Differential growth inhibition, cell cycle arrest and apoptosis of MCF-7 and MDA-MB-231 cells to holocarboxylase synthetase suppression.

Holocarboxylase synthetase (HLCS) catalyzes the covalent attachment of biotin onto the biotin-dependent carboxylases. Recent studies have shown that HLCS is over-expressed in breast cancer patients. Here we investigated the functional roles of free biotin and HLCS in supporting growth and migration of breast cancer cell lines. Depletion of biotin from culture medium markedly reduced biotinylation of the two most abundant biotin-carboxylases, acetyl-CoA carboxylase and pyruvate carboxylase. This was accompanied by a marked decrease in cell growth. Suppression of HLCS expression in the low invasive breast cancer cell line MCF-7 resulted in an 80% reduction of biotinylated ACC, but not PC. HLCS knockdown MCF-7 cell lines showed 40-50% reduction of proliferation and 35% reduction of migration, accompanied by G1 cell cycle-arrest-induced apoptosis. In contrast, knockdown of HLCS expression in the highly invasive cell line MDA-MB-231 resulted in only marginal reduction of biotinylation of both ACC and PC, accompanied by 30% reduction of proliferation and 30% reduction of migration. Our studies provide new insights to use HLCS as a novel anti-cancer drug target.

Urinary 3-hydroxyisovaleryl carnitine excretion, protein energy malnutrition and risk of all-cause mortality in kidney transplant recipients: Results from the TransplantLines cohort studies.

BACKGROUND: Leucine is an essential amino acid and a potent stimulator of muscle protein synthesis. Since muscle wasting is a major risk factor for mortality in kidney transplant recipients (KTR), dietary leucine intake might be linked to long-term mortality. Urinary 3-hydroxyisovaleryl carnitine (3-HIC) excretion, a functional marker of marginal biotin deficiency, may also serve as a marker for dietary leucine intake. OBJECTIVE: In this study we aimed to investigate the cross-sectional determinants of urinary 3-HIC excretion and to prospectively investigate the association of urinary 3-HIC excretion with all-cause mortality in KTR. DESIGN: Urinary 3-HIC excretion and plasma biotin were measured in a longitudinal cohort of 694 stable KTR. Cross-sectional and prospective analyses were performed using ordinary least squares linear regression analyses and Cox regression analyses, respectively. RESULTS: In KTR (57% male, 53 ± 13 years, estimated glomerular filtration rate 45 ± 19 mL/min/1.73 m2), urinary 3-HIC excretion (0.80 [0.57-1.16] µmol/24 h) was significantly associated with plasma biotin (std. ß = -0.17; P < 0.001). Subsequent adjustment for potential covariates revealed urinary creatinine excretion (std. ß = 0.24; P < 0.001) and urinary urea excretion (std. ß = 0.53; P < 0.001) as the primary determinant of urinary 3-HIC excretion. Whereas plasma biotin explained only 1% of the variance in urinary 3-HIC excretion, urinary urea excretion explained >45%. During median follow-up for 5.4 [4.8-6.1] years, 150 (22%) patients died. Log2-transformed urinary 3-HIC excretion was inversely associated with all-cause mortality (HR: 0.52 [0.43-0.63]; P < 0.001). This association was independent of potential confounders. CONCLUSIONS: Urinary 3-HIC excretion more strongly serves as a marker of leucine intake than of biotin status. A higher urinary 3-HIC excretion is associated with a lower risk of all-cause mortality. Future studies are warranted to explore the underlying mechanism. TRIAL REGISTRATION ID: NCT02811835. TRIAL REGISTRATION URL: https://clinicaltrials.gov/ct2/show/NCT02811835.

Hair Loss After Sleeve Gastrectomy and Effect of Biotin Supplements.

Aim: In this study, we aimed to determine the incidence of hair loss in patients who underwent laparoscopic sleeve gastrectomy (LSG), and to observe whether use of Biotin has an impact on hair loss. Methods: This study included 156 female patients who underwent LSG for obesity and completed a 1-year follow-up. All patients with vitamin deficiency were screened in the pre- and postoperative period. Hair loss was defined as the subjective perception of the women of losing a higher amount of hair when compared with normal situation. Results: Hair loss was observed in 72% of the patients after LSG (n = 112). Seventy-nine percent of the patients reported hair loss between the third and fourth-month interval, and continued for an average of 5.5 ± 2.6 months. Permanent alopecia was not observed in any of the patients. Patients who experienced hair loss and Biotin deficiency after LSG were prescribed 1000 mcg/day of Biotin for 3 months. Of these 22 patients; only 5 (23%) patients reported a remarkable decline in hair loss. In addition, 29 patients were found to take 1000 mcg/day of Biotin for average 2.5 months after onset of hair loss by their own initiative, despite optimal blood Biotin levels. Eleven (38%) patients reported a remarkable decline in hair loss. The effect of biotin use on hair loss in patients with and without biotin deficiency was compared. There was no significant difference (P = .2). Conclusion: Temporary hair loss after LSG is common. It was found that biotin supplementation used to prevent hair loss does provide low efficacy.

