Preliminary investigation of methylation status of microRNA-124a in spinal cords of rat fetuses with congenital spina bifida.

OBJECTIVE: We investigated the expression of microRNA-124a and its methylation status in the spinal cords of rats with congenital spina bifida versus rats with normal fetuses. METHODS: Real-time quantitative reverse transcription-polymerase chain reaction was used to compare the expression of microRNA-124a in the spinal cords of 42 rats with all-trans retinoic acid induced congenital spina bifida and 42 rats with normal fetuses. The DNA methylation status in the promoter region of miRNA-124a was detected using methylation specific-PCR. RESULTS: Compared with rats with normal fetuses, expression of microRNA-124a was significantly decreased in rats with congenital spina bifida fetuses. The percentages of spinal cords with DNA hypermethylation in the microRNA-124a promoter were 81% and 14% in the congenital spina bifida and normal control groups, respectively. The difference was statistically significant. Further apoptosis testing revealed increased apoptosis cell numbers in the congenital spina bifida samples. Meanwhile, the phosphorylated mitogen-activated protein kinase protein expression level dramatically decreased in the congenital spina bifida samples. CONCLUSION: Aberrant DNA methylation was responsible for down-regulation of microRNA-124a by regulating the mitogen-activated protein kinase pathway, suggesting that microRNA-124a is a potential diagnostic biomarker in congenital spina bifida.

Altered microRNA expression profiles in a rat model of spina bifida.

MicroRNAs (miRNAs) are dynamically regulated during neurodevelopment, yet few reports have examined their role in spina bifida. In this study, we used an established fetal rat model of spina bifida induced by intragastrically administering olive oil-containing all-trans retinoic acid to dams on day 10 of pregnancy. Dams that received intragastric administration of all-trans retinoic acid-free olive oil served as controls. The miRNA expression profile in the amniotic fluid of rats at 20 days of pregnancy was analyzed using an miRNA microarray assay. Compared with that in control fetuses, the expression of miRNA-9, miRNA-124a, and miRNA-138 was significantly decreased (> 2-fold), whereas the expression of miRNA-134 was significantly increased (> 4-fold) in the amniotic fluid of rats with fetuses modeling spina bifida. These results were validated using real-time quantitative reverse-transcription polymerase chain reaction. Hierarchical clustering analysis of the microarray data showed that these differentially expressed miRNAs could distinguish fetuses modeling spina bifida from control fetuses. Our bioinformatics analysis suggested that these differentially expressed miRNAs were associated with many cytological pathways, including a nervous system development signaling pathway. These findings indicate that further studies are warranted examining the role of miRNAs through their regulation of a variety of cell functional pathways in the pathogenesis of spina bifida. Such studies may provide novel targets for the early diagnosis and treatment of spina bifida.

Role of growth hormone in maturation and activation of dendritic cells via miR-200a and the Keap1/Nrf2 pathway.

OBJECTIVES: Dendritic cells (DCs) are antigen-presenting cells that participate in the immune response; recently, it has been reported that growth hormone (GH) promotes their maturation. The aim of this study was to investigate mechanisms by which GH acts on DC maturation and activation. MATERIALS AND METHODS: Human peripheral blood monocytes (HPBMs) were induced to become immature DCs and treated with GH to obtain mature DCs. An osteosarcoma mouse model was established by injection of LM8 cells to investigate anti-tumour effect of GH-induced DCs in vivo. RESULTS: After administration of GH, DCs reduced miR-200a expression and nuclear Nrf2 accumulation; miR-200a down-regulation inhibited DC maturation. Nrf2 ubiquitination level was increased by Keap1 overexpression in murine bone marrow derived dendritic cells (BMDCs), which was cancelled by miR-200a in GH exposed cells. In vivo, tumour volume was significantly reduced by GH-treated DCs and the effect was reversed by overexpression of miR-200a. CONCLUSIONS: GH promoted maturation and activation of DCs, and regulation of miR-200a played a part in this process by modulation of the Keap1/Nrf2 pathway.

Surgical management of intrahepatic vessels in children with stage III/IV hepatoblastoma.

