From zebrafish to human: A comparative approach to elucidate the role of the thyroid hormone transporter MCT8 during brain development.

Monocarboxylate transporter 8 (MCT8) facilitates transmembrane transport of thyroid hormones (THs) ensuring their action on gene expression during vertebrate neurodevelopment. A loss of MCT8 in humans results in severe psychomotor deficits associated with the Allan-Herndon-Dudley Syndrome (AHDS). However, where and when exactly a lack of MCT8 causes the neurological manifestations remains unclear because of the varying expression pattern of MCT8 between specific brain regions and cells. Here, we elaborate on the animal models that have been generated to elucidate the mechanisms underlying MCT8-deficient brain development. The absence of a clear neurological phenotype in Mct8 knockout mice made it clear that a single species would not suffice. The evolutionary conservation of TH action on neurodevelopment as well as the components regulating TH signalling however offers the opportunity to answer different aspects of MCT8 function in brain development using different vertebrate species. Moreover, the plethora of tools for genome editing available today facilitates gene silencing in these animals as well. Studies in the recently generated mct8-deficient zebrafish and Mct8/Oatp1c1 double knockout mice have put forward the current paradigm of impaired TH uptake at the level of the blood-brain barrier during peri- and postnatal development as being the main pathophysiological mechanism of AHDS. RNAi vector-based, cell-specific induction of MCT8 knockdown in the chicken embryo points to an additional function of MCT8 at the level of the neural progenitors during early brain development. Future studies including also additional in vivo models like Xenopus or in vitro approaches such as induced pluripotent stem cells will continue to help unravelling the exact role of MCT8 in developmental events. In the end, this multispecies approach will lead to a unifying thesis regarding the cellular and molecular mechanisms responsible for the neurological phenotype in AHDS patients.

Comparing treatment effects on dogs with acute hemorrhagic diarrhea syndrome: fecal microbiota transplantation, symptomatic therapy, or antibiotic treatment.

OBJECTIVE: Dogs with acute hemorrhagic diarrhea syndrome (AHDS) present with similar clinical signs and histopathological findings as dogs with parvovirosis, in which fecal microbiota transplantation (FMT) has led to a significantly faster resolution of diarrhea and shorter hospitalization times. We investigated whether FMT results in faster clinical improvement and normalization of the intestinal microbiome compared to standard treatment. ANIMALS: 32 client-owned dogs with AHDS. METHODS: A prospective, double-anonymized clinical trial included 3 groups: symptomatic treatment (n = 12), FMT treatment (FMTT; 12), and antibiotic treatment (AT; 8). Clinical improvement was determined on the basis of AHDS index, changes in the microbiome based on the dysbiosis index, and PCR results for clostridial strains. RESULTS: Overall, no significant differences in clinical scores between the treatment groups over time were detected except on day 2 (higher AHDS index in the AT group compared to FMTT group; P = .046). The dysbiosis index increased and P hiranonis decreased on day 1 in some dogs, but these changes were transient in the symptomatic treatment and FMTT groups. In the AT group, the dysbiosis index was persistently elevated and 4 of 8 dogs showed a reduced abundance of P hiranonis on day 42. In 67% of the dogs on day 1, NetF-encoding Clostridium perfringens was detected and enterotoxin-encoding strains increased, but these changes were transient in all dogs, regardless of therapy. CLINICAL RELEVANCE: Overall, in dogs with AHDS, neither FMT nor AT resulted in faster clinical improvement. In addition, C perfringens strains are self-limiting and do not require antibiotic therapy.

Evaluation of Intestinal Barrier Dysfunction with Serum Iohexol Concentration in Dogs with Acute Hemorrhagic Diarrhea Syndrome.

