Integration of Viral Genome to Human Genomic DNA in Nails of Patients with Chronic Hepatitis B Virus Infection.

Introduction: Hepatitis B virus (HBV) DNA and cytomegalovirus (CMV) DNA can be detected in patient genomes. However, it remains unknown whether viral DNA can be integrated into host genomic DNA and detected in fingernails. Methods: Nails from patients with chronic HBV infection were investigated. A total of 60 patients (male/female = 20/40, age range from 2 years to 59 years, median 15 years) were included in this study. The viral DNA levels of herpes simplex virus 1 (HSV-1), herpes simplex virus 2 (HSV-2), varicella-zoster virus (VZV), Epstein‒Barr virus (EBV), cytomegalovirus (CMV), human herpes virus 6 (HHV-6), human herpes virus 7 (HHV-7), and HBV in nails were measured with real-time PCR. Viral DNA integration into host genomic DNA was analyzed by capture-based next-generation sequencing (NGS). Moreover, virus/host chimeric sequences, which were detected by capture-based NGS, were confirmed by Sanger sequencing. Results: Of the 60 patients, 37 (62%) were positive for nail HBV DNA. All 60 patients were negative for nail HSV-1, HSV-2, VZV, CMV, EBV, or HHV-6 DNA. However, three patients were positive for nail HHV-7 DNA. All three nail HHV-7-positive patients were also positive for nail HBV DNA. The three nail samples that were positive for both HBV and HHV-7 DNA were used for viral integration analysis by capture-based NGS. One of the three nail samples showed HBV/host chimeric sequences. In addition, all three nail samples showed HHV-7/host chimeric sequences. However, these viral integration breakpoints were not confirmed by Sanger sequencing. Conclusions: Viral integrations were detected in nails by capture-based NGS. However, Sanger sequencing did not confirm any virus/host chimeric sequences. This study could not show reliable evidence of viral integration in nails.

Peripheral-dominant liver fibrosis and tumor distribution in a mouse model of congestive hepatopathy.

AIM: Congestive hepatopathy often leads to liver fibrosis and hepatocellular carcinoma. Imaging modalities provided clinical evidence that elevation of liver stiffness and tumor occurrence are mainly induced in the periphery of the liver in patients with congestive hepatopathy. However, clinical relevance of liver stiffness and liver fibrosis is unclear because liver congestion itself increases liver stiffness in congestive hepatopathy. It also unclear which factors configure such regional disparity of tumor development in patients with congestive hepatopathy. To answer these questions, we evaluated the macroscopic spatial distribution of liver fibrosis and tumors in the murine model of congestive hepatopathy. METHODS: Chronic liver congestion was induced by partial ligation of the suprahepatic inferior vena cava. Distribution of liver congestion, fibrosis, and tumors in partial ligation of the suprahepatic inferior vena cava mice were assessed by histological findings, laser microdissection (LMD)-based qPCR and enhanced computed tomography. LMD-based RNA-sequencing was performed to identify causal factors that promote tumor development in congestive hepatopathy. RESULTS: Liver fibrosis was mainly induced in the periphery of the liver and co-localized with distribution of liver congestion. Liver tumors were also induced in the periphery of the liver where liver congestion and fibrosis occurred. LMD-based RNA-sequencing revealed the upregulation of extracellular matrix/collagen fibril-, wound healing-, angiogenesis-, morphogenesis-, and cell motility-related signaling pathways in periphery of liver compared with liver center. CONCLUSIONS: Our findings showed the experimental relevance of liver congestion, fibrosis, and tumor development in congestive hepatopathy, and may provide important locational information. Macroscopic regional disparity observed in this murine model should be considered to manage patients with congestive hepatopathy.

Signature of chronic hepatitis B virus infection in nails and hair.

