha 3	Mackay et al., 1999; Rees et al., 2000; Jiang et al., 2003; Bennett et al., 2004; Burdon et al., 2004; Li et al., 2004; Devi et al., 2005; Ma et al., 2005; Addison et al., 2006; Hansen et al., 2006, 2009; Guleria et al., 2007; Santhiya et al., 2010; Zhou Z et al., 2010; Bennett and Shiels, 2011; Ding et al., 2011; Sun et al., 2011a; Yang et al., 2011a; Yao et al., 2011b; Wang and Zhu, 2012; Zhang et al., 2012; Guo et al., 2013; Ponnam et al., 2013; Zhou et al., 2013; Gillespie et al., 2014; Hu et al., 2014; Yang Z et al., 2015; Yuan et al., 2015; Li B et al., 2016; Ma AS et al., 2016; Ma MF et al., 2016; Wang et al., 2016
16p13.2	TMEM114	AD			a glycosylated transmembrane protein	Jamieson et al., 2007
16q21	HSF4	AD/AR	cortical lamellar, zonular, anterior polar, nuclear, sutural		heat-shock transcription factor	Bu et al., 2002; Smaoui et al., 2004; Forshew et al., 2005; Ke et al., 2006; Sajjad et al., 2008; Hansen et al., 2009; Gillespie et al., 2014; Lv et al., 2014; Liu et al., 2015; Behnam et al., 2016; Li D et al., 2016
17q11.2q12	CRYBA1	AD/AR	zonular sutural, pulverulent, sutural nuclear, lamellar, floriform, Y-sutural, nuclear, polymorphic		crystallin structure proteins	Kannabiran et al., 1998; Bateman et al., 2000; Burdon et al., 2004; Ferrini et al., 2004; Reddy et al., 2004; Qi et al., 2004; Lu et al., 2007; Devi et al., 2008; Gu et al., 2010; Zhu et al., 2010; Sun et al., 2011b; Yang Z et al., 2011; Yang et al., 2012; Yu et al., 2012; Gillespie et al., 2014; Zhang et al., 2014; Khan et al., 2015
17q12	UNC45B	AD	posterior subcapsular and central		a co-chaperone required for folding and accumulation of type II myosins	Hansen et al., 2014
17q24	GALK1	AR		galactokinase deficiency	galactokinase 1	Stambolian et al., 1995; Asada et al., 1999; Kalaydjieva et al., 1999; Kolosha et al., 2000; Hunter et al., 2001; Reich et al., 2002; Yasmeen et al., 2010; Singh et al., 2012; Chacon-Camacho et al., 2014; Gillespie et al., 2014
19p13.2	LONP1	AR	nuclear		a mitochondrial matrix protein that belongs to the Lon family of ATP-dependent proteases	Khan et al., 2015
19q13.13	WDR87	AR	total white		WD repeat domain 87	Khan et al., 2015
19q13.2	PRX	AD	nuclear		a protein involved in peripheral nerve myelin upkeep	Yuan et al., 2016
19q13.33	FTL	AD	iron flecks, nuclear, Y-sutural, stellate, sunflower, pulverulent, crystallin, breadcrumb-like nuclear, cortical opacities	hyperferritinemia	the light subunit of the ferritin protein	http://www.hgmd.cf.ac.uk/ac/gene.php?gene=FTL
19q13.4	LIM2	AR	congenital	Galactokinase deficiency	an eye lens-specific protein found at the junctions of lens fiber cells	Ponnam et al., 2008
19q33	SIPA1L3	AR			the signal-induced proliferation associated 1 family of genes	Greenlees et al., 2015
20p11.23p12.1	BFSP1	AR	cortical progressive, juvenile onset, nuclear		a lens-specific intermediate filament-like protein named filensin	Ramachandran et al., 2007; Wang et al., 2013; Li D et al., 2016; Ma AS et al., 2016
20q11.22	CHMP4B	AD	posterior subcapsular		a member of the chromatin-modifying protein/charged multivesicular body protein family	Shiels et al., 2007
21q22.3	LSS	AR	Total	?	catalyzes the conversion of (S)-2,3-oxidosqualene to lanosterol	Zhao et al., 2015
22q11.23	CRYBB3	AR/AD	nuclear, cortical		crystallin protein	Riazuddin et al., 2005; Hansen et al., 2009; Reis et al., 2013; Li D et al., 2016; Ma AS et al., 2016
22q11.23	CRYBB2	AD	subcapsular, zonular, nuclear, coronary, lamellar, sutural, cerulean, pulverulent, polymorphic, cortical		crystallin protein	Litt et al., 1997; Gill et al., 2000; Vanita et al., 2001; Santhiya et al., 2004; Yao et al., 2005; Bateman et al., 2007; Pauli et al., 2007; Devi et al., 2008; Li et al., 2008; Hansen et al., 2009; Lou et al., 2009; Mothobi et al., 2009; Wang et al., 2009; Santhiya et al., 2010; Wang et al., 2011b; Yao et al., 2011a; Weisschuh et al., 2012; Chen et al., 2013; Faletra et al., 2013; Garnai et al., 2014; Gillespie et al., 2014; Sun et al., 2014; Ma AS et al., 2016; Messina-Baas et al., 2016
22q12.1	CRYBB1	AR/AD	nuclear		crystallin protein	Willoughby et al., 2005; Cohen et al., 2007; Wang et al., 2007; Yang et al., 2008; Meyer et al., 2009; Wang et al., 2011b; Khan et al., 2012; Reis et al., 2013; Wu et al., 2013; Ma AS et al., 2016