Histopathologic examination of intestinal biopsies from dogs with acute hemorrhagic diarrhea syndrome (AHDS) reveals necrotizing enteritis and epithelial integrity loss. Serum iohexol measurement has been utilized to assess intestinal permeability. Our hypothesis is that dogs with AHDS have increased intestinal permeability, which is associated with the severity of clinical signs. In this prospective case-control study, 53 client-owned dogs (28 AHDS, 25 healthy controls) were evaluated. Clinical severity was assessed using the AHDS index and systemic inflammatory response syndrome (SIRS) criteria. Simultaneously, dogs received oral iohexol, and serum iohexol concentrations (SICs) were measured two hours later. Results indicated significantly higher (p = 0.002) SIC in AHDS dogs (median: 51 µg/mL; min-max: 9-246) than in healthy controls (30 µg/mL; 11-57). There was a significant positive correlation between AHDS index and SIC (rS = 0.4; p = 0.03) and a significant negative between SIC and serum albumin concentrations (Pearson r = -0.55; p = 0.01). Dogs with severe AHDS (mean 106 µg/mL; range: 17-246) demonstrated significantly higher (p = 0.002) SIC than those with mild to moderate disease (29 µg/mL; 9-54). These findings underscore the association between intestinal permeability and clinical severity in dogs with AHDS assessed by iohexol.

Defining a metagenomic threshold for detecting low abundances of Providencia alcalifaciens in canine faecal samples.

Introduction: Acute haemorrhagic diarrhoea syndrome (AHDS) in dogs is a condition of unknown aetiology. Providencia alcalifaciens is suspected to play a role in the disease as it was commonly found in dogs suffering from AHDS during a Norwegian outbreak in 2019. The role of this bacterium as a constituent of the canine gut microbiota is unknown, hence this study set out to investigate its occurrence in healthy dogs using metagenomics. Materials and methods: To decrease the likelihood of false detection, we established a metagenomic threshold for P. alcalifaciens by spiking culture-negative stool samples with a range of bacterial dilutions and analysing these by qPCR and shotgun metagenomics. The detection limit for P. alcalifaciens was determined and used to establish a metagenomic threshold. The threshold was validated on naturally contaminated faecal samples with known cultivation status for P. alcalifaciens. Finally, the metagenomic threshold was used to determine the occurrence of P. alcalifaciens in shotgun metagenomic datasets from canine faecal samples (n=362) collected in the HUNT One Health project. Results: The metagenomic assay and qPCR had a detection limit of 1.1x103 CFU P. alcalifaciens per faecal sample, which corresponded to a Cq value of 31.4 and 569 unique k-mer counts by shotgun metagenomics. Applying this metagenomic threshold to 362 faecal metagenomic datasets from healthy dogs, P. alcalifaciens was found in only 1.1% (95% CI [0.0, 6.8]) of the samples, and then in low relative abundances (median: 0.04%; range: 0.00 to 0.81%). The sensitivity of the qPCR and shotgun metagenomics assay was low, as only 40% of culture-positive samples were also positive by qPCR and metagenomics. Discussion: Using our detection limit, the occurrence of P. alcalifaciens in faecal samples from healthy dogs was low. Given the low sensitivity of the metagenomic assay, these results do not rule out a significantly higher occurrence of this bacterium at a lower abundance.

Unmet patient needs in monocarboxylate transporter 8 (MCT8) deficiency: a review.

Monocarboxylate transporter 8 (MCT8) deficiency is a rare, X-linked disorder arising from mutations in the SLC16A2 gene and resulting from dysfunctional thyroid hormone transport. This disorder is characterized by profound neurodevelopmental delay and motor disability due to a lack of thyroid hormone in the brain, and coexisting endocrinological symptoms, due to chronic thyrotoxicosis, resulting from elevated thyroid hormone outside the central nervous system (CNS). In February 2024, we reviewed the published literature to identify relevant articles reporting on the current unmet needs of patients with MCT8 deficiency. There are several main challenges in the diagnosis and treatment of MCT8 deficiency, with decreased awareness and recognition of MCT8 deficiency among healthcare professionals (HCPs) associated with misdiagnosis and delays in diagnosis. Diagnostic delay may also be attributed to other factors, including the complex symptomology of MCT8 deficiency only becoming apparent several months after birth and pathognomonic serum triiodothyronine (T3) testing not being routinely performed. For patients with MCT8 deficiency, multidisciplinary team care is vital to optimize the support provided to patients and their caregivers. Although there are currently no approved treatments specifically for MCT8 deficiency, earlier identification and diagnosis of this disorder enables earlier access to supportive care and developing treatments focused on improving outcomes and quality of life for both patients and caregivers.

Regulators of thyroid hormone availability and action in embryonic chicken brain development.