Update on Biotin Therapy in Dermatology: Time for a Change.

Biotin (vitamin B7 or H) is found in milk, nuts, egg yolks, cereals, supplements, synthesized by intestinal bacteria, and is required for gluconeogenesis, fatty acid synthesis and amino acid catabolism.

Biotin rescues mitochondrial dysfunction and neurotoxicity in a tauopathy model.

Mitochondrial and metabolic dysfunction are often implicated in neurological disease, but effective mechanism-based therapies remain elusive. We performed a genome-scale forward genetic screen in a Drosophila model of tauopathy, a class of neurodegenerative disorders characterized by the accumulation of the protein tau, and identified manipulation of the B-vitamin biotin as a potential therapeutic approach in tauopathy. We show that tau transgenic flies have an innate biotin deficiency due to tau-mediated relaxation of chromatin and consequent aberrant expression of multiple biotin-related genes, disrupting both carboxylase and mitochondrial function. Biotin depletion alone causes mitochondrial pathology and neurodegeneration in both flies and human neurons, implicating mitochondrial dysfunction as a mechanism in biotin deficiency. Finally, carboxylase biotin levels are reduced in mammalian tauopathies, including brains of human Alzheimer's disease patients. These results provide insight into pathogenic mechanisms of human biotin deficiency, the resulting effects on neuronal health, and a potential therapeutic pathway in the treatment of tau-mediated neurotoxicity.

Biotin biosynthesis affected by the NADPH oxidase and lipid metabolism is required for growth, sporulation and infectivity in the citrus fungal pathogen Alternaria alternata.

The tangerine pathotype of Alternaria alternata affects many citrus cultivars, resulting in yield losses. The capability to produce the host-selective toxin and cell-wall-degrading enzymes and to mitigate toxic reactive oxygen species is crucial for A. alternata pathogenesis to citrus. Little is known about nutrient availability within citrus tissues to the fungal pathogen. In the present study, we assess the infectivity of a biotin deficiency mutant (ΔbioB) and a complementation strain (CP36) on citrus leaves to determine how biotin impacts A. alternata pathogenesis. Growth and sporulation of ΔbioB are highly dependent on biotin. ΔbioB retains its ability to acquire and transport biotin from the surrounding environment. Growth deficiency of ΔbioB can also be partially restored by the presence of oleic acid or Tween 20, suggesting the requirement of biotin in lipid metabolism. Experimental evidence indicates that de novo biotin biosynthesis is regulated by the NADPH oxidase, implicating in the production of H2O2, and is affected by the function of peroxisomes. Three genes involved in the biosynthesis of biotin are clustered and co-regulated by biotin indicating a transcriptional feedback loop activation. Infectivity assays using fungal mycelium reveal that ΔbioB cultured on medium without biotin fails to infect citrus leaves; co-inoculation with biotin fully restores infectivity. The CP36 strain re-expressing a functional copy of bioB displays wild-type growth, sporulation and virulence. Taken together, we conclude that the attainability or accessibility of biotin is extremely restricted in citrus cells. A. alternata must be able to synthesize biotin in order to utilize nutrients for growth, colonization and development within the host.

Serum biotin level during pregnancy is associated with fetal growth and preterm delivery.

Background : Biotin is a water-soluble vitamin that plays various biological roles through histone modification, such as immune functions and fetal growth. Mammalian maternal biotin deficiency during gestation induces fetal growth restriction. Preterm infants are known to be marginal biotin deficiency. However, studies on the biotin status of pregnant women under various conditions are lacking. Method : This was a retrospective case control study to analyze serum biotin concentration during pregnancy and cord blood in normal pregnancy, preterm delivery and small-for-gestational-age (SGA). Results : Twenty pregnant women with normal term delivery, 35 with preterm delivery, 24 with SGA, and 10 non-pregnant adult women were enrolled. Serum biotin concentrations of pregnant women remained low from first to third trimester. The levels of serum biotin in cord blood showed a significant positive correlation with gestational age, and that of pregnant women showed a weak positive correlation with gestational age. The maternal serum biotin levels during second and third trimester of SGA group were significantly lower than those of normal term delivery. Conclusion : This study suggests that maternal biotin deficiency during pregnancy might be the risk of preterm labor or fetal growth restriction. Further studies are required to clarify the roles of biotin in perinatal medicine. J. Med. Invest. 67 : 170-173, February, 2020.

