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Learning objectives

Upon completion of this activity, participants should be able to:

1. List the major glomerular basement membrane components.
2. Describe the most common clinical presentations of Alport syndrome.
3. Describe the genetic patterns associated with Alport syndrome.
4. Identify the pathophysiologic basis of Alport syndrome.
5. Describe diagnostic tests for Alport syndrome.

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Alport syndrome

Alport syndrome is characterized by the familial occurrence of progressive hematuric nephritis and hearing loss.²³ The precise prevalence of the disease in the general population has not been established, but it is clearly lower than that in the selected Utah population studied by Atkin et al. (1 in 5,000 individuals).²⁴ Depending on the population studied, Alport syndrome affects 0.3–2.3% of all patients who develop end-stage renal disease (ESRD) in Europe, India or the US.

Hematuria—either macroscopic or microscopic—is the principal and constant feature of Alport syndrome.^{24, 25} It is an early-onset symptom, usually detected in childhood.^{26, 27, 28} Episodes of macroscopic hematuria precipitated by exercise or upper respiratory tract infection are observed in about 60% of patients before the age of 15 years, but are exceptional in adults. Hematuria is initially isolated; proteinuria increasing with age and progressive renal failure are observed, depending on the sex of the patient and the mode of transmission of the disorder. Hypertension does not usually develop before the onset of chronic renal insufficiency. Bilateral sensorineural hearing loss affecting high and middle frequencies is never congenital in Alport syndrome, but it can be detected during the first decade of life.^{26, 27, 28} In childhood, this hearing loss is usually progressive. Anterior lenticonus is a conical protrusion of the anterior aspect of the lens that develops progressively.²⁹ Retinal changes are characterized by the progressive appearance of asymptomatic perimacular yellowish flecks.^{30, 31} Both types of ocular lesion are specific to Alport syndrome, and are observed in about one-third of patients. Detecting such lesions can, therefore, be useful for diagnosis of the disorder. Nonspecific lesions of the cornea have also been reported in patients with Alport syndrome.³²

Ultrastructural studies have shown that the fundamental lesion of Alport syndrome involves the GBM (Figure 3).^{33, 34, 35} Characterized by thickening of the GBM (up to 800–1,200 nm) with splitting and fragmentation of the lamina densa into several strands, a 'basket weave' pattern is formed with irregular inner and outer contours. The lesion is often widespread, but it can be patchy, alternating with segments of normal or reduced thickness. In children, thinning of the GBM (to 100–200 nm) is the prevalent change and the most striking feature is the irregular appearance of the GBM that results from alternation between very thick and extremely thin segments.³⁶ Focal ruptures of the GBM repaired by newly formed basement membrane material might be observed.²⁶ Diffuse attenuation of the GBM is the only pathological finding in 10–20% of patients with Alport syndrome, regardless of age, indicating that a thin GBM is not invariably associated with benign disease.³⁷ Changes observed using light microscopy are not specific; during the early stages of the disease kidney tissue appears normal or shows minimal glomerular changes, and red blood cell casts are occasionally present. Thereafter, focal and segmental thickening of the capillary walls becomes visible, and segmental lesions of the tuft develop in an increasing number of glomeruli. These changes are associated with nonspecific tubulointerstitial lesions including foci of lipid-laden foam cells. Conventional immunofluorescence detects nothing or only faint or focal deposits of IgG, IgM or complement C3.

Figure 3 Electron microscopy images of renal tissue from patients with Alport syndrome.

(A) Thickening and splitting of the glomerular basement membrane (GBM) in an 11-year-old patient. The inner and outer contours of the GBM are 'festooned'. Magnification 30,000. (B) In tissue from a 12-year-old patient observed under low magnification, the irregular thickness of the GBM is evident. Magnification 5,400. (C) Diffuse thinning of the GBM in a 3-year-old male patient. Magnification 6,200. All sections stained with uranyl acetate and lead citrate.

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The genetics and the underlying causes of Alport syndrome remained elusive until the identification of the COL4A5 gene encoding the 5 type IV collagen chain, and detection of the first COL4A5 mutations in the most-frequent, X-linked, form of the disease.^{7, 8, 38} The genes encoding the 3(IV) and 4(IV) chains were then cloned, leading to identification of the first mutations in COL4A3 and COL4A4 in autosomal Alport syndrome.^{5, 6, 39} After many years of investigation, these major breakthroughs showed that Alport syndrome was a disorder of type IV collagen, and was heterogeneous at the genetic level. Since these discoveries, the development of specific antibodies against the different (IV) chains, and DNA analysis, have improved diagnostic approaches to the disease.

