Objective To analyze KIT gene mutations in one patient with diffuse cutaneous mastocytosis (DCM),and to provide a basis for the prediction of prognosis and selection of treatment.Methods Clinical data were collected from a boy with DCM.Peripheral blood samples were obtained from the patient,his parents and 200 unrelated healthy human controls.PCR was performed to amplify 21 coding exons and their flanking sequences of the KIT gene followed by DNA sequencing.Results A heterozygous missense mutation (c.1526A ＞ T),which leads to the mutation p.Lys509Ile,was detected in the KIT gene of the patient,but not in his parents or the healthy controls.Conclusion The heterozygous missense mutation p.Lys509Ile in the KIT gene may be a cause of DCM.

AIM: To study the effect of high fat diet on the expression of sterol regulatory element biding protein-1 (SREBP-1) and transforming growth factor β_1 (TGF-β_1) in renal tubular cells and rosiglitazone intervention. METHODS: Wistar rats were treated with high fat diet and rosiglitazone for 3 months. The serum glucose, serum insulin and serum triglyceride were detected. Oil Red O staining was used to observe the renal lipid deposit and Masson staining was for the detection of ECM accumulation. SREBP-1, TGF-β_1 and FN protein were determined by the methods of immunohistochemistry and Western blotting. SREBP-1 mRNA was detected by in situ hybridization. RESULTS: Rosiglitazone prevented effectively the increase in serum glucose, serum insulin and serum triglyceride resulted from high fat diet. High fat diet led to lipid droplet formation in renal tubular cells and interstitial ECM accumulation, which was decreased by rosiglitazone treatment. Compared to normal rats, SREBP-1 protein and SREBP-1 mRNA showed high expressions in high fat diet rats that were lowered by rosiglitazone. The precursor segment and mature segment of SREBP-1 protein were decreased by 27.39% and 27.32%. Similarly, the high expressions of TGF-β_1 and FN protein in kidney of high fat diet rats were also prevented by rosiglitazone intervention. Compared to high fat diet rats, the expression of TGF-β_1 in rosiglitazone treatment rats was lowered by 19.14%. CONCLUSION: Rosiglitazone prevents effectively the over-expression of SREBP-1 and TGF-β_1 in renal tubular cells, and decreases lipid accumulation and ECM production in rats fed with high fat diet.

Hematologic Neoplasms ; diagnosis ; pathology ; Humans

Objective To report two cases of Nagashima-type palmoplantar keratoderma(NPPK), and to identify mutations in the SERPINB7 gene. Methods Clinical data were collected from two patients with NPPK and their parents, and peripheral blood samples were obtained from the two patients, their parents and 200 unrelated healthy controls. Genomic DNA was extracted from these blood samples. PCR was performed to amplify 8 exons and their flanking sequences of the SERPINB7 gene followed by DNA sequencing. Results A homozygous mutation (c.796C > T), which led to the formation of a premature termination codon at amino acid position 266 (p.R266*), was identified in both of the two patients. However, the patients′ healthy parents were heterozygous carriers of the mutation(c.796C > T). No mutation was found in the unrelated healthy controls. Conclusion The mutation c.796C > T in the SERPINB7 gene may be responsible for NPPK in the two patients.

Objective To investigate the clinical efficacy of drug phonophoresis therapy in the treatment of acute iridocyclitis.Methods According to randomized block design,104 patients with acute iridocyclitis were divided into the control group of 52 cases (72 eyes)with 1％ atropine mydriasis,oral prednisone tablets and subconjunctival injection of dexamethasone treatment,76 eyes of 52 cases in the treatment group with 1％ atropine mydriasis,oral prednisone tablets and dexamethasone phonophoresis intraocular treatment.Results Compared with the control group,the cure rates of treatment group and control group were 84.2％ and 58.5％ respectively,there was significantly significant difference between the two groups(x2 =12.598,P =0.000),oral hormone time from the beginning to the first reduction[treatment group and control group were (5.12 ± 1.00) d and (7.32 ± 0.97) d respectively (t =-13.495,P =0.000)] and oral hormone total time [treatment group and control group were (27.82 ± 4.84) d and (35.49 ± 4.74) d respectively (t =-9.720,P =0.000)] were significantly shortened,complications decreased significantly[conjunctival edema rate (x2 =9.657,P =0.002),subconjunctival hemorrhage rate (x2 =6.601,P =0.010),conjunctival scarring rate (x2 =4.340,P =0.037),pain rate (x2 =63.419,P =0.000) and oculocardiac reflectivity rate (x2 =33.293,P =0.000)] and patient satisfaction improved significantly (treatment group and control group were 94.7％ and 69.4％ respectively) (x2 =16.333;P =0.000).Conclusion Dexamethasone phonophoresis therapy has better clinical efficacy and higher cure rate,and it is non-invasive,safe and reliable,less complications and high satisfaction in the treatment of acute iridocyclitis.

