Lysinuric Protein Intolerance and Its Nutritional and Multisystemic Challenges in Pregnancy: A Case Report and Literature Review.

Lysinuric protein intolerance (LPI) is a rare inborn error of metabolism (IEM), classified as an inherited aminoaciduria, caused by mutations in the SLC7A7 gene, leading to a defective cationic amino acid transport. The metabolic adaptations to the demands of pregnancy and delivery cause significant physiological stress, so those patients affected by IEM are at greater risk of decompensation. A 28-year-old woman with LPI had experienced 3 early miscarriages. While pregnancy was finally achieved, diverse nutritional and medical challenges emerged (food aversion, intrauterine growth restriction, bleeding risk, and preeclampsia suspicion), which put both the mother and the fetus at risk. Moreover, the patient requested a natural childbirth (epidural-free, delayed cord clamping). Although the existence of multiple safety concerns rejected this approach at first, the application of novel strategies made a successful delivery possible. This case reinforces that the woman's wish for a non-medicated, low-intervention natural birth should not be automatically discouraged because of an underlying complex metabolic condition. Achieving a successful pregnancy is conceivable thanks to the cooperation of interdisciplinary teams, but it is still important to consider the risks beforehand in order to be prepared for possible additional complications.

Plasma calprotectin is extremely high in patients with lysinuric protein intolerance.

Lysinuric protein intolerance (LPI) is a rare autosomal recessive disorder affecting the transport of cationic amino acids. Elevated plasma zinc concentrations have been described in patients with LPI. Calprotectin is a calcium- and zinc-binding protein, produced by polymorphonuclear leukocytes and monocytes. Both zinc and calprotectin have an important role in immune system. In this study, we describe plasma zinc and plasma calprotectin concentrations in Finnish LPI patients. Plasma calprotectin concentration was measured from 10 LPI patients using an enzyme-linked immunosorbent assay (ELISA) and it was remarkably high in all LPI patients (median: 622 338 µg/L) compared to that in healthy controls (608 µg/L). Plasma zinc concentration was measured by photometry and it was normal or only mildly elevated (median: 14.9 µmol/L). All the patients had decreased glomerular infiltration rate (median: 50 mL/min/1.73 m2). In conclusion, we observed extremely high plasma calprotectin concentration in patients with LPI. Mechanism of this phenomenon is unknown.

X-linked pyruvate dehydrogenase complex deficiency due to a novel PDHA1 variant associated with structural brain abnormalities in a fetus.

We report a case of pyruvate dehydrogenase E1 alpha subunit deficiency associated with a novel hemizygous PDHA1 variant presenting prenatally as multiple structural brain abnormalities in a male fetus. A healthy Finnish couple was initially referred to the Fetomaternal Medical Center because of suspected fetal choroid plexus cyst at 11 + 2 weeks of pregnancy. At 20 + 0 weeks, multiple abnormalities were observed with ultrasound including narrow thorax, slightly enlarged heart, hypoplastic cerebellum, absent cerebellar vermis and ventriculomegaly. Autopsy and genetic analyses were performed after the termination of pregnancy. The findings of macroscopic examination included cleft palate, abnormally overlapping position of fingers and toes and dysmorphic facial features. Neuropathological examination confirmed the absence of corpus callosum, cerebellar hypoplasia and ventriculomegaly. Nodular neuronal heterotopia was also observed. Trio exome sequencing revealed a novel hemizygous de novo variant c.1144C>T p.(Gln382*) in the PDHA1 gene, classified as likely pathogenic. We suggest that inherited metabolic disorders should be kept in mind as differential diagnoses in fetuses with structural brain abnormalities.

Genetic spectrum of prenatally diagnosed skeletal dysplasias in a Finnish patient cohort.

OBJECTIVE: This retrospective cohort study aims to describe the genetic spectrum of fetal skeletal dysplasias detected in a Finnish patient cohort and the diagnostic yield of various analysis methods used. METHOD: A total of 121 pregnancies with prenatally suspected or diagnosed skeletal dysplasia were analyzed between 2013 and 2020. Clinical details and findings from genetic testing were collected. RESULTS: Abnormal ultrasound triggered further testing in most cases. However, there were several cases with increased nuchal translucency and/or abnormal risk ratio in the first trimester combined screening as the initial finding. Further genetic testing was performed in 84/121 (69.4%) cases. A genetic diagnosis was confirmed in 36/84 (42.9%) cases. Half of the identified cases could be attributed to a founder mutation specific to the Finnish Disease Heritage, whereas the other half consisted of a variety of other genetic defects. CONCLUSION: In our patient cohort, the overall genetic spectrum of prenatally diagnosed skeletal dysplasias was wide. However, the impact of Finnish founder mutations was considerable, suggesting that the genetic spectrum of skeletal dysplasias may differ significantly between populations. This should be taken into consideration during the diagnostic process especially as initial ultrasound findings may be unspecific and the interpretation of ultrasound features is usually difficult.