X-linked dominant Alport syndrome

In about 85% of affected families of European origin, Alport syndrome is transmitted as an X-linked dominant trait. This mode of transmission is characterized by greater disease severity in males than in females and the absence of father-to-son transmission.

Alport syndrome is severe in hemizygous male patients, with persistent hematuria, constant progression to ESRD and a high incidence of deafness, and with anterior lenticonus affecting one-third of patients.^{37, 40} Two forms of the disease have been distinguished on the basis of the rate of progression: a 'juvenile' type, characterized by a highly stereotypical course within each affected family and by the occurrence of ESRD in men at about 20 years of age; and a 'nonprogressive' or 'adult' type in which ESRD develops at approximately 40 years of age and the disease course is much more variable.²⁴

In heterozygous female patients, hematuria can be intermittent or detected only during adulthood. The few patients in whom hematuria never develops are asymptomatic carriers.⁴¹ Proteinuria is usually mild or absent in heterozygous females, and most patients never progress to ESRD. Gross hematuria in childhood, progressive worsening of proteinuria, hearing loss (which is progressive in some women) and diffuse GBM thickening are associated with poor outcome. This type of progression is impossible to predict on the basis of family history. The risk of developing ESRD before the age of 40 years is 12% in girls and women versus 90% in boys and men; the risk of progression in women seems to increase after the age of 60 years.⁴¹ Random X-chromosome inactivation might account for the variable clinical course of the disease in female patients.

Since the identification of COL4A5,^{7, 8} more than 300 mutations have been reported in this gene. These have been de novo in 10–15% of patients. Mutations occur throughout the gene and each affected family harbors its own mutation.^{37, 38, 40, 42, 43, 44, 45, 46, 47, 48, 49, 50} Gross changes in COL4A5, including deletions of various magnitudes and locations, insertions, duplications or complex rearrangements, have been identified in 5–15% of affected families.^{37, 40, 42, 43, 44, 45} Single base changes resulting in amino acid substitutions account for about 40% of small mutations.^{37, 40, 46, 47, 48, 49, 50} Most are missense mutations in glycine codons that disrupt the Gly–X–Y repeats in the collagenous domain of COL4A5. Other missense mutations affect conserved amino acid residues, especially cysteine, in the noncollagenous domain of the 5(IV) chain. Nonsense mutations, small deletions or insertions and splice-site mutations, leading to frameshift and premature stop codons, result in absent or truncated proteins.

Phenotype–genotype correlations have been established for males with X-linked dominant Alport syndrome.^{37, 40} Large rearrangements, and all mutations that change the reading frame of the gene, are associated with juvenile type Alport syndrome, early-onset hearing defects, and a high incidence of lenticonus. The 50% renal survival rate is 20 years. By contrast, missense mutations (mostly glycine substitutions) can cause either juvenile type or adult type Alport syndrome, sometimes without deafness. Affected male patients have a 50% renal survival rate of 32 years. The effect of glycine substitutions on the phenotype seems to depend on the distance of the mutation from the noncollagenous domain, with phenotypic effects being less severe when mutations involve exons 1–20.⁴⁰ In patients with splice-site mutations, the severity of renal disease is intermediate. A new classification system based on mutation type has been proposed as a replacement for the distinction between juvenile and adult type Alport syndrome.⁴⁰ In girls and women, no distinct correlation has been observed between phenotype and genotype or severity of disease in related boys and men.⁴¹ Early prediction of prognosis in carriers of Alport syndrome is not possible.

