NLRP3 activation in macrophages promotes acute intestinal injury in neonatal necrotizing enterocolitis.

BACKGROUND: Macrophages are involved in various immune inflammatory disease conditions. This study aimed to investigate the role and mechanism of macrophages in regulating acute intestinal injury in neonatal necrotizing enterocolitis (NEC). METHODS: CD68, nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing 3 (NLRP3), cysteine aspartate-specific protease-1 (caspase-1), and interleukin-1ß (IL-1ß) in paraffin sections of intestinal tissues from NEC and control patients were detected with immunohistochemistry, immunofluorescence, and western blot. Hypertonic pet milk, hypoxia and cold stimulation were used to establish a mouse (wild type and Nlrp3-/-) model of NEC. The mouse macrophage (RAW 264.7) and rat intestinal epithelial cell-6 lines were also cultured followed by various treatments. Macrophages, intestinal epithelial cell injuries, and IL-1ß release were determined. RESULTS: Compared to the gut "healthy" patients, the intestinal lamina propria of NEC patients had high macrophage infiltration and high NLRP3, caspase-1, and IL-1ß levels. Furthermore, in vivo, the survival rate of Nlrp3-/- NEC mice was dramatically improved, the proportion of intestinal macrophages was reduced, and intestinal injury was decreased compared to those of wild-type NEC mice. NLRP3, caspase-1, and IL-1ß derived from macrophages or supernatant from cocultures of macrophages and intestinal epithelial cells also caused intestinal epithelial cell injuries. CONCLUSIONS: Macrophage activation may be essential for NEC development. NLRP3/caspase-1/IL-1ß cellular signals derived from macrophages may be the underlying mechanism of NEC development, and all these may be therapeutic targets for developing treatments for NEC.

RNAa-mediated overexpression of WT1 induces apoptosis in HepG2 cells.

AIM: Recent studies have reported that double-stranded RNA (dsRNA) can activate gene expression by targeting promoter sequence in a process termed RNA activation. The present study was conducted to evaluate the potential of WT1 induction by small activating RNA targeting the WT1 promoter (dsWT1) in the treatment of hepatocellular carcinoma. METHODS: The human hepatocellular carcinoma cell line HepG2 was transfected with dsRNA by liposomes. The expression of mRNA and protein in cells were investigated using real-time reverse real-time quantitative PCR and Western blot, respectively. Cell viability and clonogenicity were determined by MTT assay and clonogenicity assay, respectively. Cell apoptosis was evaluated by flow-cytometric analysis. RESULTS: Expressions of WT1 mRNA and protein in dsWT1 treated HepG2 cells were significantly elevated. Inhibition of cell viability by dsWT1 was dose-dependent and time-dependent. Reduction of the number and size of colonies formed were found in dsWT1 treated cells. dsWT1 induced significant apoptosis in HepG2 cells. The decreased anti-apoptotic protein Bcl-2 and elevated pro-apoptotic protein Bak expression were detected in dsWT1 treated cells. The level of pro-caspase-3 remarkably decreased and cleaved caspase-3 and PARP fragment were also detected in dsWT1 treated cells. CONCLUSION: These data show that RNAa-mediated overexpression of WT1 may have therapeutic potential in the treatment of hepatocellular carcinoma.

Association of T-helper cell cytokine level with age in patients with biliary atresia: a preliminary study.

BACKGROUND: The pathogenesis of biliary atresia (BA) is associated with an inflammatory process involving the biliary tree. This study aimed to investigate the association of T-helper cell cytokine levels with age in patients with BA. METHODS: Twenty-eight patients with BA were divided into three groups according to their age (< 2 months, 2-3 months, and ≥ 3 months). All the patients underwent Kasai portoenterostomy. Blood samples were collected from the patients preoperatively, and the liver tissue specimens were obtained during surgery. We detected serum levels of interleukin (IL)-1ß, IL-12p70, interferon (IFN)-γ, IL-6, IL-10, and transforming growth factor (TGF)-ß1 and liver expression of IL-1ß, IL-6, and TGF-ß1. RESULTS: The serum levels of IL-1ß, IL-12p70, IL-6, and IL-10 in patients aged ≥ 3 months were significantly higher than those in patients aged < 2 months. There were no significant age-related differences in the IL-1ß, IL-6 and TGF-ß1 expression levels in the liver tissue of patients with BA. CONCLUSIONS: The serum levels of IL-1ß, IL-6, IL-10 and IL-12p70 showed significant age-related differences in patients with BA. Interpretation of the role of cytokines in BA needs to take patient's age into consideration.

Outcome of children with melamine-induced urolithiasis: results of a two-year follow-up.

