Proliferation, Differentiation and Immunoregulatory Capacities of Brown and White Adipose-Derived Stem Cells from Young and Aged Mice

Background and Objectives@#Adipose tissue is a source of mesenchymal stem cells, which have the potential to differentiate into various types of cells. Adipose-derived stem cells (ADSCs) are now recognized as an accessible, abundant, and reliable stem cells suitable for tissue engineering and regenerative medicine applications. However, few literatures gave a comprehensive report on the capacities of ADSCs harvested from different sites. Especially, the capacities of ADSCs from aged mice remained unclear. In this study, we investigated several main capacities of brown adipose derived stem cells (B-ADSCs) and white adipose derived stem cells (W-ADSCs) from both young and aged mice. @*Methods@#and Results: When isolated from young mice, B-ADSCs showed a stronger proliferation rate and higher osteogenic, adipogenic and myocardial differentiation ability than W-ADSCs. Carboxy fluorescein diacetate succinimidyl ester (CFSE) labeling test suggested no significant difference in immunosuppression capacity between B-ADSCs and W-ADSCs. Similarly, no difference between these two were found in several immune related molecules, such as programmed death-ligand 1 (PD-L1), intercellular cell adhesion molecule (ICAM-1), vascular cell adhesion molecule (VCAM-1), inducible nitric oxide synthase (iNOS), tumour necrosis factor-α (TNF-α), interleukin 10 (IL10), and suppressor of cytokine signaling 1 (socs1). When isolated from aged mice, B-ADSCs also showed a stronger proliferation rate and higher osteogenic, adipogenic and myocardial differentiation ability than W-ADSCs; however, it demonstrated an attenuated immunosuppression capacity compared to W-ADSCs. @*Conclusions@#In summary, our data showed that ADSCs’ characteristics were tissue source dependent and changed with age. It provided evidence for choosing the right tissue-specific ADSCs for clinical application and fundamental research.

Comprehensive genetic diagnosis of patients with Duchenne/Becker muscular dystrophy (DMD/BMD) and pathogenicity analysis of splice site variants in the DMD gene / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are caused by mutations in the DMD gene. The aim of this study is to identify pathogenic DMD variants in probands and reduce the risk of recurrence of the disease in affected families. Variations in 100 unrelated DMD/BMD patients were detected by multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS). Pathogenic variants in DMD were successfully identified in all cases, and 11 of them were novel. The most common mutations were intragenic deletions (69%), with two hotspots located in the 5′ end (exons 2–19) and the central of the DMD gene (exons 45–55), while point mutations were observed in 22% patients. Further, c.1149+1G>A and c.1150-2A>G were confirmed by hybrid minigene splicing assay (HMSA). This two splice site mutations would lead to two aberrant DMD isoforms which give rise to severely truncated protein. Therefore, the clinical use of MLPA, NGS, and HMSA is an effective strategy to identify variants. Importantly, eight embryos were terminated pregnancies according to prenatal diagnosis and a healthy boy was successfully delivered by preimplantation genetic diagnosis (PGD). Early and accurate genetic diagnosis is essential for prenatal diagnosis/PGD to reduce the risk of recurrence of DMD in affected families.

Comprehensive genetic diagnosis of patients with Duchenne/Becker muscular dystrophy (DMD/BMD) and pathogenicity analysis of splice site variants in the DMD gene / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are caused by mutations in the DMD gene. The aim of this study is to identify pathogenic DMD variants in probands and reduce the risk of recurrence of the disease in affected families. Variations in 100 unrelated DMD/BMD patients were detected by multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS). Pathogenic variants in DMD were successfully identified in all cases, and 11 of them were novel. The most common mutations were intragenic deletions (69%), with two hotspots located in the 5' end (exons 2-19) and the central of the DMD gene (exons 45-55), while point mutations were observed in 22% patients. Further, c.1149+1G>A and c.1150-2A>G were confirmed by hybrid minigene splicing assay (HMSA). This two splice site mutations would lead to two aberrant DMD isoforms which give rise to severely truncated protein. Therefore, the clinical use of MLPA, NGS, and HMSA is an effective strategy to identify variants. Importantly, eight embryos were terminated pregnancies according to prenatal diagnosis and a healthy boy was successfully delivered by preimplantation genetic diagnosis (PGD). Early and accurate genetic diagnosis is essential for prenatal diagnosis/PGD to reduce the risk of recurrence of DMD in affected families.