Diffuse leiomyomatosis is associated with Alport syndrome in 2–5% of families with the juvenile form of the disease. The leiomyomatosis affects the esophagus, the tracheobronchial tree and the female genital tract. It is completely penetrant and fully expressed, even in female patients with mild renal disease. Development of diffuse leiomyomatosis is linked to the presence of large deletions that remove the 5' ends of both COL4A5 and COL4A6, which are contiguous genes.^{51, 52, 53} COL4A6 deletions are always limited to exons 1, 1', and 2; patients with larger COL4A6 deletions do not develop tumors.⁵³ The rare co-occurrence of Alport syndrome (A), mental retardation (M), midface hypoplasia (M), and elliptocytosis (E) is also a contiguous gene deletion syndrome (AMME).⁵⁴

Immunohistological analysis of the distribution of the different chains of type IV collagen in the basement membranes where COL4A5 is normally expressed is of the utmost importance in the diagnosis of Alport syndrome and the recognition of X-linked transmission. In most patients, the 5(IV) chain defect (absence or abnormal structure as a result of mutation) impairs protomer assembly and the formation of normal collagen IV networks.^{11, 15, 36, 37, 41, 55, 56, 57}

Within the kidney, abnormal distribution of the 5(IV) chain is observed in approximately two-thirds of patients with X-linked Alport syndrome: the 5(IV) antigen is absent from the glomerular, capsular and distal tubular basement membranes in male patients, and has a discontinuous distribution in related female patients (Figure 4). This abnormal distribution is associated with absence of the 3(IV) and 4(IV) chains that normally participate in the formation of the 345(IV)–345(IV) network. By contrast, the 1(IV) and 2(IV) chains, which are normally confined to the mesangium and the subendothelial aspect of the GBM, are present throughout the entire width of the GBM. This distribution is reminiscent of that seen in fetal kidneys. No marked changes in the renal expression of (IV) chains are detected in about one-third of patients. Results of immunolabeling are concordant within families, and correlation of abnormal labeling with the severity of the clinical and pathological phenotype is usually observed.^{15, 36, 37, 46, 55}

Figure 4 Immunohistological analysis of the renal distribution of type IV collagen chains.

The analysis was carried out in (A–D) control, X-linked (E,F) male and (G,H) female Alport syndrome patients, and (I–L) patients with autosomal recessive Alport syndrome, using antibodies to 1(IV) (A,E,I), 3(IV) (B,F,J), or 5(IV) (C,G,K,L) chains. Double labeling was made with anti-2(IV) in red, and anti-5(IV) in green (D,H). In control kidney, the 1(IV) chain is present in the mesangial matrix, Bowman's capsule, and the extraglomerular basement membranes (A). The 3(IV) and 5(IV) chains are distributed within the GBM (B and C, respectively). The Bowman's capsule is strongly 5(IV)-positive (C). In X-linked Alport syndrome, no 3(IV) expression was detected in a male patient (id for 4–5) (F), whereas the distribution is segmental in a female patient (G). In autosomal recessive Alport syndrome, no 3(IV)–5(IV) labeling is detected in the GBM (J) whereas 5(IV) is expressed in Bowman's capsule (K) and the basement membranes of the collecting ducts (L). In both types of Alport syndrome, 1(IV) is diffusely expressed in the GBM (E,I). Abbreviation: GBM, glomerular basement membrane.

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Changes in the distribution of (IV) chains are also observed in skin basement membrane; absence of the 5(IV) and the associated 6(IV) chains from the epidermal basement membrane is diagnostic of X-linked Alport syndrome in male patients (Figure 5). The distribution of 5(IV) chains in the epidermal basement membrane is, however, normal (as it is in the GBM) in about 30% of male patients. Observation of normal patterns of 5(IV) and 6(IV) localization does not facilitate definitive diagnosis in females because of the segmental distribution of the chains.^{36, 55}

Figure 5 Immunohistological analysis of the distribution of the 5(IV) collagen chain.

The analysis was carried out in the skin of (A) controls, (B) male and (C) female patients with X-linked Alport syndrome, and (D) patients with autosomal recessive Alport syndrome. The 5(IV) chain is (A) present in the epidermal basement membrane (arrow) of the skin of controls, but is (B) absent from the skin basement membrane (arrow) of the male patient with X-linked Alport syndrome. (C) Segmental labeling (single arrow) is seen in the X-linked female patient. Nonspecific labeling of the keratin layer is indicated by a double arrow. (D) Normal staining patterns of the epidermal basement membrane (single arrow) are observed in autosomal recessive Alport syndrome. Nonspecific labeling of the keratin layer is indicated by a double arrow.