BACKGROUND: Hepatitis B virus (HBV) is detected in extrahepatic tissues of individuals with HBV infection. Whether nails and hair contain HBV has been unknown. METHODS: We examined two patient groups: those with chronic HBV infection alone (n = 71), and those with both chronic HBV and hepatitis delta virus (HDV) infections (n = 15). HBV DNA in the patients' fingernails and hair were measured by real-time PCR. Hepatitis B surface antigen (HBsAg) of fingernails was evaluated by an enzyme immunoassay. HDV RNA in fingernails was measured by real-time PCR. Immunochemical staining was performed on nails. We used chimeric mice with humanized livers to evaluate the infectivity of nails. RESULTS: Of the 71 pairs of HBV-alone nail and hair samples, 70 (99%) nail and 60 (85%) hair samples were positive for ß-actin DNA. Of those 70 nail samples, 65 (93%) were HBV DNA-positive. Of the 60 hair samples, 49 (82%) were HBV DNA-positive. The serum HBV DNA level of the nail HBV DNA-positive patients was significantly higher than that of the nail HBV DNA-negative patients (p < 0.001). The hair HBV DNA-positive patients' serum HBV DNA level was significantly higher compared to the hair HBV DNA-negative patients (p < 0.001). The nail HBV DNA level was significantly higher than the hair HBV DNA level (p < 0.001). The nails and hair HBV DNA levels were correlated (r = 0.325, p < 0.05). A phylogenetic tree analysis of the complete genome sequence of HBV isolated from nails and hair identified the infection source. Of the 64 nail samples, 38 (59%) were HBsAg-positive. All 15 pairs of chronic HBV/HDV infection nail and hair samples were ß-actin DNA-positive. However, nail HBV DNA was detected in two patients (13%). None of the 15 patients were positive for hair HBV DNA. Nail HDV RNA was detected in three patients (20%). Of the 15 patients, eight (53%) were nail HBsAg-positive. HBsAg and hepatitis delta (HD) antigen were detected in the nails by immunochemical staining. Chimeric mice were not infected with PBS containing HBsAg and HBV DNA elucidated from nails. CONCLUSIONS: Nails and hair were the reservoir of HBV DNA. Moreover, nails can contain HBsAg, HDV RNA, and HD antigen.

Hepatitis B virus DNA in the fingernails and hair of children with acute hepatitis B.

Hepatitis B virus (HBV) DNA is detectable in the nails and hair of patients with chronic HBV infection. However, it remains unclear whether HBV DNA can be detectable in the nails and hair of patients with acute HBV infection. We encountered two cases of children with acute HBV infection. HBV DNA in the nails and hair from the two children was evaluated by real-time PCR. To clarify the characteristics of HBV DNA, full-length HBV genome sequencing and phylogenetic tree analysis were performed. The levels of serum HBV DNA in children of cases 1 and 2 at day 0 were 7.6 Log IU/mL and 7.4 Log IU/mL, respectively. Nail HBV DNA was detected in both children (case 1: 4.6 Log IU/mL at day 0, case 2: 5.5 Log IU/mL at day 14). Moreover, hair HBV DNA was detectable in case 2 (4.0 Log IU/mL at day 14). Serum HBV DNA became undetectable within approximately 3-4 months after the first hospital visit. After the resolution of HBV viremia, nail and hair HBV DNA became undetectable. The sequence analysis of serum, nail and hair HBV DNA showed the same HBV genotype in each case (case1: genotype C, case 2: genotype A). In case 1, 3 nucleotides were different in the full-genome HBV sequence between the serum and nails. In case 2, the full-genome HBV sequences were identical among the serum, nails and hair. In conclusion, HBV DNA was detectable in nails and hair of children with acute HBV infection.

Sphingosine-1-phosphate promotes tumor development and liver fibrosis in mouse model of congestive hepatopathy.