BACKGROUND: Hepatoblastoma (HB) is a rare childhood tumor. We investigated the effect of intraoperative management of the intrahepatic major vessels in children with HB. METHODS: Between April 2005 and August 2012, surgical resection was performed on 50 children with hepatoblastoma. These children were divided into a vessel-ligation group (n = 20) and a vessel-repair group (n = 30). In the vessel-ligation group, the intrahepatic major vessels were ligated and removed together with the tumor and the affected liver lobe/liver parenchyma. In the vessel-repair group, the affected intrahepatic major vessels were dissected and preserved as much as possible and the normal liver lobe/liver parenchyma and blood supply from these vessels were also preserved. The outcomes were analyzed by postoperative follow-up. RESULTS: In the vessel-ligation group, two patients gave up surgery, six patients underwent palliative resection, and 12 patients underwent en bloc resection; four patients died of liver failure and eight patients fully recovered and were discharged. In the vessel-repair group, all 30 patients underwent en bloc resection and were discharged after satisfactory healing. After a follow-up time of 5 - 36 months (median: 20 months), two patient in the vessel-ligation group survived and 22 patients in the vessel-repair group survived. CONCLUSIONS: Patients with HB can be successfully treated by tumor resection with vascular repair. This method prevents postoperative liver failure, ensures patient safety during the perioperative period, and allows for early chemotherapy.

[MicroRNA differential expression profile in nephroblastoma cell line versus normal embryonic kidney cell line].

OBJECTIVE: To explore the microRNA (miRNA) differential expression profile between nephroblastoma cell line G401 and normal embryonic kidney cell line CCC-HEK-1 so as to provide rationales for the role of miRNA in the pathogenesis of nephroblastoma. METHODS: Three samples from G401 cell line and another 3 samples from CCC-HEK-1 cell line were chosen as the experimental group and the control group respectively. miRNA profiles in these samples were analyzed by microarray. The threshold value used to screen up and down-regulated miRNAs was fold change ≥ 2 or ≤ 50.0%. And real-time PCR was used to validate the reliability of microarray. RESULTS: Seventy-one miRNAs were found up-regulated in the experimental group and 11 of them were more than 8 times versus control group. In addition, 59 miRNAs were found down-regulated and 11 of them were 12.5% versus control group. Overall, there were significant differences in the expression of miRNA between G401 and CCC-HEK-1 cell lines. CONCLUSIONS: There is a differential expression of miRNA between G401 and CCC-HEK-1 cell lines. It may be related with occurrence and metastasis of nephroblastoma.

Effect of growth hormone on the immune function of dendritic cells.

BACKGROUND: Dendritic cells (DCs) are one of the most important antigen presenting cells in the human body, and DCs at various stages of maturation possess different or even opposite functions. The aim of this study was to investigate the influence of growth hormones on the functional status of cord blood-derived DCs encompassing immunophenotype, ability to excrete interleukin (IL)-12 and provoke autologous leukomonocyte. METHODS: Mononuclear cells were isolated from fresh cord blood, with IL-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) used to induce and stimulate the mononuclear cells. Growth hormone at different concentrations was used to modify DCs, and then DCs morphology, number and growth status were observed. The immunophenotype of DCs was detected with a flow cytometer. The concentration of IL-12 in the DCs supernatant was determined by enzyme linked immunosorbent assay (ELISA) and DCs functional status was evaluated by autologous mixed lymphocyte reactions. RESULTS: Mononuclear cells from cord blood can be differentiated into DCs by cytokine induction and growth hormone modification. With the increase in growth hormone concentrations (5 - 100 microg/L), the expression of DCs HLA-DR, CD1alpha, CD80 and CD83 were significantly increased (P < 0.05). The ability of DCs to secrete IL-12 was significantly improved (P < 0.05), and the ability of DCs to activate autologous lymphocytes was significantly enhanced (P < 0.05). Pegvisomant was able to ablate the effects of growth hormone on DCs. CONCLUSIONS: Growth hormone may facilitate DCs induction and maturation, and improve the reproductive activity of autologous lymphocytes in a dose-dependent manner. Growth hormone may serve as a factor of modifying DCs to achieving maturity.

Effect of insulin on functional status of cord blood-derived dendritic cells and on dendritic cell-induced CTL cytotoxicity against pancreatic cancer cell lines.