Metabolomic Analysis of Liver from Dietary Biotin Deficient Mice.

Biotin is a water-soluble B-complex vitamin that functions as a cofactor of five carboxylases. Because biotin-dependent carboxylases catalyze indispensable cellular metabolic functions, biotin deficiency is considered to be involved in various pathological conditions. Moreover, biotin supplementation shows pharmacological effects in vivo. However, the precise mechanisms by which biotin deficiency induces pathological conditions remain unclear. Although abnormal metabolites are used as indicators for biotin deficiency, few comprehensive analyses of total metabolites have been reported. In this study, we analyzed the metabolomic profiles of liver extracts prepared from biotin-sufficient (BS) and -deficient (BD) mice. Thirteen of 126 metabolites showed significantly different concentrations between liver extracts from BD and BS mice. The concentrations of 5 essential amino acids, Met, Val, Thr, Ile, and Leu, and 2 conditionally essential amino acids, Cys and Tyr were significantly lower in BD mice than in BS mice. Among these, the concentrations of sulfur-containing amino acids, Cys and Met, were more than 1.5-fold lower in BD mice. The concentrations of Met metabolites, such as S-adenosylmethionine and S-adenosylhomocysteine were not significantly different between the two groups. The concentrations of glutathione and its reaction intermediates γ-Glu-Cys tendency to be lower in BD mice. The present study revealed that biotin deficiency induces an abnormal amino acids composition, especially among sulfur-containing amino acids and provide important information on the effect of biotin as a pharmacological agent.

Biotin Is Required for the Zinc Homeostasis in the Skin.

Patients with biotin deficiency present symptoms that are similar to those in patients with acrodermatitis enteropathica (inherent zinc deficiency). However, the association between biotin and zinc deficiency remains unknown. We have previously shown that epidermal keratinocytes of mice fed zinc-deficient (ZD) diets secreted more adenosine triphosphate (ATP) than those of mice fed zinc-adequate (ZA) diets and that epidermal Langerhans cells are absent in ZD mice. Langerhans cells highly express CD39, which potently hydrolyzes ATP into adenosine monophosphate (AMP). Thus, a lack of Langerhans cells in ZD mice leads to non-hydrolysis of ATP, thereby leading to the development of ATP-mediated irritant contact dermatitis. In this study, we examined if biotin-deficient (BD) mice showed the same underlying mechanisms as those in ZD mice. BD mice showed reduced serum zinc levels, disappearance of epidermal Langerhans cells, and enhanced ATP production in the skin. Consequently, irritant contact dermatitis was significantly enhanced and prolonged in BD mice. In conclusion, the findings of our study showed that biotin deficiency leads to zinc deficiency because of which patients with biotin deficiency show similar symptoms as those with acrodermatitis enteropathica.

Biotin in metabolism, gene expression, and human disease.

Biotin is a water-soluble vitamin that belongs to the vitamin B complex and which is an essential nutrient of all living organisms from bacteria to man. In eukaryotic cells biotin functions as a prosthetic group of enzymes, collectively known as biotin-dependent carboxylases that catalyze key reactions in gluconeogenesis, fatty acid synthesis, and amino acid catabolism. Enzyme-bound biotin acts as a vector to transfer a carboxyl group between donor and acceptor molecules during carboxylation reactions. In recent years, evidence has mounted that biotin also regulates gene expression through a mechanism beyond its role as a prosthetic group of carboxylases. These activities may offer a mechanistic background to a developing literature on the action of biotin in neurological disorders. This review summarizes the role of biotin in activating carboxylases and proposed mechanisms associated with a role in gene expression and in ameliorating neurological disease.

Intestinal Dysbiosis and Biotin Deprivation Induce Alopecia through Overgrowth of Lactobacillus murinus in Mice.

