The characterization and comorbidities of heterozygous Bardet-Biedl syndrome carriers.

Introduction: Bardet-Biedl syndrome (BBS) is a rare autosomal recessive disorder with clinical features of retinal dystrophy, obesity, postaxial polydactyly, renal anomalies, learning disabilities, hypogonadism, and genitourinary abnormalities. Nevertheless, previous studies on the phenotypic traits of BBS heterozygous carriers have generated inconclusive results. The aim of our study was to investigate the impact of BBS heterozygosity on carriers when compared to non-carriers within the Taiwanese population. Materials and Methods: This study follows a hospital-based case-control design. We employed the Taiwan Biobank version 2 (TWBv2) array to identify three specific loci associated with BBS (rs773862084, rs567573386, and rs199910690). In total, 716 patients were included in the case group, and they were compared to a control group of 2,864 patients who lacked BBS alleles. The control group was selected through gender and age matching at a ratio of 1:4. The association between BBS-related loci and comorbidity was assessed using logistic regression models. Results: We found that BBS heterozygous carriers exhibited a significant association with elevated BMI levels, especially the variant rs199910690 in MKS1 (p=0.0037). The prevalence of comorbidities in the carriers' group was not higher than that in the non-carriers' group. Besides, the average values of the biochemistry data showed no significant differences, except for creatinine level. Furthermore, we conducted a BMI-based analysis to identify specific risk factors for chronic kidney disease (CKD). Our findings revealed that individuals carrying the CA/AA genotype of the BBS2 rs773862084 variant or the CT/TT genotype of the MKS1 rs199910690 variant showed a reduced risk of developing CKD, irrespective of their BMI levels. When stratified by BMI level, obese males with the MKS1 rs199910690 variant and obese females with the BBS2 rs773862084 variant exhibited a negative association with CKD development. Conclusion: We found that aside from the association with overweight and obesity, heterozygous BBS mutations did not appear to increase the predisposition of individuals to comorbidities and metabolic diseases. To gain a more comprehensive understanding of the genetic susceptibility associated with Bardet-Biedl Syndrome (BBS), further research is warranted.

Spectrum of PHEX Mutations and FGF23 Profiles in a Taiwanese Cohort With X-Linked Hypophosphatemia Including 102 Patients.

BACKGROUND/AIM: X-linked hypophosphatemia (XLH), the most common form of hereditary rickets, results from loss-of-function mutations in the phosphate-regulating PHEX gene. Elevated fibroblast growth factor 23 (FGF23) contributes to hypophosphatemia in XLH. This study aimed to characterize PHEX variants and serum FGF23 profiles in Taiwanese patients with XLH. PATIENTS AND METHODS: We retrospectively reviewed the records of 102 patients clinically suspected of having hypophosphatemic rickets from 2006 to 2022. Serum intact Fibroblast growth factor-23 (iFGF23) levels were measured on clinic visit days. PHEX mutations were identified using Sanger sequencing, and negative cases were analyzed using whole-exome sequencing. RESULTS: The majority (92.1%) of patients exhibited elevated FGF23 compared with normal individuals. Among 102 patients, 44 distinct PHEX mutations were identified. Several mutations recurred in multiple unrelated Taiwanese families. We discovered a high frequency of novel PHEX mutations and identified variants associated with extreme FGF23 elevation and tumorigenesis. CONCLUSION: Our findings revealed the PHEX genotypic variants and FGF23 levels in Taiwanese patients with XLH. These results are crucial given the recent approval of burosumab, a monoclonal FGF23 antibody, for XLH therapy. This study provides key insights into the clinical management of XLH in Taiwan.

Sex-specific genetic variants associated with adult-onset inguinal hernia in a Taiwanese population.

