组织因子刺激体外培养人脐静脉内皮细胞肿瘤坏死因子α、白细胞介素1的表达

目的:组织因子介导了细胞凝集、组织损伤、血栓形成,从而参与"凝血-炎症-血栓形成"网络。组织因子是否为一种独立的前炎症介质介导炎症反应,刺激人脐静脉内皮细胞使之分泌肿瘤坏死因子α、白细胞介素1。方法:实验于2004-09/2006-02在贵阳医学院组织工程与干细胞实验中心(省级重点实验室)完成。利用重组的组织因子刺激体外培养的人脐静脉内皮细胞,并设立空白对照组、0.2mg/L组织因子刺激组和0.8mg/L组织因子刺激组,用ELISA检测不同组织因子刺激浓度在同一刺激时间(6h)其炎症介质(肿瘤坏死因子α、白细胞介素1)的表达分泌情况。结果:①组织因子可以刺激人脐静脉内皮细胞使之分泌肿瘤坏死因子α,0.2mg/L组织因子刺激组与0.8mg/L组织因子刺激组肿瘤坏死因子α水平高于空白对照组(P<0.05);0.8mg/L组织因子刺激组肿瘤坏死因子α水平高于0.2mg/L组织因子刺激组(P<0.05)。②与空白对照组相比较,0.2mg/L组织因子刺激组白细胞介素1水平未升高,0.8mg/L组织因子刺激组略有升高,但差异无显著性意义(P>0.05)。结论:组织因子可以作为一种前炎症介质介导炎症反应,使内皮细胞肿瘤坏死因子α表达增强。     

重组结缔组织生长因子对体外培养成骨细胞核结合因子α1基因表达的影响

目的:研究发现,结缔组织生长因子可促进软骨细胞和成骨细胞的增殖及其表型的发生,在骨骼发育以及骨量维持方面发挥重要作用,但其对成骨细胞的作用机制目前尚不清楚。观察重组结缔组织生长因子对体外培养人成骨细胞核结合因子α1基因表达的影响。方法:实验于2006-01/2007-01在日照市人民医院完成。①实验材料:外科手术取正常成人髂骨松质骨,患者对试验知情同意。②实验方法:体外培养正常人成骨细胞,用不同浓度0,50,100,200,1000μg/L的重组结缔组织生长因子(0μg/L作为空白对照组)干预48h后,抽提细胞总RNA和总蛋白。③实验评估:采用半定量反转录-聚合酶链反应和蛋白免疫印迹分析方法观察不同浓度重组结缔组织生长因子对体外培养人成骨细胞核结合因子α1基因表达的影响。结果:半定量反转录-聚合酶链反应和蛋白免疫印迹结果显示,50,100,200,1000μg/L重组结缔组织生长因子干预后均可显著上调成骨细胞核结合因子α1的表达,并呈明显剂量依赖关系,与空白对照组比较,差异显著(P<0.01)。结论:重组结缔组织生长因子可剂量依赖性上调成骨细胞核结合因子α1基因的表达,核结合因子α1可能参与了结缔组织生长因子对成骨细胞增殖和分化的调节。     

Practice Involves More Than Treatment: How Can Evidence‐Based Assessment Catch Up to Evidence‐Based Treatment?

[Clin Psychol Sci Prac 18: 173–177, 2011] Given the excellent work being conducted in the area of evidence‐based treatments (EBTs), it is important to consider whether other forms of evidence‐based practice are receiving concomitant attention. While significant progress has been made in the last five years to generate reviews of evidence‐based assessment (EBA) practices, this work lags behind efforts to identify EBTs. This commentary describes available data on assessment practices in clinical care settings, discusses the importance of and current status of EBA, and considers how the next generation of EBA reviews might move beyond consideration of psychometric properties to the inclusion of “effectiveness” parameters.     

Inhibition of dipeptidyl peptidase-4 by vildagliptin during glucagon-like Peptide 1 infusion increases liver glucose uptake in the conscious dog

