Disclaimer: These ACMG Standards and Guidelines are developed primarily as an educational resource for clinical laboratory geneticists to help them provide quality clinical laboratory genetic ...services. Adherence to these Standards and Guidelines is voluntary and does not necessarily assure a successful medical outcome. These Standards and Guidelines should not be considered inclusive of all proper procedures and tests or exclusive of other procedures and tests that are reasonably directed to obtaining the same results. In determining the propriety of any specific procedure or test, the clinical laboratory geneticist should apply his or her own professional judgment to the specific circumstances presented by the individual patient or specimen. Clinical laboratory geneticists are encouraged to document in the patient’s record the rationale for the use of a particular procedure or test, whether or not it is in conformance with these Standards and Guidelines. They also are advised to take notice of the date any particular guideline was adopted, and to consider other relevant medical and scientific information that becomes available after that date. It also would be prudent to consider whether intellectual property interests may restrict the performance of certain tests and other procedures.
Galactosemias are inherited disorders of galactose metabolism due to deficiency in one of the three enzymes involved in the Leloir pathway: galactose-1-phosphate uridyltransferase, galactokinase, and uridine diphosphate (UDP)-galactose-4′-epimerase. Galactose-1-phosphate uridyltransferase deficiency, or classic galactosemia, is the most frequent and the most severe of the three enzyme deficiencies; it is characterized by failure to thrive, liver failure, susceptibility to sepsis, and death, if untreated. Newborn screening for classic galactosemia has been implemented in all of the United States, while screening for galactokinase deficiency and UDP-galactose-4′-epimerase deficiency is not universal. Early identification and treatment of galactosemia leads to improved outcome. This document reviews the laboratory methods and best practices for the diagnosis of galactosemia.
•MT treatment promotes astaxanthin and lipids contents under adverse conditions.•MT treatment enhances H. pluvialis abiotic stress tolerance by regulating ROS.•Metabolomic mechanisms related to MT ...induction were elucidated in H. pluvialis.•The GABA shunt plays a key role in astaxanthin synthesis following treatment with MT.
The effect of melatonin (MT) on the coproduction of astaxanthin and lipids was studied in Haematococcus pluvialis under inductive stress conditions. The contents of astaxanthin and lipids were enhanced by 1.78- and 1.3-fold, respectively. MT treatment upregulated the transcription levels of carotenogenic, lipogenic and antioxidant system-related genes and decreased the levels of abiotic stress-induced reactive oxidative species (ROS). Further metabolomic analysis suggested that the intermediates in glycolysis and TCA cycle facilitate the accumulation of astaxanthin and lipids in algae treated with MT. Meanwhile, MT treatment upregulated the metabolite levels of the γ-aminobutyric acid (GABA) shunt, which might regulate the carbon-nitrogen balance and the antioxidant system. After MT treatment, exogenous linoleic acid, succinate, and GABA further increased the astaxanthin content. This study may help to elucidate the specific responses to MT induction in H. pluvialis and to identify novel biomarkers that may be employed to further promote astaxanthin and lipids coproduction.
We conducted a consented pilot newborn screening (NBS) for Pompe, Gaucher, Niemann-Pick A/B, Fabry, and MPS 1 to assess the suitability of these lysosomal storage disorders (LSDs) for public health ...mandated screening.
At five participating high-birth rate, ethnically diverse New York City hospitals, recruiters discussed the study with postpartum parents and documented verbal consent. Screening on consented samples was performed using multiplexed tandem mass spectrometry. Screen-positive infants underwent confirmatory enzymology, DNA testing, and biomarker quantitation when available. Affected infants are being followed for clinical management and long-term outcome.
Over 4 years, 65,605 infants participated, representing an overall consent rate of 73%. Sixty-nine infants were screen-positive. Twenty-three were confirmed true positives, all of whom were predicted to have late-onset phenotypes. Six of the 69 currently have undetermined disease status.
Our results suggest that NBS for LSDs is much more likely to detect individuals at risk for late-onset disease, similar to results from other NBS programs. This work has demonstrated the feasibility of using a novel consented pilot NBS study design that can be modified to include other disorders under consideration for public health implementation as a means to gather critical evidence for evidence-based NBS practices.
