Yutaka Kanamori

Role of Gut Cryptopatches in Early Extrathymic Maturation of Intestinal Intraepithelial T Cells

Lympho-hemopoietic progenitors residing in murine gut cryptopatches (CP) have been shown to generate intestinal intraepithelial T cells (IEL). To investigate the role of CP in progenitor maturation, we analyzed IEL in male mice with a truncated mutation of common cytokine receptor γ-chain (CRγ−/Y) in which CP were undetectable. IEL-expressing TCR-γδ (γδ-IEL) were absent, and a drastically reduced number of Thy-1highCD4+ and Thy-1highCD8αβ+ αβ-IEL were present in CRγ−/Y mice, whereas these αβ-IEL disappeared from athymic CRγ−/Y littermate mice. Athymic CRγ−/Y mice possessed a small TCR- and αEβ7 integrin-negative IEL population, characterized by the disappearance of the extrathymic CD8αα+ subset, that expressed pre-Tα, RAG-2, and TCR-Cβ but not CD3ε transcripts. These TCR− IEL from athymic CRγ−/Y mice did not undergo Dβ-Jβ and Vδ-Jδ joinings, despite normal rearrangements at the TCR-β and -δ loci in thymocytes from euthymic CRγ−/Y mice. In contrast, athymic severe combined immunodeficient mice in which CP developed normally possessed two major TCR−αEβ7+ CD8αα+ and CD8− IEL populations that expressed pre-Tα, RAG-2, TCR-Cβ, and CD3ε transcripts. These findings underscore the role of gut CP in the early extrathymic maturation of CD8αα+ IEL, including cell-surface expression of αEβ7 integrin, CD3ε gene transcription, and TCR gene rearrangements.

An animal model study for tissue-engineered trachea fabricated from a biodegradable scaffold using chondrocytes to augment repair of tracheal stenosis

INTRODUCTION We have designed an engineered graft fabricated from a biodegradable scaffold using chondrocytes and applied this construct to augment repair of tracheal stenosis. This study investigated the feasibility of using such tissue-engineered airways with autologous chondrocytes in a rabbit model. MATERIAL AND METHODS Chondrocytes were isolated and expanded from the auricular cartilage of New Zealand white rabbits, then seeded onto composite 3-layer scaffolds consisting of a collagen sheet, a polyglycolic acid mesh, and a copolymer (l-lactide/epsilon-caprolactone) coarse mesh. The engineered grafts were implanted into a 0.5 x 0.8-cm defect created in the midventral portion of the cervical trachea. Gelatin sponges that slowly released basic fibroblast growth factor (b-FGF) were then placed on the constructs, which were retrieved 1 or 3 months after implantation. RESULTS The biodegradable scaffold seeded with chondrocytes could maintain airway structure up to 3 months after implantation. Tracheal epithelial regeneration occurred in the internal lumen of this composite scaffold. Three months after implantation, staining of the sections showed cartilage accumulation in the engineered tracheal wall. CONCLUSION This composite biodegradable scaffold may be useful for developing engineered trachea. A gelatin sponge slowly releasing b-FGF might enhance chondrogenesis.

Combination therapy with Bifidobacterium breve, Lactobacillus casei, and galactooligosaccharides dramatically improved the intestinal function in a girl with short bowel syndrome: a novel synbiotics therapy for intestinal failure.

It has been well known since ancient times that fermented milk produces beneficial effects on the consumer’s health. In the last few decades, these beneficial effects have been demonstrated to be due to the metabolic action of some bacterial species, including lactobaccilli, bifidobacteria, and streptococci (1–3). Lilly et al. first introduced the term probiotics for such bacteria in 1965 (4). Probiotics are widely used as a live microbial feed supplement that beneficially affects the host animals by improving their intestinal microbial balance (5). Additionally the term prebiotics has been adopted to refer to a nondigestive food ingredient that selectively targets the growth and/or activity of one or a limited number of bacteria in the colon and, thus, has the potential to improve host health. Several types of ingredients, such as fructooligosaccharides, galactooligosaccharides, and inulin, are used as prebiotics (2, 6). Furthermore, the combined use of probiotics and prebiotics is called synbiotics therapy, but few reports concerning synbiotics have been published (7, 8). Short bowel syndrome refers to seriously adverse symptoms that are seen in patients who have been subjected to a massive bowel resection. These patients are usually malnourished and have a dilated intestine that results in intestinal bacterial overgrowth syndrome (9, 10). Regulation of intestinal bacterial overgrowth, especially pathogenic bacterial overgrowth, is very important in patients with short bowel syndrome to attain any improvement in intestinal function. For this purpose, various antibiotics have been used to eliminate the intestinal bacteria selectively. An alternative strategy for regulating intestinal bacteria is to apply probiotics and/or prebiotics. In this report, we report the use of synbiotics therapy in the treatment of a 4-year-old girl suffering from short bowel syndrome. For the synbiotics therapy, we used Bifidobacterium breve, Lactobacillus casei, and galactooligosaccharides. This novel combination therapy was expected to act synergistically for the improvement of the subject’s health. We found that the patient’s intestinal absorptive function and motility were dramatically improved by this newly designed synbiotics therapy, and she progressed satisfactorily after 2 years of the therapy. Following the case report, we discuss the beneficial effects of the new synbiotics therapy for intestinal failure.

