Early and late outcomes of congenital biliary dilatation in pediatric patients.

BACKGROUND: This study aimed to reveal the early and late postoperative complications and outcomes after surgery for congenital biliary dilatation (CBD) by reviewing cases over the past 40 years. METHODS: We retrospectively evaluated 59 patients with CBD who underwent radical surgery for complications and outcomes, based on medical records. Early complications were defined as those requiring treatment within 5 years of the initial operation. Late complications were defined as those treated more than 5 years later. RESULTS: The median age at the first surgery was 37 months. Regarding biliary reconstruction, 54 of the 59 patients (91.5%) underwent hepaticojejunostomy. Although three patients underwent cholecystoduodenostomy and one patient underwent hepaticoduodenostomy, all were converted to hepaticojejunostomy after a median of 12.5 years. One patient developed synchronous biliary carcinoma and underwent pancreaticoduodenectomy. Early complications occurred in seven patients with 10 events (surgical site infection, n = 3 bile leakage, n = 3; ileus, n = 3; bile duct obstruction, n = 1 and intussusception, n = 1). Late complications occurred in nine patients with 12 events (ileus, n = 3; anastomotic stricture, n = 3; hepatolithiasis, n = 3; asynchronous biliary carcinoma, n = 2; pancreatolithiasis, n = 1). Two of the three patients with hepatolithiasis underwent hepatectomy refractory to the endoscopic approach. Two patients developed asynchronous biliary carcinoma at 34 and 13 years after last operation; both ultimately died of the carcinoma. Only 35 patients (61.4%) underwent a follow-up examination. A total of 11 female patients (45.8%) eventually married, and all successfully gave birth. CONCLUSION: Although the long-term prognosis is excellent with complete cyst excision and hepaticojejunostomy, we emphasize the importance of long-term follow-up.

Management of refractory chylothorax in the neonatal intensive care unit: A 22-year experience.

BACKGROUND: The aim was to assess the therapeutic strategy of patients with chylothorax in a neonatal intensive care unit. METHODS: Twenty-eight infants with chylothorax were included in this study. Their clinical characteristics and outcomes were reviewed retrospectively. RESULTS: The male-to-female ratio was 1:1. The mean gestational age and birthweight were 35.1 ± 3.5 weeks and 2,692 ± 791 g, respectively. Eighteen patients were diagnosed with congenital chylothorax; chylothorax occurred postoperatively in 10 patients. Chromosomal anomalies were diagnosed in 8 patients. Six patients received surgical therapy, such as pleurodesis, thoracic duct ligation, or lymphaticovenous anastomosis. Two patients required surgery due to resistance to pleurodesis. In surgically managed patients, the daily maximum amount of pleural effusion (mL)/bodyweight (kg) ratio was significantly larger than in non-surgically managed patients: 229.0 ± 180.5 versus 59.7 ± 49.2 mL/kg. In the receiver operating characteristic analysis of the daily maximum amount of pleural effusion/bodyweight ratio, the area under the curve was 0.889 when the cut-off value was 101 mL/kg, and the sensitivity was 0.8333 and the specificity was 0.8095 (P = 0.0059). CONCLUSIONS: Pleurodesis using OK432 could become a surgical first-line therapy for chylothorax even for neonates. It was important to initiate pleurodesis for refractory chylothorax at an earlier stage. A daily chylous effusion/bodyweight ratio of >101 mL/kg was a good predictor and seemed to be a useful parameter for prompt surgical intervention.

A rare case of pyosalpinx in adolescent girl with Hirschsprung's disease who underwent transvaginal ultrasound-guided drainage.

