Objective To established a diagnostic methods to idenfity Niemann-Pick disease type C (NPC) in China. Methods Two patients aged 5 and 20 years respectively who presented progressive neurologic regression and splenomegaly were subjected to filipin staining of cultured skin fibroblasts and genetic analyses of NPC1 gene. Results Although there were differences in onset ages and clinical presentations, filipin staining of the cultured skin fibroblasts confirmed the clinical diagnosis of NPC, showing an intense punctate pattern of fluorescence concentrating around the nuclei, consistent with the accumulation of unesterified cholesterol in NPC cells. Genetic sequence analysis further verified the results of filipin staining. The case 1 was compound heterozygous for M1142T(3425T＞C),R1186H(3557T＞C)and case 2 for Q88H(264G＞T),469_470insGT.The latter 2 mutations were novel, and the possibility of polymorphisms was not supposed by analyzing 134 DNA samples obtained form normal controls. Conclusions Filipin stainning of the cultured skin fibroblasts is a reliable method to clinically diagnose NPC with a sensitivity. Genetic diagnose should be performed where genetic analysis is allowed.

In this paper, Liuwei Dihuang pill was used to study the identification of Chinese patent medicine by fluorescence sequencing typing technology. The DNA of Paeonia suffruticosa was used as template to amplify by five pair of FAM fluorescence labeling primers. Then, the amplified products were sequenced. The sequencing results were analyzed by GeneMarker V1.80 to screen the best fluorescence labeling primers. As a result, psbA-trnH fluorescence labeling primer was used to identify the raw materials of Liuwei Dihuang pill. The results showed that three kinds of raw plant medicinal materials in Liuwei Dihuang pill were able to be correctly identified by psbA-trnH fluorescence labeling primer. The fluorescence sequencing typing technology can stably and accurately distinguish raw medicinal materials in Chinese patent medicine.
DNA Primers ; chemistry ; genetics ; DNA, Plant ; chemistry ; genetics ; Drugs, Chinese Herbal ; chemistry ; standards ; Fluorescent Dyes ; chemistry ; Plants, Medicinal ; chemistry ; genetics ; Polymerase Chain Reaction ; instrumentation ; methods ; Quality Control ; Staining and Labeling

Objective To develop an HPLC method for determination of the proportion of tenacissoside A and Ⅰ in Marsdeniatenacissima.Methods Marsdeniatenacissima powder was dissoluted by methanol,treated by ultrasonic,analyzed by HPLC,using Symmetry C1s column(250 mm × 4.6 mm,5 μm).The mobile phase was acetonitrile-0.1％ phosphoric water,gradient elution (0-10 min,32％ A;10-15 min,42％A;15-28 min,50％A).The flow rate was 1 mL · min-1 The detection wavelength was 223 nm.The specificity,standard curve,accuracy,sample recovery,stability and reproducibility of the method were investigated.Results The calibration curves of tenacissoside A were linear in the rangesof 47.2-225.0 μg · mL-1.The average recoveries of three concentrations tenacissoside A were 98.82％,99.24％,99.20％ respectively.The calibration curves of tenacissoside Ⅰ were linear in the ranges of 10.5-50.0 μg · mL-1 The average recoveries of three concentrations tenacissoside Ⅰ were 98.53％,98.32％,98.70％ respectively.The content of tenacissoside A was 5.81-5.96 mg · g-1 The content of tenacissoside Ⅰ was 0.68-0.71 mg · g-1.Conclusion The established method is accurate,simple and reliable,which can be used for the quality concrol of Marsdeniatenacissima.

BACKGROUNDControl of hypersecretion of certain hormones is one of the key targets in the treatment of pituitary adenomas. RNA interference has been shown to inhibit protein expression, and thus it may represent a promising method for the treatment of pituitary adenomas. In the present study, transfection efficiency of small interfering RNA (siRNA) was optimized in human prolactinoma cells.

METHODSFirst, a method was optimized to extract highly purified human prolactinoma cells in vitro. The extracted cells were verified to retain the physiological features of prolactin (PRL) secretion. Second, three conditions for siRNA transfection were tested by the evaluation of transfection efficiency and cell viability. The proper transfection condition was verified for human prolactinoma cells. Third, the siRNA for prolactin was transfected into the human prolactinoma cells, and the suppression of PRL mRNA was evaluated by quantitative real-time reverse transcription-PCR.

RESULTSThe siRNA of 100 pmol with Lipofectamine 2000 of 5 µl for 1 × 10(6) cells was proved preferable, with transfection efficiency being 53.3% and cell viability being 69.7%. In the preliminary experiment the siRNA against PRL decreased the mRNA of PRL by 34.0%.

CONCLUSIONIt is possible to inhibit hormone hypersecretion by RNA interference, that may eventually enable therapeutic siRNA drugs developed.

Adolescent ; Adult ; Cell Line, Tumor ; Cell Separation ; Female ; Humans ; Male ; Middle Aged ; Pituitary Neoplasms ; pathology ; therapy ; Prolactinoma ; pathology ; therapy ; RNA, Small Interfering ; genetics ; Transfection

OBJECTIVETo establish an animal model of obliterative bronchiolitis (OB) after lung transplantation and investigate the pathogenesis preliminarily.

