Complete Genome Sequence of Streptococcus mitis Strain Nm-65, Isolated from a Patient with Kawasaki Disease.

Streptococcus mitis Nm-65 is a human commensal streptococcal strain of the mitis group that was isolated from the tooth surface of a patient with Kawasaki disease. The complete genome sequence of Nm-65 was obtained by means of hybrid assembly, using two next-generation sequencing data sets. The final assembly size was 2,085,837 bp, with 2,039 coding sequences.

Cytotoxic property of Streptococcus mitis strain producing two different types of cholesterol-dependent cytolysins.

Streptococcus mitis strain Nm-65 secretes an atypical 5-domain-type cholesterol-dependent cytolysin (CDC) called S. mitis-derived human platelet aggregation factor (Sm-hPAF) originally described as a platelet aggregation factor. Sm-hPAF belongs to Group III CDC that recognize both membrane cholesterol and human CD59 as the receptors, and shows preferential activity towards human cells. Draft genome analyses have shown that the Nm-65 strain also harbors a gene encoding another CDC called mitilysin (MLY). This CDC belongs to Group I CDC that recognizes only membrane cholesterol as a receptor, and it is a homolog of the pneumococcal CDC, pneumolysin. The genes encoding each CDC are located about 20 kb apart on the Nm-65 genome. Analysis of the genomic locus of these CDC-encoding genes in silico showed that the gene encoding Sm-hPAF and the region including the gene encoding MLY were both inserted into a specific locus of the S. mitis genome. The results obtained using deletion mutants of the gene(s) encoding CDC in Nm-65 indicated that each CDC contributes to both hemolysis and cytotoxicity, and that MLY is the major hemolysin/cytolysin in Nm-65. The present study aimed to determine the potential pathogenicity of an S. mitis strain that produces two CDC with different receptor recognition properties and secretion modes.

Adipose-derived stromal cells grown on a hydroxyapatite scaffold can support hematopoiesis in regenerated bone marrow in vivo.

Osteoblastic cells are a key component of the bone marrow (BM) stem cell niche and help regulate hematopoietic stem cells (HSCs). We have previously demonstrated that adipose-derived stromal cells (ADSCs) can differentiate into both osteogenic and chondrogenic cells in vitro. The current study examined whether the anatomical architecture of the BM could be regenerated in vivo by using ADSCs cultured on a hydroxyapatite (HA) scaffold. ADSCs from GFP transgenic mice were cultured in vitro on an HA scaffold. The scaffold with the attached cells was implanted subcutaneously onto the backs of C57/BL6 (Ly5.2) recipient mice. Lineage-negative (Lin-) Ly5.1 BM cells transduced with a lentiviral vector containing the luciferase (Luc) gene were intravenously administered to the recipient mice after lethal irradiation. Eight weeks after BM transplantation, the scaffolds were removed from the first recipient mice and subcutaneously implanted into lethally irradiated second recipient mice. The biodistribution and kinetics of Luc(+) Ly5.1 cells were monitored by bioluminescence imaging and flow cytometry. Luc(+) hematopoietic cells were present in the scaffolds of the secondary implanted mice for at least 8 months. Subcutaneous injection of G-CSF resulted in wide distribution of bioluminescence signals from the original scaffolds to the whole body. Therefore, BM regenerated using ADSCs grown on an HA scaffold can support HSC populations in vivo, suggesting that a functional BM niche is reconstituted. These results may have a significant impact on the development of therapeutic strategies for various hematopoietic diseases.

Efficacy of eculizumab in a patient with paroxysmal nocturnal hemoglobinuria requiring transfusions 14 years after a diagnosis in childhood.