Thyroid hormones (THs) are crucial elements in vertebrate brain development. They exert their action mainly through binding of 3,5,3'-triiodothyronine (T3) to nuclear receptors that directly influence the expression of TH-regulated genes. Intracellular TH action is therefore dependent on both the availability of T3 and its receptors. TH uptake in cells is regulated by specific TH transporters and local activation and inactivation is regulated by deiodinases. This review provides an overview of the general expression pattern of TH transporters, deiodinases and receptors during embryonic chicken brain development and compares it to the situation in mammals. It is clear that THs and their regulators are present in the embryonic brain from the early stages of development, long before the onset of embryonic thyroid gland functioning. The mechanism of TH uptake across the brain barriers during development is only partly understood. At the developing blood-brain-barrier expression of the TH-activating type 2 deiodinase is closely associated with the blood vessels, but contrary to the situation in (adult) mammals no expression of MCT8 or OATP1C1 TH transporters is found at that level in the developing chicken. At the blood-cerebrospinal fluid-barrier co-expression of the TH-inactivating type 3 deiodinase and MCT8 and OATP1C1 is found in birds and mammals. These comparative data show overlapping patterns, pointing to general mechanisms, but also indicate specific interspecies differences that may help to understand species-specific responses to regulator gene knockout/mutation.

Corrigendum: In a zebrafish biomedical model of human Allan-Herndon-Dudley syndrome impaired MTH signaling leads to decreased neural cell diversity.

[This corrects the article DOI: 10.3389/fendo.2023.1157685.].

Parent Perspectives on Complex Needs in Patients With MCT8 Deficiency: An International, Prospective, Registry Study.

CONTEXT: Monocarboxylate transporter 8 (MCT8) deficiency is a rare neurodevelopmental and metabolic disorder, with daily care posing a heavy burden on caregivers. A comprehensive overview of these complex needs and daily care challenges is lacking. DESIGN: We established an international prospective registry to systemically capture data from parents and physicians caring for patients with MCT8 deficiency. Parent-reported data on complex needs and daily care challenges were extracted. RESULTS: Between July 17, 2018, and May 16, 2022, 51 patients were registered. Difficulties in daily life care were mostly related to feeding and nutritional status (17/33 patients), limited motor skills (12/33 patients), and sleeping (11/33 patients). Dietary advice was provided for 11/36 patients. Two of 32 patients were under care of a cardiologist. Common difficulties in the diagnostic trajectory included late diagnosis (20/35 patients) and visiting a multitude of specialists (15/35 patients). Median diagnostic delay was significantly shorter in patients born in or after 2017 vs before 2017 (8 vs 19 months, P < .0001). CONCLUSIONS: Feeding and sleeping problems and limited motor skills mostly contribute to difficulties in daily care. The majority of patients did not receive professional dietary advice, although being underweight is a key disease feature, strongly linked with poor survival. Despite sudden death being a prominent cause of death, potentially related to the cardiovascular abnormalities frequently observed, patients were hardly seen by cardiologists. These findings can directly improve patient-centered multidisciplinary care and define patient-centered outcome measures for intervention studies in patients with MCT8 deficiency.

In a zebrafish biomedical model of human Allan-Herndon-Dudley syndrome impaired MTH signaling leads to decreased neural cell diversity.

Background: Maternally derived thyroid hormone (T3) is a fundamental factor for vertebrate neurodevelopment. In humans, mutations on the thyroid hormones (TH) exclusive transporter monocarboxylic acid transporter 8 (MCT8) lead to the Allan-Herndon-Dudley syndrome (AHDS). Patients with AHDS present severe underdevelopment of the central nervous system, with profound cognitive and locomotor consequences. Functional impairment of zebrafish T3 exclusive membrane transporter Mct8 phenocopies many symptoms observed in patients with AHDS, thus providing an outstanding animal model to study this human condition. In addition, it was previously shown in the zebrafish mct8 KD model that maternal T3 (MTH) acts as an integrator of different key developmental pathways during zebrafish development. Methods: Using a zebrafish Mct8 knockdown model, with consequent inhibition of maternal thyroid hormones (MTH) uptake to the target cells, we analyzed genes modulated by MTH by qPCR in a temporal series from the start of segmentation through hatching. Survival (TUNEL) and proliferation (PH3) of neural progenitor cells (dla, her2) were determined, and the cellular distribution of neural MTH-target genes in the spinal cord during development was characterized. In addition, in-vivo live imaging was performed to access NOTCH overexpression action on cell division in this AHDS model. We determined the developmental time window when MTH is required for appropriate CNS development in the zebrafish; MTH is not involved in neuroectoderm specification but is fundamental in the early stages of neurogenesis by promoting the maintenance of specific neural progenitor populations. MTH signaling is required for developing different neural cell types and maintaining spinal cord cytoarchitecture, and modulation of NOTCH signaling in a non-autonomous cell manner is involved in this process. Discussion: The findings show that MTH allows the enrichment of neural progenitor pools, regulating the cell diversity output observed by the end of embryogenesis and that Mct8 impairment restricts CNS development. This work contributes to the understanding of the cellular mechanisms underlying human AHDS.