For abbreviations, see Table 2.

^aGillespie et al., 2014.

^bAldahmesh et al., 2012.

Table 4.

Cataract genes and eye anomalies

Locus	Gene	Inheritance	Cataract phenotype	Other phenotype	Gene product	Reference
1p32	FOXE3	AR/AD	cerulean, congenital	microphthalmia, sclerocornea, aphakia, optic disc coloboma, aniridia plus, glaucoma, viteoretinal dysplasia (almost all reports)	the forkhead family of transcription factors	Semina et al., 2001; Ormestad et al., 2002; Valleix et al., 2006; Iseri et al., 2009; Ali et al., 2010; Anjum et al., 2010; Bremond-Gignac et al., 2010; Reis et al., 2010; Doucette et al., 2011; Gillespie et al., 2014; Li D et al., 2016
1q21.1	GJA8	AD/AR	nuclear, punctiform, pulverulent, jellyfish-like, star-shaped, full moon, Y-sutural, balloon-like, lamelar, zonular, nuclear, triangular, perinuclear	microcornea, glaucoma (about half of the reports)	a transmembrane connexin protein	Shiels et al., 1998; Berry et al., 1999; Polyakov et al., 2001; Willoughby et al., 2003; Zheng et al., 2005; Arora et al., 2006, 2008; Devi and Vijayalakshmi, 2006; Vanita et al., 2006; Hansen et al., 2007; Ponnam et al., 2007; Lin et al., 2008; Schmidt et al., 2008; Vanita et al., 2008; Yan et al., 2008; Graw et al., 2009; Hansen et al., 2009; Wang et al., 2009; Gao et al., 2010; Hu et al., 2010; He et al., 2011; Kumar at al., 2011; Sun et al., 2011a; Wang L et al., 2011; Li et al., 2013; Ponnam et al., 2013; Reis et al., 2013; Su et al., 2013; Chen et al., 2014; Ge et al., 2014; Gillespie et al., 2014; Mackay et al., 2014; Prokudin et al., 2014; Sun et al., 2014; Zhu et al., 2014; Chen et al., 2015; Liang et al., 2015; Yang Z et al., 2015; Ma AS et al., 2016; Min et al., 2016; Yu et al., 2016
9q21.12	TRPM3	AD		open-angle glaucoma	family of transient receptor potential channels	Bennett et al., 2014
11p13	PAX6	AD		aniridia “plus” (iris and foveal hypoplasia, nystagmus, cataract, corneal abnormalities, glaucoma)	a homeobox and paired domain-containing protein that binds DNA	LOVD PAX6 Homepage
11q13	BEST1	AD	pulverulent-like	vitreoretinochoroidopathy, microcornea, rod-cone dystrophy, cataract, posterior staphyloma	a member of the bestrophin gene family	Boon et al., 2009
14q24.3	VSX2	AR		microphthalmia, anophthalmia, iris coloboma	a homeobox protein originally described as a retina-specific transcription factor	Ferda Percin et al., 2000
15q22.32	NR2E3	AD, AR		retinitis pigmentosa-37, enhanced S-cone syndrome	a family of nuclear receptor transcription factors	Edwards et al., 2008
15q25.1	MIR184	AD	congenital, early-onset anterior polar	endothelial dystrophy, iris hypoplasia, cataract and stromal thining (EDICT) syndrome, keratoconus. Corneal anomalies	microRNAs involved in post-transcriptional regulation of gene expression	Iliff et al., 2012; Bykhovskaya et al., 2015
17p13.1	GUCY2D	AR		Leber congenital amaurosis-1	a retina-specific guanylate cyclase	Gradstein et al., 2016
21q22.3	CRYAA	AR/AD	nuclear, laminar, posterior polar, punctate, spike-like, ring-like, fan-shaped, disc-like, Y-sutural, zonular, lamellar, polymorphic, perinuclear	microcornea, iris coloboma, axial elongation, macular hypoplasia, corneal opacity	chaperone-like small heat-shock protein family	Litt et al., 1998; Pras et al., 2000; Mackay et al., 2003; Graw et al., 2006; Santhiya et al., 2006; Vanita et al., 2006; Beby et al., 2007; Hansen et al., 2007; Khan et al., 2007; Devi et al., 2008; Gu et al., 2008; Richter et al., 2008; Hansen et al., 2009; Santana et al., 2009; Zhang et al., 2009a; Li et al., 2010; Sun et al., 2011a; Su et al., 2012; Wang and Zhu, 2012; Kondo et al., 2013; Laurie et al., 2013; Reis et al., 2013; Yang et al., 2013; Kong et al., 2015; Liang et al., 2015; Javadiyan et al., 2016; Li D et al., 2016; Ma AS et al., 2016
22q12.1	CRYBA4	AD	congenital nuclear, lamellar	microcornea, microphthalmia	soluble proteins in the human lens	Billingsley et al., 2006; Zhou G et al., 2010

