Long-term survival of gastric mixed neuroendocrine-non-neuroendocrine neoplasm: Two case reports.

BACKGROUND: Gastric mixed neuroendocrine-non-neuroendocrine neoplasm (MiNEN), which consists of neuroendocrine and non-neuroendocrine components, is quite rare. Until now, most data on gastric MiNEN come from clinical cases, without large-scale retrospective studies or controlled clinical trials. Consequently, no consensus regarding the origin, molecular characteristics, or appropriate treatment of MiNEN has been reached so far. We conducted chemotherapy of irinotecan plus cisplatin (IP regimen) and surgery in two patients with gastric MiNEN, which had never been used in treating this kind of tumor, leading to their long-term survival for more than 3 and 7 years, respectively. CASE SUMMARY: We present two patients (one male and one female) with gastric MiNEN, with the primary manifestation of recurrent upper abdominal pain. After they were referred to our hospital, a diagnosis of gastric MiNEN was defined with the help of CT scan, and histopathological and immunohistochemical examinations on the samples of gastrointestinal endoscopy or radical surgery. The male patient (case 1) were found to have metastases in the reginal lymph nodes and the left liver. He received four cycles of IP regimens first, then the gastrectomy and partial left liver resection, followed by additional two cycles of IP chemotherapy. The female patient (case 2) underwent a laparoscopic gastrectomy, and received six cycles of IP regimen. She was found to have metastatic lesions in the right lung 2 years after that, and underwent video-assisted thoracoscopic surgery (VATS) of the lower lobe of the right lung. The two patients have now survived for more than 3 years and 7 years, respectively, without any evidence of recurrence or metastases. CONCLUSION: IP regimen, combined with curative-intent surgery if feasible, could be considered as the priority in the choice of front-line chemotherapy for gastric MiNEN.

Epstein-Barr virus-associated leiomyosarcoma of the larynx in an adult patient with human immunodeficiency virus infection: Case report and review of the literature.

We investigated the clinical features, treatment, and prognosis of laryngeal leiomyosarcoma (LLMS) and Epstein-Barr virus-associated (EBV-associated) LMS. We report a case of EBV-associated LLMS in an adult patient with HIV infection. We also conducted a review of the English-language literature on LLMS and EBV-associated leiomyosarcoma. To the best of our knowledge, 62 cases of LLMS and EBV-associated leiomyosarcoma have been reported to date. Of patients with LLS, 18.9% had distant metastases and 17.0% had local recurrence. The overall 5-year survival rate was 64.0%. Distant metastases affected the survival of patients with LLMS (p = 0.04). EBV-positive patients had a low survival rate (p = 0.01). Among patients with EBV-associated LMS, 8.2% had distant metastases and recurrence and the overall 5-year survival rate was 50.0%. EBV-associated LLMS is rare. The EBV infection might be a poor prognostic factor of LLMS.

Radiomics Analysis on Multiphase Contrast-Enhanced CT: A Survival Prediction Tool in Patients With Hepatocellular Carcinoma Undergoing Transarterial Chemoembolization.

Patients with HCC receiving TACE have various clinical outcomes. Several prognostic models have been proposed to predict clinical outcomes for patients with hepatocellular carcinomas (HCC) undergoing transarterial chemoembolization (TACE), but establishing an accurate prognostic model remains necessary. We aimed to develop a radiomics signature from pretreatment CT to establish a combined radiomics-clinic (CRC) model to predict survival for these patients. We compared this CRC model to the existing prognostic models in predicting patient survival. This retrospective study included multicenter data from 162 treatment-naïve patients with unresectable HCC undergoing TACE as an initial treatment from January 2007 and March 2017. We randomly allocated patients to a training cohort (n = 108) and a testing cohort (n = 54). Radiomics features were extracted from intra- and peritumoral regions on both the arterial phase and portal venous phase CT images. A radiomics signature (Rad-signature) for survival was constructed using the least absolute shrinkage and selection operator method in the training cohort. We used univariate and multivariate Cox regressions to identify associations between the Rad- signature and clinical factors of survival. From these, a CRC model was developed, validated, and further compared with previously published prognostic models including four-and-seven criteria, six-and-twelve score, hepatoma arterial-embolization prognostic scores, and albumin-bilirubin grade. The CRC model incorporated two variables: The Rad-signature (composed of features extracted from intra- and peritumoral regions on the arterial phase and portal venous phase) and tumor number. The CRC model performed better than the other seven well-recognized prognostic models, with concordance indices of 0.73 [95% confidence interval (CI) 0.68-0.79] and 0.70 [95% CI 0.62-0.82] in the training and testing cohorts, respectively. Among the seven models tested, the six-and-12 score and four-and-seven criteria performed better than the other models, with C-indices of 0.64 [95% CI 0.58-0.70] and 0.65 [95% CI 0.55-0.75] in the testing cohort, respectively. The CT radiomics signature represents an independent biomarker of survival in patients with HCC undergoing TACE, and the CRC model displayed improved predictive performance.