C-reactive protein as a tool for monitoring response to treatment in dogs with acute hemorrhagic diarrhea syndrome.

Objectives: C-reactive protein (CRP) is an established marker for systemic inflammation in dogs that is especially elevated in dogs with sepsis. Some dogs with acute hemorrhagic diarrhea syndrome (AHDS) develop bacterial translocation and consequent sepsis during hospitalization. This study aimed to evaluate the course of CRP plasma concentrations during hospitalization and its correlation with clinical and other laboratory variables in dogs with AHDS. Methods: In this prospective, observational study, CRP was evaluated on days 0, 1, 2, and 3 in 27 client-owned dogs who presented with AHDS. Clinical examination data, blood pressure, acute patient physiologic and laboratory evaluation (APPLE) full and APPLE fast scores, and canine hemorrhagic diarrhea severity (CHDS) index were measured on the same days to evaluate the severity of the disease. Results: Twenty-five of the 27 dogs were discharged from hospital. Nineteen dogs received antimicrobial treatment due to sepsis or neutropenia. CRP values were mildly elevated on day 0 (median 27.3 mg/L; 1.0-125.8 mg/L) and markedly elevated on day 1 (median 88.9 mg/L; 1.4-192.7 mg/L). CRP concentrations decreased gradually over the following days. Moreover, CRP concentrations correlated moderately with albumin, leucocyte count, neutrophil count, and APPLE full and fast scores, but not with antimicrobial treatment. Conclusion and relevance: CRP concentrations were significantly elevated in patients with AHDS. In this study population, CRP did not help in detecting the requirement of antimicrobial treatment in dogs with AHDS. Nevertheless, as CRP can monitor the response to treatment, regular analysis can guide treatment.

Frequency of signs of chronic gastrointestinal disease in dogs after an episode of acute hemorrhagic diarrhea.

BACKGROUND: Acute enteropathy is a trigger of chronic gastrointestinal (GI) disease in humans. OBJECTIVE: To report the prevalence of and explore possible risk factors for signs of chronic GI disease in dogs after an episode of acute hemorrhagic diarrhea (AHD). ANIMALS: One hundred and fifty-one dogs, 80 dogs with a historical diagnosis of AHD, 71 control dogs with no history of AHD. METHODS: In this retrospective longitudinal study, data were collected from dogs with a historical diagnosis of AHD and healthy controls matched by breed, age and sex, aged between 1 year and 15 years of age, for which a follow-up of at least 12 months after enrolment was available. Dog owners responded to a questionnaire to determine the history of signs of chronic GI disease. RESULTS: There was a higher prevalence of signs of chronic GI disease in the dogs with a previous episode of AHD compared to control dogs (AHD 28%; controls 13%; P = .03; odds ratio = 2.57; confidence interval [CI] 95% 1.12-6.31) over a similar observation time (median 4 years; range, 1-12 years). CONCLUSIONS AND CLINICAL IMPORTANCE: Severe intestinal mucosal damage and associated barrier dysfunction might trigger chronic GI disease later in life.

Long-Term Efficacy of T3 Analogue Triac in Children and Adults With MCT8 Deficiency: A Real-Life Retrospective Cohort Study.

