This study was aimed to investigate the expression level of murine double minute 4 (MDM4) mRNA in chronic lymphocytic leukemia (CLL) and its prognostic value in CLL. By means of β-actin as internal reference, the real-time quantitative RT-PCR was set up. The expression of MDM4 mRNA in 66 CLL patients was measured by fluorescence dye SYBR Green I. The dispersion of MDM4 expression ratio of groups with different prognostic factors was described by using Mann-Whitney U test. The results showed that the median MDM4 mRNA expression level was 0.037098 (0.088245-0.014875) in CLL patients. The expression level of MDM4 mRNA was significantly higher in patients with P53 gene deletion than that in patients without P53 gene deletion (0.13167 vs 0.030927) (p < 0.001), and also significantly higher in patients with P53 mutation than that in patients without P53 mutation (0.13167 vs 0.03077) (p < 0.001). MDM4 expression was also associated with Binet stages (p = 0.044) and ATM gene deletion (p = 0.046), but was not associated with LDH (p = 0.216), β(2)-MG (p = 0.314), TK1 (p = 0.300), ZAP-70 (p = 0.559), CD38 (p = 0.513) and IgVH mutation status (p = 0.333). It is concluded that the expression level of MDM4 is significantly higher in patients with P53 deletion or mutation. MDM4 expression is significantly associated with Binet stages and ATM gene deletion. MDM4 may be an important prognostic factor in CLL.
Adult ; Aged ; Aged, 80 and over ; Female ; Humans ; Leukemia, Lymphocytic, Chronic, B-Cell ; diagnosis ; genetics ; metabolism ; Male ; Middle Aged ; Nuclear Proteins ; genetics ; metabolism ; Prognosis ; Proto-Oncogene Proteins ; genetics ; metabolism ; RNA, Messenger ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Tumor Suppressor Protein p53 ; genetics

This study was purposed to establish a method for detecting miRNA expression by using fluorescent quantitative polymerase chain reaction (RQ-PCR), and to investigate the application value of this method in quantitative detection of miRNA. Total RNA was extracted from the peripheral blood or bone marrow of 82 patients with chronic lymphocytic leukemia (CLL) and 70 patients with acute leukemias (AL). Standard curves of U6 and miRNA were constructed from 10-fold serial dilutions of the cDNA, the quantitative detection was performed by using real-time quantitative PCR with SYBR Green by Roche Light Cycler. U6 RNA was used as the reference, the relative expression levels of miR-15a, miR-16-1, miR-29b, miR-181a and miR-181b in patients with CLL, while miR-128-1, miR-223, let-7b, miR-155 and miR-181a in patients with AL were analyzed by 2((-DeltaDeltaCT)) method. The relative parameters including specificity, linearity range, sensitivity and repeatability were evaluated. The results showed that the RQ-PCR assay could precisely detect the mature miRNA in as few as 10-50 ng of total RNA with the sensitivity of 0.05 ng RNA. Ct values of miRNA and U6 were all within 15 to 30. A good linear relationship (R(2) > 0.980) was obtained from the standard curves, also the efficiency of amplification was above 90%. Only a specific peak was shown on the melting curve diagram. The coefficients of variation (CV) of interrun and intrarun assay was < 4.8% and 6.3% respectively. It is concluded that the RQ-PCR is a sensitive and fast method for the detection of miRNA with its high-flux and low cost, this method will facilitate the research with detection of miRNA.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Female ; Humans ; Leukemia ; genetics ; MicroRNAs ; genetics ; Middle Aged ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity ; Young Adult

OBJECTIVETo discuss surgical skills,precautions and complications of using titanium elastic intramedullary nails in the treatment of adult midshaft clavicular fractures and evaluate the therapeutic efficacy.

METHODSFrom June 2006 to January 2009, 28 patients with completely displaced midshaft clavicular fractures (15 simple fractures, 8 wedge fractures and 5 comminuted fractures) were enrolled in the study,included 19 males and 9 females with a mean age of 39.0 years (range 19-67 years). The injury was on the left side in 14 cases and on the right side in 14 cases. The mean course of disease was 2.9 days. The shoulders of the patients were swollen, deformed and disabled before operation. X-rays revealed complete displacement of the clavicle. The mean clavicular shortening after injury was 6.76% vs. that measured after bone healing. The Constant-Murley Shoulder Scoring System was used to assess shoulder function, and the DASH Score was used to assess the disability degree of the upper arm.

