Both epilepsy and anti-seizure medications affect bone metabolism in children with self-limited epilepsy with centrotemporal spikes.

OBJECTIVE: Bone metabolism can be influenced by a range of factors. We selected children with self-limited epilepsy with centrotemporal spikes (SeLECTS) and lifestyles similar to those of healthy children to control for the confounding factors that may influence bone metabolism. We aimed to identify the specific effects of epilepsy and/or anti-seizure medications (ASMs) on bone metabolism. METHODS: Patients with SeLECTS were divided into an untreated group and a monotherapy group, and the third group was a healthy control group. We determined the levels of various biochemical markers of bone metabolism, including procollagen type I nitrogenous propeptide (PINP), alkaline phosphatase (ALP), osteocalcin (OC), collagen type I cross-linked C-telopeptide (CTX), calcium, magnesium, phosphorus, parathyroid hormone (PTH), and vitamin D3 (VD3 ). RESULTS: A total of 1487 patients (from 19 centers) were diagnosed with SeLECTS; 1032 were analyzed, including 117 patients who did not receive any ASMs (untreated group), 643 patients who received only one ASM (monotherapy group), and 272 children in the healthy control group. Except for VD3 , other bone metabolism of the three groups were different (p < .001). Bone metabolism was significantly lower in the untreated group than the healthy control group (p < .05). There were significant differences between the monotherapy and healthy control group in the level of many markers. However, when comparing the monotherapy and untreated groups, the results were different; oxcarbazepine, levetiracetam, and topiramate had no significant effect on bone metabolism. Phosphorus and magnesium were significantly lower in the valproic acid group than the untreated group (adjusted p < .05, Cliff's delta .282-.768). CTX was significantly higher in the lamotrigine group than in the untreated group (adjusted p = .012, Cliff's delta = .316). SIGNIFICANCE: Epilepsy can affect many aspects of bone metabolism. After controlling epilepsy and other confounders that affect bone metabolism, we found that the effects of ASMs on bone metabolism differed. Oxcarbazepine, levetiracetam, and topiramate did not affect bone metabolism, and lamotrigine corrected some of the abnormal markers of bone metabolism in patients with epilepsy.

Use of dual genomic sequencing to screen mitochondrial diseases in pediatrics: a retrospective analysis.

Mitochondrial diseases (MDs) were a large group multisystem disorders, attributable in part to the dual genomic control. The advent of massively sequencing has improved diagnostic rates and speed, and was increasingly being used as a first-line diagnostic test. Paediatric patients (aged < 18 years) who underwent dual genomic sequencing were enrolled in this retrospective multicentre study. We evaluated the mitochondrial disease criteria (MDC) and molecular diagnostic yield of dual genomic sequencing. Causative variants were identified in 177 out of 503 (35.2%) patients using dual genomic sequencing. Forty-six patients (9.1%) had mitochondria-related variants, including 25 patients with nuclear DNA (nDNA) variants, 15 with mitochondrial DNA (mtDNA) variants, and six with dual genomic variants (MT-ND6 and POLG; MT-ND5 and RARS2; MT-TL1 and NARS2; MT-CO2 and NDUFS1; MT-CYB and SMARCA2; and CHRNA4 and MT-CO3). Based on the MDC, 15.2% of the patients with mitochondria-related variants were classified as "unlikely to have mitochondrial disorder". Moreover, 4.5% of the patients with non-mitochondria-related variants and 1.43% with negative genetic tests, were classified as "probably having mitochondrial disorder". Dual genomic sequencing in suspected MDs provided a more comprehensive and accurate diagnosis for pediatric patients, especially for patients with dual genomic variants.

UNC13B variants associated with partial epilepsy with favourable outcome.

