Differential expression of CYP3A12 and CYP3A26 mRNAs in canine liver and intestine

The cytochrome P450 (CYP) 3A family is often considered the most important CYP subfamily with regard to drug metabolism in people. Certainly, CYP3A4 contributes to poor oral bioavailability of a number of drugs, and a tremendous amount of effort has been made in attempting to find an appropriate model system to predict the oral bioavailability of candidate drugs. The dog is a species widely used as a preclinical model for the evaluation of drug safety and pharmacokinetics. Compared with other species, little information is available on the tissue distribution of CYP enzymes. The purpose of this study was to determine the level of messenger RNA (mRNA) expression of the canine CYP3A subfamily (CYP3A12 and CYP3A26) in the liver and duodenum. Overall, expression of CYP3A mRNA was greater in the liver than in the duodenum. Hepatic expression of CYP3A26 was greater than CYP3A12 in all dogs, with CYP3A26 comprising 75.2% of the hepatic mRNA CYP3A pool. Conversely, duodenal expression of CYP3A12 was greater than CYP3A26 in all dogs, with CYP3A12 comprising 99.8% of the duodenal mRNA CYP3A pool. In summary, these results represent an important step toward the systematic comparison of human and canine CYP3A enzymes, particularly in relation to oral bioavailability of substrate drugs.     

Severe regional ischemia alters coronary flow reserve in the remote perfusion area

BACKGROUND: Clinical and experimental studies suggest that coronary flow reserve (CFR) may be abnormal in regions remote from myocardial infarction. We sought to determine the possible relation among stenosis severity, ischemic dysfunction, and impairment of CFR in remote regions. METHODS AND RESULTS: In 7 open-chest dogs, acute graded left circumflex (LCX) ischemia was created and maintained based on measurement of the transstenotic (aortic-distal LCX) pressure gradient (measured in millimeters of mercury). Regional thickening was assessed with sonomicrometers. Regional myocardial flow was assessed at rest with radiolabeled microspheres. Doppler flow probes were placed on proximal LCX and left anterior descending (LAD) arteries to measure resting flow and CFR in response to intracoronary injection of adenosine (36 microg). These parameters were assessed under baseline conditions and during transstenotic gradients of 10, 20, 30, and 40 mm Hg. Increasing LCX stenosis severity caused progressive impairment of LCX CFR: baseline (2.22+/-0.10), stenosis 10 (1.80+/-0.06), stenosis 20 (1.56+/-0.08), stenosis 30 (1.30+/-0.04), and stenosis 40 (1.17+/-0.06) (P<.01 vs. baseline). Remote LAD CFR was not altered by mild to moderate LCX stenosis (baseline [2.33+/-0.19]; stenosis 10 [2.30+/-0.25]; stenosis 20 [2.15+/-0.26]). However, critical LCX stenosis producing mild to moderate reduction in thickening in the ischemic region was associated with a significant impairment of LAD CFR: stenosis 30 (1.90+/-0.26) and stenosis 40 (1.80+/-0.22) (P<.01 vs. baseline). These changes in remote CFR persisted after correction for changes in the rate-pressure product. CONCLUSION: In an acute canine model of progressive LCX coronary stenosis, CFR was impaired in both ischemic and remote nonischemic regions in association with mild to moderate ischemic-induced regional myocardial dysfunction. Thus pharmacologic vasodilation provoked only mild heterogeneity in CFR in the presence of a critical LCX stenosis as a result of concurrent reduction of LAD CFR. This phenomenon warrants further clinical and experimental investigation because it may affect detection of flow heterogeneity during acute ischemia (which induced myocardial dysfunction).     

Comparison of methods for localization of impacted maxillary canines by panoramic radiographs

Objectives:

The objective of this study was to compare three methods for localization of impacted maxillary canines using only conventional panoramic radiographs.

Methods:

The panoramic radiographs of 94 patients (102 impacted maxillary canines) were reviewed and evaluated using the methods magnification, angulation and superimposition. The actual positions of them were decided with cone beam CT images. The predicted positions of impacted canines from the magnification and angulation methods were compared using the McNemar χ² test. Sensitivity, specificity, accuracy, positive-likelihood ratio and negative-likelihood ratio were calculated. The canine-incisor index values and α angles of palatally and bucally non-rotated impacted canines were compared using the Mann–Whitney U test.