BACKGROUND: Hydrosalpinx and pyosalpinx are rare gynecologic problems during adolescence, especially in girls without a history of sexual activity. They are even rarer in women with Hirschsprung's disease (HD). We herein report a case of pyosalpinx in an adolescent girl with HD treated by transvaginal ultrasound-guided drainage. CASE PRESENTATION: The present patient was a 12-year-old girl (weight 83 kg; height 159 cm; body mass index 32.8 kg/m2). She had undergone five laparotomies for long-segment HD by 2 years. Her menarche had occurred at 10 years. She was admitted with lower abdominal and anal pain. Computed tomography (CT), magnetic resonance imaging (MRI), and transvaginal ultrasound showed left pyosalpinx and abdominal abscess. Surgical drainage was necessary; however, she had a history of polysurgery and was severely obese, so laparotomy was considered to carry a high risk. Transvaginal ultrasound was deemed more likely to reach the abscess safely. Therefore, she was treated with transvaginal ultrasound-guided drainage by a gynecologist skilled in the procedure. She was discharged home after 52 days. One year and nine months after discharge, there was no reformation of either the abscess or pyosalpinx. CONCLUSIONS: Adolescent girls with HD are at risk of developing hydrosalpinx. Depending on the defecation status, pyosalpinx may also develop. As a less-invasive surgical treatment, transvaginal ultrasound-guided drainage can avoid laparotomy. Collaboration with a gynecologist is essential for the diagnosis and treatment of this clinical condition. Pediatric surgeons should communicate with gynecologists for such cases beginning around puberty for continuous follow-up.

The usefulness of OK-432 for the treatment of postoperative chylothorax in a low-birth-weight infant with trisomy 18.

Chylothorax is a rare but life-threatening condition in neonates. We herein report the successful use of OK-432 for a low-birth-weight infant with trisomy 18 who developed refractory chylothorax after thoracic surgery. Increasing the concentration of OK-432 seems useful in cases with a lot of pleural effusion.

A rare case of eosinophilic gastrointestinal disorders with short bowel syndrome after strangulated bowel obstruction.

BACKGROUND: Short bowel syndrome (SBS) is a rare yet costly disease with an incidence rate of 3 per million people. Herein, we report a rare case of eosinophilic gastrointestinal disorders (EGIDs) with SBS after strangulated bowel obstruction. CASE PRESENTATION: A 5-year-old male had a necrotic intestine of 340 cm resected due to strangulated bowel obstruction caused by an intestinal mesenteric hiatal hernia. The length of the residual intestine was 51 cm. Bloody stools appeared 19 days postoperatively. Colonoscopy showed diffuse redness of the colonic mucosa, and pathological findings showed moderate chronic inflammatory cellular infiltration. On blood examination, the eosinophil count was > 30%. EGIDs with short bowel syndrome (SBS) were suspected. Because his symptoms did not improve with initial nutrition therapy, he was transferred to our hospital 5 months after the operation. Prednisolone was administrated at an initial dose of 1.4 mg/kg/day, 6 days after his transfer. Bloody stools disappeared after prednisolone administration. Seven months after discharge, he had no bloody stool recurrence. CONCLUSION: The risk of developing secondary EGIDs in children with SBS should be considered, and postoperative management should include attention to abdominal symptoms and elevated eosinophil counts on blood examination.

Granulysin induces cell death with nuclear accumulation.

Exogenously added granulysin was reported to kill mammalian target cells. The sites of actions and molecular mechanisms of granulysin in target cell killing, however, are presently unclear. We here examine the effects of granulysin with the target HeLa cells transiently expressed with GFP-fused 9 kDa granulysin. Endogenously expressed GFP-fused granulysin was preferentially localized in the nucleus and induced apoptotic cell death accompanying with phosphatidylserine translocation and nuclear condensation in a caspase-independent manner. These results suggest that granulysin enters the nucleus of target cells and induces apoptosis.

Association of a new-type prostaglandin D2 receptor CRTH2 with circulating T helper 2 cells in patients with atopic dermatitis.