The unc-13 homolog B (UNC13B) gene encodes a presynaptic protein, mammalian uncoordinated 13-2 (Munc13-2), which is highly expressed in the brain-predominantly in the cerebral cortex-and plays an essential role in synaptic vesicle priming and fusion, potentially affecting neuronal excitability. However, the functional significance of the UNC13B mutation in human disease is not known. In this study, we screened for novel genetic variants in a cohort of 446 unrelated cases (families) with partial epilepsy without acquired causes by trio-based whole-exome sequencing. UNC13B variants were identified in 12 individuals affected by partial epilepsy and/or febrile seizures from eight unrelated families. The eight probands all had focal seizures and focal discharges in EEG recordings, including two patients who experienced frequent daily seizures and one who showed abnormalities in the hippocampus by brain MRI; however, all of the patients showed a favourable outcome without intellectual or developmental abnormalities. The identified UNC13B variants included one nonsense variant, two variants at or around a splice site, one compound heterozygous missense variant and four missense variants that cosegregated in the families. The frequency of UNC13B variants identified in the present study was significantly higher than that in a control cohort of Han Chinese and controls of the East Asian and all populations in the Genome Aggregation Database (gnomAD). Computational modelling, including hydrogen bond and docking analyses, suggested that the variants lead to functional impairment. In Drosophila, seizure rate and duration were increased by Unc13b knockdown compared to wild-type flies, but these effects were less pronounced than in sodium voltage-gated channel alpha subunit 1 (Scn1a) knockdown Drosophila. Electrophysiological recordings showed that excitatory neurons in Unc13b-deficient flies exhibited increased excitability. These results indicate that UNC13B is potentially associated with epilepsy. The frequent daily seizures and hippocampal abnormalities but ultimately favourable outcome under anti-epileptic therapy in our patients indicate that partial epilepsy caused by UNC13B variant is a clinically manageable condition.

[Gene mutations in unexplained infantile epileptic encephalopathy: an analysis of 47 cases].

OBJECTIVE: To investigate the characteristics of gene mutations in unexplained infantile epileptic encephalopathy (EE). METHODS: A total of 47 infants with unexplained infantile EE were enrolled, and next-generation sequencing was used to analyze gene mutations in these infants and their parents. RESULTS: Of all 47 infants, 23 were found to have gene mutations, among whom 13 had de novo mutations and 10 had heterozygous mutations inherited from their father or mother. Among the 23 infants with gene mutations, 17 were found to have the gene mutations related to EE (among whom 14 had ion channel gene mutations), 2 had the gene mutations related to congenital inherited metabolic diseases, 2 had the gene mutations related to brain structural abnormality, and 2 had the gene mutations related to mental retardation. CONCLUSIONS: Unexplained infantile EE may have gene mutations, mainly ion channel gene mutations.

[Pathological changes in the epileptogenic foci of children with intractable epilepsy].

OBJECTIVE: To investigate pathological changes in the epileptogenic foci of children with intractable epilepsy and their clinical significance. METHODS: Thirty children with intractable epilepsy were included in the study. The epileptogenic foci were surgically resected and pathological changes in the obtained specimens were observed under a light microscope (LM) and a transmission electron microscope (TEM). RESULTS: Under the LM, cortical dysplasia was found in 14 cases (47%), hippocampal sclerosis in 11 cases (37%), dysembryoplastic neuroepithelial tumor in 1 case (3%), ganglioglioma in 1 case (3%), and encephalomalacia in 3 cases (10%). The TEM observation revealed pathological changes in the ultrastructure of the hippocampus and extra-hippocampal cortex, such as changes in the number of synapses and synaptic structure, decrease in neurons and karyopyknosis, swelling and degeneration of astrocytes, and changes in mitochondrial structures. CONCLUSIONS: Pathological changes in the hippocampus and extra-hippocampal cortex, especially synaptic remodeling, may be the morphological basis for spontaneous recurrent seizures in children with intractable epilepsy. The pathological changes and epileptiform activity are related to an imbalance between excitatory and inhibitory neurotransmission.

[Effect of chorioamnionitis on brain injury in preterm infants].