Platelet function and filamin A expression in two families with novel FLNA gene mutations associated with periventricular nodular heterotopia and panlobular emphysema.

Pathogenic variants of the X-linked FLNA gene encoding filamin A protein have been associated with a wide spectrum of symptoms, including the recently described pulmonary phenotype with childhood-onset panlobular emphysema. We describe three female patients from two families with novel heterozygous FLNA variants c.5837_2del and c.508C > T. Analysis of immunofluorescence of peripheral blood smears and platelet function was performed for all patients. FLNA-negative platelets were observed, suggesting that these variants result in the loss of a functional protein product. All three patients also had periventricular nodular heterotopia and panlobular emphysema. However, they had considerably milder symptoms and later age of onset than in the previously reported cases. Therefore, patients with pathogenic FLNA variants should be studied actively for lung involvement even in the absence of pronounced respiratory symptoms. Conversely, any patient with unexplained panlobular emphysema should be analyzed for pathogenic FLNA variants. We also suggest that immunofluorescence analysis is a useful tool for investigating the pathogenicity of novel FLNA variants.

Prenatal array comparative genomic hybridization in a well-defined cohort of high-risk pregnancies. A 3-year implementation results in a public tertiary academic referral hospital.

OBJECTIVE: To find out whether the diagnostic yield of prenatal array comparative genomic hybridization (aCGH) can be improved by targeting preselected high-risk pregnancies. METHOD: All the in-house arrays ordered by the Fetomaternal Medical Center from February 2016 until December 2018 were retrospectively analyzed. The indications for array analysis included fetal structural abnormalities, increased nuchal translucency ≥3.5 mm and a chromosomal abnormality in a parent or a sibling. Common aneuploidies were excluded. RESULTS: Diagnostic yield was 15.1% in the entire patient cohort and as high as 20% in fetuses with multiple structural anomalies. The diagnostic yield was lowest in the group with isolated growth retardation. A total of 76 copy number variants (CNVs) were reported from a total of 65 samples, including 16 CNVs associated with a well-described microdeletion/microduplication syndrome, six autosomal trisomies in mosaic form, and three pathogenic single-gene deletions with dominant inheritance and 12 CNVs known to be risk factors for eg developmental delay. CONCLUSION: The diagnostic yield of aCGH was higher than what has previously been reported in less defined patient cohorts. However, the number of CNVs with unclear correlation to the fetal ultrasound findings was still relatively high. The importance of adequate pre- and posttest counseling must therefore be emphasized.

APOE Genotype Disclosure and Lifestyle Advice in a Randomized Intervention Study with Finnish Participants.

BACKGROUND: The APOE ε4 allele is associated with higher risks of cardiovascular diseases and Alzheimer disease than ε3 and ε2. OBJECTIVES: We studied the effectiveness of dietary and lifestyle guidance and personal genetic risk information [ε4 carrier (ε4+); ε4 noncarrier (ε4-)] as motivators for a healthier lifestyle. METHODS: A total of 188 healthy Finnish volunteers (82.4% women; mean ± SD age: 51.0 ± 5.6 y; BMI: 26.0 ± 3.6 kg/m2; total cholesterol: 5.2 ± 0.9 mmol/L) participated in our randomized intervention study. The participants were genotyped for APOE and divided into intervention (INT; INTε4+, n = 33; INTε4-, n = 57) and control groups (CTRL; CTRLε4+, n = 36; CTRLε4-, n = 62). Blood samples, measured observations, and questionnaire data were obtained at baseline and at 1 and 1.5 y. INT participants received their ε4 carrier status at baseline. Monthly Internet-based guidance based on the Finnish Dietary guidelines was provided for all. RESULTS: The proportion of SFAs in plasma over time fluctuated less in INTε4+ than in the other groups (P-interaction < 0.05; primary outcome). The lifestyle guidance increased vegetable consumption from 3.5 to 3.6 portions/d, improved the dietary fat quality score by 5.3%, increased the plasma n-3 (ω-3) FA proportion by 7.3%, and decreased the consumption of high-fat/high-sugar foods from 7.3 to 6.5 portions/wk and total- and LDL-cholesterol concentrations by 4.3% and 6.1%, respectively, in the entire participant population (P < 0.05; secondary outcome). Compared with the ε4- participants, ε4+ participants had 2.4% higher plasma n-6 (ω-6) FA, lower C-peptide (3.9 compared with 4.2 nmol/L × h) and sensitive C-reactive protein values, and decreased plasma malondialdehyde concentrations over time (P < 0.05; secondary outcome). CONCLUSIONS: Lifestyle guidance given to healthy Finnish participants yielded small but beneficial changes. The INTε4+ group did not seem markedly more responsive to the guidance than the other groups.This trial was registered at clinicaltrials.gov as NCT03794141.