Aim To investigate the expression of SREBP-1(sterol regulatory element binding protein-1) in the kidney of type 1 diabetic rats and the effect of insulin.Methods The type 1 diabetic models were induced by high dose of STZ and rats were randomly divided into three groups: normal control group,diabetes control group and insulin treated group.At the 2nd week end,the triglyceride(TG) content in the kidney of experimental rats was measured by the assay kit and oil Red O staining.Furthermore,the expression of SREBP-1 protein was detected by the methods of Western blot and immunohistochemistry.The analysis of SREBP-1 mRNA was performed by in situ hybridization.Results Compared with the control group,the type 1 diabetic rats' renal triglyceride content markedly increased,and the result of Oil Red O showed that lipid deposited in the renal tubular epithelium.Triglyceride content markedly decreased after insulin treatment.The difference had statistic meaning,compared with the diabetes model group.Immunohistochemistry presented the results that SREBP-1 protein was up-regulated in renal tubular epithelium of diabetic rats and insulin treatment suppressed the increasing.The results of western blot showed that the precursor and mature segments of SREBP-1 protein in kidney of diabetes group rats were about 1.86 times and 1.77 times respectively of that of normal control group rats.In situ hybridization confirmed the increasing of SREBP-1 mRNA in renal tubular epithelium in diabetic rats.The effect of insulin treatment on SREBP-1 expression was detected by the methods of Western blot and in situ hybridization and it was found that the SREBP-1 mRNA and protein of kidney were down-regulated.Compared with the normal group,the difference has statistic meaning(P

Objective To screen for genomic copy number variants(CNVs)in six neonates with Pierre Robin sequence(PRS)by Affymetrix 2.7 M chip to identify possible loci related to PRS.Methods Six neonates with PRS admitted into the Department of Neonatology,Children's Hospital of Fudan University from June 2009 to May 2010 were enrolled in this study.CNVs were detected by Cytogenetic Whole Genome 2.7 M array.Rare CNVs with potential clinical significance that deletion segments' size ＞50 kb and duplication segments' size ＞200 kb were selected based on the analysis of Chromosome Analysis Suite(ChAS)software,false positive CNVs and segments of normal population were excluded.The identified CNVs were compared with those in relative published literatures.Results(1)Among 6 PRS patients,two patients had facial deformation,two had congenital heart defects,one had congenital dysplasia of the laryngeal cartilage and one had choroidal space occupying lesion.(2)Seven rare CNVs whose size from 51-11 956 kb were identified in four neonates,including a 739 kb duplication on lp26.23-p36.22,a 6273 kb deletion on lq43-44,a 51 kb and a 55 kb deletions on 14q32.31,a 1022 kb duplication on 14q11.1-11.2,a 11 956 kb duplication on 20p13 and a 105 kb deletion on 4q23.3.(3)Published literatures showed that deletions of 1q43-44 and 14q32.31 might relate to micro/retrognathia and abnormal palate.Region of chromosome 1q43-q44 contained AKT3 and heterogeneous nuclear ribonucleoprotein U(hnRNPU)genes,and the haploinsufficiency of AKT3 and hnRNPU genes might cause developmental human microcephaly and agenesis of the corpus callosum,speech delay and seizures respectively.Region of chromosome 14q32.31 contained some C/D small nucleolar RNA,and the human imprinted 14q32 domain shared common genomic features with the imprinted 15q11-q13 loci.Conclusions This study established a method to discover whole genome CNVs in identifying novel submicroscopic deletions and duplications.Reviewing of published literatures suggested that deletions of chromosome 1q43-q44 and 14q32.31 might cause Pierre Robin sequence.