CONTEXT: Patients with mutations in thyroid hormone transporter MCT8 have developmental delay and chronic thyrotoxicosis associated with being underweight and having cardiovascular dysfunction. OBJECTIVE: Our previous trial showed improvement of key clinical and biochemical features during 1-year treatment with the T3 analogue Triac, but long-term follow-up data are needed. METHODS: In this real-life retrospective cohort study, we investigated the efficacy of Triac in MCT8-deficient patients in 33 sites. The primary endpoint was change in serum T3 concentrations from baseline to last available measurement. Secondary endpoints were changes in other thyroid parameters, anthropometric parameters, heart rate, and biochemical markers of thyroid hormone action. RESULTS: From October 15, 2014 to January 1, 2021, 67 patients (median baseline age 4.6 years; range, 0.5-66) were treated up to 6 years (median 2.2 years; range, 0.2-6.2). Mean T3 concentrations decreased from 4.58 (SD 1.11) to 1.66 (0.69) nmol/L (mean decrease 2.92 nmol/L; 95% CI, 2.61-3.23; P < 0.0001; target 1.4-2.5 nmol/L). Body-weight-for-age exceeded that of untreated historical controls (mean difference 0.72 SD; 95% CI, 0.36-1.09; P = 0.0002). Heart-rate-for-age decreased (mean difference 0.64 SD; 95% CI, 0.29-0.98; P = 0.0005). SHBG concentrations decreased from 245 (99) to 209 (92) nmol/L (mean decrease 36 nmol/L; 95% CI, 16-57; P = 0.0008). Mean creatinine concentrations increased from 32 (11) to 39 (13) µmol/L (mean increase 7 µmol/L; 95% CI, 6-9; P < 0.0001). Mean creatine kinase concentrations did not significantly change. No drug-related severe adverse events were reported. CONCLUSIONS: Key features were sustainably alleviated in patients with MCT8 deficiency across all ages, highlighting the real-life potential of Triac for MCT8 deficiency.

A retrospective study of 237 dogs hospitalized with suspected acute hemorrhagic diarrhea syndrome: Disease severity, treatment, and outcome.

BACKGROUND: Few studies have investigated management and outcome in dogs with acute hemorrhagic diarrhea syndrome (AHDS), and there is a paucity of data on dogs with concurrent signs of sepsis. OBJECTIVES: To report outcome in dogs with suspected AHDS according to disease severity and antimicrobial treatment, and to evaluate effect of fluid resuscitation on clinical criteria. ANIMALS: Two hundred thirty-seven dogs hospitalized with suspected AHDS. METHODS: Retrospective study based on medical records. Disease severity was evaluated using AHDS index, systemic inflammatory response syndrome (SIRS) criteria, and serum C-reactive protein (CRP) according to 3 treatment groups: No, 1, or 2 antimicrobials. RESULTS: Sixty-two percent received no antimicrobials, 31% received 1 antimicrobial, predominantly aminopenicillins, and 7% received 2 antimicrobials. At admission, median AHDS index was 13 (interquartile range, 11-15), which decreased significantly after the first day's hospitalization (P < .001) for all groups. Compared with no antimicrobials (7%), more dogs had ≥2 SIRS criteria in the antimicrobial groups (15% and 36%, respectively). C-reactive protein (CRP) correlated positively with AHDS index at hospitalization (P < .001). Across treatment groups, rehydration markedly reduced number of clinical SIRS criteria. Survival to discharge was 96%, lower for dogs receiving 2 antimicrobials (77%, P < .05). CONCLUSIONS AND CLINICAL IMPORTANCE: The majority of dogs hospitalized with suspected AHDS improve rapidly with symptomatic treatment only, despite signs of systemic disease on initial presentation. The often-used SIRS criteria might be a poor proxy for identifying dogs with AHDS in need of antimicrobial treatment, in particular when hypovolemic. The role of CRP in clinical decision-making or prognostication warrants further investigation.

Identification of Inhibitors Based on Molecular Docking: Thyroid Hormone Transmembrane Transporter MCT8 as a Target.