ABSTRACT
Peroxisomes are essential organelles for the specialized oxidation of a wide variety of fatty acids, but they are also able to degrade fatty acids that are typically handled by mitochondria. ...Using a combination of pharmacological inhibition and clustered regularly interspaced short palindromic repeats (CRISPR)‐CRISPR associated protein 9 genome editing technology to simultaneously manipulate peroxisomal and mitochondrial fatty acid β‐oxidation (FAO) in HEK‐293 cells, we identified essential players in the metabolic crosstalk between these organelles. Depletion of carnitine palmitoyltransferase (CPT)2 activity through pharmacological inhibition or knockout (KO) uncovered a significant residual peroxisomal oxidation of lauric and palmitic acid, leading to the production of peroxisomal acylcarnitine intermediates. Generation and analysis of additional single‐ and double‐KO cell lines revealed that the D‐bifunctional protein (HSD17B4) and the peroxisomal ABC transporter ABCD3 are essential in peroxisomal oxidation of lauric and palmitic acid. Our results indicate that peroxisomes not only accept acyl‐CoAs but can also oxidize acylcarnitines in a similar biochemical pathway. By using an Hsd17b4 KO mouse model, we demonstrated that peroxisomes contribute to the plasma acylcarnitine profile after acute inhibition of CPT2, proving in vivo relevance of this pathway. We summarize that peroxisomal FAO is important when mitochondrial FAO is defective or overloaded.—Violante, S., Achetib, N., van Roermund, C. W. T., Hagen, J., Dodatko, T., Vaz, F. M., Waterham, H. R., Chen, H., Baes, M., Yu, C., Argmann, C. A., Houten, S. M. Peroxisomes can oxidize medium‐ and long‐chain fatty acids through a pathway involving ABCD3 and HSD17B4. FASEB J. 33, 4355–4364 (2019). www.fasebj.org
Peroxisomes play an essential role in the β-oxidation of dicarboxylic acids (DCAs), which are metabolites formed upon ω-oxidation of fatty acids. Genetic evidence linking transporters and enzymes to ...specific DCA β-oxidation steps is generally lacking. Moreover, the physiological functions of DCA metabolism remain largely unknown. In this study, we aimed to characterize the DCA β-oxidation pathway in human cells, and to evaluate the biological role of DCA metabolism using mice deficient in the peroxisomal
l
-bifunctional protein (
Ehhadh
KO mice). In vitro experiments using HEK-293 KO cell lines demonstrate that ABCD3 and ACOX1 are essential in DCA β-oxidation, whereas both the bifunctional proteins (EHHADH and HSD17B4) and the thiolases (ACAA1 and SCPx) have overlapping functions and their contribution may depend on expression level. We also show that medium-chain 3-hydroxydicarboxylic aciduria is a prominent feature of EHHADH deficiency in mice most notably upon inhibition of mitochondrial fatty acid oxidation. Using stable isotope tracing methodology, we confirmed that products of peroxisomal DCA β-oxidation can be transported to mitochondria for further metabolism. Finally, we show that, in liver,
Ehhadh
KO mice have increased mRNA and protein expression of cholesterol biosynthesis enzymes with decreased (in females) or similar (in males) rate of cholesterol synthesis. We conclude that EHHADH plays an essential role in the metabolism of medium-chain DCAs and postulate that peroxisomal DCA β-oxidation is a regulator of hepatic cholesterol biosynthesis.
The acute hepatic porphyrias are inherited disorders of heme biosynthesis characterized by life-threatening acute neurovisceral attacks. Factors that induce the expression of hepatic 5-aminolevulinic ...acid synthase 1 (ALAS1) result in the accumulation of the neurotoxic porphyrin precursors 5-aminolevulinic acid (ALA) and porphobilinogen (PBG), which recent studies indicate are primarily responsible for the acute attacks. Current treatment of these attacks involves i.v. administration of hemin, but a faster-acting, more effective, and safer therapy is needed. Here, we describe preclinical studies of liver-directed small interfering RNAs (siRNAs) targeting Alas1 (Alas1-siRNAs) in a mouse model of acute intermittent porphyria, the most common acute hepatic porphyria. A single i.v. dose of Alas1-siRNA prevented the phenobarbital-induced biochemical acute attacks for approximately 2 wk. Injection of Alas1-siRNA during an induced acute attack significantly decreased plasma ALA and PBG levels within 8 h, more rapidly and effectively than a single hemin infusion. Alas1-siRNA was well tolerated and a therapeutic dose did not cause hepatic heme deficiency. These studies provide proof-of-concept for the clinical development of RNA interference therapy for the prevention and treatment of the acute attacks of the acute hepatic porphyrias.
Skeletal muscle, comprising approximately 40% of body mass, is a highly complex and heterogeneous tissue serving a multitude of functions in the organism. Non-coding RNAs (ncRNAs) are known to ...participate in skeletal muscle development as critical regulators. However, the regulatory mechanisms of ncRNAs on chicken muscle traits are not well understood. In the present study, we collected the leg muscle from male embryos of Tibetan chicken at embryonic (E) 10 and E18 for RNA sequencing. A total of 6,583 differentially expressed mRNAs (DEMs) including 3,055 down-regulated and 3,528 up-regulated were identified in E18. We identified 695 differentially expressed lncRNAs (DELs) (187 down-regulated and 508 up-regulated) and 1,906 differentially expressed circRNAs (DECs) (1,224 down-regulated and 682 up-regulated) in E18. Among the 130 differentially expressed miRNAs (DEMIs), 59 were up-regulated and 71 were down-regulated in E18. Numerous DEMs and target genes for miRNAs/lncRNAs were significantly enriched in the muscle system process and cell cycle. We constructed a miRNA-gene-pathway network by considering target relationships between genes related to skeletal muscle development and miRNAs. A competing endogenous RNA (ceRNA) network was also constructed by integrating competing relationships between DEMs, DELs, and DECs. Several DELs and DECs were predicted to regulate the ADRA1B, ATP2A2, ATP2B1, CACNA1S, CACNB4, MYLK2, and ROCK2 genes. We discovered the crosstalk between the ncRNAs and their competing mRNAs, which provides insights into ceRNA function and mechanisms in the skeletal muscle development of chicken.