Introduction: Inguinal hernia repair is one of the most common surgeries worldwide. However, there is limited information on its underlying genetic mechanism. Studies on the genetic factors related to inguinal hernia in Han Chinese are lacking. Therefore, we aimed to conduct a hospital-based study to assess the genetic factors and comorbidities underlying inguinal hernia in Taiwan. Materials and Methods: This was a retrospective case-control study. Utilizing data from the Taiwan Precision Medicine Initiative, we identified 1000 patients with inguinal hernia and 10,021 matched controls without inguinal hernia between June 2019 and June 2020. Four susceptibility loci (rs2009262, rs13091322, rs6991952, and rs3809060) associated with inguinal hernia were genotyped by the Taiwan Biobank version 2 (TWBv2) array. Inguinal hernia, surgery types, and comorbidities were obtained from the electronic health records of Taichung Veterans General Hospital. Results: Adult-onset inguinal hernia was associated with WT1 rs3809060 GT/TT genotype in males and EFEMP1 rs2009262 TC/CC genotype in females. In addition, we identified sex-specific risk factors associated with inguinal hernia; benign prostatic hyperplasia in males (OR: 3.19, 95% CI: 2.73 - 3.73, p< 0.001), chronic obstructive pulmonary disease in females (OR: 2.34, 95% CI: 1.33 - 4.11, p = 0.003) and overweight, defined by body mass index ≧24 kg/m2 (OR: 0.75, 95% CI: 0.65 - 0.86, p<0.001 in males, and OR: 0.60, 95% CI:0.37 - 0.98, p = 0.042 in females), were inversely associated with inguinal hernia. After stratifying BMI, overweight males with EFEMP1 rs2009262 TC/CC genotype exhibited a higher risk of inguinal hernia (OR: 1.31, 95% CI: 1.07 - 1.61, p = 0.01). Additionally, rs3809060 was specifically associated with male patients with direct-type inguinal hernia (OR: 1.62, 95% CI: 1.19 - 2.22, p = 0.002). Conclusion: Genetic susceptibility appears to participate in the pathogenesis of inguinal hernia in the Taiwanese population in a sex-specific manner. Future studies are needed to illuminate the complex interplay between heredity and comorbidities.

Congenital generalized lipodystrophy in Taiwan.

BACKGROUND: Congenital generalized lipodystrophy (CGL) is a rare disorder characterized by scarce adipose tissue. This disease is distributed worldwide, but little is known about these patients in the Chinese population. Here, we delineate the phenotype and prognosis of CGL in our cohort. METHODS: Patients diagnosed with CGL from 8 medical centers were reviewed. The initial presentation, laboratory findings, and molecular testing were retrospectively analyzed. RESULTS: A total of 16 patients were analyzed, and the current median age was 3.5 years (range, 9 months-17.5 years). In all patients, molecular results confirmed BSCL2 mutation. c.782dupG (p.Ile262Hisfs*12) was the most common genotype identified. All patients had triangular faces and muscular hypertrophy. In addition, 75% presented with hepatomegaly, 19% had cardiomegaly, and 44% exhibited acanthosis nigricans. Developmental delay was noted in 5 out of 9 patients (56%) with a median developmental quotient (DQ)/intelligence quotient (IQ) of 61. Thirteen patients (81.3%) had high triglyceride levels. Eight patients received leptin analysis, and 7 of them (88%) had low leptin levels. One patient exclusively received a lipid-lowering drug, 4 patients were exclusively placed on a fat-restricted diet, 5 patients were administered combination therapy, and 5 patients received no treatment. Three patients (19%) who developed diabetes mellitus received both oral hypoglycemic agents and insulin. Three patients (19%) experienced loss of ambulation and died prematurely. CONCLUSION: Our findings highlight the uniqueness of the genotype and phenotype in our cohort. Further long-term surveillance for comorbidities is necessary for early detection and management of these patients.

Chronic Iron Overload Results in Impaired Bacterial Killing of THP-1 Derived Macrophage through the Inhibition of Lysosomal Acidification.

Iron is essential for living organisms and the disturbance of iron homeostasis is associated with altered immune function. Additionally, bacterial infections can cause major complications in instances of chronic iron overload, such as patients with transfusion-dependent thalassemia. Monocytes and macrophages play important roles in maintaining systemic iron homoeostasis and in defense against invading pathogens. However, the effect of iron overload on the function of monocytes and macrophages is unclear. We elucidated the effects of chronic iron overload on human monocytic cell line (THP-1) and THP-1 derived macrophages (TDM) by continuously exposing them to high levels of iron (100 µM) to create I-THP-1 and I-TDM, respectively. Our results show that iron overload did not affect morphology or granularity of I-THP-1, but increased the granularity of I-TDM. Bactericidal assays for non-pathogenic E. coli DH5α, JM109 and pathogenic P. aeruginosa all revealed decreased efficiency with increasing iron concentration in I-TDM. The impaired P. aeruginosa killing ability of human primary monocyte derived macrophages (hMDM) was also found when cells are cultured in iron contained medium. Further studies on the bactericidal activity of I-TDM revealed lysosomal dysfunction associated with the inhibition of lysosomal acidification resulting in increasing lysosomal pH, the impairment of post-translational processing of cathepsins (especially cathepsin D), and decreased autophagic flux. These findings may explain the impaired innate immunity of thalassemic patients with chronic iron overload, suggesting the manipulation of lysosomal function as a novel therapeutic approach.

