Functional constipation is a common and frequently occurring disease in childhood. In addition to meeting the clinical di-agnostic criteria for constipation and performing the general routine examinations, there are the need to have further examinations such as colonic transit time measurement, anorectal manometry and colonoscopy in order to rule out other causes. Therefore, the purpose of this paper is to introduce several latest research progress in examinations related to constipation in children and to provide some gui-dance and references for clinicians.

Air ; analysis ; Alkenes ; analysis ; Butadienes ; analysis ; Chromatography, Gas ; methods ; Environmental Monitoring ; methods ; Workplace

Objective To establish a cellular model for the expression of the C-type lectin dendritic cellspecific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN),and to provide a basis for the functional analysis of DC-SIGN.Methods The cDNA of DC-SIGN was obtained via PCR,and cloned into the eukaryotic expression vector porcine cytomegalovirus-enhanced green fluorescent protein (PCMV-EGFP) with EGFP at the N terminal of DC-SIGN.Then,the recombinant PCMV-EGFP-DC-SIGN plasmid was transfected into HEK293T cells followed by the detection of DC-SIGN expression using PCR,Western blot and flow cytometry.Confocal microscopy was performed to localize the expression of DC-SIGN-EGFP and visualize the recognization and internalization of the Derp2 allergen by DC-SIGN.Results The recombinant fluorescent fusion protein-expressing plasmid was successfully constructed.Both PCR and Western blot confirmed the expression of DC-SIGN.Flow cytometry showed that the expression of DC-SIGN was increased by approximately 50％ in HEK293T cells transfected with the recombinant expression plasmid compared with those untransfected.As confocal microscopy showed,the green fluorescence-labelled DC-SIGN was located on the cell membrane,which could bind to the red fluorescence-labelled antigen Derp2 and internalize it into the cells.Conclusions The recombinant DC-SIGN-EGFP fusion protein is characteristically located on the surface of 293T cells,which can be recognized by the DC-SIGN-specific antibody and is capable of internalizing the allergen Derp2,and may serve as an ideal cell model for further studies on molecular function of DC-SIGN.

Objective To synthesize herpes simplex virus (HSV) envelope glycoprotein gC using gene engineering techniques,and to verify its expression.Methods Two separate parts of the HSV envelope glycoprotein gC,i.e.,GC-F and GC-R,were respectively synthesized.The GC-F and GC-R genes were synthesized,subcloned into the expression vectors pSumo-Mut (containing recognition sequences for endonucleases Stu1 and XhoI) and pCzn1 (containing recognition sequences for endonucleases NdeI and XhoI) respectively to form the recombinant plasmids pSumo-Mut-GC-F and pCzn1-GC-R.E.coli BL21 Arctic Express (DE3) cells were transformed with the two recombinant plasmids separately.Isopropyl thiogalactoside (IPTG) was used to induce the expression of target protein which was subsequently purified by nickel affinity chromatography.Finally,Western blot was performed to verify the reactivity of the synthesized protein with the sera of HSV-1-positive patients.Results Both GC-F and GC-R genes were synthesized by a total gene synthesis method.As sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) showed,the fusion proteins were mainly distributed in the sediment layer.The purity of GC-F and GC-R proteins was over 80％ after purification by affinity chromatography.Western blot showed that both of the proteins were reactive with anti-HSV-1 antibody-positive sera.Conclusions Fusion expression vectors have been constructed for the gC protein,and IPTG successfully induces its expression.Moreover,the resulting proteins could react with anti-HSV-1 antibody-positive sera,and may serve as an ideal experimental material for next functional study.