Optimization of preparation of curcumin-loaded TPGS/F127/P123 mixed micelles by uniform design methodology / 中草药

Objective: The preparation process of curcumin-loaded TPGS/F127/P123 mixed micelles was optimized with uniform design method to improve the poor solubility of curcumin in water, aiming to increase entrapment efficiency (EE), drug-loading (DL), and reduce the precipitated drug (PD). Methods: Curcumin-loaded TPGS/F127/P123 mixed micelles were prepared by thin-film hydration method with modification. Before using the uniform design, a number of preliminary experiments were conducted to identify the controlled factors such as the amount of curcumin, the dosage of TPGS, and the ratio of F127/P123. The formulation was operated by uniform design of three factors and seven levels, and its results were fitted by polynomial linear equation, the response surface, and the contour line in order to choose and verify the optimal preparation process. Results: In the optimum formulation, the dosage of curcumin was 14.0 mg, TPGS 150.0 mg, and the ratio of F127/P123 was 68: 32. The solubility of optimum formulation was (3.47 ± 0.14) mg/mL, EE (87.15 ± 4.39)%, DL (4.70 ± 0.17)%, and PD (0.33 ± 0.12)% in 48 h. Conclusion: The solubility of curcumin in TPGS/F127/P123 mixed micelles was improved after the optimization of the uniform design method, and EE and DL were also improved.

Study on the effect of inspiratory muscle training of heavy load of deep breathing trainer for exercise tolerance and breathing difficulty level of patient with COPD in stable stage / 中国医学装备

Objective: To explore the effect of adopting inspiratory muscle training of heavy load of deep breathing trainer for exercise tolerance and breathing difficulty level of patient with chronic obstructive pulmonary disease COPD in stable stage.Methods: 108 patients with COPD in stable stage were divided into observation group(54 cases)and control group(54 cases).Patients of control group received resistance training of inspiratory muscle of low load(9 cm H2O)of deep breathing trainer.And that of observation group received inspiratory muscle training of heavy load(the maximum inspiratory pressure was 60％)of deep breathing trainer,and this training was arranged at morning and evening(the time of each training was 15 min)of every day,and the training included 6 times in each week,and the total training time was 6 months.And then the exercise tolerance(6 min walk distance,6MWD),the step number of pedometer,the score of breathing scale of modified medical British research council(mMRC)and lung function were compared.Results: The 6MWD and step number of pedometer of observation group of 1 month,3 months and 6 months post training were significantly higher than that of control group(t=12.365,t=13.254,t=12.845,P＜0.05),respectively.And the score of dyspnea of mMRC of observation group of 1 month,3 months and 6 months post training were significantly better than that of control group(t=4.365,t=4.021,t=4.325,P＜0.05),respectively.And the forced expiratory volume in one second(FEV1),FEV1％and the ratio of FEV1 and FVC％of lung function of 1 month,3 months and 6 months post training were significantly better than that pre training(F=3.265,F=2.985,F=2.963,P＜0.05).And the differences of the three indicators between observation group and control group were significant(F=3.474,F=3.146,F=3.271,P＜0.05).Conclusion: For patients with COPD in stable stage,the inspiratory muscle training of heavy load of deep breathing trainer can enhance exercise tolerance of patient,and relieve their symptom of dyspnea and improve lung function of patients.