Long-term galsulfase enzyme replacement therapy in Taiwanese mucopolysaccharidosis VI patients: A case series.

BACKGROUND: Information regarding the long-term outcome of enzyme replacement therapy (ERT) with recombinant human N-acetylgalactosamine 4-sulfatase (rhASB, galsulfase, Naglazyme®, BioMarin Pharmaceutical Inc.) for Taiwanese patients with mucopolysaccharidosis (MPS) VI is limited. METHODS: Nine Taiwanese patients with MPS VI (4 males and 5 females; age range, 1.4 to 21.1 years) treated with weekly intravenous infusions of galsulfase (1.0 mg/kg) in 5 medical centers in Taiwan were reviewed. A set of biochemical and clinical assessments were evaluated annually. RESULTS: After 6.2 to 11.2 years of galsulfase treatment, 6 patients experienced improvement over baseline in the 6-minute walk test by a mean of 150 m (59% change over time), and 3 patients also increased the 3-minute stair climb test by a mean of 60 steps (46%). In a manual dexterity test, 3 patients decreased the time required to pick up 10 coins and put the coins into a cup by 15 s (33%). Shoulder range of motion in all 9 patients improved, and Joint Pain and Stiffness Questionnaire scores improved by 0.42 points (21%). Four patients showed improved pulmonary function. Five patients had positive effects on cardiac-wall diameters. Four patients had improved cardiac diastolic function. Liver and spleen sizes as measured by abdominal ultrasonography remained the same or decreased in all 9 patients. However, the severity degree of valvular stenosis or regurgitation did not show improvement despite ERT. A mean overall 69% decrease in urinary glycosaminoglycan (GAG) excretion indicated a satisfactory biomarker response. CONCLUSIONS: Long-term ERT was beneficial and safe for Taiwanese patients with MPS VI. This treatment reduced urinary GAG and had positive effects on a wide range of clinical functional assessments including endurance, mobility, joint function, pulmonary function, liver and spleen size, cardiac hypertrophy and diastolic dysfunction.

Deletion of exon 4 in the N-acetylgalactosamine-4-sulfatase gene in a Taiwanese patient with mucopolysaccharidosis type VI.

Mucopolysaccharidosis type VI (MPS VI) is a rare autosomal recessive disorder caused by a deficiency of N-acetylgalactosamine-4-sulfatase (ARSB), one of the enzymes required for the degradation of dermatan sulfate (DS). Accumulation of DS in connective tissue causes growth failure, resulting in short stature. Here, we observed a 5-year-old girl who was the only one affected member of her family and who presented with an exaggerated, convex curvature of the back at the age of one year. Abnormal excretion of DS in the urine and extremely low leukocyte ARSB activity were noted. The patient was suspected to have MPS VI. Direct DNA sequencing indicated that there was no mutation in the coding region of ARSB. However, RT-PCR analysis of RNA prepared from blood samples indicated the deletion of the entire exon 4. Further analysis of the genomic DNA by quantitative PCR confirmed a homozygous deletion of exon 4, an unusual intragenic deletion in ARSB. The deletion led to a truncated protein that lacks most of the catalytic domain. The patient received recombinant human ARSB as enzyme replacement therapy (ERT) at an early stage (2 years), and responded positively in terms of skeletal development and other developmental milestones. The early identification of type VI MPS patients and subsequent treatment with ERT may be beneficial for the clinical outcome of MPS VI patients. In addition, detailed gene analysis may enhance the ability to provide genetic counseling to families of patients affected by MPS VI.

Two frequent mutations associated with the classic form of propionic acidemia in Taiwan.

Propionyl-CoA carboxylase (PCC) is involved in the catabolism of branched chain amino acids, odd-numbered fatty acids, cholesterol, and other metabolites. PCC consists of two subunits, α and ß, encoded by the PCCA and PCCB genes, respectively. Mutations in the PCCA or PCCB subunit gene may lead to propionic acidemia. In this study, we performed mutation analysis on ten propionic acidemia patients from eight unrelated and nonconsanguineous families in Taiwan. Two PCCA mutations, c.229C→T (p.R77W) and c.1262A→C (p.Q421P), were identified in a PCCA-deficient patient. Six mutations in the PCCB gene, including c.-4156_183+3713del, c.580T→C (p.S194P), c.838dup (p.L280Pfs 11), c.1301C→T (p.A434V), c.1316A→G (P.Y439C), and c.1534C→T (p.R512C), were identified in seven PCCB-deficient families. The c.-4156_183+3713del mutation is the first known large deletion that affects the PCCB gene functions. Furthermore, the c.1301C→T and c.-4156_183+3713del mutations in the PCCB gene have not been reported previously. Clinical features demonstrated that these two frequent mutations are associated with low enzyme activity and a classic propionic acidemia phenotype.

