OBJECTIVE:To establish the method for the detection of the known glucagon-like peptide 1 receptor (GLP1R) gene mutation site rs3765467(NT_007592.16,position:39065819),and to evaluate its accuracy and practicability. METHODS:Peripheral venous blood samples of 72 healthy subjects were collected in medical examination center of our hospital during Oct. 2015-Feb. 2016. The whole blood DNA was extracted by column extraction method. After amplified by touch down PCR,high reso-lution melting(HRM)method was adopted to analyze amplified product. Sequencing verification by double stranded chain termina-tion method(Sanger sequencing method)was performed for 38 test samples. The results of 2 methods were compared. RESULTS:The results of mutation scanning showed that there were 39065817 and 39065819 polymorphism sites in amplified segments. Four types of mutations were detected by HRM method [GCG/GCG,GCA/GCG or ACG/GCG,GCA/GCA or ACG/ACG,A(G) CA(G)],but 6 types of mutations was detected by Sanger sequencing method [GCG/GCG,ACG/GCG,ACG/ACG,A(G)CA (G),GCA/GCG,GCA/GCA]. CONCLUSIONS:HRM method can identify GCG/GCG and A(G)CA(G)genotype,but can not identify GCA/GCG and ACG/GCG heterozygous mutation,GCA/GCA and ACG/ACG homozygous mutation. The method is not suitable for the detection of single nucleotide polymorphism for multiple neighboring sites. In the detection of single nucleotide mu-tation,economical and simple method should be selected after comprehensive analysis of sequence.

Objective To investigate the application of the ultrasonic guidance-assisted neonatal internal jugular vein catheterization.Methods Sixty two newborns (including low birth weight infants) receving thoracic/abdominal operation or resection of malignant tumor on the body-surface were randomly assigned to ultrasound guidance (UG) group or surface mark landmark(S) group.Newborns in both groups were all punctured with 22G venous indwelling needles to place the external casing,followed by the steel wire guidance-assisted implantation of ARROW 4F dual chamber central venous catheter.Then we compared the rate of successful insertion attempt,rate of malpositioning,complications and average operation time between the two groups.Results The rate of successful insertion attempt was 96.8%(30/31) in the UG group,significantly higher than that in the S group (32.3%,10/31),there was significant difference between the two groups(χ2=28.182,P=0.000).Malpositioning happened in 2 cases in the UG group,but 25 cases in the S group.Rate of complications was higher in the S group compared to the UG group (64.5% vs 3.2%,χ2=25.99,P=0.000).Most importantly,the average operation time was (4.366±1.137)min in the UG group,significantly shorter than that of the S group [(13.70±5.34)min,t=5.463,P=0.028)].ConclusionUltrasound guidance-assisted catheterization for neonatal internal jugular vein is safe and feasible and can dramatically improve the success rate and prevent complications.

Objective To observe the influence of coal burning fluorosis on learning and memory ability in rats and reveal its possible mechanisms. Methods Healthy 48 SD rats were divided into control, low-fluoride and high-fluoride group. All rats in fluoride exposed groups were fed with the eom polluted by drying processes with burning coal containing high level of fluoride obtained from the endemic fluorosis area to produce the animal model of fluorosis. The experiment period were 3,6 mouths, respectively. The ability of leaning and memory was measured by Morris test and cholinesterase activity detected by photometric method at 3 or 6 month after experiment, respectively. Results Fluoride contents signifieantlly influenced the escape latency, the numbers of crossing the platforms and the time of staying the platforms(the value of F was 29.29,6.47,6.50, respectively, P<0.01).In addition, the numbers of crossing the platforms and the time of staying the platforms were influenced by the exposed time(the value of F was 16.11,45.59, P<0.01). Furthermore, the fluoride contents and the exposed time had an interaction between the numbers of crossing the platforms and the time of staying the platforms (the value of F was 4.67,5.68, P<0.05 or<0.01). Three months after the experiment, the mean values of escape latency [(14.71± 4.85)s] of rats in highly fluoride exposed group were significantly prolonged as compared with controls [(9.28±4.22)s]; 6 month after the experiment, the mean values of escape latency[(12.42±8.03)s, (17.48± 8.05)s] of rats in both groups exposed to fluoride were significantly prolonged as compared to controls [(7.04± 3.29)s, P<0.05]. The decreased numbers of crossing the platforms[(1.62±0.87)number] and the declined time of staying the platforms[(16.70±5.02)s] were found in the rats exposed to high fluoride as compared to controls [(3.53±1.67 )number, (23.33±5.35)s, P<0.05]. The fluoride contents obviously influenced the activities of acetylcholinesterase and butylcolinesterase (the value of F was 12.83,13.27, P<0.01). On the other hand, the times of breeding also influnced the activities of butylcolinesterase (the value of F was 16.26, P<0.01). In 3 months of the experiment, the activities of butylcolinesterase [(0.55±0.12)kU/g] in low fluoride exposed group were significantly decreased in comparison with controls[(0.73±0.10)kU/g, P<0.05]. The activities of acetylcholinesterase[(0.62±0.42)kU/g] and butylcolioesterase[(0.58±0.10)kU/g] in high fluoride group were significantly decreased as compared to eontrois[(1.41±0.52), (0.73±0.10)kU/g, P<0.05]. The correlation analysis showed that there was a negative correlation between the cholinesterase and the escape latency(r=-0.68, P< 0.01), and a positive correlation between the cholinesterase and the time of staying the platforms(r=0.57, P< 0.01). Conclusions The ability of learning and memory in rats with coal buring fluorosis was decreased, which might be connected to the decreased activity of cholinesterase in a dose-effect correlation.