BACKGROUND: Dendritic cells (DCs) are the most important antigen-presenting cells in the human body, and DCs with different mature status possess different or even opposite functions. This study was designed to explore the influence of insulin on the functional status of cord blood-derived DCs and on DC-induced cytotoxic T lymphocyte (CTL) activity against pancreatic cancer cell lines. METHODS: Mononuclear cells were isolated from fresh cord blood. Interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) were used to induce or stimulate the mononuclear cells. Insulin at different concentrations served to modify DCs, and then DC morphology, number, and growth status were assessed. The DC immunophenotype was detected with a flow cytometer. The IL-12 in DC supernatant was determined by ELISA. DC functional status was evaluated by the autologous mixed lymphocyte reaction. T lymphocytes were induced by insulin-modified DCs to become CTLs. The CTL cytotoxicity against pancreatic cancer cell lines was determined. RESULTS: Mononuclear cells from cord blood can be differentiated into DCs by cytokine induction and insulin modification. With the increase in insulin concentration (2.5-25 mg/L), the expression of DC HLA-DR, CD1alpha, CD80, and CD83 was significantly increased, the DC ability to secrete IL-12 was significantly improved, DC function to activate autologous lymphocytes was significantly enhanced, and the cytotoxicity of CTLs induced by insulin-modified DCs against pancreatic cancer cell lines was significantly strengthened. CONCLUSIONS: Insulin may facilitate DC induction and maturation, and improve the reproductive activity of autologous lymphocytes. The cytotoxicity of CTLs induced by insulin-modified DCs against pancreatic cancer cell lines was significantly enhanced. Insulin may serve as a factor modifying DCs and inducing CTLs in vitro in insulin biotherapy.

[Detection and identification of specific serum biomarkers in papillary thyroid cancer].

OBJECTIVE: To detect and identify the potential specific serum biomarkers for diagnosis of papillary thyroid cancer. METHODS: Samples of 35 patients with papillary thyroid carcinoma, 40 patients with benign thyroid nodule and 34 healthy individuals were analyzed using the SELDI-TOF ProteinChip System and bioinfomation technology to find the differential peaks which were separated by HPLC and then further analyzed by LC-MS/MS. The protein sequences were analyzed by SEQUEST software and searched in Bioworks database. RESULTS: The top six mass-to-charge ratio (M/Z) peaks with the smallest P value were 6651, 6452, 7653, 7932, 15 106 and 15 848 Da, respectively. The 6651 and 6452 Da proteins were weakly expressed in papillary thyroid carcinoma but highly expressed in benign thyroid nodules and healthy individuals. The differences had statistical significance (P < 0.01). The 7653, 7932, 15 106, 15 848 Da proteins were highly expressed in papillary thyroid carcinoma but weakly expressed in benign thyroid nodules and healthy individuals. The differences were statistically significant (P < 0.01). Combination of these six proteins, using the method of leave-one-out to make crossing detection, the specificity of discriminating papillary thyroid carcinoma and non-cancer was 88.0%, and its sensitivity was 92.5%. The 6651 and 6452 Da proteins were identified as apolipoprotein C-I and apolipoprotein C-III, respectively. The 7653 and 15 106 Da proteins were identified as the same protein-alpha-globin, and the 7932 and 15,848 Da proteins were identified as the same protein-beta-globin. CONCLUSION: The detection of differentially expressed apolipoprotein C-I, apolipoprotein C-III, alpha-globin, and beta-globin may have utility for diagnosis of papillary thyroid carcinoma and are worthy of further investigation.

[Screening and identification analysis of serum protein biomarkers in nephroblastoma].

OBJECTIVE: To screen and characterize the serum protein biomarkers in nephroblastoma so as to establish the proteins as the specific serum biomarkers for diagnosis and prognosis monitoring. METHODS: The differential protein peaks were located by detecting serum samples of preoperative and postoperative patients and normal children using the SELDI-TOF MS technology. After purification, the differential proteins were further analyzed by LC-MS/MS and the protein sequences searched in database. RESULTS: Two peaks with m/z of 6455.5 and 6984.4 were selected as potential biomarkers. They were weakly expressed in nephroblastoma (intensity: 1029 +/- 364, 297 +/- 126) but highly expressed in normal individuals (2108 +/- 837, 753 +/- 226); another peak with m/z of 9190.8 was weakly expressed in preoperative sera (283 +/- 154) but highly expressed in serum samples of postoperative patients and normal children (5974 +/- 657, 6231 +/- 519). The protein at 6455.5 and 9190.8 were identified as apolipoprotein C-III and haptoglobin respectively. CONCLUSION: The detection of differentially expressed apolipoprotein C-III and haptoglobin may have potential utilities for serum diagnosis, malignancy classification and prognosis monitoring of nephroblastoma and is worthy of further studies and applications.