Natural history and clinical assessment of Taiwanese patients with mucopolysaccharidosis IVA.

BACKGROUND: Mucopolysaccharidosis IVA (MPS IVA) is a rare lysosomal storage disorder caused by N-acetylgalactosamine-6-sulfatase deficiency, which catalyzes a step in the catabolism of glycosaminoglycans, keratan sulfate and chondroitin-6-sulfate. This disease has a variable age of onset and rate of progression. METHODS: A retrospective analysis of medical records of 24 patients with MPS IVA (11 males, 13 females; current mean age ± SD, 12.6 ± 6.6 years; age range, 1.4-29.4 years) seen at 6 medical centers in Taiwan from January 1996 through June 2013 was performed. RESULTS: Mean ages of onset of symptoms and confirmed diagnosis were 2.0 ± 1.6 and 5.7 ± 4.5 years, respectively. The most prevalent clinical manifestations were kyphosis (100%), pectus carinatum (96%), abnormal gait (93%), striking short trunk dwarfism (92%), genu valgum (92%), and valvular heart disease (91%). Eight patients (33%) experienced at least one surgical procedure with the most common being ear tube insertion (25%), adenoidectomy (17%), tonsillectomy (13%), supraglottoplasty (13%), spinal decompression (13%), and spinal fusion (13%). The most prevalent cardiac valve abnormalities were aortic stenosis (45%) and mitral regurgitation (45%). At the time of the study, 8 out of 24 patients (33%) have died at the mean age of 17.2 ± 7.7 years. CONCLUSIONS: An understanding of the natural history involved in MPS IVA may allow early diagnosis of the disease. All affected Taiwanese patients experienced significant functional limitations. Adequate evaluations and timely management may improve clinical outcomes and quality of life.

Plasma globotriaosylsphingosine (lysoGb3) could be a biomarker for Fabry disease with a Chinese hotspot late-onset mutation (IVS4+919G>A).

BACKGROUND: Previous studies revealed a high incidence of late-onset Fabry disease mutation, IVS4+919G>A, in Taiwan. However, the natural course is largely unclear and suitable biomarkers for monitoring disease progress are unavailable. METHODS AND RESULTS: Patients carrying IVS4+919G>A or classical Fabry mutations were enrolled in this study. The subjects ranged from newborn to eighty year old adults. Plasma globotriaosylceramide (Gb3) and globotriaosylsphingosine (lysoGb3) were measured by LC-MS/MS in subjects to evaluate the sensitivity of these two biomarkers. All adult males and symptomatic females could be distinguished from healthy controls by an elevated plasma lysoGb3 level. The lysoGb3 level was also related to the left ventricular mass considering gender and age (p<0.01). Moreover, approximately 70% of male and 45% of female newborns already had an elevated plasma lysoGb3 level which increased gradually as the subjects got older (p<0.01). CONCLUSIONS: Plasma lysoGb3 is a more sensitive and reliable biomarker than plasma Gb3. LysoGb3 also correlated with age and left ventricular mass index in Fabry patients with IVS4+919G>A mutation. Because lots of infants with the IVS4+919G>A mutation already had elevated lysoGb3 levels at birth, that indicates that the development of hypertrophic cardiomyopathy may require a long and insidious course after lysoGb3 accumulation.

Unilateral agenesis of the internal carotid artery in CHARGE syndrome.

CHARGE syndrome is a multisystemic disorder comprising colobomas, heart defects, choanal atresia, retarded growth and development, genital hypoplasia, ear anomalies and deafness. The CHD7 gene on chromosome 8q12.1 was recently shown to be a major gene involved in the etiology of this syndrome. We describe a girl with CHARGE syndrome who had a novel mutation of CHD7 associated with agenesis of the left internal carotid artery. She had presented with recurrent episodes of photophobia and vomiting since the age of 6 years. Since her symptoms were well controlled by cyproheptadine, migraine-like attacks were considered. CHD7 molecular confirmation in this patient provides further evidence to support the occurrence of a vascular anomaly suggested from animal models of CHARGE syndrome with molecular delineation. We report this case to emphasize the importance of neurologic signs of photophobia and to highlight the broad clinical variability in this pleiotropic disorder.

Enzyme replacement therapy for mucopolysaccharidosis VI--experience in Taiwan.