Metabolism by the gut microbiota affects host physiology beyond the gastrointestinal tract. Here, we find that antibiotic-induced dysbiosis, in particular, overgrowth of Lactobacillus murinus (L. murinus), impaired gut metabolic function and led to the development of alopecia. While deprivation of dietary biotin per se did not affect skin physiology, its simultaneous treatment with vancomycin resulted in hair loss in specific pathogen-free (SPF) mice. Vancomycin treatment induced the accumulation of L. murinus in the gut, which consumes residual biotin and depletes available biotin in the gut. Consistently, L. murinus induced alopecia when monocolonized in germ-free mice fed a biotin-deficient diet. Supplementation of biotin can reverse established alopecia symptoms in the SPF condition, indicating that L. murinus plays a central role in the induction of hair loss via a biotin-dependent manner. Collectively, our results indicate that luminal metabolic alterations associated with gut dysbiosis and dietary modifications can compromise skin physiology.

Biotin: From Nutrition to Therapeutics.

Although frank symptomatic biotin deficiency is rare, some evidence suggests that marginal biotin deficiency occurs spontaneously in a substantial proportion of women during normal human pregnancy and might confer an increased risk of birth defects. Herein I review 1) advances in assessing biotin status, including the relation between acylcarnitine excretion and biotin status; 2) recent studies of biotin status in pregnancy; 3) advances in understanding the role of biotin in gene expression and the potential roles of biotinylated proteins that are neither histones nor carboxylases; and 4) novel large-dose biotin supplementation as therapy for multiple sclerosis. The review concludes with a summary of recent studies that have reported potentially dangerous erroneous results in individuals consuming large amounts of biotin for measurements of various plasma hormones for common clinical assays that use streptavidin-biotin technology.

Acrodermatitis caused by nutritional deficiency and metabolic disorders.

Both the metabolism and dietary intake of vitamins and minerals are essential to homeostatic function in the body. Dietary excess or deficiency, as well as genetic and acquired disorders in metabolism, can present dermatologically and systemically. More specifically, disorders in metabolism of zinc, biotin, essential fatty acids, and vitamin B, can appear with acrally distributed dermatoses. Recognition of the dermatologic manifestations associated with nutritional disorders is important for early diagnosis and management.

Screening of carnitine and biotin deficiencies on tandem mass spectrometry.

BACKGROUND: It is important to assess pediatric patients for nutritional deficiency when they are receiving specific interventions, such as enteral feeding. We focused on measurement of C0 and 3-hydroxyisovalerylcarnitine (C5-OH) with tandem mass spectrometry (MS/MS), which is performed as part of the newborn mass screening. The purpose of this study was to investigate the usefulness of MS/MS for screening carnitine and biotin deficiencies. METHODS: Forty-two children (24 boys, 18 girls) were enrolled between December 2013 and December 2015. Blood tests, including measurement of serum free carnitine via the enzyme cycling method, and acylcarnitine analysis on MS/MS of dried blood spot (DBS), were performed for the evaluation of nutrition status. RESULTS: Median patient age was 2 years (range, 2 months-14 years). Mean serum free carnitine was 41.8 ± 19.2 µmol/L. In six of the 42 patients, serum free carnitine was <20 µmol/L (range, 4.0-18.7 µmol/L). C0 and C5-OH measured on MS/MS of DBS were 33.8 ± 20.2 nmol/mL and 0.48 ± 0.22 nmol/mL, respectively. There was a strong positive correlation (r = 0.89, P < 0.001) between serum free carnitine and C0 measured on the same day. In one patient on hydrolyzed formula, C5-OH was >1.00 nmol/L. Therapy-resistant eczema was improved by treatment with additional biotin and a non-hydrolyzed formula. CONCLUSION: C0 and C5-OH, measured on MS/MS of DBS, were useful for screening carnitine and biotin deficiencies.

Selective accumulation of biotin in arterial chemoreceptors: requirement for carotid body exocytotic dopamine secretion.

