Objective To investigate the effects of Epstein-Barr virus (EBV) infection on apoptosis of cord blood monocytes-derived dendritic cells (DC) in neonates. Methods Cord blood monocytes were induced into DC by 50 ng/mL recombinant human granulocyte macrophage colony-stimulating factor(rhGM-CSF)and 10 ng/mL recombinant human interleukin-4 (rhIL-4),and DC were divided into three groups:①4G group:rhGM-CSF and rhIL-4 were added separately on the day of cell separation;②4G+0d EBV group:rhGM-CSF,rhIL-4 and B95.8 cell supernatants were added simultaneously on the day of cell separation;③4G+5d EBV group:hGM-CSF and rhIL-4 were added on the day of cell separation,and B95.8 cell supernatants were added on the 5th day.Percents of DC apoptosis were measured using Annexin V-FITC and PI staining by flow cytometry.The expression of X-linked inhibitor of apoptosis protein (XIAP) was detected by Western Blot.Results Percent of DC apoptosis in 4G+0d EBV group was significantly higher than that in 4G group on the 6th to 14th day(P<0.05),and percent of DC apoptosis in 4G+5d EBV group was significantly higher than that in 4G group on the 7th to 14th day(P<0.05).The effects of EBV infection on percent of DC apoptosis had relations with infection time points.The expression of XIAP in DC decreased significantly after EBV infection. Conclusion EBV promotes apoptosis of cord blood monocytes-derived DC,which is associated with the differentiation and maturation status of DC.

Objective To explore the immune characteristic of a patient with hypogammaglobulinaemia and low natural killer(NK)cell numbers. Methods Case histories were analyzed repeatedly to grasp important characteristics of the disease. A routine blood examination and peripheral blood immunological detection was done. Complete neutrophil function test,lymphocyte proliferation ability and cytokine profile were evaluated. Results A 3-year-old boy was referred to the hospital because of fever and abdominal pain as well as partial intestinal obstruction. Laboratory examination revealed low immunoglobulin,reduced absolute numbers of B and NK cells and deficient B cell function of lower IFN-γ and IL-12 production. The patient also had mesenteric lymphadenopathy and a caecal mass with histology suggestive of ileocecal junctional tuberculosis. The patient was infected with cytomegalovirus(CMV)and fungus,which are not common in simple antibody deficiencies. The patient was followed for four years and during this period the patient had recurrent periodontitis resulting in the loss of one tooth. Conclusions Although the precise mechanism of the defect is unclear,we speculate that the case may cast new light on primary immunodeficiencies predominantly with antibody deficiency.(J Clin Pediatr,2011,29(7):601-608)

Objective To study the clinical manifestation,diagnosis,differential diagnosis and the essentials of management and treatment of fibrodysplasia ossificans progressiva (FOP).Methods Three cases of FOP were reported.The features of clinical manifestation and radiography were studied.The literature related to FOP was reviewed.Results FOP affected young children′s age of onset was between 10 days and 2 years (mean age 1 3 years).Mean disease duration was 5 3 years (range 2～11 years),and mean age 5 3 years (range 2～11 years) with sex ratio 1∶2 (boy∶girl).Soft tissue swelling in cervical and dorsal regions with or without local pain and warmth,and low fever were the early clinical manifestations.These nodules usually disappeared spontaneously,but some of nodules gradually developed ossification.The X ray features included ectopiac ossification most frequently in the soft tissue of the upper back and neck,next,the loin,chest and extremities.Two cases showed short hallux and hallux valgus.Exacerbation of the two cases was precipitated after muscle biopsy and careless venepuncture.All patients showed progressive extra articular bony ankylosis of most joints of axial and/or appendicular skeleton with severe movement restriction.Conclusion FOP is a rare and disabling genetic disorder of connective tissue.FOP should be diagnosed as early as possible and non invasively,based upon history,clinical and radiological findings.The finding of abnormalities of the great toe is helpful to diagnose FOP so that management can be early and adequate.Manogement principle includes avoiding conditions potentially provocative of abnormal ossification.The disease should be familiar to pediatricians.