OBJECTIVE: Our purpose is to describe the characteristics and the outcome of melamine-induced renal and urinary tract injury in young children who consumed melamine-contaminated infant formula. METHODS: This is a descriptive longitudinal study over 2 years in 240 children with melamine-induced urolithiasis screened in our hospital from September 15 to October 31, 2008. Ultrasonography and serum creatinine (SCr), urea, ß2-microglobulin (MG), cystatin C (Cys C), urinary Cr (UCr), microalbumin (mALB), α1-MG, ß2-MG, n-acetyl-ß-d-glucosaminidase (NAG) and retinol-binding protein (RBP) measurements were performed. RESULTS: The children ranged in age from 1 to 82 months, and 145 were males. The largest calculus was 33 mm in diameter. X-ray diffraction pattern of the calculi displayed two diffraction peaks at 10.9° and 27.7° (2θ). Surgical management was performed in 14 patients. In 226 patients without surgical management, the calculi were passed in 59.63% patients within 1 month, in 85.40% within 6 and in 91.15% within 24 months. Increased SCr and urea levels were noted in three and six patients, respectively, at the time of diagnosis. The SCr, serum ß2-MG, and Cys C levels at the time of diagnosis were higher than those at 3 and 6 months after diagnosis (P < 0.05, respectively). The levels of mALB/UCr, NAG/UCr, and RBP/UCr at the time of diagnosis were higher than those at 3, 6, 12, and 24 months after diagnosis (P < 0.05, respectively). α1-MG/UCr and ß2-MG/UCr levels at the time of diagnosis were similar to those at 3 months after diagnosis, and significantly higher than those in the follow-up period (P < 0.05, respectively). CONCLUSION: Melamine might injure both the renal glomerulus and the tubule, and that the predominant lesion is urolithiasis. The compositions of melamine-induced urolithiasis are melamine and cyanuric acid crystals. The urolithiasis might persist for over 2 years and cause irreversible damage. Therefore, a long-term follow-up for all patients is required.

Relationship between plasma leptin levels and clinical outcomes of pediatric traumatic brain injury.

High plasma leptin level has been associated with mortality after adult intracerebral hemorrhage. The present study was undertaken to investigate the plasma leptin concentrations in children with traumatic brain injury and to analyze the correlation of leptin with pediatric traumatic brain injury outcome. Plasma leptin concentration of eighty-nine healthy children and 142 children with acute severe traumatic brain injury was measured by enzyme-linked immunosorbent assay. Twenty-six patients (18.3%) died and 42 patients (29.6%) had an unfavorable outcome (Glasgow outcome scale score of 1-3) at 6 months after traumatic brain injury. Upon admission, plasma leptin level in patients was substantially higher than that in healthy controls. A forward stepwise logistic regression selected plasma leptin level as an independent predictor for 6-month mortality and unfavorable outcome of patients. A receiver operating characteristic curve analysis showed plasma leptin level better predicted 6-month mortality and unfavorable outcome. The prognostic value of leptin was similar to that of Glasgow Coma scale score for 6-month clinical outcomes. Thus, plasma leptin level represents a novel biomarker for predicting 6-month clinical outcome in children with traumatic brain injury.

Incidence of pediatric metachronous contralateral inguinal hernia in children aged ≥1 year.

BACKGROUND: The management of the contralateral asymptomatic side when a child with initial unilateral inguinal hernia undergoes herniorrhaphy continues to be controversial. Age less than 6 months at initial herniorrhaphy is considered as a high risk factor of the occurrence of metachronous contralateral inguinal hernia (MCIH). We performed herniorraphy for patients ≥1 year with initial unilateral hernia at one-day-set outpatient-surgery department without any intervention of contralateral groin. In this study, we reviewed the characteristics of development of MCIH in this condition and discuss the management strategies of MCIH. METHODS: The subjects of this study were children who were treated at our outpatient-surgery department from January 2006 to December 2006. A total of 2129 patients with initial unilateral hernia and aged ≥1 year underwent an ipsilateral herniorhhaphy only. Patients were followed up for the development of MCIH to 60 months. The Chi-square test was used for intergroup comparison, a level of P<0.05 was considered as statistically significant. RESULTS: Among these children 1341 (63.0%) were obtained 60 months follow-up data, 1146 (85.5%) were boys and 195 (14.5%) were girls. MCIH developed in 70 (5.2%) patients, 61 were boys and 9 were girls. In 570 patients aged 12-23 months, 43 developed MCIH (7.5%); in 564 patients aged 24-59 months, 21 developed MCIH (3.7%); and in 207 patients ≥60 months, 6 patients developed MCIH (2.9%), the difference between these groups was highly significant (P=0.004). In male patients, 30 right-sided MCIHs occurred after 423 initial left-sided herniorrhaphies (7.1%) and 31 left-sided MCIHs occurred after 723 initial right-sided herniorrhaphies (4.3%), difference between these two groups was significant (P=0.041). Seventy-seven percent of the MCIHs occurred within 1 year, 94% occurred within 2 years after initial herniorraphy. CONCLUSIONS: As the overall incidence of MCIH in patients aged ≥1 year was 5.2%, routine contralateral groin exploration is not suggested. Transinguinal laparoscopy could be considered as an alternative of conventional "wait and see" policy, especially in patients less than 2 years or left-sided initial unilateral inguinal hernia. If "wait and see" policy is adopted, patients should be closely followed up for 2 years.