Prostaglandin D2 is known to be the major prostanoid produced by allergen-activated mast cells, but its role in the formation of allergic diseases is not well established because of complexity of its receptor system and lack of appropriate inhibitors. We have recently identified a new-type prostaglandin D2 receptor, named CRTH2. Studies with normal subjects have shown that CRTH2 appears to be selectively expressed by T helper 2 cells but not T helper 1 cells among circulating CD4+ lymphocytes. The exact correlation between CRTH2 and T helper 2 cells in various disease settings and the impact of CRTH2-mediated prostaglandin D2 activities on various T helper 2 responses in vivo still remain to be elucidated, however. In this study, we investigated the correlation between CRTH2 and T helper 2 cells among circulating CD4+ lymphocytes in normal adults and patients with atopic dermatitis, a T-helper-2-involving disease. The results showed that virtually all CRTH2+CD4+ lymphocytes had a pure T helper 2 phenotype and formed not all but a large proportion of circulating T helper 2 cells for both normal and atopic dermatitis subjects. In chemotaxis assays, peripheral blood CRTH2+CD4+ lymphocytes were significantly attracted by prostaglandin D2 as well as by a typical T-helper-2-attracting chemokine, thymus and activation regulated chemokine, whereas they showed little chemotactic migration toward typical T-helper-1-attracting chemokines, macrophage inflammatory protein 1beta and interferon-gamma-inducible protein 10. Furthermore, in atopic dermatitis patients, a preferential increase of CRTH2+ cells was noted within the disease-related cutaneous lymphocyte-associated antigen-positive, but not the cutaneous lymphocyte-associated antigen-negative, CD4+ lymphocyte compartment. Our results suggest the involvement of the prostaglandin D2/CRTH2 system in both normal and pathogenic T helper 2 responses.

Rapid detection of semenogelin by one-step immunochromatographic assay for semen identification.

To identify semen in forensic samples, we developed an analytical system for one-step immunoassay that has been constructed using the concept of immunochromatography and can identify semenogelin (Sg), which originates in the seminal vesicles. The system employed monoclonal antibody (mAb) and polyclonal antibody (pAb) against recombinant Sg-II (63 kDa), which has been synthesized in insect cells using baculovirus. The two antibodies bound with the seminal plasma motility inhibitor (SPMI; 14 kDa) as a final fragment peptide of Sg. The test stick is based on the sandwich technique using the above antibodies. When serial dilutions of seminal plasma were analyzed using this test stick, the intensity of a clear immunoreactive signal peaked at 2000-fold dilution. Thereafter, the signals decreased slowly but still persisted up to 400,000-fold dilution. The Sg antigen was undetectable in saliva, urine, breast milk, serum or vaginal secretions. Also, the test stick shown did not react with animal semen samples, such as those from horses, dogs, swine and bulls. When semen samples, diluted 100,000-fold from 100 men were tested, the Sg antigenic activity was detectable in all samples. In addition, the specificity and sensitivity of the test stick for identification of semen were demonstrated by comparative forensic studies. We conclude that this immunoassay method is a useful confirmatory test for the identification of semen. The immunochromatographic system for forensic testing or research use will become available commercially soon.

Quantification of seminal plasma motility inhibitor/semenogelin in human seminal plasma.

Semenogelin I and II (Sg I and II) are the major components of human semen coagulum. The protein is rapidly cleaved after ejaculation by the chymotrypsin-like protease prostate-specific antigen (PSA), which results in the liquefaction of the semen coagulum and the progressive release of motile spermatozoa. One of the cleavage products of the protein, a 14-kDa protein, is a sperm motility inhibitor (seminal plasma motility inhibitor [SPMI]). We developed a monoclonal antibody (mAb) that is specific to the fragment of Sgs, SPMI, and a sandwich enzyme-linked immunosorbent assay (ELISA) system for the quantification of Sgs using this mAb. Then, we measured SPMI/Sg levels in human seminal plasma from healthy male volunteers (n = 100, aged 18-24 years). The mean level of SPMI/Sg in seminal plasma was 19 +/- 13 mg/mL (range, 4-68 mg/mL). Log-transformed SPMI/Sg levels were negatively correlated with the sperm motility (r = -0.229, P =.0220) and positively correlated with the total protein concentration (r = 0.793, P <.0001). This result supports that SPMI, one of the fragments of Sg, has its inhibitory effect on ejaculated spermatozoa in liquefied semen under physiological conditions.

[External quality assessment of the genetic testings for hematopoietic tumor, CML].