Validation of the 8th edition of AJCC/UICC staging system for nasopharyngeal carcinoma: Results from a non-endemic cohort with 10-year follow-up.

OBJECTIVES: This study aimed to validate the 8th edition of American Joint Committee on Cancer/Union for International Cancer Control (AJCC/UICC) TNM staging system for nasopharyngeal carcinoma (NPC) in non-endemic region. MATERIALS AND METHODS: We recruited 607 patients with histology-proven, previously untreated, non-metastatic NPC treated by intensity-modulated radiotherapy (IMRT) at our center. Harrell's concordance index (c-index) and Akaike information criterion (AIC) were applied to compare the prognostic discrimination between the 7th and 8th edition staging system. RESULTS: For T category, the local recurrence-free survival (LRFS) Kaplan-Meier curves of T1, T2 and T3 were well separated in the 8th edition; however, LRFS did not significantly differ between T3 and T4 (P = 0.166). Moreover, the 7th edition achieved higher c-index (0.702 [95% CI, 0.618-0.787] vs. 0.685 [95% CI, 0.604-0.767]) and lower AIC (766.1 vs. 770.8) than 8th edition for LRFS. With regard to N category, the 8th edition achieved higher c-index (0.796 [95% CI, 0.749-0.843] vs. 0.751 [95% CI, 0.696-0.805]) and lower AIC (1439.4 vs. 1471.9) for distant metastasis-free survival. In terms of overall stage, the 8th edition also had higher c-index (0.798 [95% CI, 0.753-0.844] vs. 0.721 [95% CI, 0.672-0.770]) and lower AIC (1963.9 vs. 2007.2) compared with the 7th edition for overall survival. Furthermore, interval validation by bootstrapping the sample randomly for ~100-1000 times also validated above findings. CONCLUSION: The 8th edition of AJCC/UICC TNM staging system achieved significantly better prognostic discrimination than the 7th edition with regard to N category and overall stage but not T category.

Diffusion-weighted imaging volume and diffusion-weighted imaging volume growth in acute stroke: associations with fluid-attenuated inversion recovery hyperintensities-diffusion-weighted imaging mismatch and functional outcome.

PURPOSE: This study assessed the association between diffusion-weighted imaging (DWI) volume and fluid-attenuated inversion recovery vascular hyperintensity (FVH)-DWI mismatch, functional outcome in patients with acute stroke patients receiving endovascular therapy, as well as the value of DWI volume in predicting functional outcome with stroke patients. METHODS: In 38 stroke patients who received endovascular therapy, FVH-DWI mismatch, DWI volume on admission, DWI volume on follow-up, DWI volume growth, the functional outcome at 3 months [modified Rankin scale (mRS)], and other clinical data were collected. Statistical analysis was performed to compare the associations with the above variables and predict functional outcome after stroke. RESULTS: Compared with no FVH-DWI mismatch group (n = 15), FVH-DWI mismatch group (n = 23) had a smaller DWI volume on admission (t = -2.980; P = 0.008), smaller DWI volume on follow-up (t = -2.911; P = 0.009), lower DWI volume growth (t = -2.328; P = 0.031). The 3-month outcome (1.87 ± 0.92) in patients with FVH-DWI mismatch was better than that (2.93 ± 1.62) of patients with no FVH-DWI mismatch (t = -2.307; P = 0.032). Spearman's rank correlation analysis revealed that FVH-DWI mismatch (r = 0.327; P = 0.045), DWI volume on admission (r = 0.414; P = 0.010), DWI volume on follow-up (r = 0.486; P = 0.002), and DWI volume growth (r = 0.467; P = 0.003) were positively correlated with mRS at 3 months. ROC analysis showed when the optimal cutoff value of DWI volume on admission was 33.50, the sensitivity and specificity for predicting functional outcome was 60 and 95.65%, respectively. CONCLUSIONS: Evaluating DWI volume on admission, DWI volume on follow-up as well as DWI volume growth comprehensively may be useful in predicting the functional outcome of acute stroke patients after thrombectomy.