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare, acquired clonal disorder characterized by chronic complement-mediated hemolysis. The humanized anti-C5 antibody eculizumab binds to the C5 protein and suppresses hemolysis by inhibiting C5b-9 generation. Here, we report on a 27-year-old woman who was found to have PNH in 1997 (at 13 years of age), without subsequent transfusions, thrombosis, or renal disorder. She had been experiencing frequent malaise and fatigue and was sometimes unable to participate in social activities. She had also experienced repeated hemolytic episodes due to infection, and the hemoglobin level had decreased from 7.0 to 5.0 g/dL several times since February 2010. Red blood cell transfusion was necessary, and 6 months later, treatment with eculizumab was started. The hemoglobin level stabilized, and the patient became transfusion-independent. Furthermore, the patient showed significant improvements in fatigue scale scores and quality of life. Six months after the start of eculizumab therapy, the percentage of PNH-type red blood cells was found to have increased from 82.0% (1.95 × 10(12) cells/L) to 89.1% (2.78 × 10(12) cells/L). Furthermore, during treatment with eculizumab, intravascular hemolysis occurred due to a viral infection accompanied by a high fever. We also observed a persistent elevation in reticulocytes and total bilirubin levels, as well as a persistent reduction in haptoglobin levels. Extravascular hemolytic findings were also observed. Because treatment with eculizumab was started at a young age (27 years) and will be continued for many years, careful observation of the patient is required.

Successful coil embolization for life-threatening hemorrhage in childhood leukemia induction therapy.

An 11-year-old boy was experienced severe life-threatening hemorrhage from a branch of the superior mesenteric artery (SMA) after acute lymphoblastic leukemia induction therapy. The patient had a history of attention deficit hyperactivity disorder (ADHD), diagnosed at 3 years of age. Subsequent to discontinuing his psychotropic medication, the patient's mental status deteriorated and treatment with midazolam for 3 weeks was necessary to allow the completion of the leukemia induction regimen. On day 51, although there was no indication of thrombocytopenia or a coagulation disorder, the patient began to hemorrhage suddenly from anal with resulting hypovolemic shock, and large-volume blood transfusion was initiated. Although upper and lower endoscopy failed to determine the location of the hemorrhage, angiography enabled us to determine that it was a branch of the SMA (the middle colic artery #6), and selective arterial embolization was used to arrest the bleeding. There could have been underlying causes, such as, a probable malformation or aneurysm in that area, although there was no indication before or after the event. This is a rare case of arterial hemorrhage from a branch of the SMA that occurred in a pediatric patient idiopathically during the induction therapy of leukemia.

A case of juvenile Sjögren's syndrome with interstitial nephritis.

Primary Sjögren's syndrome (SS) is a rare autoimmune disease, especially in children. Juvenile primary SS with interstitial nephritis is rare in Japan. We report on a 12-year-old girl in whom salivary gland swelling had recurred from the age of 5 years, SS was diagnosed at the age of 10 years, and interstitial nephritis developed at the age of 12 years. The patient presented with a chief complaint of swelling of both parotid glands. The patient had a history of recurrent parotitis from 5 years of age, with episodes recurring 5 to 6 times a year and resolving within 3 days each time. However, at the age of 11 years, the patient had continuous mild swelling of the parotid glands. Examination on admission showed bilateral nontender parotid gland swelling; mild swelling of the lower extremities, xerostomia, and xerophthalmia but no exanthem. Laboratory findings were as follows: serum protein, 10.1 g/dL; immunoglobulin (Ig) G, 3,828 mg/dL; antinuclear antibodies, 1,280-fold; anti-Ro/SS-A antibody, 512-fold; anti-Ro/SS-B antibody, 4-fold; creatinine, 0.45 mg/dL; blood ß2-microglobulin, 2.2 mg/L (slightly elevated); and cystatin C, 0.86 mg/L. Urinalysis showed proteinuria and a ß2-microglobulin concentration of 11,265 mg/L. Thus, this patient had low molecular weight proteinuria. Schirmer's test showed decreased tear secretion (5 mm), and fluorescein staining showed marked bilateral superficial punctate keratitis. A lip biopsy showed infiltration by small round cells (mild to moderate), interstitial fibrosis, loss of salivary gland parenchyma, and atrophy, with no obvious epimyoepithelial islands, leading to a diagnosis of SS. Light microscopic examination of the renal biopsy specimens showed expansion of mononuclear cell infiltration in the renal interstitium, inflammatory cell infiltration of interstitial areas with edema and mild fibrosis, and tubulitis and mononuclear cell infiltration that included many lymphocytes and plasma cells. There were no pathological findings of glomerulonephritis. Small arteries showed no obvious abnormalities. Immunofluorescent staining showed slight, nonspecific deposition of IgM, but no deposition of IgG, complement 1q, 3, or 4. On the basis of the renal biopsy showing nonspecific chronic interstitial nephritis, renal tubular atrophy, and interstitial enlargement, tubulointerstitial nephritis associated with SS was diagnosed.