Genetic testings are commonly employed in various fields of clinical medicine and the test items performed at clinical laboratories are increasing rapidly in number. They are utilized to make early and/or definite diagnoses of infectious diseases, leukemia, cancers and molecular inherited diseases and also to monitor the progress of the diseases. However, these genetic testings except for infectious diseases have been developed independently at each clinical laboratory and the test results obtained at each laboratory are not always compatible each other. Under these situations it is widely expected to construct advanced genetic testing systems that can supply standardized data at any of domestic and international clinical laboratories. For the period from April, 1999 to March, 2002 three major clinical laboratories, SRL, Inc., BML, Inc. and MBC, Inc., were consigned by JBA (Japan Bioindustry Association) to collaborate in standardizing the evaluation methods for genetic testing systems among the clinical laboratories. The aim of the study is to develop the standardized genetic testing systems and to propose them as international standard operational procedures to the ISO/TC212 working group. Although one of the most important issues for standardization is the external quality assessment, they have not been carried out in reality. In this study we evaluated the difference of the genetic testing results obtained during the year of 2000 and 2001 among the clinical laboratories. The genetic testings for hematopoietic tumor, CML were selected to be evaluated since they are widely accepted as clinically useful tests.

Mechanism of plasma glutathione peroxidase production in bovine adipocytes.

Plasma glutathione peroxidase (pGPx) is an anti-oxidative enzyme. Using the polymerase chain reaction subtraction method, we have previously identified pGPx as a large part of the genes that are expressed following adipocyte differentiation in a bovine intramuscular preadipocyte (BIP) line. Therefore, we have analyzed the mechanism of production of pGPx in adipocytes. The expression of pGPx and C/EBPdelta increases during adipogenesis, with dexamethasone being the main effector of these genes. The expression of pGPx gene has been clearly detected in BIP cells and human adipocytes, but hardly in 3T3-L1 cells. The production of pGPx in bovine tissues is greatest in kidney and in intraperitoneal fat. We consider that the transcriptional control of pGPx in cattle might be carried out by C/EBPdelta and that the expression of pGPx might be a characteristic phenomenon of bovine adipogenesis.

Effects of prostaglandin D2 on helper T cell functions.

Prostaglandin (PG) D(2) is abundantly produced by mast cells in the sites of allergic inflammations and acts on various cell types through its receptors DP and CRTH2. Among human T cells, CRTH2 is preferentially expressed on Th2-type cells. However, distribution of DP among T cells and impacts of CRTH2- and DP-mediated signals on T cell functions are presently unclear. Here, we show that CD4(+) and CD8(+) T cells producing IFN-gamma and IL-2 were reduced by DP-mediated signals, while CRTH2-mediated signals enhanced IL-2, IL-4, IL-5, and IL-13 production by Th2 cells. CRTH2 signals also caused up-regulation of CD11b and CD40L in resting Th2 cells. RT-PCR analysis revealed distribution of DP among Th cell subsets. On CRTH2(+) Th2 cells, the CRTH2-mediated PGD(2) effects were dominantly observed. Thus, PGD(2) favors Th2 functions through CRTH2 while restraining Th1 functions via DP, which may contribute to development of Th2-dominated status in allergic inflammations.

Cutting edge: agonistic effect of indomethacin on a prostaglandin D2 receptor, CRTH2.

Indomethacin is a widely used nonsteroidal anti-inflammatory drug and is generally known to exhibit its multiple biological functions by inhibiting cyclooxygenases or activating peroxisome proliferator-activated receptors. In this study, we present evidence demonstrating that the novel PGD(2) receptor chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) is another functional target for indomethacin. Indomethacin induced Ca(2+) mobilization in CRTH2-transfected K562 cells at submicromolar concentrations (approximate EC(50), 50 nM) in a G(alphai)-dependent manner as PGD(2) did. Other nonsteroidal anti-inflammatory drugs (aspirin, sulindac, diclofenac, and acemetacin) had no such effect even at micromolar concentrations. In chemotaxis assay, three CRTH2-expressing cell types, Th2 cells, eosinophils, and basophils, were all significantly attracted by indomethacin (EC(50), 50-500 nM) as well as by PGD(2) (EC(50), 2-20 nM), and the effects of indomethacin were blocked by anti-CRTH2 mAb. These results suggest the involvement of CRTH2 in mediating some of therapeutic and/or unwanted side effects of indomethacin, independently of cyclooxygenases and peroxisome proliferator-activated receptors.