Results:

The statistical analysis revealed that there was a significant difference between the magnification and angulation methods (p < 0.01). Using the magnification method, 68.00% of buccal canines and 69.57% of palatal canines could be localized correctly. The results of the angulation method were 28.57% and 84.91%, respectively. The sensitivity of the angulation method for buccal canines was very low. In the superimposition method, 82.98% of the superimposing samples were palatal.

Conclusions:

The magnification and angulation methods were not reliable methods for locating the impacted canine with a single panoramic radiograph. Magnification was more successful than the angulation method. Further research is needed on the magnification method. The image superimposition method could be used as an adjunct to others.     

犬心肌缺血模型的构建

目的 制备心肌缺血模型。方法 缝扎犬左冠状动脉第一对角枝及其供血区域的所有侧支血管，形成7—8cm2大小的缺血区域。结果 完成54例心肌缺血模型，手术、心电图、同位素证实心肌缺血，缺血区边界清晰。结论 该方法具有效果确实可靠、手术方便完全、不需特殊设备等特点，是构建心肌缺血模型的理想方法。     

Effect of nefiracetam, a neurotransmission enhancer, on primary uroepithelial cells of the canine urinary bladder.

Repeated oral treatment of dogs with a high dose of nefiracetam is reported to induce hemorrhagic lesions in the urinary bladder. To delineate its pathogenesis, we established the primary culture of uroepithelial cells of the canine urinary bladder, and then explored the effect of nefiracetam on the cultured cells. Uroepithelial cells scraped from the connective tissues of the urinary bladder of naive dogs were suspended in the minimum essential medium containing dispase, and then resuspended in the keratinocyte medium to be 6.0-7.0 x 10(5) cells/ml. Afterward, they were added to an apical chamber with a 12-mm transwell filter, cultured for three days, and recultured in the keratinocyte medium containing 1 mM CaCl(2) for 20 days. Microscopically, these cultured cells consisted of three cell layers with high transepithelial electric resistance (TER; > 10,000 ohm-cm(2)). Immunofluorescence observations revealed ZO-1 and E-cadherin bands, and electron microscopic examinations displayed the superficial cells with the assembly of tight junctions. When the effect of nefiracetam and its five main metabolites (M-3, M-10, M-11, M-18, and M-20) on TER and the ZO-1 band was assessed using cultured cells, only M-18 significantly reduced TER in the coculture for 48 h or more. Both M-10 and M-18 exhibited a deformation of uroepithelial cells and a slight reduction of the ZO-1 band from 120 h later. In conclusion, this culture system possesses both functional and morphological features of the uroepithelium reflected in vivo, and M-18 may play a pivotal role in the impairment of uroepithelial cells, leading to the onset of the urinary bladder lesion in dogs due to nefiracetam.     

Expression and mapping of lubricin in canine flexor tendon.

Matrix composition and the biomechanical environment are intimately interdependent in most connective tissues. Lubricin has many distinct biological functions, including lubrication, antiadhesion, and cytoprotection in cartilage, tendons, and other tissues. This study investigated the distribution of lubricin in the canine flexor digitorum profundus (FDP) tendon by immunohistochemistry. Lubricin was found both on the tendon surface and at the interface of collagen fiber bundles within the tendon, where the cells are subjected to shear force in addition to tension and compression. The expression of lubricin in regions of the canine flexor tendon that differ in mechanical or nutritional environment was also investigated using RT-PCR. Six N-terminal splicing variants were identified from six distinct anatomical regions of flexor tendon. The variants with larger sizes were noted in regions subjected to significant shear and compressive forces. Lubricin is ubiquitous in the FDP tendon, with variations in distribution and splicing that appear to correspond to discrete anatomic locations that differ by mechanical or nutritional environment.     

Enhanced histologic repair in a central wound in the anterior cruciate ligament with a collagen-platelet-rich plasma scaffold.