Effects of hemoglobin variants HbJ Bangkok, HbE, HbG Taipei, and HbH on analysis of glycated hemoglobin via ion-exchange high-performance liquid chromatography.

BACKGROUND: To explore the effects of HbJ Bangkok, HbE, HbG Taipei, and α-thalassemia HbH on the results of HbA1c assessment using ion-exchange high-performance liquid chromatography (IE-HPLC). METHODS: We enrolled five patients in which the results of the IE-HPLC HbA1c assay were inconsistent with the average levels of FBG. We performed hemoglobin capillary (Hb) electrophoresis using whole-blood samples. We also sequenced the genes encoding Hb using dideoxy-mediated chain termination and analyzed HbA1c using borate affinity HPLC (BA-HPLC) and turbidimetric inhibition immunoassay (TINIA). RESULTS: Two patients had the HbJ Bangkok variant. Hb genotypes of these patients were ß41-42 /ßJ Bangkok and ßN /ßJ Bangkok , and the content of HbJ Bangkok was 93.9% and 52.4%, respectively. The remaining three patients had the following: HbE (ßN /ßE Hb genotype, 23.6% HbE content), HbG Taipei (ßN /ßG Taipei Hb genotype, 39.4% HbG Taipei content), and α-thalassemia HbH (6.1% HbH content, 2.8% Hb Bart's content). In the patients with ß-thalassemia and HbJ Bangkok variants, the presence of the variants interfered with the results of HbA1c analyses using IE-HPLC and TINIA; in the remaining four patients, there was interference with the results of HbA1c IE-HPLC but not with the TINIA assay. There was no interference with BA-HPLC HbA1c results. CONCLUSIONS: HbJ Bangkok, HbE, HbG Taipei Hb, and α-thalassemia HbH disease cause varying degrees of interference with the analysis of HbA1c using IE-HPLC. In these patients, we suggest using methods free from such interference for the analysis of HbA1c and other indicators to monitor blood glucose levels.

Evaluation of the Interference of Hemoglobin Variant J-Bangkok on Glycated Hemoglobin (HbA1c) Measurement by Five Different Methods.

Objective The interference of the hemoglobin variant (Hb J-Bangkok) was evaluated on 4 different glycated hemoglobin assays and compared with a reference immuno assay. Methods An overall test of coincidence of 2 least-squares linear regression lines was performed to determine whether the presence of Hb J-Bangkok caused a statistically significant difference in HbA1c results compared with a reference immuno assay. Statistical analysis was performed on the difference of the estimated average glucose calculated from HbA1c values and fasting plasma glucose in the Hb J-Bangkok variant group using the different detection systems. Deming regression analysis was used to determinate whether Hb J-Bangkok had a significant interference on HbA1c results using an HbA1c±10% relative bias at 6% and 9% HbA1c as evaluation limits. Results Turbidimetric inhibition immunoassay method, and enzymatic methods were not affected by Hb J-Bangkok. However, Hb J-Bangkok showed statistically significant interference to the two ion-exchange high-performance liquid chromatography methods. Conclusion When performing HbA1c tests, clinical laboratory personnel should identify the Hb variant and select the appropriate methods or use alternative indicators.

Percutaneous liver biopsy: retrospective study of primary and secondary hepatic lymphoma in twenty-one patients.