RESULTSClosed operation with titanium elastic intramedullary nails was undertaken in 26 cases, and open reduction was performed in the remaining two cases. Satisfactory reduction was achieved in all patients, who were followed up for a mean of 10 months (range 6-15 months). The mean union time was 11.5 weeks. No severe complication occurred in any patient. The mean clavicular shortening after bone healing was 3.38%, which was significantly different as compared with the mean clavicular shortening after injury. Constant-Murley Shoulder Score was (97.0 +/- 4.2), and DASH score was (3.4 +/- 4.8). Anatomical reduction, functional recovery and appearance were satisfactory in all patients.

CONCLUSIONTreatment of adult midshaft clavicular fractures with titanium elastic intramedullary nails is advantageous and may prove to be an alternative option for nonsurgical treatment and plate osteosynthesis of midshaft clavicular fractures in adults.

Adult ; Aged ; Clavicle ; injuries ; surgery ; Female ; Fracture Fixation, Intramedullary ; methods ; Fractures, Bone ; surgery ; Humans ; Male ; Middle Aged

OBJECTIVETo examine the effects of various concentration of nicotine on bone marrow stromal cells (BMSCs) proliferation and differentiation of cartilaginous in vitro.

METHODSBMSCs was obtained from femoral bone and tibia of New-Zealand albino rabbit. The cells of the 3rd generation were used in study. Different concentration of nicotine (0, 1 x 10(-7), 1 x 10(-6), 1 x 10(-5) M) were added into BMSCs. BMSCs proliferation was analyzed by MTT assay at the 1, 4, 7, 14 days. The expression of collagen type II and aggrecan as the marker genes of cartilaginous differentiation from BMSCs were detected by reverse transcriptase-polymerase chain reaction (RT-PCR).

RESULTSMicroscope showed that BMSCs transformed from round to fusiform shape. The concentration of nicotine in 1 x 10(-7), 1 x 10(-6) M had a significant positive effect on cell proliferation and the expression of type II collagen in a time-dependent manner when supplemented in commonly used induction media (P<0.05). Concentrations of nicotine in 1 x 10(-7) can promote the expression of aggrecan at the 7th day after induction,and in 1 x 10(-5) M may inhibit the expression of type II collagen and aggrecan.

CONCLUSIONIt was implied that local application of nicotine at an appropriate concentration may be a promising approach for enhancing cartilaginous differentiation capacity of BMSCs in cartilage tissue engineering.

Aggrecans ; genetics ; Animals ; Bone Marrow Cells ; drug effects ; physiology ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Chondrocytes ; cytology ; drug effects ; Collagen Type II ; genetics ; Male ; Nicotine ; pharmacology ; RNA, Messenger ; analysis ; Rabbits ; Stromal Cells ; drug effects ; physiology

OBJECTIVETo evaluate the feasibility and therapeutic effect of treating sternoclavicular joint dislocation by K-wire and tension band wire fixation, and to improve the safety and stability of this technique.

METHODSThis study consisted of 9 cases, 6 males and 3 females with the mean age of 25 years (range, 9-62 years). The causes were traffic accident in 7 cases, falling in 1 case and fight in 1 case. The duration from injury to operation was 2 hours to 7 days. There were 5 left dislocations and 4 right dislocations; 8 anterior dislocations and 1 posterior dislocation, including one combined with left scapular fracture and one with left olecranon fracture. Open reduction and internal fixation using K-wires and tension band wires were performed to treat dislocations.

RESULTSAll patients were followed up for 6 to 24 months, 10 months on average. According to Rockwood's rating scale on postoperative sternoclavicular joint, 8 cases achieved excellent outcomes with an average score of 13.88, and the rest case achieved a good outcome with the score of 12. Anatomical reduction was obtained in all cases. There were no such postoperative complications as severe infection, injury to blood vessel and nerve, failure of fixation, etc. Patients were all satisfied with the anatomical reduction and functional recovery.