A Long-Term Follow-Up Study on Disclosing Genetic Risk Information (APOE) to Promote Healthy Lifestyles in Finland.

AIM: This observational follow-up study was designed to assess the long-term behavioural and clinical effects of receiving personal genetic risk information. The information disclosed was the carrier status of the apolipoprotein E (APOE)alleles, which differentially contribute to the genetic risk for cardiovascular disease (CVD) and Alzheimer's disease. METHODS: This study forms a continuum with a previous 1-year intervention (2010-2011) monitoring the effects of disclosing the carrier status of the APOE ε4risk allele. The follow-up measurements, performed 5.5 years post-intervention, included clinical measurements (blood values and anthropomorphic parameters) and questionnaires (psychological and behavioural factors). The participants were healthy adult volunteers, aged 26-73 years (n = 70) who had participated in the previous intervention, and received their APOE allele status either at the beginning (former test group) or the end of the intervention (former control group). RESULTS: Personal genetic risk information resulted in a moderate health-conscious change in diet and had a slight positive long-term effect on clinical factors, particularly the serum lipids. These improvements were subsequent to the disclosure of genetic information and occurred mainly in the APOE ε4-positive members of the former control group, that is, those who were at increased genetic risk for CVD but had not been informed of their status before the end of the intervention. In contrast, changes in the values and health behaviour of the APOE ε4-positive individuals in the former test group, who had already changed their health behaviour during the previous intervention as a result of being informed of their carrier status, varied more: some continued to improve, some remained at their previously improved level, and some relapsed slightly. Both groups had nonetheless displayed an improvement immediately subsequent to the disclosure of their personal genetic risk. CONCLUSION: Receiving information on increased personal genetic risk (carrier status of APOE ε4)for CVD provided the motivation for improvements in health behaviour. The resulting changes, while modest, in most cases remained visible even after a number of years.

A homozygous FANCM mutation underlies a familial case of non-syndromic primary ovarian insufficiency.

Primary Ovarian Insufficiency (POI) affects ~1% of women under forty. Exome sequencing of two Finnish sisters with non-syndromic POI revealed a homozygous mutation in FANCM, leading to a truncated protein (p.Gln1701*). FANCM is a DNA-damage response gene whose heterozygous mutations predispose to breast cancer. Compared to the mother's cells, the patients' lymphocytes displayed higher levels of basal and mitomycin C (MMC)-induced chromosomal abnormalities. Their lymphoblasts were hypersensitive to MMC and MMC-induced monoubiquitination of FANCD2 was impaired. Genetic complementation of patient's cells with wild-type FANCM improved their resistance to MMC re-establishing FANCD2 monoubiquitination. FANCM was more strongly expressed in human fetal germ cells than in somatic cells. FANCM protein was preferentially expressed along the chromosomes in pachytene cells, which undergo meiotic recombination. This mutation may provoke meiotic defects leading to a depleted follicular stock, as in Fancm-/- mice. Our findings document the first Mendelian phenotype due to a biallelic FANCM mutation.

Novel COL1A1 Mutation c.3290G>T Associated With Severe Form of Osteogenesis Imperfecta in a Fetus.

Osteogenesis imperfecta is a genetically and clinically heterogenous group of skeletal dysplasias characterized by bone fragility. Its severity ranges from nearly asymptomatic individuals to perinatal lethality. The majority of cases are caused by mutations in either the COL1A1 or the COL1A2 gene coding for alpha 1 and alpha 2 chains of collagen type 1, respectively, and a large number of pathogenic variants of these genes has been identified. We describe a novel COL1A1 mutation associated with prenatally diagnosed severe form of osteogenesis imperfecta.

Affected female carriers of MTM1 mutations display a wide spectrum of clinical and pathological involvement: delineating diagnostic clues.