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Autosomal recessive Alport syndrome

Alport syndrome is transmitted as an autosomal recessive trait in about 10–15% of families affected by the disease. The clinical and morphological features are identical in the autosomal recessive and X-linked forms. Autosomal recessive Alport syndrome is usually severe; nephritis progresses to early-onset ESRD, hearing impairment affects the majority of patients, and ocular lesions may or may not be present. An autosomal recessive mode of inheritance is indicated by one or more of the following criteria: severe disease in young females; consanguinity in the family; absence of severe renal disease in the parents of a patient; microscopic hematuria in the father of an affected male; and immunohistochemical findings (see below). Homozygous or compound heterozygous mutations in COL4A3 or COL4A4 have been detected in several kindreds.^{39, 58, 59, 60, 61} As with COL4A5, there are no mutation hotspots and each family carries its own distinctive mutations.

The distribution of type IV collagen chains, detected by immunohistochemistry, can be normal in autosomal recessive Alport syndrome, but a peculiar distribution pattern of 3(IV) to 6(IV) chains is observed in the skin and kidneys of most patients.⁶² This pattern is characterized by the absence of 3, 4 and 5(IV) chains from the GBM contrasting with the persistence of 5(IV) chains in Bowman's capsules, collecting ducts and epidermal basement membranes (Figures 4 and 5). These findings show that the expression of 5(IV) chains is defective only in those basement membranes in which the three chains are associated within the 345(IV)–345(IV) network.

There is a broad spectrum of phenotypes in heterozygous carriers of autosomal recessive Alport syndrome. Some individuals are completely asymptomatic, whereas others present with persistent or intermittent microscopic hematuria (associated with thinning of the GBM in the few cases that have been examined); these symptoms are the typical clinicopathological features of 'benign familial hematuria' (see later section).⁶³ Progression to renal failure has been observed in a few heterozygous carriers.^{60, 61} This observation has implications for the consideration of relatives as living donors for kidney transplantation in recessive Alport syndrome.

Autosomal dominant Alport syndrome

Autosomal dominant inheritance of Alport syndrome, characterized by male-to-male transmission and similar disease severity in men and women, has been observed in a few families. The clinical phenotype is variable and milder than that of the X-linked dominant form. Progression to ESRD and hearing defect are not always seen and usually occur after 50 years of age; no ocular involvement has been reported. Thickening and splitting of the GBM, or diffuse thinning, have been observed, but the distribution of (IV) chains is normal. Mutations in the COL4A3 or COL4A4 genes have been reported in six unrelated families.^{64, 65}

Benign familial hematuria with thinning of the glomerular basement membrane

The exact prevalence of benign familial hematuria (BFH) is not known (estimates range from 1% to 10% of the population), but it is the most common cause of persistent hematuria.^{66, 67} BFH is inherited in an autosomal dominant manner, and is characterized by familial occurrence of persistent or recurrent hematuria that is often detected in childhood. By definition, no significant proteinuria, no progression to renal failure and no extrarenal symptoms are observed, and the prognosis is excellent.⁶³ The renal distribution of type IV collagen chains is normal. The GBM is uniformly thin. As such, the term 'thin basement membrane nephropathy' (TBMN) is currently used as an alternative term for BFH. The use of a descriptive term (TBMN) for this disorder can cause confusion, however, because of the nonspecificity of thinning of the GBM, which can also be evident in patients with Alport syndrome, including adults. Moreover, progression to ESRD has been reported in patients affected with the so-called TBMN. Thinning of the GBM can be a chance finding in recently transplanted kidneys from asymptomatic individuals, and has been observed in renal biopsy tissue from patients with other diseases such as IgA nephropathy or minimal change nephrotic syndrome.^{66, 68} Whichever term is used to describe the condition, diagnosis and prediction of a benign course of BFH/TBMN can be difficult as they are based on a series of negative findings and the presence of a nonspecific ultrastructural lesion on renal biopsy. Comprehensive familial investigations and regular follow-up assessments are of the utmost importance if BFH/TBMN is to be correctly distinguished from progressive nephritis. The onset of proteinuria or extrarenal symptoms should lead to reconsideration of the benign prognosis.⁶⁸