Mean hemoglobin levels in venous blood samples and prevalence of anemia in Japanese elementary and junior high school students.

Screening for anemia has been performed in schools in Japan for over 30 years. The long-term effect of the nuclear power plant disaster on the prevalence of anemia in school age children is unknown. This research was performed to evaluate the prevalence of anemia in school age children and to determine grade-level and gender-related reference hemoglobin (Hb) levels prior to the nuclear disaster. Data for this research were obtained from results of screening for anemia obtained by venous blood sampling in schools in 2002. Mean Hb levels were calculated for each grade level (elementary school grades 1-6 and junior high school years 1-3) and according to gender, and the prevalence of anemia was determined. In our research, Tokyo Health Service Association guidelines were used to determine reference Hb levels for anemia. We demonstrated that Hb levels in boys increased with age during childhood and adolescence (from 13.1 ± 0.7 g/dL in 7 year olds to 14.9 ± 1.1 g/dL in 15 year olds); in girls, Hb levels peaked at menarche (13.7 ± 0.8 g/dL in 12 year olds), decreasing slightly thereafter (13.4 ± 1.1 g/dL in 15 year olds). The prevalence of anemia was 0.26% in elementary school boys, 0.27% in elementary school girls, and 1.21% in junior high school boys. The prevalence of anemia in second- and third-year junior high school girls was lower than that in first-year junior high school girls. Among all junior high school girls, 5.73% had mild anemia. Iron-deficiency anemia is the commonest type of anemia in high school girls, secondary to the relative lack of iron due to menstruation, the growth spurt and exercise. Appropriate dietary therapy and treatment of anemia, together with education about the dietary prevention of anemia, are important to reduce the prevalence of anemia in high school students. When complete blood counts are performed in regions thought to be affected by the Fukushima nuclear power plant disaster, our report can serve as a reference during evaluation of Hb levels.

HTLV-I-infected breast milk macrophages inhibit monocyte differentiation to dendritic cells.

Human T-cell leukemia virus type I (HTLV-I), a causative agent of adult T-cell leukemia (ATL), is transmitted from mother to child, predominantly by breastfeeding. Oral HTLV-I infection and infection early in life are associated with a subsequent risk of ATL. Although the pathogenic mechanisms of ATL remain largely unknown, the host immune system seems to play an important role in HTLV-I pathogenesis. Previous studies have shown that monocytes from ATL patients had reduced capacity for dendritic cell (DC) differentiation. Therefore, we performed the present study to clarify the mechanisms responsible for the impairment of DC differentiation using HTLV-I-infected breast milk macrophages (HTLV-BrMMø). We found that when CD14⁺ monocytes were cultured with GM-CSF and IL-4 in the presence of HTLV-BrMMø, they altered the surface phenotype of immature DCs and the stimulatory capacity of T-cell proliferation. The presence of HTLV-BrMMø significantly blocked the increased expression of CD1a, CD1b, CD11b, DC-SIGN, and HLA-DR; however, increased expression of CD1d and CD86 was observed. These effects could be partially replicated by incubation with culture supernatants from HTLV-BrMMø. The impairment of monocyte differentiation might be not due to HTLV-I infection of monocytes, but might be due to unknown soluble factors. Since other HTLV-I-infected cells exhibited similar inhibitory effects on monocyte differentiation to DCs, we speculated that HTLV-I infection might cause the production of some inhibitory cytokines in infected cells. Identifying the factors responsible for the impairment of monocyte differentiation to DCs may be helpful to understand HTLV-I pathogenesis.