[Growth hormone influence on functional status of cord blood-derived dendritic cells].

OBJECTIVE: To detect the effect of functional status and immunophenotype of dendritic cell modified with growth hormone (GH) from by cord blood. METHODS: Monocyte was isolated from cord blood, DCs were induced with IL-4 & GM-CSF and modified with different concentration of GH. The morphology, count and growth status of DC were observed appearance. And the immunophenotype of DC was detected with flow cytometry. IL-12 in supernatant was measured with ELISA. The immuno-activity was determined with autologous mixed lymphocyte reaction. RESULTS: Mature DC were induced from adherent lymphomonocyte under the action of IL-4, GM-CSF and GH of different densities (5, 25, 100 ng/ml). GH increased the positive expression of CD83 (31% +/- 4%, 34% +/- 4%, 34% +/- 5%), CD80 (26% +/- 4%, 29% +/- 6%, 30% +/- 6%)and HLA-DR (45% +/- 8%, 49% +/- 6%, 50% +/- 5%) (P < 0.05), enhanced the ability of secreting IL-12 (P < 0.05), and boosted significantly the immuno-activation of DC to autologous lymphocyte (P < 0.05). CONCLUSION: GH could promote the differentiation, maturation and the expression of costimulatory molecules, increase the IL-12 production in DC and enhanced the ability of DC to stimulate the proliferation of lymphocyte within a certain dose range.

Detection and significance of serum protein marker of Hirschsprung disease.

OBJECTIVE: The objective of this study was to identify a specific fingerprint chromatogram model of serum proteins for early screening and diagnosis of Hirschsprung disease. METHODS: To detect the protein mass spectrograms of 78 serum specimens (42 specimens of Hirschsprung disease, 16 specimens of adhesive ileus including appendicitis and Meckel diverticulum after operation and inflammatory bowel disease, and 20 specimens of normal control subjects), we used surface-enhanced laser desorption/ionization time of flight mass spectrometry technology, combined with bioinformatics methods (support vector machine) to develop and compare protein mass spectrograms from serum samples. RESULTS: We identified 3 protein markers, the mass-to-charge ratio of which is positioned at 3221.7, 5639.2, and 6884.2 from the fingerprint chromatogram model of serum protein for early screening and diagnosis of Hirschsprung disease. The markers had 100% sensitivity and specificity. CONCLUSION: The fingerprint chromatogram model of serum protein using surface-enhanced laser desorption/ionization time of flight mass spectrometry technology combining support vector machine is a new method of early screening and diagnosis of Hirschsprung disease that is worthy of additional research and application.

Effects of cytotoxic T lymphocytes on hepatoma cell line SMMC-7721 induced by different subsets of dendritic cells in vitro.

BACKGROUND: Dendritic cells (DCs) loaded with complex antigen are always used to induce cytotoxic T lymphocytes (CTLs) which have a specific anti-tumor activity. However, CTLs can assault autologous cells induced by DCs loaded with autologous antigen. This study aimed to explore how to weaken the autoimmune reaction induced by DC vaccine by combining mature DC (mDC) activating immunity and immature DC (imDC) leading to immune tolerance to make hepatocellular carcinoma (HCC) vaccine in vitro. METHODS: DC progenitors derived from human peripheral blood were assigned to two groups. One was cultured to mDC and pulsed with frozen-thawed antigen (FTA) of human HCC cell line SMMC-7721 cells (mDC group), and the other was cultured to imDC and pulsed with FTA of human liver cell line L-02 cells (imDC group). The morphology of DCs was monitored and cells phenotypes including HLA-DR, CD80, CD1alpha, CD83 were assayed by flowcytometry (FCM). The concentrations of interleukin-12 (IL-12) in the supernatant were assayed by ELISA. Methyl thiazolyl tetrazolium (MTT) was used to evaluate T cell proliferation induced by mDC and imDC and the killing rate of CTL induced by mDC and imDC respectively/together on SMMC-7721 and L-02 cells. RESULTS: Compared with the imDC group, the mDC group was characterized by the following: increased secretion of IL-12 (P<0.05); higher expression of HLA-DR, CD1alpha, CD80, CD83; and stronger activity in stimulating proliferation of isogenic T cells (P<0.05). CTL induced by the mDC group had a significant killing response to SMMC-7721 as well as a higher killing rate for L-02 (P>0.05). CTL induced by mDC and imDC together had a higher killing response to SMMC-7721, but a lower killing rate for L-02 (P<0.01). CONCLUSIONS: CTL induced by mDC and imDC together has a higher antigen-specific killing response in vitro than that induced by mDC alone. This may be of greater clinical value.