BACKGROUND AND AIMS: Chronic liver congestion reflecting right-sided heart failure (RHF), Budd-Chiari syndrome, or Fontan-associated liver disease (FALD) is involved in liver fibrosis and HCC. However, molecular mechanisms of fibrosis and HCC in chronic liver congestion remain poorly understood. APPROACH AND RESULTS: Here, we first demonstrated that chronic liver congestion promoted HCC and metastatic liver tumor growth using murine model of chronic liver congestion by partial inferior vena cava ligation (pIVCL). As the initial step triggering HCC promotion and fibrosis, gut-derived lipopolysaccharide (LPS) appeared to induce LSECs capillarization in mice and in vitro. LSEC capillarization was also confirmed in patients with FALD. Mitogenic factor, sphingosine-1-phosphate (S1P), was increased in congestive liver and expression of sphingosine kinase 1, a major synthetase of S1P, was increased in capillarized LSECs after pIVCL. Inhibition of S1P receptor (S1PR) 1 (Ex26) and S1PR2 (JTE013) mitigated HCC development and liver fibrosis, respectively. Antimicrobial treatment lowered portal blood LPS concentration, LSEC capillarization, and liver S1P concentration accompanied by reduction of HCC development and fibrosis in the congestive liver. CONCLUSIONS: In conclusion, chronic liver congestion promotes HCC development and liver fibrosis by S1P production from LPS-induced capillarized LSECs. Careful treatment of both RHF and liver cancer might be necessary for patients with RHF with primary or metastatic liver cancer.

Cluster of Differentiation 44 Promotes Liver Fibrosis and Serves as a Biomarker in Congestive Hepatopathy.

Congestive hepatopathy (CH) with chronic passive congestion is characterized by the progression of liver fibrosis without prominent inflammation and hepatocellular damage. Currently, the lack of reliable biomarkers for liver fibrosis in CH often precludes the clinical management of patients with CH. To explore fibrosis biomarkers, we performed proteome analysis on serum exosomes isolated from patients with CH after the Fontan procedure. Exosomal cluster of differentiation (CD)44 levels were increased in patients with CH compared to healthy volunteers and was accompanied by increases in serum levels of soluble CD44 and CD44 expression in the liver. To address the roles of CD44 in CH, we established a mouse model of chronic liver congestion by partial inferior vena cava ligation (pIVCL) that mimics CH by fibrosis progression with less inflammation and cellular damage. In the pIVCL mice, enhanced CD44 expression in hepatic stellate cells (HSCs) and deposition of its ligand hyaluronan were observed in the liver. Blood levels of soluble CD44 were correlated with liver fibrosis. The blockade of CD44 with specific antibody inhibited liver fibrosis in pIVCL mice and was accompanied by a reduction in S100 calcium-binding protein A4 expression following activation of HSCs. Conclusion: Chronic liver congestion promotes fibrosis through CD44. This identifies CD44 as a novel biomarker and therapeutic target of liver fibrosis in patients with CH.

Pharmacotherapy options for managing hepatitis B in children.

INTRODUCTION: To eliminate viral hepatitis by 2030, the World Health Organization (WHO) launched the first global health sector strategy on viral hepatitis, with particular focus given to hepatitis B and C in 2016. To achieve the reduction of mortality in children, it is indispensable to know which children should be treated and how to treat them. AREA COVERED: In this article, the authors review the antiviral treatment of children with chronic hepatitis B virus (HBV) infection including antivirals available for children with chronic HBV infection. EXPERT OPINION: The approvals of nucleos(t)ide analogues (NAs) and pegylated interferon (PEG-IFN) for children have lowered a hurdle to the initiation of antiviral treatment in children. The international guidelines use nearly the same criteria of antiviral treatment for children with chronic HBV infection, but the WHO guidelines provide a cautious stance on the antiviral treatment of children. Not only PEG-IFN but also NAs with a high genetic barrier to drug resistance should be the first-line treatment for children. In settings with limited medical resources, NAs can be the first-line treatment for children. Although the concept of an 'immune-tolerant phase' is challenged, evidence is not sufficient to recommend the treatment of HBeAg-positive immune-tolerant children.