BACKGROUND: Hepatic lymphoma (HL) is categorized as primary and secondary hepatic lymphoma (PHL and SHL). This disorder can present as hepatic mass or mass-like lesion. Chemotherapy often is the first line treatment for patients with HL. Thus, an accurate pre-management histological diagnosis is essential to potentially improve clinical outcomes. The present study was to explore the prevalence of HL in ultrasound guided liver biopsies for hepatic mass or mass-like lesions, to investigate HL associated clinicopathological features, to raise the awareness of early recognition and proper diagnosis of this entity, and to assess specimen adequacy in needle core biopsy. METHODS: Twenty-one cases of HL were enrolled. Clinical and pathological characteristics were evaluated, quality of biopsies was assessed and pertinent literature was reviewed. RESULTS: HL was diagnosed in 0.94% of 2242 liver biopsy cases with ambiguous clinical presentation, laboratory tests and image studies. There were two cases of PHL (0.09%), and nineteen cases of SHL (0.85%). Histopathologically, diffuse large B-cell lymphoma was the most common type, followed by B-cell lymphoma not otherwise specified, T-cell lymphoma, Hodgkin's lymphoma, and B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma. Additionally, three lymphocytic infiltration patterns were documented microscopically. The nodular infiltration was the most common type. CONCLUSIONS: HL is a rare entity and histopathology along with ancillary tests remains the only way to make the diagnosis. Clinicians' awareness of this entity and early liver biopsy are essential in patient management.

[Effect of Inhibiting and Activating Wnt Signalling Pathway on NSC67657-inducing Monocytic Differentiation of HL-60 Cells].

OBJECTIVE: To investigate the effect of inhibiting and activating Wnt signalling pathway on monocyte differentiation of HL-60 cells induced with a new steroidal drug NSC67657 and its possible mechamism. METHODS: The HL-60 cells were treated with 5, 10 and 20 µmol/L XAV-939 (inhibitor of Wnt signalling pathway) for 3 days, and with 10, 20 and 30 mmol/L LiCl (activator of Wnt signalling pathway) for 1 day; the expression levels of down-stream genes and proteins of Wnt signolling pathway were detected by RT-PCR and Western blot, respectively; the expression of cell surface differentiation antigen CD14 and early apoptosis of HL-60 cells was detected by flow cytometry, moreover the most suitable concentration of Wnt inhibitor and activator for HL-60 cells was determined. Then the HL-60 cells with inhibited and activated Wnt pathway were treated with NSC67657 of 10 µmol/L for 3 days; the expression levels of CD14 and down-stream target proteins of Wnt signalling pathway in blank control (culture mediam) group, simple NSC67657-treated group, NSC67657 combined with inhibitor group and NSC67657 combined activator group were compared and analyzed. RESULTS: 20 µmol/L XAV-939 and 20 mmol/L LiCl could effectively inhibit and activate Wnt signalling pathway of HL-60 cells respectively, could significantly down- and up-regulate the expression of cyclinD1, TCF1 and c-Jun genes (P < 0.05) and proteins (P < 0.05); moreover, the number of CD10(+) HL-60 cells in these conditions was below 1%, no early apoptosis of HL-60 cells was found. In the simple NSC67657-treated groups, the expression of cyclinD1, TCF1 and c-Jun proteins was down-regulated (P < 0.05), and the percentage of CD14(+) HL-60 cells accounted for 62.13 ± 9.44; after the HL-60 cells were treated with XAV-939, the NSC67657 could more significantly down-regulate the expression of cyclinD1, TCF1 and c-Jun proteins and the percentage of CD14(+) HL-60 cell accounted for 84.17 ± 5.39%, as compared with simple NSC67657-treated group; as compared with blank controls group, the expression of cyclinD1, TCF1 and c-Jun proteins was more obviously down-regulated and the percentage of CD14(+) HL-60 cells decreased to 33.99 ± 8.37% in NSC67657 combined LiC1 streated group, but which were higher than those in simple NSC67657-treated group (P < 0.05). CONCLUSION: 20 µmol/L XAV-939 and 20 mmol/L LiCl as effective inhabitor and activator of Wnt signalling pathway respectively can significantly down- and up-regulate the expression of Wnt down-stream pathway target genes and proteins. The influence of XAV-939 and LiC1 on differentiation of HL-60 cells induced by NSC67657 suggests that Wnt signalling pathway plays a key role in monocyte differentiction of HL-60 cells induced by NSC67657.

Unusual synchronous skeletal muscle and lung metastasis in papillary thyroid cancer: A case report and review of the literature.