Background Herpes stromal keratitis (HSK) is a common infectious ocular surface disease,with a higher recurrent rate,especially in necrotizing HSK.The diagnosis of HSK primarily depends on signs and symptoms,and specific laboratory diagnostic is lack.Objective This study was to clarify the expression of herpes simplex virus (HSV) in the corneal epithelium scrapings and tears of necrotizing HSK patients.Methods Thirty eyes of 30 patients with necrotizing HSK were enrolled in Nanjing Hospital of Nanjing Medical Hospital from September 2012 to September 2013 under the patient's informed consent.The eyes were examined by slit lamp microscope and scored.HSK patients received local and systemic therapy for 8 weeks and then an oral maintenance dose for 6 months.Corneal epithelial scrapings and tears samples were collected for HSV DNA detection by real-time PCR before and the 1 st,2nd,4th,6th and 8th week after therapy respectively.The difference of HSV positive rate was compared between corneal epithelium scrapings and tears samples using Chi-square test.Multilevel mixed effective model was employed to evaluate HSV concentration change in the samples at various time points in the HSV-positive patients of initial visit.The correlation between HSV concentration and clinical score was analyzed by Spearman rank correlation.Results HSV-positive rate was 46.4％ (13/30) in the corneal epithelial scrapings and 13.3％(4/30) in the tear samples,showing a significant difference between them (P =0.006).HSV-positive rate was significantly lower in the corneal epithelial scrapings 1 week,2 weeks and 4 weeks after treatment than before (P =0.001,0.003,0.004),and no HSV was detected 6 weeks and 8 weeks after treatment.No significant change in HSV-positive rate in tear samples in 1 week and 2 weeks after treatment in comparison with before treatment (P =1.000,0.583),and no HSV was detected after 4 weeks following treatment.The HSV concentration was 2460 (2 165-636500)/ml in initial 13 HSV-positive eyes of corneal epithelial scrapings and 0 (0-1150)/ml in initial 13 HSV-positive eyes of tear samples.Multilevel mixed effective model determined that HSV concentration was significantly lower in corneal epithelial scraping than that in tear (P =0.005),and HSV concentration was reduced with the lapse of time (P =0.001),with the faster rate of decline in the corneal epithelial scrapings (P =0.049).A positive correlation was found between initial HSV concentration and clinical scores (rs =0.844,P =0.000).Conclusiors Real-time PCR appears to be a powerful molecular tool for the detection of HSV in the HSK,especially in corneal epithelial scrapings of lesion.The initial positive outcome of viral DNA in corneal epithelial scrapings predicts a severe clinical procedure.

Objective To develop a flow cytometrie assay to for diagnosis of X-Linked Hyper.1gM Syndrome(XHIM).Methods Heparinized peripheral blood obtained from patient,mother and a healthy control was diluted with RPMI- 1640(unstimulated control)or with RPMI-1640 containing 15μl PMA(1 rig/μl)and 6 trl ionomycin(50 ng/μl)(stimulated cell).Using directly labeled antibodies,we have examined CD40 ligand levels on CD3+ CD8- lymphocyte surface,and CD69 levels on CD;lymphocyte Surface to determine whether the cells were activated.Results CD69 levels on CD3+ lymphocyte surface from stimulated group and from unstimulated group were above 96% and below 3%,respectively.CD40L levels Oil CD3 CDs-T lymphocyte surface from stimulated group were 0.8% (patient),60.04%(mother) and 62.87%(healthy contr01).CD40L levels on CD3+ CD8-T lymphocyte surface from unstimulated group were 0.88% (patient),4.15%(mother)and 5.51%(healthy contr01).Conclusion This flow cytometric assayis accurate and convenient,which Can be used in neonatal screening.