For abbreviations, see Table 2.

Embryological Development of the Lens

Briefly, from mouse studies, the lens development starts at day 22 of gestation (4 mm embryonic stage) from the surface ectoderm. Pax6 and Sox2 genes, in the optic vesicle, induce the surface ectoderm to form the lens placode, which invaginates and forms the lens vesicle [Kamachi et al., 2001; Kondoh et al., 2004]. By day 40 of gestation (10 mm stage), the lens vesicle is completely separated from the surface ectoderm. Group B1 SOX proteins (SOX1, SOX2, and SOX3) activate γ-crystallin genes in the mouse lens, producing the crystallins, water-soluble proteins that comprise over 90% of the proteins of the lens [Hoehenwarter et al., 2006]. Elongating cells of the posterior end of the lens vesicle form the primary fibers, which become the embryonic nucleus in the mature lens. In the final phase of lens differentiation, several fibroblast growth factors seem to be required [McAvoy and Chamberlain, 1989; Lovicu and Overbeek, 1998]. The lens continues growing after birth, with the new secondary fibers generated from the equatorial cells of the lens epithelium (germinative zone). The lens epithelial cells synthesize crystallins and lose their nuclei to become mature lens fibers. Disruption of this delicate process due to toxic, metabolic, infectious, or genetic agents results in CC.

Syndromic and Nonsyndromic Cataracts

A rough division of hereditary cataracts comprises 2 parts: syndromic and nonsyndromic cataracts. If only the lens is implicated in the pathological process, it is defined as nonsyndromic cataracts. If there are more organs implicated, it results in syndromic cataracts. Around 100 genes throughout the genome are in the OMIM database explaining the different syndromes caused by different genes, either associated with cataracts or as part of them (Table 1). Nevertheless, genetically it is sometimes difficult to establish this difference because clinically one gene defect can result in either lens affection or systemic disease. Some examples of this variance (Table 2) are found in the affected AGK gene, which results in either an isolated cataract or in Senger syndrome (OMIM 212350) [Aldahmesh et al., 2012]; the MAF gene, in which a molecular defect can result in cataracts or in Aymé-Gripp syndrome (OMIM 601088) [Narumi et al., 2014; Sun et al., 2014; Ma AS et al., 2016], or in the NHS gene alteration which can lead to isolated cataracts or Nance-Horan syndrome (OMIM 302350) [Coccia et al., 2009; Li D et al., 2016]. The scope of this review does not include syndromic cataracts. It is only briefly referenced because some genes (WFS1, AGK, EYA1, CRYAB, COL4A1, FBN1, MAF, and NHS) are also described in previous reports with CC and no other anomalies (Table 2).