BACKGROUND:In the treatment of tibial plateau fractures, because of the variety of fracture, the complexity of anatomical changes, X-ray films or three-dimensional CT scan limited by two-dimensional plane, increases the difficulty in preoperative plan and surgical treatment. The application of three-dimensional (3D) printing technology has attracted attention in the department of orthopedics. OBJECTIVE:To explore the auxiliary role of 3D printing technique in preoperative plan and treatment for tibial plateau fractures. METHODS:Thirty patients with tibial plateau comminuted fractures were enroled in this study and divided into two groups: experimental and control groups, with 15 patients in each group. In the experimental group, patients underwent 3D CT scan, which was stored in DICOM format, and processed by Mimics software. Data were converted into STL format, entered 3D printer, and a 1:1 entity size of the fracture model was made, in accordance with repair plan of 3D fracture model. Operation time and intraoperative blood loss were compared between the two groups. At 12 months after treatment, their outcomes were assessed using Rasmussen evaluation criteria. RESULTS AND CONCLUSION: The 3D printing fracture models of 1:1 ratio identified fracture type and made a repair program before surgery in the experimental group. Operation time and intraoperative blood loss were significantly less in the experimental group than in the control group (P < 0.05). After surgery, patients were folowed up for 12 to 18 months. The healing time was 3-5 months, averagely 4.3 months. At 12 months after treatment, the Rasmussen evaluation criteria results showed that the excelent and good rate was significantly higher in the experimental group than in the control group (P < 0.05). These results suggest that the fracture model of 3D can help to make the operation plan. The treatment of tibial plateau fractures is more precise, personalized and visual.

BACKGROUND:Titanium and titanium aloy are used mostly in artificial joints, fracture fixation, and oral transplantation, while there are complex cases of insufficient bone mass in these areas. The deepened research of stem cels offers a solution for bone injury to promote new bone formation. The biocompatibility of titanium and stem cels and optimization of titanium surface modification have aroused people's attention. OBJECTIVE:To investigate whether the biocompatibility of titanium and human adipose-derived mesenchymal stem cels can be improved by type I colagen modification of titanium sheets. METHODS:The experiment was divided into two groups. Modification group: titanium sheet was modified with type I colagen; control group: titanium sheet was not modified with type I colagen. Human adipose-derived mesenchymal stem cels at passage 6 were implanted into titanium sheet in two groups. Then we calculated the number of adherent cels in two groups at 1, 2 and 4 hours after implantation, and compared the celladhesion rate. MTT assay was used to observe the proliferation of cels on titanium sheet at 2, 4, 6 and 8 days after implantation. DNA and protein content of cels were detected at 3, 6, 9 days after implantation. The growth of human adipose-derived mesenchymal stem cels seeded upon the titanium sheets was observed under scanning electron microscope at 6 days. RESULTS AND CONCLUSION:When the cels were cultured for 1 hour and 2 hours, the number of adherent cels in the modification group was higher than in the control group (P < 0.05). The absorbance of cels in two groups was increased as the culture time, as detected by MTT assay. The modification group had a significantly higher absorbance value than the control group at 4, 6, 8 days (P < 0.05). DNA and protein contents of the cels in the modification group were higher than that in control group at 6 and 9 days (P < 0.05). At 6 days, the number of adherent cels and secretion of adherent stromal cellmatrix in the modification group were significantly better than that in control group, observed by scanning electron microscopy. Type I colagen modified titanium sheets have good surface activity and biocompatibility, and can promote the proliferation of human adipose-derived mesenchymal stem cels.