The anterior cruciate ligament (ACL) of the knee is an intra-articular ligament that fails to heal after primary repair. The medial collateral ligament (MCL) of the knee is an extra-articular ligament that heals uneventfully in the majority of cases. Why these two ligaments have such different responses to injury remains unclear. In this article, we address two hypotheses: first, that the histologic response to injury is different in intra-articular and extra-articular ligaments, and second, that the response of the intra-articular ligaments can be altered by placing a collagen-platelet-rich plasma (collagen-PRP) hydrogel in the wound site. Wounds were created in extra-articular ligaments (MCL and/or patellar ligament) and an intra-articular ligament (ACL) in canine knees, and the histologic response to injury evaluated at 3 days (n = 3), 7 days (n = 4), 3 weeks (n = 5), and 6 weeks (n = 5). In the 3-week (n = 5) and 6-week (n = 5) animals, bilateral central wounds were made in the ACLs and the wounds in one knee of each animal treated with a collagen-PRP hydrogel while the contralateral side was untreated. Extra-articular ligament wounds had greater filling of the wound site and increased presence in the wound site of fibrinogen, fibronectin, PDGF-A, TGF-beta1, FGF-2, and von Willebrand's factor when compared to intra-articular ligament wounds. Treatment of the intra-articular wound with a collagen-PRP hydrogel resulted in increased filling of the wound site with repair tissue that had similar profiles of growth factor and protein expression to the extra-articular ligament wounds. The use of a collagen-PRP scaffold can ameliorate histologic differences noted between healing extra-articular ligamentous wounds and nonhealing intra-articular ligamentous wounds. This study supports the hypothesis that premature scaffold failure may play a key role in the normally expected failure of the ACL to heal after injury.     

急性脑梗死动脉溶栓治疗前后的质子磁共振波谱实验研究

目的 评价质子磁共振波谱(proton magnetic resonance spectroscopy,1HMRS)在家犬实验性脑缺血性损伤和动脉溶栓治疗中的应用价值.方法 18条杂种成年家犬,自体血栓法建立局灶性脑缺血模型后随机分成未溶栓组(A组)和B1(1.5 h溶栓)、B3(3.5 h溶栓)、B6(6.5 h溶栓)、B9(9.5 h溶栓)、B12(12.5 h溶栓)6组,每组3条.每条动物均在栓塞后1、3、6、9、12、24 h行轴位DWI、MRS扫描.各组犬随机抽取1条于栓塞后24 h处死取脑做病理学检查.结果 未溶栓组NAA,Cho及Cr峰在脑缺血后均表现为下降,其中NAA峰1～6 h内下降最迅速,Cho及Cr下降幅度不如NAA明显;Lac峰在脑缺血24 h内随梗死时间的延长而持续升高.溶栓组B1、B3、B6、B9组Lac峰升高程度比A组明显减小(P＜0.05);NAA峰下降幅度减小(P＜0.01);Lac/Cho值与24 h神经功能评分存在着高度的相关性.光镜下,未溶栓组出现大量神经元细胞坏死性改变,溶栓组神经元细胞形态改变程度轻,残存细胞数目明显增多.结论 质子磁共振波谱能敏感地反映犬局灶性脑缺血后脑内代谢的动态演变过程,并可监测溶栓治疗效果.     

在体评价犬和猪冠状动脉内皮细胞的功能

[目的]在体观察缺血再灌注对犬冠状动脉内皮细胞功能的影响以及猪冠状动脉对血管活性药物的反应性.[方法]动脉麻醉后开胸、心包吊床,之后分离冠状动脉左前降支.犬作第一对角支插管,猪作冠状动脉左回旋支插管.犬冠状动脉左前降支缺血30 min后再灌注60 min,猪不作缺血再灌注.分别向冠状动脉内注射乙酰胆碱和硝酸甘油,以注药后平均冠状动脉血流量变化的百分率来评价冠状动脉内皮细胞依赖性和非依赖性的舒张功能.[结果]注乙酰胆碱后,基线状态下犬平均冠状动脉血流量从(17±4)mL/min升至(41±12)mL/min(P＜0.05);但缺血再灌注后舒血管反应幅度降低,其中再灌注15 min时间点平均冠状动脉血流量增幅水平明显低于基线增幅水平(142%±54%vs 87%±57%,P＜0.05).注乙酰胆碱后,基线状态下猪平均冠状动脉血流量在(11±3)s内从(21±11)mL/min迅速降至(12±8)mL/min,在(42±10)s内又回升至(23±13)mL/min.硝酸甘油使基线状态下犬平均冠状动脉血流量从(18±5)mL/min升至(40±22)mL/min(P＜0.05),猪的冠状动脉也呈类似反应.[结论]乙酰胆碱引起犬冠状动脉呈舒血管反应,缺血再灌注使犬冠状动脉内皮细胞功能明显受损.猪冠状动脉对乙酰胆碱呈先收缩后舒张的"双向反应".硝酸甘油则引起犬或猪的冠状动脉均呈舒张反应.