Clinical Efficacy of Autologous Transplantation of Hematopoietic Stem Cells Cryopreserved by Ladder-style Freezing from Low Temperature Refrigerator to Liquid Nitrogen / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the efficacy of hematopoietic stem cells cryopreserved by ladder-style freezing from low temperature refrigerator to liquid nitrogen in treatment of hematological malignancies, and to analyze the survival condition of patients after hematopoietic stem cell transplantation.</p><p><b>METHODS</b>The coyoprotectant formed by 3% hydroxyethyl starch, 4% albumin and 5% dimethyl sulfoxide (DMSO) was need for cryopreservation of hematopoietic stem cells,which were first placed in -80C low temperature refrigerator and then were stored in -196C liquid nitrogen tank. 98 cases of hemafologic malignancies (io cases of ALL, 24 cases of AML, L-cases of MM and 53 case of malignant lymphoma) were selected from January 2002 to December 2016, and recived transplantatin auto-hematopoiehc stem cells cryopresorved by above-mentined method. The overall survival rate (OS), progression-free survival (PFS) were analyzed statistically.</p><p><b>RESULTS</b>One case failed in implantation due to intracranial hemorrhage and the other 97 cases all succeeded in hematopoietic reconstitution. The average time needed for neutrophil count ≥0.5×10/L was 9.24±1.89 d, and the average time needed for blood platelet ≥20×10/L without platelet transfusion for 3 days was 11.04±1.84 d. The median survival time was 47.6 months (1-80 months). The 3 and 5 year OS rates were (97.2±1.9) %, (84.2±4.6) % and (77.8±5.6) %, respectively. 3- and 5-year PFS of patients were (74.4±5.1)% and (61.2±6.2)%.</p><p><b>CONCLUSION</b>ladder-style freezing from low temperature refrigerator to liquid nitrogen can reach the same clinical transplantation effect with traditional programmed cooling freezing method in autologous hematopoietic stem cells transplantation. moreover the incidence of complications after transpeantatim does not show increase.</p>

Clinical monitoring of chromosome karyotype and fusion gene expression after allogeneic hematopoietic stem cell transplantation in chronic myelocytic leukemia / 中国组织工程研究

BACKGROUND:It has been reported that 70％ of patients with chronic myeloid leukemia (CML) are negative for cytogenetic and genetic markers within 1-5 months after allogeneic hematopoietic stem cell transplantation (allo-HSCT),but there are still some patients who have repeatedly varied outcomes in cytogenetic and genetic marker detection.Overall,the negative rate is up to 89.5％ at 3-12 months after allo-HSCT.OBJECTIVE:To monitor the changes in cytogenetic and genetic marker expression and to explore the prognostic significance in CML patients undergoing allo-HSCT.METHODS:Seventeen CML patients who had undergone allo-HSCT were enrolled.Chromosome G banding pattern of the bone marrow from these patients were analyzed using short-term culture method and direct method at 30 days,2,3,4,6,12,24,36,48,60,72 months after allo-HSCT.Dual-color fluorescence in situ hybridization was used to detect bcr-abl fusion gene;bcr-abl expressions in primary bone marrow ceils from CML patients were detected using RQ-PCR.Results and conclusion:There were 8/17 cases of male patient/male donor and 7/17cases of male patient/female donor (compatriots).46XX karyotype (women) was detected by multiple reexaminations after transplantation,and there was no Y chromosome or other aberration of chromosome karyotype in their karyotype.Among the 17 cases,1 case of female patient/female donor (compatriots) and 1 case of female patient/male donor (unrelated) manifested 46 XY chromosome karyotype and bcr-abl positive at 1 month after transplantation;after 4 months,these two cases still maintained 46 XY chromosome karyotype but bcr-abl negative;after 4-96 months,the karyotype continued to remain as 46 XY,and bcr-abl (-).Among the 17 cases,1 case of male patient/male donor of full-matched compatriot (brother) manifested that Ph chromosomal bcr-abl gene continuously expressed within 1-12 months after allo-HSCT;then the cases was given donor lymphocyte infusion,and the bcr-abl expression returned to be negative at 48 months after transplantation.To conclude,chromosomal karyotype analysis and bcr-abl fusion gene monitoring provide important reference value for subsequent treatment options and prognosis judgment for CML patients with allo-HSCT.