AIMS: To identify natural inhibitors against MCT8 for Allan-Herndon-Dudley Syndrome. BACKGROUND: Monocarboxylate Transporter 8 (MCT8) is a Thyroid Hormone (TH) transporter which is highly expressed in the liver and brain. Mutations in the MCT8 gene (SLC16A2) cause a syndrome of psychomotor retardation in humans, known as Allan-Herndon-Dudley syndrome (AHDS). Currently, no treatment is available for AHDS. Therefore, there is a need to discover new inhibitors of MCT8 for treating AHDS. OBJECTIVE: Considering the importance of natural compounds in drug discovery, this study aimed to identify potential natural inhibitors against MCT8. METHODS: As Protein-ligand interactions play a key role in structure based drug design, this study screened 24 natural kinase inhibitors and investigated their binding affinity against MCT8 by using molecular docking. The modelled 3D structure of MCT8 docked with 24 compounds using PyRX through Autodock Vina. Drug-likeness studies were made using Swiss ADME and Lipinski's rule of five was performed. Triac, desipramine and silychristin were used as the positive controls. Binding energies of the selected compounds were compared with that of positive controls. RESULT: The results showed that emodin exhibited best binding energy of -8.6 kcal/mol followed by helenaquinol, cercosporamide and resveratrol. Moreover, it was observed that emodin and helenaquinol exhibit higher binding energy than the positive controls. Cercosporamide and resveratrol exhibited higher binding energy than triac and desipramine and showed the binding energy similar to silychristin. CONCLUSION: This study reveals that these compounds could be promising candidates for further evaluation for AHDS prevention.

The Role of Thyroid Function Tests in Diagnosing Allan-herndon-dudley Syndrome Revisited: A Novel Iran-based Mutation.

INTRODUCTION: Allan-Herndon-Dudley Syndrome (AHDS) is a rare X-linked recessive intellectual disability condition with neuromuscular involvements. Altered thyroid function tests are major milestones in AHDS diagnosis. However, due to phenotypic variations in the levels of thyroid hormones in AHDS patients, we believe that the disorder is often underdiagnosed. Here, we reported a 3.5-year-old boy with an AHDS diagnosis and healthy thyroid hormones. METHODS: Whole-Exome sequencing followed by data analysis was performed on the patient's sample. The mutation was confirmed by Sanger sequencing in the patient and his mother. RESULTS: We reported a 3.5-year-old boy with AHDS diagnosis and a novel synonymous missense mutation (c. 1026G>A) in the SLC16A2 gene manifesting normal levels of T3, T4, and TSH. The mutation causes no change in amino acid sequence; however, it affects splicing through alteration of an exonic splicing enhancer. To the best of our knowledge, there are only 3 similar reports in the literature reporting AHDS diagnosis and normal levels of thyroid hormones. CONCLUSION: The altered levels of thyroid hormones are notable but not necessary markers for diagnosing AHDS. The candidate diagnosis of AHDS should be considered in patients with X-linked recessive intellectual disability syndrome with neuromuscular involvements irrespective of levels of thyroid hormones; otherwise, it could lead to the under-diagnosis of the disorder.

Impact of Changes in Gastrointestinal Microbiota in Canine and Feline Digestive Diseases.

The intestinal microbiome is an important immune and metabolic organ in health and disease. Recent molecular and metabolomic approaches have provided a better characterization of different types of dysbiosis, including mucosa-adherent bacteria and functional changes in the microbiome. This article summarizes recent advances in assessment of dysbiosis, the importance of the bile acid-converting Clostridium hiranonis as an important beneficial bacterium in the canine gut, and different therapeutic approaches to dysbiosis.

Monocarboxylate Transporter 8 Deficiency: From Pathophysiological Understanding to Therapy Development.

Genetic defects in the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) result in MCT8 deficiency. This disorder is characterized by a combination of severe intellectual and motor disability, caused by decreased cerebral thyroid hormone signalling, and a chronic thyrotoxic state in peripheral tissues, caused by exposure to elevated serum T3 concentrations. In particular, MCT8 plays a crucial role in the transport of thyroid hormone across the blood-brain-barrier. The life expectancy of patients with MCT8 deficiency is strongly impaired. Absence of head control and being underweight at a young age, which are considered proxies of the severity of the neurocognitive and peripheral phenotype, respectively, are associated with higher mortality rate. The thyroid hormone analogue triiodothyroacetic acid is able to effectively and safely ameliorate the peripheral thyrotoxicosis; its effect on the neurocognitive phenotype is currently under investigation. Other possible therapies are at a pre-clinical stage. This review provides an overview of the current understanding of the physiological role of MCT8 and the pathophysiology, key clinical characteristics and developing treatment options for MCT8 deficiency.

Clinical and Functional Consequences of C-Terminal Variants in MCT8: A Case Series.