OBJECTIVE—This study investigated the acute effects of treatment with vildagliptin on dipeptidyl peptidase-4 (DPP-4) activity, glucagon-like peptide 1 (GLP-1) concentration, pancreatic hormone levels, and glucose metabolism. The primary aims were to determine the effects of DPP-4 inhibition on GLP-1 clearance and on hepatic glucose uptake.RESEARCH DESIGN AND METHODS—Fasted conscious dogs were studied in the presence (n = 6) or absence (control, n = 6) of oral vildagliptin (1 mg/kg). In both groups, GLP-1 was infused into the portal vein (1 pmol · kg⁻¹ · min⁻¹) for 240 min. During the same time, glucose was delivered into the portal vein at 4 mg · kg⁻¹ · min⁻¹ and into a peripheral vein at a variable rate to maintain the arterial plasma glucose level at 160 mg/dl.RESULTS—Vildagliptin fully inhibited DPP-4 over the 4-h experimental period. GLP-1 concentrations were increased in the vildagliptin-treated group (50 ± 3 vs. 85 ± 7 pmol/l in the portal vein in control and vildagliptin-treated dogs, respectively; P < 0.05) as a result of a 40% decrease in GLP-1 clearance (38 ± 5 and 22 ± 2 ml · kg⁻¹ · min⁻¹, respectively; P < 0.05). Although hepatic insulin and glucagon levels were not significantly altered, there was a tendency for plasma insulin to be greater (hepatic levels were 73 ± 10 vs. 88 ± 15 μU/ml, respectively). During vildagliptin treatment, net hepatic glucose uptake was threefold greater than in the control group. This effect was greater than that predicted by the change in insulin.CONCLUSIONS—Vildagliptin fully inhibited DPP-4 activity, reduced GLP-1 clearance by 40%, and increased hepatic glucose disposal by means beyond the effects of GLP-1 on insulin and glucagon secretion.Glucagon-like peptide 1 (GLP-1) is a gut-derived hormone shown to enhance glucose-dependent insulin secretion, suppress inappropriately high glucagon secretion, slow gastric emptying, and reduce food intake (1). In some type 2 diabetic patients, GLP-1 levels are reduced, and elevation of GLP-1 by continuous infusion of the peptide leads to reductions in fasting glucose, postprandial glucose excursions, and A1C (2). The therapeutic potential of GLP-1 is limited, however, because it is rapidly inactivated by dipeptidyl peptidase-4 (DPP-4) (3,4).Vildagliptin is an orally effective selective DPP-4 inhibitor. In diabetic patients, vildagliptin improved glycemic control, increased the plasma insulin–to–glucagon molar ratio, and reduced A1C levels (5,6). During a meal tolerance test, it augmented insulin secretion and decreased glucagon release, resulting in enhanced suppression of endogenous glucose production compared with placebo (7).Ingested glucose and endogenously secreted GLP-1 are released from the gut into the hepatic portal vein, which then perfuses the liver. Typically, studies have investigated the effects of DPP-4 inhibition after a meal, when GLP-1 secretion is increased. In the present study, GLP-1 and glucose were infused directly into the hepatic portal vein in the presence or absence of DPP-4 inhibition. The first aim was to examine the effect of vildagliptin on GLP-1 clearance under these carefully controlled conditions. In addition, although GLP-1 can increase glucose disposal by stimulation of insulin secretion, the hormone has been suggested to affect glucose metabolism by actions over and above its effects on the pancreas. Therefore, the second aim of this study was to investigate the effect of DPP-4 inhibition on glucose disposal, in particular by the liver.     

Elevated IL-6 levels in the synovial fluid of osteoarthritis patients stem from plasma cells

OBJECTIVE: To analyse the levels of interleukin-6 (IL-6) in the synovial fluids and sera of patients with osteoarthritis (OA) and to identify the IL-6-secreting cells. METHODS: Serum, synovial fluid, synovial tissue, and articular cartilage samples were collected from 49 OA patients with end-stage knee or hip OA who underwent joint replacement surgery. Serum and synovial fluid levels of IL-6 were measured by enzyme-linked immunosorbent assay (ELISA) and IL-6-secreting cells were identified by immunohistochemistry. RESULTS: Eight out of 49 patients (16%) exhibited elevated IL-6 levels in the synovial fluids, averaging at 2022+/-526 pg/mL, while the levels in the rest of the patients averaged at 132+/-19 pg/mL. The sera levels of all patients were comparable in the 10 pg/mL range. Immunohistochemical analyses revealed plasma cells in the synovial lining of the high producers as the source of IL-6. CONCLUSIONS: Synovial fluid IL-6 levels may help to classify OA patients and may point to a subgroup with a particular impact from their immune system.     

The effect of vagal cooling on canine hepatic glucose metabolism in the presence of hyperglycemia of peripheral origin

We examined the role of vagus nerves in the transmission of the portal glucose signal in conscious dogs. At time 0, somatostatin infusion was started along with intraportal insulin and glucagon at 4-fold basal and basal rates, respectively. Glucose was infused via a peripheral vein to create hyperglycemia ( approximately 2 fold basal). At t = 90, hollow coils around the vagus nerves were perfused with -10 degrees C or 37 degrees C solution in the vagally cooled (COOL) and sham-cooled (SHAM) groups, respectively (n = 6 per group). Effectiveness of vagal blockade was demonstrated by increase in heart rate during perfusion in the COOL vs SHAM groups (183 +/- 3 vs 102 +/- 5 beats per minute, respectively) and by prolapse of the third eyelid in the COOL group. Arterial plasma insulin (22 +/- 2 and 24 +/- 3 micro U/mL) and glucagon (37 +/- 5 and 40 +/- 4 pg/mL) concentrations did not change significantly between the first experimental period and the coil perfusion period in either the SHAM or COOL group, respectively. The hepatic glucose load throughout the entire experiment was 46 +/- 1 and 50 +/- 2 mg . kg(-1) . min(-1) in the SHAM and COOL groups, respectively. Net hepatic glucose uptake (NHGU) did not differ in the SHAM and COOL groups before (2.2 +/- 0.5 and 2.9 +/- 0.8 mg . kg(-1) . min(-1), respectively) or during the cooling period (3.0 +/- 0.5 and 3.4 +/- 0.6 mg . kg(-1) . min(-1), respectively). Likewise, net hepatic glucose fractional extraction and nonhepatic glucose uptake and clearance were not different between groups during coil perfusion. Interruption of vagal signaling in the presence of hyperinsulinemia and hyperglycemia resulting from peripheral glucose infusion did not affect NHGU, further supporting our previous suggestion that vagal input to the liver is not a primary determinant of NHGU.     