OBJECTIVETo study the correlation of insulin resistance (IR) with TCM syndrome type and activity of fibrinolytic system in patients with coronary arterial disease (CAD).

METHODSOne hundred and twelve CAD patients were classified according to TCM Syndrome into 4 types, the Xin-blood stasis (XBS) type, the phelgm blocking Xin-channel (PBXC) type, the Qi-insufficiency with blood stasis (QIBS) type and the both Qi-Yin deficiency (QYD) type. Patients' fasting blood glucose (FBG), fasting blood insulin (FIns) were measured, the insulin sensitive index (ISI) calculated. Data were compared between various types, also with those obtained from 30 healthy persons in the control group respectively. Moreover, activity of tissue plasminogen activator (t-PA) and content of plasminogen activator inhibitor-1 (PAI-1) were determined in 90 patients selected from the 112 to conduct linear correlation analysis of IR with t-PA activity and PAI-1 content.

RESULTSFBG and FIns levels in the CAD patients were higher than those in the healthy control significantly (P < 0.01); ISI in the 4 syndrome type of CAD patients were all lower than that in the control (P < 0.01). IR existed in all the 4 types, but the level in the XBS type and the PBXC type was more severe than in the other two types. Correlation analysis showed that IR was correlated with t-PA activity and PAI-1 content (P < 0.01).

CONCLUSIONIR often exists in CAD patients, the severity of IR varies in patients of different TCM syndrome types, and IR is correlated with the abnormality of fibrinolytic system activity.

Adult ; Aged ; Aged, 80 and over ; Angina Pectoris ; blood ; diagnosis ; Diagnosis, Differential ; Female ; Fibrinolysis ; Humans ; Insulin Resistance ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Myocardial Infarction ; blood ; diagnosis ; Plasminogen Activator Inhibitor 1 ; metabolism ; Tissue Plasminogen Activator ; metabolism

BACKGROUND:As a noticeable tissue engineering material of polyhy droxyalka noates family, poly (3-hydroxybutyrate-co-4-hydroxybutyrate)(P3HB4HB) exhibitsgood biocompatibility, adhesion and mechanicalproperties, presenting aextensive application future in tissue-engineered research.
OBJECTIVE:To investigate the biocompat ibilityin vitroand ectopic osteogenic differentiationin vivoof P3HB4HB and human bone marrow mesenchymal stem cels.
METHODS:Passage 5human bone marrow mesenchymal stem cels transplanted ontothe three-dimensional P3HB4HB scaffoldwereincubated with osteogenic induction medium (test group)or with no osteogenic induction(control group), respectively. After 5-day incubation, thecelgrowth was assessed by acridine orange staining and scanning electron microscopy; after14-day incubation, both kinds of cel-scaffold composites were subcutaneously implanted into the nude mice. At 16 weeks after implantation, the cel-scaffold composites were removed to observeectopic osteogenic differentiationin vivousing hematoxylin-eosin staining, von Kossa staining and colagen type I immunohistochemical staining.
RESULTS AND CONCLUSION:Acridine orange staining showed that cels adhered wel on the surface of the scaffold;under thescanning electron microscope, induced celsgrew wel on the P3HB4HB scaffold and produced abundant extracelular matrixes. In addition, at 16 weeks after implantation, there were osteoidtissues in the test group, positive for von Kossa staining as wel as colagen type I immunohistochemical staining;furthermore, hematoxylin-eosin staining showednumerous osteoblasts and bone lacunas. In contrast, no bone tissues appeared in the control group. To conclude, P3HB4HB is a suitable material for bone tissue engineering.

To study the contribution of T cell subsets in the pathogenesis of Murine hepatitis virus Type3 (MHV-3) induced chronic viral hepatitis in C3H/Hej mice, ninety C3H/Hej mice were chosen to individually receive 10 plaque forming units (PFU) of MHV-3 intraperitoneally. The changes of virus titer and pathology in liver tissue were examined by standard plaque assay and by the hematoxylin/eosin (HE) staining method from 2 days post MHV-3 infection. The ratios of T cell subsets including CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4-CD8-, CD3+CD4+CD25+, CD3+CD4+CD25- and CD3+CD4-CD25+ T lymphocyte of total T lymphocytes in blood, spleen and liver were examined at 0, 2, 4, 6,8, 10, 12, 15, 20, 25, 30, 40 days post MHV-3 infection by flow cytosorting. We observed that the virus titer raised and showed persistent virus duplications and inflammatory changes in the livers of C3H/Hej mice from 2 days post MHV-3 infection. The double negative T cell (DN Treg cell) and CD4+CD25+ T cell ratios increased significantly from 2 days post MHV-3 infection in C3H/Hej mice, and CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4+CD25- and CD3+CD4-CD25+ T cell ratios decreased accordingly. In conclusion, the changes of virus titer and pathology in the livers of C3H/Hej mice post MHV-3 suggest their contribution to viral persistence. Further characterizations of DN Treg cells are that infection indicates that MHV-3 could induce the chronic inflammation in livers of C3H/Hej mice.The increase of the DN Treg cell and CD4+CD25+ T cell ratios in C3H/Hej mice post MHV-3 infection suggests that DN Treg cells and CD4+CD25+ T cells may both have important suppressive immunomodulation functions in the development of chronic viral hepatitis and have important roles in the virus persistent infection. Further characterizations of DNT cell and CD4+CD25+ T cell are under investigation.