High Dose of Pegylated Interferon for the Treatment of Chronic Hepatitis B in Children Infected With Genotype C.

Chronic hepatitis B virus (HBV) genotype C infection is unlikely to show a good response to interferon (IFN). However, it is unknown whether a high dose of pegylated IFN (PEG-IFN) treatment would be effective for hepatitis B e antigen (HBeAg)-positive children with chronic HBV genotype C infection. Methods: HBeAg-positive children and adolescents with chronic HBV genotype C infection were eligible for this study. To increase the dose of PEG-IFN, all patients received PEG-IFN-α-2a (180 µg) without dose adjustment on the basis of body surface area for 48 weeks and were followed up for 24 weeks after the completion of treatment. Results: Thirteen patients (median age, 9 years) were enrolled prospectively for this study. One patient dropped out, and the remaining 12 patients were evaluated. Of the 12 patients, 11 received PEG-IFN of 180 µg/1.73 m2 or more (median, 287 µg/1.73 m2). Eight (67%) experienced HBeAg seroconversion, and 1 (8%) achieved hepatitis B surface antigen (HBsAg) loss at the end of follow-up. There was a significant difference in the decrease of hepatitis B surface antigen levels from the baseline to week 24 of treatment between the responders and the nonresponders. Serum cytokines and chemokines were measured in 10 patients. The levels of C-X-C motif chemokine ligand 9, 10, 11, and 13 in the responders tended to be higher than those in the nonresponders during the first 24 weeks of treatment. Conclusions: A high dose of PEG-IFN treatment was effective and safe. A decrease in the hepatitis B surface antigen level from baseline to week 24 of treatment might be a predictor of HBeAg seroconversion.

Discordance of hepatitis B vaccination policies for healthcare workers between the USA, the UK, and Germany.

The hepatitis B (HB) vaccine is effective for the prevention of HB virus infection. It has been widely accepted that an anti-HB surface antibody (HBs) level ≥10 mIU/mL is protective against HB virus infection. Although transient infection can occur in individuals who attain a peak level of anti-HBs ≥10 mIU/mL after primary vaccination, long-term follow-up studies show that successful primary vaccination can prevent individuals from acute clinical hepatitis and chronic infection. Healthcare workers (HCWs) are at-risk individuals. Based on the accumulated data, the USA considers an anti-HBs level ≥10 mIU/mL to constitute successful vaccination for HCWs. In contrast, because some anti-HBs assays cannot accurately measure in the low anti-HBs range, including 10 mIU/mL, the UK and Germany consider an anti-HBs level ≥100 mIU/mL to constitute successful vaccination for HCWs. In the USA and UK, a booster dose is unnecessary for HCWs after successful vaccination. In Germany, anti-HBs testing is recommended for HCWs who are at particularly high individual exposure risk 10 years after successful primary immunization, and a booster dose is offered if the anti-HBs level has declined to ˂100 mIU/mL. The differences in the goal of HB vaccination, reliability of anti-HBs assays, and use of booster vaccination cause discordance in HB vaccination policies for HCWs.

Deep sequencing of hepatitis B surface antigen gene in the preserved umbilical cords in immunoprophylaxis failure against mother-to-child HBV transmission.