A phase I trial to determine the safety of imatinib in combination with vatalanib in patients with advanced malignancies

The role of tyrosine kinase inhibitors (TKIs) in the treatment of advanced malignancies is well established. Imatinib and vatalanib are oral TKIs with different mechanisms of action. This trial sought to establish the safety, tolerability, and maximum tolerated dose (MTD) of the two agents in combination. Secondary objectives included determination of potential pharmacologic interactions among vatalanib and imatinib and observation of antitumor activity. Patients with biopsy-proven advanced refractory solid tumors were enrolled in this single-center dose-escalation trial. Patients initially received imatinib and vatalanib once daily following a 14-day run-in period of daily oral vatalanib only, and were observed for a full 28-day treatment cycle prior to dose escalation. An amendment divided the vatalanib dose into two daily doses and gradually escalated the dose over a 2-3 week period. Patients continued combination therapy until disease progression or intolerable toxicity. Forty-five patients were enrolled between September 2004 and November 2007. As of September 2009, a total of 247 cycles of treatment had been administered (range: 1 -44+, median = 2 ). The MTD was determined to be vatalanib 1250 mg daily and imatinib 400 mg daily. Thirty-five patients (78%) were evaluable for response; 2 patients achieved PR, while 14 patients had SD ( 10 had stable disease ≥ 6 cycles). The combination of vatalanib and imatinib was well tolerated. Twice-daily vatalanib dosing improved tolerability and ease of full-dose administration. These results suggest that vatalanib-containing combinations may be active and tolerable, warranting further study.     

Metabolism and Renal Elimination of Gaboxadol in Humans: Role of UDP-Glucuronosyltransferases and Transporters

Purpose  Gaboxadol, a selective extrasynaptic agonist of the delta-containing γ-aminobutyric acid type A (GABA_A) receptor, is excreted in humans into the urine as parent drug and glucuronide conjugate. The goal of this study was to identify the UDP-Glucuronosyltransferase (UGT) enzymes and the transporters involved in the metabolism and active renal secretion of gaboxadol and its metabolite in humans.Methods. The structure of the glucuronide conjugate of gaboxadol in human urine was identified by LC/MS/MS. Human recombinant UGT isoforms were used to identify the enzymes responsible for the glucuronidation of gaboxadol. Transport of gaboxadol and its glucuronide was evaluated using cell lines and membrane vesicles expressing human organic anion transporters hOAT1 and hOAT3, organic cation transporter hOCT2, and the multidrug resistance proteins MRP2 and MRP4.Results. Our study indicated that the gaboxadol-O-glucuronide was the major metabolite excreted in human urine. UGT1A9, and to a lesser extent UGT1A6, UGT1A7 and UGT1A8, catalyzed the O-glucuronidation of gaboxadol in vitro. Gaboxadol was transported by hOAT1, but not by hOCT2, hOAT3, MRP2, and MRP4. Gaboxadol-O-glucuronide was transported by MRP4, but not MRP2.Conlusion. Gaboxadol could be taken up into the kidney by hOAT1 followed by glucuronidation and efflux of the conjugate into urine via MRP4.     

Mass spectrometric and physiological validation of a sensitive, automated, direct immunoassay for serum estradiol using the Architect

BACKGROUND: Measurement of estradiol (E(2)) plays a critical role in the diagnosis and clinical management of reproductive disorders. The challenge for all currently available direct methods for measuring E(2) is to provide accuracy and precision across a wide dynamic range. METHODS: We describe the development and multi-site performance evaluation of a direct E(2) assay on the Architect i2000. Assay performance and method comparisons were performed by testing specimens from men, healthy women with regular menstrual cycles, and post-menopausal women using the Architect assay and isotope dilution, gas chromatography-mass spectrometry (ID/GC-MS). Reference intervals were established by testing prospectively collected daily blood draws from 42 healthy women, 72 postmenopausal women and 101 males. RESULTS: No unexpected cross-reactivity or interference was observed for over 40 compounds tested. Recovery was 100+/-10% in the presence of estrone and estriol. Functional sensitivity (%CV<20%) was <15 pg/ml.(1) The imprecision of the assay was <7.1% (total CV), <2.5%, and <2.3% for control sera containing 45, 190, and 600 pg/ml estradiol, respectively. The assay had a correlation of y=1.033 x+0.3156, r(2)=0.99, n=131 compared to ID/GC-MS. Reference intervals for the current Architect Estradiol assay are reported. CONCLUSIONS: Format changes resulted in dramatic improvement in the performance and accuracy of this direct, fully automated assay. The assay is standardized by ID/GC-MS. The assay is clinically useful for serum concentrations from 15 to >4000 pg/ml.