2.1. Molecular Basis of Gitelman Syndrome and Clinical Consequences
As mentioned previously, GS and BS share pathological conditions, including hypokalemia, hypochloremic metabolic alkalosis, hyperreninemia and secondary hyperaldosteronism
[3][4]. However, the main difference is that GS also presents with hypocalciuria. This is due to an increase in Ca
2+ reabsorption in an attempt to compensate for the loss of salts
[20]. This compensatory process does not occur in patients with BS, since electrolyte dysregulation in the TAL causes a lack of the positive light gradient necessary for paracellular Ca
2+ reabsorption
[20][21]. Active transport of Ca
2+ remains unaltered in GS
[20][21].
Another relevant molecular characteristic is the presence of hypomagnesemia, mainly in cases of GS. However, this anomaly is not a complete differentiation between GS and BS
[22] since it can also be found in certain cases of BS
[23]. Although this phenomenon has been studied for more than 20 years
[21][24][25], the reason for this magnesium loss is not completely understood. One of the most accepted hypotheses points to the alteration in the expression of the TRPM6 channel
[26], the principal channel by which Mg
2+ is reabsorbed in DCT (
Figure 1). Moreover, there is a strict correlation between hypomagnesemia and chondrocalcinosis
[27].
Figure 1. Proteins and channels implicated in the pathogenesis of Gitelman and Bartter syndromes. The electrolyte transports of the most important channels for the diseases are represented as well as the channels related to the inhibition by thiazide (NCC) and furosemide diuretics (NKCC2). Each disease is accompanied by the causative gene (in capital letters, brackets and italics), whereas the corresponding protein is indicated above the channel (only in capital letters). NCC: Solute carrier family 12 member 3; MAGED2: Melanoma-associated antigen D2; TRPM6: Transient receptor potential cation channel subfamily M member 6; CLCNKB: Chloride channel protein ClC-Kb; NKCC2: Solute carrier family 12 member 1; BSND: Barttin; CLCNKA: Chloride channel protein ClC-Ka; and ROMK: ATP-sensitive inward rectifier potassium channel 1. The positive charge of the DCT makes an electrochemical gradient from the luminal tubule from the interstitium possible.
Mg
2+ is a cofactor of the group of pyrophosphatases, particularly for alkaline phosphatase
[28]. A decrease in the concentration of magnesium causes a dysfunction of these proteins, which increases the levels of pyrophosphate. Inorganic pyrophosphate binds to Ca
2+ ions by ionic interaction, resulting in crystal formation. These crystals are deposited over time, eventually causing chondrocalcinosis
[29]. In fact, magnesium is a crucial factor in the prognosis of GS, due to the possible development of such pathological conditions
[27].
Historically, GS has been considered a benign disease, which was generally diagnosed incidentally by the presence of cramps, extreme fatigue, tetany or muscle weakness
[5]. However, there is a wide phenotypic variability among patients, ranging from asymptomatic individuals to individuals with a severe phenotype
[22]. Furthermore, it was considered as a “desirable disease” in the sense that patients typically have normal or low blood pressure
[22][30]. For instance, a recent case report described a kidney transplant from a donor with GS, in which the possibility of lowering the blood pressure in the recipient was considered beneficial
[31]. However, clinical surgeries like this can be seen as controversial.
Similarly, GS (as well as BS) was considered as a human model of hypotension, since it was seen that carriers of pathogenic heterozygous mutations in their respective genes were associated with a lower blood pressure than that of the control population
[32]. Interestingly, a study performed in 2000 had already anticipated that some pathogenic mutations in
SLC12A3 could protect against hypertension, which coincides with the literature
[31][32][33][34][35][36]. Nevertheless, the presence of some homozygous variants is associated with primary hypertension
[37]. A relevant study confirmed the possibility that Gitelman’s patients may develop hypertension due to continuous activation of the RAAS axis
[38]. The role of zygosity in this process has yet to be elucidated.