METHODSTracheal segments (5 cartilaginous rings each) were transplanted from SD rats to SD rats (Group I) or to Wistar rats (Group II and III). Grafts were implanted into an abdominal cavity and wrapped in the omentum. Animals in Group I and II did not receive CsA, animals in Group III received CsA daily by gastro-tube at 10 mg.kg(-1).d(-1) from beginning to end. Grafts were harvested on day 3, 14, 28 after transplantation as representative time points for 3 phases of injury in the evolution of allograft airway obliteration, then examined histological changes and gene expression of T-helper 1-and T-helper 2-type cytokines [Th1: interleukin-2 (IL-2), interferon-gamma (IFN-gamma); Th2: interleukin-4 (IL-4), interleukin-10 (IL-10)] in grafts. At the same time, effects of CsA were observed on the above-mentioned indices.

RESULTSThere was no significant difference in histological changes on day 3 after transplantation among 3 groups (P > 0.05). Tracheas in Group I approached to normal morphology on day 14 after transplantation. Airway epithelium of Group II and III almost lost completely on day 14 after transplantation. There was no significant difference between Group II and Group III (P > 0.05), but there were significant differences between Group I and Group II or Group III. The cross-sectional area of the tracheal lumen was narrowed by approximately (5.0 +/- 1.2)%, (28.5 +/- 5.0)% and (19.4 +/- 2.9)% respectively on day 14 after transplantation in Group I, II and III, there were significant differences among 3 groups. On day 14 after transplantation, tracheas in Group I revealed few lymphocytic infiltration, but it showed dense lymphocytic infiltration in Group II. Tracheas in Group III have much more lymphocyte infiltration than that in Group I, but much less than that in Group II. There were significant differences among 3 groups, too (P < 0.01). The tracheal lumen revealed almost total luminal obstruction (94.8 +/- 3.6)% on day 28 after transplantation in Group II. The cross-sectional area of the tracheal lumen was narrowed by approximately (3.7 +/- 0.8)% and (36.6 +/- 7.6)% respectively in Group I and III on day 28. There were significant differences among 3 groups (P < 0.01). Compared with that on day 14, lymphocytic infiltration had decreased gradually on day 28 in Group II and III. There were significant differences among 3 groups all the same (P < 0.01). In Group II, expression of IL-2, IFN-gamma, IL-4, and IL-10 were much higher than that in Group I. Expression of Th1 cytokines was increased to a greater extent than that of Th2 cytokines in Group II compared with Group I. Allografts in Group III expressed significantly less IL-2 gene transcripts than that in Group II over all the points. There was no significant difference between Group II and III in IFN-gamma, IL-4, and IL-10 gene expression.

CONCLUSIONSCompared with isografts, allografts have more obvious changes, such as epithelial damage, fibroproliferation and lymphocytic infiltration. Th1 and Th2 lymphocyte subtypes contribute to the development of obliterative bronchiolitis in heterotopic trachea transplant model of rat, and changes of their cytokines gene expression may be involved in the pathogenesis. CsA could reduce the development of fibroproliferation and lymphocyte infiltration markedly, but it could not protect airway epithelium. CsA inhibits IL-2 gene transcripts, so it can reduce development of the pathologic lesion of obliterative bronchiolitis to a certain degree.

Abdominal Cavity ; surgery ; Animals ; Bronchiolitis Obliterans ; etiology ; pathology ; prevention & control ; Cyclosporine ; pharmacology ; Disease Models, Animal ; Gene Expression ; Immunosuppressive Agents ; pharmacology ; Interferon-gamma ; genetics ; Interleukin-10 ; genetics ; Interleukin-2 ; genetics ; Interleukin-4 ; genetics ; Lung Transplantation ; adverse effects ; methods ; Postoperative Complications ; etiology ; pathology ; prevention & control ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Reverse Transcriptase Polymerase Chain Reaction ; Trachea ; metabolism ; pathology ; transplantation ; Transplantation, Homologous

BACKGROUNDMiddle mediastinal masses comprise a wide variety of tumors but may also reflect lymphadenopathy, and thus remain an interesting diagnostic challenge. We performed positron emission tomography (PET) of mediastinal masses in order to evaluate the ability of PET to predict the malignancy of these tumors. We compared histologic findings, videomediastinoscopy, computed tomography (CT), and PET-CT in patients with mediastinal disease.

METHODSThirty-two patients were evaluated with CT, PET-CT and videomediastionoscopy, and all studies were performed within four weeks in each patient. (11)C-choline as a PET tracer was used to visualize masses. PET data were evaluated using the standardized uptake value (SUV) and were compared with pathologic data.