BACKGROUND: Vaccine escape mutants (VEMs) are one of the causes of breakthrough infections in the mother-to-child transmission of hepatitis B virus (HBV). We hypothesized that VEMs existing as minor populations in the maternal blood are associated with breakthrough infections in children. We sought to determine whether VEMs exist as minor populations in the preserved umbilical cords of children with breakthrough infections. CASE PRESENTATION: Two families (Family 1: three children, Family 2: two children) were enrolled. Despite immunoprophylaxis, a breakthrough infection occurred in two Family 1 children and two Family 2 children. Preserved umbilical cords, serum, and nails were used for the HBV DNA analysis. To detect VEMs, we performed direct and deep sequencing of hepatitis B surface antigen gene. The direct sequencing showed that there were no VEMs in the serum of the children or mother of Family 1 and family 2, but it identified a G145A mutant in the nails of the mother of Family 2. In Family 1, deep sequencing detected a T143S mutant as a minor population (1.7-2.0%) in the umbilical cords and serum of all three children and in the serum of the mother. A T126A mutant was also detected in the umbilical cord (9.2%) and serum (7.0%) of the first-born child of Family 1. In Family 2, the deep sequencing showed no VEMs in the umbilical cords, but it detected D144A (2.5%) and G145A (11.2%) mutants in the serum of the 2nd-born child. CONCLUSIONS: VEMs were present as minor populations in the preserved umbilical cords of children with breakthrough infections. The VEMs did not become major populations after the breakthrough infections. The evolution of VEMs from a minor form to a major form might not be a prerequisite for breakthrough infections in mother-to-child transmission.

Long-term outcomes of pediatric-onset primary sclerosing cholangitis: A single-center experience in Japan.

AIM: Primary sclerosing cholangitis (PSC) is very rare in Japan. Although a large-scale cohort study of 781 pediatric-onset PSC patients in Europe and North America showed that the 5-year survival with native liver was 88%, the long-term outcomes of pediatric-onset PSC in Japan are unknown. Here, we evaluated the clinical outcomes of pediatric-onset PSC in Japan. METHODS: We carried out a retrospective cohort study with a medical records review of pediatric PSC patients diagnosed between 1986 and 2017 at a single center. The PSC diagnoses were based on cholangiography, liver histology, and biochemical findings. The patients' survival was analyzed using the Kaplan-Meier method. Prognostic factors were determined by univariate and multivariate analyses using the Cox proportional hazards regression model. RESULTS: We identified 39 pediatric-onset PSC patients (22 boys, 17 girls). The median age at diagnosis was 9 years (interquartile range 6.0-13.5 years). The median follow-up period was 5.5 years (interquartile range 3.4-8.7 years). The phenotypes of PSC-autoimmune hepatitis, PSC-inflammatory bowel disease, and small-duct PSC were diagnosed in 13 (33.3%), 36 out of 38 (94.8%), and three (7.7%) patients, respectively. The 5-year liver transplantation-free survival of the whole cohort was 93.5%. Nine patients underwent liver transplantation, and four of these nine cases resulted in death. Both the univariate and multivariate analyses showed that the phenotype of "PSC-autoimmune hepatitis overlap" was an independent poor prognostic factor. CONCLUSIONS: The overall survival of pediatric-onset PSC in Japan was comparable to those in Western countries. The phenotype of PSC-autoimmune hepatitis was identified as a prognostic factor associated with a poorer long-term outcome.

Liver disease secondary to congenital heart disease in children.

Introduction: Hepatic fibrosis and hepatocellular carcinoma (HCC) can develop in children with congenital heart disease. Although hepatic fibrosis and HCC are prone to develop after the Fontan operation, they can also develop in patients suffering from congenital heart disease who have not undergone Fontan operation. Area covered: The history of cardiac hepatopathy including Fontan-associated liver disease is described. Patient characteristics, liver histology, imaging examinations and blood tests are reviewed to elucidate the mechanism of cardiac hepatopathy. In addition, a flowchart for the follow-up management of cardiac hepatopathy in children with congenital heart disease is proposed. Expert opinion: Congestion and low cardiac output are the main causes of cardiac hepatopathy. Advanced hepatic fibrosis is presumed to be associated with HCC. HCC can develop in both adolescents and young adults. Regardless of whether the Fontan operation is performed, children with a functional single ventricle and chronic heart failure should be regularly examined for cardiac hepatopathy. There is no single reliable laboratory parameter to accurately detect cardiac hepatopathy; hepatic fibrosis indices and elastography have shown inconsistent results for detection of this disease. Further studies using liver specimen-confirmed patients and standardization of evaluation protocols are required to clarify the pathogenesis of cardiac hepatopathy.