Nonsyndromic Cataracts

Around 35 genes have been strongly associated with inherited CC only and with no other systemic anomalies; they show autosomal dominant and autosomal recessive inheritance patterns (Table 3). However, in some of these cases (EPHA2, BFSP2, HSF4, CRYBA1, CRYBB1, and CRYBB3), hereditary CC can present both patterns of autosomal inheritance (dominant or recessive). A great variety of functions is described in all affected genes that produce nonsyndromic inherited CC. Protein functions encompass structural to enzymatic processes including channels, phagosomes, RNA processing, transcription factors, and regulation of intracellular signaling (Table 3).

The cataract phenotype by itself is not an efficient indicator of the affected gene or mutation, since identical cataracts can result from mutations at different loci and may have several inheritance patterns. Conversely, various cataract types can be found in the same gene affection, presenting clinical heterogeneity. In addition, some nonsyndromic cataracts could include the presence of lens opacities only or lens opacities with ocular abnormalities and with no systemic affection, i.e., microcornea, aniridia, vitreoretinochoroidopathy, microphthalmia, anophthalmia, iris hypoplasia, or rod-cone dystrophy (Table 4).

Crystallins

Crystallins are divided in α, β, and γ and constitute around 90% of the soluble proteins of the lens with an asymmetric and biphasic distribution [Augusteyn, 2010]. α-crystallins (αA-crystallin and αB-crystallin) belong to the chaperone-like small heat shock protein family (sHSPs). They block the formation of stress protein aggregates to avoid toxic effects [Clark et al., 2012]. The sHSPs are large, polydisperse, and do not crystallize, thus playing an important role in the lens. It is likely that the high refractive index of a lens with no compromise of transparency is due to the ability to form extended lattices of the water-insoluble α-crystallin [Slingsby et al., 2013]. Outside of the lens, α-crystallin is expressed principally in the retina, the muscle, and the brain. In the retina, it seems to be implicated in cytoprotection, cell survival, inflammation, and autophagy [Kannan et al., 2012]. A great diversity of mutations associated with cataracts has been reported in the CRYAA and CRYAB genes, whose products are αA-crystallin and αB-crystallin, respectively. In approximately half of the cases, CRYAA gene defects are associated with eye anomalies, whereas CRYAB gene defects are associated with myofibrillar myopathy (Tables 1, 4); in both cases, the rest of the reports connect lens opacities with great clinical heterogeneity (Table 4). Most cases show an autosomal dominant pattern, and in a few cases, a recessive autosomal inheritance.

The β- and γ-crystallins consist of 4 similarly folded Greek key motifs organized into 2 domains. The Greek key motif is a structure of 4 β-sheets, folded in an antiparallel form that generates a folded compact domain [Richardson, 1977]. The values and gradients in the refractive index of lens proteins seem to be related to the Greek key motif [Zhao et al., 2011]. The central portion and the embryonic nuclear region of the lens are rich in β- and γ-crystallins. In humans, γ-crystallins comprise from γA to γF. Now, γS-crystallin (βS-crystallin) is also included in this group; γE and γF are pseudogenes. Interestingly, CRYBA, CRYBB, CRYGA, CRYGB, CRYGC, CRYGD, and CRYGS gene mutations are only associated with inherited CC and no other systemic or eye anomalies (Table 3). In Figure 2, the normal CYGD crystallin structure is shown.

Fig. 2.

The structure of CRYGD, from https://swissmodel.expasy.org/.