Fluid-attenuated inversion recovery vascular hyperintensity: Correlation with other radiologic findings in acute ischemic stroke with middle cerebral artery occlusion

Objective: To assess fluid-attenuated inversion recovery (FLAIR) vascular hyper-intensity (FVH) and explore its relationship with CT perfusion (CTP) penumbral/infarct core mismatch ratio and diffusion weighted imaging (DWI) final infarct volume in acute ischemic stroke (AIS) patients with middle cerebral artery occlusion (MCAO). Methods: The CTP and MRI images of 38 AIS patients with MCAO were reviewed. The FVH score (longitudinal direction) [FVH score (L)] and FVH score (transverse direction) [FVH score (T)] were quantified on the FLAIR images. The FVH score (L) (range, 0-16) was based on a rostrocaudal extension of FVH and the FVH score (T) (range, 0-3) was based on FVH supply of the occluded MCA territory. The mismatch ratio was calculated from the ratio of the [mean transit time - cerebral blood volume (CBV)] lesion/CBV lesion on the CTP images. The DWI infarct volume was measured on the DWI images. Results: The mismatch ratio was larger for the group of FVH score (L)=7~8 than those of FVH score (L)=5~6 and FVH score (L)=3~4 (p=0.03), whereas the DWI infarct volume was smaller (p=0.04). Similarly, the mismatch ratio of FVH score (T)=2~3 group was larger than FVH score (T)=1 group (p=0.01), whereas the DWI infarct volume was smaller (p=0.02). Both FVH score (L) and FVH score (T) correlated positively with mismatch ratio (P=0.02, P=0.001, respectively), but negatively with DWI infarct volume (P=0.03, P=0.004, respectively). Conclusions: Higher FVH score is associated with larger mismatch ratio and smaller DWI infarct volume in AIS patients with MCAO. FLAIR vascular hyperintensity may represent collateral arterial circulation, and may play a role in protecting the ischemic penumbra.

MicroRNA-20a Promotes Osteogenic Differentiation of C3H/10T1/2 Cells through Regulating CKIP-1 Expression / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of microRNA-20a(MiR-20a) on osteogenic differentiation of mouse C3H/10T1/2 cells and its regulatory mechanism.</p><p><b>METHODS</b>Osteogenic differentiation of C3H/10T1/2 cells were identified by ALP staining and qRT-PCR. MiR-20a mimics and CKIP-1 siRNA were transfected into C3H/10T1/2 cells respectively with lipo3000. The expression of osteoblast marker genes, miR-20a and CKIP-1 were quantitatively assessed by qRT-PCR.</p><p><b>RESULTS</b>miR-20a expression was up-regulated during osteoblast differentiation of C3H/10T1/2 cells. Overexpression of miR-20a promoted osteogenic differentiation. Furthermore, miR-20a inhibited the expression of bone formation negative regulator CKIP-1. Additionally, CKIP-1 knockdown promoted osteogenic differentiation.</p><p><b>CONCLUSION</b>MiR-20a promotes osteogenic differentiation of C3H/10T1/2 cells possibly through inhibiting the expression of CKIP-1.</p>

A novel panel of serum miR-21/miR-155/miR-365 as a potential diagnostic biomarker for breast cancer