CONCLUSIONSThe technique of K-wire and tension band wire fixation is safe, simple, effective, less invasive and has been successfully used in orthopedic surgery. It is effective in treating sternoclavicular joint dislocation though it has some disadvantages.

Adolescent ; Adult ; Bone Wires ; Child ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Joint Dislocations ; surgery ; Male ; Middle Aged ; Radiography ; Sternoclavicular Joint ; diagnostic imaging ; injuries ; surgery

OBJECTIVETo evaluate the preliminary clinical outcomes of coccygectomy in patients with coccydynia after a failure of conservative treatment.

METHODSFrom May 2002 to January 2010, 31 patients with coccydynia were treated by coccygectomy in our department after conservative measures had failed to produce significant relief. A questionnaire, which included the extent of relief in the painful area, improvement in quality of life, intensity of pain in the sitting position, and pain score during daily activities, was used to evaluate the results.

RESULTSAll patients were followed up for 1 to 6 years (mean 3.3 years). The results were excellent in 20 patients (64.5%), good in 7 patients (22.6%), moderate in 3 patients (9.7%) and poor in 1 patient (3.2%). The excellent and good rates amounted to 87.1%. All patients except one had complete resolution of their symptoms and were subjectively highly satisfied with the outcomes of the surgery. Only 2 cases of superficial infection were observed postoperatively.

CONCLUSIONCoccygectomy is a feasible management option for patients with coccygodynia that has no response to conservative treatments.

Activities of Daily Living ; Adult ; Aged ; Coccyx ; surgery ; Female ; Humans ; Low Back Pain ; diagnostic imaging ; surgery ; Male ; Middle Aged ; Pain Measurement ; Radiography

OBJECTIVETo observe proliferation and differentiation of osteoblasts cultured in the plane on appropriate electrical stimulation, to specify whether it promote the proliferation, and observe expression of BMP-2 on electrical stimulation.

METHODSOsteoblasts were extracted from the skull of rabbit offspring and cultured. Cells after the 2nd generations were cultured. In experimental group, cells had electrical stimulation, and same stimulation time and intensity were given. In control group cells had not electrical stimulation. The proliferation and differentiation were detected at different time, and BMP-2 protein expression was analyzed.

RESULTSThe cell morphology of experimental group in 8 days under the light microscope was observed and showed a lot of proliferation of osteoblasts, pleomorphic changes, in 6 to 8 days a small amount of Calcified spots was also observed; while in the control group, proliferation was slower. Differentiation of the experimental group was significantly, alkaline phosphatase staining and calcium nodules were positive, quantitative analysis of alkaline phosphatase increaseed significantly. Experimental group showed that BMP-2 was gradually increased by immunohistochemistry analysis.

CONCLUSIONElectrical stimulation can promote the proliferation and differentiation of osteoblasts and achieved the increasement the number of cells in short-term, intracellular staining by immunohistochemistry showed the increasement in expression of BMP-2.

Animals ; Bone Morphogenetic Protein 2 ; metabolism ; Cell Culture Techniques ; methods ; Cell Differentiation ; Cell Proliferation ; Electric Stimulation ; Gene Expression Regulation ; Osteoblasts ; cytology ; metabolism ; Rabbits

OBJECTIVETo evaluate the in vitro and in vivo osteogenic capability of adipose-derived stromal cells (ASCs).

METHODSASCs were isolated from New Zealand white rabbits and determined by alkaline phosphatase (ALP) staining, von Kossa staining and alizarin red staining. Some specific markers of osteogenic differentiation, including ALP, osteocalcin (OCN), osteopontin (OPN) were examined by reverse transcription-polymerase chain reaction (RT-PCR). In vivo, demineralized bone matrix (DBM)-ASCs composites were implanted into the rabbit calvarial defects created at each side of the longitudinal midline. After 6 weeks, histologic properties of the transplants were analyzed.