In 1996, a heterozygous COL4A4 mutation was detected in a large family that presented with BFH/TBMN.⁶⁹ Since then, linkage to, or mutations in, the COL4A3 and COL4A4 genes have been found in 40% of families affected by BFH/TBMN.^{66, 67, 70, 71} This finding offers a new approach to the diagnosis of the disorder; however, while detection of such mutations indicates a type IV collagen disorder, it does not exclude a diagnosis of Alport syndrome as similar mutations are harbored by people with autosomal dominant Alport syndrome. In addition, these findings confirm that BFH/TBMN represents the heterozygous state of autosomal recessive Alport syndrome. In some families, linkage to COL4A3–COL4A4 has been excluded, indicating that BFH/TBMN is genetically heterogeneous.⁷² It should be noted, however, that linkage analysis in BFH/TBMN is difficult because hematuria might be intermittent or absent in individuals with a disease-associated mutation. Conversely, hematuria could be the result of coincidental causes in individuals in whom renal biopsy has not been performed to confirm BFH/TBMN. In families in which only females (or young male children) have isolated hematuria, the existence of mutations in COL4A5, leading to diagnosis of X-linked Alport syndrome, might explain this symptom. At this time, no other locus for BFH/TBMN, besides COL4A3–COL4A4, has been identified.

Spectrum of phenotypes associated with heterozygous COL4A3 or COL4A4 mutations

Heterozygous mutations of COL4A3 or COL4A4 are associated with a diverse range of phenotypes.^{73, 74} These vary from absence of symptoms (asymptomatic carriers of autosomal recessive Alport syndrome) to isolated hematuria (BFH/TBMN), intermediate forms of nephropathy with hematuria, proteinuria without progression to renal failure, and autosomal dominant Alport syndrome with late progression to renal failure and sometimes development of hearing defects. The GBM can be thin, or thick and split. All these variants can be grouped under the term 'collagen type IV(3–4) nephropathy'.⁷⁴ To date, no phenotype–genotype correlations have been established, and prediction of prognosis is not possible on the basis of molecular analysis. More studies are needed to determine if the type and location of the mutation, the presence of additional variants in type IV collagen genes, and other genes or environmental factors can explain the observed clinical heterogeneity.

Epstein and Fechtner syndromes

Epstein and Fechtner syndromes are not type IV collagen disorders. They have, however, long been regarded as variants of autosomal dominant Alport syndrome because of the misleading association of hereditary nephritis and deafness with macrothrombocytopenia (in Epstein syndrome) and with cataracts and small blue leukocyte inclusions (in Fechtner syndrome). Both Epstein and Fechtner syndromes are linked to mutations in MYH9, which encodes the nonmuscle myosin heavy chain IIA.⁷⁵

Diagnosis

Precise diagnosis of Alport syndrome and detection of its mode of transmission are straightforward if clinical examination of the patient and familial investigations are informative. This knowledge can facilitate genetic counseling and, if there is a demand, detection of heterozygotes and prenatal diagnosis by mutation screening or linkage analysis. Frequently, however, the situation is not so straightforward. In some patients, glomerular disease seems to be sporadic, even after screening of first degree relatives for hematuria. Early onset of hematuria, and detection of a hearing defect or retinal abnormalities during systematic examination, make Alport syndrome highly probable. The diagnosis should be confirmed by gene sequencing, but in the absence of information on the mode of transmission, the first diagnostic procedure should be a skin biopsy; abnormal expression of the 5(IV) chain indicates mutation of COL4A5.

For patients who present with sporadic hematuria associated with proteinuria, but who do not have any extrarenal symptoms of Alport syndrome variants, many different diagnoses can be considered. In this context, renal biopsy facilitates exclusion of other hematuric glomerular diseases (most frequently IgA nephropathy) and identification of Alport syndrome lesions. Immunohistochemical analysis of the distribution of type IV collagen chains can point the physician towards the genetic defect. Clinical presentation of isolated hematuria in females and young male children within a family indicates a differential diagnosis of BFH/TBMN and Alport syndrome. Distinguishing between these two disorders is important because of their very different prognoses. Different management options are available, including regular follow-up examinations of patients focusing on proteinuria, retinal changes or audiologic defects, and more-active approaches based on immunohistological analysis of type IV collagen gene expression and genetic investigations.

Theoretically, identification of the underlying mutation is the gold standard for diagnosis. Mutation screening has not yet been implemented on a large scale because it is expensive and time-consuming, owing to the large size of the type IV collagen genes (48–52 exons) and the wide range of known mutations. Depending on the technique used, 50–90% of mutations can be identified.^{45, 46, 48, 49} A new screening strategy involving direct sequencing of COL4A5 complementary DNA amplified from hair-root RNA samples has been proposed.⁷⁶ This simple, fast and efficient method could fundamentally change the diagnostic approach to X-linked Alport syndrome. Such a strategy cannot, however, be applied to screening of COL4A3 and COL4A4 because these genes are not expressed in skin basement membranes. Sequencing techniques are improving rapidly, and DNA-based testing for type IV collagen gene mutations could be commercially available in the near future. Identifying mutations is important for prognostic and genetic counseling of patients and their families, and for discussions of living-related kidney donation. Early detection of disease is a prerequisite for the development of therapeutic approaches that have been shown to be effective in animal models.