Objective To investigate the effect of selectively upward placement of acetabular implants on limb length and post?operative function of developmental dysplasia of the hip patients with shortened legs during total hip arthroplasty (THA). Methods Twenty?six cases of developmental dysplasia of the hip received THA between January 2008 and December 2013, in?cluding 12 cases of Crowe typeⅠ, 8 of Crowe typeⅡ, 6 of Crowe typeⅢ. There were 5 males and 21 females with an average age of 62.7 years (range, 36-80 years). The left hip was involved in 9 cases and the right hip in 17 cases. The preoperative mean Har?ris score was 42.30±12.84, and the preoperative mean WOMAC score was 59.08±13.84 at the last follow?up. The anteroposterior X?ray films and CT scan of the pelvis, anteroposterior and lateral X?ray films of the femur, and TraumaCad analysis were conducted routinely preoperation. More than 70%of the bone?implant interface was covered by appropriate upward distance of acetabular im?plant. Results The follow?up time ranged from 6 to 73 months (mean, 36 months). The Harris score improved to 91.18±7.09, and WOMAC score reduced to 9.85±3.75. According to postoperative measurement, affected limb had been lengthened by 0-5 mm in 8 cases, 6-10 mm in 5 cases, 11-15 mm in 5 cases,>15 mm in 7 cases, and shortening increased 1 mm in 1 case, but the average lengthening was 9.23±7.54 mm. The upward distance of acetabular implant was 0-5 mm in 10 cases, 6-10 mm in 7 cases and>10 mm in 9 cases. The average lengthening was 6.60±6.72 mm in patients having 0-5 mm upward distance, 11.90±5.64 mm in patients having 6-10 mm upward distance and 10.11 ± 9.35 mm in patients having>15 mm upward distance, showing no significant differ?ence. The leg length discrepancy was-3.70±6.43 mm in patients having 0-5 mm upward distance, 1.71±6.24 mm in patients having 6-10 mm upward distance and 0.56 ± 7.70 mm in patients having>15 mm upward distance, showing no significant difference. Con?clusion The limb length could be improved by selectively upward placement of acetabular implants in developmental dysplasia of the hip patients with anatomically abnormal acetabulum during THA, with reasonable preoperative design and corrective operation.

A 55-year-old male patient presented with tense bullae on the extremities and trunk.Histological examination revealed subepidermal vesicles and superficial dermal infiltration of eosinophils and lymphocytes.The patient was primarily diagnosed with bullous pemphigoid.However,serum autoantibodies of the patient bound to the dermal side of salt-split skin,and no serum antibodies against BP180,BP230 or type Ⅶ collagen were detected by enzyme-linked immunosorbent assay.Hence,the diagnoses of bullous pemphigoid and epidermolysis bullosa acquisita were excluded.As Western blot and immunoprecipitation analysis showed,there existed antibodies capable of binding to a dermal antigen with a relative molecular mass of 200 000 in the serum of the patient.Based on the above findings,the patient was diagnosed as anti-laminin γ1 (p200) pemphigoid.

Objective To evaluate the reversal effect of a cholinergic receptor agonist on acantholysis in pemphigus,and to investigate its mechanism.Methods Human HaCaT keratinocytes were co-cultured with pemphigus vulgaris immunoglobulin G (PV-IgG) to establish a cell model of pemphigus,then classified into two groups to be incubated with the cholinergic receptor agonist carbachol for 12 hours (test group) or remain untreated (control group).Cell dissociation assay was performed to quantitatively estimate the reversal effect of carbachol on acantholysis,and immunofluorescence assay to qualitatively assess the changes of desmosomal proteins.Radio-immunoprecipitation assay (RIPA) lysis buffer and Triton X-100 were used to lyse HaCaT cells to obtain total proteins and cytoplasmic proteins,and Western blot was conducted to determine the expression levels of adhesion-related proteins desmoglein 3 (Dsg3) and plakoglobin (PG) on the surface of HaCaT cells,as well as the phosphorylation levels of p38 mitogen activated protein kinase (p38 MAPK) and epidermal growth factor receptor (EGFR) at different time points.Quantitative polymerase chain reaction (qPCR) was performed to detect the mRNA expressions of the above surface proteins,and coimmunoprecipitation assay to qualitatively evaluate the interaction between Dsg3 and PG.Results The number of cell debris was significantly lower in the test group than in the control group (18.67 ± 2.52 vs.46.67 ± 2.03,t =11.22,P＜0.01).Immunofluorescence assay showed that carbachol could reverse the internalization of desmosomal molecules induced by PV-IgG.In the pemphigus cell model,the levels of total Dsg3 and PG as well as non-desmosomal Dsg3 were decreased,while the level of non-desmosomal PG increased,and the interaction between Dsg3 and PG was attenuated.When the pemphigus cell model was co-cultured with carbachol,these above changes were reversed.Carbachol also increased the mRNA levels (expressed as 2-△△Ct) of Dsg3 and PG from 1.428 ± 0.215 and 1.563 ± 0.247 in the control group to 4.974 ± 0.948 (t =3.65,P =0.01) and 13.420 ± 1.715 (t =6.85,P ＜ 0.01) in the test group respectively.In phosphorylation assay,carbachol inhibited the phosphorylation of EGFR,but had no significant effect on that of p38 MAPK.Conclusions The cholinergic receptor agonist carbachol can reverse acantholysis in pemphigus,likely by inhibiting the internalization of Dsg3 and PG,enhancing their expressions and interaction,and suppressing the phosphorylation of the key signaling molecule for acantholysis,EGFR.