KEY POINTS: Biotin, a vitamin whose main role is as a coenzyme for carboxylases, accumulates at unusually large amounts within cells of the carotid body (CB). In biotin-deficient rats biotin rapidly disappears from the blood; however, it remains at relatively high levels in CB glomus cells. The CB contains high levels of mRNA for SLC5a6, a biotin transporter, and SLC19a3, a thiamine transporter regulated by biotin. Animals with biotin deficiency exhibit pronounced metabolic lactic acidosis. Remarkably, glomus cells from these animals have normal electrical and neurochemical properties. However, they show a marked decrease in the size of quantal dopaminergic secretory events. Inhibitors of the vesicular monoamine transporter 2 (VMAT2) mimic the effect of biotin deficiency. In biotin-deficient animals, VMAT2 protein expression decreases in parallel with biotin depletion in CB cells. These data suggest that dopamine transport and/or storage in small secretory granules in glomus cells depend on biotin. ABSTRACT: Biotin is a water-soluble vitamin required for the function of carboxylases as well as for the regulation of gene expression. Here, we report that biotin accumulates in unusually large amounts in cells of arterial chemoreceptors, carotid body (CB) and adrenal medulla (AM). We show in a biotin-deficient rat model that the vitamin rapidly disappears from the blood and other tissues (including the AM), while remaining at relatively high levels in the CB. We have also observed that, in comparison with other peripheral neural tissues, CB cells contain high levels of SLC5a6, a biotin transporter, and SLC19a3, a thiamine transporter regulated by biotin. Biotin-deficient rats show a syndrome characterized by marked weight loss, metabolic lactic acidosis, aciduria and accelerated breathing with normal responsiveness to hypoxia. Remarkably, CB cells from biotin-deficient animals have normal electrophysiological and neurochemical (ATP levels and catecholamine synthesis) properties; however, they exhibit a marked decrease in the size of quantal catecholaminergic secretory events, which is not seen in AM cells. A similar differential secretory dysfunction is observed in CB cells treated with tetrabenazine, a selective inhibitor of the vesicular monoamine transporter 2 (VMAT2). VMAT2 is highly expressed in glomus cells (in comparison with VMAT1), and in biotin-deficient animals VMAT2 protein expression decreases in parallel with the decrease of biotin accumulated in CB cells. These data suggest that biotin has an essential role in the homeostasis of dopaminergic transmission modulating the transport and/or storage of transmitters within small secretory granules in glomus cells.

Effects of Biotin Deficiency on Biotinylated Proteins and Biotin-Related Genes in the Rat Brain.

Biotin is a water-soluble vitamin that functions as a cofactor for biotin-dependent carboxylases. The biochemical and physiological roles of biotin in brain regions have not yet been investigated sufficiently in vivo. Thus, in order to clarify the function of biotin in the brain, we herein examined biotin contents, biotinylated protein expression (e.g. holocarboxylases), and biotin-related gene expression in the brain of biotin-deficient rats. Three-week-old male Wistar rats were divided into a control group, biotin-deficient group, and pair-fed group. Rats were fed experimental diets from 3 wk old for 8 wk, and the cortex, hippocampus, striatum, hypothalamus, and cerebellum were then collected. In the biotin-deficient group, the maintenance of total biotin and holocarboxylases, increases in the bound form of biotin and biotinidase activity, and the expression of an unknown biotinylated protein were observed in the cortex. In other regions, total and free biotin contents decreased, holocarboxylase expression was maintained, and bound biotin and biotinidase activity remained unchanged. Biotin-related gene (pyruvate carboxylase, sodium-dependent multivitamin transporter, holocarboxylase synthetase, and biotinidase) expression in the cortex and hippocampus also remained unchanged among the dietary groups. These results suggest that biotin may be related to cortex functions by binding protein, and the effects of a biotin deficiency and the importance of biotin differ among the different brain regions.

Biotin deprivation impairs mitochondrial structure and function and has implications for inherited metabolic disorders.

Certain inborn errors of metabolism result from deficiencies in biotin containing enzymes. These disorders are mimicked by dietary absence or insufficiency of biotin, ATP deficit being a major effect,whose responsible mechanisms have not been thoroughly studied. Here we show that in rats and cultured cells it is the result of reduced TCA cycle flow, partly due to deficient anaplerotic biotin-dependent pyruvate carboxylase. This is accompanied by diminished flow through the electron transport chain, augmented by deficient cytochrome c oxidase (complex IV) activity with decreased cytochromes and reduced oxidative phosphorylation. There was also severe mitochondrial damage accompanied by decrease of mitochondria, associated with toxic levels of propionyl CoA as shown by carnitine supplementation studies, which explains the apparently paradoxical mitochondrial diminution in the face of the energy sensor AMPK activation, known to induce mitochondria biogenesis. This idea was supported by experiments on AMPK knockout mouse embryonic fibroblasts (MEFs). The multifactorial ATP deficit also provides a plausible basis for the cardiomyopathy in patients with propionic acidemia, and other diseases.Additionally, systemic inflammation concomitant to the toxic state might explain our findings of enhanced IL-6, STAT3 and HIF-1α, associated with an increase of mitophagic BNIP3 and PINK proteins, which may further increase mitophagy. Together our results imply core mechanisms of energy deficit in several inherited metabolic disorders.