RESULTSThere were 13 men and 19 women aged from 21 to 74 (mean 45.2) years. Among the patients with mediastinal diseases, sarcoidosis was diagnosed in 12 patients, tuberculosis in 5 patients, lymphoma in 5 patients, and noncaseating granulomata without classical "sarcoid" finding in 3 patients. N2 or N3 nodal metastasis was revealed in 6 of 7 patients who had non-small cell lung cancer or suspected lung cancer, and one was negative (the pathological diagnosis was reactive hyperplasia). The accuracies for correctly diagnosing mediastinal masses for CT, PET-CT and videomediastinoscopy were 38% (12/32), 63% (20/32), and 91% (29/32) respectively. The diagnostic accuracy of videomediastinoscopy was superior to that of PET-CT (chi(2) = 11.130, P < 0.001). The SUVs were similar among these diseases. On the other hand, if the diagnostic classification was benign vs malignancy, the accuracies for CT, PET-CT and videomediastinoscopy were 53% (17/32), 75% (24/32), 100% (32/32) respectively. The diagnostic accuracy of videomediastinoscopy was superior to that of PET-CT (chi(2) = 22.042, P < 0.001). The SUV of malignant lesions (6.9, 3.2 - 9.8; n = 11) appeared to be higher than that of benign lesions (4.9, 2.9 - 8.3; n = 21), however, this difference was not statistically significant (P = 0.054).

CONCLUSIONSTo diagnose lesions located in the middle mediastinum, videomediastinoscopy possesses the highest diagnostic accuracy, and therefore remains the gold standard. PET-CT is valuable for differential diagnosis of benign vs malignant lesions, CT alone or PET alone (SUV) may provide misdiagnosis in a substantial proportion of patients with mediastinal masses.

Adult ; Aged ; Carbon Radioisotopes ; Female ; Humans ; Male ; Mediastinal Diseases ; diagnosis ; Mediastinoscopy ; methods ; Middle Aged ; Positron-Emission Tomography ; Tomography, X-Ray Computed ; Video Recording

OBJECTIVETo study the function of F10 gene, a novel hydaditiform mole-related gene.

METHODSA549 cell line was transfected with the F10 gene of forward or reverse sequence or with the empty vector, respectively. The cellular mRNA was extracted after 24 h of transfection to screen for the differentially expressed genes among the 3 transfected and the control cells using differential display-polymerase chain reaction (ddPCR).

RESULTSThe bands representing differentially expressed genes were amplified from the cells, and the products were linked to T-Vector for sequence analysis. Several genes were screened by Blasting and their expressions were confirmed by fluorescent quantitative PCR.

CONCLUSIONF10 gene is functionally related to cell proliferation and apoptosis.

Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Epithelial Cells ; metabolism ; Female ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Humans ; Hydatidiform Mole ; genetics ; Hydatidiform Mole, Invasive ; genetics ; Lung Neoplasms ; genetics ; pathology ; Oncogenes ; genetics ; Pregnancy ; Transfection ; Uterine Neoplasms ; genetics

OBJECTIVE: To analyze the evaluation and treatment of blood loss during total hip arthroplasty (THA) and total knee arthroplasty (TKA) in the aged . METHODS: We retrospectively surveyed the blood loss and the rehabilitation of 46 cases of TKA and 146 cases of THA older than 60. RESULTS: In the group younger than 70, the mean total blood loss of THA was 1425 mL and the hidden hemorrhage 729 mL (51%); following the TKA, the mean total loss was 1386 mL and the hidden hemorrhage was 890 mL (64%). In the group 70 and older, the mean total blood loss of THA was 1435 mL and the hidden hemorrhage was 769 mL (53%)û following the TKA, the mean total loss was 1380 mL and the hidden hemorrhage was 910 mL (65%). The difference of hidden hemorrhage between the THA and the TKA was both significant by different (p< 0.05). Age played an important part in the THA group (p< 0.05), but not in the TKA group (> 0.05). CONCLUSION The aged have poor resistance to blood loss. Hidden hemorrhage in the TKA or THA perhaps is the primary part of the blood loss. Prompt treatment is helpful for the rehabilitation.
Age Factors ; Aged ; Arthroplasty, Replacement, Hip ; adverse effects ; methods ; Arthroplasty, Replacement, Knee ; adverse effects ; methods ; Blood Loss, Surgical ; statistics & numerical data ; Female ; Hemorrhage ; blood ; etiology ; therapy ; Humans ; Male ; Middle Aged ; Postoperative Complications ; blood ; etiology ; Retrospective Studies

OBJECTIVETo explore the feasibility of creating a 3D-CAD model of craniofacial prostheses through mirror technique to repair the unilateral craniofacial defects and restore craniofacial symmetry.

METHODSPatients with unilateral craniofacial defects underwent spiral CT scanning. CAD3-D image was reconstructed ad 3-D CAD model of craniofacial prosthesis was created with mirror technique, Boolean operation and rapid prototyping technique. Then the prosthesis made of bioactive artificial bone was made through plaster cavity block.

RESULTS15 cases were treated with no complications. Good symmetry was achieved after operation.

CONCLUSIONSDesigning the craniofacial prosthesis with mirror technique guarantees excellent functional and cosmetic results for repairing the unilateral craniofacial defects.

Adolescent ; Adult ; Child ; Computer-Aided Design ; Female ; Humans ; Imaging, Three-Dimensional ; Male ; Middle Aged ; Prosthesis Design ; methods ; Skull ; diagnostic imaging ; pathology ; surgery ; Tomography, Spiral Computed ; Young Adult