OBJECTIVE: To explore the association between chorioamnionitis and brain injury in preterm infants. METHODS: A total of 88 preterm infants (28-34 weeks), who were born between June 2008 and June 2011, were divided into a case group (n=41) and a control group (n=47) according to whether or not they had chorioamnionitis. All the infants were examined by brain ultrasonography periodically after birth and underwent brain diffusion weighted imaging (DWI) between 3 and 7 days after birth. The two groups were compared in terms of the incidence of periventricular leukomalacia (PVL) and periventricular and intraventricular hemorrhage (PVH-IVH) by brain magnetic resonance imaging (MRI) at the corrected gestational age of 40 weeks. RESULTS: There was statistical significance in the incidence of PVL between the case and the control groups (32% vs 6%; P<0.05), but no significant difference in the incidence of PVH-IVH between the two groups (27% vs 23%; P>0.05). CONCLUSIONS: Chorioamnionitis is associated with brain injury in preterm infants, increasing the incidence of PVL but having little influence over the incidence of PVH-IVH.

[Mutations in NPHS2 in familial steroid-resistant nephrotic syndrome in Southern Chinese Han ethnic group].

OBJECTIVE: Mutations in NPHS2 mapped to 1q25-q31 and encoding podocin, which is exclusively expressed in glomerular podocytes, are responsible for autosomal recessive familial steroid-resistant nephrotic syndrome (SRNS) with minor glomerular abnormalities or focal segmental glomerulosclerosis. Different groups from European and North American countries have screened NPHS2 mutations in familial SRNS with recessive inheritance, documenting a mutation detection rate of 45% - 55% in families. This study aimed to examine mutations in the NPHS2 gene in Southern Chinese Han ethnic group patients with familial SRNS. METHODS: Genomic DNA from 3 probands from Southern Chinese Han families with autosomal recessive SRNS, and their siblings and parents was isolated and analyzed for all eight exons, exon-intron boundaries and promoter of NPHS2 using the polymerase chain reaction and direct sequencing. RESULTS: No mutation of NPHS2 in all eight exons and exon-intron boundaries was identified in the 3 probands. However, a polymorphism of 954T > C in exon 8 was detected in all the 3 probands and some controls, and 5 variants of NPHS2 promoter, -1709G > A, -1000A > T, -670C > T, -116C > T and -51G > T, were identified in some patients and controls, indicating that these variants are polymorphisms. One heterozygous variant of NPHS2 promoter, -1715A > G, was also identified in one proband and her mother whose urinalyses were normal, whereas it was not found in any of the 50 controls. There was no significant difference in the allelic frequencies of -1709G > A, -1000A > T, -670C > T, -116C > T and -51G > T polymorphisms between the patients and controls. CONCLUSION: NPHS2 mutations are not a major cause of familial steroid-resistant nephrotic syndrome in Southern Chinese Han ethnic group included in the study.

[Neuroprotective effects of topiramate and folic acid on young rats with kindling-induced epilepsy].

OBJECTIVE: To study the neuroprotective effects of topiramate (TPM) alone or together with folic acid (FA) on young rats with kindling-induced epilepsy. METHODS: Rat models of epilepsy were prepared by pentylenetetrazol (PTZ)-induced kindling. Seventy-two 3-week-old male Wistar rats were randomly divided into 6 groups: four TPM-treated epilepsy groups (TPM 20, 40 or 80 mg/kg/d and TPM 40 mg/kg/d + FA 5 mg/kg/d), a positive control group (untreated epilepsy group) and a negative control group (normal control group). After two months of administration, behaviors of the rats were recorded; serum levels of neuron-specific enolase (NSE) were measured using ELISA; pathological changes in the hippocampus were observed. RESULTS: The frequency of convulsion seizures in the 20, 40 and 80 mg TPM treatment and TPM+FA groups was 44.7 +/- 2.9, 44.3 +/- 3.1, 42.7 +/- 3.2, and 40.8 +/- 3.7 respectively, which were significantly lower than that in the positive control group (48.4 +/- 3.7) (P <0.01). Twenty, forty and eighty mg TPM treatment and TPM+FA treatment significantly reduced NSE levels from 35.71 +/- 5.97 microg/L of the control group to 27.40+/- 6.40, 24.79 +/- 6.22, 21.47 +/- 6.87 and 22.55 +/- 7.02 microg/L respectively (P <0.05). Neuronal apoptosis in the CA3 and CA1 regions were alleviated in the four TPM treatment groups compared with positive control. The number of necrotic neurons was progressively reduced with the increased dose of TPM. The 40 mg TPM+FA treatment group showed less necrotic neurons in the CA3 and CA1 regions than the 40 mg TPM alone treatment group. CONCLUSIONS: TPM has protective effects against epilepsy-induced neuronal damage. The effect is dose-dependent. A combination of TPM and FA can produce a synergistic effect.