Loss of fibrocystin promotes interleukin-8-dependent proliferation and CTGF production of biliary epithelium.

BACKGROUND & AIMS: Congenital hepatic fibrosis (CHF) is a genetic liver disease resulting in abnormal proliferation of cholangiocytes and progressive hepatic fibrosis. CHF is caused by mutations in the PKHD1 gene and the subsequent dysfunction of the protein it encodes, fibrocystin. However, the underlying molecular mechanism of CHF, which is quite different from liver cirrhosis, remains unclear. This study investigated the molecular mechanism of CHF pathophysiology using a genetically engineered human induced pluripotent stem (iPS) cell model to aid the discovery of novel therapeutic agents for CHF. METHODS: PKHD1-knockout (PKHD1-KO) and heterozygously mutated PKHD1 iPS clones were established by RNA-guided genome editing using the CRISPR/Cas9 system. The iPS clones were differentiated into cholangiocyte-like cells in cysts (cholangiocytic cysts [CCs]) in a 3D-culture system. RESULTS: The CCs were composed of a monolayer of cholangiocyte-like cells. The proliferation of PKHD1-KO CCs was significantly increased by interleukin-8 (IL-8) secreted in an autocrine manner. IL-8 production was significantly elevated in PKHD1-KO CCs due to mitogen-activated protein kinase pathway activation caused by fibrocystin deficiency. The production of connective tissue growth factor (CTGF) was also increased in PKHD1-KO CCs in an IL-8-dependent manner. Furthermore, validation analysis demonstrated that both the serum IL-8 level and the expression of IL-8 and CTGF in the liver samples were significantly increased in patients with CHF, consistent with our in vitro human iPS-disease model of CHF. CONCLUSIONS: Loss of fibrocystin function promotes IL-8-dependent proliferation of, and CTGF production by, human cholangiocytes, suggesting that IL-8 and CTGF are essential for the pathogenesis of CHF. IL-8 and CTGF are candidate molecular targets for the treatment of CHF. LAY SUMMARY: Congenital hepatic fibrosis (CHF) is a genetic liver disease caused by mutations of the PKHD1 gene. Dysfunction of the protein it encodes, fibrocystin, is closely associated with CHF pathogenesis. Using an in vitro human induced pluripotent stem cell model and patient samples, we showed that the loss of fibrocystin function promotes proliferation of cholangiocytes and the production of connective tissue growth factor (CTGF) in an interleukin 8 (IL-8)-dependent manner. These results suggest that IL-8 and CTGF are essential for the pathogenesis of CHF.

Serum Amino Acid Profiling in Citrin-Deficient Children Exhibiting Normal Liver Function During the Apparently Healthy Period.

BACKGROUND: Citrin (mitochondrial aspartate-glutamate transporter) deficiency causes the failures in both carbohydrate-energy metabolism and the urea cycle, and the alterations in the serum levels of several amino acids in the stages of newborn (NICCD) and adult (CTLN2). However, the clinical manifestations are resolved between the NICCD and CTLN2, but the reasons are still unclear. This study evaluated the serum amino acid profile in citrin-deficient children during the healthy stage. METHODS: Using HPLC-MS/MS analysis, serum amino acids were evaluated among 20 citrin-deficient children aged 5-13 years exhibiting normal liver function and 35 age-matched healthy controls. RESULTS: The alterations in serum amino acids characterized in the NICCD and CTLN2 stages were not observed in the citrin-deficient children. Amino acids involved in the urea cycle, including arginine, ornithine, citrulline, and aspartate, were comparable in the citrin-deficient children to the respective control levels, but serum urea was twofold higher, suggestive of a functional urea cycle. The blood sugar level was normal, but glucogenic amino acids and glutamine were significantly decreased in the citrin-deficient children compared to those in the controls. In addition, significant increases of ketogenic amino acids, branched-chain amino acids (BCAAs), a valine intermediate 3-hydroxyisobutyrate, and ß-alanine were also found in the citrin-deficient children. CONCLUSION: The profile of serum amino acids in the citrin-deficient children during the healthy stage showed different characteristics from the NICCD and CTLN2 stages, suggesting that the failures in both urea cycle function and energy metabolism might be compensated by amino acid metabolism. SYNOPSIS: In the citrin-deficient children during the healthy stage, the characteristics of serum amino acids, including decrease of glucogenic amino acids, and increase of ketogenic amino acids, BCAAs, valine intermediate, and ß-alanine, were found by comparison to the age-matched healthy control children, and it suggested that the characteristic alteration of serum amino acids may be resulted from compensation for energy metabolism and ammonia detoxification.