In addition to hypertension, several phenotypes have also been related to this disease. For example, it has been stated that patients with GS have a greater predisposition to viral infections
[39][40], and they are more prone to develop type II diabetes mellitus
[26][41][42]. Moreover, it was recently postulated that they also have an abnormal glycosylation pattern in angiotensin converting enzyme 2 (ACE2), which leads to the activation of RAAS (also seen in BS patients)
[35].
Long-term population studies might provide a better understanding and anticipation of the comorbidities that Gitelman’s patients might face. It is essential to make a great effort in the study of genotype-phenotype correlations, emphasizing the correlations with different levels of blood pressure values.
2.2. Molecular Basis of Bartter Syndrome and Clinical Consequences
In terms of presentation, Bartter Syndrome has been traditionally grouped into neonatal or classic Bartter Syndrome. The neonatal refers to a severe form with an antenatal presentation that leads to serious polyuria. Consequently, polyhydramnios, premature delivery and severe cases of electrolyte and water loss occur
[43]. The classic type refers to a more subtle presentation that can occur at any time, but typically in early childhood, with polyuria, polydipsia, volume contraction and muscle weakness
[23]. Currently, the different forms of Bartter Syndrome are classified into six subtypes based on the underlying gene. Moreover, the different subtypes can be further grouped into three categories, based on the similarity between the main molecular mechanisms in which the encoded products participate and their associated pathophysiology: BS type 1 and 2; BS types 3 and 4; and BS type 5.
The proteins NKCC2 (BS type 1) and ROMK (BS type 2) are the main players in the reabsorption of solutes in the TAL
[44]. When they are disrupted, the physiological abnormalities lead to an early phenotype, usually with manifestations appearing during the prenatal stage
[45]. The main consequences of its dysfunction in the embryonic stage includes electrolyte imbalances that can cause polyuria due to isosthenuria, polyhydramnios, preterm birth
[46] as well as subsequent growth retardation, serious episodes of salt loss, hypercalciuria and metabolic alkalosis with hypokalemia and hypochloremia
[45]. Although type 1 and type 2 share most of these symptoms, the appearance of episodes of early transient hyperkalemia is seen mostly in type 2
[47].
Newborns with mutations in the
KCNJ1 gene may not be able to excrete potassium via ROMK and other non-canonical mechanisms restore potassium levels later
[45]. Due to the strict correlation and functional of the transporters in the TAL, the loss of function of the ROMK channels (BS type 2) could lead to the inactivation of NKCC2 (BS type 1)
[48], which could justify the overlapping phenotype in both types. Furthermore, an interaction of both channels has also been demonstrated in the secretion of uromodulin protein
[49][50], which is decreased in patients with type 2 Bartter
[50]. Thus, these results point to a possible role for uromodulin in tubular disorders.
In addition, new studies on the function of NKCC2 channels have shown that stoichiometry can change due to mutations in the
SLC12A1 gene
[51]. The type of mutation can determine differences in electrolyte disorders, which would explain the high phenotypic variability between individuals. For example, particular genetic alterations could mean that the Na
+ K
+ 2Cl
− cotransporter changes to the unique Na
+ Cl
− transport. Genotype-phenotype correlation studies of large cohorts could help determine the relevance of the type of mutations in relation to the phenotype.
BS type 3 is caused by genetic abnormalities in
CLCNKB gene. The transporter encoded by this gene mediates the reabsorption of chloride from tubular cells to peritubular capillaries in TAL and DCT
[2][23]. This type of BS is one of the most investigated, due to the clinical similarity with GS and the need to differentiate them for an accurate diagnosis. It is characterized by enormous clinical variability. Thus, the first manifestations can appear at any time, from the antenatal to the adult stage, and truncating variants are mainly associated with early onset of the disease
[23].