PURPOSE: Insufficient sensitivity and specificity prevent the use of most existing biomarkers for early detection of breast cancer. Recently, it was reported that serum microRNAs (miRNAs) may be potential biomarkers in many cancer diseases. In this study, we investigated whether serum levels of 5 miRNAs including miR-21, miR-125b, miR-145, miR-155, and miR-365 could discriminate breast cancer patients and healthy controls. METHODS: Serum levels of miRNAs were measured by using quantitative real-time polymerase chain reaction in 99 breast cancer patients and 21 healthy controls. The abundance change of serum miRNAs were also evaluated following surgical resection in 20 breast cancer patients. Receiver operating characteristic (ROC) curve analysis was performed to assess the sensitivity and specificity of miRNAs as diagnostic biomarkers. RESULTS: Serum levels of miR-21 and miR-155 was significantly higher, while miR-365 was significantly lower in breast cancer as compared with healthy controls. The serum levels of miR-21 and miR-155 significantly decreased following surgical resection. Additionally, the serum level of miR-155 at stages I and II was significantly higher compared to stage III. The serum miR-145 level was remarkably higher in progesterone receptor (PR)-positive patients than PR-negative. The positivity of miR-21, miR-155, and miR-365 was high compared to CA 153 and CEA in breast cancer. ROC curve analyses of a combination of miR-21, miR-155, and miR-365 yielded much higher area under curve and enhanced sensitivity and specificity in comparison to each miRNA alone. CONCLUSION: The combination of serum miR-21/miR-155/miR-365 may potentially serve as a sensitive and specific biomarker that enables differentiation of breast cancer from healthy controls.

Contents of seven indicative constituents in seven kinds of Gentiana macrophylla by UPLC / 中草药

Objective: An ultra performance liquid chromatographic (UPLC) method was developed for simultaneously determining seven constituents, such as loganic acid, swertiamarin, 6'-O-β-D-glucosyl gentiopicroside, gentiopicroside, sweroside, isoorientin, and isovitexin, from the medicinal plants of Gentiana macrophylla. Methods: The separation was performed on an Acquity UPLC® BEH C18 column (50 mm ×2.1 mm, 1.7 μm) through a gradient elution of methanol-0.04% aqueous phosphorite at a flow rate of 0.3 mL/min. The detection wavelength was 242 nm, and the column temperature was set at 30℃. Results: For the seven analytes, loganic acid, swertiamarin, 6'-O-β-D-glucosyl gentiopicroside, gentiopicroside, sweroside, isoorientin, and sovitexin, a good linearity (r≥0.9995) was obtained in the range of 2.100-537.100, 1.050-270.000, 0.920-236.000, 11.100-2830.000, 0.750-192.000, 0.167-102.000, and 0.216-52.800 μg, respectively. Their average recoveries (n=6) were 97.83%-100.08%, respevtively, with RSD values less than or equal to 3.76%. Conclusion: The UPLC method is simpler and more effective than HPLC, and can be used for the simultaneous determination of seven indicative constituents in medicinal plants of G. macrophylla.

Analysis of commercial specifications and grades of wild and cultivated Gentianae Macrophyllae Radix based on multi-indicative constituents / 中国中药杂志

The study is aimed to analyze the commercial specifications and grades of wild and cultivated Gentianae Macrophllae Radix based on multi-indicative constituents. The seven kinds of main chemical components containing in Gentianae Macrophyllae Radix were determined by UPLC, and then the quality levels of chemical component of Gentianae Macrophyllae Radix were clustered and classified by modern statistical methods (canonical correspondence analysis, Fisher discriminant analysis and so on). The quality indices were selected and their correlations were analyzed. Lastly, comprehensively quantitative grade division for quality under different commodity-specifications and different grades of same commodity-specifications of wild and planting were divided. The results provide a basis for a reasonable division of specification and grade of the commodity of Gentianae Macrophyllae Radix. The range of quality evaluation of main index components (gentiopicrin, loganin acid and swertiamarin) was proposed, and the Herbal Quality Index (HQI) was introduced. The rank discriminant function was established based on the quality by Fisher discriminant analysis. According to the analysis, the quality of wild and cultivated Luobojiao, one of the commercial specification of Gentianae Macrophyllae Radix was the best, Mahuajiao, the other commercial specification, was average , Xiaoqinjiao was inferior. Among grades, the quality of first-class cultivated Luobojiao was the worst, of second class secondary, and the third class the best; The quality of the first-class of wild Luobojiao was secondary, and the second-class the best; The quality of the second-class of Mahuajiao was secondary, and the first-class was the best; the quality of first-class Xiaoqinjiao was secondary, and the second-class was the better one between the two grades, but not obvious significantly. The method provides a new idea and method for evaluation of comprehensively quantitative on the quality of Gentianae Macrophyllae Radix.