Internal jugular phlebectasia in two siblings: Manometric and histopathologic studies of the pathogenesis

Two brothers, 4 years and 6 years of age, presented with a swelling in the right side of the neck. Ultrasonography and venography confirmed a diagnosis of phlebectasia of the right internal jugular vein (IJV). In an attempt to elucidate the etiology of this rare lesion, venous pressures in both the dilated right IJV and in the left IJV were taken under general anesthesia with intratracheal intubation at the time of surgery in each patient. No significant difference in pressure elevation with increase of intrathoracic pressure by overinflating the breathing bag was observed between the right and left IJV, suggesting that there would be no mechanical obstructive process generated on exertion, in each case. Microscopic examinations of a dissected portion of the dilated IJV showed paucity of muscle layer of the vein wall in the younger patient, and absence of that in the elder. Therefore, we assume that congenital muscle defect of the right IJV wall, rather than mechanical obstruction in the lower neck or the mediastinum, might cause phlebectasia. To our knowledge, this is the first report of IJV phlebectasia in siblings.

New gut associated lymphoid tissue "cryptopatches" breed murine intestinal intraepithelial T cell precursors.

Numerous mouse intraepithelial T cells (IEL) bearing either TCR-αΒ or TCR-γδ have been shown to develop somewhere in the intestinal mucosa without passing through the thymus. However, just where these T cells develop has been much less clear and has remained an open question to date. In an effort to investigate this issue, we carried out immunohistochemical study on the murine gastrointestinal tract and identified numerous tiny lymphoid tissues (~1,650 tissues/intestine) in the cryptal region of the small and large intestinal mucosa except for the stomach in which clusters of c-kit+ IL-7R+ Thy1+ lympho-hemopoietic progenitors accumulated (cryptopatches). The cryptopatch cells isolated from the small intestine, which were c-kit positive (c-kit+) but lineage marker negative (Lin), gave rise to TCR-αΒ and TCR-γδ IELs following in vivo transfer or tissue engraftment into 2 Gy-irradiated severe combined immunodeficient mice. In contrast, cells isolated from Peyer’s patches and mesenteric lymph nodes, which belong in the same intestinal immune compartment but lack c-kit+Lin- cells, failed to do so. These results in conjunction with the findings of electron microscopic analysis provide direct evidence of a local intestinal T cell precursor that develops in the cryptopatches.

The current profile and outcome of congenital diaphragmatic hernia: a nationwide survey in Japan.

BACKGROUND/PURPOSE Few nationwide surveys have been reported regarding the perinatal status, clinical course and postnatal outcome of cases with congenital diaphragmatic hernia (CDH). The aim of this study was to review the current profile and the outcomes of a large cohort of CDH cases in Japan. METHODS A nationwide retrospective cohort study was conducted on neonates diagnosed to have CDH between January 2006 and December 2010. The questionnaires were sent to 159 representative regional institutions and 109 (68.6%) institutions responded to the preliminary survey which had 674 cases. Eleven institutions which had 60 CDH neonates did not respond to the second questionnaire, and 26 institutions had no cases. Finally, 614 CDH neonates from 72 institutions had been collected and were used in the detailed survey. The perinatal status, clinical course and the postnatal outcome were reviewed. Survival was defined as infants alive at hospital discharge, at the time of transfer or still in the hospital at the time of questionnaire, which was confirmed during the period from July 2011 to November 2011 by the investigators. RESULTS Four hundred sixty-three (75.4%) of 614 CDH neonates survived in this study. The overall survival rate of neonates with isolated CDH was 84.0%. A total of 444 (72.0%) patients were prenatally diagnosed, and had a survival rate of 70.8%. Four hundred thirty-three (70.9%) patients were treated with high-frequency oscillatory ventilation (HFOV) as the initial ventilation, 344 (56.0%) patients received inhaled nitric oxide (iNO) and 43 (7.0%) required extracorporeal membrane oxygenation (ECMO). The overall survival rates of the CDH neonates who had been treated using HFOV, iNO and ECMO were 74.3%, 68.3% and 37.2%, respectively. CONCLUSIONS This study demonstrated that the current status for CDH treatment in Japan and the overall survival rate were comparable to those of recent reports from other countries.