A neonate with reduced cytomegalovirus DNA copy number and marked improvement of hearing in the treatment of congenital cytomegalovirus infection.

Congenital cytomegalovirus (CMV) infection can cause severe permanent disabilities. A mother who is seronegative before conception but acquires infection during pregnancy is a risk factor for congenital infection. We describe a neonate in whom congenital CMV infection was diagnosed at birth and confirmed with DNA quantitation by means of the polymerase chain reaction, was accompanied by cerebral ventriculomegaly and severe hearing loss, and was treated with ganciclovir/valganciclovir for 6 weeks. Initially, cerebral ventriculomegaly and calcification were also found with computed tomography, and severe hearing loss was detected with auditory brainstem response testing. After treatment, CMV DNA decreased in copy number and became undetectable. No marked side effects occurred after treatment. Surprisingly, 1 year after treatment, neurological and motor development was equivalent to that in a healthy infant. Audiometry indicated that auditory ability would improve with rehabilitation, speech and language therapy, and cochlear implantation. Single-photon emission computed tomography showed marked improvement 6 months after treatment. This case provides compelling evidence that a reliable diagnosis of congenital CMV infections coupled with a prompt and appropriate treatment program can prevent permanent disability. It is, therefore, important to establish a more effective strategy for the management of congenital CMV infection.

Successful gene therapy in utero for lethal murine hypophosphatasia.

Hypophosphatasia (HPP), caused by mutations in the gene ALPL encoding tissue-nonspecific alkaline phosphatase (TNALP), is an inherited systemic skeletal disease characterized by mineralization defects of bones and teeth. The clinical severity of HPP varies widely, from a lethal perinatal form to mild odontohypophosphatasia showing only dental manifestations. HPP model mice (Akp2(-/-)) phenotypically mimic the severe infantile form of human HPP; they appear normal at birth but die by 2 weeks of age because of growth failure, hypomineralization, and epileptic seizures. In the present study, we investigated the feasibility of fetal gene therapy using the lethal HPP model mice. On day 15 of gestation, the fetuses of HPP model mice underwent transuterine intraperitoneal injection of adeno-associated virus serotype 9 (AAV9) expressing bone-targeted TNALP. Treated and delivered mice showed normal weight gain and seizure-free survival for at least 8 weeks. Vector sequence was detected in systemic organs including bone at 14 days of age. ALP activities in plasma and bone were consistently high. Enhanced mineralization was demonstrated on X-ray images of the chest and forepaw. Our data clearly demonstrate that systemic injection of AAV9 in utero is an effective strategy for the treatment of lethal HPP mice. Fetal gene therapy may be an important choice after prenatal diagnosis of life-threatening HPP.

Fibromyalgia and chronic fatigue syndrome in children.

BACKGROUND: Fibromyalgia (FM) is characterized by widespread persistent pain and the presence of multiple discrete tender points. Chronic fatigue syndrome (CFS) is a syndrome characterized by debilitating fatigue associated with a variable number of non-specific complaints. Because neither condition had necessarily been recognized in children until recently, those patients have been treated as having school refusal without being diagnosed as having either syndrome. There is a considerable overlap of clinical symptoms between these two syndromes. It is therefore controversial as to whether these syndromes have the same pathogenesis or not. The aim of the present study was to clarify the relationship between these syndromes in children. METHODS: Fifteen patients with FM and 21 patients with CFS were investigated both clinically and immunologically. Immunological assessments included thorough analysis of autoantibodies using several techniques. RESULTS: Anti-nuclear antibody titers were higher and the prevalence of anti-Sa antibody was far more frequent in CFS patients than in FM patients. CONCLUSION: CFS and FM are different from each other at least in childhood, from an immunological aspect, although some patients could have both conditions.

Tumor necrosis factor-α-induced mononuclear cell death may contribute to polymorphonuclear cell predominance in the cerebrospinal fluid of patients with bacterial meningitis.