Objective To evaluate the bacteriostasis of sulfamethoxazole(SMZ)combined with trimethoprim(TMP)against My‐cobacterium tuberculosis(MTB) in vitro ,so as to provide basis for clinical application .Methods The minimal inhibitory concentra‐tion(MIC) value of TMP/SMZ against MTB ,including standard sensitive strain(H37Rv) ,clinical sensitive strains(20 strains) and clinical multiple‐drug‐resistance strains(MDR ,30 strains) ,was detected by using MPP observation method .Results The MIC val‐ue of standard strain H37Rv was TNP(0 .5 μg/mL)+SMZ(9 .5 μg/mL) .The growth of 40 strains(accounted for 80% ) of clinical isolates ,including 17 sensitive strains and 23 MDR strains ,could be inhibited by TMP(1 μg/mL)+ SMZ(19 μg/mL) compound . Conclusion TMP combined with SMZ may has good antibacterial activity for strains of MTB in vitro .

Objective To study the mutation of STK11 gene in a Chinese family and a sporadic patient with Peutz-Jeghers syndrome (PJS),and to provide a basis for genetic diagnosis and counseling.Methods One sporadic patient and two patients from a family with PJS were collected,all of whom had typical mucosal pigmentation and gastrointestinal polyposis.Blood samples were obtained from the two patients and six unaffected relatives in this family,the sporadic patient,and 100 healthy controls.DNA was extracted,and PCR was performed to amplify nine exons and their adjacent introns in the STK11 gene followed by direct sequencing.The sequencing results were aligned to the published sequence of STK11 gene from Genbank.Results No mutation was found in the STK11 gene of any of the patients,unaffected relatives,or healthy controls.Conclusions Genetic heterogeneity exists in Peutz-Jeghers syndrome,hinting that there may be other causative genes or sites for this entity.

Objective To assess the outcome of vesicoureteral reflux after augmentation cystoplasty in patients with neurogenic bladder.Methods Between January 2008 and January 2014,a total of 25 patients,with a hypocompliant bladder associated with vesicoureteral reflux confirmed by video-urodynamics preoperatively,were recruited in this study.They all had undergone bladder augmentation with a generous detubularized segment of bowel at our institution.No effort had been made to correct existing reflux.Preoperatively assessment included urinalysis,kidney function tests,ultrasonography,video-urodynamic evaluation.All patients had various degrees of vesicoureteral reflux.The status of vesicoureteral reflux and bladder function were studied by video-urodynamic.Results Mean follow-up was 2.2 years (range 0.5 to 5.5 years).The video-urodynamics manifested a significant improvement of bladder capacity,diminution of intravesical pressure and resolution of reflux after bladder augmentation.Of the 25 patients,20 (80％) no longer had reflux,3 (12％) had improvement,2 (8％) had no change.Sixteen of 18 with grades Ⅰ to Ⅲ (89％),all refluxing units with grade Ⅳ to grade Ⅴ (100％) showed complete cessation of reflux.Symptomatic urinary infection was not found after surgery.Conclusions Augmentation enterocystoplasty without ureteral reimplantation is effective and adequate for patients with high pressure and hypocompliant neurogenic bladder.Therefore,ureteral reimplantation is not necessary underwent when augmentation enterocystoplasty is recommended to patients with neurogenic bladder and vesicoureteral reflux.