OBJECTIVETo discuss the expression and significance of glial cell-derived neurotrophic factor (GDFN) receptor alpha 1 gene (GFR alpha 1) in the recovery spermatogenesis of mice.

METHODSAdult Kunming mice were injected intraperitoneally with 2 doses of busulfan (10 mg/kg) 24 days apart so as to establish the recovery spermatogenesis model. Testes were harvested 1 w, 2 w, 3 w, 4 w, 6 w, 8 w and 10 w after the second injection, and normal testes were used as control. The recovery spermatogenesis was observed by light and electron microscopy, and the GFR alpha 1 mRNA was measured by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization.

RESULTSThe expression of GFR alpha 1 mRNA increased significantly at 1 w and reached its peak at 2 w after the second injection [(104.72 +/- 24.4)% vs normal control, P < 0.01]; its expression reduced significantly at 3 w and reached its valley at 4 w [(20.77 +/- 4.25)% vs normal control, P < 0.01], and then increased gradually and restored to the normal level at 10 w. GFR alpha 1 mRNA was mainly expressed by undifferentiated spermatogonia.

CONCLUSIONSIn the course of recovery spermatogenesis, the expression of GFR alpha 1 plays a key role in turning the spermatogonial stem cell reactivity to GDNF, which promotes self-renewal at a high level, or results in differentiation at a low level.

Animals ; Glial Cell Line-Derived Neurotrophic Factor Receptors ; In Situ Hybridization ; Male ; Mice ; Proto-Oncogene Proteins ; genetics ; Proto-Oncogene Proteins c-ret ; RNA, Messenger ; analysis ; Receptor Protein-Tyrosine Kinases ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Spermatogenesis ; Testis ; pathology

Abstract: Mycobacterium senegalense is one of the major pathogens causing bovine farcy, and reports of its infection in human are rare. Here is a report on a woman who had been taking hormones and immunosuppressants for a long time for SLE and underwent abdominal soft tissue infection with Mycobacterium senegalense after abdominal liposuction, to provide reference for clinical diagnosis and treatment. The patient, female, 32 years old, has a history of SLE for more than 2 years, and currently takes "methylprednisolone, hydroxychloroquine, and mycophenolate mofetil" regularly. Nine months before the patient was admitted to the hospital, she once performed abdominal, waist and buttock liposuction in a medical beauty institution. One month after the operation, several masses gradually appeared on the abdominal wall, accompanied by tenderness, one of the masses had obvious fluctuation on palpation and purulent fluid could be drawn out. The location of the abdominal wall mass was consistent with the insertion site of the liposuction needle. After the onset of the disease, the patient went to the medical beauty institution for puncture of the abdominal wall mass, and 5 mL of purulent fluid was pierced and sent for bacterial culture, and cultured "Mycobacterium Senegalense", after 3 days of treatment with "cephalosporin" antibiotics (specifically unknown), the symptoms did not improve, so she went to the second affiliated hospital of hainan medical college. After completing the relevant examinations during the hospitalization in our hospital, in order to clarify the etiology, another abdominal puncture to extract pus was performed, the mycobacterial culture + identification results: Mycobacterium senegalense. Consistent with the out-of-hospital results, the diagnosis of Mycobacterium senegalense infection was confirmed. After 3 months of treatment with "cefoxitin, azithromycin, amikacin, and levofloxacin", the patient's abdominal wall soft tissue infection was cured. Trauma or invasive procedures can lead to skin, muscle, or bone infection with nontuberculous mycobacteria (NTM), which can manifest as chronic painless nodules that progress to purulent folliculitis and abscesses. NTM infection should be suspected when the patient's wound has been exposed to water, there is a history of surgery, and empirical anti-infection is ineffective. This is the first case of Mycobacterium senegalense infection caused by medical beauty, which tell people that they should be cautious when choosing medical aesthetic projects and medical aesthetic institutions.