Quality regionalization study on Gentianae Macrophyllae Radix / 中国中药杂志

The 79 samples of Gentianae Macrophyllae Radix were collected based on the distributed information by document literature. Based on sample information, and using the regression model of Gentianae Macrophyllae Radix index component and environmental factors, and combined with the prediction results of ecological suitability by MaxEnt and principal component analysis results of index component, the space distribution of Gentianae Macrophyllae Radix was estimated with the spatial analysis function of ArcGIS. The results showed that it had a higher comprehensive quality in south of Shaanxi, south of Gansu, middle of Sichuan and southeast of Xizang. The study results were coinciding with the producing regions of Gentianae Macrophyllae Radix. It can provide reference for Gentianae Macrophyllae Radix resource conservation, development and utilization.

Ecology suitability study of Chinese materia medica Gentianae Macrophyllae Radix / 中国中药杂志

This paper is aimed to predict ecology suitability distribution of Gentianae Macrophyllae Radix and search the main ecological factors affecting the suitability distribution. The 313 distribution information about G. macrophylla, 186 distribution information about G. straminea, 343 distribution information about G. dauricaand 131 distribution information about G. crasicaulis were collected though investigation and network sharing platform data . The ecology suitable distribution factors for production Gentianae Macrophyllae Radix was analyzed respectively by the software of ArcGIS and MaxEnt with 55 environmental factors. The result of MaxEnt prediction was very well (AUC was above 0.9). The results of predominant factors analysis showed that precipitation and altitude were all the major factors impacting the ecology suitable of Getiana Macrophylla Radix production. G. macrophylla ecology suitable region was mainly concentrated in south of Gansu, Shanxi, central of Shaanxi and east of Qinghai provinces. G. straminea ecology suitable region was mainly concentrated in southwest of Gansu, east of Qinghai, north and northwest of Sichuan, east of Xizang province. G. daurica ecology suitable region was mainly concentrated in south and southwest of Gansu, east of Qinghai, Shanxi and north of Shaanxi province. G. crasicaulis ecology suitable region was mainly concentrated in Sichuan and north of Yunnan, east of Xizang, south of Gansu and east of Qinghai province. The ecological suitability distribution result of Gentianae Macrophyllae Radix was consistent with each species actual distribution. The study could provide reference for the collection and protection of wild resources, meanwhile, provide the basis for the selection of cultivation area of Gentiana Macrophylla Radix.

Prognostic value of the ABCD2 score on long-term follow-up of transient ischemic attack using the new tissue-based definition

The ABCD2 score is validated for evaluating short-term stroke risk after transient ischemic attack (TIA); however, whether it is able to predict the long-term risk of vascular outcome remains uncertain. Recently a new tissue-based definition of TIA has been proposed. The ABCD2 scores of 145 TIA patients admitted to our hospital were retrospectively calculated and stratified into two categories: ≤ 3 points (low risk); 4-7 points (moderate-high risk). At a median follow-up of 81 months, new vascular events were recorded. Follow-up data were available in 107 patients. Seventy one patients had a moderate-high ABCD2 score. Sixty six patients experienced a cerebral ischemic event; 8 a myocardial infarction; 7 died of cerebrovascular or cardiovascular cause. Moderate-high ABCD2 score was significantly associated with the further cerebral ischemic events (hazard ratio [HR], 1.755; 95% confidence interval [CI], 1.019 to 3.024) and with the combined endpoint (HR, 1.818; 95% CI, 1.079 to 3.063). Our study shows that the ABCD2 score may also be used to predict long-term vascular outcome after tissue-based definition of TIA. Moderate-high ABCD2 score is associated with an increased general vascular risk in the long-term follow-up after TIA.

The role of double-balloon enteroscopy following capsule endoscopy in diagnosis of obscure Small intestinal diseases.