BACKGROUND: Bacterial meningitis is characterized by a marked predominance of polymorphonuclear leukocytes (PMNs: segmented granulocytes or neutrophils) in the cerebrospinal fluid (CSF), whereas aseptic meningitis is characterized by a predominance of mononuclear leukocytes (MNs: lymphocytes or monocytes). However, the pathophysiology of this predominance of PMNs in the CSF of patients with bacterial meningitis has never, to our knowledge, been clearly described. METHODS: To investigate the predominant cell components of CSF from pediatric patients with bacterial meningitis, we investigated cell death in association with levels of tumor necrosis factor-alpha (TNF-α) in the CSF, using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) assay and flow cytometry. RESULTS: The MTT assay of the CSF revealed that the PMNs had survived, while the MNs rapidly had undergone cell death. Although PMNs survived in CSF with high levels of TNF-α, PMN apoptosis was demonstrated with flow cytometry. CONCLUSIONS: The present study suggests that the pathophysiology of PMN predominance in the CSF of patients in the acute phase of bacterial meningitis is related to the rapid cell death of MNs and the survival of PMNs brought about by high levels of TNF-α.

Infantile case of early manifestation of SLE-like symptoms in complete C1q deficiency.

C1q deficiency is a rare complement deficiency in the early part of the complement cascade. Patients with C1q deficiency have severe recurring life-threatening infections and systemic lupus erythematosus (SLE)-like symptoms. We report on a boy with recurrent life-threatening infections and SLE-like recurrent skin conditions before 2 years of age. Immunological studies revealed an undetectable level of C1q. No abnormality was observed in the urine, but renal biopsy showed segmental granulonephritis. However, the changes observed were atypical for SLE nephritis. This case of C1q deficiency was unusual because the SLE-like symptoms appeared earlier than that normally seen in complement deficiency. Therefore, this case provides insights into the development of autoimmune disease, particularly in the early phase of component deficiency, and in managing renal disease that may develop in the future.

Production of autoantibodies by murine B-1a cells stimulated with Helicobacter pylori urease through toll-like receptor 2 signaling.

Helicobacter pylori infection is associated with several autoimmune diseases, in which autoantibody-producing B cells must be activated. Among these B cells, CD5-positive B-1a cells from BALB/c mice were confirmed to secrete autoantibodies when cocultured with purified H. pylori urease in the absence of T cells. To determine the mechanisms for autoantibody production, CD5-positive B-1a cells were sorted from murine spleen cells and stimulated with either purified H. pylori urease or H. pylori coated onto plates (referred to hereafter as plate-coated H. pylori), and autoantibody production was measured by enzyme-linked immunosorbent assay (ELISA). Complete urease was not secreted from H. pylori but was visually expressed over the bacterium-like endotoxin. Urease-positive plated-coated H. pylori stimulated B-1a cells to produce autoantibodies, although urease-deficient isotype-matched H. pylori did not. Autoantibody secretion by B-1a cells was inhibited when bacteria were pretreated with anti-H. pylori urease-specific antibody having neutralizing ability against urease enzymatic activity but not with anti-H. pylori urease-specific antibody without neutralizing capacity. The B-1a cells externally express various Toll-like receptors (TLRs): TLR1, TLR2, TLR4, and TLR6. Among the TLRs, blocking of TLR2 on B-1a cells with a specific monoclonal antibody (MAb), T2.5, inhibited autoantibody secretion when B-1a cells were stimulated with plate-coated H. pylori or H. pylori urease. Moreover, B-1a cells from TLR2-knockout mice did not produce those autoantibodies. The present study provides evidence that functional urease expressed on the surface of H. pylori will directly stimulate B-1a cells via innate TLR2 to produce various autoantibodies and may induce autoimmune disorders.

Identification of a novel biomarker candidate, a 4.8-kDa peptide fragment from a neurosecretory protein VGF precursor, by proteomic analysis of cerebrospinal fluid from children with acute encephalopathy using SELDI-TOF-MS.