Thoracic epidural catheter insertion using the caudal approach assisted with an electrical nerve stimulator in young children

Objectives We evaluated whether thoracic epidural catheter placement using the caudal approach and assisted with an electrical stimulator could be performed in young children. Methods Ten young children (1-4 years) who underwent abdominal surgeries were studied. Under general anesthesia without muscle relaxants, caudal catheter placement was performed using an 18-gauge Crawford-type needle and a 20-gauge radiopaque epidural catheter with a stainless-steel stylet. A metal adapter and a 3-way stopcock were attached to the catheter to connect to an electrical stimulator and to inject physiological saline. Electrical stimulation was performed intermittently while advancing the catheter until it reached the target length. The catheter position was confirmed on postoperative roentgenogram. Results The mean age of the subjects was 32.2 ± 10.1 months (13-48 months), and the height was 85.3 ± 6.1 cm (72-93 cm). In 9 of 10 patients, an epidural catheter could be placed at the first insertion. In 1 patient, the catheter could be placed successfully at the second insertion. The electrical current required for muscle contraction at the target length was 5.8 ± 1.5 mA. Conclusion Electrical stimulation reliably indicated the location of the catheter tip. This technique for thoracic epidural catheter insertion was easy to perform and could be used in young children.

Anaerobic dominant flora was reconstructed by synbiotics in an infant with MRSA enteritis

Many cases of fulminant enteritis caused by Staphylococcus aureus have been reported since the 1950s. 1 Such cases usually had a severe clinical course with intestinal necrosis, perforation and systemic sepsis. 2 Characteristic of most cases is a history of treatment with prophylactic antibiotics after abdominal surgery. 3 Recently, methicillin-sensitive S. aureus was reported to be replaced by methicillin-resistant S. aureus (MRSA) as a cause of severe enteritis worldwide. 4 Vancomycin is currently an effective antibiotic for MRSA infections but recently, the appearance of vancomycin-intermediate MRSA was reported in Japan. 5 It is now imperative to seek a completely different strategy to control antibiotic-resistant pathogenic bacteria in the body. The idea of controlling intestinal bacterial flora using living organisms such as Bifidobacterium , Lactobacillus and Enterococcus was recently proposed, and is a very promising alternative strategy for controlling antibiotic-resistant bacteria in the intestine. 6 Such beneficial living bacteria for host animals are called probiotics and food supplements as an energy source for probiotics are called prebiotics. 7 Furthermore, the combined use of probiotics and prebiotics was proposed to be a more effective treatment than either probiotics or prebiotics used alone; such combinations are called synbiotics. 7

Outcomes of prenatally diagnosed sacrococcygeal teratomas: the results of a Japanese nationwide survey

BACKGROUND/PURPOSE Few large multicenter surveys have been performed on sacrococcygeal teratomas (SCTs) describing both the prenatal and postnatal courses. The aim of this study was to review and report on the prenatal surveillance and postnatal outcome of a large cohort of fetuses with SCTs in Japan. METHODS A nationwide retrospective cohort study was conducted on 97 fetuses prenatally diagnosed with SCTs between 2000 and 2009. The prenatal course, perinatal data, and postnatal outcome were reviewed. RESULTS Eleven pregnancies were terminated before 22 weeks of gestation. Of the 86 remaining fetuses, 3 died in utero, and 83 were delivered. Three infants died before surgery, and 8 infants died after excisional surgery. The overall mortality was 26%, with a mortality excluding terminations of 16%. The gestational age at delivery was younger than 28 weeks in 5, 28 to 31 weeks in 13, 32 to 36 weeks in 27, and 37 weeks or more in 37 cases, with mortality rates of 60%, 38%, 11%, and 0%, respectively. The tumor component was predominantly cystic in 54 and predominantly solid in 32 cases, with mortality rates of 2% and 33%, respectively. CONCLUSIONS The overall mortality of prenatally diagnosed SCTs excluding terminations was 16%. Early delivery and predominantly solid component tumors were associated with an increased risk of mortality.

A Female Infant Who Had both Complete VACTERL Association and MURCS Association : Report of a Case

A 41-day-old female infant with VACTERL association was transferred to the pediatric intensive care unit of our hospital. She had been delivered at 36 weeks gestation by spontaneous vaginal delivery and weighed 2340 g. Esophageal atresia type A with long gap, anal atresia, cardiac anomaly (atrial septal defect and patent ductus arteriosus), thoracic vertebral dysplasia, left renal agenesis, and minor anomalies (left-side facial nerve palsy, left-side difficulty in hearing, and the absence of the right thenar) had been diagnosed by various examinations. She was transferred to our hospital to receive treatment for heart failure due to a cardiac anomaly. We recognized vaginal atresia during a radical operation for anal atresia (rectovestibular fistula) at 8 months of age. Furthermore, magnetic resonance imaging (MRI) revealed agenesis of the uterus. MURCS association includes Mullerian duct aplasia or hypoplasia, renal aplasia, and cervicothoracic somite dysplasia. This is the first case of complete VACTERL association combined with MURCS association.