The central feature of BS type 3 is severe hypochloremia
[45]. CLCNKB is one of the channels necessary for the reabsorption of NaCl
[23] in TAL, and its function has to be intact for the reabsorption of chlorine in DCT
[52][53]. In fact, its total genetic inactivation is incompatible with life
[54]. An incorrect reabsorption can affect the functionality in the HCO3
−/Cl
− exchanger
[45], and thus chloride homeostasis is totally damaged in this segment of the nephron. CLCNKA cannot compensate for the loss of function in CLCNKB, on the basis that
CLCNKA expression is decreased in orthologous
CLCNKB null mice
[52].
BS type 4 can be caused by mutations in
BSND (type 4a) or digenic recessive mutations in CLCK– channels (
CLCNKB and
CLCNKA, Type 4b). In both cases, congenital deafness is a differential symptom
[55][56], due to the loss of potential load in the inner ear and the incorrect function of chloride channels.
Furthermore, the BSND is not properly y localized when CLCNK channels are absent
[54]. BSND is a mandatory subunit for the normal function of all CLCNK channels
[12][56], so the phenotype of BS type 4 is more severe than BS type 3. Interestingly, although BSND is also present in DCT, BS type 4 is generally not confused with GS, as is the case with type 3 BS. In contrast to the other types of BS, renal failure is mainly associated with patients with type 4 BS
[12].
Since the first discovery in 2004 of recessive digenic inheritance of the
CLCNKB and
CLCNKA genes
[14][15], only a few cases of BS type 4b have been reported since then. This is because these types of cases, in addition to being rare, are not usually included in population-based studies of BS. This complexity can add another intriguing aspect. Given that large heterozygous deletions of the physically contiguous genes,
CLCNKA to
CLCNKB have been reported, the question remains, is CLCNKA actually associated with the disease? Until now, no homozygous or heterozygous patients with
CLCNKA have been phenotypically reported.
The homozygous inactivation of the orthologous
CLCNKA gene in animal models resembles the phenotype of diabetes insipidus, although the mice do not lose salt
[57]. As in the case of Alport syndrome associated with the contiguous deletion of the
COL4A5 and
COL4A6 genes
[58], excluding the association of the
COL4A6 gene with said syn-drome
[59], it would be interesting to identify the clinical or molecular relevance of the inactivation of the
CLCNKA gene alone in BS. Recently, the p.R83G variant in
CLCNKA has been postulated as the putative gene loci for a major incidence of heart failure in dilated cardiopathy
[60]. Except for this association, thus far, no possible phenotypes directly related to the functional deficiency of the
CLCNKA gene have been identified.Bartter syndrome type 5.
The clinical characteristics of patients with mutations in the
MAGED2 gene are very similar to classic Bartter, highlighting polyuria, hyperreninism and hyperaldosteronism. However, the most relevant clinical findings consist of severe polyhydramnios, premature birth and perinatal complications. Despite starting as a severe form of Bartter syndrome
[13], phenotypic restoration occurs spontaneously, without the need for any specific treatment.
The explanation for this temporal manifestation lies in the fact that the apical localization of NCC (
SLC12A3) and NKCC2 (
SLC12A1) depends on MAGED2 during the developmental stages in humans
[13]. It is logical that the diminished ubieties for both cotransporters bring about a severe phenotype
[61]. Two theories have been put forward on the pathophysiology related to MAGED deficiency. The first is that MAGED2 binds to Hsp40 to regulate endoplasmic reticulum-associated degradation (ERAD), and therefore mutations in
MAGED2 cause alterations in this process
[62] and NCC and NKCC2 are retained intracellularly.
The second postulates that the mutations in
MAGED2 prevent sufficient concentrations of cyclic adenosine monophosphate (cAMP) for the correct function of the antidiuretic hormone (ADH), causing the mislocalization of the channels
[61]. It is essential to bear in mind that ERAD, lysosomal degradation and the specific ubiquitination of unfolded and immature NCC and NKCC2 have also been widely described as key mechanisms of protein expression and localization
[63][64][65][66], for which further study of Bartter’s disease type 5 will contribute to a greater understanding of these fundamental processes. Similarly, the presence of mutations in the
MAGED2 gene could explain the cases of idiopathic polyhydramnios and unresolved prenatal tubulopathies
[67].