X-linked myotubular myopathy (XLMTM), a severe congenital myopathy, is caused by mutations in the MTM1 gene located on the X chromosome. A majority of affected males die in the early postnatal period, whereas female carriers are believed to be usually asymptomatic. Nevertheless, several affected females have been reported. To assess the phenotypic and pathological spectra of carrier females and to delineate diagnostic clues, we characterized 17 new unrelated affected females and performed a detailed comparison with previously reported cases at the clinical, muscle imaging, histological, ultrastructural and molecular levels. Taken together, the analysis of this large cohort of 43 cases highlights a wide spectrum of clinical severity ranging from severe neonatal and generalized weakness, similar to XLMTM male, to milder adult forms. Several females show a decline in respiratory function. Asymmetric weakness is a noteworthy frequent specific feature potentially correlated to an increased prevalence of highly skewed X inactivation. Asymmetry of growth was also noted. Other diagnostic clues include facial weakness, ptosis and ophthalmoplegia, skeletal and joint abnormalities, and histopathological signs that are hallmarks of centronuclear myopathy such as centralized nuclei and necklace fibers. The histopathological findings also demonstrate a general disorganization of muscle structure in addition to these specific hallmarks. Thus, MTM1 mutations in carrier females define a specific myopathy, which may be independent of the presence of an XLMTM male in the family. As several of the reported affected females carry large heterozygous MTM1 deletions not detectable by Sanger sequencing, and as milder phenotypes present as adult-onset limb-girdle myopathy, the prevalence of this myopathy is likely to be greatly underestimated. This report should aid diagnosis and thus the clinical management and genetic counseling of MTM1 carrier females. Furthermore, the clinical and pathological history of this cohort may be useful for therapeutic projects in males with XLMTM, as it illustrates the spectrum of possible evolution of the disease in patients surviving long term.

Correction: B-Cell Responses to Human Bocaviruses 1-4: New Insights from a Childhood Follow-Up Study.

[This corrects the article DOI: 10.1371/journal.pone.0139096.].

Inhaled Sargramostim Induces Resolution of Pulmonary Alveolar Proteinosis in Lysinuric Protein Intolerance.

Pulmonary alveolar proteinosis (PAP) is a potentially fatal complication of lysinuric protein intolerance (LPI), an inherited disorder of cationic amino acid transport. The patients often present with mild respiratory symptoms, which may rapidly progress to acute respiratory failure responding poorly to conventional treatment with steroids and bronchoalveolar lavations (BALs). The pathogenesis of PAP in LPI is still largely unclear. In previous studies, we have shown disturbances in the function and activity of alveolar macrophages of these patients, suggesting that increasing the activity and the number of macrophages by recombinant human GM-CSF (rhuGM-CSF) might be beneficial in this patient group.Two LPI patients with complicated PAP were treated with experimental inhaled rhuGM-CSF (sargramostim) after poor response to maximal conventional therapy. BAL fluid and cell samples from one patient were studied with light microscopy and transmission electron microscopy.Excellent response to therapy was observed in patient 1 with no compliance problems or side effects. Macrophages with myelin figure-like structures were seen in her BAL sample. Slight improvement of the pulmonary function was evident also in patient 2, but the role of sargramostim could not be properly evaluated due to the complicated clinical situation.In conclusion, inhaled rhuGM-CSF might be of benefit in patients with LPI-associated PAP.

Imbalance of plasma amino acids, metabolites and lipids in patients with lysinuric protein intolerance (LPI).

BACKGROUND: Lysinuric protein intolerance (LPI [MIM 222700]) is an aminoaciduria with defective transport of cationic amino acids in epithelial cells in the small intestine and proximal kidney tubules due to mutations in the SLC7A7 gene. LPI is characterized by protein malnutrition, failure to thrive and hyperammonemia. Many patients also suffer from combined hyperlipidemia and chronic kidney disease (CKD) with an unknown etiology. METHODS: Here, we studied the plasma metabolomes of the Finnish LPI patients (n=26) and healthy control individuals (n=19) using a targeted platform for analysis of amino acids as well as two analytical platforms with comprehensive coverage of molecular lipids and polar metabolites. RESULTS: Our results demonstrated that LPI patients have a dichotomy of amino acid profiles, with both decreased essential and increased non-essential amino acids. Altered levels of metabolites participating in pathways such as sugar, energy, amino acid and lipid metabolism were observed. Furthermore, of these metabolites, myo-inositol, threonic acid, 2,5-furandicarboxylic acid, galactaric acid, 4-hydroxyphenylacetic acid, indole-3-acetic acid and beta-aminoisobutyric acid associated significantly (P<0.001) with the CKD status. Lipid analysis showed reduced levels of phosphatidylcholines and elevated levels of triacylglycerols, of which long-chain triacylglycerols associated (P<0.01) with CKD. CONCLUSIONS: This study revealed an amino acid imbalance affecting the basic cellular metabolism, disturbances in plasma lipid composition suggesting hepatic steatosis and fibrosis and novel metabolites correlating with CKD in LPI. In addition, the CKD-associated metabolite profile along with increased nitrite plasma levels suggests that LPI may be characterized by increased oxidative stress and apoptosis, altered microbial metabolism in the intestine and uremic toxicity.