Cholesterol Metabolism Is Enhanced in the Liver and Brain of Children With Citrin Deficiency.

Context: Citrin-deficient infants present neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD), which resolves at 12 months. Thereafter, they have normal liver function associated with hypercholesterolemia, and a preference for lipid-rich carbohydrate-restricted diets. However, some develop adult-onset type II citrullinemia, which is associated with metabolic abnormalities. Objectives: To identify the causes of hypercholesterolemia in citrin-deficient children post-NICCD. Design and Setting: We determined the concentrations of sterol markers of cholesterol synthesis, absorption, and catabolism by liquid chromatography-electrospray ionization-tandem mass spectrometry and evaluated serum lipoprotein profiles. Subjects: Twenty citrin-deficient children aged 5 to 13 years and 37 age-matched healthy children. Intervention: None. Main Outcome Measures: Relationship between serum lipoproteins and sterol markers of cholesterol metabolism. Results: The citrin-deficient group had a significantly higher high-density lipoprotein cholesterol (HDL-C) concentration than did the control group (78 ± 11 mg/dL vs 62 ± 14 mg/dL, P < 0.001), whereas the two groups had similar low-density lipoprotein cholesterol and triglyceride concentrations. The concentrations of markers of cholesterol synthesis (lathosterol and 7-dehydrocholesterol) and bile acids synthesis (7α-hydroxycholesterol and 27-hydroxycholesterol) were 1.5- to 2.8-fold and 1.5- to 3.9-fold, respectively, higher in the citrin-deficient group than in the control group. The concentration of 24S-hydroxycholesterol, a marker of cholesterol catabolism in the brain, was 2.5-fold higher in the citrin-deficient group. In both groups, the HDL-C concentration was significantly positively correlated with that of 27-hydroxycholesterol, the first product of the alternative bile acid synthesis pathway. Conclusions: HDL-C and sterol marker concentrations are elevated in citrin-deficient children post-NICCD. Moreover, cholesterol synthesis and elimination are markedly enhanced in the liver and brain of citrin-deficient children.

Treatment with medium chain fatty acids milk of CD36-deficient preschool children.

OBJECTIVE: CD36 deficiency is characterized by limited cellular long chain fatty acid uptake in the skeletal and cardiac muscles and often causes energy crisis in these muscles. However, suitable treatment for CD36 deficiency remains to be established. The aim of this study was to evaluate the clinical and metabolic effects of medium chain triacylglycerols (MCTs) in two CD36-deficient preschool children who often developed fasting hypoglycemia and exercise-induced myalgia. METHODS: Fasting blood glucose, total ketone bodies, and free fatty acids were examined and compared for usual supper diets and for diets with replacement of one component with 2 g/kg of 9% MCT-containing milk (MCT milk). Changes in serum creatine kinase and alanine aminotransferase levels, resulting from replacement of glucose water intake with 1 g/kg of MCT milk and determined by using bicycle pedaling tasks, were examined and compared. Hypoglycemic and/or myalgia episodes in daily life were also investigated. RESULTS: Biochemically, participants' blood glucose and total ketone bodies levels after overnight fasting substantially increased after dietary suppers containing MCT milk. Increases in serum creatine kinase and alanine aminotransferase levels resulting from the bicycle pedaling task were suppressed by MCT milk. Hypoglycemia leading to unconsciousness and tachycardia before breakfast decreased after introduction of dietary suppers containing MCT milk. Occurrence of myalgia in the lower limbs also decreased after intakes of MCT milk before long and/or strenuous exercising. CONCLUSION: Our results suggest that MCTs can prevent fasting hypoglycemia and exercise-induced myalgia in CD36-deficient young children.