Treatment

Progression to ESRD is ineluctable in males with X-linked Alport syndrome and in all patients with the autosomal recessive form of the disease. Renal transplantation is generally a satisfactory treatment, but about 2.5% of patients develop anti-GBM glomerulonephritis leading to rapid graft loss.^{36, 37, 68} Temporary or prolonged alleviation of proteinuria has been achieved by blockade of the renin–angiotensin system, but no data are available on the long-term evolution of renal function in response to this treatment. There has been controversy over the effects of ciclosporin in these patients.

Several therapeutic approaches have been tested on canine or murine models of X-linked and autosomal Alport syndrome. In Col4a3^-/- mice, transgenesis of the human COL4A3–COL4A4 locus restores the expression of type IV collagen chains and rescues the Alport phenotype.⁷⁷ Transplantation of wild-type bone-marrow-derived stem cells into irradiated Col4a3^-/- mice results in the recruitment of wild-type podocytes and mesangial cells, partial restoration of 3(IV) expression in the GBM, and improvement in glomerular structure and function.^{78, 79} Pharmacologic treatments have also been tested, and it seems that angiotensin-converting-enzyme inhibitors and angiotensin-receptor-1 antagonists,⁸⁰ chemokine receptor 1 blockade,⁸¹ statins⁸² and metalloprotease inhibitors⁸³ alleviate proteinuria and prolong the survival of treated animals, if administered before severe renal lesions develop. Accordingly, it will be necessary to be able to recognize Alport syndrome early in the course of disease development if trials of similar therapies in humans are to prove successful.

COL4A1 and familial hematuria

No mutations of COL4A1 or COL4A2 have been detected in patients with Alport syndrome or isolated BFH/TBMN.⁸⁴ Conversely, COL4A1 mutations have been identified in a mouse model of, and in families affected by, porencephaly,^{85, 86} and in a novel rare autosomal dominant syndrome known as hereditary angiopathy with nephropathy, aneurysms and cramps (HANAC). This syndrome is characterized by angiopathy with retinal tortuosities, aneurysms, muscular cramps, and nephropathy (manifesting as hematuria or cysts).⁸⁷ As such, investigation of patients with unexplained hematuria should include a search for extrarenal symptoms of HANAC, especially retinal abnormalities. If extrarenal symptoms of HANAC are detected, an attempt to identify COL4A1 mutations should be made.

Pierson syndrome

In 1963, Pierson et al. reported the curious association in siblings of eye abnormalities with microcoria and congenital nephrotic syndrome progressing rapidly to ESRD.⁸⁸ This association was also observed in a few neonates. The glomerular lesions were classified as mesangial sclerosis, with diffuse alteration of the GBM. Hypotonia and psychomotor retardation developed in the few patients who survived for several months after birth. Analysis of two large unrelated consanguineous families with several affected children led to identification of the genetic defect in 2004.⁸⁹ This defect involves the LAMB2 gene encoding the 2 chain of laminin, which is expressed at high levels in the GBM, synaptic basal laminae and basement membranes of the eye. More recently, recessive missense mutations have also been detected in two patients in a consanguineous family with isolated congenital nephrotic syndrome, a finding that expands the clinical spectrum of LAMB2-associated disorders.⁹⁰ Mice lacking laminin 2 develop massive proteinuria, and their retinal and neuromuscular differentiation is abnormal.⁹¹ Interestingly, the onset of proteinuria precedes podocyte abnormalities, showing that the GBM acts as a protein barrier and that the glomerular slit diaphragm alone is not sufficient to prevent the passage of albumin.⁹²

Nail–patella syndrome

Nail–patella syndrome (NPS), also known as hereditary osteo-onychodysplasia, is a rare autosomal dominant disorder (affecting 1 in 50,000 individuals) characterized by an association between nail hypoplasia or dysplasia and bone abnormalities that primarily affect the knees, elbows and pelvis.^{93, 94} Normal-tension glaucoma and sensorineural hearing impairment have been recognized as additional features of the disease.⁹⁴ The prognosis depends on the presence and severity of renal involvement, which is observed in 30–40% of patients. Renal involvement usually manifests as proteinuria, sometimes with hematuria. Progression to kidney failure occurs in about 30% of patients with renal symptoms, usually many years after the discovery of proteinuria, but in some cases during childhood.⁹⁵