Application of serum protein fingerprint in diagnosis of papillary thyroid carcinoma.

To find new biomarkers and establish serum protein fingerprint models for early diagnosis and preoperative staging of papillary thyroid carcinoma, we employed SELDI-TOF-MS and bioinformatics tools. A total of 116 samples were analyzed in this study. The first 80 samples were analyzed by SELDI-TOF-MS and two biomarker patterns were identified. Pattern 1 distinguishes patients with papillary thyroid carcinoma from healthy individuals. Pattern 2 distinguishes papillary thyroid carcinoma from benign thyroid nodes. The remaining 29 samples were analyzed on the second day and served as an independent test set. The analysis of this independent test set yielded a specificity of 80.0% and a sensitivity of 88.9% for pattern 1 and a specificity of 80.0% and a sensitivity of 80.0% for pattern 2. Two additional biomarker patterns were identified to distinguish different stages of the papillary thyroid carcinoma (pattern 3) with an accuracy of 77.1% and different pathological types of thyroid carcinoma (pattern 4) with an accuracy of 88.1%. Taken together, the SELDI-TOF-MS technique combined with bioinformatics approaches can not only facilitate the discovery of better biomarkers for papillary thyroid carcinoma but also provide a useful tool for molecular diagnosis in the future.

[Application of serum protein fingerprint model and support vector machine in diagnosis of thyroid cancer].

OBJECTIVE: To detect new biomarkers and to establish a serum protein fingerprint model for early detection and diagnosis of thyroid cancer. METHODS: The serum samples of 40 thyroid cancer patients, 9 thyroid adenoma patient, and 32 healthy individuals were randomly divided into 2 sets: training set (n = 66, including 32 thyroid cancer patients, 9 thyroid adenoma patients, and 25 healthy individuals) and test set (n = 15). The serum protein were bound to WCX2 chip and tested by surface enhanced laser desorption/ionization time of flight-mass spectrometry (SELDI-TOF-MS). The data of spectra were analyzed by support vector machine (SVM) to establish a diagnostic model. RESULTS: The detective model combined with 3 biomarkers could differentiate the serum of thyroid cancer from that of healthy individual with a specificity of 86% and a sensitivity of 100%. The diagnostic model combined with 3 biomarkers could differentiate thyroid cancer from thyroid adenoma with a specificity of 88.9% and a sensitivity of 96.9%. The positive predictive value to differentiate papillary thyroid carcinoma from the thyroid cancer of other types was 97%, and the positive predictive value of thyroid carcinoma of other pathological types was 71%. CONCLUSION: The combination of SELDI with bioinformatics tools is a novel, effective, and highly specific and sensitive method for thyroid cancer detection and diagnosis.

[Detecting biomarkers from serum in nephroblastoma patients with support vector machine].

OBJECTIVE: To find new biomarkers and to establish serum protein fingerprint models for early detection and diagnosis of nephroblastoma by SELDI-TOF-MS and bioinformatics tools. METHODS: Seventy five serum samples from 35 nephroblastoma patients, 30 children's abdominal solid tumor patients, and 20 healthy children were bound to WCX2 protein chip and tested by surface enhanced laser desorption/ionization time of flight-mass spectrometry (SELDI-TOF-MS). The data of spectra were analyzed by support vector machine(SVM). RESULTS: Four peaks with m/z of 6984.5, 6455.5, 6914.0, 3256.7 were selected as potential biomarkers. The detective model combined with 2 biomarkers could separate nephroblastoma from the healthy group with a sensitivity of 100%, and a specificity of 100%. The diagnostic model combined with 2 biomarkers could separate nephroblastoma from other child's abdominal solid tumors with a sensitivity of 93.3%, and a specificity of 100%. CONCLUSION: High sensitivity and specificity achieved by this method show great potential for early diagnosis of nephroblastoma, and screening for new tumor biomarkers.

[Clinical study of multiple zonal aganglionosis in long segment Hirschsprung's disease].