Objective To evaluate the effect of total glucosides of paeony (TGP) on the expression of interleukin-18 (IL-18) in human HaCaT keratinocytes,and to explore the roles of extracelluar signal-regulated protein kinase1/2 (ERK1/2) and c-Jun N-terminal kinase 1/2 (JNK1/2) signaling pathways in the effect.Methods Some cultured human HaCaT keratinocytes were classified into three groups:control group treated with dimethyl sulfoxide (0.031％),TGP groups treated with 6 different concentrations (0.5,2.5,12.5,62.5,125.0 and 312.5 mg/L) of TGP respectively,inhibitor groups treated with TGP of 125 mg/L after 2-hour pretreatment with PD98059 (an ERK1/2 inhibitor) and SP600125 (a JNK1/2 inhibitor) of 10 μmol/L respectively.After additional culture for 48 hours,reverse transcription (RT)-PCR was performed to measure the mRNA expression level of IL-18,and enzymelinked immunosorbent assay (ELISA) to determine the level of IL-18 protein in the culture supematant of HaCaT cells.Some HaCaT keratinocytes were classified into two groups to be treated with TGP of 125 mg/L for 15,30 and 60 minutes with or without the pretreatment with PD98059 and SP600125 of 10 μmol/L; then,Western blot was carried out to determine the phosphorylation levels of ERK1/2 and JNK1/2 in HaCaT cells.Results The levels of IL-18 mRNA and protein in culture supernatant were significantly increased by TGP of 0.5 and 2.5 mg/L,but decreased by TGP of 62.5 and 125.0 mg/L,and TGP of 125.0 mg/L showed the strongest inhibitory effect.After treatment with TGP of 125.0 mg/L,the level of phosphorylated ERK1/2 in HaCaT cells peaked at 15 minutes (0.448 ± 0.018),decreased to 0.213 ± 0.005 at 30 minutes and 0.217 ± 0.005 at 60 minutes,with significant differences between TGP-treated and untreated cells at 15 minutes (0.448 ± 0.018 vs.0.204 ± 0.005,P＜ 0.05) but not at 30 or 60 minutes (both P ＞ 0.05).The phosphorylation level of ERK1/2 was 0.237 ± 0.010 in HaCaT cells pretreated with PD98059 prior to the treatment with TGP,significantly different from that in HaCaT cells treated with TGP only (P ＜0.01).TGP of 125.0 mg/L had no obvious effect on JNK phosphorylation,and there was no significant difference in the level of phosphorylated JNK1/2 between HaCaT cells untreated and those treated with TGP of 125.0 mg/L for different durations (all P ＞ 0.05).Conclusions TGP can inhibit the expression of IL-18 mRNA and protein in HaCaT cells,likely through the ERK1/2 signaling pathway.

Objective To measure serum levels of 25-hydroxyvitamin D (25HVD) in patients with chronic urticaria (CU),and to explore its role in the occurrence of CU.Methods Peripheral blood samples were obtained from 50 patients with CU and 40 healthy controls.The urticaria activity score (UAS) was used to assess the severity of CU and to group the patients with CU.Enzyme-linked immunosorbent assay (ELISA) was performed to measure the serum levels of 25HVD,interferon-γ (IFN-γ),interleukin-4 (IL-4) and immunoglobulin E (IgE).Statistical analysis was carried out by t test,rank sum test and linear correlation analysis.Results The serum level of 25HVD was significantly lower in the patients with CU than in the healthy controls (15.20 ± 7.72 vs.21.54 ± 8.31 μg/L,t =3.75,P ＜ 0.05).Moreover,there was a significant difference in the distribution of serum 25HVD levels between the patients and healthy controls (H =17.9,P ＜ 0.05).However,no significant difference was observed in the serum level of 25HVD between severe and mild CU groups (15.57 ± 7.38 vs.14.86 ± 6.28 μg/L,t =0.37,P ＞ 0.05).Compared with the control group,the patient group showed significantly decreased serum levels of IFN-γ (t =15.34,P ＜ 0.05),but increased serum levels of IL-4 and IgE (t =6.54,4.88,respectively,both P ＜ 0.05).Among the patients with CU,the serum level of 25HVD was positively correlated with that of IFN-γ(r =0.738,P ＜ 0.05),but negatively correlated with that of IL-4 (r =-0.689,P ＜ 0.05),and uncorrelated with that of IgE (r =-0.271,P ＞ 0.05).Conclusion The serum level of 25HVD evidently decreased in patients with CU,and it may participate in the occurrence of CU by mediating the Th 1/Th2 imbalance.