BACKGROUND: Acute encephalopathy includes rapid deterioration and has a poor prognosis. Early intervention is essential to prevent progression of the disease and subsequent neurologic complications. However, in the acute period, true encephalopathy cannot easily be differentiated from febrile seizures, especially febrile seizures of the complex type. Thus, an early diagnostic marker has been sought in order to enable early intervention. The purpose of this study was to identify a novel marker candidate protein differentially expressed in the cerebrospinal fluid (CSF) of children with encephalopathy using proteomic analysis. METHODS: For detection of biomarkers, CSF samples were obtained from 13 children with acute encephalopathy and 42 children with febrile seizure. Mass spectral data were generated by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) technology, which is currently applied in many fields of biological and medical sciences. Diagnosis was made by at least two pediatric neurologists based on the clinical findings and routine examinations. All specimens were collected for diagnostic tests and the remaining portion of the specimens were used for the SELDI-TOF MS investigations. RESULTS: In experiment 1, CSF from patients with febrile seizures (n = 28), patients with encephalopathy (n = 8) (including influenza encephalopathy (n = 3), encephalopathy due to rotavirus (n = 1), human herpes virus 6 (n = 1)) were used for the SELDI analysis. In experiment 2, SELDI analysis was performed on CSF from a second set of febrile seizure patients (n = 14) and encephalopathy patients (n = 5). We found that the peak with an m/z of 4810 contributed the most to the separation of the two groups. After purification and identification of the 4.8-kDa protein, a 4.8-kDa proteolytic peptide fragment from the neurosecretory protein VGF precursor (VGF4.8) was identified as a novel biomarker for encephalopathy. CONCLUSIONS: Expression of VGF4.8 has been reported to be decreased in pathologically degenerative changes such as Alzheimer's disease, amyotrophic lateral sclerosis (ALS), frontotemporal dementia, and encephalopathy. Thus, the VGF4.8 peptide might be a novel marker for degenerative brain conditions.

High mobility group box 1 in cerebrospinal fluid from several neurological diseases at early time points.

The present study aimed to elucidate the possible role of High Mobility Group Box 1 (HMGB1), which is a candidate prognostic marker in diseases that combine inflammation and tissue injury, in acute encephalopathy. HMGB1 in cerebrospinal fluid (CSF) obtained on admission from eight children with acute encephalopathy, and 16 children with febrile seizure, eight children with bacterial/aseptic meningitis, and eight children with fever without neurological symptoms were analyzed using enzyme-linked immunosorbent assay (ELISA). We found no increase in HMGB1 in CSF from acute encephalopathy or in CSF from febrile seizure or fever without neurological complications at early time points, while marked elevation of HMGB1 was seen in CSF from bacterial and aseptic meningitis. In conclusion, HMGB1 is a poor disease marker for acute encephalopathy.

A 6-year-old girl with hemoglobin H disease.

Hemoglobin H (HbH) disease is the severe nonfatal form of α-thalassemia syndrome. It is usually caused by molecular defects of 3 of 4 α-globin genes (--/-α) which cause α-globin expression to be decreased. HbH disease is rare in Japan. Here, we report on a 6-year-old girl with HbH disease who had profound hypochromatic and microcytic anemia. Analysis of the α-globin genes of the patient's family showed that the father, who was Japanese, had an abnormal gene with a 3.7-kb deletion (-α(3.7)/αα), and the mother, who was Filipino, had a deletion removing both α-globin genes of the Filipino type (--(FIL)/αα). Neither parent had anemia. The patient was found to have HbH disease with a heterozygous genetic abnormality (--(FIL)/-α(3.7)). Recently, the number of marriages of Japanese to natives of areas where thalassemia is epidemic has increased. Therefore, the incidence of HbH disease can be expected to increase in Japan. Long-term follow-up will be needed to evaluate the long-term complications and to improve the quality of life of patients with HbH disease.

Enhanced expression of cytokines/chemokines in cerebrospinal fluids in mumps meningitis in children.