EPH2

The EPHA2 (ephrin receptor A2) gene encodes a transmembrane tyrosine kinase receptor (epithelial cell) which contains an extracellular ligand-binding domain and is expressed in the human lens [Pasquale, 2010]. EPH2, with its ligands ephrin-A1 or ephrin-A5, has a role in cell adhesion and cell repulsion [Miao and Wang, 2009]. Inhibition of EPHA2 induces apoptosis and abrogates tumorigenic growth of tumor cells [Amato et al., 2014]. Apparently, the downstream signaling of activated EPHA2 promotes the antioxidative capacity of lens epithelial cells to eradicate the overproduction of reactive oxygen species [Yang J et al., 2015]. It is probable that the loss of EPHA2 function could affect the structural stability of the cell, cell-to-cell crosstalk, protein folding and transcriptional activation [Park et al., 2013]. Thus, the cytoprotective and antiapoptotic functions of EPHA2 in the lens indicate the possible role of EPHA2 in avoiding lens opacity. Practically, all mutations reported in the EPH2 gene are associated exclusively with inherited CC and no other anomalies (some cataracts are age related, as well). Only one study reports microcornea with mild dysmorphic features [Gillespie et al., 2014]. Both autosomal recessive and dominant patterns as well as clinical heterogeneity are present (Table 3). Figure 3 shows the structure of the EPH2 protein.

Fig. 3.

The structure of EPHA2, from https://swissmodel.expasy.org/.

GJA3 and GJA8

The GJA3 (gap junction alpha-3) gene encodes connexin 46, whereas the GJA8 (gap junction alpha-8) gene encodes connexin 50, gap junction channel proteins that play an important role in lens cell homeostasis. Connexin proteins have 4 transmembrane domains, 3 intracellular regions, and 2 extracellular loops. Six connexin subunits form 1 connexon. Connexin 46 and connexin 50 are expressed in the lens fiber cells and are essential for the coupling of mature fibers in the central core of the lens [Gong et al., 1997; Martinez-Wittinghan et al., 2004]. Connexins allow the interchange of ions and low-molecular-weight molecules between contiguous cells. Connexin 46 and connexin 50 are the major components of human lens fiber cells, and their molecular defects result in about 20% of nonsyndromic cataract reports [Shiels et al., 2010]. GJA3 gene mutations produce only cataracts with no other manifestations, unlike GJA8 that includes the presence of microcornea in half of the reports (Tables 3, 4). The structure of connexin 46 is shown in Figure 4.

Fig. 4.

The structure of GJA3, from https://swissmodel.expasy.org/.

MIP

The MIP (major intrinsic protein) gene encodes aquaporin 0 (AQP0), the most abundant protein in the fiber cell membrane that functions as a water channel and belongs to the superfamily of AQPs [Shiels et al., 2001]. AQPs create a microcirculation in the avascular lens to nourish central fiber cells, maintaining their transparency and homeostasis [Gao et al., 2013]. AQP0 accounts for about 45% of the total plasma proteins [Bassnett et al., 2009]. Homozygous or heterozygous loss of AQP0 will reduce the water permeability of lens fiber cells producing lens opacities in mice [Shiels et al., 2001]. Apart from the intermolecular contacts between AQP0 monomers, AQP0 interacts with other proteins in lens fiber cells as crystallins and connexins [Liu and Liang 2008; Liu et al. 2011]. Mutations in the MIP gene result only in cataracts with no other manifestations, and all of them are inherited in an autosomal dominant pattern (Table 3). The structure of the AQP0 channel is shown in Figure 5.

Fig. 5.

The structure of MIP, from https://swissmodel.expasy.org/.

FYCO1

FYCO (FYVE and coiled-coil domain containing 1), a PI(3)P-, Rab7-, and LC3-binding protein, mediates microtubule plus end-directed vesicle transport of autophagosomes, a required process for autolysosome formation [Pankiv et al., 2010]. FYCO1 is expressed in the lens epithelium and fiber cells (principally in nuclear fibers) in newborn mice, suggesting that autophagy is important in lens fiber cell differentiation [Brennan et al., 2012]. When FYCO1 is lacking, phagosomes stay p40phox⁺ longer and produce more reactive oxygen. This represents FYCO1's participation in the immunity role of regulating the phagosome maturation process and production of reactive oxygen, processes necessary for handling extracellular pathogens [Ma et al., 2014]. All mutations reported in the FYCO1 gene are inherited in an autosomal recessive pattern and have no other systemic or eye anomalies (Table 3). Figure 6 shows the structure of the FYCO1 protein.