Objective To establish two nested PCR assays for detection of Trichomonas vaginalis in urine samples from male patients with urethritis,and to evaluate their diagnostic value.Methods One thousand and eighty-eight male patients with urethritis were enrolled from sexually transmitted disease (STD) clinic in the Hospital of Dermatology,Chinese Academy of Medical Sciences and Peking Union Medical College between April 2011 and December 2013.Urethral swabs were collected followed by smear testing,wet mount microscopic examination of Trichomonas vaginalis,and cultivation of Neisseria gonorrhoeae.Urine specimens were also obtained from these patients followed by DNA extraction.Two nested PCR assays were developed and performed to amplify the repeat genomic sequence and β-tubulin gene of Trichomonas vaginalis.Results Trichomonas vaginalis was detected in none of these swab specimens by wet mount microscopy,but in 29 (2.67％) of the urine specimens by either of the two nested PCR assays.Moreover,the positive specimens detected by the two nested PCR assays were completely consistent.Conclusion Compared with wet mount microscopy,nested PCR has higher sensitivity and specificity in detection of Trichomonas vaginalis in urine samples from male patients.

Objective To analyze the clinical characteristics and treatment of acne inversa.Methods Seventeen outpatients with acne inversa were collected in the Institute of Dermatology,Chinese Academy of Medical Sciences and Peking Union Medical College from January l,2012 to December 31,2012.The general condition,clinical feature and treatment of these patients were retrospectively analyzed.Results All the patients were male with the age at onset being about 20 years and disease duration varying from 2 to 50 years.Characteristic clinical manifestations were recurrent tender inflammatory papules,nodules,abscesses,fistulae and sinus tracts in the neck,axillary fossa,groin,perineum and buttocks.Among these patients,10 had a family history and seven were sporadic with mild symptoms.Oral tretinoin combined with antibiotics were the main treatment,and surgical treatment was usually used for severe patients.Conclusions Acne inversa is mainly manifested as abscess,sinus and scars in areas bearing apocrine sweat glands,and therapeutic regimen should be selected according to the severity of lesions.

Objective To investigate the relationship between disease severity and enzyme-linked immunosorbent assay (ELISA) index values of anti-desmoglein (Dsg) 1 and-Dsg3 antibodies in 56 patients with pemphigus,and to characterize fluctuations in anti-Dsg1 and-Dsg3 antibodies in different forms of pemphigus.Methods Fifty-six patients with pemphigus (including 36 cases of pemphigus vulgaris (PV) and 20 cases of pemphigus foliaceus (PF)) were enrolled in this study.Blood samples were obtained from these patients before treatment and at the following four time points:when the condition was relieved and the taper of glucocorticoids began,the dose of glucocorticoids was tapered to half of their initial dose,the maintenance treatment started,and when the duration of maintenance treatment reached two years.ELISA was performed to determine the levels of anti-Dsg1 and-Dsg3 antibodies in these serum samples.Spearman correlation analysis was carried out to assess the relationship between disease severity and ELISA index values,and independent sample's t test to compare the levels of anti-Dsg antibodies among these time points.Results The severity of pemphigus was correlated with anti-Dsg antibody ELISA index values.Both anti-Dsg1 and anti-Dsg3 ELISA index values were significantly reduced at the remission of pemphigus compared with those before treatment (all P ＜ 0.01).At the end of the two-year maintenance treatment,10 (50％) patients with PF and 7 (19.4％) patients with PV became negative for anti-Dsg1 ELISA,whereas only 1 (2.7％) patient with PV became negative for anti-Dsg3 ELISA.Conclusions Anti-Dsg antibody ELISA index value is correlated with disease severity in patients with pemphigus,which may serve as a useful marker for assessing disease severity and activity as well as evaluating therapeutic efficacy.