Papillary thyroid cancer (PTC) frequently metastasizes to the cervical lymph region and less often to the lung and bone. Metastasis to the skeletal muscles from PTC is extremely rare, especially concurrent lung and skeletal muscle metastases. The present study reports the case of a 31-year-old man with synchronous metastasis to the skeletal muscle and lung from PTC, six years following total thyroidectomy and consecutive 131Iodine treatments. Magnetic resonance imaging (MRI) revealed a 1.7×1.2×1.5 cm mass in the left gastrocnemius muscle, indicating a neurogenic tumor. The mass was subsequently resected and confirmed via histopathology to be metastatic PTC. We propose that, in the follow-up of patients with PTC, the measurable serum thyroglobulin level, whole body scan and other imaging modalities including MRI or positron emission tomography/computed tomography, must be closely monitored for potential distant metastases, particularly in cases of PTC with aggressive pathological behavior.

[Association of polymorphisms in signal transducer and activator of transcription 4 gene and the susceptibility to unexplained recurrent spontaneous abortions].

OBJECTIVE: To investigate the association between the polymorphisms of signal transducer and activator of transcription 4 (STAT4) gene and the susceptibility to unexplained recurrent spontaneous abortion(URSA). METHODS: PCR-restriction fragment length polymorphism (PCR-RFLP) was used to detect genotype 3 loca (rs7574865 G/T, rs10181656 C/G and rs16833431 C/T) polymorphism of STAT4 in 246 URSA cases (URSA group) and 183 normal controls (control group) . RESULTS: (1)The frequencies of rs7574865 were genotype G/G of 36.2% (89/246) in URSA group and 46.4% (85/183) in control group, genotype G/T of 47.2% (116/246) in URSA group and 45.4% (83/183) in control group, and genotype T/T of 16.7% (41/246) in URSA group and 8.2% (15/183) in control group, which reached statistical difference (P < 0.05). The frequencies of rs10181656 were genotype CC of 36.6% (90/246) in URSA group and 46.4% (85/183) in control group, genotype C/G of 48.0% (118/246) in URSA group and 44.8% (82/183) in control group, and genotype G/G of 15.4% (38/246) in URSA group and 8.7% (16/183) in control group, which reached statistical difference (P < 0.05). The carriers of rs7574865 T allele and rs10181656 G allele increased the risk of URSA (OR = 1.51, 1.44, all P < 0.05).(2) There was no different distribution in 3 genotypes (C/C, C/T, T/T) and 2 alleles (C and T) of rs16833431 C/T between URSA patients and normal controls (P = 0.43,0.48). (3) Timated haplotype frequency distribution of rs7574865 G/T and rs10181656 C/G showed haplotype G-T conferring the susceptibility to URSA (OR = 1.49, P < 0.01), but haplotype C-G could provide protection on URSA (OR = 0.68, P < 0.01). CONCLUSION: Polymorphisms of STAT4 gene might confer the susceptibility to URSA by altering STAT4 function and (or) its expression.

[Expressing trend of NME3 protein in acute myeloid leukemia HL-60 cells and patients' bone marrow].

To verify the differential expression of non-metastasis cell 3 (NME3) protein in HL-60 cells when they were induced to differentiate into monocyte and granulocyte like cells, and study its value in diagnosis of acute myeloid leukemia, all-trans retinoic acid (ATRA) and a new steroidal drug NSC67657 were employed to induce acute myeloid leukemia HL-60 cells into monocyte and granulocyte like cells. Then the cell differentiating direction was observed by chemical staining, the degree of differentiation was determined by surface antigen CD11b/CD14 detection, and the apoptosis was excluded by phosphatidylserine valgus analysis, by which cellular differentiating model was constructed. Furthermore, RT-PCR and Western blot were employed to verify the differentially expression of NME3 before and after differentiation of HL-60 cells. At last, samples from bone marrow nucleated cells of 26 patients with myeloid leukemia, which were diagnosed definitely by clinical doctors, and 5 normal people were chosen. Then the expressing trend of NME3 protein in these testing groups was analyzed by means of comparison. The results showed that ATRA (2 µmol/L for 5 d) and NSC67657 (10 µmol/L for 5 d) could induce HL-60 cells to differentiate into monocyte and granulocyte like cells above 90% without cell apoptosis. The expression of NME3 gene and protein were down-regulated by the inducers, which was accorded with the screening results that was got using proteomics technology in the former research. The expression of NME3 protein in bone marrow from acute myeloid leukemia patients was elevated significantly as compared to normal persons. It is concluded that the expression level of NME3 protein is down-regulated after cellular differentiation, according with the changing trend in leukemia patients, which imply that NME3 protein may be a potential biomarker for diagnosis of acute myeloid leukemia.