RESULTSASCs were successfully induced into osteogenesis. ALP staining, von Kossa staining and alizarin red staining showed positive results. The expressions of ALP, OCN and OPN were detected in ASCs after cultivation in osteogenic medium. Extensive new bone was observed in the defects transplanted with DBM-ASCs composites.

CONCLUSIONASCs have the potential to differentiate into osteogenic lineage and DBM-ASCs constructs are a promising method for regeneration in bone defects.

Adipose Tissue ; Animals ; Cell Differentiation ; Cells, Cultured ; Osteogenesis ; Rabbits ; Stromal Cells

OBJECTIVEStudies showed elastic stable intramedullary nailing (ESIN) of displaced midclavicular fractures has excellent outcomes, as well as high complication rates and specific problems. The aim was to discuss ESIN of midshaft clavicular fractures.

METHODSTotally 60 eligible patients (aged 18-63 years) were randomized to either ESIN group or non-operative group between January 2007 and May 2008. Clavicular shortening was measured after trauma and osseous consolidation. Radiographic union and complications were assessed. Function analysis including Constant shoulder scores and disabilities of the arm, shoulder and hand (DASH) scores were performed after a 15-month follow-up.

RESULTSESIN led to a signifcantly shorter time to union, especially for simple fractures. In ESIN group, all patients got fracture union, of which 5 cases had medial skin irritation and 1 patient needed revision surgery because of implant failure. In the nonoperative group, there were 3 nonunion cases and 2 symptomatic malunions developed requiring corrective osteotomy. At 15 months after intramedullary stabilization, patients in the ESIN group were more satisfied with the appearance of the shoulder and overall outcome, and they benefited a lot from the great improvement of post-traumatic clavicular shortening. Furthermore, DASH scores were lower and Constant scores were significantly higher in contrast to the non-operative group.

CONCLUSIONESIN is a safe minimally invasive surgical technique with lower complication rate, faster return to daily activities, excellent cosmetic and better functional results, restoration of clavicular length for treating mid-shaft clavicular fractures, resulting in high overall satisfaction, which can be regard as an alternative to plate fixation or nonoperative treatment of mid-shaft clavicular fractures.

Adult ; Clavicle ; Fracture Fixation, Intramedullary ; Humans ; Nails ; Titanium ; Treatment Outcome

In order to investigate the relation of serum immunoglobulin (Ig) level to age, sex, disease stages and prognosis in chronic lymphocytic leukemia (CLL) patients, the levels of serum IgG, IgA and IgM in 83 CLL patients were detected by immune rate turbidimetry. the expressions of CD38 and ZAP-70 in CLL cells were determined by multi-parameter flow cytometry (FCM). The results showed that among the 83 CLL patients, the IgG, IgA and IgM levels were reduced in 12 (14.5%), 26 (31.3%) and 34 cases (41.0%) respectively. The incidence of Ig reduction was higher in Binet C stage than that in Binet A and B (p=0.011). There was higher incidence of Ig reduction in high-risk group of Rai stage than that in low-risk group (p=0.011). The positive rate of CD38 or ZAP-70 was significantly higher in Ig reduction group than that in normal Ig level group (p=0.033 and p=0.038 respectively). The positive rate of CD38 and ZAP-70 was also higer in advanced disease, and among them the positive rate of ZAP-70 expression in Binet C stage was obviously higer than Binet A and B (p=0.047). Nonetheless, there was no significant relationship of Ig level with gender and age (p>0.05). It is concluded that the function of humoral immunity in CLL patient is closely related to the disease stages. The serum Ig detection seems important for evaluating immunologic state and prognosis of CLL patients.
ADP-ribosyl Cyclase 1 ; metabolism ; Adult ; Aged ; Aged, 80 and over ; Female ; Humans ; Immunity, Humoral ; Immunoglobulin A ; blood ; Immunoglobulin G ; blood ; Immunoglobulin M ; blood ; Immunoglobulins ; blood ; Leukemia, Lymphocytic, Chronic, B-Cell ; blood ; immunology ; Male ; Middle Aged ; Prognosis ; ZAP-70 Protein-Tyrosine Kinase ; metabolism