Information regarding the clinical outcome of enzyme replacement therapy (ERT) with recombinant human N-acetylgalactosamine 4-sulfatase (rhASB) for mucopolysaccharidosis (MPS) VI in Asian patients is limited. We reviewed nine Taiwanese patients with MPS VI (four males and five females; age range 1.4-21.1 years) treated with weekly intravenous infusions of rhASB (1.0 mg/kg) for at least 2 years. We assessed the biochemical and clinical response every 3 months. After 2 years of treatment, seven patients experienced improvement over baseline in the 6-min walk by a mean of 69.3 m (27.3%), and seven also increased the 3-min stair climb by a mean of 47 steps (35.7%). Shoulder range of motion in all patients improved, and Joint Pain and Stiffness Questionnaire scores improved by 0.597 points (30.5%). Four patients had improved pulmonary function [forced expiratory volume in 1 s increased by 0.130 L (26.3%) and forced vital capacity by 0.148 L (27.6%)]. The respiratory disturbance index decreased in the four patients who underwent polysomnography. A mean overall 51% decrease in urinary glycosaminoglycan excretion indicated a satisfactory biochemical response. ERT was well tolerated by all patients. This treatment is thus beneficial and appears to be safe for treatment of MPS VI in Taiwanese patients.

Complex rearrangements between chromosomes 6, 10, and 11 with multiple deletions at breakpoints.

Here we report on a girl with minor facial anomalies, cleft palate, seizures, microcephaly, psychomotor retardation, and a congenital heart defect. Complex of cytogenetic methods [GTG-banding, spectral karyotyping (SKY), fluorescence in situ hybridization (FISH), multicolor banding (mBAND), and comparative genomic hybridization (array CGH)] showed complex chromosomal rearrangements (CCRs) involving chromosomes 6, 10, and 11 and 4 deletions at the breakpoints. Her father had an unrelated translocation between chromosomes 3 and 16, suggesting the possibility of an autosomal dominant trait that predisposes to complex synapses and recombination between multiple chromosomes during meiosis. This study demonstrates the power of combining available chromosome analysis technologies in resolving CCR.

Cytotoxicity of calotropin is through caspase activation and downregulation of anti-apoptotic proteins in K562 cells.

Calotropin is one of cardenolides isolated from milkweed used for medicinal purposes in many Asian countries. Whereas calotropin possesses cytotoxicity against several cancer cells, the mechanisms of action remain unclear. We set out to evaluate the cytotoxic mechanism of calotropin on human chronic myeloid leukemia K562 cells. Calotropin inhibited the growth of K562 cells in a time- and dose-dependent manner by G(2)/M phase arrest. It upregulated the expression of p27 leading to this arrest by downregulating the G2/M regulatory proteins, cyclins A and B, and by upregulating the cdk inhibitor, p27. Furthermore, it downregulated anti-apoptotic signaling (XIAP and survivin) and survival pathways (p-Akt and NFkappaB), leading to caspase-3 activation which resulted in the induction of apoptosis. In all, calotropin exerted its anticancer activity on K562 cells by modulating the pro-survival signaling that leads to induction of apoptosis.

A recurrent ITGA9 missense mutation in human fetuses with severe chylothorax: possible correlation with poor response to fetal therapy.

OBJECTIVES: To assess the possible correlations between the reported candidate genes (VEGFR3, FOXC2, ITGA9 and ITGB1) and the clinical response in fetuses with severe congenital chylothorax (CC) treated by prenatal OK-432 pleurodesis. METHODS: We studied 12 unrelated fetuses with severe CC, receiving fetal therapy by OK-432 pleurodesis. Genotyping of the candidate genes and the clinical parameters of these 12 fetuses were investigated. Additional 96 control individuals were enrolled to evaluate the possible polymorphisms at these candidate genes in population. RESULTS: A recurrent heterozygous missense mutation (c.1210G>A, p.G404S) was identified in the beta-propeller domain of integrin alpha(9) (ITGA9), a cell adhesion receptor, in four of the five fetuses who failed to respond to the OK-432 treatment. Computer modeling of the p.G404S substitution supported the deleterious nature of this mutation. Family analyses in three affected fetuses demonstrated that the heterozygous mutant allele is of parental origin, suggesting an autosomal recessive inheritance of this genetic defect. CONCLUSIONS: To the best of our knowledge, this is the first insight into the possible link between ITGA9 and CC in human fetuses. The identification of pathogenetic mutations and their possible link to the clinical responses of particular treatments may contribute to better pregnancy counseling and management.