Generation and neuronal differentiation of induced pluripotent stem cells in Cdyl-/- mice.

Chromodomain on Y-like (CDYL) is a chromodomain protein that has sequence homology to members of the enoyl CoA hydratase family. Although the chromodomain of CDYL has been implicated in chromatin remodeling during mammalian spermatogenesis, the function of the Cdyl gene remains unclear. Recently, induced pluripotent stem cells (iPS cells) have been derived from somatic cells by the forced expression of several transcription factors. iPS cells resemble embryonic stem cells in many respects. Therefore, iPS cells represent a powerful tool for the study of gene function. In this study, we have investigated whether iPS cells derived from Cdyl-/- and Cdyl+/+ fibroblasts have different characteristics. Our results showed that both Cdyl-/- and Cdyl+/+ fibroblasts could be induced to become iPS cells, but the spontaneous neuronal differentiation capacity of Cdyl-/- iPS cells was much greater than that of the Cdyl+/+ iPS cells. These results provide some insight into the molecular function of the Cdyl gene, showing that it inhibited the neuronal differentiation of iPS cells.

[Amino-terminal pro-brain natriuretic peptide and brain natriuretic peptide measurements under various detection conditions in patients with chronic heart failure].

OBJECTIVE: To find the potential interference factors for the detection of NT-proBNP and BNP in patients with chronic heart failure. METHODS: EP15-A2 issued by Clinical and Laboratory Standards Institute (CLSI) was employed to compare the precision and accuracy of commercial NT-proBNP and BNP analyzer electrochemiluminescence immunoassay system Cobas E601 and chemiluminescence system ADVIA Centaur. Moreover, NT-proBNP and BNP were detected in different time interval and in different interfered sampling conditions (haematolysis, choloplania, lipemia). NT-proBNP and BNP of 203 patients with heart failure or heart failure complicated with acute cerebral infarction were analyzed to find the deviation caused by patients' endogenous factors. RESULTS: The precision and accuracy were comparable for NT-proBNP and BNP detection using Cobas E601 and ADVIA Centaur (total-CV below 2.9% and 3.5%, the deviation from definite value below 2.38% and 3.91%). The most suitable sample type for NT-proBNP and BNP detection was serum and EDTA-anticoagulant plasma. The detection results of NT-proBNP and BNP were comparable for at least 120 min post sampling and not affected by Hb (2 g/L), DB (428 µmol/L) and chyle (2000 FIU). NT-proBNP was significantly higher in heart failure patients complicated with cerebral infarction (P = 0.003) than in heart failure patients. BNP was significantly higher in heart failure grade III patients complicated with cerebral infarction (P < 0.01). CONCLUSIONS: Cobas E601 and ADVIA Centaur supplied satisfactory detection of NT-proBNP and BNP in patients with chronic heart failure with strong anti-interference capacity. The diagnostic value of NT-proBNP and BNP for chronic heart failure should be analyzed objectively in the presence of complicating diseases.

Shape dependence of gold nanoparticles on in vivo acute toxicological effects and biodistribution.

The toxicity and biodistribution in vivo of various morphologies of Au nanoparticles (AuNPs) were studied by using KM mice. The quantitative analysis of Au in each tissue of mice was done by using the Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Sphere-shaped AuNPs displayed the best biocompatibility, compared with rod- and cube-shaped of AuNPs, and rod-shaped AuNPs was more toxic than cube-shaped AuNPs. In vivo biodistribution study revealed all AuNPs were preferentially accumulated in organ of liver and spleen. The findings from this study thus revealed that the toxicity and biodistribution in vivo of AuNPs are shape dependent.