BACKGROUND: The mumps virus is frequently the causative agent in aseptic meningitis and mumps has still prevailed in Japan. We compared data obtained from patients with mumps meningitis and patients with aseptic meningitis caused by other viruses in order to identify mumps meningitis-specific cytokine/chemokine alterations in cerebrospinal fluid (CSF). METHODS: We elucidated the cytokine/chemokine network based on the cytokine/chemokine profiles in CSF from children with mumps meningitis and meningitis due to other viral infections using multiplex cytokine measurement. Seventeen cytokines/chemokines, namely interleukin (IL)-1ß, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12 (p70), IL-13, IL-17, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, granulocyte colony-stimulating factor (G-CSF), granulocyte monocyte colony-stimulating factor (GM-CSF), monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1ß (MIP-1ß), were measured simultaneously in CSF supernatants from eight children with mumps meningitis, 11 children with other types of viral meningitis and eight children with fever without neurological complications such as convulsion. RESULTS: We found that IL-8, IL-10, IL-12, IL-13 and IFN-γ showed a statistically significant increase in CSF from mumps meningitis when compared to other types of viral meningitis and fever without neurological complications. CONCLUSION: Mumps meningitis may induce a distinct immunological response when compared with other types of viral meningitis.

Rescue of severe infantile hypophosphatasia mice by AAV-mediated sustained expression of soluble alkaline phosphatase.

Hypophosphatasia (HPP) is an inherited disease caused by a deficiency of tissue-nonspecific alkaline phosphatase (TNALP). The major symptom of human HPP is hypomineralization, rickets, or osteomalacia, although the clinical severity is highly variable. The phenotypes of TNALP knockout (Akp2(-/-)) mice mimic those of the severe infantile form of HPP. Akp2(-/-) mice appear normal at birth, but they develop growth failure, epileptic seizures, and hypomineralization and die by 20 days of age. Previously, we have shown that the phenotype of Akp2(-/-) mice can be prevented by enzyme replacement of bone-targeted TNALP in which deca-aspartates are linked to the C-terminus of soluble TNALP (TNALP-D10). In the present study, we evaluated the therapeutic effects of adeno-associated virus serotype 8 (AAV8) vectors that express various forms of TNALP, including TNALP-D10, soluble TNALP tagged with the Flag epitopes (TNALP-F), and native glycosylphosphatidylinositol-anchored TNALP (TNALP-N). A single intravenous injection of 5×10(10) vector genomes of AAV8-TNALP-D10 into Akp2(-/-) mice at day 1 resulted in prolonged survival and phenotypic correction. When AAV8-TNALP-F was injected into neonatal Akp2(-/-) mice, they also survived without epileptic seizures. Interestingly, survival effects were observed in some animals treated with AAV8-TNALP-N. All surviving Akp2(-/-) mice showed a healthy appearance and a normal activity with mature bone mineralization on X-rays. These results suggest that sustained alkaline phosphatase activity in plasma is essential and sufficient for the rescue of Akp2(-/-) mice. AAV8-mediated systemic gene therapy appears to be an effective treatment for the infantile form of human HPP.

An 8-year-old girl with a recurrence of obstructive sleep apnea syndrome caused by hypertrophy of tubal tonsils 4 years after adenotonsillectomy.

Obstructive sleep apnea syndrome affects 1% to 2% of children. It is caused mainly by upper airway obstruction and manifests as snoring and sleep disturbance. Adenotonsillectomy can improve quality of life because airway obstruction occurs when both tonsils and adenoids are enlarged. We describe an 8-year-old girl with a recurrence of obstructive sleep apnea syndrome caused by hypertrophy of the tubal tonsils 4 years after adenotonsillectomy. The findings from this case highlight the importance of 1) identifying hypertrophy of the residual adenoid and compensatory hypertrophy of the tubal tonsils in patients with obstructive sleep apnea syndrome after adenotonsillectomy and 2) determining the optimal timing of adenotonsillectomy with respect to both the severity of obstructive sleep apnea and compensatory hypertrophy of other lymphoid tissue of Waldeyer's ring, as growth of such tissues is most active during the first several years of life.