Treatment with granulocyte colony-stimulating factor in the refeeding phase of anorexia nervosa complicated with severe neutropenia and sepsis: a case report.

Leukopenia is frequently observed in patients with anorexia nervosa (AN). However, serious infectious diseases are infrequent in patients with AN. We present the case of a 13-year-old girl with AN complicated with severe neutropenia and sepsis upon refeeding. Her body mass index was 11 kg/m2 and her absolute neutrophil count (ANC) was 1555/µL on admission. After the initiation of refeeding, her ANC gradually declined to 346/µL. High fever occurred and a blood culture tested positive for Enterobacter cloacae. Although fever subsided with administration of antibiotics, human recombinant granulocyte colony-stimulating factor (G-CSF) was administered to improve her severe neutropenia. After administration of G-CSF, initially for 5 days and again for 3 days, her ANC became normal and stable. The combination of neutropenia and low body mass index may contribute to severe bacterial infection. Usually, hematological abnormalities such as neutropenia improve spontaneously with the recovery of nutritional status in AN patients. Therefore, it is difficult to determine the indication for treatment with G-SCF. Although a careful consideration is required, treatment with G-CSF is effective in AN patients with sepsis caused by severe neutropenia.

Usefulness of serum Wisteria floribunda agglutinin-positive Mac-2 binding protein in children with primary sclerosing cholangitis.

BACKGROUND: Wisteria floribunda agglutinin-positive Mac-2 binding protein (WFA+ -M2BP) is a novel serum marker of hepatic fibrosis in adults with chronic hepatitis C. However, it remains unclear whether serum WFA+ -M2BP levels are associated with the progression of liver histology in primary sclerosing cholangitis (PSC). METHODS: Twenty-eight children and adolescents with pediatric-onset PSC (male : female patient ratio, 20:8; median age at diagnosis, 9 years) were enrolled in this study. The relation between serum WFA+ -M2BP levels and clinical characteristics was retrospectively evaluated. Moreover, receiver operating characteristic (ROC) analysis was used to determine whether serum WFA+ -M2BP levels could be a reliable marker to identify PSC patients with advanced liver histology. RESULTS: According to the Ludwig classification of liver histological stage, 28 patients were classified into the four stages. The WFA+ -M2BP level, aspartate aminotransferase (AST) to platelet ratio index (APRI), and hyaluronic acid correlated significantly with liver histological stage. Moreover, WFA+ -M2BP showed a significant positive correlation (P < 0.05) with autoimmune hepatitis overlap, AST, alanine aminotransferase (ALT), γ-glutamyltransferase, total bilirubin, immunoglobulin G, APRI, and hyaluronic acid. A ROC analysis was undertaken to distinguish the patients with advanced stage disease (stage 3-4) from those with early stage disease (stage 0-2). It showed that WFA+ -M2BP yielded the highest area under the ROC curve value (0.898) among four surrogate makers (APRI, 0.850; Fibrosis-4 index, 0.806; and AST/ALT ratio, 0.802). Moreover, WFA+ -M2BP yielded the highest sensitivity, specificity, positive predictive value, and negative predictive value among the four markers. CONCLUSIONS: Serum WFA+ -M2BP levels are useful to identify patients with advanced liver histology in pediatric PSC.