OBJECTIVE: To discuss pathogeny and diagnosis and management of multiple zonal aganglionosis in Hirschsprung's disease. METHODS: Records of 3 children, aging 5 days, 29 days and 18 months, 3 boys, with multiple zonal aganglionosis in long segment Hirschsprung's disease between 1987-2005 were reviewed retrospectively. Total colectomy and Soave's operations were performed. RESULTS: 3 children were diagnosed HD before surgery, but the convulsive stenosis in distal ileum and proximal ascending colon were detected during surgery. The aganglionic cells in the stenosis gut were confirmed by pathologic diagnosis. CONCLUSIONS: The total colon and ileum should be detected carefully during surgery in children with long segment Hirschsprung's disease. It is believed not sound that the neuroblastic cells stop moving from neuroectoderm to gut in early gestation in HD, but it is also believed that some other causes in the course of gestation might interfere the normal growth of the ganglionic cells.

[Detection of methylation of hMSH2 gene promoter region of esophageal cancer].

OBJECTIVE: To detect methylation in promoter region of hMSH2 gene in esophageal cancer. METHODS: Specimens of cancer and normal tissues freshly removed from 32 cases of esophageal cancer patients without previous radiotherapy, chemotherapy or other treatment were preserved at -80 degrees C within 30 min. Methylation specific PCR (MSP) was used to detect methylation of mismatch repair gene (MMR) hMSH2 in promoter region in esophageal cancer and normal esophageal tissues. RESULTS: The frequencies of methylation of hMSH2 gene in promoter region of cancer and normal esophageal tissues were 32.4% (11/32) and 0/30 (0%), respectively, and significant difference was found between the two groups (P < 0.01). The frequency of methylation in elder patients (> or = 70 years old) was significantly higher than that in younger patients (< 70 years old) (P < 0.05). Methylation was less frequently found in grade I-II (18.2%) than in grade III-IV (70.0%) (P < 0.05). CONCLUSION: Methylation of hMSH2 gene in promoter region is related to patients' age and histopathological grade of the esophageal cancer.

[Expression of hMSH2 gene in esophageal cancer].

OBJECTIVE: To observe the expression of DNA mismatch repair gene hMSH2 mRNA in esophageal cancer tissues. METHODS: This study included 32 esophageal cancer patients who received no previous radiotherapy, chemotherapy or other treatments. Within 30 min following surgical removal of the tumor tissues, specimens of the tumor, the tissue adjacent to the tumor and normal tissue at the esophageal stump (1 cmx1 cmx1 cm in size for each specimen) were obtained for examining hMSH2 expression with hMSH2 ISH detection kit. RESULTS: The positivity rate of hMSH2 was 46.88% in the esophageal cancer tissues, 53.12% in the adjacent tissues, and 84.38% in normal tissues at the esophageal stump, showing significant difference of the former two tissues from the normal tissue (P<0.05). No significant correlation was noted between the positivity rate of hMSH2 and such factors as the patients' age, sex, tumor size, tumor location, pathological type, histological grade, lymphatic metastasis or degree of tumor invasion (P>0.05). CONCLUSION: The deletion of hMSH2 is an early event in the development of esophageal cancer.

A study of prognosis in adult onset Still's disease patients / 中华风湿病学杂志

Objective To investigate the potential clinical factors associated with the prognosis and relapse of adult onset Still's disease(AOSD).Methods The factors possibly influencing the prognosis and relapse of AOSD were analyzed by logistic regression and COX regression in the cohort study.Ninety-six con- secutive inpatients of AOSD diagnosed based on Yamaguchi criteria in the hospital from March 1996 to September 2004 were included in the study.Results Nine cases(9.4%)were lost during the follow-up. Eleven patients(12.6%)were diagnosed as other diseases(5 with other rheumatic diseases,4 with tumor and 2 with infections)in the 87 follow-up cases.In 76 cases,3 patients(3.9%)died and 33 patients(43.4%) got remission over one year after treatment.Splenomegaly(OR=3.14,95%CI=1.01～9.74)and treated with methotrexate(OR=0.22,95%CI=0.07～0.67)were associated with the prognosis from the logistic regression analysis of the 76 cases.The serum ferritin(RR=I.05,95%CI=1.01～1.08)and treated with methotrexate (RR=0.13,95%CI=0.02～0.76)were associated with relapse from the COX regression analysis of the 61 remis- sion cases.Conclusion We need to be very cautious in the follow-up of AOSD patients because some of them may change to other diseases.Methotrexate may be an importent therapy of AOSD not only in improve- ment the prognosis but also in reduction of relapse.