OBJECTIVE: The aim of this study was to evaluate the detection rate accuracy of Double-balloon Enteroscopy (DBE) after Capsule Endoscopy (CE) in patients with suspected small bowel diseases. METHODOLOGY: From January 2009 to March 2012, sixty-two patients with obscure small bowel diseases who underwent CE followed by DBE were included in this study. Introduction of the endoscope by DBE was either orally or anally according to CE. RESULTS: Sixty-two patients are reported. The overall detection rate of small bowel diseases using CE was 70.9% (44/62). Sixty-eight DBE procedures following capsule endoscopy were carried out, There was no significant difference (χ(2)=0.6739, P>0.05) of Positive findings between CE and CE +DBE. Furthermore, the detection rate of small bowel diseases in patients with obscure small intestinal bleeding using CE +DBE (90.9%, 30/33) was superior to that of CE (78.8%, 26/33); χ(2)=1.8857, P>0.05. CONCLUSIONS: Capsule Endoscopy (CE) can cover the whole GI tract and provide the selection of the route of Double-balloon enteroscopy (DBE). DBE can also serve as a good complementary approach after an initial imaging using CE. It can verify the findings of CE and provide therapeutic intervention. Using of CE followed by DBE is effective in the diagnosis and management of patients with obscure small bowel diseases.

Small Endoscopic Sphincterotomy plus Large-Balloon Dilation for Removal of Large Common Bile Duct Stones during ERCP.

OBJECTIVE: This study compared the therapeutic benefits and complication rates of small endoscopic sphincterotomy plus large-balloon dilation (ESLBD) with those of endoscopic sphincterotomy (EST) alone for large bile duct stones. METHODS: We compared prospectively ESLBD group (n=63) with conventional EST group (n=69) for the treatment of large bile duct stones (≥15mm). Mechanical lithotripsy was performed when the stone could not be removed using a normal basket. We compared the rates of stone removal, frequency of mechanical lithotripsy use, procedure-related complications, and recurrent stones. RESULTS: A total of 132 patients were reviewed in the study. The mean age of the patients was 67.9 years. The two groups showed significant differences in complete stone removal during the first session (80.9 vs. 60.8%; P = 0.046), the use of mechanical lithotripsy (7.94 vs. 24.6%; P = 0.041), and less duration of admission (P =0.045). After ERCP, there were some instances of oozing in both groups, All patients recovered completely, 14 patients had recurrent common bile duct stones among the follow-up duration. CONCLUSION: The ESLBD technique seems to be a feasible and safe alternative technique for conventional EST and EBD and has no more Post-ERCP complications.

A new method for isolating and culturing mouse bone marrow mesenchymal stem cells / 中国实验血液学杂志

This study was purposed to establish a convenient and efficient method for isolating and culturing mouse bone marrow mesenchymal stem cells (MSC). The femurs and tibias of mouse were taken under sterile condition. MSC were isolated and cultured with flushing- out bone marrow or collagenase-digested bone fragment or bone marrow plus bone fragment. MSC colony number and size were compared. Immunophenotype and differentiation ability were tested to identify MSC. The results showed that colonies from bone marrow plus bone fragment group came out earliest and the colony number was 20 ± 4 at day 4; there were 11.5 ± 2.5 colonies in collagenase-digested bone fragment group and 9.5 ± 1.5 in flushing- out bone marrow group. The total cell yields of MSC after passaging showed best in bone marrow plus bone fragment group. Flow cytometry data showed the cultured cells expressed Sca-1, CD44 and CD29, not expressed pan-leukocyte surface marker CD45 and endothelial cell marker CD31. The isolated and cultured MSC could differentiate into osteoblast at the osteogenic differentiation condition, or adipocyte at adipogenic differentiation condition. It is concluded that the method of bone marrow plus bone fragment is convenient and efficient for isolating and culturing MSC.