Light microscopy of renal tissue from people with NPS reveals no specific changes. The hallmark of the disease, observed by electron microscopy, is the presence of clusters of fibrillar type III collagen irregularly distributed within thick GBM segments and the mesangial matrix.^{96, 97, 98} Staining with phosphotungstic acid is often needed to reveal the collagen bundles; when standard staining techniques are used, the GBM has a mottled appearance. The extent and distribution of GBM lesions vary widely from case to case. There is no correlation between these microscopical features and patient age or the presence or severity of renal symptoms.⁹⁸

It was initially assumed that the primary defect in NPS affected one GBM component only; however, LMX1B, which encodes a LIM–homeodomain transcription factor with a key role in development—especially in establishing the dorsoventral patterning of limbs—was identified as the mutated gene in NPS.^{99, 100, 101} Mutations are located in the LIM or homeodomain regions and are thought to result in haploinsufficiency. Female gender and mutations in the homeodomain seem to be associated with a higher risk of developing nephropathy.⁹⁴

In the kidney, LMX1B is expressed specifically in podocytes, from the S-shaped body stage of development onwards.¹⁰⁰ Compared with normal conditions, expression of several genes is downregulated in the podocytes of Lmx1b-null mice, indicating that LMX1B has an important regulatory role in podocyte differentiation and function, and providing an explanation for the development of glomerular disease in patients with NPS.^{102, 103, 104} None of the changes in glomerular expression observed in mice (i.e. downregulated expression of type IV collagen 3 and 4 chains, podocin or CD2-associated protein) was detected in people with NPS and severe glomerular disease, however.⁹⁵

Collagen type III glomerulopathy

Accumulation of type III collagen in the glomerular extracellular matrix has been detected in some proteinuric patients—mostly Japanese—in the absence of any other symptoms of NPS.^{105, 106, 107, 108, 109} In contrast to NPS glomerulopathy, diffuse glomerular changes (i.e. marked expansion of the mesangial matrix and thickening of the capillary walls) can be observed under the light microscope. At the ultrastructural level, the mesangial matrix and the subendothelial aspect of the GBM are enlarged and have a mottled appearance due to the presence of fibrillar collagen (which can be visualized after staining with phosphotungstic acid). Unlike the lesions observed in NPS, the lamina densa is usually preserved.

The clinical features of collagen type III glomerulopathy are highly variable. Two forms of the disease can be distinguished on the basis of age at onset of symptoms. In Japanese and a few Caucasian patients, the disease is usually sporadic and first symptoms—persistent proteinuria, with or without hypertension—are detected in adulthood.^{106, 108} The severity of proteinuria and serum creatinine concentration increase slowly, and renal dysfunction is a late event. Glomeruli are strikingly enlarged as a result of massive deposits of type III collagen. An elevated serum level of the type III procollagen peptide seems to be a good marker for the disease, indicating increased synthesis of type III collagen, but the fundamental underlying defect remains unknown.

Onset of first symptoms of collagen type III glomerulopathy in early childhood indicates autosomal recessive transmission of the disease.^{105, 107} Under light microscopy, subendothelial enlargement of the capillary walls is moderate and might mimic diffuse thrombotic microangiopathy. This variant of the disease is severe. Collagen type III glomerulopathy is characterized by progressive worsening of proteinuria; nephrotic syndrome eventually develops, and early occurrence of hypertension and progression to renal failure are observed in most children. Anemia of the hemolytic type and unexplained respiratory symptoms have been reported, as has abrupt progression to ESRD resulting from superimposed hemolytic–uremic syndrome. Interestingly, collagen type III glomerulopathy has been reported in one patient who presented with an inherited factor H deficiency.¹⁰⁹ Collagen type III glomerulopathy associated with factor H deficiency was also observed in our laboratory (unpublished data), in a young patient with consanguineous parents. Such findings indicate that factors that contribute to familial hemolytic–uremic syndromes might be involved in collagen type III glomerulopathy. These factors should be systematically evaluated in patients with the latter disorder.