[An experimental study on hepatotoxicity of topiramate in young rats].

OBJECTIVE: Topiramate (TPM) has an evident efficacy in the treatment of childhood epilepsy for multiple pharmacologic properties. However it was reported that it may cause adverse effects such as liver failure and hepatitis, which arouses the attention of the medical field. This study aimed to investigate the hepatotoxicity of low-dosage, high-dosage TPM or TPM along with valproate sodium (VPA) in aspects of biochemistry indexes, oxidative stress indexes and liver pathomorphology in young rats. METHODS: Sixty 3-week-old male Wistar rats were randomly assigned into five groups of 12 rats (Groups A-E). The rats in the experimental groups (Groups A-C) were administered intragastrically with TPM 40 mg/(kg.d), 80 mg/(kg.d) and TPM 40 mg/(kg.d) plus VPA 300 mg/(kg.d) respectively. The rats in the negative control group (Group D) were administered with the same volume of distilled water. The ones in the positive control group (Group E) were treated by injection of 10% carbon tetrachloride dissolved in olive oil subcutaneously at a dose of 5 mL/kg twice a week. After 3-month administration, the changes of body weight and liver pathomorphology were observed; biochemical markers in serum and indexes of oxidative stress in liver homogenate associated with hepatotoxicity were examined. RESULTS: The body weights of rats in the experimental groups were significantly lower than that of rats in the negative control group. The levels of serum alanine aminotransferase, alkaline phosphatase and the content of malondialdehyde, and the activity of superoxide dismutase in liver tissues did not change significantly in the experimental groups. The contents of glutathion in the high dosage of TPM group (29.85 +/- 1.62 mg/g prot) or in the TPM plus VPA group (29.63 +/- 4.47 mg/g prot) were significantly reduced compared with those of the negative control group (33.09 +/- 1.69 mg/g prot) and that of the low dosage of TPM group (32.43 +/- 2.11 mg/g prot) (both P < 0.05). In the histopathological examination, extensive steatosis and diffuse punctate necrosis of hepatocytes distributed in the portal area were found by microscopy in the positive control group. There were granular degeneration of some hepatocytes near the central veins of hepatic lobules in the low dosage of TPM group and punctate necrosis of some hepatocytes in the high dosage of TPM group. In the TPM plus VPA group, histological examination showed granular degeneration and fatty degeneration of partial liver cells and punctate necrosis of some hepatocytes. CONCLUSIONS: Long-term use of TPM can decrease antioxidant capacity of organism, resulting in slight pathological changes of liver tissues. High dosage of TPM or TPM along with VPA administration enhances the risk of the side effects.

[Treatment of hemolytic uremic syndrome after acute stage].