Microarray analysis of pancreatic gene expression during biotin repletion in biotin-deficient rats.

Biotin is a B vitamin involved in multiple metabolic pathways. In humans, biotin deficiency is relatively rare but can cause dermatitis, alopecia, and perosis. Low biotin levels occur in individuals with type-2 diabetes, and supplementation with biotin plus chromium may improve blood sugar control. The acute effect on pancreatic gene expression of biotin repletion following chronic deficiency is unclear, therefore we induced biotin deficiency in adult male rats by feeding them a 20% raw egg white diet for 6 weeks. Animals were then randomized into 2 groups: one group received a single biotin supplement and returned to normal chow lacking egg white, while the second group remained on the depletion diet. After 1 week, pancreata were removed from biotin-deficient (BD) and biotin-repleted (BR) animals and RNA was isolated for microarray analysis. Biotin depletion altered gene expression in a manner indicative of inflammation, fibrosis, and defective pancreatic function. Conversely, biotin repletion activated numerous repair and anti-inflammatory pathways, reduced fibrotic gene expression, and induced multiple genes involved in pancreatic endocrine and exocrine function. A subset of the results was confirmed by quantitative real-time PCR analysis, as well as by treatment of pancreatic AR42J cells with biotin. The results indicate that biotin repletion, even after lengthy deficiency, results in the rapid induction of repair processes in the pancreas.

Biotin starvation causes mitochondrial protein hyperacetylation and partial rescue by the SIRT3-like deacetylase Hst4p.

The essential vitamin biotin is a covalent and tenaciously attached prosthetic group in several carboxylases that play important roles in the regulation of energy metabolism. Here we describe increased acetyl-CoA levels and mitochondrial hyperacetylation as downstream metabolic effects of biotin deficiency. Upregulated mitochondrial acetylation sites correlate with the cellular deficiency of the Hst4p deacetylase, and a biotin-starvation-induced accumulation of Hst4p in mitochondria supports a role for Hst4p in lowering mitochondrial acetylation. We show that biotin starvation and knockout of Hst4p cause alterations in cellular respiration and an increase in reactive oxygen species (ROS). These results suggest that Hst4p plays a pivotal role in biotin metabolism and cellular energy homeostasis, and supports that Hst4p is a functional yeast homologue of the sirtuin deacetylase SIRT3. With biotin deficiency being involved in various metabolic disorders, this study provides valuable insight into the metabolic effects biotin exerts on eukaryotic cells.

Screening of Inherited Metabolic Disorders in Infants with Infantile Spasms.

The objective of this study is to explore the incidence of inherited metabolic disorders (IMD) in infants with infantile spasms (IS), with an attempt to improve the early diagnosis and etiological and symptomatic treatment. Urine and blood samples were collected from 60 IS patients and analyzed for the quantification of amino acids, organic acids, and fatty acids by gas chromatography-mass spectrometry and tandem mass spectrum. Routine urine tests, hepatic function tests, blood biochemistry, brain imaging, as well as examinations of the brain stem auditory/visual evoked potentials were also examined. In addition to antiepileptic therapy, etiological and symptomatic treatments were also conducted in infants with confirmed IMD and the follow-up lasted for 6 months in these pediatric patients. Metabolic disorders were found in 28 (46.67 %) of 60 IS infants, among them 13 (21.67 %) were confirmed to be with IMD. Twelve of these 13 IS patients with definite IMD diagnoses (92.31 %) experienced varying degrees of delayed development of intelligence and motor function, 8 patients (61.54 %) had abnormal cranial CT or MRI findings, 11 patients (84.61 %) had abnormal brain stem evoked potentials, 4 patients (30.77 %) had abnormal hepatic functions, 3 patients (23.07 %) had abnormal blood biochemistry, 2 patients (15.38 %) had positive (+ to ++) results for routine urine ketones, and 2 patients (15.38 %) had skin lesions. After treatment in children who were diagnosed IMD, the well controlled epileptic seizures and the satisfactory developments in mental and motor were found in 4 cases of methylmalonic acidemia, 2 cases of classical phenylketonuria, and one case of biotin deficiency disease, glutaric acidemia type I, and 4-hydroxybutyric aciduria in each. IMD is a key biological cause in IS. Early screening for IMD is warranted in IS infants to facilitate the improvement for the prognosis and an early etiological treatment.