[Study on association of functional polymorphisms in Foxp3 gene with the susceptibility to unexplained recurrent spontaneous abortion].

OBJECTIVE: To investigate the association between the functional polymorphisms of Foxp3 gene and unexplained recurrent spontaneous abortion (URSA). METHODS: PCR-restriction fragment length polymorphism (rs3761548, rs2294021) and PCR with sequence-specific primers (rs2232365, rs5902434) were used to detect four polymorphisms of Foxp3 in 146 URSA cases and 112 normal controls. RESULTS: (1) The frequencies of rs3761548A/C were 10.3%, 22.3% in genotype C/C, 38.4%, 40.2% in genotype A/C and 51.4%, 37.5% in genotype A/A between URSA patients and normal controls; the frequencies of rs2232365A/G were 5.5%, 15.2% in genotype A/A, 47.9%, 50.0% in genotype A/G, 46.6%, 34.8% in genotype G/G between URSA patients and normal controls; they all reached statistical difference (P < 0.05). The carriers of rs3761548A allele and rs2232365G allele increased the risk of URSA (OR = 1.73, 1.61;all P < 0.05). (2) There was no difference in the genotypic distribution of rs5902434del/ATT polymorphism between cases and controls (P = 0.10), but the frequency of del allele in URSA was statistically increased than that of controls (71.2%, 62.5%;OR = 1.49, P = 0.04). (3) There was no different distribution in 3 genotypes (C/C, T/C, T/T) and 2 alleles (T and C) of rs2294021T/C between URSA patients and normal controls (P = 0.18 and 0.08). (4) Estimated haplotype frequency distribution of rs5902434del/ATT, rs3761548A/C and rs22323565A/G showed haplotype del-A-G conferring the susceptibility to URSA (OR = 2.51, P < 0.01) but haplotype del-C-G and ATT-A-A could provide protection on URSA (OR = 0.18, 0.22; all P < 0.01). CONCLUSION: Functional polymorphisms of Foxp3 gene could probably confer the susceptibility to URSA, by altering Foxp3 function and (or) its expression.

[Study on the mechanism of THP-1 cell differentiation imduced by a new steroidal drug NSC67657].

OBJECTIVE: To study the potential mechanism of the new steroidal drug NSC67657 induced leukemic cells differentiation. METHODS: Cell proliferation was assayed by MTT assay. Surface antigen CD14 on THP-1 cells treated by NSC67657 at different time different concentration, was detected by flow cytometry (FCM). The expression of beta-catenin- interacting protein 1 (ICAT) gene and protein were detected by RT-PCR and Western blot. Eukaryotic expressing vector pDsRed-ICAT was constructed and transfected into HL60 cell line. FCM, Wright's staining and electronmicroscope were employed to analyse the differentiation of transfected THP-1 cells after they were treated with NSC67657 for 24 hours. RESULTS: The proliferation of THP-1 cells was significantly inhibited by NSC67657 treatment. The level of CD14 expression was elevated in line with the increasing drug concentration and treatment time. 10 µmol/L NSC67657 treatment for five days was the optimal condition for the induction of THP-1 cells differentiation, when the CD14(+) THP-1 cells were more than 90%. Morphological study indentified the THP-1 cells of monocytic differentiation. The eukaryotic expressing vector pDSRed-ICAT was successfully constructed, and almost 90% positive clone could be obtained after G418 screening. Electro-transfection was employed for transfecting the vector into THP-1 cells. After the transfection the expression of ICAT gene and protein was increased. On the NSC67657 treatment, there was not significant difference in CD14 expression on transfected THP-1 cells compared to that on the control groups. After 24 h treatment, the transfected THP-1 cells remained in early differentiated stage. CONCLUSION: NSC67657 can induce THP-1 cell to monocytic differentiation and activate the expression of ICAT gene, but overexpression of ICAT itself is not sufficient to induce such differentiation.

A stem cell-based tool for small molecule screening in adipogenesis.