OBJECTIVE: Hemolytic uremic syndrome (HUS) is a common primary disease that can cause acute renal failure in childhood. Renal disease is the most important long-term complication in patients who survived the acute stage of HUS. Use of angiotensin-converting enzyme inhibitors (ACEI) and a restricted protein intake may be beneficial to the patients. However, it is not established whether such patients should be treated with steroids and immunosuppressors. The present study aimed to probe into the benefit of using steroid and immunosuppressor in patients after acute stage of HUS. METHODS: The subjects included 17 patients (aged 9 months to 15 years, 12 males, 5 females) with HUS. Thirteen patients recovered from the acute stage of HUS, and underwent continuative treatment and follow-up. All the patients were treated with ACEI and early restriction of protein intake. Additionally, 2 children manifested as glomerulonephritis, one was treated with triperygium glycosides. Other 11 children who manifested as nephrotic syndrome were treated with prednisone, among them 5 children had no response or had incomplete response to prednisone, for these children short-term high dose cyclophosphamide or methylprednisolone pulse treatment were added; in 3 of the children short-term high dose methylprednisolone treatment was applied additionally for membranoproliferative glomerulonephritis and/or focal segmental glomerulosclerosis and crescentic glomerulonephritis. RESULTS: After follow-up for 2 months to 8 years, 4 patients with milder disease recovered, their blood pressure, renal function and urinalysis became normal, but 1 patient had recurrence. Among 9 patients with severe disease, 6 maintained normal blood pressure, recovered renal function and urinalysis, the other 3 patients failed to comply with treatment protocol and died during the 3rd, 9th and 13th month. The remainder (4 cases) gave up therapy and died on the 27th to 48th days of the course. CONCLUSION: The treatment applied in this study could improve the prognosis of patients after acute phase of HUS evidently by using the steroid and immuno suppressor according to clinical classification and pathological findings. It is recommended that triperygium glycosides is beneficial to children with glomerulonephritis, proteinuria and hematuria after acute stage of HUS. Adjustment of therapeutic schedule based on pathological findings after renal biopsy is helpful. To the patients with progressive renal failure who have no response to the steroid and immunosuppressors, steroid and immunosuppressor should be discontinued and dialysis treatment should be applied. Protocol compliance is also an important factor.

[Effect of glutamine on small intestinal repair in weanling rats after chronic diarrhea].

OBJECTIVE: To investigate the nutrient effect of glutamine on small intestinal repair in weanling rats after chronic diarrhea. METHODS: Forty 21-day-old wistar rats were randomly divided into five groups (8 in each). Animal model of chronic diarrhea was induced by a lactose enriched diet in the weanling Wistar rat, normal control group was fed with a standard semipurified diet, and after 14 days the rats in both groups were killed to test the establishment of the model. After the establishment of the model, the other groups were fed with the standard semipurified diet to recover for 7 days, and were randomly divided into three groups: non-intervention group, glutamine (Gln)-intervention group and control group. Glutamine concentrations in blood was detected by high-performance liquid chromatography (HPLC). Morphological changes including villus height and villus surface area of the jejunum were measured under a light microscope and electron microscope, expression of proliferating cell nuclear antigen (PCNA) as an index of cell proliferation was observed using immunohistochemical staining and image analysis. RESULTS: The diarrhea rate in model group was 100 percent, average diarrhea index was 1.16 +/- 0.06, but both diarrhea rate and average diarrhea index in control group were 0 (P < 0.01), which affirmed establishment of the model. There was significant decrease of body weight, plasma Gln concentration, villus height, villus surface area and expression of PCNA in non-intervened group compared with the control group (P < 0.01). There was still significant decrease of body weight, villus height and villus surface area in Gln-intervened group compared with control group (P < 0.01), but plasma Gln concentration and expression of PCNA in Gln-intervened group had recovered to normal (P > 0.05). And compared with non-intervened group, except for body weight (P > 0.05), plasma glutamine, villus height, villus surface area and expression of PCNA were all significantly increased in Gln-intervened group. CONCLUSION: Chronic diarrhea can induce malnutrition and reduce the villus height, villus surface area, expression of PCNA and plasm glutamine concentration. Oral glutamine could improve the proliferation of crypt cell and promote repair of intestinal mucosa after chronic diarrhea.