Objective To detect the mutation of CARD15 gene in a patient with sarcoidosis and tuberculosis.Methods Clinical data were collected from a 32-year-old male patient with early-onset sarcoidosis and tuberculosis.Peripheral blood was obtained from the patient,both of his parents,and 102 healthy controls.All the 12 exons of the coding regions as well as flanking intronic sequences of the CARD15 gene were amplified by PCR followed by direct sequencing.The resulted sequences were blasted against the reference sequences of CARD15 gene.Results Both clinical features and histopathological findings of the patient were consistent with sarcoidosis.Furthermore,the patient presented with flexion contractures of fingers and toes,as well as iridocyclitis.A heterozygous missense recurrent mutation c.1000C ＞ T (p.R334W) was detected in exon 4 of the CARD15 gene in the patient,but not in either of his parents or any of the 102 healthy controls.Conclusions A p.R334W mutation in the CARD15 is identified in the patient,which may be responsible for the clinical phenotype of earlyonset sarcoidosis.Gene analysis may be a useful method to clarify the etiology of early-onset sarcoidosis.

Objective To investigate the role of Ras-related C3 botulinum toxin substrate 1/mitogen-activated protein kinase/extracellular signal-regulated kinase (Rac 1/MAPK/ERK) signal pathway in rats with ventilator induced lung injury (VILI) and its mechanism.Methods Thirty Sprague-Dawley (SD) rats were randomly divided into spontaneous respiration group,normal tidal volume (VT) group and high VT group with 10 rats in each group.The rats in spontaneous respiration group were kept their spontaneous breathing.The rats in normal VT group and high VT group were performed tracheal intubation after tracheostomy,and underwent mechanical ventilation on bilateral lungs with 6 mL/kg and 40 mL/kg VT respectively with maintenance anesthesia.After 4-hour ventilation,heart blood,bronchoalveolar lavage fluid (BALF) and lung tissues were harvested.The levels of interleukins (IL-1β,IL-6),tumor necrosis factor-α (TNF-α),myeloperoxidase (MPO) and macrophage inflammatory protein-2 (MIP-2) in serum and BALF were determined by enzyme linked immunosorbent assay (ELISA).Lung wet/dry radio (W/D) was determined.The lung tissues were stained with hematoxylin and eosin (HE),and pathological changes were observed,and pathological scores were evaluated.The ultra structure changes in type Ⅱ alveolar epithelial cells (AEC Ⅱ)were observed with transmission electron microscope.The positive expressions of phosphorylation of extracellular signal-regulated kinase (p-ERK) were determined by immunohistochemistry,and those of Racl and F-actin were determined by immunofluorescence.The mRNA expressions of ERK and Rac1 were determined by real-time fluorescent quantitation reverse transcription-polymerase chain reaction (RT-qPCR),and protein expressions of Rac-1,p-ERK and F-actin were determined by Western Blot.Results ① Compared with spontaneous breathing group,lung W/D in both mechanical ventilation groups was significantly increased,with more significant increase in the high VT group (6.64 ± 0.88 vs.1.79 ± 0.36,P ＜ 0.01).② There was no obvious pathological changes in the lung tissue and AEC Ⅱ of the spontaneously breathing group.In the normal VT group,there was slight edema and infiltration of inflammatory cells;AEC Ⅱ had less lamellar bodies and uniform distribution of the villi of the alveolar epithelium.In the high VT group,the edema of the lung tissue,the widening of the pulmonary septum,the alveolus congestion,the infiltration of inflammatory cells,and alveolar structure disorder were found;and AEC Ⅱ was irregular,the number of lamellar bodies in the plastids was decreased and was unevenly distributed.The pulmonary histopathological score in the high VT group was significantly higher than that in the spontaneous breathing group and the normal VT group (12.00 ± 2.00 vs.6.00 ± 1.51,8.50 ± 0.53,both P ＜ 0.01).③ Compared with spontaneous breathing group,IL-1β,IL-6,TNF-α,MPO,and MIP-2in serum and BALF in both mechanical ventilation groups were significantly increased,with more siguificant increase in the high VT group [serum IL-1 β (ng/L):104.2 ± 15.1 vs.20.3 ± 8.3,IL-6 (ng/L):46.6 ± 11.5 vs.22.7 ± 7.5,TNF-α (ng/L):39.4±6.5 vs.5.4± 1.9,MPO (ng/L):0.66±0.24 vs.0.06±0.03,MIP-2 (ng/L):109.2±25.8 vs.22.8±8.4;BALF IL-1 β (ng/L):121.5 ± 25.6 vs.24.0 ± 7.5,IL-6 (ng/L):136.7 ± 32.7 vs.31.4 ± 10.5,TNF-α (ng/L):98.0 ± 14.8vs.10.1 ±2.6,MPO (ng/L):0.80±0.31 vs.0.08±0.04,MIP-2 (ng/L):144.4±28.9 vs.41.2±20.7;all P ＜ 0.01].④ There were only a few p-ERK,Rac1 and F-actin positive expressions in the spontaneous breathing group.The positive expressions in normal VT group were increased.In high VT group,the positive expression of p-ERK was significantly increased;Rac1 and F-actin were mainly distributed in the cell membrane and cytoplasm respectively,the positive expressions were further enhanced.⑤ The gene expressions of ERK and Rac1,and protein expressions of p-ERK,Rac1 and F-actin in the high VT group were significantly higher than those in the spontaneous breathing group and normal VT group [ERK mRNA (2-△△Ct):8.23±2.83 vs.1,3.02± 1.38,p-ERK protein (gray value):1.15±0.36 vs.0.61 ±0.23,0.88±0.22;Rac1 mRNA (2-△△Ct):4.45 ±2.26 vs.1,1.22±0.39,Rac1 protein (gray value):0.91 ±0.16 vs.0.48±0.11,0.55 ± 0.10;F-actin protein (gray value):0.70± 0.09 vs.0.49 ± 0.08,0.55 ± 0.04;all P ＜ 0.01].Conclusion F-actin expression in lung tissue was up-regulated in rats with VILI,which resulted in reconstruction of AEC Ⅱ cyto-skeleton,and variation of cell membrane permeability through Rac 1 /MAPK/ERK sigualing pathway during VILI.