Techniques for small molecule screening are widely used in biological mechanism study and drug discovery. Here, we reported a novel adipocyte differentiation assay for small molecule selection, based on human mesenchymal stem cells (hMSCs) transduced with fluorescence reporter gene driven by adipogenic specific promoter--adipocyte Protein 2 (aP2; also namely Fatty Acid Binding Protein 4, FABP4). During normal adipogenic induction as well as adipogenic inhibition by Ly294002, we confirmed that the intensity of green fluorescence protein corresponded well to the expression level of aP2 gene. Furthermore, this variation of green fluorescence protein intensity can be read simply through fluorescence spectrophotometer. By testing another two small molecules in adipogenesis--Troglitazone and CHIR99021, we proved that this is a simple and sensitive method, which could be applied in adipocyte biology, drug discovery and toxicological study in the future.

[Effect of C/EBPalpha on the monocytic differentiation of HL60 cells induced by NSC67657].

OBJECTIVE: To figure out the function of C/EBPalpha in the monocytic differentiation of HL60 cells induced by a new steroidal drug NSC67657. METHODS: The differentiation of HL60 cells was induced by NSC67657, and the cell surface antigen CD14 expression was detected by flow cytometry. The gene and protein expressions of CCAAT enhancer binding protein alpha (C/EBPalpha) before and after the induction of cell differentiation were determined by RT-PCR and Western blot. Eukaryotic expressing vector pDsRed-ICAT was constructed and transfected into HL60 cells, and its expression was verified. The effect of C/EBPalpha overexpression in HL60 cells was assessed by MTT assay, Wright's staining and flow cytometry before and after NSC67657 transfection. RESULTS: HL60 cells could be induced into monocytes by 10 micromol/L ATRA within 5 days, and the coverage of CD14 positive cells reached 93.9% after 5 days of drug treatment. The eukaryotic expressing vector was successfully constructed, and over 90% positive clones were obtained after screening by G418 and electrotransfection. The results of proliferative analysis, chemical staining, ultrastructural observation, and CD11b detection confirmed that HL60 cells could be induced into granulocytic differentiation by overexpression of C/EBPalpha protein. Moreover, in the drug treatment group, transfected cells could not be induced into monocytic differentiation, and their granulocytic differentiation was also inhibited. CONCLUSION: The monocytic differentiation of HL60 cells induced by NSC67657 may not be via the regulation by C/EBPalpha protein-mediated signal transduction. However, the overexpression of CEBPalpha may inhibit the process of NSC67657-induced monocytic differentiation in HL60 cells.

[Effects of herba ephedrae, honey-fried herba ephedrae and maxingshigan decoction on autonomic activities of mice].

OBJECTIVE: To discuss the significance of processing and complex prescription with Herba Ephedrae, the effects of Herba Ephedrae, Honey-fried Herba Ephedrae and Maxingshigan decoction on autonomic activities in mice were compared. METHODS: 110 female Kunming mice were divided into 11 groups, namely normal saline group (NS), ephedrine group (E), high dose Herba Ephedrae group (MH-H), moderate dose Herba Ephedrae group (MH-M), low dose Herba Ephedrae group (MH-L) ,high dose Honey-fried Herba Ephedrae group (ZMH-H), moderate dose Honey-fried Herba Ephedrae group (ZMH-M),low dose Honey-fried Herba Ephedrae group (ZMH-L), high dose Maxingshigan decoction group (MX-H), moderate dose Maxingshigan decoction group (MX-M) and low dose Maxingshigan decoction group (MX-L). The numbers of autonomic activity in 15 minutes before intragastric administration (ig), and after ig 0.5, 1, 2, 3, 4 h were determined. RESULTS: There was interaction between time and groups. There were very significant differences between before ig and all time-points after ig (P < 0.01). 30 minutes after ig,there were significant differences between E and NS,E and ZMH-L,E and MX-L (P < 0.05). 1 h after ig, there were significant differences between MX-M and NS (P < 0.05). 3 h after ig, there were significant differences between MX-M and MH-L (P < 0.05). 30 minutes after ig, both ZMH-L and MX-L could reduce the number of autonomic activity in some extent compare with MH-L. 1 h or 2 h after ig, ZMH-L could reduce the number of autonomic activity in some extent compare with MH-L 4 h after ig, MX-L could reduce the number of autonomic activity in some extent compare with MH-L. CONCLUSION: Under the condition of this study, regarding autonomic activity as index, both ZMH-L and MX-L can reduce center stimulation in some extent.