HCCR-2 enhances the division and proliferation of SMMC 7721 via PI3K/Akt pathway / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To observe the biological changes of SMMC 7721 cell line which was stably transfected with HCCR-2 gene and to study the molecular mechanism of HCCR-2 expression in SMMC 7721 cell line.</p><p><b>METHODS</b>SMMC7721 cells were transfected with HCCR-2-pEGFP-N1 and pEGFP-N1 by lipofectmine 2000 and the transfectants were selected and confirmed using Western-blot technique. Cell cycles were tested by flow cytometry. The reproductive activity was detected by MTT assay. SMMC 7721 cells were routinely cultured with treatment of increasing concentrations (0, 50, 100, 200 ng/ml) of EGF for 24 h. Cells were pretreated with the specific inhibitor of PI3K (LY294002) and then cultured with EGF (100 ng/ml) for 24h and the expression of HCCR protein in these cells were measured by Western blot. Another set of SMMC 7721 cells were transfected with DN-Akt-pcDNA3.1 (dominant negative Akt kinase) and pcDNA3.1 by lipofectmine 2000. The transfectants were selected and confirmed using Western-blot technique as before. HCCR-2 and bcl-2 were measured on protein level by Western blot.</p><p><b>RESULTS</b>SMMC 7721 cells were stably transfected with HCCR-2 gene. HCCR-2 gene transfection can increase the proportion of S-phase cells (21.62%+/-1.33% vs 15.76%+/-0.73%, P<0.01) and decrease the cell apoptosis (1.28%+/-0.16% vs 7.72%+/-0.23%, P<0.01). MTT assay showed the growth of HCCR-2 gene transfected cells was faster than that of empty vecter transfected cells. The HCCR-2 protein was up-regulated in a dose-dependent manner in the cells cultured with different concentrations of EGF for 24 h. Treatment of SMMC 7721 cells with specific inhibitor of PI3K (LY294002) suppressed EGF-induced HCCR-2 expression. HCCR-2 protein was down-regulated in dominant negative Akt transfectants.</p><p><b>CONCLUSIONS</b>EGF upregulated HCCR-2 protein expression in SMMC 7721 cell line via PI3K/Akt pathway. The overexpression of HCCR-2 could enhance the division and proliferation of SMMC 7721.</p>

Treatment strategy and pattern evaluation of 314 patients with recurrence of rectal cancer / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To evaluate the treatment strategy, prognosis and pattern of recurrence in patients with rectal cancer.</p><p><b>METHODS</b>From May 1979 to November 2006, 314 patients with recurrence after rectal cancer resection were included in this study. Patients were divided into two groups: local recurrence (LR) and distant metastasis (DM). The clinicopathologic features, treatment strategies and prognosis were analyzed.</p><p><b>RESULTS</b>Of the 314 patients with recurrence, 168 (53.5%) were LR with a mean recurrence-free interval (RFI) of (24.7+/-1.9) months and 146 (46.5%) were DM with a mean RFI of (22.7+/-1.9) months. Compared to the DM group, the patients in the LR group showed no significant difference in clinicopathological data except the time to recurrence (P<0.01), primary tumor location (P=0.043), and the postoperative use of chemoradiotherapy (P=0.007). Mean recurrence-specific survival(RSS) was (24.7+/-1.9) months for LR and the 3- and 5-year survival rates were 0.48 and 0.25. The 3- and 5-year survival rates in patients with DM were 0.33 and 0.16 with a mean RSS of (22.7+/-1.9) months. The difference was statistically significant (P<0.01). Cox regression analysis for RSS showed that the time to recurrence, TNM stage, and treatment strategy (including procedure and the use of postoperative chemoradiation) were independently prognosis factors for the patients with recurrence rectal cancer (all P<0.01). Subgroup analyses revealed no significant differences in RFI or RSS among different subgroups within either LR or DM groups.</p><p><b>CONCLUSIONS</b>Patients of rectal cancer with LR have a better survival than those with DM. Moreover, radical resection can improve the prognosis of patients with recurrence of rectal cancer, especially for patients with early TNM stage of the primary tumor and later period of recurrence.</p>