Differential expression of granulysin and perforin by NK cells in cancer patients and correlation of impaired granulysin expression with progression of cancer.

Granulysin has been identified as an effector molecule co-localized with perforin in the cytotoxic granules of cytotoxic T lymphocytes and natural killer (NK) cells, and has been reported to kill intracellular pathogens in infected cells in the presence of perforin and to induce a cytotoxic effect against tumor cells. The aim of the present study was to elucidate whether intracellular expression of granulysin and perforin by NK cells might be associated with progression of cancer. Flow cytometric analysis demonstrated high levels of perforin and granulysin expression by CD3(-) CD16(+) cells in healthy controls. In contrast, cancer patients exhibited significantly decreased levels of granulysin expression ( P<0.005), despite having equally high levels of perforin expression in comparison with healthy controls. The tumor-free patients expressed granulysin at levels similar to healthy controls, while the progressive tumor-bearing patients expressed remarkably lower levels of granulysin compared to healthy controls ( P<0.0001). Similarly, patients with an advanced performance status had significantly fewer granulysin-positive NK cells than healthy controls. Meanwhile, a considerable number of the tumor-bearing patients showed a decrease in the number of circulating NK cells, and a correlation between impaired granulysin expression and reduced circulating NK cells was observed. These findings suggest that the tumor-bearing patients with impaired granulysin expression were in an immunosuppressive state. In conclusion, impaired expression of granulysin by NK cells correlates with progression of cancer, and determination of granulysin expression might prove informative for assessing the immunological condition of cancer patients.

Cloning and expression of type XII collagen isoforms during bovine adipogenesis.

In order to isolate candidate genes involved in bovine adipocyte differentiation, we have constructed a subtraction library from a clonal bovine intra-muscular pre-adipocyte (BIP) cell line using the suppression subtractive hybridization method. We have isolated a set of subtracted cDNA fragments whose respective mRNA levels are up-regulated during the adipogenic differentiation of BIP cells, and cloned cDNAs from a differentiated BIP-lambda ZAP II cDNA library. Two cDNA clones were highly homologous to the sequence of mouse and human type XII collagen alpha-1, determined by a BLAST homology search. As type XII collagen has been reported to have four types of splicing isoform, two clones were determined to be XII-1 and XII-2 splicing isoforms, respectively, because of a difference in the C-terminal NC1 domain. From the expression analysis of type XII collagen, the XIIA-2 isoform was mainly expressed in differentiated BIP cells and adipose tissues. Although the function of type XII collagen has not been established as yet, these results suggest that type XII collagen may be associated with adipocyte differentiation and adipose formation in cattle and is a potentially useful marker for adipogenesis.

Granulysin in human serum as a marker of cell-mediated immunity.

Granulysin is a cytolytic granule protein of natural killer (NK) cells and cytotoxic T lymphocytes (CTL) with a broad range of antimicrobial and tumoricidal activities. Two molecular forms of granulysin, the 15-kDa precursor and 9-kDa mature form, are produced in these cells. In this study, we developed monoclonal antibodies against granulysin and found that the 15-kDa granulysin is spontaneously secreted by peripheral blood NK and T cells via a non-granule exocytotic pathway. When NK cells killed the target cells, the released granulysin levels in culture supernatants significantly increased through the granule exocytosis. The granulysin protein was found in the sera of healthy individuals at an average concentration of 3.7 +/- 3.2 ng/ml (age 0-99 years, n=244). The serum levels of granulysin were transiently highly elevated among patients with acute viral infections. In addition, the serum granulysin levels in patients with severe immunodeficiency treated bycell therapy fluctuated proportionately to the improvement of other immunological parameters. Our results suggest that granulysin is well associated with diverse activities of NK cells and CTL in physiological and pathological settings and could be a useful novel serum marker to evaluate the overall status of host cellular immunity.