Fig. 6.

The structure of FYCO1, from https://swissmodel.expasy.org/.

BFSP1 and BFSP2

BFSP1 (filesin, CP115) and BFSP2 (phakinin, CP49), lens-specific proteins, are the principal components of beaded filaments, which are unique cytoskeletal lens structures [Alizadeh et al., 2003]. BFSP1 and BFSP2 genes (beaded filament structural protein 1 and 2) encode these proteins. Apparently, the beaded filament is required to maintain cell morphology, 3-dimensional membrane architecture and lens transparency during fetal development and fiber cell differentiation [Alizadeh et al., 2002]. Both genes are associated with cataracts with no other anomalies: the BFS1 gene inherited in an autosomal recessive pattern and the BFS2 gene inherited in autosomal dominant and recessive patterns. Figure 7 shows the BFSP1 structure.

Fig. 7.

The structure of BFSP1, from https://swissmodel.expasy.org/.

Nonsyndromic Cataracts with Eye Anomalies

In some cases, nonsyndromic cataracts are associated with eye anomalies; about 15 genes have been reported in the literature with this condition (Tables 3, 4), but only a few genes have consistently presented this association (BEST1, VSX2, NR2E3, MIR184, and GUCY2D). The rest of the genes (CRYGB, CRYGA, FYCO1, BFSP2, CRYGS, WDR36, GCNT2, RRAGA, CTPL1, TDRD7, VIM, MIP, GJA3, TMEM114, HSF4, CRYBA1, UNC45B, LONP1, SIPA1L3, WDR87, LIM2, BFSP1, CHMP4B, and LSS) have also been reported with inherited CC exclusively. The principal eye anomalies correspond to microcornea, aniridia, microphthalmia, or vitreoretinal dysplasia.

IRE, GALK1 and GCNT2

The hyperferritinaemia-cataract syndrome (OMIM 600886), an autosomal dominant condition, presents high serum ferritin levels and bilateral CCs. Mutations in the I-ferritin gene that encodes the iron-responsive element (IRE) results in hyperferritinaemia-cataract syndrome (http://www.hgmd.cf.ac.uk/ac/gene.php?gene=FTL). The IRE in the ferritin mRNAs is a structure in the 5′-untranslated region. The increase of iron diminishes the binding of trans-acting iron-regulatory proteins with the ferritin mRNA structure, resulting in increased ferritin mRNA translation. Mutations in the IRE block this IRE-mediated regulation of ferritin production with the subsequent hyperferritinemia [Brooks et al., 2009]; this gene defect produces CC. Galactokinase 1 converts galactose into galactose-1-phosphate, when galactokinase deficiency is present, the accumulating galactose is converted to galactitol by aldose reductase and the cataractogenesis is observed. In this case, only CC with no other manifestations is present (Table 3). The human blood i and I antigens are linear and branched repeats of N-acetyllactosamine, respectively. The I-branching beta-1,6-N-acetylglucosaminyltransferase enzyme catalyzes the conversion of the i to the I antigen. Null phenotype of I (the adult i phenotype) is associated with CC. The gene defect in the I locus leads to the formation of CC with no other clinical manifestations (Table 3).

Diagnosis of Inherited Cataracts

As previously described, hereditary CC is a difficult condition to diagnose genetically due to more than 100 genes that have been associated with it. This task is even more difficult if no specific inherited pattern is observed in the pedigree. When systemic anomalies are present in the clinical symptomatology, syndromic cataracts are a possible diagnosis. The presence of cataracts with no other alterations reduces the genes involved to 35. Fortunately, the new molecular tools are very useful in identifying the gene affection with the support of clinical data. Next-generation sequencing (NGS) certainly represents an excellent tool for analyzing inherited cataracts. Some physicians believe the technique should not be used in cases of monogenic diseases due to high costs and the fact that it generates volumes of data which can be difficult to interpret. The analysis would be easier to interpret if the potentially affected gene is preselected. NGS enables identifying mutations throughout the human genome by facilitating the genetic diagnosis of hereditary diseases, of course including CCs, and has opened a new pathway in genetic diagnosis.

Disclosure Statement

The authors report no conflicts of interest.