CONTEXT: Genetic variants in SLC16A2, encoding the thyroid hormone transporter MCT8, can cause intellectual and motor disability and abnormal serum thyroid function tests, known as MCT8 deficiency. The C-terminal domain of MCT8 is poorly conserved, which complicates prediction of the deleteriousness of variants in this region. We studied the functional consequences of 5 novel variants within this domain and their relation to the clinical phenotypes. METHODS: We enrolled male subjects with intellectual disability in whom genetic variants were identified in exon 6 of SLC16A2. The impact of identified variants was evaluated in transiently transfected cell lines and patient-derived fibroblasts. RESULTS: Seven individuals from 5 families harbored potentially deleterious variants affecting the C-terminal domain of MCT8. Two boys with clinical features considered atypical for MCT8 deficiency had a missense variant [c.1724A>G;p.(His575Arg) or c.1796A>G;p.(Asn599Ser)] that did not affect MCT8 function in transfected cells or patient-derived fibroblasts, challenging a causal relationship. Two brothers with classical MCT8 deficiency had a truncating c.1695delT;p.(Val566*) variant that completely inactivated MCT8 in vitro. The 3 other boys had relatively less-severe clinical features and harbored frameshift variants that elongate the MCT8 protein [c.1805delT;p.(Leu602HisfsTer680) and c.del1826-1835;p.(Pro609GlnfsTer676)] and retained ~50% residual activity. Additional truncating variants within transmembrane domain 12 were fully inactivating, whereas those within the intracellular C-terminal tail were tolerated. CONCLUSIONS: Variants affecting the intracellular C-terminal tail of MCT8 are likely benign unless they cause frameshifts that elongate the MCT8 protein. These findings provide clinical guidance in the assessment of the pathogenicity of variants within the C-terminal domain of MCT8.

Insights Into the Mechanism of MCT8 Oligomerization.

Mutations in the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) result in MCT8 deficiency, characterized by severe intellectual and motor disability. The MCT8 protein is predicted to have 12 transmembrane domains (TMDs) and is expressed as monomers, homodimers, and homo-oligomers. This study aimed to delineate the mechanism of MCT8 oligomerization. Coimmunoprecipitation studies demonstrated that lithium dodecyl sulfate effectively disrupts MCT8 protein complexes, indicating the involvement of non-covalent interactions. Successive C-terminal truncations of the MCT8 protein altered the oligomerization pattern only if introduced in the N-terminal half of the protein (TMD1-6). The truncation at extracellular loop 1 (E206X) still allowed homodimerization, but completely abrogated homo-oligomerization, whereas both were preserved by the C231X mutant (at TMD2), suggesting that the minimally required oligomerization sites are located proximal of Cys231. However, mutant constructs lacking the intracellular N-terminus or TMD1 and 2 were still capable to form homo-oligomers. Therefore, other domains distal of Cys231 are also likely to be involved in the formation of extensive multidomain interactions. This hypothesis was supported by structural modeling. Despite multiple approaches, MCT8 oligomerization could not be fully abrogated unless a substantial part of the protein was removed, precluding detailed studies into its functional role. Together, our findings suggest that MCT8 oligomerization involves extensive noncovalent interactions between the N-terminal halves of MCT8 proteins. Most mutations identified in patients with MCT8 deficiency have only minor effects on MCT8 oligomerization and, thus, impaired oligomerization does not appear to be an important pathogenic mechanism.

Thyroid Hormone Transporters.

Thyroid hormone transporters at the plasma membrane govern intracellular bioavailability of thyroid hormone. Monocarboxylate transporter (MCT) 8 and MCT10, organic anion transporting polypeptide (OATP) 1C1, and SLC17A4 are currently known as transporters displaying the highest specificity toward thyroid hormones. Structure-function studies using homology modeling and mutational screens have led to better understanding of the molecular basis of thyroid hormone transport. Mutations in MCT8 and in OATP1C1 have been associated with clinical disorders. Different animal models have provided insight into the functional role of thyroid hormone transporters, in particular MCT8. Different treatment strategies for MCT8 deficiency have been explored, of which thyroid hormone analogue therapy is currently applied in patients. Future studies may reveal the identity of as-yet-undiscovered thyroid hormone transporters. Complementary studies employing animal and human models will provide further insight into the role of transporters in health and disease. (Endocrine Reviews 41: 1 - 55, 2020).