Urine Beta2-Microglobulin Is an Early Marker of Renal Involvement in LPI.

OBJECTIVE: Lysinuric protein intolerance (LPI) is a rare autosomal recessive disorder affecting the transport of cationic amino acids. It has previously been shown that approximately one third of the Finnish LPI patients have impaired renal function. The aim of this study was to analyse in detail urine beta2-microglobulin values, renal dysfunction, oral L-citrulline doses and plasma citrulline concentrations in Finnish LPI patients. METHODS AND RESULTS: Of the 41 Finnish LPI patients, 56% had proteinuria and 53% hematuria. Mean plasma creatinine concentration was elevated in 48%, serum cystatin C in 62%, and urine beta2-microglobulin in 90% of the patients. Seventeen per cent of the patients developed ESRD, and five of them received a kidney transplant. L-citrulline doses and fasting plasma citrulline concentrations were similar in adult LPI patients with decreased and normal GFR (mean ± SD 79.5 ± 29.2 vs. 82.4 ± 21.9 mg/kg/day, P = 0.619, and 80.3 ± 20.1 vs. 64.8 ± 23.0 µmol/l, P = 0.362, respectively). CONCLUSIONS: Urine beta2-microglobulin is a sensitive early marker of renal involvement, and it should be monitored regularly in LPI patients. Weight-based oral L-citrulline doses and plasma citrulline concentrations were not associated with renal function. LPI patients with ESRD were successfully treated with dialysis and kidney transplantation.

B-Cell Responses to Human Bocaviruses 1-4: New Insights from a Childhood Follow-Up Study.

Human bocaviruses (HBoVs) 1-4 are recently discovered, antigenically similar parvoviruses. We examined the hypothesis that the antigenic similarity of these viruses could give rise to clinically and diagnostically important immunological interactions. IgG and IgM EIAs as well as qPCR were used to study ~2000 sera collected from infancy to early adolescence at 3-6-month intervals from 109 children whose symptoms were recorded. We found that HBoV1-4-specific seroprevalences at age 6 years were 80%, 48%, 10%, and 0%, respectively. HBoV1 infections resulted in significantly weaker IgG responses among children who had pre-existing HBoV2 IgG, and vice versa. Furthermore, we documented a complete absence of virus type-specific immune responses in six viremic children who had pre-existing IgG for another bocavirus, indicating that not all HBoV infections can be diagnosed serologically. Our results strongly indicate that interactions between consecutive HBoV infections affect HBoV immunity via a phenomenon called "original antigenic sin", cross-protection, or both; however, without evident clinical consequences but with important ramifications for the serodiagnosis of HBoV infections. Serological data is likely to underestimate human exposure to these viruses.

Dysfunction in macrophage toll-like receptor signaling caused by an inborn error of cationic amino acid transport.

Amino acids, especially arginine, are vital for the well-being and activity of immune cells, and disruption of amino acid balance may weaken immunity and predispose to infectious and autoimmune diseases. We present here a model of an inborn aminoaciduria, lysinuric protein intolerance (LPI), in which a single mutation in y(+)LAT1 cationic amino acid transporter gene SLC7A7 leads to a multisystem disease characterized by immunological complications, life-threatening pulmonary alveolar proteinosis and nephropathy. Macrophages are suggested to play a central role in LPI in the development of these severe secondary symptoms. We thus studied the effect of the Finnish y(+)LAT1 mutation on monocyte-derived macrophages where toll-like receptors (TLRs) act as the key molecules in innate immune response against external pathogens. The function of LPI patient and control macrophage TLR signaling was examined by stimulating the TLR2/1, TLR4 and TLR9 pathways with their associated pathogen-associated molecular patterns. Downregulation in expression of TLR9, IRF7, IRF3 and IFNB1 and in secretion of IFN-α was detected, suggesting an impaired response to TLR9 stimulation. In addition, secretion of TNF-α, IL-12 and IL-1RA by TLR2/1 stimulation and IL-12 and IL-1RA by TLR4 stimulation was increased in the LPI patients. LPI macrophages secreted significantly less nitric oxide than control macrophages, whereas plasma concentrations of inflammatory chemokines CXCL8, CXCL9 and CXCL10 were elevated in the LPI patients. In conclusion, our results strengthen the relevance of macrophages in the pathogenesis of LPI and, furthermore, suggest that cationic amino acid transport plays an important role in the regulation of innate immune responses.