Objective To observe the ultrastructural features of recessive dystrophic epidermolysis bullosa inversa(RDEB-Ⅰ)and to detect the mutations of COL7A1 gene in a family with RDEB-Ⅰ.Methods A 24-year-old male patient complained of recurrent vesicles in the skin for 24 years.The lesions began as generalized pruritic vesicles and bullae soon after birth,with a predilection for areas subject to friction,and showed a trend to be worse in summer but mild in winter.No photosensitivity was observed.When he was 3 to 4 years old,the lesions were decreased in number,with the only involvement of the trunk and abdomen;thereafter,the lesions were improved year by year.The patient suffered from nephritis at the age of 5 years,which progressed into renal failure at the age of 15 years.He received renal transplantation and was given long-term oral tacrolimus and mycophenolate mofetil,which leaded to an improvement in the lesions.The family history was unremarkable,and the marriage between her parents was not consanguineous.Dermatological examination revealed large areas of irregularly-marginated,hypopigmented,atrophic scar on the waist,back and abdomen with onychodystrophy involving multiple nails.No vesicles were observed.Immunofluorescence antigen mapping and transmission electron microscopy were conducted to observe the expression of type Ⅶ collagen in and ultrastructure of cutaneous lesions from the patient.Venous blood samples were obtained from the patient as well as his parents and 3 sisters,and drill biopsy specimens were obtained from the margin of vesicular lesions and unaffected anterior tibial skin of the patient.DNA specimens were obtained from the blood samples of the family members and 150 unrelated healthy controls,and RNA was extracted from the biopsy samples of the patient.PCR and direct sequencing were carried out to detect mutations in COL7A1 gene,and reverse transcription-PCR was conducted to confirm the mutation at mRNA level.Results Skin cleavage was observed under lamina densa in the dermis,with a decrease in anchoring fibrils and expression of type Ⅶ collagen in the lesions of the patient.A heterozygous synonymous mutation c.C5499T which created a new splicing site and leaded to a premature terminal codon,as well as a heterozygous missense mutation c.C6205T(C-T transition at codon 2069:CGT to TGT)which leaded to the substitution of arginine by cysteine,were identified in the COL7A1 gene of the proband and all of his sisters,but not in any of the unrelated controls.The c.C5499T and c.C6205T mutations were inherited from the patient's father and mother respectively.Conclusion The compound heterozygous mutations c.C6205T and c.C5499T may be responsible for RDEB-Ⅰ in this patient.