Peroxisomes metabolize a specific subset of fatty acids, which include dicarboxylic fatty acids (DCAs) generated by ω‐oxidation. Data obtained in vitro suggest that the peroxisomal transporter ABCD3 ...(also known as PMP70) mediates the transport of DCAs into the peroxisome, but in vivo evidence to support this role is lacking. In this work, we studied an Abcd3 KO mouse model generated by CRISPR‐Cas9 technology using targeted and untargeted metabolomics, histology, immunoblotting, and stable isotope tracing technology. We show that ABCD3 functions in hepatic DCA metabolism and uncover a novel role for this peroxisomal transporter in lipid homeostasis. The Abcd3 KO mouse presents with increased hepatic long‐chain DCAs, increased urine medium‐chain DCAs, lipodystrophy, enhanced hepatic cholesterol synthesis and decreased hepatic de novo lipogenesis. Moreover, our study suggests that DCAs are metabolized by mitochondrial fatty acid β‐oxidation when ABCD3 is not functional, reflecting the importance of the metabolic compartmentalization and communication between peroxisomes and mitochondria. In summary, this study provides data on the role of the peroxisomal transporter ABCD3 in hepatic lipid homeostasis and DCA metabolism, and the consequences of peroxisomal dysfunction for the liver.
Objective To investigate the knowledge, attitude, and practice (KAP) towards pediatric vitamin D deficiency (VitD) among parents and explore the risk factors of their knowledge, attitude, and ...practice. Methods This cross-sectional study enrolled parents in our Hospital between November 2022 and January 2023. Results A total of 621 valid questionnaires were collected in this study. The knowledge, attitude, and practice scores were 6.13 ± 3.07 (theoretical score range: 0–13), 31.13 ± 6.20 (theoretical score range: 9–45), and 27.47 ± 4.21 (theoretical score range: 9–45), respectively; the mean knowledge score was <60%, indicating poor knowledge. Commercial and service industry workers and a monthly income ≥5,000 CNY were independently associated with sufficient knowledge (all P < 0.05). The knowledge score, ethnic minorities, divorced/widows, and spouses with a master's degree or above were independently associated with positive attitudes (all P < 0.05). The attitude score, female, non-urban, undergraduate education, commercial and service industry worker, and office worker were independently associated with proactive practice (all P < 0.05). Those characteristics could help design future KAP interventions on vitD deficiency. Conclusions This study demonstrated poor knowledge, positive attitude, and proactive practice regarding pediatric VitD deficiency among parents. Targeted interventions and educational programs should be developed to improve parental knowledge.
A reliable biomarker is urgently needed in the diagnosis and management of acid sphingomyelinase deficiency (ASMD, also known as Niemann Pick A, A/B, and B). Lyso-sphingomyelin (LSM) has previously ...been proposed as a biomarker for this disease. However, existing studies have not investigated the relationship between LSM levels and clinical subtype or severity. The purpose of this study is to address this gap in knowledge.
We present a cross-sectional study of 28 patients with ASMD, enrolled in an ongoing natural history study at the Icahn School of Medicine at Mount Sinai and The Children's Hospital at Montefiore. Plasma LSM levels from 28 patients were analyzed, including 7 patients with the infantile neurovisceral phenotype (ASMD type A), 3 patients with chronic neurovisceral disease (ASMD type A/B) and 18 patients with chronic visceral ASMD (ASMD type B). The association between LSM levels and clinical subtype, dichotomized as infantile (type A) or chronic (type A/B and B), was analyzed using the Wilcoxon rank sum test. In secondary analysis, the association between LSM levels and clinical severity among the chronic ASMD patients was analyzed using the Kruskal-Wallis test.
LSM levels were elevated in all patients with ASMD when compared to a reference range of (0.04–3.8 (ng/mL)). Median LSM levels were higher in patients with infantile ASMD (386 ng/mL 314, 605) compared to chronic ASMD (133 ng/mL 90, 209), p < .001. Additionally, among individuals with chronic ASMD there was a positive association between LSM level and clinical severity (p = .01, p for trend <0.001).
We identified greater LSM elevations in patients with infantile ASMD compared to those with chronic ASMD. Among patients with chronic ASMD, LSM levels were positively associated with clinical severity. These data support investigation of LSM as a biomarker for ASMD. Future studies are required to determine if LSM levels are predictive of phenotype in pre-symptomatic patients and how such levels correlate in response to treatment.
•Lyso-sphingomyelin (LSM) has previously been proposed as a biomarker for acid sphingomyelinase deficiency (ASMD).•Existing studies have not investigated the relationship between degree of LSM elevation and clinical subtype or severity.•We present a cross-sectional study of 28 patients with ASMD enrolled in an ongoing natural history study.•LSM levels were elevated in all patients with ASMD compared to reference ranges.•Median LSM levels were higher in patients with infantile ASMD compared to chronic ASMD.•Among individuals with chronic ASMD, there was a positive association between LSM level and clinical severity.
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